Somatic Therapy of a Mouse SMA Model with a U7 snRNA Gene Correcting SMN2 Splicing

Abstract

Spinal Muscular Atrophy is due to the loss of SMN1 gene function. The duplicate gene SMN2 produces some, but not enough, SMN protein because most transcripts lack exon 7. Thus, promoting the inclusion of this exon is a therapeutic option. We show that a somatic gene therapy using the gene for a modified U7 RNA which stimulates this splicing has a profound and persistent therapeutic effect on the phenotype of a severe Spinal Muscular Atrophy mouse model. To this end, the U7 gene and vector and the production of pure, highly concentrated self-complementary (sc) adenovirus-associated virus 9 vector particles were optimized. Introduction of the functional vector into motoneurons of newborn Spinal Muscular Atrophy mice by intracerebroventricular injection led to a highly significant, dose-dependent increase in life span and improvement of muscle functions. Besides the central nervous system, the therapeutic U7 RNA was expressed in the heart and liver which may additionally have contributed to the observed therapeutic efficacy. This approach provides an additional therapeutic option for Spinal Muscular Atrophy and could also be adapted to treat other diseases of the central nervous system with regulatory small RNA genes.

A New AAV10-U7-Mediated Gene Therapy Prolongs Survival and Restores Function in an ALS Mouse Model

“…This finite therapeutic effect observed in treated SOD1 G93A mice is likely due to the accumulation of a residual amount of protein produced by the unskipped mRNA, attributable to the described prion-like properties of mutant hSOD1. 37,38 Complete SOD1 suppression could be achieved by combining several U7-AS cassettes, as recently described by Schümperli and colleagues 39 for SMA or by coupling other methods to suppress SOD1, such as RNA interference or single chain antibodies 40 counteracting protein aggregation.…”

<p>Nucleic Acid Therapy for β-Thalassemia</p>

“… 38 Not only in β-thalassemia, but the modified U7 snRNAs has also been extensively explored in several RNA mis-splicing diseases including DMD and SMA. 46 – 52 …”

The progress of AAV-mediated gene therapy in neuromuscular disorders

“…In another study, a construct with modified U7 snRNA gene carrying single stranded AON complementary to SMN exon 7 and an additional sequence able to recruit the positive splicing factor SRSF1 (U7-ESE-B) has been generated to stimulate exon 7 inclusion and SMN protein production [77,78]. The construct of scAAV9 vectors containing four tandem copies of the U7-ESE-B gene significantly increased the lifespan and muscle functions when injected into the cerebral ventricles of newborn SMA mice, with results comparable to the treatment of SMN1…”

“…cDNA-scAAV9 gene therapy and splicing-switching-AON therapy. However, it should be noted that a slightly higher dose of the scAAV9-4xsU7 vectors (4 × 10 13 vg/kg) compared to the scAAV9-SMN1 cDNA vectors (2 × 10 13 vg/kg) was needed to increase the average survival of the SMA transgenic mice beyond 100 days [78,79] 3.3 AAV-gene therapy in other Muscular Dystrophies…”