Safety Pharmacology Models Research Articles

Quantified proarrhythmic potential of selected human embryonic stem cell-derived cardiomyocytes

To improve proarrhythmic predictability of preclinical models, we assessed whether human ventricular-like embryonic stem cell-derived cardiomyocytes (hESC-CMs) can be selected following a standardized protocol. Also, we quantified their arrhythmogenic response and compared this to a contemporary used rabbit Purkinje fiber (PF) model. Multiple transmembrane action potentials (AP) were recorded from 164 hESC-CM clusters (9 different batches), and 12 isolated PFs from New Zealand White rabbits. AP duration (APD), early afterdepolarizations (EADs), triangulation (T), and short-term variability of repolarization (STV) were determined on application of the IKr blocker E-4031 (0.03/0.1/0.3/1 μM). Isoproterenol (0.1 μM) was used to assess adrenergic response. To validate the phenotype, RNA isolated from atrial- and ventricular-like clusters (n=8) was analyzed using low-density Taqman arrays. Based on initial experiments, slow beating rate (<50 bpm) and long APD (>200 ms) were used to select 31 ventricular-like clusters. E-4031 (1 μM) prolonged APD (31/31) and induced EADs only in clusters with APD90>300 ms (11/16). EADs were associated with increased T (1.6±0.2 vs 2.0±0.3⁎) and STV (2.7±1.5 vs 6.9±1.9⁎). Rabbit PF reacted in a similar way with regards to EADs (5/12), increased T (1.3±0.1 vs 1.9±0.4⁎), and STV (1.2±0.9 vs 7.1±5.6⁎). According to ROC values, hESC-CMs (STV 0.91) could predict EADs at least equivalent to PF (STV 0.69). Isoproterenol shortened APD and completely suppressed EADs. Gene expression analysis revealed that HCN1/2, KCNA5, and GJA5 were higher⁎ in atrial/nodal-like cells, whereas KCNJ2 and SCN1B were higher⁎ in ventricular-like cells (⁎P<0.05). Selection of hESC-CM clusters with a ventricular-like phenotype can be standardized. The proarrhythmic results are qualitatively and quantitatively comparable between hESC-CMs and rabbit PF. Our results indicate that additional validation of this new safety pharmacology model is warranted.

Video-electroencephalography in conscious monkeys using telemetry and computer analysis: Refinement of a safety pharmacology model

Video-electroencephalography in conscious monkeys using telemetry and computer analysis: Refinement of a safety pharmacology model

A framework to assess the translation of safety pharmacology data to humans

This article outlines a strategy for collecting accurate data for the determination of the sensitivity, specificity and predictive value of safety pharmacology models. This entails performing a retrospective analysis on commonly used safety pharmacology endpoints and an objective assessment of new non-clinical models. Such assessments require a systematic quantitative analysis of safety pharmacology parameters as well as clinical Phase I adverse events. Once the sensitivity, specificity and predictive capacity of models have been determined, they can be aligned within specific phases of the drug discovery and development pipeline for maximal impact, or removed from the screening cascade altogether. Furthermore, data will contribute to evidence-based decision-making based on the knowledge of the model sensitivity and specificity. This strategy should therefore contribute to the reduction of candidate drug attrition and a more appropriate use of animals. More data are needed to increase the power of analysis and enable more accurate comparisons of models e.g. pharmacokinetic/phamacodynamic (PK/PD) relationships as well as non-clinical and clinical outcomes for determining concordance. This task requires the collaboration and agreement of pharmaceutical companies to share data anonymously on proprietary and candidate drugs.

Video-electroencephalography in conscious non human primate using radiotelemetry and computerized analysis: Refinement of a safety pharmacology model

