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Regioselective synthesis of 2- and 4-diarylpyridine ethers and their inhibitory activities against phosphodiesterase 4B
Journal of Molecular Structure 1196 (2019) 455e461
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Journal of Molecular Structure journal homepage: http://www.elsevier.com/locate/molstruc
Regioselective synthesis of 2- and 4-diarylpyridine ethers and their inhibitory activities against phosphodiesterase 4B Xinyun Zhao a, 1, Huihui Chen a, 1, Suting Xing a, Wei Yuan a, Lamei Wu a, Xi Chen a, *, Chang-Guo Zhan b, ** a b
College of Chemistry & Materials Science, South-Central University for Nationalities, Wuhan, 430074, PR China College of Pharmacy, University of Kentucky, Lexington, KY, 40536, USA
a r t i c l e i n f o
a b s t r a c t
Article history: Received 16 April 2019 Received in revised form 8 June 2019 Accepted 25 June 2019 Available online 28 June 2019
Two diaryl ether regioisomers of pyridine were prepared through the CuI/TMEDA/Cs2CO3-catalyzed reaction of 2,4-dibromopyridine with phenol derivatives under nitrogen atmosphere. The polar solvent DMSO gave 4-isomer as the major product whereas less polar toluene resulted in 2-isomer as a major product. Structures of regioisomers were confirmed by single crystal X-ray diffraction analysis. 4regioisomer shows higher biological activity against phosphodiesterase 4B (PDE4B) than that of 2isomer. Molecular docking simulations revealed that the PDE4B-inhibitory activity difference between the two regioisomers was mainly attributed to the atomic charge difference on the eOe linker. © 2019 Elsevier B.V. All rights reserved.
Keywords: Regioselectivity 2,4-Dibromopyridine Diaryl ethers PDE4B inhibitory activity X-ray crystal structure
1. Introduction Aryl ether is the core scaffold of many medicinal and/or natural product molecules, which are widely used in many fields including medicine and molecular biology. [1e7] Among these compounds, diaryl ethers containing pyridine moiety have attracted considerable attention due to their high biological activities. Thus, the preparation of such compounds is the main focus of research interest. The typical strategy for the synthesis of such compounds, follows aromatic nucleophilic substitution of 2,4-dihalogen pyridine (X ¼ F, Cl, Br). Regioselectivity of 2,4-dihalogen pyridine can introduce different functional groups at 2 or 4- position of pyridine ring. The reaction conditions and regioselectivity obtained for 2,4dihalogen pyridine are summarized in Table 1. As can be seen from this Table, aryl groups, [3,8] aryloxy groups, [9,10] arylamino groups, [11] arylacetylene groups [12] or other interesting functional groups can be selectively introduced to the 2- or 4- position of pyridine ring. For the synthesis of aryl ether with pyridine
* Corresponding author. ** Corresponding author. E-mail addresses: [email protected] (X. Chen), [email protected] (C.-G. Zhan). 1 These authors contributed equally to this work. https://doi.org/10.1016/j.molstruc.2019.06.086 0022-2860/© 2019 Elsevier B.V. All rights reserved.
moiety, 4-substituted product is preferred, when the reactions are performed under polar solvent environments. The exception is the nucleophilic substitution reaction of aryloxy groups carried out by Zhou and his coworkers [9] in which a 2-substituted aryloxypyridine product was reported to be a dominant product for the reaction in DMSO. Since 2- and 4- substituted aryloxypyridine products are regioisomers, which can lead to some confusion for characterization. However, for the design and/or synthesis of drug molecules, the accurate characterization of molecular structures is crucial. If a wrong structure was assigned, the structure-based computational molecule design would be meaningless. [13] So it is critical to study the regioselectivity of such type of reaction and unambiguously characterize the obtained products. Thus to reinvestigate the topic in detail, we firstly carried out the same reaction between 2,4-dibromopyridine and 4-methylphenol by using the same reaction conditions reported by Zhou and his coworkers (Scheme 1). [9] We further confirmed this regioselective behavior through proper X-ray crystal analysis of the product to avoid any ambiguity. Our experimental results showed that the 4-substituted aryloxypyridine product was the dominant product in DMSO solvent, and not the 2-substituted aryloxypyridines reported. In order to achieve 2-regioisomer with higher yield, the reaction of 2,4dibromopyridine with 4-methylphenol was performed in different solvents. In solvent toluene, another 2- regioisomer was a
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Table 1 Reported regioselective reactions of 2,4-dihalopyridine in literature.
X
R
Conditions
Preferential product
Ref
F/Cl Cl Cl Cl Br
OEt NHPh NHPh 4-CF3C6H4-
NaOEt/EtOH, DMF or DMSO Pd(OAc)2, Cs2CO3, Xantphos/microwave/100 C NaH/DMA 4-CF3C6H4MgBr/CrCl2 Pd(CF3COO)2/CuI/PPh3/i-Pr2NH
4-isomer 2-isomer 4-isomer 2-isomer 2-isomer
[10] [11] [11] [8] [12]
Br Br Br
Ar OAr 4-CH3C6H4O-
Pd(PPh3)4/aqTlOH CuI/TMEDA/Cs2CO3/DMSO CuI/TMEDA/Cs2CO3/DMSO
2-isomer 2-isomer 4-isomer
[3] [9] This work
Scheme 1. CuI catalyzed regioselective C-O coupling of 2,4-dibromopyridine with 4-methylphenol.Reagents and condition: CuI/TMEDA/Cs2CO3/toluene, N2, 110 C for (A), 78%; CuI/ TMEDA/Cs2CO3/DMSO, N2, 110 C for (B), 64%.
preferred product. The regioselectivity of the reaction of 2,4dibromopyridine with 4-methylphenol can be achieved by controlling the solvents polarity. Our experiments showed that 4substitued products was dominant in polar DMSO solvents, and the 2-regioisomers is dominant, if the reaction is performed in nonpolar toluene solvent. In addition, quantum mechanical calculations were carried out to find out the cause of regioselectivity for the reaction of 2,4-dibromopyridine and 4-methylphenol. This proven conclusion promoted us to further explore the regioselectivity of the reaction of 2,4-dibromopyridine with methyl (4-hydroxy-phenyl)acetate. Another two 2- and 4- regioisomers (1 and 2) were obtained in solvents DMSO and toluene (Scheme 2). We continued to synthesize two regioisomers 5 and 6 by following Suzuki reactions and hydrolysis reactions, which were our target molecules for biological inhibitory activity against phosphodiesterase 4B (PDE4B). [14e16] Further, molecular docking simulations were also performed to study the binding of regioisomers 5 and 6 with PDE4B enzyme and to explore the effects of different isomers on catalytic activity of PDE4B. 2. Experiments and calculation details 2.1. Chemical synthesis and biological activity tests All chemicals are commercially available. Solvents were dried in a routine way and redistilled. The NMR spectrum was recorded on a Varian 400 MHz NMR spectrometer. Chemical shift values are given in ppm relative to tetramethyl silane (TMS). High-resolution mass spectrometry (HRMS) was recorded on Agilent LC-Q-TOFMS-6520 instrument. Melting points were recorded on an X-6 digital melting point apparatus. The crystal structures of the title
compounds were determined on the Bruker Apex CCD X-ray instrument. 2.1.1. General procedure for the copper (I)-catalyzed regioselective coupling reaction of 2,4-dibromopyridine and methyl (4-hydroxyphenyl)acetate Synthesis of methyl 2-(4- ((2-bromo-4-pyridinyl)oxy) phenyl) acetate 1 in DMSO: CuI (18.6 mg, 0.098 mmol), TMEDA (11.4 mg, 0.098 mmol) were dissolved in dry DMSO (2 mL), and stirred at room temperature for 30 min under nitrogen atmosphere. cesium carbonate (638.6 mg, 1.96 mmol) and methyl (4-hydroxy-phenyl)-acetate (152.9 mg, 0.92 mmol) were added into the reaction mixture and stirred for another 4 h at room temperature. Finally, solution of 2,4dibromopyridine (236.8 mg, 0.99 mmol) in dried DMSO (3 mL) solvent was added to the reaction mixture. The reaction mixture was placed in oil-bath which was preheated to 110 C under nitrogen atmosphere. The reaction progress was monitored using TLC. After completion of the reaction, the reaction mixture was cooled to room temperature, and was filtered off. The mixture was diluted with dichloromethane (25 mL), which was washed with brine (3 30 mL). The organic phase was dried over sodium sulfate and the product was purified by column chromatography to give the title compound as pale yellow liquid. Yield: 75%. 1H NMR (400 MHz, CDCl3) d 8.21 (d, J ¼ 5.6 Hz, 1H), 7.36 (d, J ¼ 8.4 Hz, 2H), 7.05 (d, J ¼ 8.4 Hz, 2H), 6.99 (d, J ¼ 2.0 Hz, 1H), 6.83 (dd, J1 ¼ 2.0 Hz, J2 ¼ 5.6 Hz, 1H), 3.74 (s, 3H), 3.67 (s, 2H); 13 C NMR (101 MHz, CDCl3) d 171.7, 166.09, 152.6, 151.1, 143.0, 131.8, 131.3, 131.3, 120.9, 120.9, 115.7, 111.9, 52.2, 40.4. HRMS (ESI): Calcd. þ for C14H13BrNOþ 3 [MþH] : 322.0079; Found: 322.0073. Synthesis of methyl 2-(4-((4-bromo-2-pyridinyl)oxy) phenyl)
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Scheme 2. Reagents and reaction conditions: (a) CuI, TMEDA, Cs2CO3, DMSO, N2, 110 C; (b) CuI, TMEDA, Cs2CO3, Toluene, N2, 110 C; (c) PPh3, PdCl2, KOH, CH3CN, N2, 70 C; (d) KOH, CH3OH, RT.
acetate 2 in toluene: CuI (18.6 mg, 0.1 mmol) and TMEDA (11.4 mg, 0.1 mmol) was mixed in dried toluene (5 mL) and stirred for 30 min at room temperature. Cesium carbonate (638.6 mg, 2 mmol) and methyl (4hydroxy-phenyl)-acetate (152.7 mg, 0.92 mmol) were added into the mixture and stirred at room temperature for another 4 h. Finally, solution of 2,4-dibromopyridine (236.8 mg, 0.92 mmol) in dried toluene (3 mL) was added. The reaction mixture was placed in oil-bath which was preheated to 110 Cunder nitrogen atmosphere for 24 h. Reaction progress was monitored by TLC. On completion, reaction mixture was cooled to room temperature. The solvent was evaporated under vacuum and the residue was dissolved in dichloromethane (20 mL) and filtered and filtrate was purified by column chromatography, to obtain a pale yellow liquid (yield, 51%). 1 H NMR (400 MHz, CDCl3) d 8.01 (d, J ¼ 5.6 Hz, 1H), 7.33 (d, J ¼ 8.4 Hz, 2H), 7.15 (dd, J1 ¼1.2 Hz, J2 ¼ 5.2 Hz,1H), 7.11-7.09 (m, 3H), 3.71 (s, 3H), 3.64 (s, 2H); 13C NMR (101 MHz, CDCl3) d171.8, 164.3, 152.7, 148.2, 134.7, 130.8, 130.7, 130.7, 122.0, 121.4, 121.4, 114.7, þ 52.1, 40.6. HRMS (ESI): Calcd. for C14H13BrNOþ 3 [MþH] : 322.0079; Found: 322.0072.
