Benzenesulfonamide derivative as well as preparation method and application thereof

Abstract

The invention discloses a benzenesulfonamide derivative, the structural formula of which is as follows: # imgabs0 #. The synthesis process is simple, the conditions of no water, no oxygen and the like are not needed, and individual reactions are ingeniously combined with microwave conditions, so that the yield of partial reactions is high, and the consumed time is short. Activity research shows that the compound has the activity of inhibiting proliferation and migration of tumor cells, and can obviously inhibit growth of tumors in tumor-bearing mice at the dosage of 10mg / kg; in addition, the compound also has the activity of reducing blood sugar and blood fat. Therefore, the compound has a good application prospect.

Description

Technical Field

[0001] The invention belongs to the technical field of compound synthesis and pharmaceutical application, in particular to a benzenesulfonamide derivative and a preparation method and application thereof. Background Art

[0002] Benzenesulfonamide compounds refer to a class of compounds containing a benzene ring and a sulfonamide functional group. In 1932, German scientists discovered that Prontosil had an inhibitory effect on Streptococcus pneumoniae, of which sulfonamide was the effective active part, thus opening the door to sulfonamide antibiotics. With the emergence of the phenomenon of antibiotic abuse, the problem of drug resistance has gradually become prominent, resulting in increased use of antibiotics, reduced efficacy, and some other adverse reactions, which has restricted the development and application of antibiotics. In the process of further in-depth research on the structure-activity relationship and mechanism of action of sulfonamide compounds, it was found that in addition to antibacterial activity, this type of compound also has biological activities such as anti-tumor, anti-depression, anti-inflammatory, anti-diabetes, and inhibition of carbonic anhydrase. Among them, the anti-tumor activity of sulfonamide compounds has attracted much attention.

[0003] Studies have shown that sulfonamide compounds can reduce cancer cell proliferation and induce cell apoptosis. For example, Dubois et al. found that sulfonamide carbonic anhydrase IX inhibitors combined with radiotherapy can effectively inhibit HT-29 cell activity. Hao Mingyue et al. synthesized a new sulfonamide tumor inhibitor, explored the effect of the new sulfonamide tumor inhibitor on the apoptosis of cervical cancer Hela cells, and further studied its mechanism of action. Yin Luo et al. synthesized and evaluated the anti-microtubule polymerization activity and in vitro cell activity of new sulfonamide derivatives with cinnamic acid acyl groups. This series of compounds showed effective microtubule polymerization inhibition activity and anti-proliferation activity against MCF-7 cells. Owa et al. reported a new class of sulfonamide anti-tumor compounds with indigo as the lead compound. These compounds can reversibly bind to the colchicine binding site on β-tubulin. Sun Hua et al. reported that 1,4-naphthoquinone derivatives have good activity in inhibiting human colon cancer cells, and their derivatives have sulfonamide groups in their structures.

[0004] Through searching, we found the following patent publications related to the patent application of this invention:

[0005] 1. A 1,4-naphthoquinone derivative and its preparation method and application, application number 201711376664.8, authorized in January 2020. The main research content disclosed in this patent is a class of 1,4-naphthoquinone derivatives, some of which contain sulfonamide fragments, and these compounds have anti-colon cancer and α-glucosidase inhibitory activity. The defect of this patent is that the activity and solubility of the compounds need to be further improved. This patent is the basis of our research group's preliminary research, and its protection scope does not cover the content of this patent. The content of this patent is a new structure with more ideal activity and solubility that was discovered after the synthesis and activity evaluation of a large number of compounds.

[0006] 2. A sulfonamide derivative and its preparation method and application, application number 202010688883.5. The main research content disclosed in this patent is a class of sulfonamide derivative compounds with anti-colon cancer and α-glucosidase inhibitory activity. This patent is also the preliminary research basis of our research topic. It is based on patent 1 (201711376664.8) to structurally modify the 1,4-naphthoquinone parent nucleus to obtain a series of derivatives with new structures. The defect of this patent is that the synthesis yield of the compound is low and the activity of the compound needs to be further improved. The content of this patent is a new structure discovered after a lot of experimental exploration, in addition to retaining the benzenesulfonamide functional group, and structurally changing multiple pharmacophores. The protection scope of patent 2 does not cover the content of this patent.

[0007] By comparison, the present invention is essentially different from the above-mentioned disclosed patents. First, there is a structural difference, and the present patent is not within the protection scope of the above-mentioned patents. Second, the above-mentioned two patents have problems such as insufficient activity, low synthesis yield, and poor water solubility. Therefore, the above-mentioned patent issues are the motivation for the technical solution of the present invention. Summary of the invention

[0008] The purpose of the present invention is to overcome the problems existing in the prior art and provide a benzenesulfonamide derivative and a preparation method and application thereof.

[0009] The technical solution adopted by the present invention to solve the technical problem is:

[0010] A benzenesulfonamide derivative, the structural formula of the derivative is as follows:

[0011]

[0012] Among them, RNR' is and its hydrochloride, wherein n1 is equal to or not equal to n2, n1=a natural number from 2 to 4, n2=a natural number from 2 to 4, and R1 is an alkyl group;

[0013] Alternatively, RNR' is and its hydrochloride, wherein n1 is equal to or not equal to n2, n1=a natural number from 2 to 4, n2=a natural number from 2 to 4, and m=a natural number from 1 to 3;

[0014] Alternatively, RNR' is

[0015] Ar Wherein R=H or halogen; or, Ar is Wherein R=H, methyl, trifluoromethyl, halogen; or, Ar is Wherein R=H, methyl, trifluoromethyl, halogen; or, Ar is

[0016] Furthermore, the R1 is methyl, ethyl, propyl or butyl.

[0017] The preparation method of the benzenesulfonamide derivatives as described above has the following reaction route:

[0018]

[0019] Among them, RNHR1 is

[0020] or, Among them, RNHR1 is

[0021] or, Among them, RNHR1 is

[0022] or,

[0023] or,

[0024] or,

[0025] or,

[0026] Further, the following steps are included:

[0027] The fatty amine raw material RNHR1 reacts with acetylaminobenzenesulfonyl chloride to obtain VIII, and the acetyl protecting group of the amine group is removed under alkaline conditions to obtain IX, and IX reacts with a halogenated or trifluoromethanesulfonate (TfO) substituted aromatic group to obtain a benzenesulfonamide derivative. In some of the steps, microwave reaction conditions can improve the yield and shorten the reaction time, and then the product I is dissolved in methanol and hydrogen chloride gas is introduced, and the precipitated solid is the corresponding hydrochloride of the product I.

[0028] The use of the benzenesulfonamide derivatives as described above in the preparation of drugs for treating human colon cancer.

[0029] The use of the benzenesulfonamide derivatives as described above in the preparation of anti-tumor drugs.

[0030] The use of the benzenesulfonamide derivatives as described above in the preparation of drugs for treating glucose and lipid metabolism disorders.

[0031] The use of the benzenesulfonamide derivatives as described above in the preparation of drugs for treating blood lipids.

[0032] The use of the benzenesulfonamide derivatives as described above in the preparation of therapeutic hypoglycemic drugs.

[0033] The beneficial effects achieved by the present invention are:

[0034] 1. The benzenesulfonamide derivatives of the present invention have good anti-human colon cancer and liver cancer cell activities. The benzenesulfonamide derivatives of the present invention also have hypoglycemic and hypolipidemic activities, that is, they can alleviate the comprehensive disorder of sugar and lipid metabolism.

[0035] 2. The reaction of the method of the present invention does not require anhydrous and oxygen-free operation, is easy to operate, and the raw materials and reagents are cheap and easily available, which is suitable for large-scale production and development.

[0036] 3. The key reaction steps in the method of the present invention cleverly use microwave reaction conditions to increase the yield and shorten the reaction time.

[0037] 4. Some compounds of the present invention are prepared into hydrochloride salts, which increase water solubility, improve lipid-water partition coefficient, and further enhance drugability.

