Skip to main content

IL-6 down-regulates HLA class II expression and IL-12 production of human dendritic cells to impair activation of antigen-specific CD4+ T cells

Abstract

Immunosuppression in tumor microenvironments critically affects the success of cancer immunotherapy. Here, we focused on the role of interleukin (IL)-6/signal transducer and activator of transcription (STAT3) signaling cascade in immune regulation by human dendritic cells (DCs). IL-6-conditioned monocyte-derived DCs (MoDCs) impaired the presenting ability of cancer-related antigens. Interferon (IFN)-γ production attenuated by CD4+ T cells co-cultured with IL-6-conditioned MoDCs corresponded with decreased DC IL-12p70 production. Human leukocyte antigen (HLA)-DR and CD86 expression was significantly reduced in CD11b+CD11c+ cells obtained from peripheral blood mononuclear cells (PBMCs) of healthy donors by IL-6 treatment and was STAT3 dependent. Arginase-1 (ARG1), lysosomal protease, cathepsin L (CTSL), and cyclooxygenase-2 (COX2) were involved in the reduction of surface HLA-DR expression. Gene expressions of ARG1, CTSL, COX2, and IL6 were higher in tumor-infiltrating CD11b+CD11c+ cells compared with PBMCs isolated from colorectal cancer patients. Expression of surface HLA-DR and CD86 on CD11b+CD11c+ cells was down-regulated, and T cell-stimulating ability was attenuated compared with PBMCs, suggesting that an immunosuppressive phenotype might be induced by IL-6, ARG1, CTSL, and COX2 in tumor sites of colorectal cancer patients. There was a relationship between HLA-DR expression levels in tumor tissues and the size of CD4+ T and CD8+ T cell compartments. Our findings indicate that IL-6 causes a dysfunction in human DCs that activates cancer antigen-specific Th cells, suggesting that blocking the IL-6/STAT3 signaling pathway might be a promising strategy to improve cancer immunotherapy.

This is a preview of subscription content, access via your institution.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Abbreviations

7AAD:

7-Amino-actinomycin D

ARG1:

Arginase-1

CFSE:

Carboxyfluorescein succinimidyl ester

COX2:

Cyclooxygenase-2

CTL:

Cytotoxic T lymphocyte

CTSL:

Cathepsin L

DC:

Dendritic cell

DMSO:

Dimethyl sulfoxide

ELISA:

Enzyme-linked immunosorbent assay

ERK:

Extracellular signal-related kinase

FACS:

Fluorescence-activated cell sorting

GAPDH:

Glyceraldehyde 3-phosphate dehydrogenase

GM-CSF:

Granulocyte macrophage colony-stimulating factor

gp130:

Glycoprotein 130

HLA:

Human leukocyte antigen

IHC:

Immunohistochemistry

IL-6R:

Interleukin-6 receptor

IL:

Interleukin

JAK:

Janus kinase

mAb:

Monoclonal antibody

MAPK:

Mitogen-activated protein kinase

MDSC:

Myeloid-derived suppressor cell

Median FI:

Median fluorescence intensity

MFI:

Mean fluorescence intensity

MHC:

Major histocompatibility complex

MoDC:

Monocyte-derived dendritic cell

nor-NOHA:

Nw-hydroxy-l-arginine

PBS:

Phosphate-buffered saline

PCR:

Polymerase chain reaction

PD-1:

Programmed cell death protein 1

PD-L1:

Programmed cell death ligand 1

PI3 K:

Phosphoinositide 3-kinase

SD:

Standard deviation

STAT3:

Signal transducer and activator of transcription 3

STEAP:

6-transmembrane epithelial antigen of prostate

TGF:

Transforming growth factor

Th:

Helper T

TIL:

Tumor-infiltrating lymphocyte

References

  1. 1.

    Savage PA, Malchow S, Leventhal DS (2013) Basic principles of tumor-associated regulatory T cell biology. Trends Immunol 34:33–40. doi:10.1016/j.it.2012.08.005

    PubMed Central  Article  CAS  PubMed  Google Scholar 

  2. 2.

