Biomedical subjects

John T Brandt

Publications and source records attributed to John T Brandt.

15 recordsLinked to original sources

PubMed2007

A comparison of prasugrel and clopidogrel loading doses on platelet function: magnitude of platelet inhibition is related to active metabolite formation.

BACKGROUND: The aim of this study was to compare rate of onset, magnitude, and consistency of platelet inhibition after administration of prasugrel or clopidogrel and to relate platelet inhibition to systemic exposure to each active metabolite. Thienopyridines are prodrugs, metabolized in vivo to active metabolites that inhibit the platelet P2Y12 adenosine diphosphate (ADP) receptor. METHODS: This was an open-label, 2-way, crossover study that randomized healthy subjects (n = 68) to an oral loading dose (LD) of prasugrel 60 mg or clopidogrel 300 mg. Platelet aggregation response to 5 and 20 micromol/L of ADP was measured by turbidometric aggregometry. Plasma concentrations of the active metabolites of prasugrel and clopidogrel were quantified by liquid chromatography with tandem mass spectrometry detection methods. RESULTS: Inhibition of platelet aggregation (IPA) after prasugrel was significantly higher (P < .01) than that after clopidogrel from 15 minutes through 24 hours (5 micromol/L ADP) and from 30 minutes through 24 hours (20 micromol/L ADP). For 20 micromol/L ADP, the median time to reach > or = 20% IPA was 30 minutes for prasugrel and 1.5 hours for clopidogrel (P < .001). The maximum IPA was 84.1% +/- 9.5% with prasugrel versus 48.9% +/- 27.0% with clopidogrel for 5 micromol/L ADP and 78.8% +/- 9.2% versus 35.0% +/- 24.5%, respectively, for 20 micromol/L ADP (P < .001). Response to prasugrel was more consistent compared to clopidogrel (P < .01). The lower IPA response to clopidogrel was associated with lower plasma concentrations of its active metabolite (P < .001). CONCLUSIONS: Prasugrel 60 mg LD results in more rapid, potent, and consistent inhibition of platelet function than clopidogrel 300 mg LD. Lower IPA responses to clopidogrel were associated with lower concentrations of its active metabolite.

Adult↗

PubMed2006

A multiple dose study of prasugrel (CS-747), a novel thienopyridine P2Y12 inhibitor, compared with clopidogrel in healthy humans.

AIMS: This double-blind, placebo-controlled trial was designed to evaluate the pharmacodynamics, pharmacokinetics, safety, and tolerability of prasugrel (CS-747, LY640315), a novel thienopyridine P2Y(12) ADP receptor antagonist compared with clopidogrel, during multiple oral dosing in healthy subjects. METHODS: Thirty subjects received placebo, prasugrel 5 mg, 10 mg, or 20 mg, or clopidogrel 75 mg orally, daily for 10 days. Platelet aggregation, bleeding time, and prasugrel metabolites were measured and adverse events were recorded. RESULTS: Inhibition of ADP-induced platelet aggregation reached steady state by day 3 following prasugrel 10 and 20 mg compared with 5 days for clopidogrel 75 mg or prasugrel 5 mg. Compared with placebo, at 24 h after the last dose of study drug, inhibition of platelet aggregation using (20 microm) ADP was significantly higher in the prasugrel 10 mg group (58.2 +/- 4.9% vs. 9.2 +/- 4.0%, P < 0.001) with no difference in the clopidogrel group (15.7 +/- 6.8% vs. 9.2 +/- 4.0%, P = 0.78). With 5 microm ADP, inhibition of platelet aggregation with prasugrel 10 mg and clopidogrel 75 mg was significantly higher than with placebo (prasugrel 10 mg, 70.5 +/- 4.7%; clopidogrel 75 mg, 36.5 +/- 9.0%; vs. placebo, 11.3 +/- 5.1%; P < 0.0001 and P = 0.02). On day 10 at 4 h postdose, bleeding time was prolonged with prasugrel 10 mg (prasugrel 10 mg, 706 +/- 252 s vs. placebo, 221 +/- 38 s, P = 0.05) but not with clopidogrel (283 +/- 56 s, P = 0.98). There were no clinically significant bleeding events, serious adverse events, or discontinuations of the study drug. CONCLUSIONS: Compared with clopidogrel 75 mg, prasugrel 10 mg and 20 mg daily for 10 days resulted in more rapid, more consistent, and higher levels of platelet inhibition.

