Abstract
The structure activity relationships of 2-oxy pyridine derivatives in the C-region of N-(6-trifluoromethyl- pyridin-3-ylmethyl) 2-(3-fluoro-4-methylsulfonylaminophenyl)propanamides as hTRPVl antagonists were investigated. The analysis indicated that the lipophilicity of the 2-oxy substituents was critical for potent antagonism and 4 or 5 carbons appeared to be optimal for activity. Multiple compounds proved to have comparable activity to 1, which had been reported as the most potent antagonist for capsaicin activity among the previous series of compounds. Further analysis of compounds 22 (2-isobutyloxy) and 53 (2-benzyloxy) in the formalin test in mice demonstrated strong analgesic activity with full efficacy. Docking analysis of 53S using our hTRPVl homology model indicated that the A- and B-region 2-(3-fluoro-4-methylsulfonylaminophenyl)propanamide made important hydrophobic and hydrogen bonding interactions with Tyr511 and that the C-region 6-trifluoromethyl and 2-benzyloxy groups of pyridine occupied the two hydrophobic binding pockets, respectively.
Keywords: TRPV1 antagonists, Analgesic, Molecular modeling, Capsaicin
1. Introduction
The transient receptor potential VI (TRPV1) receptor is a nonselective cation channel with high Ca2+ permeability which functions as a molecular integrator of nociceptive stimuli [1]. The receptor is activated by endogenous agonists including protons [2], noxious heat [3], and inflammatory lipid mediators [4,5] as well as by natural products such as capsaicin (CAP) [6] and resiniferatoxin (RTX) [7]. The increase in intracellular Ca2+ uponTRPVl activation causes excitation of the primary sensory neurons and the consequent central perception of pain. TRPVl antagonists, by inhibiting this transmission of nociceptive signaling from the periphery to the CNS, have thus attracted much attention as potential analgesics. In recent years a number of TRPVl antagonists have been developed as novel analgesic and anti-inflammatory agents, particularly for the treatment of neuropathic pains [8]. The clinical development and therapeutic potential of TRPVl antagonists have been extensively reviewed [9–15].
Recently, we investigated a series of N-(2-amino-6-trifluoro methyl-pyridin-3-ylmethyl)2-(3-fluoro-4-methylsulfonylaminophenyl)propanamides, designed by combining previously identified pharmacophoric elements, as hTRPVl antagonists [16]. Among them, compound 1 showed the most potent antagonism with Ki(CAP) = 0.3 nM and IC50(pH) = 8.4 nM, being thus 150-fold and 230fold better than precedent lead 2 [17] for CAP and pH antagonism, respectively (Fig. 1). In addition, the compound 1S, the S-form of 1, was found to be highly selective for TRPV1 with Ki(CAP) = 0.2 nM and displayed strong analgesic activity in a neuropathic pain model with almost no side effects. Consistent with its mechanism of action in vivo being through TRPV1, compound 1S blocked capsaicin-induced hypothermia but caused modest TRPV1-related hyperthermia in mice.
Fig. 1.
Lead TRPV1 antagonists.
Our structural analysis indicated that the enhanced potency of 1 compared to 2 was attributable to a new hydrophobic interaction with the receptor, afforded by the additional 4-methylpiperidine moiety in 1. Specifically, docking analysis using the hTRPV1 homology model which we developed indicated that the 4-methylpiperidinyl group in the C-region of 1 interacted with a hydrophobic region on the receptor composed of Met514 and Leu515 [16].
As a continuation of our SAR analysis of the 2-substituent in the N-(6-trifluoromethyl-pyridin-3-ylmethyl) C-region, we have investigated its 2-oxy derivatives (Fig. 2) as hTRPV1 antagonists. For a selected potent antagonist in the series, we further characterized analgesic activity in the formalin pain model and performed molecular modeling with our hTRPV1 homology model.
Fig. 2.
General structure of the designed compounds.
2. Results and discussion
2.1. Chemistry
The syntheses of the final compounds are represented in Scheme 1. 2-Oxy substituted pyridines 5 were synthesized either by O-alkylation of pyridone 3 [16] or by the nucleophilic substitution of various alcohols with 2-chloropyridine 4 [16]. Pyridines 5 containing cyclopropyl substituents were prepared from the corresponding alkenes by the Simmons–Smith reaction, as shown in Scheme 2. The nitrile groups of 5 were reduced to yield the corresponding primary amines 6. The amines were coupled with 2-(3-fluoro-4-methylsulfonylaminophenyl)propionic acid 7 [17] to provide the final compounds 8–71.
Scheme 1.
Syntheses of 2-(3-fluoro-4-methylsulfonylaminophenyl)propanamide analogs. Reagents and conditions: (a) [Method A] RBr, K2CO3, 18-crown-6, DMF, heat; (b) [Method B] ROH, DBU, 1,4-dioxane or CH3CN, reflux: [Method C] ROH, NaH, THF, reflux: [Method D] ROH, Cs2CO3, DMF, heat: [Method E] ROH, KF, DMF, heat; (c) [Method F] 2 M BH3-SMe2 in THF: [Method G] H2, 10% Pd-C, AcOH, MeOH: [Method H] LAH, diethylether: [Method I] NaBH4, NiCl2–6H2O, MeOH; (d) EDC, HOBt, DMF.
Scheme 2.
Syntheses of the 2-cyclopropylmethyloxy pyridine C-region. Reagents and conditions: (a) diiodomethane, diethylzinc, CH2Cl2.
2.2. Structure–activity relationship (SAR) analysis
The synthesized TRPV1 ligands were evaluated in vitro for antagonism as measured by inhibition of activation by capsaicin (CAP) and pH as indicated. The assays were conducted using a fluorometric imaging plate reader (FLIPR) with human TRPV1 heterologously expressed in Chinese hamster ovary (CHO) cells [16]. The results are summarized in Tables 1–5, together with the potencies of the previously reported parent antagonist 1.
Table 1.
In vitro hTRPV1 antagonistic activities for straight 2-alkyloxy derivatives.
 |
| R | Ki [CAP] (nM) | IC50 [pH] (nM) | R | Ki [CAP] (nM) | IC50 [pH] (nM) | ||
|---|---|---|---|---|---|---|---|
| 1 | 0.3 | 8.4 | 13R |  |
WE | WE | |
| 8 |  |
68.8 | WE | 14 |  |
1.6 | 183 |
| 9 |  |
12.7 | 277 | 15 |  |
1 | 43 |
| 10 |  |
0.9 | 90.1 | 16 |  |
49 | WE |
| 11 |  |
0.3 | 30.5 | 17 |  |
50 | WE |
| 12 |  |
0.7 | 27.3 | 18 |  |
15.3 | WE |
| 13 |  |
0.5 | 29.9 | 19 |  |
9.7 | 462 |
| 13S |  |
0.4 | 12.3 | 20 |  |
1.5 | 161 |
WE: weakly effective (1 μM < Ki or IC50 < 10 μM).
NE: not effective (Ki or IC50 > 10 μM).
Table 5.
In vitro hTRPV1 antagonistic activities for 2-aryloxy and arylmethyloxy derivatives.
| R | Ki [CAP] (nM) | IC50 [pH] (nM) | R | Ki [CAP] (nM) | IC50 [pH] (nM) | ||
|---|---|---|---|---|---|---|---|
| 51 |  |
6.4 | WE | 62 |  |
2.6 | 653 |
| 52 |  |
2.8 | WE | 63 |  |
0.8 | 69 |
| 53 |  |
0.5 | 18.6 | 64 |  |
4.9 | 95 |
| 54 |  |
0.5 | 27.6 | 65 |  |
0.8 | 43.7 |
| 55 |  |
1.5 | 53 | 66 |  |
1.0 | 93 |
| 56 |  |
3.7 | 165 | 67 |  |
2.6 | 114 |
| 57 |  |
16 | 337 | 68 |  |
0.7 | 76 |
| 58 |  |
2.1 | 182 | 69 |  |
29.5 | WE |
| 59 |  |
2.5 | 25 | 70 |  |
142 | WE |
| 60 |  |
2.5 | 132 | 71 |  |
51 | WE |
| 61 |  |
3.2 | 66.1 |
WE: weakly effective (1 μM < Ki or IC50 < 10 μM).
NE: not effective (Ki or IC50 > 10 μM).
To investigate the SAR for 2-oxy derivatives of the pyridine C-region we began with the straight 2-alkyloxy derivatives (Table 1). Starting from the 2-methoxy derivative 8, the antagonistic activity was enhanced sharply as the number of carbons in the chain increased until reaching a maximum with the 2-butoxy derivative 11, which was as potent as 1 with Ki(CAP) = 0.3 nM. The activity stayed constant upon further chain lengthening through the hexyloxy derivative 13, with all showing highly potent antagonism (11–13). The two stereoisomers of 13 showed stereospecificity for their antagonistic activity. 13S was the eutomer and 13R was the distomer, consistent with previous findings [16,18]. Unsaturation of the alkyl chain led to a decrease in activity. For example, the Z-alkene isomers 14 and 15 were ca. 5-fold and 1.5-fold less potent than the alkyl surrogates 11 and 12, respectively, for CAP antagonism. Introduction of oxygen to the alkyl chain caused a dramatic loss of activity. For example, the oxygenated analogs 16 and 19 exhibited 160-fold and 20-fold less potent CAP antagonism than did the parents 11 and 13, respectively. Similarly, 17 and 18, analogs of 12 oxygenated at different positions along the alkyl chain, were 70-fold and 22-fold less potent than the parent, respectively. However, addition of a lipophilic phenyl group to 18 to yield compound 20 led to potent antagonism. The SAR analysis for this series of straight 2-alkyloxy group thus indicated that the lipophilicity of side chain was critical for antagonism, probably due to its hydrophobic interaction with the receptor, and the optimal number of carbons appeared to be four.
Next, the SAR of branched 2-alkyloxy derivatives was investigated (Table 2). Similar to the findings for the derivatives with straight alkyl chains, the branched 4 carbon derivative 22 showed the most potent antagonism, but with high potency being retained with longer substituents up to the 9-carbon derivative 28. The comparison of activity between straight and branched alkyl derivatives indicated that the straight alkyl derivatives generally showed slightly better antagonism than did the corresponding branched ones (for example, 10 vs 21, 11 vs 22, 12 vs 23/25 and 13 vs 24). Interestingly, compounds 25–28 were found to have a unique profile in which they highly antagonized the activation by capsaicin but not by pH. This is an important distinction. Hyperthermia associated with antagonism is a side effect of concern. SAR studies have shown that antagonists may differ in their relative activities against agonism by capsaicin, by low pH, and by elevated temperature. Antagonism of the response to low pH is the predominant predictor of whether an antagonist may induce hyperthermia as a side effect, so antagonists blocking the response to capsaicin but not low pH may be of particular interest for further evaluation [19].
Table 2.
In vitro hTRPV1 antagonistic activities for branched 2-alkyloxy derivatives.
| R | Ki [CAP] (nM) | IC50 [pH] (nM) | R | Ki [CAP] (nM) | IC50 [pH] (nM) | ||
|---|---|---|---|---|---|---|---|
| 21 |  |
2.1 | 93.4 | 25 |  |
1.1 | NE |
| 22 |  |
0.5 | 52.6 | 26 |  |
0.7 | WE |
| 23 |  |
0.8 | 15.1 | 27 |  |
0.5 | WE |
| 24 |  |
0.9 | 23.8 | 28 |  |
0.7 | WE |
WE: weakly effective (1 μM < Ki or IC50 < 10 μM).
NE: not effective (Ki or IC50 > 10 μM).
