Drug Metabolism and Disposition : the Biological Fate of Chemicals

In vitro models that can faithfully replicate critical aspects of kidney tubule function such as directional drug transport are in high demand in pharmacology and toxicology. Accordingly, development and validation of new models is underway. The objective of this study was to characterize physiological and transport functions of various sources of human renal proximal tubule epithelial cells (RPTECs). We tested TERT1-immortalized RPTEC, including OAT1-, OCT2- or OAT3-overexpressing variants, and primary RPTECs...

The accurate prediction of human clearance is an important task during drug development. The proportion of low clearance compounds has increased in drug development pipelines across the industry since such compounds may be dosed in lower amounts and at lower frequency. Such compounds present new challenges to in vitro systems used for clearance extrapolation. In this study we compared the accuracy of clearance predictions of suspension culture to four different long-term stable in vitro liver models, including HepaRG sandwich culture, the Hµrel stochastic co-culture, the Hepatopac micropatterned co-culture (MPCC) and a micro-array spheroid culture...

Extrapolating in vivo hepatic clearance from in vitro uptake data is a known challenge, especially for OATP substrates, and the well-stirred model (WSM) commonly yields systematic under-predictions for those anionic drugs hypothetically due to "albumin-mediated hepatic drug uptake". In the present study, we demonstrate that the WSM incorporating the dynamic free fraction ( f D ), a measure of drug protein binding affinity, performs reasonably well in predicting hepatic clearance of OATP substrates. For a selection of anionic drugs including atorvastatin, fluvastatin, pravastatin, rosuvastatin, pitavastatin, cerivastatin, and repaglinide, this dynamic well-stirred model (dWSM) correctly predicts hepatic plasma clearance within 2-fold error for six out of seven OATP substrates examined...

CYP4Z1, a highly expressed CYP gene in breast cancer, was one of the last CYPs to be identified in the human genome, some twenty years ago. CYP4 enzymes typically catalyze w-hydroxylation and metabolize w3 and w6 polyunsaturated fatty acids (PUFAs) to bioactive lipid metabolites that can influence tumor growth and metastasis. These attributes of CYP4Z1 make it an attractive target for new chemotherapeutic drug design, as a potential biomarker for selection of patients that might respond favorably to drugs and for developing enzyme inhibitors as potential therapeutic agents...

VX is an organophosphate acetylcholinesterase (AChE) inhibitor and while it is one of the most toxic AChE inhibitors known the extent of metabolism in humans is not currently well understood. The known metabolism in humans is limited to the metabolite identification from a single victim of the Osaka poisoning in 1994, which allowed for the identification of several metabolic products. VX has been reported to be metabolized in vitro by paraoxonase-1 and phosphotriesterase, although their binding constants are many orders of magnitude above the LD50 , suggesting limited physiological relevance...

In the area of drug development and clinical pharmacotherapy, a profound understanding of the pharmacokinetics and potential adverse reactions associated with the drug under investigation is paramount. Essential to this endeavour is a comprehensive understanding about interindividual variations in ADME genetics and the predictive capabilities of in vitro systems, shedding light on metabolite formation and the risk of adverse drug reactions (ADRs). Both the domains of pharmacogenomics and the advancement of in vitro systems are experiencing rapid expansion...

Background Metastasis is the most common pathway of cancer death. The lack of effective predictors of breast metastasis is a pressing issue in clinical practice. Therefore, exploring the mechanism of breast cancer metastasis to uncover reliable predictors is very important for the clinical treatment of breast cancer patients. Methods TMT quantitative proteomics technology was used to detect protein content in primary breast tumor tissue samples from patients with metastatic and non-metastatic breast cancer at diagnosis...

Aldehyde oxidase (AO) is a molybdenum cofactor-containing cytosolic enzyme that has gained prominence due to its involvement in the developmental failure of several drug candidates in first-in-human trials. Unlike cytochrome P450s (P450) and glucuronosyltransferase, AO substrates have been plagued by poor in vitro to in vivo extrapolation, leading to low systemic exposures and underprediction of human dose. However, apart from measuring a drug's AO clearance rates, it is also important to determine the relative contribution to metabolism by this enzyme (fm,AO )...

The human UDP-glucuronosyltransferases (UGTs) have crucial roles in metabolizing and clearing numerous small lipophilic compounds. The UGT1A locus generates nine UGT1A mRNAs, 65 spliced transcripts and 34 circular RNAs. In this study, our analysis of published UGT-CaptureSeq datasets identified novel splice junctions that predict 24 variant UGT1A transcripts derived from ligation of exon 2 to unique sequences within the UGT1A first-exon region using cryptic donor splice sites. Of these variants, seven (1A1_n1, 1A3_n3, 1A4_n4, 1A5_n1, 1A8_n2, 1A9_n2v, 1A10_n7) are predicted to encode UGT1A proteins with truncated aglycone-binding domains...

Cytochrome P450 1A2 (CYP1A2) is a known tumor suppressor in hepatocellular carcinoma (HCC), but its expression is repressed in HCC and the underlying mechanism is unclear. In this study, we investigated the epigenetic mechanisms of CYP1A2 repression and potential therapeutic implications. In HCC tumor tissues, the methylation rates of CYP1A2 CpG island (CGI) and DNMT3A protein levels were significantly higher, and there was a clear negative correlation between DNMT3A and CYP1A2 protein expression. Knockdown of DNMT3A by siRNA significantly increased CYP1A2 expression in HCC cells...

