Among class A G protein-coupled receptors (GPCR), the human adenosine A2A receptor (hA2AAR) remains an attractive drug target. However, translation of A2AAR ligands into the clinic has proved challenging and an improved understanding of A2AAR pharmacology could promote development of more efficacious therapies. Subtype-selective fluorescent probes would allow detailed real-time pharmacological investigations both in vitro and in vivo. In the present study, two families of fluorescent probes were designed around the known hA2AAR selective antagonist preladenant (SCH 420814). Both families of fluorescent antagonists retained affinity at the hA2AAR, selectivity over all other adenosine receptor subtypes and allowed clear visualization of specific receptor localization through confocal imaging. Furthermore, the Alexa Fluor 647-labeled conjugate allowed measurement of ligand binding affinities of unlabeled hA2AAR antagonists using a bioluminescence resonance energy transfer (NanoBRET) assay. The fluorescent ligands developed here can therefore be applied to a range of fluorescence-based techniques to further interrogate hA2AAR pharmacology and signaling.

中文翻译：

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亚型选择性荧光配体作为人类腺苷A2A受体的药理研究工具。

在AG类蛋白偶联受体（GPCR）中，人腺苷A2A受体（hA2AAR）仍然是有吸引力的药物靶标。然而，已证明将A2AAR配体翻译到临床中具有挑战性，并且对A2AAR药理学的更好理解可以促进更有效疗法的发展。亚型选择性荧光探针将允许在体内和体外进行详细的实时药理研究。在本研究中，围绕已知的hA2AAR选择性拮抗剂表面活性剂（SCH 420814）设计了两个荧光探针家族。荧光拮抗剂的两个家族都保留了对hA2AAR的亲和力，对所有其他腺苷受体亚型的选择性，并允许通过共聚焦成像清楚地观察特定受体的定位。此外，Alexa Fluor 647标记的缀合物允许使用生物发光共振能量转移（NanoBRET）分析方法测量未标记的hA2AAR拮抗剂的配体结合亲和力。因此，此处开发的荧光配体可应用于多种基于荧光的技术，以进一步审视hA2AAR药理学和信号传导。