Genetic Study of Low-Density Lipoprotein Receptor Gene and Apolipoprotein B-100 Gene among Malaysian Patients with Familial Hypercholesterolaemia
Keywords:Familial hypercholesterolaemia, LDLR gene, APOB-100 gene, DHPLC, Malaysians
Background: Autosomal dominant hypercholesterolaemia (ADH) is a genetic disorder that is mainly resulted from defects in the low-density lipoprotein receptor (LDLR) and apolipoprotein B-100(APOB-100) genes. Few studies of familial hypercholesterolaemia (FH) have been conducted in Malaysia, that makes the underlying main defect remains not well understood.
Objectives: This study was aimed to describe the molecular aspects of FH among Malaysian subjects.
Methods and Findings: We studied a group of 74 familial hypercholesterolaemic patients and 77 healthy control subjects. The promoter region and the 18 exons of the low-density lipoprotein receptor gene were screened by denaturing high-performance liquid chromatography (DHPLC) to detect small deletions, insertions Â and nucleotide substitutions, while DNA sequencing was applied to look for gene variants in amplicons Â of exon 26 and 29 in the APOB -100 gene. A total of five gene sequence variants in the LDLR gene were reported in 54.1% of the studied patients. P.Arg471Arg variant has the highest frequency of 20.3% among the study subjects. One intronic mutation (c.313+1G>A) and one missense mutation (p.Arg 385Try) were found to be pathogenic, while the other three variants were reported to be non-pathogenic by the in silico analyses. Nine variants were reported in the APOB-100 gene among familial hypercholesterolaemic patients with a non-significant difference in their frequency from the control subjects.
Conclusions: five LDLR gene sequence variants were reported. However, nine polymorphisms were stated in the APOB -100 gene. However they were not associated with FH phenotype among this cohort. These findings suggested that LDLR gene mutation is the major genetic cause for FH among Malaysians. The results offer information on the genetic spectrum of FH among Malaysian cohort which can serve as a platform for further genetic studies.
Austin MA, Hutter CM, Zimmern RL, Humphries SE. Genetic causes of monogenic heterozygous familial hypercholesterolaemia: a HuGE prevalence review. Am J Epidemiol. 2004;160(5):407â€“20.
Hovingh GK, Davidson MH, Kastelein J J P, O'Connor AM. Diagnosis and treatment of familial hypercholesterolaemia. EHJ. 2013; 34(13): 962-971.
Marks D, Thorogood M, Neil HA, Humphries SE. A review on the diagnosis, natural history, and treatment of familial hypercholesterolaemia. Atherosclerosis. 2003;168(1):1-14.
Sjouke B, Kusters DM, Kindt I, Besseling J, Joep C. Defesche, et al. Homozygous autosomal dominant hypercholesterolaemia in the Netherlands: prevalence, genotype-phenotype relationship, and clinical outcome. Eur Heart J. 2015;36(9):560.
Ademi Z, Watts GF, Pang J, Sijbrands EJ, van Bockxmeer FM, O'Leary P, et al. Cascade screening based on genetic testing is cost-effective: evidence for the implementation of models of care for familial hypercholesterolemia. J Clin Lipidol. 2014;8(4):390â€“400.
Marais AD. Familial hypercholesterolaemia. Clin Biochem Rev. 2004;25(1): 49â€“68.
Marianne Benn, Gerald F. Watts, Anne Tybjaerg-Hansen, and BÃ¸rge G. Nordestgaard. Familial hypercholesterolemia in the Danish general population: prevalence, coronary artery disease, and cholesterol-lowering medication. J Clin Endocrinol Metab. 2012; 97(11):3956â€“3964.
Soutar AK, Naoumova RP. Mechanisms of disease: genetic causes of familial hypercholesterolaemia. Nat Clin Pract Cardiovasc. 2007; 4(4): 214â€“225.
Goldstein JL, Brown MS. Binding and degradation of low density lipoproteins by cultured human fibroblasts. Comparison of cells from a normal subject and from a patient with homozygous familial hypercholesterolaemia. J Biol Chem. 1974; 249(16):5153-62.
Hobbs HH, Brown MS, Goldstein JL. Molecular genetics of the LDL receptor gene in familial hypercholesterolaemia. Hum Mutat. 1992;1(6):445-66.
University College London Low Density Lipoprotein Familial Hypercholesterolaemia Database. http://www.ucl.ac.uk/fh.
Soutar AK, Myant NB, Thompson GR. Simultaneous measurement of apolipoprotein B turnover in very-low-and low-density lipoproteins in familial hypercholesterolaemia. Atherosclerosis. 1977; 28(3):247-256.
