Cancer cell proliferation inhibiting composition and processed food

Abstract

It is known that phytol contained in watermelon seedlings has a cancer cell proliferation inhibition effect. However, the problem is that in order to exhibit a cancer cell proliferation inhibiting effect, a large dose is required. A composition for inhibiting proliferation of cancer cells, which contains, as an active ingredient, at least one of a compound having a structure represented by formula (123) or a pharmaceutically acceptable salt thereof, has a higher inhibitory effect on proliferation of cancer cells than phytol, has a low inhibitory effect on proliferation of normal cells, and can be used as an anticancer agent having excellent selectivity for cancer cells. And the '120' # imgabs0 # R is alkyl with 1 to 3 carbon atoms.

Description

Technical Field

[0001] The present invention relates to a pharmaceutical composition and processed food having a thiochlorophyll substance and having an inhibitory effect on cancer cell proliferation. Background Art

[0002] It is known that an extract of watermelon seedlings has an inhibitory effect on cancer cell proliferation (Patent Document 1). This document shows that among the extracts of watermelon seedlings, especially phytol and lutein are effective for this effect. Moreover, since these substances do not affect normal cells, it is expected that they can provide an anti-cancer pharmaceutical composition with few side effects. Prior Art Documents Patent Documents

[0003] Patent Document 1: International Publication No. 2017 / 131175 Summary of the Invention Technical Problem to be Solved by the Invention

[0004] Phytol and lutein are expected to be used as an anti-cancer pharmaceutical composition. However, the problem is that a relatively large dosage is required to exert the effect. Technical Solution for Solving the Technical Problem​​​​​​​​​​​​​​​​​​​​​​

[0011]

Chemistry 3

[0012]

Chemistry 4

[0013]

Chemistry 5

[0014]

Chemistry 6

[0015] The cancer cell proliferation inhibitory composition of the present invention can inhibit the growth of cancer cells. Therefore, it can be advantageously used as an anticancer pharmaceutical composition. Furthermore, since the processed food of the present invention contains a compound that has a cancer cell proliferation inhibitory effect, regular consumption as a supplement can prevent cancer. DETAILED DESCRIPTION

[0016] The following examples illustrate the cancer cell proliferation inhibitory composition and processed food according to the present invention. The following description merely illustrates one embodiment and one example of the present invention and is not intended to limit the present invention. The following description may be modified without departing from the spirit of the present invention.

[0017] The cancer cell proliferation inhibitory composition of the present invention is composed of compounds having structures represented by formula (1) to formula (5) or pharmaceutically acceptable salts thereof.

[0018]

Chemistry 7

[0019]

Chemistry 8

[0020]

Chemistry 9

[0021]

Chemistry 10

[0022]

Chemistry 11

[0023] The compound of formula (1) is referred to as "PT42," the compound of formula (2) is referred to as "PT43," the compound of formula (3) is referred to as "PT44," the compound of formula (4) is referred to as "PT40," and the compound of formula (5) is referred to as "PT41." Furthermore, formulas (1), (2), and (3) are collectively referred to as formula (123).

[0024]

Chemistry 12

[0025] Here, R is a C1-C3 alkyl group.

[0026] When these compounds are used as compositions (pharmaceutical compositions) for inhibiting cancer cell proliferation, in addition to being used alone, they can also be used as salts mixed with pharmaceutically acceptable acids in solvents such as water, methanol, ethanol, and acetone. Examples of pharmaceutically acceptable acids include inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, and nitric acid, and organic acids such as acetic acid, propionic acid, oxalic acid, succinic acid, lactic acid, malic acid, tartaric acid, citric acid, maleic acid, fumaric acid, methanesulfonic acid, p-toluenesulfonic acid, and ascorbic acid.

[0027] The administration method of the cancer cell proliferation inhibitory composition of the present invention when used as a pharmaceutical composition is not particularly limited and can be any oral or parenteral administration method. Furthermore, the composition can be formulated into a suitable dosage form depending on the administration method, for example, injections, oral preparations such as capsules, tablets, granules, powders, pills, and fine granules, rectal preparations, oily suppositories, and aqueous suppositories.

