JP5553306B2 - Radiation protection agent - Google Patents

Description

  The present invention relates to a medicament useful for protection against radiation damage by radiation exposure or radiation therapy.

Workers at nuclear power plants, non-destructive inspectors, clinical technologists handling radiopharmaceuticals, doctors engaged in radiotherapy such as X-ray examinations and cancer, and radiological technologists are always exposed to radiation during work. There is a possibility. In addition, when a nuclear power plant accident occurs, workers and nearby residents may be exposed to a large amount of radiation at once.
When exposed to radiation, oxygen radicals are generated in the living body, and the generated oxygen radicals cause damage such as cell death and mutation. Depending on the absorbed dose of radiation, the hematopoietic / immune system, digestive system, respiratory system, central nervous system, etc. may be damaged, and the exposed person may die due to this.

  In addition, cancer patients undergoing radiation therapy receive a large amount of radiation in the affected area. At this time, normal tissues around the affected area also receive radiation, so oxygen radicals generated by the radiation cause hematopoiesis / immune system, digestive system. May cause trouble.

  For such radiation damage, Patent Document 1 discloses that 2-pyranone derivatives increase specific blood cells that are known to decrease due to immune abnormalities. Patent Document 2 discloses that the survival rate of X-irradiated mice is significantly increased by administration of 1,2-phenyl-1,2-benzoisoselenazol-3 (2H) -one. ing. However, these agents are still unsatisfactory as radiation protection agents that protect against radiation exposure and damage associated with radiation therapy.

  In addition, previous studies by the present inventors have revealed that nitric oxide is involved in the acquisition of transient radiation resistance and the radiation adaptive response (Non-Patent Documents 1 to 4). It was revealed that treatment of wtp53 cells with isosorbide nitrate, which is a nitric oxide generator, acquired transient radiation resistance (Non-patent Document 3).

  On the other hand, nitroprusside is a known nitric oxide generator and is marketed as a blood pressure lowering agent. Moreover, blood homocysteine concentration inhibitory action (patent document 3) is known as a pharmacological effect of nitroprusside. However, it is not known that this compound is useful as an agent for protecting against radiation damage.

JP-A-5-213758 Japanese Patent Laid-Open No. 1-135718 Japanese Patent Laid-Open No. 2003-95959

Int J Radiat Biol. 2000 Dec; 76 (12): 1649-57 Radiat Res. 2001 Mar; 155 (3): 387-96 Cancer Res. 2007 Sep; 67 (18): 8574-9 Int J Radiat Oncol Biol Phys. 2008 Jun; 71 (2): 550-8

  In view of the above circumstances, the problem to be solved by the present invention is to provide a medicine that prevents or treats radiation exposure or a disorder associated with radiation therapy and increases the survival rate.

  As a result of intensive studies to solve the above problems, the present inventor has unexpectedly been useful for protecting against radiation damage because nitroprusside, which has been conventionally known as a therapeutic agent for hypertension and angina pectoris, is effective. It has been found that the survival rate of the woken individual is increased.

  In addition, an effective dosing schedule for protection from radiation damage was investigated. The half-life of the reaction product of oxygen radicals generated by radiation is usually said to be 5 minutes to 20 hours. Since a good radioprotective effect was observed by administering nitroprusside immediately after irradiation, it was speculated that nitric oxide produced from nitroprusside would eliminate oxygen radicals generated by radiation. However, after further detailed examination, surprisingly, in addition to administration of nitroprusside immediately after irradiation, additional nitroprusside was administered at the time when the generation of oxygen radicals due to radiation had already ended (after 1 day after irradiation). By doing so, it was found that the radiation protection effect was enhanced. Such enhancement of the radiation damage protective effect was confirmed only for nitroprusside among the nitric oxide generators tried.

