ISSN 2090-424X Journal of Basic and Applied Chemistry www.textroad.com
*Corresponding Author: Dr. Helmy I. Heiba, Professor of Applied Organic Chemistry, Department of Drug Radiation
Research, National Center for Radiation Research and Technology, Atomic Energy Authority, P.O. Box 29, Nasr City, Cairo, Egypt. E-mail: [email protected]
Synthesis of Some Novel Sulfonamides Containing Biologically Active Alkanoic Acid, Acetamide, Thiazole, and Pyrrole Moieties of Expected Antitumor and
Radiosensitizing Activities
Mostafa M. Ghorab1, Fatma A. Ragab2, Helmy I. Heiba3*, Hebaallah M. Agha3
1 Medicinal, Aromatic and Poisonous Plants Research Center (MAPPRC), College of Pharmacy, King Saud University, Saudi Arabia
2 Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Cairo University, Cairo, Egypt 3 Department of Drug Radiation Research, National Center for Radiation Research and Technology, Atomic
Energy Authority, Cairo, Egypt
Accepted 9 May, 2011.
ABSTRACT Several novel N4 substituted sulfapyridine derivatives were synthesized by condensation of 2-Chloro-N-(4-(N-pyridin-2-ylsulfamoyl)phenyl) acetamide 2 with different amino acids, amines, malononitrile, or ammonium thiocyanate to yield the corresponding alkanoic acid derivatives 3a-f, acetamide derivatives 4- 8, pyrrole derivatives 9- 12, and thiazolidinone derivative 13. The newly synthesized compounds were screened in-vitro against human breast cancer cell line (MCF-7). Compounds 3b, 10 and 12 showed the highest activity with IC50 values 26.1, 22.4, 22.1 µM compared to doxorubicin as a reference drug. The promising compounds 3b, 10 and 12 showed the ability to sensitize cancer cells to the lethal effects of ionizing radiation. KEY WORDS: Sulfonamides, Cytotoxic activity and γ-radiation.
INTRODUCTION
A large number of structurally novel sulfonamide derivatives have ultimately been reported to show substantial in-vitro and in-vivo antitumor activity [1-8]. (E7010), (ER-34410), (E7070) are examples in advanced clinical trials. It is also reported that different N1 substituted sulfonamide show antiproliferative activities [9-11]. Recently, a series of sulfapyridine derivatives having thiazole moiety were found to exhibit cytotoxic activity against breast carcinoma cell line (MCF-7), where the 4-[(pyridin-2-ylamino) sulfonyl] benzene was considered as the pharmacophoric moiety [12]. From literature survey, it was reported that sulfonamides bearing amino acid moieties were reported to possess antitumor activity [13]. In the present work, some new N4 alkanoic acid, acetamide, pyrrole and thiazole derivatives attached to a sulfonamide moiety were synthesized with the aim of obtaining hybrid molecules displaying potent antitumor activity. Also the synergism of the most potent synthesized derivatives with γ- radiation was studied. Since it is reported that combining chemotherapy with radiotherapy is effective for cancer patients as it decreases the side effects of both drugs and radiation, consequently, this study was conducted to evaluate the ability of the tested compounds as cytotoxic agents to enhance the cell killing effect of γ-radiation to predict the augmenting effect of γ –irradiation [14].
