Synthesis of adamantane-based formic acid derivative and antitumor activity research of adamantane-based formic acid derivative

Abstract

The invention discloses synthesis of a novel drug small molecular formula (1) containing an adamantane structure and an antitumor activity research # imgabs0 #, wherein R is # imgabs1 #. 3-bromo-1-adamantanecarboxylic acid is used as an initial raw material, and then free radical substitution, oxidation reaction and demethylation reaction are performed to obtain 5-(adamantane-1-yl)-2-hydroxybenzaldehyde; secondly, 5-(adamantane-1-yl)-2-hydroxybenzaldehyde and amino are subjected to a condensation reaction, and a target compound is obtained. A CCK-8 experiment is carried out on a target compound to obtain a conclusion that the target compound 8a-8f and 5-fluorouracil are tested at the concentration of 20 [mu] M, and the result shows that the compound 8c has the best inhibition effect on HeLa and A549 cells, and the compound 8a has the better inhibition effect on HepG2 cells. IC50 value calculation is carried out on a target compound under various concentrations, the IC50 value of 8c on HeLa and A549 cells is minimum and the inhibition effect is the best, and the IC50 value of 8a on HepG2 cells is minimum and the inhibition effect is the best.

Description

Technical Field

[0001] The present invention relates to the synthesis of a small drug molecule containing an adamantane structure and the study of its anti-tumor activity. Background Art

[0002] Cancer is considered the second most deadly disease. According to the World Cancer Report 2021 released by the World Health Organization (WHO) on February 3, 2021, the global incidence and mortality of cancer are increasing at an alarming rate, with an average of 1 in 8 deaths caused by cancer. The report predicts that global cancer cases will continue to grow, from 14 million in 2021 to 19 million in 2025, and will reach 24 million in 2035. In developing and developed countries, lung cancer, breast cancer, colon cancer and melanoma are the most common types of cancer. In developed countries, lung cancer and breast cancer are more common. Traditional tumor treatments include surgical treatment, radiotherapy, and chemotherapy. However, it is well known that chemotherapy can cause many adverse reactions, such as: bone marrow suppression (blood cell reduction), gastrointestinal reactions, hair loss, fever, rash, cardiotoxicity, liver toxicity, neurotoxicity, urinary tract toxicity, local phlebitis, teratogenicity, mutagenicity, carcinogenicity, etc., so it is imperative to continuously find better anti-tumor drugs.

[0003] Adamantane is mainly used for the synthesis of specific drugs such as anticancer and antitumor drugs. It is a highly lipid-soluble alicyclic hydrocarbon, and the skeleton of adamantane is non-toxic, which can be used to change its pharmacological activity. According to previous studies, the introduction of amino groups into molecules can usually improve water solubility and enhance their anticancer activity. Since different substituents have different physicochemical properties, which in turn determine the properties of the target compound, the inhibitory effect on tumor cells will also have different effects. The study of these compounds will provide valuable information for the synthesis and research of new small molecule anticancer inhibitors. Summary of the invention

[0004] The invention discloses a method for preparing a small molecule drug containing an adamantane structure. A CCK-8 method is used to perform a toxicity experiment on cells to evaluate the anti-proliferation effect of the compound on A549, HeLa, MCF-7, Hep G2 and normal mouse fibroblast L929.

[0005] The present invention adopts the following technical scheme, containing a small drug molecule of adamantane structure, the structural formula is:

[0006] In the present invention, R is R1, R2, and R3 are each independently H, OCH3, or C2H4O2.

[0007] The present invention uses 3-bromo-1-adamantanecarboxylic acid as a starting material, undergoes a typical Friedel-Crafts alkylation reaction and a Wittig reaction, and finally undergoes a hydrolysis reaction and other multi-step reactions to synthesize 3-((benzyloxy)carbonyl)adamantan-1-yl)phenoxy)acrylic acid, and the structural formula is:

[0008] In the present invention, 3-((benzyloxy)carbonyl)adamantan-1-yl)phenoxy)acrylic acid and different amine compounds are coupled to obtain a drug small molecule containing an adamantane structure: formula (1).

