Infectious diseases can wreck the human body in lots of different ways. Fortunately for most, the immune system comes out swinging — custom making antibodies to identify and neutralize foreign threats. Now, scientists at the biotechnology company GigaGen are leveraging the immune system’s capabilities to create next-generation antibody drugs: recombinant polyclonal antibodies for viral diseases such as hepatitis B and COVID-19.
How the human body responds to infection fascinates GigaGen chief executive officer Carter Keller. “The immune system is really amazing,” he said. “It creates this rapid response to some infection or foreign element, hitting that element hard and rapidly from all different angles.”
To leverage the power of the immune system against viral threats, Keller and the GigaGen team decided to go after a specific type of antibody: polyclonal antibodies, which are a mixture of different antibodies produced by B cells (1). Unlike monoclonal antibodies, which can identify and bind to one marker or epitope on a foreign agent like a virus, polyclonal antibodies can bind to an array of epitopes on the same infectious agent.
According to David Smith, a virologist at the University of California, San Diego who is not affiliated with GigaGen, polyclonal antibodies offer a potential treatment security blanket in the case of mutation. “Let’s say a pathogen has evolved one of its epitopes to make it not susceptible to the antibody,” he said. “If you still had three or four hanging out there, they would have a better chance of neutralizing the pathogen versus a monoclonal, where you’ve put all your eggs in one basket.”
GigaGen’s approach to making antibodies relies on a recombinant strategy. They generate the antibodies in a laboratory from gene sequences, cutting away the need for animal or human donors as the antibody production factories. By combining thousands of these antibody gene sequences and expressing them in cells, they created a polyclonal mixture of antibodies. “Rather than just taking one part of that immune response and turning it into a monoclonal antibody or small molecule, we made a technology that captures the entirety of the antibody immune response and recreated it,” said Keller.
Targeting hepatitis B
While a vaccine to prevent hepatitis B infection is available, there is no cure for those who are already infected, and chronic infection can lead to liver damage or death (2,3). To create a polyclonal antibody therapy for hepatitis B, the researchers at GigaGen vaccinated people with the hepatitis B vaccine. This allowed the participants to naturally generate a strong immune response against the virus, after which they donated several milliliters of blood. The scientists then used a microfluidic droplet system to isolate individual B cells from the blood samples. Within each B cell, the team used polymerase chain reaction to isolate the sequences for the heavy and light chains of the antibody. After adding some additional sequences to the antibody to prepare it for gene expression, they put that sequence into a plasmid that they could then use for large-scale antibody manufacturing.
In large-scale monoclonal antibody production, batch variability is a big problem. One of the advantages of manufacturing a polyclonal antibody mixture is that because there is so much existing diversity in the antibody pool, batch variability due to production differences decreases.
Having a plasmid that encodes the antibody opened the door to even more possibilities. According to Sheila Keating, GigaGen’s vice president of immunology, GigaGen researchers can transfect those plasmids into yeast cells to generate a library of antibodies. “Once the yeast is expressing the antibody sequences, we can stain those yeast for fluorescently labelled antigen. So, for hepatitis B, we would use hepatitis B antigen,” she explained. “Then we would identify each antibody that was specifically binding to the surface antigens.”
This method allowed the scientists to select for the antibodies with the highest binding potency to hepatitis B viral antigens, essentially creating a super-strong pool of antibodies. “Putting it into yeast is really a powerful step,” added Keller. “It allows us to have this immortal library of antibodies that we can do a lot with, and to figure out what subsets of antibodies we want to include in our final polyclonal mixture.”
In the end, the GigaGen team included around 2,000 polyclonal antibodies in their mixture against hepatitis B. “We know that we have captured the diversity of many different samples to create that product,” Keller said.
To test if the polyclonal antibody mixture actually bound to and inhibited the virus, the team asked how well the antibodies bound to liver cells that presented the receptor for hepatitis B. They also used antibody-binding assays to test the mixture’s binding potency to the hepatitis B antigen. According to Keller, the polyclonal antibody mixture bound to hepatitis B antigen approximately 1,000 times more potently than unmodified plasma isolated from people vaccinated with the hepatitis B vaccine. The process of enriching for those super strong antibodies produced a more effective binding response.
In a mouse model of hepatitis B infection, Keating noted that the polyclonal antibody mixture neutralized and cleared the viral DNA along with its antigens. She is interested to see how the immune system itself reacts to this. “The secondary mechanism of action of any antibody that binds to hepatitis B antigen is to induce an immune response against the virus,” she said. The purpose of the immune response would then be to clear out the infected cells that are tagged by the antibody, allowing the patient to achieve functional cure where viral replication is suppressed enough so that it doesn’t cause active disease.
Those immune system reactions are best seen in the clinic in real patients. GigaGen aims to submit an Investigational New Drug filing and to initiate clinical studies beginning next year.
Looking at other viral diseases
Besides hepatitis B, the GigaGen team also applied their polyclonal antibody technology to other diseases, namely COVID-19. For COVID-19, the team used blood isolated from patients who had already recovered from the virus, and then they enriched for those antibodies that bound most strongly to the virus. As with hepatitis B, they used yeast libraries to select the best antibodies. The final polyclonal antibody mixture, which contained around 12,500 antibodies, neutralized SARS-CoV-2 at a rate more than 200 times higher than the patient’s unenriched plasma (4).
Having so many antibodies in the mixture proved beneficial, especially as SARS-CoV-2 rapidly mutated. “We found that we could neutralize multiple variants,” said Keating. “Having over 12,000 antibodies targeting the virus has allowed us to do that.” She is interested in seeing if these antibody mixtures have any utility in treating COVID-19 and long COVID in particular, given the millions of patients in the US alone who need treatments.
The hepatitis B space hasn’t had any new drugs in a long time.
- Sheila Keating, GigaGen
The team has also generated antibody mixtures against Zika virus and other respiratory pathogens (4). They said that their yeast plasmid library-based manufacturing approach can target many diseases, making it easily applicable with a quick change in initial blood donor.
Smith concurs. “We’ve known how to make monoclonal and polyclonal antibodies for a long time,” he said. “It’s all about finding the right cells that have made it so that we can extract it out. And the technologies are getting a lot better.”
For now, though, the GigaGen team is buckling down on their hepatitis B antibody therapy. “The hepatitis B space hasn’t had any new drugs in a long time,” Keating said. “The new drugs that are getting closer have not demonstrated clearance of antigen, and we want to be part of that new wave of drugs with a completely new mechanism of action.”
References
- Struble, E. B., Rawson, J. M. O., Stantchev, T., Scott, D. & Shapiro, M. A. Uses and Challenges of Antiviral Polyclonal and Monoclonal Antibody Therapies. Pharmaceutics 15, 1538 (2023).
- Hodgens, A. & Marathi, R. Hepatitis B Vaccine, in StatPearls (StatPearls Publishing, 2023).
- Iannacone, M. & Guidotti, L. G. Immunobiology and pathogenesis of hepatitis B virus infection. Nat Rev Immunol 22, 19–32 (2022).
- Keating, S. M. et al. Generation of recombinant hyperimmune globulins from diverse B-cell repertoires. Nat Biotechnol 39, 989–999 (2021).
