Identification of non-adherence to adjuvant letrozole using a population pharmacokinetics approach in hormone receptor-positive breast cancer patients.

BACKGROUND: Letrozole, an aromatase inhibitor metabolised via CYP2A6 and CYP3A4/5 enzymes, is used as adjuvant therapy for women with hormone receptor (HR)-positive early breast cancer. The objective of this study was to quantify the impact of CYP2A6 genotype on letrozole pharmacokinetics (PK), to identify non-adherent patients using a population approach and explore the possibility of a relationship between non-adherence and early relapse. METHODS: Breast cancer patients enrolled in the prospective PHACS study (ClinicalTrials.gov NCT01127295) and treated with adjuvant letrozole 2.5 mg/day were included. Trough letrozole concentrations (Css,trough) were measured every 6 months for 3 years by a validated LC-MS/MS method. Concentration-time data were analysed using non-linear mixed effects modelling. Three methods were evaluated for identification of non-adherent subjects using the base PK model. RESULTS: 617 patients contributing 2534 plasma concentrations were included and led to a one-compartment PK model with linear absorption and elimination. Model-based methods identified 28 % of patients as non-adherent based on high fluctuations of their Css,trough compared to 3 % based on patient declarations. The covariate analysis performed in adherent subjects revealed that CYP2A6 intermediate (IM) and slow metabolisers (SM) had 21 % (CI95 % = 12 - 30 %) and 46 % (CI95 % = 41 - 51 %) lower apparent clearance, respectively, compared to normal and ultrarapid metabolisers (NM+UM). Early relapse (19 patients) was not associated with model-estimated, concentration-based or declared adherence in the total population (p = 0.41, p = 0.37 and p = 0.45, respectively). CONCLUSIONS: These findings will help future investigations focusing on the exposure-efficacy relationship for letrozole in adjuvant setting.

The Frequency of CYP2D6 and CYP3A4/5 Genotypes and The Impact of Their Allele Translation and Phenoconversion-Predicted Enzyme Activity on Risperidone Pharmacokinetics in Saudi Children with Autism.

Data on the role of CYP2D6 and CYP3A4/5 polymorphisms in relation to risperidone (RIS) pharmacokinetics (PK) in children are relatively limited and inconsistent. This is partially attributable to the limited coverage of CYP2D6 and CYP3A4/5 metabolizer phenotypes, particularly those of poor and ultrarapid metabolizers (PMs and UMs), which has led to calls for studies of populations with a non-European background that may carry variants that are less frequent in Europeans. Children ≤ 18 years old with at least 8 weeks of a RIS-based regimen were recruited from three autism centers in Riyadh, Saudi Arabia. The primary outcomes measured were plasma concentrations of RIS and 9-hydroxyrisperidone (9-OH-RIS) and their dose-adjusted (C/D) ratios as a function of phenotypes and activity score (AS). For accurate DNA genotyping, targeted pharmacogenomic testing with the Axiom PharmacoFocus Array was performed via examination of a broad collection of probesets targeting CYP2D6 and CYP3A4/5 variants. The frequency of genotypes/phenotypes and the impact of their allele translation and phenoconversion-predicted enzyme activity were examined. The final cohort included 83 individuals. The most common CYP2D6 phenotype in our population was normal metabolizers (NMs, 66.3%). Inconsistent with some previous studies, the three phenotypes of intermediate metabolizers (IMs), NMs, and UMs were significantly different in terms of RIS concentration, the RIS/9-OH-RIS ratio, the RIS C/D ratio and the 9-OH-RIS C/D ratio. According to AS analyses, there were statistically significant differences in the RIS concentration (P = 0.013), RIS/9-OH-RIS ratio (P < 0.001) and RIS C/D ratio (P = 0.030) when patients were categorized into AS ≤ 1 vs. AS > 1. None of the CYP3A4/5 star allele translated phenotypes revealed a significant influence on any of the RIS PK parameters. Notably, neither CYP2D6 nor CYP3A4/5 phenotyping demonstrated a significant impact on the total active moiety, suggesting that other gene variants could modulate RIS PK. The study confirmed the previously reported partial impact of the CYP2D6 gene on RIS PK. However, future studies using contemporary genotyping techniques targeting a wide range of variants in other candidate genes must be conducted to further examine their interactive effects on RIS PK and the clinical response.

