Inhibitory effect of imperatorin on dabrafenib metabolism in vitro and in vivo.

Dabrafenib is a BRAF inhibitor that has been demonstrated to be efficacious in the treatment of melanoma and non-small-cell lung cancer patients with BRAF V600E mutations. The objective of this study was to investigate the effects of 51 traditional Chinese medicines on the metabolism of dabrafenib and to further investigate the inhibitory effect of imperatorin. The quantification of dabrafenib and its metabolite hydroxy-dabrafenib was carried out using a sensitive, rapid, and accurate assay method based on ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS). The results of in vitro experiments showed that 20 drugs inhibited the metabolism of dabrafenib by more than 80 %. In a further study of imperatorin on dabrafenib, the half-maximal inhibitory concentration (IC50) values of imperatorin on dabrafenib were 0.22 µM and 3.68 µM in rat liver microsomes (RLM) and human liver microsomes (HLM), respectively, while the inhibition mechanisms were non-competitive and mixed type inhibition, respectively. The results of in vivo experiments demonstrated that in the presence of imperatorin, the AUC(0-t), AUC(0-∞), Cmax, and Tmax of dabrafenib were increased by 2.38-, 2.26-, 1.05-, and 6.10-fold, respectively, while CLz/F was decreased by 67.9 %. In addition, Tmax of hydroxy-dabrafenib was increased by 1.4-fold. The results of the research showed that imperatorin had a consistent inhibitory effect on dabrafenib in vitro and in vivo. When the concurrent use of dabrafenib and imperatorin is unavoidable, clinicians should closely monitor for potential adverse events and make timely adjustments to the administered dosage.

Functional Assessment of CYP3A4 and CYP2C19 Genetic Polymorphisms on the Metabolism of Clothianidin In vitro.

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.

Effects of the antitumor drugs adagrasib and asciminib on apixaban metabolism in vitro and in vivo.

Apixaban is an oral anticoagulant that directly inhibits the target Factor Xa (FXa). In this study, we focused on the in vivo and in vitro effects of adagrasib and asciminib on apixaban metabolism, to discover potential drug-drug interactions (DDI) and explore their inhibitory mechanisms. The levels of apixaban and its metabolite, O-desmethyl-apixaban (M2), were determined by ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS). In vitro evaluation, the maximum half inhibitory concentration (IC50) of adagrasib in rat liver microsomes (RLM) and human liver microsomes (HLM) against apixaban was 7.99 µM and 117.40 µM, respectively. The IC50 value of asciminib against apixaban in RLM and HLM was 4.28 µM and 18.42 µM, respectively. The results of the analysis on inhibition mechanisms showed that adagrasib inhibited the metabolism of apixaban through a non-competitive mechanism, while asciminib inhibited the metabolism of apixaban through a mixed mechanism. Moreover, the interaction of apixaban with adagrasib and asciminib in Sprague-Dawley (SD) rats was also investigated. It was found that the pharmacokinetic characteristics of apixaban were significantly changed when combined with these two antitumor drugs, where AUC(0-t), AUC(0-∞), t1/2, Tmax, and Cmax were increased, while CLz/F was significantly decreased. But both drugs did not appear to affect the metabolism of M2 in a significant way. Consistent results from in vitro and in vivo demonstrated that both adagrasib and asciminib inhibited the metabolism of apixaban. It provided reference data for the future clinical individualization of apixaban.

Simultaneous measurement of COVID-19 treatment drugs (nirmatrelvir and ritonavir) in rat plasma by UPLC-MS/MS and its application to a pharmacokinetic study.

PAXLOVID™ (Co-packaging of Nirmatrelvir with Ritonavir) has been approved for the treatment of Coronavirus Disease 2019 (COVID-19). The goal of the experiment was to create an accurate and straightforward analytical method using ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) to simultaneously quantify nirmatrelvir and ritonavir in rat plasma, and to investigate the pharmacokinetic profiles of these drugs in rats. After protein precipitation using acetonitrile, nirmatrelvir, ritonavir, and the internal standard (IS) lopinavir were separated using ultra performance liquid chromatography (UPLC). This separation was achieved with a mobile phase composed of acetonitrile and an aqueous solution of 0.1% formic acid, using a reversed-phase column with a binary gradient elution. Using multiple reaction monitoring (MRM) technology, the analytes were detected in the positive electrospray ionization mode. Favorable linearity was observed in the calibration range of 2.0-10000 ng/mL for nirmatrelvir and 1.0-5000 ng/mL for ritonavir, respectively, within plasma samples. The lower limits of quantification (LLOQ) attained were 2.0 ng/mL for nirmatrelvir and 1.0 ng/mL for ritonavir, respectively. Both drugs demonstrated inter-day and intra-day precision below 15%, with accuracies ranging from -7.6% to 13.2%. Analytes were extracted with recoveries higher than 90.7% and without significant matrix effects. Likewise, the stability was found to meet the requirements of the analytical method under different conditions. This UPLC-MS/MS method, characterized by enabling accurate and precise quantification of nirmatrelvir and ritonavir in plasma, was effectively utilized for in vivo pharmacokinetic studies in rats.

