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Team 3 (Qai, Sin En, Kennice, Reuben & Pei Xin)

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Team 3 (Qai, Sin En, Kennice, Reuben & Pei Xin)

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Which is better?

Disposable or Fixed-in-place stainless steel bioreactor?

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Cultivation container is plastic (polyethylene)

must be approved by the Food and Drug Administration (FDA)

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1)wave bioreactor(rocking)

-rocker platform uses the wave motion to liquid inside and this will allow the cells to get accessed to the nutrients needed to grow

2)stirred bioreactor

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1) Speed and ease of deployment

2) No cleaning

3) Increase plant flexibility

4) Contamination is exceedingly rare

5) Fast changeover resulting in greater batches/facility/yield

6) Reduced capital expenditure for equipment and facility

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Advantages & Disadvantages  of pre-sterilized units  (Kennice)

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Advantages & Disadvantages  of pre-sterilized units  (Kennice)

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why use a disposable bioreactor (Sin En)

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why use a disposable bioreactor (Sin En)

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-greatly reduce the additional cost and time of sterilization

-featuring lesser parts in a fixed in place bioreactor hence it is less expensive to maintain on the mechanical end

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available in large capacities

more advanced measurement & control

less dependence on supplier

lower leakage risk during culture

customizable designs

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1) Capacity limit - not for large-scale manufacturing

2) Require ongoing consumable usage forecast

3) Quality by design - consider extractable and leachable study data

4) Relatively new technology

5) More responsibility for operators

6) Damage to single-use bags may cause leakage

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requires CIP/SIP

higher probability for contamination to occur

high maintenance cost

slow turnover rate

water & chemical discharge waste

inflexible infrastructure

cumbersome routine cleaning

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https://solidabiotech.com/product/compact-bioreactor-2/

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Factors like: Scale up, Time, Amount of products, etc, can determine which would be better

Using Disposable bioreactors can help reduce the risk of cross-contamination and can help save time (shorter lead times). However, due to different shapes, agitation principles, and gassing strategies, scale-up and scale-down can be challenging.

With geometrical properties and long experience in process engineering knowledge, fixed-in-place bioreactors are still better than disposable bioreactors.

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-heat transfer jackets and internal coils provide aggressive heating and cooling capability

-pressure and full vacuum capability

-operate as closed system that requires little or no external environment

-an agitator system”, “A foam control system”, “Baffles system”, “PH & temperature control system”, “Fermenter vessel”, “Aeration system” and “An Impeller system”

-sparger, impeller

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https://biopharma-asia.com/products/predictable-scaling-in-single-use-bioreactor-format-ge-healthcare-life-sciences/

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1) No pump or impeller needed to induce circulation flow

2) Inoculum, media, and buffers are aseptically transferred into the bag via a sterile tube fuser made of thermoplastics connection, resulting in exceptional strength

3) Significantly reduce the cost of validation and cleaning requirements

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Used for producing viral vectors, vaccines, and other cell-based products expressed by adherent cells

ADVANTAGE

1) Helps simplify downstream purification processes as fixed beds acts as a natural size exclusion filter for turbid biomolecules, reducing the need for further downstream clarification

2) Offers a fully closed, automated system of cell culture (lesser responsibility for operators), and contains a large surface area for cell attachment

3) Low shear force with homogenous microenvironment -> more concentrated harvest / product but less in volume -> simplified downstream process

https://www.biopharminternational.com/view/exploring-the-impact-of-fixed-bed-bioreactors-in-upstream-processing

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Innovations in bioreactor design are prompted by the need to optimize cell culture reactions. Cell culture processes must be robust and scalable, and yet economical. A look at the fixed-bed bioreactor (FBB) can help with understanding their impact on process development and scale up of biologics.

FBB apart from STR

Looking at the history of bioreactors for bioprocessing, the first bioreactors were developed from the chemical industry, broadly speaking, explains Alex Chatel, senior marketing and product manager, Univercells Technologies. These early bioreactors were large, stainless-steel stirred-tank (STR) bioreactors designed for industrial production of biologics.

