Bioequivalence of fixed-dose combination RIN®-150 to each reference drug in loose combination.

BACKGROUND: RIN(®)-150 is a fixed-dose combination (FDC) tablet containing rifampicin (RMP, 150 mg) and isoniazid (INH, 75 mg) developed for the treatment of tuberculosis. SETTING: This study was conducted at a single center: the Pfizer Clinical Research Unit in Singapore. OBJECTIVE: To demonstrate bioequivalence of each drug component between RIN-150 and individual products in a loose combination. DESIGN: This was a randomized, open-label, single-dose, two-way crossover study. Subjects received single doses of RIN-150 or two individual reference products under fasting conditions in a crossover fashion, with at least 7 days washout between doses. The primary measures for comparison were peak plasma concentration (Cmax) and the area under plasma concentration-time curve (AUC). RESULTS: Of 28 subjects enrolled, 26 completed the study. The adjusted geometric mean ratios of Cmax and AUClast between the FDC and single-drug references and 90% confidence intervals were respectively 91.63% (90%CI 83.13-101.01) and 95.45% (90%CI 92.07-98.94) for RMP, and 107.58% (90%CI 96.07-120.47) and 103.45% (90%CI 99.33-107.75) for INH. Both formulations were generally well tolerated in this study. CONCLUSION: The RIN-150 FDC tablet formulation is bioequivalent to the two single-drug references for RMP and INH at equivalent doses.

Pharmacokinetics of azithromycin in serum, bronchial washings, alveolar macrophages and lung tissue following a single oral dose of extended or immediate release formulations of azithromycin.

OBJECTIVES: Antibacterial efficacy of azithromycin could be improved by achieving higher concentrations at the sites of infection. Azithromycin extended release (azithromycin-ER) formulation was developed to enable a higher dosage of 2 g to be administered as a single oral dose without decreasing the safety profile. The aim of this study was to compare the pharmacokinetics of azithromycin in serum, epithelial lining fluid (ELF), alveolar macrophages (AMs) and lung tissue following a single oral dose of azithromycin-ER or azithromycin immediate release (azithromycin-IR) formulation. PATIENTS AND METHODS: A total of 64 patients, diagnosed with lung cancer, requiring open-chest surgery for lung resection, completed the study. Subjects were randomized to receive oral administration of either a single 2 g dose of azithromycin-ER (32 subjects) or a single 500 mg dose of azithromycin-IR (32 subjects). Simultaneously, subjects within each treatment group were randomized to one of eight specific nominal post-dose time points for bronchoalveolar lavage and lung tissue sampling. Results For azithromycin-IR formulation, the AUC(0-24) in serum, ELF, AMs and lung tissue was 3.1, 2.3, 1674 mg.h/L and 130 mg.h/kg, respectively. For azithromycin-ER formulation, the AUC(0-24) in serum, ELF, AMs and lung tissue were 10.0, 17.6, 7028 mg.h/L and 505 mg.h/kg, respectively. The AUC(0-24) ratio following administration of azithromycin-ER relative to azithromycin-IR was 3.2, 7.7, 4.2 and 3.9 in serum, ELF, AMs and lung tissue, respectively. CONCLUSIONS: Within the first 24 h, a single 2 g azithromycin-ER dose produced dose-related increase in systemic exposure compared with a single 500 mg azithromycin-IR dose, which resulted in higher levels of azithromycin in ELF, AMs and lung tissue. Both formulations had similar safety profiles. By achieving high azithromycin exposure early in the course of treatment, without compromising tolerability, azithromycin-ER shows the potential for improved antibacterial efficacy compared with azithromycin-IR.

Comparative pharmacokinetics of voriconazole administered orally as either crushed or whole tablets.

