The paradigm of one drug against multiple targets, known as unimolecular polypharmacology, offers the potential to improve efficacy while overcoming some adverse events associated with the treatment. This approach is best exemplified by targeting two or three class B1 G protein-coupled receptors, namely, glucagon-like peptide-1 receptor (GLP-1R), glucagon receptor (GCGR) and glucose-dependent insulinotropic polypeptide receptor for treatment of type 2 diabetes and obesity. Some of the dual and triple agonists have already shown initial successes in clinical trials, although the molecular mechanisms underlying their multiplexed pharmacology remain elusive. In this study we employed structure-based site-directed mutagenesis together with pharmacological assays to compare agonist efficacy across two key signaling pathways, cAMP accumulation and ERK1/2 phosphorylation (pERK1/2). Three dual agonists (peptide 15, MEDI0382 and SAR425899) and one triple agonist (peptide 20) were evaluated at GLP-1R and GCGR, relative to the native peptidic ligands (GLP-1 and glucagon). Our results reveal the existence of residue networks crucial for unimolecular agonist-mediated receptor activation and their distinct signaling patterns, which might be useful to the rational design of biased drug leads.

一种药物针对多个靶标的范例，称为单分子多药理学，提供了提高疗效同时克服与治疗相关的一些不良事件的潜力。这种方法最好的例子是靶向两种或三种 B1 G 蛋白偶联受体，即胰高血糖素样肽-1 受体 (GLP-1R)、胰高血糖素受体 (GCGR) 和葡萄糖依赖性促胰岛素多肽受体，用于治疗 2 型糖尿病和肥胖症。一些双重和三重激动剂已经在临床试验中取得初步成功，尽管其多重药理学背后的分子机制仍然难以捉摸。在这项研究中，我们采用基于结构的定点诱变和药理学分析来比较激动剂在两个关键信号通路中的功效，cAMP 积累和 ERK1/2 磷酸化 (pERK1/2)。相对于天然肽配体（GLP-1 和胰高血糖素），在 GLP-1R 和 GCGR 评估了三种双重激动剂（肽 15、MEDI0382 和 SAR425899）和一种三重激动剂（肽 20）。我们的结果揭示了对单分子激动剂介导的受体激活至关重要的残基网络的存在及其独特的信号模式，这可能有助于偏向药物先导物的合理设计。