CN108017614A - DPP-4 inhibitor and its application in treatment diabetes B medicine is prepared - Google Patents

Abstract

The present invention relates to three novel DPP-4 inhibitors and their applications in the field of drugs for treating type 2 diabetes. The DPP-4 inhibitors have one or more than two of the following three compounds as active ingredients, and the structures of the three compounds : The compound increases the levels of incretin-like peptide-1 and glucose-dependent insulin-releasing peptide by inhibiting the activity of dipeptidyl peptidase-4, and has a good therapeutic effect on type 2 diabetes.

Description

DPP-4 inhibitor and its application in preparation of medicine for treating type 2 diabetes

technical field

The present invention relates to biomedicine, specifically three aromatic ring derivatives 2-{[6-[(3R)-3-amino-1-piperidinyl]-3,4-dihydro-3- (3,5-Difluorobenzyl)-2,4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile (1), 2-{[6-[(3R )-3-amino-1-piperidinyl]-3,4-dihydro-3-(3,5-dimethylbenzyl)-2,4-dioxo-1(2H)-pyrimidinyl] Methyl}-4-fluorobenzonitrile (2), 2-{[6-[(3R)-3-amino-1-piperidinyl]-3,4-dihydro-3-(3,4- Dimethoxybenzyl)-2,4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile (3) and its pharmacological activity and pharmaceutical use. The above compounds and their derivatives can be used as DPP-4 inhibitors to treat type 2 diabetes.

Background technique

Diabetes is a metabolic disorder with a rapidly increasing number of patients. According to the latest data from the International Diabetes Federation, as of 2017, there are approximately 425 million adult diabetics in the world, and it is predicted that this number will increase to 700 million by 2045. The treatment of diabetes is particularly urgent.

Dipeptidyl peptidase IV (DPP-4), also known as CD26, is a multifunctional transmembrane serine protease that degrades incretin-like peptide-1 (GLP-1) and glucose-dependent insulinotropic peptide (GIP). ) to regulate glucose metabolism. Oral dipeptidyl peptidase IV inhibitors can prolong the half-life of GLP-1 and GIP in the body, the level of GLP-1 and GIP by inhibiting the activity of dipeptidyl peptidase IV, prolong their hypoglycemic action time, and inhibit pancreatic Glucagon secretion, prolongs the duration of stimulation of GLP-1 on insulin secretion, and exerts hypoglycemic effect. DPP-4 inhibitors can stably control blood sugar, improve β-cell function, and will not cause weight gain in patients, avoid the risk of hypoglycemia, have significant advantages in drug safety, and are generally considered to be the most promising treatment for type 2 diabetes new target.

Natural products of marine origin have been an important source of hypoglycemic drugs. Phenyl-substituted compounds (aromatic ring-substituted compounds) isolated from the red alga Laurencia similis in tropical and subtropical regions have an IC ₅₀ of hypoglycemic activity of 2.66 (Wang LJ et al., Rsc Advances.5(2015), 48822); The aromatic ring compounds isolated from the ethanol extract of Rhodomela confervoides also have significant hypoglycemic activity (Shi DY et al., China J. Chin. Mater. Med. 33 (2008), 2238). Accordingly, we synthesized 2-{[6-[(3R)-3-amino-1-piperidinyl]-3,4-dihydro- 3-(3,5-difluorobenzyl)-2,4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile (1), 2-{[6-[ (3R)-3-amino-1-piperidinyl]-3,4-dihydro-3-(3,5-dimethylbenzyl)-2,4-dioxo-1(2H)-pyrimidine Base] methyl}-4-fluorobenzonitrile (2), 2-{[6-[(3R)-3-amino-1-piperidinyl]-3,4-dihydro-3-(3, 4-Dimethoxybenzyl)-2,4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile (3).

Contents of the invention

The purpose of the present invention is to provide three novel DPP-4 inhibitors. The three compounds prolong the action time of GLP-1 and GIP by inhibiting the activity of DPP-4, so as to achieve the purpose of lowering blood sugar and can be used for treating type 2 diabetes.

To achieve the above object, the technical scheme adopted in the present invention is as follows:

DPP-4 inhibitor, which uses one or more than two of the following three compounds as active ingredients, and the structural formulas of the three compounds are as follows:

That is, 2-{[6-[(3R)-3-amino-1-piperidinyl]-3,4-dihydro-3-(3,5-difluorobenzyl)-2,4-dioxo -1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile (1), 2-{[6-[(3R)-3-amino-1-piperidinyl]-3,4-di Hydrogen-3-(3,5-dimethylbenzyl)-2,4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile (2), 2-{[ 6-[(3R)-3-amino-1-piperidinyl]-3,4-dihydro-3-(3,4-dimethoxybenzyl)-2,4-dioxo-1( 2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile (3).

