Accurate and consistent variant classification is imperative for incorporation of rapidly developing sequencing technologies into genomic medicine for improved patient care. An essential requirement ...for achieving standardized and reliable variant interpretation is data sharing, facilitated by a centralized open‐source database. Familial hypercholesterolemia (FH) is an exemplar of the utility of such a resource: it has a high incidence, a favorable prognosis with early intervention and treatment, and cascade screening can be offered to families if a causative variant is identified. ClinVar, an NCBI‐funded resource, has become the primary repository for clinically relevant variants in Mendelian disease, including FH. Here, we present the concerted efforts made by the Clinical Genome Resource, through the FH Variant Curation Expert Panel and global FH community, to increase submission of FH‐associated variants into ClinVar. Variant‐level data was categorized by submitter, variant characteristics, classification method, and available supporting data. To further reform interpretation of FH‐associated variants, areas for improvement in variant submissions were identified; these include a need for more detailed submissions and submission of supporting variant‐level data, both retrospectively and prospectively. Collaborating to provide thorough, reliable evidence‐based variant interpretation will ultimately improve the care of FH patients.
Here, we present the recent efforts made by the Clinical Genome Resource consortium, along with various global familial hypercholesterolemia (FH) researchers, to update the number and characterization of FH‐associated variants now present on the ClinVar database. Specifically, we break down the number of FH variants hosted on ClinVar by gene, location, type, and classification; in addition to providing variant‐level characterizations. We then discuss the implications learned from these variant‐level and aggregate results.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SBCE, SBMB, UL, UM, UPUK
We assessed the associations of serum, red blood cell membranes (RBCM) and dietary long-chain n-3 polyunsaturated fatty acids (LC-PUFAs) with neovascular age-related macular degeneration (AMD).
We ...included 290 patients of the Nutritional AMD Treatment 2 Study (NAT2) with neovascular AMD in one eye and early AMD lesions in the other eye, and 144 normal vision controls without AMD. Dietary intake of seafood was estimated by food frequency questionnaire. Eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) composition in serum and RBCM were determined by gas chromatography from 12-hour fasting blood samples and was expressed as percentages of total fatty acids profile. Logistic regressions estimated associations of neovascular AMD with dietary intake of seafood and circulating n-3 LC-PUFAs.
Dietary oily fish and seafood intake were significantly lower in AMD patients than in controls. After adjustment for all potential confounders (age, sex, CFH Y402H, ARMS2 A69S, and ApoE4 polymorphisms, plasma triglycerides, hypertension, hypercholesterolemia, and family history of AMD), serum EPA was associated significantly with a lower risk for neovascular AMD (odds ratio OR = 0.41; 95% confidence interval CI, 0.22-0.77; P = 0.005). Analysis of RBCM revealed that EPA and EPA+DHA were associated significantly with a lower risk for neovascular AMD (OR = 0.25; 95% CI, 0.13-0.47; P < 0.0001 and OR = 0.52; 95% CI, 0.29-0.94; P = 0.03, respectively).
The RBCM EPA and EPA+DHA, as long-term biomarkers of n-3 dietary PUFA status, were associated strongly with neovascular AMD and may represent an objective marker identifying subjects at high risk for neovascular AMD, who may most benefit from nutritional interventions. (http://www.controlled-trials.com/isrctn number, ISRCTN98246501).
