Protease-Sensitive Pancreatic Lipase Variants Are Associated With Early Onset Chronic Pancreatitis.

OBJECTIVES: Premature activation of the digestive protease trypsin within the pancreatic parenchyma is a critical factor in the pathogenesis of pancreatitis. Alterations in genes that affect intrapancreatic trypsin activity are associated with chronic pancreatitis (CP). Recently, carboxyl ester lipase emerged as a trypsin-independent risk gene. Here, we evaluated pancreatic lipase (PNLIP) as a potential novel susceptibility gene for CP. METHODS: We analyzed all 13 PNLIP exons in 429 nonalcoholic patients with CP and 600 control subjects from Germany, in 632 patients and 957 controls from France, and in 223 patients and 1,070 controls from Japan by DNA sequencing. Additionally, we analyzed selected exons in further 545 patients with CP and 1,849 controls originating from Germany, United States, and India. We assessed the cellular secretion, lipase activity, and proteolytic stability of recombinant PNLIP variants. RESULTS: In the German discovery cohort, 8/429 (1.9%) patients and 2/600 (0.3%) controls carried a PNLIP missense variant (P = 0.02, odds ratio [OR] = 5.7, 95% confidence interval [CI] = 1.1-38.9). Variants detected in patients were prone to proteolytic degradation by trypsin and chymotrypsin. In the French replication cohort, protease-sensitive variants were also enriched in patients with early-onset CP (5/632 [0.8%]) vs controls (1/957 [0.1%]) (P = 0.04, OR = 7.6, 95% CI = 0.9-172.9). In contrast, we detected no protease-sensitive variants in the non-European populations. In the combined European data, protease-sensitive variants were found in 13/1,163 cases (1.1%) and in 3/3,000 controls (0.1%) (OR = 11.3, 95% CI = 3.0-49.9, P < 0.0001). CONCLUSIONS: Our data indicate that protease-sensitive PNLIP variants are novel genetic risk factors for the development of CP.

Genetic association and differential expression of PITX2 with acute appendicitis.

Appendicitis affects 9% of Americans and is the most common diagnosis requiring hospitalization of both children and adults. We performed a genome-wide association study of self-reported appendectomy with 18,773 affected adults and 114,907 unaffected adults of European American ancestry. A significant association with appendectomy was observed at 4q25 near the gene PITX2 (rs2129979, p value = 8.82 × 10-14) and was replicated in an independent sample of Caucasians (59 affected, 607 unaffected; p value = 0.005). Meta-analysis of the associated variant across our two cohorts and cohorts from Iceland and the Netherlands (in which this association had previously been reported) showed strong cumulative evidence of association (OR = 1.12; 95% CI 1.09-1.14; p value = 1.81 × 10-23) and some evidence for effect heterogeneity (p value = 0.03). Eight other loci were identified at suggestive significance in the discovery GWAS. Associations were followed up by measuring gene expression across resected appendices with varying levels of inflammation (N = 75). We measured expression of 27 genes based on physical proximity to the GWAS signals, evidence of being targeted by eQTLs near the signals according to RegulomeDB (score = 1), or both. Four of the 27 genes (including PITX2) showed significant evidence (p values < 0.0033) of differential expression across categories of appendix inflammation. An additional ten genes showed nominal evidence (p value < 0.05) of differential expression, which, together with the significant genes, is more than expected by chance (p value = 6.6 × 10-12). PITX2 impacts morphological development of intestinal tissue, promotes an anti-oxidant response, and its expression correlates with levels of intestinal bacteria and colonic inflammation. Further studies of the role of PITX2 in appendicitis are warranted.

Associating Symptom Phenotype and Genotype in Preeclampsia.

