Identification of reversible OATP1B1 and time-dependent CYP3A4 inhibition as the major risk factors for drug-induced cholestasis (DIC).

Hepatic bile acid regulation is a multifaceted process modulated by several hepatic transporters and enzymes. Drug-induced cholestasis (DIC), a main type of drug-induced liver injury (DILI), denotes any drug-mediated condition in which hepatic bile flow is impaired. Our ability in translating preclinical toxicological findings to human DIC risk is currently very limited, mainly due to important interspecies differences. Accordingly, the anticipation of clinical DIC with available in vitro or in silico models is also challenging, due to the complexity of the bile acid homeostasis. Herein, we assessed the in vitro inhibition potential of 47 marketed drugs with various degrees of reported DILI severity towards all metabolic and transport mechanisms currently known to be involved in the hepatic regulation of bile acids. The reported DILI concern and/or cholestatic annotation correlated with the number of investigated processes being inhibited. Furthermore, we employed univariate and multivariate statistical methods to determine the important processes for DILI discrimination. We identified time-dependent inhibition (TDI) of cytochrome P450 (CYP) 3A4 and reversible inhibition of the organic anion transporting polypeptide (OATP) 1B1 as the major risk factors for DIC among the tested mechanisms related to bile acid transport and metabolism. These results were consistent across multiple statistical methods and DILI classification systems applied in our dataset. We anticipate that our assessment of the two most important processes in the development of cholestasis will enable a risk assessment for DIC to be efficiently integrated into the preclinical development process.

Role of Cytochrome P450 3A4 in Cancer Drug Resistance: Challenges and Opportunities.

One of the biggest obstacles to the treatment of diseases, particularly serious conditions like cancer, is therapeutic resistance. The process of drug resistance is influenced by a number of important variables, including MDR genes, drug efflux, low-quality medications, inadequate dosage, etc. Drug resistance must be addressed, and new combinations based on the pharmacokinetics/pharmacodynamics (PK-PD) characteristics of the partner pharmaceuticals must be developed in order to extend the half-lives of already available medications. The primary mechanism of drug elimination is hepatic biotransformation of medicines by cytochrome P450 (CYP) enzymes; of these CYPs, CYP3A4 makes up 30-40% of all known cytochromes that metabolize medications. Induction or inhibition of CYP3A4-mediated metabolism affects the pharmacokinetics of most anticancer drugs, but these details are not fully understood and highlighted because of the complexity of tumor microenvironments and various influencing patient related factors. The involvement of CYPs, particularly CYP3A4 and other drug-metabolizing enzymes, in cancer medication resistance will be covered in the current review.

Effects of CYP3A4 genetic polymorphisms on the pharmacokinetics and efficacy of perampanel in Chinese pediatric patients with epilepsy.

OBJECTIVE: This study was the first to evaluate the effect of CYP3A4 gene polymorphisms on the plasma concentration and effectiveness of perampanel (PER) in Chinese pediatric patients with epilepsy. METHODS: We enrolled 102 patients for this investigation. The steady-state concentration was determined after patients maintained a consistent PER dosing regimen for at least 21 days. Plasma PER concentrations were measured using liquid chromatography-tandem mass spectrometry. Leftover samples from standard therapeutic drug monitoring were allocated for genotyping analysis. The primary measure of efficacy was the rate of seizure reduction with PER treatment at the final check-up. RESULTS: The CYP3A4×10 GC phenotype exhibited the highest average plasma concentration of PER at 491.1 ±â€¯328.1 ng/mL, in contrast to the CC phenotype at 334.0 ±â€¯161.1 ng/mL. The incidence of adverse events was most prominent in the CYP3A4×1 G TT and CYP3A4×10 GC groups, with rates of 77.8 % (7 of 9 patients) and 50.0 % (46 of 92 patients), respectively. Moreover, the percentage of patients for whom PER was deemed ineffective was least in the CYP3A4×1 G TT and CYP3A4×10 CC groups, recorded at 11.1 % (1 of 9 patients) and 10.0 % (1 of 10 patients), respectively. There was a significant correlation between PER plasma concentration and either exposure or toxicity (both with p < 0.05). We suggest a plasma concentration range of 625-900 ng/mL as a suitable reference for PER in Chinese patients with epilepsy. CONCLUSION: The CYP3A4×10 gene's genetic polymorphisms influence plasma concentrations of PER in Chinese pediatric patients with epilepsy. Given that both efficacy and potential toxicity are closely tied to plasma PER levels, the CYP3A4 genetic phenotype should be factored in when prescribing PER to patients with epilepsy.

