The Conserved YPX3L Motif in the BK Polyomavirus VP1 Protein Is Important for Viral Particle Assembly but Not for Its Secretion into Extracellular Vesicles.

The BK polyomavirus (BKPyV) is a small DNA non-enveloped virus whose infection is asymptomatic in most of the world's adult population. However, in cases of immunosuppression, the reactivation of the virus can cause various complications, and in particular, nephropathies in kidney transplant recipients or hemorrhagic cystitis in bone marrow transplant recipients. Recently, it was demonstrated that BKPyV virions can use extracellular vesicles to collectively traffic in and out of cells, thus exiting producing cells without cell lysis and entering target cells by diversified entry routes. By a comparison to other naked viruses, we investigated the possibility that BKPyV virions recruit the Endosomal-Sorting Complexes Required for Transport (ESCRT) machinery through late domains in order to hijack extracellular vesicles. We identified a single potential late domain in the BKPyV structural proteins, a YPX3L motif in the VP1 protein, and used pseudovirions to study the effect of point mutations found in a BKPyV clinical isolate or known to ablate the interaction of such a domain with the ESCRT machinery. Our results suggest that this domain is not involved in BKPyV association with extracellular vesicles but is crucial for capsomere interaction and thus viral particle assembly.

Shortcomings of services for persons with severe and persistent mental health challenges: a qualitative study of service users and family carers.

Introduction: The opinions of service users and carers are crucial to identifying ways to innovate and implement system change. This study aims to explore the views and experiences of service users and carerson the services they have used for their mental health challenges and their suggestions for service reform. Methods: Twenty participants (15 carers and 5 service users) were interviewed for the study. Results: Eight categories emerged from the data. They were: Several gaps in the system, Barriers to accessing services, Services are not fit for purpose, Services operate in isolation, System is not person focused, Service users and carers are treated poorly, Services are overloaded and under resourced and Recommendations for service reform. Respondents reported that a persistent lack of funding and resources for mental health services was a main cause of these shortcomings. Respondents also noted that innovations were needed to re-orient services to enable continuity of care, and training of mental health professionals was needed for a better understanding of the needs of service users and their carers. Discussion: Additional research is needed with larger and more diverse samples to further explore these findings.

The economic burden experienced by carers of children who had a critical deterioration at a tertiary children's hospital in the United Kingdom (the DETECT study): an online survey.

BACKGROUND: Unplanned critical care admissions following in-hospital deterioration in children are expected to impose a significant burden for carers across a number of dimensions. One dimension relates to the financial and economic impact associated with the admission, from both direct out-of-pocket expenditures, as well as indirect costs, reflecting productivity losses. A robust assessment of these costs is key to understand the wider impact of interventions aiming to reduce in-patient deterioration. This work aims to determine the economic burden imposed on carers caring for hospitalised children that experience critical deterioration events. METHODS: Descriptive study with quantitative approach. Carers responded to an online survey between July 2020 and April 2021. The survey was developed by the research team and piloted before use. The sample comprised 71 carers of children admitted to a critical care unit following in-patient deterioration, at a tertiary children's hospital in the UK. The survey provides a characterisation of the carer's household and estimates of direct non-medical costs grouped in five different expenditure categories. Productivity losses can also be estimated based on the reported information. RESULTS: Most carers reported expenditures associated to the child's admission in the week preceding the survey completion. Two-thirds of working carers had missed at least one workday in the week prior to the survey completion. Moreover, eight in ten carers reported having had to travel from home to the hospital at least once a week. These expenditures, on average, amount to £164 per week, grouped in five categories (38% each to travelling costs and to food and drink costs, with accommodation, childcare, and parking representing 12%, 7% and 5%, respectively). Additionally, weekly productivity losses for working carers are estimated at £195. CONCLUSION: Unplanned critical care admissions for children impose a substantial financial burden for carers. Moreover, productivity losses imply a subsequent cost to society. Even though subsidised hospital parking and on-site accommodation at the hospital contribute to minimising such expenditure, the overall impact for carers remains high. Interventions aiming at reducing emergency critical care admissions, or their length, can be crucial to further contribute to the reduction of this burden. TRIAL REGISTRATION: Current Controlled Trials ISRCTN61279068, date of registration 07/06/2019, retrospectively registered.

The Digital Bytes Project: Digital Storytelling as a Tool for Challenging Stigma and Making Connections in a Forensic Mental Health Setting.

