Search

JNJ-10450232 (NTM-006) is a new molecular entity that is structurally related to acetaminophen. A comprehensive non-clinical safety program was conducted to support first-in-human and clinical efficacy studies based on preclinical data suggesting that the compound has comparable or enhanced antinociceptive and antipyretic efficacy without causing hepatotoxicity at supratherapeutic doses. No hepatic toxicity was noted in a mouse model sensitive to acetaminophen hepatotoxicity or in rats, dogs, and non-human primates in 28-day repeat dose toxicity studies at and above doses/exposures at which acetaminophen is known to cause hepatotoxicity. In the 28-day toxicity studies, all treatment-related findings were monitorable and reversible. Methemoglobinemia, which was observed in dogs and to a lesser extent in rats, is also observed with acetaminophen. This finding is considered not relevant to humans due to species differences in metabolism. Thyroid hypertrophy and hyperplasia were also observed in dogs and were shown to be a consequence of a species-specific UGT induction also demonstrated with increased thyroid hormone metabolism. Indirect bilirubin elevation was observed in rats as a result of UGT1A1 Inhibition. JNJ-10450232 (NTM-006) had no toxicologically relevant findings in safety pharmacology or genotoxicity studies. Together, these data supported progressing into safety and efficacy studies in humans., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Johnson & Johnson employed all authors at the time of the study, and each author has owned, or currently owns, Johnson & Johnson stock., (Copyright © 2023. Published by Elsevier Inc.)

Introduction: People with epilepsy are at heightened risk of sudden death compared to the general population. The leading cause of epilepsy-related premature mortality is a sudden unexpected death in epilepsy (SUDEP). The mechanism of SUDEP remains largely unresolved and the lack of preclinical models to study the potential mechanism underlying SUDEP is a problem., Method: By combining electroencephalographic (EEG) and electrocardiogram (ECG) measurements within a well described LQT1 dog model, we investigated the effect of the proconvulsive compound pentylenetetrazol (PTZ), and its link to the induction of Torsades de Pointes (TdP)., Results: Pre-treatment with the potent and selective I Ks blocker JNJ 282 induced a pronounced QT (QTc) prolongation in anaesthetized dogs (Long QT syndrome type 1 or LQT1 group) compared to dogs that were not treated (control group). Subsequent PTZ administration induced spiking on the EEG signal and seizures in both groups, but only R-on-T, salvo and TdP were observed in dogs of the LQT1 group., Conclusion: Our results show that a proconvulsive drug can trigger TdP-like cardiac arrhythmias, in conditions of compromised repolarization in the heart (I ks blockade). In man, TdP arrythmia's can often lead to ventricular fibrillation (VF) and sudden death. This observation suggests that long QT-intervals (genetic or drug induced) could potentially be one of the risk factors for SUDEP in epileptic patients., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Introduction: Calcium-based screening of hiPS-CMs is a useful preclinical safety evaluation platform with the ability to generate robust signals that facilitates high-throughput screening and data analysis. However, due to the potential inherent toxicities, it is important to understand potential effects of different calcium-sensitive dyes on the hiPS-CMs model., Methods: We compared three calcium-sensitive fluorescence dyes (Cal520, ACTOne and Calcium 5) for their impact on the variability, the beating properties and the pharmacological responses of hiPS-CMs using the Hamamatsu FDSS/μCell imaging platform. Direct effects of three dyes on the electrophysiological properties of hiPS-CMs were evaluated with the multi-electrode array (MEA) Axion Maestro platform., Results: We propose a specific experimental protocol for each dye which gives the most optimal assay conditions to minimize variability and possible adverse effects. We showed that Cal520 had the smallest effect on hiPS-CMs together with the longest-lasting stable amplitude signal (up to 4 h). Although all dyes had a (minor) acute effect on hiPS-CMs, in the form of reduced beat rate and prolonged field potential duration, the selection of the dye did not influence the pharmacological response of four cardioactive drugs (dofetilide, moxifloxacin, nimodipine and isoprenaline)., Discussion: In conclusion, we have documented that different calcium sensitive dyes have only minor direct (acute) effects on hiPS-CMs with Cal520 showing the least effects and the longest lasting signal amplitude. Importantly, drug-induced pharmacological responses in hiPS-CMs were comparable between the three dyes. These findings should help further improve the robustness of the hiPS-CMs-based calcium transient assay as a predictive, preclinical cardiac safety evaluation tool., (Copyright © 2018 Elsevier Inc. All rights reserved.)