[0009] The invention is characterized by nuclear magnetic resonance, high-resolution mass spectrometry, infrared spectrum and the like, and the results fully confirm the synthesis of the compound. BRIEF DESCRIPTION OF THE DRAWINGS

[0010] Figure 1 It is a synthetic route; Figure 2 It is the target target product of the synthetic route; Figure 3 is the hydrogen spectrum of the intermediate compound; Figure 4-Figure 9 It is the hydrogen and carbon spectra of the target compound; Fig.10 is the inhibition rate of the target compound and 5-fluorouracil on different cancer cells at a concentration of 20 μM; Fig.11 It is the IC50 value graph of the target compound; DETAILED DESCRIPTION

[0011] The following is a specific and detailed description of an embodiment of the present invention. This embodiment is implemented on the premise of the technical solution of the present invention, and a detailed implementation method and a specific operation process are given. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principle of the present invention, and these improvements and modifications should also be regarded as the scope of protection of the present invention.

[0012] Example 1: Synthesis of 3-((benzyloxy)carbonyl)adamantan-1-yl)phenoxy)acrylic acid (1) Synthesis of benzyl-3-(4-(3-methoxy-3-oxoprop-1-en-1-yl)oxy)phenyl)adamantane-1-carboxylate (Intermediate Compound 5) Take a 100mL single-mouth bottle, add 0.1g of benzyl-3-(4-hydroxyphenyl)adamantane-1-carboxylate, 0.14g of triphenylphosphine, and a certain amount of toluene to dissolve it at room temperature, then add 100μL of methyl propiolate, transfer the entire reaction system to a 115°C oil pan, connect a straight condenser, introduce nitrogen, time, and react for 3 hours. After the reaction is completed, cool to room temperature, concentrate, and elute with an eluent (V petroleum ether: V ethyl acetate = 5:1) through column chromatography. After elution, the solvent is spun out with a rotary evaporator, and the residue is placed in a vacuum drying oven to dry to obtain a white solid. Yield: 55%. 1H NMR(400MHz,Chloroform-d)δ7.80(d,J=12.2Hz,1H),7.34(s,7H),7.01(d,J=8.6Hz,2H),5.54(d,J=12 .2Hz,1H),5.12(s,2H),3.73(s,3H),2.25(s,2H),2.04(s,2H),1.97(s,4H),1.87(s,4H),1.74(s,2H). (2) Synthesis of 3-(4-(3-((benzyloxy)carbonyl)adamantan-1-yl)phenoxy)acrylic acid Take a 100mL single-mouth bottle, add 0.1g of benzyl-3-(4-((-3-methoxy-3-oxoprop-1-en-1-yl)oxy)phenyl)adamantane-1-carboxylate at room temperature, add 1.47g of sodium hydroxide, add 10mL of water, and a certain amount of anhydrous ethanol as the reaction solvent, transfer the entire reaction system to an 80 degree Celsius oil pan, connect a condenser, protect with nitrogen, and reflux for 4 hours. After the reaction is completed, cool it to room temperature, remove the rotor, remove the ethanol, and then add diluted concentrated hydrochloric acid to make the pH value acidic and precipitate flocculent precipitate, filter, and place the filter cake in a vacuum drying oven to dry overnight to obtain a white solid. Yield: 98%. 1H NMR(400MHz,Chloroform-d)δ7.88(d,J=12.2Hz,1H),7.37(s,2H),7.35(s,5H),7.03(d,J=8.5Hz,2H),5.51 (d,J=12.2Hz,1H),5.12(s,2H),2.25(s,2H),2.04(s,2H),1.96(d,J=5.8Hz,4H),1.88(s,4H),1.74(s,2H).