[0028] Furthermore, when the cancer cell proliferation inhibitory composition of the present invention is used as a pharmaceutical composition, the ratio of the active ingredient can be varied depending on the application. That is, the composition can be administered at a concentration high enough to inhibit the growth of the target cancer cells.

[0029] In addition, the cancer cell proliferation inhibitory composition of the present invention can also be provided as a processed food. Examples of processed foods include not only general processed foods such as snacks or health foods, such as candy, chewing gum, jelly, biscuits, cookies, rice crackers, bread, noodles, processed fish and meat products, tea, soft drinks, coffee drinks, milk drinks, whey drinks, lactic acid bacteria drinks, yogurt, ice cream, and pudding, but also health functional foods such as specific health foods or nutritional functional foods specified in the health functional food system of the Ministry of Health, Labor and Welfare, and further processed foods such as nutritional supplements, feeds, and food additives.

[0030] The processed food according to the present invention can be prepared by adding the cancer cell proliferation inhibitory composition to the raw materials of these processed foods. Example

[0031] <1. Synthesis of Compounds> First, (E)-1-Bromo-3,7,11,15-tetramethylhexadec-2-ene represented by formula (C3): (E)-1-bromo-3,7,11,15-tetramethylhexadec-2-ene was synthesized as a starting material.

[0032]

Chemistry 13

[0033] To a 200 mL two-necked flask, phytol (CAS No. 7541-49-3) (500 mg, 1.69 mmol) dissolved in dry Et2O (diethyl ether) (9 mL) was added and stirred at 0°C for a period of time. Phytol (CAS No. 7541-49-3) was then added dropwise, and the mixture was stirred at the same temperature for 30 minutes. Phytol has the structure of formula (C1).

[0034]

Chemistry 14

[0035] The reaction was confirmed by TLC (Hexane / AcOEt = 10 / 1). After confirming the disappearance of the starting material on TLC, saturated aqueous sodium bicarbonate solution was added dropwise to stop the reaction. Extraction was performed with ethyl acetate, and the organic layer was collected, neutralized with saturated aqueous sodium bicarbonate solution, washed with brine, and dried over anhydrous sodium sulfate. The sodium sulfate was removed by natural filtration, and the solution was concentrated under reduced pressure on a rotary evaporator to obtain 660.8 mg of a reddish-brown oily crude product. No purification was performed.

[0036] The NMR spectrum of the product obtained by the above steps is shown below. (Reddish brown oil) 1 H NMR (400 MHz, CDCl3): δ = 0.82-0.89 (m, 14H), 1.02-1.41 (m, 21H), 1.50-1.62 (m, 2H), 1.72 (d, 3H, J = 1.3 Hz), 2.02 (t, 2H, J = 7.6 Hz), 4.04 (d, 2H, J = 8.4 Hz), 5.53 (t, 1H, J = 8.4 Hz). These results indicate that the product has the structure of Formula (C1). Hereinafter, the compound of Formula (C3) will be referred to as "PT7."

[0037] <1-1> Synthesis of Formula (C2) (referred to as "PT37") To a dichloromethane solution (7 mL) of sodium thiomethoxide (258 mg, 3.7 mol, 2.0 eq) was added a dichloromethane solution (7 mL) of PT7 (0.66 g, 1.8 mmol, 1.0 eq). The mixture was reacted at the same temperature for 2 hours, then allowed to return to room temperature over 16 hours. Completion of the reaction was confirmed by TLC (developed with hexane). Water was added, and the mixture was extracted three times with dichloromethane. The recovered organic layer was dried over sodium sulfate, and the solvent was removed in vacuo. PT37 (346 mg, 63%) was obtained by purification via silica gel column chromatography (hexane / ethyl acetate = 80 / 1) as a colorless oil.

