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Amid the COVID-19 pandemic, antibodies have been under the spotlight as potential therapeutics. Antibodies patrol the body and bind to their specific target region, known as an epitope, on an antigen. In humans, antibodies consist of 2 heavy chains and 2 light chains held together by disulfide bonds (see graphical abstract). However, as with most products of evolution, the structure of antibodies is not universal. Interestingly, camelids such as dromedary camels and alpacas produce antibodies that consist of only 2 heavy chains (see graphical abstract). Given the smaller size and unique shape of camelid antibodies, they can infiltrate into antigen’s cavities that are inaccessible to human antibodies. This feature of camelid antibodies piques the research interest because they may interact with a greater range of epitopes on a target protein.
Hong and colleagues were particularly interested in studying the translational potential of dromedary camel antibodies because these camels are natural reservoirs of the Middle East Respiratory Syndrome Coronavirus (MERS-CoV), a relative of SARS-CoV-2. Instead of using the full camel antibodies, their investigation homed in on the antigen-binding region of the camel antibody – the nanobody. The goal was to identify nanobodies that best inhibit the activity of spike protein by binding to angiotensin-converting enzyme 2 (ACE2) to prevent SARS-CoV-2 entry.
To identify the nanobody with a strong affinity for SARS-CoV-2 spike protein, the scientists utilized a technique known as phage display that allowed them to express camelid nanobody on bacteriophages. They isolated white blood cells from 6 camels and amplified the camel nanobody gene from these cells. These nanobody genes were then inserted into bacteriophages, which subsequently express the encoded protein on their surface. Using this technique, a vast library of more than 1011 different nanobody phages was created in this process! Though not all nanobodies were spike protein-specific, as the immune system encounters countless other pathogens in the environment. Therefore, the scientists screened for nanobody phages that bound most strongly to the spike protein, which led them to 6 candidates for further testing.
In response to the rapidly evolving SARS-CoV-2, Hong and colleagues tested their nanobody candidates against the numerous variants of concern at that time including the wild-type, alpha, beta, and gamma. They created fusion proteins consisting of the camelid nanobody and human Fc (hFc) fragment (see graphical abstract). Then nanobody-hFc cocktails were made from different combinations of the 6 nanobody candidates to determine the most effective combination. They discovered that the combination of two of the candidates showed the best neutralization against all 4 tested SARS-CoV-2 variants. This synergistic effect was a result of the two candidates binding strongly to two non-overlapping epitopes of the spike protein, thereby minimizing their physical interference with each other.
To translate these in vitro findings into living systems, a mouse model bearing human-like ACE2 receptors was used to test the nanobody-hFc efficacy. These mice were exposed to a lethal dose of either beta or delta variant. When these mice received the nanobody-hFc cocktail (consisting of the two selected candidates), they all survived the beta variant challenge and experienced no weight loss. However, the cocktail appeared less protective against the delta variant, resulting in 75% survival after a higher dose was administered, which is in every way better than 100% mortality!
Nevertheless, this investigation brought forth another valuable tool in our battle against SARS-CoV-2. It is merely incredible that the camels readily produce broadly neutralizing nanobodies against not only the wild-type SARS-CoV-2 but also its variants!
Source:
Article author: YongGuang Jiang. Yong is an IRTA postbaccalaureate fellow at the National Institute of Health. His research is focused on studying the role of plasmin and neutrophil elastase in hematopoietic recovery.
Editor: Dr. Sutonuka Bhar. Sutonuka is a scientist working on biotherapeutics development. She achieved her doctoral degree from University of Florida where her work focused on host immune responses against viruses and bacterial membrane vesicles.
Check out Antibuddies’ blog post “A Camel to the Rescue: Nanobodies against SARS-CoV-2″.
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