2-amino-5-bromo-5-[1-bromo-2-oxo-2-(4-methylphenyl)-ethyl]-4-oxo-4,5-dihydrofuran-3-carboxylic acid nitrile exhibiting antifungal activity against candidaauris

Abstract

FIELD: antifungal medicine.SUBSTANCE: invention relates to a compound - 2-amino-5-bromo-5-[1-bromo-2-oxo-2-(4-methylphenyl)ethyl]-4-oxo-4,5-dihydrofuran-3-carboxylic acid nitrile of the formula:exhibiting antifungal activity.EFFECT: creation of a new compound with antifungal action against Candida auris and low toxicity.1 cl, 1 tbl, 2 ex

Description

[0001] The invention relates to biologically active compounds, namely to 2-amino-5-bromo-5-[1-bromo-2-oxo-2-(4-methylphenyl)ethyl]-4-oxo-4,5-dihydrofuran-3-carboxylic acid nitrile of the formula:

[0002]

[0003] possesses antifungal activity and low toxicity, which suggests the possibility of its use in medicine as a medicinal antifungal drug.

[0004] A structural analogue of the compound is known, obtained by the interaction of 2-amino-4-oxo-5-[2-oxo-2-(4-methylphenyl)ethylidene]-4,5-dihydrofuran-3-carboxylic acid nitrile with 10% excess of bromine in glacial acetic acid [Ivanov, D.V. Use of (5Z)-2-amino-5-bromo-5-(1-bromo-2-oxo-2-phenylethylidene)-4-oxo-1H-4,5-dihydrofuran-3-carboxylic acid nitrile as an agent exhibiting antifungal activity / D.V. Ivanov, N.M. Igidov, V.V. Novikova, R.R. Makhmudov / / Patent No. 2815046 C2 Russian Federation, IPC C07D 307 / 68, A61K 31 / 341, A61P 31 / 04.: No. 2021134026: declared 11 / 22 / 2021: published 03 / 11 / 2024].

[0005] The aim of this invention is to find a compound in the series of 2-amino-5-bromo-5-(2-aryl-1-bromo-2-oxoethyl)-4-oxo-4,5-dihydrofuran-3-carboxylic acid nitriles that has a pronounced antifungal effect against the current species of Candida auris in combination with low toxicity.

[0006] Non-albicans species of yeast micromycetes, previously considered non-pathogenic or minimally virulent, are now one of the main causes of morbidity and mortality in immunocompromised individuals. The frequency of occurrence of each species varies depending on geographic differences in different countries. Thus, according to Singh DP et al.

[14] , 59.9% of isolates isolated in superficial candidiasis belonged to non-albicans species. Similar data indicating cases of overall predominance of Candidanon-albicans representatives in the etiology of candidiasis of various localizations were obtained in studies [KA Al-Sofyani, MS Uddin, HS Alghamdi, D. El-Hossary, Acta Sci. Paediatr., 3, 37-47 (2020); doi: 10.31080 / aspe.2020.03.0318; I. Mareković, S. Pleško, V. Rezo Vranješ et al.,J. Fungi, 7(267) (2021); doi: 10.3390 / jof7040267; TK Ngouana, RM Toghueo, IF Kenfack et al.,J. Mycol.Med., 29(3), 233-238 (2019); doi: 10.1016 / j.mycmed.2019.06.003; E. Seyoum, A. Bitew, A.Mihret, BMC Infect. Dis., 20(231) (2020); doi: 10.1186 / s12879-020-4883-5]. The emergence of new threats in the form of C. auris, first detected in patients in 2009, also requires close attention and motivates the search for new effective molecules. C. auris is a ubiquitous yeast pathogen that can cause invasive candidiasis of the central nervous system, eyes, bones, internal organs, and candidemia. Infections caused by C. auris are a global health problem due to the pathogen's resistance to antifungal drugs such as fluconazole and amphotericin B. These diseases have an overall mortality rate of 30% to 72% [DH Caceres, K. Forsberg, RM Welsh et al., J. Fungi (Basel),5(4), 111 (2019); doi: 10.3390 / jof5040111]. Most C. auris strains are resistant to one or more classes of antifungal drugs, about 30% to two classes, and some strains to three main classes of antifungal drugs [Akter et al., Adv. Med. Psychol.Public Health, 1(4), 255–266 (2024); doi: 10.5281 / zenodo.11095062]. Most cases of mycosis caused by C. auris are nosocomial. Currently, Candida auris is included by WHO experts in the List of Priority Fungal Pathogens to guide public health research, development, and action [I. Mareković, S. Pleško, V. Rezo Vranješ et al., J. Fungi, 7(267) (2021); doi: 10.3390 / jof7040267; TK Ngouana, RM Toghueo, IF Kenfack et al., J. Mycol. Med., 29(3), 233–238 (2019); doi: 10.1016 / j.mycmed.2019.06.003; E. Seyoum, A. Bitew, A. Mihret, BMC Infect. Dis., 20(231) (2020); doi:10.1186 / s12879-020-4883-5].

