糖尿病状态下药物摄取转运体,CYP450酶和外排转运体功能与表达差异性改变机制及其对药物处置影响

It is well known that drug disposition is dependent on orderly coordination of influx transporters, drug metabolism enzymes and efflux transporters of drugs. We aimed to investigate that diabetes differentially altered function and expression of hepatic and intestinal influx transporters(OATPs), CYP450(such as CYP3A) and efflux transporters(such as P-GP) as well as mechanism leading to these differential alterations using type 2 diabetic rats as model. Function and expression of these targets in intestine and liver of diabetic rats were measured using corresponding probes, Western blot and QT-PCR, respectively. The mechanisms resulting in the differential alterations among influx transporters, CYP450s, and efflux transporters were also documented. The individual contribution of these targets to pharmacokinetics as well as combined effects were quantitatively predicted. Cell lines or primarily cultured cells were used to investigate which compounds in serum of diabetic rats led to differential alterations in function and expression of influx transporters, CYP450s and efflux transporters. The cell lines with gene transfection and gene silencing were used to investigate cross-talk among influx transporters, CYP450 and efflux transporters. Effects of differential alterations in drug transporters and CYP450s on pharmacokinetics of drug such as atorvastatin as well as their roles in hepatotoxicity of atorvastatin under diabetic status were documented. A physiologically-based pharmacokinetic model(PBPK) characterizing both diabetes and data on function/expression of influx transporters and CYP450s as well as efflux transporters was developed and applied to quantitatively predict pharmacokinetics of model drugs such as atorvastatin in diabetic rats and patients. The predicted results were further compared with observed data to try to resolve the scientific problem how to quantitative predict pharmacokinetics from in vitro to in vivo and animal to human based on both disease and alterations in CYP450s/transporters. The results may highlight differential alterations in influx transporters, CYP450s and efflux transporters induced by diabetes , their mechanism leading to differential alterations, as guidance for drug use and prediction of pharmacokinetics in diabetic patients.

药物处置依赖药物摄取转运体，代谢酶和外排转运体间有序协同作用。以2型糖尿病大鼠, 用探针和分子生物学技术，从摄取转运体(OATPs),CYP450酶(CYP3A)和外排转运体(P-GP)环节研究糖尿病鼠肝和肠CYP450酶和转运体功能与表达差异性改变机制，CYP450酶和转运体改变各自对药物处置贡献和整合效应。用细胞研究糖尿病血清中引起这些改变的成分；用基因沉默和转染研究摄取转运体，CYP450酶和外排转运体间交互作用；CYP450酶和转运体差异性改变对阿托伐他汀处置影响及其与肝毒性增加的关系。根据CYP450酶和转运体各自对药物处置贡献，建立基于疾病兼顾摄取转运体，CYP450酶和外排转运体改变的生理模型，预测阿托伐他汀等药物在糖尿病鼠和人中处置，解决基于疾病的CYP450酶和转运体改变药物处置预测问题。对于阐明糖尿病引起CYP450酶和转运体差异性改变机制，临床合理用药等有重要意义

药物处置依赖药物摄取转运体，代谢酶和外排转运体间有序协同作用。用高脂饲养加低剂量链硫脲菌素诱导糖尿病(DM)大鼠研究: 1）DM鼠肠/肝有机阴离子转运蛋白(Oatps)，P-糖蛋白(P-gp)，乳腺癌耐药蛋白(Bcrp)和Cyp450功能与表达改变及机制；2)代谢酶和转运体改变对阿托伐他汀，头孢氨苄，伐昔洛韦和那格列奈体内处置影响；3）DM鼠肝Oatp1b2和Cyp3a1改变与阿托伐他汀肝毒性增加关系；4）用基于疾病兼顾转运体-代谢酶联盟生理药代动力学(PBPK)模型预测体内药物处置。结果:1）DM鼠肝Oatp1b2, Cyp3a1和 P-gp功能与表达显著上调，而肠Oatp1a5, Cyp3a1和P-gp功能与表达则显著下调。肠Bcrp功能与表达呈糖尿病程依赖性，10天DM鼠肠Bcrp功能与表达降低，22天DM鼠肠Bcrp功能与表达上调。灌胃阿托伐他汀在DM鼠内处置改变呈病程依赖性，10天DM鼠血浆中阿托伐他汀暴露水平显著增加， 22天DM鼠血浆中阿托伐他汀暴露水平显著降低。2)用基于代谢酶-转运体联盟PBPK 模型定量预测肝/肠代谢酶和转运体各自改变对阿托伐他汀处置改变贡献和整合效应。模型分析显示转运体和代谢酶对阿托伐他汀处置贡献：肠Bcrp>肝Oatp1b2>肝Cyp3a1>肠Cyp3a1>肠P-gp>肠Oatp1a5。糖尿病程依赖性口服阿托伐他汀处置改变主要归功于肠Bcrp变化。PBPK模型也可用于预测人肝/肠代谢酶和转运体功能与表达改变后药物处置。3)MD鼠肝Oatp1b2和Cyp3a1表达与活性上调，增加阿伐他汀肝摄取和代谢，产生大量ROS，增加其肝毒性。4) DM鼠肠类肽转运体1(Pept1)和单羧酸转运体6(MCT6)功能与表达下调，导致灌胃类肽药物(头孢氨苄和伐昔洛韦)和那格列奈暴露显著降低。5)DM鼠肠CYP450s和转运体功能与表达改变可能与肠道内环境改变如短链脂肪酸(SCFAs)浓度增加、胆汁酸盐水平紊乱有关。SCFAs通过激活PPAR而下调MCT6表达,但上调BCRP表达。SCFAs通过抑制HDAC/NF-kB通路而下调P-gp表达。胆酸盐通过激活FXR下调Pept1表达。其成果对于阐明DM状态下药物代谢酶和转运体表达与功能各自改变对药物处置改变贡献,整合效应及对药物体内处置和活性/毒性的影响，对临床合理用药和药物不良反应预测有重意义