The purpose of the present invention is to provide the application of three kinds of aromatic ring derivatives derived from marine sources in the preparation of medicines for treating type 2 diabetes.

Another object of the present invention is to provide a pharmaceutical composition for preparing or preventing type 2 diabetes mellitus, using one or two of compounds 1-3 or a pharmaceutically acceptable salt thereof as an active ingredient.

The above-mentioned pharmaceutical composition (i.e. DPP-4 inhibitor), its composition contains therapeutically effective amount of one or two of the compounds 1-3 or their pharmaceutically acceptable salts, said compound in the pharmaceutical composition It can be used alone or in combination with other medicines. The pharmaceutical composition contains 0.1-99%, preferably 0.5-90%, of the compound of the present invention, and the rest are pharmaceutically acceptable carriers and/or excipients.

The pharmaceutical carrier or excipient is one or more solid, semi-solid and liquid diluents, fillers and pharmaceutical preparation adjuvants. The pharmaceutical composition of the present invention is used in a dose per body weight. The medicine of the present invention can be administered in three forms of injection (intravenous injection, intramuscular injection), oral administration and external application.

The compound increases the levels of incretin-like peptide-1 and glucose-dependent insulin releasing peptide by inhibiting the activity of dipeptidyl peptidase-4, and has a good therapeutic effect on type 2 diabetes.

The present invention has the following advantages:

The target compound is a compound with significant DPP-4 enzyme inhibitory activity, and is a new drug for the treatment of type 2 diabetes; the present invention introduces marine-sourced aromatic ring derivatives, which are rich in resources, low in toxicity, and have good hypoglycemic activity. The high-yield and low-cost synthesis of the target compound is achieved, and the process operation is simple, and has a good prospect for industrial production.

Description of drawings

Figure 1 is a dose-dependent graph of compound 1 inhibiting DPP-4.

Fig. 2 is a dose-dependent diagram of compound 2 inhibiting DPP-4.

Fig. 3 is a dose-dependent diagram of compound 3 inhibiting DPP-4.

1. Compound synthesis and structure identification

Using 6-chlorouracil and 2-cyano-5-fluorobenzyl bromide as starting materials, the intermediate was synthesized by substitution reaction; then three different aromatic ring derivatives bromobenzyl were generated on the 1-position nitrogen of pyrimidinedione Aromatic ring derivatives are substituted to generate aromatic ring derivatives to replace intermediates; then react with (R)-3-(Boc-amino)piperidine at the 6-position of pyrimidinedione, and finally remove tert-butoxycarbonyl with trifluoroacetic acid to obtain The crude product was purified by silica gel column chromatography with chloroform-methanol-triethylamine system as eluent to obtain a white solid. The compound 2-{[6-[(3R)-3-amino-1-piperidinyl]-3,4-dihydro-3-(3,5-difluorobenzyl)- 2,4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile (1), 2-{[6-[(3R)-3-amino-1-piperidinyl ]-3,4-dihydro-3-(3,5-dimethylbenzyl)-2,4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile ( 2), 2-{[6-[(3R)-3-amino-1-piperidinyl]-3,4-dihydro-3-(3,4-dimethoxybenzyl)-2,4 -Dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile (3); English names are: 2-[6-(3(R)-aminopiperidin-1-yl)- 3-(3,5-difluorobenzyl)-2,4-dioxo-3,4-dihydro-2H-pyrimidin-1-ylmethyl]-4-fluorobenzonitrile(1),2-[6-(3(R)-aminopiperidin -1-y l)-3-(3,5-dimethyl)-2,4-dioxo-3,4-dihydro-2H-pyrimidin-1-ylmethyl]-4-fluorobe nzonitrole(2),2-[6- (3(R)-aminopiperidin-1-yl)-3-(3,4-dimethoxybenzyl)-2,4-diox o-3,4-dihydro-2H-pyrimidin-1-ylmethyl]-4-fluorobenzonitrile(3 ).

2. Determination of inhibitory activity of dipeptidyl peptidase 4

A novel small molecule fluorescent probe with aggregation-induced luminescent properties was used to screen DPP-4 inhibitor activity. The small-molecule fluorescent probe itself does not emit fluorescence in the reaction system. When DPP-4 cuts off the N-terminal dipeptide of the fluorescent probe, the water solubility of the residue decreases, and aggregation occurs to generate fluorescence. The method has high sensitivity and specificity for the detection of DPP-4 activity.

