Bioavailability of three rufinamide oral suspensions compared with the marketed 400-mg tablet formulation: results from a randomized-sequence, open-label, four-period, four-sequence crossover study in healthy subjects.

BACKGROUND: Rufinamide is indicated for the adjunctive treatment of seizures associated with Lennox-Gastaut syndrome in patients aged ≥4 years. OBJECTIVES: The primary purpose of this study was to compare the relative bioavailability and other pharmacokinetics of rufinamide administered as a 400-mg tablet formulation (reference) with 10 mL of a newly developed 40-mg/mL suspension (test) manufactured using 3 different homogenization speeds in healthy subjects under fed conditions. The study also explored whether homogenization speed had any effect on rufinamide pharmacokinetics when administered as a suspension formulation. METHODS: This was a randomized, open-label, crossover, single-dose study in healthy, fed subjects aged 18 to 55 years (inclusive), conducted at a single center in the United Kingdom. Subjects were randomized to 1 of 4 treatment sequences, with each sequence consisting of 4 treatment periods. In each treatment period, subjects received a single dose of either the reference product (400-mg rufinamide tablet) or the test product (10 mL of rufinamide suspension [40 mg/mL] manufactured using 3 different homogenization speeds [1800, 2100, and 3000 revolutions per minute (rpm)]). Serial blood samples were collected for 72 hours after dosing for the measurement of rufinamide in plasma. Primary comparisons between test (suspension) and reference (tablet) formulations focused on AUC from 0 to 72 hours (AUC(0-72 h)) and C(max). The formulations were considered bioequivalent if the ratios of geometric least squares means and associated 90% CIs of AUC(0-72 h) and C(max) were within the predetermined range of 80%-125%, according to the US Food and Drug Administration (FDA) and European Medicines Agency (EMA) requirements. Tolerability was assessed by subject interviews, physical examinations, and laboratory tests. RESULTS: Twenty-four healthy subjects were randomized: 8 were male and 16 were female; 22 white, 1 black, and 1 Asian subjects were enrolled. Mean (SD) age was 29.8 (10.0) years. Mean weight was 68.2 (11.0) kg, and mean body mass index was 23.6 (3.0) kg/m(2). Twenty-one subjects completed the study; 2 subjects discontinued because of adverse events (both urinary tract infections considered unrelated to treatment) and 1 because of protocol deviation. The 72-hour pharmacokinetic data for the last complete treatment period before discontinuation were included in group means. The geometric least squares mean C(max) value for the reference tablet formulation was 4840.24 ng/mL; and 4254.87, 4204.29, and 4418.44 ng/mL for the 1800-, 2100-, and 3000-rpm test suspensions, respectively. The ratios of the geometric least squares mean values (test/reference) for C(max) were 0.88 (90% CI, 0.84-0.92), 0.87 (0.83-0.91) and 0.91 (0.88-0.95) for the 1800-, 2100-, and 3000-rpm suspensions, respectively, compared with the tablet formulation. The geometric least squares mean AUC(0-72 h) values were 75,960.48 ng · h/mL for the tablet formulation and 74,279.02, 73,746.03, and 73,701.17 ng · h/mL for the 1800-, 2100-, and 3000-rpm suspensions, respectively. The ratios of the geometric least squares mean values (test/reference) for AUC(0-72 h) were 0.98 (90% CI, 0.95-1.00), 0.97 (0.95-1.00) and 0.97 (0.95-0.99) for the 1800-, 2100-, and 3000-rpm suspensions, respectively, compared with the tablet formulation. The ratios and associated 90% CI limits (for test suspensions to the reference tablet) for AUC(0-72 h) and C(max) were within the FDA and EMA criteria for assuming bioequivalence to the 400 mg tablet. Comparisons among the 3 rufinamide test suspensions also met the regulatory criteria for assuming bioequivalence to one another. Treatment-emergent adverse events (TEAEs) were experienced by 18.2% (4/22) of subjects treated with the 400-mg tablet, 21.7% (5/23) of subjects treated with the 1800-rpm suspension, 26.1% (6/23) of subjects treated with the 2100-rpm suspension, and 8.7% (2/23) of subjects treated with the 3000-rpm suspension. Overall, 54.2% (13/24) of subjects experienced a TEAE; all TEAEs were mild or moderate in severity, with headache being the most frequently reported (37.5% [9/24]). There were no serious adverse events or deaths. CONCLUSION: This single-dose study in a small population of fed, healthy subjects found no statistically significant differences in relative bioavailability among each of the 3 test suspensions and the currently marketed 400-mg tablet formulation of rufinamide, meeting FDA and EMA regulatory requirements for assuming bioequivalence.

The haemotoxicity of azathioprine in repeat dose studies in the female CD-1 mouse.

