Ozagrel-RGDF tetramer and preparation and application thereof

Abstract

The invention discloses an ozagrel-RGDF tetramer with the following structural formula, namely [ozagrel-RGDF] 4, and discloses a preparation method of the ozagrel-RGDF tetramer and application of the ozagrel-RGDF tetramer to treatment of arterial thrombotic diseases. Experiments prove that the [ozagrel-RGDF] 4 disclosed by the invention not only has a good anti-arterial thrombosis effect, but also has no bleeding side effect. Therefore, the invention provides a safe and effective technical means for arterial thrombosis resistance.

Description

Technical Field

[0001] This invention discloses an ozagrel-RGDF tetramer, namely [ozagrel-RGDF]4, and its preparation method and its application in the preparation of anti-arterial thrombotic drugs. This invention belongs to the field of biomedicine. Background Technology

[0002] Cardiovascular disease (CVD) is considered a global public health problem due to its high mortality and disability rates. Although the pathogenesis of CVD is diverse, it is now widely accepted that arterial thrombosis is a significant contributing factor.

[0003] Thromboxane A2 (TXA2) is a metabolite of arachidonic acid and can cause vasoconstriction and platelet aggregation. Ozagrel is a selective thromboxane A2 synthase inhibitor that primarily works by inhibiting platelet activation and promoting vasodilation. Although the National Medical Products Administration has approved Ozagrel injection for the treatment of acute thrombotic cerebral infarction and cerebral infarction-associated motor dysfunction, the risk of severe intracranial hemorrhage necessitates close monitoring of patients.

[0004] The inventors recognized that the efficacy of ozagrel in treating acute thrombotic cerebral infarction and cerebral infarction-associated motor dysfunction suggests it is a potential antithrombotic pharmacophore. It is well known that GPIIb / IIIa on the surface of activated platelets binds to fibrinogen, inhibiting platelet aggregation. Furthermore, Arg-Gly-Asp-Phe is a competitive ligand for fibrinogen. The inventors hypothesized that conjugates of ozagrel and Arg-Gly-Asp-Phe might be excellent antithrombotic agents. Based on the experience that self-assemblies enhance biological activity and the self-assembly properties of conjugates of ozagrel and Arg-Gly-Asp-Phe, this invention proposes [ozagrel-RGDF]4. Summary of the Invention

[0005] The technical problem to be solved by this invention is to confirm that [ozagrel-RGDF]4 has excellent anti-arterial thrombotic effects. To achieve this objective, this invention employs the following three technical means.

[0006] The first technical means is to propose the ozagrel-RGDF tetramer of the present invention, namely [ozagrel-RGDF]4.

[0007]

[0008] The second technical approach involves proposing a method for preparing [ozagrel-RGDF]4, which includes the following seven steps:

[0009] 1) Preparation of Boc-Arg(NO2)-Gly-OBzl;

[0010] 2) Preparation of Boc-Arg(NO2)-Gly;

[0011] 3) Preparation of Boc-Asp(OBzl)-Phe-OBzl;

[0012] 4)Boc-Arg(NO2)-Gly-Asp(OBzl)-Phe-OBzl;

[0013] 5) Preparation of ozagrel-Arg(NO2)-Gly-Asp(OBzl)-Phe-OBzl;

[0014] 6) Preparation of ozagrel-Arg-Gly-Asp-Phe;

[0015] 7) Preparation of [Ozagrel-RGDF]4.

[0016] The third technical approach is to confirm the application of [ozagrel-RGDF]4 in the preparation of anti-arterial thrombotic drugs without bleeding side effects.

[0017] Compared with the prior art, the beneficial effects of the present invention are:

[0018] Experiments have shown that the [ozagrel-RGDF]4 of this invention not only has a good anti-arterial thrombotic effect but also has no bleeding side effects. Therefore, this invention provides a safe and effective technical means for anti-arterial thrombosis and enriches the selection of anti-arterial thrombotic drugs. Attached Figure Description

[0019] Figure 1 The synthetic route for ozagrel-RGDF is as follows: (i) DCC, HOBt, NMM, THF; (ii) ethyl acetate solution of 4N hydrogen chloride; (iii) Pd / C, MeOH; (iv) piperidine, DMF; (v) HATU, HOBt, NMM, THF; (vi) TFMSA, TFA.

