Innovative approaches to cardiovascular safety pharmacology assessment.

This editorial prefaces the annual themed issue on safety pharmacology (SP) methods which has been published since 2004 in the Journal of Pharmacological and Toxicological Methods (JPTM). Here we highlight content derived from the 2023 Safety Pharmacology Society (SPS) meeting held in Brussels, Belgium. The meeting generated 138 abstracts, reproduced in the current volume of JPTM. As in prior years, the manuscripts reflect various areas of innovation in SP including in silico modeling of stroke volume, cardiac output and systemic vascular resistance, computational approaches that compare drug-induced proarrhythmic sensitivity of human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs), an evaluation of the utility of the corrected J-Tpeak and Tpeak-to-Tend parameters from the ECG as potential proarrhythmia biomarkers, and the applicability of nonclinical concentration-QTc (C-QTc) modeling of data derived from the conduct of the in vivo QTc study as a component of the core battery of safety pharmacology studies.

Development of a pharmaceutical database as an aid to the nonclinical detection of drug-induced cardiac toxicity.

The Health and Environmental Sciences Institute (HESI) Cardiac Safety Committee designed and created a publicly accessible database with an initial set of 128 pharmacologically defined pharmaceutical agents, many with known cardiotoxic properties. The database includes specific information about each compound that could be useful in evaluating hypotheses around mechanisms of drug-induced cardiac toxicity or for development of novel cardiovascular safety assays. Data on each of the compounds was obtained from published literature and online sources (e.g., DrugBank.ca and International Union of Basic and Clinical Pharmacology (IUPHAR) / British Pharmacological Society (BPS) Guide to PHARMACOLOGY) and was curated by 10 subject matter experts. The database includes information such as compound name, pharmacological mode of action, characterized cardiac mode of action, type of cardiac toxicity, known clinical cardiac toxicity profile, animal models used to evaluate the cardiotoxicity profile, routes of administration, and toxicokinetic parameters (i.e., Cmax). Data from both nonclinical and clinical studies are included for each compound. The user-friendly web interface allows for multiple approaches to search the database and is also intended to provide a means for the submission of new data/compounds from relevant users. This will ensure that the database is constantly updated and remains current. Such a data repository will not only aid the HESI working groups in defining drugs for use in any future studies, but safety scientists can also use the database as a vehicle of support for broader cardiovascular safety studies or exploring mechanisms of toxicity associated with certain pharmacological modes of action.

Evaluation of the translation of multiple cardiovascular regulatory mechanisms in the anesthetized dog.

The strategic and targeted use of an anesthetized canine cardiovascular model early in drug discovery enables a comprehensive cardiovascular and electrophysiological assessment of potential safety liabilities and guides compound selection prior to initiation of chronic toxicological studies. An ideal model would enable exposure-response relationships to guide safety margin calculations, have a low threshold to initiate, and have quick delivery of decision quality data. We have aimed to profile compounds with diverse mechanism of actions (MoAs) of "non-QT" cardiovascular drug effects and evaluate the ability of nonclinical in vivo cardiovascular models to detect clinically reported effects. The hemodynamic effects of 11 drugs (atropine, itraconazole, atenolol, ivabradine, milrinone, enalaprilat, fasudil, amlodipine, prazosin, amiloride, and hydrochlorothiazide) were profiled in an anesthetized dog cardiovascular model. Derived parameters included: heart rate, an index of left ventricular contractility, mean arterial pressure, systemic vascular resistance, and cardiac output. Species specific plasma protein data was generated (human, dog) and utilized to calculate free drug concentrations. Using the anesthetized dog cardiovascular model, 10 of the 11 drugs displayed the predicted changes in CV parameters based on their primary MoAs and corresponding clinically described effects. Interestingly but not unexpected, 1 of 11 failed to display their predicted CV pattern which is likely due to a delay in pharmacodynamic effect that is beyond the duration of the experimental model (hydrochlorothiazide). The analysis from the current study supports the strategic use of the anesthetized dog model early in the drug discovery process for a comprehensive cardiovascular evaluation with good translation to human.

Preclinical cardiovascular safety assessment of pharmacology-toxicology relationship for a set of novel kinase inhibitors.

