Search

BACKGROUND: Disease-free survival (DFS) is often a primary endpoint of randomized trials of adjuvant therapies for breast cancer, but long-term follow-up of DFS and especially overall survival (OS) remain important. When the primary DFS results favor the experimental arm, patients (pts) assigned to the control group may select the option to crossover to receive the experimental treatment via protocol amendment. Such "selective crossover" disturbs the integrity of the randomized comparison for any efficacy endpoints that rely on further follow-up. Selective crossover, which is motivated by positive results having been observed in the current trial, is distinct from so-called "unplanned crossover," which refers to non-adherence to protocol. In this abstract, we discuss the consequences of selective crossover for trials evaluating adjuvant trastuzumab, using the HERA (HERceptin Adjuvant) trial as an example, and present a variety of alternative analysis approaches. METHODS: HERA enrolled 5102 women with HER2-positive early breast cancer who had completed all surgery and (neo)adjuvant chemotherapy to compare 1 or 2 years of trastuzumab treatment vs observation. After a positive first interim analysis at 1y median follow-up (MFU) showed that 1 year of trastuzumab significantly improved DFS vs observation [MJ Piccart-Gebhart et al; NEJM 2005], event-free patients in the observation group were offered crossover to receive trastuzumab. 885 (52%) of the 1698 pts in the observation group selectively crossed over to trastuzumab. RESULTS: Previously reported intention-to-treat (ITT) analysis of HERA at 4y MFU showed a decreasing effectiveness of trastuzumab with respect to DFS compared with those at 2y MFU [L Gianni et al, Lancet Oncol 2011; I Smith et al, Lancet 2007]. In addition, the ITT analysis of OS at 4y MFU showed little effect of trastuzumab, while the analysis artificially censoring follow-up in the observation group at the time of selective crossover showed a substantial OS advantage for trastuzumab. The dependent censored analysis of OS is clearly biased in favor of trastuzumab because data for pts who remain event-free can be censored at the time of crossover, while data for the sicker pts in the observation group (those who relapsed) cannot be censored due to crossover. The issues related to the ITT and dependent censored analyses will be reviewed and discussed. Alternative analytic approaches designed to estimate the treatment effect that would have been observed had there been no selective crossover will be presented. The methods include the inverse probability of censoring weighted (IPCW) approach, and randomization-based estimators under the accelerated failure time model. HERA data to about 8y MFU (available fall 2012) will be used to illustrate approaches. CONCLUSION: Alternative methods addressing selective crossover are required to estimate the trastuzumab effect for updated analyses of DFS and OS for HERA, and for any other large randomized trial with positive interim results. [ABSTRACT FROM AUTHOR]

Background: Cardiac hypertrophy is characterized by remodeling of the myocardium, which involves alterations in the ECM (extracellular matrix) and cardiomyocyte structure. These alterations critically contribute to impaired contractility and relaxation, ultimately leading to heart failure. Emerging evidence implicates that extracellular signaling molecules are critically involved in the pathogenesis of cardiac hypertrophy and remodeling. The immunophilin CyPA (cyclophilin A) has been identified as a potential culprit. In this study, we aimed to unravel the interplay between eCyPA (extracellular CyPA) and myocardial dysfunction and evaluate the therapeutic potential of inhibiting its extracellular accumulation to improve heart function., Methods: Employing a multidisciplinary approach encompassing in silico, in vitro, in vivo, and ex vivo experiments we studied a mouse model of cardiac hypertrophy and human heart specimen to decipher the interaction of CyPA and the cardiac microenvironment in highly relevant pre-/clinical settings. Myocardial expression of CyPA (immunohistology) and the inflammatory transcriptome (NanoString) was analyzed in human cardiac tissue derived from patients with nonischemic, noninflammatory congestive heart failure (n=187). These analyses were paralleled by a mouse model of Ang (angiotensin) II-induced heart failure, which was assessed by functional (echocardiography), structural (immunohistology, atomic force microscopy), and biomolecular (Raman spectroscopy) analyses. The effect of inhibiting eCyPA in the cardiac microenvironment was evaluated using a newly developed neutralizing anti-eCyPA monoclonal antibody., Results: We observed a significant accumulation of eCyPA in both human and murine-failing hearts. Importantly, higher eCyPA expression was associated with poor clinical outcomes in patients ( P =0.043) and contractile dysfunction in mice (Pearson correlation coefficient, -0.73). Further, myocardial expression of eCyPA was critically associated with an increase in myocardial hypertrophy, inflammation, fibrosis, stiffness, and cardiac dysfunction in vivo. Antibody-based inhibition of eCyPA prevented (Ang II)-induced myocardial remodeling and dysfunction in mice., Conclusions: Our study provides strong evidence of the pathogenic role of eCyPA in remodeling, myocardial stiffening, and dysfunction in heart failure. The findings suggest that antibody-based inhibition of eCyPA may offer a novel therapeutic strategy for nonischemic heart failure. Further research is needed to evaluate the translational potential of these interventions in human patients with cardiac hypertrophy., Competing Interests: None.

Aims: Cyclophilin A (CyPA) induces leucocyte recruitment and platelet activation upon release into the extracellular space. Extracellular CyPA therefore plays a critical role in immuno-inflammatory responses in tissue injury and thrombosis upon platelet activation. To date, CD147 (EMMPRIN) has been described as the primary receptor mediating extracellular effects of CyPA in platelets and leucocytes. The receptor for advanced glycation end products (RAGE) shares inflammatory and prothrombotic properties and has also been found to have similar ligands as CD147. In this study, we investigated the role of RAGE as a previously unknown interaction partner for CyPA., Methods and Results: Confocal imaging, proximity ligation, co-immunoprecipitation, and atomic force microscopy were performed and demonstrated an interaction of CyPA with RAGE on the cell surface. Static and dynamic cell adhesion and chemotaxis assays towards extracellular CyPA using human leucocytes and leucocytes from RAGE-deficient Ager-/- mice were conducted. Inhibition of RAGE abrogated CyPA-induced effects on leucocyte adhesion and chemotaxis in vitro. Accordingly, Ager-/- mice showed reduced leucocyte recruitment and endothelial adhesion towards CyPA in vivo. In wild-type mice, we observed a downregulation of RAGE on leucocytes when endogenous extracellular CyPA was reduced. We furthermore evaluated the role of RAGE for platelet activation and thrombus formation upon CyPA stimulation. CyPA-induced activation of platelets was found to be dependent on RAGE, as inhibition of RAGE, as well as platelets from Ager-/- mice showed a diminished activation and thrombus formation upon CyPA stimulation. CyPA-induced signalling through RAGE was found to involve central signalling pathways including the adaptor protein MyD88, intracellular Ca2+ signalling, and NF-κB activation., Conclusion: We propose RAGE as a hitherto unknown receptor for CyPA mediating leucocyte as well as platelet activation. The CyPA-RAGE interaction thus represents a novel mechanism in thrombo-inflammation., Competing Interests: Conflict of interest: D.H. received compensation for travel expenses and participation fees from Abbott and speaker fees from Pfizer., (© The Author(s) 2024. Published by Oxford University Press on behalf of the European Society of Cardiology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Introduction: In causal inference, the correct formulation of the scientific question of interest is a crucial step. The purpose of this study was to apply causal inference principles to external control analysis using observational data and illustrate the process to define the estimand attributes. Methods: This study compared long-term survival outcomes of a pooled set of three previously reported randomized phase 3 trials studying patients with metastatic non-small cell lung cancer receiving front-line chemotherapy and similar patients treated with front-line chemotherapy as part of routine clinical care. Causal inference frameworks were applied to define the estimand aligned with the research question and select the estimator to estimate the estimand of interest. Results: The estimand attributes of the ideal trial were defined using the estimand framework. The target trial framework was used to address specific issues in defining the estimand attributes using observational data from a nationwide electronic health record-derived de-identified database. The two frameworks combined allow to clearly define the estimand and the aligned estimator while accounting for key baseline confounders, index date, and receipt of subsequent therapies. The hazard ratio estimate (point estimate with 95% confidence interval) comparing the randomized clinical trial pooled control arm with the external control was close to 1, which is indicative of similar survival between the two arms. Discussion: The proposed combined framework provides clarity on the causal contrast of interest and the estimator to adopt, and thus facilitates design and interpretation of the analyses., Competing Interests: QL, PM, AS and OH were employed by Flatiron Health, Inc., an independent member of the Roche Group, and stock ownership in Roche. OH was employed by and owning stock in Regeneron Pharmaceuticals, Inc. LP, KR, and DH were employed by F. Hoffmann-La Roche and stock ownership in Roche. NP was employed by Genentech, Inc., and stock ownership at Roche/Genentech. The authors declare that this study received funding from Flatiron Health, Inc. The funder was involved in the study design, collection, analysis, interpretation of data, the writing of this article, and the decision to submit it for publication., (Copyright © 2024 Polito, Liang, Pal, Mpofu, Sawas, Humblet, Rufibach and Heinzmann.)

