Application of micromolecular diterpenoid compound in preparation of medicine for preventing and treating acute pancreatitis

Abstract

The invention relates to application of a small-molecule diterpene compound in preparation of a medicine for preventing and treating acute pancreatitis. The small-molecule diterpene compound is selected from a compound with a structure as shown in a formula (I) or pharmaceutically acceptable salt, ester and solvate of the compound. The research finds that the traditional Chinese medicine composition has an excellent effect of preventing and treating acute pancreatitis, can inhibit expression of inflammatory factors of human peripheral blood mononuclear cells induced by lipopolysaccharide, and has a good anti-inflammatory effect; meanwhile, the composition can improve pancreatic tissue pathological changes of mice with acute pancreatitis, alleviate edema injury, alleviate inflammation reaction and inhibit expression of lipase in serum of the mice. The invention provides a theoretical basis for researching a treatment strategy of acute pancreatitis, and provides an embedding point for preparing a new medicine for preventing or treating acute pancreatitis.

Description

Technical Field

[0001] The present invention belongs to the field of biomedicine technology and relates to the use of a small molecule diterpene compound in the preparation of a drug for preventing and treating acute pancreatitis, and specifically relates to the use of a compound having a structure represented by formula (I) or a pharmaceutically acceptable salt, ester or solvate thereof in the preparation of a drug for preventing and treating acute pancreatitis. Background Art

[0002] Acute pancreatitis (ALI) refers to an acute abdomen caused by abnormal activation of pancreatic enzymes that produce digestive effects on the pancreas and surrounding organs, characterized by local inflammatory reactions in the pancreas, and may even lead to organ dysfunction. Acute pancreatitis may be complicated by one or more organ dysfunctions, with respiratory and renal function impairment being more common. Globally, acute pancreatitis is a common digestive system emergency that requires hospitalization, and its incidence varies in certain regions. In recent years, the incidence of acute pancreatitis has been on the rise, and clinical attention should be paid.

[0003] Current studies have revealed that the pathogenesis of acute pancreatitis involves premature activation of trypsinogen, pathological calcium overload, pancreatic microcirculatory disorders, NF-kB activation, leukocyte infiltration, and impaired autophagy. Although timely clinical management, such as fasting, nutritional support, and fluid therapy, is effective in improving the treatment of patients with acute pancreatitis, more in-depth mechanistic studies are needed to explore more potential targets for acute pancreatitis.

[0004] Early treatment of acute pancreatitis mainly includes fluid therapy, analgesia and nutritional support, and treatment for the cause and early complications. Regarding the drug treatment of acute pancreatitis, there is still a lack of high-quality clinical evidence on the therapeutic value of protease inhibitors and pancreatic enzyme inhibitors, such as somatostatin and its analogs in acute pancreatitis; Chinese medicine (rhubarb, mirabilite and compound preparations, such as Qingyi Decoction, Dachengqi Decoction, etc.) can help promote the recovery of gastrointestinal function in patients and relieve abdominal pain and bloating symptoms, and can be used as an option.

[0005] The late treatment of acute pancreatitis is mainly aimed at its various local complications. At present, there is still a lack of specific drugs for pancreatitis. The strategies for treating acute pancreatitis in existing technologies are still very limited. It is very meaningful to develop more treatment strategies for acute pancreatitis. Summary of the invention

[0006] In view of the deficiencies in the prior art, the purpose of the present invention is to provide an application of a small molecule diterpene compound in the preparation of a drug for preventing and treating acute pancreatitis, specifically to provide an application of a compound having a structure shown in formula (I) or a pharmaceutically acceptable salt, ester, or solvate thereof in the preparation of a drug for preventing and treating acute pancreatitis.

[0007] In order to achieve the purpose of the invention, the present invention adopts the following technical solutions:

[0008] In a first aspect, the present invention provides an application of a small molecule diterpene compound in the preparation of a drug for preventing and treating acute pancreatitis, wherein the small molecule diterpene compound is selected from a compound having a structure represented by formula (I) or a pharmaceutically acceptable salt, ester, or solvate thereof.

