[3P-79] Capsid protein engineering of Adeno-Associated Virus by peptide insertion to improve target specificity
Adeno-Associated Virus (AAV) is a non-pathogenic virus with low immunogenicity which is attracting attention as a promising vector for gene therapy. The limitation of AAV, however, is its broad tropism in vivo which leads to the issue of off-target gene transduction. An approach to overcome this issue is capsid protein engineering of AAV by insertion of foreign peptide sequences to eliminate endogenous capsid-receptor interaction while granting novel interaction. Recently, we have developed LassoGraft technology, a method to grant novel receptor-binding specificities onto any protein by inserting the internal sequence of macrocyclic peptides selected against various target receptors. We applied this method to AAV by first selecting two protruding loops on AAV capsid which showed that insertion of a model peptide PA-tag had a negligible effect on capsid assembly but eliminated natural infectivity. Then we lasso-grafted macrocyclic peptides selected against two receptor molecules (Plexin B1 and MET) to the selected sites on AAV. Successful retargeting of the engineered AAV towards the target receptors was confirmed, suggesting the general applicability of this approach to change or improve AAV's cellular tropism.