Design and synthesis of spiro[pyrrolidine-3,3′-quinoline]-2,2′-dione derivatives as novel antifungal agents targeting chitin synthase

Abstract

A novel spiro [pyrrolidine-3′,3′-quinoline]-2,2′-dione scaffold was constructed using fragments of quinoline and pyrrolidine. Subsequently, two series of derivatives were designed based on this scaffold. The enzyme inhibition experiments revealed that all designed compounds had moderate to good inhibitory activity against chitin synthase (CHS). The inhibitory effects of compounds 5i, 5j, 8i and 8n were approximately equal to that of control drug polyoxin B (PB, IP = 86.4 ± 2.9 %, IC50 = 0.082 ± 0.013 mM) which is a well-established CHS inhibitor. The results from enzyme kinetic parameters assays proved that these compounds act as non-competitive inhibitors of CHS. The sorbitol protection experiments demonstrated the tested compounds disrupted the synthesis of cell wall, which further verified that the target of these compounds is CHS. The experiments of antimicrobial showed that compounds 5b, 5f, 5i, 5j, 8f, 8i, 8m, 8n and 8o had strong antifungal activity against the four tested pathogenic fungi strains frequently emerging in clinical setting, with MIC values ranging from 4 to 32 μg/mL, which were either superior to or comparable with those of PB or fluconazole. Furthermore, these compounds displayed synergistic or additive effects when combined with fluconazole and these active compounds also showed promising activity against fluconazole-resistant and micafungin-resistant fungi variants. The result of antimicrobial experiments indicated that these compounds had minimal activity to tested bacterial strains. This suggests that they had selective antifungal activity. The results of ADME prediction, in conjunction with the cytotoxicity assay results, indicated that these compounds had favorable pharmacokinetic profiles and low toxicity. In addition, molecular docking studies illustrated that the compound had a strong affinity with the CHS, which was consistent with the results of enzymatic assays. These findings indicated that the designed compounds are non-competitive inhibitors of CHS with good selectivity and broad-spectrum antifungal activity, and possess significant antifungal activity against drug-resistant fungi, suggesting their potential as lead compounds for the development of novel drugs against drug-resistant fungi.