JP2001316389A - Pyrrolopyridinium derivative - Google Patents

Abstract

PROBLEM TO BE SOLVED: To obtain a new AGE, an antibody prepared by using the AGE as a hapten and to provide a method for examination for measuring the AGE and a method for evaluating pharmacodynamic effects of medicine effective for diabetes-related diseases, renal insufficiency, dialysis complication, aging, diseases following aging, Alzheimer's disease, etc. SOLUTION: This pyrrolopyridinium derivative is represented by general formula (I), formula (II) or formula (III) [R' and R" are the same or different and are each an amino group or an alkyl group which may contain a protective group-containing amino group and/or a carboxy group]. Consequently the pyrrolopyridinium derivative is not only a new AGE having a completely new structural skeleton different from that of a conventional AGE, but also is considered to be a cross-linked substance formed through a saccharification reaction and an oxidcition reaction from its characteristic structure, has a hemiacetal structure having reactivity in the molecule, is expected to have usefulness different from that of a conventional AGE and is extremely useful in the field requiring various AGEs.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

The present invention relates to a novel pyrrolopyridinium derivative, an antibody prepared using the derivative as a hapten, a test method for measuring the derivative, and diabetes-related diseases, renal failure, dialysis complications, aging, and aging. The present invention relates to a method for evaluating the efficacy of a drug effective for accompanying diseases, Alzheimer's disease, and the like.

[0002]

2. Description of the Related Art The Maillard reaction is an initial stage from the formation of a Schiff base between an amino group of a protein and an aldehyde group of a reducing sugar until stabilization due to Amadori rearrangement, and further dehydration / condensation / fragmentation / oxidation.・ Through a series of complex reactions such as cyclization, fluorescence, browning, intermolecular cross-link formation,
It can be distinguished into two stages, that is, the late stage of transition to the Maillard reaction late product (AGE), which has characteristics such as recognition by endothelial cell and macrophage receptors.

[0003] This reaction has been mainly studied in the field of food chemistry. In 1968, glycosyl hemoglobin (HbA1c), a small component of hemoglobin, was identified in vivo and found to increase in diabetic patients. That is, it has been found that the Maillard reaction also occurs in biological proteins, and this has triggered the biomedical significance of the Maillard reaction.

In recent years, this AGE produced especially at a late stage
Attention has been focused on the relationship between AGEs and various diseases, and studies on AGE have been promoted. Up to now, N ^epsilon - Carboxymethyl lysine (CML), pentosidine, pyrraline, N ^epsilon -
Several AGEs have been reported, such as carboxyethyl lysine (CEL), glyoxal lysine dimer (GOLD), and methylglyoxal lysine dimer (MOLD). The present inventors and their collaborators also
Pyridinium derivatives called ^TM (JP-A-6-730)
No. 57) and naphthyridinium derivatives as disclosed in JP-A-8-48686;
No. 5 discloses pyrrolonaphthyridinium derivatives and the like as novel AGEs.

As an antibody recognizing these AGEs, for example, 6D12 (Kumamoto Antibody Research Institute) which is an anti-CML monoclonal antibody is commercially available. Actually, using these antibodies, human plasma, human Serum, human urine, atherosclerotic tissue, kidney tissue of diabetic nephropathy, eye tissue of elderly, brain tissue of Alzheimer's disease, etc. Research is being conducted, and research on the significance of its existence is ongoing.

[0006] Regarding the relationship between AGE and various diseases, first, the fluorescence, which is a characteristic change of AGE, is used as an index.
The correlation between E and various diseases has been investigated, and further studies have been made using antibodies against the above-mentioned AGE, and the involvement of AGE in various diseases has been elucidated. For example, a review by Horiuchi et al., One of the present inventors, "AGE (Glycation) and Disease" (BIO Clinica 11 (5), p.314 (199)
See 6)).

