cytochrome-c-t and Lymphoma--T-Cell

The p53 tumor suppressor protein has been implicated as an activator of apoptosis. In order to investigate the effect of chelerythrine and staurosporine on the activation of p53-dependent/-independent pathways of Dalton lymphoma (DL) cell death, cells were treated with chelerythrine and staurosporine for 1 h, 3 h and 6 h, respectively. It was found that treatment with chelerythrine and staurosporine increased the expression of total-p53/phospho-53 (ser-15) significantly at protein and mRNA levels, which resulted in activation of the p53-dependent apoptotic pathway in DL cells. In addition, increased activities of cyt-c, caspase-9 and caspase-3 and degradation of DNA into fragments confirmed activation of the p53-independent apoptotic pathway in p53 knockdown RNAi-DL cells. In brief, the present study demonstrated activation of p53-dependent/-independent apoptotic pathways in DL cells. Therefore, targeting of p53-dependent/-independent apoptotic pathways may lead to the possibility of designing and developing better therapeutic regimens to treat DL and other human cancers.

Topics: Animals; Antineoplastic Agents; Apoptosis; Benzophenanthridines; Blotting, Western; Caspase 3; Caspase 9; Cell Line, Tumor; Cytochromes c; Female; Gene Expression Regulation, Neoplastic; Humans; Lymphoma, T-Cell; Male; Mice, Inbred BALB C; Phosphorylation; Reverse Transcriptase Polymerase Chain Reaction; RNA Interference; Signal Transduction; Staurosporine; Tumor Suppressor Protein p53

Chelerythrine is a well-known protein kinase C inhibitor and potential antiproliferative and antitumor pharmacological agent. Chelerythrine inhibits/suppresses the HSF1 phosphorylation by inhibiting PKC and blocks the nuclear migration and subsequent synthesis of hsp70 leading to reduced cell viability and activation of apoptotic machinery. Chelerythrine is also known to enhance the production of reactive oxygen intermediate that is strong activator of apoptosis in high concentration. Therefore, the present study intended to investigate the role of chelerythrine-induced reactive oxygen intermediate on the viability and apoptosis of Dalton's lymphoma cells. Enhanced production of reactive oxygen species in Dalton's lymphoma (DL) cells was observed upon treatment of chelerythrine only which was seen completely abolished on treatment of mitochondrial complex inhibitors rotenone and malonate, and anti-oxidant, N-acetyl-L-cysteine. Increased number of DL cells undergoing apoptosis, as observed by fluorescent microscopy and flow cytometry analysis, in chelerythrine only-treated group was seen that was significantly inhibited on treatment of mitochondrial complex inhibitors and anti-oxidants. Staurosporine, on the other hand, does not lead to enhanced production of reactive oxygen intermediate in DL cells.

Topics: Animals; Antineoplastic Agents; Apoptosis; Benzophenanthridines; Caspases; Cell Line, Tumor; Cytochromes c; Disease Models, Animal; Humans; Hydrogen Peroxide; Lymphoma, T-Cell; Membrane Potential, Mitochondrial; Mice; Mitochondria; Models, Biological; Reactive Oxygen Species

The inhibitors of apoptosis (IAP) are important regulators of apoptosis. However, little is known about the capacity of Smac mimetics (IAP inhibitor) to overcome virally associated-lymphoma's (VAL) resistance to apoptosis. Here, we explored the pro-apoptotic effect of a novel Smac mimetic, RMT5265.2HCL (RMT) in VAL cells. RMT improved the sensitivity to apoptosis in EBV- and to some extend in HTLV-1- but not in HHV-8-VAL. Furthermore, we identified that RMT promotes caspase 3 and 9 cleavage by inhibiting XIAP and inducing the mitochondrial efflux of Smac and cytochrome C. This investigation further support exploring the use of Smac inhibitors in VAL.

