Journal of pharmacological and toxicological methods最新文献

{"title":"Predicting human cardiac QT alterations and pro-arrhythmic effects of compounds with a 3D beating heart-on-chip platform","authors":"Roberta Visone ,&nbsp;Caterina Pernici ,&nbsp;Ferran Lozano ,&nbsp;Marco Rasponi ,&nbsp;Paola Occhetta","doi":"10.1016/j.vascn.2025.107628","DOIUrl":"10.1016/j.vascn.2025.107628","url":null,"abstract":"<div><div>Detecting cardiac QT alterations and pro-arrhythmic effects of compounds during early stages of drug development process is still critical. Hence the development of relevant in-vitro preclinical models resembling the human heart is highly envisioned, so to effectively predict repolarization risks in humans. Here we described a human functional 3D cardiac model developed within a beating Organ-on-Chip (OoC) platform integrating fit-to-purpose assays for detecting drug-induced electrophysiological alterations in pre-clinical stages. The model, named uHeart, was developed and qualified for functional cardiotoxicity screening by following the latest International Council for Harmonization of Technical Requirements for Pharmaceuticals for Human Use S7B guidelines. uHeart platform encompasses two patented technologies: i) an actuating mechanism that mechanically trains microtissues by providing a physiological uniaxial cyclic strain (i.e., 10 % stretching, 1 Hz) and ii) a system of integrated electrodes able to read electrophysiological cardiac signals (i.e., Field Potential FP). Human induced pluripotent stem cell derived cardiomyocytes (h-iPSC-CMs, iCell) and human dermal fibroblasts (h-DF), 75 %–25 % ratio, were embedded in fibrin hydrogel (125*10^6 cells/ml) and cultured for up to 10 days. Upon achievement of functional microtissues synchronously and spontaneously beating, 11 drugs listed in the Comprehensive in vitro Proarrhytmia Assay (CiPA) and affecting single or multiple cardiac ion-channels were selected to qualify the model. Drug-induced alterations were evaluated at incremental doses and by analyzing the FP morphology (i.e., beating period-BP, spike amplitude-AMP, FP-duration-FPD) and the onset of arrhythmic events. DMSO and Aspirin were used as vehicle and negative controls, respectively. Microtissues beat synchronously after 5 days, and FP signals showed the typical depolarization and re-polarization spikes. Ikr blockers (e.g., Dofetilide, Quinidine) prolonged the FPD at concentration near the Cmax. ICaL blockers (e.g., Verapamil, Nifedipine) shortened it at 5 and 50 nM respectively. Mexiletine, blocking INa, statistically decreased the AMP at 10 μM. Both Terfenadine and Dofetilide elicited arrhythmic events, matching FDA labels indications. Overall, the system showed 83.3 % sensitivity, 100 % specificity and 91.6 % accuracy in detecting FPD prolongation. DMSO (up to 0.5 % <em>w</em>/<em>v</em>) and Aspirin (up to 100 μM) did not statistically alter the repolarization time. uHeart generates functional 3D cardiac in-vitro models, predicting compound specific toxic effects at concentrations near the Cmax, thus resulting suitable to perform functional cardiotoxicity preclinical drug screening.</div></div>","PeriodicalId":16767,"journal":{"name":"Journal of pharmacological and toxicological methods","volume":"133 ","pages":"Article 107628"},"PeriodicalIF":1.3,"publicationDate":"2025-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143927389","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Using SPAR to analyze arterial blood pressure waves, following fostamatinib and entospletinib administration in radiotelemetry in rats, to aid in cardiovascular safety testing","authors":"Marieke Van Daele ,&nbsp;Miquel Serna Pascual ,&nbsp;Gurleen Virk ,&nbsp;Matt Skinner ,&nbsp;Manasi Nandi ,&nbsp;Stephen J. Hill ,&nbsp;Jeanette Woolard","doi":"10.1016/j.vascn.2025.107609","DOIUrl":"10.1016/j.vascn.2025.107609","url":null,"abstract":"&lt;div&gt;&lt;div&gt;Cardiovascular safety liabilities represent a leading cause of drug attrition, and improved preclinical measurements are essential to predict drug-related toxicities (Cook et al., 2014; Weaver &amp; Valentin, 2019). Presently, radiotelemetry approaches recording blood pressure are routinely used in preclinical in vivo assessments. Typically, single points of these waveforms (e.g. mean arterial pressure (MAP)) are used for analysis. Although MAP provides important insights, this approach overlooks potential information hidden in changes in waveform morphology and variability. The Symmetric Projection Attractor Reconstruction (SPAR) is a novel mathematical method that enables detailed wave