Introduction Electroencephalography (EEG) investigations are occasionally required as follow-up studies for safety pharmacology core battery (S7A). Video-EEG monitoring is a standard diagnostic tool in humans but limited data is available on its use in telemetered freely moving macaque monkeys for safety pharmacology investigations. While proconvulsant risk evaluations are routinely conducted in rodents, pharmacological or pharmacokinetic considerations lead to the use of non human primates in toxicology and safety pharmacology in some cases. Methods Cynomolgus monkeys were instrumented with telemetry implants. Placement of EEG electrode was based on the 10–20 system using three derivations (C3-O1, Cz-Oz and C4-O2). EEG trace analysis was carried out using NeuroScore software. After 24 h of continuous video-EEG monitoring, animals received pentylenetetrazole (PTZ, 10 mg/kg/15 min) until convulsions were noted. Convulsions were immediately treated with diazepam (1.0 mg/kg). A seizure detection protocol with a dynamic spike train threshold was used for the entire EEG monitoring period (total of 44 h) including periods when PTZ was administered. Spectral analysis was done to quantify the absolute and relative amplitude of EEG frequency bands (delta, theta, alpha, sigma and beta waves). Sleep stages were quantified and EEGs during seizures were analyzed using fast Fourier transformation (FFT) to assess dominant frequencies. Results Spike trains were detected by computerized analysis in all animals presenting PTZ-induced seizures while paroxysmal activities were systematically predictive (at least 4-min prior to generalized seizures). Beta activity increased with visual stimulation using monkey treats. Characteristics of EEG for all sleep stages (I, II, III and IV) were present in all animals. Delta activity was predominant in normal awake EEG as well as in all sleep stages. Seizure peak frequency was 3–6 Hz on FFT, corresponding to the discharge of the underlying generator. Discussion EEG-video monitoring can be useful when using non human primates to characterize neurological adverse effects with unpredictable onset. Computerized video-EEG analysis was a valuable tool for safety pharmacology investigations including proconvulsant risk assessment, spectral analysis of frequency bands and sleep stage determination.

Improvement of cardiac efficacy and safety models in drug discovery by the use of stem cell-derived cardiomyocytes

Background: The pharmaceutical industry suffers from high attrition rates during late phases of drug development. Improved models for early evaluation of drug efficacy and safety are needed to address this problem. Recent developments have illustrated that human stem cell-derived cardiomyocytes are attractive for using as a model system for different cardiac diseases and as a model for screening, safety pharmacology and toxicology. Objective: In this review, we discuss contemporary drug discovery models and their characteristics for cardiac efficacy testing and safety assessment. Additionally, we evaluate various sources of stem cells and how these cells could potentially improve early screening and safety models. Conclusion: We conclude that human stem cells offer a source of physiologically relevant cells that show great potential as a future tool in cardiac drug discovery. However, some technical challenges related to cell differentiation and production and also to validation of improved platforms remain and must be overcome before successful application can become a reality.

Conscious and anesthetized non-human primate safety pharmacology models: Hemodynamic sensitivity comparison

Introduction Drug-induced cardiovascular effects identified in conscious cynomolgus monkeys equipped with tethers and prepared for radiotelemetry were compared with results from anesthetized non-human primate (cynomolgus and rhesus) models. Methods Remifentanil (4.0 μg/kg, bolus), esmolol (2.0 mg/kg, bolus) and dopamine (0.05 mg/kg/min, 30 min infusion) were given intravenously to all models. Results Remifentanil decreased heart rate (HR), systolic, mean and diastolic systemic arterial pressures (SAP) in anesthetized animals while conscious monkeys presented an increase in HR, systolic, mean and diastolic SAP, as seen in humans for the respective state of consciousness (conscious and anesthetized). Esmolol decreased HR, systolic, mean and diastolic SAP in anesthetized monkeys while only HR, systolic and mean SAP achieved a statistically significant decrease in the conscious model. The amplitude of SAP reduction was greater in anesthetized models, while the amplitude of HR reduction was greater in the conscious and anesthetized cynomolgus models than in the anesthetized rhesus model. Dopamine induced a significant increase in HR, systolic, mean and diastolic SAP in anesthetized models without any statistically significant effect on HR and SAP in the conscious model. Discussion The amplitude of hemodynamic and chronotropic alterations induced by positive control drugs was generally greater in anesthetized than in conscious models and statistical significance was achieved more often with the anesthetized models. These results suggest that an anesthetized model may be valuable as part of a drug screening program for cardiovascular safety evaluations in addition to a conscious model.

