The adenosine receptor (AR) subtypes A 2A and A 2B are rhodopsin-like G s protein-coupled receptors whose expression is highly regulated under pathological, eg hypoxic, ischemic and inflammatory conditions. Both receptors play important roles in inflammatory and neurodegenerative diseases, are blocked by caffeine, and have now become major drug targets in immuno-oncology. By Förster resonance energy transfer (FRET), bioluminescence resonance energy transfer (BRET), bimolecular fluorescence complementation (BiFC) and proximity ligation assays (PLA) we demonstrated A 2A-A 2B AR heteromeric complex formation. Moreover we observed a dramatically altered pharmacology of the A 2A AR when co-expressed with the A 2B AR (A 2B≥ A 2A) in recombinant as well as in native cells. In the presence of A 2B ARs, A 2A-selective ligands lost high affinity binding to A 2A ARs and displayed strongly reduced potency in cAMP accumulation and dynamic mass redistribution (DMR) assays. These results have major implications for the use of A 2A AR ligands as drugs as they will fail to modulate the receptor in an A 2A-A 2B heteromer context. Accordingly, A 2A-A 2B AR heteromers represent novel pharmacological targets.