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In vitro enzyme kinetics and NMR-based product elucidation for glutathione S-conjugation of the anticancer unsymmetrical bisacridine C-2028 in liver microsomes and cytosol: major role of glutathione S-transferase M1-1 isoenzyme
Abstract
This work is the next step in studying the interplay between C-2028 (anticancer-active unsymmetrical bisacridine developed in our group) and the glutathione S-transferase/glutathione (GST/GSH) system. Here, we analyzed the concentration- and pH-dependent GSH conjugation of C-2028 in rat liver microsomes and cytosol. We also applied three recombinant human GST isoenzymes, which altered expression was found in various tumors. The formation of GSH S-conjugate of C-2028 in liver subfractions followed Michaelis-Menten kinetics. We found that C-2028 was conjugated with GSH preferentially by GSTM1-1, revealing a sigmoidal kinetic model. Using a colorimetric assay (MTT test), we initially assessed the cellular GST/GSH-dependent biotransformation of C-2028 in relation to cytotoxicity against Du-145 human prostate cancer cells in the presence or absence of the modulator of GSH biosynthesis. Pretreatment of cells with buthionine sulfoximine resulted in a cytotoxicity decrease, suggesting a possible GSH-mediated bioactivation process. Altogether, our results confirmed the importance of GSH conjugation in C-2028 metabolism, which humans must consider when planning a treatment strategy. Finally, nuclear magnetic resonance spectroscopy elucidated the structure of the GSH-derived product of C-2028. Hence, synthesizing the compound standard necessary for further advanced biological and bioanalytical investigations will be achievable.
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Authors (8)
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Aleksandra Pawłowicz
- Instytut Chemii Bioorganicznej Polskiej Akademii Nauk w Poznaniu
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Witold Andrałojć dr
- Instytut Chemii Bioorganicznej Polskiej Akademii Nauk w Poznaniu
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- Accepted or Published Version
- DOI:
- Digital Object Identifier (open in new tab) 10.3390/molecules28196812
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- Category:
- Articles
- Type:
- artykuły w czasopismach
- Published in:
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MOLECULES
no. 28,
ISSN: 1420-3049 - Language:
- English
- Publication year:
- 2023
- Bibliographic description:
- Potęga A., Rafalska D., Kazimierczyk D., Kosno M., Pawłowicz A., Andrałojć W., Paluszkiewicz E., Laskowski T.: In vitro enzyme kinetics and NMR-based product elucidation for glutathione S-conjugation of the anticancer unsymmetrical bisacridine C-2028 in liver microsomes and cytosol: major role of glutathione S-transferase M1-1 isoenzyme// MOLECULES -Vol. 28,iss. 19 (2023), s.6812-
- DOI:
- Digital Object Identifier (open in new tab) 10.3390/molecules28196812
- Sources of funding:
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- COST_FREE
- Verified by:
- Gdańsk University of Technology
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