The PTPN2/PTPN1 inhibitor ABBV-CLS-484 unleashes potent anti-tumour immunity.

Immune checkpoint blockade is effective for some patients with cancer, but most are refractory to current immunotherapies and new approaches are needed to overcome resistance1,2. The protein tyrosine phosphatases PTPN2 and PTPN1 are central regulators of inflammation, and their genetic deletion in either tumour cells or immune cells promotes anti-tumour immunity3-6. However, phosphatases are challenging drug targets; in particular, the active site has been considered undruggable. Here we present the discovery and characterization of ABBV-CLS-484 (AC484), a first-in-class, orally bioavailable, potent PTPN2 and PTPN1 active-site inhibitor. AC484 treatment in vitro amplifies the response to interferon and promotes the activation and function of several immune cell subsets. In mouse models of cancer resistant to PD-1 blockade, AC484 monotherapy generates potent anti-tumour immunity. We show that AC484 inflames the tumour microenvironment and promotes natural killer cell and CD8+ T cell function by enhancing JAK-STAT signalling and reducing T cell dysfunction. Inhibitors of PTPN2 and PTPN1 offer a promising new strategy for cancer immunotherapy and are currently being evaluated in patients with advanced solid tumours (ClinicalTrials.gov identifier NCT04777994 ). More broadly, our study shows that small-molecule inhibitors of key intracellular immune regulators can achieve efficacy comparable to or exceeding that of antibody-based immune checkpoint blockade in preclinical models. Finally, to our knowledge, AC484 represents the first active-site phosphatase inhibitor to enter clinical evaluation for cancer immunotherapy and may pave the way for additional therapeutics that target this important class of enzymes.

Microcephaly gene links trithorax and REST/NRSF to control neural stem cell proliferation and differentiation.

Microcephaly is a neurodevelopmental disorder causing significantly reduced cerebral cortex size. Many known microcephaly gene products localize to centrosomes, regulating cell fate and proliferation. Here, we identify and characterize a nuclear zinc finger protein, ZNF335/NIF-1, as a causative gene for severe microcephaly, small somatic size, and neonatal death. Znf335 null mice are embryonically lethal, and conditional knockout leads to severely reduced cortical size. RNA-interference and postmortem human studies show that ZNF335 is essential for neural progenitor self-renewal, neurogenesis, and neuronal differentiation. ZNF335 is a component of a vertebrate-specific, trithorax H3K4-methylation complex, directly regulating REST/NRSF, a master regulator of neural gene expression and cell fate, as well as other essential neural-specific genes. Our results reveal ZNF335 as an essential link between H3K4 complexes and REST/NRSF and provide the first direct genetic evidence that this pathway regulates human neurogenesis and neuronal differentiation.

Milrinone as Compared with Dobutamine in the Treatment of Cardiogenic Shock.

BACKGROUND: Cardiogenic shock is associated with substantial morbidity and mortality. Although inotropic support is a mainstay of medical therapy for cardiogenic shock, little evidence exists to guide the selection of inotropic agents in clinical practice. METHODS: We randomly assigned patients with cardiogenic shock to receive milrinone or dobutamine in a double-blind fashion. The primary outcome was a composite of in-hospital death from any cause, resuscitated cardiac arrest, receipt of a cardiac transplant or mechanical circulatory support, nonfatal myocardial infarction, transient ischemic attack or stroke diagnosed by a neurologist, or initiation of renal replacement therapy. Secondary outcomes included the individual components of the primary composite outcome. RESULTS: A total of 192 participants (96 in each group) were enrolled. The treatment groups did not differ significantly with respect to the primary outcome; a primary outcome event occurred in 47 participants (49%) in the milrinone group and in 52 participants (54%) in the dobutamine group (relative risk, 0.90; 95% confidence interval [CI], 0.69 to 1.19; P = 0.47). There were also no significant differences between the groups with respect to secondary outcomes, including in-hospital death (37% and 43% of the participants, respectively; relative risk, 0.85; 95% CI, 0.60 to 1.21), resuscitated cardiac arrest (7% and 9%; hazard ratio, 0.78; 95% CI, 0.29 to 2.07), receipt of mechanical circulatory support (12% and 15%; hazard ratio, 0.78; 95% CI, 0.36 to 1.71), or initiation of renal replacement therapy (22% and 17%; hazard ratio, 1.39; 95% CI, 0.73 to 2.67). CONCLUSIONS: In patients with cardiogenic shock, no significant difference between milrinone and dobutamine was found with respect to the primary composite outcome or important secondary outcomes. (Funded by the Innovation Fund of the Alternative Funding Plan for the Academic Health Sciences Centres of Ontario; ClinicalTrials.gov number, NCT03207165.).

