Synthetic consensus DNA vaccines against Merkel cell polyomavirus large and small T antigens induce robust polyfunctional T cell responses

Merkel cell carcinoma (MCC) is an aggressive form of skin cancer with a high mortality rate and limited therapeutic options. Approximately 80% of MCC cases are associated with Merkel cell polyomavirus (MCPyV). The Large T antigen (LTAg) and Small T antigen (STAg) of MCPyV are persistently expressed in tumors and important for viral maintenance and oncogenic transformation of infected cells, making them attractive targets for therapeutic vaccination. Here, we describe the development and preclinical evaluation of DNA vaccines against MCPyV LTAg (LTAgvax) and STAg (STAgvax). Vaccines were designed based on highly conserved regions across MCPyV strains and demonstrated robust expression in vitro . Immunization elicited strong antigen specific polyfunctional immune responses, including both CD4+ and CD8+ T cells producing IFNγ, TNFα, and IL2. We observed CD8+ T cells with cytotoxic potential, evidenced by expression of T-bet and CD107a. Importantly, these immune responses were observed in inbred and outbred mouse models, suggesting broad immunogenicity across diverse genetic backgrounds, enhancing the translational relevance of both vaccines. In a minimal residual disease challenge, LTAgvax controlled growth of B16 tumors transduced to express LTAg in mice, improving survival of tumor bearing mice. In a therapeutic challenge, LTAgvax extensively remodeled the tumor microenvironment, with fewer immunosuppressive immune cells and enhanced CD4+ and CD8+ T cell infiltration. The intratumoral CD8+ T cells were more activated and a greater percentage of them had the effector memory phenotype suggesting greater capacity to fight the tumor. This led to improved survival of mice which was further enhanced when mice were co-treated with LTAgvax and anti-PD1 antibody. Together, these findings demonstrate that MCPyV T antigen targeting DNA vaccines induce robust, multifunctional immune responses and control tumor growth. Further development of these vaccines for immunotherapy of MCPyV+ MCC is warranted.