Adolescent↗

PubMed2006

Prasugrel achieves greater inhibition of platelet aggregation and a lower rate of non-responders compared with clopidogrel in aspirin-treated patients with stable coronary artery disease.

AIMS: This study was designed to compare the degree of inhibition of platelet aggregation (IPA) of prasugrel with that of clopidogrel in stable aspirin-treated patients with coronary artery disease (CAD). METHODS AND RESULTS: Subjects (n=101) were randomly assigned to the following loading dose (LD) (day 1)/maintenance dose (MD) (days 2-28) combinations: prasugrel, 40 mg/5 mg; 40 mg/7.5 mg; 60 mg/10 mg; 60 mg/15 mg; or clopidogrel, 300 mg/75 mg. Turbidometric platelet aggregation was measured at multiple timepoints during the study. At 4 h after dosing, with 20 microM ADP, both prasugrel LDs achieved significantly higher mean IPA levels (60.6% and 68.4 vs. 30.0%, respectively; all P<0.0001) and lower percentage (3 vs. 52%, P<0.0001) of pharmacodynamic non-responders (defined as IPA <20%) than clopidogrel. Prasugrel 10 and 15 mg MDs achieved consistently higher mean IPA than clopidogrel 75 mg at day 28 (all P<0.0001). At pre-MD on day 28, there were no non-responders in the 10 and 15 mg prasugrel group, compared with 45% in the clopidogrel group (P=0.0007). CONCLUSION: In this population, prasugrel (40-60 mg LD and 10-15 mg MD) achieves greater IPA and a lower proportion of pharmacodynamic non-responders compared with the approved clopidogrel dosing.

Adult↗

PubMed2006

External peer review quality assurance testing in von Willebrand disease: the recent experience of the United States College of American Pathologists proficiency testing program.

The U.S. College of American Pathologists (CAP) has conducted a focused study of the proficiency testing for von Willebrand disease (vWD) analysis from 2003 to 2005. This report summarizes the findings regarding the accuracy and precision of the various assays at different analyte levels, as well as the influence of the reference material used to construct the assay standard curve. The results show that testing of von Willebrand factor (vWF):antigen (vWF:Ag) and ristocetin cofactor activity (vWF:RCo) is reasonably accurate, with all-method mean values falling within 3.2 and 5.6%, respectively, of the International Society on Thrombosis and Haemostasis Secondary Coagulation Standard (lot 2) assigned values. vWF:Ag measurements are reasonably precise (all-method coefficients of variation [CVs] = 10.7 to 15.1%), even at lower levels of vWF. The highest precision was observed for immunoturbidometric assays (CVs, 6.3 to 9.7%). vWF:RCo measurements are less precise (all-method CVs, 23.3 to 30.9%). The reference materials used in the standard curves for immunoturbidometric vWF:Ag assays appear to have accurately assigned vWF values for the majority of commercial suppliers.

Antigens↗

PubMed2006

Platelet inhibitory activity and pharmacokinetics of prasugrel (CS-747) a novel thienopyridine P2Y12 inhibitor: a multiple-dose study in healthy humans.