    Gabrilovich DI, Ostrand-Rosenberg S, Bronte V (2012) Coordinated regulation of myeloid cells by tumours. Nat Rev Immunol 12:253–268. doi:10.1038/nri3175

    PubMed Central  Article  CAS  PubMed  Google Scholar 

  3. 3.

    Motz GT, Coukos G (2013) Deciphering and reversing tumor immune suppression. Immunity 39:61–73. doi:10.1016/j.immuni.2013.07.005

    PubMed Central  Article  CAS  PubMed  Google Scholar 

  4. 4.

    Hodi FS, O’Day SJ, McDermott DF, Weber RW, Sosman JA, Haanen JB, Gonzalez R, Robert C, Schadendorf D, Hassel JC, Akerley W, van den Eertwegh AJM, Lutzky J, Lorigan P, Vaubel JM, Linette GP, Hogg D, Ottensmeier CH, Lebbé C, Peschel C, Quirt I, Clark JI, Wolchok JD, Weber JS, Tian J, Yellin MJ, Nichol GM, Hoos A, Urba WJ (2010) Improved survival with ipilimumab in patients with metastatic melanoma. N Engl J Med 363:711–723. doi:10.1056/NEJMoa1003466

    PubMed Central  Article  CAS  PubMed  Google Scholar 

  5. 5.

    Wolchok JD, Kluger H, Callahan MK, Postow MA, Rizvi NA, Lesokhin AM, Segal NH, Ariyan CE, Gordon RA, Reed K, Burke MM, Caldwell A, Kronenberg SA, Agunwamba BU, Zhang X, Lowy I, Inzunza HD, Feely W, Horak CE, Hong Q, Korman AJ, Wigginton JM, Gupta A, Sznol M (2013) Nivolumab plus ipilimumab in advanced melanoma. N Engl J Med 369:122–133. doi:10.1056/NEJMoa1302369

    Article  CAS  PubMed  Google Scholar 

  6. 6.

    Brahmer JR, Tykodi SS, Chow LQM, Hwu WJ, Topalian SL, Hwu P, Drake CG, Camacho LH, Kauh J, Odunsi K, Pitot HC, Hamid O, Bhatia S, Martins R, Eaton K, Chen S, Salay TM, Alaparthy S, Grosso JF, Korman AJ, Parker SM, Agrawal S, Goldberg SM, Pardoll DM, Gupta A, Wigginton JM (2012) Safety and activity of anti-PD-L1 antibody in patients with advanced cancer. N Engl J Med 366:2455–2465. doi:10.1056/NEJMoa1200694

    PubMed Central  Article  CAS  PubMed  Google Scholar 

  7. 7.

    Palucka K, Banchereau J (2012) Cancer immunotherapy via dendritic cells. Nat Rev Cancer 12:265–277. doi:10.1038/nrc3258

    PubMed Central  Article  CAS  PubMed  Google Scholar 

  8. 8.

    Palucka K, Banchereau J (2013) Dendritic-cell-based therapeutic cancer vaccines. Immunity 39:38–48. doi:10.1016/j.immuni.2013.07.004

    PubMed Central  Article  CAS  PubMed  Google Scholar 

  9. 9.

    Steinman RM, Banchereau J (2007) Taking dendritic cells into medicine. Nature 449:419–426

    Article  CAS  PubMed  Google Scholar 

  10. 10.

    Steinman RM, Mellman I (2004) Immunotherapy: bewitched, bothered, and bewildered no more. Science 305:197–200. doi:10.1126/science.1099688

    Article  CAS  PubMed  Google Scholar 

  11. 11.

    Hirano T, Ishihara K, Hibi M (2000) Roles of STAT3 in mediating the cell growth, differentiation and survival signals relayed through the IL-6 family of cytokine receptors. Oncogene 19:2548–2556

    Article  CAS  PubMed  Google Scholar 

  12. 12.

