Immunohistochemistry for granzyme B (Cell Signaling Technology), IDO-1 (Sigma Aldrich), UBE2C (Abcam), beta catenin (Ventana), and IDH1 R132H (Dianova) expression was performed using the conditions outlined in Table S5. granzyme B was selected for validation due to common use of this marker in diagnostic laboratories, after confirmation that GZMA and GZMB transcript expression were closely correlated in previously published MCC expression profiles [17 (link)]. Potential prognostic markers (granzyme, IDO1, UBE2C) were evaluated using previously described tissue microarrays and whole sections on primary tumors,[26 (link)] alongside additional primary tumor cases, to achieve adequate statistical power. Immunostained slides were scanned at 20x magnification on a Vectra Polaris (PerkinElmer). Scoring was performed on representative digitized tumor fields selected by a board-certified dermatopathologist (P.W.H.) using the Positive Cell Detection analysis in QuPATH software to quantitate either density of positive inflammatory cells (granzyme, IDO1) or H-score expression on tumor cells (UBE2C, beta-catenin). Beta-catenin and IDH1 R132H immunohistochemistry were performed on representative mutant and wild type tumors.