Abstract
The concern of COVID-19 persists due to the continuous emergence of variants and the potential spillover of animal coronaviruses. The broad-spectrum neutralizing antibodies play a pivotal role in the prevention and treatment of coronavirus (CoV) infections. Here, we constructed 18 bi-specific antibodies (bsAbs) using 9 antibodies isolated from COVID-19 convalescents and vaccinated individuals, designed as dual variable domain immunoglobulin (DVD-Ig). A bsAb 5-HI showed a high binding capability to the S1 subunit of spike and exhibited breadth and potency against pseudotyped SARS-CoV-2 variants of concerns (VOCs) and SARS-related-CoVs (SARSr-CoVs), with half maximal effective concentration (EC
50
) of 0.028–3.444 nM and 50% inhibitory concentration (IC
50
) of 0.008-0.800 nM. In addition, it retained neutralization potency against the peudotyped virus of recently prevalent JN.1 strain (IC
50
, 12.74 nM). We found that the parental antibodies showed weak or no binding to the receptor binding domain (RBD) of the SARS-CoV, EG.5.1, and JN.1. However, the 5-HI maintained the binding with RBD and prevented the binding between hACE2 and RBD (IC
50
for the RBD of SARS-CoV, 1.067 nM; EG.5.1, 0.423 nM; JN.1, 0.223 nM). In neutralization assays with the authentic virus, we found that the 5-HI effectively neutralized Omicron variants XBB.1.5 (IC
50
, 0.308 nM), EG.5.1 (IC
50
, 0.129 nM), and JN.1 (IC
50
, 13.692 nM), while its parental antibodies showed weakened or no neutralization. Therefore, the 5-HI represents a promising candidate for further development in the treatment and prevention of ongoing evolved SARS-CoV-2 VOCs and other SARSr-CoVs that potentially emerge in the future.