AAV PHP.eB-Syn-GFP

Directed evolution is a receptor-independent strategy based on the generation of AAV libraries containing cap gene mutations that confer novel tropisms to AAVs for unknown receptors on target cells. Cre recombination-based AAV targeted evolution (CREATE) is a platform based on the selective recovery of capsid viral genomes from target cell nuclei expressing Cre recombinase in vivo or in vitro during screening campaigns, enabling the isolation of novel AAV variants with altered tropism. With this new technology using AAV9-based capsids, a family of AAV-PHP capsids emerged, among which AAV-PHP.B exhibited significantly improved tropism for various tissues in mice and was able to enter target cell populations that were difficult to reach due to their location (e.g., sympathetic nerves, tuberosity, dorsal root and cardiac ganglia) or their widespread distribution (e.g., enteric nervous system). Further engineering diversification of AAV-PHP.B resulted in AAV-PHP.eB, which has the same peptide insert as AAV-PHP.B but with additional flanking substitutions. This variant efficiently transduced the mouse central nervous system (CNS), showing 55-76% neuronal transduction (depending on the region), a >2.5-fold increase in transduction compared to AAV-PHP.B, with similar percentages of glial cells. High CNS tropism was also observed in rats.