The production of cell and gene therapies (CGTs) requires many steps, and each one determines the quality of the final product, especially its patient safety and efficacy. The key technologies used in producing these therapies range from advanced biological procedures to specialized laboratory tools. To explore some of the most interesting advances in this field, GEN talked with eight experts. Here’s a summary of what we learned.
“A key challenge for cell therapies continues to be the industrialization and scale-up of the manufacturing process,” says Martin Westberg, vice president and general manager of cell therapy at Cytiva. “They are complex therapeutics with equally complex and diverse manufacturing processes.”
To improve the production of cell and gene therapies, some companies design complete platforms for this specific application. Shown here is Cytiva’s Sefia cell therapy manufacturing platform, which automates key steps in the process.
So, platforms, reagents, and software work best when they are designed specifically for the production of CGTs. As Westberg says, “Standardizing and industrializing these processes will help alleviate the issues related to cost and access, issues that will only continue to grow as the field moves closer to application in broader oncology indications, as well as indications such as autoimmune diseases.”
To provide a fit-for-purpose approach, Cytiva created its Sefia cell therapy manufacturing platform. “It was developed to automate the cell therapy manufacturing workflow by using a modular and digitally integrated platform, which combines two functionally closed systems together with single-use kits and dedicated applications,” Westberg says. “The Sefia Select system automates cell-isolation, harvest, and formulation steps, while the Sefia expansion system automates cell-activation, transduction, and cell-expansion steps.”
To optimize this platform, Cytiva leveraged two key partnerships. “To understand the needs of the industry and how to better design the technologies and solutions needed to scale-up manufacturing significantly, we developed the Sefia platform as part of a collaboration with Kite, a Gilead company, that is a global leader in cell therapies,” Westberg says. As part of ongoing improvements in scale-up opportunities with this platform, Cytiva recently announced a partnership with U.K.-based Cellular Origins to provide robotic automation of the manufacturing process.
Other companies also take a platform approach to standardizing and accelerating the production of CGTs. For instance, Ori Biotech developed the IRO platform, which automates activation, transduction, expansion, and harvest. Moreover, this platform accommodates a wide variety of cell therapies, including CAR T, and works with many cell types.
“The IRO platform was developed to really solve the scalability challenge in cell and gene therapies,” says Jason C. Foster, Ori Biotech’s CEO. “We’re making personalized, living medicines for specific patients.”
Plus, CGTs must meet quality criteria. Today, as many as two out of five batches of these therapies “don’t meet the specifications that the FDA has set for them,” Foster says. “Our expectation is the IRO platform will drop failure rates from 40% today to less than five percent.”
The fast pace of change in cell and gene therapies requires a flexible production platform. Here, a scientist loads a bioreactor into Ori Biotech’s IRO platform, which was designed to be flexible. It works with many methods of cell therapy and accommodates many cell types.
Even while reducing the failure rate, this platform can speed up the production of therapies. On average, it takes one to two days off production time. It also increases throughput by 10 times and reduces the cost of goods by about 50%. “So, we can get 10 times as many products from the same footprint for half the cost,” Foster says. “That is a big step forward in making these products more affordable and also hopefully moving them to earlier-line therapy.”
Ori Biotech also designed IRO to be flexible. “The platform can adapt to different cell types,” says Foster. “We have no idea what tomorrow’s cell-therapy manufacturers are going to want in their system, so IRO is designed to be flexible so we can handle and adapt to different cell types and future advancements to deliver the best biological and clinical performance.”
A gene therapy can be delivered with a DNA plasmid, which is circular DNA produced in bacteria, to repair a flaw in a patient’s chromosomes. Scientists at
Aldevron have been making plasmids for 25 years, and Robert Reames, the company’s vice president of technical operations, points out three key challenges.
