Download Biological therapy

Biological therapy for the
manipulation of complement system
Prohászka Zoltán,
IIIrd Department of Medicine, Research Laboratory
Semmelweis University
[email protected]
Biological therapy
• Biological therapy refers to the use of medication that is tailored to
specifically target an immune mediator of disease or induce an
immunological mechanism to cure a disease.
• Targeted therapy in clinical immunology (or oncology) refers to
medications acting through specific molecular targets to achieve
immunomodulation or oncolysis, in contrast to less specific
treatments, like steroids or cytostatica.
• Specific form of targeted therapy is the substitutional therapy with
purified „factors”, like coagulation factors in haemophilia, or insulin
therapy
• Biological response modifiers (BRMs) are substances influencing
biological functions, like interferons, interleukins, growth factors and
colony stimulating factors
• Vaccination
Milestones in biological therapy
•
Serum therapy for diphtheria (1890)
The first therapeutic approach, that was created with the understanding
of the etiopathogensis of disease
Edwin Klebs
1834-1913
Corynebacterium diphtheriae
Klebs-Löffler bacillus (1883)
Emil von Behring 1854-1917
Nobel Prize in Physiology and Medicine, 1901:
Orvosi Nobel díj, 1901: "for his work on serum
therapy, especially its application against
diphtheria, by which he has opened a new road in
the domain of medical science and thereby placed
in the hands of the physician a victorious weapon
against illness and deaths". Diphteria antitoxin,
1890
Tom, the Horse (1894, London)
Johannes Bókay Jr
1858-1937
„based on an international
mandate, he checked the safity
of the diphteria antitoxin”
1894
Milestones in biological therapy
•
•
•
Serum therapy for diphtheria (1890)
Treatment for agammaglobulinemia with purified immunogobulin G (1952)
The development of monoclonal antibody (mAb) technology by Köhler and
Milstein (1975) leading to the approval of the first therapeutic murine mAb,
Muromonab-OKT3 (1986), for the prevention of transplantation rejection.
Niels K Jerne
Georges JF Köhler
César Milstein
In 1984, the Nobel Prize in Physiology
and Medicine was awarded jointly to
Niels K. Jerne, Georges J.F. Köhler
and César Milstein
"for theories concerning the
specificity in development and
control of the immune system and
the discovery of the principle for
production of monoclonal
antibodies".
Milestones in biological therapy
•
•
•
•
Serum therapy for diphtheria (1890)
Treatment for agammaglobulinemia with purified immunogobulin G (1952)
The development of monoclonal antibody (mAb) technology by Köhler and
Milstein (1975) leading to the approval of the first therapeutic murine mAb,
Muromonab-OKT3 (1986), for the prevention of transplantation rejection.
Moreover, the progress of molecular and transgenic technologies has
enabled the development of
– chimeric mAb, Abciximab-ReoPro (Gp IIb-IIIa, 1994) and Rituximab-Rituxan
(CD20, 1997),
– humanized (complementarity-determining region; CDR-grafted) mAb,
Trastuzumab-Herceptin (Her2/Neu, 1998) and Infliximab-Remicade (TNFa, 1998)
– fully human mAb, phage display–derived Adalimumab-Humira (TNFa, 2002) and
transgenic mouse-derived Panitumumab-Vectibix (EGFR, 2006)
•
The progress of development of these substances has found a niche in the
management of various severe diseases, including cancerous, autoimmune
and inflammatory syndromes.
