Tubulin-Targeted Drug Action: Functional Significance of Class II and Class IVb β-Tubulin in Vinca Alkaloid Sensitivity

Aberrant expression of beta-tubulin isotypes is frequently described in tumor tissues and tubulin-binding agent (TBA)-resistant cell lines. There is limited understanding of the role of specific P-tubulin isotypes in cellular sensitivity to TBAs, and to gain insights into the functional role of beta II- and beta IVb-tubulin, we examined these isotypes in lung cancer cell lines NCI-H460 (H460) and Calu-6. Drug-treated clonogenic assays revealed that small interfering RNA-mediated knockdown of either beta II- or beta IVb-tubulin hypersensitized the lung cancer cell lines to Vinca alkaloids, with the effects more pronounced following beta IVb-tubulin knockdown. In contrast, there was no change in paclitaxel sensitivity following knockdown of either isotype. Cell cycle analysis revealed a greater propensity for the beta II- and beta IVb-tubulin knockdown cells to undergo G-M cell cycle block following 5 nmol/L vincristine treatment, with the beta IVb knockdown cells being more sensitive than the beta II-tubulin knockdown cells compared with control. In contrast to beta II-tubulin knockdown, beta IVb-tubulin knockdown cells showed a significant increase in the sub-G, population (cell death) following treatment with both 5 and 40 nmol/L of vincristine compared with controls. Importantly, PtVb-tubulin knockdown in H460 cells caused a significant dose-dependent increase in Annexin V staining in response to vincristine but not paclitaxel. Therefore, increased sensitivity to induction of apoptosis is one mechanism underlying the Vinca alkaloid hypersensitivity. This study provides direct evidence that beta II- or beta IVb-tubulins have functionally distinct roles and expression of these isotypes may serve as strong predictors of Vinca alkaloid response and resistance. [Cancer Res 2008;68(23):9817-24]