CYP3A4 Research Articles

Evaluation of the effects of atazanavir-ritonavir on the pharmacokinetics of lumefantrine in patients living with HIV in Lagos University Teaching Hospital, South-Western Nigeria.

Atazanavir-ritonavir (ATVr)-based antiretroviral therapy and artemether-lumefantrine (AL) are commonly used drugs for the treatment of human immune deficiency virus (HIV) infection and malaria respectively. However, interaction of both drugs, with Cytochrome P 3A4 (CYP 3A4) isoenzyme, may spawn clinically significant pharmacokinetic interactions. This study evaluated the effects of atazanavir-ritonavir on the pharmacokinetics of lumefantrine. In a case-control study, twenty participants having Plasmodium falciparum malaria were recruited and divided into two groups (ATVr-arm, n=10; and control-arm, n= 10). All the participants were administered six oral doses of AL 80-480 mg (Coartem). Thereafter, their blood samples were collected at different time intervals over seven days. The concentration of lumefantrine in each sample was quantified with high-performance liquid chromatography (HPLC) and used to determine its pharmacokinetic parameters which were compared between the test and control groups. ATVr increased the mean day 7 concentration of lumefantrine (ATVr 3847.09 ± 893.35 ng/mL, control 1374.53 ± 265.55 ng/mL, p = 0.016) and the area under its plasma concentration-time curve (ATVr 670529.57 ± 157172.93 ng.h/mL, control 447976.28 ± 80886.99 ng.h/mL, p = 0.224) by 179.88 % and 49.68 %, respectively, but decreased its mean maximum plasma drug concentration (Cmax) (ATVr 13725.70 ± 2658.44 ng/mL, control 15380.48 ± 2332.62 ng/mL, p = 0.645) by 10.76 %. ATVr increased drug exposure and day 7 plasma concentration of lumefantrine. AL is therefore considered effective for the treatment of malaria in patients taking ATVr-based regimen. However, the safety associated with the interaction requires further elucidation. Clin ClinicalTrials.gov Identifier: NCT04531072, August 27, 2020. "Retrospectively registered".

Effects of Shengmai San on key enzymes involved in hepatic and intestinal drug metabolism in rats

Ethnopharmacological relevanceShengmai San (SMS) has been commonly used as a traditional Chinese medicine for the treatment of cardiovascular disorders, of which drug interactions need to be assessed for the safety concern. There is little evidence for the alterations of hepatic and intestinal drug-metabolizing enzymes after repeated SMS treatments to assess drug interactions. Aim of the studyThe studies aim to illustrate the effects of repeated treatments with SMS on cytochrome P450s (CYPs), reduced nicotinamide adenine dinucleotide (phosphate)-quinone oxidoreductase (NQO), uridine diphosphate-glucuronosyltransferase (UGT), and glutathione S-transferase (GST) using in vivo rat model. Materials and methodsThe SMS was prepared using Schisandrae Fructus, Ginseng Radix, and Ophiopogonis Radix (OR) (1:2:2). Chromatographic analyses of decoctions were performed using ultra-performance liquid chromatography (UPLC) and LC-mass spectrometry. Sprague-Dawley rats were orally treated with the SMS and its component herbal decoctions for 2 or 3 weeks. Hepatic and intestinal enzyme activities were determined. CYP3A expression and the kinetics of intestinal nifedipine oxidation (NFO, a CYP3A marker reaction) were determined. ResultsSchisandrol A, schisandrin B, ginsenoside Rb1 and ophiopogonin D were identified in SMS. SMS selectively suppressed intestinal, but not hepatic, NFO activity in a dose- and time-dependent manner. Hepatic and intestinal UGT, NQO and GST activities were not affected. A 3-week SMS treatment decreased the maximal velocity of intestinal NFO by 50%, while the CYP3A protein level remained unchanged. Among SMS component herbs, the decoction of OR decreased intestinal NFO activity. ConclusionsThese findings demonstrate that 3-week treatment with SMS and OR suppress intestinal, but not hepatic CYP3A function. It suggested that the potential interactions of SMS with CYP 3A drug substrates should be noticed, especially the drugs whose bioavailability depends heavily on intestinal CYP3A.