The present invention has been completed based on the above findings. That is, the present invention is as follows.
[1] A radiation protective agent containing nitroprusside or a pharmacologically acceptable salt thereof.
[2] The radiation injury protective agent according to [1], wherein administration is started within 60 minutes before or after radiation exposure.
[3] The radiation damage protective agent according to [2], which is additionally administered one or more times after one day after radiation exposure and within 10 days.
[4] The radiation damage protective agent according to [3], which is additionally administered on the next day after radiation exposure.
[5] The radiation protection agent according to [4], which is further administered 2 days after radiation exposure, [6] The further administration, 6 to 10 days after radiation exposure, according to [4] or [5] Radiation protection agent.
[7] The radiation injury protective agent according to any one of [1] to [6], wherein nitroprusside is administered at a dose of 0.9 to 3.6 mg / kg per administration.
[8] The radiation damage protective agent according to any one of [1] to [7], wherein the radiation damage is acute radiation damage.
[9] The radiation damage protective agent according to [8], wherein the acute radiation damage is a hematopoietic / immune system disorder.

  ADVANTAGE OF THE INVENTION According to this invention, the improvement effect of a hematopoietic / immune system function, the life extension effect, and the improvement effect of a survival rate are acquired, and the radiation damage protective agent useful for the disorder | damage | failure accompanying radiation exposure or radiation therapy can be provided. Moreover, it has low toxicity and can protect against radiation damage at low doses. Therefore, it is useful from the viewpoint of reducing radiation damage and maintaining health not only for treatments when exposed to radiation in a short period of time, but also for patients who need long-term radiation therapy such as cancer, and workers engaged in radiation-related work. It is.

The survival rate after X-ray irradiation of mice treated with sodium nitroprusside after X-ray irradiation (4 times in total, immediately after irradiation, 1 day, 2 days, and 7 days after X-ray irradiation) and mice only with X-ray irradiation are shown. FIG. White blood cells of mice treated with sodium nitroprusside after X-ray irradiation (4 times in total, immediately after irradiation, 1 day, 2 days and 7 days after X-ray irradiation) and 14 days after X-ray irradiation It is a figure which shows the value of erythrocytes and platelets. It is a figure which shows the survival days after X-ray irradiation of the mouse | mouth which administered sodium nitroprusside, isosorbide nitrate, nicorandil, nitroglycerine, or nipradilol, and the mouse | mouth (NT) which does not administer a chemical | medical agent only after X-ray irradiation. . Moreover, the control in the figure is a mouse that was not irradiated with X-rays and was administered only with the drug according to the administration schedule. X-rays of a mouse administered with sodium nitroprusside, isosorbide nitrate, nicorandil, nitroglycerin, or nipradilol only immediately after X-ray irradiation and one day after X-ray irradiation, and a mouse (NT) not administered with a drug only by X-ray irradiation. It is a figure which shows the survival days after a beam irradiation. Moreover, the control in the figure is a mouse that was not irradiated with X-rays and was administered only with the drug according to the administration schedule. Survival days after X-ray irradiation for mice administered with sodium nitroprusside and mice not administered with drug (NT) only 3 times after X-ray irradiation and 1 day and 2 days after X-ray irradiation. FIG. Moreover, the control in the figure is a mouse that was not irradiated with X-rays and was administered only with the drug according to the administration schedule. Mice that received sodium nitroprusside, isosorbide nitrate, nicorandil, nitroglycerin, or nipradilol immediately after X-ray irradiation and 1 day, 2 days, and 7 days after X-ray irradiation, and mice that received no X-ray irradiation alone It is a figure which shows the survival days after X-ray irradiation of (NT). Moreover, the control in the figure is a mouse that was not irradiated with X-rays and was administered only with the drug according to the administration schedule.

  The present invention provides a radiation damage protective agent containing nitroprusside or a pharmacologically acceptable salt thereof.

Nitroprusside in the present invention is a compound represented by the formula (Fe (CN) ₅ NO) ²⁻ and has an action of releasing nitric oxide. When exposed to radiation, oxygen radicals are generated in the body, causing various disorders. Although not theoretically constrained, nitric oxide can eliminate oxygen radicals generated by radiation by radical reaction, and thus nitroprusside is considered to have a radiation damage protection ability.

  Examples of pharmacologically acceptable salts of nitroprusside include salts with inorganic bases and salts with organic bases.

  Preferable examples of the salt with an inorganic base include alkali metal salts such as sodium salt and potassium salt; alkaline earth metal salts such as calcium salt and magnesium salt; and aluminum salt and ammonium salt.