Cardiovascular toxicity is one of the more common causes of attrition in preclinical and clinical drug development. Preclinical cardiovascular safety assessment involves numerous in vitro and in vivo endpoints which are being continually reviewed and improved to lower the incidence of cardiovascular toxicity that manifests only after the initiation of clinical trials. An example of notable preclinical toxicity is necrosis in the papillary muscle of the left ventricle in dogs that is induced by exaggerated pharmacological effects of vasodilators or positive inotropic/vasodilating off-target drug effects. Two distinct, small-molecule inhibitors that target an intracellular kinase, Compound A and Compound B, were profiled in 2-week dose-range finding and 4-week toxicity studies. Serum cardiac troponin (cTnI) was evaluated after a single dose and after 2-week and 4-week repeat dose studies with each kinase inhibitor. Acute effects on hemodynamic (heart rate, blood pressures, left ventricular contractility) and electrocardiographic (QTcV, PR, QRS intervals) endpoints by each inhibitor were assessed in an anesthetized dog cardiovascular model. Cardiovascular degeneration/necrosis with and without fibrosis was observed in dogs and correlated to increases in serum cTnI in repeat-dose toxicity studies. At the same doses used in toxicologic assessments, both kinase inhibitors produced sustained increases in heart rate, left ventricular contractility, and cardiac output, and decreases in mean arterial pressure. Cardiac pathology findings associated with these 2 kinase inhibitors were accompanied not only by cardiac troponin elevations but also associated with hemodynamic changes, highlighting the importance of the link of the physiologic-toxicologic interplay in cardiovascular safety assessment.

Systemic juvenile idiopathic arthritis: The Great Ormond Street Hospital experience (2005-2021).

Systemic juvenile idiopathic arthritis (sJIA) is a complex, systemic inflammatory disorder driven by both innate and adaptive immunity. Improved understanding of sJIA pathophysiology has led to recent therapeutic advances including a growing evidence base for the earlier use of IL-1 or IL-6 blockade as first-line treatment. We conducted a retrospective case notes review of patients diagnosed with sJIA over a 16-year period (October 2005-October 2021) at Great Ormond Street Hospital for Children. We describe the clinical presentation, therapeutic interventions, complications, and remission rates at different timepoints over the disease course. We examined our data, which spanned a period of changing therapeutic landscape, to try and identify potential therapeutic signals in patients who received biologic treatment early in the disease course compared to those who did not. A total of 76-children (female n = 40, 53%) were diagnosed with sJIA, median age 4.5 years (range 0.6-14.1); 36% (27/76) presented with suspected or confirmed macrophage activation syndrome. A biologic disease-modifying anti-rheumatic drug (bDMARD) alone was commenced as first-line treatment in 28% (n = 21/76) of the cohort; however, at last review, 84% (n = 64/76) had received treatment with a bDMARD. Clinically inactive disease (CID) was achieved by 88% (n = 67/76) of the cohort at last review; however, only 32% (24/76) achieved treatment-free CID. At 1-year follow-up, CID was achieved in a significantly greater proportion of children who received treatment with a bDMARD within 3 months of diagnosis compared to those who did not (90% vs. 53%, p = 0.002). Based on an ever-increasing evidence base for the earlier use of bDMARD in sJIA and our experience of the largest UK single-centre case series described to date, we now propose a new therapeutic pathway for children diagnosed with sJIA in the UK based on early use of bDMARDs. Reappraisal of the current National Health Service commissioning pathway for sJIA is now urgently required.

Safety pharmacology 2023 and implementation of the ICH E14/S7B Q&A guidance document.

This editorial prefaces the annual themed issue on safety pharmacology (SP) methods published since 2004 in the Journal of Pharmacological and Toxicological Methods (JPTM). We highlight here the content derived from the recent 2022 Safety Pharmacology Society (SPS) and Canadian Society of Pharmacology and Therapeutics (CSPT) joint meeting held in Montreal, Quebec, Canada. The meeting also generated 179 abstracts (reproduced in the current volume of JPTM). As in previous years the manuscripts reflect various areas of innovation in SP including a comparison of the sensitivity of cross-over and parallel study designs for QTc assessment, use of human-induced pluripotent stem cell (hi-PSC) neuronal cell preparations for use in neuropharmacological safety screening, and hiPSC derived cardiac myocytes in assessing inotropic adversity. With respect to the latter, we anticipate the emergence of a large data set of positive and negative controls that will test whether the imperative to miniaturize, humanize and create a high throughput process is offset by any loss of precision and accuracy.

