Application of Proglumide

Abstract

The present invention discloses a new application of proglumide in the preparation of drugs for preventing and treating venous embolism. By inhibiting the activity of P2Y 14 receptors, it can effectively relieve venous thrombosis and has no significant effect on bleeding time and platelet aggregation ability. Furthermore, it can be prepared into drugs for preventing and treating venous embolism caused by factors such as surgical operations, trauma, pregnancy, cancer, and long-term bedridden state, and significantly reduces the bleeding risk caused by traditional anticoagulant drugs. It can be used as a single-agent preparation or in combination with a compound preparation, having multiple effects of prevention and treatment and wide applications.

Description

Technical Field

[0001] The present invention relates to an application of proglumide, and in particular to a new application of proglumide as a drug for preventing or treating venous thrombosis. Background Art

[0002] Proglumide is also known as Proglumide (C 18 H 26 N2O4, CAS No.: 6620-60-6, ), belongs to antacids and drugs for treating peptic ulcers. It inhibits the secretion of gastric acid and pepsin by competitively inhibiting gastrin receptors, protects the gastric mucosa and promotes healing. It can be used to treat gastric ulcers, duodenal ulcers, gastritis and other digestive tract diseases. At the same time, proglumide can non-selectively antagonize cholecystokinin receptors, promote bile secretion, and has a strong choleretic effect. In addition, in non-alcoholic fatty hepatitis, proglumide may improve non-alcoholic fatty hepatitis by interacting with bile acid receptors and changing the characteristics of intestinal microorganisms; proglumide can also enhance the analgesic effect of analgesics such as celecoxib and morphine, and reduce rats' tolerance to morphine.

[0003] Venous thromboembolism (VTE) includes deep venous thrombosis (DVT) and its complication pulmonary embolism (PE). It refers to a disease in which blood abnormally coagulates in the deep veins, blocking blood return and accompanied by inflammatory changes in the vein wall. It is common in the deep veins of the lower extremities. PE occurs when a thrombus breaks off from its initial location and reaches the lungs, which may lead to respiratory insufficiency or even death. Currently, drug treatments for VTE mainly include thrombolytic therapy and anticoagulant therapy, but both have a high risk of bleeding. Summary of the invention

[0004] Purpose of the invention: In view of the problem that existing drugs affect the coagulation system and platelet function, which makes bleeding more likely to occur, the present invention aims to provide a new application of proglumide and overcome the above problems.

[0005] Technical solution: In the present invention, proglumide is used in the preparation of drugs for preventing or treating venous thrombosis, specifically in the preparation of drugs for preventing or treating deep vein thrombosis or pulmonary embolism.

[0006] The specific application population can be patients who already have related diseases or people with potential risks of related diseases.

[0007] Among them, the above-mentioned venous thrombosis is venous thrombosis caused by trauma, surgery, pregnancy, postpartum period, long-term immobilization, tumor, oral contraceptives, intravenous catheterization, nephrotic syndrome, Crohn's disease, polycythemia, macroglobulinemia or myelodysplastic syndrome.

[0008] The above-mentioned proglumide drugs can inhibit deep vein thrombosis or embolus shedding in new applications, especially without affecting the coagulation system or platelet function, because it inhibits P2Y 14 Receptors play a corresponding role.

[0009] Studies have shown that immune dysregulation plays a key role in the process of thrombosis. 14 The receptor is a Gi protein-coupled receptor that inhibits cyclic adenosine monophosphate and belongs to the P2Y purinergic receptor family. It can be activated by uridine-5'-diphosphoglucose (UDPG) and other endogenous UDP-sugars. 14 The receptors are widely distributed in tissues and expressed in smooth muscle cell subpopulations of the pancreas, gastrointestinal tract, blood vessels, lungs and uterus. They participate in the body's immune response and allergic diseases and are closely related to the occurrence and development of diseases such as acute gouty arthritis, asthma, liver fibrosis, and renal ischemia-reperfusion injury. This invention first discovered that proglumide inhibits P2Y 14 Receptors play a role in alleviating venous thrombosis.

[0010] The present invention constructs a rat deep vein thrombosis model and finds that P2Y 14 Receptor deletion can effectively reduce venous thrombosis. 14 Different concentrations of the test substance were added to the HEK293 cell line with the receptor in advance, and PPTN was used as the positive control to detect the intracellular cAMP content. The IC 50 and the relative inhibition rate of cAMP. The results showed that proglumide had an inhibitory effect on P2Y 14 In summary, the present invention has proved through in vitro and in vivo experiments that proglumide can inhibit P2Y 14 Receptors relieve venous thrombosis.

[0011] In the application of the present invention, compared with aspirin, proglumide has no significant effect on the bleeding time and platelet aggregation function of rat tails. Compared with existing antithrombotic drugs, proglumide does not affect the coagulation system and platelet function, and has clinical application value for preparing new antithrombotic drugs.

[0012] In the above new application, proglumide can also be used in combination with other drugs for preventing or treating venous embolism, specifically, a single preparation of proglumide can be used in combination with other drugs for preventing or treating venous embolism, or a compound preparation of proglumide and other active ingredients with the effect of preventing or treating venous embolism can be used in combination. In the above drug combination, the inhibition of deep vein thrombosis or embolus shedding can still be achieved without affecting the coagulation system or platelet function.

