A CCR4 antagonist reverses the tumor-promoting microenvironment of renal cancer
Chiara Berlato, Moddasar N. Khan, Tiziana Schioppa, Richard Thompson, Eleni Maniati, Anne Montfort, Maryam Jangani, Monica Canosa, Hagen Kulbe, Urs B. Hagemann, Alexander R. Duncan, Laura Fletcher, Robert W. Wilkinson, Thomas Powles, Sergio A. Quezada, Frances R. Balkwill
Chiara Berlato, Moddasar N. Khan, Tiziana Schioppa, Richard Thompson, Eleni Maniati, Anne Montfort, Maryam Jangani, Monica Canosa, Hagen Kulbe, Urs B. Hagemann, Alexander R. Duncan, Laura Fletcher, Robert W. Wilkinson, Thomas Powles, Sergio A. Quezada, Frances R. Balkwill
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Research Article Immunology Oncology

A CCR4 antagonist reverses the tumor-promoting microenvironment of renal cancer

Abstract

Elevated expression of the chemokine receptor CCR4 in tumors is associated with poor prognosis in several cancers. Here, we have determined that CCR4 was highly expressed in human renal cell carcinoma (RCC) biopsies and observed abnormal levels of CCR4 ligands in RCC patient plasma. An antagonistic anti-CCR4 antibody had antitumor activity in the RENCA mouse model of RCC. CCR4 inhibition did not reduce the proportion of infiltrating leukocytes in the tumor microenvironment but altered the phenotype of myeloid cells, increased NK cell and Th1 cytokine levels, and reduced immature myeloid cell infiltrate and blood chemokine levels. In spite of prominent changes in the myeloid compartment, the anti-CCR4 antibody did not affect RENCA tumors in T cell–deficient mice, and treatment with an anti–class II MHC antibody abrogated its antitumor activity. We concluded that the effects of the anti-CCR4 antibody required the adaptive immune system and CD4+ T cells. Moreover, CCL17-induced IFN-γ production was reduced when Th1-polarized normal CD4+ T cells were exposed to the CCR4 ligand, evidencing the involvement of CCR4 in Th1/Th2 regulation. The anti-CCR4 antibody, alone or in combination with other immune modulators, is a potential treatment approach to human solid cancers with high levels of CCR4-expressing tumor-infiltrating leukocytes and abnormal plasma CCR4 ligand levels.

Authors

Chiara Berlato, Moddasar N. Khan, Tiziana Schioppa, Richard Thompson, Eleni Maniati, Anne Montfort, Maryam Jangani, Monica Canosa, Hagen Kulbe, Urs B. Hagemann, Alexander R. Duncan, Laura Fletcher, Robert W. Wilkinson, Thomas Powles, Sergio A. Quezada, Frances R. Balkwill

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Figure 5

Other effects of anti-CCR4 antibody in the RENCA tumor model.

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Other effects of anti-CCR4 antibody in the RENCA tumor model. BALB/c mic...
BALB/c mice were injected with RENCA-luc cells and treated with Affi-5 (T) or isotype control (C). Mice were sacrificed 17 days after surgery, and tumors or spleens were dissociated and characterized by flow cytometry. (A) Percentage of NK cells (CD45+CD3–DX5+) among the CD45+ population in tumors; 2-tailed Student’s t test, **P = 0.0064, 4 independent experiments pooled together (n = 12 for C, n = 15 for T). (B) The number of NK cells/mg of tumor was also significantly higher with treatment (2-tailed Student’s t test, *P = 0.032, n = 12 for C, n = 15 for T). (C) Percentage of MDSCs (CD45+CD11+Gr1+) among the CD45+ infiltrate for tumors from control and treated animals. Two populations of MDSCs (Gr1hi and Gr1int) were identified and analyzed separately. Two-tailed Student’s t test, **P = 0.0021 and ***P = 0.0065; 2 experiments pooled together, n = 7 for C and n = 6 for T. (D) The fold change in the number of MDSCs/mg of tumor was also significantly lower in the tumors from treated animals (4 experiments, 2-tailed Student’s t test, *P = 0.017, n = 12 for C, n = 11 for T). (E) Percentage of MDSCs (CD45+CD11b+Gr1+) among the CD45+ infiltrate for spleens from control and treated animals. Two populations of MDSCs (Gr1hiLy6Cint and Ly6ChiGr1int) were identified and analyzed separately. Two-tailed Student’s t test, P = 0.075 for Gr1hi, *P = 0.018 for Ly6Chi, n = 3. (F) Naive CD3+ cells isolated from spleen of healthy mice (5 × 104/well) were pre-labeled with CFSE and activated with anti-CD3– and anti-CD28–coated beads at a ratio 1:2 beads/CD3 cells in the presence of freshly isolated MDSCs from tumors. Cells were cocultured for 3 days, and CD4+ and CD8+ T cell proliferation was measured by CFSE dye dilution from 2 independent experiments, each pooling MDSCs from 2–3 tumors. Proliferation was inhibited significantly (1-way ANOVA, P = 0.0012 for CD4, P = 0.0058 for CD8, at MDSC/T cell ratios of 10:1, 1:1, 1:2).