2.1.2. Suzuki coupling reaction for synthesis of methyl 2-(4-((2-(5chloro-2-thiophenyl)-4-pyridinyl)oxy)phenyl)acetate 3 and methyl methyl 2-(4-((4-(5-chloro-2-thiophenyl)-2-pyridinyl)oxy)phenyl) acetate 4 PdCl2 (43.4 mg,0.24 mmol) and PPh3 (257 mg,0.98 mmol) were
dissolved in dried CH3CN(10 mL), KOH (274.4 mg,4.9 mmol) and 5chlorothiophene-2-boronic acid pinacol ester (659.7 mg,2.69 mmol) were added to the above mixture and then the intermediate (1 or 2) (790 mg, 2.45 mmol) was added, under nitrogen atmosphere. The reaction mixture was placed in oil-bath which was preheated to 70 C. Reaction progress was monitored by TLC. On completion, the reaction mixture was cooled to room temperature, filtered and filtrate was purified by chromatography to give the title compound. Compound 3: white solid, yield 74%, m.p. 120.1e121.7 C. 1H NMR (400 MHz, CDCl3) d 8.36 (d, J ¼ 5.6 Hz, 1H), 7.35 (d, J ¼ 8.4 Hz, 2H), 7.25 (d, J ¼ 4.0 Hz, 1H), 7.12 (d, J ¼ 2.4 Hz, 1H), 7.07 (d, J ¼ 8.4 Hz, 2H), 6.89 (d, J ¼ 4.0 Hz, 1H), 6.69 (dd, J ¼ 5.6, 2.4 Hz, 1H), 3.74 (s, 3H), 3.67 (s, 2H); 13C NMR (101 MHz, CDCl3) d171.8, 165.5, 153.5, 153.1, 151.0, 132.9, 131.3, 131.2, 131.2, 127.2, 127.2, 124.0, 120.8, 120.8, 110.8, 106.5, 52.2, 40.4; IR(KBr, cm1): 1625, 1501, 1432, 1317, 1088, 803, 692; HRMS (ESI): Calcd. for C18H15ClNO3Sþ [MþH]þ: 360.0461; Found: 360.0456. Compound 4: yellow solid, yield 48%. mp79.3e80.8 C. 1H NMR (400 MHz, CDCl3) d 8.14 (d, J ¼ 5.2Hz,1H), 7.33 (d, J ¼ 7.6Hz,2H), 7.27 (s,1H), 7.08e7.12 (m,3H), 7.00 (s,1H), 6.95 (d, J ¼ 3.6Hz,1H), 3.71 (s,3H), 3.64 (s,2H); 13C NMR (101 MHz, CDCl3) d171.8, 164.5, 153.1, 148.3, 144.1, 139.3, 132.1, 130.6, 130.4, 127.6, 125.0, 121.3, 114.8, 106.9, 52.0, 40.6; IR(KBr, cm1): 1752, 1604, 1550, 1513, 1442, 1401, 1303, 1234, 1157, 802, 751; HRMS (ESI): Calcd. for C18H15ClNO3Sþ [MþH]þ: 360.0461; Found: 360.0454.
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2.1.3. General procedure for synthesis of 2-(4-((2-(5-chloro-2thiophenyl) -4-pyridinyl)oxy)phenyl)acetic acid 5 and 2-(4-((4-(5chloro-2-thiophenyl) -2-pyridinyl)oxy)phenyl)acetic acid 6 Intermediates (3 or 4) (96.4 mg,0.27 mmol) was dissolved into methanol (6 mL), and aqueous KOH (30 mg, 0.54 mmol, dissolved into 1 mL of H2O) was added dropwise into the above ester solution,the mixture was stirred at room temperature. The reaction progress was monitored by TLC. On completion, the solvent was evaporated under vacuum and water was added into the residue. The solution was neutralized by diluted hydrochloride solution till pH 6-7 under ice-bath condition. The resulted solid was filtered and purified by column chromatography to give the target compounds 5 and 6. Compound 5: yellow solid, yield 82%. mp, 195.0e196.8 C, 1H NMR (400 MHz, DMSO‑d6) d 8.37 (d, J ¼ 5.6 Hz, 1H), 7.68 (d, J ¼ 4.0 Hz, 1H), 7.60 (s, 1H), 7.38 (d, J ¼ 8.4 Hz, 2H), 7.16 (t, J ¼ 5.9 Hz, 3H), 6.71 (dd, J ¼ 5.6, 2.4 Hz, 1H), 3.61 (s, 2H); 13C NMR (101 MHz, DMSO‑d6) d173.2, 173.2, 165.5, 153.4, 153.4, 152.8, 152.8, 151.8, 143.5, 14.5, 133.0, 131.8, 131.8, 131.3, 128.7, 128.7, 125.7, 125.7, 120.6, 111.2, 111.1, 107.3; IR(KBr, cm1): 2921, 1621, 1468, 1404, 1302, 1221, 1161, 1079, 890, 813; HRMS (ESI): Calcd. for C17H13ClNO3Sþ [MþH]þ: 346.0305; Found: 346.0298. Compound 6: yellow solid, yield 79%. mp, 164.5e166.4 C. 1H NMR (400 MHz, DMSO‑d6) d 12.38 (br, 1H), 8.13 (d, J ¼ 5.2 Hz, 1H), 7.76 (d, J ¼ 4.4 Hz, 1H), 7.35 (d, J ¼ 5.6 Hz, 1H), 7.32e7.26 (m, 4H), 7.09 (d, J ¼ 8.4 Hz, 2H), 3.59 (s, 2H); 13C NMR (101 MHz, DMSO‑d6) d173.2, 164.5, 152.9, 148.7, 144.0, 139.2, 131.8, 131.0, 130.9, 129.1, 127.3, 121.5, 115.4, 107.0, 40.5; IR(KBr, cm1): 3201, 1689, 1608, 1548, 1441, 1397, 1304, 1224, 1179, 1076, 788, 678; HRMS (ESI): Calcd. for C17H13ClNO3Sþ [MþH]þ: 346.0305; Found: 346.0300. 2.1.4. Synthesis of 4-bromo-2-p-tolyloxy-pyridine (A) and 2bromo-4-p-tolyloxy- pyridine (B) Synthesis of compound A. The procedure is the same as that of synthesis of compound 2 to give white solid. Yield 64%. mp: 47.0e48.9 C; 1H NMR (400 MHz, CDCl3) d 8.01 (d, J ¼ 5.2 Hz, 1H), 7.21 (d, J ¼ 8.0 Hz, 2H), 7.12 (d, J ¼ 5.6 Hz, 1H), 7.07 (s, 1H), 7.01 (d, J ¼ 8.0 Hz, 2H), 2.36 (s, 3H).13C NMR (101 MHz, CDCl3) d164.7, 151.3, 148.3, 134.8, 134.6, 130.4, 130.3, 121.7, 121.2, 121.2, 114.5, 20.9. HRMS (ESI): Calcd. for C12H11BrNOþ [MþH]þ: 264.0024; Found: 264.0018. Synthesis of compound B The procedure is the same as that of synthesis of compound 1 to give white solid. Yield 78%. Mp:69.2e70.8 C(63.1e64.7 C [9]); 1H NMR (CDCl3,400 MHz): d, 8.18 (d, J ¼ 6.0 Hz, 1H), 7.23 (d, J ¼ 8.0 Hz, 2H), 6.97 (m, 3H), 6.81e6.79 (m, 1H), 2.38 (s, 3H); 13C NMR (101 MHz, CDCl3) d166.3, 151.0, 151.0, 143.0, 135.8, 130.9, 130.9, 120.7, 120.6, 115.4, 111.7, 20.9. Calcd. for C12H11BrNOþ [MþH]þ: 264.0024; Found: 264.0020. 2.1.5. Biological assay against PDE4B PDE4B enzyme inhibitory data from BPS Bioscience Inc. (San Diego, California, USA) using fluorescence polarization method. Tested compounds were dissolved in DMSO and diluted in assay buffer (final DMSO concentration 1%, final inhibitor concentration 10 mM). 2.2. Molecular simulations 2.2.1. Quantum chemistry calculation Compounds A, B, and 5 were chosen for the study. The initial geometries of these molecules were firstly constructed by using the GaussView [17]. For molecules with multiple torsion bonds, the initial geometries were built in the most extended form. The constructed geometries were optimized by using the Density Functional Theory (DFT) Becke's three-parameter hybrid exchange functional and the Lee-Yang-Parr correlation functional (B3LYP)
[18e20] and 6-31 þ G* basis set, followed by vibrational frequency analysis at the same level of theory to ensure that the optimized geometries are indeed associated with local minima on the potential energy surface. To explore the origin of regioselectivity for the reaction of 2,4-dibromopyridine coupling with phenol derivatives, the optimized geometries of A and B were used to carry out single-point energy calculations using the M06-2X [21,22] method and the 6-311þþG(2d, 2p) basis set. For all the computation mentioned above, default criteria were used for the SCF iteration and for geometrical optimization. All the quantum chemistry calculations described above were performed by using Gaussian 09 software [23] installed on a local Linux 64 clusters. 2.2.2. Molecular docking In an effort to gain an understanding of the structural basis for the PDE4 inhibitory activities observed, we further studied the binding mode of these two compounds 5 and 6 through molecular docking. For this purpose, the crystal structure of PDE4B in complex with a [1,3,5]triazine derivative (PDB ID: 4kp6) was selected as the receptor for molecular docking. Before docking, the ligand and water molecules were removed from the complex structure, except for four water molecules and a hydroxide ion that bound with the metal ions Zn2þ and Mg2þ at the catalytic pocket. Then hydrogen atoms were added by using the Leap tools implemented in AMBER software. The molecular structure of compounds 5 and 6 were constructed by the GaussView followed by geometrical optimization at PM3 level. The optimized structures as well as the crystal structure of 5 were then used for docking study. For the receptor and each ligand, the nonpolar hydrogen atoms were merged and Gasteiger charges were added. Then AUTODOCK4.2 [24] program was used to search for the most favorable binding mode of the ligands and PDE4 catalytic domain. Coordinates of Mg2þ was set as grid center and grid size was set to 60 60 60 with spacing of 0.375 Å. During the docking process, atoms in the receptor were frozen. 100 docking runs were performed for each ligand and the conformations with the lowest binding free energies were selected for analysis. Default values were used for other docking parameters. 3. Results and discussion 3.1. Regioselectivity of 2,4-dibromopyridine for synthesis of diaryl ethers In order to further explore the regioselectivity of 2,4dibromopyridine coupling with phenol derivatives, an model reaction from Zhou's work carried out to investigate the regioselectivity of 2,4-dibromopyridine with 4-methylphenol in the presence of CuI/TMEDA/Cs2CO3 in DMSO or toluene (Scheme 1). [9] TLC was used to monitor the progress of the reactions and the separation of products was performed by column chromatography. Detailed solvent optimized reaction conditions can be seen in Table S1 of Supporting Information. The experimental procedure in DMSO was the same as that of the reported one. In DMSO reaction system, the starting material 2,4-dibromopyridine disappeared completely, but in solvent toluene, a small amounts of 2,4dibromopyridine was left and recovered. In solvent toluene, the isolated yields of 2-regioisomer (compound A, 64%), other 4regioisomer (compound B, 3.7%) and small amounts of disubstituted product in the solvent toluene were obtained. The isolated yields of 2-regioisomer (compound A, 3.7%), 4-regioisomer (compound B, 78%) and small amounts of disubstituted product in DMSO system were achieved. From the 1H NMR data of compounds A and B, the signal at 8.00 ppm appears in 2-regioisomeric derivative whereas the same signal is shifted to 8.17 ppm in 4-regioisomeric derivative (See NMR S1 and S3). Surprisingly, Zhou and
