Design And Synthesis Novel Quinolin-2(1H)-one Derivatives As Potential Chitin Synthase Inhibitors

Fungal infection seriously threatens human health and safety.The limited clinical efficacy of currently used antifungal drugs is mainly due to limitations in pharmacokinetics and pharmacodynamics.Chitin is synthesized from substrate UDP-GlcNAc under the catalysis of chitin synthase.It is an indispensable component of the fungal cell wall.If the biosynthesis of chitin is blocked,the fungal cell wall will not be synthesized normally and this can lead the fungal cells die.Chitin-related substances do not exist in mammalian cells,so chitin synthase can be an ideal target for design of antifungal drugs.Quinolones are an important class of heterocyclic compounds and part of the quinoline alkaloid family.It is a class of compounds with a wide range of biological activities.It has significant biological activities in antiviral,antibacterial,anti-inflammatory,anti-diabetic and anti-cancer.3-Amino-3,4-dihydroquinolin-2（1H）-one has attracted wide attention because of their excellent activity.Therefore,introducing active fragments into drug molecules could be improve the metabolic stability,biocompatibility,selectivity and affinity for the enzyme.Based on the current research progress of chitin synthase,a series of 3-amino-3,4-dihydroquinoline-2（1H）-one compounds were designed and synthesized,and their antibacterial activity in vitro and inhibitory activity against chitin synthase were tested.The main work is summarized as follows:1.Preparation of 3-amino-3,4-dihydroquinolin-2（1H）-one.o-Nitrotoluene reacted with NBS in carbon tetrachlorideto produce 1-（bromomethyl）-2-nitrobenzene.The product combined with diethyl acetylaminomalonate under strong alkaline condition to afford diethyl 2-（2-nitrobenzyl）-2-acetamidomalonate which then was decarboxylation in acidic solution at 100 ^oC to produce 2-amino-3-（2-nitrophenyl）propanoic acid hydrochloride which was esterified with methanol at presence of sulfoxide chloride.The obtained methyl 2-amino-3-（2-nitrophenyl）propanoate hydrochloride was reduced by iron powder to offer 3-amino-3,4-dihydroquinolin-2（1H）-one（compound 6）.2.Preparation of quinoline-2（1H）-one piperazine derivatives.Compound 6 was used as the raw material which reacted with chloroacetyl chloride,and the obtained compound reacted with piperazine hydrochloride.Then the obtained product combined with a series of aromatic carboxylic acid to obtain the target compounds 9a-s.3.Preparation of quinoline-2（1H）-oneα,β-unsaturated carbonyl derivatives.Aromatic amine as a starting material combined with maleic anhydride.The obtained compound further reacted with compound 6 to obtain the target compounds 12a-s.4.Discussed the synthesis conditions of compounds.5.There are 97 synthesized compounds in this thesis,including 52 unreported compounds and 50 target compounds.The structures of the compounds were confirmed by modern spectroscopy such as ¹H NMR,¹³C NMR,LC-MS or HRMS.6.All target compounds have been evaluated antifungal and antibacterial activity in vitro.Antifungal experiments in vitro suggested that the quinoline-2（1H）-one piperazine compounds had better antifungal activity against Candida albicans（ATCC90023）and Candida albicans（ATCC 76615）than against other selected strains.To Candida albicans（ATCC 90023）,the compound 9e with MIC value of 8μg/mL showed more excellent potency when comparable with polyoxin B.Compound 9g showed excellent potency against Candida albicans（ATCC 76615）,which was 4-fold more potency than that of polyoxin B（MIC=16μg/mL）.The antifungal activity of quinolin-2（1H）-oneα,β-unsaturated carbonyl compounds against Candida albicans（ATCC 90023）and Candida albicans（ATCC 76615）was superior to against other strains.Compounds 12b,12d,12e and 12l showed the equal potency of fluconazole against Candida albicans（ATCC 90023）,which was more potency than polyoxin B.Compounds 12a and 12d exhibited good potency against Candida albicans（ATCC76615）,which was 2-fold more potency than that of polyoxin B.Compared with 12a-s,all of the control compounds exhibited weaker antifungal potency.7.Compounds with excellent antifungal activity were selected for combination drug experiments using checkerboard method.Test results showed that the combined use of drugs can improve antifungal activity.Among quinoline-2（1H）-one piperazine compounds,the combinations 9c-fluconazole,9e-fluconazole,9g-fluconazole and9p-fluconazole displayed synergistic or additive effect for selected strains whose FIC values ranged from 0.5-2.0.The combination 9p-fluconazole exhibited strongest synergistic effect against Candida albicans（ATCC 90023）with FIC value of 0.5.The combinations 9c-polyoxin B,9e-polyoxin B,9g-polyoxin B and 9p-polyoxin B displayed moderate to excellent additive effect to selected strains whose FIC values ranged from 1.0-2.0.Among quinoline-2（1H）-oneα,β-unsaturated carbonyl compounds,the representative compounds 12b,12d,12e and 12s were chosen to combination use with fluconazole and polyoxin B respectively.The combinations12e-fluconazole and 12e-polyoxin B showed strong synergistic effects against the selected strains.The combination 12e-fluconazole showed especially excellent synergistic effect against Candida albicans（ATCC 90023）with FIC value 0.4.8.The inhibition percentages（IP）of all compounds against chitin synthase were screened.The test results showed that all synthesized compounds exhibited chitin synthase inhibitory potency.There are nine compounds with inhibitory rate higher than50%in the first series compounds.Most of the quinoline-2（1H）-oneα,β-unsaturated carbonyl compounds showed excellent inhibitory activity on chitin synthase.12b,12d,and 12s had good activity especially,and their IC₅₀ values were 0.09 mM,0.16 mM,and0.19 mM,respectively.9.The kinetic parameters of the enzymatic reaction were determined.