Rapid and Improved Analytical Strategies for Antibody Based Therapeutics using FabRICATOR Digestion

Dr Jonathan Sjögren, Scientific R&D, Genovis, Lund, Sweden

The specificity of the streptococcal IdeS protease (FabRICATOR) has spurred multiple analytical strategies for detailed characterization of therapeutic monoclonal antibodies and related products such as antibody drug conjugates, Fc-fusion proteins, bispecific antibodies, biosimilars, antibody mixtures etc. The enzyme rapidly cleaves at a specific site in the hinge region of IgG generating a F(ab’)2 fragments and two Fc/2 fragments. The cleaved antibody can further be reduced to obtain antibody subunits of 25 kDa, namely the Fd, the LC and the Fc/2 fragments. The size of the fragments allows high-resolution mass spectrometry data to be obtained in a liquid chromatography MS analytical setup, called middle-down. The presentation will cover multiple analytical strategies based on FabRICATOR digestion to monitor critical quality attributes such as oxidation, glycosylation, sequence modifications and others that may occur during discovery, development or production of antibody based therapeutics.