FGF receptor genes and breast cancer susceptibility: results from the Breast Cancer Association Consortium.

BACKGROUND: Breast cancer is one of the most common malignancies in women. Genome-wide association studies have identified FGFR2 as a breast cancer susceptibility gene. Common variation in other fibroblast growth factor (FGF) receptors might also modify risk. We tested this hypothesis by studying genotyped single-nucleotide polymorphisms (SNPs) and imputed SNPs in FGFR1, FGFR3, FGFR4 and FGFRL1 in the Breast Cancer Association Consortium. METHODS: Data were combined from 49 studies, including 53 835 cases and 50 156 controls, of which 89 050 (46 450 cases and 42 600 controls) were of European ancestry, 12 893 (6269 cases and 6624 controls) of Asian and 2048 (1116 cases and 932 controls) of African ancestry. Associations with risk of breast cancer, overall and by disease sub-type, were assessed using unconditional logistic regression. RESULTS: Little evidence of association with breast cancer risk was observed for SNPs in the FGF receptor genes. The strongest evidence in European women was for rs743682 in FGFR3; the estimated per-allele odds ratio was 1.05 (95% confidence interval=1.02-1.09, P=0.0020), which is substantially lower than that observed for SNPs in FGFR2. CONCLUSION: Our results suggest that common variants in the other FGF receptors are not associated with risk of breast cancer to the degree observed for FGFR2.

The -629C>A polymorphism in the CETP gene does not explain the association of TaqIB polymorphism with risk and age of myocardial infarction in Icelandic men.

The aim of this study was to examine whether the well-established effect of the common TaqIB polymorphism in intron 1 of the gene for cholesterol ester transfer protein (CETP) on high density lipoprotein cholesterol (HDL-C) concentration and increased risk of myocardial infarction (MI), could be explained by the recently identified -629C>A functional polymorphism in the promoter. Non-fatal MI cases (388 male) and a control group of 794 healthy men were recruited from the 30 year long prospective Reykjavik Study. In the healthy men the frequency of the TaqIB B2 allele was 0.47 (95% CI: 0.44-0.50) and there was a strong allelic association with the -629A allele (D=-0.21, P<0.0001), which had a frequency of 0.52 (95% CI: 0.49-0.56). B2B2 homozygotes displayed 15% higher HDL-C levels than subjects homozygous for the B1 allele (P<0.0001). Homozygotes for the -629A allele displayed 14% higher HDL-C concentrations than subjects homozygous for the -629C allele (P<0.0001). The frequencies of the alleles associated with lower HDL-C were significantly higher in cases compared with controls, 0.59 versus 0.53 (TaqIB B1) and 0.52 versus 0.48 (-629 C) respectively (P<0.05 for both). There was a significantly higher risk for MI in B1B1 homozygotes (OR=1.44, 95% CI: 1.10-1.87, P<0.01), compared to the other genotypes combined. This was not observed for the CC homozygotes (OR=1.16, 95% CI: 0.87-1.54). In addition, homozygotes for the TaqI B2 allele experienced a first MI 2 years later than men with other genotypes, 59 versus 61 years (P<0.05). This effect was not seen for the promoter polymorphism. These results strongly confirm the role of the CETP gene and the TaqIB variant as a risk factor for MI and suggest that another functional polymorphism is yet to be discovered in the CETP gene, that will explain the effect on MI associated with TaqIB observed in this study.

Analysis of the association of a heat shock protein70-1 gene promoter polymorphism with myocardial infarction and coronary risk traits.

Heat shock proteins (HSP) are induced during coronary ischaemia, and abnormal expression of one HSP gene may cause hypertension in rats. We examined association of a promoter polymorphism in the major stress-inducible hsp70 gene (hsp70-1 or HSP70A1) on chromosome 6 (p21.3) with coronary disease traits. This C-->A base substitution (AAACCCC) is at nucleotide position-110 in the heat shock transcription factor binding site (heat shock element, HSE). The first study sample (ECTIM), recruited from Belfast and three centers in France, consisted of 578 myocardial infarction cases and 698 age-matched controls. The frequency of the A-110 allele was 0.381 (95% CI = 0.35-0.41) and 0.384 (95% CI = 0.36-0.41) in cases and controls respectively. Homozygotes for the rarer A-110 allele had a higher BMI (27.3 kg/m2 +/- 3.9) compared with homozygotes for the common C-110 allele (26.3 kg/m2 +/- 3.3). The rarer homozygotes were shorter and heavier than the common homozygotes. A follow-up study involved 1431 healthy, middle aged men from the UK (NPHS II group). The frequency of the A-110 allele was 0.385 (95% CI = 0.37-0.40), and there was no association of genotype with BMI. Thus there appears to be no strong association of the Hsp70-1 promoter polymorphism with risk of myocardial infarction, BMI or any coronary disease traits analysed here.

