The rat telemetry assay and venous catheter access buttons for use in cardiovascular safety pharmacology assessments - Surgical methods, refinements and colony maintenance.

INTRODUCTION: Rat telemetry is the assay of choice to assess the potential effects of novel drug candidates on cardiovascular parameters during early drug discovery. Telemetry device implantation can be combined with venous catheter and access button implantation when intravenous administration of the drug substance is required. METHODS: Rats (Sprague Dawley or Han Wistar) were implanted with telemetry devices for arterial blood pressure measurement using either direct aortic catheterisation (n = 131) or aortic catheterisation via the femoral artery (n = 17). Bipolar leads for ECG recording were also implanted in some of the animals (n = 102). Femoral vein catheters and access buttons were implanted as a separate surgery after the initial telemetry implantation (n = 43). RESULTS: 128 animals (86%) were implanted successfully with telemetry devices without any notable surgical or post-surgical problems. When considering the 2 different catheterisation methods separately, the success rate of the direct aortic approach was 88% compared to 76% with the aortic placement via the femoral artery. Lameness was the most common post-surgical problem. Blood loss during surgery and ischaemic patches on the tail were also observed at a low incidence with the direct aortic approach. Catheter pull-out occurred in some rats before the first signal check reducing the overall success rate for blood pressure measurement using the direct aortic approach to 85%. A 95% success rate was observed for catheter and access button implantation. DISCUSSION: A high success rate is possible when implanting telemetry devices in rats with and without venous catheters and access buttons. We have attempted to provide solutions to problems and describe refinements to the procedure which may further improve surgical outcomes.

Differentiating multichannel block on the guinea pig ECG: Use of Tpeak-Tend and J-Tpeak.

INTRODUCTION: The anaesthetised guinea pig is a well characterised assay for early assessment of drug effects on ventricular repolarisation and risk of Torsade de Pointes (TdP). We assessed whether a selective hERG blocker with known TdP risk could be differentiated from lower risk, balanced ion channel blockers in the guinea pig, using corrected QT (QTc) interval alongside novel electrocardiogram (ECG) biomarkers J-Tpeakc and Tpeak-Tend. Effects were compared with previous clinical investigations at similar plasma concentrations and with another index of TdP risk, the electromechanical window (EMW). METHODS: Twenty-two Dunkin Hartley guinea pigs anaesthetised with sodium pentobarbitone were instrumented for haemodynamic measurement and ECG recording. Three ascending doses of vehicle (n = 6), dofetilide (2, 6 or 20 µg/kg; n = 7), ranolazine (2, 6 or 20 mg/kg; n = 5) or verapamil (0.1, 0.3 or 1.0 mg/kg; n = 4) were administered intravenously. RESULTS: As reported in previous clinical studies, dofetilide induced dose-dependent increases in QTc interval, with increases in both J-TpeakC or Tpeak-Tend, while verapamil caused no significant increase in QTc interval, J-TpeakC or Tpeak-Tend. Ranolazine caused dose-dependent increases in QTc interval and corrected J-Tpeakc, but had no effect on Tpeak-Tend, which is in contrast to the effects reported in humans at similar concentrations. Only dofetilide caused a clear, dose-related decrease in the EMW. DISCUSSION: These findings suggest that measurements of J-Tpeakc and Tpeak-Tend in addition to QT interval, may help differentiate pure hERG channel blockers with high risk of TdP from lower risk, multichannel blockers.

INSPIRE: A European training network to foster research and training in cardiovascular safety pharmacology.

Safety pharmacology is an essential part of drug development aiming to identify, evaluate and investigate undesirable pharmacodynamic properties of a drug primarily prior to clinical trials. In particular, cardiovascular adverse drug reactions (ADR) have halted many drug development programs. Safety pharmacology has successfully implemented a screening strategy to detect cardiovascular liabilities, but there is room for further refinement. In this setting, we present the INSPIRE project, a European Training Network in safety pharmacology for Early Stage Researchers (ESRs), funded by the European Commission's H2020-MSCA-ITN programme. INSPIRE has recruited 15 ESR fellows that will conduct an individual PhD-research project for a period of 36 months. INSPIRE aims to be complementary to ongoing research initiatives. With this as a goal, an inventory of collaborative research initiatives in safety pharmacology was created and the ESR projects have been designed to be complementary to this roadmap. Overall, INSPIRE aims to improve cardiovascular safety evaluation, either by investigating technological innovations or by adding mechanistic insight in emerging safety concerns, as observed in the field of cardio-oncology. Finally, in addition to its hands-on research pillar, INSPIRE will organize a number of summer schools and workshops that will be open to the wider community as well. In summary, INSPIRE aims to foster both research and training in safety pharmacology and hopes to inspire the future generation of safety scientists.

Model for End-Stage Liver Disease Score Predicts Development of First Episode of Spontaneous Bacterial Peritonitis in Patients With Cirrhosis.

