As the industry moves toward continuous bioprocessing, more effective ways are being developed to validate viral clearance. Researchers at Asahi Kasei Bioprocess America have developed a small-scale inline virus spiking and mixing model to validate connected, continuous chromatography and virus filtration systems, without a surge vessel in between.
Ioana Pintescu, research associate and first author on a recent paper, tells GEN this new model defines the “viral clearance capabilities of a virus filter undergoing continuous filtration and what limits we can achieve.” It could allow for fully-continuous bioprocessing, and, based on results from other semi- or fully-continuous downstream processing examples, could save time and money, improve efficiency, and enhance value.
Pintescu and colleagues Julie Kozaili, PhD, principal scientist, and Daniel Strauss, PhD, director of R&D, connected the polishing chromatography and virus filtration steps to create a small-scale virus clearance validation model for continuous virus filtration. The model operated in a high-throughput, constant flow environment for 72 hours. With a viral limit of detection at 0.73 log TCID50/mL, it achieved complete removal of a porcine parvovirus (PPV) and a logarithmic reduction value (LRV) of 5.5 or greater.
As they explain, the challenge for continuous bioprocessing engineers is that most existing virus filtration validation steps were designed for batch processing. Continuous processing conditions, however, are different. The steady movement of product through the system contributes to variations in product feed that may increase system pressure and foul filters, as well as to fluctuations in pH, conductivity, and product concentration. Surge tanks are one solution, but they claim valuable floor space and can constrain operational flexibility and efficiency.
In developing this model, Pintescu and colleagues examined worst-case models and runs that were more representative of actual downstream processes for bovine serum albumin (BSA) and mAb protein production.
“This connected-chromatography-to-virus-filtration model was able to process both BSA and mAb protein solutions without disruption or pressure excursions over 72 hours,” they report.
Achieving that result required “simultaneously accommodating multiple sets of operating parameters and constraints,” they acknowledge. Working stage-by-stage, they set the maximum operating pressure (0.3 megapascals – MPa) based on the parameters of the pump, the column system, and the virus filter, and set a flow rate of 0.3 mL/s for BSA filtrations and 0.165 mL/min for mAb filtrations. “Significantly lower flow rates were selected for the syringe pump,” she notes. Pressure increased gradually throughout the experiment, but there were no overpressure or backpressure issues at any point during the 72-hour run-time.
The system responded comparably to spiked and non-spiked BSA and mAb runs and delivered complete clearance, starting from virus load titers of either 6.25 or 6.38 logTCID50 (tissue culture infectious dose)/mL.
“These results demonstrate that extended filtration run times and high throughput volumes posed no problem for the virus filter in this setup,” they report. “PPV LRV greater than five was achieved for every run, and the filters provided robust viral clearance under the selected conditions.”
As Pintescu says, “Other R&D facilities can reproduce a similar model, so long as they have the correct materials. However, there is limited development on connecting virus filtration to a continuous system for a variety of limitations, including the use or removal of a surge tank. More optimization is needed, and new skid systems will need to be designed to accommodate a continuous feed going through downstream unit operation uninterrupted.”