Detailed ways

In order to better understand the essence of the present invention, the following examples of the present invention will be used to illustrate the results of the pharmacological effects of these five compounds, but the present invention is not limited by these examples.

The synthetic route of compound is specifically shown as follows:

(a) The molar ratio of 2-cyano-5-fluorobenzyl bromide to compound 6 is 1:1～1:1.5, diisopropylethylamine, the solvent is N,N-dimethylformamide, room temperature; (b ) The molar ratio of compound 18 (or 19, or 20) to compound 7 is 1:1～1:1.5, potassium carbonate, potassium iodide, room temperature; (c) (R)-3-(Boc-amino)piperidine and compound 21 (or 22, or 23) with a molar ratio of 1:1~1:1.5, potassium carbonate, solvent N,N-dimethylformamide, inert gas protection, 80~100°C; (d) trifluoroacetic acid, solvent Dichloromethane, room temperature;

Example 1 "2-{[6-[(3R)-3-amino-1-piperidinyl]-3,4-dihydro-3-(3,5-difluorobenzyl)-2,4- Chemical synthesis and structural identification of dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile (compound 1)"

(1) Synthesis and structure identification of 2-{[6-chloro-3,4-dihydro-2,4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile

Under stirring at room temperature, 13.53 mL of diisopropylethylamine was added to 80 mL of N,N-dimethylformamide solution in which 10 g of 6-chlorouracil was dissolved, and 20 mL of 16 g of 2-cyano- 5-Fluorobenzyl bromide (mass concentration 15-20%) in N,N-dimethylformamide solution, after stirring continuously for 24 hours under dark conditions, TLC detection, after the raw material point disappears, the reaction ends, add 10 times the amount of distilled water, a precipitate precipitated out, the addition was completed in 20 minutes, and the stirring was continued for 15 minutes. The precipitate was filtered, washed with ice water, and drained to obtain a white solid. The mixture was recrystallized with absolute ethanol to obtain 14.78 g of a white solid. After spectral analysis, it was confirmed that the compound was 2-{[6-chloro-3,4-dihydro-2,4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile;

(2) 2-{[6-Chloro-3,4-dihydro-3-(3,5-difluorobenzyl)-2,4-dioxo-1(2H)-pyrimidinyl]methyl} Chemical Synthesis of -4-Fluorobenzonitrile

Dissolve 1 g of 2-{[6-chloro-3,4-dihydro-2,4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzyl in 5 mL at room temperature Add 0.59 g of anhydrous potassium carbonate and 0.10 g of potassium iodide to the N,N-dimethylformamide solution of nitrile, and add 5 mL of N,N-dimethylformamide dissolved in 0.82 g of 3,5-difluorobenzyl bromide dropwise. Amide solution, reacted in the dark for 24 hours, TLC detection, after the raw material point disappeared, add 10 times the amount of distilled water to the reaction solution, extract 3 times with ethyl acetate, combine the organic phase, and wash 3 times with 20% saline, organic The phase was dried over anhydrous magnesium sulfate and concentrated under reduced pressure. After separation by column chromatography, 1.09 g of a white solid was obtained after eluting with petroleum ether: ethyl acetate 10:1. After spectral analysis, the compound was confirmed to be 2-{[6-chloro-3,4-dihydro-3-(3,5-difluorobenzyl)-2,4-dioxo-1(2H)-pyrimidine Base] methyl}-4-fluorobenzonitrile.

(3) 2-{[6-[(3R)-3-Boc-amino-1-piperidinyl]-3,4-dihydro-3-(3,5-difluorobenzyl)-2,4 Chemical Synthesis of -Dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile

In a 50mL three-necked flask, add 0.51g of anhydrous potassium carbonate and 0.55g of (R)-3-(Boc-amino)piperidine to 15mL of 1g of 2-{[6-chloro-3,4-bis Hydrogen-3-(3,5-difluorobenzyl)-2,4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile N,N-Dimethylformazol In the amide solution, protected by argon, reflux reaction at 90°C for 3 hours, detected by TLC, after the disappearance of the raw material point, the reaction solution was cooled to room temperature. Add 10 times the amount of distilled water, extract 3 times with ethyl acetate, combine the organic phases, and wash 3 times with 20% saline, the organic phases are dried with anhydrous magnesium sulfate, concentrated under reduced pressure, the mixture is separated by column chromatography, petroleum ether : Ethyl acetate 4:1 eluted to give 1.2 grams of white solid. After spectral analysis, it was confirmed that the compound was 2-{[6-[(3R)-3-Boc-amino-1-piperidinyl]-3,4-dihydro-3-(3,5-difluorobenzyl )-2,4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile;