Focal pulsed field ablation (PFA) is a novel technique for treating cardiac arrhythmias. It has demonstrated positive results in initial studies and has a good safety profile. In recent studies, PFA was often utilized for first-time pulmonary vein isolation (PVI) and was performed under general anesthesia. In our study, we assessed the feasibility, safety, acute procedural efficacy, and efficiency of focal PFA under deep sedation in patients, 80% of whom had undergone at least one left atrial ablation previously. We treated 30 patients (71 ± 7, 46% male) using the CENTAURI system for various atrial arrhythmias, including atrial fibrillation, typical and atypical atrial flutter, and focal atrial tachycardia. The average procedure and fluoroscopy times were 122 ± 43 min and 9 ± 7 min, respectively. A total of 83.33% of patients received additional line ablations beyond PVI, specifically targeting the posterior box and anterior mitral line. All ablations were successfully performed in deep sedation with only one major and one minor complication observed. The major complication was a vasospasm of the right coronary artery during ablation of the cavotricuspid isthmus, which was treated successfully with intracoronary nitroglycerin. All patients could be discharged in sinus rhythm. Moreover, adenosine appears effective in identifying dormant conduction in some patients after focal PFA. In conclusion, focal PFA is an effective approach for complex left atrial ablations under deep sedation, offering both high efficacy and efficiency with a reliable safety profile. Studies on long-term outcomes are needed.

Spatial transcriptomics of histological sections have revolutionized research in life sciences and enabled unprecedented insights into genetic processes involved in tissue reorganization. However, in contrast to genomic analysis, the actual biomolecular composition of the sample has fallen behind, leaving a gap of potentially highly valuable information. Raman microspectroscopy provides untargeted spatiomolecular information at high resolution, capable of filling this gap. In this study we demonstrate spatially resolved Raman "spectromics" to reveal homogeneity, heterogeneity and dynamics of cell matrix on molecular levels by repurposing state-of-the-art bioinformatic analysis tools commonly used for transcriptomic analyses. By exploring sections of murine myocardial infarction and cardiac hypertrophy, we identify myocardial subclusters when spatially approaching the pathology, and define the surrounding metabolic and cellular (immune-) landscape. Our innovative, label-free, non-invasive "spectromics" approach could therefore open perspectives for a profound characterization of histological samples, while additionally allowing the combination with consecutive downstream analyses of the very same specimen., (© 2023. Springer Nature Limited.)

During megakaryopoiesis, megakaryocytes (MK) undergo cellular morphological changes with strong modification of membrane composition and lipid signaling. Here we adopt a lipid-centric multiomics approach to create a quantitative map of the MK lipidome during maturation and proplatelet formation. Data reveal that MK differentiation is driven by an increased fatty acyl import and de novo lipid synthesis, resulting in an anionic membrane phenotype. Pharmacological perturbation of fatty acid import and phospholipid synthesis blocked membrane remodeling and directly reduced MK polyploidization and proplatelet formation resulting in thrombocytopenia. The anionic lipid shift during megakaryopoiesis was paralleled by lipid-dependent relocalization of the scaffold protein CKIP-1 and recruitment of the kinase CK2α to the plasma membrane, which seems to be essential for sufficient platelet biogenesis. Overall, this study provides a framework to understand how the MK lipidome is altered during maturation and the impact of MK membrane lipid remodeling on MK kinase signaling involved in thrombopoiesis., Competing Interests: Competing interests The authors declare no competing financial interests.

Purpose: Pathologic complete response (pCR) has prognostic importance and is frequently used as a primary end point, but doubts remain about its validity as a surrogate for event-free survival (EFS) and overall survival (OS) in human epidermal growth factor receptor 2 (HER2)-positive, early breast cancer., Methods: We obtained individual-patient data from randomized trials of neoadjuvant anti-HER2 therapy that enrolled at least 100 patients, had data for pCR, EFS, and OS, and a median follow-up of at least 3 years. We quantified the patient-level association between pCR (defined as ypT0/Tis ypN0) and both EFS and OS using odds ratios (ORs, with ORs >1.00 indicating a benefit from achieving a pCR). We quantified the trial-level association between treatment effects on pCR and on EFS and OS using R 2 (with values above 0.75 considered as indicating strong associations)., Results: Eleven of 15 eligible trials had data for analysis (3,980 patients, with a median follow-up of 62 months). Considering all trials, we found strong patient-level associations, with ORs of 2.64 (95% CI, 2.20 to 3.07) for EFS and 3.15 (95% CI, 2.38 to 3.91) for OS; however, trial-level associations were weak, with an unadjusted R 2 of 0.23 (95% CI, 0 to 0.66) for EFS and 0.02 (95% CI, 0 to 0.17) for OS. We found qualitatively similar results when grouping trials according to different clinical questions, when analyzing only patients with hormone receptor-negative disease, and when using a more stringent definition of pCR (ypT0 ypN0)., Conclusion: Although pCR may be useful for patient management, it cannot be considered as a surrogate for EFS or OS in neoadjuvant trials of HER2-positive, operable breast cancer.

Purpose: The achievement of pathologic complete response (pCR) is strongly prognostic for event-free survival (EFS) and overall survival (OS) in patients with early breast cancer (EBC), and adapting postneoadjuvant therapy improves long-term outcomes for patients with HER2-positive disease not achieving pCR. We sought to investigate prognostic factors for EFS and OS among patients with and without pCR after neoadjuvant systemic treatment consisting of chemotherapy plus anti-HER2 therapy., Materials and Methods: We used individual data from 3,710 patients randomly assigned in 11 neoadjuvant trials for HER2-positive EBC with ≥100 patients enrolled, available data for pCR, EFS, and OS, and follow-up ≥3 years. We assessed baseline clinical tumor size (cT) and clinical nodal status (cN) as prognostic factors using stratified (by trial and treatment) Cox models separately for hormone receptor-positive versus hormone receptor-negative disease, and for patients who had pCR (pCR+; ypT0/is, ypN0) versus patients who did not achieve a pCR (pCR-)., Results: The median follow-up overall was 61.2 months. In pCR+ patients, cT and cN were significant independent prognostic factors for EFS, whereas only cT was a significant predictor for OS. In pCR- patients, cT, cN, and hormone receptor status were significant independent predictors for both EFS and OS. Regardless of hormone receptor status, cT, and cN, the 5-year EFS/OS rates were higher in pCR+ patients than in pCR- patients. In most subsets with regards to hormone receptor and pCR status, cT and cN were independent prognostic factors for both EFS and OS, including pCR+ patients., Conclusion: These results confirm that patients achieving pCR have far better survival outcomes than patients who do not. The traditional poor prognostic features, namely tumor size and nodal status, remain important even after a pCR.

