European Journal of Medicinal Chemistry 90 (2015) 45e52
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Short communication
Synthesis and in vitro antiproliferative activity of new 1,3,4-oxadiazole derivatives possessing sulfonamide moiety Mahmoud M. Gamal El-Din a, b, c, Mohammed I. El-Gamal d, Mohammed S. Abdel-Maksoud a, b, c, Kyung Ho Yoo e, Chang-Hyun Oh a, b, * a
Center for Biomaterials, Korea Institute of Science and Technology, PO Box 131, Cheongryang, Seoul 130-650, Republic of Korea Department of Biomolecular Science, University of Science and Technology, 113 Gwahangno, Yuseong-gu, Daejeon 305-333, Republic of Korea Pharmaceutical and Drug Industries Research Division, National Research Centre, Dokki, Giza 12622, Egypt d Department of Medicinal Chemistry, Faculty of Pharmacy, University of Mansoura, Mansoura 35516, Egypt e Chemical Kinomics Research Center, Korea Institute of Science and Technology, PO Box 131, Cheongryang, Seoul 130-650, Republic of Korea b c
a r t i c l e i n f o
a b s t r a c t
Article history: Received 10 June 2014 Received in revised form 3 November 2014 Accepted 5 November 2014 Available online 6 November 2014
Synthesis of a new series of 1,3,4-oxadiazole derivatives possessing sulfonamide moiety is described. Their in vitro antiproliferative activities against NCI-58 human cancer cell lines of nine different cancer types were tested. Compound 1k with p-methoxybenzenesulfonamido moiety showed the highest mean %inhibition value over the 58 cell line panel at 10 mM concentration. It showed broad-spectrum antiproliferative activity over many cell lines of different cancer types. For instance, compound 1k inhibited the growth of T-47D breast cancer cell line by 90.47% at 10 mM. And it inhibited growth of SR leukemia, SK-MEL-5 melanoma, and MDA-MB-468 breast cancer cell lines by more than 80% at the same test concentration. Compound 1k showed superior activity than Paclitaxel and Gefitinib against the most sensitive cell lines. © 2014 Elsevier Masson SAS. All rights reserved.
Keywords: Antiproliferative activity 1,3,4-Oxadiazole Sulfonamide
1. Introduction Cancer is a collection of different life-threatening diseases characterized by uncontrolled growth of cells, leading to invasion of surrounding tissue and often metastasizing to other parts of the body. According to the statistics, cancer is the second most common cause of death worldwide after cardiovascular diseases [http:// www.cdc.gov/nchs/fastats/lcod.htm]. Despite the availability of improved drugs including targeted cancer therapies, the worldwide cancer burden is expected to increase by as much as 15 million new cancer cases per year by 2020, according to the World Health Organization (WHO), unless further preventive measures are put into practice [1,2]. The development of new anticancer drugs represents a major interest and challenge to the contemporary medicinal chemistry. Much attention has been paid to the chemistry and biological activities of 1,3,4-oxadiazole nucleus. Several compounds possessing 1,3,4-oxadiazole scaffold have been recently reported as potential antiproliferative agents [3e11]. Apart from anticancer
* Corresponding author. Center for Biomaterials, Korea Institute of Science and Technology, PO Box 131, Cheongryang, Seoul 130-650, Republic of Korea. E-mail address: [email protected] (C.-H. Oh). http://dx.doi.org/10.1016/j.ejmech.2014.11.011 0223-5234/© 2014 Elsevier Masson SAS. All rights reserved.
activity, other biological activities have been reported for 1,3,4oxadiazole derivatives such as antidiabetic [12], antitubercular [13], antifungal [14], antiinflammatory [15], and antibacterial activities [16]. Furthermore, many sulfonamide derivatives have been highlighted as anticancer agents [17e22]. In the present study, we report synthesis of a new series of 1,3,4oxadiazole derivatives possessing terminal sulfonamide moiety. Their in vitro antiproliferative activities against NCI-58 cancer cell line panel are reported.
2. Results and discussion 2.1. Chemistry Synthesis of the target compounds 1aeq was achieved through the pathway illustrated in Scheme 1. Refluxing the benzoate esters 2aec with hydrazine monohydrate in ethanol afforded the corresponding benzohydrazide derivatives 3aec [23]. Cyclization to 2(chloromethyl)-5-aryl-1,3,4-oxadiazole analogs 4aec was carried out through refluxing the hydrazides 3aec with chloroacetic acid in phosphorus oxychloride [24]. Nucleophilic substitution of chloro group of compounds 4aec with 2-fluoro-4-nitrophenol was done by reflux in acetonitrile in the presence of potassium carbonate to
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Scheme 1. Reagents and conditions: a) hydrazine monohydrate, EtOH, reflux, overnight; b) ClCH2COOH, POCl3, reflux, 6 h; c) 2-fluoro-4-nitrophenol, K2CO3, acetonitrile, reflux, 36 h; d) Raney Ni, H2, THF, rt, 24 h; e) alkyl or aryl sulfonyl chloride, C5H5N, CH2Cl2, rt, overnight.
obtain the ether derivatives 5aec. The nitro group of compounds 5aec was reduced to amino using Raney Nickel in hydrogen atmosphere. Treatment of the aniline derivatives 6aec with the appropriate alkyl or aryl sulfonyl chloride derivatives in presence of pyridine as a base afforded the target sulfonamide derivatives 1aeq. 2.2. In vitro antiproliferative activity Structures of the synthesized target compounds were submitted to National Cancer Institute (NCI), Bethesda, Maryland, USA [25], and the thirteen compounds shown in Table 2 were selected on the basis of degree of structural variation and computer modeling techniques for evaluation of their antineoplastic activity. The selected compounds were subjected to in vitro anticancer assay against tumor cells in a full panel of 58 cell lines taken from nine different tissues (blood, lung, colon, CNS, skin, ovary, kidney, prostate, and breast). The compounds were tested at a single-dose concentration of 10 mM, and the percentages of growth inhibition over the 58 tested cell lines were determined. The mean % growth of the NCI-58 cancer cell line panel after treatment with each of the tested compounds is illustrated in Table 1. Upon comparing the effect of aryl ring directly attached to 1,3,4oxadiazole ring on activity, it was found that compounds 1m and 1o possessing p-methoxyphenyl were more active than compounds 1b and 1d with phenyl ring. Compounds 1h and 1k containing pchlorophenyl showed higher antiproliferative activity than the corresponding p-methoxyphenyl analogs 1n and 1q. Compound 1g with p-chlorophenyl was more active that the corresponding methoxy derivative 1l, and 1l exerted higher activity than 1a with unsubstituted phenyl ring. So it can be concluded that p-chlorophenyl is the most optimum and unsubstituted phenyl is the least optimum for activity of this series of compounds. This can be attributed to steric and/or electronic differences between chloro, methoxy, and hydrogen groups. The effect of alkyl(aryl)sulfonamide moiety on activity was also investigated. Among aliphatic derivatives, n-propylsulfonamido compounds 1b and 1m were more active than the corresponding methylsulfonamido analogs 1a and 1l. So homologation to a longer chain alkyl group could be favorable for activity, may be due to
stronger hydrophobic interaction at the receptor site and/or the increased lipophilicity enhanced penetration into the cancer cells. The influences of aliphatic and aromatic sulfonamido moieties on activity were compared. The aromatic derivatives were found generally more active than aliphatic analogs (compound 1d more active than 1a and 1b), (compounds 1h, 1j, and 1k more active than 1g), and (compounds 1oeq more active than 1l and 1m). Furthermore, substituted phenyl moieties were more optimal for activity than unsubstituted phenyl (compounds 1j and 1k were more active than 1h, and 1oeq showed higher activity than 1n). The pmethoxybenzenesulfonamido compounds 1k and 1q exerted superior activity compared to the corresponding p-toluenesulfonamido derivatives 1j and 1p. This may be rationalized that methoxy Table 1 Mean %growth of the 58 cancer cell line panel after treatment with the tested target compounds (10 mM concentration).
Compound no.
R1
R2
Mean %growth
1a 1b 1d 1g 1h 1j 1k 1l 1m 1n 1o 1p 1q
H H H Cl Cl Cl Cl OMe OMe OMe OMe OMe OMe
CH3 n-Pr 4-Cl(C6H4) CH3 C6H5 4-Tolyl 4-MeO(C6H4) CH3 n-Pr C6H5 4-Cl(C6H4) 4-Tolyl 4-MeO(C6H4)
100.73 98.89 97.14 93.74 91.95 84.99 57.53 97.56 91.00 97.55 73.29 83.27 72.40
The bold figure indicates the most active compound.
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Table 2 %Inhibition results exerted by the most potential compounds over the most sensitive cell lines.
Compound no.
R1
R2
Cell line
Cancer type
% Inhibition at 10 mM
1k
4-Cl
4-OMe(C6H4)
1o
4-OMe
4-Cl(C6H4)
1p
4-OMe
4-Tolyl
1q
4-OMe
4-OMe(C6H4)
K-562 MOLT-4 RPMI-8226 SR A549 NCI-H23 NCI-H522 SK-MEL-2 SK-MEL-5 UACC-257 UACC -62 T-47D MDA-MB-468 MCF-7 A-498 NCI-H522 T-47D MDA-MB-468 RPMI-8226 T-47D MDA-MB-468 UACC-62 RPMI-8226 CCRF-CEM NCI-H522 SK-MEL 5 T-47D
Leukemia Leukemia Leukemia Leukemia Lung cancer Lung cancer Lung cancer Melanoma Melanoma Melanoma Melanoma Breast cancer Breast cancer Breast cancer Renal cancer Lung cancer Breast cancer Breast cancer leukemia Breast cancer Breast cancer Melanoma Leukemia Leukemia Lung cancer Melanoma Breast cancer
56.82 63.99 68.36 81.58 52.58 56.58 56.57 59.40 84.32 76.66 57.66 90.47 84.83 57.22 53.03 72.66 59.48 54.52 55.35 71.88 40.25 40.96 56.64 56.91 50.84 56.06 68.44
The bold figures indicate the highest growth inhibition percentages.
group on this terminal ring may be essential as hydrogen bond acceptor at the receptor site. This enhances affinity with the receptor, and hence the antiproliferative activity improves. Among all the target derivatives, compound 1k with p-chlorophenyl on the oxadiazole ring and p-methoxybenzenesulfonamido terminal moiety showed the best activity. So those moieties together are the most favorable for antiproliferative activity of this series of compounds.
The %inhibition values of the most active compounds over the most sensitive cell lines are summarized in Table 2. T-47D breast cancer cell line was the most sensitive cell line to this series of compounds. The most active compounds against T-47D were 1k and 1p with %inhibitions of 90.47% and 71.88%, respectively. MDAMB-468 breast cancer cell line was also sensitive to the most potential compounds. Fig. 1 illustrates the %inhibition values of the tested target compounds against T-47D and MDA-MB-468 cell lines. Compounds 1k and 1o were active against both cell lines,
Fig. 1. Inhibitory effects of the tested target compounds at 10 mM concentration over T-47D and MDA-MB-468 breast cancer cell lines.
