Abstract 198: Helicobacter pylori infection via miR-328 suppression and CD44 expression in gastric mucosa causes gastric cancer initiation and progression

Abstract

Abstract Backgrounds: We previously revealed that CD44, cancer stem-like marker, interacts with xCT, a glutamate-cystine transporter, and controls the intracellular level of reduced glutathione. Helicobacter pylori (HP) infection is known to be a major cause of gastric cancer and be associated with increased expression of CD44 on gastric epithelial cells, but precise mechanism of CD44 up-regulation still has uncertain. On the other hand, microRNAs (miRNAs) are noncoding RNAs that regulate molecular pathways in cancer by targeting various genes. The aim of the current study was to identify the miRNA regulating CD44 expression in H. pylori-infected gastric mucosa during the development of preneoplastic lesions. Methods and Results: To investigate the regulation of CD44 through miRNA, we performed 384 miRNA quantitative RT-PCR array analyses in six human gastrointestinal cancer cell lines that differ in CD44 expression status. We identified miR-328 which is predicted as CD44-targeted and established the functional relationship between miR-328 and CD44. MiR-328 overexpression decreased CD44 expression and led to inhibition of cancer cell growth and impairing chemo- and ROS (reactive oxygen species) - resistance. We performed luciferase assay using vectors containing CD44 3′- UTR, which might be directly bound by miR-328, using the miRanda algorithm, and the results indicated that miR-328 regulated CD44 expression by directly targeting its 3′-UTR. We polarized THP-1 that human monocytic leukemia cell line to M1- and M2 macrophages, and detected ROS production of its by using 2′,7′-dichlorofluorescein diacetate, and ROS induced miR-328 down-regulation and CD44 up-regulation by co-culture gastric cancer cell lines with M1- and M2-polarized THP-1 macrophages. We further analyzed the association between miR-328-CD44 pathway and HP infection in human gastric mucosa. We identified that HP infection correlated with miR-328 down-regulation and CD44 up-regulation. Lastly, we examined the DNA methylation of the miR-328 5′-flanking region by bisulfite pyrosequencing in human gastric mucosa, but HP infection were not related with the DNA methylation level in this region. Conclusions: We show that inflammatory response with HP infection causes CD44 expression via miR-328 suppression, resulting in cancer initiation and progression, but HP infection was unrelated to the DNA methylation in the miR-328 5′-flanking region. Citation Format: Keisuke Miyake, Takatsugu Ishimoto, Hidetaka Sugihara, Kojiro Eto, Daisuke Izumi, Junji Kurashige, Yukiharu Hiyoshi, Shiro Iwagami, Yoshifumi Baba, Yuji Miyamoto, Naoya Yoshida, Hideo Baba. Helicobacter pylori infection via miR-328 suppression and CD44 expression in gastric mucosa causes gastric cancer initiation and progression. [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 198. doi:10.1158/1538-7445.AM2015-198