Macrophages Are More Potent Immune Suppressors Ex Vivo Than Immature Myeloid-Derived Suppressor Cells Induced by Metastatic Murine Mammary Carcinomas

Abstract

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−Ly6CmidLy6G+ 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 (Mϕs) 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 Mϕs, reveal that Mϕs 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.

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

“…M-MDSCs are more immunosuppressive than G-MDSCs [22], and we previously reported that macrophages infiltrating the lungs of 4T1 tumor-bearing mice are potently immunosuppressive [35]. Importantly, the decrease of M-MDSCs and macrophages in the lungs after primary 4T1 tumor resection (Fig.…”

“…Clinical studies have demonstrated the efficacy of pharmacological strategies to reduce MDSC number (e.g., sunitinib) [59], to inhibit MDSC suppressive function (e.g., sildenafil) [60], or to differentiate MDSCs into mature myeloid cells (e.g., all-trans retinoic acid or 25hydroxyvitamin D 3 ) [28][29][30] in a variety of human cancers. We have previously shown that ATRA-mediated differentiation of MDSCs can promote metastatic tumor growth by generation of highly immunosuppressive macrophages [35], and therefore, strategies to target or inhibit MDSCs may produce more predictable outcomes. Directly targeting MDSCs with 5-fluorouracil [61] or gemcitabine [62] in various murine models of cancer significantly enhances T cell-dependent antitumor immunity, suggesting that therapeutics which target MDSCs may work synergistically with T cell-targeted therapies.…”

“…Clonogenic assays from disaggregated lung tissue were performed as previously reported [34,35]. Briefly, singlecell suspensions derived from lung tissue were washed in PBS, and aliquots of 3 × 10 3 to 10 6 cells were plated in triplicate in medium containing 60 μM 6-thioguanine to select for the 6-thioguanine-resistant 4T1 tumor cells.…”

Immunotherapeutic Blockade of Macrophage Clever-1 Reactivates the CD8+ T-cell Response against Immunosuppressive Tumors

“…Although DCs are reportedly more efficient at T-cell priming than TAMs, macrophages can also prime CD8 þ T cells to generate cytotoxic effector cells and CD8 þ T-cell memory in vivo (32,35). Moreover, tumorigenic TAMs are extremely potent immunosuppressors both individually and through sheer numbers (32,36,37). For example, tumor-infiltrating CD8 þ T cells mostly come into contact with TAMs because of their high frequency, and TAMs can directly induce CD8 þ T-cell apoptosis and physically restrict CD8 þ T cells from reaching their target cells ) 32 , 38 , 39…”

Location, location, location: functional and phenotypic heterogeneity between tumor-infiltrating and non-infiltrating myeloid-derived suppressor cells

“…These data highlight the potent immunosuppressive functions of macrophages and support the development of therapeutic strategies to enhance antitumor immunity by targeting inhibitory myeloid cells as a collective group. 12 Moreover, in analogy to the M1/M2 polarization in macrophages, MDSCs also show an M1/M2 classification in the tumor microenvironment. Different studies have shown that MDSCs present within the tumor microenvironment exhibit M2 characteristics, which accelerates tumor growth which is mediated by enhanced arginase activity, an increased secretion of immunosuppressive cytokines and the induction of angiogenesis.…”