This double-blind, placebo-controlled trial evaluated the safety, pharmacodynamics, and pharmacokinetics of prasugrel (CS-747, LY640315), a novel thienopyridine P2Y12 ADP receptor antagonist, during multiple oral dosing in healthy subjects. Eighteen subjects received placebo, or prasugrel 2.5 or 10 mg, orally, daily for 10 days. Adverse events were recorded and blood samples for measurement of platelet aggregation, bleeding time, and prasugrel metabolite concentrations were obtained. Two bleeding events were experienced in the prasugrel 10 mg dose group and one in the placebo group. Neither of the events was considered serious. ADP-induced platelet aggregation accumulated over the 10-day study period, reaching a steady state by days 2-4 following administration of prasugrel 10 mg daily. Limited inhibition of platelet aggregation was obtained with prasugrel 2.5 mg. In the 10-mg dose group at day 5, 4 h postdose, with 20 microM ADP as the agonist, inhibition of platelet aggregation was 61.2 +/- 5.6 vs. 17.9 +/- 6.2% in the placebo group (P < 0.01). Recovery of platelet aggregation was observed 48 h after the last dose of prasugrel 10 mg, but was only partial, consistent with irreversible platelet inhibition. The mean bleeding time was prolonged by day 5 at 4 h postdose in all subjects in the prasugrel 10 mg dose group (prasugrel 10 mg, 1058 +/- 412 s vs. placebo, 196 +/- 74 s; P < 0.001). The maximum plasma concentration of metabolites increased with prasugrel dose and there was no accumulation of metabolites over the 10-day study period. This study indicated that prasugrel 10 mg daily for 10 days was well tolerated and at steady-state provided sustained high levels of inhibition of platelet aggregation.

Administration, Oral↗

PubMed2006

Platelet inhibitory activity and pharmacokinetics of prasugrel (CS-747) a novel thienopyridine P2Y12 inhibitor: a single ascending dose study in healthy humans.

We assessed the tolerability, pharmacodynamics as measured by inhibition of platelet aggregation (IPA), and pharmacokinetics of prasugrel (CS-747, LY640315), a novel thienopyridine antiplatelet agent in healthy volunteers. Twenty-four subjects were randomized into four groups of six in a double-blind, placebo-controlled trial. One subject in each group received placebo and five subjects received prasugrel orally at single doses of 2.5, 10, 30, or 75 mg. The IPA, assessed using 5 and 20 microM ADP, was periodically measured over a 7-day period by light transmission aggregometry. Plasma concentrations for three major metabolites, R-95913, R-106583, and R-100932, were measured. There were no serious adverse events and no clinically significant changes noted in any laboratory or clinical evaluations in any subject. At 1 h after prasugrel 30 and 75 mg, platelet aggregation induced by 20 microM ADP was inhibited by 43.5 +/- 7.8 and 43.2 +/- 15.7%, respectively, and this inhibition was significantly greater than that following placebo (5.9 +/- 3.5%) (P < 0.05 for both doses). The degree of inhibition observed at 2 h was slightly higher with both prasugrel 30 and 75 mg (59.8 +/- 9.9 and 57.0 +/- 7.2%) and was maintained through the subsequent 22 h. At 24 h, maximal platelet aggregation induced by 20 microM ADP was reduced to 0.05 for 2.5 and 10 mg prasugrel vs. placebo). With prasugrel 75 mg at 4 h postdose, there was a significant increase in the mean bleeding time compared to placebo (682 vs. 161 s; P < 0.05). Prasugrel metabolites obeyed linear pharmacokinetics and the three metabolites appeared in the plasma soon after administration, reaching maximum levels at approximately 1 h. In conclusion, prasugrel 30 and 75 mg were well tolerated and achieved a consistently high level of platelet inhibition with a fast onset of action.

Adenosine Diphosphate↗

PubMed2006

The platelet inhibitory effects and pharmacokinetics of prasugrel after administration of loading and maintenance doses in healthy subjects.

