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Expansion and characteristics of human T regulatory type 1 cells in co-cultures simulating tumor microenvironment

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Cancer Immunology, Immunotherapy Aims and scope Submit manuscript

Abstract

Objective

Chronic inflammation and cancer development are associated with dysregulated immune responses and the presence of regulatory T cells (Treg). To study the role of Treg in tumor cell escape from immune surveillance, an in vitro model simulating the tumor microenvironment and promoting the induction and expansion of IL-10+ Treg type 1 (Tr1) was established.

Methods

An in vitro co-culture system (IVA) included an irradiated head and neck squamous cell carcinoma cell line, immature dendritic cells (iDC), CD4+CD25− T cells and cytokines, IL-2 (10 IU/ml), IL-10 (20 IU/ml), IL-15 (20 IU/ml) ± 1 nM rapamycin. Autologous iDC and CD4+CD25− T cells were obtained from the peripheral blood of 15 normal donors. Co-cultures were expanded for 10 days. Proliferating lymphocytes were phenotyped by multi-color flow cytometry. Their suppressor function was measured in CFSE inhibition assays ± neutralizing anti-IL-10 mAb and using transwell cultures. Culture supernatants were tested for IL-4, IL-10, TGF-β and IFN-γ in ELISA.

Results

In the IVA, low doses of IL-2, IL-10 and IL-15 promoted induction and expansion of CD3+CD4+CD25−IL2Rβ+IL2Rγ+FoxP3+CTLA-4+IL-10+ cells with suppressor activity (mean suppression ± SD = 58 ± 12%). These suppressor cells produced IL-10 (mean ± SD = 535 ± 12 pg/ml) and TGF-β (mean ± SD = 512 ± 38 pg/ml), but no IL-4 or IFN-γ. Suppressor function of co-cultures correlated with the percent of expanding IL-10+ Tr1 cells (r 2 = 0.9; P < 0.001). The addition of rapamycin enriched Tr1 cells in all co-cultures. Neutralizing anti-IL-10 mAb abolished suppressive activity. Suppression was cell-contact independent.

Conclusion

The tumor microenvironment promotes generation of Tr1 cells which have the phenotype distinct from that of CD4+CD25highFoxP3+ nTreg and mediate IL-10 dependent immune suppression in a cell-contact independent manner. Tr1 cells may play a critical role in cancer progression.

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Acknowledgments

Research described in this article was supported in part by Philip Morris USA, Inc. and Philip Morris International (SL, RZ and TLW) and by the NIH grant PO-1 DE12321 (TLW).

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Authors and Affiliations

  1. Research Pavilion at the Hillman Cancer Center, University of Pittsburgh Cancer Institute, 5117 Centre Avenue, Suite 1.27, Pittsburgh, PA, 15213-1863, USA

    Christoph Bergmann, Laura Strauss & Theresa L. Whiteside

  2. Department of Otorhinolaryngology, Ludwig-Maximilians-University of Munich, Marchioninistr. 15, 81377, Munich, Germany

    Reinhard Zeidler

  3. Department of Otorhinolaryngology, University of Duisburg-Essen, Hufelandstrasse 55, 45127, Essen, Germany

    Stephan Lang

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  1. Christoph Bergmann
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  5. Theresa L. Whiteside
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Correspondence to Theresa L. Whiteside.

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Bergmann, C., Strauss, L., Zeidler, R. et al. Expansion and characteristics of human T regulatory type 1 cells in co-cultures simulating tumor microenvironment. Cancer Immunol Immunother 56, 1429–1442 (2007). https://doi.org/10.1007/s00262-007-0280-9

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  • DOI: https://doi.org/10.1007/s00262-007-0280-9

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