Preeclampsia is a complex genetic disorder with an incompletely understood pathogenesis. Its phenotype may be better elucidated by integrating symptoms. This study aimed to identify symptoms by gestational age and associations with novel preeclampsia candidate genes. Women with a history of preeclampsia recruited from The Preeclampsia Registry completed clinical/demographic, symptom surveys and provided medical records. DNA extracted from saliva was processed with multiplexed assays for eight single-nucleotide polymorphisms (SNPs) selected to tag candidate genes and/or located in symptom susceptibility regions. Groups with versus without symptoms were compared using χ2. Associations between SNPs and symptoms were analyzed as genotype categories and presence/absence of the variant allele. Logistic regression modeling was conducted with exploratory p = .05. In 114 participants, 113 reported at least 1 of the 18 symptoms. Symptoms varied by trimester. Nine symptoms were associated with seven SNPs. Visual disturbances were associated with three SNPs and nausea/vomiting with two SNPs. Modeling adjustment for maternal age and parity resulted in 15 associations between 9 symptoms and 8 SNPs. Medical records demonstrated 100% concordance with self-reported diagnosis and 48% concordance with reported severity. Findings indicated novel symptom-genotype associations in preeclampsia. The small sample was self-selected, but results support future studies including medical records review. When validated, these results may lead to holistic phenotyping of women to characterize subsets of preeclampsia. This approach may optimize health in pregnancy and later life for mothers and offspring through prediction, prevention, and precision nursing care.

TCGA Expedition: A Data Acquisition and Management System for TCGA Data.

BACKGROUND: The Cancer Genome Atlas Project (TCGA) is a National Cancer Institute effort to profile at least 500 cases of 20 different tumor types using genomic platforms and to make these data, both raw and processed, available to all researchers. TCGA data are currently over 1.2 Petabyte in size and include whole genome sequence (WGS), whole exome sequence, methylation, RNA expression, proteomic, and clinical datasets. Publicly accessible TCGA data are released through public portals, but many challenges exist in navigating and using data obtained from these sites. We developed TCGA Expedition to support the research community focused on computational methods for cancer research. Data obtained, versioned, and archived using TCGA Expedition supports command line access at high-performance computing facilities as well as some functionality with third party tools. For a subset of TCGA data collected at University of Pittsburgh, we also re-associate TCGA data with de-identified data from the electronic health records. Here we describe the software as well as the architecture of our repository, methods for loading of TCGA data to multiple platforms, and security and regulatory controls that conform to federal best practices. RESULTS: TCGA Expedition software consists of a set of scripts written in Bash, Python and Java that download, extract, harmonize, version and store all TCGA data and metadata. The software generates a versioned, participant- and sample-centered, local TCGA data directory with metadata structures that directly reference the local data files as well as the original data files. The software supports flexible searches of the data via a web portal, user-centric data tracking tools, and data provenance tools. Using this software, we created a collaborative repository, the Pittsburgh Genome Resource Repository (PGRR) that enabled investigators at our institution to work with all TCGA data formats, and to interrogate these data with analysis pipelines, and associated tools. WGS data are especially challenging for individual investigators to use, due to issues with downloading, storage, and processing; having locally accessible WGS BAM files has proven invaluable. CONCLUSION: Our open-source, freely available TCGA Expedition software can be used to create a local collaborative infrastructure for acquiring, managing, and analyzing TCGA data and other large public datasets.

Genetic Basis of Irritant Susceptibility in Health Care Workers.

OBJECTIVE: The aim of this study was to investigate the association of single nucleotide polymorphisms (SNPs) within genes involved in inflammation, skin barrier integrity, signaling/pattern recognition, and antioxidant defense with irritant susceptibility in a group of health care workers. METHODS: The 536 volunteer subjects were genotyped for selected SNPs and patch tested with three model irritants: sodium lauryl sulfate (SLS), sodium hydroxide (NaOH), and benzalkonium chloride (BKC). Genotyping was performed on genomic DNA using Illumina Goldengate custom panels. RESULTS: The ACACB (rs2268387, rs16934132, rs2284685), NTRK2 (rs10868231), NTRK3 (rs1347424), IL22 (rs1179251), PLAU (rs2227564), EGFR (rs6593202), and FGF2 (rs308439) SNPs showed an association with skin response to tested irritants in different genetic models (all at P < 0.001). Functional annotations identified two SNPs in PLAU (rs2227564) and ACACB (rs2284685) genes with a potential impact on gene regulation. In addition, EGF (rs10029654), EGFR (rs12718939), CXCL12 (rs197452), and VCAM1 (rs3917018) genes showed an association with hand dermatitis (P < 0.005). CONCLUSIONS: The results demonstrate that genetic variations in genes related to inflammation and skin homeostasis can influence responses to irritants and may explain inter-individual variation in the development of subsequent contact dermatitis.