[0013] The chemical form of proglumide used specifically can be its free base or free acid, or its pharmaceutically acceptable salt, more specifically, a salt formed by proglumide and hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, methanesulfonic acid, benzenesulfonic acid, p-toluenesulfonic acid, naphthalenesulfonic acid, citric acid, malic acid, tartaric acid, lactic acid, pyruvic acid, acetic acid, maleic acid, fumaric acid, succinic acid, salicylic acid, phenylacetic acid, mandelic acid, acidic amino acids; or a salt formed by an inorganic base containing a basic metal cation, an alkaline earth metal cation or an ammonium cation salt, or an organic base such as triethylamine, trimethylamine, 4-DMAP, pyridine, morpholine, piperazine, N-methylpiperazine, or a basic amino acid; it can be a single salt among the above salts, or a mixture of the above salts. The physical form can be a crystalline state, an amorphous state, or a eutectic state; it can be a single form among the above forms, or a mixture of the above forms.

[0014] The specific dosage forms of the drugs used are tablets, capsules, granules, powders, powder injections, pills, emulsions, suspensions, solutions, sprays, patches, sublingual tablets, etc.

[0015] Beneficial effects: Compared with the prior art, the present invention has the following significant advantages:

[0016] 1. Ability to inhibit P2Y 14 It inhibits the activity of IL-27 receptors and slows down the formation of venous thrombosis. Its activity is not significantly different from that of the positive drug aspirin, and it does not affect the coagulation system and platelet aggregation function, significantly reducing the risk of bleeding.

[0017] 2. It can be used as a single preparation or in combination with a compound preparation. It has multiple preventive and therapeutic effects and is widely used. BRIEF DESCRIPTION OF THE DRAWINGS

[0018] Figure 1 For wild type and P2Y 14 Wet weight of IVC thrombus in receptor knockout rats;

[0019] Figure 2 is the inhibition rate of cAMP generation by different concentrations of proglumide;

[0020] Figure 3 is the wet weight of the inferior vena cava thrombus in rats after administration of proglumide;

[0021] Figure 4 This is the change of tail bleeding time in rats after administration of proglumide;

[0022] Figure 5 It is the maximum aggregation rate of rat platelets after incubation of platelets with proglumide. DETAILED DESCRIPTION

[0023] The technical solution of the present invention is further described below in conjunction with embodiments.

[0024] Example 1: P2Y 14 Effects of receptor deficiency on deep vein thrombosis model induced by inferior vena cava stenosis in rats

[0025] Male SPF wild type (WT) and P2Y 14 Receptor knockout (P2Y 14 R - / - )SD rats, weighing 200±20g, free access to water and food, 12h lighting per day, and an ambient temperature of 25±2℃. Rats were fasted for 12h before the experiment, anesthetized by intraperitoneal injection, and subjected to median laparotomy to expose the inferior vena cava. A 29G needle was placed along the inferior vena cava, followed by ligation with 5-0 non-absorbable sutures at the branch of the left renal vein. The needle was removed, causing a 90% reduction in blood flow in the inferior vena cava of the rats. The inferior vena cava thrombus was removed 48h after surgery, and the wet weight of the thrombus in the rats was measured.

[0026] The results are as follows Figure 1 As shown, compared with WT rats, P2Y 14 R - / - The wet weight of the inferior vena cava thrombus in rats was significantly reduced, indicating that P2Y 14 R plays an important role in the pathogenesis of venous thrombosis. * P<0.05, n=10.

[0027] Example 2: Compounds as P2Y 14 Evaluation of inhibitory activity of receptor inhibitors

[0028] Stable P2Y 14 HEK293 cells of the recipient (purchased from Jiangsu Keygen Biotechnology Co., Ltd.) were cultured in DMEM medium (containing 10% fetal bovine serum, 100 U / ml penicillin and 100 μg / ml streptomycin) and inoculated into 384-well culture plates one day before the experiment at a seeding density of 1×10 4cells / well, and the cells were cultured at 37°C, 95% O2, and 5% CO2. Before the experiment, the culture medium was discarded and replaced with serum-free culture medium. IBMX (500μM) and Ro 20-1724 (100μM) were added to inhibit PDEs activity to ensure that cAMP was at a higher level. The AC agonist Forskolin (30μM) was used to stimulate the production of cellular cAMP, and different concentrations of proglumide (0.01, 0.1, 1, 10, 100nM) were added in advance. PPTN (CAS No.: 1160271-30-6) was used as a positive control. At the same time, 10μM P2Y 14 The receptor agonist UDPG was used to detect the intracellular cAMP content according to the instructions of the cAMP GloTM Assay Kit (PROMEGA Co. Ltd, USA) after 30 minutes. The IC 50 The results are shown in Figure 2 and Table 1.