A method for assessing and monitoring consistency of nonclinical ECG analysis.

Cardiovascular safety is a key area of concern for new drugs in development, and the collection and analysis of electrocardiograms (ECGs) is a standard and major component of nonclinical testing. Digital data capture technology allows for high-throughput and long-duration ECG collections, resulting in large volumes of data. Consistent analysis of these ECG data is critical for detecting meaningful changes during nonclinical drug development. We developed a method to assess the consistency of nonclinical ECG analysis for a group of analysts over time. Eight­lead ECGs were collected from conscious dogs using Ponemah (v5.2, DSI). Analysts placed Pstart, Qstart, Rpeak, Send, and Tend marks on six waveforms for each animal. The ECG files were randomized and re-marked under blinded conditions 4 to 14 days following initial mark placement. Averages of each parameter measured (RR interval, QRS duration, PR interval, and QT interval) were compiled for each marking session and analyst. A Gage R&R evaluation was completed. Graphical output from the Gage R&R evaluation showed distinct variability on group and individual analyst levels. Differences in inter- and intra-analyst variability (reproducibility and repeatability, respectively) were observed between trained analysts and analysts in training. The Gage R&R method is an effective tool for assessing consistency of digital ECG mark placement at a group level. Furthermore, it is able to identify areas of improvement for individual ECG analysts and to assess ECG analyst consistency during their training period. The assessment results are useful for facilitating discussions on best practices and maintaining consistency of mark placement.

Male genital lichen sclerosus associated with urological interventions and microincontinence: a case series of 21 patients.

BACKGROUND: Male genital lichen sclerosus (MGLSc) is an acquired, chronic, inflammatory cutaneous disease associated with significant morbidity and squamous cell carcinoma of the penis. Consideration of all of the evidence suggests that chronic exposure of susceptible epithelium to urinary occlusion by the foreskin is the most likely pathomechanism. MGLSc never occurs in men who were circumcised at birth, and has been associated with trauma, instrumentation and anatomical abnormalities, e.g. frank hypospadia that results in microincontinence. AIM: To describe 21 patients who developed MGLSc following urological diagnoses and procedures. METHODS: We conducted a retrospective review of patients with a diagnosis of MGLSc whose symptoms related to urological procedures who attended or saw one of the authors (CBB) privately during the period June-October 2018. RESULTS: In total, 21 patients (mean age 59 years) were identified. The referrals came from the local urology departments, primary care or extramural dermatology services. Most of the patients were uncircumcised men. All had developed symptoms and signs of MGLSc within 5 years following their urological procedure; on examination, 30% of the patients were found to have damp penile skin due to microincontinence. Of the 21 patients, 10 had undergone radical prostatectomy for prostate cancer, 4 had a diagnosis of Peyronie disease, 4 had undergone multiple cystoscopies and urethroscopies, 2 had undergone surgery on the bladder neck and 1 had undergone implantation of a penile prosthesis to treat erectile dysfunction. CONCLUSION: This case series further strengthens the urinary occlusion hypothesis for the causation of MGLSc. It is important to recognize that urological interventions can create incompetence of the naviculomeatal valve post voiding. In uncircumcised men, this creates a risk factor for MGLSc that was not previously present. Occlusion, the phenomenon of koebnerization and currently unelucidated epithelial susceptibility factors lead to inflammation, sclerosis and cancer. Patients and urologists should be aware of these possibilities and preventative measures instituted, e.g. adaptive voiding habits and barrier protection.

Automated blood sampling in canine telemetry studies: Enabling enhanced assessments of cardiovascular liabilities and safety margins.