Activation of PXR causes drug interactions with Paxlovid in transgenic mice.

Paxlovid is a nirmatrelvir (NMV) and ritonavir (RTV) co-packaged medication used for the treatment of coronavirus disease 2019 (COVID-19). The active component of Paxlovid is NMV and RTV is a pharmacokinetic booster. Our work aimed to investigate the drug/herb-drug interactions associated with Paxlovid and provide mechanism-based guidance for the clinical use of Paxlovid. By using recombinant human cytochrome P450s (CYPs), we confirmed that CYP3A4 and 3A5 are the major enzymes responsible for NMV metabolism. The role of CYP3A in Paxlovid metabolism were further verified in Cyp3a-null mice, which showed that the deficiency of CYP3A significantly suppressed the metabolism of NMV and RTV. Pregnane X receptor (PXR) is a ligand-dependent transcription factor that upregulates CYP3A4/5 expression. We next explored the impact of drug- and herb-mediated PXR activation on Paxlovid metabolism in a transgenic mouse model expressing human PXR and CYP3A4/5. We found that PXR activation increased CYP3A4/5 expression, accelerated NMV metabolism, and reduced the systemic exposure of NMV. In summary, our work demonstrated that PXR activation can cause drug interactions with Paxlovid, suggesting that PXR-activating drugs and herbs should be used cautiously in COVID-19 patients receiving Paxlovid.

Mutant CYP3A4/5 Correlated with Clinical Outcomes by Affecting Rivaroxaban Pharmacokinetics and Pharmacodynamics in Patients with Atrial Fibrillation.

PURPOSE: This study was designed to investigate the impact of single-nucleotide polymorphism-encoded cytochrome P450 enzymes (CYP3A4/5) on clinical outcomes of rivaroxaban in patients with non-valvular atrial fibrillation (NVAF) based on pharmacokinetics and pharmacodynamics (PK/PD) aspects. METHOD: A prospective study enrolling 165 rivaroxaban-treated patients with NVAF was conducted. Genotyping of CYP3A4 (rs2242480, rs2246709, rs3735451, and rs4646440) and CYP3A5 (rs776746) was performed to explore their impact on the trough plasma concentrations (Ctrough) of rivaroxaban, coagulation indicators at the Ctrough including activated partial thromboplastin time (APTT) and prothrombin time (PT), and clinical outcomes. RESULTS: Patients with mutant genotype CYP3A4 (rs2242480, rs2246709, and rs3735451) and CYP3A5 (rs776746) had higher levels of rivaroxaban Ctrough, PT values than that of wild-type. Furthermore, a positive relationship was revealed between Ctrough and PT (r = 0.212, p = 0.007), while no significant correlation was found between Ctrough and APTT. Regarding the clinical outcomes, the minor allele carriers on rs3735451 and the minor allele (A) carriers on rs2246709 were associated with higher incidence of minor bleeding (p = 0.028 and p = 0.038, respectively) and were identified as the independent risk factors of minor bleeding treated with rivaroxaban (p = 0.024 and p = 0.036, respectively), with the receiver operating characteristic (ROC) curve validated (AUC = 0.8956, 95% CI: 0.829-0.962). CONCLUSION: The CYP3A4 polymorphisms (rs2242480, rs2246709, and rs3735451) and CYP3A5 rs776746 were associated with variations in rivaroxaban PK/PD. The minor allele (C) carriers on rs3735451 and the minor allele (A) carriers on rs2246709 were correlated with clinical outcomes.

Pharmacokinetic Evaluation of Tacrolimus in Chinese Adult Patients during the Early Stages Post-Lung Transplantation.

