JP2015078163A - Oral administration-type oxidation stress disease treatment agent comprising high-molecular cyclic nitroxide radical compound as active ingredient - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a pharmaceutical formulation which can effectively treat arterial sclerosis.SOLUTION: The present invention provides a formulation for treatment of cardiovascular disease mediated by oxidation stress comprising a high-molecular cyclic nitroxide radical compound showing no responsivity to pH as an active ingredient for oral administration.

Description

  In recent years, excessive reactive oxygen species (ROS) are produced in vivo due to irregular lifestyle, aging, social stress, etc., and various chronic diseases such as arteriosclerosis and diabetes, Alzheimer's and Parkinson's disease, etc. Giving intractable diseases has become clear. Although studies have been made on the prevention and treatment of chronic diseases using small molecule antioxidants, high concentrations of small molecule antioxidants spread throughout the body after administration and have side effects. In particular, it has entered the cell and its mitochondria and has a strong toxicity because it inhibits redox reactions essential for living bodies such as electron transport systems. In addition, since renal excretion and metabolism are fast, there is a problem that it is difficult to increase the effective concentration in vivo.

  Such a problem can be solved, for example, by a polymerized cyclic nitroxide radical compound formed by covalently bonding a cyclic nitroxide radical, which can be said to be a low molecular weight antioxidant, with a hydrophobic region of an amphiphilic polymer (Patent Document 1). reference). In addition, the polymerized cyclic nitroxide radical compound occupies the core region of the cyclic nitroxide radical-binding segment by self-assembly in an aqueous medium (water, buffered aqueous solution, physiological saline, water-mixed organic solvent, etc.) On the other hand, so-called polymer micelle-like nanoparticles in which the hydrophilic segment occupies the shell region can be formed and solubilized or homogeneously dispersed in an aqueous medium. In Patent Document 1, it is assumed that these properties are used exclusively to prepare injections (intravenous and intraarterial injections) and local administration agents (including brain implants).

  On the other hand, nanoparticles prepared from a polymer nitroxide radical compound such as polyethylene glycol-b-poly (cyclic nitroxide radical covalently bonded styrene) were administered to ulcerative colitis model mice induced with dextran sulfate (DDS). It has been published that diarrhea, diarrhea, weight loss score caused by DDS, and ulcerative colitis disorder are significantly reduced and more strongly suppressed than low molecular weight cyclic nitroxide radical compounds ( Patent Document 2). Further, in Patent Publication 2, in the case of nanoparticles derived from those in which the covalent bond is via an imino bond (—NH—) and those via an ether bond (—O—), the former (also referred to as N-RNP) is used. In comparison, the latter (also referred to as O-RNP) is described to have a higher effect on ulcerative colitis. This is because the latter nanoparticles are more resistant to disintegration or dissociation than the former nanoparticles due to a decrease in pH in the digestive tract due to oral administration, so that they stay in the digestive tract without being taken into the blood. It is understood that.

  Conversely, when N-RNP is administered orally, the nanoparticles disintegrate under acidic pH conditions, and the polymerized cyclic nitroxide radical compound can migrate into the bloodstream and can remain in the bloodstream for a long time. Thus, it has been published that it can be delivered to cancer, inflammatory sites, and oxidative stress disorder sites to treat arthritis, Alzheimer, and arteriosclerosis (Patent Document 3).

WO 2009/133647 JP2012-111700A JP2013-159560A

Patent Document 1 describes that treatment (specifically, prevention or treatment) of cerebral infarction, myocardial infarction, and the like can be performed by intravenous administration of N-RNP. However, such administration results in appreciable blood pressure fluctuations, albeit significantly lower than the low molecular weight cyclic nitroxide radical. On the other hand, according to Patent Document 3, O-RNP (or RNP-O) that does not decay depending on pH is not taken into the blood at all, but N-RNP that decays depending on pH is effectively blood. It is described that it is taken in and exhibits the intended drug efficacy, and that it is not toxic when administered orally to arteriosclerosis model mice. However, it would be desirable to provide formulations that can be administered orally more safely if possible.