This article reports on the findings of a study that explores the utility of digital storytelling as a narrative practice and learning tool for social work in an Australian secure forensic mental health hospital. The short digital stories, or Digital Bytes Project, centered on capturing the lived experience, hopes and perspectives of the hospital's service users by giving voice to their experiences through digital technology. The project was collaboratively designed and co-delivered with social work students, hospital staff, and service users. It aimed to not only destigmatize people with lived experiences of mental distress and criminal justice system involvement but also to give staff and students further insights into understanding who they are working with. Through a series of 11 semi-structured, one on one interviews, this research aims to explore social work student and forensic mental health staff experiences and perceptions in relation to the utility and impact of these digital bytes, reflecting on how the prototype bytes may have impacted their learnings, or practice, including how they then interact with service users. This research investigates how these digital bytes could be used further within forensic mental health organisations and contexts. The research findings demonstrate the overall value of digital bytes in challenging different kinds of stigma, shifting power dynamics and staff perspectives; strengthening rapport and understanding through enhancing engagement and sharing power between students, staff, and consumers; as well as providing insight into the utility of digital bytes for learning and making connections between theory and practice. The preliminary findings from this research suggest the need for greater accessibility, integration, and consideration of these digital tools, with their potential to be translated across multiple human service sectors.

Using technology to reduce critical deterioration (the DETECT study): a cost analysis of care costs at a tertiary children's hospital in the United Kingdom.

BACKGROUND: Electronic early warning systems have been used in adults for many years to prevent critical deterioration events (CDEs). However, implementation of similar technologies for monitoring children across the entire hospital poses additional challenges. While the concept of such technologies is promising, their cost-effectiveness is not established for use in children. In this study we investigate the potential for direct cost savings arising from the implementation of the DETECT surveillance system. METHODS: Data were collected at a tertiary children's hospital in the United Kingdom. We rely on the comparison between patients in the baseline period (March 2018 to February 2019) and patients in the post-intervention period (March 2020 to July 2021). These provided a matched cohort of 19,562 hospital admissions for each group. From these admissions, 324 and 286 CDEs were observed in the baseline and post-intervention period, respectively. Hospital reported costs and Health Related Group (HRG) National Costs were used to estimate overall expenditure associated with CDEs for both groups of patients. RESULTS: Comparing post-intervention with baseline data we found a reduction in the total number of critical care days, driven by an overall reduction in the number of CDEs, however without statistical significance. Using hospital reported costs adjusted for the Covid-19 impact, we estimate a non-significant reduction of total expenditure from £16.0 million to £14.3 million (corresponding to £1.7 million of savings - 11%). Additionally, using HRG average costs, we estimated a non-significant reduction of total expenditure from £8.2 million to £ 7.2 million (corresponding to £1.1 million of savings - 13%). DISCUSSION AND CONCLUSION: Unplanned critical care admissions for children not only impose a substantial burden on patients and families but are also costly for hospitals. Interventions aimed at reducing emergency critical care admissions can be crucial to contribute to the reduction of these episodes' costs. Even though cost reductions were identified in our sample, our results do not support the hypothesis that reducing CDEs using technology leads to a significant reduction on hospital costs. TRIAL REGISTRATION: Current Controlled Trials ISRCTN61279068, date of registration 07/06/2019, retrospectively registered.

Molecular targeting of the UDP-glucuronosyltransferase enzymes in high-eukaryotic translation initiation factor 4E refractory/relapsed acute myeloid leukemia patients: a randomized phase II trial of vismodegib, ribavirin with or without decitabine.

