Stressful conditions in the tumor microenvironment induce autophagy in cancer cells to promote their survival, however, autophagy also causes post-translational modification of proteins which are recognized by the immune system. In particular, modified self-antigens can trigger CD4+ T cell responses that can be exploited to boost antitumor immune defenses. We have previously investigated the ability of CD4 cells to target tumor-specific self-antigens modified by citrullination, which converts arginine residues in proteins to citrulline. These studies showed that vimentin, which is frequently expressed in cells during epithelial-to-mesenchymal transition of metastasizing epithelial tumors, is citrullinated and is a good target for anti-tumor immunity (Ref. 1). Immunization with citrullinated vimentin peptides induced IFNγ- and granzyme B-secreting CD4 T cells in response to autophagic tumor targets. Remarkably, a single immunization with modified peptide, up to 14 days after tumor implant, resulted in long term survival in 60-90% of animals with no associated toxicity. This antitumor response was dependent on CD4 cells and not CD8+ T cells. Due to its ubiquitous expression and abundance in most cells, the glycolytic enzyme α-enolase is a protein that is often citrullinated and degraded during autophagy and may represent a further novel antitumor target. In this study we demonstrate that immunization of C57Bl, HLA-DR4 and HLA-DP4 transgenic mice with citrullinated enolase peptides induces strong Th1/cytotoxic CD4 responses that efficiently target tumor cells. The Th1 cell repertoire to citrullinated enolase is also detectable in healthy donors and cancer patients. Immunization of mice with citrullinated enolase peptides led to tumor therapy in HLA-DP4 mice with established B16-DP4 tumors (70% survival p = 0.0058) and in HLA-DR4 transgenic mice with established B16F1-DR4 melanoma (50% survival; p = 0.0048) or Pan02-DR4 pancreatic tumors (survival 50%; p = 0.0076). The response was partially mediated by CD4 cytotoxic T cells as tumor therapy was observed against the HLA-DR4-expressing lung tumor LLC2 (40% survival; p = 0.0142) but no survival advantage was witnessed against LLC2 tumors which do not express class II MHC. As MHC-II is not expressed by the majority of tumors unless induced by IFNγ we designed an HLA-DR4 construct under expression of an IFNγ inducible promoter. Immunization of HLA-DR4 mice with citrullinated enolase peptides led to tumor therapy against the established B16F1-IFNγ inducible DR4 melanoma (90% survival p>0.0001). These results suggest that, similar to citrullinated vimentin, citrullinated α-enolase is a promising novel target for human cancer immunotherapy. References1. Brentville VA, Metheringham RL, Gunn B, Symonds P, Daniels I, Gijon M, Cook K, Xue W, Durrant LG (2016). Citrullinated vimentin presented on MHC-II in tumor cells is a target for CD4+ T cell-mediated antitumor immunity. Cancer Research 2016 Feb 1;76(3):548-60.
Citation Format: Katherine Cook, Ian Daniels, Victoria Brentville, Rachael Metheringham, Wei Xue, Peter Symonds, Tracy Pitt, Mohammed Gijon, Lindy G. Durrant. Citrullinated α-enolase as a novel target for cancer immunotherapy [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 A035.
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