Replication of human coronaviruses SARS-CoV, HCoV-NL63 and HCoV-229E is inhibited by the drug FK506

Abstract

Recent research has shown that Coronavirus (CoV) replication depends on active immunophilin pathways. Here we demonstrate that the drug FK506 (Tacrolimus) inhibited strongly the growth of human coronaviruses SARS-CoV, HCoV-NL63 and HCoV-229E at low, non-cytotoxic concentrations in cell culture. As shown by plaque titration, qPCR, Luciferase- and green fluorescent protein (GFP) reporter gene expression, replication was diminished by several orders of magnitude. Knockdown of the cellular FK506-binding proteins FKBP1A and FKBP1B in CaCo2 cells prevented replication of HCoV-NL63, suggesting the requirement of these members of the immunophilin family for virus growth.

Fig. 1

Effect of FK506 on SARS-CoV in VeroFM and on HCoV-NL63 replication in CaCo2 cells by qPCR (A–D) and plaque titration (E and F). Data shown are mean values of representative experiments performed in at least triplicates. Left Y-axes represent the percentage of reduction of virus replication in linear scale (A, C and E) and in log scale (B, D and F) at the indicated inhibitor concentrations, respectively. Percentage of cell viability with the mock-treated cells set to 100% are shown on the right Y-axes. FK506 concentrations used for each virus are given on the X-axis.

Fig. 2

Effect of FK506 on HCoV-229E-LUC replication in HuH7 cells by Renilla activity measurement (A and B) and on HCoV-229E-GFP by quantification of GFP fluorescence (C and D). Data shown are mean values (RLU) of a representative experiment performed in triplicate. The left Y-axis represents the percentage of reduction of virus replication in linear scale (A) and in log scale (B), respectively, at the indicated inhibitor concentrations given on the X-axis. Percentage of cell viability with the mock-treated cells set to 100% are shown on the right Y-axis (A). Virus reduction values in log scale are given on the Y-axis of (B). Quantification of GFP fluorescence was used as a semiquantitative measure of virus reduction at the indicated inhibitor concentrations (C and D).

Fig. 3

Growth of HCoV-NL63 on CaCo2-ΔPPIB, -ΔFKBP1A, -ΔFKBP1B knockdown mutant cells. Knockdown of gene expression was analysed by qPCR (A). Virus growth was tested by plaque titration on the knockdown cells (B).