Abstract The adult mammalian brain entails a reservoir of neural stem cells (NSCs) generating glial cells and neurons. However, NSCs become increasingly quiescent with age, which hampers their regenerative capacity. New means are therefore required to genetically modify adult NSCs for re-enabling endogenous brain repair. Recombinant adeno-associated viruses (AAVs) are ideal gene therapy vectors due to an excellent safety profile and high transduction efficiency. We thus conducted a high-throughput screening of 177 intraventricularly injected barcoded AAV variants profiled by RNA sequencing. Quantification of barcoded AAV mRNAs identified two synthetic capsids, AAV9_A2 and AAV1_P5, both of which transduce active and quiescent NSCs. Further optimization of AAV1_P5 by judicious selection of promoter and dose of injected viral genomes enabled labeling of 30-60% of the NSC compartment, which was validated by FACS analyses and single cell RNA sequencing. Importantly, transduced NSC readily produced neurons. The present study identifies AAV variants with a high regional tropism towards the v-SVZ with high efficiency in targeting adult NSCs, thereby paving the way for preclinical testing of regenerative gene therapy.

In vivo high-throughput screening of novel adeno-associated viral capsids targeting adult neural stem cells in the subventricular zone