Introduction and clearance of beta-glucan in the downstream processing of monoclonal antibodies.

ABSTRACT

ß-Glucan process-related impurities can be introduced into biopharmaceutical products via upstream or downstream processing or via excipients. This study obtained a comprehensive process-mapping dataset for five monoclonal antibodies to assess ß-glucan introduction and clearance during development and production runs at various scales. Overall, 198 data points were available for analysis. The greatest ß-glucan concentrations were found in the depth-filtration filtrate (37-2,745 pg/ml). Load volume correlated with ß-glucan concentration in the filtrate, whereas flush volume was of secondary importance. Cation-exchange chromatography significantly cleared ß-glucans. Furthermore, ß-glucan leaching from the Planova 20N virus removal filter was reduced by increasing the flush volume (1 vs. 10 L/m2 ). ß-glucan concentrations after filter flush with 10 L/m2 were consistently <10 pg/ml. No or only limited ß-glucan clearance was attained via ultrafiltration/diafiltration (UF/DF). However, during the first run with monoclonal antibody (mAb) 4, ß-glucan concentration in the UF/DF retentate was 10.8 pg/mg, potentially due to ß-glucan leaching from the first run with a regenerated cellulose membrane. Overall, ß-glucan levels in the final mAb drug substance were 1-12 pg/mg. Assuming high doses of 1,000-5,000 mg, a ß-glucan contamination at 20 pg/mg would translate to 20-100 ng/dose, which is below the previously suggested threshold for product safety (≤500 ng/dose).