Breast Cancer Cell-Derived Adenosine Enhances Generation and Suppressor Function of Human Adaptive Regulatory T Cells.

Magis Mandapathil,Stephan Lang,Miroslaw J Szczepanski,Theresa L Whiteside,Edwin K Jackson

doi:10.3390/jpm11080754

Abstract

Introduction: Adaptive regulatory T cells (Tr1) are induced in the periphery by environmental stimuli. CD73 expression and adenosine (ADO) production by tumor cells may influence Tr1 generation and their immunosuppressive activity. Material and Methods: Tr1 were generated in co-cultures of CD4+CD25neg T cells, autologous immature dendritic cells (iDC), and irradiated ADO-producing CD73+ or non-producing CD73neg breast cancer (BrCa) cell lines (TU). The expression of ectonucleotidases and other surface markers on Tr1 was determined by flow cytometry. Tr1-mediated suppression of proliferation was evaluated in CFSE-based assays. Luciferase-based ATP detection assays and mass spectrometry were used to measure ATP hydrolysis and ADO levels. Cytokine levels were measured by ELISA or Luminex. CD73 expression on tumor cells or T cells in TU tissues was assessed by immunofluorescence. Results: CD73+ TU induced higher numbers of Tr1 cells (p < 0.01) than CD73neg TU. Tr1TU73+ hydrolyzed more exogenous ATP, produced more ADO, and mediated higher suppression than Tr1TU73neg (p < 0.05 for all). ARL67156, an ectonucleotidase inhibitor, and ZM241385, A2A receptor antagonist, reduced suppression of proliferation mediated by Tr1TU73+ cells (p < 0.01). Basal-like primary BrCa cells expressed higher levels of ectonucleotidases and induced more Tr1 than less aggressive primary luminal-like BrCa. Conclusion: BrCa producing ADO (CD73+ TU) favor the induction of Tr1, which expresses CD39 and CD73, hydrolyzes ATP to ADO, and effectively suppresses anti-tumor immunity.

Highlights

Introduction iationsHuman tumors develop a variety of mechanisms that down-regulate anti-tumor responses promoting tumor escape from the host immune system [1]
The expression of CD73 and CD39 on the surface of seven breast cancer (BrCa) cell lines was evaluated by flow cytometry
MDA-MB231 (98% CD73+ ) and MCF-7 (CD73neg ) cell lines were selected for further studies as the representative BrCa cells for surface expression of CD73 or its absence, respectively

Summary

Introduction

Human tumors develop a variety of mechanisms that down-regulate anti-tumor responses promoting tumor escape from the host immune system [1]. Similar to other solid tumors, breast cancers (BrCa) create a tumor microenvironment enriched in various immunosuppressive factors such as adenosine (ADO), which favor tumor growth by reducing anti-tumor immune responses [2]. ADO, a purine nucleoside, may be present at high concentrations in tumor tissues and body fluids [3]. ADO extracellular concentrations rise dramatically in response to tissue damage, hypoxia, or increased ATP levels, conditions commonly present in solid tumors [4]. Extracellular ADO is a product of ATP hydrolysis by ectonucleotidases, CD39, an ectonucleotidase triphosphate diphosphohydrolase (NTPDase), and CD73, a 50 ectonucleotidase. Ectonucleotidases are expressed by different tumors and presumably protect tumor cells from ATP-mediated toxicity [5].

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Conclusion