Central Venous Oxygen Saturation for Estimating Mixed Venous Oxygen Saturation and Cardiac Index in the ICU: A Systematic Review and Meta-Analysis.

OBJECTIVES: The objectives of our systematic review and meta-analyses were to determine the diagnostic accuracy of central venous oxygen saturation (Scv o2 ) in estimating mixed venous oxygen saturation (Sv o2 ) and cardiac index in critically ill patients. DATA SOURCES: A systematic search using MEDLINE, Cochrane Central Register of Controlled Trials, and Embase was completed on May 6, 2024. STUDY SELECTION: Studies of patients in the ICU for whom Scv o2 and at least one reference standard test was performed (thermodilution and/or Sv o2 ) were included. DATA EXTRACTION: Individual patient data were used to calculate the pooled intraclass correlation coefficient (ICC) for Sv o2 and Spearman correlation for cardiac index. The Quality Assessment of Diagnostic Accuracy Studies-2 and Grading Recommendations Assessment, Development, and Evaluation tools were used for the risk of bias and certainty of evidence assessments. DATA SYNTHESIS: Of 3427 studies, a total of 18 studies with 1971 patients were identified. We meta-analyzed 16 studies (1335 patients) that used Sv o2 as a reference and three studies (166 patients) that used thermodilution as reference. The ICC for reference Sv o2 was 0.83 (95% CI, 0.75-0.89) with a mean difference of 2.98% toward Scv o2 . The Spearman rank correlation for reference cardiac index is 0.47 (95% CI, 0.46-0.48; p < 0.0001). CONCLUSIONS: There is moderate reliability for Scv o2 in predicting Sv o2 in critical care patients with variability based on sampling site and presence of sepsis. There is limited evidence on the independent use of Scv o2 in predicting cardiac index.

Elevated and sustained expression of the transcription factors Egr1 and Egr2 controls NKT lineage differentiation in response to TCR signaling.

Interactions driven by the T cell antigen receptor (TCR) determine the lineage fate of CD4(+)CD8(+) thymocytes, but the molecular mechanisms that induce the lineage-determining transcription factors are unknown. Here we found that TCR-induced transcription factors Egr2 and Egr1 had higher and more-prolonged expression in precursors of the natural killer T (NKT) than in cells of conventional lineages. Chromatin immunoprecipitation followed by deep sequencing showed that Egr2 directly bound and activated the promoter of Zbtb16, which encodes the NKT lineage-specific transcription factor PLZF. Egr2 also bound the promoter of Il2rb, which encodes the interleukin 2 (IL-2) receptor ß-chain, and controlled the responsiveness to IL-15, which signals the terminal differentiation of the NKT lineage. Thus, we propose that persistent higher expression of Egr2 specifies the early and late stages of NKT lineage differentiation, providing a discriminating mechanism that enables TCR signaling to 'instruct' a thymic lineage.

Inotrope versus placebo therapy in cardiogenic shock: Rationale and study design of the CAPITAL DOREMI2 trial.

BACKGROUND: Cardiogenic shock (CS) is a state of end-organ hypoperfusion related to cardiac dysfunction. Current guidelines recommend consideration of inotrope therapy in patients with CS, however no robust data support their use. The purpose of the CAPITAL DOREMI2 trial is to examine the efficacy and safety of inotrope therapy against placebo in the initial resuscitation of patients with CS. METHODS AND DESIGN: This is a multi-center, double-blind, randomized, placebo-controlled trial comparing single-agent inotrope therapy to placebo in patients with CS. A total of 346 participants with Society for Cardiovascular Angiography and Interventions class C or D CS will be randomized in a 1:1 fashion to inotrope or placebo therapy, which will be administered over a 12-hour period. After this period, participants will continue open-label therapies at the discretion of the treating team. The primary outcome is a composite of all-cause in-hospital death, and, as measured during the 12-hour intervention period, any of: sustained hypotension or high dose vasopressor requirements, lactate greater than 3.5 mmol/L at 6 hours or thereafter, need for mechanical circulatory support, arrhythmia leading to emergent electrical cardioversion, and resuscitated cardiac arrest. All participants will be followed for the duration of their hospitalization, and secondary outcomes will be assessed at the time of discharge. IMPLICATION: This trial will be the first to establish the safety and efficacy of inotrope therapy against placebo in a population of patients with CS and has the potential to alter the standard care provided to this group of patients.

