Enhanced suppressive capacity of tumor-infiltrating myeloid-derived suppressor cells compared with their peripheral counterparts
Sarah K. Maenhout
Department of Immunology-Physiology, Laboratory of Molecular and Cellular Therapy, Vrije Universiteit Brussel, Jette, Belgium
Search for more papers by this authorSandra Van Lint
Department of Immunology-Physiology, Laboratory of Molecular and Cellular Therapy, Vrije Universiteit Brussel, Jette, Belgium
Search for more papers by this authorPerpetua U. Emeagi
Department of Immunology-Physiology, Laboratory of Molecular and Cellular Therapy, Vrije Universiteit Brussel, Jette, Belgium
Search for more papers by this authorKris Thielemans
Department of Immunology-Physiology, Laboratory of Molecular and Cellular Therapy, Vrije Universiteit Brussel, Jette, Belgium
Search for more papers by this authorCorresponding Author
Joeri L. Aerts
Department of Immunology-Physiology, Laboratory of Molecular and Cellular Therapy, Vrije Universiteit Brussel, Jette, Belgium
Correspondence to: J.L. Aerts, Laboratory of Molecular and Cellular Therapy, Vrije Universiteit Brussel, Laarbeeklaan 103E, 1090 Jette, Belgium, Tel.: +32-2-477-45-64, Fax: +32-2-477-45-68, E-mail: [email protected]Search for more papers by this authorSarah K. Maenhout
Department of Immunology-Physiology, Laboratory of Molecular and Cellular Therapy, Vrije Universiteit Brussel, Jette, Belgium
Search for more papers by this authorSandra Van Lint
Department of Immunology-Physiology, Laboratory of Molecular and Cellular Therapy, Vrije Universiteit Brussel, Jette, Belgium
Search for more papers by this authorPerpetua U. Emeagi
Department of Immunology-Physiology, Laboratory of Molecular and Cellular Therapy, Vrije Universiteit Brussel, Jette, Belgium
Search for more papers by this authorKris Thielemans
Department of Immunology-Physiology, Laboratory of Molecular and Cellular Therapy, Vrije Universiteit Brussel, Jette, Belgium
Search for more papers by this authorCorresponding Author
Joeri L. Aerts
Department of Immunology-Physiology, Laboratory of Molecular and Cellular Therapy, Vrije Universiteit Brussel, Jette, Belgium
Correspondence to: J.L. Aerts, Laboratory of Molecular and Cellular Therapy, Vrije Universiteit Brussel, Laarbeeklaan 103E, 1090 Jette, Belgium, Tel.: +32-2-477-45-64, Fax: +32-2-477-45-68, E-mail: [email protected]Search for more papers by this authorAbstract
Although the main site of action for myeloid-derived suppressor cells (MDSCs) is most likely the tumor microenvironment, so far the study of these cells has been largely restricted to spleen-derived MDSCs. In this study, we compared the suppressive capacity of splenic and tumor-derived MDSCs in different subcutaneous mouse tumor models. We investigated which suppressive mechanisms were involved. Finally, we investigated whether MDSCs and regulatory T cells (Treg) cooperate in the suppression of T-cell responses. In all models, splenic granulocytic MDSCs (grMDSC) strongly suppress CD4+ T-cell proliferation while the suppressive effect on CD8+ T cells is less pronounced. Splenic monocytic MDSCs (moMDSC) have a lower suppressive capacity, compared to grMDSC, on both CD4+ and CD8+ T-cell proliferation. Both grMDSC and moMDSC isolated from the tumor have a much stronger suppressive activity compared to MDSCs isolated from the spleen of tumor-bearing mice, associated with a higher NO2− production by the tumor-derived moMDSC and arginase activity for both subsets. The expression of CD80 is also elevated on tumor-derived grMDSC compared with their peripheral counterparts. Direct contact with tumor cells is required for the upregulation of CD80 and CD80+ MDSCs are more suppressive than CD80− MDSCs. Coculture of Treg and MDSCs leads to a stronger suppression of CD8+ T-cell proliferation compared to the suppression observed by Treg or MDSCs alone. Thus, we showed that tumor-infiltrating MDSCs possess a stronger suppressive capacity than their peripheral counterparts and that various suppressive mechanisms account for this difference.
Abstract
What's new?
Attempts to wield the body's immune system against cancer often fail. One reason is the suppression of T cells by myeloid-derived suppressor cells (MDSCs). This study investigated exactly how MDSCs thwart T cells. They found that MDSCs isolated from the solid tumor were far more potent against T cells than those from the spleen, and that they express more CD80. Furthermore, when MDSCs were cultured together with regulatory T cells, that improved their ability to suppress T cells. These findings suggest possible ways to counter the immunosuppressive tumor microenvironment.