[0007] For example, aging is considered to be a process in which various proteins and DNAs in tissues are modified or changed and cannot be decomposed / removed and accumulate to cause dysfunction. It is listed as one of the causes. In fact, for example, the fluorescence of acid hydrolyzate of skin collagen and lens protein increases with age, indicating that excessive AGE formation of lens protein is involved in the development of cataract. In addition, inhibition of enzyme activity and degradation of intracellular protein by AGE formation, impaired substance transport due to AGE-formed cytoskeletal protein, DNA mutation caused by cross-linking between protein and DNA by AGE, and the like have been observed. It has been reported that anti-AGE antibodies react with the water-soluble and alkali-soluble fractions of normal human lens crystallin in a concentration-dependent manner, and that there is a very high correlation between the reactivity and the age. At present, AGEs have been reported to have increased AGE formation in the Descemet membrane by aging, and reports that AGE immune response of pyramidal neurons was correlated with age by immunohistochemical search using antibodies. It is being recognized as a factor in the aging phenomenon.

[0008] It has been revealed that the fluorescence of diabetic patients is significantly higher than that of healthy subjects. Furthermore, AGE of diabetic patients has been observed in renal mesangial regions, blood vessel walls, renal tissues such as tubules and coronary atherosclerotic lesions. Accumulation has been reported, and immunostaining using anti-AGE antibodies has shown that AGEs show almost the same distribution in both experimental diabetic rats and human diabetic peripheral nerves. It recognized.

[0009] It has also been clarified that excessive formation of AGE in a living body is a cause of diabetic complications. For example, the lens protein crystallin AG
E formation is considered to be a cause of diabetic cataract. AGEs have also been shown to be deeply involved in diabetic nephropathy.
Hyperglycemia converts the glomerular basement membrane collagen of the kidney into AGEs, which easily binds to albumin, immunoglobulins, low-density lipoproteins, etc. It has also been pointed out that AGEs accumulate in mesangial cells and may be involved in inflammation such as receptor-mediated cytokine secretion. Recently, it has been reported that an AGE production inhibitor is useful for diabetic nephropathy. Further, it has been reported that nerve myelin protein is converted to AGE, suggesting that AGE is also involved in diabetic neuropathy. Thus, the correlation between the occurrence of diabetic complications such as diabetic nephropathy, arteriosclerosis, neurosis, retinopathy, cataract and the like and AGE has been clarified. A
Regarding GE and diabetic complications, see, for example, a review by Morisaki et al., “AGE and diabetic complications” (Latest Medicine, 49, 2, p.2
48 (1994)).

[0010] In addition, an increase in fluorescence in serum proteins of dialysis patients and the accumulation of AGE at carpal tunnel amyloid deposition sites in dialysis patients indicate that AGE is associated with dialysis-related amyloidosis. Have been.

[0011] Furthermore, monocyte, macrophage, mesangial cells and endothelial cells have scavenger receptors which recognize AGE proteins.
There are also reports suggesting that GE recognition causes cytokine release, increased vascular permeability, abnormal blood flow, etc., suggesting a link between AGE and pathologies such as inflammation, capillary occlusion, and arteriosclerosis. In addition, since AGEs are strongly detected in senile plaques, amyloid and neurofibrillary tangles, which are characteristically found in the brain of Alzheimer's disease patients, it is considered that they are deeply involved in the onset and progression of Alzheimer's disease. I have.

In addition, free radicals (active oxygen) have recently attracted attention as a cause of tissue damage such as aging and diabetes and the formation of lesions of Alzheimer's disease. The Amadori rearrangement product generated by the initial reaction of the Maillard reaction is oxidized by active oxygen to form AGE, but it is becoming clear that it is also a source of active oxygen. It has been reported in a neuronal tumor cell line (SK-N-SH) that the neurofibrillary tangle (PHF) tau protein also generates active oxygen by glycation and activates the transcription factor NF-κB. In addition, it has been shown that active oxygen-scavenging enzymes such as Cu-Zn-superoxide dismutase (Cu-Zn-SOD) are converted into AGEs to decrease their activity and cause an increase in oxidative stress. From these points, it is suggested that AGE is related to aging, diabetes, and Alzheimer's disease.

However, in view of the above-described AGE generation process, it is presumed that AGEs having various structures are generated in a living body depending on each situation, and that the distribution of these AGEs in the tissue is also different. Has been guessed. In order to find an optimum AGE for a variety of situations in a living body, it has been desired by those skilled in the art to have a large number of AGEs having different properties. Accordingly, there has been a demand for an AGE that is generated under a condition different from that of the conventional AGE, that is, a new AGE having a structural skeleton different from that of the conventional AGE.