[0020] Figure 2 The FT-ICR-MS and qCID spectra of [Ozagre-RGDF]4 are shown. Detailed Implementation

[0021] To further illustrate the present invention, a series of embodiments are given below. These embodiments are purely illustrative and are only used to specifically describe the present invention, and should not be construed as limiting the present invention.

[0022] Example 1: Preparation of Ozagrel-Arg-Gly-Asp-Phe

[0023] Example 1.1 Preparation of Boc-Arg(NO2)-Gly-OBzl

[0024] Dissolve 3.00 g of Boc-Arg(NO2) (9.40 mmol) in 25 mL of anhydrous tetrahydrofuran. Under ice-water cooling, add 2.30 g of N,N'-dicyclohexylcarbodiimide (DCC, 11.17 mmol) and 1.50 g of 1-hydroxybenzotriazole (HOBt, 11.11 mmol) to the resulting solution. Stir for 30 minutes under ice-water cooling until a colorless, insoluble substance appears in the solution. Under ice-water cooling, add 2.30 g of HCl·Gly-OBzl (11.41 mmol) to the above reaction solution, and adjust the pH to 9 with N-methylmorpholine (NMM). Stir the reaction mixture at room temperature for 12 hours; TLC (dichloromethane / methanol = 20 / 1) shows the disappearance of Boc-Arg(NO2). Filter the reaction mixture, and concentrate the filtrate under reduced pressure. The residue was dissolved in 100 mL of ethyl acetate. The resulting solution was washed with saturated sodium bicarbonate aqueous solution (20 mL × 3), saturated sodium chloride aqueous solution (20 mL × 3), 5% potassium bisulfate aqueous solution (20 mL × 3), saturated sodium chloride aqueous solution (20 mL × 3), saturated sodium bicarbonate aqueous solution (20 mL × 3), and saturated sodium chloride aqueous solution (20 mL × 3). The washed ethyl acetate solution was separated and dried over anhydrous sodium sulfate for 12 hours. The solution was filtered and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (dichloromethane / methanol = 30 / 1) to give 4.03 g of the title compound (92%) as a colorless solid.

[0025] Example 1.2 Preparation of Boc-Arg(NO2)-Gly

[0026] 1.68 g of Boc-Arg(NO2)-Gly-OBzl (3.35 mmol) was dissolved in 20 mL of methanol. A 2N NaOH aqueous solution was added dropwise to the resulting solution under ice-water cooling to adjust the pH to 13, and the mixture was stirred for 3 hours. TLC (ethyl acetate / water / glacial acetic acid = 4 / 1 / 1) showed the disappearance of Boc-Arg(NO2)-Gly-OBzl. The reaction mixture was cooled under ice-water and the pH was adjusted to 2 with a 4N hydrochloric acid aqueous solution. Insoluble matter was filtered off. The filtrate was extracted with ethyl acetate (30 mL × 3). The extracted ethyl acetate solution was separated and dried over anhydrous sodium sulfate for 12 hours. The solution was filtered and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (dichloromethane / methanol = 30 / 1) to give 1.2 g (89%) of the title compound as a colorless solid.

[0027] Example 1.3 Preparation of Boc-Asp(OBzl)-Phe-OBzl

[0028] The title compound was obtained as a colorless solid from 808 mg Boc-Asp(OBzl) (2.50 mmol) and 728 mg HCl·Phe-OBzl (2.50 mmol) using the procedure in Example 1.1. 1.52 g (87%) of the title compound was obtained.