[0038] The NMR chemical shift and mass spectrometry analysis results of the product obtained by the above steps are shown below. 1 H NMR (CDCl 3, 400MHz): δ5.23(1H,tq,J=7.9,1.2Hz,CH at C2),3.13(2H,d,J=7.8Hz,CH2 atC1),2.03(3H,s,-SCH3),2.01(2H,t,J=7.6Hz,CH2 at C4),1.64(3H,d,J=0.7Hz,CH3 atC3),1.57-1.47(3H,sept,J=6.6Hz,CH at C15),1.46-0.99(21H,m),0.87(6H,d,J=6.7Hz,CH3 at C15),0.85(3H,d,J=6.6Hz,CH3 at C11 or C7),0.84(3H,d,J=6.6Hz,CH3at C11or C7). 13 C NMR (CDCl 3, 100MHz): δ139.4,120.0,39.9,39.4,37.5,37.4,37.4,37.3,36.6,32.8,32.7,31.2,28.0,25.3,24.8,24.5,22.7,22.6,19.8,16.0,14.5. The above measurements confirmed that it was the compound of formula (C2).

[0039]

Chemistry 15

[0040] <1-2> (4) Synthesis of formula (“PT40”) To a solution of PT37 (Formula (C2)) (170 mg, 0.52 mmol, 1.0 eq) in acetonitrile (2.6 mL) was added an aqueous solution of sodium hypochlorite pentahydrate (250 mg, 1.5 mmol, 2.2 eq) (0.27 mL), and the mixture was stirred for 20 minutes. The progress of the reaction was confirmed by TLC (developed with chloroform / methanol = 19:1). After completion of the reaction, an aqueous sodium sulfite solution was added, and the mixture was extracted three times with ethyl acetate. The recovered organic layer was dried over sodium sulfate, and the solvent was removed in vacuo. PT40 (38 mg, 16%) was obtained by purification via silica gel column chromatography (chloroform) as a colorless oil.

[0041] The NMR chemical shift and mass spectrometry analysis results of the product obtained by the above steps are shown below. 1 H NMR (CDCl 3, 400MHz): δ5.24(1H,tq,J=8.0,1.1Hz,CH at C2),3.58(1H,dd,J=13.0,8.0Hz,CH2at C1),3.43(1H,dd,J=13.0,8.0Hz,CH2at C1),2.53(3H,s,-SCH3),2.06(2H,t,J=7.3Hz,CH2 at C4),1.72(3H,s,CH3 at C3),1.52(1H,sept,J=6.6Hz,CH at C15),1.46-0.99(25H,m),0.87(6H,d,J=6.6Hz,CH3 at C15),0.85(3H,d,J=6.6Hz,CH3 at C11 or C7),0.84(3H,d,J=6.6Hz,CH3 at C11 or C7). 13 C NMR (CDCl 3, 100MHz): δ145.9,110.7,53.6,40.1,39.4,37.4,37.4,37.3,37.3,36.6,32. 8,32.7,28.0,25.2,24.8,24.4,22.7,22.6,19.8,19.7,16.9.HRMSm / z[M+Na] + C 21 H 42 The calculated value of OS is 365.2854; the measured value is 365.2858.

[0042] <1-3> Synthesis of PT41 (Formula (5)) To a toluene solution (4.4 mL) of PT37 (Formula (C2)) (480 mg, 1.5 mmol, 1.0 eq) was added an aqueous solution (1.3 mL) of sodium hypochlorite pentahydrate (580 mg, 3.5 mmol, 2.4 eq), and the mixture was stirred for 3 hours. The progress of the reaction was confirmed by TLC (developed with hexane / ethyl acetate = 5:1). After completion of the reaction, an aqueous sodium sulfite solution was added, and the mixture was extracted three times with ethyl acetate. The recovered organic layer was dried over sodium sulfate, and the solvent was removed in vacuo. PT41 (45 mg, 9%) was obtained as a colorless oil by purification via silica gel column chromatography (hexane / ethyl acetate = 50:1).

