Nutri-pharmacogenomics of warfarin anticoagulation therapy: VKORC1 genotype-dependent influence of dietary vitamin K intake.

Warfarin is the most widely prescribed oral anticoagulant, but large interindividual variations exist in the dose required to achieve comparable therapeutic effects. Several clinical and genetic variables have been identified that influence warfarin dosing. However, interactions between genotype and nutrition remain uncertain in terms of dietary vitamin K intake. To investigate genotype-nutrient interactions in warfarin anticoagulation therapy, 202 consecutive outpatients (M/F = 142/60, mean age, 69 years) undergoing treatment with warfarin were enrolled. Prevalent single nucleotide polymorphisms in VKORC1 and CYP2C9 were genotyped, and dietary vitamin K intake during the week preceding the blood sampling was quantitatively estimated by a dietitian-assisted questionnaire. Patients were classified according to low, medium, or high vitamin K intake. The mean daily warfarin dose in subjects with a VKORC1-1639 A/A genotype was significantly smaller than that with a -1639A/G genotype (2.74 vs. 3.91 mg/day, respectively, p < 0.0001). Dose requirements did not differ between subjects with a CYP2C9 *1/*3 genotype versus a CYP2C9 *1/*1 genotype. In subjects with a variant VKORC1-1639 G allele, the mean daily warfarin dose was significantly attenuated by low vitamin K intake compared with medium and high intake after adjustment for covariates (3.4 vs. 5.0 vs. 4.0 mg/day, respectively, p = 0.028). No such genotype effects were observed in homozygous patients for the VKORC1-1639 A allele. The results of the present study suggest that the capacity of dietary vitamin K intake to influence warfarin dose requirements during anticoagulation therapy is VKORC1 genotype-dependent, at least in part.

Response to pre-dilatation with POBA can predict target lesion revascularization after DCB angioplasty for de novo small coronary artery lesions.

OBJECTIVE: To assess the determinants of target lesion revascularization (TLR) after drug-coated balloon (DCB) angioplasty for de novo small coronary artery lesions. METHODS: This retrospective study enrolled consecutive lesions from patients that were in a stable condition and had undergone successful DCB treatment for de novo small coronary artery lesions. The study endpoint was TLR and major adverse cardiac events at 12 months. RESULTS: A total of 68 patients with 83 lesions were enrolled in the study. Of these, 11 (13.3%) lesions required TLR. Mean ± SD pre-dilatation balloon diameters were similar in the non-TLR (2.33 ± 0.72 mm) and TLR (2.18 ± 0.36 mm) groups. A comparison of the two groups showed that post/pre-lumen area ratio during pre-dilatation (%) by plain old balloon angioplasty (POBA) was significantly and negatively associated with TLR and the optimal cut-off point was 170%. Cox proportional hazard and multivariate regression analyses showed that post/pre-lumen area ratio was the only independent predictor of TLR (hazard ratio 0.9318; 95% confidence interval 0.9001, 0.9645). CONCLUSION: Greater pre-dilatation using POBA, assessed as the post/pre-lumen area ratio, may be independently associated with a lower 12-month TLR rate in patients undergoing DCB angioplasty for de novo small coronary lesions.

The circulating furin-cleaved/mature PCSK9 ratio has a potential prognostic significance in statin-naïve patients with acute ST elevation myocardial infarction.

Background and aims: Proprotein convertase subtilisin/kexin type 9 (PCSK9) circulates as mature and furin-cleaved forms, but their biological functions are uncertain. We investigated whether their levels associate with prognosis in patients with acute ST elevation myocardial infarction (STEMI). Methods: We enrolled 160 statin-naïve patients with acute STEMI and followed for 3 years. PCSK9 subtype levels were determined by an enzyme-linked immunosorbent assay before and at five timepoints up to 48 h after emergent coronary intervention. The occurrence of coronary and cardiac events was compared between subjects stratified by the PCSK9 level. Results: One hundred and twenty-six patients completed 3 years of follow-up. In the acute phase, both PCSK9 subtype levels decreased, and thereafter increased from 6 to 48 h (mature: from 198 ± 67 to 334 ± 116 ng/mL, furin-cleaved: from 20 ± 7 to 39 ± 16 ng/mL, both p < 0.01). Major cardiac events occurred in 46 patients. The furin-cleaved/mature PCSK9 ratio at 48 h after coronary intervention predicted the likelihood of experiencing of events; patients in the third tertile had lower event-free survival than those in the first and second tetiles in Kaplan-Meier analysis (p = 0.004). Multivariate Cox regression analysis revealed that this ratio had a greater impact (HR: 1.92; 95% CI: 1.06-3.45, p = 0.03) on events than other known atherosclerosis risk factors. Conclusions: The furin-cleaved/mature PCSK9 ratio was associated with 3-year cardiovascular events in statin-naïve patients with acute STEMI, suggesting a potential link between furin cleavage process of PCSK9 and its effect on prognosis. (249 words).