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coworkers have given the data similar to 4-regioisomer (for compound 3b, page 331 in Zhou's work) with signal at 8.17 ppm, though claiming as 2-regioisimer. The NMR results of compound B show that 1H NMR and 13C NMR data are consistent with Zhou's work in DMSO solvent. [9] In order to further confirm the regioselectivity of such kind of this reaction, single crystal of compound B suitable for X-ray analysis was grown in ethanol/acetone (1:1) mixture. Crystallographic data for the reported crystal structure was deposited at the Cambridge Crystallographic Data Centre with CCDC number 1534716 (Fig. 1). From Fig. 1, it is clear that the phenoxy group of compound B is located at the 4-position of pyridine, and not 2position of pyridine, as reported in literature. Our single crystal X-ray analysis on compound B exhibits greatly different from the reported one. In solvent DMSO, preferential C4-substituted isomer of pyridine (compound B in Scheme 1) was obtained. Detailed structural information about crystallographic data of compound B can be seen in Table S2, S3 and S4. In these experiments, preferential C2- substituted isomer of pyridine (compound A in Scheme 1) was obtained in solvent toluene with optimized yield 64%. Regioisomer A crystals suitable for single crystal X-ray diffraction analysis can't be obtained although many attempts were tried. Moreover, different regioisomers also exhibit different physical property such as polarity, which will eventually leading to different crystal stacking modes and melting points. As will be seen in the subsequent sections, regioisomers A and B show significantly different polarities for A (1.52 Debye) and for B (5.63 Debye), then they display different melting points for A (47.0e48.9 C) and for B (69.2e70.8 C). According to the above regioselective results, further structural modification was made by introducing 4-CH2COOMe into the benzene ring. Depicted in Scheme 2 are the regioselectively synthetic routes for diaryl ethers 1 and 2, as well as their downstream products 5 and 6. 4-phenoxy substituted regioisomer 1 and 2phenyloxy substituted regioisomer 2 was firstly prepared selectively through the copper-catalyzed regioselective coupling reaction of 2,4-dibromopyridine and methyl 4-hydroxyphenylacetate in solvents DMSO and toluene, respectively. This is similar to the optimized conditions of Scheme 1. For synthesis of compounds 1 and 2, in toluene solvent, the yields of 2-regioisomer (compound 2, 51%) and 4- regioisomer (compound 1, 14%) were obtained. In DMSO solvent, the yields of 2-regioisomer (compound 2, 11%) and 4- regioisomer (compound 1, 75%) were obtained. In DMSO and toluene system, small amounts of disubstituted product was obtained. Intermediates 3 and 4 were obtained through Suzuki coupling reactions between regioisomers 1 or 2 and 5chlorothiophene-2-boronic acid pinacol ester in the presence of PdCl2/PPh3 with KOH in dried CH3CN. [25] The difference of yields
Fig. 1. X-ray crystal structure of compound B from 2,4-dibromopyridine reacting with 4-methylphenol in CuI/TMEDA/Cs2CO3/DMSO under nitrogen atmosphere.
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for compounds 3 and 4 depends on the reactive activity of bromine atoms at the pyridine within the regioisomers 1 and 2. Different positions of aryloxy substitutents at pyridine lead to different reactive activity of bromine atoms. Compounds 5 and 6 were obtained by hydrolyzing the corresponding esters 3 and 4. Single crystal of regioisomer 5 was grown in methanol at 298 K. Then the crystal structure of 5 was determined by single crystal X-ray diffraction analysis (Fig. 2). The structure has been deposited in Cambridge Crystallographic Data Centre with deposition CCDC numbers:1534718. Other corresponding structural information can be seen in Tables S5, S6, S7 of Supporting Information. Numerous attempts failed to grow regioisomer 6 crystal suitable for single crystal X-ray diffraction analysis. From the structure of target compound 5, it is clear that the phenoxy group is located at the 4-position of pyridine ring. Since regioisomer 1 is the raw material for the synthesis of 5, regioisomer 1 should also be a 4-phenoxyl substituted regioisomer. This conclusion is inconsistent with the work reported by Zhou et al., in which the 2-regioisomer product was reported to be dominant under the same condition. [9] 3.2. Results of the DFT calculations. The DFT calculations at M06-2X/6-311þþG(2d, 2p) level demonstrated that the compound A was slightly more stable than B by ~0.2 kcal/mol. These calculations also show that the dipole moment of A is 1.52 Debye, which is much smaller than that of compound B (5.63 Debye). On the other hand, the dipole moment of toluene and DMSO are predicted to be 0.45 and 6.07 Debye, respectively. According to the principle of like dissolves like, compound A are more easy to dissolve in solvent toluene, and compound B are easy to dissolve in DMSO. Hence, in DMSO compound B gains more stability than A, which may lead to high yields of A in this solvent. In addition to the polarity of the solvent, there maybe are other factors leading to the difference between the yields of the products A and B. In addition, the optimized geometrical parameters of 5 were compared with the corresponding ones in the X-ray crystal structure. These comparisons are collected in Fig. 3. As can be seen from this figure, they match each other well for compound 5, suggesting that the quantum chemistry calculation results in this study are reliable. 3.3. Docking and PDE4B enzyme activity of diaryl ethers regioisomers The real structural assignment of aryloxy group on the pyridine core structure has been crucial for structure-based drug design and discovery. We have confirmed the real structure of diaryl ethers derivatives with a pyridine moiety using single crystal X-ray diffraction technology. Docking study of regioisomers 5 and 6 with PDE4 enzyme were performed. As can be seen from Fig. 4, the docking conformation obtained by using crystal structure of 5 (colored by purple scheme) is nearly identical to one obtained by using gas-phase optimized geometries of 5 (color by blue scheme), suggesting that the gas-phase optimized geometrical parameters are reliable. Molecular docking also reveals that target compounds 5 and 6 bind with PDE4 in similar binding modes (Fig. 4). Both ligands were fitted in a cavity formed by His234, Met347, Leu393, Asp392, Ser394, Asn395, Pro396, Thr397, Tyr403, Ile410, Phe414, Met431, Ser442, Gln443, Phe446, Ile450 and Val451 residues. The thiophene and pyridine rings of both ligands are lodged in the hydrophobic pocket surrounded by the side chains of Ile410, Ile450, Phe414, and Phe446 residues, causing a high degree of surface complementarities. The carboxylic groups were fitted in a
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Fig. 2. The crystal structure of the target compound 5.
Collected in Table 2 are the estimated binding free energies (DGcal bind) along with the corresponding experimental inhibitory ratio at 10 mM inhibitor concentration. As can be seen from the Table, the DGcal bind predicted by the AUTODOCK are consistent with the corresponding experimental data, suggesting that the present binding modes of these compounds are reasonable. Further energy decomposition analysis showed that the difference in DGcal bind is caused by the hydrogen bonding interactions formed between the -O- linker and the Gln443 glutamine switch (not shown). Charge analysis shows that the atomic charge (0.331) of this oxygen in 5 is significantly more negative than the counterpart (0.292) in 6, leading to stronger hydrogen bonding interactions in 5. 4. Conclusions Fig. 3. Selected geometrical parameters of 5 from (A) gas-phase geometry optimization at the B3LYP/6-31 þ G* level and (B) X-ray crystal structure.
