P53 status determines IRF-1 induction of p21 and growth inhibition in HCT116 colon cancer cells

Abstract

IRF-1 is a tumor suppressor which appears to have greater growth inhibition in cancer cells versus non-malignant cells; we hypothesize that cancer cells which have mutant or absent p53 are inhibited more by IRF-1 than non-malignant cells that have wild-type p53. Methods: HCT116 human colon cancer cells with wild-type p53 (p53 wt) and HCT116 cells in which p53 was deleted in both alleles (p53 ko) were used. Cells were treated for 4h with recombinant adenovirus expressing IRF-1 (Ad-IRF-1), empty adenoviral vector (ψ5), or mock, and media changed. Cells were harvested at 24h for western blot for IRF-1, p21, and β-actin, and assessed for growth inhibition at 72h by CellTiter-glo assay. Growth inhibition data is reported as % growth inhibition ± SEM of each treatment compared to mock treatment. Results: Ad-IRF-1 at 100 MOI produced equivalent or more IRF-1 in the p53 wt cell line versus the p53 ko cell line (Fig. B), but demonstrated less growth inhibition (26% versus 62%, Fig. A). This was seen in a dose-related manner with Ad-IRF-1 while ψ5 had no effect on growth inhibition. p21 levels by western were increased by Ad-IRF-1 versus ψ5 control in p53 ko cells, but were slightly decreased in p53 wt cells (Fig. B). p21 levels were higher at baseline in p53 wt cells; p21 is known to be induced by p53. Conclusions: IRF-1 upregulates p21 in p53 ko cells but not in p53 wt cells, and has less growth inhibition in p53 wt cells, consistent with the known function of p21. This may be due to protein-protein interactions or competitive binding in the promoter region of p21.