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Feliza Mirasol

Hollie Barton, senior director, Chromatographic Services, for the PPD clinical research business of Thermo Fisher Scientific, spoke with 

®) at AAPS PharmSci 360 on the need for training the current and the new generation of analytical experts for the bio/pharma industry. With the emergence and growth of newer modalities of therapeutics, such as cell and gene therapies, messenger RNA-based and other nucleic acid-based therapeutics, and more advanced forms of antibodies, to name a few, 

the industry has been feeling the void of a lack in technical and analytic expertise

 (1). Barton highlights at least one approach toward filling that void and ensuring the future of analytical expertise.

“It used to be, when folks joined the BioA labs, they went through what was referred to as a new hire training, where they were just exposed to safety, GLP [good laboratory practice], the basics. What we're seeing now is an [addition] of trainers directly associated with functional teams they're supporting. These trainers are actual lab scientists who are coming in, and the first things that they're doing is exposing our folks to protocols, methodology, therapeutic areas, and helping them to understand exactly what they're working on,” Barton explains.

By having these in-house trainers available to staff, she continues, the staff are being advanced at a much quicker rate. “We're also seeing a lot of mentoring and one-on-one training that is being provided to get everybody where they need to be as quickly as possible,” Barton remarks.

AAPS PharmSci 360 ran Oct. 22–25 in Orlando, Fl.

analytical challenges in meeting turnaround times

1. Mirasol, F. Biologics Testing Highlights Need for Analytical Skills. 

Articles

Hollie Barton, senior director, Chromatographic Services, for the PPD clinical research business of Thermo Fisher Scientific, highlights one approach to ensuring the continuation of analytical expertise.

Ensuring the Next Generation of Analytical Experts (AAPS PharmSci 360)

Scale up in gene therapy manufacturing currently revolves around viral vector scale up. Although some processes are similar to those in monoclonal antibody (mAb) production (e.g., cell culture to produce the viral vectors), the manufacture of viral vectors carries its own challenges, some of which may be addressed based on the history of mAb manufacture.

A look at the similarities between mAb manufacture and viral vector manufacture show that both products rely on a similar synthesis process: a biochemical reaction-based process and the use of cell culture vessels/bioreactors, according to Naveenganesh Muralidharan, senior manager, manufacturing science and technology group, AGC Biologics, and Suparna Sanyal, director of Commercial Development, Cell and Gene Therapy, Lonza.

With the exception of substrates used for their manufacture, viral vectors and mAbs share in common all the other physiological factors for production, such as temperature, pH, dissolved oxygen, and shear (either from mixing or from sparging gas into the bioreactor for oxygenation). In the production of both, the same process monitoring targets must also be addressed, such as minimizing toxic metabolite accumulation during the cell culture process and sensitivity to times when feed must be added as well as the specific rates at which feed must be added, Muralidharan emphasizes.

Other bioprocessing challenges include cell growth strategy and media optimization in addition to pH control, dissolved oxygen control, mixing, and shear stress, Sanyal adds.

However, one unique challenge that viral vector manufacture poses is that—in the case of lentivirus vectors—lentivirus allows for permanent integration of the vector into the host cell DNA, says Bruce Lahn, PhD, chief scientist, VectorBuilder, a US-headquartered biotech company specializing in gene delivery technologies. “This allows for theoretical ‘permanent’ expression of the transgene in the cells,” Lahn notes. Although other viral vectors, such as an adeno-associated virus (AAV) vector, can be more “transient” than lentivirus, meaning that, although AAV-mediated expression of transgenes, for example, has been sustainable for years, the AAV vector does not integrate into the host genome.

“AAV has a significantly lower immune reaction than lentivirus,” says Lahn, adding that “clinical trials with lentivirus were initially unsuccessful due to the significant patient immune response to the lentivirus itself.” Whereas, AAV clinical trials have demonstrated the safety of AAV vectors, which produce a much lower immune response than lentivirus, he explains.

Lahn furthermore notes that the differences between types of viral vectors (e.g., lentivirus vs. AAV) can be both advantageous and disadvantageous, depending on the end goal of the therapy in development. “If you are looking to target only a specific organ or tissue, then AAV would be your choice. For example, a lot of research has been done to target AAV specifically to retinal cells for the treatment of various vision disorders as well as target it to the brain to deliver therapeutic genes for neurological disorders,” he explains.

