Figure 2. WP1130 treatment inhibits norovirus replication.

(A, B) WP1130 treatment inhibits MNV-1 infection in (A) RAW 264.7 (RAW) cells or (B) bone marrow-derived macrophages (BMDMs). Cells were infected with MNV-1 (MOI 5) in the presence of 5 µM WP1130 or DMSO (−). Cells were incubated with WP1130 30 min prior to infection (pre) or at the indicated times post-infection. Virus titers were determined by plague assay 8 hours (RAW) or 12 hours (BMDMs) post-infection. (C) MNV-1 attachment to murine macrophages is not significantly altered by WP1130 treatment. MNV-1 (MOI 5) was incubated for 1 hour on ice with RAW cells treated with 5 µM WP1130 or DMSO. Virus attachment was quantified by qRT-PCR. (D) WP1130 does not inhibit MNV-1 entry. RAW cells were infected with neutral red-containing MNV-1 (MOI 0.001) for 60 min at room temperature and then illuminated with white light. Cells were either treated prior to infection (pre-treatment) or treated for 90 minutes after infection (post-treatment) with 5 µM WP1130 or DMSO. To show the dynamic range of the assay, cells treated with DMSO were also illuminated with white light at the same time as infection was initiated (0 min). (E) WP1130 treatment inhibits MNV-1 replication. MNV-1 genomic RNA was transfected into RAW cells and quantified either 12 hours (Input) or 24 hours later using qRT-PCR. Cell were treated with DMSO or 5 µM WP1130 for the final twelve hours. MNV-1 genome copy number was normalized to DMSO-treated samples. (F) WP1130 treatment inhibits Norwalk virus replication. Norwalk virus replicon-bearing HG23 cells were treated with DMSO, 5 µM WP1130, or 100 µg/ml Ribavirin for 24 hours and Norwalk virus genomes quantitated by qRT-PCR. Norwalk virus genome copy number was normalized to DMSO-treated samples. In all cases, data from at least three independent experiments with two experimental replicates per condition are presented as means +/− S.E.M. *P<0.05, **P<0.01 and *** P<0.001, N.S. non-significant.