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借助体外代谢孵育体系考察了金松双黄酮(sciadopitysin,SP)对12种人尿苷二磷酸葡糖醛酸转移酶(UGTs)的抑制作用,通过体外-体内外推(IV-IVE)预测体内药物-药物相互作用(DDI)的风险。以混合人肝微粒体(HLM)及重组表达的人UGTs作为酶源,选用4-甲基伞形酮(4-MU)、三氟拉嗪(TFP)、N-3-羧丙基-4-羟基-1,8-萘酰亚胺(NCHN)分别为UGTs酶的广谱探针底物、UGT1A4和UGT1A1的特异性探针底物,评估金松双黄酮对12种人UGT酶的抑制作用。通过非线性拟合求得半数最大抑制浓度IC50和抑制动力学常数Ki及抑制类型;并基于体外参数预测了金松双黄酮通过抑制UGT1A1引发的潜在DDI风险。体外抑制实验表明,金松双黄酮对UGT1A1、UGT1A3、UGT1A8和UGT1A10有较强的抑制作用,当终浓度为10μmol·L~(-1)时,UGT1A1、UGT1A3、UGT1A8和UGT1A10的残余活性均小于30%。金松双黄酮对UGT1A1、UGT1A3、UGT1A8和UGT1A10的半数最大抑制浓度IC50为0.20~1.34μmol·L~(-1),抑制动力学常数Ki为0.07~2.12μmol·L~(-1)。口服金松双黄酮240 mg·d~(-1)可导致UGT1A1底物的AUC增加19%~147%。金松双黄酮可竞争性的抑制UGT1A1、UGT1A3、UGT1A8和UGT1A10催化的4-MU-O-葡糖醛酸化反应及UGT1A1催化的NCHN-O-葡糖醛酸化反应。金松双黄酮对UGT1A1的强抑制作用有可能减缓UGT1A1底物的代谢,进而引发DDI风险。上述研究提示金松双黄酮与临床药物联合应用时,应该特别注意其通过抑制UGT酶而引发的DDI。
The inhibitory effect of sciadopitysin (SP) on 12 human uridine diphosphate glucuronosyltransferases (UGTs) was investigated by means of in vitro metabolic incubation system, and in vivo drug (IV-IVE) - Risk of drug interactions (DDI). Using mixed human liver microsomes (HLM) and recombinant human UGTs as enzyme source, 4-methylumbelliferone (4-MU), trifluoperazine (TFP) and N-3- - Hydroxy-1,8-naphthalimide (NCHN) are broad-spectrum probe substrates for UGTs enzymes, UGT1A4 and UGT1A1-specific probe substrates, respectively, to evaluate the inhibitory effect of Ginseng biflavone on 12 human UGT enzymes . The half maximal inhibitory concentrations (IC50), inhibition kinetic constants (Ki), and inhibition types were determined by non-linear fitting. The potential DDI risk caused by the inhibition of UGT1A1 was also estimated based on in vitro parameters. In vitro inhibition experiments showed that Ginseng biflavonoids had a strong inhibitory effect on UGT1A1, UGT1A3, UGT1A8 and UGT1A10. The residual activities of UGT1A1, UGT1A3, UGT1A8 and UGT1A10 were all less than 30 when the final concentration was 10μmol·L -1 %. The IC50 value of the biflavone for UGT1A1, UGT1A3, UGT1A8 and UGT1A10 was 0.20 ~ 1.34μmol·L -1, and the inhibition kinetic constant Ki was 0.07 ~ 2.12μmol·L -1. Oral ginsenoside 240 mg · d ~ (-1) increased the AUC of UGT1A1 substrate by 19% -147%. Ginseng biflavonoids competitively inhibit 4-MU-O-glucuronidation catalyzed by UGT1A1, UGT1A3, UGT1A8 and UGT1A10 and UGT1A1-catalyzed NCHN-O-glucuronidation. The strong inhibition of UGT1A1 by Ginseng biflavonoids may reduce the metabolism of UGT1A1 substrate, which may lead to the risk of DDI. The above study suggests that when combined with clinical drugs, the special reference should be made to DDI induced by UGT enzyme inhibition.