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  • Midazolam oral clearance is used for characterization


    Midazolam oral clearance is used for characterization of both intestinal and hepatic CYP3A activity. However, the identification and subsequent use of metabolic markers of CYP3A activity can eliminate the need for unnecessary administration of probe drugs such as midazolam. Such metabolic markers would be particularly beneficial in vulnerable populations such as pregnant women, children, and the elderly. Among the previously reported metabolic markers for CYP3A activity, plasma 4β-hydroxycholesterol (4β-OH-cholesterol) and ratios of urinary 6β-hydroxycortisol/cortisol (6β-OH-cortisol/cortisol) and 6β-hydroxycortisone/cortisone (6β-OH-cortisone/cortisone) have been reported as reliable metabolic markers of CYP3A activity [13], [14]. In this study, we aimed to determine changes in CYP3A activity following and during pregnancy using metabolic markers.
    Materials and methods
    Discussion 4β-OH-cholesterol is a relatively sensitive marker that can be used in the assessment of CYP3A activity in the induction state, but not in the inhibition state, because of its long apparent GSK2269557 sale ranging from 60 h to 17 days [18]. A previous study has suggested that both 4β-OH-cholesterol and 4β-OH-cholesterol/cholesterol can be used as plasma markers of CYP3A activity when there is no significant increase in plasma cholesterol concentrations [19]. Since the plasma concentration of cholesterol is known to increase following and during pregnancy [17], we quantified both 4β-OH-cholesterol and cholesterol to evaluate CYP3A activity. To this end, we calculated the 4β-OH-cholesterol/cholesterol ratio to evaluate CYP3A activity in pregnant women. We found that the plasma concentrations of both cholesterol and 4β-OH-cholesterol significantly increased as pregnancy progressed (1st trimester < 2nd trimester < 3rd trimester). However, the 4β-OH-cholesterol/cholesterol ratio among the different groups showed that CYP3A activity in pregnant women was higher than that in non-pregnant women, but there was no difference in this ratio between the different pregnancy trimesters. Thus, the 4β-OH-cholesterol/cholesterol ratio helped confirm the increased CYP3A activity in pregnant women compared with that in non-pregnant women. Additionally, this study supports the potential usefulness of 4β-OH-cholesterol/cholesterol rather than 4β-OH-cholesterol to measure CYP3A activity in pregnant women where cholesterol levels increase. Previously, Herbert et al. observed significantly higher midazolam CL/F during pregnancy than during postpartum [9]. However, CYP3A activity during pregnancy, when separated into trimesters, was not investigated before. Our results in this study showed that there is no significant change in CYP3A activity during pregnancy. To support this result, we simulated pharmacokinetics of 1 mg i.v. midazolam following and during pregnancy using the Simcyp simulator pregnancy population module in a generated pregnant women cohort in which the basic demographics were adjusted to the demographics of subjects enrolled in this study (Supplementary Fig. 2). According to the simulated midazolam clearance, a gold standard for CYP3A activity, there seemed to be an increase in CYP3A activity following pregnancy. However, there was no difference of CYP3A activity during pregnancy. The simulated midazolam clearance showed results consistent with those in this study. The urinary markers 6β-OH-cortisol/cortisol and 6β-OH-cortisone/cortisone are known to be metabolized in the liver predominantly by CYP3A4 and excreted through urine [20], [21]. Unlike 4β-OH-cholesterol, these urinary markers have been reported to sensitively reflect CYP3A activity in both the inhibition and the induction states [22]. However, one limitation of using these urinary markers is that the levels of these corticosteroids can show inter- and intra-individual variations in certain circumstances such as kidney malfunction [20]. In this study, when each of the quantified 6β-hydroxylated corticosteroids (6β-OH-cortisol, 6β-OH-cortisone) was analyzed during different pregnancy trimesters, they seemed to indicate a significant increase in CYP3A activity with the progress of pregnancy. However, urinary markers are calculated as the ratio of 6β-hydroxylated metabolite level over parental corticosteroid (cortisol, cortisone) level to avoid inter-individual differences in basal corticosteroid concentrations. Moreover, a previous study has reported renal function-dependent excretion of free cortisol [23]. Since pregnancy dramatically increases renal clearance [24], urinary markers might be significantly affected by it. Furthermore, the spot urine samples used in this study were collected at uncontrolled time points during hospital visits, which may have affected urinary cortisol and cortisone concentrations, which are known to vary throughout the day [25]. Therefore, the use of urinary markers in the evaluation of CYP3A activity in pregnant women may not be the best approach.