Identification of a New Susceptibility Locus for Systemic Lupus Erythematosus on Chromosome 12 in Individuals of European Ancestry.

OBJECTIVE: Genome-wide association studies (GWAS) in individuals of European ancestry identified a number of systemic lupus erythematosus (SLE) susceptibility loci using earlier versions of high-density genotyping platforms. Followup studies on suggestive GWAS regions using larger samples and more markers identified additional SLE loci in subjects of European descent. This multistage study was undertaken to identify novel SLE loci. METHODS: In stage 1, we conducted a new GWAS of SLE in a North American case-control sample of subjects of European ancestry (n = 1,166) genotyped on Affymetrix Genome-Wide Human SNP Array 6.0. In stage 2, we further investigated top new suggestive GWAS hits by in silico evaluation and meta-analysis using an additional data set of subjects of European descent (>2,500 individuals), followed by replication of top meta-analysis findings in another data set of subjects of European descent (>10,000 individuals) in stage 3. RESULTS: As expected, our GWAS revealed the most significant associations at the major histocompatibility complex locus (6p21), which easily surpassed the genome-wide significance threshold (P < 5 × 10(-8)). Several other SLE signals/loci previously implicated in Caucasians and/or Asians were also confirmed in the stage 1 discovery sample, and the strongest signals were observed at 2q32/STAT4 (P = 3.6 × 10(-7)) and at 8p23/BLK (P = 8.1 × 10(-6)). Stage 2 meta-analyses identified a new genome-wide significant SLE locus at 12q12 (meta P = 3.1 × 10(-8)), which was replicated in stage 3. CONCLUSION: Our multistage study identified and replicated a new SLE locus that warrants further followup in additional studies. Publicly available databases suggest that this newly identified SLE signal falls within a functionally relevant genomic region and near biologically important genes.

Resequencing of the CETP gene in American whites and African blacks: Association of rare and common variants with HDL-cholesterol levels.

BACKGROUND: Cholesteryl ester transfer protein (CETP) plays a crucial role in lipid metabolism. Associations of common CETP variants with variation in plasma lipid levels, and/or CETP mass/activity have been extensively studied and well-documented; however, the effects of uncommon/rare CETP variants on plasma lipid profile remain undefined. Hence, resequencing of the gene in extreme phenotypes and follow-up rare-variant association analyses are essential to fill this gap. OBJECTIVE: To identify common and uncommon/rare variants in the CETP gene by resequencing the entire gene and test the effects of both common and uncommon/rare CETP variants on plasma lipid traits in two genetically distinct populations. METHODS AND RESULTS: The entire CETP gene plus flanking regions were resequenced in 190 individuals comprising 95 non-Hispanic whites (NHWs) and 95 African blacks with extreme HDL-C levels. A total of 279 sequence variants were identified, of which 25 were novel. Selected variants were genotyped in the entire samples of 623 NHWs and 788 African blacks and 184 QC-passed variants were tested in relation to plasma lipid traits by using gene-based, single-site, haplotype and rare variant association analyses (SKAT-O). Two novel and independent associations of rs1968905 and rs289740 with HDL-C were identified in African blacks. Using SKAT-O analysis, we also identified rare variants with minor allele frequency <0.01 to be associated with HDL-C in both NHWs (P=0.024) and African blacks (P=0.009). CONCLUSIONS: Our results point out that in addition to the common CETP variants, rare genetic variants in the CETP gene also contribute to the phenotypic variation of HDL-C in the general population.

Genetic contribution of SCARB1 variants to lipid traits in African Blacks: a candidate gene association study.

