Role of GILT and MHC class II in melanoma cells on regulating the anti-tumor immune response

Abstract

Abstract The MHC class I antigen presentation pathway in melanoma cells has a well-established role in immune-mediated destruction of melanoma. However, the role of the MHC class II pathway in melanoma cells is not fully understood. Gamma-interferon-inducible lysosomal thiol reductase (GILT) is critical for MHC class II-restricted presentation of multiple melanoma antigens by antigen presenting cells. While GILT and MHC class II expression is typically limited to antigen presenting cells, GILT and MHC class II can be expressed constitutively or induced by IFN-γ in melanoma cells. In human melanoma specimens, high GILT expression and an active and intact MHC class II pathway are associated with improved survival. The goal of this project is to investigate a causal role for GILT and MHC class II in melanoma cells, using Yale University Mouse Melanoma (YUMM) lines YUMM2.1, YUMMER1.7, and YUMMERG. These cell lines constitutively expressed GILT, and MHC class II expression was IFN-γ-inducible in a subset of cells under serum-free conditions. We have genetically engineered GILT−/− YUMM2.1 cells and MHC class II (H2Ab1)−/− YUMM2.1 cells via CRISPR/Cas9 and verified the deletion. We have generated lentiviral constructs expressing GILT alone, CIITA alone, and dual expression of H2Ab1 and CIITA, to allow for the generation of melanoma cell lines with and without GILT and MHC class II. Transduction of GILT−/− YUMM2.1 cells with GILT resulted in high transduction efficiency and overexpression. Thus, we have developed models to determine the effect of GILT and MHC class II expression in melanoma cells on regulating in vivo tumor growth and the anti-tumor immune response. This work was supported by a Basic Science Award from the Harry J. Lloyd Charitable Trust and a Merit Review Award I01-BX005336 from the US Department of Veterans Affairs (VA), Biomedical Laboratory Research and Development Service (K.T.H.). The contents do not represent the views of the VA or the US Government.