Acute sensitization of leukemia cells to vinblastine by small-molecule inhibitors of anti-apoptotic Bcl-2 family proteins

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Chemotherapeutic agents induce alterations in intracellular signal transduction cascades that culminate in the activation of BH3-only proteins and ultimately initiation of the apoptotic program. Here, the relationship between the expression of anti-apoptotic Bcl-2 family proteins and apoptosis in ML-1, NB4, and THP-1 leukemia cells treated with vinblastine (VB) was investigated. VB dissociates microtubules and usually induces M phase arrest prior to cell death. In ML-1 cells, the onset and extent of apoptotic cell death induced by VB (or other vinca alkaloids) can be dramatically enhanced by inhibiting the extracellular signal-regulated kinase (ERK) pathway with PD98059. This rapid induction of apoptosis is dependent on Jun N-terminal kinase (JNK) and occurs at all phases of the cell cycle. No sensitization was seen with paclitaxel which failed to activate JNK. Incubation with VB rapidly induces Mcl-1 in an ERK-dependent manner, while the levels of Bcl-2 remain unchanged; Bcl-X_L is minimally expressed. To confirm that Mcl-1 protects ML-1 cells from VB, we combined VB with GX15-070 (obatoclax), a small-molecule inhibitor of Mcl-1, Bcl-2, and Bcl-X_L. The cells were acutely sensitized to VB by the addition of GX15-070 with 100% of cells undergoing apoptosis within 6 hours. In NB4 cells, VB only slightly induces Mcl-1 but no induction is seen in THP-1 cells. Additionally, neither NB4 nor THP-1 cells were sensitized to VB by either PD98059 or GX15-070, the latter possibly because Bcl-X_L has been reported as a mechanism of resistance to GX15-070. Accordingly, we postulated that the NB4 and THP-1 cells might be using Bcl-2 or Bcl-X_L for protection from VB. We incubated the cell lines with ABT-737, a small molecule inhibitor of Bcl-2 and Bcl-X_L, but not Mcl-1. ABT-737 acutely sensitized the NB4 cells to VB, with almost 100% apoptosis within 4 hours. Neither ML-1 nor THP-1 cells were sensitized by ABT-737. The lack of sensitization in ML-1 cells is consistent with expression of Mcl-1 as a known mechanism of resistance to ABT-737, but the reason for the lack of response in THP-1 cells remains to be determined. The results demonstrate that the efficacy of vinca alkaloids may be dramatically increased when combined with an appropriate inhibitor of the particular Bcl-2 proteins that each tumor uses to protect itself. Furthermore, as these drug combinations convert the M phase-dependent activity of vinca alkaloids to a cell cycle phase-independent cell death, these combinations may enhance killing of quiescent cancer cells.