Preventing ATP Degradation by ASO-Mediated Knockdown of CD39 and CD73 Results in A2aR-Independent Rescue of T Cell Proliferation

Julia Festag,Tamara Thelemann,Monika Schell,Stefanie Raith,Sven Michel,Frank Jaschinski,Richard Klar

doi:10.1016/j.omtn.2020.06.020

Julia Festag, Tamara Thelemann + Show 5 more

Open Access

https://doi.org/10.1016/j.omtn.2020.06.020

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Abstract

The adenosine axis contributes to the suppression of antitumor immune responses. The ectonucleotidase CD39 degrades extracellular adenosine triphosphate (ATP) to adenosine monophosphate (AMP), which is degraded to adenosine by CD73. Adenosine binds to, e.g., the A2a receptor (A2aR), which reportedly suppresses effector immune cells. We investigated effects of ATP, AMP, and adenosine analogs on T cell proliferation, apoptosis, and proinflammatory cytokine secretion. CD39 and CD73 expression were suppressed using antisense oligonucleotides (ASOs), and A2aR was blocked using small molecules. Addition of ATP to T cells reduced proliferation and induced apoptosis. Intriguingly, those effects were reverted by suppression of CD39 and/or CD73 expression but not A2aR inhibition. Adenosine analogs did not suppress proliferation but inhibited secretion of proinflammatory cytokines. Here, we suggest that suppression of T cell proliferation is not directly mediated by A2aR but by intracellular downstream metabolites of adenosine, as blockade of the equilibrative nucleoside transporter (ENT) or adenosine kinase rescued proliferation and prevented induction of apoptosis. In conclusion, adenosine might primarily affect cytokine secretion directly via adenosine receptors, whereas adenosine metabolites might impair T cell proliferation and induce apoptosis. Therefore, inhibition of CD39 and/or CD73 has evident advantages over A2aR blockade to fully revert suppression of antitumor immune responses by the adenosine axis.

Highlights

Tumors can escape immune recognition and destruction by a plethora of immunosuppressive mechanisms
We target the expression of the ectonucleotidases CD39 and CD73 by antisense oligonucleotides (ASOs) and demonstrate that T cells treated with CD39- or CD73-specific ASOs are protected from the inhibition of proliferation induced by adenosine triphosphate (ATP) degradation products
We observed that ASO-mediated knockdown of CD39 and/or CD73 rescued T cells from ATPinduced suppression of proliferation up to a concentration of 400 mM ATP, which is a relevant concentration measured at tumor sites as a source of immunosuppressive adenosine.[36]

Summary

Introduction

Tumors can escape immune recognition and destruction by a plethora of immunosuppressive mechanisms. The unleashing of tumor-specific immune responses by blocking immunosuppressive pathways has emerged as a promising treatment option in the last years. Despite the remarkable success of blocking immune checkpoints, like programmed death 1 (PD-1), using monoclonal antibodies, only a minority of patients benefit from the currently available immunotherapies.[1] The adenosine axis has emerged as a promising therapeutic target to enhance antitumor immunity. Degradation of extracellular immune-stimulating adenosine triphosphate (ATP) by the ectonucleotidase CD39 to adenosine monophosphate (AMP) is followed by generation of immunosuppressive adenosine by CD73. Adenosine binds to adenosine receptors, e.g., the A2a receptor (A2aR) or with lower affinity to the A2b receptor (A2bR), which increases intracellular cyclic AMP (cAMP) levels, resulting in suppression of immune cell function.[2]

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