[0009]

[0010] The molecular structure and synthesis method of the compound represented by formula (I) involved in the present invention have been recorded in the text disclosed in patent WO2016150207A1, and will not be described in detail here.

[0011] The present invention creatively finds that the compound of the structure shown in formula (I) or its pharmaceutically acceptable salt, ester, solvate has excellent effect of preventing and treating acute pancreatitis. It can inhibit the expression of tumor necrosis factor α (TNF-α), interleukin 1β (IL-β), interleukin 6 (IL-6) and interleukin 18 (IL-18) in human peripheral blood mononuclear cells (hPBMC) induced by lipopolysaccharide (LPS), and has good anti-inflammatory effect; at the same time, it shows good preventive and therapeutic effects in the acute pancreatitis mouse model induced by caerulein, can improve the pathological changes of pancreatic tissue in acute pancreatitis mice, reduce edema damage, reduce inflammatory response, and inhibit the expression of lipase in mouse serum. The present invention provides a theoretical basis for studying the treatment strategy of acute pancreatitis and provides an insertion point for preparing new drugs for preventing or treating acute pancreatitis.

[0012] Preferably, the pharmaceutically acceptable salt includes any one of tartrate, stearate, oxalate, citrate, lactate, sorbate, fumarate, formate, acetate, benzoate, benzenesulfonate, ethanesulfonate, resinate, trifluoroacetate, maleate, methanesulfonate, fumarate, amino acid salt, nicotinate, phosphate, sulfate, hydrochloride or hydrobromide.

[0013] In the present invention, the drug inhibits the expression of inflammatory factors TNF-α, IL-1β, IL-6 and IL-18.

[0014] In the present invention, the drug inhibits the expression of lipase in serum.

[0015] Preferably, the medicament alleviates pancreatic tissue edema and pancreatic tissue damage.

[0016] In the present invention, the dosage form of the drug is any pharmaceutically acceptable dosage form.

[0017] Preferably, the dosage form is selected from any one of suspension, granules, capsules, powders, tablets, emulsions, solutions, pills, injections, suppositories, enemas, aerosols, sprays, patches or drops.

[0018] The drugs in the above-mentioned various dosage forms can be prepared according to conventional methods in the pharmaceutical field.

[0019] Preferably, the medicament further comprises a pharmaceutically acceptable excipient.

[0020] Preferably, the auxiliary materials include any one or a combination of at least two of carriers, diluents, excipients, binders, wetting agents, disintegrants, emulsifiers, cosolvents, solubilizers, osmotic pressure regulators, surfactants, coating materials, colorants, pH regulators, antioxidants, antibacterial agents or buffers.

[0021] In a second aspect, the present invention provides an application of a small molecule diterpene compound in the preparation of an inflammatory factor inhibitor, wherein the small molecule diterpene compound is selected from a compound having a structure represented by formula (I) or a pharmaceutically acceptable salt, ester, or solvate thereof.

[0022]

[0023] Preferably, the inflammatory factor includes any one of TNF-α, IL-1β, IL-6 or IL-18, or a combination of at least two thereof.

[0024] In a third aspect, the present invention provides an application of a small molecule diterpene compound in the preparation of a lipase inhibitor, wherein the small molecule diterpene compound is selected from a compound having a structure represented by formula (I) or a pharmaceutically acceptable salt, ester, or solvate thereof.

[0025]

[0026] In a fourth aspect, the present invention provides a method for inhibiting the expression of inflammatory factors in cells for non-therapeutic purposes, the method comprising: mixing and incubating cells with an effective dose of a small molecule diterpene compound, wherein the small molecule diterpene compound is selected from a compound of the structure shown in formula (I) or a pharmaceutically acceptable salt, ester, or solvate thereof.

[0027]

[0028] The method claimed for protection by the present invention is not directly implemented on living human or animal bodies, but on simple in vitro cells. It only protects the way of inhibiting the expression of inflammatory factors in cells (cellular level); at the same time, this method is not a process of eliminating the cause or lesion, nor is it a treatment method directly used to improve the health of human or animal bodies, but is for theoretical research on inflammatory pathways.