BACKGROUND: Although tacrolimus has been widely used in patients undergoing lung transplantation, few studies have reported the pharmacokinetics of tacrolimus in Chinese patients after lung transplantation. Thus, we aimed to investigate the pharmacokinetics and influential factors in this patient cohort in the early stage after lung transplantation. METHODS: We enrolled 14 adult lung transplant recipients who were treated with tacrolimus and then intensively collected blood samples within a 12-h dosing interval. The pharmacokinetic parameters of tacrolimus were calculated using non-compartmental analysis, and the influence of pathophysiological characteristics and CYP3A5*3 and CYP3A4*1G genotypes on the pharmacokinetics of tacrolimus was assessed. Using linear regression analysis, we investigated the correlation between tacrolimus concentration at different sampling points and measured the area under the time-concentration curve (AUC0-12h). RESULTS: Geometric mean of apparent clearance (CL/F) was 18.13 ± 1.65 L/h in non-CYP3A5*3/*3 carriers, five times higher than that in CYP3A5*3/*3 carriers (p < 0.001). Furthermore, the tacrolimus concentration 4 h after administration had the strongest correlation with AUC0-12h (R2 = 0.979). CONCLUSION: The pharmacokinetics of tacrolimus varied largely between patients during the early stage post-transplantation, which could be partially explained by CYP3A5*3 genetic polymorphisms.

Activation of PXR causes drug interactions with Paxlovid in transgenic mice

Paxlovid is a nirmatrelvir (NMV) and ritonavir (RTV) co-packaged medication used for the treatment of coronavirus disease 2019 (COVID-19). The active component of Paxlovid is NMV and RTV is a pharmacokinetic booster. Our work aimed to investigate the drug/herb-drug interactions associated with Paxlovid and provide mechanism-based guidance for the clinical use of Paxlovid. By using recombinant human cytochrome P450s (CYPs), we confirmed that CYP3A4 and 3A5 are the major enzymes responsible for NMV metabolism. The role of CYP3A in Paxlovid metabolism were further verified in Cyp3a-null mice, which showed that the deficiency of CYP3A significantly suppressed the metabolism of NMV and RTV. Pregnane X receptor (PXR) is a ligand-dependent transcription factor that upregulates CYP3A4/5 expression. We next explored the impact of drug- and herb-mediated PXR activation on Paxlovid metabolism in a transgenic mouse model expressing human PXR and CYP3A4/5. We found that PXR activation increased CYP3A4/5 expression, accelerated NMV metabolism, and reduced the systemic exposure of NMV. In summary, our work demonstrated that PXR activation can cause drug interactions with Paxlovid, suggesting that PXR-activating drugs and herbs should be used cautiously in COVID-19 patients receiving Paxlovid.

Evaluation of hepatic CYP3A enzyme activity using endogenous markers in lung cancer patients treated with cisplatin, dexamethasone, and aprepitant.

PURPOSE: Aprepitant is used with dexamethasone and 5-HT3 receptor antagonists as an antiemetic treatment for chemotherapy, including cisplatin. Aprepitant is a substrate of cytochrome P450 (CYP) 3A4 and is known to cause its inhibition and induction. In addition, dexamethasone is a CYP3A4 substrate that induces CYP3A4 and CYP3A5 expression. In this study, we aimed to quantitatively evaluate the profile of CYP3A activity using its endogenous markers in non-small cell lung cancer patients receiving a standard cisplatin regimen with antiemetics, including aprepitant. METHODS: Urinary 11ß-hydroxytestosterone (11ß-OHT)/testosterone concentration ratio and plasma 4ß-hydroxycholesterol (4ß-OHC) concentrations were measured before and after cisplatin treatment (days 1, 4, and 8). CYP3A5 was genotyped, and plasma aprepitant concentrations were measured on day 4 to examine its influence on CYP3A endogenous markers. RESULTS: The urinary 11ß-OHT/testosterone concentration ratio in the 35 patients included in this study increased by 2.65-fold and 1.21-fold on days 4 and 8 compared with day 1, respectively. Their plasma 4ß-OHC concentration increased by 1.46-fold and 1.66-fold, respectively. The mean plasma aprepitant concentration on day 4 was 1,451 ng/mL, which is far lower than its inhibitory constant. The allele frequencies of CYP3A5*1 and CYP3A5*3 were 0.229 and 0.771, respectively. In patients with the CYP3A5*1 allele, the plasma 4ß-OHC concentration was significantly lower at baseline but more potently increased with chemotherapy. CONCLUSION: CYP3A activity was significantly induced from day 4 to day 8 in patients receiving cisplatin and three antiemetic drugs.

