Cholesteryl Ester Transfer Protein Gene Mutations Research Articles

Novel mutations of cholesteryl ester transfer protein (CETP) gene in Japanese hyperalphalipoproteinemic subjects

BackgroundThe half of hyperalphalipoproteinemia (HALP) in Japan is caused by CETP gene mutations. Other than two prevalent mutations (D442G and Intron 14 splicing donor site +1G>A), some rare CETP mutations are found in Japanese HALP subjects. MethodsCETP gene analysis of genomic DNA from subjects was performed by restriction fragment length polymorphism (RFLP) and sequencing analysis. Mutations which were suspected to cause a splicing defect or a protein secretion defect were investigated in COS-1 cells transfected with a CETP minigene construct or a cDNA expression vector. ResultsEach of three subjects was identified as a carrier of CETP gene mutation of a compound heterozygote of c.653_654delGGinsAAAC and Intron 14 splicing donor site +1G>A, a heterozygote of c.658G>A or a homozygote of L261R. The c.658G>A mutation was located at the last nucleotide of exon 7, and it was confirmed to cause splicing abnormality revealed by the CETP minigene analysis. The L261R CETP was not secreted to conditioned media of the cells. ConclusionsThree novel CETP gene mutations are responsible for HALP by CETP deficiency. It is predicted that there are more rare CETP gene mutations in Japanese, and these multiple rare mutations alone or a combination with each of prevalent mutations is responsible for mild-to-moderate or marked HALP, respectively.

A novel homozygous mutation in CETP gene as a cause of CETP deficiency in a caucasian kindred

Objective To analyze the cholesteryl ester transfer protein (CETP) gene and the plasma HDL phenotype in a Caucasian subject with extremely elevated plasma high density lipoprotein-cholesterol (HDL-C). Methods and results The proband, a 63-year-old male of Swedish ancestry with elevated HDL-C (208 mg/dl) and apoA-I (and 272 mg/dl), was found to be homozygous for a point mutation in exon 2 of CETP gene (c.109 C > T) resulting in a premature termination codon (R37X). Plasma CETP mass and activity were undetectable. Plasma HDL were characterized by predominance of large HDL with enhanced preβ-HDL content. The proband's sons, heterozygotes for the mutation, had reduced plasma CETP activity and moderately elevated HDL-C. Serum of CETP deficient subjects showed a normal or enhanced cholesterol efflux capacity via ABCG1/SR-BI; cholesterol efflux via ABCA1 and macrophage cholesterol removal were lower than normal. The proband was healthy and had no atherosclerotic plaques in carotid or femoral arteries. Conclusion Complete CETP deficiency caused by mutations in CETP gene is exceedingly rare in Caucasians; the description of this single case indicates that CETP deficiency does not predispose to atherosclerosis in the absence of major cardiovascular risk factors.

CETP gene mutation (D442G) increases low-density lipoprotein particle size in patients with coronary heart disease

Background: Small, dense low-density lipoprotein (LDL) in subjects with the atherogenic pattern B has been established as a risk factor of atherosclerosis. Cholesteryl ester transfer protein (CETP) plays an important role in the transfer and exchange of cholesteryl esters and triglycerides between the lipoprotein classes of human plasma. It has been shown that CETP can also change the particle size of high-density lipoprotein (HDL) and LDL subfractions in vitro. Previous clinical studies about CETP gene mutations mainly focused on abnormalities in HDL, few involved those in LDL. Objectives: To investigate the effect of the D442G mutation in the CETP gene on major peak size of LDL particles in patients with coronary heart diseases (CHD). Methods: D442G mutation in the CETP gene was detected using the PCR-RFLP. LDL particles sizes were analyzed by 2–16% nondenaturing polyacrylamide gradient gels in CHD patients with D442G mutation in the CETP gene. Results: Six heterozygotes and one homozygote were found to have the D442G mutation among 200 CHD patients. The frequency of this mutation was 3.5%. The major peak size of LDL in patients with gene mutation ( n=7) was significantly larger than that in patients without the mutation ( n=40) (26.92±0.79 nm vs. 25.71±0.66 nm, respectively; P<0.01). All the patients with the gene mutation expressed pattern A, whereas only about half of the patients without the mutation expressed this pattern. The patients with gene mutation had decreased plasma CETP concentration, while increased concentration of HDL-C and apolipoprotein A-I compared with controls. Conclusions: CETP gene mutation (D442G) increases LDL particle size. This suggests that CETP play an antiatherogenic role.

