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BJID 2007; 11 (October) Volume 11 • 1 ISSN 1413-8670 Supplement 1 • October 2007 THE BRAZILIAN JOURNAL OF INFECTIOUS DISEASES An Official Publication of the Brazilian Society of Infectious Diseases EDITOR Anastácio Q. Sousa Consensus of the Brazilian Society of Infectious Diseases on the Management and Treatment of Hepatitis C PUBLISHED BY CONTEXTO October 2007 Printed in Brazil www.bjid.com.br 2 BJID 2007; 11 (October) THE BRAZILIAN SOCIETY OF INFECTIOUS DISEASES The Brazilian Society of Infectious Diseases is conducted for scientific purposes, for the advancement and promulgation of knowledge relevant to infectious diseases. OFFICERS 2007-2008 President João Silva de Mendonça Vice President Denise Vantil Marangoni First Secretary Juvêncio José D. Furtado Second Secretary Érico Antônio Gomes de Arruda First Treasurer Roberto Marcio da Costa Florim Second Treasurer Flávio de Queiroz Telles Filho Federal Federal Alagoas - Maria Raquel dos A.S. Guimarães Amazonas - Eucides Batista da Silva Bahia - Adriano Oliveira Ceará - Anastácio de Queiroz Sousa Distrito Federal - Dea Márcia da Silva M. Pereira Espírito Santo - Carlos Urbano Gonçalves Ferreira Jr. Goiás - Marcelo Cecílio Daher Maranhão - Graça Viana Mato Grosso do Sul - José Ivan de A. Aguiar Minas Gerais - Carlos Ernesto Ferreira Starling Pará - Helena Andrade Zeferino Brígido Paraíba - Luciana Holmes Simões Paraná - Alceu Fontana Pacheco Jr. Pernambuco - Martha Maria Romeiro F. Fonseca Piauí - Kelsen Dantas Eulálio Rio de Janeiro - J. Samuel Kierszembaum Rio Grande do Norte - Hênio Godeiro Lacerda Rio Grande do Sul - Luciano Goldani Rondônia - André Luis de Freitas Alves Santa Catarina - Silvia Cristina C. Flores São Paulo - Maria Luiza Moretti Sergipe - Márcia Maria Macedo Lima Tocantins - Hertz Ward de Oliveira www.bjid.com.br BJID 2007; 11 (October) 3 THE BRAZILIAN JOURNAL OF INFECTIOUS DISEASES An Official Publication of the Brazilian Society of Infectious Diseases EDITOR John R. David (US) Jorge Arias (BR) Jorge Luiz Nobre Rodrigues(BR) Jorge Luiz Sampaio (BR) José Wellington Oliveira Lima (BR) Kleber Luz (BR) Marcelo Ferreira (BR) Marcos Antônio de Ávila Vitória (BR) Maria Aparecida Shikanai Yasuda (BR) Maria Rita Elmor (BR) Mark Wainberg (CA) Mauro Schechter (BR) Mitermayer Galvão dos Reis (BR) Naftale Katz (BR) Raimundo Paraná (BR) Reinaldo Salomão (BR) Ricardo Diaz (BR) Richard Guerrant (US) Richard Locksley (US) Richard B. Roberts (US) Robério Dias Leite (BR) Robert Schooley (US) Rod Hay (GB) Rodolfo Teixeira (BR) Rogério de J. Pedro (BR) Selma Maria Bezerra Jerônimo (BR) Sérgio Cimerman (BR) Sérgio Coutinho (BR) Sylvia Lemos Hinrichsen (BR) Timothy Inglis (AUS) Warren D. Johnson, Jr. (US) Zilton Andrade (BR) Anastácio Q. Sousa ASSOCIATE EDITORS Adauto Castelo (BR) André Villela Lomar (BR) Antônio C. Pignatari (BR) Carlos Brites Alves (BR) Hélio Sader (BR) João Silva de Mendonça (BR) Márcio Nucci (BR) Roberto Badaró (BR) Roberto Focaccia (BR) EDITORIAL BOARD Achilea L. Bittencourt (BR) Antônio Alci Barone (BR) Antônio Andrade (BR) Antônio Campos Neto (BR) Antônio Carlos Nicodemo (BR) Arnaldo Colombo (BR) Caio Mendes (BR) Celso Ramos Filho (BR) Cláudio Sérgio Pannuti (BR) Dirceu Grecco (BR) Edgard Marcelino de Carvalho (BR) Eduardo Netto (BR) Érico Antônio Gomes de Arruda (BR) Esper Georges Kallas (BR) Eurico de Arruda Neto (BR) Flávia Rossi (BR) Guido Levi (BR) Henry Masur (US) Jeffrey Shaw (BR) PRODUCTION STAFF Luciana Bastianelli, Managing Editor Andréia Lima, Submissions Manager Taís Cupertino, Secretary www.bjid.com.br 4 BJID 2007; 11 (October) The Brazilian Journal of Infectious Diseases is an official publication of the Brazilian Society of Infectious Diseases and is published bimonthly by Contexto - Rua Alfredo Magalhães, 04/ Barra, 40140-140, Salvador-Bahia-Brazil. The editorial offices are at Contexto. Editorial Office Correspondence concerning subscriptions, advertisements, claims for missing issues, changes of address and communications to the editors should be addressed to Dr. Anastácio Q. Sousa, The Brazilian Journal of Infectious Diseases - BJID, Rua Alfredo Magalhães, 04/Barra, 40140-140, Salvador-BahiaBrazil; phone: (55 71) 3264-2971; fax (55 71) 32643326; or should be sent by e-mail: [email protected]. Contributors please consult the Instructions for Authors in this issue or visit www.bjid.com.br Sponsors This supplement was supported by: • Produtos Roche Químicos e Farmacêuticos S/A Permissions Copyright 2007 by The Brazilian Journal of Infectious Diseases and Contexto Publishing. All rights reserved. Except as authorized in the accompanying statement, no part of the BJID may be reproduced in any form or by any electronic or mechanical means, including information storage and retrieval systems, without the publisher's written permission. Authorization to photocopy items for internal or personal use, or the internal or personal use by specific clients is granted by The Brazilian Journal of Infectious Diseases and Contexto Publishing for libraries and other users. This authorization does not extend to other kinds of copying such as copying for general distribution, for advertising or promotional purposes, for creating new collective works, or for resale. Postmaster Send address changes to BJID, Rua Alfredo Magalhães, 04/Barra, 40140-140, Salvador-Bahia-Brazil. Subscriptions Subscription rates (1 year) for The Brazilian Journal of Infectious Diseases (BJID): Domestic Foreign (R$) (US$) Individuals 100 100 Institutions 150 150 Special* 60 60 *Students, interns, residents and fellows. • Schering-Plough do Brasil Information by Contexto: Home-page: www.bjid.com.br. E-mail: [email protected] Phone/Fax: (55 71) 3264-2971 / 3264-3326. The opinions expressed in this publication are those of the authors and are not attributable to the sponsors or to the publisher, editor, associate editors, or editorial board of The Brazilian Journal of Infectious Diseases. COVER: Liver cells infected with the hepatitis C virus. © 2007 by The Brazilian Journal of Infectious Diseases and Contexto Publishing. All rights reserved. Now Indexed in INDEX MEDICUS/PUBMED/MEDLINE, EMBASE/EXCERPTAMEDICA, LATINDEX, LILACS, SUBIS DATABASE, SIIC, SCIELO, CAS, SOCIEDAD IBEROAMERICANA DE INFORMACIÓN CIENTIFICA, ELSEVIER SCIENCE - BIBLIOGRAPHIC DATABASES DIVISION, CABI PUBLISHING, ULRICH’S PERIODICALS DIRECTORY www.bjid.com.br BJID 2007; 11 (October) THE BR AZILIAN BRAZILIAN 5 JOURNAL OF INFECTIOUS Volume 11 • Supplement 1 DISEASES October 2007 Consensus of the Brazilian Society of Infectious Diseases on the Management and Treatment of Hepatitis C ........... 1 Therapeutic Approach to Acute Hepatitis C ................... 49 Brazilian Society of Infectious Diseases HCV Consensus Group Epidemiological Aspects of Hepatitis C in Brazil .............. 6 Treatment of Chronic Hepatitis C in Treatment-Naïve Patients ........................................................................... 53 Decio Diament Marcelo Simão Ferreira Sexual Transmission of HCV ............................................. 8 Kleber Dias do Prado Retreatment of Hepatitis C Patients Who Previously Experienced Treatment Failure ......................................... 58 Hepatitis C Virus Perinatal Transmission ......................... 10 Fernando Lopes Gonçales Jr. Umbeliana Barbosa de Oliveira Maintenance Treatment for the Modulation of Liver Fibrosis ........................................................................... 60 H e p a t i t i s C : Vi r o l o g i c a l A s p e c t s a n d P r a c t i c a l Implications .................................................................... 12 Antonio Alci Barone Rodrigo Nogueira Angerami and Fernando Lopes Gonçales Júnior Evaldo Stanislau Affonso de Araújo and Antonio Alci Barone Pathogenesis of Hepatitis C – HCV Consensus 2007 ...... 14 Treatment of Patients Infected with Hepatitis C Virus and Presenting Extrahepatic Manifestations .......................... 64 Ana Tereza R. Viso Fátima Mitiko Tengan1 and Antonio Alci Barone Antifibrotic Therapy in Chronic Hepatitis C .................... 20 Rinaldo Focaccia Siciliano and Antonio Alci Barone H e p a t i t i s C Tr e a t m e n t B e f o r e a n d A f t e r L i v e r Transplant ..................................................................... 69 Laboratory Testing for Hepatitis C .................................. 21 Edson Abdala, Daniela Rosa Magalhães Gotardo, Patrícia Rodrigues Bonazzi and Telésforo Bacchella Neiva Sellan Lopes Gonçales and Fernando Lopes Gonçales Junior Adverse Event Management ........................................... 74 Hepatitis C: Genotyping ................................................... 25 Aline Gonzalez Vigani Norma de Paula Cavalheiro Treatment Options in the Management of Thrombocytopenia in Patients Infected with HCV .......................................... 79 Noninvasive Means of Diagnosing Liver Fibrosis in Hepatitis C ...................................................................... 28 Eduardo Sellan Lopes Gonçales, Adriana Flávia Feltrim Angerami and Fernando Lopes Gonçales Junior Chronic Hepatitis C: Pathological Anatomy ..................... 32 André Cosme de Oliveira Therapeutic Perspectives for Hepatitis C ......................... 81 Evaldo Stanislau Affonso de Araújo, Antonio Alci Barone and JeanMichel Pawlotsky Evandro Sobroza de Mello and Venâncio Avancini Ferreira Alves Co-Infection with Hepatitis B Virus and Hepatitis C Virus ................................................................................ 37 Heloísa Pedrosa Mitre and João Silva de Mendonça Co-Infection with Hepatitis C Virus and Human T Lymphocyte Virus ................................................................................. 40 Instructions for Authors Carlos Brites Alves Basic Guidelines for the Treatment of HIV/HVC CoInfection ......................................................................... 42 Statement of Editorial Policy Edgard De Bortholi Basic Aspects of the Treatment for Hepatitis C: Mechanisms of Action of Interferon Alpha and Ribavirin and the Bases of Individualization .............................................................. 47 Carlos Eduardo de Melo, Evaldo Stanislau Affonso de Araújo and Antonio Alci Barone Checklist for Submitted Manuscripts www.bjid.com.br 6 BJID 2007; 11 (October) BJID on line The Brazilian Journal of Infectious Diseases (BJID) is now present in a new Web Site: www.bjid.com.br. The BJID is an official publication of The Brazilian Society of Infectious Diseases and it is a vehicle for original publications in this field. The BJID is published bimonthly by Contexto Publishing (Salvador/BA, Brazil) since 1997. The aim of this site is to familiarize visitors with BJID' contents by providing online subscriptions, review process, submission process, abstracts, past publications, and extend the BJID´s instructions for authors to infectious-disease specialists worldwide. Full instructions for authors are provided in English and Portuguese. This new website is divided into the following sections: Home (A new design was established for BJID, including spaces for advertising) Editors (This section includes all the editorial board of BJID) Subscriptions (In this section, the visitor could subscribe the BJID online) Instructions to Authors (This section is in Portuguese and in English, and can help the authors who want to submit papers to BJID to follow. There are the rules for publishing in the Journal). Index Listing (The visitors can check the status of the BJID in the medicus indexa round the world. In a nearly future, we will include a service to check the impact of each article published in BJID) Abstracts (The visitors have free access to the abstracts of the current year. For example: every each issue, the abstracts will be available to be consulted. If the visitors would like the full text, they need to ask the permission to our office). Publications (In this section, it will be available the full text of last volumes 1997 to 2003 in pdf format) Sponsors (This space is reserved to the advertising board of the BJID and the institutions that support the Journal. The visitors can be addressed directly to the sponsor they want by a link) About us (This section inform the visitors about the production staff of the Journal, the address, phone number, e-mail, and the contacts of the BJID) Links (Interesting links about infectious diseases and institutions) This website reserved a new space to the authors who have already submitted papers to the Journal and/or those have papers in the submission process in BJID with the following sections: On-line Submission Process (The authors now could send the papers by e-mail, following the specific rules described in this section. Also, the authors now can consult the review process of the submitted articles via on-line. Immedialely after the submission of the article, the main author will receive a login and a password, with which the author will be able to consult the status of the article. If other authors want to check the status of their article via on-line, they can also register a proper login and password. On-line Review Process (The reviewers now can receive the article and make all the edition process by internet. The reviewer will receive a message with a login an a password. With them, he could access the entire article. After he comments in a specific form, it will be sent to the author automatically. The authors now can consult the review process of the submitted articles via online. Immediately after the submission of the article, the main author will receive a login and a password, with which the author will be able to consult the status of the article. If other authors want to check the status of their article via on-line, they can also register a proper login and password. www.bjid.com.br BJID 2007; 11 Supplement 1 (October) 1 Consensus of the Brazilian Society of Infectious Diseases on the Management and Treatment of Hepatitis C Evaldo Stanislau Affonso de Araújo, João Silva Mendonça, Antonio Alci Barone, Fernando Lopes Gonçales Junior, Marcelo Simão Ferreira*, Roberto Focaccia, Jean-Michel Pawlotsky§* and Brazilian Society of Infectious Diseases HCV Consensus Group Each year, and every day, the results of clinical trials and basic research provide us with a great deal of new information regarding viral hepatitis. We on the Viral Hepatitis Committee of the Sociedade Brasileira de Infectologia (SBI, Brazilian Society of Infectious Diseases) have been working to standardize the major issues surrounding dayto-day practice in treating patients infected with the hepatitis B or C virus (HBV or HCV). We have decided to address, in alternate years, HBV, together with hepatitis delta (HDV), and HCV, in our annual ‘Consensus’ on clinical management. Last year, we published the first HBV Consensus (BJID, 2007 (11):2-6). This year, we submit our HCV Consensus, which primarily serves to update the 2002 and 2004 SPI Consensuses. We distributed the principal topics among the Committee members, revised their work and compiled it into a Proceedings Supplement (to be published together with the BJID), which elucidates the highlights of the Consensus. A deeper review was written and referenced (it is our advice to the reader to read the Proceedings as well). A meeting was then held in Mogi das Cruzes in order to discuss, in a very practical and directed way, the issues most relevant to the Consensus, from public policies to the most complex therapeutic points. The results are summarized in a question/answer, topic/ statement format in this issue of the BJID. The main message of our statement was Figure 1. HCV Consensus Group. that we need to have the courage to act in favor of life. Many of us have adopted certain practices based on very new knowledge despite a lack of formal or official policies to support such practices. Some of us have been awaiting new compounds while patients are dying of chronic liver diseases. Unfortunately, the news from the battlefield is not so good. New compounds have been very disappointing (low potency, viral resistance, ineffective without interferon and various side effects, some serious). It is also difficult to incorporate new policies into everyday practice. However, strategies such as optimizing the use of pegylated interferon/ribavirin and encouraging treatment compliance, as well as finding new ways to monitor and slow liver disease progression, are effective and should be put into practice. Most importantly, the low-dose maintenance of pegylated interferon seems to be very promising, and the use of interferon alpha has saved lives. That is why we choose to expound upon what we believe to be the current standard of care and the gold standard for dealing with this hard to treat virus, as well as with the chronic complications of HCV infection. Our position will be re-evaluated over the next two years. Until then, we are confident that our guide will be of great value to the readers. Finally, we would like to thank Roche and Schering Plough for the educational grants provided to the SBI. However, we must stress that neither company attempted to influence any of the decisions made by our consensus group. The Brazilian Society of Infectious Diseases HCV Consensus Group Social and Collective Impact of the Decision-Making Process Serologic triage for hepatitis C virus (HCV): When and how? Diagnosis ‘campaigns’? Most vulnerable groups? It is recommended that detection ‘campaigns’ be carried out for more vulnerable groups 1, individuals in certain #Brazilian Society Infectious Diseases HCV Consensus Group: Ana Tereza Rodrigues Viso, Aline Vigani, André Cosme Oliveira, Carlos Eduardo de Melo, Carlos Brites Alves, Décio Diament, Edson Abdala, Edgard De Bortholi, Evandro Sobroza de Mello, Eduardo Sellan Gonçalves, Fátima Mitiko Tengan, Heloísa Pedrosa Mitre*, Kleber Dias Prado, Neiva Sellan Gonçales, Norma de Paula Cavalheiro, Rinaldo Focaccia Siciliano, Rodrigo Nogueira Angerami*, Umbeliana Barbosa de Oliveira, Venâncio Avancini Ferreira Alves*. § Special guest. *Members who did not participate of the Consensus Meeting occurred in the Blue Tree Park Hotel, Mogi das Cruzes/SP, on August 25th-26th, 2007. occupations (health professionals, technicians whose work potentially puts them in contact with blood), institutionalized individuals (prisoners, under-age felons, etc.), and the family members of patients with hepatitis C, as well as those infected with HIV. We emphasize that such individuals should be clearly informed that the triage diagnosis should be confirmed by a method with more sensitivity and specificity, as well as that confirmation of the diagnosis will not necessary imply treatment of the infection. Age is a factor to be considered, 1 History of blood product transfusion, history of major surgery, unsafe parenteral exposure (reusable syringes, sharing of utensils during the use of licit or illicit injected substances, undergoing diagnostic or esthetic therapeutic procedures involving reusable or inadequately sterilized material, e.g., tattoo, acupuncture, piercing, manicure, dental treatment, etc.), and sharing of utensils during the use of inhaled drugs. www.bjid.com.br 2 Consensus of Hepatitis C Carried Out by SBI since older people are more likely to have been exposed to the disease over the course of their lifetime. The infected health professional: are their activities restricted in any way? A priori, the activity of health professionals with hepatitis C should not be restricted. However, strict application and educational measures are recommended regarding the biosafety guidelines, and it should be noted that more data are needed before a definitive recommendation can be made. Is hepatitis C a sexually transmitted disease (STD)? Although hepatitis C is not conceptually an STD, sexual transmission is possible, albeit uncommon, and the risk of such transmission increases when the individual presents genital lesions or HIV positivity, as well as when the individual engages in risky sexual behavior. In such situations, the use of condoms is recommended. Heterosexual monogamous couples who present discrepant serologic results can use condoms of their own accord after being properly instructed. We would like to issue a warning regarding the risk of transmission in the household through the communal use of utensils contaminated with blood. Hepatitis C during pregnancy and childbirth: To get pregnant or not? Cesarean or vaginal delivery? Breastfeeding? Pregnancy is not contraindicated in women of childbearing age infected with HCV, although contraindications related to the period of treatment should be respected. Regarding the type of delivery, the decision should be made by an obstetrician. It should be noted, however, that a high viral load of HCV can be a relevant factor in this decision. Nevertheless, at the present, we cannot recommend a definitive course of action. Breastfeeding is allowed, although the nipple should be carefully prepared, and breastfeeding should be discontinued if fissures appear or bleeding occurs. Pregnant women coinfected with HCV and HIV present a clearly greater risk of perinatal HCV transmission and therefore constitute an exception to these permissions. Prevention Unapparent HCV transmission in society and in the health care environment: What should we recommend to Health Oversight Agencies? We recommend extremely rigorous inspection of health and esthetic institutes, as well as continuing education of the professionals who work in this area. We should also emphasize the need for ongoing education of health professionals at all levels of patient care – from basic care to highly complex treatments. Vaccines: Which and when? Susceptible individuals with hepatitis C should be vaccinated against hepatitis A and B. There is a real need to make vaccines against hepatitis A available in the public health care system. BJID 2007; 11 Supplement 1 (October) The Laboratory in the Era of Individualized Treatment Quantification of viral loads for all genotypes? Yes, it is recommended that the viral load of all patients be quantified. Which quantification method should be used? It is recommended that the method used be reproducible, be sensitive, and present ample linearity. ‘Moments of decision’: At baseline (One measurement? When? Several measurements?), as well as at weeks 4, 12, and 24; end-of-treatment response (ETR), sustained virologic response (SVR)...What else? Baseline viral load should be determined prior to but as close as possible to the initiation of the treatment. It can be determined only once provided that the test is performed under ideal technical conditions. At week 4, the same test will be qualitative in order to define the presence of rapid virologic response (RVR), defined as the detection of no viral RNA. At week 12, it should be quantitative, adopting the criteria of no early virologic response (EVR) (a drop in viral load ≤ 2log10), partial EVR (drop ≥ 2log10), and complete EVR (no viral RNA detected). At week 24, the test will be qualitative. If viral RNA is detected, the treatment instituted exclusively for virologic purposes will have to be interrupted. Determination of the ETR – emphasizing the differences in duration in different genotypes and patients – as well as of the SVR, must be qualitative. The SVR should be determined at 24 weeks after the end of the treatment. It should be noted that monitoring will be unnecessary during the treatment in the cases in which RVR occurred and adequate treatment compliance is maintained. ‘Week 12’ SVR? Determination of the SVR at 12 weeks after the end of the treatment is not currently considered a useful or valid measure. Metabolic alterations: Homeostasis model assessment (HOMA) and glucose tolerance test: when to order, how to interpret Various studies indicate the role of hepatitis C as a factor implicated in the development of type 2 diabetes in patients at high risk (male gender, over 40 years of age, and overweight). In experimental models, HCV was found to induce insulin resistance, including increased production of tumor necrosis factor as one of the contributing factors. Insulin resistance is also associated with the development of steatosis and progression of liver fibrosis, principally in patients infected with HCV genotype 1. Therefore, there seems to be an association between insulin resistance and the characteristics associated with patients presenting a worse response to the hepatitis C treatment: cirrhosis, obesity, concomitant infection with HIV (taking antiretroviral drugs), etc. In summary: HCV promotes insulin resistance, which leads to steatosis, fibrosis, and resistance to treatment with interferon alpha (IFN-α). The HOMA mathematical model [(serum levels of fasting insulin vs. serum levels of fasting glucose)/22.5] has proven useful in the evaluation of sensitivity to insulin; however, it www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Consensus of Hepatitis C Carried Out by SBI has not been completely standardized for all clinical situations, which is why its routine use is not recommended, except in the situations that will be reviewed now (metabolic alterations) and in the Proceedings. Noninvasive monitoring of fibrosis Value of simplicity: Thrombocytopenia as a marker of liver cirrhosis Thrombocytopenia in an individual with hepatitis C indicates moderate to severe liver fibrosis. The sensitivity of this diagnostic marker of advanced fibrosis increases when it is used in combination with other biochemical tests, such as determination of the aspartate aminotransferase to platelet ratio index, calculated using the formula: aspartate aminotransferase/alanine aminotransferase ratio + platelets. When is it indicated? Noninvasive monitoring of liver fibrosis should be performed in cases of (i) contraindication or difficult access to liver biopsy, (ii) the need for such monitoring in order to hasten the performance of a new biopsy in patients at greater risk of progression (immunocompromised patients) or in individuals presenting stage F1 fibrosis, and (iii) in patients with liver cirrhosis under treatment for modulation of fibrosis (IFN maintenance). How to measure: Serum panels vs. Fibroscan Sensitivity and positive predictive value are increased by combining the methods. Liver biopsy is dead. . . Long live liver biopsy! Biopsy for all genotypes? And for all patients? Yes, except for clinical contraindications, the biopsy should be performed for all patients presenting detectable levels of HCV RNA. The central issue is the absolute need for the biopsy sample to be representative of the hepatic parenchyma, since inappropriate biopsies frequently result in understaging of the disease. It is highly recommended that a needle biopsy be performed (wedge biopsies produce subcapsular samples that cannot be used in the staging of fibrosis nor in the staging of inflammation); needles that produce very thin biopsies highly limit architectural staging and should not be used. Trucut 14gauge needles or needles that produce samples of equivalent diameter, measuring at least 1.5 cm long and/or 10 represented portspaces, are recommended. If the biopsy is performed during a surgical procedure, it should be done at the outset of the procedure in order to avoid the artifacts that surgery can produce in the liver tissue. The biopsy report should include a staging system and scoring (METAVIR or SBP, preferably). It should be noted, however, that the pattern of inflammatory alterations (portal, interface, and lobular components) should be described in detail, since it has become increasingly important in predicting the evolution of the disease as well as in the differential diagnosis with other diseases. In addition, the principal role of biopsy is to rule out other liver diseases – steatohepatitis (alcoholic or nonalcoholic), for example, frequently co-exists with hepatitis C and is known to have a significant impact on the evolution of the disease. 3 Value of the imaging and serum level methods in the diagnosis of liver fibrosis: who needs endoscopy? Prior to biopsy, patients with indirect evidence of portal hypertension (ultrasonographic signs, thrombocytopenia) should be submitted to endoscopy of the upper digestive tract for detection of esophageal varicose veins that would render a liver biopsy unnecessary in the diagnosis of liver cirrhosis. Clinical Management in Borderline Situations Portal hypertension in clinical practice: Clinical management of pretreatment thrombocytopenia There is no conclusive evidence on the management of pretreatment thrombocytopenia; however, some alternatives can be considered and are reviewed in the Proceedings. Liver transplant in clinical practice: post-transplant limitations. Fibrogenesis, treatment after transplant, rejection control and live donor Liver disease caused by HCV corresponds to almost half of the indications for liver transplant. Currently, in Brazil, the distribution of the organ follows a criterion of severity, using the model for end-stage liver disease (MELD). After the transplant, up to 80% of the patients experience histologic recurrence, and the pre-transplant viral load is one of the most significant risk factors. For this reason, pre-transplant treatment should always be considered, even in patients with decompensated cirrhosis, provided that the treatment is given in a specialized center and with an active transplant treatment team. Post-transplant treatment for chronic hepatitis is generally indicated when fibrosis is ≥ 2 or when periportal activity is ≥ 3. Although the duration of treatment should be at least 48 weeks, it should be individualized according to the virologic response profile. Treatment Nondrug and nonspecific HCV treatment: metabolic syndrome, nonalcoholic steatohepatitis, diabetes, obesity – evidence for the use of diet, hypolipidemic agents, and antidiabetic medication It is recommended that the conditions associated with worsening of liver fibrosis and lower SVR rate, such as obesity, type 2 diabetes, nonalcoholic steatohepatitis, steatosis, dyslipidemia, and metabolic syndrome, be brought under control before antiviral treatment is instituted. Patients with concomitant diseases; how to use IFN-α and specific measures for drug users and patients with mental disease. Socially marginalized individuals and prisoners. Individuals with kidney disease in pre- and post-kidney transplant phases. Individuals with auto-immune disease and extrahepatic manifestations At facilities where a multiprofessional approach is taken or at specialized centers, both of which allow appropriate monitoring and control of all clinical situations of the underlying disease, as well as of those potentiated or triggered by HCV treatment, it is recommended that HCV treatment be instituted first. It is noteworthy that, in the case of auto-immune manifestations (cryoglobulinemia and auto-immune hepatitis in particular), treatment for HCV should be given – observing the premises www.bjid.com.br 4 Consensus of Hepatitis C Carried Out by SBI BJID 2007; 11 Supplement 1 (October) above. Corticosteroids and other immunosuppressants can be used concomitantly when indicated. a high viral load (> 850,000 IU/mL). The dose of PEG-IFN-α-2b should be 1.5 μg/kg/week. Ribavirin: Always a full dose? And how much (0.8 g, 1 g, 11 mg/kg/day, 15 mg/kg/day)? Ribavirin should always be given in a full dose. The recommended dose is 15 mg/kg/day or 1 g for patients who weigh < 75 kg. The minimum dose to be considered in adjustments is 11 mg/kg/day. Short, long, standard treatment ... Always individualize? In patients presenting a RVR, no comorbidities (severe steatosis or cirrhosis), and a low viral load (< 250,000 IU/ mL for genotype 1), the duration of the treatment can be reduced: to 12 weeks for genotypes 2 and 3; and to 24 weeks for genotype 1. Patients with a high viral load and complete EVR should be treated for the standard duration, and patients presenting a slow response (no RVR, partial EVR and negative at week 24) should be treated for an additional 24 weeks. Common adverse effects: how to deal with anemia, neutropenia and thrombocytopenia before the treatment, during the treatment, and triggered by the treatment? What are the basal limits for patients with and without cirrhosis to be treated? How can the effects be reversed? The minimum criteria for treatment should be hemoglobin ≥ 10 g/dL, neutrophils > 1500/mm3 and platelets > 60,000/mm3. Patients presenting a drop in hemoglobin to < 10 g/dL or a drop ≥ 3.5 g/dL with clinical manifestations should be given Erythropoietin in a dose of 40,000 IU/week until the end of the treatment or until anemia is under control. Patients in whom neutrophils drop to < 750/mm3 should receive filgrastim in a dose of 300 µg one to three times a week until neutropenia is under control. In patients presenting a drop in platelets to < 25.000/mm3, the treatment should be discontinued. Adjustment regimens of the dose of IFN with intermediate values are presented in the Proceedings. When to treat: the patient, the doctor, the system ... The patient wants to be treated. How can we deal with this? There is no definite position on this situation. We emphasize the need to give clear and exhaustive information regarding the indications for treatment and the factors associated with the progression of the disease. Is early indication of treatment worthwhile? In which cases? Treatment can be considered for individuals with level 1 structural lesion (F1 METAVIR), who present incomplete septa, A3 activity (METAVIR), who are under 60 years of age, and present co-factors associated with risk of progression (obesity, nonalcoholic steatohepatitis, etc.). Use of IFN and ribavirin The patient, the virus, and the medication: Relevant factors in predicting response before and during the treatment There has been no relevant alteration regarding the elements described. Please see the concepts listed below regarding individualization based on EVR. Is conventional treatment with IFN-α dead? If not, when should we use it? It is the understanding of the Brazilian Society of Infectious Diseases that treatment with conventional IFN-α is no longer justifiable and should not be performed. Dose of IFN, the ‘burden’ of the ‘weight’ The dose of pegylated IFN-α-2a (PEG-IFN-α-2a) should be 270 μg/week for patients weighing > 85 kg and presenting The fear of suspending treatment: How should we deal with that decision? We emphasize that, if the objective of the treatment is virologic, the absence of EVR and viral detection at week 24 necessarily imply the interruption of the treatment. Maintenance treatment with IFN: When and how? Is ‘watchful waiting’ still a valid concept? Maintenance treatment with PEG-IFN-α is considered for the following patients: those with structural lesion > F3 and/ or signs of portal hypertension (esophageal varicose veins, splenomegaly, dilated blood vessels, etc.) and/or platelet counts < 110,000/mm3 who are categorized as Child-Pugh class A or B, with no history of severe or potentially uncontrollable decompensation; those without hepatocellular carcinoma; relapsers; and partial responders or nonresponders to the combination of PEG-IFN-α and ribavirin (or to IFN only when ribavirin is contraindicated) administered with adequated compliance and for a minimum of 12 weeks. Maintenance treatment with PEG-IFN-α should also be considered for patients for whom the full dose treatment is contraindicated. Child-Pugh class C patients on the transplant waiting list should be treated in specialized centers. Proposed regimen2: PEG-IFN-α-2b, 0.5-1.0 μg/kg/week subcutaneously (sc); PEGIFN-α-2a, 90 μg/week (sc). Duration2: Minimum of 24 months, indefinite, or even until there is an antiviral treatment that is proven to be safe and efficient. Monitoring the occurrence of complications (hepatocellular carcinoma, gastrointestinal bleeding, encephalopathy, etc.) Monotherapy with ribavirin, although previously described, cannot be recommended at the moment, due to insufficient evidence. Impact of reduction in the dose of IFN and/or ribavirin at various moments of the treatment Every effort should be made to always maintain full treatment. Positive and negative predictive values in clinical practice: patients mono- and co-infected with HIV Positive predictivity data in the presence of RVR and negative predictivity data in the absence of EVR are equally 2 Dose schedule and duration were suggested based on preliminary data and should be re-evaluated taking into account the results of ongoing studies. www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Consensus of Hepatitis C Carried Out by SBI valid for HCV mono-infected patients and patients co-infected with HCV and HIV. Retreatment: When and how? Retreatment will always be considered when the previous treatment was considered suboptimal for the current concepts (poor compliance, insufficient doses, inappropriate interruptions, inadequate management of adverse events, uncontrolled comorbidities, etc.) and when there is recurrence. The treatment in true nonresponders to PEG-IFN and ribavirin presents such a low chance of success that it cannot be routinely recommended. Due to the fact that relapsers/nonresponders to initial treatment constitute a quite heterogeneous group, it is necessary to qualify and carefully select the patients that should be retreated. Various factors that might have influenced the nonresponse should be modified before the initiation of or even during the new therapeutic cycle. Currently, patients considered less likely to respond to retreatment are those who are true nonresponders, those who are of the Black race, those infected with genotype 1, those with high viral loads, those with advanced liver disease, and those who present intercurrent conditions (obesity, etc.) Patients previously treated with IFN as monotherapy or with the combination of IFN and ribavirin stand a greater chance of presenting an SVR than do nonresponders to the combination of PEG-IFN and ribavirin. Those who experienced recurrence during the treatment (breakthrough) or after the treatment fare better than do true nonresponders. Patients who are noncompliant with previous treatment, as well as those who required reduced doses of IFN or ribavirin due to cytopenia or other adverse effects, usually respond better to retreatment than do those who received full doses. Of course, the factors responsible for noncompliance should be eliminated, and reductions in medication doses should be properly approached. In nonresponders, it is recommended that erythropoietin and filgrastim be started sooner, and further dose reductions should be avoided at all costs. Users of drugs or alcohol who, due to their addiction, did not adequately comply with all the phases of the previous treatment, can more adequately respond to retreatment provided that these co-factors are nullified. This also applies to patients who did not receive adequate social or cultural support. Patients who suffer from obesity, insulin resistance, dyslipidemia, steatosis, anemia, or liver diseases (e.g., hemochromatosis) should be retreated, preferably after the proper diagnosis and treatment of these concomitant conditions. Doses of PEG-IFN should be the same as those used for treatment-naïve patients. We believe that, in retreatment regimens, the doses of ribavirin should be as high as possible. Due to the paucity of studies with large patient samples, we recommend that the duration of retreatment be 48 weeks for all genotypes. At the moment, there are no consolidated data in the literature to support using higher doses of PEG-IFN or ribavirin, using induction doses, or extending the treatment time to more than 48 weeks in retreatment cases. Regarding the week 12 rule, there is strong evidence that patients who do not present negativity for HCV RNA by this time will have very little chance of presenting an SVR, and their treatment should be interrupted. 5 IFN: Backbone of the current treatment. And the future? Perspectives and frustrations Preliminary results of developing therapies show that IFNα will still be the active principle of the treatment for many years. For this reason, we have adopted several of the practices that are presented here. Co-infections Antiretroviral therapy 2007 and HCV treatment: relevant interactions Although ribavirin can reduce the concentration of some antiretroviral nucleoside analog reverse transcriptase inhibitors (NARTIs), there seem to be no clinical consequences. The adverse effects of NARTIs have been associated with HCV co-infection, female gender, obesity and prolonged exposure to these drugs. The principal combinations with less hepatic repercussion among NARTIs are as follows: lamivudine + abacavir; lamivudine + zidovudine; lamivudine + abacavir + zidovudine, and emtricitabine + tenofovir. However, current studies indicate that ribavirin interferes with the effect of abacavir. Therefore, ribavirin should be used with caution in patients receiving the highly active antiretroviral therapy regimen and being treated for HCV. Greater toxicity is principally caused by concomitant use of didanosine and ribavirin. CD4 counts: Minimum limits for treatment and decision-making in patients with reconstituted immunity or immunocompetent patients Precise indication of therapy in HIV/HCV co-infected patients can be made in patients with CD4 counts > 350 cells/ mm3.. In patients with CD4 counts between 200 and 350 cells/ mm3, the decision to treat HCV infection should take into account other factors, such as the duration of HCV infection, the severity of the liver disease, the level of HIV suppression and classical predictors of treatment response to HCV, such as genotype and viral load. The SVR can be predicted when HCV RNA is undetectable in serum by week 4 of treatment. However, a < 2log10 IU/mL reduction in viral load by week 12 and/or viremia detected at week 24 predict a lack of virologic response, and discontinuation of the treatment is indicated. Ongoing studies are evaluating 72-week maintenance therapy in co-infected patients and week-24 nonresponders. This might be the truly appropriate treatment duration for HIV-positive patients, even with longer time and smaller fractionated doses. Is the immunocompetent HIV/HCV co-infected patient ‘monoinfected’? Yes, the evolution is quite similar. Multiple viral infections: Who should we treat first and how? Care should be given to the dominant virus between HBV and HCV. When treating the dominant virus, the other might become active. We need to quantify the HBV DNA in order to manage this situation. The HTLV apparently modulates the host immune response, a fact that should be considered and which is detailed in the Proceedings. There is no consensus regarding the correct course of action in multiple co-infections. www.bjid.com.br 6 BJID 2007; 11 Supplement 1 (October) Epidemiological Aspects of Hepatitis C in Brazil Decio Diament Emílio Ribas Institute of Infectious Diseases; São Paulo, SP, Brazil The true dimension of the epidemiological situation of chronic hepatitis C in Brazil remains unknown. According to data from the National Ministry of Health, 52,489 cases were diagnosed in the period from 1994 to 2005. However, the data regarding the final year of this series are incomplete. There is a trend toward an increase in the number of diagnosed cases during this period, since diagnostic techniques have become more widely available, both in public and private health care facilities (Figure 1). Figure 1. Number of diagnosed cases of hepatitis C reported by the National Ministry of Health, from 1994 to 2005, totaling 52,489 cases. Most diagnosed cases occur in the Southeast, followed by the South, Central-west, Northeast and North (Figure 2). Figure 2. Distribution of cases by macroregion (National Ministry of Health). The Brazilian Journal of Infectious Diseases 2007;11 (5) Suppl. 1:6-7. © 2007 by The Brazilian Journal of Infectious Diseases and Contexto Publishing. All rights reserved. This distribution reflects the greater availability of resources as well as the highly populated areas in the southeastern and southern regions of the country. However, serological surveys carried out in the Central-west and Northeast, as well as in the Federal District, showed prevalence rates similar to those of a previous survey carried out in the city of São Paulo (Figure 3). Figure 3. Prevalence of hepatitis C in serological surveys in several regions of the country (Galizzi Filho J, personal communication, for the data of the surveys on the Centralwest, Federal District and Northeast; Focaccia R., personal communication, for the data on São Paulo). Data obtained in these serological surveys indicate prevalence rates from 0.28 to 2.61%. According to the 2000 census taken by the Brazilian Institute of Geography and Statistics, Brazil has a population of approximately 170 million. It has been shown that, of those testing positive for infection with the hepatitis C virus (HCV), approximately 80% have the chronic form of hepatitis C. Based on these data, we can estimate that there are 400,000 to 3,800,000 cases of chronic hepatitis C in Brazil. Comparing these numbers with the data from the National Ministry of Health, we can conclude that there are a great number of undiagnosed cases of the disease. Since there has been a trend toward an increase in the number of diagnosed cases, there will be a significant increase in the demand for diagnosis and treatment of chronic hepatitis C in the future. Another interesting fact is that only 17,204 (33%) of the 52,493 cases of chronic hepatitis reported to the Center for Epidemiological Surveillance of São Paulo, from 1998 to 2006, were confirmed as being hepatitis C. Another 10,690 cases (20%) were classified as inconclusive for hepatitis C, 2099 (4%) were excluded, and 416 (0.79%) were cases of HBV/HCV co-infection. The remaining cases were confirmed as hepatitis B, HBV/HDV co-infection, or cases under investigation. www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Hepatitis C in Brazil Of the 17,204 confirmed cases, the principal form of contamination was parenteral (use of injected illicit drugs, as well as blood and blood product transfusions), followed by sexual transmission. Work-related accidents accounted for few of the cases (Figure 4). Figure 4. Distribution of cases of chronic hepatitis C by form of transmission in 17,204 individuals in the state of São Paulo, from 1998 to 2006 (Source: Center for Epidemiological Surveillance, São Paulo). Transmission was classified as unknown in 40% of the reported and confirmed cases, as can be seen in Figure 4. This suggests that the investigation of the form of transmission should be improved, and that items such as administration of medication using nondisposable syringes, acupuncture, tattoos, and piercings should be included in the epidemiological investigation. Another unusual fact is sexual transmission in 10% of the cases, well above the expected, which is less than 1%. Another aspect to be considered is the occurrence of hepatitis C in groups of differentiated risk, that is, unusual 7 cases such as those described in Figure 4. The first group would be that of the manicurists, since it is supposed that the community use of instruments by these professionals would expose them to a higher risk of contamination. There is no evidence that this could happen, and a recent serological survey conducted in the city of São Paulo in beauty salons located at shopping malls in various neighborhoods did not demonstrate prevalence rates different from those of the population in general (Oliveira, ACDS – personal communication). Another group is that of the dentists. In Brazil, localized serological surveys with small samples of patients have shown a low prevalence rate, ranging from 0.4 to 0.7%. However, a study on the level of knowledge of these professionals about the disease has shown it is very low, demonstrating the need of greater emphasis on training and continuing education, aiming at making them capable of properly protecting themselves, as well as of preventing transmission to patients and perhaps even help the diagnosis, since several extrahepatic manifestations appear in the mouth cavity, such as oral liquen planus, erythema nodosum, etc. References 1. Bellíssimo-Rodrigues W.T., Machado A.A., Bellíssimo-Rodrigues F., et al. Prevalence of hepatitis B and C among Brazilian dentists. Infect Control Hosp Epidemiol 2006;27:887-8. 2. h t t p : / / p o r t a l . s a u d e . g o v . b r / p o r t a l / s a u d e / visualizar_texto.cfm?idtxt=25340. Acessado em 24/08/2007. 3. http://www.cve.saude.sp.gov.br/htm/hepa_home.html. Acessado em 24/08/2007. 4. Leão J.C., Teo C.G., Porter S.R. HCV infection: aspects of epidemiology and transmission relevant to oral health care workers. Int J Oral Maxillofac Surg 2006;35(4):295-300. 5. Takahama A.J., Tatsch F., Tannus G., Lopes M.A. Hepatitis C: incidence and knowledge among Brazilian dentists. Community Dent Health 2005;22(3):184-7. www.bjid.com.br 8 BJID 2007; 11 Supplement 1 (October) Sexual Transmission of HCV Kleber Dias do Prado Emílio Ribas Institute of Infectious Diseases; São Paulo, SP, Brazil Although there is evidence that sexual transmission of HCV occurs, this form of transmission is of secondary epidemiological importance when compared to percutaneous forms of transmission [1]. Among the evidence that supports the possibility of sexual transmission of HCV we can list the following: 1- Case reports of acute hepatitis C with anti-HCV seroconversion in sexual partners of individuals infected by HCV, excluding mechanisms of nonsexual transmission and with high genomic homology among viral strains infecting sexual partners [2-4]. 2- Detection of HCV RNA in semen, vaginal secretion and cervical secretion, despite low titles in most cases [5-8]. 3- Data from the Centers for Disease Control and Prevention demonstrating that, from 1995 to 2000, 18% of the cases of acute HCV infection in the United States occurred in patients reporting sexual contact with an individual infected with HCV in the preceding 6 months or multiple sexual contacts as the only risk factors for acquiring the infection [1]. The risk of acquiring HCV through sexual contact differs among subgroups of individuals: We can distinguish two main risk subgroups: 1. Individuals who have multiple sexual partners or who engage in sexual practices that might lead to mucosal trauma: sex professionals; men who have sex with men (MSM); and patients treated in clinics specializing in the treatment of sexually transmitted diseases (STDs). 2. Stable monogamous heterosexual sexual partners of individuals chronically infected with HCV. In general, rates of anti-HCV incidence and prevalence are higher in the first subgroup. This may be due to differences in sexual practices among the groups, but also to nonsexual factors (sharing personal objects, tattoos, use of illicit drugs, etc.) [1]. Seroprevalence studies in the United States demonstrated median positive anti-HCV rates in 6% of women who were sex professionals, as well as in 4% of MSM, 4% of clients of STD clinics and 4% of participants in HIV surveillance studies. Studies conducted in other parts of the world have obtained similar results [9-15]. The following risk factors were identified: having had a high number of recent and lifetime sex partners; engaging in unsafe sexual practices; being infected with HIV; and having an STD. This indicates that sexual activity in general is a risk factor for HCV The Brazilian Journal of Infectious Diseases 2007;11 (5) Suppl. 1:8-9. © 2007 by The Brazilian Journal of Infectious Diseases and Contexto Publishing. All rights reserved. transmission [1]. However it is essential to mention that the results of some studies contradict these findings. Studying the prevalence and incidence of positivity for anti-HCV antibodies in a cohort of 1085 HIV-positive Canadian MSM, Alary et al. found values of 2.9% and 0.038/100 individuals/year, respectively, both significantly associated with the use of injection drugs. The authors considered sexual transmission of HCV to be rare in this group [16]. In a study conducted in Thailand, Taketa et al. assessed the prevalence of anti-HCV in injection drug users, sex professionals and individuals with STDs. The prevalence was 85%, 2% and 0%, respectively, with a very low or null transmission rate in the last two groups [17]. Marincovich et al. prospectively studied a group of 171 couples discordant for HIV and HCV. The index cases were 152 men and 19 women, whereas the spouses were 152 women and 19 men. Fortythree per cent had engaged in unprotected vaginal and/or anal sex, 15% always used a condom but reported incidents in which the condom broke or slipped off during sexual contact, and 22% had performed unprotected orogenital sex. There was only one case of HIV seroconversion and no cases of HCV seroconversion during the follow-up of 529 individuals/ year. There were 31 cases of pregnancy, 2 of them in women infected with HCV. This study suggests that the rate of HCV transmission is low or null among heterosexuals, even when the partner is infected with HIV [18]. For the second subgroup, the best studies are those excluding percutaneous factors of infection and evaluating genotypes and genomic sequence of viral strains in anti-HCV concordant couples. In those studies, the prevalence of HCV was estimated at 2.8-11% in the Asian Southeast, 0-6.3% in Northern Europe and 2.7% in the United States [1]. In one of the first studies to use genotyping and analysis of the sequence of nucleotides of the hypervariable E2 region, Zylberberg et al., studying 24 anti-HCV concordant couples, reduced to 3 couples the possible cases of sexual transmission of HCV. Nevertheless, nonsexual factors could not be ruled out and might have contributed to HCV transmission between couples [19]. In Iran, Hajiani et al. studied the HCV transmission rate for home contacts with no percutaneous risk factors. The rates found were 1.33% for the contacts and 1% for the controls (p>0.06). Only 2 of 59 spouses presented evidence of infection (3.39%). The authors conclude that intrafamily transmission is possible, although not common [20]. In a recent study, McMahon et al. determined that the transmission of HCV in 265 heterosexual couples using drugs in New York City was associated with the use of injection drugs by the couple, although not with the pattern of sexual activity [21]. Along the same lines, Boonyarad et al., studying 160 spouses infected with chronic hepatitis C (106 women www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Sexual Transmission of HCV and 54 men, all monogamous, stable heterosexual couples), verified that only 3 individuals (1.88%) tested positive for anti-HCV antibodies and HCV RNA after a mean period of 23 ± 5 years of unprotected sexual exposure. Nevertheless, in those 3 individuals, genotyping and sequence analysis did not clearly identify the same viral strains that infected their respective partners. The authors concluded that sexual transmission of HCV is rare [22]. Finally, Vandelli et al. conducted a large prospective study in which 895 monogamous and stable heterosexual couples were evaluated for 10 years [23]. Among the spouses, the authors identified 3 cases of seroconversion, none of which were attributed to sexual contact with the partner: in one case, the genotypes were different; in the other two, there were discrepancies in the sequence and phylogenetic analysis). Therefore, the authors conclude that the rate of sexual transmission of HCV is very low or even null in these patients. Therefore, it seems unnecessary to recommend condom use in this population. These couples did not practice anal sex, neither sex during menstruation nor used condoms [23]. The risk of sexual transmission of HCV ranges from 00.6%/year for heterosexual couples in monogamous, stable relationships to 1%/year to individuals with several sex partners [1]. Therefore, the formal, systematic recommendation of condom use is only necessary for the latter group. Condom use is also justified in HCV-positive individuals presenting concomitant STDs, having sex during menstruation or engaging in sexual practices that can traumatize the mucosal surfaces (anal sex, fisting, etc.) In conclusion, the sharing of objects potentially contaminated with blood, such as razor blades, scissors, nail clippers, cuticle trimmers and tooth brushes, is not recommended [1]. References 1. Terrault N.A. Sexual activity as a risk factor for hepatitis C. Hepatology 2002;36:S99-105. 2. Halfon P., Riflet H., Renou C., et al. Molecular evidence of maleto-female sexual transmission of hepatitis C virus after vaginal and anal intercourse. J Clin Microbiol 2001;39:1204-6. 3. Nakayama H., Sugai Y., Ikeya S., et al. Molecular investigation of interspousal transmission of hepatitis C virus in two Japanese patients Who acquired acute hepatitis C after 40 or 42 years of marriage. J Med Virol 2005;75(2):258-66. 4. Lai K.W., Young K.C., Cheng P.N., et al. Interspousal transmission of hepatitis C virus: application of comparing the variability of HVR1 nucleotide region. Hepatogastroenterology 2004;51(57):791-5. 5. Leruez-Ville M., Kunstmann J.M., De Almeida M., et al. Detection of hepatitis C virus in the semen of infected men. Lancet 2000;356:42-3. 6. Manavi M., Watkins-Riedel T., Kucera E., et al. Evidence of hepatitis C virus in cervical smears. J Infect 1999;38:60-1. 