Inhibitors of mToR Mediate Anti-Myeloma Activity by Apoptosis through the Extrinsic Pathway.

The mammalian target of rapamycin (mToR) plays a crucial role in cell growth due to its role as nutrient dependent regulator of important cytokine signalling pathways. In multiple myeloma (MM) mToR is involved in the AKT pathway which can be activated by the loss of the tumor suppressor PTEN or stimulation with growth and survival factors such as interleukin-6 (IL-6) and insulin-like growth factor-1 (IGF-1). mToR can be blocked by rapamycin or everolimus. When various human plasmacytoma lines were tested, rapamycin and everolimus induced a dose-dependent growth inhibition. The addition of bone marrow stromal cells, IL-6 or IGF-1 was unable to prevent the inhibitory activity. Growth inhibition was mediated not only by G1 cell cycle arrest but also by an induction of apoptosis as measured by propidium iodide staining. Further analysis was performed to elucidate the mechanism of apoptosis in the particular sensitive plasma cell line INA-6. mToR inhibition induced exclusively cleavage of pro-caspase 8 (extrinsic pathway) but not of pro-caspase 9 (intrinsic pathway). To confirm this observation, INA-6 cells were transfected with BCL-XL, an inhibitor of the intrinsic pathway, and treated with rapamycin. The transfectants showed similar sensitivity to rapamycin as the paternal cell line. Rapamycin was also tested in the INA-6 SCID mouse xenograft model.

Rapamycin was given orally for two weeks starting 24 h after tumor inoculation. A significant survival benefit of the rapamycin treated group compared to control animals (p= 0.0004) was observed. Six out of 14 treated mice did not develop plasmacytomas during the observation period of 149 days. In addition, even short term treatment of plasmacytoma bearing mice led to a significant shrinkage of the tumor mass. Histological examination revealed an increase of apoptotic plasma cells according to morphology after 72 h of treatment compared to controls. Induction of apoptosis in tumor cells was confirmed by immunohistological staining using an antibody specific for the human cleaved form of poly (ADP-ribose) polymerase. Since drugs inhibiting mToR are inducing apoptosis in human malignant plasma cells not only in vitro but also in vivo, the inhibition of mToR may represent an attractive new concept for multiple myeloma therapy.