Ebselen Not Only Inhibits Clostridioides difficile Toxins but Displays Redox-Associated Cellular Killing.
Ebselen Not Only Inhibits Clostridioides difficile Toxins but Displays Redox-Associated Cellular Killing.
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DOI：
10.1128/spectrum.00448-21
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发表时间：
2021-10-31
影响因子：
3.7
通讯作者：
Hurdle JG
中科院分区：
文献类型：
--
作者：
Marreddy RKR;Olaitan AO;May JN;Dong M;Hurdle JG
Ebselen是一种反应性有机硒化合物，通过与TcdA和TcdB毒素的半胱氨酸蛋白酶结构域共价结合而抑制TcdA和TcdB毒素。提示依布硒对难辨梭状芽胞杆菌缺乏抗菌活性。然而，这种看法与艰难梭菌具有可能成为潜在靶标的基本含半胱氨酸酶，以及已报道的依布硒对其他物种的抗菌活性相矛盾。因此，我们重新评估了抗C.依布硒的难治性。药敏试验表明，其活性要么被威尔金斯-查尔格伦琼脂中发现的丙酮酸略微降低，要么被布鲁氏菌琼脂中的血液消除。在脑心灌注琼脂中，除078型固有耐药株(MICMIC32~128MICg/ml)外，Ebselen对大多数艰难梭菌(MIC值为2~8 μg/ml)均有抑制作用。对于艰难梭菌R20291，当浓度低于其最低杀菌浓度16 μg/ml时，依布硒可抑制毒素和孢子的产生。转录组分析表明，ebselen改变了氧化还原相关过程和半胱氨酸代谢，增强了Stickland Pro代谢的表达，可能会从NADH中再生NAD+。在细胞分析中，ebselen诱导半胱氨酸摄取，耗尽非蛋白硫醇，并扰乱NAD+/NADH比率。综上所述，ebselen通过多靶点作用杀死艰难梭菌细胞，其中包括破坏细胞内的氧化还原，这与ebselen是一个反应性分子是一致的。然而，这些抗菌作用在治疗急性艰难梭菌感染(CDI)中的生理学相关性可能会受到宿主因素的破坏，例如血液，这些宿主因素保护艰难梭菌不被ebselen杀死。重要性我们证明ebselen通过破坏致病艰难梭菌的氧化还原动态平衡，改变NAD+和NADH的正常浓度来杀死致病艰难梭菌，而NAD+和NADH是细胞中各种代谢功能的关键。然而，这种抗菌作用受到宿主成分，即血液的阻碍。未来发现的ebselen类似物或机械上类似的化合物，在血液中保持活性，可能是CDI的药物线索或研究艰难梭菌体内氧化还原生物学的探针。
Ebselen, a reactive organoselenium compound, was shown to inhibit toxins TcdA and TcdB by covalently binding to their cysteine protease domains. It was suggested that ebselen lacked antimicrobial activity against Clostridioides difficile. However, this perception conflicts with C. difficile having essential cysteine-containing enzymes that could be potential targets and the reported antimicrobial activity of ebselen against other species. Hence, we reevaluated the anti-C. difficile properties of ebselen. Susceptibility testing revealed that its activity was either slightly reduced by pyruvate found in Wilkins-Chalgren agar or obliterated by blood in brucella agar. In brain heart infusion (BHI) agar, ebselen inhibited most C. difficile strains (MICs of 2 to 8 μg/ml), except for ribotype 078 that was intrinsically resistant (MIC = 32 to 128 μg/ml). Against C. difficile R20291, at concentrations below its minimal bactericidal concentration (MBC), 16 μg/ml, ebselen inhibited production of toxins and spores. Transcriptome analysis revealed that ebselen altered redox-associated processes and cysteine metabolism and enhanced expression of Stickland proline metabolism, likely to regenerate NAD+ from NADH. In cellular assays, ebselen induced uptake of cysteine, depleted nonprotein thiols, and disrupted the NAD+/NADH ratio. Taken together, killing of C. difficile cells by ebselen occurs by a multitarget action that includes disrupting intracellular redox, which is consistent with ebselen being a reactive molecule. However, the physiological relevance of these antimicrobial actions in treating acute C. difficile infection (CDI) is likely to be undermined by host factors, such as blood, which protect C. difficile from killing by ebselen. IMPORTANCE We show that ebselen kills pathogenic C. difficile by disrupting its redox homeostasis, changing the normal concentrations of NAD+ and NADH, which are critical for various metabolic functions in cells. However, this antimicrobial action is hampered by host components, namely, blood. Future discovery of ebselen analogues, or mechanistically similar compounds, that remain active in blood could be drug leads for CDI or probes to study C. difficile redox biology in vivo.