Mechanism of doxorubicin cardiotoxicity evaluated by integrating multiple molecular effects into a biophysical model.

BACKGROUND AND PURPOSE: Doxorubicin (DOX) is an effective cancer therapeutic agent but causes therapy-limiting cardiotoxicity. The effects of DOX and its metabolite doxorubicinol (DOXL) on individual channels have been well characterized in isolation. However, it is unknown how the action and interaction of affected channels combine to generate the phenotypic cardiotoxic outcome. We sought to develop an in silico model that links drug effects on channels to action potential duration (APD) and intracellular Ca2+ concentration in order to address this gap in knowledge. EXPERIMENTAL APPROACH: We first propose two methods to obtain, from published values, consensus drug effects on the currents of individual channels, transporters and pumps. Separately, we obtained equivalent values for APD and Ca2+ concentration (the readouts used as surrogates for cardiotoxicity). Once derived, the consensus effects on the currents were incorporated into established biophysical models of the cardiac myocyte and were refined adjusting the sarcoplasmic reticulum Ca2+ leak current (ILeak ) until the consensus effects on APD and Ca2+ dynamics were replicated. Using factorial analysis, we then quantified the relative contribution of each channel to DOX and DOXL cardiotoxicity. KEY RESULTS: The factorial analysis identified the rapid delayed rectifying K+ current, the L-type Ca2+ current and the sarcoplasmic reticulum ILeak as the targets primarily responsible for the cardiotoxic effects on APD and Ca2+ dynamics. CONCLUSIONS AND IMPLICATIONS: This study provides insight into the mechanisms of DOX-induced cardiotoxicity and a framework for the development of future diagnostic and therapeutic strategies.

Experimental recovery of a qubit from partial collapse.

We describe and implement a method to restore the state of a single qubit, in principle perfectly, after it has partially collapsed. The method resembles the classical Hahn spin echo but works on a wider class of relaxation processes, in which the quantum state partially leaves the computational Hilbert space. It is not guaranteed to work every time, but successful outcomes are heralded. We demonstrate, using a single trapped ion, a better performance from this recovery method than can be obtained employing projection and postselection alone. The demonstration features a novel qubit implementation that permits both partial collapse and coherent manipulations with high fidelity.

Methodological innovations expand the safety pharmacology horizon.

Almost uniquely in pharmacology, drug safety assessment is driven by the need for elaboration and validation of methods for detecting drug actions. This is the 9th consecutive year that the Journal of Pharmacological and Toxicological Methods (JPTM) has published themed issues arising from the annual meeting of the Safety Pharmacology Society (SPS). The SPS is now past its 10th year as a distinct (from pharmacology to toxicology) discipline that integrates safety pharmacologists from industry with those in academia and the various global regulatory authorities. The themes of the 2011 meeting were (i) the bridging of safety assessment of a new chemical entity (NCE) between all the parties involved, (ii) applied technologies and (iii) translation. This issue of JPTM reflects these themes. The content is informed by the regulatory guidance documents (S7A and S7B) that apply prior to first in human (FIH) studies, which emphasize the importance of seeking model validation. The manuscripts encompass a broad spectrum of safety pharmacology topics including application of state-of-the-art techniques for study conduct and data processing and evaluation. This includes some exciting novel integrated core battery study designs, refinements in hemodynamic assessment, arrhythmia analysis algorithms, and additionally an overview of safety immunopharmacology, and a brief survey discussing similarities and differences in business models that pharmaceutical companies employ in safety pharmacology, together with SPS recommendations on 'best practice' for the conduct of a non-clinical cardiovascular assessment of a NCE.

Innovation in safety pharmacology testing.

This issue of the Journal of Pharmacological and Toxicological Methods (JPTM) is themed. It is the eighth in a series, arising from the Annual Safety Pharmacology Society (SPS) meeting. The SPS is now in its 10th year as an independent branch of biological sciences (distinct from pharmacology and toxicology) and is the primary forum for driving advances in safety pharmacology. The theme of the meeting and this journal issue is innovation, and the focus is non-clinical safety assessment of new chemical entity (NCEs). The content is informed by regulatory guidance documents (S7A and S7B) prior to first in human (FIH) studies. The manuscripts cover a broad spectrum of safety pharmacology topics from theory to practice, with interrogation of state-of-the-art techniques, and profiling of methods that are in development for safety assessment. Philosophical and strategic issues are addressed, with consideration of the use of novel methods for population pharmacokinetic (PK) analysis, abuse liability, electrocardiogram (ECG) analysis algorithms, in vitro cardiac slice preparations, human pluripotent stem cells, and a brief discussion regarding the assessment of changes in the QRS complex of the ECG indicative of drug-induced blockade of cardiac sodium channels. Safety pharmacology methods continue to evolve.

