Regulation of Hepatic P450s by Anti-Cholesterol Drugs

描述（由申请人提供）：胆固醇生物合成途径的内源性代谢物现在被认为不仅是化学中间体，而且是肝细胞生理的内分泌调节剂，可以改变核受体的活性。最近，许多“抗胆固醇药物”被证明会导致生物活性代谢物在肝脏中积累，导致肝脏基因表达改变。例如，我们发现角鲨烯合成酶抑制剂（如角鲨estatin 1）通过引起内源性法内甾体积累来选择性地诱导细胞色素P450 2B （CYP2B）基因表达，从而激活构成型雄烷受体（CAR）。我们认为，角鲨烯合成酶抑制剂的生物活性有助于治疗和毒性作用。我们假设，角鲨烯合成酶抑制介导的法脂类积累导致肝细胞基因表达的显著变化，这种变化从啮齿动物到人类，从原代培养到体内都是保守的。许多这些变化（例如，CYP2B诱导）可归因于法尼醇积累和CAR激活，并且这些变化通过抑制醇脱氢酶（ADH）催化的法尼醇代谢而增强。其他变化（如Slc13A3诱导）是由法尼醇衍生代谢物和car独立机制介导的。本提案的具体目的是：(1)定义和比较（即在大鼠，小鼠和人之间）肝细胞对角鲨烯合酶抑制的整体基因表达反应，并确定CAR是否介导这种反应的组成部分，(2)确定我们在原代培养的人肝细胞中发现的角鲨烯合酶抑制对人肝细胞基因表达的影响是否在体内维持（来自Aim 1）。(3)确定一种或多种ADH酶是否负责将肝细胞中的法尼醇转化为法尼醇，从而确定ADH活性是否是角鲨烯合酶抑制后实现的法尼醇水平和对基因表达的影响的关键决定因素；(4)确定角鲨烯合酶抑制是否调节个体人类CAR同型体的活性。正常人肝细胞的内源性法内甾体代谢和信号传导基本上尚未被探索。这些研究将为类法内酯对肝细胞脂质代谢的有益作用以及促进肝毒性的作用提供新的信息。这一信息将直接关系到未来角鲨烯合成酶抑制剂作为抗胆固醇药物的作用。

DESCRIPTION (provided by applicant): Endogenous metabolites of the cholesterol biosynthetic pathway are now recognized to function not only as chemical intermediates, but also as intracrine regulators of hepatocyte physiology, which modify the activities of nuclear receptors. Recently, many "anti-cholesterol drugs" have been shown to cause bioactive metabolites to accumulate in the liver, resulting in altered hepatic gene expression. For example, we found that inhibitors of squalene synthase (e.g., squalestatin 1) selectively induce cytochrome P450 2B (CYP2B) gene expression by causing an endogenous farnesoid to accumulate, which activates the constitutive androstane receptor (CAR). We propose that the biological activities of the farnesoids contribute to the therapeutic as well as the toxic effects of squalene synthase inhibitors. We hypothesize that farnesoid accumulation mediated by squalene synthase inhibition causes distinctive changes in hepatocellular gene expression that are conserved from rodent to human and from primary culture to in vivo. Many of these changes (e.g., CYP2B induction) are attributable to farnesol accumulation and CAR activation, and these changes are enhanced by inhibition of alcohol dehydrogenase (ADH)-catalyzed farnesol metabolism. Other changes (e.g., Slc13A3 induction) are mediated by a farnesol-derived metabolite(s) and a CAR-independent mechanism(s). The specific aims of this proposal are (1) to define and compare (i.e., among rat, mouse, and human) the global gene expression response of the hepatocyte to squalene synthase inhibition, and to determine whether CAR mediates the components of this response, (2) to determine whether the effects of squalene synthase inhibition on human hepatocellular gene expression that we identify in primary cultured human hepatocytes (from Aim 1) are maintained in vivo, (3) to determine whether one or more ADH enzymes are responsible for converting farnesol to farnesal in hepatocytes, and hence whether ADH activity is a critical determinant of the farnesoid levels and effects on gene expression that are achieved following squalene synthase inhibition, and (4) to determine whether squalene synthase inhibition modulates the activities of individual human CAR isoforms. Endogenous farnesoid metabolism and signaling in normal human hepatocytes has been essentially unexplored. These studies will provide new information about the abilities of farnesoids to produce beneficial effects on hepatocellular lipid metabolism, as well as effects that promote hepatotoxicity. This information will be directly relevant to the future role of squalene synthase inhibitors as anti-cholesterol drugs.