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SENSITIZATION: Management Strategies Khalid Al Meshari, MD, FACP Director Renal and Pancreas Transplant Program King Faisal Specialist Hospital and Research Centre Layout 1. Definition 2. Scope of the problem 3. Detection techniques 4. Management Strategies, KFSH experience 5. Future Layout Definition Sensitization 1. Sensitization is defined as the presence of clinically relevant Abs in recipient sera against donor’ antigens 2. Relevance is determined by the reactivity of these Abs with donor’ lymphocytes expressing these antigens (positive CXM) Sensitization 1. methods: NIH-CDC, AHG-CDC Flow Cytometry Solid phase 2. How much Ab: Titer (1:2, 1:4, 1:8) Fluorescence intensity (MFI) Molecule of equivalent soluble fluorescence (MESF) Clinical Relevance of the Detection of HLA Antibody by Different Techniques T-Cell (Class I) B-Cell (Class II) CDC (IgG) Hyperacute rejection Flow (IgG) Acute antibody Acute antibody ‘mediated’ rejection ‘mediated’ rejection Single antigen beads Relevance unknown *Hyperacute rejection – if very high titre only Acute antibody ‘mediated’ rejection* Relevance unknown Immunological Memory: A Barrier to Transplant Decreased chance of receiving kidney Majority of those waiting longer are women 35 1 30 0.8 25 0.6 % 0.4 20 15 10 0.2 0 5 0 No sensitization Sensitized Odds No sensitization Men Sensitized Women Association of Preformed PRA with Various Characteristics of Patients Preformed PRA Characteristics Proportion female Proportion with retransplant Mean (SE) pretransplant blood transfusions None (n=3001) 1-50% (n=803) >50% (n=244) P 1014 (34%) 361 (45%) 154 (53%) <0.0001 159 (5%) 112 (14%) 97 (40%) <0.0001 3.47 (0.15) 6.01 (0.43) 10.7 (1.12) <0.0001 Opelz G. Lancet 2005;365:1570-76 TARGET ANTIGENS HLA or HLA-related antigens Major MHC antigens MHC class I (HLA-A, B, C) MHC class II (HLA-DR, DP, DQ) Minor MHC antigens MHC Class I polypeptide related sequences A (MICA) and B (MICB) Non-HLA related antigens Angiotensin II type 1 receptor Endothelium/monocyte antigens Perlecan Collagen Type IV, and VI Agrin Vimentin Myosin ABO blood group antigens Antigens Targeted by Anti-Donor Antibodies ABO MHC Antigen Class II Polymorphism: MHC Class I Minor Tissue HLA Specific Antigens Antigens Limited Very High Very High Limited Varies Vasc. Endothelium + + + + + Parenchyma + - + + + APC + + + + - T Cells + - + + - Antigens Targeted by Anti-Donor Antibodies ABO MHC Antigen Class II Polymorphism MHC Class I Limited Very High Very High Vasc. Endothelium + + + Parenchyma + - + APC + + + T Cells + - + HLA Antibodies MHC Class II Polymorphism: MHC Class I Very High Very High Vascular Endothelium + + Parenchyma - + APC + + T Cells - + Humoral Alloreactivity Pre Transplant Preformed Ab´s historic current time Humoral Alloreactivity Post Transplant De novo Ab´s de novo time Humoral Alloreactivity Pre Tx Preformed Ab’s Historic Current Post Tx De novo Ab’s Memory Response De novo Time But What Really is De Novo De novo Time De-Novo Anti-HLA Antibody may result from… Pre-transplant antibody present at low levels Recall memory response Inadequate immunosuppression Inflammation Pathways to HLA Sensitization • • • • • • • • Pregnancies Previous transplant Blood transfusions Use of tissue allografts for vascular reconstruction in congenital heart surgery CMV infection Bacterial infections Vaccination Left ventricular assist devices Antigens Targeted by Anti-Donor Antibodies ABO Antigen Polymorphism Limited Vasc. Endothelium + Parenchyma + APC + T Cells + The ABO Blood Group Antigens NAG Precursor oligosaccharide Gal H NAG Gal H antigen Gal A B