Drug resistance underpins poor outcomes in many malignancies including refractory and relapsed acute myeloid leukemia (R/R AML). Glucuronidation is a common mechanism of drug inactivation impacting many AML therapies, e.g., cytarabine, decitabine, azacytidine and venetoclax. In AML cells, the capacity for glucuronidation arises from increased production of the UDP-glucuronosyltransferase 1A (UGT1A) enzymes. UGT1A elevation was first observed in AML patients who relapsed after response to ribavirin, a drug used to target the eukaryotic translation initiation factor eIF4E, and subsequently in patients who relapsed on cytarabine. UGT1A elevation resulted from increased expression of the sonic-hedgehog transcription factor GLI1. Vismodegib inhibited GLI1, decreased UGT1A levels, reduced glucuronidation of ribavirin and cytarabine, and re-sensitized cells to these drugs. Here, we examined if UGT1A protein levels, and thus glucuronidation activity, were targetable in humans and if this corresponded to clinical response. We conducted a phase II trial using vismodegib with ribavirin, with or without decitabine, in largely heavily pre-treated patients with high-eIF4E AML. Pre-therapy molecular assessment of patients' blasts indicated highly elevated UGT1A levels relative to healthy volunteers. Among patients with partial response, blast response or prolonged stable disease, vismodegib reduced UGT1A levels, which corresponded to effective targeting of eIF4E by ribavirin. In all, our studies are the first to demonstrate that UGT1A protein, and thus glucuronidation, are targetable in humans. These studies pave the way for the development of therapies that impair glucuronidation, one of the most common drug deactivation modalities. Clinicaltrials.gov: NCT02073838.

Parents' experiences and perceptions of the acceptability of a whole-hospital, pro-active electronic pediatric early warning system (the DETECT study): A qualitative interview study.

Background: Failure to recognize and respond to clinical deterioration in a timely and effective manner is an urgent safety concern, driving the need for early identification systems to be embedded in the care of children in hospital. Pediatric early warning systems (PEWS) or PEW scores alert health professionals (HPs) to signs of deterioration, trigger a review and escalate care as needed. PEW scoring allows HPs to record a child's vital signs and other key data including parent concern. Aim: This study aimed to explore the experiences and perceptions of parents about the acceptability of a newly implemented electronic surveillance system (the DETECT surveillance system), and factors that influenced acceptability and their awareness around signs of clinical deterioration and raising concern. Methods: Descriptive, qualitative semi-structured telephone interviews were undertaken with parents of children who had experienced a critical deterioration event (CDE) (n = 19) and parents of those who had not experienced a CDE (non-CDE parents) (n = 17). Data were collected between February 2020 and February 2021. Results: Qualitative data were analyzed using generic thematic analysis. Analysis revealed an overarching theme of trust as a key factor that underpinned all aspects of children's vital signs being recorded and monitored. The main themes reflect three domains of parents' trust: trust in themselves, trust in the HPs, and trust in the technology. Conclusion: Parents' experiences and perceptions of the acceptability of a whole-hospital, pro-active electronic pediatric early warning system (The DETECT system) were positive; they found it acceptable and welcomed the use of new technology to support the care of their child.

Clinical utility and acceptability of a whole-hospital, pro-active electronic paediatric early warning system (the DETECT study): A prospective e-survey of parents and health professionals.

BACKGROUND: Paediatric early warning systems (PEWS) are a means of tracking physiological state and alerting healthcare professionals about signs of deterioration, triggering a clinical review and/or escalation of care of children. A proactive end-to-end deterioration solution (the DETECT surveillance system) with an embedded e-PEWS that included sepsis screening was introduced across a tertiary children's hospital. One component of the implementation programme was a sub-study to determine an understanding of the DETECT e-PEWS in terms of its clinical utility and its acceptability. AIM: This study aimed to examine how parents and health professionals view and engage with the DETECT e-PEWS apps, with a particular focus on its clinical utility and its acceptability. METHOD: A prospective, closed (tick box or sliding scale) and open (text based) question, e-survey of parents (n = 137) and health professionals (n = 151) with experience of DETECT e-PEWS. Data were collected between February 2020 and February 2021. RESULTS: Quantitative data were analysed using descriptive and inferential statistics and qualitative data with generic thematic analysis. Overall, both clinical utility and acceptability (across seven constructs) were high across both stakeholder groups although some challenges to utility (e.g., sensitivity of triggers within specific patient populations) and acceptability (e.g., burden related to having to carry extra technology) were identified. CONCLUSION: Despite the multifaceted nature of the intervention and the complexity of implementation across a hospital, the system demonstrated clinical utility and acceptability across two key groups of stakeholders: parents and health professionals.

Health professionals' initial experiences and perceptions of the acceptability of a whole-hospital, pro-active electronic paediatric early warning system (the DETECT study): a qualitative interview study.