Rapid determination of apolipoprotein E genotype using a heteroduplex generator.

The apoE gene exhibits two common polymorphisms that have been associated with both coronary artery disease and Alzheimer's disease. The polymorphisms create the three allelic isoforms E2, E3, and E4 which are encoded by Cys;-Cys, Cys;-Arg, and Arg;-Arg at amino acid positions 112 and 158, respectively. Numerous methods have been described to identify these three apoE alleles although there are disadvantages and ambiguities associated with all of them. Here we describe a method by which the two common apoE polymorphisms can be identified simultaneously. The method involves PCR of the region containing the two polymorphic sites, followed by hybridization of this PCR product to a synthetic molecule called a universal heteroduplex generator (UHG). The UHG is used to induce heteroduplex formation which is visualized on a non-denaturing mini-gel using ethidium bromide staining. This technique which can also identify other rare mutations in the amplified region of DNA under investigation, is an unequivocal method of genotyping and is simpler and faster than many methods, including using restriction enzyme digestion.

High-throughput method for determination of apolipoprotein E genotypes with use of restriction digestion analysis by microplate array diagonal gel electrophoresis.

Molecular epidemiological research has identified the association of a common apolipoprotein E (apo E) isoform (E4 as opposed to E3), with risk both of coronary artery disease and of Alzheimer dementia. In addition, the role of apo E genotype (usually E2/E2) in Type III hyperlipidemia is well known. However, both for diagnostic and research purposes, apo E genotyping is cumbersome. The preferred approach is electrophoretic sizing of restriction digestion fragments, enabling simultaneous analysis of the two codons (112 and 158) that represent the six common genotypes (E2/E2; E2/E3; E2/E4; E3/E3; E3/E4; E4/E4). However, the consequent demands of high-yield PCR, high-resolution, high-throughput electrophoresis, and sufficient detection sensitivity have left shortfalls in published protocols. In conjunction with a high-throughput electrophoresis system we described recently, microplate array diagonal gel electrophoresis (MADGE), we have constructed extensively optimized, simplified protocols for DNA isolation from mouthwash samples for PCR setup and high-yield PCR, for restriction digestion, and for subsequent MADGE gel image analysis. The integral system enables one worker to readily undertake apo E genotyping of as many as hundreds of DNA samples per day, without special equipment.

Spectrum of LDL receptor gene mutations in heterozygous familial hypercholesterolemia.

Familial hypercholesterolemia by usual definition reflects mutations of the LDL-receptor gene. Extensive molecular characterization of mutations ascertained mainly through homozygotes (the Dallas collection) has been presented by Hobbs et al. (Hum Mutat 1:445-446, 1992). This paper catalogues a spectrum of 134 mutations (27 novel mutations in 45 patients, 24 previously described mutations in 89 patients) ascertained through heterozygotes from the analysis of 791 patients with definite, probable, or possible FH, mainly from the UK, using high-throughput modifications of the single-strand conformation polymorphism technique. From a composite database of LDL receptor gene mutations complied from these two sets and from the literature, deductions are made about ascertainment bias, mutation rates, and molecular heterogeneity. Calculations suggest that there may be a large number of rare amino acid variants in the general population not causing classic FH. Approaches to, and feasibility of, molecular diagnostics are considered.

W64R mutation in beta-3-adrenergic receptor gene and weight in a large population sample.

We have investigated the W64R (Trp64Arg) mutation in the beta-3-adrenergic receptor in 2270 healthy British males aged 50-61 y. The frequency of the rare R allele was 0.07 (95% confidence interval (CI) 0.06-0.08). The men showed an absence of association between W64R genotype and weight or height, both in the whole sample and in each quintile of the body mass index (BMI), and there was no association with tendency to gain weight. The W64R heterozygous state appears not to be a major contributing factor to obesity in the general population.