[0043] The NMR chemical shift and mass spectrometry analysis results of the product obtained by the above steps are shown below. 1 H NMR (CDCl 3, 400MHz): δ5.35(1H,tq,J=7.9,1.2Hz,CH at C2),3.73(2H,d,J=7.9Hz,CH2 atC1),2.82(3H,s,-SCH3),2.09(2H,t,J=7.2Hz,CH2 at C4),1.74(3H,s,CH3 at C3),1.52(1H,sept,J=6.6Hz,CH at C15),1.47-0.99(21H,m),0.87(6H,d,J=6.5Hz,CH3 at C15),0.85(3H,d,J=5.4Hz,CH3 at C11 or C7),0.84(3H,d,J=6.6Hz,CH3 at C11 or C7). 13 CNMR (CDCl 3, 100MHz): δ146.1,109.8,54.1,39.4,38.7,38.4,36.8,36.7,36.6,36.0,32. 1,32.0,27.3,24.4,24.1,23.8,22.1,22.0,19.1,19.0,16.0.HRMSm / z[M+Na] + C 21 H 42 The calculated value of O2S is 381.2803; the measured value is 381.2786.

[0044] <1-4> Synthesis of PT42 (Formula (1)) To a dry dichloromethane solution (0.68 mL) of phytol (CAS No. 7541-49-3) (200 mg, 0.67 mmol, 1.0 eq) at -20°C was added dimethyl sulfide (500 μL, 420 mg, 6.7 mmol, 10 eq). The mixture was warmed to 0°C over 30 minutes, and a solution of tetrafluoroborate in ether (54 wt%, 249 μL, 296 mg, 3.37 mmol, 5.0 eq) was added. The mixture was then allowed to return to room temperature over 4 hours, and after an additional 20 hours, completion of the reaction was confirmed by TLC (developed with chloroform / methanol = 9:2). Ethyl acetate was added, and the mixture was washed twice with saturated sodium bicarbonate solution. The recovered organic layer was dried over sodium sulfate, and the solvent was removed in vacuo. The product was suspended in hexane and transferred to a centrifugal precipitation tube. After centrifugation, the supernatant was removed to obtain PT42 (243 mg, 84%) as a white solid.

[0045] The NMR chemical shift and mass spectrometry analysis results of the product obtained by the above steps are shown below. 1 H NMR (CDCl 3, 400MHz): δ5.23(1H,tq,J=8.1,1.1Hz,CH at C2),4.05(1H,d,J=8.1Hz,CH2 atC1),2.85(6H,s,-S + CH3),2.11(2H,m,CH2 at C4),1.81(3H,d,J=0.88Hz,CH3 at C3),1.52(1H,sept,J=6.6Hz,CH at C15),1.47-0.99(18H,m),0.87(6H,d,J=4.6Hz,CH3 at C15),0.85(3H,d,J=6.5Hz,CH3 at C11 or C7),0.85(3H,d,J=7.5Hz,CH3 at C11 or C7). 13 CNMR (CDCl 3, 100MHz): δ153.1,107.6,40.4,40.3,39.4,37.4,37.4,37.3,36.8,32.8,32.7,28.0,25.3,24.8,24.5,22.9,22.6,19.7,19.6,17.0. 19 F NMR (CDCl 3, 376MHz): δ-149.85,-149.90.HRMSm / z[M] + C 22 H 45 The calculated value of S is 341.3242; the measured value is 341.3237.

[0046] <1-5> Synthesis of PT43 (Formula (2)) To a dry dichloromethane solution (0.68 mL) of phytol (CAS No. 7541-49-3) (200 mg, 0.67 mmol, 1.0 eq) at -20°C was added diethyl sulfide (720 μL, 610 mg, 6.7 mmol, 10 eq). The mixture was warmed to 0°C over 30 minutes, and a solution of tetrafluoroborate in ether (54 wt%, 249 μL, 296 mg, 3.37 mmol, 5.0 eq) was added. The mixture was then allowed to return to room temperature over 4 hours, and after an additional 20 hours, completion of the reaction was confirmed by TLC (developed with chloroform / methanol = 9:2). Ethyl acetate was added, and the mixture was washed three times with saturated aqueous sodium bicarbonate. The recovered organic layer was dried over sodium sulfate, and the solvent was removed in vacuo to obtain PT43 (263 mg, 85%) as a dark brown oil.

