Abstract 1408: CSTF2-induced alternative polyadenylation of GPC3 RNA contributes to hepatocellular carcinoma

Abstract Alternative polyadenylation (APA) involves the addition of a polyadenylate (poly-A) tail to the 3' end of an mRNA molecule, which is essential for mRNA stability and translation into proteins. This mechanism is critical for gene expression and function, particularly in cancer. Glypican-3 (GPC3) is significantly overexpressed in hepatocellular carcinoma (HCC) and is associated with poor prognosis. APA profiling analysis showed that the 3’ UTR of GPC3 is shortened, which strongly correlates with increased GPC3 expression in HCC. Disruption of GPC3 led to inhibited cellular growth and increased apoptotic cell death in HCC cells, while ectopic overexpression of a GPC3 plasmid stimulated growth and proliferation in these cells. Furthermore, Cleavage Stimulation Factor Subunit 2 (CSTF2), a component of the cleavage stimulation factor complex, was found to promote the shortening of GPC3's 3' UTR through APA regulation in HCC cells. CSTF2 expression also correlated with GPC3 levels and the dPDUI (Percentage of Distal PolyA Site Usage Index) of GPC3 in HCC. Knockdown of CSTF2 resulted in decreased GPC3 expression, which inhibited cell growth and proliferation while increasing apoptotic cell death in HCC cells. Additionally, CSTF2 knockdown led to lengthening of the GPC3 3’ UTR, whereas overexpression of CSTF2 shows the opposite effect. Using predictive analysis with online tools TargetScan, miRDB, and miRabel, 40 miRNAs were identified as potential binders to the GPC3 3’ UTR. In vitro functional analysis demonstrated that miR-96-5p and miR-140-5p sustain the normal expression of GPC3 in hepatocytes, and that the shortening of the 3’ UTR of GPC3 RNA contributes to hepatocarcinogenesis. Overall, these findings suggest that the aberrant overexpression of GPC3, driven by selective APA regulation mediated by CSTF2, contributes to the malignant transformation and growth of hepatocytes, providing a novel therapeutic target for liver malignancies. Citation Format: Suk Woo Nam, Soyoung Jeon, Sang Yean Kim, Min Jeong Na, Jin Woong Ha. CSTF2-induced alternative polyadenylation of GPC3 RNA contributes to hepatocellular carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 1408.