Familial Hypercholesterolaemia in Portugal: A Need for More Diagnostic Screening

Mini Review

J Blood Disord. 2014;1(1): 3.

Familial Hypercholesterolaemia in Portugal: A Need for More Diagnostic Screening

Kazachkova N*

Department of Biology, University of the Azores, Portugal

*Corresponding author: Kazachkova N, Department of Biology, Institute for Molecular and Cell Biology (IBMC), University of Porto and Centre of Research in Natural Resources (CIRN), University of the Azores, Rua Mãe de Deus-Apartado 1422, 9501-801 Ponta Delgada, Azores, Portugal.

Received: July 30, 2014; Accepted: September 12, 2014; Published: September 17, 2014


Familial hypercholesterolaemia; Lipoprotein metabolism; Coronary artery disease; LDLR; APOB; PSCK9


FH: Familial Hypercholesterolaemia; LDL: Low Density Lipoprotein; CHD: Coronary Heart Disease; TC: Total Cholesterol; LDL-C: LDL Cholesterol; HeFH: Heterozygous FH; HoFH: Homozygous FH; ARH: Autosomal Recessive Hypercholesterolemia; LDLR: Low Density Lipoprotein Receptor; APOB: apolipoprotein B; PCSK9: Proprotein Convertase Subtilisin/Kenin Type 9; OMIM: Online Mendelian Inheritance In Man; RT-PCR: Reverse Transcriptase Polymerase Chain Reaction; DHPLC: Denaturing High Pressure Liquid Chromatography; MLPA: Multiplex Ligation-Dependent Probe Amplification; RNA: Ribonucleic Acid


Familial Hypercholesterolaemia (FH) (MIM 144010) is an autosomal dominant disorder of lipoprotein metabolism, which is characterized by the increased level of circulating Low Density Lipoprotein (LDL) cholesterol that leads to lipid accumulations in arteries, and sometimes in tendons (xanthomas) and around the eyes (xanthelasmas), resulting in premature atherosclerosis and increased risk of Coronary Heart Disease (CHD [1]. If remains untreated, FH can increase the cumulative risk for CHD by up to 20-fold (men are at 20% higher risk than women), the relative risk to develop premature CHD for men and women with FH aged between 20 and 39 years old is about 100-fold increased compared to the normal population [1,2]. These data demonstrate a serious need for diagnostic screening for the disease. The treatment with cholesterol-lowering medications will promote the arrest of atherosclerosis progression and prevention of CHD.

Currently, FH remains to be under diagnosed and undertreated in the general population. Based on prevalences between 1/500 and 1/200, 14-34 million individuals worldwide have FH [3]. Recent evaluation of the extent to which FH is under diagnosed and undertreated worldwide demonstrated that <1% of FH casesare diagnosed in most countries, except the Netherlands (71% of diagnosed FH cases), Norway (43%), Iceland (19%), Switzerland (13%), the UK (12%), and Spain (6%) [3]. Although Portugal should have about 20,000 cases, this disease is severely under diagnosed in the country, it is estimated that 80% of FH patients are not even clinically identified [4].

Nowadays, FH can be diagnosed either clinically or genetically. Both of these methods have their advantages and disadvantages. Clinical diagnosis can be useful in diagnosing relatives of known FH patients and avoiding the assessment of patients, who are probably not FH cases. Moreover, the cost of clinical diagnosis is low. However, clinical diagnosis cannot distinguish between FH cases due to mutations in different genes, between Heterozygous FH (HeFH) and homorozygous FH (HoFH), Autosomal Recessive Hypercholesterolemia (ARH) [5] or non-familial hypercholesterolemia (secondary hypercholesterolemia, sitosterolemia, etc.), it can miss asymptomatic FH cases (e.g. in the pediatric population) [3,6]. While genetic testing can provide not only the early diagnosis, but also a strategy to prevent the risk of cardiovascular complications. It can diagnose asymptomatic FH cases, distinguish between all types of FH cases and have a prognostic significance for succeeding generations. The high cost, the polygenetic nature of the disease and affection of the phenotype by many non-genetic factors are among limitations of genetic testing [3,6]. Most probably, the optimal and most effective diagnosis should be based both on clinical and genetic tests. For example cascade screening is a strategy that already proved to be costeffective in the FH diagnosis. It is characterized by the combination of initial clinical diagnosis with subsequent genetic testing by the next scheme: genetic test confirms the mutation in the index patient, then first degree relatives are screened for the same mutation and new confirmed cases are treated as new index cases and their first degree relatives are screened [6].

So far, there are no single internationally accepted criteria for the clinical diagnosis of FH [6]. Three main criteria are currently used for FH diagnostics: Dutch Criteria (the Netherlands), MEDPED Criteria (USA), and Simon Broome Criteria (UK). The US criteria are based on lipid levels and age, while the UK and Dutch criteria classify FH cases as definite, probable, and possible using not only lipid levels, but also physical signs, family and personal history [6]. According to Simon Broome Criteria, which is the most widely used by physicians, plasma Total Cholesterol (TC) and LDL-C levels in FH cases range from 7.5 mmol/L (290 mg/dl) and 4.9 mmol/L (190 mg/dl), respectively, in adults, and from 6.7 mmol/L (260 mg/dl) and 4 mmol/L (155 mg/dl), respectively, in children [6].