The SAR of 2-cycloalkyloxy derivatives was investigated next (Table 3). The analysis indicated that the SAR pattern was similar to that of the above series, and compound 30 with 5 carbons was optimal for antagonism. To examine the effect of substitution on the cyclic ring, the cyclohexyl derivatives of 31 were further investigated. Whereas the 4-trifluoromethyl group in 32 enhanced antagonism compared to 31, the 4-methyl group in 33 reduced activity, suggesting that lipophilicity is a contributor to the activity. The 4-ethyl and 4-t-butyl derivatives, 34 and 35, reduced the antagonism, probably due to steric repulsion with the receptor. However, the 4,4-dimethyl and 3,5-dimethyl analogs, 36 and 37, retained good antagonism. The results indicated that lipophilicity at the 4-position of the cyclohexyl ring was important for potent antagonism, but there were steric constraints such that size bigger than methyl led to the reduction of activity. A similar SAR pattern was found for the 4-position of piperidine ring in the C-region previously [16]. Introduction of nitrogen at the 4-position in 38 led to dramatic loss of activity. However, its activity was recovered by adding the lipophilic Boc group to the nitrogen, as in 39. Compound 40, the phenyl-fused analog of 30, still displayed good CAP antagonism, indicating that the phenyl group was tolerated.
Table 3.
In vitro hTRPV1 antagonistic activities for 2-cycloalkyloxy derivatives.
| R | Ki [CAP] (nM) | IC50 [pH] (nM) | R | Ki [CAP] (nM) | IC50 [pH] (nM) | ||
|---|---|---|---|---|---|---|---|
| 29 |  |
1.4 | 69.8 | 35 |  |
7.7 | 242 |
| 30 |  |
0.9 | 28.7 | 36 |  |
0.9 | 140 |
| 31 |  |
1.3 | 43.4 | 37 |  |
3 | 174 |
| 32 |  |
0.3 | 14 | 38 |  |
WE | NE |
| 33 |  |
2.9 | 43.1 | 39 |  |
1.3 | 293 |
| 34 |  |
4.8 | 125 | 40 |  |
2.7 | WE |
WE: weakly effective (1 μM < Ki or IC50 < 10 μM).
NE: not effective (Ki or IC50 > 10 μM).
Next, the SAR of 2-cycloalkylmethyloxy derivatives was investigated. This series was designed by inserting a methyl group into the compounds of the 2-cycloalkyloxy series of Table 3. Generally the insertion led to a 2- to 5-fold enhancement in antagonism compared to that of the corresponding parent compounds (for example, 29 vs 44, 30 vs 45, 31 vs 46, and 33 vs 47). Of particular interest, compounds 42, 44–46 and 50 displayed high potency in the sub-nanomolar range. Compounds 48–50 demonstrated a similar SAR pattern to that shown in Table 3 in which the introduction of a nitrogen at the 4-position abolished the activity. Conversely, compound 50 exhibited excellent antagonism, suggesting that the N-Boc moiety made an appropriate hydrophobic interaction with the receptor.
Finally, we sought to evaluate the SAR of 2-aryloxy and 2-arylmethyloxy derivatives. In the 2-aryloxy series, the 2-phenyloxy derivative 51 exhibited reasonable antagonism and its 4-methylation, providing 52, further increased activity. The 2-arylmethyloxy derivatives showed better antagonism compared to the corresponding 2-aryloxy ones (51 vs 54, 52 vs 55), as described above for the 2-cycloalkyloxy and 2-cycloalkylmethyloxy series. The 2-phenylethyloxy derivative 54 was found to be as potent as 53. Further optimization was conducted with the 2-benzyloxy derivative 53 to examine the effect of substitution. Introduction of 4-substituents on the benzyl group, including alkyls (55–58), halogens (59–60), trifluoromethyl (61) and methoxy (62), decrease the antagonism slightly. In the other hand, 3-substituted benzyl derivatives showed better antagonism compared to the corresponding 4-substituted surrogates (for example, 59 vs 63 for F, 60 vs 64 for Cl, 62 vs 65 for OCH3). 3,5-Disubstituted derivatives displayed activity similar to that of the corresponding 3-substituted ones (for example, 63 vs 66 for F, 64 vs 67 for Cl, 65 vs 68 for OCH3). 2-Pyridinylmethyloxy derivatives (69–71) were much less potent than the 2-benzyloxy derivative 53, confirming that incorporation of nitrogen into the ring was detrimental to antagonism.
Detailed in vitro activity of 22 and 53, the two selected antagonists in this study, was investigated for multiple TRPV1 activators including capsaicin, pH, heat (45 °C) and N-arachidonoyl dopamine (NADA), and compared to the activity of lead 1 (Table 6). Both showed excellent antagonism of all four TRPV1 activators and comparable activities to 1. Particularly, compound 53 exhibited excellent potency toward NADA activation.
Table 6.
In vitro hTRPV1 antagonistic activities for 1, 22 and 53 to multiple activators.
| Activators | 1 (nM) | 22 (nM) | 53 (nM) |
|---|---|---|---|
| CAP (f) Ki | 0.3 | 0.5 | 0.5 |
| pH (IC50) | 8.4 | 52.6 | 18.6 |
| Heat 45 °C (IC50) | 4.5 | 67.5 | 52.3 |
| NADA (f) Ki | 0.23 | 0.29 | 0.001 |
2.3. Analgesic activity
We evaluated the in vivo analgesic activities of two selected antagonists, 22 and 53, in the formalin test [20] in mice upon oral administration. Compound 22 showed a significant antinociceptive effect, with 63.4 ± 11.5% and 60.8 ± 9.9% (mean ± SEM) inhibition of response at the doses of 0.1 and 0.3 mg/kg, respectively (p < 0.05). Compound 53 inhibited the nociceptive response by 41.8 ± 17.3% and 54.1 ± 26.3% at the doses of 0.1 and 0.3 mg/kg, respectively (p < 0.05). Since we had observed that TRPV1 knockout mice showed approximately 50% of the magnitude of response in the formalin test as was seen in wild-type mice (unpublished observations), the inhibition of the formalin response that we found for the two antagonists would correspond to the expected result for full TRPV1 blockade.
2.4. Molecular modeling
Using our human TRPV1 (hTRPV1) model [16], built based on our rat TRPV1 (rTRPV1) model [21], we performed a flexible docking study of compound 53S, the active isomer of 53. As shown in Fig. 3, a binding mode generally similar to that of 1S [16] was obtained with the S form of compound 53. The sulfonylaminobenzyl group (A-region) occupied the deep bottom hole and was involved in a hydrophobic interaction with Tyr511. A fluorine atom of the A-region participated in hydrogen bonding with Ser512 and Tyr555 and NH of the sulfonamide group made hydrogen bonds with Ser512. The amide group (B region) made a hydrogen bond with Tyr511 and also contributed to the appropriate positioning of the C-region for the hydrophobic interaction. In addition, the 3-trifluoromethyl group (C-region) extended toward the upper hydrophobic areas composed of Leu547 and Thr550, forming hydrophobic interactions. Furthermore, the 2-benzyloxy group in the C-region made an additional hydrophobic interaction with the hydrophobic region composed of Met514 and Leu515.
Fig. 3.
Flexible docking result of 53S in the hTRPVl model. (A) Binding mode of 53S, the S isomer of 53. The key interacting residues are marked and displayed as capped-stick with carbon atoms in white. The helices are colored by gray and the helices of the adjacent monomer are displayed in line ribbon. Compound 53S is depicted as ball-and-stick with carbon atoms in magenta. The van der Waals surface of the ligand is presented with its lipophilic potential property. Hydrogen bonds are shown as black dashed lines and non-polai hydrogens are undisplayed for clarity. (B) Surface representations of the docked ligand and hTRPVl. The Fast Connolly surface of hTRPVl was generated by MOLCAD and colored by the lipophilic potential property. The surface of hTRPVl is Z-clipped and that of the ligand is in its carbon color for clarity. (C) Van der Waals surface of the ligand colored by its lipophilic potential property.
3. Conclusion
The structure activity relationship of 2-oxy pyridine derivatives in the C-region of N-(6-trifluoromethyl-pyridin-3-ylmethyl) 2-(3-fluoro-4-methylsulfonylaminophenyl)propanamides as hTRPV1 antagonists was investigated. The analysis indicated that the lipophilicity of the 2-oxy substituents was a key determinant of antagonism. Generally, as the lipophilicity of the 2-oxy substituents increased, the antagonism was enhanced until it reached a maximal value, after which it remained constant. The number of 4 or 5 carbons appeared to be optimal for activity. Numerous compounds (11, 13, 22, 27, 32, 42, 44, 45, 50, 53 and 54) were found to have comparable potency to 1, which had recently been reported as the most potent antagonist of a previous series, with a range of Ki(CAP) = 0.3–0.5 nM. Among the compounds, 22 (2-isobutyloxy) and 53 (2-benzyloxy) demonstrated strong analgesic activity in the formalin test in mice with full efficacy. Docking analysis of 53S with our hTRPVl homology model indicated that the 2-(3-fluoro-4-methylsulfonylaminophenyl)propanamide (A and B-regions) made critical hydrophobic and hydrogen bonding interactions with Tyr511, and the 6-trifluoromethyl and 2-benzyloxy groups of pyridine (C-region) interacted the two hydrophobic regions composed of Leu547/Thr550 and Met514/Leu515, respectively.
4. Experimental
4.1. Chemistry
All chemical reagents were commercially available. Melting points were determined on a Buchi Melting Point B-540 apparatus and are uncorrected. Silica gel column chromatography was performed on silica gel 60, 230–400 mesh, Merck. Nuclear magnetic resonance (1H NMR) spectra were recorded on a JEOL JNM-LA 300 and Bruker Avance 400 MHz FT-NMR spectrometer. Chemical shifts are reported in ppm units with Me4Si as a reference standard. Mass spectra were recorded on a VG Trio-2 GC-MS and 6460 Triple Quad LC/MS. Elemental analyses were performed with an EA lll0 Automatic Elemental Analyzer, CE Instruments.
4.1.1. General procedure for the synthesis of 5
4.1.1.1. Method A (for 29–40).
To a stirred solution of 3 (1.0 mmol) in N,N-dimethylformamide (6 mL) at 0 °C was added 18-crown-6 ether (cat.), potassium carbonate (2 mmol) followed by bromide compound (l.2 mmol). The reaction mixture was heated at 60–90 °C for overnight and extracted with EtOAc several times. The combined organic layers were washed with brine, dried over MgSO4, and concentrated in vacuo. The residue was purified by flash column chromatography on silica gel using EtOAc:hexane (1:2) as eluant.
4.1.1.2. Method B (for 8–13, 21–28 and 51–58).
To a stirred solution of 4 (1.0 mmol) in 1,4-dioxane or acetonitrile (5 mL) at 0 °Cwas added 1,8-diazabicyclo[5.4.0]undec-7-ene (2 mmol) followed by alcohol compound (l.2 mmol). The reaction mixture was stirred at 50 °C for overnight, and extracted with EtOAc for several times. The combined organic layers were washed with brine, dried over MgSO4, and concentrated in vacuo. The residue was purified by flash column chromatography on silica gel using EtOAc:hexane (1:2) as eluant.
4.1.1.3. Method C (for 16–20 and 41–43).
To a stirred solution of 4 (1.0 mmol) in anhydrous THF (5 mL) at 0 °C was added sodium hydride (2.5 mmol) followed by alcohol compound (1.2 mmol). The reaction mixture was stirred at 50 ° C for 5 h, and extracted with EtOAc several times. The combined organic layers were washed with brine, dried over MgSO4, and concentrated in vacuo. The residue was purified by flash column chromatography on silica gel using EtOAc:hexane (1:2) as eluant.
4.1.1.4. Method D (for 14–15, 48–50 and 59–71).
To a stirred solution of 4 (1.0 mmol) in N,N-dimethylformamide (5 mL) at 0 °Cwas added cesium carbonate (3.0 mmol) followed by alcohol compound (1.2 mmol). The reaction mixture was stirred at 60–90 ° C for overnight, and extracted with EtOAc several times. The combined organic layers were washed with brine, dried over MgSO4, and concentrated in vacuo. The residue was purified by flash column chromatography on silica gel using EtOAc:hexane (1:2) as eluant.