BACKGROUND: High-density lipoprotein cholesterol (HDL-C) exerts many anti-atherogenic properties including its role in reverse cholesterol transport (RCT). Scavenger receptor class B member 1 (SCARB1) plays a key role in RCT by selective uptake of HDL cholesteryl esters. We aimed to explore the genetic contribution of SCARB1 to affecting lipid levels in African Blacks from Nigeria. METHODS: We resequenced 13 exons and exon-intron boundaries of SCARB1 in 95 individuals with extreme HDL-C levels using Sanger method. Then, we genotyped 147 selected variants (78 sequence variants, 69 HapMap tagSNPs, and 2 previously reported relevant variants) in the entire sample of 788 African Blacks using either the iPLEX Gold or TaqMan methods. A total of 137 successfully genotyped variants were further evaluated for association with major lipid traits. RESULTS: The initial gene-based analysis demonstrated evidence of association with HDL-C and apolipoprotein A-I (ApoA-I). The follow-up single-site analysis revealed nominal evidence of novel associations of nine common variants with HDL-C and/or ApoA-I (P < 0.05). The strongest association was between rs11057851 and HDL-C (P = 0.0043), which remained significant after controlling for multiple testing using false discovery rate. Rare variant association testing revealed a group of 23 rare variants (frequencies ≤ 1%) associated with HDL-C (P = 0.0478). Haplotype analysis identified four SCARB1 regions associated with HDL-C (global P < 0.05). CONCLUSIONS: To our knowledge, this is the first report of a comprehensive association study of SCARB1 variations with lipid traits in an African Black population. Our results showed the consistent association of SCARB1 variants with HDL-C across various association analyses, supporting the role of SCARB1 in lipoprotein-lipid regulatory mechanism.

Needs Assessment for Research Use of High-Throughput Sequencing at a Large Academic Medical Center.

Next Generation Sequencing (NGS) methods are driving profound changes in biomedical research, with a growing impact on patient care. Many academic medical centers are evaluating potential models to prepare for the rapid increase in NGS information needs. This study sought to investigate (1) how and where sequencing data is generated and analyzed, (2) research objectives and goals for NGS, (3) workforce capacity and unmet needs, (4) storage capacity and unmet needs, (5) available and anticipated funding resources, and (6) future challenges. As a precursor to informed decision making at our institution, we undertook a systematic needs assessment of investigators using survey methods. We recruited 331 investigators from over 60 departments and divisions at the University of Pittsburgh Schools of Health Sciences and had 140 respondents, or a 42% response rate. Results suggest that both sequencing and analysis bottlenecks currently exist. Significant educational needs were identified, including both investigator-focused needs, such as selection of NGS methods suitable for specific research objectives, and program-focused needs, such as support for training an analytic workforce. The absence of centralized infrastructure was identified as an important institutional gap. Key principles for organizations managing this change were formulated based on the survey responses. This needs assessment provides an in-depth case study which may be useful to other academic medical centers as they identify and plan for future needs.

Genome-Wide Association Study and Linkage Analysis of the Healthy Aging Index.

BACKGROUND: The Healthy Aging Index (HAI) is a tool for measuring the extent of health and disease across multiple systems. METHODS: We conducted a genome-wide association study and a genome-wide linkage analysis to map quantitative trait loci associated with the HAI and a modified HAI weighted for mortality risk in 3,140 individuals selected for familial longevity from the Long Life Family Study. The genome-wide association study used the Long Life Family Study as the discovery cohort and individuals from the Cardiovascular Health Study and the Framingham Heart Study as replication cohorts. RESULTS: There were no genome-wide significant findings from the genome-wide association study; however, several single-nucleotide polymorphisms near ZNF704 on chromosome 8q21.13 were suggestively associated with the HAI in the Long Life Family Study (p < 10(-) (6)) and nominally replicated in the Cardiovascular Health Study and Framingham Heart Study. Linkage results revealed significant evidence (log-odds score = 3.36) for a quantitative trait locus for mortality-optimized HAI in women on chromosome 9p24-p23. However, results of fine-mapping studies did not implicate any specific candidate genes within this region of interest. CONCLUSIONS: ZNF704 may be a potential candidate gene for studies of the genetic underpinnings of longevity.

Resequencing of LPL in African Blacks and associations with lipoprotein-lipid levels.