[0007] Our previously obtained data on the antifungal activity of brominated esters of 2-amino-5-(1-bromo-2-aryl-2-oxoethylidene)-4-oxo-1H-4,5-dihydrofuran-3-carboxylic acids against C. auris (MIC 7.8-15.6 mg / l) [Vasiliev, V.O. Antifungal activity of ethyl esters of 2-amino-5-(1-bromo-2-aryl-2-oxoethylidene)-4-oxo-1H-4,5-dihydrofuran-3-carboxylic acids / V.O. Vasiliev, V.V. Novikova, D.V. Ivanov, N.M. Igidov / / Experimental and clinical pharmacology. - 2025. - Vol. 88, No. 8. - P. 3-5. - DOI 10.30906 / 0869-2092-2025-88-8-3-5. - EDN NNRDWQ] indicate the prospects of searching for new highly active compounds in the series of bromo derivatives of 4,5-dihydrofuran-3-carboxylic acids.

[0008] The stated objective is achieved by obtaining 2-amino-5-bromo-5-[1-bromo-2-oxo-2-(4-methylphenyl)ethyl]-4-oxo-4,5-dihydrofuran-3-carboxylic acid nitrile by reacting 2-amino-4-oxo-5-[2-oxo-2-(4-methylphenyl)ethylidene]-4,5-dihydro-3-furancarboxylic acid nitrile II with bromine upon boiling in a glacial acetic acid medium according to the scheme:

[0009]

[0010] Example 1. Obtaining the claimed compound I.

[0011] A mixture of 0.64 g (0.0025 mol) of 2-amino-4-oxo-5-[2-oxo-2-(4-methylphenyl)ethylidene]-4,5-dihydro-3-furancarboxylic acid nitrile II and 0.48 g of bromine (0.003 mol) in 30 ml of glacial acetic acid was boiled for 5 minutes until the starting compounds dissolved. The precipitate that formed upon cooling was filtered off and recrystallized from alcohol. 0.58 g (56%) of a colorless crystalline substance Ic T пл. 203-205°С. C 14 H 10 Br2N2O3. IR spectrum, ν, cm -1 (crystals): 3323 ( NH ), 3116 ( NH ), 2232 (CN), 1739 (C=O), 1722 (C=O), 1632 (C=C). NMR spectrum, δ, ppm, DMSO-d6: 2.40 s (3H, CH3), 6.37 s (1H, CH), 7.36 d (2H аром , J=7.9), 7.99 d (2H аром , J=7.9), 10.14 ush. s (1H, NH), 10.19 br. s (1H, NH). NMR spectrum 13 C, δ, ppm, DMSO-d6: 21.7, 50.6, 91.5, 112.9, 129.6, 129.8, 130.0, 131.0, 146.1, 175.2, 186.2, 190.4.

[0012] The claimed compound is a colorless crystalline substance, soluble in dimethyl sulfoxide, dimethylformamide, soluble when heated in alcohol, dioxane, insoluble in hexane and water.

