Background and Aims: Familial Hypercholesterolemia (FH) is a common genetic disorder of lipid metabolism associated to increased CAD risk. Three genes are associated with FH (LDLR, APOB, PCSK9). Variants in FH phenocopies genes (LDLRAP1, APOE, LIPA, ABCG5, ABCG8), LDL-C polygenic risk score (PRS) and hyper-Lp(a) can mimic the FH phenotype. In the present work we intend to unravel the genetic background in individuals with clinical diagnosis of FH. Methods: A biochemical and genetic study was performed to 1005 patients with clinical diagnosis of FH referred to the Portuguese FH Study until December 2021. Since 2017, genetic diagnosis is performed by an NGS panel with 8 genes and 6-SNPs to determine PRS. Results: FH was genetically confirmed in 41% of the cases. In the FH-negative cohort (N=590), 33% (N=192) present Lp(a)>50mg/dl, 17% (N=102) have high PRS, 1% (N=7) have other monogenic cause and 1% (N=6) have one pathogenic variant in ABCG5/ABCG8. Additionally, 5% (N=32) carry heterozygous VUS in either LDLR, APOB or PCSK9 and 5% (N=29) carry heterozygous variants of unknown significance (VUS) in FH phenocopies genes. No identifiable cause of dyslipidemia was found in the remaining 38% patients. Conclusions: Overall, FH was confirmed genetically in 41% of the cohort. In 50% of the FH negatives the FH phenotype can be caused by Hyper-Lp(a) or high PRS. A small part of patients has pathogenic variants in ABCG5/8 in heterozygosity and this can be the cause of hypercholesterolemia and should be further investigated. This extended NGS panel is important to identify FH/FH-phenocopies and therefore personalize each patient’s treatment.

Background and Aims: Sitosterolemia is an autosomal recessive disorder caused by variants in ABCG5/8genes and is characterized by severely elevated plasma plant sterols, causing xanthomas and premature cardiovascular disease. The aim of this study is to present 12 clinical cases of sitosterolemia in Iberoamerican countries. Methods: We report seven index cases, and five relatives identified by cascade screening. Clinical and laboratory characteristics were determined in different countries. All individuals were sequenced for the ABCG5/8 genes either by sanger or next generation sequencing. Whenever possible sterols chromatography was performed. Results: All index children and 2 index adults presented xanthomas and children had a mean LDL of 570 mg/dL. The geographical distribution of the 12 cases is as follows: five from Spain (four compound heterozygous and one true homozygous, all in ABCG8); three from Portugal (all true homozygous, two in ABCG8 and one in ABCG5); one from Argentina (true homozygous in ABCG8) and three from Uruguay (all true homozygous in ABCG8). Six index cases present variants in ABCG8 gene (two missense and four nonsense variants) and only one presents a homozygosity in the ABCG5gene (frameshift variant). Six cases (4 index and 2 relatives) (50%) were diagnosed in childhood, the remaining were identified as adults. Conclusions: There is a genetic heterogeneity of sitosterolemia in Iberoamerican countries. The treatment for sitosterolemia is specific and completely different from the other types of hypercholesterolemia, so a correct and timely diagnosis is crucial to avoid delays in treatment that could lead to poor clinical outcomes.

Background and Aims: Familial hypercholesterolemia (FH) is a common inherited lipid disorder that predisposes to cardiovascular disease (CVD). Despite most cascade screening programs are initiated by adult index cases, reverse cascade screening pediatric index cases is starting to be described. Therefore, we aimed to assess the outcome of adult cascade screening and child reverse cascade screening strategies in families from the Portuguese FH Study (PFHS). Methods: The PFHS database was consulted, and 423 index cases genetically identified with FH (224 adults and 199 children) and their 997 relatives referred to the PFHS were analysed. Results: From 224 adults with FH, 485 relatives were enrolled for cascade screening and 290 were identified with FH. From 199 paediatric cases with FH, 512 relatives were screened and 286 were identified with FH. Child reverse cascade screening presented a slightly higher diagnostic rate than adult cascade screening, 1.44 vs 1.29 new cases with FH per index case, and the age of the relatives identified was younger, 29 vs 37 years. For 94% of index children, relatives were referred (2.56 relatives per index), in contrast with the adult cohort whereas only 70% were referred with family-members (2.17 relatives per index). Conclusions: Overall, both screening approaches constitute valuable tools to identify new cases with FH, but the child reverse cascade screening creates the opportunity for more relatives to be tested at a younger age. It is crucial to improve relatives' recruitment rate since early identification allows a correct FH diagnosis and treatment to prevent CVD.

