mAb

Presented byGajeshwar prasadM.S (Pharm.)Pharmacology & Toxicologys

Under the guidance of

Dr. (Prof.) Mangala Lahkar,

Chief Academic Coordinator

National Institute of Pharmaceutical Education and Research

INTRODUCTION

Antibodies are group of glycoprotein molecules present in bloodserum and tissue fluid.

These are produced by a specialized group of cells called B-lymphocytes (plasma cells) in mammals.

Antibodies are a part of defence system to protect the body against theinvading foreign substance namely antigen (antibody generator orimmunogen).

Each antigen has specific antigenic determinants (epitopes) sitelocated on it .Antibodies have complementary determiningregion(CDRs) which are mainly responsible for the antibodyspecificity.

In response to an antigen (with several different epitopes), B-lymphocytes gear up and produce many different antibodies.Antibodies which can react with same antigen known as polyclonalantibodies.

STRUCTURE OF ANTIBODY

HISTORY OF MONOCLONAL ANTIBODY

1964 Littlefield developed a way to isolate hybrid cells from 2

parent cell lines using the hypoxanthine-aminopterin-thymidine

(HAT) selection media

1975 Kohler and Milstein provided the most outstanding proof

of the clonal selection theory by fusion of normal and

malignant cells. This resulted in the first monoclonal

antibodies, for which they received the Nobel Prize in 1984.

DISCOVERY!!!

The idea of a "magic bullet" was first proposed by Paul

Ehrlich at beginning of 20th century.

He postulated that if a compound could be made that selectively

targeted a disease-causing organism, then a toxin for that organism could

be delivered along with agent of selectivity.

DISCOVERY!!!

George Kohler & Cesar Milstein in 1975 shared the Nobel Prize in Physiology or Medicine in 1984

for discovery of hybridoma technology.

MONOCLONAL ANTIBODY

They are monospecific antibodies that are the same because they are

made by identical immune cells that are all clones of a unique parent

cell

They have monovalent affinity, in that they bind to the

same epitope.

Given almost any substance, it is possible to produce monoclonal

antibodies that specifically bind to that substance; they can then serve

to detect that substance.

TYPES OF MONO-CLONAL ANTIBODIES

Throughout the

progression of monoclonal

drug development

there have been four

major antibody types

developed:

First generation

murine

Second generation

Chimeric

Humanised

human.

1- According To Evolution 2- According to design

Naked Monoclonal

Antibody

Conjugated Monoclonal

Antibody

Immune-Toxin Monoclonal

Antibody

THE MYELOMA THEORY

In the 1970’s the B-cell cancer myeloma was discovered, and it was understood that these cancerous B-cells all produce a single type of antibody.

This was used to study the structure of antibodies, but it was not possible to produce identical antibodies specific to a given antigen.

MYELOMA CELL

Lost the ability to synthesize hypoxanthine-guanine-

phosphoribosyl transferase (HGPRT).

This enzyme enables cells to synthesize purines using an

extracellular source of hypoxanthine as a precursor.

Ordinarily, the absence of HGPRT is not a problem for the cell

because cells have an alternate (de novo)pathway that they can use

to synthesize purines.

However, when cells are exposed to aminopterin (a folic

acid analog), they are unable to use de novo pathway and are now

fully dependent on HGPRT for survival.

B cell

B cell has the enzyme HGPRT

But B cells die soon

They do not have the capacity to grow indefinitely because of

their limited life span

Scanning Electron Microscopic view of

a B cell

HYBRIDOMA TECHNOLOGY1. Immunisation of a mouse

2. Isolation of B cells from the spleen.

3. Cultivation of myeloma cells.

4. Fusion of myeloma and B cells.

5. Separation of cell lines.

6. Screening of suitable cell lines.

7. in vitro (a) or in vivo (b) multiplication.

8. Harvesting

ADVANTAGES1)Homogeneity:

Monoclonal antibody represents a single antibody molecule that binds to antigens with the same affinity and promote the same effectors functions.

2) Specificity:

• The product of a single hybridoma reacts with the same epitopeon antigens.

3) Immunizing Antigen:

• Need not be characterized and is ultimately not needed in large quantities to produce large quantities of antibody.

4) Selection:

• It is possible to select for specific epitope specificities and generate antibodies against a wider range of antigenic determinants.

5) Antibody Production:

• Unlimited quantities of a single well-defined monospecificreagent

DISADVANTAGES

Average affinity of monoclonal antibodies are generally lower than

polyclonal antibodies

Monoclonals against conformational epitopes on

native proteins may lose reactivity with antigens.