Genome-wide association studies have identified several loci associated with plasma lipid levels but those common variants together account only for a small proportion of the genetic variance of lipid traits. It has been hypothesized that the remaining heritability may partly be explained by rare variants with strong effect sizes. Here, we have comprehensively investigated the associations of both common and uncommon/rare variants in the lipoprotein lipase (LPL) gene in relation to plasma lipoprotein-lipid levels in African Blacks (ABs). For variant discovery purposes, the entire LPL gene and flanking regions were resequenced in 95 ABs with extreme high-density lipoprotein cholesterol (HDL-C) levels. A total of 308 variants were identified, of which 64 were novel. Selected common tagSNPs and uncommon/rare variants were genotyped in the entire sample (n=788), and 126 QC-passed variants were evaluated for their associations with lipoprotein-lipid levels by using single-site, haplotype and rare variant (SKAT-O) association analyses. We found eight not highly correlated (r(2)<0.40) signals (rs1801177:G>A, rs8176337:G>C, rs74304285:G>A, rs252:delA, rs316:C>A, rs329:A>G, rs12679834:T>C, and rs4921684:C>T) nominally (P<0.05) associated with lipid traits (HDL-C, LDL-C, ApoA1 or ApoB levels) in our sample. The most significant SNP, rs252:delA, represented a novel association observed with LDL-C (P=0.002) and ApoB (P=0.012). For TG and LDL-C, the haplotype analysis was more informative than the single-site analysis. The SKAT-O analysis revealed that the bin (group) containing 22 rare variants with MAF≤0.01 exhibited nominal association with TG (P=0.039) and LDL-C (P=0.027). Our study indicates that both common and uncommon/rare LPL variants/haplotypes may affect plasma lipoprotein-lipid levels in general African population.

Comprehensive evaluation of the association of APOE genetic variation with plasma lipoprotein traits in U.S. whites and African blacks.

Although common APOE genetic variation has a major influence on plasma LDL-cholesterol, its role in affecting HDL-cholesterol and triglycerides is not well established. Recent genome-wide association studies suggest that APOE also affects plasma variation in HDL-cholesterol and triglycerides. It is thus important to resequence the APOE gene to identify both common and uncommon variants that affect plasma lipid profile. Here, we have sequenced the APOE gene in 190 subjects with extreme HDL-cholesterol levels selected from two well-defined epidemiological samples of U.S. non-Hispanic Whites (NHWs) and African Blacks followed by genotyping of identified variants in the entire datasets (623 NHWs, 788 African Blacks) and association analyses with major lipid traits. We identified a total of 40 sequence variants, of which 10 are novel. A total of 32 variants, including common tagSNPs (≥5% frequency) and all uncommon variants (<5% frequency) were successfully genotyped and considered for genotype-phenotype associations. Other than the established associations of APOE*2 and APOE*4 with LDL-cholesterol, we have identified additional independent associations with LDL-cholesterol. We have also identified multiple associations of uncommon and common APOE variants with HDL-cholesterol and triglycerides. Our comprehensive sequencing and genotype-phenotype analyses indicate that APOE genetic variation impacts HDL-cholesterol and triglycerides in addition to affecting LDL-cholesterol.

A rare duplication on chromosome 16p11.2 is identified in patients with psychosis in Alzheimer's disease.

Epidemiological and genetic studies suggest that schizophrenia and autism may share genetic links. Besides common single nucleotide polymorphisms, recent data suggest that some rare copy number variants (CNVs) are risk factors for both disorders. Because we have previously found that schizophrenia and psychosis in Alzheimer's disease (AD+P) share some genetic risk, we investigated whether CNVs reported in schizophrenia and autism are also linked to AD+P. We searched for CNVs associated with AD+P in 7 recurrent CNV regions that have been previously identified across autism and schizophrenia, using the Illumina HumanOmni1-Quad BeadChip. A chromosome 16p11.2 duplication CNV (chr16: 29,554,843-30,105,652) was identified in 2 of 440 AD+P subjects, but not in 136 AD subjects without psychosis, or in 593 AD subjects with intermediate psychosis status, or in 855 non-AD individuals. The frequency of this duplication CNV in AD+P (0.46%) was similar to that reported previously in schizophrenia (0.46%). This duplication CNV was further validated using the NanoString nCounter CNV Custom CodeSets. The 16p11.2 duplication has been associated with developmental delay, intellectual disability, behavioral problems, autism, schizophrenia (SCZ), and bipolar disorder. These two AD+P patients had no personal of, nor any identified family history of, SCZ, bipolar disorder and autism. To the best of our knowledge, our case report is the first suggestion that 16p11.2 duplication is also linked to AD+P. Although rare, this CNV may have an important role in the development of psychosis.

