Abstract
BackgroundVenetoclax, a small molecule BH3 mimetic which inhibits the anti-apoptotic protein Bcl-2, and idasanutlin, a selective MDM2 antagonist, have both shown activity as single-agent treatments in pre-clinical and clinical studies in acute myeloid leukemia (AML). In this study, we deliver the rationale and molecular basis for the combination of idasanutlin and venetoclax for treatment of p53 wild-type AML.MethodsThe effect of idasanutlin and venetoclax combination on cell viability, apoptosis, and cell cycle progression was investigated in vitro using established AML cell lines. In vivo efficacy was demonstrated in subcutaneous and orthotopic xenograft models generated in female nude or non-obese diabetic/severe combined immunodeficiency (NOD/SCID) mice. Mode-of-action analyses were performed by means of cell cycle kinetic studies, RNA sequencing as well as western blotting experiments.ResultsCombination treatment with venetoclax and idasanutlin results in synergistic anti-tumor activity compared with the respective single-agent treatments in vitro, in p53 wild-type AML cell lines, and leads to strongly superior efficacy in vivo, in subcutaneous and orthotopic AML models. The inhibitory effects of idasanutlin were cell-cycle dependent, with cells arresting in G1 in consecutive cycles and the induction of apoptosis only evident after cells had gone through at least two cell cycles. Combination treatment with venetoclax removed this dependency, resulting in an acceleration of cell death kinetics. As expected, gene expression studies using RNA sequencing showed significant alterations to pathways associated with p53 signaling and cell cycle arrest (CCND1 pathway) in response to idasanutlin treatment. Only few gene expression changes were observed for venetoclax treatment and combination treatment, indicating that their effects are mediated mainly at the post-transcriptional level. Protein expression studies demonstrated that inhibition of the anti-apoptotic protein Mcl-1 contributed to the activity of venetoclax and idasanutlin, with earlier inhibition of Mcl-1 in response to combination treatment contributing to the superior combined activity. The role of Mcl-1 was confirmed by small hairpin RNA gene knockdown studies.ConclusionsOur findings provide functional and molecular insight on the superior anti-tumor activity of combined idasanutlin and venetoclax treatment in AML and support its further exploration in clinical studies.Electronic supplementary materialThe online version of this article (doi:10.1186/s13045-016-0280-3) contains supplementary material, which is available to authorized users.
Highlights
Venetoclax, a small molecule BCL-2 homolog domain 3 (BH3) mimetic which inhibits the anti-apoptotic protein Bcl-2, and idasanutlin, a selective mouse double minute 2 homolog (MDM2) antagonist, have both shown activity as single-agent treatments in pre-clinical and clinical studies in acute myeloid leukemia (AML)
MV4-11 and MOLM13 cells were cultured in RPMI-1640 supplemented with 20 % fetal calf serum (FCS; Gibco), HL-60 cells were cultured in RPMI-1640 supplemented with 10 % FCS, and OCI-AML-3 cells were cultured in alpha-MEM supplemented with 20 % FCS
Venetoclax has enhanced specificity compared with earlier BH3 mimetics, which interact with the BH3 domains of Bcl-2, Bcl-xL, and Bcl-w
Summary
Venetoclax, a small molecule BH3 mimetic which inhibits the anti-apoptotic protein Bcl-2, and idasanutlin, a selective MDM2 antagonist, have both shown activity as single-agent treatments in pre-clinical and clinical studies in acute myeloid leukemia (AML). Acute myeloid leukemia (AML) is the most common acute leukemia in adults and, high remission rates are achieved following induction chemotherapy, the majority of patients eventually relapses and becomes resistant to treatment [1]. Venetoclax (GDC-0199/ABT-199) is an orally bioavailable novel small-molecule BCL-2 homolog domain 3 (BH3) mimetic, designed to selectively inhibit the anti-apoptotic protein Bcl-2. Pre-clinical studies show that venetoclax has significant anti-tumor activity in AML models and that venetoclax-dependent inhibition of Bcl-2 can prime AML cells for responsiveness to chemotherapy [17]
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