-
Chemical Research in Toxicology Feb 2021Electrophilically reactive drug metabolites are implicated in many adverse drug reactions. In this mechanism-termed bioactivation-metabolic enzymes convert drugs into...
Electrophilically reactive drug metabolites are implicated in many adverse drug reactions. In this mechanism-termed bioactivation-metabolic enzymes convert drugs into reactive metabolites that often conjugate to nucleophilic sites within biological macromolecules like proteins. Toxic metabolite-product adducts induce severe immune responses that can cause sometimes fatal disorders, most commonly in the form of liver injury, blood dyscrasia, or the dermatologic conditions toxic epidermal necrolysis and Stevens-Johnson syndrome. This study models four of the most common metabolic transformations that result in bioactivation: quinone formation, epoxidation, thiophene sulfur-oxidation, and nitroaromatic reduction, by synthesizing models of metabolism and reactivity. First, the metabolism models predict the formation probabilities of all possible metabolites among the pathways studied. Second, the exact structures of these metabolites are enumerated. Third, using these structures, the reactivity model predicts the reactivity of each metabolite. Finally, a feedfoward neural network converts the metabolism and reactivity predictions to a bioactivation prediction for each possible metabolite. These bioactivation predictions represent the joint probability that a metabolite forms and that this metabolite subsequently conjugates to protein or glutathione. Among molecules bioactivated by these pathways, we predicted the correct pathway with an AUC accuracy of 89.98%. Furthermore, the model predicts whether molecules will be bioactivated, distinguishing bioactivated and nonbioactivated molecules with 81.06% AUC. We applied this algorithm to withdrawn drugs. The known bioactivation pathways of alclofenac and benzbromarone were identified by the algorithm, and high probability bioactivation pathways not yet confirmed were identified for safrazine, zimelidine, and astemizole. This bioactivation model-the first of its kind that jointly considers both metabolism and reactivity-enables drug candidates to be quickly evaluated for a toxicity risk that often evades detection during preclinical trials. The XenoSite bioactivation model is available at http://swami.wustl.edu/xenosite/p/bioactivation.
Topics: Epoxy Compounds; Humans; Models, Biological; Molecular Structure; Nitrobenzenes; Oxidation-Reduction; Quinones; Sulfur; Thiophenes
PubMed: 33496184
DOI: 10.1021/acs.chemrestox.0c00417 -
Journal of Medicinal Chemistry Jun 2021Disruption of EZH2-embryonic ectoderm development (EED) protein-protein interaction (PPI) is a new promising cancer therapeutic strategy. We have previously reported the...
Disruption of EZH2-embryonic ectoderm development (EED) protein-protein interaction (PPI) is a new promising cancer therapeutic strategy. We have previously reported the discovery of astemizole, a small-molecule inhibitor targeting the EZH2-EED PPI. Herein, we report the cocrystal structure of EED in complex with astemizole at 2.15 Å. The structure elucidates the detailed binding mode of astemizole to EED and provides a structure-guided design for the discovery of a novel EZH2-EED interaction inhibitor, DC-PRC2in-01, with an affinity of 4.56 μM. DC-PRC2in-01 destabilizes the PRC2 complex, thereby leading to the degradation of PRC2 core proteins and the decrease of global H3K27me3 levels in cancer cells. The proliferation of PRC2-driven lymphomas cells is effectively inhibited, and the cell cycle is arrested in the G0/G1 phase. Together, these data demonstrate that DC-PRC2in-01 could be an effective chemical probe for investigating the PRC2-related physiology and pathology and providing a promising chemical scaffold for further development.
Topics: Astemizole; Cell Line, Tumor; Cell Proliferation; Drug Repositioning; Enhancer of Zeste Homolog 2 Protein; Enzyme Inhibitors; Humans; Molecular Docking Simulation; Molecular Structure; Polycomb Repressive Complex 2; Protein Binding; Structure-Activity Relationship
PubMed: 34077206
DOI: 10.1021/acs.jmedchem.0c02261 -
Methods in Molecular Biology (Clifton,... 2022Human ether-a-go-go-related gene (hERG) channel plays an essential role in the repolarization of the cardiac action potential. Genetic mutations and some chemicals/drugs...
