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Myeloid-derived suppressor cells (MDSCs) have been identified as a heterogeneous cell population that expand during tumor progression and infiltrate the tumor, peripheral blood, as well as ascites of ovarian cancer patients. MDSCs contribute to tumor immune tolerance via inhibition of T-cell proliferation/activation. It is also known that high tumor cell glucocorticoid receptor (GR) expression is associated with a relatively poor prognosis in ovarian cancer patients. We hypothesized that ovarian tumor cell GR expression and activity may modulate tumor cell cytokine secretion thereby leading to increased MDSC generation and recruitment, thus creating an immunosuppressive tumor microenvironment. Our preliminary data confirms GR-mediated secretion of pro-tumorigenic cytokines including G-CSF, TGF-b, and MCP-1 in high-grade ovarian cancer models. We also observed GR-mediated downregulation of IL-2 and IFN-g, cytokines required for anti-tumor T-cell function. Importantly, treatment of cells with a competitive GR antagonist reversed this immunosuppressive secretome effect. Moreover, culturing healthy peripheral blood mononuclear cells (PBMCs) with tumor conditioned media from GR activated ovarian cancer cells demonstrated that secreted immunomodulatory factors have the capacity to promote differentiation of human MDSCs from PBMCs. Based on the insight provided by these data, we propose to determine whether ovarian tumor cell-intrinsic GR activation and resulting MDSC generation has an inhibitory effect on cytotoxic T-cell function, contributing to earlier relapse of GR-positive ovarian cancers, and eventually whether this mechanism may be targeted to improve patient outcomes in this subset of ovarian cancers. Supported by grant from CPRIT

Dendritic cells (DCs) loaded with killed allogeneic melanoma cells can cross-prime naive CD8+ T cells to differentiate into melanoma-specific CTLs in 3-wk cultures. In this study we show that DCs loaded with killed melanoma cells that were heated to 42°C before killing are more efficient in cross-priming of naive CD8+ T cells than DCs loaded with unheated killed melanoma cells. The enhanced cross-priming was demonstrated by several parameters: 1) induction of naive CD8+ T cell differentiation in 2-wk cultures, 2) enhanced killing of melanoma peptide-pulsed T2 cells, 3) enhanced killing of HLA-A*0201+ melanoma cells in a standard 4-h chromium release assay, and 4) enhanced capacity to prevent tumor growth in vitro in a tumor regression assay. Two mechanisms might explain the hyperthermia-induced enhanced cross-priming. First, heat-treated melanoma cells expressed increased levels of 70-kDa heat shock protein (HSP70), and enhanced cross-priming could be reproduced by overexpression of HSP70 in melanoma cells transduced with HSP70 encoding lentiviral vector. Second, hyperthermia resulted in the increased transcription of several tumor Ag-associated Ags, including MAGE-B3, -B4, -A8, and -A10. Thus, heat treatment of tumor cells permits enhanced cross-priming, possibly via up-regulation of both HSPs and tumor Ag expression.