Abstract 763: Resistance to cabazitaxel is associated with ABCB1/P-glycoprotein activation, alterations in β-tubulin content and dynamics, reduced BRCA1, and a mesenchymal phenotype in MCF-7 human breast cancer variants

Abstract

In order to identify determinants of cellular response to this new taxane, we derived a resistant variant from the MCF-7 human breast cancer cell line by stepwise selection in cabazitaxel. The 33-fold resistance to cabazitaxel in this variant, MCF-7/CTAX, was associated with ABCB1/P-glycoprotein (P-gp) activation, but also 3-fold residual resistance to taxanes after modulation with the P-gp inhibitor PSC-833, 2 μM. Another variant was established by co-selecting in the presence of PSC-833 (MCF-7/CTAX-P). This variant was negative for ABCB1 transcripts and accumulated parental levels of rhodamine-123, BODIPY-labeled paclitaxel and [3H]-docetaxel, indicating that the taxane resistance (9-fold) observed in this cell line is not mediated by transporters. However, we observed a 34% reduction in bound fluorescent-labeled paclitaxel in MCF-7/CTAX-P relative to parental controls by flow cytometry. These cells are hypersensitive to depolymerizing agents (vincas and colchicine) indicating a change in tubulin dynamic instability, and we observed reduced baseline tubulin polymer in untreated MCF-7/CTAX-P cells, and impaired tubulin polymerization in response to taxane exposure (cabazitaxel or docetaxel) relative to MCF-7 parental cells. Quantitative PCR and immunoblotting confirmed elevated levels of the class III (TUBB3) β-tubulin isotype in both MCF-7/CTAX and MCF-7/CTAX-P. Reduced BRCA1 content was observed in both MCF-7 variants, which could affect taxane response via the regulation of the mitotic spindle checkpoint. Gene silencing with specific siRNAs confirmed that reduced TUBB3 sensitizes cells to cabazitaxel, and reduced BRCA1 results in taxane resistance. In addition, altered epithelial-mesenchymal transition markers (elevated Vimentin, reduced E-cadherin) are associated with cabazitaxel resistance in these MCF-7 variants. In summary, cabazitaxel resistance mechanisms include MDR (although at a lower level than paclitaxel and docetaxel), and alterations in microtubule dynamicity, as manifested by higher expression of TUBB3, decreased BRCA1, and by the induction of EMT. Citation Format: George E. Duran, Yan C. Wang, E Brian Francisco, Francisco J. Martinez, Branimir I. Sikic. Resistance to cabazitaxel is associated with ABCB1/P-glycoprotein activation, alterations in β-tubulin content and dynamics, reduced BRCA1, and a mesenchymal phenotype in MCF-7 human breast cancer variants. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 763. doi:10.1158/1538-7445.AM2014-763