Abstract 3346: Astrocytoma initiation by Rb family inactivation induced dedifferentiation of mature astrocytes.

Abstract

Abstract Astrocytomas are the most common primary human malignant brain tumors. Despite standard-of-care surgery, radiation, and chemotherapy, no curative treatment for these devastating diseases is currently available. Tumor-initiating cells (TICs), a small population of tumor cells is shown to be responsible for astrocytoma initiation and progression. Though their presence has been convincingly shown, there is considerable debate about their origin. By using genetically engineered mice, we hypothesis that loss of Rb family, induces dedifferentiation of mature astrocytes which are capable of evolving into TICs. Our lab has developed an adult mouse model where inactivation of retinoblastoma (Rb) family proteins by expressing N-terminal 121aa of large T antigen (T121), initiates grade II astrocytoma. pRb inactivation and mutant KrasG12D activation leads to grade III astrocytoma and further PTEN inactivation results in Grade IV glioblastomas. We have utilized the Rb family inactivation model to investigate the dedifferentiation of mature astrocytes. We have performed IHC using progenitor markers, Sox2 and nestin and a proliferation marker Ki67 and observe the presence of these markers coinciding with T121 expression suggesting a possible dedifferentiation of mature astrocytes. Quantitative analysis of the overlap between Sox2 and Ki-67 with T121 indicates that the marker presence is specific to T121 expression. In addition to these markers, we also show that S100α an astrocyte differentiation marker is downregulated in pRb inactivated cells further supporting the hypothesis of possible dedifferentiation of mature astrocytes. Since T121 expression is induced in all GFAP expressing cells, there is a possibility that the GFAP expressing cells in the germinal areas such as SVZ and dentate gyrus could be migrating into the cortex and form the observed progenitor cells. To address this issue we used lenti-Cre to focally induce T121 expression in the cortex. Sox2 expression, along with the reduced expression of differentiation marker S100α in these cells, suggests that the dedifferentiation events are independent of the germinal zones in the adult brain. As a functional assay to identify progenitor/ stem cells we performed neurosphere assay. In our assay conditions, wildtype adult cortex was unable to generate any spheres compared to T121 expressing cells. We found that the longer the post induction time the higher the sphere forming ability of T121 cells. The multilineage ability of these T spheres cells was confirmed by inducing differentiation into different neural lineages. To test the tumor forming ability of the sphere cells, we have performed subcutaneous injections and awaiting the results. Thus we conclude that in our system inactivation of Rb family members induced dedifferentiation in mature astrocytes. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3346. doi:1538-7445.AM2012-3346