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Tumor-associated myeloid cells can be activated in vitro and in vivo to mediate antitumor effects

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Abstract

Tumor growth is often accompanied by the accumulation of myeloid cells in the tumors and lymphoid organs. These cells can suppress T cell immunity, thereby posing an obstacle to T cell-targeted cancer immunotherapy. In this study, we tested the possibility of activating tumor-associated myeloid cells to mediate antitumor effects. Using the peritoneal model of B16 melanoma, we show that peritoneal cells (PEC) in tumor-bearing mice (TBM) had reduced ability to secrete nitric oxide (NO) following in vitro stimulation with interferon gamma and lipopolysaccharide, as compared to PEC from control mice. This reduced function of PEC was accompanied by the influx of CD11b+ Gr-1+ myeloid cells to the peritoneal cavity. Nonadherent PEC were responsible for most of the NO production in TBM, whereas in naïve mice NO was mainly secreted by adherent CD11b+ F4/80+ macrophages. Sorted CD11b+ Gr-1− monocytic and CD11b+ Gr-1+ granulocytic PEC from TBM had a reduced ability to secrete NO following in vitro stimulation (compared to naïve PEC), but effectively suppressed proliferation of tumor cells in vitro. In vivo, treatment of mice bearing established peritoneal B16 tumors with anti-CD40 and CpG resulted in activation of tumor-associated PEC, reduction in local tumor burden and prolongation of mouse survival. Inhibition of NO did not abrogate the antitumor effects of stimulated myeloid cells. Taken together, the results indicate that in tumor-bearing hosts, tumor-associated myeloid cells can be activated to mediate antitumor effects.

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Abbreviations

IFN-γ:

Interferon gamma

Mϕ:

Macrophages

PEC:

Peritoneal cells

TLR:

Toll-like receptor

TAM:

Tumor-associated macrophages

TNFα:

Tumor necrosis factor alpha

TBM:

Tumor-bearing mice

References

  1. De Souza AP, Bonorino C (2009) Tumor immunosuppressive environment: effects on tumor-specific and nontumor antigen immune responses. Expert Rev Anticancer Ther 9:1317–1332 (Review)

    Google Scholar 

  2. Ostrand-Rosenberg S, Sinha P, Danna EA, Miller S, Davis C, Dissanayake SK (2004) Antagonists of tumor-specific immunity: tumor-induced immune suppression and host genes that co-opt the anti-tumor immune response. Breast Dis 20:127–135

    PubMed  CAS  Google Scholar 

  3. Mantovani A, Allavena P, Sica A (2004) Tumour-associated macrophages as a prototypic type II polarized phagocyte population: role in tumour progression. Eur J Cancer 40:1660–1667

    Article  PubMed  CAS  Google Scholar 

  4. Mullins DW, Martins RS, Elgert KD (2003) Tumor-derived cytokines dysregulate macrophage interferon-gamma responsiveness and interferon regulatory factor-8 expression. Exp Biol Med (Maywood) 228:270–277

    CAS  Google Scholar 

  5. Jaffe ML, Arai H, Nabel GJ (1996) Mechanisms of tumor-induced immunosuppression: evidence for contact-dependent T cell suppression by monocytes. Mol Med 2:692–701

    PubMed  CAS  Google Scholar 

  6. Taams LS, van Amelsfort JM, Tiemessen MM, Jacobs KM, de Jong EC, Akbar AN, Bijlsma JW, Lafeber FP (2005) Modulation of monocyte/macrophage function by human CD4+CD25+ regulatory T cells. Hum Immunol 66:222–230

    Article  PubMed  CAS  Google Scholar 

  7. Danna EA, Sinha P, Gilbert M, Clements VK, Pulaski BA, Ostrand-Rosenberg S (2004) Surgical removal of primary tumor reverses tumor-induced immunosuppression despite the presence of metastatic disease. Cancer Res 64:2205–2211

    Article  PubMed  CAS  Google Scholar 

  8. Sinha P, Clements VK, Ostrand-Rosenberg S (2005) Reduction of myeloid-derived suppressor cells and induction of M1 macrophages facilitate the rejection of established metastatic disease. J Immunol 174:636–645

    PubMed  CAS  Google Scholar 

  9. Talmadge JE (2007) Pathways mediating the expansion and immunosuppressive activity of myeloid-derived suppressor cells and their relevance to cancer therapy. Clin Cancer Res 13:5243–5248