Prasugrel (CS-747, LY640315), a novel thienopyridine, is a potent and orally active antiplatelet agent in vivo. The aims of this double-blind, double-dummy, placebo-controlled, randomized, parallel group phase 1 study were to investigate the antiplatelet effects of prasugrel after oral administration of a loading dose (LD) and subsequent 20 days of once-daily maintenance dosing (MD), to characterize the pharmacokinetics of prasugrel metabolites with an LD/MD regimen, and to assess the safety and tolerability of prasugrel in healthy subjects. Subjects were randomly assigned in a 1:1:1 ratio to prasugrel 40 mg LD/7.5 mg MD (n = 11), prasugrel 60 mg LD/15 mg MD (n = 10), or placebo LD/placebo MD (n = 11). Prasugrel 40 and 60 mg LDs provided rapid and consistent inhibition of 20 microM adenosine diphosphate (ADP)-stimulated platelet aggregation. Prasugrel 7.5 and 15 mg MDs maintained inhibition in a dose-dependent manner. The pharmacokinetic data indicate that exposure to prasugrel metabolites occurs rapidly after dosing and is consistent with dose proportionality. Within the limitations of this study, the safety and tolerability results suggest that prasugrel is well tolerated when dosed as an initial LD followed by a lower daily MD for 20 days. Prasugrel LDs and MDs provide rapid and sustained inhibition of ADP-mediated platelet aggregation.

Adenosine Diphosphate↗

PubMed2006

Effect of fondaparinux on coagulation assays: results of College of American Pathologists proficiency testing.

CONTEXT: Fondaparinux, a factor Xa inhibitor, is approved for thromboprophylaxis after orthopedic surgery and for treatment of venous thromboembolism. It may also be efficacious, safe, and cost-effective for other patients; thus, more widespread use of fondaparinux is likely. The effect of fondaparinux on coagulation testing needs to be thoroughly examined. OBJECTIVE: To report the effects of fondaparinux on coagulation tests (prothrombin time, activated partial thromboplastin time, fibrinogen, antithrombin, factor VIII, thrombin time, anti-factor Xa) across diverse methodologies. DESIGN: Samples with different concentrations of fondaparinux (0, 0.4, 0.8, and 2.0 microg/mL) were sent to laboratories participating in the College of American Pathologists Comprehensive Coagulation proficiency survey (N = 898). Laboratory-specific methods were used to assay coagulation parameters. RESULTS: Prophylactic or therapeutic fondaparinux prolonged the prothrombin time by approximately 1 second and the activated partial thromboplastin time by 4 to 5 seconds, and reduced factor VIII from 119% to 107% and 102%, respectively. Supratherapeutic fondaparinux reduced factor VIII to 85%. The activated partial thromboplastin time was prolonged in 19%, 29%, and 52% of laboratories with prophylactic, therapeutic, and supratherapeutic fondaparinux levels, respectively. Fibrinogen, antithrombin, and thrombin time assays did not show clinically significant changes. When measuring fondaparinux concentration using an anti-factor Xa assay, the most accurate results were obtained when fondaparinux was used as the calibrator. CONCLUSIONS: Fondaparinux, even in prophylactic doses, slightly prolongs the prothrombin time and activated partial thromboplastin time and can interfere with factor VIII assays, but it has no clinically relevant effect on fibrinogen, antithrombin, or thrombin time. A fondaparinux standard curve should be used for reporting fondaparinux levels using an anti-factor Xa assay.

Anticoagulants↗

PubMed2006

Correlation between the papanicolaou stain and the Wright-Giemsa stain in body fluids: a quality assurance study.

OBJECTIVE: To compare the use of Papanicolaou and Wright-Giemsa stains for the evaluation of body fluids in cytology and hematology laboratories and determine whether other factors account for discrepancies in diagnosis. STUDY DESIGN: We retrospectively reviewed cytopathology reports of peritoneal, pleural, and cerebrospinal fluids received by hematology and cytology laboratories for 1 year. Cases were divided into 3 categories-benign, atypical, and malignant--and slides of discrepant diagnoses were reviewed. RESULTS: During this period, 198 of 3212 (0.61%) cases received by the hematology laboratory and 252 of 4402 (0.57%) cases received by the cytology laboratory were diagnosed as malignant or atypical. Of 3212 cases simultaneously received by the cytology and hematology laboratories, 17 diagnosed as malignant by hematology were diagnosed benign by cytology (sensitivity 96%). Sixteen cases diagnosed as malignant by cytology were diagnosed as benign by hematology (sensitivity 97%). No benign cases were diagnosed as malignant (specificity 100%). Review of the glass slides of the discrepant cases revealed 8 cases undercalled by hematology and 7 cases undercalled by cytology. CONCLUSION: Papanicolaou stain is superior for carcinoma and Wright-Giemsa stain for hematopoietic disorders, but used together they may reduce false negative results. Delays in processing, staining technique, and interobserver variability contribute to discrepancy.