    Hirano T, Yasukawa K, Harada H, Taga T, Watanabe Y, Matsuda T, Kashiwamura S, Nakajima K, Koyama K, Iwamatsu A (1986) Complementary DNA for a novel human interleukin (BSF-2) that induces B lymphocytes to produce immunoglobulin. Nature 324:73–76. doi:10.1038/324073a0

    Article  CAS  PubMed  Google Scholar 

  13. 13.

    Kitamura H, Kamon H, Sawa SI, Park SJ, Katunuma N, Ishihara K, Murakami M, Hirano T (2005) IL-6-STAT3 controls intracellular MHC class II αβ dimer level through cathepsin S activity in dendritic cells. Immunity 23:491–502. doi:10.1016/j.immuni.2005.09.010

    Article  CAS  PubMed  Google Scholar 

  14. 14.

    Park SJ, Nakagawa T, Kitamura H, Atsumi T, Kamon H, Sawa SI, Kamimura D, Ueda N, Iwakura Y, Ishihara K, Murakami M, Hirano T (2004) IL-6 regulates in vivo dendritic cell differentiation through STAT3 activation. J Immunol 173:3844–3854. doi:10.4049/jimmunol.173.6.3844

    Article  CAS  PubMed  Google Scholar 

  15. 15.

    Sumida K, Wakita D, Narita Y, Masuko K, Terada S, Watanabe K, Satoh T, Kitamura H, Nishimura T (2012) Anti-IL-6 receptor mAb eliminates myeloid-derived suppressor cells and inhibits tumor growth by enhancing T-cell responses. Eur J Immunol 42:2060–2072. doi:10.1002/eji.201142335

    Article  CAS  PubMed  Google Scholar 

  16. 16.

    Narita Y, Kitamura H, Wakita D, Sumida K, Masuko K, Terada S, Nakano K, Nishimura T (2013) The key role of IL-6-arginase cascade for inducing dendritic cell-dependent CD4(+) T cell dysfunction in tumor-bearing mice. J Immunol 190:812–820. doi:10.4049/jimmunol.1103797

    Article  CAS  PubMed  Google Scholar 

  17. 17.

    Ohtake J, Ohkuri T, Togashi Y, Kitamura H, Okuno K, Nishimura T (2014) Identification of novel helper epitope peptides of Survivin cancer-associated antigen applicable to developing helper/killer-hybrid epitope long peptide cancer vaccine. Immunol Lett 161:20–30. doi:10.1016/j.imlet.2014.04.010

    Article  CAS  PubMed  Google Scholar 

  18. 18.

    Hayashi S, Kumai T, Matsuda Y, Aoki N, Sato S, Kimura K, Kitada M, Tateno M, Celis E, Kobayashi H (2011) Six-transmembrane epithelial antigen of the prostate and enhancer of zeste homolog 2 as immunotherapeutic targets for lung cancer. J Transl Med 9:191. doi:10.1186/1479-5876-9-191

    PubMed Central  Article  CAS  PubMed  Google Scholar 

  19. 19.

    Knüpfer H, Preiss R (2010) Serum interleukin-6 levels in colorectal cancer patients–a summary of published results. Int J Colorectal Dis 25:135–140. doi:10.1007/s00384-009-0818-8

    Article  PubMed  Google Scholar 

  20. 20.

    Mitsunaga S, Ikeda M, Shimizu S, Ohno I, Furuse J, Inagaki M, Higashi S, Kato H, Terao K, Ochiai A (2013) Serum levels of IL-6 and IL-1β can predict the efficacy of gemcitabine in patients with advanced pancreatic cancer. Br J Cancer 108:2063–2069. doi:10.1038/bjc.2013.174

    PubMed Central  Article  CAS  PubMed  Google Scholar 

  21. 21.

    Gupta N, Goswami B, Mittal P (2012) Effect of standard anthracycline based neoadjuvant chemotherapy on circulating levels of serum IL-6 in patients of locally advanced carcinoma breast—a prospective study. Int J Surg 10:638–640. doi:10.1016/j.ijsu.2012.11.007

    Article  PubMed  Google Scholar 

  22. 22.