In the present study, we investigated an impact of the stimulation rate on the detection of the proarrhythmic potential of 10 reference compounds with effects on different cardiac ion channels in the isolated arterially-perfused rabbit left ventricular wedge preparation. The compounds were tested in the wedge model using two distinct protocols; including baseline stimulation at 1-Hz followed by a brief period at 0.5-Hz, either without an additional brief period of 2-Hz stimulation (i.e. Protocol 1) or with 2-Hz stimulation (i.e. Protocol 2). As expected, QT-prolonging drugs (ibutilide and quinidine) prolonged the QT interval, similarly increased the Torsades de Pointes (TdP) score, and elicited early afterdepolarizations (EADs) in both protocols. HMR1556 and JNJ-303 (IKs blockers) also prolonged the QT interval up to 1μM similarly in both protocols. Nifedipine (Ca(2+) antagonist) shortened the QT interval, and reduced force of contraction similarly in both protocols. However, Na(+) channel blockers (Ia, Ib, Ic) widened the QRS duration more in Protocol 2 than in Protocol 1. Furthermore, it was only possible to detect non-TdP-like ventricular tachycardia/fibrillation (VT/VF) induced by Na(+) blockers and by QT-shortening drugs (levcromakalim and mallotoxin) using the 2-Hz stimulation (Protocol 2). Our data suggest that the inclusion of a brief period of fast stimulation at 2Hz is critical for detecting drug-induced slowing of conduction (QRS widening), QT shortening and associated (non-TdP-like) VT/VF, which are distinct from the QT prolongation/TdP proarrhythmia in isolated, arterially-perfused rabbit left ventricular wedges., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Introduction: With the recent development of more sensitive biomarkers to assess kidney injury preclinically, a survey was designed i) to investigate what strategies are used to investigate renal toxicity in both ICH S7A compliant Safety Pharmacology (SP) studies after a single dose of a compound and within repeat-dose toxicity studies by large pharmaceutical companies today; ii) to understand whether renal SP studies have impact or utility in drug development and/or if it may be more appropriate to assess renal effects after multiple doses of compounds; iii) to ascertain how much mechanistic work is performed by the top 15 largest pharmaceutical companies (as determined by R&D revenue size); iv) to gain an insight into the impact of the validation of DIKI biomarkers and their introduction in the safety evaluation paradigm; and v) to understand the impact of renal/urinary safety study data on progression of projects., Methods: Two short anonymous surveys were submitted to SP leaders of the top 15 pharmaceutical companies, as defined by 2012 R&D portfolio size. Fourteen multiple choice questions were designed to explore the strategies used to investigate renal effects in both ICH S7A compliant SP studies and within toxicology studies., Results: A 67% and 60% response rate was obtained in the first and second surveys, respectively. Nine out of ten respondent companies conduct renal excretory measurements (eg. urine analysis) in toxicology studies whereas only five out of ten conduct specific renal SP studies; and all of those 5 also conduct the renal excretory measurements in toxicology studies. These companies measure and/or calculate a variety of parameters as part of these studies, and also on a case by case basis include regulatory qualified and non-qualified DIKI biomarkers. Finally, only one company has used renal/urinary functional data alone to stop a project, whereas the majority of respondents combine renal data with other target organ assessments to form an integrated decision-making set., Conclusion: These short surveys highlighted areas of similarity: a) urinary measurements are most commonly taken on repeat-dose toxicity studies, and b) renal SP studies are less often utilised. The two major differences are a) lack of consistent use of DIKI biomarkers in urinary safety studies and b) the way large pharmaceutical companies assess renal function. Finally, suggestions were made to improve the safety assessment methods for determining the safety of compounds with potential renal liability., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Introduction: Preclinical proarrhythmic risk assessment of drug candidates is focused predominantly on arrhythmias arising from repolarization abnormalities. However, drug-induced cardiac conduction slowing is associated with significant risk of life-threatening ventricular arrhythmias, particularly in