By Kurt Brorson and Mo Heidaran, Parexel Regulatory & Access
Coronavirus COVID-19 has been all over the news recently. This virus is part of a larger family called Coronaviridae, which includes viruses that can infect only animals, cause mild colds or be communicable and pathogenic like the SARS virus of the early 2000’s. COVID-19 seems to be more on the SARS end of this scale, causing life-threatening respiratory infections.
While the main concern for the public now is the spread of human infections, other concerns stem from the spread- for example what about the virus safety of biologics intended for human use? Can this virus get into the medicinal supply, like HIV and hepatitis virus did in the 1980’s? We will argue below that the risk is extremely low, but not nonexistent. Steps can be implemented to mitigate the risk. Keep in mind that the biologics industry includes conventional biotechnology products like monoclonal antibodies and recombinant therapeutic proteins, vaccines against a plethora of bacterial and viral targets, plasma derived products and a whole new class of cell and gene therapies. Each of these have unique considerations from a viral safety standpoint.
Monoclonal antibodies and recombinant therapeutic protein manufacturing have two main strategies to assure virus safety: source material testing and process clearance.
A large segment of the biotech products are grown and produced in mammalian cell culture, most notably CHO cells. CHO cells in bioreactor culture have been found over the years to be capable of being infected by viruses and propagating them. This has been manifested by occasional facility contaminations by Mouse Minute Virus (MMV), a nuisance parvovirus probably carried into the process train via raw materials. These contaminations have been detected by impacts on cell culture and routine testing post-culture. They have been economically catastrophic, but without patient impact so far as they were detected in time.
Can Coronaviruses propagate in CHO cells? The science here is mixed.
A 2003 US Patent issued to the Schering-Plough Veterinary Corporation appeared to assert that Bovine Coronavirus (BCV) can grow in CHO-K1 cells, largely when the media is supplemented with bovine serum. In contrast, a 2010 Biotechnology and Bioengineering manuscript from Baxter Biosciences reported that they were unable to propagate Mouse Hepatitis Virus, another Coronavirus, in CHO cell culture. How well these two distantly related Coronaviruses predict the behavior of COVID-19 in the rigorous serum-free environment of a bioreactor culture is unknown. However, they indicate that CHO can propagate at least some types of Coronavirus under the right circumstances.
How could a Coronavirus get into the culture in the first place?
Cell banks are produced months to years in advance and screened rigorously for viruses. Thus, they are an unlikely source for Coronavirus as they have existed before the recent COVID-19 outbreak and have been screened. In the biotech industry, there has been wide implementation of barrier technology like HTST, irradiation and barrier virus filters for preparation of culture raw materials. These measures have been effective for control of other nuisance viruses, such as Cache Valley Virus. There is no reason to believe that they would not be effective barriers to COVID-19 as well.
The only remaining vulnerable point of entry could be pre-culture manipulations when shake flasks, etc. are opened under an LAF. Being a respiratory virus, carelessness here by operators could pose a Coronavirus risk. Use of gloves, facemasks and other gowning during these sensitive manipulations of open culture, as well as a regulatory prohibition on personnel with symptoms of illness from entering facilities are a prudent risk mitigations already in place in biotech facilities.
Harvest testing for cytopathogenic effects (CPEs) on a panel of indicator cells, including the production cell type, is an added safety measure for bioprocessing. Although the reactivity of Coronaviruses in the in vitro adventitious agent (AVA) test has not been systematically studies, the 2003 US patent cited above appears to report that BCV will cause both CPEs in CHO cells and Hemagglutination. This argues that the standard AVA test may be effective for picking up Coronavirus contaminations, if they include at least CHO cells.
Process clearance of Coronaviruses during biotechnology manufacture is likely to be very robust.
Clearance evaluation of smaller and more hardy viruses like parvoviruses is an industry standard practice, as outlined in ICH Q5A (1998). A typical bioprocess can remove 9-10+ log10 of hardy and small viruses like Parvovirus, and several log10 more of larger enveloped viruses like murine retrovirus. Coronaviruses are ss (+) strand RNA viruses that are enveloped and relatively large (100-160 nm, depending on the type). The large size assures complete clearance by both small and large virus retentive filters that are standard in bioprocessing. Some biotech manufacturing schemes include detergent inactivation steps, which would likely dissolve their membranes. Some Coronaviruses are low pH resistant down to 3.0, so the low pH inactivation steps employed in monoclonal antibody manufacturing should not be assumed to be effective.