OBJECTIVE: To examine whether baseline model for end-stage liver disease (MELD) score in patients with cirrhosis and ascites predicts the future development of first spontaneous bacterial peritonitis (SBP) episode. METHODS: A retrospective case-control study was performed at three academic centers to select patients admitted with first SBP episode (cases) and those with ascites admitted for decompensation without SBP (controls). Medical records from these centers were reviewed between January 1, 2008, and December 31, 2013. Cases and controls were matched (1:2) for age, sex, and race. Conditional logistic recession models were built to determine whether baseline MELD score (within a month before hospitalization) predicts first SBP episode. RESULTS: Of 697 patients (308, 230, and 159 from centers A, B, and C, respectively), cases and controls were matched in 94%, 89%, and 100% at three respective centers. In the pooled sample, probability of SBP was 11%, 31%, 71%, and 93% at baseline MELD scores less than or equal to 10, from 11 to 20, from 21 to 30, and greater than 30, respectively. Compared with MELD score less than or equal to 10, patients with MELD scores from 11 to 20, 21 to 30, and greater than 30 had six- (3- to 11-), 29- (12- to 69-), and 115- (22- to 598-) folds (95% CI) risk of SBP, respectively. Based on different MELD score cutoff points, MELD score greater than 17 was most accurate in predicting SBP occurrence. Analyzing 315 patients (152 cases) with available data on ascitic fluid protein level controlling for age, sex, and center, MELD score but not ascitic fluid protein associated with first SBP episode with respective odds ratios of 1.20 (1.14 to 1.26) and 0.88 (0.70 to 1.11). CONCLUSION: Baseline MELD score predicts first SBP episode in patients with cirrhosis and ascites.

Social-housing and use of double-decker cages in rat telemetry studies.

Rat telemetry is widely used for biomedical research purposes and is used routinely in early pre-clinical drug development to screen for the potential cardiovascular risk of candidate drugs. Historically, these studies have been conducted in individually housed conditions which can impact significantly on an animal's welfare. Here we present data from a survey of pharmaceutical companies and contract research organisations to define current industry practices relating to the housing of rats during telemetry studies and to expand and complement a similar project in non-rodents. Results of the survey showed that 75% of respondents socially house rats on non-recording days of telemetry studies, whereas on recording days only 46% of respondents socially house the animals. When social housing is used on rat telemetry studies, rats are usually housed with an unrecorded companion animal. We also present and compare data from a telemetry study in standard individually ventilated cages (IVCs) with a study using new double-decker IVCs, both conducted using a companion animal approach. Telemetry signals were successfully collected from the double-decker IVCs without a loss of signal quality whilst offering a more spacious environment that allowed the animals to exhibit natural behaviours including full upright posture. Cardiovascular responses following pharmacological intervention with verapamil were similar when assessed in the standard and double-decker cages. Power analysis was conducted on pooled data from the studies in socially housed animals with preliminary results showing the power of detection of drug-induced effects is equivalent to previously published data in individually housed rats. This illustrates that telemetry recordings can be made from rats in socially housed conditions within standard or larger double-decker cages for the for the collection of cardiovascular telemetry data.

Quantifying the relationship between inhibition of VEGF receptor 2, drug-induced blood pressure elevation and hypertension.

BACKGROUND AND PURPOSE: Several anti-angiogenic cancer drugs that inhibit VEGF receptor (VEGFR) signalling for efficacy are associated with a 15-60% incidence of hypertension. Tyrosine kinase inhibitors (TKIs) that have off-target activity at VEGFR-2 may also cause blood pressure elevation as an undesirable side effect. Therefore, the ability to translate VEGFR-2 off-target potency into blood pressure elevation would be useful in development of novel TKIs. Here, we have sought to quantify the relationship between VEGFR-2 inhibition and blood pressure elevation for a range of kinase inhibitors. EXPERIMENTAL APPROACH: Porcine aortic endothelial cells overexpressing VEGFR-2 (PAE) were used to determine IC50 for VEGFR-2 phosphorylation. These IC50 values were compared with published reports of exposure attained during clinical use and the corresponding incidence of all-grade hypertension. Unbound average plasma concentration (Cav,u ) was selected to be the most appropriate pharmacokinetic parameter. The pharmacokinetic-pharmacodynamic (PKPD) relationship for blood pressure elevation was investigated for selected kinase inhibitors, using data derived either from clinical papers or from rat telemetry experiments. KEY RESULTS: All-grade hypertension was predominantly observed when the Cav,u was >0.1-fold of the VEGFR-2 (PAE) IC50 . Furthermore, based on the PKPD analysis, an exposure-dependent blood pressure elevation >1 mmHg was observed only when the Cav,u was >0.1-fold of the VEGFR-2 (PAE) IC50 . CONCLUSIONS AND IMPLICATIONS: Taken together, these data show that the risk of blood pressure elevation is proportional to the amount of VEGFR-2 inhibition, and a margin of >10-fold between VEGFR-2 IC50 and Cav,u appears to confer a minimal risk of hypertension.

A multi-site comparison of in vivo safety pharmacology studies conducted to support ICH S7A & B regulatory submissions.

INTRODUCTION: Parts A and B of the ICH S7 guidelines on safety pharmacology describe the in vivo studies that must be conducted prior to first time in man administration of any new pharmaceutical. ICH S7A requires a consideration of the sensitivity and reproducibility of the test systems used. This could encompass maintaining a dataset of historical pre-dose values, power analyses, as well as a demonstration of acceptable model sensitivity and robust pharmacological validation. During the process of outsourcing safety pharmacology studies to Charles River Laboratories, AstraZeneca set out to ensure that models were performed identically in each facility and saw this as an opportunity to review the inter-laboratory variability of these essential models. METHODS: The five in vivo studies outsourced were the conscious dog telemetry model for cardiovascular assessment, the rat whole body plethysmography model for respiratory assessment, the rat modified Irwin screen for central nervous system assessment, the rat charcoal meal study for gastrointestinal assessment and the rat metabolic cage study for assessment of renal function. Each study was validated with known reference compounds and data were compared across facilities. Statistical power was also calculated for each model. RESULTS: The results obtained indicated that each of the studies could be performed with comparable statistical power and could achieve a similar outcome, independent of facility. DISCUSSION: The consistency of results obtained from these models across multiple facilities was high thus providing confidence that the models can be run in different facilities and maintain compliance with ICH S7A and B.