Antisense oligonucleotides to class III β-tubulin sensitize drug-resistant cells to Taxol

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Abstract
A major impediment to the successful use of Taxol in the treatment of cancer is the development of drug resistance. The major cellular target of Taxol is the microtubule that is comprised of α- and β-tubulin heterodimers. Binding sites for Taxol have been delineated on the β-tubulin subunit that has six isotypes. We have recently described increased expression of the brain-specific human class III β-tubulin isotype, encoded by the Hβ4 gene, in both Taxol-resistant ovarian tumours and non-small-cell lung cancer cell lines. To evaluate directly the role of the class III β-tubulin isotype in mediating Taxol resistance, antisense phosphorothioate oligodeoxynucleotides (ODN) targeted against various regions of the Hβ4 gene have been designed and examined for their efficacy in reducing Hβ4 gene and protein expression. Taxol-resistant lung cancer cells, A549-T24, which are 17-fold resistant to Taxol and display a fourfold increase in Hβ4 expression compared to the parental A549 cells, were treated with 1 μM antisense ODNs. Two ODNs, AS1 and AS3, were found to reduce mRNA expression by 40–50%, as determined by reverse transcription polymerase chain reaction. A concentration-dependent reduction in Hβ4 mRNA expression was demonstrated with AS1 ODN. Immunofluorescence staining of cells treated with AS1 ODN revealed a decrease in class III protein expression which corresponded to a 39% increase in sensitivity to Taxol (P < 0.005). These findings support an important role for Hβ4 (class III) β-tubulin expression in Taxol resistance and have potential implications for the treatment of Taxol-resistant tumours.