Sensitization of human breast cancer cells to gemcitabine by the protein kinase C modulator bryostatin 1

Purpose: Protein kinase C (PKC) plays an important role in cell proliferation, differentiation, and apoptosis. The interaction between the PKC modulator bryostatin 1 (BRYO), and gemcitabine in human breast cancer MCF-7 and MDA-MB-231 cells and in the non-transformed MCF-10A human breast epithelial cells was investigated. Methods: Immunoblotting was used to determine the expression of PKC isoenzymes and proteins involved in the cell cycle and apoptosis. MTT, ELISA and flow cytometry assays were used to determine cell survival. Results: Treatment of cells with BRYO 200 nM resulted in a significant downregulation of cytoplasmic PKC in all three cell lines. However, the expression of membranous PKC was differentially affected in these cells. BRYO (1-200 nM) had no significant effects on cell viability in any of the cell lines. Nevertheless, BRYO significantly enhanced the anti-proliferative and apoptotic effects of gemcitabine in the MCF-7 and MDA-MB-231 cells, but not in the MCF-10A cells. This was associated with significant reduction in the bcl-2/bax ratio. There was a significant upregulation of p53, p21 ^waf1, and p27 in MCF7 and MCF-10A cells treated with the combination of gemcitabine and BRYO compared to gemcitabine-treated cells. Conclusions: The potentiation of the effect of gemcitabine by BRYO was demonstrated in MCF-7 and MDA-MB-231 cells and was associated with a specific pattern of PKC modulation. Further investigation of the role of specific isoforms of PKC in the downstream molecular events of gemcitabine-induced cytotoxicity is warranted.