[1]许榜田,方大树,谢雨晴,等.苯氧乙酸类化合物对人谷胱甘肽-S-转移酶Mu抑制性能表征[J].陆军军医大学学报(原第三军医大学学报),2022,44(18):1826-1834.
 XU Bangtian,FANG Dashu,XIE Yuqing,et al.Characterization of phenoxyacetic acid compounds in inhibition of human glutathione-S-transferase Mu [J].J Amry Med Univ (J Third Mil Med Univ),2022,44(18):1826-1834.
点击复制

苯氧乙酸类化合物对人谷胱甘肽-S-转移酶Mu抑制性能表征(/HTML )
分享到:

陆军军医大学学报(原第三军医大学学报)[ISSN:1000-5404/CN:51-1095/R]

卷:
44卷
期数:
2022年第18期
页码:
1826-1834
栏目:
基础医学
出版日期:
2022-09-30

文章信息/Info

Title:
Characterization of phenoxyacetic acid compounds in inhibition of human glutathione-S-transferase Mu 
 
作者:
许榜田方大树谢雨晴杨晓兰
重庆医科大学附属大学城医院药学部;重庆医科大学检验医学院,临床检验诊断学教育部重点实验室
 
Author(s):
XU Bangtian1 FANG Dashu1 XIE Yuqing1 YANG Xiaolan2

1Department of Pharmaceutics, University-Town Hospital of Chongqing Medical University, Chongqing, 401331; 2Key Laboratory of Clinical Laboratory and Diagnostics of Ministry of Education, College of Laboratory Medicine, Chongqing Medical University, Chongqing, 400016, China

关键词:
苯氧乙酸类化合物谷胱甘肽-S-转移酶Mu抑制性能表征
Keywords:
 
分类号:
R345; R914.4; R962
文献标志码:
A
摘要:

目的设计合成4个苯氧乙酸类化合物,通过不同方法表征其对人谷胱甘肽-S-转移酶Mu(glutathione-S-transferase Mu, GSTM)的抑制性能,考察苯氧乙酸类化合物与GSTM之间的构效关系。方法设计合成含α,β-不饱和酮结构的4-丙烯酰苯氧乙酰乙胺(Ⅰa)、N,N’-(4-(丙烯酰基)苯氧乙酰基)丁二胺(Ⅱa)和不含α,β-不饱和酮结构的4-丙酰苯氧乙酰乙胺(Ⅰb)、N,N’-(4-(丙酰基)苯氧乙酰基)丁二胺(Ⅱb),利用初速度法表征4个化合物对GSTM的半抑制浓度(50% inhibiting concentration, IC50)和表观抑制常数(inhibition constant, Ki),利用荧光光谱法表征4个化合物对GSTM的解离常数(dissociation constant, Kd)。结果含α,β-不饱和酮结构的Ⅰa、Ⅱa可与谷胱甘肽(glutathione, GSH)发生亲电加成反应,与酶和GSH孵育原位生成产物后,其对GSTM的IC50分别为 (67±6) μmol/L、(0.10±0.02) μmol/L (n=2);不含α,β-不饱和酮结构的Ⅰb、Ⅱb对GSTM无明显抑制作用。Ⅰa相对GSH的Ki为(19.9±1.6) μmol/L,相对CDNB的Ki为(9.1±0.7) μmol/L (n=3);Ⅱa相对GSH的Ki为(0.063±0.005) μmol/L,相对CDNB的Ki为(0.079±0.006) μmol/L (n=3);可见Ⅱa较Ⅰa的Ki低(P<0.05)。荧光光谱法测得Ⅰa、Ⅰb、Ⅱa、Ⅱb的Kd分别为(41.5±1.8) μmol/L、(38.9±1.7) μmol/L、(18.0±0.9) μmol/L、(19.5±1.0) μmol (n=2);可见Ⅰa与Ⅰb、Ⅱa与Ⅱb之间的Kd无显著差异(P>0.05),而Ⅱa较Ⅰa、Ⅱb较Ⅰb的Kd低(P<0.05)。结论α,β-不饱和酮结构是苯氧乙酸类化合物抑制作用的必须官能团;二价抑制剂Ⅱa、Ⅱb较单价抑制剂Ⅰa、Ⅰb对GSTM具有更高亲和力,Ⅱa的产物较Ⅰa的产物抑制作用更强。

