Our studies reported here, and an independent study in B16 murine tumors, have shown that blocking VEGF can result in increased trafficking of tumor-reactive T cells to tumors (23). PI3K inhibitor improved the efficacy of both anti-PD-1 and anti-CTLA4 antibodies in murine models. Together these findings demonstrate that PTEN loss promotes immune resistance and support the rationale to explore combinations of immunotherapies and PI3K-AKT pathway inhibitors. which is mutated in ~50% of melanomas, modulates the immune microenvironment to perturb T Mouse monoclonal to INHA cell-mediated anti-tumor responses. Mutant increases the expression of IL-1 and IL-1 by tumor cells, which increases the expression of PD-L1 and PD-L2 in tumor-associated fibroblasts and suppresses the function of tumor-infiltrating T cells (TILs) (4). BRAF Xanthone (Genicide) inhibition increases the expression of melanocytic antigens (5) and inhibits VEGF production by melanoma cells, thereby enhancing trafficking of tumor-reactive T cells to tumors (6). Clinical trials evaluating the safety and efficacy of BRAF inhibitors in combination with immunotherapies are currently underway. In addition, activation of the -catenin pathway, another oncogenic pathway, was found to be associated with poor tumor infiltration of T cells in a recent publication (7). Together, these results indicate that the impact of tumor-intrinsic pathways is not always confined to tumor cells and can be extended to anti-tumor immune responses, especially T cell responses. The phosphatidylinositol 3-kinase (PI3K) pathway plays a critical role in cancer by regulating several critical cellular processes, including proliferation and survival. One of the most common ways that this pathway is activated in cancer is by loss of expression of the tumor suppressor PTEN, which is a lipid phosphatase that dampens the activity of PI3K signaling. Loss of PTEN corresponds with increased activation of the PI3K-AKT pathway in multiple tumor types (8). Loss of PTEN occurs in up to 30% of melanomas, frequently in tumors with a concurrent activating mutation (9). While expression of mutant alone fails to transform melanocytes, invasive and spontaneously metastatic lesions develop when this is complemented by loss of PTEN in mouse models (10, 11). Loss of PTEN in melanoma patients with mutations is associated with worse outcomes in stage III patients, and in stage IV patients treated with FDA-approved BRAF inhibitors (12, 13). Several studies have demonstrated that melanoma cell lines with loss of PTEN can be growth arrested by BRAF and MEK inhibitors but that they are resistant to apoptosis induction (14, 15). These studies support that PTEN loss identifies a distinct, clinically significant subset of melanomas. In this study, we evaluated the impact of loss of PTEN on T cell-mediated anti-tumor responses. Our studies in preclinical models and clinical specimens demonstrate that loss of PTEN promotes resistance to immunotherapy in melanoma. Our findings provide new insights into the role of PTEN in cancer and identify new strategies to increase the efficacy of immunotherapy in patients. RESULTS Silencing PTEN expression in melanoma reduces T cell-mediated tumor killing and mutations, we silenced PTEN expression in established (Fig. 1B). To evaluate the effects of PTEN loss on T cell-mediated anti-tumor activity, we used an established ACT murine model (6) (Fig.1C). PTEN loss significantly reduced the accumulation of transferred tumor-reactive T cells in A375 melanoma tumors (Fig.1DCE). The adoptively transferred pmel-1 T cells showed significantly reduced therapeutic activity in mice bearing PTEN-silenced tumors when compared to mice bearing PTEN-expressing tumors (Fig.1F, G). Similarly impaired T cell-mediated anti-tumor activity against PTEN-silenced Xanthone (Genicide) tumors was also observed in the context of concurrent treatment with a selective BRAF inhibitor (Supplementary Fig. S1BCF). Collectively, our and studies indicate that PTEN loss can cause resistance to T cell-mediated anti-tumor immune responses. Open in a separate window Figure 1 Reduced T cell-mediated anti-tumor activity against PTEN-silenced melanoma cells(A) PTEN expression and AKT activation in A375/GH cells with and without PTEN silencing. Two PTEN-silenced tumor cell lines (17 and 60) were independently produced by two shRNAs Xanthone (Genicide) targeting PTEN (shPTEN). Tumor cells expression scrambled shRNA (shNS) were served as control (PTEN-WT Tu). (B) T cell-induced apoptosis rate of melanoma tumor cells with and without PTEN silencing. A375/GH/shPTEN and A375/GH/shNS tumor cells were co-cultured with tumor-reactive pmel-1 T cells, at different ratios of effector and target cells (E:T). The cleavage of caspase-3 in tumor cells was determined by flow cytometry. (C) Experimental setup of the murine ACT protocol to evaluate T cell-mediated anti-tumor activity. (D) T cell infiltration of melanoma tumors with and without PTEN silencing < 0.05..