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Fig. 2. % Inhibition expressed by compounds 1k (Fig. 2a), 1o (Fig. 2b), and 1q (Fig. 2c) at a single-dose concentration of 10 mM over all cell lines of the NCI cancer cell line panel of nine different cancer types.
while compounds 1p and 1q showed higher inhibitory effect over T-47D. Among all the target compounds, compound 1k showed the most promising results. It exerted broad-spectrum antiproliferative activity against different cell lines of different cancer types; leukemia, melanoma, and breast cancer. So it can be considered as a
potential lead compound for future development of broadspectrum anticancer agents. Among all the tested derivatives, compounds 1k, 1o, and 1q showed the highest mean inhibitions. The percentages of inhibition of these four compounds over each tested cell line of the NCI-58 panel at 10 mM concentration are depicted in Fig. 2. The three
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tetramethylsilane as an internal standard. Melting points were obtained on a Walden Precision Apparatus Electrothermal 9300 apparatus and are uncorrected. Solvents and liquid reagents were transferred using hypodermic syringes. Purity % of all the target compounds were determined by HPLC and found to be >95%. All solvents and reagents were commercially available and used without further purification. 4.2. Synthesis of benzohydrazide derivatives 3aec [23]
Fig. 3. Comparison of %inhibition values expressed by compound 1k, Paclitaxel, and Gefitinib against the most sensitive cell lines towards compound 1k.
compounds exerted broad-spectrum antiproliferative activities against different cell lines of different cancer types. Among them, compound 1k was the most active. It showed the highest %inhibition values with more than 70% inhibition against SR leukemia cell line, SK-MEL-5 and UACC-257 melanoma cell lines, and T-47D and MDA-MB-468 breast cancer cell lines. The results of compound 1k against those five cell lines were compared with Paclitaxel and Gefitinib as reference standard drugs, as illustrated in Fig. 3. The results of Paclitaxel and Gefitinib were obtained from NCI datawarehouse index [26], and are inserted in Fig. 3. Compound 1k was more active than both reference compounds against SK-MEL-5 and T-47D cell lines. It showed higher activity also than Gefitinib against SR and UACC-257 cell lines. 3. Conclusion In this study, a series of new 1,3,4-oxadiazole derivatives possessing terminal sulfonamide moiety was synthesized. The target compounds were tested for in vitro antiproliferative activities over NCI-58 cancer cell line panel of nine different cancer types. Among them, compound 1k possessing p-chlorophenyl ring attached to 1,3,4-oxadiazole ring and p-methoxybenzenesulfonamido terminal moiety showed the most promising results at a single-dose concentration of 10 mM. It exerted broad-spectrum antiproliferative activities over different cell lines of different cancer types. The % inhibition values of compound 1k over T-47D breast, SR leukemia, SK-MEL-5 melanoma, and MDA-MB-468 breast cancer cell lines were 90.47%, 81.58%, 84.32%, and 84.83%, respectively. T-47D and MDA-MB-468 breast cancer cell lines were found to be highly sensitive to the most active compounds. The in vitro antiproliferative activity indicated that compound 1k could be a promising lead for future development of active anticancer agents. It was more active than Paclitaxel and Gefitinib against SK-MEL-5 melanoma and T-47D breast cancer cell lines. It exerted also higher activity than Gefitinib against SR leukemia and UACC-257 melanoma cell lines. Further modifications of this series of 1,3,4oxadiazole compounds in order to improve their antiproliferative activity are currently in progress. 4. Experimental 4.1. General The target compounds were purified by column chromatography using silica gel (0.040e0.063 mm, 230e400 mesh) and technical grade solvents. 1H NMR and 13C NMR spectra were recorded on a Bruker Avance 400 or 300 spectrometer using
To a solution of the appropriate methyl benzoate ester (1.0 mmol) in ethanol (30 mL), hydrazine monohydrate (0.15 g, 3.0 mmol) was added. The reaction mixture was heated under reflux overnight. After completion of the reaction, the solvent was evaporated under reduced pressure, and the residue was washed with water (2 3 mL), and the obtained solid was filtered and dried to give benzohydrazide derivatives 3aec. Benzohydrazide (3a): yield 70%; 1H NMR (400 MHz, DMSO-d6) d 9.80 (s, 1H), 7.84 (d, 2H, J ¼ 8.0 Hz), 7.47e7.45 (m, 3H), 4.52 (s, 2H); 13 C NMR (100 MHz, DMSO-d6) d 165.9, 133.3, 131.0, 128.3, 126.9. 4-Chlorobenzohydrazide (3b): yield 82.4%; 1H NMR (400 MHz, DMSO-d6) d 9.86 (s, 1H), 7.84 (d, 2H, J ¼ 8.8 Hz), 7.53 (d, 2H, J ¼ 8.4 Hz), 4.52 (s, 2H); 13C NMR (100 MHz, DMSO-d6) d 164.7, 135.8, 132.0, 128.8, 128.4. 4-Methoxybenzohydrazide (3c): yield 87%; 1H NMR (400 MHz, DMSO-d6) d 9.65 (s, 1H), 7.83 (d, 2H, J ¼ 8.8 Hz), 6.98 (d, 2H, J ¼ 8.8 Hz), 4.47 (s, 2H), 3.80 (s, 3H); 13C NMR (100 MHz, DMSO-d6) d 170.9, 166.6, 133.9, 130.7, 118.7, 60.5. 4.3. Synthesis of 2-(chloromethyl)-5-substituted phenyl-1,3,4oxadiazole (4aec) To a mixture of compound 3aec (3.0 mmol) and chloroacetic acid (0.26 g, 2.75 mmol), phosphorus oxychloride (1.5 g, 1 mL, 10 mmol) was added and the mixture was refluxed for 5e6 h. The reaction mixture was cooled to rt then with ice bath, and saturated solution of sodium bicarbonate was added dropwise thereto till pH 9. The precipitated product was filtered, washed with plenty of water, and then dried to give the title compounds. 2-(Chloromethyl)-5-phenyl-1,3,4-oxadiazole (4a): yield 63.5%; 1 H NMR (400 MHz, DMSO-d6) d 8.03 (dd, 2H, J ¼ 1.6, J ¼ 8.4 Hz), 7.70e7.61 (m, 3H), 5.16 (s, 2H); 13C NMR (100 MHz, DMSO-d6) d 164.9, 162.8, 132.4, 129.5, 126.7, 122.8, 33.2. 2-(Chloromethyl)-5-(4-chlorophenyl)-1,3,4-oxadiazole (4b): yield 71.3%; 1H NMR (400 MHz, DMSO-d6) d 8.00 (d, 2H, J ¼ 8.8 Hz), 7.66 (d, 2H, J ¼ 8.8 Hz), 5.15 (s, 2H); 13C NMR (100 MHz, DMSO-d6) d 164.2, 162.9, 137.1, 129.6, 128.4, 121.7, 33.2. 2-(Chloromethyl)-5-(4-methoxyphenyl)-1,3,4-oxadiazole (4c): yield 72.5% 1H NMR (400 MHz, DMSO-d6) d 7.95 (d, 2H, J ¼ 8.8 Hz), 7.14 (d, 2H, J ¼ 8.8 Hz), 5.13 (s, 2H); 13C NMR (100 MHz, DMSO-d6) d 164.9, 162.2, 128.5, 128.4, 115.1, 114.7, 55.5, 33.2. 4.4. Synthesis of 2-((2-fluoro-4-nitrophenoxy)methyl)-5substitutedphenyl-1,3,4-oxadiazole (5aec) To a solution of compound 4aec (0.5 mmol) and 2-fluoro-4nitrophenol (0.1 g, 0.6 mmol) in dry acetonitrile (50 mL), anhydrous potassium carbonate (0.2 g, 1.5 mmol) was added and the mixture was refluxed for 24e36 h till completion of the reaction. The solvent was evaporated under reduced pressure till dryness, and the residue was partitioned between water (50 mL) and ethyl acetate (50 mL). The organic layer was separated, and the aqueous layer was extracted with water (3 50 mL). The combined organic extracts were washed with brine and then dried over anhydrous sodium sulfate. The solvent was evaporated under reduced
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pressure to the get the corresponding nitro compound. The product was crystalized from methanol:ethyl acetate 4:1 v/v to give the pure title compounds. 4.4.1. 2-((2-Fluoro-4-nitrophenoxy)methyl)-5-phenyl-1,3,4oxadiazole (5a) yield 61.2%; mp 139e141 C; 1H NMR (400 MHz, DMSO-d6) d 8.27e8.20 (m, 2H), 8.05 (d, 2H, J ¼ 8.4 Hz), 7.71e7.63 (m, 4H), 5.84 (s, 2H); 13C NMR (100 MHz, DMSO-d6) d 164.9, 161.6, 151.7, 150.9 (d, JCF ¼ 10 Hz), 149.2, 141.2 (d, JCF ¼ 8 Hz), 132.3, 129.5, 126.7, 122.9, 121.2 (d, JCF ¼ 4 Hz), 115.1, 112.5, 112.2, 60.9. 4.4.2. 2-(4-Chlorophenyl)-5-((2-fluoro-4-nitrophenoxy)methyl)1,3,4-oxadiazole (5b) yield 47.3%; mp 142e143 C; 1H NMR (400 MHz, DMSO-d6) d 8.24e8.17 (m, 2H), 8.03 (d, 2H, J ¼ 8 Hz), 7.70e7.65 (m, 3H), 5.80 (s, 2H); 13C NMR (100 MHz, DMSO-d6) d 164.2, 161.8, 151.7, 150.8 (d, JCF ¼ 11 Hz), 149.2, 141.2 (d, JCF ¼ 8 Hz), 137.1, 129.7, 128.5, 121.8, 121.2 (d, JCF ¼ 4 Hz), 115.1, 112.5, 112.2, 60.9. 4.4.3. 2-((2-Fluoro-4-nitrophenoxy)methyl)-5-(4-methoxyphenyl)1,3,4-oxadiazole (5c) yield 55.6%; mp 136e138 C; 1H NMR (400 MHz, DMSO-d6) d 8.23e8.16 (m, 2H), 7.95 (d, 2H, J ¼ 8.8 Hz), 7.66 (t, 1H, J ¼ 8.4 Hz), 7.15 (d, 2H, J ¼ 9.2 Hz), 5.77 (s, 2H), 3.86 (s, 3H); 13C NMR (100 MHz, DMSO-d6) d 164.9, 162.3, 161.0, 151.7, 150.9 (d, JCF ¼ 10 Hz), 149.2, 141.2 (d, JCF ¼ 8 Hz), 128.5, 121.2 (d, JCF ¼ 3 Hz), 115.1, 115.0, 114.9, 112.4, 112.2, 60.9, 55.5. 4.5. Synthesis of 3-fluoro-4-(5-substituted phenyl-1, 3, 4-oxadiazol2-yl)methoxy)benzamine (6aec) To a solution of the compound 5aec (0.5 mmol) in a mixture of THF and methanol (2:1 v/v, 150 mL), freshly activated Raney nickel (10% eq) was added. The mixture was stirred in hydrogen atmosphere at rt for 24 h. The catalyst was cautiously filtered, and the solvent was removed under reduced pressure to give the amine in quantitative yield. 4.5.1. 3-Fluoro-4-((5-phenyl-1,3,4-oxadiazol-2-yl)methoxy)aniline (6a) yield 85%; mp 132e133 C; 1H NMR (400 MHz, CD3OD) d 8.08 (d, 2H, J ¼ 6.4 Hz), 7.63e7.59 (m, 3H), 7.00 (t, 1H, J ¼ 8.4 Hz), 6.52e6.45 (m, 2H), 5.30 (s, 2H); 13C NMR (100 MHz, CD3OD) d 167.2.9, 164.7, 157.1, 154.6, 146.5 (d, JCF ¼ 10 Hz), 138.3 (d, JCF ¼ 7 Hz), 133.5, 130.5, 130.4, 128.0, 124.6, 121.5, 111.9 (d, JCF ¼ 4 Hz), 104.5, 64.4. 4.5.2. 4-((5-(4-Chlorophenyl)-1,3,4-oxadiazol-2-yl)methoxy)-3fluoroaniline (6b) yield 89%; mp 143e145 C; 1H NMR (400 MHz, DMSO-d6) d 8.05 (d, 2H, J ¼ 6.4 Hz), 7.46 (t, 1H, J ¼ 8.4 Hz), 6.56 (d, 1H, J ¼ 8.4 Hz), 6.46 (d, 1H, J ¼ 8.8 Hz), 5.27 (s, 2H); 13C NMR (100 MHz, DMSO-d6) d 166.8, 165.7, 162.3, 157.1, 144.6 (d, JCF ¼ 11 Hz), 138.2 (d, JCF ¼ 10 Hz), 121.4, 118.0, 115.5, 111.0 (d, JCF ¼ 4 Hz), 105.0, 104.8, 63.6. 4.5.3. 3-Fluoro-4-((5-(4-methoxyphenyl)-1,3,4-oxadiazol-2-yl) methoxy)aniline (6c) yield 93.2%; mp 92e94 C; 1H NMR (400 MHz, DMSO-d6) d 8.01 (d, 2H, J ¼ 8.4 Hz), 6.94 (d, 2H, J ¼ 8.4 Hz), 6.91 (t, 1H, J ¼ 8.4 Hz), 6.45 (dd, 1H, J ¼ 2.4, J ¼ 12.8 Hz), 6.34 (d, 1H, J ¼ 8.8 Hz), 5.24 (s, 2H), 3.88 (s, 3H), 2.15 (s, 2H); 13C NMR (100 MHz, DMSO-d6) d 165.8, 162.6, 161.9, 155.6, 143.3 (d, JCF ¼ 9 Hz), 137.6 (d, JCF ¼ 12 Hz), 120.2, 116.0, 114.5, 110.5 (d, JCF ¼ 3 Hz), 104.0, 103.8, 63.3, 55.5.
