Abstract 4270: Heterogeneous radiosensitivity among phenotypes within intracerebral xenografts initiated from glioblastoma stem like cells

Abstract

Abstract Intra-tumor heterogeneity is a well established characteristic of human glioblastoma (GB), yet whether this heterogeneity extends to cellular radiosensitivity has not been clearly defined. In contrast to established glioma cell lines, brain tumor xenografts initiated from GB stem like cells are comprised of a mixture of stem cells, progenitors and cells in various states of differentiation, thus providing a clinically relevant model system for defining radiosensitivity as a function of phenotype. Of particular interest is the radiosensitivity of stem like versus non-stem like GB cells. To address this issue, CD133+ stem like cells isolated from a GB neurosphere culture were used to initiate brain tumors in nude mice; we then compared the in vivo radiosensitivity of GB stem like cells (CD133+) versus that of non-stem like GB cells (CD133-). To define radioresponse on an individual cell basis, we used γH2AX foci, which provides an indicator of DNA double strand break (DSB) induction and repair. At the onset of tumor induced morbidity, mice were irradiated (6Gy) and tumors collected at times out to 24h. The maximum number of γH2AX foci induced occurred at 0.5h followed by a rapid reduction to approach control levels by 6h after irradiation. Using dual label immunohistochemistry, γH2AX induction and dispersal were defined specifically in CD133+ and CD133- cells within a given tumor. Whereas each phenotype had a similar time course of foci induction and dispersal, the maximum number of foci induced in CD133+ cells was significantly less than that detected in CD133- tumor cells. Similar results were obtained when radiation induced 53BP1 nuclear foci were analyzed. These data indicate that the initial level of DSBs induced in CD133+ cells is significantly less than that in CD133- cells, which suggests that the stem like phenotype is relatively radioresistant. As an additional approach to evaluating the putative phenotypic difference in radiosensitivity, tumors were irradiated (12Gy) on day 21 post-implantation and % CD133+ cells determined at the time of irradiation and at the onset of morbidity. On the day of irradiation, >60% of the cells within brain tumors initiated from CD133+ cells retained CD133 expression and at morbidity tumors in unirradiated mice contained approximately 10% CD133+ cells. In irradiated mice, however, tumors collected at morbidity contained greater than 20% CD133+ cells. Thus, based on the analysis of individual cell radiosensitivity (nuclear foci) and the relative increase in CD133+ cells, these in vivo results suggest that within this brain tumor xenograft model the stem cell like phenotype comprises a radioresistant subpopulation. Moreover, this study illustrates the potential for combining γH2AX foci and phenotype immunohistochemical analyses as an approach for investigating the intra-tumor heterogeneity in radiosensitivity. 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 4270.