Targeting the apoptotic machinery of pediatric acute lymphoblastic leukemia in vivo using the BH3-mimetic ABT-737

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Despite improvements in the treatment of pediatric acute lymphoblastic leukemia (ALL), new therapies are required for patients who relapse. Over-expression of anti-apoptotic Bcl-2 family proteins may contribute to poor outcome. The efficacy of ABT-737, a small molecule BH-3 mimetic that inhibits anti-apoptotic Bcl-2 family members, was evaluated in vivo and in vitro, as a single agent and in combination with conventional drugs, against a panel of 10 pediatric ALL biopsies established as xenografts in immune-deficient mice. ABT-737 significantly delayed the in vivo progression of 7/10 xenografts when tested at only 25% of its maximum tolerated dose (MTD, 25 mg/kg Mon-Fri x 4 weeks). In vitro, xenograft cells were exquisitely sensitive to ABT-737 (IC50 <50 nM by MTT assay), and exposure to 100 nM ABT-737 caused loss of mitochondrial transmembrane potential, caspase-3/7 activation and loss of viability in the majority of cells within 16 h. Using fixed-ratio combination cytotoxicity assays, ABT-737 exhibited broad and strong synergy (Combination Indices, CIs, <0.3) in vitro with the established drugs vincristine, etoposide, topotecan, and L-asparaginase against a chemoresistant xenograft, ALL-19, derived from a child who died within 11 months of diagnosis. Surprisingly, ABT-737 was only synergistic with the glucocorticoid, dexamethasone, against dexamethasone-sensitive (IC5010 µM, n=5) xenografts, indicating that ABT-737 is unlikely to reverse glucocorticoid-resistant ALL in vivo. Although ABT-737 caused greater than additive efficacy in vivo against ALL-19 when combined with vincristine and etoposide, it exerted its best effects when combined with L-asparaginase or topotecan: while L-asparaginase at its MTD (2,000 IU/kg Mon-Fri x 4 weeks) was ineffective against ALL-19, when a lower dose (1,500 IU/kg) was combined with ABT-737 (25 mg/kg) leukemia progression was delayed 26.7 days more than if the effects were additive; moreover, the combination of topotecan (1 mg/kg Mon-Fri x 4 weeks) and ABT-737 delayed the progression of ALL-19 21.2 days greater than expected. In vitro, L-asparaginase down-regulated Mcl-1 within 1 h, while topotecan rapidly up-regulated p53 in ALL-19. Therefore, we reasoned that combining L-asparaginase, topotecan and ABT-737 would target the apoptotic machinery by non-overlapping mechanisms. Consistent with this hypothesis, the 3 drugs exerted strong synergy (CI<0.3) in vitro against ALL-19. Moreover, in vivo they delayed the progression of ALL-19 by 52 days greater than expected if the drugs were additive. This study has shown that the novel BH-3 mimetic ABT-737 augments the in vivo effects of drugs commonly used to treat ALL, and that rational targeting of the apoptotic machinery can induce durable remissions in chemoresistant pediatric ALL xenografts derived from patients with aggressive and fatal disease.