[An in vitro study on metabolic interaction between main active components of Salviae Miltiorrhizae Radix et Rhizoma and tacrolimus].

The purpose of this study was to investigate the inhibitory effects of the main active components of Salviae Miltiorrhizae Radix et Rhizoma on the metabolism of tacrolimus mediated by CYP3 A4/5 enzyme, so as to predict the potential drug-drug interaction(DDI) in clinical use. First, the reversible inhibitory activities of five active components of Salviae Miltiorrhizae Radix et Rhizoma(tanshinone Ⅰ, tanshinone Ⅱ_A, cryptotanshinone, salvianolic acid B, dihydrotanshinone Ⅰ) on the metabolism of tacrolimus was investigated by using human liver microsomes(HLM) and recombinant human CYP3 A4/5 enzyme in vitro, then the dose-dependent inhibition of CYP3 A4/5 activity was calculated in HLM. Finally, the time-dependent inhibition(TDI) activities of five active components were studied in HLM through the robust single point inhibition test. In addition, a simple and rapid liquid chromatography tandem mass spectrometry method(HPLC-MS/MS) for the determination of tacrolimus was established in this study. The results showed that dihydrotanshinone Ⅰ had a strong inhibitory effect on the metabolism of tacrolimus in both HLM and rCYP3 A4/5 enzyme systems, and the inhibitory potential IC_(50) in HLM was 6.0 µmol·L~(-1), while the other four active components of Salviae Miltiorrhizae Radix et Rhizoma exhibited relatively weak inhibition on CYP3 A4/5 activity with inhibition rate less than 30% at 10 µmol·L~(-1). Furthermore, the TDI activity of five active components of Salviae Miltiorrhizae Radix et Rhizoma at 50 µmol·L~(-1) was 5.5%-15.9%. The above results suggested that clinical DDI between tacrolimus and Salviae Miltiorrhizae Radix et Rhizoma may occur when the active components of Salviae Miltiorrhizae Radix et Rhizoma achieved a relative high concentration in human. In conclusion, this study provided a data reference for the research on drug interaction of tacrolimus and Salviae Miltiorrhizae Radix et Rhizoma as well as rational drug use in clinical practice.

The potential of Atorvastatin for chronic lung diseases therapy.

Atorvastatin (ATO) is of the statin class and is used as an orally administered lipid-lowering drug. ATO is a reversible synthetic competitive inhibitor of 3-hydroxy-3-methyl-glutaryl-CoA (HMG-CoA) reductase thus leading to a reduction in cholesterol synthesis. It has recently been demonstrated that ATO has different pharmacological actions, which are unrelated to its lipid-lowering effects and has the ability to treat chronic airway diseases. This paper reviews the potential of ATO as an anti-inflammatory, antioxidant, and anti-proliferative agent after oral or inhaled administration. This paper discusses the advantages and disadvantages of using ATO under conditions associated with those found in the airways. This treatment could potentially be used to support the formulating of ATO as an inhaler for the treatment of chronic respiratory diseases.

Pharmacokinetic functions of human induced pluripotent stem cell-derived small intestinal epithelial cells.