Characterization of cytochrome P450s (CYP)-overexpressing HepG2 cells for assessing drug and chemical-induced liver toxicity

Hepatic metabolism catalyzed by the cytochrome P450 (CYP) superfamily affects liver toxicity associated with exposures to natural compounds and xenobiotic agents. Previously we generated a battery of HepG2-derived stable cell lines that individually express 14 CYPs (1A1, 1A2, 1B1, 2A6, 2B6, 2C8, 2C9, 2C18, 2C19, 2D6, 2E1, 3A4, 3A5, and 3A7). In this study, we comprehensively characterized each cell line for its CYP expression and enzyme activity. Specifically, we measured the mRNA expression, protein expression, and metabolite formation. Using CYP3A4, 2D6, and 2C9-overexpressing cells as representatives, we examined the stability of these cells in long-term cultures for up to 10 passages. The results showed that CYPs can be stably overexpressed for up to 10 cell culture passages without losing their activities. The robustness of responses to stimuli among the cells at different passages was also investigated in CYP3A4-overexpressing cells and the response to amiodarone and dronedarone showed no difference between the cells at the passage 2 and 10. Moreover, the mRNA expression level of most CYPs was higher in CYP-overexpressing HepG2 cells than that in HepaRG cells and primary human hepatocytes. This study confirmed the stability of CYP-overexpressing HepG2 cell lines and provided useful information for a broader use of these cells in pharmacologic and toxicologic research.

Elucidation of plasma protein binding, blood partitioning, permeability, CYP phenotyping and CYP inhibition studies of Withanone using validated UPLC method: An active constituent of neuroprotective herb Ashwagandha

Ethnopharmacological relevanceWithanone (WN), an active constituent of Withania somnifera commonly called Ashwagandha has remarkable pharmacological responses along with neurological activities. However, for a better understanding of the pharmacokinetic and pharmacodynamic behavior of WN, a comprehensive in-vitro ADME (absorption, distribution, metabolism, and excretion) studies are necessary. Aim of the studyA precise, accurate, and sensitive reverse-phase ultra-performance liquid chromatographic method of WN was developed and validated in rat plasma for the first time. The developed method was successfully applied to the in-vitro ADME investigation of WN. Material and methodsThe passive permeability of WN was assayed using PAMPA plates and the plasma protein binding (PPB) was performed using the equilibrium dialysis method. Pooled liver microsomes of rat (RLM) and human (HLM) were used for the microsomal stability, CYP phenotyping, and inhibition studies. CYP phenotyping was evaluated using the specific inhibitors. CYP inhibition study was performed using specific probe substrates along with WN or specific inhibitors. ResultsWN was found to be stable in the simulated gastric and intestinal environment and has a high passive permeability at pH 4.0 and 7.0 in PAMPA assay. The PPB of WN at 5 and 20 μg/mL concentrations were found to be high i.e. 82.01 ± 1.44 and 88.02 ± 1.15%, respectively. The in vitro half-life of WN in RLM and HLM was found to be 59.63 ± 2.50 and 68.42 ± 2.19 min, respectively. CYP phenotyping results showed that WN was extensively metabolized by CYP 3A4 and1A2 enzymes in RLM and HLM. However, the results of CYP Inhibition studies showed that none of the CYP isoenzymes were potentially inhibited by WN in RLM and HLM. ConclusionThe in vitro results of pH-dependent stability, plasma stability, permeability, PPB, blood partitioning, microsomal stability, CYP phenotyping, and CYP inhibition studies demonstrated that WN could be a better phytochemical for neurological disorders.

Identification of CYP 2A6 inhibitors in an effort to mitigate the harmful effects of the phytochemical nicotine.

Aim:In this study, our goal was to study the inhibition of nicotine metabolism by P450 2A6, as a means for reduction in tobacco use and consequently the prevention of smoking-related cancers. Nicotine, a phytochemical, is an addictive stimulant, responsible for the tobacco-dependence in smokers. Many of the other phytochemicals in tobacco, including polycyclic aromatic hydrocarbons, N-nitrosamines, and aromatic amines, are potent systemic carcinogens. Tobacco smoking causes about one of every five deaths in the United States annually. Nicotine plasma concentration is maintained by the smokers’ smoking behavior within a small range. Nicotine is metabolized by cytochrome P450s 2A6 and 2A13 to cotinine. This metabolism causes a decrease in nicotine plasma levels, which in turn leads to increased tobacco smoking, and increased exposure to the tobacco carcinogens.Methods:Using the phytochemical nicotine as a lead structure, and taking its interactions with the P450 2A6 binding pocket into consideration, new pyridine derivatives were designed and synthesized as potential selective mechanism-based inhibitors for this enzyme.Results:The design and synthesis of two series of novel pyridine-based compounds, with varying substituents and substitution locations on the pyridine ring, as well as their inhibitory activities on cytochrome P450 2A6 and their interactions with its active site are discussed here. Substitutions at position 3 of the pyridine ring with an imidazole or propargyl ether containing group showed the most optimal interactions with the P4502A6 active site.Conclusion:The pyridine compounds with an imidazole or propargyl ether containing substituent on position 3 were found to be promising lead compounds for further development. Hydrogen-bonding interactions were determined to be crucial for effective binding of these molecules within the P450 2A6 active site.

