Chapter 11 Medical Biotechnology and Gene Therapy

1. Chapter 11Medical Biotechnology and Gene Therapy BTEC 3301

2. How is biotechnology used in Medicine? Biotechnology is responsible for hundreds of medical diagnostic tests that detect disease earlier, enduring a more accurate, beneficial and cost-effective outcome to disease.

3. How is biotechnology used in Medicine? • There are more than 350 biotechnology-based drug products and vaccines currently in human trials and more than 100 therapeutic drug products already on the market, with many more in development. • Some of these drugs are designed to treat cancer, heart disease, diabetes, multiple sclerosis, HIV-AIDS and obesity.

4. Therapeutic proteins • Proteins made by genetic engineering processes that patients cannot make themselves due to defective genes • Therapeutic proteins can be produced by • Bacteria • Animals • Plants

5. With genetic manipulation, pigs, cows, goats, and other animals can produce human proteins in its milk • Proteins can also be engineered in eggs • Drug delivery is simple and inexpensive

6. Why Hunt for Human Gene Disorders? • 6 dysfunctional genes per person • 30% pediatric admissions to hospital due to genetic problems • 12% adult admissions to hospital due to genetic problems • 70-75% of products, prevention and gene therapy goes for human species • There are 4000 known genetic disorders and of these >200 are enzymatic defects

7. Few common examples of human diseases caused by defective genes:

8. Genetic Engineering of a Milk Protein

9. What is Gene Therapy? It encompasses repairing or replacing defective genes and making tumors more susceptible to other kinds of treatment. Thus, gene therapy’s potential for preventing and curing disease is vast

10. What is Gene Therapy? • Gene therapy is the insertion of genetic material into cells to prevent, control, or cure disease, especially genetic disorders. • Gene therapy is "a therapeutic technique in which a functioning gene is inserted into the somatic cells of a patient to correct an inborn genetic error or to provide a new function to the cell"

11. What is Gene Therapy? • There have been many human gene therapy clinical trials, involving over 700 patients world-wide (by 1996), for several different diseases including several cancers. • In the USA the trials must be approved by the Recombinant DNA Advisory Committee (RAC) and the FDA

12. What is Gene Therapy? Gene therapy is a technique for correcting defective genes responsible for disease development. Researchers may use one of several approaches for correcting faulty genes Carriers of therapeutic genes include:

13. What is Gene Therapy? Carriers of therapeutic genes include: • harmless viruses that have undergone genetic alteration and can carry selected genetic material into human cells; and • liposomes—injectable microscopic fatty globules that can enclose and protect DNA segments (e.g., a "suicide gene" for insertion into cancer cells.) • Genes, which are carried on chromosomes, are the basic physical and functional units of heredity.

14. What is Gene Therapy? Carriers of therapeutic genes include: • Genes are specific sequences of bases that encode instructions on how to make proteins. • Although genes get a lot of attention, it’s the proteins that perform most life functions and even make up the majority of cellular structures. • When genes are altered so that the encoded proteins are unable to carry out their normal functions, genetic disorders can result.

15. How it done? • A normal gene may be inserted into a nonspecific location within the genome to replace a nonfunctional gene. This approach is most common. • An abnormal gene could be swapped for a normal gene through homologous recombination. • The abnormal gene could be repaired through selective reverse mutation, which returns the gene to its normal function. • The regulation (the degree to which a gene is turned on or off) of a particular gene could be altered.

16. Two types of Gene Therapy "platforms": 1. Ex vivo therapies: • Ex vivo therapies involve treating cells that have been removed from a patient with a functional gene to restore protein activity. The cells are then returned to the patients to have a therapeutic benefit. (ADA, sickle cell disease - blood cell diseases or immune system diseases)

17. Two types of Gene Therapy "platforms": 2. In vivo therapies: • The use of in vivo therapies generally involves injection or inhalation of a product that contains a therapeutic gene together with some sort of gene delivery system.

18. How does gene therapy work? Gene Delivery System: • Therapeutic genes and • an efficient gene delivery system • a way to get the correct genes into the correct cells .

19. How does gene therapy work? Gene Delivery System: • Viruses attack their hosts to insert their genetic material into the genetic material of the host. • This genetic material contains instructions to produce these viruses. The host cell will carry out these instructions and produce the viruses. This is how viruses spread, in general.