Abstract:

ObjectiveTo design and synthesize 4 phenoxyacetic acid compounds, characterize their inhibitory potencies on human glutathione-S-transferase Mu (glutathione-S-transferase Mu, GSTM) by different methods, and investigate the structure-activity relationship between phenoxyacetic acid compounds and GSTM. MethodsFour phenoxyacetic acid compounds were designed and synthesized, and they are ethyl 2-(4-acryloylphenoxy)acetate (Ⅰa) and N,N’-(butane-1,4-diyl)-bis-(4-acryloyl phenoxyacetic amide) (Ⅱa) containing α,β-unsaturated ketone structure, and ethyl 2-(4-propionylphenoxy)acetate (Ⅰb), N,N’-(butane-1,4-diyl)-bis-(4-propionyl phenoxyacetic amide) (Ⅱb) not containing α,β-unsaturated ketone structure. They were characterized by the half-maximal inhibitory concentration (IC50) and apparent inhibition constant (Ki) using initial velocity method, and the dissociation constants (Kd) for GSTM utilizing fluorescence analysis. ResultsⅠa and Ⅱa contained α,β-unsaturated ketone structure that undergoes electrophilic addition reaction with GSH. After incubation with enzyme and GSH, Ⅰa and Ⅱa generated products in situ, with a IC50 value of 67.00±6.00 and 0.10±0.02 μmol/L for GSTM respectively (n=2). Ⅰb and Ⅱb without α, β -unsaturated ketone structures had no inhibitory potencies for GSTM. The Ki of Ⅰa against GSH was 19.9±1.6 μmol/L, and the Ki against CDNB was 9.1±0.7 μmol/L (n=3). The Ki of Ⅱa against GSH was 0.063±0.005 μmol/L, and against CDNB was 0.079±0.006 μmol/L (n=3). The Ki of Ⅱa was lower than that ofⅠa (P<0.05). The Kd of Ⅰa, Ⅰb, Ⅱa, and Ⅱb measured by fluorescence analysis were 41.5±1.8, 38.9±1.7, 18.0±0.9, and 19.5±1.0 μmol/L, respectively (n=2). The Kd  between Ⅰa and Ⅰb, and between Ⅱa and Ⅱb had no significant differences(P>0.05). The Kd of Ⅱa was lower than  that of Ⅰa ,and the Kd of Ⅱb was lower than that of Ⅰb (P<0.05). Conclusionα,β-unsaturated ketone structure is the essential functional group of phenoxyacetic acid compounds for inhibition on GSTM. Comparing with monovalent Ⅰa and Ⅰb, divalent Ⅱa and Ⅱb have higher affinity for GSTM. The product of Ⅱa has inhibitory potency significantly higher than that of Ⅰa.

参考文献/References:

[1]AYBEK H, TEMEL Y, AHMEDB M, et al. Deciphering of the effect of chemotherapeutic agents on human glutathione S-transferase enzyme and MCF-7 cell line[J]. Protein Pept Lett, 2020, 27(9): 888-894. DOI: 10.2174/0929866527 666200413101017.
 
[2]刘芳, 宋玉芳, 张学辉, 等. 多药耐药基因表达与上皮性卵巢癌化疗耐药的相关性[J]. 中国医刊, 2021, 56(5): 561-564. DOI: 10.3969/j.issn.1008-1070.2021.05.028. 
 
LIU F, SONG Y F, ZHANG X H, et al. Correlation between multidrug resistance gene expression and chemoresistance in epithelial ovarian cancer [J]. Chin J Med, 2021, 56(5): 561-564. DOI: 10.3969/j.issn.1008-1070.2021.05.028.
 