[0014]

SUMMARY OF THE INVENTION An object of the present invention is to provide a novel AGE having a structural skeleton different from that of conventional AGE, an antibody prepared using the AGE as a hapten, a test method for measuring the AGE, and a diabetes-related disease. The present invention provides a method for evaluating the efficacy of a drug effective for renal failure, dialysis complications, aging, diseases associated with aging, Alzheimer's disease, and the like.

[0015]

Means for Solving the Problems The present inventors use the non-enzymatic saccharification model reactants of proteins to evaluate the saccharification reaction and oxidation reaction as an index for diagnosing the onset / progress of diabetic complications, dialysis complications or aging. As a result of research on the crosslinked substance formed through the both, a novel pyrrolopyridinium derivative having a completely new structural skeleton different from the conventional AGE was found, and the present invention was completed.

[0016]

BEST MODE FOR CARRYING OUT THE INVENTION The pyrrolopyridinium derivative of the present invention is a compound represented by the following general formula (I), (II) or (III).

Embedded image

Embedded image

Embedded image [In the formula, R ′ and R ″ are the same or different and each represents an amino group, an amino group having a protecting group, and / or an alkyl group optionally having a carboxyl group.]

The alkyl group in R 'and R "of the above formula (I), (II) or (III) is preferably methyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl, t-butyl. , Pentyl, isopentyl, neopentyl, t-pentyl, hexyl,
Examples thereof include a linear or branched alkyl group having 1 to 6 carbon atoms such as dimethylbutyl.

The alkyl group may have an amino group, an amino group having a protecting group and / or a carboxyl group, and the protecting group for the amino group is commonly used in the field of peptide synthesis chemistry and the like. Protecting groups are available and include, for example, acetyl, benzyloxycarbonyl, p-methoxybenzyloxycarbonyl, p-chlorobenzyloxycarbonyl, p-nitrobenzyloxycarbonyl,
p-phenylazobenzyloxycarbonyl, p-methoxyphenylazobenzyloxycarbonyl, t-butoxycarbonyl (Boc), p-toluenesulfonyl (Tos), tertiary amyloxycarbonyl, p-biphenylisopropyloxycarbonyl, diisopropylmethyloxycarbo And groups such as nyl and formyl.

Among the substances of the present invention represented by the general formulas (I), (II) and (III), the pyrrolopyridinium derivatives wherein R 'and R "are an alkyl group having an amino group and / or a carboxyl group are: It can be easily bound as a hapten to a carrier protein, etc., and is particularly useful when preparing an antibody. Carriers to be bound to a hapten for preparing an antibody include proteins such as serum albumin and limpet blood chromoprotein. Commonly used carriers such as polymers such as polylysine can be used.

The pyrrolopyridinium derivative of the present invention includes salts represented by the above general formulas (I), (II) and (III), and includes, for example, hydrochloric acid, sulfuric acid, nitric acid, hydrobromic acid, phosphoric acid, Perchloric acid, thiocyanic acid, boric acid, formic acid, acetic acid, haloacetic acid, propionic acid, glycolic acid, citric acid, tartaric acid, succinic acid, gluconic acid, lactic acid, malonic acid, fumaric acid, anthranilic acid, benzoic acid, cinnamon Acid, p-toluenesulfonic acid, naphthalenesulfonic acid, addition salt with acid with an acid such as sulfanilic acid, addition salt with an organic base such as ammonia or organic amine, or alkali metal such as sodium or potassium, calcium, magnesium, barium And salts with metals such as alkaline earth metals or aluminum and zinc. These salts can be produced from the free pyrrolopyridinium derivatives of the present invention by known methods, or can be mutually converted.

The present invention also provides a compound of the present invention in which stereoisomers such as cis-trans, optical isomer and conformer exist, or when present in the form of hydrate or complex compound. Any of the stereoisomers, hydrates,
It also includes complex compounds.

