Determination of the absolute bioavailability of oral imatinib using a stable isotopically labeled intravenous imatinib-d8 microdose.

PURPOSE: The aim of this study was to ascertain whether the absolute bioavailability of oral imatinib (Glivec®) during steady state plasma pharmacokinetics in cancer patients could be determined through a concomitant intravenous administration of a single 100 µg microdose of deuterium labeled imatinib (imatinib-d8). Secondly, the usefulness of liquid chromatography-tandem mass spectrometry (LC-MS/MS) was investigated for simultaneous analysis of orally and intravenously administered imatinib. METHODS: Included patients were on a stable daily dose of 400 mg oral imatinib prior to study participation. On day 1, patients received a 100 µg intravenous imatinib-d8 microdose 2.5 h after intake of the oral dose. Plasma samples were collected for 48 h. Imatinib and imatinib-d8 concentrations were simultaneously quantified using a validated LC-MS/MS assay. The absolute bioavailability was calculated by comparing the dose-normalized exposure with unlabeled and stable isotopically labeled imatinib in plasma. RESULTS: A total of six patients were enrolled. All patients had a history of gastrointestinal stromal tumors (GIST). The median absolute bioavailability of oral imatinib at steady state was 76% (range 44-106%). Imatinib and imatinib-d8 plasma concentrations were quantified in all collected plasma samples, with no samples below the limit of quantification for imatinib-d8. CONCLUSION: The absolute bioavailability of imatinib was successfully estimated at steady state plasma pharmacokinetics using the stable isotopically labeled microdose trial design. This study exhibits the use of a stable isotopically labeled intravenous microdose to determine the absolute bioavailability of an oral anticancer agent in patients with LC-MS/MS as the analytical tool.

Fast and Straightforward Method for the Quantification of Pazopanib in Human Plasma Using LC-MS/MS.

BACKGROUND: Pazopanib is an angiogenesis inhibitor approved for renal cell carcinoma and soft-tissue sarcoma. Studies indicate that treatment with pazopanib could be optimized by adapting the dose based on measured pazopanib plasma concentrations. METHODS: We describe the validation and clinical application of a fast and straightforward method for the quantification of pazopanib in human plasma for the purpose of therapeutic drug monitoring and bioanalytical support of clinical trials. Stable isotopically labeled C,H3-pazopanib was used as internal standard. Plasma samples were prepared for analysis by protein precipitation using methanol and diluted with 10 mmol/L ammonium hydroxide buffer. Chromatographic separation was performed on a C18 column using isocratic elution with ammonium hydroxide in water and methanol. For detection, a tandem mass spectrometer, equipped with a turbo ion spray interface was used in positive ion mode at m/z 438 → m/z 357 for pazopanib and m/z 442 → m/z 361 for the internal standard. RESULTS: Final runtime was 2.5 minutes. All validated parameters were within pre-established limits and fulfilled the FDA and EMA requirements for bioanalytical method validation. After completion of the validation, the routine application of the method was tested by analyzing clinical study samples that were collected for the purpose of therapeutic drug monitoring. CONCLUSIONS: In conclusion, the described method was successfully validated and was found to be robust for routine application to analyze samples from cancer patients treated with pazopanib.

Drug Transporters ABCB1 (P-gp) and OATP, but not Drug-Metabolizing Enzyme CYP3A4, Affect the Pharmacokinetics of the Psychoactive Alkaloid Ibogaine and its Metabolites.

The psychedelic alkaloid ibogaine is increasingly used as an oral treatment for substance use disorders, despite being unlicensed in most countries and having reported adverse events. Using wild-type and genetically modified mice, we investigated the impact of mouse (m)Abcb1a/1b and Abcg2 drug efflux transporters, human and mouse OATP drug uptake transporters, and the CYP3A drug-metabolizing complex on the pharmacokinetics of ibogaine and its main metabolites. Following oral ibogaine administration (10 mg/kg) to mice, we observed a rapid and extensive conversion of ibogaine to noribogaine (active metabolite) and noribogaine glucuronide. Mouse Abcb1a/1b, in combination with mAbcg2, modestly restricted the systemic exposure (plasma AUC) and peak plasma concentration (Cmax) of ibogaine. Accordingly, we found a ∼2-fold decrease in the relative recovery of ibogaine in the small intestine with fecal content in the absence of both transporters compared to the wild-type situation. Ibogaine presented good intrinsic brain penetration even in wild-type mice (brain-to-plasma ratio of 3.4). However, this was further increased by 1.5-fold in Abcb1a/1b;Abcg2 -/- mice, but not in Abcg2 -/- mice, revealing a stronger effect of mAbcb1a/1b in restricting ibogaine brain penetration. The studied human OATP transporters showed no major impact on ibogaine plasma and tissue disposition, but the mOatp1a/1b proteins modestly affected the plasma exposure of ibogaine metabolites and the tissue disposition of noribogaine glucuronide. No considerable role of mouse Cyp3a knockout or transgenic human CYP3A4 overexpression was observed in the pharmacokinetics of ibogaine and its metabolites. In summary, ABCB1, in combination with ABCG2, limits the oral availability of ibogaine, possibly by mediating its hepatobiliary and/or direct intestinal excretion. Moreover, ABCB1 restricts ibogaine brain penetration. Variation in ABCB1/ABCG2 activity due to genetic variation and/or pharmacologic inhibition might therefore affect ibogaine exposure in patients, but only to a limited extent. The insignificant impact of human CYP3A4 and OATP1B1/1B3 transporters may be clinically advantageous for ibogaine and noribogaine use, as it decreases the risks of undesirable drug interactions or interindividual variation related to CYP3A4 and/or OATP activity.

