New synthetic flavone exerts antitumor activity on leukemia cell lines.

MARIANO G. CARDENAS; NORA M. MARDER; ELIDA M. ALVAREZ; LEONOR P. ROGUIN

WASHINGTON, USA

Congreso; 101 Annual Metting of the American Association of Cancer Research; 2010

American Association for Cancer Research

Flavonoids are polyphenolic compounds found in dietary components of vegetable origin. They exhibit multiple biological activities, such as antiproliferative, anxiolytic, antiallergic, antiangiogenic and antioxidant actions. We have previously demonstrated that the synthetic flavone 2´-nitroflavone (2´-NF) exerts a strong antimitogenic activity in vitro in various human and murine adenocarinoma cell lines, without disturbing the proliferation of non-neoplastic cells. We also studied the in vivo efficacy of this compound in a breast cancer murine model. In order to evaluate the effect of 2´-NF in haematological diseases, we first tested its antiproliferative potency in different human and murine leukaemia and lymphoma cell lines. In addition, to elucidate the mechanism of action we explored the cell cycle progression and apoptosis induction either in a human as well as a murine cell line. Methods and results: The antiproliferative activity of 2’-NF in several haematological cell lines was initially evaluated at a 20 µM concentration by the MTS assay. IC50 values were then determined from dose-response curves. Among human cell lines, HL-60 cells (acute promyelocytic leukaemia) were the most sensitive to 2´-NF treatment, showing an IC50 of 1±0.5 µM. Among murine cell lines, similar behaviour was observed for NFS-60 (myelogenous leukaemia) and LB02 (T cell leukaemia) cells (IC50 ≈ 8 µM). When peripheral blood mononuclear cells (PBMC) from healthy donors were used as controls, it was shown that PBMC growth was not significantly affected up to a concentration of 80 µM 2’-NF, suggesting a selective action of the nitroflavone derivative on tumor cells. Different experimental assays were then performed to evaluate the in vitro apoptotic response to 2´-NF. Thus, when HL-60 cells were treated in the presence or absence of 20 µM 2´-NF, cell cycle arrest at G2/M phase was detected after 6 h, followed by an evident increment in the proportion of hypodyploid cells in a time dependent manner (3±1% control vs. 38±1% at 14 h; 3±1% control vs. 54±5% at 24 h). A typical DNA ladder fragmentation pattern was visualized by agarose gel electrophoresis at 24 h and morphological changes of apoptotic cells, such as chromatin condensation, cell shrinkage and membrane blebbing, were observed by fluorescence microscopy. Although the expression of antiapoptotic Bcl-2 and Bcl-XL proteins remained unchanged, a 2 fold increment of the pro-apoptotic protein Bax was obtained after 9 h of exposure to 2´-NF. A significant increase in caspase-3 activity was also detected after 9 h of treatment. By using various experimental approaches, we also demonstrated that 2’-NF induced an apoptotic response in murine LB02 cells. Conclusion: Our results suggest that 2’-NF is a potent and selective antitumor compound, which induces cell cycle arrest and apoptosis in human leukaemia cells. The potential therapeutic use of 2’-NF will be next explored in a leukaemia mouse model.