YAP/TAZ Activation Drives Uveal Melanoma Initiation and Progression

Uveal melanoma (UM), the most common ocular malignancy, is characterized by GNAQ/11 mutations. Hippo/YAP and Ras/mitogen-activated protein kinase (MAPK) emerge as two important signaling pathways downstream of G protein alpha subunits of the Q class (GalphaQ/11)-mediated transformation, although whether and how they contribute to UM genesis in vivo remain unclear. Here, we adapt an adeno-associated virus (AAV)-based ocular injection method to directly deliver Cre recombinase into the mouse uveal tract and demonstrate that Lats1/2 kinases suppress UM formation specifically in uveal melanocytes. We find that genetic activation of YAP, but not Kras, is sufficient to initiate UM. We show that YAP/TAZ activation induced by Lats1/2 deletion cooperates with Kras to promote UM progression via downstream transcriptional reinforcement. Furthermore, dual inhibition of YAP/TAZ and Ras/MAPK synergizes to suppress oncogenic growth of human UM cells. Our data highlight the functional significance of Lats-YAP/TAZ in UM initiation and progression in vivo and suggest combination inhibition of YAP/TAZ and Ras/MAPK as a new therapeutic strategy for UM

Clinically Actionable Hypercholesterolemia and Hypertriglyceridemia in Children with Nonalcoholic Fatty Liver Disease

OBJECTIVE: To determine the percentage of children with nonalcoholic fatty liver disease (NAFLD) in whom intervention for low-density lipoprotein cholesterol or triglycerides was indicated based on National Heart, Lung, and Blood Institute guidelines. STUDY DESIGN: This multicenter, longitudinal cohort study included children with NAFLD enrolled in the National Institute of Diabetes and Digestive and Kidney Diseases Nonalcoholic Steatohepatitis Clinical Research Network. Fasting lipid profiles were obtained at diagnosis. Standardized dietary recommendations were provided. After 1 year, lipid profiles were repeated and interpreted according to National Heart, Lung, and Blood Institute Expert Panel on Integrated Guidelines for Cardiovascular Health and Risk Reduction. Main outcomes were meeting criteria for clinically actionable dyslipidemia at baseline, and either achieving lipid goal at follow-up or meeting criteria for ongoing intervention. RESULTS: There were 585 participants, with a mean age of 12.8 years. The prevalence of children warranting intervention for low-density lipoprotein cholesterol at baseline was 14%. After 1 year of recommended dietary changes, 51% achieved goal low-density lipoprotein cholesterol, 27% qualified for enhanced dietary and lifestyle modifications, and 22% met criteria for pharmacologic intervention. Elevated triglycerides were more prevalent, with 51% meeting criteria for intervention. At 1 year, 25% achieved goal triglycerides with diet and lifestyle changes, 38% met criteria for advanced dietary modifications, and 37% qualified for antihyperlipidemic medications. CONCLUSIONS: More than one-half of children with NAFLD met intervention thresholds for dyslipidemia. Based on the burden of clinically relevant dyslipidemia, lipid screening in children with NAFLD is warranted. Clinicians caring for children with NAFLD should be familiar with lipid management

STIM1 signalling controls store-operated calcium entry required for development and contractile function in skeletal muscle

It is now well established that stromal interaction molecule 1 (STIM1) is the calcium sensor of endoplasmic reticulum (ER) stores required to activate store-operated calcium entry (SOC) channels at the surface of non-excitable cells. Yet little is known about STIM1 in excitable cells such as striated muscle where the complement of calcium regulatory molecules is rather disparate from that of non-excitable cells. Here, we show that STIM1 is expressed in both myotubes and adult skeletal muscle. Myotubes lacking functional STIM1 fail to exhibit SOC and fatigue rapidly. Moreover, mice lacking functional STIM1 die perinatally from a skeletal myopathy. In addition, STIM1 haploinsufficiency confers a contractile defect only under conditions where rapid refilling of stores would be needed. These findings provide novel insight to the role of STIM1 in skeletal muscle and suggest that STIM1 has a universal role as an ER/SR calcium sensor in both excitable and non-excitable cells