Topics: Animals; Antineoplastic Agents; Apoptosis; Apoptosis Regulatory Proteins; Biomimetics; Cell Line, Tumor; Cell Transformation, Viral; Cytochromes c; Deltaretrovirus Infections; Dipeptides; Epstein-Barr Virus Infections; HEK293 Cells; Herpesvirus 4, Human; Human T-lymphotropic virus 1; Humans; Intracellular Signaling Peptides and Proteins; Lymphoma, T-Cell; Mice; Mice, Transgenic; Mitochondria; Mitochondrial Proteins; Models, Biological; Tetrazoles; Up-Regulation; X-Linked Inhibitor of Apoptosis Protein

Curcumin is known to exert its anticancer effect either by scavenging or by generating reactive oxygen species (ROS). In this study, we report that curcumin-mediated rapid generation of ROS induces apoptosis by modulating different cell survival and cell death pathways in HuT-78 cells. Curcumin induces the activation of caspase-8, -2, and -9, alteration of mitochondrial membrane potential, release of cytochrome c, and activation of caspase-3 and concomitant PARP cleavage, but the addition of caspase inhibitors only partially blocked the curcumin-mediated apoptosis. Curcumin also downregulates the expression of antiapoptotic proteins c-FLIP, Bcl-xL, cellular inhibitor of apoptosis protein, and X-linked IAP in a ROS-dependent manner. Curcumin disrupts the integrity of IKK and beclin-1 by degrading Hsp90. Degradation of IKK leads to the inhibition of constitutive NF-κB. Degradation of beclin-1 by curcumin leads to the accumulation of autophagy-specific marker, microtubule-associated protein-I light chain 3 (LC3), LC3-I. Our findings indicate that HuT-78 cells are vulnerable to oxidative stress induced by curcumin and as a result eventually undergo cell death.

Topics: Antineoplastic Agents; Apoptosis; Apoptosis Regulatory Proteins; Caspases; Cell Line, Tumor; Cell Survival; Curcumin; Cytochromes c; Down-Regulation; Enzyme Activation; HSP90 Heat-Shock Proteins; Humans; I-kappa B Kinase; Lymphoma, T-Cell; Molecular Targeted Therapy; NF-kappa B; Oxidative Stress; Protein Stability; Reactive Oxygen Species

Imiquimod is an immune response modifier currently used as a topical treatment of genital warts, basal cell carcinoma, cutaneous metastasis of malignant melanoma, and vascular tumors. We developed more efficient killers from the same family of compounds that can induce apoptosis without the prominent pro-inflammatory response associated with imiquimod. Among these new products, tk;4EAPB0203, a member of the imidazo[1,2-a]quinoxalines, exhibits an important cytotoxic activity in vitro. HTLV-I-associated adult T-cell leukemia (ATL) and HTLV-I-negative peripheral T-cell lymphomas are associated with poor prognosis. Using potentially achievable concentrations of EAPB0203, we demonstrate inhibition of cell proliferation, G2/M cell- cycle arrest, and induction of apoptosis in HTLV-I-transformed and HTLV-I-negative malignant T cells and fresh ATL cells, whereas normal resting or activated T lymphocytes were resistant. EAPB0203 treatment significantly down-regulated the antiapoptotic proteins c-IAP-1 and Bcl-XL and resulted in a significant loss of mitochondrial membrane potential, cytoplasmic release of cytochrome c, and caspase-dependent apoptosis. Moreover, in HTLV-I-transformed cells only, EAPB0203 treatment stabilized p21 and p53 proteins but had no effect on NF-kappaB activation. These results support a potential therapeutic role for EAPB0203 in ATL and HTLV-I-negative T-cell lymphomas, either as a systemic or topical therapy for skin lesions.