Imaging of pulmonary vein (PV) anatomy by angiography before pulmonary vein isolation (PVI) for atrial fibrillation (AF) has long been standard practice in many centers. Nowadays, very accurate anatomical maps can be generated by the use of high-resolution mapping catheters, and very effective ablation lesions can be generated by the use of the high-power, short-duration (HPSD) technique. In our center, PV angiography was routinely performed before PVI. However, since there is no clear evidence for this, we refrained from performing PV angiography. This study aimed to investigate whether PV angiography is still necessary when using high-resolution mapping catheters after ablation in the high-power, short-duration (HPSD) technique. A total of 139 consecutive patients with atrial fibrillation (66.25 ± 11.68 years old, 62.39% male) undergoing radiofrequency PVI were included in the study. Ablation was performed with the HPSD technique using a fixed protocol for energy delivery of 50 watts (contact force 3-20 g). We observed no significant effect on the efficacy, efficiency and complications of the ablation procedure if pulmonary vein angiography was omitted before HPSD PVI. Thus, using our protocol, it may be useful that PV angiography is avoided, especially in young patients and those with chronic renal disease.

Background: After pulmonary vein isolation (PVI) for atrial fibrillation (AF), it is common as an endpoint to demonstrate an exit block from the pulmonary veins (PVs) in addition to an entrance block into them. By using high-resolution mapping catheters, even very small signals can be detected., Objectives: We investigated whether additional exit block testing is still necessary when using high-resolution mapping catheters after ablation in high-power short-duration (HPSD) techniques., Materials and Methods: Overall, 114 patients with AF (average age, 65.14 ± 11.3 years; 65.8% male) undergoing radiofrequency PVI were included in the study. Ablation was performed with the HPSD technique using a fixed protocol for energy delivery of 50 W (contact force 3-20 g). Entrance and exit block were tested with a high-resolution mapping catheter. Isolation of the PVs was achieved in all patients., Results: Capture of local PV tissue was demonstrated in all patients after PVI and exit block was present in all patients after entrance block was detected using a high-resolution mapping catheter., Conclusion: Exit block testing in addition to the demonstration of an entrance block as an endpoint of PVI seems to have no additional benefit and might no longer be necessary when a high-resolution mapping catheter is used in HPSD ablation for PVI of AF., (© 2022. The Author(s), under exclusive licence to Springer Medizin Verlag GmbH, ein Teil von Springer Nature.)

The standard-of-care for patients with pathological complete response (pCR) after neoadjuvant human epidermal growth factor receptor 2 (HER2)-targeted therapy plus chemotherapy is continuation of HER2-targeted therapy in the adjuvant setting. Our objective was to evaluate risk of recurrence or death in these patients and determine if outcomes differed by the HER2-targeted regimen received in each setting. We analyzed patient-level data from five randomized trials evaluating trastuzumab, pertuzumab, or both as part of systemic neoadjuvant and adjuvant therapy for HER2-positive early breast cancer, and assessed event-free survival (EFS) in 1763 patients. Patients with pCR had decreased risk of an EFS event versus those with residual disease (unadjusted hazard ratio [HR] = 0.35; 95% confidence interval [CI]: 0.27-0.46). Regardless of pCR status, after adjusting for baseline factors, reduction in EFS event risk was greater in patients administered pertuzumab/trastuzumab in both settings versus those administered only trastuzumab in both settings (HR = 0.36; 95% CI: 0.26-0.49), or pertuzumab/trastuzumab in the neoadjuvant setting and only trastuzumab in the adjuvant setting (HR = 0.67; 95% CI: 0.47-0.96). Patients with pCR had longer EFS than those with residual disease. Patients treated with pertuzumab/trastuzumab in both the neoadjuvant and adjuvant settings had the lowest risk of breast cancer recurrence.

Background and Aim: The objective of this retrospective, observational, noninterventional cohort study was to investigate prognostic factors of overall survival (OS) in patients with advanced non-small cell lung cancer (aNSCLC) and to develop a novel prognostic model., Methods: A total of 4049 patients with aNSCLC diagnosed between January 2011 and February 2020 who received atezolizumab, nivolumab, or pembrolizumab as second-line monotherapy were selected from a real-world deidentified database to build the cohort. Patients could not have received first-line treatment with clinical study drug(s) nor immune checkpoint inhibitors including anti-programmed cell death 1 (PD-1)/programmed death-ligand 1 (PD-L1), and anti-cytotoxic T-lymphocyte-associated protein 4 therapies., Results: Patients had a median age of 69 years; 45% were female, 75% White, 70% had stage IV at initial diagnosis, and 70% had nonsquamous histology. A Cox proportional hazards model with lasso regularization was used to build a prognostic model for OS using 18 baseline demographic and clinical factors based on the real-world data cohort. The risk-increasing prognostic factors were abnormally low albumin and chloride levels, Eastern Cooperative Oncology Group performance status score ≥ 2, and abnormally high levels of alkaline phosphatase and white blood cells. The risk-decreasing prognostic factors were PD-L1 positivity, longer time from advanced diagnosis to start of first-line therapy, and higher systolic blood pressure. The performance of the model was validated using data from the OAK trial, and the c-index for the OAK trial validation cohort was 0.65 and 0.67 for the real-world data cohort., Conclusions: Based on baseline demographic and clinical factors from a real-world setting, this prognostic model was developed to discriminate the risk of death in patients with aNSCLC treated with checkpoint inhibitors as second-line monotherapy, and it performed well in the real-world data and clinical trial cohorts., (© 2022 F.Hoffmann-La Roche Ltd/Genentech, Inc. Cancer Reports published by Wiley Periodicals LLC.)

Clinical decision-making tends to be based on what clinicians have been taught during undergraduate and graduate programs. The aim of the present study was to assess the clinical approach and to identify the factors that influence the decision-making for dental pulp exposure among graduate students and coordinating professors in endodontics programs offered at Brazilian universities. The study used a mail-out survey developed in the Qualtrics platform, based on seven clinical reports in which dental pulp exposure was evidenced. Descriptive statistics showing vital pulp therapy (VPT) and root canal treatment (RCT) were calculated for each clinical report. Data on the participants' (n = 113) profile and variables related to clinical and radiographic characteristics of the cases were evaluated as to their potential to affect decision-making and analyzed by logistic regression (p < 0.05). VPT was likely to be indicated in cases of patients with immature teeth (OR = 0.017; 95%CI = 0.004 -0.073). RCT indications were related to the presence of symptoms (OR = 5.326; 95%CI = 1.429-19.852) and old age (OR = 21.057; 95%CI=6.809-65.120). In pulp exposure secondary to trauma, time of pulp exposure was significantly associated with RCT indication (OR=3.267; 95%CI=1.332-8.012). The present study demonstrated that patient age, root development, and symptom features were the main factors affecting participants' decision-making.

Platelet activation plays a critical role in thrombosis. Inhibition of platelet activation is a cornerstone in treatment of acute organ ischemia. Platelet ACKR3 surface expression is independently associated with all-cause mortality in CAD patients. In a novel genetic mouse strain, we show that megakaryocyte/platelet-specific deletion of ACKR3 results in enhanced platelet activation and thrombosis in vitro and in vivo. Further, we performed ischemia/reperfusion experiments (transient LAD-ligation and tMCAO) in mice to assess the impact of genetic ACKR3 deficiency in platelets on tissue injury in ischemic myocardium and brain. Loss of platelet ACKR3 enhances tissue injury in ischemic myocardium and brain and aggravates tissue inflammation. Activation of platelet-ACKR3 via specific ACKR3 agonists inhibits platelet activation and thrombus formation and attenuates tissue injury in ischemic myocardium and brain. Here we demonstrate that ACKR3 is a critical regulator of platelet activation, thrombus formation and organ injury following ischemia/reperfusion., (© 2022. The Author(s).)

