Serine protease inhibitor TPCK prevents Taxol-induced cell death and blocks c-Raf-1 and Bcl-2 phosphorylation in human breast carcinoma cells

The mechanism of Taxol-induced apoptosis was investigated in MCF-7 human br east carcinoma cells, Taxol-induced apoptosis was associated with phosphory lation of both c-Raf-1 and Bcl-2 and activation of ERK and JNK MAP kinases, The serine protease inhibitor N-tosyl-L-phenylalanine chloromethyl ketone (TPCK) effectively blocked apoptosis, but N-p-tosyl-L-lysine chloromethyl k etone (TLCK), another serine protease inhibitor, was without effect. TPCK t reatment also prevented phosphorylation of c-Raf-1 and Bcl-2 in response to Taxol treatment. The serine protease inhibitor did not alter JNK activity, but it enhanced Taxol-induced activation of ERK1/2 Treatment of cells with the inhibitor of MEK activation, PD98059, prevented Taxol-induced ERK acti vation both in the presence and absence of TPCK, but did not influence surv ival of either Taxol- or Taxol plus TPCK-treated cells. In addition, PD9805 9 had no effect on c-Raf-1 or Bcl-2 phosphorylation, Thus, while the Taxol- induced phosphorylations of c-Raf-1 and Bcl-2 proteins appear to be coupled , these events can be disassociated from ERK1/2 activation, In summary, the se findings suggest that phosphorylation of c-Raf-1 and Bcl-2, but not ERK1 /2, are important signaling events in Taxol-induced apoptosis of MCF-7 brea st cancer cells and that a TPCK inhibitable protease(s) is required for the se processes.