[0029] Example 1.4 Preparation of HCl·Asp(OBzl)-Phe-OBzl

[0030] Under ice-water cooling, 1.00 g of Boc-Asp(OBzl)-Phe-OBzl was dissolved in 10 mL of ethyl acetate solution with 4N hydrogen chloride and stirred for 4 hours. TLC (ethyl acetate / water / glacial acetic acid = 4 / 1 / 1) showed the disappearance of Boc-Asp(OBzl)-Phe-OBzl. The reaction mixture was concentrated under reduced pressure, and the residue was dissolved in dried ethyl acetate and concentrated under reduced pressure to remove as much free hydrogen chloride as possible. This operation was repeated three times, and the residue was suspended in anhydrous diethyl ether (30 mL × 3). The supernatant diethyl ether was discarded to give 830 mg (97%) of the title compound as a colorless solid.

[0031] Example 1.5 Preparation of Boc-Arg(NO2)-Gly-Asp(OBzl)-Phe-OBzl

[0032] The title compound, as a colorless solid, was obtained from 1.00 g of Boc-Arg(NO2)-Gly (2.66 mmol) and 1.02 g of HCl·Asp(OBzl)-Phe-OBzl (2.22 mmol) using the procedure described in Example 1.1. TOF / MS: 819.3992 [M+H] + ; 1H NMR (300MHz, DMSO-d6) δ / ppm=8.39(d,J=7.5Hz,1H),8.21(d,J=8.3Hz,1H),8.02(d,J=5.4Hz,1H),7.33(d ,J=11.2Hz,8H),7.27(m,5H),7.19(d,J=7.5Hz,2H),6.99(d,J=7.6Hz,1H),5.01(m,4H),4.73(dt,J1=8.5 Hz,J2=5.1Hz,1H),4.50(q,J=7.4Hz,1H),3.95(s,1H),3.70(d,J=5.3Hz,2H),3.13(s,1H),3.02(dt,J1=8 .1Hz, J2=5.0Hz, 3H), 2.72 (dd, J1=6.1Hz, J2=5.1Hz, 1H), 2.56 (s, 1H), 1.59 (d, J=6.7Hz, 4H), 1.37 (s, 9H).

[0033] Example 1.6 Preparation of HCl·Arg(NO2)-Gly-Asp(OBzl)-Phe-OBzl

[0034] The title compound was obtained as a colorless solid from 1.00 g of Boc-Arg(NO2)-Gly-Asp(OBzl)-Phe-OBzl (1.22 mmol) using the operation of Example 1.4.

[0035] Example 1.7 Preparation of Ozagrel-Arg(NO2)-Gly-Asp(OBzl)-Phe-OBzl

[0036] The title compound, as a colorless solid, was obtained from 0.25 g ozag (1.10 mmol) and 0.95 g HCl·Arg(NO2)-Gly-Asp(OBzl)-Phe-OBzl (1.32 mmol) using the procedure described in Example 1.1. ESI-MS (m / e): 951.3 [M+Na] + MPa 118-119℃; 1H NMR (300MHz, DMSO-d6) δ / ppm=8.45(s,1H),8.31(d,J=11.3Hz,1H),8.00(t,J=9.2Hz,1H),7.91(d,J=11.9Hz,1H),7.74(s,1H),7.61(m,3H ),7.49(dd,J1=5.1Hz,J2=0.7Hz,1H),7.34(m,11H),7.19(m,8H),6.93(d,J=7.5Hz,1H),6.49(d,J=5.1Hz,1H),5.87(t,J=4.2Hz,1H),5.50 (dt,J1=6.1Hz,J2=1.0Hz,1H),5.31(dt,J1=6.1Hz,J2=1.0Hz,1H),5.11(m,4H),4.76(dt,J1=11.3Hz,J2=7.0Hz,1H),4.52(dt,J1=11.1Hz ,J2=6.1Hz,1H),4.12(dt,J1=10.7Hz,J2=6.9Hz,1H),4.00(d,J=9.1Hz,2H),3.22(m,2H),2.99(m,2H),2.71(d,J=7.0Hz,2H),1.59(m,4H).