Background. Carotenoids and docosahexaenoic acid (DHA) were identified as essential components for eye health and are both naturally present in eggs. Objective. We aimed to evaluate the effect of the ...daily consumption of two eggs enriched with lutein/zeaxanthin and DHA on macular pigment optical density (MPOD) and on circulating xanthophyll and fatty acid concentrations in healthy participants. Methods. Ninety-nine healthy volunteers consumed either two standard eggs or two enriched eggs per day for 4 months. MPOD was measured at baseline (V0) and at follow-up (V4) using a modified confocal scanning laser ophthalmoscope (primary outcome). Blood samples were collected to determine total plasma and lipoprotein fatty acids and lutein/zeaxanthin compositions at V0 and V4 (secondary outcomes). Results. A slight but significant increase in MPOD was observed for all study participants consuming two eggs per day for 4 months at all eccentricities (0.5°, 1°, 2°, and 4°). Plasma and lipoprotein lutein, zeaxanthin, and DHA concentrations significantly increased in both groups but were greater in the enriched group (for the enriched group (V0 vs. V4): lutein, 167 vs. 369 ng/mL; zeaxanthin, 17.7 vs. 29.2 ng/mL; DHA, 1.89 vs. 2.56% of total fatty acids). Interestingly, lutein from high-density lipoprotein (HDL) was strongly correlated with MPOD at 0.5 and 1° eccentricities (rho = 0.385, p = 0.008, and rho = 0.461, p = 0.001, respectively). Conclusions. MPOD was slightly increased in both groups. Lutein, zeaxanthin, and DHA plasma concentrations were strongly enhanced in the enriched group compared with the standard group. A significant correlation was found between MPOD level and lutein concentration in HDL.
Apolipoprotein E (Apo E) is a 36Kda glycoprotein involved in lipid transport. It exists in 3 major isoforms: E2, E3 and E4. ApoE status is known to be a major risk factor for late-onset Alzheimer's ...and cardiovascular diseases. Genotyping is commonly used to obtain ApoE status but can show technical issues with ambiguous determinations. Phenotyping can be an alternative, not requiring genetic material. We evaluated the ability to accurately type ApoE isoforms by 2 phenotyping tests in comparison with genotyping.
Two phenotyping techniques were used: (1) LC–MS/MS detection of 4 ApoE specific peptides (6490 Agilent triple quadripole): After its denaturation, serum was either reduced and alkylated, or only diluted, and then trypsin digested. Before analysis, desalting, evaporation and resuspension were performed. (2) Isoelectric focusing and immunoprecipitation: serum samples were neuraminidase digested, delipidated and electrophoresed on Hydragel ApoE (Sebia agarose gel) using Hydrasys 2 Scan instrument (Sebia, Lisses, France). ApoE isoforms bands were directly immunofixed in the gel using a polyclonal anti human ApoE antibody. Then, incubation of the gel with HRP secondary antibody followed by TTF1/TTF2 substrate allowed the visualization of ApoE bands. The results of the two techniques were compared to genotyping.
Sera from 35 patients previously genotyped were analyzed with the 2 phenotyping techniques. 100% concordance between both phenotyping assays was obtained for the tested phenotypes (E2/E2, E2/E3, E2/E4, E3/E3, E3/E4, E4/E4). When compared to genotyping, 3 samples were discordant. After reanalyzing them by both phenotyping tests and DNA sequencing, 2/3 discrepancies were confirmed. Those can be explained by variants or rare ApoE alleles or by unidentified technical issues. 102 additional samples were then tested on LC–MS/MS only and compared to genotyping. The data showed 100% concordance.
Our 2 phenotyping methods represent a valuable alternative to genotyping. LC–MS/MS has the advantage of being fully specific, with identification of the different isoforms and can be considered as a reference method. Sebia isofocusing technique was concordant with LC–MS/MS. Plus, it is a rapid, semi-automated assay that can be easily implemented in clinical laboratories.
•A comparison of two complementary techniques (quantitative mass spectrometry and isofocusing) for the phenotyping of Apolipoprotein E is proposed.•Apolipoprotein E known to be a major risk factor for late-onset Alzheimer's and cardiovascular diseases•These 2 methods allow the of use small volume (2 to 20μL) of residual blood from samples collected for routine clinical tests, thus enabling retrospective studies with preserved body fluids.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK, ZRSKP
OBJECTIVE—Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a central player in the regulation of cholesterol homeostasis, increasing the low-density lipoprotein (LDL) receptor degradation. ...Our study aimed at exploring the pathogenic consequences in vivo and in vitro of a PCSK9 prodomain mutation found in a family with hypobetalipoproteinemia (FHBL).
