title: Antibody-conjugated drug assay for protease-cleavable antibody-drug conjugates.
abstract: BACKGROUND
  Antibody-drug conjugates (ADCs) require multiple assays to characterize their PK. These assays can separately evaluate the ADC by quantifying the antibody or the conjugated drug and may give different answers due to assay measurement differences, heterogeneous nature of ADCs and potential biotransformations that occur in vivo.
  
  
  RESULTS
  We present a new version of the antibody-conjugated drug assay for valine-citrulline-linked monomethylauristatin E (vcMMAE) ADCs. A stable isotope-labeled internal standard, protein A affinity capture and solid-phase cleavage of MMAE using papain was used prior to LC-MS/MS analysis.
  
  
  CONCLUSIONS
  The assay was used to assess the difference in ex vivo drug-linker stability of native-cysteine versus engineered cysteine ADCs and to determine the number of drugs per antibody of a native-cysteine ADC in vivo.
authors.affiliation: Seattle Genetics, Inc., 21823 30th Drive SE, Bothell, WA 98021, USA.
authors.forename: Russell J
authors.initials: RJ
authors.lastname: Sanderson
authors.affiliation: Seattle Genetics, Inc., 21823 30th Drive SE, Bothell, WA 98021, USA.
authors.forename: Nicole D
authors.initials: ND
authors.lastname: Nicholas
authors.affiliation: Seattle Genetics, Inc., 21823 30th Drive SE, Bothell, WA 98021, USA.
authors.forename: Cassandra
authors.initials: C
authors.lastname: Baker Lee
authors.affiliation: Seattle Genetics, Inc., 21823 30th Drive SE, Bothell, WA 98021, USA.
authors.forename: Shawna Mae
authors.initials: SM
authors.lastname: Hengel
authors.affiliation: Seattle Genetics, Inc., 21823 30th Drive SE, Bothell, WA 98021, USA.
authors.forename: Robert P
authors.initials: RP
authors.lastname: Lyon
authors.affiliation: Seattle Genetics, Inc., 21823 30th Drive SE, Bothell, WA 98021, USA.
authors.forename: Dennis R
authors.initials: DR
authors.lastname: Benjamin
authors.affiliation: Seattle Genetics, Inc., 21823 30th Drive SE, Bothell, WA 98021, USA.
authors.forename: Stephen C
authors.initials: SC
authors.lastname: Alley
mesh_terms: D000818:Animals; D001681:Biological Assay / Q000379:methods*; D002956:Citrulline / Q000737:chemistry; D004355:Drug Stability; D005260:Female; D006801:Humans; D018796:Immunoconjugates / Q000737:chemistry* / Q000378:metabolism* / Q000493:pharmacokinetics; D009842:Oligopeptides / Q000737:chemistry; D010206:Papain / Q000378:metabolism*; D051381:Rats; D014633:Valine / Q000737:chemistry
chemical_list: D018796:Immunoconjugates; D009842:Oligopeptides; D002956:Citrulline; D010206:Papain; D014633:Valine; C495575:monomethyl auristatin E