    Article  PubMed  CAS  Google Scholar 

  10. Gabrilovich DI, Nagaraj S (2009) Myeloid-derived suppressor cells as regulators of the immune system. Nat Rev Immunol 9:162–174 (Review)

    Google Scholar 

  11. Peranzoni E, Zilio S, Marigo I, Dolcetti L, Zanovello P, Mandruzzato S, Bronte V (2010) Myeloid-derived suppressor cell heterogeneity and subset definition. Curr Opin Immunol 22:238–244 (Review)

    Google Scholar 

  12. Apolloni E, Bronte V, Mazzoni A, Serafini P, Cabrelle A, Segal DM, Young HA, Zanovello P (2000) Immortalized myeloid suppressor cells trigger apoptosis in antigen-activated T lymphocytes. J Immunol 165:6723–6730

    PubMed  CAS  Google Scholar 

  13. Huang B, Pan PY, Li Q, Sato AI, Levy DE, Bromberg J, Divino CM, Chen SH (2006) 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 66:1123–1131

    Article  PubMed  CAS  Google Scholar 

  14. Sinha P, Clements VK, Fulton AM, Ostrand-Rosenberg S (2007) Prostaglandin E2 promotes tumor progression by inducing myeloid-derived suppressor cells. Cancer Res 67:4507–4513

    Article  PubMed  CAS  Google Scholar 

  15. Popovic PJ, Zeh HJ III, Ochoa JB (2007) Arginine and immunity. J Nutr 137:1681S–1686S (Review)

    Google Scholar 

  16. Sinha P, Clements VK, Bunt SK, Albelda SM, Ostrand-Rosenberg S (2007) Cross-talk between myeloid-derived suppressor cells and macrophages subverts tumor immunity toward a type 2 response. J Immunol 179:977–983

    PubMed  CAS  Google Scholar 

  17. Leek RD, Harris AL (2002) Tumor-associated macrophages in breast cancer. J Mammary Gland Biol Neoplasia 7:177–189

    Article  PubMed  Google Scholar 

  18. Knowles H, Leek R, Harris AL (2004) Macrophage infiltration and angiogenesis in human malignancy. Novartis Found Symp 256:189–200

    Article  PubMed  CAS  Google Scholar 

  19. Guiducci C, Vicari AP, Sangaletti S, Trinchieri G, Colombo MP (2005) Redirecting in vivo elicited tumor infiltrating macrophages and dendritic cells towards tumor rejection. Cancer Res 65:3437–3446

    PubMed  CAS  Google Scholar 

  20. Buhtoiarov IN, Lum H, Berke G, Sondel PM, Rakhmilevich AL (2006) Synergistic activation of macrophages via CD40 and TLR9 results in T cell independent antitumor effects. J Immunol 176:309–318

    PubMed  CAS  Google Scholar 

  21. Wu Q, Buhtoiarov IN, Sondel PM, Rakhmilevich AL, Ranheim EA (2009) Tumoricidal effects of activated macrophages in a mouse model of chronic lymphocytic leukemia. J Immunol 182:6771–6778

    Article  PubMed  CAS  Google Scholar 

  22. Johnson EE, Buhtoiarov IN, Baldeshwiler MJ, Felder MAR, Van Rooijen N, Sondel PM, Rakhmilevich AL (2011) Enhanced T cell-independent antitumor effect of cyclophosphamide combined with anti-CD40 mAb and CpG in mice. J Immunother 34:76–84

    Article  PubMed  CAS  Google Scholar 

  23. Van Rooijen N, Sanders A (1994) Liposome mediated depletion of macrophages: mechanism of action, preparation of liposomes and applications. J Immunol Methods 174:83–93

    Article  PubMed  Google Scholar 

  24. Buhtoiarov IN, Lum H, Berke G, Paulnock D, Sondel PM, Rakhmilevich AL (2005) CD40 ligation induces antitumor reactivity of murine macrophages via an IFN gamma-dependent mechanism. J Immunol 174:6013–6022

    PubMed  CAS  Google Scholar 

  25. Elkord E, Alcantar-Orozco EM, Dovedi SJ, Tran DQ, Hawkins RE, Gilham DE (2010) T regulatory cells in cancer: recent advances and therapeutic potential. Expert Opin Biol Ther 10:1573–1586

