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.
Conclusion: 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.
Keywords: CDR; LC–MS; MS; asparagine deamidation; aspartic acid isomerization; immunoprecipitation; in vivo deamidation; in vivo isomerization; mAb therapeutic; monoclonal antibody; therapeutic protein quantification.