Discovery of Novel TASK-3 Channel Blockers Using a Pharmacophore-Based Virtual Screening.

TASK-3 is a two-pore domain potassium (K2P) channel highly expressed in the hippocampus, cerebellum, and cortex. TASK-3 has been identified as an oncogenic potassium channel and it is overexpressed in different cancer types. For this reason, the development of new TASK-3 blockers could influence the pharmacological treatment of cancer and several neurological conditions. In the present work, we searched for novel TASK-3 blockers by using a virtual screening protocol that includes pharmacophore modeling, molecular docking, and free energy calculations. With this protocol, 19 potential TASK-3 blockers were identified. These molecules were tested in TASK-3 using patch clamp, and one blocker (DR16) was identified with an IC50 = 56.8 ± 3.9 µM. Using DR16 as a scaffold, we designed DR16.1, a novel TASK-3 inhibitor, with an IC50 = 14.2 ± 3.4 µM. Our finding takes on greater relevance considering that not many inhibitory TASK-3 modulators have been reported in the scientific literature until today. These two novel TASK-3 channel inhibitors (DR16 and DR16.1) are the first compounds found using a pharmacophore-based virtual screening and rational drug design protocol.

Evaluating dalfampridine for the treatment of relapsing-remitting multiple sclerosis: does it add to the treatment armamentarium?

Introduction: Multiple sclerosis (MS) is a demyelinating disease, causing axonal damage and disability. Dalfampridine (DAL) is an extended-release formulation of 4-aminopyridine (4AP) and broad-spectrum voltage-dependent potassium channel blocker that is reported to improve motor, visual and cognitive functions. Furthermore, it is presently the only approved drug for walking impairment in MS. Areas covered: Herein, the authors evaluate DAL as a relapsing-remitting MS treatment, reporting and commenting on all aspects of the drug including its chemistry, safety, pharmacokinetics, and cost-effectiveness. A bibliographic search was performed on PubMed using the terms 'dalfampridine OR fampridine OR 4-aminopyridine'. Expert opinion: Evidence from post-marketing studies suggests that DAL, consistent with the effects of 4AP, may not only improve walking speed, but also arm function, fatigue, mood and cognition through restored nerve conduction in central nervous system demyelinated areas. Long-term safety data confirm that the approved dose of 10 mg twice daily is generally well tolerated. However, despite the reported efficacy, the extent of the benefits is limited in real life activities, although significant improvements have been demonstrated in the clinical setting. Patients often complain of side effects (such as cramps and painful paraesthesia) or lack of efficacy. Also, its considerably higher pricing in comparison to 4AP represents an important limitation.

Quantitative determination of talatisamine and its pharmacokinetics and bioavailability in mouse plasma by UPLC-MS/MS.

Talatisamine, as the efficacy ingredient of Aconitum, was known as a novel specific blocker for the delayed rectifier K+ channels in rat hippocampal neurons. In this study, a rapid, selective and reproducible UPLC-MS/MS separation method was established and fully validated for the quantitative determination of talatisamine levels in ICR (Institute of Cancer Research) mouse blood. A total of 24 healthy male ICR mice were divided into four groups that was administered talatisamine via intravenous at a dose of 1 mg/kg and oral administration of three doses (2, 4, 8 mg/kg). All blood samples were protein precipitate by using acetonitrile with an internal standard (IS) deltaline. The effective chromatographic separation was carried out through an UPLC BEH C18 analytical column (2.1 mm × 50 mm, 1.7 µm) with an initial mobile phase that consisted of acetonitrile and 10 mmol/L ammonium acetate aqueous solution (containing 0.1% formic acid) with a gradient elution pumped at a flow rate of 0.4 mL/min. Also, an electrospray ionization (ESI) was applied to quantify the talatisamine in the positive ions mode. The method validation demonstrated good linearity over the range of 1-1000 ng/mL (r2 ≥ 0.9993) for talatisamine in mouse blood with a lower limit of quantification (LLOQ) at 1 ng/mL. The accuracy values of the method were within 89.4% to 113.3%, and the matrix effects were between 103.2% and 106.3%. The mean extraction recoveries for talatisamine obtained from four concentrations of QC blood samples were exceeded 71.7%, and the relative standard deviation (RSD) both of intra- and inter-day precision values for replicate quality control samples did not exceed 15% respectively for all analytes during the assay validation. This method was successfully applied to the evaluation of the pharmacokinetic of talatisamine, regardless of intragastric or intravenous administration in mice. Based on the pharmacokinetics data, the bioavailability of talatisamine in mice was >65.0% after oral administration, exhibiting an excellent oral absorption.

