Abstract 3280: Type I IFN receptor signaling in myeloid and T cells drives antitumor immunity elicited by intratumoral delivery of immunogenic recombinant modified vaccinia virus Ankara

Type I interferons (IFNs) are critical in mediating antitumoral and antiviral responses. However, the roles of IFN and IFNAR signaling within the tumor microenvironment remain poorly understood. To address this, we utilized a Cre-loxP system to generate IFNAR1 conditional knockout mice, enabling us to dissect the role of IFNAR signaling in innate and adaptive immune cells during antitumor responses elicited by immune-stimulating vaccinia viruses. Our first-generation virus, MQ710, was developed by deleting E5R, which encodes a cGAS inhibitor, from the non-replicative modified vaccinia virus Ankara (MVA) and incorporating transgenes for membrane-bound Flt3L and OX40L. MQ710 has shown promising early results in a Phase I clinical trial for solid tumors. Building on this platform, the second-generation virus, MQ833, was engineered by deleting two additional viral immunosuppressive genes (E3L and WR199) and introducing extracellular matrix-anchored murine interleukin-12 (IL-12) as the third transgene. MQ833 was found to be more effective than its oncolytic vaccinia counterpart, OV-IL12, expressing the same transgenes in various tumor models, including murine ovarian cancer. The third-generation virus, MQ954, incorporated further modifications to MQ833, including deletion of C12L and insertion of human granulocyte colony-stimulating factor, G-CSF, also anchored to the extracellular matrix. Intratumoral (IT) administration of MQ954 elicited more potent antitumor effects than MQ833 in preclinical models of malignant peripheral nerve sheath tumor (MPNST) and bladder cancer. Using Irf3-/-Irf7-/- and Stat2-/- mice, we found that type I IFN production and signaling are critical for the antitumor effects induced by these viruses. We evaluated the efficacy of MQ833 and MQ954 in LyzcreIfnar1fl/fl, FcgrcreIfnar1fl/fl, MRP8creIfnar1fl/fl, Foxp3creIfnar1fl/fl, and CD4creIfnar1fl/fl mice, in which Ifnar1 gene is depleted exclusively in myeloid cells, monocytes/macrophages, neutrophils, and regulatory T cells, respectively, and compared with Ifnar1fl/fl controls. Our findings revealed that IFNAR signaling in myeloid cells, regulatory T cells (Tregs), and CD4/CD8 T cells is essential for the observed antitumor effects. Single-cell RNA sequencing of CD45+ cells isolated from MB49 tumors from conditional knock-out mice and controls treated with MQ954 twice three days apart further highlighted the pivotal role of type I IFN signaling in monocyte recruitment, macrophage and neutrophil activation and the differentiation of stem-like T cells into effector T cells. This study underscores the multifaceted role of type I IFN signaling across diverse immune cell types in the context of virus-based cancer immunotherapy and provides valuable insights into the intricate immune mechanisms driving antitumor responses. Citation Format: Yueqi Wang, Shanza Baseer Tariq, Ning Yang, Shuaitong Liu, Bharath Salla, Zeda Zhang, Juan Yan, Weiyi Wang, Yi Wang, Gregory Mazo, Juan Angelo-Lozano, Jiahu Wang, Tuo Zhang, Jenny Zhaoying Xiang, Juan C. Osorio, Jeffrey V. Ravetch, Ming O. Li, Ping Chi, Scott W. Lowe, Charles M. Rice, Liang Deng. Type I IFN receptor signaling in myeloid and T cells drives antitumor immunity elicited by intratumoral delivery of immunogenic recombinant modified vaccinia virus Ankara [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 3280.