[0038] 5. Activity studies have shown that this type of compound has the activity of inhibiting tumor cell proliferation and migration, and can significantly inhibit tumor growth in tumor-bearing mice at a dose of 10 mg / kg; in addition, this type of compound also has the activity of lowering blood sugar and blood lipids. Therefore, it shows that this compound has good application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0039] Figure 1 is the H NMR spectrum of compound 1 in deuterated methanol;

[0040] Figure 2 is the H NMR spectrum of compound 2 in deuterated dichloromethane;

[0041] Figure 3 is the H NMR spectrum of compound 3 in deuterated dimethyl sulfoxide;

[0042] Figure 4is the H NMR spectrum of compound 4 in deuterated chloroform;

[0043] Figure 5 is the H NMR spectrum of compound 5 in deuterated methanol;

[0044] Figure 6 is the H NMR spectrum of compound 6 in deuterated chloroform;

[0045] Figure 7 is the H NMR spectrum of compound 7 in deuterated acetone;

[0046] Figure 8 is the H NMR spectrum of compound 8 in deuterated chloroform;

[0047] Fig. 9 is the H NMR spectrum of compound 9 in deuterated chloroform;

[0048] Fig.10 is the H NMR spectrum of compound 10 in deuterated dimethyl sulfoxide;

[0049] Fig.11 is the H NMR spectrum of compound 11 in deuterated chloroform;

[0050] Fig.12 is the H NMR spectrum of compound 12 in deuterated dimethyl sulfoxide;

[0051] Fig.13 is the H NMR spectrum of compound 13 in deuterated dimethyl sulfoxide;

[0052] Fig.14 is the H NMR spectrum of compound 14 in deuterated acetone;

[0053] Fig.15 is the H NMR spectrum of compound 15 in deuterated chloroform;

[0054] Fig.16 is the H NMR spectrum of compound 16 in deuterated chloroform;

[0055] Fig.17 is the H NMR spectrum of compound 17 in deuterated chloroform;

[0056] Fig.18 is the H NMR spectrum of compound 18 in deuterated chloroform;

[0057] Fig.19 is the H NMR spectrum of compound 19 in deuterated chloroform;

[0058] Fig. 20 The statistical results of the effect of compound 3a of the present invention on the migration of SW480 and HCT116 cells are shown in Table 1. Compared with the model group, ***P<0.01, **P<0.05, *P<0.01;

[0059] Fig.21 The effect of compounds 3a and 3 of the present invention on cellular glucose consumption; compared with the model group, ***P<0.01;

[0060] Fig. 22 The effects of compounds 3a and 3 of the present invention on the cellular lipid content; compared with the model group, **P<0.05, *P<0.01.

[0061] Fig.23 The graph is a curve showing the effect of compound 3a of the present invention on the tumor volume change of tumor-bearing mice; compared with the model group, ***P<0.01;

[0062] Fig.24 The graph shows the inhibitory effect of different doses of compound 3a of the present invention on the tumor appearance of tumor-bearing mice. DETAILED DESCRIPTION

[0063] For a better understanding of the present invention, the present invention is further described in detail below in conjunction with embodiments, but the scope of protection claimed by the present invention is not limited to the scope represented by the embodiments.

[0064] The raw materials used in the present invention, unless otherwise specified, are all conventional commercially available products. The methods used in the present invention, unless otherwise specified, are all conventional methods in the art. The quality of each substance used in the present invention is the quality for conventional use.

[0065] A benzenesulfonamide derivative, the structural formula of the derivative is as follows:

[0066]

[0067] Among them, RNR' is and its hydrochloride, wherein n1 is equal to or not equal to n2, n1=a natural number from 2 to 4, n2=a natural number from 2 to 4, and R1 is an alkyl group;

[0068] Alternatively, RNR' is and its hydrochloride, wherein n1 is equal to or not equal to n2, n1=a natural number from 2 to 4, n2=a natural number from 2 to 4, and m=a natural number from 1 to 3;

[0069] Alternatively, RNR' is

[0070] Ar Wherein R=H or halogen; or, Ar is Wherein R=H, methyl, trifluoromethyl, halogen; or, Ar is Wherein R=H, methyl, trifluoromethyl, halogen; or, Ar is

[0071] Preferably, the R1 is methyl, ethyl, propyl or butyl.

[0072] The preparation method of the benzenesulfonamide derivatives as described above has the following reaction route:

[0073]

[0074] Among them, RNHR1 is

[0075] or, Among them, RNHR1 is

[0076] or, Among them, RNHR1 is

[0077] or,

[0078] or,

[0079] or,

[0080] or,

[0081] Preferably, the method comprises the following steps:

[0082] The fatty amine raw material RNHR1 reacts with acetylaminobenzenesulfonyl chloride to obtain VIII, and the acetyl protecting group of the amine group is removed under alkaline conditions to obtain IX, and IX reacts with a halogenated or trifluoromethanesulfonate (TfO) substituted aromatic group to obtain a benzenesulfonamide derivative. In some of the steps, microwave reaction conditions can improve the yield and shorten the reaction time, and then the product I is dissolved in methanol and hydrogen chloride gas is introduced, and the precipitated solid is the corresponding hydrochloride of the product I.

[0083] The use of the benzenesulfonamide derivatives as described above in the preparation of drugs for treating human colon cancer.

[0084] The use of the benzenesulfonamide derivatives as described above in the preparation of anti-tumor drugs.

[0085] The use of the benzenesulfonamide derivatives as described above in the preparation of drugs for treating glucose and lipid metabolism disorders.

[0086] The use of the benzenesulfonamide derivatives as described above in the preparation of drugs for treating blood lipids.

[0087] The use of the benzenesulfonamide derivatives as described above in the preparation of therapeutic hypoglycemic drugs.

[0088] Specifically, the relevant preparation and detection are as follows:

[0089] Example 1VIII Synthesis of Side Chain Intermediates

[0090] The specific reaction route is as follows:

[0091]

[0092] General synthesis method of intermediates VIII-a to VIII-i:

[0093] Under ice bath conditions, 4-acetamidobenzenesulfonyl chloride (1.50mmol) was dissolved in 10mL of anhydrous dichloromethane, and different amine raw materials (1.00mmol) were added under stirring, and then triethylamine (2.00mmol) was added, and the reaction was carried out at room temperature for 5 to 10 hours. After the reaction was completed by thin layer chromatography monitoring, 50mL of water was added, and then 50mL of dichloromethane was used for extraction three times, and the organic phases were combined and dried over anhydrous Na2SO4, and the desiccant was filtered to remove the solvent, and the crude product was separated and purified by silica gel column to obtain intermediates VIII-a to VIII-i, as follows:

[0094] (1) Synthesis of (S)-N-(4-((3-(dimethylamino)pyrrolidin-1-yl)sulfonyl)phenyl)acetamide (VIII-a)

[0095] The amino raw material is (S)-3-dimethylaminopyrrolidine dihydrochloride. According to the above general synthesis method, white solid VIII-a is obtained with a yield of 91.46%. 1 H-NMR (400MHz, CDCl3) δ7.83 (s, 1H), 7.76 (d, J = 8.4Hz, 2H), 7.70 (d, J = 8.8Hz, 2H), 3.48-3.52 (m, 1H), 3.34-3.40 (m, 1H), 3. 34-3.35(m,1H),2.95(t,J=9.0Hz,1H),2.59-2.67(m,1H),2.22(s,3H),2.17(s,6H),1.99-2.03(m,1H),1.60-1.67(m,1H).

[0096] (2) Synthesis of tert-butyl 4-(4-acetylaminophenyl)sulfonyl)piperazine-1-carboxylate (VIII-b)

[0097] The amino raw material is mono-Boc piperazine. According to the above general synthesis method, white solid VIII-b is obtained with a yield of 95.64%. 1 H-NMR (400MHz, CDCl3) δ7.69 (s, 4H), 7.54 (s, 1H), 3.50 (t, J = 5.0Hz, 4H), 2.95 (t, J = 4.8Hz, 4H), 2.23 (s, 3H), 1.41 (s, 9H).

[0098] (3) Synthesis of N-(4-((4-methylpiperazin-1-yl)sulfonyl)phenyl)acetamide (VIII-c)

[0099] The amino raw material is N-methylpiperazine. According to the above general synthesis method, white solid VIII-c is obtained with a yield of 96.86%. 1 H-NMR (400MHz, CDCl3) δ7.71(d,J=9.2Hz,2H),7.68(d,J=9.2Hz,2H),7.35(s,1H),3.03(s,4H),2.47(t,J=5.0Hz,4H),2.64(s,3H),2.23(s,3H).

