T-cell Lymphoma Cell Line Hut78 Research Articles

Effects of acitretin and interferon on the proliferative activity of and interleukin-15 expression in a human cutaneous T-cell lymphoma cell line Hut78

Objective To evaluate effects of acitretin and interferon-α (INF-α) alone or in combination on the proliferative activity of and interleukin-15 expression in human cutaneous T-cell lymphoma Hut78 cells. Methods Cultured Hut78 cells were divided into several groups, including blank control group, negative control group, dimethyl sulphoxide (DMSO) group and experimental groups. Cells in experimental groups were additionally classified into several subgroups to be treated with acitretin (0.1-10 μmol/L, acitretin groups) or INF-α (5 000-20 000 IU/ml, INF-α groups) alone, or the combination of 1.0 μmol/L acitretin and IFN-α at concentrations of 5 000-20 000 IU/ml (combination groups) , for 24, 48 and 72 hours. Subsequently, cell counting kit 8 (CCK8) assay was performed to assess the proliferative activity of Hut78 cells, and enzyme-linked immunosorbent assay (ELISA) to measure the expression of IL-15 in these cells. Results The proliferative activity of and IL-15 expression in Hut78 cells were both obviously suppressed in the acitretin groups and combination groups compared with the DMSO group, as well as in the INF-α groups compared with the negative control group, and the inhibitory effects gradually increased with the increase in acitretin or INF-α concentrations and treatment durations. As repeated measures analysis of variance revealed, there was a significant difference in both proliferation inhibition rates and IL-15 expression among different treatment durations and among different concentrations of acitretin or INF-α (all P < 0.05) , and there was an interaction effect between treatment durations and drug concentrations (all P < 0.05). A significant difference was observed in both proliferation inhibition rates and IL-15 expression at 24, 48 and 72 hours when the 1.0-μmol/L acitretin+ 10 000/20 000-IU/ml IFN-α group was compared with the 1.0-μmol/L acitretin group and 10 000/20 000 IU/ml IFN-α group (all P < 0.05). There was also a significant difference in IL-15 expression at 24, 48 and 72 hours between the 1.0-μmol/L acitretin+ 50 000-IU/ml IFN-α group and 5 000-IU/ml IFN-α group (all P < 0.05) . Conclusions Acitretin and IFN-α both can inhibit the proliferation of and IL-15 expression in Hut78 cells, the inhibitory effects are enhanced with the increase in drug concentrations and treatment durations, and the combination of acitretin and IFN-α appears to have stronger inhibitory effects than acitretin or IFN-α alone. Key words: Lymphoma, T-cell, cutaneous; Acitretin; Interferon-alpha; Interleukin-15; Cell proliferation; Cell line, tumor; Hut78 cells

Effects of chidamide combined with curcumin on human cutaneous T-cell lymphoma cell line Hut78 and their molecular mechanisms

Objective To evaluate the inhibitory effect of chidamide combined with curcumin on the proliferation of cutaneous T-cell lymphoma (CTCL) cell line Hut78, as well as their promotive effect on its apoptosis, and to explore their therapeutic mechanisms in CTCL. Methods Some Hut78 cells were treated with different concentrations of chidamide (0.3, 0.6, 1.2, 2.4 μmol/L) and 10 μmol/L curcumin alone or the combination of 1.2 μmol/L chidamide and 10 μmol/L curcumin for 24, 48 and 72 hours separately. MTS assay was conducted to estimate cell viability at each time point. After selection of chidamide concentrations, some Hut78 cells were treated with chidamide (0.6 and 1.2 μmol/L) and curcumin (10 μmol/L) alone or in combination (1.2 μmol/L chidamide and 10 μmol/L curcumin) for 24 hours, then, flow cytometry was performed to detect cell apoptosis and analyze cell cycle, real-time (RT) -PCR and Western-blot analysis were conducted to quantify the mRNA and protein expressions of apoptosis-associated genes Fas, caspase 8, nuclear factor (NF) -κB p65 as well as cell cycle-associated genes P21, CDK2 and cyclin E respectively. Statistical analysis was carried out by repeated-measures analysis of variance, one-way analysis of variance and the least significant difference (LSD) -t test. Results Chidamide could significantly inhibit the proliferation of Hut78 cells in a dose-dependent and time-dependent manner (F = 266.558, 564.966, respectively, both P 0.05) . Western-blot analysis showed that protein expressions of Fas, caspase 8 and P21 significantly increased, but those of NF-κB p65, CDK2 and cyclin E significantly decreased in the combined treatment group compared with the 0.6-, 1.2-μmol/L chidamide groups and blank control group receiving no treatment, which were in accordance with the above changes in mRNA expressions of these genes. Conclusion Chidamide can inhibit the growth of the CTCL cell line Hut78 by directly decelerating cell proliferation and inducing cell apoptosis, and the combibation with curcumin can markedly enhance the inhibitory effect of chidamide on the growth of Hut78 cells. Key words: Lymphoma, T-cell, cutaneous; Curcumin; Cell proliferation; Apoptosis; Chidamide; Hut-78 cell