analysis, transforming lengthy waveform recordings into contained visual 2D representations (attractors) (Nandi &amp; Aston, 2020). By subsequent analysis of key features of the attractor (e.g. rQ25, indicating the opening of the attractor), subtle changes in the cardiovascular system can be detected. The objective of the present study was to explore in-depth characterization of arterial blood pressure waves after drug administration. Fostamatinib, a Syk-inhibitor causing hypertension due to off-target VEGFR2-inhibition, and entospletinib, designed for better selectivity and less adverse effects, are presented as an example of how in-depth wave analysis could improve cardiovascular safety testing. Male Wistar rats were instrumented with Stellar telemetry implants. Arterial blood pressure was recorded over two consecutive days, for at least 1 h before and up to 23 h after daily administration of fostamatinib (20 mg/kg, p.o.) or entospletinib (6 mg/kg, p.o), alongside a vehicle control (10 mL/kg, p.o.). Fostamatinib increased MAP significantly on day 2 (&lt;em&gt;p&lt;/em&gt; &lt; 0.05, Two-way ANOVA); there was no significant effect on MAP on day 1. Entospletinib did not affect MAP significantly. Fostamatinib caused changes in waveform morphology, that were quantified using attractor parameters. Attractor opening increased significantly from day 1 of fostamatinib administration (e.g. rQ25 &lt;em&gt;p&lt;/em&gt; &lt; 0.05, Two-way ANOVA). These fostamatinib-induced attractor changes were opposite to the ones observed with vasodilators, associated with a decrease in attractor opening, therefore indicating that fostamatinib caused a vasoconstriction. This is consistent with literature, describing a fostamatinib-induced vasoconstriction due to VEGFR2-inhibition (Skinner et al. 2014). Moreover, entospletinib had similar but smaller effects on attractor opening (delta rQ25 &lt;em&gt;p&lt;/em&gt; &lt; 0.0005 on day 1, Two-way ANOVA). Given that SPAR could detect fostamatinib-induced cardiovascular effects earlier than MAP, and SPAR could detect entospletinib-induced changes in a more sensitive way than MAP, these data suggest that more extensive blood pressure waveform analysis using SPAR in telemetered rats may provide nuanced information that could aid in informed decision making and earlier de","PeriodicalId":16767,"journal":{"name":"Journal of pharmacological and toxicological methods","volume":"133 ","pages":"Article 107609"},"PeriodicalIF":1.3,"publicationDate":"2025-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143927604","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Characterization of drugs targeting the Mu opioid receptor (MOR) using multivariate quantitative electroencephalography (qEEG) and sleep/wake metrics","authors":"Monica R. Metea ,&nbsp;Ernesto P. Solar ,&nbsp;Bethany E. Pierce ,&nbsp;Robert W. Gould","doi":"10.1016/j.vascn.2025.107667","DOIUrl":"10.1016/j.vascn.2025.107667","url":null,"abstract":"<div><div><em>In vivo</em> qEEG assessments support rapid decision-making on compound/dose selection and safety in the course of preclinical drug development, based on the sensitivity of EEG to drug-induced brain oscillatory and synaptic changes. The present study utilizes a streamlined ‘workbench’ platform approach to generate multivariate qEEG and sleep metrics to characterize changes in oscillatory brain activity of mu opioid receptor (MOR) partial and full agonists spanning antinociceptive and reinforcing dose ranges. Male Sprague-Dawley rats were implanted with wireless EEG devices and administered saline, full MOR agonists, morphine (0.1–10 mg/kg i.p.) or oxycodone (1–5.6 mg/kg, i.p.), partial agonist, buprenorphine (0.05–0.5 mg/kg, s.c.) or a MOR antagonist, naltrexone (0.3–3 mg/kg s.c.). 24-h EEG recordings were sleep-scored followed by state-dependent EEG spectral power analyses. EEG spectral distributions were computed for each vigilance state (wake, REM, NREM). Statistical analyses were performed for each compound and a principal component analysis (PCA)-based algorithm was employed to reduce data dimensionality and identify components representing the maximum variance over the combined multivariate dataset. All agonists led to dose-related increases in time awake although high doses produced altered oscillatory activity associated with sedation, whereas naltrexone increased sleep duration. Principal components enabled class separation aligned with analgesic potency. These data support the feasibility of using a sensitive and streamlined multivariate qEEG approach for fast characterization of CNS active drugs, with potential future applications for aiding rapid analgesic screening and development aimed at reducing abuse liability.