  When the present inventors orally administer O-RNP, which is considered not to be taken into blood at all by oral administration and is localized in the gastrointestinal tract, to an arteriosclerosis model mouse, surprisingly, circulation such as arteriosclerosis occurs. It was found to be effective for chronic diseases of the organ system. As mentioned above, if O-RNP is not taken up into the blood or is very difficult to take up, it could be used more safely than N-RNP.

  Accordingly, the present invention provides a preparation for the prevention or treatment of diseases mediated by oxidative stress, which comprises as an active ingredient the oral administration of a polymerized cyclic nitroxide radical compound represented by the following formula I.

In which A represents unsubstituted or substituted C ₁ -C ₁₂ alkyl, and when substituted, the substituent represents a formyl group or a group of the formula R ¹ R ² CH—, wherein R ¹ and R ² is _{independently,} C 1 -C ₄ alkoxy or ^{R 1} and ^{R 2} are -OCH ₂ CH ₂ O together _{-, - O (CH 2)} 3 O- or -O _{(CH 2)} 4 O -
L ₁ represents a bond or a linking group,
L ₂ represents —C _1-6 alkylene-O— (C _1-6 alkylene) _q , q is an integer of 0 or 1,
X is at least 50% of the total number of X is 2,2,6,6-tetramethylpiperidin-1-oxyl-4-yl, 2,2,5,5-tetramethylpyrrolidine-1-oxyl- 3-yl, 2,2,5,5-tetramethylpyrrolin-1-oxyl-3-yl and 2,4,4-trimethyl-1,3-oxazolidine-3-oxyl-2-yl, 2,4, A residue of a cyclic nitroxide radical compound selected from the group consisting of 4-trimethyl-1,3-thiazolidin-3-oxyl-2-yl and 2,4,4-trimethyl-imidazolindin-3-oxyl-2-yl; And when X is other than the above residues, -L ₃ -X is a methyl, methyl halide or hydroxymethyl group,
m represents an integer of 15 to 10,000, and n represents an integer of 3 to 3,000.

In a preferred embodiment, the linking group of L _{1 in} the above formula is (CH ₂ ) _c S, (CH ₂ ) _c S (CH ₂ ) _d , (CH ₂ ) _c NH, — (CH ₂ ) _c NH _{(CH 2) d, (CH} 2) c NHCO, (CH 2) c NHCO (CH 2) d, (CH 2) c NHC (= S) NH, (CH 2) c NHC (= S) NH (CH _{2) d, (CH 2)} c NHC (= O) NH, (CH 2) c NHC (= O) NH (CH 2) d, (CH 2) c OCO, (CH 2) c OCO (CH 2) _{d, (CH 2) c O} , (CH 2) c O (CH 2) d, is selected from the group consisting of _{(CH 2) c,} where c and d are an integer of 1 to 5 independently.

  In another preferred embodiment, X in the above formula is represented by the following formula:

And R ′ represents a methyl group.

  In another preferred embodiment, the disease mediated by oxidative stress is arteriosclerosis.

In addition to the above, the present inventors recently succeeded in normalizing enterobacteria by oral administration of RNP ^O and reported (Journal of Gastroenterology, in press). On the other hand, in recent years, the correlation between intestinal bacteria and systemic diseases has been greatly closed (Valentina Tremaroli, Fredrik Baucked, “Functional interactions between the gutt microbiota and host13.Nat. Bungei Shunju, May issue “Intestinal bacteria determine lifespan”, Tetsuichiro Muto, Yoshimi Kanno, Shuntaro Torigoe).

  Conventionally, although treatment and prevention of oxidative stress diseases using low molecular weight antioxidants have been carried out, most of them are absorbed into the body and exert their medicinal effects, but at the same time they have side effects. Although not limited by theory, in the present invention, a novel pharmaceutical that is localized in the gastrointestinal tract, particularly the intestinal mucosa, and can treat chronic cardiovascular diseases by controlling intestinal bacteria. A new orally administered preparation that suppresses side effects and is safe and secure is provided.

