Vinblastine and paclitaxel are both microtubule-interfering agents (MIA) that cause mitotic arrest and apoptosis. We have previously shown that inhibition of MEK/ERK signaling with PD98059 can acutely sensitize ML-1 leukemic cells to vinblastine with more than 70% of the cells undergoing apoptosis within 3 h (Cancer Res. 61:1533, 2001). This sensitization has been linked to the ability of vinblastine to activate JNK, and led to the conclusion that acute apoptosis can occur when JNK is activated and ERK is inhibited. Incubation with cycloheximide also sensitized ML-1 cells to vinblastine suggesting that rapid upregulation of proteins might protect against apoptosis. To better understand the mechanism underlying this sensitization, we performed a microarray experiment (Affymetrix, 22,000 genes) to determine the spectrum of genes induced by vinblastine that could potentially be protecting the cells. We observed that 271 genes were elevated >3 fold by a 4 h incubation with vinblastine. One gene of particular interest that was identified is Mcl-1, an anti-apoptotic member of the Bcl-2 family. When ML-1 cells were incubated with vinblastine for 4 h, Mcl-1 was elevated only at concentrations that also induced phosphorylation and activation of JNK. In contrast, neither Mcl-1 nor phospho-JNK was observed in cells incubated with paclitaxel, and no sensitization to vinblastine was observed. A time course experiment showed that phospho-JNK was elevated within 1 h of treatment while Mcl-1 was elevated within 2 h of treatment with vinblastine. The addition of the MEK inhibitor, PD98059, suppressed vinblastine-mediated induction of Mcl-1 protein and the cells underwent acute apoptosis. Addition of the JNK inhibitor, SP600125, also prevented vinblastine-mediated induction of Mcl-1 protein. However, the addition of SP600125 also protected the cells from acute apoptosis demonstrating that JNK also has a pro-apoptotic function in this model. The results show that Mcl-1 is rapidly induced by vinblastine in an ERK and JNK co-regulated manner. This may prevent cells from dying during G1, S or G2 phase of the cell cycle. When Mcl-1 expression is inhibited by PD98059, cells succumb to vinblastine at all phases of the cell cycle. Experiments are currently underway to ablate Mcl-1 expression and confirm its critical role in protecting cells from the acute apoptosis mediated by vinblastine.

[Proc Amer Assoc Cancer Res, Volume 46, 2005]