Cytochrome P450 content in primary tumors and liver metastases of patients with metastatic colorectal cancer.

To determine the content of low-spin form of cytochrome P450in primary tumors and liver metastases of the patients with metastatic colorectal cancer (mCRC) and and assess its prognostic significance. The levels of the oxidized and low-spin forms of cytochrome P450in the tissues of patients with mCRC were studied by electron paramagnetic resonance. To detect CYP1A2and CYP1B1isoforms, Western blot analysis was used. The activity of metalloprotreinases was studied by gelatine zymography. In the liver metastases and tissues adjacent to metastasis, the levels of low-spin forms of cytochrome P450 are lower than in the samples of conventionally normal liver tissue. Western blot analysis revealed that low-spin form of cytochrome P450in primary tumors and liver metastases detected by electron paramagnetic resonance is attributed largely to CYP1B1isoform. The content of low-spin form of cytochrome P450inversely correlated with the the activity of gelatinases (matrix metalloproteinase-2 and -9). The survival of patients with high levels of the low-spin form of cytochrome P450in tumor tissues was higher than that of patients with low levels of the enzyme (105months vs 61months). In the primary tumors and liver metastases of patients with mCRC, the content of the low-spin form of cytochrome P450decreased, which correlated inversely with patient's survival.

Mugil liza (Valenciennes, 1836) biomarker responses in a hypersaline lagoon in southeastern Brazil before and after a microalgal bloom

Araruama lagoon is one of the largest permanent hypersaline lagoons in the world and has been impacted for twenty years by sewage dumping. In early 2005, a Prasinophyceae (Pseudoscourfieldia sp.) phytoplankton bloom occurred, leading to fish mortality during subsequent summers (2006 and 2007). In this context, the aim of the present study was to accompany this historical period through the use of biochemical biomarkers in mullet (Mugil liza) during and after the algal bloom. Mullet were collected from Araruama Lagoon in June and July 2005 and August 2007, as well as from a reference lagoon with no algal blooms, Saquarema Lagoon. Several enzymatic activities in liver were analyzed, namely glutathione peroxidase (GPX), catalase (CAT), total glutathione S-transferase (GSTt), specific GSTπ (pi) and GSTμ (mi) classes and ethoxyresorufin-O-deethylase (EROD), as well as muscle acetylcholinesterase activity (AChE). In 2005, only EROD activity was significantly increased at Araruama Lagoon, indicating induction of the biotransformation enzyme system (CYP 1A1). GSTπ, on the other hand, was lower in the Araruama Lagoon, especially in 2005, when compared to mullet from Saquarema lagoon (reference area). These results highlight the importance of developing actions that go beyond the physical-chemical monitoring of these environments, since the study of ecological relationships and the physiology of organisms affected by algal blooms also include biochemical parameters to evaluate new approaches to icthyotoxicity.

In vitro investigation of metabolic fate of \u03b1-mangostin and gartanin via skin permeation by LC-MS/MS and in silico evaluation of the metabolites by ADMET predictor\u2122

BackgroundMangosteen, Garciniam angostana L., is a juicy fruit commonly found in Thailand. The rinds of Garciniam angostana L.have been used as a traditional medicine for the treatment of trauma, diarrhea and skin infection. It is also used in dermatological product such as in cosmetics. The mangosteen pericarp can be used to extract valuable bioactive xanthone compounds such as α-mangostin and gartanin. This study is aimed to predict the metabolism of α-mangostin and gartanin using in silico and in vitro skin permeation strategies.MethodsBased on their 2D molecular structures, metabolites of those compounds were predicted in silico using ADMET Predictor™. The Km and Vmax, for 5 important recombinant CYP isozymes 1A2, 2C9, 2C19, 2D6 and 3A4 were predicted. Moreover, the in vitro investigation of metabolites produced during skin permeation using human epidermal keratinocyte cells, neonatal (HEKn cells) was performed by LC-MS/MS.ResultsIt was found that the results derived from in silico were in excellent alignment with those obtained from in vitro studies for both compounds. The prediction referred that gartanin and α-mangostin were the substrate of CYP1A2, 2C9, 2C19 and 3A. In the investigation of α-mangostin metabolites by LC-MS/MS system, the MW of the parent compound was increased from 411.200 to 459.185 Da. Therefore, α-mangostin might be metabolized via tri-oxidation process. The increased molecular weight of parent compound (397.200 to 477.157 Da) illustrated that gartanin might be conjugated to sulfated derivatives.ConclusionsIn all the studies, α-mangostin and gartanin were predicted to be.metabolized via phase I and phase II metabolism (sulfation), respectively.

