Familial Hypercholesterolemia Mutations Research Articles

Clinical and genetic diagnosis of familial hypercholesterolaemia in patients undergoing coronary angiography: the Ludwigshafen Risk and Cardiovascular Health Study.

To investigate the prevalence of familial hypercholesterolaemia (FH) and compare the performance of clinical criteria and genetic testing in patients undergoing coronary angiography. The prevalence of FH was determined with the Dutch Lipid Clinical Network (DLCN), US-MEDPED, Simon Broome (SB) criteria, the 'Familial Hypercholesterolaemia Case Ascertainment Tool' (FAMCAT), and a clinical algorithm. Genetic screening was conducted with a custom array from Affymetrix (CARRENAL array) harboring 944 FH mutations. The study cohort consisted of 3267 patients (78.6% with coronary artery disease [CAD]). FH was diagnosed in 2.8%, 2.2%, 3.9%, and 7.9% using the DLCN, US-MEDPED, SB criteria, and the FAMCAT. The clinical algorithm identified the same patients as the SB criteria. Pathogenic FH mutations were found in 1.2% (1.2% in patients with CAD, 1.0% in patients without CAD). FH was more frequently diagnosed in younger patients. With genetic testing as reference, the clinical criteria achieved areas under the ROC curve (AUCs) in the range of 0.56-0.68. Using only LDL-C corrected for statin intake, an AUC of 0.68 was achieved. FH is up to fourfold more prevalent in patients undergoing coronary angiography than in contemporary cohorts representing the general population. Different clinical criteria yield substantially different diagnosis rates, overestimating the prevalence of FH compared to genetic testing. LDL-C testing alone may be sufficient to raise the suspicion of FH, which then needs to be corroborated by genetic testing.

CTCA in children with severe heterozygous familial hypercholesterolaemia: Screening for subclinical atherosclerosis

Familial hypercholesterolemia (FH) is one of the most common genetically inherited disorders in the world. Children with severe heterozygous FH (HeFH), i.e. untreated low-density lipoprotein cholesterol (LDL-C) levels above the 90th percentile for age and sex among FH mutation carriers, can have LDL-C levels that overlap levels of children with homozygous FH (HoFH), but treatment regimen and cardiovascular follow-up to prevent cardiovascular disease are less intensive in children with severe HeFH. In children with HoFH, subclinical atherosclerosis can already be present using computed tomography coronary angiography (CTCA). The question remains whether this is also the case in children with severe HeFH who have a high exposure to elevated LDL-C levels from birth onwards as well. We calculated the cumulative LDL-C exposure (CEtotal [mmol]) in four children with severe HeFH and performed computed tomography coronary angiography (CTCA). These children, aged 13, 14, 15 and 18 years, had CEtotal of 71.3, 97.8, 103.6 and 136.1 mmol, respectively. None of them showed abnormalities on cardiovascular imaging, despite high LDL-C exposure. The results of this study, do not give us an indication to recommend performing CTCA routinely in children with severe HeFH.

Aortic Wall Thickness as a Surrogate for Subclinical Atherosclerosis in Familial and Nonfamilial Hypercholesterolemia: Quantitative 3D Magnetic Resonance Imaging Study and Interrelations with Computed Tomography Calcium Scores, and Carotid Ultrasonography.

We aimed to compare the extent of subclinical atherosclerosis in the ascending and descending aortas by measuring wall area and thickness using 3D cardiovascular magnetic resonance imaging (aAWAI and dAWAI) in patients with asymptomatic familial hypercholesterolemia (FH) and nonfamilial hypercholesterolemia (NFH). We also aimed to establish the interrelations of CMR parameters with other subclinical atherosclerosis measurements, such as calcium scores, obtained using computed tomography in coronary arteries (CCS) and ascending and descending aorta (TCSasc and TCSdsc), as well as the carotid intima-media thicknesses (cIMT) using ultrasonography. A total of 60 patients with FH (29 men and 31 women), with a mean age of 52.3 ± 9.6 years, were analyzed. A subclinical atherosclerosis assessment was also performed on a group consisting of 30 age- and gender-matched patients with NFH, with a mean age of 52.5 ± 7.9 years. We found the ascending and descending aortic wall areas and thicknesses in the FH group to be significantly increased than those of the NFH group. A multivariate logistic regression analysis showed that a positive FH mutation value was a strong predictor of high aAWAI and dAWAI independent of the LDL cholesterol level. Correlations across CMR atherosclerotic parameters, calcium scores, and cIMT in the FH and NFH groups, were significant but low. Most of the atherosclerosis tests with high results belonged to the FH group. We found that patients with documented heterozygous FH had a higher atherosclerosis burden in the aorta compared to patients with severe hypercholesterolemia without FH gene mutation. Atherosclerosis is not severe in asymptomatic patients with FH, but is more pronounced and also more diffuse than in patients with NFH. The etiology of hypercholesterolemia, and not just cholesterol levels, plays a significant role in determining the degree of subclinical atherosclerosis.