a setting of cardiac ischemia. Therefore, we optimized and characterized an anesthetized dog model to evaluate the potential proarrhythmic risk of drug candidates in ischemic heart disease patients., Methods: Anesthetized dogs were instrumented with atrial and ventricular epicardial electrodes for pacing and measurement of conduction times, and a balloon occluder and flow probe placed around the left anterior descending coronary artery (LAD) distal to the first branch. Conduction times, ECG intervals and incidence of arrhythmias were assessed serially at the end of each dose infusion (flecainide: 0.32, 0.63, 1.25, 2.5 and 5mg/kg, i.v.; dofetilide:1.25, 2.5, 5, 10 and 20 μg/kg, i.v.; or vehicle; n=6/group) both during normal flow (with and without rapid pacing) and during 5-min LAD occlusion (with and without rapid pacing). Compound X, a development candidate with mild conduction slowing activity, was also evaluated., Results: Flecainide produced pronounced, dose-dependent slowing of conduction that was exacerbated during ischemia and rapid pacing. In addition, ventricular tachycardia (VT) and fibrillation (VF) occurred in 4 of 6 dogs (3 VF @ 0.63 mg/kg; 1VT @ 2.5mg/kg). In contrast, no animals in the vehicle group developed arrhythmias. Dofetilide, a potent IKr blocker that does not slow conduction, prolonged QT interval but did not cause further conduction slowing during ischemia with or without pacing and there were no arrhythmias. Compound X, like flecainide, produced marked conduction slowing and arrhythmias (VT, VF) during ischemia and pacing., Discussion: This model may be useful to more accurately define shifts in safety margins in a setting of ischemia and increased cardiac demand for drugs that slow conduction., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Introduction: Preclinical concentration-effect (pharmacokinetic-pharmacodynamic, PKPD) modeling has successfully quantified QT effects of several drugs known for significant QT prolongation. This study investigated its sensitivity for detecting small magnitudes of QT-prolongation in a typical preclinical cardiovascular (CV) safety study in the conscious telemetered dog (crossover study in 4-8 animals receiving a vehicle and three dose levels). Results were compared with conventional statistical analysis (analysis of covariance, ANCOVA)., Methods: A PKPD model predicting individual QTc was first developed from vehicle arms of 28 typical CV studies and one positive control study (sotalol). The model quantified between-animal, inter-occasion and within-animal variability and described QTc over 24h as a function of circadian variation and drug concentration. This "true" model was used to repeatedly (n = 500) simulate studies with typical drug-induced QTc prolongation (∆QTc) of 1 to 12 ms at high-dose peak concentrations. Simulated studies were re-analyzed by both PKPD analysis (with varying complexity) and ANCOVA. Sensitivity (power) was calculated as the percentage of studies in which a significant (α = 0.05) drug effect was found. One simulation scenario did not include a concentration-effect relationship and served to investigate false-positive rates. Exposure-effect relationships were derived from both PKPD analysis (linear concentration-effect) and ANCOVA (linear trend test for dose) and compared., Results: PKPD analysis/ANCOVA had a sensitivity of 80% to detect the effects of 7/13 ms (n = 4), 5/10 ms (n = 6) and 4.5/8 ms (n = 8), respectively. The false-positive rate was much higher using ANCOVA (40%) compared to PKPD analysis (1%). Typical drug effects were more precisely predicted using estimated concentration-effect slopes (± 1.5-2.8 ms) than dose-effect slopes (± 3.3-3.7 ms)., Discussion: Preclinical PKPD analysis can increase the confidence in the quantification of small QTc effects and potentially allow reducing the number of animals while maintaining the required study sensitivity. This underscores the value of PKPD modeling in preclinical safety testing., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Introduction: Regarding evaluation of drug-induced changes in left ventricular contractility in safety pharmacology there is still a gap in knowledge between preclinically and clinically used measurements., Methods: As a step towards translation of preclinical to clinical outcomes, this study in telemetered dogs was initiated to compare indexes of contractility, such as LV dP/dt(max) (contractility measured as the maximum raise of pressure in the left ventricle) and LV dP/dt(max)/P (contractility measured as the maximum raise of pressure in