Abstract 15740: Concomitant Use of Dronedarone and Direct Oral Anticoagulants and Risk of Bleeding in Patients With Atrial Fibrillation: An Analysis of the U.S. Truven Health MarketScan Database

Introduction: Dronedarone (DR), a P-gp and CYP 3A4 inhibitor may increase exposure and the risk of bleeding when combined with direct oral anticoagulants (DOACs). Objective: To examine the association between concomitant use of DR and the DOACs, apixaban (A), dabigatran (D), and rivaroxaban (R), and risk of bleeding compared to DOAC monotherapy in patients with atrial fibrillation (AF). Methods: A retrospective cohort study using a U.S. claims database, Truven Health MarketScan identified new users of A, D, and R in patients with AF ≥18 years from Jan 1, 2007 to Sep 30, 2017. Bleeding was defined as hospitalization or emergency room visit with a primary diagnosis of gastrointestinal (GI) bleeding, intracranial hemorrhage (ICH), or bleeding at other sites. Risk of overall and by type of bleeding was examined in concomitant users of DOAC and DR compared to patients using DOAC alone after adjusting for covariates of interest and applying propensity score (PS) trimming via Cox proportional hazards modeling. Results: Among concomitant users of DR and A (1,932), D (3,117), and R (2,395), crude incidence rates of bleeding per 1,000 person-years were 17.2, 37.8, 61.8, respectively versus 26.8, 31.3, and 44.9 in users of A (51,420), D (42,312), and R (57,300) alone. Incidence rates stratified by PS showed higher bleeding incidence in concomitant users of DR with D or R, but not with A. No increased bleeding risk was associated with use of DR and A vs A alone [Adjusted Hazard ratio (aHR): 0.69 (95% CI: 0.40, 1.17), p=0.16]. A modestly increased risk of GI bleeding but not overall bleeding was associated with combined use of DR and D vs D alone [aHR bleeding: 1.18 (95% CI: 0.89, 1.56), p=0.26; aHR GI bleeding: 1.40 (95% CI: 1.01, 1.93); p=0.04]. An increased risk of overall bleeding, driven by GI bleeding, was associated with combined use of DR and R vs R alone [aHR bleeding:1.31 (95% CI: 1.01, 1.69); p=0.04; aHR GI bleeding:1.39 (95% CI: 0.98, 1.95); p=0.06]. There was no increase in the risk of ICH associated with combined use of DR and any DOAC. Conclusions: Concomitant treatment with DR and A showed no increased risk of bleeding, but DR increased the risk of GI bleeding when given with D or R, and of overall bleeding only with R. Concomitant treatment with DR and any DOAC did not increase ICH risk.

Serum Organochlorine Pesticides Level CYP1A1 Genetic Polymorphism & Risk of Prostate Cancer Cases Control Study

Background: Agriculture is the backbone of the economic and social development in the Sudan. 80% of the population depends for their living on agriculture. Organochlorines (OC's) were the first to be used against cotton pests, and control termites in sugarcane. In cancer CYP1A1 have dealt primarily with the metabolic activation of pro-carcinogens including polycyclic aromatic hydrocarbons and estradiol. Aim: The aim of this study is to determine the serum Organochlorine pesticides and PCBs (OC's) level, Cytochrome (CYP) 1A1 genetic polymorphisms and risk for Sudanese prostate cancer patients together with control groups, attending the Wad-Madani teaching hospital, Gezira State- Central Sudan. Method: 100 prostatic cancer patients and 100 controls (over 50 years old) will be studied. A questionnaire will be designed and Clinical examination will be performed including weight and heights. From venous blood (2.5ml) Serum will be separated and store at -80 ОC until analyses. The Organochlorine pesticides residues in serum will be measured by using a modified QuEChERS method by GC/MS. Soil, water and common food samples samples will be collected and storage until the analysis. For CYP1A1 genotyping examination, 2.5 ml of whole blood will be collected in 5 ml EDTA vacutainer tube. Buffy coat separate from plasma and store at -80 ОC until DNA extraction. DNA extraction for blood will be done by QIAamp DNA Mini Kits. PCR Technique will be used for laboratory genetic analysis by using Genotypes Restriction Enzymes Length Polymorphisms (RFLP). Estrogen (E1) level will be analyzed by Elisa technique with Elisa Kits. The study will help to examine one of the most important risk factors for prostate cancer in Sudan (Organochlorine & PCBs pesticides). Find the mutations for Cytochrome P-450 CYP (1A1) genotypes and the relationship between Estrogen (E1) levels and other risk factors.

