Abstract 2310: Down-regulating BCL-2 gene expression in a resistant GIST cell line by a RNA G-quadruplex interacting small molecule ligand

Abstract

Abstract We have previously shown that KIT protein expression in the gastrointestinal tumour (GIST) cell line GIST882 can be down-regulated using a small molecule approach that targets expression at the transcriptional level. The small-molecule naphthalene diimide derivative ND1 developed as a G-quadruplex telomere targeting agent can inhibit telomerase activity and can also reduce c-kit mRNA levels, consistent with a dual G-quadruplex mechanism of action in this cell line. An Imatinib (Gleevec) resistant patient-derived cell line GIST48 expresses high levels of the anti-apoptotic protein bcl-2. The compound ND1 down-regulates the expression of bcl-2 in this resistant cell line, while having no discernable effect on c-kit protein expression levels or telomerase activity, both associated with DNA quadruplex stabilization. Instead we observe that translation of the apoptosis-related protein is impeded, while mRNA levels remain unaffected. The identification of a G-rich quadruplex-forming sequence in the 5’-UTR region of the bcl-2 gene, upstream of the translation start site, lead us to the development of a dual luciferase reporter assay. Utilizing a psiCHECK-2 vector we inserted the G-rich 67 base pair bcl-2 5’-UTR gene sequence upstream of the Renilla luciferase start codon. In vitro experiments have shown that the G-rich sequence can form a stable RNA G-quadruplex and interfere with translation. Our in vivo transfection experiments with ND1 in a GIST cell line shows concentration-dependent inhibition of bcl-2 protein expression. RNA G-quadruplex formation and ligand stabilization may thus be a novel anti-cancer route for the down-regulation of the anti-apoptotic protein bcl-2 in chemo-resistant cancers. 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 2310. doi:10.1158/1538-7445.AM2011-2310