Protein kinase C isoform expression and activity alter paclitaxel resistance in vitro

Objective. The aim of this study was to assess the relationship of protein kinase C (PKC) isoform expression and functional activity to the developmen t of multidrug resistance in gynecologic malignancies. Methods. Paclitaxel-resistant subclones (T30 and T30-Res) of the Mes-sa hum an uterine sarcoma cell line were selected through exposure to paclitaxel i n vitro. Indices of relative drug resistance were determined by the MTT (3- [4,5-dimethyl-2-thiazolyl]-2,5-diphenyl-2H-tetrazolium bromide) assay. Diff erences in the expression pattern of PKC isoforms were assessed by Western blot of cell lysates. Finally, the influence of PKC activity (i.e., translo cation to the plasma membrane, confirmed by Western blot of plasma membrane bound protein) on resistance to paclitaxel was examined with the MTT assay in cells preincubated with PMA. Results. The indices of relative paclitaxel resistance of Mes-sa, Mes-sa-T3 0, and Mes-sa-T30-Res were 1-, 5-, and 11-fold, respectively. Five (alpha, gamma, iota, lambda, and mu) of the 11 known PKC isoforms were detected in all cell lysates. Only PKC-alpha and PKC-gamma expression increased with in creasing indices of paclitaxel resistance. Interestingly, PMA induction of PKC activity reversed resistance to paclitaxel in all cell lines by 2- to 3 -fold, and this reversal of drug resistance was associated with a time-depe ndent translocation of PKC-alpha and PKC-gamma to the plasma membrane compa rtment. Conclusions. Increased expression of only the PKC-alpha and PKC-gamma isofo rms correlates with increasing levels of paclitaxel resistance in Mes-sa ce lls in this in vitro experimental model. However, increased functional acti vity of these and other PKC isoforms leads to reversal in paclitaxel resist ance. Therefore, PKC activating mechanisms normally present in primary tumo r cells may be compromised in drug-resistant clones, (C) 1999 Academic Pres s.