Pancreatic cancer is poised to become the second leading cause of cancer-related deaths globally. This alarming trend is primarily due to the lack of effective treatments once surgery is no longer an option. Therefore, it is crucial to develop new anticancer strategies to improve patient outcomes. Oncolytic viruses offer a promising approach as they specifically infect, replicate, and spread within tumors, causing oncolysis of tumor cells and stimulating antitumor immunity. However, the effectiveness of these viruses is heavily dependent on their accessibility to the tumor ecosystem, a factor that has been insufficiently studied. Our research addresses this critical gap by focusing on the remodeling of the permeability imposed by the pancreatic tumor microenvironment and its implications for the transport and biodistribution of oncolytic viruses. Utilizing the patented technology PRESTO, we characterized the permeability and elasticity of ex vivo tissue slices from mice. Additionally, we constructed in vitro tissues using type I collagen and/or Matrigel, embedding pancreatic cancer cells to accurately mimic in vivo conditions. Our 3D in vitro model allowed us to dissect the role of poromechanic properties in the delivery of oncolytic viruses, advancing our understanding of how to enhance their therapeutic efficacy.