To develop a novel intestinal drug absorption system using intestinal epithelial cells derived from human induced pluripotent stem (iPS) cells, the cells must possess sufficient pharmacokinetic functions. However, the CYP3A4/5 activities of human iPS cell-derived small intestinal epithelial cells prepared using conventional differentiation methods is low. Further, studies of the CYP3A4/5 activities of human iPS-derived and primary small intestinal cells are not available. To fill this gap in our knowledge, here we used forskolin to develop a new differentiation protocol that activates adenosine monophosphate signaling. mRNA expressions of human iPS cell-derived small intestinal epithelial cells, such as small intestine markers, drug-metabolizing enzymes, and drug transporters, were comparable to or greater than those of the adult small intestine. The activities of CYP3A4/5 in the differentiated cells were equal to those of human primary small intestinal cells. The differentiated cells had P-glycoprotein and PEPT1 activities equivalent to those of Caco-2 cells. Differentiated cells were superior to Caco-2 cells for predicting the membrane permeability of drugs that were absorbed through a paracellular pathway and via drug transporters. In summary, here we produced human iPS cell-derived small intestinal epithelial cells with CYP3A4/5 activities equivalent to those of human primary small intestinal cells.

The cytochrome P450 metabolic profiling of SMU-B in vitro, a novel small molecule tyrosine kinase inhibitor.

A novel small molecule tyrosine kinase inhibitor 6-[6-Amino-5-[(1R)-1-(2,6-dichloro-3-fluorophenyl)ethoxy]-3-pyridyl]-1'-methylspiro[indoline-3,4'-piperidine]-2-one (SMU-B) had good activity against ALK (anaplastic lymphoma kinase) and ROS1 (c-ros oncogene 1) targets in non-small-cell lung cancer. The excellent bioactivity of SMU-B highlights the importance of determining its metabolic traits, which could provide meaningful information for further pharmacokinetic studies of SMU-B. In this work, we studied the metabolism of SMU-B in human liver microsomes. Three metabolites of SMU-B were identified by a quadrupole-time of flight tandem mass spectrometer (Q-TOF-MS), and the metabolic pathways of SMU-B were demethylation, dehydrogenation and oxidation. CYP3A4/5 was the principal isoform involved in SMU-B metabolism, as shown by chemical inhibition and recombination human enzyme studies. Additionally, a predication with a molecular docking model confirmed that SMU-B could interact with the active sites of CYP3A4 and CYP3A5. This study illuminates the metabolic profile of the anti-tumor drug SMU-B, which will accelerate its clinical use.

Influences of ABC transporter and CYP3A4/5 genetic polymorphisms on the pharmacokinetics of lenvatinib in Chinese healthy subjects.

PURPOSE: To investigate whether the CYP3A4/5 and ABC transporter genetic polymorphisms could affect the pharmacokinetics of lenvatinib in Chinese healthy subjects. METHODS: Thirty-two healthy Chinese volunteers were enrolled and took oral administration of 8 mg lenvatinib. Plasma concentration of lenvatinib was determined by UPLC-MS/MS, the CYP3A4*1G, CYP3A5*3, ABCB1 (3435 C>T, 1236 C>T, 2677 G>T/A), ABCG2 (421 C>A, 34 G>A), and ABCC2-24 C>T genotypes were determined by SnapShot Technique. RESULTS: In ABCB1 3435T carriers (n = 19), AUC0-120h (815.7 (701.9-923.9) ng·h/mL) and AUC0-∞ (843.3 (722.2-977.7) ng·h/mL) were significantly higher than ABCB1 3435CC homozygous subjects (n = 13, 575.3 (513.7-756.9) ng·h/mL and 590.0 (540.5-782.0) ng·h/mL, respectively); on the contrary, the clearance (CL/F) of ABCB1 3435T carriers was significantly lower (9.5 (8.2-11.1) L/h vs. 13.6 (10.4-14.8) L/h). And the Cmax in CYP3A4*1G/*1G allele carrier subjects was higher than *1 carrier (73.4 ng/mL vs. 53.5 (46.1-60.6) ng/mL), but did not reach the level of significantly statistical difference. Genetic polymorphisms of ABCC2, ABCG2, and CYP3A5 could not influence pharmacokinetic parameters of lenvatinib. CONCLUSIONS: This work presented an evidence that the ABCB1 3435 C>T polymorphism could significantly affect the exposure and clearance of lenvatinib. These findings may explain the reasons for the huge inter-individual differences in lenvatinib, and should contribute to clinical individualized treatment.

