Nanobodies, or single-domain antibodies, are smaller and simpler than conventional antibodies, but they have huge potential. Since this potential is being explored by multiple research teams, any one team has difficulty producing standout work. Yet researchers from two Göttingen institutions, the Max Planck Institute (MPI) for Biophysical Chemistry and the University Medical Center, assert that they have developed nanobodies that are truly special.

These nanobodies can block the SARS-CoV-2 virus, but that’s not especially distinctive. Nanobodies developed by other researchers can do that, too. What sets the nanobodies from Göttingen apart, the Göttingen researchers pointed out, is that they can bind and neutralize the virus up to 1,000 times better than previously developed nanobodies. The Göttingen nanobodies can also, in the researchers’ words, “tolerate the K417N/T, E484K, N501Y, and L452R immune-escape mutations found in the Alpha, Beta, Gamma, Epsilon, Iota, and Delta/Kappa lineages.”

And that’s not all. The Göttingen researchers reported that they have optimized their nanobodies for stability and resistance to extreme heat. These qualities, together with the ease with which nanobodies can be manufactured, make the Göttingen nanobodies especially attractive as a way to counter COVID-19. Treatments for COVID-19 are needed in vast quantities. In addition, these treatments often need to tolerate less-than-ideal transport and storage conditions.

Finally, the Göttingen researchers have discovered that their antibodies enforce native folding of the SARS-CoV-2 Spike protein’s receptor-binding domain (RBD) in the Eshcerichia coli cytosol, where RBD folding normally fails. “Such fold-promoting nanobodies,” the Göttingen researchers suggest, “may allow for simplified production of vaccines and their adaptation to viral escape-mutations.”

All of these advantages were reported in The EMBO Journal, in a paper titled, “Neutralization of SARS-CoV-2 by highly potent, hyperthermostable, and mutation-tolerant nanobodies.” The paper described how 45 SARS-CoV-2-blocking nanobodies, or VHH antibodies, were isolated from alpaca immune libraries. It also describes how these monomeric species were used to construct nanobody tandems and trimers.

“The most effective VHH antibody neutralizes SARS-CoV-2 at 17–50 pM concentration (0.2–0.7 µg/L), binds the open and closed states of the Spike, and shows a tight RBD interaction in the X-ray and cryo-EM structures,” the paper’s authors detailed. “The best VHH trimers neutralize even at 40 ng/L.”

The figure shows how two of the newly developed nanobodies (blue and magenta) bind to the receptor-binding domain (green) of the coronavirus spike protein (grey), thus preventing infection with SARS-CoV-2 and its variants. The nanobodies originate from alpacas and are smaller and simpler than conventional antibodies. [Max Planck Institute for Biophysical Chemistry]
One of the paper’s co-corresponding authors, Dirk Görlich, PhD, director at the MPI for biophysical chemistry, said that the nanobodies “combine extreme stability and outstanding efficacy against the virus and its Alpha, Beta, Gamma, and Delta mutants.” The paper’s other co-corresponding author, Matthias Dobbelstein, MD, professor and director of the UMG’s Institute of Molecular Oncology, added that the nanobodies can “withstand temperatures of up to 95°C without losing their function or forming aggregates.”

These properties indicate that the nanobodies might remain active in the body long enough to be effective. Also, heat-resistant nanobodies are easier to produce, process, and store.

“Our single nanobodies are potentially suitable for inhalation and thus for direct virus neutralization in the respiratory tract,” Dobbelstein noted. “In addition, because they are very small, they could readily penetrate tissues and prevent the virus from spreading further at the site of infection.”

“With the nanobody triad, we literally join forces: In an ideal scenario, each of the three nanobodies attaches to one of the three binding domains,” observed Thomas Güttler, PhD, a scientist in Görlich’s team and the paper’s lead author. “This creates a virtually irreversible bond. The triple will not let release the spike protein and neutralizes the virus even up to 30,000-fold better than the single nanobodies.” Another advantage: The larger size of the nanobody triad expectedly delays renal excretion. This keeps them in the body for longer and promises a longer-lasting therapeutic effect.

The tandem designs, according to Metin Aksu, PhD, a researcher in Görlich’s team and a co-author of the study, combine two nanobodies that target different parts of the receptor-binding domain and together can bind the spike protein. “Such tandems,” he added, “are extremely resistant to virus mutations and the resulting ‘immune escape’ because they bind the viral spike so strongly.”

For all nanobody variants—monomeric, double, and triple—the researchers found that very small amounts are sufficient to stop the pathogen. If used as a drug, this would allow for a low dosage and thus for fewer side effects and lower production costs.

The Göttingen team is currently preparing the nanobodies for therapeutic use. Dobbelstein emphasized: “We want to test the nanobodies as soon as possible for safe use as a drug so that they can be of benefit to those seriously ill with COVID-19 and those who have not been vaccinated or cannot build up an effective immunity.”

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