Body Fluids↗

PubMed2005

Effect of double antigen bridging immunoassay format on antigen coating concentration dependence and implications for designing immunogenicity assays for monoclonal antibodies.

The double antigen bridging immunoassay has been used extensively for detection of immunogenicity responses to therapeutic monoclonal antibodies. We have analyzed parameters affecting performance of this type of immunoassay including microtiter plate antigen coating concentration, enzyme-labeled antigen conjugate dilution and assay format (one-step versus two-step). We present results demonstrating that the format of the assay has a significant impact on the optimal parameters to maximize assay performance. A one-step assay format achieves maximal sensitivity across a broad range of coating concentrations and at a lower concentration of conjugate than that in a two-step format. In contrast, a two-step format requires very low coating concentrations and higher conjugate concentrations to achieve maximal sensitivity and suffers from significantly reduced sensitivity at higher coating concentrations. Together, these findings indicate that a one-step assay format can greatly reduce the effect of coating concentration variation on assay performance.

Antibodies, Monoclonal↗

PubMed2005

Development, validation, and implementation of a multiplex immunoassay for the simultaneous determination of five cytokines in human serum.

Quantification of biomarkers can provide important information about the safety and efficacy of candidate drugs. Unfortunately, limited sample volume and excess costs often limit analysis of multiple biomarkers. We developed, optimized, validated, and implemented a multiplex immunoassay for simultaneous measurement of multiple circulating cytokines: IL-1beta, TNFalpha, IL-6, IL-8, and IL-10. Multiplex immuoassays were performed using the Luminex LabMAP instrument. Capture antibodies for each cytokine were covalently bound to distinct microsphere subsets distinguished by differing dye ratios. The concentration of each individual cytokine determined by measuring orange fluorescence produced by a complex of a biotinylated cytokine-specific antibody and streptavidin-phycoerythrin. The lower limit of quantification for all assays was 20 pg/mL with the exception of IL-8 which was 100 pg/mL. The inter-assay precision was less than 25%CV for all analytes at all control levels both pre-study and in-study. The percent recovery ranged from 83 to 108% pre-study and 90 to 125% in-study. In a linearity assessment, a 15,000 pg/mL multi-analyte control could be diluted 1:50 and maintain expected accuracy. We measured the cytokine concentrations in more than 2000 serum samples from patients with sepsis. Multiplex results for IL-6 were compared to a conventional commercially available ELISA kit. The degree of agreement between the two methods as measured by the concordance correlation coefficient was 84.5%. Multiplex results were 2.36-fold higher than ELISA values on the average. After adjusting for this mean difference, the 95% empirical limits of agreement for the ratio of individual sample values were 0.33, 2.65. This multiplex immunoassay provided simultaneous measurement of circulating cytokines using 80% less patient specimen compared to traditional approaches and at a significantly decreased cost. Efficient use of this platform requires process improvements to fully maximize the positive impact of multiplex assays in clinical drug development.

Cytokines↗

PubMed2005

Differential reactivity of anti-primate and anti-human secondary antibodies against human and monkey immunoglobulins: implications for determining the sensitivity of immunogenicity assays.

Development of immunogenicity assays for assessment of human antibodies to therapeutic proteins requires a quantitative determination of assay sensitivity. In the absence of true human positive controls, this is usually accomplished by utilizing affinity-purified antibodies from non-human primates or monoclonal antibodies. In the former case, it is generally considered that non-human primate antibodies will be recognized equally to human antibodies by secondary anti-human immunoglobulin reagents used in immunogenicity assays. We present results here demonstrating that this is not the case. In reality, anti-human immunoglobulin secondary antibodies do not recognize primate immunoglobulins as well as human immunoglobulins. As a result, the use of affinity purified primate antibodies to determine the sensitivity of an immunogenicity assay will likely result in the true sensitivity of the assay being underestimated.

