title: A liquid chromatography/tandem mass spectrometry method for determination of obatoclax in human plasma.
abstract: We describe a selective and highly sensitive high-performance liquid chromatography-electrospray ionization-collision induced dissociation-tandem mass spectrometry (HPLC-ESI-CID-MS/MS) assay for the pan-antiapoptotic Bcl-2 family inhibitor, obatoclax, in human plasma. Fifty μL plasma specimens were prepared by addition of extraction solution consisting of Mobile Phase A (10mM ammonium formate (aq) titrated to pH of 3.0 with formic acid): Mobile Phase B (100% methanol) (20:80, v,v) and internal standard followed by centrifugation. HPLC separations were performed on a Waters, YMC-Pack™, ODS-AQ™ S-3 analytical column with LC mobile phase A and B. Linearity and sensitivity was assessed over a linear range of 0.04-25ng/ml at eleven concentrations. The lower limit of quantification for obatoclax was 0.04ng/mL. The intra-day precision based on the standard deviation of replicates of quality control (QC) samples ranged from 0.9 to 5.1% and the accuracy ranged from 98.9 to 106.8%. Stability studies performed replicate sets of QC samples (0.1ng/mL, 2.5ng/mL, and 15ng/mL) showed that obatoclax in human plasma was stable at room temperature for 24h as well as at -80°C for 1m and 2y. Stability was also demonstrated after 3 freeze/thaw cycles (RT to -80°C). The analytical method showed excellent sensitivity, precision, and accuracy. This method is robust and has been successfully employed in a Children's Oncology Group Phase 1 Consortium study of obatoclax in children with cancer.
authors.affiliation: Division of Clinical Pharmacology and Therapeutics, The Children's Hospital of Philadelphia, United States of America.
authors.forename: Ganesh S
authors.initials: GS
authors.lastname: Moorthy
authors.affiliation: Division of Clinical Pharmacology and Therapeutics, The Children's Hospital of Philadelphia, United States of America; Division of Pediatric Hematology/Oncology University Hospitals Case Medical Center, Rainbow Babies & Children's Hospital, United States of America; Division of Oncology, The Children's Hospital of Philadelphia, United States of America. Electronic address: Robin.Norris@UHhospitals.org.
authors.forename: Robin E
authors.initials: RE
authors.lastname: Norris
authors.affiliation: Division of Clinical Pharmacology and Therapeutics, The Children's Hospital of Philadelphia, United States of America; Division of Oncology, The Children's Hospital of Philadelphia, United States of America.
authors.forename: Peter C
authors.initials: PC
authors.lastname: Adamson
authors.affiliation: Division of Clinical Pharmacology and Therapeutics, The Children's Hospital of Philadelphia, United States of America; Division of Oncology, The Children's Hospital of Philadelphia, United States of America.
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Fox
mesh_terms: D017209:Apoptosis; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004355:Drug Stability; D006801:Humans; D007211:Indoles; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000037:antagonists & inhibitors; D011758:Pyrroles / Q000097:blood*; D011786:Quality Control; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D007211:Indoles; D019253:Proto-Oncogene Proteins c-bcl-2; D011758:Pyrroles; C520962:obatoclax
keywords: Cancer; LC-MS/MS; Obatoclax; Pediatric; Plasma