METHODS AND RESULTS—A white 49-year-old diabetic man had profound FBHL (LDLC16 mg/dL) whereas his daughter and sister displayed a milder phenotype (LDLC 44 mg/dL and 57 mg/dL, respectively), all otherwise healthy with a normal liver function. A monoallelic PCSK9 double-mutant R104C/V114A cosegregated with FBHL, with no mutation found at other FHBL-causing loci. A dose-effect was also found in FBHL relatives for plasma APOB and PCSK9 (very-low to undetectable in proband, ≈50% decreased in sister and daughter) and LDL catabolic rate (256% and 88% increased in proband and daughter). Transient transfection in hepatocytes showed severely impaired processing and secretion of the double mutant which acted as a dominant negative over secretion of wild-type PCSK9.
CONCLUSION—These results show that heterozygous PCSK9 missense mutations may associate with profound hypobetalipoproteinemia and constitute the first direct evidence in human that decrease of plasma LDLC concentrations associated to PCSK9 LOF mutations are attributable to an increased clearance rate of LDL.
Genetic susceptibility could be modified by environmental factors and may also influence differential responses to treatments for age-related macular degeneration (AMD). We investigated whether ...genotype could influence response to docosahexaenoic acid (DHA)-supplementation in the occurrence of choroidal new vessels (CNV).
The Nutritional AMD Treatment 2 (NAT2) study was a randomized, placebo-controlled, double-blind, parallel, comparative study, including 250 patients aged 55 to 85 years with early lesions of age-related maculopathy, visual acuity better than 0.4 Logarithm of Minimum Angle of Resolution units in the study eye and neovascular AMD in the fellow eye. Patients were randomized at baseline to receive either 3 daily fish-oil capsules, each containing 280 mg DHA, 90 mg EPA and 2 mg Vitamin E, or placebo.
Patients carrying the risk allele (C) for CFH Y402H had no statistically significant increased risk for developing CNV in the study eye (Hazard Ratio (HR)=0.97; 95% Confidence Interval (CI): 0.54-1.76 for heterozygous and HR=1.29; 95%CI: 0.69-2.40 for homozygous). Patients carrying the risk allele (T) for ARMS2 A69S had no statistically significant increased risk for developing CNV in the study eye (HR=1.68; 95%CI: 0.91-3.12) for heterozygous and HR=1.78; 95%CI: 0.90-3.52 for homozygous). A significant interaction was observed between CFH Y402H and DHA-supplementation (p=0.01). We showed a protective effect of DHA-supplementation among homozygous non-risk patients. Among these patients, occurrence of CNV was 38.2% in placebo group versus 16.7% in DHA group (p=0.008).
These results suggest that a genetic predisposition to AMD conferred by the CFH Y402H variant limits the benefit provided by DHA supplementation.
ISRCTN registry 98246501.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
To evaluate the dynamic remodeling of drusen in subjects with unilateral neovascular age-related macular degeneration (AMD) receiving a three-year course of oral docosahexaenoic acid (DHA) or ...placebo.
Institutional setting.
Three hundred subjects with age-related maculopathy and neovascular AMD in the fellow eye were randomly assigned to receive either 840 mg/day DHA or placebo for 3 years. Main outcome measures of this post-hoc sub-group analysis were progression of drusen number, total diameter, and total area on fundus photography, and their association with DHA supplementation, socio-demographic and genetic characteristics.
Drusen progression was analyzed in 167 subjects that did not develop CNV (87 that received DHA and 80 that received placebo). None of the drusen remodeling outcomes were significantly associated with DHA supplementation. Total drusen diameter reduction in the inner subfield was significantly associated with age (older patients: r = -0.17; p = 0.003). Women showed a tendency to decreased total drusen diameter in the inner subfield with CFH polymorphism (p = 0.03), where women with TT genotype tended to have a greater reduction in drusen diameter than other genotypes (CC and CT). Drusen area in the inner subfield was more reduced in older patients (r = -0.17) and in women (p = 0.01). Drusen number showed no significant trends.