NAG A antigen Gal NAGA NAG Fuc Gal NAGA Fuc B antigen Blood group (phenotype) Genotypes Antigens Antibodies to ABO in serum A AA, AO A Anti-B B BB, BO B Anti-A AB AB A and B None O OO H Anti-A and anti-B Complement-Fixing Antibodies The structure of IgM Antigen Idiotype Epitope Fab (variable region) C1q-binding region J Chain C3b- and C4bbinding region CH2 CH2 CH3 CH3 FC Fc receptor types I, II, and III Antibodies involved in Pre-transplant Humoral Allo-immunity Antigenic target Type of Antigen Antibody Sensitizing events MHC Protein IgG or mix of IgG and IgM Previous transplant, pregnancy, or transfusion ABO Carbohydrate IgM or mix of IgM and IgG “Natural” (crossreaction with gut bacterial flora) Nature of the Alloantibody Response -HLA Isoagg Previous exposure T-cell help Repertoire Clonality YES YES Extensive Polyclonal Limited Polyclonal Naturally No autoproduced reactive T-cell? Layout Scope of the problem Defining the Scope of HLA Sensitization • 34% of USA waiting list is sensitized • Frequency of highly sensitized patients (14%) is increasing – Increasing number waiting for regraft – More sensitive techniques for detecting Ab • Some highly sensitized patients will never be transplanted • Many sensitized patients have living donors who are eliminated because of a positive XM Patients on the UNOS Wait-List 31 JAN ‘05 vs. Patients Receiving Deceased Donor Grafts Dec ’99 to NOV ‘04 Current PRA% 0 – 19% 20 – 79% 80 – 100% Waiting # 44,889 7,934 5,754 77% 13% 10% % 23% Transplanted # 34,036 3,613 1,882 % 86% 9% 5% 14% OPTN 18 FEB 2005 Waiting Time (days) Waiting Times for Sensitized Renal Candidates 3000 0-9 10-79 80+ 2500 2000 1500 1000 500 0 97-98 www.optn.org July 2006 99-00 Years Listed 01-02 <half transplanted Reasons Patients Removed From UNOS Wait-list Dec 1999 to Nov 2004 Current PRA% 0-19% 20-79% 80-100% D D Transplant 47% (33,998) 42% (3,611) 37% (1,879) Living Donor Transplant 20% (14,853) 12% (1,002) 5% ( 276) Condition Deteriorated 4% ( 3,065) 7% ( 561) 8% ( 409) Died 17% (12,290) 27% (2,292) 37% (1,906) 6% (4,216) 1% ( 401) 2% (1,221) 3% (2,302) <1% ( 243) 7% ( 595) <1% ( 61) 3% ( 235) 2% ( 141) <1% ( 24) 7% ( 383) 1% ( 55) 3% ( 147) 1% ( 55) <1% ( 15) Other • Not clarified • Medically unsuitable • Refused Transplant • Tx’d Kid-Pan or Panc • Condition Improved OPTN 25 Feb 2005 Defining Scope of ABO Incompatibility • Based on blood group frequencies in the USA there is a 35% chance that any 2 individuals will be ABO incompatible • Blood type incompatible recipients have natural occurring antibodies against carbohydrate epitopes on disparate blood group molecules which can result in hyperacute rejection • Up to 1/3 of potential live donors are excluded on the basis of ABO incompatibility • About 1500 patients/yr in the US present with a willing but ABO incompatible live donor Kidney Median Waiting Times by ABO 2500 97-98 99-00 01-02 2000 1500 1000 500 0 O A B OPTN/UNOS Data as of 5-14-06 AB Immunologic Barriers • Sensitization to HLA – – – – Increases waiting time Associated with reduced graft survival Affects most, if not all transplanted organs Greater impact on women • ABO – Increases waiting time – Affects differ among ethnic groups because of differences in ABO distribution – ABOi Tx can experience hyperacute rejection Problems with Sensitized Patients • Hyperacute Rejection • Primary Nonfunction • Delayed Function • Accelerated Humoral Rejection – Not easily controlled by drugs – May cause lasting damage • Acute Cellular Rejection • Chronic Rejection Correlation of Immediate Graft Failures with Crossmatch Test 80% loss if XM+ 15 loss if XM180 Patients 150 42% loss if Ab+ 