WO2015027911A1

WO2015027911A1 - Compound for treating diabetes

Info

Publication number: WO2015027911A1
Application number: PCT/CN2014/085270
Authority: WO
Inventors: 谢永美; 魏于全; 耿福能
Original assignee: 四川好医生药业集团有限公司; 四川大学
Priority date: 2013-08-30
Filing date: 2014-08-27
Publication date: 2015-03-05
Also published as: CN103435612B; CN103435612A

Abstract

Provided are a compound as presented in formula I, or pharmaceutically acceptable salts, hydrates, or solvates thereof. Also provided are new uses of the compound, or pharmaceutically acceptable salts, hydrates, or solvates thereof. The deuterated compound prepared in the present invention can substantially improve the weight, fasting glucose, transaminase, and fasting serum insulin of obesity patients, increase insulin sensitivity, improve lipid levels and oral glucose tolerance, and effectively treat fibrosis-related diseases. The compound can be well absorbed in body and has high bioavailability that is conducive for medical effectiveness, and a prolonged half-life, thereby reducing the number of administrations, lowering toxic side effects, and providing a new, safe, and reliable option for clinical medicine.

Description

Compound for treating diabetes

The present invention relates to a method of treating diabetes.

Background technique

According to statistics, the number of patients with diabetes in China has reached 92.4 million, ranking first in the world (Yang W, et al. N Engl J Med, 2010, 362(12): 1090-1 101) _o Chinese diabetes treatment costs per year As much as 173.4 billion yuan, direct medical expenses due to diabetes account for 13% of China's total medical expenditure (Alcorn T, et al. Lancet, 2012, 379 (9833): 2227-2228). It can be seen that diabetes not only seriously jeopardizes the health of the people, but also brings a heavy economic burden to the country. Therefore, prevention and treatment of diabetes is an urgent task.

Depending on the etiology and clinical manifestations, diabetes is divided into four main types: insulin-dependent (type 1), non-insulin-dependent (type 2), malnutrition-related and secondary diabetes. Among them, type 2 diabetes (T2D) accounts for more than 90% of the total number of people with diabetes. Therefore, the research of T2D therapeutic drugs is the focus and hot spot. At present, T2D therapeutic drugs commonly used in clinical practice include insulin, biguanide, sulfonylurea, glycosidase inhibitor, thiazolyldione, glitazone and glinide, but they often have different degrees of side effects, such as Hypoglycemia, weight gain, cardiovascular side effects, etc. Therefore, the development of new anti-diabetic drugs that act on new targets, avoid the side effects of traditional anti-diabetic drugs, and protect pancreatic β-cells has become a hot spot at home and abroad. After efforts, a number of anti-diabetic drugs with new mechanisms of action have been approved for marketing, such as: dipeptidyl peptidase-4 (DPP-4) inhibitor and glucagon-like peptide-1 (GLP-1). Body agonists, such drugs have the disadvantage of causing side effects such as pancreatitis, nasopharyngitis, respiratory infections, headaches, allergies and the like. Therefore, the research on high-efficiency and low-toxic T2D innovative drugs is of great significance.

Summary of the invention

It is an object of the present invention to provide a novel compound for the treatment of diabetes. Another object of the invention is to provide the use of this novel compound. Specifically, the present invention provides a compound of Formula I, or a pharmaceutically acceptable salt, hydrate or solvate thereof,

Formula I

Rl R2 R3 are each independently selected from H, hydrazine, C1 C3 fluorenyl, partially deuterated or fully deuterated C1 C3 fluorenyl;

c=c

R4 is selected from -CH - H H C≡C or not;

R5 R6 are independently selected from H

Wherein, R7~R9 are independently selected from the group consisting of H, hydrazine, halogen, C 1~C4 Or, R5, CH, R6, and its associated N-N NR _{12 -} NON

— NN ^R 13

, R11 is selected from a non-deuterated, partially or fully deuterated C1~C4 sulfhydryl group, and R12 is selected from H, C1-C4 sulfhydryl groups,

a thiol group selected from H or C1 to C4;

0

R15 is selected from H, C1 C4 fluorenyl or decyloxy, _C-R _16;

R16 is selected from the group consisting of C1 to C4.

Further, R1, R2, and R3 are each independently selected from the group consisting of H, 氘, C1 C3 垸, partial or full C C1 C3 垸

R4 is selected from

Rio

R ₅ and R ₆ together with the N to which they are bonded form an OR, and R 10 and R 11 are selected from C 1 to C 4 alkyl groups which are undeuterated, partially or fully deuterated;

Wherein at least one of R1, R2, R3, R10, R11 is 氘 or 氘.

—— c ⁼ c—

Further, R4 is selected from ^H H . Further, the thiol group of C1 to C3 and C1 C4 is a methyl group or an ethyl group.

R1, R2 and R3 are each independently selected from H or hydrazine; R10 and R11 are each independently selected from methyl group,

Triterpene methyl.

Further, the compound is selected from one of the following:

0 s;: ; , ... , AA, D into ■ , Ί , ,

::

D1 D3

D ■ 0 0 one.

A , 00 3⁄4 . .',. 3⁄4 ^

ΌϋΗ _3. K ^¾

f, n ', '

'CD,

4, · D3⁄4

D1S D'! 017

Preferably, the compound is selected from the group consisting of

Wherein the pharmaceutically acceptable salt is an acid salt, a hydrobromide salt, a phosphate, a sulfate, a methanesulfonate, a p-toluenesulfonate, a citrate, a benzoate, or a rich salt of the compound. Citrate, succinate, acetate.

The research of the invention shows that the salt of the above compound can also lower the blood sugar of the type 2 diabetes, improve the oral glucose tolerance, and has obvious therapeutic effect on the type 2 diabetic fatty liver, thereby effectively reducing the deposition of glycogen and collagen in the kidney, thereby May delay the progression of glomerular sclerosis and tubular fibrosis in diabetic nephropathy.

The present inventors have also found that the salt formation of the above compounds increases the water solubility of the compound more or less, and is also advantageous for improving bioavailability. Among them, different compounds with different acids will have different effects on the bioavailability of the drug. In a specific embodiment of the present invention, the mesylate or p-toluenesulfonate of the compound has a higher bioavailability than other salts. To be significant, for example, the mesylate salt of the D1 compound or the p-toluenesulfonate of the D2 compound.

The present invention also provides the use of the above compound, or a pharmaceutically acceptable salt, hydrate or solvate thereof, for the manufacture of a medicament for improving insulin sensitivity, protecting liver and reducing enzymes, or improving blood sugar, blood fat or body weight.

The hepatoprotective enzyme of the invention can be used for preventing or treating fatty liver or diabetes with fatty liver.

Further, the medicament is a medicament for treating diabetes, hyperlipemia, obesity or fatty liver.

Further, the fatty liver is diabetic fatty liver or primary fatty liver.

Further, the drug is a drug for alleviating or treating diabetic nephropathy.

The present invention also provides the use of the above compound or a pharmaceutically acceptable salt, hydrate or solvate thereof for the preparation of a medicament for treating pulmonary fibrosis or liver fibrosis.

The present invention also provides a pharmaceutical composition prepared by using the above compound or a pharmaceutically acceptable salt, hydrate or solvate thereof as an active ingredient, together with pharmaceutically acceptable excipients or auxiliary ingredients. preparation.

Wherein the preparation is an oral, sublingual, topical, implant, inhalation, injection or transdermal absorption preparation.

