Eya3 promotes breast tumor–associated immune suppression via threonine phosphatase–mediated PD-L1 upregulation
Rebecca L. Vartuli, Hengbo Zhou, Lingdi Zhang, Rani K. Powers, Jared Klarquist, Pratyaydipta Rudra, Melanie Y. Vincent, Debashis Ghosh, James C. Costello, Ross M. Kedl, Jill E. Slansky, Rui Zhao, Heide L. Ford
Rebecca L. Vartuli, Hengbo Zhou, Lingdi Zhang, Rani K. Powers, Jared Klarquist, Pratyaydipta Rudra, Melanie Y. Vincent, Debashis Ghosh, James C. Costello, Ross M. Kedl, Jill E. Slansky, Rui Zhao, Heide L. Ford
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Research Article Immunology Oncology

Eya3 promotes breast tumor–associated immune suppression via threonine phosphatase–mediated PD-L1 upregulation

Abstract

Eya proteins are critical developmental regulators that are highly expressed in embryogenesis but downregulated after development. Amplification and/or re-expression of Eyas occurs in many tumor types. In breast cancer, Eyas regulate tumor progression by acting as transcriptional cofactors and tyrosine phosphatases. Intriguingly, Eyas harbor a separate threonine (Thr) phosphatase activity, which was previously implicated in innate immunity. Here we describe what we believe to be a novel role for Eya3 in mediating triple-negative breast cancer–associated immune suppression. Eya3 loss decreases tumor growth in immune-competent mice and is associated with increased numbers of infiltrated CD8+ T cells, which, when depleted, reverse the effects of Eya3 knockdown. Mechanistically, Eya3 utilizes its Thr phosphatase activity to dephosphorylate Myc at pT58, resulting in a stabilized form. We show that Myc is required for Eya3-mediated increases in PD-L1, and that rescue of PD-L1 in Eya3-knockdown cells restores tumor progression. Finally, we demonstrate that Eya3 significantly correlates with PD-L1 in human breast tumors, and that tumors expressing high levels of Eya3 have a decreased CD8+ T cell signature. Our data uncover a role for Eya3 in mediating tumor-associated immune suppression, and suggest that its inhibition may enhance checkpoint therapies.

Authors

Rebecca L. Vartuli, Hengbo Zhou, Lingdi Zhang, Rani K. Powers, Jared Klarquist, Pratyaydipta Rudra, Melanie Y. Vincent, Debashis Ghosh, James C. Costello, Ross M. Kedl, Jill E. Slansky, Rui Zhao, Heide L. Ford

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Figure 5

Eya3 regulation of CD8+ T cells is required for increased 66cl4 mammary carcinoma growth.

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Eya3 regulation of CD8+ T cells is required for increased 66cl4 mammary ...
(A) Percentage of CD8+ cells in 50 μl blood from animals treated with rat IgG or CD8α-depleting antibody as measured by flow cytometry. Blood was isolated from 10 mice per group. Data represent mean ± SEM. Significance was measured using a 2-tailed Student’s t test. (B) Tumor volume of 66cl4-SCR and Eya3-KD tumors treated with rat IgG or CD8+-depleting antibody, as measured using calipers. Each point represents mean of tumors from mice in that condition ± SEM, and a mixed effects model was used to measure significance. n = 5 mice per group. Solid lines and filled symbols represent mice treated with IgG antibody. Dotted lines and open symbols represent mice treated with anti-CD8+ antibody. (C) Fold change of tumor growth between IgG-treated 66cl4-SCR and SCR2 tumors and CD8+-treated 66cl4-SCR and SCR2 tumors and fold change of tumor growth between IgG-treated 66cl4-Eya3 KD2 and KD3 tumors and CD8+-treated 66cl4-Eya3 KD2 and KD3 tumors. Data represent mean ± SEM. Significance was measured using a 2-tailed Student’s t test of average fold change of CD8+/IgG tumor size averaged for every time point over the experimental time course. n = 5 mice per group. *P < 0.05, ***P < 0.001.