Steroid-tacrolimus drug-drug interaction and the effect of CYP3A genotypes.

AIMS: Tacrolimus, metabolized by CYP3A4 and CYP3A5 enzymes, is susceptible to drug-drug interactions (DDI). Steroids induce CYP3A genes to increase tacrolimus clearance, but the effect is variable. We hypothesized that the extent of the steroid-tacrolimus DDI differs by CYP3A4/5 genotypes. METHODS: Kidney transplant recipients (n = 2462) were classified by the number of loss of function alleles (LOF) (CYP3A5*3, *6 and *7 and CYP3A4*22) and steroid use at each tacrolimus trough in the first 6 months post-transplant. A population pharmacokinetic analysis was performed by nonlinear mixed-effect modelling (NONMEM) and stepwise covariate modelling to define significant covariates affecting tacrolimus clearance. A stochastic simulation was performed and translated into a Shiny application with the mrgsolve and Shiny packages in R. RESULTS: Steroids were associated with modestly higher (3%-11.8%) tacrolimus clearance. Patients with 0-LOF alleles receiving steroids showed the greatest increase (11.8%) in clearance compared to no steroids, whereas those with 2-LOFs had a negligible increase (2.6%) in the presence of steroids. Steroid use increased tacrolimus clearance by 5% and 10.3% in patients with 1-LOF and 3/4-LOFs, respectively. CONCLUSIONS: Steroids increase the clearance of tacrolimus but vary slightly by CYP3A genotype. This is important in individuals of African ancestry who are more likely to carry no LOF alleles, may more commonly receive steroid treatment, and will need higher tacrolimus doses.

1ß-Hydroxydeoxycholic Acid as an Endogenous Biomarker in Human Plasma for Assessment of CYP3A Clinical Drug-Drug Interaction Potential.

4ß-Hydroxycholesterol (4ß-HC) in plasma has been used as a biomarker to assess CYP3A drug-drug interaction (DDI) potential during drug development. However, due to the long half-life and narrow dynamic range of 4ß-HC, its use has been limited to the identification of CYP3A inducers, but not CYP3A inhibitors. The formation of 1ß-hydroxydeoxycholic acid (1ß-OH DCA) from deoxycholic acid (DCA) is mediated by CYP3A, thus 1ß-OH DCA can potentially serve as an alternative to 4ß-HC for assessment of CYP3A DDI potential. To study this feasibility, we developed a sensitive LC-MS/MS method for the simultaneous quantitation of 1ß-OH DCA and its glycine and taurine conjugates in human plasma with the LLOQ of 50 pg/mL, which enabled the quantitation of basal levels and further reduction. The method was applied to a DDI study to assess how 1ß-OH DCA and its glycine and taurine conjugates would respond to CYP3A induction or inhibition. Rifampin induction resulted in an increase of 1ß-OH DCA and its conjugates in plasma, with 6.8-, 7.8-, 8.3-, 10.3-fold increases of AUCLST, AUC24h, Cmax and mean concentrations for total 1ß-OH DCA (total of all three forms), respectively. Importantly, inhibition with itraconazole resulted in notable reduction of these biomarkers, with 84%, 85%, 82%, 81% reductions of AUCLST, AUC24h, Cmax and mean concentrations for total 1ß-OH DCA, respectively. This preliminary data demonstrates for the first time that total 1ß-OH DCA in plasma has the potential to serve as a biomarker for CYP3A DDI assessment in early clinical development and may provide key advantages over 4ß-HC. Significance Statement We have reported the use of total 1ß-Hydroxydeoxycholic Acid (1ß-OH DCA) (sum of 1ß-OH DCA and its glycine and taurine conjugates) plasma concentration as a biomarker for CYP3A activity. Itraconazole inhibition led to an 81-85% decrease of total 1ß-OH DCA plasma exposures, while rifampin induction led to a 6.8-10.3 fold increase of total 1ß-OH DCA plasma exposures. Using 1ß-OH DCA exposures in plasma also provides benefit of allowing PK and biomarker assessment using the same matrix, thus simplify collection procedures.