The pharmaceutically acceptable excipient of the present invention means a substance contained in a dosage form other than the active ingredient, including but not limited to a filler (diluent), a lubricant (glidant or anti-adhesive), a dispersing agent, Wetting agents, binders, conditioners, solubilizers, antioxidants, bacteriostats, emulsifiers, disintegrators, and the like. The binder comprises syrup, gum arabic, gelatin, sorbitol, tragacanth, cellulose and derivatives thereof (such as microcrystalline cellulose, sodium carboxymethylcellulose, ethylcellulose or hydroxypropylmethylcellulose) , gelatin paste, syrup, starch slurry or polyvinylpyrrolidone; fillers include lactose, powdered sugar, dextrin, starch and its derivatives, cellulose and its derivatives, inorganic calcium salts (such as calcium sulfate, calcium phosphate , calcium hydrogen phosphate, precipitated calcium carbonate, etc.), sorbitol or glycine; lubricants include micronized silica gel, magnesium stearate, talc, aluminum hydroxide, boric acid, hydrogenated vegetable oil, polyethylene glycol, etc.; disintegrants included Starch and its derivatives (such as sodium carboxymethyl starch, sodium starch glycolate, pregelatinized starch, modified starch, hydroxypropyl starch, corn starch, etc.), polyvinylpyrrolidone or microcrystalline cellulose; Containing sodium dodecyl sulfate, water or alcohol; antioxidants include sodium sulfite, sodium bisulfite, sodium metabisulfite, dibutyl benzoic acid, etc.; bacteriostatic agent containing 0.5% phenol, 0.3% cresol 0.5% chlorobutanol, etc.; the regulator includes hydrochloric acid, citric acid, potassium hydroxide (sodium), sodium citrate and a buffer (including sodium phosphate and disodium hydrogen phosphate); the emulsifier comprises a poly Sorbate-80, sorbitan acid, Pluronic F-68, lecithin, soybean phospholipid, etc.; solubilizers include Tween-80, bile, glycerin, and the like.

The pharmaceutically acceptable auxiliary component, which has a certain physiological activity, but the addition of the component does not change the dominant position of the above compound or derivative in the course of disease treatment, but only plays an auxiliary effect, and these auxiliary functions are only The use of known activity of this component is an adjuvant treatment that is commonly used in the medical field. If the above auxiliary components are used in combination with the compound of the present invention, it should still fall within the scope of protection of the present invention.

The deuterated compound prepared by the invention can obviously improve the body weight, fasting blood glucose, transaminase and fasting serum insulin of obese patients, improve insulin sensitivity, improve blood lipid index and oral glucose tolerance; the compound has good absorption in vivo and high bioavailability, It is beneficial to the exertion of the drug, and the half-life is prolonged, which can reduce the number of administrations, reduce the side effects, and provide a safe and reliable new choice for clinical use.

It is apparent that various other modifications, substitutions or changes can be made in the form of the above-described embodiments of the present invention without departing from the spirit and scope of the invention.

The above content of the present invention will be further described in detail below by way of specific embodiments. But should not understand this The scope of the above-described subject matter of the present invention is limited to the following embodiments. Any technique implemented based on the above description of the present invention is within the scope of the present invention.

DRAWINGS

Figure 1 Liver HE, where NS: saline group; Normal: normal group; SIS3: control compound

Figure 2 Daily basic food intake

Figure 3 weight change time

Figure 4 fasting blood glucose

Figure 5 glucose tolerance

Figure 6 Liver HE (x200 times)

Figure 7 Adipose tissue HE

Figure 8 Kidney PAS staining (x400 times)

Figure 9 Kidney Masson staining (x400 times)

Figure 10 Lung HE staining

Figure l lMasson staining

Figure 12 Comparison of total cells in BALF

Detailed ways

Example 1 Synthesis of Compound D1

Synthesis of compound 2

Under N ₂ protection, DMF was added to a three-necked flask containing 5 g of Compound 1 and 1.25 g of NaH. After stirring at 0 ° C for 10 min, CD ₃ I was slowly added dropwise and stirred at room temperature overnight. After the completion of the reaction, the reaction was completed, and water was slowly added dropwise to the reaction flask, followed by extraction with EA, and the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated, and then purified to give a pale yellow solid 3.95 g, yield 78.9%

MR (400 MHz, CDC1 ₃ ) δ 8.36 (t, 1H), 7.92 (q, 1H), 7.57 - 7.42 (m, 5H), 7.11-7.07 (m, 1H), 6.52 (s, 1H).

Synthesis of Compound 3

Under the protection of N ₂ , 3.94 g of Compound 2 was dissolved in DMF, and 2.4 mL of POCl ₃ was slowly added dropwise at 0 ° C. After stirring for 3 hours, the reaction was monitored by TLC. After the reaction of the raw materials was completed, 50 mL of ice water was added to the reaction flask, and the pH was adjusted to be greater than 14 with a 1 mol/L NaOH solution, and solids were precipitated, suction filtered, and dried to give a pale yellow solid 3.85 g, yield 86% 1H MR (400 MHz, CDC) δ 9.77 (s, 1H), 8.67-8.65 (m, 1H), 8.47-8.45 (m, 1H), 7.60-7.53 (m, 5H), 7.32-7.29 (m, 1H).

Synthesis of Compound 4 After 1.82 g of Compound 3 and 1.0 g of malonic acid were dissolved in 30 mL of pyridine, 4.8 mL of hexahydropyridine was added, and the reaction was refluxed for 4 h. After the reaction was stopped, water was added, and the pH was adjusted to 6 with a 1 mol/L HCl solution, and the solid was precipitated, suction filtered, and dried to give a pale yellow solid, 1.56 g, yield 73% ₀ 1H MR (400 MHz, DMSO- ₆ ) δ 12.04 ( s, IH), 8.43-8.41 (m, 2H), 7.68 - 7.57 (m, 5H), 7.44 (d, IH), 7.32 (q, IH), 6.41 (d, 1H).

Synthesis of Compound 6 (D1)

27.8 mg (0.1 mol) of compound 4, 27 mg (0.2 mmol) of HOBT, 57.5 mg (0.3 mmol) of EDCI, dissolved in 2 mL of DMF, added 0.025 mL of triethylamine, stirred at room temperature for 30 minutes; then added 24 mg (0.12) Methyl) Compound 5, stirred at room temperature overnight. After the completion of the reaction, water was added, and EA was extracted three times (15 mL each time). The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, Ipi MR (400 MHz, CDC1 ₃ ) δ 8.44 (d, IH), 8.24 (s, IH), 7.77 (d, IH), 7.55-7.44 (m, 5H), 7.24 (s, IH), 6.87 (d , IH), 6.64 (s, 2H), 4.73 (s, 2H), 3.87 (s, 6H), 3.86-3.83 (m, 2H), 2.86-2.82 (m, 2H).

Example 2 Synthesis of Compound D2

-

Oh,

Synthesis of Compound 7

Under a nitrogen atmosphere, DMF was added to a three-necked flask containing 5 g of Compound 1 and 1.25 g of NaH. After stirring at 0 ° C for 10 min, CH ₃ I was slowly added dropwise and stirred at room temperature overnight. After the completion of the reaction, water was slowly added dropwise to the reaction flask, followed by extraction with EA. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and then evaporated to give a pale yellow product of about 4.05 g, yield of 79.5%.

Synthesis of Compound 8

Under the protection of nitrogen, 4.05 g of compound 7 was dissolved in DMF, and 2.4 mL of P0C1 ₃ was slowly added dropwise at 0 ° C. After stirring for 3 h, the reaction was monitored by TLC. After the reaction of the starting material was completed, 50 mL of ice water was added to the reaction flask, and the pH was adjusted to be more than 14 with a 1 mol/L NaOH solution, and the solid was precipitated, suction filtered, and dried to give a pale yellow solid, 3.83 g, yield 83.7%.

Synthesis of compound 9

After 1.82 g of Compound 8 and 1.0 g of malonic acid were dissolved in 30 mL of pyridine, 4.8 mL of hexahydropyridine was added and refluxed for about 4 hours. After the reaction was stopped, water was added, and the pH was adjusted to 6 with a 1 mol/L HCl solution to precipitate a solid, which was filtered and dried to give a pale yellow solid, 1.61 g, yield 75.2%.

Synthesis of Compound 11

27.8 mg (0.1 mmol) of compound 9, 27 mg (0.2 mmol) of HOBT and 57.5 mg (0.3 mmol) of EDCI, dissolved in 2 mL of DMF, and then added 0.05 mL of triethylamine, stirred at room temperature for 30 minutes; then added 29.5 mg ( 0.12 mmol) 1,2,3,4-tetrahydro-6,7-isoquinolindiol hydrobromide (10), stirred at room temperature overnight. TLC monitoring, after completion of the reaction, adding water, EA extraction After three times (15 mL each time), the organic layer was washed with saturated brine, dried over anhydrous sodium sulfate, dried and evaporated.