Effect of a common mutation (D442G) of the cholesteryl ester transfer protein gene on lipids and lipoproteins in children.

Cholesteryl ester transfer protein (CETP) is thought to regulate plasma HDL. Patients with CETP deficiency caused by mutation of the CETP gene [D442G; a missense mutation (Asp442-->Gly)] have been reported to show high plasma HDL levels. However, there are no data available on children with D442G. To determine the effects of plasma CETP and CETP gene mutation (D442G) on lipids and lipoproteins in children, we screened children by PCR and restriction fragment length polymorphism analysis of the CETP gene. Plasma lipids, apolipoproteins, and CETP mass levels were also determined. In the current study, 22 children with D442G were found (21 heterozygotes and a homozygote). A homozygous child showed high plasma HDL level and very low plasma CETP mass. In heterozygous children, plasma concentrations of HDL cholesterol, apo A-I and apo A-II were not increased, whereas plasma CETP mass was significantly decreased. Plasma CETP mass in heterozygous children was correlated with plasma concentrations of total cholesterol, LDL cholesterol, and apo B. Plasma CETP mass in children without D442G was not correlated with the plasma concentration of any lipid or apolipoprotein. All of these data suggest that the D442G mutation, by itself, might not affect HDL metabolism in children. The CETP mass required for efficient HDL-cholesteryl ester clearance in children may be less than that in older subjects.

Effects of hepatic lipase gene promoter nucleotide variations on serum HDL cholesterol concentration in the general Japanese population.

Genetic factors may play a major role in determining serum high-density lipoprotein (HDL) cholesterol (HDL-C) levels in the general population. Cholesteryl ester transfer protein (CETP) is a strong genetic factor as a determinant of HDL-C levels in Japanese, whereas hepatic lipase (HL) plays a predominant role in Caucasian populations. We investigated the effects of HL gene promoter polymorphisms on HDL-C levels in a general population of Japanese men (n = 299). An HL promoter polymorphism of -514C/T explained a considerable variance of HDL-C (2.9%), as compared with CETP mutations of D442G and IVS14 + 1G > A (3.6% and 1.9%). HL promoter variation of the -514C/C genotype, reported to have high HL activity, had significant effects on decreasing HDL-C levels (-3.8mg/dl), but -514T allele carriers had a weak effect on increasing HDL-C levels. The frequency of the -514T allele was three times higher (0.50) in the Japanese than in Caucasian populations (0.15-0.19). Thus, the higher frequency of the HL -514T allele, along with CETP gene mutations, could explain about 9% of phenotypic variability of HDL-C. These genetic attributes may be among the many factors that contribute to the relatively higher serum HDL-C levels in Japanese subjects.

Cholesteryl ester transfer protein deficiency is correlated with thyroid disease.

We investigated whether cholesteryl ester transfer protein (CETP) deficiency is a risk factor for atherosclerosis or other disease. Ninety-seven individuals with increased levels of high density lipoprotein-cholesterol (HDL-C) (≥85mg/dl) were selected. CETP deficiency was assessed in all 97 subjects by polymerase chain reaction (PCR) and either single stranded conformation polymorphism (SSCP) or restriction fragment length polymorphism (RFLP). Of the 97 subjects, 7 (7%) were heterozygous for the intron 14 splicing defect, which decreases CETP activity, and 9 (9%) were heterozygous for D442G missense mutation, which has no effect on CETP activity. One patient was found to be a compound heterozygote for the intron 14 and D442G mutations. These mutations were found in only 3 atherosclerotic patients; 2 carried the D442G mutation, and 1 had the intron 14 mutation. Although CETP activity was not associated with atherosclerosis, we discovered a novel relation between CETP gene mutations and thyroid gland disorders. Specifically, 5 of 7 subjects with CETP gene intron 14 mutation had thyroid disease. Regarding the D442G mutation, there was no difference between subjects with and without thyroid disease. It is possible that CETP activity affects thyroid function.

