CCL5 production in lung cancer cells leads to an altered immune microenvironment and promotes tumor development.

Current immunotherapies for lung cancer are only effective in a subset of patients. Identifying tumor-derived factors that facilitate immunosuppression offers the opportunity to develop novel strategies to supplement and improve current therapeutics. We sought to determine whether expression of driver oncogenes in lung cancer cells affects cytokine secretion, alters the local immune environment, and influences lung tumor progression. We demonstrate that oncogenic EGFR and KRAS mutations, which are early events in lung tumourigenesis, can drive cytokine and chemokine production by cancer cells. One of the most prominent changes was in CCL5, which was rapidly induced by KRASG12V or EGFRL858R expression, through MAPK activation. Immunocompetent mice implanted with syngeneic KRAS-mutant lung cancer cells deficient in CCL5 have decreased regulatory T cells (Tregs), evidence of T cell exhaustion, and reduced lung tumor burden, indicating tumor-cell CCL5 production contributes to an immune suppressive environment in the lungs. Furthermore, high CCL5 expression correlates with poor prognosis, immunosuppressive regulatory T cells, and alteration to CD8 effector function in lung adenocarcinoma patients. Our data support targeting CCL5 or CCL5 receptors on immune suppressive cells to prevent formation of an immune suppressive tumor microenvironment that promotes lung cancer progression and immunotherapy insensitivity.

Targeting myeloid-derived suppressor cells in combination with primary mammary tumor resection reduces metastatic growth in the lungs.

BACKGROUND: Solid tumors produce proteins that can induce the accumulation of bone marrow-derived cells in various tissues, and these cells can enhance metastatic tumor growth by several mechanisms. 4T1 murine mammary tumors are known to produce granulocyte colony-stimulating factor (G-CSF) and increase the numbers of immunosuppressive CD11b+Gr1+ myeloid-derived suppressor cells (MDSCs) in tissues such as the spleen and lungs of tumor-bearing mice. While surgical resection of primary tumors decreases MDSC levels in the spleen, the longevity and impact of MDSCs and other immune cells in the lungs after tumor resection have been less studied. METHODS: We used mass cytometry time of flight (CyTOF) and flow cytometry to quantify MDSCs in the spleen, peripheral blood, and lungs of mice bearing orthotopic murine mammary tumors. We also tested the effect of primary tumor resection and/or gemcitabine treatment on the levels of MDSCs, other immune suppressor and effector cells, and metastatic tumor cells in the lungs. RESULTS: We have found that, similar to mice with 4T1 tumors, mice bearing metastatic 4T07 tumors also exhibit accumulation of CD11b+Gr1+ MDSCs in the spleen and lungs, while tissues of mice with non-metastatic 67NR tumors do not contain MDSCs. Mice with orthotopically implanted 4T1 tumors have increased granulocytic (G-) MDSCs, monocytic (M-) MDSCs, macrophages, eosinophils, and NK cells in the lungs. Resection of primary 4T1 tumors decreases G-MDSCs, M-MDSCs, and macrophages in the lungs within 48 h, but significant numbers of functional immunosuppressive G-MDSCs persist in the lungs for 2 weeks after tumor resection, indicative of an environment that can promote metastatic tumor growth. The chemotherapeutic agent gemcitabine depletes G-MDSCs, M-MDSCs, macrophages, and eosinophils in the lungs of 4T1 tumor-bearing mice, and we found that treating mice with gemcitabine after primary tumor resection decreases residual G-MDSCs in the lungs and decreases subsequent metastatic growth. CONCLUSIONS: Our data support the development of therapeutic strategies to target MDSCs and to monitor MDSC levels before and after primary tumor resection to enhance the effectiveness of immune-based therapies and improve the treatment of metastatic breast cancer in the clinic.

Emerging roles of T helper 17 and regulatory T cells in lung cancer progression and metastasis.

Lung cancer is a leading cause of cancer-related deaths worldwide. Lung cancer risk factors, including smoking and exposure to environmental carcinogens, have been linked to chronic inflammation. An integral feature of inflammation is the activation, expansion and infiltration of diverse immune cell types, including CD4+ T cells. Within this T cell subset are immunosuppressive regulatory T (Treg) cells and pro-inflammatory T helper 17 (Th17) cells that act in a fine balance to regulate appropriate adaptive immune responses.In the context of lung cancer, evidence suggests that Tregs promote metastasis and metastatic tumor foci development. Additionally, Th17 cells have been shown to be an integral component of the inflammatory milieu in the tumor microenvironment, and potentially involved in promoting distinct lung tumor phenotypes. Studies have shown that the composition of Tregs and Th17 cells are altered in the tumor microenvironment, and that these two CD4+ T cell subsets play active roles in promoting lung cancer progression and metastasis.We review current knowledge on the influence of Treg and Th17 cells on lung cancer tumorigenesis, progression, metastasis and prognosis. Furthermore, we discuss the potential biological and clinical implications of the balance among Treg/Th17 cells in the context of the lung tumor microenvironment and highlight the potential prognostic function and relationship to metastasis in lung cancer.

