DG9-conjugated morpholino rescues phenotype in SMA mice by reaching the CNS via a subcutaneous administration

Antisense oligonucleotide (AO)-mediated therapy is a promising strategy to treat several neurological diseases including spinal muscular atrophy (SMA). However, limited delivery to the central nervous system (CNS) with AOs administered intravenously or subcutaneously is a major challenge. Here we demonstrate a single subcutaneous administration of cell-penetrating peptide DG9 conjugated to an AO called phosphorodiamidate morpholino oligomers (PMOs) reaches the CNS and significantly prolonged the median survival compared to unconjugated PMO and R6G-PMO in a severe SMA mouse model. Treated mice exhibited significantly higher expression of full-length SMN2 expression (FL-SMN2) in both the CNS and systemic tissues compared to non-treated and unmodified AO-treated mice. The treatment ameliorated the atrophic musculature and improved breathing function accompanied by improved muscle strength and innervation at the neuromuscular junction with no signs of apparent toxicity. We also demonstrated DG9-conjugated PMO localizes in nuclei in the spinal cord and brain after subcutaneous injections. Our data identify DG9 peptide conjugation as a powerful way to improve the efficacy of AO-mediated splice modulation. Finally, DG9-PMO is a promising therapeutic option to treat SMA and other neurological diseases, overcoming the necessity for intrathecal injections and treating body-wide tissues without apparent toxicity.