A Low Prevalence of Coronary Heart Disease among Subjects with Increased High-Density Lipoprotein Cholesterol Levels, Including Those with Plasma Cholesteryl Ester Transfer Protein Deficiency

Background.Use of genetic analysis may improve the predictive value of risk factors for disease. A high plasma level of high-density lipoprotein (HDL) cholesterol is a strong negative risk factor for coronary heart disease (CHD). Cholesteryl ester transfer protein (CETP) deficiency causes increased levels of HDL cholesterol. However, recent studies suggest that CETP deficiency is a risk factor for CHD despite elevated HDL cholesterol levels.Methods.Plasma lipid levels, CHD prevalence, resting electrocardiograms, and common CETP gene mutations were analyzed cross-sectionally in a population of 19,044 male and 29,487 female Japanese subjects (ages 45–79 years).Results.High HDL cholesterol levels (serum HDL cholesterol ≥80 mg/dl, ≥95th percentile) were found in 6 and 5% of Japanese men and women, respectively. In the group with HDL cholesterol ≥80 mg/dl, common CETP gene mutations were identified in 23–24% of men and 31–49% of women. The prevalence of CHD in the group with high HDL cholesterol (≥80 mg/dl) was low among both men (1.0%) and women (1.3%). There was no difference in CHD prevalence between hyper-HDL-cholesterolemic subjects with and without CETP mutations.Conclusions.Subjects with very high HDL levels (HDL cholesterol ≥80 mg/dl) as well as mild-to-moderate HDL elevations (60–79 mg/dl) appear to be protected against CHD, whether or not they have CETP deficiency, a genetic cause of elevated HDL.

Relationship of HDL and coronary heart disease to a common amino acid polymorphism in the cholesteryl ester transfer protein in men with and without hypertriglyceridemia

Plasma triglyceride (TG) levels are inversely related to HDL-cholesterol levels and subjects with high TG and low HDL cholesterol have increased coronary heart disease (CHD) risk. Plasma cholesteryl ester transfer protein (CETP) transfers cholesteryl esters from HDL to TG-rich lipoproteins. In this study we determined the relationship between a common CETP amino acid polymorphism (I405V) and CETP and HDL levels and CHD prevalence in 576 men of Japanese ancestry in the Honolulu Heart Program cohort. This conservative substitution was associated with altered plasma CETP concentration (1.95 +/- 0.54, 1.91 +/- 0.57, and 1.77 +/- 0.57 microg/ml for the II, IV and VV genotypes, respectively). The distribution of plasma CETP concentrations among the VV, but not II, men appeared bimodal (P < 0.01), suggesting the presence of a functionally significant CETP gene mutation(s) in a subset of V alleles. HDL-C levels were higher in VV than IV for II men (55.4 +/- 17.4, 51.3 +/- 16.6, 51.1 +/- 17.0 mg/dl, P < 0.04). However, the increase in HDL was only significant in VV men with plasma TG > 165 mg/dl. Although CHD prevalence was not significantly different among the three genotypes in this population, in the subpopulation with high plasma TG, CHD prevalence appeared higher among VV than IV or II subjects (38% vs. 27% vs. 18%, P < 0.05 for an interaction of genotype and plasma TG levels). In fresh plasma from a separate group of normolipidemic subjects, the V/I polymorphism was not associated with any change in plasma CETP specific activity. The data suggest that a widespread and common CETP gene mutation(s) in linkage disequilibrium with 405V causes low CETP. Among hypertriglyceridemic men this is associated with higher HDL and possibly with increased CHD.