2% loss if Ab- 4 27 120 90 164 60 34 30 48 36 18 4 24 23 9 0 0 Preformed antibodies No preformed antibodies Random donor lymphocytes 6 +XM -XM Donor lymphocytes Patel, Terasaki. NEJM 280: 1969 1-year graft survival analysis of kidney transplants from cadaver donors in relation to PRA Opelz G, Lancet 2005 ;365:1570-6 Opelz G. Lancet 2005;365:1570-76 10-Year follow-up of kidney grafts from HLA-identical sibling donors Death censored functional graft survival of HLA-identical sibling transplants KFSH DD List 60 58 Listed patients HSP 50 40 37 32 30 20 23 17 53% 10 0 40% 16 43% 5 2 O A B AB 40% KFSH DD List 150 Listed patients HSP 132 125 100 75 50 25 0 58 44% Layout Detection techniques Antibody Mediated Rejection Endothelium PMN Anti-HLA Ab C1 Fc Receptor Mediated Binding C’ Activation ADCC CDC C4 C4a + C4b Donor HLA MAC Platelet microthrombi C4d Evolution of Detection Techniques • 1999 – – – – T-AHG-CDC screening No solid phase assays Many important DSA not detected AMR histology not well characterized 2009 Transplant Programs have seen a revolution in technology • • • • • HLA typing HLA Ab Screen HLA Ab Specificity Donor Specific HLA Ab C4d staining → Molecular (low to high (allele) resolution) → Solid Phase (increase sensitivity) → Solid Phase (increase in resolution) → Flow Crossmatch (increase sensitivity) → More accurate Diagnosis of AMR Some issues to consider next: • Needs to standardize HLA Ab quantitation • Further studies needed to define relative risks of low levels of donor specific antibodies (e.g. detected by solid phase only) • Further studies needed to validate significance of HLA Cw, DP, DQ antibodies, and non-HLA Ab (e.g. MICA, MICB) Stages of Humoral Alloreactivity Banff Conference 2001/2003 Humoral Rejection Conference, NIH, April 2003 I. Latent Humoral Response Circulating Antibody II. Silent Humoral Rejection Circulating C4d Antibody deposition Accommodation? Pre-rejection? Circulating C4d III. Subclinical Humoral Rejection Antibody deposition Tissue Injury Circulating C4d Antibody deposition Tissue Injury IV. Humoral Rejection Graft Dysfunction Takemoto SK, et al. Am J Transplant. 2004; 4: 1033-1041 (with permission from E. Akalin) Layout Management Strategies Management Strategies Matching Strategies • Regional or national sharing of crossmatch trays • Mandatory sharing of 0-antigen mismatched kidneys • Acceptable mismatch / virtual crossmatch / HLA matchmaker • Paired kidney exchange Desensitization Diagnostics • Flow or Luminex® PRA • PD flow crossmatch • HLASA by solid phase assay • C4d staining Therapeutics • Apheresis • IVIG • Rituximab • Thymoglobuline® Acceptable Mismatch Recognition that matching is not for everyone. 85% of DD Txs are mismatched. Focus on appropriate mismatching rather than looking for a perfect HLA “match”. Shift emphasis to antibody evaluation for the identification of “acceptable mismatches”. Requires detailed evaluation of the patient’s HLA antibody specificities. Exposure to IPA and NIMA During Pregnancy Can Lead to Either Immunization or Tolerization Immunization Adult exposure to IPA Maternal site NIMA/IMA Tolerance Fetal exposure to NIMA Possible mechanisms Chimerism Privileged site: modulation of APC Soluble HLA Immune deviation Regulatory T cells: HLA-DR sharing? Anti-HLA Ab towards paternal HLA molecules Non-Inherited Maternal HLA Antigens are Often “Acceptable” Mismatches NIMA NIPA Antibodies 46 72 No antibodies 43 6 p<0.001 Donor Selection on Basis of Acceptable Mismatches Patient: A24,31; B27,51; DR4,- and 100% PRA AM: A25, A26, B44 Possible kidney donors: A25, A31; B27, B51; DR4 A26, A31; B27, B51; DR4 A24, A25; B27, B51; DR4 