Overcoming Viral Clearance Challenges

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Ensuring the viral safety of biopharmaceuticals is a crucial component of the drug development process. As the diversity of biologic modalities increases, the challenges to viral clearance are rising as well. In January 2024, FDA issued final guidance, including recommendations for both testing and clearance of adventitious viral agents in recombinant proteins, antibody-based drug products, subunit vaccines, virus-like particles, viral vectors, cytokines, and similar products (1). Standards and techniques are outlined for endogenous and adventitious virus detection and identification, testing of unprocessed bulk product, and the design and performance of viral clearance studies, with emphasis placed on the need to establish specific strategies on a case-by-case basis and the importance of thoroughly characterizing starting materials and conducting risk assessments.

Many challenges to successful viral clearance

Achieving successful viral clearance of biologics involves overcoming several complex challenges such as oversimplification/lack of consideration during development, material availability (viral clearance studies can require a large quantity of material, especially during late phase), timeline pressures, contract development and manufacturing organization (CDMO)/contract research organization (CRO) capacity, and implementation of compliant methods and technologies. “These challenges highlight the need for meticulous planning, robust process design, and continuous monitoring to ensure the safety and efficacy of biologic products,” says Alex Harley, subject matter expert in viral clearance at FUJIFILM Diosynth Biotechnologies.

It is necessary, according to Yuanyuan Chen, a senior director of biosafety testing at WuXi Biologics, to analyze all potential routes of viral contamination for the product, including endogenous viruses from the cell line and adventitious viruses from raw materials or introduced during production. For murine cell-derived products, he emphasizes the importance of viral testing of unprocessed bulk (UPB) to determine the number of retrovirus-like particles (RVLPs).

Assessing these potential sources of contamination is complex as it encompasses many disciplines (raw materials, operator training, process hygiene, facility design/engineering), contends Lee R. Stevens, global manufacturing, science, and technology senior engineer in Thermo Fisher Scientific’s Pharma Services business. “Effective preventive strategies are complex, often require complicated designs, and need to be deployed rigorously,” he says. Accurate risk assessment is important, though, so application of excessive layers of control measures can be avoided.

Selecting the right clearance technologies that are effective in removing or inactivating likely virus contaminants, compatible with the manufacturing process, and do not negatively impact the product is essential as well, but can be equally difficult, Chen observes. Developing and qualifying a scale-down model with performance that accurately reflects the actual manufacturing process can also be challenging, he notes.

For instance, large processing ranges that support a wide design space help to ensure a robust process that consistently achieves the required level of viral reduction, but identifying and testing these conditions can be complex and may require preliminary studies and risk assessments, comments Mun Peak Nyon, process development supervisor in Thermo Fisher Scientific’s Pharma Services business.

In addition, because different products have different requirements, including mature modalities such as monoclonal antibodies (mAbs), a product-specific and risk-based approach must be applied to viral clearance study design, which must take into consideration multiple factors, such as the drug declaration stage, unit operations to be evaluated, model virus panel, and virus detection methods, according to Chen. “Inadequate experience or expertise can lead to design flaws and may cause repeat studies, which are time-consuming and costly,” notes Nyon.

Furthermore, many studies are implemented by a contract laboratory, so development of the study must be accomplished in collaboration with the service provider. “Effective communication between the sponsor and the third-party vendor is crucial for aligning on study protocols, timelines, and expectations. Detailed planning is required to coordinate the shipment of materials, scheduling of laboratory time, and execution of the study. Miscommunication or poor planning can lead to delays, errors, and increased costs,” Nyon says.

For Sungchul Yoo, senior director of drug substance manufacturing at Samsung Biologics, the most salient challenge in drug manufacturing from a quality standpoint is the prevention of virus carryover from earlier downstream processes to later stages. “If virus breach occurs, issues [such as] batch failure could hinder the entire manufacturing process, as operators would have to reprocess certain steps if applicable. Identifying and resolving virus carryover early on, ideally at the start of the downstream process, helps avoid production delays and ensures on-time delivery of drugs where needed,” he explains.

Finally, comprehensive documentation to demonstrate compliance with regulatory requirements is crucial, and poor documentation practices can lead to data discrepancies, regulatory non-compliance, and challenges in validating study results, according to Nyon.

It is also worth noting, says Chen, that viral clearance for cell and gene therapies (CGTs) can present additional challenges. While dedicated unit operations for viral clearance exist for mAbs, most CGT products lack such dedicated steps in their purification processes. Similarly, continuous processing creates additional complexities related to establishing an appropriate scale-down model and the efficient execution of viral clearance studies in third-party labs given that continuous processes run for much longer periods and often involve new dedicated equipment, such as multi-column chromatography.