Fig. 4. Binding modes of ligands 5 and 6 in the active site of PDE4. Ligands rendered with blue and green schemes are docking conformations obtained by using gas-phase optimized geometries of 5 and 6, respectively. Ligand rendered with purple scheme is docking conformation obtained by using experimental crystal structure of 5.
Diaryl ethers derivatives with pyridine moiety were regioselectively synthesized from 2,4-dihalopyridine through three steps sequence, including CuI-catalyzed regioselective reactions in different solvents, Suzuki coupling reactions and esters hydrolysis. The real structure of 4-regioisomer 5 was confirmed by single crystal X-ray diffraction analysis, i.e. 2,4-dibromopyridine can react with methyl 4-hydroxyphenylacetate to selectively give dominant 4-regioisomer of pyridine in the presence of CuI/TMEDA/Cs2CO3/ N2/DMSO. Different regioisomers 5 and 6 were docked into PDE4B binding pocket and showed different binding energy in spite of similar binding modes with different hydrogen bonding interaction contribution of -O- linker. 4-regioisomer 5 has higher inhibitory activity than that of 2-regioisomer 6 at 10 mM ligand concentration. In order to further confirm the CuI-catalyzed regioselectivity of 2,4dibromopyridine in DMSO, The reaction of 2,4-dibromopyridine and 4-methylphenol was carried out under the same reaction condition to give predominantly the 4-regioselective isomer which structure has been confirmed by single crystal X-ray diffraction analysis. The regioselectivity of 2,4-dihalopyridine in different solvents maybe correlate with the polarity of products diaryl ethers. It is clear that regioselectivity of a chemical reaction and real structures of medicinal molecules are very important for structurebased drugs design. Our current research will provide a reasonable basis for the synthesis of regioselective diaryl ethers and the design of diaryl ether type PDE4 inhibitors.
Table 2 Calculated binding free energies in comparison with available experimental data.
hydrophilic pocket formed by His234, Asp392, Ser394, Asn395, Thr397, and Tyr403. Despite the common features of the binding modes, the two thiophene rings flipped relatively to each other by about 180 .
Regioisomers 5 6 (b)
a DGcal bind
6.6 6.2
Inhibitory ratio obtained at 10 mM ligand concentration. Binding free energies predicted by AUTODOCK.
a
Inhibitory ratio (%) 64 26
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Acknowledgements The work was supported by the National Natural Science Foundation of China (grant number 21273089) and the Fundamental Research Funds for the Central Universities, South-Central University of Nationalities (CZW17004). Appendix A. Supplementary data Supplementary data to this article can be found online at https://doi.org/10.1016/j.molstruc.2019.06.086. References Debreczeni, R.A. Norman, [1] F.W. Goldberg, R.A. Ward, S.J. Powell, J.E. N.J. Roberts, A.P. Dishington, H.J. Gingell, K.F. Wickson, A.L. Roberts, J. Med. Chem. 52 (2009) 7901e7905. [2] R.J. Patch, L.L. Searle, A.J. Kim, D. De, X. Zhu, H.B. Askari, J.C. O'Neill, M.C. Abad, D. Rentzeperis, J. Liu, M. Kemmerer, L. Lin, J. Kasturi, J.G. Geisler, J.M. Lenhard, M.R. Player, M.D. Gaul, J. Med. Chem. 54 (2011) 788e808. mez, M.M. Cid, Tetrahedron 62 (2006) 11063e11072. [3] C. Sicre, J.L. Alonso-Go [4] N.A. Swain, D. Batchelor, S. Beaudoin, B.M. Bechle, P.A. Bradley, A.D. Brown, B. Brown, K.J. Butcher, R.P. Butt, M.L. Chapman, S. Denton, D. Ellis, S.R.G. Galan, S.M. Gaulier, B.S. Greener, M.J. de Groot, M.S. Glossop, I.K. Gurrell, J. Hannam, M.S. Johnson, Z. Lin, C.J. Markworth, B.E. Marron, D.S. Millan, S. Nakagawa, A. Pike, D. Printzenhoff, D.J. Rawson, S.J. Ransley, S.M. Reister, K. Sasaki, R.I. Storer, P.A. Stupple, C.W. West, J. Med. Chem. 60 (2017) 7029e7042. [5] Z.K. Sweeney, S.F. Harris, N. Arora, H. Javanbakht, Y. Li, J. Fretland, J.P. Davidson, J.R. Billedeau, S.K. Gleason, D. Hirschfeld, J.J. Kennedy-Smith, T. Mirzadegan, R. Roetz, M. Smith, S. Sperry, J.M. Suh, J. Wu, S. Tsing, ~ or, A. Paul, G. Su, G. Heilek, J.Q. Hang, A.S. Zhou, J.A. Jernelius, F.A.G. Villasen J. Zhang, K. Klumpp, J. Med. Chem. 51 (2008) 7449e7458. [6] C.L. Yeates, J.F. Batchelor, E.C. Capon, N.J. Cheesman, M. Fry, A.T. Hudson, M. Pudney, H. Trimming, J. Woolven, J.M. Bueno, J. Chicharro, E. Fern andez, mez de las Heras, E. Herreros, M.L. Leo n, J.M. Fiandor, D. Gargallo-Viola, F. Go J. Med. Chem. 51 (2008) 2845e2852. [7] Z. Zhan, J. Ai, Q. Liu, Y. Ji, T. Chen, Y. Xu, M. Geng, W. Duan, ACS Med. Chem. Lett. 5 (2014) 673e678. [8] A.K. Steib, O.M. Kuzmina, S. Fernandez, S. Malhotra, P. Knochel, Chem. Eur J. 21
461
(2015) 1961e1965. [9] Q. Zhou, B. Zhang, T. Du, H. Gu, Y. Ye, H. Jiang, R. Chen, Tetrahedron 69 (2013) 327e333. [10] M.D. Wendt, A.R. Kunzer, Tetrahedron Lett. 51 (2010) 3041e3044. [11] R.J. Burton, M.L. Crowther, N.J. Fazakerley, S.M. Fillery, B.M. Hayter, J.G. Kettle, C.A. McMillan, P. Perkins, P. Robins, P.M. Smith, E.J. Williams, G.L. Wrigley, Tetrahedron Lett. 54 (2013) 6900e6904. [12] B. Zhang, R. Chen, H. Jiang, Q. Zhou, F. Qiu, D. Han, R. Li, W. Tang, A. Zhong, J. Zhang, X. Yu, Tetrahedron 72 (2016) 2813e2817. [13] X. Zhao, X. Chen, G.-F. Yang, C.-G. Zhan, J. Phys. Chem. B 114 (2010) 6968e6972. [14] C. De Savi, R.J. Cox, D.J. Warner, A.R. Cook, M.R. Dickinson, A. McDonough, L.C. Morrill, B. Parker, G. Andrews, S.S. Young, P.S. Gilmour, R. Riley, M.S. Dearman, J. Med. Chem. 57 (2014) 4661e4676. [15] X. Zhao, X. Chen, C. Zhan, You Ji Hua Xue 29 (2009) 159e165. [16] X. Chen, X. Zhao, Y. Xiong, J. Liu, C.-G. Zhan, J. Phys. Chem. B 115 (2011) 12208e12219. [17] R. Dennington, T.A. Keith, J.M. Millam, Semichem Inc: Shawnee Mission, KS, 2009. [18] P.J. Stephens, F.J. Devlin, C.F. Chabalowski, M.J. Frisch, J. Phys. Chem. 98 (1994) 11623e11627. [19] A.D. Becke, J. Chem. Phys. 98 (1993) 5648e5652. [20] C. Lee, W. Yang, R.G. Parr, Phys. Rev. B 37 (1988) 785e789. [21] Y. Zhao, D.G. Truhlar, Theoretical Chemistry Accounts 120 (2008) 215e241. [22] Y. Zhao, D.G. Truhlar, Acc. Chem. Res. 41 (2008) 157e167. [23] M.J.T. Frisch, G.W., H.B. Schlegel, G.E. Scuseria, M.A. Robb, J.R. Cheeseman, G. Scalmani, V. Barone, B. Mennucci, G.A. Petersson, H. Nakatsuji, M. Caricato, X. Li, H.P. Hratchian, A.F. Izmaylov, J. Bloino, G. Zheng, J.L. Sonnenberg, M. Hada, M. Ehara, K. Toyota, R. Fukuda, J. Hasegawa, M. Ishida, T. Nakajima, Y. Honda, O. Kitao, H. Nakai, T. Vreven, J.A. Montgomery Jr., J.E. Peralta, F. Ogliaro, M. Bearpark, J.J. Heyd, E. Brothers, K.N. Kudin, V.N. Staroverov, R. Kobayashi, J. Normand, K. Raghavachari, A. Rendell, J.C. Burant, S.S. Iyengar, J. Tomasi, M. Cossi, N. Rega, J.M. Millam, M. Klene, J.E. Knox, J.B. Cross, V. Bakken, C. Adamo, J. Jaramillo, R. Gomperts, R.E. Stratmann, O. Yazyev, A.J. Austin, R. Cammi, C. Pomelli, J.W. Ochterski, R.L. Martin, K. Morokuma, V.G. Zakrzewski, G.A. Voth, P. Salvador, J.J. Dannenberg, S. Dapprich, € Farkas, J.B. Foresman, J.V. Ortiz, J. Cioslowski, D.J. Fox, A.D. Daniels, O. Gaussian, Inc.: Wallingford CT, 2009. [24] G.M. Morris, R. Huey, W. Lindstrom, M.F. Sanner, R.K. Belew, D.S. Goodsell, A.J. Olson, J. Comput. Chem. 30 (2009) 2785e2791. [25] Q. Zhou, B. Zhang, L. Su, T. Jiang, R. Chen, T. Du, Y. Ye, J. Shen, G. Dai, D. Han, H. Jiang, Tetrahedron 69 (2013) 10996e11003.