The first challenge is scale. “We’re talking about supporting personalized medicines, where instead of making one large batch of grams or hundreds of grams for thousands of patients, we’re talking about needing to scale out to produce dozens or hundreds of small-scale individual batches.” Second, the therapeutic sequences keep increasing in complexity. As examples, Reames mentions the use of DNA plasmids to produce adeno-associated viruses (AAVs) for gene therapy and in mRNA manufacturing, where the production of complex sequences of DNA creates upstream challenges in ensuring the fidelity of the sequences. Third, he says, is “the desire that customers have for more stringent purification specifications.” That means making a very pure product that is free of unwanted components, such as endotoxins or non-supercoiled DNA.
One of the ways that Aldevron addresses these challenges is with new technologies for downstream purification. “Specifically, some new membrane technologies are utilized for the initial capture and polishing purification steps of plasmids,” Reames says. “With these technologies, you’re able to process the material quicker and reduce the costs associated with producing that material by having lower-cost materials.”
These purification technologies were designed for smaller molecules, such as recombinant proteins or monoclonal antibodies. So, Aldevron invested in “a lot of process development optimization of these technologies to make them work specifically for plasmids,” Reames explains.
The development of a gene therapy depends on assays to evaluate the safety and efficacy of the treatment, as well as its quality. For example, assays are used to “measure the overall expression levels, functionality, and integration of the therapeutic gene in target cells,” says Despoina Lymperopoulou, PhD, senior manager, pharma analytics at Thermo Fisher Scientific. “Additionally, assays help detect any impurities or contamination that may arise from the production of the therapy.”
As examples, Lymperopoulou points out Thermo Fisher’s recent MycoSEQ Plus qPCR, AAV genomic titer dPCR, and residual HEK293 dPCR assays. “These assays help safeguard manufacturing integrity by detecting mycoplasma contamination to protect cell viability and prevent production disruptions, enabling precise viral genome quantification for accurate dosing, and minimizing risks associated with residual host cell DNA, including potential immunogenicity or oncogenicity,” Lymperopoulou says.
Developing these types of assays, however, poses significant challenges. “For the AAV dPCR assay, selecting a highly specific target compatible with various AAV vectors was critical,” Lymperopoulou explains. “We identified a target that does not cross-react with non-AAV vectors or host cell DNA, ensuring high specificity.” In addition, Thermo Fisher needed to consider sample preparation for this assay. “Since both extraction-free and extraction-based approaches have distinct advantages, we developed protocols that offer customers the flexibility to choose the method that best suits their needs,” Lymperopoulou says.
In the world of assays, though, improvements are always being developed. As one example, Lymperopoulou mentions rapid sterility assays, which “will be critical for early detection and mitigation of contamination risks in costly batch production, ensuring the safety and compliance of the final product.”
Other companies are also working on better assays for use in producing CGTs. For instance, Marjorie Smithhisler, director of commercial development for cell therapy at Lonza, says, “We have developed and continue to grow a library of standard assays for both viral vector and cell-therapy projects. Our assay library took years of experience and expertise to develop, and benefits all of our customers, who won’t need to develop these assays from scratch,” Smithhisler says. “These standard offerings enable faster turnaround time on important budgetary decisions, as well as to meet timelines and milestones for therapeutic development.” Lonza’s services cover all phases of viral-vector work and cell-therapy assays from feasibility through qualification for platform and non-platform assays, and also include exosome-based characterization, which is an important aspect to better understand these new therapeutics, Smithhisler notes.
The manufacturer must run analytics throughout the production of a CGT, and that gets more complicated as more therapies are being produced for more patients.
To address the analytics bottleneck, “Automation would allow for increased capacity, with reduced error and variability as the number of batches increases for cell and gene therapy manufacturing processes that need to meet the expected worldwide demand,” says Darrin
Kuijstermans, PhD, head of process development laboratories at Lonza’s site in Geleen, Netherlands. “In addition, with the introduction of fully or semi-automated analytics, we can reduce one of the main bottlenecks in cell and gene therapies in regard to quality-control capacity challenges, making sure resource requirements supported by analytics automation help address capacity and scalability concerns for manufacturing.”
