Tiny immune proteins known as nanobodies may encumber the SAR-CoV-2 virus, preventing it from latching onto host cells. This exciting possibility has been demonstrated by several research groups, almost exclusively in studies using in vitro assays. A new study, however, has shown that nanobodies may prevent COVID-19 in vivo. Moreover, the new study reports on the efficacy of nanobody cocktails.

The new study was led by Wai-Hong Tham, PhD, an associate professor of infectious diseases at the Walter and Eliza Hall Institute (WEHI) in Australia. Tham and her WEHI team, in collaboration with scientists at the Doherty Institute and the Kirby Institute, found that nanobody cocktails consisting of two noncompeting nanobodies were able to block angiotensin-converting enzyme 2 (ACE2) engagement with receptor-binding domain (RBD) variants present in human populations. The cocktails potently neutralized both wild-type SARS-CoV-2 and the N501Y D614G variant at low concentrations.

Details appeared online in the Proceedings of the National Academy of Sciences (PNAS), in an article titled, “Nanobody cocktails potently neutralize SARS-CoV-2 D614G N501Y variant and protect mice.”

“Epitope mapping, X-ray crystallography, and cryo-electron microscopy revealed two distinct antigenic sites and showed two neutralizing nanobodies from different epitope classes bound simultaneously to the spike trimer,” the article’s authors wrote. “Prophylactic administration of either single nanobody-Fc or as mixtures reduced viral loads by up to 104-fold in mice infected with the N501Y D614G SARS-CoV-2 virus.”

Essentially, by mapping nanobodies, the scientists were able to identify a nanobody that recognized the SARS-CoV-2 virus, including emerging global variants of concern. The nanobody also recognized the original SARS-CoV virus (which causes SARS), indicating it may provide cross-protection against these two human coronaviruses. These results, the authors concluded, indicate that nanobodies could provide an alternative to conventional antibody treatments for COVID-19.

Antibody-based therapies harness the ability of antibodies to bind other proteins—that is, disease-associated proteins—tightly and specifically. Diminutive antibodies known as nanobodies are produced naturally by alpacas in response to infection.

As part of the current research, WEHI researchers oversaw the immunization of a group of alpacas in regional Victoria with a synthetic, noninfectious part of the SARS-CoV-2 “spike” protein.

“The synthetic spike protein is not infectious and does not cause the alpacas to develop disease,” said Tham. “But it allows the alpacas to develop nanobodies. We can then extract the gene sequences encoding the nanobodies and use this to produce millions of types of nanobodies in the laboratory, and then select the ones that best bind to the spike protein.”

The leading nanobodies that block virus entry were then combined into a “nanobody cocktail.”

“By combining the two leading nanobodies into this nanobody cocktail,” Tham explained, “we were able to test its effectiveness at blocking SARS-CoV-2 from entering cells and reducing viral loads in preclinical models.”

By mapping the nanobodies, the research team was able to identify a nanobody that recognized the SARS-CoV-2 virus, including emerging global variants of concern. The nanobody was also effective against the original SARS virus (SARS-CoV), indicating it may provide cross-protection against these two globally significant human coronaviruses.

“In the wake of COVID-19, there is a lot of discussion about pandemic preparedness,” Tham pointed out. “Nanobodies that are able to bind to other human beta-coronaviruses—including SARS-CoV-2, SARS-CoV, and MERS—could prove effective against future coronaviruses as well.”