FIG 2.

KPT-335 inhibits RSV replication. (A) A549 cells were treated with DMSO or increasing concentrations of KPT-335 for 72 h. At 70 h posttreatment, 20 μl of CellTiter Blue reagent was added to each well and incubated for 2 h at 37°C. The plate was shaken for 10 s, and fluorescence emission was measured at 560/590 nm. The average fluorescence value of the culture medium background (wells containing no cells) was subtracted from each experimental well. The mean fluorescence emission values of the wells containing DMSO only were considered 100% viability and were used to calculate percent relative cell viability of the wells containing KPT-335. The percent viability versus the log₁₀ concentration of KPT-335 was plotted using GraphPad Prism, and the values were fitted to a nonlinear regression curve to determine the CC₅₀. The data shown are from triplicate samples from one experiment and are representative of three independent experiments. (B) A549 cells were infected with RSV A2 (MOI of 0.5) and treated with increasing amounts of KPT-335 from 2 h p.i. Samples were collected at 72 h p.i., and titers of infectious RSV were determined by immuno-plaque assay as described in Materials and Methods. Average RSV titer in infected samples treated with DMSO was taken as 100% and served as the control; this value was used to calculate percent infection of the samples treated with KPT-335. The percent infection versus the log₁₀ concentration of KPT-335 was plotted using GraphPad Prism, and the values were fitted to a nonlinear regression curve to determine the IC₅₀. The data shown are from triplicate samples from one experiment and are representative of three independent experiments.