[0100] (4) Synthesis of N-(4-((3,5-dimethylpiperidin-1-yl)sulfonyl)phenyl)acetamide (VIII-d)

[0101] The amino raw material is 3,5-dimethylpiperidine. According to the above general synthesis method, white solid VIII-d is obtained with a yield of 92.84%. 1 H-NMR (400MHz, CDCl3) δ7.71(d,J=8.8Hz,2H),7.67(d,J=9.2Hz,2H),7.41(s,1H),3.71(d,J=8.8Hz, 2H), 2.23 (s, 3H), 1.65-1.78 (m, 4H), 0.98 (d, J = 6.8Hz, 1H), 0.84 (d, J = 6.0Hz, 6H), 0.41-0.50 (m, 1H).

[0102] (5) Synthesis of N-(4-(aza-1-ylsulfonyl)phenyl)acetamide (VIII-e)

[0103] The amino raw material is cycloheximide. According to the above general synthesis method, white solid VIII-e is obtained with a yield of 90.85%.

[0104] (6) Synthesis of N-(4-((4-methyl-1,4-diazepin-1-yl)sulfonyl)phenyl)acetamide (VIII-f)

[0105] The amino raw material is N-methyl homopiperazine. According to the above general synthesis method, white solid VIII-f is obtained with a yield of 86.31%. 1 H-NMR (400MHz, MeOD) δ7.82 (d, J = 8.8 Hz, 2H), 7.77 (d, J = 9.2 Hz, 2H), 3.58 (s, 2H), 3.37-3.43 (m, 6H), 2.91 (s, 3H), 2.16 (s, 3H), 2.12-2.15 (m, 2H).

[0106] (7) Synthesis of N-(4-(azo-1-ylsulfonyl)phenyl)acetamide (VIII-g)

[0107] The amino raw material is cyclooctane imine. According to the above general synthesis method, white solid VIII-g is obtained with a yield of 89.48%. 1 H-NMR (400MHz, CDCl3) δ7.73(d,J=8.4Hz,2H),7.65(d,J=8.4Hz,2H),7.46(s,1H),3.05(t,J=5.0Hz,4H),2.22(s,3H),1.72(s,8H),1.63(s,4H).

[0108] (8) Synthesis of N-(4-(N-(quinolin-8-yl)sulfamoyl)phenyl)acetamide (VIII-h)

[0109] The amino raw material is 8-aminoquinoline. According to the above general synthesis method, white solid VIII-h is obtained with a yield of 79.82%. 1 H-NMR (400MHz, CDCl3) δ9.21(s,1H),8.76(dd,J=1.2,1.6Hz,1H),8.09(dd,J=1.6,1.2Hz,1H),7.85( d,J=8.8Hz,2H),7.81(dd,J=1.6,2.0Hz,1H),7.50(d,J=8.4Hz,2H),7.40-7.47(m,3H),2.14(s,3H).

[0110] (9) Synthesis of N-(4-((4-phenylpiperazin-1-yl)sulfonyl)phenyl)acetamide (VIII-i)

[0111] The amino raw material is N-phenylpiperazine. According to the above general synthesis method, white solid VIII-i is obtained with a yield of 91.36%. 1H-NMR (400MHz, DMSO-d6) δ10.39(s,1H),7.84(d,J=8.8Hz,2H),7.70(d,J=8.8Hz,2H),7.19(t,J=7.8Hz,2H ), 6.90 (d, J = 8.4Hz, 2H), 6.79 (t, J = 7.4Hz, 1H), 3.19 (t, J = 4.6Hz, 4H), 2.98 (t, J = 4.6Hz, 4H), 2.09 (s, 3H).

[0112] (10) Synthesis of N-(4-((4-benzylpiperazin-1-yl)sulfonyl)phenyl)acetamide (VIII-j)

[0113] Dissolve mono-Boc piperazine (1117 mg, 6 mmol) in 15 mL acetonitrile, add anhydrous Na2CO3 (795 mg, 7.5 mmol), add benzyl bromide (855 mg, 5 mmol) while stirring, and react at room temperature for 5 h. After the reaction is complete, add 50 mL of water, extract three times with 50 mL of dichloromethane, combine the organic phases, add anhydrous Na2SO4 to dry the organic phase, filter to remove the desiccant, and evaporate the solvent. Use silica gel column chromatography to separate and purify to obtain the intermediate product. The intermediate is dissolved in 5 mL of dichloromethane, and 2 mL of trifluoroacetic acid is added dropwise under ice bath, and reacted for 5 h. After the reaction is completed, the excess trifluoroacetic acid and solvent are evaporated under reduced pressure. Subsequently, it was dissolved with 5 mL of dichloromethane, and 4-acetylbenzenesulfonyl chloride (882 mg, 3.78 mmol) and triethylamine (1.1 mL, 7.56 mmol) were added under stirring in an ice bath. The reaction was changed to room temperature. After the reaction was complete, 50 mL of water was added, and the mixture was extracted three times with 50 mL of dichloromethane. The organic phases were combined, and anhydrous Na2SO4 was added to dry the organic phase. The desiccant was filtered out, and the solvent was evaporated. The crude product was separated and purified by column chromatography to obtain compound VIII-j with a yield of 76.81% as a white solid. 1 H-NMR (400MHz, CDCl3) δ7.66-7.71(m,4H),7.38(s,1H),7.22-7.28(m,4H),3.48(s,2H),3.01(s,4H),2.52(t,J=4.8Hz,4H),2.23(s,3H).

[0114] (11) Synthesis of N-(4-((4-cinnamylpiperazin-1-yl)sulfonyl)phenyl)acetamide (VIII-k)

[0115] Using cinnamyl bromide as raw material, the synthesis method is the same as the synthesis method of compound VIII-j to obtain white solid VIII-k with a yield of 79.66%. 1H-NMR (400MHz, DMSO-d6) δ10.40(s,1H),7.83(d,J=8.8Hz,2H),7.66(d,J=8.8Hz,2H),7.40(d,J=7.2Hz,2H),7.30(t,J=7.4Hz,2 H),7.22(t,J=7.2Hz,1H),6.50(d,J=16Hz,1H),6.15-6.22(m,1H),3.08(d,J=5.4Hz,2H),2.88(s,4H),2.46(s,4H),2.10(s,3H).

[0116] (12) Synthesis of N-(4-((3-(dimethylamino)azetidin-1-yl)sulfonyl)phenyl)acetamide (VIII-1)

[0117] Dissolve 1-tert-butyloxycarbonyl-3-azetidinone (200 mg, 1.17 mmol) in 5 mL of anhydrous dichloromethane, add dimethylamine hydrochloride (143 mg, 1.75 mmol) and triethylamine (0.4 mL, 2.95 mmol) while stirring, stir at room temperature for 3-5 hours, then add sodium cyanoborohydride (1.75 mmol), and react at room temperature for 9.5 hours. After the reaction is complete, pour the reaction solution into 50 mL of water, extract three times with 50 mL of dichloromethane: methanol = 10:1 (volume ratio), combine the organic phases, add anhydrous Na2SO4 to dry, filter to remove the desiccant, and evaporate the solvent. Then dissolve it with 5 mL of anhydrous dichloromethane, drop 1 mL of trifluoroacetic acid, and stir for 3 hours. After the reaction is completed, remove the excess trifluoroacetic acid and solvent under reduced pressure. Then, it was dissolved with 5 mL of dichloromethane, and 4-acetylbenzenesulfonyl chloride (409 mg, 1.75 mmol) and triethylamine (0.5 mL, 3.51 mmol) were added under stirring in an ice bath, and the reaction was changed to room temperature. After the reaction was complete, 50 mL of water was added, and the mixture was extracted three times with 50 mL of dichloromethane. After the organic phases were combined, anhydrous Na2SO4 was added for drying, the desiccant was removed by filtration, and the solvent was evaporated. The crude product was separated and purified by silica gel column chromatography to obtain compound VIII-1 with a yield of 64.26% as a white solid. 1 H-NMR (400MHz, CDCl3) δ7.78(d,J=8.6Hz,2H),7.73(d,J=8.4Hz,2H),7.47(s,1H),3.81( t, J=7.6Hz, 2H), 3.55 (d, J=7.2Hz, 2H), 3.03 (d, J=6.6Hz, 1H), 2.34 (s, 3H), 2.03 (s, 6H).

