kv 8.31 passive immunisierung und antikörpertherapien konteptvorlesung 3 themenblock 8 beda m....
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KV 8.3 1
Passive Immunisierung und Antikörpertherapien
Konteptvorlesung 3
Themenblock 8
Beda M. Stadler
Institut für Immunologie
Diese Folien stehen auch als PPT Files zum download bereit unter:http://www.iib.unibe.ch/wiki/?p=teaching/medical_students&l=de
Protein Pharmaceutika 2011
• 134 FDA-zugelassene
biotechnologisch hergestellte
(Proteine/Peptide) Medikamente
• Protein Pharmaceutika kosten
derzeit ca. $50 Milliarden/Jahr
2
3
Top Ten Monoclonal Antibodies2009/2010
Generic Name Brands® IndicationsSales $ billion2009 / 2010
Infliximab Remicade RA,UC,CD,Ps,PsA,AS 6.9 8.0
Bevacizumab Avastin Colon cancer 5.9 6.8
Rituximab Rituxan NHL,RA 5.8 6.7
Adalimumab Humira RA,Ps,JIA,PsA,AS,CD 5.4 6.5
Trastuzumab Herceptin Breast Cancer 5.0 5.5
Cetuximab Erbitux Colon, head and neck cancer 2.5 3.2
Ranibizumab Lucentis Wet macular degeneration 2.4 3.1
Natalizumab Tysabri Multiple sclerosis 1.0 1.7
Omalizumab Xolair Allergic Asthma 0.9 1.1
Palivizumab Synagis RSV 1.1 1.0
Thearapeutische Antikörper auch zu teuer für uns?
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Übersicht / Lernziele
• Antikörper Produktion einst• Passive Immuntherapie einst • Monoklonale /
rekombinante Antikörper• Antikörper als Therapie für:
• Immunsuppression• Krebs Therapie• Immunmodulation• Anti-Adhäsionstherapien
• Rückblick und ZukunftPaul Ehrlich vor 100 Jahren
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Passive Immuntherapie einst
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Passive Immuntherapie mit polyklonalen Antikörpern
Multispender human Gammaglobulin oder Pferd Antitoxin
Tetanus
Pferd AntiveninSchlangenbiss
Multispender human GammaglobulinTollwut
Multispender human GammaglobulinMasern
Multispender human GammaglobulinHepatitis A and B
Pferd AntitoxinDiphtherie
Pferd AntitoxinBotulismus
Pferd AntiveninSchwarze Spinnenbiss
TherapeutikumKrankheit
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Monoklonale AntikörperHybridoma Technologie
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Therapeutische Maus Antikörper
Probleme• Immunogenizität
(humane anti-Maus AK, HAMA)
• keine Effektor Mechanismen (Maus Fc)
• Unerwartete Toxizität (Reaktivitiät mit normalem Gewebe, Kreuzreaktion mit anderen Antigenen)
• Verlust von Tumorantigen (z.B. anti-idiotyp Behandlung von Lymphomen)
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Immunogenicität von Infliximab(chimärer monokl. Ak gegen TNFα)
Human anti-mouse Ab
Human/mouse chimeric Ab (Infliximab)
• 61% of patients had detectable antibodies after the fifth infusion of Infliximab
• Incidence of the formation of antibodies to Infliximab is reduced with immuno-suppressive therapy
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Diversity of Antibodies is Generated by Gene RearrangementDank Wissen um Grundlagen
rekombinante Antikörper
Die Domänenstruktur hilft zum Umbau von Antikörpern
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Fab
Fv
Fc
Hinge
VL
VH
CL
CH
CH2
CH3
(a) Immunoglobulin G
VHVL
scFv IgG
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Ezzell, Scientific American
Humanisierung von Maus Antikörpern
HumanMaus Chimäre Humanisiert
Therapeutische Antikörper
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Generic name Trade name Target Type
Muromonab Orthoclone OKT3 CD3 Murine
Abciximab ReoPro Platelet GP IIb/IIIa Murine
Rituximab Rituxan CD20 Chimeric
Daclizumab Zenapax IL2Ra Humanized
Basiliximab Simulect IL2R Chimeric
Palivizumab Synagis RSV Humanized
Infliximab Remicade TNFa Chimeric
Trastuzumab Herceptin Her2/neu/ErbB2 Humanized
Gemtuzumab Mylotarg CD33 Humanized
Alemtuzumab Campath-1H CD52 Humanized
Endrecolomab Panorex 17A-1 Humanized
Ibritumomab tiuxetan Zevalin CD20 Chimeric
Adalimumab Humira TNFa Human
Omalizumab Xolair IgE Humanized
Centuximab Erbitux EGFR/ErbB1/Her1 Humanized
Bevacisumab Avastin VEGF Humanized
Efalizumab Raptiva CD11a Humanized
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VHVL VHVLVHVL
CH1CL