Next, an example of the method for producing the compound of the present invention will be described. A compound in which R ′ is NH ₂ or R ″ is NH ₂ (amine components: R ′ and R ″ each represent the same group as described above) is exemplified by a compound such as glucose, fructose, galactose, mannose and deoxyglucose. The compound of the present invention can be obtained by coexisting with saccharides such as carbohydrates, amino sugars such as glucosamine and galactosamine, and oligosaccharides such as saccharose. Further, the compound of the present invention can also be obtained by carrying out a mixed reaction using a protein, a peptide or the like as an amine component, followed by an acid hydrolysis treatment.

The reaction conditions such as reaction temperature, reaction time, pH and the like are not particularly set, and can be appropriately set. The simplest thing to do is leave it at room temperature,
The reaction can be promoted by heating or the like. The obtained compound of the present invention can be purified by ordinary means such as distillation, chromatography, recrystallization and the like.

The pyrrolopyridinium derivative of the present invention can be used for the production of bovine serum albumin according to the method described in, for example, CLIN. CHEM. 40/9, 1766-1773 (1994), which is a report on an antibody against pentosidine, one of the AGEs. (BSA)
Polyclonal antibody or monoclonal antibody of the pyrrolopyridinium derivative of the present invention can be obtained by a commonly used method such as immunizing mice, rabbits and the like by polymerizing with limpet blood chromoprotein (KLH) or the like.

Using the thus obtained antibody, the pyrrolopyridinium derivative of the present invention can be measured by a known method such as ELISA or immunostaining.
Further, the pyrrolopyridinium derivative of the present invention can also be measured directly by HPLC or the like, for example, using its fluorescence wavelength.

Using the amount of the pyrrolopyridinium derivative of the present invention thus measured as an index, the degree of accumulation of the late product of the Maillard reaction in the living body was evaluated, and diabetes, diabetic complications, dialysis-related complications, amyloidosis, aging Alternatively, the degree of progress of a disease associated with aging can be evaluated.
Also, by measuring the amount of the pyrrolopyridinium derivative of the present invention in the presence or absence of the test drug,
The efficacy of a test drug for diabetes, diabetic complications, dialysis-related complications, amyloidosis, aging or diseases associated with aging can also be evaluated.

The object to be measured includes living tissue, living tissue extract, body fluid and / or urine, and preferably human skin tissue, human skin tissue extract, human lens extract, human atherosclerotic tissue, diabetic kidney Kidney tissue of a patient with the disease, tissue of the elderly, brain tissue of an Alzheimer's disease patient, human blood, human plasma, human serum, human urine, etc., and particularly preferably human skin tissue, human skin tissue extract, and human Examples include plasma, human serum, and human urine.

Hereinafter, preferred embodiments of the present invention will be described. (1) Pyrrolopyridinium derivatives represented by the general formula (I), (II) or (III), stereoisomers thereof, and salts thereof. (2) The pyrrolopyridinium derivative according to the above (1) represented by the general formula (I), (II) or (III) and a salt thereof. (3) The pyrrolopyridinium derivative according to any one of the above (1) and (2) represented by the general formula (I) and a salt thereof. (4) The pyrrolopyridinium derivative according to any one of the above (1) and (2) represented by the general formula (II) and a salt thereof. (5) The pyrrolopyridinium derivative according to any one of the above (1) and (2) represented by the general formula (III) and a salt thereof. (6) An antibody prepared using the pyrrolopyridinium derivative according to any one of (1) to (5) as a hapten. (7) An inspection method for measuring the pyrrolopyridinium derivative according to any one of (1) to (5). (8) The test method according to (7), wherein the pyrrolopyridinium derivative is measured in a living tissue, a living tissue extract, a body fluid, and / or urine. (9) The test method according to any one of the above (7) or (8), wherein the pyrrolopyridinium derivative is measured in a body fluid and / or urine. (10) The test method according to (9), wherein the body fluid and / or urine is any of human blood, human plasma, human serum, and human urine. (11) The test method according to any one of the above (7) or (8), wherein a pyrrolopyridinium derivative of a biological tissue and / or a biological tissue extract is measured. (12) The above (1), wherein the living tissue and / or the living tissue extract is either human skin tissue or human skin tissue extract.
1) The inspection method described. (13) The test method according to any one of (7) to (12), wherein the antibody according to (6) is used. (14) The test method according to any one of the above (7) to (13), wherein a pyrrolopyridinium derivative is measured in order to evaluate a production amount and / or an accumulation amount of a Maillard reaction late product in a living body. (15) A drug for treating diabetes using the pyrrolopyridinium derivative according to any one of (1) to (5) as an indicator,
A method for evaluating the efficacy of a drug for treating diabetes complications, a drug for treating dialysis-related complications, a drug for treating amyloidosis, an antiaging drug, or a drug for treating diseases associated with aging. (16) A therapeutic drug for diabetes using the antibody according to (6) above,
A method for evaluating the efficacy of a drug for treating diabetes complications, a drug for treating dialysis-related complications, a drug for treating amyloidosis, an antiaging drug, or a drug for treating diseases associated with aging. (17) A drug for treating diabetes using the pyrrolopyridinium derivative according to any one of (1) to (5) as an index,
A therapeutic agent for diabetes complications, a therapeutic agent for dialysis-related complications, a therapeutic agent for amyloidosis, an antiaging drug, or a method for diagnosing an aging-related disease. (18) A therapeutic drug for diabetes using the antibody according to (6) above,
A therapeutic agent for diabetes complications, a therapeutic agent for dialysis-related complications, a therapeutic agent for amyloidosis, an antiaging drug, or a method for diagnosing an aging-related disease.