BACKGROUND: Paediatric early warning systems (PEWS) alert health professionals to signs of a child's deterioration with the intention of triggering an urgent review and escalating care. They can reduce unplanned critical care transfer, cardiac arrest, and death. Electronic systems may be superior to paper-based systems. The objective of the study was to critically explore the initial experiences and perceptions of health professionals about the acceptability of DETECT e-PEWS, and what factors influence its acceptability. METHODS: A descriptive qualitative study (part of The DETECT study) was undertaken February 2020-2021. Single, semi-structured telephone interviews were used. The setting was a tertiary children's hospital, UK. The participants were health professionals working in study setting and using DETECT e-PEWS. Sampling was undertaken using a mix of convenience and snowballing techniques. Participants represented two user-groups: 'documenting vital signs' (D-VS) and 'responding to vital signs' (R-VS). Perceptions of clinical utility and acceptability of DETECT e-PEWS were derived from thematic analysis of transcripts. RESULTS: Fourteen HPs (12 nurses, 2 doctors) participated; seven in D-VS and seven in the R-VS group. Three main themes were identified: complying with DETECT e-PEWS, circumventing DETECT e-PEWS, and disregarding DETECT e-PEWS. Overall clinical utility and acceptability were deemed good for HPs in the D-VS group but there was diversity in perception in the R-VS group (nurses found it more acceptable than doctors). Compliance was better in the D-VS group where use of DETECT e-PEWS was mandated and used more consistently. Some health professionals circumvented DETECT e-PEWS and fell back into old habits. Doctors (R-VS) did not consistently engage with DETECT e-PEWS, which reduced the acceptability of the system, even in those who thought the system brought benefits. CONCLUSIONS: Speed and accuracy of real-time data, automation of triggering alerts and improved situational awareness were key factors that contributed to the acceptability of DETECT e-PEWS. Mandating use of both recording and responding aspects of DETECT e-PEWS is needed to ensure full implementation.

The psychosocial complexities of acute burn patients in an Australian trauma hospital.

INTRODUCTION: Psychosocial risk and protective factors specific to acute burn patients have been shown to impact on longer-term health outcomes. Yet the nature and impact of such factors throughout the acute hospital admission phase have not been extensively examined to date. This study analysed the psychosocial pre-admission, hospitalisation and discharge factors for a sample of burn patients at an Australian specialist acute burns unit (The Alfred) and the relationship with their admission length. METHODS: A retrospective audit was undertaken of psychosocial factors documented in patient medical records, Victorian Adult Burns Service registry data, and social work files (January-December 2014). RESULTS: Two hundred and forty-nine patients were identified, with a mean age of 42.83 years and a preponderance of men (73.9%). The majority (71%) had a low burn severity (<10%), and an average of eight psychosocial factors. Independent of the severity of the burn, two psychosocial domains were strongly predictive of length of stay - coping with treatment and discharge planning - along with pre-existing psychosocial factors and family support. CONCLUSION: A diversity of psychosocial factors was identified, highlighting the clinical complexity of this patient group. Given three domains of admission-related psychosocial factors were predictive of the length of patient stay, more targeted psychosocial interventions in these areas may ensure patients and their families are supported more effectively throughout an admission, and length of stay costs may also be reduced.

An Immunodominant and Conserved B-Cell Epitope in the Envelope of Simian Foamy Virus Recognized by Humans Infected with Zoonotic Strains from Apes.

Cross-species transmission of simian foamy viruses (SFVs) from nonhuman primates (NHPs) to humans is currently ongoing. These zoonotic retroviruses establish lifelong persistent infection in their human hosts. SFV are apparently nonpathogenic in vivo, with ubiquitous in vitro tropism. Here, we aimed to identify envelope B-cell epitopes that are recognized following a zoonotic SFV infection. We screened a library of 169 peptides covering the external portion of the envelope from the prototype foamy virus (SFVpsc_huHSRV.13) for recognition by samples from 52 Central African hunters (16 uninfected and 36 infected with chimpanzee, gorilla, or Cercopithecus SFV). We demonstrate the specific recognition of peptide N96-V110 located in the leader peptide, gp18LP Forty-three variant peptides with truncations, alanine substitutions, or amino acid changes found in other SFV species were tested. We mapped the epitope between positions 98 and 108 and defined six amino acids essential for recognition. Most plasma samples from SFV-infected humans cross-reacted with sequences from apes and Old World monkey SFV species. The magnitude of binding to peptide N96-V110 was significantly higher for samples of individuals infected with a chimpanzee or gorilla SFV than those infected with a Cercopithecus SFV. In conclusion, we have been the first to define an immunodominant B-cell epitope recognized by humans following zoonotic SFV infection.IMPORTANCE Foamy viruses are the oldest known retroviruses and have been mostly described to be nonpathogenic in their natural animal hosts. SFVs can be transmitted to humans, in whom they establish persistent infection, like the simian lenti- and deltaviruses that led to the emergence of two major human pathogens, human immunodeficiency virus type 1 and human T-lymphotropic virus type 1. This is the first identification of an SFV-specific B-cell epitope recognized by human plasma samples. The immunodominant epitope lies in gp18LP, probably at the base of the envelope trimers. The NHP species the most genetically related to humans transmitted SFV strains that induced the strongest antibody responses. Importantly, this epitope is well conserved across SFV species that infect African and Asian NHPs.

