AAV9-mediated Intra-striatal Delivery ofGNAO1Reduces Hyperlocomotion inGnao1Heterozygous R209H Mutant Mice

Abstract ID 91058 Poster Board 567 Mutations in the GNAO1 gene, which encodes the abundant brain G-protein Gαo, result in neurologic disorders characterized by developmental delay, epilepsy, and movement abnormalities. There are over 50 mutant alleles associated with GNAO1 disorders; the R209H mutation results in dystonia, choreoathetosis, and developmental delay without seizures. There has been success in controlling these motor symptoms with deep brain stimulation and some medications, but there is a clear need for genetic therapies to provide more definitive treatments. Mice heterozygous for the human mutant allele (Gnao1+/R209H) exhibit hyperactivity in open field tests but no seizures. We developed self-complimentary adeno-associated virus vectors (scAAV9) expressing two splice variants of human GNAO1 Gαo isoforms 1 (GoA, GNAO1.1) and 2 (GoB, GNAO1.2) under a chicken-β-actin promoter. A reporter virus expressing HA-tagged GNAO1.1 showed enriched expression of the human protein in striatum, with some expression in somatosensory cortex. For the therapeutic study, bilateral intra-striatal injections of untagged vectors (scAAV9-GNAO1.1 or scAAV9-GNAO1.2) reduced mutation-associated hyperactivity in open field tests (Total movement: WT 3,500 ± 200 cm, Gnao1+/R209H + Vehicle 11,900 ± 600 cm, Gnao1+/R209H + scAAV9-GNAO1.1 8,100 ± 400 cm, ***p< 0.001 mutant ± AAV treatment; Gnao1+/R209H + scAAV9-GNAO1.2 8,700 ± 600 cm, **p< 0.01 mutant ± AAV treatment; Two-way ANOVA ****p < 0.0001) compared to injection with vehicle (phosphate-buffered saline or scAAV9-GFP). GNAO1 overexpression did not increase seizure susceptibility, a potential side-effect of GNAO1 vector treatment. This represents the first report of successful preclinical gene therapy for GNAO1 encephalopathy applied in vivo. Further studies are needed to uncover the molecular mechanism that results in behavior improvements after scAAV9-mediated G αo expression and to refine the vector design. Supported by a grant from The Bow Foundation.