Effect of Nigella sativa oil and ciprofloxacin against bacterial infection on gene expression in Nile tilapia (Oreochromis niloticus) blood

The effect of both Nigella sativa oil (NSO) and ciprofloxacin (Cip) was investigated by evaluating their therapeutic and immunity properties on serum biochemical parameters, and the gene expression levels of hepcidin, interleukin 1β (IL-1β) and cytochrome P450 1A (CYP 1A) in O. niloticus blood against the bacterial infection and the search for antibiotic substitutes. Twelve experimental groups of Nile tilapia were included: control groups (T1–4), Aeromonas hydrophila (Ah) groups (T5–8) and Pseudomonas fluorescens (Ps) groups (T9–12). The antibiogram test showed that the Ah and Ps groups had a high sensitivity to ciprofloxacin, gatifloxacin, lomefloxacin, norfloxacin, gentamicin, and kanamycin. The mortality rate was 43.75% and 37.5% in untreated Ah (T5) and Ps (T9) groups, respectively, but was reduced by the use of NSO and Cip. The fish infected by Ps (T9-T12) showed a significant increase in serum total protein, albumin, and globulin levels, compared to the uninfected control group (T1). A significant increase in the relative hepcidin expression was observed in the Ah infected groups than in the Ps infected groups. Hepcidin transcripts were increased in each of Cip and NSO in the infected groups. The relative expression of the IL-1β gene was significantly increased in the Cip-treated groups, but it decreased in the NSO-treated groups after the bacterial infection, compared to the untreated infected groups. Both Cip and NSO led to a decrease in the level of CYP1A gene expression in the infected fish groups. The study showed that NSO has a therapeutic effect against pathogens just as ciprofloxacin.

Paclitaxel Nanoparticles Induce Apoptosis and Regulate TXR1, CYP3A4 and CYP2C8 in Breast Cancer and Hepatoma Cells

Although the anticancer potentials of water-insoluble drugs are improved by nanoformulation, other intervening factors may contribute in the drug efficacy. This work was designated to explore the effect of paclitaxel-loaded Poly(Lactic-co-Glycolic Acid) (PLGA) nanoparticles on the viability of cancer cells, the expression of Taxol Resistance gene I (TXR1) and paclitaxel metabolizing genes. Paclitaxel loaded PLGA Nanoparticles (PTX-NPs) were prepared, physically characterized and used in the treatment of breast adenocarcinoma cells (MCF-7) and hepatoma cells (HepG2). Cells viability and apoptosis were investigated. In parallel, RNA was isolated, reverse transcribed and used to monitor the expression levels of TXR1, CYP 3A4 and CYP2C8 genes. PTX-NPs were characterized by transmission electron microscopy to be of a nano-size sphere-like shape. FTIR analysis revealed good coupling between PTX and PLGA. The encapsulation efficiency was 99% and the drug release demonstrated a progressive releasing phase followed by slower and sustained releasing phases. Although HepG2 cells demonstrated more resistance to PTX than MCF-7 cells, both cell types were more responsive to PTX-NPS compared to PTX. The IC50 values decreased from 19.3 to 6.7 in breast cancer cells and from 42.5 to 13.1μg/ml in hepatoma cells. The apoptosis was the key mechanism in both cells, where at least 44% of cells underwent apoptosis. The expression of TXR1 decreased when either cells were treated with PTX-NPs, respectively, meanwhile the expressions of CYP3A4 and CYP2C8 were increased. Taken together, this in vitro study reports the associations between the enhanced responsiveness of MCF-7 and HepG2 cells to PLGA-loaded paclitaxel nanoparticles and the accompanying decrease in the cells resistance to the PTX and its enhanced metabolism.

Cardiac Arrest Associated With Combination Cisapride and Itraconazole Therapy.

We report a case of cardiac arrest associated with cisapride in combination with itraconazole and provide a brief review of pertinent literature. Cisapride (Propulsid; Janssen Pharmaceuticals, Titusville, NJ), a gastrointestinal prokinetic drug, has recently been reported to prolong the QT interval. Itraconazole, an oral antifungal agent, is an inhibitor of cytochrome P450 (CYP3A4) metabolism and may elevate serum drug levels of compounds metabolized by this pathway. A 31-year-old woman had a witnessed cardiac arrest while taking the combination of cisapride and itraconazole. Following resucitation, the prolonged QT interval returned to normal after withdrawal of both agents. Echocardiography and cardiac catheterization were within normal limits; electrophysiologic testing failed to induce ventricular tachycardia/ventricular fibrillation. She has had no documented arrhythmias since the arrest. This combination can now be added to a growing list of drugs that may cause torsades de pointes and sudden cardiac death.