A shared DNA-damage-response pathway for induction of stem-cell death by UVB and by gamma irradiation.

Both UVB radiation and DNA-breaking agents were previously reported to kill Arabidopsis stem cells. We demonstrate that death induced by UVB or by ionizing radiation (IR) requires Suppressor of Gamma Response 1 (SOG1), a transcription factor already found to govern many responses to these agents in Arabidopsis. DNA-damage responses (DDRs) triggered primarily by replication-blocking photoadducts or double-strand-breaks thus converge to a shared programmed-cell-death (PCD) pathway. Both UVB- and IR-induced PCD also require functional DDR protein kinases. Employment of atr atm mutants (uniquely available in Arabidopsis) shows that either ATR (which recognizes ssDNA) or ATM (which recognizes DSBs) suffices for PCD induction by either agent. Thus, DNA damage made by UVB or by IR engenders both ATM-activating and ATR-activating structures. The elevated PCD in UVB-irradiated atr and atm mutants suggests that in wt plants ATR and/or ATM may activate both pathways that avert PCD and those that elicit it. The similar PCD levels induced by roughly 30,000 unrepaired photoadducts vs. 20 IR-induced DSBs indicate that DDR damage-tolerance activities in this model stem-cell niche are remarkably efficient.

Non-clinical models: validation, study design and statistical consideration in safety pharmacology.

The current issue of the Journal of Pharmacological and Toxicological Methods (JPTM) focuses exclusively on safety pharmacology methods. This is the 7th year the Journal has published on this topic. Methods and models that specifically relate to methods relating to the assessment of the safety profile of a new chemical entity (NCE) prior to first in human (FIH) studies are described. Since the Journal started publishing on this topic there has been a major effort by safety pharmacologists, toxicologists and regulatory scientists within Industry (both large and small Pharma as well as Biotechnology companies) and also from Contract Research Organizations (CRO) to publish the surgical details of the non-clinical methods utilized but also provide important details related to standard and non-standard (or integrated) study models and designs. These details from core battery and secondary (or ancillary) drug safety assessment methods used in drug development programs have been the focus of these special issues and have been an attempt to provide validation of methods. Similarly, the safety pharmacology issues of the Journal provide the most relevant forum for scientists to present novel and modified methods with direct applicability to determination of drug safety-directly to the safety pharmacology scientific community. The content of the manuscripts in this issue includes the introduction of additional important surgical methods, novel data capture and data analysis methods, improved study design and effects of positive control compounds with known activity in the model.

Is it worthwhile to offer repeat hydrodilatation for frozen shoulder after 6 weeks?

AIMS: Frozen shoulder is a common problem for which hydrodilatation is an established treatment option although the optimal regime is not yet established. We asked 'is it worthwhile to offer a repeat hydrodilatation procedure after 6 weeks?' METHODS: A total of 36 patients diagnosed with frozen shoulder and undergoing hydrodilatation were evaluated. All patients had an initial hydrodilatation and were offered a repeat procedure after 6 weeks. Of the total patients, 22 chose to undergo a repeat procedure. The Shoulder Disability Questionnaire UK Score and Oxford Shoulder Score were used to evaluate pre- and postprocedure outcome. The mean follow up was 15.4 months. RESULTS: The Oxford Shoulder Score improved by a significant level after both single (30.5-17.1) and repeat hydrodilatation (36.0-21.4). No significant change was observed after either procedure when using the Shoulder Disability Questionnaire UK Score as an outcome measure. CONCLUSIONS: The data are suggestive that routine hydrodilatation after 6 weeks is not appropriate and should be instead offered to those patients who can tolerate it and who have not fully benefitted after a single procedure.

Principles of safety pharmacology.

