Aminoflavone (4H-1-benzopyran-4-one, 5-amino-2-(4-amino-3-fluorophenyl)-6,8-difluoro-7-methyl; AF; NSC 686288) is a new anticancer drug that recently entered Phase I clinical trials. Previous work of our laboratory has demonstrated that the antiproliferative effect of AF in MCF-7 human breast cancer cells is mediated by the Aryl hydrocarbon Receptor (AhR). Treatment of MCF-7 cells with AF resulted in AhR nuclear translocation generating an increase in AhR-target genes expression, such as CYP1A1. It has been demonstrated that CYP1A1 induction is necessary for the conversion of AF pro-drug into several metabolites that cause cell apoptosis.

AF also exhibits noteworthy evidence of antitumor activity in vitro against neoplastic cells of renal origin. In vivo, the drug produced tumor-free, long term survivors in mice bearing subcutaneous renal xenografts. These studies performed on renal cell lines and renal cell carcinoma isolates obtained from patients’ tumors suggest that renal papillary tumors were generally more sensitive to AF than the clear cell tumors. AF treatment of sensitive renal cells, in contrast to resistant cells, promotes CYP1A1 induction, covalent binding of AF-reactive intermediates to several macromolecules and apoptosis.

Based on this evidence, the aim of this study was to evaluate the role of AhR pathway in the antiproliferative effect of AF in human renal cancer cells.

Our results show that AF (0.1-1 μM) inhibited cell growth evaluated by MTS assay in a dose dependent manner in TK10 (47 ± 5% of control, p<0.01) and SN12-c (47 ± 6% of control, p<0.01) human renal cells but not in ACHN cells (96 ± 4% of control). AF treatment (1 μM, 72 h) also induced apoptosis in TK10 and SN12c cells but not in ACHN cells.

In TK10 and SN12-c sensitive cell lines AF (1 μM) induced time dependent AhR nuclear translocation evaluated by Western blot and immunofluorescence. This effect was not seen in resistant ACHN cells. In addition, AF induced AhR transcriptional activity shown by induction of XRE-Luciferase in TK10 (AF 10 μM p<0,01 vs control) and SN12-c (AF 0,1 μM p<0,05 vs control) sensitive cells but not in ACHN cells.

Finally, the antiproliferative effect of AF was abrogated by preincubation of TK10 (80 ± 16%, p<0.01 vs AF) and SN12-c (63 ± 6%, p<0.01 vs AF) cells with the AhR antagonist, α-napthoflavone (αNF, 1 μM). In other experiments, a different AhR antagonist, 4,7-phenanthroline (10 μM), partially reversed growth inhibition by AF in TK10 cells.

These data indicate that AhR-mediated signal transduction is necessary for the cytotoxic effect of AF in sensitive renal tumor cells. Therefore, AhR translocation could be included as a marker of sensitivity to AF in future phase II trials.

AF acting through the AhR pathway is a promising new targeted treatment for renal cancer cases that are refractory to other therapeutic agents currently used in the clinic.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4453.