20. How does gene therapy work? Gene Delivery System: • In addition to the instructions producing the components of the virus itself, viruses can carry additional genes containing instructions for creating other kinds of proteins. • In theory, if we insert a gene that is missing from a patient in a virus, and infect that patient with the virus, the virus will spread the missing gene in all the cells of the patient. • The missing gene is now replaced and the disease is cured

21. How does gene therapy work? The 7 requirements of a successful gene delivery system: • A practical gene delivery system (the vector) must meet seven characteristics: • Efficient in introducing the genes to recipient cells • Capable of achieving long term expression or short term expression (depending on use) • Flexible with respect to the target tissues it can deliver to

22. How does gene therapy work? The 7 requirements of a successful gene delivery system: • A practical gene delivery system (the vector) must meet seven characteristics: • Harmless to the patient • Able to handle a wide range of therapeutic genes • Able to demonstrate a dose-response relationship • Supplied in a form familiar and comfortable to the medical community. The biggest hurdle to overcome (still) of gene therapy is finding effective, safe vectors

23. Types of vectors Types of viruses are currently used as vectors in gene therapy: retroviruses, adenoviruses adeno-associated viruses and Herpes simplex viruses They differ in their mechanisms of action and results:

24. Types of vectors viruses are currently used as vectors in gene therapy: • Retroviruses – • A class of viruses that can create double-stranded DNA copies of their RNA genomes. These copies of its genome can be integrated into the chromosomes of host cells. Human immunodeficiency virus (HIV) is a retrovirus

25. Types of vectors Three types of viruses are currently used as vectors in gene therapy: • Adenoviruses – • A class of viruses with double-stranded DNA genomes that cause respiratory, intestinal, and eye infections in humans. The virus that causes the common cold is an adenovirus.

26. Types of vectors Three types of viruses are currently used as vectors in gene therapy: • Adeno-associated viruses – • A class of small, single-stranded DNA viruses that can insert their genetic material at a specific site on chromosome 19.

27. Types of vectors Three types of viruses are currently used as vectors in gene therapy: • Herpes simplex viruses– • A class of double-stranded DNA viruses that infect a particular cell type, neurons. Herpes simplex virus type 1 is a common human pathogen that causes cold sores & eye infection

28. Non-virus-mediated delivery system Direct introduction: • There are several nonviral options for gene delivery. • The simplest method is the direct introduction of therapeutic DNA into target cells. • This approach is limited in its application because it can be used only with certain tissues and requires large amounts of DNA.

29. Non-virus-mediated delivery system Artificial lipid: • Another nonviral approach involves the creation of an artificial lipid ,a liposome, which carries the therapeutic DNA which delivers the DNA through the target cell's membrane.

30. Non-virus-mediated delivery system Special cell receptors as chemical links: • Therapeutic DNA also can get inside target cells by chemically linking the DNA to a molecule that will bind to special cell receptors. • Once bound to these receptors, the therapeutic DNA constructs are engulfed by the cell membrane and passed into the interior of the target cell (Less effective).

31. What is the current status of gene therapy research? • The Food and Drug Administration (FDA) has not yet approved any human gene therapy product for sale. • Current gene therapy is experimental and has not proven very successful in clinical trials. • Little progress has been made since the first gene therapy clinical trial began in 1990

32. What is the current status of gene therapy research? • In 1999, gene therapy suffered a major setback with the death of 18-year-old Jesse Gelsinger. Jesse was participating in a gene therapy trial for ornithinetranscarboxylase deficiency (OTCD) also known as Urea cycle. • Jesse died from multiple organ failures 4 days after starting the treatment. His death is believed to have been triggered by a severe immune response to the adenovirus carrier.

33. The urea cycle is the sole source of endogenous production of arginine and it is the principal mechanism for the clearance of waste nitrogen resulting from protein turnover. http://www.ornl.gov/sci/techresources/Human_Genome/medicine/genetherapy.shtml

34. What is the current status of gene therapy research? • Another major blow came in January 2003, when the FDA placed a temporary halt on all gene therapy trials using retroviral vectors in blood stem cells. • Gene Therapy X-SCID Poses Substantial Cancer Risk

35. What is the current status of gene therapy research? • FDA took this action after it learned that a second child treated in a French gene therapy trial had developed a leukemia-like condition. • Another child developed leukemia-like conditions (August 2002 ) when tested for combined immunodeficiency disease (X-SCID), also known as "bubble baby syndrome: • “http://www.medicalnewstoday.com/articles/42339.php

36. What factors have kept gene therapy from becoming an effective treatment for genetic disease? Short-lived nature of gene therapy: • Before gene therapy can become a permanent cure for any condition, the therapeutic DNA introduced into target cells must remain functional and the cells containing the therapeutic DNA must be long-lived and stable.