Synthesis and anticancer properties of a hybrid molecule with the testosterone and estradiol head-groups.

This is the first report on a unique hybrid molecule made of estradiol and testosterone (TS). This distinctive hybrid molecule (1) was designed to interact with both the estrogen receptor (ER) and the androgen receptor (AR) found in hormone-dependent female and male cancer cells, and was synthesized using ethynylestradiol (17EE) as the estrogenic component and 7α-(4-azido-but-2-enyl)-4-androsten-17ß-ol-3-one as the androgenic counterpart in a seven-step reaction with âˆ¼ 26 % overall yield. We reasoned that the dual receptor binding ability could allow 1 to act as an antihormone. This was tested on hormone-dependent and hormone-independent breast cancer (BCa) and prostate cancer (PCa) cells. The antiproliferative activity was also assessed on colon and skin cancer cells. We found that 1 was active against MCF7 (ER + ) BCa cells (IC50 of 4.9 µM), had lower inhibitory potency on LNCaP (AR + ) PCa cells (IC50 > 5 µM) and no effect on PC3 and DU145 (AR-) PCa cells. This suggests that the estrogenic component of 1 can interact with the ER on MCF7 cells more effectively than the androgenic component with the AR on LNCaP PCa cells, possibly due to a suboptimal spacer or linkage site(s). Nonetheless, the hybrid 1 was active against colon (HT-29) and melanoma (M21) cancer cells (IC50 of 3.5 µM and 2.3 µM, respectively), and had low cross-reactivity with the drug- and androgen-metabolizing cytochrome P450 3A4 (CYP3A4, IC50 ≫ 5 µM). These findings demonstrate the anticancer potential of 1 and warrant further explorations on this new type of hybrids.

Estimation of CDK inhibitors by RP-HPLC: application for pharmacokinetic interactions studies with PPIs.

Background: The study investigated pharmacokinetic interactions between palbociclib and ribociclib with proton pump inhibitors (PPIs) using the reverse-phase high-performance liquid chromatography (RP-HPLC) method. Methods: Developed RP-HPLC method quantified palbociclib and ribociclib in biological matrices. In vitro metabolic stability assays and in vivo studies in rats evaluated effect of omeprazole and esomeprazole on pharmacokinetics of palbociclib and ribociclib. Results: The RP-HPLC method was sensitive, accurate and linear. Esomeprazole and omeprazole decreased metabolic clearance of palbociclib and ribociclib by several folds. In vivo, esomeprazole elevated Cmax of palbociclib and ribociclib by 90.1% and 86.4%, whereas omeprazole reduced it by 32.0% and 16.8%, respectively. Conclusion: The RP-HPLC method was used to analyze in vitro and in vivo samples. Long-term treatment with PPIs affects pharmacokinetics of palbociclib and ribociclib, necessitating optimal chemotherapy regimen.

[Box: see text].

Functional assessment of CYP3A4 and CYP2C19 genetic polymorphisms on the metabolism of clothianidin invitro.

Clothianidin, classified as a second-generation neonicotinoid, has achieved extensive application due to its high efficacy against insect pests. This broad-spectrum usage has resulted in its frequent detection in environmental surveys. CYP2C19 and CYP3A4 are crucial for converting clothianidin to desmethyl-clothianidin (dm-clothianidin). The expression of these CYP450s can be significantly influenced by genetic polymorphisms. The objective of our research was to examine the catalytic effects of 27 CYP3A4 variants and 31 CYP2C19 variants on the metabolism of clothianidin within recombinant insect microsomes. These variants were assessed through a well-established incubation procedure. In addition, the concentration of its metabolite dm-clothianidin was quantified by employing an ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS). Lastly, the kinetic parameters of these CYP3A4 and CYP2C19 variants were calculated by applying Michaelis-Menten kinetic analysis to fit the data. The observed changes in enzyme activity were related to the metabolic transformation of clothianidin to dm-clothianidin. In the CYP2C19 metabolic pathway, one variant (CYP2C19.23) showed no notable change in intrinsic clearance (CLint), four variants (CYP2C19.29, .30, .31 and L16F) demonstrated a marked increase in CLint (110.86-183.46 %), and the remaining 25 variants exhibited a considerable decrease in CLint (26.38-89.79 %), with a maximum decrease of 73.62 % (CYP2C19.6). In the CYP3A4 metabolic pathway, 26 variants demonstrated significantly reduced CLint (10.54-52.52 %), with a maximum decrease of 89.46 % (CYP3A4.20). Our results suggested that most variants of CYP3A4 and CYP2C19 significantly altered the enzymatic activities associated with clothianidin metabolism to various degrees. This study provides new insights into assessing the metabolic behavior of pesticides and delivers crucial data that can guide clinical detoxification strategies.