  Preferable examples of the salt with an organic base include salts with trimethylamine, triethylamine, pyridine, picoline, ethanolamine, diethanolamine, triethanolamine, dicyclohexylamine, N, N-dibenzylethylenediamine and the like.

Preferable examples of the pharmacologically acceptable salt of nitroprusside include sodium salt (Na ₂ Fe (CN) ₅ NO) and potassium salt (K ₂ Fe (CN) ₅ NO) of nitroprusside.

  Nitroprusside or a pharmacologically acceptable salt thereof may be crystalline or non-crystalline and may exist in the form of hydrates and / or solvates. And / or solvates are also encompassed by “nitroprusside or a pharmaceutically acceptable salt thereof”. Compounds containing various amounts of water obtained by methods such as stoichiometric amounts of hydrates and lyophilization are also within the scope of the present invention. The sodium salt or potassium salt of nitroprusside is usually used in the form of a dihydrate.

The radiation in the present invention includes α rays, β rays, γ rays emitted from radioactive substances, artificially created X rays, proton rays, carbon rays, neutral rays, and electron rays. This is a disorder associated with such radiation exposure or radiation therapy, and does not include exposure from naturally occurring radiation sources (natural exposure). For example, acute and / or late radiation damage caused by systemic radiation exposure due to a nuclear accident or nuclear explosion, or acute radiation exposure due to local radiation exposure due to radiation or a radiation exposure accident for medical purposes such as cancer treatment And / or late radiation damage. Among these, it is preferable to use for protection against local or systemic acute radiation damage, and particularly preferred is a hematopoietic / immunological system disorder.
In addition, the protection in the present invention includes prevention and / or treatment of such radiation exposure and damage associated with radiation therapy, but is preferably used for treatment.

  The radiation damage protective agent of the present invention contains nitroprusside or a pharmacologically acceptable salt thereof as an active ingredient, and can also be contained as a mixture with any other active ingredient for treatment. The radiation protection agent is produced by any method well known in the technical field of pharmaceutics by mixing the active ingredient with one or more pharmacologically acceptable carriers.

  As the pharmacologically acceptable carrier, various organic or inorganic carrier substances commonly used as pharmaceutical materials are used. Specific examples thereof include excipients, lubricants, binders, disintegrants, liquids in solid preparations. Solvents, solubilizers, suspending agents, tonicity agents, buffers, soothing agents and the like in the preparation can be mentioned. In formulation, formulation additives such as preservatives, antioxidants, coloring agents, sweeteners and the like may be used as necessary.

  In addition, it is desirable to use a route of administration that is most effective in the treatment, and it is usually administered parenterally or orally such as transdermally or intravenously. Formulations suitable for parenteral administration preferably comprise a sterile aqueous solution containing the active compound that is isotonic with the blood of the recipient. For example, in the case of an injection, a solution for injection is prepared using a carrier comprising a salt solution, a glucose solution, or a mixture of salt water and a glucose solution. To these parenteral agents, a solubilizing agent, a buffering agent, a pH adjusting agent, a tonicity agent, a soothing agent, a preservative, and the like can be further added as necessary. A formulation suitable for parenteral administration may be prepared by suspending nitroprusside in the present invention or a pharmacologically acceptable salt thereof in distilled water for injection or vegetable oil. Depending on the case, a base, a suspending agent, a thickener and the like can be added. In addition, a suitable formulation for parenteral may be a form in which the powder or lyophilized product of nitroprusside in the present invention or a pharmacologically acceptable salt thereof is dissolved upon use, and if necessary, an excipient Etc. can be added. Examples of oral preparations include tablets (including sublingual tablets and orally disintegrating agents), capsules (including soft capsules and microcapsules), powders, granules, troches, syrups, emulsions, suspensions, and the like. . These preparations may be controlled-release preparations such as immediate-release preparations or sustained-release preparations (eg, sustained-release microcapsules).

  In addition, since the antihypertensive agent (injection) which uses sodium nitroprusside as an active ingredient is already used clinically as nitroprusside in this invention, or its pharmacologically acceptable salt, [Nitopro (Maruishi Pharmaceutical Co., Ltd. ), Nitropress (manufactured by Abbott Co., Ltd.), Nipride (manufactured by Roche Co., Ltd.)], and the radiation damage protective agent of the present invention, the above-mentioned commercial preparation can be used as it is.

