Abstract 1147: RNA-targeting approaches to modulate alternative RNA splicing in cancer

Abstract

Abstract Alternative RNA splicing (AS) is a critical step in gene expression regulation. This differential inclusion of sequences in the mRNA is a source of functional transcriptomic diversity that allows multiple RNA isoforms to be expressed from the same gene. Dysregulation of AS is hallmark of cancer and leads to the expression of RNA isoforms that promote tumor initiation and maintenance. Understanding how tumor-associated RNA isoforms impact cancer biology is needed for the development of cancer therapies targeting AS. Here, we describe two approaches to modulate AS in cancer cells. First, we utilize CRISPR-Artificial Splicing Factors (CASFx), splicing factor domains fused to catalytically inactive Cas13 proteins, to direct splicing activity to a specific site using a guide RNA (gRNA). We demonstrate that this approach can be used to promote either exon inclusion or skipping in mRNA from several cancer-associated genes (e.g., HRAS, EP400, TRA2β, SRSF3) depending on the gRNA binding location and the splicing-factor domain fusion. Second, we develop antisense oligonucleotides (ASOs) that target a non-coding poison exon in the oncogenic splicing factor TRA2β. This exon functions as a regulatory “poison exon”, which induces RNA degradation when included, and is differentially spliced in multiple tumor types compared to their corresponding adjacent normal tissue. TRA2β-targeting ASOs promoting poison exon inclusion cause decreased TRA2β protein expression, increased cell death, and decreased proliferation across multiple cancer cell lines. Additionally, compared to cancer cells, normal cell lines were less affected by TRA2β-targeting ASO toxicity, demonstrating a therapeutic window for using this technology to specifically target cancer cells. Citation Format: Nathan K. Leclair, Laura Urbanski, Mattia Brugiolo, Marina Yurieva, Brenton R. Graveley, Albert Cheng, Olga Anczukow. RNA-targeting approaches to modulate alternative RNA splicing in cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1147.