Azathioprine (AZA) is a cytotoxic immunosuppressive drug used in the prevention of rejection in organ transplants and the treatment of auto-immune diseases. However, AZA is haemotoxic causing significant bone marrow depression. The present studies were to characterize the haemotoxicity of AZA in the female CD-1 mouse. In Experiment 1, a dose-ranging study, with AZA gavaged daily for 10 days, clinical evidence of toxicity was evident at 125 mg/kg and above. Experiment 2 was a dose-response study with AZA gavaged daily for 10 days at 40-120 mg/kg. At day 1 after the final dose, AZA induced a dose-related pancytopaenia, reduced femoral marrow cellularity, increases in serum levels of the cytokine fms-like tyrosine kinase 3 ligand, reduction in granulocyte-monocyte colony-forming units and erythroid colonies, and increased bone marrow apoptosis. Histology demonstrated hepatocyte hypertrophy, thymic atrophy, reduced splenic extramedullary haemopoiesis, and reduced cellularity of sternal bone marrow. In Experiment 3, AZA was dosed for 10 days at 100 mg/kg with autopsies at 1, 3, 9, 22, 29, 43 and 57 days postdosing. At 1, 3 and 9 days, haematological parameters reflected changes in Experiment 2. At 22/29 days, many blood parameters were returning towards normal; at 43/57 days, most parameters compared with controls. However, there was some evidence of a persistent (i.e. residual/late-stage) mild reduction in RBC and erythroid progenitor cell counts at day 43/57. We conclude that the CD-1 mouse provides an acceptable model for the haemotoxicity of AZA in man.

Serum FLT-3 ligand in a busulphan-induced model of chronic bone marrow hypoplasia in the female CD-1 mouse.

The concentration of the cytokine fms-like tyrosine kinase-3 ligand (FL) is elevated in the plasma of patients treated with chemotherapy or radiotherapy for malignant conditions. In addition, plasma FL is increased in patients with bone marrow failure resulting from stem-cell defects (e.g. aplastic anaemia). Our goal in the present study was to measure the concentration of serum FL in mice treated with the chemotherapeutic agent busulphan (BU) to induce bone marrow depression and relate changes in FL to effects on haemopoiesis. Female CD-1 mice were treated with BU (9.0 mg/kg) or vehicle by intraperitoneal injection on 10 occasions over 21 days. Animals were autopsied on days 1, 23, 72, 119 and 177 postdosing. A full blood count was performed, and serum prepared for FL analysis. Femoral marrow cell suspensions were prepared to assess the total femoral nucleated cell count (FNCC) and the number of committed haemopoietic progenitor cells (CFU-C). On days 1 and 23 postdosing, significant decreases were evident in many peripheral blood parameters; the FNCC and CFU-C were also reduced in BU-treated mice, in conjunction with increases in serum FL levels. On days 72, 119 and 177 postdosing, several peripheral blood and bone marrow parameters remained reduced and the concentration of serum FL continued to be significantly increased. Linear regression analysis demonstrated significant correlations between the concentration of serum FL in BU-treated mice and peripheral blood and bone marrow parameters; this suggests the possible use of serum FL as a potential biomarker for drug-induced bone marrow injury.

Characterization of troponin responses in isoproterenol-induced cardiac injury in the Hanover Wistar rat.

The investigations aimed to evaluate the usefulness of cardiac troponins as biomarkers of acute myocardial injury in the rat. Serum from female Hanover Wistar rats treated with a single intraperitoneal (IP) injection of isoproterenol (ISO) was assayed for cardiac troponin I (cTnI) (ACS: 180SE, Bayer), cTnI (Immulite 2000, Diagnostic Products Corporation) and cardiac troponin T (cTnT) (Elecsys 2010, Roche). In a time-course study (50.0 mg/kg ISO), serum cTnI (ACS:180SE) and cTnT increased above control levels at 1 hour postdosing, peaking at 2 hours (cTnI, 4.30 microg/L; cTnT, 1.79 microg/L), and declined to baseline by 48 hours, with histologic cardiac lesions first seen at 4 hours postdosing. The Immulite 2000 assay gave minimal cTnI signals, indicating poor immunoreactivity towards rat cTnI. In a dose-response study (0.25 to 20.0 mg/kg ISO), there was a trend for increasing cTnI (ACS:180SE) values with increasing ISO dose levels at 2 hours postdosing. By 24 hours, cTnI levels returned to baseline although chronic cardiac myodegeneration was present. We conclude that serum cTnI and cTnT levels are sensitive and specific biomarkers for detecting ISO induced myocardial injury in the rat. Serum troponin values reflect the development of histopathologic lesions; however peak troponin levels precede maximal lesion severity.