Discovery of Small Molecule Renal Outer Medullary Potassium (ROMK) Channel Inhibitors: A Brief History of Medicinal Chemistry Approaches To Develop Novel Diuretic Therapeutics.

The renal outer medullary potassium (ROMK) channel is a member of the inwardly rectifying family of potassium (Kir, Kir1.1) channels. It is primarily expressed in two regions of the kidney, the cortical collecting duct (CCD) and the thick ascending loop of Henle (TALH). At the CCD it tightly regulates potassium secretion while controlling potassium recycling in TALH. As loss-of-function mutations lead to salt wasting and low blood pressure, it has been surmised that inhibitors of ROMK would represent a target for new and improved diuretics for the treatment of hypertension and heart failure. In this review, we discuss and provide an overview of the medicinal chemistry approaches toward the development of small molecule ROMK inhibitors over the past decade.

Treatment of internuclear ophthalmoparesis in multiple sclerosis with fampridine: A randomized double-blind, placebo-controlled cross-over trial.

AIM: To examine whether the velocity of saccadic eye movements in internuclear ophthalmoparesis (INO) improves with fampridine treatment in patients with multiple sclerosis (MS). METHODS: Randomized, double-blind, placebo-controlled, cross-over trial with fampridine in patients with MS and INO. Horizontal saccades were recorded at baseline and at multiple time points post-dose. Main outcome measures were the change of peak velocity versional dysconjugacy index (PV-VDI) and first-pass amplitude VDI (FPA-VDI). Both parameters were compared between fampridine and placebo using a mixed model analysis of variance taking patients as their own control. Pharmacokinetics was determined by serial blood sampling. RESULTS: Thirteen patients had a bilateral and 10 had a unilateral INO. One patient had an INO of abduction (posterior INO of Lutz) and was excluded. Fampridine significantly reduced both PV-VDI (-17.4%, 95% CI: -22.4%, -12.1%; P < 0.0001) and FPA-VDI (-12.5%, 95% CI: -18.9%, -5.5%; P < 0.01). Pharmacokinetics demonstrated that testing coincided with the average tmax at 2.08 hours (SD 45 minutes). The main adverse event reported after administration of fampridine was dizziness (61%). CONCLUSION: Fampridine improves saccadic eye movements due to INO in MS. Treatment response to fampridine may gauge patient selection for inclusion to remyelination strategies in MS using saccadic eye movements as primary outcome measure.

Comparative Randomized, Single-Dose, Two-Way Crossover Open-Label Study to Determine the Bioequivalence of Two Formulations of Dalfampridine Tablets.

Dalfampridine is a medication that is approved by the US Food and Drug Administration to improve walking impairments in patients with multiple sclerosis (MS). The branded dalfampridine is enormously expensive; hence, the availability of generic dalfampridine will provide better access to the medication, especially for uninsured patients with MS. Bioequivalence studies are demanded by the regulatory authorities to allow the marketing of new generics of dalfampridine. The aim of this study was to assess the bioavailability of the generic (test) and branded (reference) formulations of 10 mg dalfampridine of extended-release tablets after oral administration to healthy adults under fed conditions. The current report methodology was based on a comparative, randomized, single-dose, 2-way crossover open-label study design. Twenty-seven subjects were given a single dose of the test dalfampridine tablet and completed the clinical study. The pharmacokinetic parameters Cmax and AUC0→t, Kel , AUC0→∞ , tmax , and t1/2el were estimated to prove bioequivalence. The confidence intervals for the log-transformed test/reference ratios for dalfampridine 100.96% (97.09%-104.97%) and 99.77% (95.81%-103.87%) for Cmax and AUC0→∞ , respectively, were within the allowed limit specified by the regulatory authorities (80%-125%). Hence, clinically, the test tablet can be prescribed as an alternative to the reference for the indication of improving walking impairments in patients with MS.