[3]WANG C H, WU H T, CHENG H M,et al. Inhibition of glutathione S-transferase M1 by new gabosine analogues is essential for overcoming cisplatin resistance in lung cancer cells[J]. J Med Chem, 2011, 54(24): 8574-8581. DOI: 10.1021/jm201131n.
 
[4]CARLSTEN C, SAGOO G S, FRODSHAM A J,et al. Glutathione S-transferase M1 (GSTM1) polymorphisms and lung cancer: a literature-based systematic HuGE review and meta-analysis[J]. Am J Epidemiol, 2008, 167(7): 759-774. DOI: 10.1093/aje/kwm383.
 
[5]DI PAOLO V, FULCI C, ROTILI D,et al. Characterization of water-soluble esters of nitrobenzoxadiazole-based GSTP1-1 inhibitors for cancer treatment[J]. Biochem Pharmacol, 2020, 178: 114060. DOI: 10.1016/j.bcp.2020.114060.
 
[6]SINGH R R, REINDL K M. Glutathione S-transferases in cancer[J]. Antioxidants (Basel), 2021, 10(5): 701. DOI: 10.3390/antiox10050701.
 
[7]IERSEL M L, PLOEMEN J P, STRUIK I,et al. Inhibition of glutathione S-transferase activity in human melanoma cells by alpha, beta-unsaturated carbonyl derivatives. Effects of acrolein, cinnamaldehyde, citral, crotonaldehyde, curcumin, ethacrynic acid, and trans-2-hexenal[J]. Chem Biol Interact, 1996, 102(2): 117-132. DOI: 10.1016/s0009-2797(96)03739-8.
 
[8]XIA C, LU J P, XU B T,et al. Design and characterization of a labeling reagent for covalent immobilization of glutathione-S-transferase[J]. Nanosci Nanotechnol Lett, 2019, 11(11): 1547-1560. DOI: 10.1166/nnl.2019.3044.
 
[9]AYNA A, KHOSNAW L, TEMEL Y,et al. Antibiotics as inhibitor of glutathione S-transferase: biological evaluation and molecular structure studies[J]. Curr Drug Metab, 2021, 22(4): 308-314. DOI: 10.2174/1389200222666210118102700.
 
[10]HANSSON J, BERHANE K, CASTRO V M,et al. Sensitization of human melanoma cells to the cytotoxic effect of melphalan by the glutathione transferase inhibitor ethacrynic acid[J]. Cancer Res, 1991, 51(1): 94-98.
 
[11]黄玲萍, 谢丽霞, 邱钰超, 等. 依他尼酸联合顺铂化疗促肺癌A549细胞凋亡的作用及机制研究[J]. 第三军医大学学报, 2017, 39(17): 1720-1727. DOI: 10.16016/j.1000-5404.201701068. 
 
HUANG L P, XIE L X, QIU Y C, et al. Ethacrynic acid promotes apoptosis in lung cancer A549 cells when combined with cisplatin chemotherapy[J]. J Third Mil Med Univ, 2017, 39(17): 1720-1727. DOI: 10.16016/j.1000-5404.201701068.
 
[12]PLOEMEN J H, VAN OMMEN B, VAN BLADEREN P J. Inhibition of rat and human glutathione S-transferase isoenzymes by ethacrynic acid and its glutathione conjugate[J]. Biochem Pharmacol, 1990, 40(7): 1631-1635. DOI: 10.1016/0006-2952(90)90465-w.
 
[13]TARS K, OLIN B, MANNERVIK B. Structural basis for featuring of steroid isomerase activity in alpha class glutathione transferases[J]. J Mol Biol, 2010, 397(1): 332-340. DOI: 10.1016/j.jmb.2010.01.023.
 
[14]杨宪峰, 张灵, 杨晓兰, 等. 融合标签谷胱甘肽S-转硫酶高亲和配体的设计与筛选[J]. 重庆医科大学学报, 2012, 37(9): 791-795. DOI: 10.3969/j.issn.0253-3626.2012.09.011. 
 