[0037] Example 1.8 Preparation of Ozagrel-Arg-Gly-Asp-Phe Monomer

[0038] Under ice-water cooling, 0.10 g of ozagrel-Arg(NO2)-Gly-Asp(OBzl)-Phe-OBzl (0.11 mmol) was thoroughly mixed with 1000 μL of trifluoroacetic acid, followed by the addition of 200 μL of trifluoromethanesulfonic acid and stirring for 50 minutes. TLC (ethyl acetate / water / glacial acetic acid = 4 / 1 / 1) showed the disappearance of ozagrel-Arg(NO2)-Gly-Asp(OBzl)-Phe-OBzl. 15 mL of ice-cooled diethyl ether was added to the reaction mixture, causing a large amount of colorless solid to precipitate, and stirring was continued for 30 minutes. The solvent was removed by filtration, and the resulting solid was dissolved in 0.2 mL of distilled water. The pH of the resulting solution was adjusted to 7 by adding saturated sodium bicarbonate solution. After filtration to remove insoluble matter, the solution was purified by C18 column chromatography, and the solvent was removed by freeze-drying to give 0.05 g (67%) of the title compound as a colorless solid. ESI(+) / FT-MS(m / e):704.31126[M+H] + ; Mp187-188℃; 1HNMR(300MHz,DMSO-d6)δ / ppm=10.01(s,1H),8.77(d,J=7.8Hz,1H),8.40(d,J=9.9Hz,2H),7.77(s,1H), 7.54(d,J=7.9Hz,2H),7.38(m,2H),7.20(m,10H),7.05(s,1H),6.92(s,1H),6.78(d,J=15.8Hz,1H),5.21 (s,2H),4.39(p,J=7.5Hz,2H),4.19(q,J=6.2Hz,1H),3.87(dd,J1=6.6Hz,J2=3.1Hz,1H),3.61(dd,J1=6. 4Hz,J2=3.0Hz,2H),2.99(m,5H),2.42(m,3H),1.87(d,J=7.4Hz,1H),1.66(s,1H),1.49(t,J=7.2Hz,2H); 13 C NMR (75MHz, DMSO-d) 6) δ / ppm=176.29,173.38,172.37,170.48,168.87,165.32,157.94,139.59 ,139.40,138.47,137.96,135.04,130.06,129.23,128.50,128.31,127.9 1,125.73,120.09,56.05,52.90,51.32,49.63,43.28,41.38,40.84,40. 57,40.29,40.01,39.73,39.45,39.17,38.66,37.88,31.13,24.57;IR(cm -1 ):3273.31,1590.28,1515.21,1393.06,1345.66,1268.96,1229.79,1108.35,1078.29,1029.49,983.51,916.06,814.96,725.30,700.05,661.84.

[0039] Example 2: Preparation of Ozagrel-Arg-Gly-Asp-Phe Tetramer

[0040] 100 mg of ozagrel-Arg-Gly-Asp-Phe monomer was dissolved in 5 mL of ultrapure water. The resulting solution was vortexed for 15 minutes and then sonicated for 4 hours until it became completely colorless and transparent. This colorless and transparent solution was concentrated to dryness under reduced pressure to obtain the ozagrel-Arg-Gly-Asp-Phe tetramer, namely [ozagrel-Arg-Gly-Asp-Phe]4. The structure of [ozagrel-Arg-Gly-Asp-Phe]4 was confirmed by ultra-high resolution Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR-MS).

[0041] Figure 2 The FT-ICR-MS of A showed a [Ozagrel-Arg-Gly-Asp-Phe]+H peak at 704.31568 (theoretical value 704.31563), a divalent [Ozagrel-Arg-Gly-Asp-Phe]3+H peak at 1055.96921 (theoretical value 1055.96955), and a divalent [Ozagrel-Arg-Gly-Asp-Phe]4+H peak at 1407.61776 (theoretical value 1407.62345). To clarify the relationship between the [Ozag-Arg-Gly-Asp-Phe]+H peak, the [Ozag-Arg-Gly-Asp-Phe]3+H peak, and the [Ozag-Arg-Gly-Asp-Phe]4+H peak, the qCID spectrum of the [Ozag-Arg-Gly-Asp-Phe]4+H peak was determined.

