title: Bioanalytical method development and validation for a large peptide HIV fusion inhibitor (Enfuvirtide, T-20) and its metabolite in human plasma using LC-MS/MS.
abstract: A method for measuring a human immunodeficiency virus (HIV) cell membrane fusion inhibitor (T-20/Ro 29-9800) and its metabolite (M-20/Ro 50-6343) in human plasma by liquid chromatography tandem mass spectrometry (LC-MS/MS) was developed. The relatively large peptide analytes and their corresponding deuterated (d(10)) peptides used as internal standard were isolated from plasma by protein precipitation with two volumes of acetonitrile to plasma. A large pore size reversed-phase C(18) column was employed to elute the peptides. A triple quadrupole mass spectrometer with electrospray interface operating in positive ion and multiple reaction monitoring modes with transitions m/z 1124-->1343 for both T-20 and M-20 was utilized for peak detection. The advantages of the method were a simple sample preparation, specific and sensitive MS/MS detection, and a wide dynamic range of 10-2000 ng/ml for T-20. The method was validated and used for analyzing samples from clinical studies to provide pharmacokinetic profiles of the HIV fusion inhibitor peptide drug and its metabolite.
authors.affiliation: Hoffmann-La Roche, 340 Kingsland Street, Nutley, NJ 07110, USA.
authors.forename: D
authors.initials: D
authors.lastname: Chang
authors.forename: S J
authors.initials: SJ
authors.lastname: Kolis
authors.forename: K H
authors.initials: KH
authors.lastname: Linderholm
authors.forename: T F
authors.initials: TF
authors.lastname: Julian
authors.forename: R
authors.initials: R
authors.lastname: Nachi
authors.forename: A M
authors.initials: AM
authors.lastname: Dzerk
authors.forename: P P
authors.initials: PP
authors.lastname: Lin
authors.forename: J W
authors.initials: JW
authors.lastname: Lee
authors.forename: S K
authors.initials: SK
authors.lastname: Bansal
mesh_terms: D000327:Adsorption; D000595:Amino Acid Sequence; D002138:Calibration; D002853:Chromatography, Liquid / Q000379:methods*; D004355:Drug Stability; D015507:Drugs, Investigational / Q000032:analysis / Q000737:chemistry / Q000627:therapeutic use; D000077560:Enfuvirtide; D015700:HIV Envelope Protein gp41 / Q000097:blood*; D023581:HIV Fusion Inhibitors / Q000097:blood* / Q000737:chemistry / Q000493:pharmacokinetics; D015658:HIV Infections / Q000097:blood / Q000188:drug therapy; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods*; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000097:blood* / Q000493:pharmacokinetics / Q000592:standards; D012015:Reference Standards; D015203:Reproducibility of Results; D013678:Technology, Pharmaceutical / Q000379:methods
chemical_list: D015507:Drugs, Investigational; D015700:HIV Envelope Protein gp41; D023581:HIV Fusion Inhibitors; D010446:Peptide Fragments; D000077560:Enfuvirtide