Animals↗

PubMed2003

Survival advantage associated with heterozygous factor V Leiden mutation in patients with severe sepsis and in mouse endotoxemia.

Sepsis is associated with systemic inflammation, coagulopathy, and disrupted protein C (PC) pathway function. The effect of prothrombotic polymorphism, factor V Leiden (Arg506Gln; FV Leiden), was examined in a large clinical trial (PROWESS) of severe sepsis and a mouse endotoxemia model. In PROWESS, 4.1% (n = 65) of patients were heterozygous FV Leiden (VL+/-) carriers. The 28-day mortality was lower in VL+/- (13.9%) than in non-FV Leiden (VL-/-; 27.9%) patients (P =.013). The mortality benefit of recombinant human activated PC (rhAPC) treatment was similar in VL+/- (placebo, 15.6%; rhAPC,12.1%) and VL-/- patients (placebo, 31.0%; rhAPC, 24.7%; interaction P =.981). VL+/- status did not appear to influence baseline biomarkers of coagulopathy and inflammation or disease severity, with the exception that vasopressor usage was less in VL+/- patients (46.2% versus 63.0%; P =.009). In a median lethal dose (40 mg/kg) endotoxin mouse model, VL+/- mice had lower mortality than wild-type mice (19% versus 57%; P =.008), whereas the mortality of homozygous (VL+/+) mice was almost identical to that of wild-type mice (65% versus 57%; P =.76). The findings suggest that FV Leiden constitutes a rare example of a balanced gene polymorphism that maintains the FV Leiden mutation in the general gene pool due to a survival advantage of VL+/- in severe sepsis.

Aged↗

PubMed2002

Laboratory diagnosis of dysfibrinogenemia.

Dysfibrinogenemia is a coagulation disorder caused by a variety of structural abnormalities in the fibrinogen molecule that result in abnormal fibrinogen function. It can be inherited or acquired. The inherited form is associated with increased risk of bleeding, thrombosis, or both in the same patient or family. Traditionally, dysfibrinogenemia is diagnosed by abnormal tests of fibrin clot formation; the thrombin time and reptilase time are the screening tests, and the fibrinogen clotting activity-antigen ratio is the confirmatory test. The inherited form is diagnosed by demonstrating similar laboratory test abnormalities in family members, and if necessary by analysis of the fibrinogen protein or fibrinogen genes in the patient. The acquired form is diagnosed by demonstrating abnormal liver function tests and by ruling out dysfibrinogenemia in family members. This article reviews the laboratory testing of dysfibrinogenemia and presents an algorithm for sequential test selection that can be used for diagnosis.

Algorithms↗

PubMed2002

Plasminogen and tissue-type plasminogen activator deficiency as risk factors for thromboembolic disease.

OBJECTIVE: To review the published evidence for an association between a deficiency of plasminogen or tissue-type plasminogen activator (tPA) and the risk of thrombosis. DATA SOURCES: Review of the medical literature, with an emphasis on the last 10 years. DATA EXTRACTION AND SYNTHESIS: After an initial assessment of the literature, including review of clinical study design and laboratory methods, a draft manuscript summarizing the findings was prepared and circulated to participants in the College of American Pathologists Conference on Diagnostic Issues in Thrombophilia. The key findings and each recommendation were presented for discussion at the conference. Recommendations were accepted if a consensus of the 27 experts attending the conference was reached. The results of the discussion were used to revise the manuscript into its final form. CONCLUSIONS: The consensus of the conference was that routine laboratory assessment of plasminogen and tPA concentration in patients with thrombophilia is not warranted at this time. Analysis of plasminogen and tPA gene alterations in patients with thrombophilia is also not warranted at this time. Determination of plasminogen concentration should be performed in patients suspected of having ligneous conjunctivitis.

Blood Coagulation Tests↗