Several keys to ensuring success

Overcoming the many challenges to achieving effective viral clearance in biologics manufacturing requires consideration of several important factors. Primary among them, according to Harley, is balancing product quality, purity, and yield, as some viral clearance techniques can be detrimental to product quality, and some processes may produce great yields and quality, but they may not provide the needed levels of clearance.

Well-devised viral clearance studies using a representative down-scale model and robustly scalable technologies that result in efficacious viral clearance are a must, says Nyon. Stevens also observes that prevention of adventitious contamination, in particular throughout the cell culture and after final viral clearance, and detection at the bulk harvest is needed to assess baseline viral load to the downstream.

Planning appropriate timelines that allow for demonstration of viral clearance through spiking studies, which can take several months, is also important, Harley observes. “Investing in the development of the best viral clearance strategy avoids oversimplification and ensures production of data needed to support effective decision-making and documentation,” he says.

Preventing virus carryover in downstream operations requires the performance of rigorous quality risk assessments, agrees Yoo. “Such an assessment involves precise measurements of the potential virus carryover risk starting from the design phase, in which plans for physical segregation are outlined, through implementation of appropriate mitigation actions. Those actions may include the use of single-use aseptic connections to provide closed systems and separation of areas from other processing activities using secluded air handling units,” he says.

Assessing worst-case parameters in viral clearance studies is crucial when developing a viral clearance strategy, adds Nyon. “Testing under worst-case parameters ensures the process reliably removes viruses, protecting patient health, and meets strict regulatory requirements,” he observes. In addition, Nyon notes that conditions for viral-clearance processes should be selected carefully to ensure the expected viral clearance, manufacturability, and validation, but still offer sufficient flexibility to allow for process improvements through the lifecycle of the molecule.

Model virus stocks used for spiking studies should, meanwhile, be of high-purity and high-titer to minimize the spiking percentage and make scale-down models more representative of the actual manufacturing process. Using ultra-pure virus stocks prepared by special processes enables spiking percentages of less than 1% for the viral filtration and polishing chromatography steps of viral clearance studies, says Chen. “Using these stocks significantly reduces abnormal phenomena such as filter clogging and flux decay, enables higher log reduction values (LRVs), and ensures more accurate and reliable results and the smooth progress of relevant studies,” Chen observes.

Choosing the right method

Successful viral clearance typically requires the use of multiple, complementary viral inactivation and removal methods. Usually, three or more unit operations using orthogonal clearance principles are needed to achieve the cumulative viral clearance required, observes Stevens.

Selection of the appropriate downstream unit operations is therefore a crucial element to a successful viral clearance strategy, according to Chen. “For recombinant proteins, for instance,” he notes, “chromatography steps (including affinity chromatography, ion exchange chromatography, and mix-mode chromatography) used to remove impurities such as host-cell proteins do have some limited viral removal effect, but dedicated steps such as low-pH incubation, solvent/detergent treatment, and virus-retentive filtration are primary techniques.”

When selecting specific viral clearance technologies, Chen adds, attention should be paid first to whether these technologies will have an adverse impact on product quality and then to their feasibility and compatibility with the downstream process. It is also important to use effective and well-understood technologies.

That is because the choice of viral clearance technologies is highly molecule- and process-dependent, comments Matthias D. Kaeser, process development director in Thermo Fisher Scientific’s Pharma Services business. “While using platform processes is a good approach to speed up viral clearance development and can often simplify the selection of appropriate technologies, intimate knowledge about the target molecule and the processing steps will help when developing successful viral clearance strategies for particularly sensitive or difficult-to-purify molecules,” he says.

Implementing a testing strategy that can detect known and unknown contaminants in a process is an equally important element to include in a successful viral clearance strategy, according to Harley. “Doing so can provide an understanding of the RVLP load entering the downstream process and confirm the absence of adventitious agents,” he explains. The RVLP loading can then be used to determine the estimated number of virus-like particles per dose or product for mAb processes.

To prevent virus carryover, Yoo highlights the importance of physical serration. “Separating process areas for pre-viral and post-viral stages ensures no overlapping use of equipment and routes taken,” he says. A pressure cascade system should also be employed to keep consistent airflow and best support physical segregation. If the same equipment is used, Yoo emphasizes the need to clean and steam-sanitize the equipment using a validated cycle.

Leveraging advances in technology

While the general techniques for ensuring viral clearance have not changed significantly over time, the devices and approaches used to implement those techniques have advanced in many ways.Recent examples include single-use membrane chromatography, new virus retention filters, new detergents for viral inactivation, viral surrogate kits, and use of advanced analytical technologies.