Efficacy and safety of corticosteroids in cardiac arrest: a systematic review, meta-analysis and trial sequential analysis of randomized control trials.

BACKGROUND: Post-cardiac arrest, outcomes for most patients are poor, regardless of setting. Many patients who do achieve spontaneous return of circulation require vasopressor therapy to maintain organ perfusion. There is some evidence to support the use of corticosteroids in cardiac arrest. RESEARCH QUESTION: Assess the efficacy and safety of corticosteroids in patients following in- and out-of-hospital cardiac arrest. STUDY DESIGN AND METHODS: We searched databases CINAHL, EMBASE, LILACS, MEDLINE, Web of Science, CENTRAL, ClinicalTrails.gov, and ICTRP. We included randomized controlled trials (RCTs) that examined the efficacy and safety of corticosteroids, as compared to placebo or usual care in patients post-cardiac arrest. We pooled estimates of effect size using random effects meta-analysis and report relative risk (RR) with 95% confidence intervals (CIs). We assessed risk of bias (ROB) for the included trials using the modified Cochrane ROB tool and rated the certainty of evidence using Grading of Recommendations Assessment, Development and Evaluation methodology. RESULTS: We included 8 RCTs (n = 2213 patients). Corticosteroids administered post-cardiac arrest had an uncertain effect on mortality measured at the longest point of follow-up (RR 0.96, 95% CI 0.90-1.02, very low certainty, required information size not met using trial sequential analysis). Corticosteroids probably increase return of spontaneous circulation (ROSC) (RR 1.32, 95% CI 1.18-1.47, moderate certainty) and may increase the likelihood of survival with good functional outcome (RR 1.49, 95% CI 0.87-2.54, low certainty). Corticosteroids may decrease the risk of ventilator associated pneumonia (RR 0.76, 95% CI 0.46-1.09, low certainty), may increase renal failure (RR 1.29, 95% CI 0.84-1.99, low certainty), and have an uncertain effect on bleeding (RR 2.04, 95% CI 0.53-7.84, very low certainty) and peritonitis (RR 10.54, 95% CI 2.99-37.19, very low certainty). CONCLUSIONS: In patients during or after cardiac arrest, corticosteroids have an uncertain effect on mortality but probably increase ROSC and may increase the likelihood of survival with good functional outcome at hospital discharge. Corticosteroids may decrease ventilator associated pneumonia, may increase renal failure, and have an uncertain effect on bleeding and peritonitis. However, the pooled evidence examining these outcomes was sparse and imprecision contributed to low or very low certainty of evidence.

Transcatheter mitral valve repair for inotrope dependent cardiogenic shock - Design and rationale of the CAPITAL MINOS trial.

BACKGROUND: Functional mitral regurgitation (MR) is an important clinical consideration in patients with heart failure. Transcatheter edge-to-edge repair (TEER) has emerged as a useful therapeutic tool for patients with chronic heart failure, however the role of TEER in patients with cardiogenic shock (CS) and MR has not yet been studied in a randomized trial. The Transcatheter Mitral Valve Repair for Inotrope Dependent Cardiogenic Shock (CAPITAL MINOS) trial was therefore designed to determine if TEER improves clinical outcomes in the CS population. METHODS AND DESIGN: The CAPITAL MINOS trial is an open-label, multi-center randomized clinical trial comparing TEER to medical therapy in patients with CS and MR. A total of 144 patients with Society for Cardiovascular Angiography and Interventions (SCAI) class C or D CS and at least 3+ MR will be randomized in a 1:1 ratio to TEER or medical therapy alone. The primary outcome will be a composite of in-hospital all-cause mortality, cardiac transplantation, implantation of durable left ventricular assist device, or discharge on palliative inotropic therapy. Patients will be followed for the duration of their index hospitalization for the primary outcome. Secondary outcomes include 6 month mortality. IMPLICATIONS: The CAPITAL MINOS trial will determine whether TEER improves outcomes in patients with CS and MR and will be an important step in optimizing treatment for this high-risk patient population.