Impact of genetic variants in human scavenger receptor class B type I (SCARB1) on plasma lipid traits.

BACKGROUND: Scavenger receptor class B type 1 (SCARB1) plays an important role in high-density lipoprotein cholesterol (HDL-C) metabolism in selective cholesteryl ester uptake and in free cholesterol cellular efflux. METHODS AND RESULTS: This study aims to identify common (minor allele frequency ≥5%) and low-frequency/rare (minor allele frequency <5%) variants, using resequencing all 13 exons and exon-intron boundaries of SCARB1 in 95 individuals with extreme HDL-C levels selected from a population-based sample of 623 US non-Hispanic whites. The sequencing step identified 44 variants, of which 11 were novel with minor allele frequency <1%. Seventy-six variants (40 sequence variants, 32 common HapMap tag single nucleotide polymorphisms, and 4 relevant variants) were selected for genotyping in the total sample of 623 subjects followed by association analyses with lipid traits. Seven variants were nominally associated with apolipoprotein B (apoB; n=4) or HDL-C (n=3; P<0.05). Three variants associated with apoB remained significant after controlling false discovery rate. The most significant association was observed between rs4765615 and apoB (P=0.0059), while rs11057844 showed the strongest association with HDL-C (P=0.0035). A set of 17 rare variants (minor allele frequency ≤1%) showed significant association with apoB (P=0.0284). Haplotype analysis revealed 4 regions significantly associated with either apoB or HDL-C. CONCLUSIONS: Our findings provide new information about the genetic role of SCARB1 in affecting plasma apoB levels in addition to its established role in HDL-C metabolism.

Effects of multiple genetic loci on age at onset in late-onset Alzheimer disease: a genome-wide association study.

IMPORTANCE: Because APOE locus variants contribute to risk of late-onset Alzheimer disease (LOAD) and to differences in age at onset (AAO), it is important to know whether other established LOAD risk loci also affect AAO in affected participants. OBJECTIVES: To investigate the effects of known Alzheimer disease risk loci in modifying AAO and to estimate their cumulative effect on AAO variation using data from genome-wide association studies in the Alzheimer Disease Genetics Consortium. DESIGN, SETTING, AND PARTICIPANTS: The Alzheimer Disease Genetics Consortium comprises 14 case-control, prospective, and family-based data sets with data on 9162 participants of white race/ethnicity with Alzheimer disease occurring after age 60 years who also had complete AAO information, gathered between 1989 and 2011 at multiple sites by participating studies. Data on genotyped or imputed single-nucleotide polymorphisms most significantly associated with risk at 10 confirmed LOAD loci were examined in linear modeling of AAO, and individual data set results were combined using a random-effects, inverse variance-weighted meta-analysis approach to determine whether they contribute to variation in AAO. Aggregate effects of all risk loci on AAO were examined in a burden analysis using genotype scores weighted by risk effect sizes. MAIN OUTCOMES AND MEASURES: Age at disease onset abstracted from medical records among participants with LOAD diagnosed per standard criteria. RESULTS: Analysis confirmed the association of APOE with earlier AAO (P = 3.3 × 10(-96)), with associations in CR1 (rs6701713, P = 7.2 × 10(-4)), BIN1 (rs7561528, P = 4.8 × 10(-4)), and PICALM (rs561655, P = 2.2 × 10(-3)) reaching statistical significance (P < .005). Risk alleles individually reduced AAO by 3 to 6 months. Burden analyses demonstrated that APOE contributes to 3.7% of the variation in AAO (R(2) = 0.256) over baseline (R(2) = 0.221), whereas the other 9 loci together contribute to 2.2% of the variation (R(2) = 0.242). CONCLUSIONS AND RELEVANCE: We confirmed an association of APOE (OMIM 107741) variants with AAO among affected participants with LOAD and observed novel associations of CR1 (OMIM 120620), BIN1 (OMIM 601248), and PICALM (OMIM 603025) with AAO. In contrast to earlier hypothetical modeling, we show that the combined effects of Alzheimer disease risk variants on AAO are on the scale of, but do not exceed, the APOE effect. While the aggregate effects of risk loci on AAO may be significant, additional genetic contributions to AAO are individually likely to be small.

