Genetics 34: Gene Therapy and Treatment 2023 PDF

34: Gene Therapy and Treatment of Genetic Disease Robin T. Varghese Ph.D. [email protected] Learning Objective 1. 2. 3. 4. 5. 6. Identify limitations of precision medicine. Identify RNAi as a therapeutic tool. Identify genome editing technologies and their potentials. Distinguish between genome...

34: Gene Therapy and Treatment of Genetic Disease Robin T. Varghese Ph.D. [email protected] Learning Objective 1. 2. 3. 4. 5. 6. Identify limitations of precision medicine. Identify RNAi as a therapeutic tool. Identify genome editing technologies and their potentials. Distinguish between genome editing technologies. Identify CRISPR components and applications. Identify the challenges of genome editing What is a genetic disease? • Genetic disorders occur when an inheritable alteration affects your genes or when you have the wrong amount of genetic material Types: • Chromosomal disorders (e.g., Down syndrome, Turner’s syndrome) • Single gene (e.g., Sickle cell disease, Cystic fibrosis) • Multifactorial (e.g., schizophrenia, cancer) • Mitochondrial (e.g., LHON, MERRF) 2019 2021 2022 6,501 4,150 7,001 4,526 7,268 4,704 https://www.omim.org/statistics/geneMap The Molecular Analysis for Therapy Choice (NCI-MATCH) Precision medicine looks at the genetics, environment, and lifestyle of a person to select treatment that could work best for them. • Rapidly advancing across health landscape • Increased public awareness and acceptance At the same time, we need to be cautious – not only in ensuring the safety and efficacy of health technologies but also exaggerated optimism. Why such limitations in precision medicine? • Most of the human genome isn’t being actively studied – There are around 20,000 human protein-coding genes, but recent studies have suggested scientists actively study only about 2,000 of them – So, there is lots we don’t know • We may understand the genetic lesion that is the cause of a disease or trait, but there is no therapy – e.g., there is no clinical action-ability • Expensive, ( BUT price of sequencing is dropping quickly) – But the time/cost of accurately interpreting sequencing results is still high • Still exploring more recent precision medicine technologies which involve gene therapy and genome editing. How do we currently treat genetic diseases? Strategies for treating genetic disease • Clinicians and patients frequently treat symptoms e.g., – Codeine for pain management – Organ transplants e.g., – Lung transplants for Cystic fibrosis patients – Bone marrow transplantation, to treat certain blood and immune system disorders • Dietary and Lifestyle changes (proper nutrition, weight control, exercise) can address the causes of diseases like diabetes, hypertension • Causes of genetic diseases can also be targeted/ treated – Gene therapy is specifically designed to treat the molecular cause of the disease, e.g., – RNA interference (RNAi) – Genome editing e.g., CRISPR Strategies for treating genetic disease Management of Genetic Disorders Lifestyle Adjustments • Familial hypercholesterolemia: Exercise and a low-fat diet • Hereditary pancreatitis: avoid smoking and alcohol • Hereditary Cancer syndromes: Frequent cancer screening and caution w/ DNA damaging factors Management of Genetic Disorders Dietary Restrictions • Celiac Disease-Gluten is a protein in rye, barley, and wheat. For people with celiac disease the gluten irritates the gut, leading to inflammation that goes into overdrive and can attack the intestinal villi. — remove gluten from diet. • Galactosemia—remove galactose from diet • Phenylketonuria (PKU) –restrict dietary phenylalanine OR treat with supplemental BH4 (depending on gene mutation) Strategies for treating genetic disease Supply the missing product to manage disease • Congenital Hypothyroidism: a partial or complete loss of function of the thyroid gland that affects infants from birth à supply thyroxine as treatment • Hemophilia: caused by various mutations in the clotting cascade, which leads to poor blood clotting following injury. à supply concentrates of clotting factor VIII (for hemophilia A) or clotting factor IX (for hemophilia B) • Type 1 Diabetes: loss of beta