4.5.4. N-(3-fluoro-4-((5-phenyl-1,3,4-oxadiazol-2-yl)methoxy) phenyl)methanesulfonamide (1a) To a solution of the compound 6a (0.18 mmol) in anhydrous THF, catalytic amount of pyridine was added in ice cooled bath and was stirred for 15 min then Methanesulfonyl chloride (0.025 g, 0.22 mmol) was added and the reaction mixture was stirred at room temperature for 24 h. When the reaction was completed, the solvent was completely removed and the residue was partitioned between water and ethyl acetate. The organic layer was separated and the aqueous layer was extracted with ethyl acetate (3 5 mL) and the combined organic extracts was washed with brine, separated, dried over anhydrous sodium sulfate after evaporation of the solvent, the residue was purified by column chromatography (silica gel, hexane-ethyl acetate 3:1 v/v) to give the compound as solid 25 mg 39% yield; mp 163e165 C; 1H NMR (300 MHz, CDCl3) d 8.08 (d, 2H, J ¼ 6.1 Hz), 7.54 (d, 2H, J ¼ 6.1 Hz), 7.17 (t, 2H, J ¼ 9.2 Hz), 6.94 (d, 1H, J ¼ 6.3 Hz), 6.70 (s, 1H), 5.39 (s, 2H), 3.00 (s, 3H); 13C NMR (100 MHz, CD3OD) d 167.3, 164.1, 155.7, 153.3, 144.0 (d, JCF ¼ 12 Hz), 135.0 (d, JCF ¼ 9 Hz), 133.5, 130.5, 128.1, 124.5, 119.2, 118.1 (d, JCF ¼ 3 Hz), 111.1, 63.1, 39.2. Compounds 1beq were prepared with the same method from 1a and the appropriate sulfonyl chloride derivative. 4.5.5. N-(3-fluoro-4-((5-phenyl-1, 3, 4-oxadiazol-2-yl)methoxy) phenyl)propan-1-sulfonamide (1b) Yield 41%; mp 169e171 C; 1H NMR (400 MHz, CDCl3) d 8.08 (d, 2H, J ¼ 8.4 Hz), 7.57e7.50 (m, 3H), 7.17e7.13 (m, 2H), 6.90 (d, 2H, J ¼ 8.4 Hz), 6.46 (s, 1H), 5.38 (s, 2H), 3.06e3.02 (m, 2H), 1.87e1.82 (m,2H), 1.03 (t, 3H, J ¼ 7.2 Hz); 13C NMR (100 MHz, CDCl3) d 166.1, 161.9, 154.5, 152.0, 142.9 (d, JCF ¼ 11 Hz), 132.6 (d, JCF ¼ 9 Hz), 132.3, 129.2, 127.2, 123.3, 118.1, 116.9 (d, JCF ¼ 2 Hz), 110.4, 62.0, 53.4, 17.2, 12.8. 4.5.6. N-(3-fluoro-4-((5-phenyl-1,3,4-oxadiazol-2-yl)methoxy) phenyl)benzenesulfonamide (1c) Yield 34%; mp 137e139 C; 1H NMR (400 MHz, CDCl3) d 8.08 (d, 2H, J ¼ 7.9 Hz), 7.58 (d, 2H, J ¼ 8.1 Hz), 7.63e7.43 (m, 5H), 7.40 (t, 2H, J ¼ 4.3 Hz), 7.02 (t, 2H, J ¼ 4.4 Hz), 5.33 (s, 2H); 13C NMR (100 MHz, CDCl3) d 166.0, 161.9, 154.2, 151.7, 143.2 (d, JCF ¼ 11 Hz), 138.7, 133.1, 132.3, 132.0 (d, JCF ¼ 9 Hz), 129.1, 127.1, 123.2, 118.2 (d, JCF ¼ 5 Hz), 117.5, 111.6, 111.4, 61.9. 4.5.7. 4-Chloro-N-(3-fluoro-4-((5-phenyl-1,3,4-oxadiazol-2-yl) methoxy)phenyl)benzenesulfonamide (1d) Yield 43%; mp 115e117 C; 1H NMR (400 MHz, CDCl3) d 8.05 (d, 2H, J ¼ 8 Hz), 7.83 (s, 1H), 7.67 (d, 2H, J ¼ 8.8 Hz), 7.58e7.49 (m,3H), 7.37 (d, 2H, J ¼ 8.4 Hz), 7.04 (t, 2H, J ¼ 8 Hz), 6.78 (d, 1H, J ¼ 8.4 Hz), 5.35 (s, 2H); 13C NMR (100 MHz, CDCl3) d 166.1, 161.9, 154.2, 151.7, 143.3 (d, JCF ¼ 11 Hz), 139.7, 137.3, 132.3, 131.7 (d, JCF ¼ 9 Hz), 129.4, 129.2, 128.7, 127.2, 123.1, 118.4 (d, JCF ¼ 3 Hz), 117.5, 111.8, 111.6, 61.8; MS [m/z þ Na] 482.0. 4.5.8. N-(3-fluoro-4-((5-phenyl-1,3,4-oxadiazol-2-yl)methoxy) phenyl)-4-methylbenzenesulfonamide (1e) Yield 41%; mp 119e121 C; 1H NMR (400 MHz, CDCl3) d 8.05 (d, 2H, J ¼ 8 Hz), 7.73 (s, 1H), 7.63 (d, 2H, J ¼ 8 Hz), 7.55e7.47 (m,3H), 7.20 (d, 2H, J ¼ 12 Hz), 7.04e6.98 (m, 2H), 6.78 (d, 1H, J ¼ 8 Hz), 5.33 (s, 2H), 3.80 (s, 3H); 13C NMR (100 MHz, CDCl3) d 166.0, 161.9, 154.1, 151.7, 144.1, 143.0 (d, JCF ¼ 11 Hz), 135.7, 132.2 (d, JCF ¼ 5 Hz), 129.7, 129.1, 127.2, 123.3, 118.0 (d, JCF ¼ 4 Hz), 117.4, 111.3, 111.1, 61.9, 21.5. 4.5.9. N-(3-fluoro-4-((5-phenyl-1,3,4-oxadiazol-2-yl)methoxy) phenyl)-4-methoxybenzenesulfonamide (1f) Yield 45%; mp 125e127 C; 1H NMR (400 MHz, CDCl3) d 8.05 (d, 2H, J ¼ 8 Hz), 7.68 (d, 2H, J ¼ 5.6 Hz), 7. 55e7.48 (m, 4H), 7.03e6.98
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(m, 2H), 6.87 (d, 2H, J ¼ 9.2 Hz), 6.78 (d, 1H, J ¼ 8.2 Hz), 5.32 (s, 2H), 2.34 (s, 3H); 13C NMR (100 MHz, CDCl3) d 166.0, 163.2, 161.9, 154.2, 151.7, 143.1 (d, JCF ¼ 11 Hz), 132.2 (d, JCF ¼ 8 Hz), 130.2, 129.4, 129.1, 127.2, 123.3, 118.1 (d, JCF ¼ 3 Hz), 117.5, 114.3, 111.4, 111.2, 61.9, 55.6; MS [m/z þ Na] 478.0. Compounds 1gek were prepared with the same method from 6b and the appropriate sulfonyl chlorides. 4.5.10. N-(4-((5-(4-Chlorophenyl)-1,3,4eoxadiazol-2-yl)methoxy)3-fluorophenyl)methanesulfonamide (1g) Yield 30%; mp 130e132 C; 1H NMR (400 MHz, CD3OD) d 8.05 (d, 2H, J ¼ 6.8 Hz), 7.66e7.58 (m, 3H), 7.29 (t, 1H, J ¼ 8.4 Hz), 7.13 (dd, 1H, J ¼ 2.4, J ¼ 12.4 Hz), 7.0e7.03 (m, 1H), 5.47 (s, 2H), 2.97 (s, 3H); MS [m/z þ Na] 419.9. 4.5.11. N-(4-((5-(4-Chlorophenyl)-1,3,4eoxadiazol-2-yl)methoxy)3-fluorophenyl)benzenesulfonamide (1h) Yield 27%; mp 170e172 C; 1H NMR (400 MHz, CD3OD) d 8.05 (d, 2H, J ¼ 8 Hz), 7.74 (d, 2H, J ¼ 6.8 Hz), 7.63e7.58 (m, 3H), 7.50 (d, 2H, J ¼ 8 Hz), 7.15 (t, 2H, J ¼ 8.8 Hz), 6.97 (dd, 1H, J ¼ 2.4, 2.4 Hz), 6.84 (d, 1H, J ¼ 7.6 Hz), 5.40 (s, 2H); MS [m/z þ Na] 482.0. 4.5.12. 4-Chloro-N-(4-((5-(4-chlorophenyl)-1,3,4eoxadiazol-2-yl) methoxy)-3-fluorophenyl)benzenesulfonamide (1i) Yield 28%; mp 173e175 C; 1H NMR (400 MHz, CD3OD) d 7.93 (d, 2H, J ¼ 8.4 Hz), 7.58 (d, 2H, J ¼ 8.4 Hz), 7.50 (d, 2H, J ¼ 8.4 Hz), 7.50 (d, 2H, J ¼ 8.4 Hz), 7.05 (t, 2H, J ¼ 8.8 Hz), 6.75 (dd, 1H, J ¼ 2.4, 2.4 Hz), 6.69e6.71 (m, 1H), 5.29 (s, 2H); MS [m/z þ Na] 516.0. 4.5.13. N-(4-((5-(4-Chlorophenyl)-1,3,4eoxadiazol-2-yl)methoxy)3-fluorophenyl)-4-methylbenzenesulfonamide (1j) Yield 43%; mp 152e154 C; 1H NMR (400 MHz, CDCl3) d 8.00 (d, 2H, J ¼ 8 Hz), 7.64 (d, 2H, J ¼ 8 Hz), 7.49 (d, 2H, J ¼ 8 Hz), 7.21 (d, 2H, J ¼ 8 Hz), 7.03e6.98 (m, 2H), 6.80e6.78 (m, 1H), 5.32 (s, 2H), 2.36 (s, 3H); 13C NMR (100 MHz, CDCl3) d 165.2, 162.0, 154.1, 151.7, 144.1, 143.0 (d, JCF ¼ 11 Hz), 138.8, 132.2 (d, JCF ¼ 9 Hz), 129.8, 129.6, 128.4, 127.2, 121.7, 117.9 (d, JCF ¼ 3 Hz), 117.5, 111.3, 111.1, 61.8, 21.5; MS [m/ z þ Na] 495.9. 4.5.14. N-(4-((5-(4-Chlorophenyl)-1,3,4eoxadiazol-2-yl)methoxy)3-fluorophenyl)-4-methoxybenzenesulfonamide (1k) Yield 40%; mp 146e148 C; 1H NMR (400 MHz, CDCl3) d 8.01 (d, 2H, J ¼ 8 Hz), 7.68 (d, 2H, J ¼ 8 Hz), 7.51 (d, 2H, J ¼ 8.6 Hz), 7.04e6.97 (m, 2H), 6.89 (d, 2H, J ¼ 8 Hz), 6.76 (d, 2H, J ¼ 8 Hz), 5.32 (s, 2H), 3.83 (s, 3H); 13C NMR (100 MHz, CDCl3) d 163.5, 161.6, 160.3, 152.5, 150.0, 141.3 (d, JCF ¼ 11 Hz), 136.9, 130.7 (d, JCF ¼ 9 Hz), 128.5, 127.8, 126.7, 119.9, 116.3 (d, JCF ¼ 3 Hz), 115.8, 112.6, 109.6, 60.1, 53.9; MS [m/ z þ Na] 508.0. Compounds 1leq were prepared with the same method from 6c and the appropriate sulfonyl chlorides. 4.5.15. N-(3-fluoro-4-((5-(4-methoxyphenyl)-1,3,4eoxadiazol-2yl)methoxy)phenyl)methanesulfonamide (1l) Yield 36%; mp 122e124 C; 1H NMR (400 MHz, CDCl3) d 8.01 (d, 2H, J ¼ 8 Hz), 7.44 (s, 1H), 7.15 (t, 1H, J ¼ 8 Hz), 7.02e6.96 (m, 3H), 5.36 (s, 2H), 3.88 (s, 3H), 2.99 (s, 3H); 13C NMR (100 MHz, CDCl3) d 166.0, 162.8, 161.3, 154.5, 151.9, 143.1 (d, JCF ¼ 10 Hz), 132.4 (d, JCF ¼ 9 Hz), 129.0, 117.9, 117.4 (d, JCF ¼ 4 Hz), 114.6, 110.9, 110.7, 61.9, 55.5, 39.3. 4.5.16. N-(3-fluoro-4-((5-(4-methoxyphenyl)-1,3,4eoxadiazol-2yl)methoxy)phenyl)propane-1-sulfonamide (1m) Yield 38%; mp 132e134 C; 1H NMR (400 MHz, CDCl3) d 8.10 (d, 2H, J ¼ 8.2 Hz), 7.65 (s, 1H), 7.08e7.22 (m, 2H), 7.07e6.98 (m, 3H), 5.38 (s, 2H), 3.92 (s, 3H), 3.15 (t, 2H, J ¼ 8.2 Hz), 2.00e1.82 (m, 2H),
51
