LipidSeq: a next-generation clinical resequencing panel for monogenic dyslipidemias.

We report the design of a targeted resequencing panel for monogenic dyslipidemias, LipidSeq, for the purpose of replacing Sanger sequencing in the clinical detection of dyslipidemia-causing variants. We also evaluate the performance of the LipidSeq approach versus Sanger sequencing in 84 patients with a range of phenotypes including extreme blood lipid concentrations as well as additional dyslipidemias and related metabolic disorders. The panel performs well, with high concordance (95.2%) in samples with known mutations based on Sanger sequencing and a high detection rate (57.9%) of mutations likely to be causative for disease in samples not previously sequenced. Clinical implementation of LipidSeq has the potential to aid in the molecular diagnosis of patients with monogenic dyslipidemias with a high degree of speed and accuracy and at lower cost than either Sanger sequencing or whole exome sequencing. Furthermore, LipidSeq will help to provide a more focused picture of monogenic and polygenic contributors that underlie dyslipidemia while excluding the discovery of incidental pathogenic clinically actionable variants in nonmetabolism-related genes, such as oncogenes, that would otherwise be identified by a whole exome approach, thus minimizing potential ethical issues.

Sortilin: an unusual suspect in cholesterol metabolism: from GWAS identification to in vivo biochemical analyses, sortilin has been identified as a novel mediator of human lipoprotein metabolism.

The concentration of low-density lipoprotein (LDL) cholesterol (C) in plasma is a key determinant of cardiovascular disease risk and human genetic studies have long endeavoured to elucidate the pathways that regulate LDL metabolism. Massive genome-wide association studies (GWASs) of common genetic variation associated with LDL-C in the population have implicated SORT1 in LDL metabolism. Using experimental paradigms and standards appropriate for understanding the mechanisms by which common variants alter phenotypic expression, three recent publications have presented divergent and even contradictory findings. Interestingly, although these reports each linked SORT1 to LDL metabolism, they did not agree on a mechanism to explain the association. Here, we review recent mechanistic studies of SORT1 - the first gene identified by GWAS as a determinant of plasma LDL-C to be evaluated mechanistically.

Lipoprotein(a): more interesting than ever after 50 years.

PURPOSE OF REVIEW: Lipoprotein(a) [Lp(a)] is a risk factor for cardiovascular disease; we highlight the most recent research initiatives that have sought to define Lp(a)-dependent pathogenicity as well as pharmacologic approaches to lowering Lp(a). RECENT FINDINGS: Recent large-scale meta-analyses have confirmed elevated Lp(a) concentrations to be a moderate but consistent prospective coronary heart disease (CHD) risk factor. The Mendelian randomization approach has also associated LPA variants with Lp(a) concentration and CHD risk. Discoveries linking Lp(a) to oxidized phospholipid burden have implicated a proinflammatory role for Lp(a) hinting at a new mechanism underlying the association with CHD risk, which adds to previous atherogenic and thrombogenic mechanisms. Most existing Lp(a)-lowering drug treatments almost always show simultaneous effects on other lipoproteins, making it difficult to assign any clinical outcome specifically to the effects of Lp(a) lowering. Early experiments with antisense oligonucleotides targeting apolipoprotein(a) mRNA seem to indicate the pleiotropic effects of Lp(a) reduction on LDL and HDL in mice. The mechanism linking Lp(a) concentration with concentrations of other blood lipids remains unknown but may provide an insight into Lp(a) metabolism. SUMMARY: Despite the wealth of epidemiologic evidence supporting Lp(a) concentration as a CHD risk factor, the lack of a definitive functional mechanism involving an Lp(a)-dependent pathway in CHD pathogenesis has limited the potential clinical connotation of Lp(a). However, the application of novel technologies to the long-standing mysteries of Lp(a) biology seems to provide the opportunity for expanding our understanding of Lp(a) and its complex role in cardiovascular health.

The case-finding study: A novel community-based research recruitment approach for engaging participants with early cognitive decline.