7. Pekler V.A., Robbins W.A., Nyamathi A., et al. Use of versant TMA and bDNA 3.0 assays to detect and quantify hepatitis C virus in semen. J Clin Lab Anal 2003;17(6):264-70. 9 8. Bélec L., Legoff J., Si-Mohamed A., et al. Cell-associated, nonreplicating strand (+) hepatitis C virus-RNA shedding in cervicovaginal secretions from chronically HCV-infected women. J Clin Virol 2003;27(3):247-51. 9. Van de Laar T.J., Van der Bij A.K., Prins M., et al. Increase in HCV incidence among men who have sex with men in Amsterdam most likely caused by sexual transmission. J Infect Dis 2007;196(2):230-8. 10. Danta M., Brown D., Bhagani S., et al. Recent epidemic of acute hepatitis C virus in HIV-positive men who have sex with men linked to high-risk sexual behaviours. AIDS 2007;21(8):983-91. 11. Götz H.M., Van Doornum G., Niesters H.G., et al. A cluster of acute hepatitis C virus infection among men who have sex with men: results from contact tracing and public health implications. AIDS 2005;19(9):969-74. 12. D’Oliveira A. Jr., Voirin N., Allard R., et al. Prevalence and sexual risk of hepatitis C virus infection when human immunodeficiency virus was acquired through sexual intercourse among patients of the Lyon University Hospitals, France, 1992-2002. J Viral Hepat 2005;12(3):330-2. 13. De los Angeles Pando M., Biglione M.M., Toscano M.F., et al. Human immunodeficiency virus type 1 and other viral coinfections among young heterosexual men and women in Argentina. Am J Trop Med Hyg 2004;71(2):153-9. 14. Marx M.A., Murugavel K.G., Tarwater P.M., et al. Association of hepatitis C virus infection with sexual exposure in southern India. Clin Infect Dis 2003;37(4):514-20. 15. Russi J.C., Serra M., Viñoles J., et al. Sexual transmission of hepatitis B virus, hepatitis C virus, and human immunodeficiency virus type 1 infections among male transvestite commercial sex workers in Montevideo, Uruguay. Am J Trop Med Hyg 2003;68(6):716-20. 16. Alary M., Joly J.R., Vincelette J., et al. Lack of evidence of sexual transmission of hepatitis C virus in a prospective cohort study of men who have sex with men. Am J Public Health 2005;95(3):502-5. 17. Taketa K., Ikeda S., Suganuma M., et al. Differential seroprevalences of hepatitis C virus, hepatitis B virus and human immunodeficiency virus among intravenous drug users, commercial sex workers and patients with sexually transmitted diseases in Chiang Mai, Thailand. Hepatol Res 2003;27(1):6-12. 18. Marincovich B., Castilla J., Del Romero J., et al. Absence of hepatitis C virus transmission in a prospective cohort of heterosexual serodiscordant couples. Sex Transm Infect 2003;79(2):160-2. 19. Zylberberg H., Thiers V., Lagorce D., et al. Epidemiological and virological analysis of couples infected with hepatitis C virus. Gut 1999;45(1):112-6. 20. Hajiani E., Masjedizadeh R., Hashemi J., et al. Hepatitis c virus transmission and its risk factors within families of patients infected with hepatitis C virus in southern Iran: Khuzestan. World J Gastroenterol 2006;12(43):7025-8. 21. McMahon J.M., Pouget E.R., Tortu S. Individual and couple-level risk factors for hepatitis C infection among heterosexual drug users: a multilevel dyadic analysis. J Infect Dis 2007;195(11):1556-9. 22. Boonyarad V., Chutaputti A., Choeichareon S., et al. Interspousal transmission of hepatitis C in Thailand. J Gastroenterol 2003;38(11):1053-9. 23. Vandelli C., Renzo F., Romanò L., et al. Lack of evidence of sexual transmission of hepatitis C among monogamous couples: results of a 10-year prospective follow-up study. Am J Gastroenterol 2004;99(5):855-9. www.bjid.com.br 10 BJID 2007; 11 Supplement 1 (October) Hepatitis C Virus Perinatal Transmission Umbeliana Barbosa de Oliveira Emílio Ribas Institute of Infectious Diseases; São Paulo, SP, Brazil The hepatitis C virus (HCV) is the most frequent cause of chronic hepatic disease. Its principal route of transmission is exposure to contaminated blood. Perinatal transmission is one of the less common modes of infection with HCV. Incidence Vertical transmission of HCV has been reported in numerous studies. However, the estimated rate of vertical transmission varies considerably due to several factors: • Study methodology; • Selection of maternal population; • Risks involved in the transmission, such as coinfection with HIV and high levels of HCV RNA. A review of the literature carried out in 1998 identified 976 children from 28 studies with follow-up periods that were sufficiently long to estimate the transmission rate. In those studies, the vertical transmission rate was below 10% in cases involving HIV-negative mothers. The risk of vertical transmission increased significantly in cases involving HIVpositive mothers. It is believed that, in these cases, the mother tends to present higher levels of HCV viremia, increasing the risk of transmission. In a more recent review (published in 2001), the authors searched through studies published in the 1992-2000 period and identified 77 studies. They found clinical and demographic variables that influenced the transmission rate. They concluded that co-infection with HIV is the most significant factor associated with the risk of vertical HCV transmission. Findings regarding other possible risk factors, such as HCV genotype, type of delivery, and breastfeeding, have been inconclusive. Risk Factors As previously mentioned, the principal risk factor for vertical transmission of HCV is concomitant infection with HIV. Viremia Level Some studies have shown that viral load can be an important determinant factor for vertical transmission of HCV. It has been demonstrated that women with viral loads lower than 1 × 105 copies per mL have a lower risk of vertical transmission, whether co-infected with HIV or not. One study that compared viral loads among HCV RNA-positive mothers whose children were infected with HCV or not showed that, in cases of probable vertical transmission, the maternal viral load was ten times higher than in those cases in which transmission did not occur. However, there are also inconsistent data in two large studies that found no significant differences between the HCV RNA levels of the mothers who transmitted HCV and those of mothers who did not. The Brazilian Journal of Infectious Diseases 2007;11 (5) Suppl. 1:10-11. © 2007 by The Brazilian Journal of Infectious Diseases and Contexto Publishing. All rights reserved. Co-Infection with HIV Numerous studies have shown that the HCV vertical transmission rate is higher among HIV-positive women that among those who are HIV-negative (19% vs. 4%). This is in part explained by the fact that HIV-positive women present higher viral loads of HCV. One study showed that the average level of HCV RNA in women co-infected with HIV was ten times higher than in women mono-infected with HCV. This shows that the increase in the co-infection risk can be nullified after adjusting the risk for the viral load, and antiretroviral therapy for HIV can reduce the risk of HCV transmission. One study that compared HIV-positive women and HIVnegative women with similar viral loads of HCV showed that the risk of HCV transmission was similar in both groups. In that study, all HIV-positive women were receiving antiretroviral therapy. Another study showed that children infected with HIV are at a higher risk of being infected with HCV than are those who are HIV-negative (17.1% vs. 5.4%). The explanation for these data remains unclear. Genotype There is no evidence that HCV genotype influences vertical transmission. The data collected to date do not allow us to establish a relationship between HCV genotype and vertical transmission risk. Breastfeeding It is possible to detect HCV RNA in breast milk and colostrum. However, HCV transmission through breastfeeding has not been documented. A likely explanation would be that HCV is inactivated by the effect of gastric acidity and that the HCV RNA levels in breast milk are very low. Therefore, breastfeeding does not increase the risk that HCV will be transmitted from an infected mother to her child. The American College of Obstetricians and Gynecologists, as well as the American Academy of Pediatrics, allows breastfeeding by mothers who are infected with HCV. It is important to emphasize that breastfeeding is not recommended in case of nipple fissure, due to the possibility of bleeding. Type of Delivery The impact that the type of delivery has on perinatal transmission of HCV is completely unknown. Vaginal delivery has been associated with an increase in the transmission risk. In a study carried out by Lin et al. in 1994, 70 pregnant women were evaluated. The vertical transmission rate among children who were born by vaginal delivery was 32%, compared to 6% in cases of cesarean section. However, the high transmission rate in the Lin et al. study was associated with the fact that a large proportion of the women evaluated (76%) were coinfected with HIV and HCV. Therefore, cesarean section is associated with a decrease in the HCV transmission risk in co- www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Hepatitis C Virus Perinatal Transmission infected women. The higher risk of transmission in vaginal delivery might be associated with exposure of the newborn to HCV-contaminated blood during its passage through the vagina. However, the exposure to blood can sometimes be greater in cesarean section. Many studies are needed in order to estimate the risk of perinatal transmission of HCV. Currently, cesarean sections are not recommended for pregnant women infected with HCV. Other risk factors associated with delivery are rupture of the placental membrane six hours or more before delivery and invasive procedures such as internal monitoring of the fetus (Blood 2000; 96:2045). Clinical Manifestations Newborns infected with HCV are generally asymptomatic. In these cases, a significant proportion presents normal alanine aminotransferase (ALT) levels. One study monitored 104 children with vertical transmission upon birth for approximately 49 months (J Infect Dis 2000;181:419). Although HCV RNA levels were positive for HCV in 90% of these children, the clinical manifestations were rare. None of them presented hepatic insufficiency. The majority presented persistent or transitory increase in ALT levels. Levels of ALT were, in general, normal or slightly increased at birth, evolving to an increase in the fourth to sixth months of age, remaining high for two years, and declining significantly thereafter. In that study, 20 children were submitted to hepatic biopsy. In all cases, evidence of chronic hepatitis was found. The degree of inflammation varied from mild to moderate. In three cases some level of fibrosis was found. Other studies suggest that children infected with verticallytransmitted HCV are generally asymptomatic. However, such transmission is often accompanied by an increase in ALT levels during the first 6 to 12 months of life. One large study monitored 266 perinatally HCV-infected children for an average of 4.2 years. In that study, approximately 20% of the children presented spontaneous clearance of HCV, and 80% evolved to chronic infection. Clearance of HCV is defined as a loss in the polymerase chain reaction positivity for HCV RNA in those children who maintained detectable HCV RNA in the first year of life. Most of those children were asymptomatic. Further studies involving long-term follow-up evaluation are needed in order to determine what proportion of children with chronic hepatitis C caused by vertical transmission will develop hepatic insufficiency and hepatocarcinoma. Diagnosis The initial diagnosis of infection with HCV in adults is made through anti-HCV antibody testing. However, this test presents limitations when used in children. This is primarily due to the passage of IgG from the mother to the child through the placental barrier. Therefore, the presence of anti-HCV in the serum of the child does not necessarily indicate HCV infection. Clearance of the anti-HCV passively acquired from the mother can take more than 12 11 months, although it occurs within 12 months in 95% of the cases. Second, the presence of maternal anti-HCV does not necessarily mean that the mother is chronically infected with HCV. A diagnosis of vertical transmission is made by identifying HCV RNA in the mother and child. It is important to emphasize that the HCV RNA test can also present false-positive or false-negative results. A falsepositive result can occur as a result of contamination of positive samples. A false-negative result occurs due to a loss of RNA during sample storage or to an error in RNA extraction. A consensus from the National Institutes of Health recommends that children born to mothers testing positive for HCV should be submitted to the HCV RNA test on two occasions: between 2 and 6 months of life and after 15 months of life (together with anti-HCV tests on the latter occasion). Conclusions The incidence of vertical transmission by HCV is approximately 2% to 5%. The risk is higher in the following situations: co-infection with HIV; and high maternal viral load of HCV. There are still no effective interventions to reduce the risk of HCV transmission from the mother to the child. As yet, HCV testing is not recommended for pregnant women. References 1. Manzini P., Saracco G., Cerchier A., et al. Human immunodeficiency virus infection as risk factor for mother-tochild hepatitis C virus transmission; persistence of anti-hepatitis C virus in children is associated with the mother’s anti-hepatitis C virus immunoblotting pattern. Hepatology 1995;21:328. 2. Thomas S.L., Newell M.L., Peckham C.S., et al. A review of hepatitis C virus (HCV) vertical transmission: risks of transmission to infants born to mothers with and without HCV viraemia or human immunodeficiency virus infection. Int J Epidemiol 1998;27:108. 3. Yeung L.T., King S.M., Roberts E.A. Mother-to-infant transmission of hepatitis C virus. Hepatology 2001;34:223. 4. Terrault N. Epidemiological evidence for perinatal transmission of hepatitis C virus. Viral Hepatitis Reviews 1998;4:245. 5. Lin H.H., Kao J.H., Hsu H.Y., et al. Possible role of high-titer maternal viremia in perinatal transmission of hepatitis C virus. J Infect Dis 1994;169:638. 6. Kumar R.M., Shahul S. Role of breast-feeding in transmission of hepatitis C virus to infants of HCV-infected mothers. J Hepatol 1998;29:191. 7. Azzari C., Resti M., Moriondo M., et al. Vertical transmission of HCV is related to maternal peripheral blood mononuclear cell infection. Blood 2000;96:2045. 8. Tovo P.A., Pembrey L.J., Newell M.L. Persistence Rate and Progression of Vertically Acquired Hepatitis C Infection. J Infect Dis 2000;181:419. 9. Vogt M., Lang T., Frosner G., et al. Prevalence and clinical outcome of hepatitis C infection in children who underwent cardiac surgery before the implementation of blood-donor screening. N Engl J Med 1999;341:866. 10. Three broad modalities in the natural history of vertically acquired hepatitis C virus infection. Clin Infect Dis 2005;41:45. www.bjid.com.br 12 BJID 2007; 11 Supplement 1 (October) Hepatitis C: Virological Aspects and Practical Implications Antonio Alci Barone Laboratory of Hepatitis, LIM 47 DMIP-HC-FMUSP; São Paulo, SP, Brazil The hepatitis C virus (HCV) is a member of the Hepacivirus genus of the family Flaviviridae. Other important pathogens for humans, such as the dengue virus and the yellow fever virus, belong to this family, as do infectious agents in animals, such as the bovine viral diarrhea virus and the classical swine flu virus. The shape of HCV is spherical (50 nm in diameter), and it has an enveloped nucleocapsid and a single-stranded RNA genome of positive polarity. The HCV genome, with approximately 10,000 nucleotides, consists of a single long open reading frame (ORF) flanked by two noncoding regions (NCRs) at the initial 5’ end, at which the internal ribosome entry segment (IRES) is located, and at the 3’ end. Its translation yields a large polyprotein (with nearly 3000 amino acids) that is processed by viral and host cell proteases into structural proteins, including the core, envelope 1 (E1), E2, and p7 proteins, as well as the nonstructural proteins NS2, NS3, NS4, and NS5. Recently, an alternate reading frame that codifies an F protein with more than 160 amino acids has been identified. However, its expression in natural HCV infection has not been confirmed. Structural proteins are cleaved by enzymes of the parasitized cell. Envelope proteins are extensively glycosylated and are involved in the binding with receptors as well as in the entrance and fusion of the virus. The function of p7 protein remains unknown. Nonstructural proteins, initially NS2 and subsequently NS3, undergo self-cleavage and position themselves in transmembrane domains across the host cell membrane and into the cytosol or lumen (Figure 1). Unlike that of the hepatitis B virus (HBV), the HCV genome does not invade the infected cell nucleus. After the binding through receptors (CD81, a tetraspanin, and the low density lipoprotein receptor), the HCV genome acts directly as an mRNA in the cytoplasm, where the translation is initiated through the IRES in the 5’ NCR. The protein produced is subsequently processed by the cell enzymes and by the enzymes within the virus itself, yielding structural and nonstructural proteins. After synthesis and maturation, these nonstructural proteins and the RNA form replication complexes that combine with the membrane and catalyze the translation of intermediate negative strands of RNA, from which positivestrand progeny are generated. The genomic RNA and capsid proteins unite, forming the nucleocapsid, which is transported in cytoplasmic vesicles. While passing through the Golgi complex, these vesicles assemble with the other particles and undergo exocytosis and cell release (Figure 2). The study of the HCV genome, even in samples obtained from a single individual, reveals great heterogeneity among the HCV genotypes. Genotypes in which multiple mutants The Brazilian Journal of Infectious Diseases 2007;11 (5) Suppl. 1:12-13. © 2007 by The Brazilian Journal of Infectious Diseases and Contexto Publishing. All rights reserved. coexist have been designated quasispecies. The multiple mutations represent a rapid and very efficient mechanism for the virus to evade the immune response and persist in the host. The selection process and the process of adaptation to the host have led to the evolution to different HCV genotypes. The classification system most commonly used is that proposed by Simmonds et al. and is based on the similarity of the sequence of nucleotides using the following criteria: similarity lower than 72% characterizes a new genotype; similarity between 75 and 86% characterizes a new subgenotype. There are 6 genotypes, which are numbered from 1 to 6, with subgenotypes 1a, 1b, 1c, 2a, 2b, 2c, 3a, 3b, 4a, 5a, and 6a. Although the criterion is based on molecular biology, this classification has practical, pathogenetic, epidemiological, and treatment-related implications. Therefore, subgenotype 1a is more prevalent in the USA, 1b in Japan, 3a in Scotland, and 4a in Egypt/ Zaire. In Brazil, genotype 1 is found in approximately 60% of the patients, followed by genotype 3, which is found in 20 to 30%, and genotype 2, which is found in a lower percentage. Subgenotype 1b can cause severe forms of the infection and, similar to genotype 4, does not respond as well to treatment with IFN-α. Therefore, genotypes 1 and 4 should be treated for 48 weeks. The important advances in the knowledge of viral hepatitis B and C are a consequence of some facts that will be discussed herein. The Use of Chimpanzees as a Model for the Study of Viral Hepatitis Although chimpanzees are not natural hosts for these viruses, they reproduce the disease, thus allowing important discoveries: • Epidemiological studies: knowledge of these diseases as communicable; • Infectivity and titers of pools of HCV and HBV, obtained from the infected animals; • Infectivity of the molecular clones of HCV and importance of genetic elements specific to HCV; • Neutralization capacity of antibodies specific for HBV and HCV; • Protective immunity, tested through re-exposure to the viruses; • Mutants that escape to humoral and cellular immunity could be recognized. However, the use of nonhuman primates has advantages and disadvantages. Advantages The only animal susceptible to the acute and chronic forms of the disease; non-selected population; and sequential biopsies. www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Virological Aspects of Hepatitis C Figure 1. Organization of the HCV genome, polyprotein processing, and protein topology [1]. Figure 3. Systems used in the HCV in vitro study [1]. Disadvantages Ethical considerations regarding the use of primates; cost and availability; rarity of vertical transmission; more benign disease presentation; and weaker, more limited immune response Use of Replicons In HCV, subgenomic RNA replicons are those in which the structural region has been replaced by the neomycin phosphotransferase gene, and the translation of the nonstructural proteins is regulated by the IRES of the encephalomyocarditis virus. Through this experimental approach, it became possible for the first time to effectively and efficiently replicate HCV in vitro in cultured Huh-7 human hepatoma cells. Interestingly, some simple amino acid substitutions can increase the replication efficiency by up to 10,000 times in all of the nonstructural proteins. The replicon system allowed the clarification of important aspects of the virus life cycle, as well as simplifying the evaluation of new antiviral strategies. However, some aspects of the virus life cycle cannot be studied using this system. 13 Figure 2. HCV cycle in the host cell [1]. Efficient Cell Culture Systems for HCV A 32-year-old male patient of Asian origin presented a profile consistent with fulminant hepatitis. Using reverse transcriptase polymerase chain reaction, HCV RNA was detected in the serum during the acute phase and not during the remission phase. Using these samples, the complete HCV genome was recovered and cloned. This strain, designated JFH1, has 9678 bp in the genome, with a single long ORF (nt 341-9439), and encodes 3033 amino acids The transfection of this sample into HUH-7.5.1 cultured cells allowed three independent groups of researchers to obtain HCV infecting particles, for tissue culture as well as for chimpanzee. These studies [2-4] were published, between June and July of 2005, in the following journals: Proceedings of the National Academy of Sciences, Nature Medicine, and Science. The infectious virions of HCV obtained in this way have been used to infect laboratory animals and naïve cells. Such infection can be monitored by the detection of the expression of the NS5A, through analysis of reporter genes, or by direct measurement of viral RNA (Figure 3). In conclusion, more recent findings regarding the virological aspects of HCV have greatly increased the possibility of thoroughly studying this agent and especially its relationship with the human host. Based on this approach, it will be possible to better understand how to combat the virus in chronically infected patients, thus preventing the progression of the disease and its consequences. References 1. Tellinghuissen T.L., et al. Studying Hepatitis C virus: making the best of a bad virus. Journal of Virology 2007;81(17):8853-67. 2. Barthenschlager B., et al. Efficient hepatitis C virus cell culture: what a difference the host cell makes. PNAS 2005;102(28):973940. 3. Wakita T., et al. Production of infectious hepatitis C virus in tissue culture from a cloned viral genome. Nature medicine 2005;11(7):791-6. 4. Lindenbach B.D., et al. Complete replication of hepatitis C virus in cell culture. Science 2005;309:623-6. 5. Kato T., et al. Sequence analysis of Hepatitis C virus isolated from a fulminant hepatitis patient. J Med Virol 2001;64:334-9. www.bjid.com.br 14 BJID 2007; 11 Supplement 1 (October) Pathogenesis of Hepatitis C – HCV Consensus 2007 Ana Tereza R. Viso Laboratory of Hepatitis, LIM 47 DMIP-HC-FMUSP; São Paulo, SP, Brazil The chronic hepatitis C virus (HCV) infects approximately 130 million people worldwide [1]. It is estimated that approximately 15% of HCV-infected individuals eliminate the virus spontaneously, that 25% develop a mild form of the disease, and that 60% develop the chronic progressive form [2]. The elimination or persistence of HCV infection depends on the balance between the effectiveness, specificity and rapidity of the innate and adaptive immune responses, as well as on the HCV replication rate [3]. Persistence of HCV can also be caused by infection at privileged (extrahepatic) sites, viral inhibition of antigen presentation, selective immune suppression, negative regulation of HCV gene expression, viral mutations, immune exhaustion of T cells and the incomplete differentiation of memory T cells [4,5]. Fibrosis is the principal complication of chronic hepatitis C, and it is estimated that 20% of patients develop cirrhosis over a period of 10, 20 or 30 years [2,6]. The progression of fibrosis increases morbidity and mortality in chronic hepatitis C [7], since it can lead to death due to complications caused by cirrhosis or hepatocarcinoma [2]. Various studies have associated the progression of fibrosis in hepatitis C with diverse factors such as: the kinetics and pathogenicity of HCV; host-HCV interaction; intrinsic host factors such as demographic profile, body mass index and diabetes mellitus; host exposure to external factors; and the form of HCV acquisition. Life Cycle and Pathogenicity of HCV Belonging to the Flaviviridae family, HCV is a small enveloped virus [8]. Its genome consists of one RNA molecule that is composed of two terminal regions, 5’- and 3’untranslated regions, and between these there is a single open reading frame that encodes a polyprotein with approximately 3000 amino acids. This polyprotein cleaves at the N-terminal side of three structural proteins, the nucleocapsid (core), envelope 1 (E1) and envelope 2 (E2), all of which are involved in the architectural organization of HCV. At the carboxylterminal side, the polyprotein cleaves to six nonstructural proteins, NS2, NS3, NS4 (NS4A and NS4B), NS5 (NS5A and NS5B) and NS6, which are responsible for the life cycle of the virus [9]. After entering a susceptible host, HCV invades, infects and replicates within the blood stream, repeating the process in various tissues, as well as in peripheral B and T lymphocytes, as it proceeds to the liver by tropism, passing through various tissues such as those of the pancreas, thyroid, adrenal glands, The Brazilian Journal of Infectious Diseases 2007;11 (5) Suppl. 1:14-19. © 2007 by The Brazilian Journal of Infectious Diseases and Contexto Publishing. All rights reserved. spleen and bone marrow [10-12]. Since HCV can also directly infect the lymphatic tissue, its stimulation can lead to the development of B-cell lymphomas [13]. It is known that the liver is the principal site of HCV replication, and various studies have shown that this virus infects approximately 10% of hepatic cells [5]. Infection with HCV at extrahepatic sites can promote the appearance of HCV variants [14,15], thereby decreasing the chance that the immune system will recognize the virus. To enter the host cell, HCV E2 and E1 proteins recognize and bond with the CD81 receptors present on the surface of hepatocytes and lymphocytes [16,17]. Circulating HCV particles are accompanied by low-density and very lowdensity lipoproteins, which prompts discussion in the literature regarding the possibility that low-density lipoprotein is also a viral receptor [15]. After the interaction of the virus envelope with the host cell membrane, HCV enters the cell through endocytosis. In the cytoplasm, the messenger RNA then undergoes translation, and polyproteins are processed; the HCV RNA then replicates, after which the new viral ‘RNA’s are packaged and transported to the surface of the host cell so that they can disseminate and complete a new cycle [18]. The HCV replication rate is high, approximately 1 × 1012 virions per day; this, together with its high mutation rate, estimated at 10-3 nucleotide substitutions per year, leads to great heterogeneity in its presentations, which are known as quasispecies [8]. The selection of and host adaptation to HCV quasispecies have given rise to distinct genotypes [19] whose classification is based on the similarity of the sequence of nucleotides: similarity below 69% characterizes a new viral type; and similarity between 75 and 80% characterizes a subtype [20]. The progression of fibrosis in chronic hepatitis C has been associated with the diversity of HCV quasispecies [21]. The production of new viruses is counterbalanced by the destruction of infected cells through tissue apoptosis or degradation in peripheral blood, since the half-life of the virus in peripheral blood is approximately 2.7 hours [7]. Experimental studies have shown that NS3 and NS5 proteins induce apoptosis in infected hepatocytes [22]. In individuals infected with HCV, the persistence of the virus can be attributed the large inoculum and the high rate of viral replication, which allow the virus to evade the host immune response [4,23]. There is controversy over whether the sequence of nucleotides is directly associated with more intense hepatic lesions [6,24,25]. There is some evidence of direct cytopathic lesion caused by HCV, including HCV-induced histological lesions with scant inflammatory infiltrate [26-28], fulminant hepatitis C after chemotherapy in liver transplants [29] and HCV-related acute cholestatic syndrome after renal transplantation [30]. www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Pathogenesis of Hepatitis C Studies suggest that specific genotypes, such as genotype 1, can be more cytopathic [31] or can induce more rapid progression of the disease than do other genotypes [32]. Genotype 1 has been shown to be the genotype most strongly associated with chronic HCV infection [33]. The risk of cirrhosis and hepatocarcinoma has been shown to be greater in individuals presenting genotype 1b than in those presenting genotypes 2 and 3 [34]. However, other authors have stated that HCV genotype and viral load do not influence the progression of the disease [2,6]. It is known that steatosis is a cofactor that influences the progression of fibrosis in chronic hepatitis C [35]. Various studies have directly associated steatosis with HCV genotype 3 [36,37]. Therefore, genotype 3 is considered cytopathic [38,39]. Kumar et al. identified the reduction of steatosis as the only variable predictive of the virological response to the treatment of chronic hepatitis C in individuals infected with HCV genotype 3 [38]. Innate Immune Response to HCV The innate immune response to HCV is responsible for the activation of cytokines such as interferon (IFN) which activate antiviral proteins that inhibit the replication of the virus while the adaptive immune response to HCV neutralizes viral particles and destroys infected cells [40]. Studies of HCVinfected chimpanzees that eliminate the virus without the specific T-cell immune response suggest that, in some cases, the innate immune response might be sufficient to destroy the infection [41]. The RNA of HCV is recognized by the innate immune response through the Toll-like receptor which responds with the production of IFN-1α and IFN-1β [5,42]. IFN-1 stimulates the nitric oxide synthase enzyme that is expressed in hepatocytes and macrophages as the isoform inducible nitric oxide synthase [43]. Patients with HCV who are treated with IFN present higher levels of inducible nitric oxide synthase, which have been correlated with lower serum levels of alanine aminotransferase [44]. In addition, IFN-1 induces the production of various proteins such as protein kinase (PKR), 2’,5’-oligoadenylate synthetase (OAS) and the Mx protein [44]. These proteins are responsible for the expression of the genes that inhibit the replication of this virus within hepatocytes in an attempt to destroy the infection [7,40]. When IFN bonds with the IFN receptor on the surface of the infected cell, it activates the Janus kinase, which induces phosphorylation of cytoplasmic proteins known as signal transducers and activators of transcription (STATs), specifically STAT 1 and STAT 2. The STATs form a dimer that directs itself to the cell nucleus where it forms a complex with the p48 protein, which is a stimulation factor for IFN-stimulated gene factor 3. That complex bonds with the IFN-stimulated response element ISRE, which is an RNA-polymerase promoter complex, and there is a stimulus of the genes responsible for the production of antiviral response proteins and of major histocompatibility complex (MHC) proteins [44]. 15 Some individuals present genetic alterations in the STATs or in the Janus kinase that would impede the formation of antiviral proteins [45]. Various viral proteins have shown a capacity to escape the effect of IFN, as evidenced by the high rate of resistance to treatment with IFN-α seen among individuals with hepatitis C [44,45]. There are various characteristics of HCV that allow it to evade the innate immune response: • The viral replication complex appears to be composed of a membrane that is highly resistant to in vitro proteases and nucleases, which protects HCV from detection by the innate immune response [40]. • The HCV core protein interacts with diverse cell factors, including the tumor necrosis factor (TNF) receptor, which decreases the cytolytic activity of T cells [46] • Core proteins impede the antiviral activity of IFN, as do NS3/4A and NS5A proteins [40]. • The NS3/4A proteins can impede the recognition of the Toll-like receptor [5]. • The NS5A and E2 proteins can bind to PKR, thereby blocking its activity [5]. • Multiple mutations in the IFN-sensitivity-determining region (ISDR) modify the NS5A region, which inhibits the phosphorylation of PKR, thereby impeding its antiviral activity [45]. • The E2 region of HCV contains a sequence of eight amino acids identical to those of PKR, and this sequence is more common in genotype 1 than in genotypes 2 and 3, which probably accounts for the fact that individuals infected with genotype 1 present greater resistance to treatment with IFN [44]. • Mutations in the ISDR sequence of NS5A suppress the antiviral action of OAS. • Levels of this protein are lower in nonresponders to treatment with IFN [44]. The liver cell populations that participate in the innate immune response are the natural killer (NK) cells, NK T cells, Kupffer cells and dendritic cells [5]. The NK cells respond minutes or hours after HCV infection by polarizing of the granules in the direction of the infected cells as well as by releasing perforins that fragment the nuclei of infected cells and induce apoptosis [47]. They inhibit viral replication with the production of IFN gamma (IFN-γ), which recruits intrahepatic inflammatory cells and stimulates the T-helper 1 (Th1) response [48], thereby inducing the necrosis or apoptosis of the HCVinfected cell [49]. Studies suggest that HCV inhibits receptor genes in the activation of NK cells, decreasing the activity of these cells by reducing their number and function in chronically infected individuals [40]. The NK cells also have the capacity to increase the functions of dendritic cells in the presence of hepatic cells, although that capacity is impaired in NK cells derived from patients with chronic hepatitis C, in which the production of interleukin (IL)-10 and transforming growth factor beta (TGF-β) can inhibit the activity of dendritic cells [50]. www.bjid.com.br 16 Pathogenesis of Hepatitis C After HCV enters the host cell, the binding of the E2 glycoprotein with the CD81 receptor of NK cells inhibits the function of the NK cells [7,51], which alters the immune response to HCV infection. The E2 glycoprotein also inhibits cytotoxicity and the production of IFNγ by NK cells [40]. Various studies have suggested that the failure of dendritic cells to recognize HCV contributes to the persistence of hepatitis C [50,52-54]. Humoral Immune Response to HCV After HCV infection, there is expression of the hypervariable NS1/E2 region on the surface of the virus, which stimulates B cells to produce high antibody titers of antibodies with the objective of destroying the permanence of the virus [44]. The appearance of anti-HCV antibodies is significantly delayed, and these antibodies can first be detected from 7 to 31 weeks after infection [7]. The host applies selective pressure on HCV, and this stimulates high nucleotide variation, as well as the appearance of mutations in the envelope proteins, from which the virus selects genomic variants in an attempt to eliminate the site of immune response recognition [55]. The great quantity of HCV quasispecies formed allows the virus to evade the humoral immune response, and the effect of HCVneutralizing antibodies appears to be insufficient to control the infection [7], which therefore persists [15]. Similar to what occurs in auto-immune type 2 hepatitis, HCV can mimic the immune system, leading to viral escape or postinfection immunity [4,56]. Anti-HCV antibodies have been implicated in tissue damage due to the formation of immunocomplexes such as antinuclear antibodies [57], autoantibodies that act against cytochrome P450 and antibodies that act against the liver and kidney [4]. The deposition of immunocomplexes has been related to the appearance of extrahepatic manifestations, such as arthritis, cryoglobulinemia [58], vasculitis, glomerulonephritis, Sicca syndrome and itchiness, all of which cause considerable morbidity [57]. There is evidence that HCV infection can be resolved by the cell response with specific CD4+ and CD8+ T cells when there is no formation of antibodies against this virus [59,60], showing that the humoral immune response is not always involved in the response to HCV infection. Cell Response to HCV Since there is a weak humoral immune response to HCV, it is believed that the reactivity of cytotoxic T-lymphocytes (CTLs) or CD8+ T cells is fundamental to viral elimination [61,62], and that impairment of this reactivity is one of the factors responsible for the chronicity of the infection [7,63,64]. The CD8+ T cells can eliminate HCV from the liver through two mechanisms: inducement of apoptosis in infected hepatocytes; and suppression of replication by the production of IFN-γ [22,65]. The CTL response is less vigorous in chronically infected patients than in those presenting acute infection [4]. This can be the result of BJID 2007; 11 Supplement 1 (October) immunologic tolerance or exhaustion of the CD8+ T cell response to the high viral load that persists in individuals chronically infected with HCV [4]. In addition to CD8+ T cells, CD4+ T cells seem to be involved in the viral damage mediated by the increased expression of MHC class II molecules. Some studies have attributed the vigorous and long-lasting response of CD4+ T cells to the elimination of HCV in the acute form the infection [4,66]. However, the loss of the specific CD4+ T cell reactivity to HCV has been associated with the persistence of the virus and the progression of liver damage [67,68]. In acute HCV infection, the peak in serum levels of transaminases corresponds with the cell response, which suggests that the hepatic lesion is immune-mediated [5,64]. It is known that, after activation, T cells initiate clonal proliferation by secreting cytokines and other substances that can affect hepatic function in a variety of ways [69]. Various cytokines act as mediators in the inflammation caused by chronic hepatitis C and have been related to hepatocyte death, i.e. cholestasis and fibrosis, and paradoxically play a role in regeneration following hepatic injury [69,70]. It is argued that the imbalance between the production of Th1 and Th2 cytokines is related to the progression of chronic hepatitis C. The expression of Th1 cytokines such as IL-2 and TNF-α has been shown to be related to the more aggressive presentation of hepatic disease, whereas the expression of Th2 cytokines such as IL-10 has been shown to be related to the milder presentation [71]. The production of TNF-α is one of the earliest events in hepatic injury and is the ‘trigger’ for the production of other cytokines [72], as well as being implicated in the inducement of hepatocyte apoptosis in viral hepatitis [73]. The levels of cytokines such as IFN-γ, TNF-α, IL-6 and IL-8 are elevated in individuals with chronic hepatitis C [7482], and some authors have shown that this increase is proportional to the extent of the damage, histologically [26,76,83,84]. There is evidence that IL-4 can modulate the immune response in HCV-infected individuals [75], principally through the activity of Th2 cells. It has been shown that IL-10 can suppress proliferation in the Th1 and Th2 responses, as well as inducing anergy [85]. There is evidence that IL-10 levels increase in chronic hepatitis C [75]. Some studies report reduced inflammatory activity [86], and others report that administration of IL-10 to such patients causes fibrosis [87]. Various studies have shown that TGF-β is increased in chronic hepatitis C and is involved in the progression of fibrosis, which has been challenged by other authors [88,89]. It has been suggested that TGF-β and IL-10 act as immunosuppressive agents in the liver [90]. In addition, both have been shown to inhibit the immune response and regulate the activity of dendritic cells [91], which can establish a balance between the Th1 and Th2 responses in chronic diseases [92]. www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Pathogenesis of Hepatitis C Host Factors Associated with the Persistence and Progression of Hepatitis C In HCV infection, the genetic constitution and immune ‘status’ of the host are important factors in the persistence and progression of the virus [23,93], since they influence antigen recognition and presentation, as well as the type of Th response [94]. Some MHC class II alleles, such as DR5, have been associated with a lower incidence of cirrhosis in individuals chronically infected with HCV [94]. Rehermann et al. [95] identified CTLs restricted by histocompatibility leukocyte antigen A2 in 97% of chronic hepatitis C patients, compared with 2% of anti-HCV-negative controls. It is speculated that the MHC class II molecule presentation of antigens is deficient in HCV-infected cells, since some viral proteins inhibit the presentation of the antigen through IFN-induced negative immunoregulation [44]. Some pro-inflammatory cytokines appear to be associated with the viral infection response as well as with the expression of specific haplotypes [94], such as IL-10 haplotypes, which can be predictors of spontaneous elimination of HCV [96]. However, there is disagreement in the literature, since other authors did not find evidence for polymorphism in the studied genes being considered as a relevant factor in the elimination of HCV or in the response to treatment [97-99]. The influence of demographic data such as age, gender [6,34] and ‘race’ [100,101] in the progression of hepatitis C can be due to genetic variations existent among those. Some studies report that HCV positivity increases with age [34,101], thereby leading to a greater chance of progression of the disease [6,34]. The male gender is more prevalent in most studies on hepatitis C [6,101] and, in addition, it was associated with the progression of the disease to cirrhosis [6,34]. Some studies suggest that Afro-Americans, due to a greater propensity to chronicity, resistance to treatment (higher percentage of genotype 1) and development of hepatocarcinoma, present a worse evolution of hepatitis C than do Caucasian-Americans [100,102]. Analyzing 99 chronic HCV-infected individuals and 31 individuals who had spontaneously eliminated HCV, Sugimoto et al. [103] found evidence that the CD4+ T-cell response was less vigorous in Afro-Americans than in Caucasian-Americans, with a predominance of the Th2 response and maintenance of the infection. The evolution of hepatitis C in different ethnicities could be due to genetic factors, such as the presence of HLA class II alleles, which could define the spontaneous elimination of HCV [104]. There are various extrinsic host factors that are related to the progression of chronic hepatitis C: alcohol abuse; smoking [6,34,105-109]; the endovenous acquisition of HCV; and coinfection with other viruses such as HIV, HBV and human T-cell lymphotropic virus [6,110,111]. The prevalence of HCV infection is higher among individuals who consume alcohol [112,113]. Studies suggest that alcohol increases the ability of HCV to enter and persist 17 within the organism [112]. Other studies argue that alcohol intake affects some components of the immune response [112] and can alter the inflammatory response of cytokines, thereby increasing viremia, which can be an important cofactor in the development of hepatocarcinoma [114]. In addition, alcohol intake in HCV-infected individuals increases hepatic steatosis and induces apoptosis [107,112,115]. Smoking, in addition to increasing inflammatory activity and hepatic fibroses [109], can induce direct injury to the liver, as well as causing indirect damage (toxic effect), and can have immunological effects (production of IL-1, IL-6 and TNF-a, which cause liver damage). References 1. Alter M.J. Epidemiology of hepatitis C virus infection. World J Gastroenterol 2007;13(17):2436-41. 2. EASL International Consensus Conference on Hepatitis C: Paris 26-28, February 1999. Consensus Statement. J Hepatology 1999;30:956-61. 3. Brown P.M.J., Neuman M.G. Immunopathogenesis of hepatitis C viral infection: Th1/Th2 responses and the role of cytokines. Clin Biochemistry 2001;34:167-71. 4. Cerny A., Chisari F.V. Pathogenesis of chronic hepatitis C: Immunological features of hepatic injury and viral persistence. Hepatology 1999;30:595-601. 5. Dustin L.B., Rice C.M. Flying under the radar: the immunobiology of hepatitis C. Annu Rev Immunol 2007;25:71-99. 6. Poynard T., et al. Natural history of liver fibrosis progression in patients with chronic hepatitis C. The OBSVIRC, METAVIR, CLINIVIR, and DOSVIRC groups. Lancet 1997;394:825-32. 7. Pawlotsky J.M. Pathophysiology of hepatitis C infection and related liver disease. Trends Microbiol 2004;12(2):96-102. 8. Major M.E., et al. Hepatology 1997;25:1527-38. 9. McGarvey M.J., et al. Structure and molecular virology. In: Zuckerman AJ, Toma HC, editors. Viral hepatitis. 2nd ed. London: Churchill Livingstone; 1998. p.253-70. 10. Koziel M.J., et al. Intrahepatic cytotoxic T Lymphocytes specific for hepatitis C virus in persons with chronic hepatitis. J Immunol 1992;149(10):3339-44. 11. Gowans E.J. Distribution of markers of hepatitis C virus infection throughout the body. Semm Liver Dis 2000;20:85-102. 12. MacDonald A.J., et al. CD4 T helper type 1 and regulatory T cells induced against the same epitopes on the core protein in hepatitis C virus-infected persons. J Infect Dis 2002;185:720-7. 13. Ferri C., et al. Hepatitis C vírus infection and B-cell lymphomas. Eur J Can 1994;30:1591-2. 14. Maggi F., et al. Differences in hepatitis C virus quasispecies composition between liver, peripheral blood mononuclear cells and plasma. J Gen Virol 1997;78(Pt 7):1521-5. 15. Giannini C., Bréchot C. Hepatitis C virus biology. Cell Death and Differentiation 2003;(10):S27-38. 16. Pileri P., et al. Binding of hepatitis C virus to CD81. Science 1998;282:938-41. 17. Polyak S.J. Hepatitis C virus-cell interactions and their role in pathogenesis. Clin Liver Dis 2003; 7:67-88. 18. Lindenbach B.D., Rice C.M. Unravelling hepatitis C virus replication from genome to function. Nature 2005;436(18):933-8. 19. Simmonds P., et al. Classification of hepatitis C virus into six major genotypes and a series of subtypes by phylogenetic analysis of the NS-5 region. J General Virology 1993;74:2391-9. 20. Simmonds P., et al. Consensus Proposals for a unified system of nomenclature of hepatitis C virus genotypes. Hepatology 2005;42(4):963-73. www.bjid.com.br 18 Pathogenesis of Hepatitis C 21. Wang X.H., et al. Progression of fibrosis during chronic hepatitis C is associated with rapid virus evolution. J Virol 2007;81(12):6513-22. 22. Herzer K., et al. Hepatitis viruses: live and let die. Liver Int 2007;27(3):293-301. 23. Gremion C., Cerny A. Hepatitis C virus and the immune system: a concise review. Rev Med Virol 2005;15(4):235-68. 24. Haruna Y., et al. Detection of hepatitis C virus RNA in liver tissues by an in situ hybridization technique. J Hepatol 1993;18(1):96-100. 25. Brillanti S., et al. Persistent hepatitis C viraemia without liver disease. Lancet 1993;341(8843):464-5. 26. Miller R.H., Purcell R.H. Hepatitis C virus shares amino acid sequence similarity with pestiviruses and flaviviruses as well as members of two plant virus supergroups. Proc Natl Acad Sci USA 1990;87:2057-61. 27. Gonzalez-Peralta R.P., et al. Optimization for the detection of hepatitis C virus antigens in the liver. J Hepatol 1994;20:143-47. 28. Barone A.A., et al. Response in patients with chronic HCV hepatitis to treatment with interferon-alpha. Braz J Infect Dis 1999;3(3):118-28. 29. Naoumov N.V. Hepatitis C virus-specific CD4+ T cells: do they help or damage? Gastroenterology 1999;117:1012-4. 30. Delladetsima J.K., et al. Cholestatic syndrome with bile duct damage and loss in renal transplant recipients with HCV infection. Liver 2001;21(2):81-8. 31. Dusheiko G., et al. Hepatitis C virus genotypes: an investigation of typo-specific differences in geographic origin and disease. Hepatology 1994;19:13-8. 32. Kobayashi M., et al. The natural course of chronic hepatitis C: a comparison between patients with genotypes 1 and 2 hepatitis C viruses. Hepatology 1996;23(4):695-9. 33. Amoroso P., et al. Correlation between virus genotype and chronicity rate in acute hepatitis C. J Hepatology 1998;28:936-44. 34. Bellentani S., et al. Clinical course and risk factors of hepatitis C virus related liver disease in the general population: report from the Dionysos study. Gut 1999;44(6):874-80. 35. Cholet et al. Factors associated with liver steatosis and fibrosis in chronic hepatitis C patients. Gastroenterol Clin Biol 2004;28:272-8. 36. Rubbia-Brandt L., et al. Steatosis affects chronic hepatitis C progression in a genotype specific way. Gut 2004;53(3):406-12. 37. Sharma P., et al. Hepatic steatosis in hepatitis C virus genotype 3 infection: does it correlate with body mass index, fibrosis, and HCV risk factors? Dig Dis Sci 2004;49(1):25-9. 38. Kumar D., et al. Hepatitis C virus genotype 3 is cytopathic to hepatocytes: reversal of hepatic steatosis after sustained therapeutic response. Hepatology 2002;36:1466-72. 39. Negro F. Mechanisms and significance of liver steatosis in hepatitis C virus infection. World J Gastroenterol 2006;12(42):6756-65. 40. Thimme R., et al. A target on the move: innate and adaptive immune escape strategies of hepatitis C virus. Antiviral Res 2006;69(3):129-41. 41. Lloyd A.R., et al. Host and viral factors in the immunopathogenesis of primary hepatitis C virus infection. Immunol Cell Biol 2007;85(1):24-32. 42. Mizukoshi E., Rehermann B. Immune responses and immunity in hepatitis C virus infection. J Gastroenterol 2001;36(12):799-808. 43. Samuel C.E. Antiviral actions of interferons. Clin Microbil Rev 2001;14(4):778-809. 44. Taylor D.R., et al. Hepatitis C virus and interferon resistance. Microbes and Infection 2000;2:1743-56. 45. Tan S.-L., Katze M.G. How hepatitis C virus counteracts the interferon response: the jury is still out on NS5A. Virology 2001;284:1-12. 46. Chen C.M., et al. Direct interaction of hepatitis C virus core protein with the cellular lymphotoxin-beta receptor modulates the signal pathway of the lymphotoxin-beta receptor. J Virol 1997;71(12):9417-26. BJID 2007; 11 Supplement 1 (October) 47. Rehermann B., Chisari F.V. Cell mediated immune response to the hepatitis C virus. Curr Top Microbiol Immunol 2000;242:299-325. 48. Janeway C.A., et al. Imunobiologia: o sistema imune na saúde e na doença. Tradução Ana Cristina Arámburu da Silva, Cristina Bonorino, Denise Cantarelli Machado, Gaby Renard, Isabel Cristina Ribas Werlang, Moisés Evandro Bauer. 6ª ed. Porto Alegre: Artmed, 2007. Cap. 1, p.37-101: Imunidade Inata. 49. Winnock M., et al. Liver-associated lymphocytes: role in tumor defense. Semin Liver Dis 1993;13:81-92. 50. Jinushi M., et al. Negative regulation of NK cell activities by inhibitory receptor CD94/NKG2A leads to altered NK cell-induced modulation of dendritic cell functions in chronic hepatitis C virus infection. J Immunol 2004;15;173(10):6072-81. 51. Crotta S., et al. Inhibition of natural killer cells through engagement of CD81 by the major hepatitis C virus envelope protein. J Exp Med 2002;195(1):35-41. 52. Longman R.S., et al. Presence of functional dendritic cells in patients chronically infected with hepatitis C virus. Blood 2004;103(3):1026-9. 53. Bain C., et al. Impaired Assostimulatory function of dendritic cells in chronic hepatitis C infection. Gastroenterlol 2001;120(2):512-24. 54. Sarobe P., et al. Abnormal priming of CD4+ T cells by dendritic cells expressing hepatitis C virus core and E1 proteins. J Virol 2002;76(10):5062-70. 55. Botarelli P., et al. T-Lymphocyte response to hepatitis C virus in different clinical courses of infection. Gastroenterology 1993;104:580-7. 56. Cerny A., Chisari F.V. Hepatitis C and Autoimmune Hepatitis. Hepatology 2000;31(3):811-2. 57. Cacoub P., et al (Multivirc Group). Extrahepatic manifestations of chronic hepatitis C. Arthritis Rheum 1999;42:2204-12. 58. Agnello V. Mixed cryoglobulinemia and hepatitis C virus. Hosp Pract 1995;30(3):35-42. 59. Takaki A., et al. Cellular immune responses persist and humoral responses decrease two decades after recovery from a singlesource outbreak of hepatitis C. Nat Med 2000; 6(5):578-82. 60. Lechner et al. Analysis of successful immune responses in persons infected with hepatitis C virus. J Exp Med 2000;191(9):14991512. 61. Missale G., et al. Different clinical behaviors of acute hepatitis C virus infection associated with different vigor of the anti-viral cell-mediated immune response. J Clin Invest 1996;98:706-14. 62. Gruner N.H., et al. Association of hepatitis C virus-specific CD8+ T cells with viral clearance in acute hepatitis C. J Infect Dis 2000;181(5):1528-36. 63. Chisari F.V. Cytotoxic T cells and viral hepatitis. J Clin Invest 1997;100(12):S19-S24. 64. Thimme R., et al. Determinants of viral clearance and persistence during acute hepatitis C virus infection. J Exp Med 2001;194(10):1395-406. 65. Kanto T., Hayashi N. Immunopathogenesis of hepatitis C virus infection: multifaceted strategies subverting innate and adaptive immunity. Intern Med 2006;45(4):183-91. 66. Wertheimer A.M., et al. Novel CD4+ and CD8+ T-cell determinants within the NS3 protein in subjects with spontaneously resolved HCV infection. Hepatology 2003;37(3):577-89. 67. Carucci P., et al. Hepatitis C virus-specific T helper cell responses in recurrent hepatitis C after liver transplantation. J Hepatol 1997;26(suppl):145. 68. Gerlach J.T., et al. Recurrence of hepatitis C virus after loss of virus-specific CD4+ T-cell response in acute hepatitis C. Gastroenterology 1999;117:933-41. 69. Kerr J.F.R., et al. The nature of piecemeal necrosis in chronic active hepatitis. Lancet 1979;20:827-8. 70. Marcellin P., et al. Fibrosis and disease progression in hepatitis C. Hepatology 2002;36:S47-56. www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Pathogenesis of Hepatitis C 7 1 . Napoli J., et al. Progressive liver injury in chronic hepatitis C infection correlates with increased intrahepatic expression of Th1-associated cytokines. Hepatology 1996 ;24:759-65. 72. Tilg H., et al. How to modulate inflammatory cytokines in liver diseases. Liver Int 2006;26(9):1029-39. 73. Ding W.X., Yin X.M. Dissection of the multiple mechanisms of TNF-alpha-induced apoptosis in liver injury. J Cell Mol Med 2004;8(4):445-54. 74. Tilg H., et al. Serun levels of cytokines in chronic liver diseases. Gastroenterology 1992;103:264-74. 75. Cacciarelli T.V., et al. Immunoregulatory cytokines in chronic hepatitis C virus infection: pre- and posttreatment with interferon alfa. Hepatology 1996;24:6-9. 76. Malaguarnera M., et al. Serum interleukin 6 concentrations in chronic hepatitis C patients before and after interferon-alpha treatment. Int J Clin Pharmacol Ther 1997;35(9):385-8. 77. Oyanagi Y., et al. Enhanced expression of interleukin-6 in chronic hepatitis C. Liver 1999;19(6):464-72. 78. Biró L., et al. Changes in the acute phase complement component and IL-6 levels inpatients with chronic hepatitis C receiving interferon alfa-2b. Immunol Lett 2000;72:69-74. 79. Polyak S.J., et al. Elevated levels of interleukin-8 in serum are associated with hepatitis C virus infection and resistance to interferon therapy. J Virol 2001;75(13):6209-11. 80. Lapinski T.W. The levels of IL-1 beta, IL-4 and IL-6 in the serum and the liver tissue of chronic HCV-infected patients. Arch Immunol Ther Exp. (Warsz) 2001;49(4):311-6. 81. Kasprzak et al. Expression of cytokines (TNF-α, IL-1 α, and IL2) in chronic hepatitis C: comparative hybridocytochemical and immunocytochemical study in children and adult patients. J Histochemistry and Cytochemistry 2004;52(1):29-38. 82. Kamal S.M., et al. Kinetics of Intrahepatic hepatitis C virus (HCV)specific CD4+ T cell responser in HCV and Schistosoma mansoni coinfection: relation to progression of liver fibrosis. J Infect Dis 2004;189:1140-50. 