Showing an inspiring knack for innovation under pressure, the global scientific community has developed promising tests and treatments for COVID-19 in the span of just a few months. But moving medical technologies from conception to deployment at such an unprecedented rate comes with a multitude of hurdles, one of which is the obvious challenge of scale. How do companies turn a handful of prototypes or a few flasks of drug-secreting cells into a mass-produced product ready for market?
This is the primary focus for process engineer scientists at the Advanced Biofuels and Bioproducts Process Development Unit (ABPDU) at Lawrence Berkeley National Laboratory (Berkeley Lab). The ABPDU was founded to serve as a resource of equipment and expertise for companies or institutions that are using biological processes to generate exciting and potentially revolutionary products. Since opening their doors in 2012, the unit has collaborated with more than 62 groups to accelerate the development of sustainable biofuels, smart materials, innovative foods, and next-generation medicines.
Eager to apply their biomanufacturing know-how to the current crisis, ABPDU teams have been working during the shelter-in-place orders to help two companies scale-up production of new technologies that could aid in ending the pandemic.
Rapid, portable test kits using CRISPR
Although diagnostic testing for COVID-19 is becoming increasingly accessible, the test kits currently in use at hospitals, clinics, and walk-in centers around the world have many limitations. To detect the presence of coronavirus (SARS-CoV-2) from nasal and throat swabs, most of these kits have to be analyzed in centralized laboratories, with trained personnel and sophisticated equipment. Hoping to shift the paradigm, Argentina-based startup Caspr Biotech is developing a streamlined testing approach based on the increasingly powerful CRISPR gene editing system.
“The company has recently shown how well their system works, and its potential is pretty big,” said project lead and ABPDU principal investigator Eric Sundstrom. “The limiting factor for them at this time is the production of the CRISPR Cas-12 enzyme system. Our role here is to work in parallel with them and design an approach for scaled-up production of the protein, so that they have enough for more widespread clinical testing. Then, hopefully, we can help transfer the technology to a manufacturer who can produce it at the commercial scale.”
Like many biological products and medicines, Caspr’s proprietary Cas-12 enzyme is produced by specially engineered microbes through fermentation. The enzyme works as a detection system because it binds to a specific sequence of DNA – in this case it is engineered to target part of the SARS-CoV-2 genome – and then chops the DNA at that location. After cleaving the viral DNA, the Cas-12 enzyme will start to indiscriminately chop up any DNA that it encounters, so each test kit contains special DNA fragments that are bound with fluorescent proteins. This way, if the Cas-12 enzymes encounter the target viral DNA sequence in the blood sample added to the kit, the fluorescent molecules will be freed, and therefore activated, producing a light-based signal that indicates the individual is positive for a COVID-19 infection.
The company’s CRISPR Cas-12 enzyme is capable of seeking out very low levels of coronavirus DNA and has negligible rates of binding to other DNA sequences, according to as-of-yet unpublished evaluations of the system using samples collected from COVID-19 patients in the U.S. and other countries. These properties make the test highly sensitive and specific; meaning that the test results should have very few false negatives and no false positives. Furthermore, the kit analyzing equipment is small, portable, and can process a sample in less than 40 minutes. Current tests take several hours.
When Caspr approached the ABPDU for scale-up help in early April, they were already in the midst of validation testing (a step necessary for approval by regulatory agencies such as the Food and Drug Administration [FDA]) yet were only able to produce tiny batches of their enzyme at a time. “The company was at the smallest scale when they came to ABPDU, so working on this project has been an excellent opportunity for our team to rapidly dissect a challenge and build new processes,” said Laura Fernandez, a Berkeley Lab process engineer at the ABPDU.
Fernandez and senior process engineer Jan-Philip Prahl first scaled Caspr’s fermentation process up to their highly automated 250-milliliter bioreactor system, then successfully designed a fermentation protocol for 10-liter bioreactors. The insights generated by the 10-liter run will allow the company to further scale to commercial volumes, which are typically hundreds or thousands of liters. Two other team members are now developing a process for purifying the protein from the other molecules made by the fermenting microbes.
“We found that ABPDU is the ideal partner for the optimization of the fermentation at the initial scale of 10 liters and for helping us with the tech transfer process to larger industrial players to continue the production,” said Caspr co-founder and CEO Franco Goytia. “We are submitting to the FDA for Emergency Use Authorization in the next few weeks, while in parallel scaling up our production capacity to hopefully make our solution widely available in the U.S. and the rest of the world in the second half of 2020.”
A faster approach for antibody production
The co-founder and CEO of Swiftscale Biologics, a San Francisco-based startup, reached out to the ABPDU in early March for help scaling up their rapid neutralizing antibody development and manufacturing platform. Neutralizing antibodies are proteins produced by immune cells that allow the body to detect and combat pathogens. They work by binding to specific features on the outside of virus particles, thereby inhibiting the virus’ ability to infect cells or tagging it for destruction by specialized engulfing immune cells.
Antibody infusions – of either survivor-donated blood plasma or antibodies produced through biomanufacturing – are frequently used to treat infectious diseases in cases where the patient’s immune system is struggling, and evidence from across the world shows that donated plasma is currently the most effective treatment for COVID-19. But unfortunately, there is not nearly enough donated plasma to treat all patients in need, and lab-grown copies of SARS-CoV-2 antibodies are still being developed and tested in clinical trials.
“Traditional manufacturing processes for biologics are too time-consuming to meet the needs of the current outbreak,” said Swiftscale CEO David Mace. “To meet this urgent need, Swiftscale is building a platform that can produce these life-saving therapies in a fraction of that time. We can then leverage this platform to accelerate the work of our partners who are developing promising biologics.”
Akash Narani, principal process engineer at ABPDU and lead on the team collaborating with Swiftscale, explained that the company is using a “cell-free” approach that expedites the screening and development of antibodies and then leverages specially engineered microbes that can produce active therapeutic antibodies with previously unachievable productivity. This approach is less costly and time-consuming to develop and scale compared with the standard way to produce antibodies for human medicines and diagnostics, which relies on cultured mammalian cells.
“Swiftscale is envisioning a new way to develop and manufacture biologics. Cell-free technology is still relatively new, but if done right, they can lead to rapid prototyping [identification of the most effective antibodies] and rapid production,” said Narani.
Narani and his colleagues began hands-on work at the ABPDU on May 18. Their role is to design and test an efficient process to take Swiftscale’s 250-mililiter fermentations, performed in collaboration with Culture Biosciences, up to 300 liters – a 1,000-fold increase in production volume. Adding to the challenge, the company is still continually optimizing the parts of the production process that come before and after the fermentation stage.
“The scale-up production is a crucial step in the research process and I’m glad we could contribute to such important work and that our efforts are helping research to move quickly,” said Carolina Araujo-Barcelos, an ABPDU fermentation researcher.
Ethan Oksen, a senior research associate, added: “We hope that Swiftscale can apply the insights and data we generated to move quickly to production scale, where they can generate neutralizing antibodies for clinical trials.”
This work also included the efforts of Berkeley Lab researchers Gregory Bontemps, Shawn Chang, Chang Dou, Asun Oka, Isaac Wolf, and Jipeng Yan. The COVID-19 research performed at the ABPDU is supported by Caspr Biotech and Swiftscale Biologics. ABPDU is funded by the Department of Energy’s Bioenergy Technologies Office (BETO).