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ACS Medicinal Chemistry Letters Apr 2024Toward addressing the cardiotoxicity liability associated with the antimalarial drug astemizole (AST, hERG IC = 0.0042 μM) and its derivatives, we designed and...
Toward addressing the cardiotoxicity liability associated with the antimalarial drug astemizole (AST, hERG IC = 0.0042 μM) and its derivatives, we designed and synthesized analogues based on compound ( NF54 IC = 0.012 μM; hERG IC = 0.63 μM), our previously identified 3-trifluoromethyl-1,2,4-oxadiazole AST analogue. Compound retained multistage antiplasmodium activity (ABS NF54 IC = 0.017 μM; gametocytes iGc/LGc IC = 1.24/1.39 μM, and liver-stage HepG2 IC = 2.30 μM), good microsomal metabolic stability (MLM CL < 11 μL·min·mg, < 0.33), and solubility (150 μM). It shows a ∼6-fold and >6000-fold higher selectivity against human ether-á-go-go-related gene higher selectively potential over hERG relative to and AST, respectively. Despite the excellent antiplasmodium activity profile, efficacy in the mouse infection model was diminished, attributable to suboptimal oral bioavailability ( = 14.9%) at 10 mg·kg resulting from poor permeability (log = -0.82). No cross-resistance was observed against 44 common mutant lines, suggesting activity via a novel mechanism of action.
PubMed: 38628794
DOI: 10.1021/acsmedchemlett.3c00496 -
European Journal of Pharmaceutical... Sep 2021Extrahepatic CYP2J2 metabolism of arachidonic acid (AA) to bioactive regioisomeric epoxyeicosatrienoic acids (EETs) is implicated in both physiological and pathological...
Extrahepatic CYP2J2 metabolism of arachidonic acid (AA) to bioactive regioisomeric epoxyeicosatrienoic acids (EETs) is implicated in both physiological and pathological conditions. Here, we aimed to characterize atypical substrate inhibition kinetics of this endogenous metabolic pathway and its reversible inhibition by xenobiotic inhibitors when AA is used as the physiologically-relevant substrate vis-à-vis conventional probe substrate astemizole (AST). As compared to typical Michaelis-Menten kinetics observed for AST, complete substrate inhibition was observed for CYP2J2 metabolism of AA to 14,15-EET whereby velocity of the reaction declined significantly at concentrations of AA above 20-30 µM with an estimated substrate inhibition constant (K) of 31 µM. In silico sequential docking of two AA substrates to orthosteric (OBS) and adjacent secondary binding sites (SBS) within a 3-dimensional homology model of CYP2J2 revealed favorable and comparable binding poses of glide-scores -3.1 and -3.8 respectively. Molecular dynamics (MD) simulations ascertained CYP2J2 conformational stability with dual AA substrate binding as time-dependent root mean squared deviation (RMSD) of protein Cα atoms and ligand heavy atoms stabilized to a plateau in all but one trajectory (n=6). The distance between heme-iron and ω6 (C14, C15) double bond of AA in OBS also increased from 7.5 ± 1.4 Å to 8.5 ± 1.8 Å when CYP2J2 was simulated with only AA in OBS versus the presence of AA in both OBS and SBS (p<0.001), supporting the observed in vitro substrate inhibition phenomenon. Poor correlation was observed between inhibitory constants (K) determined for a panel of nine competitive and mixed mode xenobiotic inhibitors against CYP2J2 metabolism of AA as compared to AST, whereby 4 out of 9 drugs had a greater than 5-fold difference between K values. Nonlinear Eadie-Hofstee plots illustrated that complete substrate inhibition of CYP2J2 by AA was not attenuated even at high concentrations of xenobiotic inhibitors which further corroborates that CYP2J2 may accommodate three or more ligands simultaneously. In light of the atypical kinetics, our results highlight the importance of using physiologically-relevant substrates in in vitro enzymatic inhibition assays for the characterization of xenobiotic-endobiotic interactions which is applicable to other complex endogenous metabolic pathways beyond CYP2J2 metabolism of AA to EETs. The accurate determination of K would further facilitate the association of xenobiotic-endobiotic interactions to observed therapeutic or toxic outcomes.
Topics: Arachidonic Acid; Cytochrome P-450 Enzyme Inhibitors; Cytochrome P-450 Enzyme System; Kinetics; Xenobiotics
PubMed: 34044117
DOI: 10.1016/j.ejps.2021.105889 -
European Journal of Pharmacology Jul 2019Chloroquine (CQ) is an old antimalarial drug currently being investigated for its anti-tumor properties. As chloroquine has been shown to inhibits several potassium...
Chloroquine (CQ) is an old antimalarial drug currently being investigated for its anti-tumor properties. As chloroquine has been shown to inhibits several potassium channels, we decided to study its effect on the tumor-related Kv10.1 channel by using patch-clamp electrophysiology and cell migration assays. We found that chloroquine inhibited Kv10.1 channels transiently expressed in HEK-293 cells in a concentration- and voltage-dependent manner acting from the cytoplasmic side of the plasma membrane. Chloroquine also inhibited the outward potassium currents from MDA-MB-231 cells, which are mainly carried through Kv10.1 channels as was confirmed using astemizole. Additionally, chloroquine decreased MDA-MB-231 cell migration in the in vitro scratch wound healing assay. In conclusion, our data suggest that chloroquine decreases MDA-MB-231 cell migration by inhibiting Kv10.1 channels. The inhibition of Kv10.1 channels could represent another mechanism of the antitumoral action of chloroquine, besides autophagy inhibition and tumor vessel normalization.
