Preston Sarah, Jabbar Abdul, Nowell Cameron, Joachim Anja, Ruttkowski Bärbel, Baell Jonathan, Cardno Tony, Korhonen Pasi K, Piedrafita David, Ansell Brendan R E, Jex Aaron R, Hofmann Andreas, Gasser Robin B
Faculty of Veterinary and Agricultural Sciences, The University of Melbourne, Parkville, Victoria 3010, Australia.
Medicinal Chemistry, Monash University Institute of Pharmaceutical Sciences (MIPS), Monash University, Parkville, Victoria 3052, Australia.
Int J Parasitol. 2015 Apr;45(5):333-43. doi: 10.1016/j.ijpara.2015.01.007. Epub 2015 Mar 5.
Due to major problems with drug resistance in parasitic nematodes of animals, there is a substantial need and excellent opportunities to develop new anthelmintics via genomic-guided and/or repurposing approaches. In the present study, we established a practical and cost-effective whole-organism assay for the in vitro-screening of compounds for activity against parasitic stages of the nematode Haemonchus contortus (barber's pole worm). The assay is based on the use of exsheathed L3 (xL3) and L4 stages of H. contortus of small ruminants (sheep and goats). Using this assay, we screened a panel of 522 well-curated kinase inhibitors (GlaxoSmithKline, USA; code: PKIS2) for activity against H. contortus by measuring the inhibition of larval motility using an automated image analysis system. We identified two chemicals within the compound classes biphenyl amides and pyrazolo[1,5-α]pyridines, which reproducibly inhibit both xL3 and L4 motility and development, with IC50s of 14-47 μM. Given that these inhibitors were designed as anti-inflammatory drugs for use in humans and fit the Lipinski rule-of-five (including bioavailability), they show promise for hit-to-lead optimisation and repurposing for use against parasitic nematodes. The screening assay established here has significant advantages over conventional methods, particularly in terms of ease of use, throughput, time and cost. Although not yet fully automated, the current assay is readily suited to the screening of hundreds to thousands of compounds for subsequent hit-to-lead optimisation. The current assay is highly adaptable to many parasites of socioeconomic importance, including those causing neglected tropical diseases. This aspect is of major relevance, given the urgent need to deliver the goals of the London Declaration (http://unitingtocombatntds.org/resource/london-declaration) through the rapid and efficient repurposing of compounds in public-private partnerships.
由于动物寄生线虫存在严重的耐药性问题，通过基因组引导和/或重新利用的方法开发新型驱虫药具有迫切需求和绝佳机遇。在本研究中，我们建立了一种实用且经济高效的全生物体检测方法，用于体外筛选对捻转血矛线虫（捻转胃虫）寄生阶段具有活性的化合物。该检测方法基于使用小反刍动物（绵羊和山羊）的捻转血矛线虫脱鞘L3（xL3）和L4阶段。利用此检测方法，我们通过使用自动图像分析系统测量幼虫运动抑制情况，筛选了一组精心挑选的522种激酶抑制剂（美国葛兰素史克公司；代码：PKIS2）对捻转血矛线虫的活性。我们在联苯酰胺和吡唑并[1,5-α]吡啶化合物类别中鉴定出两种化学物质，它们可重复性地抑制xL3和L4的运动及发育，IC50为14 - 47 μM。鉴于这些抑制剂被设计用作人类抗炎药物且符合Lipinski五规则（包括生物利用度），它们在从活性分子到先导化合物的优化以及重新用于对抗寄生线虫方面显示出前景。此处建立的筛选检测方法相较于传统方法具有显著优势，特别是在易用性、通量、时间和成本方面。尽管目前尚未完全自动化，但当前的检测方法很适合筛选数百至数千种化合物以进行后续的从活性分子到先导化合物的优化。当前的检测方法高度适用于许多具有社会经济重要性的寄生虫，包括那些导致被忽视热带病的寄生虫。考虑到迫切需要通过公私伙伴关系中化合物的快速高效重新利用来实现《伦敦宣言》（http://unitingtocombatntds.org/resource/london-declaration）的目标，这一点具有重大意义。
