JPH07309746A - Injection containing cysteine derivative - Google Patents

Abstract

PURPOSE:To obtain an injection having thrombogenesis inhibiting action and thrombolytic action and capable of effectively using for removal of thrombus and prevention of re-obstruction in cardiac infarction and cerebral infarction. CONSTITUTION:This injection contains a cysteine derivative such as cysteine, cysteine methyl ester, cysteine ethyl ester, N-acetyl-L(D)-cysteine or glutathione as a thrombogenesis inhibitor and/or a thrombolytic agent. Further, a tissue plasminogen activator (tPA) may be blended therein.

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to an injectable preparation containing a cysteine derivative, which is used to prevent reocclusion of blood vessels after thrombolysis in myocardial infarction or cerebral infarction and to more effectively carry out thrombolytic therapy.

[0002]

BACKGROUND OF THE INVENTION Tissue plasminogen activator (tPA) for removing thrombus generated in blood vessels during myocardial infarction or cerebral infarction,
Percutaneous coronary angiolysis (PTCR) and percutaneous coronary dilatation (PTCA) using a thrombolytic agent such as urokinase are widely used. At this time, even if the blood vessel is opened, reocclusion due to the generation of platelet thrombus or fibrin clot at the damaged site of the blood vessel becomes a problem. In order to prevent this, drugs such as heparin which is an anticoagulant and aspirin which is an antiplatelet agent, ticlopidine, etc. are used, but they are weak in terms of efficacy, and when a high dose is used, side effects such as bleeding tendency are brought about, and more There is a need for effective and safe drugs.

[0003]

[Means for Solving the Problems] Cysteine, which is a sulfur-containing amino acid, is a biological component and is known to have an antioxidative effect by itself or in peptides or proteins such as glutathione, and is also a nutritional component. Even if it is administered in a large amount, it is used as a protein source without causing serious side effects.

The present inventors have been studying the action of cysteine derivatives in a model animal for thrombus formation. Surprisingly, the cysteine derivative alone shows a lytic action of thrombus produced together with a thrombus formation inhibitory action. ,Also,
It was found that the time to the subsequent reocclusion can be significantly prolonged by using it in combination with tPA and the thrombolytic action of tPA can be more effectively exerted, and the present invention has been completed. The above problems can be solved by the present invention described below.

(1) An injection containing a cysteine derivative as a thrombus formation inhibitor and / or a thrombolytic agent.

(2) The injection according to (1) above, wherein the cysteine derivative is at least one selected from the group consisting of cysteine, cysteine methyl ester, cysteine ethyl ester, N-acetyl-L (D) -cysteine and glutathione. Agent.

(3) The injectable preparation according to (1) or (2) above, which contains a thrombolytic agent and / or a blood anticoagulant other than the cysteine derivative, and the cysteine derivative.

(4) The injectable agent according to the above (1) to (3), wherein the thrombolytic agent other than the cysteine derivative is a tissue plasminogen activator.

In the present invention, the cysteine derivative is used as a thrombus formation inhibitor and / or thrombolytic agent, and is used alone or in combination with other thrombolytic agents and / or blood anticoagulants to remove thrombus during myocardial infarction or cerebral infarction. And it is effective in preventing reclosure.

As the cysteine derivative of the present invention, cysteine, ester derivative of cysteine, acetyl derivative or other various derivatives can be used without any limitation. Moreover, you may use these as salts, such as a hydrochloride and a sulfate, depending on the case.

Specifically, L-cysteine hydrochloride and DL-
Cysteine hydrochloride monohydrate, L-cysteine methyl ester hydrochloric acid, L-cysteine ethyl ester hydrochloric acid,
Examples include N-acetyl-L-cysteine and glutathione.

In the present invention, the content of the cysteine derivative is 1.0 to 200 mg / ml, more preferably 10 to 100.
Contained in mg / ml. If the amount is less than 1.0 mg / ml, it cannot be expected that the thrombus generated will be dissolved.

In the present invention, when the above-mentioned cysteine derivative and tPA are used in combination, the time until the subsequent re-occlusion is significantly extended and the thrombolytic action and thrombus formation inhibiting action are made more effective.

