JP6645695B2 - Imidazopyridine amine compound, its production method and use - Google Patents

Description

  The present invention relates to an imidazopyridine amine compound having a nerve cell differentiation promoting action, a method for producing the same, and a use thereof.

  Imidazole, one of the nitrogen-containing aromatic heterocyclic compounds containing nitrogen atoms at the first and third positions on the 5-membered ring, has been widely applied in organic synthesis, and various compounds containing an imidazole structure are also used as pharmaceuticals Have been. Examples include anti-ulcer drugs (eg, cimetidine), anti-hypertensive drugs (eg, losartan), and anti-asthmatic drugs (eg, ozagrel). Therefore, application of the compound having an imidazole structure to pharmaceuticals is expected.

  In addition, advances in stem cell research have led to the development of a system (in vitro) that reproduces the development and differentiation of the central nervous system in the brain using pluripotent stem cells, and has developed Alzheimer's disease and There is a need for the development of new pharmaceutical compositions and agents for promoting the differentiation of pluripotent stem cells into nerve cells for the prevention and cure of neurodegenerative diseases represented by Parkinson's disease and the like.

  On the other hand, it has been reported that Ageladine A and its derivatives derived from marine organisms are isolated or synthesized and have strong anti-angiogenic activity (Patent Document 1, Non-Patent Document 1). Ageladine A is an imidazopyridine amine derivative having a dipromopyrrole ring. However, methods for total synthesis of Ageladine A and methods for synthesizing derivatives thereof are limited, and only very limited compounds of the derivatives have been synthesized. Further, the physiological activities of these compounds are not known.

U.S. Patent Publication 2011/0152313 A1

Shengule S R et al., J. Med.Chem. 2011, 54, 2492-2501.

  An imidazopyridine amine compound or a pharmaceutically acceptable salt or solvate thereof, which has an excellent neuronal cell differentiation promoting action and high safety, and a method for producing the same, and for preventing or treating a neurodegenerative disease thereof And a method for screening for a compound that regulates differentiation of a pluripotent stem cell into a neural cell, and a compound that regulates differentiation into a neural cell.

  The present inventors have developed a new method for synthesizing imidazopyridine amine compounds. Therefore, the present inventors synthesized a novel imidazopyridine amine compound, evaluated its neuronal cell-inducing activity and cytotoxicity, and confirmed its superior neuronal cell differentiation promoting action and high safety. The present invention has been completed.

Specifically, the present invention provides a compound represented by the following formula 1, or a pharmaceutically acceptable salt or solvate thereof.

In the above equation 1,
R ₁ , R ₂ , R ₃ and X may be the same or different and are each independently H, halogen, cyano, nitro, optionally substituted amino, optionally substituted acyl, substituted Optionally substituted alkoxy, optionally substituted carbamoyl, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted cycloalkyl, substituted Optionally substituted cycloalkenyl, optionally substituted aryl, optionally substituted arylalkyl, optionally substituted aralkyl, optionally substituted heterocycloalkyl, optionally substituted heteroaryl Or, represents an optionally substituted heteroarylalkyl,
R ₁ and R ₂ or R ₁ and R ₃ may be taken together with the N atom to form an unsubstituted or substituted 5- to 8-membered heterocycle;
m is 0, 1 or 2;
Ring A represents an optionally substituted carbocycle or heterocycle.

In the compound represented by the formula 1 of the present invention,
R ₃ is selected from optionally substituted phenyl, optionally substituted naphthyl or optionally substituted ethyl,
The ring A may be one ring selected from the following formulas 2 to 4.





Here, Y, Y ′, Y ″ and Y ″ ″ may be the same or different, and each is independently halogen, cyano, nitro, optionally substituted amino, substituted Optionally substituted acyl, optionally substituted alkoxy, optionally substituted carbamoyl, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted Cycloalkyl, optionally substituted cycloalkenyl, optionally substituted aryl, optionally substituted arylalkyl, optionally substituted aralkyl, optionally substituted heterocycloalkyl, substituted Represents an optionally substituted heteroaryl, or an optionally substituted heteroarylalkyl,
p is 0 or an integer of 1 to 4,
q is 0, 1 or 2,
r is 0 or an integer of 1 to 4,
s is 0 or an integer of 1 to 3,
May be represented.

The compound of the present invention may be a compound represented by the following formulas 5 to 22, or a pharmaceutically acceptable salt or solvate thereof.

The present invention also provides a method for producing an imidazopyridine amine compound represented by the following reaction formula 1,
(i) Using the substituted amine derivative (1) as a raw material, guanidination (3) by reaction with a cyanamide compound (2),
(ii) Next, reacting with a conjugated dialdehyde compound (4) to form an imidazole compound (5),
(iii) Next, an imine is formed with the ammonium salt (6) and the aldehyde compound (7), and the imidazopyridine ring is formed by advancing the subsequent aza electron cyclization reaction to produce an imidazopyridine amine compound (8). A manufacturing method is provided.

  In the manufacturing method, the manufacturing method may be a one-pot method.

  In addition, the present invention provides a medicament containing a compound represented by the above formula 1 or 5 to 22 or a pharmaceutically acceptable salt or solvate thereof as an active ingredient.

  The present invention also provides a pharmaceutical composition for preventing or treating a neurodegenerative disease, comprising as an active ingredient a compound represented by Formula 1 or Formula 5 to Formula 22 or a pharmaceutically acceptable salt or solvate thereof. I will provide a.

  In the pharmaceutical composition of the present invention, the neurodegenerative disease may be selected from Alzheimer's disease, mild cognitive impairment, Parkinson's disease, dementia with Lewy bodies, and multiple sclerosis.

  In addition, the present invention promotes the differentiation of pluripotent stem cells into nerve cells, comprising the compound of the above formula 1 or the formulas 5 to 22 or a pharmaceutically acceptable salt or solvate thereof as an active ingredient. For promoting neuronal differentiation.

  Further, the present invention provides a method for inhibiting differentiation into nerve cells, which comprises the step of adding a compound represented by the above formula 1 or the formula 5 or a pharmaceutically acceptable salt or solvate thereof to cultured cells in vitro. Methods for screening for compounds that modulate are provided.

