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The curcuminoid CLEFMA selectively induces cell death in H441 lung adenocarcinoma cells via oxidative stress

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CLEFMA or 4-[3,5-bis(2-chlorobenzylidene-4-oxo-piperidine-1-yl)-4-oxo-2-butenoic acid] is a curcuminoid being developed as an anticancer drug. We recently reported that it potently inhibits proliferation of various cancer cells. In this project, we investigated the effect of CLEFMA on gene expression profile in H441 lung adenocarcinoma cells, and studied its mechanism of action. In microarray data, we observed a deregulation of genes involved in redox and glutamate metabolism. Based on the affected ontologies, we hypothesized that antiproliferative activity of CLEFMA could be a result of the induction of reactive oxygen species (ROS). We tested this hypothesis by determining the levels of glutathione (GSH) and ROS in H441 cells treated with CLEFMA. We observed a rapid depletion of intracellular GSH/GSSG ratio. Using a cell-permeable fluorogenic substrate, we found that CLEFMA significantly induced ROS in a time- and dose-dependent manner (p < 0.05). Flow-cytometry with a mitochondria-selective fluorescent reporter of ROS indicated that the CLEFMA-induced ROS was of mitochondrial origin. In contrast to the cancer cells, the normal lung fibroblasts (CCL-151) did not show any increase in ROS and were resistant to CLEFMA-induced cell death. Furthermore, the addition of antioxidants, such as catalase, superoxide dismutase and N-acetylcysteine, rescued cancer cells from CLEFMA-induced cell death. Gene expression pathway analysis suggested that a transcription factor regulator Nrf2 is a pivotal molecule in the CLEFMA-induced deregulation of redox pathways. The immunoblotting of Nrf2 showed that CLEFMA treatment resulted in phosphorylation and nuclear translocation of Nrf2 in a time-dependent fashion. Based on these results, we conclude that induction of ROS is critical for the antiproliferative activity of CLEFMA and the Nrf2-mediated oxidative stress response fails to salvage H441 cells.

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Acknowledgements

The work was partially funded by a grant from the National Cancer Institute (1R03 CA143614-01).

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Authors and Affiliations

  1. Department of Pharmaceutical Sciences, University of Oklahoma Health Science Center, 1110 North Stonewall Avenue, Oklahoma, OK, 73117, USA

    Kaustuv Sahoo & Vibhudutta Awasthi

  2. Arthritis & Immunology Department, Oklahoma Medical Research Foundation, 825 N.E. 13th Street, Mailstop #24, Oklahoma, OK, 73104, USA

    Mikhail G. Dozmorov

  3. Department of Molecular and Integrative Physiology, University of Kansas Medical Center, Kansas, KS, 66160, USA

    Shrikant Anant

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  1. Kaustuv Sahoo
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Correspondence to Vibhudutta Awasthi.

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Sahoo, K., Dozmorov, M.G., Anant, S. et al. The curcuminoid CLEFMA selectively induces cell death in H441 lung adenocarcinoma cells via oxidative stress. Invest New Drugs 30, 558–567 (2012). https://doi.org/10.1007/s10637-010-9610-4

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