Safety Pharmacology is a rapidly developing discipline that uses the basic principles of pharmacology in a regulatory-driven process to generate data to inform risk/benefit assessment. The aim of Safety Pharmacology is to characterize the pharmacodynamic/pharmacokinetic (PK/PD) relationship of a drug's adverse effects using continuously evolving methodology. Unlike toxicology, Safety Pharmacology includes within its remit a regulatory requirement to predict the risk of rare lethal events. This gives Safety Pharmacology its unique character. The key issues for Safety Pharmacology are detection of an adverse effect liability, projection of the data into safety margin calculation and finally clinical safety monitoring. This article sets out to explain the drivers for Safety Pharmacology so that the wider pharmacology community is better placed to understand the discipline. It concludes with a summary of principles that may help inform future resolution of unmet needs (especially establishing model validation for accurate risk assessment). Subsequent articles in this issue of the journal address specific aspects of Safety Pharmacology to explore the issues of model choice, the burden of proof and to highlight areas of intensive activity (such as testing for drug-induced rare event liability, and the challenge of testing the safety of so-called biologics (antibodies, gene therapy and so on.).

High-fidelity readout of trapped-ion qubits.

We demonstrate single-shot qubit readout with a fidelity sufficient for fault-tolerant quantum computation. For an optical qubit stored in 40Ca+ we achieve 99.991(1)% average readout fidelity in 10(6) trials, using time-resolved photon counting. An adaptive measurement technique allows 99.99% fidelity to be reached in 145 micros average detection time. For 43Ca+, we propose and implement an optical pumping scheme to transfer a long-lived hyperfine qubit to the optical qubit, capable of a theoretical fidelity of 99.95% in 10 micros. We achieve 99.87(4)% transfer fidelity and 99.77(3)% net readout fidelity.

Nitric oxide fails to confer endogenous antiarrhythmic cardioprotection in the primate heart in vitro.

BACKGROUND AND PURPOSE: The role of nitric oxide (NO) in cardiac pathophysiology remains controversial. According to data from several studies using rat and rabbit isolated hearts, NO is an endogenous cardioprotectant against reperfusion-induced ventricular fibrillation (VF). Thus, if cardiac NO production is abolished by perfusion with L-N(G)-nitro-L-arginine methylester (L-NAME) (100 microM) there is a concomittant increase in the incidence of reperfusion-induced VF, with L-NAME's effects on NO and VF prevented by L- (but not D-) arginine co-perfusion. To make a better estimate of the clinical relevance of these findings, 100 microM L-NAME was tested in primate hearts under similar conditions. EXPERIMENTAL APPROACH: Marmoset (Callithrix jaccus) hearts, isolated and perfused, were subjected to 60 min left regional ischaemia followed by 10 min reperfusion in vitro. The ECG was recorded and NO in coronary effluent measured by chemiluminescence. KEY RESULTS: L-NAME (100 micro M) decreased NO in coronary effluent throughout ischaemia and reperfusion (e.g. from 3720+/-777 pmol min(-1) g(-1) in controls to 699+/-98 pmol min(-1) g(-1) after 5 min of ischaemia) and, during ischaemia, lowered coronary flow and reduced heart rate, actions identical to those seen in rat and rabbit hearts. However, the incidence of reperfusion-induced VF was unchanged (20%, with or without L-NAME). CONCLUSIONS AND IMPLICATIONS: A species difference exists in the effectiveness of endogenous NO to protect hearts against reperfusion-induced VF. The present primate data, which presumably take precedence over rat and rabbit data, cast doubt on the clinical relevance of NO as an endogenous, antiarrhythmic, cardioprotectant.

Nupafant, a PAF-antagonist prototype for suppression of ventricular fibrillation without liability for QT prolongation?

BACKGROUND AND PURPOSE: PAF antagonists inhibit ischaemia-induced ventricular fibrillation (VF) in animals. However, unfavourable ancillary actions (on QT interval and coronary flow) have been reported with the PAF antagonist, BN-50739. If these are class actions, they would preclude development of PAF antagonists as novel anti-VF drugs. Our purpose was to examine this proposition using the hitherto untested PAF antagonist, nupafant. EXPERIMENTAL APPROACH: Two rat heart preparations (Langendorff and 'dual coronary' perfusion) were used to assay nupafant's effects on ischaemia-induced VF, coronary flow and QT interval, and to test for the site-selectivity necessary if any effects on VF are caused by PAF antagonism. KEY RESULTS: Global (whole-heart) delivery of 10 microM nupafant, reduced the incidence of ischaemia-induced VF and widened QT interval without affecting coronary flow. Importantly, lower concentrations (0.1 and 1 microM) had no effect on VF, yet widened QT almost identically to 10 microM nupafant. When nupafant was delivered selectively to (and entrapped within) the involved region it partially protected against VF (P<0.05). This occurred without change in QT interval. Selective nupafant delivery to the uninvolved region was without effect. CONCLUSIONS AND IMPLICATIONS: Nupafant protects against ischaemia-induced VF primarily by site-selective actions in the ischaemic region but, unlike BN-50739, the effect is unrelated to its QT widening action, and is not compromised by any effect on coronary flow. This establishes proof of concept that VF suppression by PAF antagonism need not invariably be associated with QT prolongation or vasodilatation, justifying further development of this drug class.