Schumaker VN, Phillips ML, Chatterton JE. Apolipoprotein B and low-density lipoprotein structure: implications for biosynthesis of triglyceride-rich lipoproteins. Advances in Protein Chemistry. 1994;45:205- 248.
Abifadel M, Varret M, Rabes J, Allard D , Ouguerram K , Devillers M, et al . Mutations in PCSK9 cause autosomal dominant hypercholesterolaemia. Nat Genet. 2003;34(2): 154-6.
Al-Allaf FA, Coutelle C, Waddington SN, David AL, Harbottle R, Themis M . LDLR-Gene therapy for familial hypercholesterolaemia: problems, progress, and perspectives. Int Arch Med. 2010; 1(3)3: 36.
Nordestgaard BG, Chapman MJ,. Humphries SE,Ginsberg HN, Masana L, Descamps OS , et al. Familial hypercholesterolaemia is underdiagnosed and undertreated in the general population: guidance for clinicians to prevent coronary heart disease. Eur Heart J. 2013;34(45):3478â€“90.
Jarvik GP, Brunzell JD, Motulsky AG. Frequent detection of familial hypercholesterolaemia mutations in familial combined hyperlipidaemia . J Am Coll Cardiol. 2008; 52(19): 1554â€“1556
Shi Z, Yuan B, Zhao D, Taylor AW, Lin J, Watts GF. Familial hypercholesterolaemia in China: Prevalence and evidence of underdetection and undertreatment in a community population. Int J Cardiol. 2014;174(3):834â€“836.
Watts GF, Gidding S, Wierzbicki AS, Toth PP, Alonso R, W. Brownh WV, et al. Integrated guidance on the care of familial hypercholesterolaemia from the International FH Foundation. Int J Cardiol. 2014;171(3):309â€“325.
Harada-Shiba M, Arai H, Oikawa S, Ohta T, Okada T, Okamura T, et al. Guidelines for the management of familial hypercholesterolaemia. J Atheroscler Thromb. 2012;19(12):1043â€“1060.
Lye SH, Chahil JK, Bagali P, Alex L, Vadivelu J, Ahmad WA, et al . Genetic Polymorphisms in LDLR, APOB, PCSK9 and other lipid related genes associated with familial hypercholesterolaemia in Malaysia. PLOS ONE. 2013; 8(4): 60729.
Al-Khateeb A, Zahri MK, Mohamed MS, Sasongko TH, Ibrahim S, Yosuf Z, et al. Analysis of sequence variations in low-density lipoprotein receptor gene among Malaysian patients with familial hypercholesterolaemia. BMC Med Genet. 2011; 12: 40.
Khoo KL, Tan H, Liew YM. Familial hyperlipidaemia in Malaysian children. Med J Malaysia. 2000; 55(2): 249â€“258.
Khoo KL, Van Acker P, Tan H, Deslypere JP. Genetic causes of familial hypercholesterolaemia in a Malaysian population. Med J Malaysia. 2000; 55(4): 409â€“418.
Khoo KL, van Acker P, Defesche JC, Tan H, van de Kerkhof L, Heijnen-van Eijk SJ, et al. Low-density lipoprotein receptor gene mutations in a Southeast Asian population with familial hypercholesterolaemia. Clin Genet. 2000;58(2): 98â€“105.
Khoo KL, Tan H, Liew YM, Deslypere JP, Janus E. Lipids and coronary heart disease in Asia. Atherosclerosis. 2003;169(1): 1â€“10.
Azian M, Hapizah MN, Khalid BA, Khalid Y, Rosli A, Jamal R, et al. Use of the denaturing gradient gel electrophoresis (DGGE) method for mutationalscreening of patients with familial hypercholesterolaemia (FH) and Familial defective apolipoprotein B100 (FDB). Malays J Pathol 2006; 28(1): 7â€“15
Scientific Steering Committee on behalf of the Simon Broome Register Group. Risk of fatal coronary heart disease in familial hypercholesterolaemia. Bmj. 1991; 303(6807):893-896.
(NCEP): Executive Summary of The Third Report of The National Cholesterol Education Program (NCEP) Expert Panel on Detection, Evaluation, And Treatment of High Blood Cholesterol In Adults (Adult Treatment Panel III). JAMA. 2001; 285(19):2486-2497.
Friedewald, W. T., Levy, R. I. & Fredrickson, D. S. Estimation of the concentration of low-density lipoprotein cholesterol in plasma, without use of the preparative ultracentrifuge. Clin Chem. 1972;18 (6): 499-502.
Bodamer, O. A., Bercovich, D., Schlabach, M., Ballantyne, C., Zoch, D., & Beaudet, A. L. Use of denaturing HPLC to provide efficient detection of mutations causing familial hypercholesterolemia. Clin Chem. 2002; 48(11): 1913- 1918.