  In the screening method of the present invention, the cultured cells may be pluripotent stem cells.

  The present invention provides a production method capable of synthesizing an imidazopyridine amine compound in one pot, a compound synthesized by the production method, a neuronal cell differentiation promoting action, a use of the compound as a medicament, and a pharmaceutical composition containing the compound.

The figure showing the result of having evaluated the concentration-dependent nerve cell differentiation promotion effect of the compound 1-3 and the harmine which is a positive control compound. The figure showing the result of having evaluated the astrocyte differentiation promotion effect of compound 1-3 and harmin.

1. Compound, or a pharmaceutically acceptable salt or solvate thereof As one embodiment of the present invention, a compound represented by the following formula 1 or a pharmaceutically acceptable salt or solvate thereof is provided. .



In the above equation 1,
R ₁ , R ₂ , R ₃ and X may be the same or substituted, and each is independently H, halogen, cyano, nitro, optionally substituted amino, substituted Optionally substituted acyl, optionally substituted alkoxy, optionally substituted carbamoyl, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted Good cycloalkyl, optionally substituted cycloalkenyl, optionally substituted aryl, optionally substituted arylalkyl, optionally substituted aralkyl, optionally substituted heterocycloalkyl, substituted Represents an optionally substituted heteroaryl, or an optionally substituted heteroarylalkyl,
R ₁ and R ₂ or R ₁ and R ₃ may be taken together with the N atom to form an unsubstituted or substituted 5- to 8-membered heterocycle;
m is 0, 1 or 2;
Ring A represents an optionally substituted carbocycle or heterocycle.

  In the present specification, the halogen includes any atom of F, Cl, Br or I.

  In the present specification, “amino” includes a primary amino group, a secondary amino group, a tertiary amino group, and a quaternary amino group.

  In the present specification, examples of “acyl” include, unless otherwise specified, for example, formyl; carboxyl; carbamoyl; alkyl-carbonyl; alkoxy-carbonyl; cycloalkyl-carbonyl; aryl-carbonyl; aralkyl-carbonyl; -Carbonyl; aralkyloxy-carbonyl; mono- or di-alkyl-carbamoyl; mono- or di-aryl-carbamoyl; mono- or di-cycloalkyl-carbamoyl; mono- or di-aralkyl-carbamoyl.

  In the present specification, “alkoxy” includes, for example, methoxy, ethoxy, propoxy, isopropoxy, butoxy, isobutoxy, tert-butoxy, pentyloxy, hexyloxy and the like, unless otherwise specified.

  In the present specification, unless otherwise specified, the term "carbamoyl" includes, for example, "mono- or di-alkyl-carbamoyl", "mono- or di-aryl-carbamoyl", "mono- or di-cycloalkyl-". Carbamoyl "," mono- or di-aralkyl-carbamoyl "," mono- or di-5heterocyclic-carbamoyl "and the like.

  In the present specification, examples of “alkyl” include, unless otherwise specified, for example, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl, tert-butyl, pentyl, isopentyl, neopentyl, hexyl, and the like. .

  In the present specification, unless otherwise specified, “alkenyl” includes, for example, vinyl, propenyl, isopropenyl, 2-buten-1-yl, 4-penten-1-yl, 5-hexen-1-yl And the like.

  In the present specification, “alkynyl” includes, for example, 2-butyn-1-yl, 4-pentyn-1-yl, 5-hexyn-1-yl and the like unless otherwise specified.

  In the present specification, “cycloalkyl” includes, for example, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl and the like unless otherwise specified.

  In the present specification, the term “cycloalkenyl” includes, for example, a C 3-8 cycloalkenyl group (eg, 2-cyclopropenyl, 2-cyclobutenyl, 2-cyclopentenyl, 3-cyclopentenyl, 2-cyclopentenyl) unless otherwise specified. Hexenyl, 3-cyclohexenyl, 2-cycloheptenyl, 3-cycloheptenyl, 4-cycloheptenyl, 2-cyclooctenyl, 3-cyclooctenyl, and 4-cyclooctenyl, which may be further condensed with an aryl group such as a phenyl group. For example, indenyl, benzocyclohexenyl, benzocycloheptenyl, benzocyclooctenyl, etc. Among them, preferred are 2-cyclohexenyl, 2-cycloheptenyl, cyclooctyl, indenyl and the like.

  In the present specification, “aryl” includes, for example, phenyl, 1-naphthyl, 2-naphthyl, 2-biphenylyl, 3-biphenylyl, 4-biphenylyl, 2-anthryl and the like, unless otherwise specified. The C6-14 aryl may be partially saturated, and examples of the partially saturated C6-14 aryl include tetrahydronaphthyl.

  In the present specification, “arylalkyl” means a group in which the above “alkyl” is substituted by the above “aryl”, and specifically, for example, benzyl, 1-phenethyl, 2-phenethyl, naphthylmethyl, 1-naphthylethyl , 2-naphthylethyl.

  In the present specification, “aralkyl” includes, unless otherwise specified, for example, benzyl, phenethyl, diphenylmethyl, 1-naphthylmethyl, 2-naphthylmethyl, 2,2-diphenylethyl, 3-phenylpropyl, 4-phenylpropyl Examples include phenylbutyl, 5-phenylpentyl, 2-biphenylylmethyl, 3-biphenylylmethyl, 4-biphenylylmethyl and the like.

  As used herein, the term “heterocycloalkyl” refers to a monocyclic 3- to 7-membered ring containing 1 to 4 heteroatoms consisting of an oxygen atom, a nitrogen atom or a sulfur atom, or the same or different monocyclic ring. A ring is a condensed polycyclic heterocyclic group.Specific examples include piperidinyl, pyrrolidinyl, piperazinyl, tetrahydrofuryl, tetrahydropyranyl, morphonyl, atidinyl, imidazolidinyl, oxazolidinyl, hexahydropyrrolitinyl, octahydroindine Examples include tinyl, octahydroquinolitinyl, octahydroindolyl, and oxo-oxidized forms of these substituents.