Including coronary artery calcium scores to guide treatment in a specialist lipid clinic setting

Abstract Funding Acknowledgements Type of funding sources: None. Background The Multi-Ethnic Study of Atherosclerosis demonstrated that coronary artery calcium (CAC) scoring has proven to be an effective tool at guiding lipid lowering therapy in patients with dyslipidaemia who are at risk of atherosclerotic cardiovascular disease (ASCVD) in the setting of a cohort study. The 2019 joint ESC/EAS dyslipidaemia guidelines identify a number of risk modifiers to stratify ASCVD risk from low to very-high including Lipoprotein (a) [Lp(a)] and the identification of familial hypercholesterolaemia (FH) mutations. Purpose 1) To determine whether CAC scoring can improve the risk assessment of ASCVD in a lipid clinic setting and therefore act as a more effective guide for clinicians when implementing lipid lowering therapy. 2) To compare CAC scores in patients grouped by Lp(a) levels and FH mutations. Methods A retrospective cohort analysis of patients attending a specialist lipid clinic from July to December 2021 in a tertiary referral centre. Data was collected in July/August 2022. Patients were divided into two groups, those with a CAC score of 0 and greater than 0. Data collected included patient risk factors and modifiers as described in the 2019 joint ESC/EAS dyslipidaemia guidelines, serum Lp(a) levels and FH mutation status. Statistical analysis with SPSS software applied the Mann-Whitney and Chi Squared test. Results 49.4% patients (n=41) had a CAC score of greater than 0. Of the patients classified as very-high risk, 91.3% (n=21) had a CAC score of greater than 0. For those classified as high risk, 26.5% (n=9) had a CAC score greater than 0. The mean Lp(a) plasma level was 124nmol/L for a CAC score of 0, compared to 249nmol/L for a CAC score of greater than 0 (U=562,p=0.02). 23 patients had FH mutations, of which 47.8% (n=11) had CAC scores of greater than 0 (χ2 = 2.15, p= 0.340). Conclusion This initial analysis describes CAC scores in patients attending a specialist lipid clinic classified by ASCVD risk and risk modifiers. It suggests CAC scores have a role, either additional or confirmatory, in ASCVD risk assessment. Further analysis is required to include additional risk modifiers, expand the cohort and preform a more granular stratification of the group by CAC score.

Impact of Healthy Lifestyle in Patients With Familial Hypercholesterolemia.

Pathogenic mutations are associated with poor outcomes in patients with familial hypercholesterolemia (FH). However, data on the effects of a healthy lifestyle on FH phenotypes are limited. The authors investigated the interaction between a healthy lifestyle and FH mutation with prognosis in patients with FH. We investigated the associations of the interaction between genotypes and lifestyle, with the occurrence of major adverse cardiac events (MACE), such as cardiovascular-related mortality, myocardial infarction, unstable angina, and coronary artery revascularization, in patients with FH. We assessed their lifestyle based on 4 questionnaires (healthy dietary pattern, regular exercise, not smoking, and absence of obesity). The Cox proportional hazards model was used to assess the risk for MACE. The median follow-up duration was 12.6 (IQR: 9.5-17.9) years. During the follow-up duration, 179 MACE were observed. Independent of classic risk factors, FH mutation and lifestyle score were significantly associated with MACE (HR: 2.73; 95%CI: 1.03-4.43; P = 0.02; and HR: 0.69, 95%CI: 0.40-0.98, P = 0.033, respectively). The estimated risk of coronary artery disease by 75 years of age varied according to lifestyle, ranging from 21.0% among noncarriers with a favorable lifestyle to 32.1% among noncarriers with an unfavorable lifestyle and ranging from 29.0% among carriers with a favorable lifestyle to 55.4% among carriers with an unfavorable lifestyle. A healthy lifestyle was associated with reduced risk for MACE among patients with FH with or without genetic diagnosis.