Inotrope and vasopressor use in cardiogenic shock: what, when and why?

PURPOSE OF REVIEW: Despite increasing interest in the management of cardiogenic shock (CS), mortality rates remain unacceptably high. The mainstay of supportive treatment includes vasopressors and inotropes. These medications are recommended in international guidelines and are widely used despite limited evidence supporting safety and efficacy in CS. RECENT FINDINGS: The OptimaCC trial further supports that norepinephrine should continue to be the first-line vasopressor of choice in CS. The CAPITAL DOREMI trial found that milrinone is not superior to dobutamine in reducing morbidity and mortality in CS. Two studies currently underway will offer the first evidence of the necessity of inotrope therapy in placebo-controlled trials: CAPITAL DOREMI2 will randomize CS patients to inotrope or placebo in the initial resuscitation of shock to evaluate the efficacy of inotrope therapy and LevoHeartShock will examine the efficacy of levosimendan against placebo in early CS requiring vasopressor therapy. SUMMARY: Review of the current literature fails to show significant mortality benefit with any specific vasopressor or inotropic in CS patients. The upcoming DOREMI 2 and levosimendan versus placebo trials will further tackle the question of inotrope necessity in CS. At this time, inotrope selection should be guided by physician experience, availability, cost, and most importantly, individual patients' response to therapy.

Mechanical circulatory support in the treatment of cardiogenic shock.

PURPOSE OF REVIEW: Cardiogenic shock is a condition that is characterized by end-organ hypoperfusion secondary to reduced cardiac output, and is associated with substantial mortality. The mainstay of therapy for cardiogenic shock is reversal of the underlying cause, and concomitant supportive care with vasoactive medications (vasopressors and inotropes). Patients who continue to deteriorate despite these measures may require mechanical circulatory support (MCS). Here, we review the devices available for MCS, and their associated benefits and risks. RECENT FINDINGS: Despite growing use worldwide, there is little randomized evidence supporting the routine use of any specific device for MCS in cardiogenic shock. A large randomized trial of the intra-aortic balloon pump did not demonstrate short- or long-term improvement in mortality. The TandemHeart and Impella devices which assist in left ventricular unloading have only been evaluated in small randomized trials, which showed an increase in adverse events without improvement in mortality. Finally, venoarterial extracorporeal membrane oxygenation (provides both circulatory and respiratory support) and is currently being evaluated in large randomized clinical trials. SUMMARY: Various devices for MCS in cardiogenic shock are available, but routine use is not supported by high-quality randomized evidence. Given the resources required for initiation of MCS, use of these treatments should be limited to centers experienced in advanced cardiac care, and future research should focus on what role (if any) these devices have in clinical practice.

Inotropes, vasopressors, and mechanical circulatory support for treatment of cardiogenic shock complicating myocardial infarction: a systematic review and network meta-analysis.

PURPOSE: To compare the relative efficacy of supportive therapies (inotropes, vasopressors, and mechanical circulatory support [MCS]) for adult patients with cardiogenic shock complicating acute myocardial infarction. SOURCE: We conducted a systematic review and network meta-analysis and searched six databases from inception to December 2021 for randomized clinical trials (RCTs). We evaluated inotropes, vasopressors, and MCS in separate networks. Two reviewers performed screening, full-text review, and extraction. We used the Grading of Recommendations Assessment, Development, and Evaluation (GRADE) framework to rate the certainty in findings. The critical outcome of interest was 30-day all-cause mortality. PRINCIPAL FINDINGS: We included 17 RCTs. Among inotropes (seven RCTs, 1,145 patients), levosimendan probably reduces mortality compared with placebo (odds ratio [OR], 0.53; 95% confidence interval [CI], 0.33 to 0.87; moderate certainty), but primarily in lower severity shock. Milrinone (OR, 0.52; 95% CI, 0.19 to 1.39; low certainty) and dobutamine (OR, 0.67, 95% CI, 0.30 to 1.49; low certainty) may have no effect on mortality compared with placebo. With regard to MCS (eight RCTs, 856 patients), there may be no effect on mortality with an intra-aortic balloon pump (IABP) (OR, 0.94; 95% CI, 0.69 to 1.28; low certainty) or percutaneous MCS (pMCS) (OR, 0.96; 95% CI, 0.47 to 1.98; low certainty), compared with a strategy involving no MCS. Intra-aortic balloon pump use was associated with less major bleeding compared with pMCS. We found only two RCTs evaluating vasopressors, yielding insufficient data for meta-analysis. CONCLUSION: The results of this systematic review and network meta-analysis indicate that levosimendan reduces mortality compared with placebo among patients with low severity cardiogenic shock. Intra-aortic balloon pump and pMCS had no effect on mortality compared with a strategy of no MCS, but pMCS was associated with higher rates of major bleeding. STUDY REGISTRATION: Center for Open Science ( https://osf.io/ky2gr ); registered 10 November 2020.