Voriconazole is a triazole antifungal agent used to treat serious, invasive fungal infections including aspergillosis and candidemia. Limitations with existing formulations of voriconazole including restricted utility in patients with renal dysfunction (intravenous preparation) and the unavailability of an oral suspension in some countries make the administration of crushed tablets desirable in many clinical scenarios. However, concerns that this approach may alter the systemic absorption of voriconazole exist. Therefore, an open-label, randomized, two-way crossover comparative pharmacokinetic (PK) study using healthy volunteers was performed to compare these methods of tablet administration. In a random sequence, subjects received voriconazole tablets either crushed or whole. The voriconazole dose was 400 mg every 12 h for 1 day orally followed by 200 mg every 12 h orally for 5.5 days. Study periods were separated by 7 days. PK parameters were determined by the noncompartmental method. An equivalence approach with no-effect boundaries of 80 to 125% was used to assess bioequivalence. Twenty healthy subjects (10 males; aged 20 to 43 years) were enrolled in and completed the study. The adjusted mean areas under the plasma concentration-time curve from 0 to tau, where tau equals 12 h, for the crushed and whole tablet groups were 9,793 and 11,164 ng . h/ml, respectively (ratio, 87.72; 90% confidence interval [CI], 80.97, 95.04). The ratio of the maximum concentration of drug in serum for the crushed tablet versus whole tablet arms was 94.94 (90% CI, 86.51, 104.22). The only difference noted between groups was a slightly faster time to maximum concentration of drug in serum when subjects received crushed tablets, 0.5 h versus 1.5 h (90% CI, -0.75, -0.25). Treatment-related adverse events occurred in 12 subjects receiving whole tablets and 9 subjects receiving crushed tablets; all were mild. The administration of crushed voriconazole tablets is bioequivalent to whole-tablet administration.

Absence of clinically relevant drug interactions following simultaneous administration of didanosine-encapsulated, enteric-coated bead formulation with either itraconazole or fluconazole.

This open-label, two-way crossover study was undertaken to determine whether the enteric formulation of didanosine influences the pharmacokinetics of itraconazole or fluconazole, two agents frequently used to treat fungal infections that occur with HIV infection, and whose bioavailability may be influenced by changes in gastric pH. Healthy subjects were randomized to Treatment A (200-mg itraconazole or 200-mg fluconazole) or Treatment B (same dose of itraconazole or fluconazole with 400 mg of didanosine as an encapsulated, enteric-coated bead formulation). In the itraconazole study, a lack of interaction was concluded if the 90% confidence interval (CI) of the ratio of the geometric means of log-transformed C(max) and AUC(0-T) values of itraconazole and hydroxyitraconazole, the active metabolite of itraconazole, were contained entirely between 0.75 and 1.33. In the fluconazole study, the equivalence interval for C(max) and AUC(0-T) was 0.80-1.25. The data showed that for itraconazole the point estimate and 90% CI of the ratios of C(max) and AUC(0-T) values were 0.98 (0.79, 1.20) and 0.88 (0.71, 1.09), respectively; for hydroxyitraconazole the respective values were 0.91 (0.76, 1.08) and 0.85 (0.68, 1.06). In the fluconazole study, the point estimate and 90% CI of the ratios of C(max) and AUC(0-T) values were 0.98 (0.93, 1.03) and 1.01 (0.99, 1.03), respectively. The T(max) for itraconazole, hydroxyitraconazole, and fluconazole were similar between treatments. Both studies indicated a lack of clinically significant interactions of the didanosine formulation with itraconazole or fluconazole. These results showed that the encapsulated, enteric-coated bead formulation of didanosine can be concomitantly administered with drugs, such as the azole antifungal agents, whose bioavailability may be influenced by interaction with antacids.

Stereoselective metabolism and pharmacokinetics of tegafur.

