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. 2021 Dec;101(Pt B):108187.
doi: 10.1016/j.intimp.2021.108187. Epub 2021 Sep 28.

The SARS CoV-2 spike directed non-neutralizing polyclonal antibodies cross-react with Human immunodeficiency virus (HIV-1) gp41

Reshma Perween  1 , Murugavelu PraveenKumar  1 , Tripti Shrivastava  1 , Hilal Ahmed Parray  1 , Vanshika Singh  1 , Swarandeep Singh  2 , Adarsh Chiranjivi  1 , Kamini Jakhar  1 , Sudipta Sonar  1 , Mahima Tiwari  1 , Reema  1 , Anil Kumar Panchal  1 , Chandresh Sharma  1 , Deepak Kumar Rathore  1 , Shubbir Ahamed  1 , Sweety Samal  1 , Shailendra Mani  1 , Sankar Bhattacharyya  1 , Supratik Das  1 , Kalpana Luthra  2 , Rajesh Kumar  3
Affiliations

The SARS CoV-2 spike directed non-neutralizing polyclonal antibodies cross-react with Human immunodeficiency virus (HIV-1) gp41

Reshma Perween et al. Int Immunopharmacol. 2021 Dec.

Abstract

Cross-reactivity among the two diverse viruses is believed to originate from the concept of antibodies recognizing similar epitopes on the two viral surfaces. Cross-reactive antibody responses have been seen in previous variants of SARS and SARS-CoV-2, but little is known about the cross reactivity with other similar RNA viruses like HIV-1. In the present study, we examined the reactivity the SARS-CoV-2 directed antibodies, via spike, immunized mice sera and demonstrated whether they conferred any cross-reactive neutralization against HIV-1. Our findings show that SARS-CoV-2 spike immunized mice antibodies cross-react with the HIV-1 Env protein. Cross-neutralization among the two viruses is uncommon, suggesting the presence of a non-neutralizing antibody response to conserved epitopes amongst the two viruses. Our results indicate, that SARS-CoV-2 spike antibody cross reactivity is targeted towards the gp41 region of the HIV-1 Env (gp160) protein. Overall, our investigation not only answers a crucial question about the understanding of cross-reactive epitopes of antibodies generated in different viral infections, but also provides critical evidence for developing vaccine immunogens and novel treatment strategies with enhanced efficacy capable of recognising diverse pathogens with similar antigenic features.

Keywords: Class I viruses; Cross-reactivity; Gp41; HIV-1; Non-neutralizing antibodies; SARS-CoV2.

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Conflict of interest statement

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Figures

Fig. 1
Fig. 1
Cross-reactive binding of SARS-CoV-2 antibodies to HIV-1 glycoproteins. A (I and II) & B. In ELISA and Western blot research, polyclonal antibodies from spike immunised mouse sera were examined for cross-reactive binding to gp140 protein (from clade C J41 foldon & clade B JRFL foldon). The sera collected from N protein (SARS-CoV-2) immunized mice and pre-immune sera was included here as assay negative control. The immunoblot and ELISA experiments were repeated a minimum of 3 times. Data is presented as the mean ± SD, and differences between groups were determined by two-way analysis of variance (ANOVA) followed by Tukey’s post hoc tests using GraphPad Prism 7. Statistical significance between the control and different groups is shown as *P < 0.05, **P < 0.01, ***P < 0.0001, ****P < 0.0001 and ns (non-significant).
Fig. 2
Fig. 2
FACS analysis for cross reactive binding antibodies between SARS-CoV-2 and HIV-1: A). The FACS-based cell surface antibody staining assay with HIV-1 efficiently cleaved, functional Envs 4-2. J41 (clade C), JRFL and JRCSF (clades B) with mice spike hyperimmune sera. A representative histogram showing binding of spike immunized mice polyclonal sera to cell surface expressed HIV-1 protein. (B) GMFI data for the binding of HIV cell surface expressed Env to mice SARS-CoV-2 serum samples. All the experiments were done in duplicate and repeated at least two times. Statistical significance between Pre-immune and different treated groups is shown as ****P < 0.0001.
Fig. 3
Fig. 3
A). Cross neutralization potential of SARS-CoV-2, HIV-1 polyclonal antibodies and effect of depletion on cross reactive binding antibodies on cross neutralization: Cross-neutralization potential of sera was assessed against pseudotyped viruses expressing HIV-1 clones representing different subtypes. The value represents the serum neutralization titers. In the case of SARS-CoV-2, a CPE based assay was performed and for HIV-1, a TZM-bl cell neutralization assay was carried out. Neutralization assays were done in duplicates and repeated at least two independent times B). The extent of binding of the depleted and undepleted spike serum with magnetic beads coated with HIV-1J41 gp140 protein was accessed by ELISA. C). Degree of shift in sensitivity of Wuhan SARS-CoV2 viruses to plasma depleted with HIV-1 Env (J41 gp140). The value in the graph represents the CPE value. No change in neutralization of plasma samples was observed in depleted and undepleted samples. Statistical significance was determined using student t-test and p < 0.05 was considered significant and ns (non-significant).
Fig. 4
Fig. 4
A & B). HIV-1 gp140 from 4-2J41 (clade C), JRFL (clade B) proteins were treated with PNGase and deglycosylated. The spike immunized mice sera bound with both glycosylated and deglycosylated forms of gp140 protein. In a parallel blot, an equal amount of proteins was loaded and Anti-His tag antibody was used for probing. C). Cross-reactivity of binding was assessed in presence of methyl α-d-mannopyranoside. Plates were coated with SARS-CoV-2 Spike protein and incubated with dilutions of methyl α-d-mannopyranoside along with a constant amount of the indicated antibodies. Antibody binding was quantified via ELISA. Statistical significance between the control and other experimental groups was estimated by two-way analysis of variance (ANOVA) followed by Tukey’s post hoc ergo propter hoc tests using GraphPad Prism 7. Statistical significance between control and different groups is shown as, ****P < 0.0001 and ns (non-significant).
Fig. 5
Fig. 5
A & B). Accessing potential role of glycosylation in cross-reactivity of Anti-SARS-C0V2 polyclonal antibodies: Cross-reactivity of spike immunized mice sera was tested for gp120 and gp140 protein ELISA and in Western blot analysis. The spike immunized mice sera specifically bound to the gp140 protein and did not cross-react with the gp120 protein. The same blot was restriped and blotted with PGT121 mAb to insure equal loading of experimental proteins. C). The extent of binding of spike and RBD immunized mice sera was tested for HIV-1 MPER peptide.
Fig. 6
Fig. 6
A). Bidirectional immunoreactivity of HIV-1 gp140 immunized sera to the SARS-CoV-2 Spike. Serial dilutions of HIV-1 gp140 immunized rabbit sera were assessed for SARS-CoV-2 S protein binding. Milk coated wells were used as negative control in ELISA binding assay. B). Purified spike proteins were detected on Western blot by using gp140 immunized rabbit sera as primary antibody followed by using HRP conjugated anti Rabbit-Fc antibody. C). RBD-ACE2 competition assay suggests that HIV-1 polyclonal antibodies bind to RBD epitopes that are non-overlapping with ACE2 binding site. D). Spike protein was treated with PNGase and deglycosylated. The HIV-1 gp140 foldon immunized rabbit sera bound with both glycosylated and deglycosylated forms of spike protein. Statistical significance was determined using student t-test and p < 0.05 was considered significant and ns (non-significant).
Fig. 7
Fig. 7
Isotyping of cross-reactive binding antibodies: An ELISA binding assay was used to assess the isotyping of cross-reactive antibodies in spike mice immune serum.
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