Investigating Aglycosylated Antibody Stability and Retention On Cation Exchange Chromatography

Full-length aglycosylated antibodies have been developed as candidates in several potential therapeutic areas. Purification of these molecules using preparative cation exchange chromatography revealed temperature- and pH-dependent yield losses which are not characteristic of typical glycosylated antibodies. Further investigation suggested that single amino acid substitutions used to eliminate the N-linked glycosylation site in CHO expression systems, rather than the presence or absence of the glycosylation, were responsible for the diminished yields. Several biophysical techniques such as differential scanning calorimetry, circular dichroism, and analytical ultracentrifugation were used in an attempt to correlate solution-phase stability with retention on cation exchange sorbents. Results from this analysis have potential implications for the engineering of therapeutic protein sequences and the selection of mammalian versus bacterial expression systems.