[0047] The NMR chemical shift and mass spectrometry analysis results of the product obtained by the above steps are shown below. 1 H NMR (CDCl 3, 400MHz): δ5.23(1H,tq,J=8.1,1.1Hz,CH at C2),4.08(1H,d,J=8.0Hz,CH2 atC1),3.37(2H,q,J=7.5Hz,-S + CH2), 3.36(2H,q,J=7.4Hz,-S + CH2),2.09(2H,m,CH2 at C4),1.82(3H,d,J=0.88Hz,CH3at C3),1.52(1H,sept,J=6.6Hz,CH at C15),1.52(6H,t,J=7.5Hz,-S + CH2CH3),1.46-0.97(19H,m),0.87(6H,d,J=6.4Hz,CH3 at C15),0.85(3H,d,J=6.1Hz,CH3 at C11 or C7),0.85(3H,d,J=6.6Hz,CH3 at C11or C7). 13 C NMR (CDCl 3, 100MHz): δ151.9,108.5,40.2,39.4,37.9,37.4,37.4,37.3,36.8,33.5 ,32.8,32.7,28.0,25.2,24.8,24.5,22.7,22.6,19.7,19.6,17.0,9.7. 19F NMR (CDCl 3, 376MHz): δ-149.79,-149.85.HRMSm / z[M] + C 24 H 49 The calculated value of S is 369.3555; the measured value is 369.3552.

[0048] <1-6> Synthesis of PT44 (Formula (3)) To a solution of phytol (200 mg, 0.67 mmol, 1.0 eq) in dry dichloromethane (0.68 mL) at -20°C was added diisopropyl sulfide (970 μL, 800 mg, 6.7 mmol, 10 eq). The mixture was warmed to 0°C over 30 minutes, and a solution of tetrafluoroborate in ether (54 wt%, 249 μL, 296 mg, 3.37 mmol, 5.0 eq) was added. The mixture was then allowed to return to room temperature over 4 hours, and after another 20 hours, the reaction was confirmed to be complete by TLC (developed with chloroform / methanol = 9:2). Saturated aqueous sodium bicarbonate solution was added, and the mixture was extracted three times with ethyl acetate. The recovered organic layer was dried over sodium sulfate, and the solvent was removed in vacuo. The product was suspended in hexane and transferred to a centrifugal precipitation tube. After centrifugation, the supernatant was removed to obtain PT44 (247 mg, 76%) as a light brown solid.

[0049] The NMR chemical shift and mass spectrometry analysis results of the product obtained by the above steps are shown below. 1 H NMR (CDCl 3, 400MHz): δ5.24(1H,tq,J=7.8,1.1Hz,CH at C2),4.07(1H,d,J=7.7Hz,CH2 atC1),3.94(2H,sept,J=6.8Hz,-S + CH),2.07(2H,m,CH2 at C4),1.81(3H,s,CH3 at C3),1.62(6H,d,J=6.8Hz,-S + CHCH3), 1.60(6H, d, J=6.9Hz,-S + CHCH3),1.52(1H,sept,J=6.6Hz,CH at C15),1.46-0.99(20H,m),0.87(6H,d,J=6.9Hz,CH3 at C15),0.85(3H,d,J=7.2Hz,CH3 at C11 or C7),0.84(3H,d,J=6.7Hz,CH3 at C11 or C7). 13 C NMR (CDCl3, 100MHz): δ149.2,110.7,44.4,40.1,39.3,37.4,37.4,37.3,36.7,34.2 ,32.8,32.6,28.0,25.0,24.8,24.4,22.6,19.7,19.6,19.3,18.4,16.8. 19 F NMR (CDCl 3, 376MHz): δ-150.40,-150.45.HRMSm / z[M] + C 26 H 53 The calculated value of S is 397.3868; the measured value is 397.3870.

