Although clonal neo-antigen burden is associated with improved response to immune therapy, the functional basis for this remains unclear.,Here we study this question in a novel controlled mouse melanoma model that enables us to explore the effects of intra-tumor heterogeneity (ITH) on tumor aggressiveness and immunity independent of tumor mutational burden.,Induction of UVB-derived mutations yields highly aggressive tumors with decreased anti-tumor activity.,However, single-cell-derived tumors with reduced ITH are swiftly rejected.,Their rejection is accompanied by increased T cell reactivity and a less suppressive microenvironment.,Using phylogenetic analyses and mixing experiments of single-cell clones, we dissect two characteristics of ITH: the number of clones forming the tumor and their clonal diversity.,Our analysis of melanoma patient tumor data recapitulates our results in terms of overall survival and response to immune checkpoint therapy.,These findings highlight the importance of clonal mutations in robust immune surveillance and the need to quantify patient ITH to determine the response to checkpoint blockade.,•Novel mouse system to uncouple tumor mutational load and tumor heterogeneity•Lower tumor heterogeneity leads to decreased tumor growth because of immune rejection•Both clone numbers and their genetic diversity mediate tumor growth and rejection•Tumor heterogeneity is linked to patient survival and checkpoint blockade response,Novel mouse system to uncouple tumor mutational load and tumor heterogeneity,Lower tumor heterogeneity leads to decreased tumor growth because of immune rejection,Both clone numbers and their genetic diversity mediate tumor growth and rejection,Tumor heterogeneity is linked to patient survival and checkpoint blockade response,Syngeneic mouse models of melanoma and human patient data are used to explore the effect of intra-tumor heterogeneity on immune response and demonstrate that heterogeneity predicts immunotherapy outcomes better than mutational burden.
Melanoma represents a significant malignancy in humans and dogs.,Different from genetically engineered models, sporadic canine melanocytic neoplasms share several characteristics with human disease that could make dogs a more relevant preclinical model.,Canine melanomas rarely arise in sun-exposed sites.,Most occur in the oral cavity, with a subset having intra-epithelial malignant melanocytes mimicking the in situ component of human mucosal melanoma.,The spectrum of canine melanocytic neoplasia includes benign lesions with some analogy to nevi, as well as invasive primary melanoma, and widespread metastasis.,Growing evidence of distinct subtypes in humans, differing in somatic and predisposing germ-line genetic alterations, cell of origin, epidemiology, relationship to ultraviolet radiation and progression from benign to malignant tumors, may also exist in dogs.,Canine and human mucosal melanomas appear to harbor BRAF, NRAS, and c-kit mutations uncommonly, compared with human cutaneous melanomas, although both species share AKT and MAPK signaling activation.,We conclude that there is significant overlap in the clinical and histopathological features of canine and human mucosal melanomas.,This represents opportunity to explore canine oral cavity melanoma as a preclinical model.