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Synthesis and NHE1 inhibitory activity of ligustrazine derivatives
Chinese Chemical Letters 18 (2007) 539–541 www.elsevier.com/locate/cclet
Synthesis and NHE1 inhibitory activity of ligustrazine derivatives Mei Ren a, Yun Gen Xu a,*, Na Wen b, Da Yong Zhang a, Wei Yi Hua a a
b
Center of Drug Discovery, China Pharmaceutical University, Nanjing 210009, China Department of Pharmacology, China Pharmaceutical University, Nanjing 210009, China Received 29 December 2006
Abstract A series of novel derivatives of ligustrazine linked with substituted benzoyl guanidine were synthesized. These compounds have not been reported in literature, and their chemical structures were confirmed by IR, 1H NMR and MS. The results of NHE1 inhibitory activity test showed that compounds I2, I3, I4, I6, and I7 possess more potent NHE1 inhibitory activity than cariporide. # 2007 Yun Gen Xu. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved. Keywords: Na+/H+ exchanger inhibitors; Ligustrazine derivatives; Benzoyl guanidine derivatives; Synthesis
Ligustrazine (tetramethylpyrazine, TMP, Fig. 1) is a major active component of the Chinese traditional medicine Chuanxiong (Ligusticum wallichii Franchat), which was introduced on the market during the early 1970s and can now be synthesized chemically. Ligustrazine is widely used in China as a calcium channel antagonist for the treatment of coronary atherosclerotic disease and ischemic cerebrovascular disease [1]. Ligustrazine also shows good protective effect on myocardial injury during ischemia and reperfusion [2]. The Na+/H+ exchanger (NHE) comprises a family of membrane proteins that are involved in the transport of H+ in exchange for Na+ [3]. They are regulated by the intracellular pH through interaction of H+ with a sensor site of the exchanger protein. To date, nine NHE isoforms (NHE1–NHE9) have been identified in various organs in the human body. NHE1 is the ubiquitously distributed in tissue, and is the predominant NHE isoform in the myocardial cell. It is rapidly activated during ischemia by intracellular H+ accumulation that is exchanged for Na+. The high intracellular Na+ level leads to an increase in intracellular Ca2+ via Na+/Ca2+ exchange [4]. At the cardiac level, this cellular Ca2+ overload is involved in ischemic and reperfusion injuries like myocardial infarction activation, stunning and tissue necrosis [5]. Thus, seeking of effective NHE1 inhibitors to prevent cardiomyocyte death, angina and arrhythmia caused by ischemia-reperfusion episodes has become a focus in new drug research. Acyl guanidine is the common character existed in most of NHE1 inhibitors, such as cariporide (Fig. 1), and it is considered as the functional group for keeping the drugs’ potential. Based on the structure of cariporide, ligustrazine was modified by substituting one of the methyl groups with various benzoyl guanidine (Table 1). Compounds I1–7 [6] were synthesized from tetramethylpyrazine 1 by bromization with NBS to form 2-bromomethyl-3,5,6-trimethylpyrazine 2 which was reacted with ethyl benzoate derivatives 3 to afford intermediates 4, followed by treatment with guanidine (Scheme 1).
* Corresponding author. E-mail address: [email protected] (Y.G. Xu). 1001-8417/$ – see front matter # 2007 Yun Gen Xu. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved. doi:10.1016/j.cclet.2007.03.020
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M. Ren et al. / Chinese Chemical Letters 18 (2007) 539–541
Fig. 1. Structures of ligustrazine and cariporide. Table 1 Chemical structure of 3a–g, 4a–g and ligustrazine derivatives I1–7 Compound
–O–a
Ra
3a, 4a, I1 3b, 4b, I2 3c, 4c, I3 3d, 4d, I4 3e, 4e, I5 3f, 4f, I6 3g, 4g, I7
4 4 4 4 4 2 2
H 3-Cl 3-NO2 3-Br 3-OCH3 H 4-CF3
a
Based on benzoyl guanidine.
Scheme 1. Synthetic route of target compounds I1–7. Reagents and conditions: (a) NBS, CCl4, hn, reflux; (b) Na2CO3, acetone or DMF, reflux. (c) (i) Guanidine, isopropyl alcohol, reflux; (ii) HCl.
