Melanoma microsatellites exhibit a metastatic signature by spatial transcriptomics and overexpress mediators of...

Microsatellites in melanoma are physically separate from the primary tumor, are hypothesized to arise by lymphovascular invasion similar to other metastases, and have not been previously studied at the molecular level. We compared gene expression in microsatellites with the matched primary melanoma in two patients using spatial transcriptomics. We identified recurrently altered genes in microsatellites contributing to vascular invasion (ITGA4, MCAM), survival in the circulation (CXCL8, PDGFRB, CDH1), vascular exit (ITGB2), survival at metastatic sites (C3, TGBFI), matrix remodeling (MMP9, VCAN, FN1, BGN), and angiogenesis (EMILIN2). Gene set enrichment analysis confirmed a metastatic signature in comparison to genes upregulated and downregulated in distant melanoma metastases, and identified pathway enrichment for NFKB, CDH1, and ZEB1 signaling. Microsatellites exhibited increased host immune responses and four recurrently overexpressed mediators of immune evasion were present (PAEP, GDF15, CD74, HLA-DRA) implying selection pressure from the increased immune response. Seven additional non-recurrent mediators of immune evasion were identified in microsatellites, overexpressed up to 58-fold in microsatellites. PAEP was 275-fold and 25-fold overexpressed, respectively, in microsatellites in two cases. We studied PAEP protein expression by immunohistochemistry in a larger group of 12 and found overexpression in microsatellites in five of 12 patients. PAEP was detected by immunohistochemistry in a microsatellite and also in a subclone within its primary, raising the possibility that the immunostain identifies the subclone in the primary responsible for the microsatellite. These preliminary findings suggest that melanoma microsatellites are true metastases at the level of gene expression, accounting for the sharply increased risk of disease progression in patients with microsatellites.