"While they offer size and scale, they lack flexibility and require extensive cleaning regimes (and associated quality assurance and control activities). To accommodate more flexibility, reduce operational and capital expenditures, and de-risk commercialization, novel single-use STRs were developed," Chatel says.

The single-use STRs are disposed of after each batch, removing the need for cleaning regimes, which in turn reduces the need for hard piping, cleaning utilities, and other associated personnel. Chatel notes that, while an improvement, the STR design itself is by nature not linearly scalable and not well suited to handling delicate cells or processes, especially at scale. This limitation leads to complex and time-consuming scale-up studies, he states.

In STRs, Chatel explains, cells are suspended in a liquid media and are constantly mixed by an impeller. In single-use FBBs, however, cells are "entrapped" in a solid but porous matrix, and cells are at a much higher density. As opposed to STRs, the cells don't move around the bioreactor in the latter, but, rather, an impeller circulates media containing nutrients throughout the fixed-bed matrix.

"Biomass immobilization has several advantages, such as providing a low-shear environment for cell growth and sensitive products and removing the need for cell retention devices for perfusion processes," Chatel states. He goes on to explain that first-generation FBBs are made up of randomly packed bed matrix. This construct can be problematic because it provides an inhomogeneous environment for cell anchorage and growth, which leads to variable productivity and product quality, according to Chatel. Newer FBBs, however, provide a structured fixed bed with a homogenous three-dimensional environment, ensuring homogeneous cell distribution and growth. These newer FBBs are equally suitable for adherent and suspension cells both in serum and serum-free conditions.

Finally, structured FBBs are scalable by design and offer higher throughputs than conventional STRs, an advantage that enables cost-efficient production. "In one study conducted by Univercells Technologies (to be published), producing an AAV-2 [adeno-associated virus-2] in [an FBB] (scale-X nitro 600, Univercells Technologies, which has a 600-m growth surface and 60 L working volume) equates in productivity to a STR capacity of 3600 L and results in a cost of goods (CoG) reduction of 24% compared to the STR process," Chatel says.

Andrew Laskowski, global product manager, iCELLis Bioreactors, Pall, reinforces the notion that FBBs are compact upstream manufacturing technologies that have been optimized for the cultivation of adherent cells. The primary feature of these bioreactors is an immobilized surface where cells adhere and proliferate. FBBs are primarily used to manufacture viral vectors or viral vaccines, remarks Laskowski.

"These products can also be manufactured in suspension cells using a conventional stainless-steel or single-use stirred bioreactor, but suspension processes require additional development time to adapt suspension cells lines, and product titer is generally lower than adherent cell processes. FBB therefore enable companies to bring products to clinic quicker and with a more efficient processes," Laskowski observes.

Going with the workflow

As FBBs are mostly used in the production of viral vectors, viral vaccines, and other cell-based products expressed by adherent cells, these bioreactors streamline large-scale workflows by facilitating easy sterile media addition, cell addition, and infection/transfection, notes Laskowski. "Cells are immobilized on the surface of the fixed bed, enabling FBB to act as natural cell retention devices, facilitating the easy implementation of perfusion," he states.

Laskowski also points out that FBBs help simplify downstream purification processes, as the fixed bed acts as a natural size exclusion filter for turbid biomolecules, reducing the need for further downstream clarification.

Chatel adds that some structured FBBs (e.g., the scale-X family of bioreactors, Univercells Technologies) also enable cell harvest, meaning they can be used to generate the inoculum of larger-scale systems. "They thus play a central role in biomanufacturing processes, as the choice of upstream technology significantly impacts the rest of the manufacturing process," he states.

FBBs convey specific advantages to the overall biomanufacturing process. For example, they offer a fully closed, automated system of cell culture and contain a large surface area for cell attachment in a small footprint, Laskowski explains. Furthermore, FBBs are integrated with analytical technologies, such as pH sensors, dissolved oxygen sensors, temperature sensors, and (in the case of the iCELLis bioreactor, Pall) a biomass sensor to measure cell density.

