An AAV variant selected through NHP screens robustly transduces the brain and drives secreted protein expression in NHPs and mice

Abstract

Recent work has shown that prolonged expression of recombinant proteins after adeno-associated virus (AAV)–mediated delivery of gene therapy to long-lived, ventricle-lining ependymal cells can profoundly affect disease phenotypes in animal models of neurodegenerative diseases. Here, we performed in vivo screens of millions of peptide-modified capsid variants of AAV1, AAV2, and AAV9 parental serotypes in adult nonhuman primates (NHPs) to identify capsids with potent transduction of key brain tissues, including ependyma, after intracerebroventricular injection. Through these screens, we identified an AAV capsid, AAV-Ep⁺, with markedly increased potency in transducing ependymal cells and cerebral neurons in NHPs. AAV-Ep⁺’s potency was conserved in three species of NHP, two mouse strains, and human neurons derived from induced pluripotent stem cells. To apply AAV-Ep⁺ to the treatment of ceroid lipofuscinosis type 2 disease, a lysosomal storage disorder caused by loss-of-function mutations in tripeptidyl-peptidase 1 (TPP1), we used the capsid to package the human TPP1 transgene (AAV-Ep⁺.hTPP1) and delivered the construct by intracerebroventricular injection into mice lacking TPP1 activity. AAV-Ep⁺ provided robust and therapeutically relevant TPP1 protein concentrations in these mice, significantly improving tremor and life span. In NHPs, high cerebrospinal fluid (CSF) TPP1 concentrations were achieved after intracerebroventricular delivery of AAV-Ep⁺.hTPP1 at a total dose of 1 × 10¹² viral genomes, which was more than 30× lower than previously reported doses in NHPs. These results suggest that AAV-Ep⁺ may be a potent vector for gene therapy applications where CSF protein expression is required.