A potent neutralizing nanobody against SARS-CoV-2 with inhaled delivery potential

Summary The outbreak of COVID-19 has emerged as a global pandemic. The unprecedented scale and severity call for rapid development of effective prophylactics or therapeutics. We here reported Nanobody (Nb) phage display libraries derived from four camels immunized with the SARS-CoV-2 spike receptor-binding domain (RBD), from which 381 Nbs were identified to recognize SARS-CoV-2-RBD. Furthermore, seven Nbs were shown to block interaction of human angiotensin converting enzyme 2 (ACE2) with SARS-CoV-2-RBD-variants, bat-SL-CoV-WIV1-RBD and SARS-CoV-1-RBD. Among the seven candidates, Nb11-59 exhibited the highest activity against authentic SARS-CoV-2 with ND50 of 0.55 μg/mL. Nb11-59 can be produced on a large-scale in Pichia pastoris, with 20 g/L titer and 99.36% purity. It also showed good stability profile, and nebulization did not impact its stability. Overall, Nb11-59 might be a promising prophylactic and therapeutic molecule against COVID-19, especially through inhalation delivery. Graphical Abstract Highlights 381 Nanobodies were identified to recognize SARS-CoV-2-RBD including several mutants Nb11-59 exhibited potent antiviral activity against authentic SARS-CoV-2 with ND50 of 0.55 μg/mL Nb11-59 can be large-scale produced in Pichia pastoris with titers reached 20 g/L Nb11-59 showed a good stability and could be developed as an inhaled drug to treat COVID-19.