the left ventricle, corrected for pressure) (telemetry; both commonly preclinically used) and EF (ejection fraction) and FS (fractional shortening) (echocardiography; both commonly clinically used). Different inotropic states were induced by minoxidil, milrinone, isoprenaline, clonidine, atenolol and verapamil., Results: Both techniques demonstrated reproducible changes in contractility which showed a clear linear association. A change in LV dP/dt(max) of 1000 mmHg/s (in the range of 2500 to 7500 mmHg/s; in healthy dogs) corresponded with a change in ejection fraction of approximately 7% and a fractional shortening of approximately 6%. A change of 10/s LV dP/dt(max)/P (in the range of 35 to 85/s; in healthy dogs) corresponded with a change in ejection fraction of approximately 7% and a fractional shortening of 7%., Discussion: The correlation found in this study could potentially enable a better--translational--assessment of the clinical relevance of changes in contractility indices measured with telemetry devices in preclinical safety studies., (© 2013.)

Introduction: In the present study, we investigated whether a new biomarker - index of cardiac electrophysiological balance (iCEB=QT/QRS) - could predict drug-induced cardiac arrhythmias (CAs), including ventricular tachycardia/ventricular fibrillation (VT/VF) and Torsades de Pointes (TdPs)., Methods: The rabbit left ventricular arterially-perfused-wedge was used to investigate whether the simple iCEB measured from the ECG is reflective of the more difficult measurement of λ (effective refractory period×conduction velocity) for predicting CAs induced by a number of drugs., Results: Dofetilide concentration-dependently increased iCEB and λ, predicting potential risk of drug-induced incidence of early afterdepolarizations (EADs) starting at 0.01μM. Digoxin (1 and 5μM), encainide (5 and 20μM) and propoxyphene (10 and 100μM) markedly reduced both iCEB and λ, predicting their ability to induce non-TdP-like VT/VF. At 10μM, both NS1643 and levcromakalim significantly decreased λ and iCEB, which was preceded with presence of non-TdP-like VT/VF. Isoprenaline (0.05 to 0.5μM) significantly reduced both λ and iCEB, which was associated with a high incidence of non-TdP-like VT/VF in most preparations. Other biomarkers (i.e. transmural dispersion of T-wave and instability of the QT interval) predicted only dofetilide-induced long QT and EADs, but did not predict drug-induced risk of non-TdP-like VT/VF., Discussion: Our data from 7 reference drugs of known pro-arrhythmic effects suggests that 1) this non-invasive iCEB predicts potential risk of drug-induced CAs beyond long QT and TdP; 2) iCEB is more useful than the current biomarkers (i.e. transmural dispersion and instability) in predicting potential risks for drug-induced non-TdP-like VT/VF., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Current methods to measure physiological properties of cardiomyocytes and predict fatal arrhythmias that can cause sudden death, such as Torsade de Pointes, lack either the automation and throughput needed for early-stage drug discovery and/or have poor predictive value. To increase throughput and predictive power of in vitro assays, we developed kinetic imaging cytometry (KIC) for automated cell-by-cell analyses via intracellular fluorescence Ca²⁺ indicators. The KIC instrument simultaneously records and analyzes intracellular calcium concentration [Ca²⁺](i) at 30-ms resolution from hundreds of individual cells/well of 96-well plates in seconds, providing kinetic details not previously possible with well averaging technologies such as plate readers. Analyses of human embryonic stem cell and induced pluripotent stem cell-derived cardiomyocytes revealed effects of known cardiotoxic and arrhythmogenic drugs on kinetic parameters of Ca²⁺ dynamics, suggesting that KIC will aid in the assessment of cardiotoxic risk and in the elucidation of pathogenic mechanisms of heart disease associated with drugs treatment and/or genetic background., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Introduction: How does safety pharmacology operate in large pharmaceutical companies today? By understanding our current position, can we prepare safety pharmacology to successfully navigate the complex process of drug discovery and development?, Methods: A short anonymous survey was conducted, by invitation, to safety pharmacology representatives of the top 12 pharmaceutical companies, as defined by 2009 revenue figures. A series of multiple choice questions was designed to explore group size, accountabilities, roles