Figure 4

In vitro potency of 1R-2b. (A) IFN-α mediation of phosphorylation of STAT1, ERK1/2 and AKT in ACHN cells was performed as described in Materials and Methods. Cells were exposed to the indicated amounts of 1R-2b or rhIFN-α2a for 30 min (p-STAT1) or 60 min (p-ERK1/2 and p-AKT). Fold-increase in phosphorylation was calculated relative to total protein loading controls and normalized to untreated cells. (B) NUB1 expression was determined as described in Materials and Methods. ACHN or 786-O cells were exposed to 3000 U/mL of 1R-2b or rhIFN-α2a for 24 h. Up-regulation was determined relative to β-actin loading control and normalized against untreated cells. (C) Growth inhibition was performed as described in Materials and Methods in complete media containing 10% FBS. A dose/response curve was generated with 1R-2b or rhIFN-α2a ranging from 1×10⁵ to 0.26 U/mL. Graphs show growth relative to untreated control and represent the mean ± standard deviation.