[0029] Table 1. Effects of Proglumide on P2Y 14 Inhibitory activity of receptors

[0030] Inhibition rate (%) Relative inhibition rate (%, PPTN 100.84%) <![CDATA[IC 50 (nM)]]> 102.37 101.52 67.11

[0031] The results showed that the relative inhibition rate of proglumide on cAMP (100 nM) was 101.52%; the half inhibitory concentration IC 50 It is 67.11nM.

[0032] Example 3: Pharmacological study of proglumide in a rat deep vein thrombosis model induced by inferior vena cava stenosis

[0033] Male SPF SD rats, weighing 200±20g, were given free water and food, and were illuminated for 12h per day at an ambient temperature of 25±2°C. The animals were randomly divided into three groups: a model group, an aspirin positive control group (10mg / kg), and a proglumide group (10mg / kg), with 8 rats in each group. The animal model construction method was the same as in Example 1, and aspirin and proglumide were administered by gavage 1h before surgery and 24h after surgery, respectively. The inferior vena cava thrombus was removed 48h after surgery, and the wet weight of the thrombus in the rats was measured.

[0034] The results are as follows Figure 3 As shown in Figure 2, the wet weight of thrombus in rats treated with proglumide was significantly reduced compared with the solvent control group, but there was no significant difference with the aspirin treatment group ( ** P<0.01, n=8).

[0035] Example 4: Pharmacological evaluation of the effect of proglumide on physiological hemostasis in rats

[0036] Male SPF SD rats, weighing 200±20g, were given free water and food, and were illuminated for 12h per day at an ambient temperature of 25±2℃. The animals were randomly divided into three groups: solvent control group, aspirin positive control group (10mg / kg) and proglumide group (10mg / kg), with 5 rats in each group. The rats were fasted for 12h before the experiment, and aspirin and proglumide were administered by gavage 1h in advance. The rat tails were cut off at 5mm from the tip of the tail, and the wound was placed in 37℃ saline. The time from tail cutting to bleeding cessation was recorded, which was the bleeding time.

[0037] The results are as follows Figure 4 As shown in Figure 2, compared with the solvent control group, the administration of proglumide did not prolong the tail bleeding time of rats, while the tail bleeding time of rats was significantly increased after aspirin administration ( *** P<0.001, ns=non-significance, n=5).

[0038] Example 5: ADP-induced platelet aggregation experiment

[0039] Male SD rats were fasted for 12 h before the experiment. After intraperitoneal injection of anesthesia, the laparotomy was performed. Blood was collected from the abdominal aorta using a sodium citrate anticoagulant tube. After mixing, the upper platelet-rich plasma was collected and placed in another test tube, which was then centrifuged at 600 g for 6 min to obtain platelets. Platelets were resuspended in platelet-poor plasma and the platelet count was adjusted to 3 × 10 8 / mL. Platelets need to be left at 37°C for 30 minutes to restore activity, and platelet aggregation is detected by optical turbidimetry. Before the measurement, the instrument transmittance is adjusted to zero with platelet-poor plasma, 300 μL of platelets and 3 μL of blank solvent or drug (working concentration 1 mg / mL) are added to the cuvette, and after gentle pipetting and mixing, they are pre-incubated at 37°C for 5 minutes. The cuvette to be tested is then placed in the detection channel after zeroing, and the inducer ADP (working concentration 10 μM) is added to immediately test platelet aggregation, and the maximum platelet aggregation rate within 5 minutes is recorded.

[0040] The results are as follows Figure 5 As shown in the figure, aspirin exerts its antiplatelet effect by inhibiting cyclooxygenase activity and reducing thromboxane A2 synthesis. As can be seen from the figure, aspirin pretreatment significantly inhibited ADP-induced platelet aggregation in rats, while proglumide had no significant effect on the platelet aggregation function of rats ( ** P<0.01, ns=non-significance, n=4).

Claims

1. Use of proglumide in the preparation of a drug for preventing or treating venous thrombosis, wherein the venous thrombosis is deep vein thrombosis or pulmonary embolism.

2. The use according to claim 1, characterized in that: The venous thrombosis is caused by trauma, surgery, pregnancy, puerperium, long-term immobilization, tumor, oral contraceptives, intravenous catheterization, nephrotic syndrome, Crohn's disease, polycythemia, macroglobulinemia or myelodysplastic syndrome.

3. The use according to claim 1, characterized in that: The drug inhibits the formation of deep vein thrombosis or the dislodgement of emboli.

4. The use according to claim 3, characterized in that: The drug inhibits the formation of deep vein thrombosis or the shedding of emboli without affecting the coagulation system or platelet function.

5. The use according to claim 1, characterized in that: The drug inhibits P2Y 14 Receptor.

6. The use according to claim 1, characterized in that: Proglumide is used in combination with other drugs to prevent or treat venous thrombosis.

7. The use according to claim 6, characterized in that: Medicines containing proglumide as the sole active ingredient are used in combination with other medicines for the prevention or treatment of venous thrombosis.

8. The use according to claim 6, characterized in that: The compound medicine composition is formed by proglumide and other active ingredients having the function of preventing or treating venous thrombosis.

9. The use according to claim 6, characterized in that: The combination of the drugs inhibits the formation of deep vein thrombosis or the shedding of emboli without affecting the coagulation system or platelet function.

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