Transcript anti-cancer therapy

ANTI-CANCER
THERAPY
MONOCLONAL ANTIBODIES
BY: FIROUZEH KAMALI
Conventional Anti-Cancer Therapy
Chemotherapy: Imperfect
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Systematic nature of cytoxicity
Agents lack intrinsic anti-tumor selectivity
Anti-proliferative mechanism on cells in cycle,
rather than specific toxicity directed towards
particular cancer cell
Host toxicity: treatment discontinued at dose
levels well below dose required to kill all
viable tumor cells
HISTORY
Emil von Behring in 1890
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Discovered antibodies
Paul Ehrlich (16 years later)
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Coined phrase, “magic bullets and poisoned arrows”:
use of antibodies to specifically target toxic
substances in pathogenic substances
Kohler and Milstein in 1975
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Discovery of monoclonal antibodies (mAb) directed
against well-characterized antigens
Use of DNA bio-engineered technologies within last
25 years
Rationale
mAb as efficient carriers for delivery of antitumor agents
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Enhanced vascular permeability of circulating
macromolecules for tumor tissue and subsequent
accumulation in solid tumors
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Normal tissue: blood vessels have intact endothelial layer that
permits passage of small molecules but not entry of
macromolecules (like mAb)
Tumor tissue: blood vessels leaky, so small and large
molecules have access to malignant tissue
-tumor tissue generally do not have a lymphatic drainage
system; therefore, macromolecules are retained and can
accumulate in solid tumors
Patho-physiology of Tumor Tissue
Angiogenesis
Hypervasculature
Impaired lymphatic drainage
***Due to these characteristics, tumors can
be exploited for tumor-selective drug
delivery****
Genetic Engineering
Remove or modify effector functions of mAb: used to
avoid unwanted side effects
Use mAb in their natural, fragmented, chemically
modified, or recombinant forms
Use of phage display antibody libraries or transgenic
animals
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Identify animals that make desired antibodies
Animals must be immunized using the cellular antigens and
immunization procedures used to generate conventional
antibodies
Perform cell fusions to generate clones and isolate stable
clones, making mAb
Most mAb used in the clinical setting were generated
in mice
Structure of Antibody
Presently, all intact therapeutic antibodies are murine
immunoglobulins of the IgG class
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Murine immunoglobulin = glycoprotein that has a Y-shaped
structure: 2 identical polypeptide heavy chains and 2 identical
light chains linked by an S-S bond
Chimeric antibody = genetically engineered construct containing
a mouse Fab portion and a human Fc portion
3 main components
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Two identical Fabs (fragment-antigen binding site): the arms of
the Y
An Fc (for fragment crystallizable), the stem of the Y
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Constant region responsible for triggering effector functions that
eliminate the antigen-associated cells
Constant region must be tailored to match requirements of the antibody
(depending on which antigen you want it to bind to)
IgG structure
3 MECHANISMS RESULTING IN
APOPTOSIS
Antigen cross-linking
Activation of death receptors
Blockade of ligand-receptor growth or
survival pathways
1. Antigen Cross-Linking
Target growth factor receptor
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Antagonize ligand-receptor signaling
Growth-factor signaling mediated by the
receptor tyrosine kinase is inhibited
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EGFR (epidermal growth factor receptor)
IGF-1R (insulin-like growth factor-1 receptor)
FGFR (fibroblast growth factor receptor)
PDGFR (platelet-derived growth factor receptor)
VEGFR (vascular endothelial growth factor)
Results in arrest of tumor cell growth
2. Activation of death receptors
Cross-link targeted surface antigens on
tumor cells and antibody agonists that
mimic ligand-mediated activation of
specific receptors
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Response: intracellular Ca II ions increase
Activate caspase-3 and caspase-9 (involved
in cell apoptosis)
APOPTOSIS PATHWAY
3. Delivery of Cytotoxic Agents
Physically link antibodies to toxic
substances for delivery
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Radio-immunoconjugates (aim of delivering
radiation directly to the tumor)
Toxin-immunoconjugates (deliver toxins
intracellularly)
Antibody-directed enzyme pro-drug therapy
(ADEPT): localize enzymes to tumor cell
