Enhancing mesothelin CAR T cell therapy for pancreatic cancer with an oncolytic herpes virus boosting CAR target antigen expression

Abstract Mesothelin (MSLN) is a prominent target antigen for CAR T cell therapy due to its extensive expression in various solid tumors, including pancreatic cancer. However, the therapeutic efficacy of MSLN-targeted CAR T cell therapy has been limited in clinical trials for pancreatic cancer, often resulting in temporary stable disease as the best response. The heterogeneous expression of MSLN and its loss over time, along with the immunosuppressive tumor microenvironment (TME), are key factors restricting effectiveness. Oncolytic viruses are emerging cancer therapies that replicate in tumor cells and remodel the TME into an immunogenic state. Here, we engineered an oncolytic herpes simplex virus type 1 expressing human MSLN (HSV-MSLN) and evaluated its combination with MSLN-CAR T cells in a murine pancreatic ductal adenocarcinoma model. In vitro, HSV-MSLN effectively induced MSLN expression on murine pancreatic cancer cells, with subsequent cell lysis. In co-culture, HSV-MSLN-infected cancer cells activated MSLN-CAR T cells, which effectively eliminated the infected cells. In vivo, HSV-MSLN delivered MSLN on the tumor cell surface and reprogrammed the TME toward an immunogenic state. The combination therapy significantly enhanced antitumor efficacy, inducing activated, proliferative CD8 + CAR T cells and reducing PD-1 + TIM-3 + exhausted endogenous CD8 + T cells and regulatory T cells in tumors. Furthermore, the combination therapy increased migratory XCR1 + CD103 + dendritic cells (DCs) in tumors and tumor-draining lymph nodes (TDLNs) while expanding CD44 + CD8 + T cells with central and effector memory phenotypes. Taken together, these results demonstrate that HSV-MSLN reprograms immune cells in the TME and TDLNs and synergizes with MSLN-CAR T cells to enhance antitumor responses, leading to a more robust therapeutic effect. Graphical abstract MSLN-CAR T cell therapy showed limited efficacy against pancreatic cancer due to heterogeneous antigen expression and an immunosuppressive TME. Here, we engineer the oncolytic virus HSV-MSLN to deliver MSLN and reshape the TME, combining it with MSLN-CAR T cells. Combination therapy reduces tumor burden, enhances CAR T cell activation and proliferation, and decreases exhaustion. Additionally, it increases CD4 + and CD8 + T cell infiltration, enhances CD8 + T cell activation, reduces exhaustion, and decreases Tregs in the tumor. It also elevates migratory DCs in tumors and TDLNs and expands central and effector memory CD8⁺ T cells, promoting a dynamic immune cycle that amplifies antitumor immunity.