Frequent loss of RUNX3 gene expression in human bile duct and pancreatic cancer cell lines

Abstract

RUNX3, a Runt domain transcription factor involved in TGF-b signaling, is a candidate tumor-suppressor gene localized in 1p36, a region commonly deleted in a wide variety of human tumors, including those of the stomach, bile duct, and pancreas. Recently, frequent inactivation of RUNX3 has been demonstrated in human gastric carcinomas. In this study, to examine the involvement of RUNX3 abnormalities in tumorigenesis of bile duct as well as pancreatic cancers, we investigated not only the expression but also methylation status of RUNX3 in 10 human bile duct and 12 pancreatic cancer cell lines. Seven (70%) of the bile duct and nine (75%) of the pancreatic cancer cell lines exhibited no expression of RUNX3 by both Northern blot analysis and the reverse transcriptase polymerase chain reaction. All of the 16 cell lines that did not express RUNX3 also showed methylation of the promoter CpG island of the gene, whereas the six cell lines that showed RUNX3 expression were not methylated or only partially methylated in the RUNX3 promoter region. Moreover, treatment with the methylation inhibitor 5 0 -aza-2 0 -deoxycitidine activated RUNX3 mRNA expression in all of 16 cancer cell lines that originally lacked RUNX3 expression. Finally, hemizygous deletion of RUNX3, as detected by fluorescence in situ hybridization, was found in 15 of the 16 cancer cell lines that lacked RUNX3 expression. These data suggest that the inactivation of RUNX3 plays an important role in bile duct and pancreatic carcinogenesis, and that methylation is a common mechanism by which the gene is inactivated.

Enhancer of Zeste Homologue 2 (EZH2) Down-regulates RUNX3 by Increasing Histone H3 Methylation

“…line MKN28, breast cancer line MCF-7, prostate cancer line LNCap, colon cancer line DLD1, and pancreatic cancer line MiaPaca2, EZH2 mRNA was suppressed with EZH2 siRNA in all five cancer cell lines, in which RUNX3 is silenced (7,11,12,27,28 (Fig. 1A, f-j).…”

“…2B) (7,11,12,27). We then investigated whether the status of DNA methylation in the RUNX3 promoter region changed after knockdown of EZH2 and resulted in restoration of RUNX3 gene expression.…”

“…RUNX3-R122C is a mutation located in the conserved Runt domain that was discovered in a case of gastric cancer and it abolishes the tumor suppressive activity of RUNX3 (7). Subsequent studies have revealed that RUNX3 inactivation is not limited to gastric cancer, and frequent inactivation of RUNX3 due to DNA hypermethylation has been reported in various other cancers, including lung cancer (8), hepatocellular carcinoma (9), breast cancer (10), colon cancer (11), pancreatic cancer (12), bile duct cancer (12), prostate cancer (13), and laryngeal cancer (10). Thus, RUNX3 is primarily inactivated by epigenetic silencing, rather than by mutations or deletions, suggesting that RUNX3 can be reactivated and serve as a good gene for drug targeting.…”

Transcriptional cross-regulation of RUNX1 by RUNX3 in human B cells

“…RUNX2 (AML-3) is essential in osteogenesis (Komori et al, 1997;Fujita et al, 2004;Yoshida et al, 2004), whereas RUNX3 (AML-2) has important functions in neurogenesis Levanon et al, 2002), growth regulation of gastric epithelial cells (Li et al, 2002), TGF-b signalling (Shi and Stavnezer, 1998;Fainaru et al, 2004) and thymopoiesis (Taniuchi et al, 2002;Woolf et al, 2003). Recent studies have shown that loss of RUNX3 expression, usually through allelic loss and epigenetic changes, is associated with hepatocellular carcinoma, testicular yolk sac tumours, gastric and pancreatic cancers (Guo et al, 2002;Kato et al, 2003;Wada et al, 2004;Xiao and Liu, 2004), RUNX3, therefore, has important tumour suppressor function as an integral part of TGFb apoptotic signalling pathways.…”

RUNX3 Modulates DNA Damage-mediated Phosphorylation of Tumor Suppressor p53 at Ser-15 and Acts as a Co-activator for p53

“…Subsequent studies revealed that the frequent reduction of RUNX3 expression levels is also observed in several human cancers such as lung cancer, breast cancer, colon cancer, pancreatic cancer, and prostate cancer, which might be attributed to promoter hypermethylation (7)(8)(9)(10)(11)(12)(13), indicating that the down-regulation of RUNX3 is not restricted to gastric cancer. Intriguingly, Yano et al (15) demonstrated that, during transforming growth factor-␤-mediated apoptotic cell death, RUNX3 has an ability to transactivate pro-apoptotic Bim (Bcl-2-interacting mediator of cell death) (14) in gastric cancer-derived cell lines.…”