Background and Aims: Familial hypercholesterolemia (FH) is an autosomal semi-dominant disease associated with pathogenic (P) or likely pathogenic (LP) variants in LDLR, APOB and PCSK9 genes. Here we report the first results of the detection rate of monogenic variants in FH patients in Latvia. Methods: Whole genome sequencing (WGS) with 30x coverage was performed in index cases selected from the Latvian Registry of FH. The diagnosis was defined according to the Dutch Lipid Clinic Network criteria. LDLR, APOB, PCSK9, LDLRAP1, ABCG5, ABCG8, LIPA, LPA, CYP27A1, APOE genes were analyzed. Here only variants annotated as P/LP using the FH Variant Curation Expert Panel (VCEP) guidelines for LDLR and adaptations for APOB and PCSK9 are reported. The statistical analysis was performed with IBM SPSS, version 22. Results: Among 164 patients, 66.7% were women, mean age was 52.9+/-11.4 years and mean highest documented LDL-cholesterol level was 7.5+/-1.7 mmol/L. The diagnosis was definite, probable and possible FH in 57 (34.8%), 105 (64.0%) and 2 (1.2%) patients, respectively. A total 15 P/LP variants were found in 34 patients (diagnostic yield 20.7%), 14 in LDLR and 1 in APOB gene. Additionally, 10 VUS were also detected in LDLR. Conclusions: Despite high clinical likelihood of FH, confirmed P/LP variants were detected only in 20.7% of patients. Future studies will extend to internal analysis of other variants based on VCEP criteria, impact of polygenic mechanisms, and WGS in a larger Latvian sample.

Background and Aims: Familial hypercholesterolemia (FH) is the most common monogenic disorder of lipid metabolism. Genetic testing can confirm the clinical diagnosis, but there are currently over 3300 different variants in LDLR deposited in ClinVar and about ∼400 had conflicting classifications of pathogenicity. Here, we present the progress of LDLR variant classification by the FH Variant Curation Expert Panel (VCEP), composed of 13 reviewers, 17 curators, and 12 associated labs, with our LDLR consensus variant classification guidelines. Methods: Variants with conflicting classifications and other variants in the same codon (required to properly classify conflicting variants) are prioritized. Associated labs send internal variant case-level data, which is uploaded into the Variant Curation Interface (VCI) and supplemented by literature evidence. Each variant is assessed by one (very experienced) or two curators and approved by three reviewers before being officially published to ClinVar. Results: As of December 2022, we have completed classification of 316 LDLR variants. Of those with prior conflicting classifications (n=165), 33% were classified as Pathogenic/Likely pathogenic (P/LP), 9% as Benign/Likely benign (B/LB), 55% as Variant of Uncertain Significance (VUS) by insufficient evidence and only 3% remained conflicting. Of the remaining 135 variants, 53% were classified as P/LP, 2% as B/LB and 45% as VUS. Until May 2023, we will evaluate 451 LDLR variants, 247 of them with prior conflicting classifications. Conclusions: Ultimately, efforts of the FH VCEP hope to improve FH genetic diagnosis, which relies on accurate LDLR variant classification. FH VCEP’s guidelines significantly decrease conflicting classifications, which will be especially helpful to the FH community.

Abstract Issue/problem Healthcare (HC) can significantly benefit from genomic information for earlier, accurate diagnosis, effective personalized treatment with less adverse events, and accurate profiling of individuals for disease prevention. However, European countries are currently at variable maturity stages regarding the implementation of genomic medicine (GM) in healthcare, hindering the equitable delivery of personalized medicine to citizens across borders. Description of the problem The European 1+Million Genomes Initiative (1+MG) aims to provide cross-border access to quality genomic information and related clinical data, to advance data-driven research and HC solutions to benefit citizens. This initiative is encouraging countries to develop national GM strategies, but guidance for successful implementation is needed. In this context, the Beyond 1 Million Genomes, a supporting action to the 1+MG initiative, organized three Country Exchange Visits (CEV) to discuss critical issues, share experiences and best practices, for the implementation of sustainable GM strategies in healthcare. Results The United Kingdom, Estonia and Finland, which have advanced GM programs, hosted CEV describing progress and lessons learnt. Representatives of 1+MG signatory countries participated in these events and were able to present country level progress. The resulting Policy Brief (PB) captures key issues discussed at the CEVs, with real-life examples, and proposes policy recommendations for the successful implementation of GM in European healthcare systems. Lessons Sustainable GM implementation in HC systems requires: 1) Patient and citizens trust and engagement; 2) Sustainable infrastructure and data regulation, with solid ethical and legal frameworks; 3) Capacity building of healthcare professionals; 4) A strong ecosystem involving all stakeholders, and encouraging synergies between healthcare, research and industry to promote continuous innovation. Key messages • The implementation of GM in healthcare will take countries further towards making personalized medicine a reality, with remarkable health and socioeconomic benefits for patients and healthcare systems. • Promoting cooperation, capacity building and sharing of best practices is crucial to reduce asymmetries between countries, which constrains effective and equitable cross-border personalized medicine.