Human ether-a-go-go-related gene (hERG) channel plays an essential role in the repolarization of the cardiac action potential. Genetic mutations and some chemicals/drugs interfere with hERG channel activity, which may prolong the QT interval and potentially cause long QT syndrome. The FluxOR™ thallium flux assay performed in two cell lines, U2OS and HEK293, with stable hERG expression can be used to identify compounds that inhibit hERG channel activity. This chapter describes a cell-based hERG channel inhibition assay that has been optimized and performed in a 1536-well plate format. The homogeneous and robust assay can be used to identify compounds that inhibit hERG channel activity.
Topics: Action Potentials; Ether-A-Go-Go Potassium Channels; HEK293 Cells; Humans; Long QT Syndrome; Research Design
PubMed: 35294752
DOI: 10.1007/978-1-0716-2213-1_3 -
International Journal of Molecular... Sep 2022Despite the significant progress made towards comprehending the deregulated signatures in lung cancer, these vary from study to study. We reanalyzed 25 studies from the...
Despite the significant progress made towards comprehending the deregulated signatures in lung cancer, these vary from study to study. We reanalyzed 25 studies from the Gene Expression Omnibus (GEO) to detect and annotate co-deregulated signatures in lung cancer and in single-gene or single-drug perturbation experiments. We aimed to decipher the networks that these co-deregulated genes (co-DEGs) form along with their upstream regulators. Differential expression and upstream regulators were computed using Characteristic Direction and Systems Biology tools, including GEO2Enrichr and X2K. Co-deregulated gene expression profiles were further validated across different molecular and immune subtypes in lung adenocarcinoma (TCGA-LUAD) and lung adenocarcinoma (TCGA-LUSC) datasets, as well as using immunohistochemistry data from the Human Protein Atlas, before being subjected to subsequent GO and KEGG enrichment analysis. The functional alterations of the co-upregulated genes in lung cancer were mostly related to immune response regulating the cell surface signaling pathway, in contrast to the co-downregulated genes, which were related to S-nitrosylation. Networks of hub proteins across the co-DEGs consisted of overlapping TFs (SOX2, MYC, KAT2A) and kinases (MAPK14, CSNK2A1 and CDKs). Furthermore, using Connectivity Map we highlighted putative repurposing drugs, including valproic acid, betonicine and astemizole. Similarly, we analyzed the co-DEG signatures in single-gene and single-drug perturbation experiments in lung cancer cell lines. In summary, we identified critical co-DEGs in lung cancer providing an innovative framework for their potential use in developing personalized therapeutic strategies.
Topics: Adenocarcinoma of Lung; Astemizole; Gene Expression Regulation, Neoplastic; Humans; Lung Neoplasms; Mitogen-Activated Protein Kinase 14; Transcription Factors; Valproic Acid
PubMed: 36142846
DOI: 10.3390/ijms231810933 -
European Journal of Medicinal Chemistry Jun 2022The development of inhibitors of key biological mechanisms involved in multidrug resistance (MDR) burden meets an important medical need but still represents a... (Review)
Review
The development of inhibitors of key biological mechanisms involved in multidrug resistance (MDR) burden meets an important medical need but still represents a challenging task. Major MDR targets in both bacterial and cancer cells are multidrug efflux systems. Several aspects should be considered in the attempt to design efficient inhibitors of these systems such as toxicity, stability, permeability as a few examples. In order to successfully design promising new compounds, a full understanding of the efflux mechanism is required, from both biological and structural points of view. It is nowadays well established that the success rate in classical drug design and biological evaluation improves when combined with in silico methodologies. In this review, we focus on the biological evaluation and molecular mechanistic insights of inhibitors of the drug efflux activity of the Hedgehog receptor Patched1 (Ptch1). Ptch1 is known to be over-expressed in many types of cancers, but its activity and role in the resistance to chemotherapy of cancer cells have been highlighted only recently. Remarkably, due to its peculiar efflux mechanism, inhibition of Ptch1 was shown to be particularly relevant for improving the efficacy of chemotherapy without concomitant toxicity for healthy cells or potential side effects. To date, three compounds have been identified as efficient Ptch1 inhibitors, namely astemizole, methiothepin and panicein A hydroquinone. Due to the chemical and structural differences of these molecules, the hit-to-lead drug design is not straightforward. This review describes how the merging of in vitro, in vivo and in silico studies provides molecular details that could contribute to the rational design of new Ptch1 inhibitors.