    Article  PubMed  CAS  Google Scholar 

  26. Hegardt P, Widegren B, Sjögren HO (2000) Nitric-oxide-dependent systemic immunosuppression in animals with progressively growing malignant gliomas. Cell Immunol 200:116–127

    Article  PubMed  CAS  Google Scholar 

  27. Youn JI, Nagaraj S, Collazo M, Gabrilovich DI (2008) Subsets of myeloid-derived suppressor cells in tumor-bearing mice. J Immunol 181:5791–5802

    PubMed  CAS  Google Scholar 

  28. Jia W, Jackson-Cook C, Graf MR (2010) Tumor-infiltrating, myeloid-derived suppressor cells inhibit T cell activity by nitric oxide production in an intracranial rat glioma + vaccination model. J Neuroimmunol 223:20–30

    Article  PubMed  CAS  Google Scholar 

  29. Ortega A, Carretero J, Obrador E, Estrela JM (2008) Tumoricidal activity of endothelium-derived NO and the survival of metastatic cells with high GSH and Bcl-2 levels. Nitric Oxide 19:107–114

    Article  PubMed  CAS  Google Scholar 

  30. Vicetti Miguel RD, Cherpes TL, Watson LJ, McKenna KC (2010) CTL induction of tumoricidal nitric oxide production by intratumoral macrophages is critical for tumor elimination. J Immunol 185:6706–6718

    Article  PubMed  Google Scholar 

  31. Lum HD, Buhtoiarov IN, Schmidt BE, Berke G, Paulnock DM, Sondel PM, Rakhmilevich AL (2006) Tumoristatic effects of anti-CD40 mAb-activated macrophages involve nitric oxide and tumor-necrosis factor-α. Immunology 118:261–270

    Article  PubMed  CAS  Google Scholar 

  32. Buhtoiarov IN, Sondel PM, Wigginton JM, Buhtoiarova TN, Yanke EM, Mahvi DA, Rakhmilevich AL (2011) Antitumor synergy of cytotoxic chemotherapy and anti-CD40 plus CpG-ODN immunotherapy through repolarization of tumor associated macrophages. Immunology 132:226–239

    Article  PubMed  CAS  Google Scholar 

  33. Dinapoli MR, Calderon CL, Lopez DM (1996) The altered tumoricidal capacity of macrophages isolated from tumor-bearing mice is related to reduce expression of the inducible nitric oxide synthase gene. J Exp Med 183:1323–1329

    Article  PubMed  CAS  Google Scholar 

  34. Naama HA, McCarter MD, Mack VE, Evoy DA, Hill AD, Shou J, Daly JM (2001) Suppression of macrophage nitric oxide production by melanoma: mediation by a melanoma-derived product. Melanoma Res 11:229–238

    Article  PubMed  CAS  Google Scholar 

  35. Atochina O, Daly-Engel T, Piskorska D, McGuire E, Harn DA (2001) A schistosome-expressed immunomodulatory glycoconjugate expands peritoneal Gr1(+) macrophages that suppress naive CD4(+) T cell proliferation via an IFN-gamma and nitric oxide-dependent mechanism. J Immunol 167:4293–4302

    PubMed  CAS  Google Scholar 

  36. Dunay IR, Damatta RA, Fux B, Presti R, Greco S, Colonna M, Sibley LD (2008) Gr1(+) inflammatory monocytes are required for mucosal resistance to the pathogen Toxoplasma gondii. Immunity 29:306–317

    Article  PubMed  CAS  Google Scholar 

  37. Caso R, Silvera R, Carrio R, Iragavarapu-Charyulu V, Gonzalez-Perez RR, Torroella-Kouri M (2010) Blood monocytes from mammary tumor-bearing mice: early targets of tumor-induced immune suppression? Int J Oncol 37:891–900

    PubMed  CAS  Google Scholar 

  38. Greifenberg V, Ribechini E, Rössner S, Lutz MB (2009) Myeloid-derived suppressor cell activation by combined LPS and IFN-gamma treatment impairs DC development. Eur J Immunol 39:2865–2876

    Article  PubMed  CAS  Google Scholar 

  39. Pelaez B, Campillo JA, Lopez-Asenjo JA, Subiza JL (2001) Cyclophosphamide induces the development of early myeloid cells suppressing tumor cell growth by a nitric oxide-dependent mechanism. J Immunol 166:6608–6615

