In Vitro Effects of Some Chemotherapeutic Drugs on Rat Erythrocytes Glutathione S-transferase (GST) Enzyme

Year 2024, Volume: 13 Issue: 4, 17 - 22, 30.12.2024

Barzan Ahmed , Yusuf Temel , Mehmet Çiftci

https://doi.org/10.46810/tdfd.1454573

Abstract

After heart disease, cancer is the leading cause of death worldwide. Currently, breast, lung, bowel, and prostate cancer are the most common cancers in the worldwide. By stopping cancer cells from dividing, spreading, growing, making more cells, and then destroying them, chemotherapy drugs are used to treat diseases caused by cancer. The glutathione S-transferase enzyme is responsible for the detoxification of xenobiotic molecules produced by the body during cancer treatment. In this study, glutathione S-transferase enzyme (GST, EC: 2.5.1.18) was extracted from the erythrocytes of rats by affinity column chromatography in one step. The gel electrophoresis (SDS-PAGE) was used to verify the GST enzyme's purity, A single protein band was obtained. The GST enzyme was purified with 22.5 EU/mg specific activity, 237.14 purification-fold, and 48.98% purification yield. The subsequently investigation study into the in vitro effects of 5-fluorouracil (5-FU) and cyclophosphamide (CP) chemotherapy drugs on purified GST enzyme activity revealed that both 5-fluorouracil and cyclophosphamide increased GST activity at a concentration range of (0.385 to 15.4 mM, and 19.15 to 191.5 mM) respectively.

Keywords

Rat erythrocyte, Glutathione S-transferase, Purification, Chemotherapeutic Drugs, Enzyme activity.