In summary, procedures mandated by ICH Q5A (1998) assure the safety of biotech products with respect to Coronavirus. Propagation of COVID-19 in CHO cell bioreactor cultures is unlikely but not impossible based on experience with other Coronaviruses. However, they are very unlikely to get introduced in the first place. Routine harvest testing is in place should detect contamination. If present, Coronavirus would be completely removed from the product stream by virus filtration and other steps if they were.
Plasma derived products also rely on rigorous testing and validated clearance of viruses.
For these products, the panels of viruses used for clearance validation studies are even larger than those employed for biotech products. Coronaviruses target endothelial cells and they tend to cause mostly respiratory, GI or neuronal infections in patients. However, this does not mean that blood accumulation of COVID-19 is impossible as some Coronaviruses also infect blood cells like macrophages. However, process clearance of Coronaviruses in this context is also expected to be robust. Most plasma products include a solvent/detergent inactivation step which is highly effective for inactivation of lipid enveloped viruses like Coronaviruses. Plasma derived product processing also usually includes virus retentive filters, as described above likely to remove all Coronaviruses given their size. Thus, it is highly likely that plasma derived products that use these two steps have negligible risk for Coronavirus.
Cell therapy and gene therapy products are produced using widely different schemes, not always with extensive downstream processing that can inactivate or remove viruses.
In addition, it is virtually impossible in some cases to subject cell and gene therapy drug products to viral inactivation steps as the process of inactivation has shown to adversely impact the product quality. Similarly, virus filters can remove the actual product and thus can’t be used in this context. For these reasons, the emphasis to control spread of communicable diseases for cell therapy product including allogeneic CAR T products are placed on establishing Donor Eligibility (DE) for Human cells, Tissues, and Cellular and Tissue-Based Products (HCTPs) that is collected from allogeneic donors.
Currently, DE requirements according to 21 CFR 1271 subpart C does not include testing of donors for Coronavirus but FDA has been very proactive in changing the requirements as the public is confronted with new infectious agents. For example, recently the agency issued several guidance documents in which screening for Zika Virus was added to the list of Relevant Communicable Disease Agents and Diseases (RCDAD) for HCTPs and blood derived product must now be tested for Zika virus according to 21 CFR 610.40.
The question is if Coronavirus poses a significant risk of disease transmission when products are derived from allogeneic sources or cultured and manipulated in the presence of human derived components. The answer to these questions remains to be answered. However, in view of the complexity of these products the risk of disease transmission for Coronaviruses will need to be evaluated on a case-specific approach using a risk and science-based approach, including specific testing as described below.
Vaccines are diverse and will require a case-specific approach.
Some vaccines are produced in bacterial systems, so there is no virus risk for these. Certain viral vaccines on the other hand are often propagated in mammalian cells like Vero et al. which are susceptible to infection by other viruses. Extensive testing is a key component for the viral safety of these products. As discussed above, one should not assume that Coronaviruses won’t replicate in mammalian production cells, especially in complex media with serum and other biological molecules. Testing, for example by a sensitive PCR test for Coronaviruses, would add a needed level of safety assurance in this context. In our experience, close collaboration with subject matter experts, as well as contract testing labs, is key for robust viral safety strategies, no matter the product category.
Parexel’s consulting group can provide our clients with advice when developing comprehensive virus risk mitigation strategies for biopharmaceutical and biologics development. Our recommendations are based on a deep knowledge and experience in viral clearance in bioprocessing. As mentioned above, regulators and the biologics industry has been confronted with and developed risk mitigation strategies against emerging viruses in the past; some maybe even more scary like Zika virus which causes developmental abnormalities in kids of mothers who were infected. We think that while COVID-19 may a theoretical risk in the context of biologics manufacturing and control, it is very low because of standards and practices already in place as described above. Parexel stands ready to assist companies to develop tailored virus risk-mitigation strategies based on deep expertise and that consider a number of criteria including:
- What class is the product and are platform technologies available?
- What is the stage of product development (e.g. BLA vs. IND)?
- What specific unit operations will be used downstream? What cell substrates will be used for production?
- Finally, which global regions are targeted for product introduction?