Considerations in evaluating new plastic film for single-use technologies and film development

The increase in adoption of single-use bioprocess technology for manufacturing biopharmaceuticals has been driven by the many advantages it offers, such as increased flexibility. However, the materials of construction can be more complex than conventional stainless steel vessels. Single-use containers are constructed from plastic films, which are often composed of several layers of polymers with additives for processing and performance.

The diversity of applications for single-use containers requires film that can achieve a wide variety of performance attributes such as mechanical strength, flexibility, biocompatibility, and suitable gas barrier properties to name but a few. The right balance of chemical composition and film architecture is critical for achieving desired performance across many applications. Also, in light of industry reports of a leachate, originating from bioprocess containers, that can negatively impact cell culture growth with certain sensitive cell lines, it has become imperative to understand the profile of extractable compounds resulting from the film chemistry. Over the past several years, GE Healthcare has worked closely with biopharmaceutical manufacturers to understand their applications and resulting requirements for bioprocess film performance. This investment in learning has led to the establishment of a scientifically driven film development process focused on the innovation of bioprocess films that are fit for purpose. The process is built on a strong foundation of knowledge in science, engineering, and bioprocess applications and incorporates key learnings about extractables and  leachables, as well as considerations for security of supply via supply chain management.

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Question 1

How has GE applied its expertise in film development?

A good example of this is GE's most recent film innovation, Bioclear 11. It is a low-antioxidant film developed as a response to the need in the industry for improved cell growth performance with certain sensitive CHO cell lines. The film is used in Cellbag™ bioreactors and M*bag mixing chambers from GE Healthcare's Life Sciences business. Bioclear 11 was purposefully designed to have all the physical and mechanical attributes essential for WAVE™ bioreactor applications. Through extensive testing, Bioclear 11 has been proven to meet all the same requirements as Bioclear 10 film, but consistently outperforms Bioclear 10 in its ability to support TBPP-sensitive CHO cells. The innovation of Bioclear 11 is an example of how intelligent design, strong relationships with customers and suppliers, as well as increased material controls can be combined to provide solutions for unmet industry needs.

Question 2

Does the lower antioxidant level affect the shelf-life of Bioclear 11?

A shelf-life study was conducted with Bioclear 11 Cellbag bioreactors. They were found to have the same 2-year shelf-life as Bioclear 10 Cellbag bioreactors.

Question 3

What cell-line was used to evaluate cell culture performance with Bioclear™ 11 and how sensitive is it to the degradation product of TBPP?

The improved cell growth performance for Bioclear 11 Cellbag bioreactors was verified by using a CHO DG44 cell line. It showed sensitivity to the bDtBPP degradation product of TBPP at levels as low as down to 0.1 mg/L (100 ppb).

Question 4

What are the important factors to consider when evaluating film for single-use applications?

Defining the critical to quality attributes needed in single-use applications is the first important consideration. Once those are established, an in depth understanding of the materials of construction is needed in order to be able to make the right selection of materials. This includes knowledge about the physical and mechanical attributes, as well as the chemical structures and degradation profiles.

Question 5

What about leak resistance? As you have made changes to reduce leachables how has this impacted the strength of the bags?

In order to ensure a film will have optimal strength for bioprocess applications, the polymer resins and additives must be carefully selected and optimized. A combination of chemistry, biology, and engineering design factors must be considered to achieve the critical to quality attributes of a bioprocess film.

In the case of Bioclear™11, specifically designed for Wave™ Bioreactors, the concentration of the antioxidant tris(2,4-di-tert-butylphenyl)phosphite) was lowered to reduce the level of extractables. However, extensive testing was done to confirm the reduction in antioxidant concentration did not impact the form, fit, or function of the film. The physical properties of the film, such as tensile strength and modulus, are unchanged. For more details about the comparability study of our Bioclear™ films, click the link below.

www.gelifesciences.com/gehcls_images/GELS/Related%20Content/Files/1435051374327/litdoc29144238_20150623112241.pdf

Film selection for bioprocess applications requires consideration of how the design of the film influences the chemical, biological, and engineering performance attributes. At GE Healthcare, through collaboration with our Global Research Centers, we draw upon our strong foundation of material science knowledge and combine it with our understanding of bioprocess application to advance our scientific acumen in film development.

Question 6

In your opinion when is the best time to evaluate extractables and leachables in your manufacturing process?

