Implementing high-temperature short-time media treatment in commercial-scale cell culture manufacturing processes.

The production of therapeutic proteins by mammalian cell culture is complex and sets high requirements for process, facility, and equipment design, as well as rigorous regulatory and quality standards. One particular point of concern and significant risk to supply chain is the susceptibility to contamination such as bacteria, fungi, mycoplasma, and viruses. Several technologies have been developed to create barriers for these agents to enter the process, e.g. filtration, UV inactivation, and temperature inactivation. However, if not implemented during development of the manufacturing process, these types of process changes can have significant impact on process performance if not managed appropriately. This article describes the implementation of the high-temperature short-time (HTST) treatment of cell culture media as an additional safety barrier against adventitious agents during the transfer of a large-scale commercial cell culture manufacturing process. The necessary steps and experiments, as well as subsequent results during qualification runs and routine manufacturing, are shown.

Single pass tangential flow filtration to debottleneck downstream processing for therapeutic antibody production.

As the therapeutic monoclonal antibody (mAb) market continues to grow, optimizing production processes is becoming more critical in improving efficiencies and reducing cost-of-goods in large-scale production. With the recent trends of increasing cell culture titers from upstream process improvements, downstream capacity has become the bottleneck in many existing manufacturing facilities. Single Pass Tangential Flow Filtration (SPTFF) is an emerging technology, which is potentially useful in debottlenecking downstream capacity, especially when the pool tank size is a limiting factor. It can be integrated as part of an existing purification process, after a column chromatography step or a filtration step, without introducing a new unit operation. In this study, SPTFF technology was systematically evaluated for reducing process intermediate volumes from 2× to 10× with multiple mAbs and the impact of SPTFF on product quality, and process yield was analyzed. Finally, the potential fit into the typical 3-column industry platform antibody purification process and its implementation in a commercial scale manufacturing facility were also evaluated. Our data indicate that using SPTFF to concentrate protein pools is a simple, flexible, and robust operation, which can be implemented at various scales to improve antibody purification process capacity.

Isolation and characterization of therapeutic antibody charge variants using cation exchange displacement chromatography.

In this report, we have demonstrated the isolation and enrichment of charge variants of a monoclonal antibody IgG1 using cation exchange displacement chromatography. We successfully achieved the separation of acidic, main and basic charge variants with high recovery (>70%) and purity (>90%) by using a commercially available stationary phase in conjunction with a commercially available displacer. In addition, we have isolated and enriched a trace methionine-oxidized variant of the monoclonal antibody allowing a secondary means of identification of this variant while providing sufficient enrichment for further analysis, stability tests and potency determination. Further characterization of the displacement trains by SEC indicate the possibility of enrichment of high and low molecular weight species. Glycan analysis of the displacement fractions indicates minimal variation in glycan distribution patterns among a wide spectrum of charge variants. These results provide a case study demonstrating the utility of cation exchange displacement chromatography as a viable approach to isolate and enrich antibody charge variants for enhanced molecular characterization.