[0050] <1-7> Synthesis of PT49 (Formula (6)) To a dry dichloromethane solution (0.68 mL) of phytol (200 mg, 0.67 mmol, 1.0 eq) at -20°C was added di-tert-butyl sulfide (1.2 mL, 0.99 g, 6.7 mmol, 10 eq). The mixture was heated to 0°C over 30 minutes, and a solution of tetrafluoroborate in ether (54 wt%, 249 μL, 296 mg, 3.37 mmol, 5.0 eq) was added. The mixture was then allowed to return to room temperature over 4 hours, and after another 20 hours, the reaction was confirmed to be complete by TLC (developed with chloroform / methanol = 9:2). Ethyl acetate was added, and the mixture was washed twice with saturated sodium bicarbonate aqueous solution. The recovered organic layer was dried over sodium sulfate, and the solvent was removed in vacuo. The product was suspended in hexane and transferred to a centrifugal precipitation tube. After centrifugation, the supernatant was removed to obtain the crude product PT49 as a light brown solid.

[0051]

Chemistry 16

[0052] Prepare 2×10 550 μL of Jurkat cell suspension with a concentration of 10 cells / mL was inoculated into each well of a 96-well plate. The DMSO solution of the evaluation sample was diluted 500 times with culture medium. Then, 50 μL of the diluted sample solution was immediately added to start cell treatment. After 72 hours of culture in the incubator, 10 μL of the viable cell assay reagent SF was added. After a 2-3 hour color development reaction in the incubator, the absorbance at 450 nm and 630 nm was measured using a microplate reader, and the cell viability was calculated using the following formula (F1).

[0053] Cell survival rate (%) = [(As-Ab) / (Ac-Ab)] × 100...(F1) In addition, As, Ab, and Ac are as follows. As: absorbance of the evaluation well Ab: absorbance of negative control well Ac: absorbance of control well

[0054] The absorbance in the above formula is the absorbance at 450nm (denoted as A 450 ) minus the absorbance at 630 nm (denoted as A 630 ) and the difference "A 450 -A 630 In addition, the two concentrations that were most closely sandwiched between 50% cell viability were expressed logarithmically, plotted, and linear fitting was performed to calculate the IC 50 The results are shown in Table 1.

[0055] [Table 1]

[0056] Phytol (Formula (C1)) and PT37 (Formula (C2)) are shown as comparative examples. IC values ​​of phytol (Formula (C1)), PT37 (Formula (C2)), PT40 (Formula (4)), and PT41 (Formula (5)) for Jurkat cells 50 The values ​​were 12.8 μM, 100 μM or more, 30 μM, and 100 μM or more, respectively. The new substance PT40 (Formula (4)) also has a cancer cell proliferation inhibitory effect.

[0057] IC values ​​of PT42 (Formula (1)), PT43 (Formula (2)), and PT44 (Formula (3)) for Jurkat cells 50 The values ​​were 1.9 μM, 0.55 μM, and 0.41 μM, respectively, showing a very high inhibitory effect on cancer cell proliferation. Possibility of industrial application

[0058] The cancer cell proliferation inhibitory composition according to the present invention can be advantageously used for the treatment or prevention of cancer.

Claims

1. A compound having a structure represented by formula (123): 【Chemistry 100】 R is a C1-C3 alkyl group.

2. A composition for inhibiting cancer cell proliferation, comprising at least one of the compounds according to claim 1 or pharmaceutically acceptable salts thereof as an active ingredient. 3 . A processed food comprising at least one of the compounds according to claim 1 or pharmaceutically acceptable salts thereof.

4. Compounds having the structure of formula (4): 【Chemistry 101】 5. Compounds having the structure of formula (5): 【Chemistry 102】

Images