surfaces
General Drug Delivery System
Drug molecules
bound to
macromolecule
through spacer
molecule
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Drug released from
macromolecule after
cellular uptake of the
conjugate
Targeting moiety =
monoclonal antibody
TOXIN IMMUNOCONJUGATES
Cell surface antigen must internalize upon mAb binding
When drug is released, it interferes with protein
synthesis to induce apoptosis
3 methods to attach cytotoxic drug to variable regions of
mAb
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a. Couple drug to lysine moieties in the mAb
b. Generation of aldehyde groups by oxidizing the carbohydrate
region and subsequent reaction with amino-containing drugs or
drug derivatives
c. Couple drugs to sulfhydryl groups by selectively reducing the
interchain disulfides near the Fc region of the mAb
Direct attachment of mAb to drug
by S-S bonding
Immunoconjugate
BR96-doxorubicin conjugate
(BR96-DOX)
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Promising toxinimmunoconjugate
mouse/human chimeric mAb
Targets antigen overexpressed on surface of
human carcinoma cells of
breast, colon, lung, and ovary
Disulfide reduction attaches
mAb to drug, BR96
Dose that can be safely
administered every 3 weeks is
insufficient
Other examples of toxinimmunoconjugates
KS1/4-MTX
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Conjugate of methotrexate (MTX)
Coupling of MTX to the lysine moieties of the mAb
No significant clinical response
KS1/4-DAVLB
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Conjugate of vinca alkaloid derivatives
Vinca alkaloid derivatives attached to amino groups of
lysine residues on KS1/4 mAb
No significant clinical response
Why are these toxinimmunoconjugates unsuccessful?
Cause gastrointestinal toxicity
Inner regions of solid tumors poorly
vascularized and have low blood flow
(reduce amount of immunoconjugate
reaching these parts of the tumor)
Antigen expression is heterogenous on
tumor cells
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Restricts the amount of cells that can be
effectively targeted by antibody conjugates
ADEPT ENZYMES (Antibodydirected enzyme pro-drug therapy)
Chemically link the mAb to the enzyme of
interest; can also be a fusion protein produced
recombinantly with the antibody variable region
genes and the gene coding the enzyme
Convert subsequently administered anti-cancer
pro-drugs into active anti-tumor agents
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Upon binding to targeted enzymes, it is converted into
active drug
Anti-growth factor mAb Therapy
Angiogenesis
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Formation of nascent blood vessels
VEGF
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One of the most upregulated antigens in cancer
Protect endothelial cells from apoptosis via activation of PKC
pathways and upregulation of anti-apoptotic proteins such as
Bcl-2
Activity mediated by tyrosine kinase receptors, VEGFR 1 and
VEGFR 2
Functions indirectly as survival factor for tumor cells
Inhibit VEGF signaling
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Block the receptor
Inhibits tumor growth and metastasis
Deprives tumors of nutrient-providing blood vessels
RITUXIMAB (Rituxan)
1st therapeutic mAb approved by FDA in 1997
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High-level expression of the gene encoding Rituximab was found
a mouse-chimeric mAb
Contains the human IgG1 and murine variable regions that target CD20
B-cell antigen
CD20 antigen function: cell cycle progression
Binding Rituximab to CD-20 causes: autophosphorylation, activation of
serine/tyrosine protein kinases, and induction of oncogene expression --induces apoptosis
Response rates of 50% to 70% in follicular lymphomas
Response rates of 90% to 100% when used in combination with
various chemotherpay procedures
Concluded that the dose of 4, once-weekly 375 mg/m squared IV
infusions of Rituximab was safe and effective in patients with
relapse or refractory B non-Hodgkin’s lymphoma
Toxic effects of Rituximab
Short-lived mild
reactions to infusion
after first treatment:
fever, chills, rigors,
rash, and nausea
Factors affecting pharmacokinetic
parameters
Circulating target antigens (which can lead to
rapid clearance)
Antigen-antibody internalization in cells (which
affect serum clearance and half-life)
Antibody size and domains with the Fc region
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Fragments have shorter half-lives and more rapid
clearance rates than their full-sized immunoglobulins
FUTURE
Researchers hope to define the optimal
combinations of the use of mAb with
conventional chemotherapeutic agents
and with radiation therapy
Determine best therapy candidates and
expand clinical trials to other tumor types