Abstract 3317: Radiation induced multinucleated giant cells: A novel therapeutic target to prevent survival and relapse of glioblastoma

Abstract

Abstract In this study, we address the fundamental issue of radiation resistance in Glioblastoma Multiforme (GBM) a highly malignant form of brain tumor. However, because of the unavailability of multiple biopsies of GBM patients after treatment, we are limited in our understanding of resistance mechanisms. To circumvent this problem we recapitulated clinical scenario of GBM resistance in a cellular model developed from fresh naive primary GBM patient samples and cell lines. Using our model we show that upon lethal dose of radiation, small percentage of GBM cells survive, are non-apoptotic and transiently become non- proliferative. They are arrested in G2/M phase of cell cycle mediated by inhibitory phosphorylation of Cdk1(Y15) and p21. Surprisingly, we find although non -proliferative, these cells are highly motile and undergo homotypic cell-cell fusions at high frequency to form multinucleated and giant cells (MNGCs). Cell fusion leads to the induction of senescence as seen by β-galactosidase staining and accordingly high expression of SASPs (senescence associated secretory proteins like GM-CSF, SCF, IL-6 and IL-8) is also observed. SASPs provide survival and proliferation signals to the cells in an autocrine manner. Accordingly, we find enhanced expression of anti-apoptotic genes BIRC3 and Bcl-xL and unaltered levels of Bax (pro-apoptotic protein) mRNA in the resistant cells. MNGCs also activate AKT, which is a survival signal and a central convergence node of signals downstream to SASPs. AKT activation along with Bcl-xL is reported to regulate apoptosis synergistically. Additionally, resistant cells also undergo active DNA damage repair mediated by sensory kinases ATM and ATR. Interestingly, irrespective of the phase of cell cycle, they adopt Non Homologous End Joining (NHEJ) as their pathway of choice to repair their double strand breaks in DNA. Since we find increased presence of H3K36me2 marks known to modulate NHEJ, a possible link between NHEJ and this modification in GBM is worth exploring. Finally, unlike previous reports we demonstrate that MNGCs do not die of mitotic catastrophe instead escape senescence, undergo normal cell division and proliferation giving rise to mono-nucleated relapse cells with unaltered ploidy compared to parent cells. Importantly, we also show the potential ability to prevent the relapse in Glioma by disrupting the non-proliferative state of radiation resistant cells using mitotic inducer (MK1775) and selectively ablating growth of MNGCs by cytokinesis inhibitor. In summary, our data provides a mechanistic explanation for clinical observations and novel insights into an unexplored multi-step process of radiation survival and relapse in Glioblastoma We identified homotypic cell fusions of resistant Glioma cells as a novel non-genetic mechanism to sustain survival and relapse and generate rationale for novel combinatorial targeted therapies. Citation Format: Ekjot Kaur, Jacinth Rajendra, Sanket Shah, Jyothi Nair, Ankit Khushwaha, Aliasgar Moiyadi, Shilpee Dutt. Radiation induced multinucleated giant cells: A novel therapeutic target to prevent survival and relapse of glioblastoma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3317. doi:10.1158/1538-7445.AM2015-3317