  In the present specification, “heterocycloaryl” means, for example, having 2 to 12 carbon atoms and containing 1 to 4 hetero atoms composed of an oxygen atom, a nitrogen atom or a sulfur atom, unless otherwise specified. Or a polycyclic heteroaryl group in which two or more of the same or different monocyclic rings are fused, such as pyrrole, furyl, thienyl, imidazolyl, thiazolyl, pyrazyl, indolyl, quinolyl, isoquinolyl , Tetrazolyl, pyridinyl, pyrazolyl, pyridazinyl, pyrimidinyl groups and the like.

  In the present specification, “heteroarylalkyl” includes, for example, a pyridylmethyl group, a pyridylethyl group, a thienylmethyl group, a thienylethyl group, and the like, unless otherwise specified.

  In the present specification, the term "carbocyclic" refers to a cyclic substituent having carbon as a constituent atom, and unless otherwise specified, for example, cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclopentadienyl, cyclohexyl, cyclohexyl Hexenyl, cyclohexadienyl, cycloheptanyl, cyclooctanyl and the like can be mentioned.

  In the present specification, as a “heterocycle” (“heterocycle”), unless otherwise specified, for example, one or two kinds selected from a nitrogen atom, a sulfur atom and an oxygen atom other than a carbon atom as a ring-constituting atom A 5- to 14-membered (monocyclic, bicyclic or tricyclic) heterocycle containing from 1 to 4 heteroatoms, preferably (i) a 5- to 14-membered (preferably 5- to 10-membered) aromatic heterocycle, (Ii) 5- to 10-membered non-aromatic heterocyclic ring and the like. Among them, a 5- or 6-membered aromatic heterocycle is preferable. Specifically, for example, thienyl (eg, 2-thienyl, 3-thienyl), furyl (eg, 2-furyl, 3-furyl), pyridyl (eg, 2-pyridyl, 3-pyridyl, 4-pyridyl), Thiazolyl (eg, 2-thiazolyl, 4-thiazolyl, 5-thiazolyl), oxazolyl (eg, 2-oxazolyl, 4-oxazolyl, 5-oxazolyl), pyrazinyl, pyrimidinyl (eg, 2-pyrimidinyl, 4-pyrimidinyl), pyrrolyl (Eg, 1-pyrrolyl, 2-pyrrolyl, 3-pyrrolyl), imidazolyl (eg, 1-imidazolyl, 2-imidazolyl, 4-imidazolyl), pyrazolyl (eg, 1-pyrazolyl, 3-pyrazolyl, 4-pyrazolyl), Triazolyl (eg, 1-triazolyl, 2-triazolyl), tetrazolyl, pyridazinyl (eg, 3-pyridazi) , 4-pyridazinyl), isothiazolyl (eg, 3-isothiazolyl, 4-isothiazolyl, 5-isothiazolyl), isoxazolyl (eg, 3-isoxazolyl, 4-isoxazolyl, 5-isoxazolyl), indolyl (eg, 1-indolyl, 2 -Indolyl, 3-indolyl), 2-benzothiazolyl, 2-benzoxazolyl, benzimidazolyl (eg, 1-benzimidazolyl, 2-benzimidazolyl), benzo [b] thienyl (eg, 2-benzo [b] thienyl) , 3-benzo [b] thienyl), benzo [b] furanyl (eg, 2-benzo [b] furanyl, 3-benzo [b] furanyl), quinolyl (eg, 2-quinolyl, 3-quinolyl, 4-quinolyl) , 5-quinolyl, 8-quinolyl), isoquinolyl (eg, 1-isoquinolyl, 3-iso Aromatic heterocycles such as noryl, 4-isoquinolyl, 5-isoquinolyl); for example, pyrrolidinyl (eg, 1-pyrrolidinyl, 2-pyrrolidinyl, 3-pyrrolidinyl), oxazolidinyl (eg, 2-oxazolidinyl), imidazolinyl (eg, 1) -Imidazolinyl, 2-imidazolinyl, 4-imidazolinyl), piperidinyl (eg, piperidino, 2-piperidinyl, 3-piperidinyl, 4-piperidinyl), piperazinyl (eg, 1-piperazinyl, 2-piperazinyl), morpholinyl (eg, 2- Non-aromatic heterocycles such as morpholinyl, 3-morpholinyl, morpholino), thiomorpholinyl (eg, 2-thiomorpholinyl, 3-thiomorpholinyl, thiomorpholino), and tetrahydropyranyl.

  As used herein, the term "optionally substituted" means (i) unsubstituted or (ii) the above-mentioned "halogen", "cyano", "nitro", "amino", "acyl", "Alkoxy", "carbamoyl", "alkyl", "alkenyl", "alkynyl", "cycloalkyl", "cycloalkenyl", "aryl", "arylalkyl", "aralkyl", "heterocycloalkyl", " One or two or more identical or different substituents selected from “heteroaryl” or “heteroarylalkyl” are substituted alone or in a combined state by combining two or more substituents. Refers to the case of having a configuration.

The compound represented by the formula 1 can be synthesized by the following reaction route.

  Using the substituted amine derivative (1) as a raw material, (i) guanidination (3) by reaction with cyanamide (2), and further (ii) reaction with conjugated dialdehyde (4) to form an imine, (iii) ) Construct an imidazole ring by intramolecular conjugate addition reaction. Then, (iv) by the action of ammonium salt, the equilibrium between imine and enamine with the other aldehyde, and further (v) imine formation with the newly introduced aldehyde, followed by (vi) aza A dihydropyridine ring is formed by advancing the electron cyclization reaction. Furthermore, (vii) after conversion to a pyridine ring by natural oxidation, finally (viii) removal of the aryl group derived from aniline.

In the above-mentioned production method, all steps leading to the compound of the present invention can be synthesized by a one-pot method by introducing a reagent into a flask at appropriate time intervals.

  Preferred examples of the compound represented by the formula 1 include the following compounds 1 to 22, but are not limited thereto.

Compound 1: 4- (pyridin-2-yl) -1H-imidazo [4,5-c] pyridin-2-amine is represented by the following formula 5.

Compound 2: 1-phenyl-4- (pyridin-2-yl) -1H-imidazo [4,5-c] pyridin-2-amine is represented by the following formula 6.

Compound 3: methyl (2S) -2- (acetylamino) 5- [2-amino-4- (pyridin-2-yl) -1H-imidazo [4,5-c] pyridin-1-yl] pentanoate is as follows: It is expressed by Equation 7.

