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Phenyl 4-(2-oxo-3-alkylimidazolidin-1-yl)benzenesulfonates (PAIB-SOs) are new antimitotic prodrugs bioactivated by the enzyme CYP1A1 into phenyl 4-(2-oxo-3-imidazolidin-1-yl)benzenesulfonates (PIB-SOs) that are potently (nM) and selectively inhibiting the growth of CYP1A1-expressing breast cancer cells. The screening program identified three PAIB-SOs designated as CEU-835, CEU-934, and CEU-938 for further pharmacokinetic studies. A novel method for their quantification in CD-1® IGS female mouse plasma using a Waters UHPLC ACQUITY Arc system was developed and validated. Their analytical measurements ranged from 7.3 to 1484 ng/mL. They exhibited linearity between 7.3 and 1484 ng/mL. They also showed good resolutions (2.21–4.99) and good theoretical plate numbers (14707–70580). Moreover, they were robust at wavelengths between 273 and 310 nm and sensitive from 2 to 62 AU/ng mL−1. This new method is indispensable to assess PAIB-SOs half-life in mouse plasma and for the quantification of PAIB-SOs in other biological matrices. [ABSTRACT FROM AUTHOR]

Aims: Some population pharmacokinetic models have been developed using height to explain some of the interindividual variability in tobramycin pharmacokinetics in cystic fibrosis patients. However, their predictive performance when extrapolated to other clinical centres is unclear. Therefore, the aim of this study was to externally evaluate the predictability of tobramycin population pharmacokinetic models with an independent dataset and perform simulations using previously recommended height‐based dosing regimens. Methods: A literature search was conducted through the PubMed database to identify relevant population pharmacokinetic models. Tobramycin plasma concentration data from April 2014 to November 2019 were retrospectively collected from the Institut universitaire de cardiologie et de pneumologie de Québec, Canada. External evaluations were performed using NONMEM® v7.5 and RStudio® v1.3.1073. Monte Carlo simulations were performed to evaluate the probability of target attainment of Cmax/MIC ratios for several dosing regimens. Results: The validation dataset included 27 patients and 143 concentration samples. Three models were evaluated. Only the ones by Crass et al. and Alghanem et al. performed satisfactorily in terms of prediction‐based diagnostics with MDPE values of −3.4% and 29.3% and MDAPE values of 19.0 and 29.5%, respectively. In simulation‐based evaluations, both pcVPC and NPDE showed no evidence of model misspecification. Our simulations suggest that patients treated with a once‐daily dose of 3.4 mg/cm should produce peak and trough levels consistent with current guidelines. Conclusion: Our results show that the models by Crass et al. and Alghanem et al. are appropriate for simulation‐based applications to aid individualized dosing in our population and that height‐based dosing regimens could be considered in cystic fibrosis patients. [ABSTRACT FROM AUTHOR]

New Findings: What is the role of SCN5A‐C683R?SCN5A‐C683R is a novel variant associated with an uncommon phenotype of adrenaline‐triggered ventricular arrhythmia in the absence of a distinct ECG phenotype.What is the main finding and its importance?Functional studies demonstrated that NaV1.5/C683R results in a mixed electrophysiological phenotype with gain‐of‐function (GOF) and loss‐of‐function (LOF) properties compared with NaV1.5/wild type. Gain‐of‐function properties are characterized by a significant increase of the maximal current density and a hyperpolarizing shift of the steady‐state activation. The LOF effect of NaV1.5/C683R is characterized by increased closed‐state inactivation. Electrophysiological properties and clinical manifestation of SCN5A‐C683R are different from long‐QT‐3 or Brugada syndrome and might represent a distinct inherited arrhythmia syndrome. Mutations of SCN5Ahave been identified as the genetic substrate of various inherited arrhythmia syndromes, including long‐QT‐3 and Brugada syndrome. We recently identified a novel SCN5A variant (C683R) in two genetically unrelated families. The index patients of both families experienced adrenaline‐triggered ventricular arrhythmia with cardiac arrest but did not show a specific ECG phenotype, raising the hypothesis that SCN5A‐C683R might be a susceptibility variant and the genetic substrate of distinct inherited arrhythmia. We conducted functional cellular studies to characterize the electrophysiological properties of NaV1.5/C683R in order to explore the potential pathogenicity of this novel variant. The C683R variant was engineered by site‐directed mutagenesis. NaV1.5/wild type (WT) and NaV1.5/C683R were expressed in tsA201 cells. Electrophysiological characterization of C683R was performed using the whole‐cell patch‐clamp technique. Adrenergic stimulation was mimicked by exposure to the protein kinase A activator 8‐CPT‐cAMP. The impact of β‐blockers was tested by exposing NaV1.5/WT and NaV1.5/C683R currents to propranolol and nadolol. C683R resulted in a co‐association of gain‐of‐function and loss‐of‐function properties of NaV1.5. Gain‐of‐function properties were characterized by a significant increase of the maximal NaV1.5 current density compared with NaV1.5/WT (861 ± 309 vs. 627 ± 489 pA/pF; P < 0.05, n ≥ 9) that was potentiated in NaV1.5/C683R with 8‐CPT‐cAMP stimulation (869 ± 287 vs. 607 ± 320 pA/pF; P < 0.05, n ≥ 12). C683R also resulted in a significant hyperpolarizing shift in the voltage of steady‐state activation (−65.4 ± 3.0 vs. −57.2 ± 4.8 mV; P < 0.001), resulting in an increased window current compared with WT. The loss‐of‐function effect of NaV1.5/C683R was characterized by significantly increased closed‐state inactivation compared with NaV1.5/WT (P < 0.05). C683R is a novel SCN5A variant resulting in a co‐association of gain‐of‐function and loss‐of‐function properties of the cardiac sodium channel NaV1.5. The phenotype is characterized by adrenaline‐triggered ventricular arrhythmias. Electrophysiological properties and clinical manifestations are different from long‐QT‐3 or Brugada syndrome and might represent a distinct inherited arrhythmia syndrome. [ABSTRACT FROM AUTHOR]