INTRODUCTION: Innovative recruitment strategies are needed to better engage potential research participants at a preclinical stage of cognitive decline. METHODS: Local newspaper advertisements attracted community-dwelling people ≥55 years with memory concerns, who were interested in research, to self-refer for cognitive assessment and discuss cognitive research involvement. Respondents completed telephone screening and then attended an in-person cognitive screening assessment with a study partner. Case conferencing with a clinician researcher characterized a "clinical suspicion" of the participant's cognitive concern. RESULTS: Of 209 respondents who underwent in-person assessment, 203 participants were classified as having subjective cognitive decline (47%), mild cognitive impairment (44%), or dementia (9%). Thirty percent of participants were enrolled in observational studies or randomized controlled trials. DISCUSSION: Community-based engagement, cognitive screening, and case conferencing effectively combined to identify research participants at risk of cognitive decline and recruited participants into cognitive research studies. Those not recruited continued to be followed up longitudinally.

Targeted exonic sequencing of GWAS loci in the high extremes of the plasma lipids distribution.

OBJECTIVE: Genome-wide association studies (GWAS) for plasma lipid levels have mapped numerous genomic loci, with each region often containing many protein-coding genes. Targeted re-sequencing of exons is a strategy to pinpoint causal variants and genes. METHODS: We performed solution-based hybrid selection of 9008 exons at 939 genes within 95 GWAS loci for plasma lipid levels and sequenced using next-generation sequencing technology individuals with extremely high as well as low to normal levels of low-density lipoprotein cholesterol (LDL-C, n = 311; mean low = 71 mg/dl versus high = 241 mg/dl), triglycerides (TG, n = 308; mean low = 75 mg/dl versus high = 1938 mg/dl), and high-density lipoprotein cholesterol (HDL-C, n = 684; mean low = 32 mg/dl versus high = 102 mg/dl). We identified 15,002 missense, nonsense, or splice site variants with a frequency <5%. We tested whether coding sequence variants, individually or aggregated within a gene, were associated with plasma lipid levels. To replicate findings, we performed sequencing in independent participants (n = 6424). RESULTS: Across discovery and replication sequencing, we found 6 variants with significant associations with plasma lipids. Of these, one was a novel association: p.Ser147Asn variant in APOA4 (14.3% frequency, TG OR = 0.49, P = 7.1 × 10(-4)) with TG. In gene-level association analyses where rare variants within each gene are collapsed, APOC3 (P = 2.1 × 10(-5)) and LDLR (P = 5.0 × 10(-12)) were associated with plasma lipids. CONCLUSIONS: After sequencing genes from 95 GWAS loci in participants with extremely high plasma lipid levels, we identified one new coding variant associated with TG. These results provide insight regarding design of similar sequencing studies with respect to sample size, follow-up, and analysis methodology.

Common low-density lipoprotein receptor p.G116S variant has a large effect on plasma low-density lipoprotein cholesterol in circumpolar inuit populations.

BACKGROUND: Inuit are considered to be vulnerable to cardiovascular disease because their lifestyles are becoming more Westernized. During sequence analysis of Inuit individuals at extremes of lipid traits, we identified 2 nonsynonymous variants in low-density lipoprotein receptor (LDLR), namely p.G116S and p.R730W. METHODS AND RESULTS: Genotyping these variants in 3324 Inuit from Alaska, Canada, and Greenland showed they were common, with allele frequencies 10% to 15%. Only p.G116S was associated with dyslipidemia: the increase in LDL cholesterol was 0.54 mmol/L (20.9 mg/dL) per allele (P=5.6×10(-49)), which was >3× larger than the largest effect sizes seen with other common variants in other populations. Carriers of p.G116S had a 3.02-fold increased risk of hypercholesterolemia (95% confidence interval, 2.34-3.90; P=1.7×10(-17)), but did not have classical familial hypercholesterolemia. In vitro, p.G116S showed 60% reduced ligand-binding activity compared with wild-type receptor. In contrast, p.R730W was associated with neither LDL cholesterol level nor altered in vitro activity. CONCLUSIONS: LDLR p.G116S is thus unique: a common dysfunctional variant in Inuit whose large effect on LDL cholesterol may have public health implications.