A24, A26; B27, B51; DR4 A24, A31; B44, B51; DR4 A24, A31; B27, B44; DR4 A25, A31; B44, B51; DR4 A26, A31; B44, B51; DR4 A25, A31; B27, B44; DR4 A26, A31; B27, B44; DR4 A24, A25; B44, B51; DR4 A24, A26; B44, B51; DR4 A25, A31; B27, B44; DR4 etc. Current Approaches to Determine Acceptable Mismatches • Cross-matches with a patient specific panel. (Leiden) • Use of single antigen expressing cell lines (SALs) (Leiden) • Consider non-inherited maternal HLA antigens • Single antigen beads • HLA Matchmaker program HLA Matchmaker • The HLA Type of the antibody producer determines what structural components of an immunizing HLA antigen that will be seen as nonself • Donor HLA-A, B mismatches are defined by triplets of amino acid residues (epitopes) on alloantibody-accessible sites of HLA molecules HLA Matching on the Triplet Level Donor mm B18 Patient B7 Immune system of the recipient recognizes: A single HLA mismatch or 11 triplet mismatches HLA Matching on the Triplet Level Donor mm B18 Patient B7 B52 A33 Immune system of the recipient recognizes: No triplet mismatches ! Allocation of Cadaver Kidneys to Highly Sensitized Patients Class et al. Transplantation 2004;78:190-193 40 35 30 25 20 AM 15 ETKAS 10 5 0 <6 >6 >12 >24 >36 Waiting time in months AM: allocation through the Acceptable Mismatch Program (n=57) KAS: Kidney Allocation System ET: allocation through the standard Eurotransplant system (n=11) Graft Survival Class et al. Transplantation 2004;78:190-193 100 80 60 AM 40 <5% PRA 5-85% PRA 20 >85% PRA 0 12 24 Post Tx [m] Comparison of graft survival between acceptably mismatched patients (n=112) and nonsensitized (5% panel reactive antibody (PRA), sensitized (6%-84% PRA), and HS (85% PRA) recipients whose transplants were arranged through the ET-KAS (14.328). GS, graft survival. Virtual Crossmatch Virtual Crossmatching: • Determining the presence of donorspecific HLA antibodies (DSA) without performing a crossmatch, by comparing the HLA type of the donor with the HLA antibody specificities of the recipient. Emory Algorithm for Evaluating the Sensitized Patient AJT Oct. 2006 Depictions of Kidney Paired Donations Conventional Kidney Paired Donation Donor Recipient Blood Type A Blood Type B Intended Pairs Blood Type Incompatible Blood Type B Blood Type A Unconventional Kidney Paired Donation Donor Recipient Blood Type A Blood Type O Blood Type O Blood Type A Positive Cross-match Matched Pairing Domino Paired Donation Donor 1: NDD incompatible Donor 2 Recipient 1 Recipient 2 1st eligible recipient From UNOS match run Montgomery et al. Lancet. 2006; 368: 419 NEAD Chains Donor 1: NDD incompatible Donor 2 Recipient 1 incompatible Donor X Recipient X A Nonsimultaneous,extended,Altruistic-Donor Chain Rees M, etal. New Eng J Med 2009;360 (11):1096-1101 1 2 3 4 5 6 7 8 9 10 Transplant Date: JUL 2007 JUL 2007 SEP 2007 SEP 2007 FEB 2008 FEB 2008 FEB 2008 FEB 2008 MAR 2008 MAR 2008 Recipient’s State: AZ OH OH OH MD MD MD NC MD OH O O A A B A A A A A O A A B A A A AB A AB 52% 0% 23% 0% 82% 78% 64% 3% 100% 46% Cauc Cauc Cauc Cauc Cauc Hispanic Cauc Cauc Cauc AA Brother Sister Husband Wife Daughter Father Wife Husband Friend Friend Brother Brother Mother Daughter Recipient’s Sex and ABO type: Donor’s Sex and ABO type: O Recipient’s PRA: Recipient’s Ethnicity: Recipient-to-donor Relationship: Wife Husband The initiating donor was an unpaired altruistic donor from Michigan. The recipient of Transplant 6 required desensitization to HLA DSA by T and B cell flow cytometry. 