(4) 2-{[6-[(3R)-3-amino-1-piperidinyl]-3,4-dihydro-3-(3,5-difluorobenzyl)-2,4-di Chemical Synthesis of Oxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile

Under stirring at 0°C, add 4.5 mL of dichloromethane solution dissolved with 0.9 mL of trifluoroacetic acid into 5 mL with 0.5 g of 2-{[6-[(3R)-3-Boc-amino-1-piperidinyl]- 3,4-Dihydro-3-(3,5-difluorobenzyl)-2,4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile in dichloromethane In the solution, TLC detects that after the raw material point disappears, add ice water to quench the reaction, adjust the pH to alkaline with 10% NaOH solution, add 30mL dichloromethane to extract 3 times, combine the organic phases, and add 20% saline After washing three times, the organic phase was dried over anhydrous magnesium sulfate and concentrated under reduced pressure. The mixture was separated by column chromatography and eluted with dichloromethane-methanol-triethylamine system to obtain 0.35 g of white solid. After spectral analysis, it was confirmed that the compound was 2-{[6-[(3R)-3-amino-1-piperidinyl]-3,4-dihydro-3-(3,5-difluorobenzyl)- 2,4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile, referred to as compound 1.

The physical and chemical properties of the compound are as follows: white solid, melting point 70-72°C; H NMR spectrum: ¹ H NMR (500MHz, CDCl ₃ ) δ7.65 (dd, J=8.5, 5.3Hz, 1H), 7.12–7.02 (m ,1H),6.99–6.85(m,1H),6.84(s,1H),6.82(s,1H),6.65(t,J=9.0Hz,1H),5.40(s,1H),5.23(t, J＝10.0Hz, 2H), 4.99(s, 2H), 3.98(s, 1H), 3.29–3.17(m, 2H), 2.94(d, J＝12.0Hz, 1H), 2.70(s, 2H), 2.03(d,J=13.0Hz,1H),1.82(s,1H),1.66–1.59(m,1H),1.45(d,J=25.0Hz,1H),1.26(d,J=8.0Hz,1H ). Carbon NMR spectrum: ¹³ C NMR (126MHz, CDCl ₃ ) δ165.2 (d, J = 12.7Hz), 163.8 (d, J = 12.7Hz), 162.5, 161.9 (d, J = 12.4Hz), 159.7, 152.1, 143.9(d, J=8.7Hz), 140.2(d, J=8.9Hz), 135.6(d, J=9.5Hz), 116.5, 115.9(d, J=22.7Hz), 115.4(d, J=22.7Hz), 111.3(dd, J=19.7, 6.0Hz), 106.6(d, J=3.2Hz), 103.1(dd, J=25.2, 26.5Hz), 90.2, 56.1, 51.8, 47.2, 46.6, 43.6, 30.4, 29.7.

Example 2 "2-{[6-[(3R)-3-amino-1-piperidinyl]-3,4-dihydro-3-(3,5-dimethylbenzyl)-2,4 Total chemical synthesis and structural identification of -dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile (compound 2)"

(1) 2-{[6-chloro-3,4-dihydro-3-(3,5-dimethylbenzyl)-2,4-dioxo-1(2H)-pyrimidinyl]methyl Chemical Synthesis of }-4-Fluorobenzonitrile

Dissolve 1 g of 2-{[6-chloro-3,4-dihydro-2,4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzyl in 5 mL at room temperature Add 0.59 g of anhydrous potassium carbonate and 0.10 g of potassium iodide to the N,N-dimethylformamide solution of nitrile, and add 5 mL of N,N-dimethylbenzyl bromide dissolved in 0.79 g of 3,5-dimethylbenzyl bromide dropwise. Formamide solution, reacted in the dark for 24 hours, TLC detection, after the raw material point disappeared, added 10 times the amount of distilled water to the reaction solution, extracted 3 times with ethyl acetate, combined the organic phase, and washed 3 times with 20% saline, The organic phase was dried over anhydrous magnesium sulfate and concentrated under reduced pressure. After column chromatography, eluting with petroleum ether: ethyl acetate 8:1, 1.15 g of white solid was obtained. After spectral analysis, it was confirmed that the compound was 2-{[6-chloro-3,4-dihydro-3-(3,5-dimethylbenzyl)-2,4-dioxo-1(2H)- pyrimidinyl]methyl}-4-fluorobenzonitrile.