[0118] Example 2 Synthesis of Compound 1

[0119] The specific synthetic route is as follows:

[0120]

[0121] The above side chain intermediate VIII-1 (1.00mmol) was dissolved in 10mL methanol, and 2mL sodium hydroxide aqueous solution (5M) was added, and heated to reflux. After the reaction was complete, it was extracted three times with 50mL dichloromethane, and the organic phases were combined and dried with anhydrous Na2SO4. The desiccant was filtered to remove the desiccant, and the solvent was evaporated to obtain a crude product. The above crude product (51mg, 0.20mmol) was dissolved in 2mL ethanol, and 2,3-dichloro-1,4-naphthoquinone (136mg, 0.60mmol) was added. A microwave reactor was used to react at 70℃ for 4h, and the solvent was evaporated. The product 1 was separated and purified by column chromatography. Yield: 89%. The structural parameters of compound 1 are as follows: 1 H-NMR (400MHz, MeOD) δ8.15(t,J=7.8Hz,2H),7.85(s,2H),7.82(d,J=8.8Hz,2H),7.28(d,J=8.4Hz,2H),4.01-4.06(m,5H),2.82(s,6H); 13 C-NMR (100MHz, CDCl3) δ179.7,177.7,144.5,142.2,134.5,133.4,132.0,13 0.6,128.6,126.6,126.4,122.2,119.6,54.2,52.0,39.8.HRMS(ESI-TOF)m / z calcd.forC 21 H 21 ClN3O4S[M+H] + :446.0941,found446.0941.

[0122] Compound 1 is dissolved in methanol, hydrogen chloride gas is introduced, and the solid precipitated in the system is the corresponding hydrochloride 1a.

[0123] Example 3 Synthesis of Compound 2

[0124]

[0125] The synthesis method of compound 2 is the same as the synthesis method of compound 1, and the side chain is selected as VIII-a (1.00 mmol), and the yield is 88%. The structural parameters of compound 2 are as follows: 1H-NMR (400MHz, CDCl3) δ8.21-8.23(m,1H),8.14-8.17(m,1H),7.80-7.83(m,3H),7.77(s,1H),7.71-7.76(m,1H),7.14(d,J=8.8Hz,2H),3.53-3 .57(m,1H),3.37-3.43(m,1H),3.26-3.32(m,1H),2.98(t,J=9.2Hz,1H) ,2.58-2.66(m,1H),2.17(s,6H),2.00-2.05(m,1H),1.64-1.72(m,1H); 13 C-NMR (100MHz, CDCl3) δ180.2,177.4,141.4,140.7,135.3,133.5,132.3,132.3,12 9.8,127.4,127.2,127.2,123.0,118.1,65.1,51.8,47.0,29.9.HRMS(ESI-TOF)m / z calcd.for C 22 H 23 ClN3O4S[M+H] + :446.1098,found446.1097.

[0126] Dissolve compound 2 in methanol, introduce hydrogen chloride gas, and the solid precipitated in the system is the corresponding hydrochloride 2a.

[0127] Example 4 Synthesis of Compound 3

[0128]

[0129] The synthesis method of compound 3 is the same as the synthesis method of compound 1, and the side chain is selected as VIII-c, and the yield is 84%. The structural parameters of compound 3 are as follows: 1 H-NMR (400MHz, DMSO-d6) δ9.62 (s, 1H), 8.07 (dd, J = 1.2, 1.2Hz, 2H), 7.87-7.91 (m, 1H), 7.83-7. 87(m,1H),7.64(d,J=8.4Hz,2H),7.29(d,J=8.8Hz,2H),2.89(s,4H),2.37(s,4H),2.16(s,3H); 13C-NMR (100MHz, DMSO-d6) δ180.4,177.5,144.3,143.1,135.2,134.1,132.2,131.0,129.1,128.2,127.1,126.7,122.7,119.6,53.9,46.2,45.7.

[0130] Compound 3 is dissolved in methanol, hydrogen chloride gas is introduced, and the solid precipitated in the system is the corresponding hydrochloride 3a.

[0131] Example 5 Synthesis of Compound 4

[0132]

[0133] The synthesis method of compound 4 is the same as the synthesis method of compound 1, and the side chain is selected as VIII-d, and the yield is 85%. The structural parameters of compound 4 are as follows: 1 H-NMR (400MHz, CDCl3) δ8.22(d,J=8.0Hz,1H),8.16(d,J=7.6Hz,1H),7.82(t,7.6Hz,1H),7.74(d,J=8.4Hz,2H),7.73(s,1H),7.69 (s,1H),7.14(d,J=8.8Hz,2H),3.75(d,J=7.6Hz,2H),1.75(m,4H),0.99(d,J=5.8Hz,1H),0.86(d,J=6.0Hz,6H),0.86-0.99(m,1H); 13 C-NMR (100MHz, CDCl3) δ180.2,177.4,141.3,140.8,135.3,132.4,132.3,129.8 ,128.1,127.4,127.2,123.0,118.1,52.9,41.4,30.9,19.0.HRMS(ESI-TOF)m / z calcd.for C 23 H 24 ClN2O4S[M+H] + :459.1145, found 4459.1142.

[0134] Example 6 Synthesis of Compound 5

[0135]

[0136] The synthesis method of compound 5 is the same as the synthesis method of compound 1 above, and the side chain is selected as VIII-f, and the yield is 95%; the structural parameters of compound 4 are as follows: 1H-NMR(400MHz,MeOD)δ8.12-8.16(m,2H),7.85-7.87(m,1H),7.81-7.83(m,1H),7.79(d,J=8 .4Hz,2H),7.24(d,J=8.8Hz,1H),3.61(s,2H),3.41-3.45(m,6H),2.94(s,3H),2.16(s,2H); 13 C-NMR (100MHz, MeOD) δ181.1,179.0,145.0,143.7,135.9,134.6,133.7,133.4,131.9,128. 6,127.9,127.7,124.0,120.0,59.1,56.3,48.3,47.6,45.0,44.6,25.9.HRMS(ESI-TOF)m / z calcd.for C 22 H 23 ClN3O4S[M+H] + :440.1098,found 440.1098.

[0137] Example 7 Synthesis of Compound 6

[0138]

[0139] The synthesis method of compound 6 is the same as the synthesis method of compound 1, and the side chain is selected as VIII-g, and the yield is 56%. The structural parameters of compound 6 are as follows: 1 H-NMR (400MHz, CDCl3) δ8.22(d,J=8.0Hz,1H),8.15(d,J=7.6Hz,1H),7.82(t,J=8.4Hz,2H),7.73(d,J= 7.6Hz,1H),7.68(s,1H),7.12(d,J=8.4Hz,2H),3.30(t,J=5.4Hz,4H),1.73-1.76(m,8H),1.62(s,4H); 13 C-NMR (100MHz, CDCl3) δ180.3,177.4,141.0,140.8,135.3,134.1,133.5,132.3,12 9.8,127.8,127.4,127.2,123.0,117.9,50.2,27.2,25.9,25.0.HRMS(ESI-TOF)m / z calcd.for C 24 H 26 ClN2O4S[M+H] + :473.1302,found473.1298.

[0140] Example 8 Synthesis of compound 7.

[0141]

[0142] The synthesis method of compound 7 is the same as the synthesis method of compound 1, and the side chain is selected as VIII-i, and the yield is 56%. The structural parameters of compound 7 are as follows: 1 H-NMR (400MHz, Acetone-d6) δ8.80(s,1H),8.13(t,J=7.6Hz,2H),7.89-7.93(m,1H),7.84-7.87(m,1H),7.79(d,J=8.8Hz,2H),7 .41(d,J=8.8Hz,2H),7.20-7.24(m,2H),6.95(d,J=8.0Hz,2H),6.81-6.84(m,1H),3.28(t,J=4.8Hz,4H),3.15(t,J=5.0Hz,4H); 13 C-NMR (100MHz, CDCl3) δ180.3,177.4,150.7,141.8,140.7,135.3,133.6,132.3,131.1,129 .9,129.3,128.5,127.5,127.3,122.8,121.0,118.6,117.0,48.2,46.2.HRMS(ESI-TOF)m / z calcd.for C 26 H 23 ClN3O4S[M+H] + :508.1098, found 508.1092.

