Abstract 2817: Dietary chemopreventive agent (curcumin), hypomethylates 5′ promoter of DCLK1 gene in human colon cancer cells and restores expression of long isoform of DCLK1

Abstract

Abstract Curcumin, a dietary chemopreventive agent, inhibits proliferation of colon cancer cells by inhibiting multiple oncogenic pathways. Curcumin has also emerged as a non-toxic epigenetic modulator in recent years. Direct interaction of curcumin with DNA methyl transferases (DMTs), inhibits their enzymatic activity, resulting in hypomethylation of specific genes. DCLK1 is a cancer stem cell (CSC) marker, critically required for colon tumorgenesis and for maintaining tumoregenic/metastatic potential of colon cancers. The 5′ promoter of DCLK1 gene in human colorectal cancers (hCRCs) is epigenetically silenced due to DNA methylation, resulting in loss of expression of long isoform of DCLK1 (DCLK1-L). However, a short isoform (DCLK1-S) is expressed from an alternate promoter within IntronV, in response to several oncogenic pathways, including activation of NF-κB pathway, and is required for maintaining tumorigenic/metastatic potential of hCRC cells. Since a subset of DCLK1+CSCs survive inhibitory effects of curcumin, resulting in relapse of the disease (Can Res, 2014), we hypothesized that curcumin will likely inhibit DCLK1-S expression, by inhibiting NF-κB pathway, but cause re-expression of DCLK1-L by hypomethylating the 5′promoter. A representative hCRC cell line, devoid of DCLK1-L expression due to hypermethylation of 5′promoter (HCT116), was used. Dose and time dependent effects of curcumin were examined on 2D/3D growths of HCT116 cells. As expected, proliferation of curcumin treated cells was significantly decreased in a time and dose dependent manner. DCLK1-L was re-expressed, corresponding to de-methylation of 5′ promoter; curcumin (5-10μM) was optimally effective as an epigenetic modifier at 72h. Higher doses were required for inhibiting DCLK1-S expression and corresponded with loss of binding of NF-κBp65 to the NF-κB cis element in IntronV promoter of DCLK1 gene. The loss in DCLK1-S expression, likely explains significant inhibitory effects of curcumin on the growth of colon cancers and CSCs, as reported by many investigators. Re-expression of DCLK1-L, marker of quiescent stem cells in normal colons, may allow curcumin-treated cancer cells to survive and become dormant/quiescent. It is speculated that ‘CSCs’, re-expressing DCLK1-L, are initially induced into dormancy, but revert back to expressing DCLK1-S after a period of dormancy, resulting in relapse of the disease, as previously reported; this intriguing possibility will be examined in future studies. Citation Format: Shubhashish Sarkar, Malaney O'Connell, Pomila singh. Dietary chemopreventive agent (curcumin), hypomethylates 5′ promoter of DCLK1 gene in human colon cancer cells and restores expression of long isoform of DCLK1. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2817. doi:10.1158/1538-7445.AM2015-2817