Early-stage Myeloid-derived Suppressor Cells Research Articles

SOCS3 deficiency-dependent autophagy repression promotes the survival of early-stage myeloid-derived suppressor cells in breast cancer by activating the Wnt/mTOR pathway.

Early-stage myeloid-derived suppressor cells are a newly defined subset of myeloid-derived suppressor cells in breast cancer tissues and related to poor prognosis in patients with breast cancer. Compared with classical myeloid-derived suppressor cells, early-stage myeloid-derived suppressor cells display exceptional immunosuppressive ability and accumulate in the tumor microenvironment to suppress innate and adaptive immunity. Previously, we demonstrated that early-stage myeloid-derived suppressor cells were SOCS3 deficiency dependent and correlated with differentiation arrest in the myeloid lineage. Autophagy is a major regulator of myeloid differentiation, but the mechanism by which autophagy regulates the development of early-stage myeloid-derived suppressor cells has not been elucidated. Here, we constructed EO771 mammary tumor-bearing conditional myeloid SOCS3 knockout mice (SOCS3MyeKO) characterized by abundant tumor-infiltrating early-stage myeloid-derived suppressor cells and exacerbated immunosuppression in vitro and in vivo. We found that early-stage myeloid-derived suppressor cells isolated from SOCS3MyeKO mice showed differentiation arrest in the myeloid lineage, which was caused by limited autophagy activation in an Wnt/mTOR-dependent manner. RNA sequencing and microRNA microarray assays revealed that miR-155-induced C/EBPβ downregulation activated the Wnt/mTOR pathway and promoted autophagy repression and differentiation arrest in early-stage myeloid-derived suppressor cells. Furthermore, inhibition of Wnt/mTOR signaling suppressed both tumor growth and the immunosuppressive functions of early-stage myeloid-derived suppressor cells. Thus, SOCS3 deficiency-dependent autophagy repression and their regulatory mechanisms could contribute to the immunosuppressive tumor microenvironment. Our study proposes a novel mechanism for promoting early-stage myeloid-derived suppressor cell survival, which might shed new light on a potential target of oncologic therapy.

Myeloid-derived suppressor cells in multiple myeloma patients at the immune recovery and after high-dose chemotherapy and hematopoietic stem cell transplantation

Introduction. Myeloid-derived suppressor cells (MDSCs) play an important role in restriction of the immune response and are associated with a poor prognosis in cancer. Mobilization of hematopoietic stem cells (HSCs) before high-dose chemotherapy (HCT) with autologous HSC transplantation (auto-HSCT) is accompanied by a signifcant increase in MDSC counts in peripheral blood and apheresis product of multiple myeloma (MM) patients. However, quantitative changes of these cells at the post-transplant and their role at the immune recovery remain unexplored.The study was aimed to analyze the dynamics of circulating MDSC counts and the expression of suppressor molecule arginase-1 in patients with MM in the frst 12 months after HCT and auto-HSCT and evaluate association between MDSCs and transplantation outcomes.Material and Methods. The study included 44 MM patients who underwent HCT and auto-HSCT. The relative number of granulocytic MDSCs (G-MDSCs), monocytic MDSCs (M-MDSCs), and early-stage MDSCs (E-MDSCs), as well as the expression of arginase-1 in each of MDSC subsets was evaluated by fow cytometry in patient peripheral blood samples.Results. At the engraftment (day +12 – +16, leukocytes >1×109 /l), M-MDSC relative count was increased (pU=0.038), as well as the relative (pU=0.003) and absolute (pU˂0.0001) counts of G-MDSCs, decreasing after 6 months down to pre-transplant values (рU=0.007, рU=0.024 and рU=0.02, respectively) and remaining at the same level at the 12-month follow-up period. The absolute count of E-MDSCs by the time of the engraftment decreased transiently (pU=0.004 vs before HCT), gradually recovering by 12-month follow-up (pU=0.032 vs day +12 – +16). The remission within 12 months in the group with G-MDSCs˂0.17 % at the engraftment was observed in 67 ± 11 % of patients, with G-MDSCs >0.17 % – in 94 ± 6 % of patients (p=0.049). During the 12-month post-transplant, the number of M-MDSCs expressing arginase-1 has been increasing, with a tendency to lower values at the engraftment in patients with early MM relapse (pU=0.09).Conclusion. The association of early MM relapse after auto-HSCT with the lower count of G-MDSCs and the lower count of arginase-1+ M-MDSCs at the engraftment suggests that MDSCs is involved in the restriction of homeostatic proliferation as a factor for more effective immune recovery.

