Discovery of New Anti-amoeba Therapeutics

新抗阿米巴疗法的发现

基本信息

批准号：
9495660
负责人：
William David Nes
金额：
$ 31.94万
依托单位：
TEXAS TECH UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-06-22 至 2020-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9495660
关键词：
ADME Study Acanthamoeba Acanthamoeba Keratitis Active Sites Amebiasis Amebicides Amino Acid Sequence Amoeba genus Amphotericin B Anabolism Animals Antifungal Agents Azoles Binding Binding Sites Biochemical Biological Availability Catalysis Characteristics Cholesterol Collaborations Combined Modality Therapy Complex Cornea Crystallization Cultured Cells Cyst Disease Drug Targeting Drug usage Effectiveness Electronics Enzymes Ergosterol Genome Goals Growth Growth Inhibitors Health Health Sciences Healthcare Human In Vitro Incubated Infection Ketoconazole Lanosterol Lead Medical Medical center Metabolic Control Metabolism Methyltransferase Mus Naegleria Naegleria fowleri Orthologous Gene Outcome Parasites Pathogenicity Pathway interactions Pharmacology Pharmacotherapy Phase Property Protein Isoforms Protozoa Reaction Recombinant Proteins Research Research Personnel Rodent Model Roentgen Rays Series Side Specificity Steroids Sterols Structure Testing Texas Therapeutic Toxic effect Universities Use Effectiveness Vision analog base cholesterol biosynthesis design drug candidate druggable target enzyme biosynthesis fungus improved in vitro testing in vivo inhibitor/antagonist lead candidate medical schools methyl group molecular recognition mouse model novel pathogen preference primary amebic meningoencephalitis public health relevance

项目摘要

DESCRIPTION (provided by applicant): Amebiasis and related vision-threatening infections caused by amoebae is a major contributor to diarrheal diseases, primary amoebic meningoencephalitis and cornea problems. Sequencing of amoeba genomes has evoked a broad search for new, pathogen-specific drug targets, of which the sterol C24-methyltransferase (24-SMT) is a clear candidate since it is synthesized in protozoa but not in animals. Notably, 24-SMT is responsible for the introduction of the methyl group at C24 into the ergosterol side chain. The cholesterol side chain is missing this structural feature which is crucial to ergosterol function. In the R21 phase, we will incubate a series of mechanism-based inhibitors of 24-SMT that differ in the sterol frame and electronics of the side chain with cultured cells of Acanthamoeba and Naegleria and determine which compounds are the most potent inhibitors of ergosterol biosynthesis, trophozoites growth and cyst formation. Additionally, to make improvements to existing drug therapy for treating amoeba infections we will further evaluate in vitro representative anti-fungal azoles that target sterol 14-demethylase (14-SDM; CYP51). Therefore, we will characterize the substrate preference and product outcome of cloned enzymes and use these enzymes to determine inhibitor specificity, binding and covalent inactivation properties. The R33 phase will be undertaken with the proof-of-concept demonstrated. We will test in vitro whether our lead molecules that inhibit 24-SMT in combination with traditional chemotherapeutics have synergistic activities. Additionally, in collaboration with investigators at Meharry Medical College and UTSouthwestern Medical Center, we will evaluate our lead candidate drugs in a mouse model of Acanthamoeba keratitis or primary amebic meningoencephalitis due to Naegleri fowleri. With the aid of a collaborator at Texas Tech University Health Sciences Center, crystal structures of 24-SMT and 14- SDM complexed with relevant inhibitors will be generated to identify binding sites with certainty and should reveal interactions involved with catalysis. ADME/toxicity properties will be evaluated by the bioavailability and metabolism of 3H-inhibitor fed to healthy mice. The overall goal of these studies is to establish mechanism-based inhibitors as a novel class of anti- amoeba agents and to develop synergistic partners of steroidal inhibitors and antifungal agents (medical azole or amphotericin B) that target ergosterol biosynthesis and processing. Specific therapeutic combinations of these compounds could achieve optimal amebacidal effectiveness that thereby, provide for better healthcare.

描述（由申请人提供）：阿米巴病和由阿米巴引起的相关威胁视力的感染是腹泻病、原发性阿米巴脑膜脑炎和角膜问题的主要原因，阿米巴基因组的测序引发了对新的病原体特异性药物靶点的广泛搜索。其中甾醇 C24-甲基转移酶 (24-SMT) 是一个明显的候选者，因为它是在原生动物中合成的，而不是在动物中合成的。值得注意的是，24-SMT 负责将 C24 处的甲基引入麦角甾醇侧链，而胆固醇侧链缺少这一对麦角甾醇功能至关重要的结构特征，我们将在 R21 阶段孵化一系列机制。基于 24-SMT 的抑制剂，其甾醇框架和侧链电子与棘阿米巴和耐格里阿米巴培养细胞不同，并确定哪些化合物最有效此外，为了改进治疗阿米巴感染的现有药物疗法，我们将进一步评估针对甾醇 14-去甲基酶（14-SDM；CYP51）的体外代表性抗真菌唑类药物。我们将表征克隆酶的底物偏好和产物结果，并使用这些酶来确定 R33 相的抑制剂特异性、结合和共价失活特性。我们将在体外测试我们的抑制 24-SMT 的先导分子与传统化疗药物联合使用是否具有协同活性。我们将在德克萨斯理工大学的合作者的帮助下，在棘阿米巴角膜炎或由福氏耐格里阿米巴引起的原发性阿米巴脑膜脑炎小鼠模型中评估我们的主要候选药物。健康科学中心，将生成与相关抑制剂复合的 24-SMT 和 14-SDM 的晶体结构，以确定确定的结合位点，并应揭示与催化相关的相互作用/毒性特性将通过 3H- 的生物利用度和代谢进行评估。这些研究的总体目标是建立基于机制的抑制剂作为一类新型抗阿米巴药物，并开发类固醇抑制剂和抗真菌药物的协同伙伴。（医用唑或两性霉素 B）针对麦角甾醇的生物合成和加工。这些化合物的特定治疗组合可以实现最佳的阿米巴药效果，从而提供更好的医疗保健。