A Quarter of a Century Later: What is Dofetilide's Clinical Role Today?

Dofetilide is a class III antiarrhythmic agent approved by the Food and Drug Administration for the conversion of atrial fibrillation and atrial flutter and maintenance of sinus rhythm in symptomatic patients with persistent arrhythmia. Drug trials showed neutral mortality in post-myocardial infarction patients and those with heart failure. This is a review of postmarket data, including real-world efficacy and safety in a variety of populations. Dofetilide has been used off-label with success in patients with paroxysmal atrial fibrillation and atrial flutter, as well as atrial tachycardia and ventricular tachycardia. The real-world acute conversion rate of atrial fibrillation and atrial flutter is higher than that reported in clinical trials. Dofetilide has an acceptable safety profile when initiated (or reloaded) under hospital monitoring and dosed according to creatinine clearance. Dofetilide is well tolerated and a good choice for patients with acceptable renal function and a normal QT interval, especially if atrioventricular nodal blockade needs to be avoided.

Fampridine is a Substrate and Inhibitor of Human OCT2, but not of Human MATE1, or MATE2K.

PURPOSE: The renal clearance of fampridine (Fampyra®, or Ampyra®) significantly exceeds the glomerular filtration rate, suggesting active renal secretion is likely the major elimination pathway. The goal of this study was to identify the renal transporters that are involved in the renal active secretion, and elucidate the active renal secretion mechanism of fampridine. METHODS: The uptake of fampridine to HEK-293 cells overexpressing human OCT2, MATE1 or MATE2K was determined in the absence and presence of Cimetidine, the prototypical inhibitor of the transporters. The inhibition potential of fampridine on the renal transporters was evaluated by determining the uptake of TEA and Metformin, the probe substrates of the transporters of OCT2 and MATEs, respectively, in the absence or presence of fampridine. RESULTS: Significant time- and concentration-dependent uptake of fampridine by human OCT2 was observed. The Km and Vmax were determined as 51.0 ± 17.1 µM and 1107 ± 136 pmole/min/106 cells, respectively. Fampridine also inhibited OCT2 mediated uptake of Metformin with estimated IC50 of 66.8 µM. In contrast, there was not significant uptake of fampridine by human MATE1 or MATE2K, and fampridine did not inhibit MATE1 or MATE2K mediated uptake of TEA. CONCLUSION: The studies indicated fampridine is a substrate and inhibitor of OCT2, but not MATE1 or MATE2K. Results from the study suggested the active renal secretion of fampridine is mediated by human OCT2 but not MATE1 or MATE2K. To our knowledge, fampridine is the first reported substrate specific to OCT2 but not to MATE1 or MATE2K.

4-Chloro-3-nitro-N-butylbenzenesulfonamide acts on KV3.1 channels by an open-channel blocker mechanism.

The effects of 4-chloro-3-nitro-N-butylbenzenesulfonamide (SMD2) on KV3.1 channels, heterologous expressed in L-929 cells, were studied with the whole cell patch-clamp technique. SMD2 blocks KV3.1 in a reversible and use-dependent manner, with IC50 around 10 µM, and a Hill coefficient around 2. Although the conductance vs. voltage relationship in control condition can be described by a single Boltzmann function, two terms are necessary to describe the data in the presence of SMD2. The activation and deactivation time constants are weakly voltage dependent both for control and in the presence of SMD2. SMD2 does not change the channel selectivity and tail currents show a typical crossover phenomenon. The time course of inactivation has a fast and a slow component, and SMD2 significantly decreased their values. Steady-state inactivation is best described by a Boltzmann equation with V 1/2 (the voltage where the probability to find the channels in the inactivated state is 50%) and K (slope factor) equals to -22.9 ± 1.5 mV and 5.3 ± 0.9 mV for control, and -30.3 ± 1.3 mV and 6 ± 0.8 mV for SMD2, respectively. The action of SMD2 is enhanced by high frequency stimulation, and by the time the channel stays open. Taken together, our results suggest that SMD2 blocks the open conformation of KV3.1. From a pharmacological and therapeutic point of view, N-alkylsulfonamides may constitute a new class of pharmacological modulators of KV3.1.