Pharmacokinetic Drug Interactions of an Orally Available TRH Analog (Rovatirelin) With a CYP3A4/5 and P-Glycoprotein Inhibitor (Itraconazole).

The effects of itraconazole on the pharmacokinetics of rovatirelin were investigated in an open-label, single-sequence drug-drug interaction study in 16 healthy subjects. Subjects were administered a single oral dose of rovatirelin (1.6 mg) on day 1 and day 15. From day 8 through 16, subjects received daily oral doses of itraconazole (200 mg/day). Concentrations of rovatirelin and (thiazolylalanyl)methylpyrrolidine (TAMP), the major metabolite of rovatirelin formed by cytochrome P450 (CYP) 3A4/5, were determined in plasma and urine. Pharmacokinetic parameters were used to evaluate the drug-drug interaction potential of rovatirelin as a victim. With coadministration, maximum concentration (Cmax ) and area under the concentration-time curve extrapolated to infinity (AUCinf ) of rovatirelin increased 3.05-fold and 2.82-fold, respectively, and the 90% confidence intervals of the ratios for Cmax (2.64-3.52) and AUCinf (2.47-3.23) did not fall within the 0.8-1.25 boundaries. Urinary excretion of rovatirelin increased at almost the same ratio as the AUCinf ratio with coadministration; however, renal clearance did not change. Cmax , AUCinf , and urinary excretion of TAMP were decreased by coadministration. Itraconazole has the potential to inhibit drug transport via intestinal P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP); therefore, substrate assessments of rovatirelin for the 2 transporters were evaluated using Caco-2 cell monolayers. In vitro studies showed that rovatirelin is a substrate for P-gp but not for BCRP. The current study shows that itraconazole's effect on rovatirelin pharmacokinetics is mediated through inhibition of CYP3A4/5 and intestinal P-gp.

Tamsulosin Associated with Interstitial Lung Damage in CYP2D6 Variant Alleles Carriers.

Drugs are serious but underestimated causative agents of interstitial lung disease (ILD). Both cytotoxic and immune mechanisms may be involved in drug-induced ILD (DI-ILD). We aimed to investigate whether polymorphisms of relevant CYP enzymes involved in the metabolization of tamsulosin might explain the pathologic mechanism of the DI-ILD in the cases with suspected tamsulosin DI-ILD. We collected 22 tamsulosin-associated DI-ILD cases at two ILD Expertise Centers in the Netherlands between 2009 and 2020. CYP2D6, CYP2C9, CYP2C19, CYP3A4, and CYP3A5 single nucleotide polymorphisms were genotyped and compared with a control group of 78 healthy Caucasian male volunteers. Nine cases were phenotyped as CYP2D6 poor metabolizers and 13 as CYP2D6 intermediate metabolizers. The phenotypes of the cases differed significantly from those of the healthy controls, with more poor metabolizers. After withdrawal of tamsulosin, the pulmonary condition of three cases had improved, six patients had stabilized, and one patient stabilized after reducing the tamsulosin dose. The described 22 cases suggest that an association between the presence of CYP2D6 allelic variants and tamsulosin-associated ILD is highly likely. These cases highlight the importance of both clinical and genetic risk stratification aimed to achieve a more accurate prevention of DI-ILD in the future and enhance the quality of life of patients.

Influence of Germline Genetics on Tacrolimus Pharmacokinetics and Pharmacodynamics in Allogeneic Hematopoietic Stem Cell Transplant Patients.