[0143] Example 9 Synthesis of Compound 8

[0144]

[0145] The synthesis method of compound 8 is the same as the synthesis method of compound 1, and the side chain is selected as VIII-j. The yield is 53%. The structural parameters of compound 8 are as follows: 1H-NMR (400MHz, CDCl3) δ8.23(d,J=7.6Hz,1H),8.16(d,J=7.6Hz,1H),7.80-7.84(m,1H),7.75-7.77(m,1H),7.72(d,J=8.4Hz ,2H),7.69(s,1H),7.28-7.29(m,2H),7.24-7.26(m,1H),7.12(d,J=8.4Hz,2H,.350(s,2H),3.07(s,4H),2.52-2.55(m,4H); 13 C-NMR (100MHz, CDCl3) δ141.5,140.7,137.4,135.3,133.5,132.3,131.4,129.8,12 9.1,128.4,127.4,127.4,127.2,122.8,118.4,62.6,52.1,46.1.HRMS(ESI-TOF)m / z calcd.for C 27 H 25 ClN3O4S[M+H] + :522.1254, found 522.1250.

[0146] Example 10 Synthesis of Compound 9

[0147]

[0148] The synthesis method of compound 9 is the same as the synthesis method of compound 1, and the side chain is selected as VIII-k. The yield is 66%. The structural parameters of compound 9 are as follows: 1 H-NMR (400MHz, CDCl3) δ8.22(d,J=7.6Hz,1H),8.16(d,J=7.6Hz,1H),7.81(t,J=7.4Hz,1H),7.75(t,J=8.2Hz,3H),7.69(s,1H),732-7.36(m, 2H),7.26-7.30(m,2H),7.23(s,1H),7.12(d,J=8.4Hz,2H),6.52(d,J= 16.0Hz,1H),6.15-6.19(m,1H),3.16(s,2H),3.12(m,4H),2.61(m,4H); 13C-NMR (100MHz, CDCl3) δ180.2,177.4,141.6,140.6,136.5,135.3,133.5,132.3,131.1,129.8 ,128.6,128.4,127.8,127.4,127.2,126.4,122.7,118.5,60.6,52.1,46.0.HRMS(ESI-TOF)m / z calcd.for C 29 H 27 ClN3O4S[M+H] + :548.1411, found 548.1408.

[0149] Example 11 Synthesis of Compound 10

[0150] The specific preparation method is as follows:

[0151]

[0152] Select the side chain VIII-e (1.00mmol) and dissolve it in 10mL methanol, add 2mL sodium hydroxide aqueous solution (5M), heat to reflux, after the reaction is complete, extract it three times with 50mL dichloromethane, combine the organic phases, add anhydrous Na2SO4 to dry, filter to remove the desiccant, evaporate the solvent to obtain a crude product. The crude product (0.20mmol) is dissolved in 2mL THF / H2O (volume ratio of 3:1), add 2,4-dichloroquinazoline (0.30mmol) and sodium acetate (0.60mmol) under ice bath conditions, and react in a sealed tube at 100℃. After the reaction is complete, cool to room temperature, add 20mL of water, extract it three times with 50mL dichloromethane, combine the organic phases, dry it with anhydrous Na2SO4, filter and spin dry the solvent, and separate and purify it by column chromatography to obtain the final product 10. Yield: 78%; Product 10 structural parameters: 1 H-NMR (400MHz, DMSO-d6) δ10.43(s,1H),8.63(d,J=8.4Hz,1H),8.11(d,J=8.4Hz,2H),7.94(t,J=7.6Hz,1H),7.8 5(d,J=8.8Hz,2H),7.78(d,J=8.4Hz,2H),7.71(t,J=7.4Hz,1H),3.24(t,J=5.8Hz,4H),1.65(s,4H),1.52(s,4H); 13C-NMR(100MHz,DMSO-d6)δ159.6,156.2,151.5,142.6,134.9,134.4,128.0,127.5,127.4,124.1,122.5,114.3,48.2,29.0,26.8.HRMS(ESI-TOF)m / z calcd.for C 20 H 22 ClN4O2S[M+H] + :417.1152, found 417.1151.

[0153] Example 12 Synthesis of Compound 11

[0154]

[0155] The synthesis method of Example 11 is the same as that of the above-mentioned compound 10, and the side chain VIII-g (1.00 mmol) is selected. Yield: 45%; Structural parameters of compound 11: 1 H-NMR (400MHz, CDCl3) δ8.01(d,J=8.4Hz,1H),7.97(s,1H),7.92(d,J=8.8Hz,2H),7.88-7.89( m,2H),7.85(d,J=8.4Hz,2H),7.59-7.63(m.1H),3.10-3.13(m,4H),1.75(s,8H),1.67(s,4H); 13 C-NMR (100MHz, CDCl3) δ158.3,156.7,151.6,141.3,134.3,133.5,128.6,12 8.5,127.3,121.2,120.7,113.5,50.3,27.3,26.0,25.1.HRMS(ESI-TOF)m / z calcd.for C 22 H 26 ClN4O2S[M+H] + :445.1465,found445.1463.

[0156] Example 13 Synthesis of Compound 12

[0157] The synthetic route is as follows:

[0158]

[0159] Select side chain VIII-e (1.00mmol) and dissolve it in 10mL methanol, add 2mL sodium hydroxide solution (5M), heat and reflux, after the reaction is complete, extract it three times with 50mL dichloromethane, combine the organic phases and add anhydrous Na2SO4 to dry, filter to remove the desiccant, evaporate the solvent to obtain a crude product. Take the above crude product (0.50mmol) and dissolve it in 2mL DMF, add 2-chloroquinazoline-4 (3H) -one (1.0mmol), use a microwave reactor, microwave reaction at 150℃ for 3h, cool to room temperature, pour into 10mL ice water, collect the precipitated solid, separate and purify by column chromatography to obtain product 12. Yield: 90%; Product 12 structural parameters: 1 H-NMR (400MHz, DMSO-d6) δ10.95(s,1H),9.15(s,1H),8.00(d,J=8.0Hz,1H),7.96(d,J=8.8Hz,2H),7.69-7. 71(m,1H),7.47(d,J=8.0Hz,1H),7.29(t,J=7.4Hz,1H),3.19-3.35(m,4H),1.63(s,4H),1.49-1.51(m,4H); 13 C-NMR(100MHz,DMSO-d6)δ162.0,149.9,147.3,143.3,135.1,132.2,128.4,126.4,126.0,124.2,119.2,48.2,29.0,26.8.HRMS(ESI-TOF)m / z calcd.for C 20 H 23 N4O3S[M+H] + :399.1491, found 399.1486.

[0160] Example 14 Synthesis of Compound 13

[0161]

[0162] The synthesis method of Example 14 is the same as that of the above-mentioned compound 12, and the side chain VIII-g (1.00 mmol) is selected. Yield: 50%; Structural parameters of compound 13: 1H-NMR (400MHz, DMSO-d6) δ10.95(s,1H),9.15(s,1H),7.99(d,J=8.4Hz,2H),7.98(d,J=8.8Hz,2H),7.75(d,J=8.8Hz,2H ),7.70(d,J=7.2Hz,1H),7.47(d,J=8.0Hz,1H),7.29(t,J=7.4Hz,1H),3.02(d,J=4.8Hz,4H),1.66(s,4H),1.59(s,8H); 13 C-NMR(100MHz,DMSO-d6)δ179.8,162.0,149.9,147.3,143.4,135.1,130.8, 128.7,126.4,126.0,124.2,119.2,5.0,27.2,26.0,25.0.HRMS(ESI-TOF)m / z calcd.for C 22 H 27 N4O3S[M+H] + :427.1804,found427.1802.