1.07 (t, 2H, J ¼ 8.2 Hz); 13C NMR (100 MHz, CDCl3) d 166.0, 162.7, 161.4, 154.5, 151.9, 142.8 (d, JCF ¼ 11 Hz), 132.7 (d, JCF ¼ 9 Hz), 128.9, 117.9, 116.8 (d, JCF ¼ 3 Hz), 114.6, 110.4, 110.2, 61.9, 55.5, 53.4, 17.2, 12.8. 4.5.17. N-(3-fluoro-4-((5-(4-methoxyphenyl)-1,3,4eoxadiazol-2yl)methoxy)phenyl)benzenesulfonamide (1n) Yield 30%; mp 138e140 C; 1H NMR (400 MHz, CDCl3) d 8.01 (d, 2H, J ¼ 8.2 Hz), 7.75 (d, 2H, J ¼ 8.2 Hz), 7.50 (s, 1H), 7.48e7.35 (m, 2H), 6.92e7.09 (m, 4H), 5.38 (s, 2H), 7.81e7.71 (m, 1H), 5.32 (s, 2H), 3.89 (s, 3H); MS [m/z þ Na] 478.2. 4.5.18. 4-Chloro-N-(3-fluoro-4-((5-(4-methoxyphenyl)1,3,4eoxadiazol-2-yl)methoxy)phenyl)benzenesulfonamide (1o) Yield 32%; mp 164e166 C; 1H NMR (400 MHz, CDCl3) d 8.01 (d, 2H, J ¼ 7.6 Hz), 7.69 (d, 2H, J ¼ 8.4 Hz), 7.39 (d, 2H, J ¼ 8.4 Hz), 7.06e7.02 (m, 4H), 6.81 (d, 1H, J ¼ 7.6 Hz), 5.35 (s, 2H), 3.89 (s, 3H); 13 C NMR (100 MHz, CDCl3) d 166.1, 162.8, 161.2, 154.3, 150.6, 144.0, 139.8 (d, JCF ¼ 11 Hz), 137.2, 131.3 (d, JCF ¼ 9 Hz), 129.4, 129.0, 128.6, 127.2, 118.5 (d, JCF ¼ 4 Hz), 117.4, 115.6, 114.6, 112.0, 111.7, 61.7, 55.5; MS [m/z þ Na] 512.0. 4.5.19. N-(3-fluoro-4-((5-(4-methoxyphenyl)-1,3,4eoxadiazol-2yl)methoxy)phenyl)-4-methylbenzenesulfonamide (1p) Yield 42%; mp 136e138 C; 1H NMR (400 MHz, CDCl3) d 8.99 (d, 2H, J ¼ 8.8 Hz), 7.63 (d, 2H, J ¼ 10.8 Hz), 7.20 (d, 2H, J ¼ 7.6 Hz), 7.03e6.98 (m, 4H), 6.77 (d, 1H, J ¼ 9.6 Hz), 5.32 (s, 2H), 3.87 (s, 3H), 2.35 (s, 3H); 13C NMR (100 MHz, CDCl3) d 165.9, 162.7, 161.4, 154.1, 151.6, 144.0, 143.1 (d, JCF ¼ 11 Hz), 135.8, 132.1 (d, JCF ¼ 9 Hz), 129.7, 128.9, 127.2, 118.1 (d, JCF ¼ 4 Hz), 117.4, 115.7, 114.6, 111.4, 111.2, 61.8, 55.5, 21.5; MS [m/z þ Na] 492.0. 4.5.20. N-(3-fluoro-4-((5-(4-methoxyphenyl)-1,3,4eoxadiazol-2yl)methoxy)phenyl)-4-methoxybenzenesulfonamide (1q) Yield 43%; mp 124e126 C; 1H NMR (300 MHz, CDCl3) d 8.99 (d, 2H, J ¼ 8.8 Hz), 7.68 (d, 2H, J ¼ 12 Hz), 7.47 (s, 1H), 7.10e6.95 (m, 4H), 6.87 (d, 2H, d ¼ 12 Hz), 5.31 (s, 2H), 3.87 (s, 3H), 3.81 (s, 3H); 13C NMR (100 MHz, CDCl3) d 165.9, 163.2, 162.7, 154.1, 151.7, 143.1 (d, JCF ¼ 11 Hz), 132.2 (d, JCF ¼ 9 Hz), 130.3, 129.4, 128.9, 118.0 (d, JCF ¼ 3 Hz), 117.4, 115.7, 114.6, 114.3, 111.4, 111.3, 61.9, 55.5, 55.4. 4.6. Cancer cell line screening at the NCI Screening against the cancer cell lines was carried out at the National Cancer Institute (NCI), Bethesda, Maryland, USA, applying the standard protocol of the NCI [27]. The human cell lines are grown in RPMI 1640 medium containing 5% fetal bovine serum and 2 mM L-glutamine. For a typical screening experiment, cells are inoculated into 96-well microtiter plates in 100 mL at plating densities ranging from 5000 to 40,000 cells/well depending on the doubling time of individual cell lines. After cell inoculation, the microtiter plates are incubated at 37 C, 5% CO2, 95% air and 100% relative humidity for 24 h prior to addition of experimental drugs. After 24 h, two plates of each cell line are fixed in situ with TCA, to represent a measurement of the cell population for each cell line at the time of drug addition (Tz). Experimental drugs are solubilized in dimethyl sulfoxide at 400-fold the desired final maximum test concentration and stored frozen prior to use. At the time of drug addition, an aliquot of frozen concentrate is thawed and diluted to twice the desired final maximum test concentration with complete medium containing 50 mg/mL gentamicin. Additional four, 10-fold or 1/2 log serial dilutions are made to provide a total of five drug concentrations plus control. Aliquots of 100 mL of these different drug dilutions are added to the appropriate microtiter wells already containing 100 mL of medium, resulting in the required final drug
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concentrations. Following drug addition, the plates are incubated for an additional 48 h at 37 C, 5% CO2, 95% air, and 100% relative humidity. For adherent cells, the assay is terminated by the addition of cold TCA. Cells are fixed in situ by the gentle addition of 50 mL of cold 50% (w/v) TCA (final concentration, 10% TCA) and incubated for 60 min at 4 C. The supernatant is discarded, and the plates are washed five times with tap water and air dried. Sulforhodamine B (SRB) solution (100 mL) at 0.4% (w/v) in 1% acetic acid is added to each well, and plates are kept for 10 min at room temperature. After staining, unbound dye is removed by washing five times with 1% acetic acid and the plates are air dried. Bound stain is subsequently solubilized with 10 mM trizma base, and the absorbance is read on an automated plate reader at a wavelength of 515 nm. For suspension cells, the methodology is the same except that the assay is terminated by fixing settled cells at the bottom of the wells by gently adding 50 mL of 80% TCA (final concentration, 16% TCA). Using the seven absorbance measurements [time zero, (Tz), control growth, (C), and test growth in the presence of drug at the five concentration levels (Ti)], the percentage growth is calculated at each of the drug concentrations levels. Percentage growth inhibition is calculated as: [(Ti Tz)/(C Tz)] 100 for concentrations for which Ti Tz [(Ti Tz)/Tz] 100 for concentrations for which Ti < Tz. Acknowledgments This work was supported by Korea Institute of Science and Technology (KIST), KIST Project (2E23720). We would like to thank the National Cancer Institute (NCI), Bethesda, Maryland, USA, for performing the in vitro anticancer testing over the cell lines. Appendix A. Supplementary data Supplementary data related to this article can be found at http:// dx.doi.org/10.1016/j.ejmech.2014.11.011. References [1] H. Frankish, Lancet 361 (2003) 1278.
[2] D. Belpomme, P. Irigaray, A.J. Sasco, J.A. Newby, V. Howard, R. Clapp, L. Hardell, Int. J. Oncol. 30 (2007) 1037e1049. [3] R. Gudipati, R.N.R. Anreddy, S. Manda, Saudi Pharm. J. 19 (2011) 153e158. [4] M. Rashid, A. Husain, R. Mishra, Eur. J. Med. Chem. 54 (2012) 855e866. [5] A. Husain, M. Rashid, R. Mishra, S. Parveen, D.-S. Shin, D. Kumar, Bioorg. Med. Chem. Lett. 22 (2012) 5438e5444. [6] P. Puthiyapurayil, B. Poojary, C. Chikkanna, S.K. Buridipad, Eur. J. Med. Chem. 53 (2012) 203e210. [7] J. Sun, H. Zhu, Z.-M. Yang, H.-L. Zhu, Eur. J. Med. Chem. 60 (2013) 23e28. [8] S. Zhang, Y. Luo, L.-Q. He, Z.-J. Liu, A.-Q. Jiang, Y.-H. Yang, H.-L. Zhu, Bioorg. Med. Chem. 21 (2013) 3723e3729. [9] N.B. Patel, A.C. Purohit, D.P. Rajani, R. Moo-Puc, G. Rivera, Eur. J. Med. Chem. 62 (2013) 677e687. [10] S. Rapolu, M. Alla, V.R. Bommena, R. Murthy, N. Jain, V.R. Bommareddy, M.R. Bommineni, Eur. J. Med. Chem. 66 (2013) 91e100. [11] P. Li, L. Shi, X. Yang, L. Yang, X.-W. Chen, F. Wu, Q.-C. Shi, W.-M. Xu, M. He, D.Y. Hu, B.-A. Song, Bioorg. Med. Chem. Lett. 24 (2014) 1677e1680. [12] Z. Hadady, M. Toth, L. Somsak, ARKIVOC 7 (2004) 140e149. [13] M.J. Ahsan, J.G. Samy, H. Khalilullah, M.S. Nomani, P. Saraswat, R. Gaur, A. Singh, Bioorg. Med. Chem. Lett. 21 (2011) 7246e7250. [14] F. Liu, X.Q. Luo, B.A. Song, P.S. Bhadury, S. Yang, L.H. Jin, W. Xue, D.Y. Hu, Bioorg. Med. Chem. 16 (2008) 3632e3640. [15] V. Jakubkiene, M.M. Burbuliene, E. Udrenaite, V. Garaliene, P. Vainilavicius, Pharmazie 57 (2002) 610e613. [16] N.C. Desai, N. Bhatt, H. Somani, A. Trivedi, Eur. J. Med. Chem. 67 (2013) 54e59. [17] P.N. Ibrahim, D.R. Artis, R. Bremer, G. Habets, S. Mamo, M. Nespi, C. Zhang, J. Zhang, Y.-L. Zhu, R. Zuckerman, B. West, Y. Suzuki, J. Tsai, K.-P. Hirth, G. Bollag, W. Spevak, H. Cho, S.J. Gillette, G. Wu, H. Zhu, S. Shi, WO2007002433, Jan 4th, 2007. [18] M.M. Ghorab, M.A. Shaaban, H.M. Refaat, H.I. Heiba, S.S. Ibrahim, Eur. J. Med. Chem. 53 (2012) 403e407. [19] J. Lal, S.K. Gupta, D. Thavaselvam, D.D. Agarwal, Eur. J. Med. Chem. 64 (2013) 579e588. [20] Z.-C. Wang, Y.-J. Qin, P.-F. Wang, Y.-A. Yang, Q. Wen, X. Zhang, H.-Y. Qiu, Y.T. Duan, Y.-T. Wang, Y.-L. Sang, H.-L. Zhu, Eur. J. Med. Chem. 66 (2013) 1e11. [21] X.-M. Wang, J. Xu, Y.-P. Li, H. Li, C.-S. Jiang, G.-D. Yang, S.-M. Lu, S.-Q. Zhang, Eur. J. Med. Chem. 67 (2013) 243e251. [22] F. Xu, H. Xu, X. Wang, L. Zhang, Q. Wen, Y. Zhang, W. Xu, Bioorg. Med. Chem. 22 (2014) 1487e1495. da, A.L.P. de Miranda, [23] P.C. Lima, L.M. Lima, K.C.M. da Silva, P.H.O. Le C.A.M. Fraga, E.J. Barreiro, Eur. J. Med. Chem. 35 (2000) 187e203. [24] S. Cao, X.H. Quin, G.H. Song, B. Chia, Z.S. Jiang, J. Agric. Food Chem. 51 (2003) 152e155. [25] NCI website: www.dtp.nci.nih.gov. (retrieved on Oct. 31st, 2014). [26] DTP Data Search: http://dtp.nci.nih.gov/dtpstandard/dwindex/index.jsp. (retrieved on Oct. 31st, 2014). [27] DTP Human Tumor Cell Line Screen Process: http://www.dtp.nci.nih.gov/ branches/btb/ivclsp.html. (retrieved on Oct. 31st, 2014).