RéSUMé: OBJECTIF: Comparer l'efficacité relative des thérapies de soutien (inotropes, vasopresseurs et assistance circulatoire mécanique [ACM]) chez les patients adultes atteints d'un choc cardiogénique compliquant un infarctus aigu du myocarde. SOURCES: Nous avons réalisé une revue systématique et une méta-analyse en réseau et effectué des recherches dans six bases de données depuis leur création jusqu'à décembre 2021 pour en tirer les études randomisées contrôlées (ERC). Nous avons évalué les inotropes, les vasopresseurs et les ACM dans des réseaux distincts. Deux réviseurs ont effectué la recherche, l'évaluation du texte intégral et l'extraction. Nous avons utilisé le système de notation GRADE (Grading of Recommendations Assessment, Development, and Evaluation) pour évaluer la certitude des résultats. Le critère d'évaluation d'intérêt était la mortalité toutes causes confondues à 30 jours. CONSTATATIONS PRINCIPALES: Nous avons inclus 17 ERC. Parmi les inotropes (sept ERC, 1145 patients), le lévosimendan a probablement réduit la mortalité par rapport au placebo (rapport de cotes [RC], 0,53; intervalle de confiance [IC] à 95 %, 0,33 à 0,87; certitude modérée), mais principalement en cas de choc de sévérité moindre. La milrinone (RC, 0,52; IC 95 %, 0,19 à 1,39; certitude faible) et la dobutamine (RC, 0,67, IC 95 %, 0,30 à 1,49; certitude faible) pourraient n'avoir aucun effet sur la mortalité par rapport au placebo. En ce qui concerne l'ACM (huit ERC, 856 patients), il pourrait n'y avoir aucun effet sur la mortalité avec un ballon intra-aortique (IABP) (RC, 0,94; IC 95 %, 0,69 à 1,28; certitude faible) ou un ACM percutané (ACMp) (RC, 0,96; IC 95 %, 0,47 à 1,98; certitude faible), par rapport à une stratégie sans ACM. L'utilisation d'un ballon intra-aortique était associée à moins de saignements majeurs par rapport à une ACMp. Nous n'avons trouvé que deux ERC évaluant les vasopresseurs, ce qui n'a pas fourni suffisamment de données pour la méta-analyse. CONCLUSION: Les résultats de cette revue systématique et de la méta-analyse en réseau indiquent que le lévosimendan réduit la mortalité par rapport au placebo chez les patients présentant un choc cardiogénique de faible gravité. Le ballon intra-aortique et l'ACMp n'ont eu aucun effet sur la mortalité par rapport à une stratégie sans ACM, mais l'ACMp était associée à des taux plus élevés de saignements majeurs. ENREGISTREMENT DE L'éTUDE: Center for Open Science ( https://osf.io/ky2gr ); enregistrée le 10 novembre 2020.

Effect of different maternal positions on comfort of antenatal mothers and fetal parameters during nonstress test.

Providing comfort to an antenatal mother is one of the critical components of the nonstress test (NST). An experimental study was conducted on 450 antenatal mothers beyond 34 weeks, who were randomly allocated into three groups - supine, left lateral, and semi-fowler's position, to explore the association of selected maternal positions on NST results and to find the impact of these positions on the comfort level of antenatal mothers. The fetal parameters were assessed by observing and recording biophysiological measurements from the NST and the maternal comfort of different positions by the Verbal Numerical Rating Scale. The data analyzed reveal a significant association between maternal positions and fetal parameters at P < 0.05 level. Antenatal women who were in semi-fowler's position exhibited more comfort and fetal NST reactivity as noted by significant variation in the baseline fetal heart rate, beat-to-beat variability, acceleration, deceleration, and fetal movements.

Impact of baseline beta-blocker use on inotrope response and clinical outcomes in cardiogenic shock: a subgroup analysis of the DOREMI trial.