3 The recipient of Transplant 9 required desensitization to blood group (AHG titer of 1:8) 1 2 Daughter Mother Mother Daughter What is Desensitization? • Lower baseline DSA to below threshold of renal endothelial cell injury at time of transplant • Prevent memory response • Maintain low DSA level posttransplant – Detect and treat response early Desensitization as a Solution • Goal of desensitization is to reduce relevant antibodies to a level where a previously “incompatible” transplant can be performed safely and successfully. • Requirements – Accurate and thorough assessment of a patient’s immunologic risk – Timely and careful monitoring both pre- and post-transplant – Open and timely communication between clinical and laboratory staffs Desensitization Therapeutics Diagnostics • Flow or Luminex® PRA • Apheresis: • PD flow crossmatch • IVIG • SA by solid phase assay • Rituximab • C4d staining • Thymoglobuline® Specialized, dedicated Staff Transplant Team Transplant Surgeon Challenges to the Clinicians: Develop new therapeutic strategies to improve outcomes Transplant Physician Clinical Laboratory Challenges to the Labs: Researcher Challenges to the Researchers: Provide insights into the mechanisms underlying allograft injury Provide diagnostics tools to predict and monitor for allograft injury Provide environment for translation of basic science into clinical practice Risk Assessment • Sensitization History – Transplants: Ag exposure / immunologic response – Pregnancies: determine paternal HLA – Recent or large numbers of transfusions • Current Ab: specificity and titer – Specificity: DSA or not – Titer: correlates best with risk and treatment needed • Immunologic Profile of Current Tx – Number of mismatches – Child → mother, husband → wife – Nature of mismatch: repeat, highly immunogenic Zachary AA and Hart JM in Handbook of Human Immunology, 1997 Zachary et al., in Manual of Clinical Laboratory Immunology, 6th Edition, 2003 Desensitization High-dose IVIG Jordan SC, et al. Am J Transplant 2006; 6:459-66. Low-dose IVIG and Pheresis Zachary AA, et al. Hum Immunol 2005; 66:364-70. Rituximab Pescovitz MD, et al. Am J Transplnt 2005; 5(suppl11):324 Rituximab / High-dose IVIG Ashley A, et al. NEJM 2008;359:242-251 Controlling B &T Memory T and B – cell memory occurs together HLA A2 Pregnancy associated sensitization Anti-HLA A2 Ab Transplantation (1996) 62:672 B Transfusion associated sensitization IFNg Transplantation (1990) 45:987 Th APC IL-2 CTL Antibody and T-cell mediated rejection occur together Ab mediated rejection: T2 + T3 Tubulitis 50% (12/24) of cases Transplantation (1996) 61:1586 Fas Granzyme B Perforin Strategy Use new solid phase technologies to fully define level of risk then proceed to modulate the immune system prior to and around the time of transplant •Requires approaches specific to T & B-cell memory T-cell B-cell Plasma cell HLA Ab • IVIG - + - ++ • Pheresis - - - ++ • Rituximab - +++ - - +++ ++ + - • Thymoglobulin M. Zand, Transplantation 2005 Activity of rATG, Alemtuzumab, and Rituximab in Debulking the B-Cell Mass Naïve Activated Memory Pre-Plasma Cell Plasma Cell CD19 +++ +++ +++ +/- - CD20 ++ ++ ++ +/- - CD27 - +/- + +++ +++ CD38 - + +/- ++ +++ CD40 ++ ++ ++ ++ - CD52 + + - - - CD80/86 - ++ ++ +++ - CD95 + + + ++ + HLA-DR + ++ ++ ++ - sIgD ++ ++ +/- - - - +/- ++ ++ + sIgM/G rTAG Rituximab Alemtuzumab Anti B-Cell Activity Induced apoptosis in Resting B-cell Activated B-cell Plasma cell Rabbit Anti-thymocyte Globulin Yes Yes Yes Rituximab (anti-CD20) No Yes No Sirolimus No Yes No Mycophenolate Mofetil Inhibits B cell proliferation only Cyclosporine No direct activity on resting or activated B cells The Limitations of Immunomodulation (desensitization) • May not be available to all patients – Center expertise – High