(611d) Non-Protein a All-Membrane Process for Monoclonal Antibody Purification and Efforts on Using Novel Nonwoven Material and Device to Further Improve Efficiency

The downstream processing in mAb manufacturing is facing challenges of increasing production efficiency and lowering production cost, while mAb capture from the harvest by Protein A is the most expensive process. In this study, we developed three different types of membranes with polybutylene terephthalate (PBT) nonwoven as substrate and established a non-protein A three-step nonwoven membrane chromatography for monoclonal antibody purification. It is comprised of a mAb (IgG1) capture step with a novel multimodal anion exchange ligand without any adjustment of CHO supernatant, viral inactivation, a flow-through anion exchange polishing step, and a final bind-and-elute multimodal cation exchange polishing and concentration step. This three-step membrane chromatography reduced HCP from 327000 ppm in the supernatant to 93.5 ppm (3.5 LRV), DNA from 994.7 ppm to 6.8 ppb (5.3 LRV), % aggregates from 5.4% to 0.4% with no detectable fragments, thereby meeting the common requirements (HCP<100 ppm, DNA<10 ppb, % aggregates<1%) for mAb products. A side-by-side comparison with the standard mAb platform (protein A resin and two ion exchange resins) showed that the three-step membrane process outperformed the standard process with significantly shorter process time (3.8 h vs. 13.1 h), 14% higher overall recovery (88.3% vs. 77.5%), fewer steps (no diafiltration for buffer exchange) and fewer aggregates in the final product despite similar HCP and DNA removal.

To improve the purification process we are currently working in two different areas: i) using a novel nonwoven substrate partially composed of smaller fibers to prepare ion exchange membranes and ii) using a membrane cassette, specifically designed for chromatographic separation. With the novel nonwoven substrate both cation and anion exchange membranes were prepared. The cation exchanger was designed to capture the mAb and exhibited a constant 10% dynamic binding capacities (DBC) of ≈100 mg human IgG at 0.25, 0.5 and 1.0 min residence times, which are comparable to that of the PBT based ion exchange membranes, but with much lower pressure drops. The anion exchanger, which is currently under investigation for acidity impurities removal in mAb polishing, has also shown high capacity in binding pure BSA that was used for basic membrane characterization. On the other hand, ChromaWeb® (SPF Technologies, Inc.) cassette which is designed to provide high-resolution and linearly scalable chromatographic performance was employed to be the membrane holder for an anion exchange membrane (PBT-TEA). The packed membranes showed a 10%DBC of 74-84 mg BSA/mL at 0.1 to 1.0 min residence time. It was found that elution pool size remained largely constant from 2.7 to 3.0 membrane volumes (MVs) from 1.0 to 0.1 min residence time, and the elution pool size decreased from 3.7 MVs at 0.3 M NaCl in 20 mM Tris-HCl pH 7.0 to 2.7 MV at 1 M NaCl. Taken together, optimization of the nonwoven substrate structure in terms of fiber diameter, web thickness, and web configuration, as well as the use of a favorable device configuration, are effective to improve the performance of nonwoven chromatographic membranes and provide a significant improvement on downstream processing of mAbs.