Lurasidone – pharmacodynamic and pharmacokinetic properties, clinical potential and interaction risk

Lurasidone is a novel second-generation antipsychotic approved for the treatment of schizophrenia and bipolar depression in adults. It displays high affinity for D2 and 5-HT2A and 5-HT7 receptors, moderate affinity for 5-HT1A and α2C-noradrenergic receptors, and negligible affinity for histamine H1 and muscarinic M1 receptors. It acts as potent D2, 5-HT2A and 5-HT7 antagonist and partial 5-HT1A agonist. Lurasidone taken orally is rapidly absorbed with the time to maximum concentration of 1-3 hours. Lurasidone should be taken with food (at least 350 kcal) due to limited absorption. The mean elimination half-life of lurasidone is 18.1–25.5 hours for doses 20–80 mg/day and 28.8–37.4 hours for doses of 120–160 mg/day. Steady-state is reached within 7 days. The drug is metabolised via CYP 3A4 and excreted mainly in faeces (67–80%) and with urine (about 8–19%). The use of lurasidone with strong inhibitors or inducers of CYP 3A4 (e.g. ketoconazole, erythromycin, or carbamazepine, respectively) is contraindicated. In the case of combined treatment of lurasidone and moderate inhibitors of CYP 3A4, the dose of lurasidone should be decreased to 40 mg/day. Lurasidone is an inhibitor of P-glycoprotein and could increase the level of digoxin and potentate the side effects risk of this drug. Pomelo, grapefruit, or a large amount of oranges should be avoided in the diet during treatment with lurasidone. Pharmacodynamic properties of lurasidone underlie its antipsychotic, antidepressant, precognitive, and sleep-awake rhythm normalising activity.

Abstract 1770: A new DOT1L inhibitor with in vivo activity in mouse models of MLL-translocated leukemia

Abstract Rearrangements in the mixed lineage leukemia (MLL) gene define a distinct, aggressive form of acute leukemia with poor prognosis. The MLL gene encodes for a SET domain histone methyl transferase that catalyzes the methylation of histone 3 lysine 4 (H3K4) at specific gene loci, thus regulating transcription of developmental genes including HOX genes. In disease-linked translocations, the catalytic SET domain is lost and the remaining part fused to a variety of partners, e.g. AF4, AF9, AF10 and ENL. These MLL fusion proteins directly interact with disruptor of telomeric silencing 1-like protein (DOT1L), the only known H3K79 methyltransferase. The H3K79me2 mark is broadly associated with active transcription. Mislocated enzymatic activity of DOT1L causes local H3K79 hypermethylation, misexpression of leukomogenic genes, e.g. HOXA9 and MEIS1, and the aberrant maintenance of a stem cell-like state. Current treatment options of MLL are limited to chemotherapy and allogeneic hematopoietic stem cell transplantation, and outcomes remain poor. The first and only clinical DOT1L inhibitor Pinometostat (EPZ-5676), an S-Adenosyl Methionine (SAM) competitive inhibitor, showed only modest activity and emerging resistance in adult acute leukemia. Pinometostat is no oral drug, but requires administration as continuous infusion to achieve sufficient exposure and sustained target inhibition. So far, there is no approved DOT1L inhibitor and the need remains to develop effective DOT1L inhibitors. We report the identification of new SAM-competitive, structurally SAM-unrelated DOT1L inhibitors with subnanomolar biochemical potency. Compounds with nanomolar cellular activity and good exposure in mouse were tested in tumor xenograft models. Compounds 8 and 9 showed excellent blood exposure after a single dose. However, repeated dosing led to reduced exposure due to cytochrome P450 (Cyp450) 3A4 induction, insufficient pharmacodynamics (PD) and lack of efficacy. Further modification resulted in compound 10 that could be administered orally and was stable upon repeated dosing. A 300 mg/kg dose covered efficacious blood exposure for 24 h, but was not tolerated by tumor-bearing mice. Unfortunately, a 6-fold reduced dose did not achieve sufficient PD modulation and efficacy. Additional activities led to compound 11, which lost its oral bioavailability and had to be administered subcutaneously. Nevertheless, compound 11 achieved tumor growth inhibition in the MV4-11 and Molm-13 xenograft models in mice. In conclusion, we progressed in making DOT1L inhibitors with improved PK properties, but further optimization is required to generate a viable clinical candidate. The limited efficacy obtained with compound 11 and lack of efficacy observed with subcutaneous administration of EPZ-5676 around the maximally tolerated dose illustrate the difficulty to achieve a sustained level of DOT1L inhibitor in vivo to suppress DOT1L activity sufficiently. Furthermore, H3K79me2 and efficacy in vivo seem disconnected. Citation Format: Andreas Weiss, Frederic Stauffer, Clemens Clemens, Henrik Möbitz, Christian Ragot, Kim S. Beyer, Keith Calkins, Claudio Thoma, Daniel Guthy, Michael Kiffe, Bernard Van Eerdenbrugh, William R. Sellers, Francesco Hofmann, Ralph Tiedt, Christoph Gaul. A new DOT1L inhibitor with in vivo activity in mouse models of MLL-translocated leukemia [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1770.