Validation of respiratory safety pharmacology models: Conscious and anesthetized beagle dogs

Introduction Installation, operation and performance qualifications were performed on a test system for respiratory monitoring. Methods For performance qualification, conscious dogs received saline (0.2 mL/kg, iv, n = 12), albuterol (100 μg/kg, inhalation, n = 5), methacholine (2.0 and 8.0 μg/kg, iv, n = 8) and remifentanil (4.0 μg/kg, iv, n = 7). Following anesthesia with propofol infusion, dogs received saline (iv, n = 15), albuterol (100 μg/kg, inhalation, n = 8), methacholine (8.0 μg/kg, iv, n = 8), remifentanil (4.0 μg/kg, iv, n = 7), and cholecystokinine tetrapeptide (CCK-4) (10 μg/kg, iv, n = 7) and were exposed to hypoxic gas mixture (10% oxygen) ( n = 12). Results Saline had no significant respiratory effect. Albuterol increased tidal volume (TV) (+ 28%, p < 0.05) and minute ventilation (MV) (+ 96%, p < 0.01) in conscious dogs. In anesthetized dogs, MV was significantly increased (+ 23%, p < 0.05) but the difference was not statistically significant for TV and respiratory rate (RR). Methacholine at 2.0 μg/kg increased MV (+ 45%, p < 0.01) in conscious animals while 8.0 μg/kg increased RR (+ 66%, p < 0.01), TV (+ 24%, p < 0.05) and MV (+ 88%, p < 0.05). In anesthetized dogs, methacholine increased RR (+ 51%, p < 0.05), MV (+ 34%, p < 0.05), lung elastance (+ 36.9%, p < 0.01), and resistance (+ 45.8%, p < 0.01). Remifentanil decreased MV in conscious dogs (− 68%, p < 0.01) while transient apnea was observed in all anesthetized dogs. CCK-4 increased RR (+ 328%, p < 0.01) and MV (+ 127%, p < 0.05) and decreased TV (− 58%, p < 0.01). Exposure to hypoxic gas mixture increased MV and RR ( p < 0.01). Baseline MV was lower ( p < 0.05) in anesthetized than in conscious dogs. Discussion Arterial blood gas values, particularly SaO 2, presented a limited sensitivity to detect any ventilation disturbance, but allowed confirmation of both ventilatory compensatory phenomenon (when present) and initial pharmacologic drug effect. These results also highlight the greater sensitivity of the conscious model when compared to anesthetized dogs.

Investigation of the mechanism by which ketanserin increases cardiac action potential duration

In the current ICH S7B guideline, in vitro evaluation of proarrhythmic liability is limited to the risk of QT interval prolongation, whilst the effect of new chemical entities on cardiac conductivity is often overlooked. The aim of this work was to compare the effects of the sodium channel blocker, lidocaine in three in vitro safety pharmacology models: hNav1.5 channel test, atrial action potential (AP) and Purkinje fiber AP and to identify the most sensitive model for detecting cardiac conduction slowing.Whole-cell patch‐clamp methods were used to record the sodium current (INa) encoded by hNav1.5 in stably transfected HEK293 cells at ambient temperature. Transmembrane APs were recorded in rabbit Purkinje fibers and rabbit and guinea-pig left stimulated atria at physiological temperature. Parameters involved in depolarization or repolarization were reported.Lidocaine (from 10 to 1000 μM) decreased the amplitude of INa (IC50: 256 ± 37 μM) in a concentration-dependent manner. In the Purkinje fiber assay, lidocaine (10, 30 and 100 μM) had no effects on maximal upstroke velocity (Vmax), but shortened AP duration at 90% repolarization (APD90). At 30 and 100 μM, lidocaine also increased AP triangulation. In guinea-pig atria, lidocaine decreased Vmax starting from 30 μM and conduction velocity (CV) at 100 μM, but had no effects on other parameters. In rabbit atria, lidocaine decreased Vmax and CV at 100 μM without affecting APD90. The effects of 100 μM lidocaine on Vmax and CV were more marked in rabbit than in guinea-pig atria.Rabbit atria are more sensitive than rabbit Purkinje fibers or guinea-pig atria for detecting lidocaine-induced cardiac conduction slowing. These data suggest that isolated rabbit atria in addition to the hNav1.5 channel assay could be relevant models to predict drug-induced conduction slowing.

Assessment of absolute volume from diameter of the intact canine left ventricular cavity.

Assessment of absolute volume from diameter of the intact canine left ventricular cavity.