Subclinical atherosclerosis determined by coronary artery calcium deposition in patients with clinical familial hypercholesterolemia

Background and aimsLimited knowledge exists regarding the association between coronary artery calcium (CAC) deposition in patients with clinical familial hypercholesterolemia (FH) and FH subtypes such as polygenic causes. We studied CAC score in patients with clinical FH and subtypes including polygenic causes of FH compared to healthy controls. MethodsIn a case-control study, we identified potential clinical FH cases registered with an LDL-C >6.7 mmol/l within a nationwide clinical laboratory database on the Faroe Islands and invited them for diagnostic evaluation according to clinical FH scoring systems. Controls were identified in the background population. All subjects were aged 18–75 years and without a history of cardiovascular disease. FH mutation testing and genotypes of twelve LDL-C associated single nucleotide polymorphisms were determined using conventional methods in selected individuals. CAC scores were assessed by cardiac CT. Odds ratios obtained using multivariate logistic regression were used as measures of association. ResultsA total of 120 clinical FH patients and 117 age- and sex-matched controls were recruited. We found a very low frequency of monogenic FH (3%), but a high level of polygenic FH (60%) in those genetically tested (54%). There was a statistically significant association between the CAC score and a diagnosis of clinical FH with the highest observed odds ratio of 5.59 (95% CI 1.65; 18.94, p = 0.006) in those with a CAC score ≥300 compared to those with a CAC of zero. In supplemental analyses, there was a strong association between CAC scores and clinical FH of a polygenic cause. ConclusionWe found a statistically significant association between CAC levels and clinical FH with the highest observed risk estimates among clinical FH cases of a presumed polygenic cause.

Expanding the genetic spectrum for Chinese familial hypercholesterolemia population with six genetic mutations identified using a next-generation sequencing-based laboratory-developed screening test.

This study was to reveal the prevalence of definite familial hypercholesterolemia (FH) in the hospital-visiting population, determine the pathogenic mutation detection rate in clinically diagnosed definite FH patients, and expand the FH mutation spectrum in China. Blood lipid profiles of 41,803 patients visiting the hospital were investigated and 4967 patients with clinical diagnoses of other metabolic diseases were excluded. One hundred and seventy-three (0.41%) received a definite diagnosis of FH according to the Dutch Lipid Clinical Network Criteria-Chinese Revised Version (DLCN-CRV), and 18 patients subsequently agreed to undergo genetic testing. A next-generation sequencing (NGS)-based laboratory-developed test covering the exonic regions of 24 lipid metabolism-related genes was conducted alongside in silico analyses to identify possible FH mutations in 16 definite FH patients, according to the American College of Medical Genetics and Genomics (ACMG) criteria. Sanger sequencing was used to confirm mutations, and SWISS-MODEL was used to simulate the molecular structures of the confirmed protein-carrying mutations. The FH prevalence was 0.41% for the 41,803 individuals (DLCN-CRV grade >8) and 25% of definite FH patients carried six FH pathogenic mutations (≥ACMG Class 4). All genetic variants were confirmed by Sanger sequencing. Five pathogenic variants on the LDLR gene (NM_000527: c.C1783T: p.R595W, c.T493G: p.W165G, c.G1879A: p.A627T, c.G682T: p.E228X, and exon10: c.G1432A: p.G478R) and one pathogenic variant on APOB (NM_000384: c.C10579T: p.R3527W) in 25% of the identified definite FH patients. Two pathogenic mutations, c.T493G (p.W165G) and c.C1783T (p.R595W), were added to the current genetic spectrum of FH in China. This study contributes to improving the current FH detection rate and genetic screening strategies; it provides new directions for treatment, management, and drug development.

Impact of variants of uncertain significance of LDL receptor on phenotypes of familial hypercholesterolemia

BackgroundData on the effect of variants of uncertain significance (VUS) of LDL receptor (LDLR) on familial hypercholesterolemia (FH) phenotype is limited. ObjectiveTo investigate the associations between genotypes and phenotypes, including low-density lipoprotein (LDL) cholesterol level and occurrence of major adverse cardiac events (MACEs), in FH patients (N = 1050, male/female = 490/560). MethodsWe retrospectively assessed the data of patients with FH admitted at Kanazawa University Hospital between 1990 and 2020. Based on genotype, the patients were divided into patients without variants, with VUS of LDLR, and with pathogenic variants. Cox proportional hazard model was used to identify the factors associated with MACEs. ResultsThe median follow-up duration was 12.6 years (interquartile range: 9.5–17.9 years). Altogether, 777 patients had FH mutation and 273 had pathogenic mutation, with 92 having VUS. Over the follow-up duration, 175 MACEs were observed. LDL cholesterol level was found to be significantly higher in patients with pathogenic variants (251 mg/dL) than in patients with VUS (225 mg/dL) and without variants (203 mg/dL). Pathogenic variants and VUS are significantly associated with MACEs (hazard ratio [HR] = 1.52, 95% confidence interval [CI] = 1.02–2.02, P = 0.033 and HR = 3.18, 95% CI = 2.00–4.36, P = 1.9 × 10−5, relative to patients without any variants, respectively), independent of classical risk factors. ConclusionVUS of LDLR was significantly associated with poor outcomes in FH patients. Genetic testing is useful for the diagnosis and risk stratification of FH patients.