Enzyme immunoassay for cholesteryl ester transfer protein in human serum

We developed a new simple sandwich-type enzyme immunoassay to measure cholesteryl ester transfer protein (CETP) mass in human serum. In assay validation, Intra- and Inter-assay coefficients of variation were 2.7 to 5.7% and 2.2 to 12.2%, respectively. There was no cross-reactivity with various lipoproteins (apo A-I, apo A-II, apo B, apo C-III). A good correlation between CETP mass and CETP activity (n=46, correlation coefficient=0.88) was observed. This assay provided a specific and reproducible method for measuring CETP mass in samples. The average value of CETP in the normal sera of 41 males was 1.8±0.6 μg/ml (mean±S.D.) and that of 37 females was 2.0±0.5 μg/ml. In the study of patients with the CETP gene mutation (Int 14A and D442G), our results on the value of plasma CETP mass reflected to genetic CETP deficiency. In conclusion, this assay for CETP mass in human serum may be a useful tool for clinical investigations involving lipid metabolism related to disease.

Genetic cholesteryl ester transfer protein deficiency is extremely frequent in the Omagari area of Japan. Marked hyperalphalipoproteinemia caused by CETP gene mutation is not associated with longevity.

Low levels of HDL cholesterol have been clearly demonstrated to be associated with an increased incidence of coronary heart disease, strongly suggesting that HDL particles have an antiatherogenic function. However, little information has been available concerning the atherogenicity of a marked hyperalphalipoproteinemia (HALP). There is no agreement about whether plasma cholesteryl ester transfer protein (CETP) deficiency is associated with an antiatherogenic state or not, although this disorder was reported to be one of the major causes of marked HALP. In the current study, we have found a unique area (Omagari City, Akita Prefecture, Japan) where CETP deficiency caused by a G-to-A mutation at the 5' splice donor site of intron 14 in the CETP gene is extremely frequent. In Omagari City, the mutation was detected more than 20 times more frequently and the prevalence of a marked HALP with plasma HDL cholesterol > or = 2.58 mmol/L (100 mg/dL) was 5 to 10 times higher than in other areas of Japan. This discovery has made it possible to perform a large population-based study concerning the atherogenicity of a marked elevation of HDL cholesterol in a genetically more homogeneous population. There was a statistically significant U-shaped relationship between HDL cholesterol levels and the incidence of ischemic changes in electrocardiograms. In cases of HDL cholesterol < 1.81 mmol/L (70 mg/dL), the incidence increased in proportion to the levels of HDL cholesterol. The frequency of the CETP gene mutation was higher in patients with coronary heart disease than in healthy control subjects. In subjects aged > 80 years, the prevalence of both marked HALP and the intron 14 splicing defect was significantly lower than in the younger generation. The current study indicated for the first time that a marked HALP caused by CETP gene mutation may not represent a longevity syndrome, suggesting the importance of reevaluation of the clinical significance and pathophysiology of a marked HALP.

Cholesteryl ester transfer protein activity enhances plasma cholesteryl ester formation. Studies in CETP transgenic mice and human genetic CETP deficiency.

The plasma cholesteryl ester transfer protein (CETP) promotes the removal of HDL cholesteryl esters and is thought to stimulate reverse cholesterol transport (RCT). However, mechanisms by which CETP may stimulate RCT are poorly understood. Thus, we examined the relationship between plasma CETP expression and plasma cholesteryl ester formation in CETP transgenic (Tg) mice, hamsters, and human subjects with genetic CETP deficiency. Incubation of CETP Tg mouse plasma showed a 20% to 40% increase in plasma cholesterol esterification rate (CER, P < .05) compared with control mice. Injection of a neutralizing CETP monoclonal antibody (MAb) (TP2) into natural flanking region CETP Tg mice resulted in an increase in plasma free cholesterol (FC) concentration, FC/CE ratio, FC/phosphatidylcholine ratio, and hepatic CETP mRNA. In hamsters, CETP inhibition also resulted in an increase in plasma FC/phosphatidylcholine ratio and increased CETP mRNA in adipose tissue. In humans with two common CETP gene mutations (an intron 14 splicing defect and a D442G missense mutation), mean plasma CERs were 39 and 60, respectively, compared with 89 nmol x mL-1 x h-1 in normal subjects. By contrast, lecithin:cholesterol acyltransferase (LCAT) mass was normal in CETP-deficient subjects. MAb neutralization of CETP activity in incubated human plasma did not alter the LCAT reaction, even after supplementation with discoidal HDL and VLDL. Thus, genetic alterations in CETP levels lead to secondary changes in the plasma LCAT reaction, possibly because of remodeling of HDL by CETP acting in concert with other factors in vivo. In human genetic CETP deficiency, a moderate impairment in the plasma LCAT reaction may contribute to a defect in RCT, providing a potential mechanism to explain the recently observed excess of coronary heart disease in these subjects.