</div></div>","PeriodicalId":16767,"journal":{"name":"Journal of pharmacological and toxicological methods","volume":"133 ","pages":"Article 107667"},"PeriodicalIF":1.3,"publicationDate":"2025-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143927614","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"In vitro systems for the assessment of chronic cardiotoxic effects: News from the HESI stem cell working group","authors":"Matthias Gossmann ,&nbsp;Ulrich Thomas ,&nbsp;Elena Dragicevic ,&nbsp;Ravi Vaidyanathan ,&nbsp;Bettina Lickiss ,&nbsp;Ouissame Filali ,&nbsp;Sonja Stoelzle-Feix ,&nbsp;Peter Linder","doi":"10.1016/j.vascn.2025.107687","DOIUrl":"10.1016/j.vascn.2025.107687","url":null,"abstract":"<div><div>The great promise of human induced pluripotent stem cell-derived cardiomyocytes (hiPS-CMs) as <em>in vitro</em> models for the assessment of proarrhythmic risk, was extensively studied by the Comprehensive <em>In Vitro</em> Proarrhythmia Assay (CiPA) initiative. Since long-term exposure of cancer-related therapeutics have been linked to alterations of cardiac function in patients, the Stem Cell Working Group as part of the Health and Environmental Science Institute (HESI) endeavors to gain further insight into chronic cardiotoxicity. The objective of the study was to optimize non-clinical safety assessment strategies of chronic cardiotoxicity by testing prolonged exposure of reference compounds on cell-based assay systems using hiPSC-CMs. Here, we present an extract of the HESI chronic cardiotoxicity study with 8 different compounds known to act <em>via</em> diverse mechanisms of actions (MoAs) in cardiomyocytes, affecting cardiac energetics (doxorubicin, erlotinib, sunitinib), contractility (BMS-986094, Nilotinib), electrophysiology (pentamidine), and myofilament organization (endothelin-1, vincristine). Each compound was tested on iCell Cardiomyocytes² (FUJIFILM Cellular Dynamics Inc.) at four different concentrations over a time frame of 144 h. Changes in contractile properties were assessed using FLEXcyte technology, a 96-well assay system with specialized plates that enable monitoring cardiac contractility in a label-free manner. The results show alterations in cardiac function related to the MoA of the respective compound, as well as time- and/or dose-dependent effects. For instance, Doxorubicin and BMS 986094 treatment both exhibit a concentration-dependent decrease in amplitude over time, by acting either through mitochondrial toxicity (Doxorubicin) or impaired calcium handling (BMS 986094). This excerpt of the HESI Stem Cell Working Group study underlines the potential of <em>in vitro</em> systems to address contractile function of hiPSC-CMs for chronic safety pharmacological studies of compounds with diverse MoAs.</div></div>","PeriodicalId":16767,"journal":{"name":"Journal of pharmacological and toxicological methods","volume":"133 ","pages":"Article 107687"},"PeriodicalIF":1.3,"publicationDate":"2025-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143927616","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Recommendations on the measurement of electrocardiogram and hemodynamic parameters in restrained non-rodent species in regulatory safety assessment studies","authors":"Jean-Pierre Valentin ,&nbsp;Annie Delaunois ,&nbsp;C. Michael Foley ,&nbsp;Kim A. Henderson ,&nbsp;Pierre Lainee ,&nbsp;Derek J. Leishman ,&nbsp;Michael K. Pugsley ,&nbsp;Sridharan Rajamani ,&nbsp;Christopher P. Regan ,&nbsp;Eric I. Rossman ,&nbsp;Rebecca Ross ,&nbsp;Stephen D. Tichenor ,&nbsp;Hugo M. Vargas ,&nbsp;Todd A. Wisialowski","doi":"10.1016/j.vascn.2025.107601","DOIUrl":"10.1016/j.vascn.2025.107601","url":null,"abstract":"<div><div>Life supportive cardiovascular (CV) parameters [e.g., arterial blood pressure (BP), electrocardiogram (ECG), heart rate (HR)] are recorded in non-rodent safety pharmacology and toxicology studies and data are used to support pharmaceutical drug development in accordance with several guidelines (e.g., ICH M3, S6, S7, S9). Various methodological approaches are used to collect CV parameters on toxicology studies, including fully implantable telemetry, non-invasive jacket telemetry, and short-duration restraint-based measurement from surface ECG leads and a blood pressure cuff, but their robustness and applicability domain differ widely. The challenge of using conscious animals, especially non-human primates, is that arousal behavior, locomotion, body temperature, and sympathetic activation are significant sources