Abstract Protein synthesis has a key role of maintaining cellular homeostasis such as growth and metabolism. However, dysregulation of translational process is a frequent feature of cancer and contributes to cancer progression. Since oncogenes and tumor suppressor genes are regulated by the translation machinery, therapeutic drugs which target the translation machinery have the possibility for overcoming cancer. We then focused on the hypusine cascade which is related to translational process. In our previous study, we showed that overexpression of the deoxyhypusine synthase (DHPS), an enzyme contained in the hypusine cascade, promoted cancer progression such as migration and invasion ability through RhoA signaling pathway. A DHPS inhibitor, GC7, showed significant decrease in the tumor formation in vivo. In particular, eukaryoutec initiation factor 5A (eIF5A) which is activated by hypusination has important roles of the translational regulation such as initiation and elongation of protein synthesis and nucleocytoplasmic transport of mRNA. We tried to identify the target genes of eIF5A by combining RNA-ChIP method with RNA sequencing analysis. As a result, candidate genes which bound to eIF5A were extracted by bioinformatics analysis, and redundant binding motifs for eIF5A in mRNAs emerged. These findings and future analyses would reveal the regulation of translational machinery by eIF5A which could be an important target for cancer therapy. Citation Format: Tomoki Muramatsu, Kousuke Tanimoto, Johji Inazawa. Exploring target genes of eukaryotic initiation factor 5A in cancer using RNA sequence analysis. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1992.