2′-Nitroflavone induces cell cycle arrest and apoptosis in HeLa human cervical carcinoma cells

The mechanism of antitumor action of a synthetic nitroflavone derivative, 2′-nitroflavone, was evaluated in vitro in HeLa human cervix adenocarcinoma cells. We showed that the nitroflavone derivative slowed down the cell cycle at the S phase and increase the population of cells at the G2/M phase after 24 h of incubation. The treatment with 2′-nitroflavone also induced an apoptotic response, characterized by an increase of the sub-G1 fraction of cells, by cells with chromatin condensation and membrane blebbing, by a typical ladder of DNA fragmentation and by detection of apoptotic cells stained with Annexin V. The observed apoptosis was regulated by caspase-8 and -9, both contributing to the activation of the effector caspase-3. In addition, inhibitors of caspase-8 or -9 partially protected HeLa cells from 2′-nitroflavone-induced cell death. We also found that 2′-nitroflavone did not affect the total amount of Bax and Bcl-2 proteins, although a translocation of Bax from cytosol to mitochondria was evident after 6 h of exposure. Furthermore, 2′-nitroflavone decreased the expression of the anti-apoptotic Bcl-XL protein, induced the release of cytochrome C to cytosol and increased the levels of Fas and Fas-L. Our results indicated that both death receptor and mitochondria-dependent pathways are involved in the apoptotic cell death triggered by 2′-nitroflavone and suggest that this derivative could be a potentially useful agent for the treatment of certain malignancies.