83. Shimoda K., et al. Interleukin-8 and hIRH (SDF1-alpha/PBSF) mRNA expression and histological activity index in patients with chronic hepatitis C. Hepatology 1998;28(1):108-15. 84. Neuman M.G., et al. Kinetics of serum cytokines reflect changes in the severity of chronic hepatitis C presenting minimal fibrosis. J Viral Hepat 2002;9:134-40. 85. Taylor A., et al. Mechanisms of immune suppression by interleukin10 and transforming growth factor-beta: the role of T regulatory cells. Immunology 2006;117(4):433-42. 86. Nelson D.R., et al. Long-term interleukin 10 therapy in chronic hepatitis C patients has a proviral and anti-inflammatory effect. Hepatology 2003;38(4):859-68. 87. Nelson D.R., et al. Interleukin 10 treatment reduces fibrosis in patients with chronic hepatitis C: a pilot trial of inferferon nonresponders. Gastroenterology 2000;118:655-60. 88. Roulot D., et al. Quantitative analysis of transforming growth factor beta 1 messenger RNAin the liver of patients with chronic hepatitis C. Absence of correlation between high levels and severity of disease. Hepatology 1995;21:298-304. 89. Nelson D.R., et al. Transforming growth factor-beta 1 in chronic hepatitis C. J Viral Hepat 1997;4(1):29-35. 90. Knolle P.A., Gerken G. Local control of the immune response in the liver. Immunol Rev 2000;174:21-34. 91. Racanelli V., Rehermann B. The liver as an immunological organ. Hepatology 2006;43:S54-62. 92. Yazdanbakhsh M., et al. Allergy, parasites, and the hygiene hypothesis. Science 2002;296(5567):490-4. 93. Chitturi S., George J. Predictors of liver-related complications in patients with chronic hepatitis C. Ann Med 2000;32:588-91. 94. Thursz M.R., et al. Host factors in chronic viral hepatitis. Semin Liver Dis 1997;17:345-50. 95. Rehermann B., et al. Quantitative analysis of the peripheral blood cytotoxic T lymphocyte response in patients with chronic hepatitis C virus infection. J Clin Invest 1996;98:1432-40. 96. Mangia A., et al. IL-10 haplotypes as possible predictors of spontaneous clearance of HCV infection. Cytokine 2004;25(3):103-9. 97. Bozkaya H., et al. Circulating IL-2 and IL-10 in chronic active hepatitis C with respect to the response to IFN treatment. Infection 2000;28:309-13. 98. Constantini P.K., et al. Interleukin-1, interleukin-10 and tumour necrosis factor-alpha gene polymorphisms in hepatitis C virus infection: an investigation of the relationships with spontaneous viral clearance and response to alpha-interferon therapy. Liver 2002;22(5):404-12. 99. Minton E.J., et al; Trent Hepatitis C Study Group. Clearance of hepatitis C virus is not associated with single nucleotide polymorphisms in the IL-1, -6, or -10 genes. Hum Immunol 2005;66(2):127-32. 100. Reddy K.R., et al. Racial differences in responses to therapy with interferon in chronic hepatitis C. Consensus Interferon Study Group. Hepatology 1999;30:787-93. 101. Lepe R., et al. Ethnic differences in the presentation of chronic hepatitis C. J Viral Hepat 2006;13(2):116-20. 102. El-Serag H.B. Epidemiology of hepatocellular carcinoma. Clin Liver Dis 2001;5:87-107. 103. Sugimoto K., et al. Influence of ethnicity in the outcome of hepatitis C virus infection and cellular immune response. Hepatology 2003;37:590-9. 104. Azocar J., et al. MHC class II genes in HCV viral clearance of hepatitis C infected Hispanic patients. Hum Immunol 2003;64(1):99-102. 105. Pessione F., et al. Cigarette smoking and hepatic lesions in patients with chronic hepatitis C. Hepatology 2001;34:121-5. 106. Campollo R O. Hepatitis C virus infection and alcohol. Rev Gastroenterol Mex 2002;67 Suppl 2:S80-3. 107. Vento S., Cainelli F. Does hepatitis C virus cause severe liver disease only in people who drink alcohol? Lancet Infectious Dis 2002;2(5):303-9. 108. Monto A., et al. Risks of a range of alcohol intake on hepatitis crelated fibrosis. Hepatology 2004;39:826-34. 109. El-Zayadi A.R. Heavy smoking and liver. World J Gastroenterol 2006;12(38):6098-101. 110. Kishihara Y., et al. Human T lymphotropic virus type I infection influences hepatitis C virus clearance. J Infect Dis 2001;184:1114-9. 111. Einav S., Koziel M.J. Immunopathogenesis of hepatitis C virus in the immunosuppressed host. Transpl Infect Dis 2002;4(2):85-92. 112. Campollo R.O. Hepatitis C virus infection and alcohol. Rev Gastroenterol Mex 2002;67(Suppl 2):S80-3. 113. Sanchez Avila J.F. Hepatitis C and addictions. Rev Gastroenterol Mex 2002;67(Suppl2):S84-7. 114. Schiff E. The alcoholic patient with hepatitis C virus infection. Am J Med 1999;107(6B):95S-9. 115. Bhattacharya R., Shuhart M.C. Hepatitis C and alcohol: interactions, outcomes, and implications. J Clin Gastroenterol 2003;36(3):242-52. www.bjid.com.br 19 20 BJID 2007; 11 Supplement 1 (October) Antifibrotic Therapy in Chronic Hepatitis C Rinaldo Focaccia Siciliano and Antonio Alci Barone Laboratory of Hepatitis, LIM 47 DMIP-HC-FMUSP; São Paulo, SP, Brazil The most pronounced histological characteristics of chronic hepatitis C are hepatocellular necrosis, inflammatory infiltrate and fibrosis. Unlike the first two, which present fluctuations, the evolution of liver fibrosis progressively evolves toward pathological sequelae due to the chronic liver injury induced by hepatitis C virus (HCV). Fibrosis results from the accumulation of extracellular matrix components, which leads to distortion of the liver architecture, alterations in hepatic microcirculation, and cellular dysfunction. This pathological hepatic process develops slowly and progressively, producing clinical repercussions only in its final stage, liver cirrhosis, which can take decades. Therefore, determination of the degree of fibrosis through liver biopsy or progression rate is crucial to the understanding of the natural history of chronic hepatitis C. Some authors have identified host factors that are associated with the accelerated progression of liver fibrosis in HCV infection. The most important are being over 40 years of age at the time of HCV infection, being male, consuming excessive quantities of alcohol, and presenting conditions that lead to immunodeficiency, such as HIV infection or transplant. Other factors also seem to contribute to a more rapid progression toward fibrosis, such as liver steatosis, coinfection with hepatitis B virus (HBV), obesity, and diabetes mellitus. Recent advances in clinical studies and basic science have brought new perspectives to the development of therapies that can curb the progression of the fibrogenic process or even promote reversion of liver fibrosis. These advances occurred after the recognition of fibrogenic cell types in the liver, as well as of the principal sites of fibrosis formation and its reversibility potential. Fibrosis seems to result from an imbalance between the synthesis and degradation of the extracellular matrix, resulting in the accumulation of conjunctive tissue in the liver. This process is triggered and sustained by the chronic liver damage caused by HCV and leads to the disruption of the normal liver architecture, culminating in the development of cirrhosis/ hepatic insufficiency. Stellate cells are the principal source of extracellular matrix in the damaged liver tissue. In a normal liver, they are present in the Disse space and are the principal reservoirs of vitamin A. Chronic hepatic injury secondary to HCV, through the increase of free radicals and fibrogenic mediators, leads to the activation of stellate cells, as well as to their proliferation and differentiation into myofibroblasts, when they acquire contractile, pro-inflammatory and fibrogenic properties. Once activated, they migrate to the sites of injury The Brazilian Journal of Infectious Diseases 2007;11 (5) Suppl. 1:20-21. © 2007 by The Brazilian Journal of Infectious Diseases and Contexto Publishing. All rights reserved. with the objective of effecting repair by secreting a large quantity of extracellular matrix. Activated stellate cells also produce cytokines that perpetuate their activation status, as well as being stimulated by the apoptosis of damaged hepatocytes. The leukocytes attracted into the hepatic inflammatory process secondary to HCV infection also induce the production of collagen by stellate cells. These activated cells, in turn, have a pro-inflammatory effect that feeds a vicious cycle. Under physiological conditions, the excess extracellular matrix is degraded through regulation by metalloproteinases. Activated hepatic myofibroblasts, in addition to producing large quantities of Type I and Type III collagen, secrete tissue inhibitors of metalloproteinases, which block the collagenolytic activity. We have begun to comprehend various parts of this process of accumulation of extracellular matrix in the liver tissue, although there are still many gaps in our understanding. In addition to the efficient collagenolytic activity of the metalloproteinases, the degradation of the extracellular matrix in the liver can also occur through the activities of neutrophils, macrophages and stellate cells themselves; however, the importance and modulation of each have yet to be clarified. The activation of stellate cells, as well as their proliferation and fibrogenic activity, is regulated by various soluble growth factors, such as platelet-derived growth factor, transformation growth factor beta, and endothelin-1. Determining the mechanisms involved in the process of liver fibrosis has led to a new perspective on the development of antifibrotic drugs in animal models. However, there is still an obstacle to be transposed before positive laboratory results can be reproduced in humans. New lines of research attempt to attenuate the activation of stellate cells, inhibit some of their properties when activated, promote their apoptosis, or stimulate the degradation of the extracellular matrix. No specific antifibrotic therapies have been approved for use in humans; however, considerable effort has been made in laboratory studies in this promising area. In clinical practice, the treatment of hepatitis C with the combination of pegylated interferon alpha and ribavirin is the only pharmacological regimen currently available that can modify the natural evolution of liver fibrosis. This does not simply occur by suppressing the viremia (achieving a sustained virological response) but possibly by the antifibrotic role of pegylated interferon alpha. Recent clinical studies show remission of liver fibrosis or reduction of its progression rate in patients under treatment with therapeutic regimens containing interferon alpha or pegylated interferon. This effect is even more apparent among hepatitis C patients presenting a biochemical or virological response after therapy. Notably, www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Antifibrotics and Chronic Hepatitis C most of these studies are retrospective, and their principal objective was the evaluation of the virologic response to the treatment with interferon alpha. Although liver fibrosis is a dynamic process, its remission is slow and demands prolonged therapy and follow-up evaluation. Multicentric prospective comparative studies that evaluate the histological impact of maintenance treatment with pegylated interferon in low doses versus placebo in patients with hepatitis C who are nonresponsive to treatment are currently being carried out. Preliminary results of these studies are quite promising; reduction of fibrosis or clinical complications resulting from liver cirrhosis can be seen after approximately two years of therapy with pegylated interferon. The final results, with histological analysis, after long-term follow-up treatment and inclusion of a large number of patients, will consolidate an evaluation of safety, clinical and histological benefits, and cost-effectiveness ratio of the prolonged use of pegylated interferon in low doses in hepatitis C. The role of the physician in the positive modification of the natural history of hepatitis C-related liver fibrosis should not be restricted to the removal of HCV as a hepatic aggressive/ pro-inflammatory factor (achieving a sustained virologic response) or to the use of interferon as a potential antifibrotic agent. In daily practice, other strategies aimed at attenuating the progression of liver fibrosis are based on the effects on modifiable factors that can alter the natural history of HCV: 21 drinking cessation; prevention and control of obesity; and prevention of HIV or HBV co-infection. Much progress has been achieved in basic science regarding the understanding of the biological mechanisms that lead to the development of liver fibrosis. However, clinical trials to validate new drugs or antifibrotic strategies are warranted. References 1. Albanis E, Friedman S,L. Antifibrotic agents for liver disease. Am J Transplant 2006;6(1):12-9. 2. Friedman S.L., Rockey D.C., Bissell D.M. Hepatic fibrosis 2006: report of the Third AASLD Single Topic Conference. Hepatology 2007;45(1):242-9. 3. Bataller R., Brenner D.A. Liver fibrosis. J Clin Invest 2005;115(2):209-18. 4. Marcellin P., Asselah T., Boyer N. Fibrosis and disease progression in hepatitis C. Hepatology 2002;36(5 Suppl 1):S47-56. 5. Friedman S.L., Bansal M.B. Reversal of hepatic fibrosis - fact or fantasy? Hepatology 2006;43(2 Suppl 1):S82-8. 6. Everson G.T., Hoefs J.C., Seeff L.B., et al. Impact of disease severity on outcome of antiviral therapy for chronic hepatitis C: Lessons from the HALT-C trial. Hepatology 2006;44(6):1675-84. 7. Kaiser P., Hass H., Lutze B., et al. Long-term low dose treatment with pegylated interferon alpha 2b leads to a significant reduction in fibrosis and inflammatory score in chronic hepatitis C nonresponder patients with fibrosis or cirrhosis. 57th Annual Meeting of the American Association for the Study of Liver Diseases. Massachusetts – USA 2006. 8. Afdhal N., Freilich B., Levine R., et al. Colchicine versus peginterferon long-term (COPILOT) trial: interim analysis of clinical outcomes at year 2. Hepatology 2004;40:238A. www.bjid.com.br 22 BJID 2007; 11 Supplement 1 (October) Laboratory Testing for Hepatitis C Neiva Sellan Lopes Gonçales and Fernando Lopes Gonçales Junior Hepatitis Study Group – MI/FCM/UNICAMP; Campinas, SP, Brazil Serological Detection of Hepatitis C Virus Serological diagnosis of patients infected with the hepatitis C virus (HCV) can be performed using two categories of tests: indirect tests, which detect antibodies against HCV; and direct tests, which detect, quantify, or characterize components of the viral particle, such as HCV RNA testing and testing for detection of the HCV core antigen. Anti-HCV antibodies are usually detected using third- and fourth-generation immunoenzymatic assays – enzyme immunoassay (EIA)/enzyme-linked immunosorbent assay (ELISA) 3 and EIA/ELISA 4, respectively – which contain HCV core antigens and HCV nonstructural genes. The specificity of the EIA tests available on the market that detect anti-HCV was determined to be higher than 99%, whereas their sensitivity, which was more difficult to determine due to the lack of gold standard tests with high sensitivity, was 9599% [1]. However, false-positive results for anti-HCV can occasionally occur, especially in populations with prevalence rates below 10% [2-4]. There are many reasons why laboratories do not routinely use a supplementary test based on immunoblot analysis, such as the recombinant immunoblot assay, to complement the diagnosis of HCV infection. In addition to the high cost of such a test, the lack of laboratory standards that can evaluate its performance and interpretation, in conjunction with its actual accuracy, is among the principal reasons. Furthermore, this type of test does not distinguish past from present infection, and its use is only indicated for confirmation of EIA results. In contrast, the use of nucleic acid testing (NAT) makes it possible to differentiate between viremic and nonviremic individuals by detection of HCV RNA, allowing the clinician a differentiated approach to anti-HCV-positive individuals. However, there can be situations in which HCV RNA is not detected (negative HCV RNA) and the individual has active infection with HCV. This can occur in individuals in whom anti-HCV antibody titers are high and RNA titers are low [5]. Therefore, HCV RNA might not be detectable in certain individuals in the acute phase of the disease. However, these findings are transient, and chronic infection can develop [6]. In addition, HCV RNA intermittent positivity has been observed in individuals chronically infected with HCV [6-8]. Negativity of HCV RNA results can indicate resolved infection. In 15 to 25% of those anti-HCV positive individuals who acquired the infection after 45 years of age, the infection resolves spontaneously. This percentage increases to 40-45% in those who acquired the HCV infection in childhood or young adulthood [9]. Different tests based on polymerase chain reaction (PCR) have been developed to directly detect the viral particle. One characteristic of real-time PCR is amplification coupled with The Brazilian Journal of Infectious Diseases 2007;11 (5) Suppl. 1:22-24. © 2007 by The Brazilian Journal of Infectious Diseases and Contexto Publishing. All rights reserved. detection, which allows the evaluation of the number of viral genomes at the onset of and throughout the reaction. Qualitative detection of HCV RNA by reverse transcriptase (RT)-PCR is generally accepted as the most sensitive and standardized test to date [10,11]. Nevertheless, there is variability among the results from different laboratories, as evidenced by the use of international panels of proficiency. The accuracy and reliability of the results are directly related to the laboratory procedures adopted in the performance of the tests [12]. The lack of preliminary care in sample collection, in conjunction with the time involved in preparing and separating the samples, can result in incorrect results. It is extremely important that all laboratory procedures comply with Good Laboratory Practice and strictly follow the protocols standardized by the manufacturers of the diagnostic kits and reagents. The gold standard consists of the careful use of NAT, standardized for detection of HCV RNA, together with EIAs (specificity in conjunction with sensitivity). An alternative to aid diagnosis is the use of the ratio between optical density and cut-off value (OD/COV) or the sample/cut-off ratio as an indicator of the true positivity of the test. Studies carried out in Brazil show that, in EIAs, reagents with OD/COV greater than 3 are repeatedly associated with 100% true-positive results (positive predictive value) and present approximately 92% positivity for HCV RNA by RTPCR [13]. In terms of the population studied, the positive predictive value is increased when accompanied by risk factors, high levels of alanine aminotransferase (ALT), or liver disease. In immunocompetent patients, EIAs present excellent reproducibility; however, in hemodialyzed or immunocompromised patients, EIA sensitivity is significantly reduced [14]. In low-risk populations, such as blood donors, or in random population screening, i.e., in populations that do not present risk factors for the acquisition of HCV infection, negative EIA results are sufficient to rule out the presence of HCV. However, false-positive results can occur in these populations. In such cases, a qualitative study of HCV RNA should be performed to confirm the diagnosis. In high-risk populations, when there is clinical suspicion of HCV infection, positive EIA results confirm the exposure to HCV. A qualitative study of HCV RNA should be performed to distinguish individuals with chronic infection from those who have eliminated the HCV spontaneously. In patients with chronic hepatitis of unknown cause and negative anti-HCV EIA results, especially in immunocompromised patients [14], a qualitative study of HCV RNA should be performed. The presence of HCV RNA confirms the diagnosis, although a negative result does not rule out HCV infection. In such cases, it is recommended that a new HCV RNA study be performed six months after the first study. Detection of the HCV core antigen by EIA can be an alternative for early diagnosis of HCV infection. www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Laboratory Testing for Hepatitis C The HCV core antigen ELISA was developed to be used as a serological screening test to detect the HCV core antigen, especially during the immunological window period, when antibodies are not detected. This assay was found to have sensitivity close to that of NAT, with a mean difference in detection of one to two days [15]. Based on this assay, a new assay was developed to detect and quantify HCV core antigen. The modifications made to this new assay, such as the dissociation of immune complexes, which allows the detection of free antigens and core antigen antibodies, and the change in the signal amplification, through the modification of the conjugate, have increased the sensitivity of the test. Studies have demonstrated that this test can reduce the immunological window by 3.3 days in comparison with the previous test (i.e., the HCV core antigen ELISA). This increase in sensitivity has led to a significant (58-day) decrease in the size of the immunological window. The difference between this EIA and PCR was only 0.24 days [16]. This test can be considered a viable alternative to detecting viremia directly when NAT cannot be used for reasons of cost, organization, emergency, or logistic difficulties. Tests that allow simultaneous or combined detection of HCV core antigen and antibody in a single assay are currently available on the market. These tests, known as HCV Ag/Ab combo assays, have high sensitivity and specificity, reducing the duration of the immunological window (during which antibodies are not detected) by up to 12 days [17]. Studies carried out using this assay showed sensitivity close to that of NAT, with a mean difference in detection of 1 to 2 days [18]. The use of NAT in the diagnosis of HCV infection makes it possible to distinguish viremic from nonviremic individuals through the detection of HCV RNA. Therefore, these tests can be considered a plausible future solution in the screening of blood donors, organ transplantation programs, and cases of occupational exposure, in which a rapid and low-cost diagnosis is necessary. In order to standardize the tests, the World Health Organization and the United States National Institute for Biological Standards and Controls have established a standard measure known as the international unit (IU). Assays for qualitative detection of HCV RNA are important tools because they are significantly more sensitive than are most quantitative tests. Qualitative assays are based on the principle of target amplification using either PCR or transcription-mediated amplification. The cut-off value of the lower limit of detection of HCV RNA of these commercial assays is 50 IU/mL and 6 IU/ mL, respectively [19]. The specificity of these essays exceeds 99%. A positive test for HCV RNA confirms active replication of HCV. Clinical and laboratory follow-up with study of HCV RNS should be performed to confirm the absence of active replication of HCV. Once HCV infection is confirmed, performing further qualitative tests for HCV RNA in patients submitted to clinical follow-up evaluation but not receiving treatment has no diagnostic utility. The quantification of HCV RNA can be performed by target amplification using PCR or by signal amplification using branched DNA (bDNA). In these commercial assays, the cutoff value of the lower limit of quantification of HCV RNA ranges from 600 to 615 IU/mL, and the upper linear limit ranges from 850,000 to 7,700,000 IU/mL [20]. The standardization in IU does not represent the actual number of viral particles in the preparation. There are significant variations among commercial assays. The dynamics of each assay should be observed, and appropriate dilutions of the material being analyzed should be performed to ensure the accuracy of the quantification. The ideal assay for HCV RNA should have a lower detection limit of 5 to 50 IU/mL and a linearity curve of 6 to 7 log10. Traditional assays for detection of viral load, such as bDNA and Roche Monitor, present detection limits of 615 IU/ mL and 600 IU/mL respectively [21,22], which are inadequate to define end-of-treatment response or sustained virological response. Real-time PCR assays are a promising tool due to their sensitivity and broad range of linearity. Cobas Taqman 48 HCV assay is a quantitative assay that has a detection limit of 10 to 100 IU/mL, which makes it well suited for use in followup treatment (at the initiation and at week 12) [23]. 23 Acute Infection and Cutting/Piercing Accidents After exposure to HCV, anti-HCV antibodies can be detected by EIA in 50 to 70% of the patients at the onset of symptoms, this percentage increasing to approximately 90% after 3 months. Routinely, HCV RNA can be detected between post-exposure weeks 1 and 3, remaining at detectable levels when symptom onset occurs. From post-infection week 2 to post-infection week 8, levels of ALT rise, and this increase is accompanied by the appearance of hepatocytic lesions. Vertical Transmission An important question is that of exactly how mother-tochild transmission of the HCV infection is defined. In many children born to mothers with chronic hepatitis C, anti-HCV (IgG) is detectable in the blood. These antibodies are acquired through passive transplacental transfer. These passively acquired antibodies will remain detectable for the first 12 to 15 months of life. Therefore, the criterion to identify mother-tochild transmission of HCV infection is the detection of antiHCV and HCV RNA in the blood of the child after the age of 18 months. Chronic Infection In patients with chronic hepatitis C, the diagnosis of chronicity is based on the detection of anti-HCV and HCV RNA in the blood, using techniques of high sensitivity, and is confirmed through liver biopsy. Loss of anti-HCV and isolated presence of HCV RNA are uncommon in immunocompetent patients with chronic hepatitis C. However, these findings can occur in hemodialyzed patients and in severely immunocompromised patients Follow-Up Treatment Some patients with detectable HCV RNA should be considered for treatment. Genotyping should be performed at the initiation of treatment in order to define treatment duration, since, according to treatment protocols, patients infected with genotype 2 or 3 should be treated for 24 www.bjid.com.br 24 Laboratory Testing for Hepatitis C BJID 2007; 11 Supplement 1 (October) weeks, whereas those infected with genotype 1 should be treated for 48 weeks [24]. A considerable limitation in the evaluation of patients with chronic infection with HCV has been the lack of standardization of the tests for detection of HCV RNA. A significant difference has been observed in the assays used, both in terms of sensitivity (upper limit of detection) and in terms of dynamics. These differences are observed not only among the different assays but also among different laboratories performing a given assay. Therefore, it is important that, throughout the clinical follow-up of a patient receiving specific treatment, the same assays and, if possible, the same laboratory always be used [25-28]. Quantification of HCV RNA should be performed in the pretreatment sample and in the week-12 sample in order to evaluate the predictive value of the treatment response. Since the qualitative study of HCV RNA presents a lower limit of detection of 50 IU/mL, it should be used at week 4 of treatment, as a predictor of sustained virologic response (SVR), then again, to detect the SVR, at the end of treatment and at 6 months after the end of treatment. Therefore, presenting negative PCR results by week 4 of treatment has a high predictive value for achieving an SVR. 13. Gonçales N.S.L., Costa F.F., Vassalo J., Gonçales Jr., F.L. Diagnosis of hepatitis C in Brazilian blood donors using a reverse transcriptase nested polymerase chain reaction: comparison with enzyme immunoassay and recombinant protein immunoblot assay. Rev Inst Med trop S Paulo 2000;42(5):263-7. 14. Lakshmi V., Reddy A.K., Dakshinamurty K.V. Evaluation of commercially available third-generation anti-hepatitis C virus enzyme-linked immunosorbent assay in patients on haemodialysis. Indian J Med Microbiol 2007;25:140-2. 15. Tanaka E.C., Ohue K., Aoyagi K., et al. Evaluation of a new enzyme immunoassay for hepatitis C virus (HCV) core antigen with clinical sensitivity approximating that of genomic amplification of HCV RNA. Hepatology 2000;32:388-93. 16. Laperche S., Le Marrec N., Simon N., et al. A new HCV core antigen assay based on disassociation of immune complexes: an alternative to molecular biology in the diagnosis of early HCV infection. Transfusion 2003;43:958-62. 17. Laperche S., Elghouzzi M.-H., More P., et al. Is an assay for simultaneous detection of hepatitis C virus core antigen and antibody a valuable alternative to nucleic acid testing? Transfusion 2005;45:1965-72. 18. Laperche S., Le Marrec N., Girault A., et al.. Simultaneous detection of hepatitis C virus (HCV) core antigen and anti-HCV antibodies improves the early detection of HCV infection. J Clin Microbiol 2005;43:3877-83. 19. Desombere I., Van Vlierberghe H., Couvert S., et al. Comparison of qualitative (COBAS AMPLICOR HCV 2.0 versus VERSANT HCV RNA) and quantitative (COBAS AMPLICOR HCV monitor 2.0 versus VERSANT HCV RNA 3.0) assays for hepatitis C virus (HCV) RNA detection and quantification: impact on diagnosis and treatment of HCV infections. J Clin Microbiol 2005;43(6):2590-2. 20. Elbeik T, Surtihadi J, Destree M et al. Multicenter evaluation of performance characteristics of the Bayer VERSANT HCV RNA 3.0 assay (bDNA). J Clin Microbiol 2004;42:563-9. 21. Ve i l l o n P. , P a y a n C . , P i c c h i o G. , e t a l . C o m p a r a t i v e evaluation of the total hepatitis C virus core antigen, branched-DNA, and Amplicor Monitor assays in determining viremia for patients with chronic hepatitis C during interferon plus ribarin combination therapy. J Clin Microbiol 2003;41:3212-20. 22. Nolte F.S., Fried M.W., Shiffman M.L., et al. Prospective multicenter clinical evaluation of AMPLICOR and COBAS AMPLICOR hepatitis C virus tests. J Clin Microbiol 2001;39:4005-12. 23. Konnick E.Q., Willians S., Ashwood E.R., et al. Evaluation of Cobas hepatitis C virus (HCV) Taqman anlyte –specific reagent assay nd comparison to the Cobas Amplicor HCV monitor v2.0 and Versant HCV bDNA 3.0 assays. J Clin Microbiol 2005;43:2133-40. 24. Fried M.W., Shiffman M.L., Reddy C., et al. Peginterferon alfa-2a plus ribavirin for chronic hepatitis C virus infection. N Engl J Med 2002;347:975-92. 25. Shiffman M.L., Ferreira-Gonzalez A., Reddy K.R., et al. Comparison of three commercially available assays for HCV RNA using the International Units standard: Implications for management of patients with chronic hepatitis C virus infection in clinical practice. Am J Gastroenterol 2003;98:1159-66. 26. Podzorski R.P. Molecular testing in the diagnosis and management of hepatitis C virus infection. Arch Pathol Lab Med 2002;126:285-90. 27. Schirm J., van Loon A.M., Valentine-Thon E., et al. External quality assessment program for qualitative and quantitative detection of hepatitis C virus in diagnostic virology. J Clin Microbiol 2002;40:2973-80. 28. Pawlotsky J.M. Diagnostic testing in hepatitis C virus infection: viral kinetics and genomics. Semin Liver Dis 2003;23:3-11. References 1. Alter M.J., Kuhnert W.L., Finelli L. Guidelines for Laboratory Testing and Result Reporting of antibody to hepatitis C Virus. MMWR 2003;52:1-15. 2. Kleinman S., Alter H., Bush M., et al. Increased detection of hepatitis C virus (HCV) – infected blood donors by a multiple-antigen HCV enzyme immunoassay. Transfusion 1992;32:805-613. 3. Conry-Cantilena C., VanRaden M., Gibble J., et al. Route of infection, viremia, and liver disease in blood donors found to have hepatitis C virus infection. N Engl J Med 1996;334:1691-6. 4. Hyams K.C., Riddle J., Rubertone M., et al. Prevalence and incidence of hepatitis C virus infection in the US military: a seroepidemiologic survey of 21000 troops. Am J Epidemiol 2001;153:764-70. 5. Busch M.P., Kleinmam S.H., Jackson B., et al. Nucleic acid testing of blood donors for transfusion-transmitted infectious diseases: report of Interorganization Task Force on Nucleic Acid Amplification of Blood Donors. Transfusion 2000;40:143-59. 6. Thomas D.L., Astemborski J., Rai R.M., et al. Natural history of hepatitis C virus infection: host, viral, and environmental factors. JAMA 2000;284:450-6. 7. Alter M.J., Margolis H.S., Krawczynski K., et al. Natural history of community-acquired hepatitis C in the United States. N Engl J Med 1992;327:1899-905. 8. Larghi A., Zuin M., Crosignani A., et al. Outcome of an out break of acute hepatitis C among healthy volunteers participating in pharmacokinetics studies. Hepatology 2002;36:993-1000. 9. Alter H.J., Seef L.B. Recovery, persistence, and sequelae in hepatitis C virus infection: a perspective on long term outcome. Semin Liver Dis 2000;20:17-35. 10. Pawlotsky J.M. Molecular diagnosis of viral hepatitis. Gastroenterology 2002;122:1554-68. 11. Pawlotsky J.M. Use and interpretation of virological tests for hepatitis C. Hepatology 2002;36:S65-S73. 12. Callendo A.M., Valsamakis A., Zhou Y., et al. Multilaboratory comparison of hepatitis C virus viral load assays. J Clin Microbiol 2006;44:1726-32. www.bjid.com.br BJID 2007; 11 Supplement 1 (October) 25 Hepatitis C: Genotyping Norma de Paula Cavalheiro Laboratory of Hepatitis, LIM 47 DMIP-HC-FMUSP; São Paulo, SP, Brazil The heterogeneity of the hepatitis C virus (HCV) genome was described at the beginning of the 1990s. Subsequently, genotyping tests were developed in order to delineate and differentiate these variants, leading to various classifications in different parts of the world. However, in 1994, in a consensus publication, criteria were established and a universal classification was consolidated, comprising the identification of 6 large groups or viral genotypes, in addition to over 70 different subtypes distributed worldwide. It was established that genotypes differ from one another in 31% to 33%, as do the subtypes in 20% to 25%. The standardization in HCV classification was in accordance with the uniformity of data published in scientific and epidemiological studies on evolution and pathogenesis. Due to genomic peculiarities of different populations in the world, databases for centralization and collection of information on HCV circulating genomes were created, by region: the first in Japan by Prof. Masashi Mizokami et al. (http://s2as02.genes.nig.ac.jp/); the second in the European Union by Prof. Gilbert Deleage et al. (http://euhcvdb.fr/); and the third in the United States by Dr. Carla Kuiken et al. (http:/ /hcv.lanl.gov/ or http://hcv-db.org). The accessibility of these databases, which are consulted and updated by researchers worldwide, helps standardize terms for HCV viral genotypes and subtypes [1]. Prior to 1994, new variants were identified in Vietnam, Thailand, Burma and Indonesia. These viral genotypes were originally classified as genotypes 7, 8, 9, 10 and 11. These variants were reclassified in 2005; genotype 10a came to be denominated subtype 3k, and genotypes 7a, 7d, 7b, 7e/7c, 11a, 9a, 9b, 9c, 8b and 8a became subtypes 6e, 6c, 6d, 6f, 6g, 6h, 6i, 6j, 6k and 6l, respectively [1]. In Brazil, HCV genotype 1 is predominant, being identified in 70% of the infected population, followed by genotypes 3 (in 25%) and 2 (in approximately 5%). In the southern region, the profile is differentiated, the prevalence of genotype 3 being comparable to that of genotype 1 [2,3]. The HCV viral genotype can be determined in a clinical sample in different forms, and the regions of HCV genome considered appropriate include the core, E1, NS4 and NS5 regions, as well as the 5’UTR, as mentioned in innumerable studies [4,5]. The most direct method, considered the gold standard, is the sequencing of the HCV genome in a certain region, sufficiently divergent for different genotypes and subtypes to be distinguished [5]. The Brazilian Journal of Infectious Diseases 2007;11 (5) Suppl. 1:25-27. © 2007 by The Brazilian Journal of Infectious Diseases and Contexto Publishing. All rights reserved. In practice, there are two categories of assays for the diagnosis of HCV viral types: serotyping and genotyping [6,7]. The enzyme-linked immunosorbent assay, also known as the indirect method (serotyping), commercialized only for research, discriminates among the six HCV genotypes, and not the subtypes. It presents genotype-specific antibodies for different HCV genotypes (Serotype HCV 5’NC – AbbottMurex) in the NS4 region of the genome. This test omits the determination of the quantity of circulating viral particles and presents lower sensitivity and specificity in the diagnosis of HCV genotypes, when compared with genotyping. Its performance is also limited in the evaluation of immunocompromised patients [3,7,8]. Genotyping (the direct method) analyzes the sequences of various regions of the genome in the differentiation of HCV genotypes and subtypes. Therefore, it depends on samples that present viral particles for analysis. These techniques are: sequencing, hybridization with genotype-specific probes, and real-time polymerase chain reaction (PCR) [5,7,9]. Reverse hybridization, distributed under the commercial name Line Probe Assay (LIPA - Versant HCV Genotype assay 1.0/2.0; Innogenetics, Ghent, Belgium; distributed by Siemens Medical Solutions Diagnostics, Tarrytown, NY, USA), as described by Lieven Stuyver et al. in 1993, uses nitrocellulose tapes, on which oligonucleotide probes of region 5’NC, complementary to each specific HCV genotype/subtype, are immobilized. These tapes are hybridized under high stringency conditions, and the PCR product is marked with biotinylated primers in the amplification process. After hybridization, an avidin conjugate will bind to the biotinylated hybrid. The substrate will yield the formation of a colored product, which will be deposited on the tape and reveal the viral genotype present in the sample. Genotypes 1 through 6, as well as subtypes 1a, 1b, 1a/1b, 2a/2c, 2b, 3a-c, 4a-h, 5a, 6a and 10a, are discriminated. The results are evaluated through visual assessment. A new version of LIPA (version 2.0) adds probes of the core region of HCV and enables distinction of subtypes 1c of 6 and subtypes 1a and 1b, which, in the former version, was not possible for all the samples analyzed, since region 5’NC is extremely conserved and may not present sufficient diversity for the distinction of these subtypes. The interpretation of results is also performed by assessing the nitrocellulose tapes with a scanner, and the images are analyzed using a computer program. The new version is in the registration phase for distribution in Brazil [7,10,11]. Direct sequencing is the most complete information on the variations of the sequences analyzed. In addition, it is an ideal and definitive method for the study of viral genetic variability. Moreover, the study of viral dynamics in the context www.bjid.com.br 26 Hepatitis C: Genotyping of quasispecies, during the natural history of the disease or as consequence of antiviral therapy, depends principally on the direct sequencing of certain regions of the HCV genome. Furthermore, it is considered the gold standard for determining HCV genotypes and subtypes [7]. The sequencing technique for HCV genotyping consists of PCR amplification of part of the viral genome, especially of the 5’NC, NS5B and core regions. These regions are sufficiently conserved for the development of reliable primers and, at the same time, present diversity for the discrimination of viral genotypes and subtypes. The PCR employed for sequencing comes from PCR-HCV qualitative or quantitative PCR HCV products, the volume of DNA and the high purity of the resulting tapes being essential for the success of the analyses. This second PCR is performed specifically to adjust or mark these products for reading in sequencers. The assessment of both tapes, positive and negative, is essential in order to resolve possible ambiguities during the evaluation of the sequencings [7]. The sequences provided by the equipment should be analyzed, and there are international databases available for consultation, for this purpose. These databases are also used for the centralization and collection of sequences worldwide. The database recommended for Brazil is the European site http://euhcvdb.fr/, probably because of the epidemiological pathway followed during the Brazilian colonization period. The genotyping test TRUGENE-SIEMENS HCV 5’NC Genotyping Kit (Siemens Medical Solutions Diagnostics, Tarrytown, NY, USA) offers software used in conjunction with the equipment, with a previously selected genomic library that analyzes the sequences of region 5’NC immediately after sequencing. In addition to the genotyping results, it presents the homology with patterns of HCV genotypes and subtypes based on region 5’NC. This methodology is only available to researchers [1,7,12]. The real time PCR-HCV method is a quantitative molecular diagnostic test that uses TaqMan technology. In this test, genotype-specific probes marked for HCV typing were employed in order to identify genotypes 1, 2 and 3a. Another test, also based on TaqMan technology, can identify genotypes 1 to 4 and presents specific probes for subtypes 1a/b, 2a/b/c, 3a and 4b/c/d. Both methodologies are based on region 5’NC [9,13-16]. Commercially, but only for research, there is the Abbott Real-Time PCR HCV Assay (Abbott Diagnostics Europe, Wiesbaden, Germany) test, which also determines the viral genotype of the samples for HCV genotypes 1a, 1b, 2a, 2b, 3, 4, 5 and 6. This methodology for the genotyping of HCV is associated with viral quantification, levels below 6053 IU/mL compromising the efficiency and sensitivity of the test [9]. The identification of genotypes is clinically important, and treatment protocols recommend that genotype information be obtained in order to delineate the duration and type of the medication to be used. The literature mentions that genotypes BJID 2007; 11 Supplement 1 (October) 1 and 4 are considered more resistant than are genotypes 2 and 3, and that the standard treatment consists of interferon combined with ribavirin. Another disadvantage of genotypes 1 and 4 is that they present a worse prognosis of evolution of the disease. For treatment-naïve patients, infection with genotype 1, 4 or 5 should be treated for 12 months, compared with 6 months for infection with genotype 2 or 3. The Consensus Group recommends pegylated interferon combined with ribavirin for genotype 1. For re-treatment, pegylated interferon associated with ribavirin is recommended, regardless of the genotype, in the usual doses, respecting the duration recommended for genotype 1 for other genotypes. In cases of HIV/HCV co-infection, the response to antiviral therapy for HCV is not favorable: 14%-38% for genotype 1; and 43%-73% for genotypes 2 and 3 [1,7,17,18]. The advantages of genotyping methods include reliability and the opportunity to obtain important information on the molecular pathogenesis of HCV [7]. The impact of HCV heterogeneity and its different genotypes on the everyday clinical management of HCV chronic infection has not been completely established, nor has its role as an epidemiological marker been clarified. The sensitivity and specificity of serological and virological tests can also be influenced by the heterogeneity of HCV, which justifies constant evolution in the study of patients and differentiation techniques of HCV genotypes and subtypes [4]. References 1. Simmonds P., Bukh J., Combet C., et al. Consensus proposals for a unified system of nomenclature of hepatitis C virus genotypes. Hepatology 2005;42:962-73. 2. Campiotto S., Pinho J.R.R., Carrilho F.J., et al. Geographic distribution of hepatitis C virus genotypes in Brazil. Brazilian Journal of Medical and Biological Research 2005;38:41-9. 3. Cavalheiro N.P., Barone A.A., Tengan F.M. HCV Serotypes in brazilian patients. Int J Infect Dis 2002;6:228-32. 4. Pawlotsky J.-M. Diagnostic tests for hepatitis C. J Hepatol 1999;31 Suppl. 1:71-9,. 5. Laperche S., Lunel F., Izopet J., et al. Comparison of Hepatitis C Virus NS5b and 5_ Noncoding Gene Sequencing Methods in a Multicenter Study. Journal of Clinical Microbiology 2005:733-9. 6. Pawlotsky J.-M., Prescott L., Simmonds P., et al. Serological determination of hepatitis C virus genotype: Comparison with a standardized genotyping assay. J Clin Microbiol 1997;35:1734-9. 7. Stéphane C., Pawlotsky J.-M. Hepatitis C virus: Virology, diagnosis and management of antiviral therapy. World J Gastroenterol 2007;13(17):2461-6. 8. Prescott L.E., Simmonds P. Serological genotyping using synthetic peptides derived from the NS4 region. In: LAU, JOHNSON YIU-NAM. Hepatitis C protocols: methods in molecular medicine. Totowa, Humana Press, cap.17, p.199-205, 1998. 9. Raymond H.W., Carol Cimmins. Evaluation of the Abbott Molecular Diagnostics Real Time PCR Assay for HCV Quantitative Viral Load and HCV Genotyping. Poster S30, Clinical Virology Symposium, 2004. 10. Nadarajah R., Khan Y., Miller S.A., Brooks G.F. Evolution of a new generation Line-Probe Assay that detects 5’untranslated and Core regions to genotype and subtype Hepatitis C Virus. Am J Clin Pathol 2007;128:300-4. www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Hepatitis C: Genotyping 11. Bouchardeau F., Cantaloube J.F., Chevaliez S., et al. Improvement of Hepatitis C Virus (HCV) Genotype Determination with the New Version of the INNO-LiPA HCV Assay. Journal of Clinical Microbiology 2007;45:1140-5. 12. Nolte F.S., Green A.M., Fiebelkorn K.R., et al. Clinical evaluation of two methods for genotyping Hepatitis C virus based on analysis of the 5’noncoding region. Journal of Clinical Microbiology 2003;41:1558-64. 13. Lindh M., Hannoun C. Genotyping of hepatitis C virus by Taqman real-time PCR. Journal of Clinical Virology 2005;34:108-14. 14. Moghaddam A., Reinton N., Dalgard O. A rapid real-time PCR assay for determination virus genotypes 1, 2 and 3a. Journal of Viral Hepatitis 2006;13:222-9. 27 15. Rolfe K.J., Alexander G.J.M., Tim G.W., et al. A real-time Taqman method for hepatitis C virus genotyping. Journal of Clinical Virology 2005;34:115-21. 16. Mukaide M., Tanaka Y., Kakuda H., et al. New combination test for hepatitis C virus genotype and viral load determination using Amplicor GT HCV MONITOR test v2.0. World J Gastroenterol 2005;11(4):469-75. 17. Poynard T., Marcellin P., Lee S.S., et al. Randomized trial of interferon alfa 2b plus ribavirin for 48 weeks or for 24 weeks versus interferon alfa 2bplus placebo for 48 weeks for treatment of chronic infectious hepatitis C virus. Lancet 1998;352:1426-32. 18. Tom Wong, Samuel S.L. Hepatitis C: a review for primary care physicians. CMAJ 2006;174(5):649-59. www.bjid.com.br 28 BJID 2007; 11 Supplement 1 (October) Noninvasive Means of Diagnosing Liver Fibrosis in Hepatitis C Eduardo Sellan Lopes Gonçales, Adriana Flávia Feltrim Angerami and Fernando Lopes Gonçales Junior Study Group on Hepatitis, Infectious Diseases Division, UNICAMP; Campinas,SP Liver biopsy is still considered the gold standard for staging fibrosis in chronic liver diseases. However, liver biopsy is an invasive procedure, and complications occur in 0.6%-5% of patients [1,2]. In addition, to perform the procedure there is a need for additional resources such as ultrasonography. Therefore, as a rule, patients undergoing liver biopsy are hospitalized for at least 6 hours [3]. Recent studies involving patients with chronic hepatitis C showed that fragments of technically inadequate hepatic tissue frequently lead to the underestimation of the stage of liver fibrosis [4]. That rate of diagnostic error can vary from 10%-30% depending on the study [5]. In addition, in developed countries, there is greater patient resistance to undergoing biopsy. In Brazil there is an additional factor, which is that patients are obligated to submit to liver biopsy for indication of treatment, except in clinically confirmed cases of hepatic cirrhosis according to the Ministry of Health guidelines. For all of these reasons, an increasing number of studies are being conducted in order to evaluate the effectiveness of noninvasive markers for staging liver fibrosis. The noninvasive methods used in the largest number of published studies are the calculation of two indices - the aspartate aminotransferase (AST) to platelet ratio index (APRI) and the FibroTest index - and the FibroScan test. The effectiveness of the various methods evaluated in various studies revealed quite heterogeneous results. The APRI method and the Forns index are unable to stage a large percentage of patients, and their accuracies do not exceed 80-85%. Therefore, a considerable number of patients are required to undergo liver biopsy. Otherwise, approximately 20% would be incorrectly diagnosed. The efficacy of those methods encounters difficulty regarding standardization and the definition of cut-off values for each degree of fibrosis. cut-off values are used. The absence of cirrhosis (Ishak stage 0-4) is defined as values lower than 1, and cirrhosis (Ishak stage 5-6) is defined as values higher than 2 [6]. The formula for calculating the APRI test is as follows: APRI The APRI was developed by Wai et al. [6] and is calculated based on AST levels and platelet counts. According to the results obtained in that study, the lower and upper cut-off values for the definition of significant fibrosis and cirrhosis are determined. Through analysis of the results, the positive and negative predictive values for the presence or absence of significant fibrosis or cirrhosis are also determined. To evaluate significant fibrosis, the following cut-off values are used: lower than 0.5 (absence of significant fibrosis, Ishak stage 0-2); and higher than 1.5 (presence of significant fibrosis, Ishak stage 3-6)[6]. To evaluate cirrhosis, different FibroScan - A New Noninvasive Method The evaluation of the degree of liver fibrosis is of fundamental importance to the prognosis, follow-up and therapeutic decision-making for patients with chronic liver disease. Biopsy is an invasive method and occasionally (although rarely) results in complications. In addition, the biopsy results, from an anatomical-pathological point of view, are often evaluated subjectively [8,9]. Nevertheless, biopsy continues to be the gold standard by which fibrosis is staged and evaluated There are various studies on noninvasive options in the staging, evaluation and monitoring of liver fibrosis. FibroScan is a new method, still only available on a small scale, which presents better results in various studies with respect to differentiating between cirrhotic and noncirrhotic patients. It The Brazilian Journal of Infectious Diseases 2007;11 (5) Suppl. 1:28-31. © 2007 by The Brazilian Journal of Infectious Diseases and Contexto Publishing. All rights reserved. APRI = AST(/LSN) × 100 / Platelets (109/L) Table 1 shows the APRI values obtained. Table 1. The aspartate aminotransferase to platelet ratio index results obtained by Wai et al. No Fibrosis Fibrosis No Cirrhosis Cirrhosis Cut-Off < 0.5 > 1.5 < 1.0 > 2.0 PPV 64% 91% 35% 65% NPV 90% 65% 100% 95% PPV=positive predictive value; NPV=negative predictive value. Therefore, the aforementioned Wai et al. showed that the APRI has a high positive predictive value to identify patients with significant fibrosis and a high negative predictive value to rule out cirrhosis. That study also showed that it is possible to predict the presence or absence of significant fibrosis in 51% of patients and to predict the presence or absence of cirrhosis in 81% of patients [6]. FibroTest FibroTest combines and analyzes the serum levels of five factors in patients with chronic hepatitis C. Those five factors are bilirubin, gamma-glutamyl transferase, apolipoprotein A1, alpha-2-macroglobulin and haptoglobin. The results obtained are evaluated through a formula which predicts and classifies them as F0-1, F2-3 and F4 [7]. www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Noninvasive Diagnosis of Fibrosis in Hepatitis C 29 Figure 1. Algorithm proposed by Sebastiani et al. (EASL) [14], in which the aspartate aminotransferase to platelet ratio index (APRI) method is used in conjunction with FibroTest. ALT=alanine aminotransferase; UDE=upper digestive tract endoscopy; US=ultrasound. Chronic hepatitis C with elevated ALT levels APRI F0-F1 F≥ ≥2 Unclassified FibroTest F≥ ≥2 F0-F1 Biopsy ≥F2) Significant fibrosis (≥ F0-F1 is considered a quick and easy noninvasive procedure for diagnosing cirrhosis and has been presented as an alternative to liver biopsy in patients with a formal contraindication [8]. FibroScan measures hepatic elasticity through a transducer positioned intercostally on the skin over the right lobe of the liver. The transducer transmits low amplitude and low frequency vibration pulses to the hepatic tissue. This vibration pulses propagate an elastic wave whose velocity is directly related to the elasticity of the tissue. Results are given in kilopascals (kPa). Some studies have shown that body mass index and age of the patient, as well as the level of experience on the part of the health professional, can influence the FibroScan results of patients with chronic hepatitis C. The body mass index and steatosis can affect the evaluation of fibrosis, although some studies have shown that those factors are minimized if the test is repeated a fair number of times. Some studies recommend a total of five measurements to validate the results [10]. For the detection of fibrosis ≥ F2, FibroScan presents 85.2% sensitivity, 90.7% specificity, 93.8% positive predictive value, 78.8% negative predictive value and 87.7% diagnostic power. For the detection of cirrhosis, the test presents 78.3% sensitivity, 98.2% specificity, 97.8% positive predictive value, 81.6% negative predictive value and 88.2% diagnostic power [11-13]. Despite the reasonable quantity of published studies, few have compared the methods in a randomized manner. There are two studies that propose algorithms for evaluation. The first study, published by the European Association for the www.bjid.com.br 30 Noninvasive Diagnosis of Fibrosis in Hepatitis C BJID 2007; 11 Supplement 1 (October) Figure 2. Algorithm proposed by Castéra et al. [15], in which the FibroTest results are evaluated together with the FibroScan results. Detectable HCV RNA FibroScan/ FibroTest Concordance between the two methods No Yes Liver biopsy No liver biopsy Treatment or monitoring Fibrosis absent or minimal (<F2) Moderate fibrosis (F2) Severe fibrosis or ≥F3) cirrhosis (≥ Monitoring Treatment Treatment + UDE and US every 6 months Study of the Liver (EASL), aims to standardize and compare the various methods based on serum markers of liver fibrosis. In its conclusion, the study proposes the use of an organogram that could reduce the need for liver biopsy by 60%-70% [14]. That study, conducted by Sebastiani et al. [14], evaluated the capacity of different methods to diagnose significant fibrosis (METAVIR fibrosis score ≥ 2) in patients with normal or high levels of transaminase and to diagnose cirrhosis. The results obtained show that significant fibrosis can be diagnosed with an accuracy of 94% using the APRI as the first screening test, followed by FibroTest in patients who were not classified through the APRI method, thereby limiting biopsy to only those patients in whom the degree of fibrosis is classified as F0-F1 using noninvasive methods [14]. Cirrhosis can also be diagnosed through this algorithm (95% accuracy). The authors considered that in their original study they might have obtained highly favorable results due www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Noninvasive Diagnosis of Fibrosis in Hepatitis C The study conducted by Castéra et al. proposes the combination of FibroScan and FibroTest as a screening method for significant fibrosis. When there was concordance between the methods, which occurred in 70%-80% of patients, the compatibility with the liver biopsy was 84% in diagnosing fibrosis ≥ F2, 95% in diagnosing fibrosis ≥ F3, and a 94% in diagnosing cirrhosis (F4). Therefore, much still needs to be studied in relation to the noninvasive methods for estimating the degree of fibrosis, which is why it is necessary to carry out randomized comparative studies involving different patient populations. Table 2. FibroScan results Variable Sensitivity Specificity PPV NPV ≥F2 67% 89% 95% 48% Degree of fibrosis ≥F3 73% 91% 87% 81% F4 87% 91% 77% 95% PPV=positive predictive value; NPV=negative predictive value. to the fact that the majority of patients presented significant fibrosis. The principal limitation of these markers is the difficulty in obtaining confirmation for patients with F0-F1 fibrosis. Therefore, an algorithm for the evaluation of fibrosis that attempts to define which patients are not required to undergo liver biopsy was developed. Another study conducted by Castéra et al. [15] compared the effectiveness of transitory hepatic elastography (FibroScan, Echosens, Paris, France) in relation to the APRI and FibroTest. It succeeded in showing that FibroScan has a great capacity to diagnose significant fibrosis (≥ F2). The results can be seen in Table 2. Therefore, it can be seen that FibroScan presents a high positive predictive value for patients with fibrosis ≥ F2 and an excellent negative predictive value for patients with hepatic cirrhosis. It is undoubtedly a good method for demonstrating significant fibrosis or for ruling out hepatic cirrhosis. 