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A Combined Ultrafiltration/Diafiltration Process for the Purification of Oncolytic Measles Virus
Daniel Loewe1, Hauke Dieken1, Tanja A Grein1
1Institute of Bioprocess Engineering and Pharmaceutical Technology, University of Applied Sciences Mittelhessen, 35390 Giessen, Germany.
Abstract:
Measles virus (MV) is an important representative of a new class of cancer therapeutics known as oncolytic viruses. However, process intensification for the downstream purification of this fragile product is challenging. We previously found that a mid-range molecular weight cut-off (300 kDa) is optimal for the concentration of MV. Here, we tested continuous and discontinuous diafiltration for the purification of MV prepared in two different media to determine the influence of high and low protein loads. We found that a concentration step before diafiltration improved process economy and MV yield when using either serum-containing or serum-free medium. We also found that discontinuous diafiltration conferred a slight benefit in terms of the permeate flow, reflecting the repetitive dilution steps and the ability to break down parts of the fouling layer on the membrane. In summary, the combined ultrafiltration/diafiltration process is suitable for the purification of MV, resulting in the recovery of ~50% infectious virus particles with a total concentration factor of 8 when using 5 diavolumes of buffer.
Insights
Purifying measles virus (MV) for cancer therapy is challenging. A combined ultrafiltration/diafiltration process effectively concentrates and purifies MV, yielding about 50% infectious virus particles.
Area of Science:
- Virology
- Biotechnology
- Cancer Therapeutics
Background:
- Measles virus (MV) shows promise as an oncolytic virus therapy for cancer.
- Downstream purification of fragile biological products like MV presents significant challenges.
- Previous work identified a 300 kDa molecular weight cut-off as optimal for MV concentration.
Purpose of the Study:
- To evaluate continuous versus discontinuous diafiltration for MV purification.
- To assess the impact of different media compositions (serum-containing vs. serum-free) on purification.
- To determine the influence of high and low protein loads on the purification process.
Main Methods:
- Ultrafiltration and diafiltration techniques were employed for MV purification.
- Both continuous and discontinuous diafiltration modes were tested.
- MV was prepared in both serum-containing and serum-free media to simulate different protein loads.
Main Results:
- A pre-concentration step significantly improved process economy and MV yield, regardless of the medium used.
- Discontinuous diafiltration showed a slight advantage in permeate flow rate, attributed to dilution and fouling mitigation.
- The combined ultrafiltration/diafiltration process achieved a concentration factor of 8.
- Approximately 50% of infectious MV particles were recovered using 5 diavolumes of buffer.
Conclusions:
- The integrated ultrafiltration/diafiltration process is a viable method for purifying oncolytic measles virus.
- Process optimization, including a concentration step and discontinuous diafiltration, enhances yield and efficiency.
- This purification strategy is suitable for fragile oncolytic viruses like MV, supporting their therapeutic development.