Cyclophilin A (CyPA) is widely expressed by all prokaryotic and eukaryotic cells. Upon activation, CyPA can be released into the extracellular space to engage in a variety of functions, such as interaction with the CD147 receptor, that contribute to the pathogenesis of cardiovascular diseases. CyPA was recently found to undergo acetylation at K82 and K125, two lysine residues conserved in most species, and these modifications are required for secretion of CyPA in response to cell activation in vascular smooth muscle cells. Herein we addressed whether acetylation at these sites is also required for the release of CyPA from platelets based on the potential for local delivery of CyPA that may exacerbate cardiovascular disease events. Western blot analyses confirmed the presence of CyPA in human and mouse platelets. Thrombin stimulation resulted in CyPA release from platelets; however, no acetylation was observed-neither in cell lysates nor in supernatants of both untreated and activated platelets, nor after immunoprecipitation of CyPA from platelets. Shotgun proteomics detected two CyPA peptide precursors in the recombinant protein, acetylated at K28, but again, no acetylation was found in CyPA derived from resting or stimulated platelets. Our findings suggest that acetylation of CyPA is not a major protein modification in platelets and that CyPA acetylation is not required for its secretion from platelets.

Background: Acute respiratory distress syndrome (ARDS) is considered the main cause of COVID-19 associated morbidity and mortality. Early and reliable risk stratification is of crucial clinical importance in order to identify persons at risk for developing a severe course of disease. Deceleration capacity (DC) of heart rate as a marker of cardiac autonomic function predicts outcome in persons with myocardial infarction and heart failure. We hypothesized that reduced modulation of heart rate may be helpful in identifying persons with COVID-19 at risk for developing ARDS., Methods: We prospectively enrolled 60 consecutive COVID-19 positive persons presenting at the University Hospital of Tuebingen. Arterial blood gas analysis and 24 h-Holter ECG recordings were performed and analyzed at admission. The primary end point was defined as development of ARDS with regards to the Berlin classification., Results: 61.7% (37 of 60 persons) developed an ARDS. In persons with ARDS DC was significantly reduced when compared to persons with milder course of infection (3.2 ms vs. 6.6 ms, p < 0.001). DC achieved a good discrimination performance (AUC = 0.76) for ARDS in COVID-19 persons. In a multivariate analysis, decreased DC was associated with the development of ARDS., Conclusion: Our data suggest a promising role of DC to risk stratification in COVID-19., Competing Interests: Declaration of Competing Interest Previously, we submitted an interim analysis of impaired cardiac function in COVID-19 positive persons to a different journal, which has been accepted for publication on May 28th (Rath D, Petersen-Uribe Á, Avdiu A, et al. Impaired cardiac function is associated with mortality in persons with acute COVID-19 infection. Clin Res Cardiol. 2020;1-9). However, the endpoint in this subanalysis differed and fewer persons were enrolled. Furthermore, additional biosignal analyses from 24 h ECG Holter recordings were performed in the current manuscript., (Copyright © 2021 Elsevier Inc. All rights reserved.)

COVID-19 may lead to severe acute respiratory distress syndrome (ARDS) resulting in increased morbidity and mortality. Heart failure and/or pre-existing cardiovascular disease may correlate with poor outcomes and thus require special attention from treating physicians. The present study sought to investigate a possible impact of impaired myocardial function as well as myocardial distress markers on mortality or ARDS with need for mechanical ventilation in 157 consecutive patients with confirmed SARS-CoV-2 infection. All patients were admitted and treated at the University Hospital of Tübingen, Germany, during the first wave of the pandemic. Electrocardiography, echocardiography, and routine blood sampling were performed at hospital admission. Impaired left-ventricular and right-ventricular function, tricuspid regurgitation > grade 1, and elevated RV-pressure as well as thrombotic and myocardial distress markers (D-dimers, NT-pro-BNP, and troponin-I) were associated with mechanical ventilation and/or all-cause mortality. Impaired cardiac function is more frequent amidst ARDS, leading to subsequent need for mechanical ventilation, and thus denotes a poor outcome in COVID-19. Since a causal treatment for SARS-CoV-2 infection is still lacking, guideline-compliant cardiovascular evaluation and treatment remains the best approach to improve outcomes in COVID-19 patients with cardiovascular comorbidities., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Petersen-Uribe, Avdiu, Martus, Witzel, Jaeger, Zdanyte, Heinzmann, Tavlaki, Warm, Geisler, Müller, Gawaz and Rath.)

One of the challenges in the design of confirmatory trials is to deal with uncertainties regarding the optimal target population for a novel drug. Adaptive enrichment designs (AED) which allow for a data-driven selection of one or more prespecified biomarker subpopulations at an interim analysis have been proposed in this setting but practical case studies of AEDs are still relatively rare. We present the design of an AED with a binary endpoint in the highly dynamic setting of cancer immunotherapy. The trial was initiated as a conventional trial in early triple-negative breast cancer but amended to an AED based on emerging data external to the trial suggesting that PD-L1 status could be a predictive biomarker. Operating characteristics are discussed including the concept of a minimal detectable difference, that is, the smallest observed treatment effect that would lead to a statistically significant result in at least one of the target populations at the interim or the final analysis, respectively, in the setting of AED., (© 2020 John Wiley & Sons Ltd.)

Background: COVID-19 infection may cause severe respiratory distress and is associated with increased morbidity and mortality. Impaired cardiac function and/or pre-existing cardiovascular disease may be associated with poor prognosis. In the present study, we report a comprehensive cardiovascular characterization in the first consecutive collective of patients that was admitted and treated at the University Hospital of Tübingen, Germany., Methods: 123 consecutive patients with COVID-19 were included. Routine blood sampling, transthoracic echocardiography and electrocardiography were performed at hospital admission., Results: We found that impaired left-ventricular and right-ventricular function as well as tricuspid regurgitation > grade 1 were significantly associated with higher mortality. Furthermore, elevated levels of myocardial distress markers (troponin-I and NT pro-BNP) were associated with poor prognosis in this patient collective., Conclusion: Impaired cardiac function is associated with poor prognosis in COVID-19 positive patients. Consequently, treatment of these patients should include careful guideline-conform cardiovascular evaluation and treatment. Thus, formation of a competent Cardio-COVID-19 team may represent a major clinical measure to optimize therapy of cardiovascular patients during this pandemic.

Surface receptor-mediated adhesion is a fundamental step in the recruitment of leukocytes and platelets, as well as platelet-leukocyte interactions. The surface receptor CD147 is crucially involved in host defense against self-derived and invading targets, as well as in thrombosis. In the current study, we describe the previously unknown interaction of CD147 with integrin αMβ2 (Mac-1) in this context. Using binding assays, we were able to show a stable interaction of CD147 with Mac-1 in vitro. Leukocytes from Mac-1 -/- and CD147 +/- mice showed a markedly reduced static adhesion to CD147- and Mac-1-coated surfaces, respectively, compared to wild-type mice. Similarly, we observed reduced rolling and adhesion of monocytes under flow conditions when cells were pre-treated with antibodies against Mac-1 or CD147. Additionally, as assessed by antibody inhibition experiments, CD147 mediated the dynamic adhesion of platelets to Mac-1-coated surfaces. The interaction of CD147 with Mac-1 is a previously undescribed mechanism facilitating the adhesion of leukocytes and platelets., Competing Interests: The authors declare no conflict of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results.