There is no one size fits all approach for determining the best time to evaluate extractables and leachables. A drug safety profile should be clinical-phase appropriate and based on patient risk.

Common inputs include:

  1. user experience and historical data on the single-use systems,
  2. supplier biocompatibility data,
  3. supplier extractables data, and
  4. user-generated in-process leachables data.

It is important to review of all aspects of the manufacturing process in combination with consideration of the formulation chemistry and the materials of construction of the bioprocess equipment it comes into contact with. The extractables profiles for the single-use technologies used in the manufacturing process provide a useful source of information to aid in the evaluation process. The data for GE Healthcare single-use technologies can be accessed through the following website: www.gelifescisences.com/rsf. The specific pathway a company will take blends many other product and process inputs, and is greatly influenced by the company's perception of risk.

Question 7

Are bag decisions somewhat custom, I mean depending on the sensitivity of the cell line, drug product, etc. do you have recommendations about the make up of the bags that should be used?

GE Healthcare offers a variety of standard single-use assemblies; in addition, users can request custom solutions (film, ports, tubing configurations) designed to meet the needs of their specific process.

Each bioprocess application presents unique requirements and challenges for bioprocess film. The film must be able to meet the critical to quality attributes required for optimal performance in a given bioprocess application. Over the past several years we have developed a good understanding of the film chemistry that affects cell growth, and we have taken substantial steps in both film design and film evaluation – through cell culture, chemical, and physical attributes testing. For example, the Bioclear™ 11 film used in WAVE Cellbag™ bioreactors has optimal flexural properties for the stresses imposed by the rocking motion and wave impact forces, and also has a low antioxidant profile for better compatibility with certain sensitive cell lines. Film selection requires consideration of how the design of the film influences the chemical, biological, and engineering performance characteristics. However, it is still good practice for a user to evaluate the suitability of a film with their specific cell lines and processes.

Question 8

How concerned are you about leachables/extractables in single use connectors and tubing? Do you have plans in these areas?

One should be concerned about extractables and leachables coming from all components of a single-use assembly. As such, we have extractables data on connectors, tubing and all other components. This information can be found at the following website: www.gelifescisences.com/rsf. However, one should also evaluate the potential impacts of extractables and leachables by using a risk based approach that takes into account the interactions between process, fluid stream, and the single-use assembly. Such an assessment often leads to a reduced emphasis on smaller components such as connectors, and a greater focus on the major components, such as film.

Question 9

How has GE developed the capabilities to work with customers?

As the implementation of single-use technology continues to grow and expand into new applications, the industry will be presented with new challenges and requirements. In recent years, GE Healthcare has invested in expanding our internal capabilities for characterizing the materials used in our single-use systems. Through collaboration with our Global Research Centers, we draw upon our strong foundation of material science knowledge and combine it with our understanding of bioprocess application to be able to tackle both current and future challenges with bioprocess films and design the next generation of film technology. A great focus will remain in partnering and collaborating with end-users and experts in order to optimize single use technologies including plastic films.

Question 10

We are a lab scaling up from benchtop glass bioreactors to disposable bags, but are still working at fairly small scale. What kind of film questions should we be asking as we make the transition?

Selecting a single-use assembly for an intended application requires consideration of a number of factors. For example, operating parameters such as temperature, humidity, working volume, and run time are key inputs. In addition, knowledge about the sensitivity of a particular system to the ingress of gases (O2 or CO2), loss of water, or shifts in pH must also be factored into the selection process. Evaluating these inputs in comparison to the performance attributes of a single-use assembly can help determine the suitability of the system for the specific application. Some of the key performance attributes of single-use assemblies include tensile properties, biocompatibility, gas barrier properties, extractables profile, puncture resistance, operating temperature range, chemical compatibility, and film clarity to name but a few. At GE Healthcare, we have advanced our scientific evaluation of films, combining the chemical and physical attribute testing with cell culture growth performance. Our systems have been implemented into many cell culture and fermentation processes. In addition, we partner with customers to understand their applications to help us provide solutions for their performance demands.

Question 11

We are a company just starting to work on risk assessment for extractables and leachables after moving from stainless steel, what do you see are the biggest challenges? Any advice on avoiding potential pitfalls?