Prevalence of premature coronary artery disease (CAD) according to genetic familial hypercholesterolaemia (FH) diagnosis in Lithuania

Background and Aims : FH is known to be a predisposing cause of premature CAD. We aimed to determine the distribution of premature CAD according to genetic diagnosis of FH in Lithuania.Methods: Prospective observational cohort study enrolled patients with clinically diagnosed FH according to Dutch Lipid Clinic Network (DLCN) criteria treated in Vilnius University Hospital Santaros Klinikos during the period of 2016-2021. Premature CAD was defined as occuring in men younger than 55 years and women younger than 60 years. Data of 60 study patients were included in the analysis. Obstructive Atherosclerotic CAD was defined as the presence of stenosis ≥50% in at least one coronary vessel in Coronary Computed Tomography Angiography (CCTA) or coronary angiography, as well as performed percutaneous coronary intervention (PCI) or coronary arteries bypass grafting (CABG). Genetic testing was performed using genomic DNA, which was enzymatically fragmented, and regions of interest were enriched using DNA capture probes. The final indexed libraries were sequenced on an Illumina platform. The prevalence of CAD according to genetic diagnosis of FH was analysed. Statistical analysis was performed using R (v. 4.0.4) program package.Results: Of 60 examined patients 28,3% (n=17) had the genetic diagnosis of FH and 71,7% (n=43) had no FH mutation. Premature CAD was found in 47% (n=8) patients with genetic diagnosis of FH and in 19% (n=8) with no mutations determined (p=0,049).Conclusions: Premature CAD was more prevalent among patients with genetically confirmed FH. Background and Aims : FH is known to be a predisposing cause of premature CAD. We aimed to determine the distribution of premature CAD according to genetic diagnosis of FH in Lithuania. Methods: Prospective observational cohort study enrolled patients with clinically diagnosed FH according to Dutch Lipid Clinic Network (DLCN) criteria treated in Vilnius University Hospital Santaros Klinikos during the period of 2016-2021. Premature CAD was defined as occuring in men younger than 55 years and women younger than 60 years. Data of 60 study patients were included in the analysis. Obstructive Atherosclerotic CAD was defined as the presence of stenosis ≥50% in at least one coronary vessel in Coronary Computed Tomography Angiography (CCTA) or coronary angiography, as well as performed percutaneous coronary intervention (PCI) or coronary arteries bypass grafting (CABG). Genetic testing was performed using genomic DNA, which was enzymatically fragmented, and regions of interest were enriched using DNA capture probes. The final indexed libraries were sequenced on an Illumina platform. The prevalence of CAD according to genetic diagnosis of FH was analysed. Statistical analysis was performed using R (v. 4.0.4) program package. Results: Of 60 examined patients 28,3% (n=17) had the genetic diagnosis of FH and 71,7% (n=43) had no FH mutation. Premature CAD was found in 47% (n=8) patients with genetic diagnosis of FH and in 19% (n=8) with no mutations determined (p=0,049). Conclusions: Premature CAD was more prevalent among patients with genetically confirmed FH.

The Prevalence and Genetic Spectrum of Familial Hypercholesterolemia in Qatar Based on Whole Genome Sequencing of 14,000 Subjects.

Familial hypercholesterolemia (FH) is an inherited disease characterized by reduced efficiency of low-density lipoprotein-cholesterol (LDL-C) removal from the blood and, consequently, an increased risk of life-threatening early cardiovascular complications. In Qatar, the prevalence of FH has not been determined and the disease, as in many countries, is largely underdiagnosed. In this study, we combined whole-genome sequencing data from the Qatar Genome Program with deep phenotype data from Qatar Biobank for 14,056 subjects to determine the genetic spectrum and estimate the prevalence of FH in Qatar. We used the Dutch Lipid Clinic Network (DLCN) as a diagnostic tool and scrutinized 11 FH-related genes for known pathogenic and possibly pathogenic mutations. Results revealed an estimated prevalence of 0.8% (1:125) for definite/probable cases of FH in the Qatari population. We detected 16 known pathogenic/likely pathogenic mutations in LDLR and one in PCSK9; all in a heterozygous state with high penetrance. The most common mutation was rs1064793799 (c.313+3A >C) followed by rs771019366 (p.Asp90Gly); both in LDLR. In addition, we identified 18 highly penetrant possibly pathogenic variants, of which 5 were Qatari-specific, in LDLR, APOB, PCSK9 and APOE, which are predicted to be among the top 1% most deleterious mutations in the human genome but further validations are required to confirm their pathogenicity. We did not detect any homozygous FH or autosomal recessive mutations in our study cohort. This pioneering study provides a reliable estimate of FH prevalence in Qatar based on a significantly large population-based cohort, whilst uncovering the spectrum of genetic variants associated with FH.