Titers – Deceased donors more difficult • Short term outcomes may be worse – Higher rejection than the general transplant population • Long term outcomes are not known • Burden of immunosuppression Limitations of Anti-CD20 Treatment • CD20 expression: – Low on pre-B cells, high in normal B cells, progressive loss in plasma B cells – Anti-CD20 depletes mature and memory B cells but not plasma cells nor pre-existing Ab titers • Vascular access to B cell compartment: – Blood > lymph nodes/spleen > peritoneal cavity – Follicular > MZ > GC • Survival factors in the B cell microenvironment: – BAFF/ BLyS B cell survival factors Which Option is Best for an Individual Incompatible Pair Defining the Immunologic Phenotype How difficult will they be to match? How difficult will they be to desensitize? How Difficult Will They Be to Desensitize? (+) XM Major Factors DSA titer (depth ) and multiplicity (breadth) Repeat mismatches Sensitized from previous transplant(s) How Difficult Will They Be to Desensitize? ABOi Major Factors Iso-hemoagglutinin Titer Donor Type: Difficulty: A1 >B >A2 Matching Donor/Recipient Pair to Transplant Modality KPD Desensitization Easy-to-match pair • O donor • Low PRA Difficult-to-match pair • AB donor • Broad sensitization Difficult-to-desensitize • High titer DSA • High immunologic risk Easy-to-desensitize • Low titer DSA • Low immunologic risk Highly sensitized No live donor available Desensitization High dose IVIg Wait for (-) XM DD kidney Live donor available Desensitization PP/IVIg or IVIg Tx Kidney Paired donation Desensitization with KPD Tx The Cost Segev,etal JAMA,2005;293:1883-1890 Per-Patient Analysis of Transplant Costs and Desensitization While Awaiting Donation From the Deceased Donor List vs Kidney Paired Donation (KPD) Years on Waitlist* Cost of Dialysis† Cost of Desensitizationŧ Cost of KPD§ Highly sensitized 6.73 $485,038 $233,717 $204,738 Blood type O 4.85 $374,605 $201,544 $172,565 Blood type A 2.97 $264,172 $169,372 $140,393 Blood type B 5.48 $411,309 $212,237 $183,258 Blood type AB 1.63 $185,453 $146,439 $117,460 Recipient type * Average years awaiting deceased donor donation are based on UNOS data. † Average per patient cost of dialysis calculated according to Medicare payments of $58,758/year for dialysis, for the number of years the average patient awaits a donation from the deceased donor waitlist, plus $89,508 for the cost of transplantation and the first year of postoperative care. Ŧ Average per patient cost of desensitization calculated according to Medicare payments of $28,979 for desensitization. $89,508 for transplantation and the first year of post operative care, and $17,118/year of immunosuppression for the number of years the patient would have awaited a donation from the deceased donor waitlist. § Average per patient cost of a patient successfully matched through a KPD calculated according to Medicare payments of $89,508 for the cost of transplantation and the first year of post operative care, and $17,118/year of immunosuppression for the number of years the patient would have awaite donation from the deceased donor waitlist. KFSH & RC Experience Desensitization Measures of Success • Prevention of hyper-acute AMR • Minimization and reversal of antibody mediated rejection • Induction of a state of relative donor specific unresponsiveness or a state of accommodation • patient and graft survival comparable to sensitized CXM negative recipients • Prevention of neoplasia • Prevention and/or minimization of opportunistic infections Diagnostic Algorithm KFSH&RC Sensitization present Clinical relevance to donor’s HLA Strength of reaction to donor’s