Synthesis of Compound 12 (D2)

Under the protection, the above crude product and an appropriate amount of NaH were dissolved in DMF, stirred at 0 ° C for 30 minutes, and then slowly added with CD ₃ I, and reacted at room temperature. After the completion of the reaction, water was slowly added dropwise to the reaction flask, followed by extraction with EA, and the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and evaporated 1H MR (400 MHz, CDC1 ₃ ) δ 8.44 (d, IH), 8.25 (s, IH), 7.77 (d, IH), 7.55-7.44 (m, 5H), 7.24 (s, IH), 6.87 (d , IH), 6.64 (s, 2H), 4.73 (s, 2H), 3.87 -3.83 (m, 2H), 3.76 (s, 3H), 2.86-2.82 (m, 2H).

Example 3

Synthesis of compound 13

Take 28.1 mg (0.1 mmol) of compound 4, 27 mg (0.2 mmol) of HOBT, 57.5 mg (0.3 mmol) of EDCI, dissolve with 2 mL of DMF, add 0.05 mL of triethylamine, stir at room temperature for 30 minutes; then add 29.5 mg (0.12) Methyl) 1,2,3,4-tetrahydro-6,7-isoquinolindiol hydrobromide (10), stirred at room temperature overnight. After the reaction was completed, water was added, and EA was extracted three times (15 mL each time). The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, Synthesis of Compound 14 (D3)

Under the protection of N ₂ , the above crude product and an appropriate amount of NaH were dissolved in DMF, stirred at 0 ° C for 30 minutes, and then slowly added with CD ₃ I, and reacted at room temperature. After completion of the reaction, water was slowly added dropwise to the reaction flask, followed by extraction with EA. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, 1H MR (400 MHz, CDC1 ₃ ) δ 8.44 (d, IH), 8.25 (s, IH), 7.77 (d, IH), 7.59-7.40 (m, 5H), 7.24 (s, IH), 6.88 (d , IH), 6.65 (s, 2H), 4.73 (s, 2H), 3.87-3.83 (m, 2H), 2.86 -2.82 (m, 2H). Example 4 Synthesis of Compound D4 Strict HO m _ν Λ, _{: ΟΗ} '-' - ' _丫 f _{Γ| s}

Synthesis of CX ~~ and VIII n compounds 16

27.8 mg (0.1 mmol) of compound 9, 27 mg (0.2 mmol) of HOBT, 57.5 mg (0.3 mmol) of EDCI, dissolved in 2 mL of DMF, added 0.05 mL of triethylamine, stirred at room temperature for 30 minutes; then added 26 mg (0.12 mmol) 1,2,3,4-Tetrahydro-6-methoxy-7-isoquinolinol hydrochloride (15), stirred at room temperature overnight. TLC was monitored. After completion of the reaction, water was added and EA was extracted three times (15 mL each time). The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and then evaporated. Synthesis of Compound 17 (D4)

Under the protection, the above crude product and an appropriate amount of NaH were dissolved in DMF, and stirred at 0 ° C for 30 minutes, and then CD ₃ I was slowly added dropwise. The reaction was carried out at room temperature. After the completion of the reaction, water was slowly added dropwise to the reaction flask, followed by extraction with EA. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, 1H MR (400 MHz, CDC1 ₃ ) δ 8.45 (d _: 1H), 8.24 (s, 1H), 7.76 (d, 1H), 7.60-7.42 (m, 5H), 7.24 (s, 1H), 6.87 (d , 1H), 6.64 (s, 2H), 4.73 (s, 2H) _: 3.89 (s, 3H), 3.87-3.84 (m, 2H), 3.77 (s, 3H), 2.87 -2.83 (m, 2H).

Example

Synthesis of Compound 18

28.1 mg (0.1 mmol) of compound 4, 27 mg (0.2 mmol) of HOBT, 57.5 mg (0.3 mmol) of EDCI, dissolved in 2 mL of DMF, added 0.05 mL of triethylamine, stirred at room temperature for 30 minutes; then added 26 mg (0.12 mmol) 1,2,3,4-Tetrahydro-6-methoxy-7-isoquinolinol hydrochloride (15), stirred at room temperature overnight. TLC was monitored. After completion of the reaction, water was added and EA was extracted three times (15 mL each time). The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and then evaporated. Synthesis of Compound 19 (D5)

Under the protection of N ₂ , the above crude product and an appropriate amount of NaH were dissolved in DMF, stirred at 0 ° C for 30 minutes, and then slowly added with CD ₃ I, and reacted at room temperature. After completion of the reaction, water was slowly added dropwise to the reaction flask, followed by extraction with EA, and the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, 1H MR (400 MHz, CDC1 ₃ ) δ 8.44 (d, 1H), 8.25 (s, 1H), 7.76 (d, 1H), 7.60-7.42 (m, 5H), 7.24 (s, 1H), 6.87 (d , 1H), 6.65 (s, 2H), 4.73 (s, 2H), 3.87-3.83 (m, 2H), 3.85 (s, 3H), 2.87 - 2.84 (m, 2H).

Example 6 Synthesis of Compound D6

Synthesis of Compound 21

28.1 mg (0.1 mmol) of compound 4, 27 mg (0.2 mmol) of HOBT, 57.5 mg (0.3 mmol) of EDCI, dissolved in 2 mL of DMF, added 0.05 mL of triethylamine, stirred at room temperature for 30 minutes; then added 26 mg (0.12 mmol) 1,2,3,4-Tetrahydro-7-methoxy-6-isoquinolinol hydrochloride (20), stirred at room temperature overnight. TLC was monitored. After completion of the reaction, water was added and EA was extracted three times (15 mL each time). The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and then evaporated. Synthesis of Compound 22 (D6)

Under the protection, the above crude product and an appropriate amount of NaH were dissolved in DMF, stirred at 0 ° C for 30 minutes, and then slowly added with CD ₃ I, and reacted at room temperature. After completion of the reaction, water was slowly added dropwise to the reaction flask, followed by extraction with EA, and the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, 1H MR (400 MHz, CDC1 ₃ ) δ 8.45 (d,

1H), 8.25(s, 1H), 7.77 (d, 1H), 7.61-7.45 (m, 5H), 7.24 (s, 1H), 6.88 (d, 1H), 6.65 (s, 2H), 4.73 (s , 2H),

3.87-3.83 (m, 2H), 3.85 (s, 3H), 2.87 - 2.84 (m, 2H).

Synthesis of Compound D7 of Example 7

S3

Synthesis of Compound 23

Take 28.1 mg (0.1 mmol) of compound 4, 27 mg (0.2 mmol) of HOBT, 57.5 mg (0.3 mmol) of EDCI, dissolve with 2 mL of DMF, add 0.05 mL of triethylamine, stir at room temperature for 30 minutes; then add 26 mg (0.12 mmol) 1,2,3,4-Tetrahydro-7-methoxy-6-isoquinolinol hydrochloride (20), stirred at room temperature overnight. TLC was monitored. After completion of the reaction, water was added and EA was extracted three times (15 mL each time). The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and then evaporated. Synthesis of Compound 24 (D7)

Under the protection, the above crude product and an appropriate amount of NaH were dissolved in DMF, and stirred at 0 ° C for 30 minutes, and then CD ₃ I was slowly added dropwise.

M... - Reaction at room temperature. After completion of the reaction, water was slowly added dropwise to the reaction flask, followed by extraction with EA, and the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, 1H MR (400 MHz, CDC1 ₃ ) δ 8.44 (d, IH), 8.25 (s, IH), 7.76 (d, IH), 7.60-7.42 (m, 5H), 7.23 (s, IH), 6.87 (d , (H, 2H)

Example 8 Synthesis of Compound D8

COOH 0,. c3⁄4, Π 27.8 mg (0.1 mmol) of compound 4, 27 mg (0.2 mmol) HOBT, 57.5 mg (0.3 mmol) EDCI, dissolved in 2 mL of DMF, added 0.05 mL of triethylamine, stirred at room temperature for 30 minutes. Then, 0.014 mL (0.12 mmol) of N-methylpiperazine was added, and the mixture was stirred at room temperature overnight. After TLC monitoring, after completion of the reaction, water was added, and EA was extracted three times (15 mL each time). The organic phase was washed with saturated brine and dried over anhydrous sodium sulfate. . NMR (400 MHz, CDC1 ₃ ) δ 8.42 (dd, IH), 8.20 (dd, IH), 7.74 (d, IH), 7.58-7.41 (m, 5H), 7.26-7.21 (m, IH), 6.80 ( d, IH), 3.68 (d, 4H), 2.43 (s, 4H), 2.32 (s, 3H).

