Discovery of Novel α,β-Unsaturated Amide Derivatives as Candidate Antifungals to Overcome Fungal Resistance

In our previous study, coumarin-containing CYP51 inhibitor A32 demonstrated potent antiresistance activity. However, compound A32 demonstrated unsatisfied metabolic stability, necessitating modifications to overcome these limitations. In this study, α,β-unsaturated amides were used to replace the unstable coumarin ring, which increased metabolic stability by four times while maintaining antifungal activity, including activity against resistant strains. Subsequently, the sterol composition analysis and morphological observation experiments indicated that the target of these novel compounds is lanosterol 14α-demethylase (CYP51). Meanwhile, biofilm growth was inhibited and resistance genes (ERG11, CDR1, CDR2, and MDR1) expression was downregulated to find out how the antiresistance works. Importantly, compound C07 demonstrated the capacity to stimulate reactive oxygen species, thus displaying potent fungicidal activity. Moreover, C07 exhibited encouraging effectiveness in vivo following intraperitoneal administration. Additionally, the most potent compound C07 showed satisfactory pharmacokinetic properties and low toxicity. These α,β-unsaturated amide derivatives, particularly C07, are potential candidates for treating azole-resistant candidiasis.在我们此前的研究中，含香豆素（coumarin）的CYP51抑制剂A32展现出强效的抗耐药活性。然而，化合物A32的代谢稳定性欠佳，亟需对其进行结构修饰以克服该类缺陷。本研究中，我们采用α,β-不饱和酰胺取代不稳定的香豆素环，使代谢稳定性提升四倍，同时保留了抗真菌活性，包括针对耐药菌株的活性。随后，甾醇组成分析与形态学观察实验结果表明，此类新型化合物的作用靶点为羊毛甾醇14α-去甲基酶（CYP51）。与此同时，为阐明该类化合物的抗耐药机制，我们发现其可抑制生物膜生成，并下调耐药基因（ERG11、CDR1、CDR2及MDR1）的表达。尤为关键的是，化合物C07能够诱导活性氧（reactive oxygen species）的产生，因此展现出强效的杀真菌活性。此外，经腹腔给药后，C07在体内展现出令人满意的抗真菌效果。除此之外，活性最强的化合物C07还具备良好的药代动力学特性，且毒性较低。此类α,β-不饱和酰胺衍生物，尤其是C07，可作为治疗唑类耐药念珠菌病的潜在候选药物。