Validation and implementation of drug-dependent antibody assays in clinical trials for safety monitoring of patients dosed with roxifiban, an orally bioavailable glycoprotein IIb/IIIa antagonist.

Thrombocytopenia exposes patients to increased bleeding risk. This serious adverse event was observed with a frequency of approximately 2% in early clinical trials with the potent, orally bioavailable glycoprotein (GP) IIb/IIIa receptor antagonist roxifiban. We previously reported that drug-dependent antibodies (DDAbs) to GP IIb/IIIa are the main cause of thrombocytopenia with roxifiban. Two ELISA assays for detection of free DDAbs (in citrate plasma) and total DDAbs (in EDTA plasma to elute platelet bound DDAbs) were developed and analytically validated. These tests served two purposes during the clinical development program, to pre-screen patients for pre-existing antibodies and monitor patients for increasing antibody titers as a surrogate for eminent thrombocytopenia. The free DDAb assay showed inter and intra-assay precision of 5-12 and 12-14% CV, respectively. The total DDAb assay showed a precision of 5-10 and 4-12% CV, respectively. Three cycles of freeze-thaw did not significantly alter DDAb values in citrate plasma, EDTA plasma or extraction solution. The clinical qualifications of the two assays were conducted in two phase II clinical trials in coronary arterial disease (CAD) patients dosed with roxifiban. Both assays have demonstrated clinical sensitivity of nearly 99-100% and clinical specificity of nearly 95%.

The use of roxifiban (DMP754), a novel oral platelet glycoprotein IIb/IIIa receptor inhibitor, in patients with stable coronary artery disease.

INTRODUCTION: Intravenous platelet glycoprotein (GP) IIb/IIIa receptor inhibitors have a significant beneficial impact on the outcomes of patients undergoing high-risk coronary interventions and in the stabilization of patients with unstable angina pectoris refractory to conventional medical treatment. The role of long-term treatment with oral platelet GP IIb/IIIa receptor inhibitors in patients with coronary artery disease is unproven. This study examined the dose-response effect on inhibition of platelet aggregation by roxifiban (DMP754), a novel oral platelet GP IIb/IIIa receptor inhibitor, and its safety and tolerability in patients with a history of chronic stable angina pectoris. METHODS: Ninety-eight patients were randomized to receive either a placebo or 1 of 8 oral dosages of roxifiban. Twenty-two patients were enrolled in multiple-dose regimens, bringing the total study population to 120. The oral dosages were 0.25, 0.5, 0.75, 1, 1.25, 1.5, 2, or 2.5 mg/day for up to 30 days. RESULTS: Pharmacodynamic response of roxifiban was clearly dose-dependent. Platelet aggregation inhibition in response to 10 micromol/L slope adenosine diphosphate was sustained throughout the study period (up to 1 month). No serious adverse events, including significant major bleeding events, were associated with roxifiban treatment. Minor bleeding was reported in 5% of participants in the placebo group (1 of 21 cases) versus 26% in the study group (26 of 99 cases). Incidence of minor bleeding associated with roxifiban 2 and 2.5 mg/day was significantly (p < or = 0.05) greater than that with placebo. Adverse events, including gastrointestinal disorders, platelet and clotting disorders, and urinary tract disorders, were observed in 1 of 21 cases (5%) in the placebo group and in 12 of 99 cases (12%) in the study group. Reversible thrombocytopenia without other complications developed in two patients. CONCLUSIONS: Roxifiban-induced inhibition of platelet aggregation was dose-dependent and sustained throughout the study period: higher drug dosages correlated with higher levels of platelet inhibition and higher incidence of minor bleeding events. No serious adverse events were observed at any dosage. Thus, roxifiban appears to be a potent oral platelet GP IIb/IIIa receptor inhibitor that is clinically well-tolerated and deserves further study as a new treatment strategy in patients with chronic stable angina pectoris.

Prospective testing for drug-dependent antibodies reduces the incidence of thrombocytopenia observed with the small molecule glycoprotein IIb/IIIa antagonist roxifiban: implications for the etiology of thrombocytopenia.

Thrombocytopenia is a relatively common side effect observed during glycoprotein (GP) IIb/IIIa antagonist therapy. With the oral antagonist roxifiban, we observed thrombocytopenia, defined as 50% reduction of platelets over predose values or below 90 000/microL (9 x 10(10)/L), with a frequency of 2% (8 of 386). Thrombocytopenia occurred either early (days 2 to 4) or delayed (days 11 to 16). No additional cases were observed with up to 6 months of treatment. Retrospective analysis provided evidence for drug-dependent antibodies (DDABs) to GP IIb/IIIa in 5 of 6 subjects, suggestive of an immune etiology of thrombocytopenia. The hypothesis that excluding patients based on positive DDAB reaction would reduce the frequency of thrombocytopenia was tested. Patients were screened for DDABs during the study qualification period and, overall, 3.9% of the patients were excluded based on pre-existing DDAB concentrations above a statistically defined medical decision limit. An additional 2.6% were excluded based on therapy-related antibody production during the first 2 weeks. With antibody testing, 0.2% of patients (2 of 1044) developed immune-mediated thrombocytopenia. One case developed a rapidly increasing antibody concentration and presented with thrombocytopenia despite discontinuation of roxifiban therapy. The second case was related to a false-negative test result. The frequency of thrombocytopenia was statistically significantly reduced from 2% to 0.2% (P =.0007) comparing nonscreened and screened patients. Testing for DDABs can reduce the frequency of thrombocytopenia in patients treated with roxifiban and, by analogy, other GP IIb/IIIa antagonists. Thus, DDAB testing may be employed to increase the safety of GP IIb/IIIa antagonists.