HLA Diagnostic Algorithm KFSH&RC Sensitization present • LAB screen® mixed • LAB screen ® class I & II Clinical relevance to donor’s HLA • LAB screen ® single antigen class I & II • Molecular typing of donor’s HLA ( low resolution and occasionally high resolution) Strength of reaction to donor’s HLA • SFI, MESF • PD FCXM (T and/or B IgG) • CDC / CD AHG CXM (T and/or B IgG) Management Approach KFSH&RC Protocol Induction of desensitization High risk: • Positive CDC/AHG CDC CXM • Positive PD FCXM • Positive DSA Low risk: • Negative CDC/AHG CDC CXM • Positive PD FCXM • Positive DSA • Antibody depleting: IA • Antibody modulation: High dose IVIG • Anti-B cell xx Rituximab® • Antibody modulation: High dose IVIG • Anti-B cell xx Rituximab® Pre-conditioning Regimen High Risk Patients High dose IVIG (2.0gm/ kg) Daily Immunoadsorption : 3.5 plasma volume/ treatment 5 – 10 treatments Rituximab Single dose of 500mg IV MMF 500 mg BID -14 -7 -1 0 1 Pre-conditioning Regimen Moderate Risk HD IVIG (2.0gm/ kg) Rituximab Single dose of 500mg IV -14 -2 0 Pre-conditioning Regimen Low Risk IVIG (1.0gm/ kg) -1 0 Measures To Control Activated T & B Lymphocytes during the Pre-adaptation Phase Steroids (Standard Dose) Tacrolimus 0.15 – 0.3 mg/kg BID Target level: 10-15 ng /dl MMF 1 gm BID Thymoglobulin 1.5 mg/kg/day 0 1 2 Weeks Post Engraftment 3 4 Monitoring Post-Transplant: Intense monitoring 1 month 0 • High index of clinical suspicion for AMR • Class I or class II SA Luminex® • Renal allograft biopsy with C4d Staining Monitoring Post-Transplant: Routine Monitoring 3 * 6 ♣* 12 ♣* 24 ♣* Months *Protocol biopsy ♣*Protocol biopsy and SA Luminex® 36 ♣ Outcomes of Desensitization Protocols at Different Centers Long-Term Outcomes of Desensitization Protocols for HS ESRD Patients Trans. Program Patient # Protocol Used Patient Survival (3-5 Yrs) Graft Survival (3-5 Yrs) AR Rates Mean SCr (3-5 Yrs) Mayo Clinic #94 HD IVIG PE/IVIG LD 97% at 5 Yrs 80% at 5 Yrs. 35% 1.6 0.6 mg/dl JHU #90 PE + CMV-Ig 95% at 3 Yrs. 80.9% at 3 Yrs. 62% 1.2 0.3 mg/dl CSMC #96 HD IVIG 97% at 5 Yrs. 87% at 5 Yrs. 36% 1.5 0.4 mg/dl KFSH #85 Anti CD20Ab HD IVIG IA or PE/IVIG LD 98% at 5 Yrs 94% at 5Yrs 21% AMR* 13% ACR* 0.9 ± 0.4 mg/ dl Adapted from Jordan and Pescovitz Clin J Am Soc Nephrol 1:421-432, 2006 *Both clinical and subclinical rejections Pre-conditioning Regimen ABO Incompatible Tx KFSH&RC Protocol Plasma exchange (PE): 1.5 plasma volume qod 4 – 10 treatments each followed by small dose IVIG Rituximab Single dose of 500mg IV PE followed by high dose IVIG MMF 500 mg BID -14 -7 0 1 2 3 4 Recent Published Reports of ABOIncompatible Kidney Transplantation Reference Patients 1-Yr graft survival (%) Comment Schneulle and van der Woude (1998) 108 NA Meta-analysis of A2 donors Nelson et al. (1998) 46 94 10-yr experience with A2 and A2B donors Alkhunaizi et al. (1999) 15 93 A2 donors with plasmapheresis Sorensen et al. (2001) 15 93 A2 donors without plasmapheresis Gloor et al. (2003) 10 80 Included high titer anti-A recipients Shimura et al. (2000) 67 81 A1 and B donors; 8 yr graft survival, 66% Gloor et al. (2003) 8 100 Splenectomy and plasmapheresis Almeshari et al (2009) 7 100 Plasmapheresis / high dose IVIG/Rituximab A2 and A2B donors Non-A2 donors a Two year graft survival. Layout Future Development of B Lineage Immune Response and Memory Plasmablast/ short-lived plasma cell TH germinal center Memory B cell antigen TH Naïve B cells TH Activated B cells Long-lived plasma cell short-lived plasma cell Development of B Cells in the Germinal Center T helper B cell cell Antigen Cytokines Proliferation Somatic hypermutatino of IgV genes – selection