Speeding up production in small, dynamic biopharma facilities

In the biopharma industry, rapid product roll-out is critical, as the demand grows for monoclonal antibodies (mAbs), bacteria-produced vaccines and therapeutic mammalian cells. The yearly output of biopharma protein products amounts to more than 17 million liters worldwide. Innovative solutions like the GEA kytero single-use perfusion separators, designed for highly intensified, continuous cell separation, answer the needs of producers looking to capitalize on this dynamic market and grow their success with robust and easily scalable upstream processes. The new perfusion separators offer a transformative approach to multiply product output, achieve unprecedented productivity and serve medical institutions and patients more quickly. 

Continuous means cost-efficient

Continuous processing optimizes production time and reduces the effort involved in starting biopharma process runs. Operators no longer need to wait for the end of a batch run to separate the cells and collect the target protein – the active pharmaceutical ingredient – from the product-containing medium. Instead of discarding the cells, they return them to the bioreactor, allowing the production to run for weeks – rather than hours or days – and yielding product from day one. 

This innovation enables new products to reach the market faster and established product to be delivered without losing valuable time. Importantly, the same cultured cells are reused over the entire multi-week process run and do not have to be freshly cultivated for successive batches. 

Continuous means cost-efficient, and in the past only larger operations have been able to benefit from this approach. That is about to change with the new single-use perfusion separators, which make this technology available for smaller pharma companies characterized by limited floorspace, few labor resources, complex workflow demands and fierce competition. These companies tend to depend on modern single-use equipment for their key processing tasks. Elaborate cleaning cycles that take up additional space, utilities, workforce and time would be overwhelming for them. In 2021, single-use separators marked the first time worldwide that easily applicable disk stack separation technology for batch processing became accessible to smaller R&D or production outfits, offering high efficiency, superior flexibility and uncomplicated operation. The new single-use perfusion separators in the series – the first of their kind – achieve the same for small facilities with their eyes set on continuous production.

Existing biopharma processes 

Culturing cells in the bioreactor and subsequently harvesting the product protein is the key upstream process in biopharma production. Following exponential cell growth in the reactor, the cells need to be taken from the vessel and separated from the culture medium – a process that perfusion processing can optimize. However, making the switch to a perfusion setup has in the past appeared forbiddingly expensive and risk prone. This is a main reason why many pharma manufacturers – even larger facilities – have so far resisted. Single-use perfusion units present an opportunity to enter perfusion processing with comparatively little effort and cost, removing many of the typical entry barriers for this process. A survey of the typical processing options gives the following picture:

Preparing the inoculum (the startup cells)

No matter which process setup is used for cell cultivation and separation, the preparation is the same: Cells from a cryopreserved stock vial are pre-cultivated in shake flasks placed in an incubator shaker. Passaging (removing the medium and transferring cells into fresh growth medium) can be done at standardized intervals (e.g., every other day) to boost propagation of the cell strain before inoculation. 

Option 1: Batch processing

For R&D purposes a simple batch process can be the most efficient method to culture and separate the cells. The cell culture can be active for less than seven days, or even just a few hours, and is then fed into a small separator or other cell retention unit to separate the cells and isolate the desired product. The key advantage of this procedure is that it is easy to set up and easy to protect from contamination. 

Option 2: Fed-batch processing

Batch-based cell cultivation processes can only last for a limited time before the nutrients in the substrate medium are spent and toxic byproducts of the cell metabolism exceed a prescribed limit, affecting the viability of the cells. With a fed-batch process, this time limit can be extended to more than 15 days. This is done by repeatedly replacing the medium in the bioreactor with fresh substrate from a separate tank as prescribed by complex schedules. This procedure removes toxic particles and adds further nutrients sufficiently to keep the batch active, but the feed volume must be carefully balanced without diluting the culture. After the entire batch run, the product is harvested in the same way as in the standard batch process. Fed-batch processing, while demanding, is suitable for commercial-capacity production. It delivers high-quality results with sufficient cell density and product titer.

Option 3: Perfusion processing

Using a perfusion setup with a solution like the new GEA kytero perfusion separator allows for cell culturing operations to be extended to weeks. The medium in the bioreactor, containing cells, cell products and nutrients, is continuously fed into the perfusion separator, instantly yielding valuable product. Spent medium is collected with the harvested product at the same rate that fresh medium is being added to the reactor, keeping the culture medium at a constant volume in the bioreactor. The exchange rate is called vessel volume per day. The separated cells are returned from the separator unit, equipped with a special port, to the bioreactor, at no loss of viability. 

Facilitating perfusion: High-efficient disk stack separation 

Several cell retention methods are available to provide the required separation of the cells, including alternating tangential flow filtration or tangential flow filtration, which separates particles according to size, while disk stack separation employs centrifugal force to separate particles according to mass. 

The latter technology, refined by GEA separation engineers over many decades and used in industrial pharma operations around the world, offers advantages in terms of efficiency, flexibility and space. Disk stack separation delivers exceptionally high cell density and product yield with 99 percent solids capture. It can be fine-tuned to small differences in mass and thus accommodate intricate automated separation tasks that can be changed on a day-to-day basis. Friction-free and low-shear mechanical concepts ensure minimal temperature increase for maximum cell viability.

Groundbreaking single-use convenience

In the GEA kytero series the company’s proven disk stack technology has been transformed into particularly compact machines with an easy-to-handle single-use element that contains the entire disk stack technology. This element and all corresponding feed and discharge lines – that is, all parts in contact with the product – are simply exchanged after a production run for maximum convenience and safety from contamination. Gamma-treated exchange cartridges are optionally available. 

The newly introduced perfusion separators in this series offer the same advantages as the original fed-batch separation models: No hygienic cleaning or autoclaving is required, so the machine can be ready for the next continuous process run in a matter of minutes. The separators do not require any utilities except power and air supply. They fit into every facility, especially in comparison with classic filtration systems. As with all disk-stack separators, no floorspace or labor resources must be reserved for changing filters.
 
In addition to the innovation of a compact, closed, 100-percent contamination-proof separating unit system, the GEA kytero machines feature a no-contact drive system, providing ideal support to the protection of biocontainment. 

Starting up and scaling up with perfusion

GEA kytero perfusion separators are available in two variants for up to 500-liter and 2,000-liter bioreactors and thus enable process treatment with economically reduced quantities of expensive cell strains. A new separator for the smallest R&D lab applications, suitable for both batch and perfusion test processes, is expected to be introduced in early 2025. 

Test users of GEA kytero machines have found they support overcoming current limitations. The test and production results achieved with these process components can be scaled up to larger single-use model variants and even to industrial production with classic stainless-steel separators in the renowned GEA pharma aseptic and GEA pharma pure series – always maintaining the technical principle of disk stack separation, which ensures a safe process transfer and facilitates validation.

Biopharma manufacturers aiming to establish perfusion processes and expand their position in the dynamic pharma market are encouraged to explore the benefits of single-use perfusion technology in their operations.