(2) 2-{[6-[(3R)-3-Boc-amino-1-piperidinyl]-3,4-dihydro-3-(3,5-dimethylbenzyl)-2, Chemical Synthesis of 4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile

In a 50mL three-necked flask, add 0.49g of anhydrous potassium carbonate and 0.52g of (R)-3-(Boc-amino)piperidine to 15mL of 1g of 2-{[6-chloro-3,4-bis Hydrogen-3-(3,5-dimethylbenzyl)-2,4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile N,N-dimethyl In the formamide solution, protected by argon, reflux reaction at 90° C. for 4 hours, detected by TLC, after the starting point disappeared, the reaction solution was cooled to room temperature. Add 10 times the amount of distilled water, extract 3 times with ethyl acetate, combine the organic phases, and wash 3 times with 20% saline, the organic phases are dried with anhydrous magnesium sulfate, concentrated under reduced pressure, the mixture is separated by column chromatography, petroleum ether : Ethyl acetate 3:1 eluted to give 1.18 grams of white solid. After spectral analysis, it was confirmed that the compound was 2-{[6-[(3R)-3-Boc-amino-1-piperidinyl]-3,4-dihydro-3-(3,5-dimethylbenzyl base)-2,4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile;

(3) 2-{[6-[(3R)-3-amino-1-piperidinyl]-3,4-dihydro-3-(3,5-dimethylbenzyl)-2,4- Chemical Synthesis of Dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile

Under stirring at 0°C, add 4.5 mL of dichloromethane solution dissolved with 0.9 mL of trifluoroacetic acid into 5 mL with 0.5 g of 2-{[6-[(3R)-3-Boc-amino-1-piperidinyl]- 3,4-Dihydro-3-(3,5-dimethylbenzyl)-2,4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile dichloro In methane solution, TLC detection, after the raw material point disappears, add ice water to quench the reaction, adjust the pH to alkaline with 10% NaOH solution, add 30mL dichloromethane to extract 3 times, combine the organic phase, the mass concentration is 20% Washed with brine three times, the organic phase was dried over anhydrous magnesium sulfate, concentrated under reduced pressure, the mixture was separated by column chromatography, and eluted with dichloromethane-methanol-triethylamine system to obtain 0.34 g of white solid. After spectral analysis, the compound was confirmed to be 2-{[6-[(3R)-3-amino-1-piperidinyl]-3,4-dihydro-3-(3,5-dimethylbenzyl) -2,4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile, referred to as compound 2.

The physical and chemical properties of the compound are as follows: white solid, melting point 115-117°C; hydrogen nuclear magnetic resonance spectrum: ¹ H NMR (500MHz, CDCl ₃ ) δ7.63 (dd, J = 8.5, 5.3 Hz, 1H), 7.03 (t, J ＝7.0Hz,1H),6.90(s,1H),6.90(s,1H),6.88(s,1H),6.84(s,1H),5.44(s,1H),5.24–5.16(m,2H) ,4.94(s,2H),3.43(s,1H),3.41(s,1H),3.38(s,1H),2.92(s,1H),2.86(d,J=11.5Hz,1H),2.65( s,1H),2.22(s,6H),2.10(s,1H),1.79(s,1H),1.66–1.56(m,2H),1.27–1.23(m,1H). C NMR: ¹³ C NMR (126MHz, CDCl ₃ ) δ165.1 (d, J=259.56Hz), 163.0, 159.2, 152.1, 144.2 (d, J=8.82Hz), 137.9, 136.4, 135.6 (d, J=10.8Hz), 129.3, 126.0, 116.7, 115.8 (d, J=23.94Hz), 115.5 (d, J=25.2Hz), 106.6, 90.6, 54.0, 53.4, 51.8, 47.3, 46.6, 44.4, 28.6, 21.2.