[0163] Example 15 Synthesis of Compound 14

[0164] The specific synthetic route is as follows:

[0165]

[0166] Select side chain VIII-e (1.00mmol) and dissolve it in 10mL methanol, add 2mL sodium hydroxide aqueous solution (5M), heat to reflux, and after the reaction is complete, extract it three times with 50mL dichloromethane, combine the organic phases, add anhydrous Na2SO4 to dry, filter to remove the desiccant, evaporate the solvent to obtain a crude product. Take the above crude product (0.5mmol) and dissolve it in 2mL DMF, add 2,4-dichloro-5,6,7,8-tetrahydroquinazoline (1.0mmol), palladium acetate (0.1mmol), 1,1'-binaphthyl-2,2'-bisdiphenylphosphine (0.1mmol) and cesium carbonate (1.0mmol), use a microwave reactor, microwave reaction at 100℃ for 3h, cool to room temperature, pour into 10mL ice water, collect the precipitated solid, separate and purify by column chromatography to obtain product 14. Yield: 45%; Structural parameters: 1H-NMR (400MHz, Acetone-d6) δ8.28(s,1H),8.00(d,J=8.8Hz,2H),7.77(d,J=8.8Hz,2H),3.28(t,J=5.8Hz,4H ),2.70(t,J=6.0Hz,2H),2.65(t,J=6.2Hz,2H),1.84-1.90(m,4H),1.70(d,J=3.6Hz,4H),1.57-1.60(m,4H); 13 C-NMR (100MHz, Acetone-d6) δ167.3,160.6,157.1,143.9,134.8,128.6,121.6,114.1,48.9,32.7,27.6,22.6,22.5,18.9.

[0167] Example 16 Synthesis of Compound 15

[0168]

[0169] The synthesis method of Example 16 is the same as that of the above-mentioned compound 14, and the side chain VIII-g (1.00 mmol) is selected. Yield: 45%; Structural parameters of compound 15: 1 H-NMR (400MHz, CDCl3) δ7.78 (s, 4H), 6.63 (s, 1H), 3.09 (t, J = 5.0Hz, 4H), 2.80 (t, J = 6.0Hz, 2H),2.52(t,J=6.0Hz,2H),1.93-1.96(m,2H),1.85-1.89(m,2H),1.74(s,8H),1.65(s,4H); 13 C-NMR (100MHz, CDCl3) δ166.8,158.9,157.0,141.9,132.7,128.6,120.1,112.0,50.2,27.3,26.0,25.0,22.2,21.9,21.7.HRMS(ESI-TOF)m / zcalcd.for C 22 H 30 ClN4O2S[M+H] + :449.1778,found 449.1778.

[0170] Example 17 Synthesis of Compound 16

[0171]

[0172] Select side chain VIII-e (1.00mmol) and dissolve it in 10mL methanol, add 2mL sodium hydroxide aqueous solution (5M), heat and reflux, after the reaction is complete, extract it three times with 50mL dichloromethane, combine the organic phases and add anhydrous Na2SO4 to dry, filter to remove the desiccant, evaporate the solvent to obtain a crude product. Take the above crude product (0.5mmol) and dissolve it in 2mL DMF, add 2-phenyl-7,8-dihydro-6H-thiopyrano[3,2-d]pyrimidin-4-yl trifluoromethanesulfonate (1.0mmol), react at 100℃ for 3h, cool to room temperature, pour into 10mL ice water, collect the precipitated solid, separate and purify by column chromatography to obtain the final product 16. Yield: 40%; Product 16 structural parameters: 1 H-NMR (400MHz, CDCl3) δ8.34(d,J=5.6Hz,2H),7.90(d,J=8.8Hz,2H),7.81(d,J=9.6Hz,2H),7.46(d,J=5.6Hz,3H),6.81(s ,1H),3.30(t,J=5.8Hz,4H),3.16(t,J=6.4Hz,2H),3.00(t,J=6.2Hz,2H),2.30(t,J=5.8Hz,2H),1.73(s,4H),1.59(s,4H); 13 C-NMR (100MHz, CDCl3) δ160.1,156.8,155.1,142.9,137.8,133.1,130.2,128.5,1 28.2,127.8,199.6,110.1,48.3,32.2,29.2,27.8,27.0,23.2.HRMS(ESI-TOF)m / z calcd.forC 25 H 29 N4O2S2[M+H] + :481.1732,found 481.1732.

[0173] Example 18 Synthesis of Compound 17

[0174]

[0175] The synthesis method of Example 18 is the same as that of the above-mentioned compound 16, and the side chain VIII-g (1.00 mmol) is selected. Yield: 40%; Structural parameters of compound 17: 1H-NMR (400MHz, CDCl3) δ8.34-8.36(m,2H),7.91(d,J=8.4Hz,2H),7.82(d,J=8.8Hz,2H),7.47(d,J=6.0Hz,2H),7.46(s,1H),6. 81(s,1H),3.18(t,J=5.8Hz,2H),3.11(t,J=5.0Hz,2H),3.01(t,J=6.4Hz,2H),2.32(t,J=6.0Hz,2H),1.75(s,8H),1.66(s,4H); 13 C-NMR (100MHz, CDCl3) δ160.1,158.8,155.1,143.0,137.8,131.6,130.2,128.5, 127.8,119.5,110.1,50.2,32.2,27.8,27.3,26.0,25.0,23.2.HRMS(ESI-TOF)m / z calcd.for C 27 H 33 N4O2S2[M+H] + :509.2045,found 509.2045.

[0176] Example 19 Synthesis of Compound 18

[0177]

[0178] The synthesis method of Example 19 is the same as that of the above-mentioned compound 16. The side chain VIII-e (1.00 mmol) is selected and reacted with 2-(4-(trifluoromethyl)phenyl)-7,8-dihydro-6H-thiopyrano[3,2-d]pyrimidin-4-yl trifluoromethanesulfonate to obtain compound 18. Yield: 44%; Structural parameters of compound 18: 1 H-NMR (400MHz, CDCl3) δ8.45(d,J=8.0Hz,2H),7.81-7.88(m,4H),7.72(d,J=8.4Hz,2H),3.31(t,J=5.8Hz,2H),3.3 1(t,J=5.8Hz,4H),3.20(t,J=5.8Hz,2H),3.02(t,J=6.4Hz,2H),2.30-2.36(m,2H),1.74(s,4H),1.59-1.62(m,4H); 13C-NMR (100MHz, CDCl3) δ160.0,157.2,155.1,142.6,141.1,133.5,131.8,131.5,128.2,128.0,1 25.6,125.4,125.4,122.9,119.8,111.2,48.3,32.2,29.2,27.8,26.9,23.0.HRMS(ESI-TOF)m / z calcd.for C 26 H 28 F3N4O2S2[M+H] + :549.1606,found549.1606.

[0179] Example 20 Synthesis of Compound 19

[0180]

[0181] The synthesis method of Example 20 is the same as that of the above-mentioned compound 16. The side chain VIII-e (1.00 mmol) is selected and reacted with 2-(4-(trifluoromethyl)phenyl)-7,8-dihydro-5H-thiopyrano[4,3-d]pyrimidin-4-yl trifluoromethanesulfonate to obtain compound 19. Yield: 47%; Structural parameters of compound 19: 1 H-NMR (400MHz, CDCl3) δ8.47(d,J=8.0Hz,2H),7.80-7.85(m,4H),7.72(d,J=8.4Hz,2H),6.63(s,1H),3.70( s,2H),3.32(t,J=6.0Hz,4H),3.25(t,J=5.8Hz,2H),3.01(t,J=6.0Hz,2H),1.75(s,4H),1.60-1.62(m,4H); 13 C-NMR (100MHz, CDCl3) δ163.8,159.7,157.0,142.4,133.9,132.2,128.3,128.2,1 25.5,125.4,120.3,110.4,48.3,34.2,29.2,26.9,25.0,23.5.HRMS(ESI-TOF)m / z calcd.forC 26 H 28 F3N4O2S2[M+H] + :549.1606,found 549.1606.

[0182] Example 21 Experiment on the inhibition of tumor cell proliferation by compound 1-19 and its hydrochloride

[0183] Tumor cells (SW480, HCT116, HepG2) were cultured in 1% penicillin-streptomycin solution and 10% fetal bovine serum in IMDM cell culture medium at 37°C in a constant temperature incubator containing 5% CO2. Tumor cells in the logarithmic growth phase were taken and the cell concentration was adjusted to 5×10 4 Cells / mL were inoculated on a 96-well plate, 100 μL per well, and blank wells and control wells were set up at the same time. After 24 hours of culture, compounds with final concentrations of 0.001, 0.01, 0.1, 1, and 10 μM were added, 0.5 μL per well, and 3 replicates were set up for each drug concentration. Blank wells are simple culture medium wells without cells, DMSO, and compounds. Control wells are complete culture medium containing only the same concentration of DMSO added to act on cells. The cells were placed in a 37°C, 5% CO2 constant temperature incubator. After 6 h, 12 h, 24 h, and 48 h, 20 μL of 5 mg / mL MTT solution (prepared with PBS and sterilized by filtration with a 0.22 μm filter membrane) was added to each well. The cells were placed in a 37°C, 5% CO2 constant temperature incubator and incubated for 4 h. The culture was terminated, and the culture supernatant in the wells was carefully removed. 100 μL of DMSO was added to each well. The cells were mixed by repeated pipetting. After being placed at 37°C for 10 min, the purple crystals were fully dissolved. The absorbance (OD) value of each well was measured with a microplate reader (490 nm, 630 nm). The cell inhibition rate was calculated according to the following formula.