Expression of Inhibitory Molecules (Arginase-1, IDO, and PD-L1) by Myeloid-Derived Suppressor Cells in Multiple Myeloma Patients in Remission.

We studied suppressor potential of myeloid-derived suppressor cells (MDSC) in multiple myeloma patients, including before and after mobilization of hematopoietic stem cells (HSC), by evaluating the expression of arginase-1 (Arg1), indolamine-2,3-dioxygenase (IDO), and PD-L1 in MDSC subsets. The study included 20 multiple myeloma patients in remission, 5 patients with progression, as well as 10 sex-and age-matched healthy donors. The expression of Arg1, IDO, and PD-L1 in circulating granulocytic MDSC (G-MDSC, Lin-HLA-DR-CD33+CD66b+), monocytic MDSC (M-MDSC, CD14+HLA-DRlow/-), and early-stage MDSC (E-MDSC, Lin-HLA-DR-CD33+CD66b-) was evaluated by flow cytometry. Multiple myeloma patients in remission were characterized by reduced expression of Arg1 in M-MDSC in comparison with donors. The expression of Arg1 in M-MDSC depended on the number of induction therapy lines performed and was significantly lower in patients who received ⩾2 lines and responded with remission. Patients with multiple myeloma progression (resistant to therapy) showed significantly increased expression of Arg1 and PD-L1 in M-MDSC, as well as increased expression of Arg1 in E-MDSC. After G-CSF-induced mobilization of HSC, the content of circulating Arg1-expressing M-MDSC increased significantly. Considering the presence of MDSC in apheresis products, MDSC suppressive activity is discussed as a factor affecting the outcomes of autologous HSC transplantation in multiple myeloma patients.

Circulating PMN-MDSC level positively correlates with a poor prognosis in patients with metastatic hormone-sensitive prostate cancer

IntroductionThis study aims to investigated for the first time the role of myeloid-derived suppressor cells (MDSCs) in metastatic-hormone sensitive prostate cancer (mHSPC), which has not been investigated previously.Materials and methodsThis was a prospective observational cohort study. MDSC subsets in peripheral blood samples were classified and evaluated by flow cytometry as early-stage MDSCs (e-MDSCs), polymorphonuclear MDSCs (PMN-MDSCs), and monocytic MDSCs (M-MDSCs). The prostate-specific antigen progression free survival (PSA–PFS) and overall survival (OS) were evaluated to assess the prognostic value of each of the MDSC subsets. The immune cell dynamics and gene expression alteration were analyzed by single-cell RNA-sequencing (scRNA-seq) in a representative case.ResultsThirty-one mHSPC patients and 11 healthy controls (HCs) were included in this study. There were significantly more PMN/M-MDSCs in mHSPC patients than in HCs (p <0.05) before treatment, but the numbers became similar to those in HCs after treatment. Although there were no marked differences in the high and low ratios of e-MDSCs and M-MDSCs, patients with a high ratio of PMN-MDSCs (≥0.30%) had a poorer PSA–PFS and OS than those with a low ratio (<0.30%) (p <0.05). scRNA-seq showed that the expression of genes implicated in tumor progression was upregulated in a representative mHSPC case.ConclusionsA high frequency of PMN-MDSCs correlated with poor prognosis in mHSPC patients. PMN-MDSCs and their highly expressed genes are potential novel therapeutic targets for mHSPC.

