Clinical significance of personalized tacrolimus dosing by adjusting to donor CYP3A-status in liver transplant recipients.

Donor's CYP3A-status (CYP3A5 genotype and CYP3A4 expression) can provide prognostic information regarding tacrolimus-metabolizing capacity of the liver graft and initial tacrolimus dosing for therapeutic blood concentrations in liver transplants. The present work prospectively investigated whether CYP3A-status guided tacrolimus therapy has any potential clinical benefit for recipients in the early postoperative period. METHODS: The contribution of preliminary assaying of donor CYP3A-status to the optimization of initial tacrolimus therapy and to the reduction of adverse events (acute rejection, infection, nephrotoxicity) was investigated in 112 liver transplant recipients (CYPtest group) comparing to 101 control patients on tacrolimus concentration guided therapy. RESULTS: The time for achieving therapeutic tacrolimus concentration was significantly reduced, confirming potential benefit of initial tacrolimus therapy adjusted to donor's CYP3A-status over classical clinical practice of tacrolimus concentration guided treatment (4 vs 8 days, P < 0.0001). Acute rejection episodes (3.6 vs 23.8%, P < 0.0001) and tacrolimus induced nephrotoxicity (8 vs 27%, P = 0.0004) were less frequent in CYPtest group than in control patients, whereas occurrence of infectious disease was not influenced by tacrolimus dosing strategy (3.6 vs 5.9% in CYPtest and control groups, P > 0.05). Acute rejection was often accompanied with tacrolimus blood concentrations lower than 10 ng mL-1 (20/24 of control and 2/4 of CYPtest patients), while nephrotoxicity was associated with high tacrolimus concentrations (>20 ng mL-1 ) in the first week after transplantation (13/27 of control and 2/9 of CYPtest patients). CONCLUSION: CYP3A-status guided therapy significantly improved the risk of misdosing induced early adverse effects (acute rejection, nephrotoxicity).

Adverse events in a newborn on valproate therapy due to loss-of-function mutations in CYP2C9.

An increased risk of valproate-induced toxicity has been reported in children, particularly in those younger than 2 years of age. Significant variations in valproate pharmacokinetics and shifts in the metabolic pathways towards CYP2C9-dependent metabolism seem to play some role in the age-related differences in the incidence of adverse events. We present the case of a premature patient with moderate hemorrhage in the subependymal region (grade II - intraventricular hemorrhage without ventricular dilatation), several myoclonic episodes in her right upper arm (series of jerks lasting milliseconds), and epileptiform abnormalities on the EEG (localized spike-and-wave in the left frontal region with preserved background activity who was treated with valproate. Serious side effects, consisting of bone marrow depression, hyperammonemia, and serum alkaline phosphatase elevation, were observed seventeen days after the beginning of valproate therapy. The toxic symptoms were likely the consequence of a reduced ability to metabolize valproate. The patient was demonstrated to carry two loss-of-function mutations in CYP2C9 (CYP2C9*3/*3) resulting in exaggerated blood concentrations of valproate. The present case highlights the importance of assaying inborn errors in CYP2C9 gene in pediatric patients to avoid valproate-evoked serious side effects.

Clinical significance of CYP2C9-status guided valproic acid therapy in children.

OBJECTIVES: Valproic acid (VPA)-induced adverse effects, which are sometimes serious in children, can be associated with alterations in VPA metabolism. VPA-evoked toxicity is attributed to both the parent compound and its unsaturated metabolites, primarily formed by the cytochrome P450 (CYP)2C9 enzyme. Thus, patients' CYP2C9-status may account for the predisposition to adverse reactions, and testing CYP2C9-status may contribute to the improvement and rationalization of VPA therapy in children. METHODS: In the CYPtest group, children's CYP2C9-status was screened before initiating antiepileptic therapy. CYP2C9-status was estimated by the identification of defective CYP2C9 allelic variants (CYP2C9*2, CYP2C9*3) and current CYP2C9 expression in patients' leukocytes, which reflects hepatic CYP2C9 activities. When the results of CYP2C9 genotyping and CYP2C9 expression were combined, the patients' VPA-metabolizing capacity was predicted, and VPA dosing was adjusted to the patients' CYP2C9-status. Clinical and biochemical parameters, such as VPA serum levels, blood cell counts, liver function parameters, and adverse effects in patients of CYPtest group were compared with those of the control group treated with VPA according to conventional clinical practice. RESULTS: CYP2C9-guided treatment significantly reduced VPA misdosing and consequently decreased the ratio of patients out of the range of target VPA blood concentrations. In the CYPtest group of children who received CYP2C9-status adapted dose, serum alkaline phosphatase (ALP) level and the ratio of patients with abnormal ALP levels were substantially lower than in the control group. The incidence of serious side effects, notably hyperammonemia, was reduced in the CYPtest group; however, some other side effects, such as weight changes and somnolence, could not be avoided. SIGNIFICANCE: The knowledge of pediatric patients' CYP2C9-status can contribute to the optimization of VPA dosing and to the avoidance of misdosing-induced side effects.

