PMC

Structures. I: 2-MeOE2, II: STX140, III: STX243, IV: paclitaxel, V: vinorelbine, VI: ENMD1198 and VII: colchicine.

STX140, STX243, 2-MeOE2, paclitaxel, ENMD1198, vinorelbine and colchicine chemosensitivity in MDA-MB-231 cells. MDA-MB-231 cells were transfected with the mock vector or the non-targeting siRNA (control), (A) pALTER-TUBB3 or siRNA-tubb3 and treated with STX140, (B) STX243, (C) paclitaxel, (D) 2-MeOE2, (E) ENMD1198, (F) vinorelbine or (G) colchicine for 4 days. The percent proliferation was determined by AlamarBlue assay and results presented are the average of three or four independent experiments (n=3–4) done in triplicate, ^*P<0.05, ^**P<0.01 and ^***P<0.001.

Fluorescent images of cells overexpressing class III β-tubulin after drug exposure. MCF-7-control, MCF-7-TUBB3, MDA-MB-231-control and MDA-MB-231-TUBB3 cells were treated for 48 h with STX140, paclitaxel or colchicine at concentrations close to IC₅₀ values obtained in control cells and stained with FITC-anti-tubulin (tubulin) and Hoechst 33342 (nucleus). Images were taken using a Zeiss inverted microscope at × 200 magnification.

STX140, STX243, 2-MeOE2, paclitaxel, ENMD1198, vinorelbine and colchicine chemosensitivity in MCF-7 cells. MCF-7 cells were transfected with the mock vector or the non-targeting siRNA (control), (A) pALTER-TUBB3 or siRNA-tubb3 and exposed to STX140, (B) STX243, (C) paclitaxel, (D) 2-MeOE2, (E) ENMD1198, (F) vinorelbine or (G) colchicine for 4 days. The percent proliferation was determined by AlamarBlue assay and results presented are the average of three or four independent experiments (n=3–4) done in triplicate, ^*P<0.05 and ^**P<0.01.

Class III β-tubulin mRNA and protein expression. (A) MCF-7 and MDA-MB-231 cells were transfected with the mock vector or the non-targeting siRNA (control), pALTER-TUBB3 or siRNA-tubb3 and the expression of class III β-tubulin mRNA was quantified by RT–PCR. Three or four independent experiments (n=3–4) were carried out in duplicate and the results presented are the average of the Ct values relative to the control Ct values, bars: mean±s.d., ^*P<0.05 and ^**P<0.01. The expression of class III β-tubulin protein, total β-tubulin and total α-tubulin was analysed in (B) MCF-7 and (C) in MDA-MB-231 24 h and 5 days after transfection by SDS–PAGE and immunoblotting. (1) Control 24 h, (2) pALTER-TUBB3 24 h, (3) siRNA-tubb3 24 h, (4) control 96 h, (5) pALTER-TUBB3 96 h and (6) siRNA-tubb3 96 h.