- Email: [email protected]
Mo-P6:450 Beta2-adrenergic receptor gene polymorphisms do not predict cardiovascular disease in the general
Mon&ty, June 19, 2006: Poster Session P6 Genetics of cardiovascular disease with higher VLDL-TG levels. Increased frequencies of both I (0.56, P<0.01) and A (0.44, P<0.01) alleles were found in group of patients with high VLDL-TG/-apo B ratio. Conclusions: The study shows the association of the common Ala54Thr and FABP-2p-ID SNPs with increased levels of VLDL-TG and high VLDLTG/VLDL-apo B ratio. It implicates their influence on VLDL-1, the main predictor of small dense LDL. Funding: Supported by grant IGA MH CR No. NtU8149-3.
Mo-P6:449
A NEW OX40L PROMOTER HAPLOTYPE SHOWING ASSOCIATION WITH CORONARY ARTERY D I S E A S E IN T H E P R O C A R D I S T R I O F A M I L Y A N D SCARF COHORTS
M. Ria I , The Procaxdis Consortmm - , EG. Olsson , O. Bengtsson 3, E Eriksson I , A. Hamsten I , J. Lagercrantz I . 1Atherosclerosis Research Unit,
King Gustaf V Research btstitute, Department of Medicbte, Karolbtska bzstitute, Stockholm, Sweden: 2Department of Cardiovascular Medicine, Universi~ of O~ford, O~ford, United Kingdom: ~Department of Molecular Biology, AstraZeneca R &D, Molndal, Sweden Objective: We have previously identified T-cell activator OX40 ligand (OX40L) as a polymorphic gene contributing, through as yet unknown mechanisms, to the risk of developing precocious coronary artery disease (CAD) and MI in two cohorts. Our approach was to use data from a mouse atherosderosis model to positionally identify candidate genes in a human context (Wang, et al. 2005). Here we present further fine mapping of the gene, in order to identify the physiological genetic variant. We also test the proposed association of OX40L with CAD in PROCARDIS families in order to unravel mechanisms behind the genetic component of cardiovascular diseases. M e t h o d s : Studies were performed in trios where proband suffered of precocious MI or symptomatic acute coronary syndrome (SACS). Genetic variants affecting potential regulatory regions were identified in silico. Linkage disequilibrium blocks structure was determined; haplotype analysis and Transmission Disequlibrium Test (TDT) were performed. Results: New, potentially functional, SNPs were identified. Furthermore these genetic variants were shown to be part of a haplotype associated with MI in the SCARF cohort. The new haplotype were also analysed in two-generation families, confirming a significant association with CAD. Conclusions: Our results, together with our previos case:control study, strongly reinforces the contention that this haplotype, defined by at least two SNPs, may be causally related to MI and/or CAD. Funding: Swedish Heart-Lung foundation, Swedish Medical Research Council, Swedish Society for Medical Research, Karolinska Institute and AstraZeneca Sweden.
JMo-P6:450
I BETA2-ADRENERGIC RECEPTOR GENE POLYMORPHISMS DO NOT PREDICT C A R D I O V A S C U L A R D I S E A S E IN T H E G E N E R A L
A.A. Sethi, B.G. Nordestgaard, A. Tybjaerg-Hansen. Universi~ of
Copenhagen, Copenhagen, Denmark Objectives: Single Nucleotide Polymorphisms (SNP) in the beta2-adrenergic receptor gene have been associated with dramatic reductions in adenylate cyclase activity and decreased cardiac function in animal models. In humans, these SNPs have been associated with altered vascular response to sympa-thetic stimulation and with outcome of congestive heart failure in patients with functional heart failure class II-IV. We tested the hypothesis that Glyl6Arg, Gln27Glu, and Thr164Ile in the beta2-adrenergic receptor gene were associated with cardiovascular disease in 9,224 individuals from the general population. M e t h o d s : Abovementioned SNPs were genotyped in 1,110 cases with ischemic heart disease, 490 cases with ischemic cere-brovascular disease, and 7,976 controls from the general population, The Copenhagen City Heart Study. Results: Allele frequencies of 16Arg, 27Glu, and 164Ile were 0.38, 0.44, and 0.01, respectively. Genotype was not associ-ated with risk of ischemic heart disease, irrespective of whether we studied SNPs alone, or combined, women or men, and whether age alone or in addi-tion other cardiovascular risk factors were allowed for. Similarly, genotype did not predict risk of ischemic cerebrovascular disease, irrespective of whether we examined SNPs alone, or combined, women or men, and whether allowing for age or in addition for other risk factors. Conclusions: In two cohort studies none of the SNPs in the beta2-
145
adrenergic receptor gene, considered separately or combined, were associated with risk of ischemic heart or cerebrovascular disease in women or men. Funding: Nothing to dis-dose of commercial nature.