Hepatic stellate cells (HSC) are the major site of vitamin A (retinol) esterification and subsequent storage as retinyl esters within lipid droplets. However, retinyl esters become depleted in many pathophysiological states, including acute and chronic liver injuries. Recently, using a liver slice culture system as a model of acute liver injury and fibrogenesis, a time-dependent increase and decrease in the apparent formation of the bioactive retinoid all- trans -retinoic acid ( at RA) and retinyl palmitate was measured, respectively...

Organic anion transporting polypeptide (OATP) 1B1 and OATP1B3 (collectively, OATP1B) transporters encoded by the solute carrier organic anion transporter ( SLCO) genes mediate uptake of multiple pharmaceutical compounds. Nonalcoholic steatohepatitis (NASH), a severe form of nonalcoholic fatty liver disease (NAFLD), decreases OATP1B abundance. This research characterized the pathological and pharmacokinetics effects of three diet- and one chemical-induced NAFLD model in male and female humanized OATP1B mice, which comprises knock-out of rodent Oatp orthologs and insertion of human SLCO1B1 and SLCO1B3...

Drug-induced liver injury (DILI) is a frequent cause of clinical trial failures during drug development. While inhibiting BSEP is a well-documented DILI mechanism, interference with genes related to bile acid (BA) metabolism and transport can further complicate DILI development. Here, the effects of twenty-eight compounds on genes associated with BA metabolism and transport were evaluated, including those with discontinued development or use, boxed warnings, and clean labels for DILI. The study also included rifampicin and omeprazole, PXR and AhR ligands, and four mitogen-activated protein kinase kinase (MEK1/2) inhibitors...

Bupropion is used for treating depression, obesity, seasonal affective disorder, and for smoking cessation. Bupropion is commonly-prescribed, but has complex pharmacokinetics and interindividual variability in metabolism and bioactivation may influence therapeutic response, tolerability and safety. Bupropion is extensively and stereoselectively metabolized, the metabolites are pharmacologically active, and allelic variation in CYP2B6 affects clinical hydroxylation of single-dose bupropion. Genetic effects on stereoselective disposition of steady-state bupropion are not known...

The determination of metabolic stability is critical for drug discovery programs, allowing for the optimization of chemical entities and compound prioritization. As such, it is common to perform high-volume in vitro metabolic stability experiments early in the lead optimization process to understand metabolic liabilities. Additional metabolite identification experiments are subsequently performed for a more comprehensive understanding of the metabolic clearance routes to aid medicinal chemists in the structural design of compounds...

In vitro-in vivo extrapolation (IVIVE) allows prediction of clinical outcomes across populations from in vitro data using specific scalars tailored to the biological characteristics of each population. This study experimentally determined scalars for patients with varying degrees of non-alcoholic fatty liver disease (NAFLD), ranging from fatty liver to non-alcoholic steatohepatitis (NASH) and cirrhosis. Microsomal, S9 and cytosol fractions were extracted from 36 histologically normal and 66 NAFLD livers (27 NAFL, 13 NASH, and 26 NASH with cirrhosis)...

Fluorination of organic compounds plays an important role in the chemical and pharmaceutical industry and is often applied in order to improve physicochemical parameters or modify pharmacological properties. While oxidative and reductive defluorination have been shown to be responsible for the metabolic degradation of organofluorine compounds, the involvement of hydrolytic mechanisms catalyzed by human enzymes has not been reported so far. Here, we investigated the enzymatic defluorination of terminally monofluorinated aliphates with [1-(5-fluoropentyl)-1 H -indol-3-yl]-1-naphthalenyl-methanone (AM-2201) as a model substance...

In vitro time-dependent inhibition (TDI) kinetic parameters for cytochrome P450 (CYP) 1A2, 2B6, 2C8, 2C9, 2C19, and 2D6, were determined in pooled human liver microsomes for 19 drugs (and 2 metabolites) for which clinical drug-drug interactions (DDI) are known. In vitro TDI data were incorporated into the projection of the magnitude of DDIs using mechanistic static models and Simcyp®. Results suggest that for the mechanistic static model, use of estimated average unbound exit concentration of the inhibitor from the liver resulted in a successful prediction of observed magnitude of clinical DDIs and was similar to Simcyp®...

Organic anion transporting polypeptide (OATP1B) plays a key role in the hepatic clearance of a majority of high molecular weight (MW) acids and zwitterions. Here, we evaluated the role of OATP1B-mediated uptake in the clearance of novel hypoxia-inducible factor prolyl hydroxylase inhibitors ("Dustats"), which are typically low MW (300-400 daltons) aliphatic carboxylic acids. Five acid dustats, namely daprodustat, desidustat, enarodustat, roxadustat and vadadustat, showed specific transport by OATP1B1/1B3 in transporter-transfected HEK293 cells...

The regulation of drug-metabolizing enzymes and transporters by cytokines has been extensively studied, in vitro and in clinic. Cytokine-mediated suppression of CYPs or drug transporters may increase or decrease the systemic clearance of drug substrates that are primarily cleared via these pathways; neutralization of cytokines by therapeutic proteins may thereby alter systemic exposures of such drug substrates. The FDA recommends evaluating such clinical drug interactions during clinical development and has provided labeling recommendations for therapeutic proteins...