Highly Parallel Genome-Wide Expression Analysis of Single Mammalian Cells

We have developed a high-throughput amplification method for generating robust gene expression profiles using single cell or low RNA inputs.The method uses tagged priming and template-switching, resulting in the incorporation of universal PCR priming sites at both ends of the synthesized cDNA for global PCR amplification. Coupled with a whole-genome gene expression microarray platform, we routinely obtain expression correlation values of R(2)~0.76-0.80 between individual cells and R(2)~0.69 between 50 pg total RNA replicates. Expression profiles generated from single cells or 50 pg total RNA correlate well with that generated with higher input (1 ng total RNA) (R(2)~0.80). Also, the assay is sufficiently sensitive to detect, in a single cell, approximately 63% of the number of genes detected with 1 ng input, with approximately 97% of the genes detected in the single-cell input also detected in the higher input.In summary, our method facilitates whole-genome gene expression profiling in contexts where starting material is extremely limiting, particularly in areas such as the study of progenitor cells in early development and tumor stem cell biology

XPO1 expression worsens the prognosis of unfavorable DLBCL that can be effectively targeted by selinexor in the absence of mutant p53

Additional file 1. Table S1: Clinicopathologic and molecular characteristics of DLBCL patients with high or low XPO1 expression. Table S2: Significantly differentially expressed genes between XPO1high and XPO1low DLBCL patients with concurrent TP53 mutation and high MYC expression. Figure S1: Biomarker study for XPO1 and selinexor. (A–B) XPO1high expression showed significant adverse prognostic impact in the ABC subtype but not the GCB subtype of DLBCL. (C) XPO1high expression showed a trend of unfavorable prognostic effect on PFS in MYC-rearranged (MYC-R+) DLBCL. (D) XPO1high expression was associated with significantly poorer survival in DLBCL patients with wild type (Wt) TP53. (E) ABC-DLBCL and GCB-DLBCL cells showed similar sensitivity to the cytotoxicity of selinexor. (F) TP53 mutation (Mut-TP53) significantly reduced the anti-lymphoma efficacy of selinexor in HGBCL-DH cells. IC50 values were calculated by GraphPad Prism 8 based on the cell viability data after 72-hour treatment

Phase II study of the farnesyltransferase inhibitor R115777 in advanced melanoma (CALGB 500104)

BACKGROUND: Multiple farnesylated proteins are involved in signal transduction in cancer. Farnesyltransferase inhibitors (FTIs) have been developed as a strategy to inhibit the function of these proteins. As FTIs inhibit proliferation of melanoma cell lines, we undertook a study to assess the impact of a FTI in advanced melanoma. As farnesylated proteins are also important for T cell activation, measurement of effects on T cell function was also pursued. METHODS: A 3-stage trial design was developed with a maximum of 40 patients and early stopping if there were no responders in the first 14, or fewer than 2 responders in the first 28 patients. Eligibility included performance status of 0–1, no prior chemotherapy, at most 1 prior immunotherapy, no brain metastases, and presence of at least 2 cutaneous lesions amenable to biopsy. R115777 was administered twice per day for 21 days of a 28-day cycle. Patients were evaluated every 2 cycles by RECIST. Blood and tumor were analyzed pre-treatment and during week 7. RESULTS: Fourteen patients were enrolled. Two patients had grade 3 toxicities, which included myelosuppression, nausea/vomiting, elevated BUN, and anorexia. There were no clinical responses. All patients analyzed showed potent inhibition of FT activity (85-98%) in tumor tissue; inhibition of phosphorylated ERK and Akt was also observed. T cells showed evidence of FT inhibition and diminished IFN-γ production. CONCLUSIONS: Despite potent target inhibition, R115777 showed no evidence of clinical activity in this cohort of melanoma patients. Inhibition of T cell function by FTIs has potential clinical implications. Clinicaltrials.gov number NCT0006012

Topoisomerase II alpha gene copy loss has adverse prognostic significance in ERBB2-amplified breast cancer: a retrospective study of paraffin-embedded tumor specimens and medical charts

<p>Abstract</p> <p>Background</p> <p>Amplification of the <it>ERBB2 </it>(<it>Her-2/neu</it>) oncogene, which occurs in approximately 25% of breast carcinomas, is a known negative prognostic factor. Available data indicate that a variable number of nearby genes on chromosome 17q may be co-amplified or deleted, forming a continuous amplicon of variable size. In approximately 25% of these patients, the amplicon extends to the gene for <it>topoisomerase II alpha </it>(<it>TOP2A</it>), a target for anthracyclines. We sought to understand the significance of these associated genomic changes for breast cancer prognosis and predicting response to therapy.</p> <p>Methods and patients</p> <p>Archival tissue samples from 63 breast cancer patients with <it>ERBB2 </it>amplification, stages 0–IV, were previously analyzed with FISH probes for genes located near <it>ERBB2</it>. In the present study, the clinical outcome data were determined for all patients presenting at stages I–III for whom adequate clinical follow up was available.</p> <p>Results</p> <p>Four amplicon patterns (Classes) were identified. These were significantly associated with the clinical outcome, specifically, recurrence of breast cancer. The Amplicon class IV with deleted <it>TOP2A </it>had 67% (6/9) cases with recurrence, whereas the other three classes combined had only 12% (3/25) cases (p-value = 0.004) at the time of last follow-up. <it>TOP2A </it>deletion was also significantly associated with time to recurrence (p-value = 0.0002). After adjusting for age in Cox regression analysis, the association between <it>TOP2A </it>deletion and time to recurrence remains strongly significant (p-value = 0.002) whereas the association with survival is marginally significant (p-value = 0.06).</p> <p>Conclusion</p> <p><it>TOP2A </it>deletion is associated with poor prognosis in <it>ERBB2</it>-amplified breast carcinomas. Clarification of the mechanism of this association will require additional study.</p

