Synthesis, screening and validation of cysteine-reactive fragments as chikungunya virus protease inhibitors

IF 2.5 4区 医学 Q3 CHEMISTRY, MEDICINAL
Junming He , Henry Illingworth , Sven Ullrich , Pritha Ghosh , Jennifer Ton , Colin J. Jackson , Christoph Nitsche
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Abstract

Alphaviruses like the Chikungunya virus cause severe outbreaks; however, no specific treatments are available. Their viral replication depends on the nsP2 cysteine protease, a promising but underexplored target for drug discovery. In this study, we report a covalent fragment screening against Chikungunya virus nsP2 protease, resulting in the identification of three inhibitors that can serve as starting points for future drug development. Careful validation proved indispensable in eliminating false-positive hits from a Förster resonance energy transfer (FRET)-based inhibition assay, wherein interference was caused by the inner filter effect between the fluorescent substrate and coloured compounds. Jump-dilution experiments accompanied by reactivity studies with cysteine and the recombinant protein indicate covalent inhibition via thia-Michael addition. We further demonstrate cross-inhibition of the related alphavirus nsP2 protease from Sindbis virus. The study provides early insights into nsP2 inhibition by electrophilic fragments featuring non-promiscuous N-arylacrylamides, thus advancing the search for antivirals targeting Chikungunya and other alphaviruses of concern.

Abstract Image

基孔肯雅病毒蛋白酶抑制剂半胱氨酸反应片段的合成、筛选和验证。
甲病毒如基孔肯雅病毒引起严重疫情;然而,目前还没有具体的治疗方法。它们的病毒复制依赖于nsP2半胱氨酸蛋白酶,这是一个很有希望但尚未被开发的药物发现靶点。在这项研究中,我们报道了针对基孔肯雅病毒nsP2蛋白酶的共价片段筛选,从而鉴定出三种抑制剂,可以作为未来药物开发的起点。仔细的验证证明,在消除假阳性击中从Förster共振能量转移(FRET)为基础的抑制试验是必不可少的,其中干扰是由荧光底物和有色化合物之间的内部过滤效应引起的。跳跃稀释实验以及与半胱氨酸和重组蛋白的反应性研究表明,通过添加thia-Michael可以抑制共价。我们进一步证实了与Sindbis病毒相关的甲病毒nsP2蛋白酶的交叉抑制作用。该研究提供了非混杂n-芳基丙烯酰胺亲电片段对nsP2抑制的早期见解,从而推进了针对基孔肯雅病毒和其他令人关注的甲型病毒的抗病毒药物的研究。
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来源期刊
CiteScore
5.70
自引率
3.70%
发文量
463
审稿时长
27 days
期刊介绍: Bioorganic & Medicinal Chemistry Letters presents preliminary experimental or theoretical research results of outstanding significance and timeliness on all aspects of science at the interface of chemistry and biology and on major advances in drug design and development. The journal publishes articles in the form of communications reporting experimental or theoretical results of special interest, and strives to provide maximum dissemination to a large, international audience.
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