    PubMed  CAS  Google Scholar 

  40. Beatty GL, Chiorean EG, Fishman MP, Saboury B, Teitelbaum UR, Sun W, Huhn RD, Song W, Li D, Sharp LL, Torigian DA, O’Dwyer PJ, Vonderheide RH (2011) CD40 agonists alter tumor stroma and show efficacy against pancreatic carcinoma in mice and humans. Science 331:1612–1616

    Article  PubMed  CAS  Google Scholar 

  41. Dolcetti L, Peranzoni E, Ugel S, Marigo I, Fernandez Gomez A, Mesa C, Geilich M, Winkels G, Traggiai E, Casati A, Grassi F, Bronte V (2010) Hierarchy of immunosuppressive strength among myeloid-derived suppressor cell subsets is determined by GM-CSF. Eur J Immunol 40:22–35

    Article  PubMed  CAS  Google Scholar 

  42. Sunderkötter C, Nikolic T, Dillon MJ, van Rooijen N, Stehling M, Drevets DA, Leenen PJM (2004) Subpopulations of mouse blood monocytes differ in maturation stage and inflammatory response. J Immunol 172:4410–4417

    PubMed  Google Scholar 

  43. Cui SJ, Reichner JS, Mateo RB, Albina JE (1994) Activated murine macrophages induce apoptosis in tumor-cells through nitric-oxide-dependent or nitric-oxide-independent mechanisms. Cancer Res 54:2462–2467

    PubMed  CAS  Google Scholar 

  44. Klostergaard J, Leroux ME, Hung MC (1991) Cellular-models of macrophage tumoricidal effector mechanisms in vitro: characterization of cytolytic responses to tumor-necrosis-factor and nitric-oxide pathways in vitro. J Immunol 147:2802–2808

    PubMed  CAS  Google Scholar 

  45. Decker T, Lohmannmatthes ML, Gifford GE (1987) Cell-associated tumor-necrosis-factor (TNF) as a killing mechanism of activated cytotoxic macrophages. J Immunol 138:957–962

    PubMed  CAS  Google Scholar 

  46. Ellyard JI, Quah BJ, Simson L, Parish CR (2010) Alternatively activated macrophage possess antitumor cytotoxicity that is induced by IL-4 and mediated by arginase-1. J Immunother 33:443–452

    Article  PubMed  CAS  Google Scholar 

  47. Egilmez NK, Harden JL, Virtuoso LP, Schwendener RA, Kilinc MO (2011) Nitric oxide short-circuits interleukin-12-mediated tumor regression. Cancer Immunol Immunother 60:839–845

    Article  PubMed  CAS  Google Scholar 

  48. Fukumura D, Kashiwagi S, Jain RK (2006) The role of nitric oxide in tumour progression. Nat Rev Cancer 6:521–534 (Review)

    Google Scholar 

  49. Wang GY, Ji B, Wang X, Gu JH (2005) Anti-cancer effect of iNOS inhibitor and its correlation with angiogenesis in gastric cancer. World J Gastroenterol 1:3830–3833

    Google Scholar 

  50. Sikora AG, Gelbard A, Davies MA, Sano D, Ekmekcioglu S, Kwon J, Hailemichael Y, Jayaraman P, Myers JN, Grimm EA, Overwijk WW (2010) Targeted inhibition of inducible nitric oxide synthase inhibits growth of human melanoma in vivo and synergizes with chemotherapy. Clin Cancer Res 16:1834–1844

    Article  PubMed  CAS  Google Scholar 

  51. Liao S, Cheng G, Conner DA, Huang Y, Kucherlapati RS, Munn LL, Ruddle NH, Jain RK, Fukumura D, Padera TP (2011) Impaired lymphatic contraction associated with immunosuppression. Proc Natl Acad Sci USA 108:18784–18789

    Article  PubMed  CAS  Google Scholar 

  52. Tsung K, Dolan JP, Tsung YL, Norton JA (2002) Macrophages as effector cells in interleukin 12-induced T cell-dependent tumor rejection. Cancer Res 62:5069–5075

    PubMed  CAS  Google Scholar 

  53. Weiss JM, Ridnour LA, Back T, Hussain SP, He P, Maciag AE, Keefer LK, Murphy WJ, Harris CC, Wink DA, Wiltrout RH (2010) Macrophage-dependent nitric oxide expression regulates tumor cell detachment and metastasis after IL-2/anti-CD40 immunotherapy. J Exp Med 207:2455–2467