[0042] Figure 2 The qCID of the [Ozagre-Arg-Gly-Asp-Phe]4+H peak of B shows a divalent [Ozagre-Arg-Gly-Asp-Phe]3+H peak at 1055.96934 (theoretical value 1055.96955), and a [Ozagre-Arg-Gly-Asp-Phe]+H peak at 704.31126 (theoretical value 704.31563). In other words, [Ozagre-Arg-Gly-Asp-Phe]4 is the only form in which Ozagre-Arg-Gly-Asp-Phe exists.

[0043] Example 3: Evaluation of the antivenous thrombotic activity of ozagrel-Arg-Gly-Asp-Phe tetramer

[0044] The antithrombotic activity of ozagrel-Arg-Gly-Asp-Phe tetramer was evaluated in a rat model of inferior vena cava ligation. Male SD rats (250±20g) were rested and fasted for one day at 25°C. Afterwards, they were randomly divided into groups of 11 rats each. Rats were administered either orally with a solution of warfarin and saline (positive control, 4.87 μmol / kg), or orally with saline (blank control, 10 mL / kg), or intravenously with a solution of ozagrel and saline (pharmacophore control, 43 μmol / kg), or orally with a solution of ozagrel-Arg-Gly-Asp-Phe tetramer and saline (0.43 μmol / kg). Thirty minutes later, the rats were anesthetized by intraperitoneal injection of 20% urethane solution. The anesthetized rats were fixed in a supine position on a surgical board, and the skin was prepared and disinfected. Then, the abdominal cavity was opened along the linea alba. The abdominal incision extends from the coagulation gland to expose a corner of the liver. Organs such as the small intestine are removed from the abdominal cavity and wrapped with gauze soaked in physiological saline. The connective tissue surrounding the blood vessels is bluntly dissected to expose the inferior vena cava and its branches. The abdominal aorta and inferior vena cava are separated below the renal vein, and the inferior vena cava is ligated at its junction with the left renal vein using sutures moistened with physiological saline. Organs such as the intestine, wrapped in gauze soaked in physiological saline, are returned to the abdominal cavity according to their anatomical position, and the abdominal cavity is sutured layer by layer. Because warfarin-treated rats are prone to death, the abdominal cavity is opened 30 minutes after suturing. Branches are ligated one by one, and a 2 cm section of the inferior vena cava is removed from the ligation point at the junction of the inferior vena cava and the left renal vein. The thrombus is then removed from the inferior vena cava and accurately weighed.

[0045] Table 1 shows the venous thrombus weights, indicating that the antithrombotic activity of the oral dose of 0.43 μmol / kg ozagrel-Arg-Gly-Asp-Phe tetramer was not significantly different from that of normal saline (P>0.05). Table 1 also shows that the oral dose of warfarin at 4.87 μmol / kg and the intravenous dose of ozagrel at 43 μmol / kg exhibited significant antithrombotic activity (P<0.01 compared to the normal saline group). Therefore, 0.43 μmol / kg [ozagrel-Arg-Gly-Asp-Phe]4 has no antithrombotic effect.

[0046] Table 1. Effects of Ozagrel-Arg-Gly-Asp-Phe on Venous Thrombosis

[0047] Therapeutic agents dose Severity of venous thrombosis, mean ± SD mg physiological saline 10mL / kg 17.6±2.5 warfarin 4.87 μmol / kg 9.1±1.3 Ozagre 43 μmol / kg 12.04±1.2 [[ozagrel-Arg-Gly-Asp-Phe]4] 0.43 μmol / kg 16.4 ± 2.2 a ]]

[0048] a) Compared with physiological saline, P>0.05; compared with warfarin and ozagrel, P<0.01; n=11.