Mechanisms of CFTR functional variants that impair regulated bicarbonate permeation and increase risk for pancreatitis but not for cystic fibrosis.

CFTR is a dynamically regulated anion channel. Intracellular WNK1-SPAK activation causes CFTR to change permeability and conductance characteristics from a chloride-preferring to bicarbonate-preferring channel through unknown mechanisms. Two severe CFTR mutations (CFTRsev) cause complete loss of CFTR function and result in cystic fibrosis (CF), a severe genetic disorder affecting sweat glands, nasal sinuses, lungs, pancreas, liver, intestines, and male reproductive system. We hypothesize that those CFTR mutations that disrupt the WNK1-SPAK activation mechanisms cause a selective, bicarbonate defect in channel function (CFTRBD) affecting organs that utilize CFTR for bicarbonate secretion (e.g. the pancreas, nasal sinus, vas deferens) but do not cause typical CF. To understand the structural and functional requirements of the CFTR bicarbonate-preferring channel, we (a) screened 984 well-phenotyped pancreatitis cases for candidate CFTRBD mutations from among 81 previously described CFTR variants; (b) conducted electrophysiology studies on clones of variants found in pancreatitis but not CF; (c) computationally constructed a new, complete structural model of CFTR for molecular dynamics simulation of wild-type and mutant variants; and (d) tested the newly defined CFTRBD variants for disease in non-pancreas organs utilizing CFTR for bicarbonate secretion. Nine variants (CFTR R74Q, R75Q, R117H, R170H, L967S, L997F, D1152H, S1235R, and D1270N) not associated with typical CF were associated with pancreatitis (OR 1.5, p = 0.002). Clones expressed in HEK 293T cells had normal chloride but not bicarbonate permeability and conductance with WNK1-SPAK activation. Molecular dynamics simulations suggest physical restriction of the CFTR channel and altered dynamic channel regulation. Comparing pancreatitis patients and controls, CFTRBD increased risk for rhinosinusitis (OR 2.3, p<0.005) and male infertility (OR 395, p<<0.0001). WNK1-SPAK pathway-activated increases in CFTR bicarbonate permeability are altered by CFTRBD variants through multiple mechanisms. CFTRBD variants are associated with clinically significant disorders of the pancreas, sinuses, and male reproductive system.

Connecting the dots: potential of data integration to identify regulatory SNPs in late-onset Alzheimer's disease GWAS findings.

Late-onset Alzheimer's disease (LOAD) is a multifactorial disorder with over twenty loci associated with disease risk. Given the number of genome-wide significant variants that fall outside of coding regions, it is possible that some of these variants alter some function of gene expression rather than tagging coding variants that alter protein structure and/or function. RegulomeDB is a database that annotates regulatory functions of genetic variants. In this study, we utilized RegulomeDB to investigate potential regulatory functions of lead single nucleotide polymorphisms (SNPs) identified in five genome-wide association studies (GWAS) of risk and age-at onset (AAO) of LOAD, as well as SNPs in LD (r2≥0.80) with the lead GWAS SNPs. Of a total 614 SNPs examined, 394 returned RegulomeDB scores of 1-6. Of those 394 variants, 34 showed strong evidence of regulatory function (RegulomeDB score <3), and only 3 of them were genome-wide significant SNPs (ZCWPW1/rs1476679, CLU/rs1532278 and ABCA7/rs3764650). This study further supports the assumption that some of the non-coding GWAS SNPs are true associations rather than tagged associations and demonstrates the application of RegulomeDB to GWAS data.

The application of network label propagation to rank biomarkers in genome-wide Alzheimer's data.