Example 3 "2-{[6-[(3R)-3-amino-1-piperidinyl]-3,4-dihydro-3-(3,4-dimethoxybenzyl)-2, Total Chemical Synthesis and Structure Identification of 4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile (Compound 3)"

(1) 2-{[6-chloro-3,4-dihydro-3-(3,4-dimethoxybenzyl)-2,4-dioxo-1(2H)-pyrimidinyl]methanol Chemical Synthesis of Di}-4-Fluorobenzonitrile

Dissolve 1 g of 2-{[6-chloro-3,4-dihydro-2,4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzyl in 5 mL at room temperature Add 0.54 g of anhydrous potassium carbonate and 0.10 g of potassium iodide to the N,N-dimethylformamide solution of nitrile, and add 5 mL of N,N-dimethylformamide dissolved in 0.75 g of 3,4-dimethoxybenzyl bromide dropwise. base formamide solution, reacted in the dark for 24 hours, TLC detection, after the raw material point disappeared, add 10 times the amount of distilled water to the reaction solution, extract with ethyl acetate 3 times, combine the organic phase, and wash 3 times with 20% saline , the organic phase was dried over anhydrous magnesium sulfate, and concentrated under reduced pressure. After separation by column chromatography, 1.15 g of a white solid was obtained after elution with petroleum ether: ethyl acetate 3:1. After spectral analysis, the compound was confirmed to be 2-{[6-chloro-3,4-dihydro-3-(3,4-dimethoxybenzyl)-2,4-dioxo-1(2H) -pyrimidinyl]methyl}-4-fluorobenzonitrile.

(2) 2-{[6-[(3R)-3-Boc-amino-1-piperidinyl]-3,4-dihydro-3-(3,4-dimethoxybenzyl)-2 ,Synthesis of 4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile

In a 50mL three-necked flask, add 0.62g of anhydrous potassium carbonate and 0.6g of (R)-3-(Boc-amino)piperidine to 15mL of 1.28g of 2-{[6-chloro-3,4-di Hydrogen-3-(3,4-dimethoxybenzyl)-2,4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile N,N-dimethyl In the methyl formamide solution, under the protection of argon, the reaction was refluxed at 90° C. for 3 hours, and detected by TLC. After the starting point disappeared, the reaction solution was cooled to room temperature. Add 10 times the amount of distilled water, extract 3 times with ethyl acetate, combine the organic phases, and wash 3 times with 20% saline, the organic phases are dried with anhydrous magnesium sulfate, concentrated under reduced pressure, the mixture is separated by column chromatography, petroleum ether : Ethyl acetate 3:2 eluted to give 1.16 g of white solid. After spectral analysis, it was confirmed that the compound was 2-{[6-[(3R)-3-Boc-amino-1-piperidinyl]-3,4-dihydro-3-(3,4-dimethoxy Benzyl)-2,4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile;

(3) 2-{[6-[(3R)-3-amino-1-piperidinyl]-3,4-dihydro-3-(3,4-dimethoxybenzyl)-2,4 Chemical Synthesis of -Dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile

Under stirring at 0°C, add 4.5 mL of dichloromethane solution dissolved with 0.9 mL of trifluoroacetic acid into 5 mL with 0.55 g of 2-{[6-[(3R)-3-Boc-amino-1-piperidinyl]- 3,4-dihydro-3-(3,4-dimethoxybenzyl)-2,4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile In the methyl chloride solution, TLC detects that after the raw material point disappears, add ice water to quench the reaction, adjust the pH to alkaline with a 10% NaOH solution, add 30 mL of dichloromethane to extract 3 times, and combine the organic phases with a mass concentration of 20 % brine was washed 3 times, the organic phase was dried over anhydrous magnesium sulfate, and concentrated under reduced pressure. The mixture was separated by column chromatography and eluted with dichloromethane-methanol-triethylamine system to obtain 0.39 g of white solid. After spectral analysis, it was confirmed that the compound was 2-{[6-[(3R)-3-amino-1-piperidinyl]-3,4-dihydro-3-(3,4-dimethoxybenzyl )-2,4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile, referred to as compound 3.

The physical and chemical properties of the compound are as follows: white solid, melting point 75-77°C; hydrogen nuclear magnetic resonance spectrum: ¹ H NMR (500MHz, CDCl ₃ ) δ7.67 (dd, J = 8.5, 5.3 Hz, 1H), 7.07 (dd, J =8.0,2.1Hz,1H),7.05(s,1H),7.03(d,J=8.0Hz,1H),6.80(d,J=2.0Hz,1H),6.77(d,J=8.0Hz,1H ),5.37(s,1H),5.25(s,2H),5.02(s,2H),3.84(s,3H),3.83(s,3H),3.00(d,J=11.0Hz,1H),2.91 (d,J=4.5Hz,1H),2.90(s,1H),2.59(s,1H),2.39(s,1H),1.93(d,J=9.5Hz,1H),1.76(dd,J= 10.0, 3.9Hz, 1H), 1.60(dd, J＝10.5, 3.4Hz, 1H), 1.50(s, 2H), 1.24(d, J＝5.0Hz, 1H). C NMR spectrum: ¹³ C NMR( 126MHz, CDCl ₃ ) δ165.2(d, J=259.6Hz), 162.6, 159.4, 152.5, 148.7, 148.5, 144.4(d, J=8.7Hz), 135.5(d, J=9.6Hz), 129.4, 121.8 ,116.3,115.6(d,J=22.7Hz),114.5(d,J=24.0Hz),112.6,110.8,106.7,90.8,59.4,55.8,53.4,51.8,47.3,46.2,44.2,33.3,23.2.

