The future of biopharmaceutical manufacturing platforms is to implement the use of small-scale disposable single-use (SU) plastic wave-reactor bags in parallel to the commonly used 10,000 L stainless steel bioreactors. This will see the industry undergo a facelift with both large multi-billion dollar facilities and smaller, more flexible, sites being constructed with the ability to produce numerous therapies annually without reduced contamination or regulatory issues. It has been estimated that up to 40% in capital cost reduction is possible for facilities adopting disposable technology.

However, the replacement of stainless steel with polyethylene-based plastics brings with it a cohort of new materials into the bioprocessing pipeline. These materials are uncharacterised and have been observed to leach breakdown products into the contact media which raises numerous safety concerns for the patient. The mammalian cell engineering group in the National Institute for Cellular Biotechnology works in collaboration with the National Institute for Bioprocessing Research and Training (NIBRT), Trinity College Dublin (TCD) and the Synthesis and Solid-State Pharmaceutical Cluster (SSPC) on identifying the effects of leachable compounds from disposable bioreactors that threaten the efficiency of the production of biopharmaceuticals.

The Chinese Hamster Ovary (CHO) cell has been the pre-eminent mammalian cell line used for the production of biotherapeutic agents within the biopharmaceutical industry for the past three decades. This cell type dominates as it produces high-quality complex protein therapeutics with human-like post-translational modifications which ensure effective drug therapies with a high degree of safety. Legacy products such as Herceptin and Enbrel for the treatment of breast cancer and rheumatoid arthritis are some examples of life-saving therapies produced in CHO cells. There is constant pressure on the industry for innovation in research and development to come up with ways to further enhance the production capacity of CHO cells in order to meet global demand. With g/L yields being common place these days, CHO cells (like brewing beer) are grown in vast multi-thousand litre stainless steel bioreactors as a means to produce enough material for the clinic. However, with the increasing incidence of disease such as cancer, outputs within the industry is not always sufficient to meet this demand.

Pfizer Grange Castle is the largest single investment the company has made in Ireland at over €1.8 billion in the area of biotechnology and drug manufacturing. Costs associated with capital investment and the cost per production run all contribute to the significant cost of these life-saving therapies, some of which the health services cannot support. It is not surprising that with figures like these, the biopharmaceutical market is set to be valued in excess of $386.7 billion by 2019 and contributes €65 billion in exports to the Irish economy ¹.

Here in the National Institute for Cellular Biotechnology, we are involved in extensive research in collaboration with industrial partners such as Biogen Idec and Eli Lilly to genetically engineer CHO cells using a variety of methods such as microRNAs and CRISPR-Cas9 as a means to enhance bioprocess phenotypes such as growth and longevity which will increase production yields. One such engineering success we’ve had is in the form of DECOY-7 technology which is a miRNA sponge construct which depletes the miRNA, miR-7, and enhances maximal cell density ultimately doubling product yield

Running in parallel to innovation in cell line development and genetic engineering of CHO, are also advances in process development and in the material components required throughout the production pipeline. By reducing costs associated with manufacturing, profits can be boosted while drug costs being reduced indirectly. This has stimulated the interest in the industry for the widespread implementation of disposable technology.

Single-Use disposable bioreactors

Given the huge capital costs associated with setting up a new recombinant drug manufacturing facility, such as the announcement of Bristol-Meyer Squibb breaking ground in Dublin at a cost of €750 million, there is a drive in the industry to develop and implement the use of disposable single-use bioreactors in place of the current stainless steel bioreactors. These polyethylene-based films have numerous advantages over their stainless steel counterparts including reduced capital investment, higher turnover in relation to the elimination of cleaning and sterilisation required for the reusable stainless steel reactors between production runs, increased flexibility within single manufacturing sites and environmental benefits. However, despite the attractive aspects listed, very few manufactures actually implement this technology. This is due in part to cytotoxic compounds to leach from the plastic materials into the cell culture media and ending up in drug product thereby putting the patient at risk. A recent infographic published by Thermo Fisher Scientific summarises the key issues around disposable technology.

After this observation, all material components that come into contact with the drug product must receive approval for the drug regulatory body for safety before market distribution. Supported by Science Foundation Ireland (SFI), our research group in the NICB in collaboration with the Synthesis and Solid-State Pharmaceutical Cluster (SSPC) in the University of Limerick are focusing on the effects of these leachable compounds on the CHO cell itself thereby threatening the efficiency of the bioprocess. One such example of a leachable is the breakdown product bis(2,4-di-tert-butylphenyl)-phosphate or bDtBPP, which breaks down from the compound Irgafos 168 found in polyethylene-based films. Irgafos 168 is a secondary antioxidant critical for the manufacturing process of these disposable plastic bioreactor bags. In addition to our academic collaborators in NIBRT and TCD, we work closely with our 7 industrial partners (Allergan Pharmaceuticals Ireland, BioMarin Manufacturing Ireland Ltd., Eli Lilly and Company, Genzyme Ireland Ltd. (Sanofi), Janssen Biologics, Merck Sharp & Dohme (MSD) and Pfizer Ireland Pharmaceuticals) to explore the potential risks associated with disposable bioreactors.

We have identified that this leachable compound bDtBPP is detrimental to CHO cell growth at concentrations that have previously been observed to accumulate in culture media incubated in contact with these plastic materials (see below).

What this means is, if disposable technology was to replace stainless steel bioreactors, this could be to the detriment of process efficiency ultimately making less therapeutic drug per production run. By trying to save costs, leachable compounds such as bDtBPP threaten to make drug production more expensive which would have a direct impact on treatment costs. Projects like ours in the NICB strive to settle any safety concerns surrounding single-use technology before its full adoption by the biopharmaceutical industry.

Our work has been accepted for publication in the journal Biotechnology Progress. Assessment of the risks associated with leachables from disposable technologies will facilitate in changing the face of the biopharmaceutical industry and usher in the Facility of the Future.

References

1. Ireland Pharmaceuticals and Healthcare Report BMI Q3 2015, Published May 2015, ISSN 2052-5079, www.bmiresearch.com.