Phenoconversion of CYP2C9 in epilepsy limits the predictive value of CYP2C9 genotype in optimizing valproate therapy.

AIM: Since prominent role in valproate metabolism is assigned to CYP2C9 in pediatric patients, the association between children's CYP2C9-status and serum valproate concentrations or dose-requirements was evaluated. MATERIALS & METHODS: The contribution of CYP2C9 genotype and CYP2C9 expression in children (n = 50, Caucasian) with epilepsy to valproate pharmacokinetics was analyzed. RESULTS: Valproate concentrations were significantly lower in normal expressers with CYP2C9*1/*1 than in low expressers or in patients carrying polymorphic CYP2C9 alleles. Consistently, the dose-requirement was substantially higher in normal expressers carrying CYP2C9*1/*1 (33.3 mg/kg vs 13.8-17.8 mg/kg, p < 0.0001). Low CYP2C9 expression significantly increased the ratio of poor metabolizers predictable from CYP2C9 genotype (by 46%). CONCLUSION: Due to the substantial downregulation of CYP2C9 expression in epilepsy, inferring patients' valproate metabolizing phenotype merely from CYP2C9 genotype results in false prediction.

Dehydroepiandrosterone post-transcriptionally modifies CYP1A2 induction involving androgen receptor.

The pharmacological dosage of dehydroepiandrosterone (DHEA) protects against chemically induced carcinogenesis. The chemoprotective activity of DHEA is attributed to its inhibitory potential for the expression of CYP1A enzymes, which are highly responsible for metabolic activation of several mutagenic and carcinogenic chemicals. The present work investigated whether the chemoprevention by DHEA was due to diminished transcriptional activation of CYP1A genes or to the post-transcriptional modulation of CYP1A expression. In primary human hepatocytes, DHEA diminished the increase in CYP1A activities (7-ethoxyresorufin O-dealkylation and phenacetin O-dealkylation) and in CYP1A2 mRNA level induced by 3-methylcholanthrene, but did not alter the amount of CYP1A1 and CYP1B1 mRNA. The androgen receptor seemed to be involved in DHEA-mediated diminishment of CYP1A2 induction, which was attenuated in the presence of bicalutamide, the androgen receptor antagonist. The potential role of the glucocorticoid receptor and estrogen receptor in DHEA-mediated decrease in CYP1A2 induction was excluded. The developed computational model of CYP1A2 induction kinetics and CYP1A2 mRNA degradation proposed that a post-transcriptional mechanism was likely to be the primary mechanism of the DHEA-mediated diminishment of CYP1A2 induction. The hypothesis was confirmed by the results of actinomycin D-chase experiments in MCF-7 and LNCaP cells, displaying that the degradation rates of CYP1A2 mRNA were significantly higher in the cells exposed to DHEA. The novel findings on DHEA-mediated modulation of CYP1A2 mRNA stability may account for the beneficial effects of DHEA by decreasing the metabolic activation of pro-carcinogenic compounds.

Estimation of drug-metabolizing capacity by cytochrome P450 genotyping and expression.

Many undesired side effects or therapeutic failures of drugs are the result of differences or changes in drug metabolism, primarily depending on the levels and activities of cytochrome P450 (P450) enzymes. To assess whether P450 expression profiles can reflect hepatic drug metabolism, we compared P450 mRNA levels in the liver or peripheral leukocytes with the corresponding hepatic P450 activities. A preliminary P450 genotyping for the most frequent polymorphisms in white populations (CYP2C9*2, CYP2C9*3, CYP2C19*2, CYP2C19*3, CYP2D6*3, CYP2D6*4, CYP2D6*6, and CYP3A5*3) was carried out before P450 phenotyping, excluding the donors with nonfunctional alleles of CYP2C9, CYP2C19, and CYP2D6 and those with a functional CYP3A5*1 allele from a correlation analysis. The hepatic mRNA levels of CYP1A2, CYP2B6, CYP2C9, CYP2C19, CYP2D6, and CYP3A4 displayed a strong association with P450 activities in the liver, whereas the expression of CYP1A2, CYP2C9, CYP2C19, and CYP3A4 in leukocytes was proven to reflect the hepatic activities of these P450 species. The leukocytes were found to be inappropriate cells for the assessment of hepatic CYP2B6 and CYP2D6 activities. Combining the results of P450 genotyping and phenotyping analyses, patients' drug-metabolizing capacities can be estimated by the P450 expression in the liver and in leukocytes with some limitations. Patients' genetic and nongenetic variations in P450 status can guide the appropriate selection of drugs and the optimal dose, minimizing the risk of harmful side effects and ensuring a successful outcome of drug therapy.