YANG X F, ZHANG L, YANG X L, et al. Design and screening of high-affinity ligands for glutathione S-transferase as fusion tag[J]. J Chongqing Med Univ, 2012, 37(9): 791-795. DOI: 10.3969/j.issn.0253-3626.2012.09.011.
 
[15]许榜田, 赵语, 龙友琦, 等. 两种依他尼酸衍生物对人类谷胱甘肽-S-转移酶Mu的抑制动力学研究[J]. 重庆医科大学学报, 2020, 45(8): 1144-1150. DOI: 10.13406/j.cnki.cyxb.002211. 
 
XU B T, ZHAO Y, LONG Y Q, et al. A study of the inhibition kinetics of human glutathione-S-transferase Mu by two ethacrynic acid derivatives[J]. J Chongqing Med Univ, 2020, 45(8): 1144-1150. DOI: 10.13406/j.cnki.cyxb.002211.
 
[16]同婷婷, 段昌园, 李欣蓬, 等. 双依他尼酸乙醇胺对谷胱甘肽S-转硫酶mu的选择性抑制作用[J]. 基因组学与应用生物学, 2021, 40(1): 406-413. DOI: 10.13417/j.gab.040.000406. 
 
TONG T T, DUAN C Y, LI X P, et al. Characterization of aminoethanol di-ethacrynic acid as a selective divalent potent inhibitor of glutathione S-transferase mu[J]. Genom Appl Biol, 2021, 40(1): 406-413. DOI: 10.13417/j.gab.040.000406.
 
[17]刘芳, 杨晓兰, 廖飞. GSTM潜抑制剂双依他尼酰乙二胺对人耐顺铂卵巢癌细胞COC1/DDP的抗肿瘤活性[J]. 基因组学与应用生物学, 2016, 35(6): 1266-1271. DOI: 10.13417/j.gab.035.001266.
 
LIU F, YANG X L, LIAO F. Anti-tumor activity of short divalent ethacrynic amide on human ovarian cancer cell line COC1/DDP[J]. Genom Appl Biol, 2016, 35(6): 1266-1271. DOI: 10.13417/j.gab.035.001266.
 
[18]HOU J F, LIU F X, WU N,et al. Efficient biodegradation of chlorophenols in aqueous phase by magnetically immobilized aniline-degrading Rhodococcus rhodochrous strain[J]. J Nanobiotechnology, 2016, 14: 5. DOI: 10.1186/s12951-016-0158-0.
 
[19]BRADFORD M M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding[J]. Anal Biochem, 1976, 72: 248-254. DOI: 10.1006/abio.1976.9999.
 
[20]HABIG W H, PABST M J, JAKOBY W B. Glutathione S-transferases. The first enzymatic step in mercapturic acid formation[J]. J Biol Chem, 1974, 249(22): 7130-7139.
 
[21]LIAO F, LI J C, KANG G F, et al. Measurement of mouse liver glutathione S-transferase activity by the integrated method[J]. J Med Coll PLA, 2003, 18(5): 295-300.
 
[22]YANG X L, DU Z Y, PU J, et al. Classification of difference between inhibition constants of an inhibitor to facilitate identifying the inhibition type[J]. J Enzyme Inhib Med Chem, 2013, 28(1): 205-213. DOI: 10.3109/147563 66.2011.645240.
 
[23]许金钩, 王尊本. 荧光分析法[M].第3版. 北京: 科学出版社, 2006: 68-69.
 
XU J G, WANG Z B. Fluorescence analysis[M]. 3rd ed. Beijing: Science Press, 2006: 68-69.
 
[24]XU B T, TONG T T, WANG X,et al. Short divalent ethacrynic amides as pro-inhibitors of glutathione S-transferase isozyme Mu and potent sensitisers of cisplatin-resistant ovarian cancers[J]. J Enzyme Inhib Med Chem, 2022, 37(1): 728-742. DOI: 10.1080/14756366.2022.2038591.
 

更新日期/Last Update: 2022-09-23