37. What factors have kept gene therapy from becoming an effective treatment for genetic disease? Short-lived nature of gene therapy: • Problems with integrating therapeutic DNA into the genome and the rapidly dividing nature of many cells prevent gene therapy from achieving any long-term benefits. Patients will have to undergo multiple rounds of gene therapy.

38. What factors have kept gene therapy from becoming an effective treatment for genetic disease? Immune response : • Anytime a foreign object is introduced into human tissues, the immune system is designed to attack the invader. • The risk of stimulating the immune system in a way that reduces gene therapy effectiveness is always a potential risk. • Furthermore, the immune system's enhanced response to invaders makes it difficult for gene therapy to be repeated in patients.

39. What factors have kept gene therapy from becoming an effective treatment for genetic disease? Problems with viral vectors: • Viruses, while the carrier of choice in most gene therapy studies, present a variety of potential problems to the patient -toxicity, immune and inflammatory responses, and gene control and targeting issues. • In addition, there is always the fear that the viral vector, once inside the patient, may recover its ability to cause disease and become virulent

40. What factors have kept gene therapy from becoming an effective treatment for genetic disease? Multigene disorders : • Conditions or disorders that arise from mutations in a single gene are the best candidates for gene therapy. • Unfortunately, some the most commonly occurring disorders, such as heart disease, high blood pressure, Alzheimer's disease, arthritis, and diabetes, are caused by the combined effects of variations in many genes. • Multigene or multifactorial disorders such as these would be especially difficult to treat effectively using gene therapy.

41. What are some of the ethical considerations for using gene therapy? Some Questions to Consider............... • What is normal and what is a disability or disorder, and who decides? • Are disabilities diseases? Do they need to be cured or prevented? • Does searching for a cure demean the lives of individuals presently affected by disabilities?

42. What are some of the ethical considerations for using gene therapy? Some Questions to Consider............... • Is somatic gene therapy more or less ethical than germline gene therapy? • Preliminary attempts of gene therapy are exorbitantly expensive. • Who will have access to these therapies? Who will pay for the cost ?

44. Gene Therapy Links and Research Institutes: For more information on different types of genetic disease and gene therapy: Must visit these sites to enrich your knowledge about what are being done around you, if not for the present but for our children:

45. Reading Materials • It is a MUST that you read the rest of the slides which gives more examples of gene therapies and disasters • What are some recent developments in gene therapy research?

46. What are some recent developments in gene therapy research? Some Questions to Consider............... • University of California, Los Angeles, research team gets genes into the brain using liposomes coated in a polymer call polyethylene glycol (PEG). The transfer of genes into the brain is a significant achievement because viral vectors are too big to get across the "blood-brain barrier." This method has potential for treating Parkinson's disease. See Undercover genes slip into the brain at NewScientist.com (March 20, 2003).

47. What are some recent developments in gene therapy research? Some Questions to Consider............... • RNA interference or gene silencing may be a new way to treat Huntington's. Short pieces of double-stranded RNA (short, interfering RNAs or siRNAs) are used by cells to degrade RNA of a particular sequence. If a siRNA is designed to match the RNA copied from a faulty gene, then the abnormal protein product of that gene will not be produced. See Gene therapy may switch off Huntington's at NewScientist.com (March 13, 2003).

48. What are some recent developments in gene therapy research? Some Questions to Consider............... • New gene therapy approach repairs errors in messenger RNA derived from defective genes. Technique has potential to treat the blood disorder thalassaemia, cystic fibrosis, and some cancers. See Subtle gene therapy tackles blood disorder at NewScientist.com (October 11, 2002).

49. What are some recent developments in gene therapy research? Some Questions to Consider............... • Gene therapy for treating children with X-SCID (sever combined immunodeficiency) or the "bubble boy" disease is stopped in France when the treatment causes leukemia in one of the patients. See 'Miracle' gene therapy trial halted at NewScientist.com (October 3, 2002).

50. What are some recent developments in gene therapy research? Some Questions to Consider............... • Researchers at Case Western Reserve University and Copernicus Therapeutics are able to create tiny liposomes 25 nanometers across that can carry therapeutic DNA through pores in the nuclear membrane. See DNA nanoballs boost gene therapy at NewScientist.com (May 12, 2002).