4.1.1.5. Method E (for 44–47).
To a stirred solution of 4 (1.0 mmol) in N,N-dimethylformamide (5 mL) was added potassium fluoride (2.0 mmol) followed by alcohol compound (1.2 mmol). The reaction mixture was stirred at 60–90 °C for overnight and extracted with EtOAc several times. The combined organic layers were washed with brine, dried over MgSO4, and concentrated in vacuo. The residue was purified by flash column chromatography on silica gel using EtOAc:hexane (1:2) as eluant.
4.1.2. General procedure for nitrile reduction
4.1.2.1. Method F (for 16–20, 29–37 and 59–68).
To a stirred solution of nitrile (2.0 mmol) in anhydrous THF (1 mL) was added 2 M BH3SMe2 inTHF (3 mL, 3 equiv) at room temperature. After being refluxed for 8 h, the reaction mixture was cooled to room temperature and 2 M HCl solution was added. Then the mixture was refluxed for 30 min, cooled to room temperature, neutralized by 2 M NaOH solution and extracted with EtOAc several times. The combined organic layers were washed with brine, dried over MgSO4, and concentrated in vacuo. The residue was purified by flash column chromatography on silica gel using CH2Cl2:MeOH (10:1) as eluant.
4.1.2.2. Method G (for 8–13, 21–28 and 51–58).
A suspension of nitrile (5.0 mmol) and 10% Pd/C (500 mg) and concentrated HCl (3 mL) in MeOH (30 mL) was hydrogenated under a balloon of hydrogen for 6 h at room temperature and filtered through Celite. The filtrate was concentrated in vacuo and the residue was purified by flash column chromatography on silica gel using EtOAc eluant.
4.1.2.3. Method H (for 14–5, 41–47 and 69–71).
To a cooled solution of nitrile (1.0 mmol) in anhydrous THF at 0 °C (5 mL) was added slowly lithium aluminum hydride (3.0 mmol) in portion wise. The mixture was refluxed for overnight, cooled to room temperature and quenched by dropwise addition of water. The solution was filtered through Celite and dried over MgSO4, filtered, and concentrated in vacuo. The residue was purified by flash column chromatography on silica gel using CH2Cl2:MeOH (10:1) as eluant.
4.1.2.4. Method I (for 38–40 and 48–50).
To a stirred solution of nitrile (1.0 mmol) and NiCl26H2O (2.0 mmol) in MeOH (8 mL) was added sodium borohydride (4.0 mmol) slowly. The mixture was refluxed for 12 h and then cooled to room temperature. The solution was filtered through Celite and dried over MgSO4, filtered, and concentrated in vacuo. The residue was purified by flash column chromatography on silica gel using CH2Cl2:MeOH (10:1) as eluant.
4.1.3. General procedure for amide coupling
A mixture of acid (5.0 mmol), amine (5.5 mmol), N-(3-dimethylaminopropyl)-N′ -ethyl-carbodiimide hydrochloride (6.0 mmol), and 1-hydrxybenzotriazole hydrate in N,N-dime-thylformamide (20 mL) was stirred for 12 h at room temperature. The reaction mixture was extracted with EtOAc several times. The combined organic extracts were washed with 1 M HCl (25 mL) and brine (25 mL), dried over MgSO4, filtered, and concentrated in vacuo. The residue was purified by flash column chromatography on silica gel using EtOAc:hexane (1:2) as eluant.
4.1.3.1. N-((2-Methoxy-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (8).
Yield 81%, white solid, mp 122–124 °C; 1H NMR (CDCl3) δ 7.60 (d, J = 7.3 Hz, 1H), 7.52 (dd, J = 8.3, 8.3 Hz, 1H), 7.21 (d, J = 7.3 Hz, 1H), 7.12–7.06 (m, 2H), 6.50 (bs, NH), 6.00 (bt, NH), 4.37 (d, J = 6.1 Hz, 2H), 3.95 (s, 3H), 3.52 (q, J = 7.1 Hz, 1H), 3.04 (s, 3H), 1.49 (d, J = 7.0 Hz, 3H); MS (FAB) m/z 450 (M + H). Anal. Calcd for C18H19F4N3O4S: C, 48.10; H, 4.26; N, 9.35. Found: C, 48.27; H, 4.24; N, 9.33.
4.1.3.2. N-((2-Ethoxy-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (9).
Yield 68%, white solid, mp 138–140 °C; 1H NMR (CDCl3) δ 7.58 (d, J = 7.3 Hz, 1H), 7.51 (dd, J = 8.2, 8.2 Hz, 1H), 7.19 (d, J = 7.3 Hz, 1H), 7.12–7.05 (m, 2H), 6.58 (bs, NH), 6.02 (bt, NH), 4.44–4.36 (m, 4H), 3.53 (q, J = 7.0 Hz, 1H), 3.03 (s, 3H), 1.49 (d, J = 7.1 Hz, 3H), 1.34 (t, 3H, J = 7.1 Hz); MS (FAB) m/z 464 (M + H). Anal. Calcd for C19H21F4N3O4S: C, 49.24; H, 4.57; N, 9.07. Found: C, 49.40; H, 4.56; N, 9.04.
4.1.3.3. N-((2-Propoxy-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (10).
Yield 78%, white solid, mp 88–90 °C; 1H NMR (CDCl3) δ 7.58 (d, J = 7.5 Hz, 1H), 7.52 (dd, J = 8.2, 8.2 Hz, 1H), 7.19 (d, J = 7.3 Hz, 1H), 7.12–7.05 (m, 2H), 6.50 (bs, NH), 5.97 (bt, NH), 4.39–4.23 (m, 4H), 3.52 (q, J = 7.1 Hz, 1H), 3.03 (s, 3H), 1.74 (m, 2H), 0.99 (t, J = 7.3 Hz, 3H); MS (FAB) m/z 478 (M + H). Anal. Calcd for C20H23F4N3O4S: C, 50.31; H, 4.86; N, 8.80. Found: C, 50.14; H, 4.88; N, 8.83.
4.1.3.4. N-((2-Butoxy-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (11).
Yield 77%, white solid, mp 77–79 °C; 1H NMR (300 MHz, CDCl3) δ 7.57 (d, J = 7.3 Hz, 1H), 7.52 (dd, J = 8.1, 8.1 Hz, 1H), 7.19 (d, J = 7.5 Hz, 1H), 7.12–7.05 (m, 2H), 6.46 (bs, NH), 5.96 (bt, NH), 4.39–4.28 (m, 4H), 3.51 (q, J = 7.1 Hz, 1H), 3.03 (s, 3H), 1.75–1.66 (m, 2H), 1.49 (d, J = 7.1 Hz, 3H), 1.49–1.37 (m, 2H), 0.98 (t, J = 9.5 Hz, 3H); MS (FAB) m/z 492 (M + H). Anal. Calcd for C21H25F4N3O4S: C, 51.32; H, 5.13; N, 8.55. Found: C, 51.49; H, 5.11; N, 8.52.
4.1.3.5. N-((2-Pentyloxy-6-(trifluoromethyl)pyridin-3-yl) methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (12).
Yield 71%, white solid, mp 58–60 °C; 1H NMR(CDCl3) δ 7.57 (d, J = 7.5 Hz, 1H), 7.52 (dd, J = 8.2, 8.2 Hz, 1H), 7.19 (d, J = 7.4 Hz, 1H), 7.12–7.05 (m, 2H), 6.45 (bs, NH), 5.98 (bt, NH), 4.38–4.29 (m, 4H), 3.51 (q, J = 7.0 Hz, 1H), 3.03 (s, 3H), 1.77–1.67 (m, 2H), 1.49 (d, J = 7.1 Hz, 3H), 1.43–1.35 (m, 4H), 0.93 (t, J = 7.1 Hz, 3H);MS (FAB) m/z 506 (M + H). Anal. Calcd for C22H27F4N3O4S: C, 52.27; H, 5.38; N, 8.31. Found: C, 52.41; H, 5.36; N, 8.28.
4.1.3.6. N-((2-Hexyloxy-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (13).
Yield 90%, white solid, mp 84–86 °C; 1H NMR (CDCl3) δ 7.60 (d, J = 7.3 Hz, 1H), 7.52 (dd, J = 8.3, 8.3 Hz, 1H), 7.21 (d, J = 7.3 Hz, 1H), 7.12–7.06 (m, 2H), 6.50 (bs, NH), 6.00 (bt, NH), 4.37 (d, J = 6.1 Hz, 2h), 3.95 (s, 3H), 3.52 (q, J = 7.1 Hz, 1H), 3.04 (s, 3H), 1.49 (d, J = 7.0 Hz, 3H); MS (FAB) m/z 520 (M + H). Anal. Calcd for C23H29F4N3O4S: C, 53.17; H, 5.63; N, 8.09. Found: C, 53.00; H, 5.64; N, 8.13.
4.1.3.7. (Z)-N-((2-(But-2-en-1-yloxy)-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (14).
Yield 72%, white solid, mp 110 °C; 1H NMR (CDCl3) δ 7.59 (d, J = 8.2 Hz, 1H), 7.52 (dd, J = 8.3 Hz, 1H), 7.20 (d, J = 7.5 Hz, 1H), 7.09 (dd, J = 11.5,1.8 Hz, 1H), 7.06 (d, J = 9.1 Hz, 1H), 6.48 (bs, 1H), 6.01 (bt, 1H), 5.76 (m, 1H), 5.58 (m, 1H), 4.94 (d, J = 6.8 Hz, 2H), 4.37 (d, J = 6.0 Hz, 2H), 3.50 (q, J = 7.0 Hz, 1H), 3.02 (s,3H), 1.77 (d, J = 7.0 Hz, 3H), 1.48 (d, J = 7.1 Hz, 3H); MS (FAB) m/z 490 (M + H). Anal. Calcd for C21H23F4N3O4S: C, 51.53; H, 4.74; N, 8.58. Found: C, 51.36; H, 4.75; N, 8.61.
4.1.3.8. (Z)-N-((2-(Pent-2-en-1-yloxy)-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (15).
Yield 66%, white solid, mp 107 °C; 1H NMR (CDCl3) δ 7.59 (d, J = 7.4 Hz, 1H), 7.51 (dd, J = 8.4 Hz, 1H), 7.20 (d, J = 8.4 Hz, 1H), 7.09 (dd, J = 11.5,1.8 Hz, 1H), 7.06 (d, J = 9.2 Hz, 1H), 6.49 (s, 1H), 6.01 (bt, 1H), 5.67 (m, 1H), 5.52 (m, 1H), 4.92 (d, J = 4.7 Hz, 2H), 4.37 (d, J = 6.4 Hz, 2H), 3.50 (q, J = 6.8 Hz, 1H), 3.03 (s, 3H), 2.20 (m, 2H), 1.48 (d, J = 7.1 Hz, 3H), 1.02 (t, J = 7.5 Hz, 3H); MS (FAB) m/z 504 (M + H). Anal. Calcd for C22H25F4N3O4S: C, 52.48; H, 5.00; N, 8.35. Found: C, 52.69; H, 4.99; N, 8.33.
4.1.3.9. N-((2-(2-Methoxyethoxy)-6-(trifluoromethyl)pyridin-3-yl) methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (16).
Yield 68%, white solid, mp 109 °C; 1H NMR (CDCl3) δ 7.65 (d, J = 7.5 Hz, 1H), 7.51 (t, J = 8.4 Hz, 1H), 7.23 (d, J = 7.2 Hz, 1H), 7.12 (m, 2H), 6.54 (bt, 1H), 6.45 (bs, 1H), 4.56 (m, 2H), 4.41 (t, J = 3.6 Hz, 2H), 3.74 (t, J = 4.5 Hz, 2H), 3.48 (q, J = 7.2 Hz, 1H), 3.43 (s, 3H), 3.01 (s, 3H), 1.47 (d, J = 6.9 Hz, 3H); MS (FAB) m/z 494 (M + H). Anal. Calcd for C20H23F4N3O5S: C, 48.68; H, 4.70; N, 8.52. Found: C, 48.49; H, 4.71; N, 8.55.