Potent neutralizing antibodies in humans infected with zoonotic simian foamy viruses target conserved epitopes located in the dimorphic domain of the surface envelope protein.

Human diseases of zoonotic origin are a major public health problem. Simian foamy viruses (SFVs) are complex retroviruses which are currently spilling over to humans. Replication-competent SFVs persist over the lifetime of their human hosts, without spreading to secondary hosts, suggesting the presence of efficient immune control. Accordingly, we aimed to perform an in-depth characterization of neutralizing antibodies raised by humans infected with a zoonotic SFV. We quantified the neutralizing capacity of plasma samples from 58 SFV-infected hunters against primary zoonotic gorilla and chimpanzee SFV strains, and laboratory-adapted chimpanzee SFV. The genotype of the strain infecting each hunter was identified by direct sequencing of the env gene amplified from the buffy coat with genotype-specific primers. Foamy virus vector particles (FVV) enveloped by wild-type and chimeric gorilla SFV were used to map the envelope region targeted by antibodies. Here, we showed high titers of neutralizing antibodies in the plasma of most SFV-infected individuals. Neutralizing antibodies target the dimorphic portion of the envelope protein surface domain. Epitopes recognized by neutralizing antibodies have been conserved during the cospeciation of SFV with their nonhuman primate host. Greater neutralization breadth in plasma samples of SFV-infected humans was statistically associated with smaller SFV-related hematological changes. The neutralization patterns provide evidence for persistent expression of viral proteins and a high prevalence of coinfection. In conclusion, neutralizing antibodies raised against zoonotic SFV target immunodominant and conserved epitopes located in the receptor binding domain. These properties support their potential role in restricting the spread of SFV in the human population.

Pulmonary hypertension-targeted therapies in heart failure: A systematic review and meta-analysis.

BACKGROUND: Pulmonary hypertension (PH) due to left heart failure (HF) is the most common form of PH. However, treatment is unclear because there are conflicting results about safety and efficacy of PH-targeted therapies. OBJECTIVES: To assess the effects of PH-targeted therapy on exercise capacity in HF patients. METHODS: MEDLINE, EMBASE and the Cochrane Library were searched from January 1990 to July 2017 for randomized controlled trials comparing PH-targeted therapies to conventional therapy in HF. The primary outcome was to assess the effects on exercise capacity. Secondary outcomes included mortality, hospitalisation, NT-proBNP levels, echocardiographic and hemodynamics parameters and discontinuation rate. RESULTS: 22 studies were included (n = 5448), including 3, 8 and 11 studies with low, high and unknown risk of bias, respectively. PH-targeted therapies were associated with an improvement of exercise capacity (standardized mean difference 0.29;95%CI:0.08-0.50, p = 0.006). Pre-specified subgroup analyses found that this improvement was predominantly observed in studies evaluating phosphodiesterase-5 inhibitors and prostanoids and in patients with reduced ejection fraction. Moreover, systolic pulmonary artery pressure measured by echocardiography was improved (mean difference: -7.5mmHg; [95%CI]: -14.9,-0.1, p = 0.05), which was also entirely driven by studies evaluating phosphodiesterase-5 inhibitors. However, PH-targeted therapies were associated with an increased treatment discontinuation rates and a potential increase in mortality compared to standard treatment. CONCLUSIONS: In conclusion, PH-targeted therapies and especially phosphodiesterase-5 inhibitors may improve exercise capacity in patients with HF. However, an increase in adverse outcomes was likely. Moreover, most studies were at high or unknown risk of bias, precluding confident conclusions about the effects of PH-targeted therapies.

Human central nervous system astrocytes support survival and activation of B cells: implications for MS pathogenesis.

