Abstract 3431: βIII-tubulin in glioblastoma: An emerging multifactorial survival factor role in chemotherapy response and tumor formation.
Authors: Gorjana Mitic, Maria Tsoli, David S. Ziegler, Maria Kavallaris
Journal: Cancer Research (2013)

Abstract Glioblastoma (GBM) is an aggressive primary brain tumour with a very poor prognosis. Current treatment is usually surgical resection, radiotherapy and chemotherapy, however, GBM's highly resistant nature renders treatment ineffective. ßIII-tubulin is a neuronal specific microtubule protein that is aberrantly expressed in GBM and it has been implicated in a range of epithelial cancers with poor prognosis and high aggressiveness1. In gliomas, its high expression correlates with higher tumour grade2, however the functional relevance of this expression is unknown. Our aim was to investigate the functional and mechanistic role of ßIII-tubulin in GBM drug sensitivity and tumorigenesis. Methods: To study the functional role of βIII-tubulin in GBM, siRNA gene knockdown was used in two GBM cell lines, U87 and U87vIII and in a primary GBM neurosphere culture. Non-targeting siRNA was used as a control. Knockdown levels were assessed by western blotting. Apoptosis was measured by AnnexinV staining. Drug sensitivity of cell lines was tested using drug-treated clonogenic assays. Senescence was assessed by using cytoplasmic staining for sa-β-galactosidase. Neurosphere formation was assessed by soft-agar growth assay. Results: Specific βIII-tubulin knockdown achieved a 94% and 52% reduction in ßIII-tubulin levels in GBM cell lines and neurospheres respectively, compared to control siRNA. βIII-tubulin knockdown in U87vIII cells resulted in a significant increase in sensitivity to both the DNA-damaging agent temozolomide (TMZ) and tubulin binding agents epothilone B and paclitaxel compared to control (p&amp;lt;0.01). Increased sensitivity to TMZ following βIII-tubulin knockdown in U87 and U87vIII cells was associated with a significant increase in drug-induced apoptosis (p&amp;lt;0.01). TMZ is known to induce senescence3,4 and βIII-tubulin knockdown significantly increased TMZ-induced senescence in U87 (βIII-tub siRNA: 84.1±2.6%, p&amp;lt;0.05, compared to ctrl siRNA: 58.1±7.1%) and U87vIII (βIII-tub siRNA: 83.9±4.5%, p&amp;lt;0.01, compared to ctrl siRNA: 56.3±4.3%) cells. Further validation using a 3D-growth model of primary human GBM neurospheres, showed that neurosphere formation was significantly reduced (30.7%, p&amp;lt;0.01) upon βIII-tubulin knockdown compared to ctrl siRNA. Conclusions: Our data strongly suggests that aberrant expression of βIII-tubulin in glioblastoma may play an important role in intrinsic drug sensitivity to broad classes of chemotherapy drugs. Targeting βIII-tubulin increases temozolomide sensitivity by enhancing drug-induced apoptosis and senescence. Furthermore, we have identified a potential role for βIII-tubulin in GBM tumorigenesis. 1 Kavallaris. Nature Rev Cancer, 10:194-204, 2010 2 Katsetos et al. Arch Pathol Lab Med, 125:613-624, 2001 3 Mhaidat. Br J Cancer, 97:1225-1233, 2007 4 Gunther. Br J Cancer, 88:463-469, 2003 Citation Format: Gorjana Mitic, Maria Tsoli, David S. Ziegler, Maria Kavallaris. βIII-tubulin in glioblastoma: An emerging multifactorial survival factor role in chemotherapy response and tumor formation. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3431. doi:10.1158/1538-7445.AM2013-3431