Association study of ABCA1 polymorphisms in singapore populations 1

Numerous epidemiological data have shown the risk of CAD is considerably increased in individuals with low levels of plasma high density lipoprotein-cholesterol HDL-C with each 1 mg/dL 0

1 Introduction and Objectives

1.1 Introduction

Coronary artery disease (CAD) is a leading cause of death and disability in many industrialized nations and by 2020, it is projected to be the single most important killer in developing countries (Lopez and Murray, 1998) Numerous epidemiological data have shown the risk of CAD is considerably increased in individuals with low levels of plasma high density lipoprotein-cholesterol (HDL-C) with each 1 mg/dL (0.026 mM) reduction in HDL-C translating into an increased CAD risk of 2-3%, even after adjustment for other lipid and non-lipid risk factors (Gordon and Rifkind, 1989) Plasma HDL-C levels are modulated by both genes and environment, with genetic risk factors alone estimated to contribute 40-70% of the variation in HDL-C in the general population (Ordovas, 2002; Wang and Paigen, 2002) Much of the atheroprotective effect of HDL has been ascribed

to its role in reverse cholesterol transport (RCT), a process by which excess cholesterol

in peripheral tissues is delivered to the liver and steriodogenic organs Genes that are involved in various aspects of HDL metabolism including RCT have been examined for their associations with plasma lipids, especially HDL-C and CAD Generally, for genes that show consistent and significant associations between genotypes and phenotypes, only 5% or less of the variability in HDL is attributed to each locus (Ordovas, 2002)

Tangier disease was the first genetic HDL deficiency disorder to be described (Fredrickson et al., 1961) Independent studies using skin fibroblasts from Tangier disease patients (Walter et al., 1994; Francis et al., 1995; Rogler et al., 1995; Remaley et al., 1997) as well the more common familial hypoalphalipoproteinemia (FHA; Marcil et al., 1999), have shown a marked impairment in cholesterol efflux Prominent biochemical

features in Tangier disease and FHA patients are reduced levels of HDL-C and apolipoprotein AI (ApoAI) Tangier disease patients also show excessive accumulation of cholesterol esters in lymphoid tissues Genetic analyses ruled out any defects in the ApoAI gene in Tangier disease patients (Makrides et al., 1988; Serfaty-Lacrosniere et al., 1994) In 1999, the molecular defect in these HDL deficiency syndromes was mapped to the ABCA1 gene which belongs to the superfamily of ATP binding cassette transporters (Bodzioch et al., 1999; Brooks-Wilson et al., 1999; Lawn et al., 1999; Marcil et al., 1999; Remaley et al., 1999; Rust et al., 1999)

Cholesterol efflux is generally regarded as rate-limiting for HDL biogenesis, and plasma HDL-C levels are consistently inversely correlated with CAD Therefore ABCA1 is an excellent candidate gene to investigate whether common sequence variants in the gene are associated with CAD susceptibility and plasma lipids At the time of this study was initiated in early 2000, knowledge as to the extent of common genetic variations in the ABCA1 gene was confined to only two reports (Pullinger et al 2000; Wang et al., 2000) Far less was known about the contribution of ABCA1 SNPs to inter-individual differences

in CAD susceptibility and intermediate phenotype variation, with only Wang et al (2000) reporting an association of M883I with HDL-C levels in normolipidemic Inuits

The two major objectives of the dissertation were:

1 Identification of ABCA1 polymorphisms that might be useful in a genetic association study; and

2 Association analysis of ABCA1 SNPs with CAD and plasma lipid levels in Singapore Chinese, Malays and Indians

For the SNP discovery phase, we combined in silico and experimental

approaches DHPLC was the method of choice for a quick scan of the coding region of

ABCA1 The ABCA1 proximal promoter was sequenced The human EST database was also mined for potential candidate ABCA1 SNPs using the SNPFINDER program (Buetow et al., 1999)

A case-control approach was adopted for the association study Males from all the three major races of Singapore, namely, Chinese, Malays and Indians, were genotyped for seven selected SNPs in the ABCA1 gene The SNPs were analyzed individually as well as in combination for frequency differences between cases and control groups for each ethnic group separately Associatiosn with concentrations of plasma HDL-C, ApoAI and triglycerides, were also investigated but only in control individuals in order to eliminate confounding due to effects of hypolipidemia treatment in cases By examining individuals from diverse ethnic origins, it might be established if the results of any associations among ABCA1 SNPs, CAD and plasma lipids would be applicable to a more generalized population The results reveal the presence of ethnic-specific ABCA1 associations with CAD as well as plasma lipids