Dynamic drusen remodeling with net reduction in drusen load over three years was found in patients with exudative AMD in one eye and drusen in the other eye (study-eye). This reduction was correlated with increased age and female gender, and showed a tendency to be influenced by CFH genotype, but did not appear to be affected by DHA supplementation.
Controlled-Trials.com ISRCTN98246501.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Major genetic factors for age-related macular degeneration (AMD) have recently been identified as susceptibility risk factors, including variants in the CFH gene and the ARMS2 LOC387715/HTRA1locus. ...Our purpose was to perform a case-control study in two populations among individuals who did not carry risk variants for CFHY402H and LOC387715 A69S (ARMS2), called "study" individuals, in order to identify new genetic risk factors. Based on a candidate gene approach, we analyzed SNP rs5888 of the SCARB1 gene, coding for SRBI, which is involved in the lipid and lutein pathways. This study was conducted in a French series of 1241 AMD patients and 297 controls, and in a North American series of 1257 patients with advanced AMD and 1732 controls. Among these individuals, we identified 61 French patients, 77 French controls, 85 North American patients and 338 North American controls who did not carry the CFH nor ARMS2 polymorphisms. An association between AMD and the SCARB1 gene was seen among the study subjects. The genotypic distribution of the rs5888 polymorphism was significantly different between cases and controls in the French population (p<0.006). Heterozygosity at the rs5888 SNP increased risk of AMD compared to the CC genotypes in the French study population (odds ratio (OR) = 3.5, CI95%: 1.4-8.9, p<0.01) and after pooling the 2 populations (OR = 2.9, 95% CI: 1.6-5.3, p<0.002). Subgroup analysis in exudative forms of AMD revealed a pooled OR of 3.6 for individuals heterozygous for rs5888 (95% CI: 1.7-7.6, p<0.0015). These results suggest the possible contribution of SCARB1, a new genetic factor in AMD, and implicate a role for cholesterol and antioxidant micronutrient (lutein and vitamin E) metabolism in AMD.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Despite documented benefits of lipid-lowering treatment in women, a considerable number are undertreated, and fewer achieve treatment targets vs. men.
Data were combined from 27 double-blind, active ...or placebo-controlled studies that randomized adult hypercholesterolemic patients to statin or statin+ezetimibe. Consistency of treatment effect among men (n = 11,295) and women (n = 10,499) was assessed and percent of men and women was calculated to evaluate the between-treatment ability to achieve specified treatment levels between sexes.
Baseline lipids and hs-CRP were generally higher in women vs. men. Between-treatment differences were significant for both sexes (all p < 0.001 except apolipoprotein A-I in men = 0.0389). Men treated with ezetimibe+statin experienced significantly greater changes in LDL-C (p = 0.0066), non-HDL-C, total cholesterol, triglycerides, HDL-C, apolipoprotein A-I (all p < 0.0001) and apolipoprotein B (p = 0.0055) compared with women treated with ezetimibe+statin. The odds of achieving LDL-C < 100 mg/dL, apolipoprotein B < 90 mg/dL and the dual target LDL-C < 100 mg/dL & apoliprotein B < 90 mg/dL was significantly greater for women vs. men and the odds of achieving hs-CRP < 1 and < 2 mg/L and dual specified levels of LDL-C < 100 mg/dL and hs-CRP < 2 mg/L were significantly greater for men vs. women. Women reported significantly more gall-bladder-related, gastrointestinal-related, and allergic reaction or rash-related adverse events (AEs) vs. men (no differences between treatments). Men reported significantly more CK elevations (no differences between treatments) and hepatitis-related AEs vs. women (significantly more with ezetimibe+simvastatin vs. statin).
These results suggest that small sex-related differences may exist in response to lipid-lowering treatment and achievement of specified lipid and hs-CRP levels, which may have implications when managing hypercholesterolemia in women.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
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