  The administration target of the radiation protection agent of the present invention is a mammal, and examples thereof include humans, dogs, cats, cows, horses, pigs, etc., preferably humans.

The radiation damage protective agent of the present invention is preferably administered immediately before or immediately after exposure to radiation and generation of oxygen radicals in a living body, specifically, within 60 minutes before or after radiation exposure, preferably 30 Administration is started within minutes, more preferably within 15 minutes, even more preferably within 10 minutes, particularly preferably within 5 minutes. Further, it is preferable from the viewpoint of prolonging the life and improving the survival rate at the time when the generation of oxygen radicals by radiation has already ended after radiation exposure (after one day from radiation exposure). Specifically, after one day after radiation exposure and within 10 days (more preferably within 8 days, particularly preferably within 7 days), one more time (for example, once, preferably twice, more preferably 3 times). It is preferable to administer additional doses. The interval between administrations is usually 0.5 days or more (for example, 1 day), but is limited to this as long as the radiation protection effect is enhanced as compared with single administration within 60 minutes before or after radiation exposure. Not. For example, the additional administration is preferably performed on the next day after the radiation exposure, and more preferably on the next day and 2 days after the radiation exposure.
Further, in addition to the additional administration on the next day of radiation exposure, or the next day and 2 days later, the additional administration 6 to 10 days after radiation exposure (preferably 7 days later) is a viewpoint of further improving the life extension effect and survival rate. To preferred. Specific examples of administration schedules include a total of 4 administrations within 60 minutes before or after radiation exposure, 1 day, 2 days and 7 days after radiation exposure.
Note that the above description of additional administration does not exclude administration at other times.

  The dose of the radiation protection agent of the present invention can be appropriately selected according to the purpose of protection (prevention or treatment) of radiation injury and conditions such as the age and condition of the patient, but nitroprusside or its drug in vivo. It is preferable to administer such that the salt concentration that is physically acceptable is 15 to 60 μM (preferably 30 to 60 μM) immediately after administration. Specifically, the dose is set on the assumption that nitroprusside is equally distributed to the extracellular fluid, the average extracellular fluid volume, the molecular weight of nitroprusside (251.95, 297.95 when sodium nitroprusside is used) and the above. It can be calculated from the concentration immediately after administration of nitroprusside in vivo. The average amount of extracellular fluid is obtained from body weight (kg) × 0.2 for humans and body weight (kg) × 0.35 for mice, with the specific gravity of the extracellular fluid being 1.00. However, a low dose that does not cause a blood pressure lowering effect is preferable, and the dose per administration is 0.9 to 3.6 mg, more preferably 1.8 to 3.6 mg as nitroprusside per kg body weight, Particularly preferred is 2.3 to 3.6 mg. Further, the dose limiting rate is 2.0 to 2.5 μg / kg / min.

Hereinafter, the present invention will be described more specifically with reference to examples. The present invention is not limited by these.
The materials used in the examples are collectively described below.

<Normal human fibroblasts>
AG1522 cells were used. AG1522 cells were cultured using Dulbecco's modified medium containing 15% fetal calf serum and 20 mM HEPES.
<Normal mouse>
Eight week old jcl: ICR mice were used. jcl: ICR mice were raised on commercial chow.
<Nitroprusside>
Nitopro (registered trademark, sodium nitroprusside injection solution) manufactured by Maruishi Pharmaceutical Co., Ltd. was used.
<Isosorbide nitrate>
Nitrol (registered trademark) manufactured by Eisai Co., Ltd. was used.
<Nitroglycerin>
Millisroll (registered trademark) manufactured by Nippon Kayaku Co., Ltd. was used.
<Nicorandil>
Nicorandil intravenous infusion manufactured by Fuji Pharmaceutical Co., Ltd. was used.
<Nipradilol>
Niprazirol ophthalmic solution 0.25% “Nitten” manufactured by Nitten Co., Ltd. was used.

  Further, in the examples, “the sodium nitroprusside was administered so that the concentration immediately after administration in vivo was 30 μM” means that the sodium nitrate is equally distributed to the extracellular fluid, and the average extracellular fluid The dose was calculated from the amount, the molecular weight of sodium nitroprusside (297.95), and the concentration immediately after administration of sodium nitroprusside (30 μM) in vivo, and administered. The average amount of extracellular fluid was determined from mouse body weight (kg) × 0.35, with the specific gravity of the mouse extracellular fluid being 1.00.