BACKGROUND: Cardiogenic shock (CS) is associated with significant morbidity and mortality. The impact of beta-blocker (BB) use on patients who develop CS remains unknown. We sought to evaluate the clinical outcomes and hemodynamic response profiles in patients treated with BB in the 24 h prior to the development of CS. METHODS: Patients with CS enrolled in the DObutamine compaREd to MIlrinone trial were analyzed. The primary outcome was a composite of all-cause mortality, resuscitated cardiac arrest, need for cardiac transplant or mechanical circulatory support, non-fatal myocardial infarction, transient ischemic attack or stroke, or initiation of renal replacement therapy. Secondary outcomes included the individual components of the primary composite and hemodynamic response profiles derived from pulmonary artery catheters. RESULTS: Among 192 participants, 93 patients (48%) had received BB therapy. The primary outcome occurred in 47 patients (51%) in the BB group and in 52 (53%) in the no BB group (RR 0.96; 95% CI 0.73-1.27; P = 0.78) throughout the in-hospital period. There were fewer early deaths in the BB group (RR 0.41; 95% CI 0.18-0.95; P = 0.03). There were no differences in other individual components of the primary outcome or in hemodynamic response between the two groups throughout the remainder of the hospitalization. CONCLUSIONS: BB therapy in the 24 h preceding the development of CS did not negatively influence clinical outcomes or hemodynamic parameters. On the contrary, BB use was associated with fewer deaths in the early resuscitation period, suggesting a paradoxically protective effect in patients with CS. Trial registration ClinicalTrials.gov Identifier: NCT03207165.

Immunomodulatory Effects of Vitamin D Supplementation in a Deficient Population.

In addition to its canonical functions, vitamin D has been proposed to be an important mediator of the immune system. Despite ample sunshine, vitamin D deficiency is prevalent (>80%) in the Middle East, resulting in a high rate of supplementation. However, the underlying molecular mechanisms of the specific regimen prescribed and the potential factors affecting an individual's response to vitamin D supplementation are not well characterized. Our objective is to describe the changes in the blood transcriptome and explore the potential mechanisms associated with vitamin D3 supplementation in one hundred vitamin D-deficient women who were given a weekly oral dose (50,000 IU) of vitamin D3 for three months. A high-throughput targeted PCR, composed of 264 genes representing the important blood transcriptomic fingerprints of health and disease states, was performed on pre and post-supplementation blood samples to profile the molecular response to vitamin D3. We identified 54 differentially expressed genes that were strongly modulated by vitamin D3 supplementation. Network analyses showed significant changes in the immune-related pathways such as TLR4/CD14 and IFN receptors, and catabolic processes related to NF-kB, which were subsequently confirmed by gene ontology enrichment analyses. We proposed a model for vitamin D3 response based on the expression changes of molecules involved in the receptor-mediated intra-cellular signaling pathways and the ensuing predicted effects on cytokine production. Overall, vitamin D3 has a strong effect on the immune system, G-coupled protein receptor signaling, and the ubiquitin system. We highlighted the major molecular changes and biological processes induced by vitamin D3, which will help to further investigate the effectiveness of vitamin D3 supplementation among individuals in the Middle East as well as other regions.

Differential responsiveness to BRAF inhibitors of melanoma cell lines BRAF V600E-mutated.

BACKGROUND: Most mutations in melanoma affect one critical amino acid on BRAF gene, resulting in the V600E substitution. Patient management is often based on the use of specific inhibitors targeting this mutation. METHODS: DNA and RNA mutation status was assessed in 15 melanoma cell lines by Sanger sequencing and RNA-seq. We tested the cell lines responsiveness to BRAF inhibitors (vemurafenib and PLX4720, BRAF-specific and sorafenib, BRAF non-specific). Cell proliferation was assessed by MTT colorimetric assay. BRAF V600E RNA expression was assessed by qPCR. Expression level of phosphorylated-ERK protein was assessed by Western Blotting as marker of BRAF activation. RESULTS: Three cell lines were discordant in the mutation detection (BRAF V600E at DNA level/Sanger sequencing and BRAF WT on RNA-seq). We initially postulated that those cell lines may express only the WT allele at the RNA level although mutated at the DNA level. A more careful analysis showed that they express low level of BRAF RNA and the expression may be in favor of the WT allele. We tested whether the discordant cell lines responded differently to BRAF-specific inhibitors. Their proliferation rate decreased after treatment with vemurafenib and PLX4720 but was not affected by sorafenib, suggesting a BRAF V600E biological behavior. Yet, responsiveness to the BRAF specific inhibitors was lower as compared to the control. Western Blot analysis revealed a decreased expression of p-ERK protein in the BRAF V600E control cell line and in the discordant cell lines upon treatment with BRAF-specific inhibitors. The discordant cell lines showed a lower responsiveness to BRAF inhibitors when compared to the BRAF V600E control cell line. The results obtained from the inhibition experiment and molecular analyses were also confirmed in three additional cell lines. CONCLUSION: Cell lines carrying V600E mutation at the DNA level may respond differently to BRAF targeted treatment potentially due to a lower V600E RNA expression.