Impairment of renal phosphate elimination in chronic kidney disease (CKD) leads to enhanced plasma and tissue phosphate concentration, which in turn up-regulates transcription factor NFAT5 and serum & glucocorticoid-inducible kinase SGK1. The kinase upregulates ORAI1, a Ca 2+ -channel accomplishing store-operated Ca 2+ -entry (SOCE). ORAI1 is stimulated following intracellular store depletion by Ca 2+ -sensors STIM1 and/or STIM2. In megakaryocytes and blood platelets SOCE and thus ORAI1 are powerful regulators of activity. The present study explored whether the phosphate-donor ß-glycerophosphate augments NFAT5, ORAI1,2,3 and/or STIM1,2 expressions and thus SOCE in megakaryocytes. Human megakaryocytic Meg01cells were exposed to 2 mM of phosphate-donor ß-glycerophosphate for 24 hours. Platelets were isolated from blood samples of patients with impaired kidney function or control volunteers. Transcript levels were estimated utilizing q-RT-PCR, cytosolic Ca 2+ -concentration ([Ca 2+ ] i ) by Fura-2-fluorescence, and SOCE from increase of [Ca 2+ ] i following re-addition of extracellular Ca 2+ after store depletion with thapsigargin (1 µM). NFAT5 and ORAI1 protein abundance was estimated with Western blots. As a result, ß-glycerophosphate increased NFAT5, ORAI1/2/3, STIM1/2 transcript levels, as well as SOCE. Transcript levels of NFAT5, SGK1, ORAI1/2/3, and STIM1/2 as well as NFAT5 and ORAI1 protein abundance were significantly higher in platelets isolated from patients with impaired kidney function than in platelets from control volunteers. In conclusion, phosphate-donor ß-glycerophosphate triggers a signaling cascade of NFAT5/SGK1/ORAI/STIM, thus up-regulating store-operated Ca 2+ -entry.

For a trial with primary endpoint overall survival for a molecule with curative potential, statistical methods that rely on the proportional hazards assumption may underestimate the power and the time to final analysis. We show how a cure proportion model can be used to get the necessary number of events and appropriate timing via simulation. If phase 1 results for the new drug are exceptional and/or the medical need in the target population is high, a phase 3 trial might be initiated after phase 1. Building in a futility interim analysis into such a pivotal trial may mitigate the uncertainty of moving directly to phase 3. However, if cure is possible, overall survival might not be mature enough at the interim to support a futility decision. We propose to base this decision on an intermediate endpoint that is sufficiently associated with survival. Planning for such an interim can be interpreted as making a randomized phase 2 trial a part of the pivotal trial: If stopped at the interim, the trial data would be analyzed, and a decision on a subsequent phase 3 trial would be made. If the trial continues at the interim, then the phase 3 trial is already underway. To select a futility boundary, a mechanistic simulation model that connects the intermediate endpoint and survival is proposed. We illustrate how this approach was used to design a pivotal randomized trial in acute myeloid leukemia and discuss historical data that informed the simulation model and operational challenges when implementing it., (© 2019 John Wiley & Sons, Ltd.)

Background: Cyclophilin A (CyPA) is an important intracellular molecule mediating essential cellular functions such as signaling and protein folding. Enhanced CyPA platelet surface expression is associated with hypertension and hypercholesterolemia in patients with stable coronary artery disease (CAD). In patients with acute myocardial infarction CyPA platelet surface expression is significantly decreased. The aim of this study was to investigate possible associations of CyPA platelet surface expression and a clinically relevant CyPA single-nucleotide polymorphism (CyPA PPIA rs6850) with prognosis in patients with symptomatic cardiovascular disease., Materials and Methods: Blood was obtained from 335 consecutive patients with symptomatic CAD. All patients were followed up for 1080 days for endpoints all-cause death, myocardial infarction (MI), ischemic stroke, and bleeding. The primary combined endpoint was defined as a composite of all-cause death and/or MI and/or ischemic stroke. Cyclophilin A platelet surface expression levels less than or equal to the median were significantly associated with a worse prognosis (combined endpoint and all-cause death) when compared to CyPA greater than the median. Genotyping for CyPA PPIA rs6850 was performed in 752 patients with symptomatic CAD. Homozygous carriers of the minor allele showed a significantly worse cumulative event-free survival for both combined endpoint and MI when compared to carriers of the major allele., Conclusion: The CyPA platelet surface expression is associated with mortality whereas CyPA PPIA rs6850 is associated with recurrent MI in patients with symptomatic CAD. Cyclophilin A might offer a new biomarker for risk stratification and tailoring therapies in patients with cardiovascular disease., (© 2019 International Society on Thrombosis and Haemostasis.)

Background: Mitral regurgitation (MR) has a severe impact on hemodynamics and induces severe structural changes in the left atrium. Atrial remodeling is known to alter excitability and conduction in the atrium facilitating atrial fibrillation and atrial flutter. PMVR is a feasible and highly effective procedure to reduce MR in high-risk patients, which are likely to suffer from atrial rhythm disturbances. So far, electroanatomical changes after PMVR have not been studied., Hypothesis: In the current study, we investigated changes in surface electrocardiograms (ECGs) of patients undergoing PMVR for reduction of MR., Methods: We evaluated 104 surface ECGs from patients in sinus rhythm undergoing PMVR. P wave duration, P wave amplitude, PR interval, QRS duration, QRS axis, and QT interval were evaluated before and after PMVR and at follow-up., Results: We found no changes in QRS duration, QRS axis, and QT interval after successful PMVR. However, P wave duration, amplitude, and PR interval were significantly decreased after reduction of MR through PMVR (P < .05, respectively)., Conclusion: The data we provide offers insight into changes in atrial conduction after reduction of MR using PMVR in patients with sinus rhythm., (© 2019 The Authors. Clinical Cardiology published by Wiley Periodicals, Inc.)

Introduction: The role of cryoballoon (CB) pulmonary vein isolation (PVI) for patients with persistent atrial fibrillation (AF) is controversial, since long-term success can be poor. We performed left atrial voltage mapping before CB PVI and determined AF-free survival depending on the extent of low-voltage areas (LVAs)., Methods and Results: We consecutively enrolled 60 patients with persistent AF (average age, 60.6 ± 12.9 years; CHA 2 DS 2 VASc score, 2.3 ± 1.6; and left atrial size 46.0 ± 5.2 mm) who were planned for CB PVI. Before ablation, we performed left atrial voltage mapping (Abbott EnSite Precision or Velocity). LVAs were defined if local bipolar signal amplitudes were less than 0.5 mV during sinus rhythm. Thirty-seven patients did not show significant LVAs (<10%), while 12 patients had LVAs between 10% and 30% and 11 patients showed substantial LVAs greater than 30% of the left atrial area. CB PVI could be successfully performed in all patients. A 7-day holter monitoring was obtained 3, 6, and 12 months after ablation. After a 12-month follow-up time, 83.8% of patients without LVAs (<10%) were free of atrial fibrillation, while 50.0% of patients with 10% to 30% LVAs and 9.1% of patients with LVAs more than 30% had stable sinus rhythm. The degree of atrial fibrosis correlated with the risk of AF recurrence., Conclusion: In patients with persistent AF undergoing CB PVI, the extent of left atrial LVAs predicts an AF-free survival. CB PVI seems to be a highly effective treatment for patients with persistent AF without atrial fibrosis., (© 2019 Wiley Periodicals, Inc.)

