Regulation of Hepatic P450s by Anti-Cholesterol Drugs

描述(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作为化学中间体发挥作用，而且还作为肝细胞生理的内分泌调节物质，改变核受体的活性。最近，许多降胆固醇药物被证明能使生物活性代谢物在肝脏内积聚，导致肝脏基因表达改变。例如，我们发现角鲨烯合成酶的抑制剂(例如，角鲨素1)选择性地诱导细胞色素P450 2B(CYP2B)基因的表达，这是通过引起内源性法尼醇的积累，从而激活构成的雄烷受体(CAR)。我们认为，法尼醇类化合物的生物活性有助于角鲨烯合成酶抑制剂的治疗和毒性作用。我们假设，由角鲨烯合成酶抑制介导的法尼醇类积聚导致肝细胞基因表达的独特变化，这些基因表达从啮齿动物到人类以及从原代培养到体内都是保守的。这些变化中的许多(例如，CYP2B的诱导)归因于法尼醇的积累和CAR的激活，并且这些变化被抑制酒精脱氢酶(ADH)催化的法尼醇代谢而增强。其他变化(例如，SLC13A3诱导)由法呢醇衍生的代谢物(S)和不依赖CAR的机制(S)介导。该建议的具体目的是(1)确定和比较(即，在大鼠、小鼠和人之间)肝细胞对角鲨烯合成酶抑制的全局基因表达反应，并确定CAR是否介导了该反应的组成部分，(2)确定角鲨烯合成酶抑制对我们在原代培养的人肝细胞中发现的人肝细胞基因表达的影响(来自目标1)是否在体内保持，(3)确定是否一个或多个ADH酶负责将肝细胞中的法尼醇转化为法尼醇，因此，ADH活性是否是抑制角鲨烯合成酶后达到的法尼醇水平和对基因表达的影响的关键决定因素，以及(4)确定抑制角鲨烯合成酶是否调节单个人CAR亚型的活性。正常人肝细胞的内源性法尼醇代谢和信号转导基本上是未知的。这些研究将为法尼醇类化合物对肝细胞脂质代谢产生有益影响的能力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角鲨烯合成酶抑制剂作为抗胆固醇药物的未来作用。 与公共卫生相关：在美国，冠心病是导致死亡的主要原因，每年导致的死亡人数超过所有形式癌症的总和，而低密度脂蛋白水平升高是导致冠心病的主要风险因素。使用他汀类药物的降胆固醇药物疗法现在是一种被证明可以降低患冠心病风险的方法，而且由于促进高胆固醇血症的行为很普遍，他汀类药物被许多人服用。然而，某些患者对他汀类药物治疗没有反应，另一些患者由于出现严重的药物不良反应而不能服用药物。因此，需要替代的降胆固醇药物。这一项目的公共卫生意义在于，它将增加我们对一类新型降胆固醇药物潜在效用的了解。

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. PUBLIC HEALTH RELEVANCE: Coronary heart disease is the leading cause of mortality in the United States, causing more deaths annually than all forms of cancer combined, and elevated low-density lipoprotein levels are a major risk factor for the development of coronary heart disease. Anti-cholesterol drug therapy, using statins, is now a proven approach for reducing the risk of developing coronary heart disease, and since behaviors that promote hypercholesterolemia are prevalent, statins are consumed by many people. However, certain patients do not respond to statin treatment and others cannot take the drugs due to the development of severe adverse drug reactions. Therefore alternative anti-cholesterol drugs are needed. The public health relevance of this project is that it will increase our knowledge about the potential utility of a novel class of anti-cholesterol drugs.