Background and Aims : Dyslipidaemia is a group of disorders of lipid metabolism characterized by abnormal lipid concentrations and are associated to serious conditions that can be prevented by early identification of patients. This project aims to characterize 96 Portuguese patients with clinical diagnoses of 3 main familial dyslipidemias.Methods: Molecular analysis (19 genes of lipid metabolism) was performed by Next Generation Sequencing. NGS Library was run on an Illumina NextSeq.Conclusions: This project contributed to a personalized diagnosis and treatment of Portuguese individuals improving patients’ prognosis. Since several variants of uncertain significance were found and might constitute a genetic cause of dyslipidemia, functional studies will be essential to investigate their impact. Background and Aims : Dyslipidaemia is a group of disorders of lipid metabolism characterized by abnormal lipid concentrations and are associated to serious conditions that can be prevented by early identification of patients. This project aims to characterize 96 Portuguese patients with clinical diagnoses of 3 main familial dyslipidemias. Methods: Molecular analysis (19 genes of lipid metabolism) was performed by Next Generation Sequencing. NGS Library was run on an Illumina NextSeq. Conclusions: This project contributed to a personalized diagnosis and treatment of Portuguese individuals improving patients’ prognosis. Since several variants of uncertain significance were found and might constitute a genetic cause of dyslipidemia, functional studies will be essential to investigate their impact.

Background and Aims : Familial Hypercholesterolemia (FH) is a monogenic, common autosomal disorder of lipid metabolism. Genetic diagnosis includes the study of 3 genes: LDLR, APOB, PCSK9, but 50-60% of clinical FH patients present a negative result. The present work aims determine if the cause of hypercholesterolemia in FH negative individuals can be explained by a polygenic contribution and compare three different LDL-C genetic risk score (GRS) in clinical FH patients.Methods: A total of 48 index cases (ICs) with clinical diagnosis of FH (Simon Broome criteria) was analysed by a NGS panel and were negative for FH genes. Selected SNPs associated with raising plasma LDL-C in 3 GRS were also included in the panel: 6-SNPs (Futema et al., 2015), 12-SNPs (Talmud et al., 2013) and 10-SNPs (Wang et al., 2016). High polygenic risk score (PRS) was considered for GRS≥0.76, GRS>0.73 and GRS≥1.96, respectively.Results: Among the 48 FH negative ICs (15 children + 32 adults), a high PRS was identified in 19 (6-SNPs GRS), 41 (12-SNPs GRS) and 9 ICs (10-SNPs GRSs). In addition, 9 ICs have a 6-SNPs GRS<0.51.Conclusions: There are large differences in the GRS identification of polygenic hypercholesterolaemia. The 12-SNPs GRS identifies 85% of all FH negative individuals with a polygenic hypercholesterolemia compared to just 19% using the 10-SNPs GRS. There is a need to develop more accurate GRS for polygenic hypercholesterolaemia. The 6-SNPs GRS additionally identifies that 19% of the FH negative ICs have a low PRS, highlighting that they can have an unidentified monogenic cause. Background and Aims : Familial Hypercholesterolemia (FH) is a monogenic, common autosomal disorder of lipid metabolism. Genetic diagnosis includes the study of 3 genes: LDLR, APOB, PCSK9, but 50-60% of clinical FH patients present a negative result. The present work aims determine if the cause of hypercholesterolemia in FH negative individuals can be explained by a polygenic contribution and compare three different LDL-C genetic risk score (GRS) in clinical FH patients. Methods: A total of 48 index cases (ICs) with clinical diagnosis of FH (Simon Broome criteria) was analysed by a NGS panel and were negative for FH genes. Selected SNPs associated with raising plasma LDL-C in 3 GRS were also included in the panel: 6-SNPs (Futema et al., 2015), 12-SNPs (Talmud et al., 2013) and 10-SNPs (Wang et al., 2016). High polygenic risk score (PRS) was considered for GRS≥0.76, GRS>0.73 and GRS≥1.96, respectively. Results: Among the 48 FH negative ICs (15 children + 32 adults), a high PRS was identified in 19 (6-SNPs GRS), 41 (12-SNPs GRS) and 9 ICs (10-SNPs GRSs). In addition, 9 ICs have a 6-SNPs GRS<0.51. Conclusions: There are large differences in the GRS identification of polygenic hypercholesterolaemia. The 12-SNPs GRS identifies 85% of all FH negative individuals with a polygenic hypercholesterolemia compared to just 19% using the 10-SNPs GRS. There is a need to develop more accurate GRS for polygenic hypercholesterolaemia. The 6-SNPs GRS additionally identifies that 19% of the FH negative ICs have a low PRS, highlighting that they can have an unidentified monogenic cause.