New-onset atrial fibrillation and associated outcomes and resource use among critically ill adults-a multicenter retrospective cohort study.

BACKGROUND: New-onset atrial fibrillation (NOAF) is commonly encountered in critically ill adults. Evidence evaluating the association between NOAF and patient-important outcomes in this population is conflicting. Furthermore, little is known regarding the association between NOAF and resource use or hospital costs. METHODS: Retrospective analysis (2011-2016) of a prospectively collected registry from two Canadian hospitals of consecutive ICU patients aged ≥ 18 years. We excluded patients with a known history of AF prior to hospital admission. Any occurrence of atrial fibrillation (AF) was prospectively recorded by bedside nurses. The primary outcome was hospital mortality, and we used multivariable logistic regression to adjust for confounders. We used a generalized linear model to evaluate contributors to total cost. RESULTS: We included 15,014 patients, and 1541 (10.3%) had NOAF during their ICU admission. While NOAF was not associated with increased odds of hospital death among the entire cohort (adjusted odds ratio [aOR] 1.02 [95% confidence interval [CI] 0.97-1.08]), an interaction was noted between NOAF and sepsis, and the presence of both was associated with higher odds of hospital mortality (aOR 1.28 [95% CI 1.09-1.36]) than either alone. Patients with NOAF had higher total costs (cost ratio [CR] 1.09 [95% CI 1.02-1.20]). Among patients with NOAF, treatment with a rhythm-control strategy was associated with higher costs (CR 1.24 [95% CI 1.07-1.40]). CONCLUSIONS: While NOAF was not associated with death or requiring discharge to long-term care among critically ill patients, it was associated with increased length of stay in ICU and increased total costs.

Long-term mortality and costs following use of Impella® for mechanical circulatory support: a population-based cohort study.

PURPOSE: The Impella® device is a form of mechanical circulatory support (MCS) used in critically ill adults with cardiogenic shock. We sought to evaluate short- and long-term outcomes following the use of Impella, including mortality, healthcare utilization, and costs. METHODS: Population-based, retrospective cohort study of adult patients (≥ 16 yr) receiving Impella in Ontario, Canada (1 April 2012-31March 2019). We captured outcomes through linkage to health administrative databases. The primary outcome was mortality during hospitalization. Secondary outcomes included mortality at 30 days, 90 days, and one year following Impella insertion. We analyzed health system costs in Canadian dollars in the year following the date of the index admission, including the costs of inpatient admission. RESULTS: We included 162 patients. Mean (standard deviation) age was 59.2 (14.5) yr, and 73.5% of patients were male. Median [interquartile range (IQR)] time to Impella insertion from date of hospital admission was 2 [1-9] days. In-hospital mortality was 56.8%, and a significant proportion of patients were bridged to a ventricular assist device (67.9%). Mortality at one year was 61.7%. Among hospital survivors, only 38.6% were discharged home independently. Median [IQR] total cost in the year following admission among all patients was $88,397 [32,718-225,628], of which $66,529 [22,789-183,165] was attributed to inpatient care. CONCLUSIONS: In-hospital mortality among patients with cardiogenic shock receiving Impella is high, but with minimal increase at one year. While Impella patients accrued substantial costs, these largely reflected inpatient costs, and not costs incurred following hospital discharge.

