Background: Glioblastoma is an aggressive malignant brain tumour with a dismal prognosis. Current treatments are ineffective and offer no realistic chance of cure. ßIII-tubulin is a neuronal specific β-tubulin isotype, a structural component of microtubules, that is not expressed in glial cells. Its aberrant expression in various cancers is associated with drug resistance and poor clinical outcomes (1). In gliomas, high ßIII-tubulin expression correlates with higher tumour grade, however its role in the biology of gliomas is unknown. The aim of this study was to investigate the role of ßIII-tubulin in glioblastoma drug resistance and tumourigenesis.
Results: Two glioblastoma cell lines, U87 and U87vIII, were selected for functional studies. To examine the role of βIII-tubulin in glioblastoma chemo-sensitivity and tumourigenesis, siRNA gene silencing was used. Potent and specific knockdown achieved a 94% reduction in ßIII-tubulin levels compared to non-targeting control siRNA. Drug-treated clonogenic assays using βIII-tubulin knock-down cells resulted in a significant increase in sensitivity to DNA-damaging agent temozolomide and tubulin-binding agents epothilone B and paclitaxel, in U87vIII cells (p<0.01) compared to control siRNA transfected cells. Additionally, knockdown of βIII-tubulin led to increased apoptosis after temozolomide treatment compared to control siRNA.
To examine the potential role of ßIII-tubulin in glioblastoma tumourigenicity, anchorage-independent growth (soft agar assay) was used. A significant decrease (p<0.01) in anchorage-independent growth in U87vIII cells was observed following βIII-tubulin knock-down. Further studies are currently ongoing to examine the effects of βIII-tubulin knockdown in U87 cells and on primary glioblastoma neurosphere formation.
Conclusion: βIII-tubulin is expressed in glioblastoma cell lines and its expression contributes to intrinsic drug resistance to two different classes of drugs: DNA-damaging and tubulin-binding agents. Our data suggests that expression of βIII-tubulin may also be contributing to tumourigenic phenotype of this devastating disease.
1. Kavallaris M. Microtubules and resistance to tubulin-binding agents. Nat Rev Cancer 2010; 10: 194-204.