"Compared to conventional 2D flatware manufacturing methods, these properties of fixed-bed bioreactors result in a decrease in labor and operational costs, process footprint and facility utilization reduction, decreased risk of contamination due to open handling, and an increase in process assurance, consistency, and robustness," Laskowski asserts.

Structured FBBs are also highly intensified, meaning that, for an equivalent production throughput, the working volume is much smaller than in STRs, says Chatel. This result is achieved through drastically increased volumetric cell concentrations thanks to the immobilization of the cell mass within the fixed bed, he explains.

Furthermore, the cell growth support matrix can act as a filter, helping pre-clarify the harvest material from cell debris and other process impurities. Most importantly, however, FBBs provide a low-shear, homogenous microenvironment for cell growth, which typically leads to higher productivities (two- to four-fold increases have been observed) (1). "The resulting harvest is therefore not only more concentrated in product and of higher quality, but of greatly reduced volume compared to an equivalent STR process, resulting in a simplified downstream process," says Chatel.

Tackling scale up

Scale-up of the biomanufacturing process typically takes places in steps, with key process parameters and product quality attributes confirmed at increasingly large scale, Chatel explains. "For processes based on STR, the change in volume between the laboratory and the final scale can be 100 to 1000-fold. The scale-up activity is often empirical, time-consuming and can result in trade-offs between reaching the final scale and meeting target productivity, for instance," he states.

When scaling up using FBBs, however, the change in volume does not exceed tens of liters. "The inherently scalable bioreactor design also significantly simplifies the process, thus accelerating and de-risking scale-up," Chatel adds.

Meanwhile, Laskowski points out that FBBs are the most scalable technology currently available to cultivate adherent cells. Unlike 2D flatware, which scales out rather than scales up, he explains, FBBs are available in many different sizes (e.g., iCELLis bioreactor, Pall, available in sizes from 0.5 m 2 to 500 m ²)."The performance at these different scales is maintained by increasing the diameter of the vessels but conserving the geometry of the fixed bed and the linear speed of cell culture media through the fixed bed. Contrary to 2D flatware, as batch sizes in FBB increase, the number of operators and process footprint remains relatively unchanged, providing FBB manufacturers with the advantage of economies of scale," Laskowski states.

On the process development front, there is some influence from FBBs. As Laskowski notes, FBBs require a bit of additional upfront capital investment to purchase the process development-scale hardware and automation systems. He estimates the investment cost to be in the ballpark of approximately $100,000. After the initial capital investment, the cost in raw materials per batch is comparable to flatware at a similar scale, he says.

Yet, there are significant advantages to using FBBs for process development. Laskowski points out that it is possible to use the provided analytical tools to better optimize the cultivation conditions for cell growth and production. "The bioreactors are fully automated," he states, "freeing up scientists to work on other projects."

Finally, the process optimized in an FBB can be more easily scaled up to manufacture clinical or commercial batches, allowing FBB users to achieve important milestones more quickly, Laskowski adds. "For example, as they disclosed in a presentation at ASGCT in 2019, Novartis Gene Therapies used the iCELLis bioreactor to commercialize their gene therapy product, Zolgensma, in just under 38 months," Laskowski says.

Meanwhile, Chatel explains that cells adapt well to the protective environment in a structured FBB, which means that initial process transfers from smaller-scale laboratory systems or other bioreactors is simple and effective with equivalent performance and without the extra step of conducting process optimization. Thus, the transfer of a benchtop process to clinical and commercial production is simplified, he emphasizes. He points to one study (to be published) in which a process using a suspension human embryonic kidney 293 (HEK293) cell line producing an adenovirus product in a 2-L STR was transferred to a bench-top system (scale-X hydro, 2.4 m, Univercells Technologies). This process was scaled-up to commercial production (scale-X nitro, 200 m, Univercells Technologies) in less than nine months while increasing product titers.

