JPH05310711A - Spin label compound - Google Patents

Abstract

PURPOSE:To provide a new spin label compound useful as a probe for getting an image of an organ in a living body by the electron spin resonance of the benzodiazepine receptor distribution in the brain. CONSTITUTION:The spin label compound of formula I [A is group of formula II to formula VII (R<1> is H, halogen, nitro, cyano or CF3; R<2> is H or halogen; R<3> is H or methyl; Me is methyl; Et is ethyl); B is group of formula VIII (X is single bond or methylene); n is 1 or 2], e.g. 7-chloro-1,3-dihydro-5- phenyl-1-[N-(2,2,6,6-tetramethylpiperidin-1-oxyl-4-yl)carbamoylmethyl] -2H-1,4 benzodiazepin-2-one. The compound of formula I can be produced e.g. by reacting a compound of the formula A-H with a compound of formula IX (Y is reactive ester residue of alcohol).

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a novel spin-label compound useful as a probe for obtaining a biological image of benzodiazepine receptor distribution in the brain by electron spin resonance (ESR) method.

[0002]

2. Description of the Related Art The ESR method is a powerful means for directly detecting paramagnetic species such as free radicals and transition metal ions, and is basically suitable for in vivo measurement of living bodies. However, the X-band (about 9.5 GHz) has been used as the microwave in the ESR device which has been generally used so far, and the dielectric loss due to water is large, and the measurement of a large-capacity biological sample is impossible. there were. Recently, L-band (390 ~
ES using low-frequency microwave called 1550 MHz)
An R instrument (hereinafter referred to as "L-band ESR instrument") has been developed to enable in vivo measurement of free radicals in a large-volume sample containing a large amount of water, particularly a biological sample [eg Pharmacia, 27 , 710 to 715 ( 1991)].

By the way, when human diseases develop, various abnormalities occur in local physiological and biochemical processes, which progress to cause impairment of higher-order functions and various symptoms.
Symptoms develop. In the brain, intracellular signaling functions are expressed by binding neurotransmitters such as dopamine, serotonin, γ-aminobutyric acid (GABA), and acetylcholine to their receptors, but when receptor binding is abnormal Manifests various mental disorders.

The benzodiazepine receptor present in the brain of mammals forms a complex with the inhibitor of GABA which is an inhibitory neurotransmitter (hereinafter referred to as "GABA receptor") and Cl ^- channel, and the action expression of GABA in the living body. Are known to be involved in.

[0005]

If a biological image of benzodiazepine receptor distribution in the brain can be obtained by the ESR method, which is a non-invasive measurement method, GABA receptors such as epilepsy, Parkinson's disease, cerebral ischemia, and cerebral edema are involved. It is useful for the study of pathological conditions that are considered to occur and the screening of drugs effective against these diseases or symptoms. Spin-labeled compounds as probes for obtaining biological images,
It must have a sufficient half-life, cross the blood-brain barrier, and selectively bind to the benzodiazepine receptor.

[0006]

Means for Solving the Problems The present inventors have used a benzodiazepine derivative and its similar compounds for the purpose of developing a spin-label compound that can be effectively used to obtain an ESR image of benzodiazepine receptor distribution in the brain. , A benzodiazepine derivative having a hydrogen atom at the 1-position and an oxo group at the 2-position or an oxo group at the 2-position and a hydroxyl group at the 3-position By spin-labeling the hydrogen atom at the 1-position or the hydroxyl group at the 3-position of the similar compound, E of the distribution of benzodiazepine receptors in the brain
The present invention has been completed by finding that a spin label compound useful for obtaining an image by the SR method can be obtained.

According to the present invention, a spin label compound represented by the following chemical formula 4 is provided.

[0008]

[Image Omitted] A- (CH ₂ ) _n CONH-B [wherein A is (a), (b),
Means any group selected from (c), (d), (e) and (f),

[0009]

[Chemical 5]

(Wherein R ¹ is a hydrogen atom, a halogen atom,
A nitro group, a cyano group or a trifluoromethyl group is meant, R ² is a hydrogen atom or a halogen atom, R ³ is a hydrogen atom or a methyl group, Me is a methyl group and Et is an ethyl group. Means )

B means a group represented by the following chemical formula 6,

[0012]

[Chemical 6]

(In the formula, X means a single bond or a methylene group, Me means a methyl group.) N means 1 or 2. ]

The halogen atom in the above chemical formula 5 means fluorine, chlorine, bromine and iodine, but chlorine and bromine are preferable. Among the compounds of the present invention represented by the above chemical formula 4, those in which n is 1 are preferable.