OBJECTIVES: UFT is composed of racemic tegafur (FT), a prodrug of 5-fluorouracil (5-FU), and uracil in a fixed molar combination (1:4). FT contains a chiral center and has two stereoisomers, R-FT and S-FT. The objectives of this study were to assess the stereoselectivity in the metabolism of FT to 5-FU in vitro and to determine stereoselective differences in the disposition of FT in vivo. METHODOLOGY: R-FT, S-FT, and racemic FT were incubated with pooled human liver microsomes and S-9 fraction for a period of up to 30 min for in vitro studies. For pharmacokinetics, plasma samples were obtained from fasted cancer patients over a period of 24 h after oral administration of 200 mg UFT. Samples from in vitro studies and patient plasma samples were analyzed for FT using a validated achiral and a chiral assay, and for 5-FU using a validated GC/MS assay. RESULTS: R-FT was metabolized at a rate 5.6-times faster than S-FT by human liver microsomes. Similarly, stereoselective metabolism of R-FT was also seen in the S-9 incubations. In cancer patients, the peak plasma concentrations (C(max)) and the time to reach C(max) (T(max)) were similar for the two isomers after the administration of UFT suggesting no apparent differences in their absorption kinetics. However, the area under the curve from zero to infinity [AUC(INF)] and the terminal elimination half-life (T-HALF) values for R-FT were about 4.6- and 4.4-fold lower compared to S-FT, respectively, suggesting the preferential elimination of R-FT. The T-HALF of racemic FT (8.3 h) was comparable to the T-HALF of S-FT (10.3 h) which indicated that the kinetics of the racemate are governed by S-FT. The active cytotoxic moiety, 5-FU, exhibited formation rate limited kinetics from R-FT because the T-HALF of 5-FU (3.4 h) was similar to that of R-FT (2.4 h). CONCLUSIONS: The R-isomer of FT is preferentially metabolized to 5-FU compared to the S-isomer in vitro. The distinct kinetic profiles of the stereoisomers of FT following the administration of UFT is apparently due to the stereoselective disposition of the R-isomer relative to the S-isomer. These data suggest that the R-isomer of FT is worthy of further preclinical and clinical evaluation for safety, efficacy, and pharmacokinetics.

Pharmacokinetics of stavudine and didanosine coadministered with nelfinavir in human immunodeficiency virus-exposed neonates.

We evaluated the pharmacokinetics of stavudine (d4T) and didanosine (ddI) in neonates. Eight neonates born to human immunodeficiency virus-infected mothers were enrolled to receive 1 mg of d4T per kg of body weight twice daily and 100 mg of ddI per m(2) once daily in combination with nelfinavir for 4 weeks after birth. Pharmacokinetic evaluations were performed at 14 and 28 days of age. For d4T, on days 14 and 28, the median areas under the concentration-time curves from 0 to 12 h (AUC(0-12)s) were 1,866 and 1,603, ng x h/ml, respectively, and the median peak concentrations (C(max)s) were 463 and 507 ng/ml, respectively. For ddI, on days 14 and 28, the median AUC(0-10)s were 1,573 and 1,562 h x ng/ml, respectively, and the median C(max)s were 627 and 687 ng/ml, respectively. Systemic levels of exposure to d4T were comparable to those seen in children, suggesting that the pediatric dose of 1 mg/kg twice daily is appropriate for neonates at 2 to 4 weeks of age. Levels of exposure to ddI were modestly higher than those seen in children. Whether this observation warrants a reduction of the ddI dose in neonates is unclear.

Effect of food on the oral bioavailability of UFT and leucovorin in cancer patients.

UFT is composed of tegafur (FT), a prodrug of 5-fluorouracil (5-FU), and uracil in a fixed combination (1:4). In conjunction with leucovorin, UFT is being developed for the first-line oral treatment of metastatic colorectal cancer. The effect of food on the oral bioavailability of UFT (2 x 100 mg capsules; dose in terms of FT) and leucovorin (2 x 15 mg tablets) was evaluated in a single-dose, randomized, two-way crossover study. Patients (n = 25) were assigned to receive both drugs after an overnight fast or 5 min after completion of a high-fat meal (721 calories) with a 3-day washout period between treatments; then they were permitted to continue on oral UFT/leucovorin therapy for safety assessment. UFT (300 mg/m2/day as three divided doses) and leucovorin (90 mg/day as three divided doses) were given for 28 days. After a 7-day rest, the 28-day cycle was repeated. Pharmacokinetics (n = 22 patients) were determined for FT, 5-FU, uracil, leucovorin, and 5-methyltetrahydrofolate (an active metabolite of leucovorin). The absence of food-effect on peak plasma concentration (CMAX) and the area under the curve (AUC) was concluded if the 90% confidence interval for the ratio of the treatment means was entirely contained in 0.75-1.33. Administration of UFT with food resulted in a 34% decrease in CMAX of FT, whereas the AUC of FT remained unchanged. Food decreased the CMAX and AUC values of uracil and 5-FU by 37-76%. On the contrary, the CMAX and AUC values of leucovorin and 5-methyltetrahydrofolate were increased by 14-60% with food. Time to reach CMAX for all analytes was significantly (P < or = 0.001) delayed by food. Except for the AUCs of FT, the statistical criterion for concluding a lack of food-effect was not met. These data suggest that UFT/leucovorin should not be dosed simultaneously with food. It is recommended that food should not be consumed for 1 h before and after an oral dose of UFT and leucovorin in a manner similar to pivotal Phase III trials. The 28-day oral regimen of UFT and leucovorin was generally well tolerated in the population studied.

