Potent Thiophene Antagonists of Human Complement C3a Receptor with Anti-Inflammatory Activity

Abstract

Structure–activity relationships for a series of small-molecule thiophenes resulted in potent and selective antagonism of human Complement C3a receptor. The compounds are about 100-fold more potent than the most reported antagonist SB290157. A new compound JR14a was among the most potent of the new antagonists in vitro, assessed by (a) inhibition of intracellular calcium release (IC50 10 nM) induced in human monocyte-derived macrophages by 100 nM C3a, (b) inhibition of β-hexosaminidase secretion (IC50 8 nM) from human LAD2 mast cells degranulated by 100 nM C3a, and (c) selectivity for human C3aR over C5aR. JR14a was metabolically stable in rat plasma and in rat liver microsomes and efficacious in rats when given orally to suppress rat paw inflammation, macrophage and mast cell activation, and histopathology induced by intraplantar paw administration of a C3aR agonist. Potent C3aR antagonists are now available for interrogating C3a receptor activation and suppressing C3aR-mediated inflammation in mammalian physiology and disease.

The ‘C3aR antagonist’ SB290157 is a partial C5aR2 agonist

“…The discrepancy could be explained by the competitive nature of SB290157-mediated inhibition. The lower C3a used in the present study, 5 nM versus 100 nM in Rowley et al (32), would give rise to a lower apparent IC 50 value of SB290157 (33). In marked contrast to the CHO-C3aR data, SB290157 did not display any agonistic activity on ERK signalling in HMDMs at the highest possible dose tested of 100 μM, although human C3a displayed an EC 50 of 0.21 nM ( Figure 3B ), in accordance with the previous data (23, 32).…”

“…The lower C3a used in the present study, 5 nM versus 100 nM in Rowley et al (32), would give rise to a lower apparent IC 50 value of SB290157 (33). In marked contrast to the CHO-C3aR data, SB290157 did not display any agonistic activity on ERK signalling in HMDMs at the highest possible dose tested of 100 μM, although human C3a displayed an EC 50 of 0.21 nM ( Figure 3B ), in accordance with the previous data (23, 32). We also performed dose titrations of SB290157 to ensure the absence of agonism was not a result of the ERK signalling being down-regulated, as experienced by higher concentrations of human C3a ( Figure 3B ).…”

“…We therefore assessed the ability of SB290157 to inhibit this C3a-mediated signalling pathway, and found that this compound dose-dependently inhibited C3a-induced ERK signalling in HMDMs ( Figure 3A ). The IC 50 of SB290157 (236 nM), was ~16-fold more potent than previously reported data (IC 50 = 3.8 μM), determined using an intracellular calcium mobilisation assay in the same cells (32). The discrepancy could be explained by the competitive nature of SB290157-mediated inhibition.…”

“…Thus, our data further caution against the use of this compound as the sole tool to delinate C3aR biology and function. Until further validated selective C3aR inhbitory tools become available (32), we strongly recommend the use of other genetic approaches such as gene-knockout mice, or gene knockdown approaches, to validate roles for C3aR. If SB290157 usage is absolutely required, an in vivo dose of no more than 1 mg/kg ( i.p. )…”

Tackling COVID-19 infection through complement-targeted immunotherapy

“…During the past 10 years, the Fairlie lab has developed sophisticated approaches to design small molecule agonist and antagonists from proteins including C3a (Reid et al, 2013). In this context, they have recently reported on the new compound JR14a, a very potent C3aR antagonist that is 100-fold more potent than SB290157 (Rowley et al, 2020). This molecule awaits broad preclinical testing in animal models of inflammation.…”