Subpial Gene Delivery of synapsin‐promoted Caveolin‐1 Prolongs Survival in hSOD G93A mice Model of ALS

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Introduction Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease resulting from loss of upper and lower motor neurons in brain and spinal cord . Familial (FALS) and sporadic (SALS) forms account for 90-95% and 5-10% of ALS cases, respectively. Due to the unknown etiology of SALS, interventions that afford neuroprotection and promote functional neuronal and synaptic plasticity are urgently needed. Recent clinical findings now shows an ALS-linked risk variant in the non-coding enhancer region of caveolin-1 (Cav-1), resulting in reduced Cav-1 protein expression and disruption of membrane/lipid rafts (MLRs). The present study tested whether spinal cord delivery of neuron-targeted Cav-1 (using adeno-associated virus serotype 9 (AAV9) encoding a synapsin-promoted Cav-1 construct (i.e., SynCav1)) via subpial lumber route of administration would improve motor function and extend survival in a mutant humanized mouse model of ALS (hSOD1G93A). Methods AAV9-SynCav1 subpial lumber injection (10 ul; 1.9x1013 genomic copies ml-1) at the lumber 1 level with thoracic 13 laminectomy was performed at 8 weeks (wk) of age. Weekly body weight (BW) was recorded and survival was determined using Kaplan-Meier curve. Running wheel (RW) tests were recorded at both 8 wk (pre-surgery baseline) and 16 wk (symptomatic age). BW and RW values were expressed as mean ± standard error while Kaplan-Meier was expressed as meidan days (d) survival. Statistical significance was defined as p < 0.05. Results Immunoblot assay revealed a significant decrease in Cav-1 protein expression in end-stage hSOD1G93A lumbar spinal cord tissue. Subpial SynCav1 delivery preserved Cav-1 expression without affecting misfolded SOD expression as determined by primary antibody for Cav-1 (#3267, 1:1000, Cell Signaling Technologies, Danvers, MA, USA, Denvor). Both male and female hSOD1G93A mice that received SynCav1 exhibited significantly greater survival compared to non-treated hSOD1G93A mice (male median survival 181 d for SynCav1-hSOD1G93A vs 158 d non-treated hSOD1G93A, p=0.002; female median survival 180 d for SynCav1-hSOD1G93Avs 167 d non-treated hSOD1G93A, p=0.008). No significant difference in BW or RW was measured among the groups. Conclusion SynCav1 gene delivery to the spinal cord via subpial administration significantly prolongs survival in the aggressive hSOD1G93A mouse model of ALS. We did not observe improvements on body weight nor motor function with SynCav1, possibly due to the aggressive nature of this particular transgenic disease model. From week 8 to week12, body weight from surgery group is even lower compare to non-treated hSOD1G93A mice due to surgery. Future direction is to optimize the dose of SynCav1 while simultaneously using a less invasive route of administration (e.g., intrathecal) in order to minimize 1) physical trauma with subpial procedure and 2) any surgery-associated inflammatory response, both of which could contribute to rapid disease progression.