INTRODUCTION: A successful integration of automated blood sampling (ABS) into the telemetry instrumented canine cardiovascular model is presented in this study. This combined model provides an efficient means to quickly gain understanding of potential effects on key cardiovascular parameters in dog while providing a complete Pharmacokinetic/Pharmacodynamic (PK/PD) profile for discovery compounds without handling artifacts, reducing the need for a separate pharmacokinetic study. METHODS: Male beagle dogs were chronically implanted with telemetry devices (PhysioTel™ model D70-PCTP) and vascular access ports (SPMID-GRIDAC-5NC). BASi Culex-L automated blood sampling (Bioanalytical Systems, Inc) system was used to collect blood samples at multiple time points. A series of four use cases utilizing four different test compounds and analytical endpoints are described to illustrate some of the potential applications of the technique. RESULTS: In the four presented use cases, automated blood sampling in telemetry instrumented dogs provides simultaneous cardiovascular (heart rate, arterial blood pressure, and left ventricular pressure), electrophysiological assessment (QTc, PR, and QRS intervals), body temperature, and animal activity, while collecting multiple blood samples for drug analysis. CONCLUSION: The combination of automated blood sampling with cardiovascular telemetry monitoring is a novel capability designed to support safety pharmacology cardiovascular assessment of discovery molecules. By combining telemetry and high-fidelity ABS, the model provides an enhanced PK/PD understanding of drug-induced hemodynamic and electrocardiographic effects of discovery compounds in conscious beagles in the same experimental session. Importantly, the model can reduce the need for a separate pharmacokinetic study (positive reduction 3R impact), reduces compound syntheses requirements, and shorten development timelines. Furthermore, implementation of this approach has also improved animal welfare by reducing the animal handling during a study, thereby reducing stress and associated data artifacts (positive refinement 3R impact).

Automated blood sampling in canine telemetry model: Enhanced assessment of immune liabilities.

INTRODUCTION: This manuscript presents a successful integration of multi-timepoint biomarker blood sampling (e.g., cytokines) in a conscious dog cardiovascular study using automated blood sampling via vascular access ports in telemetry instrumented dogs. In addition to determining plasma exposure of the test compound, the assessment of biomarkers of interest allows for more comprehensive preclinical evaluation on a traditional conscious dog cardiovascular (CV) telemetry study especially for immunology and immune-oncology molecules. This model system provides a rapid and efficient means to quickly gain understanding of potential effects on key cardiovascular parameters in large species that are commonly used for preclinical safety evaluations while collecting multiple blood samples for drug and cytokine analysis. METHODS: Male beagle dogs were chronically implanted with telemetry devices (PhysioTel™ model D70-PCTP) and vascular access ports (SPMID-GRIDAC-5NC). BASi Culex-L automated blood sampling (ABS) (Bioanalytical Systems, Inc) system was used to collect blood samples at multiple time points for cytokine analysis. Four beagles received low-dose lipopolysaccharide solution (LPS) (0.1 and 0.5 µg/mL). The following cytokines were measured by Milliplex® map Canine Cytokine Magnetic Bead Panel: Interleukin (IL) 2, IL-6, IL-7, IL-8, IL-10, IL-15, IL-18, TNF-α, MCP-1, KC-like, GM-CSF, IFN gamma, and IP10. RESULTS: Low dose LPS administration induced a pronounced dose-dependent, transient release of key inflammatory cytokines (IL-2, IL-6, IL-10, TNF-α, MCP-1, and KC-like). Cytokine responses were similar to other canine and human endotoxin models. LPS administration led to an increase in body temperature, heart rate, and mean arterial pressure, as well as a decrease in QTcV interval. CONCLUSION: Successful incorporation of cytokine analysis in telemetry instrumented dogs with vascular access ports allows for translational PK/PD modeling of both efficacy and safety of compounds in the immunology as well as the immune-oncology therapeutic areas designed to modulate the immune system. Remote collection of blood samples simultaneously with CV endpoints is a significant enhancement for assessment of biomarkers that are sensitive to animal handling and excitement associated with room disturbances which are obligatory with manual blood collection. Furthermore, implementing this approach has also refined our animal welfare procedure by reducing the handling during a study and thereby reducing stress (positive refinement 3R impact).

Automated blood sampling in canine telemetry studies: Enabling enhanced assessments of cardiovascular liabilities and safety margins.

