Therapeutic strategy for spinal muscular atrophy by combining gene supplementation and genome editing

Defect in the SMN1 gene causes spinal muscular atrophy (SMA), which shows loss of motor nerve cells, muscle weakness and atrophy. While current treatment strategies including small molecules or viral vectors improved motor function and survival, an ultimate treatment is highly challenging to correct the endogenous mutations by genome-editing technologies. We have developed the CRISPR-Cas9 based homology-independent targeted integration (HITI) strategy, which allowed for DNA knock-in in both dividing and non-dividing cells in vivo. Here, we demonstrated a gene correction of mutant mSmn1 by using the HITI strategy to treat SMA model mice. To provide the long-term therapeutic benefits of gene correction, we established a unique approach to treat SMA by combination with gene cDNA supplementation and editing (Gene-DUET). As a proof of concept of therapeutic approach for SMA, we demonstrated the advantageous effects including motor function and survival improvement by Gene-DUET strategy in SMA model mice. The Gene-DUET represents a powerful strategy for treating inherited diseases especially for neurodegenerative disorders as well as neuromuscular diseases. To investigate the global transcriptional alterations by HITI, cDNA and DUET treatments in SMA mice compared to untreated SMA mice and heterozygous mice.