4.1.3.10. N-((2-(2-Ethoxyethoxy)-6-(trifluoromethyl)pyridin-3-yl) methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (17).
Yield 70% white solid, mp 118 °C; 1H NMR (CDCl3) δ 7.63 (d, J = 7.6 Hz, 1H), 7.52 (t, J = 8.3 Hz, 1H), 7.24 (d, J = 7.4 Hz, 1H), 7.11 (m, 2H), 6.55 (bt, 1H), 6.46 (bs, 1H), 4.55 (m, 2H), 4.40 (t, J = 3.6 Hz, 2H), 3.73 (t, J = 4.5 Hz, 2H), 3.63 (q, J = 7.2 Hz, 2H), 3.47 (q, J = 7.2 Hz, 1h), 3.44 (s, 3H), 3.01 (s, 3H), 1.42 (d, J = 6.8 Hz, 3H); MS (FAB) m/z 508 (M + H). Anal. Calcd for C21H25F4N3O5S: C, 49.70; H, 4.97; N, 8.28. Found: C, 49.51; H, 4.99; N, 8.31.
4.1.3.11. N-((2-(3-Methoxypropoxy)-6-(trifluoromethyl)pyridin-3-yl) methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (18).
Yield 77%, white solid, mp 112 °C; 1H NMR (CDCl3) δ 7.61 (d, J = 7.3 Hz, 1H), 7.49 (dd, J = 8.1 Hz, 1H), 7.20 (d, J = 7.3 Hz, 1H), 7.02–7.11 (m, 2H), 6.44 (bt, 1H), 4.47–4.50 (m, 2H), 4.34 (d, J = 6.0 Hz, 2H), 3.42–3.61 (m, 3H), 3.36 (s, 3H), 3.03 (s, 3H), 1.89–2.01 (m, 2H), 1.47 (d, J = 7.1 Hz, 3H); MS (FAB) m/z 508 (M + H). Anal. Calcd for C21H25F4N3O5S: C, 49.70; H, 4.97; N, 8.28. Found: C, 49.88; H, 4.96; N, 8.24.
4.1.3.12. N-((2-(3-Ethoxypropoxy)-6-(trifluoromethyl)pyridin-3-yl) methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (19).
Yield 60%, white solid, mp 113 °C; 1H NMR (CDCl3) δ 7.61 (d, J = 7.5 Hz, 1H), 7.50 (dd, J = 8.3 Hz, 1H), 7.20 (d, J = 7.5 Hz, 1H), 7.01–7.11 (m, 2H), 6.35 (bt, H), 4.37–4.50 (m, 2H), 4.35 (d, J = 6.0 Hz, 2H), 3.47–3.60 (m, 5H), 3.03 (s, 3H), 1.90–2.01 (m, 2H), 1.47 (d, J = 7.0 Hz, 3H), 1.20 (t, J = 7.1 Hz, 3H); MS (FAB) m/z 522 (M + H). Anal. Calcd for C22H27F4N3O5S: C, 50.67; H, 5.22; N, 8.06. Found: C, 50.49; H, 5.24; N, 8.09.
4.1.3.13. N-((2-(2-Phenoxyethoxy)-6-(trifluoromethyl)pyridin-3-yl) methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (20).
Yield 68%, white solid, mp 70 °C; 1H NMR (CDCl3) δ 7.66 (d, J = 7.3 Hz, 1H), 7.41 (dd, J = 8.4, 8.4 Hz, 1H), 7.22–7.35 (m, 3H), 6.88–7.05(m, 5H), 6.42 (bs, 1H), 6.21 (bt, 1H), 4.63–4.82 (m, 2H), 4.27–4.42 (m, 4H), 3.34 (q, J = 7.1 Hz, 1H), 2.99 (s, 3H), 1.38 (d, J = 7.0 Hz, 3H); MS (FAB) m/z 556 (M + H). Anal. Calcd for C25H25F4N3O5S: C, 54.05; H, 4.54; N, 7.56. Found: C, 54.23; H, 4.52; N, 7.53.
4.1.3.14. N-((2-Isopropoxy-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (21).
Yield 81%, white solid, mp 66–68 °C; 1H NMR (CDCl3) δ 7.56 (d, J = 7.5 Hz, 1H), 7.50 (dd, J = 8.1 Hz, 1H), 7.16 (d, J = 7.5 Hz, 1H), 7.12–7.05 (m, 2H), 6.70 (bs, NH), 6.05 (bt, NH), 5.36 (m, 1H), 3.53 (q, J = 7.1 Hz, 1H), 3.03 (s, 3H), 1.49 (d, J = 7.1 Hz, 3H), 1.30 (t, 6H); MS (FAB) m/z 478 (M + H). Anal. Calcd for C20H23F4N3O4S: C, 50.31; H, 4.86; N, 8.80. Found: C, 50.49; H, 4.84; N, 8.77.
4.1.3.15. N-((2-Isobutoxy-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (22).
Yield 84%, white solid, mp 62–64 °C; 1H NMR (CDCl3) δ 7.60 (d, J = 7.3 Hz, 1H), 7.52 (dd, J = 8.3, 8.3 Hz, 1H), 7.21 (d, J = 7.3 Hz, 1H), 7.12–7.06 (m, 2H), 6.50 (bs, NH), 6.00 (bt, NH), 4.37 (d, J = 6.1 Hz, 2h), 3.95 (s, 3H), 3.52 (q, J = 7.1 Hz, 1H), 3.04 (s, 3H), 1.49 (d, J = 7.0 Hz, 3H); MS (FAB) m/z 492 (M + H). Anal. Calcd for C21H25F4N3O4S: C, 51.32; H, 5.13; N, 8.55. Found: C, 51.15; H, 5.15; N, 8.58.
4.1.3.16. N-((2-Isopentyloxy-6-(trifluoromethyl)pyridin-3-yl) methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (23).
Yield 68%, white solid, mp 136–139 °C; 1H NMR (CD3OD) δ 7.40–7.49 (m, 2H), 7.14–7.23 (m, 3H), 4.38 (t, J = 7.2 Hz, 2H), 4.31 (d, J = 7.5 Hz, 2H), 3.71 (q, J = 6.9 Hz, 1H), 2.97 (s, 3H), 1.76 (m, 1H), 1.64 (q, J = 6.6 Hz, 2H), 1.45 (d, J = 6.9 Hz, 3H), 0.95 (d, J = 6.6 Hz, 6H); MS (FAB) m/z 506 (M + H). Anal. Calcd for C22H27F4N3O4S: C, 52.27; H, 5.38; N, 8.31. Found: C, 52.06; H, 5.40; N, 8.35.
4.1.3.17. N-((2-(3,3-Dimethylbutoxy)-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (24).
Yield 70%, white solid, mp 64–66 °C; 1H NMR (CD3OD) δ 7.40–7.48 (m, 2H), 7.13–7.22 (m, 3H), 4.42 (t, J = 7.5 Hz, 2H), 4.31 (d, J = 7.2 Hz, 2H), 3.72 (q, J = 6.9 Hz, 1H), 2.97 (s, 3H), 1.68 (t, J = 7.2 Hz, 2H), 1.45 (d, J = 6.9 Hz, 3H), 0.97 (s, 9H); MS (FAB) m/z 520 (M + H). Anal. Calcd for C23H29F4N3O4S: C, 53.17; H, 5.63; N, 8.09. Found: C, 53.30; H, 5.62; N, 8.06.
4.1.3.18. N-((2-(Pentan-3-yloxy)-6-(trifluoromethyl)pyridin-3-yl) methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (25).
Yield 65%, colorless oil; 1H NMR (CDCl3) δ 7.55 (d, J = 7.5 Hz, 1H), 7.49 (t, J = 8.4 Hz, 1H), 7.12 (m, 3H), 6.25 (t, J = 5.7 Hz, 1H), 5.20 (m, 1H), 4.42 (m, 2H), 3.58 (q, J = 6.9 Hz, 1H), 3.01 (s, 3H), 1.70 (m, 4H), 1.49 (d, J = 6.9 Hz, 3H), 0.90 (m, 6H); MS (FAB) m/z 506 (M + H). Anal. Calcd for C22H27F4N3O4S: C, 52.27; H, 5.38; N, 8.31. Found: C, 52.09; H, 5.40; N, 8.34.
4.1.3.19. N-((2-(Heptan-4-yloxy)-6-(trifluoromethyl)pyridin-3-yl) methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (26).
Yield 68%, colorless oil; 1H NMR (CDCl3) δ 7.56 (d, J = 7.4 Hz, 1H), 7.48 (t, J = 8.5 Hz, 1H), 7.11 (m, 3H), 6.24 (t, J = 5.8 Hz, 1H), 5.21 (m, 1H), 4.43 (m, 2H), 3.59 (q, J = 6.8 Hz, 1H), 3.02 (s, 3H), 1.71 (m, 4H), 1.48 (d, J = 6.9 Hz, 3H), 1.25 (m, 4H), 0.92 (m, 6H); MS (FAB) m/z 534 (M + H). Anal. Calcd for C24H31F4N3O4S: C, 54.02; H, 5.86; N, 7.88. Found: C, 54.22; H, 5.85; N, 7.85.
4.1.3.20. N-((2-(Nonan-5-yloxy)-6-(trifluoromethyl)pyridin-3-yl) methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (27).
Yield 57%, pale yellow oil; 1HNMR(CDCl3) δ 7.49–7.56(m,2H), 7.15 (d, J = 7.7 Hz, 1H), 7.08 (t, J = 5.9 Hz, 1H), 6.47 (bs, 1H), 5.98 (bt, 1H), 5.29 (m, 1H), 4.37 (m, 2H), 3.49 (q, J = 7.0 Hz, 1H), 3.03 (s, 3H), 1.57 (m, 2H), 1.49 (d, J = 7.0 Hz, 3H), 1.24–1.31 (m, 8H), 0.88–0.90 (m, 6H); MS (FAB) m/z 562 (M + H). Anal. Calcd for C26H35F4N3O4S: C, 55.60; H, 6.28; N, 7.48. Found: C, 55.78; H, 6.27; N, 7.45.
4.1.3.21. N-((2-(2,6-Dimethylheptan-4-yloxy)-6-(trifluoromethyl) pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl) propanamide (28).
Yield 67%, colorless oil; 1H NMR (CDCl3) δ 7.57 (d, J = 7.5 Hz, 1H), 7.49 (t, J = 8.6 Hz, 1H), 7.12 (m, 3H), 6.25 (t, J = 5.8 Hz, 1H), 5.22 (m, 1H), 4.44 (m, 2H), 3.57 (q, J = 6.9 Hz, 1H), 3.03 (s, 3H), 1.71 (m, 4H), 1.49 (d, J = 6.8 Hz, 3H), 1.24 (m, 2H), 0.92 (m, 12H); MS (FAB) m/z 562 (M + H). Anal. Calcd for C26H35F4N3O4S: C, 55.60; H, 6.28; N, 7.48. Found: C, 55.78; H, 6.26; N, 7.44.
4.1.3.22. N-((2-Cyclobutoxy-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide(29).
Yield 80%, white solid, mp 76–78 °C; 1HNMR(CDCl3) δ 7.58–7.51 (m, 2H), 7.18(d,J = 7.3 Hz, 1H), 7.13 –7.07 (m, 2H), 6.50(bs, NH), 6.00(bt, NH), 5.20 (m,1H), 4.37 (d, J = 6.2 Hz, 2H), 3.56 (q, J = 7.0 Hz, 1H), 3.03 (s, 3H), 2.50–2.40 (m, 2H), 2.05–1.65 (m, 4H), 1.50 (d, J = 7.1 Hz, 3H); MS (FAB) m/z 490 (M + H). Anal. Calcd for C21H23F4N3O4S: C, 51.53; H, 4.74; N, 8.58. Found: C, 51.35; H, 4.76; N, 8.61.