islet cells from the pancreas à replacement with insulin • Vitamin D-dependent rickets: is a disorder in the biosynthesis of vitamin D or its receptor activity which affects bone development i.e., leads to softening and weakening of the bones. àAdminister vitamin D as treatment. What is gene therapy? Gene therapy seeks to modify or manipulate the expression of gene/s to produce a therapeutic effect. • Replacing a mutated gene that causes disease with a healthy copy of the gene. • Reducing (“knocking down”) gene expression or Inactivating (“knocking out),” a mutated gene that is functioning improperly. • Introducing a new gene into the body to help fight a disease. • Gene editing – a permanent manipulation of a gene in a patient’s genome Although gene therapy is a promising treatment option for several diseases (including inherited disorders, some types of cancer, and certain viral infections), the technique remains risky and is still under study to make sure that it will be safe and effective. Gene therapy is currently being tested only for diseases that have no other cures. https://ghr.nlm.nih.gov/primer/therapy/genetherapy Strategies for treating genetic disease How does gene therapy work? Gene therapy is designed to introduce, remove, or change genetic material into cells to compensate for abnormal genes or to make a beneficial protein with the intention of direct therapeutic effect. The delivery of genetic material into cells can be accomplished by multiple methods. The two major classes are recombinant viruses (viral vectors) and non-viral (e.g., naked DNA, electroporation, nanoparticles). Gene Therapy September 14, 1990- NIH performed 1st approved gene therapy procedure: • Ashanti DeSilva a 4-year girl born with adenosine deaminase (ADA) deficiency, a form of severe combined immunodeficiency (SCID) • People with SCID lack virtually all immune protection from pathogens and are prone to persistent infections that can be life-threatening • Without treatment, children usually do not survive past age 2 • Ashanti received a healthy version of the gene that produces ADA using a viral vector and she is still alive today! • The trial seemed successful, and many more trials were conducted in SCID and other diseases. Strategies for treating genetic disease The Death of Jesse Gelsinger • • • • • 1999 - first person identified as having died in a clinical trial (U. Penn) for gene therapy Gelsinger suffered from ornithine transcarbamylase deficiency(OTC gene), an X-linked genetic disease of the liver, the symptoms of which include an inability to metabolize ammonia – a byproduct of protein breakdown. Four days after injection with adenoviral vector carrying a corrected gene Jesse died of immune response Controversy – FDA determined co-investigator Dr. James M. Wilson (Director of the Institute for Human Gene Therapy) broke several rules of conduct: – Inclusion of Gelsinger as a substitute for another volunteer who dropped out, despite Gelsinger's having high ammonia levels that should have led to his exclusion from the trial. – Failure by the university to report that two patients had experienced serious side effects from the gene therapy. – Failure to disclose, in the informed-consent documentation, the deaths of monkeys given a similar treatment. Both Wilson and the University are reported to have had financial stakes in the research. Examples of Gene Therapy- Replacing Gene Product • Luxturna (voretigene neparvovec-rzyl): adeno-associated virus vector-based gene therapy indicated for Leber congenital amaurosis (LCA) with confirmed biallelic RPE65 mutation-associated retinal dystrophy • w/ viable retinal cells • $425,000 per eye **First in-vivo gene therapy approved by FDA https://www.zolgensma-hcp.com/patient-profiles/start-trial-patient-stories/ • Zolgensma (onasemnogene abeparvovec-xioi) adeno-associated virus vector-based gene therapy Treatment of Spinal Muscular Atropy (Type 1) • One time treatment for the SMN1 gene • $2.125 million (USD) per treatment, making it the most expensive medication in the world as of June 2022 Find full list on: https://www.fda.gov/vaccines-blood-biologics/cellular-gene-therapy-products/approved-cellular-and-gene-therapy-products 