Tacrolimus exhibits high inter-patient pharmacokinetics (PK) variability, as well as a narrow therapeutic index, and therefore requires therapeutic drug monitoring. Germline mutations in cytochrome P450 isoforms 4 and 5 genes (CYP3A4/5) and the ATP-binding cassette B1 gene (ABCB1) may contribute to interindividual tacrolimus PK variability, which may impact clinical outcomes among allogeneic hematopoietic stem cell transplantation (HSCT) patients. In this study, 252 adult patients who received tacrolimus for acute graft versus host disease (aGVHD) prophylaxis after allogeneic HSCT were genotyped to evaluate if germline genetic variants associated with tacrolimus PK and pharmacodynamic (PD) variability. Significant associations were detected between germline variants in CYP3A4/5 and ABCB1 and PK endpoints (e.g., median steady-state tacrolimus concentrations and time to goal tacrolimus concentration). However, significant associations were not observed between CYP3A4/5 or ABCB1 germline variants and PD endpoints (e.g., aGVHD and treatment-emergent nephrotoxicity). Decreased age and CYP3A5*1/*1 genotype were independently associated with subtherapeutic tacrolimus trough concentrations while CYP3A5*1*3 or CYP3A5*3/*3 genotypes, myeloablative allogeneic HSCT conditioning regimen (MAC) and increased weight were independently associated with supratherapeutic tacrolimus trough concentrations. Future lines of prospective research inquiry are warranted to use both germline genetic and clinical data to develop precision dosing tools that will optimize both tacrolimus dosing and clinical outcomes among adult HSCT patients.

Hormonal status affects plasma exposure of tamoxifen and its main metabolites in tamoxifen-treated breast cancer patients.

BACKGROUND: Tamoxifen is considered a prodrug of its active metabolite endoxifen, which is dependent on the CYP2D6 and CYP3A enzymes. Tamoxifen pharmacokinetic variability influences endoxifen exposure and, consequently, its clinical outcome. This study investigated the impact of hormonal status on the pharmacokinetics of tamoxifen and its metabolites in TAM-treated breast cancer patients. METHODS: TAM-treated breast cancer patients (n = 40) previously believed to have CYP3A activity within the normal range based on oral midazolam and phenotyped as CYP2D6 normal metabolizers using oral metoprolol were divided into two groups according to premenopausal (n = 20; aged 35-50 years) or postmenopausal (n = 20; aged 60-79 years) status. All patients were treated with 20 mg/day tamoxifen for at least three months. Serial plasma samples were collected within the 24 h dose interval for analysis of unchanged tamoxifen, endoxifen, 4-hydroxytamoxifen and N-desmethyltamoxifen quantified by LC-MS/MS. CYP activities were assessed using midazolam apparent clearance (CYP3A) and the metoprolol/alfa-hydroxymetoprolol plasma metabolic ratio (CYP2D6). CYP3A4, CYP3A5 and CYP2D6 SNPs and copy number variation were investigated using TaqMan assays. RESULTS: Postmenopausal status increased steady-state plasma concentrations (Css) of tamoxifen (116.95 vs 201.23 ng/mL), endoxifen (8.01 vs 18.87 ng/mL), N-desmethyltamoxifen (485.16 vs 843.88 ng/mL) and 4-hydroxytamoxifen (2.67 vs 4.11 ng/mL). The final regression models included hormonal status as the only predictor for Css of tamoxifen [ß-coef ± SE, p-value (75.03 ± 17.71, p = 0.0001)] and 4-hydroxytamoxifen (1.7822 ± 0.4385, p = 0.0002), while endoxifen Css included hormonal status (8.578 ± 3.402, p = 0.02) and race (11.945 ± 2.836, p = 0.007). For N-desmethyltamoxifen Css, the final model was correlated with hormonal status (286.259 ± 76.766, p = 0.0007) and weight (- 8.585 ± 3.060, p = 0.008). CONCLUSION: The premenopausal status was associated with decreased endoxifen plasma concentrations by 135% compared to postmenopausal status. Thus, the endoxifen plasma concentrations should be monitored mainly in the premenopausal period to maintain plasma levels above the efficacy threshold value. TRIAL REGISTRATION: RBR-7tqc7k.

CYP3A4/5 mediates the metabolic detoxification of humantenmine, a highly toxic alkaloid from Gelsemium elegans Benth.