The decades of experience by the biopharma industry in the scale-up of mAb manufacturing can offer some lessons that have been applied to viral vector manufacturing. “The tools, technologies, know-how, and infrastructure for efficient cell expansion, seed train generation and scaling up production using single-use bioreactors has been well established in current good manufacturing practices (CGMP) processes for mAbs,” says Sanyal. Because of this, best practices such as cell banking, viability, safety and stability assessment of cell lines, cell growth strategies, metabolic analysis, and production parameter optimization can be easily leveraged for viral vector manufacturing, she emphasizes.

“Over the years of experience in mAb manufacturing processes, companies have templated scale-up activities in biologics. However, viral vector manufacturing processes are novel and highly variable from one manufacturer to other,” states Muralidharan. Therefore, what the industry has established thus far with regards to mAb manufacturing is an in-depth product knowledge for defining critical quality attributes and characterizing the critical process parameters of biomanufacturing processes. The industry also holds a detailed understanding of the variability introduced from raw materials and has well-established process operations for mAb process, says Muralidharan. However, this same depth of knowledge has yet to be established for new therapeutic technologies such as in-vitro gene therapies. As such, setting the release specifications for viral vector manufacturing based on the currently limited process and product knowledge of these molecules remains a challenging task, Muralidharan points out.

Other unique challenges associated with viral vector manufacture include the presence of impurities when produced. As Muralidharan explains, viral vectors manufactured from cell-substrate production systems can contain residual amounts of impurities such as host-cell DNA (hcDNA) and plasmid DNA (pDNA). In the final product, the presence of either impurity presents safety risks to treated patients. To address this risk, global regulatory agencies recommend tight, quantifiable limits for DNA impurities for protein therapeutics (10 ng/dose), according to Muralidharan. “However, because of the inherent nature of hcDNA and pDNA encapsidation in viral capsids, FDA permits viral vector manufacturers to minimize the amount and size of DNA impurities based on risk assessment, rather than requiring quantifiable DNA impurity limits,” he states.

Manufacturers, therefore, define their own hcDNA and pDNA specification limits for final products based on risk assessments to ensure patient safety. Such assessments are not necessarily straightforward, however, and require methods and materials that might not be readily available and that are yet to be established, Muralidharan adds.

“The production process for mAbs is largely standardized and cost-efficient with ease of scalability, using suspension bioreactors and stable cell lines,” says Sanyal. The processes used for viral vector manufacture, in comparison, are more diverse and bespoke. “The majority of viral vector production processes utilize transient transfection processes, which are difficult to scale up and require the use of costly plasmid DNAs,” she points out.

Sanyal also brings up the fact that the production of viral vectors for gene therapies is an emerging market and predominantly targets rare genetic disorders. As such, the target patient population is rather small and requires only small amounts of viral vector for the early phase clinical trials, or even in post-commercialization, she says. “For this reason, many therapeutic developers continue to utilize 2D adherent cell culture processes that are faster to implement but not scalable or cost-effective. As the gene therapy market matures, there is an increase in the recent approvals of viral-vector-based products and a concomitant shift towards the adoption of suspension bioreactor-based production processes to meet market demand,” she adds.

Because transferring non-optimized small-scale R&D processes to commercial scale can lead to much uncertainty, according to Muralidharan, and that uncertainty can lead to a high risk of failure to pass through process performance qualification (PPQ) and meeting that validated state consistently during commercial production, it makes sense to have smart steps in place for when scale up moves forward.

“The smart step is to start planning for commercialization as early as the preclinical testing phase. This early planning for commercialization will lead to developing scalable process optimization work during early development. The key for successful commercial process starts there,” says Muralidharan.

Meanwhile, Sanyal points to the transition to a 3D suspension, bioreactor-based process as a key first step in scaling up viral vector manufacturing. The next smart step from there is process optimization for upstream unit operations to maximize viral vector productivity per cell. Following that, subsequent optimization of downstream purification to help maximize yield, purity, and product quality. “The use of perfusion-based processes can also improve production in some cases,” Sanyal says. “Additionally, the use of staggered campaign manufacturing strategies can be utilized to maximize production throughput to meet commercial demand.”