[0029]

EXAMPLES Examples of the pyrrolopyridinium derivative of the present invention will be shown below, but the present invention is not limited thereto.

Embodiment 1 79.2g of glucose (400mM) and γ
-45.3 g (400 mM) of aminobutyric acid in 250 mM phosphate buffer (pH
7.3) Dissolved in 1.1 L and allowed to stand at 37 ° C for 60 days. Reaction liquid 33
Without concentrating 0 mL, Amb was made acidic (around pH 2) with trifluoroacetic acid (TFA) and equilibrated with ion-exchanged water.
It was added to an erlite XAD-2 column (organo: φ57 mm × 750 mm). This was further eluted with ion-exchanged water and fractionated 100 mL at a time. The obtained fractions were sequentially analyzed by HPLC chromatography, and fractions containing Compound 1 were collected and concentrated to dryness. Next, Develosil ODSLOP equilibrated with 0.2% TFA
-45S (Nomura Chemical), 7% acetonitrile / 0.2% TF
Eluted with A. The eluate is 15 mL in the fraction collector
Each fraction was collected by reverse phase HPLC (column: Ineartsil ODS-
3V; φ4.6mm x 250mm, solvent: 20% methanol / 0.2% TFA,
(Flow rate: 0.8 mL / min). The fraction containing Compound 1 was collected and concentrated to dryness. The fraction was further subjected to Develosil OD.
Purification with S LOP-45S (5% acetonitrile / 0.2% TFA) was repeated. Reversed phase HPLC (column: Ineartsil ODS-3V; φ
4.6mm x 250mm, solvent: 20% methanol / 0.2% TFA, flow rate:
Based on the analysis results at 0.8 mL / min), three fractions were collected for each component, and two fractions were subsequently subjected to purification (Fr
-1: 340mg, Fr-2: 400mg). Fr-1 was subjected to reverse phase HPLC (column:
Inertsil ODS-3; φ20mm x 250mm, flow rate: 10.0 mL / mi
n, eluent: 8% acetonitrile / 0.2% TFA), 280 nm
The main component, Compound 1 (8- (1,2-Dihyd
roxyethyl) -6,9-dihydroxy-2,5-bis (3-carboxypropyl)-
2,6,8,9-tetrahydro-7-oxa-2-aza-5-azoniabenzo [cd] az
ulene). Concentrate this under reduced pressure, about 5 mL
Sep-Pak Va equilibrated with ion exchanged water
c Added to C18 Cartridge, washed thoroughly with ion-exchanged water to remove trifluoroacetic acid, and eluted with 50% acetonitrile. This was concentrated to dryness to obtain Compound 1 (95 mg). Fr-2 was subjected to reverse phase HPLC (column: Inertsil ODS-3; φ2
0 mm x 250 mm, flow rate: 8.0 mL / min, eluent: 12% acetonitrile / 0.2% TFA) to obtain a fraction containing a main component while detecting UV absorption at 280 nm. Concentrate this under reduced pressure, about 5 mL
Sep-Pak Va equilibrated with ion exchanged water
c Added to C18 Cartridge, washed thoroughly with ion-exchanged water to remove trifluoroacetic acid, and eluted with 50% acetonitrile. This was concentrated to dryness to give compound 2 (3- (3,4
-Dihydroxytetrahydrofuran-2-yl) -1,5-bis (3-carboxyp
ropyl) -1H-pyrrolo [3,2-c] pyridinium, 72 mg).

