Abstract 2110: Modulating NRAS mRNA translation by nucleic acid clamp-mediated stabilization of the 5′- UTR G-quadruplex

Abstract

Abstract NRAS, a proto-oncogene belonging to the RAS oncogene superfamily, is a intracellular signal cascade mediator, with intrinsic GTPase activity, functioning to initiate the MAPK cell signaling pathway. Among the RAS oncogene superfamily that is mutated in approximately 30% of all cancers, the NRAS mutation accounts for about 15% of RAS related human malignancies, particularly myeloid leukemias and cutaneous melanomas. Recently, a stable G-quadruplex (G4) was previously identified in the 5′-untranslated region (UTR) of NRAS mRNA. This G4 functioned as a translational silencer, downregulating NRAS protein and leading to the hypothesis that a small nucleic acid clamp could be optimized to facilitate the formation of specifically this G4, decrease NRAS expression, and facilitate cell death in melanoma cells. In the present study, a set of clamps were designed and examined for their ability to facilitate the formation of a G4 in the 5′-UTR sequence of NRAS. These clamps are comprised of ribonucleotides complementary to the RNA 5′- and 3′-regions flanking the G4-forming sequence held together by a small linker element. Based on binding studies consisting of electromobility shift assays and circular dichroism, an optimal flank length and linker length was chosen for cellular studies. Effects on cellular viability and NRAS expression are being examined in leukemia and melanoma cells, and RNA ChIP-Seq is being used to localize the intracellular activity of the clamps to the 5′-UTR. These nucleic acid clamps offer a high degree of selectivity for G4s over linear RNA facilitated by the small linker region; moreover, the approach of complementary nucleotides, versus small molecule-mediated G4 stabilization, allows for specificity for just one G4 out of all of the intracellular structures. This approach not only provides a solid base for developing the clamp into a non-small molecule cancer therapy, but also opens a new window for modulating oncogenic gene expression at the transcriptional or translational levels. The current study, in particular, offers a novel approach to anti-NRAS-mediated therapies for the treatment of myeloid leukemias and cutaneous melanomas. Citation Format: Taisen Hao, Tracy A. Brooks. Modulating NRAS mRNA translation by nucleic acid clamp-mediated stabilization of the 5′- UTR G-quadruplex. [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 2110. doi:10.1158/1538-7445.AM2015-2110