RéSUMé: OBJECTIF: Le dispositif Impella® est un système d'assistance circulatoire mécanique utilisé chez les adultes en état critique souffrant d'un choc cardiogénique. Nous avons souhaité évaluer les issues à court et à long terme liées à l'utilisation de l'Impella, notamment la mortalité, l'utilisation des soins de santé et les coûts. MéTHODE: Nous avons réalisé une étude de cohorte rétrospective basée sur une population de patients adultes (≥ 16 ans) recevant un Impella en Ontario, au Canada (1er avril 2012­31 mars 2019). Nous avons saisi les données pertinentes à nos critères d'évaluation en créant des liens avec les bases de données administratives de santé. Le critère d'évaluation principal était la mortalité pendant l'hospitalisation. Les critères d'évaluation secondaires comportaient la mortalité à 30 jours, 90 jours et un an après l'insertion de l'Impella. Nous avons analysé les coûts pour le système de santé en dollars canadiens au cours de l'année suivant la date d'admission initiale, notamment les coûts hospitaliers. RéSULTATS: Nous avons inclus 162 patients. L'âge moyen (écart type) était de 59,2 (14,5) ans, et 73,5 % des patients étaient des hommes. Le temps médian [écart interquartile (ÉIQ)] jusqu'à insertion de l'Impella depuis l'admission à l'hôpital était de 2 [1­9] jours. La mortalité hospitalière était de 56,8 %, et un dispositif d'assistance ventriculaire a été implanté chez une importante proportion de patients (67,9 %). La mortalité à un an était de 61,7 %. Parmi les survivants hospitaliers, seuls 38,6 % ont reçu leur congé pour un retour indépendant à la maison. Le coût total médian [ÉIQ] pour l'année suivant l'admission parmi tous les patients était de 88 397 $ [32 718 ­ 225 628 $], dont 66 259 $ [22 789 ­ 183 165 $] étaient attribuables aux soins hospitaliers. CONCLUSION: La mortalité hospitalière parmi les patients en choc cardiogénique chez lesquels un Impella est installé est élevée, mais on n'observe qu'une augmentation minimale à un an. Alors que les patients ayant reçu un Impella ont engendré des coûts importants, ces coûts reflètent principalement les coûts hospitaliers et non les coûts encourus après le congé de l'hôpital.

Monoallelic expression in melanoma.

BACKGROUND: Monoallelic expression (MAE) is a frequent genomic phenomenon in normal tissues, however its role in cancer is yet to be fully understood. MAE is defined as the expression of a gene that is restricted to one allele in the presence of a diploid heterozygous genome. Constitutive MAE occurs for imprinted genes, odorant receptors and random X inactivation. Several studies in normal tissues have showed MAE in approximately 5-20% of the cases. However, little information exists on the MAE rate in cancer. In this study we assessed the presence and rate of MAE in melanoma. The genetic basis of melanoma has been studied in depth over the past decades, leading to the identification of mutations/genetic alterations responsible for melanoma development. METHODS: To examine the role of MAE in melanoma we used 15 melanoma cell lines and compared their RNA-seq data with genotyping data obtained by the parental TIL (tumor infiltrating lymphocytes). Genotyping was performed using the Illumina HumanOmni1 beadchip. The RNA-seq library preparation and sequencing was performed using the Illumina TruSeq Stranded Total RNA Human Kit and subsequently sequenced using a HiSeq 2500 according to manufacturer's guidelines. By comparing genotyping data with RNA-seq data, we identified SNPs in which DNA genotypes were heterozygous and corresponding RNA genotypes were homozygous. All homozygous DNA genotypes were removed prior to the analysis. To confirm the validity to detect MAE, we examined heterozygous DNA genotypes from X chromosome of female samples as well as for imprinted and olfactory receptor genes and confirmed MAE. RESULTS: MAE was detected in all 15 cell lines although to a different rate. When looking at the B-allele frequencies we found a preferential pattern of complete monoallelic expression rather then differential monoallelic expression across the 15 melanoma cell lines. As some samples showed high differences in the homozygous and heterozygous call rate, we looked at the single chromosomes and showed that MAE may be explained by underlying large copy number imbalances in some instances. Interestingly these regions included genes known to play a role in melanoma initiation and progression. Nevertheless, some chromosome regions showed MAE without CN imbalances suggesting that additional mechanisms (including epigenetic silencing) may explain MAE in melanoma. CONCLUSION: The biological implications of MAE are yet to be realized. Nevertheless, our findings suggest that MAE is a common phenomenon in melanoma cell lines. Further analyses are currently being undertaken to evaluate whether MAE is gene/pathway specific and to understand whether MAE can be employed by cancers to achieve a more aggressive phenotype.