Supporting Information
Additional Supporting Information may be found in the online version of this article.
| Filename | Description |
|---|---|
| ijc28449-sup-0001-suppinfo01.tif533.4 KB | Supplementary Figure 1: Representative FACS plot of the cell sort of grMDSC and moMDSC. First, myeloid cells were gated based on their forward scatter and side scatter. Within the CD11b+ cell population both the Ly6G+Ly6Cint granulocytic MDSCs (upper panel) and the Ly6C+LY6G− momocytic MDSCs (lower panel) were sorted to high purity. One representative FACS plot for all the cell sort experiments is shown and a purity of at least 90% for both cell populations was achieved in each of the performed experiments. |
| ijc28449-sup-0002-suppinfo02.tif772.2 KB | Supplementary Figure 2: MDSCs isolated from the spleen of LLC or MO4 tumor-bearing mice suppress proliferation of both CD4+ and CD8+ T cells. Sorted MDSCs were cultured at different ratios [ranging from 1:1 to 1:8 (MDSCs:splenocytes)] with 2×105 CellTrace violet labeled splenocytes form healthy mice in the presence of anti-CD3/CD28 beads for 3 days after which proliferation was determined. A, Overview of the percentage suppression of CD4+ and CD8+ T-cell proliferation in the presence of MDSC-G (upper panels) or MDSC-M (lower panel) in the MO4 model and the LLC model (B). Three independent expeiments wete performed and results are presented as mean ± SEM for the MO4 model. For the LLC model, 2 independent experiments were performed for the MDSC-M and results are presented as the mean. For the MDSC-G, 3 independent experiment were performed and results are presented aa mean ± SEM. *, statistically significant differences from values of T-cell proliferation in the absence of MDSCs (P < 0.005). NS, no statistically significant differences from values of T-cell proliferation in the absece of MDSCs. |
| ijc28449-sup-0003-suppinfo03.tif692.4 KB | Supplementary Figure 3: MoMDSC isolated from within the tumor microenvironment possess a stronger suppressive capacity compared to their peripheral counterparts in the MO4 model. MDSC-M were isolated from the spleen and tumor microenvironment of MO4 tumor-bearing mice and cultured with 2×105 Cell Trace Violet labelled splenocytes from healthy mice in the presece of anti-CD3/CD28 beads for 3 days after which proliferation of CD8+ (upper panel) and CD4+ (lower panel) T cells was determined by flow cytometry. Two independent experiments were performed and results are presented as mean. |
| ijc28449-sup-0004-suppinfo04.tif500.9 KB | Supplementary Figure 4: Representative FACS plot of the cell sort of Treg. A, Based on the expression of CD4 and CD25 the double positive T-cells were sorted to high purity. One representative FACS plot for all the cell sort experiments is shown and a purity of at least 90% was achieved in each of the performed experiments. B, The sorted cells were stained with an antibody against FoxP3 in order to confirm that the sorted cells were Treg. One representative FACS plot is shown. |
| ijc28449-sup-0005-suppinfo05.tif694.3 KB | Supplementary Figure 5 : Inhibition of T-cell proliferation by Treg isolated from the spleen of E.G7-OVA tumor-bearing mice. Treg were isolated from the spleen of E.G7-OVA tumor-bearing mice and used in a suppression assay. A, After 3 days, proliferation of CD8+ T cells was determined by flow cytometry. One representative experiment is shown. B, proliferation of CD8+ T cells after co-culture with different ratios (ranging from 1:2 to 1:8 (Treg:splenocytes)) of Treg and percentage of suppression by Treg was calculated. Results of at least 3 independent experiments are presented as mean ° SEM. *, statistically significant differences from values of T-cell proliferation in the absence of Treg (p < 0.05). NS, no statistically significantn differences from values of T-cell proliferation in the absence of Treg. C, IFNβ, TNF-α and IL-2 production by splenocytes was determined after 3 days of culture with Treg. Results are presented as the mean of 3 independent experiments. |
Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.
References
- 1 Pardoll D. Does the immune system see tumors as foreign or self? Annu Rev Immunol 2003; 21: 807–39.
- 2 Zitvogel L, Tesniere A, Kroemer G. Cancer despite immunosurveillance: immunoselection and immunosubversion. Nat Rev Immunol 2006; 6: 715–27.
- 3 Gabrilovich DI, Nagaraj S. Myeloid-derived suppressor cells as regulators of the immune system. Nat Rev Immunol 2009; 9: 162–74.