4.1.3.23. N-((2-Cyclopentyloxy-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (30).
Yield 79%, white solid, mp 67–69 °C; 1H NMR (CDCl3) δ 7.56 (d, J = 7.5 Hz, 1H), 7.50 (dd, J = 8.1, 8.1 Hz, 1H), 7.16 (d, J = 7.5 Hz, 1H), 7.12–7.05 (m, 2H), 6.70 (bs, NH), 6.05 (bt, NH), 5.36 (m, 1H), 3.53 (q, J = 7.1 Hz, 1H), 3.03 (s, 3H), 1.49 (d, J = 7.1 Hz, 3H), 1.30 (t, 6H); MS (FAB) m/z 504 (M + H). Anal. Calcd for C22H25F4N3O4S: C, 52.48; H, 5.00; N, 8.35. Found: C, 52.67; H, 4.98; N, 8.33.
4.1.3.24. N-((2-Cyclohexyloxy-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (31).
Yield 80%, white solid, mp 81–83 °C; 1H NMR (CDCl3) δ 7.56 (d, J = 7.5 Hz, 1H), 7.50 (dd, J = 8.1 Hz, 1H), 7.16 (d, J = 7.5 Hz, 1H), 7.12–7.05 (m, 2H), 6.70 (bs, NH), 6.05 (bt, NH), 5.36 (m,1H), 3.53 (q, J = 7.1 Hz, 1H), 3.03 (s, 3H), 1.49 (d, J = 7.1 Hz, 3H), 1.30 (t, 6H); MS (FAB) m/z 518 (M + H). Anal. Calcd for C23H27F4N3O4S: C, 53.38; H, 5.26; N, 8.12. Found: C, 53.59; H, 5.23; N, 8.07.
4.1.3.25. N-((6-Trifluoromethyl-2-(4-(trifluoromethyl)cyclohexyloxy) pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (32).
Yield 30%, white solid, mp 66–68 °C; 1H NMR (CDCl3) δ 7.59 (d, J = 7.0 Hz, 1H), 7.51 (dd, J = 8.2 Hz, 1H), 7.20 (d, J = 7.5 Hz, 1H), 7.13–7.05 (m, 2H), 6.50 (bs, NH), 5.91 (bt, NH), 5.43 (m, 1H), 4.39 (m, 2H), 3.51 (q, J = 6.6 Hz, 1H), 3.03 (s, 3H), 2.20–2.08 (m, 3H), 1.85–1.77 (m, 2H), 1.63–1.50 (m, 4H), 1.49 (d, J = 7.1 Hz, 3H); MS (FAB) m/z 586 (M + H). Anal. Calcd for C24H26F7N3O4S: C, 49.23; H, 4.48; N, 7.18. Found: C, 49.50; H, 4.51; N, 7.14.
4.1.3.26. N-((2-(4-Methylcyclohexyloxy)-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl )propanamide (33).
Yield 78%, white solid, mp 107–109 °C; 1H NMR (CDCl3) δ 7.57–7.49 (m, 2H), 7.16 (d, J = 7.3 Hz, 1H), 7.12–7.05 (m, 2H), 6.48 (bs, NH), 5.99 (bt, NH), 5.00 (m,1H), 4.34 (d, J = 5.8 Hz, 2H), 3.51 (q, J = 6.8 Hz,1H), 3.03 (s, 3H), 2.12–2.00 (m, 2H), 1.80–1.72 (m, 2H), 1.50–1.10(m, 5H), 1.48 (d, J = 7.1 Hz, 3H), 0.94 (d, J = 6.6 Hz, 3H); MS (FAB) m/z 532 (M + H). Anal. Calcd for C24H29F4N3O4S: C, 54.23; H, 5.50; N, 7.91. Found: C, 54.40; H, 5.51; N, 7.95.
4.1.3.27. N-((2-(4-Ethylcyclohexyloxy)-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (34).
Yield 65%, white solid, mp 123–125 °C; 1H NMR (CDCl3) δ 7.57–7.50 (m, 2H), 7.16 (d, J = 7.3 Hz, 1H), 7.12–7.05 (m, 2H), 6.47 (bs, NH), 5.99 (bt, NH), 5.00 (m, 1H), 4.34 (m, 2H), 3.52 (q, J = 7.5 Hz, 1H), 3.03 (s, 3H), 2.13–2.03 (m, 2H), 1.87–1.80 (m, 2H), 1.49 (d, J = 7.1 Hz, 3H), 1.32–1.04 (m, 7H), 0.91 (t, J = 7.1 Hz, 3H); MS (FAB) m/z 546 (M + H). Anal. Calcd for C25H31F4N3O4S: C, 55.04; H, 5.73; N, 7.70. Found: C, 55.20; H, 5.72; N, 7.68.
4.1.3.28. N-((2-(4-tert-Butylcyclohexyloxy)-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (35).
Yield 69%, white solid, mp 83–85 °C; 1H NMR (CDCl3) δ 7.57–7.50 (m, 2H), 7.16 (d, J = 7.3 Hz, 1H), 7.12–7.05 (m, 2H), 6.46 (bs, NH), 5.98 (bt, NH), 4.96 (m, 1H), 4.34 (m, 2H), 3.51 (q, J = 7.1 Hz, 1H), 3.03 (s, 3H), 2.20–2.10 (m, 2H), 1.88–1.80 (m, 2H), 1.49 (d, J = 7.1 Hz, 3h), 1.30–1.00 (m, 5H), 0.89 (s, 9H); MS (FAB) m/z 574 (M + H). Anal. Calcd for C27H35F4N3O4S: C, 56.53; H, 6.15; N, 7.33 Found: C, 56.38; H, 6.17; N, 7.35.
4.1.3.29. N-((2-(4,4-Dimethylcyclohexyloxy)-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (36).
Yield 63%, white solid, mp 86 °C; 1H NMR (CDCl3) δ 7.56–7.50 (m, 2H), 7.17 (d, J = 7.2 Hz, 1H), 7.11–7.05 (m, 2H), 6.44 (bs, NH), 5.97 (bt, NH), 4.96 (m, 1H), 4.33 (m, 2H), 3.52 (q, J = 7.2 Hz, 1H), 3.03 (s, 3H), 2.20–2.10 (m, 2H), 1.88–1.80 (m, 2H), 1.49 (d, J = 7.1 Hz, 3H), 1.30–1.00 (m, 4H), 0.96 (d, J = 6.3 Hz, 6H); MS (FAB) m/z 546 (M + H). Anal. Calcd for C25H31F4N3O4S: C, 55.04; H, 5.73; N, 7.70. Found: C, 54.90; H, 5.74; N, 7.73.
4.1.3.30. N-((2-(3,5-Dimethylcyclohexyloxy)-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl) propanamide (37).
Yield 66%, white solid, mp 84 °C; 1H NMR (CDCl3) δ 7.55–7.51 (m, 2H), 7.18 (d, J = 7.4 Hz, 1H), 7.10–7.06 (m, 2H), 6.42 (bs, NH), 5.98 (bt, NH), 4.95 (m, 1H), 4.34 (m, 2H), 3.53 (q, J = 7.2 Hz, 1H), 3.03 (s, 3H), 2.22–2.10 (m, 2H), 1.88–1.80 (m, 2H), 1.48 (d, J = 7.1 Hz, 3H), 1.29 (m, 1H), 1.28–1.00 (m, 3H), 0.96 (d, J = 6.3 Hz, 6H); MS (FAB) m/z 546 (M + H). Anal. Calcd for C25H31F4N3O4S: C, 55.04; H, 5.73; N, 7.70. Found: C, 54.89; H, 5.74; N, 7.73.
4.1.3.31. N-((2-(Piperidin-4-yloxy)-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (38).
Yield 69%, white solid, mp 80–82 °C; 1H NMR (CDCl3) δ 7.58 (d, J = 7.5 Hz, 1H), 7.41 (dd, J = 8.3 Hz, 1H), 7.27 (d, J = 7.5 Hz, 1H), 7.19–7.11 (m, 2H), 5.29 (m,1H), 4.36 (m, 2H), 3.71 (q, J = 7.0 Hz,1H), 3.20 (m, 2H), 3.01–2.90 (m, 2H), 2.97 (s, 3H), 2.06 (m, 2H), 1.81 (m, 2H), 1.45 (d, J = 7.1 Hz, 3H); MS (FAB) m/z 519 (M + H). Anal. Calcd for C22H26F4N4O4S: C, 50.96; H, 5.05; N, 10.81. Found: C, 50.78; H, 5.07; N, 10.84.
4.1.3.32. tert-Butyl 4-(3-((2-(3-fluoro-4-(methylsulfonamido)phenyl) propanamido)methyl )-6-(trifluoromethyl)pyridin-2-yloxy)piperidine-1-carboxylate (39).
Yield 75%, white solid, mp 82–84 °C; 1H NMR (CDCl3) δ 7.60 (d, J = 7.3 Hz, 1H), 7.50 (dd, J = 8.2 Hz, 1H), 7.20 (d, J = 7.3 Hz, 1H), 7.13–7.04 (m, 2H), 5.87 (bt, NH), 5.24 (m, 1H), 4.36 (d, 2H), 3.70–3.62 (m, 2H), 3.54 (q, J = 7.7 Hz, 1H), 3.28–3.17 (m, 2H), 3.04 (s, 3H), 1.98–1.88 (m, 2H), 1.54–1.40 (m, 2H), 1.51 (d, 3H), 1.50 (s, 9H); MS (FAB) m/z 619 (M + H). Anal. Calcd for C27H34F4N4O6S: C, 52.42; H, 5.54; N, 9.06. Found: C, 52.59; H, 5.52; N, 9.03.
4.1.3.33. N-((2-(2,3-Dihydro-1H-inden-2-yloxy)-6-(trifluoromethyl) pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl) propanamide (40).
Yield 83%, white solid, mp 135 °C; 1H NMR (CDCl3) δ 7.59 (d, J = 7.3 Hz, 1H), 7.46 (dd, J = 8.2 Hz, 1H), 7.29–7.24 (m, 5H), 6.99 (dd, J = 2.0 Hz, 1H), 6.91 (d, J = 8.8 Hz, 1H), 6.35 (bs, 1H), 5.89 (m, 1H), 5.79 (bt, 1H), 4.27 (m, 2H), 3.43 (dd, J = 5.5 Hz, 2H), 3.08–3.04 (m, 3H), 3.00 (s, 3H), 1.31 (d, J = 7.1 Hz, 3H); MS (FAB) m/z 552 (M + H). Anal. Calcd for C26H25F4N3O4S: C, 56.62; H, 4.57; N, 7.62. Found: C, 56.79; H, 4.56; N, 7.59.
4.1.3.34. N-((2-(Cyclopropylmethoxy)-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (41).
Yield 81%, white solid, mp 89–91 °C; 1H NMR (CDCl3) δ 7.60 (d, J = 7.3 Hz, 1H), 7.52 (dd, J = 8.3 Hz, 1H), 7.21 (d, J = 7.3 Hz, 1H), 7.12–7.06 (m, 2H), 6.50 (bs, NH), 6.00 (bt, NH), 4.37 (d, J = 6.1 Hz, 2H), 3.95 (s, 3H), 3.52 (q, J = 7.1 Hz, 1H), 3.04 (s, 3H), 1.49 (d, J = 7.0 Hz, 3H); MS (FAB) m/z 490 (M + H). Anal. Calcd for C21H23F4N3O4S: C, 51.53; H, 4.74; N, 8.58. Found: C, 51.70; H, 4.73; N, 8.55.
