Influence of Genetic Polymorphisms in P2Y12 Receptor Signaling Pathway on Antiplatelet Response to Clopidogrel in Coronary Heart Disease

Backgrounds: Remarkable interindividual variability in clopidogrel response is observed, genetic polymorphisms in P2RY12 and its signal pathway is supposed to affect clopidogrel response in CHD patients. Methods: 539 CHD patients treated with clopidogrel were recruited. The platelet reaction index (PRI) indicated by VASP-P level were detected in 12-24h after clopidogrel loading dose (LD) or within 5-7 days after initiation of maintain dose (MD) clopidogrel. A total of 13 SNPs in relevant genes were genotyped in sample A (239 CHD patients). The SNPs which have significant differences in PRI will be validated in another sample (sample B, 300 CHD patients). Results: CYP2C19*2 increased the risk of clopidogrel resistance significantly. When CYP2C19*2 and CYP2C19*3 were considered, CYP2C19 loss of function (LOF) alleles were associated with more obviously increased the risk of clopidogrel resistance; P2RY12 rs6809699 C>A polymorphism was also associated with increased risk of clopidogrel resistance (AA vs CC: P=0.0398). This difference still existed after stratification by CYP2C19 genotypes. It was also validated in sample B. The association was also still significant even in the case of stratification by CYP2C19 genotypes in all patients (sample A+B). Conclusion: Our data suggest that P2RY12 rs6809699 is associated with clopidogrel resistance in CHD patients. Meanwhile, the rs6809699 AA genotype can increase on-treatment platelet activity independent of CYP2C19 LOF polymorphisms.

miR-122/SIRT1 axis regulates chondrocyte extracellular matrix degradation in osteoarthritis

Background/Aims: MicroRNAs (miRNAs) are involved in the pathogenesis of osteoarthritis (OA). The present study aimed to investigate the potential function of miR-122 in the development of OA and its potential molecular mechanisms. Methods: The expression of miR-122, silent information regulator 1 (SIRT1), collagen II, aggrecan, matrix metalloproteinase (MMP) 13 (MMP13) and ADAMTS4 in OA cartilage was detected by RT-qPCR. Target gene prediction and screening, luciferase reporter assay were used to verify downstream target genes of miR-122. Results: Compared with osteonecrosis, the expression of miR-122 was significantly increased in OA cartilage, while the expression of SIRT1 was significantly decreased. Overexpression of miR-122 increased the expression of extracellular matrix (ECM) catabolic factors, for example disintegrins, MMPs and metalloproteinases with platelet reaction protein motifs, and inhibited the expression of synthetic metabolic genes such as collagen II and aggregating proteoglycan. Inhibition of miR-122 expression had the opposite effect. Furthermore, SIRT1 was identified as a direct target of miR-122. SIRT1 was significantly inhibited by miR-122 overexpression. Knockdown of SIRT1 reversed the degradation of chondrocyte ECM by miR-122 inhibitors. Conclusion: The miR-122/SIRT1 axis can regulate the degradation of ECM in OA, thus providing new insights into the treatment of OA.

Precision of VerifyNow P2Y12 Assessment of Clopidogrel Response in Patients Undergoing Cerebral Aneurysm Flow Diversion