Implementing this approach to analytics, though, depends on “the additional skill set requirements in automation engineering and adding this new category to essential resource needs apart from the bioprocess engineers and biologists currently on hand,” Kuijstermans says. “Other aspects to consider are the data integrity requirements and thus making sure the automated platforms can meet regulatory expectations upon implementation.”
In addition to producing CGTs, a bioprocessor needs something to put them in. As an alternative to cryopreservation bags (“cryobags”), BioLife Solutions developed its CellSeal CryoCases specifically for CGTs. These containers are rigid because they are made from cyclic olefin copolymer plastic. Compared to using pliable plastic bags as containers, the CryoCases have “less fracture risk,” says Sean Werner, PhD, the company’s chief technology officer for cell processing.
The BioLife Solutions CellSeal CryoCase (inset) is a rigid cell and gene therapy primary container that can be sealed and cryogenically frozen in a lab. These containers work with various automated platforms, as shown here (right) where five CryoCases were filled by the Signata CT-5 a semi-automated fill and finish system.
In addition, these cases include ports for closed filling and retrieval of a therapy, which can be kept in long-term storage at cryogenic temperatures, such as those reached when storing the cases in vapor-phase liquid nitrogen. Moreover, one CryoCase can hold up to 75 milliliters of a therapy.
This technology also eliminates some steps required when storing a therapy in a cryobag. After filling a cryobag, air bubbles must be removed, and that increases the chances of damaging the container. With CryoCases, there are no air bubbles to remove. “So, you remove the fracture risk that’s related to air bubbles and the high level of handling requirements and difficulty in handling bags in preparation for cryo-storage,” Werner says.
BioLife Solutions Frozen CryoCase
The CryoCases also simplify handling in other ways. For example, BioLife designed its containers to be grabbed by a robotic arm. “We’re working with a number of automation partners focused on cell and gene therapies,” Werner says.
In many steps in the production of CGTs, devices must be connected. Nonetheless, “The industry has had to rely on older techniques like biosafety cabinets or tube welding for sterile processing,” says Nick Johnson, business unit director at CPC (Colder Products Company) Biopharma. “There is a growing need for more efficient sterile processing techniques.”
CPC created what Johnson describes as “the industry’s smallest aseptic connectors—MicroCNX Series Connectors—which are designed to provide a simple, efficient method of sterile connection of tubing used for small-format assemblies.”
Manufacturers of CGTs often use 1/8 and 1/16 inch flow paths, and the small tubing can create its own set of challenges. “Welding 1/16-inch tubes together requires time-consuming precision to ensure the tubing’s flow path remains open after clamping and heat application,” says Johnson. “Any offset of the tubing at the weld point can lead to leakage and/or contamination risk.” Several features of MicroCNX address these challenges. For example, Johnson points out that CPC’s connectors “do not require extra tubing, and the connectors can be quickly joined outside of a biosafety cabinet.”
Various stages of producing cell and gene therapies depend on reliable connectors. For example, CPC’s MicroCNX ULT Series connectors shown here help to simplify sterile processing. These connectors can be used in applications that require freezing to temperatures as low as −190°C.
CPC’s connectors also work well across a range of environmental conditions. “Many cell therapy processes are dropping into -150°C or lower temperature ranges to support cell health, shelf life, and transportation needs,” says Johnson. “So in the past year, CPC has introduced two new ultra-low temperature aseptic connectors: the MicroCNX ULT Series and the MicroCNX Nano Series,” which can be frozen to -190°C using vaporized liquid nitrogen.
For existing setups, it’s easy to adopt CPC’s connectors. As Johnson notes, “Our MicroCNX connectors are compatible with the common tubing types deployed in developing advanced therapies, including silicone, thermoplastic elastomer, and most polyvinyl chloride.”
As described above, the production of CGTs depends on a wide range of technologies, from tools and techniques to assays and platforms. Optimizing the production of any of these therapies requires a complete ecosystem of interacting tools and techniques. And just like ecosystems in nature, this one continues to evolve.
The post Improving CGT Production to Speed Delivery and Lower Cost appeared first on GEN - Genetic Engineering and Biotechnology News.