4.1.3.35. N-((2-((2-Methylcyclopropyl)methoxy)-6-(trifluoromethyl) pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl) propanamide (42).
Yield 75%, white solid, mp 96–97 °C; 1H NMR (CD3OD) 5 7.48 (d, J = 7.5 Hz, 1H), 7.43 (dd, J = 8.1 Hz, 1H), 7.15–7.23 (m, 3H), 4.34 (d, J = 5.1 Hz, 2H), 4.20 (d, J = 7.1 Hz, 2H), 3.73 (q, J = 6.9 Hz, 1H), 2.98 (s, 3H), 1.46 (d, J = 7.1 Hz, 3H), 1.04 (d, J = 6.0 Hz, 3H), 0.95 (m, 1H), 0.78 (m,1H), 0.51 (m,1H), 0.31 (m,1H); MS (FAB) m/z 504 (M + h). Anal. Calcd for C22H25F4N3O4S: C, 52.48; H, 5.00; N, 8.35. Found: C, 52.63; H, 5.01; N, 8.37.
4.1.3.36. N-((2-((2,2-Dimethylcyclopropyl)methoxy)-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fl uoro-4-(methylsulfonamido)phenyl)propanamide (43).
Yield 72%, white solid, mp 63–65 °C; 1H NMR (CDCl3) δ 7.58 (d, J = 7.3 Hz, 1H), 7.51 (dd, J = 8.4 Hz, 1H), 7.19 (d, J = 7.3 Hz, 1H), 7.14–7.05 (m, 2H), 6.54 (bs, NH), 6.07 (bt, NH), 4.57–4.33 (m, 3H), 4.24–4.15 (m, 1H), 3.51 (q, J = 7.1 Hz, 1H), 3.03 (s, 3H), 1.49 (dd, J = 1.7 Hz, 3H), 1.13 (d, J = 1.5 Hz, 3H), 1.09 (s, 3H), 1.06–0.95 (m, 1H), 0.57 (dd, J = 8.4, 4.4 Hz, 1H), 0.28 (m,1H); MS (FAB) m/z 518 (M + H). Anal. Calcd for C23H27F4N3O4S: C, 53.38; H, 5.26; N, 8.12. Found: C, 53.55; H, 5.24; N, 8.10.
4.1.3.37. N-((2-(Cyclobutylmethoxy)-6-(trifluoromethyl)pyridin-3-yl) methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (44).
Yield 79%, white solid, mp 100–102 °C; 1H NMR(CDCl3) δ 7.57 (d, J = 7.5 Hz, 1H), 7.50 (dd, J = 8.2, 8.2 Hz, 1H), 7.19 (d, J = 7.3 Hz, 1H), 7.12–7.04 (m, 2H), 6.64 (bs, NH), 6.02 (bt, NH), 4.45–4.26 (m, 4H), 3.51 (q, J = 7.1 Hz, 1H), 3.03 (s, 3H), 2.71 (m, 1H), 2.13–1.79 (m, 6h), 1.48 (d, J = 7.1 Hz, 3H); MS (FAB) m/z 504 (M + H). Anal. Calcd for C22H25F4N3O4S: C, 52.48; H, 5.00; N, 8.35. Found: C, 52.67; H, 4.99; N, 8.32.
4.1.3.38. N-((2-(Cyclopentylmethoxy)-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (45).
Yield 70%, white solid, mp 112–114 °C; 1H NMR (CDCl3) δ 7.58–7.51 (m, 2H), 7.18 (d, J = 7.3 Hz, 1H), 7.13–7.07 (m, 2H), 6.50 (bs, NH), 6.00 (bt, NH), 5.20 (m, 1H), 4.37 (d, J = 6.2 Hz, 2H), 3.56 (q, J = 7.0 Hz, 1H), 3.03 (s, 3H), 2.50–2.40 (m, 2H), 2.05–1.65 (m, 4h), 1.50 (d, J = 7.1 Hz, 3H); MS (FAB) m/z 518 (M + H). Anal. Calcd for C23H27F4N3O4S: C, 53.38; H, 5.26; N, 8.12. Found: C, 53.55; H, 5.24; N, 8.09.
4.1.3.39. N-((2-(Cyclohexylmethoxy)-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (46).
Yield 73%, white solid, mp 108–110 °C; 1H NMR(CDCl3) δ 7.56 (d, J = 7.3 Hz, 1H), 7.51 (dd, J = 8.3, 8.3 Hz, 1H), 7.18 (d, J = 7.3 Hz, 1H), 7.12–7.04 (m, 2H), 6.57 (bs, NH), 5.99 (bt, NH), 4.38 (m, 2H), 4.16 (m, 2H), 3.51 (q, J = 7.1 Hz, 1H), 3.03 (s, 3H), 1.82–1.67 (m, 5H), 1.49 (d, J = 7.1 Hz, 3H), 1.32–1.00 (m, 6H); MS (FAB) m/z 532 (M + H). Anal. Calcd for C24H29F4N3O4S: C, 54.23; H, 5.50; N, 7.91. Found: C, 54.05; H, 5.51; N, 7.94.
4.1.3.40. N-((2-((4-Methylcyclohexyl)methoxy)-6-(trifluoromethyl) pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (47).
Yield 73%, white solid, mp 71–73 °C; 1H NMR (CDCl3) 5 7.56 (d, J = 7.3 Hz, 1H), 7.50 (dd,J = 8.2, 8.2 Hz, 1H), 7.18 (d, J = 7.5 Hz, 1H), 7.12–7.04 (m, 2H), 6.59 (bs, NH), 6.00 (bt, NH), 4.45–4.11 (m, 4H), 3.51 (q, J = 7.1 Hz, 1H), 3.03 (s, 3H), 1.95–1.25 (m, 12H), 1.10–0.90 (m, 4H); MS (FAB) m/z 546 (M + H). Anal. Calcd for C25H31F4N3O4S: C, 55.04; H, 5.73; N, 7.70. Found: C, 54.89; H, 5.75; N, 7.74.
4.1.3.41. N-((2-(Piperidin-4-ylmethoxy)-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (48).
Yield 66%, white solid, mp 52–54 °C; 1H NMR (CD3OD) δ 7.58 (d, J = 7.9 Hz, 1H), 7.43 (dd, J = 8.3, 8.3 Hz, 1H), 7.29 (d, J = 7.4 Hz, 1H), 7.22–7.15 (m, 2H), 4.47–4.23 (m, 4H), 3.73 (q, J = 7.1 Hz, 1H), 3.43–3.36 (m, 2H), 3.05–2.93 (m, 2H), 3.00 (s, 3H), 2.04–1.96 (m, 3H), 1.53–1.45 (m, 2H), 1.46 (d, J = 7.1 Hz, 3H); MS (FAB) m/z 533 (M + H). Anal. Calcd for C23H28F4N4O4S: C, 51.87; H, 5.30; N, 10.52. Found: C, 51.69; H, 5.32; N, 10.55.
4.1.3.42. tert-Butyl 4-((3-((2-(3-fluoro-4-(methylsulfonamido) phenyl)propanamido)methyl)-6-(trifluoromethyl)pyridin-2-yloxy)methyl)piperidine-1-carboxylate (49).
Yield 52%, white solid, mp 59–61 °C; 1H NMR (CDCl3) δ 7.60 (d, J = 7.5 Hz, 1H), 7.52 (dd, J = 8.4, 8.4 Hz, 1H), 7.21 (d, J = 7.5 Hz, 1H), 7.12–7.05 (m, 2H), 5.83 (bs, NH), 4.37 (d, J = 5.9 Hz, 2H), 4.25–4.07 (m, 4H), 3.53 (q, J = 6.4 Hz, 1h), 3.4 (s, 3H), 2.78–2.63 (m, 2H), 1.90 (m,1H), 1.68–1.55 (m, 2H), 1.48 (s, 9H), 1.25–1.05 (m, 2H); MS (fAB) m/z 633 (M + H). Anal. Calcd for C28H36F4N4O6S: C, 53.16; H, 5.74; N, 8.86. Found: C, 52.96; H, 5.76; N, 8.90.
4.1.3.43. tert-Butyl 2-(2-(3-((2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamido)methyl)-6-(trifluoromethyl)pyridin-2-yloxy)ethyl)piperidine-1-carboxylate (50).
Yield 64%, white solid, mp 60 °C; 1H NMR (CDCl3) δ 7.62 (d, J = 7.9 Hz, 1H), 7.50 (dd, J = 8.3, 8.4 Hz, 1H), 7.21 (d, J = 7.7 Hz, 1H), 7.15–7.05 (m, 2H), 5.85 (bs, NH), 4.36 (d, J = 5.6 Hz, 2H), 4.22–4.05 (m, 4H), 3.50 (q, J = 6.4 Hz, 1H), 3.03 (s, 3H), 2.79–2.63 (m, 2H), 1.92 (m, 2H), 1.65–1.55 (m, 2H), 1.44 (s, 9H), 1.22–1.05 (m, 2H); MS (FAB) m/z 647 (M + H). Anal. Calcd for C29H38F4N4O6S: C, 53.86; H, 5.92; N, 8.66. Found: C, 53.69; H, 5.94; N, 8.69.
4.1.3.44. N-((2-Phenoxy-6-(trifluoromethyl)pyridin-3-yl)methyl)-2- (3-fluoro-4-(methylsulfonamido)phenyl)propanamide (51).
Yield 80%, white solid, mp 95–97 °C; 1H NMR (CD3OD) δ 7.75 (d, J = 7.2 Hz, 1H), 7.00–7.49 (m, J = 7.8 Hz, 9H), 6.26 (bt, 1H), 4.51 (d, J = 5.7 Hz, 2H), 3.56 (q, J = 6.9 Hz, 1H), 3.01 (s, 3H), 1.48 (d, J = 6.9 Hz, 3H); MS (fAB) m/z 512 (M + H). Anal. Calcd for C23H21F4N3O4S: C, 54.01; H, 4.14; N, 8.22. Found: C, 53.83; H, 4.16; N, 8.25.
4.1.3.45. N-((2-(p-Tolyloxy)-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (52).
Yield 78%, white solid, mp 167 °C; 1H NMR (CDCl3) δ 7.74 (d, J = 7.5 Hz, 1H), 7.47 (dd, J = 8.4, 8.4 Hz, 1H), 7.31 (d, J = 7.9 Hz, 1H), 7.18 (d, J = 8.8 Hz, 2H), 7.01 (m, 2H), 6.91 (m, 2H), 4.49 (m, 2H), 3.58 (q, J = 7.0 Hz, 1H), 2.94 (s, 3H), 1.49 (d, J = 7.1 Hz, 3H); MS (FAB) m/z 526 (M + H). Anal. Calcd for C24H23F4N3O4S: C, 54.85; H, 4.41; N, 8.00. Found: C, 54.67; H, 4.43; N, 8.03.
4.1.3.46. N-((2-Benzyloxy-6-(trifluoromethyl)pyridin-3-yl)methyl)-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (53).
Yield 71%, white solid, mp 115 °C; 1H NMR (CDCl3) δ 7.62 (d, J = 7.1 Hz, 1H), 7.47 (dd, J = 8.4, 8.4 Hz, 1H), 7.44–7.36 (m, 5H), 7.24 (d, J = 7.5 Hz, 1H), 7.04 (dd, J = 11.2,1.8 Hz, 1H), 6.97 (d, J = 8.4 Hz, 1H), 6.42 (bs, 1H), 5.96 (bt, 1H), 5.41 (m, 2H), 4.39 (m, 2H), 3.41 (q, J = 7.1 Hz, 1H), 3.01 (s, 3H), 1.42 (d, J = 7.1 Hz, 3H); MS (FAB) m/z 526 (M + H). Anal. Calcd for C24H23F4N3O4S: C, 54.85; H, 4.41; N, 8.00. Found: C, 54.66; H, 4.43; N, 8.03.