Optimisation of mutation prediction in suspected familial hypercholesterolaemia

Background: Familial hypercholesterolaemia (FH) mutation positive subjects have double the cardiovascular risk of low-density-lipoprotein-cholesterol (LDL-C) matched controls and are able to undergo cascade family screening. For best targeting of resources, it is desirable to assign an optimal strategy for mutation detection.

Clinical Diagnostic Criteria of Familial Hypercholesterolemia - A Comparison of the Japan Atherosclerosis Society and Dutch Lipid Clinic Network Criteria.

This study is aimed to compare the efficacy of the 2017 Japan Atherosclerosis Society (JAS) familial hypercholesterolemia (FH) criteria, which focuses on only 3 essential clinical manifestations, with that of Dutch Lipid Clinic Network (DLCN) FH criteria, which adopts a scoring system of multiple elements.Methods and Results:A total of 680 Japanese dyslipidemic participants (51% men) were enrolled between 2006 and 2018, all of whom had full evaluations of low-density lipoprotein (LDL) cholesterol, Achilles tendon X-rays, family history records, and genetic analysis of FH-associated genes (LDLR,APOB, andPCSK9). Predictive values for the existence of FH mutations by both clinical criteria were evaluated. Overall, 173 FH patients were clinically diagnosed by using the 2017 JAS criteria and 100, 57, 156, and 367 subjects were also diagnosed as having definite, probable, possible, and unlikely FH by the DLCN FH criteria, respectively. The positive and negative likelihood ratio predicting the presence of FH mutations by using the 2017 JAS FH criteria were 19.8 and 0.143, respectively; whereas, using the DLCN criteria of definite, probable, and possible FH, the ratios were 29.2 and 0.489, 9.70 and 0.332, and 3.43 and 0.040, respectively. Among Japanese patients, the JAS 2017 FH criteria is considered superior to diagnose FH mutation-positive patients and simultaneously rule out FH mutation-negative patients compared with the DLCN FH criteria.

Prevalence and Impact of Apolipoprotein E7 on LDL Cholesterol Among Patients With Familial Hypercholesterolemia.

Background: It has been suggested that a rare mutant apolipoprotein E7, APOE7 (p.Glu262Lys, p.Glu263Lys), has been identified to be associated with hyperlipoproteinemia in the general population. Moreover, its prevalence has been shown to be 0.005–0.06%. However, there are no prior data regarding its prevalence and impact on serum lipids in patients with familial hypercholesterolemia (FH).Methods: We recruited 1,138 patients with clinically diagnosed FH [mean age = 48, men = 512, median low-density lipoprotein (LDL) cholesterol = 231 mg/dl]. The coding regions of three FH genes (LDLR, APOB, and PCSK9) and apolipoprotein E (APOE) gene were sequenced. We investigated the prevalence and impact of APOE7 mutant on serum lipid levels in patients with FH.Results: We identified 29 patients (2.5 %) with a mutant APOE7 (heterozygote), which is apparently much higher than that of the general population. Moreover, when we focus on those without FH mutation (n = 540), we identified 21 patients (3.9 %) with a mutant APOE7. Patients with a mutant APOE7 exhibited significantly higher median LDL cholesterol and triglyceride levels compared with those without this rare mutant (249 vs. 218 mg/dl, p < 0.05, 216 vs. 164 mg/dl, p < 0.05, respectively). Moreover, LDL cholesterol levels in the APOE7-oligogenic FH individuals, with a pathogenic mutation in FH genes and APOE7 mutant, were significantly higher than that in monogenic FH patients (265 vs. 245 mg/dl, p < 0.05).Conclusion: We identified more patients with a mutant APOE7 than expected among those diagnosed with FH clinically, especially among those without FH-causing mutation. This implies a mutant APOE7 may be one of the causes FH, especially among those without FH mutations.