In Children with Nonalcoholic Fatty Liver Disease, Zone 1 Steatosis is Associated with Advanced Fibrosis

Background & Aims Focal zone 1 steatosis, although rare in adults with nonalcoholic fatty liver disease (NAFLD), does occur in children with NAFLD. We investigated whether focal zone 1 steatosis and focal zone 3 steatosis are distinct subphenotypes of pediatric NAFLD. We aimed to determine associations between the zonality of steatosis and demographic, clinical, and histologic features in children with NAFLD. Methods We performed a cross-sectional study of baseline data from 813 children (age <18 years; mean age, 12.8 ± 2.7 years). The subjects had biopsy-proven NAFLD and were enrolled in the Nonalcoholic Steatohepatitis Clinical Research Network. Liver histology was reviewed using the Nonalcoholic Steatohepatitis Clinical Research Network scoring system. Results Zone 1 steatosis was present in 18% of children with NAFLD (n = 146) and zone 3 steatosis was present in 32% (n = 244). Children with zone 1 steatosis were significantly younger (10 vs 14 years; P < .001) and a significantly higher proportion had any fibrosis (81% vs 51%; P < .001) or advanced fibrosis (13% vs 5%; P < .001) compared with children with zone 3 steatosis. In contrast, children with zone 3 steatosis were significantly more likely to have steatohepatitis (30% vs 6% in children with zone 1 steatosis; P < .001). Conclusions Children with zone 1 or zone 3 distribution of steatosis have an important subphenotype of pediatric NAFLD. Children with zone 1 steatosis are more likely to have advanced fibrosis and children with zone 3 steatosis are more likely to have steatohepatitis. To achieve a comprehensive understanding of pediatric NAFLD, studies of pathophysiology, natural history, and response to treatment should account for the zonality of steatosis

Co-evolution of a broadly neutralizing HIV-1 antibody and founder virus

Current HIV-1 vaccines elicit strain-specific neutralizing antibodies. However, cross-reactive neutralizing antibodies arise in ~20% of HIV-1-infected individuals, and details of their generation could provide a roadmap for effective vaccination. Here we report the isolation, evolution and structure of a broadly neutralizing antibody from an African donor followed from time of infection. The mature antibody, CH103, neutralized ~55% of HIV-1 isolates, and its co-crystal structure with gp120 revealed a novel loop-based mechanism of CD4-binding site recognition. Virus and antibody gene sequencing revealed concomitant virus evolution and antibody maturation. Notably, the CH103-lineage unmutated common ancestor avidly bound the transmitted/founder HIV-1 envelope glycoprotein, and evolution of antibody neutralization breadth was preceded by extensive viral diversification in and near the CH103 epitope. These data elucidate the viral and antibody evolution leading to induction of a lineage of HIV-1 broadly neutralizing antibodies and provide insights into strategies to elicit similar antibodies via vaccination

Automated telephone communication systems for preventive healthcare and management of long-term conditions