Cytotoxic effects of bortezomib combined with fludarabine in T-cell lymphoma cell line HUT-78 in vitro

Cytotoxic effects of bortezomib combined with fludarabine in T-cell lymphoma cell line HUT-78 in vitro

Schedule-dependent inhibition of T-cell lymphoma cells by cotreatment with the mTOR inhibitor everolimus and anticancer drugs

Everolimus (RAD001) is a novel mammalian target of rapamycin (mTOR) inhibitor, and anti-proliferative activity in various malignancies has been reported. This study evaluated the anti-tumor effects and schedule-dependent synergism of everolimus in combination with other chemotherapeutic agents in T-cell lymphoma cell lines. Human T-cell lymphoma cell lines Hut-78 and Jurkat were treated with increasing doses of everolimus, alone or in combination with doxorubicin, etoposide, vincristine, or bortezomib, using different dosing schedules. Anti-tumor effects were measured by assays for cell proliferation, apoptosis, and cell cycle distribution. Drug interactions were determined by median effect analysis. Exposure to everolimus alone induced G1 phase cell cycle arrest without significant apoptosis. With certain dosing schedules, everolimus showed synergism with doxorubicin, etoposide, and bortezomib, but antagonism with vincristine. Cytotoxic synergism was observed following cotreatment with doxorubicin and everolimus, bortezomib and everolimus, doxorubicin followed by everolimus, and bortezomib followed by everolimus. By contrast, cell exposure to everolimus followed by doxorubicin or followed by bortezomib resulted in antagonistic effects. Sequential exposure to doxorubicin or bortezomib followed by everolimus effectively prevented potential negative interactions, and resulted in drug synergism. Drug combination synergisms or antagonisms were associated with variable effects on the cell cycle distribution. Everolimus effectively inhibited the growth of T-cell lymphoma cells in vitro. Specific schedule-dependent combinations of everolimus with other anti-tumor agents which avoid potential drug antagonism and produce effective synergism may lead to clinically effective treatments for T-cell lymphoma.

The Selective Protein Kinase C β Inhibitor Enzastaurin Induces Apoptosis in Cutaneous T-Cell Lymphoma Cell Lines through the AKT Pathway