Background and Aims: Familial Hypercholesterolemia (FH) confers a high risk for cardiovascular disease (CVD) with several modulating characteristics, which makes it the perfect candidate for personalised medicine approaches.

Background and Aims: Familial hypercholesterolemia (FH) is due to mutations in LDLR, APOB and PCSK9, however, about 50% of clinical FH patients do not have an identifiable genetic cause. MicroRNAs (small non-coding RNAs) are negative regulators of gene expression and creation of new biding sites can be the cause of hypercholesterolaemia in mutation-negative families. The present work aims to analyse miRNAs targets as regulators of genes involved in lipid metabolism in FH patients.

Background and Aims: Familial hypercholesterolemia (FH) is the most common genetic disorder conferring an increased cardiovascular risk due to cholesterol accumulation since birth. FH patients have usually mutations in LDLR, APOB or PCSK9 genes, but in about 50% a variant causing disease is not identified. The 5' and 3' untranslated regions (UTRs) and promoter of these genes is poorly studied. The aim of this project is to perform an in vitro characterization of variants in 5' UTR and promoter of LDLR gene.

Background and Aims: Familial hypercholesterolemia (FH) is an inherited disorder of lipid metabolism, characterized by increased low density lipoprotein cholesterol (LDLc) levels. Additional information regarding lipid profile may however be of great importance, when developing FH diagnosis models. The main purpose of this work was to test several logistic regression models, including a predefined set of clinical variables, together with progressively more specific sets of biochemical variables.

Background and Aims: Familial hypercholesterolaemia (FH) is one of the most common genetic disorders, with a frequency around 1:250 in most populations. FH phenotype has a considerable genetic heterogeneity and phenotypic variability, and it is not uncommon to observe a wide range of lipid levels among carriers of an identical FH-causing mutation. Here we characterise two LDLR missense variants [c.1A>T/p.(Met1Leu) and c.1A>C/p.(Met1Leu)] in the initiation codon exhibiting distinct functional phenotypes.

Background and Aims: Mutations in the Low Density Lipoprotein Receptor (LDLR) gene are the major cause of familial hypercholesterolaemia (FH), with over 2600 variants described. However, less than 15% of the LDLR variants identified in clinical FH patients have functional evidences to prove their pathogenicity. The aim of the present work is to stablish a quantitative high-throughput in vitro microscopy approach to functionally characterize rare LDLR variants.

Background and Aims: Familial hypercholesterolemia (FH) is a heritable disorder characterized by elevated low-density lipoprotein (LDL) cholesterol levels and premature cardiovascular disease. Increasingly, patients with a clinical suspicion of FH are being offered genetic testing as part of diagnosis. However, determining pathogenicity of identified DNA variants remains a major challenge; this process is often rudimentary and differs among laboratories. To improve accuracy, concordance and standardization, the ClinGen FH Variant Curation Expert Panel (VCEP) has been tasked with developing FH-specific variant interpretation guidelines, derived from American College of Medical Genetics and Genomics and Association for Molecular Pathology (ACMG/AMP) recommendations (2015).

Background and Aims: Familial chylomicronemia syndrome (FCS) is a rare autosomal disorder of lipoprotein metabolism. It is characterized by marked elevation of triglyceride and chylomicron levels, lipaemic plasma, recurrent pancreatitis, eruptive xanthoma, hepatosplenomegaly, and lipemia retinalis. All genes associated with FCS (LPL, APOC2, APOA5, LMF1 and GPHBP1) have an effect on the activity of lipoprotein lipase (LPL). The aim of this study is to present all cases with FCS clinical diagnosis, studied in our laboratory.

Background and Aims: Statins are the standard treatment for dyslipidaemia disorders, but there is a wide (10-70%) variation in patient response to statin treatment and several documented serious adverse effects. Studies of variants in genes regulating drug absorption, metabolism, pharmacodynamics and excretion mechanisms, have been implicated as reasons for this variability. The aim of this study is to determine the prevalence of pharmacogenetic relevant genotypes in the Portuguese population and in a sample of high cardiovascular risk patients: familial hypercholesterolemia (FH) subjects. Genetic associations with biomarkers in lipid metabolism will be studied.

Background and Aims: High LDL-C values may be due to monogenic variants in LDLR, APOB, PCSK9 genes which lead to familial hypercholesterolemia (FH), but also to a polygenic origin that arises from polymorphisms in different genes related to cholesterol metabolism. Aim: to apply a genetic risk score (GRS) with SNPs:6 in a sample of hypercholesterolemic patients from the FH Detection Program in Argentina: Da Vinci Study

Background and Aims: Familial hypercholesterolemia (FH) is an autosomal dominant disorder associated with high levels of LDL-c and premature CHD (pCHD). Identification of FH in pediatric age is essential for a timely diagnosis and management. This study aims to highlight the importance of FH diagnosis in children.