Influence of immunogenicity on the pharmacokinetics of BMS-191352, a Pseudomonas exotoxin immunoconjugate, in rats and dogs.

BMS-191352 is an immunotoxin construct of modified Pseudomonas exotoxin conjugated to a fragment of the BR96 monoclonal antibody. We have investigated the potential for immunogenicity of BMS-191352 and its influence on the pharmacokinetics in rats and dogs. BMS-191352 was administered intravenously at doses of 0.75, 1.5, and 3 mg m(-2) once every two days for a total of five doses in rats, and 1.2, 2.4, and 4.8 mg m(-2) once every three days for a total of five doses in dogs. Blood samples were collected on days 1 and 9 in rats, and on days 1, 7, and 13 in dogs to monitor pharmacokinetics and anti-BMS-191352 immune response. Plasma concentrations of BMS-191352 and serum anti-BMS-191352 antibody titre were determined using ELISA assays. Pharmacokinetics were assessed using a non-compartmental method. Anti-BMS-191352 antibodies were not observed in rats within the drug administration interval. In all dogs, except one, markedly higher anti-BMS-191352 antibody titres were observed on day 13 compared with days 1 and 7, and its magnitude was independent of BMS-191352 dose. The single dose kinetics of BMS-191352 in rats and dogs were linear and the drug exposures were generally dose proportional. Mean half-life, total body clearance, and volume of distribution were 1.74 h, 3.35 mL min(-1) m(-2), and 0.27 Lm(-2) in rats, respectively, and 4.27 h, 6.28 mL min(-1) m(-2), 1.19 L m(-2) in dogs, respectively. The multiple-dose (day 9) kinetics in rats were similar to the single-dose kinetics. In dogs, the disposition of BMS-191352 on day 7 was similar to that on day 1; however, there was a precipitous reduction in the systemic drug exposure (by 5- to 110-fold) and marked increase in drug clearance on day 13. These changes in the kinetics of BMS-191352 were attributed to the generation of anti-BMS-191352 antibodies. In the one dog that did not develop anti-BMS-191352 antibodies, the pharmacokinetics were unchanged. The pharmacokinetics of BMS-191352 may be perturbed due to an immune response thus restricting the therapeutic utility of the immunotoxin.

BR96 sFv-PE40 immunotoxin: nonclinical safety assessment.

BR96 sFv-PE40, a recombinant DNA-derived fusion protein composed of the heavy- and light-chain variable region domains of the monoclonal antibody BR96 and the translocation and catalytic domains of Pseudomonas exotoxin A, is being developed for the treatment of solid tumors expressing cell surface Lewis(y)-related antigens. Single- and repeat-dose intravenous toxicity studies in rats and dogs and a comparative ex vivo tissue-binding study with rat, dog, and human tissues were conducted to assess the toxicity of BR96 sFv-PE40 and to estimate a safe starting dose in humans. Additional studies were performed to investigate the prevention of pulmonary vascular-leak syndrome, the dose-limiting toxicity of BR96 sFv-PE40 in rats, and the immunogenicity of BR96 sFv-PE40. In single-dose studies in rats, the vascular leak appeared to be primarily confined to the lungs; however, with a repeat-dose regimen (every other day for 5 doses) other organs including the brain and heart were involved at lethal doses (12-15 mg/m2 cumulative). Single doses of 1.8 mg/m2 and a cumulative 3.8 mg/m2 dose (0.75 mg/m2, every other day for 5 doses) were generally well tolerated in rats. These doses are significantly greater than doses required to cure rodents bearing human tumor xenografts. In dogs, the major target organ following single or repeated doses (every 3 days for 5 doses) was the pancreas. Morphologic changes in the exocrine pancreas ranged from atrophy with single-cell necrosis to diffuse acinar necrosis. After a 1-mo dose-free observation period, no residual pancreatic toxicity was observed in dogs given single doses up to 6.0 mg/m2 or 5 doses of 2.4 mg/m2 (12 mg/m2 cumulative). No significant pancreatic toxicity was observed at doses <0.6 mg/m2 in high Lewis(y)-expressing dogs. Assessment of trypsinlike immunoreactivity was useful in monitoring changes in pancreatic function. The immunogenicity of BR96 sFv-PE40 could be inhibited by combined treatment with an immunosuppressant in dogs, thus maintaining exposure to BR96 sFv-PE40.

