目的 基于信号转导和转录活化因子3（STAT3）信号通路探讨三氟甲基化二氢喹喔啉酮类化合物S-3-（三氟甲基）-6，7-双［4-（三氟甲基）苯基］-3，4-二氢喹啉-2（1H）-酮（2o）对人结直肠癌细胞生长的抑制作用。方法 采用 MTT法检测三氟甲基化二氢喹喔啉酮类化合物 2b、2e、2f、2g、2k、2l、2m、2o、2q（20 μmo·L^-1）作 用 48 h 对 人 结 直 肠 癌 细 胞DLD-1 存活率的影响，检测化合物2o（0.5、1.0、2.0、5.0、7.5、10.0、12.5、15.0、17.5 μmo·L^-1）作用48 h对人结直肠癌细胞HCT116、RKO和DLD-1存活率的影响；克隆形成实验检测化合物2o（2.5、5.0、10.0 μmo·L^-1）对RKO、DLD-1、HCT116细胞增殖的影响；流式细胞术和 Hoechst染色检测化合物 2o 对细胞凋亡的影响；细胞划痕实验检测化合物 2o 对 DLD-1 和RKO 细胞迁移率的影响；Western blotting 法检测化合物 2o 对 RKO 细胞 p-STAT3、STAT3、骨髓白血病细胞分化蛋白（MCL）-1、生存素（Survivin）、B淋巴细胞瘤-2（Bcl-2）、Bcl-2相关X蛋白（Bax）蛋白表达水平的影响，检测化合物2o对白细胞介素-6（IL-6）刺激下p-STAT3、STAT3蛋白表达水平的影响。结果 化合物2o对DLD-1细胞的杀伤能力较其他化 合 物 强 ； 化 合 物 2o 对 HCT116、 RKO 和 DLD-1 细 胞 的 半 数 抑 制 浓 度 （IC₅₀） 分 别 为 （3.573±0.172）、（8.056±0.458）、（6.226±0.458）μmo·L^-1。与对照组比较，化合物2o明显降低了HCT116、RKO和DLD-1细胞的集落形成能力；显著降低DLD-1和RKO细胞迁移率（P＜0.01、0.001）；明显增加RKO、DLD-1细胞凋亡率；明显增加HCT116和RKO细胞核亮染比例；明显降低p-STAT3、MCL-1、Survivin、Bcl-2蛋白表达，明显升高Bax蛋白表达，对STAT3蛋白表达无明显影响。与对照组比较，IL-6刺激的模型组p-STAT3蛋白表达明显升高，STAT3蛋白表达无明显变化；与模型组比较，随着化合物2o浓度的升高，p-STAT3蛋白表达明显降低，STAT3蛋白表达无明显变化。结论 化合物2o可能通过下调STAT3信号通路发挥抗人结直肠癌作用。

Objective To investigate the inhibitory effect of S-3- (trifluoromethyl) -6, 7-bis(4- (trifluoromethyl)phenyl) -3, 4- dihydroquinoxalin-2(1H)-one on the occurrence and development of human colorectal cancer cells based on signal transducer and activator of transcription 3 (STAT3) signaling pathway. Methods MTT methods was used to detected the effects of trifluoromethylated dihydroquinoxalinones 2b, 2e, 2f, 2g, 2k, 2l, 2m, 2o, 2q (20 μmo·L^-1) treatment for 48 hours on the survival rate of human colorectal cancer cell line DLD-1, and the effects of compound 2o (0.5, 1.0, 2.0, 5.0, 7.5, 10.0, 12.5, 15.0, 17.5 μmo·L^-1) treatment for 48 h on the survival rates of human colorectal cancer cells HCT116, RKO, and DLD-1. Clonogenesis assay detected effect of compound 2o (2.5, 5.0, 10.0 μmo·L^-1) on the proliferation of RKO, DLD-1, and HCT116 cells. Flow cytometry and Hoechst staining were used to detect the effect of compound 2o on apoptosis. The effect of compound 2o on the migration rate of DLD-1 and RKO cells was detected by cell scratch test. Western blotting was used to detect the effects of compound 2o on the expression of p-STAT3, STAT3, myeloid leukemia cell differentiation protein (MCL)-1, Survivin, B-lymphomatoma-2 (Bcl-2), and Bcl-2 related X protein (Bax) proteins in RKO cells, and to detect the effects of compound 2o on the expression of p-STAT3, STAT3 proteins stimulated by interleukin-6 (IL-6). Results Compound 2o had stronger killing ability on DLD-1 cells than other compounds. The semi maximum inhibitory concentrations (IC₅₀) of compound 2o on HCT116, RKO, and DLD-1 cells were (3.573 ± 0.172), (8.056 ± 0.458), and (6.226 ± 0.458) μmo·L^-1 respectively. Compared with the control group, compound 2o apparently decreased the colony forming ability of HCT116, RKO, and DLD-1 cells, significantly decreased the migration rates of DLD-1 and RKO cells (P＜0.01, 0.001), apparently increased the apoptosis rate of RKO and DLD-1 cells, apparently increased the ratio of HCT116 and RKO nuclei to bright staining, apparently decreased the expression of p-STAT3, MCL-1, Survivin, and Bcl-2 proteins, apparently increased the expression of Bax protein, and had no significant impact on STAT3 protein expression. Compared with the control group, the expression of p-STAT3 protein in the model group stimulated by IL-6 was significantly increased, while the expression of STAT3 protein had no significant change. Compared with the model group, with the increase of compound 2o concentration, the expression of pSTAT3 protein significantly decreased, while the expression of STAT3 protein did not significantly change. Conclusion Our data suggested that compound 2o may play an anti human colorectal cancer role by downregulating STAT3 signaling pathway.

R965

大理药业股份有限公司横向课题（KJHX1603）