To study systemic potential of the selected compounds
To study systemic potential of the selected compounds as EP1 receptor antagonist, they were evaluated with regard to the sulprostane-induced increase of intravesical bladder pressure in rats. Some of the analogs, 13 and 15–17, which were selected on the basis of their in vitro EP1 receptor affinity, were evaluated for their in vivo potency after intraduodenal (id) administration as described in Table 6. These four analogs were found to be effective in an animal model after id administration (1mg/kg). Their potency was equipotent or slightly more potent relative to 2.
Table 7 describes the binding affinities of 13 and 15–17 for the other prostanoid receptors, hTP, hDP, mFP, and hIP. These analogs were also evaluated for their ability to antagonize the hEP1 receptor. As expected, all the compounds were proved to have potent hEP1 receptor antagonist activity, whereas 13 and 16–17 had weak affinity for the mFP receptor and 15 did not show any affinity at a concentration of 10μM. As a result, all the selected analogs were found to be highly selective EP1 receptor antagonist.
As shown in Table 8, 13 and 15–17 were tested for their inhibition of cytochrome P450 isozymes. Compound 13 produced 24% and 13% inhibition against 2C9 and 3A4, respectively, whereas 15 produced stronger inhibition (44%) against 2C9 and weaker inhibition (18%) against 2C19. Compound 17 exhibited weak inhibitory activity (13%) against 2C9. Benzoic ddr1 synthesis analogs 15 and 16 had significantly stronger inhibitory activity than 1, and 3-methylbenzoic analogs 13 and 17 had equipotent to slightly less potent inhibitory activity against 2C9 relative to 15 and 16. Analogs 13 and 15–17 had significantly weaker inhibitory activity against 2C9 and 2C19 relative to 2. These analogs seem to be particularly promising as drug candidates because of their reduced inhibitory activity against 3A4, given that the 3A4 isozyme is known to metabolize many drugs. On the basis of the data provided above, the potential risk of harmful drug interactions and/or risk of tolerance developing for 13 and 15–17 could be significantly less than that for 2.
In conclusion, we have discovered EP1 receptor-selective antagonists 13 and 15–17, which are promising clinical candidates, starting with a molecular design based on the structural hybridization of 1 and 2, which show moderate in vivo activity with no P450 inhibition and an increased in vivo activity with strong P450 inhibition, respectively. The 4,5-dimethylbenzene moiety of 8, 12, 15, and 17, and the indane moiety of 9, 13, 16, and 18 were newly identified as surrogates for a trifluoromethylbenzene moiety of 1 and 2. As illustrated by 13 and 17, 3-methylbenzoic acid analogs were found to have slightly reduced P450 inhibitory activity with preserved antagonist activity.
Biological assay method
Introduction Prostanoid receptors are known to be members of the G-protein coupled receptor superfamily. Recently, eight prostanoid receptors (EP1, EP2, EP3, EP4, IP, TP, DP, and FP) have been cloned and characterized. The characterization of these receptors at the molecular level has resulted in renewed interest in this field, but selective agonists and antagonists of human prostanoid receptors are only available in some limited field.3, 4, 5, 6 As a result, the correlation of specific receptors with various pathologies is currently being established by using potent but poorly selective ligands. Recent studies have suggested that the EP1 subtype receptor mediates the induction of pain, pyrexia, allodynia and diuresis. Based on this information, compounds that are selective antagonists of this receptor are predicted to be useful as analgesics, antipyretics, and agents to treat hyperalgesia and pollakisuria. A few antagonists, such as ZD-4953, ZD-6416, and ZD-6804, entered clinical development for indications related to hyperalgesia, but their development was suspended for unknown reasons. Our search for a subtype-selective EP1 receptor antagonist started with screening our in-house compound library. 4-(2-Arylsulfonylaminobenzoyl)amino benzoic acids 2–4 were the new chemical leads derived from the initial chemical lead 1, which was identified by screening. Here we report on the discovery process (Fig. 1) for a new class of 4-( 2-[alkyl(phenylsulfonyl)amino]phenoxy methyl)benzoic acids 21–23 which are potent subtype-selective EP1 receptor antagonists.