Example 9 Synthesis of Compound D9

27.8 mg (0.1 mmol) of compound 4, 27 mg (0.2 mmol) of HOBT, 57.5 mg (0.3 mmol) of EDCI, dissolved in 2 mL of DMF, added 0.025 mL of triethylamine, stirred at room temperature for 30 minutes; then added O.OllmL ( 0.12 mmol) Hexahydropyridine was stirred at room temperature overnight. After the completion of the reaction, water was added, and EA was extracted three times (15 mL each time). The organic phase was washed with saturated brine and dried over anhydrous sodium sulfate. 1H MR (400 MHz, CDC1 ₃ ) δ 8.42 (d, IH), 8.23 (d, IH), 7.72 (d, IH), 7.58-7.41 (m, 5H), 7.25- 7.20 (m, IH), 6.84 (d, IH), 3.59 (m, 4H), 1.81-1.45 (m, 6H). Example 10 Synthesis of Compound D10

27.8 mg (0.1 mmol) of compound 4, 27 mg (0.2 mmol) of HOBT, 57.5 mg (0.3 mmol) of EDCI, dissolved in 2 mL of DMF, added 0.025 mL of triethylamine, stirred at room temperature for 30 minutes; then added O.OllmL ( 0.12 mmol) morpholine, stirred at room temperature overnight. After the reaction was completed, water was added, and EA was extracted three times (15 mL each time). The organic phase was washed with saturated brine and dried over anhydrous sodium sulfate 1H MR (400 MHz, CDC1 ₃ ) δ 8.43 (d, 1H), 8.21 (d, 1H), 7.76 (d, 1H), 7.59-7.38 (m, 5H), 7.25-7.20 (m, 1H), 6.76 (d, 1H), 3.71 (s, 4H), 3.66 (s, 4H).

Example 11 Synthesis of Compound Dll

27.8 mg (0.1 mmol) of compound 4, 27 mg (0.2 mmol) of HOBT, 57.5 mg (0.3 mmol) of EDCI, dissolved in 2 mL of DMF, added 0.025 mL of triethylamine, stirred at room temperature for 30 minutes; then added 0.015 mL (0.12) Methyl) N-methylhomopiperazine, stirred at room temperature overnight. After the completion of the reaction, water was added, and EA was added three times (15 mL each time), and the organic layer was washed with saturated brine, dried over anhydrous sodium sulfate, and then concentrated to give a white solid product 26.4 mg, yield 70%.

1H NMR (400 MHz, CDC1 ₃ ) 5 8.42 (d, 1H), 8.17 (t, 1H), 7.77 (d, 1H), 7.61-7.38 (m, 5H), 7.25-7.19 (m, 1H), 6.76 (t, 1H), 3.84-3.62 (m, 4H), 2.75-7.53 (m, 4H), 2.39 (s, 3H), 2.07-1.88 (m, 2H).

Example 12 Synthesis of Compound D12

OCH;

,COOH ₍ ..ί ^: ' 0 ί=:= ^; _}

丫 3⁄4U "OCH

29 30 (012) 28 mg (0.1 mmol) of compound 4, 27 mg (0.2 mmol) of HOBT, 57.5 mg (0.3 mmol) of EDCI, dissolved in 1 mL of DMF, then added 0.025 mL of triethylamine, stirred at room temperature for 0.5 h and then added 19 mg ( 0.12 mmol) 3,5-dimethoxyaniline (29), stirring was continued overnight. After adding 20 ml of EA, it was washed with a saturated NaCI solution, dried over anhydrous sodium sulfate, filtered, and concentrated, and then purified to give a yellow powder 30 mg, yield 71%. 1H MR (400 MHz, CDC1 ₃ )5 8.42 (dd, 1H), 8.23 (d, 1H), 7.78 (d, 1H), 7.57-7.49 (m, 3H), 7.46-7.37 (m, 3H), 7.21 (dd, 1H), 6.88 (s, 2H), 6.52 (d, 1H), 6.23 (t, 1H), 3.77 (s, 6H). Example 13 Synthesis of Compound D13 Fly A Production-v

, ί :

I 丄

'Ρ ί

31 28 mg (0.1 mmol) of compound 4, 27 mg (0.2 mmol) of HOBT, 57.5 mg (0.3 mmol) of EDCI, dissolved in 1 mL of DMF, added 0.025 mL of triethylamine, stirred at room temperature for 0.5 h, then added 15 mg (0.12 mmol). Chloroaniline (31) was stirred overnight. After adding 20 ml of EA, it was washed with a saturated NaCI solution, dried over anhydrous sodium sulfate, filtered, and concentrated, and then purified to give a yellow powder, 24 mg, yield 61%. 1H employment R (400 MHz, CDC1 ₃ ) δ 8.39 (dd, IH), 8.14 (d, IH), 7.83 (s, IH), 7.76 (d, IH), 7.63-7.46 (m, 5H), 7.41- 7.33 (m, 2H), 7.30-7.20 (m, 2H), 7.16-7.10 (m, IH), 6.53 (d, 1H). Example 14 Synthesis of Compound D14

Synthesis of 33 3 3S(D14) Compound 34

278 mg (1 mmol) of compound 4 was placed in a flask, force B 575 mg (3 mmol) EDCI, 270 mg (2 mmol) HOBT, dissolved in 10 ml of DMF, and then 0.25 ml (1.5 mmol) of triethylamine was added dropwise, stirred at room temperature for 30 min, then 224 mg was added. (1.2 mmol) N-Boc piperazine (33), stirred at room temperature overnight. To the reaction system, 200 ml of ethyl acetate was added, and the mixture was washed twice with a saturated aqueous solution of sodium chloride, dried over anhydrous sodium sulfate, and filtered, and then concentrated to give white powder, 400 mg, yield 89%.

Synthesis of Compound 35 (D14)

223 mg (0.5 mmol) of compound 34 was placed in a 25 ml round bottom flask, first dissolved in 2 ml of dry dichloromethane, and 3 ml of dry dichloromethane and 1.25 ml of trifluoroacetic acid were mixed and added dropwise to the reaction system at room temperature. Stir for 10 h. Saturated NaHC0 ₃ solution and extracted with of dichloromethane, washed with saturated NaCl solution, dried over anhydrous sodium sulfate, filtered and concentrated to give a white powder 130mg, yield 74%. 1H MR (400 MHz, CDCI3) δ 8.43 (d, IH), 8.19 (d, IH), 7.74 (d, IH), 7.58 -7.42 (m, 5H), 7.23 (dd, IH), 6.77 (d, IH), 3.70 (s, 4H), 2.96 (s, 4H).

Example 15 Synthesis of Compound D15

36 3S

Take 18 mg (0.05 mmol) of compound 35 and 19 mg (0.1 mmol) of m-bromoanisole (36), 3 mg (0.0025 mmol) of Pd ₂ (dba) ₃ , 15 mg (0.15 mmol) of sodium tert-butoxide, 4 mg (0.005 mmol) BINAP was placed in a reaction tube, and after replacing nitrogen, 1 ml of toluene was added and reacted at 100 ° C overnight. After concentration and column chromatography, a white powder (yield: 18 mg, yield: 79%). 1H MR (400 MHz, CDCI3) δ 8.43 (d, IH), 8.22 (d, IH), 7.77 (d, IH), 7.59-7.38 (m, 5H), 7.23 (dd, IH), 7.19 (d, IH), 6.83 (d, IH), 6.55 (d, IH), 6.47 (d, 2H), 3.91-3.75 (m, 7H), 3.29-3.16 (m, 4H). Example 12 Synthesis of Compound D12

35 3β

Take 18 mg (0.05 mmol) of compound 35 and 20 mg (0.1 mmol) of iodobenzene (38), 3 mg (0.0025 mmol) of Pd ₂ (dba) ₃ , 15 mg (0.15 mmol) of sodium tert-butoxide, 4 mg (0.005 mmol) of BINAP In a reaction tube, after replacing nitrogen gas, 1 ml of toluene was added, and the reaction was carried out at 100 ° C overnight. After concentration, the column was passed to give a brown powder (yield: 17 mg, yield 77%). 1H MR (400 MHz, CDC1 ₃ ;>5 8.43 (d, IH), 8.25-8.19 (m, IH), 7.77 (d, IH), 7.58-7.40 (m, 5H), 7.29 (t, 2H), 7.25-7.20 (m, IH), 6.94 (d, 2H), 6.91 (s, IH), 6.84 (d, IH), 3.83 (s, 4H), 3.27-3.15 (m, 4H).

