The vaccine was formulated in 100-L single-dose vials of WI-SARS containing the equivalent of either 0.5, 1.0, or 1.5?g of S protein. without adjuvant was poorly immunogenic in mice; a second dose resulted in a significant boost in antibody levels, even in the absence of adjuvant. The use of adjuvants resulted in higher antibody titers, with the AS01B-adjuvanted vaccine being slightly more immunogenic than the AS03A-adjuvanted vaccine. Two doses of WI-SARS with and without Adjuvant Systems were highly efficacious in mice. In hamsters, two doses of WI-SARS with and without AS01B were immunogenic, and two doses of 2?g of WI-SARS with and without the adjuvant provided complete protection from early challenge. Although antibody titers had declined in all groups of vaccinated hamsters 18 wk after the second dose, the vaccinated hamsters were still partially protected from wild-type virus challenge. Vaccine with adjuvant provided better protection than non-adjuvanted WI-SARS vaccine at this later time point. Enhanced disease was not observed in the lungs or liver of hamsters following SARS-CoV challenge, regardless of the level of serum neutralizing antibodies. Introduction Severe acute respiratory syndrome (SARS) was first recognized in Asia in early 2003 and caused 8000 cases and 774 deaths before the outbreak ended in July 2003 (1). The causative agent was a newly-identified coronavirus (CoV) that spread to humans from a yet unidentified animal reservoir through civet cats and possibly other infected animals (2C4). The spike (S) glycoprotein of SARS-CoV is the attachment protein and the target of the protective neutralizing antibody response (5). Several SARS-CoV vaccines have been developed using different vaccine platforms, including whole inactivated, subunit, DNA, vectored, and live virus vaccines (6C8). The immunogenicity and efficacy of some of these experimental vaccines have been evaluated in animal models such as mice, hamsters, ferrets, and non-human primates, and a few of them have been evaluated in phase I clinical trials (6,9,10). In general, vaccines that elicit a robust serum neutralizing antibody response in animal models provide protection from challenge with virus (11). The inclusion of aluminum salts and saponin as adjuvants has been reported to enhance the immunogenicity of subunit and inactivated virus vaccines (12,13). The mouse is a reasonable model for screening candidate SARS vaccines because SARS-CoV replicates to high titers in the respiratory tract of mice following intranasal inoculation (14). Furthermore, mice develop a neutralizing antibody response that confers protection Clarithromycin against subsequent challenge (14). However, young mice do not develop clinical illness, pneumonitis is transient, and the virus is cleared by about day 5 post-challenge (14). Therefore, the efficacy of a SARS vaccine can only be judged by quantitative virology in this model. Hamsters are an excellent model for SARS-CoV infection and vaccine efficacy since they support high levels of viral replication and histopathological changes in respiratory tissues, and demonstrate clinical signs (reduced activity) following intranasal inoculation (6,11,15). Efficient replication of SARS-CoV in the respiratory tract following intranasal inoculation of golden Syrian hamsters leads to high virus titers in the lungs from days 1 through 7 post-infection Clarithromycin (p.i.), with peak titers occurring at day 3 p.i. Pronounced pneumonitis accompanies SARS-CoV infection on days Igf2 3C5 p.i., and consolidation of up to one-third of the lungs occurs at days 5C7 p.i. Viral replication and pulmonary pathology coincide with greatly reduced physical activity of the hamsters (16). Thus the efficacy of a SARS vaccine can be judged by quantitative virology and histopathological changes in the lungs of hamsters. In this study, we evaluated the immunogenicity and efficacy of a -propiolactone (BPL)-inactivated whole virion SARS-CoV vaccine in BALB/c mice and golden Syrian hamsters. We specifically examined three variables: (1) the response to different doses of antigen, (2) the effect Clarithromycin of different adjuvants, and (3) the duration of protection, which was evaluated by administering challenge virus to vaccinated hamsters at 4 or 18 wk post-vaccination. Materials and Methods Viruses and cells SARS-CoV (Urbani strain 200300592) was obtained from the Centers for Disease Control and Prevention, Atlanta, GA. Virus stock was produced in Vero cells (CCL81, obtained from ATCC) in serum-free conditions using OptiPRO Clarithromycin medium (Invitrogen, Paisley, Scotland). For vaccine preparation, Vero cells were grown in serum-free Clarithromycin conditions and infected with a dilution of 1 1:100 (v:v) of the SARS-CoV virus stock in 120?mL of OptiPRO medium per tray of cells (Nunc, Roskilde, Denmark). The cells were incubated at 37C for 72?h. At 3 days p.i., when the cytopathic effect (CPE) was visible, the virus was harvested by collecting cell supernatants. The infectious titer of the virus was determined using a 50% cell culture infectious dose (CCID50) method. Purification and inactivation of SARS-CoV Vaccine purification was then performed. After low-speed.

The vaccine was formulated in 100-L single-dose vials of WI-SARS containing the equivalent of either 0