Replication-competent influenza virus with a protein-responsive multiplication ability

Applications of influenza A viruses (IAV) for virotherapy and biotechnology have accelerated substantially with the development of reverse genetic technology and advances in the understanding of packaging signals. While the use of a replication-competent IAV is particularly promising, owing to its efficient transmission to organ depths with high infectivity, there is also a risk that its multiplication cannot be controlled in a cell-type-specific manner, causing an infectious disease. Therefore, here a simple and effective replication-competent IAV-based cell-targeting system has been developed. It was demonstrated that the activity of the ribonucleoprotein complex (RNP) of IAV could be regulated by the interaction between the endogenous protein and a nanobody fused to the subunit of RNA-dependent RNA polymerase (RdRp). To validate the feasibility of the method, it was demonstrated that RNP containing RdRp fused with Nb139, a nanobody against p53, is inactive in HEK293T cells expressing endogenous p53, but active in p53-defective Saos-2 cells. Finally, a replication-competent IAV was successfully generated that multiplies only in p53-defective tumor cells and an IAV vector was developed that can deliver a foreign gene in cell type-specific manner. The method is flexible because the nanobody can be easily altered to target a different cell type, offering a valuable platform for virotherapy and biotechnology.