[Marked PT-INR prolongation associated with appetite loss due to urinary tract infection in a late elderly case with chronic atrial fibrillation].

An 80-year old woman presented with macroscopic hematuria on June 4(th), 2008. She had been suffering from general malaise and appetite loss since about 10 days previously. She had received anticoagulant therapy with warfarin due to chronic atrial fibrillation and PT-INR was well controlled between 1.6-2.2. When she presented, PT-INR was 12.88, and urinary tract infection (UTI) and hypoalbuminemia (2.2 g/dl) were observed. Therefore, warfarin therapy was discontinued, and antibiotics and vitamin K were administered. Normalization of PT-INR resulted in the disappearance of hematuria and UTI improved as a result of antibiotics administration. As the appetite loss improved, for serum albumin level increased. The previous dose of warfarin achieved PT-INR around 1.8. Her drug compliance had been good, and she took no drug nor food which could interact with warfarin. We also found no liver dysfunction, acute renal failure, malignancy, nor hyper- or hypo-thyroidism. Hypoalbuminemia caused by appetite loss due to UTI seems very likely to increase concentration of circulating free warfarin resulting in extreme prolongation of PT-INR. Our findings in the present case may suggest that we should pay more attention on changes of drug pharmacokinetics in elderly patients because of their poor adaptation to their circumstances such as infection or dehydration.

Microdosing clinical study to clarify pharmacokinetic and pharmacogenetic characteristics of atorvastatin in Japanese hypercholesterolemic patients.

The present study investigated whether the clinical pharmacokinetics of atorvastatin can be predicted from the results of microdosing study in Japanese patients with hypercholesterolemia whose SLCO1B1 and ABCG2 polymorphisms were analyzed. Forty seven statin-naive patients clinically indicated to LDL cholesterol-lowering therapy with atorvastatin were enrolled in a two-period crossover study. Microdose (100 µg) of atorvastatin was orally administered followed by therapeutic dose (10 mg) administration. Transport studies were performed with BCRP-expressing membrane vesicles. The dose-normalized plasma AUC following the therapeutic dose of atorvastatin was positively correlated with that following its microdose, but the AUC increased more than dose proportionally from microdose to therapeutic dose. The patients carrying SLCO1B1 c.521TC showed a significantly higher AUC compared with those carrying c.521TT following the microdose (175%) and therapeutic dose (139%). On the other hand, SLCO1B1 c.388G or ABCG2 c.421A variant alleles did not significantly affect the pharmacokinetics of atorvastatin. Atorvastatin showed ATP-dependent transport in BCRP-expressing membrane vesicles and it inhibited rosuvastatin transport with Ki of 6.3 ± 2.9 µM (mean ± SD). Microdosing study appears to be feasible to roughly estimate the pharmacokinetic and pharmacogenetic profiles of atorvastatin following the oral therapeutic dose in hypercholesterolemic patients.

Circulating malondialdehyde-modified low-density lipoprotein (MDA-LDL) as a novel predictor of clinical outcome after endovascular therapy in patients with peripheral artery disease (PAD).

