Gene Therapy for Spinal Muscular Atrophy Tuesday, November 14, 2017 Cheng Shan, Brandon Mikhael, Olivia Gallagher, and Kathy Nguyen PHM142 Fall 2016 Coordinator: Dr. Jeffrey Henderson Instructor: Dr. David Hampson

What is Spinal Muscular Atrophy (SMA)? 3 Spinal muscular atrophy is a rare neuromuscular disease in the autosomal 5q gene1 Main effect is loss of motor neurons, causing muscular atrophy and eventually respiratory failure1 Children and infants are the most affected1 Affects 1 in 6000 - 1-10,000 live births2 MacKenzie, Alex. "Genetic therapy for spinal muscular atrophy." Nature biotechnology 28.3 (2010): 235-237. D'Amico, A., Mercuri, E., Tiziano, F. D., & Bertini, E. (2011). Spinal muscular atrophy. Orphanet journal of rare diseases, 6(1), 71. Picture from https://www.premilife.com/diseases/brain-and-nerves-system/spinal-muscular-atrophy/ not really sure how to cite a picture from a website tho...

SMN-1 (Survival Motor Neuron -1) gene is usually homozygously deleted1 Root Causes SMN-1 (Survival Motor Neuron -1) gene is usually homozygously deleted1 SMN protein is involved with much of RNA metabolism1 The absence of the SMN gene is lethal in all other animal species embryonically1 Humans have the presence of a SMN-2 gene, which produces a small amount of SMN protein permitting initial survival1 MacKenzie, Alex. "Genetic therapy for spinal muscular atrophy." Nature biotechnology 28.3 (2010): 235-237.

Exact role of SMN is unclear1 SMN Function 4 Exact role of SMN is unclear1 SMN protein deficiency may cause a deficient pre-synaptic transcriptome, causing denervation1 Greater SMN-2 copy number is associated with a better outlook1 1.MacKenzie, Alex. "Genetic therapy for spinal muscular atrophy." Nature biotechnology 28.3 (2010): 235-237.

Targets defective or missing genes What is Gene Therapy? Targets defective or missing genes Introduction of nucleic acids (genetic material) into the cell Viruses often used to deliver genetic material5 Main goal is to restore function with no adverse effects Gene Therapy - A Revolution in Progress: Human Genetics and Medical Research. (n.d.). Retrieved November 14, 2017, from https://history.nih.gov/exhibits/genetics/sect4.htm

Gene Augmentations for SMA 6 Different approaches to gene therapy Use of adeno-associated virus to deliver full length SMN6 Use of histone deacetylase inhibitors or small molecules to promote production of full length SMN from SMN26 Wood, M. J., Talbot, K., & Bowerman, M. (2017). Spinal muscular atrophy: antisense oligonucleotide therapy opens the door to an integrated therapeutic landscape. Human Molecular Genetics.

Gene Therapy for SMA - Mouse Model 1 Intravenous administration of adeno- associated virus serotype 9 (scAAV9) with SMN complementary DNA that encodes for survival motor neuron (SMN)7 Coding region of recombinant virus forms intramolecular double-stranded DNA template7 Inducing expression of SMN counters effects of SMA and extends survival7

Gene Therapy for SMA - Human Trials Study performed in April 20147 15 infants with SMA17 Given single dose of vector containing scAAV9 along with human survival motor neuron gene (hSMN) and a chicken beta-active promoter (scAAV9.CB.hSMN) (AVXS-101)7 Vector delivered in normal saline through intravenous infusion7 Self-complementary feature of vector with hybrid promoter enables rapid and sustained expression of SMN7

Gene Therapy for SMA - Human Trials RESULTS: All 15 patients alive and event-free at 20 months of age7 In high-dose cohort, scores on the CHOP INTEND (Children’s Hospital of Philadelphia Infant Test of Neuromuscular Disorders) scale of motor function all increased7 12 patients that received high doses: 11 sat unassisted, 9 rolled over, 11 fed orally and could speak, and 2 walked independently7

Gene Therapy = Therapeutic Strategy to treat SMN Increased survival7 Achievement of motor milestones7 Improved motor function7

Adverse Effects of Gene Therapy 7 There are many adverse events related to scAAv9 gene therapy7 56 serious adverse events were observed in 13 patients7

Limitations of Gene Therapy The presence of anti-AAV9 antibodies hinders universal application of scAAV9 gene therapy8 Antibodies may be present prior to therapy or in some cases, be developed as a result of therapy8 Antibodies can be detected at birth8 Anti-AAV9 causes vector clearance by the immune system and therefore could make scAAV9 gene therapy ineffective8

Overcoming the Limitations of Gene Therapy Potential methods to overcome the limitations of anti-AAV9 antibodies: 1. Inducing transient immunosuppression8 2. Designing AAV9 variants that that are resistant to antibodies8 Further research and development of gene therapy are required to make treatment more universal

Summary Slide Spinal muscular atrophy is a rare neuromuscular disease1 Characterized by loss of motor neurons, causing muscular atrophy and eventually respiratory failure1 Usually caused by the homozygous deletion of the SMN-1 (Survival Motor Neuron-1) gene11 Humans have SMN-2 gene which produces a small amount of SMN protein permitting initial survival1 Gene therapy targets defective or missing genes by introducing nucleic acids (genetic material) into the cell using viruses Gene therapy for spinal muscular atrophy involves the intravenous administration of adeno-associated virus serotype 9 (scAAV9) with SMN complementary DNA that encodes for survival motor neuron (SMN)7 This allows for improved motor function and increased survival7 The presence of anti-AAV9 antibodies hinders universal application of scAAV9 gene therapy by causing vector clearance8 Potential methods to overcome this include inducing transient immunosuppression and designing variants that are resistant to antibodies8 Further research and development of gene therapy are required to make treatment more universal

References MacKenzie, A. (2010). Genetic therapy for spinal muscular atrophy. Nature Biotechnology, 28, 235-7. D’Amico, A., Mercuri, E., Tiziano, F. D., & Bertini, E. (2011). Spinal muscular atrophy. Orphanet journal of rare diseases, 6(1), 71. Spinal Muscular Atrophy [Online image]. 2016. Retrieved November 12, 2017 from http://libguides.gwumc.edu/c.php?g=27779&p=170351. Baiono, M. T., & Ambiel, C. R. (2010). Spinal muscular atrophy: diagnosis, treatment and future prospects. Journal de pediatria, 86(4), 261-70. Gene Therapy - A Revolution in Progress: Human Genetics and Medical Research. (n.d.). Retrieved November 14, 2017, from https://history.nih.gov/exhibits/genetics/sect4.htm. Wood, M. J., Talbot, K. & Bowerman, M. (2017). Spinal muscular atrophy: antisense oligonucleotide therapy opens the door to an integrated therapeutic landscape. Human Molecular Genetics, 26(R2), R151-59. Mendell, J. R., Al-Zaidy, S., Shell, R., Arnold, W. D., Rodino-Klapac, L. R., Prior, T. W., … Kaspar, B. K. (2017). Single-Dose Gene-Replacement Therapy for Spinal Muscular Atrophy. The New England Journal of Medicine, 377(18), 1713-22. Jeune, V. L., Joergensen, J.A., Hajjar, R.J. & Weber, T. (2013). Pre-existing Anti-Adeno-Associated Virus Antibodies as a Challenge in AAV Gene Therapy. Hum Gene Ther Methods, 24(2), 59-67.