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Regioselective synthesis of 2- and 4-diarylpyridine ethers and their inhibitory activities against phosphodiesterase 4B Xinyun Zhao a, 1, Huihui Chen a, 1, Suting Xing a, Wei Yuan a, Lamei Wu a, Xi Chen a, *, Chang-Guo Zhan b, ** a b
College of Chemistry & Materials Science, South-Central University for Nationalities, Wuhan, 430074, PR China College of Pharmacy, University of Kentucky, Lexington, KY, 40536, USA
a r t i c l e i n f o
a b s t r a c t
Article history: Received 16 April 2019 Received in revised form 8 June 2019 Accepted 25 June 2019 Available online 28 June 2019
Two diaryl ether regioisomers of pyridine were prepared through the CuI/TMEDA/Cs2CO3-catalyzed reaction of 2,4-dibromopyridine with phenol derivatives under nitrogen atmosphere. The polar solvent DMSO gave 4-isomer as the major product whereas less polar toluene resulted in 2-isomer as a major product. Structures of regioisomers were confirmed by single crystal X-ray diffraction analysis. 4regioisomer shows higher biological activity against phosphodiesterase 4B (PDE4B) than that of 2isomer. Molecular docking simulations revealed that the PDE4B-inhibitory activity difference between the two regioisomers was mainly attributed to the atomic charge difference on the eOe linker. © 2019 Elsevier B.V. All rights reserved.
Keywords: Regioselectivity 2,4-Dibromopyridine Diaryl ethers PDE4B inhibitory activity X-ray crystal structure
1. Introduction Aryl ether is the core scaffold of many medicinal and/or natural product molecules, which are widely used in many fields including medicine and molecular biology. [1e7] Among these compounds, diaryl ethers containing pyridine moiety have attracted considerable attention due to their high biological activities. Thus, the preparation of such compounds is the main focus of research interest. The typical strategy for the synthesis of such compounds, follows aromatic nucleophilic substitution of 2,4-dihalogen pyridine (X ¼ F, Cl, Br). Regioselectivity of 2,4-dihalogen pyridine can introduce different functional groups at 2 or 4- position of pyridine ring. The reaction conditions and regioselectivity obtained for 2,4dihalogen pyridine are summarized in Table 1. As can be seen from this Table, aryl groups, [3,8] aryloxy groups, [9,10] arylamino groups, [11] arylacetylene groups [12] or other interesting functional groups can be selectively introduced to the 2- or 4- position of pyridine ring. For the synthesis of aryl ether with pyridine
* Corresponding author. ** Corresponding author. E-mail addresses: [email protected] (X. Chen), [email protected] (C.-G. Zhan). 1 These authors contributed equally to this work. https://doi.org/10.1016/j.molstruc.2019.06.086 0022-2860/© 2019 Elsevier B.V. All rights reserved.
moiety, 4-substituted product is preferred, when the reactions are performed under polar solvent environments. The exception is the nucleophilic substitution reaction of aryloxy groups carried out by Zhou and his coworkers [9] in which a 2-substituted aryloxypyridine product was reported to be a dominant product for the reaction in DMSO. Since 2- and 4- substituted aryloxypyridine products are regioisomers, which can lead to some confusion for characterization. However, for the design and/or synthesis of drug molecules, the accurate characterization of molecular structures is crucial. If a wrong structure was assigned, the structure-based computational molecule design would be meaningless. [13] So it is critical to study the regioselectivity of such type of reaction and unambiguously characterize the obtained products. Thus to reinvestigate the topic in detail, we firstly carried out the same reaction between 2,4-dibromopyridine and 4-methylphenol by using the same reaction conditions reported by Zhou and his coworkers (Scheme 1). [9] We further confirmed this regioselective behavior through proper X-ray crystal analysis of the product to avoid any ambiguity. Our experimental results showed that the 4-substituted aryloxypyridine product was the dominant product in DMSO solvent, and not the 2-substituted aryloxypyridines reported. In order to achieve 2-regioisomer with higher yield, the reaction of 2,4dibromopyridine with 4-methylphenol was performed in different solvents. In solvent toluene, another 2- regioisomer was a
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Table 1 Reported regioselective reactions of 2,4-dihalopyridine in literature.
X
R
Conditions
Preferential product
Ref
F/Cl Cl Cl Cl Br
OEt NHPh NHPh 4-CF3C6H4-
NaOEt/EtOH, DMF or DMSO Pd(OAc)2, Cs2CO3, Xantphos/microwave/100 C NaH/DMA 4-CF3C6H4MgBr/CrCl2 Pd(CF3COO)2/CuI/PPh3/i-Pr2NH
4-isomer 2-isomer 4-isomer 2-isomer 2-isomer
[10] [11] [11] [8] [12]
Br Br Br
Ar OAr 4-CH3C6H4O-
Pd(PPh3)4/aqTlOH CuI/TMEDA/Cs2CO3/DMSO CuI/TMEDA/Cs2CO3/DMSO
2-isomer 2-isomer 4-isomer
[3] [9] This work
Scheme 1. CuI catalyzed regioselective C-O coupling of 2,4-dibromopyridine with 4-methylphenol.Reagents and condition: CuI/TMEDA/Cs2CO3/toluene, N2, 110 C for (A), 78%; CuI/ TMEDA/Cs2CO3/DMSO, N2, 110 C for (B), 64%.
preferred product. The regioselectivity of the reaction of 2,4dibromopyridine with 4-methylphenol can be achieved by controlling the solvents polarity. Our experiments showed that 4substitued products was dominant in polar DMSO solvents, and the 2-regioisomers is dominant, if the reaction is performed in nonpolar toluene solvent. In addition, quantum mechanical calculations were carried out to find out the cause of regioselectivity for the reaction of 2,4-dibromopyridine and 4-methylphenol. This proven conclusion promoted us to further explore the regioselectivity of the reaction of 2,4-dibromopyridine with methyl (4-hydroxy-phenyl)acetate. Another two 2- and 4- regioisomers (1 and 2) were obtained in solvents DMSO and toluene (Scheme 2). We continued to synthesize two regioisomers 5 and 6 by following Suzuki reactions and hydrolysis reactions, which were our target molecules for biological inhibitory activity against phosphodiesterase 4B (PDE4B). [14e16] Further, molecular docking simulations were also performed to study the binding of regioisomers 5 and 6 with PDE4B enzyme and to explore the effects of different isomers on catalytic activity of PDE4B. 2. Experiments and calculation details 2.1. Chemical synthesis and biological activity tests All chemicals are commercially available. Solvents were dried in a routine way and redistilled. The NMR spectrum was recorded on a Varian 400 MHz NMR spectrometer. Chemical shift values are given in ppm relative to tetramethyl silane (TMS). High-resolution mass spectrometry (HRMS) was recorded on Agilent LC-Q-TOFMS-6520 instrument. Melting points were recorded on an X-6 digital melting point apparatus. The crystal structures of the title
compounds were determined on the Bruker Apex CCD X-ray instrument. 2.1.1. General procedure for the copper (I)-catalyzed regioselective coupling reaction of 2,4-dibromopyridine and methyl (4-hydroxyphenyl)acetate Synthesis of methyl 2-(4- ((2-bromo-4-pyridinyl)oxy) phenyl) acetate 1 in DMSO: CuI (18.6 mg, 0.098 mmol), TMEDA (11.4 mg, 0.098 mmol) were dissolved in dry DMSO (2 mL), and stirred at room temperature for 30 min under nitrogen atmosphere. cesium carbonate (638.6 mg, 1.96 mmol) and methyl (4-hydroxy-phenyl)-acetate (152.9 mg, 0.92 mmol) were added into the reaction mixture and stirred for another 4 h at room temperature. Finally, solution of 2,4dibromopyridine (236.8 mg, 0.99 mmol) in dried DMSO (3 mL) solvent was added to the reaction mixture. The reaction mixture was placed in oil-bath which was preheated to 110 C under nitrogen atmosphere. The reaction progress was monitored using TLC. After completion of the reaction, the reaction mixture was cooled to room temperature, and was filtered off. The mixture was diluted with dichloromethane (25 mL), which was washed with brine (3 30 mL). The organic phase was dried over sodium sulfate and the product was purified by column chromatography to give the title compound as pale yellow liquid. Yield: 75%. 1H NMR (400 MHz, CDCl3) d 8.21 (d, J ¼ 5.6 Hz, 1H), 7.36 (d, J ¼ 8.4 Hz, 2H), 7.05 (d, J ¼ 8.4 Hz, 2H), 6.99 (d, J ¼ 2.0 Hz, 1H), 6.83 (dd, J1 ¼ 2.0 Hz, J2 ¼ 5.6 Hz, 1H), 3.74 (s, 3H), 3.67 (s, 2H); 13 C NMR (101 MHz, CDCl3) d 171.7, 166.09, 152.6, 151.1, 143.0, 131.8, 131.3, 131.3, 120.9, 120.9, 115.7, 111.9, 52.2, 40.4. HRMS (ESI): Calcd. þ for C14H13BrNOþ 3 [MþH] : 322.0079; Found: 322.0073. Synthesis of methyl 2-(4-((4-bromo-2-pyridinyl)oxy) phenyl)
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457
Scheme 2. Reagents and reaction conditions: (a) CuI, TMEDA, Cs2CO3, DMSO, N2, 110 C; (b) CuI, TMEDA, Cs2CO3, Toluene, N2, 110 C; (c) PPh3, PdCl2, KOH, CH3CN, N2, 70 C; (d) KOH, CH3OH, RT.
acetate 2 in toluene: CuI (18.6 mg, 0.1 mmol) and TMEDA (11.4 mg, 0.1 mmol) was mixed in dried toluene (5 mL) and stirred for 30 min at room temperature. Cesium carbonate (638.6 mg, 2 mmol) and methyl (4hydroxy-phenyl)-acetate (152.7 mg, 0.92 mmol) were added into the mixture and stirred at room temperature for another 4 h. Finally, solution of 2,4-dibromopyridine (236.8 mg, 0.92 mmol) in dried toluene (3 mL) was added. The reaction mixture was placed in oil-bath which was preheated to 110 Cunder nitrogen atmosphere for 24 h. Reaction progress was monitored by TLC. On completion, reaction mixture was cooled to room temperature. The solvent was evaporated under vacuum and the residue was dissolved in dichloromethane (20 mL) and filtered and filtrate was purified by column chromatography, to obtain a pale yellow liquid (yield, 51%). 1 H NMR (400 MHz, CDCl3) d 8.01 (d, J ¼ 5.6 Hz, 1H), 7.33 (d, J ¼ 8.4 Hz, 2H), 7.15 (dd, J1 ¼1.2 Hz, J2 ¼ 5.2 Hz,1H), 7.11-7.09 (m, 3H), 3.71 (s, 3H), 3.64 (s, 2H); 13C NMR (101 MHz, CDCl3) d171.8, 164.3, 152.7, 148.2, 134.7, 130.8, 130.7, 130.7, 122.0, 121.4, 121.4, 114.7, þ 52.1, 40.6. HRMS (ESI): Calcd. for C14H13BrNOþ 3 [MþH] : 322.0079; Found: 322.0072.
