Diminutive proteins called DARPins have been engineered that are capable of binding to SARS-CoV-2. They neutralized several SARS-CoV-2 variants of concern, and in a mouse model of COVID-19, intranasal administration of one of the DARPins reduced viral burden in respiratory passages. These findings, which were reported in Nature Chemical Biology, suggest that DARPins are promising candidates for the prevention and treatment of infection by SARS-CoV-2.

DARPins, or designed ankyrin repeat proteins, are genetically engineered antibody mimetic proteins derived from naturally occurring ankyrin proteins, which participate in various kinds of high-affinity protein-protein interactions. DARPins are smaller than conventional protein biologics, about one-eighth the size of monoclonal antibodies. Moreover, they are less prone to “go bad” during prolonged storage at moderate-to-high temperatures and can be made in large quantities at low cost. Accordingly, DARPins are, potentially, more affordable than conventional protein biologics.

The SARS-CoV-2-binding DARPins were engineered by scientists in the laboratory of Zhilei Chen, PhD, at Texas A&M University School of Medicine. In an article titled, “A potent and broad neutralization of SARS-CoV-2 variants of concern by DARPins,” the scientists described how they engineered and selected two synthetic proteins—FSR16m and FSR22—for the possible treatment of SARS-CoV-2 infection.

“FSR16m and FSR22 are trimeric proteins composed of DARPin SR16m or SR22 fused with a T4 foldon,” the article’s authors indicated. “Despite selection by a spike protein from a now historical SARS-CoV-2 strain, FSR16m and FSR22 exhibit broad-spectrum neutralization of SARS-CoV-2 strains, inhibiting authentic B.1.351, B.1.617.2, and BA.1.1 viruses.”

When the scientists conducted cryo-electron microscopy experiments, they determined that their DARPins recognized a region of the receptor-binding domain overlapping a critical portion of the angiotensin-converting enzyme 2 (ACE2)-binding surface. When the scientists intranasally administered the DARPins to mice inoculated B.1.617.2, the mice showed less weight loss and 10–100-fold lower viral burden in upper and lower respiratory tracts.

“This study offers the possibility of an on-demand nasal spray able to tackle COVID-19 either before or after virus exposure,” Chen said. Her team’s discovery provides another, potentially lower-cost therapeutic option for those who cannot receive traditional vaccines or are considered high risk.

“The ability of these DARPins to exhibit increased neutralization potency toward SARS-CoV-2 variants contrasts with many human-derived anti-SARS-CoV-2 monoclonal antibodies, which lose neutralization potency,” the article’s authors noted. “The following reasons may account for this phenomenon: (a) [The potency of] in vitro engineered DARPins … appears to be less affected by the virus evolution history. (b) [Due] to its much smaller size, a trimer DARPin can easily engage all three monomers in a spike trimer concurrently. (c) [Both] DARPins engage key residues within the hACE2-binding interface. As emerging natural variants of concern tend to exhibit higher infectivity and ACE2 binding affinity, these variants may become more susceptible to binding and neutralization by our DARPin molecules.”

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