31 References 1. Perrault J., McGill D.B., Ott B.J., Taylor W.F. Liver biopsy: complications in 1000 inpatients. Gastroenterology 1978;74:103-6. 2. Gunneson T.J., Menon K.V., Wiesner R.H., et al. Ultrasoundassisted percutaneous liver biopsy perfomed by a physician assistant. Am J Gastroenterol 2002;97:1472-5. 3. Wong J.B., Koff R.S. Watchful waiting with periodic liver biopsy versus immediate empirical therapy for histologically mild chronic hepatitis C. A cost-effectiveness analysis. Ann Intern Med 2000;133:665-75. 4. Colloredo G., Guido M., Sonzogni A., Leandro G. Impact of liver biopsy size on histological evaluation of chronic viral hepatitis: the smaller the sample, the milder the disease. J Hepatol 2003;39:239-44. 5. Poniachik J., Bernstein D.E., Reddy K.R., et al. The role of laparoscopy in the diagnosis of cirrhosis. Gastrointest Endosc 1996;43:568-71. 6. Wai C.T., Greenson J.K., Fontana R.J., et al. A simple noninvasive índex can predict both significant fibrosis and cirrhosis in patients with chronic hepatitis C. Hepatology 2003;38:518-26. 7. Poynard T., McHutchison J., Manns M., et al. Biochemical surrogate markers of liver fibrosis and activity in a randomized trial of peginterferon alfa-2b and ribavirin. Hepatology 2003;38:481-92. 8. Transient elastography: a new surrogate marker of liver fibrosis influenced by major changes of transaminases. Journal of Viral Hepatitis 2007;14:360-9. 9. Diagnosis of hepatic steatosis and fibrosis by transient elastography in asymptomatic healthy individuals: a prospective study of living related potencial liver donors. Journal of Gastroenterol 2007;42:382-8. 10. Usefulneness of elastometry in evaluating the extents of liver fibrosis in hemophiliacs coinfected with hepatitis C and human immunodeficiency virus. Hepatology research 2006;35:135-9. 11. Features associated with success rate and performance of fibroscan measurements for the diagnosis of cirrhosis in HCV patients: a prospective study of 935 patients. Journal of Hepatology 2007;46:628-34. 12. Transient elastography: a valid alternative to biopsy in patients with chronic liver disease. Journal compilation 2006;24:513-8. 13. Do not trivialize the Fibroscan examination, value its accuracy. Journal of Hepatology 2007;46. 14. Sebastiani G., Vario A., Guido M., et al. Stepwise combination algorithms of non-invasive markers to diagnose significant fibrosis in chronic hepatitis C. Journal of Hepatology 2006;44:686-93. 15. Castéra L., Vergniol J., Foucher J., et al. Gastroenterology 2005;128:343-50. www.bjid.com.br 32 BJID 2007; 11 Supplement 1 (October) Chronic Hepatitis C: Pathological Anatomy Evandro Sobroza de Mello and Venâncio Avancini Ferreira Alves Pathological Anatomy Division, Hospital das Clínicas of São Paulo; LIM-14: Hepatic Pathology, University of São Paulo School of Medicine; CICAP – Hospital Alemão Oswaldo Cruz; São Paulo, SP, Brazil In infections with the hepatitis C virus (HCV), there is a wide spectrum of histological alterations that can affect the liver, from acute hepatitis to mild reactive phenomena to more severe forms, including chronic hepatitis with varying degrees of inflammation/fibrosis, cirrhosis, and hepatocellular carcinoma. In cases of acute hepatitis C, biopsies are rare, pathologists focusing their attention on the chronic form of the disease. The histological diagnosis of chronic hepatitis through liver biopsy remains extremely important in the management of patients infected with HCV, since it is the cornerstone of the detection of liver disease caused by the virus as well as the determination of the intensity of this disease. It should therefore be added to the diagnosis of infection made using serologic methods. The basic parameter for the histological diagnosis of chronic hepatitis is the presence of portal inflammatory infiltrate, with predominance of lymphocytes, usually with variations in the number of plasmocytes and histiocytes. This inflammation is accompanied by periportal activity of varying degrees (also denominated interface activity or piecemeal necrosis), parenchymal activity (lobular) and fibrosis. There are various classification systems using in the scoring and staging of chronic hepatitis [4,9,11,13,18,21,34]. Many of those systems are of historical importance. According to directive no. 863, issued by the São Paulo State Secretary of Health on November 4, 2002, it is recommended that one of two chronic hepatitis classification systems be used: the Sociedade Brasileira de Patologia (SBP, Brazilian Society of Pathology) system [13] or the METAVIR system [1,4]. These two systems are in fact very similar, and they both take into account the previously mentioned basic aspects of chronic hepatitis: periportal activity, lobular activity, and fibrosis. In addition to these, the classification system proposed by Ishak in 1995 [18] has been widely used in international literature. The Ishak system is an update of the system proposed by the same author in 1981, which gained popularity and was commonly referred to as the Knodell system [21] (a designation that should no longer be used), has been widely used in international literature. Table 1 shows an approximate correspondence between these systems, both for fibrosis (architectural alteration) and for periportal/lobular activity. Protocol of Histological Evaluation for Liver Biopsies of Patients with Chronic viral Hepatitis This protocol can be applied to several etiologies of chronic hepatitis, including, in addition to HCV, HBV, autoThe Brazilian Journal of Infectious Diseases 2007;11 (5) Suppl. 1:32-36. © 2007 by The Brazilian Journal of Infectious Diseases and Contexto Publishing. All rights reserved. immune hepatitis and, less frequently, Wilson’s disease or some forms of drug-induced hepatitis. The protocol is based on the criteria of the SBP National Consensus of Chronic Hepatitis [13]. 1) Sample type (needle biopsy, wedge biopsy, resected surgical sample, other): 2) Sample size Number of portal spaces in the biopsy: _____ 3) Histological variables: • Portal fibrosis: ( ) 0 (absent) ( ) 1 (discrete, without septum formation) ( ) 2 (with portal-portal septa) ( ) 3 (with portal-portal and portal-central septa, with formation of nodules – in ‘nodular transformation’) ( ) 4 (cirrhosis) • Portal inflammation ( ) 0 (absent) ( ) 1 (discrete) ( ) 2 (moderate) ( ) 3 (pronounced) ( ) 4 (very pronounced) • Periportal activity (interface activity) ( ) 0 (absent) ( ) 1 (presence of spillover only) ( ) 2 (discrete piecemeal necrosis – occasional foci in some portal spaces) ( ) 3 (moderate piecemeal necrosis – occasional foci in many portal spaces or innumerable foci in few portal spaces) ( ) 4 (pronounced piecemeal necrosis - innumerable foci in many portal spaces) • Parenchymal activity ( ) 0 (absent) ( ) 1 (tumefaction, lymphocyte sinusoidal infiltrate and occasional foci of lytic hepatocytic necrosis) ( ) 2 (innumerable foci of lytic hepatocytic necrosis) ( ) 3 (occasional areas of confluent necrosis) ( ) 4 (innumerable areas of confluent necrosis or areas of panacinar necrosis) • Histological evidence of association with other conditions: ( ) level ______ siderosis ( ) steatohepatitis markers ( ) others:______ Nature and Size of the Liver Biopsy Surgical biopsies performed with forceps generate subcapsular samples and should be discouraged, since the portal spaces in this location are frequently large, and it is www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Chronic Hepatitis C – Pathological Anatomy 33 Table 1. Approximate equivalence of the most widely used classification systems for the staging and scoring of chronic hepatitis Architectural Alteration (Fibrosis)* SBP, 2000 METAVIR, 1994 Ishak, 1995 0 1 2 3 4 0 1 2 3 4 0 1 or 2 3 4 or 5 6 Inflammatory Activity** SBP, 2000 and Ishak, 1995 Parenchymal activity 0 or 1 0 or 1 2 2 2 3 3 4 METAVIR, 1994 Periportal activity A 0 1 or 2 0–1 2 3–4 0–2 3–4 0–4 0 1 1 2 3 2 3 3 SBP= Sociedade Brasileira de Patologia (SBP, Brazilian Society of Pathology). *Maximum Ishak score, 6; maximum METAVIR score, 4; maximum SBP score, 4. **Corresponds to periportal and parenchymal activity, independently, for SBP and Ishak, and mixed periportal and lobular for METAVIR; in the METAVIR classification, the activity score reaches 3, whereas in Ishak and SBP it reaches 4. difficult or impossible to correctly evaluate the presence of fibrosis. Even during the surgical procedure, therefore, liver biopsy should be performed with a needle. In addition, the biopsy should preferably be performed at the outset of the surgery in order to prevent alterations secondary to surgical manipulation. Data in the literature demonstrate that the size of the needle biopsy greatly influences the result of the analysis [8,10,15,33,35]. Samples measuring 3.0 cm or more in length, show hepatitis with mild activity, as a result, in only 50% of the cases; 1.5-cm long samples, in 60%; and those measuring 1.0 cm or less, in almost 90% of the cases [8]. Other authors have also considered 1.5 cm as the minimum size for diagnosis in needle liver biopsy [35]. Thin needles have also provided inferior results [8,33]. Bedossa et al. [3] only achieved a precision plateau with 2.5-cm long biopsies. Therefore, 1.5cm long biopsies should be considered the minimum necessary size and, ideally, they should measure 2.5 cm or more. Larger diameter needles, such as Tru-Cut needles, are also recommended. Steatosis Approximately 50% of the biopsy samples collected from patients with HCV present steatosis [17,40]. The evaluation of the presence of steatosis, its scoring, and the evaluation of the presence of associated steatohepatitis has gradually become more important [6,7,12,16,19,20,24,29-32,36,40]. The spectrum of steatosis, steatohepatitis and cirrhosis has been denominated nonalcoholic fatty liver disease (NAFLD). Although NAFLD is common in the population in general, concomitance between NAFLD and HCV is 2-3 times greater than what would be expected only at random [24]. In patients with chronic HCV infection, steatosis has been attributed to a series of factors usually associated with NAFLD, including high body mass index, insulin resistance and old age [16,26,31]. Evidence also indicates that steatosis contributes to the progression of fibrosis in a pattern similar to that seen in NAFLD [7,16,17,40]. It has been suggested that steatosis can also result from the viral cytopathic effect, especially in patients infected with genotype 3. In a series of patients with genotype 3 and steatosis, a sustained virological response led to regression of steatosis in 91% of the cases, a much higher index than the 19% observed for those who did not present sustained virological response [6], making the cytopathic effect a more consistent cause of steatosis. Other authors have reported similar results [22,36]. In HCV-positive patients, it is currently essential to characterize steatosis and related injuries, especially the presence and quantification of perisinusoidal and centrilobular fibrosis, which characterizes steatohepatitis. The lesson we learn from steatosis is that, in HCV-infected patients, biopsy is an instrument for the detection of liver diseases, whether associated with the virus or not, and that we should be prepared for other (probably less common) liver diseases that might be present in a particular case. Histopathological Aspects of Post-Transplant HCV Recurrence Immediate post-transplant virological recurrence of HCV is universal, and the progression of the disease is more rapid www.bjid.com.br 34 Chronic Hepatitis C – Pathological Anatomy than in non-transplanted patients [22A]. Long-term studies have shown that 70-90% of cases will present histological recurrence of the disease [20A,23A]. In most cases, hepatitis C histologically manifests in the same manner before the transplant: with portal and parenchymal inflammation, aggression of the interface with piecemeal necrosis, and fibrosis at portal spaces. Steatosis and ductal injury are also common findings. Earlier findings are often predominantly lobular, with inflammation and apoptotic bodies (acute hepatitis), and steatosis is occasionally the first histological manifestation [4A]. In a small proportion of cases, hepatitis C can result in a severe, rapidly progressive cholestatic pattern, which leads to loss of the graft [36A,41,42]. Due to the low tolerance of transplanted individuals to the treatment with interferon and ribavirin [24A,45], the evaluation of the biopsy is crucial for indication of treatment. Some anatomopathological aspects can be useful in predicting its evolution: histological recurrence in less than six months [46]; level of inflammatory activity [23A,44]; marked ballooning of hepatocytes; and cholestasis [43]. Histopathological Criteria for Possible Predictive Value of Worse Evolution In chronic hepatitis, the contribution of the histopathological analysis of the samples collected by liver biopsy is currently considered decisive for diagnosis, for staging of the architectural damage, and for determining the level of necroinflammatory activity, assuming a decisive role in indicating the therapy with antiviral agents. In our view, in addition to reports on that decision, as summarized in the METAVIR, Ishak, Scheuer, and Desmet classification systems, or, among us, the SBP/Brazilian Society of Hepatology consensus led by Gayotto, most recent evidence brings back the need of a detailed report of each of the principal forms of liver damage, and there have been studies that demonstrate a more rapid evolution of cases that present, among other predictive factors, more interface activity, confluent necrosis of hepatocytes, and steatosis [47,48]. A study involving 106 patients with initial biopsy presenting architectural staging 0 or 1 and re-biopsied after a mean interval of 7.8 years (minimum of 48 months) [48] revealed progression of architectural damage in 64 cases (60.4%), suggesting the need for therapeutic intervention, even in infected individuals not yet presenting significant alterations to the hepatic architecture. Among the predictive factors for progression of the injury, those authors highlight the level of necroinflammatory activity: 31.2% of the cases with moderate activity (A2) presented progression, which only occurred in 2.3% of those without activity (A0) and the presence of steatosis (progression in 87.5% of the cases with > 30% cells with steatosis, in 80% of those with < 30% steatosis, and in only 48.6% without steatosis). Other authors also emphasize the presence and extent of steatosis as a risk factor for progression of injuries in chronic hepatitis C, either resulting from viral cytopathic effect, as proposed for genotype 3a [49], or associated with coexistent BJID 2007; 11 Supplement 1 (October) steatohepatitis, alcoholic [50] or nonalcoholic [47]. A recent meta-analysis including 3068 Italian patients infected with HCV and submitted to biopsy [51] confirmed that steatosis is independently associated with genotype 3, fibrosis, diabetes, inflammatory liver activity, drinking, body mass index, and older age. Important experience was brought to debate in the most recent European Hepatology Congress: analyzing predictive factors of damage progression in 563 cases of HCV with mean intervals between biopsies of 5.4 years [52], it was determined that, in contrast to generic approaches that suggested that liver damage progress in a relatively uniform, linear manner, the speed of progression varied considerably in each patient. These authors, selecting statistically significant variables, identified the risk of progression associated with various architectural alterations (Table 2).We find, therefore, that important current studies demonstrate the outstanding contribution of histopathological findings in the discrimination of differentiated progression risk in patients chronically infected with HCV. More than dividing patients in classes that deserve antiviral treatment or not, most recent evidence point to the need of reviewing the systems of histological scoring, and the pathologist should inform, in addition to the stage of architectural alteration, the level of each type of necroinflammatory damage in each acinar compartment of the liver. Table 2. Progression risks of architectural alterations Factor Age > 50 years Brindging necrosis Confluent necrosis Piecemeal necrosis Steatosis Moderate/Pronounced Discrete Perivenular fibrosis Relative Risk 1.6 4.0 3.4 2.9 3.8 2.1 2.6 References 1. The French METAVIR Cooperative Study Group. Intraobserver and interobserver variations in liver biopsy interpretation in patients with chronic hepatitis C. Hepatology 1994;20:15-20. 2. Bain V.G., Bonacini M., Govindarajan S., et al. A multicentre study of the usefulness of liver biopsy in hepatitis C. J Viral Hepat 2004;11:375-82. 3. Bedossa P., Dargere D., Paradis V. Sampling variability of liver fibrosis in chronic hepatitis C. Hepatology 2003;38:1449-57. 4. Bedossa P., Poynard T. An algorithm for the grading of activity in chronic hepatitis C. The METAVIR Cooperative Study Group. Hepatology 1996;24:289-93. 4a. Baiocchi L., Tisone G., Palmieri G., et al. Hepatic steatosis: a specific sign of hepatitis C reinfection after liver transplantation. Liver Transpl Surg 1998;4:441-7. 5. Berg T., Sarrazin C., Hinrichsen H., et al. Does noninvasive staging of fibrosis challenge liver biopsy as a gold standard in chronic hepatitis C? Hepatology 2004;39:1456-7; author reply 14571458. www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Chronic Hepatitis C – Pathological Anatomy 6. Castera L., Hezode C., Roudot-Thoraval F., et al. Effect of antiviral treatment on evolution of liver steatosis in patients with chronic hepatitis C: indirect evidence of a role of hepatitis C virus genotype 3 in steatosis. Gut 2004 ;53:420-4. 7. Cholet F., Nousbaum J.B., Richecoeur M., et al. Factors associated with liver steatosis and fibrosis in chronic hepatitis C patients. Gastroenterol Clin Biol 2004;28:272-8. 8. Colloredo G., Guido M., Sonzogni A., Leandro G. Impact of liver biopsy size on histological evaluation of chronic viral hepatitis: the smaller the sample, the milder the disease. J Hepatol 2003;39:239-44. 9. De Groote J., Desmet V.J., Gedigk P., et al. A classification of chronic hepatitis. Lancet 1968;2:626-8. 10. Demetris A.J., Ruppert K. Pathologist’s perspective on liver needle biopsy size? J Hepatol 2003;39:275-7. 11. Desmet V.J., Gerber M., Hoofnagle J.H., et al. Classification of chronic hepatitis: diagnosis, grading and staging. Hepatology 1994;19:1513-20. 12. Fiore G., Fera G., Napoli N., et al. Liver steatosis and chronic hepatitis C: a spurious association? Eur J Gastroenterol Hepatol 1996;8:125-9. 13. Gayotto L.C.C., Alves V.A., Cerski C.T., et al. Visão Histórica e consenso nacional sobre a classificação das hepatites crônicas projeto do clube de patologia hepática da Sociedade brasileira de Patologia aprovado pela Sociedade Brasileira de Hepatologia. GED 2000;19:137-40. 14. Giannini E., Testa R. Noninvasive diagnosis of fibrosis: the truth is rarely pure and never simple. Hepatology 2003;38:1312-13; author reply 1313. 15. Holund B., Poulsen H., Schlichting P. Reproducibility of liver biopsy diagnosis in relation to the size of the specimen. Scand J Gastroenterol 1980;15:329-35. 16. Hu K.Q., Kyulo N.L., Esrailian E., et al. Overweight and obesity, hepatic steatosis, and progression of chronic hepatitis C: a retrospective study on a large cohort of patients in the United States. J Hepatol 2004;40:147-54. 17. Hwang S.J., Luo J.C., Chu C.W., et al. Hepatic steatosis in chronic hepatitis C virus infection: prevalence and clinical correlation. J Gastroenterol Hepatol 2001;16:190-5. 18. Ishak K., Baptista A., Bianchi L., et al. Histological grading and staging of chronic hepatitis. J Hepatol 1995;22:696-9. 19. Ismail F.W., Hamid S.S. Hepatic steatosis and hepatitis C. J Pak Med Assoc 2004;54:108-9. 20. Khokhar N., Mushtaq M., Mukhtar A.S., Ilahi F. Steatosis and chronic hepatitis C virus infection. J Pak Med Assoc 2004;54:110-2. 20a. Feray C., Gigou M., Samuel D., et al. The course of hepatitis C virus infection after liver transplantation. Hepatology 1994;20:1137-43. 21. Knodell R.G., Ishak K.G., Black W.C., et al. Formulation and application of a numerical scoring system for assessing histological activity in asymptomatic chronic active hepatitis. Hepatology 1981;1:431-5. 22. Kumar D., Farrell G.C., Fung C., George J. Hepatitis C virus genotype 3 is cytopathic to hepatocytes: Reversal of hepatic steatosis after sustained therapeutic response. Hepatology 2002;36:1266-72. 22a. Gane E. The natural history and outcome of liver transplantation in hepatitis C virus-infected recipients. Liver Transpl 2003;9:S28-34. 23. Lebensztejn D.M., Kaczmarski M., Sobaniec-Lotowska M., Barwijuk-Machala M. Blind liver biopsy in children-diagnostic significance and complications in authors’ own material. Med Sci Monit 2000;6:1155-8. 23a. Gane E.J., Portmann B.C., Naoumov N.V., et al. Long-term outcome of hepatitis C infection after liver transplantation. N Engl J Med 1996;334:815-20. 35 24. Lonardo A., Adinolfi L.E., Loria P., et al. Steatosis and hepatitis C virus: mechanisms and significance for hepatic and extrahepatic disease. Gastroenterology 2004;126:586-97. 24a. Garcia-Retortillo M., Forns X. Prevention and treatment of hepatitis C virus recurrence after liver transplantation. J Hepatol 2004;41:2-10. 25. Lu L.G., Zeng M.D., Wan M.B., et al. Grading and staging of hepatic fibrosis, and its relationship with noninvasive diagnostic parameters. World J Gastroenterol 2003;9:2574-8. 26. Monto A., Alonzo J., Watson J.J., et al. Steatosis in chronic hepatitis C: relative contributions of obesity, diabetes mellitus, and alcohol. Hepatology 2002;36:729-36. 27. Myers R.P., Benhamou Y., Imbert-Bismut F., et al. Serum biochemical markers accurately predict liver fibrosis in HIV and hepatitis C virus co-infected patients. AIDS 2003;17:721-5. 28. Myers R.P., Tainturier M.H., Ratziu V., et al. Prediction of liver histological lesions with biochemical markers in patients with chronic hepatitis B. J Hepatol 2003;39:222-30. 29. Negro F. Hepatitis C virus and liver steatosis: when fat is not beautiful. J Hepatol 2004;40:533-5. 30. Patton H.M., Patel K., Behling C., et al. The impact of steatosis on disease progression and early and sustained treatment response in chronic hepatitis C patients. J Hepatol 2004;40:484-90. 31. Ramalho F. Hepatitis C virus infection and liver steatosis. Antiviral Res 2003;60:125-7. 32. Rubbia-Brandt L., Fabris P., Paganin S., et al. Steatosis affects chronic hepatitis C progression in a genotype specific way. Gut 2004;53:406-12. 33. Scheuer P.J. Liver biopsy size matters in chronic hepatitis: bigger is better. Hepatology 2003;38:1356-8. 34. Scheuer P.J. The nomenclature of chronic hepatitis: time for a change. J Hepatol 1995;22:112-4. 35. Schlichting P., Holund B., Poulsen H. Liver biopsy in chronic aggressive hepatitis. Diagnostic reproducibility in relation to size of specimen. Scand J Gastroenterol 1983;18:27-32. 36. Sharma P., Balan V., Hernandez J., Rosati M, Williams J, RodriguezLuna H, Schwartz J, Harrison Eet al. Hepatic steatosis in hepatitis C virus genotype 3 infection: does it correlate with body mass index, fibrosis, and HCV risk factors? Dig Dis Sci 2004;49:25-9. 36a. Kaneko J., Sugawara Y., Akamatsu N., et al. Cholestatic hepatitis due to hepatitis C virus after a living donor liver transplantation. Hepatogastroenterology 2004;51:243-4. 37. Terjung B., Lemnitzer I., Dumoulin F.L., et al. Bleeding complications after percutaneous liver biopsy. An analysis of risk factors. Digestion 2003;67:138-45. 38. Wai C.T., Greenson J.K., Fontana R.J., et al. A simple noninvasive index can predict both significant fibrosis and cirrhosis in patients with chronic hepatitis C. Hepatology 2003;38:518-26. 39. Wawrzynowicz-Syczewska M., Kruszewski T., Boron-Kaczmarska A. Complications of percutaneous liver biopsy. Rom J Gastroenterol 2002;11:105-7. 40. Wyatt J., Baker H., Prasad P., et al. Steatosis and fibrosis in patients with chronic hepatitis C. J Clin Pathol 2004;57:402-6. 41. Pessoa M.G., Bzowej N., Berenguer M., et al. Evolution of hepatitis C virus quasispecies in patients with severe cholestatic hepatitis after liver transplantation. Hepatology 1999;30:1513-20. 42. Troppmann C., Rossaro L., Perez R.V., McVicar J.P. Early, rapidly progressive cholestatic hepatitis C reinfection and graft loss after adult living donor liver transplantation. Am J Transplant 2003;3:239-40. 43. Pelletier S.J., Iezzoni J.C., et al. Prediction of liver allograft fibrosis after transplantation for hepatitis C virus: persistent elevation of serum transaminase levels versus necroinflammatory activity. Liver Transpl 2000;6:44-53. 44. Prieto M., Berenguer M., Rayon J.M., et al. High incidence of allograft cirrhosis in hepatitis C virus genotype 1b infection following transplantation: relationship with rejection episodes. Hepatology 1999;29:250-6. www.bjid.com.br 36 Chronic Hepatitis C – Pathological Anatomy 45. Samuel D., Bizollon T., Feray C., et al. Interferon-alpha 2b plus ribavirin in patients with chronic hepatitis C after liver transplantation: a randomized study. Gastroenterology 2003;124:642-50. 46. Testa G., Crippin J.S., Netto G.J., et al. Liver transplantation for hepatitis C: recurrence and disease progression in 300 patients. Liver Transpl 2000;6:553-61. 47. Westin J., Nordlinder H., Lagging M., et al. Steatosis accelerates fibrosis development over time in hepatitis C vírus genotype 3 infected patients. J Hepatol 2002;37:837-42. 48. Boccato S., Pistis R., Noventa F., et al. Fibrosis Progression in initially mild chronic hepatitis C. J Viral Hepat 2006;13:297-302. BJID 2007; 11 Supplement 1 (October) 49. Rubbia-Brandt L., Leandro G., Spahr L., et al. Liver steatosis in chronic hepatitis C: a morphological sign suggesting infection with HCV genotype 3. Histopathology 2001;39:119-24. 50. Pessione F., Degos F., Marcellin P., et al. Effect of alcohol consumption on serum hepatitis C virus RNA and histological lesions in chronic hepatitis C. Hepatology 1998;27:1717-22. 51. Leandro G., Mangia A., Hui J., et al. Relationship between steatosis, inflammation, and fibrosis in chronic hepatitis C: a meta-analysis of individual patient data. Gastroenterology 2006;130:1636-42. 52. Silini E.M., Cavallero A., Dal Bello B., et al. Modelling liver fibrosis progression in chronic hepatitis C: A study of 563 patients with sequential liver biopsies. J. Hepatol 2006;44(S2):S36-81. www.bjid.com.br BJID 2007; 11 Supplement 1 (October) 37 Co-Infection with Hepatitis B Virus and Hepatitis C Virus Heloísa Pedrosa Mitre and João Silva de Mendonça Infectious Diseases Division of the Hospital for State Civil Servants; São Paulo, SP, Brazil The hepatitis B virus (HBV) and the hepatitis C virus (HCV) share common transmission pathways. Therefore, co-infection can be expected. The World Health Organization estimates that, worldwide, 170 million people are infected with HCV, and 350 million people are infected with HBV. However, the number of individuals co-infected with both viruses is unknown. Although various studies have evaluated small numbers of co-infected individuals, the inclusion criteria, parameters assessed, and study designs are not uniform. In addition, ethnicity, local epidemiology, and viral genotypes are also diverse. Therefore, the conclusions of a specific study, in principle, should not be widespread. In cases of co-infection with HBV and HCV, the replication of either virus can be inhibited, just as either virus can be dominant or the dominance can alternate between the two. It is more common for HBV to appear to be suppressed by HCV. The chronologies of the two infections have an influence on which virus will be dominant. Molecular biology techniques (to determine levels of HCV RNA and HBV DNA) have facilitated the definition of the interaction between the two viruses. Co-infections can appear in various manners: a) Simultaneous acute infection with HBV and HCV: presupposes same source and transmission pathway. The number of studies is small, but indicates that the interaction between the two viruses is similar to that which occurs in chronic infections. There are descriptions of cases in which there is a delay in the identification of the Hepatitis B surface antigen (HBsAg), lower levels of alanine aminotransferase (ALT), and lower HBV antigenemia, which can be attributed to suppression of HBV activity by HCV. b) Superinfection by one virus, the other virus being chronically present: it should be suspected above all in individuals with risk factors, such as the use of illicit intravenous drugs, multi-transfused individuals, and those living in areas of high HBV prevalence. Superinfection by HCV: infection by HCV in a patient that is already infected with HBV. This is known in Asian countries, where the prevalence of HBV is high. The viral suppression of the HCV may occur, although the viral suppression of the HBV is more frequent, noted by lower levels of HBV DNA and lower DNA polymerase activity, as well as by the hepatic expression of HBsAg and hepatitis B core antigen (HBcAg), clearance of the hepatitis B e antigen (HBeAg) or even of the HBsAg. The HCV core antigen seems to affect the transcription of HBV and, as a result, its replication, which is reported to be more accentuated for the HCV genotype 1. The Brazilian Journal of Infectious Diseases 2007;11 (5) Suppl. 1:37-39. © 2007 by The Brazilian Journal of Infectious Diseases and Contexto Publishing. All rights reserved. After HBV clearance, HCV can persist, resulting in chronic hepatitis. In addition, there is the possibility of evolution to severe disease, with a risk of death. Superinfection by HBV: infection by HBV in an individual that is already chronically infected with HCV. The HCV RNA levels are lower, and, in a study conducted in Italy, HCV clearance was higher in co-infected individuals (71%) than in mono-infected individuals (14%). However, the HBV DNA levels can be lower than those of mono-infected individuals, indicating HCV interference. Therefore, one virus can induce the clearance of another. The evolution to severe forms of the disease, with fulminant hepatitis profiles, has also been described. c) Asymptomatic infection with HBV: there are reports of patients infected with HCV, with low levels of HBV DNA, reactive anti-HBc, however, non-reactive HBsAg, HBeAg, anti-HBe, and anti-HBs, configuring co-infection with HCV with asymptomatic HBV. These individuals evolve with high ALT levels and high histological activity. There are reports in which biopsies from such patients were evaluated, and cirrhosis was found in 33% of the cases, compared with 19% for patients presenting chronic mono-infections. The data suggest that the evolution of the disease is more severe in co-infected individuals. There are various potential outcomes: a) Clearance of both viruses: negative HBsAg and, eventually, appearance of anti-HBs. The anti-HCV remains reactive. However, both HBV DNA and HCV RNA become undetectable. This is the best evolution for both viruses. b) Fulminant hepatitis: several studies indicate that individuals co-infected with HBV and HCV have a higher risk of evolution to fulminant hepatitis. In a prospective study involving patients with acute hepatitis C, of those who died, 23% were chronically infected with HBV, whereas only 2.9% were infected with HBV (OR=10.2). In a study conducted in France, among 40 patients with fulminant or sub-fulminant hepatitis, 12.5% presented acute hepatitis B or C, and 7.5% presented superinfection with HCV. c) Chronic hepatitis: of all possible events, this is the one that occurs most frequently and presents treatment possibilities. Among the chronic cases of hepatitis, several immunological and molecular-biological profiles can be identified: Active HBV and HCV: detectable HBV DNA and HCV RNA in serum. This situation has higher possibility of evolution to cirrhosis and hepatic decompensation. In these cases, treatment may be considered, either with interferon (IFN) + ribavirin (RBV) or IFN + lamivudine (LMV). www.bjid.com.br 38 HBV/HCV Co-Infection Inactive HBV and active HCV: with evolution to undetectable HBV DNA and detectable HCV RNA. If treatment is indicated, IFN + RBV is the treatment of choice. Active HBV and inactive HCV or previous infection with HCV: with reactive HBsAg and HBeAg, detectable HBV DNA and undetectable HCV RNA. If indicated, therapeutic options may be IFN or IFN + LMV. Cirrhosis: co-infected individuals have a higher risk of evolution to cirrhosis if compared with mono-infected individuals (44% and 21%, respectively) as well as a higher risk of chronic hepatitis decompensation. Hepatocellular carcinoma: evidence indicates that the possibility of evolution to hepatocellular carcinoma (HCC) is higher in co-infected individuals. A prospective study comprising 290 individuals with cirrhosis concluded that, in the univariate and multivariate analyses, co-infection with HBV and HCV is a predictive factor for the development of HCC. These conclusions were confirmed in subsequent studies, in which the incidence of HCC in individuals coinfected with HBV and HCV was 2 and 3.7 cases/100 individuals/year, respectively. The cumulative risk of HCC development in 10 years was 45% in co-infected individuals, 16% in individuals monoinfected with HBV, and 28% in individuals mono-infected with HCV. Treatment The treatment guidelines have been defined in monoinfected individuals, among others, by professional associations of liver researchers, such as the Asian-Pacific Association for the Study of the Liver, the European Association for the Study of the Liver, and the American Association for the Study of Liver Diseases. According to these guidelines, the choice of pegylated interferon over conventional interferon was defined in association with ribavirin in cases of chronic infection with HCV, and, in isolation, in cases of chronic infection with HBV, when the administration of antiviral drugs is also considered. On the other hand, the treatment of individuals co-infected with HBV and HCV is complex due to the interaction between both viruses, and between both viruses and the host immunologic system. Standards for these treatments are not yet available. However, the therapeutic regimen should be carefully chosen and should be based on serologic markers, viremia levels, histology, and, above all, definition of the dominant virus. Individuals with uncompensated cirrhosis should be referred to facilities specializing in liver transplantation. Interferon Since IFN is an immunomodulating drug, with antiviral and antiproliferative effect, it is effective against HBV and HCV. It is also the most widely studied therapeutic option. The first study in co-infected individuals, carried out in the 1990s, suggests that higher doses (9 MU 3 times a week) are more effective for the clearance of HBV or HCV. BJID 2007; 11 Supplement 1 (October) If suppression of one of the viruses is achieved, there is the possibility of reactivation of the other, since the suppressor effect of the former has been removed. Pegylated IFN, which is more effective in mono-infected individuals, can have a similar effect in co-infected individuals. In the infection with HCV and asymptomatic HBV infection, the histological changes and activity are typically pronounced. The response to treatment is less favorable and respondents present recurrence (downregulation?). IFN + RBV: this is the regimen that performs best in individuals mono-infected with HCV. There have been various studies, involving small samples of co-infected individuals, reporting rates of sustained biochemical response and sustained virologic response (SVR) that are comparable to those of mono-infected individuals, especially if HCV is the dominant virus. Due to the viral interaction, it is extremely important that HBV activity is monitored. It is known that at least half of the patients present HBV reactivation at the beginning of the treatment, and that 45% will have ‘flare-ups’. Therefore, we should pay attention to HBV, even if HBV DNA is undetectable at the beginning of the treatment. Over the course of the disease, HBV DNA can become undetectable in 10% to 30% of the patients. IFN + LMV: small studies suggest that the addition of LMV is useful, above all in patients with active HBV. One such study described eight HBeAg-reactive patients, in whom HBV DNA and HCV RNA were detected, submitted to a 12-month course of IFN + LMV, followed by an additional six months of treatment with LMV. In approximately one-third of the patients, there was HBV and HBeAg clearance. In half of the patients, there was normalization of ALT, HCV clearance, and an SVR. It would be premature to venture an opinion on what is the most appropriate regimen, considering that further studies, involving larger patient samples, are still needed. Adefovir and entecavir: there have been no studies involving co-infected individuals. These might be good options in patients with dominant HBV. Liver transplantation: little experience with co-infected individuals. There are reports of higher survival compared with individuals mono-infected with HBV, with suggestion of the benefit of HCV suppression over HBV in immunosuppression after transplantation. In order to define the risks and benefits, it is necessary to await the results of further, larger studies. Triple Co-Infection a) HBV, HCV, and hepatitis D virus (HDV): only patients who are already infected with HBV can acquire HDV. The triple infection can occur in certain geographic areas where the prevalence of infection with HBV is high. The evolution to severe disease is described, the desired treatment being that that consists of high doses of IFN for a long period and, still, with poor responses. www.bjid.com.br BJID 2007; 11 Supplement 1 (October) HBV/HCV Co-Infection There are few relevant studies in the literature, and all of those involved small study samples, which precludes the standardization of the procedure in these cases. There are reports of treatment of seven patients with achievement of sustained biochemical response in two patients, and worsening with side effects in two other patients. b) HBV, HCV, and HIV: in these cases, the complexity of the HBV/HCV interaction is added to the effect of the HIV on the host immunologic system. It is known that, in HCV/HIV and HBV/HIV co-infections, the evolution of the hepatic disease is faster and potentially more severe. In addition, after the introduction of the highdose anti-HIV therapy known as highly active antiretroviral therapy, the mortality rate for chronic liver diseases has increased. There are few studies on the treatment of individuals coinfected with HBV, HCV, and HIV. The rate at which an SVR is achieved has been reported to be 17% with the use of IFN, compared with 25% for IFN + RBV. It must be borne in mind that, due to the frequent use of LMV in the antiretroviral therapy for HIV, HBV resistance to LMV is high in co-infected individuals. In one small study, the effect against the dominant virus, HBV or HCV, did not lead to the reactivation of the other, except in one case involving a patient with a very low CD4 lymphocyte count. 39 Studies of new antiviral drugs that are more effective against HBV (tenofovir, entecavir, adefovir, etc.) in HBV/ HIV or HBV/HCV/HIV co-infected individuals might inform therapeutic decisions regarding the complex treatment of co-infections involving HBV. References 1. Chakravarti A., et al. Characteristics of dual infection of hepatitis B and C viruses among patients with chronic liver disease: a study from tertiary care hospital. Trop Gastroenterol 2005;26:183-7. 2. Crockett S.D., et al. Natural story and treatment of hepatitis B virus and hepatitis C virus coinfection. Ann Clin Microbiol Antimicrob 2005;4:1-12. 3. French A.L., et al. Isolated hepatitis B core antibody is associated with HIV and ongoind but not resolved hepatitis C virus infection in a cohort of US women. JID 2007;195:1437-41. 4. Liu Z., et al. Hepatitis B virus (HBV) and hepatitis C virus (HCV) dual infection. Int J Med Sci 2006;3:57-62. 5. Sagnelli E., et al. Virologic and clinical expressions of reciprocal inhibitory effect of hepatitis B, C, and Delta viruses in patients with chronic hepatitis. Hepatology 2000;32:1106-10. 6. Sagnelli E., et al. HBV superinfection in hepatitis C cirus chronic carriers, viral interaction, and clinical course. Hepatology 2002;36:1285-91. 7. Sagnelli E., et al. Hepatitis C virus superinfection in hepatitis B virus chronic carriers: a reciprocal viral interaction and a variable clinical course. J Clin Virol 2006;35:317-20. 8. Soriano V., et al. Treatment of chronic hepatitis B or C in HIVinfected patients with dual viral hepatitis. JID 2007;195:1181-3. 9. Wang Y.M., et al. Suppression of hepatitis C virus by hepatitis B virus in coinfected patients at the National University Hospital of Singapore. J Gastroenterol 1999;34:481-5. www.bjid.com.br 40 BJID 2007; 11 Supplement 1 (October) Co-Infection with Hepatitis C Virus and Human T Lymphocyte Virus Carlos Brites Alves Federal University of Bahia, School of Medicine; Salvador, BA, Brazil Most individuals infected with human T lymphocyte virus (HTLV) type 1 or 2 will not develop the disease related to this virus, remaining asymptomatic for the rest of their lives. This fact has important implications for prospective counseling and evaluation of this population. Individuals infected with this virus, once identified, should be submitted to anamnesis and complete physical examination (in order to identify early manifestations of the disease and probable forms of acquiring the infection) and should be periodically evaluated every 6-12 months. It is recommended that injection drug users be tested for other pathogens common to this population, such as the hepatitis B virus, the hepatitis C virus (HCV), HIV, etc. It is recommended that partners of sexually active individuals be tested for HTLV. Children of women infected with HTLV-1 should be tested. Follow-up evaluations should include the following periodic laboratory tests (every 6-12 months): complete blood workups with platelet counts; parasitological stool examination (testing for Strongyloides); urine and urine sediment test (urinary infection). Although the proviral load of HTLV-1 is still under evaluation and has yet to be validated, some studies recommend annual quantification. Special Situations • In asymptomatic cases that present evidence of HTLVrelated systemic disease, such as dermatological alterations, hyperreflexia, clone or Babinski sign: • Serum calcium level, immunophenotyping of T lymphocytes (CD3, CD4, CD8, histocompatibility leukocyte antigen-DR, CD38 and CD56); diffuse histiocytic lymphoma, creatine phosphokinase, folate and vitamin B12 levels; free thyroxine 4 and thyroid stimulating hormone; and study of somatosensitive evoked potentials. Healthy individuals infected with HTLV-1 should be counseled regarding the transmission mechanisms of the infection and be reassured that the probability of developing the disease in the future is low. If necessary, they should be referred for specialized psychological follow-up evaluation. Currently, there is no indication – based on scientific evidence – that any certain type of specific anti-HTLV-1 pharmacological intervention plays a role in the prophylaxis of HTLV-related diseases. Therefore, there is no indication for the use of immunomodulatory, immunosuppressant or antiretroviral drugs in asymptomatic individuals infected with HTLV (National Ministry of Health, guide of clinical management of the HTLV-infected patient, 2004). The Brazilian Journal of Infectious Diseases 2007;11 (5) Suppl. 1:40-41. © 2007 by The Brazilian Journal of Infectious Diseases and Contexto Publishing. All rights reserved. HTLV/HCV Co-Infection Infection with HCV is frequently detected in HTLV-infected individuals and vice versa, as seen in other commonly transmitted pathogens. Co-infection with HTLV-2 and HCV in patients who are drug users has been reported, principally in cohorts in the northern hemisphere [1]. A study carried out in Paraná, Brazil revealed a strong association (OR=22.60; 95% CI: 10.35-49.35) between these two pathogens, probably reflecting shared transmission forms [2]. The prevalence of co-infection also seems to increase in individuals infected with HIV in Brazil. Segurado et al. [3] demonstrated that HCV infection was an independent risk factor for HTLV infection (adjusted OR=6.43, p=0.02). Interactions Between HCV and HTLV in Co-Infected Individuals: Potential Clinical Implications There are few studies on the effects of co-infection with HCV and HTLV. Hisada et al. [4] demonstrated that the co-infection with HTLV is associated with greater viral load of HCV. In a study conducted in Japan, HCV/HTLV-1 co-infected individuals were found to be at a higher risk of incidental liver disease (RR = 5.9), hepatocarcinoma and death (RR = 21.9), as well as for developing diabetes [5]. In addition, co-infected individuals have been shown to present a higher frequency of anergy to purified protein derivative, although with no statistical significance, suggesting a differentiated immunomodulatory effect in this population [6]. A recent study conducted in the state of Bahia, Brazil revealed a high prevalence of HTLV/HCV co-infection. Although it did not evaluate the clinical impact, it showed the relevance of this association. In practical terms, there is no established recommendation for the management of HCV/HTLV co-infection. Nevertheless, the analysis what evidence there is suggests that this association can result in significant modifications in the natural history of HCV, increasing the viral load of HCV, as well as increasing the morbidity and mortality associated with this infection. Therefore, the co-infected patient requires special attention regarding the clinical evolution of hepatitis C and the markers of the infection. In addition to monitoring the parameters related to HTLV infection, especially the neurologic alterations secondary to the infection, we should carefully evaluate the stage of the liver disease, and routinely evaluate the viral load of HCV. Attention should be given to potential alterations of glucose metabolism, since there seems to be an increased tendency toward these problems in the co-infected individual. For coinfected patients, routine evaluations of fasting glycemia, as well as glucose tolerance tests, could be necessary. www.bjid.com.br BJID 2007; 11 Supplement 1 (October) HCV/HTLV Co-Infection References 1. Zunt J.R., Tapia K., Thiede H., et al. HTLV-2 infection in injection drug users in King County, Washington, Scand J Infect Dis 2006;38(8):654-63. 2. Morimoto H.K., Caterino-De-Araujo A., Morimoto A.A., et al. Seroprevalence and risk factors for human T cell lymphotropic virus type 1 and 2 infection in human immunodeficiency virusinfected patients attending AIDS referral center health units in Londrina and other communities in Paraná, Brazil (AIDS Res Hum Retroviruses 2005;21(4):256-62. 3. Segurado A.C., Braga P., Etzel A., Cardoso M.R. Hepatitis C virus coinfection in a cohort of HIV-infected individuals from Santos, Brazil: seroprevalence and associated factors. AIDS Patient Care STDS 2004;18(3):135-43. 41 4. H i s a d a M . , C h a t t e r j e e N . , Z h a n g M . , e t a l . I n c r e a s e d hepatitis C virus load among injection drug users infected with human immunodeficiency virus and human T lymphotropic virus type II. J Infect Dis 2003;188(6):891-7. 5. Boschi-Pinto C., Stuver S., Okayama A., et al. A follow-up study of morbidity and mortality associated with hepatitis C virus infection and its interaction with human T lymphotropic virus type I in Miyazaki, Japan. J Infect Dis 2000;182(1):379-80. 6. Hisada M., Shima T., Okayama A., et al. Suppression of skin reactivity to purified protein derivative by hepatitis C virus among HTLV-1 carriers in Japan.J Acquir Immune Defic Syndr Hum Retrovirol 1998;19(4):421-5. www.bjid.com.br 42 BJID 2007; 11 Supplement 1 (October) Basic Guidelines for the Treatment of HIV/HVC Co-Infection Edgard De Bortholi Emílio Ribas Institute of Infectious Diseases; São Paulo, SP, Brazil The treatment of HIV/HCV co-infection presents many questions that have not yet been answered or on which there is no consensus. Since it is a recently introduced issue, the guidelines in the literature are divergent on some points. The still fragmented knowledge and lack of long-term worldwide experience in the treatment of such co-infections has forced referral facilities to constantly update their approaches. Some guidelines for the management of the HIV/HCV coinfected patient have been proposed, and new recommendations are particularly necessary: • Management of patients with persistently normal aminotransferase levels. • Definition and quantification of liver fibrosis: when and how? • Predictors of the response to anti-HCV therapy in coinfected patients. • Therapeutic doses of pegylated interferon and ribavirin. • Treatment duration. • Treatment of nonresponsive and recidivist patients. • Treatment of acute infection in HIV-positive patients. • HIV/HCV/HBV co-infected patients. • Interaction between antiretroviral drugs and anti-HCV therapeutics. • Antiretroviral hepatotoxicity in co-infected patients. • Antiretroviral drugs and recommended doses in hepatic insufficiency. Management of Patients with Persistently Normal Aminotransferase Establishing the persistence of normal aminotransferase levels in HCV-infected patients is difficult, especially in coinfected patients. Fluctuations in the levels of aspartate aminotransferase and alanine aminotransferase are common in this group of patients due to several factors, among which are the use of drugs of hepatotoxic potential, alcohol abuse, and infection with other opportunistic agents. In contrast to mono-infected patients, who present persistently normal alanine aminotransferase levels (~25%), co-infected patients present levels of 7-9%. However, of such patients, 25-40% present advanced liver fibrosis, which leads to liver cirrhosis. The rapid evolution of fibrosis in co-infected patients, even in those with normal transaminase levels, indicates treatment, based on patient motivation, duration of the disease, fibrosis stage, and viral load of HCV. The Brazilian Journal of Infectious Diseases 2007;11 (5) Suppl. 