Wanshura, Leah E. Colvin

Abstract Activation of the Sonic hedgehog (Shh) pathway and increased expression of Gli1 play an important role in proliferation and transformation of granule cell progenitors (GCP) in the developing cerebellum. Medulloblastomas arising from cerebellar GCPs are frequently driven by Shh pathway–activating mutations; however, molecular mechanisms of Shh pathway dysregulation and transformation of neural progenitors remain poorly defined. We report that the transcription factor and oncogene Snail1 (Sna1) is directly induced by Shh pathway activity in GCPs, murine medulloblastomas, and human medulloblastoma cells. Enforced expression of Sna1 was sufficient to induce GCPs and medulloblastoma cell proliferation in the absence of Shh/Gli1 exposure. In addition, enforced expression of Sna1 increased transformation of medulloblastoma cells in vitro and in vivo. Analysis of potential Sna1 targets in neural cells revealed a novel Sna1 target, N-Myc, a transcription factor known to play a role in Shh-mediated GCP proliferation and medulloblastoma formation. We found that Sna1 directly induced transcription of N-Myc in human medulloblastoma cells and that depletion of N-Myc ablated the Sna1-induced proliferation and transformation. Taken together, these results provide further insight into the mechanism of Shh-induced transformation of neural progenitor cells and suggest that induction of Sna1 may serve to amplify the oncogenic potential of Shh pathway activation through N-Myc induction. Cancer Res; 71(15); 5336–45. ©2011 AACR.

Abstract Activation of the Sonic Hedgehog (SHH) pathway and increased expression of the Gli1 transcription factor have been implicated in the tumorigenesis of both sporadic and syndromic medulloblastoma, a malignant pediatric brain tumor that arises from cerebellar progenitor cells. While SHH pathway activity is required for normal neural stem cell proliferation, molecular mechanisms of SHH pathway dysregulation and transformation of neural precursor cells remain poorly defined. In epithelial cells, Snail1 was found to be induced by Gli1 expression and is believed to promote Gli1-mediated transformation (Li et al. 2006), however the role of Snail1 in neural cells is not well understood. We have identified Snail1 as a putative target of Gli1 in neural stem cells. We hypothesize that Snail1 participates in transduction of SHH/Gli1 signaling in the nervous system and plays a role in proliferation of neuronal progenitor cells in the cerebellum. Mice haploinsufficient for Patched (Ptc) have constitutive activation of the SHH pathway and a subset develop medulloblastoma. Quantitative RT-PCR analyses of mRNA isolated from early postnatal untransformed cerebella show that Snail1 mRNA is increased in vivo in Ptc+/- mice versus wild-type (WT) siblings.. Enforced expression of Gli1 in cultured WT murine granule precursor cells (GPCs) induced Snail1 mRNA.. Enforced expression of Snail1 was also sufficient to induce proliferation of GCPs in the absence of SHH/Gli1 exposure. Given that activation of SHH/Gli1-pathway induces proliferation of GPCs and is associated with medulloblastoma formation, we hypothesized that Snail1 may play a role in the proliferation and transformation of neural progenitor cells. We have found that the stem-like cells generated from medulloblastomas in Ptc+/- mice express higher levels of Snail mRNA than tumor parenchyma. Quantitative RT-PCR analysis demonstrates that Snail1 mRNA is expressed at baseline and is dramatically induced in human medulloblastoma cell lines after exposure to SHH. Enforced expression of Snail1 further increased proliferation of medulloblastoma cells as demonstrated by EdU incorporation. Analysis of potential Snail1 targets in human medulloblastoma cells reveals that β-catenin and N-Myc protein are induced by Snail1 overexpression. N-Myc is induced by SHH in GPCs (Kenney et al. 2003) and is known to induce proliferation in GPCs during both normal cerebellar development and medulloblastoma formation. Together these results give further insight into the mechanism of SHH-induced proliferation of neural progenitor and suggest that Snail1 may contribute to malignant transformation of GCPs through N-Myc. These data establish Snail1 as a target of SHH/Gli1 pathway activity in both normal and neoplastic cells derived from cerebellum. Ongoing experiments will further elucidate how this novel proliferative process affects transformation of neural cells. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 7.