Abstract 4022: Novel evidence for m6A methylation regulators as prognostic biomarkers and potential therapeutic targets in gastric cancer
Authors: Tadanobu Shimura, Raju Kandimalla, Yuji Toiyama, Yoshinaga Okugawa, Masato Kusunoki, Ajay Goel
Journal: Cancer Research (2019)

Abstract Purpose: N6-methyladenosine (m6A) post-transcriptional RNA modification represents the most abundant epigenetic alteration in eukaryotic cells. These m6A modifications are regulated by a cascade of enzymes and cofactors, categorized as ‘writers’ [methyltransferase-like 3 (METTL3), METTL14, and Wilms’ tumor 1-associating protein (WTAP1)], ‘erasers’ [fat mass and obesity-associated protein (FTO) and alkylated DNA repair protein AlkB homolog 5 (ALKBH5)] and ‘readers’ [YTHDF1, and YTHDF2 (YTH N6-Methyladenosine RNA Binding Protein)]. While emerging evidence indicates that these enzymes play a central role in RNA metabolism (e.g. RNA stability, translation, splicing, transport and localization), their clinical significance, if any, remains unclear. Herein, we for the first time, systematically unraveled the functional role, as well as the clinical significance of m6a regulators in gastric cancer (GC). Experimental design: First, we investigated the expression of the seven m6A regulator genes (FTO, METTL3, METTL14, WTAP, ALKBH5, YTHDF1, and YTHDF2) in a large cohort of 171 GC patients by qRT-PCR assays. Subsequently, a multivariate Cox-regression model was developed using the 7-gene panel to evaluate its predictive potential, followed by Kaplan Meier analyses for survival outcomes. In addition, we undertook a series of functional studies to investigate the oncogenic role of FTO in GC cells, and subsequent validation of our findings in an animal model. Results: Gastric cancer patients with low-expression of METTL3, METTL14, ALKBH5, WTAP and YTHDF1 demonstrated significantly poor OS (p=0.02, 0.001, 0.01, 0.02 and 0.02, respectively), while patients with high FTO expression exhibited markedly worse OS (p&amp;lt;0.0001). Furthermore, the cumulative risk-score derived from these gene panel also significantly associated with poor OS, with a corresponding hazard ratio of 5.47 (95% CI: 3.18-9.41, p&amp;lt;0.0001). We observed that FTO expression was frequently up-regulated in GC cell lines, particularly in those with epithelial-mesenchymal-transition (EMT) features. Functional studies with FTO knockdown in HGC27 and AGS cells inhibited cell proliferation (p=0.005, and &amp;lt;0.0001, respectively) and migratory potential (p&amp;lt;0.0001, respectively); while its overexpression in MKN28 cells resulted in enhanced proliferation and migration (p&amp;lt;0.0001, and 0.007, respectively). Finally, confirming our in-vitro findings, FTO suppression led to significant tumor growth inhibition in a xenograft animal model (p&amp;lt;0.0001). Conclusions: We provide first evidence that m6A regulators may serve as important prognostic biomarkers in GC patients. Our functional studies reveal that FTO is an important oncogene, and may be a novel therapeutic target in patients with gastric cancer. Citation Format: Tadanobu Shimura, Raju Kandimalla, Yuji Toiyama, Yoshinaga Okugawa, Masato Kusunoki, Ajay Goel. Novel evidence for m6A methylation regulators as prognostic biomarkers and potential therapeutic targets in gastric cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4022.