of variability that impact CV parameters, thus telemetry methods are the preferred approach. Historically, toxicology studies have incorporated short-duration restraint methods (manual; chemical sedation) to collect brief periods (“a snapshot at one timepoint”) of CV endpoints in a large number of non-rodents (<em>N</em> ≥ 24) after repeat dosing. Non-rodent species, however, demonstrate increased BP and HR in response to room entry and/or manual restraint, which confounds the interpretation of drug-related effects. Published case examples and company experiences indicate that CV waveforms collected under restraint are of low quality, can vary based on body position and sensor placement (skin electrode; pressure cuff), as well as the type of sedative and dose used. In addition, the data captured can be misleading or incongruent with CV safety pharmacology findings in the same animal species. As a result, restraint-induced stress lowers the sensitivity to detect CV changes as measured over a short-duration and has a higher probability for false negative findings. Although functional CV evaluations in restrained animals have been included in toxicology studies for decades, there is no industry consensus on methods, assay performance, or value (translation) for risk assessment. This communication will review the available literature and leverage pharmaceutical and Contract Research Organization (CRO) experiences, and propose recommendations, with an emphasis on short-duration techniques, from the ICH E14/S7B Industry Support Group with the aim to identify how, when, and if short term restraint-based CV data are valuable for nonclinical safety assessment.</div></div>","PeriodicalId":16767,"journal":{"name":"Journal of pharmacological and toxicological methods","volume":"133 ","pages":"Article 107601"},"PeriodicalIF":1.3,"publicationDate":"2025-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143927787","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Organelle profiling of compound-induced effects in human iPSC-derived cardiomyocytes in combination with electrophysiology assays for in vitro prediction of cardiac safety","authors":"Brigitta R. Szabo ,&nbsp;Elena Matsa ,&nbsp;Georgios Kosmidis ,&nbsp;Paul Volders","doi":"10.1016/j.vascn.2025.107652","DOIUrl":"10.1016/j.vascn.2025.107652","url":null,"abstract":"<div><div>Methods with suboptimal efficiency for detecting cardiovascular side-effects burden the pharmaceutical industry. As scalable, human-based alternatives to traditional models, hiPSC-CMs show great promise in the field. However, for widespread industrial application, high quality validation studies are critical. Our goal was to establish a highly predictive in vitro hiPSC-CM drug screening protocol leveraging the power of morphological profiling multiplexed with established electrophysiological readouts (multi-electrode array; MEA). Three healthy control hiPSC-CM lines were cultured in serum-free conditions and then treated with a library of seventeen compounds at ranges comparable to maximal clinical plasma concentrations. High content imaging assays for sarcomeres, mitochondria, DNA damage, Golgi, endoplasmic reticulum, gap junctions, peroxisomes and lysosomes were validated in a 384 well plate format. Morphological data was analyzed in combination with MEA recordings. For deeper mechanistic insight, RNA sequencing was also performed. As expected, positive control, doxorubicin reduced viability up to 70 %. Investigating its effect on each readout served as a key step in establishing proof of concept. In accordance with its mechanism of action, dose-dependent increase in γH2AX (marker of DNA damage) was present with minor differences in sensitivity between cell lines. Lysosomal, nucleolar, and gap junction morphology was affected as well. Electrophysiological activity was altered even at low concentrations, while arrhythmia/quiescence was detected &gt;1 μM. With 0.1 μM Doxorubicin, 125 genes were differentially expressed compared to vehicle, several of which were involved in cellular responses to DNA damage and the p53 pathway. Notably, TOP2A, a marker of DNA stress and a target of doxorubicin, was significantly downregulated. For all 17 drugs tested, each parameter was examined. All parameters were collated for bioinformatic analysis. Collecting such an elaborate set of features is fundamental for profiling assays. First, principal component analysis revealed compound effects that otherwise remained hidden. Second, to build a predictive cardiac safety score, partial least squares-discriminant analysis revealed specific spatial clustering of compounds with potential toxicity. Hence, morphological analysis in combination with traditional readouts and bioinformatics enables deeper understanding and in vitro prediction of compound activity and toxicity. Drug-induced deregulation in pathways provides mechanistic explanations for the structural and functional changes in hiPSC-CMs.