Conceptual diagram of polymerized cyclic nitroxide nanoparticles radical compound forms used (O-RNP or RNP ^O) and Comparative nanoparticles (N-RNP or RNP ^N) in the present invention. The graph which shows the ESR signal intensity | strength in blood after oral administration of O-RNP and N-RNP. FIG. 5 shows the results obtained by administering O-RNP and N-RNP to a model animal at a concentration of 1200 mg / kg, respectively, collecting blood at each indicated time, and measuring the electron spin resonance (ESR) spectrum. The graph which shows the pharmacokinetics after oral administration using O-RNP and N-RNP labeled with the radioisotope ^125I . This is a result of removing the stomach, small intestine, cecum, large intestine, liver and blood at regular intervals and measuring the radiation dose with a gamma counter. The graph showing the weight transition of the animal (WTD / RNP-O) group after Western diet load single (WTD group) to the LDLR-deficient mouse and RNP ^O in addition to the load. The graph showing the measurement result of the white fat weight (n = 9-10) in the WTD group and WTD / RNP-O group of FIG. The graph showing the measurement result of the weight of a liver (n = 9-10) instead of the white fat weight of FIG. The graph showing the arteriosclerosis treatment effect by oral administration of RNP ^O (photograph which changes to the arterial lesion of the full length of aorta) and the photograph which changes to the figure (SUDAN IV dyeing | staining figure of an aorta).

<Detailed Description of the Invention>
In order to facilitate the understanding of the present invention, the conceptual diagram shown in FIG. 1 and the data shown in FIG. 2 to FIG. 3 are taken using typical compounds and comparative compounds used in the present invention as examples. This will be described with reference to FIGS. We make nanoparticles that disintegrate in response to changes in pH in aqueous media (water, buffered water, saline, water-miscible organic solvent-containing aqueous solutions, etc.) and non-disintegrating nanoparticles When these two nanoparticles are orally administered, the pH-disintegrating nanoparticles (N-RNP) (comparative example) are effectively taken into the blood as shown in FIGS. 2 and 3, but do not disintegrate at the pH. Nanoparticles (O-RNP) as the active ingredient of the present invention are not substantially taken into the blood. Nevertheless, O-RNP used as an active ingredient in the present invention has an effect on circulatory system diseases mediated by oxidative stress as described later. As a typical example of such a circulatory system disease, the present invention will be described more specifically by taking arteriosclerosis as an example. Once it can be confirmed that O-RNP can prevent or treat arteriosclerosis by oral administration. It is as if it was confirmed that other cardiovascular diseases in which O-RNP or N-RNP was shown to be effective by intravenous infusion can be treated by oral administration of O-RNP in Patent Document 1 and the like I will.

  Accordingly, cardiovascular diseases mediated by oxidative stress referred to in the present invention include arteriosclerosis, cerebral infarction, myocardial infarction, angina pectoris, diabetes, cancer, aging and the like.

  In the context of the present invention, the term arteriosclerosis is used in the sense commonly used in the art, but generally refers to a condition caused by arteriosclerosis, where arteriosclerosis is thickened and hardened by the arteries. Means state. In the context of the present invention, arteriosclerosis and arteriosclerosis may be used interchangeably.

  In the context of the present invention, a prophylactic or therapeutic formulation broadly means a pharmaceutical formulation for use on animals, preferably mammals, more preferably humans.

  The polymerized cyclic nitroxide radical compound that can be more suitably used in the present invention is a compound of the formula I wherein X is independently at least 50%, preferably 70% or more, more preferably 85% of the total number of X. Or more, most preferably 95% or more or 100%, and m is 15 to 10,000, preferably 20 to 5,000, more preferably 20 to 1,000, most preferably 20. And an integer of ˜500 and n is independently 3 to 1,000, preferably 3 to 500, and more preferably 3 to 100.