Example 4 dipeptidyl peptidase-4 inhibitory activity assay

The compounds 1-3 to be tested were dissolved in DMSO respectively, and diluted with HEPES buffer solution to prepare test solutions with different concentrations. The experiment was divided into sample group (containing 10 μmol L ^-1 small molecule fluorescent probe tetraphenylethylene, 5 U L ^-1 DPP-4 solution, 10 μL test compound solution), sample control group (containing 10 μmol L ^-1 small Molecular fluorescent probe tetraphenylethylene, 25mg L ^-1 solution of the compound to be tested), control group (containing 10 μmol L ^-1 small molecule fluorescent probe tetraphenylethylene, 5U L ^-1 DPP-4 solution), For the blank group (10 μmol·L ^-1 small molecule fluorescent probe tetraphenylethylene), the reaction solvent was HEPES buffer (0.5 mmol·L ^-1 , pH 7.0), and the total reaction volume was 100 μL. The positive control drug was Sitagliptin. After incubation at 37°C for 30 minutes, the fluorescence intensity of each group was detected by a microplate reader (λex=320nm, λem=450nm), and the inhibition rate of each component to DPP-4 was calculated according to the following formula, wherein I represents the fluorescence intensity. Experimental results with express.

Table 1 dipeptidyl peptidase-4 inhibitory rate (%)

As shown in Figure 1-3.

The test results show that the above compounds have a significant inhibitory effect on dipeptidyl peptidase-4, and have potential clinical application prospects for anti-type 2 diabetes.

Example 5: Preparation of compound 1-3 injection

Compounds 1-3 were dissolved with a small amount of DMSO (weight ratio: 1:0.1-1:0.5, here 1:0.4), and then added water for injection (weight ratio: 1:20-1:200, in This is 1:200), finely filtered, potted and sterilized to make injection solution.

Embodiment 6: Preparation of compound 1～3 powder injection

Compounds 1-3 were respectively dissolved in a small amount of DMSO (weight ratio: 1:0.1-1:0.5, here 1:0.5), and then dissolved in sterile water for injection (weight ratio: 1:20-1 :60, here is 1:60), stir to dissolve, filter with sterile suction filter funnel, then aseptic fine filter, subpackage in ampoules, after low-temperature freeze-drying, aseptically melt-seal to obtain powder injection.

Embodiment 7: the preparation of compound 1～3 powder

Compounds 1 to 3 were added to the excipient at a weight ratio of 9:1 to the excipient (weight ratio, Tween 80: propylene glycol: cyclodextrin: lactose = 1:2:4:12) to prepare powder.

Embodiment 8: Preparation of compound 1～3 tablet

Compounds 1～3 are respectively according to its and excipient (weight ratio, hypromellose E5: microcrystalline cellulose MCC102: magnesium stearate: (8% povidone K30)=15:15:2:0.1 ) with a weight ratio of 5:1, adding excipients, granulating and compressing into tablets.

Embodiment 9: Preparation of compound 1-3 oral liquid

Compounds 1-3 were added to distilled water containing 20% simple syrup and 0.1% sodium benzoate, respectively, and prepared into oral liquid with a concentration of 15 μg/mL according to the conventional oral liquid preparation method.

Embodiment 10: Preparation of compound 1～3 capsules

Compounds 1-3 were respectively mixed with excipients (weight ratio, medicinal starch: glucose: hydrolyzed gelatin: glycine = 30:10:1:1) at a weight ratio of 5:1 to make capsules.

Embodiment 11: Preparation of compound 1～3 pills

Compounds 1-3 were respectively mixed with excipients (weight ratio, PEG2000:PEG6000=4:1) at a weight ratio of 1:10 to prepare pills.