Mo-P6:4511S I N G L E - S T R A N D
CONFORMATION POLYMORPHISM FOR THE MUTATION SCANNING OF PROPROTEIN CONVERTASE SUBTILISIN KEXIN 9 IN F A M I L I A L H Y P E R C H O L E S T E R O L A E M I A
S. Mapplebeck 1 , R. Whittall 2 , C.S. Hubbart 2, M. Thomas 1 , S.E. Humphries 2. 1 Clinical Biochemistry, Royal Free Hospital, London,
United Kingdom." 2Cardiovascular Genetics, Universi~ College, LotMon, United Kingdom Objective: Familial hypercholesterolaemia (FH) is a common disorder of lipoprotein metabolism. In the majority of patients, FH arises due to mutations in the gene encoding the LDL receptor (LDLR) with 2-5% of cases caused by the R3500Q change, within the apoB gene (APOB). Several nucleotide changes within proprotein convertase subtilisin kexin 9 (PCSK9) are reported to segregate with hypercholesterolaemia in patients lacking changes in either the L D L R or APOB. The objective of this study was to develop a fluorescencebased single strand conformation polymorphism (SSCP) method for the rapid mutation scanning of PCSK9. M e t h o d s : 300 UK FH patients, lacking mutations in either the L D L R or APOB, were examined for nucleotide changes in PCSK9. Exons were amplified by fluorescent-PCR and screened by capillary SSCP and heteroduplex analysis (HA). Those samples displaying differences in electrophoretic mobility were sequenced. Results: SSCP identified the majority of the reported nucleotide changes within PCSK9. In addition, within exon 2, a different SSCP pattern arose due to the base change G > A at position 650 (D129N) with a prevelence of 0.9% in these FH patients. Digestion with NRUI in 538 healthy UK men confirmed this nudeotide change as confined to FH subjects. Conclusions: We have successfuly performed SSCP/HA using fluorescently-labdled primers for the rapid detection of nucleotide changes in PCSK9. However, further optimisation is required to improve both the sensitivity and specificity of this method. Funding: Funded by the British Heart Foundation.
Mo-P6:452
G E N O T Y P E S AT T H E M A T R I X METALLOPROTEINASE (MMP) LOCI FOR MMP9 AND MMP12 ARE ASSOCIATED WITH CAROTID IMT MEASURES OF PLAQUE STABILITY
A. Panayiotou 1, M. Griffin 2 , N. Georgiou 2 , D. Bond 2, T. Tyllis 2, S.E. Humphries 3, A.N. Nicolaides 1,2.1 University of Cyprus, Nicosia,
Cyprus: 2 Vascular Screening atzd Diagnostic Center; Nicosia, Cyprus: 3Cardiovascular Genetics, Ucl, LotMon, United Kingdom Objective: Unstable atherosderotic plaques axe hypoechoic on ultrasound whereas stable plaques are hyperechoic. The aim of the study was to determine the association between MMP polymorphisms and plaque echodensity. M e t h o d s : Carotid and femoral bifurcations have been scanned with highresolution ultrasound in 516 volunteers over the age of 40. Total plaque thickness (TPT) (the sum of plaque thickness in every artery in cm) and the mean plaque type (MPT) using the Widder classification, with type 1 being the most hyperechoic and type 5 the most hypoechoic, were recorded. G e n t typing was performed with the TaqMan method and the MMP9 (R279Q), MMP12 (-82A>G), MMP7 (-181A>G), MMP1 (1G/2G) polymorphisms were determined. Results: There was no association between MMP7 or MMP1 genotypes with TPT or MPT. However, subjects heterozygous for the MMP9 and MMP12 genotypes had a higher plaque thickness (median, IQR) (0.38, 0.14-0.68) and (0.41, 0.15-0.74) respectively than those homozygous for the common allele (0.28, 0-0.595) and (0.29, 0-0.62) (P<0.01). In a multiple regression analysis both MMP9 and MMP12 heterozygous genotypes were independent predictors of TPT (OR 1.68, CI 1.16-2.45 and OR 1.54, CI 1.0-2.3) and could explain 3.2% of the variability in TPT. In a linear logistic analysis, MMP12 genotype was the only significant predictor of MPT (OR 1.54, CI 1.03-2.3). These findings remained significant after correcting for age. Conclusions: The MMP9 (R279Q) and the MMP12 (-82A>G) polymorphisms are associated with increased plaque thickness and reduced echodensity, features that are characteristic of the unstable plaque.
