Figure 5. DiFiD induces G₂/M arrest and apoptosis.

(A) Cell cycle analysis was performed in HN5 and FaDu cell lines after 24 or 48 h treatment with IC₅₀ concentrations of DiFiD, (HN5=750 nM; FaDu=1.5 μM). Figure is representative of three independent studies. (B) Representative immunoblot of HN5 cells treated with 750 nM DiFiD for 24 hours demonstrates expression of cyclin B1, CDC2, Pro- and cleaved-PARP protein with GAPDH used as a loading control. (C) Representative immunoblot of FaDu cells treated with 1.50 μM DiFiD for 24 hours demonstrates expression of cyclin B1, CDC2, Pro- and cleaved-PARP protein with GAPDH used as a loading control. (D) Annexin V assay of HN5 cells following treatment with DiFiD at the IC₅₀ dose. HN5 cells were treated with DiFiD at the IC₅₀ dose for 48 h, stained with Annexin V (FITC) and PI, and analyzed by flow cytometry. (E) Caspase 3/7 assay demonstrates caspase cleavage in HN5 cells 48 h post treatment with DiFiD. Data represents cumulative results from three independent experiments with ± SEM ***p<0.001.