Supplier may serve as a good source of information and to orient you to how other drug manufacturers manage the topic. A web or literature search will lead to articles that can help to familiarize you. As well, there are several conferences every year on the topic. To complement the educational aspect, you should then embark on your own risk assessment on the topic. It is important to keep in mind that there are several different versions of risk including: product risk, process risk, patient risk. As well, it is essential to understand your organization's general stance on managing risk. At the extremes, some companies will be very comfortable with technical arguments as to why something is not an issue, whereas others will take the opposite view, leaving nothing to first-principles arguments. Lastly, it is important to include representation from several different functions during the risk assessment/management exercise. When it comes to understanding the extractables / leachables risk associated with single-use technology, some key questions that are top of mind of many users include: the materials of construction of the assembly, the methodology used in the supplier's extractables studies, and the security of supply and change notification process related to the materials of the assembly. Gathering this information is important for avoiding potential pitfalls related to unexpected outcomes with extractables and leachables.

Question 12

What film selection advice do you have for vaccine manufacturing and specifically virus production?

Standard principles related to understanding, managing, and mitigating risk apply to vaccine manufacturing. One might for example contend that virus adsorption to a plastic surface present a risk to be addressed. It might therefore be appropriate to ask the supplier if they have any information to confirm the issue or mitigate the concern as it relates to their specific products. It might also be appropriate to assess the issue experimentally, to move from a supplier's claims related to general applicability of a product in the industry, and to a process-specific qualification of a given piece of equipment. Single-use technologies, (i.e. Xcellerex™ XDR bioreactors, Cellbag™ bioreactors, and liquid handling systems), have been implemented in various applications including vaccine manufacturing. Please contact your GE representative to discuss existing options and any concerns you have regarding different technologies or biosafety levels.

Question 13

How does your film interact with lipids, is there a film recommendation related to lipids?

The interactions between lipids and plastic film depend on the chemical structure of the lipids as well as the materials of construction of the film. If the polymers that make up the film are more hydrophobic in nature, the propensity for lipid adsorption / absorption is higher. It is also important to consider the physicochemical properties of the solution formulation (i.e. polarity, ionic strength, surfactant content etc.) in order to determine if the solution or the plastic film will provide a more favorable environment for the lipids to reside. Lipids are common in cell culture processes and the vast majority of them do not seem to have an issue related to lipid depletion when run in single-use systems. We therefore conclude that there is no systemic issue. That being said, on rare occasions, some users may have had to optimize media formulations in order to counterbalance potential negative effects from lipid depletion. GE Healthcare has the capability to test and optimize your cell culture media and processes on a custom request.

Question 14

Why doesn’t the industry move to standardized testing for single-use films and bags?

Although single-use technology is broadly accepted in pharmaceutical manufacturing, the industry continues to evolve with respect to the qualification requirements for these systems. Industry-wide discussions aimed at aligning on standardized approaches for qualifying single-use technologies have been ongoing for several years. Standard methodologies exist in some areas, such as ASTM methods for tensile properties and the USP testing for Class VI plastics. Progress is being made in several other areas, such as extactables testing protocols and cell culture evaluations, through joint efforts of pharmaceutical manufacturers and single-use suppliers. GE Healthcare is actively participating in many activities with members of BPSA and BPOG in order to drive toward the goal of standardization. As the implementation of single-use technology continues to grow, the industry will be presented with new challenges and requirements. In recent years, GE Healthcare has invested in expanding our internal capabilities for characterizing the materials used in our single-use systems. Through collaboration with our Global Research Centers, we draw upon our strong foundation of material science knowledge and combine it with our understanding of bioprocess application to be able to tackle both current and future challenges with bioprocess films and design the next generation of film technology.

Question 15

In your opinion do you think that there is another bDtBPP type compound that we haven’t identified yet or do you think the key leachables have been identified and are being addressed.