The alkyl group in C ₁ -C ₁₂ alkyl or C ₁ -C ₄ alkoxy can be linear or branched and includes, but is not limited to, methyl, ethyl, propyl, isopropyl, butyl, isobutyl. The former can further be hexyl, decyl, octyl, undecyl. C _1-6 alkylene include, but are not limited to, methylene, 1,2-ethanediyl, 1,2-propanediyl, 1,3-propanediyl, 1,4-butanediyl, etc. corresponding _{C 1-} Mention may be made of _6- alkyl diyl groups.

  Such a polymer cyclic nitroxide radical compound is described in Patent Document 1 and known per se, and can be produced by the described method. Further, as described in Patent Document 1, such a polymeric cyclic nitroxide radical compound is self-assembled in an aqueous medium, and as described above, a shell region or a surface layer is formed with a hydrophilic polyethylene (PEG) segment. It is understood that a so-called core-shell type polymer micelle in which a cyclic nitroxide radical-bonding segment forms a core region, and an average diameter by DLS measurement generally forms a nanoparticle having 30 nm to 100 nm. Has been. The polymer micelle or polymer micelle particle referred to in the present invention means such a nanoparticle.

  In the preparation of the present invention, depending on the dosage form, the polymer cyclic nitroxide radical compound can exist in the form of a self-assembled polymer micelle as described above, for example, in an aqueous medium. Alternatively, the solid preparation may contain the compound in any state, but it is preferable to include a compound which once formed a polymer micelle in an aqueous medium and then dried by freeze-drying or the like.

  As long as the purpose of the present invention is met, the preparation can contain diluents, excipients, and additives that can be included in oral preparations well known in the art, but the lyophilized product itself. You can also. When the preparation for oral administration of the present invention is a solid, the polymer cyclic nitroxide radical compound is converted into sucrose, lactose, mannitol, cellulose, trehalose, maltitol, dextran, starch, agar, gelatin, casein, albumin, glyceride, etc. It can contain in the 1 or more types of combination chosen from. In addition, other inert diluents, lubricants such as magnesium stearate, parabens, sorbic acid, preservatives such as α-tocopherol, antioxidants such as cysteine, disintegrants, binders, buffers, Sweeteners and the like can also be included.

  Liquid preparations for oral administration include physiologically acceptable emulsions, syrups, elixirs, suspensions and solution preparations. These preparations may contain commonly used inert diluents such as water, especially sterile water, deionized water. Such an aqueous solution may contain the above-mentioned sugars and polyethylene glycol having a molecular weight of about 200 to 100,000.

The oral dose of the polymerized cyclic nitroxide radical compound varies depending on the age, severity, responsiveness, etc. of the patient to be treated, but in general, in vitro and in vivo ( In vivo) Based on EC ₅₀ values confirmed to be effective in animal models, a specialist or clinician can determine. Although not limited, in general, it can be 0.01 μg to 1000 mg, preferably 0.1 μg to 500 mg, more preferably 10 μg to 100 mg per kg body weight. The dosing schedule can also be determined with reference to the results of in vivo animal model studies.

<Specific examples for carrying out the invention>
Hereinafter, the present invention will be specifically described, but the present invention is not intended to be limited to these embodiments. For the sake of brevity, the polymerized cyclic nitroxide radical compound includes methoxy produced according to Production Example 3 of the above-mentioned Patent Document 1 (the disclosure of which is incorporated herein by reference). An example using nanoparticles prepared from PEG-b-PCMS-O-TEMPO according to Production Example 5 (hereinafter referred to as RNP 2 ^O ) will be mainly described.

Mn of methoxy PEG-b-PCMS-O-TEMPO = 9000. Among them, the molecular weight of PEG was 5000, and the molecular weight of PCMS-O-TEMPO was 4000. The average RNP ^O diameter was 40 nm.

