Single-Dose Intrathecal rAAV2-Retro Delivering Anti-hSOD1 miRNA Confers Robust Neuroprotection in Adult ALS Mice

Amyotrophic lateral sclerosis (ALS) kills within a few years of onset, and the drugs licensed for treatment change that course only marginally. Silencing SOD1 has become a credible treatment route, but two gaps hold this approach back: an efficient vector platform for spinal cord transduction with limited peripheral transduction and evidence on whether adding neurotrophic support to gene silencing has any effect. We built an rAAV2-retro platform for intrathecal delivery. One intrathecal dose of an eGFP-expressing vector transduced all spinal cord segments and several brain regions, with a clear preference for spinal motor neurons and low expression in the liver and heart. Side by side in adult mice, we then used this platform to compare miRNA-mediated silencing of human SOD1, overexpression of IGF1, and the two together in SOD1G93A animals. A single injection of rAAV2-retro carrying an anti-hSOD1 miRNA extended median survival by 25 days, improved motor performance, protected neuromuscular junctions, spared α-motor neurons, and damped gliosis and apoptosis. IGF1 on its own also lengthened survival and limited neuronal loss, but it left motor performance unchanged. The benefit of IGF1 over miRNA alone was observed on most endpoints under the conditions tested; a modest, nominally significant improvement in hindlimb clasping was observed, which should be interpreted as exploratory given the number of comparisons performed. Silencing of hSOD1 therefore accounts for most of the benefit, and no statistically detectable additional benefit was observed for IGF1 on top of hSOD1 silencing under the conditions tested. A single intrathecal dose of rAAV2-retro-miRNA is thus an effective gene silencing approach in the SOD1G93A mouse model, with limited peripheral transduction in the liver and heart observed in C57BL/6J mice. However, these data also show the difficulty of achieving additional benefit by adding IGF1 under the conditions tested.