Combined genetic and chemical capsid modifications enable flexible and efficient de- and retargeting of adenovirus vectors

Numerous attempts to target viral gene therapy vectors to specific cells have met with limited success. Here we describe a novel virus vector-targeting platform based on a unique combination of genetic and chemical vector particle modifications to overcome typical restrictions in virus vector targeting. We genetically introduced cysteines at solvent-exposed positions of the adenovirus capsid. The corresponding thiol groups were highly reactive, and we established procedures for controlled covalent coupling to them of protein and nonprotein ligands. After the coupling of transferrin, the particles were efficiently targeted to the transferrin receptor pathway. Depending on the chemistry used, ligands could be coupled under the formation of thioether or disulfide bonds, the latter allowing for separation of ligand and particle after cell entry in the endosome. Furthermore, this technology could be efficiently combined with vector shielding for true retargeting: after amino-PEGylation of the vector particles the genetically introduced thiols were still accessible for ligand coupling, and particles could be retargeted to the transferrin receptor. Since this platform is robust, can be scaled, is compatible with industrial standards, and can integrate chemically diverse molecules as ligands, it may be used for clinical gene therapy and, potentially, also for vaccination.