Toshiyuki MIYATA, PhD National Cardiovascular Center Research Institute, Osaka, Japan,the 19th Great Wall International Congress of Cardiology, October 24, 2008,Genetic risk factors for venous thromboembolism in Japanese,Advanced age is a risk for venous thromboembolism (VTE),All VTE,Blood, 110, 3097-3101, 2007,among residents of Olmsted County, Minnesota, from 1966 to 1990,PE,DVT alone,Male,Female,Incidence rates of VTE increase dramatically at about age 55, and, by age 80, are nearly 1 in 100 per year, approximately 1000-fold higher than for those aged 45 or younger.,Risk factors for venous thromboembolism N Engl J Med, 358, 1037-52, 2008,Acquired factors Reduced mobility Advanced age Cancer Acute medical illness Major surgery Trauma Spinal cord injury Pregnancy and postpartum period Polycythemia vera Antiphospholipid antibody syndrome Oral contraceptives,Hormone-replacement therapy Heparins Chemotherapy Obesity Central venous catheterization Immobilizer or cast Hereditary factors Antithrombin deficiency Protein C deficiency Protein S deficiency Factor V Leiden Prothrombin gene mutation,Thrombo- modulin,endothelium,EPCR,PC,PS,prothrombin,FVa,FXa,FVa FVIIIa,HSPG,thrombin,APC,Protein C/protein S system,Proteoglycan sulfate system,Heparan sulfate,FVi FVIIIi,IIa,AT,Anticoagulant Systems on Endothelium,Caucasian,Japanese,Factor V Leiden Prothrombin G20210A,Deficiencies of protein C, protein S, antithrombin,Low frequency variation,Deficiencies of protein C, protein S, antithrombin,Very rare mutation,Genetic risk factors for venous thromboembolism,Protein S K196E,Tokyo,General population, Suita study at NCVC,VTE patients: Sub-group of Blood Coagulation Abnormality Study Group,Suita, Osaka,NCVC, Osaka U,Nagoya U,Jichi Med U,Keio Univ,Kyoto Pref Med U,About 170 Japanese VTE patients,Case-control study for VTE,more than 3,500 individuals from a general population randomly selected from Suita city residents,Kimura et al., Blood, 2006,VTE,139 20 1 160 0.069 3290 332 17 3639 0.050 2.179 0.336 Pro Ser,Major homo Hetero Minor homo Total MAF Major homo Hetero Minor homo Total MAF 2 P Major allele Minor allele,General population(Suita study),152 9 0 161 0.028 3501 149 0 3650 0.020 0.987 0.320 Ala Thr,146 13 2 161 0.053 3585 66 0 3651 0.009 75.464 0.0001 Lys Glu,FXII -4CT,61 69 30 160 0.403 1468 1686 497 3651 0.367 3.402 0.183 4G 5G,ADAMTS13 P475S,PLG A620T,PS K196E,63 75 23 161 0.376 1513 1651 486 3650 0.359 0.372 0.830 T C,PAI-1 4G/5G,MAF: minor allele frequency,Case-control association study using Japanese VTE patients,Protein S K196E as a Risk for VTE,Population based-control n (%),Geno- types,VTE group n (%),Protein S K196E (Protein S Tokushima),KK KE EE Total KK KE+EE total,3585 (98.2) 66 (1.8) 0 (0.0) 3651 (100.0) 3585 (98.2) 66 (1.8) 3651 (100.0),2 = 75.464, P0.0001,2 = 41.807, P0.0001 OR=5.58 (95% CI 2.90-9.46),146 (90.7) 13 (8.1) 2 (1.2) 161 (100.0) 146 (90.7) 15 (9.3) 161 (100.0),OR: odds ratio, CI: confidence interval,Gla,EGF-like 1-4,Sex hormone-binding globulin,635 aa,Protein S K196E Protein S Tokushima,Lys196Glu,PS K196E mutation has been identified in Japanese DVT patients in 1993 by two independent groups. This mutation was referred to as protein S Tokushima. It is present in the second EGF-like domain of protein S. In vitro study showed that this mutant exhibits the loss of the APC cofactor activity and the prothrombinase inhibitory activity.,Yamazaki et al, 1993, Shigekiyo et al, 1993, Hayashi et al, 1994,Tokyo,Nagoya U 3 hetero / 182 individuals Heterozygosity: 1.65% Allele freq.: 0.82%,Kyushu U 5 hetero / 304 individuals Heterozygosity: 1.64% Allele freq.: 0.82%,Geographical distribution of PROS1 K196E mutation,Natl Cardio Vasc Ctr at Suita 66 heterozygotes / 3,651 individuals Heterozygosity: 1.81% Allele freq.: 0.90%,Heterozygotes,N=34,Wild-types,N=1,828,Staclot