Topics: Drug Design; Drug Resistance, Multiple; Drug Resistance, Neoplasm; Hedgehog Proteins; Humans; Neoplasms; Patched-1 Receptor
PubMed: 35421658
DOI: 10.1016/j.ejmech.2022.114306 -
Toxicological Sciences : An Official... Oct 2022Validation of risk-stratification method for the chronic atrioventricular block cynomolgus monkey model and its mechanistic interpretation was performed using 6...
Validation of Risk-Stratification Method for the Chronic Atrioventricular Block Cynomolgus Monkey Model and Its Mechanistic Interpretation Using 6 Drugs With Pharmacologically Distinct Profile.
Validation of risk-stratification method for the chronic atrioventricular block cynomolgus monkey model and its mechanistic interpretation was performed using 6 pharmacologically distinct drugs. The following drugs were orally administered in conscious state, astemizole: 1, 5, and 10 mg/kg (n = 6); haloperidol: 1, 10, and 30 mg/kg (n = 5); amiodarone: 30 mg/kg (n = 4); famotidine: 10 mg/kg (n = 4); levofloxacin: 100 mg/kg (n = 4); and tolterodine: 0.2, 1, and 4.5 mg/kg (n = 4). Astemizole of 5 and 10 mg/kg significantly prolonged ΔΔQTcF, whereas no significant change was observed by the others. Torsade de pointes (TdP) was induced by astemizole of 5 and 10 mg/kg in 3/6 and 6/6, and by haloperidol of 10 and 30 mg/kg in 1/5 and 1/5, respectively, which was not observed in the others. Torsadogenic risk of the drugs was quantified using the criteria for the monkey model specified in our previous study. Namely, high-risk drugs induced TdP at ≤ 3 times of their maximum clinical daily dose. Intermediate-risk drugs did not induce TdP at this dose range, but induced it at higher doses. Low/no-risk drugs never induced TdP at any dose tested. The magnitude of risk was intermediate for astemizole and haloperidol, and low/no risk for the others. The prespecified, risk-stratification method for the monkey model may solve the issue existing between nonclinical models and patients with labile repolarization, which can reinforce the regulatory decision-making and labeling at time of marketing application of nondouble-negative drug candidate (hERG assay positive and/or in vivo QT study positive).
Topics: Animals; Atrioventricular Block; Macaca fascicularis; Astemizole; Haloperidol; Torsades de Pointes; DNA-Binding Proteins; Electrocardiography
PubMed: 35993620
DOI: 10.1093/toxsci/kfac088 -
Biophysical Journal May 2020High-throughput in vitro drug assays have been impacted by recent advances in human induced pluripotent stem cell-derived cardiomyocyte (hiPSC-CM) technology and by...