Importance: Confirmation of long-term comparability between subcutaneous and intravenous trastuzumab is essential., Objective: To evaluate efficacy and safety of subcutaneous trastuzumab compared with that of intravenous trastuzumab for patients with ERBB2 (HER2)-positive early breast cancer after 6 years' follow-up in the HannaH (Enhanced Treatment With Neoadjuvant Herceptin) trial., Design, Setting, and Participants: Open-label, prospective, multicenter, international, neoadjuvant-adjuvant, randomized, phase 3 noninferiority clinical trial (primary end points: pathologic complete response and serum trough concentration predose cycle 8) conducted for 596 patients with ERBB2-positive early breast cancer enrolled from October 19, 2009, to December 1, 2010., Interventions: Eligible patients received 8 cycles of chemotherapy (4 cycles of docetaxel, 75 mg/m2, followed by 4 cycles of fluorouracil, 500 mg/m2, epirubicin, 75 mg/m2, and cyclophosphamide, 500 mg/m2) with either fixed-dose subcutaneous trastuzumab, 600 mg, or intravenous trastuzumab (loading dose, 8 mg/kg; maintenance dose, 6 mg/kg) every 3 weeks in the neoadjuvant setting. Patients received an additional 10 cycles of subcutaneous trastuzumab or intravenous trastuzumab (according to their initial randomization) after surgery in the adjuvant setting to complete 1 year of anti-ERBB2 therapy., Main Outcomes and Measures: Event-free and overall survival rates were calculated using the Kaplan-Meier method. Hazard ratios were estimated by Cox proportional hazards regression. Adverse events and serious adverse events were graded per standard criteria., Results: In total, 294 women (mean [SD] age, 50.3 [11.1] years) treated with subcutaneous trastuzumab and 297 women (mean [SD] age, 49.5 [10.8] years) treated with intravenous trastuzumab were included in respective intention-to-treat populations. Six-year event-free survival rates (65% in both study groups; hazard ratio, 0.98; 95% CI, 0.74-1.29) and overall survival rates (84% in both study groups; hazard ratio, 0.94; 95% CI, 0.61-1.45) were similar between the subcutaneous and intravenous trastuzumab groups. Patients achieving a total pathologic complete response had longer event-free survival and higher 6-year overall survival rates than those with residual disease. Incidence of adverse events (290 of 297 [97.6%] vs 282 of 298 [94.6%]), grade 3 or higher adverse events (158 of 297 [53.2%] vs 160 of 298 [53.7%]), cardiac events (44 of 297 [14.8%] vs 42 of 298 [14.1%]), and serious adverse events (65 of 297 [21.9%] vs 45 of 298 [15.1%]) was comparable between the subcutaneous and intravenous trastuzumab treatment groups., Conclusions and Relevance: This final analysis of the HannaH trial further confirms the comparable efficacy and safety of subcutaneous and intravenous trastuzumab and highlights the suitability of subcutaneous trastuzumab as an alternative route of administration for patients with ERBB2-positive early breast cancer., Trial Registration: ClinicalTrials.gov identifier: NCT00950300.

Background/aims: Tachycardiomyopathy (TCM) is a largely reversible form of non-ischemic heart failure. The underlying mechanism are, however, still today poorly understood. Recent data indicate distinct changes in mitochondrial distribution in these patients, compared to other non-ischemic cardiomyopathies.This study investigated underlying mechanisms in mitochondrial dynamics in endomyocardial biopsy samples (EMB) from patients with TCM and compared them to patients with dilated cardiomyopathy (DCM), which show similar clinical features., Methods: Focused mRNA analyses were performed on routinely obtained paraffinfixed EMB specimen from patients fulfilling TCM diagnosis criteria, as well as patients with DCM to elucidate regulatory changes in mitochondrial fusion, fission and mitophagy., Results: In patients with TCM we were able to identify mRNA of Mitofusin 1 and 2, two effector proteins regulating mitochondrial fusion, to be strongly upregulated compared to patients with DCM. Conclusively, we did not find differences in the mRNA expression of mitochondrial fission regulators including DRP1, Fis1, MFF, MiD49, and MiD51. Furthermore, we did not find significant changes in PINK1 expression, an important mediator for mitochondrial autophagy., Conclusion: The mRNA upregulation of Mitofusin 1 and 2 provides first insight into the complex changes of mitochondrial dynamics in cardiomyocytes of patients with reversible heart failure due to TCM., Competing Interests: The authors declare no conflicts of interest., (© Copyright by the Author(s). Published by Cell Physiol Biochem Press.)

Background: Atrial fibrosis is a hallmark of arrhythmogenic structural remodeling in patients with persistent atrial fibrillation (AF) and is negatively correlated with procedure outcome in patients undergoing ablation. However, noninvasive methods to determine the extent of atrial fibrosis are limited. Here, we used microRNA (miRNA) expression analysis to detect markers of left atrial low-voltage areas (LVAs) in patients with persistent AF undergoing catheter ablation., Methods: We performed 3-dimensional voltage mapping in 102 patients (average age 62.1±13.1 years, CHA 2 DS 2 -VASc score of 2.3±1.6, LA size 41.5±5.7 mm) undergoing ablation for persistent AF and determined the extent of left atrial low-voltage. LVAs were defined if bipolar electrogram amplitudes were <0.5 mV during sinus rhythm. Before ablation, we obtained a blood sample, isolated miRNAs, and profiled them on a miRCURY LNA Universal RT microRNA PCR Human panel., Results: Sixty-nine miRNAs were identified in all samples, with an average of 123 miRNAs detectable per sample. We found that the serum concentration of miR-21, a miRNA that has been previously linked to cardiac fibrosis development, was strongly associated with the extent of LVAs determined by voltage mapping. We could confirm that LVAs were negatively correlated with ablation success in a 1-year follow-up. In addition, miR-21 serum levels were associated with AF-free survival after catheter ablation., Conclusions: Circulating miR-21 correlates with left atrial LVAs and is associated with procedure outcome in patients with persistent AF undergoing ablation., (© 2018 The Authors.)

Background/aims: Fibrotic remodeling of the atria plays a key role in the pathogenesis of atrial fibrillation (AF). As little is known about the contribution of circulating monocytes in atrial remodeling and the pathophysiology of AF, we investigated profibrotic factors in different subsets of circulating monocytes obtained from patients with atrial fibrillation undergoing catheter ablation., Methods: A 3D high density voltage mapping was performed in sinus rhythm to evaluate the extent of low-voltage areas (LVAs) in the atria of 71 patients with persistent AF. Low-voltage was defined as signals of < 0.5mV during sinus rhythm. Prior to ablation, blood was drawn and monocytes were analyzed by FACS. Based on the expression of CD14 and CD16, three subgroups including CD14++ CD16- ('classical'), CD14++ CD16+ ('intermediate'), and CD14+ CD16++ ('non-classical') were analyzed for the expression of TGFb, CD147, and MMP-9, representing pivotal profibrotic pathways in myocardial remodeling., Results: Expression of TGFb was increased in CD14+ monocytes of patients with extensive LVAs compared to patients with a low extend of LVAs. While CD14++ CD16- monocytes showed no difference, CD14++ CD16+ and CD14+ CD16++ monocytes showed a strong increase of TGFb abundance. Although CD147 and MMP-9 are strongly associated with myocardial fibrosis, we found no difference in expression between the two groups in any monocyte subsets., Conclusion: TGFb is specifically upregulated on CD14++ CD16+ and CD14+ CD16++ monocytes in patients with extensive LVAs undergoing catheter ablation., (© 2018 The Author(s). Published by S. Karger AG, Basel.)

Cyclophilin A (CyPA) is involved in the pathophysiology of several inflammatory and cardiovascular diseases. To our knowledge, there is no specific inhibitor targeting extracellular CyPA without affecting other extracellular cyclophilins or intracellular CyPA functions. In this study, we developed an antibody-based inhibitor of extracellular CyPA and analysed its effects in vitro and in vivo . To generate a specific antibody, mice and rats were immunized with a peptide containing the extracellular matrix metalloproteinase inducer binding site and various antibody clones were selected and purified. At first, antibodies were tested for their binding capacity to recombinant CyPA and their functional activity. The clone 8H7-mAb was chosen for further experiments. 8H7-mAb reduced the CyPA-induced migration of inflammatory cells in vitro and in vivo . Furthermore, 8H7-mAb revealed strong antithrombotic effects by inhibiting CyPA-dependent activation of platelets and thrombus formation in vitro and in vivo . Surprisingly, 8H7-mAb did not influence in vivo tail bleeding time or in vitro whole blood coagulation parameters. Our study provides first evidence that antibody-based inhibition of extracellular CyPA inhibits thrombosis and thromboinflammation without affecting blood homeostasis. Thus, 8H7-mAb may be a promising compound for thrombi modulation in inflammatory diseases to prevent organ dysfunction., Competing Interests: Disclosure: None., (Schattauer GmbH Stuttgart.)