Topics: Breast Neoplasms; Cell Line, Tumor; Cell Membrane; Cell Movement; Chloroquine; Cytoplasm; Ether-A-Go-Go Potassium Channels; HEK293 Cells; Humans; Potassium Channel Blockers
PubMed: 31082369
DOI: 10.1016/j.ejphar.2019.05.017 -
Journal of Pharmacological and... Sep 2020Screening compounds for activity on the hERG channel using patch clamp is a crucial part of safety testing. Automated patch clamp (APC) is becoming widely accepted as an...
Reliable identification of cardiac liability in drug discovery using automated patch clamp: Benchmarking best practices and calibration standards for improved proarrhythmic assessment.
INTRODUCTION
Screening compounds for activity on the hERG channel using patch clamp is a crucial part of safety testing. Automated patch clamp (APC) is becoming widely accepted as an alternative to manual patch clamp in order to increase throughput whilst maintaining data quality. In order to standardize APC experiments, we have investigated the effects on IC values under different conditions using several devices across multiple sites.
METHODS
APC instruments SyncroPatch 384i, SyncroPatch 384PE and Patchliner, were used to record hERG expressed in HEK or CHO cells. Up to 27 CiPA compounds were used to investigate effects of voltage protocol, incubation time, labware and time between compound preparation and experiment on IC values.
RESULTS
All IC values of 21 compounds recorded on the SyncroPatch 384PE correlated well with IC values from the literature (Kramer et al., 2013) regardless of voltage protocol or labware, when compounds were used immediately after preparation, but potency of astemizole decreased if prepared in Teflon or polypropylene (PP) compound plates 2-3 h prior to experiments. Slow acting compounds such as dofetilide, astemizole, and terfenadine required extended incubation times of at least 6 min to reach steady state and therefore, stable IC values.
DISCUSSION
Assessing the influence of different experimental conditions on hERG assay reliability, we conclude that either the step-ramp protocol recommended by CiPA or a standard 2-s step-pulse protocol can be used to record hERG; a minimum incubation time of 5 min should be used and although glass, Teflon, PP or polystyrene (PS) compound plates can be used for experiments, caution should be taken if using Teflon, PS or PP vessels as some adsorption can occur if experiments are not performed immediately after preparation. Our recommendations are not limited to the APC devices described in this report, but could also be extended to other APC devices.
Topics: Animals; Arrhythmias, Cardiac; Astemizole; Benchmarking; CHO Cells; Calibration; Cardiovascular Agents; Cell Line; Cricetulus; Drug Discovery; Drug Evaluation, Preclinical; ERG1 Potassium Channel; HEK293 Cells; Heart; Humans; Patch-Clamp Techniques; Phenethylamines; Polypropylenes; Polytetrafluoroethylene; Reference Standards; Reproducibility of Results; Sulfonamides; Terfenadine
PubMed: 32565325
DOI: 10.1016/j.vascn.2020.106884 -
Expert Opinion on Drug Safety Jan 2021: This study aimed to measure the association of various H1-antihistamines (H1A) with Torsade de Pointes (TdP), and present a comprehensive overview of H1A-induced...
: This study aimed to measure the association of various H1-antihistamines (H1A) with Torsade de Pointes (TdP), and present a comprehensive overview of H1A-induced TdP cases reported to the Food and Drug Administration Adverse Event Reporting System (FAERS). : All H1A-induced TdP cases (n = 406) were retrieved from the FAERS database using the preferred term 'Torsade de Pointes' of MedDRA version-22 from 1990 to 2019. Four data-mining algorithms were used for disproportionality analysis: Reporting Odds Ratio (ROR); Proportional Reporting Ratio (PRR), Empirical Bayes Geometric Mean (EBGM), and Information Content (IC). H1A with >3 TdP cases were included. : A total of 12 signals (Astemizole, cetirizine, chlorpheniramine, clemastine, desloratadine, diphenhydramine, hydroxyzine, loratadine, meclizine, promethazine, terfenadine, and trimeprazine) were identified including six new signals (cetirizine, chlorpheniramine, clemastine, desloratadine, loratadine, and meclizine). The number of risk factors (p = 0.031) and concomitant QT-prolonging drugs (p = <0.001) were significantly lower among new signals vs old signals. Moreover, new signals were strongly associated with QT-prolongation, cardiac reactions, and electrolyte abnormalities as compared with old signals. : Our study found the increased torsadogenic potential of new signals compared with previously known old signals, hence necessitating clinical studies to determine the actual torsadogenic potential of newly identified signals.
Topics: Adolescent; Adult; Adverse Drug Reaction Reporting Systems; Aged; Algorithms; Child; Child, Preschool; Data Mining; Databases, Factual; Female; Histamine H1 Antagonists; Humans; Infant; Long QT Syndrome; Male; Middle Aged; Pharmacovigilance; Risk Factors; Torsades de Pointes; United States; United States Food and Drug Administration; Young Adult
PubMed: 33141610
DOI: 10.1080/14740338.2021.1846717