Ramensky V, Bork P, Sunyaev S: Human non-synonymous SNPs: server and survey. Nucleic Acids Res. 2002; 30(17):3894-3900.
A. C. M. Jansen , E. S. Van Aalst-cohen, M. W. Tanck, M. D. Trip , P. J . Lansberg, A. H. Liem, et al . The contribution of classical risk factors to cardiovascular disease in familial hypercholesterolaemia: data in 2400 patients Journal of Internal Medicine. 2004; 256: 482â€“490
S. W., Defesche, J. C., Hutten, B. A., Kastelein, J. J. & Vissers, M. N. Functionality of sequence variants in the genes coding for the low-density lipoprotein receptor and apolipoprotein B in individuals with inherited hypercholesterolaemia. Hum Mutat. 2010; 31(6): 752-60.
Punzalan FE, Sy RG, Santos RS, Cutiongco EM, Gosiengfiao S, Fadriguilan E, et al. A: Low density lipoproteinâ€“receptor (LDL-R) gene mutations among Filipinos with familial hypercholesterolaemia. J Atheroscler Thromb. 2005; 12(5):276-283.
Thiart R, Scholtz CL, Vergotine J, Hoogendijk CF, de Villiers JN, Nissen H,Brusgaard K, et al: Predominance of a 6bp deletion in exon 2 of the LDL receptor gene in Africans with familial hypercholesterolaemia. Journal of medical genetics 2000; 37(7):514-519.
PatrÃcia Mendes TC, Ramos A, Raposo M, Kazachkova N, Margarida Medeiros A, Bruges-Armas J ,et al . Familial hypercholesterolemia: Molecular characterization of possible cases from the Azores Islands (Portugal). Meta Gene. 2014; 2:638â€“645
Zakharova FM1, Golubkov VI, Mandel'shtam MIu, LipovetskiÄ BM, GaÄtskhoki VS. Identification of novel missense mutation G571E, novel silent mutation H229H, nonsense mutation C74X, and four single nucleotide polymorphisms in the low-density lipoprotein receptor in patients with familial hypercholesterolemia from St. Petersburg. Bioorg Khim. 2001;27(5):393-6.
Shapiro MB, Senapathy P. RNA splice junctions of different classes of eukaryotes: sequence statistics and functional implications in gene expression. Nucleic Acids Res 1987;15(17):7155-7174.
Cameron J, Holla OL, Kulseth MA, Leren TP, Berge KE. Splice-site mutation c.313Ã¾1, G.A in intron 3 of the LDL receptor gene results in transcripts with skipping of exon 3 and inclusion of intron 3. Clin Chim Acta. 2009;403:131â€“5
Alicezah MK, Razali R, Rahman T , Hoh BP, Suhana NH, HM Nawawi ,et al . Homozygous familial hypercholesterolemia. Malaysian J Pathol 2014; 36(2) : 131 â€“ 137
Reshef, A., Nissen, H., Triger, L., Hensen, T. S., Eliav, O., Schurr, D., et al. Molecular genetics of familial hypercholesterolaemia in Israel. Hum Genet. 1996; 98(5): 581-586.
Yu. A. Tatishcheva1, M. Yu. Mandelshtam1, V. I. Golubkov1, B. M. Lipovetsky, and V. S. Gaitskhoki. Four New Mutations and Two Polymorphic Variants of the Low-Density Lipoprotein Receptor Gene in Familial Hypercholesterolemia Patients from St. Petersburg. Russian Journal of Genetics. 2001; 37( 9): 1082â€“1086
Sauna, Z. E. & Kimchi-Sarfaty, C. Understanding the contribution of synonymous mutations to human disease. Nat Rev Genet. 2011; 12: 683â€“691
Chmara M, Wasag B, Zuk M, Kubalska J, Wegrzyn A, Bednarska-Makaruk M, et al : Molecular characterization of Polish patients with familial hypercholesterolaemia: novel and recurrent LDLR mutations. J Appl Genet. 2010; 51(1):95-106.
Al-Khateeb A, Mohamed M, Yusof Z, Zilfalil B. Molecular description of familial defective APOB-100 in Malaysia. Biochem Genet. 2013;51 (9-10):811â€“823.
Innerarity TL, Weisgraber KH, Arnold KS, Mahley RW, Krauss RM, Vega GL, et al. Familial defective apolipoprotein B-100: low-density lipoproteins with abnormal receptor binding .Proc Natl Acad Sci USA. 1987; 84 (19):6919â€“6923.
Hansen PS. Familial defective apolipoprotein B-100. Dan Med Bull. 1998; 45(4):370â€“382.
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