Someday soon, we may have llamas to thank for stopping the COVID-19 pandemic, specifically, we may express our gratitude to a pet llama named Wally. From Wally’s blood, scientists at the University of Pittsburgh School of Medicine have isolated “nanobodies” that bind to the spike proteins on the SARS-CoV-2 virus, disabling its entry into cells. And if the viruses cannot enter cells, they cannot reproduce and spread. Wally’s nanobodies, after some manipulation by the researchers, stop the virus in its tracks.
There’s more good news. The research team believes the nanobodies can be manufactured in large quantities, rapidly distributed globally, and administered by inhalers, getting them right where they are needed most: in the respiratory tract and the lungs. The nanobodies appear to have a long shelf life, are inexpensive and safe. What’s not to like about Wally’s gift to the world?
These special llama antibodies, called “nanobodies,” are much smaller than human antibodies and many times more effective at neutralizing the SARS-CoV-2 virus. They’re also much more stable.
“Nature is our best inventor,” said senior author Yi Shi, assistant professor of cell biology at Pitt. “The technology we developed surveys SARS-CoV-2 neutralizing nanobodies at an unprecedented scale, which allowed us to quickly discover thousands of nanobodies with unrivaled affinity and specificity.” [Emphasis added.]
The story first hit the media August 8 in a preprint on bioRxiv, accompanied August 11 by news from the University of California San Francisco (UCSF) with a good short video explaining how the nanobodies bind to the spike proteins of the coronavirus. UCSF’s research, very similar to Pitt’s, indicates international interest in this approach. Indeed, researchers at Hebrew University in Jerusalem are participating with Shi’s team. This is not a meteoric story making a quick flash and then disappearing into the void like so many cancer cures and memory boosters that never make it to market. Although clinical trials were not announced in either press release or the paper in Science, demand for this kind of rapidly deployable and inexpensive treatment could drive government support. The UCSF team is seeking a patent for their engineered nanobodies, which they named AeroNabs.
Antibodies vs. Nanobodies
Many have heard about antibody treatments for COVID-19, such as Regeneron, but nanobodies (sometimes abbreviated Nb or Nabs) have several advantages. Yi Shi, one of the authors of the Science paper, explains.
Shi’s nanobodies can sit at room temperature for six weeks and tolerate being fashioned into an inhalable mist to deliver antiviral therapy directly into the lungs where they’re most needed. Since SARS-CoV-2 is a respiratory virus, the nanobodies could find and latch onto it in the respiratory system, before it even has a chance to do damage.
In contrast, traditional SARS-CoV-2 antibodies require an IV, which dilutes the product throughout the body, necessitating a much larger dose and costing patients and insurers around $100,000 per treatment course.
“Nanobodies could potentially cost much less,” said Shi. “They’re ideal for addressing the urgency and magnitude of the current crisis.”
Wally the Llama provided the raw material which was potent already. Then researchers applied some genetic engineering to make the natural nanobody binding even stronger.
In collaboration with Cheng Zhang at Pitt and Dina Schneidman-Duhovny at the Hebrew University of Jerusalem, the team found that their nanobodies use a variety of mechanisms to block SARS-CoV-2 infection. This makes nanobodies ripe for bioengineering. For instance, nanobodies that bind to different regions on the SARS-CoV-2 virus can be linked together, like a Swiss army knife, in case one part of the virus mutates and becomes drug-resistant.
Medical researchers commonly look for therapeutic agents in animals. Monoclonal antibodies, for instance, can be “fished” in animals using phage display methods. Among mammals, camelids (including camels, llamas, and alpacas) have an unusual way of producing small single-domain antibodies with higher specificity to individual proteins. Being a tenth the size of human antibodies, it makes them extremely potent and stable.
“As a virologist, it’s incredible to see how harnessing the quirkiness of llama antibody generation can be translated into the creation of a potent nanoweapon against clinical isolates of SARS-CoV-2,” said study coauthor and CVR Director Paul Duprex.
The research team injected some of the virus spike protein into Wally. About two months later, they collected hundreds of nanobodies his body had produced to fight the invader. Then they found the ones that bound to the coronavirus most strongly.
Then, with the help of Pitt’s Center for Vaccine Research (CVR), the scientists exposed their nanobodies to live SARS-CoV-2 virus and found that just a fraction of a nanogram could neutralize enough virus to spare a million human cells from being infected.
Nanobody search efforts are not new, but the particles are difficult to find. Shi’s team at Pitt developed new technology within the past three years for identifying, characterizing, and sorting through thousands of Nbs rapidly. That’s how they were able to find ones with the highest affinity and, within 3 months, produce Nbs that bind more strongly by orders of magnitude than any others that have been produced. The paper in Science ends on a very optimistic note.
Here, in vivo antibody affinity maturation [in Wally] followed by advanced proteomics enabled the rapid discovery of a diverse repertoire of high-affinity RBD [receptor binding domain] Nbs, including an ultrapotent neutralizer with sub-picomolar affinity, which is unprecedented for natural, single-domain antibodies. We demonstrated the simplicity and versatility of Nb bioengineering and the outstanding physicochemical properties of the monomeric Nbs and their multivalent forms. To our knowledge, the multivalent constructs represent the most potent SARS-CoV-2 neutralizers to date. Flexible and efficient administration, such as inhalation may further improve their antiviral efficacy while minimizing the dose, cost, and potential toxicityfor clinical applications. The high sequence similarity between Nbs and human IgGs [immunoglobulins] may restrain the immunogenicity. It is possible to fuse the antiviral Nbs with highly stable, albumin-Nb constructs to improve pharmacokinetics. These high-quality Nbs can also be applied as rapid and economic point-of-care diagnostics. We envision that the Nb technology described here will contribute to curbing the current pandemic and possibly a future event.
The most challenging part of the work was done by Wally the Llama. His immune system had to recognize the invader and then manufacture tiny particles that could fit onto its unique shape, so that it could be neutralized. The rest of the work by humans was just refining what the intelligent design in Wally’s immune system had accomplished.
For Operation Warp Speed, the world’s leading pharmaceutical companies have revved up their search for a vaccine from four years to under one year. Wally found the key within two months. Put this potentially lifesaving discovery in the ID column.