Abstract 239: Assessment of immunotherapeutic efficacy of entinostat by using the 3d-explore ex vivo platform utilizing tumoroids of fresh patient tumor samples with intact tumor microenvironment

Abstract Background: Entinostat, a selective histone deacetylase 1/3 inhibitor has been shown to promote a robust anti-tumor response and increased neoantigen expression and antigen-specific T cell activation in preclinical tumor models. Entinostat plays a role in neutralizing myeloid derived suppressor cells (MDSC) to improve anti-tumor immune response. Entinostat is currently being tested in later-stage clinical trials in various types of cancer. Using a novel 3D ex-vivo platform with fresh patient tumor samples we assessed the therapeutic efficacy of Entinostat with intact stromal components and tumor immune microenvironment. Methods: All tumor samples were obtained with patient consent and relevant IRB approval. Unpropagated 3D tumoroids with intact TME measuring 150 µm in size were prepared from fresh tumor samples from renal cell carcinoma (RCC) patients using a proprietary technology developed at Nilogen Oncosystems. Tumoroids prepared from each patient’s tumor sample were pooled to represent the tumor heterogeneity and treated ex vivo with Entinostat for 48h to detect treatment-mediated changes in tumor immune cell composition including lymphoid and myeloid markers and their activation status. Results: Tumoroids treated with Entinostat increased NK, NKT, CD4 and CD8+T cells compared to untreated control. Ex vivo treatment with Entinostat led to an increase in CD69 and Granzyme B expression in tumor resident CD4+ T cells. Immune checkpoint receptors, 41BB and PD-1 expression was increased in CD8+T cells followed by Entinostat treatment. Decreased monocytic MDSC was seen in Entinostat treated tumoroids compared to untreated control and this was associated with decreased GM-CSF, IL-4, IL-6 cytokine levels and increased IFN-γ and IL-2, quantified by multiplex cytokine assay. Interestingly, the tumor cell killing study revealed a higher percentage of death in early stage MDSC (eMDSC) indicating a direct effect of Entinostat on MDSC survival. Conclusions: These results demonstrate that Entinostat treatment leads to a broad range of immune cell activation and MDSC inhibition in RCC. These data provide a mechanistic rationale utilizing our 3D tumoroid platform for discovery of potential biomarkers of drug sensitivity and to identify novel immunotherapeutic approaches. Clinical applications of 3D-EXplore may help to identify RCC patients who may likely benefit from Entinostat treatment. Citation Format: Krithika Nandakumar Kodumudi, Brittany Bunch, Jared Ehrhart, Matt Weitzman, Olivia MacIntosh, Kelly Sussman, Soner Altiok. Assessment of immunotherapeutic efficacy of entinostat by using the 3d-explore ex vivo platform utilizing tumoroids of fresh patient tumor samples with intact tumor microenvironment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 239.

Expansion of myeloid-derived suppressor cells in the peripheral blood of patients with ankylosing spondylitis

