Cell type specific gene delivery using chemically modified AAV vectors

In the last years the use of Adeno-Associated Viruses (AAVs) as carriers for gene transfer became increasingly popular for both research and clinical applications. Besides their excellent safety profile, the existence of various AAV serotypes characterized by different proprieties is exploited in the context of gene therapy to target a broad range of diseases. However, currently used techniques to re-direct AAV tropism still present many drawbacks. With this project, we have developed a tool that could enable cell-type specific reprogramming of viral tropism based of post-production chemical modifications of the vectors. By using AAV2 as representative serotype, we patented a method to chemically coupling targeting ligands by exploiting naturally occurring functional groups present on viral capsid. In particular, we have employed different heterobifunctional crosslinker molecules to couple the lectins Wheat Germ Agglutinin (WGA) and Griffonia Simplicifolia Isolectin B4 (IB4), the protein Nerve Growth Factor (NGF) and the Protease-Activated Receptor 1 (PAR1) agonist peptide SFLLRN directly with the amine groups (-NH2) present on lysine residues that compose AAV viral proteins (VPs). After in vitro validation and optimization of modified vectors, their efficiency and acquired specificity were assessed in a mouse model, where we were able to confirm correct re-targeting of viral tropism and acquired accurate cell-type specificity. Moreover, we have explored other possible applications of our tool, including boosting transduction efficiency of different AAV serotypes and conferring particular proprieties such as increased permeability for cellular barriers. Due to its versatility, the technology we have developed could be exploited in different fields of basic and applied research and could have a major impact on gene therapy approaches. Furthermore, being it based on simple click chemistry reactions, it could allow for streamlined modification of different class of viruses, thus offering a valid and more flexible alternative to currently used methods to re-direct viral tropism based on genetic modifications.