A fit-for-purpose platform
“A key challenge for cell therapies continues to be the industrialization and scale-up of the manufacturing process,” says Martin Westberg, vice president and general manager of cell therapy at Cytiva. “They are complex therapeutics with equally complex and diverse manufacturing processes.”
To improve the production of cell and gene therapies, some companies design complete platforms for this specific application. Shown here is Cytiva’s Sefia cell therapy manufacturing platform, which automates key steps in the process.
So, platforms, reagents, and software work best when they are designed specifically for the production of CGTs. As Westberg says, “Standardizing and industrializing these processes will help alleviate the issues related to cost and access, issues that will only continue to grow as the field moves closer to application in broader oncology indications, as well as indications such as autoimmune diseases.”
To provide a fit-for-purpose approach, Cytiva created its Sefia cell therapy manufacturing platform. “It was developed to automate the cell therapy manufacturing workflow by using a modular and digitally integrated platform, which combines two functionally closed systems together with single-use kits and dedicated applications,” Westberg says. “The Sefia Select system automates cell-isolation, harvest, and formulation steps, while the Sefia expansion system automates cell-activation, transduction, and cell-expansion steps.”
To optimize this platform, Cytiva leveraged two key partnerships. “To understand the needs of the industry and how to better design the technologies and solutions needed to scale-up manufacturing significantly, we developed the Sefia platform as part of a collaboration with Kite, a Gilead company, that is a global leader in cell therapies,” Westberg says. As part of ongoing improvements in scale-up opportunities with this platform, Cytiva recently announced a partnership with U.K.-based Cellular Origins to provide robotic automation of the manufacturing process.
Reducing failure rates
Other companies also take a platform approach to standardizing and accelerating the production of CGTs. For instance, Ori Biotech developed the IRO platform, which automates activation, transduction, expansion, and harvest. Moreover, this platform accommodates a wide variety of cell therapies, including CAR T, and works with many cell types.
“The IRO platform was developed to really solve the scalability challenge in cell and gene therapies,” says Jason C. Foster, Ori Biotech’s CEO. “We’re making personalized, living medicines for specific patients.”
Plus, CGTs must meet quality criteria. Today, as many as two out of five batches of these therapies “don’t meet the specifications that the FDA has set for them,” Foster says. “Our expectation is the IRO platform will drop failure rates from 40% today to less than five percent.”
The fast pace of change in cell and gene therapies requires a flexible production platform. Here, a scientist loads a bioreactor into Ori Biotech’s IRO platform, which was designed to be flexible. It works with many methods of cell therapy and accommodates many cell types.
Even while reducing the failure rate, this platform can speed up the production of therapies. On average, it takes one to two days off production time. It also increases throughput by 10 times and reduces the cost of goods by about 50%. “So, we can get 10 times as many products from the same footprint for half the cost,” Foster says. “That is a big step forward in making these products more affordable and also hopefully moving them to earlier-line therapy.”
Ori Biotech also designed IRO to be flexible. “The platform can adapt to different cell types,” says Foster. “We have no idea what tomorrow’s cell-therapy manufacturers are going to want in their system, so IRO is designed to be flexible so we can handle and adapt to different cell types and future advancements to deliver the best biological and clinical performance.”
Producing purer plasmids
A gene therapy can be delivered with a DNA plasmid, which is circular DNA produced in bacteria, to repair a flaw in a patient’s chromosomes. Scientists at
Aldevron have been making plasmids for 25 years, and Robert Reames, the company’s vice president of technical operations, points out three key challenges.
The first challenge is scale. “We’re talking about supporting personalized medicines, where instead of making one large batch of grams or hundreds of grams for thousands of patients, we’re talking about needing to scale out to produce dozens or hundreds of small-scale individual batches.” Second, the therapeutic sequences keep increasing in complexity. As examples, Reames mentions the use of DNA plasmids to produce adeno-associated viruses (AAVs) for gene therapy and in mRNA manufacturing, where the production of complex sequences of DNA creates upstream challenges in ensuring the fidelity of the sequences. Third, he says, is “the desire that customers have for more stringent purification specifications.” That means making a very pure product that is free of unwanted components, such as endotoxins or non-supercoiled DNA.