Example 17 Synthesis of Compound D17

40 41 ξ01 ?5 18 mg (0.05 mmol) of compound 35 and 20 mg (0.1 mmol) 4-bromoacetophenone (40), 3 mg (0.0025 mmol) Pd ₂ (dba) ₃ , 15 mg (0.15 mmol) tert-butanol Sodium, 4 mg (0.005 mmol) of BINAP was placed in a reaction tube, and after replacing nitrogen, 1 ml of toluene was added, and the reaction was carried out at 100 ° C overnight. After concentration, the column was passed to give a pale yellow powder (yield: 77%).

Example 18 Synthesis of Compound D18

Compound 42 synthesis

0.37 g (1.8 mmol) of compound 2 was placed in a 25 ml round bottom flask. After replacing nitrogen, 1 ml of DMF was added to dissolve. 0.5 ml of DMF dissolved trifluoroacetic anhydride was added dropwise at 0 ° C, and stirring was continued for 5 hours. It was mixed with 3 ml of water, extracted with EA, and concentrated. 5 ml of a 10% aqueous NaOH solution and 5 ml of methanol were added, and the mixture was stirred at 100 ° C overnight. After cooling to room temperature, add water and wash with EA. The aqueous layer was acidified with 1 M HCl and filtered. The compound 42, white powder 300 mg was obtained in a yield of 65%.

Synthesis of Compound 43 (D18)

25 mg (0.1 mmol) of compound 42 was added, 27 mg (0.2 mmol) of hydrazine, 57.5 mg (0.3 mmol) of EDCI was added, dissolved in 1 ml of DMF, triethylamine 25 μl was added, and stirred at room temperature for 0.5 h, then 15 mg (0.12 mmol) of p-chloro The aniline was stirred overnight. After adding 20 ml of EA, it was washed with a saturated NaCl solution, dried over anhydrous sodium sulfate, filtered, and concentrated, and then purified by column chromatography to give white powder 20 mg, yield 55 1H MR (400 MHz, CDC1 ₃ ) δ 8.47 (dd, IH), 8.45- 8.39 (m, IH), 7.93 (d, IH), 7.49 -7.39 (m, 5H), 7.37 (d„ IH), 7.28 (dd, 3H).

Example 19 Synthesis of Compound D19

25 mg (0.1 mmol) of compound 42 was added, and 27 mg (0.2 mmol) of hydrazine, 57.5 mg (0.3 mmol) of EDCI was added, dissolved in 1 ml of DMF, and triethylamine 25 μl was added thereto. After stirring at room temperature for 0.5 h, 15 mg (0.12 mmol) of 3 was added. 5-Dimethylaniline (44) was stirred overnight. After adding 20 ml of EA, it was washed with a saturated NaCI solution, dried over anhydrous sodium sulfate, filtered, and evaporated 1H MR (400 MHz, CDC1 ₃ ) δ 8.76 (dd, 1H), 8.57 (dd, 1H), 8.52 (dd, 1H), 8.43 (dd, 1H), 8.03 (d, 1H), 7.74-7.65 (m , 3H), 7.63-7.44 (m, 4H), 2.22 (s, 6H).

Example 20 Compound

25 mg (0.1 mmol) of compound 42 was added, 57.5 mg (0.3 mmol) of EDCI, 27 mg (0.2 mmol) of HOBT was added, dissolved in 1 ml of DMF, and triethylamine 25 μl was added thereto. After stirring at room temperature for 0.5 h, 18 mg (0.12 mmol) of 3 was added. 5-Dimethoxyaniline (29), stirring was continued overnight. After adding 20 ml of EA, it was washed with a saturated NaCl solution, dried over anhydrous sodium sulfate, and concentrated, and then passed to afford white powder 25 mg, yield 64%. 1H MR (400 MHz, CDCI3) δ 8.49 (d, 1H), 8.45 (d, 1H), 7.96 (d, 1H), 7.47-7.30 (m, 9H), 3.66 (s, 6H).

Example 21 Preparation of Dl Hydrochloride

The lg compound D1 was weighed, 20 ml of dichloromethane was added, and the mixture was stirred and dissolved. 80 ml of a 2 M HCl solution of ethyl acetate was added dropwise to the solution under ice-cooling. During the stirring, a yellow solid was precipitated, stirred for 1 hour, filtered, and washed with diethyl ether. After vacuum drying, 0.93 g of D1 hydrochloride was obtained in a yield of 86.2%. Elemental analysis: C, 68.10%; H, 6.27%; N, 8.58% (theoretical: C, 68.21%; H, 6.34%; N, 8.52%)

Example 22 Preparation of D1 p-toluenesulfonate

Weigh lg compound Dl, add 20ml of dichloromethane, stir and dissolve, add 0.41g of p-toluenesulfonic acid to the solution under ice bath, stir to dissolve, add 50ml of anhydrous ether to the solution, and solid precipitate during stirring. Filtration, washing with diethyl ether, and drying in vacuo to give D1 p-toluenesulfonate 1. lg, yield 79.9%. Elemental analysis: C, 66.72%; H, 6.01%; N, 6.78% (theoretical: C, 66.86%; H, 6.09%; N, 6.68%).

Example 23 Preparation of D1 Mesylate

Weigh lg compound Dl, add 20ml of dichloromethane, stir to dissolve, add 0.23g of methanesulfonic acid to the solution under ice bath, add 50ml of anhydrous ether to the solution, solid precipitate during stirring, suction filtration, use ether After washing, vacuum drying of 0.9 g of D1 methanesulfonate, yield 74.4%. Elemental analysis: C, 63.09%; H, 6.11%; N, 7.71% (theoretical: C, 63.02%; H, 6.20%; N, 7.60%) Example 24 Preparation of Dl Benzoate

The lg compound D1 was weighed, 20 ml of dichloromethane was added, stirred and dissolved, 0.29 g of benzoic acid was added to the solution under ice bath, 50 ml of anhydrous diethyl ether was added to the solution, and solids were precipitated during the stirring, suction filtration, and diethyl ether washing. After drying, 0.85 g of D1 mesylate was obtained in a yield of 67.1%. Elemental analysis: C, 71.92%; H, 6.19%; N, 7.34% (theoretical: C, 72.64%; H, 6.27%; N, 7.26%)

Example 25 Preparation of D1 Citrate

Weigh lg compound Dl, add 20ml of dichloromethane, stir and dissolve, add 0.46g citric acid to the solution under ice bath, fully dissolve, add 50ml of anhydrous ether to the solution, precipitate solid during the stirring, suction filtration, The mixture was washed with diethyl ether and dried under vacuum to give 1.17 g of D1 lemon salt, with a yield of 82.5%. Elemental analysis: C, 62.83%; H, 5.89%; N, 6.58% (theoretical: C, 62.95%;

H, 5.90%; N, 6.48%

Example 26 Preparation of D2 Hydrochloride

Weigh lg compound D2, add 20ml of dichloromethane, stir and dissolve, add 80ml of 2M HCl solution in ethyl acetate to the solution under ice bath, precipitate yellow solid during stirring, stir for 1 hour, suction filtration, wash with ether After drying in vacuo, 0.91 g of D2 hydrochloride was obtained, and the yield was 84.3%. Elemental analysis: C, 67.68%; H, 6.87%; N, 8.54% (Theoretical values: C, 67.80%; H, 6.91%; N, 8.47%)

Example 27 Preparation of D2 p-toluenesulfonate

Weigh lg compound D2, add 20 ml of dichloromethane, stir and dissolve, add 0.41 g of p-toluenesulfonic acid to the solution under ice bath, stir to dissolve, add 50 ml of anhydrous ether to the solution, and precipitate solids during the stirring. Filtration, washing with diethyl ether and drying in vacuo gave 1.05 g of D? Elemental analysis: C, 65.82%; H, 6.47%; N, 6.73% (theoretical: C, 66.54%; H, 6.54%; N, 6.65%).