Genetics 100 for cardiologists: basics of genome-wide association studies.

The spring of 2012 marked the fifth anniversary of the widespread appearance in the biomedical literature of genome-wide association studies (GWAS) of diseases of adulthood. Articles reporting GWAS results now regularly appear in dozens of general medicine and cardiology journals. As of August 2012, more than 1700 published GWAS have reported findings across a range of human diseases. Many of these reported new genetic determinants of cardiovascular disease, including coronary artery disease and its risk factors such as diabetes, dyslipidemia, and hypertension. Though GWAS reports follow a standard format, superficially they can appear intimidating to most nongeneticists, whom we suspect often skip over them. Considering the importance of GWAS in cardiovascular science and medicine, and because they show no signs of fading, it is important for cardiovascular medical personnel and scientists to understand GWAS fundamentals. In this article, we provide a roadmap for the nonexpert reader to navigate through GWAS of cardiovascular disease. We cover the basic essentials needed to understand GWAS: underlying theory, mechanics, analysis and display, interpretation, and relevance. Areas covered include the relationship between GWAS and standard epidemiologic study design, the concepts of DNA sequence variation and linkage disequilibrium, the particular statistical considerations in studies involving many independent variables and large sample sizes, the meaning and interpretation of Manhattan plots, and the biologic and clinical significance of GWAS-based discoveries. We conclude with comments about the limitations of GWAS and about what to look for in the "post-GWAS era."

Genetic determinants of "cognitive impairment, no dementia".

Dementia is a heritable condition with devastating effects on both patients and their caregivers. Studies have identified genetic variants associated with increased susceptibility to late-onset Alzheimer disease (AD)-related dementia; however, no studies have assessed whether genetic variation is associated with the early stages of cognitive decline. Given that cerebrovascular disease is an established mechanism in which chronic ischemia increases susceptibility to dementia, we assessed whether genetic variation associated with either cardio-metabolic or AD-related traits is associated with an early stage of cognitive decline called "cognitive impairment, no dementia" (CIND). We studied 484 CIND patients and 459 cognitively healthy controls selected from the Canadian Study of Health and Aging. We tested for association between ~200,000 genetic variants selected from genes associated with cardio-metabolic traits and CIND status using the Cardio-MetaboChip. We also assessed whether AD-related variants and APOE alleles were associated with CIND status, either individually or as part of a composite genetic risk score. We identified a potential association between the ZNF608/GRAMD3 locus, specifically the rs1439568 polymorphism and CIND status (major allele odds ratio [OR] = 1.51; p = 8.4 × 10(-6)). AD-related variants were not associated with CIND status, however APOE E4 allele frequency was significantly higher in CIND patients versus healthy controls (OR = 1.35; p = 0.044). We identified a potential association between the ZNF608/GRAMD3 locus and CIND status, although AD-related variants were not associated with CIND. Additional replication of this association signal is invited.

Western Database of Lipid Variants (WDLV): a catalogue of genetic variants in monogenic dyslipidemias.

BACKGROUND: Next-generation sequencing (NGS) is nearing routine clinical application, especially for diagnosis of rare monogenic cardiovascular diseases. But NGS uncovers so much variation in an individual genome that filtering steps are required to streamline data management. The first step is to determine whether a potential disease-causing variant has been observed previously in affected patients. METHODS: To facilitate this step for lipid disorders, we developed the Western Database of Lipid Variants (WDLV) of 2776 variants in 24 genes that cause monogenic dyslipoproteinemias, including conditions characterized primarily by either high or low low-density lipoprotein cholesterol, high or low high-density lipoprotein cholesterol, high triglyceride, and some miscellaneous disorders. We incorporated quality-control steps to maximize the likelihood that a listed mutation was disease causing. RESULTS: The details of each mutation found in a dyslipidemia, together with a mutation map of the causative genes, are shown in graphical display items. CONCLUSIONS: WDLV will help clinicians and researchers determine the potential pathogenicity of mutations discovered by DNA sequencing of patients or research participants with lipid disorders.