Increased coronary heart disease in Japanese-American men with mutation in the cholesteryl ester transfer protein gene despite increased HDL levels.

Plasma high density lipoprotein (HDL) levels are strongly genetically determined and show a general inverse relationship with coronary heart disease (CHD). The cholesteryl ester transfer protein (CETP) mediates the transfer of cholesteryl esters from HDL to other lipoproteins and is a key participant in the reverse transport of cholesterol from the periphery to the liver. A high prevalence of two different CETP gene mutations (D442G, 5.1%; intron 14G:A, 0.5%), was found in 3,469 men of Japanese ancestry in the Honolulu Heart Program and mutations were associated with decreased CETP (-35%) and increased HDL chol levels (+10% for D442G). However, the overall prevalence of definite CHD was 21% in men with mutations and 16% in men without mutations. The relative risk (RR) of CHD was 1.43 in men with mutations (P < .05); after adjustment for CHD risk factors, the RR was 1.55 (P = .02); after additional adjustment for HDL levels, the RR was 1.68 (P = .008). Similar RR values were obtained for the D442G mutation alone. Increased CHD in men with mutations was primarily observed for HDL chol 41-60 mg/dl; for HDL chol > 60 mg/dl men with and without mutations had low CHD prevalence. Thus, genetic CETP deficiency appears to be an independent risk factor for CHD, primarily due to increased CHD prevalence in men with the D442G mutation and HDL cholesterol between 41 and 60 mg/dl. The findings suggest that both HDL concentration and the dynamics of cholesterol transport through HDL (i.e., reverse cholesterol transport) determine the anti-atherogenicity of the HDL fraction.

Cholesteryl ester transfer protein: an enigmatic protein.

The cholesteryl ester transfer protein (CETP) mediates the transfer of cholesteryl esters from high-density lipoproteins (HDL) into very-low-density lipoproteins (VLDL) with a reciprocal exchange of triglycerides. Plasma CETP is mainly bound to HDL and is involved in the interconversion of these lipoproteins. In experimental models such as transgenic mice, CETP activity decreases HDL cholesterol and increases the cholesteryl ester content of apo B-containing lipoproteins. In humans, CETP activity and concentration are positively correlated with VLDL-LDL cholesterol. Clinical studies suggest that the effect of CETP on HDL cholesterol depends on the amount of acceptor lipoproteins. CETP activity is negatively correlated with HDL cholesterol only in hypertriglyceridemic states. Various CETP gene mutations have been reported, they induce hyper-alpha-lipoproteinemia. On the other hand, the impact of the variability of CETP gene on HDL cholesterol variations in Caucasians is controversial. CETP is often involved in secondary dyslipidemia and is susceptible to modify the composition of plasma lipoproteins in an atherogenic way. The real impact of CETP activity on atherosclerosis is still unknown. CETP is susceptible to play a proatherogenic role since it mediates a redistribution of plasma cholesterol from lipoproteins associated with a protection against atherosclerosis into the proatherogenic apo B-containing lipoproteins. However, CETP mediates one of the steps of the reverse cholesterol transport, an antiatherogenic process that channels cholesterol from peripheral tissues back to the liver.