References

• Rasul MF, Hussen BM, Salihi A, Ismael BS, Jalal PJ, Zanichelli A, Jamali E, Baniahmad A, Ghafouri-Fard S, Basiri A, Taheri M. Strategies to overcome the main challenges of the use of CRISPR/Cas9 as a replacement for cancer therapy. Mol Cancer. 2022; 21(1) 64: 2-30.
• Focaccetti C, Bruno A, Magnani E, Bartolini D, Principi E, Dallaglio K, Bucci EO, Finzi G, Sessa F., Noonan DM, Albini A. Effects of 5-fluorouracil on morphology, cell cycle, proliferation, apoptosis, autophagy and ROS production in endothelial cells and cardiomyocytes. PloS one. 2015; 10(2): 0115686.
• Jahani M, Azadbakht M, Norooznezhad F, Mansouri K. L-arginine alters the effect of 5-fluorouracil on breast cancer cells in favor of apoptosis. Biomed Pharmacother. 2017; 88: 114-123.   Ince S, Kucukkurt I, Demirel HH, Acaroz DA, Akbel E, Cigerci IH, Protective effects of boron on cyclophosphamide induced lipid peroxidation and genotoxicity in rats. Chemosphere. 2014; 108:197-204.
• Temel Y, Çağlayan C, Ahmed BM, Kandemir FM, Çiftci M., The effects of chrysin and naringin on cyclophosphamide-induced erythrocyte damage in rats: biochemical evaluation of some enzyme activities in vivo and in vitro. Naunyn Schmiedebergs Arch. 2021; 394: 645-654.
• Orhan H, Şahin G, Glutatyon S-Transferazların klinik ve toksikolojik önemi. Türkiye Klinikleri Tıp Bilimleri. 1995; 15: 303-15.
• Ballatori N, Krance SM, Notenboom S, Shi S, Tieu K, Hammond CL. Glutathione dysregulation and the etiology and progression of human diseases. Biol. Chem. 2009; 390: 191-214.
• Kaur G, Gupta SK, Singh P, Ali V, Kumar V, Verma M. Drug-metabolizing enzymes: role in drug resistance in cancer. Clin Transl Oncol. 2020; 22: 1667-1680.
• Lu J, Holmgren A, The thioredoxin antioxidant system. Free Radic Biol Med. 2014; 66: 75-87.
• Singh RR, Reindl KM. Glutathione S-transferases in cancer. Antioxid. 2021; 10(5): 701-710.
• Temel Y, Taysi MŞ, The effect of mercury chloride and boric acid on rat erythrocyte enzymes. Biol. Trace Elem. Res. 2019; 191(1): 177-182.
• Aybek H, Temel Y, Ahmed BM, Ağca CA, Çiftci M. Deciphering of the effect of chemotherapeutic agents on human glutathione S-transferase enzyme and MCF-7 cell line. Protein Peptide Lett. 2020; 27(9): 888-894.
• Taysi MŞ, Temel Y. Glutathione S-transferase: Purification and characterization from quail (Coturnix coturnix japonica) liver and the impact of some metal ions on enzyme activity. Bionanosci. 2021; 11: 91-98.
• Ayna A, Khosnaw L, Temel Y, Ciftci M. Antibiotics as inhibitor of glutathione S-transferase: biological evaluation and molecular structure studies. Curr. Drug Metab. 2021; 22(4): 308-314.
• Temel Y, Kocyigit UM. Purification of glucose‐6‐phosphate dehydrogenase from rat (Rattus norvegicus) erythrocytes and inhibition effects of some metal ions on enzyme activity. J Biochem Mol Toxicol. 2017; 31(9): e21927.
• Habig WH, Pabst MJ, Jakoby WB. Glutathione S-transferases: the first enzymatic step in mercapturic acid formation. J. Biol. Chem. 1974; 249(22): 7130-7139.
• Türkan F, Huyut Z, Taslimi P, Gülçin, İ. Investigation of the effects of cephalosporin antibiotics on glutathione S-transferase activity in different tissues of rats in vivo conditions in order to drug development research. Drug Chem. Toxicol. 2020; 43(4): 423-428. Temel Y, Koçyigit UM, Taysı MŞ, Gökalp F, Gürdere MB, Budak Y, Ceylan M, Gülçin İ, Çiftci, M. Purification of glutathione S‐transferase enzyme from quail liver tissue and inhibition effects of (3aR, 4S, 7R, 7aS)‐2‐(4‐((E)‐3‐(aryl) acryloyl) phenyl)‐3a, 4, 7, 7a‐tetrahydro‐1H‐4, 7‐methanoisoindole‐1, 3 (2H)‐dione derivatives on the enzyme activity. Biochem Mol Toxicol. 2018; 32(3); e22034.
• Aksoy M, Ozaslan MS, Kufrevioglu OI, Purification of glutathione S-transferase from Van Lake fish (Chalcalburnus tarichii Pallas) muscle and investigation of some metal ions effect on enzyme activity. J. Enzyme Inhib. Med. Chem. 2016; 31(4): 546-550.
• Kruger NJ. The Bradford method for protein quantitation. The protein protocols handbook. Springer; 2009. P. 17-24.
• Bradford MM. A rapid and sensitive method for the quantition of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976; 72 (1–2): 248–251.
• Laemmli UK. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970; 227(5259): 680-685.
• Townsend DM, Tew KD. The role of glutathione-S-transferase in anti-cancer drug resistance. Oncogene. 2003; 22(47);7369-7375.
• Batt AM, Magdalou J, Vincent-Viry M, Ouzzine M, Fournel-Gigleux S, Galteau MM, Siest G. Drug metabolizing enzymes related to laboratory medicine: Cytochromes P-450 and UDP-glcuronosyltransferases. Clin Chim Acta. 1994;226(2): 171-190.
• Temel Y, Ayna A, Hamdi Shafeeq I, Ciftci M. In vitro effects of some antibiotics on glucose-6-phosphate dehydrogenase from rat (Rattus norvegicus) erythrocyte. Drug Chem. Toxicol. 2020; 43(2): 219-223.
• Bayindir S, Ayna A, Temel Y, Ciftci M. The synthesis of new oxindoles as analogs of natural product 3, 3′-bis (indolyl) oxindole and in vitro evaluation of the enzyme activity of G6PD and 6PGD. Turk. J. Chem. 2018; 42(2): 332-345.
• Bayindir S, Temel Y, Ayna A, Ciftci M. The synthesis of N‐benzoylindoles as inhibitors of rat erythrocyte glucose‐6‐phosphate dehydrogenase and 6‐phosphogluconate dehydrogenase. J Biochem Mol Toxicol. 2018; 32(9): e22193.
• Pljesa-Ercegovac, M., Savic-Radojevic, A., Matic, M., Coric, V., Djukic, T., Radic, T. and Simic, T. Glutathione transferases: potential targets to overcome chemoresistance in solid tumors. International J Molecular Sci. 2018; 19(12): 3785.
• Dierickx PJ. Purification and characterization of glutathione S-transferase from the human hepatoma derived PLC/PRF/5 cell line. Biomed res. 1989; 10(4); 301-306.
• Gadagbui, B.K. and James, M.O. Activities of affinity-isolated glutathione S-transferase (GST) from channel catfish whole intestine. Aquat Toxicol. 2000; 49(1-2): 27-37.
• Riol MM, Valinas MN, Fernandez MG, Lopez MP. Glutathione S-transferases from rainbow trout liver and freshly isolated hepatocytes: purification and characterization. Comp Biochem Physiol C Toxicol Pharmacol. 2001; 128(2): 227-235.
• Huang Q, Liang L, Wei T, Zhang D, Zeng QY. Purification and partial characterization of glutathione transferase from the teleost Monopterus albus. Comp Biochem Physiol C Toxicol Pharmacol. 2008; 147(1): 96-100.
• Hamed RR, Maharem TM, Abdel-Meguid N, Sabry GM, Abdalla AM, Guneidy RA. Purification and biochemical characterization of glutathione S-transferase from Down syndrome and normal children erythrocytes: A comparative study. Res. Dev. Disabil. 2011; 32(5): 1470-1482.
• Lebda M, Taha N, Noeman S, Korshom M, El-Wahab Mandour A. Purification and Characterization of Glutathione-S-Transferase from Rat′ s Liver: Effect of Carbon Tetrachloride and Camel′ s Milk. J Chromat. Separation Techniq. 2012; 3(4): 2-8.
• Howie AF, Hayes JD, Beckett GJ. Purification of acidic glutathione S-transferases from human lung, placenta and erythrocyte and the development of a specific radioimmunoassay for their measurement. Clinica chimica acta. 1988; 177(1): 65-75.
• Akkemik E, Taser P, Bayindir A, Budak H, Ciftci M. Purification and characterization of glutathione S-transferase from turkey liver and inhibition effects of some metal ions on enzyme activity. Environ Toxicol Pharmacol. 2012; 34(3): 888-894.
• Senjo M, Ishibashi T. Purification and characterization of glutathione S-transferase from rat brain cytosol: identification of four isozymes and evidence for absence of the Ya subunit. Biomed Res. 1986; 7(1): 19-26.
• Temel Y. The in vitro effect of 5-FU and Tamoxifen Chemotherapeutics on penthose phosphate pathway enzymes. Cumhuriyet Sci J. 2021; 42(2): 245-251.
• Erat M, Şakiroğlu H. The effect of some antineoplastic agents on glutathione S-transferase from human erythrocytes. J Enzyme Inhib Med Chem. 2013; 28(4): 711-716.