[0049] Example 4: Evaluation of the anti-arterial thrombotic activity of ozagrel-Arg-Gly-Asp-Phe tetramer

[0050] 1) Pull the polyethylene tube into a thin tube with one end at an angle, with a fixed length of 10.0 cm, and insert it into the right jugular vein (larger diameter) and the left carotid artery (smaller diameter); the middle section of the polyethylene tube is fixed to a length of 8.0 cm, with the thrombus line pressed against the direction of carotid artery cannulation, and the tube must be filled with heparin before cannulation.

[0051] 2) Male SD rats weighing 200±20g were acclimatized to their environment and fasted for one day before surgery. They were randomly divided into four groups: saline group (blank control, oral dose of 0.3mL / 100g, 10 rats), aspirin group (positive control, oral dose of 167μmol / kg, 10 rats), ozagrel group (pharmacophore control, intravenous injection dose of 43μmol / kg, 10 rats), and [ozagrel-Arg-Gly-Asp-Phe] group (oral dose of 0.43μmol / kg, 10 rats). After 30 minutes, the rats were anesthetized by intraperitoneal injection of 20% urethane solution (7mL / kg), and surgery began 2 minutes later. During the surgery, the rat was placed supine on a fixation board. The skin of the neck was cut open, and the right common carotid artery and left jugular vein were separated. A precisely weighed silk thread was placed under the blood vessel and ligated at the distal end. A small incision was made at the distal end of the vein, and a cannula was inserted into the vein. Heparin was injected, and then the syringe used for heparin injection was removed. The suture was then secured, and the proximal end of the artery was clamped with an arterial clamp. A small incision was made at the distal end of the artery, and the arterial end was ligated. After securing the suture, the arterial clamp was released, establishing an extracorporeal circulation bypass. After 15 minutes of circulation, the vein was cut to observe whether blood circulation was normal. If blood circulation was normal, the silk thread with thrombus attached was removed from the arterial end. Uncoagulated blood was absorbed with filter paper, and the silk thread with thrombus attached was precisely weighed. The weight of the thrombus was obtained by subtracting the weight of the silk thread from the weight of the silk thread with thrombus attached. The data are listed in Table 2. Arterial blood was collected, centrifuged at 3000g for 10 minutes, and measured using the method in Example 5.

[0052] Table 2 shows that the thrombus weights indicate that [ozagrel-Arg-Gly-Asp-Phe]4 effectively inhibited arterial thrombosis in rats at an oral dose of 0.43 μmol / kg, exhibiting significantly greater anti-arterial thrombotic activity compared to saline and intravenous ozagrel at a dose of 43 μmol / kg (P<0.01); and comparable to the anti-arterial thrombotic activity of oral aspirin at a dose of 167 μmol / kg (P>0.05). This is an unexpected technical effect. Therefore, [ozagrel-Arg-Gly-Asp-Phe]4 possesses excellent anti-arterial thrombotic activity.

[0053] Table 2. Effects of [Ozagrel-Arg-Gly-Asp-Phe]4 on arterial thrombosis in rats.

[0054] Therapeutic agents dose Thrombus weight: mean ± SD mg physiological saline 10mL / kg 32.58±2.21 aspirin 167 μmol / kg 20.70±1.73 Ozagre 43 μmol / kg 26.24±1.22 [[ozagrel-Arg-Gly-Asp-Phe]4] 0.43 μmol / kg 20.55 ± 0.84 a ]]

[0055] a) Compared with normal saline and intravenous ozagrel at a dose of 43 μmol / kg, P < 0.01; compared with oral aspirin at a dose of 167 μmol / kg, P > 0.05; n = 10.

[0056] Example 5: Evaluation of the bleeding side effects of ozagrel-Arg-Gly-Asp-Phe tetramer

[0057] The hemorrhage side effects of rat plasma obtained from Example 4, which was administered with saline, orally administered with 0.43 μmol / kg dose of [ozagrel-Arg-Gly-Asp-Phe]4, and intravenously injected with 43 μmol / kg dose of ozagrel were assessed.

