The Substitutions L50F, E166A, and L167F in SARS-CoV-2 3CLpro Are Selected by a Protease Inhibitor In Vitro and Confer Resistance To Nirmatrelvir.
The Substitutions L50F, E166A, and L167F in SARS-CoV-2 3CLpro Are Selected by a Protease Inhibitor In Vitro and Confer Resistance To Nirmatrelvir.
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作者：
SARS-CoV-2主要蛋白酶（3CLpro）在病毒生命周期中具有不可或缺的作用，是治疗COVID-19的治疗靶点。需要确定3CL前体抑制剂选择耐药变体的潜力。因此，SARS-CoV-2在增加浓度的ALG-097161存在下体外传代，ALG-097161是在3CLpro药物发现计划的背景下设计的探针化合物。我们鉴定了3CLpro（L50 F E166 A L167 F）中的氨基酸取代组合，其与ALG-097161、Nirmatrelvir（PF-07321332）、PF-00835231和恩西曲韦的50%有效浓度（EC 50）值增加>20倍相关。虽然两个单一的取代（E166 A和L167 F）提供了低水平的抗性抑制剂在生化试验中，三重突变体的结果在最高水平的抗性（6×至72×）。所有取代都与酶促3CLpro活性的显著损失相关，表明病毒适应性降低。结构生物学分析表明，不同的取代减少了抑制剂/酶相互作用的数量，同时保持底物的结合。这些观察结果对于解释临床环境中对3CLpro抑制剂的耐药性发展非常重要。
The SARS-CoV-2 main protease (3CLpro) has an indispensable role in the viral life cycle and is a therapeutic target for the treatment of COVID-19. The potential of 3CLpro-inhibitors to select for drug-resistant variants needs to be established. Therefore, SARS-CoV-2 was passaged in vitro in the presence of increasing concentrations of ALG-097161, a probe compound designed in the context of a 3CLpro drug discovery program. We identified a combination of amino acid substitutions in 3CLpro (L50F E166A L167F) that is associated with a >20× increase in 50% effective concentration (EC50) values for ALG-097161, nirmatrelvir (PF-07321332), PF-00835231, and ensitrelvir. While two of the single substitutions (E166A and L167F) provide low-level resistance to the inhibitors in a biochemical assay, the triple mutant results in the highest levels of resistance (6× to 72×). All substitutions are associated with a significant loss of enzymatic 3CLpro activity, suggesting a reduction in viral fitness. Structural biology analysis indicates that the different substitutions reduce the number of inhibitor/enzyme interactions while the binding of the substrate is maintained. These observations will be important for the interpretation of resistance development to 3CLpro inhibitors in the clinical setting.
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DOI：
10.1074/jbc.m115.651463
发表时间：
2015-08-07
期刊：
The Journal of biological chemistry
影响因子：
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作者：
Tomar S;Johnston ML;St John SE;Osswald HL;Nyalapatla PR;Paul LN;Ghosh AK;Denison MR;Mesecar AD
通讯作者：
Mesecar AD
影响因子：
4.9
作者：
通讯作者：
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影响因子：
7.7
作者：
Flynn, Julia M.;Samant, Neha;Schneider-Nachum, Gily;Barkan, David T.;Yilmaz, Nese Kurt;Schiffer, Celia A.;Moquin, Stephanie A.;Dovala, Dustin;Bolon, Daniel N. A.
通讯作者：
Bolon, Daniel N. A.
影响因子：
16.6
作者：
Abdelnabi R;Foo CS;Jochmans D;Vangeel L;De Jonghe S;Augustijns P;Mols R;Weynand B;Wattanakul T;Hoglund RM;Tarning J;Mowbray CE;Sjö P;Escudié F;Scandale I;Chatelain E;Neyts J
通讯作者：
Neyts J
影响因子：
7.6
作者：
通讯作者：
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