MATERIALS AND METHODS
Chemistry
Melting points are uncorrected and were determined on a Stuart melting point apparatus (Stuart Scientific, Redhill, UK). Elemental analysis (C, H, N) were performed on Perkin-Elmer 2400 analyser (Perkin-Elmer, Norwalk, CT, USA) at the microanalytical laboratories of the Faculty of Science, Cairo University. All compounds were within ±0.4% of the theoretical values. The IR spectra (KBr) were measured on Shimadzu IR 110 spectrophotometer (Shimadzu, Koyoto, Japan), 1H-NMR spectra were obtained on a Bruker proton NMR-Avance 300 (300 MHz) (Bruker, Munuch, Germany), in DMSO-d6 as a solvent, using tetramethylsilane (TMS) as internal standard. Mass spectra were run on JEOL JMS AX-500 mass spectrometer (JEOL JMS, Tokyo, Japan). All reactions were monitored by thin layer chromatograph (TLC)
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using precoated Aluminium sheets Silica gel Merck 60 F254. Ethyl acetate/Cyclohexane (2.5:7.5 mL) mixture was used as eluting solvent and and TLC sheets were visualized by UV lamp (Merck, Damstadt, Germany). 2-(2-oxo-2-(4-(N-pyridin-2-ylsulfamoyl) phenylamino) ethylamino) alkanoic acid (3a-f)
General procedure: An amino acid namely; glycin, alanin, isoleucine, aspartic acid, phenylalanine, tyrosine (0.0096 mol) and sodium carbonate (0.57 g, 0.0054 mol) were dissolved in water (10 mL), and the solution was adjusted to pH 9.0–9.5. The chloro derivative 2 (1.56 g, 0.0048 mol) was then added, and the mixture was stirred at 100 ◦ C for 6h. The reaction mixture was left overnight at room temperature, filtered and then treated with formic acid. The solid obtained was crystallized from dioxane [15]. 2-(2-oxo-2-(4-(N-pyridin-2-ylsulfamoyl) phenylamino) ethylamino) acetic acid (3a)
General procedure: A mixture of chloro compound (2) (0.65g, 0.002 mol) and different amines namely; diethylamine, 3-aminopropanol, piperidine, morpholine, and 4-chloroaniline (0.002 mol) in ethanol (20 ml) was refluxed for 2 h. The reaction mixture was concentrated under vacuum and poured onto cold water (100 ml), the obtained solid was filtered, washed with water and crystallized from acetonitrile to give (4- 8), respectively. 2-(Diethylamino)-N-(4-(N-pyridin-2-ylsulfamoyl) phenyl) acetamide (4)
Microanalytical and spectral data are as reported [17]. N-(3-Cyano-5-oxo-1-(4-(N-pyridin-2-ylsulfamoyl)phenyl)-4,5-dihydro-1H-pyrrol-2-yl) acetamide (10), N-Acetyl-N-(3-Cyano-5-oxo-1-(4-(N-pyridin-2-ylsulfamoyl) phenyl)-4,5-dihydro-1H-pyrrol-2-yl) acetamide (11)
A solution of compound 9 (0.355 g, 0.001 mol) in acetic anhydride (20 ml) was refluxed for 2 h, the reaction mixture was then concentrated, and the solid separated was crystallized from ethanol to give 10, while compound 11 was obtained by refluxing for 15 h. Microanalytical and spectral data of 10: Yield 65%, m.p. 160-162°C, anal. Calcd. for C18H15N5O4S (397), C, 54.40; H, 3.80; N, 17.62. Found: C, 54.66; H, 4.94; N, 17.37. IR (KBr, cm-1): 3446, 3351 (NH), 3001 (CH arom.), 2933, 2856 (CH aliph.), 2215 (C≡N), 1716, 1639 (2C=O), 1369, 1174(SO2). 1H-NMR (DMSO-d6, δ ppm): 2.0 [s, 3H, COCH3], 2.70 [s, 2H, CH2, pyrrole], 6.79- 8.10 [m, 9H, Ar-H + NHCO, D2O exchangeable], 10.78 [s, 1H, SO2NH, D2O exchangeable]. MS (m/z): 397 (M+, 0.61%), 92 (100%). Microanalytical and spectral data of 11: Yield 68%, m.p. 220-222°C, anal. Calcd. for C20H17N5O5S (439), C, 54.66; H, 3.90; N, 15.94. Found: C, 54.98; H, 3.67; N, 15.62. IR (KBr, cm-1): 3455 (NH), 3035 (CH arom.), 2929, 2858 (CH aliph.), 2215 (C≡N), 1710, 1625 (3 C=O), 1371, 1172(SO2). 1H-NMR (DMSO-d6, δ ppm): 1.92 [s, 6H, 2COCH3], 2.76 [s, 2H, CH2, pyrrole], 6.85- 8.20 [m, 9H, Ar-H + NHCO, D2O exchangeable], 10.79 [s, 1H, SO2NH, D2O exchangeable]. MS (m/z): 439 (M+, 2.10 %), 62 (100%). 4-(4-Cyano-5-(3-ethylthioureido)-2-oxo-2,3-dihydro-1H-pyrrol-1-yl)-N-(pyridin-2-yl)benzenesulfonamide(12)
