MECHANISMS OF INDUCTIVE DRUG INTERACTIONS WITH ANTI-HIV PROTEASE INHIBITORS

抗HIV蛋白水解酶抑制剂(PRI)是治疗艾滋病的一线药物，目前常规与其他抗HIV药物联合使用，包括其他蛋白酶抑制剂、核苷(如叠氮胸苷)和非核苷逆转录酶抑制剂(如地拉韦林)。PrI是细胞色素P3A酶和多药耐药(MDR)转运蛋白MDR1和MRP2的底物。在处方PRI中遇到的一个重大临床问题是它们倾向于产生临床上显著的药物相互作用，包括诱导性和抑制性， 对艾滋病患者进行常规的药物治疗。到目前为止，药物与PrI相互作用的机制和程度的研究重点一直是Pris在体外和体内对CYP3A酶的有效抑制能力。然而，人们对PRI诱导各种药物清除的机制以及它们诱导自身清除(自身诱导)的机制知之甚少。中使用的PRI的 临床上，利托那韦和奈非那韦最常被报道为诱导性药物相互作用和自身诱导。利托那韦最令人惊讶的是诱导性药物相互作用和自身诱导，因为利托那韦在体外可以灭活细胞色素P3A酶，并在体内有效地抑制细胞色素P3A4/5，这些酶负责清除利托那韦和其他PRI。与利福平一样，利托那韦似乎是一种广谱的酶和转运蛋白诱导剂。这一观察结果与利托那韦是人类孤儿的优秀配体/激活剂是一致的 核受体，hPXR(孕烷X受体)。令人惊讶的是，奈非那韦不是hPXR的配体/激活剂，但它是一种自身诱导剂，并导致诱导性药物相互作用。因此，我们假设利托那韦和奈非那韦通过不同的机制自动诱导和引起诱导性药物相互作用。利托那韦主要通过激活hPXR来调节其诱导效应，而奈非那韦则通过其他机制(例如激活其他孤儿核受体)来调节其诱导效应。这一建议的具体目的是为了验证这一假说，并阐明PrI在体外和体内诱导CYP酶和MDR转运蛋白的机制和程度。 具体目标 1.利用人肝细胞和人肠道细胞系，研究Pris诱导的细胞色素P450酶和多药耐药转运蛋白的表达。 2.确定利托那韦和奈非那韦在体外诱导的细胞周期蛋白和多药耐药转运蛋白在体内是否也有类似程度的诱导。 3.确定MDR1基因的已知变异是否决定利托那韦和奈非那韦在体内的分布，从而决定这些PrI对MDR转运体和CyP酶的体内诱导。 目的：探讨利托那韦和奈非那韦诱导细胞色素P450酶和多药耐药转运蛋白的分子机制。

Anti-HIV protease inhibitors (PRIs) are frontline drugs in the treatment of AIDS, and are now routinely administered in combination with other anti-HIV drugs including other protease inhibitors, nucleoside (e.g. azidothymidine) and non-nucleoside reverse transcriptase inhibitors (e.g. delavirdine). PRIs are substrates of CYP3A enzymes and of the multidrug resistance (MDR) transporters, MDR1 and MRP2. A significant clinical problem encountered in prescribing PRIs is their propensity to produce clinically significant drug interactions, both inductive and inhibitory, with drugs routinely administered to people with AIDS. To date, the focus of studies on the mechanisms and extent of drug interactions with PRIs has been on the capacity of the PRIs to potently inhibit CYP3A enzymes, both in vitro and in vivo. However, little is known about the mechanisms by which PRIs induce clearance of various drugs, as well as the mechanisms by which they cause induction of their own clearance (autoinduction). Of the PRIs used in the clinic, clinically significant inductive drug interactions and autoinduction have been reported most frequently with ritonavir and nelfinavir. Inductive drug interactions and autoinduction by ritonavir are most surprising as ritonavir inactivates CYP3A enzymes in vitro and potently inhibits CYP3A4/5 in vivo, enzymes responsible for the clearance of ritonavir and other PRIs. Like rifampin, ritonavir appears to be a broad-spectrum inducer of enzymes and transporters. This observation is consistent with ritonavir being an excellent ligand/activator of the human orphan nuclear receptor, hPXR (pregnane X receptor). Surprisingly, nelfinavir is NOT a ligand/activator of hPXR, and yet it is an autoinducer and causes inductive drug interactions. Thus, we have hypothesized that ritonavir and nelfinavir autoinduce and cause inductive drug interactions via different mechanisms. Ritonavir mediates its inductive effects primarily by activating hPXR, while nelfinavir does so via other mechanisms (e.g. activating other orphan nuclear receptors). The specific aims of this proposal are directed towards testing this hypothesis and towards elucidating the mechanisms and extent of in vitro and in vivo induction by PRIs of CYP enzymes and MDR transporters. Specific Aims 1. To determine, using human hepatocytes and human intestinal cell lines, the CYP enzymes and MDR transporters that are significantly induced by PRIs. 2. To determine if CYPs and MDR transporters significantly induced by ritonavir and nelfinavir in vitro, are induced to a similar extent in vivo in humans. 3. To determine if known variants of the MDR1 gene determine the in vivo disposition of ritonavir and nelfinavir and, therefore, the in vivo induction by these PRIs of MDR transporters and CYP enzymes. To determine the molecular mechanisms by which CYP enzymes and MDR transporters are induced by ritonavir and nelfinavir.