The Frequencies distribution of CYP3A5 rs776746 and ABCB1 rs1045642 polymorphisms in the west Algerian population and relationships with pharmacogenetics.

Introduction: Pharmacogenetic markers, such as the ATP Binding Cassette (ABCB1) and cytochrome P450 (CYP) 3A5 enzymes, play a crucial role in personalized medicine by influencing drug efficacy and toxicity based on individuals' or populations' genetic variations.This study aims to investigate the genetic polymorphisms of CYP3A5 (rs776746) and ABCB1 (rs1045642) in the West Algerian population and compare the genotypes and allelic distributions with those of various ethnic groups. Methods: The study involved 472 unrelated healthy subjects from the Western Algerian population. DNA genotyping was performed using TaqMan allelic discrimination assay. The variants in our population were compared to those in other ethnic groups available in the 1000 Genomes Project. Genotype and allele frequencies were calculated using the chi-square test and the Hardy-Weinberg equilibrium (HWE). Results: The minor allele frequencies were found to be 0.21 for CYP3A5 6986A and 0.34 for ABCB1 3435T. These frequencies were similar to those observed in North African populations, while notable differences were observed in comparison to certain Caucasian and African populations. Conclusion: The difference in the allelic and genotypic distribution of these polymorphisms emphasize the need for dose adjustments in drugs metabolized by CYP3A5 and transported by ABCB1 to optimize treatments outcomes.

Differential distribution of ivacaftor and its metabolites in plasma and human airway epithelia.

Ivacaftor is the first clinically approved monotherapy potentiator to treat CFTR channel dysfunction in people with cystic fibrosis. Ivacaftor (Iva) is a critical component for all current modulator therapies, including highly effective modulator therapies. Clinical studies show that CF patients on ivacaftor-containing therapies present various clinical responses, off-target effects, and adverse reactions, which could be related to metabolites of the compound. In this study, we reported the concentrations of Iva and two of its major metabolites (M1-Iva and M6-Iva) in capillary plasma and estimated M1-Iva and M6-Iva metabolic activity via the metabolite parent ratio in capillary plasma over 12 hours. We also used the ratio of capillary plasma versus human nasal epithelial cell concentrations to evaluate entry into epithelial cells in vivo. M6-Iva was rarely detected by LC-MS/MS in epithelial cells from participants taking ivacaftor, although it was detected in plasma. To further explore this discrepancy, we performed in vitro studies, which showed that M1-Iva, but not M6-Iva, readily crossed 16HBE cell membranes. Our studies also suggest that metabolism of these compounds is unlikely to occur in airway epithelia despite evidence of expression of metabolism enzymes. Overall, our data provide evidence that there are differences between capillary and cellular concentrations of these compounds that may inform future studies of clinical response and off-target effects.

The influence of drug-induced metabolic enzyme activity inhibition and CYP3A4 gene polymorphism on aumolertinib metabolism.