Coronary aneurysms and ankylosing spondylitis: what stenting option is truly optimal?

Vascular inflammation, particularly aortitis, is a known rare manifestation of ankylosing spondylitis. This condition has not been reported to date in the coronary circulation. We present the first documented case of a coronary aneurysm in a patient with ankylosing spondylitis. This association may limit treatment options in such patients, as newer drug-eluting stents may potentiate degeneration in established aneurysmal coronary arteries.

A new method of arthroscopic reconstruction of the dislocated acromio-clavicular joint.

BACKGROUND: Symptomatic total acromio-clavicular joint dislocation (Rockwood et al. types III-VI) may be treated by surgical reconstruction. AIM: To describe an arthroscopically assisted technique to reconstruct anatomically the coraco-clavicular ligaments in acute or chronic (> 6 weeks) acromio-clavicular joint dislocation. METHODS: This new technique involves arthroscopic exposure of the coracoid process. Prior to introducing this technique, cadaveric studies were undertaken. RESULTS: Five patients underwent this procedure. All engaged in regular sports or manual-type work. All patients were discharged the same day with the shoulder immobilised for 4 weeks, with no heavy lifting for 3 months. All patients were pain-free at 6 weeks with full function and maximum Constant scores at 3 months. There have been no complications. CONCLUSIONS: A new, safe technique is described which provides a cosmetically acceptable, anatomically solid reconstruction of the coraco-clavicular ligaments.

Acute lateral dislocation of the patella: correlation of ultrasound scanning with operative findings.

Acute lateral dislocation of the patella has been associated with disruption of the medial restraints of the patella. Following non-operative management there is a re-dislocation rate of up to 44%. The purpose of this study was to test whether sonography is a reliable method of assessing the medial retinaculum after acute dislocation of the patella. Ten patients following acute patellar dislocation had an ultrasound scan (USS) performed by an experienced musculoskeletal radiologist. Each patient subsequently had an examination under anaesthetic, arthroscopy, and repair of the ruptured structures. The ultrasound reports were compared to the surgical findings to determine the accuracy of this investigation. USS located deficiencies in the ligamentous attachments to the medial border of the patella and the presence of avulsed bony fragments, all of which were confirmed at operation. The sonographic diagnosis of haematoma or torn fibres in the vastus medialis obliquus (VMO) corresponded with our operative findings. The most significant findings were the correlation of free fluid around the medial collateral ligament (MCL) with avulsion of the femoral attachment of the medial patellofemoral ligament (MPFL) and the presence of avulsed fragments of bone from the medial border of the patella.

Validation of three myocardial jeopardy scores in a population-based cardiac catheterization cohort.

BACKGROUND: The Jeopardy Score from Duke University and the Myocardial Jeopardy Index from the Bypass Angioplasty Revascularization Investigation (BARI) have been validated but never applied to a large unselected cohort. We assessed the prognostic value of these existing jeopardy scores, along with that of a new Lesion Score developed for the Alberta Provincial Project for Outcome Assessment in Coronary Heart Disease (APPROACH), a clinical data collection initiative capturing all patients undergoing cardiac catheterization in the province of Alberta. METHODS: The predictive value of these three scores were compared in a cohort of >20,000 patients (9922 treated medically, 6334 treated with percutaneous intervention, and 3811 treated with bypass surgery). Scores were considered individually in logistic regression models for their ability to predict outcome and then added to models containing sociodemographic data, comorbidities, ejection fraction, indication for procedure, and descriptors of coronary anatomy. RESULTS: All scores were found to be predictive of 1-year mortality, especially when patients are treated medically or with percutaneous intervention. In these patients, the APPROACH Lesion Score performed slightly better than the other jeopardy scores. The Duke Jeopardy Score was most predictive in those patients undergoing coronary bypass surgery. CONCLUSIONS: Myocardial jeopardy scores provide independent prognostic information for patients with ischemic heart disease, especially if those patients are treated medically or with percutaneous intervention. These scores represent potentially valuable tools in cardiovascular outcome studies. The APPROACH Lesion Score may perform slightly better than previously developed jeopardy scores.