BACKGROUND AND AIMS: Despite advances in the treatment of peripheral artery disease (PAD), cardiovascular events and death rates remain high. This study aimed at identifying markers of outcome in patients with PAD undergoing endovascular therapy (EVT). METHODS: Consecutive patients undergoing EVT were recruited. Markers of oxidative stress (malondialdehyde-modified low-density lipoprotein [MDA-LDL]), inflammation (IL-6; high-sensitivity C-reactive protein [hsCRP]) and fibrinolysis (D-dimer) were measured pre-EVT and at post-EVT time-points to 36 h. Clinical follow-up assessed major cardiovascular and/or limb events. RESULTS: In the 35 patients enrolled, mean MDA-LDL levels decreased from a baseline level of 106.2 U/L to 72.6 U/L immediately post-EVT (p<0.0001); levels remained significantly reduced at all time-points. IL-6, hsCRP and D-dimer increased and were significantly higher at the 36 h time-point. A significant, negative association was seen between decreased MDA-LDL and pre-EVT hsCRP levels (r = -0.42, p=0.012). Clinical follow-up data were obtained for a mean period of 16 months. MDA-LDL ratios (obtained by comparison of post- and pre-EVT values) allowed assessment of high (≥0.495) and low (<0.495) ratio groups. A significantly higher rate of major adverse events, including limb-related events or death, was seen in the low ratio group (p<0.001). Cox proportional hazard analysis including traditional risk factors indicated that this ratio is a significant predictor of clinical endpoints (HR = 0.4210, p=0.0154). An association with clinical outcome was not observed with the other candidate biomarkers. CONCLUSIONS: Assessment of pre- and post-EVT MDA-LDL levels is a promising marker of clinical outcome in patients with PAD.

A case of coronary rupture and pseudoaneurysm formation after fracture of implanted paclitaxel-eluting stents.

A 48-year-old man who had undergone implantation of two paclitaxel-eluting stents (PESs) at the right coronary artery was admitted to our hospital with progressive dyspnea. In the coronary care unit, he developed cardiogenic shock due to cardiac tamponade treated by pericardiocentesis. A coronary angiogram showed a large pseudoaneurysm at the site of the previously implanted stents, suggesting coronary rupture due to implanted stent fracture. The pseudoaneurysm was completely sealed by polytetrafluoroethylene-covered stent implantation. Although this case is very rare, coronary rupture by stent fracture should be considered when cardiac tamponade occurs after drug-eluting stent implantation, especially PES.

Chronic total occlusions of the right coronary and left anterior descending coronary arteries in a young adult patient with antiphospholipid syndrome.

A 36-year-old male appeared to have an old myocardial infarction on electrocardiogram, and coronary angiography (CAG) was performed. The CAG showed total occlusions of the right coronary artery and left anterior descending artery. He was successfully treated with drug-eluting stent implantation for both occluded coronary arteries. Such serious coronary lesions are uncommon for his young age. The patient was diagnosed as having antiphospholipid syndrome (APS) based on elevation of anticardiolipin antibody and anti-ß2 glycoprotein I antibody. Two years after stent implantation, the patient was well without ischemia or thrombosis. APS should be considered a potential cause of serious coronary disease in young adults. .

A possible role for HLA-DRB1*04:06 in statin-related myopathy in Japanese patients.

Statin-related myopathy (SRM) is a clinically important adverse reaction. Recent pharmacogenetic research, mainly in non-Asian populations, have indicated clinical relevance of some of genetic biomarkers to SRM, but predictive markers for SRM in Asian populations including Japanese has not yet been established. This study was aimed to identify clinically important genetic markers associated with SRM in Japanese patients. Allele frequencies of the three reported candidate markers - SLCO1B1 rs4149056, RYR2 rs2819742, and GATM rs9806699 - and carrier frequencies of HLA types were compared between patients with SRM patients (n = 52) and healthy Japanese subjects (n = 2878 or 86 (for rs9806699) as controls). No significant association of RYR2, SLCO1B1, and GATM variants with SRM were observed in our Japanese patients, but a significant association was detected for HLA-DRB1*04:06 with SRM (odds ratio: 3.19; 95% confidence interval: 1.53-6.66). This study suggested that HLA-DRB1*04:06 might be associated with SRM onset in a Japanese population. Further studies are required to validate these results.

Pharmacogenomics of statin responsiveness.

Statins are widely prescribed and are established as first-line therapy for the primary and secondary prevention of coronary artery disease. However, the benefit of treatment varies between patients. Genetic variation can contribute to interindividual variations in clinical efficacy of drug therapy, and significant progress has been made in identifying common genetic polymorphisms that influence responsiveness to statin therapy. To date, >30 candidate genes related to pharmacokinetics and pharmacodynamics of statins have been investigated as potential determinants of drug responsiveness in terms of low-density lipoprotein cholesterol lowering. Genetic variation at gene loci that affect intestinal cholesterol absorption include apolipoprotein (apo) E; adenosine triphosphate-binding cassette transporter G5 and G8; cholesterol production, such as 3-hydroxy-3-methylglutaryl coenzyme A reductase; and lipoprotein catabolism, such as apoB and the low-density lipoprotein receptor, all may play a role in modulating responsivesness as well genes involved in metabolism of statins such as cytochrome P450. However, there is considerable variation in results reported, and the data suggest that combined analysis of multiple genetic variants in several genes, all of which have possible functional significance, is more likely to give significant results than single gene studies in small sample populations. In the future, pharmacogenomic studies with a greater number of participants (>2,000 participants) should provide a better picture as to who is most likely and who is least likely to benefit from statin therapy.

