A new high-performance liquid chromatography-tandem mass spectrometry method for the determination of paclitaxel and 6α-hydroxy-paclitaxel in human plasma: Development, validation and application in a clinical pharmacokinetic study

<div><p>Paclitaxel belongs to the taxanes family and it is used, alone or in multidrug regimens, for the therapy of several solid tumours, such as breast-, lung-, head and neck-, and ovarian cancer. Standard dosing of chemotherapy does not take into account the many inter-patient differences that make drug exposure highly variable, thus leading to the insurgence of severe toxicity. This is particularly true for paclitaxel considering that a relationship between haematological toxicity and plasma exposure was found. Therefore, in order to treat patients with the correct dose of paclitaxel, improving the overall benefit–risk ratio, Therapeutic Drug Monitoring is necessary. In order to quantify paclitaxel and its main metabolite, 6α-hydroxy-paclitaxel, in patients’ plasma, we developed a new, sensitive and specific HPLC–MS/MS method applicable to all paclitaxel dosages used in clinical routine. The developed method used a small volume of plasma sample and is based on quick protein precipitation. The chromatographic separation of the analytes was achieved with a SunFire^™ C18 column (3.5 μM, 92 Å, 2,1 x 150 mm); the mobile phases were 0.1% formic acid/bidistilled water and 0.1% formic acid/acetonitrile. The electrospray ionization source worked in positive ion mode and the mass spectrometer operated in selected reaction monitoring mode. Our bioanalytical method was successfully validated according to the FDA-EMA guidelines on bioanalytical method validation. The calibration curves resulted linear (R² ≥0.9948) over the concentration ranges (1–10000 ng/mL for paclitaxel and 1–1000 ng/mL for 6α-hydroxy-paclitaxel) and were characterized by a good accuracy and precision. The intra- and inter-day precision and accuracy were determined on three quality control concentrations for paclitaxel and 6α-hydroxy-paclitaxel and resulted respectively <9.9% and within 91.1–114.8%. In addition, to further verify the assay reproducibility, we tested this method by re-analysing the incurred samples. This bioanalytical method was employed with success to a genotype-guided phase Ib study of weekly paclitaxel in ovarian cancer patients treated with a wide range of drug’s dosages.</p></div

List of publications related to LC-MS/MS methods for the quantification of PTX in human plasma samples.

<p>List of publications related to LC-MS/MS methods for the quantification of PTX in human plasma samples.</p

Recovery of the analytes and the IS from human plasma.

<p>Recovery of the analytes and the IS from human plasma.</p

Source- and compound-dependent parameters and ion transitions of each analyte and IS used for the mass spectrometer method.

<p>Source- and compound-dependent parameters and ion transitions of each analyte and IS used for the mass spectrometer method.</p

Chemical structures of PTX, 6α-OH-PTX, and DTX (docetaxel) used as IS.

<p>Chemical structures of PTX, 6α-OH-PTX, and DTX (docetaxel) used as IS.</p

MS/MS mass spectra of PTX and 6α-OH-PTX with chemical structures and identification of the main fragment ions.

<p>MS/MS mass spectra of PTX and 6α-OH-PTX with chemical structures and identification of the main fragment ions.</p

Intra and inter-day precision and accuracy of the method for the analysis of PTX and its metabolite 6α-OH-PTX in human plasma samples.

<p>Intra and inter-day precision and accuracy of the method for the analysis of PTX and its metabolite 6α-OH-PTX in human plasma samples.</p

Plasma concentration-<i>vs</i>-time profiles of PTX and its main metabolite 6α-OH-PTX in three patients with advanced ovarian cancer.

<p>Patient 1, 2, and 3 received 110 mg/m² of PTX as 1-h intravenous infusion during the first chemotherapy cycle.</p

Re-analysis of incurred plasma samples of one patient treated at the dose of 100 mg/m² of PTX during the first chemotherapy cycle.

<p>Re-analysis of incurred plasma samples of one patient treated at the dose of 100 mg/m² of PTX during the first chemotherapy cycle.</p