Distribution and kinetics of the Kv1.3-blocking peptide HsTX1[R14A] in experimental rats.

The peptide HsTX1[R14A] is a potent and selective blocker of the voltage-gated potassium channel Kv1.3, which is a highly promising target for the treatment of autoimmune diseases and other conditions. In order to assess the biodistribution of this peptide, it was conjugated with NOTA and radiolabelled with copper-64. [64Cu]Cu-NOTA-HsTX1[R14A] was synthesised in high radiochemical purity and yield. The radiotracer was evaluated in vitro and in vivo. The biodistribution and PET studies after intravenous and subcutaneous injections showed similar patterns and kinetics. The hydrophilic peptide was rapidly distributed, showed low accumulation in most of the organs and tissues, and demonstrated high molecular stability in vitro and in vivo. The most prominent accumulation occurred in the epiphyseal plates of trabecular bones. The high stability and bioavailability, low normal-tissue uptake of [64Cu]Cu-NOTA-HsTX1[R14A], and accumulation in regions of up-regulated Kv channels both in vitro and in vivo demonstrate that HsTX1[R14A] represents a valuable lead for conditions treatable by blockade of the voltage-gated potassium channel Kv1.3. The pharmacokinetics shows that both intravenous and subcutaneous applications are viable routes for the delivery of this potent peptide.

Population Pharmacokinetics/Pharmacodynamics of 3,4-Diaminopyridine Free Base in Patients With Lambert-Eaton Myasthenia.

Lambert-Eaton myasthenia (LEM) is a rare autoimmune disorder associated with debilitating muscle weakness. There are limited treatment options and 3,4-diaminopyridine (3,4-DAP) free base is an investigational orphan drug used to treat LEM-related weakness. We performed a population pharmacokinetic/pharmacodynamic (PK/PD) analysis using 3,4-DAP and metabolite concentrations collected from a phase II study in patients with LEM. The Triple Timed Up & Go (3TUG) assessment, which measures lower extremity weakness, was the primary outcome measure. A total of 1,270 PK samples (49 patients) and 1,091 3TUG data points (32 randomized patients) were included in the PK/PD analysis. A two-compartment and one-compartment model for parent and metabolite, respectively, described the PK data well. Body weight and serum creatinine partially explained the variability in clearance for the final PK model. A fractional inhibitory maximum effect (Emax ) model characterized the exposure-response relationship well. The PK/PD model was applied to identify a suggested dosing approach for 3,4-DAP free base.

Acetylator Status Impacts Amifampridine Phosphate (Firdapse™) Pharmacokinetics and Exposure to a Greater Extent Than Renal Function.

