Aryl Hydrocarbon Receptor Mediates Sensitivity of MCF-7 Breast Cancer Cells to Antitumor Agent 2-(4-Amino-3-methylphenyl) Benzothiazole

Abstract

2-(4-Amino-3-methylphenyl) benzothiazole (NSC 674495; DF 203) demonstrates drug uptake and metabolism by tumor cells sensitive to the antiproliferative activity of the drug [J Med Chem 1999;42:4172-4184]. In insensitive cells, little metabolism occurs. Because CYP1A1 can metabolize DF 203, the aryl hydrocarbon receptor (AhR) may mediate drug action. We demonstrate here that DF 203 increases CYP1A1 and CYP1B1 transcription in sensitive MCF-7 cells, accompanied by AhR translocation to the nucleus, increase in xenobiotic-responsive element (XRE)-driven luciferase activity, and induction of protein/DNA complexes on the XRE sequence of the CYP1A1 promoter. MDA-MB-435 and PC3 cells, resistant to DF 203, did not show drug-induced CYP1A1 and CYP1B1 gene expression. AhR was observed to be constitutively localized in the nucleus, with no induction of XRE-driven luciferase activity in transiently transfected cells and weak or no induction of protein/DNA complexes on the XRE sequence of CYP1A1. Taken together, these data elucidate a novel basis for antitumor drug action: induction in sensitive cells of a metabolizing system for the drug itself. These results suggest that clarification of the basis for differential engagement of AhR-related signaling in different tumor cell types may aid in further preclinical development and perhaps early clinical studies.

DNA damage and cell cycle arrest induced by 2-(4-amino-3-methylphenyl)-5-fluorobenzothiazole (5F 203, NSC 703786) is attenuated in aryl hydrocarbon receptor deficient MCF-7 cells

“…Treated cells were subsequently maintained for an additional 48 h, and cellular protein was measured by the sulphforhodamine-B (SRB) assay as described previously (Monks et al, 1991). Briefly, cells were fixed with trichloroacetic acid and stained with SRB.…”

“…Experiments were repeated at least three times. Stained cells were then visualised on a Zeiss Axiovert microscope using  63 objective and images were captured with an Optronics CCD camera (Loaiza-Pérez et al, 2002).…”

“…Evaluation of CYP1A1 and CYP1B1 gene expression in MCF-7 and AH R100 cells was performed by real-time RT -PCR, using 15 cycles of PCR, primers and probes described in a previous report (Loaiza-Pérez et al, 2002). RNA was isolated using Qiagen kits (Qiagen, Valencia, CA, USA).…”

“…In contrast, nonresponsive cells, for example, breast carcinoma MDA-MB-435 or prostate carcinoma PC-3 cells, showed no activation of the AhR and no induction of CYP1A1 after drug treatment (Loaiza-Pérez et al, 2002). We also have reported a suitable model to investigate the role of the AhR in mediating drug cytotoxicity (Loaiza-Pérez et al, 2001), consisting of AhR-deficient AH R100 cells, derived from MCF-7 human breast epithelial cancer cells by continuous exposure to escalating concentrations of benzo[a]pyrene .…”

“…In contrast, nonresponsive cells, for example, breast carcinoma MDA-MB-435 or prostate carcinoma PC-3 cells, showed no activation of the AhR and no induction of CYP1A1 after drug treatment (Loaiza-Pérez et al, 2002). We also have reported a suitable model to investigate the role of the AhR in mediating drug cytotoxicity (Loaiza-Pérez et al, 2001), consisting of AhR-deficient AH R100 cells, derived from MCF-7 human breast epithelial cancer cells by continuous exposure to escalating concentrations of benzo[a]pyrene . AH R100 cells display relative resistance to the cytotoxic effects of several polycyclic aromatic hydrocarbons (PAHs), such as benzo [a]pyrene and dimethylbenz [a]anthracene (DMBA), due to impairment of the AhR signal transduction pathway Ciolino et al, 2002).…”