ABCA1 gene variation and heart disease risk reduction in the elderly during pravastatin treatment.

AIMS: Our goals were to examine the relationships of a specific ATP-binding cassette transporter A1 (ABCA1) variant, rs2230806 (R219K), on baseline lipids, low-density lipoprotein cholesterol (LDL-C) lowering due to pravastatin, baseline heart disease, and cardiac endpoints on trial. METHODS AND RESULTS: The ABCA1 R219K variant was assessed in 5414 participants in PROSPER (PROspective Study of Pravastatin in the Elderly at Risk) (mean age 75.3 years), who had been randomized to pravastatin 40 mg/day or placebo and followed for a mean of 3.2 years. Of these subjects 47.6% carried the variant, with 40.0% carrying one allele, and 7.6% carrying both alleles. No effects on baseline LDL-C levels were noted, but mean HDL-C increased modestly according to the number of variant alleles being present (1.27 vs 1.28 vs 1.30 mmol/L, p = 0.024). No relationships between the presence or absence of this variant and statin induced LDL-C lowering response or CHD at baseline were noted. However within trial those with the variant as compared to those without the variant, the overall adjusted hazard ratio for new cardiovascular disease (fatal CHD, non-fatal myocardial infarction, or fatal or non-fatal stroke) was 1.22 (95% CI 1.06-1.40, p = 0.006), while for those in the pravastatin group it was 1.41 (1.15-1.73, p = 0.001), and for those in the placebo group it was 1.08 (0.89-1.30, p = 0.447) (p for interaction 0.058). CONCLUSION: Our data indicate that subjects with the ABCA1 R219K variant may get significantly less heart disease risk reduction from pravastatin treatment than those without the variant.

Increased levels of the oxidative stress marker, nitrotyrosine in patients with provocation test-induced coronary vasospasm.

BACKGROUND: Endothelial dysfunction of the coronary arteries caused by oxidative stress plays an important role in the pathogenesis of coronary vasospasm. However, it is not clear whether circulating biomarkers for oxidative stress are altered after coronary vasospasm. We investigated temporal changes in the levels of oxidative stress biomarkers after coronary vasospasm induced by intracoronary acetylcholine provocation testing, resulting in transient myocardial ischemia. METHODS AND RESULTS: Thirty consecutive patients with suspected vasospastic angina pectoris (VSAP) were enrolled in the study. Patients were categorized into the VSAP-positive group (n=14) and the VSAP-negative group (n=16) on the basis of test results. Serum samples were examined for the levels of the oxidative stress markers 4-hydroxynonenal (HNE) and nitrotyrosine (NT) before, and 15min, 3h, and 12h after the provocation test. The serum HNE levels did not change in either group after the test. The serum NT levels in the VSAP-positive group significantly increased at 3h and 12h after the test (11.3±3.3µg/ml at 3h, p=0.015, and 12.1±5.7µg/ml at 12h, p=0.03), as compared with baseline (8.1±3.2µg/ml). In the VSAP-negative group, the serum NT levels significantly decreased from baseline at each of the 3 time points. CONCLUSIONS: Serum NT significantly increased after coronary vasospasm induced by acetylcholine provocation, suggesting that serum NT could be a biomarker of transient myocardial ischemia and could contribute to the development of VSAP.

Genetic variation at the SLCO1B1 gene locus and low density lipoprotein cholesterol lowering response to pravastatin in the elderly.