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry journal homepage: http://www.elsevier.com/locate/ejmech
Short communication
Synthesis and in vitro antiproliferative activity of new 1,3,4-oxadiazole derivatives possessing sulfonamide moiety Mahmoud M. Gamal El-Din a, b, c, Mohammed I. El-Gamal d, Mohammed S. Abdel-Maksoud a, b, c, Kyung Ho Yoo e, Chang-Hyun Oh a, b, * a
Center for Biomaterials, Korea Institute of Science and Technology, PO Box 131, Cheongryang, Seoul 130-650, Republic of Korea Department of Biomolecular Science, University of Science and Technology, 113 Gwahangno, Yuseong-gu, Daejeon 305-333, Republic of Korea Pharmaceutical and Drug Industries Research Division, National Research Centre, Dokki, Giza 12622, Egypt d Department of Medicinal Chemistry, Faculty of Pharmacy, University of Mansoura, Mansoura 35516, Egypt e Chemical Kinomics Research Center, Korea Institute of Science and Technology, PO Box 131, Cheongryang, Seoul 130-650, Republic of Korea b c
a r t i c l e i n f o
a b s t r a c t
Article history: Received 10 June 2014 Received in revised form 3 November 2014 Accepted 5 November 2014 Available online 6 November 2014
Synthesis of a new series of 1,3,4-oxadiazole derivatives possessing sulfonamide moiety is described. Their in vitro antiproliferative activities against NCI-58 human cancer cell lines of nine different cancer types were tested. Compound 1k with p-methoxybenzenesulfonamido moiety showed the highest mean %inhibition value over the 58 cell line panel at 10 mM concentration. It showed broad-spectrum antiproliferative activity over many cell lines of different cancer types. For instance, compound 1k inhibited the growth of T-47D breast cancer cell line by 90.47% at 10 mM. And it inhibited growth of SR leukemia, SK-MEL-5 melanoma, and MDA-MB-468 breast cancer cell lines by more than 80% at the same test concentration. Compound 1k showed superior activity than Paclitaxel and Gefitinib against the most sensitive cell lines. © 2014 Elsevier Masson SAS. All rights reserved.
Keywords: Antiproliferative activity 1,3,4-Oxadiazole Sulfonamide
1. Introduction Cancer is a collection of different life-threatening diseases characterized by uncontrolled growth of cells, leading to invasion of surrounding tissue and often metastasizing to other parts of the body. According to the statistics, cancer is the second most common cause of death worldwide after cardiovascular diseases [http:// www.cdc.gov/nchs/fastats/lcod.htm]. Despite the availability of improved drugs including targeted cancer therapies, the worldwide cancer burden is expected to increase by as much as 15 million new cancer cases per year by 2020, according to the World Health Organization (WHO), unless further preventive measures are put into practice [1,2]. The development of new anticancer drugs represents a major interest and challenge to the contemporary medicinal chemistry. Much attention has been paid to the chemistry and biological activities of 1,3,4-oxadiazole nucleus. Several compounds possessing 1,3,4-oxadiazole scaffold have been recently reported as potential antiproliferative agents [3e11]. Apart from anticancer
* Corresponding author. Center for Biomaterials, Korea Institute of Science and Technology, PO Box 131, Cheongryang, Seoul 130-650, Republic of Korea. E-mail address: [email protected] (C.-H. Oh). http://dx.doi.org/10.1016/j.ejmech.2014.11.011 0223-5234/© 2014 Elsevier Masson SAS. All rights reserved.
activity, other biological activities have been reported for 1,3,4oxadiazole derivatives such as antidiabetic [12], antitubercular [13], antifungal [14], antiinflammatory [15], and antibacterial activities [16]. Furthermore, many sulfonamide derivatives have been highlighted as anticancer agents [17e22]. In the present study, we report synthesis of a new series of 1,3,4oxadiazole derivatives possessing terminal sulfonamide moiety. Their in vitro antiproliferative activities against NCI-58 cancer cell line panel are reported.
2. Results and discussion 2.1. Chemistry Synthesis of the target compounds 1aeq was achieved through the pathway illustrated in Scheme 1. Refluxing the benzoate esters 2aec with hydrazine monohydrate in ethanol afforded the corresponding benzohydrazide derivatives 3aec [23]. Cyclization to 2(chloromethyl)-5-aryl-1,3,4-oxadiazole analogs 4aec was carried out through refluxing the hydrazides 3aec with chloroacetic acid in phosphorus oxychloride [24]. Nucleophilic substitution of chloro group of compounds 4aec with 2-fluoro-4-nitrophenol was done by reflux in acetonitrile in the presence of potassium carbonate to
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Scheme 1. Reagents and conditions: a) hydrazine monohydrate, EtOH, reflux, overnight; b) ClCH2COOH, POCl3, reflux, 6 h; c) 2-fluoro-4-nitrophenol, K2CO3, acetonitrile, reflux, 36 h; d) Raney Ni, H2, THF, rt, 24 h; e) alkyl or aryl sulfonyl chloride, C5H5N, CH2Cl2, rt, overnight.
obtain the ether derivatives 5aec. The nitro group of compounds 5aec was reduced to amino using Raney Nickel in hydrogen atmosphere. Treatment of the aniline derivatives 6aec with the appropriate alkyl or aryl sulfonyl chloride derivatives in presence of pyridine as a base afforded the target sulfonamide derivatives 1aeq. 2.2. In vitro antiproliferative activity Structures of the synthesized target compounds were submitted to National Cancer Institute (NCI), Bethesda, Maryland, USA [25], and the thirteen compounds shown in Table 2 were selected on the basis of degree of structural variation and computer modeling techniques for evaluation of their antineoplastic activity. The selected compounds were subjected to in vitro anticancer assay against tumor cells in a full panel of 58 cell lines taken from nine different tissues (blood, lung, colon, CNS, skin, ovary, kidney, prostate, and breast). The compounds were tested at a single-dose concentration of 10 mM, and the percentages of growth inhibition over the 58 tested cell lines were determined. The mean % growth of the NCI-58 cancer cell line panel after treatment with each of the tested compounds is illustrated in Table 1. Upon comparing the effect of aryl ring directly attached to 1,3,4oxadiazole ring on activity, it was found that compounds 1m and 1o possessing p-methoxyphenyl were more active than compounds 1b and 1d with phenyl ring. Compounds 1h and 1k containing pchlorophenyl showed higher antiproliferative activity than the corresponding p-methoxyphenyl analogs 1n and 1q. Compound 1g with p-chlorophenyl was more active that the corresponding methoxy derivative 1l, and 1l exerted higher activity than 1a with unsubstituted phenyl ring. So it can be concluded that p-chlorophenyl is the most optimum and unsubstituted phenyl is the least optimum for activity of this series of compounds. This can be attributed to steric and/or electronic differences between chloro, methoxy, and hydrogen groups. The effect of alkyl(aryl)sulfonamide moiety on activity was also investigated. Among aliphatic derivatives, n-propylsulfonamido compounds 1b and 1m were more active than the corresponding methylsulfonamido analogs 1a and 1l. So homologation to a longer chain alkyl group could be favorable for activity, may be due to
stronger hydrophobic interaction at the receptor site and/or the increased lipophilicity enhanced penetration into the cancer cells. The influences of aliphatic and aromatic sulfonamido moieties on activity were compared. The aromatic derivatives were found generally more active than aliphatic analogs (compound 1d more active than 1a and 1b), (compounds 1h, 1j, and 1k more active than 1g), and (compounds 1oeq more active than 1l and 1m). Furthermore, substituted phenyl moieties were more optimal for activity than unsubstituted phenyl (compounds 1j and 1k were more active than 1h, and 1oeq showed higher activity than 1n). The pmethoxybenzenesulfonamido compounds 1k and 1q exerted superior activity compared to the corresponding p-toluenesulfonamido derivatives 1j and 1p. This may be rationalized that methoxy Table 1 Mean %growth of the 58 cancer cell line panel after treatment with the tested target compounds (10 mM concentration).
Compound no.
R1
R2
Mean %growth
1a 1b 1d 1g 1h 1j 1k 1l 1m 1n 1o 1p 1q
H H H Cl Cl Cl Cl OMe OMe OMe OMe OMe OMe
CH3 n-Pr 4-Cl(C6H4) CH3 C6H5 4-Tolyl 4-MeO(C6H4) CH3 n-Pr C6H5 4-Cl(C6H4) 4-Tolyl 4-MeO(C6H4)
100.73 98.89 97.14 93.74 91.95 84.99 57.53 97.56 91.00 97.55 73.29 83.27 72.40
The bold figure indicates the most active compound.
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Table 2 %Inhibition results exerted by the most potential compounds over the most sensitive cell lines.
Compound no.
R1
R2
Cell line
Cancer type
% Inhibition at 10 mM
1k
4-Cl
4-OMe(C6H4)
1o
4-OMe
4-Cl(C6H4)
1p
4-OMe
4-Tolyl
1q
4-OMe
4-OMe(C6H4)
K-562 MOLT-4 RPMI-8226 SR A549 NCI-H23 NCI-H522 SK-MEL-2 SK-MEL-5 UACC-257 UACC -62 T-47D MDA-MB-468 MCF-7 A-498 NCI-H522 T-47D MDA-MB-468 RPMI-8226 T-47D MDA-MB-468 UACC-62 RPMI-8226 CCRF-CEM NCI-H522 SK-MEL 5 T-47D
Leukemia Leukemia Leukemia Leukemia Lung cancer Lung cancer Lung cancer Melanoma Melanoma Melanoma Melanoma Breast cancer Breast cancer Breast cancer Renal cancer Lung cancer Breast cancer Breast cancer leukemia Breast cancer Breast cancer Melanoma Leukemia Leukemia Lung cancer Melanoma Breast cancer
56.82 63.99 68.36 81.58 52.58 56.58 56.57 59.40 84.32 76.66 57.66 90.47 84.83 57.22 53.03 72.66 59.48 54.52 55.35 71.88 40.25 40.96 56.64 56.91 50.84 56.06 68.44
The bold figures indicate the highest growth inhibition percentages.
group on this terminal ring may be essential as hydrogen bond acceptor at the receptor site. This enhances affinity with the receptor, and hence the antiproliferative activity improves. Among all the target derivatives, compound 1k with p-chlorophenyl on the oxadiazole ring and p-methoxybenzenesulfonamido terminal moiety showed the best activity. So those moieties together are the most favorable for antiproliferative activity of this series of compounds.
The %inhibition values of the most active compounds over the most sensitive cell lines are summarized in Table 2. T-47D breast cancer cell line was the most sensitive cell line to this series of compounds. The most active compounds against T-47D were 1k and 1p with %inhibitions of 90.47% and 71.88%, respectively. MDAMB-468 breast cancer cell line was also sensitive to the most potential compounds. Fig. 1 illustrates the %inhibition values of the tested target compounds against T-47D and MDA-MB-468 cell lines. Compounds 1k and 1o were active against both cell lines,
Fig. 1. Inhibitory effects of the tested target compounds at 10 mM concentration over T-47D and MDA-MB-468 breast cancer cell lines.
