Diabetes mellitus (DM) is associated with increased platelet activation and reduced platelet inhibition by clopidogrel. Prostaglandin E1 (PGE1) stimulates adenyl cyclase activity in platelets and increases cyclic AMP concentrations, which inhibit Ca2+release and platelet aggregation induced by P2Y1 receptor activation. PGE1 is included in the VerifyNow P2Y12 assay to suppress P2Y1 induced platelet aggregation. We hypothesized that diabetes mellitus may be associated with altered response to PGE1 in subjects treated with clopidogrel. Subjects with established coronary artery disease who were taking clopidogrel 75 mg daily and aspirin for >14 days were enrolled (n = 96). Diabetic (n = 34) were compared with non-diabetic subjects (n = 62). VerifyNow P2Y12 assay and light transmittance aggregometry (LTA) were performed using ADP as agonist with and without addition of PGE1. Genomic DNA was genotyped for common cytochrome P450 (CYP) 2C19 variants using Taqman assays. Residual on-treatment platelet aggregation induced by 20 μM ADP was not significantly different between subjects with and without DM. Addition of 22 nM and 88 nM PGE1 to 20 μM ADP resulted in a significant reduction of maximal platelet aggregation (MPA). Residual LTA platelet aggregation with PGE1 and VerifyNow P2Y12 platelet reactivity were significantly higher in subjects with DM than those without DM and in carriers of CYP 2C19⋆2 polymorphism. We conclude that an impaired inhibitory response to PGE1 may contribute to the high platelet reactivity phenotype in subjects with DM treated with clopidogrel. Addition of PGE1 to ADP agonist platelet assays may identify subjects with blunted inhibitory response to prostaglandins and result in a higher proportion of subjects with DM being classified as non-responders.

Recent candidate gene studies using a human liver bank and in vivo validation in healthy volunteers identified polymorphisms in cytochrome P450 (CYP) 3A4 gene (CYP3A4*22), Ah-receptor nuclear translocator (ARNT), and peroxisome proliferator-activated receptor-α (PPAR-α) genes that are associated with the CYP3A4 phenotype. We hypothesized that the variants identified in these genes may be associated with altered clopidogrel response, since generation of clopidogrel active metabolite is, partially mediated by CYP3A activity. Blood samples from 211 subjects, of mixed racial background, with established coronary artery disease, who had received clopidogrel, were analyzed. Platelet aggregation was determined using light transmittance aggregometry (LTA). Genotyping for CYP2C19*2, CYP3A4*22, PPAR-α (rs4253728, rs4823613), and ARNT (rs2134688) variant alleles was performed using Taqman® assays. CYP2C19*2 genotype was associated with increased on-treatment platelet aggregation (adenosine diphosphate 20 μM; P=0.025). No significant difference in on-treatment platelet aggregation, as measured by LTA during therapy with clopidogrel, was demonstrated among the different genotypes of CYP3A4*22, PPAR-α, and ARNT. These findings suggest that clopidogrel platelet inhibition is not influenced by the genetic variants that have previously been associated with reduced CYP3A4 activity.

BackgroundThe metabolic activation of clopidogrel is a two-step process. It has been suggested that paraoxonase-1 (PON1) is a rate-limiting enzyme in the conversion of 2-oxo- clopidogrel to an active thiol metabolite. Conflicting results have been reported in regard to (1) the association of a common polymorphism of PON1 (Q192R) with reduced rates of coronary stent thrombosis in patients taking clopidogrel and (2) its effects on platelet inhibition in patient populations of European descent.MethodsBlood samples from 151 subjects of mixed racial background with established coronary artery disease and who received clopidogrel were analyzed. Platelet aggregation was determined with light transmittance aggregometry and VerifyNow® P2Y12 assay. Genotyping for cytochrome P450 2C19 (CYP2C19)*2 and *3 and PON1 (Q192R) polymorphisms was performed.ResultsCarriers of CYP2C19*2 alleles exhibited lower levels of platelet inhibition and higher on-treatment platelet aggregation than noncarriers. There was no significant difference in platelet aggregation among PON1 Q192R genotypes. Homozygous carriers of the wild-type variant of PON1 (QQ192) had similar on-treatment platelet reactivity to carriers of increased-function variant alleles during maintenance clopidogrel dosing, as well as after administration of a clopidogrel 600 mg loading dose.ConclusionCYP2C19*2 allele is associated with impaired platelet inhibition by clopidogrel and high on-treatment platelet aggregation. PON1 (Q192R) polymorphism does not appear to be a significant determinant of clopidogrel response.

Factor XIII (FXIII) is necessary for cross linking of fibrin strands and generation of stable fibrin clot. FXIII Val34Leu is a common genetic single nucleotide polymorphism that has been associated with accelerated fibrin stabilization and reduced rate of fibrinolysis. The contribution of Val34Leu to long term risk of recurrent myocardial infarction (MI) in patients with coronary stenting has not been conclusively established. The objective of the study was to examine the effects of Val34Leu on fibrin generation, platelet aggregation, and long term clinical outcomes in patients with coronary artery disease treated with dual antiplatelet therapy. Patients with angiographically documented coronary artery disease who were treated with aspirin and clopidogrel were enrolled (n = 211). Light transmittance aggregometry and plasma fibrin clot formation using thrombelastography (TEG) were determined. Genotyping of Val34Leu was performed using Taqman assay. Clinical events during follow up were recorded. Homozygous carriers of 34Leu variant had significantly shorter fibrin clot formation time as compared to wild type individuals (TEG K: 1.27 ± 0.3 vs. 1.68 ± 1.1 min, p = 0.011). The Val34Leu variant was associated with gene dose dependent increased risk of MI (log rank, p = 0.002) or occurrence of composite of MI and CV death (log rank, p = 0.005) with highest event rates observed in homozygous carriers of 34Leu. In summary, FXIII Val34Leu polymorphism was associated with increased rate of fibrin stabilization in homozygous carriers of the variant and may increase risk of recurrent MI and death in patients with angiographically established coronary artery disease treated with dual antiplatelet therapy.

Mesothelioma is a rare pulmonary malignancy commonly associated with asbestos exposure. Its presentation is insidious and non-specific, with complaints of chest pain, dyspnea and cough. Chest X-ray may demonstrate unilateral pleural effusion. CT and PET scans may highlight nodular pleural plaques. Diagnosis often times is difficult with negative imaging and negative pleural fluid studies. In rare cases, hydropneumothoraces may be seen. We report a case of malignant pleural mesothelioma presenting as recurrent hydropneumothorax with negative CT scan of the chest for pleural abnormalities and negative pleural fluid studies.