Example 2 Compound 1 (19.6 mg) was added to 1 mL of 2N H 2
It was dissolved in 3 mL of Cl and stirred at room temperature. The reaction was performed over time by reversed-phase HPLC (column: Inertsil ODS-3V; φ 4.6 mm x 250 mm,
Flow rate: 0.8 mL / min, Eluent: 12% acetonitrile / 0.2% T
FA). After 12 hours, the reaction solution was concentrated to dryness under reduced pressure, and the residue was subjected to reverse phase HPLC (column: Inertsil ODS-3; φ
Purification with 10mmx250mm, flow rate: 4.0 mL / min, eluent: 12% acetonitrile / 0.2% TFA) to obtain the raw material and product fractions, each of which was concentrated under reduced pressure and ion exchanged when it became about 5 mL Sep-Pak Vac C18 Cartridg equilibrated with water
e, washed thoroughly with ion-exchanged water to remove trifluoroacetic acid, and eluted with 50% acetonitrile.
Concentrate to dryness and concentrate the starting material and compound 3 (8- (1,2-Dihydroxyet
hyl) -1,5-bis (3-carboxypropyl) -2,6,8,9-tetrahydro-7
-oxa-2-aza-5-azoniabenzo [cd] cyclopenta [h] azulene-
6,8-diol, 6.9 mg) was obtained.

Embodiment 3 FIG. 27.0 g (200 mM) of glucose and N
28.2 g (200 mM) of ^α -acetyl lysine was dissolved in 750 mL of a 250 mM phosphate buffer (pH 7.3) and allowed to stand at 37 ° C. for 56 days.
The reaction solution was acidified (about pH 2) with trifluoroacetic acid without concentration, and added to a sulfonic acid type ion exchange resin (Diaion PK-216: Mitsubishi Kasei). The column was washed with water and eluted with 2N aqueous ammonia. This fraction was concentrated to dryness and further developed with Develosil ODS LOP-45 equilibrated with 0.2% TFA.
S (Nomura Chemical), 10% methanol / 0.2% TFA, 20%
The methanol content was sequentially increased with methanol / 0.2% TFA, and the fraction eluted with 20% methanol / 0.2% TFA was further subjected to STR.
Purified by ODS-II column (Shimadzu Techno Research)
Compound 4 and compound 5 were obtained.

Embodiment 4 FIG. Using N ^α -t-butoxycarbonyl-L-lysine as an amine component, a mixed reaction with glucose was carried out in the same manner as described above, followed by separation and purification, followed by TFA
And left to stand for 30 minutes to remove the third butoxycarbonyl group, to give compound 6.

The physical properties of the obtained compound of the present invention are shown below. In the same manner as in the above example, 1-position or all carbons
^A labeled substance was produced using ¹³ C-labeled glucose and used for structural analysis. Fluorescence spectrum is 650-40
Ultraviolet absorption (UV) spectra were measured in a methanol by Fluorescence Spectrophotometer (Hitachi) and by DU-650 (Beckman). Sputtered ion mass spectroscopy (SIMS) was measured using M-80B (Hitachi) using glycerin as a matrix. Nuclear magnetic resonance (NMR) spectrum is in ARX-500 (Bruker) in heavy water
And the proton has a resonance frequency of 500.13 MHz
¹³ C sets the resonance frequency 125.77 MHz to 0.00.
ppm. ¹ H-NMR spectrum and ¹³ C-N
Assignment of MR spectrum is ¹ H- ¹ H COSY, HMQ
C, determined by two-dimensional NMR such as HMBC.

Compound 1 Fluorescence spectrum: Exmax = 325 nm, EMmax = 440 nm SIMS: m / z 439 NMR data: as shown in Table 1.