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Fig. 2. % Inhibition expressed by compounds 1k (Fig. 2a), 1o (Fig. 2b), and 1q (Fig. 2c) at a single-dose concentration of 10 mM over all cell lines of the NCI cancer cell line panel of nine different cancer types.
while compounds 1p and 1q showed higher inhibitory effect over T-47D. Among all the target compounds, compound 1k showed the most promising results. It exerted broad-spectrum antiproliferative activity against different cell lines of different cancer types; leukemia, melanoma, and breast cancer. So it can be considered as a
potential lead compound for future development of broadspectrum anticancer agents. Among all the tested derivatives, compounds 1k, 1o, and 1q showed the highest mean inhibitions. The percentages of inhibition of these four compounds over each tested cell line of the NCI-58 panel at 10 mM concentration are depicted in Fig. 2. The three
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49
tetramethylsilane as an internal standard. Melting points were obtained on a Walden Precision Apparatus Electrothermal 9300 apparatus and are uncorrected. Solvents and liquid reagents were transferred using hypodermic syringes. Purity % of all the target compounds were determined by HPLC and found to be >95%. All solvents and reagents were commercially available and used without further purification. 4.2. Synthesis of benzohydrazide derivatives 3aec [23]
Fig. 3. Comparison of %inhibition values expressed by compound 1k, Paclitaxel, and Gefitinib against the most sensitive cell lines towards compound 1k.
compounds exerted broad-spectrum antiproliferative activities against different cell lines of different cancer types. Among them, compound 1k was the most active. It showed the highest %inhibition values with more than 70% inhibition against SR leukemia cell line, SK-MEL-5 and UACC-257 melanoma cell lines, and T-47D and MDA-MB-468 breast cancer cell lines. The results of compound 1k against those five cell lines were compared with Paclitaxel and Gefitinib as reference standard drugs, as illustrated in Fig. 3. The results of Paclitaxel and Gefitinib were obtained from NCI datawarehouse index [26], and are inserted in Fig. 3. Compound 1k was more active than both reference compounds against SK-MEL-5 and T-47D cell lines. It showed higher activity also than Gefitinib against SR and UACC-257 cell lines. 3. Conclusion In this study, a series of new 1,3,4-oxadiazole derivatives possessing terminal sulfonamide moiety was synthesized. The target compounds were tested for in vitro antiproliferative activities over NCI-58 cancer cell line panel of nine different cancer types. Among them, compound 1k possessing p-chlorophenyl ring attached to 1,3,4-oxadiazole ring and p-methoxybenzenesulfonamido terminal moiety showed the most promising results at a single-dose concentration of 10 mM. It exerted broad-spectrum antiproliferative activities over different cell lines of different cancer types. The % inhibition values of compound 1k over T-47D breast, SR leukemia, SK-MEL-5 melanoma, and MDA-MB-468 breast cancer cell lines were 90.47%, 81.58%, 84.32%, and 84.83%, respectively. T-47D and MDA-MB-468 breast cancer cell lines were found to be highly sensitive to the most active compounds. The in vitro antiproliferative activity indicated that compound 1k could be a promising lead for future development of active anticancer agents. It was more active than Paclitaxel and Gefitinib against SK-MEL-5 melanoma and T-47D breast cancer cell lines. It exerted also higher activity than Gefitinib against SR leukemia and UACC-257 melanoma cell lines. Further modifications of this series of 1,3,4oxadiazole compounds in order to improve their antiproliferative activity are currently in progress. 4. Experimental 4.1. General The target compounds were purified by column chromatography using silica gel (0.040e0.063 mm, 230e400 mesh) and technical grade solvents. 1H NMR and 13C NMR spectra were recorded on a Bruker Avance 400 or 300 spectrometer using
To a solution of the appropriate methyl benzoate ester (1.0 mmol) in ethanol (30 mL), hydrazine monohydrate (0.15 g, 3.0 mmol) was added. The reaction mixture was heated under reflux overnight. After completion of the reaction, the solvent was evaporated under reduced pressure, and the residue was washed with water (2 3 mL), and the obtained solid was filtered and dried to give benzohydrazide derivatives 3aec. Benzohydrazide (3a): yield 70%; 1H NMR (400 MHz, DMSO-d6) d 9.80 (s, 1H), 7.84 (d, 2H, J ¼ 8.0 Hz), 7.47e7.45 (m, 3H), 4.52 (s, 2H); 13 C NMR (100 MHz, DMSO-d6) d 165.9, 133.3, 131.0, 128.3, 126.9. 4-Chlorobenzohydrazide (3b): yield 82.4%; 1H NMR (400 MHz, DMSO-d6) d 9.86 (s, 1H), 7.84 (d, 2H, J ¼ 8.8 Hz), 7.53 (d, 2H, J ¼ 8.4 Hz), 4.52 (s, 2H); 13C NMR (100 MHz, DMSO-d6) d 164.7, 135.8, 132.0, 128.8, 128.4. 4-Methoxybenzohydrazide (3c): yield 87%; 1H NMR (400 MHz, DMSO-d6) d 9.65 (s, 1H), 7.83 (d, 2H, J ¼ 8.8 Hz), 6.98 (d, 2H, J ¼ 8.8 Hz), 4.47 (s, 2H), 3.80 (s, 3H); 13C NMR (100 MHz, DMSO-d6) d 170.9, 166.6, 133.9, 130.7, 118.7, 60.5. 4.3. Synthesis of 2-(chloromethyl)-5-substituted phenyl-1,3,4oxadiazole (4aec) To a mixture of compound 3aec (3.0 mmol) and chloroacetic acid (0.26 g, 2.75 mmol), phosphorus oxychloride (1.5 g, 1 mL, 10 mmol) was added and the mixture was refluxed for 5e6 h. The reaction mixture was cooled to rt then with ice bath, and saturated solution of sodium bicarbonate was added dropwise thereto till pH 9. The precipitated product was filtered, washed with plenty of water, and then dried to give the title compounds. 2-(Chloromethyl)-5-phenyl-1,3,4-oxadiazole (4a): yield 63.5%; 1 H NMR (400 MHz, DMSO-d6) d 8.03 (dd, 2H, J ¼ 1.6, J ¼ 8.4 Hz), 7.70e7.61 (m, 3H), 5.16 (s, 2H); 13C NMR (100 MHz, DMSO-d6) d 164.9, 162.8, 132.4, 129.5, 126.7, 122.8, 33.2. 2-(Chloromethyl)-5-(4-chlorophenyl)-1,3,4-oxadiazole (4b): yield 71.3%; 1H NMR (400 MHz, DMSO-d6) d 8.00 (d, 2H, J ¼ 8.8 Hz), 7.66 (d, 2H, J ¼ 8.8 Hz), 5.15 (s, 2H); 13C NMR (100 MHz, DMSO-d6) d 164.2, 162.9, 137.1, 129.6, 128.4, 121.7, 33.2. 2-(Chloromethyl)-5-(4-methoxyphenyl)-1,3,4-oxadiazole (4c): yield 72.5% 1H NMR (400 MHz, DMSO-d6) d 7.95 (d, 2H, J ¼ 8.8 Hz), 7.14 (d, 2H, J ¼ 8.8 Hz), 5.13 (s, 2H); 13C NMR (100 MHz, DMSO-d6) d 164.9, 162.2, 128.5, 128.4, 115.1, 114.7, 55.5, 33.2. 4.4. Synthesis of 2-((2-fluoro-4-nitrophenoxy)methyl)-5substitutedphenyl-1,3,4-oxadiazole (5aec) To a solution of compound 4aec (0.5 mmol) and 2-fluoro-4nitrophenol (0.1 g, 0.6 mmol) in dry acetonitrile (50 mL), anhydrous potassium carbonate (0.2 g, 1.5 mmol) was added and the mixture was refluxed for 24e36 h till completion of the reaction. The solvent was evaporated under reduced pressure till dryness, and the residue was partitioned between water (50 mL) and ethyl acetate (50 mL). The organic layer was separated, and the aqueous layer was extracted with water (3 50 mL). The combined organic extracts were washed with brine and then dried over anhydrous sodium sulfate. The solvent was evaporated under reduced
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pressure to the get the corresponding nitro compound. The product was crystalized from methanol:ethyl acetate 4:1 v/v to give the pure title compounds. 4.4.1. 2-((2-Fluoro-4-nitrophenoxy)methyl)-5-phenyl-1,3,4oxadiazole (5a) yield 61.2%; mp 139e141 C; 1H NMR (400 MHz, DMSO-d6) d 8.27e8.20 (m, 2H), 8.05 (d, 2H, J ¼ 8.4 Hz), 7.71e7.63 (m, 4H), 5.84 (s, 2H); 13C NMR (100 MHz, DMSO-d6) d 164.9, 161.6, 151.7, 150.9 (d, JCF ¼ 10 Hz), 149.2, 141.2 (d, JCF ¼ 8 Hz), 132.3, 129.5, 126.7, 122.9, 121.2 (d, JCF ¼ 4 Hz), 115.1, 112.5, 112.2, 60.9. 4.4.2. 2-(4-Chlorophenyl)-5-((2-fluoro-4-nitrophenoxy)methyl)1,3,4-oxadiazole (5b) yield 47.3%; mp 142e143 C; 1H NMR (400 MHz, DMSO-d6) d 8.24e8.17 (m, 2H), 8.03 (d, 2H, J ¼ 8 Hz), 7.70e7.65 (m, 3H), 5.80 (s, 2H); 13C NMR (100 MHz, DMSO-d6) d 164.2, 161.8, 151.7, 150.8 (d, JCF ¼ 11 Hz), 149.2, 141.2 (d, JCF ¼ 8 Hz), 137.1, 129.7, 128.5, 121.8, 121.2 (d, JCF ¼ 4 Hz), 115.1, 112.5, 112.2, 60.9. 4.4.3. 2-((2-Fluoro-4-nitrophenoxy)methyl)-5-(4-methoxyphenyl)1,3,4-oxadiazole (5c) yield 55.6%; mp 136e138 C; 1H NMR (400 MHz, DMSO-d6) d 8.23e8.16 (m, 2H), 7.95 (d, 2H, J ¼ 8.8 Hz), 7.66 (t, 1H, J ¼ 8.4 Hz), 7.15 (d, 2H, J ¼ 9.2 Hz), 5.77 (s, 2H), 3.86 (s, 3H); 13C NMR (100 MHz, DMSO-d6) d 164.9, 162.3, 161.0, 151.7, 150.9 (d, JCF ¼ 10 Hz), 149.2, 141.2 (d, JCF ¼ 8 Hz), 128.5, 121.2 (d, JCF ¼ 3 Hz), 115.1, 115.0, 114.9, 112.4, 112.2, 60.9, 55.5. 4.5. Synthesis of 3-fluoro-4-(5-substituted phenyl-1, 3, 4-oxadiazol2-yl)methoxy)benzamine (6aec) To a solution of the compound 5aec (0.5 mmol) in a mixture of THF and methanol (2:1 v/v, 150 mL), freshly activated Raney nickel (10% eq) was added. The mixture was stirred in hydrogen atmosphere at rt for 24 h. The catalyst was cautiously filtered, and the solvent was removed under reduced pressure to give the amine in quantitative yield. 4.5.1. 3-Fluoro-4-((5-phenyl-1,3,4-oxadiazol-2-yl)methoxy)aniline (6a) yield 85%; mp 132e133 C; 1H NMR (400 MHz, CD3OD) d 8.08 (d, 2H, J ¼ 6.4 Hz), 7.63e7.59 (m, 3H), 7.00 (t, 1H, J ¼ 8.4 Hz), 6.52e6.45 (m, 2H), 5.30 (s, 2H); 13C NMR (100 MHz, CD3OD) d 167.2.9, 164.7, 157.1, 154.6, 146.5 (d, JCF ¼ 10 Hz), 138.3 (d, JCF ¼ 7 Hz), 133.5, 130.5, 130.4, 128.0, 124.6, 121.5, 111.9 (d, JCF ¼ 4 Hz), 104.5, 64.4. 4.5.2. 4-((5-(4-Chlorophenyl)-1,3,4-oxadiazol-2-yl)methoxy)-3fluoroaniline (6b) yield 89%; mp 143e145 C; 1H NMR (400 MHz, DMSO-d6) d 8.05 (d, 2H, J ¼ 6.4 Hz), 7.46 (t, 1H, J ¼ 8.4 Hz), 6.56 (d, 1H, J ¼ 8.4 Hz), 6.46 (d, 1H, J ¼ 8.8 Hz), 5.27 (s, 2H); 13C NMR (100 MHz, DMSO-d6) d 166.8, 165.7, 162.3, 157.1, 144.6 (d, JCF ¼ 11 Hz), 138.2 (d, JCF ¼ 10 Hz), 121.4, 118.0, 115.5, 111.0 (d, JCF ¼ 4 Hz), 105.0, 104.8, 63.6. 4.5.3. 3-Fluoro-4-((5-(4-methoxyphenyl)-1,3,4-oxadiazol-2-yl) methoxy)aniline (6c) yield 93.2%; mp 92e94 C; 1H NMR (400 MHz, DMSO-d6) d 8.01 (d, 2H, J ¼ 8.4 Hz), 6.94 (d, 2H, J ¼ 8.4 Hz), 6.91 (t, 1H, J ¼ 8.4 Hz), 6.45 (dd, 1H, J ¼ 2.4, J ¼ 12.8 Hz), 6.34 (d, 1H, J ¼ 8.8 Hz), 5.24 (s, 2H), 3.88 (s, 3H), 2.15 (s, 2H); 13C NMR (100 MHz, DMSO-d6) d 165.8, 162.6, 161.9, 155.6, 143.3 (d, JCF ¼ 9 Hz), 137.6 (d, JCF ¼ 12 Hz), 120.2, 116.0, 114.5, 110.5 (d, JCF ¼ 3 Hz), 104.0, 103.8, 63.3, 55.5.