Long term follow-up of children with familial hypercholesterolemia and relatively normal LDL-cholesterol at diagnosis

Familial hypercholesterolemia (FH) is a genetic disorder with high low-density lipoprotein cholesterol (LDL-C) levels and high risk of cardiovascular disease. The long-term importance of carrying an FH mutation despite having relatively normal LDL-C levels in childhood is not known. We investigated the development of LDL-C levels and need of statin therapy in children with an FH mutation, with pretreatment LDL-C ≤ 4.1 mmol/L (~160 mg/dL), followed-up at lipid clinics in Oslo, Norway and Rotterdam, The Netherlands. Of 742 FH children, 109 (15%) had pretreatment LDL-C ≤ 4.1 mmol/L (~160 mg/dL) [mean (SD) 3.5 (0.5) mmol/L; (~130 (19) mg/dL)] measured at 11.8 (3.9) years of age [mean age (SD)]. After 8.2 (5.2) years [mean (SD)] of follow-up, 71.6% had started statin treatment. Therefore, all children carrying an FH mutation, independent of cholesterol levels, should receive follow-up at specialized lipid clinics for optimal and individualized treatment.

Cardiovascular Events According To Maternal Or Paternal Heritability Of Genetic Mutation In Familial Hypercholesterolemia

Familial hypercholesterolemia (FH) is a genetic autosomal disease, characterized by high levels of cholesterol since birth, that leads to a very high cardiovascular (CV) risk. In patients with the heterozygous FH (HeFH), in case of a maternal transmission, fetus is exposed to high maternal cholesterol levels during pregnancy. Based on the Developmental Origins of Health and Disease concept, our objective was to study the effect of heritability of genetic FH mutation on the occurrence of CV events in adults with HeFH. We included 138 HeHF patients with a genetic diagnosis and a clear familial history of hypercholesterolemia among 513 patients included in the Marseille's REFERCHOL register. Occurrence of CV events were compared by maternal (MH group, 64 patients) or paternal (PH group, 74 patients) heritability. The logistic regression method was used to determine the predictors of CV events. At inclusion, parameters were comparable between groups except more hypertension (25% vs. 9.4%, P = 0.01), more treatment by anti-PCSK9 (20.3 vs. 5.4%, P = 0.008) and less familial early CV events (28.6 vs. 51.4%, P = 0.007) in the MH group. Regarding CV events, 35.9% of MH patients and 27% of PH patients had a CV event ( P = 0.26). Predictive factors of CV event were sex ( P = 0.0004), age at start of statins ( P = 0.0003), maximal LDL-c ( P = 0.009) and Lp(a) ( P = 0.007). On multivariate analysis, heritability was not statistically associated with CV risk (PH risk vs. MH risk: OR = 0.34, P = 0.076). Sex, age at start of statins and Lp(a) were still associated with CV events occurrence (OR = 0.3, P = 0.044 for female; OR = 1.08, P = 0.001 for a later start of statins; OR = 6.98, P = 0.0096 for Lp(a)). Heritability has no significant effect on the occurrence of CV event during adulthood although paternal heritability seems to be protective. We are recruiting all HeFH patients from French FH register to improve the power of the study and confirm the observed trend.

Genomic analyses of all known putative familial hypercholesterolemia mutations showcase the value and limitations of public familial hypercholesterolemia mutation databases

Abstract Background/Introduction Familial hypercholesterolemia (FH) is genetically very heterogeneous and genomic and locus-specific public databases describing putative FH mutations are assumed to be of limited clinical utility because of classification errors. Purpose A description of all currently known publicly available putative FH mutations in order to determine the reliability of the classification of FH mutations described. Methods The LOVD and ClinVar databases were interrogated for the phenotype and genes of interest. Additional information on each variant was obtained using the Bioconductor toolset, gnomAD, and GETex. Results Currently know putative FH variants included 4,529 variants (97.2%) in the classical FH-genes LDLR (61%), PCSK9 (10%), and APOB (29%). Single nucleotide variants constituted 83% and 17% were copy number variants. Exonic variants contributed 78%, 14% of the variants were intronic, 7% large CNV, and 1% in upstream or downstream regions. Of the 4,529 variants, 45% were classified as pathogenic or likely-pathogenic (Fig. 1a). The ratio of exonic/intronic variants was 10.1 for pathogenic variants, 6.9 for likely pathogenic variants, 2.8 for likely benign and 1.4 for benign variants (p-value for class difference &amp;lt;2x10–6). For 502 frameshift mutations in exons that are particularly damaging, only 93.2% were classified as pathogenic or likely-pathogenic and 1.7% as benign or likely benign (Fig. 1b). Across 222 exon-covering deletions of &amp;gt;100 nucleotides, also particularly damaging, only 90.5% were classified as pathogenic or likely-pathogenic. Of the 4,529 variants, 1,561 (34.5%) were polymorphic in gnomAD. For these variants, the gnomAD sample size was on average 227420 (26102–282902). Of all 1,561 polymorphic variants in gnomAD 182 (11.6%) were classified as pathogenic or likely-pathogenic (Fig. 1c) and the variant frequency ranged from 10–4 to 10–6 (average 1.8x10–5), 43 with a frequency &amp;gt;1.8x10–5. Among variants found in gnomAD and classified as non-pathogenic, we observed ∼2000x higher frequencies (average 0.04). Of the 4,529 variants, 100 matched eQTLs or sQTLs in GTEx, none was annotated as pathogenic (Fig. 1d). Conclusion We review all currently known putative FH mutations with pathogenic or likely-pathogenic variants being mostly exonic. Highly damaging mutations are largely classified as pathogenic or likely-pathogenic, but up to 9% are not classified as pathogenic in the two public FH mutation databases. Assuming an FH population prevalence of 1/250 and 2000 pathogenic variants with the most frequent variant 10x more frequent than the average, we expect most pathogenic FH mutations at frequencies &amp;lt;2x10–5. We found 46 pathogenic or likely pathogenic variants to have a frequencies of &amp;gt;2x10–5 in the general population, evidence for misclassification. No pathogenic FH variant was found among GETEx eQTLs or sQTLs. Our data showcase the utility and weaknesses of the current public FH mutation databases. FH variants overview Funding Acknowledgement Type of funding source: Public hospital(s). Main funding source(s): Hôpitaux Universitaire de Genève - Fonds privés

