Genistein mediates the selective radiosensitizing effect in NSCLC A549 cells via inhibiting methylation of the keap1 gene promoter region

Oncotarget. 2016 May 10;7(19):27267-79. doi: 10.18632/oncotarget.8403.

Abstract

Non-small cell lung cancer (NSCLC) cells often possess a hypermethylated Keap1 promoter, which decreases Keap1 mRNA and protein expression levels, thus impairing the Nrf2-Keap1 pathway and thereby leading to chemo- or radio-resistance. In this study, we showed that genistein selectively exhibited a radiosensitizing effect on NSCLC A549 cells but not on normal lung fibroblast MRC-5 cells. Genistein caused oxidative stress in A549 cells rather than MRC-5 cells, as determined by the oxidation of the ROS-sensitive probe DCFH-DA and oxidative damage marked by MDA, PCO or 8-OHdG content. In A549 instead of MRC-5 cells, genistein reduced the level of methylation in the Keap1 promoter region, leading to an increased mRNA expression, thus effectively inhibited the transcription of Nrf2 to the nucleus, which suppressed the Nrf2-dependent antioxidant and resulted in the upregulation of ROS. Importantly, when combined with radiation, genistein further increased the ROS levels in A549 cells whereas decreasing the radiation-induced oxidative stress in MRC-5 cells, possibly via increasing the expression levels of Nrf2, GSH and HO-1. Moreover, radiation combined with genistein significantly increased cell apoptosis in A549 but not MRC-5 cells. Together, the results herein show that the intrinsic difference in the redox status of A549 and MRC-5 cells could be the target for genistein to selectively sensitize A549 cells to radiation, thereby leading to an increase in radiosensitivity for A549 cells.

Keywords: Keap1/Nrf2; genistein; methylation; oxidative stress; selective radiosensitivity.

MeSH terms

  • A549 Cells
  • Apoptosis / drug effects
  • Apoptosis / genetics
  • Apoptosis / radiation effects
  • Carcinoma, Non-Small-Cell Lung / genetics
  • Carcinoma, Non-Small-Cell Lung / metabolism
  • Carcinoma, Non-Small-Cell Lung / pathology
  • Cell Line
  • Cell Survival / drug effects
  • Cell Survival / genetics
  • Cell Survival / radiation effects
  • DNA Methylation / drug effects*
  • DNA Methylation / radiation effects
  • Fibroblasts / cytology
  • Fibroblasts / metabolism
  • Gene Expression Regulation, Neoplastic / drug effects
  • Gene Expression Regulation, Neoplastic / radiation effects
  • Genistein / pharmacology*
  • Humans
  • Kelch-Like ECH-Associated Protein 1 / genetics*
  • Lung Neoplasms / genetics
  • Lung Neoplasms / metabolism
  • Lung Neoplasms / pathology
  • Oxidation-Reduction / drug effects
  • Oxidation-Reduction / radiation effects
  • Oxidative Stress / drug effects*
  • Oxidative Stress / radiation effects
  • Promoter Regions, Genetic / genetics*
  • Radiation Tolerance / drug effects
  • X-Rays

Substances

  • KEAP1 protein, human
  • Kelch-Like ECH-Associated Protein 1
  • Genistein