Design and search for prospective diuretics (CA II Inhibitors) among aroylhydrazones of esters quinone oxime using in silico and in vivo methodology

The design and search for new selective inhibitors of CA II with a better pharmacological profile, which would cause minimal electrolyte disturbances in the body, remains an urgent problem of medical chemistry and pharmacology today. It is important that the discovered new classes of inhibitors do not always contain the main “pharmacophoric” function (sulfamide), which is characteristic of “classic” drugs (Acetazolamide, Methazolamide, Ethoxzolamide, Dorzolamide and others), but are derivatives of phenols, polyamines, coumarins/thiocoumarins, ureas, thioureas, hydroxamates, etc. These molecules also bind in the active site of the enzyme, but do not interact directly with the catalytic zinc ion or interact through zinc-coordinated water molecules/hydroxide ion. However, this leads to an increase in their selectivity and, as a result, pharmacological action. Continuing the search for compounds that affect urination, we were interested in aroylhydrazones of esters of quinone oxime. Firstly, they are characterized by certain structural features (dynamic and geometric isomerism); secondly, they exhibit redox properties; thirdly, the presence of aromatic fragments makes it possible to create a voluminous combinatorial library for analysis. These compounds are ligands in complexation reactions, and an additional increase in the number of hydrogen acceptors in the molecule due to structural modification will improve ligand-enzymatic interactions with carbonic anhydrase (CAII) and, as a result, reveal new promising diuretics. The aim – design and search for potential diuretics (CA II inhibitors) among aroylhydrazones of esters of quinone oxime using in silico, traditional synthesis and in vivo methodologies. Methods of organic synthesis, physico-chemical methods of analysis of organic compounds (NMR 1H-spectroscopy, elemental analysis). Prediction of affinity to the biological target, prediction of toxicity and lipophilicity of the combinatorial library of benzohydrazides O-aroyl esters of quinone oxime using computer services. The study of compounds affecting the excretory function of rat kidneys was carried out according to the generally accepted method of E.B.Berkhin with water load. The investigation of the probable mechanism was carried out using flexible molecular docking, as an approach to search for molecules that have affinity for human carbonic anhydrase type II (CA II). Macromolecular data of the crystal structure of CA II (PDB ID – 3HS4) were downloaded from the Protein Data Bank (PDB). The design was developed and the search for diuretic agents among benzohydrazides of O-aroyl esters of quinone oximes was developed using in silico methods (prediction of affinity, lipophilicity, toxicity and enzyme-ligand interactions), traditional organic synthesis, and in vivo methods (effect on excretory function of rat kidneys). The synthesis of benzohydrazides of O-aroyl esters of quinone oxime was carried out by the interaction of aroylhydrazines with 4-[(aroylimino)]cyclohexa-2,5-dien-1-ones. The structure of the synthesized compounds was confirmed by elemental analysis and 1H NMR spectra. Studies of the effect of synthesized compounds on the excretory function of rat kidneys allowed us to identify a number of promising compounds among aroylhydrazones of quinonexime esters, which increase daily diuresis by 54.2-352.8% compared to the control group. At the same time, it was established that the most active was N'-(4-[(2-chlorobenzoyloxy)imino]cyclohexa-2,5-dien-1-ylidene)-3-nitrobenzohydrazide, which increased daily diuresis by 352.8% in comparison with the control group, while exceeding the effect of “Hydrochlorothiazide” (170.8%). The developed and implemented strategy for the search for diuretics among benzohydrazides of O-aroylesters of quinone oxime allowed the identification of an effective compound, which in terms of diuretic effect exceeds the comparison drug “Hydrochlorothiazide”. Visualization of the molecular docking of the active compounds showed that their geometry makes it difficult to place them in the pocket of the active site of CA II, but the pronounced diuretic effect can also be associated with their ability to form coordination bonds with the zinc cation. The obtained results justify the further targeted search for potential diuretics among this class of compounds for a more detailed understanding and study of the mechanism of action

N-arylsulfonyl-2-aroylamino-1,4-quinone imines and their hydrogenated analogues: prediction of toxicity and prospects for use as diuretics