1:42-46. © 2007 by The Brazilian Journal of Infectious Diseases and Contexto Publishing. All rights reserved. Definition and Quantification of Liver Fibrosis: When and How? Various studies have demonstrated the rapidity of liver fibrosis progression in HIV/HCV co-infected patients. Such patients, even those presenting little or no fibrosis, should undergo histological evaluation at least every two years. Unfortunately, since it is an invasive procedure, liver biopsy might present complications resulting from technical performance. Pathologist reports are often made difficult by the small size of the liver fragments obtained in the biopsy, which has repercussions for the indication of anti-HCV therapy. Despite the disadvantages of liver biopsy, it remains the principal technique for determining the severity of hepatic injury. Noninvasive procedures to assess the level of the liver fibrosis are currently divided into two categories: imaging techniques, such as elastometry (FibroScan); and the use of biochemical markers (Fibrotest, APRISHASTA, FIB-4, and Forn Index). These procedures are accurate at discriminating between the absence of fibrosis and advanced fibrosis but are not very precise at evaluating the intermediate stages of the fibrosis. They present good predictive value for advanced liver fibrosis and cirrhosis. Biochemical marker determination in co-infected patients is of little utility, given the inflammatory nature of the HIV disease and the fact that, in this population, hepatotoxic drugs are used. Such drugs interfere with the serum markers of fibrosis in various ways: atazanavir elevates bilirubin levels; non-nucleoside analog reverse transcriptase inhibitors affect gamma glutamyl-transferase levels; and some protease inhibitors elevate cholesterol levels. Due to technical difficulties, complications and the number of times that the degree of fibrosis must be determined, liver biopsy might soon be replaced by FibroScan for the evaluation of co-infected patients, at least in one of their follow-up visits. Predictors of the Response to Anti-HCV Therapy in CoInfected Patients The current trend in the treatment of HCV infection is to individualize the approach. All HIV/HCV co-infected patients should undergo HCV genotyping before any therapeutic decision is made. Since the first attempts at treatment of HCV infection in HIV-positive patients, a small percentage of patients presenting a sustained virological response at the end of the treatment has been demonstrated, especially those with manifested immunosuppression with low CD4 counts. These patients present high viral loads of HCV, lower CD8 anti-HCV responses, greater frequency of liver steatosis caused by the www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Treatment of HIV/HCV Co-Infection use of alcohol and hepatotoxic drugs (resulting from a lower kinetic response to HCV treatment), a higher percentage of adverse effects, and worse treatment compliance. In order to achieve an early virological response, it is necessary to interfere with the natural and adaptive responses, which are reduced in co-infected patients, inducing a delay in viral clearance, which translates to a late virological response. High levels of HCV RNA in co-infected patients might also explain the low rate at which a sustained virological response is achieved. Approximately 30% of co-infected patients do not present a significant reduction in HCV viremia during the first month of treatment with pegylated interferon and ribavirin. The precise indication for treatment in HIV/HCV coinfected patients is CD4 counts greater than 350 cells/mm3. In patients with CD4 counts between 200 and 350 cells/mm3, the decision to treat the HCV infection should take other factors into account, such as the duration of the HCV infection, the severity of the liver disease, the degree of HIV suppression, and the classic predictors of the response to the HCV treatment (genotype and viral load). A sustained virological response can be predicted based on the HCV RNA serum negativity in week 4 of treatment. However, a reduction of less than 2 log IU/mL in the viral load of HCV in week 12 or the presence of detectable viremia in week 24 predicts the absence of a virological response, and the treatment should be discontinued. Ongoing studies focus on the maintenance of the treatment for 72 weeks in co-infected patients who are nonresponsive at week 24. Perhaps this, or even a longer time with smaller and fractionated doses, is the real treatment time for HIVpositive patients. Contraindications and Special Populations The patients with hepatic decompensation (ascites, digestive bleeding, hepatic encephalopathy, etc.) cannot be treated with interferon due to the high risk of developing serious complications. In these patients, the possibility of a liver transplant should be considered, even though this possibility is still remote because of the inherent difficulties of immunosuppression and HIV infection. However, the patients with compensated (Child-Pugh class A or B) cirrhosis can be treated since they are the ones that benefit most from the treatment. As to intravenous drug users and chronic alcoholics, the treatment should be postponed until the habits are controlled, and such patients should be referred to a detoxification program. Therapeutic Doses of Pegylated Interferon and Ribavirin To date, the efficacy of high doses of pegylated interferon in the treatment of HIV/HCV co-infected patients has not been confirmed, and we should therefore await the results of future investigations. Ribavirin induces errors in the viral replication cycle, and this effect is relevant, principally in HIV-positive patients, in whom 43 the mediated immune response is impaired. Various authors have demonstrated that the dose of ribavirin is directly related to achieving a sustained virological response. High doses of this drug are fundamental to the maintenance of viral suppression, especially in the first weeks of the treatment. Therefore, the recommended dose for the treatment of HCV is 15 mg/kg/day (the minimal dose for adjustment is 11 mg/kg/dose). Treatment Duration The current consensuses recommend that the duration of treatment for HIV/HCV co-infected patients be 48 weeks regardless of the genotype. However, values of HCV RNA > 2 log IU/mL at week 12 have a negative predictive value similar to that observed for mono-infected patients. Nevertheless, recent studies have questioned this simplistic view of the treatment time. When patients infected with genotype 2 or 3 test negative for HCV RNA in week 4 and maintain their negativity until week 12, the treatment time is reduced to 24 weeks. When patients infected with genotype 1 or 4 are in the same situation, the duration of treatment is extended to 48 weeks. For the patients who, despite positivity in week 4, present a drop in HCV RNA > 2 log IU/mL by week 12, the polymerase chain reaction (PCR) for qualitative HCV should be repeated in week 12. If the PCR is negative, and the genotype is 2 or 3, the treatment should continue to week 48. However, if the genotype is 1 or 4, the treatment should be maintained until week 72. If the qualitative PCR for HCV is still positive at week 24, or there is an HCV RNA reduction < 2 log IU/mL in week 12, the treatment should be discontinued, regardless of the genotype. Using erythropoietin and filgrastim has been a good strategy, even in co-infected patients, for the treatment of anemia and neutropenia, respectively. Treatment of Nonresponsive and Recidivist Patients At least to date, the HIV/HCV co-infected patients, in contrast with mono-infected patients, are not candidates for liver transplant, due to the disease progression and the absence of a therapeutic response. A growing number of co-infected patients who have already undergone treatment with standard interferon with or without ribavirin, especially those with progression of the fibrosis staging, can now be retreated with pegylated interferon and ribavirin. For the patients who have already used pegylated interferon and ribavirin and did not respond to the treatment or presented recurrence, there is currently no regimen for infection control. Despite the absence of a virological response in these patients, 35-43% of them present a reduction in the degree of liver fibrosis, demonstrating the antifibrotic effect of interferon. These data provide a rationale if the maintenance of interferon, even in small doses, for a prolonged time, has no effect on the progression of the fibrosis, even in the patients in whom HCV has not been eradicated. www.bjid.com.br 44 Treatment of HIV/HCV Co-Infection New anti-HCV drugs are urgently needed, especially for this group of patients, who depend exclusively on clinical treatment. Treatment of Acute Infection in HIV-Positive Patients Outbreaks of acute HCV infection in homosexuals have been reported in some European cities. Despite the knowledge of the low transmissibility of HCV through sex, sexual practices that generate traumatic lesions and genital ulcerations have been associated with the infection. The natural history of HCV infection in HIV patients has demonstrated its evolution to chronicity. Therefore, early therapeutic intervention (in the acute phase of infection) is particularly indicated in these cases, although treatment should not be instituted earlier than 12 weeks after the exposure, due to the possibility of spontaneous viral clearance. However, a delay in starting treatment might result in a reduction of the therapeutic response. The treatment of acute HCV infection in HIV-positive patients seems to provide a pattern of lower virological response when compared to HIV-negative patients. However, the viral clearance pattern obtained in HIV-positive patients in the acute phase is greater than that seen in those presenting chronic infection. Acute infection in HIV-positive patients should be treated with pegylated interferon and ribavirin for 24 weeks. HIV/HCV/HBV Co-Infected Patients In HIV-positive patients living in developed countries, the prevalence of multiple viral hepatitis (HBV/VCV; VBV/VDV; VBV/ HCV/HDV) is lower than 3%. However, this is still higher than that seen in the general population. Patients presenting HBV/ HCV co-infection seem to present reciprocal inhibition of viral replication, with one of the viruses being predominant. However, this predominance might oscillate from one virus to the other. Nevertheless, in patients with severe immunosuppression, replication of all of the viruses might occur simultaneously. In HIV-positive patients with good immune status, the interference seems to favor HCV, to the detriment of HBV. The progression of the liver disease seems to be more accelerated in HIV-positive patients infected with both HBV and HCV. In addition, these individuals are more likely to develop hepatocarcinoma. Using interferon to treat chronic HDV in HIV-positive patients is rarely effective. There is no established consensus as to the approach to treating multiple viral hepatitis. Few studies have examined the efficacy and safety of the combination of pegylated interferon and ribavirin for treating multiple infections in HIVpositive patients. When possible, all of the viruses involved in the hepatitis should be treated. Interaction between Antiretroviral Medications and AntiHCV Therapy The highly active antiretroviral therapy (HAART) regimen is associated with increased survival due to the fact that it BJID 2007; 11 Supplement 1 (October) controls HIV and slows the progression of the hepatic disease. The better prognosis that the HAART regimen confers on HIV/ HCV co-infected patients is probably due to deceleration of the progression of hepatic fibrosis. This has principally been demonstrated in HIV patients treated with protease inhibitors. Some studies have suggested the early introduction of antiretroviral therapy in HIV/HCV co-infected patients, considering the beginning of the HAART regimen in patients with CD4 counts > 350 cells/mm3, particularly in male patients and in those who acquired HCV at an advanced age, which would decrease the effects resulting from immune reconstitution and the slowing of the progression of HCVinduced hepatic disease. The principal complication in the treatment of HIV-infected patients is the interaction between ribavirin and some antiretroviral drugs. Ribavirin can decrease the concentrations of some nucleoside analog reverse transcriptase inhibitors (NRTIs), although there seem to be no clinical consequences. The adverse effects of the NRTIs have been associated with HCV co-infection, female gender, obesity, and prolonged exposure to these drugs. The principal NRTI combinations associated with lower impact on the liver are as follows: lamivudine+abacavir (3TC/ABC); 3TC+zidovudine (AZT); 3TC+ABC+AZT; and emtricitabine + tenofovir. However, recent studies have indicated that ribavirin interferes with the effects of ABC, and, therefore, its use in the HAART regimen for HCV patients should be considered with caution. The greatest toxicity is that resulting from the concomitant use of didanosine and ribavirin, worsening with the addition of stavudine. These combinations increase the possibility of mitochondrial toxicity and are potentiated by the presence of HIV and HCV. In patients treated with these antiretroviral drugs combined with ribavirin, lactic acidosis, pancreatitis and hepatic decompensation have been reported. Therefore, it is recommended that replacements for these antiretroviral drugs be found for patients who will initiate the treatment for HCV. In addition, AZT should be replaced, when possible, due to frequent, severe anemia and neutropenia when AZT is administered together with ribavirin. It is important to emphasize that the antiretroviral drugs, as well as the drugs used in the treatment and prophylaxis of the various opportunistic infections, are metabolized by the liver and, therefore, many of these metabolites can increase the risk of hepatic lesion. The chart below shows the principal interactions between the drugs used, facilitating the choice of antiretroviral drugs in the clinical approach to co-infected patients. The combination of ritonavir and saquinavir has proven to be highly hepatotoxic, and its use is not currently recommended, especially in HIV/HCV co-infected patients with concomitant tuberculosis, in whom noncompliance with treatment and, principally, the elevation of transaminase levels result from the discontinuation of the treatment with antituberculosis drugs or the introduction of alternative www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Treatment of HIV/HCV Co-Infection Chart 1. Drug interactions Antiretroviral drugs Adverse effects Therapeutic alternative Zidovudine Anemia and neutropenia Stavudine Didanosine Hepatotoxicity Mitochondrial toxicity Hepatotoxicity Mitochondrial toxicity Stavudine Nevirapine Hepatotoxicity Ribavirin Hepatotoxicity Interaction with anti-HCV therapy Potentiates the probability of anemia caused by ribavirin Lamivudine or efavirenz Ribavirin inhibits the phosphorylation of this drug Lamivudine or efavirenz Should not be combined with didanosine due to potentiation of mitochondrial toxicity Efavirenz Do not combine with didanosine and stavudine; potentiation of mitochondrial toxicity Tolerated in low doses Increases the probability of when used in combination intolerance to the antiretroviral with other protease inhibitors therapy. HCV=hepatitis C virus. Chart 2. Antiretrovirals and recommended doses in liver failure Name Hepatic metabolism Recommendations Yes Yes No No Unknown No Yes Mild (A*)=200 mg twice a day / moderate/severe=contraindicated Not recommended No need for dose adjustment -recommended No need for dose adjustment -recommended No need for dose adjustment -recommended No need for dose adjustment -recommended No need for dose adjustment -recommended No No need for dose adjustment -recommended Yes Yes Yes Not recommended Not recommended Mild (A*)/moderate (B*)=No need for dose adjustment -recommended Severe (C*)=contraindicated Protease inhibitors Atazanavir Yes Fosamprenavir Yes Indinavir Nelfinavir Ritonavir Yes Yes Yes Mild (A*)=No need for dose adjustment - recommended Moderate (B*)=300 mg/day. Severe (C*)=contraindicated Mild/moderate=700 mg 2xday Severe=contraindicated (because the dose cannot be reduced to lower than 700 mg) Mild/moderate=600 mg 3× day. Severe=not recommended Not recommended Mild/moderate=No need for dose adjustment -recommended Severe=not recommended Mild/ moderate=not recommended. Severe=contraindicated NRTIs Abacavir Didanosine Emtricitabine Lamivudine Stavudine Zalcitabine Zidovudine NtRTIs Tenofovir NNRTIs Delavirdine Efavirenz Nevirapine Saquinavir Yes Boosters w/ ritonavir Atazanavir/ritonavir Yes Fosamprenavir/ritonavir Lopinavir/ritonavir Yes Saquinavir/ritonavir Yes Tipranavir/ritonavir Yes Fusion inhibitors Enfuvirtide Unknown Not recommended at some levels of hepatic insufficiency because it has not been adequately studied Yes Not recommended Not recommended Mild/moderate=Not recommended. Severe=contraindicated Mild=No need for dose adjustment - recommended Moderate/severe=contraindicated Not recommended *Child-Pugh class. NRTIs=nucleoside analog reverse transcriptase inhibitors; NtRTIs=nucleotide analog reverse transcriptase inhibitors; NNRTIs=non-nucleoside analog reverse transcriptase inhibitors. www.bjid.com.br 45 46 Treatment of HIV/HCV Co-Infection regimens that are less efficient in the resolution of infection with Mycobacterium tuberculosis. The data currently available are insufficient to determine the nature of the interaction of atazanavir, tenofovir, fosamprenavir, and tipranavir with the therapeutic regimen in HCV infection. References 1. Arends J.E., Boucher C.A.B., Hoepelman A.I.M. Hepatitis C virus and human imunodeficiency virus coinfected: where do we stand? Journal of Medicine 2005;63:156-63. BJID 2007; 11 Supplement 1 (October) 2. Lai, et al. Antiretroviral medication considerations for individuals coinfected with HIV and hepatitis C virus. AIDS Patient Care and STDs 2006;20(10):678-92. 3. Rockstroh J.K. Influence of viral hepatitis on HIV infection. Journal of Hepatology 2006;44:S25-S7. 4. Sancho A.R., Soriano Vicente. Coinfección por el VIH y el virus de la hepatitis C. http//www.doyma. Acessado em 26/02/2007. 5. Soriano V., et al. Care of patients coinfected with HIV and hepatitis C virus: 2007 updated recomendations from the HCV-HIV international panel. AIDS 2007,21:1073-89. 6. Thomas L.D. Options for treatment of hepatitis C in HIV-infected persons. Journal of Hepatology 2006;44:S40-S3. www.bjid.com.br BJID 2007; 11 Supplement 1 (October) 47 Basic Aspects of the Treatment for Hepatitis C: Mechanisms of Action of Interferon Alpha and Ribavirin and the Bases of Individualization Carlos Eduardo de Melo, Evaldo Stanislau Affonso de Araújo and Antonio Alci Barone Laboratory of Hepatitis, LIM 47 DCMIP-HC-FMUSP; São Paulo, SP, Brazil Pharmacological Characteristics of Interferons and Ribavirin The treatment of patients with chronic hepatitis C has developed considerably in recent years. However, it is still based on the use of interferon alpha (IFN-α) as an antiviral and immunomodulatory agent against the hepatitis C virus (HCV). The IFNs are a family of proteins that are naturally produced by the cells of the immune system. The IFN-á protein presents antiviral, antiproliferative and immunomodulatory activity [1-3]. Its mechanism of biological action occurs through the activation of specific genes, influencing cell growth and division, as well as modulating some immune system activities. Therefore, IFNs have an indirect antiviral effect on HCV [2,4]. Commercially, IFN-α is produced by means of recombinant DNA techniques and is available in preparations of two distinct subtypes (IFN-α 2a or IFN-α 2b), which can be combined with other molecules, such as polyethylene glycolor, more recently, albumin [5,6]. The only difference between IFN-α 2a and IFN-α 2b is in the amino acid present at position 23 of the protein: IFN-α 2a has a lysine at that position, whereas IFN-α 2b has an arginine [7]. After the binding with its specific receptor (IFNAR) on the surface of the target cells, IFN-α activates an intracellular signaling cascade, which takes the induction of IFN-stimulated genes (ISGs), establishing a non-virus-specific antiviral state inside the cell [3,7]. The principal signaling mechanism used by IFN-α is the so-called Janus kinase/signal transducers and activators of transcription (Jak/STAT) pathway [3]. Therefore, two cytoplasmatic proteins with the activity of tyrosine kinase associated with IFNAR, activated Jak1 and tyrosine kinase 2 (Tyk2), are activated by the dimerization of the receptors. Activated Jak1 and Tyk2 perform the phosphorylation of STAT1 and STAT2, respectively. The phosphorylated STAT1 and STAT2 bond with the protein p48 forming IFN-stimulated gene factor 3 (ISGF3), which translocates into the nucleus and bonds with IFN-stimulated regulatory element in the sequences which promote a variety of genes inducible by IFN-α including antiviral proteins such as 2’5’-oligoadenylate synthetase (2’5’OAS), protein kinase RNA, and Mx protein [1,3,7,8]. The absorption of IFN-α (2a or 2b) is high (above 80%) when administrated intramuscularly or subcutaneously. The concentration typically peaks at 3-12 h after administration [9]. The Brazilian Journal of Infectious Diseases 2007;11 (5) Suppl. 1:47-48. © 2007 by The Brazilian Journal of Infectious Diseases and Contexto Publishing. All rights reserved. The metabolism and elimination of IFN-α occurs principally via the kidneys, with a half-life of 3-8 h [9]. Pharmacological Characteristics of the Pegylated Interferons Pegylated IFNs (PEG-IFNs) are produced through the binding of an inert molecule of polyethylene glycol to the recombinant IFN-á, thus reducing the renal clearance, altering the metabolism and increasing the half-life of the IFN molecule, although maintaining all of its immunostimulatory characteristics [10,11]. The two PEG-IFNs currently available are produced with polyethylene glycol molecules of different complexities. PEGIFN-α 2b consists of the binding of IFN-α 2b with a linear PEG chain, forming a 12-kDa molecule. PEG-IFN-α 2a is formed by the binding of two 20-kDa chains with IFN-α 2a, resulting in a complex 40-kDa molecule [6]. The differences in the chemical structure of the two PEGIFN-α formulations are associated with significant differences in the pharmacological characteristics of the two drugs. The PEG-IFN-α 2b (12 kDa) is more rapidly absorbed (with an absorption half-life of 4.6 h), presents a wide volume of body distribution (approximately 0.99 L/ kg) and a mean elimination time of 40 h. However, PEG-IFNα 2a (40 kDa) is absorbed more slowly (absorption half-life, 50 h), its distribution is restricted to well-vascularized organs with good perfusion, such as the liver, and it remains detectable in the serum for one week (approximately 65 h elimination half-life) [6,12,13]. Pharmacological Characteristics of Ribavirin Ribavirin is a synthetic nucleoside which is structurally similar to guanosine [14,15]. Ribavirin enters into the eukaryotic cells rapidly and, after it undergoes intracellular phosphorylation, shows virustatic activity against a broad spectrum of DNA and RNA viruses [14,15]. The exact mechanism of the antiviral action of ribavirin has not yet been totally elucidated [1,16]. However, some studies suggest the following possible mechanisms: a) direct inhibition of HCV replication; b) inhibition of the enzyme inosine monophosphate dehydrogenase of the host; c) induction of mutagenesis in the viral RNA; d) immunomodulation by the induction of a T helper 1 (Th1)-type immune response Ribavirin is rapidly absorbed (half-life of approximately 2 h) and widely distributed throughout the body after its oral administration; its metabolization occurs principally via the kidneys [16]. www.bjid.com.br 48 Treatment for Hepatitis C: Basic Aspects Treatment with IFN-α has as a success-defining characteristic, progressively more extensive and vigorous immune stimulation. The more rapid the stimulation is, the greater are the chances of success. The study of mononuclear cells ex vivo and in vivo demonstrated that, 3-6 h after the administration of conventional IFN-α, 516 genes were upregulated, of which 88 with actions directly linked to immune functions [17]; the same phenomenon was observed for PEGIFN, also differentiating responders from nonresponders using the intensity of expression in certain IFN-inducible genes (2’5’OAS, MX1, IRF-7 and TLR-7), greater in the responders and lesser in the Afro-Americans [18,19]. The final pathway of the phenomenon triggered in the cell nucleus is the activation of effector cells. An initial activation of the innate immunity (natural killer cells) is supposedly necessary for the early reduction of the viremia – the greater and the more rapid it is, the more closely it associates with achieving a sustained virological response (also differentiating rapid responders from slow responders). Progressively, as of week 4 of the treatment, the effective immune stimulation induced by IFN with the respective reduction of the viremia would enable the specific defense mechanisms (CD4+ and CD8+ cells) which, in turn, would be in charge of disposing of the residual infected cells (hepatocytes and extrahepatic cells) [20]. In fact, Pillai et al. clearly showed that the magnitude and diversity of the cellular response was associated with early and sustained virological responses [21], in contrast to other authors who only associated the Th1-type cellular response with the initial viremia [22]. At any rate, it is clear that patients presenting a rapid and vigorous initial response have greater chances of success. However, patients presenting a slower response need more long-term stimulation. Therein reside the bases for the individualization of treatment. It is equally clear that this initial virological response depends on the gene stimulus induced by IFN-α. Whether or not these phenomena imply differences associated with the different types of IFNs used in clinical practice has yet to be answered. However, initial evidence was provided by the analysis of the expression of mRNA of inducible IFN genes in two groups of patients exposed to the two existing types of PEG-IFN, suggesting that, despite lower plasma exposure, the patients who used PEG-IFN-α 2b expressed their genes more vigorously, emphasizing the relevance of the intracellular environment in the response to HCV treatment [23]. References 1. Feld J.J., Hoofnagle J.H. Mechanism of action of interferon and ribavirin in treatment of hepatitis C. Nature 2005;436(7053):967-72. 2. Peters M. Actions of cytokines on the immune response and viral interactions: an overview. Hepatology 1996,23(4):909-16. 3. Wohnsland A., Hofmann W.P., Sarrazin C. Viral determinants of resistance to treatment in patients with hepatitis C. Clin Microbiol Rev 2007,20(1):23-38. BJID 2007; 11 Supplement 1 (October) 4. Souvignet C., Lejeune O., Trepo C. Interferon-based treatment of chronic hepatitis C. Biochimie 2007;89(6-7):894-8. 5. Chemmanur A.T., Wu G.Y. Drug evaluation: Albuferon-alpha—an antiviral interferon-alpha/albumin fusion protein. Curr Opin Investig Drugs 2006;7(8):750-8. 6. Foster G.R. Review article: pegylated interferons: chemical and clinical differences. Aliment Pharmacol Ther 2004;20(8):825-30. 7. Pestka S. The human interferon alpha species and receptors. Biopolymers 2000;55(4):254-87. 8. Pawlotsky J.-M. Mechanisms of antiviral treatment efficacy and failure in chronic hepatitis C. Antiviral Res 2003;59(1):1-11. 9. USP DI® Volume I: Drug Information for the Health Care Professional [database on CD-ROM]. Version 5.1. Greenwood Village, Colo: Thomson Micromedex. 10. Reddy K.R., Wright T.L., Pockros P.J., et al. Efficacy and safety of pegylated (40-kd) interferon alpha-2a compared with interferon alpha-2a in noncirrhotic patients with chronic hepatitis C. Hepatology 2001;33(2):433-8. 11. Strader D.B., Wright T., Thomas D.L., et al. Diagnosis, management, and treatment of hepatitis C. Hepatology 2004,39(4):1147-71. 12. Bailon P., Palleroni A., Schaffer C.A., et al. Rational design of a potent, long-lasting form of interferon: a 40 kDa branched polyethylene glycol-conjugated interferon alpha-2a for the treatment of hepatitis C. Bioconjug Chem 2001;12(2):195-202. 13. Glue P., Fang J.W., Rouzier-Panis R., et al. Pegylated interferonalpha2b: pharmacokinetics, pharmacodynamics, safety, and preliminary efficacy data. Hepatitis C Intervention Therapy Group. Clin Pharmacol Ther 2000;68(5):556-67. 14. Graci J.D., Cameron C.E. Mechanisms of action of ribavirin against distinct viruses. Rev Med Virol 2006;16(1):37-48. 15. Leyssen P., De Clercq E., Neyts J. Perspectives for the treatment of infections with Flaviviridae. Clin Microbiol Rev 2000;13(1):67-82. 16. Parker W.B. Metabolism and antiviral activity of ribavirin. Virus Res 2005;107(2):165-71. 17. Ji X., Cheung R., Cooper S., et al. Interferon alfa regulated gene expression in patients initiating interferon treatment for chronic hepatitis C. Gastroenterology 2003;37:610-21. 18. Taylor M.W., Tsukahara T., Brodsky L., et al. Changes in gene expression during pegylated interferon and ribavirin therapy of chronic hepatitis C virus distinguish responders from non responders to antiviral therapy. Journal of Virology 2007;81(7):3391-3401. 19. He X.-S., Ji X., Hale M.B., et al. Global transcriptional response to interferon is a determinant of HCV treatment outcome and is modified by race. Hepatology 2006;44:352-9. 20. Tang K.H., Herrmann E., Cooksley H., et al. Relationship between early HCV kinetics and T-cell reactivity in chronic hepatitis C genotype 1 during peginterferon and ribavirin therapy. Journal of Hepatology 2005;43:776-82. 21. Pillai V., Lee W.M., Thiele D.L., et al. Clinical responders to antiviral therapy of chronic HCV infection show elevated antiviral CD4+ and CD8+ T-cell responses. Journal of Viral Hepatitis 2007;14:318-29. 22. Aberle J.H., Perstinger G., Weseslindtner L., et al. CD4+ T cell responses in patients with chronic hepatitis C undergoing peginterferon/ribavirin therapy correlate with faster, but not sustained, viral clearance. The Journal of Infectious Diseases 2007;195:1315-9. 23. Silva M., Poo J., Wagner F., et al. A randomized trial to compare the pharmacokinetic, pharmacodynamic, and antiviral effects of peginterferon alfa-2b and peginterferon alfa-2a in patients with chronic hepatitis C (COMPARE). Journal of Hepatology 2006;45:204-13. www.bjid.com.br BJID 2007; 11 Supplement 1 (October) 49 Therapeutic Approach to Acute Hepatitis C Rodrigo Nogueira Angerami 1 and Fernando Lopes Gonçales Júnior 2 Epidemiological Surveillance Unit, Hospital Epidemiology Service, Hospital das Clínicas, State University at Campinas and Campinas Referral Center for STD/AIDS; 2Study Group on Hepatitis, Infectious Diseases Division, Clinical Medicine Department, School of Medical Sciences, State University at Campinas; Campinas, Brazil 1 It is estimated that 150 to 200 million individuals are currently infected with the hepatitis C virus (HCV) [1-3], and that, annually, there are 3 to 4 million new cases of infection worldwide [4]. In view of the impossibility of immunoprevention [5] - either through vaccines or the use of post-exposure immunoglobulin - and the risk of chronicity in individuals exposed to the disease, which is estimated to be between 50% and 85% [2,4,6-9,33], the definition of strategies aimed at early detection and treatment of infected individuals, preferably during the acute phase of the infection [2,4], has been widely discussed in recent years. In individuals presenting symptoms consistent with acute hepatitis C (AHC), it is known that the possibility of spontaneous viral clearance, among various factors, depends especially on a potent initial cellular immune response mediated by cytotoxic T lymphocytes - initially by HCVspecific CD8+ T lymphocytes and, subsequently, CD4+ T lymphocytes [10-12]. Other factors associated with a higher rate of sustained virological response (SVR) are the specific characteristics of the patients. Individuals who are Asian or Caucasian, female, young, and HIV-negative, as well as presenting a rapid drop in HCV RNA levels and presenting specific human leukocyte antigen class II alleles, together with the previously mentioned T cytotoxic response, also respond better to treatment [16,17,33]. Better responses are also found in cases of infections with shorter incubation period produced by smaller inoculates. In addition, in a prospective study conducted by Santantoni et al., a higher rate of spontaneous viral clearance was observed in patients infected with genotype 3 [7]. A diagnosis of acute infection with HCV remains a rare event in clinical practice. In 70-80% of infected patients, AHC is asymptomatic, and 75% are anicteric [4,8,13-15,33]. Individuals with AHC present elevated serum levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST), with or without a clinical profile of acute hepatitis, detection of (previously undetectable) HCV RNA and seroconversion for anti-HCV antibodies. It is estimated that infection with HCV accounts for only 20% of all diagnosed cases of acute hepatitis [13]. In the United States, there are 40,000 cases of AHC annually [12], and only a small percentage of those are clinically diagnosed in this phase. Considering the high risk of developing chronic hepatitis C and the favorable evidence, in terms of therapeutic response, it is fundamental to improve our capacity to properly detect and treat cases of acute infection with HCV [12]. At-Risk Populations Currently, it is thought that, in 90% of cases of infection with HCV, it is possible to identify the associated risk factors [14]. The Brazilian Journal of Infectious Diseases 2007;11 (5) Suppl. 1:49-52. © 2007 by The Brazilian Journal of Infectious Diseases and Contexto Publishing. All rights reserved. In the last two decades, especially after serologic triage was adopted as obligatory for blood donors, and due to the increased accuracy of serologic tests, there has been a drastic reduction in the number of new infections with HCV through blood and blood products. This resulted in significant changes in the epidemiological pattern of the disease [2,16,18]. However, new infections still occur via parenteral route, and, less frequently, by sexual or vertical transmission. Currently, the principal groups at risk for infection with HCV are as follows: health professionals exposed through cut or puncture accidents involving sharp objects contaminated with infectious material; intravenous drug users; individuals engaging in sex with HCV-positive partners; patients on hemodialysis; and patients submitted to various invasive procedures [4,12,14,18-21]. In the United States, intravenous drug users account for 68% of the new cases of HCV infection, compared with 18% for individuals engaging in sex with HCV-positive partners, 4% for health professionals exposed through cut or puncture accidents, 1% for individuals infected through other routes, and 9% for those in whom the source of infection cannot be identified [22]. Currently, it is believed that the risk of vertical transmission is significantly potentialized in cases of HIV/HCV co-infection, in which the rate of such transmission can be as high as 17% [37]. Special attention has been given to the occupational risk of infection with HCV in health professionals, and the estimated risk of cut or puncture accidents with sharp objects contaminated with infectious materials ranges from 1% to 2% [5,22]. Although risks, impacts and recommendations are still controversial, it is estimated that the risk of sexual transmission ranges from 0% to 3% [34,37], increasing especially in specific situations, such as co-infection with HIV or other sexually transmitted diseases, and between homosexual partners. It is also known that the risk of HCV infection is higher in individuals with chronic kidney disease under treatment with hemodialysis. It is believed that, among such individuals treated in the United States, the prevalence of anti-HCV reactivity ranges from 15% to 50% [37]. The definition of protocols for clinical and laboratory periodic follow-up evaluation in these segments at increased risk is fundamental in order to improve the detection of AHC. Laboratory Diagnosis The laboratory techniques most commonly used in the detection of anti-HCV antibodies are fundamentally based on third- and fourth-generation enzyme-linked immunosorbent assays. Such serologic tests have demonstrated great specificity (> 99%) and sensitivity (95%99%) [38]. However, one of the possible limitations in the diagnosis of acute infection with HCV results from the possibility of late seroconversion, occurring at 4 to 10 weeks after exposure [23]. Regarding detection of anti-HCV www.bjid.com.br 50 Therapeutic Approach to Acute Hepatitis C antibodies, we observe 50% to 70% positivity at symptom onset; in addition, from postinfection month 3 onward, antibodies are detected in 90% of the cases [38]. However, HCV RNA determination constitutes an important tool for early detection of acute infections, since HCV is detectable by 1 to 3 weeks after transmission [23,38]. Immediately after HCV RNA detection and concomitantly with the appearance of possible signs and symptoms - 2 to 8 weeks after infection - it is possible to observe increased ALT levels [38]. Detection of HCV RNA, in the absence of anti-HCV antibodies, strongly suggests acute infection, especially when subsequent anti-HCV seroconversion is observed. The use of transcription-mediated amplification (TMA), together with other available techniques, constitutes an additional strategy for early detection of acute HCV infection [23]. Therapeutic Approaches The high rate of chronicity observed after infection with HCV justifies the increasing interest in possible therapeutic approaches in cases of acute infection. Various approaches have been proposed in literature, all of which aim at preventing the progression to chronic disease. Nevertheless, to date, the ideal treatment regimen to be adopted in clinical practice has not been established [4,6,8,9,14,18,19,24]. Comparing results from different clinical trials is difficult for the following reasons: acute hepatitis C is frequently asymptomatic; different criteria are used in case definition; samples of patients are small and heterogeneous; there are no control groups; different types and doses of interferon (IFN) are used; there are different end points, and follow-up time varies enormously. Therefore, it is impossible to define an ideal therapeutic regimen. When we evaluate the data available in literature, we observe excellent results in patients treated in the acute phase of the infection, even under monotherapy [15,16]. The principal post-treatment outcome measure is the rate at which patients achieve an SVR, defined as the absence of HCV RNA detected in serum by qualitative polymerase chain reaction (PCR) at 24 weeks after the end of the treatment [25,26]. Among patients treated in the acute phase of the infection, this rate ranges from 37% to 98% [2,4,8,18,20,24], which is higher than the 54%56% observed among patients with chronic hepatitis C, even among those under combined therapy with pegylated IFN (PEGIFN) and ribavirin (RBV) [25,26]. Various trials evaluate the efficacy of different regimens using conventional IFN alpha (IFN α) in the treatment of AHC. The best results to date were those reported by Jaeckel et al., whose treatment regimen consisted of monotherapy with IFN α-2b with inducement - 5 million IU/day, s.c., for 4 weeks, followed by 5 million IU, 3 times a week, for 20 weeks [2]. In that study, the use of IFN, even without RBV, made it possible to obtain high SVR levels. However, the Jaeckel et al. study raises the following considerations: 68% of the patients included in the study were icteric and therefore presented a higher probability of having self-limited infection, which could, in part, explain the high SVR rate reported. As previously mentioned, symptomatic individuals can evolve to spontaneous viral clearance. In addition, Kamal et al. observed a higher SVR rate in treated symptomatic individuals when compared to those asymptomatic individuals treated BJID 2007; 11 Supplement 1 (October) with the same therapeutic regimen (96% and 76%, respectively) [8]. However, Santantonio et al. reported an SVR rate of 94% among individuals who did not present spontaneous viral clearance by week 12 after acute symptom onset, when treated with PEG-IFN α-2b, in monotherapy, for 24 weeks [12]. Such finding corroborates other reports demonstrating that it is safe to adopt the expected conduct up to week 21 in order to await possible spontaneous viral clearance [15,16,20,28,36]. In view of these facts, it should be noted that, among individuals acutely infected with HCV, those who are asymptomatic, especially those who are anicteric and present normal ALT levels, are more likely to present spontaneous viral clearance [8,12,14,18,20] - typically between weeks 5 and 12 [27,28]. This viral clearance can occur, in rare situations (in 10%-50% of cases), up to 24 weeks after symptom onset [8,9,12-15,17,18,21]. Various authors suggest that the treatment be initiated, at most, 12 to 16 weeks after symptom onset, avoiding unnecessary exposure to IFN in patients who can clear HCV spontaneously [4,10,12,18,33]. However, some authors suggest that, in cases of asymptomatic acute infection, treatment should be introduced at the time of diagnosis [12]. It should be noted, however, that lower SVR rates have been observed in cases of late treatment. These rates were initially described by Nomura et al. [18] who, using IFN α (6 million IU, i.m., daily, for four weeks), reported an SVR in 87% of early treated individuals - 8 weeks after detection of symptoms - and in 40% of patients treated after AHC symptom onset. This study demonstrated that early introduction of treatment enabled a high SVR rate, even in short-term treatment regimen. More recently, in a study conducted by Kamal et al., the SVR rate in individuals treated after week 20 was 76.6%, whereas, in individuals who started therapy at weeks 8 and 10, the rates were, respectively, 95.3% and 93.2% [20]. Similarly to what was observed in the treatment of chronic hepatitis C [14,29,30], negative HCV RNA at treatment week 1 was associated with a greater probability of achieving an SVR [31]. In the study carried out by Kamal et al., achieving a rapid virological response (RVR), defined as HCV RNA negativity or a ≥ 2 log10 drop in HCV RNA, by treatment week 4, was found to have positive and negative predictive values of 88% and 98%, respectively, for achieving an SVR. In another study, involving individuals with AHC treated with IFN as monotherapy in daily doses for 4 weeks, HCV RNA negativity in the first week of treatment was found to have a positive predictive value of 87% for achieving an SVR [18]. The combination of PEG-IFN α and RBV is currently considered the first-line treatment regimen for patients with chronic hepatitis C, those infected with genotype 2 or 3 and treated thusly achieving an SVR at a rate of 82% [20]. However, the efficacy, safety, duration and appropriate timing of PEG-IFN treatment in cases of AHC have not yet been well established. A multicenter, prospective, randomized controlled study, using PEG-IFN α-2b and involving a 48-week post-treatment follow-up period, reported SVR rates of 95.3% and 93.2%, respectively, in groups of patients in which treatment was initiated at week 8 or week 12 after symptom onset [20]. Studies that compared treatment results obtained with conventional IFN to those obtained with PEG-IFN found similarly high www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Therapeutic Approach to Acute Hepatitis C efficacy in both formulations [4]. Kamal et al., comparing the efficacy of PEG-IFN α, with and without RBV, reported 85% and 80% SVR rates, respectively [20]. It should be highlighted, however, that no evidence justifies the routine use of the combination with RBV up to the present moment. In a multicenter study, patients with acute hepatitis C received PEG-IFN α-2b (in a dose of 1.0-1.5 μg/kg/week) initiated at the time of diagnosis and continued for 12 weeks; HCV RNA negativity was found in 87% at treatment week 4, in 91% at the end of the treatment (week 12), and in 73% at 24 weeks after the end of the treatment [9]. In patients treated with higher doses of PEG-IFN (≥ 1.2 μg/kg/week for 12 weeks), the same authors reported an 84% SVR rate, comparable to that found for 24-week regimens [9]. In a prospective, uncontrolled study, De Rosa et al., using PEG-IFN α-2b (1.01.6 μg/kg/week, during 12 weeks) initiated immediately after diagnosis, reported an overall rate of 74%, although higher rates were reported when higher doses of PEG-IFN were used (82% when ≥ 1.33 μg/kg/week doses were used) [21]. Similar to what was observed in the treatment for chronic hepatitis C, compliance to treatment regimen has proven an important SVR predictive factor in the treatment of AHC. This was demonstrated in a multicenter study, in which a 71% overall response rate was found, compared with an 89% rate found in the population with expected compliance to treatment [32]. When SVR rates were evaluated in difficult-to-treat populations, two studies involving intravenous drug users reported SVR rates between 72% and 74% [9.21]. In both situations, regular follow-up and multidisciplinary approach were used in order to provide adequate compliance to the proposed treatment regimen. Kamal et al. reported that HCV genotype is also an element to be considered as an SVR predictive factor in patients with AHC. Therefore, similar to what was reported in cases of chronic hepatitis C, there were associations among genotypes, treatment time, and SVR rates, in patients with acute hepatitis C. Distinct SVR rates were found, according to the genotype involved: 63.8% for genotype 1; and 100% for genotypes 2 and 3. However, when a prolonged (24-week) treatment regimen was adopted, an SVR rate of 88% was observed in patients infected with genotype 1, higher than the 38% and 60% reported for 8-week and 12-week regimens, respectively [8]. In addition to genotypes other than genotype 1, relevant positive predictive factors for achieving an SVR include lower viral load at the start of treatment, early initiation of treatment, and rapid negative HCV RNA after the initiation of treatment [8,18]. Calleri et al. reported that, in addition to achieving an RVR (by week 4 of treatment) and low pre-treatment viremia, PEG-IFN doses ≥ 1.2 mg/kg/week were positive predictive factors for achieving an SVR [9]. It is not currently possible to determine the best treatment regimen for AHC cases. Nor can we establish a minimum follow-up time to be adopted after the end of the treatment. Wiegand et al., in a prospective study using biochemical, ultrasound, and virological monitoring (HCV-PCR, TMA and RNA detection in peripheral mononuclear cells) of patients treated with IFN α, in monotherapy, did not report any evidence of viral recurrence or hepatocytic lesion during the period studied (mean, 135 weeks; range, 52-224 weeks) [10]. However, as suggested by Alberti et al. [16] in a meta-analysis 51 and subsequently by other authors, the minimum clinical, biochemical, and virological follow-up time should not be less than 48 weeks. Conclusions High SVR rates have been reported in patients treated for acute hepatitis C. In view of this, various national and international consensuses have recommended the treatment of this infection [6,14,19,34,35]. However, there is no consensus regarding the proper timing of the initiation of treatment onset duration of treatment. The best treatment regimen to be used, as well as the length of the post-treatment follow-up period, is also debatable. Taking into account the fact that acute hepatitis C is generally asymptomatic, serologic triage and often HCV RNA testing are universally recommended strategies for correct diagnostic approach of intravenous drug users, patients infected with HIV, patients on hemodialysis, children of mothers with HCV, sexual partners of HCV-positive individuals, and health professionals having been exposed to HCV. These segments are currently considered the principal at-risk groups for acute infection with HCV and, therefore, potentially considered for treatment. Diagnostic laboratory tests should include anti-HCV antibody tests (detectable between post-infection weeks 4 and 10), determination of serum levels of aminotransferases such as ALT (high between post-exposure weeks 2 and 5) and especially detection of HCV RNA (detectable by 1 week after infection). The performance of liver biopsy for the diagnosis of acute hepatitis C is restricted to situations in which the clinical profile is consistent, although anti-HCV seroconversion or recent HCV RNA detection are not characteristic. Individuals with acute hepatitis C are those who present increased ALT, accompanied or not by clinical profile consistent with acute hepatitis, with detection of (previously undetectable) HCV RNA and seroconversion for anti-HCV. Taking into account the possibility of spontaneous viral clearance of HCV, especially reported in symptomatic individuals with acute infection, it is recommended that, in symptomatic individuals, the treatment be initiated from week 12 after symptom onset onward. In the case of asymptomatic patients with acute infection, treatment should be initiated at the time of diagnosis, since there is less probability of spontaneous viral clearance and due to the fact that late specific therapy is associated with a lower SVR rate. Both IFN formulations - conventional IFN and PEG-IFN are considered efficacious options for the treatment of acute hepatitis C. However, there has been an increasing tendency toward using PEG-IFN as a first option, especially in individuals infected with genotype 1. The option of high doses of PEG-IFN should be especially considered in situations in which short-course treatment regimens are used or in cases of infection with genotype 1. When there is an option for using the conventional IFN treatment regimen, adopting a daily induction dose, followed by reduction of the number of doses, up to the end of the treatment, seems to be the most efficacious option. There is as yet no conclusive evidence that the combination of IFN and RBV is superior to monotherapy with IFN. www.bjid.com.br 52 Therapeutic Approach to Acute Hepatitis C Therefore, as observed in studies of chronic hepatitis C with HCV genotype 1, prolonged treatment regimens are recommended in cases of acute hepatitis C with HCV genotype 1, although 24-week treatments can also be considered. In acute infections with genotype 2 or 3, short-course treatments (e.g., 12-week courses) can be feasible options. Prolonged (24-week) treatment or combined therapy with RBV can be considered for patients who do not present an early virological response in the first 4 weeks of treatment. Monotherapy regimens, with shorter duration and supervised administration of IFN, especially in groups of difficult-totreat patients, would be desirable. The ideal duration of post-treatment evaluation - clinical, biochemical (ALT levels) and virological (detection of HCV RNA) - has not yet been well established. However, durations of at least 48 weeks should be considered. References 1. Alter M.J., Kruszon-Moran D., Nainan O.V. et al. The prevalence of hepatitis C virus infection in the United States, 1988 through 1994. N Engl J Med 1999;341:556-62. 2. Jaeckel E., Cornberg M., Wedemeyer H., et al. Treatment of acute hepatitis C with interferon alfa-2b. N Engl J Med 2001;345:1452-7. 3. Kamal S.M., Ismail A., Graham C.S., et al. Pegylated interferon alpha therapy in acute hepatitis C: relation to hepatitis C virus-specific T cell response kinetics. Hepatology 2004;39:1721-31. 4. Weigrand K., Stremmel W., Encke J. Treatment of hepatitis C virus infection. World Journal of Gastroenterology 2007;13(13):1897-1905. 5. Updated US Public Health Service guideline for the management of occupational exposures to HBV, HCV and HIV and recommendations for postexposure prophylaxis. MMWR 2001;50: RR1. 6. EASL International Consensus Conference on Hepatitis C, Consensus Statement, 1999. J Hepatol 1999;30:956-61. 7. Santantonio T., Fasano M., Sinisi E., et al. Efficacy of a 24week course of peg-interferon a-2b monotherapy in patients with acute hepatitis C after failure of spontaneous clearance. J Hepatol 2005;42:329-33. 8. Kamal S.M., Moustafa K.N., Chen J., et al. Duration of peginterferon therapy in acute hepatitis C: a randomized trial. Hepatology 2006;43:923-31. 9. Calleri G., Cariti G., Gaiottino F., et al. A short course of pegylated interferon-a in acute hepatitis C. Journal of Viral Hepatitis 2007;14:116-21. 10. Wiegand J., Jäckel E., Cornberg M., et al. Long-term followup after successful interferon therapy of acute hepatitis C. Hepatology 2004;40:98-107. 11. McKiernan S.M., Hagan R., Curry M., et al. Distinct MHC class I and II alleles are associated with hepatitis C viral clearance, originating from a single source. Hepatology 2004;108-14. 12. Santantonio T., Medda E., Ferrari C., et al. Risk factors and outcome among a large patient cohort with communityacquired acute hepatitis C in Italy. Clin Infect Dis 2006;43:1154-9. 13. Afdhal N.H. The natural history of hepatitis C. Seminars in Liver Diseases 2004;24,suppl. 2:3-8. 14. Strader D.B., Wright T., Thomas D.L., Seeff L.B. AASLD Practice Guideline – Diagnosis, Management and Treatment of Hepatitis C. Hepatology 2004;39:1147-71. 15. Heller T., Rehermann B. Acute hepatitis C: a multifaceted disease. Seminars in Liver Diseases 2005;25:7-17. BJID 2007; 11 Supplement 1 (October) 16. Alberti A., Boccato S., Vario A., Benvegnù L. Therapy of acute hepatitis. Hepatology 2002;5:S195-S200. 17. Micallef J.M., Kaldor J.M., Dore G.J. Spontaneous viral clearance following acute hepatitis C infection: a systematic review of longitudinal studies. Journal of Viral Hepatitis, 2006;13:34-41. 18. Nomura H., Sou S., Tanimoto H., et al. Short-term interferonalfa therapy for acute hepatitis C: a randomized controlled trial. Hepatology 2004;39:1213-19. 19. Consensus Statements on the Prevention and Management of Hepatitis B and Hepatitis C in the Asia-Pacific Region. Journal of Gastroenterology and Hepatology 2000;15:815-41. 20. Kamal S.M., Fouly A.E., Kamel R.R., et al. Peginterferon alfa-2b therapy in acute hepatitis C: impact of onset of therapy on sustained virologic response. Gastroenterology 2006;130:632-8. 21. De Rosa F.G., Bargiacchi O., Audagnotto S., et al. Twelveweek treatment of acute hepatitis C virus with pegylated interferon-a-2b in injection drug users. Clinical Infectious Diseases 2007;45:583-8. 22. Alter M.J. Prevention of spread of hepatitis C. Hepatology 2002;36:suppl.1:593-8. 23. Mondelli M.U., Cerino A., Cividini A. Acute hepatitis C: diagnosis and management. J Hepatol 2005;42:S108-S14. 24. Corey K.E., Ross A.S., Wurcel A., et al. Outcomes and treatment of acute hepatitis C virus infection in a United States population. Clinical Gastroenterology and Hepatology 2006;4:1278-82. 25. Manns M.P., Mc Hutchison J.G., Gordon S.C., et al. Peginterferon alfa-2b plus ribavirin compared with interferon alfa-2b plus ribavirin for initial treatment of chronic hepatitis C: a randomized trial. Lancet 2001;358:958-65. 26. Fried M.W., Shiffman M.L., Reddy K.R., et al. Peginterferon alfa-2a plus ribavirin for chronic hepatitis C virus infection. N Engl J Med 2002;347:975-82. 27. Hofer H., Watkins-Riedel T., Janata O., et al. Spontaneous viral clearance in patients with acute hepatitis C can be predicted by repeated measurements of serum viral load. Hepatology 2003;37:60-4. 28. Gerlach J.T., Diepolder H.M., Zachoval R., et al. Acute hepatitis C: high rate of both spontaneous and treatmentinduced viral clearance. Gastroenterology 2003;125:80-8. 29. Davis G.I., Wong J.B., McHutchinson J.G., et al. Early virologic response to treatment with peginterferon alfa-2b plus ribavirin in patients with chronic hepatitis C. Hepatology 2003;38:645-52. 30. Firenci P. Predicting the therapeutic response in patients with chronic hepatitis C: the role of viral kinetics studies. J Antimicrob Chemother 2004;53:15-8. 31. Yamaji K., Hayashi J., Kawakami Y., et al. Hepatitis C viral RNA status at two weeks of therapy predicts the eventual response. J Clin Gastroenterol 1998;26:193-9. 32. Wiegand J., Baggish P., Boecher W., et al. Early monotherapy with pegylated interferon alpha-2b for acute hepatitis infection: the HEP-NET acute-HCV II study. Hepatology 2006;43:250-6. 33. Jaeckel E., Cornberg M., Waldemeyer H., et al. Acute hepatitis C: to treat or not to treat? Hepatology 2002;35:1538-40. 34. Management of hepatitis C: 2002. NIH Consensus Development Conference, 2002. 35. Angerami R.N., Stucchi R., Gonçales N.S.L., Gonçales Jr. F.L.G. Hepatite C aguda. II Consenso da Sociedade Paulista de Infectologia para Manuseio e Terapia da Hepatite C 2004:34-6. 36. Zekry A., Patel K., Mc Hutchison J.G. Treatment of acute hepatitis C infection: more pieces of the puzzle. J Hepatol 2005;42:293-6. 37. Focaccia R., Galante V.C., Oliveira U.B. Hepatite C – Epidemiologia. In: Tratado de Hepatites Virais. Ed Roberto Focaccia, 2ª Edição, Editora Atheneu, 2007:211-16. 38. Consenso da Sociedade Paulista de Infectologia para Manuseio e Terapia da Hepatite C, 2004. www.bjid.com.br BJID 2007; 11 Supplement 1 (October) 53 Treatment of Chronic Hepatitis C in Treatment-Naïve Patients Marcelo Simão Ferreira Federal University of Uberlândia School of Medicine; Uberlândia, MG, Brazil After the initial acute phase of infection, 50-85% of patients infected with the hepatitis C virus (HCV) develop the chronic form of the disease, which, in 20-30% of cases, will evolve to cirrhosis, liver failure or hepatocellular carcinoma, albeit after several decades. Once this infection has been established, it rarely resolves spontaneously. It is known that, during the chronic phase, the more severe forms of this viral infection can be induced by various cofactors: chronic alcoholism; coinfection with HIV or the hepatitis B virus; liver biopsy-proven steatosis (or steatohepatitis); and advanced age. The main objective for treating this disease is, therefore, to prevent the occurrence of late complications, by means of the eradication of HCV, which can be achieved in just over half of the cases treated with the currently available drugs. Treatment for Chronic Hepatitis C: Drugs and Treatment Response Patterns The currently recommended treatment for the chronic forms of hepatitis C is the combination of interferon alpha (IFN-α) and ribavirin. The former is a cytosine that is a component of the innate response of the human host. Various genes involved in the immune response are induced/stimulated by IFN-α, resulting in the activation of natural killer cells, maturation of dendritic cells, and proliferation of memory cells, as well as in the prevention of apoptosis of T cells. The hepatocellular injury seen in chronic hepatitis C is not due to the cytopathic effect of HCV. It is immunomediated by natural killer cells and CD8 T lymphocytes, which are activated by the action of IFN-α. Ribavirin is an oral nucleoside analog with antiviral effects against various pathogens (respiratory syncytial virus, arenavirus, etc.) Although its mechanism of action in HCV remains unclear, it seems that ribavirin causes the virus to mutate rapidly to forms that are more easily killed, as well as depleting intracellular adenosine triphosphate, which is essential for the synthesis of viral RNA. Immunomodulatory effects have been also attributed to this drug. The use of IFN-α was approved for hepatitis C treatment in 1991. At that time, the response to this immunotherapy was very low (< 20%). However, when the combination of ribavirin and IFN-α began to be used, the proportion of patients in whom a sustained virological response (SVR) was achieved grew to 40-45%. Even at that time, treatment responses were confirmed by detecting HCV RNA through molecular tests such as polymerase chain reaction (PCR). In this context, the infection is considered eradicated when there is an SVR, which is defined as the absence of serum HCV RNA in a sensitive test (qualitative PCR) at the end of treatment and at six months The Brazilian Journal of Infectious Diseases 2007;11 (5) Suppl. 