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

 描述(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、内阿米巴和滴虫不能从头合成FA。它们从宿主和/或微环境中清除FA，但具有激活FA的ACS和有限FA延伸的FA延伸酶(ELO)。我们发现ACS抑制剂Triacsin C(TriaC)在体外能抑制贾第虫体内的ACS酶活性和寄生虫的生长，表明ACSS可以作为新的药物靶点。在这个项目中，我们将充分开发针对贾第虫和其他厌氧原虫的新疗法的潜力，通过靶向寄生虫的ACS酶来实现以下目标。在R21阶段，我们将：1)确定GiACS酶的生化特性，并建立高通量筛选(HTS)方法来测试FDA批准的药物；2)合成TriaC的类似物，用于测试抗GiACS的有效性和SAR分析；3)测试TriaC和TOP HITS对来自三个贾第虫组合的菌株的有效性；以及4)通过测试TRIAC和TOP HITS对其他厌氧原虫，包括棘阿米巴、内阿米巴和毛滴虫的疗效，评估ACS是否可以作为广谱药物靶点。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类似物和顶击类似物，用于体外和SAR分析，并通过贾第虫功能基因组学分析验证其药物作用机制；3)从其他对ACS抑制敏感的原虫中鉴定ACSS，用于筛选已知药物和进行SAR分析；以及4)评估TOP HITS在体外和体内的预效ADME试验以及在动物模型中对贾第鞭毛虫的有效性。我们最近报道，ACS可能成为隐孢子虫的一个新的药物靶点。我们的初步数据也证实了TriaC对贾第虫和棘阿米巴的疗效，这有力地支持了ACS作为原虫寄生虫的广谱药物靶点。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，该项目的成功完成有望确定一些已知的药物和新化合物，这些药物和新化合物对贾第虫和其他厌氧原生动物有效，用于重新调整用途和/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.