MOLECULAR CHARACTERIZATION AND SPECTRUM OF MUTATIONS ASSOCIATED WITH AUTOSOMAL DOMINANT HYPERCHOLESTEROLEMIA

Introduction- Familial Hypercholesterolemia (FH) is a common genetic disease that is inherited in an autosomal dominant manner. FH can be caused by mutations in the low-density lipoprotein receptor gene (LDLR) in 80% of cases, in its main ligand apolipoprotein B gene (ApoB-100) in 5% of cases, in the propotein convertase subtilin/kexin type 9 gene (PCSK9), an enzyme involved in LDLR degradation, in 1-2% of cases, while the genetic alteration is still unidentified in 15% of cases. FH leads to premature coronary heart disease(CHD), namely myocardial infarction and angina pectoris, due to elevated plasma low-density lipoprotein cholesterol (LDL-C) levels and high levels of total cholesterol (TC). About 1 in 500 individuals in the general population is affected, making it the most common monogenic form of hypercholesterolemia. Patients with homozygous alleles for defective LDLR are very rare, the frequency is about 1:106. Three scientific groups have developed diagnostic tools for FH: The US MedPed Program (Make Early Diagnosis, Prevent Early Death), the Simon Broome Register Group in the United Kingdom , and the Dutch Lipid Clinic Network (DLCNC). The major difference between these systems is the use of different cut-offs for premature CHD. The US MedPed Program based on validated age and sex adjusted for serum cholesterol cut-off points. The Simon Broome criteria take into account that TC and LDL-C levels differ for adults and children. The criteria also take into account of evidence of dominant transmission and the age of onset of coronary disease in the kindred. Using this approach, cases are categorized as ‘definite’ and ‘possible’. The DLCNC score is based on a family history of hypercholesterolemia, premature CHD, clinical features, LDL-C levels and DNA analysis. It uses a numeric score for each criterion, and classifies individuals as “definite”, “probable” or “possible” FH. A diagnosis is considered definite if the score is greater than 8, considered probable if the score is between 6 and 8 points, and considered possible when the score is between 3 and 5 points. Aims- To determine the frequency and spectrum of mutations causing FH in patients attending the different Italian lipid clinics. To investigate genotype-phenotype correlations in FH carrying different mutations of the LDL-R gene. To identify the presence of a likely polygenic cause due to the inheritance of LDL-C-raising SNPs. In addition, to evaluate which of diagnostic criteria DCLN or Simon Broome is more accurate to detect patients with mutations in our population. Results- Mutations were found in 214/322 subjects with a mutation rate of 66%. Out of 214 mutated subjects about 95.3% were carriers of LDLR mutations, 2.8% of APOB mutations and 1.8% of PCSK9 mutations. In the LDLR gene we found 58 different mutations of which 5 were novels (c.102C>A; c.892A>G; c.1277T>G; c.694+1G>C c.1070_1071dupAG). Regarding the APOB gene we identified four different mutations of which two novels (p.Val3306Ile and p.Trp3633Arg ). In the PCSK9 gene we found four different variations of which two novels (p.Pro331Ala, p.Arg499Hys). The gradually increase of the TC and LDL-C among patients with different types of mutation shows that the type of the LDLR mutation influences the lipid profile. Patients with radical mutations show a worse lipid profile than missense carriers allowing a prognostic evaluation for physicians. In a proportion of patients with the FH phenotype but without mutations in the main candidate genes, there is a likely polygenic cause due to the inheritance of LDL-C-raising SNPs which increases LDL-C concentration in patients. In addition, we examined the reliability of the Dutch Lipid Clinic Network (DCLN) score and Simon Broome criteria, to identify patients with a high or low probability of carrying an FH-causing mutation. The DCLN criteria was more accurate than Simon Broom showing very high sensitivity and specificity to detect patients with mutations. Conclusions- The results of this study represent an update of FH genetic background in an Italian population of patients from southern regions. These data enlarge the spectrum of mutations causing FH. The correlation between mutation types and lipid profile underlines importance of genetic screening as a prognostic tool. The comparison of the Dutch Lipid Clinic Network (DCLN) score and Simon Broome revealed that genetic screening is also useful to confirm the diagnosis, especially in patients with an uncertain phenotype.