[0184] Cell survival rate (%) = (OD of experimental group - OD of blank group) / (OD of control group - OD of blank group) × 100%

[0185] According to the MTT results, the linear regression equation was obtained and the IC of each compound was calculated. 50 value (half effective inhibitory concentration).

[0186] The in vitro antitumor activity results of compound 1-19 as its hydrochloride are shown in Table 1.

[0187] Table 1. In vitro antitumor activity of compounds 1-19 as their hydrochloride salts

[0188]

[0189] Note: a Camptothecin was used as a positive control.

[0190] The results in Table 1 show that compounds 1-19 all have certain inhibitory activity against human colon cancer cells. In the process of cell inhibitory activity testing, it was found that the solubility of hydrochloride 1a, 2a and 3a was better than that of the corresponding unsalted precursor compounds, and the inhibitory activity did not change much.

[0191] Example 22 Experiment on the inhibition of tumor cell proliferation by compound 3a

[0192] Representative compound 3a was selected for further antitumor activity evaluation. In order to observe whether compound 3a can effectively inhibit the migration of cancer cells, an in vitro scratch test was used for evaluation. Fig. 20 As shown, compound 3a has the ability to inhibit cancer cell migration. At 12h, 2μM compound 3a showed preliminary inhibitory activity on both HCT-116 and SW480 cells. After 24h of administration, the HCT-116 and SW480 cells in the control group migrated by about 40% and 30%, respectively, and 2μM compound 3a effectively inhibited cell migration. At 48h, without drug intervention, the migration distance of the compound reached more than 50%, and 0.5μM compound 3a can effectively inhibit HCT-116 and SW480 cells. The inhibitory effect of 2μM is the most significant, and it is concentration-dependent.

[0193] Example 23 Evaluation of the Cellular Glucose Consumption Promoting Activity of the Compounds

[0194] Glucose consumption experiments were used to evaluate the amount of glucose consumption promoted by compounds at different concentrations, thereby reflecting the hypoglycemic activity at the cellular level. Metformin was used as a positive control. The specific method was as follows: HepG2 cells in the logarithmic growth phase were routinely digested and cultured in DMEM low-glucose medium. The inoculation cell density was 5×10 4 100 μL / mL was added to a 96-well culture plate, and each well was placed in an incubator for overnight culture. After the cells adhered to the wall, the supernatant was discarded, washed once with 1×PBS, starved for 24 hours with serum-free DMEM high-glucose medium, washed once with 1×PBS, and cultured with DMEM high-glucose medium. A drug control group (culture medium + test substance + cells), a negative control group (culture medium + DMSO + cells) and a blank control group (simple culture medium + test substance) + a positive control group (culture medium + metformin + cells) were set up. Three replicate wells were set up for each concentration, and 0.5 μL of dilution containing different concentrations of the test substance was added to each well. The cells were placed in a carbon dioxide incubator and continued to be cultured. After 24 hours, the relative glucose consumption was detected using a glucose determination kit.

[0195] First, representative compounds 3 and 3a were selected to evaluate the cytotoxicity of the compounds after 24 h of treatment. At a concentration of 1 μM, the cell survival rate was about 80%. Therefore, 1 μM and 0.2 μM concentrations were selected to test their effects on the amount of glucose consumed by cells. Fig.21 As shown, compound 3a can significantly promote glucose consumption at both 1 μM and 0.2 μM, and compound 3 can significantly promote glucose consumption at 1 μM, and its activity is comparable to that of the positive control metformin at a concentration of 500 μM, indicating that both compounds have good activity in promoting glucose consumption.

[0196] Further, the activity of compound 1-19 and its hydrochloride in promoting glucose consumption was evaluated at a concentration of 1 μM, as shown in Table 2.

[0197] Table 2. Glucose consumption promoting activity of compounds 1-19 as their hydrochloride salts

[0198]

[0199] Note: Metformin was used as a positive control and the tested concentration was 500 μM.

[0200] As shown in Table 2, all tested compounds have a certain degree of promoting activity on cellular glucose consumption at a concentration of 1 μM, indicating that this class of compounds has the potential to lower blood sugar. In addition, tumor cell proliferation requires a lot of energy, which affects glucose consumption and may be related to inhibiting tumor proliferation.

[0201] Example 24 Study on the lipid-lowering activity of the compound at the cellular level

[0202] Oil red O staining was used to evaluate the activity of different concentrations of representative compounds in reducing fat content at the cellular level. Lovastatin was used as a positive control. The specific method was as follows: HepG2 cells in the logarithmic growth phase were taken and cultured with DMEM low-glucose medium at a cell density of 1×10 5 The cells were inoculated with 2 mL / mL in a 6-well plate, 2 mL per well, and cultured in a 37°C incubator overnight. After the cells adhered, the supernatant was discarded, the cells were washed once with 1×PBS, starved with serum-free DMEM high-glucose medium for 24 h, washed once with 1×PBS, and induced with the prepared DMEM low-glucose complete medium (containing 1% BSA) containing inducer (0.5% sodium oleate and 0.25% sodium palmitate) for 24 h to establish a HepG2 cell fat accumulation model, and a drug control group (inducer + test substance + cells), a negative control group (inducer + DMSO + cells) and a blank control group (DMEM low-glucose medium solution containing only 1% BSA) and a positive control group (inducer + lovastatin + cells) were set up. Three replicate wells were set for each concentration, and 10 μL of dilution containing different concentrations of the test substance was added to each well. The cells were placed in a carbon dioxide incubator and continued to be cultured for 24 hours, then washed three times with 1×PBS, fixed with 2 mL of 4% paraformaldehyde per well for 30 minutes, washed three times with 1×PBS, and acted on the cells with 2 mL of 60% isopropanol per well for 5 minutes to increase the permeability of the cells. They were stained with 2 mL of Oil Red O per well for 1 hour in the dark at room temperature, washed with distilled water for 4 times, and then 1 mL of isopropanol was added to each well for binding for 10 minutes, washed out with shaking, and the absorbance was measured at 492 nm with an enzyme reader.

[0203] Since the compound had little effect on cell viability at 1 μM and 0.2 μM concentrations at 24 h, the lipid-lowering activity was still tested at the above two concentrations. Fig. 22 As shown, compounds 3a and 3 at a concentration of 1 μM could significantly reduce the intracellular lipid content.

[0204] Further, the lipid-lowering activity of compound 1-19 and its hydrochloride at the cellular level was evaluated at a concentration of 1 μM, as shown in Table 3.

[0205] Table 3. Glucose consumption promoting activity of compounds 1-19 as their hydrochloride salts

[0206]

[0207] Note: Lovastatin was used as a positive control and the test concentration was 10 μM.

[0208] As shown in Table 3, all the tested compounds at a concentration of 1 μM had a certain reducing effect on the increase of intracellular lipids induced by cell inducers, indicating that this type of compound also has a certain activity in lowering blood lipids.

[0209] Example 25 Study on the in vivo antitumor activity of compound 3a

[0210] (1) Establishment of Balb / C nude mouse colon cancer model

[0211] HCT116 cells in the logarithmic growth phase and in good condition (viable cell count>95%) were selected, centrifuged at 1000 rpm for 5 min to obtain cell pellets, washed twice with pre-cooled phosphate buffer and counted. The cell number was adjusted according to the cell density using pre-cooled physiological saline to adjust the cell concentration to 5×10 6 The suspension of 100 μg / mL was placed in a tube and placed on ice for later use.