A mixture of compound 9 (0.355 g, 0.001 mol) and ethyl isothiocyanate (0.086 g, 0.001 mol) in dimethylformamide (20 ml), and a catalytic amount of triethylamine, was refluxed for 10 h. The reaction mixture was cooled then poured onto cold water, and the solid obtained was crystallized from ethanol to give 12. Yield 54 %, m.p. 173-175°C, anal. Calcd. for C19H18N6O3S2 (442), C, 51.57; H, 4.10; N, 18.99. Found: C, 51.35; H, 4.46; N, 18.65. IR (KBr, cm-1): 3471, 3336 (NH), 3070 (CH arom.), 2927, 2863 (CH aliph.), 2192 (C≡N), 1646 (C=O), 1257 (C=S), 1384, 1135(SO2). 1H-NMR (DMSO-d6, δ ppm): 1.16[t, 3H, CH3], 2.74 [s, 2H, CH2, pyrrole], 4.45 [q, 2H, CH2], 6.87- 8.21 [m, 10H, Ar-H + 2 NH, D2O exchangeable], 10.75 [s, 1H, SO2NH, D2O exchangeable]. MS (m/z): 442 (M+, 2.84%), 65 (100%). 4-(4-Oxo-4, 5-dihydrothiazol-2-ylamino)-N-(pyridin-2-yl) benzenesulfonamide (13)
A solution of compound 2 (0.325g, 0.001 mol) and Ammonium thiocyanate (0.076g, 0.001 mol) in ethanol (20 ml) was refluxed for 3 h. The reaction mixture was filtered while hot and the obtained solid was crystallized from dioxane to give 13: Yield 85 %, m.p. 240-242°C, anal. Calcd. for C14H12N4O3S2 (348): C, 48.26; H, 3.47; N, 16.08. Found: C, 48.59; H, 3.26; N, 16.32. IR (KBr, cm-1): 3448-3366 (OH), 3286, 3191 (NH), 3047(CH arom.), 1397, 1137 (SO2). 1H-NMR (DMSO-d6, δ ppm): 6.86 [s, 1H, CH, thiazole], 6.91-8.00 [m, 9H, Ar-H + NH, D2O exchangeable] 10.84 [s, 1H, SO2NH, D2O exchangeable], 11.71 [s, 1H, OH, D2O exchangeable]. MS (m/z): 348 (M+, 1.7 %), 184 (100%). Biological Testing Facilities
The in-vitro anticancer screening was done at the National Cancer Institute, Cairo University, Egypt. Irradiation was performed in the National Center for Radiation Research and Technology, Atomic Energy Authority, Cairo, Egypt using Gamma cell-40 (137Cs) source. In-vitro anticancer screening
The cytotoxic activity for the newly synthesized compounds was evaluated against human breast cancer cell line (MCF-7) in-vitro using the Sulfo-Rhodamine-Bstain (SRB) assay by the method of Skehan et al. [18]. The relation between surviving fraction and drug concentration (μM /L) is plotted to get the best fitting survival curve for breast tumor cell line after the specified time. The molar concentration required for 50% inhibition of cell viability (IC50) was calculated by solving the third order polynomial equation using simple designed software written on ‘Matlab R2008a’ program. The results are given in Table 1.
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Table 1 – In-vitro anticancer screening of the synthesized compounds against human breast cancer cell line (MCF-7). Cpd. No. Compound concentration (µM) IC50 (µM)
#: Each value is the mean of three experiments ± standard error.
Radiosensitizing activity
The most active compounds were reevaluated again in combination with γ-radiation using the SRB assay. Cells were subjected to a single dose of γ-radiation at a dose level of 8 Gy with a dose rate of 2Gy/min. The surviving fractions were expressed as means ± standard error. The results were analyzed using 1-way ANOVA test and the percentage of change in the surviving fraction from control for each compound was calculated considering the control as 100% in the surviving fraction. The results are given in Table 2.