and responsibilities of group members, outsourcing policy and publication record., Results: A 92% response rate was obtained. Six out of 11 companies have 10 to 30 full time equivalents in safety pharmacology, who hold similar roles and responsibilities; although the majority of members are not qualified at PhD level or equivalent. Accountabilities were similar across companies and all groups have accountability for core battery in vivo studies and problem solving activities but differences do exist for example with in vitro safety screening and pharmacodynamic/pharmokinetic modeling (PK/PD). The majority of companies outsource less than 25% of studies, with in vitro profiling being the most commonly outsourced activity. Finally, safety pharmacology groups are publishing 1 to 4 articles each year., Conclusion: This short survey has highlighted areas of similarity and differences in the way large pharmaceutical companies operate safety pharmacology., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Introduction: The aim of the present study was to evaluate direct/acute effects of arsenic trioxide on action potentials (APs) in isolated cardiac tissues, and to investigate if the choice of species and tissue and the duration of the perfusion play a role in arsenic-induced acute/direct prolongation of AP/QT., Methods and Results: Direct electrophysiological effects of arsenic trioxide were measured in cardiac tissues isolated from four different species using micro-electrode recording. Arsenic (after 30 to 95 min perfusion at 10 μM) significantly prolonged APD(90), increased triangulation of the AP and elicited early afterdepolarizations (EADs) only in isolated guinea-pig and dog Purkinje fibers but not in rabbit and porcine (minipig) Purkinje fibers. Arsenic induced a prolongation of the APD(90) and increases in triangulation and the occurrence of EADs was not observed in papillary muscles of guinea-pigs and rabbits. Arsenic at 4 increasing concentrations from 0.1 μM to 10 μM at the standard perfusion-time of 15 min per concentration, and after a continuous 90-min perfusion at 1 μM and 1 Hz did not induce these direct effects on APD(90), triangulation and EADs in isolated guinea-pig Purkinje fibers, but it at 1 µM elicited EADs in 2 out of 7 preparations after 90 min at 0.2 Hz., Discussion: The present study demonstrates that the choice of species and cardiac tissue as well as perfusion-time play important roles in arsenic-induced direct/acute effects on APD(90) and induction of EADs in vitro., (Copyright © 2012. Published by Elsevier Inc.)

Introduction: The purpose of this study was: (1) to characterize and assess the incidence of spontaneous arrhythmias in totally naive Beagle dogs (n=51; 10 males+41 females): (2) to study the effects of permanent ECG- and LVP-probe telemetry implants both acutely and up to 233days after surgery in a subset of 11 female Beagle dogs., Methods: Naive ECG assessments were conducted by means of 6 external telemetry leads in jacketed dogs. Telemetry ECG recordings were captured by means of implanted telemetry devices suitable for ECG, LVP and aortic blood pressure recording. Experienced laboratory personnel visually evaluated all 22h ECG recordings at different time points after implantation and evaluated the incidence and type of arrhythmia., Results: The 51 healthy and totally naive Beagle dogs showed a prevalence of: 49.0% 2°AVB; 58.8% single atrial premature complexes; 17.6% junctional tachycardia; 27.5% ventricular complexes; 13.7% ventricular escape complexes; 21.6% ventricular premature complexes; 3.9% runs of ventricular complexes; 3.9% runs of ventricular escape complexes. As such, a high percentage of clinically normal Beagle dogs showed different types of arrhythmias when ECG's obtained by external telemetry leads were fully evaluated for a 22h period. The chronic implantation of a ventricular probe through the apex of the heart in 11 dogs only, resulted in higher incidences and frequencies of ventricular episodes, which (in some dogs) extended up to 8weeks. Eight months after surgery none of the implanted dogs showed ventricular tachycardia and only 10% had single ventricular or ventricular premature complexes at low frequencies., Discussion: A thorough evaluation of the ECG's of Beagle dogs selected for telemetry implantation can help to avoid inherent arrhythmia-burdened dogs being implanted and used in studies where these arrhythmias will confound drug assessment by increasing the number of potential false positives., (Copyright © 2011 Elsevier Inc. All rights reserved.)