Compound 4: 1- (4-fluorophenyl) -4- (pyridin-2-yl) -1H-imidazo [4,5-c] pyridin-2-amine is represented by the following formula 8.

Compound 5: 1- (4-methyoxyphenyl) -4- (pyridin-2-yl) -1H-imidazo [4,5-c] pyridin-2-amine is represented by the following formula 9.

Compound 6: 1- (3,4-dimethyoxyphenyl) -4- (pyridin-2-yl) -1H-imidazo [4,5-c] pyridin-2-amine is represented by the following formula 10.

Compound 7: 1- (2,4-dimethyoxyphenyl) -4- (pyridin-2-yl) -1H-imidazo [4,5-c] pyridin-2-amine is represented by the following formula 11.

Compound 8: 4- (pyridin-2-yl) -1- (3,4,5-trimthoxyphenyl) -1H-imidazo [4,5-c] pyridin-2-amine is represented by the following formula 12.

Compound 9: 4- [2-amino-4- (pyridin-2-yl) -1H-imidazo [4,5-c] pyridin-1-yl] phenol is represented by the following formula 13.

Compound 10: 1- (4-tert-butylphenyl) -4- (pyridin-2-yl) -1H-imidazo [4,5-c] pyridin-2-amine is represented by the following formula 14.

Compound 11: 4- (pyridin-2-yl) -1- [4- (trifluoromethyl) phenyl] -1H-imidazo [4,5-c] pyridin-2-amine is represented by the following formula 15.

Compound 12: 1- (2,4-dinitrophenyl) -4- (pyridin-2-yl) -1H-imidazo [4,5-c] pyridin-2-amine is represented by the following formula 16.

Compound 13: 1- (naphthalen-1-yl) -4- (pyridin-2-yl) -1H-imidazo [4,5-c] pyridin-2-amine is represented by the following formula 17.

Compound 14: 1- (3H-indol-6-yl) -4- (pyridin-2-yl) -1H-imidazo [4,5-c] pyridin-2-amine is represented by the following formula 18.

Compound 15: 4- (isoquinolin-1-yl) -1-phenyl-1H-imidazo [4,5-c] pyridin-2-amine is represented by the following formula 19.

Compound 16: 1- (3,4-dimethyoxyphenyl) -4- [6- (trifluoromethyl) pyridin-2-yl) -1H-imidazo [4,5-c] pyridin-2-amine is represented by the following formula 20: Is done.

Compound 17: 4- (4,5-dibromo-2H-pyrrol-2-yl) -1- (3,4-dimethoxyphenyl) -1H-imidazo [4,5-c] pyridin-2-amine has the following formula 21.

Compound 18: 4- [2-amino-4- (4,5-dibromo-2H-pyrrol-2-yl) -1H-imidazo [4,5-c] pyridin-1-yl] phenol has the following formula 22 It is represented by

  Pharmaceutically acceptable salts include, for example, metal salts, ammonium salts, salts with organic bases, salts with inorganic acids, salts with organic acids, salts with basic or acidic amino acids, and the like, It is not limited to these.

  Preferable examples of the metal salt include an alkali metal salt such as a sodium salt and a potassium salt; an alkaline earth metal salt such as a calcium salt, a magnesium salt and a barium salt; an aluminum salt.

  Preferred examples of the salt with an organic base include salts with trimethylamine, triethylamine, pyridine, ethanolamine, diethanolamine, triethanolamine and the like.

  Preferable examples of salts with inorganic acids include salts with hydrochloric acid, hydrobromic acid, nitric acid, sulfuric acid, phosphoric acid, and the like.

  Preferred examples of salts with organic acids include formic acid, acetic acid, trifluoroacetic acid, phthalic acid, fumaric acid, oxalic acid, tartaric acid, maleic acid, citric acid, succinic acid, malic acid, methanesulfonic acid, benzenesulfonic acid , P-toluenesulfonic acid and the like.

  Preferable examples of the salt with a basic amino acid include salts with arginine, lysine, ornithine and the like, and preferable examples of the salt with an acidic amino acid include salts with aspartic acid, glutamic acid and the like.

  The pharmaceutically acceptable solvate is not particularly limited as long as it forms a solvate with the compound represented by Formula 1 or the compounds 1 to 18, but is not limited to hydrate or ethanol. Japanese is preferred.

2. Pharmaceuticals and Pharmaceutical Compositions for Prevention or Treatment of Neurodegenerative Diseases In another embodiment of the present invention, one of the compounds represented by Formula 1 or one of the compounds 1 to 18 is administered alone. For the prevention or treatment of neurodegenerative diseases by administering simultaneously or at different times in combination with two or more kinds of multiple compounds or other neuronal cell differentiation promoting agents (eg, All-trans retinoic acid), etc. It can be used as a pharmaceutical composition.

  In the present invention, the neural cell differentiation promoting action refers to an action of promoting the efficiency of differentiation into neural cells in the process of inducing differentiation of neural stem cells such as mouse embryonic stem (ES) cells-derived neural stem cells into neural cells. Say.

  The neuronal differentiation promoting action in the present invention is an action in vitro, and if the action is observed in vitro, it can be expected that the action has a neuronal differentiation promoting action in vivo or in an actual living body.

  As a pharmaceutical preparation containing the compound of the present invention, or a pharmaceutically acceptable salt or solvate thereof as an active ingredient, an oral preparation or a parenteral preparation can be prepared and used.

  The compound represented by the formula 1 or the compound 1 to 18, preferably, at least one of the compound 1 to 18 or a pharmaceutically acceptable salt or solvate thereof, for example, with a solvent such as physiological saline. It can be diluted to a predetermined concentration and dose, added with a pharmaceutically acceptable excipient, and administered to a patient as a pharmaceutical preparation.

  The pharmaceutical preparation is a pharmaceutical composition containing the compound having the activity of promoting neuronal differentiation, or a salt or solvate thereof as an active ingredient, and is preferably a pharmaceutical composition for preventing or treating a neurodegenerative disease. But is not limited to this.