Expansion of myeloid-derived suppressor cells (MDSCs) due to impaired differentiation of myeloid progenitor cells under conditions of inflammation was described in a number of autoimmune diseases, including rheumatoid arthritis, systemic lupus erythematosus, and type 1 diabetes mellitus. Studying the role of MDSCs in ankylosing spondylitis is an important issue, given that increased concentration of proinflammatory mediators in this pathology can also cause myelopoiesis disorders. The aim of present work was to study the quantitative content of MDSC subpopulations in patients with different clinical phenotypes and activity of AS. 37 patients, including 10 patients without peripheral skeletal lesions (axial form) and 27 patients with simultaneous lesions of spine and peripheral joints (peripheral form) were recruited into the study. The control group consisted of 32 age/sex-related healthy donors. Evaluation of granulocytic (LinHLA-DRCD33+CD66b+; G-MDSC), monocytic (CD14+HLA-DRlow/-; M-MDSC) and early-stage MDSCs (LinHLA-DRCD33+CD66b- ; E-MDSC) was performed using corresponding antibodies (BD Biosciences, USA) in the population of peripheral blood mononuclear cells by flow cytometry. In general, the AS patients were characterized by an increased relative and absolute amount of M-MDSC (p = 0.00002 and p = 0.00003, respectively) and G-MDSC (p = 0.0002 and p = 0.0006, respectively). Patient gender, age, and HLA-B27 expression did not significantly affect the content of these cells in peripheral blood. An increase in the median values of M-MDSC was detected both in patients with axial (Ме 5.0 (3.2-6.3) versus 2.4 (1.7-3.5) %; p = 0.001) and peripheral form (Ме 5.0 (3.0-7.0) versus 2.4 (1.7-3.5) %; p = 0.0002) AS. At the same time, the G-MDSC expansion was observed only in patients with involvement of peripheral joints (Ме 0.16 (0.07-0.3) % versus 0.05 (0.04-0.09) %; p = 0.0001). The relative contents of E-MDSC, M-MDSC and G-MDSC in the axial form of AS was in direct correlation with the activity of the disease (R = 0.58, p = 0.02; R = 0.73, p = 0.08 and R = 0.65 p = 0.04, respectively). This relationship was not observed in peripheral form of AS. The data obtained suggest a potential involvement of MDSCs in pathogenesis and phenotypic heterogeneity of AS. Simultaneously, the revealed direct correlation between the MDSC contents and the disease activity suggests a decrease in suppressive activity and/or appearance of pro-inflammatory activity in MDSC, thus requiring further research in the field.

Increased Levels of Circulating Monocytic- and Early-Stage Myeloid-Derived Suppressor Cells (MDSC) in Acute Myeloid Leukemia.

To investigate the levels of circulating myeloid-derived suppressor cells (MDSC) in patients with primary acute myeloid leukemia (AML), and to explore the relationship between the number of MDSC and AML. Peripheral blood samples from 29 patients with primary AML and 30 healthy controls were collected. CD33, CD11b, HLA-DR, CD14, and CD15 were used to label cells, and flow cytometry was used to analyze the numbers of total MDSC and subgroups eMDSC (early-stage MDSC), M-MDSC (monocytic MDSCs), PMN-MDSC (polymorphonuclear-MDSCs) or G-MDSC (granulocytic-MDSC) via two gating strategies. Presence of MDSC in AML was determined after assessment of clinical data. Phenotypic analysis of MDSC by the two gating strategies was consistent. Compared with healthy controls, the numbers of total MDSC (CD33+CD11b+ HLA-DR-) and G-MDSC (CD33+CD11b+ HLA-DR-CD14¬-CD15+ or CD14¬-CD15+ CD11b+) in peripheral blood of AML patients were lower (p < 0.05), while numbers of M-MDSC (CD33+CD11b+ HLA-DR-CD14+CD15- or HLA-DR-/LOWCD14+) and eMDSC (CD33+CD11b+ HLA-DR-/LOWCD14-CD15-) were higher (p < 0.05). The levels of G-MDSC in peripheral blood of AML-M2 patients were higher than those in other subtypes, along with total MDSC, while the levels of eMDSC and M-MDSC in AML-M3 patients were higher than those in other subtypes. The high frequency of HLA-DR-/LOWCD14+M-MDSC and CD33+CD11b+ HLA-DR-/LOWCD14-CD15- eMDSC in peripheral blood of AML patients indicates potential for MDSC as a diagnostic index in AML.

Immunophenotyping of Circulating Myeloid-Derived Suppressor Cells (MDSC) in the Peripheral Blood of Cancer Patients.

Myeloid-derived suppressor cells (MDSC) are a heterogeneous group of pathologically expanded myeloid cells with immunosuppressive activity. According to their phenotype, MDSC can be divided into three major subpopulations: early stage MDSC (e-MDSC), lacking myeloid lineage markers, monocytic MDSC (M-MDSC), and granulocytic MDSC (PMN-MDSC). Additionally, PMN-MDSC can be subdivided based on their activation and differentiation status, although it is not clear how this status contributes to immunosuppression and disease pathology. Here, we describe an immunophenotyping and gating strategy for the identification and isolation of MDSC subsets based on fluorescence-activated cell sorting. This method allows direct comparison of MDSC subsets in clinical settings.

