Abstract 2484: Aurora B silencing attenuates radiation resistance through PARP and identification of a missense mutation in aurora B as a determinant of radiosensitization in gliomas
Authors: Kamalakannan Palanichamy, Disha Patel, Michael Erkkinen, Zachary Reardon, Shin-ei Noda, Arnab Chakravarti
Journal: Cancer Research (2011)

Abstract Purpose: Aurora B (Aur B), a mitotic protein located on chromosome 17p13.1, is critical for proper chromosome segregation and changes in chromosome 17p are quite common in glioblastoma multiforme (GBM). Upregulation of Aur B induces multicellularity and fosters polyploidy. Aur B reaches maximum levels later in mitosis and regulates the chromosomal passenger complex progression. It has been reported that Aur B expression markedly correlates with short survival of GBM patients and may be a prognostic factor. Targeting Aur B is an ideal strategy, since it is active during mitosis and non-proliferating cells would not be adversely affected. Materials and Methods: The cell lines used for this study include both established and primary GBM cell lines. Radiation experiments were conducted using gammacell irradiator. DNA damage was estimated using comet assay. Apoptosis, cell cycle and multinucleated polyploid cells were estimated using flow cytometry. Gene silencing and overexpression were carried out using established protocols. Results and Discussion: Aur B expression increases after radiation in GBM cells and not in normal human astrocytes as determined both at transcriptional and translational levels. The radiosensitivity of cells lines was estimated by clonogenic survival assay. Through sequencing Aur B, we discovered mutations at the 885(C) and 893(T) residues in the GBM cell lines. The 885(C) was a silent mutation and 893(T) leads to an amino acid change from methionine (M) to threonine (T). This missense mutation leads to a new phospho site. The GBM cell lines tested had heterozygous and homozygous deletions. The radiosensitivity assay conducted on these cell lines shows a differential response with regards to the nature of mutation. The cell lines which had homozygous deletions on both sites 885(C) and 893(T) exhibit an increased radioresistance under in vitro conditions when compared to one of them being a heterozygote. Further, we show that Aur B silencing inhibits cytokinesis leading to 4N or 8N DNA content in p53 functional cell lines. The endoreduplicated cells due to Aur-B inhibition leads to the loss of viability in p53 mutant/null cell lines. Overexpression of Aur B increases the DNA repair enzyme PARP and scaffolding protein XRCC1 after radiation, which are critical for the activation and recruitment of Base Excision Repair. Whereas, silencing Aur B decreases the PARP expression levels. Conclusion: In GBM cell lines, we report that Aur B knockdown sensitizes cells to radiation and over expression of Aur B leads to radioresistance. Thus far, one of the prime targets in mediating this effect was found to be PARP, since silencing and forced induction of Aur B correlates with expression levels of PARP. For the first time, we have shown that the mutational status of Aur B alters the radiation sensitivity. Further in vivo studies are underway. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2484. doi:10.1158/1538-7445.AM2011-2484