Discovery of oridonin as a novel agonist for BRS-3.

Background: Bombesin Receptor Subtype-3 (BRS-3, Bombesin-like receptor, BB) is an orphan G-protein coupled receptor (GPCR). Recent studies have shown that BRS-3 played a vital role in glucose regulation, insulin secretion, and energy homeostasis. Therefore, discovering more novel exogenous ligands with diverse structures for BRS-3 will be of great importance for target validation and drug development.

Purpose: In this study, we aim to discover new agonists of BRS-3 from our natural compound libraries, providing a new probe to study the function of BRS-3.

Study Design: Multiple cell-based assays and in vivo experiments were performed to identify the new ligand.

Methods: BRS-3 overexpression cells were coupled with FLIPR assay, homogeneous time-resolved fluorescence (HTRF) IP-ONE assay, dynamic mass redistribution (DMR) assay, β-arrestin2 recruitment assay, and western blot to determine receptor activation and downstream signaling events. To further validate the target of BRS-3, a series of in vitro and in vivo experiences were conducted, including glucose uptake, glucose transporter type 4 (GLUT4) transportation in C2C12, and oral glucose tolerance test (OGTT) in mice.

Results: We discovered and identified oridonin as a novel small molecule agonist of BRS-3, with a moderate affinity (EC of 2.236 × 10 M in calcium mobilization assay), specificity, and subtype selectivity. Further in vitro and in vivo tests demonstrated that oridonin exerted beneficial effects in glucose homeostasis through activating BRS-3.

Conclusions: Oridonin, as the discovered new ligand of BRS-3, provides a valuable tool compound to investigate BRS-3's function, especially for target validation in type 2 diabetes and obesity. Oridonin is promising as a lead compound in the treatment of metabolic disorders. Compared to the known agonists of BRS-3, we can take advantage of the multiple reported pharmacological activities of ODN as a natural product and assess whether these pharmacological activities are regulated by BRS-3. This may facilitate the discovery of novel functions of BRS-3.