| ObjectiveMantle cell lymphoma (MCL) has poor outcome and is a therapeutic challenge. Atiprimod,a cationic amphiphilic compound, inhibits the proliferation of most human tumor celllines. Our study was undertaken to evaluate the therapeuticefficacy of Atiprimod on MCLcells and elucidate the mechanism by which it induces cell apoptosis.MethodsFour human MCL cell lines, SP53, MINO, Grant 519, and Jeko-1; freshly isolatedprimary MCL cells from three MCL patients; and normal peripheral blood mononuclearcells (PBMCs) from five healthy donors were treated with Atiprimod.ResultsThe results showed that Atiprimod not only inhibited the growth of the MCL cell linesSP53, MINO, Grant 519, and Jeko-1, but also freshly isolated patient MCL cells in adose-dependent manner. Interestingly, Atiprimod also inhibited the proliferation of PHA-,PMA/ionomycin-, or anti-CD3 mAb/anti-CD28 mAb-activated, but not resting PBMCsfrom healthy donors. Atiprimod induced apoptosis in both the cell lines and primary MCLcells in time-dependent and dose-dependent manners. Notably, Atiprimod, at the sametreatment condition, was in much lower cytotoxicity on normal T cells. Atiprimod wasalso effective and therapeutic in a MCL mouse model established in severe combinedimmunodeficient (SCID) mice. SP53 and Grant 519 cells were inoculated subcutaneouslyinto the SCID mice in the number of 5×10~6 cells for each mouse, respectively. Afterpalpable tumors developed, mice were treated intraperitoneally with either vehicle alone (PBS, n=10) or Atiprimod (25 mg/kg per day, n=10) for 6 consecutive days. Tumorgrowth was significantly inhibited after Atiprimod treatment compared with vehiclecontrol (P＜0.05), and the survival time of tumor-bearing mice was significantlyprolonged in the treatment group. Western blot analysis showed that Atiprimod activatedc-Jun NH(2)-terminal protein kinases (JNK), inhibits signal transducer activator oftranscription 3 (STAT3) and IκB. Furthermore, it was found that the apoptosis of MCLcells was induced through the mitochondrial pathway involved in both the release ofcaspase-independent apoptosis-inducing factor (AIF) and the activation ofcaspase-dependent caspase-9, caspase-3, and the cleavage of PARP. The level of Bax, Bad,and phospho-Bcl-2 were up-regulated in Atiprimod-treated MCL cells. Western blotshowed that pan-caspase inhibitor (z-VAD-FMK) completely blocked cleavage of PARP.Tunel assay showed that pretreatment of cells with AIF inhibitor N-Penylmaleimide butnot Z-VAD-FMK completely prevented nucleosomal DNA fragmentation.ConclusionAtiprimod inhibits growth and induces apoptosis of MCL cells in vitro and in vivo. Cellapoptosis was induced via activation of both caspase-independent and caspase-dependentsignaling pathway. These results support the use of Atiprimod as a potential agent in MCLchemotherapy. ObjectiveEvaluated whether bortezomib (BTZ) is additive or synergistic with cyclophosphamide(CTX) and rituximab (RTX) in treatment relapsed Mantle cell lymphoma (MCL).MethodsFour human MCL cell lines SP53, MINO, Grant 519, and Jeko-1 and freshly isolatedprimary tumor cells from three MCL patients were treated with BTZ, CTX, RTXindividually or in combination of all three (BRC regimen). Cell proliferation andapoptosis were evaluated to determine if there was additive or synergistic effect of theBRC regimen. Western blot analysis was used to elucidate the molecular mechanism bywhich BTZ, RTX, CTX and BRC induces apoptosis in MCL cells. In addition, in vivoexperiments using severe combined immunodeficiency mice with human mantle celllymphoma xenografts were performed to examine the in vivo efficacy of the regimen tocontrol the growth of and eradicate MCL cells.ResultsBTZ and CTX as single agents inhibited the growth of MCL cell lines in adose-dependent manner (P＜0.01). The IC50 (inhibitory concentration at 50％) for BTZand for CTX were between 10 and 20 nM and between 5 and 20 mM, respectively.Increasing doses of BTZ with a fixed dose of RTX (10μg/mL) and CTX (10 mM)resulted in markedly synergistic growth inhibition of MCL cells (P＜0.01). The BRCregimen induced apoptosis in about 69.7％of MCL cell lines and 92.6％of primary tumorcells (P＜0.05 and P＜0.01, compared with those induced by BTZ, RTX or CTX assingle agents). Furthermore, western blotting analysis showed that BRC inducedapoptosis earlier via activation and cleavage of caspases-8, -9, and -3, and PARP ascompared with single-agent treatment. The pan-caspase inhibitor z-VAD-FMK completelyblocked apoptosis induced by BRC. In vivo studies demonstrated that BRC regimen eradicated subcutaneous tumors in MCL-bearing SCID mice and significantly prolongedthe long-term event-free survival up to 10 weeks in 70％of the mice, whereas alltumor-bearing mice receiving single-agent treatment or PBS (control) died of aggressiveMCL within 5 weeks.ConclusionCytoreductive chemotherapy with both BTZ and anti-CD20 antibody effectively inhibitedthe growth and induced apoptosis of MCL cells in vitro and in vivo. BRC regimen mayoffer a better therapeutic modality for MCL patients. Thus, our data lay the basis for aclinical trial in relapsed MCL using the BRC combinational treatment. |