1:53-57. © 2007 by The Brazilian Journal of Infectious Diseases and Contexto Publishing. All rights reserved. after the end of treatment. Patients who achieve an SVR almost always show a dramatic decrease in HCV RNA levels, defined as a ≥ 2 log10 drop or the absence of HCV RNA by 12 weeks after the initiation of treatment. This response is designated the early virological response (EVR) and has been widely used for treatment follow-up of patients infected with HCV genotype 1. The maintenance of the undetectable viral load status at the end of the treatment is designated the end-oftreatment response (ETR). A patient is considered recidivist when HCV RNA becomes undetectable during treatment but becomes positive again after the end of the treatment, whereas a patient is considered a nonresponder when HCV RNA levels remain stable or decrease < 2 log10 during treatment with the combined regimen. The most recent advance in the treatment of hepatitis C has been the development of long-acting, pegylated formulations of IFN-α (PEG-IFN-α), produced by the covalent addition of a polyethylene glycol molecule to the IFN-α molecule. This combination decreased absorption, reduced the clearance of the drug and increased its half-life. With this increased half-life, PEG-IFN-α can be administered in weekly doses. Two pegylated formulations have now been approved for the treatment of hepatitis C: PEG-IFN-α 2a (PegasysRoche), with a molecular weight of 40 kDa, and PEG-IFN-α 2b (PEG-INTRON; Schering-Plough), with a molecular weight of 12 kDa. Table 1 shows the names and doses of the drugs that are currently used for the treatment of chronic hepatitis C. Viral Kinetics After the Beginning of Treatment The most important objective of chronic hepatitis C treatment is HCV eradication. The introduction of treatment leads to a biphasic drop in the viral population. The speed at which the quantity of HCV drops differs among the treated patients, and the drops can therefore be classified as rapid or slow. Patients in whom there are rapid drops at the beginning of treatment more often achieve an SVR. During this rapid response phase, which generally occurs within the first 48 h of HCV treatment, the viral load decreases rapidly, which reflects the IFN-α inhibition of replication and the degradation of the drug in the serum. The HCV RNA titers begin to decline 8 to 12 h after the administration of the first IFN-α dose, and the drop ranges from 0.5 to 1.5 log10 within the first 48 h. Thereafter, the rate of viral load reduction slows, reflecting the clearance of the virus in the infected cells. The complete elimination of viral particles requires combined treatment for several months. During this second phase, ribavirin seems to play a crucial role in HCV depuration. Negative HCV viral load during treatment is generally followed by alanine aminotransferase normalization and improvement in the necroinflammatory activity in liver biopsy. www.bjid.com.br 54 Chronic Hepatitis C in Treatment-Naïve Patients Table1. Drugs used for the treatment of chronic hepatitis C Drugs Conventional interferon Alpha 2a Alpha 2b Consensus Pegylated interferon Peginterferon α 2a (40 kDa) Peginterferon α 2b (12 kDa) Nucleoside analogs Ribavirin Recommended doses 3 MU sc 3 × week 3 MU sc 3 × week 9 μg sc 3 × week 180 μg SC / week 1.5 μg/kg SC / week From 1,000 mg (≤ 75 kg) to 1250 mg (> 75 kg), oral, daily MU: million units; sc: subcutaneous. Treatment Protocols For adults diagnosed with chronic hepatitis C and presenting detectable serum levels of HCV RNA, together with persistent elevation of aminotransferases, histological evidence of progressive hepatic disease, no severe comorbidities, and no contraindications, treatment is recommended. All patients should be initially submitted to viral load quantification (quantitative PCR), HCV genotype identification (genotypes 1 to 6) and liver biopsy for the evaluation of necroinflammatory activity (intensity) and fibrosis (staging). The two most common methods for histological evaluation are the METAVIR and Ishak scoring systems, in which fibrosis is scored as absent (F0), only portal (F1), portal with septum formation (F2), hepatic with portalcentral and portal-portal bridging (F3), or cirrhosis (F4). Treatment is recommended for patients who present a score of at least F2. It must be borne in mind that liver biopsy is an invasive, costly, and potentially fatal procedure. For patients presenting HCV genotype 1, this procedure is useful for therapeutic decisions when there is no evidence of advanced fibrosis detected by other methods (ultrasound, etc.) However, for those presenting genotype 2 or 3 and high treatment response rates, liver biopsy might be unnecessary and might not influence the therapeutic decision, especially for those individuals who present persistently high levels of aminotransferases. The combination of PEG-IFN-α and ribavirin is the treatment currently recommended for patients with chronic hepatitis C; its efficacy in achieving an SVR is greater than that of the conventional treatment with IFN-α in isolation or with the combination of IFN-α and ribavirin (56% vs. 16% vs. 42%). As demonstrated in Table 1, subcutaneous PEG-IFN-a is administered weekly, and oral ribavirin is administered in two daily doses. The recommended dose of PEG-IFN-α 2a is 180 μg per week and that of PEG-IFN-α 2b is 1.5 μg/kg per week. Treatment duration and ribavirin dosing vary according to HCV genotype. Patients presenting HCV genotype 1 must be treated with either 1,000-mg or 1,250-mg doses of ribavirin (body weight ≤ 75 kg or > 75 kg, respectively) for 48 weeks. Patients infected with HCV genotype 2 or 3 must receive 800 mg of ribavirin daily for 24 weeks. In Brazil, ribavirin capsules BJID 2007; 11 Supplement 1 (October) contain 250 mg, and so it is not possible to administer the recommended 800-mg doses. Therefore, we believe it is prudent to prescribe 1000-mg doses for all patients infected with these genotypes. There is little information on the treatment against hepatitis C genotypes 4, 5, and 6 (which are rare in Brazil). It has been recommended that the 48-week treatment regimen be used in these cases. In the treatment for infection with HCV genotype 1, the possibility of achieving an SVR is based on the EVR, as previously defined. Negative results or a significant (≥ 2 log10) drop in HCV viral load by week 12 of treatment is indicative of an SVR, which appears in 65% of the patients treated with PEG-IFN-α 2a and in 72% of those treated with PEG-IFN-α 2b. However, among those who have not achieved an EVR by week 12 using either formulation, only 3% achieve an SVR. Therefore, the EVR is a strong negative predictor of the SVR. All patients who continue to present positivity (based on viral loads) at week 12 should be re-assessed at week 24, at which point a qualitative PCR should be performed. If results are negative, treatment should be maintained up to week 48. If is the results are still positive, treatment should be discontinued. For HCV genotypes 2 and 3, it is not generally recommended that molecular tests be performed during the 24-week treatment regimen. At the end of the combined treatment, regardless of the genotype, a qualitative PCR should be performed in order to evaluate the ETR. Chart 1 shows the algorithm for the follow-up of patients with chronic hepatitis C during the treatment with PEG-IFN-α and ribavirin. With the current treatment protocols, the rate at which an SVR is achieved ranges from 54% to 56% after the use of the combination of PEG-IFN-α (either formulation) and ribavirin, which is considerably higher than that seen with older conventional treatments (Chat 1). Among patients infected with HCV genotype 2 or 3, the rate at which an SVR is achieved ranges from 75% to 80%, compared with 40% to 61% among those infected with HCV genotype 1. Among patients with HCV genotype 1, the rate at which an SVR is achieved is lower in blacks (28%) than in Caucasians (52%). Other factors that influence treatment response, leading to lower cure rates, are high viral load levels (> 600,000 IU/mL), male gender, high body mass index, biopsy showing advanced fibrosis and high iron levels in the hepatic parenchyma. In patients infected with HCV genotype 2 or 3 and treated with a conventional regimen involving the combination of IFN-α (3 million units 3 times a week) and ribavirin, the rate at which an SVR is achieved can be similar to that of those treated with pegylated formulations and ribavirin. Therefore, this regimen is still recommended in Brazil (directive 863/2002). For such patients, the use of this treatment regimen for six months is less costly and better tolerated. Patients with liver cirrhosis have lower chances to eliminate HCV by means of the current treatment than do noncirrhotic patients. This might be due to the fact that the former present more advanced age, greater alterations in hepatic microcirculation (which prevent the adequate interaction between IFN and infected cells), and lower treatment www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Chronic Hepatitis C in Treatment-Naïve Patients 55 Chart 1. Chronic hepatitis treatment strategies Table 2. Drugs for the treatment of chronic hepatitis C Genotype PEG-IFN-α α (dose) Ribavirin 1 (4, 5, 6) α 2a: 180 μg/week α 2b: 1.5 μg/kg/week 1.0 g (< 75 kg) 1.25 g (> 75 kg) 48 Same as above 1.0 g 24 2, 3 Duration Treatment evolution (dose) (weeks) a) No response: ¯HCV RNA at week 12 < 2 log10 IU/mL or HCV RNA ⊕ by week 24 b) Rapid response: treat for 24 weeks if HCV RNA Θ by week 4 and low initial viral load (< 600,000 IU/mL) a) No response: uncommon b) Rapid response: treat for 12-16 weeks if HCV RNA Θ by week 4 PEG-IFN=pegylated interferon; SVR=sustained virological response; HCV=hepatitis C virus. www.bjid.com.br SVR (%) 40-61 75-80 56 Chronic Hepatitis C in Treatment-Naïve Patients compliance rates. These patients should be treated carefully due to the risk of decompensation of the disease during treatment and the worsening of pre-existing hematological parameters (leukopenia and platelet reduction). Patients with compensated cirrhosis should be treated if they present the minimum criteria to receive these medications: a) Total bilirubin < 1.5 g/dL b) Albumin > 3.4 g/dL c) Platelets > 75,000/mm3 d) Hemoglobin > 13 g/dL; neutrophils > 1500/mm3 e) Creatinine < 1.5 mg/dL f) Absence of ascites, together with hepatic encephalopathy. In patients with advanced fibrosis and treated with the PEG-IFN-α/ribavirin combination, the rate at which an SVR is achieved ranges from 37% to 50%, being higher (70%-75%) in those presenting HCV genotype 2 or 3, as well as in those with low viral loads. Side effects, especially thrombocytopenia and neutropenia, are more common in these patients. At this stage of the disease, cirrhotic patients who achieved an SVR are not totally protected against the risk of developing hepatocellular carcinoma. Two recent studies have confirmed a limited reduction in the risk of hepatocellular carcinoma in patients who successfully responded to the combined regimen; however, some degree of risk remains due to the carcinogenic effect of hepatic fibrosis. In addition, the complications of advanced liver disease, in these cases, occur less frequently, mortality is lower, and there will obviously be no re-infection of the organ after the liver transplant. In order to prevent re-infection in transplanted patients, those who are on a liver transplant waiting list and present decompensated cirrhosis have been treated with IFN-α or PEG-IFN-α and ribavirin for periods ranging from three to fourteen months. In these individuals, the rate at which an SVR is achieved has been low (from 20% to 25%), and morbidity/mortality due to the treatment have been considerably high. Bacterial infections, severe cytopenias, and even mortality have been reported during treatment. However, in those patients who responded to treatment, with the elimination of HCV, there was improvement in liver function, fewer episodes of decompensation and lower mortality. Benefits seem to be higher, as expected, for patients infected with HCV genotype 2 or 3, who traditionally respond better to the treatment regimen. Not every patient whose HCV RNA levels become undetectable during treatment achieves an SVR. In 10% of the treated patients, this molecular marker reappears in the serum during treatment, whereas in 20% of the treated patients, it reappears after the end of treatment (recurrence); in this context, HCV RNA becomes detectable a few weeks after the interruption of treatment, and aminotransferase levels again increase. Recurrence is more common with short treatment or when there is a delay in achieving negative HCV RNA results. In those patients who responded to treatment and achieved an SVR, long-term follow-up evaluations have shown that HCV RNA results remain negative in more than 95% of these patients, confirming the cure and the improvement in histological parameters. BJID 2007; 11 Supplement 1 (October) Modifications During the Course of Treatment of Chronic Hepatitis C In patients infected with HCV genotype 1, a 72-week combined regimen (PEG-IFN-α + ribavirin) may be beneficial for slow responders, who fail to present negative HCV RNA results by week 4 or week 12 of treatment (EVR). In a randomized study, 326 patients still presented positive results for the virus at week 4 of treatment. These patients received the medications for either 48 or 72 weeks. The rate at which an SVR was achieved was significantly higher in the group receiving the longer-duration regimen (45% vs. 32%; p = 0.01). In another randomized study, patients who did not achieve an EVR were submitted to either 48 or 72 weeks of treatment. The results showed that 29% (31/106) of the patients in the 72week group achieved an SVR, compared with only 17% (17/ 100) of those in the 48-week group (p = 0.04). Therefore, selected patients who present slow initial response to antiviral medications can be treated for longer periods. However, the treatment duration can be shortened if patients develop a rapid virological response (RVR), defined as presenting a response by week 4 of treatment. Various studies have demonstrated that negative results in the qualitative PCR by week 4 of treatment have a positive predictive value for SVR. Studies involving patients infected with HCV genotype 2 or 3 who achieve an RVR with the use of PEG-IFN-α 2b and ribavirin have demonstrated that treatment can be discontinued at 12 to 16 weeks after the beginning of treatment, since these patients presented SVR rates that were similar to those seen in the control group, which was submitted to the 24-week regimen. Recurrences were especially more common among patients infected with HCV genotype 2 and in those with high viral loads before treatment. In a recent randomized study, 150 patients infected with HCV genotype 2 were treated with PEG-IFN-α and ribavirin for either 16 or 24 weeks. The comparison showed that both groups presented high SVR rates (94% and 95%, respectively), indicating that patients infected with HCV genotype 2 can be treated for a shorter period of time. In a similar study, involving patients infected with HCV genotype 1, good results were demonstrated using a 24-week course of treatment. Patients achieving an RVR and a low viral load (< 600,000 IU/mL), treated for this short period of time, presented a high cure rate (89%), confirming the hypothesis that, even in patients infected with the more difficult to treat genotypes, it is possible that treatment can be shortened if patients are adequately selected regarding short-term regimens. Although further studies are necessary to confirm the efficacy of less prolonged therapies for the treatment of chronic hepatitis C, these current studies have already indicated that it is possible to use this strategy for patients achieving an RVR, significantly improving treatment compliance and quality of life of the patients, as well as reducing the frequency of side effects and increasing the cost-effectiveness ratio. Table 2 summarizes the current recommendations for the treatment of chronic hepatitis C, showing the recommended medications and doses, together with treatment durations and responses. www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Chronic Hepatitis C in Treatment-Naïve Patients Contraindications to and Side Effects of Treatment Absolute contraindications to the PEG-IFN-α and ribavirin treatment include pregnancy, breastfeeding and hypersensitivity to either drug. Relative contraindications, due to potential side effects are as follows: decompensated liver disease (jaundice, ascites, hepatic encephalopathy, severe coagulopathy, etc.); neuropsychiatric, pulmonary, cerebrovascular or coronary diseases; severe autoimmune diseases; malignant neoplasms; convulsions; and history of solid organ transplants. Patients addicted to injection drugs, as well as chronic alcoholics, should be advised to abandon the habit (for at least six months) prior to the beginning of the antiviral therapy. Patients diagnosed with anemia, leukopenia and platelet reduction should be treated with care, and hematological parameters should be closely monitored throughout the treatment. Side effects of interferon and ribavirin affect practically all of the patients. Table 3 shows these side effects, which are organized by their frequency. The most common side effects are fatigue, myalgia, psychological alterations (depression, anxiety, insomnia, and irritability) and worsening of hematological parameters (anemia, platelet reduction, and leukopenia). Ribavirin induces hemolytic anemia, which frequently requires dose reduction. In addition, ribavirin is teratogenic, which requires strict contraceptive control during treatment. Only 1% to 2% of patients will develop severe side effects, requiring the interruption of one or both medications. Recombinant erythropoietin (for anemia) and filgrastim (for neutropenia) should be routinely used for the control of treatment-induced cytopenias. Patients suffering from depression or mood disorders can use antidepressants or anxiolytics, with variable success. References 1. Hoofnagle J.H., Seef L.B. Peginterferon and Ribavirin for chronic hepatitis C. New England Journal of Medicine 2006;355:244451. 2. Strader D.B., Wright T., Thomas D.L., Seef L.B. Diagnosis, management and treatment of hepatitis C. Hepatology 2004;4:1147-71. 3. Iacobellis A., Siciliano M., Perri F., et al. Peginterferon alfa-2b and ribavirin in patients with hepatitis C virus and decompensated cirrhosis: a controlled study. Journal of Hepatology 2007;46:206-12. 4. Bruno S., Stroffolini T., Colombo M., et al. Sustained virological response to Interferon a is associated with improved outcome in HCV-related cirrhosis: a retrospective study 2007;45:579-87. 5. Schiff E.R. Emerging strategies for pegylated interferon combination therapy. Gastroenterology & Hepatology 2007;4(1):517-21. 6. Navasa M., Forns X. Antiviral therapy in HCV decompensated cirrhosis: to treat or not to treat? Journal of Hepatology 2007;46:185-8. 7. Everson G.T., Hoefs J.C., Seef L.B., et al. Impact of disease severity on outcome of antiviral therapy for chronic hepatitis C: lessons from the HALT-C trial. Hepatology 2006;44:1675-84. 8. Tan J., Lok A.S.F. Update on viral hepatitis: 2006. Current Opinion in Gastroenterology 2007;263-7. 57 Table 3. Side effects of interferon and ribavirin Side effects Interferon α (including PEG-IFN) Ribavirin Flu symptoms (greater with PEG-IFN-α 2a) Neutropenia Thrombocytopenia Depression, acute psychosis, suicide attempts Irritability Visual disorders Fatigue, myalgia Hypothyroidism/hyperthyroidism Headache Nausea, vomiting Itching Fever, weight loss Hearing loss Alopecia Pulmonary interstitial fibrosis Angina/myocardial infarction Bacterial infections (in cirrhotic patients) Hemolytic anemia Fatigue Pruritus Skin rash Sinusitis Fetal malformations Gout PEG-IFN=pegylated interferon. 9. Hadzyiannis S.J., Sette H., Morgan T.R., et al. Peginterferon a 2a and Ribavirin combination therapy in chronic hepatitis C. Annals of Internal Medicine 2004;140:346-55. 10. Pawlotsky J.-M. Current and future concepts in hepatitis C therapy. Seminars in Liver Disease 2005;25:72-83. 11. Mangia A., Santoro R., Minerva N., et al. Peginterferon alfa 2b and Ribavirin for 12 vs 24 weeks in HCV genotype 2 or 3. The New England Journal of Medicine 2005;325:2609-17. 12. Zenzem S., Buti M., Ferenci P., et al. Efficacy of 24 weeks treatment with chronic hepatitis C infected with genotype 1 and low pretreatment viremia. Journal of Hepatology 2006;44:97-103. 13. Delgard O., Bjoro K., Hellum K.B., et al. Treatment with pegylated interferon and ribavirin in HCV infection with genotype 2 or 3 for 14 weeks: a pilot stydy Hepatology 2004;40:1260-5. 14. Manns M., Waldemeyer H., Cornberg M. Treating viral hepatitis C: efficacy, side effects and complications. Gut 2006;55:1350-9. 15. Davis G.L. Monitoring of viral levels during therapy of hepatitis C. Hepatology 2002;36:5145-51. 16. Manns M.P., McHutchison J.G., Gordon S.C., et al. Peginterferon alfa 2b plus ribavirin for initial treatment of chronic hepatitis C: a randomized trial. Lancet 2001;358:958-65. 17. Hofmann W.P., Zeuzem S., Sarrazin C. Towards individualized antiviral therapy of patients infected with hepatitis C virus genotypes 2 and 3. Hepatology Reviews 2006;3:3-10. 18. Abergel A., Hezode C., Leroy V., et al. Peginterferon alpha-2b plus ribavirin for treatment of chronic hepatitis C with severe fibrosis: a multicentre randomized controlled trial comparing two doses of peginterferon alpha 2b. Journal of Viral Hepatitis 2006;13:811-20. 19. Hung C.H., Lee C.M., Lu S.N., et al. Long term effect of interferon alpha 2b plus ribavirin therapy on incidence of hepatocellular carcinoma in patients with hepatitis C virus-related cirrhosis. Journal of Viral Hepatitis 2006;13:409-14. 20. EASL International Consensus Conference on hepatitis C. Paris, 26-27 February 1999. Consensus statement. Journal of Hepatology 1999;3151:3-8. www.bjid.com.br 58 BJID 2007; 11 Supplement 1 (October) Retreatment of Hepatitis C Patients Who Previously Experienced Treatment Failure Fernando Lopes Gonçales Jr. Department of Viral Hepatitis Studies, State University of Campinas (Unicamp); Campinas, SP, Brazil In its different formulations, interferon (IFN) alpha combined with ribavirin (RBV) is the best treatment alternative for patients infected with the hepatitis C virus (HCV) [1]. In such patients, the objective is to achieve an end-of-treatment virological response – negative serum HCV ribonucleic acid (RNA) – followed by a sustained virological response (SVR), which is defined as HCV RNA negativity for six months after the suspension of treatment. The treatment regimens for HCV infection have evolved from monotherapy with conventional IFN alpha to combined therapy with IFN and RBV and, more recently, to combined therapy with pegylated IFN (PEG-IFN alpha-2a or alpha-2b) and RBV [2,3]. It is known that approximately 60% of the patients infected with HCV genotype 1 and nearly 40% of those infected with genotype 3 do not achieve an SVR when treated with the conventional regimen of IFN and RBV [2-7]. The availability of PEG-IFNs has reduced the percentage of patients experiencing treatment failure. In large international trials involving treatment-naïve patients, regimens that include the combination of PEG-IFN and RBV have produced significantly higher rates of SVR than those observed with the use of the conventional combination of IFN and RBV (5456% vs. 44-47%) [5-7]. Similar to what occurs in treatment-naïve patients, retreatment provides the patient with a new chance to achieve an SVR. Long-term studies have proven that the vast majority of the patients who achieve an SVR usually remain negative for HCV RNA for a long time [8]. The arguments in favor of retreating patients would be the possibility of eradicating HCV, reducing fibrosis, and decreasing the risk of evolving to hepatocarcinoma. With regard to the new antiviral agents, we do not know when they will be available or whether all patients will be able to wait several years to initiate retreatment. It is known that patients who experience relapse after treatment with conventional IFN, with or without RBV, respond better to retreatment with PEG-IFN alpha and RBV than do those presenting no response to treatment with conventional IFN (with or without RBV). Krawitt et al. observed an SVR rate of 55% in 66 relapsing patients who were retreated with PEGIFN alpha-2b (100-150 µg/week) and RBV (1000 mg/day), compared with only 20% in 116 previous nonresponders treated with the same regimen [9]. An SVR was observed in 53% of the relapsing patients infected with genotype 1, compared with 59% of the relapsing patients infected with genotypes 2 or 3. There were, therefore, no significant differences between these groups of patients. This was not The Brazilian Journal of Infectious Diseases 2007;11 (5) Suppl. 1:58-59. © 2007 by The Brazilian Journal of Infectious Diseases and Contexto Publishing. All rights reserved. observed in the previous nonresponders receiving retreatment. Of those, only 17% of the individuals infected with genotype 1, in comparison with 57% of the individuals infected with genotypes 2 or 3, achieved an SVR. Therefore, genotype had an influence on SVR in the previous nonresponders who underwent retreatment. Studies conducted in Brazil and involving nonresponders to IFN/RBV demonstrated SVR rates resulting from retreatment with the PEG-IFN alpha-2a + RBV regimen [10], as well as with the PEG-IFN alpha-2b + RBV regimen [11], that were higher than those reported in international studies. Of the relapsers treated, 57% [10] and 62% [11] achieved an SVR. These SVR rates in relapsers are higher than, for example, the rates of 41% and 59% obtained, respectively, in Canada (by Sherman et al.) and in France (by Moucari et al.) [12,13]. In analyzing the response in relapsers by genotype, the Brazilian researchers found that an SVR was achieved in 69-70% of those infected with genotype 3, compared with 43-44% of those infected with genotype 1 [10,11]. True nonresponders, that is, patients who never presented viral negativity during or at the end of the treatment, are the most difficult group to retreat. Some authors showed that, after previous treatment with conventional IFN, with or without RBV, relapsers have significantly higher SVR when retreated with PEG-IFN alpha-2b and RBV than do nonresponders to previous treatment (55% vs. 20%; p<0.001) [9]. The same was observed by Sherman et al., who found that, after retreatment with PEG-IFN alpha-2a and RBV, the SVR rate was 23% in nonresponders and 41% in relapsers [12]. The results of the retreatment of true nonresponders to PEG-IFN and RBV obtained by the Brazilian researchers are better than those reported in international studies [10,11]. However, it is very difficult to compare different studies with different populations of patients treated with different formulations of conventional IFN. Two large international trials (EPIC3 and HALT-C) are evaluating, in two different international cohorts, the responses to treatment with the combination of PEG-IFN and RBV in nonresponders to previous treatment with IFN and RBV [14,15]. The EPIC3 study is also evaluating the real meaning of the virological response at week 12 and its predictive value in nonresponders. The final results of both trials will soon be available. With regard to the use of induction doses of PEG-IFN in the first 12 weeks or the increase in the duration of the treatment of nonresponders, there are few studies in the literature. In a recent pilot study, researchers working in Spain observed that patients with genotype 1 HCV who were retreated for 72 weeks, with different induction doses of PEG- www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Retreatment of Hepatitis C Patients IFN alpha-2a in the first 12 weeks, achieved an SVR at a rate of 30-37%, which is practically twice as high as the SVR rate of 18% observed in those treated with normal doses of PEGIFN alpha-2a [16]. Since nonresponders to initial treatment/relapsers constitute a very heterogeneous group, it is necessary to carefully qualify and select the patients who should be retreated. The various factors that might have been responsible for the lack of a response should be modified/ nullified prior to the initiation of or even during the new treatment cycle. Currently, the patients who are considered the least likely to respond to retreatment are the true nonresponders, as well as those who are black, infected with genotype 1, presenting a high viral load, having advanced liver disease, or presenting intercurrent diseases, such as obesity, HIV coinfection, and coinfection with hepatitis B virus. Those who were previously treated with IFN as monotherapy or with the combination of IFN and RBV are more likely to achieve an SVR than are the nonresponders to the combination of PEG-INF and RBV. Those who had relapses during (breakthrough) or after the previous treatment also fare better than do those who are true nonresponders. Patients who were noncompliant with the previous treatment or those who needed to undergo IFN or RBV dose reduction due to cytopenias or other adverse effects usually respond better to retreatment than do those who received full doses. It is clear that the factors responsible for noncompliance should be corrected. In addition, medication dose reductions should be adequately addressed. In nonresponders, erythropoietin and filgrastim should be used earlier and further dose reductions should be avoided at all costs. Drug and alcohol users who, due to their dependence, did not satisfactorily complete all phases of the previous treatment might respond better to retreatment if these cofactors are removed. This also applies to the patients who did not have adequate social or cultural support. Obese patients, insulin-resistant patients, dyslipidemic patients, patients with steatosis, and patients with liver diseases, such as hemochromatosis, should be retreated, preferably after the diagnosis and treatment of these accompanying conditions have been appropriately addressed. The doses of PEG-IFN should be the same as those used in treatment-naïve patients. We recommend that, in retreatment, the doses of RBV be as high as possible. Due to the paucity of studies of large population samples, the duration of retreatment should be 48 weeks for all genotypes. At the moment, there are no conclusive data in the literature to support the use of higher doses of PEGIFN and RBV, the use of induction doses, or the extension of treatment duration to more than 48 weeks in such patients. With regard to the week-12 rule, there is strong evidence that patients who do not present HCV RNA negativity by that time are much less likely to achieve an SVR, and that their treatment should therefore be discontinued. 59 References 1. National Institutes of Health Consensus Development Conference Statement: Management of hepatitis C: 2002-June 10-12, 2002. Hepatology 2002;36:S3-20. 2. McHutchison J.G., Gordon S.C., Schiff E.R., et al. Interferon alfa-2b alone or in combination with ribavirin as initial treatment for chronic hepatitis C. N Engl J Med 1998;339:1485-92. 3. Poynard T., Marcellin P., Lee S.S., et al. Randomised trial of interferon alpha-2b plus ribavirin for 48 weeks or for 24 weeks versus interferon alpha-2b plus placebo for 48 weeks for treatment of chronic infection with hepatitis C virus. Lancet 1998;352:1426-32. 4. Poynard T., McHutchison J., Goodman Z., et al. Is an “a la carte” combination interferon alfa-2b plus ribavirin regimen possible for the first line treatment in patients with chronic hepatitis C? Hepatology 2000;31:211-8. 5. Fried M.W., Shiffman M.L., Reddy K.R., et al. Peginterferon alfa-2a plus ribavirin for chronic hepatitis C virus infection. N Engl J Med. 2002;347:975-82. 6. Manns M.P., McHutchison J.G., Gordon S.C., et al. Peginterferon alfa-2b plus ribavirin compared with interferon alfa-2b plus ribavirin for initial treatment of chronic hepatitis C: a randomised trial. Lancet 2001;358:958-65. 7. Hadziyannis S.J., Sette H., Morgan T.R., et al. Peginterferonalpha2a and ribavirin combination therapy in chronic hepatitis C: a randomized study of treatment duration and ribavirin dose. Ann Intern Med 2004;140:346-55. 8. M a r c e l l i n P. , B o y e r N . , G e r v a i s A . , e t a l . L o n g - t e r m histologic improvement and loss of detectable intrahepatic HCVRNA in patients with chronic hepatitis C and sustained r e s p o n s e t o i n t e r f e r o n - a l p h a t h e r a p y. A n n I n t M e d 1 9 9 7;127:875-81. 9. Krawitt E.L., Ashikaga T., Gordon S.R., et al. Peginterferon alpha-2b and ribavirin for treatment-refractory chronic hepatitis C. J Hepatol 2005;43:243-9. 10. Parise E., Cheinquer H., Crespo D., et al. Peginterferon alfa-2 a (40 KD) (PEGASYS) plus ribavirin (COPEGUS) in retreatment of chronic hepatitis C patients, nonresponders and relapsers to previous conventional interferon plus ribavirin therapy. Braz J Infect Dis 2006;10:11-6. 11. Gonçales F.L. Jr., Vigani A., Gonçales N., et al. Weight–based combination therapy with peginterferon alpha-2b and ribavirin for naïve, relapser and non-responder patients with chronic hepatitis C. Braz J Infect Dis 2006;10:311-6. 12. Sherman M., Yoshida E.M., Deschenes M., et al. Peginterferon alfa-2a (40KD) plus ribavirin in chronic hepatitis C p a t i e n t s w h o f a i l e d p r e v i o u s i n t e r f e r o n t h e r a p y. G u t 2006 ;55:1631-8. 13. Moucari R., Ripault M.P., Oules V., et al. High predictive value of early viral kinectics in retreatment with peginterferon and ribavirin of chronic hepatitis C patients non-responders to standard combination therapy. J Hepatol 2007;46:596-604. 1 4 . Shiffman M.L., Di Bisceglie A.M., Lindsay K.L., et al. Peginterferon alfa-2a and ribavirin in patients with chronic hepatitis C who have failed prior therapy. Gastroenterology 2004;126:1015-23. 1 5 . Poynard T., Schiff E., Terg R., et al. Sustained virologic response (SVR) in the EPIC3 trial: week 12 virology predicts SVR in previous interferon/ribavirin treatment failures receiving Peg-Intron/Rebetol weight based dosing. J Hepatol 2005;42 (Suppl. 2):40. 1 6 . Diago M. Peginterferon alfa-2a (40kd) (Pegasys®) and ribavirin (Copegus®) in patients infected with HCV genotype 1 who failed to respond to interferon and ribavirin: final results of the Spanish high-dose induction pilot trial. Presented at the 55 th AASLD; October 29-November 2, 2004; Boston, MA. www.bjid.com.br 60 BJID 2007; 11 Supplement 1 (October) Maintenance Treatment for the Modulation of Liver Fibrosis Evaldo Stanislau Affonso de Araújo and Antonio Alci Barone Laboratory of Hepatitis, LIM 47 DMIP-HC-FMUSP; São Paulo, SP, Brazil In many cases, the evolution of chronic infection with the hepatitis C virus (HCV) is favorable. However, progression to liver fibrosis is a common phenomenon and can lead to liver cirrhosis. Phenomena associated with liver fibrosis have been previously reviewed and are more related to the host than to viral factors [1]. In brief, the onset of fibrosis is caused by the activation of stellate cells, which acquire the shape of myofibroblasts and become the source of collagen deposition, as well as of protein matrix formation. Activated stellate cells rapidly undergo apoptosis, after which the collagen matrix is degraded and removed by the activity of metalloproteinases. There is a complex balance between the procollagen and antifibrotic factors, although the mechanisms of fibrosis regression are not fully understood [2]. However, it appears that the tissue inhibitor of metalloproteinase-1 [3] and stellate cell apoptosis [4] are crucial to maintaining this balance. The mechanism through which HCV triggers fibrosis is little understood. Apparently, hepatocyte infection triggers a state of oxidative stress and induces inflammatory cell recruitment. These phenomena lead to the activation of stellate cells and collagen deposition. In addition, HCV proteins directly activate stellate cells [4]. Liver cirrhosis and the preceding transition stage are marked phenomena in the clinical evolution of patients and have been associated with morbidity/mortality due to chronic hepatitis C. A study involving a cohort of patients chronically infected with HCV, monitored from 1991 onward, with a mean infection period of 22 years, provided evidence that, five years after the diagnosis of cirrhosis (Ishak score ≥ 4), the survival rate was 80% [5]. This rate decreased to 19% after the first hepatic decompensation [5]. In the multivariate analysis, treatment with the combination of conventional or pegylated interferon alpha with ribavirin was found to be favorably associated with survival. In other words, the treatment was associated with the most important outcome: survival! In addition, subjects presenting a virological response, whether sustained or not, presented better evolution when compared to nonresponders. This fact suggests that even transitory negative viremia levels are a favorable phenomenon and may imply lower structural alterations and activity in the liver [5]. One relevant aspect of this study was that the interruption of alcohol consumption from the moment of diagnosis on prevented the previous ingestion from having an unfavorable influence on the disease progression. This fact reinforces the importance of alcohol abstinence for patients with hepatitis C. Poynard et al. [6] retrospectively analyzed data from 3,010 patients treated with either conventional or pegylated The Brazilian Journal of Infectious Diseases 2007;11 (5) Suppl. 1:60-63. © 2007 by The Brazilian Journal of Infectious Diseases and Contexto Publishing. All rights reserved. interferon alpha, using various therapeutic regimens . The authors concluded that the treatment with pegylated interferon and ribavirin resulted in significant improvement in histology, inflammatory activity and structural alterations (fibrosis). There was improvement in the ‘fibrosis progression rate’, which is a valid concept, although reproducibility was jeopardized by the risks of sample variation. The least worsening of fibrosis was found in the optimized group receiving pegylated interferon and ribavirin (8%), and greatest degree of such worsening was found in the group receiving interferon for 24 weeks (23%) [6]. In general, fibrosis stabilized or improved even in those patients not achieving a sustained virological response (SVR). However, sample variations, a high percentage of patients with mild initial fibrosis (over 70% classified as F1), lack of paired biopsy results from all participants, lack of a control group, and principally, a relatively short follow-up period (20 months between biopsies, on average) were limitations of this analysis. Nevertheless, Poynard et al. [6] addressed the concept of cirrhosis ‘reversion’, or as the authors designated it, the ‘reversible cirrhosis stage’. This group was composed of young patients, whose structural staging changed, regressing from F4. This phenomenon occurred in 75 (49%) of the 153 cirrhotic patients [6]. The authors postulated that this stage of fibrosis would still be ‘easily’ reversed. Some of the factors that were found to be associated with the regression of fibrosis after treatment are, obviously, the initial degree of fibrosis, minimal baseline activity, achieving an SVR, being less than 40 years of age, initial viral load lower than 3.5 million copies/mL, and (a new concept at that time) body mass index (BMI) < 27 kg/m2 [6]. In another relevant meta-analysis, Cammà et al [7,8]. evaluated three randomized clinical studies comprising 1441 patients, paired biopsies being available for 1013 (70.3%). Similarly to Poynard et al. [6], these authors demonstrated improvement in hepatic fibrosis in the patients treated with pegylated interferon alpha-2a, when compared to those treated with conventional interferon, who achieved an SVR or even experienced recurrence. However, there was no improvement among nonresponders. The authors also found that a BMI higher than 30 kg/m2 was associated with the worsening of hepatic fibrosis. However, in a more representative sample – 447 (44%) of 1013 cirrhotic patients – no ‘regression of cirrhosis’ was found. Only 33% of the cirrhotic patients presented improvement in fibrosis. However, the observation period between biopsies was also short. Finally, high alanine aminotransferase level was another factor that was associated with histological improvement. More recently, Di Marco et al. [9] prospectively evaluated cirrhotic patients with portal hypertension and no previous decompensation who received pegylated interferon alpha-2b (1.0 μg/kg/week) with or without www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Maintenance Treatment for Liver Fibrosis ribavirin (0.8 g/day). Patients infected with genotype 2 or 3 showed rapid response, with negative viral RNA results by week 4 of treatment, and achieved an SVR. However, only 10 of the patients infected with genotype 1 or 4 achieved an SVR. Nevertheless, for this subgroup of patients, the predictability of early response by week 12, but not by week 4, was also valid, and high baseline viral load was a negative predictive marker for SVR. The occurrence of cytopenias was high; however, stimulating factors were not used with these patients. If stimulating factors had been available, the SVR rate associated with treatment maintenance would have certainly been better. Treatment compliance is even more critical for this group of patients and was associated with SVR [9]. Finally, it was clear that SVR is associated with better evolution. Only 6% of the patients achieving an SVR deteriorated, compared with 38% of the nonresponders [9]. Bruno et al. [10] conducted a retrospective multicenter study comprising 920 patients with compensated cirrhosis who were treated with conventional interferon alpha (from 3 to 6 million IU/3 times a week) for a year. Similarly to the prospective study conducted by Di Marco et al. [9], the authors demonstrated benefits for those who achieved an SVR, with a reduction in the risk of decompensation, occurrence of hepatocellular carcinoma (HCC) and death. They also found that a platelet count of 109,000/mL is an independent predictor of decompensation of the liver disease [10]. This factor is extremely relevant for clinical practice and could be used as a cut-off point in order to define the use of more aggressive measures, or even those that still need to be validated. Specifically regarding HCC, Cammà et al. [7,8] in 2001, pointed out that the benefit of the reduction in the incidence of HCC, albeit modest, is also more relevant among those who achieve an SVR after being treated with interferon alpha. Other authors have also reproduced data regarding the effective and tolerable treatment [11], fewer complications, reduction or negative incidence of HCC among cirrhotic patients monoinfected with HCV who achieved an SVR after the treatment with interferon alpha [12-14], and even histological response of patients co-infected with HIV [12]. Finally, even patients with severe cirrhosis under individualized, ascending-dose regimens benefit from treatment and can achieve an SVR, albeit a modest one [15-17]. Despite the innumerable benefits of the treatment with interferon alpha, we must emphasize that surveillance regarding the incidence of HCC is highly recommended, even in cirrhotic patients who have achieved an SVR [10]. There is clear evidence of improvement in fibrosis (a quantitative decrease, as well as functional gains such as lower portal hypertension) [3], greater survival, potential decrease in the incidence of HCC and hepatic complications, and even ‘cirrhosis reversion’ [6], even for nonresponders [6] or for those who presented recurrence [5-8]. Therefore, we should ask why we do not treat even those patients with more severe cirrhosis [15-17], and why we do not use the treatment with interferon alpha for objectives other than the virological. 61 In order to answer these questions, it is initially important that recent knowledge on viral kinetics and the concepts of treatment individualization – dose and duration –be considered in the therapeutic decision-making, and that the same concepts are not clearly validated for cirrhotic patients. Therefore, if our goal is to achieve an SVR, extending the treatment of nonresponders to at least 24 weeks of ‘ideal’ treatment is considered ‘futile’ [1]. However, is this true from a histological point of view, or from the perspective of the need to modulate the natural history of the disease? In order to answer these questions and in view of the previously described evidence, physicians began to consider the use of maintenance treatment with interferon alpha. However, for a conclusive analysis, a prospective evaluation would be ideal, since differences in methodologies would greatly affect the quality of the results [17]. Three principal studies (Table 1) have addressed this issue: the Hepatitis C Antiviral LongTerm Treatment against Cirrhosis (HALT-C) trial, conducted by the NIH; the Evaluation of Peg-Intron in Control of Hepatitis C Cirrhosis (EPIC)3 trial; and the Colchicine versus Peg-Interferon Long-Term (COPILOT) trial. Other, smaller, studies, such as the PROFIC-C trial, have also addressed this issue [18]. The COPILOT study evaluates patients with fibrosis classified as greater than Ishak 3 and previous nonresponders to interferon/ribavirin or pegylated interferon/ribavirin, comparing, in two branches, colchicine to pegylated interferon alpha-2b at 0.5 μg/kg/week. Preliminary analyses after a twoyear follow-up period revealed that the group using interferon presented significantly fewer hepatic complications, especially portal hypertension and upper gastrointestinal tract bleeding [6]. Alterations in HCV quantification were minimal. The EPIC3 trial has yet to produce preliminary results. However, the HALT-C study has provided a consistent amount of information. Nevertheless, data regarding the main objective of the study, fibrosis modulation, are still unavailable but should be presented at the upcoming congress of the American Association for the Study of Liver Diseases (AASLD; Afdhal, personal communication). Using the available results from HALT-C, Everson et al. [15], in 2006, emphasized the need to ‘optimize’ the treatment of cirrhotic patients, for whom the SVR rate was lower, regardless of platelet counts or the need to reduce interferon doses – or even the influence of previous treatment response. Therefore, cirrhosis is a determining factor of a lower SVR rate. Among the therapy ‘optimization’ measures that are currently available, should we consider treatment prior to the establishment of cirrhosis? What would be the criteria? We still cannot answer that, but if we consider some preliminary results of new therapies, we can predict that interferon alpha will still be the backbone of hepatitis treatment for many years. Therefore, we should certainly attempt to gain a better understanding of the potential of these treatments and use them wisely. Among the minor studies, Erhardt et al. preliminarily showed that the maintenance of a 0.35-1.0 μg/kg/week dose www.bjid.com.br 62 Maintenance Treatment for Liver Fibrosis BJID 2007; 11 Supplement 1 (October) Table 1. Maintenance studies with pegylated interferon Study HALT-C COPILOT EPIC3 Disease stage Ishak 4-6 CTP ≤ 6 1400 Placebo, IFN-α, Peg-IFN-α-2a (90 μg) Peg-IFN á-2b (0.5 μg) 3.5 years Ishak 3-6 CTP ≤ 7 800 Peg-IFN-α-2b (0.5 μg) METAVIR 2-4 CTP ≤ 6 1700 (700 cirrhotic) 4 years 3-5 years Patients (n) Treatment arms Colchicine (0.6 mg bid) Duration HALT-C=Hepatitis C Antiviral Long-Term Treatment against Cirrhosis (trial); COPILOT= Colchicine versus Peg-Interferon Long-Term (trial); EPIC3=Evaluation of Peg-Intron in Control of Hepatitis C Cirrhosis (trial) 3; CTP=Child-Turcotte-Pugh score; IFN=interferon; Peg=pegylated. of pegylated interferon alpha-2b significantly reduced, after 48 weeks, the incidence of HCC and complications due to cirrhosis when compared to the control group. In a similar study, Kaiser et al. [19] demonstrated that, in the intervention group, the fibrosis score dropped from 3.58 to 2.59 after 18 months of treatment, and to 2.36 by six months after the end of treatment. In the control group, the fibrosis score increased from 3.88 to 4.07 and to 4.79 by the same time points. Therefore, monotherapy with lower doses of pegylated interferon alpha2b effectively reduced and modulated hepatic fibrosis. The mechanism for the improvement induced by interferon is unknown and is certainly multifactorial. The elimination of the triggering agent is undoubtedly crucial, but, since even patients who suffer recurrence get better, it is possible that interferon alpha has an intrinsic antifibrotic effect, as well as inhibiting the activation of stellate cells [4]. Other possible approaches that are positively associated with the improvement of hepatic fibrosis in nonresponders – or as a coadjuvant – are the use of renin-angiotensin inhibitors (inhibiting the activation of stellate cells) and the control of metabolic syndrome [4]. Other substances that are potentially active against hepatic fibrosis are interleukin-10 and the natural herb known as Sho-saiko-to [4]. The maintenance treatment with ribavirin is clearly a discarded alternative since there are no positive effects [20]. Finally, the use of controlled phlebotomies may be an alternative in selected cases. Excessive iron in the hepatocytes, increased by the effect of HCV, has been associated with greater tissue damage [20]. The field of antifibrogenesis is in full development and has been recently reviewed in an AASLD symposium [3]. Considering all the reviewed aspects, we should finally evaluate who would benefit from the maintenance treatment. These would be the patients with extensive fibrosis (F3/F4) or cirrhosis that does not respond to the optimal standard treatment. In view of the risk of disease progression and the onset of HCC, as well as the body of evidence available, simply monitoring the progression of the disease is not an acceptable approach, neither for the patient nor for the physician [20]. Therefore, what is the ideal dose and how long should we maintain the treatment with interferon? The period has yet to be defined, and it may be indefinite (or until the appearance of a definitely efficacious, safe therapy). Biopsy monitoring every two years in association with HCC and screening for esophageal varices are acceptable standards, although normalization of alanine aminotransferase levels and HCV reduction will rarely be seen [20]. The advent of noninvasive methods of monitoring fibrosis [21,22] has transformed maintenance into an even more alternative strategy. Regarding the adopted dose, in ongoing and published studies, pegylated interferon alpha has been used in smaller-thanhabitual doses, once a week. Tolerance and safety have proven adequate. Doses from one-third to one-half of the standard size seem to be satisfactory for hepatic fibrosis modulation. However, definitive results, which are still unavailable, could alter this perception. In summary: 1. Hepatic fibrosis is a potentially reversible phenomenon, which can even provide functional benefits. 2. Even for cirrhotic patients, treatment with interferon alpha is possible, efficient and safe, although it is less efficacious if we consider the SVR. 3. Treatment with pegylated interferon alpha, even when it is ineffective, is associated with greater survival, fewer complications and lower incidence of HCC. 4. Interferon alpha has a modulatory effect on hepatic fibrosis. 5. Maintenance treatment with reduced doses of pegylated interferon alpha proved to be effective in modulating hepatic fibrosis as well as in altering the natural history of the disease. 6. Antifibrogenesis is a field that is still in development, and, in addition to interferon alpha, other measures can be adopted, such as control of metabolic syndrome and the use of renin-angiotensin inhibitors. Considering the reviewed aspects, together with the facts that the number of antiviral therapies currently in development is smaller than expected, and that none of those are yet clinically available, as well as the fact that they will still need to be combined with interferon alpha, I believe that the optimization of interferon alpha will continue to be crucial. A pharmaco-economic analysis must be certainly considered as a supporting tool in the collective decision-making. However, considering the physician-patient relationship, some reflections are pertinent and should be debated by the Hepatitis Committee of the Brazilian Society of Infectology. www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Maintenance Treatment for Liver Fibrosis Therefore, we propose the following: Individualization The dose and duration of treatment with the combination of pegylated interferon and ribavirin should be defined based on patient body weight and on the early response during therapy for patients with minimal lesions (F1) who nevertheless are at risk for progression: moderate peri-portal activity, from 40 to 60 years of age and comorbidities (nonalcoholic steatohepatitis, HIV infection or metabolic syndrome). Maintenance Treatment Considering maintenance treatment with pegylated interferon alpha for patients with structural lesions greater than F3 or portal hypertension signs (esophageal varices, enlarged spleen, dilation of vessels) or platelet counts lower than 110,000/mm3, classified as Child-Pugh class A or B, no evidence of severe or potentially uncontrollable decompensation, with no HCC, no recurrence, partial responders or nonresponders to pegylated interferon alpha in association with ribavirin (or only to interferon when there are contraindications regarding ribavirin) whose compliance can be confirmed for at least 12 weeks. This treatment should also be considered for patients who present contraindications regarding the use of full doses. Child-Pugh class C patients could be treated in specialized centers when on a transplant waiting list. Proposed regimen*: subcutaneous pegylated interferon alpha-2b at 0.5-1.0 μg/kg/week; subcutaneous pegylated interferon alpha-2a at 90 μg/ week. Duration*: at least 24 months, indefinite, or until the appearance of a definitely efficacious, safe antiviral therapy. Follow-up evaluation: Monitoring the onset of complications (HCC, digestive tract hemorrhage, encephalopathy, etc.) *Dose schedule and duration were suggested based on preliminary data and should be reassessed depending on the results of ongoing studies. References 1. Hoofnagle J.H., Seef L.B. Peginterferon and ribavirin for chronic hepatitis C. New England Journal of Medicine 2006;355(23):2444-51. 2. Arthur M.J.P.Reversibility of liver fibrosis and cirrhosis following treatment for hepatitis C.Gastroenterology 2002;122(5):15258. 3. Friedman S.L., Rockey D.C., Bissel M. Hepatic fibrosis 2006: report of the third AASLD single topic conference. Hepatology 2007;45:242-9. 63 4. Bataller R., Brenner D.A. Liver fibrosis. The Journal of Clinical Investigation 2005;115:209-18. 5. Lawson A., Hagan S., Rye K., et al. The natural history of hepatitis C with severe hepatic fibrosis. Journal of Hepatology 2007;47:37-45. 6. Poynard T., McHutchinson J., Manns M., et al. Impact of pegylated interferon alfa-2b and ribavirin on liver fibrosis in patients with chronic hepatitis C. Gastroenterology 2002;122:1303-13. 7. Cammà C., Di Bona D., Schepis F., et al. Effect of peginterferon alfa-2a on liver histology in chronic hepatitis C: a meta-analysis of individual patient data. Hepatology 2004;39:333-42. 8. Cammà C., Giunta M., Andreone P., Craxì A. Interferon and prevention of hepatocellular carcinoma in viral cirrhosis: an evidence-based approach. Journal of Hepatology 2001;34:593602. 9. Di Marco V., Almasio P.L., Ferraro D., et al. Peg-Interferon alone or combined with ribavirin in HCV cirrhosis with portal hypertension: a randomized controlled trial. Journal of Hepatology 2007 (in press). 10. Bruno S., Stroffolini T., Colombo M., et al. Sustained Virological Response to interferon-á is associated with improved outcome in HCV-related cirrhosis: a retrospective study. Hepatology 2007;45:579-87. 11. Helbling B., Jochum W., Stamenic I., et al. HCV-Related advanced fibrosis/cirrhosis: randomization controlled trial of pegylated interferon á-2a and ribavirin. Journal of Viral Hepatitis 2006;13:762-9. 12. Sarmento-Castro R., Horta A., Vasconcelos O., et al. Impacto f peginterferon alpha-2b and ribavirin treatment on liver tissue in patients with HCV or HCV-HIV co-infection. Journal of Infection 2007;54:609-16. 13. Veldt B.J., Saracco G., Boyer N., et al. Long term clinical outcome of chronic hepatitis C patients with sustained virological response to interferon monotherapy. Gut 2004;53:1504-8. 14. Coverdale S.A., Khan M.H., Byth K., et al. Effects of interferon treatment response on liver complications of chronic hepatitis C: 9-year follow-up study. American Journal of Gastroenterology 2004. 15. Everson T.G., Hoefs J.C., Seef L.B., et al. Impact of disease severity on outcome of antiviral therapy for chronic hepatitis C: lessons from the HALT-C Trial. Hepatology 2006;44:1675-84. 16. Everson T.G., Trotter J., Forman L., et al. Treatment of advanced hepatitis C with low accelerating dosage regimen of antiviral therapy. Hepatology 2005;42:255-62. 17. Everson G.T. Maintenance Interferon for chronic hepatitis C: more issues than answers? Hepatology 2000;32:436-8. 18. Schuppan D., Krebs A., Bauer M., Hahn E.G. Hepatitis C and liver fibrosis. Cell Death and Differentiation 2003;10:S59-S67. 19. Kaiser S., Hass H., Luize B., et al. Long term, low dose treatment with pegylated interferon alfa 2b leads to a significant reduction in fibrosis and inflammatory score in chronic hepatitis C nonresponder patients with fibrosis or cirrhosis.41st EASL, 2006. 20. Kelleher T.B., Afdhal N.H. Maintenance therapy for chronic hepatitis C. Current Gastroenterology Reports 2005;7:50-3. 21. Afdhal N.H., Kowdley K.V., Llovet J.M. CCO Independent Conference Coverage of the 2007 Annual Meeting of EASL. Interim analisys: peginterferon alfa-2b maintenance therapy may reduce incidence of HCV-related HCC, Cirrhosis complications, 2007. 22. Afdhal N.H., Nunes D. Evaluation of liver fibrosis: a concise review. American Journal of Gastroenterology 2004;1160-70. www.bjid.com.br 64 BJID 2007; 11 Supplement 1 (October) Treatment of Patients Infected with Hepatitis C Virus and Presenting Extrahepatic Manifestations Fátima Mitiko Tengan1 and Antonio Alci Barone University of São Paulo School of Medicine; São Paulo, SP, Brazil HCV and Mixed Cryoglobulinemia Antiviral treatment should be performed with the same medications (standard or pegylated interferon alpha (PEGIFN-α, with or without ribavirin) and similar regimens, until additional controlled studies provide further information on the treatment of mixed cryoglobulinemia (MC) related to infection with the hepatitis C virus (HCV). Data regarding antiviral treatment of MC (Table 1) show that this therapeutic approach should be the first choice due to the antiproliferative and immunomodulatory effects of IFN and the usefulness of antiviral treatment, as demonstrated in most studies. In addition, the strict correlation between virological and clinical response, as well as the positive effect of inhibiting viral replication in the expanded B-cell clones, which is considered the pathogenic basis of MC, are reasons to make this choice. However, IFN-α can also trigger or worsen autoimmune diseases [6,7]. Renal insufficiency and neuropathies can occur or be worsened, and ulcer cicatrization may be prolonged. Therefore, treatment with IFN-α should be restricted to symptomatic patients, with or without renal involvement, after the careful evaluation of clinical and laboratory characteristics regarding autoimmunity during this period. In comparison with the antiviral treatment of chronic hepatitis C, the antiviral treatment of MC is more complex due to various reasons, such as the lack of standardized treatment protocols, the higher incidence of recurrence and the contraindications to antiviral treatment (old age, severe hepatic disease, acute nephritis and disseminated vasculitis). In addition, the interpretation of laboratory findings seems to be more complex than in chronic HCV infection. In fact, biochemical markers of MC response (cryocrit, rheumatoid factor or complement activity) can be more independent of the virological response than are alanine aminotransferase (ALT) levels. At the moment, antiviral treatment is suggested as the treatment of choice for this condition, even when there is no indication of hepatic pathology. Patients with apparently benign manifestations of the disease (palpable purpura, arthralgia and mild fatigue) should not be treated or can be symptomatically treated with nonsteroidal anti-inflammatory drugs. Special attention must be given to the treatment of patients with severe MC (with acute nephritis and disseminated vasculitis).For these cases, the data are insufficient to guarantee the safety of IFN administration, and we therefore strongly suggest a cautious approach. It is preferable to use an alternative therapeutic approach to all patients for whom antiviral treatment is contraindicated or not tolerated, as well as for those who did not respond to previous treatment. Possible alternatives include the use of corticosteroids, immunosuppressants, plasmapheresis and a hypo-antigenic, or low-antigen-content, diet [8]. Treatment should be individualized for each patient according to the severity of clinical symptoms, considering other factors involved (age, comorbidities, etc.) and for a limited period of time (weeks or months) until the remission of symptoms. Any therapeutic approach must be avoided for clinically asymptomatic patients. Table 1. Therapeutic regimens used for mixed cryoglobulinemia associated with hepatitis C virus infection Treatment (months) Treatment duration response End-of-treatment Sustained virological response 3 M IU IFN 3×/week + RBV 27 NR or 3 M IU IFN 3×/week Relapsers + RBV 18 3 M IU IFN 3×/week or PEG-IFN + RBV 9 1.5 μg/kg/week PEG-IFN + RBV 18 1.0 μg/kg/week PEG-IFN + RBV 6 78% NA 12 85% NA ≥ 18 NA 70% Author Year Number Zuckerman [1] 2000 9 NR Mazzaro [2] 2003 Alric [3] 2004 Cacoub [4] 2005 Mazzaro [5] 2005 ≥ 10 12 88% 89% 44% CS: corticosteroid; NR: Nonresponders; IFN: interferon; PEG: pegylated; RBV: Ribavirin; NA: not available. The Brazilian Journal of Infectious Diseases Publishing. All rights reserved. 