The compounds listed in Table Table Table Table Table
The compounds listed in Table 1, Table 2, Table 3, Table 4, Table 5 were biologically evaluated for their inhibition of the specific binding of a radiolabeled ligand [3H]PGE2 to membrane fractions prepared from cells stably expressing each mouse prostanoid receptor. The EP1 antagonist activity of these compounds was determined by a Ca2+ assay using the mouse EP1 receptor (mEP1) expressed in mouse CHO cells in the presence of 0.1% of bovine serum albumin (BSA). During the course of screening our in-house compound library, compound 1 was found to show subtype-selective EP1 receptor affinity (Table 1). Removal of the metabolically unstable para-ester moiety and the meta-methyl residue of 1 afforded 2, which showed 5.0-fold less potent EP1 receptor affinity. Introduction of a 4-chloro residue into the anthranilic MS023 of moiety of 2 afforded 3, which had 2.7-fold more potent receptor affinity. Replacement of the phenylsulfonyl moiety of 3 with a 4-chlorophenylsulfonyl moiety provided 4, which retained its EP1 receptor affinity and showed more potent EP2 receptor affinity. Further optimization was initiated with the chemical modification of 3 and 4. Optimization of the carboxyamide moiety of the chemical leads was carried out as shown in Table 2. Transformation of the carboxyamide moiety of 3 into an aminocarbonyl moiety gave 5, which showed 10-fold less potent EP1 receptor affinity. Replacement of the carboxyamide moiety of 4 with an ethylene moiety produced 6, which retained EP1 receptor affinity and showed increased affinity for the EP3 receptor. Replacement of the carboxyamide moiety of 4 with a cis-double bond and trans-double bond led to 7 and 8, respectively. The trans-isomer 8, which is relatively close to the carboxyamide structure, showed more potent EP1 receptor affinity than the cis-isomer 7 and compound 8 retained potent EP1 receptor affinity. Replacement of the carboxyamide moiety of 4 with a triple bond led to 9, which had 2.5-fold more potent EP1 receptor affinity and showed a marked increase of EP3 receptor affinity. Replacement of the carboxyamide moiety of 4 with methyleneoxy and oxymethylene moieties provided 10 and 11, respectively, with 4.5-fold less potent and 5.3-fold more potent EP1 receptor affinity, respectively. Removal of the chloro residue from the more potent ether analog 11 gave 12, which had 1.6-fold more potent EP1 receptor affinity. Compounds 11 and 12 did not exhibit antagonist activity at 10μM, although both showed higher EP1 receptor affinity than the other compounds listed in Table 1, Table 2. Overall, phenylsulfonylamino analogs 3 and 12 tended to show better subtype selectivity than the corresponding 4-chlorophenylsulfonyl analogs 4 and 11, respectively. As illustrated in Table 3, the effect on Ki value of transformation of the phenylsulfonylamino moiety of 12 was investigated. The corresponding phenylaminosulfonyl analog 13 demonstrated a 15-fold decrease of receptor affinity. Replacement of the sulfonylamino moiety of 12 with an oxymethyl moiety led to 14, with 6.0-fold less potent EP1 receptor affinity and increased affinity for the EP2 receptor. As a result, the phenylsulfonylamino moiety was confirmed to be an optimum structure. As shown in Table 4, the effect on Ki values of substituting the aminophenoxy moiety of 12 was investigated. The 5-methyl analog 16 and the 5-trifluoromethyl analog 18 respectively retained or had slightly more potent receptor affinity than 12, while the 5-fluoro analog 15, 5-methoxy analog 17, and 4-methyl analog 19 all showed a decrease of receptor affinity. Among the compounds tested, the 5-trifluoromethyl analog 18 showed the most potent EP1 receptor affinity. The effect of N-alkylation of 18 on receptor affinity and EP1 receptor antagonist activity was investigated as shown in Table 5. N-Ethylation of 18 afforded 20, which had 8.6-fold more potent EP1 receptor affinity. In addition, 20 exhibited EP1 receptor antagonist activity (IC50=0.68μM), while 18 did not show antagonist activity at 10μM. Replacement of the N-ethyl moiety of 20 with an N-n-propyl moiety gave 21, with a significant increase of both EP1 receptor affinity and antagonist activity. The corresponding N-isopropyl analog 22 was slightly less potent in terms of both receptor affinity and antagonist activity. Replacement of the N-ethyl moiety of 20 with N-isobutyl moiety afforded 23, which showed 5.8-fold more potent EP1 receptor affinity and 5.2-fold more potent antagonist activity. Replacement of the N-ethyl moiety of 20 with an N-neopentyl moiety led to 24, which had reduced EP1 receptor affinity, and showed weak affinity for the EP3 and EP4 receptors.