Example 28 Preparation of D2 Mesylate

Weigh lg compound D2, add 20ml of dichloromethane, stir and dissolve, add 0.23g of methanesulfonic acid to the solution under ice bath, add 50ml of anhydrous ether to the solution, solid precipitate during the stirring process, suction filtration, use ether After washing and vacuum drying, 1.02 g of D2 methanesulfonate was obtained in a yield of 84.3%. Elemental analysis: C, 62.09%; H, 6.82%; N, 7.38% (theoretical: C, 62.68%; H, 6.71%; N, 7.56%)

Example 29 Preparation of D2 benzoate

Weighed lg compound D2, added 20 ml of dichloromethane, dissolved and dissolved, added 0.29 g of benzoic acid to the solution under ice bath, and added 50 ml of anhydrous ether to the solution. During the stirring, solids were precipitated, suction filtered, and washed with diethyl ether. D2 mesylate after drying

I.05g, yield 83.3%. Elemental analysis: C, 71.68%; H, 6.79%; N, 7.37% (theoretical: C, 72.27%; H, 6.76%; N, 7.22%)

Example 30 Preparation of D2 Citrate

Weigh lg compound D2, add 20 ml of dichloromethane, stir and dissolve, add 0.46 g of citric acid to the solution under ice bath, dissolve well, add 50 ml of anhydrous ether to the solution, and precipitate solids during the stirring, suction filtration, The mixture was washed with diethyl ether and dried under vacuum to give 1.27 g of D2 lemon salt, with a yield of 89.4%. Elemental analysis: C, 62.72%; H, 6.15%; N, 6.68% (theoretical: C, 62.66%; H, 6.34%; N, 6.45%) The beneficial effects of the present invention will be specifically described below by way of test examples.

Test Example 1 Preliminary pharmacodynamic screening

Five-week-old C57BL/6J mice, male, were fed with common vocabulary and high-fat vocabulary for 10 weeks. Male mice that were successfully modeled were randomly divided into groups of 5, and body weight was recorded. The administration group was given 5 mg/kg of different compounds, and the blank control group and the normal group were given the same volume of physiological saline. The rats were intraperitoneally administered once a day, and after 5 days of continuous administration, the fasting basal blood glucose and glucose tolerance of the mice were examined. Data were presented as mean ± standard deviation, and significant differences were determined by t test. When P 0.05, it is considered to have a significant difference. The test results are shown in Tables 1 and 2.

Effect of the compounds in Table 1 on fasting blood glucose

Note: NS: saline group; Normal: normal group; SIS3: control compound (CAS: 521985-36-4); *- indicates significant difference compared with untreated group (P<0.05)

Table 2 Effect of compounds on glucose tolerance (mg/dL) Group time (min)

0 30 60 120

NS 138.6±10.2 405.6±13.5 427.8±33.7 297 ±39.6

SIS3 103.7±15.0 387±10.0 369.6±41.3 170.4±19.8

Dl 99.6±4.5 457.8±32.7 403.2±28.1 166.4±20.3

D2 108.6±16.2 416.4±24.3 351±22.1 154±18.1

D3 111.6±10.2 334.8±82.0 277.2±86.6 162.2±29.6

D4 130.5 ±8.0 398 ±9.2 400.3±31.5 260.9±15.1

D5 105.9±5.4 365±21.6 428.2 ±18.2 179.5±10.8

D6 118.4±6.5 380.4±223.1 340±12.5 198.2±11.4

D7 101.9±8.3 373.6±12.0 360.5±6.7 168.2±21.0

D8 135±4.5 424 ±39.3 345 ±70.9 181±22.5

D9 120±5.4 435±36.9 407±21.5 235±7.8

D10 94.8±14.5 373.8±22.2 380.4±56.0 229.8±30.5

Dll 150±22.5 421±9.5 428±61.3 265±33.1

D12 148.2±11.7 400.2±17.8 395.4±32.8 246.5±29.0

D13 125.2 ±39.5 410.8±53.4 389.3±15.1 265.8±30.0

D14 103.5±1.3 407.4±20.9 353.4±9.2 208.8±6.5

D15 120±38.7 340 ±50.4 310±29.1 228±52.9

D16 145±18.2 378 ±49.4 335±65.3 220 ±39.1

D17 138±10.5 425±14.7 360±41.3 245±53.1

D18 92.7±14.0 403.2±20.8 360.6±42.2 191.8±81.9

D19 107.4±12.0 396 ±20.0 362.4±31.5 223.2±17.2

D20 168±23.5 435±60.1 407 ±68.1 235±36.0

Normal 70.2±3.1 331.8±37.9 241.8±35.1 123±21.1 Note: NS: blank control group; Normal: normal group; SIS3: control compound (CAS: 521985-36-4)

The results in Tables 1 and 2 show that comparing the normal group with the blank control group, the fasting blood glucose of the model mice is significantly increased, and the glucose tolerance is decreased, indicating that the model of the glycebrate disease model is successful.

Compared with the blank control group, Dl, D2, D3, D4, D5, D6, D7 and SIS3 can significantly reduce blood glucose levels in diabetic animal models. From the therapeutic effect, Dl, D2 and D3 are better than SIS3, D5 and D7 are similar to SIS3, and D4 and D6 are not as effective as SIS3.

Based on the data obtained in Test Example 1, the present invention selected Dl, D2, D3, D4, D5, D6, D7 and SIS3 for pharmacokinetic studies.

Test Example 2 Pharmacokinetic test

Test compound SIS3, compounds D1, D2, D3, D4, D5, D6, D7.

2. Drug preparation

It was prepared with 1% sodium carboxymethylcellulose and physiological saline.

3. Animal grouping and administration methods

SD rats were randomly divided into 8 groups according to the mode of administration, 5 rats in each group: SIS3, Dl, D2, D3, D4, D5, D6, D7 were administered by intragastric administration at a dose of 50 mg/kg. Rats in each group were fasted 12 hours before the experiment. After oral administration of the drug solution, blood was collected at 0.25, 0.5, 1, 1.5, 2, 3, 5, 6, 8, 10, 12, 24, 48 h, and placed in a heparinized plastic centrifuge tube.

4. Treatment of rat plasma samples

The collected blood was centrifuged at 4 °C (6000 r/min, lOmin), 100 μl of plasma was taken, 500 μl of methanol was added, vortexed for 1 min, centrifuged C13000r/min, lOmin), and the supernatant was transferred to EP. In the tube, nitrogen was blown dry, dissolved in a mobile phase, centrifuged at C13000 r/min, 2 min), and the supernatant was taken for 70 μl for HPLC analysis. Using the DAS 2.1.1 version of pharmacokinetic intelligence analysis software, the pharmacokinetic compartment model was automatically fitted and the pharmacokinetic parameters were calculated. The results are shown in Table 3.

5. Results and discussion

Table 3 Pharmacokinetic results

Drug pharmacokinetic parameters

Cmax (mg/L) Tmax (h) AUC(0-∞)(mg L*h) Ti/ ₂ (h) Control SIS3 2.09 6 13.36 4.84

Compound D1 1.61 7.33 17.96* 10.94* Compound D2 1.65 8.67 74.56* 25.45* Compound D3 1.35 6.67 43.04* 16.84* Compound D4 0.85* 5.86 7.61* 3.24

Compound D5 1.50 9.52 12.95 6.30

Compound D6 0.96* 5.34 10.20* 4.06

Compound D7 1.38 8.45 14.86 5.23

Note: SIS3: Control compound (CAS: 521985-36-4); *- indicates significant difference compared with SIS3 group (Ρ<0.05) As seen from the above table, when oral dose is 50mg/kg The pharmacokinetic parameters of Dl, D2 and D3 were significantly better than that of compound SIS3. AUC(0-∞) was 1.3 times, 5.6 times and 3.2 times of the original drug, respectively. The half-life was 2.3 times and 5.4 times respectively. 3.5 times. The absorption of D5 and D7 in vivo is comparable to that of SIS3, while the absorption of D4 and D6 in vivo is not as good as that of SIS3. It can be seen that Dl, D2 and D3 are more easily absorbed than the compound SIS3, and the bioavailability of the drug is significantly improved at the same dose, which is more conducive to the exertion of the drug, and the half-life is prolonged, which can reduce the number of administrations and reduce the side effects.

