Abstract
Cytochrome P450 (CYP) is the most important phase I drug-metabolizing enzyme, and the effect of drugs on CYP enzymes can lead to decreased pharmacological efficacy or enhanced toxicity of drugs, but there are many deficiencies in the evaluation models of CYP enzymes in vitro. Human-induced hepatocytes (hiHeps) derived from human fibroblasts by transdifferentiation have mature hepatocyte characteristics. The aim was to establish a novel evaluation system for the effect of drugs on CYP3A4, 1A2, 2B6, 2C9, and 2C19 in vitro based on hiHeps. Curcumin can inhibit many CYP enzymes in vitro, and so the inhibition of curcumin on CYP enzymes was compared by human liver microsomes, human hepatocytes, and hiHeps using UPLC-MS and the cocktail method. The results showed that the IC50 values of CYP enzymes in the hiHeps group were similar to those in the hepatocytes group, which proved the effectiveness and stability of the novel evaluation system in vitro. Subsequently, the evaluation system was applied to study the inhibitory activity of notoginseng total saponins (NS), safflower total flavonoids (SF), and the herb pair of NS–SF on five CYP enzymes. The mechanism of improving efficacy after NS and SF combined based on CYP enzymes was elucidated in vitro. The established evaluation system will become a powerful tool for the research of the effect of drugs on the activity of CYP enzymes in vitro, which has broad application prospects in drug research.
Highlights
Drug-metabolizing enzymes play a key role in the process of drug metabolism (Underhill and Khetani, 2018), especially drug-drug interaction (DDI) in which drug-metabolizing enzymes have a significant impact on the safety of clinical drugs (Akamine et al, 2019)
This study aims to set up a novel system based on Human-induced hepatocytes (hiHeps) to evaluate the effect of drugs on cytochrome P450 (CYP) enzymes in vitro, and provide new methods and ideas for drug safety/toxicity research in preclinical and clinical stages
The kinetic parameter Michaelis constant (Km) could reflect the catalytic ability of the metabolic enzyme, and provide the necessary reference value of the probe drug for the evaluation system of CYP enzyme activity in vitro (Hakkola et al, 2020; Kong et al, 2020)
Summary
Drug-metabolizing enzymes play a key role in the process of drug metabolism (Underhill and Khetani, 2018), especially drug-drug interaction (DDI) in which drug-metabolizing enzymes have a significant impact on the safety of clinical drugs (Akamine et al, 2019). With the increasing development of new drugs, a rapid, accurate, stable, and low-cost evaluation system for the effect of drugs on CYP enzymes has become important. Liver microsomes are difficult to use when simulating the complete metabolic environment in vivo and evaluating the induction ability of drugs on CYP enzymes, so it is hard to accurately and comprehensively reflect the effect of drugs on the activity of CYP enzymes (Diao and Huestis, 2019; Thiengsusuk et al, 2020). Human hepatocytes are the closest model to human liver tissue, which can be used to evaluate the induction and inhibition effect of drugs on CYP enzymes relatively accurately, but human hepatocytes are difficult to obtain, unable to subculture, expensive, and have a short duration for the activity of metabolic enzymes, which is not conducive to extensive application for evaluation of the effect of drugs on CYP enzymes in vitro (Zeilinger et al, 2016; Schink and Dehus, 2017; Inoue et al, 2020). Some evaluation systems of the effect of drugs on CYP enzymes in vitro were established based on HepG2, HepRG, and other cells (Darnell et al, 2011; Cui et al, 2014), the expression of CYP enzyme activity was not stable in these cells, resulting in poor repeatability and accuracy for evaluation of the effect of drugs on CYP enzymes in vitro
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