Claims (10)

1. DPP-4 inhibitor, characterized in that: it uses one or more of the following three compounds or pharmaceutically acceptable salts respectively corresponding to the following three compounds as active ingredients, and the three compounds have the structural formula They are as follows: 2. according to the described DPP-4 inhibitor of claim 1, it is characterized in that: Compound 1: The 1-position nitrogen of pyrimidinedione is connected to 3,5-difluorobenzyl, the chemical name in Chinese is: 2-{[6-[(3R)-3-amino-1-piperidinyl]-3, 4-Dihydro-3-(3,5-difluorobenzyl)-2,4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile; English: 2- [6-(3(R)-aminopiperidin-1-yl)-3-(3,5-difluorobenzyl)-2,4-dioxo-3,4-dihydro-2H-pyrimidin-1-ylmethyl]-4-fluorobenzonitrile ; Compound 2: The 1-position nitrogen of pyrimidinedione is connected to 3,5-dimethylbenzyl, the chemical name in Chinese is: 2-{[6-[(3R)-3-amino-1-piperidinyl]-3 ,4-dihydro-3-(3,5-dimethylbenzyl)-2,4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile; English: 2-[6-(3(R)-aminopiperidin-1-yl)-3-(3,5-dimethyl)-2,4-dioxo-3,4-dihydro-2H-pyr imidin-1-ylmethyl]- 4-fluorobenzonitrile; Compound 3: The 1-position nitrogen of pyrimidinedione is connected to 3,4-dimethoxybenzyl, the chemical name in Chinese is: 2-{[6-[(3R)-3-amino-1-piperidinyl]- 3,4-Dihydro-3-(3,4-dimethoxybenzyl)-2,4-dioxo-1(2H)-pyrimidinyl]methyl}-4-fluorobenzonitrile; English For: 2-[6-(3(R)-aminopiperidin-1-yl)-3-(3,4-dimethoxybenzyl)-2,4-dioxo-3,4-dihydr o-2H-pyrimidin-1-ylmethyl ]-4-fluorobenzonitrile; One or two of the compounds 1-3 have DPP-4 inhibitory activity, and the compounds 1-3 referred to here can be the compounds 1-3 themselves and the pharmaceutically acceptable salts of the compounds 1-3 One of such chemical equivalents, but not limited to the above chemical equivalents. 3. according to the described DPP-4 inhibitor of claim 1 or 2, it is characterized in that The DPP-4 inhibitor includes pharmaceutically acceptable drug adjuvant or carrier. 4. DPP-4 inhibitor according to claim 3, is characterized in that The DPP-4 inhibitor includes one or more of diluents, wetting agents, binders, disintegrants, lubricants, regulators and other pharmaceutically or food acceptable auxiliary materials. 5. according to the described DPP-4 inhibitor of claim 3 or 4, it is characterized in that One or two of the compounds 1-3 or their corresponding pharmaceutically acceptable salts can be mixed with pharmaceutically acceptable pharmaceutical adjuvants or carriers to make tablets, capsules, Oral liquid, granules, pills or injections, but not limited to the above dosage forms. 6. according to the described DPP-4 inhibitor of claim 3 or 4, it is characterized in that A single dose of the pharmaceutical composition contains active ingredients of one or two of the compounds 1-3 or pharmaceutically acceptable salts thereof, and the effective content of the pharmaceutical composition is 1-95%. 7. the application of the arbitrary described DPP-4 inhibitor of claim 1-6 in the treatment type 2 diabetes medicine, it is characterized in that: The DPP-4 inhibitor uses one or more of the following three compounds or pharmaceutically acceptable salts corresponding to the following three compounds as active ingredients, and the structural formulas of the three compounds are as follows: The type 2 diabetes includes diabetes caused by the development of insulin resistance or diabetes caused by a defect in insulin secretion. 8. The application according to claim 7, characterized in that The medicine includes pharmaceutically acceptable pharmaceutical auxiliary materials. 9. The application according to claim 4, characterized in that The medicine includes diluents, wetting agents, binders, disintegrants, lubricants, regulators and other pharmaceutically or food-acceptable adjuvants; One or two of the compounds 1-3 can be mixed with pharmaceutically acceptable drug carriers to make tablets, capsules, oral liquids, granules, pills or injections for the treatment of related diseases, but not limited to above dosage forms. 10. The application according to claim 4 or 9, characterized in that A single dose of the pharmaceutical composition contains one or more active ingredients of compounds 1-3 or their pharmaceutically acceptable salts, and the effective mass content of the pharmaceutical composition is 1-95%.