Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2022 Oct 25:13:1040836.
doi: 10.3389/fimmu.2022.1040836. eCollection 2022.

Cross-reactive neutralizing human monoclonal antibodies mapping to variable antigenic sites on the norovirus major capsid protein

Lauren A Ford-Siltz  1 , Kentaro Tohma  1 , Gabriela S Alvarado  2 , Joseph A Kendra  1 , Kelsey A Pilewski  1 , James E Crowe Jr  2   3 , Gabriel I Parra  1
Affiliations

Cross-reactive neutralizing human monoclonal antibodies mapping to variable antigenic sites on the norovirus major capsid protein

Lauren A Ford-Siltz et al. Front Immunol. .

Abstract

Human noroviruses are the major viral cause of acute gastroenteritis around the world. Although norovirus symptoms are in most cases mild and self-limited, severe and prolonged symptoms can occur in the elderly and in immunocompromised individuals. Thus, there is a great need for the development of specific therapeutics that can help mitigate infection. In this study, we sought to characterize a panel of human monoclonal antibodies (mAbs; NORO-123, -115, -273A, -263, -315B, and -250B) that showed carbohydrate blocking activity against the current pandemic variant, GII.4 Sydney 2012. All antibodies tested showed potent neutralization against GII.4 Sydney virus in human intestinal enteroid culture. While all mAbs recognized only GII.4 viruses, they exhibited differential binding patterns against a panel of virus-like particles (VLPs) representing major and minor GII.4 variants spanning twenty-five years. Using mutant VLPs, we mapped five of the mAbs to variable antigenic sites A (NORO-123, -263, -315B, and -250B) or C (NORO-115) on the major capsid protein. Those mapping to the antigenic site A showed blocking activity against multiple variants dating back to 1987, with one mAb (NORO-123) showing reactivity to all variants tested. NORO-115, which maps to antigenic site C, showed reactivity against multiple variants due to the low susceptibility for mutations presented by naturally-occurring variants at the proposed binding site. Notably, we show that cross-blocking and neutralizing antibodies can be elicited against variable antigenic sites. These data provide new insights into norovirus immunity and suggest potential for the development of cross-protective vaccines and therapeutics.

Keywords: GII.4; antibodies; cross-reactive; gastroenteritis; mapping; neutralization; norovirus.

PubMed Disclaimer

Conflict of interest statement

JC has served as a consultant for Luna Labs USA, Merck Sharp & Dohme Corporation, Emergent Biosolutions, and GlaxoSmithKline, and is a member of the Scientific Advisory Board of Meissa Vaccines, a former member of the Scientific Advisory Board of Gigagen Grifols and is founder of IDBiologics. The laboratory of JC received unrelated sponsored research agreements from AstraZeneca, Takeda, and IDBiologics during the conduct of the study. Vanderbilt University has applied for patents for some of the antibodies in this paper. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