Compounds I1–7 are the new compounds, and their structures were confirmed by IR, 1H NMR, MS and elemental analysis. The NHE1 inhibitory activities of seven target compounds were evaluated in platelet swelling assay (PSA) and compared to cariporide. The NHE1 inhibitory activities were measured by the decrease of optical density (OD), which induced by platelet swelling [7]. The results showed that all compounds possess NHE1 inhibitory effects. Among these, the inhibitory effects on NHE1 of compound I2, I3, I4, I6 and I7 are superior to the control cariporide (Table 2). Activity of compound I4 is 21 times more potent than cariporide. Ligustrazine showed no effect on NHE1 under this experimental condition, Table 2 IC50 of cariporide, ligustrazine and compounds I1–7 for inhibition of NHE1 Compound
IC50 (10
Cariporide I1 I3 I5 I7
6.50 20.4 0.57 3.28 1.17
8
mol/L)
Compound
IC50 (10
TMP I2 I4 I6
NA 1.47 0.30 0.66
8
mol/L)
M. Ren et al. / Chinese Chemical Letters 18 (2007) 539–541
541
suggesting that its protective effect on myocardial injury during ischemia and reperfusion was not regulated via NHE1 system. The preliminary pharmacological tests showed that p-(3,5,6-trimethylpyrazine-2-ylmethoxy) substituted benzoyl guanidine derivatives exhibit stronger inhibitory effects on NHE1 than o-(3,5,6-trimethylpyrazine-2-ylmethoxy) substituted benzoyl guanidine derivatives, and that introducing electron-withdrawing substituents to benzene ring improve the inhibitory potency of NHE1. The above results gave us useful enlightenment for further design of novel NHE1 inhibitors. In conclusion, a series of novel derivatives of ligustrazine linked with substituted benzoyl guanidine were synthesized. Some of them showed better NHE1 inhibitory activity than cariporide, and the most active one was compound I4 [8]. Structure–activity relationships were briefly discussed. The protective effect of compound I4 on myocardial injury during ischemia and reperfusion in animal models is underway. References [1] [2] [3] [4] [5] [6]
X.C. Cheng, X.Y. Liu, W.F. Xu, Drugs Future 30 (10) (2005) 1059. Z.G. Wu, J. Wuhan Inst. Chem. Technol. 25 (1) (2003) 28. L. Fliegel, H. Wang, J. Mol. Cell. Cardiol. 29 (1997) 1991. B. Masereel, L. Pochet, D. Laeckmann, Eur. J. Med. Chem. 38 (2003) 547. H.M. Piper, C. Balser, Y.V. Ladilov, et al. Basic Res. Cardiol. 91 (1996) 191. Typical procedure for the synthesis of 4a–g: The mixture of compound 2 (10 mmol), compound 3 (10 mmol) and Na2CO3 (5 mmol) in 50 mL acetone was refluxed for 48 h (check by TLC) and filtrated, and the solvent was evaporated in vacuo. Typical procedure for the synthesis of I1–7: A mixture of guanidine (3.5 mmol) and 3 mL isopropyl was stirred at 70 8C, to it was added dropwise a solution of compound 4 (0.35 mmol) dissolved in 2 mL isopropyl. The mixture was refluxed for 30 min (checked by TLC). Adjust the pH to 2 by hydrochloride, filtered, and the filtrate was evaporated in vacuo. The final product was purified by silica gel column chromatography (eluent: dichloromethane: methanol = 20:1–10:1). [7] D. Rosskopf, E. Morgenstern, W. Scholz, et al. Hypertension 9 (3) (1991) 231. [8] Selected spectroscopic data: I4: mp 175–176 8C, yield: 52.1%; IR (KBr, cm 1) 3380, 3189, 2921. 1706 (nC O), 1657 (nC N), 1595, 1551 (nC C), 1265 (nC–O–C); 1H NMR (500 MHz, DMSO-d6, d ppm): 2.50 (s, 6H, –CH3), 2.59 (s, 3H, –CH3), 5.29 (s, 2H, –CH2–O), 7.20 (d, 1H, J = 8.8 Hz, ArH), 7.65 (bs, 2H, NH), 8.06 (dd, 1H, J1 = 2.0 Hz, J2 = 8.8 Hz, ArH), 8.32 (d, 1H, J = 2.0 Hz, ArH); MS(ESI(+)70 V, m/z): 394.0 ([M+H]+, base peak). Anal. Calcd. for C16H18BrN5O2HClH2O: C 43.02, H 4.74, N 15.68, found: C 42.87, H 4.82, N 15.57.