2007;11 (5) Suppl. 1:64-68. © 2007 by The Brazilian Journal of Infectious Diseases and Contexto www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Treating Extrahepatic Manifestations of Hepatitis C Prior to the identification of HCV, corticosteroid therapy was the treatment of choice for MC, since corticosteroids, even in small doses, control most of the symptoms. However, corticosteroids can favor HCV replication, cause various side effects and not induce significant changes in the cryocrit or in the natural history of the disease. Cytostatic, immunosuppressive drugs (e.g., cyclophosphamide, chlorambucil and azathioprine) are mainly used, in combination with plasmapheresis, when there is no response to corticosteroids and during acute phases of MC (acute nephritis evolving to renal insufficiency and hyperviscosity syndrome). Various studies showed that rituximab (anti-CD20 antibody, a specific B-cell surface antigen) is efficacious for most patients with MC, with the significant improvement or resolution of MC – particularly skin lesions – and regression of clonal expansion of B cells [9]. Plasmapheresis is indicated for the removal of circulating cryoglobulins and immunocomplexes. Due to its efficacy and fast action, plasmapheresis is especially recommended in the presence of acute manifestations (cryoglobulinemic nephritis, severe sensorymotor neuropathies, skin ulcers and hyperviscosity syndrome). Combined with cyclophosphamide, it has been shown to effectively reduce the rebound effect at the end of apheresis. The low-antigen-content diet has low macromolecule content with high antigenic properties, resulting in more efficient removal of cryoglobulins by the reticuloendothelial system. This diet can improve the minor manifestations of the disease (purpura, arthralgia and paresthesias) and is generally prescribed during the initial phase of the disease. HCV and Lymphoma The inclusion of antiviral treatment seems to be rational in therapeutic regimens for non-Hodgkin’s lymphomas (NHLs) and HCV infection. This approach is supported by recent studies on low-grade lymphomas [10], and, in particular, on marginal zone lymphomas [11,12]. Vallisa et al. [10] treated 13 patients diagnosed with concomitant low-grade NHL-B and HCV infection, characterized by an indolent evolution, with PEG-IFN and ribavirin. A hematologic response was seen in most patients (complete and partial response, 75%), and this was strongly associated with the clearance or reduction of HCV viral load in serum, after the treatment, which proved to be useful for treating this pathology. Hermine et al. [11] reported that patients with concomitant HCV infection and splenic lymphoma with villous lymphocytes presented complete remission after being treated with IFN. The inclusion of a control group with patients diagnosed with the same disease but presenting no HCV infection demonstrated that, unlike the patients with HCV infection, the HCV-negative patients did not respond to the treatment with IFN. Similarly, remission of polyclonal proliferation in response to the antiviral treatment proved to be clearly associated with virological response [13]. 65 Although antiviral response seems to be an attractive tool for low-grade NHL and positive HCV, chemotherapy might be necessary for intermediate- or high-grade NHL, and the antiviral treatment can be maintained after chemotherapy [14]. The use of rituximab in NHL associated with HCV, alone or in combination (with antiviral treatment or with chemotherapy), seems very promising, especially for low-grade NHL [15-17]. Despite the limited number of described cases, it is reasonable to consider rituximab a safe and efficacious treatment for indolent B-cell lymphomas accompanied by HCV infection. HCV and the Presence of Autoantibodies From a clinical point of view, the major concern is represented by the use of IFN. It has been reported that the administration of IFN can have a negative effect on autoimmune hepatitis. Significant increases in ALT activity – even if transitory, corrected with corticosteroids and not associated with deterioration of liver function – have also been reported in HCV/anti-liver-kidney-microsomal type 1 (anti-LKM1)-positive cases treated with IFN [18,19]. Initial treatment for CS is recommended when there are high antibody titers (≥ 1:320), high globulin titers, anti-LKM1 antibodies, anti-human microsomal cytochrome P450 (CYP) IID6 257-279 antibodies and interface hepatitis with various plasmocytes. In the case of initial treatment with IFN, rigorous monitoring of ALT levels is suggested, especially in patients who are anti-LKM1-positive. HCV and Sjogren’s Syndrome In one study, 12 patients diagnosed with concomitant Sjögren’s syndrome and HVC infection were treated with IFN alone or with the combination of IFN and ribavirin [20]. Half of the patients presented improvement of the dry syndrome using the associated regimen, but none responded to IFN in isolation. Various patients presented adverse immunologic events during treatment. HCV and Arthritis Treatment decisions must be made case by case. Etiologic treatment with IFN-a and ribavirin is recommended when there is hepatic or systemic involvement, since it can occasionally induce or worsen autoimmune disturbances. The treatment leads to a significant improvement in HCV-related arthritis, even without a complete biochemical or virological response. Cryoglobulinemia-related arthritis generally responds to antiviral treatment. Considering there are few data available at the moment, the usually non-aggressive evolution of HCVrelated arthritis does not justify the use of antiviral medications as a standard treatment. HCV and Porphyria Cutanea Tarda Treatment with IFN-a seems to be less efficacious in patients with concomitant chronic HCV infection and porphyria cutanea tarda than in those with chronic HCV infection alone. [21]. The disease also responds to iron www.bjid.com.br 66 Treating Extrahepatic Manifestations of Hepatitis C depletion by phlebotomy, which can be performed prior to antiviral treatment. However, porphyria can be triggered in genetically predisposed patients treated with the association of IFN-a and ribavirin, as a consequence of hemolysis induced by ribavirin, which causes an increase in serum iron levels [22]. HCV and Lichen Planus Doutre et al. [23] reported improvement in lesions of two patients treated with IFN-α. Other authors [24-28] also reported that lesions disappeared during treatment with IFNá for several months after the end of treatment. Protzer et al. [29] reported oral and cutaneous lichen planus (LP) exacerbations during the treatment with IFN-α. Treatment was discontinued because local measures did not improve the lesions. It is generally recommended that stricter control measures be taken when patients with previous manifestations of LP receive IFN. There are no detailed reports on the effect of the combination of IFN and ribavirin in patients with concomitant LP and HCV infection. HCV and Thyroid The treatment with IFN-α may trigger the formation of autoantibodies in patients with HCV and exacerbate thyroid dysfunction in patients with pre-existing antibodies [30-33]. Changes are generally detected after three months of treatment and disappear when treatment is discontinued [34]. In severe cases, treatment must be discontinued, particularly in patients with hypothyroidism. Alternatively, in patients previously receiving thyroid medications, it can be useful to increase the dose during the antiviral treatment [35]. Low antibody titers are not an indication for treatment discontinuation. The possibility of good treatment control generally allows the continuity of antiviral treatment. Prior to treatment, levels of thyroid hormones, including thyroidstimulating hormone (TSH), as well as anti-thrombopoietin antibodies, must be monitored. In addition, it is opportune to perform regular TSH monitoring during treatment. When there are altered values, the decision of continuing or discontinuing the treatment must be made case by case. HCV and Type 2 Diabetes Mellitus Considering that hyperglycemia was considered an independent risk factor for the ‘nonresponse’ to antiviral treatment of chronic hepatitis C [36], and that abnormal glucose levels can be associated with host-related factors, such as age, gender, alcohol use, ethnicity, obesity and resistance to insulin, it is recommended that glucose levels in blood be controlled (by means of medications or changes in lifestyle), prior to the administration of antiviral treatment. BJID 2007; 11 Supplement 1 (October) However, during the acute phase of kidney disease (when renal insufficiency and systemic manifestations are present), it is recommended that antiviral treatment be avoided or discontinued. In this case, measures aimed at reducing the inflammatory activity of renal lesions (corticosteroid therapy), removing circulating cryoglobulins (plasmapheresis) and reducing the formation of new antibodies (cyclophosphamide administration) are used [41-45]. Under these conditions, favorable outcomes have been achieved with mycophenolate mofetil and, more recently, with anti-CD20 antibody (rituximab). Regarding the use of ribavirin, lower proteinuria and improved kidney function have been reported in patients treated with IFN + ribavirin [1, 46,47]. It must be highlighted that ribavirin clearance is reduced in patients with renal insufficiency, and that dialysis does not eliminate the medication. Therefore, the use of this antiviral drug in standard doses is not recommended for patients who present with creatinine clearance < 50 mL/min. HCV and Neuropathies Treatment with IFN-a is not efficacious and can worsen peripheral neuropathy in patients with chronic hepatitis C and MC [48]. A detailed review of available studies on patients with both HCV infection and peripheral neuropathy was inconclusive. There were patients who responded to corticosteroids, endovenous immunoglobulin or plasmapheresis combined with antiviral treatment [49]. HCV and Resistance to Insulin When treating patients with HCV infection, the physician has the challenge of differentiating patients with good prognoses from those with poor prognoses, especially regarding the intervention measures available. At the moment, the available data allow us to infer that the treatment of insulin resistance (reducing hyperinsulinemia) and of those factors that surely contribute to the onset and maintenance of steatosis can increase the rate at which a sustained virological response is achieved in patients with HCV infection who were treated with the combination of PEG-IFN and ribavirin. Comments Decisions regarding the treatment of the abovementioned pathologies must be taken on a case-by-case basis, since the pathogeneses of most of these clinical manifestations are unknown. Cumulative knowledge on each of the pathologies must be considered regarding the immunomodulatory effect of IFN-α, which is the standard treatment for HCV infection. One option would be the use of an immunosuppressive agent in conjunction with interferon when there are autoimmune phenomena. References HCV and Nephropathies The treatment options for HCV and nephropathies are essentially the same as those described for MC [37-40]. 1. Zuckerman E., Keren D., Slobodin G., et al. Treatment of refractory, symptomatic, hepatitis C virus related mixed cryoglobulinemia with ribavirin and interferon-alpha. J Rheumatol 2000; 27: 2172-8. www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Treating Extrahepatic Manifestations of Hepatitis C 2. Mazzaro C., Zorat F., Comar C., et al. Interferon plus ribavirin in patients with hepatitis C virus positive mixed cryoglobulinemia resistant to interferon. J Rheumatol 2003;30:1775-81. 3. Alric L., Plaisier E., Thebault S., et al. Influence of antiviral therapy in hepatitis C virus-associated cryoglobulinemic MPGN. Am J Kidney Dis 2004;43:617-23. 4. Cacoub P., Saadoun D., Limal N., et al. PEGylated interferon alfa2b and ribavirin treatment in patients with hepatitis C virusrelated systemic vasculitis. Arthritis Rheum 2005;52:911-5. 5. Mazzaro C., Zorat F., Caizzi M., et al. Treatment with peginterferon alfa-2b and ribavirin of hepatitis C virus-associated mixed cryoglobulinemia: a pilot study. J Hepatol 2005;42:632-8. 6. Cid M.C., Hernandez-Rodriguez J., Robert J., et al. Interferonalpha may exacerbate cryoglobulinemia-related ischemic manifestations: an adverse effect potentially related to its antiangiogenic activity. Arthritis Rheum 1999;42:1051-5. 7. Suzuki H., Takei T., Tsuji H., Nishikawa T. Membranoproliferative glomerulonephritis and demyelinating neuropathy caused by type II mixed cryoglobulinemia associated with HCV infection. Intern Med 2000;39:397-400. 8. Ferri C., Giuggioli D., Cazzato M., et al. HCV-related cryoglobulinemic vasculitis: an update on its etiopathogenesis and therapeutic strategies. Clin Exp Rheumatol 2003;21:S78S84. 9. Zaja F., De Vita S., Mazzaro C., et al. Efficacy and safety of rituximab in type II mixed cryoglobulinemia. Blood 2003;101:3827-34. 10. Vallisa D., Bernuzzi P., Arcaini L., et al. Role of anti-hepatitis C virus (HCV) treatment in HCV-related, low-grade, B-cell, nonHodgkin’s lymphoma: a multicenter Italian experience. J Clin Oncol 2005;23:468-73. 11. Hermine O., Lefrere F., Bronowicki J.P., et al. Regression of splenic lymphoma with villous lymphocytes after treatment of hepatitis C virus infection. N Engl J Med 2002;347:89-94. 12. Kelaidi C., Rollot F., Park S., et al. Response to antiviral treatment in hepatitis C virus-associated marginal zone lymphomas. Leukemia 2004;18:1711-6. 13. Zuckerman E., Zuckerman T., Sahar D., Streichman S. Attias therapy on t(14;18) translocation and immunoglobulin gene rearrangement in patients with chronic hepatitis C virus infection. Blood 2001;97:1555-9. 14. Gisbert J.P., Garcia-Buey L., Pajares J.M., Moreno-Otero R. Systematic review: regression of lymphoproliferative disorders after treatment for hepatitis C infection. Aliment Pharmacol Ther 2005;21:653-62. 15. Hainsworth J.D., Litchy S., Burris H.A., et al. Rituximab as firstline and maintenance therapy for patients with indolent nonHodgkin’s lymphoma. J Clin Oncol 2002;20:4261-7. 16. Somer B.G., Tsai D.E., Downs L., et al. Improvement in Sjogren’s syndrome following therapy with rituximab for marginal zone lymphoma. Arthritis Rheum 2003;49:394-8. 17. Ramos-Casals M., Lopez-Guillermo A., Brito-Zeron P., et al. Treatment of B-cell lymphoma with rituximab in two patients with Sjogren’s syndrome associated with hepatitis C virus infection. Lupus 2004;13:969-71. 18. Muratori L., Lenzi M., Cataleta M., et al. Interferon therapy in liver/kidney microsomal antibody type 1-positive patients with chronic hepatitis C. J Hepatol 1994;21:199-203. 19. Garcia-Buey L., Garcia-Monzon C., Rodriguez S., et al. Latent autoimmune hepatitis triggered during interferon therapy in patients with chronic hepatitis C. Gastroenterology 1995;108:1770-7. 20. Doffoel-Hantz V., Loustaud-Ratti V., Ramos-Casals M., et al. Evolution of Sjogren syndrome associated with hepatitis C virus when chronic hepatitis C is treated by interferon or the association of interferon and ribavirin. Rev Med Interne 2005;26:88-94. 67 21. Fernandez I., Castellano G., Salamanca R.D., et al. Porphyria cutanea tarda as a predictor of poor response to interferon alfa therapy in chronic hepatitis C. Scand J Gastroenterol 2003;38(3):314-9. 22. Jessner W., Der-Petrossian M., Christiansen L., et al. Porphyria cutanea tarda during interferon/ribavirin therapy for chronic hepatitis C. Hepatology 2002;36(5):1301-2. 23. Doutre M.S., Beylot C., Couzigou P., et al. Lichen planus and virus C hepatitis: disappearance of the lichen under interferon alfa therapy. Dermatology 1992;184:229. 24. Boccia S., Gamberini S., Dalla Libera M., et al. Lichen planus and interferon therapy for hepatitis C. Gastroenterology 1993;105:1921-2. 25. Barreca T., Corsini G., Franceschini R., et al. Lichen planus induced by interferon- alpha-2a therapy for chronic active hepatitis C.Eur. J. Gastroenterol. Hepatol 1995;7:367-8. 26. Nagao Y., Sata M., Ide T., et al. Development and exacerbation of oral lichen planus during and after interferon therapy for hepatitis C. Eur J Clin Invest 1996;26:1171-4. 27. Schlesinger T.E., Camisa C., Gay J.D., Bergfeld W.F. Oral erosive lichen planus with epidermolytic hyperkeratosis during interferon alfa-2b therapy for chronic hepatitis C virus infection. J Am Acad Dermatol 1997;36:1023-5. 28. Dalekos G.N., Christodoulou D., Kistis K.G., et al. A prospective evaluation of dermatological side-effects during alpha-interferon therapy for chronic viral hepatitis. Eur J Gastroenterol Hepatol 1998;10:933-9. 29. Protzer U., Ochsendorf F.R., Leopolder-Ochsendorf A., Holtermuller K.H. Exacerbation of lichen planus during interferon alfa-2a therapy for chronic active hepatitis C. Gastroenterology 1993;104:903-5. 30. Di Bisceglie A.M., Martin P., Kassianides C., et al. Recombinant interferon alfa therapy for chronic hepatitis C. A randomized, double-blind, placebo-controlled trial. N Engl J Med 1989;321(22):1506-10. 31. Lisker-Melman M., Bisceglie A.D., Usala S., et al. Development of thyroid disease during therapy of chronic viral hepatitis with interferon alfa. Gastroenterology 1992;102(6):2155-60. 32. Marazuela M., Garcia-Buey L., Gonzalez-Fernandez B., et al. Thyroid autoimmune disorders in patients with chronic hepatitis C before and during interferon-alpha therapy. Clin Endocrinol (Oxf) 1996;44(6):635-42. 33. Deutsch M., Dourakis S., Manesis E., et al. Thyroid abnormalities in chronic viral hepatitis and their relationship to interferon alfa therapy. Hepatology 1997;26(1):206-10. 34. Mazziotti G., Sorvillo F., Stornaiuolo G., et al. Temporal relationship between the appearance of thyroid autoantibodies and development of destructive thyroiditis in patients undergoing treatment with two different type-1 interferons for HCV-related chronic hepatitis: a prospective study. J Endocrinol Invest 2002;25(7):624-30. 35. Willson R.A. Extrahepatic manifestations of chronic viral hepatitis. Am J Gastroenterol 1997;92(1):3-17. 36. Lecoub A., Hernandez C., Simó R., et al. Glucose abnormalities are na independent risk factor for nonresponse to antiviral treatment in chronic hepatitis C. Am J Gastroentreol 2007;102:1-7. 37. Johnson R.J., Gretch D.R., Yamabe H., et al. Membranoproliferative glomerulonephritis associated with hepatitis C virus infection. N Engl J Med 1993;328:465-70. 38. Diego J.M., Roth D. Treatment of hepatitis C infection in patients with renale disease. Curr Opin Nephrol Hypert 1988;7:557. 39. Misiani R., Bellavita P., Baio P., et al. Successful treatment ofHCVassociated cryoglobulinaemic glomerulonephritis with a combination of interferon-alpha and ribavirin. Nephrol Dial Transplant 1999;14:1558-60. 40. Mazzaro C., Panarello G., Carniello S., et al. Interferon versus steroids in patients with hepatitis C virus-associated cryoglobulinaemic glomerulonephritis. Dig Liver Dis 2000;32:708-15. www.bjid.com.br 68 Treating Extrahepatic Manifestations of Hepatitis C 41. Dammacco F., Sansonno D., Han J.H., et al. Natural interferonalpha versus its combination with 6-methyl-prednisolone in the therapy of type II mixed cryoglobulinemia: a long-term, randomized, controlled study. Blood 1994;84:3336-43. 42. D’Amico G. Renal involvement in hepatitis C infection: cryoglobulinemic glomerulonephritis. Kidney Int 1998;54:650-71. 43. Bombardieri S., Ferri C., Paleologo G., et al. Prolonged plasma exchange in the treatment of renal involvement in essential mixed cryoglobulinemia. Int J Artif Organs 1983;6(Suppl. 1):47-50. 44. Ferri C., Moriconi L., Gremignai G., et al. Treatment of the renal involvement in mixed cryoglobulinemia with prolonged plasma exchange. Nephron 1986;43:246-53. BJID 2007; 11 Supplement 1 (October) 45. Madore F., Lazarus J.M., Brady H.R. Therapeutic plasma exchange in renal diseases. J Am Soc Nephrol 1996;7:367-86. 46. Jefferson J.A., Johnson R.J. Treatment of hepatitis C-associated glomerular disease. Semin Nephrol 2000;20:286-92. 47. Reed M.J., Alexander G.J., Thiru S., Smith K.G. Hepatitis Cassociated glomerulonephritis—a novel therapeutic approach. Nephrol Dial Transplant 2001;16:869-71. 48. Scelsa S.N., Herskovitz S., Reichler B. Treatment of mononeuropathy multiplex in hepatitis C virus and cryoglobulinemia. Muscle Nerve 1998;21(11):1526-9. 49. Ramos-Casals M., Trejo O., Garcia-Carrasco M., Font J. Therapeutic management of extrahepatic manifestations in patients with chronic hepatitis C virus infection. Rheumatology [Oxford) 2003;42(7):818-28. www.bjid.com.br BJID 2007; 11 Supplement 1 (October) 69 Hepatitis C Treatment Before and After Liver Transplant 1 Edson Abdala 1,2 , Daniela Rosa Magalhães Gotardo1 , Patrícia Rodrigues Bonazzi1,2 and Telésforo Bacchella 1 Liver Transplant Section, Department of Gastroenterology, University of São Paulo School of Medicine; 2Department of Infectious and Parasitic Diseases, University of São Paulo School of Medicine; São Paulo, SP, Brazil Advanced hepatic disease, either in the form of cirrhosis or hepatocellular carcinoma, caused by infection with the hepatitis C virus (HCV), is currently the main indication for liver transplant worldwide [1,2]. Hepatitis C also appears as an etiologic factor for terminal hepatic disease. However, although this procedure is defined as a standard therapy in both situations, recurrence of HCV infection is universally recognized. The potential for HCV infection to evolve in a more aggressive manner is greater among transplant patients than among immunocompetent individuals, and the reestablishment of hepatic cirrhosis in these patients can occur within five to ten years after the transplant. The progression to cirrhosis also occurs at a more accelerated pace in these patients, with decompensation rates higher than 40% a year after diagnosis [3]. Despite the risk of recurrence, cirrhosis caused by HCV has long represented a disease with good post-transplant evolution potential and low recurrence rates. In the mid 1990s, there was an increase in the rate of recurrence, which impairs the function of the graft and reduces the survival of the patient. Studies to determine the risk factors began to be carried out, and the adoption of certain measures has enabled better outcomes [4]. Natural History of Hepatitis C After Transplant Early recurrence of the HCV infection after the transplant, defined as the detection of HCV RNA in the serum or graft, is a practically universal event, observed in more than 95% of the cases. Hepatic disease recurrence is represented by a wide range of histopathological aspects, and the differential diagnosis with acute cellular rejection can delay its detection. In these cases, there is overlapping of histopathological standards, as well as immunopathogenic phenomena in common [5,6]. In the recurrence context, acute hepatitis generally occurs between one and six months after the liver transplant, at a frequency of approximately 70%. Its histopathological findings are characterized by hepatocyte edema, large-droplet steatosis, moderate lobular inflammation, and acidophilic corpuscles. Although spontaneous resolution of acute hepatitis C occurs in up to 15% of immunocompetent individuals, it is rarely observed in the transplant context [3,4]. Severe progressive cholestatic hepatitis can occur early, between one and three months after the transplant. This kind of recurrence is rarer, occurring only in 10% of the cases. Its severe evolution pattern is characterized by high levels of The Brazilian Journal of Infectious Diseases 2007;11 (5) Suppl. 1:69-73. © 2007 by The Brazilian Journal of Infectious Diseases and Contexto Publishing. All rights reserved. serum bilirubin (over 6 mg/dL), high serum levels of HCV RNA, central ballooning in the liver biopsy, low inflammatory infiltrate, and cholangiolar proliferation, without associated ductopenia, suggesting that HCV has a direct cytopathic effect. These patients evolve to rapid graft loss, and death occurs even before a new transplant attempt can be made [4]. In most cases, however, the hepatitis C recurrence is diagnosed as chronic hepatitis, with a more accelerated progression of fibrosis than that observed in the immunocompetent population, resulting in cirrhosis in 8% to 30% of patients within five years. Cirrhosis is also more aggressive in these patients, with a 65% cumulative risk of complications within three years. The histopathological findings found in the graft are similar to those found in the native liver of an individual with hepatitis C and include mixed portal infiltrate with lymphoid aggregates, periportal inflammation, varied lobular inflammation, and steatosis. These findings can be detected in 70% to 90% of patients one year after the transplant [7]. In all of these cases, however, the real recurrence rate can only be estimated through routine serial biopsies, considering that 20% to 30% of patients do not evolve to increased aminotransferase levels, and that such an increase lacks specificity, potentially resulting from other events, such as rejection, ischemia or opportunistic infections [8]. In the Liver Transplant Sector of the Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo (HCFMUSP, University of São Paulo School of Medicine Hospital das Clínicas), protocol biopsies are carried out every six months in the first year after the transplant, every year between the second and forth year after the transplant, and every three years after the sixth year of the transplant. An evaluation of 43 patients demonstrated histological recurrence caused by chronic hepatitis in 80% of cases in an average period of 9.9 months [9]. Risk Factors for the Severity of Post-Transplant HCV Recurrence of Hepatitis C The factors that determine the evolution of hepatitis C in patients submitted to liver transplant can be variables related to the donor and receptor, viral factors, and events associated with the transplant, resulting in greater severity of the disease and higher rates of graft loss. The factors that are more consistently associated with the severity of the disease are: advanced age of donors, treatment for acute cellular rejection involving pulse therapy with corticosteroids or administration of OKT3, and infection with the cytomegalovirus [3,5,7,10]. A better understanding of the factors that contribute to the progression of the disease may indicate the potentially modifiable mechanisms of its evolution. www.bjid.com.br 70 Liver Transplant and Hepatitis C Treatment Donor and Receptor Factors The use of older liver donors is a factor that negatively affects the fibrosis progression rate. Recent studies show a tendency toward a ten-year increase in the age of donors in the last decade. This measure, although applied to compensate for the low availability of organs, has been shown to have a direct influence on the degree of fibrosis in recurrent hepatitis C. Recent studies have shown that the ten-year difference in donor age (40 versus 50 years) has been associated with greater fibrosis progression (from 0.6 to 2.1 units a year) and with a decrease in the interval of appearance of cirrhosis (of up to eight years). Donor age seems to influence graft survival only in HCVpositive patients. However, there is little chance that this will change, since very few transplant programs are able to pair younger donors with HCV-positive receptors [11,12]. The involvement of immunogenetic factors is also considered, with studies that observed the association between HLA-B14 and HLA-DRB104 as beneficial to hepatitis C evolution, and the mismatch between the donor/receptor HLA-DRB1 with an increased recurrence risk [13,14]. Other donor factors that require further investigation include the use of live donors, hepatic iron content, and hepatic steatosis [15,16]. Viral Factors Some studies have associated high viral load before or soon after the transplant with the severity of post-transplant HCV recurrence. An analysis involving 284 North-American and Spanish patients showed that the pre-transplant viral load is an independent factor in the progression of fibrosis. In another study carried out in the United States, the five-year survival of patients submitted to transplant for HCV was found to be lower in patients with viral loads higher than one million mEq/mL [7]. The importance of the HCV genotype in the progression of the disease remains controversial. Although most of the studies conducted in the United States failed to show this association, a large collaborative European study showed a higher rate of progression and severity in transplant patients infected with HCV genotype 1b. One hypothesis is that, in the liver transplant context, the host immune response to HCV is stronger for the 1b genotype than for other genotypes, and that the tissue lesion is associated with this response [17]. Factors Associated with the Transplant The treatment of acute rejection episodes with the use of corticosteroid pulse therapy or anti-lymphocyte preparations has been associated with greater severity of hepatitis C recurrence. However, for such patients, the use of immunosuppressive regimens is recommended, which is sufficient to prevent moderate or severe rejection, as is the subsequent use of corticosteroid pulse therapy or OKT3 administration, but not to the point of exacerbating the hepatitis C progression or causing other long-term complications [5]. BJID 2007; 11 Supplement 1 (October) Considering that one of the hypotheses put forth to explain the more severe HCV recurrence observed in recent years is the increased potency of immunosuppressive agents, several studies have been carried out to minimize or even abandon the use of the immunosuppressive regimen. These studies have shown diverging results. However, there were differences among the studies in terms of the initial immunosuppressive doses and the dose reduction rates. In some studies, the prolonged use of maintenance corticosteroid therapy was associated with lesser severity of the hepatic disease recurrence. In this case, the method of reducing the dose was important, and gradual reductions have been associated with less aggressive forms of hepatitis C [18,19]. The use of azathioprine or mycophenolate mofetil has not been shown to have any consistent effect on HCV recurrence. Although cyclosporine has been shown to have antiviral properties in vitro, it has not shown to have any advantage over tacrolimus in clinical practice [20,21]. The long-term use of sirolimus can provide some benefit, since it has antifibrotic and potential antiviral effects, although such studies are still preliminary and do not support its preferential use in this group of patients. The use of new drugs, such as sirolimus and the interleukin 2 receptor antagonist, requires controlled and prospective studies. Therefore, the general immunosuppression status seems to be one of the possible determinant events in the course of recurrent hepatitis C [20,22,23]. Another factor that is associated with the transplant and negatively influences the post-transplant evolution of hepatitis C is the presence of infection with the cytomegalovirus, which leads to the worsening of fibrosis [24]. Pre- and Post-Transplant Approach to Treating HCV-Positive Patients Antiviral therapy is the main strategy used in treating HCVpositive patients. However, the ideal moment at which to intervene remains unknown. The authors of most studies have initiated the treatment for HCV recurrence when there is histological evidence of the disease. Alternative treatments include the use of antiviral therapy before or soon after the transplant, when there is still no clinical evidence of recurrent disease. This is known as pre-emptive therapy. Antiviral therapy is generally less efficient and less well tolerated in the transplant patient than in the immunocompetent patient. Treatment of Patients with Cirrhosis Who are on the Transplant Waiting List Viral clearance in the patient with cirrhosis, in addition to providing better expectations for the transplant (increased graft survival), can even interrupt the progression of fibrosis in these patients, and, in some cases, preclude the need for the liver transplant. Studies have proven that, although the side effect rates are high, the rate at which a sustained virological response is achieved in patients with compensated cirrhosis treated with www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Liver Transplant and Hepatitis C Treatment progressively higher doses of conventional interferon and ribavirin is approximately 22%, and can be even higher in those infected with genotype 2 or 3. When submitted to transplant, such patients do not present recurrence of the disease, confirming the validity of this therapeutic strategy [25]. In one study, carried out by Forns et al., 30 transplant waiting list patients of different functional classes were submitted to antiviral treatment. In that study, the efficacy of the treatment was evaluated on the basis of the virological response throughout the treatment period, as well as on the rate at which a sustained virological response was achieved. The simple reduction in the HCV viral load before the transplant was sufficient to avoid recurrence after the transplant (efficacy of approximately 66%). In addition, none of the patients achieving a sustained virological response experienced disease recurrence [26]. Most of the studies involving this population of patients have evaluated the efficacy of conventional interferon. Overall, they concluded that the treatment is recommended for patients with Child-Turcotte-Pugh class A or B cirrhosis and a model for end-stage liver disease (MELD) score lower than 18, or even in patients with decompensated cirrhosis. However, in the last case, following the treatment protocol in a center with support and possibility of immediate transplant [26]. In a recent study of data collected in the HCFMUSP Liver Transplant Sector, 37 transplant waiting list patients with HCVinduced cirrhosis were submitted to antiviral treatment. This population was composed of 46% women and 54% men, with a mean age of over 50 years. The predominant genotypes were 1 and 3. There was history of ascites and hepatic encephalopathy in 43.2% and 8.1% of the patients infected with genotypes 1 and 3, respectively. History of varicose digestive hemorrhage two months before the treatment was present in 5.4% of patients, and a history of spontaneous bacterial peritonitis was common (in 8.1%). Pegylated interferon was used in only five patients (all infected with genotype 1), and conventional interferon was used in the remaining patients. Both were used in combination with ribavirin. The mean duration of treatment was 7.9 months. The most common side effect associated with the treatment discontinuation was neutropenia (in 37.7%). Of the 37 patients, 14 (37.8%) presented viral load reduction of at least 2 log. In that study, the presence of compensated or decompensated cirrhosis did not affect the treatment response [27]. Recent studies describe the treatment with pegylated interferon and ribavirin in patients with decompensated cirrhosis. Although they present a considerable virological response, the frequency of severe complications necessarily leads to the need to always analyze the risk/benefit ratio before the decision to initiate treatment is made, also considering the feasibility of an emergency transplant [28,29]. Pre-Emptive Therapy Antiviral therapy before the establishment of histologically confirmed disease presents theoretical advantages, 71 considering that, immediately after the transplant, the HCV viral load and the degree of hepatic fibrosis tend to be lower. This could provide a better response to the treatment, similar to what occurs in nontransplant patients. However, this is a moment at which the immunosuppression is still high, interfering with the antiviral response, and the antiviral regimen tolerability is too low in view of all of the other post-transplant clinical complications, such as infections and cytopenias. In addition, the immunomodulatory effect of interferon can increase the risk of acute cellular rejection, which is higher in this phase of the transplant process. Another criticism of pre-emptive treatment is that it does not distinguish patients who will actually evolve to a more significant recurrence of the disease and for whom treatment is indicated, from those who might have no need of antiviral therapy after the transplant [30,31]. Controlled studies have shown that treatment with the combination of conventional interferon and ribavirin has an advantage over monotherapy with interferon. There was a delay in the appearance of recurrence in those patients, who presented viral load reduction and better histological profiles. Studies involving the use of pegylated interferon and ribavirin have also demonstrated histological improvement, although their results are generally disappointing, with sustained virological response rates of 7-13% with the use of isolated interferon, 16-33% with interferon and ribavirin, and 9% with isolated pegylated interferon [32]. This strategy is not applicable to all patients. Those with better MELD scores before the transplant seem to be the best candidates. The need to reduce the dose or even discontinue the treatment is common, typically caused by cytopenias and concomitant renal dysfunction, with secondary anemia, which makes the use of ribavirin particularly difficult [33]. Post-Transplant Treatment In general, most patients submitted to transplant for cirrhosis caused by HCV are treated after the transplant, when recurrence is already an established event. Unfortunately, most studies that support this treatment strategy have been uncontrolled, preventing the determination of the treatment risks, acute/chronic rejection rates, and even the evaluation of the therapeutic efficacy. This is a population that, in principle, presents the worst prognostic factors of evolution and treatment response, since it comprises older patients who are infected with genotype 1, have high viral loads and present more extensive fibrosis, as well as more often having a history of previous treatment. These characteristics are also associated with the fact that transplant patients present comorbidities that frequently prevent the use of full-dose therapies [34]. In this group of patients, protocol biopsies are an essential means of assessing the degree of hepatic fibrosis and should be carried out whenever clinically indicated (by an increase in aminotransferase levels) or at least on an annual basis, with the specific purpose of detecting and monitoring HCV recurrence [35]. www.bjid.com.br 72 Liver Transplant and Hepatitis C Treatment The results of this group are no less disappointing, with a sustained virological response rate of 12.5% with isolated interferon, 21% with interferon and ribavirin, and 9% with isolated pegylated interferon. In uncontrolled studies the combined use of pegylated interferon and ribavirin proved to be the best strategy, with responses between 30% and 45% [2,36-38]. The optimal duration of antiviral therapy remains undefined. Although most recent studies established treatment periods of 48 to 52 weeks, the validity of prolonging treatment in patients who achieved virological response by the end of the standard treatment period is still in question [39-41]. The advantages of therapy that begins within 6 to 24 months after the transplant, compared with pre-emptive therapy, is that these patients require less immunosuppression, present better clinical status, and are at lower risk of acute or chronic rejection [7]. The occurrence of acute or chronic rejection has not been a limiting factor to the treatment, although there are some reports on this subject [42]. The use of ribavirin as isolated therapy or as maintenance after the combined use with interferon has no subside in the literature [43]. Use of Adjuvant Therapy In this group of patients, one of the central issues is the high rate of side effects from antiviral drugs, especially cytopenia, which often requires dose reduction or even discontinuation of the treatment. The risk/benefit ratio of the use of erythropoietin or granulocyte colony-stimulating factor has not been well established. However, its use has facilitated the maintenance of antiviral treatment and the use of optimal doses of ribavirin and interferon [44]. Efforts have been made to investigate the use of ribavirin substitutes that do not cause hemolysis, such as viramidine, although controlled studies are still needed in order to determine the best strategy in relation to the adverse effects of the therapy [5]. Second Transplants in Cases of Hepatitis C Recurrence Although a second transplant is always an option in patients presenting hepatitis C recurrence, this strategy is historically associated with disappointing results. Receptor age, total bilirubin, high prothrombin time, older donor age, admission to the intensive care unit, high creatinine level, and high MELD score are predictive factors of short survival after a new transplant. Second transplants remain controversial and require comprehensive discussions in view of the low availability of organs and the use of MELD score as an organ allocation criterion, which implies that second transplants will be given to recurrent patients presenting more severe clinical profiles. In general, a second transplant is recommended if one of the variables related to recurrence, and thus the natural history of HCV recurrence, can be altered [45-47]. BJID 2007; 11 Supplement 1 (October) References 1. Gane E. The natural history and outcome of liver transplantation in hepatitis C virus-infected recipients. Liver Transplant 2003;9 (suppl3):S28-S34. 2. Teixeira R., Menezes E.G., Schiano T.D. Therapeutic management of recurrent hepatitis C after liver transplantation. Liver Int 2007;27:302-12. 3. Samuel D., Forns X., Berenguer M., et al. Report of the monothematic EASL conference on liver transplantation for viral hepatitis. J Hepatol 2006;45:127-43. 4. Rodriguez-Luna H., Douglas D.D. Natural history of hepatitis C following Liver Transplant 2004;17:363-71. 5. Wiesner R.H., Sorrell M., Villamil F., et al. Liver Transplantation Society Expert Panel. Report of the first international liver transplantation expert panel consensus conference on liver transplantation and hepatitis C. Liver Transplant 2003;9 (suppl3):S1-S9. 6. Guido M., Fagiuoli S., Tessari G., et al. Histology predicts cirrhotic evolution of post transplant hepatitis C. Gut 2002;50:697-700. 7. Terrault N.A., Berenguer M. Treating hepatitis C infection in liver transplant recipients. Liver Transplant 2006;12:1192-204. 8. Charlton M. Liver biopsy, viral kinetics and the impact of viremia on severity of hepatitis C virus recurrence. Liver Transplant 2003;9 (Suppl. 3):S58-S62. 9. Campos S.V., Bonazzi P.R., Abdala E., et al. Chronic hepatitis C recurrence after liver transplantation. Liver Transplant 2007;13 (suppl.1):S213-S14. 10. Cameron A.M., Ghobrial R.M., Hiatt J.R., et al. Effect of nonviral factors on hepatitis C recurrence after liver transplantation. Ann Surg 2006;244:563-71. 11. Berenguer M., Prieto M., San Juan F., et al. Contribution of donor age to the recent decrease in patient survival among HCV-infected liver transplant recipients. Hepatology 2002;36:202-10. 12. Machicao V., Bonatti H., Krishna M., et al. Donor age affects fibrosis progression and graft survival after liver transplantation for hepatitis C. Transplantation 2004;77:84-92. 13. Langrehr J.M., Puhl G., Bahra M., et al. Influence of donor/recipient HLA-matching on outcome and recurrence of hepatitis C after liver transplantation. Liver Transplant 2006;12:644-51. 14. Belli L.S., Burra P., Poli F., et al. HLA-DRB1 donor-recipient mismatch affects the outcome of hepatitis C disease recurrence after liver transplantation. Gastroenterology 2006;130:695-702. 15. Imber C.J., St.Peter S.D., Handa A., Friend P. Hepatic steatosis and its relationship to transplantation. Liver Transplant 2002;8:415-23. 16. Guo L., Orrego M., Rodriguez-Luna H., et al. Living donor liver transplantation for hepatitis C-related cirrhosis: no difference in histological recurrence when compared to deceased donor liver transplantation recipients. Liver Transplant 2006;12: 560-5. 17. Féray C., Caccamo L., Alexander G.J.M., et al. European Collaborative Study on factors influencing outcome after liver transplantation for hepatitis C. Gastroenterology 1999;117:619-25. 18. McCaughan G.W., Zekry A. Impact of immunosuppression on immunopathogenesis of liver damage in hepatitis C virus-infected recipients following liver transplantation. Liver Transplant 2003;9 (suppl3):S21-S7. 19. Lladó L., Xiol X., Figueras J., et al. Immunosuppression without steroids in liver transplantation is safe and reduces infection and metabolic complication: results from a prospective multicenter randomized study. J Hepatol 2006;44:710-6. 20. Berenguer M., Aguilera V., Prieto M., et al. Effect of calcineurin inhibitors on survival and histologic disease severity in HCVinfected liver transplant recipients. Liver Transplant 2006;12:762-7. www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Liver Transplant and Hepatitis C Treatment 21. Hilgard P., Kahraman A., Lehmann N., et al. Cyclosporine versus tacrolimus in patients with hepatitis C infection after liver transplantation: effects on virus replication and recurrent hepatitis. World J Gastroenterol 2006;7:697-702. 22. Jain A., Kashyap R., Demetris A.J., et al. A prospective randomized trial of mycophenolate mofetil in liver transplant recipients with hepatitis C. Liver Transplant 2002;8:40-6. 23. Tisone G., Orlando G., Cardillo A., et al. Complete weaning off immunosuppression in HCV liver transplant recipients is feasible and favourably impacts on the progression of disease recurrence. Journal of Hepatology 2006;44:702-9. 24. Chopra K.B., Demetris A.J., Blakolmer K., et al. Progression of liver fibrosis in patients with chronic hepatitis C after orthotopic liver transplantation. Transplantation 2003;76:1487-91. 25. Crippin J.S., McCashland T., Terrault N., et al. A pilot study of the tolerability and efficacy of antiviral therapy in hepatitis C virusinfected patients awaiting liver transplantation. Liver Transplant 2002;8:350-5. 26. Forns X., Garcia-Retortillo M., Serrano T., et al. Antiviral therapy of patients with decompensated cirrhosis to prevent recurrence of hepatitis C after liver transplantation. J of Hepatol 2003;39:905-15. 27. Oliveira P.R.D., Freitas A.C., Tengan F.M., et al. Hepatitis C antiviral therapy in cirrhotic patients. J Clin Virol 2006;38(suppl):S135-S6. 28. Horoldt B., Haydon G., O’Donnell K., et al. Results of combination treatment with pegylated interferon and ribavirin in cirrhotic patients with hepatitis C infection. Liver Int 2006;26:650-9. 29. Iacobellis A., Siciliano M., Perri F., et al. Peginterferon alfa-2b and ribavirin in patients with hepatitis C virus and decompensated cirrhosis: a controlled study. J Hepatol 2007;46:206-12. 30. Shiffman M.L., Vargas H.E., Everson G.T. Controversies in the management of hepatitis C virus infection after liver transplantation. Liver Transplant 2003;9(suppl3):S1129-S44. 31. Kuo A., Terrault N.A. Management of hepatitis C in liver transplant recipients. Am J Transplant 2006;6:449-58. 32. Szabo G., Katz E., Bonkowsky H.L. Management of recurrent hepatitis C after liver transplantation: a concise review. American J Gastroenterol 2000;95:2164-70. 33. Davis G.L. New approaches and therapeutic modalities for the prevention and treatment of recurrent HCV after liver transplantation. Liver Transplant 2003;9(suppl3):S114-S19. 34. Berenguer M., Palau A., Fernandez A., et al. Efficacy, predictors of response and potential risks associated with antiviral therapy in liver transplant recipients with recurrent hepatitis C. Liver Transplant 2006;12:516-19. 73 35. Sebagh M., Rifai K., Féray C., et al. All liver recipients benefit from the protocol 10-year liver biopsies. Hepatology 2003;37:1293-301. 36. Wang C.S., Ko H.H., Yoshida E.M., et al. Interferon-based combination anti-viral therapy for hepatitis C virus after liver transplantation: a review and quantitative analysis. Am J Transplant 2006;6:1586-99. 37. Murkherjee S., Lyden E. Impact of pegylated interferon alpha-2b and ribavirin on hepatic fibrosis in liver transplant patients with recurrent hepatitis C: an open-label series. Liver Int 2006;26:539-5. 38. Heydtmann M., Freshwater D., Dudley T., et al. Pegylated interferon alpha-2b for patients with HCV recurrence and graft fibrosis following liver transplantation. Am J Transplant 2006;6:825-33. 39. Lavezzo B., Franchello A., Smedile A., et al. Treatment of recurrent hepatitis C in liver transplants: efficacy of a six versus a twelve month course of interferon alfa-2b with ribavirin. J Hepatol 2002;37:247-52. 40. Rodriguez-Luna H., Khatib A., Sharma P., et al. Treatment of recurrent hepatitis C infection after liver transplantation with combination of pegylated interferon alpha-2b and ribavirin: an open-label series. Transplantation 2004:77:190-4. 41. Bizzolon T., Ahmed S.N.S., Radenne S., et al. Long term histological improvement and clearance of intrahepatic hepatitis C virus RNA following sustained response to interferon-ribavirin combination therapy in liver transplanted patients with hepatitis C recurrence. Gut 2003;52:283-7. 42. Khalili M., Vardanian A.J., Hamerski C.M., et al. Management of hepatitis C-infected liver transplant recipients at large North American centres: changes in recent years. Clin Transplant 2006;20:1-9. 43. Schiano T.D., Martin P. Management of HCV infection and liver transplantation. Int J Med Sci 2006;3:79-83. 44. Gotardo D.R.M., Abdala E., Bonazzi P.R., et al. Safety of recurrent hepatitis C treatment after liver transplantation with use of adjuvants. Liver Transplant 2007;13(suppl 1):S152. 45. McCashland T.M. Retransplantation for recurrent hepatitis C: positive aspects. Liver Transplant 2003;9(suppl 3.):S67-S72. 46. Wall W.J., Khakhar A. Retransplantation for recurrent hepatitis C: the argument against. Liver Transplant 2003;9(suppl 3):S73-S8. 47. Ercolani G., Grazi G.L., Ravaioli M., et al. Histological recurrent hepatitis C after liver transplantation: outcome and role of retransplantation. Liver Transplant 2006;12:1104-11. www.bjid.com.br 74 BJID 2007; 11 Supplement 1 (October) Adverse Event Management Aline Gonzalez Vigani Department of Viral Hepatitis Studies, State University of Campinas (Unicamp); Campinas, SP, Brazil The treatment of hepatitis C involves combining conventional interferon (IFN) or pegylated IFN (PEG-IFN) with ribavirin (RBV). These therapeutic regimens are associated with numerous adverse events, among which constitutional and neuropsychiatric symptoms, as well as hematological abnormalities, stand out [1,2]. The adverse events observed with the use of PEG-IFN or conventional IFN are similar, and the frequencies of those events are shown in Table 1 [3-5]. Adverse events resulting from the treatment of hepatitis C can jeopardize the quality of life of patients and their response to treatment. The control of those events involves medicinal and non-medicinal interventions. The latter include a reduction in the dosage of IFN or RBV and discontinuation of the treatment. A temporary or permanent reduction in the dosage of PEGIFN, conventional IFN or RBV as a result of an adverse event is necessary in approximately 30% of patients. In 10% of patients it is necessary to discontinue the treatment [4]. Hematological abnormalities (neutropenia, anemia and thrombocytopenia) and depression are the most common causes of dose reduction. The maintenance of the dosages of medications used in the treatment of hepatitis C and of the recommended course of treatment are important for the effectiveness of the therapy, Table 1. Adverse events associated with using the pegylated interferon-ribavirin combination in the treatment of hepatitis C Adverse event Reaction at the site of injection Fatigue Headache Myalgia Fever Chills Alopeica Artralgia Irritability Depression Anorexia Dermatitis Anemia Neutropenia Thrombocytopenia Approximate incidence (%) 36-58 50-64 50-62 42-56 43-56 24-48 28-36 27-35 24-35 21-34 14-32 16-21 12-22 17-20 3-6 as shown in Figure 1 [6]. Dose reduction is associated with lowering the rate at which a sustained viral response (SVR) is achieved. Early identification and strategies for controlling adverse events are important in the prevention of moderate and severe complications. These practices also attenuate the deleterious effects on the quality of life of patients and maximize the effectiveness of treatment for hepatitis C. Constitutional Symptoms Constitutional symptoms such as fatigue, headache and myalgia are the most common adverse events in patients Figure 1. Effect of dose reduction of pegylated interferon (PEG-IFN) and ribavirin (RBV) in relation to the sustained viral response (SVR) rate. treated with the combination of IFN and RBV (Table 1). Practically all patients manifest at least one such event during the administration of the initial doses of IFN. These symptoms typically disappear or become less intense after the first month of treatment. Paracetamol, acetaminophen and ibuprofen taken immediately before the injection of IFN alleviate the symptoms. Adequate hydration, together with light to moderate exercise, can help minimize these side effects. Hematological Effects In approximately 25% of patients, it is necessary at least to reduce the dosage of IFN or RBV due to abnormalities in laboratory test results [4]. Conventional IFN and PEG-IFN have both been associated with the suppression of hematopoiesis. In addition, hemolytic anemia occurs in all patients treated with RBV, although the intensity of that anemia is variable. The Brazilian Journal of Infectious Diseases 2007;11 (5) Suppl. 