MONOCLONAL ANTIBODIES

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MONOCLONAL ANTIBODIES

• SEMINAR PRESENTATION
• Performed by
• PASCHALIS KOURELIAS
• MSc BIOMEDICAL IMMUNOLOGY
• EAST LONDON UNIVERSITY
• STRATFORD 09 /12/2003

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WELCOME ABOARD

• WELCOME TO THE MOST EXCITING DISCOVERY OF MODERN
  MOLECULAR IMMUNOLOGY.
• THE JOURNEY TO THE WORLD OF MONOCLONAL ANTIBODIES
  STARTS JUST NOW !!!!!!!

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WHAT IS AN ANTIBODY ?

• ANTIBODIES or IMMUNOGLOBULINS ARE THE SECRETED
  FORM OF THE B-CELL ANTIGEN RECEPTOR (BCR)
• THEYRE MAIN ROLE IS TO BIND TO A FOREIGN
  ANTIGEN, AND THEN MEDIATE SECONDARY
  EFFECTS(ACTIVATION OF COMPLIMENT
  SYSTEM,RECRUITMENT OF PHAGOCYTES, OPSONIZATION).
• THEY ARE REMARKABLY DIVERSE AND PROVIDE DIFFERENT
  COMBINE SITES TO RECOGNIZE THE MILLIONS OF
  ANTIGENIC SHAPES IN THE ENVIRONMENT.

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WHAT IS MONOCLONAL ANTIBODY? PART 1

• MONOCLONAL ANTIBODIES (m-Abs), ARE Abs WHICH ARE
  GENERATED BY EXPANDING AND IMMORTALIZING SINGLE
  CLONES OF B-CELLS,EACH OF WHICH HAS A DEFINED
  SPECIFICITY.

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WHAT IS MONOCLONAL ANTIBODY ? PART 2

• BECAUSE OF THE DEFINED SPECIFICITY THE mAbs
  RECOGNIZE JUST ONE ONLY EPITOPE ON THE ANTIGEN.
• THE MONOCLONAL Abs WHICH DERIVED FROM ONE CLONE
  ARE ALMOST OF A SINGLE ISOTYPE

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HOW WE CAN MAKE THEM I

• THEY PRODUCED BY CELL LINES OR CLONES OBTAINED
  FROM ANIMALS (mainly mice) THAT HAVE BEEN
  IMMUNIZED WITH THE SUBSTANCE THAT IS SUBJECT OF
  STUDY.
• AN ALTERNATIVE WAY IS TO TRASFORM B-CELLS (HUMAN
  B-CELS)BY INFECTED THEM WITH EPSTEIN-BARR VIRUS.

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HOW WE CAN MAKE THEM II

• TO PRODUCED THE DESIRED mAb, THE CELLS MUST BE
  GROWN IN EITHER OF TWO WAYS
• BY INJECTION INTO THE ABNOMINAL CAVITY OF A
  SUITABLY PREPARED MOUSE.
• BY TISSUE CULTURING CELLS IN PLASTIC FLASKS.

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HOW WE CAN MAKE THEM III
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HOW WE CAN MAKE THEM
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SIGNIFICANCE-USE OF mAbs

• mAbs ARE USED EXTENSIVELY IN BIOMEDICAL RESEARCH.
• IN DIAGNOSIS OF A DISEASE, INFECTION.
• IN TREATMENT OF A DISEASE
• IN TREATMENT OF CANCER
• IN SEROLOGICAL ASSAYS
• AS MARKERS IN IDENTIFICATION OF
  PROTEINS,CARBOHYDRATES,NUCLEIC ACIDS.

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USE OF MONOCLONAL Abs

• MONOCLONAL Abs WERE ENVISAGED AS FORMING A VERY
  SPECIFIC HIGH AFFINITY DELIVERY SYSTEM FOR A
  TOXIN OR RADIOISOTOPE.
• TO DELIVER A DAMAGING STIMULUS TO THE TUMOUR CELL
  OR VIRALLY INFECTED CELL WITHOUT HAVING ANY
  DETRIMENTAL EFFECT ON NORMAL NEIGHBOURING CELLS.

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EFFECTS OF THE USE OF mAb

• BECAUSE OF MICE (FOREIGN ORIGEN) THEY COULD
  INITIATE A XENOGENEIC EFFECT.
• The production of human anti-mouse Abs
• HAMA
• LIMITED USEFULNESS AND APLLICATION AS THERAPEUTIC
  AGENTS.
• The monoclonal Abs rapidly cleared from
  recipients system, by immune response.

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HUMAN MONOCLONAL Abs

• THE PROCESS IS MUCH MORE COMPLEX IN THE
  IMMUNISATION STAGES AND IN FUSION STEP.
• MORE DIFFICULT TO PRODUCE IMMORTALISATION OF
  HUMAN ANTIDODY SECRETING CELLS.
• ALTERNATIVE OPTIONS BECOME NECESSARY, AND BECOME
  AVAILABLE THANKS TO THE MAJOR ADVANCES OF GENETIC
  ENGINEERING AND RECOMBINANT DNA TECHNOLOGY.

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CHIMERIC ANTIBODIES

• GENETIC ENGINEERING AND GENE MANIPULATION
  HUMANISED THE MOUSE MONOCLONAL Abs.
• POSSIBLE TO CLONE A V- region FROM A MOUSE
  ANTIBODY AND EXPRESS THIS GENE ALONG WITH A C-
  region OF THE REQUIRED TYPE.
• XENOGENEIC EFFECT IS LIMITED
• VAST REPERTOIRE OF V-regions OF m-Abs CAN BE
  UTILISED.

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MOSAIC V-REGIONS

• TO FURTHER HUMANISE THE MICE MOBOCLONAL Abs,
  MOSAIC V-region Abs, HAVE BEEN PRODUCED.
• ONLY PORTIONS OF MOUSE ORIGIN ARE THE CDRs.
• HUMAN FWR AND C-regions.
• LESS OF THE MOLECULE IS XENOGENEIC.
• CHOICE OF C-regions ALLOWS THE TAILORINGOF THE
  MOLECULE TO A PARTICULAR PURPOSE

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SUMMARY

• MONOCLONAL Abs DERIVED FROM JUST ONLY A SIGLE
  CLONE. ALWAYS FROM THE SAME ISOTYPE.
• GENERATE FROM MICE BY FUSING SPLEEN CELLS WITH A
  CANCER CELL (MYELOMA) or
• BY USING IN VITRO TECHNIQUES.
• HUMAN m-Abs ARE VERY LIMITED IN PRODUCTION DUE TO
  SEVERAL DIFFICULTIES.
• THEY HAVE A GREAT USE IN MOLECULE MEDICINE. IN
  DIAGNOSIS-TREATMENT OF DISEASES INCLUDING CANCER.
• SEVERAL METHODS HAVE BEEN DEVELOPED FOR HUMANISED
  THE MICE MONOCLONAL Abs.
• CHIMERIC ANTIBODIES, MOSAIC V-regions,

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ANY QUESTIONS ?

• ANY QUESTIONS ????

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REFERENCES

• 1.Day,NE. Varghese C. (2001).25 years of
  monoclonal Antibodies. Immunology Today. Volume
  32, No 8.
• 2.Avidan B, Sonneberg A.(2002). Role of
  monoclonal antibodies in molecular medicine.
• Journal of Immunology. Vol 971130-1135.
• 3. Blot W.J, Gefferson B. (2001). Humanised mice
  antibodies. Current opinion in Immunology35403-40
  9.
• 4. Gammon. MD.(2000).Monoclonal antibody. New
  England Journal Med. 405362-365.