Synthesis and NHE1 inhibitory activity of ligustrazine derivatives Mei Ren a, Yun Gen Xu a,*, Na Wen b, Da Yong Zhang a, Wei Yi Hua a a
b
Center of Drug Discovery, China Pharmaceutical University, Nanjing 210009, China Department of Pharmacology, China Pharmaceutical University, Nanjing 210009, China Received 29 December 2006
Abstract A series of novel derivatives of ligustrazine linked with substituted benzoyl guanidine were synthesized. These compounds have not been reported in literature, and their chemical structures were confirmed by IR, 1H NMR and MS. The results of NHE1 inhibitory activity test showed that compounds I2, I3, I4, I6, and I7 possess more potent NHE1 inhibitory activity than cariporide. # 2007 Yun Gen Xu. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved. Keywords: Na+/H+ exchanger inhibitors; Ligustrazine derivatives; Benzoyl guanidine derivatives; Synthesis
Ligustrazine (tetramethylpyrazine, TMP, Fig. 1) is a major active component of the Chinese traditional medicine Chuanxiong (Ligusticum wallichii Franchat), which was introduced on the market during the early 1970s and can now be synthesized chemically. Ligustrazine is widely used in China as a calcium channel antagonist for the treatment of coronary atherosclerotic disease and ischemic cerebrovascular disease [1]. Ligustrazine also shows good protective effect on myocardial injury during ischemia and reperfusion [2]. The Na+/H+ exchanger (NHE) comprises a family of membrane proteins that are involved in the transport of H+ in exchange for Na+ [3]. They are regulated by the intracellular pH through interaction of H+ with a sensor site of the exchanger protein. To date, nine NHE isoforms (NHE1–NHE9) have been identified in various organs in the human body. NHE1 is the ubiquitously distributed in tissue, and is the predominant NHE isoform in the myocardial cell. It is rapidly activated during ischemia by intracellular H+ accumulation that is exchanged for Na+. The high intracellular Na+ level leads to an increase in intracellular Ca2+ via Na+/Ca2+ exchange [4]. At the cardiac level, this cellular Ca2+ overload is involved in ischemic and reperfusion injuries like myocardial infarction activation, stunning and tissue necrosis [5]. Thus, seeking of effective NHE1 inhibitors to prevent cardiomyocyte death, angina and arrhythmia caused by ischemia-reperfusion episodes has become a focus in new drug research. Acyl guanidine is the common character existed in most of NHE1 inhibitors, such as cariporide (Fig. 1), and it is considered as the functional group for keeping the drugs’ potential. Based on the structure of cariporide, ligustrazine was modified by substituting one of the methyl groups with various benzoyl guanidine (Table 1). Compounds I1–7 [6] were synthesized from tetramethylpyrazine 1 by bromization with NBS to form 2-bromomethyl-3,5,6-trimethylpyrazine 2 which was reacted with ethyl benzoate derivatives 3 to afford intermediates 4, followed by treatment with guanidine (Scheme 1).
* Corresponding author. E-mail address: [email protected] (Y.G. Xu). 1001-8417/$ – see front matter # 2007 Yun Gen Xu. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved. doi:10.1016/j.cclet.2007.03.020
540
M. Ren et al. / Chinese Chemical Letters 18 (2007) 539–541
Fig. 1. Structures of ligustrazine and cariporide. Table 1 Chemical structure of 3a–g, 4a–g and ligustrazine derivatives I1–7 Compound
–O–a
Ra
3a, 4a, I1 3b, 4b, I2 3c, 4c, I3 3d, 4d, I4 3e, 4e, I5 3f, 4f, I6 3g, 4g, I7
4 4 4 4 4 2 2
H 3-Cl 3-NO2 3-Br 3-OCH3 H 4-CF3
a
Based on benzoyl guanidine.
Scheme 1. Synthetic route of target compounds I1–7. Reagents and conditions: (a) NBS, CCl4, hn, reflux; (b) Na2CO3, acetone or DMF, reflux. (c) (i) Guanidine, isopropyl alcohol, reflux; (ii) HCl.
