Characterization of class II MHC processing and presentation pathway in melanoma

Although many melanoma cells express class II MHC molecules constitutively or after treatment with IFNgamma, little has been known about the class II MHC processing pathways and peptides displayed. We found that melanoma cells that have similar levels of class II MHC, but deficiencies of invariant chain and DM compared with BLCL represent a common phenotype. While the class II MHC processing pathway is functional in these cells, the associated peptides derived from cytosolic source proteins were over-represented compared with BLCL. The percentage of cytoplasmic source proteins was not reduced by expression of ICP47, an inhibitor of the transporter associated with antigen processing (TAP), or of invariant chain. This indicated that the over-representation of cytosolic peptides was not due to their loading onto invariant chain free class II MHC molecules in the endoplasmic reticulum. The ability of melanoma cells with this phenotype to present a distinct set of class II MHC associated peptides compared to professional APC may influence their ability to modulate anti-tumor responses.; Defining the mechanisms for presentation of the DRB1*0401- restricted epitope gp10044-59 in melanoma is essential to understanding the peptide-based vaccine efficacy. We showed that in melanoma cells that have down-regulated components of the melanosomal pathway, the majority of gp100 is localized to LAMP-1hi/class II MHC+ conventional endosomes. Antigenically indistinguishable epitopes are generated in both early and late endosomes, despite requirement for different acidic proteases. Gp100 that traffics to the plasma membrane and is then internalized by the AP-2 adaptor protein is the only source for epitope presentation. In pigmented melanomas that contain melanosomes in addition to conventional endosomes, the majority of gp100 molecules accumulate in stage II melanosomes that are largely devoid of class II MHC, and is underrepresented in LAMP-1hi late endosomes. The presentation of the gp10044-59 epitope in pigmented melanomas was substantially lower than in non-pigmented melanomas. Gp10044-59 processing occurred entirely in early endosomes/stage I melanosomes, and not in late endosomes. These findings emphasize that gp100 44-59 presentation is a consequence of gp100 unique cell biology, whose alteration during malignant transformation may modulate melanoma immune recognition.