Abstract 985: SOCS3 deficiency blocked autophagy-dependent myeloid differentiation of early-stage myeloid-derived suppressor cells via the miR-155/C/EBPβ/Wnt axis

Abstract Early-stage myeloid-derived suppressor cells (eMDSCs) are a newly defined subset of myeloid-derived suppressor cells (MDSCs) that accumulate densely in tumors and potently promote tumor growth and metastasis by suppressing antitumor immune responses in vitro and in vivo. We previously identified a subset of eMDSCs in human breast cancer with a characteristic phenotype of Lin-HLA-DR-CD33+. We also found that SOCS3 deficiency and sustained activation of the JAK/STAT signaling pathway are critical molecular events coordinating the differentiation of eMDSCs, although the distinct molecular regulation has not been fully elucidated. Herein, we genetically constructed conditional SOCS3 knockout mice with SOCS3 deficiency specifically in the myeloid linage (SOCS3MyeKO). We delineated that the number of eMDSCs in SOCS3MyeKO mice (eMDSCsSOCS3KO) with the murine GMP phenotype increased significantly, and these cells suffered from a differentiation block in the myeloid lineage. To explore which pathways participated in dysfunctional eMDSC differentiation, we performed whole-genome RNA sequencing and miRNA microarray on CD11b+Gr-1+ cells, eMDSCsfl/fl and eMDSCsSOCS3KO to screen the potential regulatory ceRNA network in eMDSCsSOCS3KO. CD11b+Gr-1+ cells isolated from SOCS3fl/fl mouse spleens were used as mature myeloid cell controls. The transcriptome results and corresponding intervention experiment revealed that the differentiation block in eMDSCsSOCS3KO was caused by SOCS3 deficiency-mediated limited autophagy activation in an AMPK-independent manner. The results of miRNA microarray and RNA sequencing demonstrated that miR-155 overexpression and Wnt/β-catenin pathway activation were involved in the SOCS3 knockout-mediated myeloid differentiation block and autophagy repression. Further experiments revealed that miR-155 was induced by activation of the STAT3/NK-κB pathway upon SOCS3 deficiency, which consequently activated the Wnt/β-catenin pathway via targeting C/EBPβ. Furthermore, applying a specific miR-155 antagonist or the autophagy agonist rapamycin efficiently suppressed tumor growth and eMDSC infiltration in vivo. Overall, these findings indicated that SOCS3 deficiency blocked autophagy-dependent myeloid differentiation of e-MDSCs via the miR-155/C/EBPβ/Wnt axis, and thus targeted therapy against this pathway could be a potential therapeutic target in breast cancer. Citation Format: Wenwen Zhang, Mengmeng Jiang, Rui Zhang, Pengpeng Liu, Yingnan Ye, Xiubao Ren, Wenwen Yu, Jinpu Yu. SOCS3 deficiency blocked autophagy-dependent myeloid differentiation of early-stage myeloid-derived suppressor cells via the miR-155/C/EBPβ/Wnt axis [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 985.

Quantification of Early-Stage Myeloid-Derived Suppressor Cells in Cancer Requires Excluding Basophils.

Myeloid derived suppressor cells (MDSC) are a heterogeneous group of immature cells that accumulate in the peripheral blood and tumor microenvironment and are barriers to cancer therapy. MDSCs serve as prognostic biomarkers and are targets for therapy. On the basis of surface markers, three subsets of MDSCs have been defined in humans: granulocytic, monocytic, and early stage (e-MDSC). The markers attributed to e-MDSCs overlap with those of basophils, which are rare circulating myeloid cells with unrecognized roles in cancer. Thus, we asked whether e-MDSCs in circulation and the tumor microenvironment include basophils. On average, 58% of cells with e-MDSC surface markers in blood and 36% in ascites from patients with ovarian cancer were basophils based on CD123high expression and cytology, whereas cells with immature features were rare. Circulating and ascites basophils did not suppress proliferation of stimulated T cells, a key feature of MDSCs. Increased accumulation of basophils and basogranulin, a marker of basophil degranulation, were observed in ascites compared to serum in patients with newly diagnosed ovarian cancer. Basophils recruited to the tumor microenvironment may exacerbate fluid accumulation by their release of proinflammatory granular constituents that promote vascular leakage. No significant correlation was observed between peripheral basophil counts and survival in patients with ovarian cancer. Our results suggest that studies in which e-MDSCs were defined solely by surface markers should be reevaluated to exclude basophils. Both immaturity and suppression are criteria to define e-MDSCs in future studies.