BACKGROUND: Ranking and identifying biomarkers that are associated with disease from genome-wide measurements holds significant promise for understanding the genetic basis of common diseases. The large number of single nucleotide polymorphisms (SNPs) in genome-wide studies (GWAS), however, makes this task computationally challenging when the ranking is to be done in a multivariate fashion. This paper evaluates the performance of a multivariate graph-based method called label propagation (LP) that efficiently ranks SNPs in genome-wide data. RESULTS: The performance of LP was evaluated on a synthetic dataset and two late onset Alzheimer's disease (LOAD) genome-wide datasets, and the performance was compared to that of three control methods. The control methods included chi squared, which is a commonly used univariate method, as well as a Relief method called SWRF and a sparse logistic regression (SLR) method, which are both multivariate ranking methods. Performance was measured by evaluating the top-ranked SNPs in terms of classification performance, reproducibility between the two datasets, and prior evidence of being associated with LOAD.On the synthetic data LP performed comparably to the control methods. On GWAS data, LP performed significantly better than chi squared and SWRF in classification performance in the range from 10 to 1000 top-ranked SNPs for both datasets, and not significantly different from SLR. LP also had greater ranking reproducibility than chi squared, SWRF, and SLR. Among the 25 top-ranked SNPs that were identified by LP, there were 14 SNPs in one dataset that had evidence in the literature of being associated with LOAD, and 10 SNPs in the other, which was higher than for the other methods. CONCLUSION: LP performed considerably better in ranking SNPs in two high-dimensional genome-wide datasets when compared to three control methods. It had better performance in the evaluation measures we used, and is computationally efficient to be applied practically to data from genome-wide studies. These results provide support for including LP in the methods that are used to rank SNPs in genome-wide datasets.

Parental attitudes toward newborn screening for Duchenne/Becker muscular dystrophy and spinal muscular atrophy.

INTRODUCTION: Disease inclusion in the newborn screening (NBS) panel should consider the opinions of those most affected by the outcome of screening. We assessed the level and factors that affect parent attitudes regarding NBS panel inclusion of Duchenne muscular dystrophy (DMD), Becker muscular dystrophy (BMD), and spinal muscular atrophy (SMA). METHODS: The attitudes toward NBS for DMD, BMD, and SMA were surveyed and compared for 2 categories of parents, those with children affected with DMD, BMD, or SMA and expectant parents unselected for known family medical history. RESULTS: The level of support for NBS for DMD, BMD, and SMA was 95.9% among parents of children with DMD, BMD, or SMA and 92.6% among expectant parents. CONCLUSIONS: There was strong support for NBS for DMD, BMD, and SMA in both groups of parents. Given advances in diagnostics and promising therapeutic approaches, discussion of inclusion in NBS should continue.

Lipoprotein lipase gene sequencing and plasma lipid profile.

Lipoprotein lipase (LPL) plays a crucial role in lipid metabolism by hydrolyzing triglyceride (TG)-rich particles and affecting HDL cholesterol (HDL-C) levels. In this study, the entire LPL gene plus flanking regions were resequenced in individuals with extreme HDL-C/TG levels (n = 95), selected from a population-based sample of 623 US non-Hispanic White (NHW) individuals. A total of 176 sequencing variants were identified, including 28 novel variants. A subset of 64 variants [common tag single nucleotide polymorphisms (tagSNP) and selected rare variants] were genotyped in the total sample, followed by association analyses with major lipid traits. A gene-based association test including all genotyped variants revealed significant association with HDL-C (P = 0.024) and TG (P = 0.006). Our single-site analysis revealed seven independent signals (P < 0.05; r² < 0.40) with either HDL-C or TG. The most significant association was for the SNP rs295 exerting opposite effects on TG and HDL-C levels with P values of 7.5.10⁻4 and 0.002, respectively. Our work highlights some common variants and haplotypes in LPL with significant associations with lipid traits; however, the analysis of rare variants using burden tests and SKAT-O method revealed negligible effects on lipid traits. Comprehensive resequencing of LPL in larger samples is warranted to further test the role of rare variants in affecting plasma lipid levels.