Gelsemium elegans Benth., a well-known toxic herbal plant, is widely used to treat rheumatic arthritis, inflammation and other diseases. Gelsemium contains humantenmine (HMT), which is an important bioactive and toxic alkaloid. Cytochrome P450 enzymes (CYPs) play important roles in the elimination and detoxification of exogenous substances. This study aimed to investigate the roles of CYPs in the metabolism and detoxification of HMT. First, metabolic studies were performed in vitro by using human liver microsomes, selective chemical inhibitors and recombinant human CYPs. Results indicated that four metabolites, including hydroxylation and oxidation metabolites, were found in human liver microsomes and identified based on their high-resolution mass spectrum. The isoform responsible for HMT metabolism was mainly CYP3A4/5. Second, the toxicity of HMT on L02 cells in the presence of the nicotinamide adenine dinucleotide phosphate system (NADPH) was significantly less than that without NADPH system. A CYP3A4/5 activity inhibition model was established by intraperitoneally injecting ketoconazole in mice and used to evaluate the role of CYP3A4/5 in HMT detoxification. In this model, the 14-day survival rate of the mice decreased to 17% after they were intragastrically treated with HMT, along with hepatic injury and increasing alanine aminotransferase (ALT) /aspartate aminotransferase (AST) levels. Overall, CYP3A4/5 mediated the metabolism and detoxification of HMT.

Human mass balance, pharmacokinetics and metabolism of rovatirelin and identification of its metabolic enzymes in vitro.

The mass balance, pharmacokinetics and metabolism of rovatirelin were characterised in healthy male subjects after a single oral dose of [14C]rovatirelin. [14C]Rovatirelin was steadily absorbed, and the peak concentrations of radioactivity and rovatirelin were observed in plasma at 5-6 h after administration. The AUCinf of radioactivity was 4.9-fold greater than that of rovatirelin. Rovatirelin and its metabolite (thiazoylalanyl)methylpyrrolidine (TAMP) circulated in plasma as the major components. The total radioactivity recovered in urine and faeces was 89.0% of the administered dose. The principal route of elimination was excretion into faeces (50.1% of the dose), and urinary excretion was the secondary route (36.8%). Rovatirelin was extensively metabolised to 20 metabolites, and TAMP was identified as the major metabolite in plasma and excreta among its metabolites. To identify the metabolic enzymes responsible for TAMP formation, the in vitro activity was determined in human liver microsomes. The enzymatic activity depended on NADPH, and it was inhibited by ketoconazole. Furthermore, recombinant human cytochrome P450 (CYP) 3A4 and CYP3A5 displayed enzymatic activity in the assay. Therefore, CYP3A4/5 are the most important enzymes responsible for TAMP formation.

Absence of herb-drug interactions of mistletoe with the tamoxifen metabolite (E/Z)-endoxifen and cytochrome P450 3A4/5 and 2D6 in vitro.

BACKGROUND: Women diagnosed with breast cancer frequently seek complementary and alternative (CAM) treatment options that can help to cope with their disease and the side effects of conventional cancer therapy. Especially in Europe, breast cancer patients use herbal products containing mistletoe (Viscum album L.). The oldest and one of the most prescribed conventional drugs for the treatment of estrogen receptor positive breast cancer is tamoxifen. Aside from positive clinical experience with the combination of tamoxifen and mistletoe, little is known about possible herb-drug interactions (HDIs) between the two products. In the present in vitro study, we investigated the effect of standardized commercial mistletoe preparations on the activity of endoxifen, the major active metabolite of tamoxifen. METHODS: The estrogen receptor positive human breast carcinoma cell line MCF-7 was treated with (E/Z)-endoxifen hydrochloride in the presence and absence of a defined estradiol concentration. Each concentration of the drug was combined with fermented Viscum album L. extracts (VAE) at clinically relevant doses, and proliferation, apoptosis and cell cycle were analyzed. In parallel, possible inhibition of CYP3A4/5 and CYP2D6 was investigated using 50-donor mixed gender pooled human liver microsomes (HLMs). RESULTS: VAE did not inhibit endoxifen induced cytostasis and cytotoxicity. At higher concentrations, VAE showed an additive inhibitory effect. VAE preparations did not cause inhibition of CYP3A4/5 and CYP2D6 catalyzed tamoxifen metabolism. CONCLUSIONS: The in vitro results suggest that mistletoe preparations can be used in combination with tamoxifen without the risk of HDIs.