Vemurafenib is an orally administered small-molecule inhibitor of the oncogenic BRAF kinase that is indicated for the treatment of patients with unresectable or metastatic melanoma harbouring BRAF V600 mutations. Vemurafenib is absorbed rapidly after a single oral dose of 960 mg, reaching maximum drug concentration approximately 4 h after administration. Extensive accumulation occurs after multiple dosing at 960 mg twice daily. Steady state is achieved after approximately 15-21 days and exposure at steady state is relatively constant. Population pharmacokinetic analysis identified a vemurafenib half-life of ≈57 h and elimination appears to be predominantly via the hepatic route. Pharmacokinetic parameters are generally consistent regardless of age, sex or race. No dose adjustments are necessary for patients with mild or moderate hepatic or renal impairment, but the effects of severe hepatic or renal impairment on vemurafenib pharmacokinetics are uncertain. Vemurafenib appears to be a substrate and inducer of cytochrome P450 (CYP) 3A4, a moderate inhibitor of CYP1A2 and both a substrate and inhibitor of the drug efflux transporters P-glycoprotein and breast cancer resistance protein. The relationship between plasma vemurafenib concentrations and response remains to be clarified.

Background: Tachycardiomyopathy or tachycardia-induced cardiomyopathy (TCM) has been known for decades as a reversible form of nonischemic cardiomyopathy. However, its mechanism and properties remain poorly understood., Objectives: The current study investigated endomyocardial biopsy samples from patients with TCM and compared them with samples from patients with dilated cardiomyopathy (DCM) and inflammatory cardiomyopathy (ICM)., Methods: The study included 189 patients with new-onset heart failure and severely reduced ejection fraction not caused by valvular or ischemic heart disease. Nineteen patients retrospectively fulfilled common criteria of TCM, 79 patients had a diagnosis of DCM, and 91 had a diagnosis of ICM., Results: Patients with TCM, on the basis of clinical criteria, had stronger myocardial expression of major histocompatibility complex class II molecule and enhanced infiltration of CD68 + macrophages compared with patients with DCM. Furthermore, when compared with patients with ICM, the presence of T cells and macrophages was significantly reduced in TCM. Myocardial fibrosis was detected to a significantly lower degree in patients with TCM compared with patients with DCM and ICM. Electron microscopic examination revealed severe structural changes in patients with TCM. A disturbed distribution pattern of mitochondria was predominantly present in TCM. Quantitative assessment of myocyte morphology revealed significantly enhanced myocyte size compared with patients with ICM. Ribonucleic acid expression analysis identified changes in metabolic pathways among the patient groups., Conclusions: TCM is characterized by changes in cardiomyocyte and mitochondrial morphology accompanied by a macrophage-dominated cardiac inflammation. Thus, further prospective studies are warranted to characterize patients with TCM by endomyocardial biopsy more clearly., (Copyright © 2017 American College of Cardiology Foundation. Published by Elsevier Inc. All rights reserved.)

Background: Risk stratification of patients with non-ischemic dilated cardiomyopathy remains a matter of debate in the era of device implantation., Objective: We investigated associations between histopathological findings, contrast-enhanced cardiac MRI and the inducibility of ventricular tachycardia (VT) or fibrillation (VF) in programmed ventricular stimulation., Methods: 56 patients with impaired left ventricular ejection fraction (LVEF≤50%, mean 36.6±10.5%) due to non-ischemic dilated cardiomyopathy underwent cardiac MRI, programmed ventricular stimulation, and endomyocardial biopsy and were retrospectively investigated. Inducibility was defined as sustained mono- or polymorphic VT or unstable VT/VF requiring cardioversion/defibrillation. Primary study endpoint was defined as the occurrence of hemodynamically relevant VT/VF and/or adequate ICD-therapy during follow-up., Results: Endomyocardial biopsy detected cardiac fibrosis in 18 (32.1%) patients. Cardiac MRI revealed 35 (62.5%) patients with positive late gadolinium enhancement. VT/VF was induced in ten (17.9%) patients during programmed ventricular stimulation. Monomorphic VT was inducible in 70%, while 20% of patients showed polymorphic VT. One patient (10%) presented with VF. Inducibility correlated significantly with the presence of positive late gadolinium enhancement in cardiac MRI (p<0.01). We could not find a significant association between inducibility and the degree of cardiac inflammation and fibrosis in non-site directed routine right ventricular endomyocardial biopsy. During a mean follow-up of 2.6 years, nine (16.1%) patients reached the primary endpoint. Monomorphic VTs were found in 66.7% patients and were terminated by antitachycardia pacing therapy. One patient with polymorphic VT and two patients with VF received adequate therapy by an ICD-shock. However, inducibility did not correlate with the occurrence of endpoints., Conclusion: Inducibilty during programmed ventricular stimulation is associated with positive late gadolinium enhancement in cardiac MRI of patients with non-ischemic dilated cardiomyopathy. The presence of myocardial fibrosis or inflammation in undirected endomyocardial biopsy does not seem to be sufficient to predict future ventricular arrhythmias., Competing Interests: All authors take full responsibility for all aspects of the reliability and freedom from bias of the data presented and their discussed interpretation. The authors state that they have no conflict of interest. The authors have declared that no competing interests exist.

Background: In the phase III, open-label, randomised HannaH study, fixed-dose neoadjuvant-adjuvant subcutaneous trastuzumab for human epidermal growth factor receptor 2 (HER2)-positive early breast cancer was non-inferior to standard weight-based intravenous infusion in terms of serum trough concentration and pathological complete response (pCR). Evidence suggests that pCR, particularly total pCR (tpCR), is likely to predict clinical benefit. We report associations between tpCR and event-free survival (EFS) from HannaH (the largest population from a single study of patients presenting with newly diagnosed HER2-positive breast cancer treated with neoadjuvant-adjuvant trastuzumab to date) plus long-term efficacy and safety., Methods: Eligible patients received four cycles of neoadjuvant docetaxel followed by four cycles of fluorouracil/epirubicin/cyclophosphamide administered concurrently with 3-weekly subcutaneous (600 mg fixed dose) or intravenous trastuzumab (8 mg/kg loading, 6 mg/kg maintenance doses). Post-surgery, patients received adjuvant trastuzumab as randomised to complete 1 year of standard treatment. In exploratory analyses, we used Cox regression to assess associations between tpCR and EFS. EFS rates per subgroup were estimated using the Kaplan-Meier method., Findings: Three-year EFS rates were 76% for subcutaneous and 73% for intravenous trastuzumab (unstratified hazard ratio [HR] 0.95, 95% confidence interval [CI] 0.69-1.30; intention-to-treat population). Three-year overall survival rates were 92% for subcutaneous and 90% for intravenous trastuzumab (unstratified HR 0.76, 95% CI 0.44-1.32). tpCR was associated with a reduced risk of an EFS event: subcutaneous arm HR 0.38 (95% CI 0.22-0.65); intravenous arm HR 0.32 (95% CI 0.18-0.60). Results were similar for subgroups, including oestrogen receptor status. The few additional adverse events occurring during treatment-free follow-up were balanced between arms., Interpretation: Long-term efficacy supports the established non-inferiority of subcutaneous trastuzumab, and its safety profile remains consistent with the known intravenous profile. In each of HannaH's treatment arms, tpCR was associated with improved EFS, adding to evidence that tpCR is associated with clinical benefit in HER2-positive early breast cancer., (Copyright © 2016 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Background/aims: The polyamine oxidase inhibitor MDL-72527 (N1,N4-bis(2,3-butadienyl)-1,4-butanediamine) were expected to increase the abundance of spermine, a powerful inhibitor of platelet activation. Nothing is known, however, on the sensitivity of platelet function and survival to MDL-72527 exposure. The present study thus explored whether MDL-72527 modifies function and survival of platelets without and with platelet activation by collagen related peptide (CRP)., Methods: Platelets isolated from wild-type mice were exposed for 30 minutes to MDL-72527 (100 µM) with or without subsequent activation with CRP (2-5 µg/ml). Flow cytometry was employed to estimate cytosolic Ca2+-activity ([Ca2+]i) from Fluo-3 fluorescence, platelet degranulation from P-selectin abundance, integrin activation from αIIbβ3 integrin abundance, generation of reactive oxygen species (ROS) from DCFDA fluorescence, phospholipid scrambling of the cell membrane from annexin-V-binding, platelet volume from forward scatter and aggregation utilizing staining with CD9-APC and CD9-PE., Results: In the absence of CRP, exposure of platelets to MDL-72527 did not significantly modify [Ca2+]i, P-selectin abundance, αIIbβ3 integrin abundance, ROS, annexin-V-binding, and forward scatter. The addition of 2-5 µg/ml CRP was followed by significant increase of [Ca2+]i, P-selectin abundance, αIIbβ3 integrin activation, ROS abundance, annexin-V-binding, and aggregation as well as a significant decrease of forward scatter, all effects significantly blunted or virtually abolished in the presence of MDL-72527., Conclusions: MDL-72527 is a powerful inhibitor of platelet activation, apoptosis and aggregation., (© 2016 The Author(s) Published by S. Karger AG, Basel.)

