Gene therapy

Gene therapy is the process of inserting genes into cells to prevent, treat or cure wide range of diseases. Gene therapy primarily involves genetic manipulations in animals or humans to correct a disease. Gene augmentation therapy: a DNA is inserted into the Genome to replace the missing gene product.Gene inhibition therapy: the antisense gene inhibits the expression of the dominant gene.

1. GENE THERAPY Presented by, Pavan k a M.pharm, Pharmacology COPS, DayanandA sagar UNIVERSTY 1

2. contents Introduction Approaches Types of Gene Therapy Vectors used in gene therapy Applications Recent advancements in gene therapy 2

3. INTRODUCTION Gene therapy is the process of inserting genes into cells to prevent, treat or cure wide range of diseases. Gene therapy primarily involves genetic manipulations in animals or humans to correct a disease. Gene augmentation therapy : a DNA is inserted into the Genome to replace the missing gene product. Gene inhibition therapy : the antisense gene inhibits the expression of the dominant gene. 3

4. 4

5. 5

6. Somatic cell gene therapy Germ cell gene therapy Non reproductive cell / Somatic cells. These are the cells of an organism other than sperm or egg cells, e.g., bone marrow cells, blood cells, skin cells, intestinal cells. somatic cell gene therapy involves the insertion of a fully functional and expressible gene into a target somatic cell to correct a genetic disease permanently. The genetic alterations in somatic cells are not carried to the next generations. Therefore, somatic cell gene therapy is preferred . The reproductive (sex) cells of an organism constitute germ cell line. Gene therapy involving the introduction of DNA into germ cells is passed on to the successive generation. For safety, ethical and technical reasons, germ cell gene therapy is not being attempted at present. Approaches in gene therapy 6

7. Types of gene therapy Ex vivo gene therapy : This involves the transfer of genes in cultured cells (ex: bone marrow cells) which are then reintroduced into the Patient. In vivo gene therapy : The direct Transfer of genes into the cells of a particular tissue is referred to as in vivo gene therapy. 7

8. EX-VIVO GENE THERAPY The ex vivo gene therapy can be applied to only selected tissues (e.g., bone marrow) whose cells can be cultured in the laboratory. Steps involved 1.lsolate cells with genetic defect from a patient. 2.Grow the cells in culture. 3.Introduce the therapeutic gene to correct gene defect. 4.Select the genetically corrected cells (stable transformants) and grow. 5.Transplant the modified cells to the patient. 8

9. The first human gene therapy National Institutes of Health in Bethesda, Maryland, in 1990, Performed on a 4yr old girl Ashanthi DeSilva. suffering from SCID- Severe Combined Immunodeficiency. Caused due to defect in gene coding for ADA. Deoxy adenosine accumulate and destroys T lymphocytes. Disrupts immunity , suffer from infectious diseases and die at young age. Correct the deficiency of enzyme, Adenosine deaminase (ADA). Bone marrow cells from the child were transformed with an engineered retrovirus containing a functional ADA gene. The treated cells were reintroduced into the patient’s marrow. Four years later, the child was leading a normal life 9

10. synthetic ADA was administered in a complex with polyethylene glycol (PEG). For many ADA-SCID patients, injection of the ADA-PEG complex allowed some immune system development, with weight gain and reduced infection, although not full immune reconstitution. trial participants received both treatments at once, making it unclear which treatment was primarily responsible for the positive clinical outcome. 10

11. IN-VIVO GENE THERAPY The direct delivery of the therapeutic gene (DNA) into the target cells of a particular tissue of a patient constitutes in vivo gene therapy. Many tissues are the potential candidates for this approach. These include liver, muscle, skin, spleen, lung, brain and blood cells. Gene delivery can be carried out by viral or nonviral vector systems. It can be the only possible option in patients where individual cells cannot be cultured in vitro in sufficient numbers (e.g. brain cells). 11

12. In vivo gene transfer is necessary when cultured cells cannot be re-implanted in patients effectively. The success of in vivo gene therapy mostly depends on the following parameters. The efficiency of the uptake of the remedial (therapeutic) gene by the target cells. lntracellular degradation of the gene and its uptake by nucleus. The expression capability of the gene. 12

13. Example: In patients with cystic fibrosis, a protein called cystic fibrosis transmembrane regulator (CFTR) is absent due to a gene defect. In the absence of CFTR chloride ions concentrate within the cells and it draws water from surrounding. This leads to the accumulation of sticky mucous in respiratory tract and lungs. treated by in vivo replacement of defective gene by adenovirus vector . 13

14. VECtors in gene therapy The carrier particles or molecules used to deliver genes to somatic cells are referred to as vectors. 2 main classes Viral vectors Non viral vectors 14