[0029] Compared with the prior art, the present invention has the following beneficial effects:

[0030] The present invention creatively finds that the compound of the structure shown in formula (I) or its pharmaceutically acceptable salt, ester, solvate has excellent effect of preventing and treating acute pancreatitis. It can inhibit the expression of tumor necrosis factor α (TNF-α), interleukin 1β (IL-β), interleukin 6 (IL-6) and interleukin 18 (IL-18) in human peripheral blood mononuclear cells (hPBMC) induced by lipopolysaccharide (LPS), and has good anti-inflammatory effect; at the same time, it shows good preventive and therapeutic effects in the acute pancreatitis mouse model induced by caerulein, can improve the pathological changes of pancreatic tissue in acute pancreatitis mice, reduce edema damage, reduce inflammatory response, and inhibit the expression of lipase in mouse serum. The present invention provides a theoretical basis for studying the treatment strategy of acute pancreatitis and provides an insertion point for preparing new drugs for preventing or treating acute pancreatitis. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] Figure 1 This is a statistical graph showing the effects of different concentrations of compound A on the expression of inflammatory factor TNF-α in human peripheral blood mononuclear cells stimulated by LPS;

[0032] Figure 2 This is a statistical graph showing the effects of different concentrations of compound A on the expression of inflammatory factor IL-1β in human peripheral blood mononuclear cells stimulated by LPS;

[0033] Figure 3 This is a statistical diagram showing the effect of different concentrations of compound A on the expression of inflammatory factor IL-6 in human peripheral blood mononuclear cells stimulated by LPS;

[0034] Figure 4 This is a statistical graph showing the effects of different concentrations of compound A on the expression of inflammatory factor IL-18 in human peripheral blood mononuclear cells stimulated by LPS;

[0035] Figure 5 This is a statistical chart showing the effect of compound A on the expression of lipase in each group of mice after the mice with acute pancreatitis model were treated;

[0036] Figure 6 This is a typical HE pathological picture of each group of mice after compound A treated acute pancreatitis model mice;

[0037] Figure 7 This is a statistical chart of the total pathological scores of each group of mice after Compound A treated acute pancreatitis model mice. DETAILED DESCRIPTION

[0038] The technical solution of the present invention is further described below by specific implementation methods. It should be understood by those skilled in the art that the embodiments are only to help understand the present invention and should not be regarded as specific limitations of the present invention.

[0039] The process, conditions, reagents, test methods, etc. for implementing the present invention, except for the contents specifically mentioned below, are all common knowledge and common common sense in the art, and the present invention has no particular restrictions. The test methods without specifying specific conditions in the embodiments are usually carried out under conventional conditions or under conditions recommended by the manufacturer.

[0040] Unless otherwise specified, the meanings of all professional terms and scientific terms used in this specification are the same as those generally understood by technicians in the technical field to which the invention belongs. However, in case of conflict, the present specification including the definitions shall prevail.

[0041] The compound A involved in the following examples is provided by Suzhou Huyun New Drug Research and Development Co., Ltd. (which is prepared by the preparation method disclosed in WO2016150207A1), and its molecular structure is shown below:

[0042]

[0043] Example 1

[0044] This example explores the inhibitory effect of compound A on inflammatory factors, and the specific operation is as follows:

[0045] Human peripheral blood mononuclear cells (PBMCs) were resuspended in serum-free medium and the cell concentration was adjusted to 1×10 6 / mL, add LPS, 100μL of cell suspension per well was plated in a 96-well round bottom plate, and cultured at 37℃ for 1h. Then, different concentrations of compound A were added to make the final concentrations 125, 250, 500, and 1000nM, respectively, and the final concentration of LPS was 1μg / mL. After 24h of conventional culture at 37℃, the cell culture supernatant was collected by centrifugation, and the expression levels of inflammatory factors (TNF-α, IL-β, IL-6, and IL-18) in the supernatant were detected by flow cytometry. The statistical results are shown in Tables 1 and Figure 1-4 As shown (LPS-free represents the group without LPS stimulation, Ctrl represents the control group, and others represent the treatment groups at various concentrations):

[0046] Table 1

[0047]

[0048]

[0049] Compared with the group without LPS stimulation, # represents p < 0.05, ## represents p < 0.01, and ### represents p < 0.001; compared with the control group without compound A, * represents p < 0.05, ** represents p < 0.01, and *** represents p < 0.001.