[Table 1]

Compound 2 Fluorescence spectrum: EXmax = 331 nm, EMmax = 466 nm SIMS: m / z 393 NMR data: shown in Table 2.

[Table 2]

Compound 3 Fluorescence spectrum: EXmax = 324 nm, EMmax = 453 nm SIMS: m / z 421 NMR data: shown in Table 3.

[Table 3]

Compound 4 Fluorescence spectrum: EXmax = 325 nm, EMmax = 440 nm SIMS: m / z 609 NMR data: shown in Table 4.

[Table 4]

Compound 5 Fluorescence spectrum: EXmax = 324 nm, EMmax = 453 nm SIMS: m / z 591 NMR data: The results are shown in Table 5.

[Table 5]

Embodiment 5 FIG. The pyrrolopyridinium derivative of the present invention was polymerized with a limpet blood chromoprotein, and immunized in a mouse to obtain a monoclonal antibody against the substance of the present invention by a conventional method. This antibody reacted significantly with various glycated proteins produced in vitro, but did not react with the unmodified product. Furthermore, the antibody also reacted with the human lens extract, and the reaction intensity increased with age.

[0041]

The pyrrolopyridinium derivative of the present invention is not only a novel AGE having a completely new structural skeleton different from the conventional AGE, but also a cross-link formed through both saccharification reaction and oxidation reaction in view of its characteristic structure. It is considered to be a substance and further has a hemiacetal structure having a reactivity in the molecule, and is very noteworthy as a new AGE. As mentioned earlier, diabetics,
It is clear from various research results that dialysis patients are placed under high oxygen stress compared to healthy people,
A crosslinked substance such as the substance of the present invention, which is formed not only through a saccharification reaction but also through an oxidation reaction, is very highly used as a new indicator of the onset / progress of diabetes complications, dialysis complications, or aging.

Therefore, using the substance of the present invention as an index, diabetic complications such as diabetes and diabetic nephropathy, diabetic arteriosclerosis, diabetic neuropathy, diabetic cataract, diabetic retinopathy, dialysis-related complications, Alzheimer's disease It is possible to examine diseases, aging and diseases associated therewith, and to evaluate the efficacy of the compounds of the present invention in in vitro and in vivo test systems using the substance of the present invention as an index. In addition, an antibody prepared using the substance of the present invention as a hapten can be used immunochemically and immunohistochemically in the above-mentioned tests and evaluation of drug efficacy, and is extremely useful. As described above, the pyrrolopyridinium derivative of the present invention is a novel substance having a new structural skeleton containing an intramolecular hemiacetal structure that is distinctly different from known AGEs. It is very useful in the field where activity is suggested and various AGEs are required as described above.

Continued on the front page (51) Int.Cl. ⁷ Identification symbol FI Theme coat II (reference) // C12P 21/08 C12P 21/08 C07M 7:00 C07M 7:00

Claims (7)

[Claims] 1. The following general formula (I), (II) or (III)
And a stereoisomer thereof and salts thereof. Embedded image Embedded image Embedded image [In the formula, R ′ and R ″ are the same or different and each represents an amino group, an amino group having a protecting group, and / or an alkyl group optionally having a carboxyl group.] 2. An antibody prepared using the pyrrolopyridinium derivative according to claim 1 as a hapten. 3. A test method for measuring the pyrrolopyridinium derivative according to claim 1. 4. A living tissue, a living tissue extract, a body fluid and / or
4. The method according to claim 3, wherein the pyrrolopyridinium derivative is measured in urine. 5. The test method according to claim 3, wherein the antibody according to claim 2 is used. 6. Efficacy of a therapeutic drug for diabetes, a therapeutic drug for diabetic complications, a therapeutic drug for dialysis-related complications, a therapeutic drug for amyloidosis, an anti-aging drug or a therapeutic drug for aging-related diseases, using the pyrrolopyridinium derivative according to claim 1 as an indicator. Evaluation method. 7. A method for evaluating the efficacy of a therapeutic agent for diabetes, a therapeutic agent for diabetic complications, a therapeutic agent for dialysis-related complications, a therapeutic agent for amyloidosis, an anti-aging drug or a therapeutic drug for aging-related diseases using the antibody according to claim 2.