Modulation of PAR1 signalling by benzimidazole compounds

作者：Asteriti S; Daniele S; Porchia F; Dell' Anno M T; Fazzini A; Pugliesi I; Trincavelli M L; Taliani S; Martini C; Mazzoni M R; Gilchrist A^*

来源：British Journal of Pharmacology, 2012, 167(1): 80-94.

DOI：10.1111/j.1476-5381.2012.01974.x

• allosteric
• benzimidazole scaffold
• biased signalling
• endothelial cells
• functional selectivity
• Ga
• GPCR
• HMEC-1
• PAR1
• positive
• negative modulation
• protease-activated receptor
• Q94
• thrombin
• thrombin receptor

BACKGROUND AND PURPOSE Recently, a small molecule (Q94) was reported to selectively block PAR1/Gaq interaction and signalling. Here, we describe the pharmacological properties of Q94 and two analogues that share its benzimidazole scaffold (Q109, Q89). Q109 presents a modest variation from Q94 in the substituent group at the 2-position, while Q89 has quite different groups at the 1- and 2-positions. EXPERIMENTAL APPROACH Using human microvascular endothelial cells, we examined intracellular Ca2+ mobilization and inositol 1,4,5-trisphosphate accumulation as well as isoprenaline- or forskolin-stimulated cAMP production in response to thrombin. KEY RESULTS Q89 (10 mu M) produced a leftward shift in the thrombin-mediated intracellular Ca2+ mobilization concentrationresponse curve while having no effect on the Emax. Both Q94 (10 mu M) and Q109 (10 mu M) reduced intracellular Ca2+ mobilization, leading to a decrease in Emax and an increase in EC50 values. Experiments utilizing receptor-specific activating peptides confirmed that Q94 and Q109 were selective for PAR1 as they did not alter the Ca2+ response mediated by a PAR2 activating peptide. Consistent with our Ca2+ results, micromolar concentrations of either Q94 or Q109 significantly reduced thrombin-induced inositol 1,4,5-trisphosphate production. Neither Q94 nor Q109 diminished the inhibitory effects of thrombin on cAMP production, indicating they inhibit signalling selectively through the Gq pathway. Our results also suggest the 1,2-disubstituted benzimidazole derivatives act as allosteric agonists of PAR1. CONCLUSIONS AND IMPLICATIONS The Q94 and Q109 benzimidazole derivatives represent a novel scaffold for the development of new PAR1 inhibitors and provide a starting point to develop dual signalling pathway-selective positive/negative modulators of PAR1.

• 出版日期2012-9