The PAR2 inhibitor I-287 selectively targets Gαq and Gα12/13 signaling and has anti-inflammatory effects

Abstract

Protease-activated receptor-2 (PAR2) is involved in inflammatory responses and pain, therefore representing a promising therapeutic target for the treatment of immune-mediated inflammatory diseases. However, as for other GPCRs, PAR2 can activate multiple signaling pathways and those involved in inflammatory responses remain poorly defined. Here, we describe a new selective and potent PAR2 inhibitor (I-287) that shows functional selectivity by acting as a negative allosteric regulator on Gαq and Gα12/13 activity and their downstream effectors, while having no effect on Gi/o signaling and βarrestin2 engagement. Such selective inhibition of only a subset of the pathways engaged by PAR2 was found to be sufficient to block inflammation in vivo. In addition to unraveling the PAR2 signaling pathways involved in the pro-inflammatory response, our study opens the path toward the development of new functionally selective drugs with reduced liabilities that could arise from blocking all the signaling activities controlled by the receptor.

Effector membrane translocation biosensors reveal G protein and βarrestin coupling profiles of 100 therapeutically relevant GPCRs

“…Gα s -67-RlucII (Carr et al, 2014), Gα i1 -loop-RlucII and GFP10-Gγ 1 (Armando et al, 2014), Gα i2 -loop-RlucII and βarrestin2-RlucII (Quoyer et al, 2013), Gα oB -99-RlucII (Mende et al, 2018), Gα q -118-RlucII (Breton et al, 2010), Gα 12 -136-RlucII and PKN-RBD-RlucII (Namkung et al, 2018), Gα 13 -130-RlucII (Avet et al, 2020), GFP10-Gγ 2 (Gales et al, 2006), βarrestin1- RlucII (Zimmerman et al, 2012), rGFP-CAAX (Namkung et al, 2016), EPAC (Leduc et al, 2009), MyrPB-Ezrin-RlucII (Leguay et al, 2021), HA-β 2 AR (Lavoie et al, 2002), signal peptide-Flag-AT 1 (Goupil et al, 2015) and EAAC-1 (Brabet et al, 1998) were previously described. Full-length, untagged Gα subunits, Gβ 1 and Gγ 9 were purchased from cDNA Resource Center.…”

Molecular mechanisms regulating Proteinase‐Activated Receptors (PARs)

“…I‐191 was also found to inhibit biased PAR2 antagonist GB88‐induced RhoA activation, ERK1/2 phosphorylation, and inhibition of cAMP accumulation [206,207]. A related compound I‐287, an allosteric modulator, inhibited Gα q and Gα 12/13 , but not Gα i or β‐arrestin recruitment to PAR2, and exhibited anti‐inflammatory properties inhibiting PAR2‐induced secretion of IL‐8 in vitro and reducing complete Freund's adjuvant (CFA)‐induced paw inflammation in mice [208].…”

Novel proteinase-activated receptor-2 (PAR2) antagonist C391 blocks Alternaria-induced human airway epithelial signaling and asthma indicators in murine models

“…Both compounds inhibited inflammation to a similar extent in a paw edema model (Kennedy et al, 2020). The PAR2 antagonists from Vertex all inhibited PAR2-induced Gprotein but not β-arrestin signaling and show partial efficacy in mouse models of pain and subcutaneous inflammation (Avet et al, 2020;Jiang et al, 2018;Jimenez-Vargas et al, 2018). While these inhibitors show efficacy in peripheral inflammation and pain models, the ability to inhibit β-arrestin signaling appears to be important for reduction of asthma indicators, making C391 a more promising therapeutic.…”

“…There are only a handful of PAR2 antagonists that have been developed [reviewed in (Yau, Lim, Liu & Fairlie, 2016)]. These include K-14585 (Goh, Ng, Nilsson, Kanke & Plevin, 2009;Kanke et al, 2005), GB88 (Hollenberg et al, 2014;Suen et al, 2012;Suen et al, 2014), two small molecules isolated from a large screening library (AC-55541 and AC-264613; (Gardell et al, 2008)), two PAR2 allosteric antagonists AZ8838 and AZ3451 from Astra Zeneca (Cheng et al, 2017;Kennedy et al, 2020), a group of small molecules: I-343, I-191 and I-287, from Vertex (Avet et al, 2020;Jiang et al, 2018;Jimenez-Vargas et al, 2018), and C391 (Boitano et al, 2015). C391 is distinct from these other antagonists in that it requires minimal pre-incubation for inhibition of multiple PAR2 signaling pathways.…”

“…A more recent advance from our laboratory is the modification of the peptide sequence of the high potency peptidomimetic PAR2 agonist, 2-furoyl-LIGRLO-NH 2 , to form a novel PAR2 antagonist pharmacophore, compound 391 [C391; (Boitano et al, 2015)]. C391 is one of a handful of recently proposed PAR2 antagonists (Avet et al, 2020;Cheng et al, 2017;Goh, Ng, Nilsson, Kanke & Plevin, 2009;Hollenberg et al, 2014;Huang, Ni, Xi, Chu, Zhang & You, 2019;Kanke et al, 2005;Suen et al, 2012;Suen et al, 2014;Yau, Lim, Liu & Fairlie, 2016). It is distinct from other reported antagonists in that it requires minimal pre-incubation for inhibition of peptide activation of PAR2-dependent Ca 2+ signaling and it acts as an antagonist for mitogen activating protein kinase (MAPK) signaling pathways.…”