Cholesteryl ester transfer protein deficiency: Identification in the Chinese

Cholesteryl ester transfer protein (CETP) regulates cholesterol content in high-density lipoproteins (HDL) through the exchange of cholesteryl ester in HDL and triglyceride in triglyceride-rich lipoproteins. Three types of CETP deficiency have so far been reported, all in Japanese individuals. Among them, two types of mutation in the CETP gene are relatively common: (1) a guanine to adenine mutation in the intron 14 splicing donor site (114A), and (2) an Asp 442 to Gly (D442G) mutation. The 114A mutation has an estimated allele frequency of 0.81% in the Japanese. It elevates the plasma HDL-cholesterol level strikingly: typically, >120 mg/dl for homozygotes and 80-110 mg/dl for heterozygotes. In contrast, the D442G mutation has a higher estimated allele frequency (4.62%), but has a mild effect on the HDL-cholesterol level: typically, >80 mg/dl for homozygotes and 50-100 mg/dl for heterozygotes. We screened Chinese individuals for the two CETP gene mutations. 7 refs., 1 tab.

Atherosclerotic Disease in Marked Hyperalphalipoproteinemia

Hyperalphalipoproteinemia (HALP) has been regarded as a beneficial state accompanied by a longevity syndrome. However, we reported the cases of markedly hyperalphalipoproteinemic subjects with juvenile corneal opacification. The patients had reduced postheparin hepatic triglyceride lipase (HTGL) activities, and one of them has recently been identified to be homozygous for a missense mutation in exon 15 (D442: G) in the cholesteryl ester transfer protein (CETP) gene. In the current study, to elucidate the clinical significance of and atherogenicity in marked HALP, we determined the incidence of atherosclerotic cardiovascular disease (ACD) in patients with marked HALP and characterized the lipoprotein abnormalities in those who had ACD, focusing especially on CETP and HTGL. The subjects were 201 patients (111 males and 90 females) with marked HALP ( > or = 2.58 mmol/L [100 mg/dL]), 67% of whom were demonstrated to have the CETP gene mutations in the intron 14 splice donor site or in exon 15. Their mean age was 54 +/- 15 years. Plasma levels of total cholesterol, HDL cholesterol, and triglyceride in all subjects were 6.28 +/- 1.78, 3.15 +/- 0.90, and 1.08 +/- 0.53 mmol/L, respectively. Ten of the male patients (9.0%) and two of the female patients (2.2%) had apparent ACD such as myocardial infarction, angina pectoris, and peripheral vascular diseases. Ten patients with HALP who had ACD were identified to be heterozygotes for CETP deficiency. To further clarify the characteristics of marked HALP in patients with ACD, we compared the plasma lipids, lipoproteins, CETP, and HTGL activities between heterozygotes for CETP deficiency who were with and without ACD.

Enzyme-linked immunosorbent assay for cholesteryl ester transfer protein in human serum

Cholesteryl ester transfer protein (CETP) plays an important role in the reverse cholesterol transport system. An ELISA for serum CETP protein mass was developed. We have established monoclonal antibodies for CETP and used two different clones to develop the ELISA. Maximum CETP immunoreactivity was obtained in serum samples to which 0.5% SDS was added. The assay range by this method was from 30 to 2000 ng/ml. The CETP level in sera of normolipemic volunteers determined by this method was 1680 ± 700 ng/ml (mean ± S.D. of 101 normolipemic volunteers). Mean intra- and inter-assay coefficients of variation were 6–8 and 9–12%, respectively. CETP mass determined by this ELISA correlated well with cholesteryl ester transfer activity ( r = 0.77). We also tested serum samples of patients with CETP gene mutations and found that the CETP mass decreased among the patients with homozygous and heterozygous mutation. This method will help clarify the role of CETP in the metabolism of human HDL and LDL.

Plasma cholesteryl ester transfer protein and high-density lipoproteins: new insights from molecular genetic studies.