One of the ways that Aldevron addresses these challenges is with new technologies for downstream purification. “Specifically, some new membrane technologies are utilized for the initial capture and polishing purification steps of plasmids,” Reames says. “With these technologies, you’re able to process the material quicker and reduce the costs associated with producing that material by having lower-cost materials.”
These purification technologies were designed for smaller molecules, such as recombinant proteins or monoclonal antibodies. So, Aldevron invested in “a lot of process development optimization of these technologies to make them work specifically for plasmids,” Reames explains.
Innovative assays
The development of a gene therapy depends on assays to evaluate the safety and efficacy of the treatment, as well as its quality. For example, assays are used to “measure the overall expression levels, functionality, and integration of the therapeutic gene in target cells,” says Despoina Lymperopoulou, PhD, senior manager, pharma analytics at Thermo Fisher Scientific. “Additionally, assays help detect any impurities or contamination that may arise from the production of the therapy.”
As examples, Lymperopoulou points out Thermo Fisher’s recent MycoSEQ Plus qPCR, AAV genomic titer dPCR, and residual HEK293 dPCR assays. “These assays help safeguard manufacturing integrity by detecting mycoplasma contamination to protect cell viability and prevent production disruptions, enabling precise viral genome quantification for accurate dosing, and minimizing risks associated with residual host cell DNA, including potential immunogenicity or oncogenicity,” Lymperopoulou says.
Developing these types of assays, however, poses significant challenges. “For the AAV dPCR assay, selecting a highly specific target compatible with various AAV vectors was critical,” Lymperopoulou explains. “We identified a target that does not cross-react with non-AAV vectors or host cell DNA, ensuring high specificity.” In addition, Thermo Fisher needed to consider sample preparation for this assay. “Since both extraction-free and extraction-based approaches have distinct advantages, we developed protocols that offer customers the flexibility to choose the method that best suits their needs,” Lymperopoulou says.
In the world of assays, though, improvements are always being developed. As one example, Lymperopoulou mentions rapid sterility assays, which “will be critical for early detection and mitigation of contamination risks in costly batch production, ensuring the safety and compliance of the final product.”
Other companies are also working on better assays for use in producing CGTs. For instance, Marjorie Smithhisler, director of commercial development for cell therapy at Lonza, says, “We have developed and continue to grow a library of standard assays for both viral vector and cell-therapy projects. Our assay library took years of experience and expertise to develop, and benefits all of our customers, who won’t need to develop these assays from scratch,” Smithhisler says. “These standard offerings enable faster turnaround time on important budgetary decisions, as well as to meet timelines and milestones for therapeutic development.” Lonza’s services cover all phases of viral-vector work and cell-therapy assays from feasibility through qualification for platform and non-platform assays, and also include exosome-based characterization, which is an important aspect to better understand these new therapeutics, Smithhisler notes.
Automating analytics
The manufacturer must run analytics throughout the production of a CGT, and that gets more complicated as more therapies are being produced for more patients.
To address the analytics bottleneck, “Automation would allow for increased capacity, with reduced error and variability as the number of batches increases for cell and gene therapy manufacturing processes that need to meet the expected worldwide demand,” says Darrin
Kuijstermans, PhD, head of process development laboratories at Lonza’s site in Geleen, Netherlands. “In addition, with the introduction of fully or semi-automated analytics, we can reduce one of the main bottlenecks in cell and gene therapies in regard to quality-control capacity challenges, making sure resource requirements supported by analytics automation help address capacity and scalability concerns for manufacturing.”
Implementing this approach to analytics, though, depends on “the additional skill set requirements in automation engineering and adding this new category to essential resource needs apart from the bioprocess engineers and biologists currently on hand,” Kuijstermans says. “Other aspects to consider are the data integrity requirements and thus making sure the automated platforms can meet regulatory expectations upon implementation.”