PURPOSE: The purpose of this study is to evaluate safety, tolerability, and pharmacokinetic (PK) properties of amifampridine phosphate (Firdapse™) and its major inactive 3-N-acetyl metabolite in renally impaired and healthy individuals with slow acetylator (SA) and rapid acetylator (RA) phenotypes. METHODS: This was a Phase I, multicenter, open-label study of the PK properties and safety profile of amifampridine phosphate in individuals with normal, mild, moderate, or severely impaired renal function. Amifampridine phosphate was given as a single 10 mg (base equivalent) dose, and the plasma and urine PK properties of amifampridine and its 3-N-acetyl metabolite were determined. The safety profile was evaluated by monitoring adverse events (AEs), clinical laboratory tests, and physical examinations. FINDINGS: Amifampridine clearance was predominantly metabolic through N-acetylation, regardless of renal function in both acetylator phenotypes. In individuals with normal renal function, mean renal clearance represented approximately 3% and 18% of the total clearance of amifampridine in RA and SA, respectively. Large differences in amifampridine exposure were observed between acetylation phenotypes across renal function levels. Mean amifampridine exposure values of AUC0-∞ and Cmax were up to 8.8-fold higher in the SA group compared with the RA group across renal function levels. By comparison, mean AUC0-∞ was less affected by renal function within an acetylator group, only 2- to 3-fold higher in individuals with severe renal impairment (RI) compared with those with normal renal function. Exposure to amifampridine in the SA group with normal renal function was higher (AUC0-∞, approximately 1.8-fold; Cmax, approximately 4.1-fold) than the RA group with severe RI. Exposure to the inactive 3-N-acetyl metabolite was higher than amifampridine in both acetylator groups, independent of renal function level. The metabolite is cleared by renal excretion, and exposure was clearly dependent on renal function with 4.0- to 6.8-fold increases in AUC0-∞ from normal to severe RI. No new tolerability findings were observed. IMPLICATIONS: A single dose of 10 mg of amifampridine phosphate was well tolerated, independent of renal function and acetylator status. The results indicate that the PK profile of amifampridine is affected by metabolic acetylator phenotype to a greater extent than by renal function level, supporting Firdapse™ administration in individuals with RI in line with current labeling recommendations. Amifampridine should be dosed to effect per the individual patient need, altering administration frequency and dose in normal through severe RI. The therapeutic dose of amifampridine phosphate should be tailored to the individual patient needs by gradual dose titration up to the present maximum recommended dose (60-80 mg/day) or until dose-limiting AEs intervene to avoid overdosing and underdosing. EudraCT identifier: 2013-005349-35.

Improvement of hERG-ROMK index of spirocyclic ROMK inhibitors through scaffold optimization and incorporation of novel pharmacophores.

SAR in the previously described spirocyclic ROMK inhibitor series was further evolved from lead 4 by modification of the spirocyclic core and identification of novel right-side pharmacophores. In this process, it was discovered that the spiropyrrolidinone core with the carbonyl group α to the spirocenter was preferred for potent ROMK activity. Efforts aimed at decreasing hERG affinity within the series led to the discovery of multiple novel right-hand pharmacophores including 3-methoxythiadiazole, 2-methoxypyrimidine, and pyridazinone. The most promising candidate is pyridazinone analog 32 that showed an improved functional hERG/ROMK potency ratio and preclinical PK profile. In vivo evaluation of 32 demonstrated blood pressure lowering effects in the spontaneously hypertensive rat model.

Prolonged immunomodulation in inflammatory arthritis using the selective Kv1.3 channel blocker HsTX1[R14A] and its PEGylated analog.

Effector memory T lymphocytes (TEM cells) that lack expression of CCR7 are major drivers of inflammation in a number of autoimmune diseases, including multiple sclerosis and rheumatoid arthritis. The Kv1.3 potassium channel is a key regulator of CCR7- TEM cell activation. Blocking Kv1.3 inhibits TEM cell activation and attenuates inflammation in autoimmunity, and as such, Kv1.3 has emerged as a promising target for the treatment of TEM cell-mediated autoimmune diseases. The scorpion venom-derived peptide HsTX1 and its analog HsTX1[R14A] are potent Kv1.3 blockers and HsTX1[R14A] is selective for Kv1.3 over closely-related Kv1 channels. PEGylation of HsTX1[R14A] to create a Kv1.3 blocker with a long circulating half-life reduced its affinity but not its selectivity for Kv1.3, dramatically reduced its adsorption to inert surfaces, and enhanced its circulating half-life in rats. PEG-HsTX1[R14A] is equipotent to HsTX1[R14A] in preferential inhibition of human and rat CCR7- TEM cell proliferation, leaving CCR7+ naïve and central memory T cells able to proliferate. It reduced inflammation in an active delayed-type hypersensitivity model and in the pristane-induced arthritis (PIA) model of rheumatoid arthritis (RA). Importantly, a single subcutaneous dose of PEG-HsTX1[R14A] reduced inflammation in PIA for a longer period of time than the non-PEGylated HsTX1[R14A]. Together, these data indicate that HsTX1[R14A] and PEG-HsTX1[R14A] are effective in a model of RA and are therefore potential therapeutics for TEM cell-mediated autoimmune diseases. PEG-HsTX1[R14A] has the additional advantages of reduced non-specific adsorption to inert surfaces and enhanced circulating half-life.