Sokolova K.V., Podpletnia O.A., Konovalova S.O., Avdieienko A.P., Komarovska-Porokhniavets O.Z., Lubenets V.I., Kovalenko S.I. Continuing our research on compounds that affect urination, we have become interested in N-arylsulfonyl-2-aroylamino-1,4-quinone imines, which combine a quinone matrix with tolylsulfonamide and benzamide fragments with versatile biological activity in their structure, which has a promising value in preventing development of pathological processes in kidneys. Therefore, the search for low-toxic compounds with polyvector activity as a promising approach to the design of drug-like molecules has become an urgent aspect in this regard. The aim of this work was to investigate N-arylsulfonyl-2-aroylamino-1,4-quinone imines and their hydrogenated analogues as promising diuretic agents with antiradical and antibacterial activity using in silico, in vitro and in vivo methodologies. The virtual laboratory of the ProTox-II site is used to predict the toxicity of molecules. The study of compounds affecting the excretory function of the rat kidneys was carried out on 120 white Wistar rats according to the method of E.B. Berkhin under conditions of water stress and spontaneous urination. The interaction of the synthesised compounds with 2,2-diphenyl-1-picrylhydrazyl (DPPH) was used to study their antiradical activity in vitro. The antibacterial activity of the compounds was studied on test cultures of the bacteria Escherichia coli, Staphylococcus aureus, Mycobacterium luteum and the fungi Candida tenuis, Aspergillus niger by the method of serial dilutions in a liquid nutrient medium. Based on the results of the calculation, it was predicted that N-arylsulfonyl-2-aroylamino-1,4-quinone imines (2) and their hydrogenated analogues (3) have hepato-(immuno-, cyto-) toxicity, carcinogenicity (mutagenicity) similar to natural quinones and diuretics (toxicity class IV). This class of compounds has been shown to have both stimulatory and inhibitory effects on diuresis under condi­tions of water stress and spontaneous urination. At the same time, N-(5-methyl-6-oxo-3-(tosylimino)cyclohexa-1,4-dien-1-yl)benzamide (2.3) was revealed to increase daily diuresis by 67.1% compared with the control, exceeding the effect of «Furosemide» (22.2%). It was found that quinone imines (2.1-2.5) inhibited the formation of the DPPH radical by 25.99-40.09%, while their hydrogenated analogues (3.1 and 3.2) – by 61.56% and 68.28%, respectively, and are more effective acceptors of radicals. The microbiological screening revealed a number of promising compounds that inhibited the growth of S. aureus (compound 2.5, MIC 62.5 μg/ml, MBC 125.0 μg/ml), M. luteum (3.1 and 3.2, MIC 31.2 μg/ml, MBC 62.5 μg/ml) and A. niger (2.1, 2.4 and 3.2, MIC 31.2 μg/ml, MPC 62.5 μg/ml). According to the results of biological studies, among N-arylsulfonyl-2-aroylamino-1,4-quinone imines and their hydrogenated analogues, compound 2.3 has been iden­tified, which competes with «Furosemide» in potency and has high antibacterial activity against S. aureus. Other compounds show moderate antiradical activity, high antibacterial activity against M. luteum (2.1, 3.1) and antifungal activity against A. niger (2.1, 2.4, 3.2). The obtained results support the further research for diuretics with polyvector activity within this class of compounds

N-arylsulfonyl-2-aroylamino-1,4- quinonе imines and their hydrogenated analogues: prediction of toxicity and prospects for use as diuretics

Continuing our research on compounds that affect urination, we have become interested in N-arylsulfonyl-2-aroylamino-1,4-quinone imines, which combine a quinone matrix with tolylsulfonamide and benzamide fragments with versatile biological activity in their structure, which has a promising value in preventing development of pathological processes in kidneys. Therefore, the search for low-toxic compounds with polyvector activity as a promising approach to the design of drug-like molecules has become an urgent aspect in this regard. The aim of this work was to investigate N-arylsulfonyl-2-aroylamino-1,4-quinone imines and their hydrogenated analogues as promising diuretic agents with antiradical and antibacterial activity using in silico, in vitro and in vivo methodologies. The virtual laboratory of the ProTox-II site is used to predict the toxicity of molecules. The study of compounds affecting the excretory function of the rat kidneys was carried out on 120 white Wistar rats according to the method of E.B. Berkhin under conditions of water stress and spontaneous urination. The interaction of the synthesised compounds with 2,2-diphenyl-1- picrylhydrazyl (DPPH) was used to study their antiradical activity in vitro. The antibacterial activity of the compounds was studied on test cultures of the bacteria Escherichia coli, Staphylococcus aureus, Mycobacterium luteum and the fungi Candida tenuis, Aspergillus niger by the method of serial dilutions in a liquid nutrient medium. Based on the results of the calculation, it was predicted that N-arylsulfonyl-2-aroylamino-1,4-quinone imines (2) and their hydrogenated analogues (3) have hepato-(immuno-, cyto-) toxicity, carcinogenicity (mutagenicity) similar to natural quinones and diuretics (toxicity class IV). This class of compounds has been shown to have both stimulatory and inhibitory effects on diuresis under conditions of water stress and spontaneous urination. At the same time, N-(5-methyl-6-oxo-3-(tosylimino)cyclohexa-1,4-dien-1-yl)benzamide (2.3) was revealed to increase daily diuresis by 67.1% compared with the control, exceeding the effect of «Furosemide» (22.2%). It was found that quinone imines (2.1-2.5) inhibited the formation of the DPPH radical by 25.99-40.09%, while their hydrogenated analogues (3.1 and 3.2) – by 61.56% and 68.28%, respectively, and are more effective acceptors of radicals. The microbiological screening revealed a number of promising compounds that inhibited the growth of S. aureus (compound 2.5, MIC 62.5 μg/ml, MBC 125.0 μg/ml), M. luteum (3.1 and 3.2, MIC 31.2 μg/ml, MBC 62.5 μg/ml) and A. niger (2.1, 2.4 and 3.2, MIC 31.2 μg/ml, MPC 62.5 μg/ml). According to the results of biological studies, among N-arylsulfonyl-2-aroylamino-1,4- quinone imines and their hydrogenated analogues, compound 2.3 has been identified, which competes with «Furosemide» in potency and has high antibacterial activity against S. aureus. Other compounds show moderate antiradical activity, high antibacterial activity against M. luteum (2.1, 3.1) and antifungal activity against A. niger (2.1, 2.4, 3.2). The obtained results support the further research for diuretics with polyvector activity within this class of compounds