2.1.2. Suzuki coupling reaction for synthesis of methyl 2-(4-((2-(5chloro-2-thiophenyl)-4-pyridinyl)oxy)phenyl)acetate 3 and methyl methyl 2-(4-((4-(5-chloro-2-thiophenyl)-2-pyridinyl)oxy)phenyl) acetate 4 PdCl2 (43.4 mg,0.24 mmol) and PPh3 (257 mg,0.98 mmol) were
dissolved in dried CH3CN(10 mL), KOH (274.4 mg,4.9 mmol) and 5chlorothiophene-2-boronic acid pinacol ester (659.7 mg,2.69 mmol) were added to the above mixture and then the intermediate (1 or 2) (790 mg, 2.45 mmol) was added, under nitrogen atmosphere. The reaction mixture was placed in oil-bath which was preheated to 70 C. Reaction progress was monitored by TLC. On completion, the reaction mixture was cooled to room temperature, filtered and filtrate was purified by chromatography to give the title compound. Compound 3: white solid, yield 74%, m.p. 120.1e121.7 C. 1H NMR (400 MHz, CDCl3) d 8.36 (d, J ¼ 5.6 Hz, 1H), 7.35 (d, J ¼ 8.4 Hz, 2H), 7.25 (d, J ¼ 4.0 Hz, 1H), 7.12 (d, J ¼ 2.4 Hz, 1H), 7.07 (d, J ¼ 8.4 Hz, 2H), 6.89 (d, J ¼ 4.0 Hz, 1H), 6.69 (dd, J ¼ 5.6, 2.4 Hz, 1H), 3.74 (s, 3H), 3.67 (s, 2H); 13C NMR (101 MHz, CDCl3) d171.8, 165.5, 153.5, 153.1, 151.0, 132.9, 131.3, 131.2, 131.2, 127.2, 127.2, 124.0, 120.8, 120.8, 110.8, 106.5, 52.2, 40.4; IR(KBr, cm1): 1625, 1501, 1432, 1317, 1088, 803, 692; HRMS (ESI): Calcd. for C18H15ClNO3Sþ [MþH]þ: 360.0461; Found: 360.0456. Compound 4: yellow solid, yield 48%. mp79.3e80.8 C. 1H NMR (400 MHz, CDCl3) d 8.14 (d, J ¼ 5.2Hz,1H), 7.33 (d, J ¼ 7.6Hz,2H), 7.27 (s,1H), 7.08e7.12 (m,3H), 7.00 (s,1H), 6.95 (d, J ¼ 3.6Hz,1H), 3.71 (s,3H), 3.64 (s,2H); 13C NMR (101 MHz, CDCl3) d171.8, 164.5, 153.1, 148.3, 144.1, 139.3, 132.1, 130.6, 130.4, 127.6, 125.0, 121.3, 114.8, 106.9, 52.0, 40.6; IR(KBr, cm1): 1752, 1604, 1550, 1513, 1442, 1401, 1303, 1234, 1157, 802, 751; HRMS (ESI): Calcd. for C18H15ClNO3Sþ [MþH]þ: 360.0461; Found: 360.0454.
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2.1.3. General procedure for synthesis of 2-(4-((2-(5-chloro-2thiophenyl) -4-pyridinyl)oxy)phenyl)acetic acid 5 and 2-(4-((4-(5chloro-2-thiophenyl) -2-pyridinyl)oxy)phenyl)acetic acid 6 Intermediates (3 or 4) (96.4 mg,0.27 mmol) was dissolved into methanol (6 mL), and aqueous KOH (30 mg, 0.54 mmol, dissolved into 1 mL of H2O) was added dropwise into the above ester solution,the mixture was stirred at room temperature. The reaction progress was monitored by TLC. On completion, the solvent was evaporated under vacuum and water was added into the residue. The solution was neutralized by diluted hydrochloride solution till pH 6-7 under ice-bath condition. The resulted solid was filtered and purified by column chromatography to give the target compounds 5 and 6. Compound 5: yellow solid, yield 82%. mp, 195.0e196.8 C, 1H NMR (400 MHz, DMSO‑d6) d 8.37 (d, J ¼ 5.6 Hz, 1H), 7.68 (d, J ¼ 4.0 Hz, 1H), 7.60 (s, 1H), 7.38 (d, J ¼ 8.4 Hz, 2H), 7.16 (t, J ¼ 5.9 Hz, 3H), 6.71 (dd, J ¼ 5.6, 2.4 Hz, 1H), 3.61 (s, 2H); 13C NMR (101 MHz, DMSO‑d6) d173.2, 173.2, 165.5, 153.4, 153.4, 152.8, 152.8, 151.8, 143.5, 14.5, 133.0, 131.8, 131.8, 131.3, 128.7, 128.7, 125.7, 125.7, 120.6, 111.2, 111.1, 107.3; IR(KBr, cm1): 2921, 1621, 1468, 1404, 1302, 1221, 1161, 1079, 890, 813; HRMS (ESI): Calcd. for C17H13ClNO3Sþ [MþH]þ: 346.0305; Found: 346.0298. Compound 6: yellow solid, yield 79%. mp, 164.5e166.4 C. 1H NMR (400 MHz, DMSO‑d6) d 12.38 (br, 1H), 8.13 (d, J ¼ 5.2 Hz, 1H), 7.76 (d, J ¼ 4.4 Hz, 1H), 7.35 (d, J ¼ 5.6 Hz, 1H), 7.32e7.26 (m, 4H), 7.09 (d, J ¼ 8.4 Hz, 2H), 3.59 (s, 2H); 13C NMR (101 MHz, DMSO‑d6) d173.2, 164.5, 152.9, 148.7, 144.0, 139.2, 131.8, 131.0, 130.9, 129.1, 127.3, 121.5, 115.4, 107.0, 40.5; IR(KBr, cm1): 3201, 1689, 1608, 1548, 1441, 1397, 1304, 1224, 1179, 1076, 788, 678; HRMS (ESI): Calcd. for C17H13ClNO3Sþ [MþH]þ: 346.0305; Found: 346.0300. 2.1.4. Synthesis of 4-bromo-2-p-tolyloxy-pyridine (A) and 2bromo-4-p-tolyloxy- pyridine (B) Synthesis of compound A. The procedure is the same as that of synthesis of compound 2 to give white solid. Yield 64%. mp: 47.0e48.9 C; 1H NMR (400 MHz, CDCl3) d 8.01 (d, J ¼ 5.2 Hz, 1H), 7.21 (d, J ¼ 8.0 Hz, 2H), 7.12 (d, J ¼ 5.6 Hz, 1H), 7.07 (s, 1H), 7.01 (d, J ¼ 8.0 Hz, 2H), 2.36 (s, 3H).13C NMR (101 MHz, CDCl3) d164.7, 151.3, 148.3, 134.8, 134.6, 130.4, 130.3, 121.7, 121.2, 121.2, 114.5, 20.9. HRMS (ESI): Calcd. for C12H11BrNOþ [MþH]þ: 264.0024; Found: 264.0018. Synthesis of compound B The procedure is the same as that of synthesis of compound 1 to give white solid. Yield 78%. Mp:69.2e70.8 C(63.1e64.7 C [9]); 1H NMR (CDCl3,400 MHz): d, 8.18 (d, J ¼ 6.0 Hz, 1H), 7.23 (d, J ¼ 8.0 Hz, 2H), 6.97 (m, 3H), 6.81e6.79 (m, 1H), 2.38 (s, 3H); 13C NMR (101 MHz, CDCl3) d166.3, 151.0, 151.0, 143.0, 135.8, 130.9, 130.9, 120.7, 120.6, 115.4, 111.7, 20.9. Calcd. for C12H11BrNOþ [MþH]þ: 264.0024; Found: 264.0020. 2.1.5. Biological assay against PDE4B PDE4B enzyme inhibitory data from BPS Bioscience Inc. (San Diego, California, USA) using fluorescence polarization method. Tested compounds were dissolved in DMSO and diluted in assay buffer (final DMSO concentration 1%, final inhibitor concentration 10 mM). 2.2. Molecular simulations 2.2.1. Quantum chemistry calculation Compounds A, B, and 5 were chosen for the study. The initial geometries of these molecules were firstly constructed by using the GaussView [17]. For molecules with multiple torsion bonds, the initial geometries were built in the most extended form. The constructed geometries were optimized by using the Density Functional Theory (DFT) Becke's three-parameter hybrid exchange functional and the Lee-Yang-Parr correlation functional (B3LYP)
[18e20] and 6-31 þ G* basis set, followed by vibrational frequency analysis at the same level of theory to ensure that the optimized geometries are indeed associated with local minima on the potential energy surface. To explore the origin of regioselectivity for the reaction of 2,4-dibromopyridine coupling with phenol derivatives, the optimized geometries of A and B were used to carry out single-point energy calculations using the M06-2X [21,22] method and the 6-311þþG(2d, 2p) basis set. For all the computation mentioned above, default criteria were used for the SCF iteration and for geometrical optimization. All the quantum chemistry calculations described above were performed by using Gaussian 09 software [23] installed on a local Linux 64 clusters. 2.2.2. Molecular docking In an effort to gain an understanding of the structural basis for the PDE4 inhibitory activities observed, we further studied the binding mode of these two compounds 5 and 6 through molecular docking. For this purpose, the crystal structure of PDE4B in complex with a [1,3,5]triazine derivative (PDB ID: 4kp6) was selected as the receptor for molecular docking. Before docking, the ligand and water molecules were removed from the complex structure, except for four water molecules and a hydroxide ion that bound with the metal ions Zn2þ and Mg2þ at the catalytic pocket. Then hydrogen atoms were added by using the Leap tools implemented in AMBER software. The molecular structure of compounds 5 and 6 were constructed by the GaussView followed by geometrical optimization at PM3 level. The optimized structures as well as the crystal structure of 5 were then used for docking study. For the receptor and each ligand, the nonpolar hydrogen atoms were merged and Gasteiger charges were added. Then AUTODOCK4.2 [24] program was used to search for the most favorable binding mode of the ligands and PDE4 catalytic domain. Coordinates of Mg2þ was set as grid center and grid size was set to 60 60 60 with spacing of 0.375 Å. During the docking process, atoms in the receptor were frozen. 100 docking runs were performed for each ligand and the conformations with the lowest binding free energies were selected for analysis. Default values were used for other docking parameters. 3. Results and discussion 3.1. Regioselectivity of 2,4-dibromopyridine for synthesis of diaryl ethers In order to further explore the regioselectivity of 2,4dibromopyridine coupling with phenol derivatives, an model reaction from Zhou's work carried out to investigate the regioselectivity of 2,4-dibromopyridine with 4-methylphenol in the presence of CuI/TMEDA/Cs2CO3 in DMSO or toluene (Scheme 1). [9] TLC was used to monitor the progress of the reactions and the separation of products was performed by column chromatography. Detailed solvent optimized reaction conditions can be seen in Table S1 of Supporting Information. The experimental procedure in DMSO was the same as that of the reported one. In DMSO reaction system, the starting material 2,4-dibromopyridine disappeared completely, but in solvent toluene, a small amounts of 2,4dibromopyridine was left and recovered. In solvent toluene, the isolated yields of 2-regioisomer (compound A, 64%), other 4regioisomer (compound B, 3.7%) and small amounts of disubstituted product in the solvent toluene were obtained. The isolated yields of 2-regioisomer (compound A, 3.7%), 4-regioisomer (compound B, 78%) and small amounts of disubstituted product in DMSO system were achieved. From the 1H NMR data of compounds A and B, the signal at 8.00 ppm appears in 2-regioisomeric derivative whereas the same signal is shifted to 8.17 ppm in 4-regioisomeric derivative (See NMR S1 and S3). Surprisingly, Zhou and