Background Automated telephone communication systems (ATCS) can deliver voice messages and collect health-related information from patients using either their telephone’s touch-tone keypad or voice recognition software. ATCS can supplement or replace telephone contact between health professionals and patients. There are four different types of ATCS: unidirectional (one-way, non-interactive voice communication), interactive voice response (IVR) systems, ATCS with additional functions such as access to an expert to request advice (ATCS Plus) and multimodal ATCS, where the calls are delivered as part of a multicomponent intervention. Objectives To assess the effects of ATCS for preventing disease and managing long-term conditions on behavioural change, clinical, process, cognitive, patient-centred and adverse outcomes. Search methods We searched 10 electronic databases (the Cochrane Central Register of Controlled Trials; MEDLINE; Embase; PsycINFO; CINAHL; Global Health; WHOLIS; LILACS; Web of Science; and ASSIA); three grey literature sources (Dissertation Abstracts, Index to Theses, Australasian Digital Theses); and two trial registries (www.controlled-trials.com; www.clinicaltrials.gov) for papers published between 1980 and June 2015. Selection criteria Randomised, cluster- and quasi-randomised trials, interrupted time series and controlled before-and-after studies comparing ATCS interventions, with any control or another ATCS type were eligible for inclusion. Studies in all settings, for all consumers/carers, in any preventive healthcare or long term condition management role were eligible. Data collection and analysis We used standard Cochrane methods to select and extract data and to appraise eligible studies. Main results We included 132 trials (N = 4,669,689). Studies spanned across several clinical areas, assessing many comparisons based on evaluation of different ATCS types and variable comparison groups. Forty-one studies evaluated ATCS for delivering preventive healthcare, 84 for managing long-term conditions, and seven studies for appointment reminders. We downgraded our certainty in the evidence primarily because of the risk of bias for many outcomes. We judged the risk of bias arising from allocation processes to be low for just over half the studies and unclear for the remainder. We considered most studies to be at unclear risk of performance or detection bias due to blinding, while only 16% of studies were at low risk. We generally judged the risk of bias due to missing data and selective outcome reporting to be unclear. For preventive healthcare, ATCS (ATCS Plus, IVR, unidirectional) probably increase immunisation uptake in children (risk ratio (RR) 1.25, 95% confidence interval (CI) 1.18 to 1.32; 5 studies, N = 10,454; moderate certainty) and to a lesser extent in adolescents (RR 1.06, 95% CI 1.02 to 1.11; 2 studies, N = 5725; moderate certainty). The effects of ATCS in adults are unclear (RR 2.18, 95% CI 0.53 to 9.02; 2 studies, N = 1743; very low certainty). For screening, multimodal ATCS increase uptake of screening for breast cancer (RR 2.17, 95% CI 1.55 to 3.04; 2 studies, N = 462; high certainty) and colorectal cancer (CRC) (RR 2.19, 95% CI 1.88 to 2.55; 3 studies, N = 1013; high certainty) versus usual care. It may also increase osteoporosis screening. ATCS Plus interventions probably slightly increase cervical cancer screening (moderate certainty), but effects on osteoporosis screening are uncertain. IVR systems probably increase CRC screening at 6 months (RR 1.36, 95% CI 1.25 to 1.48; 2 studies, N = 16,915; moderate certainty) but not at 9 to 12 months, with probably little or no effect of IVR (RR 1.05, 95% CI 0.99, 1.11; 2 studies, 2599 participants; moderate certainty) or unidirectional ATCS on breast cancer screening. Appointment reminders delivered through IVR or unidirectional ATCS may improve attendance rates compared with no calls (low certainty). For long-term management, medication or laboratory test adherence provided the most general evidence across conditions (25 studies, data not combined). Multimodal ATCS versus usual care showed conflicting effects (positive and uncertain) on medication adherence. ATCS Plus probably slightly (versus control; moderate certainty) or probably (versus usual care; moderate certainty) improves medication adherence but may have little effect on adherence to tests (versus control). IVR probably slightly improves medication adherence versus control (moderate certainty). Compared with usual care, IVR probably improves test adherence and slightly increases medication adherence up to six months but has little or no effect at longer time points (moderate certainty). Unidirectional ATCS, compared with control, may have little effect or slightly improve medication adherence (low certainty). The evidence suggested little or no consistent effect of any ATCS type on clinical outcomes (blood pressure control, blood lipids, asthma control, therapeutic coverage) related to adherence, but only a small number of studies contributed clinical outcome data. The above results focus on areas with the most general findings across conditions. In condition-specific areas, the effects of ATCS varied, including by the type of ATCS intervention in use. Multimodal ATCS probably decrease both cancer pain and chronic pain as well as depression (moderate certainty), but other ATCS types were less effective. Depending on the type of intervention, ATCS may have small effects on outcomes for physical activity, weight management, alcohol consumption, and diabetes mellitus. ATCS have little or no effect on outcomes related to heart failure, hypertension, mental health or smoking cessation, and there is insufficient evidence to determine their effects for preventing alcohol/ substance misuse or managing illicit drug addiction, asthma, chronic obstructive pulmonary disease, HIV/AIDS, hypercholesterolaemia, obstructive sleep apnoea, spinal cord dysfunction or psychological stress in carers. Only four trials (3%) reported adverse events, and it was unclear whether these were related to the intervention