Among the compounds in which A is the group (a) in the above chemical formula 4, preferred are compounds in which R ¹ is a chlorine atom or a nitro group and R ² is a hydrogen atom. Among the compounds in which A is a group (b) in Chemical formula 4, preferred are
R ¹ is a chlorine or bromine atom, R ² is a hydrogen atom, R ³ is a hydrogen atom or a compound in which the 5-position of the oxazolidine ring portion is a methyl group. Among the compounds in which A is the group (d) in the chemical formula 4, preferred are compounds in which R ¹ is a chlorine atom.

The compound of the present invention is represented by the following chemical formula 7

## STR00007 ## A compound represented by A--H (wherein A means the same as above) and the following compound

Embedded image Y- (CH ₂ ) _n CONH-B

It can be produced by reacting with a compound represented by the formula (B and n have the same meanings as described above, and Y means a reactive ester residue of alcohol.).

Examples of the reactive ester residue of alcohol represented by Y in the above chemical formula 8 are halogen atom such as chlorine, bromine and iodine, methanesulfonyloxy,
Examples include lower alkylsulfonyloxy groups such as ethanesulfonyloxy, arylsulfonyloxy groups such as benzenesulfonyloxy, p-toluenesulfonyloxy, m-nitrobenzenesulfonyloxy.

The reaction between the compound of Chemical formula 7 and the compound of Chemical formula 8 is carried out in a suitable solvent, preferably in the presence of a base. Examples of the solvent used include N, N-dimethylformamide, tetrahydrofuran and dioxane, and examples of the base include sodium hydride. The reaction temperature is usually about 0 to 50 ° C. The product can be isolated and purified by a conventional method such as chromatography, recrystallization and reprecipitation.

The starting compound represented by the chemical formula 7 is J. Or
g. Chem., 27 , 3788 (1962), "Progress in Medicinal
Chemistry, "ed. By GP Ellis and GB West, Elsev
ier Science Publishers, BV, 1983, pp.157-163
Etc. according to the method described in the above.

The raw material compound represented by the above chemical formula 8 is commercially available or Biochemistry, 8 , 2580 (1969),
It can be produced according to the method described in J. Mol. Biol., 173 , 63 (1984) and the like.

[0022]

The compound of the present invention selectively binds to the benzodiazepine receptor present in the cranial nervous system, crosses the blood-brain barrier, and is stable while collecting the data necessary for constructing a biological image by the ESR method. Therefore, it is useful as a probe for imaging the benzodiazepine receptor distribution in the brain by the ESR method.

For example, 7-chloro-1,3-dihydro-
5-Phenyl-1- [N- (2,2,6,6-tetramethylpiperidin-1-oxyl-4-yl) carbamoylmethyl] -2H-1,4-benzodiazepin-2-one (see Examples below). Compound) can be used to obtain an image of benzodiazepine receptor distribution in the brain by the following procedure. The compound of Example was dissolved in a minimum volume of dimethylsulfoxide and then diluted with physiological saline to adjust the concentration to 3 ~.
Adjust to 15 mM. The total dose of this solution is about 15-100mg / in the carotid artery of small animals such as mice, gerbils and rats.
L-band ESR device reported by Shinichi Ishida et al. (Hiroshi Nishikawa,
Yoshikawa Toshikazu, "ESR and Free Radicals," Nihon Medical Museum, 1989, pp. 127-132), and according to their method, change the magnetic field gradient of the ESR spectrum in the head of the animal. While measuring online with a computer. Then, an image showing the distribution of benzodiazepine receptors in the brain can be obtained by constructing an image by the two-dimensional back projection method based on these data. The compounds of the examples are sufficiently stable while measuring the ESR spectrum.

Therefore, using the compound of the present invention as a probe, the benzodiazepine receptor in the brain of a disease model animal of a disease or symptom thought to involve GABA receptors such as epilepsy, Parkinson's disease, cerebral ischemia, and cerebral edema You can check the distribution state. By imaging the distribution of benzodiazepine receptors in the brain with the compound of the present invention as a probe before and after the administration of the test compound, it is possible to screen a drug effective for these diseases and symptoms.