SHIP represses lung inflammation and inhibits mammary tumor metastasis in BALB/c mice.

SH2-containing-inositol-5'-phosphatase (SHIP) is a negative regulator of the phosphatidylinositol-3-kinase pathway in hematopoietic cells and limits the development of leukemias and lymphomas. The potential role of SHIP in solid tumor development and metastasis remains unknown. While SHIP restricts the aberrant development of myeloid cells in C57BL/6 mice, there are conflicting reports regarding the effect of SHIP deletion in BALB/c mice with important consequences for determining the influence of SHIP in different model tumor systems. We generated SHIP-/- BALB/c mice and challenged them with syngeneic non-metastatic 67NR or metastatic 4T1 mammary tumors. We demonstrate that SHIP restricts the development, alternative-activation, and immunosuppressive function of myeloid cells in tumor-free and tumor-bearing BALB/c mice. Tumor-free SHIP-/- BALB/c mice exhibited pulmonary inflammation, myeloid hyperplasia, and M2-polarized macrophages and this phenotype was greatly exacerbated by 4T1, but not 67NR, tumors. 4T1-bearing SHIP-/- mice rapidly lost weight and died from necrohemorrhagic inflammatory pulmonary disease, characterized by massive infiltration of pulmonary macrophages and myeloid-derived suppressor cells that were more M2-polarized and immunosuppressive than wild-type cells. Importantly, while SHIP loss did not affect primary tumor growth, 4T1-bearing SHIP-/- mice had 7.5-fold more metastatic tumor cells in their lungs than wild-type mice, consistent with the influence of immunosuppressive myeloid cells on metastatic growth. Our findings identify the hematopoietic cell-restricted protein SHIP as an intriguing target to influence the development of solid tumor metastases, and support development of SHIP agonists to prevent the accumulation of immunosuppressive myeloid cells and tumor metastases in the lungs to improve treatment of metastatic breast cancer.

Emerging roles of regulatory T cells in tumour progression and metastasis.

The metastasis of cancer is a complex and life-threatening process that is only partially understood. Immune suppressive cells are recognized as important contributors to tumour progression and may also promote the development and growth of tumour metastases. Specifically, regulatory T cells (Tregs) have been found to promote primary tumour progression, and emerging pre-clinical data suggests that Tregs may promote metastasis and metastatic tumour growth. While the precise role that Tregs play in metastatic progression is understudied, recent findings have indicated that by suppressing innate and adaptive anti-tumour immunity, Tregs may shield tumour cells from immune detection, and thereby allow tumour cells to survive, proliferate and acquire characteristics that facilitate dissemination. This review will highlight our current understanding of Tregs in metastasis, including an overview of pre-clinical findings and discussion of clinical data regarding Tregs and therapeutic outcome. Evolving strategies to directly ablate Tregs or to inhibit their function will also be discussed. Improving our understanding of how Tregs may influence tumour metastasis may lead to novel treatments for metastatic cancer.

Macrophages are more potent immune suppressors ex vivo than immature myeloid-derived suppressor cells induced by metastatic murine mammary carcinomas.

Myeloid-derived suppressor cells (MDSCs) are emerging as potential promoters of metastatic tumor growth, and there is interest in targeting immature MDSCs by inducing their differentiation into more mature myeloid cells. We used all-trans retinoic acid (ATRA) to differentiate MDSCs in mice bearing metastatic 4T1 or 4TO7 murine mammary tumors, and assessed the immune-suppressive mechanisms and potencies of different myeloid cell subpopulations. Metastatic mammary tumors induced the accumulation of distinct populations of immature CD11b(+)Gr1(+)F4/80(-)Ly6C(mid)Ly6G(+) MDSCs ("Gr1(+) cells") and mature CD11b(+)Gr1(-)F4/80(+) cells ("F4/80(+) cells") in metastatic target organs. ATRA triggered the differentiation of Gr1(+) cells into F4/80(+) cells in the lungs and, unexpectedly, enhanced pulmonary metastatic tumor growth. We found that F4/80(+)Ly6C(-)Ly6G(-) mature macrophages (Ms) were up to 30-fold more potent immune suppressors than Gr1(+) cells on a per-cell basis, which we postulate may contribute to the increased metastatic growth observed with ATRA treatment. F4/80(+) cells and Gr1(+) cells used different reactive oxygen species (ROS)-mediated mechanisms of immunosuppression ex vivo, with F4/80(+) cells producing higher levels of ROS, which is consistent with their superior immunosuppressive abilities. These data highlight the potent immunosuppressive functions of Ms, reveal that Ms can suppress T cell responses via ROS production, and suggest that ROS inhibitors may be useful in promoting antitumor immune responses. Our findings also caution against using ATRA to modulate myeloid cell differentiation and function to treat breast cancer metastases in the lung, and support the development of therapeutic strategies to enhance antitumor immunity by targeting myeloid cells as a collective group.