Therapeutic containers
In addition to producing CGTs, a bioprocessor needs something to put them in. As an alternative to cryopreservation bags (“cryobags”), BioLife Solutions developed its CellSeal CryoCases specifically for CGTs. These containers are rigid because they are made from cyclic olefin copolymer plastic. Compared to using pliable plastic bags as containers, the CryoCases have “less fracture risk,” says Sean Werner, PhD, the company’s chief technology officer for cell processing.
The BioLife Solutions CellSeal CryoCase (inset) is a rigid cell and gene therapy primary container that can be sealed and cryogenically frozen in a lab. These containers work with various automated platforms, as shown here (right) where five CryoCases were filled by the Signata CT-5 a semi-automated fill and finish system.
In addition, these cases include ports for closed filling and retrieval of a therapy, which can be kept in long-term storage at cryogenic temperatures, such as those reached when storing the cases in vapor-phase liquid nitrogen. Moreover, one CryoCase can hold up to 75 milliliters of a therapy.
This technology also eliminates some steps required when storing a therapy in a cryobag. After filling a cryobag, air bubbles must be removed, and that increases the chances of damaging the container. With CryoCases, there are no air bubbles to remove. “So, you remove the fracture risk that’s related to air bubbles and the high level of handling requirements and difficulty in handling bags in preparation for cryo-storage,” Werner says.
BioLife Solutions Frozen CryoCase
The CryoCases also simplify handling in other ways. For example, BioLife designed its containers to be grabbed by a robotic arm. “We’re working with a number of automation partners focused on cell and gene therapies,” Werner says.
Simplifying sterile connections
In many steps in the production of CGTs, devices must be connected. Nonetheless, “The industry has had to rely on older techniques like biosafety cabinets or tube welding for sterile processing,” says Nick Johnson, business unit director at CPC (Colder Products Company) Biopharma. “There is a growing need for more efficient sterile processing techniques.”
CPC created what Johnson describes as “the industry’s smallest aseptic connectors—MicroCNX Series Connectors—which are designed to provide a simple, efficient method of sterile connection of tubing used for small-format assemblies.”
Manufacturers of CGTs often use 1/8 and 1/16 inch flow paths, and the small tubing can create its own set of challenges. “Welding 1/16-inch tubes together requires time-consuming precision to ensure the tubing’s flow path remains open after clamping and heat application,” says Johnson. “Any offset of the tubing at the weld point can lead to leakage and/or contamination risk.” Several features of MicroCNX address these challenges. For example, Johnson points out that CPC’s connectors “do not require extra tubing, and the connectors can be quickly joined outside of a biosafety cabinet.”
Various stages of producing cell and gene therapies depend on reliable connectors. For example, CPC’s MicroCNX ULT Series connectors shown here help to simplify sterile processing. These connectors can be used in applications that require freezing to temperatures as low as −190°C.
CPC’s connectors also work well across a range of environmental conditions. “Many cell therapy processes are dropping into -150°C or lower temperature ranges to support cell health, shelf life, and transportation needs,” says Johnson. “So in the past year, CPC has introduced two new ultra-low temperature aseptic connectors: the MicroCNX ULT Series and the MicroCNX Nano Series,” which can be frozen to -190°C using vaporized liquid nitrogen.
For existing setups, it’s easy to adopt CPC’s connectors. As Johnson notes, “Our MicroCNX connectors are compatible with the common tubing types deployed in developing advanced therapies, including silicone, thermoplastic elastomer, and most polyvinyl chloride.”
Optimizing the ecosystem
As described above, the production of CGTs depends on a wide range of technologies, from tools and techniques to assays and platforms. Optimizing the production of any of these therapies requires a complete ecosystem of interacting tools and techniques. And just like ecosystems in nature, this one continues to evolve.
The post Improving CGT Production to Speed Delivery and Lower Cost appeared first on GEN - Genetic Engineering and Biotechnology News.