Circulating and tumor-infiltrating arginase 1-expressing cells in gastric adenocarcinoma patients were mainly immature and monocytic Myeloid-derived suppressor cells

Myeloid-derived suppressor cells (MDSCs) are a group of heterogeneous cells derived from immature myeloid cells (IMCs). MDSCs are known to play important roles in tumor immune evasion. While we know that there are a large number of circulating and tumor-infiltrating MDSCs existing in gastric cancer (GC) patients, the phenotypic characteristics and arginase 1 (ARG1) expression levels of these MDSCs remain very unclear. In our study, flow cytometric analysis of circulating MDSCs from 20 gastric adenocarcinoma (GAC) patients found that ≥80% ARG1-expressing MDSCs were mainly early-stage MDSCs (HLA-DR−CD33+CD14−CD15−MDSCs). In addition, our investigation showed that tumor-infiltrating MDSCs from 6 GAC patients consisted of >35% ARG1-expressing naïve MDSCs (HLA-DR−CD33−CD11b−CD14−CD15−MDSCs), >15% early-stage MDSCs and >40% monocytic MDSCs (HLA-DR−CD14+MDSCs). This preliminary study describes the phenotypic characteristics and ARG1 expression levels of MDSCs from GAC patients and shows that circulating and tumor-infiltrating ARG1-expressing cells were mainly immature and monocytic MDSCs, which provides information to better understand the mechanisms that allow gastric cancer cells to evade the immune system.

Cancer exosome-derived miR-9 and miR-181a promote the development of early-stage MDSCs via interfering with SOCS3 and PIAS3 respectively in breast cancer

We previously identified that the development of early-stage myeloid-derived suppressor cells (eMDSCs) in breast cancer with high IL-6 (IL-6high) expression was correlated with the SOCS3 deficiency-dependent hyperactivation of the JAK/STAT signaling pathway. However, the regulatory mechanisms have not yet been elucidated. In this study, we aimed to investigate how the posttranscriptional regulation mediated by cancer exosome-derived miRNAs affected the JAK/STAT signaling pathway and the development of eMDSCs. Using miRNA microarray, we screened miR-9 and miR-181a which were exclusively upregulated in eMDSCs and inversely associated with SOCS3 expression. We found both miRNAs promoted the amplification of immature eMDSCs with the strong suppression on T-cell immunity in mice and humans. Furthermore, miR-9 and miR-181a promoted 4T1 tumor growth and immune escape via enhancing eMDSCs infiltration in situ. But miR-9 and miR-181a stimulated eMDSCs development by separately inhibiting SOCS3 and PIAS3, two crucial regulators in the negative feedback loop of the JAK/STAT signaling pathway. Elevated miR-9 and miR-181a in eMDSCs was derived from tumor-derived exosomes, and blocking the exosome release could fully attenuate the miRNA-mediated regulation on eMDSCs development. In summary, our findings indicated that tumor exosome-derived miR-9 and miR-181a activated the JAK/STAT signaling pathway via targeting SOCS3 and PIAS3, respectively, and thus promoted the expansion of eMDSCs which might provide potential therapeutic target for IL-6high breast cancer treatment.

Myeloid-Derived Suppressor Cells in Kidney Transplant Recipients and the Effect of Maintenance Immunotherapy.