15. Viral vectors The vectors frequently used in gene therapy are viruses, particularly retroviruses. RNA is the genetic material in retroviruses. As the retrovirus enters the host cell, it synthesizes DNA from RNA (by reverse transcription). The so formed viral DNA (referred to as provirus)gets incorporated into the DNA of the host cell. Risk Factor: some of the retroviruses can convert normal cells into cancerous ones. Therefore, it is absolutely essential to ensure that such a thing does not happen. Many viral vector systems have been developed for gene delivery These include retroviruses, adenoviruses, adeno-associated viruses and herpes simplex virus. 15

16. Non viral vectors There are certain limitations in using viral vectors in gene therapy. In addition to the prohibitive cost of maintaining the viruses, the viral proteins often induce inflammatory responses in the host. Pure DNA constructs that can be directly introduced into target tissues. Lipoplexes , lipid-DNA complexes that have DNA surrounded by lipid layers. Human artificial chromosome which can carry large DNA (one or more therapeutic genes). 16

17. Clinical applications Bone repair: Bone loss caused by trauma, neoplasia, reconstructive surgery, congenital defects or periodontal disease is a major worldwide health problem. The bone morphogenic proteins (BMPs) enable skeletal tissue formation during embryogenesis, growth, adulthood, and healing. Probably BMPs (BMPs 2, 4 and 7) are the only growth factors which can singly induce de novo bone formation both in vitro and at heterotopic sites. Bone defects in the oral and maxillofacial region can be repaired by transferring genes encoding BMP‘s . Oral Cancer: The general strategy in cancer treatment is to express a gene product that will result in cancer cell death. It can be achieved by Addition of a tumor-suppressor gene, Deletion of a defective tumor gene, Introduction of genes to inhibit tumor angiogenesis and  "Cancer vaccination" with genes for tumor antigens. 17

18. Gene therapy to grow new teeth : This approach is generally presented in terms of adding molecules to induce de novo tooth initiation in the mouth. It might be combined with gene-manipulated tooth regeneration; that is, endogenous dental cells in situ can be activated or repressed by a gene-delivery technique to produce a tooth. More than 200 genes are known to be expressed during tooth development. Cardiac disease: Gene therapy has been investigated to target angiogenesis ( the formation of new blood vessels) during cardiac surgery and to improve calcium handling mechanism in heart failure. Infectious disease: gene therapy vaccines are being developed and trailed for tackling infectious diseases including tuberculosis, malaria, HIV and influenza. 18

19. Recent advancements in gene therapy 2017 Was the Year of Gene-Therapy Breakthroughs Sickle-cell cure In March, researchers announced that a teenage boy in France had been  cured of sickle-cell disease  after receiving an experimental gene therapy developed by Bluebird Bio. Scientists removed stem cells from the boy’s bone marrow and modified them in the lab by introducing copies of a gene to prevent his red blood cells from becoming “sickled.” When the treated cells were infused back into his body, they began to make normal blood cells. Restoring sight  In December, the FDA approved the first gene therapy for an inherited disease. The treatment, called Luxturna, aims to correct a mutation responsible for a range of retinal diseases that make people gradually go blind. In human tests, the  treatment has restored vision  for more than two dozen patients who were losing their sight. 19

20. Cancer treatment: The FDA calls the treatment, made by Novartis, the “ first gene therapy ” in the U.S. The therapy is designed to treat an often-lethal type of blood and bone marrow cancer that affects children and young adults. Known as a CAR-T therapy, the approach has shown remarkable results in patients. Kymriah treats a bone marrow cancer that affects children and young adults, and Yescarta treats a type of lymphoma.  Hemophilia : BioMarin is one company working on a gene therapy that replaces the faulty gene involved in the most common type of hemophilia, effectively curing the disorder. In December, the company published  early clinical trial results  showing that nine patients who received its therapy saw substantial increases in the blood-clotting proteins absent in hemophilia.  20

21. During 2016, Italian scientists at Milan’s San Raffaele Telethon Institute for Gene Therapy  reported  that they had cured 18 children of a rare but terrible immune deficiency disease, ADA-SCID . They removed the children’s bone marrow, added a gene to make the ADA enzyme their bodies lack, and replaced it.  Technology Review  explained  how the treatment, now called Strimvelis and owned by Glaxo, took 14 years to develop and test. It was approved in Europe in May of this year. 21

22. REFERENCES Biochemistry by U. Satyanarayana , U. Chakrapani 3 rd edition http://www.rroij.com/open-access/gene-therapy-principles-and-applications-in-dentistry-5-12.php?aid=34559 https://journals.sagepub.com/doi/full/10.1177/0192623307309925 www.slideshare.com https://academic.oup.com/hmg/article/5/Supplement_1/1397/661061 22

23. 23 THANK YOU