Protein S (Diagnostica Stago),Protein S activity in wild-type and K196E heterozygous individuals,71.9+/-17.6%,87.9+/-19.8%,Protein S activity,P0.0001,Kimura et al., JTH, 4, 2010-2013, 2006,Acquired factors influencing plasma protein S activity,Gender and age dependency of plasma protein S levels,Sakata et al, JTH 2004,Men N=1252,Women N=1438,30,40,50,60,70,80,140,120,100,80,60,Protein S activity (%),Age band,Age Gender Pregnancy Oral contraceptives Anticoagulant drugs DIC Liver diseases Kidney disease,PROS1 K196E mutation is a genetic risk factor for venous thromboembolism (VTE) in Japanese. A missense mutation causing Lys196 to be replaced by Glu is located within the 2nd EGF-like domain. The odds ratio of the mutant E allele for VTE was 5.58. Individuals heterozygous for the mutant E-allele had lower (mean 16%) plasma protein S activity than wildtype subjects.,PROS1 K196E,Kimura et al., Blood, 2006, Kimura et al., JTH 2006, Miyata et al., IJH 2006,The allelic frequency of the E allele was 0.9%. We estimated a total of as many as 10,000 Japanese as homozygotes. A substantial proportion of the Japanese population carries the PROS1 E allele and is at risk of developing VTE. Therefore, individuals with the PROS1 E allele should avoid environmental risk factors known to be associated with VTE.,PROS1 K196E, cont.,Kimura et al., Blood, 2006, Kimura et al., JTH 2006, Miyata et al., IJH 2006,Caucasian,Japanese,Factor V Leiden Prothrombin G20210A,Deficiencies of protein C, protein S, antithrombin,Low frequency variation,Deficiencies of protein C, protein S, antithrombin,Very rare mutation,Genetic risk factors for venous thromboembolism,Protein S K196E,Methods: We have sequenced the entire coding regions of 3 genes in all 173 DNA samples obtained from Japanese VTE patients without any consideration of their activities and antigen levels.,ABI 3730 DNA Analyzer,Question,Prevalence of nonsynonymous mutations in PROS1, PROC, and SERPINC1 (antithrombin) in the Japanese VTE patients,Number of Mutation Carriers,55 out of 173 VTE patients (32%) carried nonsynonymous mutations,SERPINC1 14,5,PROS1 24,PROC 12,Including one patient with PROS1 gene deletion,nonsynonymous mutations Missense, Nonsense, Frame-shift, Splice-site, Inframe deletion,PS K196E mutation as a modifier,R221W V339M V339M R271W K193del,K196E K196E K196E K196E K196E,No Yes No Yes unavailable,40 25 55 39 57,PROC,PROS1,Family history,Onset age of initial VTE,5UTR,promoter,centromere,D3S3619,D3S3634,Chromosome 3,Large PROS1 deletion in 1 out of 163 VTE patients,3,4,7,11,13,2,1,5,6,8,9,12,14,10,15,PROS1,Deletion, at least 107 kb,Yin et al., Thromb Haemost 2007; 98: 783-789,A large PROS1 deletion was found in one VTE patient who showed 16% PS activity and did not have point mutations in PROS1.,Comparison of first onset age of VTE events between mutation carriers and non-carriers,55* 44.7+/-16.5,118 52.6+/-16.1,Number,P=0.0031,Carriers,Non-carriers,First onset age mean+/-SD,* Five had mutations in both PROS1 and PROC. Two were homozygotes for PROS1 K196E. One was a compound heterozygote for PROS1 K196E/R101C.,Recurrent mutations found in Japanese VTE patients,PROS1 K196E in the 2nd EGF-like domain 2 homo, 13 hetero, reduced PS activity with normal PS antigen PROC K193del at the 6th residue from the C-terminus in the light chain 4 hetero, normal PC amidolytic activity with reduced anticoagulant activity PROC V339M in the catalytic 4 hetero, reduced PC amidolytic activity,SERPINC1 14,Other genetic factors,5,PROS1 24,PROC 12,VTE,Genetic risk factors,Environmental risk factors,Obesity, Cancer, Immobilization, Hospitalization, Surgery, Pregnancy,CLOT,prevention,Risk factors for venous thromboembolism N Engl J Med, 358, 1037-52, 2008,Acquired factors Reduced mobility Advanced age Cancer Acute medical illn