Perylenylethynyl phenols: novel antiviral photosensitizers

Abstract

The invention relates to the field of biochemistry, the chemistry of bioactive and photoactive compounds and medicine, and more particularly to a multi-stage method for synthesizing individual photosensitizers which exhibit antiviral activity and are potentially applicable as a medicinal preparation in medical practice.

Description

[0001] Perylenylethynylphenols: Novel antiviral photosensitizers

[0002] 1. Description of the invention

[0003] 1.1 Field of technology to which the invention relates

[0004] The invention relates to the field of chemical technology, the chemistry of biologically and photoactive compounds and medicine, namely to a multi-stage method for obtaining biologically active compounds that exhibit antiviral activity and are potentially applicable as a medicinal product in medical practice.

[0005] 1.2 Prior Art

[0006] The Coronaviridae family includes many medically significant viral pathogens, as demonstrated by the recent outbreak of SARS-CoV-2, the causative agent of the COVID-19 pandemic, which has resulted in over 6 million deaths to date. Treatment of COVID-19 disease remains challenging, making the development of new effective drugs against acute coronavirus infections an important priority for medical research.

[0007] The versatile and light-dependent antiviral activity of the perylene-type compounds hypocrellin and hypericin has been known for several decades. Other major broad-spectrum membrane-targeted perylene compounds are the peryleneethynyl derivatives. Their mechanism was originally thought to primarily involve inhibition of biophysical fusion, but more recent data have emphasized the significant role of photosensitization in their mechanism of action. These compounds are effective photogenerators of singlet oxygen ( 1 O2), which causes significant damage to the unsaturated components of the lipid bilayer, especially when the photosensitizer and the double bonds are in close proximity (the so-called contact-dependent pathway). This effect is based on the ene oxidation reaction of lipids with subsequent cleavage of the C-C bond. 1O2-induced formation of short polar lipids dramatically changes membrane rheology and, in the case of virion membranes, their ability to fuse with the host cell membrane. As a rule, compounds generating 1 Membrane-targeted O2 exhibit remarkable antiviral effects in vitro, such as arylidene rhodanines (LJ compounds), alkyl derivatives of rose bengal, etc. Perylene and its derivatives are recognized as photogenerators of singlet oxygen. We recently determined the quantum yields of generation 1O2 in methanol for several heteroarylethynylperylenes and thienylperylenes that have shown activity against SARS-CoV-2. Non-nucleoside perylene antivirals are amphipathic compounds that consist of a lipophilic perylene moiety and polar functional groups. They are partially soluble in aqueous buffers containing a few percent DMSO, probably forming micelles. The antiviral activity of these compounds is influenced by various factors that affect not only their ability to generate singlet oxygen 1 O2, but also on the ability to penetrate lipid membranes.

[0008] The structure, method of determining the structure, method of isolation, gross formula, complete elemental formula, and biological activity of the claimed compounds are not described in the patent and scientific and technical literature.

[0009] 1.3 Essence of the invention

[0010] The essence of the invention lies in the synthesis of new photosensitizers based on perylene (Fig. 1), the establishment of their structural, physicochemical properties and antiviral activity.

[0011] Fig. 1.

[0012] The synthesis of biologically active compounds in individual form is multi-stage. It can be divided into three main stages:

[0013] 1. Synthesis of the initial 2- and 3-ethynylperylene, previously described in the literature;

[0014] 2. Synthesis, purification and characterization of 3-substituted perylenes;

[0015] 3. Synthesis, purification and characterization of 2-substituted perylenes.

[0016] The starting compounds are synthesized in strict accordance with the methods described in the literature, with subsequent comparison of the NMR spectra of the obtained compounds with the literature data. The syntheses of the starting 2- and 3-ethynylperylene, previously described in the literature, are similar in essence. The synthesis of the derivatives of 3 -substituted ethynylperylenes was carried out by introducing the starting compound - 3-ethynylperylene - into the Sonogashira reaction with 4-iodophenol, 3-iodophenol and 2-bromo-4-iodophenol, as well as copper iodide and dichlorobis(triphenylphosphine)palladium catalysts, in dimethylformamide with the addition of triethylamine. The reaction proceeded for 24 hours in an argon environment at a temperature of 80 ° C. Further two-stage purification - extraction and column chromatography on silica gel - was carried out according to standard methods. Three new compounds were obtained (Table 1).

[0017] The synthesis of 2-substituted ethynylperylene derivatives was carried out by introducing the starting compound, 2-ethynylperylene, into the Sonogashira reaction with 4-iodophenol, 3-iodophenol and 2-bromo-4-iodophenol, as well as copper iodide and dichlorobis(triphenylphosphine)palladium catalysts, in dimethylformamide with the addition of triethylamine. The reaction proceeded for 24 hours in an argon environment at a temperature of 80°C. Further two-stage purification - extraction and column chromatography on silica gel - was carried out using standard methods. Three new compounds were obtained (Table 1).