</div></div>","PeriodicalId":16767,"journal":{"name":"Journal of pharmacological and toxicological methods","volume":"133 ","pages":"Article 107652"},"PeriodicalIF":1.3,"publicationDate":"2025-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143927848","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Phenotypic-based assays for enhanced cardiac safety attrition in drug development","authors":"Margaux Boos,&nbsp;Nicolas Redon,&nbsp;Agnes Jacquet,&nbsp;Véronique Ballet,&nbsp;Ambroise Garry","doi":"10.1016/j.vascn.2025.107653","DOIUrl":"10.1016/j.vascn.2025.107653","url":null,"abstract":"&lt;div&gt;&lt;div&gt;In the field of drug discovery and development, ensuring the safety and efficacy of potential therapeutic candidates is critical. However, the attrition rate during drug development due to safety concerns remains a major challenge for the pharmaceutical industry. Unexpected toxicities still account for 20–30 % of clinical trial failures, in part due to the persistence of animal testing as the primary approach for de-risking new drugs. To address this issue, a paradigm shift has taken place towards the use of human cell-based phenotypic assays, which offer a comprehensive and biologically relevant approach to assess safety profiles and reduce the number of late-stage failures. Traditional methods for drug safety assessment often rely on biochemical and genetic assays, which do not always fully capture the dynamic phenotypic changes occurring in human cells. To address this limitation, holotomography has emerged as a promising technique that offers label-free, high-resolution imaging capabilities for the study of cellular morphology and function. Holotomography uses quantitative phase imaging techniques, such as digital holography and tomography, to generate three-dimensional reconstructions of cellular structures with nanoscale resolution. By extracting information about refractive index variations within cells, holotomography provides valuable insights into cellular architecture, intracellular dynamics, and morphological alterations induced by drug treatments. This label-free nature of holotomography enables real-time monitoring of cellular responses, minimizing disruption caused by exogenous dyes or stains. Holotomography makes it possible to examine a wide range of function, including apoptosis necrosis, lipid droplets formation, mitochondrial tracking and cell killing. To identify the compounds inducing adverse cardiovascular events, we have developed, in collaboration with Nanolive, a holotomography-based assay in a 96-well plate format, using cardiomyocytes derived from human induced-pluripotent stem cells (hiPSC-CMs) and the 3D Cell Explorer platform. In this study, we evaluated a panel of drugs inducing phospholipidosis, including amiodarone, propanolol, ellipticine, ifenprodil, metergoline, methiothepin, mibefradil, ouabain, spiperone, tamoxifen, promazine and verapamil at different doses (concentration range from 0.03 to 100 μM). After 24 h–72 h of treatment, holotomography allows for imaging of cellular structures, including the ultrastructural changes associated with phospholipidosis. It helps visualize the lamellar bodies within lysosomes and provides a detailed analysis of the intracellular lipid accumulation. Different parameters were used to quantify phospholipidosis: ratio of total lipid droplets area to the parent cell area (%), mean lipid droplets granularity per cell and mean lipid droplets dry mass per cell (pg). All compounds tested induced phospholipidic effects based on assessment of these parameters, and results ","PeriodicalId":16767,"journal":{"name":"Journal of pharmacological and toxicological methods","volume":"133 ","pages":"Article 107653"},"PeriodicalIF":1.3,"publicationDate":"2025-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143927849","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Cardiovascular functional and structural alterations induced by VEGFR-2 inhibitors axitinib and lenvatinib","authors":"Patrizia Pannucci ,&nbsp;Dustin N. Kruger ,&nbsp;Callan D. Wesley ,&nbsp;Pieter-Jan D.F. Guns ,&nbsp;Jeanette Woolard","doi":"10.1016/j.vascn.2025.107656","DOIUrl":"10.1016/j.vascn.2025.107656","url":null,"abstract":"<div><div>Antiangiogenic therapies, also