Evaluation of a new immunoassay for cystatin C, based on a double monoclonal principle, in men with normal and impaired renal function

BACKGROUND: Elevated cystatin C in blood reflects impaired glomerular filtration rate (GFR), but current cystatin C assays, based on polyclonal antibodies and immunoturbidimetric or nephelometric detection, have several limitations. We evaluated a new immunoassay based on monoclonal antibodies in samples from patients with and without chronic kidney disease (CKD). METHODS: The study enrolled 170 men without known CKD (Group A) and 104 men with CKD (Group B). All patients were assessed with iohexol clearance, plasma creatinine and plasma cystatin C by a conventional particle-enhanced immunoturbidimetric assay (PETIA) and by the new double monoclonal assay. In Group A, three serial blood draws were performed at median intervals of 4 h and 12 days between samples, to also allow assessments of the variability in cystatin C values with the new assay. Concordance correlation coefficients and the 95% limits of agreement were used to estimate the agreement of reciprocal cystatin C and reciprocal creatinine with iohexol clearance. RESULTS: Median iohexol clearance (mL/min/1.73 m(2)) was 81 [interquartile range (IQR) 70, 92] in Group A and 23 (IQR 16, 34) in Group B. The concordance correlation with GFR for the new cystatin C assay compared to the established assay was similar in Group A (0.441 versus 0.465) but higher in Group B (0.680 versus 0.593). Cystatin C measured by both assays exhibited closer agreement with GFR than creatinine. The agreement between the two cystatin C assays was high, with concordance correlations of 0.815 in Group A and 0.935 in Group B. Compared to the conventional assay, the new assay tended to yield lower values of cystatin C at the low end of the range in Group A. The new cystatin C assay exhibited small intraindividual variability across serial samples (coefficient of variation ≤ 6%). CONCLUSIONS: In this first clinical evaluation, the new cystatin C assay performed similarly to the established PETIA in patients with normal GFR and better in patients with CKD. The new assay may offer an alternative to current commercial assays to detect and monitor impaired kidney function

X-Linked Genes and Risk of Orofacial Clefts: Evidence from Two Population-Based Studies in Scandinavia

Background: Orofacial clefts are common birth defects of complex etiology, with an excess of males among babies with cleft lip and palate, and an excess of females among those with cleft palate only. Although genes on the X chromosome have been implicated in clefting, there has been no association analysis of X-linked markers. Methodology/Principal Findings: We added new functionalities in the HAPLIN statistical software to enable association analysis of X-linked markers and an exploration of various causal scenarios relevant to orofacial clefts. Genotypes for 48 SNPs in 18 candidate genes on the X chromosome were analyzed in two population-based samples from Scandinavia (562 Norwegian and 235 Danish case-parent triads). For haplotype analysis, we used a sliding-window approach and assessed isolated cleft lip with or without cleft palate (iCL/P) separately from isolated cleft palate only (iCPO). We tested three statistical models in HAPLIN, allowing for: i) the same relative risk in males and females, ii) sex-specific relative risks, and iii) X-inactivation in females. We found weak but consistent associations with the oral-facial-digital syndrome 1 (OFD1) gene (formerly known as CXORF5) in the Danish iCL/P samples across all models, but not in the Norwegian iCL/P samples. In sex-specific analyses, the association with OFD1 was in male cases only. No analyses showed associations with iCPO in either the Norwegian or the Danish sample. Conclusions: The association of OFD1 with iCL/P is plausible given the biological relevance of this gene. However, the lack of replication in the Norwegian samples highlights the need to verify these preliminary findings in other large datasets. More generally, the novel analytic methods presented here are widely applicable to investigations of the role of X-linked genes in complex traits

c-MYC expression sensitizes medulloblastoma cells to radio- and chemotherapy and has no impact on response in medulloblastoma patients

BACKGROUND: To study whether and how c-MYC expression determines response to radio- and chemotherapy in childhood medulloblastoma (MB). METHODS: We used DAOY and UW228 human MB cells engineered to stably express different levels of c-MYC, and tested whether c-MYC expression has an effect on radio- and chemosensitivity using the colorimetric 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium inner salt (MTS) assay, clonogenic survival, apoptosis assays, cell cycle analysis, and western blot assessment. In an effort to validate our results, we analyzed c-MYC mRNA expression in formalin-fixed paraffin-embedded tumor samples from well-documented patients with postoperative residual tumor and compared c-MYC mRNA expression with response to radio- and chemotherapy as examined by neuroradiological imaging. RESULTS: In DAOY - and to a lesser extent in UW228 - cells expressing high levels of c-MYC, the cytotoxicity of cisplatin, and etoposide was significantly higher when compared with DAOY/UW228 cells expressing low levels of c-MYC. Irradiation- and chemotherapy-induced apoptotic cell death was enhanced in DAOY cells expressing high levels of c-MYC. The response of 62 of 66 residual tumors was evaluable and response to postoperative radio- (14 responders (CR, PR) vs. 5 non-responders (SD, PD)) or chemotherapy (23 CR/PR vs. 20 SD/PD) was assessed. c-MYC mRNA expression was similar in primary MB samples of responders and non-responders (Mann-Whitney U test, p = 0.50, ratio 0.49, 95% CI 0.008-30.0 and p = 0.67, ratio 1.8, 95% CI 0.14-23.5, respectively). CONCLUSIONS: c-MYC sensitizes MB cells to some anti-cancer treatments in vitro. As we failed to show evidence for such an effect on postoperative residual tumors when analyzed by imaging, additional investigations in xenografts and larger MB cohorts may help to define the exact function of c-MYC in modulating response to treatment

