Colette Côté, PhD - Chief Portfolio Manager at PathoQuest and General Manager of the U.S. subsidiary
Audrey Brussel, PhD - Viral Safety Leader at PathoQuest and company representative to the International Consortium on Adventitious Agent Contamination in Biomanufacturing (CAACB)
It is striking just how much the past year has dramatically changed our lives. Today we casually talk about our life experiences as ‘pre-COVID’ or ‘post-COVID’. Mass availability of multiple, effective vaccines against the SARS-CoV-2 novel coronavirus (COVID-19) has been met with both excitement and trepidation. For many, it has brought about a general sense of relief that we are finally one step closer to a return to normalcy after an undeserved and rather abrupt disruption to our lives. For others, it has led to hesitation or even outright denial as to the safety and efficacy of such a rapidly engineered vaccine. Indeed, this global pandemic has been devastating, but it has also brought about a much-needed paradigm shift to the world. It has
fundamentally and dramatically changed the way vaccines are designed, developed, and manufactured. The rapid pace of COVID-19 vaccine development and the drive for Emergency Use Authorization (EUA) by the regulatory agencies has been unprecedented. But it has also raised questions around the long-term sustainability of the process and, more importantly, the overall patient and product safety of such a vaccine.
COVID-19: A Needed Kick in the Pants in the Evolution of Biosafety Tests and Guidelines
COVID-19 challenged the biopharmaceutical industry to accelerate development timelines without compromising biosafety. They had to accelerate their own innovation, and research and development to bring a solution to patients. Critically, biopharma had to pay extra attention to ensure the manufacturing process and their final released products remained free from adventitious contamination from COVID-19, its variants, or any other pathogens.
Despite technological advancements in the design, development, and manufacturing processes surrounding biologics,1 current biosafety testing strategies, the technologies they are dependent on, and the framework of regulatory requirements in place to ensure the safety of medicinal products all remain in a seemingly archaic state.
The biosafety testing industry as we know it continues to be driven by the same collection of aging guidance documents that have often remained untouched for decades (e.g. ICHQ5 A (R1)).2 Historically, change to these documents and their prescribed processes has come with a strong degree of reluctance. The adage of ‘if it ain’t broke, don’t fix it’ is seemingly echoed by the industry and to some degree the agencies. Not surprisingly then, change occurs at a glacial pace, with rounds of public opinion gathering before finalization and implementation of the change only a year or more later. During that time, technologies continue to evolve, further broadening the gap between rapid technological advancement and the slow pace in the administration of the guidance. This gap will no doubt continue to grow unless there is another profound paradigm shift in safety testing. Until then, accelerating the introduction and implementation of novel technologies and testing approaches, while maintaining the spirit of existing, albeit aging regulatory guidance remains a challenge. Fortunately, this is where the rapid development and the EUA of COVID-19 vaccines have been a major driving force and a boon for progress.3
Traditional biosafety testing encompasses a varied collection of methodologies. These include classical in vivo and in vitro testing along with widely embraced molecular approaches such as the broad use of the polymerase chain reaction (PCR), among others.4 While quite powerful in their early days of implementation, both in vivo and in vitro tests have become increasingly perceived as time-consuming or insufficiently sensitive or specific enough for clear identification of any offending agents.5 For example, both in vivo and in vitro tests may be limited by their susceptibility to the agent in question and may not detect every possible contamination. If and when there is a sign of infection, those tests simply indicate that a contaminant may be present without providing clear identification due to the limitations of the tests themselves. In addition, they are not specific of a contamination by viruses, leading to false positives. Furthermore, animal-based tests continue to fall out of favor with a desire for more ethical testing strategies that accommodate the 3Rs: replace, reduce and refine.6 Despite these drawbacks, both in vivo and in vitro assays remain the workhorses of the industry and at the core of many guidance documents and agency recommendations. Likewise, while standard molecular endpoint tests like PCR achieve greater specificity and sensitivity, they are limited in the ability to define if an agent is live and replicating or simply represents residual nucleic acids of no concern. Ultimately it is the collective application of these in vitro and in vivo tests that enables a clearer picture as to the overall safety of the material and its final release into the market. Unfortunately, this multi-testing approach is cumbersome and time-consuming, and not compatible with fast-tracking development timelines.
