G protein–coupled receptors display multifunctional signaling, offering the potential for agonist structures to promote conformational selectivity for biased outputs. For β₂-adrenergic receptors (β₂AR), unbiased agonists stabilize conformation(s) that evoke coupling to Gαs (cyclic adenosine monophosphate [cAMP] production/human airway smooth muscle [HASM] cell relaxation) and β-arrestin engagement, the latter acting to quench Gαs signaling, contributing to receptor desensitization/tachyphylaxis. We screened a 40-million-compound scaffold ranking library, revealing unanticipated agonists with dihydroimidazolyl-butyl-cyclic urea scaffolds. The S-stereoisomer of compound C1 shows no detectable β-arrestin engagement/signaling by four methods. However, C1-S retained Gαs signaling—a divergence of the outputs favorable for treating asthma. Functional studies with two models confirmed the biasing: β₂AR-mediated cAMP signaling underwent desensitization to the unbiased agonist albuterol but not to C1-S, and desensitization of HASM cell relaxation was observed with albuterol but not with C1-S. These HASM results indicate biologically pertinent biasing of C1-S, in the context of the relevant physiologic response, in the human cell type of interest. Thus, C1-S was apparently strongly biased away from β-arrestin, in contrast to albuterol and C5-S. C1-S structural modeling and simulations revealed binding differences compared with unbiased epinephrine at transmembrane (TM) segments 3,5,6,7 and ECL2. C1-S (R2 = cyclohexane) was repositioned in the pocket such that it lost a TM6 interaction and gained a TM7 interaction compared with the analogous unbiased C5-S (R2 = benzene group), which appears to contribute to C1-S biasing away from β-arrestin. Thus, an agnostic large chemical-space library identified agonists with receptor interactions that resulted in relevant signal splitting of β₂AR actions favorable for treating obstructive lung disease.

G 蛋白偶联受体显示出多功能信号传导，为激动剂结构提供了促进偏倚输出的构象选择性的潜力。对于 β ₂ -肾上腺素能受体 (β ₂ AR)，无偏激动剂可稳定引起与 Gα 偶联（环磷酸腺苷 [cAMP] 产生/人气道平滑肌 [HASM] 细胞松弛）和 β-抑制蛋白结合的构象，后者起到淬灭 Gαs 信号传导的作用，有助于受体脱敏/快速反应。我们筛选了一个包含 4000 万个化合物的支架排名库，揭示了具有二氢咪唑基-丁基-环脲支架的意外激动剂。通过四种方法，化合物 C1的S-立体异构体未显示出可检测到的 β-抑制蛋白接合/信号传导。然而，C1-S保留了 Gαs 信号——有利于治疗哮喘的输出差异。使用两种模型进行的功能研究证实了这种偏差：β2 _AR介导的 cAMP 信号对无偏激动剂沙丁胺醇而不是 C1 -S脱敏，并且在沙丁胺醇中观察到 HASM 细胞松弛的脱敏，但在 C1 -S中没有。这些 HASM 结果表明，在相关生理反应的背景下，在感兴趣的人类细胞类型中， C1 -S的生物学相关偏差。因此，与沙丁胺醇和 C5 - S相比，C1 -S明显远离 β-抑制素。C1 -S结构建模和模拟揭示了与跨膜 (TM) 片段 3、5、6、7 和 ECL2 的无偏肾上腺素相比的结合差异。与类似的无偏 C5 -S （R2 = 苯基）相比， C1 -S（R2 = 环己烷）在口袋中重新定位，使其失去 TM6 相互作用并获得 TM7 相互作用，这似乎有助于 C1 -S偏离来自β-抑制素。因此，一个不可知论的大型化学空间库鉴定了具有受体相互作用的激动剂，这些受体相互作用导致 β ₂ AR 作用的相关信号分裂，有利于治疗阻塞性肺病。