Abstract 5563: Novel approaches to measuring tumor-mutational burden provide insights into tumor-immune interaction in primary melanoma

Abstract Background: Tumor mutation burden (TMB) has been widely explored in metastatic melanoma as a proxy of neoantigenic load. In contrast, the role of TMB on the biology of primary melanoma disease progression is less understood. In this study, we explored the associations between the immune context of melanoma subtypes and TMB as measured by several standard and novel criteria. Methods: Whole-genome-sequencing and multiplex immunohistochemistry (mIHC) were used to characterize the genomic and immune landscape of a cohort of 116 primary melanomas, including cutaneous non-acral (n=59), acral (n=39) and mucosal (n=18) subtypes, as well as 5 mucosal metastases. mIHC was used to identify intratumoral CD8+ T-cells, memory T-cells, macrophages, dendritic cells, B-cells and PD-L1 expression. Standard measures of TMB -total mutational burden (tTMB) and non-synonymous (ns) mutational burden (nsTMB)- as well as novel measures that incorporate multiplicity and clonality -ns persistent mutation burden (ns-pTMB), and ns mutation dosage (ns-dosTMB) were calculated, and their association with immune profiles and clinical-pathological factors established. Results: In cutaneous non-acral melanomas, ns-pTMB and ns-dosTMB were positively associated with CD8+ T cell proportions (p=0.02 for both comparisons). Clonal, but not subclonal ns-pTMB from mutations present in single-copy regions of the genome were positively associated with higher tumor-resident CD103+CD8 T-cell (p=0.01) and B-cell proportions (p=0.02). Higher single-copy ns-pTMB was also associated with tumors presenting early/intermediate regression compared to late regression. In mucosal melanomas, clonal ns-dosTMB had the strongest positive association with the presence of macrophages (CD68, p=0.005). Additionally, macrophages (p=0.01), tissue resident memory T-cells (p=0.02), PDL1+ cells (p=0.03) and B-cells (p=0.048) were found to be positively associated with clonal ns-pTMB from mutations present in multiple copies, which was also positively associated with the presence of lymphatic invasion (p=0.03). Mucosal patients who recurred had lower levels of single-copy ns-pTMB (p=0.03). In acral melanomas, all measures of TMB were positively associated with the presence of dendritic cells (CD11c), with no particular role for clonality observed. In both cutaneous non-acral (p=0.046) and acral melanomas (p=0.03), but not mucosal (p=0.4), clonal ns-dosTMB was found positively associated with tumor mitotic rate. Conclusion: Novel TMB measures reveal associations between tumor and immune context that are not captured by standard TMB measures alone in primary melanomas. Different TMB measures are linked to distinct immune populations across melanoma subtypes that harbor unique genomic and immune landscapes. Clonal mutations play a dominant role in the associations of TMB with immune cell populations. Citation Format: Eva R. Shteinman, Nigel Maher, Jordan W. Conway, Grace H. Attrill, Felicity Newell, Nic Waddell, Nicholas Hayward, John V. Pearson, Georgina V. Long, Richard A. Scolyer, James S. Wilmott, Ismael A. Vergara. Novel approaches to measuring tumor-mutational burden provide insights into tumor-immune interaction in primary melanoma [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(6_Suppl):Abstract nr 5563.