The rush to bring a COVID-19 vaccine to market has been both a success story in effectiveness and a refreshing change to traditional biosafety thinking. A several month-long in vivo tests suddenly became a roadblock to accelerating the release of this specific product with billions of patients anxiously waiting. Even a traditional 28-day in vitro assay model, while more amenable than an in vivo test in the context of the 3Rs, ultimately was viewed as an impediment to rapid deployment of the product. This is true not only for healthy patients in search of prophylactic treatments like a COVID-19 vaccine, but also for critically or terminally ill patients seeking life-saving treatments envisioned by the cell and gene therapy industry. The challenge then was to find alternative, state-of-the-art testing strategies that not only fit well within the guidance requirements and recommendations but were also more sensitive, specific, and reflective of the state of the test material, thereby increasing the confidence in the safety of the product.
Next-Generation Sequencing-Based Tests are Growing in Popularity Among Biopharmas and Regulatory Agencies
The risk of not taking this call to action was too great that even the regulatory agencies recognized the need to think and act differently, while continuing to acknowledge the unique risks alternative testing strategies could present. For example, the use of Next-Generation Sequencing (NGS) as a supplement, and to some degree a replacement test, for classical in vivo or in vitro tests to detect and identify contaminants in a product suddenly transitions into an intriguing new reality. The idea here is not necessarily to skip traditional testing completely, but rather execute an alternative test method in the short term to not only fast-track release of the product, but also minimize the risk to the patient during this critical time. In the COVID-19 scenario, the ability to shave even a few days off testing and accelerate safe product release no doubt becomes a critical life-saving moment for many.
Historically, technologies like NGS were often relegated to early or upstream product development, settling comfortably in the research and development space due to its perceived novelty, the fear of discovering something unknown, and/or the inherently higher cost for applications. In reality, NGS has proven itself to be an ideal tool for expanded application in our new COVID-19 world. In its quiet decades-long history, it has been used for a wide range of applications in both the clinical and biopharma space. The most obvious application related to COVID-19 includes tracing divergence, at the individual nucleotide level, of the circulating SARS-CoV-2 strains in patients worldwide, enabling exquisite surveillance of potentially immune evading strains. But the same strategy can be also applied to screening medicinal products, including engineered COVID-19 vaccines themselves, for inadvertent contamination with infectious COVID-19, during the manufacturing process, by infected and potentially asymptomatic operators.
With the current state of the methodology, it is possible to rapidly, within a couple of weeks or less, agnostically screen samples by NGS for even low levels of a contaminating virus, not just COVID-19. Not only does this methodology significantly reduce the time to result when compared to traditional tests, but also increases the confidence of the result due to the extraordinary depth of analysis that can be achieved in short order. Current NGS methodologies also enable deeper insight into the replicative nature or biological context of a contaminant and are not just limited to revealing its presence based on a simple sequence signature. The range of possibilities for NGS in the biosafety testing space continues to broaden, with new applications being envisioned daily. COVID-19 has shown that NGS can quite easily and rapidly meet the expectations for quality control through the development and manufacturing processes.
Questions Raised and Challenges for Biomanufacturers
Soon after the COVID-19 outbreak, researchers started to describe the large tropism of SARS-CoV-2 capable of replicating in different cell types such as those in the lungs and brain, with life-threatening implications for humans.7 In the industry, manufacturers wondered if SARS-CoV-2 could replicate in cell lines used in bioproduction processes and be easily detected by the current toolbox of biosafety tests. They started looking for solutions to screen for SARS-CoV-2 at any step of the process: from research and master cell banks to downstream material and processes. Biomanufacturers needed rapid and robust testing solutions – beyond what is expected from regulatory guidelines – to mitigate the global industrial risk of unintended COVID-19 contaminations.
In this context, the agnostic nature of NGS-based testing is key, allowing for the detection of any pathogen without prior knowledge of what to look for. The growing use of NGS in the industry drives this proactive attitude in pathogen detection – instead of a reactive approach as witnessed with COVID-19. Moreover, the NGS approach is also key to detect new variants that might otherwise escape detection by more traditional tests.
Streamlining Biosafety Testing: More Safety, Less Constraints
The lessons learned from the COVID-19 pandemic highlight a new trend in biosafety and place NGS in a prime position now and in the future. Of the many advantages of the technology, two rise above in importance: its ability to significantly reduce development timelines of innovative medicines, and its agnostic approach to pathogen detection, even emerging ones.
