Cao Jing-Hua, Xu Bo, Li Min, Wu De-Zhu, Huang Wei, Cui Jing-Rong
State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Science, Peking University, Beijing, 100083, PR China.
Ai Zheng. 2007 Apr;26(4):361-6.
BACKGROUND & OBJECTIVE: Norcantharidin (NCTD), the demethylated form of cantharidin, can inhibit the proliferation of many kinds of cancer cells, but its effect is milder than those of other drugs. This study was to explore the inhibitory effect of Nd3, a derivative of norcantharidin, on the proliferation of human ovarian cancer cell line SKOV3, compare its antitumor effect with that of norcantharidin, and investigate its possible molecular mechanisms.
SKOV3 cells were treated with Nd3 and norcantharidin, separately. Cell proliferation was evaluated by sulforhodamine B (SRB) assay. Cell cycle distribution and apoptosis were detected by flow cytometry. The expression of Cdc2, Cyclin B1, Bax, and Bcl-2 was detected by Western blot.
When treated with 2.5, 5, 10, 20, 30, and 40 micromol/L Nd3 for 48 h, the inhibition rates of SKOV3 cells were 27.3%, 34.1%, 53.3%, 64.3%, 83.3%, and 96.7%, respectively, which were significantly higher than that of negative control cells (P<0.001). The 50% inhibition concentration of Nd3 was (25.1+/-2.3) micromol/L at 24 h, (21.8+/-2.8) micromol/L at 36 h, and (20.4+/-3.3) micromol/L at 48 h. When treated with 10, 20, 30, and 40 micromol/L Nd3 for 48 h, SKOV3 cells were arrested at G2/M phase at rates of 14.3%, 20.2%, 26.2%, and 27.9%; when treated with 30 micromol/L Nd3 for 12, 24, 36, and 48 h, the proportions of SKOV 3 cells at G2/M phase were 19.8%, 26.6%, 27.8%, and 32.0%. When treated with 40 micromol/L Nd3 for 48 h, the apoptosis rate of SKOV3 cells was significantly higher than that of control cells [(17.9+/-4.4)% vs. (2.5+/-2.8)%, P<0.01]. After treatment of Nd3, the expression of Cdc2, Cyclin B1, and Bcl-2 were down-regulated, and the expression of Bax was up-regulated.
Nd3 inhibits SKOV3 cell proliferation more than norcantharidin does, blocks cell cycle at G2/M phase, and induces apoptosis. The antitumor mechanism of Nd3 is related to the changes of Cdc2, Cyclin B1, Bax, and Bcl-2 expression.
去甲斑蝥素（NCTD）是斑蝥素的去甲基化形式，可抑制多种癌细胞的增殖，但其作用比其他药物更为温和。本研究旨在探讨去甲斑蝥素衍生物Nd3对人卵巢癌细胞系SKOV3增殖的抑制作用，将其抗肿瘤作用与去甲斑蝥素进行比较，并研究其可能的分子机制。
分别用Nd3和去甲斑蝥素处理SKOV3细胞。通过磺酰罗丹明B（SRB）法评估细胞增殖。采用流式细胞术检测细胞周期分布和凋亡情况。通过蛋白质免疫印迹法检测Cdc2、细胞周期蛋白B1、Bax和Bcl-2的表达。
用2.5、5、10、20、30和40 μmol/L的Nd3处理SKOV3细胞48小时后，抑制率分别为27.3%、34.1%、53.3%、64.3%、83.3%和96.7%，均显著高于阴性对照细胞（P<0.001）。Nd3在24小时的半数抑制浓度为（25.1±2.3）μmol/L，36小时为（21.8±2.8）μmol/L，48小时为（20.4±3.3）μmol/L。用10、20、30和40 μmol/L的Nd3处理SKOV3细胞48小时后，细胞阻滞于G2/M期的比例分别为14.3%、20.2%、26.2%和27.9%；用30 μmol/L的Nd3处理12、24、36和48小时后，SKOV3细胞在G2/M期的比例分别为19.8%、26.6%、27.8%和32.0%。用40 μmol/L的Nd3处理SKOV3细胞48小时后，其凋亡率显著高于对照细胞[（17.9±4.4）% 对 （2.5±2.8）%，P<0.01]。Nd3处理后，Cdc2、细胞周期蛋白B1和Bcl-2的表达下调，Bax的表达上调。
Nd3对SKOV3细胞增殖的抑制作用强于去甲斑蝥素，可将细胞周期阻滞于G2/M期并诱导凋亡。Nd3的抗肿瘤机制与Cdc2、细胞周期蛋白B1、Bax和Bcl-2表达的变化有关。
