MTOR (Mammalian Target of Rapamycin) Inhibition with RAD001 Induces Cell Cycle Arrest in Diffuse Large B Cell Lymphoma (DLBCL) Cells and Sensitizes DLBCL Cells to Rituximab Therapy.

Abstract

Purpose: DLBCL is a common lymphoma entity. Although a significant amount of patients can be cured with modern chemotherapeutic regimen, a substantial proportion of patients die due to progressive disease. Therefore, new therapeutic strategies are clearly needed. Inhibitors of MTOR (mammalian target of rapamycin) represent a new class of antiproliferative drugs with applications as immunosuppressive and anticancer agents. Extensive safety data exist on RAD001 which is already approved as an immunosuppressant in organ transplant recipients.Methods: We have analyzed the effects of MTOR inhibition in three DLBCL cell lines: SUDHL-4, DB, DOOH-2. Propidium Iodide staining, MTT- and Annexin-V assays were performed to test the effects on viability, cell cycle distribution and apoptosis. Expression of cell cycle regulatory and phosphorylation of p70 s6 kinase and 4-EBP-1 molecules was revealed in western blot experimentsResults: Rapamycin and RAD001 inhibited cell cycle progression in DLBCL cells by inducing a G1 arrest without inducing apoptosis. Phosphorylation of the main targets of MTOR, p70s6 kinase and 4-EBP-1 was reduced in cells cultured in the presence of RAD001.Cell cycle arrest was accompanied by reduced phosphorylation of the Retinoblastoma protein (RB) as well as reduced expression of cyclin D3, E and A in all cell lines. Although the effect of chemotherapeutic agents like Vincristin or Doxorubicin was not enhance by Rad001, Rituximab induced cytotoxicity was augemted in both Rituximab sensitive cell lines.Conclusion: MTOR Inhibition is a promising therapeutic strategy in DLBCL of the germinal center like subtype by inducing a G1 arrest and augments Rituximab induced cytotoxicity. Therefore, combination of these drugs might be an interesting new therapeutic approach in DLBCL patients.