The test methods and results are described below.
<Toxicity test>
Sodium nitroprusside sodium was intraperitoneally administered to normal mice so that the concentration immediately after administration in vivo was 30 μM (dose: 2.8 to 3.2 mg / kg), and this intraperitoneal administration (first time) 1 day after administration, 2 days and 7 days later, sodium nitroprusside sodium was further administered in the same manner as in the first administration, and the follow-up was observed. No death of the mice was observed (total 4 administrations, n = 3).
In addition, isosorbide nitrate, nicorandil, nitroglycerin and nipradilol, which have the same action of releasing nitric oxide as sodium nitroprusside, were performed in the same manner, but no mice died from these drugs.

<Survival after X-ray irradiation>
1. AG1522 cells
(1) For an AG1522 cell culture dish, use an X-ray generator (HITEX 150 Hitex Co., Ltd.) to irradiate 3.5 Gy at a dose rate of 1 Gy / min (with a 0.5 mm aluminum filter). When used, the survival rate was about 10% (average value of 5 dishes).
(2) Several AG1522 cell culture dishes were prepared, and 120 minutes after adding sodium nitroprusside so that the sodium nitroprusside concentration in the culture medium of each dish was 0.3 μM or 1.0 μM, an X-ray generator (HITEX 150 Hitex Co., Ltd.) was used, and X-rays were irradiated to each dish at a dose rate of 1 Gy / min at 3.5 Gy (using a 0.5 mm aluminum filter). When the sodium concentration is 0.3 μM, it is 48%, and when it is 1.0 μM, it is 57% (both are average values of 5 dishes), and the survival rate compared to (1) without adding sodium nitroprusside. Rose.
(3) As with sodium nitroprusside, isosorbide nitrate, nicorandil, nitroglycerin and nipradilol, which have the action of releasing nitric oxide, were carried out in the same manner as in (2), and the survival rate exceeded 10% (isosorbide nitrate was 24%). %, Nicorandil 27%, nitroglycerin 16%, nipradilol 29%), and the survival rate increased compared with the case where these drugs were not added (1).

2. Normal mouse test 1
(1) A normal mouse (n = 51) is irradiated with 6.5 Gy at a dose rate of 0.5 Gy / min using an X-ray generator (MBR-1520A-3, manufactured by Hitachi Medical Corporation) ( When 0.5 mm aluminum / 0.3 mm copper filter was used), the survival rate was about 30 to 40% (see FIG. 1).
(2) X-rays were irradiated to a normal mouse (n = 51) using an X-ray generator (MBR-1520A-3, manufactured by Hitachi Medical Corporation) at a dose rate of 0.5 Gy / min (6.5 Gy) 0.5 mm aluminum / 0.3 mm copper filter was used). Next, sodium nitroprusside was intraperitoneally administered to the X-irradiated mice so that the concentration immediately after administration in vivo was 30 μM (first dose, 5 minutes after X-ray irradiation, dose: 2.8). ~ 3.2 mg / kg). Further, sodium nitroprusside was intraperitoneally administered in the same manner as the first administration after 1 day, 2 days and 7 days after X-ray irradiation. As a result, the survival rate was about 80%, and the survival rate was significantly improved as compared to (1) in which sodium nitroprusside was not administered (see FIG. 1).
(3) In (1) and (2), when blood was collected on the 14th day after X-ray irradiation and a blood test was performed (n = 10), the leukocyte of a mouse to which sodium nitroprusside was administered intraperitoneally, The values of erythrocytes and platelets increased compared to the case where administration was not performed, and an effect of improving hematopoietic / immune system functions was observed (see FIG. 2).

