Use of 2-aminopyrrole as an agent with antitumor activity
A novel pyrrole derivative synthesized via a specific reaction process exhibits potent cytotoxic and antitumor activity against chemotherapy-resistant tumor cells, addressing the challenge of drug resistance and enhancing treatment efficacy.
Patent Information
- Authority / Receiving Office
- RU · RU
- Patent Type
- Patents
- Current Assignee / Owner
- FEDERALNOE GOSUDARSTVENNOE BYUDZHETNOE OBRAZOVATELNOE UCHREZHDENIE VYSSHEGO OBRAZOVANIYA PERMSKAYA GOSUDARSTVENNAYA FARMATSEVTICHESKAYA ACAD MINIST ZDRAVOOKHRANENIYA ROSSIJSKOJ FEDERATSII
- Filing Date
- 2025-11-12
- Publication Date
- 2026-07-03
AI Technical Summary
Current chemotherapy drugs face challenges with tumor cells that have developed resistance, necessitating the development of compounds with enhanced cytotoxic and antitumor activity against such resistant tumor cells.
The synthesis of a new pyrrole derivative, 2-amino-5-[2-(naphthalen-2-yl)-2-oxoethylidene]-1-(4-nitrobenzamido)-4-oxo-4,5-dihydro-1H-pyrrole-3-carboxamide, is achieved through a reaction of N-(5-(naphthalen-2-yl)-2-oxofuran-3(2H)-ylidene)-4-nitrobenzohydrazide with cyanoacetamide in the presence of triethylamine, yielding a compound with improved cytotoxic and antitumor properties.
The synthesized compound demonstrates significant cytotoxicity and antitumor activity against chemotherapy-resistant tumor cells, including ES-2 ovarian carcinoma, HCT116 human colon cancer, and 4T1 breast cancer cells, with reduced toxicity compared to its prototype brigatinib, and effectively inhibits tumor growth in vivo.
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Abstract
Description
[0001] The invention relates to the field of organic chemistry, a pyrrole derivative, namely, a new biologically active compound I - 2-amino-5-[2-(naphthalen-2-yl)-2-oxoethylidene]-1-(4-nitrobenzamido)-4-oxo-4,5-dihydro-1H-pyrrole-3-carboxamide of the formula:
[0002]
[0003] which can find application in medicine as a drug with antitumor activity.
[0004] The closest structural analogues to the claimed compound are the compounds 2-amino-1-benzamido-5-[2-(naphthalen-2-yl)-2-oxoethylidene]-4-oxo-4,5-dihydro-1H-pyrrole-3-carboxamide of formula II and 2-amino-5-(3,3-dimethyl-2-oxobutylidene)-4-oxo-1-[2-(phenylamino) benzamido]-4,5-dihydro-1H-pyrrole-3-carboxamide of formula III, which have cytotoxic and antitumor activities against human epithelial tumors [1].
[0005]
[0006] The patented compound has advantages over the above-described analogs, since it exhibits pronounced cytotoxic and antitumor activity against tumor cells that have phenotypic signs of resistance to chemotherapy drugs currently used in the treatment of oncological diseases of the corresponding localizations.
[0007] As a prototype, we took the antitumor drug brigatinib [2], LD 50 which in white nonlinear mice under experimental conditions is 96 mg / kg.
[0008] In the acute toxicity test (LD 50), compound I exhibited toxicity greater than 1800 mg / kg. Thus, the toxicity of compound I is significantly less than that of its biological prototype, brigatinib.
[0009] The technical result of the invention is the synthesis of a new, previously undescribed 2-amino-5-[2-(naphthalen-2-yl)-2-oxoethylidene]-1-(4-nitrobenzamido)-4-oxo-4,5-dihydro-1H-pyrrole-3-carboxamide (compound I), which has cytotoxic and antitumor properties against human solid tumor cells.
[0010] The technical result is achieved by obtaining 2-amino-5-[2-(naphthalen-2-yl)-2-oxoethylidene]-1-(4-nitrobenzamido)-4-oxo-4,5-dihydro-1H-pyrrole-3-carboxamide (I) by reacting N-(5-(naphthalen-2-yl)-2-oxofuran-3(2H)-ylidene)-4-nitrobenzohydrazide (IV) with cyanoacetamide in the presence of the basic catalyst triethylamine in a toluene medium according to the scheme:
[0011] Method for the preparation of 2-amino-5-[2-(naphthalen-2-yl)-2-oxoethylidene]-1-(4-nitrobenzamido)-4-oxo-4,5-dihydro-1H-pyrrole-3-carboxamide (I). A mixture of 1.94 g (0.005 mol) N-(5-(naphthalen-2-yl)-2-oxofuran-3(2H)-ylidene)-4-nitrobenzohydrazide (IV), 0.42 g (0.005 mol) cyanoacetamide and 0.5 g (0.005 mol) triethylamine was boiled in 60 ml of toluene for 40 min. The precipitate that formed upon cooling was filtered off and recrystallized from acetic acid.
[0012] 1.48 g (63%) of crystalline substance (I) with Tm 290-291°C were obtained. IR spectrum, ν / cm -1 , KBr tablets: 3439 (NH), 3394 (NH), 3275 (NH), 1698 (C=O), 1668 (C=O), 1627 (C=O), 1578 (C=C). NMR spectrum 1 H (DMSO-d6), δ, ppm: 6.92 s (1H, CH), 7.06 br s (1H, NH), 7.43 br s (1H, NH), 7.57 - 8.46 m (11H аром ), 8.79 ush. s (1H, NH), 9.28 br. s (1H, NH), 10.40 br. s (1H, NH). NMR spectrum 13C, δ, ppm, DMSO-d6: 88.3, 104.2, 123.5, 123.9, 127.4, 128.1, 128.8, 129.3, 129.8, 130.2, 131.3, 132.5, 134.9, 137.3, 141.7, 149.8, 164.5, 166.1, 167.0, 176.3, 190.8.