INTRODUCTION: A successful integration of automated blood sampling (ABS) into the telemetry instrumented canine cardiovascular model is presented in this study. This combined model provides an efficient means to quickly gain understanding of potential effects on key cardiovascular parameters in dog while providing a complete Pharmacokinetic/Pharmacodynamic (PK/PD) profile for discovery compounds without handling artifacts, reducing the need for a separate pharmacokinetic study. METHODS: Male beagle dogs were chronically implanted with telemetry devices (PhysioTel™ model D70-PCTP) and vascular access ports (SPMID-GRIDAC-5NC). BASi Culex-L automated blood sampling (Bioanalytical Systems, Inc) system was used to collect blood samples at multiple time points. A series of four use cases utilizing four different test compounds and analytical endpoints are described to illustrate some of the potential applications of the technique. RESULTS: In the four presented use cases, automated blood sampling in telemetry instrumented dogs provides simultaneous cardiovascular (heart rate, arterial blood pressure, and left ventricular pressure), electrophysiological assessment (QTc, PR, and QRS intervals), body temperature, and animal activity, while collecting multiple blood samples for drug analysis. CONCLUSION: The combination of automated blood sampling with cardiovascular telemetry monitoring is a novel capability designed to support safety pharmacology cardiovascular assessment of discovery molecules. By combining telemetry and high-fidelity ABS, the model provides an enhanced PK/PD understanding of drug-induced hemodynamic and electrocardiographic effects of discovery compounds in conscious beagles in the same experimental session. Importantly, the model can reduce the need for a separate pharmacokinetic study (positive reduction 3R impact), reduces compound syntheses requirements, and shorten development timelines. Furthermore, implementation of this approach has also improved animal welfare by reducing the animal handling during a study, thereby reducing stress and associated data artifacts (positive refinement 3R impact).

Automated Blood Sampling in a Canine Telemetry Cardiovascular Model.

Successful implementation of automated blood sampling (ABS) into a telemetry instrumented canine cardiovascular model provides simultaneous cardiovascular assessment of novel compounds while collecting multiple blood samples for analysis of drug level, cytokines, and biomarkers. Purpose-bred male Beagle dogs (n = 36) were instrumented with a dual-pressure telemetry transmitter and vascular access port. Modifications to acclimation practices, surgical procedures, and housing were required for implementation of ABS in our established cardiovascular canine telemetry colony. These modifications have increased the use and reproducibility of the model by combining early pharmacokinetic and cardiovascular studies, thus achieving both refinement and reduction from a 3R perspective. In addition, the modified model can shorten timelines and reduce the compound requirement in early stages of drug development. This telemetry-ABS model provides an efficient means to quickly identify potential effects on key cardiovascular parameters in a large animal species and to obtain a more complete pharmacokinetic-pharmacodynamic profile for discovery compounds.

Time for a Fully Integrated Nonclinical-Clinical Risk Assessment to Streamline QT Prolongation Liability Determinations: A Pharma Industry Perspective.

Defining an appropriate and efficient assessment of drug-induced corrected QT interval (QTc) prolongation (a surrogate marker of torsades de pointes arrhythmia) remains a concern of drug developers and regulators worldwide. In use for over 15 years, the nonclinical International Council for Harmonization of Technical Requirements for Pharmaceuticals for Human Use (ICH) S7B and clinical ICH E14 guidances describe three core assays (S7B: in vitro hERG current & in vivo QTc studies; E14: thorough QT study) that are used to assess the potential of drugs to cause delayed ventricular repolarization. Incorporating these assays during nonclinical or human testing of novel compounds has led to a low prevalence of QTc-prolonging drugs in clinical trials and no new drugs having been removed from the marketplace due to unexpected QTc prolongation. Despite this success, nonclinical evaluations of delayed repolarization still minimally influence ICH E14-based strategies for assessing clinical QTc prolongation and defining proarrhythmic risk. In particular, the value of ICH S7B-based "double-negative" nonclinical findings (low risk for hERG block and in vivo QTc prolongation at relevant clinical exposures) is underappreciated. These nonclinical data have additional value in assessing the risk of clinical QTc prolongation when clinical evaluations are limited by heart rate changes, low drug exposures, or high-dose safety considerations. The time has come to meaningfully merge nonclinical and clinical data to enable a more comprehensive, but flexible, clinical risk assessment strategy for QTc monitoring discussed in updated ICH E14 Questions and Answers. Implementing a fully integrated nonclinical/clinical risk assessment for compounds with double-negative nonclinical findings in the context of a low prevalence of clinical QTc prolongation would relieve the burden of unnecessary clinical QTc studies and streamline drug development.

An Industry Survey With Focus on Cardiovascular Safety Pharmacology Study Design and Data Interpretation.