    Yoshitomi M, Yutani S, Matsueda S, Ioji T, Komatsu N, Shichijo S, Yamada A, Itoh K, Sasada T, Kinoshita H (2012) Personalized peptide vaccination for advanced biliary tract cancer: IL-6, nutritional status and pre-existing antigen-specific immunity as possible biomarkers for patient prognosis. Exp Ther Med 3:463–469. doi:10.3892/etm.2011.424

    PubMed Central  CAS  PubMed  Google Scholar 

  23. 23.

    Hoentjen F (2005) STAT3 regulates NF-B recruitment to the IL-12p40 promoter in dendritic cells. Blood 105:689–696. doi:10.1182/blood-2004-04-1309

    Article  CAS  PubMed  Google Scholar 

  24. 24.

    Kortylewski M, Xin H, Kujawski M, Lee H, Liu Y, Harris T, Drake C, Pardoll D, Yu H (2009) Regulation of the IL-23 and IL-12 balance by Stat3 signaling in the tumor microenvironment. Cancer Cell 15:114–123. doi:10.1016/j.ccr.2008

    PubMed Central  Article  CAS  PubMed  Google Scholar 

  25. 25.

    Morinobu A, Gadina M, Strober W, Visconti R, Fornace A, Montagna C, Feldman GM, Nishikomori R, O’Shea JJ (2002) STAT4 serine phosphorylation is critical for IL-12-induced IFN-gamma production but not for cell proliferation. Proc Natl Acad Sci U S A 99:12281–12286. doi:10.1073/pnas.182618999

    PubMed Central  Article  CAS  PubMed  Google Scholar 

  26. 26.

    Hsich CS, Macatonia SE, Tripp CS, Wolf SF, O’Garra A, Murphy KM (1993) Development of TH1 CD4+ T cells through IL-12 produced by Listeria-induced macrophage. Science 260:547–549. doi:10.1126/science.8097338

    Article  Google Scholar 

  27. 27.

    Kaliński P, Hilkens CM, Snijders A, Snijdewint FG, Kapsenberg ML (1997) IL-12-deficient dendritic cells, generated in the presence of prostaglandin E2, promote type 2 cytokine production in maturing human naive T helper cells. J Immunol 159:28–35

    PubMed  Google Scholar 

  28. 28.

    Nishimura T, Iwakabe K, Sekimoto M, Ohmi Y, Yahata T, Nakui M, Sato T, Habu S, Tashiro H, Sato M, Ohta A (1999) Distinct role of antigen-specific T helper type 1 (Th1) and Th2 cells in tumor eradication in vivo. J Exp Med 190:617–627. doi:10.1084/jem.190.5.617

    PubMed Central  Article  CAS  PubMed  Google Scholar 

  29. 29.

    Hunder NN, Wallen H, Cao J, Hendricks DW, Reilly JZ, Rodmyre R, Jungbluth A, Gnjatic S, Thompson JA, Yee C (2008) Treatment of metastatic melanoma with autologous CD4+ T cells against NY-ESO-1. N Engl J Med 358:2698–2703. doi:10.1056/NEJMoa0800251

    PubMed Central  Article  CAS  PubMed  Google Scholar 

  30. 30.

    Langowski JL, Zhang X, Wu L, Mattson JD, Chen T, Smith K, Basham B, McClanahan T, Kastelein RA, Oft M (2006) IL-23 promotes tumour incidence and growth. Nature 442:461–465. doi:10.1038/nature04808

    Article  CAS  PubMed  Google Scholar 

  31. 31.

    Takahashi N, Ohkuri T, Homma S, Ohtake J, Wakita D, Togashi Y, Kitamura H, Todo S, Nishimura T (2012) First clinical trial of cancer vaccine therapy with artificially synthesized helper/ killer-hybrid epitope long peptide of MAGE-A4 cancer antigen. Cancer Sci 103:150–153. doi:10.1111/j.1349-7006.2011.02106.x

    Article  CAS  PubMed  Google Scholar 

  32. 32.

