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DNA vaccine and Biopharmaceutical protein Production

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  1. DNA vaccine and Biopharmaceutical protein Production Theerapol Sirinarumitr DVM, Ph.D. Department of Pathology, Faculty of Veterinary Medicine Kasetsart University

  2. Transcription Transcription Translation Translation Background Prokaryotes Eukaryotes

  3. HISTORY • Skeletal muscle cell can uptake naked DNA (plasmid carried genes) and expressed those genes (Science 247:1465-1468, 1990 by Wolff et al.)

  4. Advantages • Readily delivery • Molecularly defined reagent • Neither infectious nor capable of replication • Elicited both humoral and cell-mediated immune response

  5. Advantages • Manipulate the resulting immune response through the co-deliverly of genes encoding immunodulating cytokines or costimulatory molecules • site-directed mutagenesis for enhancing antigenic potency of individual epitope or to abolish unwanted effects of wild-type protein

  6. Purpose of vaccination • Prophylaxis • Therapeutic • infectious agent • cancer

  7. Gene delivery and Expression • Routes of administration • epidermal, mucosal, intramuscular and intravenous • Skeletal muscle and skin (Langerhans cell) • Plasmid DNA persisted episomally in myocytes

  8. Intramuscular injection and particle-mediated delivery of plasmid DNA • Particle-mediated delivery required up to 5000 times less DNA than intramuscular or intradermal inoculation of DNA (nanogram vs 25-100 microgram)

  9. Transfection efficiency does not necessarily correlate with the efficiency of immunization • Intradermal injection (low transfection efficiency) can elicit potent immune response when compared to muscle (higher transfection efficiency)

  10. Site and method of DNA delivery may affect the nature of the immune response (skeletal muscle and skin) LC and keratinocyte Proinflammatory cytokines: IL-1, TNF-a & GM-CSF Take up and process antigen Muscle can not *Dendritic cell **10 LC

  11. ID or IM injection induce Th1 bias of the immune response INF-G-producing CD4+ IgG2a isotype • Gene gun (skin or muscle) induces immune responses with Th2 bias

  12. Muscle uptake Muscle control

  13. DNA vaccine’s immunology

  14. Processing and Presentation (8-12 amino acids)

  15. 12-25 amino acid

  16. Protein-based vaccine elicitation of strong CTL

  17. Th1-type responses CD8+ CTL CD4+ Th1 cells IgG2a>IgG1 IFN-g>IL-4,IL-5,IL-10 IL-2, IL-12 Th2-type responses DTH IL-4 and IL-10

  18. Influenza, rabies, HIV, HBV, HCV & HSV • Mycobacterium tuberculosis, Mycoplasma pulmonis, and Borrelia burgdorferi • Malaria and Leishmania

  19. Genetic engineering of immune response

  20. Engineering Immune Response • Costimulatory molecule • Immunostimulatory cytokine • Immunostimulatory DNA sequences (ISS) • Biological adjuvant