An Accelerator Mass Spectrometry-Enabled Microtracer Study to Evaluate the First-Pass Effect on the Absorption of YH4808.

14 C-labeled YH4808, a novel potassium-competitive acid blocker, was intravenously administered as a microtracer at 80 µg (11.8 kBq or 320 nCi) concomitantly with the nonradiolabeled oral drug at 200 mg to determine the absolute bioavailability and to assess the effect of pharmacogenomics on the oral absorption of YH4808. The absolute bioavailability was low and highly variable (mean, 10.1%; range, 2.3-19.3%), and M3 and M8, active metabolites of YH4808, were formed 22.6- and 38.5-fold higher after oral administration than intravenous administration, respectively. The product of the fraction of an oral YH4808 dose entering the gut wall and the fraction of YH4808 passing on to the portal circulation was larger in subjects carrying the variants of the CHST3, SLC15A1, and SULT1B1 genes. A combined LC+AMS is a useful tool to construct a rich and highly informative pharmacokinetic knowledge core in early clinical drug development at a reasonable cost.

Measuring kinetics and potency of hERG block for CiPA.

INTRODUCTION: The Comprehensive in vitro Proarrhythmic Assay (CiPA) aims to update current cardiac safety testing to better evaluate arrhythmic risk. A central theme of CiPA is the use of in silico approaches to risk prediction incorporating models of drug binding to hERG. To parameterize these models, accurate in vitro measurement of potency and kinetics of block is required. The Ion Channel Working Group was tasked with: i) selecting a protocol that could measure kinetics of block and was easily implementable on automated platforms for future rollout in industry and ii) acquiring a reference dataset using the standardized protocol. METHODS: Data were acquired using a 'step depolarisation' protocol using manual patch-clamp at ambient temperature. RESULTS: Potency, kinetics and trapping characteristics of hERG block for the CiPA training panel of twelve drugs were measured. Timecourse of block and trapping characteristics could be reliably measured if the time constant for onset of block was between ~500ms and ~15s. Seven drugs, however had time courses of block faster than this cut-off. DISCUSSION: Here we describe the implementation of the standardized protocol for measurement of kinetics and potency of hERG block for CiPA. The results highlight the challenges in identifying a single protocol to measure hERG block over a range of kinetics. The dataset from this study is being used by the In Silico Working Group to develop models of drug binding for risk prediction and is freely available as a 'gold standard' ambient temperature dataset to evaluate variability across high throughput platforms.

Inhibition of the potassium channel KCa3.1 by senicapoc reverses tactile allodynia in rats with peripheral nerve injury.

Neuropathic pain is a debilitating, chronic condition with a significant unmet need for effective treatment options. Recent studies have demonstrated that in addition to neurons, non-neuronal cells such as microglia contribute to the initiation and maintenance of allodynia in rodent models of neuropathic pain. The Ca2+- activated K+ channel, KCa3.1 is critical for the activation of immune cells, including the CNS-resident microglia. In order to evaluate the role of KCa3.1 in the maintenance of mechanical allodynia following peripheral nerve injury, we used senicapoc, a stable and highly potent KCa3.1 inhibitor. In primary cultured microglia, senicapoc inhibited microglial nitric oxide and IL-1ß release. In vivo, senicapoc showed high CNS penetrance and when administered to rats with peripheral nerve injury, it significantly reversed tactile allodynia similar to the standard of care, gabapentin. In contrast to gabapentin, senicapoc achieved efficacy without any overt impact on locomotor activity. Together, the data demonstrate that the KCa3.1 inhibitor senicapoc is effective at reducing mechanical hypersensitivity in a rodent model of peripheral nerve injury.