Purpose: To characterize the population pharmacokinetics (PKs) of subcutaneous (SC) and intravenous (IV) trastuzumab in early breast cancer (EBC), assess the impact of covariates on trastuzumab PK, and evaluate fixed (nonweight-based) dosing for the SC regimen administrated via handheld syringe., Methods: Serum trastuzumab concentrations from 595 patients with HER2-positive EBC in the HannaH study (fixed 600 mg SC trastuzumab or weight-based IV trastuzumab) were analyzed using nonlinear mixed-effects modeling. Multiple logistic regression was used to assess the exposure-response relationships between PK, efficacy [pathologic complete response (pCR)], and safety [grade ≥3 adverse events (AEs)]., Results: Trastuzumab PK was described by a two-compartment model with parallel linear and nonlinear elimination and first-order SC absorption, with a bioavailability of 77 %. Estimated total clearance (CL) values were 0.18-0.22 L/day for steady-state trough/peak concentrations of 75-148 µg/mL; the estimate for central volume of distribution was 2.9 L. Body weight and alanine transaminase, while showing significant effects on PK, only explained 8% of the variability in CL. Exposure-response analyses showed no relationship between PK, pCR, and grade ≥3 AEs for either regimen., Conclusion: A fixed 600 mg SC dose of trastuzumab provides the desired exposure, with steady-state trough concentrations (35-123 μg/mL for the 5th-95th percentiles) above the historical target concentration of 20 μg/mL for efficacy. Fixed dosing is further supported by lack of an exposure-response relationship between PK, pCR, and grade ≥3 AEs. No dose adjustment per patient factors is required within the ranges studied.

Objective: Recently, we reported that extracellular cyclophilin A (CyPA) is an important agonist for platelets. Whereas soluble CyPA-levels have been associated with cardiovascular risk factors, cell-bound CyPA has not been investigated yet. In this study, we analyzed for the first time platelet-bound CyPA in patients with symptomatic coronary artery disease (CAD)., Methods and Results: blood was obtained from 388 consecutive patients: 204 with stable CAD and 184 with acute coronary syndrome (76 with unstable angina, 78 with non ST-elevation myocardial infarction (NSTEMI), and 30 with STEMI). In vitro stimulation of platelets with classical agonists revealed an enhanced expression of CyPA on the platelet surface. In patients with stable CAD, platelet-bound CyPA correlated excellently with platelet activity measured by P-selectin exposure in flow cytometry. The analysis of classical risk factors for atherosclerosis revealed that patients with hypertension and hypercholesterolemia had significantly enhanced platelet-bound CyPA, whereas diabetes and smoking were not associated with enhanced CyPA-binding to the platelet surface. In multivariate analysis, hypercholesterolemia was the only significant predictor of enhanced platelet-bound CyPA. Interestingly, in patients with acute myocardial infarction (AMI) platelet-bound CyPA was significantly decreased compared with patients with stable CAD., Conclusions: Enhanced platelet-bound CyPA is associated with hypertension and hypercholesterolemia in stable CAD patients. In patients with AMI platelet-bound CyPA is significantly decreased.

The most abundant populations of non-neoplastic cells in the glioblastoma (GBM) microenvironment are resident microglia, macrophages and infiltrating monocytes from the blood circulation. The mechanisms by which monocytes infiltrate into GBM, their fate following infiltration, and their role in GBM growth are not known. Here we tested the hypothesis that loss of the fractalkine receptor CX3CR1 in microglia and monocytes would affect gliomagenesis. Deletion of Cx3cr1 from the microenvironment resulted in increased tumor incidence and shorter survival times in glioma-bearing mice. Loss of Cx3cr1 did not affect accumulation of microglia/macrophages in peri-tumoral areas, but instead indirectly promoted the trafficking of CD11b+CD45hiCX3CR1lowLy-6ChiLy-6G-F4/80-/low circulating inflammatory monocytes into the CNS, resulting in their increased accumulation in the perivascular area. Cx3cr1-deficient microglia/macrophages and monocytes demonstrated upregulation of IL1β expression that was inversely proportional to Cx3cr1 gene dosage. The Proneural subgroup of the TCGA GBM patient dataset with high IL1β expression showed shorter survival compared to patients with low IL1β. IL1β promoted tumor growth and increased the cancer stem cell phenotype in murine and human Proneural glioma stem cells (GSCs). IL1β activated the p38 MAPK signaling pathway and expression of monocyte chemoattractant protein (MCP-1/CCL2) by tumor cells. Loss of Cx3cr1 in microglia in a monocyte-free environment had no impact on tumor growth and did not alter microglial migration. These data suggest that enhancing signaling to CX3CR1 or inhibiting IL1β signaling in intra-tumoral macrophages can be considered as potential strategies to decrease the tumor-promoting effects of monocytes in Proneural GBM.

Cyclophilins are a group of highly conserved cytosolic enzymes that have a peptidylprolyl cis/trans isomerase activity. Cyclophilin A (CyPA) can be secreted in the extracellular space by inflammatory cells and upon cell death. The presence of CyPA in patients with non-ischemic cardiomyopathy is associated with poor clinical prognosis. Here, we investigated the inhibition of extracellular CyPA in a mouse model of troponin I-induced autoimmune myocarditis using the strictly extracellular CyPA-inhibitor MM284. Since A/J mice develop severe inflammation and fibrosis after immunization with murine cardiac troponin I (mcTn I), we used this model to analyze the effects of an extracellular CyPA inhibition. As extracellular CyPA-inhibitor we used the recently described CsA-derivate MM284. In vitro studies confirmed that MM284 inhibits CyPA-induced monocytic migration and adhesion. A/J mice immunized with mcTnI were treated with MM284 or vehicle every second day. After 28 days, we found a considerable reduction of myocardial injury and fibrosis. Further analysis revealed a reduced myocardial presence of T-cells and macrophages compared to control treated animals. Whereas MMP-9 expression was reduced significantly by MM284, we observed no significant reduction of inflammatory cytokines such as IL-6 or TNFα. Extracellular CyPA plays an important role in autoimmune myocarditis for myocardial damage and fibrosis. Our data suggest a new pharmacological approach for the treatment of myocardial inflammation and reduction of cardiac fibrosis by inhibition of extracellular CyPA.