Recombinant Antigens Based on Non-Glycosylated Regions from RBD SARS-CoV-2 as Potential Vaccine Candidates against COVID-19.

Leandro Alberto Núñez-Muñoz ,Edgar Morales-Ríos,Karla Acosta-Virgen,América Padilla-Viveros,Gabriel Marcelino-Pérez,Berenice Calderón-Pérez,Ana Olivares-Martínez,Hugo González-Conchillos,Miriam Pérez-Saldívar,Daniela Roa-Velázquez,Gustavo Torres-Franco,Jorge Fernández-Hernández,Diana L Peláez-González ,Martha Espinosa‐Cantellano ,Jorge O Ramos-Flores ,José Abrahán Ramírez-Pool ,Diana Angélica Tapia-Sidas ,Roberto Ruiz‐Medrano ,Brenda Yazmín Vargas-Hernández ,Beatriz Xoconostle‐Cázares

doi:10.3390/vaccines9080928

Abstract

The Receptor-Binding Domain (RBD) of the Spike (S) protein from Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has glycosylation sites which can limit the production of reliable antigens expressed in prokaryotic platforms, due to glycan-mediated evasion of the host immune response. However, protein regions without glycosylated residues capable of inducing neutralizing antibodies could be useful for antigen production in systems that do not carry the glycosylation machinery. To test this hypothesis, the potential antigens NG06 and NG19, located within the non-glycosylated S-RBD region, were selected and expressed in Escherichia coli, purified by FPLC and employed to determine their immunogenic potential through detection of antibodies in serum from immunized rabbits, mice, and COVID-19 patients. IgG antibodies from sera of COVID-19-recovered patients detected the recombinant antigens NG06 and NG19 (A450 nm = 0.80 ± 0.33; 1.13 ± 0.33; and 0.11 ± 0.08 for and negatives controls, respectively). Also, the purified antigens were able to raise polyclonal antibodies in animal models evoking a strong immune response with neutralizing activity in mice model. This research highlights the usefulness of antigens based on the non-N-glycosylated region of RBD from SARS-CoV-2 for candidate vaccine development.

Highlights

The Coronaviridae family encompasses several viruses that can cause human diseases ranging from common cold to severe clinical manifestations, such as Middle EastCoronavirus Respiratory Syndrome (MERS-CoV), Severe Acute Respiratory Syndrome (SARS-CoV), and COVID-19 (SARS-CoV-2) [1,2]
Spike protein has been described as a target of the humoral immune response, considering its localization on the viral particle and due to experimental evidence of its high antigenicity
We performed the analysis of the secondary structure (Figure S1) and determined the physicochemical properties of the NG06 and NG19 proteins (Table S2)

Summary

Introduction

The Coronaviridae family encompasses several viruses that can cause human diseases ranging from common cold to severe clinical manifestations, such as Middle EastCoronavirus Respiratory Syndrome (MERS-CoV), Severe Acute Respiratory Syndrome (SARS-CoV), and COVID-19 (SARS-CoV-2) [1,2]. S protein is the largest known type-1 viral fusion protein and contains 22 canonical N-glycosylation sites (N-X-S/T, where X 6= P), as well as 33 O-glycosylation sites [5,6,7,8,9] which could enhance infectivity [10] This affects the host’s ability to develop an effective adaptive immune response [11], or probably causes epitope masking [12] as has been reported for other viral proteins such as the envelope protein (ENV) of HIV [13], glycoprotein 1 (GP1) of Ebola virus [14], hemagglutinin (HA) of the influenza virus [15], the glycoprotein complex of the Lassa Virus

Methods

Results

Conclusion