High-resolution melting curve analysis to establish CYP2C19∗2 single nucleotide polymorphism: Comparison with hydrolysis SNP analysis.

Clinically significant consequences of CYP2C19 polymorphism in drug metabolism require reliable genotyping methods for assaying large numbers of blood samples. Hydrolysis single nucleotide polymorphism analysis and high-resolution melting curve (HRM) analysis successfully genotyped the 114 liver donors for CYP2C19∗2. However, the HRM analysis was found to be more efficient, rapid, user- and cost-friendly method.

Drug interaction potential of 2-((3,4-dichlorophenethyl)(propyl)amino)-1-(pyridin-3-yl)ethanol (LK-935), the novel nonstatin-type cholesterol-lowering agent.

The widely prescribed lipid-lowering statins are considered to be relatively safe drugs. However, the risk of severe myopathy and drug interactions as a consequence of statin therapy provides a challenge for development of novel cholesterol-lowering agents, targeting enzymes other than HMG-CoA reductase. The novel pyridylethanol-(phenylethyl)amine derivative, (2-((3,4-dichlorophenethyl)(propyl)-amino)-1-(pyridin-3-yl)ethanol (LK-935), blocking lanosterol 14alpha-demethylase, was demonstrated to efficiently reduce cholesterol biosynthesis. The drug interaction potential of LK-935 was investigated and compared with that of atorvastatin and rosuvastatin in primary human hepatocytes. Clear evidence was provided for the induction of CYP3A4 by LK-935. LK-935 was proved to be a potent human pregnane X receptor (hPXR) activator as a prerequisite for the transcriptional activation of CYP3A4 gene; however, the rapid metabolism of LK-935 in primary hepatocytes prevented maximal CYP3A4 induction. Therefore, the induction of CYP3A4 by LK-935 may be prone to mild or negligible drug interactions. However, because CYP3A4 and also CYP2C9 play a significant role in LK-935 metabolism, the inhibition of these cytochromes P450 by coadministered drugs may lead to some increase in the LK-935 concentration required for the potent induction of CYP3A4. Rosuvastatin was found to increase human constitutive androstane receptor (hCAR)-mediated transcription of CYP3A4, CYP2C9, and CYP2B6 genes, predicting the consequent potential for drug interactions with several coadministered drugs. Activation of hCAR and hPXR by atorvastatin and the subsequent induction of not only CYP2B6 and CYP3A4 but also of CYP2C9 present an additional target by which atorvastatin, a widely used cholesterol-lowering drug, can modify the kinetics of numerous drugs.

Investigation of the CYP2C9 induction profile in human hepatocytes by combining experimental and modelling approaches.

The goal of the present review is to characterise the induction profile of CYP2C9, a member of the cytochrome P450 superfamily. Since the mechanism of CYP2C9 induction is fairly complex, with parallel processes triggered by various inducers, an evaluation of the experimental results is often a great challenge. At least three nuclear receptors, the glucocrticoid receptor (GR), the pregnane X receptor (PXR) and the constitutive androstane receptor (CAR), are known to mediate the CYP2C9 gene induction in man. However, mathematical modelling and simulation can provide an appropriate tool for the interpretation of CYP2C9 regulatory mechanisms. As an example, we present modelling and simulation approaches of the CYP2C9 gene expression in human hepatocytes treated with well-known CYP2C9 inducers: the steroid hormone precursor dehydroepiandrosterone (DHEA) and the synthetic glucocorticoid dexamethasone (DXM). The results of the analysis suggest that in addition to the potent function of GR and the further involvement of PXR and CAR activated by DXM or DHEA, an additional factor might play a role in CYP2C9 regulation by DHEA. The novel potential candidate for DHEA action in CYP2C9 induction is likely to be the estrogen receptor. Additionally, the balance of DHEA sulphation-desulphation processes should also be considered in any description of DHEA-induced CYP2C9 profiles.