摘要
目的探讨17β-羟基类固醇脱氢酶3(17β-HSD3)抑制剂的抗前列腺癌活性、药代动力学及其急性毒性。方法取对数生长期人雄激素依赖性前列腺癌LNCaP细胞,设置空白对照组(Con组)、姜黄素组(Cur组,Cur 150µmol/L)、雄激素受体(AR)抑制剂组(AR inhibitor组,AR抑制剂5µmol/L)和姜黄素类似物H7组(H7组,H7150µmol/L),分组处理后采用xCELLience RTCA DP细胞分析仪检测各组细胞活性、流式细胞术检测细胞凋亡率、Western blot法检测细胞中Caspase-3、AR蛋白表达。C57小鼠32只,采用随机数字表法分成4组(n=8),分别为空白对照组和姜黄素类似物H7高、中、低剂量组。用1%羧甲基纤维素钠(CMC-Na)溶解H7(100、50、25 mg/kg),每天灌胃1次,空白对照组小鼠灌胃1%CMC-Na,观察小鼠有无死亡及异常现象,14 d后取材,苏木素-伊红(HE)染色,观察肝脏和肾脏的病理形态学变化。SD大鼠10只,用1%CMC-Na溶解H7(5 mg/kg)灌胃后,高效液相色谱仪(HPLC)检测血药浓度,分析药代动力学参数。另取SD雄性大鼠10只,H7(5 mg/kg)灌胃30 min后,HPLC检测大鼠脑、肝、脾、肺、小肠、胃、肾、心和睾丸中药物浓度,考察药物组织分布。结果与Con组比较,Cur组、AR inhibitor组和H7组LNCaP细胞的细胞指数(CI值)显著下降,凋亡率明显升高,Caspase-3蛋白表达水平明显升高,而AR蛋白表达水平降低(P<0.05);H7组凋亡率较Cur组和AR inhibitor组明显升高(P<0.05)。连续灌胃H714 d,小鼠未出现活动增加、流涎、抽搐、昏迷等异常现象。与Con组相比,小鼠肝脏和肾脏未见明显的病理形态学变化。药代动力学参数结果显示,测出血药浓度-时间曲线下面积(AUC)为(557.31±36.12)mg/(L·h);峰浓度(Cmax)为(36.92±1.29)mg/L;最高峰时间(t_(max))为2 h;消除半衰期(t_(1/2))为(22.13±1.74)h。H7在小肠中的浓度约是脑组织中的5000倍。结论H7作为17β-HSD3抑制剂,具有明显抗激素依赖性前列腺癌的活性和良好的药代动力学参数,无明显的急性毒性作用,以胃肠吸收为主,具有进一步研究开发前景。
Objective To investigate the anti-prostate cancer activity,pharmacokinetic analysis and acute toxicity of 17β-hydroxysteroid dehydrogenase 3(17β-HSD3)inhibitor.Methods The logarithmic growth phase androgen-dependent prostate cancer LNCaP cells were divided into the blank control group(Con group),curcumin group(Cur group,Cur 150µmol/L),androgen receptor(AR)inhibitor group(AR inhibitor group,5µmol/L)and curcumin analog H7 group(H7 group,H7150µmol/L).After grouping treatment,the xCELLience RTCA DP cell analyzer was used to detect cell viability,and flow cytometry was used to detect cell apoptosis.The changes of Caspase-3 and AR protein expression levels in cells were detected by Western blot assay.Thirty-two C57 mice were divided into 4 groups(n=8 for each group),namely the blank control group and the high,medium and low-dose curcumin analog H7 groups.The high,medium and low-dose groups of mice were treated with 1%carboxymethyl sodium cellulose(CMC-Na)dissolved H7(100,50 and 25 mg/kg),and gavage once a day.At the same time,mice in the blank control group were gavage 1%CMC-Na to observe whether the mice were dead or abnormal.After 14 days,samples were taken and stained with hematoxylin-eosin(HE)to observe the pathological changes of liver and kidney.Ten SD rats were used to gavage with 1%CMC-Na dissolved H7(5 mg/kg),and then the plasma concentration was detected by high performance liquid chromatography(HPLC)to analyze the pharmacokinetic parameters.In addition,another 10 SD rats were taken and H7(5 mg/kg)was gavage for 30 min.The drug concentrations in rat brain,liver,spleen,lung,small intestine,stomach,kidney,heart and testis were detected by HPLC,and the distribution of the drug tissue was investigated.Results Compared with the Con group,the cell index(CI value)of LNCaP cells significantly decreased in the Cur group,AR inhibitor group and H7 group.The flow cytometry results showed that the apoptosis rate was significantly increased(P<0.05).The Western blot results showed that Caspase-3 protein expression levels were significantly increased(P<0.05),while AR protein expression levels decreased(P<0.05).Compared with Cur group and AR inhibitor group,flow cytometry results showed that the apoptosis rate was significantly increased in H7 group(P<0.05).After treatment with H7 for 14 days,no abnormalities such as increased activity,salivation,convulsions and coma were found in the mice.Compared with the control group,HE staining showed no obvious pathological changes in mouse liver and kidney.The results of pharmacokinetic parameters showed that the area under the curve(AUC)was(557.31±36.12)mg/(L·h),the peak concentration(Cmax)was(36.92±1.29)mg/L,the highest peak time(t_(max))was 2 h and the elimination half-life(t_(1/2))was(22.13±1.74)h.The H7 concentrations in the small intestine were 5000 times higher than that in brain tissue.Conclusion As a 17β-HSD3 inhibitor,H7 has obvious anti-hormone-dependent prostate cancer activity and good pharmacokinetic parameters,no obvious acute toxicity,mainly gastrointestinal absorption,and has further research and development prospects.
作者
韩杨
包海花
柏合
刘洁婷
孙文慧
袁晓环
HAN Yang;BAO Hai-hua;BAI He;LIU Jie-ting;SUN Wen-hui;YUAN Xiao-huan(The First Clinical Medical College,Mudanjiang Medical University,Mudanjiang 157011,China;Medical Research Center of Mudanjiang Medical University)
出处
《天津医药》
CAS
北大核心
2021年第9期938-943,共6页
Tianjin Medical Journal
基金
2018年度黑龙江省省属高等学校基本科研业务费科研项目(2018-KYYWFMY-0045)。