Myeloid-derived suppressor cells (MDSC) represent a heterogeneous group of myeloid regulatory cells that were originally described in cancer. Several studies in animal models point to MDSC as important players in the induction of allograft tolerance due to their immune modulatory function. Most of the published studies have been performed in animal models, and the data addressing MDSCs in human organ transplantation are scarce. We evaluated the phenotype and function of different MDSCs subsets in 38 kidney transplant recipients (KTRs) at different time points. Our data indicate that monocytic MDSCs (Mo-MDSC) increase in KTR at 6 and 12 months posttransplantation. On the contrary, the percentages of polymorphonuclear MDSC (PMN-MDSC) and early-stage MDSC (e-MDSC) are not significantly increased. We evaluated the immunosuppressive activity of Mo-MDSC in KTR and confirmed their ability to increase regulatory T cells (Treg) in vitro. Interestingly, when we compared the ability of Mo-MDSC to suppress T cell proliferation, we observed that tacrolimus, but not rapamycin-treated KTR, was able to inhibit CD4+ T cell proliferation in vitro. This indicates that, although mTOR inhibitors are widely regarded as supportive of regulatory responses, rapamycin may impair Mo-MDSC function, and suggests that the choice of immunosuppressive therapy may determine the tolerogenic pathway and participating immune cells that promote organ transplant acceptance in KTR.

TAM Family Receptor Kinase Inhibition Reverses MDSC-Mediated Suppression and Augments Anti-PD-1 Therapy in Melanoma.

Myeloid cell receptor tyrosine kinases TYRO3, AXL, and MERTK and their ligands, GAS6 and PROTEIN S, physiologically suppress innate immune responses, including in the tumor microenvironment. Here, we showed that myeloid-derived suppressor cells (MDSC) dramatically upregulated TYRO3, AXL, and MERTK and their ligands [monocytic MDSCs (M-MDSC)>20-fold, polymorphonuclear MDSCs (PMN-MDSC)>15-fold] in tumor-bearing mice. MDSCs from tumor-bearing Mertk-/-, Axl-/- , and Tyro3-/- mice exhibited diminished suppressive enzymatic capabilities, displayed deficits in T-cell suppression, and migrated poorly to tumor-draining lymph nodes. In coimplantation experiments using TYRO3-/-, AXL-/-, and MERTK-/- MDSCs, we showed the absence of these RTKs reversed the protumorigenic properties of MDSCs in vivo Consistent with these findings, in vivo pharmacologic TYRO3, AXL, and MERTK inhibition diminished MDSC suppressive capability, slowed tumor growth, increased CD8+ T-cell infiltration, and augmented anti-PD-1 checkpoint inhibitor immunotherapy. Mechanistically, MERTK regulated MDSC suppression and differentiation in part through regulation of STAT3 serine phosphorylation and nuclear localization. Analysis of metastatic melanoma patients demonstrated an enrichment of circulating MERTK+ and TYRO3+ M-MDSCs, PMN-MDSCs, and early-stage MDSCs (e-MDSC) relative to these MDSC populations in healthy controls. These studies demonstrated that TYRO3, AXL, and MERTK control MDSC functionality and serve as promising pharmacologic targets for regulating MDSC-mediated immune suppression in cancer patients.

SOCS3 Suppression Promoted the Recruitment of CD11b+Gr-1-F4/80-MHCII- Early-Stage Myeloid-Derived Suppressor Cells and Accelerated Interleukin-6-Related Tumor Invasion via Affecting Myeloid Differentiation in Breast Cancer.

