Developing Therapeutics against Giardia and Other Anaerobic Protozoa by Targeting Parasite Fatty Acyl-CoA Synthetase (ACS)

 描述（由申请人提供）：脂肪酸（FA）在进入后续代谢途径之前被脂肪酰辅酶A合成酶（ACS）激活。厌氧原生动物贾第虫、内阿米巴和毛滴虫不能重新合成FA。它们从宿主和/或微环境中吸收FA，但具有用于FA活化的ACS和用于有限FA延伸的FA延伸酶（ELO）。我们发现ACS抑制剂triacsin C（triaC）可以抑制贾第虫ACS酶的活性以及体外寄生虫的生长，表明ACS可以作为新的药物靶点。在本项目中，我们将充分探索通过靶向寄生虫ACS酶来开发针对贾第虫和其他厌氧原生动物的新疗法的潜力，并实现以下目标。在R21阶段，我们将：1）确定GiACS酶的生物化学特征并开发高通量筛选（HTS）测定以测试FDA批准的药物; 2）合成triaC的类似物以测试抗GiACS功效和SAR分析; 3）测试triaC和针对来自三种贾第虫聚集体的菌株的最高命中的功效;以及4）通过测试triaC和针对其他厌氧原生动物（包括阿米巴、内阿米巴和毛滴虫）的最高筛选命中的功效来评估ACS是否可以作为广谱药物靶标。 在R33阶段，我们将：1）通过ActiProbe10K文库的HTS发现新的抗GiACS抑制剂; 2）合成更多的triaC类似物和前命中的类似物，用于体外测试对贾第虫和其他原生动物的效力和SAR分析，并通过贾第虫的功能基因组学分析验证药物作用机制; 3）表征来自对ACS抑制敏感的其它原生动物的ACS，用于筛选已知药物和SAR分析;和4）在体外和体内功效前ADME测定中评价最高命中，并在动物模型中评价针对贾第虫的功效。我们最近报道，ACS可以作为一个新的药物靶点在隐孢子虫。我们的初步数据还证实了triaC对贾第虫和阿米巴的疗效，这强烈支持ACS作为原生动物寄生虫的广谱药物靶标。通过利用我们团队在寄生虫生物化学和药物化学方面的专业知识，该项目的成功完成预计将有很大的潜力来确定一些已知的药物和新型化合物对贾第虫和其他厌氧原生动物有效，用于重新利用和/或开发新的治疗方法。

 DESCRIPTION (provided by applicant): Fatty acids (FAs) are activated by fatty acyl-CoA synthetase (ACS) before they enter subsequent metabolic pathways. The anaerobic protozoa Giardia, Entamoeba and Trichomonas are unable to synthesize FAs de novo. They scavenge FAs from the host and/or microenvironment, but possess ACS for FA activation and FA elongase (ELO) for limited FA extension. We discovered that the ACS inhibitor triacsin C (triaC) could inhibit the ACS enzyme activity in Giardia as well as the parasite growth in vitro, indicatin that ACSs could serve as novel drug targets. In this project, we will fully explore the potential t develop novel therapeutics against Giardia and other anaerobic protozoa by targeting parasite ACS enzymes and achieving the following aims. In the R21 phase, we will: 1) determine the biochemical features of GiACS enzymes and develop a high- throughput screening (HTS) assay to test FDA-approved drugs; 2) synthesize analogs of triaC for testing anti-GiACS efficacy and SAR analysis; 3) test the efficacy of triaC and top hits against strains from three Giardia assemblages; and 4) assess whether ACS could serve as a broad-spectrum drug target by testing the efficacy of triaC and top screening hits against other anaerobic protozoa, including Acanthamoeba, Entamoeba and Trichomonas. In the R33 phase, we will: 1) discover novel anti-GiACS inhibitors by HTS of the ActiProbe10K library; 2) synthesize more triaC analogs and analogs of top hits for testing efficacy against Giardia and other protozoa in vitro and SAR analysis, and validate the mechanism of drug action by functional genomics analysis in Giardia; 3) characterize ACSs from other protozoa that are sensitive to the inhibition of ACS for screening known drugs and SAR analysis; and 4) evaluate top hits in in vitro and in vivo pre-efficacy ADME assays and for efficacy against Giardia in an animal model. We have recently reported that ACS could serve as a novel drug target in Cryptosporidium. Our preliminary data also confirmed the efficacy of triaC on Giardia and Acanthamoeba, which strongly supports ACS as a broad-spectrum drug target in protozoan parasites. By taking advantage of our team's expertise in parasite biochemistry and medicinal chemistry, the successful completion of this project is expected with a great potential to identify a number of known drugs and novel compounds efficacious against Giardia and other anaerobic protozoa for repurposing and/or for developing new therapeutics.