known as vascular endothelial growth factor receptor inhibitors (VEGFRIs), represent a novel approach in cancer treatment that has significantly improved cancer prognosis and survivorship. However, several cardiovascular complications have been identified as associated to these therapeutics, without being anticipated during preclinical evaluations. This discrepancy draws attention to the lack of predictive and sensitive preclinical strategies to predict the VEGFRIs-induced cardiovascular toxicities, which mainly include hypertension, cardiac dysfunction and thromboembolism. The lack of clarity regarding the extent to which VEGF inhibitors impact cardiovascular function complicates the therapeutic management of cancer patients developing such adverse effects. The severity of cardiovascular toxicities associated with VEGFR inhibitors often leads to dose reduction or interruption of these potentially lifesaving therapies. As a result, this study aims to characterize the impact of RTKIs targeting VEGFR-2 on cardiovascular physiology, as well as to improve the predictive value of existing preclinical approaches, in order to better define and limit the RTKI-induced cardiovascular toxicities. Male 10-week-old C57BL/6J mice were dosed with axitinib (12 mg.kg⁻¹), lenvatinib (4 mg.kg⁻¹) and vehicle (40 %HPβCD) as an intravenous injection for 4 consecutive days. On day 2 of the treatment period, echocardiography was performed. On day 4 animals were randomly divided into two clusters: cardiac function was assessed via PV-loop experiments and vascular reactivity was investigated using isolated organ baths. Lenvatinib evoked a significant decrease in the systolic function (with a drop of 24 % and 28 % for LVEF and FS, respectively), while also affecting LV internal diameter and wall thickness. The invasive evaluation of haemodynamic parameters via PV-loop showed that lenvatinib produced a notable increase in the slope of the end systolic pressure volume relationship. In addition, neither axitinib nor lenvatinib altered phenylephrine-induced contraction, both in presence and absence of L-NAME, as well as they did not affect Ach-induced endothelium-dependent relaxation and DEANO-induced endothelium-independent relaxation. This study showed that lenvatinib treatment is associated with systolic dysfunction and enlargement of the left ventricle, while also affecting cardiac contractility. Conversely, axitinib did not impact cardiac performance. In terms of vascular reactivity, neither axitinib and lenvatinib were associated with alteration of aortic stiffness or endothelial dysfunction.</div></div>","PeriodicalId":16767,"journal":{"name":"Journal of pharmacological and toxicological methods","volume":"133 ","pages":"Article 107656"},"PeriodicalIF":1.3,"publicationDate":"2025-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143927852","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Protocol optimization to improve performance of an automated patch clamp system on assessing hERG activity for challenging compounds","authors":"Liang Guo,&nbsp;Catherine Garrison,&nbsp;Jingsong Fan,&nbsp;Jinqing Li,&nbsp;Paul Levesque","doi":"10.1016/j.vascn.2025.107640","DOIUrl":"10.1016/j.vascn.2025.107640","url":null,"abstract":"<div><div>As block of Ikr channel (encoded by hERG gene) delays the membrane repolarization in myocardia, leading to the acquired QT prolongation and potentially a fatal ventricular arrhythmia TdP, thorough scrutiny on hERG liability is required for every small molecule prior to clinical development. This can be achieved early in compound profiling by high-throughput radiometric binding, thallium flux or automated patch clamp (APC) assays; however, screening platforms often underestimate hERG inhibition potency when compared with the gold-standard, manual patch clamp (MPC) assay. To address this issue, ~90 discovery compounds were selected, having hERG MPC IC50s spanning from 13 nM to 30 μM and exhibiting a positive correlation with hydrophobicity (cLogP) or molecular weight but not with the polar surface area (PSA). The averaged hERG IC50s from binding, thallium, or APC (SyncoPatch-384) assays were 18×, 14× or 7× of MPC IC50s, respectively, for the whole compound set, and increased to 36×, 25× or 14× for compounds with an MPC IC50s ≤1 μM, confirming suboptimal assay performance and suggesting hydrophobicity as a culprit. To evaluate the optimized APC protocol—which employed a process of on-run testing solution preparation, 3-repeat compound applications and a 35-min exposure, in comparison with a single application, typical 5-min exposure protocol—a