Identification and Characterization of Gene Mutations in Patients with Familial Hypercholesterolemia in Taiwan

Autor: Kuan-Rau Chiou, 邱寬饒
Rok vydání: 2006
Druh dokumentu: 學位論文 ; thesis
Popis: 94
Familial hypercholesterolemia (FH) is an autosomal dominant disease associated with a very high risk of coronary vascular disease. FH is mainly caused by mutations in the low-density liporotein (LDL) receptor and apolipoprotein B (apoB); it is characterized by a high concentration of LDL. The objectives of this study are firstly to set up a molecular genetic program for identification of index patients and further screening of affected family members, and secondly to characterize the LDL receptor gene mutations identified from the patients with FH. A total of 51 unrelated patients, all fitting the Simon Broome Familial Hypercholesterolemia Register diagnostic criteria, were screened for mutations in both the LDL receptor and apoB-100 genes, employing the multiplex polymerase chain reaction (PCR) and exon-by-exon DNA sequencing technique. The promoter region and all 18 exons of the LDL receptor, as well as the ligand binding domains in exon 26 and 29 of the apoB-100 gene, were amplified through PCR and were fully sequenced. A total of 14 different functional mutations in the LDL receptor gene and one mutation in the apoB-100 gene were found in 21 patients (detection rate 41%). Among these mutations, eleven had been previously reported, including D69N, D206E, D207X, C308Y, R395W, I420T, G457R, H562Y, A606T, R236W/D568N, and R3500W; three novel mutations were identified, including two missense mutations: M510K (T-to-A substitution at nucleotide (nt) 1592; codon 510; exon 11) and W512R (T-to-C substitution at nt 1597;codon 512; exon 11), respectively, and one frame shift mutation: 1953 delTA (2-bp deletion at nt1953, 1954; codon 631; exon 13). Segregation of the mutation among family members was present in each adult case. To further characterize the three novel mutations, the site-directed mutagenetic LDLR genes (pcDNA3-LDLR and its mutant derivatives) were transfected into COS-7 cells. For missense M510K, 36.2% of normal receptor activities were seen. Conversely, frameshift 1953 delTA and missense W512R led to defective proteins, with only 0-5% of normal receptor activity. Molecular genetics clearly identify index patients and family members carrying mutations. Employing this methodology, we identified 21 families with gene mutations and characterized three novel mutations causing FH in Taiwan. This facilitates a better understanding of FH among the Chinese population and makes possible diagnosis of FH at the molecular level at a presymptomatic, early ag
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