Background Dual antiplatelet therapy (DAT), most commonly with aspirin and Clopidogrel, is the standard of care for intracranial stenting, including flow diversion. Clopidogrel response varies by individual. Objective To investigate the real-world precision of VerifyNow P2Y12 assessment (Accumetrics, San Diego, California) of Clopidogrel response. Methods Using a prospectively-collected, IRB-approved cerebral aneurysm database 643 patients were identified who were treated with the Pipeline embolization device from 2011 to 2017. Patients with multiple P2Y12 assays drawn within a 24-h window were identified. A single patient could contribute multiple, independent sets. Levels drawn before a 5-d course of DAT and patients who received alternative antiplatelet agents were excluded. Therapeutic range was defined as platelet reaction units (PRU) 60–200. Results A total of 1586 P2Y12 measurements were recorded; 293 (46%) patients had more than one assay. One hundred forty (22%) patients had multiple P2Y12 measurements within 24 h. These patients accounted for 230 independent 24-h sets. The average P2Y12 fluctuation across all sets was 35 points; the 25th, 50th, and 75th percentiles were 12, 26, and 48 points, respectively. Of the 230 24-h sets of P2Y12 assays, 76% remained within their original therapeutic category: 100 (43%) all therapeutic, 54 (23%) all hypo-responsive, and 21 (9%) all hyper-responsive. Twenty-four percent of patients fluctuated between therapeutic categories when multiple P2Y12 assessments were drawn within a 24-h period: 29 (13%) between hypo-response and therapeutic, 23 (10%) between hyper-response and therapeutic, and 3 (1%) between hypo-response and hyper-response. CONCLUSION Our experience suggests P2Y12 is an often-imprecise measure, and this should be considered when utilizing P2Y12 levels for clinical decisions.

Features of platelet reaction in acute pancreatitis

Aim. To identify the features of platelet reaction in peripheral blood in patients with acute pancreatitis. Methods. We conducted a retrospective analysis of changes of platelets concentration in 19 patients with acute pancreatitis admitted to Kazan City clinical hospital №7. The age of patients ranged from 35 to 76 years, 45±12 years in average. All patients received complex treatment. The main method of surgical treatment was navigation drainage of fluid collections, peripancreatic mass and retroperitoneal space under ultrasound control performed in 11 patients. Subsequently in 4 patients due to progressive pyoinflammatory process midline laparotomy, sanitation, drainage of omental sac and purulent cavities were performed. In 4 cases aseptic pancreonecrosis resolved after conservative treatment. All patients dynamically had erythrocytes and platelets count and mean platelet volume checked. Results. Analysis of platelets concentration in peripheral blood in patients with acute pancreatitis showed multidirectional character depending on the severity of pathological process. In mild acute pancreatitis the platelet count remains within normal limits. In severe pancreatitis its number increases. The highest numbers of platelets were observed on days 10-20. On discharge platelet count remained elevated. Severe pervasive pancreonecrosis (n=4) with fatal outcome was characterized by thrombocytopenia on admission with further progression. Conclusion. The nature of the changes in platelet reaction in patients with acute pancreatitis depends on the severity of the pathological process and can be used as a criterion for evaluation of treatment effectiveness and prognosis of the outcome of the disease.

Abstract 16419: A Comparative Analysis of Prasugrel versus Ticagrelor on Rates of High On-Treatment Platelet Reactivity Following Switch From Clopidogrel Therapy

Background: Clopidogrel is characterized by non-uniform pharmacodynamic (PD) effects leading to considerable rates of high on-treatment platelet reactivity (HPR), a marker of increased atherothrombotic risk. Prasugrel and ticagrelor have greater potency and more uniform PD effects compared with clopidogrel. The effects of switching from clopidogrel to prasugrel or ticagrelor and how this impacts HPR rates has been poorly explored. Methods: This was a prospective, randomized PD study conducted in patients with coronary artery disease on maintenance aspirin (81mg/qd) and clopidogrel (75mg/qd) therapy. Patients were randomized to switch to prasugrel (60mg loading dose/10mg maintenance dose qd) or ticagrelor (180mg loading dose/90mg maintenance dose bid) for 1 week. PD assessments were performed at 5 time points: before (while on clopidogrel) and 30 min, 2 hrs, 24 hrs and 1 week after switch. HPR was defined by a platelet reactivity index (PRI) > 50% assessed by VASP and platelet reaction unit (PRU) > 208 assessed by VerifyNow P2Y12 (VN). Results: A total of 110 patients were studied. HPR on clopidogrel therapy as defined by VASP-PRI was observed in 67.3% of the overall population. HPR rates were similarly distributed among patients subsequently randomized to prasugrel vs ticagrelor (69.8% vs 64.7%, p=0.57). After switching to prasugrel or ticagrelor, HPR rates significantly reduced as early as 30 min and continued to decrease consistently in both groups over the study time period. At 1 week, HPR was found in 13.7% and 15.6% of patients randomized to prasugrel and ticagrelor, respectively (p=0.80) (Figure). Parallel findings were observed when assessing HPR rates defined by VN-PRU (Figure). Conclusions: Switching from clopidogrel to a more potent P2Y 12 inhibiting agent significantly reduces HPR rates. Reduction in HPR rates occur to a similar extent between prasugrel and ticagrelor in both the acute and maintenance phases of therapy.