- 4 Poschke I, Kiessling R. On the armament and appearances of human myeloid-derived suppressor cells. Clin Immunol 2012; 144: 250–68.
- 5 Facciabene A, Motz GT, Coukos G. T-regulatory cells: key players in tumor immune escape and angiogenesis. Cancer Res 2012; 72: 2162–71.
- 6 Gabrilovich DI, Bronte V, Chen S-H, et al. The terminology issue for myeloid-derived suppressor cells. Cancer Res 2007; 67: 425.
- 7 Ostrand-Rosenberg S. Myeloid-derived suppressor cells: more mechanisms for inhibiting antitumor immunity. Cancer Immunol Immunother 2010; 59: 1593–600.
- 8 Serafini P, Borrello I, Bronte V. Myeloid suppressor cells in cancer: recruitment, phenotype, properties, and mechanisms of immune suppression. Semin Cancer Biol 2006; 16: 53–65.
- 9 Ribechini E, Greifenberg V, Sandwick S, et al. Subsets, expansion and activation of myeloid-derived suppressor cells. Med Microbiol Immunol 2010; 199: 273–81.
- 10 Fleming TJ, Fleming M, Malek TR. Selective expression of Ly-6G on myeloid lineage cells in mouse bone marrow. RB6-8C5 mAb to Granulocyte-Differentiation Antigen (Gr-1) Detects Members of the Ly-6 Family. J Immunol 1993; 151: 2399–408.
- 11 Peranzoni E, Zilio S, Marigo I, et al. Myeloid-derived suppressor cell heterogeneity and subset definition. Curr Opin Immunol 2010; 22: 238–44.
- 12 Movahedi K, Guilliams M, Van den Bossche J, et al. Identification of discrete tumor-induced myeloid-derived suppressor cell subpopulations with distinct T cell-suppressive activity. Blood 2008; 111: 4233–44.
- 13 Apolloni E, Bronte V, Mazzoni a, et al. Immortalized myeloid suppressor cells trigger apoptosis in antigen-activated T lymphocytes. J Immunol 2000; 165: 6723–30.
- 14 Almand B, Clark JI, Nikitina E, et al. Increased production of immature myeloid cells in cancer patients: a mechanism of immunosuppression in cancer. J Immunol 2001; 166: 678–89.
- 15 Nagaraj S, Nelson A, Youn J, et al. Antigen-specific CD4(+) T cells regulate function of myeloid-derived suppressor cells in cancer via retrograde MHC class II signaling. Cancer Res 2012; 72: 928–38.
- 16 Subudhi SK, Alegre ML, Fu YX. The balance of immune responses: costimulation verse coinhibition. J Mol Med 2005; 83: 193–202.
- 17 Seliger B, Marincola FM, Ferrone S, et al. The complex role of B7 molecules in tumor immunology. Trends Mol Med 2008; 14: 550–9.
- 18 Bour-Jordan H, Esensten JH, Martinez-Llordella M, et al. Intrinsic and extrinsic control of peripheral T-cell tolerance by costimulatory molecules of the CD28/ B7 family. Immunol Rev 2011; 241: 180–205.
- 19 Zang X, Allison JP. The B7 family and cancer therapy: costimulation and coinhibition. Clin Cancer Res 2007; 13: 5271–9.
- 20 Zheng Y, Manzotti CN, Liu M, et al. CD86 and CD80 differentially modulate the suppressive function of human regulatory T cells. J Immunol 2004; 172: 2778–84.
- 21 Sinha P, Okoro C, Foell D, et al. Proinflammatory S100 proteins regulate the accumulation of myeloid-derived suppressor cells. J Immunol 2008; 181: 4666–75.
- 22 Youn JI, Nagaraj S, Collazo M, et al. Subsets of myeloid-derived suppressor cells in tumor-bearing mice. J Immunol 2008; 181: 5791–802.
- 23 Liu Y, Yu Y, Yang S, et al. Regulation of arginase I activity and expression by both PD-1 and CTLA-4 on the myeloid-derived suppressor cells. Cancer Immunol Immunother 2009; 58: 687–97.
- 24 Yang R, Cai Z, Zhang Y, et al. CD80 in immune suppression by mouse ovarian carcinoma-associated Gr-1+CD11b+ myeloid cells. Cancer Res 2006; 66: 6807–15.
- 25 Butte MJ, Keir ME, Phamduy TB, et al. PD-L1 interacts specifically with B7-1 to inhibit T cell proliferation. Immunity 2007; 27: 111–22.
- 26 Kusmartsev S, Nagaraj S, Gabrilovich DI. Tumor-associated CD8+ T cell tolerance induced by bone marrow-derived immature myeloid cells. J Immunol 2005; 175: 4583–92.