[0212] After one week of adaptive feeding, Balb / C nude mice (5 weeks old) were fixed and disinfected with 75% medical cotton balls on the right armpit of the mice. The HCT116 cell suspension was drawn up with a 1mL syringe, and after the excess bubbles were discharged, it was injected into the subcutaneous part of the right armpit of the mice. Each nude mouse was injected with 0.2mL. The needle was pulled out and the needle hole was gently pressed with a sterile cotton swab to prevent the cell fluid from flowing out with the needle.

[0213] After inoculation of HCT116 cells, the mice were observed to see if there were any abnormal changes in their diet, water intake, and body weight. The inoculation site was observed once a day to see if a nodule grew. When a substantial mass was felt, the subcutaneous tumor model was successfully constructed. When the tumor mass of the nude mouse model grew to about 1 cm in diameter, secondary transplantation preparations were performed.

[0214] Before tumor transplantation, tert-amyl alcohol was used to prepare the Avertin solution, which was prepared immediately before use and placed in a 4°C refrigerator for later use. It was preheated to 37°C and injected intraperitoneally. The anesthetic dose of Avertin was 0.4 mL / weight of nude mouse.

[0215] The tumors in nude mice were removed in a clean bench and placed in a culture dish containing physiological saline to wash the tumors. Tumors with good growth were selected, the capsule and connective tissue were removed, and they were cut into tissue blocks with a diameter of about 2 mm using sterile equipment for subsequent use.

[0216] The anesthetized mouse was placed flat in the clean bench, and the right armpit of the mouse was disinfected with 75% medical cotton balls. A 0.4 cm incision was made with sterile scissors, and a tumor was picked up with a special implant needle for tumor implantation, and then placed in the tumor implantation sleeve and inserted into the subcutaneous tissue. Suturing was performed with absorbable surgical sutures.

[0217] (2) Grouping and administration of tumor-bearing nude mice

[0218] The tumor volume of nude mice was measured every day. When the tumor volume of nude mice grew to 200mm 3 When the nude mice were about 100 μg / kg, they were grouped according to the weight and tumor size. The compound to be tested was dissolved in a glucose injection solution containing 5% Tween-80, and the administration method was intraperitoneal injection. The mice were divided into 5 groups according to the type and concentration of administration: normal group, model group, low-dose group (5 mg / kg), high-dose group (10 mg / kg), and positive control fluorouracil group (20 mg / kg). The administration cycle was: once every other day in the first week, for three consecutive times, no administration in the second week, once every other day in the second week, for two consecutive times, no administration in the third week, and the mice were killed on the 30th day.

[0219] (3) Evaluation of pharmacological and physicochemical indicators

[0220] The short diameter and long diameter of the tumor were measured and recorded every 2 days, and a curve of the overall change of tumor volume in each group of nude mice was drawn. The mice were weighed before being killed. Blood was collected from the femoral artery. After standing for 30 minutes, the blood was centrifuged at 3500rpm for 10 minutes at 4°C to collect serum. The mice were dissected, the tumors were removed, and washed in physiological saline. The excess water was absorbed with filter paper and placed on a precision balance for weighing, recording and photographing.

[0221] Relative tumor weight: organ index = tumor weight / nude mouse body weight;

[0222] Relative tumor inhibition rate: relative tumor volume (RTV) and relative tumor proliferation T / C (%) were calculated according to the following formula: RTV = Vt / V0, where Vt: tumor volume obtained by each tumor measurement; V0: initial tumor volume (first administration); T / C (%) = average RTV value of the administration group / average RTV value of the control group × 100%;

[0223] Tumor volume inhibition rate: IRTV (%) = 100% - T / C;

[0224] The tumor volume of mice was measured during the administration period, and the tumor growth curve was drawn (see Fig.23 ). The nude mice in the control group were injected with the same volume of 5% Tween-80 glucose injection, and the tumor grew faster. There was no significant difference in tumor volume between the nude mice in the 5mg / kg administration group and the nude mice in the control group on the 30th day (P>0.05), and no obvious inhibitory activity was shown. At a dose of 10mg / kg, the tumor volume of nude mice was significantly inhibited (P<0.001), the overall trend of nude mouse tumor growth was very slow, and the growth of the tumor was significantly inhibited. Nude mice in the fluorouracil group died, and all died after 12 days.

[0225] After the tumor was removed, a photo was taken. Fig.24 As shown, by direct observation, it can be found that the tumors of nude mice in the 10 mg / kg group are significantly smaller than those in the control group, and the shape is more regular. The weight of nude mouse tumors is another intuitive indicator for evaluating the effectiveness of drugs. The nude mouse tumors were weighed, and the weight of nude mouse tumors in the 10 mg / kg group was significantly reduced, only 41% of that in the control group (Table 3). In order to exclude the influence of the decrease in the weight of the mice on the tumor weight, the relative tumor weight was calculated to find that the compound was still an effective dose at 10 mg / kg. Table 3 shows that the relative tumor inhibition rate of compound 3a can reach 80% at 10 mg / kg, which is 6 times the inhibition rate of the 5 mg / kg dosage. Therefore, compound 3a has good tumor inhibition activity at a dose of 10 mg / kg and has the potential to be developed into a new anti-tumor drug.

[0226] Table 3 Tumor weight and relative tumor inhibition rate (IRTV) of compound 3a at different doses

[0227] Group Dosage (mg / kg) Tumor weight (g) IRTV(%) Model Group - 3.50±0.51 - Treatment group 1 10 1.44±0.27*** 80.17 Treatment group 2 5 3.11±0.85 12.14

[0228] In summary, the present invention synthesized a series of benzenesulfonamide derivatives, completed the optimization of the synthesis process, and evaluated the in vitro anti-tumor, hypoglycemic and hypolipidemic activities of the compounds. The synthesis process of the present invention is simple, does not require anhydrous and anaerobic conditions, and some reactions are cleverly combined with microwave conditions, so that the partial reaction yield is high and the time is short. Subsequent activity studies have shown that the compounds have the activity of inhibiting tumor cell proliferation and migration, and can significantly inhibit tumor growth at a dose of 10 mg / kg in vivo; in addition, they also have hypoglycemic and hypolipidemic activities. Therefore, it is shown that the compound has good application prospects.

[0229] Although the embodiments of the present invention are disclosed for illustrative purposes, those skilled in the art will appreciate that various substitutions, changes and modifications are possible without departing from the spirit and scope of the present invention and the appended claims. Therefore, the scope of the present invention is not limited to the contents disclosed in the embodiments.

Claims

1. A benzenesulfonamide derivative, characterized in that: The structural formula of the derivative is as follows: Among them, RNR' is and its hydrochloride, wherein n1 is equal to or not equal to n2, n1=a natural number from 2 to 4, n2=a natural number from 2 to 4, and R1 is methyl, ethyl, propyl, or butyl; Alternatively, RNR' is and its hydrochloride, wherein n1 is equal to or not equal to n2, n1=a natural number from 2 to 4, n2=a natural number from 2 to 4, and m=a natural number from 1 to 3; Alternatively, RNR' is 2. The benzenesulfonamide derivative according to claim 1, characterized in that: The specific structural formula is as follows:

3. The method for preparing the benzenesulfonamide derivatives according to claim 1 or 2, characterized in that: The reaction route is: or, Among them, RNHR1 is or, 4. The preparation method according to claim 3, characterized in that: The steps include: The fatty amine raw material RNHR1 reacts with 4-acetylaminobenzenesulfonyl chloride to obtain VIII, and the acetyl protecting group of the amino group is removed under alkaline conditions to obtain IX, and IX reacts with a halogenated aromatic group to obtain a benzenesulfonamide derivative.

5. Use of the benzenesulfonamide derivatives as claimed in claim 1 or 2 in the preparation of drugs for treating human colon cancer.

6. Use of the benzenesulfonamide derivatives as claimed in claim 1 or 2 in the preparation of drugs for treating human liver cancer.

7. Use of the benzenesulfonamide derivatives as claimed in claim 1 or 2 in the preparation of drugs for treating glucose and lipid metabolism disorders.

8. Use of the benzenesulfonamide derivatives as claimed in claim 1 or 2 in the preparation of drugs for treating hypolipidemics.

9. Use of the benzenesulfonamide derivatives according to claim 1 or 2 in the preparation of therapeutic hypoglycemic drugs.

Images