[0058] The assays were performed in accordance with the instructions for the Activated Partial Thromboplastin Time (APTT) kit, the Prothrombin Time (PT) kit, the Thrombin Time (TT) kit, and the Fibrinogen (FIB) Content Kit.

[0059] Specifically, the method can be described as follows: Place one magnetic bead, 100 μL of rat plasma, and 100 μL of APTT reagent in a sample cup, preheat at 37°C for 5 minutes, then add 100 μL of CaCl2 solution to the sample cup and measure the activated partial thromboplastin time (APTT); place one magnetic bead and 100 μL of rat plasma in a sample cup, preheat at 37°C for 5 minutes, then add 200 μL of preheated thrombin reagent (at 37°C) to the sample cup and measure the prothrombin time (PT); place one magnetic bead and 200 μL of rat plasma in a sample cup, preheat at 37°C for 3 minutes, then add 200 μL of thrombin reagent to the sample cup and measure the thrombin time (TT); place one magnetic bead and 200 μL of diluted plasma (from 100 μL of plasma using 900 μL of...) The FIB plasma diluent was placed in a sample cup and preheated at 37°C for 3 minutes. Then, 100 μL of fibrinogen was added to the sample cup, and the fibrinogen content (FIB) was measured. The results are shown in Table 3.

[0060] The data in Table 3 show that there were no significant differences in the activated partial thromboplastin time (APTT), prothrombin time (PT), thrombin time (TT), and fibrinogen (FIB) levels between rats treated with [ozagrel-Arg-Gly-Asp-Phe]4 at an oral dose of 0.43 μmol / kg and rats treated with saline (P>0.05). The data in Table 3 also show that the activated partial thromboplastin time (APTT), prothrombin time (PT), thrombin time (TT), and fibrinogen (FIB) levels in rats treated with ozagrel at an intravenous dose of 43 μmol / kg were significantly higher than those in rats treated with saline (P<0.01). In other words, [Ozagrel-Arg-Gly-Asp-Phe]4 does not have the bleeding side effect of ozagrel. This is an unexpected technical effect.

[0061] Table 3. Effects of [Ozagrel-Arg-Gly-Asp-Phe]4 on four coagulation parameters (mean ± SD)

[0062] therapeutic agents from rat plasma PT(s) TT(s) APTT(s) FIB(g / L) physiological saline 10.35±1.04 29.88±2.29 19.98±1.44 2.09±0.13 Ozagre 15.53 ± 1.56 a ]] 36.63 ± 2.40 a ]] 31.33 ± 3.49 a ]] 2.57 ± 0.19 a ]] [[ozagrel-Arg-Gly-Asp-Phe]4] <![CDATA[10.21±1.03 b ]]> <![CDATA[29.65±2.29 b ]]> <![CDATA[19.82±1.40 b ]]> <![CDATA[2.05±0.14 b ]]>

[0063] a) Compared with physiological saline, P<0.01; b) Compared with physiological saline, P>0.05; n=10.

Claims

1. The ozagrel-RGDF tetramer with the following structural formula, namely [ozagrel-RGDF]4, 2. The method for preparing the ozagrel-RGDF tetramer according to claim 1, characterized in that... The preparation method includes the following steps: 1) Preparation of Boc-Arg(NO2)-Gly-OBzl; 2) Preparation of Boc-Arg(NO2)-Gly; 3) Preparation of Boc-Asp(OBzl)-Phe-OBzl; 4)Boc-Arg(NO2)-Gly-Asp(OBzl)-Phe-OBzl; 5) Preparation of ozagrel-Arg(NO2)-Gly-Asp(OBzl)-Phe-OBzl; 6) Preparation of ozagrel-Arg-Gly-Asp-Phe; 7) Preparation of [Ozagrel-RGDF]4.

3. The use of [ozagrel-RGDF]4 as described in claim 1 or 2 in the preparation of anti-arterial thrombotic drugs.

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