Based on the data obtained in Test Example 1 and Test Example 2, the present invention selected Dl, D2 and D3 for pharmacodynamic studies.

Test Example 3 Anti-diabetes, hypolipidemic, cholesterol-lowering, weight loss and treatment of fatty liver

Test compound

SIS3, compound Dl, compound D2, compound D3

2. Experimental method: Five-week-old C57BL/6J mice, male, were fed with common vocabulary and high-fat words for 10 weeks. Male mice with successful modeling were randomly divided into groups of 5, and body weight was recorded. The administration group was given 10 mg/kg of different compounds, and the blank control group and the normal group were given the same volume of physiological saline. The drug was administered once a day by intraperitoneal injection for 8 weeks. Changes in body weight, food intake, and body temperature were recorded for different groups of mice. After 8 weeks of drug treatment, the biochemical parameters of each group of animals were examined. The results are shown in Table 4, Table 5 and Table 6. After the animals were sacrificed, liver HE staining, see Figure 1.

The therapeutic effect of the compounds in Table 4 on the diabetes model

Table 5 Effect of Compound Treatment on Glucose Tolerance (mg/dL)

Note: NS: saline group; Normal: normal group; SIS3: control compound (CAS: 521985-36-4); *- indicates significant difference compared with untreated group (P<0.05) Table 6 Effect of Compound Treatment on Insulin Tolerance Test (mg/dL)

Note: NS: saline group; Normal: normal group; SIS3: control compound (CAS: 521985-36-4); *· indicates significant difference compared with untreated group (P<0.05)

The above results showed that the compounds D1, D2 and D3 had a good therapeutic effect on diabetes without affecting the food intake. When the intraperitoneal dose was 10 mg/kg, the body weight of C57BL/6J obese mice was significantly reduced. Fasting blood glucose, transaminase and fasting serum insulin improve insulin sensitivity, improve blood lipid index and oral glucose tolerance, and have obvious therapeutic effects on diabetic fatty liver.

Test Example 4 Acute toxicity test

The present invention investigates the acute toxicity of the compounds D1, D2 and D3. The compounds D1, D2 and D3 of Examples 1, 2, and 3 were dispersed in 1% sodium carboxymethylcellulose, and ground to a suspension to prepare 300 mg/ml. Kunming mice were given lg/kg by oral gavage alone. During the 14-day observation period, mice did not die. On the 15th day of the experiment, blood biochemical tests were performed, and no obvious abnormal changes were found in blood biochemical indicators. In addition, no abnormal drug-related changes were observed in the main organs of the pathological anatomy of the mice, and the animals in the administration group did not exhibit abnormalities compared with the vehicle control group and the normal group.

Test Example 5 Solubility test

The following are examples of Dl and D2 compounds and their salts to illustrate the effect of salt formation on solubility (water solubility). The test results are shown in Table 7:

Table 7 Compound Solubility

Compound solubility (μ Ι)

Compound D1 1.439

D1 hydrochloride 37.221

D1 mesylate salt 18.921

D1 p-toluenesulfonate 10.794

D1 citrate 7.269

D1 benzoate 3.989

Compound D2 2.243

D2 hydrochloride 8.807

D2 mesylate 4.989

D2 p-toluenesulfonate 9.428

D2 citrate 2.391

D2 benzoate 3.131 As can be seen from Table 7, after salt formation, the solubility of the compound increases.

Test Example 6 Pharmacokinetic Study of Salt-Forming Compounds

From the above data, the oral absolute bioavailability of the drug D1 hydrochloride, D1 mesylate, and D1 p-toluenesulfonate was 18.5% 60.7% and 23.6%, respectively. The oral absolute bioavailability of the drug D2 hydrochloride, D2 mesylate, and D2 p-toluenesulfonate was 13.3%, 22.9%, and 37.4%, respectively.

Test Example 7 Therapeutic effect of D1 mesylate on diabetes-related diseases

The efficacy of D1 mesylate was evaluated using a spontaneous type 2 diabetes model db/db mouse. SPF male db/db mice and db/m mice were provided by Changzhou Cavans Experimental Animal Co., Ltd. Eight-week-old db/db mice were randomly divided into three groups for drug treatment trials, namely 10 in the model vehicle group, 10 in the metformin positive control group, and 10 in the drug treatment group. Ten male healthy db/m mice of the same age as the model group were selected as the normal control group. Model vehicle group: 15% propylene glycol + 35% PEG400 + normal saline, 1 time per day, volume 5ml 'kg times - ¹ ; metformin group: 1 time per day, the dose is 300mg ' kg ^ days - ¹ ; Drug treatment group: D1 mesylate was administered once a day at a dose of 50 mg_kg days- ¹ ; in the normal control group, the same volume of normal saline was administered by gavage. The drug was administered for 8 weeks, and the body weight and food intake of the different groups were recorded weekly. At the end of the experiment, fasting blood glucose and glucose tolerance were measured. After the animals were sacrificed, the liver and adipose tissue were stained with HE, and the kidneys were stained with PAS and Masson. The experimental results are as follows: As can be seen from Fig. 2, the D1 mesylate group did not affect the food intake of the mice compared with the solvent group.

As can be seen from Figure 3, D1 mesylate has a good weight-reducing effect.

The results in Figure 4 show that the drug-treated group can significantly reduce fasting blood glucose in mice (P<0.05).

The results in Figure 5 show that compared with the solvent group and the metformin group, administration of D1 mesylate at 50 mg/kg can significantly improve glucose tolerance in db/db mice.

The results in Figure 6 show that under normal light microscope, the liver tissue of the normal group is clear, the liver cells are normal and evenly distributed. In the solvent group and the metformin group, the liver tissue structure is destroyed, the liver cells are swollen, and the liver tissue contains a large amount. Fat Cells, fatty vacuoles dense, honeycomb, form severe fatty liver; 50mg/kg gavage administration of D1 mesylate treatment group is similar to the normal group, mouse liver tissue structure and cell morphology is normal, liver cell contour is clear, No significant fat vacuoles were observed in the liver tissue. Thus, D1 mesylate can significantly treat fatty liver in db/db mice.

The results in Figure 7 show that under the light microscope, the volume of fat cells in the solvent group was significantly larger than that in non-diabetic db / m mice, while the fat cells in the 1) mesylate treatment group were significantly reduced.

Figure 8 shows the results of PAS staining of kidney tissue sections. It can be seen from the figure that the renal pathology of the mice in the solvent group is thickened by the glomerular capillary basement membrane, and the mesangial area is broadened, including the increase of mesangial cells and mesangial matrix and membrane. The red glycogen deposition inside; after the drug treatment, the above pathological changes were alleviated to some extent compared with the solvent group.

Figure 9 shows the results of Masson staining of kidney tissue sections. It can be seen from the figure that there is a large amount of collagen fiber deposition (blue) in the solvent group, collagen deposition in the metformin group is similar to that in the solvent group, and a large amount of muscle fibers (red) in the drug-treated group, and normal. The similarity of the group indicates that D1 mesylate can reduce renal collagen deposition, which may delay the progression of glomerular sclerosis and tubular fibrosis in diabetic nephropathy.

The above results indicate that when D1 mesylate is administered orally at a dose of 50 mg/kg, D1 mesylate can significantly reduce blood glucose and improve oral glucose tolerance in db/db mice after 8 weeks of treatment, and /db mouse fatty liver has obvious therapeutic effect, effectively reducing the deposition of glycogen and collagen in the kidney, which may delay the progression of glomerular sclerosis and tubular fibrosis in diabetic nephropathy.