- 27 Srivastava MK, Sinha P, Clements VK. Myeloid-derived suppressor cells inhibit T-cell activation by depleting cystine and cysteine. Cancer Res 2010; 70: 68–77.
- 28 Sinha P, Clements VK, Ostrand-Rosenberg S. Interleukin-13-regulated M2 macrophages in combination with myeloid suppressor cells block immune surveillance against metastasis. Cancer Res 2005; 65: 11743–51.
- 29 Gabrilovich DI, Velders MP, Sotomayor EM, et al. Mechanism of immune dysfunction in cancer mediated by immature Gr-1+ myeloid cells. J Immunol 2001; 166: 5398–406.
- 30 Corzo Ca, Condamine T, Lu L, et al. HIF-1α regulates function and differentiation of myeloid-derived suppressor cells in the tumor microenvironment. J Exp Med 2010; 207: 2439–53.
- 31 Haverkamp JM, Crist Sa, Elzey BD, et al. In vivo suppressive function of myeloid-derived suppressor cells is limited to the inflammatory site. Eur J Immunol 2011; 41: 749–59.
- 32 Gallina G, Dolcetti L, Serafini P, et al. Tumors induce a subset of inflammatory monocytes with immunosuppressive activity on CD8+ T cells. J Clin Invest 2006; 116: 2777–90.
- 33 Schlecker E, Stojanovic A, Eisen C, et al. Tumor-infiltrating monocytic myeloid-derived suppressor cells mediate CCR5-dependent recruitment of regulatory T cells favoring tumor growth. J Immunol 2012; 189: 5602–11.
- 34 Greten TF, Manns MP, Korangy F. Myeloid derived suppressor cells in human disease. Int Immunopharmacol 2011; 11: 802–7.
- 35 Filipazzi P, Huber V, Rivoltini L. Phenotype, function and clinical implications of myeloid-derived suppressor cells in cancer patients. Cancer ImmunoI Immunother 2012; 61: 255–63.
- 36 Diaz-montero CM. Increased circulating myeloid-derived suppressor cells correlate with clinical cancer stage,metastatic tumor burden, and doxorubicin-cyclophosphamide chemotherapy. Cancer Immunol Immunother 2009; 58: 49–59.
- 37 Gros A, Turcotte S, Wunderlich JR, et al. Myeloid cells obtained from the blood but not from the tumor can suppress T cell proliferation in patients with melanoma. Clin Cancer Res 2012; 18: 5212–23.
- 38 Serafini P, Mgebroff S, Noonan K, et al. Myeloid-derived suppressor cells promote cross-tolerance in B-cell lymphoma by expanding regulatory T cells. Cancer Res 2008; 68: 5439–49.
- 39 Hoechst B, Gamrekelashvili J, Manns MP, et al. Plasticity of human Th17 cells and iTregs is orchestrated by different subsets of myeloid cells. Blood 2011; 117: 6532–41.
- 40 Tang S, Lotze MT. The power of negative thinking: which cells limit tumor immunity? Clin Cancer Res 2012; 18: 5157–9.
- 41 Tomihara K, Guo M, Shin T, et al. Antigen-specific immunity and cross-priming by epithelial ovarian carcinoma-induced CD11b(+)Gr-1(+) cells. J Immunol 2010; 184: 6151–60.
- 42 Poschke I, Mougiakakos D, Hansson J, et al. Immature immunosuppressive CD14+HLA-DR-/low cells in melanoma patients are Stat3hi and overexpress CD80, CD83, and DC-sign. Cancer Res 2010; 70: 4335–45.
- 43 Huang B, Pan PY, Li Q, et al. Gr-1+CD115+ immature myeloid suppressor cells mediate the development of tumor-induced T regulatory cells and T-cell anergy in tumor-bearing host. Cancer Res 2006; 66: 1123–31.
- 44 Fujimura T, Ring S, Umansky V, et al. Regulatory T cells stimulate B7-H1 expression in myeloid-derived suppressor cells in ret melanomas. J Invest Dermatol 2012; 132: 1239–46.
- 45 Mellor AL, Munn DH. IDO expression by dendritic cells: tolerance and tryptophan catabolism. Nat Rev Immunol 2004; 4: 762–74.
- 46 Bauer TM, Jiga LP, Chuang J-J, et al. Studying the immunosuppressive role of indoleamine 2,3-dioxygenase: tryptophan metabolites suppress rat allogeneic T-cell responses in vitro and in vivo. Transpl Int 2005; 18: 95–100.
Citing Literature
