Single-cell profiling uncovers a hypoxia-vascular-immune axis underlying poor immunotherapy response in acral versus cutaneous melanoma

Acral melanoma (AM), accounting for ~50% of melanomas in Asian populations, exhibits far poorer immunotherapy response (anti-PD-1 ORR < 20%) than cutaneous melanoma (CM, ~43.7%). While the tumor microenvironment (TME) plays a pivotal role in immunotherapy resistance, previous studies mainly focused on immune cells, neglecting stromal components like pericytes—creating a critical knowledge gap in understanding AM’s therapeutic refractoriness. We integrated multiple single-cell transcriptomic datasets from 31 acral melanoma, 13 cutaneous melanoma and 35 normal skin samples. Comprehensive bioinformatics analyses including cell type annotation, differential expression, gene functional enrichment, and cell-cell interaction analysis, were performed to delineate the tumor microenvironment differences between acral and cutaneous melanoma. Key findings were validated using spatial transcriptomics, bulk RNA-seq datasets, and functional assays including qPCR and western blot. AM had a more immunosuppressive TME, enriched in collagen-secreting RGS5+/COL3A1+ pericytes that co-localized with KDR+/PODXL+ endothelial cells, associated with vascular abnormalities and hypoxia. These collagen-secreting RGS5+/COL3A1+ pericytes (enriched in AM) were significantly negatively correlated with the infiltration ratio of CD8+ tumor-infiltrating lymphocytes (TILs); moreover, in immunotherapy cohorts, high expression of the collagen-secreting pericyte marker RGS5 was associated with poorer response to immune checkpoint blockade (ICB) treatment. We also identified a hypoxia-related NECTIN2-TIGIT immunosuppressive axis: hypoxia upregulated NECTIN2 on multiple cells, interacting with TIGIT on T cells to potentially exacerbate dysfunction. This study provides insights into vascular-stromal features associated with immunotherapy resistance in AM, highlighting RGS5+/COL3A1+ pericytes and the NECTIN2-TIGIT axis as targets, and guides development of effective combination immunotherapies for AM.