Our goal was to determine whether genetic variation at genes affecting statin metabolism or targets of statin therapy would influence low density lipoprotein (LDL) cholesterol lowering with pravastatin, baseline heart disease, or cardiac endpoints on trial. We examined associations of single nucleotide polymorphisms (SNPs) at the liver X receptor alpha (LXRA, rs12221497), and the solute carrier organic anion transporter (SLCO1B1, rs4149056 and rs2306283) gene loci with these variables. We studied 5411 participants in PROSPER (PROspective Study of Pravastatin in the Elderly at Risk) (mean age 75.3 years), who had been randomized to pravastatin 40 mg/day or placebo and were followed for a mean of 3.2 years. No relationships between genetic variation at the LXRA gene locus with statin induced LDL lowering response or other parameters were noted. Both the SLCO1B1 rs4149056 (valine for alanine at 174) and the rs2306283 (asparagine for aspartic acid at 130) SNPs affect the amino acid sequence of the SLCO1B1 gene product. No effect of the rs2306283 SNP on any of the variables was noted. However the presence of the rs4149056 SNP was associated with significantly less LDL cholesterol lowering response to pravastatin (wildtype, 71.5% of the population, -37.0%; heterozygotes, 25.8% of the population, -36.0%; and homozygotes, 2.7% of the population, -31.8%, p=0.003 at 6 months, and p=0.022 at 12 months). Our data indicate that the presence of the rs4149056 non-synonymous SNP at the SLCO1B1 gene locus can significantly decrease the pravastatin induced LDL cholesterol lowering response.

KIF6, LPA, TAS2R50, and VAMP8 genetic variation, low density lipoprotein cholesterol lowering response to pravastatin, and heart disease risk reduction in the elderly.

Single nucleotide polymorphisms (SNPs) at the KIF6 (kinesin like protein 6, rs20455 or 719Arg), LPA (lipoprotein(a), rs3798220), TAS2R50 (taste receptor type 2, member 50, rs1376251) and VAMP8 (vesicle-associated membrane protein 8, rs1010) have previously been associated with low density lipoprotein cholesterol (LDL-C) lowering response to statins, coronary heart disease (CHD) at baseline, or CHD events on trial. We examined SNPs at the KIF6 (rs20455 or 719Arg), LPA (rs3798220), TAS2R50 (rs1376251) and VAMP8 (rs1010) in 5,411 participants in PROSPER (PROspective Study of Pravastatin in the Elderly at Risk) (mean age 75.3 years), who had been randomized to pravastatin 40 mg/day or placebo and were followed for a mean of 3.2 years. No SNP was related to vascular disease at baseline. Only the KIF6 SNP was related to LDL-C lowering with homozygous Arg 719 subjects being significantly less responsive than other groups (p=0.025, -34.2 vs. -36.1%). With regard to the primary CHD endpoint on trial (fatal or non-fatal myocardial infarction or stroke), we observed a significant relationship for KIF6 719Arg homozygotes (p=0.03, hazards ratio 0.47, 12.8% of the population) in women on pravastatin only, and for TAS2R50 for the AA genotype (p=0.03, hazards ratio 1.76, 8.9% of the population), also only in women on pravastatin. Our data indicate that the assessment of KIF6 rs20455 and TAS2R50 rs1376251 genotypes are not useful for predicting statin induced cardiovascular risk reduction in men, but do predict CHD risk reduction in women in this elderly population. However, these differences are no longer significant after correction for multiple comparisons, and we do not recommend the assessment of any of these SNPs in clinical practice.

Serum deoxyribonuclease I activity can be used as a novel marker of transient myocardial ischaemia: results in vasospastic angina pectoris induced by provocation test.

AIMS: Serum deoxyribonuclease I (DNase I) activity has recently been highlighted as a potential diagnostic marker for detection of acute myocardial infarction. To evaluate whether serum DNase I activity is useful for detection of myocardial ischaemia, we investigated alteration of its levels after onset of vasospastic angina pectoris (VSAP), resulting in transient myocardial ischaemia, induced by the intracoronary ergonovine provocation test. METHODS AND RESULTS: Twenty-nine consecutive patients with suspected VSAP were subjected to the test. Patients were categorized as VSAP-positive (n = 13) or -negative (n = 16) based on development of angina. Serum samples were examined for DNase I activity before, immediately after, and 3, 6, and 24 h after the provocation tests. The serum DNase I activity increased significantly from the baseline 3 h after the provocation test in 11 patients of the VSAP-positive group whose levels of troponin T were within the normal range. Median of the percentage differences from the baseline in serum DNase I activity 3 h after the test was 32.1% (25th and 75th percentile: 28.6 and 42.0%, respectively; P = 0.000012). In the VSAP-negative group, levels of DNase I activity remained unchanged at any point of time after the provocation test. CONCLUSION: Transient myocardial ischaemia resulting from VSAP induces a significant elevation of serum DNase I activity. Therefore, serum DNase I activity may be applicable as a useful marker for detecting transient myocardial ischaemia.