Are P2Y12 reaction unit (PRU) and % inhibition index equivalent for the expression of P2Y12 inhibition by the VerifyNow® assay? Role of haematocrit and haemoglobin levels

SummaryThe results of the whole blood VerifyNow® P2Y12 assay can be expressed as platelet reaction units (PRU) or % inhibition index (%inh), but an optimal cut-off for the assessment of high on-treatment platelet reactivity (HPR) predictive of clinical events has been validated only for PRU. The aim of the study was to study the influence of haematological variables, such as platelet and leukocyte counts or haematocrit / haemoglobin, within the limits indicated by the manufacturer for assay validity, on the results of the test. We performed a comparison of PRU and %inh in a series 186 samples obtained from a clinical trial on patients under dual antiplatelet therapy. The results show that PRU significantly decreases with increasing haematocrit / haemoglobin, whereas %inh does not, due to a parallel change in PRU and iso-TRAP baseline value. PRU and % inhibition index are not equivalent for the definition of HPR, because of their different sensitivities to haematocrit / haemoglobin.

Abstract 4487: Clopidogrel Resistance in Patients Undergoing Percutaneous Coronary Intervention: One Year Outcome

Background: Clopidogrel therapy is critical in patients undergoing percutaneous coronary intervention (PCI). Data are lacking on long term outcomes of clopidogrel resistance (CR) and its definition. Methods: We identified 389 patients who underwent PCI during a 6 month period. Using the VerifyNow Point-of-care assay, P2Y12 platelet reaction units (PRU) and percent inhibition of platelet activation (IPA) were measured 16 –24 hours after the loading dose of clopidogrel (600mg for patients not on clopidogrel daily, and 300mg for patients on clopidogrel 75mg daily) but before the next day clopidogrel dose. CR was defined as PRU >200, %IPA<10% or IPA<20%. Major adverse cardiac events (MACE) included death, myocardial infarction, coronary artery bypass graft surgery and unplanned repeat coronary revascularization. Results: Mean age of patients was 65±11 years, 24% females and 31% diabetics. PRU mean value is 173±93, IPA mean value=47.5±42, CR is found in PRU>200 =36%(140), IPA<10% =8%(31), IPA <20% = 16.4%(64) patients. Presence of myocardial infarction, hypertension, hyperlipidemia, renal disease, smoking history, stable or unstable angina at admission were similar between the PRU groups and between IPA groups. Diabetes and older age were more prevalent in clopidogrel resistance patients. 1-year MACE rates are shown below. Conclusion: Clopidogrel resistance is an important predictor of long-term MACE in patients undergoing PCI, particularly when defined as Platelet Reaction Units >200.

Morphological and Functional Aspects of Cellular Hemostatic Mechanisms

SummaryTransmission, scanning and low-voltage, high resolution scanning electron microscopy have provided fundamental knowledge of relationships between plate-let structure and function. In combination with ultrastructural cytochemistry and immunocytochemistry, basic aspects of shape change, pseudopod extension, internal transformation, secretion, adhesion, aggregation, spreading, hemostatic plug formation and thrombus development have been defined. The morphological events of the platelet reaction in hemostasis have been linked to biochemical alterations. The result is a comprehensive picture of cellular aspects of hemosta-sis. This knowledge provides a basis for designing programs of treatment for heart attacks, strokes and other occlusive vascular disorders, as well as for their prevention.