Protection against ventricular fibrillation by the PAF antagonist, BN-50739, involves an ischaemia-selective mechanism.

The platelet-activating factor (PAF) antagonist BN-50739 can suppress certain cardiac arrhythmias. PAF is released from ischaemic myocardium and may contribute to initiation of ischaemia-induced ventricular fibrillation (VF). In this study we characterised the action of BN-50739 on left regional ischaemia-induced VF and examined whether effects are mediated within the ischaemic territory, or are nonspecific. In rat isolated Langendorff perfused hearts (n = 12/group), 10 microM BN-50739 reduced the incidence of ischaemia-induced VF from 75 to 17% (p<0.05). This was accompanied by QT widening and an increase in coronary flow. Heart rate and PR interval were not affected by the drug. In separate studies, isolated rat hearts were perfused by using a dual-lumen tube that allows independent delivery of solution to the left and right coronary beds. Successful regional localisation of drug delivery was confirmed by observing, before ischaemia, a regionally selective increase in coronary flow (p<0.05), measured by using two in-line flow meters. Protection against ischaemia-induced VF (p<0.05) was achieved by pretreatment with BN-50739, delivered selectively and entrapped within the involved region, but not when the drug was delivered to the uninvolved region. In conclusion, BN-50739 protects against ischaemia-induced VF by eliciting a pharmacologic action in the involved (ischaemic) myocardium. This supports the hypothesis that BN-50739 suppresses an arrhythmogenic effect of endogenous PAF released within the ischaemic tissue.

Inadequate ischaemia-selectivity limits the antiarrhythmic efficacy of mibefradil during regional ischaemia and reperfusion in the rat isolated perfused heart.

1. Mibefradil was compared with (+/-)-verapamil for effects on ischaemia- and reperfusion-induced ventricular fibrillation (VF), and the role of ischaemia-selective L-channel block was examined. Langendorff perfused rat hearts (n=12/group) were used. 2. Neither drug at up to 100 nM reduced the incidence of VF during 30 min regional ischaemia. 300 and 600 nM (+/-)-verapamil abolished VF (P<0. 05); mibefradil was effective only at 600 nM (P<0.05). Reperfusion-induced VF incidence was reduced only by 600 nM (+/-)-verapamil (P<0.05). Both drugs at >/=100 nM increased coronary flow (P<0.05) with a similar potency and maximum effectiveness. 3. In separate hearts perfused with Krebs' solution containing 3 mM K+ (the same as that used for arrhythmia studies) neither drug at up to 600 nM affected ventricular contractility. With K+ raised to 6 mM, (+/-)-verapamil >/=30 nM reduced developed pressure (P<0.05); mibefradil did so only at 600 nM (P<0.05). With K+ raised to 10 mM the effects of (+/-)-verapamil were further increased (P<0.05) and mibefradil became active at >/=100 nM (P<0.05). Likewise both drugs impaired diastolic relaxation, with raised K+ exacerbating the effects and (+/-)-verapamil being more potent and its effects more greatly exacerbated by K+. In contrast, when K+ was normal (3 mM), coronary flow was increased by each drug at >/=30 nM (P<0.05) indicating a marked vascular : myocardial selectivity. 4. In conclusion, mibefradil differed from (+/-)-verapamil in its myocardial effects only in terms of its lower potency. As mibefradil is the more potent T-channel blocker, the T-channel is unlikely to represent the molecular target for these effects. The K+ elevations that occur in the ischaemic milieu determine the ability of both drugs to block myocardial L-channels; this is sufficient to account for the drugs' actions on VF. Neither drug possesses sufficient selectivity for ischaemic myocardium versus blood vessels to permit efficacy (VF suppression without marked vasodilatation) and so inappropriate hypotension is likely to preclude the safe use of mibefradil (or similar analogue) in VF suppression, and explains the lack of clinical effectiveness of (+/-)-verapamil.