MIF inhibits myeloid derived suppressor cell mediated immunosuppression by promoting an inflammatory M1 phenotypic shift

Cancers of the oral cavity and pharynx contribute 3% of yearly cancer cases in the United States, primarily as head and neck squamous cell carcinoma (HNSCC). HNSCC presents with a five-year survival rate of 66.2%, with primary risk factors being alcohol abuse or human papillomavirus infection. Myeloid derived suppressor cells (MDSC) are a myeloid lineage cell population known to accumulate within HNSCC tumors and promote a pro-tumor microenvironment by suppressing the host immune response against tumor cells. Macrophage migration inhibitory factor (MIF) is a cytokine released by immune cells and mediates inflammatory and chemotactic effects upon interaction with its receptor, CD74, expressed by antigen presenting cells (APC), including MDSC. Given their role in promoting tumor growth, our group sought to elucidate the function of host versus tumor cell derived MIF on MDSC. Given the chemotactic effects of MIF upon APC, we hypothesized that knockdown of Mif gene expression would result in a better outcome and reduced tumor burden as a result of reduced MDSC accumulation within the primary tumor microenvironment. Using murine HNSCC cell lines LY2 and MOC2 in conjunction with Mif+/+, Mif-/- and myeloid specific MIF knockout mice (m-Mif-/-), our group has revealed an anti-immunosuppressive role for host-derived MIF through its interaction with MDSC, with tumor derived MIF being immunosuppressive in nature. After injection of the tumor cell lines into the buccal mucosa of age matched mice, we monitored tumor development until terminal sacrifice. At this point, we extracted tumor cells for flow cytometric analysis and isolated tumor associated Gr-1+ MDSC. Extracted MDSC were co-cultured with naïve T cells fluorescently labeled with CFSE, to enable quantification of suppression of T cell proliferation. Furthermore, we extracted isolated MDSC RNA for analysis by RT-qPCR. Unexpectedly, our initial finding ran contrary to our hypothesis, as we found that Mif-/- mice had a mean tumor burden of 108.3 mg compared to Mif+/+ mice who presented with a mean tumor of only 64 mg (p = 0.00624). Flow cytometry revealed increased CD11b+ F4/80+ macrophages in mice expressing MIF. Interestingly, a smaller proportion of these cells were identified as MDSC in Mif+/+ and m-Mif-/- compared to Mif-/-, determined by expression of MDSC markers Ly6G and Ly6C. Our group found that both Mif+/+ and m-Mif-/- demonstrated reduced suppression of T cell proliferation compared to Mif-/-. Gene expression analysis revealed in Mif-/- MDSC upregulation of immunosuppressive genes Arg1, Ptgs2, S100a8 and S100a9. Seeing that both Mif+/+ and m-Mif-/- demonstrate similar anti-tumor phenotypic and molecular responses to HNSCC, our group has concluded that in response to host derived, but not tumor derived, MIF, MDSC at the tumor site shift towards a more anti-tumor M1 phenotype and additionally that MIF derived from non-myeloid host sources within the tumor microenvironment are sufficient to stimulate this response.