DNA immunotherapy targeting BARF1 demonstrates therapeutic impact in novel murine carcinoma models expressing EBV latent antigens

Abstract Epstein Barr Virus (EBV) latent infection is associated with multiple types of cancer. Several clinical studies have targeted EBV antigens as immune therapeutic targets with limited efficacy of EBV+ malignancies, suggesting additional targets might be important. BamHI-A rightward frame 1 (BARF1) is an EBV antigen that is highly expressed in EBV-associated gastric carcinoma (EBVaGC) and EBV+ nasopharyngeal carcinoma (NPC) and has not been evaluated as an antigen in the context of therapeutic immunization. Here, we developed a synthetic DNA-based expression cassette as immunotherapy targeting BARF1 (pBARF1). Immunization with pBARF1 induced potent antigen-specific humoral and polyfunctional effector T cell responses in vivo. We observed significant antigen-specific increases of IFN-γ+, IFN-γ+ TNF-α+, and IFN-γ+ TNF-α+ IL-2+ populations in CD8+ T cells from the pBARF1 immunized animals in both C57BL/6 and BALB/c mice. Novel BARF1 expressing carcinoma lines that grow in immune-competent animals were developed as surrogates for human EBV+ tumors. We confirmed that BARF1 expression in these new cancer cell lines, MC38-BARF1 and CT26-BARF1, was within the range of human EBV+ cancer cell lines, SNU719 and C666-1. Immunization with pBARF1 plasmid demonstrated impact of tumor progression through induction of CD8+ T cell in these BARF1+ carcinoma models. We also observed long-term antitumor immunity, at least up to 446 days, against BARF1 in a CT26-BARF1 rechallenge study. Using IVIS in vivo imaging, we observed that pBARF1 immunized animals rapidly controlled and cleared cancer cells. These studies demonstrate the potential for pBARF1 immunity to induce antigen-specific immune responses impacting tumor progression. Further study of this immune target and these EBV+ tumor models is likely important as part of therapeutic approaches for EBV+ malignancies. Citation Format: Xizhou Zhu, Alfredo Perales-Puchalt, Krzysztof Wojtak, Ziyang Xu, Kun Yun, Pratik S. Bhojnagarwala, Devivasha Bordoloi, Daniel H. Park, Kevin Liaw, Mamadou A. Bah, Paul M. Lieberman, Ebony N. Gary, Ami Patel, David B. Weiner. DNA immunotherapy targeting BARF1 demonstrates therapeutic impact in novel murine carcinoma models expressing EBV latent antigens [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3573.