Arachidonic acid can be metabolized by cytochrome P450 (CYP450) enzymes in a tissue- and cell-specific manner to generate vasoactive products such as epoxyeicosatrienoic acids (EETs-cardioprotective) and hydroxyeicosatetraenoic acids (HETEs-cardiotoxic). Type II diabetes is a well-recognized risk factor for developing cardiovascular disease. A mouse model of Type II diabetes (C57BLKS/J-db/db) was used. After sacrifice, livers and hearts were collected, washed, and snap frozen. Total proteins were extracted. Western blots were performed to assess cardiac CYP2J and hepatic CYP2C, CYP4A, and CYP4F protein expression, respectively. Significant decreases in relative protein expression of cardiac CYP2J and hepatic CYP2C were observed in Type II diabetes animals compared to controls (CYP2J: 0.80 ± 0.03 vs. 1.05 ± 0.06, n = 20, p < 0.001); (CYP2C: 1.56 ± 0.17 vs. 2.21 ± 0.19, n = 19, p < 0.01). In contrast, significant increases in relative protein expression of both hepatic CYP4A and CYP4F were noted in Type II diabetes mice compared to controls (CYP4A: 1.06 ± 0.09 vs. 0.18 ± 0.01, n = 19, p < 0.001); (CYP4F: 2.53 ± 0.22 vs. 1.10 ± 0.07, n = 19, p < 0.001). These alterations induced by Type II diabetes in the endogenous pathway (CYP450) of arachidonic acid metabolism may increase the risk for cardiovascular disease by disrupting the fine equilibrium between cardioprotective (CYP2J/CYP2C-generated) and cardiotoxic (CYP4A/CYP4F-generated) metabolites of arachidonic acid. [ABSTRACT FROM AUTHOR]

Cardiac arrhythmias and ECG abnormalities including bradycardia, prolongation of the QT interval, and atrioventricular ( AV) conduction blocks have been extensively observed with fingolimod, the first marketed oral drug for treating the relapsing-remitting form of multiple sclerosis. This study was aiming to further elucidate the effects of fingolimod on cardiac electrophysiology at three different levels: (i) in vitro, (ii) ex vivo , and (iii) in vivo. (i) Patch-clamp experiments in whole cell configuration were performed on Cav1.2-transfected tsA201 cells exposed to fingolimod-phosphate 100 or 500 nmol/L ( n = 27 cells, total) to measure drug effect on L-type calcium current ( ICaL). (ii) Langendorff perfusion experiments were undertaken on male Hartley guinea-pigs isolated hearts ( n = 4) exposed to fingolimod 10 and 100 nmol/L to evaluate drug-induced effects on monophasic action potential duration measured at 90% repolarization ( MAPD90). (iii) Implanted cardiac telemeters were used to record ECGs in guinea-pigs ( n = 7) treated with a single dose of fingolimod 0.0625 mg/kg suspension, administered as an oral gavage. (i) In vitro cellular experiments showed that fingolimod-phosphate causes a concentration-dependent reduction in ICaL. (ii) Ex vivo Langendorff experiments revealed that fingolimod had no significant effect on MAPD90. (iii) Fingolimod caused significant prolongations of the RR, PR, QT, and QTcF intervals in vivo. Reversible AV blocks were also observed in 7/7 animals. Fingolimod possesses ICaL-blocking properties, further contributing to its AV conduction-slowing effects. These properties are also consistent with its mitigated effect on the QT interval in humans, despite previously shown HERG-blocking effect. [ABSTRACT FROM AUTHOR]