Thus, FBBs, in effect, do double duty in helping to optimize overall bioproduction. They convey volumetric and process development advantages in the upstream, while also producing targeted product that undergoes an initial filtering process prior to cell harvest. The impact on downstream processing can thus be seen early in the manufacturing life cycle.

Reference

1. Univercells Technologies, scale-X Bioreactor Demonstrates Superior Productivity for Vaccines and Gene Therapy. Press Release, Jan. 27, 2022.

About the author

Feliza Mirasol is the science editor for BioPharm International.

Article details

BioPharm International
Vol. 36, No. 4
April 2023
Pages: 18-21

Citation

When referring to this article, please cite it as Mirasol, F. Exploring the Impact of Fixed-Bed Bioreactors in Upstream Processing. BioPharm International 2023, 36 (4), 18-21.

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Exploring the Impact of Fixed-Bed Bioreactors in Upstream Processing

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The innovation of going from stainless-steel bioreactors to fixed-bed bioreactors shows an evolution in upstream optimization.

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Parts of a disposable bioreactor, stirrer type (peixin)

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Parts of a disposable bioreactor, stirrer type (peixin)

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1. impeller/agitation system

to ensure culture media and nutrients are evenly mixed

keep cells in homogenous condition

2. exhaust

release of carbon dioxide from the respiration of cells and reduces pressure in the bag

3. air in

to provide/pump in oxygen for cell respiration leading to higher survival of growth cells

4. feeding pump

provide media which contains nutrients for cells to feed on and increase the growth of cells

5. sensors probes

detect the change in pH

if pH increase, acid will be pump in to reduce pH value to neutral

if pH decrease, alkaline will be pump in to increase pH value to neutral

6. reactor tank

a tank that contains culture media that needs to be seeded and cell growth

7. thermal jacket

allows adequate heat transfer to maintain particular temperature in bioreactor

prokaryotes --> 25oC

eukaryotes --> 37oC

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For fixed-in-place bioreactor, it can be used in large scale cultivation of cells but not small scales. Fixed in place bioreactor is mostly found in biopharma industry that cultivates large scale cells. By having large scale bioreactor, it has its disadvantage as it can have cross-contamination and hard to track the origin of contamination. But it also has advantages which is environmentally friendly as it reduces the usage of disposable bioreactor and saves cost.

For disposable bioreactor, it can be used in small scale cultivation like research in lab which does not require many samples. By having small scale bioreactor, it has its advantage which reduces cross-contamination and easy identification of cross-contamination origin. But it also has it disadvantages where it is not environmentally friendly and cannot get large scale results as time consuming to kept culturing cell lines in different disposable bioreactors.

In conclusion, though both bioreactors have its advantages and disadvantages, but the use of different bioreactors applies in different context like in lab research we only need a small-scale cell culture media thus reducing the wastage. In biopharma industry that grows cell in large batches, large scale bioreactor is thus more preferred.

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Feeding strategies for Bioreactors (reuben)

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Feeding strategies for Bioreactors (reuben)

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1) Batch culture

2) Fed-batch culture

3) Continuous culture

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Charging in fresh cell-free medium into the bioreactor

cells are added to a fixed amount of nutrients

nutrients provided only at initiation process

no addition of fresh medium in between

working volume remains constant

vessel is filled with media and fermentation is started by cells inoculation

gas is continuously bubbled into the culture

acid and alkali are added as required

cells, spent medium and products are harvested at the end

processing time

microbial 12-24h

animal cells 5-7 days

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vessel partiall filled with media and fermentation is started by cell inoculation

fresh media added due to consumption of growth limiting nutrients

no withdrawal of cells or medium during cultivation period

critical elements of nutrient solution are added in small concentrations at the beginning of the fermentation

Processing time

microbial 24-36h

animal cells 7-21 days

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fresh medium is continuously fed into the bioreactor

maintains feed flow rate at suitable value to avoid washing out of cells

continuous withdrawal of cell suspension from bioreactor in continuously stirred tank reactor which offers a low cell density at a steady rate