Inhibition of the adenosine A2a receptor modulates expression of T cell coinhibitory receptors and improves effector function for enhanced checkpoint blockade and ACT in murine cancer models

Abstract

Adenosine signaling via the A2a receptor (A2aR) is emerging as an important checkpoint of immune responses. The presence of adenosine in the inflammatory milieu or generated by the CD39/CD73 axis on tissues or T regulatory cells serves to regulate immune responses. By nature of the specialized metabolism of cancer cells, adenosine levels are increased in the tumor microenvironment and contribute to tumor immune evasion. To this end, small molecule inhibitors of the A2aR are being pursued clinically to enhance immunotherapy. Herein, we demonstrate the ability of the novel A2aR antagonist, CPI-444, to dramatically enhance immunologic responses in models of checkpoint therapy and ACT in cancer. Furthermore, we demonstrate that A2aR blockade with CPI-444 decreases expression of multiple checkpoint pathways, including PD-1 and LAG-3, on both CD8+ effector T cells (Teff) and FoxP3+ CD4+ regulatory T cells (Tregs). Interestingly, our studies demonstrate that A2aR blockade likely has its most profound effects during Teff cell activation, significantly decreasing PD-1 and LAG-3 expression at the draining lymph nodes of tumor bearing mice. In contrast to previous reports using A2aR knockout models, pharmacologic blockade with CPI-444 did not impede CD8 T cell persistence or memory recall. Overall these findings not only redefine our understanding of the mechanisms by which adenosine inhibits immunity but also have important implications for the design of novel immunotherapy regimens.

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Abbreviations

A2aR:

Adenosine A2a receptor

ARG1:

Arginase1

ATCC:

American type culture collection

B16-OVA:

OVA-expressing B16 murine melanoma

dLN:

Tumor-draining lymph nodes

iNOS:

Inducible nitric oxide synthase

LM-OVA:

OVA-expressing Listeria monocytogenes

ndLN:

Non-draining lymph nodes

n.s.:

Not significant

r.o.:

Retro-orbital

Teff:

CD8+ effector T cell

Tet-OVA+:

OVA class-I tetramer+

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Acknowledgements

We thank members of the Powell lab, especially Chirag Patel, for critical discussion of the manuscript; Corvus pharmaceuticals for their generous gift of CPI-444; and Aduro Biotech for their generous gift of LM-OVA.

Funding

This work was supported in part by funds from the Bloomberg~Kimmel Institute for Cancer Immunotherapy. In addition, CPI-444 and unrestricted research funds were provided by Corvus.

Author information

Affiliations

Authors

Contributions

RDL designed and conducted the experiments and wrote the manuscript. I-MS, M-HO, I-HS, JW, and JE helped with the experiments. JDP designed the experiments and revised the manuscript.

Corresponding author

Correspondence to Jonathan D. Powell.

Ethics declarations

Conflict of interest

Jonathan D. Powell has been a paid consultant for Corvus and has equity in the company. All other authors declare that they have no conflicts of interest.

Ethical approval and ethical standards

All applicable international and national guidelines for the care of animals were followed. All mouse procedures approved by Johns Hopkins University Institutional Animal Care and Use Committee (Protocol #M016M103, approved 4/1/2016).

Animal source

C57BL/6 obtained from Charles River Laboratories (MC38 experiments) or Jackson Laboratories (ACT; 000664). OT-I and CD90.1, BALB/c mice obtained from The Jackson Laboratory. Male or female mice were used for each experiment; mice were sex and age matched accordingly.

Cell line authentication

MC38 cells were donated by CORVUS pharmaceuticals. The identity and specific pathogen free status of these cells was validated by microsatellite genotype analysis (IDEXX Bioresearch). B16-OVA melanoma cells were a gift from Hyam Levitsky. All other tumor cell lines used were obtained from the ATCC. All cell lines were mycoplasma free via ELISA-based assays performed every 6 months.

Additional information

Some of the data contained herein had been presented as an abstract and oral presentation. Proceedings of the 107th Annual Meeting of the American Association for Cancer Research (AACR); 2016 April 16–20; New Orleans, LA. Cancer Res 2016; 76: (Abstract 4364).

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Cite this article

Leone, R.D., Sun, IM., Oh, MH. et al. Inhibition of the adenosine A2a receptor modulates expression of T cell coinhibitory receptors and improves effector function for enhanced checkpoint blockade and ACT in murine cancer models. Cancer Immunol Immunother 67, 1271–1284 (2018). https://doi.org/10.1007/s00262-018-2186-0

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Keywords

  • Immunotherapy
  • Immune checkpoint
  • A2a
  • PD-1
  • Lag-3
  • Treg