Citations

Abstract Pancreatic ductal adenocarcinoma (PDAC), as with all cancer, is driven by dysregulation of genetic programs due to accumulation of somatic mutations, epigenetic modifications and changes in the micro-environment. Holistic views of perturbations in gene expression networks could illuminate key regulators and pathways for this highly lethal cancer. Toward this end, we performed massively-parallel mRNA-sequencing in normal (n = 10) and tumor (n = 8) derived pancreatic tissue samples as well as pancreatic cancer cell lines (n = 9), and determined differential gene expression (DE) patterns. Sub-network enrichment analyses of all expressed genes based on magnitude of DE identified HNF1A as the regulator of the most significantly and consistently dysregulated expression sub-network in our pancreatic tumor samples (median P = 7.56 x 10-7, median rank = 1, range = 1-25). To explore the effect of HNF1A expression in pancreatic tumor-derived cells we generated stable HNF1A-inducible clones in two pancreatic cancer cell lines. Induction of HNF1A overexpression caused severe growth inhibition (5.3-fold, P = 4.5x10-5 for MIA PaCa-2 clones; 7.2-fold, P = 2.2x10-5 for PANC-1 clones), and G0/G1 cell cycle arrest. This was accompanied by down-regulation of 49 out of 84 assayed cell cycle genes, while only 1 displayed increased expression. These data, combined with HNF1A’s direct regulation of pancreatic development and homeostasis genes (i.e. PDX1, PTF1A and NR5A2), suggest it may be an important tumor suppressor in pancreatic cells. Citation Format: Jason W. Hoskins, Marta Flandez, Jinping Jia, Hemang Parikh, Irene Collins, Mickey Emmanuel, Abdisamad Ibrahim, Wenming Xiao, John Powell, Nuria Malats, Gloria M. Petersen, Fransisco X. Real, Laufey T. Amundadottir. Transcriptome analysis in pancreatic cancer reveals a tumor suppressor function for HNF1A. [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer: Innovations in Research and Treatment; May 18-21, 2014; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2015;75(13 Suppl):Abstract nr B112.

Abstract A genome-wide association study (GWAS) of pancreatic cancer conducted within the NCI Cohort Consortium (PanScan I and II) identified pancreatic cancer susceptibility loci on chromosomes 1q32.1/NR5A2, 5p15.33/CLPTM1L/TERT, 9q34.2/ABO, and 13q22.1. The most significant single-nucleotide polymorphism (SNP) identified on 13q22.1, rs9543325, lies in a 600 kb gene desert; the nearest genes are KLF5, KLF12, PIBF1, DIS3, and BORA (265–586 kb). Imputation using the 1000 Genomes and DCEG reference datasets did not improve the GWAS signal, but produced a set of highly correlated SNPs for functional follow-up. We performed eQTL analysis to test for association between the genotypes of these functional candidate variants and expression of nearby genes. Among 100 normal derived pancreatic tissue samples, DIS3 expression showed the strongest association with a novel 30 bp indel variant in the risk locus (P = 4.0 × 104), indicating risk alleles associate with reduced DIS3 expression. Mutations in DIS3 have previously been identified in acute myeloid leukemia and multiple myeloma, and its expression has been correlated with metastatic potential in colorectal cancer, suggesting DIS3 is relevant to cancer biology. Chromosome conformation capture identified a physical interaction between the indel-containing locus and a region near the DIS3 promoter. Luciferase assay for regulatory function of this indel-containing locus revealed allele-specific silencer activity for the insertion allele. Supershift electromobility shift assay (EMSA) demonstrated binding of LEF1 specifically to the insertion allele of the indel, which contains two in silico predicted LEF1 binding elements. Finally, through immunohistochemical analysis, high DIS3 protein levels associated with better survival for pancreatic cancer patients (hazard ratio = 2.87, 95% CI = 1.49–5.53, P = 0.001). Our results suggest that at least one target gene for the pancreatic cancer risk variants on chr13q22.1 may be DIS3, and that the underlying biology may be mediated by the novel indel through a long-range repressive effect on DIS3 expression. Citation Format: Jason W. Hoskins, Abdisamad Ibrahim, Mickey Emmanuel, Hemang Parikh, Jinping Jia, Irene Collins, Kris Ylaya, Sean F. Altekruse, Stephen M. Hewitt, Gloria M. Petersen, Laufey T. Amundadottir. Functional analysis of the chr13q22.1 pancreatic cancer risk locus suggests allele-specific effects on DIS3 expression with prognostic implications. [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer: Innovations in Research and Treatment; May 18-21, 2014; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2015;75(13 Suppl):Abstract nr B111.