INTRODUCTION: The Safety Pharmacology Society (SPS) conducted a membership survey to examine industry practices related mainly to cardiovascular (CV) safety pharmacology (SP). METHODS: Questions addressed nonclinical study design, data analysis methods, drug-induced effects, and conventional and novel CV assays. RESULTS: The most frequent therapeutic area targeted by drugs developed by the companies/institutions that employ survey responders was oncology. The most frequently observed drug-mediated effects included an increased heart rate, increased arterial blood pressure, hERG (IKr) block, decreased arterial blood pressure, decreased heart rate, QTc prolongation, and changes in body temperature. Broadly implemented study practices included Latin square crossover study design with n = 4 for nonrodent CV studies, statistical analysis of data (eg, analysis of variance), use of arrhythmia detection software, and the inclusion of data from all study animals when integrating SP studies into toxicology studies. Most responders frequently used individual animal housing conditions. Responders commonly evaluated drug effects on multiple ion channels, but in silico modeling methods were used much less frequently. Most responders rarely measured the J-Tpeak interval in CV studies. Uncertainties relative to Standard for Exchange of Nonclinical Data applications for data derived from CV SP studies were common. Although available, the use of human induced pluripotent stem cell cardiomyocytes remains rare. The respiratory SP study was rarely involved with identifying drug-induced functional issues. Responders indicated that the study-derived no observed effect level was more frequently determined than the no observed adverse effect level in CV SP studies; however, a large proportion of survey responders used neither.

Increased T Cell Plasticity With Dysregulation of Follicular Helper T, Peripheral Helper T, and Treg Cell Responses in Children With Juvenile Idiopathic Arthritis and Down Syndrome-Associated Arthritis.

OBJECTIVE: Juvenile idiopathic arthritis (JIA) is the most common inflammatory arthritis in children; however, an aggressive, erosive arthritis of little-known immunologic mechanism occurs 20 times more frequently in children with Down syndrome. This study was undertaken to characterize T cell and B cell polyreactivity, follicular helper T (Tfh) cell, peripheral helper T (Tph) cell, and Treg cell responses, and synovial inflammation in Down syndrome-associated arthritis (DA). METHODS: Multiparametric flow cytometric analysis and Simplified Presentation of Incredibly Complex Evaluations (SPICE) software were used to examine peripheral blood B cell populations and T cell cytokine responses in patients with DA, JIA, Down syndrome (trisomy 21 [T21]), and in healthy controls. Tfh and Tph cell frequency and origin, in addition to Treg cell frequency, were also evaluated. Synovial inflammation was assessed by immunohistology. RESULTS: Expansion of IgM-only memory B cells was demonstrated in DA compared to JIA (mean ± SEM 22.48 ± 3.278 versus 9.011 ± 1.317; P = 0.005), paralleled by decreased frequency of transitional B cells. T cell responses in DA were characterized by marked functional plasticity, as was evident from the increased frequency of polyfunctional CD8+ Th cells (P < 0.05), CD161+ Th cells (P < 0.05), and CD8- Th cells (P < 0.001), and positivity for tumor necrosis factor, interferon-γ, interleukin-17A, or granulocyte-macrophage colony-stimulating factor, compared to all other groups. Significant expansion of CXCR3+CCR6+ (Th1/Th17) Tfh cells (P = 0.003) and CXCR3+CCR6+ Tph cells (P = 0.01), paralleled by a decrease in CXCR3-CCR6- (Th2) Tfh cells was observed in DA compared to T21. Treg cells were significantly reduced in DA compared to T21 (mean ± SEM 7.111 ± 0.9518 versus 11.96 ± 1.055 versus; P = 0.0028), with a specific reduction in the naive:memory Treg cell ratio. Marked synovial tissue inflammation and increased T cell and B cell infiltrations were demonstrated in DA compared to JIA. CONCLUSION: DA is more common and more aggressive than JIA. It is characterized by increased polyreactive Th, Tfh, and Tph cell responses, reduced Treg cell frequency, and evidence of increased synovial inflammation, all of which are potentially distinct from JIA and T21.

The application of a haemorrhage assessment tool in evaluating control of bleeding in a pilot trauma haemorrhage trial.