Pharmacogenetics of Potassium Channel Blockers.

The QT interval on surface electrocardiograms provides a model of a multicomponent integrated readout of many biological systems, including ion channels, modulatory subunits, signaling systems that modulate their activity, and mechanisms that regulate the expression of their responsible genes. The problem of drug exposure causing exaggerated QT interval prolongation and torsades de pointes highlights the multicomponent nature of cardiac repolarization and the way in which simple perturbations can yield exaggerated responses. Future directions will involve cellular approaches coupled to evolving technologies that can interrogate multicellular systems and provide a sophisticated view of mechanisms in this previously idiosyncratic drug reaction.

Increased Late Sodium Current Contributes to the Electrophysiological Effects of Chronic, but Not Acute, Dofetilide Administration.

BACKGROUND: Drugs are screened for delayed rectifier potassium current (IKr) blockade to predict long QT syndrome prolongation and arrhythmogenesis. However, single-cell studies have shown that chronic (hours) exposure to some IKr blockers (eg, dofetilide) prolongs repolarization additionally by increasing late sodium current (INa-L) via inhibition of phosphoinositide 3-kinase. We hypothesized that chronic dofetilide administration to intact dogs prolongs repolarization by blocking IKr and increasing INa-L. METHODS AND RESULTS: We continuously infused dofetilide (6-9 µg/kg bolus+6-9 µg/kg per hour IV infusion) into anesthetized dogs for 7 hours, maintaining plasma levels within the therapeutic range. In separate experiments, myocardial biopsies were taken before and during 6-hour intravenous dofetide infusion, and the level of phospho-Akt was determined. Acute and chronic dofetilide effects on action potential duration (APD) were studied in canine left ventricular subendocardial slabs using microelectrode techniques. Dofetilide monotonically increased QTc and APD throughout 6.5-hour exposure. Dofetilide infusion during ≥210 minutes inhibited Akt phosphorylation. INa-L block with lidocaine shortened QTc and APD more at 6.5 hours than at 50 minutes (QTc) or 30 minutes (APD) dofetilide administration. In comparison, moxifloxacin, an IKr blocker with no effects on phosphoinositide 3-kinase and INa-L prolonged APD acutely but no additional prolongation occurred on chronic superfusion. Lidocaine shortened APD equally during acute and chronic moxifloxacin superfusion. CONCLUSIONS: Increased INa-L contributes to chronic dofetilide effects in vivo. These data emphasize the need to include time and INa-L in evaluating the phosphoinositide 3-kinase inhibition-derived proarrhythmic potential of drugs and provide a mechanism for benefit from lidocaine administration in clinical acquired long QT syndrome.

PK/PD Modelling of the QT Interval: a Step Towards Defining the Translational Relationship Between In Vitro, Awake Beagle Dogs, and Humans.

Inhibiting the human ether-a-go-go-related gene (hERG)-encoded potassium ion channel is positively correlated with QT-interval prolongation in vivo, which is considered a risk factor for the occurrence of Torsades de Pointes (TdP). A pharmacokinetic/pharmacodynamic model was developed for four compounds that reached the clinic, to relate drug-induced QT-interval change in awake dogs and humans and to derive a translational scaling factor a 1. Overall, dogs were more sensitive than humans to QT-interval change, an a 1 of 1.5 was found, and a 10% current inhibition in vitro produced a higher percent QT-interval change in dogs as compared to humans. The QT-interval changes in dogs were predictive for humans. In vitro and in vivo information could reliably describe the effects in humans. Robust translational knowledge is likely to reduce the need for expensive thorough QT studies; therefore, expanding this work to more compounds is recommended.