If another pandemic were to strike, one hopes our lessons learned with COVID-19 better positions the industry to rapidly deploy the necessary resources and strategies to ensure a rapid and safe solution. After a health crisis such as COVID-19, the list of mandatory or precautionary measures regarding biosafety issued by agencies will likely grow, but hopefully not become more cumbersome. Staying abreast of these changes, whether proposed or actual, is critical. Today biomanufacturers have rightfully become more demanding when choosing their suppliers and partners, specifically when it comes to ensuring the prevention and control of contamination in their processes and products, regardless of the source. In this light, NGS maintains an advantage by serving as an ideal alternative technology, demonstrating a vast breadth and depth of performance, unrivaled by traditional testing strategies. It is clear that COVID-19 has paved a necessary path to further streamline biosafety testing strategies, and in doing so has enabled far greater safety with much less constraint. In this context, COVID-19 has become a necessary evil - it became a proving ground for not just NGS but other evolving technologies as well. Ultimately though, what is needed for any technology is a comprehensive evaluation as to the time and costs savings, the animals spared, and the robustness and reliability of the approach to ensure it is and remains a ‘best-in-class’ technology.
Final Thoughts
What is really exciting about NGS is we have only scratched the surface of its breadth of application, with intriguing new possibilities for use ahead of us. The implementation of NGS in the biosafety testing space has been rather a niche thus far, with slightly more than a decade of use in this field. Slowly but surely, it has and continues to gain wider acceptance, particularly as science and bioinformatics are tweaked for specific applications. Viral safety testing is a mere glimpse into its use and overall value in a biopharmaceutical setting. In truth, NGS is a twenty-year-old technology that the industry has finally awakened to, but we are barely scratching the surface when it comes to its use. NGS is innovation, opportunity, and the future. The impact that COVID-19 has on our lives will fade over time. The hope is we will not have to wait for another health crisis to unleash the full potential of this and other like technologies. Patients’ lives are in the line.
References
- Salgueiro S, Bazhenova A. Embracing Innovation in Biomanufacturing. BioProcess Interrnational. Accessed March 9, 2021.(https://bioprocessintl.com/manufacturing/single- use/embracing-innovation-in-bioprocessing-and-biomanufacturing/)
- Quality of Biotechnological Products: Viral Safety Evaluation of Biotechnology Products Derived from Cell Lines of Human or Animal Origin. ICH, Q5 (R1). Step 5 version. October, 1997. https://www.ema.europa.eu/en/documents/scientific-guideline/ich-q-5-r1-viral- safety-evaluation-biotechnology-products-derived-cell-lines-human-animal-origin_en.pdf
- Viral Safety Evaluation of Biotechnology Products Derived from Cell Lines of Human or Animal Origin. ICH, Q5A(R2). Final Concept Paper. November 18, 2019. (https://database. ich.org/sites/default/files/Q5A-R2_FinalConceptPaper_2019_1117.pdf )
- Viral Safety Evaluation of Biotechnology Products Derived from Cell Lines of Human or Animal Origin. ICH, Q5A (R1) Step 4 version. September 23, 1999. (https://database.ich.org/ sites/default/files/Q5A_R1_Guideline.pdf )
- Gombold J, Karakasidis S, Niksa P, et al. Systematic evaluation of in vitro and in vivo adventitious virus assays for the detection of viral contamination of cell banks and biological products. Vaccine. 2014;32(24):2916-2926. doi:10.1016/j.vaccine.2014.02.021 (https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4526145/)
- Legislation for the protection of animals used for scientific purposes. Directive 2010/63/ EU as amended by Regulation (EU) 2019/1010. European Commission Web site. https:// ec.europa.eu/environment/chemicals/lab_animals/legislation_en.htm. Accessed May 12, 2021.
- Liu J, Li Y, Liu Q, et al. SARS-CoV-2 cell tropism and multiorgan infection. Cell Discov. 2021;7(1):17. Published 2021 Mar 23. doi:10.1038/s41421-021-00249-2 https://www.nature.com/articles/s41421-021-00249-2
Colette Côté, PhD, is Chief Portfolio Manager at PathoQuest and General Manager of the U.S. subsidiary. Prior to joining PathoQuest, Dr. Côté was Director and Head of NGS R&D Testing Services at MilliporeSigma where she developed a wide range of novel NGS applications working closely with key stakeholders in the industry. Her work has been pivotal to the advancement of the use of NGS technology in this field. Dr. Côté received her Ph.D. in Molecular Biology, Cell Biology, and Biochemistry from Brown University and was a post-doctoral fellow at the NIH.
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