Monoclonal antibody product analysis,
historical and forecast sales growth ($m)
50,000
40,000
30,000
20,000
10,000
0
2001
Avastin
denosumab
Xolair
ustekinumab
lumiliximab
Aurograb
Removab
ABT-874
Theraloc
2002
2003
Rituxan
Lucentis
Actemra
Synagis
ipilimumab
Mylotarg
Mycograb
Bexxar
reslizumab
2004
2005
2006
Humira
Erbitux
Cimzia
pertuzumab
Humax CD20
Simulect
teplizumab
Zenapax
abagovomab
2007
2008
Herceptin
Numax
Vectibix
Raptiva
galiximab
canakinumab
epratuzumab
Humax-EGFR
Proxinium/Vicinium
2009
2010
2011
Remicade
golimumab
ocrelizumab
ReoPro
Humax CD20 (I&I)
MAB Campath (CNS)
Zevalin
belimumab
2012
2013
bapineuzumab
Tysabri
Soliris
mepolizumab
MAB Campath
Rencarex
Humax CD4
inotuzumab ozogamicin
8
Monoclonal antibody product trends - companies
50,000
40,000
30,000
20,000
10,000
0
2001
2002
2003
Genentech
Amgen
Novartis
Merck KGaA
Alexion Pharmaceuticals
Mitsubishi Tanabe
Fresenius/TRION
Viventia
2004
2005
2006
Roche
Schering-Plough
Wyeth
Bristol-Myers Squibb
Eli Lilly
Eisai
OncoScience/YM BioSciences
2007
2008
2009
Abbott Laboratories
Elan
Chugai
UCB
Bayer Schering Pharma
Wilex
Ception
2010
2011
2012
2013
Johnson & Johnson
AstraZeneca
Biogen Idec
GlaxoSmithKline
Bristol-Myers Squibb/Medarex
Genmab
Menarini
9
Outline of the lecture
• Overview on monoclonal antibodies as therapeutics
• Molecular biological technologies to manipulate and
produce human antibody based therapeutics
• Examples to highlight the application of biological
therapeutics to manipulate the complement system
The structure of human immunoglobulin G
Antigen binding
Light chain (L)
Heavy chain (H)
•
Two light chains/molecule
–
•
Variable (V) domains
(VL and VH domains)
(kappa or lambda)
Two heavy chains/molecule
–
(mu, gamma, delta, alpha
or epsilon
Constant (C) domains
(CL, CH1, CH2, CH3)
Complementarity determining
(CDR) and hypervariable regions
in the heavy and light chains
How to produce humanized or human antibodies in
large scale?
• The sequence of the variable domains (VH, VL) with the 3+3
hypervariable regions are required
– these sequences are unique, and only present in the mature B cells
(after immunization or infectious disease)
• The sequence of the constant domains are also required
– Known and available
• Genetic modification of mouse monoclonal antibodies
– Chimera production
– Humanization
• Production of human antibodies
– Hybridoma technology
– Antibody (Phage) libraries
– Transgenic animals
Induction of anti-mouse immune response in
humans
„HAMA”: human anti-mouse antibodies
•loss of functional activity of the therapeutics
•induction of side effects
•interference in immunoassays
Production of human antibodies
Single-chain Ab
Engineering of constant domains
• Constant domains determine
– The biological functions of the antibodies
• Receptor interactions (Fc receptors)
• Complement activation (IgG1: ADCC reaction and CDC)
• Neutralization (IgG4)
– In vivo half-life and access to storage pools depends on
glycosilation, which is determined by expression/production
systems
• Tissue culture: prokaryotes, yeast, insect cells, eukaryote cells
• Living organisms: transgenic plants, transgenic animals (secretion
of antibodies to milk, to serum, etc…)
– The compartment of its production
• Bloodstream
• Milk (secretory component)
Targeting the human complement system by
biological therapeutics, examples
• The complement system is a plasma serine protease system,
composed by soluble (zymogen) proteases, proteins, humoral
regulators, cell-surface regulators and cellular receptors
• It is part of the complex plasma serine protease system, including
–
–
–
–
Coagulation
Fibrinolysis
Contact (kinin-kallikrein) system