[0018] Table 1.

[0019] The obtained components are characterized by physicochemical and biological methods.

[0020] The described method allows obtaining photosensitizers in quantities and with purity sufficient for conducting physicochemical and biological studies.

[0021] 1.4 Carrying out the invention Example 1. Method for the synthesis of 3-substituted ethynylperylene derivatives. 3-ethynyl perylene (1.2 equiv.), the corresponding iodophenol (1 equiv.), bis(triphenylphosphine)dichloropalladium (0.05 equiv.) and copper(I) iodide (0.1 equiv.) were dissolved in dry DMF (100 ml). The mixture was evacuated and purged with argon 5 times, then triethylamine (5 equiv.) was added. The reaction mixture was heated to 80°C and left stirring for 24 hours (Fig. 2).

[0022] Example 2. A method for isolating the photosensitizers of 3-substituted ethynylperylenes involved diluting the reaction mixture with ethyl acetate (150 ml), washing it with 1% aqueous EDTA (200 ml), water (3 x 200 ml), and concentrated sodium chloride solution (200 ml), drying it over anhydrous sodium sulfate, filtering it, and evaporating the solvent in vacuo to give a crude solid. The residue was purified by column chromatography on silica gel with solvent gradients of 0 to 2% ethyl acetate in dichloromethane for HOPY11 and HO3PY11, and with pure toluene for HOBrPY11. The appropriate fractions were evaporated to give the desired compounds as colored solids.

[0023] Example 3. Method for the synthesis of 2-substituted ethynylperylene derivatives. 2-ethynylperylene (1.2 equiv.), the corresponding iodophenol (1 equiv.), bis(triphenylphosphine)dichloropalladium (0.05 equiv.) and copper(I) iodide (0.1 equiv.) were dissolved in dry DMF (-100 ml). The mixture was evacuated and purged with argon 5 times, then triethylamine (5 equiv.) was added. The reaction mixture was heated to 80°C and left stirring for 24 h (Fig. 3).

[0024] Example 4. A method for isolating 2-substituted ethynylperylene photosensitizers involved diluting the reaction mixture with ethyl acetate (150 ml), washing it with 1% aqueous EDTA (200 ml), water (3 x 200 ml), and concentrated sodium chloride solution (200 ml), drying it over anhydrous sodium sulfate, filtering it, and evaporating the solvent in vacuo to give a crude solid. The residue was purified by column chromatography on silica gel with solvent gradients of 0 to 2% ethyl acetate in dichloromethane for HOPY13 and HO3PY13, and with pure toluene for HOBrPY13. The appropriate fractions were evaporated to give the desired compounds as colored solids.

[0025] Example 5. Determination of the main physicochemical and spectral characteristics of the obtained photosensitizers HOPY11, HOPY13, HO3PY11, HO3PY13, HOBrPY11 and HOBrPY13.

[0026] The obtained samples HOPY11, HOPY13, HO3PY11, HO3PY13, HOBrPY11 and HOBrPY13 are solid substances of yellow, red or brown color. Perylene compounds have a characteristic absorption spectrum in the UV range:

[0027] HOPY11 (96% ETHANOL) λ mах , nm 466;

[0028] HO3PY11 (96% ethanol) λ mах , nm 463;

[0029] HOBrPY11 (96% ETHANOL) λ mах , nm 466;

[0030] HOPY13 (96% ethanol) λ mах , nm 438;

[0031] HO3PY13 (96% ETHANOL) λ mах , nm 438;

[0032] HOBrPY13 (96% ethanol) λ mах , nm 438.

[0033] When irradiated in the UV range, fluorescence is observed for all substances:

[0034] HOPY11 (96% ethanol) λ mах , nm 476;

[0035] HO3PY11 (96% ETHANOL) λ mах , nm 472;

[0036] HOBrPY11 (96% ethanol) λ mах . nm 475;

[0037] HOPY13 (96% ethanol) λmах , nm 442;

[0038] HO3PY13 (96% ethanol) λ mах , nm 440;

[0039] HOBrPY13 (96% ethanol) λ mах , nm 442.

[0040] All the obtained compounds generated singlet oxygen, the quantum yield of which was:

[0041] HOPY11 (96% ethanol) fd( 1 O2) 0.422;

[0042] HO3PY11 (96% ethanol) fd( 1 O2) 0.451 ;

[0043] HOBrPY11 (96% ethanol) fd( 1 O2) 0.282;

[0044] HOPY13 (96% ethanol) fd( 1 O2) 0.297;

[0045] HO3PY13 (96% ethanol) fd( 1 O2) 0.460;

[0046] HOBrPY13 (96% ethanol) fd( 1 O2) 0.332.