1:74-78. © 2007 by The Brazilian Journal of Infectious Diseases and Contexto Publishing. All rights reserved. Anemia Anemia resulting from the treatment of hepatitis C is a multifactorial side effect. Those factors increase the destruction of red blood cells and reduce their production. The increased destruction of red blood cells occurs due to hemolysis triggered by RBV, and the reduction in red blood www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Adverse Event Management cell production occurs due to IFN-induced suppression of bone marrow erythroid precursors [7,8]. The RBV penetrates erythrocytes, in which monophosphate, diphosphate and triphosphate are phosphorylated. Those pharmacologically active forms are incapable of passing through the erythrocyte membrane, and so remain intracellularly retained at a concentration 60 times greater than that of plasma [7,8]. The accumulated phosphorylated derivates are slowly eliminated from the erythrocytes, which have a half-life of 40 days. In contrast, the elimination of RBV from plasma is rapid, since it has a halflife of 24 hours. The mechanisms by which IFN can exacerbate RBVinduced anemia include suppression of hematopoiesis in bone marrow and an increase in the destruction of erythroid precursor cells [9]. Anemia starts and develops almost immediately after the initiation of therapy, becoming more intense after 4 to 6 weeks of treatment. The average decrease in the level of hemoglobin (Hb) is 2.5 to 3.0 g/dL, but more intense degrees of hemolysis can occur. Anemia associated with combined therapy can exacerbate other side effects resulting from the treatment of hepatitis C such as dyspnea, fatigue, dizziness and headache. Non-pharmacological control of the treatment of anemia involves dose reduction or permanent discontinuation of RBV [9]. In patients without heart disease, a reduction in the dosage of ribavirin when the levels of Hb fall below 10 g/dL is recommended, and discontinuation is recommended when Hb levels are under 8.5 g/dL (Table 2) [10]. Those recommendations are more restricted to patients with stable heart disease. In rare cases, when the level of Hb is lower than 8.5 g/dL, a transfusion of packed red blood cells, in conjunction with discontinuation of the treatment for hepatitis C, might be necessary. Between 9 and 22% of patients receiving combined treatment for hepatitis C require an RBV dose reduction due to anemia [3,4]. However, the strategy of RBV dose reduction has been associated with a modest increase in the level of Hb (˜1 g/dL) at 4-8 weeks after the reduction, as well as with a decrease in the rate at which an SVR is achieved (Figure 1) [6,11]. Hematopoietic growth factors such as alpha erythropoietin can be an alternative in relation to RBV dose reduction for the treatment of anemia arising during the treatment of hepatitis C. Alpha erythropoietin is a synthetic glycoprotein that has a mechanism of action similar to that of endogenous erythropoietin, which is a hormone produced by the renal peritubular capillaries that stimulates erythropoiesis in bone marrow. Studies indicate that therapy with erythropoietin at the dosage of 40,000 IU once a week elevates the level of Hb after one week of use and thus permits the maintenance of RBV dosage in patients that become anemic during combined therapy for hepatitis C [12,13]. In the study conducted by Afdhal et al., the initial dose of RBV was maintained in 88% of 75 the patients who received erythropoietin during treatment for hepatitis C, and Hb levels increased (from 10.6 ± 0.9 to 13.2 ± 1.2 g/dL) in those same patients [13]. The initial dose of RBV was maintained in 66% of the patients who received a placebo. The cost of alpha erythropoietin is high, but it is a therapeutic option for patients who develop an Hb level < 10 g/dL during treatment of hepatitis C [14]. Neutropenia A decrease in neutrophil counts to below normal values occurs in the majority of patients receiving conventional IFN or PEG-IFN and results from the suppression of bone marrow caused by those medications [15]. Neutropenia (neutrophil count <750 cells/mL) is more common with the use of PEGIFN than with that of conventional IFN, with values of 20% and 4%, respectively [3,4]. Intense neutropenia (<500 cells/ mm3) is also more common in patients treated with PEG-IFN than in those treated with conventional IFN, with values of 5% and 1%, respectively. After a single injection of PEG-IFN, neutrophil counts decrease, on average, 21% within the first 24 hours but generally stabilize at higher levels thereafter. Therefore, the measurement of neutrophil counts should be carried out before the administration of IFN so that more reliable values can be obtained. In patients who develop neutropenia during treatment for hepatitis C, in general, IFN dose reduction or discontinuation of treatment is necessary. Recommendations for dose reduction or discontinuation of the treatment due to neutropenia are shown in Table 2. Neutrophil counts generally return to pretreatment levels 2 to 4 weeks after the end of treatment [2]. The clinical implications of IFN-related neutropenia are not associated with an increased risk of infectious complications [16,17]. The low infection rates observed in neutropenic patients undergoing treatment for hepatitis C can reflect an early reduction in IFN dosage and, in addition, can indicate that those patients have a lower risk of infection when compared with neutropenic patients undergoing chemotherapy. Filgrastim is beginning to be used as an adjuvant therapy, at the dose of 300 µg 1 to 3 times a week, to increase neutrophil counts in patients who have less than 750 cells/mm3 during treatment for hepatitis C. However, clinical experience with filgrastim in such cases is still limited [10,15]. Filgrastim is a granulocyte-colony stimulating factor structurally similar to that produced by human cells and obtained through genetic engineering. Filgrastim interacts with receptors on the surface of myeloid progenitor cells in bone marrow, inducing proliferation, differentiation and activation of functional cells. Thrombocytopenia Thrombocytopenia observed in patients undergoing treatment for hepatitis C is a consequence of IFN-induced www.bjid.com.br 76 Adverse Event Management BJID 2007; 11 Supplement 1 (October) Table 2. Dose modifications for adverse events resulting from therapy with interferon and ribavirin according to manufacturer recommendations Hematological parameters Hemoglobina <10 g/dL <8.5 g/dL Platelets Neutrophils <25,000/mm3 <750/mm3 <500/mm3 PEG-IFN-α α 2b PEG-IFN-α α 2a IFN RBV Reduce to 11 mg/kg Discontinue Discontinue Reduce to 1 μg/kg Discontinue Discontinue Reduce to 135 μg Discontinue Discontinue Reduce by 50% Discontinue Discontinue PEG-IFN=pegylated interferon; RBV=ribavirin. aIn patients without heart disease or in patients with a history of stable heart disease undergoing combined therapy, IFN dosage should be reduced by 50% and RBV dosage should be lowered to 200 mg/day if there is a decrease in hemoglobin of ≥2 g/dL within a period of 4 weeks. Both medications should be permanently discontinued if hemoglobin drops to < 12 g/dL after dose reduction. Figure. 2. Algorithm for the pharmacological treatment of INF-induced depression. BDI=Beck Depression Inventory; CES-D=Center for Epidemiological Studies Depression Rating Scale; Zung SDS=Zung Self Rating Depression Scale. www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Adverse Event Management 77 bone marrow suppression. A drop in platelet counts during combined treatment for hepatitis C is common, but IFN dose reduction or discontinuation of treatment as a result of that drop is uncommon [2,3]. Thrombocytopenia requiring IFN dose reduction is more common in patients treated with PEGIFN and RBV than in those treated with conventional IFN and RBV, with frequency rates of 4% and 1%, respectively [18]. Recommendations for the discontinuation of PEG-IFN or conventional IFN as a result of thrombocytopenia are shown in Table 2. Platelet counts generally return to pretreatment levels by 4 weeks after the discontinuation of treatment [2]. There are ongoing studies that aim to evaluate the effectiveness and safety of growth factors that stimulate the production and maturation of megakaryocytes in thrombocytopenic patients undergoing treatment for hepatitis C. Rare cases of autoimmune thrombocytopenic purpura have been described during the treatment for hepatitis C, and the presence of that disorder should be investigated in rare cases involving an intense decrease in platelet counts. Thyroid Disease Approximately 1-6% of individuals treated with IFN develop thyroid abnormalities [23]. In all patients, an evaluation of thyroid function is recommended. Levels of thyroid-stimulating hormone should be determined before the initiation of treatment for hepatitis C, every 12 weeks during treatment and once after the end of the treatment. Individuals who develop hypothyroidism while undergoing treatment should receive hormonal replacement therapy. Neuropsychiatric Symptoms Approximately 20% to 30% of patients report depressive symptoms during treatment for hepatitis C [19]. The control of depression in patients who are candidates to receive treatment for hepatitis C is shown in Figure 2 [20]. In the pretreatment phase, it is important to establish the psychiatric status of the patient, which involves determining whether there is a history of depression, assessing the mood state and identifying abuse of illegal substances. Patients who abuse drugs or who have a complex psychiatric history involving disorders such as schizophrenia and bipolar disorder or who are receiving psychiatric treatment should be jointly accompanied by a psychiatrist during treatment for hepatitis C. Patients with current or previous depression should be evaluated before the initiation of treatment for hepatitis C, preferably through the use of scales that infer the intensity of depression. In patients with moderate depression, antidepressant medication can be given 4 weeks before the initiation of treatment for hepatitis C. Antidepressants belonging to a class of selective serotonin reuptake inhibitors (SSRIs) constitute the treatment of choice for IFN-associated depression [19]. Such antidepressants are safe and well-tolerated in patients with hepatic disease. In addition, the treatment success rate in patients with depression during treatment for hepatitis C is near 90% [18]. Fluoxetine, sertraline, citalopram, paroxetine and other SSRIs can have a slightly sedative effect, and the choice of SSRI to be used should be based on that effect and on the predominant symptomatology of the patient [21]. In patients with fatigue or cognitive slowness, fluoxetine or sertraline can be preferred over paroxetine, which tends to be less stimulating. In case of suicidal ideation or attempted suicide, the treatment of hepatitis C should be immediately suspended [22]. The dose of antidepressant medication started during treatment of hepatitis C should be slowly reduced over a 6 to 12 month period after the end of the treatment. Retinopathy The use of IFN can trigger or aggravate prior retinopathy. Subconjunctival hemorrhage and retinal hemorrhage have been reported during treatment with IFN [19]. Patients with risk factors for retinopathy such as systemic arterial hypertension and diabetes mellitus should undergo ophthalmological examination before and during therapy under the supervision of an ophthalmologist. Treatment should be discontinued in individuals who present either retinal lesions during treatment or the worsening of a prior lesion. Pulmonary Side Effects Dry cough, which can occur during treatment, has been associated with the use of RBV. In most cases the cough is tolerable, but occasionally it is necessary to discontinue the use of RBV. Cases in which cough becomes productive or is accompanied by abnormal pulmonary auscultation findings or fever, pneumonia should be investigated. Interstitial pneumonia can be severe, but it is normally reversible with the discontinuation of therapy [19]. References 1. Maddrey W.C. Safety of combination interferon alfa-2b/ribavirin therapy in chronic hepatitis C-relapsed and treatment-naive patients. Semin Liver Dis 1999;19 Suppl 1:67-75. 2. Fried M.W. Side effects of therapy of hepatitis C and their management. Hepatology 2002;36(5 Suppl 1):S237-44. 3. Manns M.P., McHutchison J.G., Gordon S.C., et al. Peginterferon alfa-2b plus ribavirin compared with interferon alfa-2b plus ribavirin for initial treatment of chronic hepatitis C: a randomised trial. Lancet 2001;22:958-65. 4. Fried M.W., Hadziyannis S.J. Treatment of chronic hepatitis C infection with peginterferons plus ribavirin. Semin Liver Dis 2004;24 Suppl 2:47-54. 5. Hadziyannis S.J., Sette H. Jr., Morgan T.R., et al. Peginterferonalpha2a and ribavirin combination therapy in chronic hepatitis C: a randomized study of treatment duration and ribavirin dose. Ann Intern Med 2004;2;140(5):346-55. 6. Shiffman M.L., Di Bisceglie A.M., Lindsay K.L., et al. Hepatitis C Antiviral Long-Term Treatment Against Cirrhosis Trial Group. Peginterferon alfa-2a and ribavirin in patients with chronic hepatitis C who have failed prior treatment. Gastroenterology 2004;126(4):1015-23. 7. Bodenheimer H.C. Jr., Lindsay K.L., Davis G.L., et al. Tolerance and efficacy of oral ribavirin treatment of chronic hepatitis C: a multicenter trial. Hepatology 1997;26(2):473-7. 8. De Franceschi L., Fattovich G., Turrini F., et al. Hemolytic anemia induced by ribavirin therapy in patients with chronic hepatitis C virus infection: role of membrane oxidative damage. Hepatology 2000;31(4):997-1004. www.bjid.com.br 78 Adverse Event Management 9. Kowdley K.V. Hematologic side effects of interferon and ribavirin therapy. J Clin Gastroenterol 2005;39(1 Suppl):S3-8. 10. Afdhal N.H. Role of epoetin alfa in maintaining ribavirin dose. Gastroenterol Clin North Am 2004;33(1 Suppl):S25-35. 11. Sulkowski M.S., Wasserman R., Brooks L., et al. Changes in haemoglobin during interferon alpha-2b plus ribavirin combination therapy for chronic hepatitis C virus infection. J Viral Hepat 2004;11(3):243-50. 12. Dieterich D.T., Wasserman R., Brau N., et al. Once-weekly epoetin alfa improves anemia and facilitates maintenance of ribavirin dosing in hepatitis C virus-infected patients receiving ribavirin plus interferon alfa. Am J Gastroenterol 2003;98(11):2491-9. 13. Afdhal N.H., Dieterich D.T., Pockros P.J., et al. Epoetin alfa maintains ribavirin dose in HCV-infected patients: a prospective, double-blind, randomized controlled study. Gastroenterology 2004;126(5):1302-11. 14. Devine E.B., Kowdley K.V., Veenstra D.L., Sullivan S.D. Management strategies for ribavirin-induced hemolytic anemia in the treatment of hepatitis C: clinical and economic implications. Value Health 2001;4(5):376-84. 15. Collantes R.S., Younossi Z.M. The use of growth factors to manage the hematologic side effects of PEG-interferon alfa and ribavirin. J Clin Gastroenterol 2005;39(1 Suppl):S9-13. BJID 2007; 11 Supplement 1 (October) 16. Sulkowski M.S. Management of the hematologic complications of hepatitis C therapy. Clin Liver Dis 2005;9(4):601-16. 17. Soza A., Everhart J.E., Ghany M.G., et al. Neutropenia during combination therapy of interferon alfa and ribavirin for chronic hepatitis C. Hepatology 2002;36(5):1273-9. 18. Aspinall R.J., Pockros P.J. The management of side-effects during therapy for hepatitis C. Aliment Pharmacol Ther 2004;20(9):917-29. 19. Russo M.W., Fried M.W. Side effects of therapy for chronic hepatitis C. Gastroenterology 2003;124(6):1711-9. 20. Raison C.L., Demetrashvili M., Capuron L., Miller A.H. Neuropsychiatric adverse effects of interferon-alpha: recognition and management. CNS Drugs 2005;19(2):105-23. 21. Edwards J.G., Anderson I. Systematic review and guide to selection of selective serotonin reuptake inhibitors. Drugs 1999;57(4):507-33. 22. Janssen H.L., Brouwer J.T., van der Mast R.C., Schalm S.W. Suicide associated with alfa-interferon therapy for chronic viral hepatitis. J Hepatol 1994;21(2):241-3. 23. Huang M.J., Tsai S.L., Huang B.Y., et al. Prevalence and significance of thyroid autoantibodies in patients with chronic hepatitis C virus infection: a prospective controlled study. Clin Endocrinol (Oxf) 1999;50(4):503-9. www.bjid.com.br BJID 2007; 11 Supplement 1 (October) 79 Treatment Options in the Management of Thrombocytopenia in Patients Infected with HCV André Cosme de Oliveira Liver Transplant Section of the Clinical Hospital of University of São Paulo School of Medicine (HC-FMUSP); São Paulo, SP, Brazil Thrombocytopenia (platelet count < 80,000/mm3) in patients with chronic hepatitis has been associated with two factors: the first is hypersplenism resulting from splenomegaly in portal hypertension [1]. The spleen continuously sequesters one-third of circulating platelets, so that splenomegaly increases the fraction of platelets trapped in the splenic sinusoids, especially when resulting from passive congestion or an increase in venous portal pressure [1]. Hypersplenism seems to be the most common cause of thrombocytopenia associated with liver cirrhosis and portal hypertension. The second mechanism is related to the decreased production of thrombopoietin, a hormone produced by hepatocytes, which regulates the development of the megakaryocyte. In cirrhosis, due to the reduction in the mass of functioning hepatocytes, there can be a reduction of thrombopoiesis in the bone marrow, leading to thrombocytopenia in the peripheral blood [1]. In some situations, patients who are otherwise eligible for HCV treatment with interferon and ribavirin cannot be so treated because their platelets counts are low, which jeopardizes the treatment [2,3]. Nevertheless, hepatitis C patients treated with interferon and ribavirin also present a drop in the platelet count as a side effect [2,3]. There is as yet no treatment consensus in the literature for the management of these patients. We therefore present some related studies that address the management of these patients, in the pre-treatment and intra-treatment phases. Patients receiving interferon alpha or peginterferon alpha can present a 30-50% reduction in the baseline platelet count, and a dose reduction is necessary in approximately 4% of the patients [2,3]. We should consider possible reduction of the dose when platelet counts drop to < 50,000/mm3, and discontinuation of treatment when < 25,000/mm3. When using peginterferon alpha-2, the weekly dose can be reduced from 180 μg to 135 μg, or even to 90 μg. While using peginterferon alpha-2b, the weekly dose can be reduced from 1.5 μg/kg to 1.0 μg/kg, or even to 0.5 μg/kg [2,3]. Human recombinant interleukin (IL)-11 (oprelvekin, Neumega®; Wyeth Laboratories, Collegeville, PA, USA) can be used as a way to stimulate the increase in the number of platelets at a dose of 5 μg/kg/day, subcutaneously, initially for 7 days. If necessary, maintenance can consist of 1 to 3 doses per week during HCV drug therapy, similarly to what is recommended for filgrastim (granulocyte colony-stimulating factor) [4]. The Brazilian Journal of Infectious Diseases 2007;11 (5) Suppl. 1:79-80. © 2007 by The Brazilian Journal of Infectious Diseases and Contexto Publishing. All rights reserved. Administration of IL-11 has multiple effects on the hematopoietic system, specifically promoting the proliferation and maturation of megakaryocytes and inducement of neutrophilia and thrombocytosis [5]. In patients with cirrhosis, its use should be restricted to those with Child-Pugh class A or B cirrhosis [5]. In a study conducted by Ghalib et al. [6], none of the patients presented increased platelet counts until day 4 of daily use of the medication, and the maximal therapeutic response was observed near day 13 (Figure 1). At 6 to 8 days after the initiation of treatment, 89% of the patients presented duplication of the initial number of platelets, and 78% of the patients presented platelet counts > 80,000/mm3. Another treatment option for thrombocytopenia in these patients is splenic embolization. Figure 1. Ratio between platelet counts and duration of treatment with interleukin-11. Splenic embolization should preferably be performed with injection of polyvinyl alcohol particles between 355 and 500 µm (Trufill PVA Embolization Particles; Cordis Corp., Johnson & Johnson Co., Miami Lakes, FL, USA) into the splenic artery and should not exceed an embolization volume of 30% of the initial spleen volume [7]. The most severe complications after embolization can be ascites, thrombosis of the splenic vein or portal vein, spontaneous bacterial peritonitis, and splenic abscess. Abdominal pain and fever are typically present postembolization [8]. Splenic embolization presented 50% of complication in embolizations < 30% of splenic volume, whereas, in embolizations > 30%, 100% of the patients presented complications [9]. Abdominal computed tomography scans can be performed 2 to 3 weeks after embolization for assessment of effectiveness of the procedure and to identify complications [10]. www.bjid.com.br 80 Thrombocytopenia in Patients Infected with HCV All patients should be previously vaccinated against pneumococci before embolization. In addition, antibiotic therapy should be initiated and maintained during hospitalization of the patient. We suggest the use of cefotaxime and ampicillin in these cases [11]. The combined treatment with interferon and ribavirin should not be initiated or reinitiated until 8 weeks after splenic embolization [12]. The use of human recombinant interleukin-11 (IL-11) (oprelvekin) and splenic embolization are both feasible treatments for these patients, taking into account the characteristics of the individual health care facilities at which the follow-up treatment is administered. References 1. Moreno A., Bárcena R., Blázquez J., et al. Thrombocytopenia can be found in patients with chronic hepatitis related to hepatitis C virus (HCV). Both hypersplenism and decreased liver production of thrombopoietin (TPO) have been hypothesized as mechanisms responsible for thrombocytopenia. Hospital Ramón y Cajal, Madrid, Spain. 2. Manns M.P., et al. Peginterferon alfa-2b plus ribavirin compared with interferon alfa-2b plus ribavirin for initial treatment of chronic hepatitis C: a randomised trial. Lancet 2001;358:958-65. 3. Fried M.W. Peginterferon alfa-2a plus ribavirin for chronic hepatitis C virus infection. N Engl J Med 2002;347:975-82. 4. Hennepin County Medical Center. Management of HCV and treatment side effects. April 10, 2005. BJID 2007; 11 Supplement 1 (October) 5. Lawitz E., Hepburn M., Casey T. A Pilot Study of Interleukin11 in Subjects with Chronic Hepatitis C and Advanced Liver Disease Nonresponsive to Antiviral Therapy. Gastroenterology Service; Infectious Diseases Service; Pathology Department; Brooke Army Medical Center, San Antonio, Texas. 6. Ghalib R., Levine C., Hassan M., et al. Recombinant Human Interleukin-11 Improves Thrombocytopenia in Patients With Cirrhosis. Hepatology 2003:1165-71. 7. Zhu K., Meng X., Li Z., et al. Partial splenic embolization using polyvinyl alcohol particles for hypersplenism in cirrhosis: A prospective randomized study. Eur J Radiol 2007;24. 8. Lee C.M., Leung T.K., Wang H.J., et al. Evaluation of the effect of partial splenic embolization on platelet values for liver cirrhosis patients with thrombocytopenia. World J Gastroenterol 2007;13(4):619-22. 9. Hayashi H., Beppu T., Masuda T., et al. Predictive factors for platelet increase after partial splenic embolization in liver cirrhosis patients. J Gastroenterol Hepatol 2007;7. 10. Lee C.M., Leung T.K., Wang H.J., et al. Evaluation of the effect of partial splenic embolization on platelet values for liver cirrhosis patients with thrombocytopenia. World J Gastroenterol 2007;13(4):619-22. 11. Foruny J.R., Blázquez J., Moreno A., et al. Safe use of pegylated interferon/ribavirin in hepatitis C virus cirrhotic patients with hypersplenism after partial splenic embolization. Eur J Gastroenterol Hepatol 2005;17(11):1157-64. 12. Pålsson B., Verbaan H. Partial splenic embolization as pretreatment for antiviral therapy in hepatitis C virus infection. Eur J Gastroenterol Hepatol 2005;17(11):1153-5. www.bjid.com.br BJID 2007; 11 Supplement 1 (October) 81 Therapeutic Perspectives for Hepatitis C OVERVIEW Evaldo Stanislau Affonso de Araújo and Antonio Alci Barone Laboratory of Hepatitis, LIM 47 DMIP-HC-FMUSP; São Paulo, SP, Brazil NEW THERAPIES FOR HEPATITIS C Jean-Michel Pawlotsky French National Reference Center for Viral Hepatitis B, C and Delta; Department of Virology, and INSERM U841, Hôpital Henri Mondor, Université Paris 12, Créteil, France Overview Hepatitis C virus (HCV) is represented by a single open reading frame of single-stranded RNA comprising approximately 3000 AA, positioned between 3’ and 5’ noncoding extremities. Transcription from the 5’ end results in a compound polyprotein, generated by structural proteins in the amino-terminal region extremity and by nonstructural proteins in the opposing terminal region. Post-transcription cleavage by viral and host cell proteases generates 10 individual HCV proteins [1]. The HCV cDNA clone was first described in 1989. Subsequently, counter to the expectations of an enormous breakthrough, there was a hiatus in the growth of knowledge regarding the molecular virology of this virus. Only in 1997 was the first functioning and complete HCV cDNA clone described and used in animal models (chimpanzees) for preliminary analyses of the viral expression using biotechnological tools. A milestone in the study of viral molecular biology was the description, in 1999, of a viral culture model in tumor cells (Huh 7) using a HCV 1b subgenomic replicon. Based on this model, despite strict limitations, other viral fragments were used in order to evaluate the dynamics of viral replication in different settings and with exposure to antiviral agents. Nevertheless, the inability to obtain a complete HCV sequence remains an obstacle to be overcome [1]. One characteristic of HCV is that is has minimal cytopathic effects, with immunomediated hepatotropic injury. However, it is also found in extrahepatic sites, where it has a short halflife (approximately 3 hours) and high serum turnover [2]. Standard treatment involves the administration of pegylated interferon alpha together with ribavirin. This treatment regimen has had modest success, a little over 50% [3], reaching 71% in cases of dose optimization and full compliance [4]. However, it could be much less, if we consider real life data, as well as characteristics of the host (race, comorbidities, immune state), of the liver disease (level of fibrosis and steatosis), of the medication (dose, interferon type) and of the virus itself. In addition, it is well known that genotype 1 has a lower response rate, and certain viral proteins can subvert the stimulus induced by interferon and ribavirin [3]. Viral kinetics, the evaluation of the decay pattern of the viremia – in terms of intensity and The Brazilian Journal of Infectious Diseases 2007;11 (5) Suppl. 1:81-87. © 2007 by The Brazilian Journal of Infectious Diseases and Contexto Publishing. All rights reserved. speed – after the initiation of the therapy, has proven to be a useful tool in the management of patients. It demonstrates the interaction among virus, drug, and host, in a relatively simple way: intense, rapid decay indicates a favorable case; insignificant, slow decay indicates cases that are unfavorable, moderate or intermediate. Such kinetic patterns allow patients to be characterized as rapid responders, slow responders, or nonresponders. This approach allows the treatment to be individualized, with ideal doses per weight and shorter, standard, or extended duration. In addition, it allows early prediction of treatment response. Therefore, individualized therapy constitutes the best therapeutic tool at the moment [5]. Of course, for the sake of effectiveness, safety, and tolerability, the current therapy is insufficient to confront the present hepatitis C epidemic. Therefore, from a therapy with poorly understood mechanisms of action that function through essentially indirect means [6], we have evolved to the specifically targeted antiviral therapy for hepatitis C [7], that is, the search for compounds that have a direct effect on the HCV life cycle. However, the preliminary results of current clinical studies have demonstrated worrisome aspects regarding safety, tolerability, and efficacy for some of these compounds [7], which is why we believe that treatment with interferon alpha will continue to lead the field for many years to come, and studies involving combinations of new compounds including interferon and, occasionally, ribavirin are already underway. A good example of this fact are the preliminary results of the PROVE-1 study recently presented in Barcelona [8], in which the group that used the combination of pegylated interferon alpha-2a 180 μg + ribavirin 1-1.2 g + telaprevir 750 mg every 8 hours, in an intention-to-treat analysis, showed a markedly greater response than that observed for the control group (receiving pegylated interferon + ribavirin), despite the fact that the subjects were infected with genotype 1. At week 4, 79% presented viremia < 10 IU/ mL. At week 12, 70% presented viremia < 10 IU/mL, whereas 39% presented viremia ≥ 10 IU/mL, emphasizing the loss of cases and the intention-to-treat analysis. Nevertheless, the incidence of rash, gastrointestinal effects, and anemia was significantly higher in the telaprevir group. The new therapeutic options are divided into ‘smallmolecule’ (protease and viral polymerase inhibitors and www.bjid.com.br 82 Therapeutic Perspectives for Hepatitis C protein inhibitors of the host – glucosidase) [9] – data summarized in Table 1 – and drugs whose targets are still indirect. In this second group, there are several that are currently in Phase I trials, specifically the Toll-like receptor agonists 7 and 9 (ANA 245 – Isatoribine – ANA 975 and Actilon) [10] whose mechanism of immune stimulation is broader than that induced by interferon alpha. The modulation of the inflammation and apoptosis by caspase inhibition is also under evaluation, and preliminary results demonstrate that aspartate aminotransferase normalizes in the patients who received it during trials ranging from Phase II to IDN-6556 [11]. There are other forms of interferon in phase I and II trials: albuferon (interferon alpha-2b fused to albumin, with more favorable posology regimen and similar efficacy to that of pegylated interferon); omega interferon; gamma interferon; interferon for oral use; and oral interferon inducers (resiquimod and imiquimod). In addition, there are nucleoside analogs similar to ribavirin (viramidine, now known as taribavirin) in Phase III, inosine monophosphate dehydrogenase inhibitors– VX98-497 (merimepodib) in Phase II for previous nonresponders [12], mycophenolate mofetil, also in Phase II for nonresponders, broad spectrum antiviral agents in Phase II (amantadine and rimantadine) and other immunomodulators in Phases I to III (histamine, thymosin alpha-1, IL-10 and IL-12). Finally, therapeutic vaccines are being studied in pre-clinical phases (E1/E2) I or II (E1, NS3NS4-NS5-core, IC41) [10]. After a brief review of developing drugs, we should bear in mind the problems and disappointments already encountered regarding the resistance developed in vitro or in vivo, in addition to the already mentioned weaknesses related to posology tolerance and complexity. Posology regimens, as already described for HIV, can be facilitated by the association with the protease inhibitor ritonavir, acting as a potent inhibitor of the enzyme system function of cytochrome P450, which would enable, in preliminary analyses, simplified posology regimens for some of the small molecules [13]. Regarding other problems, further review will bring up some relevant issues, in addition to the already evident necessary maintenance of interferon, and even ribavirin, in future therapeutic regimens. New Therapies for Hepatitis C The current standard of care for patients with chronic hepatitis C is the combination of pegylated interferon (IFN) alpha and ribavirin. This treatment is effective in approximately 80% of patients with hepatitis C virus (HCV) genotype 2 or 3 infection, but less than 50% of those with HCV genotype 1 [14]. Many new HCV drugs are at the preclinical developmental stage and several are in clinical development. Novel HCV therapies currently in development schematically belong to four categories: novel IFNs, alternatives to ribavirin, immune therapies, and specific and nonspecific inhibitors of the HCV lifecycle. BJID 2007; 11 Supplement 1 (October) Novel Therapies for Hepatitis C Novel IFNs New IFN alpha molecules are currently being developed. They are expected to yield more potent antiviral effects, and eventually more potent immunomodulatory effects, with improved pharmacokinetic and pharmacodynamic properties and, if possible, better tolerance. The potential interest in nonalpha IFNs is also being studied. Albumin-linked IFN alpha Albumin-linked IFN alpha (Albuferon®, Human Genome Sciences and Novartis) is an IFN alpha-2b molecule attached to a human albumin moiety which has a prolonged half-life that allows dosing at intervals of 2 to 4 weeks. Albumin-linked IFN alpha is able to induce an antiviral response in previous nonresponders to pegylated IFN and ribavirin combination. Results presented at the 42nd Annual Meeting of the European Association for the Study of the Liver (EASL) have shown no significant difference in the rates of sustained virological response between patients receiving pegylated IFN alpha-2a and ribavirin or various doses of albumin-linked IFN alpha administered every two or four weeks with ribavirin. Consensus IFN alpha IFN alphacon-1 (Infergen, Amgen and InterMuneYamanouchi) is a synthetic recombinant “consensus” IFN (cIFN) created by scanning the sequences of several natural alpha IFNs and assigning the most frequently observed amino acid in each corresponding position to the recombinant molecule [15]. There is no clear evidence that cIFN is superior to other alpha IFNs when both are given in equivalent doses in terms of sustained virological response. Other alpha IFNs in development Multiferon (Viragen) is a highly purified, multi-subtype natural human IFN alpha derived from human leukocytes which has already been approved for use in HCV therapy in several countries. Medusa® (Flamel Technologies) is a selfassembled poly-aminoacid nanoparticles system that can be used as a protein carrier for the development of novel longacting native protein drugs. Medusa IFN alpha-2a and Medusa IFN beta are currently in early clinical development. Various types of orally administered IFNs alpha are also currently being developed. Whether similar efficacy as with subcutaneous administration can be achieved is under study. Non-type I IFNs IFN omega, like IFN alpha or beta, is a type 1 IFN. It shares 70% homology with IFN alpha and binds to the same receptor. A stable, glycosylated form of IFN omega has been developed (Intarcia) and has been administered to patients with chronic hepatitis C in phase I and II trials. The results of a phase II trial presented at the 42nd EASL Annual Meeting have shown sustained virological response rates (HCV RNA below 50 international units (IU)/mL at week 12 post-therapy) of 36% in www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Therapeutic Perspectives for Hepatitis C 83 Table 1. Developing drugs that have a direct effect on HCV (adapted from [7,10]) Site of action Compound Input inhibitors Anti HCV Immunoglobulin (HCIg) Monoclonal antibodies (HCV-AB 68, HCVAB 65) HCV-RNA Antisense oligonucleotides transcription ISIS 14803 inhibitors AVI-4065 RNA that catalyzes the cleavage of the target RNA (Ribozymes) IRES small-molecule inhibitors Small Interfering RNA (siRNA) or short hairpin RNA (shRNA) Post-transcription Serum proteinase inhibitors processing to NS3-4A inhibitors Commercial name Development phase Posology Comments Civacir Phase II 200-400 mg/kg Poor preliminary results — Phase I — — Phase II Transitory reduction of viremia Interrupted — Heptazyme Phase I Phase I — Current Interrupted VGX-410C Phase I — Current TT033 Pre-clinical — Parenteral use — Interrupted BILN 2061 — ACH-806/Gs-9132 VX-950 Phase II (telaprevir) VO 450-750 mg PROVE –Telaprevir + every 8 h RBV+PEG-IFN: 88% with RNA < 10 IU in 12 without vs. 52% PEG+RBV HCV replication inhibitors Polymerase inhibitors NS5B (RdRp) Cyclophilin B inhibitors NS5A inhibitors Helicase inhibitors Viral assembly and release Glucosidase inhibitor (Iminosugar) SCH 503034 (boceprevir) ACH-806 and GS-9132 ITMN 191 ACH 1095 Nucleoside analogs/ Nucleotides Phase II — Phase I To initiate NM283 (Valopicitabine) –An.Nucleos. Phase II R1626 – An.Nucl. Phase II VO 200-400 mg every 8 h — Resistance Rash SCH+PEG-IFN-α-2b + RBV current Interrupted — — VO — — Potent, but GI effects VO 1.5-4.5g BID Marked reduction PEG-IFN-RBV combination under study Anemia MK-0608 Pre-clinical Non-nucleoside HCV-796 — inhibitors Phase II with PEG-IFN XTL 2125 Phase I R7128 Phase I DEBIO-025 Phase I — NIM 811 Phase I A 689 Pre-clinical — A-831 Pre-clinical — QU633 — — recombinant human antibodies UT-231B Phase II future — MX-3253 Phase II — (celgosivir) www.bjid.com.br Marked reduction without resistance Reduction of viremia — — — Poor results PEG-IFN/RBV combination 84 Therapeutic Perspectives for Hepatitis C the IFN omega-ribavirin combination arm versus 6% in patients treated with IFN omega alone. The manufacturer plans to develop an implantable infusion pump that will release a steady amount of IFN omega for approximately four weeks. In spite of in vitro results that would support a potentiating effect of IFN gamma on IFN alpha [16], the proof of concept that IFN gamma could have a utility in the treatment of chronic hepatitis C still remains to be made. IFN lambda-1 (interleukin 29 or IL-29) is a member of a novel family of cytokines that are distantly related to the IL10 family and type 1 IFNs. IFN lambda-1 exhibits dose- and time-dependent inhibition of HCV replication in various models, independent of types 1 and 2 IFN receptors and induced pathways [17]. A pegylated form of IFN lambda will soon enter clinical evaluation. Alternatives to Ribavirin Alternatives to ribavirin are needed that would have the same effects as ribavirin on infection in combination with pegylated IFN or other IFN molecules without its hemolytic properties. However, the challenge for discovering such molecules is hampered by the fact the antiviral mechanisms of ribavirin remain poorly understood. Taribavirin (Valeant Pharmaceuticals) is an amidine prodrug of ribavirin converted into ribavirin by adenosine deaminases, which are primarily present in hepatocytes. Taribavirin is thus preferentially taken up in the liver where it serves for ribavirin delivery to the major site of HCV replication, whereas it is not transported efficiently into red blood cells [18]. In two recent phase III trials in combination with pegylated IFN alpha 2a and 2b, respectively, taribavirin at a flat dose of 600 mg twice a day failed to achieve similar efficacy as weight-based dosed ribavirin in patients with chronic hepatitis C of various genotypes. The incidence of hemolytic anemia was however significantly lower with taribavirin. New trials with higher doses of taribavirin have been planned. Immune Therapies Various nonspecific immunomodulatory agents, including thymosin alpha-1 (Thymalphasin, SciClone), IL-10 or histamine (Maxym Pharmaceuticals) have been administered to patients with chronic hepatitis C, with little success. Vaccines can induce CD4+ and CD8+ T-cell responses to HCV. Preclinical and early human studies indicate that therapeutic vaccines using different forms of recombinant HCV proteins together with various adjuvants could upregulate both cellular and humoral immune responses in patients with chronic hepatitis C [19]. However, there is currently no evidence that therapeutic vaccines alone can result in changes in HCV RNA levels. It remains to be determined whether therapeutic vaccines could be useful in combination with potent antiviral molecules. Inhibitors of the HCV Life Cycle Every step of the HCV lifecycle constitutes a potential target for specific or nonspecific antiviral molecules. Many drugs are BJID 2007; 11 Supplement 1 (October) at the preclinical developmental stage and several are in clinical development, but initial trials using some of these inhibitors alone have raised concerns about their tolerability and the development of viral resistance. A number of specifically targeted therapies are now also being tested in combination with pegylated IFN alpha with or without ribavirin. Inhibitors of the early steps of the HCV life cycle Inhibition of HCV entry can be based on the use of specific antibodies that neutralize infectious particles and prevent their attachment to the receptor molecules. These include polyclonal hepatitis C immune globulins, that have been administered to prevent HCV infection in HCV-infected liver transplant recipients with little success [20], and anti-HCV monoclonal antibodies with high-affinity HCV neutralizing properties that reduce viral replication by 0.3 to 1.0 log [21,22]. Small molecule entry inhibitors can theoretically belong to two groups of drugs: molecules that specifically fix onto the HCV surface structures and neutralize the virus, or molecules that compete with infectious viral particles at the receptor level. Unfortunately, our understanding of HCV entry mechanisms remains rudimentary, hampering the development of such molecules inhibitors. Fusion could also become an interesting target for novel therapies when its mechanisms are better understood. HCV RNA translation inhibitors Several nucleic acid-based strategies have been tested, including antisense oligodeoxynucleotides, phosphorodiamidate morpholino oligomers (PMO), or ribozymes. All of them have been shown to potently inhibit HCV translation in vitro, but have been disappointing in vivo [23,24]. RNA interference initiated by small interfering RNAs (siRNA) or short hairpin RNAs (shRNA) is very specific and offers a potential to be used as antiviral against HCV. However, because of their size and chemical composition, siRNAs and shRNAs currently are not orally bioavailable and require parenteral administration. Alternative strategies currently target the three-dimensional functional internal ribosome entry site (IRES) complexed with ribosomal subunits and viral and cellular proteins with small molecule inhibitors. Inhibitors of HCV post-translational processing Highly selective, potent peptidomimetic inhibitors of HCV NS3/4A proteinase have been designed. VX-950 or telaprevir (Vertex Pharmaceuticals) and SCH 503034 or boceprevir (Schering-Plough Corporation) have now advanced to phase II clinical trials. Patients who received 750 mg of telaprevir alone every 8 hours experienced a median 4.4-log reduction in HCV RNA levels [25]. The drug is well tolerated over shortterm administration. However, viral breakthroughs occur during the second week of telaprevir administration in patients with low exposure to the drug. They are due to selection of telaprevir-resistant variants. Combination with pegylated IFN alpha with or without ribavirin could theoretically at least partly prevent telaprevir resistance. This hypothesis is currently www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Therapeutic Perspectives for Hepatitis C under study in two phase II trials. Boceprevir appears to have less potent antiviral properties than telaprevir in vivo at the doses used to date [26]. In combination, the antiviral effect of boceprevir appears to be additive to that of pegylated IFN alpha-2b [26]. A phase II clinical trial is ongoing in combination with pegylated IFN alpha and ribavirin, where high doses of boceprevir are administered to treatment-naïve patients. Other approaches have been developed to inhibit the NS3/ 4A serine proteinase function. ACH-806/GS-9132 (Achillion Pharmaceuticals and Gilead Sciences) inhibits binding of NS4A to the NS3 proteinase, therefore inhibiting polyprotein processing by preventing the formation of the active proteinase complex. ACH-806/GS-9132 has potent in vitro activity against HCV genotype 1 and administration of 300 mg twice daily for 5 days resulted in an average change in HCV RNA level from baseline of -0.9 log [27]. The development of this drug has been halted because of nephrotoxicity [27]. Inhibitors of HCV replication Inhibitors of the RNA-dependent RNA polymerase (RdRp) belong to two categories: nucleoside/nucleotide inhibitors, that target the catalytic site of the enzyme, and non-nucleoside inhibitors that target allosteric sites of the RdRp. Three RdRp inhibitors have been administered to patients in clinical trials, including two nucleoside and one non-nucleoside inhibitor. Two have been withdrawn due to toxicity. R1626 (Roche Products) induces a dose-dependent HCV RNA level reduction [28]. At very high doses, HCV RNA level decrease reaches more than 3 logs, but side-effects are frequent. R1626 in combination with pegylated IFN alpha and ribavirin has recently progressed into phase II of clinical development. Other drugs have entered phase I clinical development. The HCV RdRp has been reported to bind cyclophilin B, a cellular peptidyl-prolyl cis-trans isomerase that apparently regulates HCV replication through modulation of the RNA binding capacity of RdRp. Synthetic, non-immunosuppressive cyclophylin B inhibitors have been developed and are being tested in patients with chronic HCV infection [29]. Side-effect were however frequent and led to treatment withdrawal in several cases [16]. Another cyclophilin B inhibitor, NIM 811 (Novartis) is currently being tested in a phase I trial [30,31]. Inhibitors of virus assembly and release Iminosugars have been suggested to be able to cross cellular membranes and concentrate in the endoplasmic reticulum where they could competitively inhibit envelope proteins glycosylation and interfere with viral assembly [32]. MX-3253 or celgosivir (Migenix) has a modest antiviral effect on HCV in monotherapy. It is currently administered in combination with pegylated IFN alpha and ribavirin in a phase II clinical trial. Questions and Issues with New Therapies Among the many new avenues being explored, orally administered antiviral drugs that specifically inhibit a step of 85 the HCV lifecycle have come under the spotlight. However, although encouraging results have been published, serious issues have been raised as to the antiviral potency of these drugs, their tolerability, and the crucial problem of viral resistance. Antiviral Potency Antivirals usually enter development as soon as they show some degree of antiviral efficacy in one of the existing in vitro models. However, in vitro antiviral activity does not always translate into antiviral efficacy in vivo. There are several examples of drugs that were highly potent in vitro but failed when administered to patients. Unfortunately, studies with “negative” results are rarely published, even though they could teach us a lot. There are several possible reasons for these in vitro/in vivo discrepancies, including the use of poorly relevant preclinical models, misinterpretation of preclinical data, poor pharmacokinetics, poor delivery of a potentially potent drug to its target site, or the fact that the target is not physically accessible in infected cells in vivo. Tolerability Small-molecule viral inhibitors have been under close scrutiny for potential toxicity. The clinical development of BILN 2061 (Boehringer-Ingelheim, Ingelheim, Germany), a potent HCV NS3 serine protease inhibitor and the first drug of this type to be administered to infected patients, has been suspended because of myocardial toxicity in animals [33,34]. The development of an enormous number of potentially active HCV drug candidates was stopped before they were given to patients, owing to concerns raised by preclinical toxicity studies in vitro and in animal models. The clinical development of several drugs has been stopped because of their side-effect profile, including valopicitabine (digestive side-effects), HCV796 (ALT elevations) and ACH-806 (nephrotoxicity). Other drugs are still being developed in spite of serious side-effects. R1626 has been shown to induce a dose-dependent reduction of blood cell counts and hemoglobin levels after two weeks of administration at high doses [28], and the outcome of these effects is unknown if therapy is prolonged or if ribavirin is used in combination. Apparent safety during short-term administration does not guarantee that no serious adverse effects will occur when the drug is given for several weeks or months. Reesink et al. observed no serious adverse events during 14 days of telaprevir administration [25]. However, a recent commercial press release from Vertex Pharmaceuticals (December 13, 2006) states that, in the PROVE 1 study, a phase 2b twelve-week clinical trial of the triple combination of pegylated IFN alpha, ribavirin and telaprevir in treatment-naive patients, 3% of the patients discontinued telaprevir because of rash (rash was the most common reason for treatment discontinuation). Drug-drug interactions may also be a problem when HCV inhibitors are used in combination with pegylated IFN, with or without ribavirin. In addition, although synergy or additive efficacy may be expected, antagonism www.bjid.com.br 86 Therapeutic Perspectives for Hepatitis C can also occur. Here again, in vitro studies may give clues as to likely adverse effects but they cannot replace in vivo studies. Resistance A surprising finding has been the frequency and early timing of the emergence of resistance with all classes of antiHCV drugs. These results suggest that resistant variants are preexisting, fit, and ready to be selected by any specific HCV inhibitor. These findings disqualify HCV inhibitor monotherapy and raise major ethical issues as to whether naive or nonresponder patients should now be included in trials of these drugs in monotherapy, as there is a risk that they will be disqualified from future trials and therapies with drug combinations. Conclusion At this point, it is clear that specific HCV inhibitors should not be used alone. Combination therapy with oral antiviral drugs will require company portfolios to contain more than one such drug. And appropriate preclinical drug-drug interaction studies will have to be performed before clinical trials are initiated. This may take several years. In the meantime, new strategies are needed to improve the results of current HCV therapy. The “conservative“ approach aimed at optimizing pegylated interferon-ribavirin therapy should not be neglected. Preliminary results have shown that increasing the dose and/or the number of injections of pegylated IFN increases the response rate, and this is being further explored in ongoing trials. Likewise, increasing the dose of ribavirin has been shown to significantly improve the cure rate, and the adverse effects of ribavirin can now be partly controlled by using erythropoietin. Other trials are addressing the best way of tailoring the duration of treatment to the early virologic response (i.e. the HCV RNA decline at week 4 or even week 2). Another option is to enhance pegylated IFN-ribavirin efficacy by adding antiviral drugs with an additive or synergistic antiviral effect. Such combinations have the advantage of theoretically preventing the onset of resistance to the inhibitor, through the antiviral effect of IFN alpha. This may indeed happen in good IFN responders, although it is unclear whether IFN inhibition will be sufficient to avoid the emergence of resistance. In addition, patients with little or no response to IFN will in effect be receiving inhibitor monotherapy. Preliminary data are encouraging and ongoing trials will show how well double and triple combinations are tolerated, and whether the responses persist. References 1. Tellinghusein T.L., Evans M.J., Hahn T., et al. Studying Hepatitis C Virus: making the best of a bad virus. Journal of Virology 2007;81(17):8853-67. 2. Lindenbach B.D., Rice C.M. Unravelling hepatitis C virus replication from genome to function. Nature 2005;436(18):933-7. BJID 2007; 11 Supplement 1 (October) 3. Wohnsland A., Hofmann W.P., Sarrazin C. Viral determinants of resistance to treatment in patients with hepatitis C. Clinical Microbiologogy Reviews 2007;20(1):23-38. 4. Pawlotsky J.-M. Mechanisms of antiviral treatment efficacy and failure in chronic hepatitis C. Antiviral Research 2003;59:1-11. 5. Marcellin P., et al. Which patients with genotype 1 chronic hepatitis C can benefit from prolonged treatment with the ‘accordion’ regimen? J Hepatology 2007 (in press). 6. Goodbourn S., Didcock L., Randall R.E. Interferons: cell signalling, immune modulation, antiviral responses and virus countermeasures. Journal of General Virology 2000;(81):2341-64. 7. Pawlotsky J.-M., Chevaliez S., McHutchison J.G. The hepatitis C virus life cycle as a target for new antiviral therapies. Gastroenterology 2007;132:1979-98. 8. McHutchinson J.G., et al. EASL 2007 Late Breaker 786. 9. Harrison S.A. Small Molecule and novel treatments for chronic hepatitis C virus infection. American Journal of Gastroenterology 2007;102:1-7. 10. McHutchinson J.G., Bartenschlager R., Patel K., Pawlotsky J.-M. The face of future hepatitis C antiviral drug development: recent biological and virologic advances and their translation to drug development and clinical practice. Journal of Hepatology 2006;44:411-21. 11. Walters L., Nelson M. New therapeutic options for hepatitis C. Current Opinion on Infectious Diseases 2006;19:615-22. 12. Marcellin P., Horsmans Y., Nevens F., et al. Phase 2 study of the combination of merimepodib with peginterferon-a2b, and ribavirin in nonresponders to previous therapy for chronic hepatitis C. J Hepatology 2007 (in press). 13. Kempf D.J., Chen H.-J., Yeung J.T., et al. Pharmacokinetic boosting of VX-950, an inhibitor of HCV protease, by co-dosing with ritonavir. Journal of Hepatology 2006;44(2):S4. 14. NIH Consensus Statement on Management of Hepatitis C: 2002. NIH Consens State Sci Statements 2002;19:1-46. 15. Keeffe E.B., Hollinger F.B. Therapy of hepatitis C: consensus interferon trials. Consensus Interferon Study Group. Hepatology 1997;26:101S-7S. 16. Larkin J., Jin L., Farmen M., et al. Synergistic antiviral activity of human interferon combinations in the hepatitis C virus replicon system. J Interferon Cytokine Res 2003;23:247-57. 17. Marcello T., Grakoui A., Barba-Spaeth G., et al. Interferons alpha and lambda inhibit hepatitis C virus replication with distinct signal transduction and gene regulation kinetics. Gastroenterology 2006;131:1887-98. 18. Wu J.Z., Lin C.C., Hong Z. Ribavirin, viramidine and adenosinedeaminase-catalysed drug activation: implication for nucleoside prodrug design. J Antimicrob Chemother 2003;52:543-6. 19. Leroux-Roels G., Batens A.H., Desombere I., et al. Immunogenicity and tolerability of intradermal administration of an HCV E1based vaccine candidate in healthy volunteers and patients with resolved or ongoing chronic HCV infection. Hum Vaccin 2005;1:61-5. 20. Davis G.L., Nelson D.R., Terrault N., et al. A randomized, openlabel study to evaluate the safety and pharmacokinetics of human hepatitis C immune globulin (Civacir) in liver transplant recipients. Liver Transpl 2005;11:941-9. 21. Schiano T.D., Charlton M., Younossi Z., et al. Monoclonal antibody HCV-AbXTL68 in patients undergoing liver transplantation for HCV: results of a phase 2 randomized study. Liver Transpl 2006;12:1381-9. 22. Galun E., Terrault N.A., Eren R., et al. Clinical evaluation (Phase I) of a human monoclonal antibody against hepatitis C virus: safety and antiviral activity. J Hepatol 2007;46:37-44. 23. McHutchison J.G., Patel K., Pockros P., et al. A phase I trial of an antisense inhibitor of hepatitis C virus (ISIS 14803), administered to chronic hepatitis C patients. J Hepatol 2006;44:88-96. www.bjid.com.br BJID 2007; 11 Supplement 1 (October) Therapeutic Perspectives for Hepatitis C 24. Soler M., McHutchison J.G., Kwoh T.J., et al. Virological effects of ISIS 14803, an antisense oligonucleotide inhibitor of hepatitis C virus (HCV) internal ribosome entry site (IRES), on HCV IRES in chronic hepatitis C patients and examination of the potential role of primary and secondary HCV resistance in the outcome of treatment. Antivir Ther 2004;9:953-68. 25. Reesink H.W., Zeuzem S., Weegink C.J., et al. Rapid decline of viral RNA in hepatitis C patients treated with VX-950: a phase Ib, placebo-controlled, randomized study. Gastroenterology 2006;131:997-1002. 26. Sarrazin C., Rouzier R., Wagner F., et al. SCH 503034, a novel hepatitis C virus protease inhibitor, plus pegylated interferon á-2b for genotype 1 non-responders. Gastroenterology 2007;in press. 27. Pottage J.C., Lawitz E., Mazur D., et al. Short-term antiviral activity and safety of ACH-806 (GS-9132), an NS4A antagonist, in HCV genotype 1 infected individuals. J Hepatol 2007;in press. 28. Roberts S., Cooksley G., Dore G., et al. Results of a phase 1b, multiple dose study of R1626, a novel nucleoside analogue targeting HCV polymerase in chronic HCV genotype 1 patients. Hepatology 2006;44 (suppl. 1):692A. 87 29. Flisiak R., Orban A., Kierkus J., et al. The cyclophilin inhibitor DEBIO-025 has a potent dual anti-HIV and anti-HCV activity in treatment-naïve HIV/HCV co-infected subjects. Hepatology 2006;44 (suppl. 1):609A. 30. Ma S., Boerner J.E., Tiong Yip C., et al. NIM811, a cyclophilin inhibitor, exhibits potent in vitro activity against hepatitis C virus alone or in combination with alpha interferon. Antimicrob Agents Chemother 2006;50:2976-82. 31. Goto K., Watashi K., Murata T., et al. Evaluation of the antihepatitis C virus effects of cyclophilin inhibitors, cyclosporin A, and NIM811. Biochem Biophys Res Commun 2006;343:879-84. 32. Durantel D., Carrouee-Durantel S., Branza-Nichita N., et al. Effects of interferon, ribavirin, and iminosugar derivatives on cells persistently infected with noncytopathic bovine viral diarrhea virus. Antimicrob Agents Chemother 2004;48:497-504. 33. Hinrichsen H., Benhamou Y., Wedemeyer H., et al. Short-term antiviral efficacy of BILN 2061, a hepatitis C virus serine protease inhibitor, in hepatitis C genotype 1 patients. Gastroenterology 2004;127:1347-55. 34. Reiser M., Hinrichsen H., Benhamou Y., et al. Antiviral efficacy of NS3-serine protease inhibitor BILN-2061 in patients with chronic genotype 2 and 3 hepatitis C. Hepatology 2005;41:832-5. www.bjid.com.br 88 BJID 2007; 11 Supplement 1 (October) INSTRUCTIONS FOR AUTHORS Manuscripts for submission to The Brazilian Journal of Infectious Diseases should be sent to Anastácio Queiroz de Sousa, Editor-in-Chief, Rua Alfredo Magalhães, 04/Barra, 40140-140, SalvadorBahia-Brazil. Each manuscript will be assigned a registration number, and the author notified that the manuscript is complete and appropriate to begin the review process. Authors must indicate in a cover letter the address, telephone number, fax number, and e-mail of the corresponding author. The corresponding author will be asked to make a statement confirming that the content of the manuscript represents the views of the co-authors, that neither the corresponding author nor the co-authors have submitted duplicate or overlapping manuscripts elsewhere, and that the items indicated as personal communications in the text are supported by the referenced person. 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