| The viruses of the order Mononegavirales include important human, animal, and plant pathogens and additionally, have great potential as vaccine, oncolytic and gene therapy vectors. This dissertation focuses on two prototypic Mononegavirales, vesicular stomatitis virus (VSV) and Sendai virus (SeV), their virus-encoded cap methylation function, and potential applications as an anticancer therapy. The L protein of Mononegavirales has six conserved domains postulated to constitute the specific enzymatic activities of this multifunctional protein. We conducted a comprehensive mutational analysis by targeting the entire SeV L protein domain VI, creating twenty-four infectious L mutants. Our analysis identified several residues required for successful cap methylation and virus replication. This study confirms structural and functional similarity of this domain across different families of the order Mononegavirales. Additionally, the oncolytic potential of VSV was analyzed for the first time in a panel of human pancreatic ductal adenocarcinoma (PDA) cell lines and compared to other oncolytic viruses. VSV showed superior oncolytic abilities; however, cells were heterogeneous in their susceptibility to virus-induced oncolysis and several cell lines were resistant to all tested viruses. Four cell lines that varied in their permissiveness to VSV were tested in mice, and in vivo results closely mimicked those in vitro. While our results demonstrate VSV is a promising oncolytic agent against PDA, further studies are needed to better understand the molecular mechanisms of resistance to oncolytic virotherapy. |
