Recombinant therapeutic proteins


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Recombinant therapeutic proteins

  2. 2. 1982 Human insulin, created using recombinant DNA technology 1986 1993 Interferon alfa and muromonab-CD3 approved CBER's Office of Therapeutics Research and Review (OTRR) formed First whole chimeric antibody, rituximab, and first humanized antibody, daclizumab, approved $30 billion share of biotechnological drugs of $400 billion in yearly worldwide pharmaceutical sales An inhaled form of insulin (Exubera) approved 1997 2002 2006
  3. 3. Process of Drug Production Transfection Cell culture Purification Formulation/ Filling + Cells and plasmid Cell line Drug substance (crude) Drug substance (pure) Drug product (sterile) Cell line manufacture Bioreactor process Downstream purification Medium developmentdevelopment & scale-up Analytical characterization 5
  4. 4. Therapeutic proteins will form the back-born of future medicinal therapy
  5. 5. BACTERIAL CELLS • simple physiology • short generation times, as bacteria grow and multiply rapidly • large yields of product - up to 10 % of mass (low cost) • The expressed proteins often do not fold properly and so are biologically inactive. • The synthesised protein is often toxic to bacteria preventing the cell cultures from reaching high densities
  6. 6. Yeast cells • Yeast is a simple eukaryote and performs many of the post-translational modifications required for human proteins • Presence of active proteases that degrade foreign (expressed) proteins, thereby reducing their yield • (a solution to this problem is the construction of yeast strains from which the protease genes have been deleted).
  7. 7. INSECT CELLS • High level of expression • Correct folding • More difficult to work with • Expensive • Slow generation time • Not suitable for proteins with repetitive sequences PLANTS • • competitive cost the availability of established practices for their efficient harvesting, transporting, • sorting and processing
  8. 8. • cheap method • produce the desired proteins in vast quantities when using larger animals like cows. • long lead time to generate a herd of transgenic animals • concerns about the health of the animal. • cause serious negative health effects
  9. 9. In vitro systems • E. coli extract; plant wheatgerm extract; and mammalian sources, rabbit reticulocyte lysate. • lack both the genomic material and the cellular boundary system • contain the cellular components required for transcription and/or translation of genes.
  10. 10. Some recombinant proteins approved for human use Protein Company Disorder Factor VIII Baxter, Bayer Hemophilia A Factor IX Genetics Institute Hemophilia B Tissue plasminogen activator (TPA) Genetech Acute myocardial infarction Insulin Eli Lilly, Novo Nordisk Diabetes mellitus Human growth hormone Eli Lilly, Genetech, Upjohn, Novo Nordisk GH deficiency in children (dwarfism) Erythropoietin Amgen, Ortho Biotech Anemia DNase I Genetech Cystic fibrosis Various interferons (IFN) Schering, Biogen, Chiron, Genetech Hepatitis B and C, multiple sclerosis
  11. 11. Drugs in transgenic animals DRUG ANIMAL USED Anti-Cancer Drugs (currently in the process of making). Chickens LACTOFERRIN (Breast Milk Supplement) COWS GENETIC MODIFICIATION The anti-cancer drug is produced in the chickens eggs. Recombinant DNA targets lysosome from the breast milk and modifies it in the cow which gives a more nutritional boost for infants WHO/WHERE PRODUCED Roslin Institute in the United Kingdom is Hermitech, Inc. in Sioux Falls, South Dakota. & China
  12. 12. Schematic representation of the process used to purify ATryn from the milk of transgenic goats.
  13. 13. Recombinant Hepatitis vaccine • The hepatitis B virus (HBV) vaccine – surface antigen purified from the blood of infected individuals. – Due to safety concerns, the HBV vaccine became the first to be produced using recombinant DNA technology (1986) – Produced in bakers’ yeast (Saccharomyces cerevisiae
  14. 14. Vaccine Production at industry level to respond to a human influenza pandemic. • to respond to a human influenza pandemic. •
  15. 15. • single vector containing the coding sequences for both subunit genes. • After transfection, a genetically stable transformant producing biologically active recombinant FSH was isolated • 150–450 gene copies were present in mammalian cells. • FSH products from cell culture supernatants. • collected from a perfusion-type bioreactor • downstream purification
  16. 16. rhDNase I
  18. 18. Bio therapeutics are delicate drugs • Much larger and more complex than traditional pharmaceuticals • Composed of unstable proteins with a precise structure • Easily damaged by unfavorable temperature history during storage