The problem of drug-induced hERG channel blockade, which can lead to acquired long QT syndrome and potentially fatal arrhythmias, has exercised drug developers and regulatory authorities for over 10 years, and exacting guidelines have been put into place to test for this liability both preclinically (ICH S7B) and clinically (ICH E14). However, the I(Ks) channel, which along with the transient outward current (I(to)) is the other main potassium channel affecting cardiac repolarisation and thus the length of the QT interval, has received little attention, and potent I(Ks) blocking drugs with serious side effects could potentially enter into human testing without being detected by the existing regulatory core battery and standard screening strategies. Here we review the pharmacology of cardiac I(Ks) channel blockade and describe the discovery of a potent I(Ks) blocker whose activity was not detected by standard hERG or invitro action potential screens, but subsequently evoked unprovoked torsades de pointes (TdP) invivo in our anaesthetised dog model. We have exploited this molecule to develop a ligand binding assay to detect I(Ks) blockade at an earlier stage in drug discovery, and note that several other laboratories developing new drugs have also developed higher throughput screens to detect I(Ks) blockade (e.g., [Trepakova, E. S., Malik, M. G., Imredy, J. P., Penniman, J. R., Dech, S. J., & Salata, J. J. (2007) Application of PatchXpress planar patch clamp technology to the screening of new drug candidates for cardiac KCNQ1/KCNE1 (I(Ks)) activity. Assay Drug Development Technology 5, 617-627]). Because of the presence of I(Ks) channels in other tissues, including blood vessels and in the epithelia of intestine, kidney, lung and the cochlea, I(Ks) blockade has the potential to cause extensive side effects in addition to QT prolongation and arrhythmias. We therefore suggest that compounds selected for development should also be examined for I(Ks) liability before testing in humans. The possibility of undetected I(Ks) blockade is therefore an additional gap to that identified earlier [Lu, H. R., Vlaminckx, E., Hermans, A. N., Rohrbacher, J., Van Ammel, K., Towart, R., et al. (2008) Predicting drug-induced changes in QT interval and arrhythmias: QT-shortening drugs point to gaps in the ICH S7B Guidelines. British Journal of Pharmacology, 154, 1427-1438] in the ICH S7B regulatory guidelines.

The purpose of conducting cardiovascular safety pharmacology studies is to investigate the pharmacological profiles of new molecular entities (NMEs) and provide data that can be used for optimization of a possible new drug, and help make a selection of NMEs for clinical development. An anaesthetised dog preparation has been used for more than two decades by our department to measure multiple cardiovascular and respiratory parameters and to evaluate different scientific models, leading to more in-depth evaluation of drug-induced cardiovascular effects. An anaesthetic regime developed in house (induction with lofentanil, scopolamine and succinylcholine, and maintenance with fentanyl and etomidate) gives us a preparation free of pain and stress, with minimal effects on the cardiovascular system. This anaesthetic regime had minimal influences on circulating catecholamine levels, on the baroreflex sensitivity, and on all measured basal parameters compared to conscious dogs. All parameters were stable for at least 3 h, with acceptable tolerance intervals, evaluated over 99 safety studies with 3 vehicle treatments (saline, 10% and 20% hydroxypropyl-beta-cyclodextrin). This translates into a highly sensitive model for detecting possible drug-induced effects of NMEs with different mechanisms of action such as: Ca-, Na-, I(Kr)-, I(Ks)-channel blockers, K- and Ca-channel activators, alpha1- and beta-agonists, and muscarinic antagonists. Fentanyl in combination with etomidate is a successful anaesthetic regime in humans [Stockham, R.J., Stanley, T.H., Pace, N.L., King, K., Groen, F. & Gillmor, S.T. (1987). Induction of anaesthesia with fentanyl or fentanyl plus etomidate in high-risk patients. Journal of Cardiothoracic Anesthesia. 