  As used herein, the term "neurodegenerative disease" refers to a progressive disease in which nerve cells of the central nervous system gradually degenerate and fall into cell death.Specifically, the present invention is not limited thereto, and includes Alzheimer's disease, dementia, Mild cognitive impairment, senile dementia, Parkinson's disease, Lewy body dementia, amyotrophic lateral sclerosis, Creutzfeldt-Jakob disease, Fahren's disease, Huntington's disease and related polyglutamine elongation diseases, Menkes disease, multiple Includes sclerosis as well as Wilson's disease.

  Pharmaceutically acceptable excipients used in preparing the formulations include stabilizers, antioxidants, pH adjusters, buffers, suspending agents, emulsifiers, surfactants, and the like, which are known to those skilled in the art. Can be prepared by adding the following additives. The types of these pharmaceutical excipients and their usage and dosage are described in the Pharmaceutical Excipients Encyclopedia 2007 (edited by the Japan Pharmaceutical Excipients Association, Yakuji Nippo, July 2007), etc., prepared and used according to these descriptions. be able to.

  More specifically, tartaric acid, citric acid, succinic acid, organic acids such as fumaric acid as a stabilizer, ascorbic acid, dibutylhydroxytoluene or propyl gallate as an antioxidant, dilute hydrochloric acid or Sodium hydroxide aqueous solution or the like, as a buffer citric acid, succinic acid, fumaric acid, tartaric acid or ascorbic acid or salts thereof, glutamic acid, glutamine, glycine, aspartic acid, alanine or arginine or salts thereof, magnesium oxide, zinc oxide, water Magnesium oxide, phosphoric acid or boric acid or salts thereof, as a suspending agent or emulsifier, lecithin, sucrose fatty acid ester, polyglycerin fatty acid ester, polyoxyethylene hydrogenated castor oil, polysorbate or polyoxyethylene / polyoxypropylene copolymer Etc. Such as polysorbate 80 as a surfactant, sodium lauryl sulfate or polyoxyethylene hardened castor oil and the like, but not limited to be used.

Examples of the formulation of the pharmaceutical composition include, but are not limited to, the following.
Formulation Example 1 (manufacture of capsule)
1) Compound 3 1 mg
2) Fine powdered cellulose 10 mg
3) Lactose 18 mg
4) 1 mg of magnesium stearate
30 mg in total
The above 1), 2), 3) and 4) are mixed and filled into a gelatin capsule.

Formulation Example 2 (tablet production)
1) The above compound 31 g
2) Lactose 50 g
3) 14 g corn starch
4) 44 g of calcium carboxymethylcellulose
5) Magnesium stearate 1 g
1000 tablets total 110 g
The total amount of the above 1), 2) and 3) and 30 g of 4) are kneaded with water, dried under vacuum, and sized. 14 g of 4) and 1 g of 5) are mixed with the sized powder and tableted with a tableting machine. In this way, 1000 tablets containing 1 mg of the compound of Example 1 per tablet are obtained.

  The dose can be usually administered at about 0.01 to 30 mg / kg / day, but may be appropriately changed depending on the disease or condition of the administration subject, and the age, sex, administration method (eg, oral, parenteral) of the administration subject, etc. Is also adjusted appropriately.

  In addition, as an embodiment of the use of the present pharmaceutical composition, an effective amount of the compound represented by the formula 1 or the compound 1 to 18, preferably the compound 1 to 18 or a pharmaceutically acceptable salt or solvate thereof is used. And a method for preventing or treating a neurodegenerative disease, which comprises the step of administering to a patient having a neurodegenerative disease.

3. Nerve cell differentiation promoter As another embodiment of the present invention, a nerve cell differentiation promoter from pluripotent stem cells is provided.

  The nerve cell differentiation promoting agent is obtained by adding an additive to the compound represented by the formula 1 or the compound 1 to 18, preferably one or more compounds of the compounds 1 to 18, or a salt or solvate thereof. Can be adjusted.

  The nerve cell differentiation promoting agent is, for example, a stabilizer or an antioxidant known to those skilled in the art, provided that the activity of the compound represented by the formula 1 or the compounds 1 to 18 is not affected. It can be prepared by adding additives such as a pH adjuster, a buffer, a suspending agent, an emulsifier, and a surfactant. The types of these pharmaceutical excipients and their usage and dosage are described in the Pharmaceutical Excipients Encyclopedia 2007 (edited by the Japan Pharmaceutical Excipients Association, Yakuji Nippo, July 2007), etc., prepared and used according to these descriptions. be able to. In addition, known additives that are not described in the Pharmaceutical Excipients Dictionary 2007 (edited by the Japan Pharmaceutical Excipients Association, Yakuji Nippo-sha, July 2007) can also be used.

  Specifically, tartaric acid, citric acid, organic acids such as succinic acid or fumaric acid as stabilizers, for example, ascorbic acid, dibutylhydroxytoluene or propyl gallate as antioxidants, dilute hydrochloric acid or water as pH adjusters Sodium oxide aqueous solution or the like, as a buffer, citric acid, succinic acid, fumaric acid, tartaric acid or ascorbic acid or salts thereof, glutamic acid, glutamine, glycine, aspartic acid, alanine or arginine or salts thereof, magnesium oxide, zinc oxide, hydroxide Magnesium, phosphoric acid or boric acid or salts thereof, as a suspending or emulsifying agent, lecithin, sucrose fatty acid ester, polyglycerin fatty acid ester, polyoxyethylene hydrogenated castor oil, polysorbate or polyoxyethylene / polyoxypropylene copolymer, etc. But, Polysorbate 80 as a surface active agent, sodium lauryl sulfate or polyoxyethylene hardened castor oil and the like, but not limited to be used.

  The concentration of the neuron differentiation promoting agent expected at the time of incubation with the cultured pluripotent stem cells is 0.001 to 10 μM, but the compound represented by the formula 1 or the compound 1 to 18 alone or in combination, or another compound It may be used in combination with a nerve cell differentiation promoting agent (eg, All-trans retinoic acid) or the like, and changes as appropriate.

4. Method for Screening Compound that Regulates Neuronal Cell Differentiation-Promoting Activity As another embodiment of the present invention, a method for screening a compound that regulates neuronal cell differentiation-promoting activity is provided.