In the future, when an ESR device capable of measuring an extremely large volume biological sample such as a human head is developed, an image of the benzodiazepine receptor distribution in the brain is obtained by the ESR method using the compound of the present invention as a probe. Get, GA
It can be expected to carry out non-invasive diagnosis of diseases or symptoms involving BA receptors.

[0026]

EXAMPLES The present invention will be described in more detail below with reference to reference examples and examples, but the present invention is not limited to these examples. The compounds are identified by elemental analysis values, mass spectra, IR spectra, NMR spectra,
It was performed by high performance liquid chromatography or the like.

Reference Example 4-Production of 4-chloroacetamide-2,2,6,6-tetramethylpiperidine-1-oxyl: -4-amino-2,2,6,6-tetramethylpiperidine-1-oxyl 1.00 g And 0.59 g of triethylamine were dissolved in 16 ml of toluene, and a solution of 0.66 g of chloroacetyl chloride in 5 ml of toluene was gradually added while stirring under cooling with ice water. After stirring at room temperature for 2 hours, 2 ml of saturated saline was added to the reaction solution, and the mixture was stirred for 30 minutes and then dried over anhydrous sodium sulfate. The solvent was distilled off under reduced pressure to obtain 1 g of a crude target product. Mass spectrum m / e: 247 (M ⁺ ). IR spectrum (KBr, cm ^-1 ): 3240 (NH), 1640 (C =
O).

Example 7-Chloro-1,3-dihydro-5-phenyl-1-
[N- (2,2,6,6-tetramethylpiperidine-1
-Oxyl-4-yl) carbamoylmethyl] -2H-
Preparation of 1,4-benzodiazepin-2-one:-

[0029]

[Chemical 9]

152 mg of 63% oily sodium hydride was suspended in 15 ml of N, N-dimethylformamide, and 7-chloro-1,3-dihydro-5-phenyl-2H was stirred while cooling with ice water.
After 1.03 g of -1,4-benzodiazepin-2-one was gradually added, the mixture was stirred at room temperature for 1 hour. A solution of 4-chloroacetamide-2,2,6,6-tetramethylpiperidine-1-oxyl (1 g) in N, N-dimethylformamide (2 ml) was gradually added thereto, and the mixture was stirred at room temperature for 4 hours and then left overnight. The reaction mixture was poured into ice water, extracted with a mixed solvent of ethyl acetate and diethyl ether, washed with water, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was recrystallized from chloroform-diethyl ether, and the obtained crystals were recrystallized again from ethyl acetate to obtain 1.40 g of the desired product. Melting point 225
~ 227 ° C

Elemental analysis values (as C ₂₆ H ₃₀ ClN ₄ O ₃ ):-Theoretical value: C, 64.79; H, 6.27; Cl, 7.36; N, 11.6
2. Experimental value: C, 64.77; H, 6.34; Cl, 7.46; N, 11.5
7. Mass spectrum m / e: 481 (M ⁺ ). IR spectrum (KBr, cm ^-1 ): 3280 (NH), 1655 (C =
O).

[0032]

INDUSTRIAL APPLICABILITY The compound of the present invention has a sufficient half-life, crosses the blood-brain barrier, and selectively binds to benzodiazepine receptors present in the cranial nervous system.
It is useful as a probe for obtaining an in-vivo image of benzodiazepine receptor distribution by the R method, and therefore GAB such as epilepsy, Parkinson's disease, cerebral ischemia, cerebral edema.
It can be used for the study of pathological conditions in which the A receptor is considered to be involved and for the screening of drugs effective for these diseases or symptoms.

Claims (1)

[Claims] 1. A spin label compound represented by the following chemical formula 1. Embedded image A- (CH ₂ ) _n CONH-B [wherein A is (a), (b), or
Means any group selected from (c), (d), (e) and (f), (Wherein R ¹ represents a hydrogen atom, a halogen atom, a nitro group, a cyano group or a trifluoromethyl group, R ² represents a hydrogen atom or a halogen atom, and R ³ represents a hydrogen atom or a methyl group. , Me means a methyl group, Et means an ethyl group.) B means a group represented by the following chemical formula 3, and (In the formula, X means a single bond or a methylene group, Me means a methyl group.) N means 1 or 2. ]