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coworkers have given the data similar to 4-regioisomer (for compound 3b, page 331 in Zhou's work) with signal at 8.17 ppm, though claiming as 2-regioisimer. The NMR results of compound B show that 1H NMR and 13C NMR data are consistent with Zhou's work in DMSO solvent. [9] In order to further confirm the regioselectivity of such kind of this reaction, single crystal of compound B suitable for X-ray analysis was grown in ethanol/acetone (1:1) mixture. Crystallographic data for the reported crystal structure was deposited at the Cambridge Crystallographic Data Centre with CCDC number 1534716 (Fig. 1). From Fig. 1, it is clear that the phenoxy group of compound B is located at the 4-position of pyridine, and not 2position of pyridine, as reported in literature. Our single crystal X-ray analysis on compound B exhibits greatly different from the reported one. In solvent DMSO, preferential C4-substituted isomer of pyridine (compound B in Scheme 1) was obtained. Detailed structural information about crystallographic data of compound B can be seen in Table S2, S3 and S4. In these experiments, preferential C2- substituted isomer of pyridine (compound A in Scheme 1) was obtained in solvent toluene with optimized yield 64%. Regioisomer A crystals suitable for single crystal X-ray diffraction analysis can't be obtained although many attempts were tried. Moreover, different regioisomers also exhibit different physical property such as polarity, which will eventually leading to different crystal stacking modes and melting points. As will be seen in the subsequent sections, regioisomers A and B show significantly different polarities for A (1.52 Debye) and for B (5.63 Debye), then they display different melting points for A (47.0e48.9 C) and for B (69.2e70.8 C). According to the above regioselective results, further structural modification was made by introducing 4-CH2COOMe into the benzene ring. Depicted in Scheme 2 are the regioselectively synthetic routes for diaryl ethers 1 and 2, as well as their downstream products 5 and 6. 4-phenoxy substituted regioisomer 1 and 2phenyloxy substituted regioisomer 2 was firstly prepared selectively through the copper-catalyzed regioselective coupling reaction of 2,4-dibromopyridine and methyl 4-hydroxyphenylacetate in solvents DMSO and toluene, respectively. This is similar to the optimized conditions of Scheme 1. For synthesis of compounds 1 and 2, in toluene solvent, the yields of 2-regioisomer (compound 2, 51%) and 4- regioisomer (compound 1, 14%) were obtained. In DMSO solvent, the yields of 2-regioisomer (compound 2, 11%) and 4- regioisomer (compound 1, 75%) were obtained. In DMSO and toluene system, small amounts of disubstituted product was obtained. Intermediates 3 and 4 were obtained through Suzuki coupling reactions between regioisomers 1 or 2 and 5chlorothiophene-2-boronic acid pinacol ester in the presence of PdCl2/PPh3 with KOH in dried CH3CN. [25] The difference of yields
Fig. 1. X-ray crystal structure of compound B from 2,4-dibromopyridine reacting with 4-methylphenol in CuI/TMEDA/Cs2CO3/DMSO under nitrogen atmosphere.
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for compounds 3 and 4 depends on the reactive activity of bromine atoms at the pyridine within the regioisomers 1 and 2. Different positions of aryloxy substitutents at pyridine lead to different reactive activity of bromine atoms. Compounds 5 and 6 were obtained by hydrolyzing the corresponding esters 3 and 4. Single crystal of regioisomer 5 was grown in methanol at 298 K. Then the crystal structure of 5 was determined by single crystal X-ray diffraction analysis (Fig. 2). The structure has been deposited in Cambridge Crystallographic Data Centre with deposition CCDC numbers:1534718. Other corresponding structural information can be seen in Tables S5, S6, S7 of Supporting Information. Numerous attempts failed to grow regioisomer 6 crystal suitable for single crystal X-ray diffraction analysis. From the structure of target compound 5, it is clear that the phenoxy group is located at the 4-position of pyridine ring. Since regioisomer 1 is the raw material for the synthesis of 5, regioisomer 1 should also be a 4-phenoxyl substituted regioisomer. This conclusion is inconsistent with the work reported by Zhou et al., in which the 2-regioisomer product was reported to be dominant under the same condition. [9] 3.2. Results of the DFT calculations. The DFT calculations at M06-2X/6-311þþG(2d, 2p) level demonstrated that the compound A was slightly more stable than B by ~0.2 kcal/mol. These calculations also show that the dipole moment of A is 1.52 Debye, which is much smaller than that of compound B (5.63 Debye). On the other hand, the dipole moment of toluene and DMSO are predicted to be 0.45 and 6.07 Debye, respectively. According to the principle of like dissolves like, compound A are more easy to dissolve in solvent toluene, and compound B are easy to dissolve in DMSO. Hence, in DMSO compound B gains more stability than A, which may lead to high yields of A in this solvent. In addition to the polarity of the solvent, there maybe are other factors leading to the difference between the yields of the products A and B. In addition, the optimized geometrical parameters of 5 were compared with the corresponding ones in the X-ray crystal structure. These comparisons are collected in Fig. 3. As can be seen from this figure, they match each other well for compound 5, suggesting that the quantum chemistry calculation results in this study are reliable. 3.3. Docking and PDE4B enzyme activity of diaryl ethers regioisomers The real structural assignment of aryloxy group on the pyridine core structure has been crucial for structure-based drug design and discovery. We have confirmed the real structure of diaryl ethers derivatives with a pyridine moiety using single crystal X-ray diffraction technology. Docking study of regioisomers 5 and 6 with PDE4 enzyme were performed. As can be seen from Fig. 4, the docking conformation obtained by using crystal structure of 5 (colored by purple scheme) is nearly identical to one obtained by using gas-phase optimized geometries of 5 (color by blue scheme), suggesting that the gas-phase optimized geometrical parameters are reliable. Molecular docking also reveals that target compounds 5 and 6 bind with PDE4 in similar binding modes (Fig. 4). Both ligands were fitted in a cavity formed by His234, Met347, Leu393, Asp392, Ser394, Asn395, Pro396, Thr397, Tyr403, Ile410, Phe414, Met431, Ser442, Gln443, Phe446, Ile450 and Val451 residues. The thiophene and pyridine rings of both ligands are lodged in the hydrophobic pocket surrounded by the side chains of Ile410, Ile450, Phe414, and Phe446 residues, causing a high degree of surface complementarities. The carboxylic groups were fitted in a
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Fig. 2. The crystal structure of the target compound 5.
Collected in Table 2 are the estimated binding free energies (DGcal bind) along with the corresponding experimental inhibitory ratio at 10 mM inhibitor concentration. As can be seen from the Table, the DGcal bind predicted by the AUTODOCK are consistent with the corresponding experimental data, suggesting that the present binding modes of these compounds are reasonable. Further energy decomposition analysis showed that the difference in DGcal bind is caused by the hydrogen bonding interactions formed between the -O- linker and the Gln443 glutamine switch (not shown). Charge analysis shows that the atomic charge (0.331) of this oxygen in 5 is significantly more negative than the counterpart (0.292) in 6, leading to stronger hydrogen bonding interactions in 5. 4. Conclusions Fig. 3. Selected geometrical parameters of 5 from (A) gas-phase geometry optimization at the B3LYP/6-31 þ G* level and (B) X-ray crystal structure.