  The method for screening a compound that regulates the activity of promoting neuronal differentiation according to the present invention may be performed, for example, by using a compound represented by the above formula 1 or a compound 1 to 18, preferably a compound 1 to 18, in a cultured cell of pluripotent stem cells. Is added at a concentration that expresses neuronal differentiation promoting activity, and a test compound is also added. The cells are cultured for a certain period of time, the ratio of cells differentiated into nerve cells is measured, and a compound that regulates the nerve cell differentiation promoting activity is selected by comparing with a control group to which no test compound is added.

  The test compound in which the nerve cell differentiation promoting activity is recognized by the above method targets an action point different from the action point of the nerve cell differentiation promotion activity of the compound represented by the formula 1 or the compound 1 to 18 of the present invention. The compound can be used as a candidate for an active ingredient of a nerve cell differentiation promoting agent.

  On the other hand, a test compound which has been shown to have a neuronal cell differentiation inhibitory activity by the above method is used as a candidate for an active ingredient of a medicament for preventing or treating a disease such as a brain tumor based on the differentiation and proliferation of nerve cells such as a neuroma. it can.

  All documents mentioned herein are incorporated by reference in their entirety. The examples described herein are illustrative of embodiments of the present invention and should not be construed as limiting the scope of the invention.

<Synthesis of Compound>
Materials and Methods Regarding the synthesis route, the following compounds 1 to 18 were produced by the method represented by the following reaction formula 3.

  Using a substituted amine derivative (1) such as a substituted aniline derivative as a raw material, (i) guanidination (3) by reaction with cyanamide (2), and further (ii) reaction with conjugated dialdehyde (4) to form an imine After that, (iii) an imidazole ring (5) was constructed by an intramolecular conjugate addition reaction. Then, (iv) the ammonium salt (6) is acted on to cause the imine and enamine to equilibrate with the other aldehyde, and (v) the imine with the newly introduced aldehyde compound (7) Formation and subsequent (vi) aza electron cyclization reaction were allowed to proceed to form an imidazopyridine ring, and an imidazopyridine amine compound (8) was synthesized. Furthermore, after (vii) conversion to a pyridine ring by natural oxidation, finally (viii) removal of the aryl group derived from aniline was performed. In the synthesis of Compound 1 of the present invention, the guanidine derivative (3) in the above reaction formula was isolated, and the subsequent synthesis reaction was performed. On the other hand, for compounds 2 to 18 of the present invention, all steps were synthesized in one pot by introducing reagents into flasks at appropriate time intervals.

Compound 1: 4- (pyridin-2-yl) -1H-imidazo [4,5-c] pyridin-2-amine is represented by the following formula 5.

Compound 2: 1-phenyl-4- (pyridin-2-yl) -1H-imidazo [4,5-c] pyridin-2-amine is represented by the following formula 6.

Compound 3: methyl (2S) -2- (acetylamino) 5- [2-amino-4- (pyridin-2-yl) -1H-imidazo [4,5-c] pyridin-1-yl] pentanoate is as follows: It is expressed by Equation 7.

Compound 4: 1- (4-fluorophenyl) -4- (pyridin-2-yl) -1H-imidazo [4,5-c] pyridin-2-amine is represented by the following formula 8.

Compound 5: 1- (4-methyoxyphenyl) -4- (pyridin-2-yl) -1H-imidazo [4,5-c] pyridin-2-amine is represented by the following formula 9.

Compound 6: 1- (3,4-dimethyoxyphenyl) -4- (pyridin-2-yl) -1H-imidazo [4,5-c] pyridin-2-amine is represented by the following formula 10.

Compound 7: 1- (2,4-dimethyoxyphenyl) -4- (pyridin-2-yl) -1H-imidazo [4,5-c] pyridin-2-amine is represented by the following formula 11.

Compound 8: 4- (pyridin-2-yl) -1- (3,4,5-trimthoxyphenyl) -1H-imidazo [4,5-c] pyridin-2-amine is represented by the following formula 12.

Compound 9: 4- [2-amino-4- (pyridin-2-yl) -1H-imidazo [4,5-c] pyridin-1-yl] phenol is represented by the following formula 13.

Compound 10: 1- (4-tert-butylphenyl) -4- (pyridin-2-yl) -1H-imidazo [4,5-c] pyridin-2-amine is represented by the following formula 14.

Compound 11: 4- (pyridin-2-yl) -1- [4- (trifluoromethyl) phenyl] -1H-imidazo [4,5-c] pyridin-2-amine is represented by the following formula 15.

Compound 12: 1- (2,4-dinitrophenyl) -4- (pyridin-2-yl) -1H-imidazo [4,5-c] pyridin-2-amine is represented by the following formula 16.

Compound 13: 1- (naphthalen-1-yl) -4- (pyridin-2-yl) -1H-imidazo [4,5-c] pyridin-2-amine is represented by the following formula 17.

Compound 14: 1- (3H-indol-6-yl) -4- (pyridin-2-yl) -1H-imidazo [4,5-c] pyridin-2-amine is represented by the following formula 18.

Compound 15: 4- (isoquinolin-1-yl) -1-phenyl-1H-imidazo [4,5-c] pyridin-2-amine is represented by the following formula 19.

Compound 16: 1- (3,4-dimethyoxyphenyl) -4- [6- (trifluoromethyl) pyridin-2-yl) -1H-imidazo [4,5-c] pyridin-2-amine is represented by the following formula 20: Is done.

Compound 17: 4- (4,5-dibromo-2H-pyrrol-2-yl) -1- (3,4-dimethoxyphenyl) -1H-imidazo [4,5-c] pyridin-2-amine has the following formula 21.

Compound 18: 4- [2-amino-4- (4,5-dibromo-2H-pyrrol-2-yl) -1H-imidazo [4,5-c] pyridin-1-yl] phenol has the following formula 22 It is represented by

  Specifically, the compounds 1 to 18 were synthesized using the starting material (1) and the aldehyde compound (7) shown in Tables 1-1 to 1-3 in the reaction formula of the reaction formula 3.

  The reaction conditions for Compound 1 and Compound 5 are described below. For the synthesis of Compounds 2 to 4 and 6 to 18, the reaction conditions for Compound 5 were followed.

