GRPR inhibitors(Alpha-9 Theranostics)

The gastrin-releasing peptide receptor (GRPR) binds to ligands such as gastrin-releasing peptide (GRP) and plays a variety of biological roles. In this study, we investigated the therapeutic effect of a novel gastrin-releasing peptide receptor antagonist RH-1402 in hyperuricemia-induced kidney fibrosis and its underlying mechanisms. We conducted enzyme linked immunosorbent assay (ELISA) and immunohistochemical analyses and found that proGRP and GRPR expression levels were significantly increased in patients with hyperuricemic nephropathy (HN) and HN mice. GRPR knockdown significantly attenuated inflammatory and fibrotic responses in adenosine-treated human proximal tubule epithelial cells. GRPR knockout or GRPR conditional knockout in renal tubular epithelial cells significantly alleviated the decline in renal function and fibrosis in HN mice in vivo. RNA-seq and String database analysis revealed that GRP/GRPR promoted HN by suppressing the ABCG2/PDZK1 and increasing TGF-β/Smad3 levels by activating the NF-κB pathway. Overexpression of GRPR increased TGF-β/Smad3 levels, where as it reduced ABCG2/PDZK1 levels in adenosine-treated HK2 cells, which was reversed by the NF-κB inhibitor. Furthermore, we evaluated the therapeutic effects of the novel GRPR inhibitor RH-1402 on hyperuricaemia-induced renal injury and evaluated the inflammatory and fibrosis responses in vivo and in vitro. Pre-treatment with RH-1402 attenuated hyperuricaemia-induced renal injury, restored renal function, and suppressed renal inflammation and fibrosis. Taken together, GRPR enhances hyperuricaemia-induced tubular injury, inflammation, and renal fibrosis via ABCG2-dependent mechanisms and may serve as a promising therapeutic target for HN treatment.

GRPR is over-expressed in cancer cells and is a potential drug target for the treatment of cancer. PD176252, as the most representative non-peptide inhibitor of GRPR, can inhibit the growth of cancer cells, but its low selectivity to cancer cells and normal cells limits its further application.

The aim of this study was to design and synthesize novel GRPR inhibitor with stronger anti-cancer activity and higher affinity with GRPR than the lead compound PD176252.

A series of 1, 3, 4-oxadiazole derivatives as PD176252 analogues (4a-4j, 6a-6q) were synthesized and their cytotoxic activity was investigated on four cancer lines with high expression of GRPR (gastric (HGC-27), colon (HCT- 116), prostate (PC-3), and lung (A549)) and one human cell line (gastric mucosal epithelial (GES-1)) by MTT assay. Flow cytometry analysis and Western Blot were used to determine whether the compound induced programmed apoptosis of cancer cells. Competitive binding experiment was used to verify the affinity between GRPR and the optimal compound.

Compound 6m exhibited significant growth inhibition on all tested cancer cell lines, especially gastric cancer cells (HGC-27 cellular IC50 0.37 ± 0.04μM). Also, the selectivity of 6m to HGC-27 was much higher than that of PD176252. Flow cytometric analysis and Western Blot proved that 6m significantly promoted the apoptosis of HGC- 27 cells. Moreover, competitive binding experiment confirmed the close binding of 6m with GRPR, which indicated 6m with a higher affinity than lead compound PD176252.

Our results suggested that 6m, as a novel GRPR inhibitor, had a higher affinity with GRPR and potential anti-cancer effect than PD176252, which can be used as a template for further optimization.