Antibodies sometimes display unexpected

crossreactions with unrelated antigens.

Immune rejections

Time and effort commitment: VERY

LARGE.

ADVERSE EFFECTS

Fever

Chills

Weakness

Headache

Nausea

Vomiting

Diarrhea

Rashes

Hypo/Hypertension

APPLICATIONDiagnostic applications :

Biochemical test –

Pregnancy

Cancers

Hormonal disorder

Infectious diseases

Diagnostic imaging –

Myocardial infraction

Deep vein thrombosis

Cancers

Bacterial infection

Cont.....THERAPEUTIC APPLICATION

MAbs as Direct therapeutic agents –

Pathogenic organisms

Cancer treatment

Immunosuppression

AIDS treatment

Autoimmune diseases

MAbs as targeting agents –

Immunotoxin (cancer)

Drug delivery

Dissolving blood clot

Radioimmunotherapy (of tumors)

Cont.....

Protein purification & drugs

Counteract transplant rejection

NEW METHODS

Nowadays ,two new alternative methods are also available for

production of monoclonal antibody –

Phase Display

Transgenic mice

List of FDA’s Approved Monoclonal antibody

Antibody Brand name Target Indication year

Muromonab-

CD3Orthoclone-

Okt3

CD3; Murine

IgG2a

Reversal of kidney

transplant rejection

1986

Rituximab Rituxan CD20; Chimeric

IgG1

Non-Hodgkin's lymphoma 1997

Infliximab Remicade TNFa; Chimeric

IgG1

Crohn disease 1998

Trastuzumab Herceptin HER2;

Humanized IgG1

Breast cancer 1998

Adalimumab Humira TNF; Human

IgG1

Rheumatoid arthritis 2002

Omalizumab Xolair IgE; Humanized

IgG1

Asthma 2003

Bevacizumab Avastin VEGF;

Humanized IgG1

Colorectal cancer 2004


Antibody Brand name Target Indication year

UstekinumabStelara

IL12/23; Human IgG1 Psoriasis 2009

Tocilizumab Actemra IL6R; Humanized IgG1 Rheumatoid arthritis 2010

Brentuximab

vedotin

Adcetris CD30; Chimeric IgG1;

immunoconjugate

Hodgkin lymphoma, systemic

anaplastic large cell

lymphoma

2011

Pertuzumab Perjeta HER2; humanized IgG1 Breast Cancer 2012

Adotrastuzumab

emtansine

Kadcyla HER2; humanized

IgG1;

immunoconjugate

Breast cancer 2013

Obinutuzumab Gazyva CD20; Humanized

IgG1;

Glycoengineered

Chronic lymphocytic

leukemia

2013

List of FDA’s Approved Monoclonal antibodyAntibody Brand name Target Indication year

SiltuximabSylvant

IL-6; Chimeric IgG1 Castleman disease 2014

Vedolizumab Entyvio α4β7 integrin;

humanized

IgG1

Ulcerative colitis, Crohn

disease

2014

Ramucirumab Cyramza VEGFR2; Human

IgG1

Gastric cancer 2014

Pembrolizumab Keytruda PD1; Humanized

IgG4

Melanoma 2014

Evolocumab PCSK9; Human

IgG2

High cholesterol Review

Secukinumab IL-17a; Human IgG1 Psoriasis Review

Nivolumab PD1; Human IgG4 Melanoma, non-small cell lung

cancer

Review


Antibody Brand

name

Target Indication year

Blinatumomab CD19, CD3; Murine

bispecific tandem scFv

Acute lymphoblastic leukemia Review

Necitumumab EGFR; Human IgG1 Non-small cell lung cancer Review

Mepolizumab IL-5; Humanized IgG1 Severe eosinophilic asthma Review

CONCLUSION

Monoclonal antibodies are Antigen specific, can be produced

against any type of antigen, hence vast diagnostic applications.

Target specificity, a novel therapeutic approach particularly in

cancer.

REFERENCEJustin K.H. Liu* “The history of monoclonal antibody development -

Progress, remaining challenges and future innovations”

T.Mohammad, Funelas C.,and Suria H.“Application of Quantitative

Pharmacology in Development of Therapeutic Monoclonal

Antibodies”

Golay j., Introna M.“Mechanism of action of therapeutic monoclonal

antibodies: Promises and pitfalls of in vitro and in vivo assays”

www. wikipedia.com/monoclonal antibodies

U.satyanararayana, “Biotechnology”

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