Fig. 4. Binding modes of ligands 5 and 6 in the active site of PDE4. Ligands rendered with blue and green schemes are docking conformations obtained by using gas-phase optimized geometries of 5 and 6, respectively. Ligand rendered with purple scheme is docking conformation obtained by using experimental crystal structure of 5.
Diaryl ethers derivatives with pyridine moiety were regioselectively synthesized from 2,4-dihalopyridine through three steps sequence, including CuI-catalyzed regioselective reactions in different solvents, Suzuki coupling reactions and esters hydrolysis. The real structure of 4-regioisomer 5 was confirmed by single crystal X-ray diffraction analysis, i.e. 2,4-dibromopyridine can react with methyl 4-hydroxyphenylacetate to selectively give dominant 4-regioisomer of pyridine in the presence of CuI/TMEDA/Cs2CO3/ N2/DMSO. Different regioisomers 5 and 6 were docked into PDE4B binding pocket and showed different binding energy in spite of similar binding modes with different hydrogen bonding interaction contribution of -O- linker. 4-regioisomer 5 has higher inhibitory activity than that of 2-regioisomer 6 at 10 mM ligand concentration. In order to further confirm the CuI-catalyzed regioselectivity of 2,4dibromopyridine in DMSO, The reaction of 2,4-dibromopyridine and 4-methylphenol was carried out under the same reaction condition to give predominantly the 4-regioselective isomer which structure has been confirmed by single crystal X-ray diffraction analysis. The regioselectivity of 2,4-dihalopyridine in different solvents maybe correlate with the polarity of products diaryl ethers. It is clear that regioselectivity of a chemical reaction and real structures of medicinal molecules are very important for structurebased drugs design. Our current research will provide a reasonable basis for the synthesis of regioselective diaryl ethers and the design of diaryl ether type PDE4 inhibitors.
Table 2 Calculated binding free energies in comparison with available experimental data.
hydrophilic pocket formed by His234, Asp392, Ser394, Asn395, Thr397, and Tyr403. Despite the common features of the binding modes, the two thiophene rings flipped relatively to each other by about 180 .
Regioisomers 5 6 (b)
a DGcal bind
6.6 6.2
Inhibitory ratio obtained at 10 mM ligand concentration. Binding free energies predicted by AUTODOCK.
a
Inhibitory ratio (%) 64 26
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Acknowledgements The work was supported by the National Natural Science Foundation of China (grant number 21273089) and the Fundamental Research Funds for the Central Universities, South-Central University of Nationalities (CZW17004). Appendix A. Supplementary data Supplementary data to this article can be found online at https://doi.org/10.1016/j.molstruc.2019.06.086. References Debreczeni, R.A. Norman, [1] F.W. Goldberg, R.A. Ward, S.J. Powell, J.E. N.J. Roberts, A.P. Dishington, H.J. Gingell, K.F. Wickson, A.L. Roberts, J. Med. Chem. 52 (2009) 7901e7905. [2] R.J. Patch, L.L. Searle, A.J. Kim, D. De, X. Zhu, H.B. Askari, J.C. O'Neill, M.C. Abad, D. Rentzeperis, J. Liu, M. Kemmerer, L. Lin, J. Kasturi, J.G. Geisler, J.M. Lenhard, M.R. Player, M.D. Gaul, J. Med. Chem. 54 (2011) 788e808. mez, M.M. Cid, Tetrahedron 62 (2006) 11063e11072. [3] C. Sicre, J.L. Alonso-Go [4] N.A. Swain, D. Batchelor, S. Beaudoin, B.M. Bechle, P.A. Bradley, A.D. Brown, B. Brown, K.J. Butcher, R.P. Butt, M.L. Chapman, S. Denton, D. Ellis, S.R.G. Galan, S.M. Gaulier, B.S. Greener, M.J. de Groot, M.S. Glossop, I.K. Gurrell, J. Hannam, M.S. Johnson, Z. Lin, C.J. Markworth, B.E. Marron, D.S. Millan, S. Nakagawa, A. Pike, D. Printzenhoff, D.J. Rawson, S.J. Ransley, S.M. Reister, K. Sasaki, R.I. Storer, P.A. Stupple, C.W. West, J. Med. Chem. 60 (2017) 7029e7042. [5] Z.K. Sweeney, S.F. Harris, N. Arora, H. Javanbakht, Y. Li, J. Fretland, J.P. Davidson, J.R. Billedeau, S.K. Gleason, D. Hirschfeld, J.J. Kennedy-Smith, T. Mirzadegan, R. Roetz, M. Smith, S. Sperry, J.M. Suh, J. Wu, S. Tsing, ~ or, A. Paul, G. Su, G. Heilek, J.Q. Hang, A.S. Zhou, J.A. Jernelius, F.A.G. Villasen J. Zhang, K. Klumpp, J. Med. Chem. 51 (2008) 7449e7458. [6] C.L. Yeates, J.F. Batchelor, E.C. Capon, N.J. Cheesman, M. Fry, A.T. Hudson, M. Pudney, H. Trimming, J. Woolven, J.M. Bueno, J. Chicharro, E. Fern andez, mez de las Heras, E. Herreros, M.L. Leo n, J.M. Fiandor, D. Gargallo-Viola, F. Go J. Med. Chem. 51 (2008) 2845e2852. [7] Z. Zhan, J. Ai, Q. Liu, Y. Ji, T. Chen, Y. Xu, M. Geng, W. Duan, ACS Med. Chem. Lett. 5 (2014) 673e678. [8] A.K. Steib, O.M. Kuzmina, S. Fernandez, S. Malhotra, P. Knochel, Chem. Eur J. 21
461
(2015) 1961e1965. [9] Q. Zhou, B. Zhang, T. Du, H. Gu, Y. Ye, H. Jiang, R. Chen, Tetrahedron 69 (2013) 327e333. [10] M.D. Wendt, A.R. Kunzer, Tetrahedron Lett. 51 (2010) 3041e3044. [11] R.J. Burton, M.L. Crowther, N.J. Fazakerley, S.M. Fillery, B.M. Hayter, J.G. Kettle, C.A. McMillan, P. Perkins, P. Robins, P.M. Smith, E.J. Williams, G.L. Wrigley, Tetrahedron Lett. 54 (2013) 6900e6904. [12] B. Zhang, R. Chen, H. Jiang, Q. Zhou, F. Qiu, D. Han, R. Li, W. Tang, A. Zhong, J. Zhang, X. Yu, Tetrahedron 72 (2016) 2813e2817. [13] X. Zhao, X. Chen, G.-F. Yang, C.-G. Zhan, J. Phys. Chem. B 114 (2010) 6968e6972. [14] C. De Savi, R.J. Cox, D.J. Warner, A.R. Cook, M.R. Dickinson, A. McDonough, L.C. Morrill, B. Parker, G. Andrews, S.S. Young, P.S. Gilmour, R. Riley, M.S. Dearman, J. Med. Chem. 57 (2014) 4661e4676. [15] X. Zhao, X. Chen, C. Zhan, You Ji Hua Xue 29 (2009) 159e165. [16] X. Chen, X. Zhao, Y. Xiong, J. Liu, C.-G. Zhan, J. Phys. Chem. B 115 (2011) 12208e12219. [17] R. Dennington, T.A. Keith, J.M. Millam, Semichem Inc: Shawnee Mission, KS, 2009. [18] P.J. Stephens, F.J. Devlin, C.F. Chabalowski, M.J. Frisch, J. Phys. Chem. 98 (1994) 11623e11627. [19] A.D. Becke, J. Chem. Phys. 98 (1993) 5648e5652. [20] C. Lee, W. Yang, R.G. Parr, Phys. Rev. B 37 (1988) 785e789. [21] Y. Zhao, D.G. Truhlar, Theoretical Chemistry Accounts 120 (2008) 215e241. [22] Y. Zhao, D.G. Truhlar, Acc. Chem. Res. 41 (2008) 157e167. [23] M.J.T. Frisch, G.W., H.B. Schlegel, G.E. Scuseria, M.A. Robb, J.R. Cheeseman, G. Scalmani, V. Barone, B. Mennucci, G.A. Petersson, H. Nakatsuji, M. Caricato, X. Li, H.P. Hratchian, A.F. Izmaylov, J. Bloino, G. Zheng, J.L. Sonnenberg, M. Hada, M. Ehara, K. Toyota, R. Fukuda, J. Hasegawa, M. Ishida, T. Nakajima, Y. Honda, O. Kitao, H. Nakai, T. Vreven, J.A. Montgomery Jr., J.E. Peralta, F. Ogliaro, M. Bearpark, J.J. Heyd, E. Brothers, K.N. Kudin, V.N. Staroverov, R. Kobayashi, J. Normand, K. Raghavachari, A. Rendell, J.C. Burant, S.S. Iyengar, J. Tomasi, M. Cossi, N. Rega, J.M. Millam, M. Klene, J.E. Knox, J.B. Cross, V. Bakken, C. Adamo, J. Jaramillo, R. Gomperts, R.E. Stratmann, O. Yazyev, A.J. Austin, R. Cammi, C. Pomelli, J.W. Ochterski, R.L. Martin, K. Morokuma, V.G. Zakrzewski, G.A. Voth, P. Salvador, J.J. Dannenberg, S. Dapprich, € Farkas, J.B. Foresman, J.V. Ortiz, J. Cioslowski, D.J. Fox, A.D. Daniels, O. Gaussian, Inc.: Wallingford CT, 2009. [24] G.M. Morris, R. Huey, W. Lindstrom, M.F. Sanner, R.K. Belew, D.S. Goodsell, A.J. Olson, J. Comput. Chem. 30 (2009) 2785e2791. [25] Q. Zhou, B. Zhang, L. Su, T. Jiang, R. Chen, T. Du, Y. Ye, J. Shen, G. Dai, D. Han, H. Jiang, Tetrahedron 69 (2013) 10996e11003.