High-throughput in vitro drug assays have been impacted by recent advances in human induced pluripotent stem cell-derived cardiomyocyte (hiPSC-CM) technology and by contact-free all-optical systems simultaneously measuring action potentials (APs) and Ca transients (CaTrs). Parallel computational advances have shown that in silico simulations can predict drug effects with high accuracy. We combine these in vitro and in silico technologies and demonstrate the utility of high-throughput experimental data to refine in silico hiPSC-CM populations and to predict and explain drug action mechanisms. Optically obtained hiPSC-CM APs and CaTrs were used from spontaneous activity and under optical pacing in control and drug conditions at multiple doses. An updated version of the Paci2018 model was developed to refine the description of hiPSC-CM spontaneous electrical activity; a population of in silico hiPSC-CMs was constructed and calibrated using simultaneously recorded APs and CaTrs. We tested in silico five drugs (astemizole, dofetilide, ibutilide, bepridil, and diltiazem) and compared the outcomes to in vitro optical recordings. Our simulations showed that physiologically accurate population of models can be obtained by integrating AP and CaTr control records. Thus, constructed population of models correctly predicted the drug effects and occurrence of adverse episodes, even though the population was optimized only based on control data and in vitro drug testing data were not deployed during its calibration. Furthermore, the in silico investigation yielded mechanistic insights; e.g., through simulations, bepridil's more proarrhythmic action in adult cardiomyocytes compared to hiPSC-CMs could be traced to the different expression of ion currents in the two. Therefore, our work 1) supports the utility of all-optical electrophysiology in providing high-content data to refine experimentally calibrated populations of in silico hiPSC-CMs, 2) offers insights into certain limitations when translating results obtained in hiPSC-CMs to humans, and 3) shows the strength of combining high-throughput in vitro and population in silico approaches.
Topics: Action Potentials; Adult; Computer Simulation; Drug Evaluation; Humans; Induced Pluripotent Stem Cells; Myocytes, Cardiac
PubMed: 32298635
DOI: 10.1016/j.bpj.2020.03.018 -
Xenobiotica; the Fate of Foreign... Jul 20221. Cytochrome P450 2J2 (CYP2J2) shows high expression in extrahepatic tissues, including the heart and kidney and in tumours. Inhibition of CYP2J2 has attracted...
1. Cytochrome P450 2J2 (CYP2J2) shows high expression in extrahepatic tissues, including the heart and kidney and in tumours. Inhibition of CYP2J2 has attracted attention for cancer treatment because it metabolises arachidonic acid (AA) to epoxyeicosatrienoic acid (EET), which inhibits apoptosis and promotes tumour growth. Multi-kinase inhibitor (MKI) is a molecular-targeted drug with antitumor activities. This study aimed to clarify the inhibitory effects of MKIs on CYP2J2 activity. We also investigated whether MKIs affected CYP2J2-catalysed EET formation from AA.2. Twenty MKIs showed different inhibitory potencies against astemizole -demethylation in CYP2J2. In particular, apatinib, motesanib, and vatalanib strongly inhibited astemizole -demethylation. These three MKIs exhibited competitive inhibition with inhibition constant () values of 9.3, 15.4, and 65.0 nM, respectively. Apatinib, motesanib, and vatalanib also inhibited CYP2J2-catalysed 14,15-EET formation from AA.3. In simulations of docking to CYP2J2, the energy values of apatinib, motesanib, and vatalanib were low, and measured -84.5, -69.9, and -52.3 kcal/mol, respectively.4. In conclusion, apatinib, motesanib, and vatalanib strongly inhibited CYP2J2 activity, suggesting that the effects of a given CYP2J2 substrate may be altered upon the administration of these MKIs.
Topics: Cytochrome P-450 Enzyme System
PubMed: 36251932
DOI: 10.1080/00498254.2022.2137068 -
Drug Metabolism and Disposition: the... Nov 2022Cytochrome P450s (P450s) have been identified and analyzed in dogs and pigs, species that are often used in preclinical drug studies. Moreover, P450s are clinically...