XIV bztenmtional Symposium on Atherosclerosis, Rome, Italy, June 18-22, 2006
Mo-P6:449
A NEW OX40L PROMOTER HAPLOTYPE SHOWING ASSOCIATION WITH CORONARY ARTERY D I S E A S E IN T H E P R O C A R D I S T R I O F A M I L Y A N D SCARF COHORTS
M. Ria I , The Procaxdis Consortmm - , EG. Olsson , O. Bengtsson 3, E Eriksson I , A. Hamsten I , J. Lagercrantz I . 1Atherosclerosis Research Unit,
King Gustaf V Research btstitute, Department of Medicbte, Karolbtska bzstitute, Stockholm, Sweden: 2Department of Cardiovascular Medicine, Universi~ of O~ford, O~ford, United Kingdom: ~Department of Molecular Biology, AstraZeneca R &D, Molndal, Sweden Objective: We have previously identified T-cell activator OX40 ligand (OX40L) as a polymorphic gene contributing, through as yet unknown mechanisms, to the risk of developing precocious coronary artery disease (CAD) and MI in two cohorts. Our approach was to use data from a mouse atherosderosis model to positionally identify candidate genes in a human context (Wang, et al. 2005). Here we present further fine mapping of the gene, in order to identify the physiological genetic variant. We also test the proposed association of OX40L with CAD in PROCARDIS families in order to unravel mechanisms behind the genetic component of cardiovascular diseases. M e t h o d s : Studies were performed in trios where proband suffered of precocious MI or symptomatic acute coronary syndrome (SACS). Genetic variants affecting potential regulatory regions were identified in silico. Linkage disequilibrium blocks structure was determined; haplotype analysis and Transmission Disequlibrium Test (TDT) were performed. Results: New, potentially functional, SNPs were identified. Furthermore these genetic variants were shown to be part of a haplotype associated with MI in the SCARF cohort. The new haplotype were also analysed in two-generation families, confirming a significant association with CAD. Conclusions: Our results, together with our previos case:control study, strongly reinforces the contention that this haplotype, defined by at least two SNPs, may be causally related to MI and/or CAD. Funding: Swedish Heart-Lung foundation, Swedish Medical Research Council, Swedish Society for Medical Research, Karolinska Institute and AstraZeneca Sweden.
JMo-P6:450
I BETA2-ADRENERGIC RECEPTOR GENE POLYMORPHISMS DO NOT PREDICT C A R D I O V A S C U L A R D I S E A S E IN T H E G E N E R A L
A.A. Sethi, B.G. Nordestgaard, A. Tybjaerg-Hansen. Universi~ of
Copenhagen, Copenhagen, Denmark Objectives: Single Nucleotide Polymorphisms (SNP) in the beta2-adrenergic receptor gene have been associated with dramatic reductions in adenylate cyclase activity and decreased cardiac function in animal models. In humans, these SNPs have been associated with altered vascular response to sympa-thetic stimulation and with outcome of congestive heart failure in patients with functional heart failure class II-IV. We tested the hypothesis that Glyl6Arg, Gln27Glu, and Thr164Ile in the beta2-adrenergic receptor gene were associated with cardiovascular disease in 9,224 individuals from the general population. M e t h o d s : Abovementioned SNPs were genotyped in 1,110 cases with ischemic heart disease, 490 cases with ischemic cere-brovascular disease, and 7,976 controls from the general population, The Copenhagen City Heart Study. Results: Allele frequencies of 16Arg, 27Glu, and 164Ile were 0.38, 0.44, and 0.01, respectively. Genotype was not associ-ated with risk of ischemic heart disease, irrespective of whether we studied SNPs alone, or combined, women or men, and whether age alone or in addi-tion other cardiovascular risk factors were allowed for. Similarly, genotype did not predict risk of ischemic cerebrovascular disease, irrespective of whether we examined SNPs alone, or combined, women or men, and whether allowing for age or in addition for other risk factors. Conclusions: In two cohort studies none of the SNPs in the beta2-
145
adrenergic receptor gene, considered separately or combined, were associated with risk of ischemic heart or cerebrovascular disease in women or men. Funding: Nothing to dis-dose of commercial nature.