IIt is difficult to make predictions because there are many variable factors to consider, such as differences in the level of sensitivity of cell lines. In addition, the types of compounds present in a single-use assembly that have the potential to become leachables will vary from system to system and are highly dependent upon the materials of construction used in the assembly and the processing conditions they have been exposed to. That being said, it is possible to make some predictions of what compounds may be extracted from a given material based upon the chemical composition. For example, the films used in single-use bioprocess assemblies are often composed of several layers of polymers, such as polyethylene, polyester, nylon, and ethylene vinyl alcohol copolymers. Small molecule additives may be needed for processing and performance. Some common types of additives include antioxidants and slip agents. In addition, residual monomers and chemical degradants resulting from film processing may also remain in the film and have the potential to be extracted. The information provided by supplies about the materials of construction and the extractables profile for a single-use system can be leveraged to assess the potential structure-activity relationship for the chemical compounds and the specific cell line being used. Over the past several years, we at GE Healthcare have advanced our understanding of the film chemistry that affects cell growth and have expanded our supply chain management strategy to allow us to better trace the composition of our films. We have taken substantial steps in our scientific evaluation of films, combining the chemical and physical attribute testing with cell culture growth performance. For example, the Bioclear™ 11 film used in WAVE Cellbag™ bioreactors has been designed to have a low antioxidant profile for better compatibility with certain sensitive cell lines.

Question 16

Do you have recommendations for deciding how detailed the evaluation of extractables and leachables need to be? I know it is up to the company to decide but are there some guiding principals? Also, how is the work split between company and supplier when doing these evaluations. Is there some documentation that you will provide as part of the process?

A drug safety profile should be clinical-phase appropriate and based on patient risk. Considerations regarding extractables / leachables are an important part of the risk assessment. A number of factors need to be evaluated as part of the assessment including the interactions between process, fluid stream, and the single use assembly. The level of detail about extractables / leachables required for the assessment will depend on the users experience and historical data on the single-use systems, as well as the nature of the chemical composition of the single-use systems and the formulations that will come in contact with them. In recent years, GE Healthcare has invested in expanding our internal capabilities for characterizing the materials used in our single-use systems. We provide information about our single-use systems such as the materials of construction, biocompatibility data, as well as profiles of extractables. This type of information can be leveraged in combination with user-generated in-process leachables data as inputs into the risk assessment. More details on this topic can be found in the PDA Technical Report No. 66 Application of Single-Use Systems in Pharmaceutical Manufacturing.

Question 17

How do you match up the model solvents you use for testing with drug products?

The extraction studies conducted on single-use systems must try to account for as many extractable compounds as possible since the extraction efficiency of the fluids they are intended to contain will vary widely depending on a user's specific formulations. In light of the many different classes of compounds that can have the potential to migrate from a material, an extraction study should include suitable solvents of differing polarities and pH values in order to achieve a thorough extraction. A range of solvent polarities is needed because the compounds that have the potential to extract from a material may have different polarities, in which case their solubility in one solvent may be better than in another. In addition, pharmaceutical formulations can vary widely in chemical composition, which can result in different extraction efficiencies. A simple, but useful guide to solubility is "like dissolves like". For example, a nonpolar compound is more readily soluble in a nonpolar solvent, and therefore, more easily extracted from the matrix of a material. In some cases, the pH of the extraction solvent can have a significant impact on the removal of certain compounds. Thus, including both acidic and basic extraction solvents is important for the removal of such compounds.

Extraction studies with single-use bioprocess systems typically include the following solvents: neutral water (WFI or equivalent), pH 2 ± 0.2 (created with neutral water and HCl), pH 13 ± 0.2 (created with neutral water and NaOH or ammonium hydroxide), 50% ethanol in neutral water, 1% Polysorbate 80, and 5M NaCl. Neutral water is a polar solvent that readily solubilizes ionic compounds (i.e. salts, acids, bases etc.) and small polar organic compounds. The addition of both acidic and basic solutions provides a bracket approach to solution pH, which encourages the migration of both organic bases and acids, respectively. Mixtures of polar organic solvents such as ethanol with water allow for the extraction of a wider range of polar organic molecules. Ionic compounds and metals tend to be more readily soluble in high ionic strength solutions. The choice of a highly concentrated brine solution of 5M NaCl provides the type of ionic strength environment suitable for efficient extraction of such compounds. In order to extract non-polar molecules, surface active agents (surfactants) are used because they form aggregates in solution with hydrophobic cores (non-polar) that provide a more favorable environment for the solubilization of non-polar molecules. Polysorbate 80 is a surfactant typically used in pharmaceutical formulations to enhance the solubility of non-polar molecules; therefore, it is an effective solvent for extracting such compounds from the matrix of a material. The extraction study design implemented by GE Healthcare for single-use bioprocess systems is meant to simulate a wide variety of possible formulation compositions used in biopharmaceutical manufacturing, and therefore extract as many compounds as possible from a given material.

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