2006 Nobel Prize in Physiology/Medicine "for their discovery of RNA interference - gene silencing by double-stranded RNA" Andrew Z. Fire RNA interference- Gene Knockdown Craig C. Mello RNA Interference- Gene Knockdown Largely, can be used as a research tool to characterize the effects of knocking down single genes to identify therapeutic potential siRNA-based therapy • Only 4 small interfering RNA based therapies are FDA approved: • Patisiran (2018)- hereditary transthyretin amyloidosis (hATTR) (1st siRNA-based drug approved by FDA) • Givosiran (2019)- acute hepatic porphyria (AHP) • Lumasiran (2020)-primary hyperoxaluria type 1 (PH1) • Inclisiran (2021)- heterozygous familial hypercholesterolemia (HeFH) • Vutrisiran (2022)- hereditary transthyretin amyloidosis (hATTR) • $450,000 per year (for life) • One of the challenges facing siRNAs is their delivery. They must be delivered to and taken up by their intended target, but they have low bioavailability, requiring a larger dose for absorption, and delivery is compromised by rapid clearance. RNAi therapies with targets outside the liver have yet to make it far in clinical trials. https://www.ajmc.com/view/market-of-sirnas-poised-to-expand-beyond-3-currently-approveddrugs Genome editing • Genome editing, or genome editing with engineered nucleases is a type of genetic engineering in which DNA is inserted, deleted or replaced in the genome of a living organism using engineered nucleases, or "molecular scissors." Strategies for treating genetic disease The evolution of genome editing 1996 2010 2013 zinc finger nucleases https://www.kjim.org/journal/view.php?viewtype=pubreader&number=169769#!po=16.6667 2020 Nobel Prize in Chemistry “for Developing CRISPR Technology" Jennifer A. Doudna Emmanuelle Charpentier CRISPR https://www.youtube.com/watch?v=2pp17E4E-O8 CRISPR/Cas9 genome editing technology A CRISPR/Cas9-caused break in DNA can be repaired in four different ways, two of which open the door to inserting a new gene of choice (“knock-in”) and two of which can disable a gene (“knock-out”). CRISPR/Cas9 genome editing DuPont/Caribou Biosciences drought resistant corn and wheat Roslin Institute – swine feverresistant pigs Recombinetics– hornless dairy cattle CRISPR gene-editing in Humans On October 2016, a team led by oncologist, Lu You, at Sichuan University in Chengdu, China, delivered PD-1 modified cells into a patient with metastatic non-small-cell lung cancer: ex-vivo First human CRISPR Trial Victoria Gray was treated in 2019, making her the first person in the United States to undergo treatment for Sickle Cell Disease using a CRISPR-based therapy turning off the BCL11A gene: ex-vivo https://innovativegenomics.org/multimedia-library/meet-victoria-gray/ September 2021- FDA had accepted its Investigational New Drug application for EBT-101 as a potential functional cure for chronic HIV. EBT-101 will be a first-in-human, CRISPR-based one-time gene therapy to be evaluated in individuals with HIV Yin et al. 2017 excised the HIV-1 DNA from the genome of a "humanized" mouse that carries HIV-infected human lymphocytes Clinical Trials He Jiankui CRISPR baby scandal He Jiankui, in 2018, utilized CRISPR to edit twin embryos born in October 2018. • The parents of the children, “Mark” (HIV-positive) & “Grace” (HIV-negative) were offered IVF + CRISPR gene editing so children will have innate resistance to HIV. • CCR5 gene • December 30, 2019, the Shenzhen Nanshan District People's Court sentenced He Jiankui to three years in prison and a fine of 3 million RMB ($430,000 USD). Challenges of genome editing • It is not 100% accurate • Could have significant off-target side effects • Still very new • Ethical concerns • Eugenics potential • Creating dangerous new biologicals • Expensive so only available to few • Not regulated uniformly across the globe References • Genomics & Personalized Medicine: What Everyone Needs to Know, Snyder, Oxford Press • Genetics in Medicine, 8th ed. Nussbaum, McInnes, Willard, Elsevier • https://ghr.nlm.nih.gov/primer/therapy/genetherapy • Genome editing tools info: https://www.ptglab.com/news/blog/crisprcas9-talens-and-zfns-the-battle-in-gene-editing/ • https://clinicaltrials.gov/