BACKGROUND: The success of clinical trials of selective B cell depletion in patients with relapsing multiple sclerosis (MS) indicates B cells are important contributors to peripheral immune responses involved in the development of new relapses. Such B cell contribution to peripheral inflammation likely involves antibody-independent mechanisms. Of growing interest is the potential that B cells, within the MS central nervous system (CNS), may also contribute to the propagation of CNS-compartmentalized inflammation in progressive (non-relapsing) disease. B cells are known to persist in the inflamed MS CNS and are more recently described as concentrated in meningeal immune-cell aggregates, adjacent to the subpial cortical injury which has been associated with progressive disease. How B cells are fostered within the MS CNS and how they may contribute locally to the propagation of CNS-compartmentalized inflammation remain to be elucidated. METHODS: We considered whether activated human astrocytes might contribute to B cell survival and function through soluble factors. B cells from healthy controls (HC) and untreated MS patients were exposed to primary human astrocytes that were either maintained under basal culture conditions (non-activated) or pre-activated with standard inflammatory signals. B cell exposure to astrocytes included direct co-culture, co-culture in transwells, or exposure to astrocyte-conditioned medium. Following the different exposures, B cell survival and expression of T cell co-stimulatory molecules were assessed by flow cytometry, as was the ability of differentially exposed B cells to induce activation of allogeneic T cells. RESULTS: Secreted factors from both non-activated and activated human astrocytes robustly supported human B cell survival. Soluble products of pre-activated astrocytes also induced B cell upregulation of antigen-presenting cell machinery, and these B cells, in turn, were more efficient activators of T cells. Astrocyte-soluble factors could support survival and activation of B cell subsets implicated in MS, including memory B cells from patients with both relapsing and progressive forms of disease. CONCLUSIONS: Our findings point to a potential mechanism whereby activated astrocytes in the inflamed MS CNS not only promote a B cell fostering environment, but also actively support the ability of B cells to contribute to the propagation of CNS-compartmentalized inflammation, now thought to play key roles in progressive disease.

Mitochondrial HSP90 Accumulation Promotes Vascular Remodeling in Pulmonary Arterial Hypertension.

RATIONALE: Pulmonary arterial hypertension (PAH) is a vascular remodeling disease with a poor prognosis and limited therapeutic options. Although the mechanisms contributing to vascular remodeling in PAH are still unclear, several features, including hyperproliferation and resistance to apoptosis of pulmonary artery smooth muscle cells (PASMCs), have led to the emergence of the cancer-like concept. The molecular chaperone HSP90 (heat shock protein 90) is directly associated with malignant growth and proliferation under stress conditions. In addition to being highly expressed in the cytosol, HSP90 exists in a subcellular pool compartmentalized in the mitochondria (mtHSP90) of tumor cells, but not in normal cells, where it promotes cell survival. OBJECTIVES: We hypothesized that mtHSP90 in PAH-PASMCs represents a protective mechanism against stress, promoting their proliferation and resistance to apoptosis. METHODS: Expression and localization of HSP90 were analyzed by Western blot, immunofluorescence, and immunogold electron microscopy. In vitro, effects of mtHSP90 inhibition on mitochondrial DNA integrity, bioenergetics, cell proliferation and resistance to apoptosis were assessed. In vivo, the therapeutic potential of Gamitrinib, a mitochondria-targeted HSP90 inhibitor, was tested in fawn-hooded and monocrotaline rats. MEASUREMENTS AND MAIN RESULTS: We demonstrated that, in response to stress, HSP90 preferentially accumulates in PAH-PASMC mitochondria (dual immunostaining, immunoblot, and immunogold electron microscopy) to ensure cell survival by preserving mitochondrial DNA integrity and bioenergetic functions. Whereas cytosolic HSP90 inhibition displays a lack of absolute specificity for PAH-PASMCs, Gamitrinib decreased mitochondrial DNA content and repair capacity and bioenergetic functions, thus repressing PAH-PASMC proliferation (Ki67 labeling) and resistance to apoptosis (Annexin V assay) without affecting control cells. In vivo, Gamitrinib improves PAH in two experimental rat models (monocrotaline and fawn-hooded rat). CONCLUSIONS: Our data show for the first time that accumulation of mtHSP90 is a feature of PAH-PASMCs and a key regulator of mitochondrial homeostasis contributing to vascular remodeling in PAH.