Figure 1
Figure 1
Human mAbs isolated from individuals infected with a GII.4 Sydney 2012 variant exhibit blocking and neutralization against GII.4 Sydney 2012, but not against non-GII.4 genotypes. (A) Carbohydrate blocking of human mAbs against a GII.4 Sydney 2012 variant virus (RockvilleD1/2012). Blocking of serial dilutions of mAbs (starting at 50 μg/mL or 12.5 μg/mL) was determined by comparing the signal to that of the positive control, binding of VLPs in the absence of mAbs. (B) Neutralization of a GII.4 Sydney 2012 virus (strain 011617) with different concentrations of human mAbs. The dashed line represents 5 genome copies/µL, which is the copy number per µL for which the limit of detection is 50% (LOD50) or the “neutralization threshold”, which was defined as the average of genome copies/µL after 1 hour post-inoculation (HPI) such that samples presenting values below the line were unable to replicate and were therefore neutralized. All virus inoculations were performed in the presence of 0.5 mM glycochenodeoxycholic acid (GCDCA), a glycine-conjugated form of the primary bile acid chenodeoxycholic acid. The graph represents the average values of two independent experiments. The error bars represent the geometric standard deviation of the mean. (C) Heat map of the binding of mAbs (at 2 μg/mL) to VLPs of selected genotypes. The value in each cell is the average of the optical density at 405 nm (OD 405 nm) in duplicate wells.
Figure 2
Figure 2
Human mAbs present varying binding and blocking profiles against multiple GII.4 variants spanning over three decades. (A) Heat map of the binding of mAbs to VLPs representing major and minor GII.4 variants. Each cell shows the average of the EC50 values calculated as the half-maximal effective concentration that results in 50% binding to VLPs in duplicate wells, where 100% binding was set as the maximum average OD 405 nm value calculated from each plate at or near antibody saturation. Major pandemic variants are labeled with an asterisk. (B) Heat map of the carbohydrate blocking of human mAbs against VLPs representing major and minor GII.4 variants. Blocking of serial dilutions of mAbs was compared to the signal of the positive control, binding of VLPs in the absence of mAbs. Each cell shows the average of the EC50 values calculated as the half-maximal effective concentration that results in 50% blocking of VLPs in duplicate wells. Major pandemic variants are labeled with an asterisk.
Figure 3
Figure 3
Human mAbs NORO-123, -263, -315B, and -250B bind to antigenic site A and NORO-115 binds to antigenic site C on the major capsid protein. (A) Binding of mAbs NORO-263, -315B, and -250B to wild-type (wt) VLPs from Farmington Hills (FH 2002) and Sydney (SY 2012) variants, or to mutant VLPs with swapped antigenic sites A, C, D, E, and G. (B) Binding of NORO-115 to wt VLPs from Grimsby (GR 1995) and SY 2012 variants, or to mutant VLPs with antigenic sites A, C, D, E, and G derived from the SY 2012 variant. (C) Binding of NORO-123 to SY 2012 VLPs presenting depleted antigenic sites A or G. Values represent the average binding at 2 mg/mL of each mAb. Each bar represents an average of duplicate wells. The dashed line represents the cut-off value (OD 405 nm = 0.2). The error bars represent the standard deviation of the mean.
Figure 4
Figure 4
NORO-263, -315B, and -250B bind to motifs A(I) and/or A(II), while residues 376 and 340 contribute to the binding of NORO-115 to antigenic site C. (A) Structure of the P domain dimer (top view) with motifs A(I), A(II), and A(III) highlighted in red, pink, and maroon, respectively. The HBGA molecule is represented in dark grey. (B) Binding of mAbs NORO-263, -315B, and -250B to wt VLPs from Farmington Hills (FH 2002) and Sydney (SY 2012) variants, or to mutant VLPs with swapped motifs A(I), A(II), or A(III). Values represent the average binding at 2 μg/mL of each mAb. Each bar represents an average of duplicate wells. The dashed line represents the cut-off value (OD 405 nm = 0.2). The error bars represent the standard deviation of the mean. (C) Structure of the P domain dimer (top view) with residues 376 and 340 from antigenic site C (yellow) highlighted in gold. An HBGA molecule is represented in dark grey. (D) Binding of mAb NORO-115 to wt VLPs from Farmington Hills (FH), Camberwell (CA) variants, or to mutant VLPs presenting the backbone of FH variant with residues exchanged with the CA variant (FH 2002: E376Q and FH 2002: G340A/E376Q). The error bars represent the standard deviation of the mean. (E) Sequence alignment of antigenic sites A, C, D, E, and G from selected GII.4 viruses. CA, Camberwell; GR, Grimsby; FH, Farmington Hills; SA, Sakai; HT, Hunter; YE, Yerseke; DH, Den Haag; OS, Osaka; AP, Apeldoorn; NO, New Orleans; SY, Sydney.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

References

    1. Green KY. Norovirus infection in immunocompromised hosts. Clin Microbiol Infect (2014) 20(8):717–23. doi: 10.1111/1469-0691.12761 - DOI - PMC - PubMed
    1. Chhabra P, de Graaf M, Parra GI, Chan MC, Green K, Martella V, et al. . Updated classification of norovirus genogroups and genotypes. J Gen Virol (2019) 100(10):1393–406. doi: 10.1099/jgv.0.001318 - DOI - PMC - PubMed
    1. Parra GI, Squires RB, Karangwa CK, Johnson JA, Lepore CJ, Sosnovtsev SV, et al. . Static and evolving norovirus genotypes: Implications for epidemiology and immunity. PloS Pathog (2017) 13(1):e1006136. doi: 10.1371/journal.ppat.1006136 - DOI - PMC - PubMed
    1. Parra GI. Emergence of norovirus strains: A tale of two genes. Virus Evol (2019) 5(2):vez048. doi: 10.1093/ve/vez048 - DOI - PMC - PubMed
    1. Tohma K, Lepore CJ, Gao Y, Ford-Siltz LA, Parra GI. Population genomics of GII.4 noroviruses reveal complex diversification and new antigenic sites involved in the emergence of pandemic strains. MBio (2019) 10(5):e02202-19. doi: 10.1128/mBio.02202-19 - DOI - PMC - PubMed

Publication types