4.5.4. N-(3-fluoro-4-((5-phenyl-1,3,4-oxadiazol-2-yl)methoxy) phenyl)methanesulfonamide (1a) To a solution of the compound 6a (0.18 mmol) in anhydrous THF, catalytic amount of pyridine was added in ice cooled bath and was stirred for 15 min then Methanesulfonyl chloride (0.025 g, 0.22 mmol) was added and the reaction mixture was stirred at room temperature for 24 h. When the reaction was completed, the solvent was completely removed and the residue was partitioned between water and ethyl acetate. The organic layer was separated and the aqueous layer was extracted with ethyl acetate (3 5 mL) and the combined organic extracts was washed with brine, separated, dried over anhydrous sodium sulfate after evaporation of the solvent, the residue was purified by column chromatography (silica gel, hexane-ethyl acetate 3:1 v/v) to give the compound as solid 25 mg 39% yield; mp 163e165 C; 1H NMR (300 MHz, CDCl3) d 8.08 (d, 2H, J ¼ 6.1 Hz), 7.54 (d, 2H, J ¼ 6.1 Hz), 7.17 (t, 2H, J ¼ 9.2 Hz), 6.94 (d, 1H, J ¼ 6.3 Hz), 6.70 (s, 1H), 5.39 (s, 2H), 3.00 (s, 3H); 13C NMR (100 MHz, CD3OD) d 167.3, 164.1, 155.7, 153.3, 144.0 (d, JCF ¼ 12 Hz), 135.0 (d, JCF ¼ 9 Hz), 133.5, 130.5, 128.1, 124.5, 119.2, 118.1 (d, JCF ¼ 3 Hz), 111.1, 63.1, 39.2. Compounds 1beq were prepared with the same method from 1a and the appropriate sulfonyl chloride derivative. 4.5.5. N-(3-fluoro-4-((5-phenyl-1, 3, 4-oxadiazol-2-yl)methoxy) phenyl)propan-1-sulfonamide (1b) Yield 41%; mp 169e171 C; 1H NMR (400 MHz, CDCl3) d 8.08 (d, 2H, J ¼ 8.4 Hz), 7.57e7.50 (m, 3H), 7.17e7.13 (m, 2H), 6.90 (d, 2H, J ¼ 8.4 Hz), 6.46 (s, 1H), 5.38 (s, 2H), 3.06e3.02 (m, 2H), 1.87e1.82 (m,2H), 1.03 (t, 3H, J ¼ 7.2 Hz); 13C NMR (100 MHz, CDCl3) d 166.1, 161.9, 154.5, 152.0, 142.9 (d, JCF ¼ 11 Hz), 132.6 (d, JCF ¼ 9 Hz), 132.3, 129.2, 127.2, 123.3, 118.1, 116.9 (d, JCF ¼ 2 Hz), 110.4, 62.0, 53.4, 17.2, 12.8. 4.5.6. N-(3-fluoro-4-((5-phenyl-1,3,4-oxadiazol-2-yl)methoxy) phenyl)benzenesulfonamide (1c) Yield 34%; mp 137e139 C; 1H NMR (400 MHz, CDCl3) d 8.08 (d, 2H, J ¼ 7.9 Hz), 7.58 (d, 2H, J ¼ 8.1 Hz), 7.63e7.43 (m, 5H), 7.40 (t, 2H, J ¼ 4.3 Hz), 7.02 (t, 2H, J ¼ 4.4 Hz), 5.33 (s, 2H); 13C NMR (100 MHz, CDCl3) d 166.0, 161.9, 154.2, 151.7, 143.2 (d, JCF ¼ 11 Hz), 138.7, 133.1, 132.3, 132.0 (d, JCF ¼ 9 Hz), 129.1, 127.1, 123.2, 118.2 (d, JCF ¼ 5 Hz), 117.5, 111.6, 111.4, 61.9. 4.5.7. 4-Chloro-N-(3-fluoro-4-((5-phenyl-1,3,4-oxadiazol-2-yl) methoxy)phenyl)benzenesulfonamide (1d) Yield 43%; mp 115e117 C; 1H NMR (400 MHz, CDCl3) d 8.05 (d, 2H, J ¼ 8 Hz), 7.83 (s, 1H), 7.67 (d, 2H, J ¼ 8.8 Hz), 7.58e7.49 (m,3H), 7.37 (d, 2H, J ¼ 8.4 Hz), 7.04 (t, 2H, J ¼ 8 Hz), 6.78 (d, 1H, J ¼ 8.4 Hz), 5.35 (s, 2H); 13C NMR (100 MHz, CDCl3) d 166.1, 161.9, 154.2, 151.7, 143.3 (d, JCF ¼ 11 Hz), 139.7, 137.3, 132.3, 131.7 (d, JCF ¼ 9 Hz), 129.4, 129.2, 128.7, 127.2, 123.1, 118.4 (d, JCF ¼ 3 Hz), 117.5, 111.8, 111.6, 61.8; MS [m/z þ Na] 482.0. 4.5.8. N-(3-fluoro-4-((5-phenyl-1,3,4-oxadiazol-2-yl)methoxy) phenyl)-4-methylbenzenesulfonamide (1e) Yield 41%; mp 119e121 C; 1H NMR (400 MHz, CDCl3) d 8.05 (d, 2H, J ¼ 8 Hz), 7.73 (s, 1H), 7.63 (d, 2H, J ¼ 8 Hz), 7.55e7.47 (m,3H), 7.20 (d, 2H, J ¼ 12 Hz), 7.04e6.98 (m, 2H), 6.78 (d, 1H, J ¼ 8 Hz), 5.33 (s, 2H), 3.80 (s, 3H); 13C NMR (100 MHz, CDCl3) d 166.0, 161.9, 154.1, 151.7, 144.1, 143.0 (d, JCF ¼ 11 Hz), 135.7, 132.2 (d, JCF ¼ 5 Hz), 129.7, 129.1, 127.2, 123.3, 118.0 (d, JCF ¼ 4 Hz), 117.4, 111.3, 111.1, 61.9, 21.5. 4.5.9. N-(3-fluoro-4-((5-phenyl-1,3,4-oxadiazol-2-yl)methoxy) phenyl)-4-methoxybenzenesulfonamide (1f) Yield 45%; mp 125e127 C; 1H NMR (400 MHz, CDCl3) d 8.05 (d, 2H, J ¼ 8 Hz), 7.68 (d, 2H, J ¼ 5.6 Hz), 7. 55e7.48 (m, 4H), 7.03e6.98
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(m, 2H), 6.87 (d, 2H, J ¼ 9.2 Hz), 6.78 (d, 1H, J ¼ 8.2 Hz), 5.32 (s, 2H), 2.34 (s, 3H); 13C NMR (100 MHz, CDCl3) d 166.0, 163.2, 161.9, 154.2, 151.7, 143.1 (d, JCF ¼ 11 Hz), 132.2 (d, JCF ¼ 8 Hz), 130.2, 129.4, 129.1, 127.2, 123.3, 118.1 (d, JCF ¼ 3 Hz), 117.5, 114.3, 111.4, 111.2, 61.9, 55.6; MS [m/z þ Na] 478.0. Compounds 1gek were prepared with the same method from 6b and the appropriate sulfonyl chlorides. 4.5.10. N-(4-((5-(4-Chlorophenyl)-1,3,4eoxadiazol-2-yl)methoxy)3-fluorophenyl)methanesulfonamide (1g) Yield 30%; mp 130e132 C; 1H NMR (400 MHz, CD3OD) d 8.05 (d, 2H, J ¼ 6.8 Hz), 7.66e7.58 (m, 3H), 7.29 (t, 1H, J ¼ 8.4 Hz), 7.13 (dd, 1H, J ¼ 2.4, J ¼ 12.4 Hz), 7.0e7.03 (m, 1H), 5.47 (s, 2H), 2.97 (s, 3H); MS [m/z þ Na] 419.9. 4.5.11. N-(4-((5-(4-Chlorophenyl)-1,3,4eoxadiazol-2-yl)methoxy)3-fluorophenyl)benzenesulfonamide (1h) Yield 27%; mp 170e172 C; 1H NMR (400 MHz, CD3OD) d 8.05 (d, 2H, J ¼ 8 Hz), 7.74 (d, 2H, J ¼ 6.8 Hz), 7.63e7.58 (m, 3H), 7.50 (d, 2H, J ¼ 8 Hz), 7.15 (t, 2H, J ¼ 8.8 Hz), 6.97 (dd, 1H, J ¼ 2.4, 2.4 Hz), 6.84 (d, 1H, J ¼ 7.6 Hz), 5.40 (s, 2H); MS [m/z þ Na] 482.0. 4.5.12. 4-Chloro-N-(4-((5-(4-chlorophenyl)-1,3,4eoxadiazol-2-yl) methoxy)-3-fluorophenyl)benzenesulfonamide (1i) Yield 28%; mp 173e175 C; 1H NMR (400 MHz, CD3OD) d 7.93 (d, 2H, J ¼ 8.4 Hz), 7.58 (d, 2H, J ¼ 8.4 Hz), 7.50 (d, 2H, J ¼ 8.4 Hz), 7.50 (d, 2H, J ¼ 8.4 Hz), 7.05 (t, 2H, J ¼ 8.8 Hz), 6.75 (dd, 1H, J ¼ 2.4, 2.4 Hz), 6.69e6.71 (m, 1H), 5.29 (s, 2H); MS [m/z þ Na] 516.0. 4.5.13. N-(4-((5-(4-Chlorophenyl)-1,3,4eoxadiazol-2-yl)methoxy)3-fluorophenyl)-4-methylbenzenesulfonamide (1j) Yield 43%; mp 152e154 C; 1H NMR (400 MHz, CDCl3) d 8.00 (d, 2H, J ¼ 8 Hz), 7.64 (d, 2H, J ¼ 8 Hz), 7.49 (d, 2H, J ¼ 8 Hz), 7.21 (d, 2H, J ¼ 8 Hz), 7.03e6.98 (m, 2H), 6.80e6.78 (m, 1H), 5.32 (s, 2H), 2.36 (s, 3H); 13C NMR (100 MHz, CDCl3) d 165.2, 162.0, 154.1, 151.7, 144.1, 143.0 (d, JCF ¼ 11 Hz), 138.8, 132.2 (d, JCF ¼ 9 Hz), 129.8, 129.6, 128.4, 127.2, 121.7, 117.9 (d, JCF ¼ 3 Hz), 117.5, 111.3, 111.1, 61.8, 21.5; MS [m/ z þ Na] 495.9. 4.5.14. N-(4-((5-(4-Chlorophenyl)-1,3,4eoxadiazol-2-yl)methoxy)3-fluorophenyl)-4-methoxybenzenesulfonamide (1k) Yield 40%; mp 146e148 C; 1H NMR (400 MHz, CDCl3) d 8.01 (d, 2H, J ¼ 8 Hz), 7.68 (d, 2H, J ¼ 8 Hz), 7.51 (d, 2H, J ¼ 8.6 Hz), 7.04e6.97 (m, 2H), 6.89 (d, 2H, J ¼ 8 Hz), 6.76 (d, 2H, J ¼ 8 Hz), 5.32 (s, 2H), 3.83 (s, 3H); 13C NMR (100 MHz, CDCl3) d 163.5, 161.6, 160.3, 152.5, 150.0, 141.3 (d, JCF ¼ 11 Hz), 136.9, 130.7 (d, JCF ¼ 9 Hz), 128.5, 127.8, 126.7, 119.9, 116.3 (d, JCF ¼ 3 Hz), 115.8, 112.6, 109.6, 60.1, 53.9; MS [m/ z þ Na] 508.0. Compounds 1leq were prepared with the same method from 6c and the appropriate sulfonyl chlorides. 4.5.15. N-(3-fluoro-4-((5-(4-methoxyphenyl)-1,3,4eoxadiazol-2yl)methoxy)phenyl)methanesulfonamide (1l) Yield 36%; mp 122e124 C; 1H NMR (400 MHz, CDCl3) d 8.01 (d, 2H, J ¼ 8 Hz), 7.44 (s, 1H), 7.15 (t, 1H, J ¼ 8 Hz), 7.02e6.96 (m, 3H), 5.36 (s, 2H), 3.88 (s, 3H), 2.99 (s, 3H); 13C NMR (100 MHz, CDCl3) d 166.0, 162.8, 161.3, 154.5, 151.9, 143.1 (d, JCF ¼ 10 Hz), 132.4 (d, JCF ¼ 9 Hz), 129.0, 117.9, 117.4 (d, JCF ¼ 4 Hz), 114.6, 110.9, 110.7, 61.9, 55.5, 39.3. 4.5.16. N-(3-fluoro-4-((5-(4-methoxyphenyl)-1,3,4eoxadiazol-2yl)methoxy)phenyl)propane-1-sulfonamide (1m) Yield 38%; mp 132e134 C; 1H NMR (400 MHz, CDCl3) d 8.10 (d, 2H, J ¼ 8.2 Hz), 7.65 (s, 1H), 7.08e7.22 (m, 2H), 7.07e6.98 (m, 3H), 5.38 (s, 2H), 3.92 (s, 3H), 3.15 (t, 2H, J ¼ 8.2 Hz), 2.00e1.82 (m, 2H),
51