Comparison between different diagnostic scores for the diagnosis of familial hypercholesterolemia: assessment of their diagnostic accuracy in comparison with genetic testing

Abstract Background The Dutch Lipid Clinic Network Score (DLCN) is a useful score to guide the physician to the diagnosis of familial hypercholesterolemia (FH). However, some concerns have been pointed out about its diagnostic accuracy in pediatric population; so, other scores such as Simon Broome (SB) and US MedPed Program have been tested and currently used. Aim To test the different diagnostic and predictive accuracy between those three scores (DLCN, SM and MedPed) in relation to positivity of genetic test. Methods We enrolled 25 pediatric patients aged &amp;lt;18 yo referred to our Chieti and Rome lipid clinics for the clinical suspicion of FH. We performed the genetic analysis for searching pathogenetic mutations for FH (NGS by Lipid in Code, LIPIGEN study). Differently from the others, DLCN provides a numerical data as a diagnostic score; thus, to uniform the three scores, we chose to define subjects with a DLCN score ≥6 as likely FH, while subjects with a lower value were defined as unlikely FH. Furthermore, we calculated the SB and the MedPed score, as previously demonstrated, defining the subjects as likely or unlikely FH. Therefore, we analyzed the diagnostic accuracy of those scores in relation to FH mutation data (positive or negative). Results We recruited 25 patients, 13 males (52%), mean age of 11.8 years. Sixteen patients (64%) resulted positive for the genetic mutation The DLCN showed a sensitivity of 43%, specificity of 66%, a positive predictive value (PPV) of 0.7 and a negative predictive value (NPV) of 0.4. SB showed a sensitivity of 68%, specificity of 55%, a PPV of 0.73 and NPV of 0.5. The MedPed score showed a sensitivity of 68%, specificity of 44%, a PPV of 0.68 and NPV of 0.44. Furthermore, we elaborated a ROC curve of low-density lipoprotein cholesterol (LDL-C) values with respect to the positivity of the FH genetic test: the value that showed the best diagnostic accuracy was 190 mg / dL (sensitivity 68.8% and specificity 75%), while the value of 116.5 mg/dL demonstrated a sensitivity of 100% but with a specificity of 11%. Conclusions Our work demonstrates that none of the three scores tested has an optimal diagnostic accuracy with respect to the diagnosis of FH in the pediatric population. By comparison, SB demonstrates the best sensitivity and the best specificity. In addition, the LDL-C value of 190 mg / dL provides the best diagnostic accuracy against the diagnosis of FH in the pediatric population. Further data and studies will be needed to improve the diagnostic performance of these scores, in order to refer pediatric patients to the genetic test, for an early confirmation in order to refer pediatric patients to the genetic test, for an early FH diagnosis confirmation. Funding Acknowledgement Type of funding source: None

A reassessment of the Japanese clinical diagnostic criteria of familial hypercholesterolemia in a hospital-based cohort using comprehensive genetic analysis