FOXM1 promotes pulmonary artery smooth muscle cell expansion in pulmonary arterial hypertension.

Pulmonary arterial hypertension (PAH) is a progressive vascular remodeling disease characterized by a persistent elevation of pulmonary artery pressure, leading to right heart failure and premature death. Exaggerated proliferation and resistance to apoptosis of pulmonary artery smooth muscle cells (PASMCs) is a key component of vascular remodeling. Despite major advances in the field, current therapies for PAH remain poorly effective in reversing the disease or significantly improving long-term survival. Because the transcription factor FOXM1 is necessary for PASMC proliferation during lung morphogenesis and its overexpression stimulates proliferation and evasion of apoptosis in cancer cells, we thus hypothesized that upregulation of FOXM1 in PAH-PASMCs promotes cell expansion and vascular remodeling. Our results showed that FOXM1 was markedly increased in distal pulmonary arteries and isolated PASMCs from PAH patients compared to controls as well as in two preclinical models. In vitro, we showed that miR-204 expression regulates FOXM1 levels and that inhibition of FOXM1 reduced cell proliferation and resistance to apoptosis through diminished DNA repair mechanisms and decreased expression of the pro-remodeling factor survivin. Accordingly, inhibition of FOXM1 with thiostrepton significantly improved established PAH in two rat models. Thus, we show for the first time that FOXM1 is implicated in PAH development and represents a new promising target. KEY MESSAGES: FOXM1 is overexpressed in human PAH-PASMCs and PAH animal models. FOXM1 promotes PAH-PASMC proliferation and resistance to apoptosis. Pharmacological inhibition of FOXM1 improves established PAH in the MCT and Su/Hx rat models. FOXM1 may be a novel therapeutic target in PAH.

Trial Duration and Risk Reduction in Combination Therapy Trials for Pulmonary Arterial Hypertension: A Systematic Review.

BACKGROUND: Relative risk (RR) and number needed-to-treat (NNT) are frequently time-dependant measures. We performed a systematic review and meta-analysis to assess whether trial duration influenced the relative and absolute risk of worsening in randomized controlled trials (RCTs) comparing combination therapy (CT) of pulmonary arterial hypertension (PAH)-specific therapies vs monotherapy (MT). METHODS: We searched MEDLINE, Embase, and the Cochrane Library (January 1990 to September 2016) for RCTs assessing CT compared with MT in PAH. The primary outcome was the risk of clinical worsening. We assessed whether trial duration correlated with RR and NNT using weighted meta-regression with mixed effects. Changes in NNT overtime were also assessed using data from long-term event-driven trials. RESULTS: There were 3,801 patients throughout 15 studies included. The RR for clinical worsening positively correlated with trial duration (R2 = 0.67, P = .0002), whereas the NNT did not (mean NNT, 7; R2 = 0.02; P = .65). Among long-term event-driven trials, the mean NNT progressively decreased until 52 weeks of follow-up, being stable thereafter. Conversely, the mean RR progressively increased from approximately 0.40 at week 16 to approximately 0.68 at week 104. CONCLUSIONS: Absolute risk reduction of clinical worsening was relatively constant beyond 6 to 12 months of treatment in clinical trials comparing CT with MT in PAH. These results question the need for CT trials of very long duration in PAH.

Peripheral and local predictive immune signatures identified in a phase II trial of ipilimumab with carboplatin/paclitaxel in unresectable stage III or stage IV melanoma.