1(1), 19-23.]. In the anaesthetised dog, QT correction factors (Van de Water correction and body temperature correction) and risk factors (total, short-term and long-term instability) have been evaluated, using this regime [Van de Water, A., Verheyen, J., Xhonneux, R. & Reneman, R. (1989). An improved method to correct the QT interval of the electrocardiogram for changes in heart rate. Journal of Pharmacological Methods, 22, 207-217.; van der Linde, H.J., Van Deuren, B., Teisman, A., Towart, R. & Gallacher, D.J. (2008). The effect of changes in core body temperature on the QT interval in beagle dogs: A previously ignored phenomenon, with a method for correction. British Journal of Pharmacology, 154, 1474-1481.; van der Linde, H.J., Van de Water, A., Loots, W., Van Deuren, B., Lu, H.R., Van Ammel, K., et al. (2005) A new method to calculate the beat-to-beat instability of QT duration in drug-induced long QT in anaesthetised dogs. Journal of Pharmacological and Toxicological Methods, 52, 168-177.]. Furthermore, this anaesthetic protocol has been used to create different scientific models (long QT, short QT) with different specific end-points (ventricular fibrillation, adrenergic- or pause-dependent TdP) and also their specific precursors: e.g. aftercontractions, phase 2 EADs, phase 3 EADs, DADs, T-wave morphology changes, T-wave alternans, R-on-T, transmural and interventricular dispersion [Gallacher, D.J., Van de Water, A., van der Linde, H.J., Hermans, A.N., Lu, H.R., Towart, R., et al. (2007). In vivo mechanisms precipitating torsade de pointes in canine model of drug-induced long QT1 syndrome. Cardiovascular Research, 76-2, 247-256.]. This paper gives a brief overview of the stability, reproducibility, sensitivity and utility of a well-validated anaesthetised dog model.

Assessing drug-induced changes (particularly prolongation) in the QT interval has been the major preoccupation of safety pharmacology since its inception, under the assumption that QT widening represents a surrogate biomarker for torsades de pointes (TdeP) liability. While evidence of changes in QT remains a bane to the development of novel therapeutic agents, non-clinical and clinical methods have been developed (with a certain amount of validation) to limit this potential liability of a new chemical entity (NCE). Because of the associated withdrawal of numerous drugs from clinical use, determining whether or not a drug development candidate exhibits a TdeP liability has been the motivation in the implementation of discussions between 'pharmaceutical companies', academicians, clinicians and regulatory authorities worldwide that has led to the development of the ICHS7A and ICHS7B guidance documents (Anon, 2001, 2005). Simultaneously, it has resulted in the firm establishment of safety pharmacology as a standalone discipline within the drug development scheme (Pugsley et al., 2008). As far as TdeP liability is concerned, QT widening remains the most poignant issue, in that QT widening in humans is immediately regarded as a cause for concern, yet QT widening in preclinical models (and indeed in man) is not a quantitative predictor of TdeP liability (and indeed may not even be a qualitative predictor by itself (Pugsley et al., 2008). The present focused issue of the journal returns to safety pharmacology, and contains papers arising from the 8th annual SPS Meeting that was held in Madison, WI in 2008. Indeed, so many papers have arisen from the meeting that this issue of the Journal is only part 1. Part 2 will be published as the next issue of the Journal. Some topics which have been addressed include whether an assessment method for drugs that produce a shortened QT interval is needed, what the role of the slow component of the delayed rectifier K current (I(Ks)) should be in a safety assessment and whether safety pharmacology endpoints can or should be added to repeat dose Toxicology studies.