Quantitative investigation of drug-drug interaction between bergenin and vilazodone in rats through UPLC-MS/MS assay.

In this study, we firstly established and verified a method by ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) for the analysis of vilazodone and its metabolite M10 in rat plasma, then this method was used to explore the pharmacokinetics of vilazodone and M10 present or absence of 80 mg/kg bergenin in rats. Protein precipitation with acetonitrile was used to prepare the samples in this research. The mobile phase for liquid chromatography was consisted of 0.1% formic acid aqueous solution and acetonitrile. Brexpiprazole was used as the internal standard (IS), and the multiple reaction monitoring (MRM) mode was used for detection. The verification items required by the US Food and Drug Administration (FDA) guidelines such as selectivity, sensitivity, linearity, stability, recovery and matrix effect of this method were all met the standards. Besides, rats were used to explore the drug-drug interaction between vilazodone and bergenin, which were divided into two groups, and separately gavaged with the same-volume of carboxymethyl cellulose sodium (CMC-Na) solution and 80 mg/kg bergenin, respectively. The results showed that bergenin significantly affected the metabolism of vilazodone. It suggested that there was a potential drug-drug interaction between bergenin and vilazodone in rats. In clinical application, we should pay attention to the dose of vilazodone when in combination with bergenin.

Simultaneous determination of iguratimod and its metabolite in rat plasma using a UPLC-MS/MS method: Application for drug-drug interaction.

This aim of the work was to establish an acceptable sensitive assay based on ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) for quantitatively analyzing the plasma concentrations of iguratimod (IGR) and its metabolite M2 in rats, and to further investigate the effect of fluconazole on the pharmacokinetics of IGR and M2. The mobile phase consisted of acetonitrile and water with 0.1% formic acid, was used to separate IGR, M2 and internal standard (IS) fedratinib on a UPLC BEH C18 column (2.1 mm × 50 mm, 1.7 µm) with the flow rate of 0.4 mL/min. Positive ion mode and multiple reaction monitoring (MRM) were used to construct the quantitative analysis. The calibration standard of IGR and M2 covered 2-10000 and 1-1000 ng/mL respectively, with the lower limit of quantification (LLOQ) as 2 ng/mL and 1 ng/mL respectively. In addition, selectivity, recovery, accuracy, precision, matrix effect and stability of the method validation program were well accepted in this work. Subsequently, this approach was used to assess the effect of fluconazole on the pharmacokinetics of IGR and M2 in rats. In the presence of 20 mg/kg fluconazole (experimental group), we found the main pharmacokinetic parameters were significantly altered when compared with 2.5 mg/kg IGR alone (control group). Among them, AUC(0-∞) and Cmax of IGR in the experimental group was 1.43 and 1.08 times higher than that of the control group, respectively. Moreover, we also found that the other main pharmacokinetic parameters of M2 had no significant changes, except t1/2z and Tmax. In conclusion, fluconazole significantly altered the main pharmacokinetics of IGR and M2 in rats. It implys that we should pay more attention to the adverse reaction of IGR when the concomitant use of fluconazole and IGR occur in the future clinical practice.

Development of an UPLC-MS/MS method for quantitative analysis of abexinostat levels in rat plasma and application of pharmacokinetics.

Broad-spectrum histone deacetylase inhibitors (HDACi) have excellent anti-tumor effects, such as abexinostat, which was a novel oral HDACi that was widely used in clinical treatment. The purpose of this study was to establish a rapid and reliable method for the detection of abexinostat concentrations in rat plasma using ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS). The mobile phase we used was acetonitrile and 0.1% formic acid, and the internal standard (IS) was givinostat. Selective reaction monitoring (SRM) was used for detection with ion transitions at m/z 397.93 → 200.19 for abexinostat and m/z 422.01 → 186.11 for givinostat, respectively. The intra-day and inter-day precision of abexinostat were less than 11.5% and the intra-day and inter-day accuracy ranged from - 10.7% to 9.7% using this method. During the analysis process, the stability of the test sample was reliable. In addition, the recovery and matrix effects of this method were within acceptable limits. Finally, the method presented in this paper enabled accurate and quick determination of abexinostat levels in rat plasma from the pharmacokinetic study following gavage at a dose of 8.0 mg/kg abexinostat.