BACKGROUND: Checkpoint blockade with ipilimumab provides long-term survival to a significant proportion of patients with metastatic melanoma. New approaches to increase survival and to predict which patients will benefit from treatment are needed. This phase II trial combined ipilimumab with carboplatin/paclitaxel (CP) to assess its safety, efficacy, and to search for peripheral and tumor-based predictive biomarkers. METHODS: Thirty patients with untreated unresectable/metastatic melanoma were treated with ipilimumab and CP. Adverse events (AEs) were monitored and response to treatment was evaluated. Tumor tissue and peripheral blood were collected at specified time points to characterize tumor immune markers by immunohistochemistry and systemic immune activity by multiplex assays and flow cytometry. RESULTS: Eighty three percent of patients received all 5 cycles of CP and 93% completed ipilimumab induction. Serious AEs occurred in 13% of patients, and no treatment-related deaths were observed. Best Overall Response Rate (BORR) and Disease Control Rate (DCR) were 27 and 57%, respectively. Median overall survival was 16.2 months. Response to treatment was positively correlated with a higher tumor CD3+ infiltrate (immune score) at baseline. NRAS and BRAF mutations were less frequent in patients who experienced clinical benefit. Assessment of peripheral blood revealed that non-responders had elevated baseline levels of CXCL8 and CCL4, and a higher proportion of circulating late differentiated B cells. Pre-existing high levels of chemokines (CCL3, CCL4 and CXCL8) and advanced B cell differentiation were strongly associated with worse patient overall survival. Elevated proportions of circulating CD8+/PD-1+ T cells during treatment were associated with worse survival. CONCLUSIONS: The combination of ipilimumab and CP was well tolerated and revealed novel characteristics associated with patients likely to benefit from treatment. A pre-existing systemic inflammatory state characterized by elevation of selected chemokines and advanced B cell differentiation, was strongly associated with poor patient outcomes, revealing potential predictive circulating biomarkers. TRIAL REGISTRATION: Clinicaltrials.gov , NCT01676649 , registered on August 29, 2012.

HDAC6: A Novel Histone Deacetylase Implicated in Pulmonary Arterial Hypertension.

Pulmonary arterial hypertension (PAH) is a vascular remodeling disease with limited therapeutic options. Although exposed to stressful conditions, pulmonary artery (PA) smooth muscle cells (PASMCs) exhibit a "cancer-like" pro-proliferative and anti-apoptotic phenotype. HDAC6 is a cytoplasmic histone deacetylase regulating multiple pro-survival mechanisms and overexpressed in response to stress in cancer cells. Due to the similarities between cancer and PAH, we hypothesized that HDAC6 expression is increased in PAH-PASMCs to face stress allowing them to survive and proliferate, thus contributing to vascular remodeling in PAH. We found that HDAC6 is significantly up-regulated in lungs, distal PAs, and isolated PASMCs from PAH patients and animal models. Inhibition of HDAC6 reduced PAH-PASMC proliferation and resistance to apoptosis in vitro sparing control cells. Mechanistically, we demonstrated that HDAC6 maintains Ku70 in a hypoacetylated state, blocking the translocation of Bax to mitochondria and preventing apoptosis. In vivo, pharmacological inhibition of HDAC6 improved established PAH in two experimental models and can be safely given in combination with currently approved PAH therapies. Moreover, Hdac6 deficient mice were partially protected against chronic hypoxia-induced pulmonary hypertension. Our study shows for the first time that HDAC6 is implicated in PAH development and represents a new promising target to improve PAH.

Implication of Inflammation and Epigenetic Readers in Coronary Artery Remodeling in Patients With Pulmonary Arterial Hypertension.

OBJECTIVE: Pulmonary arterial hypertension (PAH) is a vascular disease not restricted to the lungs. Many signaling pathways described in PAH are also of importance in other vascular remodeling diseases, such as coronary artery disease (CAD). Intriguingly, CAD is 4× more prevalent in PAH compared with the global population, suggesting a link between these 2 diseases. Both PAH and CAD are associated with sustained inflammation and smooth muscle cell proliferation/apoptosis imbalance and we demonstrated in PAH that this phenotype is, in part, because of the miR-223/DNA damage/Poly[ADP-ribose] polymerase 1/miR-204 axis activation and subsequent bromodomain protein 4 (BRD4) overexpression. Interestingly, BRD4 is also a trigger for calcification and remodeling processes, both of which are important in CAD. Thus, we hypothesize that BRD4 activation in PAH influences the development of CAD. APPROACH AND RESULTS: PAH was associated with significant remodeling of the coronary arteries in both human and experimental models of the disease. As observed in PAH distal pulmonary arteries, coronary arteries of patients with PAH also exhibited increased DNA damage, inflammation, and BRD4 overexpression. In vitro, using human coronary artery smooth muscle cells from PAH, CAD and non-PAH-non-CAD patients, we showed that both PAH and CAD smooth muscle cells exhibited increased proliferation and suppressed apoptosis in a BRD4-dependent manner. In vivo, improvement of PAH by BRD4 inhibitor was associated with a reduction in coronary remodeling and interleukin-6 expression. CONCLUSIONS: Overall, this study demonstrates that increased BRD4 expression in coronary arteries of patient with PAH contributes to vascular remodeling and comorbidity development.