Functional Promoter Polymorphisms Govern Differential Expression of HMG-CoA Reductase Gene in Mouse Models of Essential Hypertension

3-Hydroxy-3-methylglutaryl-coenzyme A [HMG-CoA] reductase gene (Hmgcr) is a susceptibility gene for essential hypertension. Sequencing of the Hmgcr locus in genetically hypertensive BPH (blood pressure high), genetically hypotensive BPL (blood pressure low) and genetically normotensive BPN (blood pressure normal) mice yielded a number of single nucleotide polymorphisms (SNPs). BPH/BPL/BPN Hmgcr promoter-luciferase reporter constructs were generated and transfected into liver HepG2, ovarian CHO, kidney HEK-293 and neuronal N2A cells for functional characterization of the promoter SNPs. The BPH-Hmgcr promoter showed significantly less activity than the BPL-Hmgcr promoter under basal as well as nicotine/cholesterol-treated conditions. This finding was consistent with lower endogenous Hmgcr expression in liver and lower plasma cholesterol in BPH mice. Transfection experiments using 5′-promoter deletion constructs (strategically made to assess the functional significance of each promoter SNP) and computational analysis predicted lower binding affinities of transcription factors c-Fos, n-Myc and Max with the BPH-promoter as compared to the BPL-promoter. Corroboratively, the BPH promoter-luciferase reporter construct co-transfected with expression plasmids of these transcription factors displayed less pronounced augmentation of luciferase activity than the BPL construct, particularly at lower amounts of transcription factor plasmids. Electrophoretic mobility shift assays also showed diminished interactions of the BPH promoter with HepG2 nuclear proteins. Taken together, this study provides mechanistic basis for the differential Hmgcr expression in these mouse models of human essential hypertension and have implications for better understanding the role of this gene in regulation of blood pressure

Pan-cancer analysis of whole genomes

Cancer is driven by genetic change, and the advent of massively parallel sequencing has enabled systematic documentation of this variation at the whole-genome scale(1-3). Here we report the integrative analysis of 2,658 whole-cancer genomes and their matching normal tissues across 38 tumour types from the Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium of the International Cancer Genome Consortium (ICGC) and The Cancer Genome Atlas (TCGA). We describe the generation of the PCAWG resource, facilitated by international data sharing using compute clouds. On average, cancer genomes contained 4-5 driver mutations when combining coding and non-coding genomic elements; however, in around 5% of cases no drivers were identified, suggesting that cancer driver discovery is not yet complete. Chromothripsis, in which many clustered structural variants arise in a single catastrophic event, is frequently an early event in tumour evolution; in acral melanoma, for example, these events precede most somatic point mutations and affect several cancer-associated genes simultaneously. Cancers with abnormal telomere maintenance often originate from tissues with low replicative activity and show several mechanisms of preventing telomere attrition to critical levels. Common and rare germline variants affect patterns of somatic mutation, including point mutations, structural variants and somatic retrotransposition. A collection of papers from the PCAWG Consortium describes non-coding mutations that drive cancer beyond those in the TERT promoter(4); identifies new signatures of mutational processes that cause base substitutions, small insertions and deletions and structural variation(5,6); analyses timings and patterns of tumour evolution(7); describes the diverse transcriptional consequences of somatic mutation on splicing, expression levels, fusion genes and promoter activity(8,9); and evaluates a range of more-specialized features of cancer genomes(8,10-18).Peer reviewe

Search for strongly interacting massive particles generating trackless jets in proton-proton collisions at s = 13 TeV

A search for dark matter in the form of strongly interacting massive particles (SIMPs) using the CMS detector at the LHC is presented. The SIMPs would be produced in pairs that manifest themselves as pairs of jets without tracks. The energy fraction of jets carried by charged particles is used as a key discriminator to suppress efficiently the large multijet background, and the remaining background is estimated directly from data. The search is performed using proton-proton collision data corresponding to an integrated luminosity of 16.1 fb - 1 , collected with the CMS detector in 2016. No significant excess of events is observed above the expected background. For the simplified dark matter model under consideration, SIMPs with masses up to 100 GeV are excluded and further sensitivity is explored towards higher masses