CYP3A4, the most abundant cytochrome P450 enzyme in the human liver and small intestine, is responsible for the metabolism of about 50% of all marketed drugs. Numerous pathophysiological factors, such as diabetes and obesity, were shown to affect CYP3A activity. Evidences suggest that drug disposition is altered in type 1 (T1D) and type 2 diabetes (T2D). The objective was to evaluate the effect of T1D and T2D on hepatic and intestinal CYP3a drug-metabolizing activity/expression in mice. Hepatic and intestinal microsomes were prepared from streptozotocin-induced T1D, db/db T2D and control mice. Domperidone was selected as a probe substrate for CYP3a and formation of five of its metabolites was evaluated using high performance liquid chromatography. Hepatic CYP3a protein and mRNA expression were assessed by Western blot and reverse-transcription quantitative polymerase chain reaction respectively. Hepatic microsomal CYP3a activity was significantly increased in both T1D and T2D groups versus control group. Intestinal CYP3a activity was also significantly increased in both T1D and T2D groups. Moreover, significant increases of both hepatic CYP3a mRNAs and protein expression were observed in both T1D and T2D groups versus control group. Additional experiments with testosterone further validated the increased activity of CYP3a under the effect of both T1D and T2D. Although differences exist in the pathophysiological insults associated with T1D and T2D, our results suggest that these two distinct diseases may have the same modulating effect on the regulation of CYP3a, ultimately leading to variability in drug response, ranging from lack of effect to life-threatening toxicity. [ABSTRACT FROM AUTHOR]

Phenyl 4-(2-oxo-3-alkylimidazolidin-1-yl)benzenesulfonates (PAIB-SOs) are a new family of antimitotic prodrugs bioactivated in breast cancer cells expressing CYP1A1. In this study, we report that the 14 C-labeled prototypical PAIB-SO [ 14 C]CEU-818 and its antimitotic counterpart [ 14 C]CEU-602 are distributed in whole mouse body and they show a short half-life in mice. To circumvent this limitation, we evaluated the effect of the homologation of the alkyl side chain of the imidazolidin-2-one moiety of PAIB-SOs. Our studies evidence that PAIB-SOs bearing an n -pentyl side chain exhibit antiproliferative activity in the nanomolar-to-low-micromolar range and a high selectivity toward CYP1A1-positive breast cancer cells. Moreover, the most potent n -pentyl PAIB-SOs were significantly more stable toward rodent liver microsomes. In addition, PAIB-SOs 10 and 14 show significant antitumor activity and low toxicity in chorioallantoic membrane (CAM) assay. Our study confirms that homologation is a suitable approach to improve the rodent hepatic stability of PAIB-SOs.

Background: In humans, CYP3A drug-metabolizing enzyme subfamily is the most important. Numerous pathophysiological factors, such as diabetes and obesity, were shown to affect CYP3A activity. Often considered a precursor state for type II diabetes, metabolic syndrome exerts a modulating role on CYP3A, in our hypothesis., Objective: To evaluate the effect of metabolic syndrome on CYP3A drug-metabolizing activity/expression in guinea pigs., Methods: Hepatic microsomes were prepared from male Hartley guinea pigs fed with a control, a high-fat high sucrose (HFHS) or a high-fat high fructose diet (HFHF). Domperidone was selected as a probe substrate of CYP3A and formation of four of its metabolites was evaluated using high-performance liquid chromatography. CYP3A protein and mRNA expression were assessed by Western blot and reverse-transcription quantitative polymerase chain reaction, respectively. Hepatic fatty infiltration was evaluated using standard Oil Red O staining. Triglyceride and free fatty acid liver content were also quantified., Results: Microsomal CYP3A activity was significantly decreased in both HFHS and HFHF diet groups versus the control diet group. Significant decreases of CYP3A mRNA and protein expression were observed in both HFHS and HFHF diet groups. Oil Red O staining showed a massive liver fatty infiltration in the HFHS and HFHF diet groups, which was not observed in the control diet group. Both triglyceride and free fatty acid liver content were significantly increased in the HFHS and HFHF diet groups., Conclusion: Diet-induced metabolic syndrome decreases CYP3A expression/activity in guinea pigs. This may ultimately lead to variability in drug response, ranging from lack of effect to life-threatening toxicity.

Background: Brugada syndrome (BS) is an electrical cardiac disorder with a right bundle branch block and ST segment elevation in leads V1 to V3 on surface electrocardiograms (ECGs), and is a syndrome that may lead to sudden cardiac death., Purpose: The aim of the present study was to screen for mutations in the SCN5A gene in a family with BS, and to characterize the consequences of the mutation on channel function., Results: A heterozygous nonsense SCN5A mutation (W822X) was identified in the index patient. The mutation was confirmed in the patient's asymptomatic 16-year-old brother and 48-year-old father. The mutation was absent in the index patient's sister and mother. The ECG of the index patient showed a BS type 2 ECG phenotype, which converted into a type 1 ECG phenotype in the presence of flecainide. The ECG of the patient's brother was not typical for BS, but ajmaline treatment unmasked a type 1 ECG phenotype. The ECG of the asymptomatic father was normal at baseline and in the presence of ajmaline. No Na+ currents could be measured in tsA201 cells transfected with W822X mutant channels. Heterozygote expression showed a nearly 50% reduction in Na+ current amplitude with no significant alterations of biophysical properties, indicating a loss of functional Na+ channels, obviously without any dominant-negative activity on wild type channels., Conclusions: The haploinsufficiency of the Nav1.5 protein is the plausible explanation for the clinical BS phenotype in this family. Because the heterozygous W822X mutation theoretically leads to channel expression at one-half of the normal level, the authors suggest that a modifier gene may influence or rescue the phenotype in the asymptomatic family members.