Measuring Lipolytic Activity to Support Process Improvements to Manage Lipase-Mediated Polysorbate Degradation.

Polysorbates are critical stabilizers in biopharmaceutical protein formulations. However, they may degrade in drug substance (DS) or drug product (DP) during storage. Degradation catalyzed by lipases present in host cell proteins (HCPs) is one suspected root cause. The purpose of this study was to develop an assay to detect lipolytic activity in biopharmaceutical DS and DP formulations. The assay is based on the hydrolysis of the lipase substrate 4-methylumbelliferyl oleate to yield the fluorescent product 4-methylumbelliferone. First, the assay components and their concentrations (buffer salts and pH, solvent and inhibitor Orlistat) were established and optimized using a model lipase (Porcine pancreatic lipase) and cell culture harvest fluid that exhibited lipolytic activity. The assay was then successfully applied and thereby qualified in protein formulations and at lipase concentrations possibly encountered in actual biopharmaceutical DS and DP formulations. The lipase assay can be used to detect lipolytic activity in intermediate and final DS, for example during process optimization in downstream purification, to better and specifically reduce the level, or deplete, lipases from HCPs. The assay is also suitable to be applied during root cause investigations related to polysorbate degradation in biopharmaceutical DP.