Development of an immunoassay for BMS-191352, a single-chain immunotoxin, and its application to toxicokinetic studies.

BMS-191352 is a single-chain fusion protein composed of the variable regions of chimeric BR96 monoclonal antibody and the binding defective form of Pseudomonas Exotoxin A (PE40). The immunotoxin exhibits potent cytotoxicity against tumor cells expressing the Lewis antigen. A sensitive and specific double antibody sandwich ELISA has been developed and validated for the determination of BMS-191352 in rat and dog EDTA plasma. A monoclonal anti-PE40 antibody (EXA2-1H8) was used to capture BMS-191352 in plasma samples. The captured BMS-191352 was then detected using a biotinylated monoclonal BR96 antiidiotypic antibody (757-4-1) followed by the addition of streptavidin-horseradish peroxidase conjugate and chromogen 3,3',5,5'-tetramethylbenzidine. The optical density was measured at 450 nm. The standard curve range in rat and dog plasma was 2-32 ng/mL. The RSD for the inter- and intra-assay precision was within 9.2% and the accuracy was greater than 89.0%. The ELISA method was applied to the analysis of BMS-191352 in plasma samples from toxicokinetic studies conducted in rats and dogs. These studies revealed that the systemic exposure of BMS-191352 was dose proportional and the kinetics of BMS-191352 were linear between the dose range of 1.8-7.2 mg/m2 in the dog.

Reversal of doxorubicin resistance in multidrug resistant melanoma cells in vitro and in vivo by dipyridamole.

The occurrence of multidrug resistance (MDR) decreases the clinical utility of several anticancer agents, including doxorubicin (DOX). A transmembrane efflux pump, P-glycoprotein (P-gp), is frequently implicated in the development of MDR in tumor cells. Dipyridamole (DP), a clinically used antiplatelet drug, enhances the cytotoxicity of the anticancer drugs affected by MDR. Although this aspect has been studied extensively in cell culture models, the effectiveness of DP to overcome multidrug resistance has not been investigated using in vivo models of multidrug-resistant solid tumors. Therefore, the objective of this study was to evaluate the role of DP in the reversal of resistance to DOX in tumor-bearing mice in the context of its anti-MDR activity in vitro. For this purpose, drug-sensitive murine melanoma cells (B16V) and their DOX-selected MDR variant, B16VDXR cells, were used. In vitro, the reversal of DOX resistance of B16VDXR cells by DP was determined using clonogenic assays, and the influence of DP on the transport of DOX was evaluated by measurement of steady-state accumulation as well as efflux of DOX in B16VDXR cells. Antitumor activity of different treatments was assessed by monitoring tumor growth. Pharmacokinetics of DOX, with or without DP, were evaluated in C57BL/6 mice bearing B16V or B16VDXR tumors. DP produced a 6.4-fold reversal of resistance to DOX in vitro; this was accompanied by an increase (3.6-fold) in the steady-state intracellular accumulation of DOX and a marked reduction in the efflux of DOX from B16VDXR cells. Furthermore, a linear correlation was observed between the EC50 values and the steady-state intracellular levels of DOX in the multidrug-resistant cells. In the in vivo experiments, similar growth patterns were seen for the DOX alone and the DOX+DP groups for B16V tumors. The results with B16VDXR tumors were in sharp contrast. The DOX+DP treatment caused a significant delay in the growth of B16VDXR tumors compared to treatment with DOX alone or controls. DP did not alter the plasma pharmacokinetics of DOX in C57BL/6 mice but resulted in a significant increase in the intratumoral accumulation of DOX.

Effect of tamoxifen on mitoxantrone cytotoxicity in drug-sensitive and multidrug-resistant MCF-7 cells.

