Degradation of cellular proteins by the proteasome is critical for generation of MHC-associated peptides. The constitutive proteasome and the IFNγ-induced immunoproteasome (IP) differ in the use of three catalytic β subunits, which alters production of MHC class I epitopes. The potential for a disparate repertoire of epitopes between inflammatory/non-inflammatory tumors therefore arises. We have previously demonstrated this phenomenon as a mechanism of escape from T lymphocyte killing following changed processing of the cancer-testis antigen NY-ESO-1 (Woods et al 2016). Based on our, and others, prior data demonstrating alterations in the epitopes produced following processing of individual antigens, it follows that whole scale plasticity of the immunopeptidome could arise depending on which proteasome subtype is expressed by a tumor. Thus, the potential arises for profound change in the ability of CD8+ T lymphocytes to recognize tumor cells, depending on IFNɣ mediated inflammation. Given the change in targetability of tumor cells we observed following changed processing of a single antigen, a wholescale alteration of the immunopeptidome may represent a significant mechanism of immune escape by the tumor. Such plasticity may occur temporally or heterogeneously within a single tumor. Here, we present the first study demonstrating the effect of IFNɣ mediated inflammation on the composition of the immunopeptidome in melanoma. Using a sensitive mass spectrometry approach (DIA-SWATH) we characterize changes to the immunopeptidome of melanoma, dependent on the proteasome subtype expressed by the cell. These changes dramatically altered the landscape of the cell. Significantly, we demonstrate that while the immunopeptidome had similar overall immunogenicity in presence or absence of IFNɣ, there were changes in the ability of CD8+ T cells to recognize altered epitopes. In in vivo animal models we manipulated melanoma cells to switch proteasome subtype following treatments which induced IFNγ at the tumor site. In human biopsies we demonstrated a direct correlation between the presence of tumor infiltrating lymphocytes, and expression of immunoproteasome specific subunits. Importantly, this study highlights the effect of changes in inflammation at the tumor microenvironment as a potential mechanism of significant immune escape from T lymphocyte mediated tumor targeting. Awareness of tumor immunopeptidome plasticity may be critical to inform development of therapies involving cancer vaccination, adoptive T-lymphocyte transfer, or combination treatments including these.

Citation Format: Katherine Woods, Ralf B. Schittenhelm, Anthony W. Purcell, Andreas Behren, Jonathan Cebon. Inflammation mediates profound changes to the immunopeptidome of melanoma [abstract]. In: Proceedings of the Second CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; 2016 Sept 25-28; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(11 Suppl):Abstract nr A099.