[0050] From Table 1 and Figure 1-4 It can be seen that the expression of TNF-α, IL-β, IL-6, and IL-18 in human peripheral blood mononuclear cells is low without LPS stimulation. After 24 hours of LPS stimulation, the expression of TNF-α, IL-β, IL-6, and IL-18 increased significantly, but compound A can inhibit the expression of TNF-α, IL-β, IL-6, and IL-18. The above test results indicate that compound A has a strong anti-inflammatory ability and is expected to inhibit the level of inflammation in pancreatic tissue of patients with acute pancreatitis.

[0051] Example 2

[0052] This example explores the preventive and therapeutic effect of compound A on an acute pancreatitis mouse model, and the specific operation is as follows:

[0053] (1) Establishment of a mouse model of acute pancreatitis induced by caerulein:

[0054] Caerulein is a gastric regulatory molecule similar to cholecystokinin (CCK) in function and composition. It stimulates the over-secretion of pancreatic acinar cells, causing separation barriers between trypsinogen and lysosomal hydrolases in cells. After activation in a cathepsin B-dependent manner, the activity of pancreatic enzymes can further lead to zymogen activation, causing a series of cell damage caused by protease activity. The inflammatory response caused by pancreatic tissue self-digestion recruits inflammatory cells, releases inflammatory factors, and further causes local or even systemic severe inflammatory reactions. The acute pancreatitis mouse model was established by intraperitoneal injection of caerulein (50μg / kg) 7 times at 1h intervals.

[0055] (2) Efficacy test of compound A on acute pancreatitis mouse model:

[0056] SPF BALB / c mice were divided into 5 groups, 5 mice in each group, including blank control group, model group, model + 10 mg / kg compound A group, model + 30 mg / kg compound A group, and model + 5 mg / kg dexamethasone group (DEX) (oral administration).

[0057] Compound A or dexamethasone solution was administered to mice via tail vein once at 2h, 4h, 6h and 24h of the first modeling. Mice were killed 24h after modeling, and serum and pancreatic tissue were obtained. Serum was tested for lipase expression, and pancreatic tissue was subjected to pathological examination (pancreatic tissue was fixed in 4% formaldehyde solution for 48h, dehydrated, transparent, paraffin-embedded and sliced ​​4μm thick, and then stained with hematoxylin-eosin (HE)), and photographed and tissue scored (edema, inflammatory cell infiltration, acinar cell vacuoles and necrosis).

[0058] (3) Evaluation of the efficacy of compound A in acute pancreatitis mouse model:

[0059] (3.1) The statistical results of lipase expression in the serum of each group of mice are as follows Figure 5 As shown in the figure (NC represents the blank control group, Model represents the model group, and DEX represents the dexamethasone treatment group, where, compared with the blank control group, *** represents p < 0.001, compared with the model group, # represents p < 0.05, and ### represents p < 0.001), it can be seen from the figure that compared with the normal animal control group, the expression of mouse lipase increased after caerulein induction, and the 10mg / kg group and 30mg / kg group of compound A can downregulate the expression of lipase compared with the model group. At the same time, the dexamethasone (DEX) treatment group also significantly alleviated the downregulation of mouse lipase levels. In summary, compound A plays a role in preventing and treating acute pancreatitis by reducing the lipase level of acute pancreatitis model mice and reducing the release of lipase caused by pancreatic damage.

[0060] (3.2) Typical pathological images of pancreatic tissue of mice in each group (40×) are shown below Figure 6 The statistical diagram of pancreatic tissue pathological scores of mice in each group is shown in Figure 7 As shown in the figure, compared with the normal animal control group, pulmonary edema, inflammation and local glandular vacuoles appeared in the pancreatic tissue of mice after caerulein induction. Compared with the model group, the 10mg / kg group and 30mg / kg group of compound A can alleviate the edema of the pancreatic tissue of mice in a dose-dependent manner; the DEX-treated group also significantly alleviated the edema of the pancreatic tissue of mice. In summary, it shows that compound A plays a role in preventing and treating acute pancreatitis by reducing pulmonary edema of the pancreatic tissue of mice with acute pancreatitis model and reducing pathological damage to the pancreatic tissue.

