Abstract 4082: Anchored IL-12 synergizes with an epigenetic modulator to promote immune remodeling and overcome anti-PD1-refractory murine tumors

Abstract Background Most patients with solid malignancies harbor innate or acquired resistance to immune checkpoint blockade (ICB), prompting the need for novel therapeutic strategies. Interleukin-12 (IL-12) is a promising cytokine for cancer therapy due to its ability to bridge innate and adaptive immunity. However, a narrow therapeutic index limits the use of systemic IL-12 therapy. Here, we investigated the tumor-suppressive effects and mode of action of intra-tumorally delivered murine Interleukin-12 anchored to aluminum hydroxide (referred as mANK-101) in combination with the class I HDAC inhibitor entinostat, in various ICB-refractory murine tumor models, including CT26 (colorectal) and MOC-1 (HPV16neg). We hypothesized that combining Entinostat with an anchored form of IL-12 could overcome systemic toxicity while maintaining anti-tumor activity. Methods Entinostat and intra-tumoral mANK-101 were administered to mice bearing well-established αPD-1-refractory CT26 (colorectal) and MOC-1 (HPV16neg) tumors. Antitumor activity, survival, and protective memory upon tumor rechallenge were evaluated. Comprehensive proteomic and immune cell analysis was performed in MOC-1 tumors, tumor-draining lymph node (tdLN), and periphery. Tumor-specific T cell responses were examined. Results We demonstrate that intra-tumoral mANK-101 synergizes with entinostat to suppress multiple αPD1-refractory tumors, resulting in significant tumor eradication (62-88%), survival benefit (P 0. 0001), and protective memory, including CT26 (colon, Kras G12Dmut) and MOC-1 (oral, HPV16neg). Analysis of MOC-1 tumor-bearing mice demonstrated these effects to be associated with peripheric activation of CD8+ and NK lymphocytes, augmented polyfunctional IFNγ+/TNFα+-producing CD8+ T cells, CD8+ T cell effector memory, and tumor-specific T cell responses. Significant decrease in CD4+ Tregs and increased CD8/Treg ratio were also observed. Ongoing functional studies, proteomic and immune cell analysis at the tumor site, tdLN, and periphery, including single cell transcriptomics and epigenetic studies, will allow for a deeper understanding of the synergistic effect of mANK-101 with the epigenetic modulator Entinostat. Conclusions Collectively, these findings form a rationale for the clinical combination of intralesional delivery of ANK-101 with entinostat for patients with ICB-refractory malignancies, including colorectal and HPV16neg head and neck cancers. Citation Format: Asma S. Khelifa, Katherine L. Lothstein, Masaya Miyamoto, Rob Tighe, Sailaja Battula, Howard L. Kaufman, Jeffrey Schlom, Sofia R. Gameiro. Anchored IL-12 synergizes with an epigenetic modulator to promote immune remodeling and overcome anti-PD1-refractory murine tumors abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts) ; 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84 (6Suppl): Abstract nr 4082.