[0013] Example 2. Study of biological activity.

[0014] Acute toxicity was studied in experiments on white nonlinear mice of both sexes weighing 20-24 g, kept on a standard vivarium diet. To determine the average lethal dose, the express method of V.B. Prozorovsky was used (Prozorovsky, V.B. Statistical processing of the results of pharmacological studies / V.B. Prozorovsky / / Psychopharmacology and biological narcology. - 2007. - No. 7. - Pp. 2090-2120). The studied compounds were administered orally as a suspension in 1% starch solution in increasing doses, four consecutive doses were used for calculations. Each dose was administered to two animals. The general condition and behavior of the animals, the condition of the fur coat, and body weight were recorded. In each group of animals, the number of deaths was taken into account for the entire observation period. The results were processed statistically with the calculation of the median lethal dose (LD 50 ).

[0015] Antifungal activity was determined by two-fold serial dilutions in Sabouraud's liquid nutrient medium [Guidelines for Conducting Preclinical Studies of Drugs. - Part 1. - Moscow: Grif i K, 2012. - Pp. 576-578.]. Minimum inhibitory concentrations (MICs) were determined for the test compound (I) against the C. auris strain РКПГY 1937 / ОКМ-134 obtained from the Russian Collection of Pathogenic Fungi (St. Petersburg). Inoculations were made in Sabouraud's liquid medium with different concentrations of the test compounds. The test compound (0.05 g) was dissolved in 5 ml of dimethyl sulfoxide, 1 ml of the resulting 1:100 dilution was combined with 4 ml of Sabouraud's liquid medium. Next, a series of serial dilutions of compounds with a twofold decreasing concentration were prepared.

[0016] The cultures were grown in test tubes on slanted agar medium (solid Sabouraud medium). A 48-hour culture was used to determine antifungal activity. To prepare a working suspension of microorganisms, the grown culture was washed with isotonic sodium chloride solution and the density of the microbial suspension was determined to be 0.5 according to McFarland. A working solution with a concentration of 1-5×10 was then prepared from the resulting microbial suspension. 5 CFU / ml. This suspension was added in a quantity of 0.1 ml to wells containing serial dilutions of the compound under study. Thus, the microbial load for determining antifungal activity was 1-2.5×10 5 CFU / ml.

[0017] The results were recorded after 24 hours of incubation of the control and test tubes in a thermostat at 35°C ± 1°C. The minimum inhibitory concentration (MIC) was determined based on the absence of growth signs on the nutrient medium: the last tube with inhibited growth (transparent) corresponds to the MIC of the compound against the given strain.

[0018] The fungistatic effect of the test compound I was compared with the effect of fluconazole substance (JSC Medisorb, batch No. 260519).

[0019] The test results are shown in Table 1.

[0020] Statistical processing of the obtained results was carried out using the one-way analysis of variance method using Microsoft Excel 2010, StatGraphics Plus 5.0 software.

[0021] Table 1

[0022] Antifungal activity and acute toxicity of compound (II)

[0023] Connection Acute toxicity of compounds LD50, mg / kg Antifungal activity MIC, mg / l Candida auris RKPGY 1937 / OKM-134, The claimed compound (I) >2000 3.9-7.8* Fluconazole 1273** 62.5-250.0

[0024] Note:

[0025] * - the excess of the antifungal effect of fluconazole is statistically significant (p≤0.05).

[0026] ** National Library of Medicine: ChemIDplus:[site]. - Bethesda, 1994 -. - URL: https: / / chem.nlm.nih.gov / chemidplus / name / fluconazole - Text: electronic.

[0027] As can be seen from Table 1, the claimed compound I, with lower toxicity, surpasses the comparison drug, fluconazole, in antifungal activity against Candida auris RKPY 1937 / OKM-134 by 16-30 times.

[0028] Therefore, the claimed compound can find application in medicine as an antifungal drug in the treatment of resistant mycoses caused by this type of Candida spp.

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