The purpose of this study is to clarify the drug interaction profile of aumolertinib, and the influence of CYP3A4 genetic polymorphism on aumolertinib metabolic characteristics. Through microsomal enzyme reactions, we screened 153 drugs and identified 15 that significantly inhibited the metabolism of aumolertinib. Among them, telmisartan and carvedilol exhibited potent inhibitory activities in rat liver microsomes (RLM) and human liver microsomes (HLM). In vivo, the pharmacokinetic parameters of aumolertinib, including AUC and Cmax, were significantly altered when co-administered with carvedilol, with a notable decrease in the clearance rate CLz/F. Interestingly, the pharmacokinetic parameters of the metabolite HAS-719 exhibited a similar trend as aumolertinib when co-administered. Mechanistically, both telmisartan and carvedilol exhibited a mixed-type inhibition on the metabolism of aumolertinib. Additionally, we used a baculovirus-insect cell expression system to prepare 24 recombinant CYP3A4 microsomes and obtained enzymatic kinetic parameters using aumolertinib as a substrate. Enzyme kinetic studies obtained the kinetic parameters of various CYP3A4 variant-mediated metabolism of aumolertinib. Based on the relative clearance rates, CYP3A4.4, 5, 7, 8, 9, 12, 13, 14, 17, 18, 19, 23, 24, 33, and 34 showed significantly lower clearance rates compared to the wild-type. Among the different CYP3A4 variants, the inhibitory potency of telmisartan and carvedilol on the metabolism of aumolertinib also varied. The IC50 values of telmisartan and carvedilol in CYP3A4.1 were 6.68 ± 1.76 µM and 0.60 ± 0.25 µM, respectively, whereas in CYP3A4.12, the IC50 exceeded 100 µM. Finally, we utilized adeno-associated virus to achieve liver-specific high expression of CYP3A4*1 and CYP3A4*12. In the group with high expression of the less active CYP3A4*12, the magnitude of the drug-drug interaction was significantly attenuated. In conclusion, CYP3A4 genetic polymorphism not only influences the pharmacokinetic characteristics of aumolertinib, but also the inhibitory potency of telmisartan and carvedilol on it.

Pharmacogenetics of Carbamazepine: A Systematic Review on CYP3A4 and CYP3A5 Polymorphisms.

BACKGROUND AND OBJECTIVE: The association between carbamazepine (CBZ) metabolism and resistance in epilepsy and the genetic polymorphisms of CYP3A5 (rs776746 and rs15524) and CYP3A4 (rs2242480, rs2740574, rs35599367, rs12721627, and rs28371759) has been the subject of previous investigations with controversial results. We conducted a systematic review to assess the potential link between these polymorphisms and CBZ metabolism and resistance. METHODS: Identifying relevant studies, was carried out bay searching PubMed, Scopus, PharmGKB, EPIGAD, and PHARMAADME databases up until June 2023. The studies included in our analysis investigated the connection between CYP3A5 (rs776746 and rs15524) and CYP3A4 (rs2242480, rs2740574, rs35599367, rs12721627, and rs28371759) polymorphisms and CBZ metabolism and resistance. RESULTS: This review included a total of 23 studies and more than 2177 epilepsy patients. As a result the CYP3A4 (rs12721627 and rs28371759) polymorphisms are associated with reduced catalytic activity, where the CYP3A4 (rs2740574) polymorphism is linked to lower levels of CBZ-diol and decreased activity. It's been found also that the CYP3A5 (rs776746) polymorphism influences the dose-adjusted plasma levels of CBZ. CONCLUSION: Although these findings highlight the impact of genetic variations in the CYP3A4 and CYP3A5 genes on CBZ pharmacokinetics and pharmacodynamics, further studies across diverse populations are essential to enhance personalized epilepsy therapy in clinical settings.

Acneiform Eruptions Possibly Triggered by Clarithromycin During Sirolimus Treatment.

Acneiform eruption is the recognized dermatological side effect of sirolimus, an inhibitor of the mammalian target of rapamycin, although the pathophysiological mechanisms and dose dependency of this side effect remain unclear. This case report describes a case of a 40-year-old Japanese woman treated with systemic sirolimus who developed acneiform eruptions following the administration of clarithromycin. The acneiform eruption resolved after discontinuation of sirolimus and relapsed with the resumption. Since sirolimus and clarithromycin have a potential drug-drug interaction mediated by cytochrome P450 3A (CYP3A), this case suggests that the acneiform eruption developed in association with elevated blood levels of sirolimus. We conclude that clinicians should be aware of the possibility of developing acneiform eruption during sirolimus treatment, especially when administered with medications that inhibit CYP3A.

The Role of Intestinal Cytochrome P450s in Vitamin D Metabolism.