subset of 10 “challenging” compounds with an averaged APC-IC50 24× of MPC IC50s were selected along with cisapride and dofetilide. Cisapride showed fast inhibition kinetics with an IC50 (~10 nM) obtained both at 5-min and at 35-min exposure, whereas dofetilide exhibited a slow inhibition profile with the IC50 decreasing from 16 nM at 5-min to 0.9 nM at 35-min exposure (an 18× increase in potency). Half of 10 subset compounds showed cisapride-like fast inhibition and half showed dofetilide-like slow inhibition over 35-min exposure, which appeared to be cLogP-dependent (&lt;3.54 or &gt; 4.08, respectively). Compound loss and/or slow inhibition kinetics were identified as responsible for APC underestimation. Importantly, APC IC50s from the optimized protocol matched MPC IC50s, improved performance was further confirmed with a large compound set tested in a screening mode, capable of assessing 96 compounds per plate (3-sequential dosings/well, 12-min exposure/dose, 4-replicate wells/drug).</div></div>","PeriodicalId":16767,"journal":{"name":"Journal of pharmacological and toxicological methods","volume":"133 ","pages":"Article 107640"},"PeriodicalIF":1.3,"publicationDate":"2025-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143927856","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Recording of transcellular voltage deflections from cardiomyocytes on MEA devices using laser poration","authors":"Timm Danker,&nbsp;Udo Kraushaar,&nbsp;Jasmin Schäfer","doi":"10.1016/j.vascn.2025.107645","DOIUrl":"10.1016/j.vascn.2025.107645","url":null,"abstract":"<div><div>The occurrence of life-threatening cardiac arrhythmia induced by certain drugs is frequently preceded by an extended duration of cardiac action potentials (AP), which are often associated with minor proarrhythmic fluctuations in the membrane potential. The configuration and temporal progression of the repolarizing segment of the AP plays a crucial role in determining whether arrhythmia will occur or not. Microelectrode arrays (MEA), a well-established tool in research and cardiac safety pharmacology, provide a straightforward method for observing the effects of cardiotoxic compounds through extracellular field potentials (FP). However, despite the significant utility of MEA, the waveform of the FP does not provide a clear representation of the original AP shape. This limitation is due to the extracellular recording principle and the inherent alternating current (AC) filtering that results from it. To overcome this challenge, the IntraCell device (Foresee Biosystems) has been evaluated. This innovative device can repeatedly breach the membrane of cardiomyocytes that are cultivated on top of the MEA electrodes at various cultivation time points. This is achieved through the use of a highly focused nanosecond laser beam. The process of laser poration transforms the electrophysiological signal from FP to intracellular-like APs, referred here as laser-induced APs (liAP), and facilitates the recording of transcellular voltage deflections. Amplitudes increased by 4.1 ± 0.41 (<em>n</em> = 20, range 1.34–8.83) times, analyzed from a randomly picked subset of recordings, resulting in amplitudes between 7 and 22 mV. Although the overall pharmacological sensitivity appeared unaltered when challenged with commonly used tool compounds (e.g. Nifedipine IC50: FP 0.282 ± 0.05 μM, <em>n</em> = 3; LiAP 0.164 ± 0.04 μM n = 3; E4031 prolongation compared to control @ 0.01 μM FP 1.4 ± 0.29 times n = 3; LiAP 4.16 ± 0.55 times n = 3), this access to the intracellular environment allows for a more accurate depiction of the AP shape and a more sensitive and precise classification of proarrhythmic potentials compared to standard MEA recordings. This novel system represents a groundbreaking extension to the existing electrophysiological methods. It allows for a more accurate evaluation of cardiotoxic effects while retaining all the benefits of MEA-based recordings. These benefits include ease of use, the ability to conduct acute and chronic experiments, and the capacity for signal propagation analysis, among others. Therefore, this new device offers a significant advancement in the field of cardiac safety pharmacology and has the potential to greatly enhance our understanding of drug-induced cardiac arrhythmias.</div></div>","PeriodicalId":16767,"journal":{"name":"Journal of pharmacological and toxicological methods","volume":"133 ","pages":"Article 107645"},"PeriodicalIF":1.3,"publicationDate":"2025-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143927957","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}