Phenotypical and functional characterization of a HepG2 cell clone stably overexpressing cytochrome P450 (CYP) 2C9.

IF 1.7 Q2 MULTIDISCIPLINARY SCIENCES

BMC Research Notes Pub Date : 2026-06-30 DOI:10.1186/s13104-026-07935-y

Sarah Kammerer, Natalie Herzog, Thea Jenchen, Valentina Stock, Rebecca Hofer, Veronika Ruzsanyi, Jan-Heiner Küpper

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引用次数: 0

Abstract

Objective: We aimed to generate a HepG2 cell clone stably overexpressing cytochrome P450 (CYP) 2C9 together with an empty vector (EV) control cell clone for pharmacological and toxicological studies of substances with CYP2C9-mediated biotransformation.

Results: A new HepG2 cell clone was generated by lentiviral transduction to functionally overexpress human CYP2C9. We found high CYP2C9 transcript and protein levels based on qRT-PCR, Western blot and immunofluorescence in the cell clone. Most importantly, specific enzyme activities of 62.9 ± 2.6 pmol 4-hydroxydiclofenac/min/10⁶ cells as analyzed by diclofenac conversion via liquid chromatography-mass spectrometry were found in HepG2-CYP2C9 cells. CYP2C9 enzyme activity could successfully be inhibited by the application of the pan-CYP inhibitor 1-aminobenzotriazole or the CYP2C9-specific inhibitor sulfaphenazole. No changes in morphology or population doubling times were detected when comparing the clone to parental HepG2 cells. The corresponding EV control cell line was detected negative for CYP2C9 expression in all experimental setups and did not show any difference to the parental HepG2 cells. Thus, the newly established HepG2-CYP2C9 cell line is an appropriate tool to study metabolism and toxicity of substances depending on conversion by CYP2C9.

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稳定过表达细胞色素P450 (CYP) 2C9的HepG2细胞克隆的表型和功能特征

目的：建立稳定过表达细胞色素P450 (CYP) 2C9的HepG2细胞克隆和空载体（EV）对照细胞克隆，用于cyp2c9介导的生物转化物质的药理学和毒理学研究。结果：通过慢病毒转导，获得了一个功能过表达人CYP2C9的HepG2细胞克隆。通过qRT-PCR、Western blot和免疫荧光检测，我们发现细胞克隆中CYP2C9转录物和蛋白水平较高。最重要的是，在HepG2-CYP2C9细胞中发现了62.9±2.6 pmol 4-羟基双氯芬酸/min/106细胞（双氯芬酸转化液相色谱-质谱法）的特异酶活性。应用泛cyp抑制剂1-氨基苯并三唑或CYP2C9特异性抑制剂磺胺苯唑可成功抑制CYP2C9酶活性。当克隆与亲本HepG2细胞比较时，未检测到形态学或群体倍增次数的变化。相应的EV对照细胞系在所有实验设置中检测到CYP2C9表达为阴性，与亲本HepG2细胞没有任何差异。因此，新建立的HepG2-CYP2C9细胞系是研究物质依赖于CYP2C9转化的代谢和毒性的合适工具。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

BMC Research Notes Biochemistry, Genetics and Molecular Biology-Biochemistry, Genetics and Molecular Biology (all)

CiteScore

3.60

自引率

0.00%

发文量

363

审稿时长

15 weeks

期刊介绍： BMC Research Notes publishes scientifically valid research outputs that cannot be considered as full research or methodology articles. We support the research community across all scientific and clinical disciplines by providing an open access forum for sharing data and useful information; this includes, but is not limited to, updates to previous work, additions to established methods, short publications, null results, research proposals and data management plans.