4.1.3.47. N-((2-Phenethoxy-6-(trifluoromethyl)pyridin-3-yl)methyl)-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (54).
Yield 78%, white solid, mp 98 °C; 1H NMR (CDCl3) δ 7.60 (d, J = 7.4 Hz, 1H), 7.46 (dd, J = 8.3, 8.3 Hz, 1H), 7.33 (m, 5H), 7.19 (d, J = 7.3 Hz, 1H), 6.97 (dd, J = 11.3,1.8 Hz, 1H), 6.89 (d, J = 8.9 Hz, 1H), 6.43 (bs, 1H), 5.70 (bt, 1H), 4.66 (m, 1H), 4.50 (m, 1H), 4.28 (d, J = 6.2 Hz, 2H), 3.14–3.05 (m, 3H), 2.99 (s, 3H), 1.36 (d, J = 7.1 Hz, 3H); MS (FAB) m/z 540 (M + H). Anal. Calcd for C25H25F4N3O4S: C, 55.65; H, 4.67; N, 7.79. Found: C, 55.83; H, 4.65; N, 7.75.
4.1.3.48. N-((2-(4-Methylbenzyloxy)-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (55).
Yield 68%, white solid, mp 117–119 °C; 1H NMR(CDCl3) δ 7.61 (d, J = 7.4 Hz, 1H), 7.47 (dd, J = 8.1, 8.1 Hz, 1H), 7.31 (d, J = 8.0 Hz, 2H), 7.22 (d,J = 7.2 Hz, 1H), 7.20 (d, J = 7.9 Hz, 2H), 7.03 (dd,J = 11.5, 1.9 Hz, 1H), 6.96 (d, J = 8.6 Hz, 1H), 6.46 (bs, 1H), 5.98 (bt, 1H), 5.36 (m, 2H), 4.37 (m, 2H), 3.40 (q, J = 7.1 Hz, 1H), 3.01 (s, 3H), 2.28 (s, 3H), 1.42 (d, J = 7.0 Hz, 3H); MS (FAB) m/z 540 (M + H). Anal. Calcd for C25H25F4N3O4S: C, 55.65; H, 4.67; N, 7.79. Found: C, 55.84; H, 4.65; N, 7.76.
4.1.3.49. N-((2-(4-Ethylbenzyloxy)-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (56).
Yield 78%, white solid, mp 137 °C; 1H NMR (CDCl3) δ 7.61 (d, J = 7.7 Hz, 1H), 7.47 (dd, J = 8.3, 8.3 Hz, 1H), 7.34 (d, J = 8.1 Hz, 2H), 7.23 (d, J = 7.7 Hz, 3H), 7.03 (dd, J = 11.2, 2.1 Hz, 1H), 6.84 (d, J = 8.6 Hz, 1H), 6.40 (bs, 1H), 5.96 (bt, 1H), 5.38 (m, 2H), 4.37 (m, 2H), 3.41 (q, J = 7.1 Hz, 1H), 3.01 (s, 3H), 2.68 (q, J = 7.5 Hz, 2H), 1.41 (d, J = 7.1 Hz, 3H), 1.25 (t, J = 7.6 Hz, 3H); MS (FAB) m/z 554 (M + H). Anal. Calcd for C26H27F4N3O4S: C, 56.41; H, 4.92; N, 7.59. Found: C, 56.61; H, 4.90; N, 7.55.
4.1.3.50. N-((2-(4-Butylbenzyloxy)-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (57).
Yield 72%, white solid, mp 131 °C; 1H NMR (CDCl3) δ 7.61 (d, J = 7.5 Hz, 1H), 7.48 (dd, J = 8.2, 8.2 Hz, 1H), 7.33 (d, J = 8.1 Hz, 2H), 7.23 (d, J = 7.3 Hz, 1H), 7.10 (d, J = 7.9 Hz, 2H), 7.04 (dd, J = 11.2, 2.0 Hz, 1H), 6.97 (d, J = 8.6 Hz, 1H), 6.41 (bs, 1H), 5.94 (bt, 1H), 5.38 (m, 2H), 4.37 (m, 2H), 3.39 (q, J = 7.0 Hz, 1H), 3.01 (s, 3H), 2.63 (t, J = 7.8 Hz, 2H), 1.61 (m, 2H), 1.41 (d, J = 7.1 Hz, 3H), 1.38 (m, 2H), 0.93 (t, J = 7.3 Hz, 3H); MS (FAB) m/z 582 (M + H). Anal. Calcd for C28H31F4N3O4S: C, 57.82; H, 5.37; N, 7.22. Found: C, 57.98; H, 5.38; N, 7.20.
4.1.3.51. N-((2-(4-tert-Butylbenzyloxy)-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (58).
Yield 78%, white solid, mp 105–107 °C; 1H NMR(CDCl3) δ 7.61 (d, J = 7.1 Hz, 1H), 7.48 (dd, J = 8.3, 8.3 Hz, 1H), 7.45–7.37 (m, 4H), 7.23 (d, J = 7.3 Hz, 1H), 7.04 (dd, J = 11.2, 1.9 Hz, 1H), 6.97 (d, J = 8.1 Hz, 1H), 6.42 (bs, 1H), 5.98 (bt, 1H), 5.39 (m, 2H), 4.38 (m, 2H), 3.40 (q, J = 7.3 Hz), 3.00 (s, 3H), 1.41 (d, J = 7.1 Hz, 1H), 1.34 (s, 9H); MS (FAB) m/z 582 (M + H). Anal. Calcd for C28H31F4N3O4S: C, 57.82; H, 5.37; N, 7.22. Found: C, 57.99; H, 5.38; N, 7.19.
4.1.3.52. N-((2-(4-Fluorobenzyloxy)-6-(trifluoromethyl)pyridin-3-yl) methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (59).
Yield 68%, white solid, mp 142 °C; 1H NMR (CDCl3) δ 7.59 (d, J = 7.3 Hz, 1H), 7.48 (dd, J = 8.3, 8.3 Hz, 1H), 7.41 (m, 2H), 7.23 (d, J = 7.5 Hz, 1H), 7.06 (m, 3H), 6.99 (d, J = 8.0 Hz, 1H), 6.51 (bs, 1H), 5.93 (bt, 1H), 5.37 (m, 2H), 4.38 (m, 2H), 3.45 (q, J = 7.1 Hz, 1H), 3.02 (s, 3H), 1.44 (d, J = 7.1 Hz, 3H); MS (FAB) m/z 544 (M + H). Anal. Calcd for C24H22F5N3O4S: C, 53.04; H, 4.08; N, 7.73. Found: C, 53.20; 4.06; N, 7.70.
4.1.3.53. N-((2-(4-Chlorobenzyloxy)-6-(trifluoromethyl)pyridin-3-yl) methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (60).
Yield 74%, white solid, mp 154 °C; 1H NMR (CDCl3) δ 7.60 (d, J = 7.5 Hz, 1H), 7.48 (dd, J = 8.3, 8.3 Hz, 1H), 7.39–7.28 (m, 4H), 7.24 (d, J = 7.5 Hz, 1H), 7.05 (dd, J = 11.2, 2.0 Hz, 1H), 7.00 (d, J = 8.3 Hz, 1H), 6.48 (bs, 1H), 5.91 (bt, 1H), 5.37 (d, J = 4.7 Hz, 2H), 4.39 (m, 2H), 3.45 (q, J = 7.1 Hz, 1H), 3.02 (s, 3H), 1.44 (d, J = 7.0 Hz, 3H); MS (FAB) m/z 560 (M + H). Anal. Calcd for C24H22ClF4N3O4S: C, 51.48; H, 3.96; N, 7.50. Found: C, 51.67; H, 3.95; N, 7.47.
4.1.3.54. N-((6-(Trifluoromethyl)-2-(4-(trifluoromethyl)benzyloxy) pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl) propanamide (61).
Yield 63%, white solid, mp 167 ° C; 1H NMR (CDCl3) δ 7.64 (d, J = 8.4 Hz, 1H), 7.61 (d, J = 8.6 Hz, 2H), 7.55 (d, J = 8.1 Hz, 2H), 7.49 (dd, J = 8.2, 8.2 Hz, 1H), 7.25 (d, J = 8.3 Hz, 1H), 7.07 (dd, J = 11.2, 2.0 Hz, 1H), 7.01 (d, J = 7.9 Hz, 1H), 6.43 (bs, 1H), 5.89 (bt, 1H), 5.46 (m, 2H), 4.42 (d,J = 6.0 Hz, 2H), 3.48 (q, J = 7.1 Hz, 1H), 3.02 (s, 3H), 1.45 (d, J = 7.1 Hz, 3H); MS (FAB) m/z 594 (M + H). Anal. Calcd for C25H22F7N3O4S: C, 50.59; H, 3.74; N, 7.08. Found: C, 50.77; H, 3.72; N, 7.05.
4.1.3.55. N-((2-(4-Methoxybenzyloxy)-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (62).
Yield 67%, white solid, mp 138 °C; 1H NMR (CDCl3) δ 7.62 (d, J = 7.2 Hz, 1H), 7.49 (d, J = 8.4 Hz, 1H), 7.37 (d, J = 8.7 Hz, 2H), 7.23 (d, J = 7.2 Hz, 1H), 7.00 (m, 4H), 6.45 (bs, 1H), 5.94 (bt, 1H), 5.43 (m, 2H), 4.37 (m, 2H), 3.83 (s, 3H), 3.41 (m, 1H), 3.01 (s, 3H), 1.42 (d, J = 7.2 Hz, 3H); MS (FAB) m/z 556 (M + H). Anal. Calcd for C25H25F4N3O5S: C, 54.05; H, 4.54; N, 7.56. Found: C, 54.22; H, 4.52; N, 7.53.
4.1.3.56. N-((2-(3-Fluorobenzyloxy)-6-(trifluoromethyl)pyridin-3-yl) methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (63).
Yield 68%, white solid, mp 140 °C; 1H NMR (CDCl3) δ 7.63 (d, J = 7.5 Hz, 1H), 7.49 (d, J = 8.4 Hz, 1H), 7.36 (d, J = 6 Hz, 1H), 7.23 (m, 6H), 6.55 (bs, 1H), 5.97 (bt, 1H), 5.41 (d, J = 5.4 Hz, 2H), 4.40 (s, 2H), (q, J = 6.9 Hz, 1H), 3.01 (s, 3H), 1.45 (d, J = 7.2 Hz, 3H); MS (FAB) m/z 544 (M + H). Anal. Calcd for C24H22F5N3O4S: C, 53.04; H, 4.08; N, 7.73. Found: C, 52.91; H, 4.07; N, 7.70.
4.1.3.57. N-((2-(3-Chlorobenzyloxy)-6-(trifluoromethyl)pyridin-3-yl) methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (64).
Yield 66%, white solid, mp 155 °C; 1H NMR (CDCl3) δ 7.63 (d, J = 7.5 Hz, 1H), 7.50 (d, J = 8.4 Hz, 1H), 7.31 (m, 7H), 5.93 (bt, 1H), 5.97 (bt, 1H), 5.39 (d, J = 7.2 Hz, 2H), 4.42 (s, 2H), 3.47 (q, J = 6.9 Hz, 1H), 3.01 (s, 3H), 1.46 (d, J = 6.9 Hz, 3H); MS (FAB) m/z 560 (M + H). Anal. Calcd for C24H22ClF4N3O4S: C, 51.48; H, 3.96; N, 7.50. Found: C, 51.38; H, 3.95; N, 7.48.
