Downregulation of RUNX3 and TES by hypermethylation in glioblastoma

Abstract

Glioblastoma, the most aggressive and least treatable form of malignant glioma, is the most common human brain tumor. Although many regions of allelic loss occur in glioblastomas, relatively few tumor suppressor genes have been found mutated at such loci. To address the possibility that epigenetic alterations are an alternative means of glioblastoma gene inactivation, we coupled pharmacological manipulation of methylation with gene profiling to identify potential methylation-regulated, tumor-related genes. Duplicates of three short-term cultured glioblastomas were exposed to 5 lM 5-aza-dC for 96 h followed by cRNA hybridization to an oligonucleotide microarray (Affymetrix U133A). We based candidate gene selection on bioinformatics, reverse transcription-polymerase chain reaction (RT-PCR), bisulfite sequencing, methylationspecific PCR and matrix-assisted laser desorption/ ionization time-of-flight mass spectrometry. Two genes identified in this manner, RUNX3 and Testin (TES), were subsequently shown to harbor frequent tumor-specific epigenetic alterations in primary glioblastomas. This overall approach therefore provides a powerful means to identify candidate tumor-suppressor genes for subsequent evaluation and may lead to the identification of genes whose epigenetic dysregulation is integral to glioblastoma tumorigenesis.

RUNX3 is down‐regulated in glioma by Myc‐regulated miR‐4295

“…Prior reports have suggested that RUNX3 is frequently inactivated in human cancer cells and can be activated by hemizygous deletion of the RUNX3 gene, hypermethylation of the Runx3 promoter, or cytoplasmic sequestration of RUNX3 protein 32, 34. Furthermore, accumulating evidence demonstrated RUNX3 could inhibit the proliferation, tumourigenic and metastasis of glioma cells 21.…”

RUNX3 is multifunctional in carcinogenesis of multiple solid tumors

“…Diverse tumor tissues have been reported to exhibit RUNX3 hypermethylation and inactivation. These include tissues and cell lines that originated from gastric (Li et al, 2002;Oshimo et al, 2004), bladder (Kim et al, 2005;Wolff et al, 2008), colorectal (Ahlquist et al, 2008;Soong et al, 2009;Subramaniam et al, 2009), breast (Lau et al, 2006;Jiang et al, 2008), lung (Sato et al, 2006), pancreatic (Wada et al, 2004), brain cancers (Mueller et al, 2007), and hepatocellular carcinoma . Notably, RUNX3 methylation status is one of the five markers used to classify colorectal tumors associated with very high frequencies of CpG island methylation (CIMP), microsatellite instability, and BRAF mutation (Weisenberger et al, 2006).…”

Frequent biallelic inactivation and transcriptional silencing of the DIRAS3 gene at 1p31 in oligodendroglial tumors with 1p loss

“…18 A 324 bp-fragment covering all 22 CpG sites in CpG island I, a 379 bp-fragment covering all 21 CpG sites in CpG island II, as well as a 302-bp fragment covering 19 out of 29 CpG sites in CpG island III were individually amplified by PCR from sodium bisulfite-modified DNA (for primers see Table I). The respective PCR products were cloned using One Shot Escherichia coli cells and the TOPO TA Cloning Kit (Invitrogen Life Technologies, Carlsbad, CA).…”