B5 - Neurologic Diseases (excluding Ophthalmic and Auditory Diseases)
303: Optimized Artificial miRNAs Delivered by rAAV9 Dramatically Improve Survival, Respiratory and Motor Functions of SOD1G93A-ALS Mice
Type: Oral Abstract Session
Presentation Details
Session Title: Neurologic Diseases IV
Autosomal dominant mutations in superoxide dismutase 1 (SOD1) gene cause motor neuron degeneration and are linked to 10-20% of familial and 2% of sporadic cases of Amyotrophic Lateral Sclerosis (ALS), a fatal disease for which an effective treatment is urgently needed. While current treatments such as gene silencing using ASO or siRNA have shown limited clinical benefits with the need for repeated dosing, our study explores the use of Adeno-Associated Virus serotype 9 (AAV9) for the delivery of optimized artificial microRNAs (amiRs) targeting SOD1 in a single-doses treatment for ALS-SOD1. Our choice of a miR-33-based amiR scaffold in the design of SOD1-amiR is based on our previous studies demonstrating its gene silencing efficacy, reduced off-target activity, and fewer truncated AAV9 genomes, compared to conventional shRNAs and other scaffolds. We generated an AAV9 vector (AAV9-hSMN1-dual-amiR) using a promoter derived from the endogenous human survival motor neuron 1 (hSMN1) to drive the expression of two amiR-Sod1 embedded in the mouse and human miR-33 scaffold. The SOD1G93A mice were injected by tail vein at the dose of 1.0xe14 vg/kg at three different time points: day 60-68 (the asymptomatic stage), day 90 (non-motor symptomatic stage when mice start to lose body weight), and day 105-125 (the motor symptomatic stage). PBS-treated transgenic animals were used as controls. In comparison to a median survival of 117 days observed in PBS-injected SOD1G93A mice (n = 17), the median survival of the AAV9-hSMN1-dual-amiR treated mice (n = 17) was extended by 98.5 days when animals were injected at day 60-68.More importantly SOD1G93A mice treated at day 90 (n = 24) and day 105-125 (n = 29), showed a median survival extension of 39 and 22 days, respectively. We further investigated the SOD1G93A mice treated on day 60. SOD1 levels measured on day 105 by qRT-PCR showed a substantial reduction in the brain stem (down by 46 ± 12%; p< 0.0001), the spinal cord (down by 35-45%; P< 0.0001) in SOD1G93A mice treated with AAV9-hSMN1-dual-amiR, compared to PBS-treated controls. Reduction of SOD1 expression in motor neurons was confirmed by immunofluorescent staining. SOD1 levels in peripheral tissues such as liver, heart, quadriceps, and diaphragm from same animals were also reduced. Notably, diaphragm, crucial for respiratory function, showed a by ~90% reduction in SOD1 levels in mice treated with the AAV9-hSMN1-dual-amiR vector. To gauge respiratory function, ventilation was quantified using whole-body plethysmography on day 105, 45 days post-injection. When compared to age-matched litter mates, PBS-treated SOD1G93A mice showed hypercapnia in minute ventilation, peak expiration and inspiratory flows. In contrast, hSMN1-dual-amiR vector-treated SOD1G93A mice performed at the same level as the age-matched litter mates. Remarkably, neither rotarod nor grip tests showed any significant difference between the hSMN1-dual-amiR-treated group and age-matched healthy litter mates until day 190. Thus, a single intravenous injection of AAV9-hSMN1-dual-amiR vector significantly reduces SOD1 expression within CNS and non-CNS tissues, effectively preserves respiratory and motor functions, and extends survival by 98.5 days, 39 days, and 22 days when administered at the asymptomatic, non-motor symptomatic and motor symptomatic stages in SOD1G93A mice, respectively. These therapeutic outcomes, unprecedented in SOD1G93A mice, suggested the need for a clinical evaluation of this promising treatment strategy. G.G. and J.X. are Corresponding Authors
Plain Language Summary
Autosomal dominant mutations in superoxide dismutase 1 (SOD1) gene cause motor neuron degeneration and are linked to 10-20% of familial and 2% of sporadic Amyotrophic Lateral Sclerosis (ALS), a fatal neurodegenerative disease leading to generalized progressive muscle weakness and death from respiratory failure within 2-5 years of symptom onset. Here we use a single intravenous injection of a AAV9 vector significantly reduces SOD1 expression within CNS and non-CNS tissues, effectively preserves respiratory and motor functions, and extends survival by 98.5 days, 39 days, and 22 days when administered at the asymptomatic, non-motor symptomatic and motor symptomatic stages in SOD1G93A mice, respectively. These therapeutic outcomes, unprecedented in SOD1G93A mice, suggested the need for a clinical evaluation of this promising treatment strategy.
Fang Wan1, Jinchen He1, Hong Ma1, Veena Kumanan1, Jisun Lee1, Debora PiresFerreira1, Sha Zhu2, Gabriela Toro2, Lingzhi Zhao2, Alisha Gruntman1, Zuoshang Xu3, Joan Shen2, Guangping Gao1, Jun Xie1
1Horae Gene Therapy Center, University of Massachusetts Chan Medical School, Worcester, MA,2NeuShen Therapeutics US, Inc., Lexington, MA,3Biochemistry & Molecular Pharm, University of Massachusetts Chan Medical School, Worcester, MA"
Find This Session