Cancer immunosurveillance relies on effector/memory tumor-infiltrating CD8þ T cells with a T-helper cell 1(T H 1) profile. Evidence for a natural killer (NK) cell-based control of human malignancies is still largely missing. The KIT tyrosine kinase inhibitor imatinib mesylate markedly prolongs the survival of patients with gastrointestinal stromal tumors (GIST) by direct effects on tumor cells as well as by indirect immunostimulatory effects on T and NK cells. Here, we investigated the prognostic value of tumor-infiltrating lymphocytes (TIL) expressing CD3, Foxp3, or NKp46 (NCR1) in a cohort of patients with localized GIST. We found that CD3 þ TIL were highly activated in GIST and were especially enriched in areas of the tumor that conserve class I MHC expression despite imatinib mesylate treatment. High densities of CD3 þ TIL predicted progression-free survival (PFS) in multivariate analyses. Moreover, GIST were infiltrated by a homogeneous subset of cytokinesecreting CD56 bright (NCAM1) NK cells that accumulated in tumor foci after imatinib mesylate treatment. The density of the NK infiltrate independently predicted PFS and added prognostic information to the Miettinen score, as well as to the KIT mutational status. NK and T lymphocytes preferentially distributed to distinct areas of tumor sections and probably contributed independently to GIST immunosurveillance. These findings encourage the prospective validation of immune biomarkers for optimal risk stratification of patients with GIST. Cancer Res; 73(12); 3499-510. Ó2013 AACR.

Immunosuppressive cytokines subvert innate and adaptive immune responses during cancer progression. The inflammatory cytokine interleukin-18 (IL-18) is known to accumulate in cancer patients, but its pathophysiological role remains unclear. In this study, we show that low levels of circulating IL-18, either exogenous or tumor derived, act to suppress the NK cell arm of tumor immunosurveillance. IL-18 produced by tumor cells promotes the development of NK-controlled metastases in a PD-1-dependent manner. Accordingly, PD-1 is expressed by activated mature NK cells in lymphoid organs of tumor bearers and is upregulated by IL-18. RNAi-mediated knockdown of IL-18 in tumors, or its systemic depletion by IL-18-binding protein, are sufficient to stimulate NK cell-dependent immunosurveillance in various tumor models. Together, these results define IL-18 as an immunosuppressive cytokine in cancer. Our findings suggest novel clinical implementations of anti-PD-1 antibodies in human malignancies that produce IL-18. Cancer Res; 71(16); 5393-9. Ó2011 AACR.

Gut microbiota composition influences the clinical benefit of immune checkpoints in patients with advanced cancer but mechanisms underlying this relationship remain unclear. Molecular mechanism whereby gut microbiota influences immune responses is mainly assigned to gut microbial metabolites. Short-chain fatty acids (SCFA) are produced in large amounts in the colon through bacterial fermentation of dietary fiber. We evaluate in mice and in patients treated with anti-CTLA-4 blocking mAbs whether SCFA levels is related to clinical outcome. High blood butyrate and propionate levels are associated with resistance to CTLA-4 blockade and higher proportion of Treg cells. In mice, butyrate restrains anti-CTLA-4-induced upregulation of CD80/CD86 on dendritic cells and ICOS on T cells, accumulation of tumorspecific T cells and memory T cells. In patients, high blood butyrate levels moderate ipilimumab-induced accumulation of memory and ICOS + CD4 + T cells and IL-2 impregnation. Altogether, these results suggest that SCFA limits anti-CTLA-4 activity.

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Close proximity between cytotoxic T lymphocytes and tumour cells is required for effective immunotherapy. However, what controls the spatial distribution of T cells in the tumour microenvironment is not well understood. Here we couple digital pathology and transcriptome analysis on a large ovarian tumour cohort and develop a machine learning approach to molecularly classify and characterize tumour-immune phenotypes. Our study identifies two important hallmarks characterizing T cell excluded tumours: 1) loss of antigen presentation on tumour cells and 2) upregulation of TGFβ and activated stroma. Furthermore, we identify TGFβ as an important mediator of T cell exclusion. TGFβ reduces MHC-I expression in ovarian cancer cells in vitro. TGFβ also activates fibroblasts and induces extracellular matrix production as a potential physical barrier to hinder T cell infiltration. Our findings indicate that targeting TGFβ might be a promising strategy to overcome T cell exclusion and improve clinical benefits of cancer immunotherapy.

The cytotoxic T lymphocyte antigen-4 (CTLA-4)-blocking antibody ipilimumab induces immune-mediated longterm control of metastatic melanoma in a fraction of patients. Although ipilimumab undoubtedly exerts its therapeutic effects via immunostimulation, thus far clinically useful, immunologically relevant biomarkers that predict treatment efficiency have been elusive. Here, we show that neutralization of IL-2 or blocking the α and β subunits of the IL-2 receptor (CD25 and CD122, respectively) abolished the antitumor effects and the accompanying improvement of the ratio of intratumoral T effector versus regulatory cells (Tregs), which were otherwise induced by CTLA-4 blockade in preclinical mouse models. CTLA-4 blockade led to the reduction of a suppressive CD4 + T cell subset expressing Lag3, ICOS, IL-10 and Egr2 with a concomitant rise in IL-2-producing effector cells that lost FoxP3 expression and accumulated in regressing tumors. While recombinant IL-2 improved the therapeutic efficacy of CTLA-4 block-

During cancer development, a number of regulatory cell subsets and immunosuppressive cytokines subvert adaptive immune responses. Although it has been shown that tumor-derived interleukin (IL)-18 participates in the PD-1-dependent tumor progression in NK cell-controlled cancers, the mechanistic cues underlying this immunosuppression remain unknown. Here, we show that IL-18 converts a subset of Kit

The tumor microenvironment (TME) is a complex ecosystem composed of tumor cells, infiltrating immune cells, and stromal cells intertwined with non-cellular components. The co-evolvement and dynamic interplay within and between these components shape the tumor's distinct biology and influence its response to cancer therapies. Cancer-associated fibroblasts (CAFs) constitute vastly heterogenous stromal cells and are prominent components of the microenvironment in solid tumors. Functionally, CAFs can contribute to the malignant development and progression by diverse mechanisms, including supporting tumor cell growth by secreting growth factors, extracellular matrix remodeling, promoting angiogenesis, and by mediating tumor-promoting inflammation. [1][2][3] Clinically, activated CAFs have been associated with worse prognosis, resistance to therapies, and disease recurrence in multiple cancers. [4][5][6][7] Cancer immunotherapies have rapidly changed the therapeutic landscape for cancer. Despite impressive efficacy demonstrated in subsets of patients, most of the patients show innate or acquired resistance to these therapies. [8][9][10] In solid tumors, these challenges are especially prominent due to abundance of CAFs and a complex TME, which may have a significant impact on T cells recruitment, infiltration, and cytotoxic function within the tumor. 11 A better understanding of the mechanisms that impede immune activation may thus enhance the potential of cancer immunotherapy. Studies conducted in recent years have highlighted an emerging role of CAFs in immune regulation that shapes the tumor immune microenvironment (TIME) and influences response to cancer immunotherapies. 12,13 Analysis of CAFs at the single-cell level revealed distinct subpopulations and functional heterogeneity of CAFs. [14][15][16][17][18] Extensive crosstalk between CAFs and cellular components of the immune system has been shown to contribute to immune escape