[0047] Example 6. Determination of the composition and structure of HOPY11, HOPY13, HO3PY11, HO3PY13,

[0048] HOBrPY11 and HOBrPY13.

[0049] The gross formulas of HOPY11, HOPY13, HO3PY11, HO3PY13, HOBrPY11 and HOBrPY13 are established on the basis of high-resolution mass spectra. 1 high-resolution mass spectrum of the compounds:

[0050] 1. HOPY11 shows an ion with m / z 367.1128, which is consistent with the gross formula C 28 N 15 O- (calculated m / z = 367.1128 for [M-H]").

[0051] 2. HO3PY11 ion with m / z 369.1286 is observed, which is consistent with the gross formula C 28 H 17 O + (calculated m / z = 369.1274 for [M+H] + ).

[0052] 3. HOBrPY11 ion with m / z 447.0386 is observed, which is consistent with the gross formula C 28 N 16 VGO + (calculated m / z = 447.0379 for [M+H] + ).

[0053] 4. HOPY13 ion with m / z 369.1285 is observed, which is consistent with the gross formula C 28 H 17 O + (calculated m / z = 369.1274 for [M+H] + ).

[0054] 5. HO3PY13 ion with m / z 369.1282 is observed, which is consistent with the gross formula C 28 H 17 O + (calculated m / z = 369.1274 for [M+H] + ).

[0055] 6. HOBrPY13 ion with m / z 447.0390 is observed, which is consistent with the gross formula C 28 N 16 VGO + (calculated m / z = 447.0379 for [M+H] + ).

[0056] The structure of HOPY11, HOPY13, HO3PY11, HO3PY13, HOBrPY11 and HOBrPY13 was determined based on NMR data. Assignment of signals in NMR spectra 1 N and 13 The analysis was carried out on the basis of the assumed structure of the compounds. The spectra were recorded for solutions of individual substances in perdeuterodimethyl sulfoxide at room temperature.

[0057] HOPY11 'H NMR (500 MHz, DMSO-d6, δ, ppm, J / Hz) δ 9.92 (s, 1H, OH), 8.46 (d, 1H, J 7.2 Hz, H4'), 8.41 (d, 1H, J = 7.3 Hz, H12'), 8.39 (d, 1H, J = 7.3 Hz, H7'), 8.36 (d, 1H, J = 8.0 Hz, NH), 8.24 (d, 1H, J = 8.3 Hz, H6'), 7.85-7.82 (t, 2H, H9', H10'), 7.75 (d, 1H, J = 7.9 Hz, H2'), 7.70 (t, J = 7.7 Hz, SH, Н5'), 7.60 - 7.55 (t, 2Н, Н8', Н1 Г), 7.53 (d, 2Н, J = 8.7 Hz), 6.87 (d, 2Н, J = 8.7 Hz). 13 C NMR (126 MHz, DMSO-d6) δ 158.27, 134.19, 133.73, 133.17, 133.12, 130.97, 130.76, 130.56, 130.14, 129.89, 128.48, 128.27, 127.79, 127.60, 126.97, 125.67, 121.45, 121.35, 121.20, 120.34, 119.96, 115.83, 112.46, 96.36, 85.75.

[0058] HO3PY11. 'H NMR (500 MHz, DMSO-d6, δ, ppm, J / Hz) δ 9.74 (s, 1H), 8.45 (d, J= 7.5 Hz, SH), 8.40 (s, 1H), 8.37-8.29 (m, 2H), 8.00 (s, 1H), 7.88-7.74 (m, 3H), 7.62-7.50 (m, 3H), 7.27 (t, J= 7.9 Hz, 1H), 7.08 (d, J= 7.3 Hz, 1H), 7.02 (s, SH), 6.91-6.82 (m, 1H). 13C NMR (126 MHz, DMSO- <d6) δ 157.34, 134.17, 134.08, 131.14, 130.69, 130.67, 130.45, 129.91, 129.83, 129.79, 129.45, 128.46, 128.43, 128.18, 128.14, 127.75, 127.67, 127.64, 127.56, 127.52, 127.34, 126.81, 126.76, 123.06, 122.38, 122.28, 121.38, 120.92, 120.68, 117.81, 117.79, 116.38, 89.88, 88.85. HOBrPYll. 1 H NMR (500 MHz, DMSO-d6, δ, m.d., J / Hz) 8 10.88 (s, 1H), 8.44 (d, J= 7.5 Hz, 1H), 8.42-8.30 (m, 3H), 8.51, 81.2 (d, G= J= (m, 1H), 7.86–7.78 (m, 2H), 7.75 (d, J= 7.8 Hz, 1H), 7.68 (t, J= 7.9 Hz, 1H), 7.59–7.50 (m, 3H), 7.04 (H, J= 8.4). 13 C NMR (126 MHz, DMSO- <d6) δ 155.12, 135.76, 134.17, 133.74, 132.26, 131.08, 130.96, 130.87, 130.12, 129.84, 128.56, 128.28, 127.85, 127.76, 127.60, 126.96, 126.92, 125.75, 121.53, 121.35, 121.21,

[0059] 120.26, 119.49, 116.56, 114.27, 109.45, 94.56, 86.81.