    Harizi H, Juzan M, Grosset C, Rashedi M, Gualde N (2001) Dendritic cells issued in vitro from bone marrow produce PGE(2) that contributes to the immunomodulation induced by antigen-presenting cells. Cell Immunol 209:19–28. doi:10.1006/cimm.2001.1785

    Article  CAS  PubMed  Google Scholar 

  33. 33.

    Obermajer N, Muthuswamy R, Lesnock J, Edwards RP, Kalinski P (2011) Positive feedback between PGE2 and COX2 redirects the differentiation of human dendritic cells toward stable myeloid-derived suppressor cells. Blood 118:5498–5505. doi:10.1182/blood-2011-07-365825

    PubMed Central  Article  CAS  PubMed  Google Scholar 

  34. 34.

    Laghi L, Bianchi P, Miranda E, Balladore E, Pacetti V, Grizzi F, Allavena P, Torri V, Repici A, Santoro A, Mantovani A, Roncalli M, Malesci A (2009) CD3+ cells at the invasive margin of deeply invading (pT3–T4) colorectal cancer and risk of post-surgical metastasis: a longitudinal study. Lancet Oncol 10:877–884. doi:10.1016/S1470-2045(09)70186-X

    Article  CAS  PubMed  Google Scholar 

  35. 35.

    Huh JW, Lee JH, Kim HR (2012) Prognostic significance of tumor-infiltrating lymphocytes for patients with colorectal cancer. Arch Surg 147:366–372. doi:10.1001/archsurg.2012.35

    Article  PubMed  Google Scholar 

  36. 36.

    Bindea G, Mlecnik B, Tosolini M, Kirilovsky A, Waldner M, Obenauf AC, Angell H, Fredriksen T, Lafontaine L, Berger A, Bruneval P, Fridman WH, Becker C, Pagès F, Speicher MR, Trajanoski Z, Galon J (2013) Spatiotemporal dynamics of intratumoral immune cells reveal the immune landscape in human cancer. Immunity 39(4):782–795. doi:10.1016/j.immuni.2013.10.003

    Article  CAS  PubMed  Google Scholar 

  37. 37.

    Anitei MG, Zeitoun G, Mlecnik B, Marliot F, Haicheur N, Todosi AM, Kirilovsky A, Lagorce C, Bindea G, Ferariu D, Danciu M, Bruneval P, Scripcariu V, Chevallier JM, Zinzindohoué F, Berger A, Galon J, Pagès F (2014) Prognostic and predictive values of the immunoscore in patients with rectal cancer. Clin Cancer Res. 20(7):1891–1899. doi:10.1158/1078-0432.CCR-13-2830

    Article  PubMed  Google Scholar 

  38. 38.

    Maby P, Tougeron D, Hamieh M, Mlecnik B, Kora H, Bindea G, Angell HK, Fredriksen T, Elie N, Fauquembergue E, Drouet A, Leprince J, Benichou J, Mauillon J, Le Pessot F, Sesboué R, Frebourg T, Galon J, Latouche JB (2015) Correlation between density of CD8+ T cell infiltrates in microsatellite unstable colorectal cancers and frameshift mutations: a rationale for personalized immunotherapy. Cancer Res 75(17):3446–3455. doi:10.1158/0008-5472.CAN-14-3051

    Article  CAS  PubMed  Google Scholar 

  39. 39.

    Simpson JAD, Al-Attar A, Watson NFS, Scholefield JH, Ilyas M, Durrant LG (2010) Intratumoral T cell infiltration, MHC class I and STAT1 as biomarkers of good prognosis in colorectal cancer. Gut 59:926–933. doi:10.1136/gut.2009.194472

    Article  CAS  PubMed  Google Scholar 

  40. 40.

    Taube JM, Young GD, McMiller TL, Chen S, Salas JT, Pritchard TS, Xu H, Meeker AK, Fan J, Cheadle C, Berger AE, Pardoll DM, Topalian SL (2015) Differential expression of immune-regulatory genes associated with PD-L1 display in melanoma: implications for PD-1 pathway blockade. Clin Cancer Res 21(17):3969–3976. doi:10.1158/1078-0432.CCR-15-0244

    Article  CAS  PubMed  Google Scholar 

  41. 41.