1.07 (t, 2H, J ¼ 8.2 Hz); 13C NMR (100 MHz, CDCl3) d 166.0, 162.7, 161.4, 154.5, 151.9, 142.8 (d, JCF ¼ 11 Hz), 132.7 (d, JCF ¼ 9 Hz), 128.9, 117.9, 116.8 (d, JCF ¼ 3 Hz), 114.6, 110.4, 110.2, 61.9, 55.5, 53.4, 17.2, 12.8. 4.5.17. N-(3-fluoro-4-((5-(4-methoxyphenyl)-1,3,4eoxadiazol-2yl)methoxy)phenyl)benzenesulfonamide (1n) Yield 30%; mp 138e140 C; 1H NMR (400 MHz, CDCl3) d 8.01 (d, 2H, J ¼ 8.2 Hz), 7.75 (d, 2H, J ¼ 8.2 Hz), 7.50 (s, 1H), 7.48e7.35 (m, 2H), 6.92e7.09 (m, 4H), 5.38 (s, 2H), 7.81e7.71 (m, 1H), 5.32 (s, 2H), 3.89 (s, 3H); MS [m/z þ Na] 478.2. 4.5.18. 4-Chloro-N-(3-fluoro-4-((5-(4-methoxyphenyl)1,3,4eoxadiazol-2-yl)methoxy)phenyl)benzenesulfonamide (1o) Yield 32%; mp 164e166 C; 1H NMR (400 MHz, CDCl3) d 8.01 (d, 2H, J ¼ 7.6 Hz), 7.69 (d, 2H, J ¼ 8.4 Hz), 7.39 (d, 2H, J ¼ 8.4 Hz), 7.06e7.02 (m, 4H), 6.81 (d, 1H, J ¼ 7.6 Hz), 5.35 (s, 2H), 3.89 (s, 3H); 13 C NMR (100 MHz, CDCl3) d 166.1, 162.8, 161.2, 154.3, 150.6, 144.0, 139.8 (d, JCF ¼ 11 Hz), 137.2, 131.3 (d, JCF ¼ 9 Hz), 129.4, 129.0, 128.6, 127.2, 118.5 (d, JCF ¼ 4 Hz), 117.4, 115.6, 114.6, 112.0, 111.7, 61.7, 55.5; MS [m/z þ Na] 512.0. 4.5.19. N-(3-fluoro-4-((5-(4-methoxyphenyl)-1,3,4eoxadiazol-2yl)methoxy)phenyl)-4-methylbenzenesulfonamide (1p) Yield 42%; mp 136e138 C; 1H NMR (400 MHz, CDCl3) d 8.99 (d, 2H, J ¼ 8.8 Hz), 7.63 (d, 2H, J ¼ 10.8 Hz), 7.20 (d, 2H, J ¼ 7.6 Hz), 7.03e6.98 (m, 4H), 6.77 (d, 1H, J ¼ 9.6 Hz), 5.32 (s, 2H), 3.87 (s, 3H), 2.35 (s, 3H); 13C NMR (100 MHz, CDCl3) d 165.9, 162.7, 161.4, 154.1, 151.6, 144.0, 143.1 (d, JCF ¼ 11 Hz), 135.8, 132.1 (d, JCF ¼ 9 Hz), 129.7, 128.9, 127.2, 118.1 (d, JCF ¼ 4 Hz), 117.4, 115.7, 114.6, 111.4, 111.2, 61.8, 55.5, 21.5; MS [m/z þ Na] 492.0. 4.5.20. N-(3-fluoro-4-((5-(4-methoxyphenyl)-1,3,4eoxadiazol-2yl)methoxy)phenyl)-4-methoxybenzenesulfonamide (1q) Yield 43%; mp 124e126 C; 1H NMR (300 MHz, CDCl3) d 8.99 (d, 2H, J ¼ 8.8 Hz), 7.68 (d, 2H, J ¼ 12 Hz), 7.47 (s, 1H), 7.10e6.95 (m, 4H), 6.87 (d, 2H, d ¼ 12 Hz), 5.31 (s, 2H), 3.87 (s, 3H), 3.81 (s, 3H); 13C NMR (100 MHz, CDCl3) d 165.9, 163.2, 162.7, 154.1, 151.7, 143.1 (d, JCF ¼ 11 Hz), 132.2 (d, JCF ¼ 9 Hz), 130.3, 129.4, 128.9, 118.0 (d, JCF ¼ 3 Hz), 117.4, 115.7, 114.6, 114.3, 111.4, 111.3, 61.9, 55.5, 55.4. 4.6. Cancer cell line screening at the NCI Screening against the cancer cell lines was carried out at the National Cancer Institute (NCI), Bethesda, Maryland, USA, applying the standard protocol of the NCI [27]. The human cell lines are grown in RPMI 1640 medium containing 5% fetal bovine serum and 2 mM L-glutamine. For a typical screening experiment, cells are inoculated into 96-well microtiter plates in 100 mL at plating densities ranging from 5000 to 40,000 cells/well depending on the doubling time of individual cell lines. After cell inoculation, the microtiter plates are incubated at 37 C, 5% CO2, 95% air and 100% relative humidity for 24 h prior to addition of experimental drugs. After 24 h, two plates of each cell line are fixed in situ with TCA, to represent a measurement of the cell population for each cell line at the time of drug addition (Tz). Experimental drugs are solubilized in dimethyl sulfoxide at 400-fold the desired final maximum test concentration and stored frozen prior to use. At the time of drug addition, an aliquot of frozen concentrate is thawed and diluted to twice the desired final maximum test concentration with complete medium containing 50 mg/mL gentamicin. Additional four, 10-fold or 1/2 log serial dilutions are made to provide a total of five drug concentrations plus control. Aliquots of 100 mL of these different drug dilutions are added to the appropriate microtiter wells already containing 100 mL of medium, resulting in the required final drug
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concentrations. Following drug addition, the plates are incubated for an additional 48 h at 37 C, 5% CO2, 95% air, and 100% relative humidity. For adherent cells, the assay is terminated by the addition of cold TCA. Cells are fixed in situ by the gentle addition of 50 mL of cold 50% (w/v) TCA (final concentration, 10% TCA) and incubated for 60 min at 4 C. The supernatant is discarded, and the plates are washed five times with tap water and air dried. Sulforhodamine B (SRB) solution (100 mL) at 0.4% (w/v) in 1% acetic acid is added to each well, and plates are kept for 10 min at room temperature. After staining, unbound dye is removed by washing five times with 1% acetic acid and the plates are air dried. Bound stain is subsequently solubilized with 10 mM trizma base, and the absorbance is read on an automated plate reader at a wavelength of 515 nm. For suspension cells, the methodology is the same except that the assay is terminated by fixing settled cells at the bottom of the wells by gently adding 50 mL of 80% TCA (final concentration, 16% TCA). Using the seven absorbance measurements [time zero, (Tz), control growth, (C), and test growth in the presence of drug at the five concentration levels (Ti)], the percentage growth is calculated at each of the drug concentrations levels. Percentage growth inhibition is calculated as: [(Ti Tz)/(C Tz)] 100 for concentrations for which Ti Tz [(Ti Tz)/Tz] 100 for concentrations for which Ti < Tz. Acknowledgments This work was supported by Korea Institute of Science and Technology (KIST), KIST Project (2E23720). We would like to thank the National Cancer Institute (NCI), Bethesda, Maryland, USA, for performing the in vitro anticancer testing over the cell lines. Appendix A. Supplementary data Supplementary data related to this article can be found at http:// dx.doi.org/10.1016/j.ejmech.2014.11.011. References [1] H. Frankish, Lancet 361 (2003) 1278.
[2] D. Belpomme, P. Irigaray, A.J. Sasco, J.A. Newby, V. Howard, R. Clapp, L. Hardell, Int. J. Oncol. 30 (2007) 1037e1049. [3] R. Gudipati, R.N.R. Anreddy, S. Manda, Saudi Pharm. J. 19 (2011) 153e158. [4] M. Rashid, A. Husain, R. Mishra, Eur. J. Med. Chem. 54 (2012) 855e866. [5] A. Husain, M. Rashid, R. Mishra, S. Parveen, D.-S. Shin, D. Kumar, Bioorg. Med. Chem. Lett. 22 (2012) 5438e5444. [6] P. Puthiyapurayil, B. Poojary, C. Chikkanna, S.K. Buridipad, Eur. J. Med. Chem. 53 (2012) 203e210. [7] J. Sun, H. Zhu, Z.-M. Yang, H.-L. Zhu, Eur. J. Med. Chem. 60 (2013) 23e28. [8] S. Zhang, Y. Luo, L.-Q. He, Z.-J. Liu, A.-Q. Jiang, Y.-H. Yang, H.-L. Zhu, Bioorg. Med. Chem. 21 (2013) 3723e3729. [9] N.B. Patel, A.C. Purohit, D.P. Rajani, R. Moo-Puc, G. Rivera, Eur. J. Med. Chem. 62 (2013) 677e687. [10] S. Rapolu, M. Alla, V.R. Bommena, R. Murthy, N. Jain, V.R. Bommareddy, M.R. Bommineni, Eur. J. Med. Chem. 66 (2013) 91e100. [11] P. Li, L. Shi, X. Yang, L. Yang, X.-W. Chen, F. Wu, Q.-C. Shi, W.-M. Xu, M. He, D.Y. Hu, B.-A. Song, Bioorg. Med. Chem. Lett. 24 (2014) 1677e1680. [12] Z. Hadady, M. Toth, L. Somsak, ARKIVOC 7 (2004) 140e149. [13] M.J. Ahsan, J.G. Samy, H. Khalilullah, M.S. Nomani, P. Saraswat, R. Gaur, A. Singh, Bioorg. Med. Chem. Lett. 21 (2011) 7246e7250. [14] F. Liu, X.Q. Luo, B.A. Song, P.S. Bhadury, S. Yang, L.H. Jin, W. Xue, D.Y. Hu, Bioorg. Med. Chem. 16 (2008) 3632e3640. [15] V. Jakubkiene, M.M. Burbuliene, E. Udrenaite, V. Garaliene, P. Vainilavicius, Pharmazie 57 (2002) 610e613. [16] N.C. Desai, N. Bhatt, H. Somani, A. Trivedi, Eur. J. Med. Chem. 67 (2013) 54e59. [17] P.N. Ibrahim, D.R. Artis, R. Bremer, G. Habets, S. Mamo, M. Nespi, C. Zhang, J. Zhang, Y.-L. Zhu, R. Zuckerman, B. West, Y. Suzuki, J. Tsai, K.-P. Hirth, G. Bollag, W. Spevak, H. Cho, S.J. Gillette, G. Wu, H. Zhu, S. Shi, WO2007002433, Jan 4th, 2007. [18] M.M. Ghorab, M.A. Shaaban, H.M. Refaat, H.I. Heiba, S.S. Ibrahim, Eur. J. Med. Chem. 53 (2012) 403e407. [19] J. Lal, S.K. Gupta, D. Thavaselvam, D.D. Agarwal, Eur. J. Med. Chem. 64 (2013) 579e588. [20] Z.-C. Wang, Y.-J. Qin, P.-F. Wang, Y.-A. Yang, Q. Wen, X. Zhang, H.-Y. Qiu, Y.T. Duan, Y.-T. Wang, Y.-L. Sang, H.-L. Zhu, Eur. J. Med. Chem. 66 (2013) 1e11. [21] X.-M. Wang, J. Xu, Y.-P. Li, H. Li, C.-S. Jiang, G.-D. Yang, S.-M. Lu, S.-Q. Zhang, Eur. J. Med. Chem. 67 (2013) 243e251. [22] F. Xu, H. Xu, X. Wang, L. Zhang, Q. Wen, Y. Zhang, W. Xu, Bioorg. Med. Chem. 22 (2014) 1487e1495. da, A.L.P. de Miranda, [23] P.C. Lima, L.M. Lima, K.C.M. da Silva, P.H.O. Le C.A.M. Fraga, E.J. Barreiro, Eur. J. Med. Chem. 35 (2000) 187e203. [24] S. Cao, X.H. Quin, G.H. Song, B. Chia, Z.S. Jiang, J. Agric. Food Chem. 51 (2003) 152e155. [25] NCI website: www.dtp.nci.nih.gov. (retrieved on Oct. 31st, 2014). [26] DTP Data Search: http://dtp.nci.nih.gov/dtpstandard/dwindex/index.jsp. (retrieved on Oct. 31st, 2014). [27] DTP Human Tumor Cell Line Screen Process: http://www.dtp.nci.nih.gov/ branches/btb/ivclsp.html. (retrieved on Oct. 31st, 2014).