Genetic diagnosis of FH is currently based on the testing of defects in the low density lipoprotein receptor gene (LDLR, OMIM 606945), the gene for apolipoprotein B (APOB, OMIM 107730), and the proprotein convertase subtilisin/kenin type 9 (PCSK9, OMIM607786) gene. The frequency of FH is about 1/200-500 for HeFH and 1/160000-1000000 for HoFH [1,7]. The LDLR is responsible for the binding and subsequent cellular uptake of apolipoprotein B- and E-containing lipoproteins [8], APOB is the major protein component of triglyceride-rich and low density lipoproteins, it is responsible for carrying cholesterol to tissues [9], and PCSK9 is a serine protease that reduces both hepatic and extra hepatic LDLR levels and increases plasma LDL cholesterol [10]. Mutations in any of these genes can cause impairment of normal uptake and clearance of LDL by the liver. More than 1300 mutations (account for 60-80% of FH) are known so far in the LDLR gene [11], they include single-nucleotide mutations, copy number variations, and splicing mutations and can be divided into five distinct classes according to how they affect LDLR expression and function (null (produce no detectable protein), transport-defective, biding-defective, internalization-defective and recycling-defective); more than 20 mutations are related to FH in the PCSK9 gene (account for 0-3% of FH), and several - in the APOB gene (e.g., rs5742904; account for 1-5% of FH) [1,3,12,13].

In Portugal many patients have TC above 350 mg/dl and triglycerides below 200 mg/dl, which suggests the presence of FH in this population [14-16]. The Portuguese population has much low percentage of patients with tendon xanthomas (present in only 8.5% FH cases) than other populations where the presence of tendon xanthomas is generally considered diagnostic of FH (present in 22.5% of Spanish FH patients and 78.8% of English FH patients) [14,15]. It contributes to higher level of undiagnosed FH and makes molecular diagnostics of great importance for this population. In the Portuguese population three common genes have been analyzed for the association with FH, namely LDLR, APOB, and PCSK9 [4,11,13- 15,17]. Individuals form Azores, Madeira, and Mainland have been studied in family-based and case-based studies. Known mutations have been analyzed as well as mutation screening has been performed in these studies using such methods as sequencing, DHPLC, MLPA, RT-PCR and fragment analysis, site-directed mutagenesis, protein expression in vitro, etc. In total, 113 genetic variants (point, nonsense, splice site and missense mutations, mutations in the promoter region, deletions and insertions, polymorphisms) were described for the Portuguese population [4,11,13-15,17]. 93 genetic variants have been described for the LDLR gene, 51 of them were associated with higher risk for FH, 39 genetic variants were reported as novel [4,13- 15,17], two risk variants of seven described (among them 5 novel) have been reported for the PCSK9 gene [4,14,15,17], and five of 13 (12 of them were novel) - for the APOB gene (Table 1) [4,11,14,15,17]. Most of these data were obtained from the Portuguese Familial Hypercholesterolemia Study that was initiated at the National Institute of Health in 1999 year, aiming to perform an epidemiological study to determine the prevalence and distribution of FH in Portugal and to better understand the pathophysiology of CHD in these patients [14]. The clinical criteria for FH used in this study were adapted from the UK Simon Broome Criteria. The genetic diagnosis of FH was performed in three phases and an optional fourth phase: I - screening for the common mutations in the APOB gene and analysis of the LDLR gene, II - identification of large rearrangements in the LDLR gene using MLPA technique, III - screening of the PCSK9 gene (if no mutation was found in phases I and II), and IV (only performed when putative splicing mutations, not described before or without functional studies, are found). Analysis of relatives was performed asthe cascade screening [4]. Index cases and relatives with or without a clinical diagnosis of FH were enrolled in the Portuguese Familial Hypercholesterolemia Study. In total, genetic analysis was performed in 684 index cases and 1210 relatives. Only in 52.8% of index cases (340/49.7% in the LDLR gene, including eight homozygotes and seven compound heterozygotes, 9/1.32% in the APOB and 12/1.75% in the PCSK9) and 45.2% of relatives (529/43.72% in the LDLR gene, 13/1.07% in the APOB and 5/0.41% in the PCSK9) studied, a mutation causing disease was identified [4,14,15]. This fact can be explained by several reasons, e.g. some patients do not meet inclusion criteria, there are genetic changes in another gene(s) involved in LDLR function, regulation or LDL metabolism, the underlying gene defect causing FH in these patients is in a non-coding region of the LDLR gene that may affect its expression or RNA processing, or nonfamilial hypercholesterolemia is mistakenly taken for FH [4].

Citation: Kazachkova N. Familial Hypercholesterolaemia in Portugal: A Need for More Diagnostic Screening. J Blood Disord. 2014;1(1): 1004. ISSN 2379-8009