of B cells with high affinity for antigen Follicular dendritic cell presents antigen to B cell Death of nonselected cells Plasma cell Memory B cell Somatic hypermutation, affinity maturation and memory B cell formation Acquisition of Diversity, Specificity and Functional Capacity • Ig Gene re-arrangement • Allelic exclusion • Somatic hypermutation • Affinity maturation • Isotype switching BLyS (BAFF) / APRIL System Ligands Receptors BLyS BAFF-R BCMA APRIL Heterotrimer TACI Proteoglycans • Increased B-cell survival • Co-stimulation of B-cell proliferation • Ig class switch recombination • Enhanced APC function • Germinal centre formation • Regulation of B-cell tolerance • Sequester APRIL at cell surface to improve TACI and/or BCMA signaling? • Mediator plasma cell trafficking Dillon et al. Nature Reviews Drug Discovery 5, 235 (2006) Copyright @ 2006 Nature Publishing Group. Nature Reviews / Drug Discovery Synergistic Depletion of Memory B Cells and Plasma Cells with Anti-CD20 and BLyS (BAFF)/APRIL Blockade • Maintenance of long-lived plasma cells and serological memory despite mature and memory B cell depletion during CD20 immunotherapy in mice. Di Lillo et al. J Immunol. 2008; 180: 361 • The Dependence of Plasma cells and Independence of Memory B Cells on BAFF and APRIL. O’Conner at al. J Immunol. 2008; 180(6): 3655. B Cell Development Shortlived plasma cell Long-lived Antibody removal strategies plasma cell Plasmablast Short-lived plasma cell Clonal expansion Memory cell Space-directed strategies Activated Follicular B cell Fetal liver and bone marrow Tolerance strategies CLP Pro-B IgM μ μ Large Pre-B Small Pre-B IgM IgD Targeted cell depletion strategies Clonal expansion Immature Activated (T1/T2) Marginal Zone B (spleen) Plasma Plasmablast cell Novel Monoclonal Antibodies and Fusion Receptor Proteins That Deplete and/or Block Activation of B Cells Belimumab Atacicept BR3Fc Anti-CD40 BCR BAFF (BLyS) BR3 Epratuzumab syk CD19 Ras AME-133 PKC Ocrelizumab Ofatumumab BCMA TACI CD22 p13K APRIL Activation/Survival B Cell Atacicept Long-lived plasma cell 138+ 20- DSA Bone marrow, secondary lymphoid organs Baseline anti-donor antibody titer variable person to person, variable from time to time The Effect of Desensitization Protocols on Human Splenic B Cell Populations in Vivo Ramos, et al. AJT 2007; 7: 402 – 407. The Ubiquitin-Proteasome Pathway Elimination of Cells Bortezomib (Velcade TM, Pyz-Phe-borolLeu) O N N Pyrazinoic acid N H H N Indication Treatment of myeloma OH B OH O phenylalanine leucine boric acid Side effects GI 87% Neutropenia 20% Thrombocytopenia 35% Neuropathy 36% Proteasome Inhibition Causes Apoptosis of Human Plasma cells and is an Effective Therapy for AMR and ACR Bortezomib (Velcade®) Everly MJ,Transplantation 2008;86 (12):1754-1761 Perry DK,AJT 2009;9 (12):201-209 Proteasome Inhibitors Role of Proteasomes • T & B cell activation • Cell cycle control • Cell adhesion and migration • Apoptosis of cells of T, B, and monocyte lineage • Antigen presentation Eculizumab : Anti C5 Eculizimab C9 C5a C5 C7 C8 C5 C5 convertase C5b C5 C5b C5 C5b Poly-C9 C5 C5b C7 C7 C8 C7 C8 Plasma membrane MAC Abbas AK, et al, Cellular and Molecular Immunology, 4th edn, 2000. Strategies for Future • Avoid antibodies • Remove antibodies mechanically • Deplete or suppress B cells by targeting developmentally regulated cell surface molecules (CD20, CD19) • Impair B cell proliferation by targeting space-dependent cell receptors (TACl, BMCR) • Eliminate or suppress mature antigen-specific antibodyproducing plasma cells (Bortezomib) • B cell tolerance/combined B/T cell tolerance • Encourage graft accommodation