Abstract B186: Knockdown of a chromatid segregation mediator gene impairs tumor growth in glioblastoma multiforme.

Abstract

Abstract Glioblastoma Multiforme (GBM) is the most common primary brain malignancy and despite multimodal therapy including surgery, irradiation and chemotherapy median survival remains 12–15 months. Increasing body of evidence from our and other groups is confirming the existence of a stem-like cell population within these tumors believed to confer tumor growth, invasiveness, and therapy resistance. Adult human neural stem cells (aHNSCs) are suggested to be the cells of origin for the glioblastoma stem-like cells (GSCs). To identify new molecular targets we performed a micro-array analysis on the transcriptome of 33000 genes identifying differentially expressed genes in 5 aHNSC and 9 GSC cultures. Amongst the genes highly up-regulated in GBM, several were involved in cell division and the regulation of cell cycle. For further analysis, we chose the Chromatid Segreration Mediator (CSM), a centromere specific gene important for appropriate chromatid segregation during mitosis. This candidate gene was strongly expressed in GSCs but was almost undetectable in aHNSCs. Abundance of the CSM protein may lead to aneupleudy, which is a common feature of GBMs. The high levels of CSM mRNA were confirmed by analyzing 16 GSC cultures with micro-array and qPCR. CSM was also over-expressed on protein level, and was found to be highly immunopositive in glioblastoma biopsies compared to the normal human brain. In silico analysis, using Rembrandt, identifies CSM-gene over-expression as a negative prognostic indicator in glioma patients. In order to study the functional role of this gene in GBM, we used lentiviral-based knock-down (KD) technology to obtain specific down regulation of the gene. We transduced different shRNA constructs in 2 GBM cultures and achieved KD-efficieny of 55–65%. Growth of tumors was measured by proliferation assay and the relative number of sphere-forming cells (widely known as GSCs) was quantified with a sphere-formation assay. In our KD-cultures we observed a moderate reduction of proliferation (25%). Interestingly, the sphere forming capacity was dramatically reduced by 60–70%, indicating a significant decrease in number of GSCs. Cell-cycle analysis revealed that it was 30–40% less cells in G2/M-phase and 15–20% more cells in G0/G1-phase in the KD compared to non-silencing control. We find our results very promising and suggest that Chromatid Segregation Mediator gene might have potential as a novel therapeutic target for GBM management. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2011 Nov 12-16; San Francisco, CA. Philadelphia (PA): AACR; Mol Cancer Ther 2011;10(11 Suppl):Abstract nr B186.