Inhibition of AKT/mTOR Signaling Induces Cell Cycle Arrest and Apoptosis in Mantle Cell Lymphoma.

Mantle cell lymphoma (MCL) is a distinct type of B-cell lymphoma associated with aggressive clinical course. The PI3K/AKT pathway is activated in a variety of human malignancies including hematopoeitic tumors. Activated (serine 473-phosphorylated) AKT (pAKT) is believed to mediate its effects, at least in part, via phosphorylation/activation of the mammalian target of rapamycin (mTOR). However, the biologic significance of AKT/mTOR signaling in MCL is unknown. We hypothesized that the PI3K/AKT signaling pathway is activated in MCL and contributes to tumor cell survival through activation of mTOR and its downstream effectors 4EBP1, p70S6K, ribosomal protein (rp) S6 (rpS6) and eIF-4E. We used 3 mantle cell lymphoma cell lines (Mino, Jeko and Z-138), a PI3K inhibitor (LY294002), and small interfering RNA (siRNA) specific for mTOR gene to investigate the biologic effects after inhibition of AKT/mTOR signaling in MCL. Treatment of MCL cells with LY294002 resulted in apoptotic cell death in a dose-dependent manner that was associated with downregultion of the anti-apoptotic proteins cFLIP and Mcl-1. Inhibition of PI3K also resulted in decreased cyclin D1 levels and cell cycle arrest as shown by decreased S-phase fraction assessed by BrdU incorporation. Changes in cell cycle and apoptosis were associated with decreased phosphorylation (activation) of mTOR and its downstream targets 4EBP1, p70S6K, and rpS6. Silencing of mTOR expression, using mTOR-specific siRNA also decreased phosphorylation of mTOR signaling proteins and induced cell cycle arrest and apoptosis in MCL cell lines. These biologic effects were associated with downregulation of cyclin D1 and modulation of apoptosis regulators. In addition, expression of activated AKT, and effectors downstream of AKT/mTOR pathway (p-p70S6K, p-rpS6, 4EBP1, eIF-4E) was assessed in MCL tumors. Using a 10% cutoff to define positivity, 10/20 (33%) expressed ^Ser473pAKT, 22/22 (100%) p-p70S6K, 5/20 (25%)p- rpS6, 13/14 (93%) 4E-BP1, and 16/29 (55%) eIF-4E. p-p70S6K phosphoprotein was detected predominantly in the nucleus of tumor cells. All other phosphoproteins were expressed with a cytoplasmic pattern. We conclude that activation of PI3K/AKT may contribute to cell cycle progression and tumor cell survival in MCL cells through activation of the mTOR signaling pathway. Our data suggest that inhibition of mTOR with rapamycin analogues currently used in clinical trials may be a novel therapeutic strategy for patients with MCL.