Loss of p14 diminishes immunogenicity in melanoma via non-canonical Wnt signaling by reducing the peptide surface density.

Immunotherapy has achieved tremendous success in melanoma. However, only around 50% of advanced melanoma patients benefit from immunotherapy. Cyclin-dependent kinase inhibitor 2A (CDKN2A), encoding the two tumor-suppressor proteins p14 ARF and p16 INK4a , belongs to the most frequently inactivated gene loci in melanoma and leads to decreased T cell infiltration. While the role of p16 INK4a has been extensively investigated, knowledge about p14 ARF in melanoma is scarce. In this study, we elucidate the impact of reduced p14 ARF expression on melanoma immunogenicity. Knockdown of p14 ARF in melanoma cell lines diminished their recognition and killing by melanoma differentiation antigen (MDA)-specific T cells. Resistance was caused by a reduction of the peptide surface density of presented MDAs. Immunopeptidomic analyses revealed that antigen presentation via human leukocyte antigen class I (HLA-I) molecules was enhanced upon p14 ARF downregulation in general, but absolute and relative expression of cognate peptides was decreased. However, this phenotype is associated with a favorable outcome for melanoma patients. Limiting Wnt5a signaling reverted this phenotype, suggesting an involvement of non-canonical Wnt signaling. Taken together, our data indicate a new mechanism limiting MDA-specific T cell responses by decreasing both absolute and relative MDA-peptide presentation in melanoma.