[0013] The claimed compound is a light yellow crystalline substance, soluble in DMSO (dimethyl sulfoxide), DMF (dimethylformamide), when heated - in ethyl alcohol, acetic acid, stable during storage.
[0014] The invention is confirmed by the following graphic materials, which show:
[0015] The cell lines ES-2 (human ovarian clear cell adenocarcinoma) and HCT116 (human colon cancer) were chosen as objects for studying the cytotoxic properties of the compounds.
[0016] The cytotoxic and antitumor activity of compound I was assessed on solid tumor cell lines, namely, on 4T1 breast cancer cells [3], as well as on ES-2 ovarian carcinoma cells and HCT116 human colon cancer cells.
[0017] The cytotoxicity of the test compounds was determined fluorimetrically using a SpectraFluor Plus microplate reader (Tecan, Switzerland). The in vitro cytotoxic properties of some of the compounds were studied by spectrophotometry using resazurin (AlamarBlue), taking advantage of the ability of viable cells to convert resazurin into fluorescent resarufin [4]. Thus, the more viable cells remain after exposure to the test compound, the brighter the staining, quantified by the fluorimetric method, will be.
[0018] To study the cytotoxicity of the studied cell lines, they were seeded into wells of a flat-bottomed 96-well culture plate and cultured in RPMI-1640 medium supplemented with L-glutamine and 1% penicillin-streptomycin mixture and 10% fetal bovine serum.
[0019] Cells were incubated for 24-48 hours at 37°C in a humidified atmosphere containing 5% CO2. After reaching 70% cell confluency, the test compound was added to the culture medium at various concentrations. The antitumor drug brigatinib was used as a positive control.
[0020] Compound I was used to culture ES-2 ovarian carcinoma cells with a cytotoxic effect similar to that seen in cells cultured with brigatinib. These results indicate that compound I effectively inhibits tumor cell growth (Fig. 1, Fig. 2).
[0021] The results of the in vitro study of cytotoxic properties of ES-2 ovarian carcinoma cells are shown in Fig. 1.
[0022] Results of in vitro cytotoxic properties study using HCT116 cell culture. Fig. 2.
[0023] The cytotoxic activity of compound I was studied on a panel of tumor cell lines using the colorimetric MTS method. The half-inhibitory concentrations (IC50) of the studied compounds were calculated using the online calculator https: / / www.ic50.tk and are presented in Table 1. According to the obtained IC50 values, compound I exhibited high cytotoxic activity against the tumor cell lines ES-2, HCT116. (for example, against vinblastine, this indicator was > 800), while the resistance index of compound I ranged from 17.5 to 3.5, indicating the absence of cross-resistance of tumor cells to standard chemotherapy drugs (doxorubicin, paclitaxel, vinblastine). These cells were resistant to doxorubicin and sensitive to compound I, paclitaxel, and vinblastine (Table 1).
[0024] Table 1. IC50 values (in µM) for compound I and chemotherapeutic agents.
[0025] cell line Compound I Doxorubicin Paclitaxel Vinblastine ES-2 17,5 ± 0,6 1,8 ± 0,5 [8] 5,4 ± 1[8] 0,0003 ± 0,00004 NCT116 3,5 ± 0,4 0,22 ± 0,004 [7] 0,22 ± 0,01[7] 0,25 ± 0,019 Resistance index 1,2 162,5 [8] > 46 [8] 1,7
[0026] The antitumor activity of compound I was studied in vitro using a syngeneic model. 4T1 breast cancer cells were subcutaneously transplanted into Balb / c mice. After tumor volumes reached 50 cm 3 , groups of animals were randomized into 2 groups (n=5) and intraperitoneally injected with the vehicle (control) or compound I at a dose of 10 mg / kg on days 10, 13, 16, 19, 22, 25, and 28 after transplantation. The tumor volume in each group was measured with a caliper and calculated as length × width × width × 0.5. On the 30th day after inoculation of 4T1 cells, the tumor size in laboratory animals of the control group exceeded the same indicator in animals that received injections of compound I (Fig. 3).
[0027] Fig. 3 - Antitumor activity of compound I in the 4T1 sinograft model of Balb / c mice. Differences are significant at * p < 0.01, ** p < 0.001.
[0028] The linear dimensions of 4T1 tumors in the control and experimental groups were determined using an electronic caliper.
[0029] Antitumor activity is evidenced by significant reductions in tumor size, volume, and weight (Tables 2, 3). No signs of systemic side effects were detected in any animal group over the entire 30-day experimental period.
[0030] Table 2. Linear and weight parameters of 4T1 tumors (control).
[0031] Length, mm Width, mm Height, mm volume, mm3 weight, mg 19 8 5 760 530 12,5 9 9 1012,5 550 9,5 7,8 5 370,5 230 8 8 5 320 180 10,2 6,8 4,6 319,06 160 Average 556,41 330
[0032] Table 3. Linear and weight parameters of 4T1 tumors (I).
[0033] Length, mm Width, mm Height, mm volume, mm3 weight, mg 12 7,2 6,4 571,39 320 9,9 7,1 4,8 337,39 210 9,5 7,2 4,2 287,28 173 8,5 6,3 3,7 198,14 150 Average 348,55 213,25
[0034] Compound I also exhibited antitumor activity against 4T1 sinograft.
[0035] Thus, compound I exhibits both cytotoxic and antitumor activity against aggressive metastatic tumor cells. Compound I also demonstrated high antitumor activity against the 4T1 sinograft. This was confirmed by a significant reduction in tumor size and tumor volume in animals treated with compound I.
[0036] Bibliography
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