Quantitation of OSI-7836 (4 ' T hio-araC) and 4 ' Thio-araU in Human Plasma Using LC-MS-MS
Authors
Michael P. Sullivan 1, Christopher J.L. Buggé 1, Orlando E. Espinosa 1, Jim Shen 1, and Dan Drolet 2
1 Cedra Corporation, 8609 Cross Park Drive, Austin, TX 78754
2 OSI, Boulder, Colorado
Purpose
To develop and validate an LC-MS-MS method for the determination of OSI-7836, a nucleoside anti-cancer agent, and 4'thio-araU in human plasma in support of pharmacokinetic profiling.
Methods
This method utilized a solid phase extraction from 0.1 mL of human EDTA plasma (preserved with tetrahydrouridine) with a Supelco Discovery DSC-18 cartridge and methanol as eluant. HPLC separation was done on a PVA-SIL column (4.6 × 100 mm) with an analysis time of 4 min. The mobile phase was acetonitrile/formic acid/water 80:1:20 v/v/v at 1 mL/min. The internal standards were stable isotope forms of the two analytes to account for differences due to adsorption, extraction and instrumental performance. The extracts were analyzed on a SCIEX API-3000 triple quadrupole mass spectrometer equipped with a heated nebulizer interface. Chromatograms were acquired in positive ion MRM mode. Following validation, study samples were analyzed from a clinical trial.
Results
No interfaces were observed at or near the elution times of the analytes or their internal standards. Acceptable intraday and interday precision (<15% CV) and accuracy (<15% difference from theoretical) were obtained over a range of 1-500 ng/mL for OSI-7836 and 5-2500 ng/mL for 4'thio-araU in human plasma. The mean (n=3) correlation coefficients for OSI-7836 and 4'thio-araU were 0.9995 +/- 0.00006 and 0.9969 +/- 0.00133, respectively. The extraction recoveries were close to 90% for the analytes and internal standards. A chromatogram from an analysis of a clinical trial is presented.
Conclusion
A validated method which was robust, sensitive, specific, and accurate was developed and used to quantify OSI-7836 and 4'thio-araU in human plasma samples; this assay has also been validated in dog and mouse plasma and human urine matrices.