[0014] Example 2: Synthesis of target compounds 8a-8f (1) Synthesis of benzyl-3-(4-((3-oxo-3-(phenylamino)prop-1-en-1-yl)oxy)phenyl)adamantane-1-carboxylate (Compound 8a) Take a 100mL single-mouth bottle, add 0.1g of 3-(4-(3-((benzyloxy)carbonyl)adamantan-1-yl)phenoxy)acrylic acid, 0.13g of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDC), 0.09g of 1-hydroxybenzotriazole (HOBt), 120μL of N,N-Diisopropylethylamine (DIPEA) at room temperature, add a certain amount of dichloromethane and ultra-dry tetrahydrofuran to dissolve it, transfer the entire reaction system to a magnetic stirrer, protect with nitrogen, and react at room temperature for 1 hour. After 1 hour, add 42μL of aniline and continue to react for 12 hours. After the reaction, the product was washed several times with saturated NaCl solution and dichloromethane, dried with anhydrous magnesium sulfate for one hour, filtered after drying, concentrated, and eluted by column chromatography using an eluent (V dichloromethane: V ethyl acetate = 30:1). After elution, the solvent was removed and the residue was placed in a vacuum drying oven and dried overnight to obtain a white solid. Yield: 32%. 1H NMR(400MHz, DMSO-d6)δ9.93(s,1H),7.73(d,J=12.0Hz,1H),7.62(d,J=8.0Hz,2H),7.42(d,J=8.7Hz,2H),7.38–7.26(m,7H),7.14(d,J=8.6Hz,2 H),7.02(t,J=7.3Hz,1H),5.84(d,J=12.0Hz,1H),5.10(s,2H),2.17(s,2H),1.97(d,J=16.1Hz,3H),1.84(dd,J=20.8,11.0Hz,9H),1.68(s,2H).

[0015] (2) Synthesis of benzyl-3-(4-(E)-3-((2-methoxyphenyl)amino)-3-oxoprop-1-en-1-yl)oxy)phenyl)adamantane-1-carboxylate (Compound 8b) Take a 100mL single-mouth bottle, add 0.1g of 3-(4-(3-((benzyloxy)carbonyl)adamantan-1-yl)phenoxy)acrylic acid, 0.13g of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDC), 0.09g of 1-hydroxybenzotriazole (HOBt), 120μL of N,N-Diisopropylethylamine (DIPEA) at room temperature, add a certain amount of dichloromethane and ultra-dry tetrahydrofuran to dissolve it, transfer the entire reaction system to a magnetic stirrer, protect with nitrogen, and react at room temperature for 1 hour. After 1 hour, add 80μL of o-anisidine and continue to react for 12 hours. After the reaction, the product was washed several times with saturated NaCl solution and dichloromethane, dried with anhydrous magnesium sulfate for one hour, filtered after drying, concentrated, and eluted by column chromatography using an eluent (V dichloromethane: V ethyl acetate = 30:1). After elution, the solvent was removed and the residue was placed in a vacuum drying oven and dried overnight to obtain a white solid. Yield: 32%. 1H NMR (400MHz, Chloroform-d) δ7.85(d,J=11.7Hz,1H),7.40(d,J=18.5Hz,2H),7.33(d,J=8.7Hz,7H),7.14(s,1H),7.01(d,J=8.6Hz,2H),6.85( d,J=8.8Hz,2H),5.66(d,J=11.7Hz,1H),5.12(s,2H),3.78(s,3H),2.2 4(s,2H),2.04(s,2H),1.96(d,J=6.8Hz,4H),1.88(s,4H),1.74(s,2H).

[0016] (3) Synthesis of benzyl-3-(4-(E)-3-((3-methoxyphenyl)amino)-3-oxoprop-1-en-1-yl)oxy)phenyl)adamantane-1-carboxylate (Compound 8c) Take a 100mL single-mouth bottle, add 0.1g of 3-(4-(3-((benzyloxy)carbonyl)adamantan-1-yl)phenoxy)acrylic acid, 0.13g of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDC), 0.09g of 1-hydroxybenzotriazole (HOBt), 120μL of N,N-Diisopropylethylamine (DIPEA) at room temperature, add a certain amount of dichloromethane and ultra-dry tetrahydrofuran to dissolve it, transfer the entire reaction system to a magnetic stirrer, protect with nitrogen, and react at room temperature for 1 hour. After 1 hour, add 80μL of m-methoxyaniline and continue to react for 12 hours. After the reaction, the product was washed several times with saturated NaCl solution and dichloromethane, dried with anhydrous magnesium sulfate for one hour, filtered after drying, concentrated, and eluted by column chromatography using an eluent (V dichloromethane: V ethyl acetate = 30:1). After elution, the solvent was removed and the residue was placed in a vacuum drying oven and dried overnight to obtain a white solid. Yield: 32%. 1H NMR (400MHz, DMSO-d6) δ9.90 (s, 1H), 7.73 (d, J = 12.0Hz, 1H), 7.42 (d, J = 8.7Hz, 2H), 7.35 (p, J = 7.3Hz, 6H), 7.16 (dd, J = 17.3, 8.4Hz, 4H ),6.61(d,J=9.7Hz,1H),5.81(d,J=12.0Hz,1H),5.10(s,2H),3.72(s,3H),2.17(s,2H),1.95(s,2H),1.90–1.80(m,8H),1.69(s,2H).