Toxicological analysis of azide and cyanide for azide intoxications using gas chromatography.

Azide is a highly toxic chemical agent to human being. Accidental, but also intentional exposure to azide occurs. To be able to confirm azide ingestion, we developed a method to identify and quantify azide in biological matrices. Cyanide was included in the method to evaluate suggested in vivo production of cyanide after azide ingestion. Azide in biological matrices was first derivatized by propionic anhydride to form propionyl azide. Simultaneously, cyanide was converted into hydrogen cyanide. After thermal rearrangement of propionyl azide, ethyl isocyanate was formed, separated together with hydrogen cyanide by gas chromatography (GC) and detected using a nitrogen phosphorous detector (NPD). The method was linear from 1.0-100 µg/mL for both analytes, and azide was stable in human plasma at -20°C for at least 49 days. Azide was measured in the gastric content of two cases of suspected azide ingestion (case 1:1.2 mg/mL, case 2:1.5 mg/mL). Cyanide was only identified in the gastric content of case 1 (approximately 1.4 µg/mL). Furthermore, azide was quantified in plasma (19 µg/mL), serum (24 µg/mL), cell pellet (21 µg/mL) and urine (3.0 µg/mL) of case 2. This method can be used to confirm azide and cyanide exposure, and azide concentrations can be quantified in several biological matrices.

LC-MS/MS assay for the quantification of testosterone, dihydrotestosterone, androstenedione, cortisol and prednisone in plasma from castrated prostate cancer patients treated with abiraterone acetate or enzalutamide.

Prostate cancer is the most common malignancy among men in the Western world. Treatment of this patient population, e.g. by (chemical) castration, is primarily focused on depletion of tumor-stimulating androgens, with testosterone being the major androgenic hormone. After initial therapy, prostate cancer may progress to metastatic castration-resistant prostate cancer. Anti-hormonal drugs abiraterone acetate and enzalutamide are commonly used to treat patients with this disease as both drugs reduce tumor growth and increase time to tumor progression. To evaluate the pharmacodynamic effects of anti-hormonal drugs in this patient population, we developed an LC-MS/MS method for the quantification of testosterone, dihydrotestosterone, androstenedione, cortisol and prednisone in human plasma. The validated assay ranges from 10-10,000 pg/mL for testosterone and androstenedione, 100-10,000 pg/mL for dihydrotestosterone, 50-5000 pg/mL for cortisol and 500-50,000 pg/mL for prednisone. Intra-assay and inter-assay variabilities were within ±15% of the nominal concentrations for quality control (QC) samples at low, medium and high concentrations and within ±20% at the lower limit of quantification (LLOQ), respectively. The applicability of the method was demonstrated in plasma from patients with metastatic castrated-resistant prostate cancer using either abiraterone acetate or enzalutamide.

Development and validation of a liquid chromatography-tandem mass spectrometry assay for nine oral anticancer drugs in human plasma.

A liquid chromatography-tandem mass spectrometry assay was developed and validated for the nine oral anticancer agents alectinib, cobimetinib, lenvatinib, nintedanib, osimertinib, palbociclib, ribociclib, vismodegib and vorinostat in order to support therapeutic drug monitoring (TDM). The assay was based on reversed-phase chromatography coupled with tandem mass spectrometry operating in the positive ion mode. The assay was validated based on the guidelines on bioanalytical methods by the US Food and Drug Administration and European Medicines Agency. The method was validated over a linear range of 10-200 ng/mL for alectinib, lenvatinib, nintedanib and vismodegib; 50-1000 ng/mL for cobimetinib and palbociclib; 100-2000 ng/mL for osimertinib; 5.00-100 ng/mL for ribociclib; 25-500 ng/mL for vorinostat. Intra-assay and inter-assay bias was within ±20% for all analytes at the lower limit of quantification and within ±15% at remaining concentrations. Stability experiments showed that osimertinib is unstable in the biomatrix and should be shipped on dry-ice and stored at -20 °C until analysis. All other compounds were stable in the biomatrix. The described TDM method was successfully validated and applied for TDM in patients treated with these KIs.