Introduction: Recently we have demonstrated that to the choice of tissue type is important in identifying I(Kr) and I(Ks)-induced prolongation of the action potential. However, the differential sensitivity of cardiac tissues to other ionic current blockers or modulators is relatively unknown. The aim of the present study was therefore to evaluate tissue-specific effects of different ion channel blockers or activators on the action potential (AP), which can affect other parameters in addition to drug-induced APD/QT prolongation or shortening., Methods and Results: Electrophysiological effects were measured in isolated rabbit Purkinje fibers, papillary muscles and ventricular trabeculae using a microelectrode technique under the following conditions: block of I(to) with 4-AP (1 x 10(-3) M), block of Ca(2+) channels with diltiazem (1 x 10(-5) M), block of Na(+) channels with flecainide (1 x 10(-5) M), activation of Ca(2+) current with Bay-K-8644 (1 x 10(-5) M), activation of K(ATP) channels with levcromakalim (1 x 10(-5) M) or block of I(K1) current with BaCl(2) (n=8 to 12 for each group). 4-AP prolonged APD significantly more in the Purkinje fiber than in the papillary muscle or the ventricular trabecula. 4-AP elicited 63% incidence of early afterdepolarizations but 0% in the papillary or trabeculae. Diltiazem and flecainide shortened APD(40) and APD(50) and increased triangulation more in the Purkinje fiber, whilst having little effect on these parameters in the papillary muscle or the ventricular trabecula. Bay-K-8644 significantly prolonged APD in the ventricular trabecula, but not in the Purkinje fiber or the papillary muscle. BaCl(2) prolonged APD(90) in all tissues, but significantly shortened APD(40) only in the Purkinje fiber. Levcromakalim shortened APD in all tissues, but significantly less in the Purkinje fibers., Conclusion: The present study demonstrates that certain cardiac tissues respond differently to the same ion channel blockers/activators, which are not involved in APD/QT prolongation. As such the appropriate selection of tissue needs to be taken into careful consideration in cardiac safety assessments when exploring different mechanisms of drug-induced changes in the action potential.

Introduction: Regulatory guidelines (CPMP/986/96, ICHS7B) recommend the use of isolated cardiac tissues, including Purkinje fibers, papillary muscles and ventricular trabeculae, for detecting potential drug-induced long QT. However, the differential sensitivity of these tissues in experimental drug-induced long QT is relatively unknown. We investigated the electrophysiological characteristics of these tissue types in vitro together with their different responses to drugs that are known to induce prolongation of the QT interval in man., Methods: Electrophysiological parameters were measured in vitro using a micro-electrode technique. The isolated rabbit Purkinje fibers, papillary muscles or ventricular trabeculae were superperfused with Tyrode's solution and stimulated according to different stimulation protocols. The effects of dofetilide (1 x 10(-8) M), sertindole (1 x 10(-6) M), erythromycin (3 x 10(-4) M) and sparfloxacin (1 x 10(-4) M) were evaluated relative to solvent (n=8 to 12 for each group)., Results: In isolated Purkinje fibers, action potential duration at 90% repolarization (APD(90) at 1 Hz) was markedly prolonged by 55% (erythromycin), 103% (dofetilide), 118% (sertindole) and 88% (sparfloxacin) from baseline. The prolongation of APD(90) caused by these 4 compounds was associated with a 28% to 78% incidence of early afterdepolarizations (EADs) at 0.2 Hz only in the Purkinje fiber. In contrast, APD(90) was altered by erythromycin, dofetilide, sertindole and sparfloxacin only by +15%, +6%, -7% or +15%, respectively, in isolated papillary muscles, and by 33%, +28%, +4% or +16%, respectively, in ventricular trabeculae. EADs were not induced by these four compounds in papillary muscles or in trabeculae. Reducing the stimulation rate to 0.2 Hz resulted in a 33% prolongation of APD(90) in Purkinje fibers, while APD(90) was shortened by 10% in papillary muscles and by 20% in ventricular trabeculae., Conclusion: The present study demonstrates that the differential sensitivity of tissue types play an important role in the detection of drug-induced long APD and EADs. Indeed the Purkinje fiber was the only tissue type to display the well known phenomenon associated with I(kr) channel blockade (inverse-use dependence), when the stimulation rate was decreased below 1 Hz. Rabbit Purkinje fibers constitute the most sensitive tissue type for detecting drug-induced long action potential duration and EADs. As such the selection of tissue type needs to be taken into careful consideration in cardiac safety assessments when exploring drug induced long QT.