No abstract

Curcumin, the principal curcuminoid of tumeric, has potent anticancer activity. To determine the mechanism of curcumin-induced cytotoxicity in prostate cancer cells, we exposed PC3 prostate carcinoma cells to 25 to 100 microM curcumin for 24 to 72 h. Curcumin treatment of PC3 cells caused time- and dose-dependent induction of apoptosis and depletion of cellular reduced glutathione (GSH). Exogenous GSH and its precursor N-acetyl-cysteine, but not ascorbic acid (AA) or ebselen, decreased curcumin accumulation in PC3 cells and also prevented curcumin-induced DNA fragmentation. The failure of AA and ebselen to protect PC3 cells from curcumin-induced apoptosis argued against the involvement of reactive oxygen species; rather, GSH-mediated inhibition of curcumin-induced cytotoxicity was due to reduced curcumin accumulation in PC3 cells. Curcumin-treated PC3 cells showed apoptosis-inducing cellular ceramide accumulation and activation of p38 mitogen-activated protein kinase (MAPK) and c-jun N-terminal kinase (JNK). Caspase-3, caspase-8, and caspase-9 were activated, and cytochrome c and apoptosis-inducing factor (AIF) were released from mitochondria following curcumin treatment. Interestingly, curcumin-induced apoptosis was not prevented by p38 MAPK, JNK, or caspase inhibition. We conclude that curcumin-induced cytotoxicity was due to cellular ceramide accumulation and damage to mitochondria that resulted in apoptosis mediated by AIF and other caspase-independent processes.

Thymoquinone (TQ) from black caraway seeds has several anticancer activities; however, its effect on triple-negative breast cancer (TNBC) cells that lack functional tumor suppressor p53 is not known. Here, we explored the growth inhibitory effect of TQ on 2 TNBC cell lines with mutant p53. Cell metabolism assays showed that TQ inhibited TNBC cell growth without affecting normal cell growth. Flow cytometric analyses of TQ-treated TNBC cells showed G1 phase cell cycle arrest and apoptosis characterized by the loss of mitochondrial membrane integrity. Western blots of lysates from TQ-treated TNBC cells showed cytochrome c and apoptosis-inducing factor in the cytoplasm, as well as caspase-9 activation consistent with the mitochondrial pathway of apoptosis. Caspase-8 was also activated in TQ-treated TNBC cells, although the mechanism of activation is not clear at this time. Importantly, TQ-induced apoptosis was only partially inhibited by zVAD-fmk, indicating a role for caspase-independent effector molecules. Poly(ADP-ribose) polymerase cleavage and increased γH2AX, as well as reduced Akt phosphorylation and decreased expression of X-linked inhibitor of apoptosis, were evident in TQ-treated cells. Finally, TQ enhanced cisplatin- and docetaxel-induced cytotoxicity. These findings suggest that TQ could be useful in the management of TNBC, even when functional p53 is absent.

No abstract

Thymoquinone, which is the active constituent of the volatile oil extracted from black caraway seeds (Nigella sativa), has been reported to have antiinflammatory, antioxidant, and antineoplastic activities in vitro and in vivo. In this study we explored the mechanism by which thymoquinone inhibits breast cancer cell growth, as well as the potential of using thymoquinone in combination with conventional chemotherapeutic drugs or radiation therapy. Cytofluorimetric analysis of Oregon Green 488-stained MDA-MB-231 and MDA-MB-468 breast cancer cells following thymoquinone treatment demonstrated inhibition of cell proliferation. Colorimetric 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays performed on MDA-MB-231, MDA-MB-468, and T-47D breast cancer cells after 24, 48, and 72 h treatment with thymoquinone (0.5–10 µM) showed a time- and dose-dependent cytotoxic effect, while untransformed human mammary epithelial cells were not adversely affected by thymoquinone. Longer exposure (144 h) to lower doses of thymoquinone (0.25–1 µM) was also cytotoxic for breast carcinoma cells. Importantly, staining of thymoquinone-treated breast cancer cells with annexin-V-FLUOS/propidium iodide indicated that cell death was by apoptosis. Western blot analysis of thymoquinone-treated breast cancer cells revealed increased cytosolic cytochrome c and PARP cleavage, which suggested induction of the mitochondrial pathway of apoptosis. However, broad spectrum caspase inhibitors (Bod-D and z-VAD-fmk) did not prevent thymoquinone-induced cytotoxicity, implying caspase-independent cell death. Thymoquinone treatment also sensitized breast cancer cells to γ-radiation and the conventional chemotherapeutic agents docetaxel and cisplatin. We conclude that thymoquinone caused caspase-independent apoptosis in breast cancer cells that involved the loss of mitochondrial membrane integrity. Furthermore, the ability of thymoquinone to potentiate the cytotoxic effects of chemotherapeutic drugs and -radiation may have application in a clinical setting. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):B154.