BackgroundClinical diagnostic criteria of familial hypercholesterolemia (FH) in Japan include LDL cholesterol ​≥ ​180 ​mg/dL, Achilles tendon thickness ​≥ ​9.0 ​mm, and family history. However, few data exist regarding its validation. Design and MethodsA series of 680 participants, with a mean LDL cholesterol of 175 ​mg/dL were enrolled at Kanazawa University Hospital between 2006 and 2018. All had full assessments of, LDL cholesterol, Achilles tendon X-rays, family history records, and genetic analysis of FH-associated genes (LDLR, APOB, and PCSK9). The area under the curve (AUC) of receiver operating characteristic (ROC) curve analysis predicting the presence of FH mutations by each clinical marker were assessed. ResultsThe optimal cutoff values predicting the presence of an FH-associated mutation were 181 ​mg/dL for LDL cholesterol and ≥7.0 ​mm for Achilles tendon thickness. AUCs predicting FH mutations were 0.827 for Achilles tendon thickness ≥9.0 ​mm, 0.889 for LDL cholesterol ≥180 ​mg/dL, and 0.906 for family history. If Achilles tendon thickness ≥7.0 ​mm was used as a clinical criterion, then 41 participants (6%) were newly diagnosed with FH and 86 (12%) were newly misclassified as FH. ConclusionsCurrent clinical diagnostic criteria of FH were validated in this cohort. We recommend considering a tentative diagnosis of “potential FH” if the Achilles tendon thickness is ​≥ ​7.0 ​mm and <9.0 ​mm rather than dismissing a diagnosis of FH.

Mutation spectrum and polygenic score in German patients with familial hypercholesterolemia.

Autosomal-dominant familial hypercholesterolemia (FH) is characterized by increased plasma concentrations of low-density lipoprotein cholesterol (LDL-C) and a substantial risk to develop cardiovascular disease. Causative mutations in three major genes are known: the LDL receptor gene (LDLR), the apolipoprotein B gene (APOB) and the proprotein convertase subtilisin/kexin 9 gene (PCSK9). We clinically characterized 336 patients suspected to have FH and screened them for disease causing mutations in LDLR, APOB, and PCSK9. We genotyped six single nucleotide polymorphisms (SNPs) to calculate a polygenic risk score for the patients and 1985 controls. The 117 patients had a causative variant in one of the analyzed genes. Most variants were found in the LDLR gene (84.9%) with 11 novel mutations. The mean polygenic risk score was significantly higher in FH mutation negative subjects than in FH mutation positive patients (P < .05) and healthy controls (P < .001), whereas the score of the two latter groups did not differ significantly. However, the score explained only about 3% of the baseline LDL-C variance. We verified the previously described clinical and genetic variability of FH for German hypercholesterolemic patients. Evaluation of a six-SNP polygenic score recently proposed for clinical use suggests that it is not a reliable tool to classify hypercholesterolemic patients.

Fragment-based design of small molecule PCSK9 inhibitors using simulated annealing of chemical potential simulations.

PCSK9 is a protein secreted by the liver that binds to the low-density lipoprotein receptor (LDLR), causing LDLR internalization, decreasing the clearance of circulating LDL particles. Mutations in PCSK9 that strengthen its interactions with LDLR result in familial hypercholesterolemia (FH) and early onset atherosclerosis, while nonsense mutations of PCSK9 result in cardio-protective hypocholesterolemia. These observations led to PCSK9 inhibition for cholesterol lowering becoming a high-interest therapeutic target, with antibody drugs reaching the market. An orally-available small molecule drug is highly desirable, but inhibiting the PCSK9/LDLR protein-protein interaction (PPI) has proven challenging. Alternate approaches to finding good lead candidates are needed. Motivated by the FH mutation data on PCSK9, we found that modeling the PCSK9/LDLR interface revealed extensive electron delocalization between and within the protein partners. Based on this, we hypothesized that compounds assembled from chemical fragments could achieve the affinity required to inhibit the PCSK9/LDLR PPI if they were selected to interact with PCSK9 in a way that, like LDLR, also involves significant fractional charge transfer to form partially covalent bonds. To identify such fragments, Simulated Annealing of Chemical Potential (SACP) fragment simulations were run on multiple PCSK9 structures, using optimized partial charges for the protein. We designed a small molecule, composed of several fragments, predicted to interact at two sites on the PCSK9. This compound inhibits the PPI with 1 μM affinity. Further, we designed two similar small molecules where one allows charge delocalization though a linker and the other doesn’t. The first inhibitor with charge delocalization enhances LDLR surface expression by 60% at 10 nM, two orders of magnitude more potent than the EGF domain of LDLR. The other enhances LDLR expression by only 50% at 1 μM. This supports our conjecture that fragments can have surprisingly outsized efficacy in breaking PPI’s by achieving fractional charge transfer leading to partially covalent bonding.