Crystal structure of adenosine A_2A receptor in complex with clinical candidate Etrumadenant reveals unprecedented...

The G_s protein-coupled adenosine A_2A receptor (A_2AAR) represents an emerging drug target for cancer immunotherapy. The clinical candidate Etrumadenant was developed as an A_2AAR antagonist with ancillary blockade of the A_2BAR subtype. It constitutes a unique chemotype featuring a poly-substituted 2-amino-4-phenyl-6-triazolylpyrimidine core structure. Herein, we report two crystal structures of the A_2AAR in complex with Etrumadenant, obtained with differently thermostabilized A_2AAR constructs. This led to the discovery of an unprecedented interaction, a hydrogen bond of T88^3.36 with the cyano group of Etrumadenant. T88^3.36 is mutated in most A_2AAR constructs used for crystallization, which has prevented the discovery of its interactions. In-vitro characterization of Etrumadenant indicated low selectivity versus the A₁AR subtype, which can be rationalized by the structural data. These results will facilitate the future design of AR antagonists with desired selectivity. Moreover, they highlight the advantages of the employed A_2AAR crystallization construct that is devoid of ligand binding site mutations.