Bcl-2, Mcl-1 and p53 Expression Confer Sensitivity to Bcl-2 Inhibitor ABT-737 in Multiple Myeloma.

Bcl-2 family of proteins confers resistance to chemotherapy in multiple myeloma (MM). Identification of BH3-mimetic drugs that inactivate pro-survival targets by interfering with the interaction of Bcl-2 proteins may therefore be therapeutically useful. A recent study identify ABT-737, a potent small-molecule inhibitor of anti-apoptotic proteins Bcl-2, Bcl-x L , and Bcl-w with an affinity 2–3 orders of magnitude more potent than any previously reported compounds (Nature2005, 435:672–681). Here we show that ABT-737 triggers apoptosis in MM cells resistant to conventional therapies. The functional specificity of ABT-737 binding to Bcl-2 or Bcl-x L was confirmed by using the less active enantiomer of ABT-737, which had no significant effect on viability of cells, even at higher concentration. Washout experiments show that exposure of MM cells to ABT-737 for 3h is sufficient to cause an early and irreversible commitment to cell death. Given that ABT-737 binds Bcl-2, Bcl-x L , and Bcl-w with high affinity but has far lower affinity for Mcl-1, we asked whether the anti-MM activity of ABT-737 is regulated by the relative expression of Bcl-2, Bcl-x L , or Mcl-1. For these studies, we selected MM.1S (sensitive to ABT-737: IC 50 -2 μM) and OPM-1 (least sensitive to ABT-737; IC 50 : 10–15 μM) and examined the basal expression levels of Bcl-2, Bcl-x L , and Mcl-1. MM.1S cells, in contrast to OPM-1 cells, express high basal levels of both Bcl-2 and Bcl-x L , but low Mcl-1 protein; conversely, OPM-1 cells express high Mcl-1 and low Bcl-2 levels. Since ABT-737 cannot neutralize Mcl-1, it is likely that the high Mcl-1 and low Bcl-2/Bcl-x L expression profile in OPM-1 renders these cells less sensitive to ABT-737; whereas low Mcl-1 and high Bcl-2/Bcl-x L expression in MM.1S cells allows for efficient killing by ABT-737. Importantly, treatment of MM.1S MM cells with ABT-737 and proteasome inhibitor Bortezomib induces additive cytotoxicity (combination index = 1.0). The mechanism underlying additive anti-MM activity of ABT-737 with Bortezomib includes downregulation of Mcl-1 along with targeting Bcl-2. Our findings have clinical implications: Bortezomib is FDA approved for the treatment of MM, but prolonged treatment can be associated with toxicity (Chauhan et al. Cancer Cell2005, 8:407–419); importantly, combining ABT-737 with Bortezomib would allow for the use of lower doses of Bortezomib. Another mechanism whereby MM cells evades the cytotoxic effects of chemotherapy is via p53 mutations. Bcl-2 is linked to p53-mediated signaling, and we therefore examined whether ABT-737 is able to overcome the tumorigenic effects conferred via p53 mutations. Co-precipitations experiments show that MM.1S cells (ABT-737-sensitive) predominantly carry wild type p53, whereas OPM-1 cell line (less sensitive to ABT-737) has mutant p53. Taken together, these finding suggest that 1) sensitivity to ABT-737 correlates with Bcl-2 expression in MM cells, 2) higher expression of Mcl-1 reduces sensitivity to ABT-737 whereas other anti-MM agents that block Mcl-1 may synergize with ABT-737, and 3) ABT-737 induces apoptosis in MM with p53 (wt) or p53 (mt), albeit with differential sensitivity. A report that ABT-737 enhances the apoptotic activity of chemotherapeutic agents (Olterdorf et al. Nature2005, 435:672–681), together with our present findings, provides the framework for clinical trials of ABT-737, either alone or in combination with other anti-MM agents, to enhance efficacy, reduce toxicity, and overcome drug resistance in MM patients.