VLVH
Fv scFvFab VHdsFv
(a) Monovalent antibody fragments
Fab scFvFv dsFv VH
Mono- & plurivalente Ak-Fragmente
scFv Diabody orTriabody Tetrabody
VH VL
L
Ag
L
VH VL
L
Ag
Ag
L
L
Ag
Ag
Ag
Ag
Ag
Ag
Ag
VH VL
(b) Single chain two-domain constructs
scFv Diabody Triabody Tetrabody
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Transgene Humanisation
humane Antikörper in human Ig transgenenen Mäusen
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Der Wunsch nach humanen AK
mRNA von B Zellen
mRNA cDNA
PCRofH&Lchaingenes
PCRofH&Lchaingenes
BlutAntikörper (Fab)
Expression aufp3 des M13
Phagen
Klonierungund Rekombination
Humane Antikörper mittels Repertoire Klonierung
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Expression von Antikörpern
AntikörperDNA
AntikörperVektor
Transfektion
Zellkultur
Expression
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Industrielle Antikörper Produktion
Flask SpinnerBioreaktor
(Bench scale)Bio Reactor
(Plant)
Subkultur Produktion
1mL → 100mL → 1L → 1Lx3 → 12L → 100L → 300L → 2’000L→ 10’000L
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Ersatz von passivem IgGdurch rekombinante anti-RhD
RhD+
Erythrozytenrekombinante Antikörper
mittels phage display
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Neue Therapeutische Antikörper
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Immunsuppressive Anikörper(anti-T Zell Antikörper)
• Lymphocyte immune globulin (Atgam®)• lytic to human thymic lymphocytes; blocks T-cell responding
• Anti-CD3: (OKT3®, muromonab-CD3)• binds CD3, blocks antigen binding; depletes T-cells
• Anti-CD3: huOKT3• Humanized version of OKT3
• Anti-CD4: OKT4• less infusion-related reaction
• Anti-Tac monoclonal antibody (Zenopax®)• binds IL-2 receptor of activated T-cells causing their inactivation
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OKT3
Campath-1H
CD3
CD52
Polyclonal Anti-Thymocyte Globulin
Lymphocyte Depleting Antibody Therapies
KV 8.3 24
Immunsuppressive Anikörper(anti B-Zell Antikörper)
• Anti-CD20: Rituxan® (Rituximab)• results in mature B-cell depletion
and induces apoptosis (B-cell mediated vascular rejection) for B-cell Non Hodgkin’s Lymphoma
• anti-CD52: Campath-1H (Alemtuzumab)• depletes T and B lymphocytes,
NK cells, monocytes and macrophages
slg
DRCD19 CD20
CD22
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Side effects
• fever • chills • weakness • headache • nausea • vomiting • diarrhea • low blood pressure • rashes
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Complement pathway Tumor cell
ADCC
Anti-Idiotyp IL-1
TNF
Mac
NK
-Tox
-Tox
-Tox
-Tox
-Y-90
-Y-90-Y-90
-Y-90
-Y-9
0-Drug
-Drug
-Drug
1 2
3 4
AntikörperKrebs Therapie Strategien
5Anti
Angiogenese
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Antikörper-abhängige Zytotoxizität
(ADCC)
Apoptosis viaInduktion von intrazellulären
Signal-Pathways
Complement-abhängigeZytotoxizität
(CDC)
Ziel Zelle
Ziel ZelleZiel Zelle
NK Zelle
Strategie 1Antikörper vermittelte Zytotoxizität
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Binds HER-2 (human epidermal growth factor receptor 2), a growth factor receptor found on some tumor cells (some breast cancers, lymphomas).
Over-expression of HER2 causes increased cell growth and reproduction.
HER2 protein over-expression affects approximately 25% to 30% of breast cancer patients
z.B. Herceptin® (Trastuzumab)
normal cell tumor cell treatment
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HLA-DR
TDT
CD19
CD10
CD20
CD22
CD21
CD38
Stem cell
Pro-B cell
Pre-B cells
ImmatureB cell
MatureB cell
ActivatedB cell
Plasmacell
Neoplasias: Leukemias from B-cell Precursors (B-ALL)
B-cell Lymphomas (NHL, CLL)
Multiple Myeloma
Antigen independent Antigen dependent
Strategie 1: maligne B-Zellen
KV 8.3 30
Strategie 1anti idiotypische Antikörper
(B-Z
ell L
ymp
ho
me)
KV 8.3 31
Strategie 2: Verschiedene Isotope
Nackter Antikörper Radiomarkierter Antikörper
KV 8.3 32
Isotopen markierte Antikörper
LymphoCide (anti CD22, found on some B-cell leukemias.
Tositumomab (Anti-CD20)
Lym-1 (Oncolym). Anti- HLA-DR expressed at high levels on lymphoma cells.
