Increase of βIII- and βIVa-Tubulin Isotypes in Human Prostate Carcinoma Cells as a Result of Estramustine Resistance

Estramustine (EM), an antimicrotubule agent, is effective against hormone-refractory prostate cancer when used in combination with vinblastine or paclitaxel. To understand the effect of EM on β-tubulin, a cellular target for this class of drugs, human prostate carcinoma cells (DU-145) were made resistant to EM, and two cell lines were selected at 12- (EM-12) and 15-µm (EM-15) concentrations of the drug. These cell lines exhibited 8- to 9-fold resistance to EM and 2- to 4-fold cross-resistance to paclitaxel. Immunofluorescent staining of the cells with β-tubulin isotype-specific antibodies showed a ∼6-fold increase in the βIII-tubulin levels and moderate increase in overall β-tubulin levels in EM-resistant cells when compared to DU-145 cells. This increase of βIII isotype was confirmed by Western analysis. A reverse transcriptase-PCR assay was also employed using β-tubulin isotype-specific primers to quantify β-tubulin isotype RNA. A 4-fold increase in βIII and a 3-fold increase in βIVa transcript were seen in both EM-resistant cell lines. These results indicate that overexpression of specific β-tubulin isotypes may play a role in the cellular defense against EM and other antimicrotubule agents.