Enzastaurin displays pro-apoptotic properties against a spectrum of malignancies and is currently being investigated in clinical trials. We have investigated the effects of enzastaurin on the viability of the cutaneous T-cell lymphoma cell lines HuT-78 and HH by using 3-(4,5-dimethylthiazol-2-yl)-5(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium assay, cell cycle analysis, propidium iodide and annexin-V staining, and caspase-3-mediated proteolytic activation. Enzastaurin-treatment decreased cell viability, increased annexin V-FITC-positive cells, and increased the proportion of sub-G1 populations in both cell lines that was not reversed by the T-cell growth stimulating cytokines IL-2, IL-7, IL-15. Enzastaurin-induced cell death involved caspase-3-activated cleavage of poly(ADP-ribose) polymerase that was inhibited by the pan-caspase inhibitor ZVAD-fmk, whereas the increase in sub-G1 population was only partially inhibited by ZVAD-fmk. Furthermore, enzastaurin downregulated AKT activity and its downstream effectors GSK3beta and ribosomal protein S6. The phosphatidylinositol 3-kinase (PI3K)/AKT pathway has been implicated in the growth and survival of hematologic malignancies and inhibition of this pathway is considered as a therapeutic target. Protein kinase C activation contributes to PI3K/AKT activation, but it is unknown how enzastaurin may interfere with signaling through this pathway. These results demonstrate that enzastaurin, at clinically achievable concentrations, induces apoptosis and affects AKT signaling, and provide a rationale for further in vivo studies addressing the therapeutic efficacy in cutaneous T-cell lymphoma patients.

Inhibition of phosphatidylinositol 3 ′-kinase induces preferentially killing of PTEN-null T leukemias through AKT pathway

We examined the functional role of the phosphatidylinositol 3 ′-kinase pathway in the growth and survival of cell lines of T-cell origin. Pharmacological inhibition of PI3 ′-kinase using LY294002 resulted in apoptosis of acute lymphoblastic T-cell leukemia (T-ALL) cell lines including CEM, Jurkat, and MOLT-4. On the other hand, the cutaneous T-cell lymphoma cell line HUT-78 was found to be refractory to LY294002- inducible apoptosis. Sensitivity or resistance to pharmacological inhibitors of PI3 ′-kinase correlated with tumor suppressor PTEN gene expression, as sensitive T-ALL cells do not express PTEN and have high level of activated AKT, in contrast to HUT-78 cells. Our data demonstrate that inhibition of PI3 ′-kinase results in dephosphorylation of AKT and partial inhibition of Bcl-xL expression in T-ALL cells, but not in HUT-78 cells. Interestingly, HUT-78 cells were also found to express higher levels of Bcl-xL protein as compared to T-ALL cells. Inhibition of PI3 ′-kinase also induces release of cytochrome c from mitochondria and activation of caspase-3 and PARP in all T-ALL cell lines tested, but not in HUT-78 cells. Taken altogether, our data demonstrate that the PI3 ′-kinase/AKT pathway plays a major role in the growth and survival of PTEN-null T-ALL cells, and identify this cascade as promising target for therapeutic intervention in acute T-cell leukemias.

T-cell lymphoma as a model for the use of histone deacetylase inhibitors in cancer therapy: impact of depsipeptide on molecular markers, therapeutic targets, and mechanisms of resistance

AbstractDepsipeptide (FK228) is a novel histone deacetylase inhibitor currently in clinical trials and the first to demonstrate clinical activity in patients. Responses have been observed in patients with T-cell lymphomas, despite prior treatment with multiple chemotherapeutic agents. To better understand the effects of histone deacetylase inhibitors on T-cell lymphoma, the human T-cell lymphoma cell line HUT78 was tested for sensitivity and molecular response to depsipeptide. Treatment with depsipeptide, as well as other histone deacetylase inhibitors, caused induction of histone acetylation, induction of p21 expression, and substantial apoptosis without significant cell cycle arrest. Treatment with the caspase inhibitor z-VAD-fmk significantly inhibited depsipeptide-induced apoptosis, enabling detection of cell cycle arrest. Treatment with depsipeptide increased expression of the interleukin-2 (IL-2) receptor, and combination with the IL-2 toxin conjugate denileukin diftitox resulted in more than additive toxicity. Cells selected for resistance to depsipeptide overexpressed the multidrug resistance pump, P-glycoprotein (Pgp). However, cells selected for resistance to depsipeptide in the presence of a Pgp inhibitor had a Pgp-independent mechanism of resistance. These studies confirm the activity of depsipeptide in a T-cell lymphoma model and suggest a general sensitivity of T-cell lymphoma to histone deacetylase inhibitors, an emerging new class of anticancer agents. (Blood. 2004;103:4636-4643)