Interleukin-6 (IL-6) is an important trigger for the expansion and recruitment of myeloid-derived suppressor cells (MDSCs), which are regarded to be major coordinators of the immunosuppressive tumor microenvironment. In this study, we constructed IL-6-knockdown breast cancer mice models to explore the molecular events involved in the IL-6-mediated effects on MDSC development. We defined a subset of early-stage MDSCs (e-MDSCs) with the phenotype of CD11b+Gr-1−F4/80−MHCII− in IL-6 high-expressing 4T1 mice mammary carcinoma models, which were the precursors of CD11b+Gr-1+ conventional MDSCs. Furthermore, sustained suppression of SOCS3 and aberrant hyperactivation of the JAK/STAT signaling pathway was exclusively detected in wide-type 4T1 tumor-bearing mice, which promoted the accumulation of e-MDSCs in situ and their immunosuppressive capability in vitro. After blocking the IL-6/STAT3 signaling pathway with the IL-6 receptor antibody or STAT3 antagonist JSI-124 in tumor-bearing mice, significant shrinkage of primary tumors and decrease in lung metastatic nodules were observed in vivo, accompanied by the dramatic decrease of e-MDSC recruitment and recovery of anti-tumor T cell immunity. Thus, SOCS3 suppression accelerated the IL-6-mediated growth and metastasis of mammary carcinoma via affecting myeloid differentiation in breast cancer. Moreover, the IL-6/STAT3 signaling pathway might be a promising candidate target in developing novel therapeutic strategies to eliminate e-MDSCs and improve breast cancer prognosis.

Abstract 5081: IL-6 trans-signaling pathway promotes early-stage myeloid derived suppressor cells (eMDSCs) via SOCS3 suppression in breast cancer

Abstract IL-6 is known to be an important trigger of the expansion and recruitment of myeloid-derived suppressor cells (MDSCs) which are regarded as major coordinators of immunosuppressive microenvironment in cancer. But key molecular events are not fully understood yet. We previously identified a subset of early-stage MDSCs (eMDSCs) with immature myeloid phenotypes in IL-6 over-expressing breast cancer tissues. In this study we elucidated how tumor-derived IL-6 manipulated the development and function of eMDSCs in vitro and vivo. We isolated primary eMDSCs in situ and induced eMDSCs in vitro separately, as well as constructed IL-6 knockdown 4T1 mammary tumor-bearing mice models to explore the molecular events involved. We found that in primary human breast cancer tissues, tumor-derived IL-6 was positively correlated with the accumulation of eMDSCs in situ, which in turn accelerated tumor progression and unfavorable clinical outcomes. IL-6 significantly stimulated the amplification of eMDSCs and promoted their T cell suppressive capacity in vitro. And IL-6 trans-signaling pathway via soluble IL-6 receptor CD126 induced SOCS3 suppression and aberrant hyperphosphorylation of the JAK/STAT signaling pathway in eMDSCs, which promoted the accumulation of eMDSCs in vitro and vivo. Correspondingly, we defined a similar immature subset of CD11b+Gr-1- eMDSCs in IL-6 over-expressing 4T1 murine mammary cancer-bearing mice which displayed more potent suppression on T cell immunity than conventional CD11b+Gr-1+ MDSCs. The proportion of CD11b+Gr-1- eMDSCs in tumor tissues was highly correlated with the tumor size and the numbers of lung metastatic nodules compared to the CD11b+Gr-1+ MDSCs. Tumor-derived IL-6 inducing the inhibition of SOCS3 and the activation of the JAK/STAT signaling pathway was observed in CD11b+Gr-1- MDSCs simultaneously, which further caused the differentiation arrest of myeloid linkage and comparable immunosuppressive capacity of eMDSCs. In order to inhibit IL-6-triggerd activation of the JAK/STAT pathway, specific IL-6R blocking antibody and STAT antagonist JSI-124 were prescribed to 4T1 tumor-bearing mice. We found significant shrinkage of primary tumor and decrease of lung metastatic nodules in vivo, along with recovery of the differentiation of myeloid linkage and attenuation of eMDSC-mediated T cell suppression in vitro. Therefore, we concluded that IL-6-induced dysfunction of the SOCS feedback loop and hyperactivation of the JAK/STAT signaling pathway produced significant differentiation arrest in myeloid linkage, which promoted the accumulation of eMDSCs, the defect in local immune surveillance, and consequential tumor progression in breast cancer. Target therapy against this pathway efficiently decelerated the growth and metastasis of breast cancer in vivo which might be a promising therapeutic strategy for breast cancer treatment. Citation Format: Jinpu Yu. IL-6 trans-signaling pathway promotes early-stage myeloid derived suppressor cells (eMDSCs) via SOCS3 suppression in breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5081.