Complement system
• These systems have common activators (injury) and common
regulators (protease inhibitors)
Key biological functions of complement
Tissue macrophages
Dendritic cells
Mast cells
B cells
Monocytes
T cells
Neutrophils
Complement system
Innate immunity
Clearance
•Opsonisation
•Immunecomplexes
•Lysis of pathogens
•Apoptotic cells
•Chemotaxis
•Necrotic cells
•Inflammation
•Activation of target cells
Adaptive immunity
•Augmentation of antibody production
•T-cell response
•Depletion of self-reacting B-cells
•Induction of B-cell memory
Schematic presentation of the complement system
Classical pathway (Immunecomplexes)
Alternative pathway (Spontaneous C3 activation)
Factor B and Factor D
Regulators:
C1-inhibitor, C4-binding protein, Factor I
C3 activation
Regulators:
MCP, CD59, DAF, Factor H and Factor I
Lectin pathway (Carbohydrate structures)
Alternative pathway
amplification
C3b
Opsonization
Antigen presentation
Antibody production
C5 activation
Anaphylatoxins C3a, C5a
Inflammation
Chemotaxis
Regulators:
S protein and Clusterin
C5-C9
Terminal Pathway
Lysis
Cellular damages
Induction of apoptosis
Complement related human pathologies
•
Deficiency (genetic or acquired)
–
–
–
•
Pathological activation
–
–
–
–
•
Autoimmune diseases (immunecomplex diseases)
Transplant rejection
Ischemia/reperfusion (stroke, myocardial infarction, etc…)
Hemodialysis, on-pump cardiac operation
Dysregulated activation and consumption
–
–
•
C1-inhibitor (hereditary angioedema)
Alternative pathway regulators (Paroxysmal Nocturnal Hemoglobinuria, atypical Hemolytic
Uremic Syndrome, )
Terminal pathway components (meningitis)
Sepsis
Pathological pregnancies, preeclampsia, HELLP syndrome, DIC
Complement related biological therapies
–
–
–
Substitution of deficient factor/protein
Non-specific inhibition of pathological activation
Targeted inhibition of complement activation
Substitution therapy for HAE with C1-deficiency
Classical pathway (Immuncomplexes)
Alternative pathway (Spontaneous C3 activation)
Factor B and Factor D
Regulators:
C1-inhibitor, C4-binding protein, Factor I
C3 activation
Regulators:
MCP, CD59, DAF, Factor H and Factor I
Lectin pathway (Carbohydrate strucutures)
•Life-threatening edematous attacks
Alternative pathway
amplification
(Bradikinin overproduction)
C3b
•Acute
treatment with C1-inhibitor
Opsonization
Antigen
presentation
concentrate
Antibody production
•Purified human C1-inhibitor
C5 activation
•Cetor/Sanquin or Berinert P/Behring
•Nanofiltrated Cinryze/ViroPharma (4th
Regulators:
most expensive drug, 350.000
$/year
S protein
and Clusterin
Anaphylatoxins C3a, C5a
•Recombinant
human C1-inhibitor
Inflammation
•Rhucin/Pharming
Chemotaxis
C5-C9
Terminal Pathway
Lysis
Cellular damages
Induction of apoptosis
Inhibition of pathological complement activation
Classical pathway (Immunecomplexes)
Alternative pathway (Spontaneous C3 activation)
Factor B and Factor D
Regulators:
C1-inhibitor, C4-binding protein, Factor I
C3 activation
Regulators:
MCP, CD59, DAF, Factor H and Factor I
Lectin pathway (Carbohydrate structures)
Alternative pathway
amplification
C3b
Opsonization
Antigen presentation
Antibody production
C5 activation
sCR1
(soluble complement receptor 1)
Anaphylatoxins C3a, C5a
Inflammation
Chemotaxis
Regulators:
S protein and Clusterin
C5-C9
Terminal Pathway
Lysis
Cellular damages
Induction of apoptosis
•Inhibition of
complement activation
on multiple levels
•Aimed to be used in I/R
injury situation, i.e. bypass operation
•Lack of breakthrough
results with this drug
Inhibition of pathological complement activation
Classical pathway (Immunecomplexes)
Alternative pathway (Spontaneous C3 activation)