Table 2: In-vitro anticancer screening of compounds 3b, 10 and 12 against human breast cancer cell line (MCF-7) in combination with γ-radiation
#: Each value is the mean of three values ± Standard Error *: Significant difference from control group at p<0.001
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RESULTS AND DISCUSSION Chemistry
The starting key material 2-Chloro-N-(4-(N-pyridin-2-ylsulfamoyl) phenyl) acetamide 2 was prepared by reaction of 4-amino-N-(pyridin-2-yl) benzenesulfonamide with chloroacetylchloride [19]. The chloroaceta-mide derivative 2 was treated with the sodium salt of various amino acids under reflux to afford the corresponding 2-(2-oxo-2-(4-(N-pyridin-2-ylsulfamoyl) phenylamino) ethylamino) alkanoic acid derivatives 3a-f (scheme 1). N4 - (4-(N-pyridin-2-ylsulfamoyl) phenyl) acetamide derivatives 4-8 were obtained by refluxing chloroacetamide derivative 2 with different amines in ethanol for 2 h. (scheme 2). Compounds 6 and 7 were reported to be obtained in a yield of 70 % and 61 %, respectively. [16], while they were obtained at higher yields by conducting the reaction in dioxane. Compound 9 was reported to be obtained by treatment of compound 2 with malononitrile for 24 h. in dioxane [17]. When compound 9 was refluxed in acetic anhydride for 2 h. the corresponding monoacetyl derivative 10 was obtained, while the diacetyl derivative 11 was obtained at long time (15 h.). Refluxing compound 9 with ethyl isothiocyanate in dimethylformamide for 10 h. yielded the thioureido derivative 12. Treatment of compound 2 with ammonium thiocyanate in ethanol yielded the corresponding thiazolidinone derivative 13, via intramolecular cyclization [20] (Scheme 3).
Scheme 1 Scheme 2
Scheme 3
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In-vitro anticancer screening and Radiosensitizing activity
The in-vitro cytotoxic screening of all the synthesized compounds against human breast cancer cell line (MCF-7) showed that the propanoic acid derivative 3b, the pyrrole derivative bearing monoacetyl moiety 10 and the pyrrole derivative carrying thioureido moiety 12 were found to be the most active compounds with IC50 values 26.1, 22.4, 22.1 µM, respectively, compared to doxorubicin as a reference drug (IC50 = 26.1µM) Table 1. Reevaluation of compounds 3b, 10 or 12 in combination with γ -radiation resulted in a remarkable decrease of their IC50 values to 18.1, 20.3, and 10.8 μM, respectively, Table 2.
The pyrrole derivatives (9-12) (IC50= 22.16 – 30.31 µM) possess higher cytotoxic activity than the amino acid, the acetamide and thiazole derivatives. The most active compounds in this class are the monoacetyl derivative 10 and the thioureido derivative 12 which were found to be more active than doxorubicin, while the amino acid derivatives 3 (a-f) (IC50= 26.10 – 38.70 µM), the most active compound in this class is the alanin derivative 3b which showed activity similar to that of the reference drug, Concerning the acetamide derivatives (4-8), (IC50= 27.5 – 45.28 µM), the most active compounds in this class are the piperidine derivative 6 and the 4-chloroaniline derivative 8 which lead us to conclude that cyclic aliphatic and aromatic moieties may be promising nucleus as cytotoxic agents. The preliminary results of biological screening of the tested compounds indicate that the pyrrole derivatives were the most active on (MCF7) than the amino acid, the acetamide and thiazole derivatives.
CONCLUSION
Using combination of compounds 3b, 10 or 12 and ionizing radiation synergistically enhanced growth inhibition on breast cancer, which showed the ability of these compounds to sensitize cancer cells to the lethal effects of ionizing radiation. This illustrates the importance of the combination therapy for the patients with cancer to decrease the side effects of both drugs and γ- radiation.
The authors have declared no conflict of interest.
REFERENCES
1. Abbate, F., A. Casini, T. Owa, A. Scozzafava and C.T. Supuran, 2004. Carbonic anhydrase inhibitors: E7070, a sulfonamide anticancer agent, potently inhibits cytosolic isozymes I and II, and transmembrane, tumorassociated isozyme IX. Bioorg. Med. Chem. Lett., 14, 217-223.
2. Ghorab, M.M., F.A. Ragab, H.I. Heiba, R.K. Arafa and E.M. El-Hossary, 2011. Docking study, in vitro anticancer screening and radiosensitizing evaluation of some new fluorine-containing quinoline and pyrimidoquinoline derivatives bearing a sulfonamide moiety., Med. Chem. Res., 20, 388-400
3. Ghorab, M.M., F.A. Ragab and M.M. Hamed, 2009. Design, synthesis and anticancer evaluation of novel tetrahydroquinoline derivatives containing sulfonamide moiety. Eur. J. Med. Chem., 44, 4211–4217.
