Orthogonal technologies for viral safety

Many biopharmaceutical therapies, such as monoclonal antibodies and blood plasma factors, are produced using biological materials. This carries the risk that microorganisms may be unintentionally introduced into the manufacturing process and contaminate the product; these include (among others) bacteria, mycobacteria, transmissible spongiform encephalopathy (TSE) agents and viruses, called adventitious agents. In addition to extensive testing during the manufacturing process to demonstrate the absence of adventitious agents, it has to be proven that the manufacturing process is capable of removing them.

Introduction of multiple complementary and validated viral clearance steps in the manufacturing process, which inactivate and/or remove potential viral contaminants, are of paramount importance in developing new processes for biopharmaceutical drug manufacturing.

An orthogonal approach to viral safety in bioprocessing involves different but complementary techniques, which work together to reduce the risk of virus contamination reaching the final drug product. This may include retentive processes such as virus filtration, adsorptive processes such as chromatography, and inactivation processes under certain conditions such as low pH; that destroy the virus particles. Regulatory bodies like the European Medicines Agency (EMA) and Food and Drug Administration (FDA) require that a minimum of two orthogonal steps for viral clearance need to be employed in the manufacture of therapeutic proteins derived from biological materials. Cytiva can provide technology for three orthogonal step options in either batch or continuous processing mode.

Integrated solutions

With over 20 years of experience in providing automated systems for viral clearance, we can provide the know-how and engineering excellence for an automated system for a single orthogonal viral clearance step, an optimized downstream processing train, or a complete end-to-end solution. Whether it would be a traditional stainless-steel approach, a single-use approach, continuous or hybrid, our integrated solutions can help you find the best choice for viral clearance in your bioprocessing journey.

Virus filtration: robust filtration solutions for the removal of viruses from biotech and plasma processes

Virus filtration uses a membrane barrier to retain virus particles. It is a size-based removal method that uses a specifically designed polymeric membrane to retain virus particles on the surface and within the pores of the membrane. The critical performance parameters are typically the log reduction value (LRV) and the throughput. These parameters may be affected by many interacting factors such as viral load, protein concentration, presence of foulants, pressure, operating flux, ionic strength, and stop start interruptions. Selecting and validating a filter for viruses appropriate for the expected range of conditions, is therefore, very important.

We have developed simple to use and robust products for different challenges. For the requirements of plasma processing, Pegasus™ SV4 and Ultipor™ VF Grade DV20 filters (Pall™ Life Sciences products) have the best fit with the typical process design space; whereas for the demands of mAb processing, Pegasus Prime filters in combination with Pegasus Protect filters (Pall™ Life Sciences products) provide the most suitable solution over a wide range of conditions. Both options are available in either cartridge format for integration into traditional stainless steel processing lines, or capsule format, which may be integrated with pre-sterilized single-use manifolds.

  • Pegasus™ Prime: for mAb and recombinant processing
  • Pegasus™ Protect: for mAb and recombinant processing (prefilter)
  • Pegasus™ SV4: for plasma processing, and high-fouling mAbs and recombinants
  • Ultipor® VF Grade DV20: for various legacy and licensed processes

Virus filtration for monoclonal antibody processes

Pegasus Prime filters are innovative virus removal filters that combine high flow and robust capacity. These filters can simplify process development and manufacturing processes around virus safety and will deliver sustainable economy at all scales of monoclonal antibody manufacturing.

Advice and support for validating Pegasus Prime virus filters

Commonly asked questions include:

Q: What virus spike should I use?
A: The basis for successful spike selection is:

  • Use the purest spike available (ultracentrifugation essential)
  • Use the most sensitive assay technique (large-volume assays)
  • Spike only what is needed to measure your target log reduction value (LRV)

To learn more, download the application note “Validating Pegasus™ Prime Virus Membrane Filters: What Virus Spike Should I Use?”.

Q: How do I incorporate a prefilter in my virus clearance study?
A: One of the key complications for validation of virus filters is the use of an adsorptive prefilter that cannot be validated as part of a robust size-removal virus filtration step. The decision tree in our application note “Validating Pegasus™ Prime Virus Removal Membrane Filters: How Do I Incorporate a Prefilter in My Virus Clearance Study?” will guide you to the most appropriate test method.

Virus filtration in continuous bioprocessing

Continuous bioprocessing is rapidly gaining momentum in monoclonal antibody bioprocessing, providing potential advantages such as smaller facility footprints, lower investment costs, flexibility, and process economy. It is important to note that continuous downstream processing relies on the same fundamental concepts as batch processing. When viral filters are used in a continuous processing, operating conditions are likely to be quite different to batch mode, with much lower flow, and longer processing times in continuous processing compared to batch.

Virus filtration for plasma processes

Pegasus SV4 virus removal filters combine high viral clearance and process cost control. They demonstrate highly efficient clearance of both small "non-enveloped" and large viruses, and they offer high retention robustness to pressure interruption (also described as pressure release, or start/stop).

The Pegasus SV4 filter membrane ensures a constant and stable flow rate when used with either dilute or complex/concentrated biological fluids, which significantly improves viral filtration economy in plasma fractionation processing.

Virus inactivation: economic solutions for virus inactivation, whatever your process requirements

The inactivation of virus contaminants can be achieved by subjecting the bioprocess fluid to conditions that denature the virus protein but not the active ingredient. In the production of biologic therapies, the two most employed inactivation methods are the use of low pH or addition of detergents. The efficiency of these processes is clearly linked to the effectiveness of mixing, to bring the virus quickly into intimate contact with the denaturant, and the hold time at these conditions. Deactivation with detergents is more common in blood plasma processing, whereas low pH inactivation is more frequently applied to monoclonal antibody processing. In the latter case, a low pH hold of 1 hour at a pH of 3.5 is typically applied and is effective against enveloped viruses.

We have developed automated system technology that enables the viral inactivation at low pH to be carefully controlled in either a single-use batch or a single-use continuous mode. The Allegro™ MVP single-use system can be used in combination with our single-use mixing technology over a range of batch volumes to provide an automated solution for acid and base dosing, ensuring pH control that will inactivate viruses while avoiding denaturing conditions for the monoclonal antibody.

Virus inactivation in continuous bioprocessing

For small batch or continuous bioprocessing, we have developed the Cadence™ Virus Inactivation System. The system employs two parallel mixers and recirculation loops that work synchronously and alternately so that one set is involved in the low pH viral inactivation step while the other is filling with bioprocess fluid. The system is specifically designed so that it can work with the elution from an upstream continuous capture chromatography step and can also operate in batch mode.

Viral clearance using anion exchange membranes: scalable and ready-to-use solutions for impurity removal or capture of large molecules

Under appropriate conditions of ionic strength, anion exchange (AEX) sorbents or membranes can be used in negative or flow-through mode. This is done to remove virus contamination, which binds to the surface of the membrane while the therapeutic molecule of interest, such as a monoclonal antibody, passes through the device. The plug and play nature of membrane chromatography devices makes them particularly convenient for this type of polishing step.

Typically employed at the end of a sequence of chromatography steps, anion exchange membrane capsules can also help reduce levels of host cell protein (HCP) and DNA contamination as well as provide an orthogonal approach to viral clearance. Mustang™ Q capsules can be especially effective in this regard and have been employed successfully as part of an orthogonal strategy for virus safety in several industrial processes.

Our validation and virus filtration specialists can provide support and advice from process development through to commercialization, for the lifetime of your drug product — contact us today.

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