[0017] (4) Synthesis of benzyl-3-(4-(E)-3-((4-methoxyphenyl)amino)-3-oxoprop-1-en-1-yl)oxy)phenyl)adamantane-1-carboxylate (Compound 8d) Take a 100mL single-mouth bottle, add 0.1g of 3-(4-(3-((benzyloxy)carbonyl)adamantan-1-yl)phenoxy)acrylic acid, 0.13g of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDC), 0.09g of 1-hydroxybenzotriazole (HOBt), 120μL of N,N-Diisopropylethylamine (DIPEA) at room temperature, add a certain amount of dichloromethane and ultra-dry tetrahydrofuran to dissolve it, transfer the entire reaction system to a magnetic stirrer, protect with nitrogen, and react at room temperature for 1 hour. After 1 hour, add 0.09g of p-anisidine and continue to react for 12 hours. After the reaction, the product was washed several times with saturated NaCl solution and dichloromethane, dried with anhydrous magnesium sulfate for one hour, filtered after drying, concentrated, and eluted by column chromatography using an eluent (V dichloromethane: V ethyl acetate = 5:1). After elution, the solvent was removed and the residue was dried in a vacuum drying oven overnight to obtain a white solid. Yield: 32%. 1H NMR(400MHz, DMSO-d6)δ9.79(s,1H),7.70(d,J=11.9Hz,1H),7.53(d,J=8.7Hz,2H),7.44–7.28(m,7H),7.13(d,J=8.5Hz,2H),6.87( d,J=8.9Hz,2H),5.81–5.76(m,1H),5.10(s,2H),3.71(s,3H),2.17(s,2H),1.94(s,2H),1.84(dd,J=21.0,11.3Hz,8H),1.69(s,2H).

[0018] (5) Benzyl-3-(4-((3-((3-(methoxycarbonyl)phenyl)amino)-3-oxoprop-1-en-1-yl)oxy)phenyl)adamantane-1-carboxylate (Compound 8e) Take a 100mL single-mouth bottle, add 0.1g of 3-(4-(3-((benzyloxy)carbonyl)adamantan-1-yl)phenoxy)acrylic acid, 0.13g of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDC), 0.09g of 1-hydroxybenzotriazole (HOBt), 120μL of N,N-Diisopropylethylamine (DIPEA) at room temperature, add a certain amount of dichloromethane and ultra-dry tetrahydrofuran to dissolve it, transfer the entire reaction system to a magnetic stirrer, protect with nitrogen, and react at room temperature for 1 hour. After 1 hour, add 0.14g of methyl 2-aminobenzoate and continue to react for 12 hours. After the reaction, the product was washed several times with saturated NaCl solution and dichloromethane, dried with anhydrous magnesium sulfate for one hour, filtered after drying, concentrated, and eluted by column chromatography using an eluent (V petroleum ether: V ethyl acetate = 5:1). After elution, the solvent was removed and the residue was dried in a vacuum drying oven overnight to obtain a white solid. Yield: 35%. 1H NMR(400MHz,Chloroform-d)δ8.06(s,1H),7.90(d,J=11.7Hz,2H),7.77(d,J=7.5Hz,1H),7.34(s,8H),7.03(d,J=8.5Hz,2H), 5.66(d,J=11.7Hz,1H),5.12(s,2H),3.90(s,3H),2.24(s,2H),2.04(s,2H),1.96(d,J=8.4Hz,4H),1.88(s,4H),1.73(s,2H).