Factor B and Factor D
Regulators:
C1-inhibitor, C4-binding protein, Factor I
C3 activation
Regulators:
MCP, CD59, DAF, Factor H and Factor I
Lectin pathway (Carbohydrate structures)
Alternative pathway
amplification
C3b
Opsonization
Antigen presentation
Antibody production
IgG4
C5 activation
Eculizumab
(humanized murine anti-C5 Ab)
1st most expensive drug, 409.500 $/year
Anaphylatoxins C3a, C5a
Inflammation
Chemotaxis
Regulators:
S protein and Clusterin
C5-C9
Terminal Pathway
Lysis
Cellular damages
Induction of apoptosis
Pexelizumab
(scV anti-C5 Ab)
Current on-label indication and off-label applications for
Eculizumab
•
On-label: Paroxysmal Nocturnal Hemoglobinuria (PNH)
–
–
–
–
•
Disease of hemopoetic stem cells (clonal deletion of GPI-anchor for receptors, including
complement regulators CD59 and DAF)
Red blood cells are susceptible to episodic hemolysis mediated by complement
Chronic, progressive disease with recurrent thrombosis and organ-ischemia
Current management: regular transfusions, anticoagulation, bone-marrow transplantation,
and since 2007 targeted therapy with Eculizumab
Off-label applications: Current clinical trials with Eculizumab
–
–
–
–
–
–
–
–
–
Atypical hemolytic uremic syndrome
Age-related macular degenration
Complement-mediated injury after kidney transplantation
Dense-deposit disease, C3-nephropathy
Neuromyelitis optica
Catastrophic Antiphospholipid syndrome
Cold-agglutinin disease
ANCA-vasculitis
Sickle-cell disease
A simplified overview on the classification of thrombotic
microangiopathies (based on Besbas et al., 2006, Kidney Int.)
•
Advanced etiology, no underlying disease
–
Infections
•
•
–
Complement dysregulation
•
•
–
Shiga-like toxin producing pathogens
Neuraminidase producing pathogens
Alternative pathway dysregultaion
Thrombomodulin mutation
Failure of von-Willebrand factor processing
•
•
•
•
Acute renal failure, HUS
Critically ill, HUS
•
Acute renal failure, HUS
•
Acute neurological symptoms,
TTP
•
TMA as severe complication
Acquired ADAMTS13 inhibitory antibodies
Congenital defect of ADAMTS13 protease
–
•
Typical clinical presentation
(Upshaw-Schülman sy)
Secondary forms, underlying diseases
Laboratory tests currently used for the work-up of patients
with clinical TMA in our laboratory
•
Advanced etiology, no underlying
disease
–
–
Complement dysregulation
•
•
–
Shiga-like toxin producing pathogens
Neuraminidase producing pathogens
–
–
–
–
–
Acquired ADAMTS13 inhibitory antibodies
Congenital defect of ADAMTS13 protease
(Upshaw-Schülman sy)
Secondary forms
•
•
CFH exons 2, 4, 6, 9 14-15, 17, 18, 2023
CFI exons 3, 5-6, 9-10, 12-13
CD46 exons 5-6
C3 exons 14, 20, 26-27, 37
CFB exons 6-7
THBD in progress
Haplotype analysis
–
–
•
C3, C4, FH, FB, FI
Mutation screening
–
Alternative pathway dysregultaion
Thrombomodulin mutation
–
•
•
CH50 and WIELISA-ALT
Complement protein determination
–
Failure of von-Willebrand factor
processing
•
•
Functional complement measurements
–
•
Infections
•
•
•
CFH tag SNPs
MCP tag SNPs
Copy number determination on 1q32
(MLPA)
Screening for autoimmune form of
aHUS (anti-Factor H IgG)
Current and future therapeutic options
for patients with aHUS
Episodic occurence of disease shub
(hemolysis with fragmented erythrocytes, LDH increase , low platelet count)
Therapy:
Plasma exchange
Immunosuppression
Cytostatica
Eculizumab
900 mg/week for 4 weeks,
thereafter 1200 mg/two weeks
ESRD, dialysis, tx
The autoimmune form of atypical HUS
(Biologicals for the treatment of autoimmune disease)
• Presence of pathogenic autoantibodies against factor H
– Linked to CFHR1-3 deletion
– Binding to the functionally active N-terminal part of the molcule
