Abstract 4536: Dissecting tumor-immune microenvironment in response and resistance to immune checkpoint blockade in metastatic melanoma

Abstract Background: Immune checkpoint inhibitor (ICI) therapies have markedly improved the prognosis for patients with stage III & IV metastatic melanoma by prolonging progression-free and overall survival rates. However, the variability in immune evasion and resistance mechanisms presents significant challenges to the clinical efficacy of ICIs. This project aims to define drivers of immunotherapy response and resistance by employing advanced genomic, single-cell mRNA analyses, and spatial profiling techniques on tissue biopsies from metastatic melanoma patients. Methods: In this study, we developed a framework to analyze response and resistance, both intrinsic and acquired, via immune features in the tumor microenvironment in a standardized, uniformly processed, and deeply clinically annotated cohort of metastatic melanoma patients (n=61) treated with ICB as part of the human tumor atlas network (HTAN) initiative. From the tumor samples, we conducted single-nucleus RNA sequencing, and for a subset of the samples high-resolution spatial imaging (including protein mIHC, CODEX, and transcriptomics MERFISH). Standardized processing and data pipelines allowed for integrating genomic, transcriptomic, and spatial features to elucidate characteristics and mechanisms in tumor microenvironment and their relationships with resistance. Results: Studies of the pretreatment samples demonstrated that CD4 and CD8 T cells, particularly TCF7+ CD8 T cells, are significantly more prevalent in responders to immunotherapy. Conversely, macrophages, especially Angio-TAMs, show higher levels of enrichment in non-responders. Moreover, the presence of B cells and follicular dendritic cells in non-lymph node biopsies supports the presence of tertiary lymphoid structures within the TME. Three levels of immune enrichment were identified through spatial analysis, and samples with more immune enrichment tend to have better responses. We also identified 10 recurrent cellular neighborhoods (RCNs) and found that RCN2,4, and 7 with high lymphocyte infiltration are significantly more enriched in responders than non-responders. In addition, RCN4 is associated with immune infiltration while RCN7 is TLS-like. Conclusions: Our findings indicate that patients with low immune infiltration exhibited enrichment of macrophages associated with the hypoxia and angiogenesis phenotypes, while patients with high immune infiltrates displayed enrichment in lymphocytes, particularly TCF7+ CD8+ T cells, confirming previous findings and indicating a robust T cell-mediated immune response. This project integrates genomic, transcriptomic, and spatial features to elucidate shared tumor and microenvironmental states and their relationships with resistance, and guide more personalized and effective treatment strategies for metastatic melanoma. Citation Format: Xinyu Cui, Giuseppe Tarantino, Yiwen He, Priyanka Solanky, Kathleen Pfaff, Aaron R. Thorner, Tyler J. Aprati, Boyang Zhang, Timothy Blosser, Emily Robitschek, Jiajia Chen, Junko Tsuji, Elliot Boblitt, Allison Frangieh, Hannah M. Faulkner, Marta Holovatska, Aleigha Lawless, Michael Manos, Karla Helvie, Tatyana Sharova, Dennie Frederick, James L. Fahey, Diego Villamarin, Sachi Krishna, Chanell Mangum, Ajit J. Nirmal, Domenic Abbondanza, Cai McCann, Bruce Johnson, Alex K. Shalek, Eliezer Van Allen, Xiaowei Zhuang, Ryan Sullivan, Barbara E. Engelhardt, Samouil L. Farhi, Scott Rodig, F. Stephen Hodi, Genevieve M. Boland, David Liu. Dissecting tumor-immune microenvironment in response and resistance to immune checkpoint blockade in metastatic melanoma [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 4536.