    Angevin E, Tabernero J, Elez E, Cohen SJ, Bahleda R, van Laethem JL, Ottensmeier C, Lopez-Martin JA, Clive S, Joly F, Ray-Coquard I, Dirix L, Machiels JP, Steven N, Reddy M, Hall B, Puchalski TA, Bandekar R, van de Velde H, Tromp B, Vermeulen J, Kurzrock R (2014) A phase I/II, multiple-dose, dose-escalation study of siltuximab, an anti-interleukin-6 monoclonal antibody, in patients with advanced solid tumors. Clin Cancer Res 20:2192–2204. doi:10.1158/1078-0432.CCR-13-2200

    Article  CAS  PubMed  Google Scholar 

  42. 42.

    Voorhees PM, Manges RF, Sonneveld P, Jagannath S, Somlo G, Krishnan A, Lentzsch S, Frank RC, Zweegman S, Wijermans PW, Orlowski RZ, Kranenburg B, Hall B, Casneuf T, Qin X, van de Velde H, Xie H, Thomas SK (2013) A phase 2 multicentre study of siltuximab, an anti-interleukin-6 monoclonal antibody, in patients with relapsed or refractory multiple myeloma. Br J Haematol 161:357–366. doi:10.1111/bjh.12266

    Article  CAS  PubMed  Google Scholar 

  43. 43.

    Fizazi K, De Bono JS, Flechon A, Heidenreich A, Voog E, Davis NB, Qi M, Bandekar R, Vermeulen JT, Cornfeld M, Hudes GR (2012) Randomised phase II study of siltuximab (CNTO 328), an anti-IL-6 monoclonal antibody, in combination with mitoxantrone/prednisone versus mitoxantrone/prednisone alone in metastatic castration-resistant prostate cancer. Eur J Cancer 48:85–93. doi:10.1016/j.ejca.2011.10.014

    Article  CAS  PubMed  Google Scholar 

  44. 44.

    Coward J, Kulbe H, Chakravarty P, Leader D, Vassileva V, Leinster DA, Thompson R, Schioppa T, Nemeth J, Vermeulen J, Singh N, Avril N, Cummings J, Rexhepaj E, Jirström K, Gallagher WM, Brennan DJ, McNeish IA, Balkwill FR (2011) Interleukin-6 as a therapeutic target in human ovarian cancer. Clin Cancer Res 17:6083–6096. doi:10.1158/1078-0432.CCR-11-0945

    PubMed Central  Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

We are grateful to Ms. Ai Nishiuchi for her technical and secretarial assistance. This work was partially supported by a Grant-in-Aid for Scientific Research (25460584 to Hidemitsu Kitamura), a Research Fellowship for Young Scientists (251464 to Kentaro Sumida), a Grant-in-Aid for Translational Research Network Program from the Ministry of Education, Culture, Sports, Science, and Technology (MEXT), a Health and Labour Sciences Research Grant (11103410 to Hidemitsu Kitamura) from the Ministry of Health, Labour, and Welfare (MHLW), Japan, and by the Joint Research Program of the Institute for Genetic Medicine, Hokkaido University.

Author information

Affiliations

Authors

Corresponding author

Correspondence to Hidemitsu Kitamura.

Ethics declarations

Conflict of interest

All authors declare no conflict of interest.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary material 1 (PDF 941 kb)

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Ohno, Y., Kitamura, H., Takahashi, N. et al. IL-6 down-regulates HLA class II expression and IL-12 production of human dendritic cells to impair activation of antigen-specific CD4+ T cells. Cancer Immunol Immunother 65, 193–204 (2016). https://doi.org/10.1007/s00262-015-1791-4

Download citation

Keywords

  • Dendritic cells
  • Antigen presentation
  • HLA class II
  • IL-12
  • Helper T cells