In the present invention, urokinase, streptokinase, plasminogen proactivator, etc. may be contained as other thrombolytic agents. Furthermore, as blood anticoagulant, heparin, sodium citrate, ethylenediaminetetraacetic acid disodium salt (EDTA-2N
a), ethylenediaminetetraacetic acid dipotassium salt (EDTA
-2K), ethylenediaminetetraacetic acid tripotassium salt (ED
TA-3K) or the like may be blended.

In the present invention, the cysteine derivative and the other thrombolytic agent and / or blood anticoagulant are used as a solution, and the solvent thereof includes, but is not limited to, distilled water, physiological saline and the like. Also, the cysteine derivative and the other thrombolytic agent and / or blood anticoagulant may be used as one agent, but may be administered separately.

The method for producing the injection of the present invention is not particularly limited, and the cysteine derivative and other necessary thrombolytic agent and / or blood anticoagulant may be dissolved in the above-mentioned solvent according to a conventional method. The resulting injection is stored in a bag-shaped container made of soft plastic resin, glass, a vial made of hard plastic resin, a syringe, or the like.

Next, the injection containing the cysteine derivative of the present invention and the method of using the same will be specifically described.

The cysteine derivative is dissolved in distilled water that has been deoxygenated in advance, the insoluble matter is removed by a filter, and the mixture is dispensed into a glass vial and sealed under a nitrogen atmosphere. Regarding administration, since a catheter is inserted into the blockage site of the heart or cerebral blood vessels during PTCA, the cysteine derivative solution is continuously administered immediately after administration of tPA and urokinase.

The concentration at the time of administration varies depending on the degree of symptoms, but it is 1 to 200 mg / ml, more preferably 10 to 1
Dilute with physiological saline to 00 mg / ml. Alternatively, it may be directly administered as a mixed injection in a tPA or urokinase solution. After thrombolytic therapy or PTCA, it is continuously administered intravenously. In this case, if a syringe pump capable of continuously injecting a very small amount is used, or if infusion of body fluid such as water or electrolyte has been performed, it can be mixed into the infusion agent.

[0020]

The present invention will be described in more detail by way of examples.

Example 1 A guinea pig (Hartley, male, body weight 450 to 600 g) was anesthetized with pentobarbital, and then the common carotid artery was exposed. Attach a probe for blood flow measurement to this, ultrasonic blood flow measuring device (Advance)
Connected to. A cannula for drug administration was placed in the femoral vein. Irradiate monochromatic light (wavelength 540 nm, 500,000 lux) to the central side from the probe attachment site (light irradiation device, Hamamatsu Holonics Co., Ltd.), and stabilize the blood flow, then from the femoral vein, dye Rose Bengal (Wako Pure Chemical, 10 mg). / kg) was administered, and changes in blood flow from administration to occlusion and occlusion time were measured.

The cysteine solution was dissolved in 10 ml of deoxygenated distilled water of L-cysteine hydrochloride (Nakalalask) and filtered (pore size 0.2 μm, Millipore).
After sterilization by, the mixture was dispensed into a sterilized glass vial and sealed in a nitrogen atmosphere. At the time of administration, it was diluted with physiological saline to obtain an injection having a cysteine concentration of 100 mg / ml. Immediately after administration of rose bengal, 2 ml each of cysteine solution and physiological saline containing no cysteine were continuously infused by a syringe pump for 20 minutes. As shown in FIG. 1, when cysteine was not administered, blood flow stopped in 10 to 15 minutes, and formation of thrombus clots could be confirmed by observation of this site with a scanning electron microscope.

On the other hand, when cysteine was administered, there was no decrease in blood flow as shown in FIG. 2, and no destruction of endothelial cells or thrombus formation was observed even by observation with a scanning electron microscope.

Immediately after administration of rose bengal and administration of 1 ml of an injection having a cysteine concentration of 100 mg / ml immediately after the blood flow was stopped, resumption of blood flow was observed, and the blood flow continued for 2 hours or more.

(Example 2) Rat (SD, male, body weight 2
70 to 400 g) and the probe was attached to the common carotid artery as in Example 1. The thrombus formation was prepared by dropping 5 ul of 20% silver nitrate solution on the outside of the blood vessel. When nothing was administered, blood flow was stopped in 12 to 18 minutes as shown in FIG. 3 and thrombus formation was observed by observation with a scanning electron microscope, but 3 minutes after the blood flow was stopped, the cysteine concentration was 100 mg / ml. When 1 ml of the injection was administered, as shown in FIG. 4, the blood flow was reopened thereafter and the blood flow continued, and no endothelial cell destruction or thrombus formation was observed even by observation with a scanning electron microscope.

