ABSTRACT
In many acute leukemias, normal differentiation does not occur. However, in many cell lines derived from hematologic malignancies, differentiation or apoptosis can be induced by variety of agents. Despite advances in the treatment of Acute Lymphoblastic Leukemia [ALL], in most patients long-term survival rates remain unsatisfactory, especially in T-cell derived ALL. Thus we studied the anti-cancer effects of fenretinide, 1alpha,25[OH]2D3, and bryostatin-1 in CCRF-CEM [T-cell derived] and Nalm-6 [B-cell derived] ALL cell lines. Using MTT assays, both cell lines were shown to exhibit increased inhibition of proliferation at micro [fenretinide] and nanomolar [1alpha,25[OH]2D3, bryostatin-1] concentrations. These anti-cancer agents were shown to induce apoptosis and activate caspase-3 pathway in both ALL cell lines. Furthermore, for the first time we are reporting consistent anti-proliferative and apoptotic effects of Bryostatin-1 in ALL T-cell derived cell line with the lowest ED50 [ranging 4.6 nM - 7.4 nM]. To evaluate the differentiation induction by fenretinide, 1alpha,25[OH]2D3, and bryostatin-1 in ALL cell lines, we assayed for the expressions of CD19, CD38 markers on Nalm-6 and CD7 marker on CCRF-CEM cell line. The flow cytometric analysis showed a significant increase in expression of CD markers in response to anticancer drug treatments. To assay the effects of anti-cancer drugs on cell cycle distribution, cell cycle analysis using flow cytometry was employed. These anti-cancer drugs appear to affect the CCRF-CEM and Nalm-6 cell cycles differently [G0/G1 and G2/M, respectively]. Overall results demonstrate that the anticancer agents used in this study are strong inhibitors of ALL cell proliferation and inducers of apoptosis and differentiation in vitro. These findings may be quite helpful if these drugs are to be used for differentiation therapy of ALL patients in clinics in the future. Further studies are warranted to establish the in vivo effect of these drugs particularly in patients with T-cell derived ALL
ABSTRACT
In many acute leukemias, normal differentiation does not occur. However, in many cell lines derived from hematologic malignancies, differentiation or programmed cell death [apoptosis] can be induced by variety of agents including: Vitamin analogs, demethylating agents, cyclic AMP analogs and anti-proliferative agents. To the best of our knowledge there had been not any study specifically to analyze apoptotic and anti-proliferative effects of 4-HPR [a vitamin analog] in NB-4 cell line. To test whether this drug has activity in acute myeloid leukemia [AML], we first analyzed the anti-proliferative effect of 4-HPR in on EAML cell line [NB-4] using MTT Assay. Next we tested whether this drug induced apoptotic cell death. The abilty of this compound to induce apoptosis of cancer cells was examined by Annexin V-FITC Assay using Flow cytometry. We also analyzed the cell cycle progression by PI staining using flow cytometry. Using MTT assay, NB-4 cells exhibited increased inhibition of proliferation at micromolar concentrations of 4-HPR at 24, 48 and 71 hrs post treatment. Flow cytometry analysis indicates that 4-HPR is a potent inducer of in vitro apoptotic cell death, and cell cycle analysis revealed an increase in 5 phase population. In total, the results indicate that 4-HPR is a strong inhibitor of AML cell proliferation and a potent inducer of in vitro apoptotic cell death. Further studies are required to evaluate the in vitro effects of 4-HDR in AML blasts derived from AML patients