A Novel mTOR Kinase Inhibitor Causes Growth Inhibition, Cell Cycle Arrest, Apoptosis and Autophagic Cell Death in Mantle Cell Lymphoma Cell Lines: A Distinct Profile from Rapamycin.

Mantle cell lymphoma (MCL) is a distinct sub-type non-Hodgkin lymphoma characterized by overexpression of cyclin D1 (CCND1) in 95% of patients due to the cytogenetic change of chromosome translocation t(11;14) (q13;q32). It remains one of the most challenging lymphomas associated with shorter response duration to conventional chemotherapy as well as continuous relapses and refractory to current drugs. However, dysregulation of cyclin D1 biology alone is insufficient to develop MCL. The emerging data suggest that the mammalian target of rapamycin (mTOR) plays a crucial role in the proper transmission of proliferative and anti-apoptotic signals through the PI3K/AKT pathway that makes it an attractive therapeutic target for hematological malignances including mantle cell lymphoma. As a single agent, rapamycin analogs such as temsirolimus (CCI-779) achieved 38% overall response rate in heavily pretreated MCL and prolonged progression free survival (PFS) in relapsed and refractory mantle cell lymphoma (4.8 months in temsirolimus vs. 1.9 months in investigator’s choice, ASCO 2008). mTOR regulates two distinct complexes TORC1 and TORC2. TORC1 complex is involved in cell cycle regulation by phosphorylating p70S6K and 4E-BP1, two molecules that are important for translational control of cyclin D1 and c-myc as well as ribosomal biogenesis whereas TORC2 complex mainly regulates phospho- AKT serine 473 leading to cell survival and proliferation. mTOR kinase also negatively regulates autophagy, a process of cellular bulk protein degradation by fusion to lysosomes upon the nutrient deprivation. We have developed mTOR kinase selective inhibitors which exhibit distinct biological profile from rapamycin in many cancer cell lines. Here we demonstrate that a selective mTOR kinase inhibitor displays potent anti-proliferative activity in JeKo-1 and Mino cells associated with decreased phosphorylation of S6, p70S6K, AKT S473, 4E-BP1 as well as decreased cyclin D1 levels leading to G1 arrest. The inhibitor also promotes autophagic cell death at 72h and 96h post-treatment. Furthermore a selective mTOR kinase inhibitor but not rapamycin induces a significant apoptosis in JeKo-1 and Mino cells. The observed apoptosis is correlated with caspases mediated PARP cleavage as well as inhibition of anti-apoptotic protein Mcl-1, suggesting TORC2/AKT S473 complex may provide survival signaling for mantle cell lymphoma. A timecourse study demonstrated that JeKo-1 and Mino cells undergo apoptosis at 24h and 48h followed by significant autophagic cell death at 72h and 96h in a dose dependent manner when exposed to our mTOR kinase inhibitor. In conclusion, mTOR kinase inhibitors are able to induce G1 cell cycle arrest, caspase-dependent apoptosis and autophagic cell death that contribute to the anti-tumor activity. Therefore it may provide a powerful alternative targeted therapy for mantle cell lymphoma.