“Single-use membrane chromatography has advantages from a viral clearance standpoint as it removes a potential source of cross-contamination from re-used chromatography resins without being cost-prohibitive,” states Kaeser.

New filter technologies are also having an impact. Chen points to a second-generation parvovirus filter membrane that has been optimized in design and materials to enable more effective virus retention. “Better LRVs of parvovirus can be obtained, even in challenging conditions, such as during pressure pauses or when low pressure must be used,” he explains. In addition, Chen notes the filter can be operated stably over a wider range of pH values and chemical environments and offers higher flux capabilities for improved production efficiency.

New detergents for viral inactivation, meanwhile, are often less harsh on sensitive molecules such as enzymes and various modalities of antibodies when compared to traditional low pH inactivation, according to Kaeser. “Their use on clarified harvest is convenient and generally provides good clearance throughout the remainder of the DSP [downstream processing] process,” he says. They are also less harmful to the environment and present fewer extractable/leachable concerns compared to legacy solvent/detergent combinations, thus further facilitating their use in processing.

Viral surrogate kits, observes Harley, allow for assessment of a process’ ability to clear virus, providing information during development activities as opposed to waiting until the end of development or manufacture when it may be too late to add additional clearance steps. He does note, though, that while useful for removal, they currently cannot be used in the estimation of inactivation in a good manufacturing practice (GMP) environment due to regulatory guidelines. Harley adds that the new regulatory guidance does, however, provide a pathway for applying analytical technology advances such as next-generation sequencing and molecular methods in lieu of cell-based assays.

While not a technology advance, Kaeser believes that modular claims on viral clearance offer significant benefits. “For platform processes of monoclonal IgG [immunoglobulin G], many pharmaceutical companies are increasingly adopting modular claims during the early phases of drug development. This approach enables companies to address viral clearance requirements more efficiently,” he explains. As an example, he mentions ASTM E2888-12(2019) (Standard Practice for Process for Inactivation of Rodent Retrovirus by pH), which assures 5 log10 inactivation of murine retrovirus if the low pH virus inactivation parameters are within certain criteria. “By implementing such modular claims early on, companies can better manage regulatory expectations, reduce development timelines, and save costs,” Kaeser contends.

The ability to apply prior knowledge for viral clearance study design, particularly for well-characterized mAb platform processes and risk-based approaches for well-characterized cell lines, is also crucial for the evolution of successful viral clearance strategies, Harley adds. “These advances can give an early indication of what clearance a process can provide and allow for changes to be made before the final process is fixed,” he concludes.

Work with an experienced outsourcing partner

To avoid introducing virus contaminants into GMP facilities/operations, viral clearance studies are typically performed by third-party labs. For small and mid-sized companies that do not have in-house teams with strong viral clearance experience, relying on an experienced CDMO/CRO is also an effective means of mitigating knowledge gaps, Kaeser says.

An experienced third-party lab, according to Chen, possesses in-depth knowledge and expertise both in the field of virology and downstream processing and is familiar with the latest clearance technologies, virus titration methods, and global regulatory requirements. They also have a history of successfully conducting viral clearance studies across various types of biologics. “Combined, this extensive knowledge and experience enables these service providers to fully assess relevant risks and customize scientific viral clearance study plans according to the characteristics of specific products and processes,” he observes.

The best strategy, comments Harley, is to keep viral clearance needs in mind during development so that relevant steps can be included early in production. “Include viral clearance techniques early in development and work with an outsourcing partner to gain an understanding of the regulatory requirements for clearance studies and the time that it will take to generate clearance data,” he says.

CDMOs/CROs, adds Yoo, should also be agile, flexible, and ready to cope with any challenges. In addition, negotiation and collaboration among the manufacturer, suppliers, and third-party lab is the key to successful viral clearance, Chen notes.“Taking a case-by-case approach and working closely with all parties ensures that each aspect of the viral clearance strategy is tailored to meet the specific needs and challenges of the product, thereby enhancing its overall safety and efficacy,” he concludes.

Reference

1. FDA. Guidance for Industry: Q5A(R2) Viral Safety Evaluation of Biotechnology Products Derived From Cell Lines of Human or Animal Origin (Rockville, MD, January 2024).

About the author

Cynthia A. Challener, PhD, is a contributing editor to BioPharm International®.

Article details

BioPharm International®
Vol. 37, No. 10
November/December 2024
Pages: 18–22

Citation

When referring to this article, please cite it as Challener, C.A. Overcoming Viral Clearance Challenges. BioPharm International 2024 37 (10).