[0019] (5) Benzyl-3-(4-((3-((2-(methoxycarbonyl)phenyl)amino)-3-oxoprop-1-en-1-yl)oxy)phenyl)adamantane-1-carboxylate (Compound 8f) Take a 100mL single-mouth bottle, add 0.1g of 3-(4-(3-((benzyloxy)carbonyl)adamantan-1-yl)phenoxy)acrylic acid, 0.13g of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDC), 0.09g of 1-hydroxybenzotriazole (HOBt), 120μL of N,N-Diisopropylethylamine (DIPEA) at room temperature, add a certain amount of dichloromethane and ultra-dry tetrahydrofuran to dissolve it, transfer the entire reaction system to a magnetic stirrer, protect with nitrogen, and react at room temperature for 1 hour. After 1 hour, add 0.14g of methyl 2-aminobenzoate and continue to react for 12 hours. After the reaction, the product was washed several times with saturated NaCl solution and dichloromethane, dried with anhydrous magnesium sulfate for one hour, filtered after drying, concentrated, and eluted by column chromatography using an eluent (V petroleum ether: V ethyl acetate = 5:1). After elution, the solvent was removed and the residue was placed in a vacuum drying oven and dried overnight to obtain a white solid. Yield: 28%. 1H NMR (400MHz, DMSO-d6) δ10.15(s,1H),8.31(s,1H),7.89(d,J=8.1Hz,1H),7.78(d,J=12.0Hz,1H),7.62(d,J=7.7Hz,1H),7.35(s,8H),7.13(d ,J=8.6Hz,2H),5.83(d,J=12.0Hz,1H),5.09(s,2H),3.84(s,3H),2.15 (s,2H),1.94(s,2H),1.86(s,4H),1.81(d,J=11.7Hz,4H),1.66(s,2H).

[0020] Example 3: In vitro cytotoxicity evaluation results The present invention conducted CCK-8 experimental tests on target compounds 8a-8f, expressed HeLa, A549, HepG2 and L929, and tested target compounds 8a-8f and 5-fluorouracil at a concentration of 20 μM. As can be seen from the figure, 8c containing an amine compound structure with a methoxyl group grafted at the end of the benzene ring of an adamantane structure has the best inhibitory effect on HeLa cells and HepG2 cells, with inhibition rates of 66.88% and 70.14%, respectively, and 5-fluorouracil has inhibition rates of 58.85% and 46.66% on HeLa cells and HepG2 cells, respectively. 8a containing an amine compound structure with no substitution grafted at the end of the benzene ring of an adamantane structure has the best inhibitory effect on A549 cells, with an inhibition rate of 48.33%, while the inhibition rate of 5-fluorouracil on A549 cells is 41.13%.

[0021] The present invention calculates the IC50 values ​​of compounds 8a-8f at 40 μM, 20 μM, 10 μM, and 5 μM concentrations for HeLa, A549, and HepG2 cells, wherein compounds 8a and 8c have lower IC50 values ​​for HepG2 cells, 8c has lower IC50 value for HeLa cells, and 8a has lower IC50 value for A549 cells. For amine compounds with different groups connected at the 3rd position at the end of the benzene ring, the efficacy of compounds containing methoxy groups in HeLa cells and HepG2 cells is significantly better than that of compounds containing methyl benzoate. The difference is that in A549 cells, the efficacy of compounds without substituents is better than that of compounds containing methyl benzoate, and all target compounds are non-toxic to L929 cells. Fig.11 IC50 values ​​of target compounds 8a-8f on different cells.

Claims

1. A novel small drug molecule containing an adamantane structure (1), characterized in that: The molecular chemical structure is as follows, 2. R is R1, R2, and R3 are H, OCH3, or HCOOCH3 at corresponding positions.

3. The adamantane structure derivative according to claim 1, wherein: Target product:

4. The adamantane structure derivative according to claim 1, wherein: Synthesis route:

5. As claimed in claim 1, the use of the adamantane structure derivative formula (1) having multiple biological activities such as antiviral, antibacterial, and antitumor.

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