4.1.3.58. N-((2-(3-Methoxybenzyloxy)-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (65).
Yield 68%, white solid, mp 128 °C; 1H NMR (CDCl3) δ 7.62 (d, J = 7.7 Hz, 1H), 7.46 (dd, J = 8.3, 8.3 Hz, 1H), 7.31 (dd, J = 8.1 Hz, 1H), 7.23 (d, J = 7.4 Hz, 1H), 7.06–6.88 (m, 4H), 6.90 (m, 1H), 6.49 (bs, 1H), 5.99 (bt, 1H), 5.39 (m, 2H), 4.39 (m, 2H), 3.83 (s, 3H), 3.42 (q, J = 7.1 Hz, 1H), 3.01 (s, 3H), 1.42 (d, J = 7.1 Hz, 3H); MS (FAB) m/z 556 (M + H). Anal. Calcd for C25H25F4N3O5S: C, 54.05; H, 4.54; N, 7.56. Found: C, 54.19; H, 4.52; N, 7.53.
4.1.3.59. N-((2-(3,5-Difluorobenzyloxy)-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (66).
Yield 66%, white solid, mp 145–148 °C; 1H NMR (CDCl3) δ 7.63 (d, J = 7.5 Hz, 1H), 7.52 (t, J = 8.4 Hz, 1H), 7.27 (d, J = 3 Hz, 1H), 7.10 (m, 2H), 6.94 (m, 2H), 6.79 (t, J = 2.1 Hz, 1H), 6.47 (bs, 1H), 5.91 (bt, 1H), 5.39 (m, 2H), 4.43 (q, J = 3.3 Hz, 2H), 3.51 (q, J = 6.9 Hz, 1H), 3.02 (s, 3H), 1.48 (d, J = 6.9 Hz, 3H); MS (FAB) m/z 562 (M + H). Anal. Calcd for C24H21F6N3O4S: C, 51.34; H, 3.77; N, Found: C, 51.52; H, 3.75; N, 7.45.
4.1.3.60. N-((2-(3,5-Dichlorobenzyloxy)-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (67).
Yield 72%, white solid, mp 156–160 °C; 1H NMR (CDCl3) δ 7.63 (d, J = 7.5 Hz, 1H), 7.63 (t, J = 7.5 Hz, 1H), 7.51 (t, J = 8.1 Hz, 1H), 7.33 (bs, 3H), 7.26 (m, 1H), 7.10 (m, 2H), 6.50 (bs,1H), 5.92 (bt, 1H), 5.36 (m, 2H), 4.43 (m, 2H), 3.53 (q, J = 6.6 Hz, 1H), 3.02 (s, 3H), 1.45 (d, J = 9.6 Hz, 3H); MS (FAB) m/z 595 (M + H). Anal. Calcd for C24H21Cl2F4N3O4S: C, 48.49; H, 3.56; N, 7.07. Found: C, 48.65; H, 3.54; N, 7.03.
4.1.3.61. N-((2-(3,5-Dimethoxybenzyloxy)-6-(trifluoromethyl)pyridin-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (68).
Yield 65%, white solid, mp 148 °C; 1H NMR (CDCl3) δ 7.63 (d, J = 7.3 Hz, 1H), 7.45 (dd, J = 8.3, 8.3 Hz, 1H), 7.24 (d, J = 7.5 Hz, 1H), 7.04 (dd, J = 11.2, 2.0 Hz, 1H), 6.98 (d, J = 8.8 Hz, 1H), 6.59 (d, J = 2.2 Hz, 2H), 6.45 (t, J = 2.4 Hz, 1H), 6.00 (bt, 1H), 5.26–5.41 (m, 2H), 4.30–4.48 (m, 2H), 3.81 (s, 6H), 3.43 (q, J = 7.3 Hz, 1H), 3.01 (s, 3H), 1.43 (d, J = 7.1 Hz, 3H); MS (FAB) m/z 586 (M + H). Anal. Calcd for C26H27F4N3O6S: C, 53.33; H, 4.65; N, 7.18. Found: C, 53.50; H, 4.63; N, 7.15.
4.1.3.62. N-((2-(Pyridin-2-ylmethoxy)-6-(tnfluoromethyl)pyridm-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (69).
Yield 67%, white solid, mp 105 °C; :H NMR (CDCl3) δ 8.61 (d, J = 4.7 Hz, 1H), 7.74 (dd, J = 7.6,1.7 Hz, 1H), 7.67 (d, J = 7.3 Hz, 1H), 7.42 (dd, J = 8.2, 8.2 Hz, 1H), 7.43 (d, J = 7.7 Hz, 1H), 7.33 (m, 1H), 7.24 (d, J = 7.5 Hz, 1H), 7.09 (dd, J = 11.4, 2.0 Hz, 1H), 7.01 (d, J = 8.2 Hz, 1H), 6.54 (bs, 1H), 5.51 (m, 2H), 4.45 (d, J = 5.7 Hz, 2H), 3.58 (q, J = 7.0 Hz, 1H), 3.01 (s, 3H), 1.46 (d, J = 7.0 Hz, 3H); MS (FAB) m/z 527 (M + H). Anal. Calcd for C23H22F4N4O4S: C, 52.47; H, 4.21; N, 10.64. Found: C, 52.66; H, 4.19; N, 10.62.
4.1.3.63. N-((2-(Pyridin-3-ylmethoxy)-6-(tnfluoromethyl)pyridm-3-yl)methyl)-2-(3-fluoro-4-(methylsufonamido)phenyl)propanamide (70).
Yield 68%, white solid, mp 95–97 °C; 1H NMR (CDCl3) δ 8.69 (m, 2H), 7.79 (d, J = 7.7 Hz, 1H), 7.64 (d, J = 7.5 Hz, 1H), 7.48 (dd, J = 8.1 Hz, 1H), 7.34 (m, 1H), 7.25 (d, J = 7.5 Hz, 1H), 7.05 (dd, J = 11.4, 1.9 Hz, 1H), 7.02 (d, J = 8.4 Hz, 1H), 5.87 (bt, 1H), 5.41 (s, 2H), 4.38 (d, J = 6.2 Hz, 2H), 3.49 (q, J = 7.3 Hz, 1H), 3.04 (s, 3H), 1.46 (d, J = 7.1 Hz, 3H); MS (FAB) m/z 527 (M + H). Anal. Calcd for C23H22F4N4O4S: C, 52.47; H, 4.21; N, 10.64. Found: C, 52.63; H, 4.19; N, 10.60.
4.1.3.64. N-((2-(Pyridin-4-ylmethoxy)-6-(tnfluoromethyl)pyridm-3-yl)methyl)-2-(3-fluoro-4-(methylsulfonamido)phenyl)propanamide (71).
Yield 75%, white solid, mp 103–105 °C; 1H NMR(CDCl3) δ 8.49 (d, J = 6.2 Hz, 2H), 7.61 (d, J = 7.4 Hz, 1H), 7.49 (d, J = 6.2 Hz, 2H), 7.40 (dd, J = 8.3, 8.3 Hz, 1H), 7.32 (d, J = 7.4 Hz, 1H), 7.19 (dd, J = 11.5, 2.0 Hz, 1H), 7.14 (d, J = 8.4 Hz, 1H), 5.49 (s, 2H), 4.35 (m, 2H), 3.72 (q, J = 6.9 Hz, 1H), 2.96 (s, 3H), 1.45 (d, J = 7.0 Hz, 3H); MS (FAb) m/z 527 (M + H). Anal. Calcd for C23H22F4N4O4S: C, 52.47; H, 4.21; N, 10.64. Found: C, 52.65; H, 4.20; N, 10.61.
4.2. Animal test
4.2.1. Animals
The studies were conducted with male NMRI mice (20–35 g) and male Sprague Dawley rats (130–180 g), supplied by commercial breeders (Charles River, Sulzfeld, Germany; Iffa Credo, Brussels, Belgium; Janvier, Genest St. Isle, France). Animals were housed under a 12:12 h light-dark cycle (lights on at 06:00 a.m.), and with room temperature 20–24 °C, relative air humidity 35–70%, 15 air changes per hour, and air movement <0.2 m/s. The animals had free access to standard laboratory food (Ssniff R/M-Haltung, Ssniff Spezialdiäten GmbH, Soest, Germany) and tap water. All animals were used only once in all pain models. There were at least five days between delivery of the animals and the start of experiment. Animal testing was performed in accordance with the recommendations and policies of the International Association for the Study of Pain [22] and the German Animal Welfare Law. All study protocols were approved by the local government committee for animal research, which is also an ethics committee. Animals were assigned randomly to treatment groups. Different doses and vehicle were tested in a randomized fashion. Although the operators performing the behavioral tests were not formally ‘blinded’ with respect to the treatment, they were not aware of the study hypothesis or the nature of differences between drugs.
4.2.2. Formalin test
The formalin test is a broadly used model of chemically-induced persistent pain [20]. Generally two phases of the nociceptive response can be observed in this model: a first phase starting immediately after injection, and a second phase starting around 15 min after formalin injection. In the first phase nociceptors are activated directly by the chemical stimulation of formalin and therefore this period is considered as a model of acute pain. The second phase reflects a spinal and peripheral hypersensitization (chronic phase) and is considered as a model for chronic pain. In this study, a period within the second phase was analyzed. The test was carried out in a plexiglass box with a mirror placed behind to allow an unobstructed view of the animals. Each animal was injected with 20 μl of 1% formalin in 0.9% NaCl subcutaneously into the dorsal surface of the right hind paw. After placing the mice back into the chamber the nociceptive behavior was measured by observation during the time interval of 21–24 min post-formalin and the amount of time spent licking and biting the injected paw was counted [20]. A vehicle control group was included for each investigation. Statistical significance is tested by Kruskal–Wallis-Test. Group size was 10 animals.
4.3. Molecular modeling
The 3D structure of the ligand was generated with Concord and energy minimized using MMFF94s force field and MMFF94 charge until the rms of Powell gradient was 0.05 kcal mol−1 A−1 in SYBYL-X 1.2 (Tripos Int., St. Louis, MO, USA). The flexible docking study on our hTRPV1 model [16] was performed by GOLD v.5.0.1 (Cambridge Crystallographic Data Centre, Cambridge, UK), which uses a genetic algorithm (GA) and allows for full ligand flexibility and partial protein flexibility. The binding site was defined as 8 A around the capsaicin docked in the hTRPV1 model. The side chains of the nine residues, which are important for ligand binding, (i.e., Tyr511, Ser512, Met514, Leu515, Leu518, Phe543, Leu547, Thr550, and Asn551) were set to be flexible with ‘crystal mode’ in GOLD. The ligand was docked using the GoldScore scoring function with 30 GA runs. Other parameters were set as default. All computation calculations were undertaken on Intel® Xeon™ Quad-core 2.5 GHz workstation with Linux Cent OS release 5.5.
Table 4.
In vitro hTRPV1 antagonistic activities for 2-cycloalkylmethyloxy derivatives.
| R | Ki [CAP] (nM) | IC50 [pH] (nM) | R | Ki [CAP] (nM) | IC50 [pH] (nM) | ||
|---|---|---|---|---|---|---|---|
| 41 |  |
1.1 | 115 | 46 |  |
0.7 | l9.7 |
| 42 |  |
0.3 | 5.1 | 47 |  |
1.0 | 44.4 |
| 43 |  |
1.1 | 161 | 48 |  |
WE | NE |
| 44 |  |
0.3 | 40.5 | 49 |  |
3.4 | 342 |
| 45 |  |
0.4 | 11.6 | 50 |  |
0.3 | l6.6 |
WE: weakly effective (1 μM < Ki or IC50 < 10 μM).
NE: not effective (Ki or IC50 > 10 μM).
Acknowledgments
This research was supported by Research Grants from Grunenthal, Grants from the National Research Foundation of Korea (NRF) (R11-2007-107-02001-0), Grants from the National Leading Research Lab (NLRL) program (2011-0028885), the Ewha Global Top 5 Grant 2011 and in part by the Intramural Research Program of NIH, Center for Cancer Research, NCI (Project Z1A BC 005270).
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