Cytochrome P450s (P450s) have been identified and analyzed in dogs and pigs, species that are often used in preclinical drug studies. Moreover, P450s are clinically important for drug therapy not only in humans, but also in species under veterinary care, including dogs and cats. In the present study, seven P450s homologous to human CYP2J2, namely, dog CYP2J2; cat CYP2J2; and pig CYP2J33, CYP2J35, CYP2J91, and CYP2J93, were newly identified and characterized, along with pig CYP2J34 previously identified. The cDNAs of these CYP2Js contain open reading frames of 502 amino acids, except for CYP2J35 (498 amino acids), and share high sequence identity (77%-80%) with human CYP2J2. Phylogenetic analysis revealed that dog and cat CYP2J2 were closely related, whereas pig CYP2Js formed a cluster. All seven genes contain nine coding exons and are located in corresponding genomic regions, with the pig genes forming a gene cluster. These CYP2J2 mRNAs were predominantly expressed in the small intestine with additional expression in the kidney and brain for dog CYP2J2 and pig CYP2J91 mRNAs, respectively. All seven CYP2Js metabolized human CYP2J2 substrates terfenadine, ebastine, and astemizole, indicating that they are functional enzymes. Dog CYP2J2 and pig CYP2J34 and CYP2J35 efficiently catalyzed ebastine primary hydroxylation and secondary carebastine formation at low substrate concentrations, just as human CYP2J2 does. Velocityversus-substate plots exhibited sigmoidal relationships for dog CYP2J2, cat CYP2J2, and pig CYP2J33, indicating allosteric interactions. These results suggest that dog, cat, and pig CYP2Js have similar functional characteristics to human CYP2J2, with slight differences in ebastine and astemizole oxidations. SIGNIFICANCE STATEMENT: Dog CYP2J2; cat CYP2J2; and pig CYP2J33, CYP2J34, CYP2J35, CYP2J91, and CYP2J93, homologous to human CYP2J2, were identified and characterized by sequence, phylogenetic, and genomic structure analyses. Intestinal expression patterns of CYP2J mRNAs were characteristic in dogs, cats, and pigs. Dog, cat, and pig CYP2Js likely play roles as drug-metabolizing enzymes in the small intestine, similar to human CYP2J2.
Topics: Animals; Astemizole; Butyrophenones; Cats; Cytochrome P-450 CYP2J2; Cytochrome P-450 Enzyme System; Dogs; Humans; Phylogeny; Piperidines; Swine; Terfenadine
PubMed: 35701183
DOI: 10.1124/dmd.122.000930 -
European Journal of Pharmacology Aug 2022The transient receptor potential (TRP) channel TRPV2 is widely expressed in a variety of different cell types and tissues. However, elucidating the exact biological...
The transient receptor potential (TRP) channel TRPV2 is widely expressed in a variety of different cell types and tissues. However, elucidating the exact biological functions of TRPV2 is significantly hampered by the lack of selective pharmacological tools to modulate channel activity in vitro and in vivo. This study aimed to identify new compounds that modify TRPV2 activity via the use of a plate-based calcium imaging approach to screen a drug repurposing library. Three antihistaminic drugs, loratadine, astemizole and clemizole were identified to reduce calcium-influx evoked by the TRPV2 agonist tetrahydrocannabivarin in HEK293 cells expressing murine TRPV2. Using single-cell calcium-microfluorimetry and whole-cell patch clamp recordings, we further confirmed that all three compounds induced a concentration-dependent block of TRPV2-mediated Ca influx and whole-cell currents, with loratadine being the most potent antagonist of TRPV2. Moreover, this study demonstrated that loratadine was able to block both the human and mouse TRPV2 orthologs, without inhibiting the activity of other closely related members of the TRPV superfamily. Finally, loratadine inhibited TRPV2-dependent responses in a primary culture of mouse endometrial stromal cells and attenuated cell proliferation and migration in in vitro cell proliferation and wound healing assays. Taken together, our study revealed that the antihistaminic drugs loratadine, astemizole and clemizole target TRPV2 in a concentration-dependent manner. The identification of these antihistaminic drugs as blockers of TRPV2 may form a new starting point for the synthesis of more potent and selective TRPV2 antagonists, which could further lead to the unravelling of the physiological role of the channel.
Topics: Animals; Astemizole; Benzimidazoles; Calcium; Calcium Channel Blockers; Calcium Channels; Cell Proliferation; HEK293 Cells; Histamine Antagonists; Humans; Loratadine; Mice; Stromal Cells; TRPV Cation Channels; Transient Receptor Potential Channels
PubMed: 35714693
DOI: 10.1016/j.ejphar.2022.175086