Bispecific antibodies targeting distinct regions of the spike protein potently neutralize SARS-CoV-2 variants of concern

Abstract

The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants of concern threatens the efficacy of existing vaccines and therapeutic antibodies and underscores the need for additional antibody-based tools that potently neutralize variants by targeting multiple sites of the spike protein. We isolated 216 monoclonal antibodies targeting SARS-CoV-2 from plasmablasts and memory B cells collected from patients with coronavirus disease 2019. The three most potent antibodies targeted distinct regions of the receptor-binding domain (RBD), and all three neutralized the SARS-CoV-2 Alpha and Beta variants. The crystal structure of the most potent antibody, CV503, revealed that it binds to the ridge region of SARS-CoV-2 RBD, competes with the angiotensin converting enzyme 2 receptor, and has limited contact with key variant residues K417, E484 and N501. We designed bispecific antibodies by combining non-overlapping specificities and identified five bispecific antibodies that inhibit SARS-CoV-2 infection at concentrations of less than 1 ng/mL. Through a distinct mode of action, three bispecific antibodies crosslinked adjacent spike proteins using dual N-terminal domain-RBD specificities. One bispecific antibody was greater than 100-fold more potent than a cocktail of its parent monoclonals in vitro and prevented clinical disease in a hamster model at a 2.5 mg/kg dose. Notably, two bispecific antibodies in our panel comparably neutralized the Alpha, Beta, Gamma and Delta variants and wild-type virus. Furthermore, a bispecific antibody that neutralized the Beta variant protected hamsters against SARS-CoV-2 expressing the E484K mutation. Thus, bispecific antibodies represent a promising next-generation countermeasure against SARS-CoV-2 variants of concern.

Structural and antigenic variations in the spike protein of emerging SARS-CoV-2 variants

“…Going forward with the identification of more effective therapies against SARS-CoV-2, multiple laboratories have reported the development of bispecific antibodies by combining 2 antibodies that target nonoverlapping epitopes on the spike protein and multivalent nanobodies. These engineered molecules have been described to be highly potent in neutralizing the SARS-CoV-2 variants and suppressing mutational escape [ 119 – 122 ]. Alongside this, antibodies from the NTD and community RBD-1 to RBD-4 are an attractive choice for the therapeutic cocktails being more potent than others, but the data suggest that emerging SARS-CoV-2 variants tend to escape binding by many members of the groups associated with most potent neutralizers [ 80 , 81 ].…”

Mutations of SARS-CoV-2 spike protein: Implications on immune evasion and vaccine-induced immunity

“…Therefore, targeting multiple sites in the spike protein could be a promising strategy to cope with VOC as mutations in the NTD and other regions are suggested to affect the structure and thus the immunogenicity RBD epitopes [ 240 ]. In this regard, Cho et al [ 241 ] designed bispecific antibodies which potently neutralize VOC (Alpha, Beta, Gamma, and Delta variants) and the wild-type virus. In addition, combining RBD and NTD neutralizing antibodies is suggested to be another means of fighting with VOC [ [242] , [243] , [244] ].…”

Impact of the Double Mutants on Spike Protein of SARS-CoV-2 B.1.617 Lineage on the Human ACE2 Receptor Binding: A Structural Insight

“…The nAbs recognize RBD, or other regions on the S glycoprotein, directly or indirectly interfering with the ACE2 interaction [67]. Previous studies have shown that different Abs target different regions on the S protein [68,69]. The epitome mapping studies on the Abs specific to S protein showed that 21% of the 377 epitopes were from RBD [70].…”