Integrated pharmacokinetic/pharmacodynamic model of XV459, a potent and specific GPIIb/IIIa inhibitor, in healthy male volunteers.

Roxifiban is an oral prodrug of XV459, a potent and specific inhibitor of the glycoprotein (GP) IIb/IIIa receptor previously under investigation for the treatment of peripheral arterial disease and acute coronary care syndrome. The objective of the present analysis was to develop a pharmacokinetic/pharmacodynamic (PK/PD) model that would be used to guide dose selection in Phase 2. This was a randomized, sequential, rising multiple-dose study in 41 healthy male volunteers given doses of 0.5 to 1.25 mg daily for 7 to 10 days. Total XV459 was measured in plasma by a sensitive and specific LC/MS/MS method. The percent inhibition of platelet aggregation (%IPA) was evaluated in citrated plasma in response to 10 microM ADP using the initial slope of the response. The resulting PK data were fit to a two-compartment model with first-order absorption and saturable oral absorption. The pharmacodynamics was modeled using a direct sigmoidal Emax model. Modeling was performed using NONMEM V. Intersubject variability was moderate in both PK and PD (15.3%-18.5%), except for V2/F (64.8%). Residual variability was low at 11.8%. Platelet count influenced both CL/F and EC50. Age and weight did not explain any additional variability in either PK or PD. The model was shown to produce realistic data when used for simulation. Overall, the results suggest that XV459 concentrations in the range of 10 to 20 ng/ml will yield %IPA values in the range of 40% to 80% inhibition. Because of the pharmacodynamically mediated PK of XV459 (due to platelet binding), the EC50 and CL/F are negatively correlated, limiting the utility of plasma concentration monitoring.

Safety, tolerability, pharmacokinetics, and time course of pharmacologic response of the active metabolite of roxifiban, XV459, a glycoprotein IIb/IIIa antagonist, following oral administration in healthy volunteers.

Roxifiban is an esterprodrug that is hydrolyzed, after oral administration, to the active glycoprotein (GP) IIb/IIIa antagonist, XV459. The objectives of the study were to investigate the safety, tolerability, pharmacokinetics, and the time course of the pharmacologic response of XV459 in escalating doses of roxifiban and to assess the effect of age, loading dose of roxifiban, and aspirin pretreatment on XV459 pharmacokinetics, pharmacologic response, and safety profile in a five-part double-blind, placebo-controlled study. Healthy male volunteers (ages 18-46 years) received 7 (0.75-1.5 mg; n = 20) and 10 (0.75-1.0 mg; n = 8) multiple, oral, qd doses of roxifiban or placebo (n = 5). Healthy older male and female volunteers (ages 47-75 years) received roxifiban qd doses (0.5-0.75 mg; n = 8) or placebo (n = 3) for 7 days. Healthy male subjects (ages 18-46 years; n = 16) received a 1.5 or 1.0 mg loading dose either with or without pretreatment of 325 mg aspirin once daily for 3 days followed by single daily doses of 1.0 mg roxifiban for 6 days. Measurable plasma concentrations of XV459 appeared rapidly and were sustained throughout the dosing interval of 24 hours. The pharmacokinetics of XV459 were nonlinear. Systemic exposure of XV459 plateaued at the 1-mg dose level; plasma concentrations approached steady state in 4 to 6 days for doses greater than 1.0 mg. The time course of pharmacologic response as measured by the inhibition of platelet aggregation in response to an ex vivo 10 microM adenosine 5'-diphosphate (ADP) agonist correlated closely to the plasma concentration of XV459. Potent inhibition of ADP-induced platelet aggregation (IPA) persisted over the entire dosing interval. A clear dose response was achieved with roxifiban doses of 0.5 and 1.0 mg. For doses greater than 1.0 mg, a dose-proportional increase in IPA was not observed. Both the pharmacokinetics and pharmacologic response of XV459 exhibited low intraindividual variability (coefficient of variation [CV] < 15%) and higher interindividual variability (CV < 30%). Pretreatment with aspirin and/or a loading dose of 1.5 mg roxifiban had no significant effect on the pharmacokinetics and pharmacologic response of XV459. A dose-related increase in template bleeding time was observed at 1.25- and 1.5-mg doses of roxifiban, as compared to placebo. However, these bleeding time increases in the 1.25- and 1.5-mg dose groups were not significantly different from those at the lower dose groups. Overall, once-daily oral administration of roxifiban was fairly well tolerated and provided sustained systemic drug exposure and pharmacologic response over the entire administration interval.