title: A robust and rapid liquid chromatography tandem mass spectrometric method for the quantitative analysis of 5-azacytidine.
abstract: The DNA methyltransferase inhibitor 5-azacytidine is being evaluated clinically as an oral formulation to treat various solid tumors. A sensitive, reliable method was developed to quantitate 5-azacytidine using LC-MS/MS to perform detailed pharmacokinetic studies. The drug of interest was extracted from plasma using Oasis MCX ion exchange solid-phase extraction 96-well plates. Chromatographic separation was achieved with a YMC J'sphere M80 C18 column and isocratic elution with a methanol-water-formic acid (15:85:0.1, v/v/v) mobile phase over a 7 min total analytical run time. An AB Sciex 5500 triple quadrupole mass spectrometer operated in positive electrospray ionization mode was used for the detection of 5-azacytidine. The assay range was 5-500 ng/mL and proved to be accurate (97.8-109.1%) and precise (CV ≤ 9.8%). Tetrahydrouridine was used to stabilize 5-azacytidine in blood/plasma samples. With the addition of tetrahydrouridine, long-term frozen plasma stability for 5-azacytidine at -70°C has been determined for at least 323 days. The method was applied for the measurement of total plasma concentrations of 5-azacytidine in a cancer patient receiving a 300 mg oral daily dose.
authors.affiliation: The Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins University, Baltimore, MD, USA.
authors.forename: Nicole M
authors.initials: NM
authors.lastname: Anders
authors.affiliation: The Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins University, Baltimore, MD, USA.
authors.forename: Teresia M
authors.initials: TM
authors.lastname: Wanjiku
authors.affiliation: The Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins University, Baltimore, MD, USA.
authors.forename: Ping
authors.initials: P
authors.lastname: He
authors.affiliation: The Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins University, Baltimore, MD, USA.
authors.forename: Nilofer S
authors.initials: NS
authors.lastname: Azad
authors.affiliation: The Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins University, Baltimore, MD, USA.
authors.forename: Michelle A
authors.initials: MA
authors.lastname: Rudek
mesh_terms: D000964:Antimetabolites, Antineoplastic / Q000097:blood* / Q000737:chemistry / Q000493:pharmacokinetics / Q000627:therapeutic use; D001374:Azacitidine / Q000097:blood* / Q000737:chemistry / Q000493:pharmacokinetics / Q000627:therapeutic use; D002853:Chromatography, Liquid / Q000379:methods*; D004355:Drug Stability; D006801:Humans; D009369:Neoplasms / Q000188:drug therapy; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D000964:Antimetabolites, Antineoplastic; D001374:Azacitidine
keywords: 5-azacytidine; DNA methyltransferase inhibitor; LC/MS/MS; pharmacokinetics