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Combination immunotherapy with anti-CTLA-4 and interleukin-2 redirects regulatory T cells into tumor-draining lymph nodes and expands anti-tumor CD8+ T cells in the tumor microenvironment

  • Joseph Broucek1,
  • Tasha Hughes1,
  • Erica Huelsmann1,
  • Eugene Lusciks1,
  • Graham Hill1,
  • Janet Zayas1,
  • Joseph Poshepny1,
  • Carl Ruby1,
  • Frederick Kohlhapp1,
  • Andrew Zloza1 and
  • Howard Kaufman2
Journal for ImmunoTherapy of Cancer20142(Suppl 3):P97

https://doi.org/10.1186/2051-1426-2-S3-P97

Published: 6 November 2014

Keywords

MelanomaTumor MicroenvironmentIpilimumabTumor AreaPlacebo Treatment

Background

Monotherapy with Ipilimumab (anti-CTLA-4 antibody) and monotherapy with IL-2 (T cell stimulating cytokine) are approved for the treatment of metastatic melanoma. Combination immunotherapy has been suggested as a more potent regimen but has not been sufficiently investigated. We hypothesized that this combination may enhance therapeutic responses through alteration of the effector CD8+ T cell to CD4+FoxP3+ regulatory T cell ratio.

Methods

On day 0, C57BL/6 mice were challenged via intradermal injection with B16-F10 melanoma (120,000 cells). To track anti-tumor CD8+ T cell responses, pmel CD8+ T cells (specific against melanoma antigen gp100; 50,000 cells) were adoptively transferred via retroorbital injection. On days 3, 6, and 9 anti-CTLA-4 (100 µg in 100 µl or 100 µg IgG control) was administered via intraperitoneal injection. On days 4-8 IL-2 (100,000 units in 100 µl or 100µl PBS) was administered every 12 h. Tumor area was measured every 2-3 days until reaching 100 mm2 or until mice were sacrificed for flow cytometry analysis of T cell responses in the tumor and tumor-draining lymph nodes.

Results

Tumor growth was significantly reduced with combination anti-CTLA-4 and IL-2 treatment (average tumor area: 2mm2 on day 14) compared to anti-CTLA-4 only, IL-2 only, and placebo treatment (14, 29, and 68 mm2, respectively, on day 14) (p < 0.01 for all comparisons). On day 4, regulatory T cells were decreased by 20% in the tumor with anti-CTLA-4 therapy alone or in combination with IL-2, compared to placebo. By day 10, regulatory T cells decreased by 80% in the tumor, but increased three-fold in the tumor-draining lymph nodes, compared to placebo (p < 0.01 for all comparisons). In the tumor microenvironment, anti-tumor (pmel) CD8+ T cells were increased 2-fold with IL-2 therapy alone or in combination with anti-CTLA-4, compared to placebo (p < 0.01). This resulted in a pmel CD8+ T cell/Treg ratio of >10 with combination immunotherapy compared to ratios of 3, 1.3, and 0.7 for anti-CTLA-4 only, IL-2 only, and placebo treatments, respectively.

Conclusions

Combination immunotherapy with anti-CTLA-4 and IL-2 increases the tumor-specific CD8+ T cell/regulatory CD4+ T cell ratio in the tumor microenvironment and significantly decreases tumor growth (compared to monotherapy alone or placebo). These findings propose a previously unrecognized mechanism for anti-CTLA-4 in which regulatory CD4+ T cells are redirected out of the tumor microenvironment into the tumor-draining lymph nodes. These data highlight the potential synergistic action of anti-CTLA-4 combined with IL-2 for the treatment of melanoma.

Authors’ Affiliations

(1)
Rush University Medical Center, Chicago, USA
(2)
Rutgers Cancer Institute of New Jersey, New Brunswick, USA

Copyright

© Broucek et al.; licensee BioMed Central Ltd. 2014

This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.

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