Figure 2. Fludarabine inhibits IDO protein independently of STAT1 phosphorylation on Y710 and S727.
A- PBMC were pre-treated with the indicated concentrations of fludarabine or DMSO (vehicle) for 24 h. Cells were washed and activated for 30 min (pSTAT1) or 24 h (total STAT1 and β-actin) with 50 U/ml of IFN-γ. B- MDA-231 were pre-treated with 100 µM fludarabine before activation with 50 U/ml of IFN-γ, anti-CD3 (OKT3) or IgG2a-activated TIL supernatants (Sup.). C- 624.38mel were pre-treated with 50 µM fludarabine, and cultured with anti-CD3 (OKT3) or IgG2a-activated CD4+ T lymphocyte supernatants. B-C Cells were harvested after 30 min (pSTAT1, STAT1 and β-actin) or 24 h (IDO and β-actin). A-C Proteins were extracted for immunoblot analysis. Results are representative of three independent experiments.