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. 2021 Jan-Dec;13(1):1893426.
doi: 10.1080/19420862.2021.1893426.

Bispecific VH/Fab antibodies targeting neutralizing and non-neutralizing Spike epitopes demonstrate enhanced potency against SARS-CoV-2

Shion A Lim  1 , Josef A Gramespacher  1 , Katarina Pance  1 , Nicholas J Rettko  1 , Paige Solomon  1 , Jing Jin  2 , Irene Lui  1 , Susanna K Elledge  1 , Jia Liu  1 , Colton J Bracken  1 , Graham Simmons  2 , Xin X Zhou  1 , Kevin K Leung  1 , James A Wells  1   3   4
Affiliations

Bispecific VH/Fab antibodies targeting neutralizing and non-neutralizing Spike epitopes demonstrate enhanced potency against SARS-CoV-2

Shion A Lim et al. MAbs. 2021 Jan-Dec.

Abstract

Numerous neutralizing antibodies that target SARS-CoV-2 have been reported, and most directly block binding of the viral Spike receptor-binding domain (RBD) to angiotensin-converting enzyme II (ACE2). Here, we deliberately exploit non-neutralizing RBD antibodies, showing they can dramatically assist in neutralization when linked to neutralizing binders. We identified antigen-binding fragments (Fabs) by phage display that bind RBD, but do not block ACE2 or neutralize virus as IgGs. When these non-neutralizing Fabs were assembled into bispecific VH/Fab IgGs with a neutralizing VH domain, we observed a ~ 25-fold potency improvement in neutralizing SARS-CoV-2 compared to the mono-specific bi-valent VH-Fc alone or the cocktail of the VH-Fc and IgG. This effect was epitope-dependent, reflecting the unique geometry of the bispecific antibody toward Spike. Our results show that a bispecific antibody that combines both neutralizing and non-neutralizing epitopes on Spike-RBD is a promising and rapid engineering strategy to improve the potency of SARS-CoV-2 antibodies.

Keywords: Bispecific; COVID-19; SARS-CoV-2; knob-in-hole; neutralizing antibody.

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Figures

Figure 1.
Figure 1.
Fabs identified by phage display bind Spike RBD and Secto with high affinity outside of the ACE2 binding site
Figure 2.
Figure 2.
Bispecific VH/Fab IgGs bind with high affinity to trimeric Secto. (a) Cartoon schematic of the bispecific VH/Fab IgG antibody scaffold utilized in this study. (b-e) Multipoint BLI measurements (10 nM, 5 nM, and 2.5 nM) of the indicated bispecific antibody on Secto. (b) Bis1 (VH A01/FabC01) (c) Bis2 (VH B01/FabC01) (d) Bis3 (VH A01/FabD01) (e) Bis4 (VH B01/FabD01)
Figure 3.
Figure 3.
Bispecific VH/Fab IgGs are more potent in neutralizing SARS-CoV-2 pseudovirus than the mono-specific counterparts
Figure 4.
Figure 4.
Bispecific VH/Fab IgGs neutralize authentic SARS-CoV-2 virus more potently than the mono-specific VH-Fcs
See this image and copyright information in PMC

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