Compounds I1–7 are the new compounds, and their structures were confirmed by IR, 1H NMR, MS and elemental analysis. The NHE1 inhibitory activities of seven target compounds were evaluated in platelet swelling assay (PSA) and compared to cariporide. The NHE1 inhibitory activities were measured by the decrease of optical density (OD), which induced by platelet swelling [7]. The results showed that all compounds possess NHE1 inhibitory effects. Among these, the inhibitory effects on NHE1 of compound I2, I3, I4, I6 and I7 are superior to the control cariporide (Table 2). Activity of compound I4 is 21 times more potent than cariporide. Ligustrazine showed no effect on NHE1 under this experimental condition, Table 2 IC50 of cariporide, ligustrazine and compounds I1–7 for inhibition of NHE1 Compound
IC50 (10
Cariporide I1 I3 I5 I7
6.50 20.4 0.57 3.28 1.17
8
mol/L)
Compound
IC50 (10
TMP I2 I4 I6
NA 1.47 0.30 0.66
8
mol/L)
M. Ren et al. / Chinese Chemical Letters 18 (2007) 539–541
541
suggesting that its protective effect on myocardial injury during ischemia and reperfusion was not regulated via NHE1 system. The preliminary pharmacological tests showed that p-(3,5,6-trimethylpyrazine-2-ylmethoxy) substituted benzoyl guanidine derivatives exhibit stronger inhibitory effects on NHE1 than o-(3,5,6-trimethylpyrazine-2-ylmethoxy) substituted benzoyl guanidine derivatives, and that introducing electron-withdrawing substituents to benzene ring improve the inhibitory potency of NHE1. The above results gave us useful enlightenment for further design of novel NHE1 inhibitors. In conclusion, a series of novel derivatives of ligustrazine linked with substituted benzoyl guanidine were synthesized. Some of them showed better NHE1 inhibitory activity than cariporide, and the most active one was compound I4 [8]. Structure–activity relationships were briefly discussed. The protective effect of compound I4 on myocardial injury during ischemia and reperfusion in animal models is underway. References [1] [2] [3] [4] [5] [6]
X.C. Cheng, X.Y. Liu, W.F. Xu, Drugs Future 30 (10) (2005) 1059. Z.G. Wu, J. Wuhan Inst. Chem. Technol. 25 (1) (2003) 28. L. Fliegel, H. Wang, J. Mol. Cell. Cardiol. 29 (1997) 1991. B. Masereel, L. Pochet, D. Laeckmann, Eur. J. Med. Chem. 38 (2003) 547. H.M. Piper, C. Balser, Y.V. Ladilov, et al. Basic Res. Cardiol. 91 (1996) 191. Typical procedure for the synthesis of 4a–g: The mixture of compound 2 (10 mmol), compound 3 (10 mmol) and Na2CO3 (5 mmol) in 50 mL acetone was refluxed for 48 h (check by TLC) and filtrated, and the solvent was evaporated in vacuo. Typical procedure for the synthesis of I1–7: A mixture of guanidine (3.5 mmol) and 3 mL isopropyl was stirred at 70 8C, to it was added dropwise a solution of compound 4 (0.35 mmol) dissolved in 2 mL isopropyl. The mixture was refluxed for 30 min (checked by TLC). Adjust the pH to 2 by hydrochloride, filtered, and the filtrate was evaporated in vacuo. The final product was purified by silica gel column chromatography (eluent: dichloromethane: methanol = 20:1–10:1). [7] D. Rosskopf, E. Morgenstern, W. Scholz, et al. Hypertension 9 (3) (1991) 231. [8] Selected spectroscopic data: I4: mp 175–176 8C, yield: 52.1%; IR (KBr, cm 1) 3380, 3189, 2921. 1706 (nC O), 1657 (nC N), 1595, 1551 (nC C), 1265 (nC–O–C); 1H NMR (500 MHz, DMSO-d6, d ppm): 2.50 (s, 6H, –CH3), 2.59 (s, 3H, –CH3), 5.29 (s, 2H, –CH2–O), 7.20 (d, 1H, J = 8.8 Hz, ArH), 7.65 (bs, 2H, NH), 8.06 (dd, 1H, J1 = 2.0 Hz, J2 = 8.8 Hz, ArH), 8.32 (d, 1H, J = 2.0 Hz, ArH); MS(ESI(+)70 V, m/z): 394.0 ([M+H]+, base peak). Anal. Calcd. for C16H18BrN5O2HClH2O: C 43.02, H 4.74, N 15.68, found: C 42.87, H 4.82, N 15.57.