Mixed lineage kinase 3 (MLK3) deficiency reduces macrophage‐associated inflammation in a murine model of nonalcoholic steatohepatitis (NASH). However, the mechanistic links between MLK3 activation in hepatocytes and macrophage‐driven inflammation in NASH are uncharted. Herein, we report that MLK3 mediates the release of (C‐X‐C motif) ligand 10 (CXCL10)‐laden extracellular vesicles (EVs) from lipotoxic hepatocytes, which induce macrophage chemotaxis. Primary mouse hepatocytes (PMHs) and Huh7 cells were treated with palmitate or lysophosphatidylcholine (LPC). Released EVs were isolated by differential ultracentrifugation. LPC treatment of PMH or Huh7 cells induced release of EVs, which was prevented by either genetic or pharmacological inhibition of MLK3. Mass spectrometry identified the potent chemokine, CXCL10, in the EVs, which was markedly enriched in EVs isolated from LPC‐treated hepatocytes versus untreated cells. Green fluorescent protein (GFP)‐tagged CXCL10 was present in vesicular structures and colocalized with the red fluorescent protein (RFP)‐tagged EV marker, CD63, after LPC treatment of cotransfected Huh‐7 cells. Either genetic deletion or pharmacological inhibition of MLK3 prevented CXCL10 enrichment in EVs. Treatment of mouse bone‐marrow–derived macrophages with lipotoxic hepatocyte‐derived EVs induced macrophage chemotaxis, an effect blocked by incubation with CXCL10‐neutralizing antisera. MLK3‐deficient mice fed a NASH‐inducing diet had reduced concentrations of total plasma EVs and CXCL10 containing EVs compared to wild‐type mice. Conclusions: During hepatocyte lipotoxicity, activated MLK3 induces the release of CXCL10‐bearing vesicles from hepatocytes, which are chemotactic for macrophages. (Hepatology 2016;63:731–744)

A 37-year-old African-American woman with end-stage renal disease presumed to be secondary to diabetes mellitus type 2, on daily peritoneal dialysis, was admitted for a painful left lower extremity lesion. Examination revealed a large, dusky, tender region over the left lateral thigh. She was on warfarin for mechanical heart valves. Despite discontinuation of warfarin and placement on heparin, the lesion progressed to extend to the medial left thigh and medial and lateral right thigh. CT scan demonstrated arteriolar medial calcification and vascular calcification of the small subcutaneous vessels, without evidence of abscess or haematoma. The patient declined punch biopsy. Given the known risk factors of high calcium-phosphate and radiological findings, a diagnosis of calcific uraemic arteriolopathy was made. Phosphate-binder therapy was optimised. She was transitioned to daily haemodialysis, and sodium thiosulfate was initiated. Skin lesions demonstrated improvement at her 5 weeks posthospitalisation follow-up.

AbstractBackgroundLate‐onset Alzheimer's disease (AD) is heritable with 20 genes showing genome‐wide association in the International Genomics of Alzheimer's Project (IGAP). To identify the biology underlying the disease, we extended these genetic data in a pathway analysis.MethodsThe ALIGATOR and GSEA algorithms were used in the IGAP data to identify associated functional pathways and correlated gene expression networks in human brain.ResultsALIGATOR identified an excess of curated biological pathways showing enrichment of association. Enriched areas of biology included the immune response (P = 3.27 × 10−12 after multiple testing correction for pathways), regulation of endocytosis (P = 1.31 × 10−11), cholesterol transport (P = 2.96 × 10−9), and proteasome‐ubiquitin activity (P = 1.34 × 10−6). Correlated gene expression analysis identified four significant network modules, all related to the immune response (corrected P = .002–.05).ConclusionsThe immune response, regulation of endocytosis, cholesterol transport, and protein ubiquitination represent prime targets for AD therapeutics.

