Inhibition of severe acute respiratory syndrome coronavirus replication by niclosamide

Abstract

Antiviral agents are urgently needed to fight severe acute respiratory syndrome (SARS). We showed that niclosamide, an existing antihelminthic drug, was able to inhibit replication of a newly discovered coronavirus, SARS-CoV; viral antigen synthesis was totally abolished at a niclosamide concentration of 1.56 microM, as revealed by immunoblot analysis. Thus, niclosamide represents a promising drug candidate for the effective treatment of SARS-CoV infection.

Figures

FIG. 1.

Inhibition of SARS-CoV replication by niclosamide in Vero E6 cells. (A) Chemical structure of niclosamide (2′, 5-dichloro-4′-nitrosalicylanilide); (B) dose-dependent inhibition of SARS-CoV antigen synthesis in Vero E6 cells by niclosamide. This immunoblot analysis shows that niclosamide, at a concentration of 1.56 μM or higher, completely inhibited the synthesis of viral antigens of SARS-CoV in Vero E6 cells. The bottom panel is an immunoblot stained with antiactin antibody as an internal control. The viral antigens spike protein (S) and nucleocapsid protein (NP) are indicated.

FIG. 2.

Inhibition of viral antigen synthesis in SARS-CoV-infected Vero E6 cells as revealed by an IFA. Vero E6 cells were treated with the various concentrations of niclosamide indicated, and an IFA was performed at 48 h postinfection. Vero E6 cells were fixed and stained with the same primary antiserum directed against SARS-CoV as that used in the immunoblot analysis. Fluorescein isothiocyanate-conjugated goat anti-human immunoglobulin (Jackson ImmunoResearch Laboratoeies, Inc.) was used as the secondary antibody. The positive panel shows cells infected by SARS-CoV but without drug treatment, while the negative panel shows cells without virus infection.

FIG. 3.

Effect of niclosamide on reduction of the viral yields in the culture supernatants from SARS-CoV-infected Vero E6 cells. Viral RNA was isolated from culture supernatants from SARS-CoV-infected Vero E6 cells and analyzed by RT-PCR. PCR products of 368 bp were observed when Vero E6 cells were treated with niclosamide at concentrations of 1.56 μM and lower, whereas no PCR products could be detected when cells were treated with niclosamide at concentrations of 3.12 μM and higher. Representative results from five separate experiments are shown. Positive, cells infected by SARS-CoV without drug treatment; Negative, cells without virus infection. The RT-PCR products were quantitated by image analysis, and results are shown in Table 1.