Topics: Aminoquinolines; Antineoplastic Agents; Apoptosis; bcl-X Protein; Caspases; Cell Division; Cytochromes c; G2 Phase; Human T-lymphotropic virus 1; Humans; Imiquimod; Inflammation; Inhibitor of Apoptosis Proteins; Jurkat Cells; Leukemia-Lymphoma, Adult T-Cell; Lymphocyte Activation; Lymphoma, T-Cell; Membrane Potential, Mitochondrial; NF-kappa B; Quinoxalines; Skin Neoplasms; Tumor Suppressor Protein p53

Vinorelbine is a chemotherapeutic vinca alkaloid clinically prescribed for non-small cell lung cancer and breast cancer. Here we studied the mechanism for vinorelbine-induced apoptosis in a human T-cell lymphoma. Although vinorelbine induces DNA fragmentation that is inhibited by specific peptide inhibitors for caspases-9 and -3 in Jurkat cells, caspase-8 deficiency retards vinorelbine-induced apoptosis. Activation of caspase-8 is also observed in vinorelbine-treated cells, and the activity is diminished when the caspase-3 activity is blocked by a specific peptide inhibitor, Ac-DNLC-CHO. Blocking of the Fas receptor with an antagonistic anti-Fas antibody does not affect vinorelbine-induced DNA fragmentation. These results suggest that vinorelbine-induced apoptosis is enhanced by the activation of caspase-8 via caspase-9-mediated activation of caspase-3, but not through a Fas-triggered signal. Western blotting suggests that vinorelbine cleaves caspase-3, -9 and -8 and reduces the amount of mitochondrial cytochrome c. Caspase-8 deficiency suppresses all of these events. A downstream substrate for caspase-8, Bid, is also cleaved in vinorelbine-treated cells, but the Bid truncation is also observed in caspase-8-deficient Jurkat cells. Importantly, recombinant caspases-3 and -9, as well as caspase-8, directly cleaves recombinant Bid in vitro. These results suggest that caspases-3 and -9 participate in Bid truncation, indicating a new mechanism for vinorelbine-induces apoptosis.

Topics: Apoptosis; BH3 Interacting Domain Death Agonist Protein; Caspase 3; Caspase 8; Caspase 9; Cell Line, Tumor; Cytochromes c; Humans; Jurkat Cells; Lymphoma, T-Cell; Vinblastine; Vinorelbine

We investigated the effect of UVA-activated 8-methoxypsoralen (PUVA) on the cell line Karpas 299 derived from anaplastic large-cell lymphoma (ALCL) expressing chimeric fusion protein nucleophosmin-anaplastic lymphoma kinase (NPM/ALK). NPM/ALK activates phosphatidylinositol 3 kinase (PI3K)/Akt pathway responsible for the cell protection from apoptosis. We found that PUVA treatment first induced G2/M cell cycle arrest resulting in a decrease in the cell proliferation rate. The mitochondrial apoptosis was triggered immediately following PUVA treatment, as we judged from the unmasking of mitochondrial membrane antigen 7A6. However, the mitochondrial membrane depolarization was not observed and caspase-3 was only slightly activated. The late apoptotic events were lacking: neither translocation of phosphatidylserine to the outer side of plasma membrane nor DNA fragmentation occurred. We revealed that PUVA enhanced the expression of peroxiredoxin, stress protein endoplasmin and galectin-3. Galectin-3 has been shown to protect mitochondrial membrane integrity and prevent cytochrome c release thereby blocking the effector stage of apoptosis. We suggest that the elevated level of this protein following PUVA treatment acts in synergy with the constitutively expressed chimeric kinase NPM/ALK to block the apoptosis.

Topics: Anaplastic Lymphoma Kinase; Caspase 3; Cell Cycle Proteins; Cell Division; Cell Line; Cytochromes c; G2 Phase; Galectin 3; Heat-Shock Proteins; Lymphoma, T-Cell; Membrane Glycoproteins; Membrane Proteins; Methoxsalen; Mitochondria; Peroxidases; Peroxiredoxins; Phosphatidylinositol 3-Kinases; Protein-Tyrosine Kinases; PUVA Therapy; Receptor Protein-Tyrosine Kinases; Ultraviolet Rays