Background The greatest potential to reduce the burden of stroke is by primary prevention of first-ever stroke, which constitutes three quarters of all stroke. In addition to population-wide prevention strategies (the ‘mass’ approach), the ‘high risk’ approach aims to identify individuals at risk of stroke and to modify their risk factors, and risk, accordingly. Current methods of assessing and modifying stroke risk are difficult to access and implement by the general population, amongst whom most future strokes will arise. To help reduce the burden of stroke on individuals and the population a new app, the Stroke Riskometer™, has been developed. We aim to explore the validity of the app for predicting the risk of stroke compared with current best methods. Methods 752 stroke outcomes from a sample of 9501 individuals across three countries (New Zealand, Russia and the Netherlands) were utilized to investigate the performance of a novel stroke risk prediction tool algorithm (Stroke Riskometer™) compared with two established stroke risk score prediction algorithms (Framingham Stroke Risk Score [FSRS] and QStroke). We calculated the receiver operating characteristics (ROC) curves and area under the ROC curve (AUROC) with 95% confidence intervals, Harrels C-statistic and D-statistics for measure of discrimination, R2 statistics to indicate level of variability accounted for by each prediction algorithm, the Hosmer-Lemeshow statistic for calibration, and the sensitivity and specificity of each algorithm. Results The Stroke Riskometer™ performed well against the FSRS five-year AUROC for both males (FSRS = 75·0% (95% CI 72·3%–77·6%), Stroke Riskometer™ = 74·0(95% CI 71·3%–76·7%) and females [FSRS = 70·3% (95% CI 67·9%–72·8%, Stroke Riskometer™ = 71·5% (95% CI 69·0%–73·9%)], and better than QStroke [males–59·7% (95% CI 57·3%–62·0%) and comparable to females = 71·1% (95% CI 69·0%–73·1%)]. Discriminative ability of all algorithms was low (C-statistic ranging from 0·51–0·56, D-statistic ranging from 0·01–0·12). Hosmer-Lemeshow illustrated that all of the predicted risk scores were not well calibrated with the observed event data ( P < 0·006). Conclusions The Stroke Riskometer™ is comparable in performance for stroke prediction with FSRS and QStroke. All three algorithms performed equally poorly in predicting stroke events. The Stroke Riskometer™ will be continually developed and validated to address the need to improve the current stroke risk scoring systems to more accurately predict stroke, particularly by identifying robust ethnic/race ethnicity group and country specific risk factors.

Introduction Pulmonary hypertension (PH) is a devastating, progressive disease with increasingly debilitating symptoms and usually shortened overall life expectancy due to a narrowing of the pulmonary vasculature and consecutive right heart failure. Little is known about PH in Africa, but limited reports suggest that PH is more prevalent in Africa compared with developed countries due to the high prevalence of risk factors in the region. Methods and analysis A multinational multicentre registry-type cohort study was established and tailored to resource-constraint settings to describe disease presentation, disease severity and aetiologies of PH, comorbidities, diagnostic and therapeutic management, and the natural course of PH in Africa. PH will be diagnosed by specialist cardiologists using echocardiography (right ventricular systolic pressure >35 mm Hg, absence of pulmonary stenosis and acute right heart failure), usually accompanied by shortness of breath, fatigue, peripheral oedema and other cardiovascular symptoms, ECG and chest X-ray changes in keeping with PH as per guidelines (European Society of Cardiology and European Respiratory Society (ESC/ERS) guidelines). Additional investigations such as a CT scan, a ventilation/perfusion scan or right heart catheterisation will be performed at the discretion of the treating physician. Functional tests include a 6 min walk test and the Karnofsky Performance Score. The WHO classification system for PH will be applied to describe the different aetiologies of PH. Several substudies have been implemented within the registry to investigate specific types of PH and their outcome at up to 24 months. Data will be analysed by an independent institution following a data analyse plan. Ethics and dissemination All local ethics committees of the participating centres approved the protocol. The data will be disseminated through peer-reviewed journals at national and international conferences and public events at local care providers.