Method for synthesizing compound 1
2,4-dinitrofluorobenzene (3.0 mL, 28 mmoL) and guanidine hydrochloride (2.2 g, 23 mmoL) in anhydrous N, mixed in N- dimethylformamide (DMF, 120 mL), triethylamine (NEt _3, 6.5 mL, 47 mmol) at room temperature under a nitrogen atmosphere. After stirring for 24 hours, the mixture was slowly cooled with ice to 0 ° C., and a 1.0 M hydrochloric acid solution (200 mL) was added. After washing the mixture with chloroform (3 × 100 mL), the aqueous layer was neutralized with sodium hydrogen carbonate, and the reaction product was extracted with chloroform (4 × 300 mL). The organic layer was dried using sodium sulfate and purified under reduced pressure. The obtained crude product was recrystallized from ethyl acetate to give 2,4-dinitrophenylguanidine as yellow needles (yield: 2.8 g, 52.6%).

  A mixture of 2,4-dinitrophenylguanidine (67.0 mg, 0.297 mmol) and dialdehyde (50.0 mg, 0.595 mmol) dissolved in DMF (1.5 mL) was stirred at room temperature for 1 hour, and then ammonium chloride (79.5 mg, 1.49 mmol), methanol (1.0 mL) and pyridine-2-carbaldehyde (28.3 mg, 0.297 mmol) were added, and the mixture was stirred at 75 ° C for 2 hours. After ice cooling to room temperature, a saturated sodium hydrogen carbonate solution (1.0 mL) and 2-mercaptoethanol (1.0 mL) were added, and the mixture was further stirred at room temperature for 1 hour. Compound 1 was obtained from the resulting reaction solution using reverse phase HPLC (yield: 13.7 mg, 22%).

Method for synthesizing compound 5 In the above reaction formula 3, 4-methoxyaniline (45.5 mg, 0.3 mmol) was used as a raw material, and cyanamide (37.8 mg, 0.9 mmol) was mixed with methanol (0.3 mL). The solution (75 μL) was added. After stirring under reflux conditions for 12 hours, the mixture was cooled to room temperature and neutralized with NEt ₃ (125 μL, 0.9 mmol). Thereafter, the mixture containing dialdehyde (50.0 mg, 0.6 mmol) was stirred at room temperature for 1 hour, and pyridine-2-carbaldehyde (28.3 mg, 0.3 mmol) and NH ₄ Cl (79.5 mg, 1.49 mmol) were added. Was. After stirring under reflux conditions for 2 hours, the mixture was concentrated under reduced pressure. Purification by reverse phase HPLC gave compound 5.

  For the purification of the compound, reversed-phase high-performance liquid chromatography (reverse-phase-HPLC, general-purpose HPLC Prominence, Shimadzu Corporation (Kyoto)) was used. The column used was Nacalai Tesque, Inc. (5C18-AR300, 10 × 250 mm, (Kyoto)), and fractionated by a two-phase partitioning system containing 0.1% trifluoroacetic acid (TFA) and 0.1% TFA in acetonitrile.

The following shows the properties, yields, yields, and physical property data of each compound (IR: infrared absorption, ¹ H NMR: proton nuclear magnetic resonance, ¹³ C NMR: carbon nuclear magnetic resonance, HRMS: high-resolution mass spectrometry) Was described. IR spectra were measured using Thermo Nicolet iS5 (Thermo Fisher Scientific Inc. (USA)). ^{The 1} H NMR and ¹³ C NMR spectra were measured using the ECA600 type NMR spectrometer manufactured by JEOL Ltd. (Tokyo), the ECA500 type NMR spectrometer and the AL400 NMR type spectrometer manufactured by JEOL USA Inc. (USA) under the following conditions. Measured.

  The HRMS was measured by microTOF-QIII manufactured by BRUKER and the ionization method was Electrospray Ionization (Electron Spray Ionization: ESI) (ionization mode: positive mode, measurement temperature: 200 ° C, gas flow rate: 8.0). L / min).

Compound 1
13.7 mg of compound 1 as a white solid was obtained, and the yield was 22%. IR (neat, cm ^-1 ) 3397, 2368, 2326, 1683, 1675, 1436, 1203, 1136, 1064, 841; ¹ H NMR (500 MHz, CD ₃ OD, 25 ° C.) δ = 8.90 (d, J = 8.0 Hz, 1H), 8.85 (d, J = 3.6 Hz, 1H), 8.30 (d, J = 6.6 Hz, 1H), 8.08 (dd, J = 7.8, 7.8 Hz, 1H), 7.59-7.56 (m, 2H); ¹³ C NMR (125 MHz, CD ₃ OD, 25 ° C.) 162.6, 160.7, 150.9, 150.1, 149.1, 139.0, 135.9, 133.9, 126.6, 125.0, 107.8; HRMS (ESI) m / z calcd for C ₁₁ H ₉ N ₅ [M + H] ⁺ 212.0931, found 212.0929

Compound 2
10.2 mg of compound 2 as a brown solid was obtained, and the yield was 12%. IR (neat, cm ^-1 ) 3111, 1676, 1617, 1506, 1437, 1202, 1131, 800; ¹ H NMR (500 MHz, CD ₃ OD, 25 ° C.) δ = 9.30 (d, J = 8.0 Hz, 1H ), 8.87 (d, J = 5.0 Hz, 1H), 8.29 (d, J = 6.0 Hz, 1H), 8.10 (dd, J = 8.0, 8.0 Hz, 1H), 7.77-7.70 (m, 3H), 7.64 -7.59 (m, 3H), 7.32 (d, J = 6.0 Hz, 1H); ¹³ C NMR (125 MHz, CD ₃ OD, 25 ° C.) δ = 161.8, 161.6, 160.5, 150.8, 148.7, 139.1, 134.5, 134.2, 133.6, 132.0, 131.9, 128.3, 126.9, 126.8, 105.8; HRMS (ESI) m / z calcd for C ₁₇ H ₁₃ N ₅ [M + H] ⁺ 288.1244, found 288.1244