Test Example 8 Therapeutic effect of compounds on pulmonary fibrosis

A healthy female Wistar rat was instilled with bleomycin (BLM) by endotracheal intubation at a concentration of 5 mg/kg to prepare a pulmonary fibrosis model. Rats were then randomly divided into control group (15% propylene glycol + 35% PEG400 + normal saline;), D2 p-toluenesulfonate treatment group, and rats not injected with BLM were normal control group, 6 rats in each group. On the second day of modeling, the rats were intragastrically administered daily, the control group was given a corresponding volume of the vehicle, and the normal control group was given a corresponding volume of physiological saline. After 28 days of treatment, the rats were anesthetized, blood was taken through the abdominal aorta, and the supernatant was taken for use. The lung weight was weighed and the lung coefficient was calculated. For lung lavage, lung lavage fluid (BALF) was taken, and the cells and supernatant in BALF were separated for testing. The left lung was fixed with 4% paraformaldehyde to prepare paraffin sections for HE and Masson staining, and the right lung tissue was used to detect the content of hydroxyproline in lung tissue. All data are represented by X Shi SD and tested by SPSS 13.0 statistical software. * indicates P < 0.05. ** indicates P < 0.01. *** indicates P < 0.001.

Experimental results:

(1) Evaluation of lung coefficient

Table 9 lung coefficient of each group

Animal number rat weight (g) lung coefficient mean lung coefficient

1 415 1970 4.75

2 399 2150 5.39

3 400 2000 5.00

Normal group 5.29

4 405 2010 4.96

5 410 2120 5.17

6 308 1995 6.48

1 438 2430 5.55

2 436 2020 4.63

Sham operation group 3 425 2300 5.41 5.26

4 430 2315 5.38

5 428 2200 5.14 6 435 2380 5.47

1 195 3100 15.90

2 188 2880 15.32

3 200 1920 9.6

BLM+ solvent group 14.75

4 192 3000 15.63

5 190 2995 15.76

6 189 3080 16.30

1 375 3060 8.16

2 384 2770 7.21

BLM + (D2 toluene 3 387 2420 6.25

7.68 sulfonate group) 4 376 3290 8.75

5 382 3125 8.18

6 385 2900 7.53

It can be seen from Table 9 that the lung coefficient of the BLM group is significantly increased, indicating that the collagen deposition in the lung is more serious and the degree of pulmonary fibrosis is higher. After the intervention of D2 on the tosylate, the lung coefficient of the rat is compared with the BLM group. Significantly decreased, demonstrating that D2 p-toluenesulfonate has a significant effect in reducing collagen deposition.

(2) HE staining

As can be seen from the HE results (Fig. 10), on the 29th day, the lung tissue structure of the solvent group was destroyed, the alveolar space was significantly reduced, and the pulmonary interstitial was replaced by collagen fibers and fibroblasts to form diffuse pulmonary fibrosis. In the normal group and the sham operation group, the lung structure was normal, and the alveolar septum showed no edema, inflammation, and pulmonary fibrosis. After treatment with D2 p-toluenesulfonate, the structure of the lungs was normal, and the alveolar septum was edema, inflammation, and pulmonary fibrosis.

(3) Masson staining

As shown in Figure 11 : Masson staining showed collagen staining in blue, collagen deposition in the bleomycin solvent control group was significantly higher than that in the normal group, and collagen deposition was significantly reduced in the D2 p-toluenesulfonate-treated group compared with the bleomycin group. .

(4) Lung lavage fluid cell detection

As shown in Figure 12: The total number of cells in the BALF and the neutrophils were different and statistically significant (P < 0.01) compared with the bleomycin control group in the D2 p-toluenesulfonate-treated group. D2 p-toluenesulfonate treatment differed from normal values but there was no statistical difference.

In summary, the deuterated compound prepared by the invention can significantly improve body weight, fasting blood glucose, transaminase and fasting serum insulin in obese patients, improve insulin sensitivity, improve blood lipid index and oral glucose tolerance, and can effectively treat fibrosis-related Diseases; These compounds have good absorption in the body, high bioavailability, which is beneficial to the exertion of the drug, and the half-life is prolonged, which can reduce the number of administrations, reduce the side effects, and provide a safe and reliable new choice for clinical use.

Claims

claims

1. The compound represented by formula I or its pharmaceutically soluble solvate,

Formula I

R1, R2, and R3 are each independently selected from H, deuterium, C1 C3 alkyl, partially deuterated or fully deuterated C1 C3 alkyl;

R4 is selected from -CH -,

〇_or none;

R5 and R6 are independently selected from H

Among them, R7~R9 are independently selected from H, deuterium, halogen, and C1~C4 alkyl group.

Fully deuterated C1~C4 alkyl group, R12 is selected from H, C1-C4

R15 is selected from H, C1 C4 alkyl group or alkyloxy group, C- R _16;

R16 is selected from C1~C4 alkyl groups.

2. The compound according to claim 1 or its pharmaceutically acceptable salt, hydrate or solvate, characterized in that: R1, R2, R3 are independently selected from H, deuterium, C1 C3 alkyl, partial deuterium Substituted or fully deuterated C1 C3 alkyl group;

—— c=c—

R4 is selected from ^H H or -C≡C-;

R5, R6 and the N connected to them

, Rio and Rl l are selected from undeuterated, partially or fully deuterated C1 C4 alkyl groups;

Among them, at least one of R1, R2, R3, R10 and R11 is deuterium or a deuterium substituted product.

3. The compound according to claim 1 or 2 or its pharmaceutically acceptable salt, hydrate or solvate, characterized in that: R4 is selected

4. The compound according to claim 1 or 2 or its pharmaceutically acceptable salt, hydrate or solvate, characterized by The alkyl group of C1 to C3 and C1 to C4 is a methyl group or an ethyl group.

5. The compound according to claim 4 or its pharmaceutically acceptable salt, hydrate or solvate, characterized in that: R1, R2, R3 are each independently selected from H or deuterium; R10 and R11 are each independently selected from H or deuterium; Methyl, monodeuterium methyl, dideuterium methyl or trideuterium

6. The compound according to claim 1 or its pharmaceutically acceptable salt, hydrate or solvate, characterized in that: the compound is selected from: r,

or j

012

V. c 、ί ΐ¾ ί

0,

strict'

■

C, CD,

Di 7

° a H

-Ί

CD,.

:ΰ19

7. The compound according to claim 1 or 6 or its pharmaceutically acceptable salt, hydrate or solvate, characterized in that: the compound is selected from one of the following:

8. The compound or a pharmaceutically acceptable salt, hydrate or solvate thereof according to any one of claims 1 to 7, characterized in that: the pharmaceutically acceptable salt is hydrochloric acid of the compound Salt, hydrobromide, phosphate, sulfate, mesylate, p-toluenesulfonate, citrate, benzoate, fumarate, succinate or acetate.

9. The compound according to claim 8 or its pharmaceutically acceptable salt, hydrate or solvate, characterized in that: the pharmaceutically acceptable salt is the mesylate or p-toluene of the compound Sulfonates.

10. The compound according to claim 9 or its pharmaceutically acceptable salt, hydrate or solvate, characterized in that: the pharmaceutically acceptable salt is the methanesulfonate salt of the D1 compound or the D2 compound p-Toluenesulfonate.

11. The compound of claim 10 or its pharmaceutically acceptable salt, hydrate or solvate in the preparation of drugs for improving insulin sensitivity, protecting liver and lowering enzymes, or improving blood sugar, blood lipids or body weight. use.

12. The use according to claim 11, characterized in that: the drug is a drug for treating diabetes, hyperlipidemia, obesity or fatty liver.

13. The use according to claim 12, characterized in that: the fatty liver is diabetic fatty liver or primary fatty liver.

14. The use according to claim 12, characterized in that: the drug is a drug for alleviating or treating diabetic nephropathy.

15. Use of the compound of claim 10 or its pharmaceutically acceptable salt, hydrate or solvate in the preparation of a drug for treating pulmonary fibrosis or liver fibrosis.

16. A pharmaceutical composition, characterized in that: it is composed of the compound described in any one of claims 1 to 10 or its pharmaceutically acceptable salt, hydrate or solvate as an active ingredient, plus commonly used pharmaceutical ingredients Preparations prepared from excipients or auxiliary ingredients.