title: Quantification of in vivo site-specific Asp isomerization and Asn deamidation of mAbs in animal serum using IP-LC-MS.
abstract: BACKGROUND
  Isomerization of aspartic acid and deamidation of asparagine are two common amino acid modifications that are of particular concern if located within the complementarity-determining region of therapeutic antibodies. Questions arise as to the extent of modification occurring in circulation due to potential exposure of the therapeutic antibody to different pH regimes.
  
  
  RESULTS
  To enable evaluation of site-specific isomerization and deamidation of human mAbs in vivo, immunoprecipitation (IP) has been combined with LC-MS providing selective enrichment, separation and detection of naive and modified forms of tryptic peptides comprising complementarity-determining region sequences.
  
  
  CONCLUSIONS
  IP-LC-MS can be applied to simultaneously quantify in vivo drug concentrations and measure the extent of isomerization or deamidation in PK studies conducted during the drug discovery stage.
authors.affiliation: Bristol Myers Squibb Co, Princeton, NJ 08543, USA.
authors.forename: John T
authors.initials: JT
authors.lastname: Mehl
authors.affiliation: Bristol Myers Squibb Co, Princeton, NJ 08543, USA.
authors.forename: Bogdan G
authors.initials: BG
authors.lastname: Sleczka
authors.affiliation: Bristol Myers Squibb Co, Princeton, NJ 08543, USA.
authors.forename: Eugene F
authors.initials: EF
authors.lastname: Ciccimaro
authors.affiliation: Bristol Myers Squibb Co, Princeton, NJ 08543, USA.
authors.forename: Alexander T
authors.initials: AT
authors.lastname: Kozhich
authors.affiliation: Bristol Myers Squibb Co, Seattle, WA, USA.
authors.forename: Deb G
authors.initials: DG
authors.lastname: Gilbertson
authors.affiliation: Bristol Myers Squibb Co, Princeton, NJ 08543, USA.
authors.forename: Ragini
authors.initials: R
authors.lastname: Vuppugalla
authors.affiliation: Bristol Myers Squibb Co, Princeton, NJ 08543, USA.
authors.forename: Christine S
authors.initials: CS
authors.lastname: Huang
authors.affiliation: Bristol Myers Squibb Co, Seattle, WA, USA.
authors.forename: Brenda
authors.initials: B
authors.lastname: Stevens
authors.affiliation: Johnson & Johnson, Malvern, PA, USA.
authors.forename: Jingjie
authors.initials: J
authors.lastname: Mo
authors.affiliation: Bristol Myers Squibb Co, Princeton, NJ 08543, USA.
authors.forename: Ekaterina G
authors.initials: EG
authors.lastname: Deyanova
authors.affiliation: Bristol Myers Squibb Co, Princeton, NJ 08543, USA.
authors.forename: Yun
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Bristol Myers Squibb Co, Princeton, NJ 08543, USA.
authors.forename: Richard Yc
authors.initials: RY
authors.lastname: Huang
authors.affiliation: Bristol Myers Squibb Co, Princeton, NJ 08543, USA.
authors.forename: Guodong
authors.initials: G
authors.lastname: Chen
authors.affiliation: Bristol Myers Squibb Co, Princeton, NJ 08543, USA.
authors.forename: Timothy V
authors.initials: TV
authors.lastname: Olah
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000097:blood / Q000737:chemistry*; D001216:Asparagine / Q000032:analysis*; D001224:Aspartic Acid / Q000032:analysis*; D002853:Chromatography, Liquid / Q000379:methods; D006801:Humans; D047468:Immunoprecipitation / Q000379:methods; D007536:Isomerism; D008252:Macaca fascicularis; D008297:Male; D053719:Tandem Mass Spectrometry / Q000379:methods
chemical_list: D000911:Antibodies, Monoclonal; D001224:Aspartic Acid; D001216:Asparagine
keywords: CDR; LC–MS; MS; asparagine deamidation; aspartic acid isomerization; immunoprecipitation; in vivo deamidation; in vivo isomerization; mAb therapeutic; monoclonal antibody; therapeutic protein quantification