Recent studies in transgenic mice provide strong evidence for a direct anti-atherogenic role of high-density lipoproteins (HDL) and highlight the importance of multiple gene interactions in the regulation of HDL levels. Plasma lipid transfer processes mediated by cholesteryl ester transfer protein (CETP) have a major impact on HDL levels, as revealed in studies of human genetic CETP deficiency and CETP transgenic mice. Subsequent to the discovery of an intron 14 CETP gene splicing defect, several new CETP gene mutations have been discovered recently in Japanese and other populations. One of these is an exon 15 missense mutation, changing amino acid 442 of CETP from aspartate to glycine. Population studies in Japan indicate that CETP gene mutations are sufficiently common to have a significant influence on HDL levels in the general population. Studies in transgenic mice show that CETP expression results in decreased levels of HDL cholesterol, but that the effects of CETP on HDL apolipoprotein A-I (apoA-I) content and size show important modulation by co-expression with transgenes encoding human apoA-I, apoC-III and apoA-II. In addition to the apparent antiatherogenic phenotype of human genetic CETP deficiency, high level expression of CETP in transgenic mice leads to accelerated atherosclerosis, illustrating the pro-atherogenic potential of CETP expression.

Frequency of intron 14 splicing defect of cholesteryl ester transfer protein gene in the Japanese general population — relation between the mutation and hyperalphalipoproteinemia

Cholesteryl ester transfer protein (CETP) deficiency, which has been found only in Japan, is characterized by marked hyperalphalipoproteinemia (HALP) and abnormalities of both low density and high density lipoproteins. We have reported that this deficiency is commonly associated with a G→A mutation at the intron 14 splice donor site of the CETP gene (Yamashita et al., Biochem. Biophys. Res. Commun., 170 (1990) 1346–1351). In the current study, we determined the frequency of this mutation in Japanese subjects by using polymerase chain reaction. A single primer-template mismatch of one base pair from the CETP gene mutation permitted the introduction of a cleavage site for Nde I in mutant alleles but not in normal ones. Out of 171 patients with marked HALP whose serum HDL-cholesterol was more than 100 mg/dl, 6 (3.5%) subjects were homozygous and 48 (28.1%) were heterozygous for this mutation. Furthermore, in unrelated 512 healthy Japanese subjects, 5 (0.98%) were identified as heterozygotes. Relative allelic frequency of A at the intron 14 splice donor site was 0.0049 and the frequency of homozygous CETP deficiency was estimated to be approximately 1/42 000. These results demonstrate that this common mutation may be frequent in the Japanese population. Although HALP is very heterogenous, this mutation could be one of the major causes of marked HALP.

Increased High-Density Lipoprotein Levels Caused by a Common Cholesteryl-Ester Transfer Protein Gene Mutation

The plasma cholesteryl-ester transfer protein (CETP) catalyzes the transfer of cholesteryl esters from high-density lipoprotein (HDL) to other lipoproteins. We recently described a Japanese family with increased HDL levels and CETP deficiency due to a splicing defect of the CETP gene. To assess the frequency and phenotype of this condition, we screened 11 additional families with high HDL levels by means of a radioimmunoassay for CETP and DNA analysis. We found the same CETP gene mutation in four families from three different regions of Japan. Analysis of restriction-fragment-length polymorphisms of the mutant CETP allele showed that all probands were homozygous for the identical haplotype. Family members homozygous for CETP deficiency (n = 10) had moderate hypercholesterolemia (mean total cholesterol level [+/- SD], 7.01 +/- 0.83 mmol per liter), markedly increased levels of HDL cholesterol (4.24 +/- 1.01 mmol per liter) and apolipoprotein A-I, and decreased levels of low-density lipoprotein cholesterol (1.99 +/- 0.80 mmol per liter) and apolipoprotein B. Members heterozygous for the deficiency (n = 20), whose CETP levels were in the lower part of the normal range, had moderately increased levels of HDL cholesterol and apolipoprotein A-I and an increased ratio of HDL subclass 2 to HDL subclass 3, as compared with unaffected family members (1.5 +/- 0.8 vs. 0.7 +/- 0.4). CETP deficiency was not found in six unrelated subjects with elevated HDL cholesterol levels who were from different parts of the United States. CETP deficiency appears to be a frequent cause of increased HDL levels in the population of Japan, possibly because of a founder effect. The results that we observed in heterozygotes suggest that CETP normally plays a part in the regulation of levels of HDL subclass 2. There was no evidence of premature atherosclerosis in the families with CETP deficiency. In fact, the lipoprotein profile of persons with CETP deficiency is potentially antiatherogenic and may be associated with an increased life span.