KV 8.3 33
A conjugate of a monoclonal antibody that binds CD33 (cell-surface molecule expressed by the cancerous cells in acute myelogenous leukemia (AML) but not found on the normal stem cells needed to repopulate the bone marrow) and calicheamicin, an oligosaccharide that blocks the binding of transcription factors (proteins) to DNA and thus inhibits transcription.
Strategie 3: Mylotarg® (anti CD-33)
KV 8.3 34
Strategie 4: Enzym markierte Antikörper
Antibody directed enzyme prodrug therapy
(ADEPT)
Tumorantigen
Tumorzelle
ZytotoxischerWirkstoff
Prodrug132
4
Enzym
Antikörper
KV 8.3 35
Strategie 5: Anti-Angiogenes
KV 8.3 36
• Recombinant humanized monoclonal antibody against VEGF; prevents binding with its receptor.
• Blocks stimulation of angiogenesis stops tumor growth.
Anti-VEGF: Avastin ®
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Immunmodulatorische Antikörper
• Anti-TNF Therapie• Infliximab • Etanercept • Humira• etc (Biosimilarika)
• Anti-IgE Therapie• Xolair
KV 8.3 38
Anti-TNF-a: Infliximab
• Binds specifically to TNF-a, neutralizing its activity• Used for Crohn’s disease and rheumatoid arthritis• Side-effect: can convert a latent case of
tuberculosis into active disease
Chimeric monoclonal antibody
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Infliximab: Mechanism of Action Binds and neutralizes soluble and membrane bound TNF
TNFR2
Nucleus
NFB
transcriptionNo Signal
TNF
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Other Postulated Mechanisms of Action…
Antibody dependent cell-mediated cytotoxicity (ADCC)
Lysis of TNF-expressing cells through complement (C’) activation
Complement Activation C’
Complement Receptor
Phagocyte
T CellPhagocytosis
KV 8.3 41
EtanerceptInfliximab
TNF-a: Rezeptor oder Antikörper
KV 8.3 42
Biosimilarika: z.B. anti-TNF
Generic Name
Brand name (source)
Molecular Derivation
Development status
Infliximab
Remicade® Human/mouse chimeric anti-human TNF monoclonal Ab
Approved in US & Europe for CD & RA
Etanercept Enbrel® Human recombinant p75 (TNF receptor fusion protein)
Good for RA
CDP-571 Humicade™ Humanised anti-human TNF mAb
Trials ongoing for CD & ulcerative colitis
CDP-870 N/A(Celltech) Humanised anti-TNF mAb fragment
Phase III trials ongoing
Adalimumab Humira® Human anti-human TNF mAb
Approved for RA
Omalizumab (Xolair®)
BMC
↓ free IgE ↓ FcεRI ↓ Degranulation↓ Inflammation↓ Symptoms
IgE
FcεRI
MediatorRelease
Allergic Inflammation
Holgate S. et al., J Allergy Clin Immunol 2005 vol. 115 (3) pp. 459-65
anti-IgE
OmalizumabReduces IgE and FcεRI Levels
KV 8.3 44
Anti-IgE: Omalizumab (Xolair®)
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Anti Adhäsionstherapie
• Efalizumab (Raptiva®)• Anti-CD11a (subunit of LFA-1) inhibits
activation of T cells
• Alefacept (Amevive®)• binds to CD2 on memory effector T
lymphocytes, inhibiting their activation
• Abciximab (Reopro®)• Anti-Gp IIb/IIIa
KV 8.3 46
Efalizumab (Raptiva)
Interactions between leukocyte-function associated antigen type 1 (LFA-1) and intercellular adhesion molecules are important in the pathogenesis of psoriasis. Efalizumab, a humanized monoclonal antibody, binds to the subunit (CD11a) of LFA-1 and inhibits the activation of T cells
KV 8.3 47
Alefacept (Amevive®)
CD2
Anti-CD2
KV 8.3 48
Anti-GpIIb/IIIa Inhibits the clumping of platelets by binding the receptors on their surface that normally are linked by fibrinogen.
Abciximab (Reopro®)
Helpful in preventing reclogging of the coronaryarteries in patients who have undergone angioplasty.
KV 8.3 49
Ausblick: mehr Antikörper für…
• Transplantation• Knochenmark, Niere, Leber und Pancreas Insel Zellen.
• Krebs• Metastatic Brust Krebs, CLL, non-Hodgkin’s Lymphome.
• Autoimmunkrankheiten• Rheumatoide Arthritis, Psoriasis, Multiple Sclerosis,
Crohn’s Krankheit.
• Infektionskrankheiten• Respiratory syncytial virus, Cytomegalovirus,
Septikämien.
• Allergien: • Mehr als 150 weitere Ak in der Pipeline…
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