Tail group SAR of the imidazole
Tail group SAR of the imidazole derived analogs is shown in . The previous SAR study from the discovery of AMG 837 revealed that a simple un-substituted meta-biphenyl tail group was less favorable in terms of potency. Efforts to introduce polarity to the tail group were not successful. When a methylsulfonyl, cyano or methoxy group were added to the different positions of the C ring, all yielded various level of diminished activity (e.g., –). This was not entirely unexpected. As endogenous GPR40 ligands are FFAs, the ligand binding site in the receptor is hydrophobic in nature. It was encouraging to find that a trifluoromethoxy group at the -position of the C ring gave an imidazole head group Resiniferatoxin with similar potency as AMG 837 ( vs. AMG 837). However, the activity of is significantly reduced in the presence of 100% human serum. A breakthrough was revealed when a methyl or chlorine was introduced at the -position of the C ring (, ), yielding compounds 2-fold more potent than AMG 837. Adding one more methyl at the other position of the C ring or increasing the size of the substitution relative to the methyl group does not further improve potency ( and vs. ). With the encouraging results from compounds and , the combination of 2-methyl with a methyl, chloro or fluoro were screened (–). Compound (AM-3189) stands out by displaying greater potency both in buffer and in 100% human serum. (AM-3189) also possesses favorable drug-like properties with a calculated log (pH 7.4) of 3.01, tPSA of 64Å and more favorable lipophilic efficiency (4.4 compared to 3.4 for AMG 837). Adding a methyl group to the -positions of the B ring proximal to the C ring of , targeting restricted rotation of the rotable bond between the B ring and C ring to reduce possible conformers, yielded less potent compounds and . All tested compounds in have similar or lower than that of AMG 837, which indicates that, like AMG 837, these imidazole-derived compounds are partial agonists on the GPR40 receptor. To confirm the activity difference between a pair of enantiomers that was observed previously, the opposite enantiomer of (AM-3189) was also synthesized and was found to be >100× less active in the GPR40 aequorin assay. Similar to AMG 837, while highly potent on GPR40, (AM-3189) was highly selective over the closely related GPCRs, GPR41 and GPR43. showed no significant activity in cell-based assays against PPAR-α, -δ, and -γ, despite a possible structural resemblance to some PPAR agonists. An external panel of 64 receptors (MDS Pharma) also revealed no significant activity. Overall, was both highly potent and selective in vitro. In addition to its favorable profile in vitro, compound (AM-3189) distinguished itself by displaying an excellent pharmacokinetic profile in multiple species. As shown in , demonstrated low clearance, moderate volume of distribution, and good oral bioavailability. (AM-3189) does not penetrate the rat CNS as indicated by a rat brain to plasma ratio of 0.04 at 3h after an oral dose of 5mg/kg. The reduced CNS penetration may be due to P-glycoprotein mediated efflux as shown in . The activity of (AM-3189) was examined in in vitro insulin secretion assays, first using human islets. As was observed in recombinant cell systems, activity of and AMG 837 was similar in this assay (A). We also evaluated the activity of in islets isolated from human GPR40 knock-in mice. In these mice, the mouse GPR40 gene has been replaced with the human GPR40 gene using genetic recombination technology. increased glucose stimulated insulin secretion from human GPR40 knock-in islets (B). (AM-3189) was significantly less potent and efficacious on the rodent GPR40 receptor compared to the human receptor. In the presence of 0.01% HSA, the EC value of on rat GPR40 was 640±60nM and the (% of AMG 837) value was 41%. Similar weak activity was seen on receptors from other non-primate preclinical species, including mouse, guinea pig, hamster, and pig (data not shown). (AM-3189) was a potent agonist of dog GPR40 (EC=37nM) but had low efficacy compared to AMG 837 (=37% of AMG 837). (AM-3189) had good activity against the cynomolgus monkey receptor, with EC and (% of AMG 837) values of 29nM and 96%, respectively.