Abstract PR13: Epigenetic regulation of MYC drives dynamic transition between tumor initiating states in glioblastoma
Authors: Kozono David , Li Jie , Nitta Masayuki , Sampetrean Oltea , Ng Kimberly , Gonda David , Kushwaha Deepa , Hiroko Matsui , Harismendy Olivier , Becher Oren , Kwon Chang-Hyuk , Ligon Keith L. , Saya Hideyuki , Carter Bob S. , Pizzo Donald , Vandenberg Scott , Chen Clark C. 
Published: May 2015

Glioblastoma is the most common form of brain cancer and remains a devastating disease. Recent studies revealed significant intra-tumoral heterogeneity in the genetic and epigenetic make-up of glioblastomas. One level of heterogeneity involves subpopulations of cells capable of tumor initiation (TI). Here we provide data suggesting that the transition between TI- and non-TI is a dynamic process governed by spontaneous fluctuation in the level of MYC. In vitro culturing of sub-clones derived from long-term passaged and primary glioblastoma lines revealed that only a subset of the sub-clones possess TI capacity. The property of TI for each individual sub-clone appeared stable through serial passages. However, a small fraction of the non-TI clones will spontaneously acquire the TI ability. This phenomenon was observed both in vitro and in vivo.

Transcriptome profiling of the sub-clones with varied TI capacity revealed a gene signature enriched for genes regulated by MYC. Among various sub-clones, MYC expression levels correlated with TI capacity tightly. More direct evidence was provided by MYC overexpression which augmented TI capacity in both xenograft and genetic murine models. Reversely, MYC silencing abolished glioblastoma capacity for TI. Importantly, the sub-clones that spontaneously acquired capacity exhibited enhanced MYC expression.

In freshly resected glioblastoma specimens, overall MYC expression levels of the specimens correlated with their xenograft-forming ability. When these specimens were sub-fractionated by A2B5, a cell surface marker enriched in TI glioblastoma populations, the MYC level was significantly elevated in the A2B5+ fraction relative to the A2B5- fraction. In The Cancer Genome Atlas (TCGA) glioblastoma specimens, the expression level of a MYC signature directly correlated with mRNA signatures associated with the Cancer Stem Cell states. In dual immunofluorescence staining of clinical glioblastoma specimens, C-MYC co-stained with MIB1, suggesting that MYC expression support in vivo tumor proliferation.

Since the various sub-clones were genetically identical based on SNP array profiling, we hypothesized that fluctuations in MYC expression was regulated through epigenetic regulation. Supporting our hypothesis, the primary glioblastoma tumor lines that demonstrated high MYC levels showed higher ratios of H3K4me3 to H3K27me3 at the MYC locus. Culturing conditions that enhanced TI capacity of glioblastoma cell lines also increased the ratio of H3K4me3 to H3K27me3 and induced MYC expression.

In sum, our results suggest a threshold model in which TI capacity is driven by epigenetic regulation of MYC. MYC inhibition constitutes an attractive therapeutic target since this inhibition reduces dynamic cell state transition and reduce the complexity of the tumor heterogeneity.

This abstract is also presented as Poster B75.

Citation Format: David Kozono, Jie Li, Masayuki Nitta, Oltea Sampetrean, Kimberly Ng, David Gonda, Deepa Kushwaha, Matsui Hiroko, Olivier Harismendy, Oren Becher, Chang-Hyuk Kwon, Keith L. Ligon, Hideyuki Saya, Bob S. Carter, Donald Pizzo, Scott Vandenberg, Clark C. Chen. Epigenetic regulation of MYC drives dynamic transition between tumor initiating states in glioblastoma. [abstract]. In: Abstracts: AACR Special Conference on Cellular Heterogeneity in the Tumor Microenvironment; 2014 Feb 26-Mar 1; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2015;75(1 Suppl):Abstract nr PR13. doi:10.1158/1538-7445.CHTME14-PR13