4. Ghorab, M.M., F.A. Ragab, H.I. Heiba, H.A. Youssef and M.G. El-Gazzar, 2010. Synthesis of some new pyrazole and pyrimidine derivatives carrying a sulfonamide moiety of expected antitumor activity and study of the synergistic effect of gamma-irradiation. Arzneim. Forsch.. 60, 48-55
5. Al-Said, M.S., M.M. Ghorab, S.I. Al-qasoumi, E. M. El-Hossary and E. Noaman, 2010. Synthesis and in vitro anticancer screening of some novel 4-[2-amino-3-cyano-4-substituted-5,6,7,8-tetrahydroquinolin-1-(4H)-yl] Benzenesulfonamides Eur. J. Med .Chem., 45, 3011-3018
6. Ghorab, M.M., F.A. Ragab, H.I. Heiba, H.A. Youssef and M.G. El-Gazzar, 2010. Synthesis of novel pyrrole and pyrrolo[2,3-d]pyrimidine derivatives bearing sulfonamide moiety for evaluation as anticancer and radiosensitizing agents. Bioorg. Med. Chem. Lett., 20, 6316–6320
7. Ghorab, M.M., F.A. Ragab and M.M. Hamed, 2010. Synthesis of novel pyrrole and pyrrolo[2,3-d]pyrimidine derivatives bearing sulfonamide moiety for evaluation as anticancer and radiosensitizing agents. Arzneim. Forsch., 60, 141-148
8. Ghorab, M.M., E. Noaman, M.M. Ismail, H.I. Heiba, Y.A. Ammar and M.Y. Sayed, 2006. Novel antitumor and radioprotective sulfonamides containing pyrrolo[2,3-d]pyrimidines. Arzneim. Forsch., 56, 405-413.
9. Calvert, N., T.A. Connors and W.C.J. Ross, 1968. Aryl-2-halogenoalkylamines: XXV. Derivatives of sulphanilamide designed for selective deposition in neoplastic tissue. Eur. J. Cancer, 4, 627-636.
13
J. Basic. Appl .Chem., 1(2)8-14, 2011
10. Huang, Z., Z. Lin and J. Huang, 2001. A novel kind of antitumor drugs using sulfonamide as potent compound. Eur. J. Med. Chem., 36, 863-872.
11. Huang, Z., G. Yang, Z. Lin, and J. Huang, 2001. 2-[N1-2-pyrimidyl-aminobenzene sulfonamido]ethyl 4-bis(2-chloroethyl)aminophenylbutyrate: a potent antitumor agent. Bioorg. Med. Chem. Lett., 11 1099–1103
12. Kamel, M.M., H.I. Ali, M.M. Anwar, N.A. Mohameda and A.M. Soliman, 2010. Synthesis, antitumor activity and molecular docking study of novel Sulfonamide-Schiff’s bases, thiazolidinones, benzothiazinones and their C-nucleoside derivatives. Eur. J. Med. Chem45, 572–580.
13. Mastrolorenzo, A., A. Scozzafava and C. T. Supuran, 2000. 4-toluenesulfonylureido derivatives of amines, amino acids and dipeptides: a novel class of potential antitumor agents. Eur. J. Pharm. Sci. 11, 325–332
14. Nishimura, Y. 2004. Rational of chemotherapy. Int. J. Clin. Oncol. 9, 414-420.
15. Alqasoumi, S.I., Ragab, F.A., Alafeefy, A.M., Galal, M., Ghorab, M.M. 2009. Radioprotective and Antitumor Activity of Some Novel Amino Acids and Imidazoles Containing Thieno [2,3-d]pyrimidine Moiety”: Phosphorus, Sulfur, and Silicon. 184, 12, 3241-3257
16. Elzbawy, S. R., A.A. Abdel-Wahab and A. A. Khalaf, 1982. Synthesis of some new biologically active 2- Aminopyridine derivatives. Rev. Roum. Chim. 27, 537-544.
17. El-Gaby, M.S., A.M. Gaber, A.A. Atalla and K.A. Abd Al-Wahab, 2002. Novel synthesis and antifungal activity of pyrrole and pyrrolo[2,3-d]pyrimidine derivatives containing sulfonamido moieties. Farmaco, 57, 613-617.
18. Skehan, P., R. Storeng, D. Scudiero, A. Monks, J. McMahon, D. Vistica, J.T. Warren, H. Bokesch, S. Kenney and M.R. Boyd, 1990. New colorimetric cytotoxicity assay for anticancer-drug screening. J. Natl. Cancer Inst., 82, 1107-1112.
20. Vicini, P., A. Geronikaki, K. Anastasia, M. Incertia and F. Zania, 2006, Synthesis and antimicrobial activity of novel 2-thiazolylimino-5-arylidine-4-thiazolidinines Bioorg. Med. Chem., 14, 3859-3864.