OBJECTIVES: To determine whether it was feasible to use a haemorrhage assessment tool (HAT) within a trauma trial and whether the data obtained could differentiate patients who had achieved haemostasis. BACKGROUND: Major haemorrhage is one of the leading causes of death worldwide, affecting 40% of trauma patients. Clinical trials evaluating haemostatic interventions often use transfusion outcomes as a primary endpoint. Transfusion is highly dependent on local practice, limiting its reliability as a robust, transferable endpoint. METHODS: A five-point HAT questionnaire was applied to participants enrolled into the EFIT-1 trial. This RCT evaluated the feasibility of administering a 6 g fibrinogen concentrate to patients with severe trauma haemorrhage. RESULTS: Of participants, 98% completed a HAT; 75% participants had 'achieved haemostasis' at the time of tool completion, as determined by clinical acumen alone. HAT scores were able to differentiate which participants required transfusion after 3 h. Of participants, 56% were transfused red blood cells when they scored 0-2, compared to 17% with HAT scores between 3 and 5. CONCLUSION: This study has confirmed the feasibility of using a HAT during the emergency care of patients suffering trauma haemorrhage, and future studies should be conducted to determine its value as an endpoint in haemostasis studies.

Autosomal dominant tubulointerstitial kidney disease (ADTKD) in Ireland.

Introduction: Autosomal dominant tubulointerstitial kidney disease (ADTKD) is a rare genetic cause of renal impairment resulting from mutations in the MUC1, UMOD, HNF1B, REN, and SEC61A1 genes. Neither the national or global prevalence of these diseases has been determined. We aimed to establish a database of patients with ADTKD in Ireland and report the clinical and genetic characteristics of these families. Methods: We identified patients via the Irish Kidney Gene Project and referral to the national renal genetics clinic in Beaumont Hospital who met the clinical criteria for ADTKD (chronic kidney disease, bland urinary sediment, and autosomal dominant inheritance). Eligible patients were then invited to undergo genetic testing by a variety of methods including panel-based testing, whole exome sequencing and, in five families who met the criteria for diagnosis of ADTKD but were negative for causal genetic mutations, we analyzed urinary cell smears for the presence of MUC1fs protein. Results: We studied 54 individuals from 16 families. We identified mutations in the MUC1 gene in three families, UMOD in five families, HNF1beta in two families, and the presence of abnormal MUC1 protein in urine smears in three families (one of which was previously known to carry the genetic mutation). We were unable to identify a mutation in 4 families (3 of whom also tested negative for urinary MUC1fs). Conclusions: There are 4443 people with ESRD in Ireland, 24 of whom are members of the cohort described herein. We observe that ADTKD represents at least 0.54% of Irish ESRD patients.

Considerations for an In Vitro, Cell-Based Testing Platform for Detection of Drug-Induced Inotropic Effects in Early Drug Development. Part 2: Designing and Fabricating Microsystems for Assaying Cardiac Contractility With Physiological Relevance Using Human iPSC-Cardiomyocytes.

Contractility of the myocardium engines the pumping function of the heart and is enabled by the collective contractile activity of its muscle cells: cardiomyocytes. The effects of drugs on the contractility of human cardiomyocytes in vitro can provide mechanistic insight that can support the prediction of clinical cardiac drug effects early in drug development. Cardiomyocytes differentiated from human-induced pluripotent stem cells have high potential for overcoming the current limitations of contractility assays because they attach easily to extracellular materials and last long in culture, while having human- and patient-specific properties. Under these conditions, contractility measurements can be non-destructive and minimally invasive, which allow assaying sub-chronic effects of drugs. For this purpose, the function of cardiomyocytes in vitro must reflect physiological settings, which is not observed in cultured cardiomyocytes derived from induced pluripotent stem cells because of the fetal-like properties of their contractile machinery. Primary cardiomyocytes or tissues of human origin fully represent physiological cellular properties, but are not easily available, do not last long in culture, and do not attach easily to force sensors or mechanical actuators. Microengineered cellular systems with a more mature contractile function have been developed in the last 5 years to overcome this limitation of stem cell-derived cardiomyocytes, while simultaneously measuring contractile endpoints with integrated force sensors/actuators and image-based techniques. Known effects of engineered microenvironments on the maturity of cardiomyocyte contractility have also been discovered in the development of these systems. Based on these discoveries, we review here design criteria of microengineered platforms of cardiomyocytes derived from pluripotent stem cells for measuring contractility with higher physiological relevance. These criteria involve the use of electromechanical, chemical and morphological cues, co-culture of different cell types, and three-dimensional cellular microenvironments. We further discuss the use and the current challenges for developing and improving these novel technologies for predicting clinical effects of drugs based on contractility measurements with cardiomyocytes differentiated from induced pluripotent stem cells. Future research should establish contexts of use in drug development for novel contractility assays with stem cell-derived cardiomyocytes.