Quantitation of dolastatin-10 using HPLC/electrospray ionization mass spectrometry: application in a phase I clinical trial

A highly sensitive and specific assay for the quantitation of the anticancer agent dolastatin-10 (DOL-10) in human plasma is described. The method was based on the use of electrospray ionization-highperformance liquid chromatography/mass spectrometry (ESP-LC/MS). The analytical procedure involved extraction of plasma samples containing DOL-10 and the internal standard (DOL-15) with n-butyl chloride, which was then evaporated under nitrogen. The residue was dissolved in 50 mu l mobile phase and 10 mu l was subjected to ESP-LC/MS analysis using a C-18 microbore column. A linear gradient using water/acetonitrile was used to keep the retention times of the analytes of interest under 5 min. The method exhibited a linear range from 0.005 to 50 ng/ml with a lower limit of quantitation (LLQ) at 0.005 ng/ml. Absolute recoveries of extracted samples in the 85-90% range were obtained. The method's accuracy (less than or equal to 5% relative error) and precision (less than or equal to 10% CV) were well within industry standards. The analytical procedure was applied to extract DOL-10 metabolites from samples obtained following incubation of the drug with an activated S9 rat liver preparation. Two metabolic products were detected and were tentatively identified as a N-demethyl-DOL-10 and hydroxy-DOL-10. Structural assignments were made based on the fragmentation patterns obtained using the electrospray source to produce collision-induced dissociation (CID). The method was also applied to the measurement of DOL-10 in the plasma of patients treated with this drug. Preliminary investigation of the pharmacokinetics suggested that drug distribution and elimination may be best described by a three-compartment model with t(1/2)alpha = 0.087 h, t(1/2)beta = 0.69 h and t(1/2)gamma = 8.0 h. Plasma clearance was 3.7 1/h per m(2).