Test 2
(1) Using normal X-ray generators (MBR-1520A-3, manufactured by Hitachi Medical Corporation), X-rays were irradiated with 6.5 Gy at a dose rate of 0.5 Gy / min (0.5 mm aluminum / 0.3 mm copper filter was used).
(2) Next, sodium nitroprusside was intraperitoneally administered to the mouse irradiated with X-ray in (1) so that the concentration immediately after administration in vivo was 30 μM (total administration once, n = 3, X-ray 5 minutes after irradiation, dosage: 2.8-3.2 mg / kg, see FIG.
(3) In the same manner as (2), isosorbide nitrate, nicorandil, nitroglycerin or nipradilol was intraperitoneally administered to the mice irradiated with X-rays of (1) (n = 3 for each drug, see FIG. 3) .

Test 3
In Test 3, sodium nitroprusside, isosorbide nitrate, nicorandil, nitroglycerin and nipradilol were used as drugs. Then, in the same manner as in test 2, normal mice were irradiated with X-rays, and drugs were administered to the mice irradiated with X-rays (first administration), and the drugs were administered in the same manner as the first administration one day after X-ray irradiation. (2 doses in total, n = 3 for each drug, see FIG. 4).

Test 4
In Test 4, sodium nitroprusside was used as a drug. Then, in the same manner as in Test 2, normal mice were irradiated with X-rays, and drugs were administered to the mice irradiated with X-rays (first administration). The first administration was performed 1 day and 2 days after X-ray irradiation. The drug was administered in the same manner (total 3 times, n = 3, see FIG. 5).

Test 5
In Test 5, sodium nitroprusside, isosorbide nitrate, nicorandil, nitroglycerin and nipradilol were used as drugs. Then, in the same manner as in Test 2, normal mice were irradiated with X-rays, and drugs were administered to the X-irradiated mice (first dose). The first time after 1 day, 2 days and 7 days after X-ray irradiation. The drug was administered in the same manner as the administration (4 times in total, n = 3 for each drug, see FIG. 6).

From Tests 2 to 5, sodium nitroprusside had a high life-prolonging effect among the drugs used (see FIG. 3). In addition to the administration immediately after X-ray irradiation, the life-prolonging effect was greatly improved by further administration (see FIGS. 3 to 6). In particular, when 1 day and 2 days after X-ray irradiation were administered every day, and after 7 days after X-ray irradiation, 2 out of 3 mice survived 30 days after X-ray irradiation. (See FIG. 6).
On the other hand, isosorbide nitrate, nicorandil, nitroglycerin and nipradilol were found to have a life-prolonging effect, but a high life-prolonging effect like sodium nitroprusside was not obtained (see FIG. 3). Moreover, the improvement of the life extension effect by additional administration like sodium nitroprusside was not recognized (refer FIG. 4 and FIG. 6).

  Based on the above, sodium nitroprusside had low toxicity, and the effects of improving hematopoietic / immune system function, prolonging life and improving survival rate were obtained at low doses. Moreover, compared with sodium nitroprusside, which had the action of releasing nitric oxide as well as isosorbide nitrate, nicorandil, nitroglycerin and nipradilol, it had excellent radiation damage protection ability.

  The radiation injury protective agent of the present invention has an effect of improving hematopoietic / immune system function, prolonging life, and improving survival rate, and is useful for the prevention and treatment of radiation exposure and damage associated with radiation therapy.

Claims (9)

A therapeutic agent for radiation damage containing nitroprusside or a pharmacologically acceptable salt thereof (except when used in combination with serotonin, not a therapeutic agent for ultraviolet B wave damage). The therapeutic agent for radiation injury according to claim 1, wherein the administration is started within 60 minutes after the radiation exposure. The therapeutic agent for radiation injury according to claim 2, wherein the therapeutic agent is further administered one or more times after one day after radiation exposure and within 10 days. The therapeutic agent for radiation injury according to claim 3, which is additionally administered on the next day after radiation exposure. The therapeutic agent for radiation injury according to claim 4, which is further administered 2 days after the radiation exposure. The therapeutic agent for radiation injury according to claim 4 or 5, further administered 6 to 10 days after radiation exposure. The therapeutic agent for radiation injury according to any one of claims 1 to 6, which is administered at a dose of 0.9 to 3.6 mg / kg as nitroprusside per administration. The therapeutic agent for radiation injury according to any one of claims 1 to 7, wherein the radiation injury is acute radiation injury. The radiation disorder therapeutic agent according to claim 8, wherein the acute radiation disorder is a hematopoietic / immune system disorder.

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