The influence of the antiestrogen tamoxifen (TAM) on the activity of mitoxantrone (MXN), was evaluated against wild-type MCF-7/WT and their multidrug-resistant variant MCF-7/ADR cells. Multidrug resistance (MDR) in this cell line which was selected for resistance to Adriamycin (ADR), is associated with increased expression of P-glycoprotein (P-gp). In a clonogenic assay it was observed that TAM (1-10 microM) significantly enhanced the activity of MXN in the MCF-7/ADR but not in the drug-sensitive cell line. Isobologram analysis indicated that the effect of the combination was additive in the parental MCF-7/WT cells and strongly synergistic in the MDR MCF-7/ADR cells. Also, TAM (10 microM) caused a three-fold increase in the steady-state levels (Css) of MXN in MCF-7/ADR cells but did not modulate MXN levels in MCF-7/WT cells. The observed synergism in MCF-7/ADR cells was perhaps due to the increase in Css of MXN that may involve interaction of TAM with P-gp. The combination of MXN and TAM may be useful in the treatment of drug-sensitive and drug-resistant breast cancer.

Dipyridamole reverses the resistance to topoisomerase II inhibitors but not to antimicrotubule agents in multidrug-resistant melanoma cells.

The influence of dipyridamole (DP) on the cytotoxicity and cellular disposition of several DNA topoisomerase II (topo II) inhibitors and antimitotic agents in multidrug-resistant B16VDXR cells was examined. B16VDXR cells, derived from parental B16V cells by step-wise treatment with doxorubicin (DOX), overexpress a 170 kDa P-glycoprotein (P-gp). Additionally, the resistance to DOX in B16VDXR cells is associated with decreased frequency of DNA strand breaks compared to that in the drug-sensitive B16V cells. DP (10 microM) significantly (P < 0.01) potentiated the cytotoxicity of DOX (6.4-fold), mitoxantrone (2.3-fold), and etoposide (14-fold) in the drug-resistant B16VDXR cells. This was accompanied by a 3.7-fold and 4.2-fold increase in the total intracellular and nuclear levels of DOX, respectively. Surprisingly, no significant change in the intracellular and nuclear levels or the efflux of etoposide was observed in B16VDXR cells. Combination index (CI) analysis, however, indicated that DP interacted synergistically with DOX as well as etoposide. Further, it was intriguing to observe that DP (10 microM) failed to modulate the resistance to vincristine, vinblastine, and taxol. This was despite a significant increase in the accumulation of vinblastine (3.3-fold) and taxol (3.9-fold) in B16VDXR cells in the presence of DP (10 microM). The observed pattern of chemosensitization suggests that in addition to interaction with P-gp, the multidrug-resistance modulating activity of DP may involve P-gp independent mechanism(s). The possibilities include that (i) DP interacts with topo II or (ii) DP promotes the formation and/or obstructs the repair of DNA strand breaks caused by topo II inhibitors.

Dipyridamole modulates multidrug resistance and intracellular as well as nuclear levels of doxorubicin in B16 melanoma cells.

Simultaneous occurrence of resistance to many chemotherapeutic agents, termed multidrug resistance (MDR), is a complex phenotype. MDR occurs due to several reasons, including over-expression of a 170-kDa membrane-bound protein, called P-glycoprotein (P-gp), which apparently participates in active drug efflux. Multidrug-resistant cells also frequently exhibit an altered pattern of intracellular drug distribution, resulting in a reduction in the nuclear level of drugs such as doxorubicin (DOX). In this study, the effect of dipyridamole (DP) on drug resistance and on intracellular as well as nuclear levels of DOX in multidrug-resistant melanoma cells has been examined. For this purpose, drug-sensitive murine melanoma cells (B16V) and their multidrug-resistant variant cells, (B16VDXR; selected for resistance to DOX) which over-produce P-gp, were employed. B16VDXR cells were cross-resistant to several anti-cancer agents including etoposide (VP-16) and mitoxantrone (Mitox). DP (10 microM) significantly potentiated the cytotoxicity of DOX, VP-16 and Mitox towards multidrug-resistant B16VDXR cells but not in parental drug-sensitive B16V cells. The presence of DP resulted in a 3.7-fold increase in the total cellular level and a 4.2-fold increase in the nuclear content of DOX in the resistant cells. Isobologram analysis indicates that DP at several pharmacologically relevant concentrations synergistically potentiates the activity of DOX in B16VDXR cells.