[0060] HOPY13. 1N NMR (500 MHz, DMSO- <d6, δ м.д., J / Гц) 8 9.97 (s, 1Н), 8.42 (d, J= 7.5 Гц, Ш), 8.37-8.29 (m, ЗН), 7.94 (s, Ш), 7.84-7.73 (т, ЗН), 7.58-7.50 (т, ЗН), 7.48 (d, J= 8.3 Гц, 2Н), 6.86 (d, J= 8.3 Гц, 2Н). 13 With NMR (126 MHz, DMSO-d6) δ 158.13, 134.16, 134.13, 133.04, 131.02, 130.41, 130.15, 129.95, 129.50, 128.37, 128.11, 127.75, 127.57, 127.46, 127.11, 126.76, 122.34, 121.26, 121.15, 120.86, 115.75, 112.37, 90.49, 87.52.

[0061] HO3PY13. *H NMR (500 MHz, DMSO-d6, δ, ppm, J / Hz) 8 9.74 (s, 1H), 8.45 (d, J= 7.5 Hz, 1H), 8.40 (s, SH), 8.37-8.29 (m, 2H), 8.00 (s, 1H), 7.88-7.74 (m, 3H), 7.62-7.50 (m, 3H), 7.27 (t, J = 7.9 Hz, 1H), 7.08 (d, J = 7.3 Hz, 1H), 7.02 (s, 1H), 6.91-6.82 (m, 1H). 13With NMR (126 MHz, DMSO-d6) δ 157.34, 134.17, 134.08, 131.14, 130.69, 130.67, 130.45, 129.91, 129.83, 129.79, 129.45, 128.46, 128.43, 128.18, 128.14, 127.75, 127.67, 127.64, 127.56, 127.52, 127.34, 126.81, 126.76, 123.06, 122.38, 122.28, 121.38, 120.92, 120.68, 117.81, 117.79, 116.38, 89.88, 88.85

[0062] HOBrPY13. 1 H NMR (500 MHz, DMSO-d6, δ, ppm, J / Hz) 8 10.87 (s, 1H), 8.44-8.28 (m, 4H), 7.94 (s, 1H), 7.84-7.71 (m, 4H), 7.56-7.45 (m, 4H), 7.04 (d, J= 8.4 Hz, 1H). 13 C NMR (126 MHz, DMSO-d6) 8 155.09, 135.86, 134.24, 134.15, 132.16, 131.15, 130.50, 130.42, 129.99, 129.56, 128.53, 128.25, 127.81, 127.74, 127.60, 127.31, 126.86, 122.49, 121.39, 121.36, 121.00, 120.88,

[0063] 116.58, 114.17, 109.38, 88.88, 88.65.

[0064] Biological experiments showed that the compounds HOPY11, HOPY13, HO3PY11, HO3PY13, HOBrPY11 and HOBrPY13 have significant antiviral activity against the SARS-CoV-2 coronavirus. The obtained data are presented in Table 2, where the concentration EC 50 demonstrates the half-maximal effective concentration at which 50% of virions are inactivated, and CC50 shows the concentration of compounds required to reduce cell viability by 50%.

[0065] Table 2.

Claims

Invention formula 1. A photosensitizer expressed by the general structural formula representing 3-substituted ethynylperylenes (HOPY11, HO3PY11, HOBrPY11) and 2-substituted ethynylperylenes (HOPY13, HO3PY13, HOBrPY13), where R is: ethynyl aryl, para-substituted ethynyl phenol, meta-substituted ethynyl phenol, ethynyl bromophenol; substituent in the 2-position: ethynyl aryl, para-substituted ethynyl phenol, meta-substituted ethynyl phenol, ethynyl bromophenol; 2. A method for synthesizing derivatives of a photosensitizer according to claim 1, comprising introducing into a Sonogashira reaction with 4-iodophenol (HOPY11), 3-iodophenol (HO3PY11) and 2-bromo-4-iodophenol (HOBrPY11), with 4-iodophenol (HOPY13), 3-iodophenol (HO3PY13) and 2-bromo-4-iodophenol (HOBrPY13), with catalysis by copper iodide and palladium tetrakis-triphenylphosphine in an argon atmosphere.

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