Lack of Smoking Effects on Pharmacokinetics of Oral Paliperidone-analysis of a Naturalistic Therapeutic Drug Monitoring Sample.

Major smoking effects have been reported for a series of psychotropic agents, mainly including substrates of CYP450 1A2, although smoking may also affect alternative metabolic pathways. To our knowledge, smoking effects on paliperidone pharmacokinetics have not been assessed yet. We compared plasma concentrations of paliperidone as well as dose-corrected-plasma concentrations (C/D) from a naturalistic database between smokers and nonsmokers using nonparametrical tests, such as the Mann-Whitney U-test (MWU). Additionally, we compared light and heavy smokers with nonsmokers separately. Comparing 55 smokers with 37 nonsmokers treated with oral paliperidone, no differences in the percentage of females, age, body weight, body mass index, and daily paliperidone dose were reported (p=0.709 for χ2, p=0.26, p=0.38, p=0.67, and p=0.8 for MWU). No differences were detected in plasma concentrations or C/D values (p=0.50 and p=0.96 for MWU). Likewise, differences in daily dose, plasma concentrations, or C/D values were not significant between light smokers (n=17) and nonsmokers (p=0.61, p=0.81, and p=0.33 for MWU) or heavy smokers (n=22) and nonsmokers (p=0.874, p=0.38, and p=0.59; MWU in all cases). Paliperidone is not affected by smoking, and paliperidone dose-adjustments in smokers may not be necessary. This may be seen as an essential difference to risperidone, whose cytochrome-mediated metabolism might be affected by smoking.

Detoxification mechanism of Aconiti Lateralis Radix Praeparata combined with dried Rehmanniae Radix based on metabolic enzymes in liver

The enzymes CYP1 A2 and CYP3 A4 were measured by building a "Cocktail" probe drug and the incubation system of liver microsomes. The compatibility of Aconiti Lateralis Radix Praeparata combined with dried Rehmanniae Radix on CYP450 enzyme protein and gene expression was explored from the level of protein and molecular biology. It explored the molecular mechanism of compatibility detoxication of Aconiti Lateralis Radix Praeparata to provide scientific support for clinical safe and effective application of Aconiti Lateralis Radix Praeparata. The CYP450 enzyme activity was determined by using "Cocktail" probe drugs. The content of CYP450 enzyme was measured by CO reduction of differential spectrum method. The mRNA expression of CYP1 A2 and CYP3 A4 enzyme was detected by RT-PCR technology. Compared with the blank group, the CYP1 A2 and CYP3 A4 enzyme activity and mRNA expression were increased in the dried Rehmanniae Radix combined with Aconiti Lateralis Radix Praeparata group with significant differences(P<0.05), while the CYP3 A4 enzyme activity and mRNA expression were no influence in the Aconiti Lateralis Radix Praeparata group. The CYP3 A4 enzyme activity and mRNA expression were increased in the dried Rehmanniae Radix and the dried Rehmanniae Radix combined with Aconiti Lateralis Radix Praeparata group, and there were significant differences(P<0.05). The content of CYP450 enzyme was decreased in the Aconiti Lateralis Radix Praeparata group, contributed to extremely significant difference(P<0.01). The content of CYP450 enzyme was increased in the dried Rehmanniae Radix and the dried Rehmanniae Radix combined with Aconiti Lateralis Radix Praeparata group, and there were significant differences(P<0.05). The CYP1 A2 and CYP3 A4 enzyme activity and gene expression were enhanced after dried Rehmanniae Radix combined with Aconiti Lateralis Radix Praeparata. The metabolism of toxic ingredients of Aconiti Lateralis Radix Praeparata was accelerated to reach an effect of detoxication. The detoxication mechanism of compatibility of Aconiti Lateralis Radix Praeparata was verified from the viewpoint of liver metabolic enzymes.

Isatidis Folium alleviates acetaminophen-induced liver injury in mice by enhancing the endogenous antioxidant system.

