Abstract 2982: Reduced expression of histone demethylase KDM6B promotes pancreatic cancer progression through downregulation of C/EBPα.
Authors: Yamamoto Keisuke , Tateishi Keisuke , Kudo Yotaro , Kakiuchi Miwako , Yamamoto Shinzo , Miyabayashi Koji , Asaoka Yoshinari , Ijichi Hideaki , Omata Masao , Koike Kazuhiko 
Published: August 2014

It has become increasingly clear that epigenetic aberrations are involved in the pathogenesis of various cancer. In pancreatic cancer, however, the implication of epigenetic alteration remains elusive. In our screening analysis for epigenetic regulators that affect the phenotype of pancreatic cancer, we identified KDM6B/JMJD3, a H3K27me3 demethylase, as a candidate gene. KDM6B/JMJD3 activates INK4a/ARF locus during oncogene-induced senescence, thus it is supposed to suppress tumorigenesis; however, its role in established cancer remains unclear. In this study, our aim is to clarify the role of KDM6B in the progression of pancreatic cancer and to understand the underlying mechanisms.

We generated pancreatic cancer cell lines stably knocked down for KDM6B by shRNAs. Matrigel invasion assay, soft agar colony formation assay, tumor sphere formation assay revealed increased invasiveness, anchorage-independent growth and tumorigenicity of KDM6B-knockdown (KD) cells, respectively. To confirm these phenotypic changes in vivo, these cells were xenotransplanted into nude mice. In intrasplenic injection experiments, mice injected with KDM6B-KD cells showed significantly shorter survival. When tumor cells were orthotopically implanted into the pancreas, mice injected with KDM6B-KD cells developed massive peritoneal dissemination with hemorrhagic ascites, while no mice injected with control cells developed peritoneal dissemination.

To identify genes responsible for the phenotypic change, we performed cDNA microarray analysis. Gene set enrichment analysis (GSEA) revealed significant expression change of C/EBP-target genes upon KDM6B ablation. Among the C/EBP transcription family members, the downregulation of CEBPA, a putative tumor suppressor gene, was confirmed in KDM6B-KD cells. ChIP assay showed specific increase of H3K27me3 levels in the upstream region of CEBPA gene after KDM6B depletion. Notably, enforced expression of CEBPA rescued the increased invasiveness and tumorigenicity of KDM6B-KD cells, indicating that reduction of KDM6B expression in pancreatic cancer enhances its aggressiveness through downregulation of C/EBPα.

Notably, we identified a cell surface protein reflective of KDM6B expression and tumorigenic potential of pancreatic cancer cells; flow cytometry analysis revealed that cells with higher expression of this molecule expressed lower KDM6B and showed increased tumorigenicity. Immunohistochemical analysis of pancreatic cancer specimen showed a positive correlation between this marker expression and tumor grade, while KDM6B and C/EBPα expression was seldom observed in high grade tumor.

Collectively, these data suggest a role for the KDM6B-CEBPA axis in the regulation of tumorigenictiy of pancreatic cancer cells, providing a link between epigenetic change and pancreatic cancer progression.

Citation Format: Keisuke Yamamoto, Keisuke Tateishi, Yotaro Kudo, Miwako Kakiuchi, Shinzo Yamamoto, Koji Miyabayashi, Yoshinari Asaoka, Hideaki Ijichi, Masao Omata, Kazuhiko Koike. Reduced expression of histone demethylase KDM6B promotes pancreatic cancer progression through downregulation of C/EBPα. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2982. doi:10.1158/1538-7445.AM2013-2982

Note: This abstract was not presented at the AACR Annual Meeting 2013 because the presenter was unable to attend.