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Head and neck squamous cell carcinoma (HNSCC) are immunosuppressed solid tumors often diagnosed at advanced stages of the disease. Most HNSCC patients have a history of oral epithelial dysplasia; however, there are limited diagnostic criteria used to determine which lesions will progress to cancer. There is an unmet clinical need to determine what cellular events occur during tumor initiation that enables anti-tumor immune evasion and heightened disease activity. Amplification of SOX2, a transcription factor that regulates stem cell pluripotency and fate, is a high-risk event that correlates to cancer initiation in squamous cell carcinomas. We have previously shown that SOX2 amplification results in dampened STING-type I interferon (IFN-I) signaling, a crucial pathway for innate immune sensing of cancers. Our clinical data from a longitudinal study following SOX2-positive patients with oral epithelial dysplasia and then HNSCC diagnosis confirmed infiltration of suppressive myeloid subsets and exclusion of effector T cells upon malignant transformation. Subcutaneous implantation of Sox2-expressing tumors exhibited expansion of myeloid-derived suppressor cells and loss of infiltrating CD8+ T cells. Furthermore, tumor microenvironment displayed secretion of cytokines involved in myeloid recruitment including Csf-2 and IL1a. Moreover, expression of SOX2 in HNSCC cell lines and human-derived epithelial progenitor cells enhanced IL-1α expression. To recapitulate tumor formation in the oral cavity, we engineered a keratin-5-restricted, tamoxifen-inducible Cre to trigger Sox2 expression in the buccal mucosa of mice. We observed that immune landscape substantially shifted before the HNSCC histology occurred. Upon analysis of immune infiltrates at 2-weeks and 4-weeks post tamoxifen painting, we found the recruitment of a distinct myeloid population following malignant transformation. These early myeloid cells display a dichotomous IFN-I-low and IL-1α-high signature. Upon culturing bone marrow-derived macrophages (BMDMs) with recombinant IL-1α and then waiting four days to challenge with agonists to induce IFN-I signaling, we show that priming with IL-1α and IL-6 but not IL-10 desensitizes myeloid cells to STING stimulation. We then found that blockade of IL-1/IL1r1 signaling in our pre-clinical model delayed the influx of myeloid cells and the onset of Sox2-driven cancer initiation. From this, we propose that Sox2-mediated epithelial transformation results in expression of IL-1α which primes myeloid cells to generate an immune-tolerant niche. We employed pharmacological and genetic approaches to show that blockade of IL1 signaling during the cancer initiation stage delays the onset of Sox2-driven HNSCC. Overall, we identified a novel cancer-immune cell interaction axis where Sox2 in cancer cells drives IL1α-mediated STING suppression. Citation Format: Hulya F. Taner, Wong Gong, Luke Broses, Wanqing Chen, Jung Kuczura, Sashider Rajesh, Yee Sun Tan, Yuying Xie, Yu Leo Lei. Resolving an immune tolerogenic niche at the earliest phase of oral cancer initiation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2874.

Background: Progression from noninvasive to muscle-invasive and then metastatic bladder cancer causes >17,000 deaths each year in the United States. Unfortunately, the molecular drivers of progression remain poorly understood and no strategies to prevent progression exist. We have identified an E3 ubiquitin ligase, TRIM29, which is highly expressed in 70% of muscle invasive bladder cancers (MIBC). Global overexpression of TRIM29 in a genetically engineered mouse model (GEMM) is sufficient to induce urothelial cancer initiation and invasive progression. TRIM29 has also been shown to bind and ubiquitinate STING (Stimulator of Interferon Genes), a regulator of anti-tumor cytokine production. However, the role of TRIM29-mediated suppression of STING and innate immune responses in invasive progression was not previously understood. Hypothesis: Given the importance of Trim29 in invasive progression demonstrated in our GEMM, we hypothesized TRIM29-mediated degradation of STING creates an immune-privileged niche required for muscle invasion. Methods: To determine the requirement for Trim29-mediated immune suppression and bladder cancer invasive progression, we generated a urothelial specific tamoxifen inducible Trim29 knockout (KO) GEMM. To induce bladder tumorigenesis, N-butyl-N-(4-hydroxybutyl) nitrosamine (BBN), a urothelial specific chemical carcinogen with similarities to tobacco, was delivered to mice via drinking water. Results: As expected, BBN promoted development of muscle-invasive urothelial tumors in 50% of mice after 24 weeks. Tamoxifen-induced Trim29 KO completely blocked progression to muscle-invasion, although these mice did demonstrate development of low grade, non-muscle invasive tumors. BBN upregulated Trim29 expression in the urothelium, but only modestly increased Sting and infiltrating macrophage and T cells. In contrast, Trim29 KO mice, which did not develop invasive tumors, harbored substantially elevated Sting levels in the urothelium and substantially increased macrophage and T cell infiltration in the bladder and tumor. In vitro, TRIM29 promoted STING ubiquitination and TRIM29 knockdown increased STING protein levels in bladder cancer cell lines. TRIM29 KO in these cells also resulted in increased expression of innate immune cytokines. Conclusion: These results suggest that TRIM29-mediated downregulation of STING in the bladder results in decreased immune surveillance in the tumor microenvironment, contributing to the development of an immune privileged niche required for MIBC progression. Citation Format: Alan Kelleher, Yin Wang, Luke Broses, Yu Leo Lei, Phillip Palmbos. TRIM29-mediated immune evasion in bladder cancer invasive progression [abstract]. In: Proceedings of the AACR Special Conference: Tumor Immunology and Immunotherapy; 2022 Oct 21-24; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2022;10(12 Suppl):Abstract nr A27.