Vitamin D hydroxylation in the liver/kidney results in conversion to its physiologically active form of 1,25-dihydroxyvitamin D3 [1,25(OH)2D3]. 1,25(OH)2D3 controls gene expression through the nuclear vitamin D receptor (VDR) mainly expressed in intestinal epithelial cells. Cytochrome P450 (CYP) 24A1 is a catabolic enzyme expressed in the kidneys. Interestingly, a recently identified mutation in another CYP enzyme, CYP3A4 (gain-of-function), caused type III vitamin D-dependent rickets. CYP3A are also expressed in the intestine, but their hydroxylation activities towards vitamin D substrates are unknown. We evaluated CYP3A or CYP24A1 activities on vitamin D action in cultured cells. In addition, we examined the expression level and regulation of CYP enzymes in intestines from mice. The expression of CYP3A or CYP24A1 significantly reduced 1,25(OH)2D3-VDRE activity. Moreover, in mice, Cyp24a1 mRNA was significantly induced by 1,25(OH)2D3 in the intestine, but a mature form (approximately 55 kDa protein) was also expressed in mitochondria and induced by 1,25(OH)2D3, and this mitochondrial enzyme appears to hydroxylate 25OHD3 to 24,25(OH)2D3. Thus, CYP3A or CYP24A1 could locally attenuate 25OHD3 or 1,25(OH)2D3 action, and we suggest the small intestine is both a vitamin D target tissue, as well as a newly recognized vitamin D-metabolizing tissue.

Interaction of CYP3A4 with the inhibitor cobicistat: Structural and mechanistic insights and comparison with ritonavir.

Cobicistat is a derivative of ritonavir marketed as a pharmacoenhancer for anti-HIV therapy. This study investigated the interaction of cobicistat with the target protein, drug-metabolizing cytochrome P450 3A4 (CYP3A4), at the molecular level using spectral, kinetic, functional, and structural approaches. It was found that, similar to ritonavir, cobicistat directly coordinates to the heme via the thiazole nitrogen but its affinity and the binding rate are 2-fold lower: 0.030 µM and 0.72 s-1, respectively. The newly determined 2.5 Å crystal structure of cobicistat-bound CYP3A4 suggests that these changes arise from the inability of cobicistat to H-bond to the active site S119 and establish multiple stabilizing contacts with the F-F' connecting fragment, which becomes disordered upon steric clashing with the bulky morpholine moiety. Nonetheless, cobicistat inhibits recombinant CYP3A4 as potently as ritonavir (IC50 of 0.24 µM vs 0.22 µM, respectively) due to strong ligation to the heme and formation of extensive hydrophobic/aromatic interactions via the phenyl side-groups. To get insights into the inhibitory mechanism, the K257 residue, known to be solely and irreversibly modified by the reactive ritonavir metabolite, was substituted with alanine. Neither this nor control K266A mutation changed the extent of time-dependent inhibition of CYP3A4 by cobicistat and ritonavir, suggesting the existence of alternative inactivation mechanism(s). More importantly, K257 was found to be functionally important and contributed to CYP3A4 allosterism, possibly by modulating protein-ligand interactions through conformational dynamics.

Applying Physiologically Based Pharmacokinetic Modeling to Interpret Carbamazepine's Nonlinear Pharmacokinetics and Its Induction Potential on Cytochrome P450 3A4 and Cytochrome P450 2C9 Enzymes.

Carbamazepine (CBZ) is commonly prescribed for epilepsy and frequently used in polypharmacy. However, concerns arise regarding its ability to induce the metabolism of other drugs, including itself, potentially leading to the undertreatment of co-administered drugs. Additionally, CBZ exhibits nonlinear pharmacokinetics (PK), but the root causes have not been fully studied. This study aims to investigate the mechanisms behind CBZ's nonlinear PK and its induction potential on CYP3A4 and CYP2C9 enzymes. To achieve this, we developed and validated a physiologically based pharmacokinetic (PBPK) parent-metabolite model of CBZ and its active metabolite Carbamazepine-10,11-epoxide in GastroPlus®. The model was utilized for Drug-Drug Interaction (DDI) prediction with CYP3A4 and CYP2C9 victim drugs and to further explore the underlying mechanisms behind CBZ's nonlinear PK. The model accurately recapitulated CBZ plasma PK. Good DDI performance was demonstrated by the prediction of CBZ DDIs with quinidine, dolutegravir, phenytoin, and tolbutamide; however, with midazolam, the predicted/observed DDI AUClast ratio was 0.49 (slightly outside of the two-fold range). CBZ's nonlinear PK can be attributed to its nonlinear metabolism caused by autoinduction, as well as nonlinear absorption due to poor solubility. In further applications, the model can help understand DDI potential when CBZ serves as a CYP3A4 and CYP2C9 inducer.