Finally, Lahn discusses the benefit of having a wide variety of AAV serotypes to work with, as AAV serotypes can target specific tissues. As such, “one of the hottest trends in the gene therapy world is the work to create new AAV serotypes that can target specific tissues or cell types. This process can be done by rational design or by creating an extensive library of AAV capsids and screening for capsids that can infect cells/organs more efficiently or infect new cell types,” he explains.

Furthermore, with machine learning and artificial intelligence becoming more prevalent in scientific research, their use has shown great promise in identifying novel AAV capsids. This stands in contrast to the lentiviral system, which cannot be optimized and screened this way with “smart” tools, notes Lahn.


Volume 47, No.11
November 2023
Pages: 20–21

When referring to this article, please cite it as Mirasol, F. Gleaning Scale-Up Lessons from mAbs for Viral Vectors. 

Monoclonal antibody and viral vector manufacture share similarities, but vector scale up faces unique challenges.

Gleaning Scale-Up Lessons from mAbs for Viral Vectors

® at AAPS PharmSci 360 about how PPD is targeting automation advances to address the bio/pharma industry’s need to move more efficiently and quickly, especially when adding staff alone is not always the answer.

“2023, through the latter part of 2023, and into 2024 have been by far our biggest investment in information technology (IT),” Barton says. “We are looking at automation from a front-end to the back-end—fully automated workflows—and, so, it's pretty exciting to see some of the robotics that are showing up in the laboratories.”

Barton also expresses enthusiasm about how PPD is also adding teams, such as an informatics team, which functions between the operations group and the IT group, bridging the gap between the two groups. Having the informatics group in place results in the addition of analytic dashboards, according to Barton.

“We're using those dashboards to hone in on ensuring we are right first-time [allowing us to] focus on quality and ensure that errors that are repetitive are being addressed, and that is being done through process improvements using that automation,” Barton states.

The company has also added a group called PPI—practical process improvement. “That team is looking at efficiencies and evaluating automation, while following the workflow of the departments,” she says. Even with just the smallest changes recommended by the PPI team, such as reducing the number of steps between work stations or work areas, the company is already seeing improvements, she notes.

Hollie Barton, senior director, Chromatographic Services, for the PPD clinical research business of Thermo Fisher Scientific, reviews the company’s latest moves in targeting automation.

Targeting Automation Advances to Meet Industry Needs (AAPS PharmSci 360)

Hollie Barton with Thermo Fisher Scientific, discusses ways that labs are addressing the pressure of rapid analytical turnaround times.

Addressing the Analytical Challenge of Meeting Quick Turnaround Times (AAPS PharmSci 360)

® at AAPS PharmSci 360 where she discussed the challenges of increased demand for longer lead times from analytical labs in an environment where clinical pressures are calling for drastically reduced turnaround times. In this interview, Barton emphasizes how one such group, BioA labs, has been addressing the need to move faster when cycle times are being determined by methodology.

“Actually, the labs are kind of taking a two-tiered approach to that. They are doing some internal and some external things. Internally, we're watching BioA labs move to 24 hour, seven-day- week work shifts [but also with] fluid shifts within that. When I say fluid shifts, we're really looking at folks to work possibly five–eight hour shifts, and then the next week possibly moved to four–10 [shifts], maybe even three–twelve [shifts], to attack [the] methodology they're having to run,” Barton says.

She also notes that project management is being added to the BioA labs. Normally, those lab personnel reside at the central laboratory or clinical sites. “What we're seeing is them being placed in the BioA lab to provide a single point-of-contact with communication that's flowing through. [That] addition has really cut down on the work that the PI [principal investigator] has to do in regard to communication and point-of-contact,” Barton explains.

This slight change has allowed labs, particularly PIs, to better focus better on the science, which is where the investigators’ attention should be, Barton emphasizes. Barton also notes that, externally, she is seeing BioA labs pushing back on contracts, especially in situations where there are no central laboratories involved. “That moves the needle in the wrong direction if you don't bring in the central lab because the BioA lab should not be performing those responsibilities,” she observes.

“We're also seeing where we've added additional teams to the back-end, externally, to better look at protocols. So, we're adding site tech teams to understand exactly what we need to be quoting and looking at. We're seeing all of those things from an internal and external push on clients,” Barton concludes.

Hollie Barton, senior director, Chromatographic Services, for the PPD clinical research business of Thermo Fisher Scientific, discusses ways that labs are addressing the pressure of rapid analytical turnaround times.

