Introduction: Currently, standardmethods formeasuring cardiac output are either invasive (i.e. flowprobe) or are limited in terms of short measurement intervals and measurement variability (i.e. echocardiography). The ability to reliably measure cardiac output in a non-invasive manner in large animals would provide a valuable tool to expand functional cardiovascular endpoints in preclinical safety studies. PhysioFlow (R) is a novel method that uses waveform analysis of an impedance signal to measure cardiac output non-invasively. Unlike cardiac impedance techniques in the past, PhysioFlow (R) is not dependant on thoracic structure or basal thoracic impedance (Z(0)) and therefore this methodology is transferrable from human to animal models.Methods: Three tool compounds with known effects on cardiac output were administered to conscious beagle dogs to determine if the non-invasive PhysioFlow (R) system could detect the expected changes in stroke volume and cardiac output as determined by literature references using the current standard methodologies (e.g. aortic blood flow and thermodilution).Results: The PhysioFlow (R) system was able to detect increases in cardiac output when dosed with 20 mu g/kg of Dobutamine, a decrease in cardiac output when dosed with 0.1 mg/kg of Acepromazine, and no significant change in cardiac output when dosed with 2 mg/kg of Minoxidil. These results are within expected ranges based on published literature (Stepien et al., 1995; Taylor et al., 2007).Discussion: PhysioFlow (R), a signal morphology-based impedance cardiography, can be utilized to reliably and non-invasively measure cardiac output in beagle dogs. (C) 2016 Elsevier Inc. All rights reserved.

Purpose: Aggrecanase activity, most notably ADAMTS-5, has been widely implicated as a pathogenic factor in cartilage degradation. Therefore, highly selective and potent monoclonal antibodies (mAbs) to both ADAMTS-5 and ADAMTS-4 were developed and experimental systems were utilized to assess target engagement and modulation of disease-related and safety endpoints with the intent of selecting a candidate and supporting clinical development in osteoarthritis (OA). Methods/Results: In a surgical mouse model of OA, both ADAMTS-5 and ADAMTS-4-specific mAbs bound within cartilage following systemic administration, demonstrating access to the anticipated site of action, whereas structural disease modification and associated alleviation of pain-related behavior were only observed with ADAMTS-5 mAb treatment. Likewise, ex vivo treatment of human OA cartilage demonstrated a preferential role for ADAMTS-5 inhibition over ADAMTS-4, as measured by aggrecan-derived 374ARGS neoepitope release in explant cultures and was most evident in a distinct subset of patients where elevated 374ARGS neoepitope levels were present in explants prior to treatment. Notably, suppression of 374ARGS neoepitope release was sustained for weeks after a single treatment of human cartilage explants and in cynomolgus monkeys, consistent with a high affinity antibody/antigen interaction and slow ADAMTS-5 turnover that could translate to less frequent clinical dosing. As a result an ADAMTS-5 selective monoclonal antibody (GSK2394002) was progressed for preclinical development as a potential OA disease modifying therapeutic. While desired pharmacology was observed in explant cartilage and disease models, ADAMTS-5 is also expressed in cardiovascular (CV) tissue. Recently it was reported that knockout mice show developmental heart valve defects and altered vascular and fibroblast proteoglycan processing. Along with these observations, an initial histologic cardiac signal in cynomolgus monkeys (subendocardial hemorrhage) prompted a careful assessment of CV function in a set of dedicated safety pharmacology studies. In these studies, increased mean arterial pressure (MAP) was observed within days after >3 mg/kg dosing and ST segment elevation was noted on 24 hour continuous ECG monitoring >30 mg/kg. Surprisingly, these effects were sustained for up to 8 months following a single dose and MAP was increased upon repeated low dose administration suggesting the effects are cumulative and, unlike the systemic pharmacodynamic marker signal, once induced they may not be readily reversible. Although a potential mechanistic link between ADAMTS-5 activity and versican processing is demonstrated, the cause of these cardiovascular observations is currently unknown. In a parallel general toxicology study, there were no histologic or biochemical correlates to the CV findings and no evidence of off-target mechanisms could be identified using in vitro tests examining cardiac conductance, ion channels, or binding selectivity in protein arrays. Conclusions: The findings support a hypothesis set forth from knockout mouse studies that ADAMTS-5 is the major aggrecanase involved in cartilage degradation and confirms a link between a native biological pathway and pharmacology which translates to human tissues and non-human primate models, subsequently pointing to a specific OA patient population to target. However, our results suggest selective and potent ADAMTS-5 engagement/inhibition with GSK2394002, while demonstrating desired effects on cartilage-related endpoints, results in modulation of CV functions that pose considerable challenges for clinical development.

Introduction Assessing the effects of new chemical entities on cardiovascular function is an essential component of nonclinical safety evaluation. Regulatory guidelines advocate the use of telemetry to evaluate drug effects on the cardiovascular system of animal including electrocardiogram (ECG) waveforms. Currently, standard method is to place ECG electrodes subcutaneously and the ECG signals have a suboptimal signal mainly due to movement artifacts. In order to improve and optimize the ECG signal and minimize movement artifacts, a model for continuous monitoring of the ECG in conscious, unrestrained animals by placing ECG electrodes on the pericardium was evaluated.