Abstract 1528: Activation of NRF2/ABCC1 axis confers resistance to topoisomerase II poisons in human oral malignancies

Abstract

Abstract Etoposide (VP-16), a DNA topoisomerase II poison, is an important clinical used chemotherapeutic agent for human oral malignancies. An etoposide-resistant cell line, KB-7D, has been generated from human oral epidermoid carcinoma KB cells to investigate the mechanism of action of drug resistance in oral malignancies. Previous studies revealed that KB-7D cells were approximately 50-fold more resistant to etoposide as compared to parental KB cells. It also exhibited cross-resistant to chemotherapeutic agents such as doxorubicin. This multi-drug resistance may be caused by the over-expression of ABCC1, leading to the decrease in drug accumulation in KB-7D cells. Our current work continues the effort to investigate the mechanism of regulation of ABCC1 expression in the etoposide-derived drug resistant cells and subsequently find the molecular target that can be used to restore therapeutic efficacy in chemo-refratory cancers. Down-regulation of ABCC1 by RNA interference and a selective inhibitor, MK-571, significantly enhanced the chemosensitivity to etoposide and doxorubicin in KB and KB-7D cells. To further determine the possible transcriptional factors that regulate the ABCC1 transactivation, the promoter region of ABCC1 was examined. A NRF2 binding sequence, antioxidant responsive element (ARE), has been found locate in the ABCC1 promoter at -470 to -458 upstream from the transcription start site. Real-time RT-PCR and Western blot analyses showed that expression of NRF2 mRNA and protein in KB-7D cells was 1.4 to 1.8 folds higher than those expressed in the parental cells. In addition, Chromatin Immunoprecipitation (ChIP) analysis revealed that NRF2 directly targeted to the ARE sequence in the ABCC1 promoter region. Interestingly, down-regulation of NRF2 decreased the expression of ABCC1 and also increased the chemosensitivity of KB-7D cells against selected anti-cancer drugs. In addition, cells incubated with an NRF2 activator, tBHQ, induced nucleus accumulation of NRF2 and over-expression of ABCC1, resulting in the enhancement of drug-resistance against etoposide or doxorubcin in KB and KB-7D cells. In summary, constitutive activation of NRF2-dependent ABCC1 contributed to the causation of chemoresistance in etoposide-derived drug resistant cells. Blockage of ABCC1 expression by manipulation of the NRF2 signaling pathway enhanced the chemotherapeutic efficacy in cells. Therefore, targeting NRF2 may be able to reverse chemoresistance in chemo-refractory oral malignancies. In addition, development of NRF2 inhibitors may be a new strategy to overcome chemoresistance in human cancers. (The study was supported by grants of Department of Health DOH99-TD-C-111-004, Taiwan.) Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1528. doi:10.1158/1538-7445.AM2011-1528