[0061] The applicant declares that the present invention illustrates the application of a small molecule diterpene compound of the present invention in the preparation of a drug for preventing and treating acute pancreatitis through the above-mentioned embodiments, but the present invention is not limited to the above-mentioned embodiments, that is, it does not mean that the present invention must rely on the above-mentioned embodiments to be implemented. Those skilled in the art should understand that any improvement of the present invention, equivalent replacement of various raw materials of the product of the present invention, addition of auxiliary components, selection of specific methods, etc., all fall within the protection scope and disclosure scope of the present invention.

[0062] The preferred embodiments of the present invention are described in detail above. However, the present invention is not limited to the specific details in the above embodiments. Within the technical concept of the present invention, a variety of simple modifications can be made to the technical solution of the present invention, and these simple modifications all belong to the protection scope of the present invention.

[0063] It should also be noted that the various specific technical features described in the above specific embodiments can be combined in any suitable manner without contradiction. In order to avoid unnecessary repetition, the present invention will not further describe various possible combinations.

Claims

1. Use of a small molecule diterpene compound in the preparation of a drug for preventing and treating acute pancreatitis, characterized in that: The small molecule diterpene compound is selected from the compound with the structure shown in formula (I) or its pharmaceutically acceptable salt, ester, solvate.

2. The use according to claim 1, wherein the pharmaceutically acceptable salt comprises any one of tartrate, stearate, oxalate, citrate, lactate, sorbate, fumarate, formate, acetate, benzoate, benzenesulfonate, ethanesulfonate, resinate, trifluoroacetate, maleate, methanesulfonate, fumarate, amino acid salt, nicotinate, phosphate, sulfate, hydrochloride or hydrobromide.

3. The use according to claim 1 or 2, characterized in that: The drug inhibits the expression of inflammatory factors TNF-α, IL-1β, IL-6 and IL-18.

4. The use according to any one of claims 1 to 3, characterized in that: The drug inhibits the expression of lipase in serum; Preferably, the medicament alleviates pancreatic tissue edema and pancreatic tissue damage.

5. The use according to any one of claims 1 to 4, characterized in that: The dosage form of the drug is any pharmaceutically acceptable dosage form; Preferably, the dosage form is selected from any one of suspension, granules, capsules, powders, tablets, emulsions, solutions, pills, injections, suppositories, enemas, aerosols, sprays, patches or drops.

6. The use according to any one of claims 1 to 5, characterized in that: The drug also includes pharmaceutically acceptable excipients.

7. The use according to claim 6, characterized in that: The auxiliary materials include any one or a combination of at least two of carriers, diluents, excipients, binders, wetting agents, disintegrants, emulsifiers, cosolvents, solubilizers, osmotic pressure regulators, surfactants, coating materials, colorants, pH regulators, antioxidants, antibacterial agents or buffers.

8. Use of a small molecule diterpene compound in the preparation of an inflammatory factor inhibitor, characterized in that: The small molecule diterpene compound is selected from the compound of the structure shown in formula (I) or a pharmaceutically acceptable salt, ester, or solvate thereof; Preferably, the inflammatory factor includes any one of TNF-α, IL-1β, IL-6 or IL-18, or a combination of at least two thereof.

9. Use of a small molecule diterpene compound in the preparation of a lipase inhibitor, characterized in that: The small molecule diterpene compound is selected from the compound with the structure shown in formula (I) or its pharmaceutically acceptable salt, ester, solvate.

10. A method for inhibiting the expression of inflammatory factors in cells for non-therapeutic purposes, characterized in that: The method comprises: mixing and incubating cells with an effective dose of a small molecule diterpene compound, wherein the small molecule diterpene compound is selected from a compound of the structure shown in formula (I) or a pharmaceutically acceptable salt, ester, or solvate thereof.

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