    Article  PubMed  CAS  Google Scholar 

  54. Zoglmeier C, Bauer H, Noerenberg D, Wedekind G, Bittner P, Sandholzer N, Rapp M, Anz D, Endres S, Bourquin C (2011) CpG blocks immune suppression by myeloid-derived suppressor cells in tumor-bearing mice. Clin Cancer Res 17:1765–1775

    Article  PubMed  CAS  Google Scholar 

  55. Mandruzzato S, Solito S, Falisi E, Francescato S, Chiarion-Sileni V, Mocellin S, Zanon A, Rossi CR, Nitti D, Bronte V, Zanovello P (2009) IL4Ralpha+ myeloid-derived suppressor cell expansion in cancer patients. J Immunol 182:6562–6568

    Article  PubMed  CAS  Google Scholar 

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Acknowledgments.

We thank Songwon Seo for performing correlative statistical analysis and Jennifer Levenson for breeding EGFP transgenic mice. This work was supported by NIH-NCI grants CA87025 and CA32685, a grant from the Midwest Athletes Against Childhood Cancer (MACC) Fund and support from The Crawdaddy Foundation.

Conflict of interest

The authors declare that they have no conflict of interest.

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Authors and Affiliations

  1. Departments of Human Oncology, University of Wisconsin, 4136 WIMR, 1111 Highland Avenue, Madison, WI, 53705, USA

    Alexander L. Rakhmilevich, Mark J. Baldeshwiler, Tyler J. Van De Voort, Richard K. Yang, Nicholas A. Kalogriopoulos, David S. Koslov & Paul M. Sondel

  2. Paul P. Carbone Comprehensive Cancer Center, University of Wisconsin, Madison, WI, USA

    Alexander L. Rakhmilevich & Paul M. Sondel

  3. Department of Obstetrics and Gynecology, University of Wisconsin, Madison, WI, USA

    Mildred A. R. Felder

  4. Department of Pediatrics, University of Wisconsin, Madison, WI, USA

    Paul M. Sondel

  5. Department of Molecular Cell Biology, Vrije Universiteit Medical Centre, Van der Boechorststraat, Amsterdam, The Netherlands

    Nico Van Rooijen

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  1. Alexander L. Rakhmilevich
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  2. Mark J. Baldeshwiler
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  3. Tyler J. Van De Voort
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Correspondence to Alexander L. Rakhmilevich.

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262_2012_1236_MOESM1_ESM.doc

Supplemental figure. Effect of B16 cells in vivo and in vitro on PEC function and phenotype. A. C57BL/6 mice were injected i.p. with 1.6x107 B16 cells or left untreated. PEC were obtained 4 days later, placed in 96-well plate, and plastic-adherent cells were stimulated with IFN-γ (10 U/ml) and LPS (1 ng/ml). The supernatants were collected 48 hr later and NO activity was determined by nitrite levels. The results are presented as the Mean ± SEM nitrite concentration from 4 mice per group. B. PEC were obtained from naïve C57BL/6 mice, and 2x105 PEC were allowed to adhere to plastic for 2 hr followed by removal of nonadherent cells before adding B16 cells at indicated doses to adherent PEC in the presence of IFN-γ and LPS. NO activity in supernatants was determined 48 hr later. The results are presented as the Mean ± SEM nitrite concentration from triplicates. C. PEC obtained from naïve C57BL/6 mice (5x106) and RAW 247.7 macrophage cell line cells (2.5x106) were placed in the wells of the 6-well plates. PEC were allowed to adhere to plastic for 2 hr followed by removal of nonadherent cells. The cells were incubated in medium with or without gamma-irradiated (200 Gy) B16 cells (2.5x106) for 3 days, after which the cells were analyzed for co-expression of F4/80 and Gr-1 by flow cytometry. The representative dot plots are shown. (DOC 163 kb)

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Rakhmilevich, A.L., Baldeshwiler, M.J., Van De Voort, T.J. et al. Tumor-associated myeloid cells can be activated in vitro and in vivo to mediate antitumor effects. Cancer Immunol Immunother 61, 1683–1697 (2012). https://doi.org/10.1007/s00262-012-1236-2

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