Inactivated SARS-CoV vaccine elicits high titers of spike protein-specific antibodies that block receptor binding and virus entry

Biochemical and Biophysical Research Communications
Volume 325, Issue 2, 10 December 2004, Pages 445-452

Yuxian He, Yusen Zhou, Pamela Siddiqui, Shibo Jiang

Abstract

The only severe acute respiratory syndrome (SARS) vaccine currently being tested in clinical trial consists of inactivated severe acute respiratory syndrome-associate coronavirus (SARS-CoV). However, limited information is available about host immune responses induced by the inactivated SARS vaccine. In this study, we demonstrated that SARS-CoV inactivated by β-propiolactone elicited high titers of antibodies in the immunized mice and rabbits that recognize the spike (S) protein, especially the receptor-binding domain (RBD) in the S1 region. The antisera from the immunized animals efficiently bound to the RBD and blocked binding of RBD to angiotensin-converting enzyme 2, the functional receptor on the susceptible cells for SARS-CoV. With a sensitive and quantitative single-cycle infection assay using pseudovirus bearing the SARS-CoV S protein, we demonstrated that mouse and rabbit antisera significantly inhibited S protein-mediated virus entry with mean 50% inhibitory titers of 1:7393 and 1:2060, respectively. These data suggest that the RBD of S protein is a major neutralization determinant in the inactivated SARS vaccine which can induce potent neutralizing antibodies to block SARS-CoV entry. However, caution should be taken in using the inactivated SARS-CoV as a vaccine since it may also cause harmful immune and/or inflammatory responses.

Keywords

SARS-CoV, Vaccine, Spike protein, Receptor-binding domain, Antibodies

Interferon-beta and interferon-gamma synergistically inhibit the replication of severe acute respiratory syndrome-associated coronavirus (SARS-CoV)

Virology
Volume 329, Issue 1, 10 November 2004, Pages 11-17

Bruno Sainz Jr. , Eric C. Mossel, C. J. Peters, Robert F. Garry

Abstract

Recent studies have shown that interferon-gamma (IFN-γ) synergizes with IFN-α/β to inhibit the replication of both RNA and DNA viruses. We investigated the effects of IFNs on the replication of two strains of severe acute respiratory syndrome-associated coronavirus (SARS-CoV). While treatment of Vero E6 cells with 100 U/ml of either IFN-β or IFN-γ marginally reduced viral replication, treatment with both IFN-β and IFN-γ inhibited SARS-CoV plaque formation by 30-fold and replication by 3000-fold at 24 h and by > 1 × 105-fold at 48 and 72 h post-infection. These studies suggest that combination IFN treatment warrants further investigation as a treatment for SARS.

Keywords

Coronavirus, Interferon-gamma, Interferon-beta

Inactivation of the coronavirus that induces severe acute respiratory syndrome, SARS-CoV

Journal of Virological Methods
Volume 121, Issue 1, October 2004, Pages 85-91

Miriam E.R. Darnella, Kanta Subbaraob, Stephen M. Feinstonea, Deborah R. Taylora

Abstract

Severe acute respiratory syndrome (SARS) is a life-threatening disease caused by a novel coronavirus termed SARS-CoV. Due to the severity of this disease, the World Health Organization (WHO) recommends that manipulation of active viral cultures of SARS-CoV be performed in containment laboratories at biosafety level 3 (BSL3). The virus was inactivated by ultraviolet light (UV) at 254 nm, heat treatment of 65 °C or greater, alkaline (pH > 12) or acidic (pH < 3) conditions, formalin and glutaraldehyde treatments. We describe the kinetics of these efficient viral inactivation methods, which will allow research with SARS-CoV containing materials, that are rendered non-infectious, to be conducted at reduced safety levels.

Keywords

SARS, Coronavirus, Virus inactivation, Tissue culture

Development and evaluation of an efficient 3′-noncoding region based SARS coronavirus (SARS-CoV) RT-PCR assay for detection of SARS-CoV infections

Journal of Virological Methods
Volume 120, Issue 1, 1 September 2004, Pages 33-40

Huo-Shu H Houng, David Norwood, George V Ludwig, Wellington Sun, Minta Lin, David W Vaughn

Abstract

The severe acute respiratory syndrome (SARS) epidemic originating from China in 2002 was caused by a previously uncharacterized coronavirus that could be identified by specific RT-PCR amplification. Efforts to control future SARS outbreaks depend on the accurate and early identification of SARS-CoV infected patients. A real-time fluorogenic RT-PCR assay based on the 3′-noncoding region (3′-NCR) of SARS-CoV genome was developed as a quantitative SARS diagnostic tool. The ideal amplification efficiency of a sensitive SARS-CoV RT-PCR assay should yield an E value (PCR product concentration increase per amplification cycle) equal to 2.0. It was demonstrated that the 3′-NCR SARS-CoV based RT-PCR reactions could be formulated to reach excellent E values of 1.81, or 91% amplification efficacy. The SARS-CoV cDNA preparations derived from viral RNA extract and the cloned recombinant plasmid both exhibit the identical amplification characteristics, i.e. amplification efficacy using the same PCR formulation developed in this study. The viral genomic copy (or genomic equivalences, GE) per infectious unit (GE/pfu) of SARS-CoV used in this study was also established to be approximate 1200–1600:1. The assay’s detection sensitivity could reach 0.005 pfu or 6–8 GE per assay. It was preliminarily demonstrated that the assay could efficiently detect SARS-CoV from clinical specimens of SARS probable and suspected patients identified in Taiwan. The 3′-NCR based SARS-CoV assay demonstrated 100% diagnostic specificity testing samples of patients with acute respiratory disease from a non-SARS epidemic region.

Keywords

Severe acute respiratory syndrome, SARS coronavirus, Real-time RT-PCR, Quantitative RT-PCR, 3′-Noncoding region.

The 3D structure analysis of SARS-CoV S1 protein reveals a link to influenza virus neuraminidase and implications for drug and antibody discovery

Journal of Molecular Structure: THEOCHEM
Volume 681, Issues 1–3, 26 July 2004, Pages 137-141

Xue Wu Zhang, Yee Leng Yap

Abstract

The spike protein of SARS-associated coronavirus (SARS-CoV) is an important target for anti-SARS drug discovery. Its S1 domain is responsible for receptor binding and SARS-CoV entry into cells. In this study, we constructed a rational 3D model for S1 domain of SARS-CoV spike protein by fold recognition and molecular modeling techniques. We found that there is a structure similarity between S1 protein and influenza virus neuraminidase. Our analyses suggest that the existing anti-influenza virus inhibitors and anti-neuraminidase antibody could be used as a starting point for designing anti-SARS drugs, vaccines and antibodies. Interestingly, our prediction for antibody is consistent with a recently experimental discovery of anti-SARS antibody.

Keywords

SARS-CoV, S1 protein, Structure, Influenza virus, Inhibitor, Antibody