Isatidis Folium (IF) has been clinically combined with acetaminophen (APAP), but the rationality of combinational therapy is still ambiguous. In the present study, the protective effect and related mechanism of IF on APAP-induced hepatotoxicity were evaluated. Hepatic histopathology and blood biochemistry investigations clearly demonstrated that IF could restore APAP-induced hepatotoxicity. Liver distribution study indicated that the hepatoprotective effect of IF on APAP is attributed to the reduction of N-acetyl-p-benzoquinone imine (NAPQI) in liver, which is a known hepatotoxic metabolite of APAP. Further study suggested the reduction is not via decreasing the generation of NAPQI through inhibiting the enzyme activities of CYP 1A2, 2E1, and 3A4 but via accelerating the transformation of NAPQI to NAPQI-GSH by promoting GSH and decreasing GSSG contents in liver. Furthermore, IF significantly enhanced the hepatic activities of GSH-associated enzymes in APAP-treated mice. In summary, IF could alleviate APAP-induced hepatotoxicity by reducing the content of NAPQI via enhancing the level of GSH and the followed generation of NAPQI-GSH which might be ascribed to the upregulation of GSH-associated enzymes.

Metabolism of flutamide in diet control fischer 344 and brown Norway x F 344 rats, and its hydroxylation and conjugation by human CYP450s and UDP-glucuronosyltransferases

In our current report, flutamide (2-methyl-N-[4-nitro-3-(trifluoromethyl)-phenyl] propanamide) treated diet control and non limited-fed Fischer 344 (F344) and Brown Norway (BN) x F344 rats showed that diet control reduces spontaneous and flutamide-induced hyperplasia((1)). In this continued study, we found that serum concentrations of active metabolite of flutamide, 2-hydroxyflutamide (OH-flu), were 181 ± 26.6 ng/mL and 68 ± 8.0 ng/mL (p<0.05), in non-limited fed and diet control F344 rats. In BN x F344 rats, the serum concentrations of 2-OH-flutamide were 232 ± 57 ng/mL and 52 ± 6 ng/mL (p<0.05) in non-limited fed and diet control animals. In diet control groups, liver microsomes from flutamide-treated F344 rats showed high 7-ethoxyresorufin O-deethylase (EROD) activity, while 7-Benzoxyresorufin O-dealkylase (BROD) activity was not affected significantly. Both rat and human liver microsomes showed flutamide oxidation activity. Human liver microsomes showed 10 times higher activity than rat liver microsomes (0.673 ± 0.04 vs 0.063 ± 0.008 nmol OH-flu/min/mg protein). Microsomes from human tissues such as colon, colon cancer, kidney, bladder, pancreas, prostate, prostate cancer, or ovarian cancer, showed no or non-detectable activity for flutamide hydroxylation. CYP450 1A1, 1A2, 1B1 and 2C19 from human lymphoblastoid cell lines, oxidized flutamide to OH-flu in vitro with activities from 0.118 ± 0.005 to 0.275 ± 0.010 nmol/min/mg protein. Microsomes isolated from human kidney, colon, and liver showed UDP-glucuronosyltransferase (UGT) activity for glucuronidation of OH-flu. Human kidney showed the highest activity. Several human recombinant UGTs (1A1, 1A4, 1A6, 1A7, 1A9, and 1A10) also showed glucuronidation activity for OH-flu. It was found that UGT1A6 was more active than other human UGTs.

Drug – Drug Interactions Between Newer Anti- Retroviral Drugs And Anti Epileptics - A Review

Drug – Drug Interactions (DDIs) are the leading cause of drug toxicity and emergence of drug resistance, ultimately leading to increased burden in People Living with Human Immunodeficiency Virus (PLHIV). On an average 55 % of people on Anti Retroviral Therapy (ARVs) are co-administered with Anti Epileptic Drugs (AEDs). The introduction of newer anti-retroviral drugs such as dolutegravir, bictegravir, emtricitabine, doravirine are proven to have less side effects, high tolerability and effective decrease in the viral load, but the risk of DDIs still stands to be high. This review briefly describes about the pharmacokinetic properties of dolutegravir, bictegravir, emtricitabine, doravirine, mechanism of interaction between the above mentioned ARVs and AEDs, effect of DDIs on ARVs, effect of DDIs on interacting AEDs, outcome of DDIs and possible management of DDIs. The pharmacokinetic type of DDIs was observed between the ARVs and AEDs. The majority of DDIs were found affecting the metabolism and the absorption of the drugs. UGT1A1, CYP 3A are the two important classes of metabolic enzymes involved in the DDIs and p- glycoprotein (P-gp) is the transporter involved in the DDIs affecting the absorption. Significant interactions have been found in between the above mentioned newer ARV’s with carbamazepine, oxcarbazepine, phenytoin and phenobarbitol.