Abstract 2771: Epigenetic regulation of the cocaine and amphetamine regulated transcript (CART) in cancer

Abstract

Abstract The cocaine and amphetamine regulated transcript (CART) is well documented in the central nervous system for its role in appetite regulation and reward pathways. In recent studies CART has been proven to be a poor prognostic marker in lymph node-negative, estrogen receptor (ER)-positive breast cancer patients, and has also been shown to be associated with worse survival in small bowel carcinoid tumours. The regulation of CART in these different cancer types however remains largely unknown. The presence of a large CpG island in the putative promoter region and the lack of expression of CART in many cancer cell lines suggest a possible epigenetic mechanism of suppression. A panel of various cancer cell lines (breast, colorectal, endometrial and ovarian) were assessed for endogenous CART expression via western blot and RT-PCR. Cells were then treated with forskolin and leptin; agents which have been previously shown to induce CART expression in the CNS. Cells were also treated with CART conditioned media, which has been previously shown to be the only factor capable of inducing CART expression. Treatment was carried out with varying doses of two DNA methyltransferase inhibitors, 5-Aza-2′-deoxycytidine and RG108, followed by examination of CART expression post treatment. Bisulfite sequencing of the promoter region of CART was performed in all cell lines and a small selection of formalin fixed paraffin embedded (FFPE) tissue samples. Whole methylome sequencing was carried out to confirm this promoter region of CART and study the global effects of treatment with epigenetic modifiers and CART peptide. Assessment of a collection of cell lines via western blot has shown CART expression to be negative across the panel. Moreover, treatment with forskolin and leptin did not result in re-expression of CART. Treatment with 5-Aza-2′-deoxycytidine has however resulted in increased CART mRNA levels, and subsequent bisulfite sequencing of the promoter region of CART has revealed dense methylation at the CpG island. Treatment with CART conditioned media may also induce promoter demethylation and subsequent re-expression of CART. Collectively, this data suggests the potential involvement of DNA methylation in the regulation of CART. Citation Format: Kate Connor, Sudipto Das, Bruce Moran, Laoighse Mulrane, William M. Gallagher, Darran O’Connor. Epigenetic regulation of the cocaine and amphetamine regulated transcript (CART) in cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2771.