Current SARS-CoV-2 Omicron subvariants evade immunity from most neutralizing antibodies and vaccines. This study identified a nanobody (aS3A) cross-reactive with SARS-CoV-1 and SARS-CoV-2 RBD. A bispecific nanobody dimer (2-3-Fc), engineered by fusing aS3A-Fc to aRBD-2, exhibited potent neutralization against all major variants of concern, including BA.5. In hamsters, a single systemic dose (10 mg/kg) or a lower intranasal dose (5 mg/kg) of 2-3-Fc provided complete or drastic protection against Omicron infection, respectively. A variant, Y296-3-Fc, showed enhanced neutralization of BA.2.75. This research provides potential therapeutic and prophylactic candidates against SARS-CoV-2 variants.