Physiologically-Based Pharmacokinetic Modelling to Investigate the Effect of CYP3A4/3A5 Maturation on Tacrolimus Pharmacokinetics in Paediatric HSCT Patients.

Tacrolimus (FK506) is a cornerstone of GVHD-prophylaxis treatment in paediatrics undergoing haematopoietic stem cell transplantation (HSCT). However, due to concerns about highly inter/intra-individual variability, precision dosing of FK506 is crucial. Cytochrome P450(CYP) 3A4 and 3A5 are considered important sources of FK506 pharmacokinetic variability. Nevertheless, the impact of age-related maturation in hepatic and intestinal CYP3A4/3A5 enzymes remains unknown in paediatric HSCT patients. Physiologically-based pharmacokinetic (PBPK) models were developed and verified in adult volunteers and adult HSCT patients using GastroPlusTM (version 9.0), and then extrapolated to paediatric HSCT patients, taking into account the maturation of CYP3A4 and CYP3A5. Default CYP3A4 and CYP3A5 ontogeny profiles were updated based on the latest reports. The paediatric PBPK model was evaluated with independent data collected from Sun Yat-sen Memorial Hospital (86 paediatric HSCT patients, 1 to 16 -year-old). Simulations were performed to evaluate a reported FK506 dosing regimen in infants and children with different CYP3A5 genotypes. Extensive PBPK model validation indicated good predictability, with the predicted/observed (P/O) ratios within the range of 0.80-fold to 1.25-fold. Blood tacrolimus concentration-time curves were comparable between the real and virtual patients. Simulations showed that the higher levels of tacrolimus in 9-month-old to 3-year-old infants were mainly attributed to the CYP3A4/3A5 ontogeny profiles, which resulted in lower clearance and higher exposure relative to dose. The oral dosage of 0.1 mg/kg/day (q12 h) is considered appropriate for paediatric HSCT patients 9 months to 15 years of age with CYP3A5 *1/*1 genotypes. Lower doses were required for paediatric HSCT patients with CYP3A5 *1/*3 (0.08 mg/kg/day, q12h) or CYP3A5 *3/*3 genotypes (0.07 mg/kg/day, q12h), and analyses demonstrated 12.5%-20% decreases in ≤3-year-old patients. The study highlights the feasibility of PBPK modelling to explore age-related enzyme maturation in infants and children(≤3-year-old) undergoing HSCT and emphasizes the need to include hepatic and gut CYP3A4/3A5 maturation parameters.

Tacrolimus Personalized Therapy based on CYP3A5 Genotype in Chinese Patients with Idiopathic Inflammatory Myopathies.

OBJECTIVE: Idiopathic inflammatory myopathies (IIM) are a heterogeneous and life-threatening group of diseases, especially anti-melanoma differentiation-associated gene 5 antibody positive dermatomyositis (MDA5+ DM) is reportedly strongly associated with high mortality rate. Tacrolimus (TAC) provides an excellent therapeutic option, but the trough concentration (Cmin) -outcome relationship remains unexplored. This study was undertaken to identify optimal Cmin and individualized dose based on CYP3A5 genotype for IIM patients. METHODS: 134 IIM patients with 467 Cmin were enrolled. We examined the relationship between TAC Cmin and relapses. The receiver operating characteristic analysis was used to confirm the optimal Cmin. Analyses of factors influencing Cmin were conducted. The dose requirement based on CYP3A5 genotype was confirmed. RESULTS: TAC Cmin is strongly associated with relapses. The optimal cutoff values were 5.30, 5.85, 4.85 and 5.35 ng/ml for acute, subacute, chronic and all phase IIM patients (p = 0.001, 0.013, 0.002, and < 0.001, respectively), as well as 5.35, 5.85, 5.55 and 5.85 ng/ml for acute, subacute, chronic and all phase MDA5+ DM patients (p = 0.007, 0.001, 0.036, and < 0.001, respectively). CYP3A5 genotype was one of the significant factors influencing TAC Cmin. CYP3A5 expressers required 0.059 mg/kg/d to attain the target Cmin, while nonexpressers required 0.046 mg/kg/d (p = 0.019). CONCLUSION: TAC treatment may elicit favorable outcome in patients with IIM and MDA5+ DM when Cmin exceeded 5.35 and 5.85 ng/ml, which is crucial to lower relapse rate. The individualized dose based on the CYP3A5 genotype provides a reference for TAC personalized therapy in IIM.