(Example 3) As in Example 1, a guinea pig (Hartley, male, body weight 450 to 600 g) was attached to the common carotid artery with a probe, and rose bengal was administered under light irradiation. Five minutes after the blood flow was stopped, tPA (trade name; activacin, Kyowa Hakko) was administered, and the blood flow change was measured for 120 minutes. In the case of administration of only tPA, temporary reocclusion occurred in 23 minutes after the blood flow was opened, but the cysteine concentration was 100 mg.
When 2 ml of the injection solution of / ml was administered, the blood flow continued for an average of 88 minutes thereafter. Moreover, the same amount of cysteine and tPA were used as one drug, and as a result, similar stable blood flow was exhibited.

Example 4 The thrombolytic action of cysteine was examined as follows. Fibrinogen concentration 1.25m
To 2 ml of a g / ml solution, 0.5 U / ml thrombin was added and well stirred, and then DL-cysteine monohydrate monohydrate 0.06 mg / ml was added. Using a spectrophotometer (U-2000 Spectroscopy), a wavelength of the obtained solution of 500
The change in absorbance with time in the nm region was measured. As the fibrinogen, type 1-S obtained by separating bovine fibrinogen (Sigma) from SDS-polyacrylamide gel was used.

The results are shown in FIG. The results show that the solution added with DL-cysteine monohydrate (line A) has a significantly lower absorbance than the solution without added cysteine derivative (line D), that is, DL-cysteine monohydrate has a fibrinogen content of It was found that the change in fibrin polymer was suppressed.

DL-Cysteine hydrochloride monohydrate was added to 0.015 mg / ml (B line) and 0.03 mg / ml (C
(Line) The completely same test was performed on the added solution to examine the thrombolytic action. From the results, it was found that the inhibitory effect of DL-cysteine monohydrate monohydrate on the conversion of fibrinogen into fibrin polymer increases depending on the concentration.

(Example 5) DL-Cysteine hydrochloride monohydrate of Example 4 and L-cysteine methyl ester hydrochloride, L-cysteine ethyl ester hydrochloride, N-acetyl-L-cysteine and glutathione were used instead. The same test as in Example 4 was performed. Table 1 shows the minimum effective concentration showing the thrombolytic effect for each cysteine derivative.

[0031]

[Table 1]

As shown by the results, each cysteine derivative showed an excellent thrombolytic effect at a very low dose.

[0033]

INDUSTRIAL APPLICABILITY As described above, the injection containing the cysteine derivative of the present invention has an action of inhibiting thrombus formation and a dissolving action of thrombus generated, and prevents thrombus removal and re-occlusion during myocardial infarction or cerebral infarction. Is possible. In addition, when used together with tPA, the time until the subsequent re-occlusion is significantly extended and the thrombolytic action can be more effectively exhibited.

[Brief description of drawings]

FIG. 1 shows the results of blood flow measurement in the common carotid artery of guinea pigs when cysteine-free physiological saline was administered after rose bengal was administered.

FIG. 2 shows the results of blood flow measurement in the common carotid artery of guinea pigs when an injection containing cysteine was administered after administration of rose bengal.

FIG. 3 shows the results of blood flow measurement of the common carotid artery of a rat when nothing was administered after a 20% silver nitrate solution was dripped on the outside of a blood vessel to form a thrombus.

FIG. 4 shows the results of blood flow measurement in the common carotid artery of a rat when a 20% silver nitrate solution was dropped on the outside of a blood vessel to form a thrombus, and then an injection containing cysteine was administered.

FIG. 5 shows the time change of the absorbance in a wavelength region of 500 nm of a fibrinogen solution to which thrombin was added and then cysteine was added.

Claims (4)

[Claims] 1. An injection containing a cysteine derivative as a thrombus formation inhibitor and / or a thrombolytic agent. 2. The cysteine derivative is cysteine, cysteine methyl ester, cysteine ethyl ester,
The injection according to claim 1, which is at least one of the group consisting of N-acetyl-L (D) -cysteine and glutathione. 3. The injectable composition according to claim 1, which contains a thrombolytic agent and / or a blood anticoagulant other than the cysteine derivative, and the cysteine derivative. 4. The injection according to claim 1, wherein the thrombolytic agent other than the cysteine derivative is tissue plasminogen activator.