Mo-P6:4511S I N G L E - S T R A N D
CONFORMATION POLYMORPHISM FOR THE MUTATION SCANNING OF PROPROTEIN CONVERTASE SUBTILISIN KEXIN 9 IN F A M I L I A L H Y P E R C H O L E S T E R O L A E M I A
S. Mapplebeck 1 , R. Whittall 2 , C.S. Hubbart 2, M. Thomas 1 , S.E. Humphries 2. 1 Clinical Biochemistry, Royal Free Hospital, London,
United Kingdom." 2Cardiovascular Genetics, Universi~ College, LotMon, United Kingdom Objective: Familial hypercholesterolaemia (FH) is a common disorder of lipoprotein metabolism. In the majority of patients, FH arises due to mutations in the gene encoding the LDL receptor (LDLR) with 2-5% of cases caused by the R3500Q change, within the apoB gene (APOB). Several nucleotide changes within proprotein convertase subtilisin kexin 9 (PCSK9) are reported to segregate with hypercholesterolaemia in patients lacking changes in either the L D L R or APOB. The objective of this study was to develop a fluorescencebased single strand conformation polymorphism (SSCP) method for the rapid mutation scanning of PCSK9. M e t h o d s : 300 UK FH patients, lacking mutations in either the L D L R or APOB, were examined for nucleotide changes in PCSK9. Exons were amplified by fluorescent-PCR and screened by capillary SSCP and heteroduplex analysis (HA). Those samples displaying differences in electrophoretic mobility were sequenced. Results: SSCP identified the majority of the reported nucleotide changes within PCSK9. In addition, within exon 2, a different SSCP pattern arose due to the base change G > A at position 650 (D129N) with a prevelence of 0.9% in these FH patients. Digestion with NRUI in 538 healthy UK men confirmed this nudeotide change as confined to FH subjects. Conclusions: We have successfuly performed SSCP/HA using fluorescently-labdled primers for the rapid detection of nucleotide changes in PCSK9. However, further optimisation is required to improve both the sensitivity and specificity of this method. Funding: Funded by the British Heart Foundation.
Mo-P6:452
G E N O T Y P E S AT T H E M A T R I X METALLOPROTEINASE (MMP) LOCI FOR MMP9 AND MMP12 ARE ASSOCIATED WITH CAROTID IMT MEASURES OF PLAQUE STABILITY
A. Panayiotou 1, M. Griffin 2 , N. Georgiou 2 , D. Bond 2, T. Tyllis 2, S.E. Humphries 3, A.N. Nicolaides 1,2.1 University of Cyprus, Nicosia,
Cyprus: 2 Vascular Screening atzd Diagnostic Center; Nicosia, Cyprus: 3Cardiovascular Genetics, Ucl, LotMon, United Kingdom Objective: Unstable atherosderotic plaques axe hypoechoic on ultrasound whereas stable plaques are hyperechoic. The aim of the study was to determine the association between MMP polymorphisms and plaque echodensity. M e t h o d s : Carotid and femoral bifurcations have been scanned with highresolution ultrasound in 516 volunteers over the age of 40. Total plaque thickness (TPT) (the sum of plaque thickness in every artery in cm) and the mean plaque type (MPT) using the Widder classification, with type 1 being the most hyperechoic and type 5 the most hypoechoic, were recorded. G e n t typing was performed with the TaqMan method and the MMP9 (R279Q), MMP12 (-82A>G), MMP7 (-181A>G), MMP1 (1G/2G) polymorphisms were determined. Results: There was no association between MMP7 or MMP1 genotypes with TPT or MPT. However, subjects heterozygous for the MMP9 and MMP12 genotypes had a higher plaque thickness (median, IQR) (0.38, 0.14-0.68) and (0.41, 0.15-0.74) respectively than those homozygous for the common allele (0.28, 0-0.595) and (0.29, 0-0.62) (P<0.01). In a multiple regression analysis both MMP9 and MMP12 heterozygous genotypes were independent predictors of TPT (OR 1.68, CI 1.16-2.45 and OR 1.54, CI 1.0-2.3) and could explain 3.2% of the variability in TPT. In a linear logistic analysis, MMP12 genotype was the only significant predictor of MPT (OR 1.54, CI 1.03-2.3). These findings remained significant after correcting for age. Conclusions: The MMP9 (R279Q) and the MMP12 (-82A>G) polymorphisms are associated with increased plaque thickness and reduced echodensity, features that are characteristic of the unstable plaque.
XIV bztenmtional Symposium on Atherosclerosis, Rome, Italy, June 18-22, 2006