SC-37 * THE ROLE OF NG2 EXPRESSING PROGENITOR CELLS IN DIFFUSE INTRINSIC PONTINE GLIOMA

Abstract

Pediatric diffuse intrinsic pontine glioma (DIPG) is one of the most difficult cancers to treat. pLys27Met (K27M) driver mutation in the H3F3A (H3.3) and HIST1H3B (H3.1) genes of histone are correlated with a subgroup of DIPGs. Other genomic aberrations include p53 mutations and amplification of signaling pathways including PDGFRα. We have recently reported the involvement of Hedgehog (Hh) pathway in a subset of DIPGs. Modulation of Hh and tyrosine kinase receptors may alter the self-renewal properties of cancer stem cells (CSC). NG2 Proteoglycan positive cells that co-express PDGFRα and Olig-2 are present in adult gliomas, where NG2 contributes to the neoplastic transformation of glioma cells. We examined NG2 expression in frozen brainstem specimens of DIPGs and observed significant NG2 expression in DIPGs [10 of 14 (71 %), fold change = 33, p < 0.05)] as compared to the adjacent normal tissue. NG2 expression was associated with histone 3 K27M mutation [8 of 10 (80 %)]. Two mechanisms of NG2 regulation in DIPG were identified: i) histone 3 binds to NG2 promoter, and ii) miR 129-2 negatively regulates NG2. We detected downregulation of miR129-2 in 85.7% (6 of 7) of DIPG tumors compared to normal tissue (FC = -30.79, n = 7 pairs). We also found overall hypermethylation at 8 CpG loci corresponding to the miR129-2 promoter. NG2 knockdown in vitro (shRNA, miR129-2 or demethylating drugs) retards cellular migration. Luciferase assay in primary mouse glioma cells co-transfected with 3'UTR of NG2 and miR129-2 revealed regulation of NG2 by miR129-2. This was further confirmed in vivo by injection of NG2-dsRed transgenic mice with mir129-2 lentivirus. Orthotopic injection of NG2+ cells results in rapid tumor formation while NG2-KD cells fail to form tumors. Our study offers a potential model for the expansion of tumor stem cells and their self-renewal properties in DIPGs.