(238b) Monoclonal Antibody Capture Purification Via Precipitation: Solubility Behavior in the Peg-Zinc Precipitant System

As monoclonal antibody (mAb) upstream titers and production demands continue to increase, the current capture purification processes workhorse, protein A affinity chromatography, has become a significant manufacturing bottleneck. Precipitation is an attractive alternative bulk separation technique that takes advantage of the ever-increasing titer to lower the cost of the overall biomanufacturing process by eliminating protein A chromatography and to improve the environmental sustainability of the process by reducing process mass intensity. Here, two precipitants with orthogonal mechanisms, polyethylene glycol (PEG) 3350 as a volume excluder and zinc chloride (ZnCl₂) as a crosslinker, were examined in a high throughput screening format with six commercially relevant mAbs in both purified drug substance and harvested cell culture fluid (HCCF) formats. A simple solubility model was proposed to explain the variabilities in precipitation behaviors across a range of pure mAbs with different biophysical properties. Furthermore, with the aid of HCCF pretreatments (including pre-concentration and pre-depletion of interfering contaminants) prior to precipitation, we have successfully recovered several commercially relevant mAbs from HCCFs with high yield (over 98% mAb yield) and good quality of stable mAb (above 97% monomer contents), using less than 6mM ZnCl₂ and 3 w/v% PEG for each HCCF. In addition, interesting similarities and variabilities have been observed for multiple mAbs when they were precipitated in (1) purified format, (2) original HCCF format, and (3) HCCF with pretreatments. This high throughput precipitation screening technique, combined with HCCF pretreatments, is an effective approach to optimize and “platformize” operating conditions for monoclonal antibody capture purification via precipitation, facilitating integration of capture via precipitation into a fully continuous, alternative downstream process for mAb production.