Efficacy of ivermectin and its metabolites against Plasmodium falciparum liver stages in primary human hepatocytes.

Ivermectin, a broad-spectrum anti-parasitic drug, has been proposed as a novel vector control tool to reduce malaria transmission by mass drug administration. Ivermectin and some metabolites have mosquito-lethal effect, reducing Anopheles mosquito survival. Ivermectin inhibits liver stage development in a rodent malaria model, but no inhibition was observed in a primate malaria model or in a human malaria challenge trial. In the liver, cytochrome P450 3A4 and 3A5 enzymes metabolize ivermectin, which may impact drug efficacy. Thus, understanding ivermectin metabolism and assessing this impact on Plasmodium liver stage development is critical. Using primary human hepatocytes (PHHs), we characterized ivermectin metabolism and evaluated the efficacy of ivermectin and its primary metabolites M1 (3″-O-demethyl ivermectin) and M3 (4-hydroxymethyl ivermectin) against Plasmodium falciparum liver stages. Two different modes of ivermectin exposure were evaluated: prophylactic mode (days 0-3 post-infection) and curative mode (days 3-5 post-infection). We used two different PHH donors and modes to determine the inhibitory concentration (IC50) of ivermectin, M1, M3, and the known anti-malarial drug pyrimethamine, with IC50 values ranging from 1.391 to 14.44, 9.95-23.71, 4.767-8.384, and 0.9073-5.416 µM, respectively. In our PHH model, ivermectin and metabolites M1 and M3 demonstrated inhibitory activity against P. falciparum liver stages in curative treatment mode (days 3-5) and marginal activity in prophylactic treatment mode (days 0-3). Ivermectin had improved efficacy when co-administered with ketoconazole, a specific inhibitor of cytochrome P450 3A4 enzyme. Further studies should be performed to examine ivermectin liver stage efficacy when co-administered with CYP3A4 inhibitors and anti-malarial drugs to understand the pharmacokinetic and pharmacodynamic drug-drug interactions that enhance efficacy against human malaria parasites in vitro.

3D spheroid HepaRG and fluorescent biphasic tracer for CYP3A4-mediated antibiotic interaction monitoring in sepsis.

Cytochrome P450 3A4 (CYP3A4) is a crucial enzyme in the metabolism of xenobiotics, particularly in drug metabolism interactions (DDIs), making it a significant factor in clinical drug use. However, current assay techniques are both laborious and costly, making it difficult to construct a high-throughput monitoring method that can be used in conjunction with the clinic. This poses certain safety hazards for drug combination. Therefore, it is crucial to develop a synchronized monitoring method for the inhibition and induction of CYP3A4. In this study, we utilized 3D culture technology to develop a HepaRG cells spheroid model. The CYP450 and transporter expression, the albumin secretion, and urea synthesis capacity characteristics were analyzed. The NEN probe was utilized as a tracer molecule for CYP3A4. The fluorescence intensity of metabolites was characterized by laser confocal technique to determine the inhibition and expression of CYP3A4 in the HepaRG cell spheroid model by the antibiotics for sepsis. The results indicate that the HepaRG sphere model successfully possessed the physiological phenotype of the liver, which could be used for drug interaction monitoring. Through positive drug testing, NEN probe was able to achieve bidirectional characterization of CYP3A4 induction and inhibition. The monitoring method described in this paper was successfully applied to drug interaction monitoring of commonly used antibiotics in sepsis patients, which is a convenient and rapid monitoring method. The proposed method offers a new strategy for monitoring CYP3A4-mediated drug-drug interactions with a high-throughput assay, which will help to improve the safety of clinical drug combination.