On Oct. 30, 2023, Aragen, a contract research development and manufacturing organization (CRDMO), announced that it was setting up a new biologics manufacturing facility in Bangalore, India as part of a $30 million investment.

The new 160,000 m2 facility will include process development labs and multiple good manufacturing practice (GMP) suites as well as supporting functions such as quality control labs (analytical and microbiology). The GMP manufacturing suites will be equipped to conduct intensified bioprocessing using single-use bioreactors and will have advanced downstream purification capability, according to a company press release.

The facility will be designed to offer integrated solutions across the biomanufacturing lifecycle—from process development to process validation, analytical development, pilot production, large-scale drug substance manufacturing, and stability services. The facility will also be equipped to develop and manufacture monoclonal antibodies, therapeutic proteins, and fusion proteins to augment the company’s bioproduction capability in California.

The process development laboratory is slated to be operational by the third quarter of 2024. The first manufacturing suite will open in the third quarter of 2025. The facility will include plasmid DNA, messenger RNA, cell and gene therapy, microbial manufacturing, and additional GMP manufacturing suites in the future to meet demand.

“With the opening of our bio-manufacturing facility, we will empower our customers with even faster development timelines and support their full lifecycle journey from clone development to cell culture manufacturing …, ” said Subodh Deshmukh, CEO, Aragen Bioscience, in the press release.

“Setting up of a biologics manufacturing facility in India is part of our forward integration strategy for our US-based biologics business, allowing Aragen to offer integrated gene-to-GMP solutions to our customers. Once this facility is commissioned, Aragen will offer a unique value proposition to our biologics customers where the early stage discovery will be executed in our R&D labs in California … As the molecule advances, Aragen can tech-transfer the program to our … manufacturing facility in Bangalore, offering better COGS [cost of goods and services] to our customers,” said Manni Kantipudi, CEO, Aragen Life Sciences, in the press release.

As part of a $30 million investment, Aragen is setting up a new biologics manufacturing facility in Bangalore, India.

Aragen to Establish $30 Million Biomanufacturing Facility in India

® at AAPS PharmSci 360, Sara Fathollahi, PhD, product application specialist, DFE Pharma, gives a summary of the takeaway message from her presentation at the conference.

“My main point … was [about] how excipient suppliers (taking DFE Pharma as an example) can support a smooth transition from batch to continuous manufacturing. I believe that the pharma industry is like a big complex puzzle. Each of us—excipient suppliers, manufacturers, machinery suppliers, universities, research centers—we are pieces of this puzzle, and we are individual pieces, but, together, we make the big, impressive image of innovation,” Fathollahi explains.

In her talk, Fathollahi showed how her company supports its customers by using excipients to de-risk the manufacturing process. “What we do is that we share [internal] … data. We have historical data of all batches that we produce and that support us [in showing] which variation the customer can expect from our material. Then, we can also select batches for them, and with [those] batches—in addition to experiments that they do on their end—they can prove robustness to the regulatory bodies,” she states.

“To give you a more specific example, let's say that the manufacturer received one, five, or ten batches from us; even though, after conducting really complete characteristics on the batches they received from us, thedata that they have [are] limited to those batches. As a supplier, we have historical data, we can really provide much more information and can give insight about which variation can be expected from our product in the next five years, for example,” Fathollahi emphasizes. In addition, DFE Pharma can also select batches that are representative of the variation that can be expected from a specific product, and those batches can be tested by customers.

“This is the way that we … support a smooth transition from batch to continuous manufacturing—by addressing the challenge of raw material variability,” Fathollahi adds.

Fathollahi was featured as a speaker at AAPS PharmSci 360 where she gave her talk, “

Impact of Raw Materials Variation On Continuous Feeders Performance

.” Her presentation centered around gaining a better understanding of the natural batch-to-batch variability of raw materials; understanding the link between raw material variation and the manufacturing process; understanding which variation is present in a given raw material and what that specific variation means for the manufacturing process; assessing the risk of using raw materials (with their natural variation) on process robustness and consistency of the final product; and understanding how to derisk the use of excipients in continuous manufacturing processes.

redesign concepts for continuous manufacturing

Sara Fathollahi, PhD, product application specialist at DFE Pharma, makes the case of how excipient suppliers can support the transition from batch to continuous manufacturing.