Thymoquinone from Nigella sativa Seeds Promotes the Antitumor Activity of Noncytotoxic Doses of Topotecan in Human Colorectal Cancer Cells in Vitro.

Topotecan, a topoisomerase I inhibitor, is an anticancer drug widely used in the therapy of lung, ovarian, colorectal, and breast adenocarcinoma. Due to the primary dose-limiting toxicity of topotecan, which is myelosuppressive, it is necessary to identify other chemotherapeutic agents that can work synergistically with topotecan to increase its efficacy and limit its toxicity. Many studies have shown synergism upon the combination of topotecan with other chemotherapeutic agents such as gemcitabine. Other studies have demonstrated that pre-exposing cells to naturally occurring compounds such as thymoquinone, followed by gemcitabine or oxaliplatin, resulted in higher growth inhibition compared to treatment with gemcitabine or oxaliplatin alone. Our aim was to elucidate the underlying mechanism of action of topotecan in the survival and apoptotic pathways in human colon cancer cell lines in comparison to thymoquinone, to study the proapoptotic and antiproliferative effects of thymoquinone on the effectiveness of the chemotherapeutic agent topotecan, and to investigate the potential synergistic effect of thymoquinone with topotecan. Cells were incubated with different topotecan and thymoquinone concentrations for 24 and 48 hours in order to determine the IC50 for each drug. Combined therapy was then tested with ± 2 values for the IC50 of each drug. The reduction in proliferation was significantly dose- and time-dependent. After determining the best combination (40 µM thymoquinone and 0.6 µM topotecan), cell proteins were extracted after treatment, and the expression levels of B-cell lymphoma 2 and of its associated X protein, proteins p53 and p21, and caspase-9, caspase-3, and caspase-8 were studied by Western blot. In addition, cell cycle analysis and annexin/propidium iodide staining were performed. Both drugs induced apoptosis through a p53-independent mechanism, whereas the expression of p21 was only seen in thymoquinone treatment. Cell cycle arrest in the S phase was detected with each compound separately, while combined treatment only increased the production of fragmented DNA. Both compounds induced apoptosis through the extrinsic pathway after 24 hours; however, after 48 hours, the intrinsic pathway was activated by topotecan treatment only. In conclusion, thymoquinone increased the effectiveness of the chemotherapeutic reagent topotecan by inhibiting proliferation and lowering toxicity through p53- and Bax/Bcl2-independent mechanisms.

Effects of Nigella sativa and thymoquinone on biochemical and subcellular changes in pancreatic ß-cells of streptozotocin-induced diabetic rats.

BACKGROUND: The present study investigated the effects of Nigella sativa aqueous extract and oil, as well as thymoquinone, on serum insulin and glucose concentrations in streptozotocin (STZ) diabetic rats. METHODS: Rats were divided into five experimental groups (control, untreated STZ-diabetic, and aqueous extract-, oil-, or thymoquinone-treated diabetic rats). Treated rats received 2 mL/kg, i.p., 5%N. sativa extract, 0.2 mL/kg, i.p., N. sativa oil, or 3 mg/mL, i.p., thymoquinone 6 days/week for 30 days. Serum insulin and glucose concentrations, superoxide dismutase (SOD) levels, and pancreatic tissue malondialdehyde (MDA) were determined. Electron microscopy was used to identify any subcellular changes. RESULTS: Diabetes increased tissue MDA and serum glucose levels and decreased insulin and SOD levels. Treatment of rats with N. sativa extract and oil, as well as thymoquinone, significantly decreased the diabetes-induced increases in tissue MDA and serum glucose and significantly increased serum insulin and tissue SOD. Ultrastructurally, thymoquinone ameliorated most of the toxic effects of STZ, including segregated nucleoli, heterochromatin aggregates (indicating DNA damage), and mitochondrial vacuolization and fragmentation. The aqueous extract of N. sativa also reversed these effects of STZ, but to a lesser extent. The N. sativa oil restored normal insulin levels, but failed to decrease serum glucose concentrations to normal. CONCLUSIONS: The biochemical and ultrastructural findings suggest that N. sativa extract and thymoquinone have therapeutic and protect against STZ-diabetes by decreasing oxidative stress, thus preserving pancreatic ß-cell integrity. The hypoglycemic effect observed could be due to amelioration of ß-cell ultrastructure, thus leading to increased insulin levels. Consequently, N. sativa and thymoquinone may prove clinically useful in the treatment of diabetics and in the protection of ß-cells against oxidative stress.