Structural Requirements for Sterol 14alpha-Demethylases

甾醇 14α-脱甲基酶的结构要求

• 批准号: 9026357
• 负责人: Galina I Lepesheva
• 金额: $ 42.34万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-01-01 至 2019-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9026357
• 关键词: Achievement; Address; Adverse effects; Agriculture; Alcohols; Aldehydes; Alzheimer' s Disease; Amino Acid Sequence; Animal Model; Antifungal Agents; Aspergillus fumigatus; Azoles; Binding; Biochemical; Biological; Biological Process; Biological Response Modifier Therapy; Cancer cell line; Catalysis; Cells; Chagas Disease; Chemotherapy-Oncologic Procedure; Cholesterol; Clinical; Clinical Trials; Complex; Custom; Cytochrome P450; Data; Development; Disease; Drug Design; Drug Targeting; Drug effect disorder; Drug resistance; Electron Transport; Electrons; Enzymes; Ergosterol; Family; Formic Acids; Funding; Future; Goals; Growth; Growth and Development function; Health; Human; Industrial fungicide; Infection; Knowledge; Lead; Malignant Neoplasms; Membrane Fluidity; Membrane Proteins; Minor; Mixed Function Oxygenases; Modification; Molecular; Mycoses; NADPH-Ferrihemoprotein Reductase; Organism; Outcome; Parasites; Permeability; Pharmaceutical Preparations; Phylogeny; Phytosterols; Plants; Positioning Attribute; Process; Progress Reports; Proteins; Protozoa; Public Health; Publications; Reaction; Research; Resistance; Roentgen Rays; Sterol Biosynthesis Pathway; Sterols; Structure; Testing; Trypanosoma cruzi; Visceral; Visceral Leishmaniasis; Voriconazole; arm; base; cancer cell; cell growth; chemotherapy; cholesterol biosynthesis; design; drug candidate; drug discovery; electron donor; fungus; human disease; inhibitor/antagonist; insight; killings; member; membrane biogenesis; methyl group; microbial; novel; novel therapeutics; pathogen; peptidomimetics; pharmacophore; posaconazole; prevent; protein protein interaction; resistant strain; scaffold; screening

 DESCRIPTION (provided by applicant) Sterol 14α-demethylase (CYP51) is the monooxygenase that catalyzes a unique three step reaction (14α- methyl →14α-alcohol→14α-aldehyde→14α-demethylated product plus formic acid), removing the 14α- methyl group from the cyclized sterol precursors. CYP51 is regarded as a possible evolutionary ancestor to all members of the currently existing P450 superfamily. The CYP51 reaction is required for biosynthesis of cholesterol in humans, ergosterol in eukaryotic microbial human pathogens (such as fungi and protozoa), and various phytosterols in plants. The vast majority of the sterols are utilized for the formation of viable eukaryotic membranes (fluidity, permeability, modulation of multiple functions of integrated membrane proteins), while some sterols serve as precursors for a number of regulatory molecules essential for cell growth, development, and multiplication. Blocking sterol biosynthesis in unicellular organisms is lethal. CYP51 inhibitors (azoles) are used as the major clinical antifungal drugs and agricultural fungicides. Two of these drugs (posaconazole and ravuconazole) are presently in clinical trials for the deadly human infection caused by the protozoan pathogen Trypanosoma cruzi (Chagas disease). All of the azoles currently in use were discovered empirically, i.e. by screening their effects on fungal cell growth, and are far from ideal in terms of efficiency, safet, side effects, and sensitivity to resistance. The long-term goal of our research is to understand the molecular basis for the CYP51 functional conservation, drug action, and drug resistance. Here we propose to apply the information on the CYP51 structure/function that was gained during the previous cycles of funding to rationalize the CYP51-targeting drug discovery paradigm. We will perform protozoan/fungal/human CYP51 structure-guided derivatization of the highly potent, non-toxic experimental VNI scaffold to make pharmacologically-optimized drug candidates suitable for clinical trials for Chagas disease (including the infections caused by naturally resistant strains of the parasite) and visceral leishmaniasis (Aim 1a), as well as to develop new custom-designed inhibitors of fungal CYP51s (Aim 1b). Aim 2 will evaluate human CYP51 as a potential drug target for cholesterol- related human diseases, including cancer and Alzheimer's disease. Aim 3 of this proposal is to determine the structure of CYP51 in complex with its electron donor partner, NADPH-cytochrome P450 reductase, which will reveal the key residues involved in the protein-protein interaction, and may lead to the development of a novel type of CYP51 inhibitors. The results of this research will direct future approaches in drug discovery, enabling the creation of rationally designed species-oriented drug candidates directed at saving millions of human lives.

 描述（由申请人提供） 甾醇 14α-去甲基酶 (CYP51) 是一种单加氧酶，可催化独特的三步反应（14α-甲基→14α-醇→14α-醛→14α-去甲基化产物加甲酸），从环化甾醇前体中去除 14α-甲基。 CYP51 被认为是现有 P450 超家族所有成员的可能进化祖先。 CYP51反应是人体胆固醇、真核微生物人类病原体（例如真菌和原生动物）中麦角甾醇以及植物中各种植物甾醇的生物合成所必需的。绝大多数甾醇用于形成活的真核细胞膜（流动性、渗透性、整合膜蛋白多种功能的调节），而一些甾醇作为细胞生长、发育和增殖所必需的许多调节分子的前体。阻断单细胞生物中的甾醇生物合成是致命的。 CYP51抑制剂（唑类）是临床主要的抗真菌药物和农业杀菌剂。其中两种药物（泊沙康唑和拉夫康唑）目前正处于临床试验阶段，用于治疗由原生动物病原体克氏锥虫（恰加斯病）引起的致命人类感染。 目前使用的所有唑类药物都是凭经验发现的，即通过筛选它们对真菌细胞生长的影响，在效率、安全性、副作用和耐药性敏感性方面远非理想。我们研究的长期目标是了解 CYP51 功能保守、药物作用和耐药性的分子基础。在这里，我们建议应用在之前的资助周期中获得的有关 CYP51 结构/功能的信息来合理化 CYP51 靶向药物发现范式。我们将对高效、无毒的实验性 VNI 支架进行原生动物/真菌/人类 CYP51 结构引导的衍生化，以制备适合恰加斯病（包括由恰加斯病引起的感染）临床试验的药理学优化候选药物。 寄生虫的天然耐药菌株）和内脏利什曼病（目标 1a），以及开发新的定制设计的真菌 CYP51 抑制剂（目标 1b）。目标 2 将评估人类 CYP51 作为胆固醇相关人类疾病（包括癌症和阿尔茨海默病）的潜在药物靶点。该提案的目标 3 是确定 CYP51 与其电子供体伙伴 NADPH-细胞色素 P450 还原酶复合物的结构，这将揭示参与蛋白质-蛋白质相互作用的关键残基，并可能导致新型 CYP51 抑制剂的开发。这项研究的结果将指导未来的药物发现方法，从而能够创建合理设计的面向物种的候选药物，从而拯救数百万人的生命。