Synergistic Induction of Apoptosis by Simultaneous Disruption of the Bcl-2 and mTOR/Akt Pathways in Acute Myeloid Leukemia.

Activation of the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt)/mammalian target of rapamycin (mTOR) signaling pathway provides survival signals for leukemic cells and blockade of this pathway may facilitate cell death. Akt/mTOR activation is a frequent event in AML that translates into significantly shorter survival of AML patients (S.M. Kornblau, ASH 2007). Bcl-2 family proteins are key regulators of apoptosis that are known to promote tumorigenesis and chemoresistance. We have recently reported that ABT-737, a small molecule BH3 mimetic, effectively kills AML progenitor cells via the disruption of Bcl-2:Bax, but its activity is largely diminished in cells overexpressing Mcl-1 (M. Konopleva, Cancer Cell 2006). We have now investigated anti-leukemia efficacy of concomitant blockade of mTOR/Akt, known to regulate Mcl-1 levels, and of Bcl-2 signaling. Combined use of rapamycin derivative temsirolimus and ABT-737 induced striking apoptotic responses in AML cell lines OCI-AML3 and MOLM13 leukemic cells. Mechanistically, temsirolimus downregulated expression of the anti-apoptotic protein Mcl-1 and induced expression levels of the proapoptotic BH3-only protein Noxa which specifically neutralizes Mcl-1. These effects were seen upon prolonged temsirolimus treatment that inhibited mTORC2 complex (Zeng, Blood 2007) and downregulated Akt phosphorylation. In primary AML samples co-cultured with bone marrow-derived stromal cells, ABT-737 at 25nM induced apoptotic cell death in CD34⁺ AML progenitor cells from 13 of the 16 samples tested. Temsirolimus/ABT-737 combination markedly enhanced apoptosis induction in 8/9 primary samples that were sensitive to temsirolimus alone (specific apoptosis, temsirolimus 15±5%; ABT-737, 28±5.9%; temsirolimus+ABT, 45±5.9%, p<0.001) and in 3/7 samples resistant to temsirolimus (p=0.7). Further, combined blockade of mTOR/AKT and Bcl-2 pathway effectively induced apoptosis in 3 AML samples resistant to ABT-737 alone. Taken together, these results indicate that temsirolimus can directly alter the Bcl-2 apoptotic rheostat via induction of Noxa and decreased Mcl-1 expression sensitizing leukemia cells to apoptosis induced by BH3 mimetics. The combined use of agents interfering with AKT/mTOR and Bcl-2 pathways may be a highly effective anti-leukemia strategy that induces apoptosis in AML progenitor cells in the context of bone marrow microenvironment.