– Inhibition of the complement regulating activity of FH
• Specific therapeutic approach: inhibition of autoantibody production
by the depletion of B-cells
Rituximab (Rituxan, MabThera)
•
•
•
•
•
Anti-CD20 monclonal antibody (human-mouse chimera) developed to deplete B-cells
(treatment of lymphomas and leukemias)
The ligand of CD20 is unknown, the molecule is involved in the regulation of calcium
flux
The mechanisms of action are: induction of ADCC reaction, of complement
dependent cytotoxicity, and of apoptosis; and saturation of Fc receptors
Recently, the drug found its way to treat diseases characterized by hyperactive Bcells, producing autoantibodies
One treatment cycle (4 doses of 375 mg/m2, 1 each week) depletes CD20-pos B cells
from the periphery for ~2 years
CD20-positive B-cell depletion in
autoimmune diseases
• Rheumatological diseases
–
–
–
–
–
Rheumatoid arthritis
Systemic lupus erythematosus (SLE)
Sjögren’s syndrome
Dermatomyositis and polymyositis
Vasculitides
• Non-rheumatological autoimmune diseases
–
–
–
–
Idiopathic thrombocytopenic purpura (ITP)
Thrombotic thrombocytopenic purpura (TTP)
Autoimmune hemolytic anaemia (AIHA)
Pemphigus vulgaris and foliaceus
Perosa et al, J Intern Med, 2010
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16
250,00
109
50,00
300,00
100,00
/l
200,00
250,00
200,00
150,00
RR syst
Hgmm
IVIG
150,00
g/l
Diagnosis of TTP
0,00
RR diast
plt
Hgb
100,00
50,00
Feresis
0,00
Madách K és mtsai: Aneszteziológia és Intenzív Terápia, 2008; 38(1): 34-38
Mechanisms of action of IVIG in
autoimmune and inflammatory diseases
•
Blockade of Fc receptors on macrophages of the reticuloendothelial system of liver
and spleen
•
Restoration of the idiotypic–anti-idiotypic network
•
Suppression or neutralization of cytokines by specific antibodies in the IVIG
•
Blockage of binding of adhesion molecules on leukocytes to vascular endothelium
•
Inhibition of complement uptake on target tissues
•
Neutralization of microbial toxins
•
Saturation of the FcRn receptors to enhance the clearance of autoantibodies
•
Induction of inhibitory FcgRIIb receptors on effector macrophages
•
Neutralization of growth factors for B cells, such as B-cell activating factor
•
Inhibition of T cell–proliferative responses
•
Expansion, activation, or both of a population of Treg cells
•
Inhibition of the differentiation and maturation of dendritic cells
Ballow M, JACI, 2011
Mechanisms of action of intravenous immune globulin (IgIV) on the immune modulation of various components of the innate and adaptive immune systems.
(Adapted from Tha-In et al. Trend Immunol, 2008) DC, Dendritic cell; Mo, monocyte; NK, natural killer.
Take home messages
• Biological therapy, 2011: 29 companies, 52 products, several
hundreds of indications, 40 milliard US dollars annual turnover
• Several diseases, that were untreatable or treatable but only in nonspecific manner, are now efficiently cured or treated
• Based on continuous product development, there is
– increased efficacy (engineering of biological effects)
– decreased side-effects of novel products (100% human antibodies)
• Drugs, currently in clinical practice are increasingly used off-label,
and this will soon result in broadening of the field of indications
– rituximab for autoimmune diseases
• Alternative applications of different preparations for substitution
therapies is also spreading
– IVIG for modulation of autoimmunity and inflammation
• The appearance of generic drugs will also arrive soon (for rituximab:
1997+15=2012)
– Biosimilarity, in contrast to bioequivalency
Thank you for your attenetion!
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