Journal of Chemical Information and Modeling Article Chart 2. Chemical Structures of 10Hit Compounds FW01 FW02 FW03 FW04 FwO5 FW06 FW07 Fw08 FW09 FW10 Table 2. Summary of the Molecular Properties, Fit Values, GoldScores, and Biological Evaluations of Hit Compounds HBA IBD PH]5-HT"K(nM) 476.6 51.9±164 3475 70.5±14 FW03 0.4 1212131211 4.04 96.5±13.1 103.5±234 FwOS 4290 123.6±320 2943 225.2±20.2 344.8 445444454 544 2949±5.0 320.1±769 027D Binding data are the mean values of five to six individual experiments with 5-HTiA receptors each done in triplicate onist stimulated 3SS]GTPyS binding derived from a mean curve out of five experiments with human SHT1A receptors stably expressed in CHO cells. ND=not Hydrogen Bond Distance(A) D332N20 (a) (b) Y2,64 D3.32 s5.42 s43 Y7. 43 oI N7.39o1 o gure 5. a, Predicted binding mode of compound Fwo1(green, sticks)with S-HTIAR(white, cartoon). Hy re illustrated as otted lines. b, Evolution of the hydrogen bond distances between compound Fwol and S-htiar during the 50 ns simulation. Distance was calculated between the centroid of two equal carboxylic oxygen atoms of D3. 32 and the protonated nitrogen atom(N20)in Fwol The competitive binding assays were carried out for the binding assays. Out of the 10 compounds tested, 5 compounds purchased 45 compounds. The inhibitory potency was tested displayed K values of less than 100 nM. Novelty confirmation with each compound at a fixed concentration of 10 uM. Ten revealed that these compounds were not reported for the ompounds(Fwol-FW10, Chart 2)showed an inhibition rate affinity to 5-HTIAR. To explore agonistic or antagonistic greater than 90%, and their ki values were calculated. Table 2 properties of these compounds, compounds FwO1-FWI0 were summarizes the results of virtual screening and corresponding subjected to [S]-GTPyS function assays at 5-HTIAR. As we dxdoLor/10. 1021/c400481p. Chem. Inf Model. 2013, 53, 3202-3211The competitive binding assays were carried out for the purchased 45 compounds. The inhibitory potency was tested with each compound at a fixed concentration of 10 μM. Ten compounds (FW01-FW10, Chart 2) showed an inhibition rate greater than 90%, and their Ki values were calculated. Table 2 summarizes the results of virtual screening and corresponding binding assays. Out of the 10 compounds tested, 5 compounds displayed Ki values of less than 100 nM. Novelty confirmation revealed that these compounds were not reported for the affinity to 5-HT1AR. To explore agonistic or antagonistic properties of these compounds, compounds FW01-FW10 were subjected to [35S]-GTPγS function assays at 5-HT1AR. As we Chart 2. Chemical Structures of 10Hit Compounds Table 2. Summary of the Molecular Properties, Fit Values, GoldScores, and Biological Evaluations of Hit Compounds hit MW HBA HBD AlogP fit value GoldScore [3 H]5-HTa Ki (nM) [35S]GTPγSb EC50 (nM) FW01 476.6 5 1 5.84 3.60 50.7 51.9 ± 16.4 7 FW02 347.5 4 2 3.35 3.74 47.4 70.5 ± 14.8 77 FW03 350.4 4 1 4.04 2.96 48.5 96.5 ± 13.1 404 FW04 394.5 5 2 3.77 3.40 45.7 103.5 ± 23.4 128 FW05 429.0 4 1 6.10 3.17 50.7 123.6 ± 32.0 534 FW06 294.3 4 3 3.27 3.10 52.0 133.4 ± 9.7 434 FW07 397.5 4 1 4.60 3.70 45.2 225.2 ± 20.2 340 FW08 344.8 4 2 4.20 3.15 54.4 294.9 ± 5.0 572 FW09 446.0 5 1 4.74 3.50 59.2 320.1 ± 76.9 597 FW10 396.9 4 1 4.27 3.48 50.1 351.7 ± 19.0 NDc 5-HT 1.8 ± 0.1 3 a Binding data are the mean values of five to six individual experiments with 5-HT1A receptors each done in triplicate. b Agonist stimulated [ 35S]GTPγS binding derived from a mean curve out of five experiments with human 5-HT1A receptors stably expressed in CHO cells. c ND = not determined. Figure 5. a, Predicted binding mode of compound FW01 (green, sticks) with 5-HT1AR (white, cartoon). Hydrogen bonds are illustrated as cyan dotted lines. b, Evolution of the hydrogen bond distances between compound FW01 and 5-HT1AR during the 50 ns simulation. * Distance was calculated between the centroid of two equal carboxylic oxygen atoms of D3.32 and the protonated nitrogen atom (N20) in FW01. Journal of Chemical Information and Modeling Article 3207 dx.doi.org/10.1021/ci400481p | J. Chem. Inf. Model. 2013, 53, 3202−3211