Test Example 1: Examination of RNP ^O inhibitory effect on arteriosclerosis Preparation of Western diet-loaded LDLR (low density lipoprotein receptor) deficient mouse model and therapeutic effect LDLR deficient mice (8 males, 10 WTD groups, WTD / The effect of RNP ^O on arteriosclerosis using an arteriosclerosis model (WTD group) loaded with Western diet (34% sucrose, 21% fat, 0.15% cholesterol) on 9 RNP-O groups) It was investigated. Along with the above load, 3.125 mg / mL RNP 2 ^O was administered by drinking water with free drinking (WTD / RNP-O group). Twelve weeks after administration, changes in body weight, white fat weight, liver weight, and formation of arteriosclerotic lesions were evaluated. The respective results are shown in FIG. 4, FIG. 5, FIG. 6, and FIG.

From FIG. 4, the WTD / RNP-O group has no significant difference in weight transition compared to the WTD group, and from FIG. 5 and FIG. 6, the WTD / RNP-O group has white fat mass and There was no significant difference in liver weight, and no toxicity was observed in orally administered RNP ^O.

  From FIG. 7, it is recognized that the formation of arteriosclerotic lesions is suppressed by administration of RNPO in both the full length of the aorta and the section of the aortic origin.

Claims (5)

Formulation for the prevention or treatment of cardiovascular diseases mediated by oxidative stress, comprising as an active ingredient for the oral administration of a polymerized cyclic nitroxide radical compound represented by the following formula I:
In which A represents unsubstituted or substituted C ₁ -C ₁₂ alkyl, and when substituted, the substituent represents a formyl group or a group of the formula R ¹ R ² CH—, wherein R ¹ and R ² is _{independently,} C 1 -C ₄ alkoxy or ^{R 1} and ^{R 2} are -OCH ₂ CH ₂ O together _{-, - O (CH 2)} 3 O- or -O _{(CH 2)} 4 O -
L ₁ represents a bond or a linking group,
L ₂ represents —C _1-6 alkylene-O— (C _1-6 alkylene) _q , q is an integer of 0 or 1,
X is at least 50% of the total number of X is 2,2,6,6-tetramethylpiperidin-1-oxyl-4-yl, 2,2,5,5-tetramethylpyrrolidine-1-oxyl- 3-yl, 2,2,5,5-tetramethylpyrrolin-1-oxyl-3-yl and 2,4,4-trimethyl-1,3-oxazolidine-3-oxyl-2-yl, 2,4, A residue of a cyclic nitroxide radical compound selected from the group consisting of 4-trimethyl-1,3-thiazolidin-3-oxyl-2-yl and 2,4,4-trimethyl-imidazolindin-3-oxyl-2-yl; And when X is other than the above residues, -L ₃ -X is a methyl, methyl halide or hydroxymethyl group,
m represents an integer of 15 to 10,000, and n represents an integer of 3 to 3,000. The linking group of L ₁ is (CH ₂ ) _c S, (CH ₂ ) _c S (CH ₂ ) _d , (CH ₂ ) _c NH, — (CH ₂ ) _c NH (CH ₂ ) _d , (CH ₂ ) _c NHCO, (CH ₂ ) _c NHCO (CH ₂ ) _d , (CH ₂ ) _c NHC (═S) NH, (CH ₂ ) _c NHC (═S) NH (CH ₂ ) _d , (CH ₂ ) _c NHC ( ═O) NH, (CH ₂ ) _c NHC (═O) NH (CH ₂ ) _d , (CH ₂ ) _c OCO, (CH ₂ ) _c OCO (CH ₂ ) _d , (CH ₂ ) _c O, (CH _{2) c O (CH 2)} d, (CH 2) selected from the group consisting of _c, where c and d are integers of 1 to 5 independently, formulation according to claim 1. X is the following formula
The formulation according to claim 1 or 2, wherein R 'represents a methyl group.   The preparation according to any one of claims 1 to 3, wherein the disease mediated by oxidative stress is arteriosclerosis.   The preparation according to any one of claims 1 to 4, wherein the polymerized cyclic nitroxide radical compound represented by formula I is in the form of polymer micelle particles.