Compound 3
6.0 mg of compound 3 as a brown solid was obtained, and the yield was 5%. IR (neat, cm ^-1 ) 3198, 1734, 1684, 1653, 1558, 1507, 1419, 1229; ¹ H NMR (500 MHz, CD ₃ OD, 25 ° C.) δ = 9.14 (d, J = 8.0 Hz, 1H ), 8.83 (d, J = 4.5 Hz, 1H), 8.34 (d, J = 6.0 Hz, 1H), 8.05 (dd, J = 8.0, 7.5 Hz, 1H), 7.74 (d, J = 6.0 Hz, 1H) ), 7.56 (dd, J = 7.5, 4.5 Hz, 1H), 4.51 (dd, J = 8.5, 4.5 Hz, 1H), 4.31-4.27 (m, 2H), 3.71 (s, 3H), 1.98 (s, 3H), 1.96-1.84 (m, 3H), 1.81-1.75 (m, 1H); ¹³ C NMR (125 MHz, CD ₃ OD, 25 ° C.) δ = 173.7, 173.5, 158.2, 156.7, 150.1, 143.4, 143.1 , 140.3, 138.9, 138.3, 126.1, 124.5, 105.3, 53.2, 52.8, 42.9, 29.7, 26.0, 22.3; HRMS (ESI) m / z calcd for C ₁₉ H ₂₃ N ₆ O ₃ [M + H] ⁺ 383.1826, found 383.1828

Compound 4
2.1 mg of compound 4 as a brown solid was obtained, and the yield was 2%. IR (neat, cm ^-1 ) 3314, 2354, 1653, 1539, 1511, 1419, 1249, 1212; ¹ H NMR (500 MHz, CD ₃ OD, 25 ° C.) δ = 8.75 (d, J = 4.5 Hz, 1H ), 8.32 (d, J = 8.0 Hz, 1H), 8.24 (d, J = 5.5 Hz, 1H), 8.00 (dd, J = 8.0, 6.5 Hz, 1H), 7.59 (dd, J = 9.0, 5.0 Hz) , 2H), 7.48 (dd, J = 6.5, 4.5 Hz, 1H), 7.42 (dd, J = 9.0, 5.0 Hz, 2H), 7.02 (d, J = 5.5 Hz, 1H); ¹³ C NMR (125 MHz) , CD ₃ OD, 25 ° C) δ = 164.4 (q, J = 246.8 Hz), 158.0, 156.7, 150.2, 143.9 (q, J = 35.9 Hz), 140.1, 139.1, 138.3, 131.2, 130.6 (q, J = 9.5 Hz), 126.1, 124.6, 118.6, 118.4, 105.4; HRMS (ESI) m / z calcd for C ₁₇ H ₁₃ FN ₅ [M + H] ⁺ 306.1150, found 306.1151

Compound 5
21 mg of compound 5 as a yellow solid was obtained, and the yield was 37%. IR (neat, cm ^-1 ) 3104, 1684, 1653, 1559, 1517, 1457, 1254, 1201, 1130; ¹ H NMR (500 MHz, CD ₃ OD, 25 ° C.) δ = 9.25 (d, J = 8.0 Hz) , 1H), 8.85 (d, J = 4.5 Hz, 1H), 8.27 (d, J = 6.0 Hz, 1H), 8.08 (dd, J = 8.0, 8.0 Hz, 1H), 7.58 (dd, J = 8.0, 4.5 Hz, 1H), 7.52 (d, J = 9.0 Hz, 2H), 7.27-7.24 (m, 3H), 3.93 (s, 3H); ¹³ C NMR (125 MHz, CD ₃ OD, 25 ° C.) δ = 163.0, 162.8, 160.7, 150.8, 148.8, 139.0, 134.6, 134.2, 129.7, 126.8, 126.6, 125.6, 119.3, 117.0, 105.7, 56.3; HRMS (ESI) m / z calcd for C ₁₈ H ₁₆ N ₅ O [M + H] ⁺ 318.1349, found 318.1351

Compound 6
34.7 mg of compound 6 as a yellow solid was obtained, and the yield was 23%. IR (neat, cm ⁻¹ ) 3067, 1684, 1653, 1517, 1457, 1269, 1200, 1131, 1023, 799; ¹ H NMR (500 MHz, CD ₃ OD, 25 ° C.) δ = 9.18 (d, J = 8.0 Hz, 1H), 8.85 (d, J = 4.5 Hz, 1H), 8.27 (d, J = 6.5 Hz, 1H), 8.08 (dd, J = 8.0, 8.0 Hz, 1H), 7.58 (dd, J = 8.0, 4.5 Hz, 1H), 7.29 (d, J = 6.5 Hz, 1H), 7.24 (d, J = 8.0 Hz, 1H), 7.18 (d, J = 2.5 Hz, 1H), 7.26 (dd, J = 8.0, 2.5 Hz, 1 H), 3.96 (s, 3 H), 3.90 (s, 3 H); ¹³ C NMR (125 MHz, CD ₃ OD, 25 ° C.) δ = 160.4, 152.3, 152.1, 150.8, 149.8, 148.4, 139.3, 139.0, 135.2 (2C), 126.6 (2C), 126.0, 121.0, 113.7, 111.8, 105.8, 56.8, 56.7; HRMS (ESI) m / z calcd for C ₁₉ H ₁₈ N ₅ O ₂ [M + H] ⁺ 348.1455, found 348.1456

Compound 7
9.9 mg of compound 7 as a brown solid was obtained, and the yield was 10%. IR (neat, cm ^-1 ) 3236, 1734, 1684, 1559, 1521, 1507, 1204, 1132; ¹ H NMR (500 MHz, CD ₃ OD, 25 ° C.) δ = 9.24 (d, J = 8.0 Hz, 1H ), 8.86 (d, J = 4.5 Hz, 1H), 8.26 (d, J = 6.5 Hz, 1H), 8.08 (dd, J = 8.0, 8.0 Hz, 1H), 7.58 (dd, J = 8.0, 4.5 Hz , 1H), 7.44 (d, J = 9.0 Hz, 1H), 7.17 (d, J = 6.5 Hz, 1H), 6.88 (d, J = 2.5 Hz, 1H), 6.80 (dd, J = 9.0, 2.5 Hz) , 1H), 3.93 (s, 3H), 3.82 (s, 3H); ¹³ C NMR (125 MHz, CD ₃ OD, 25 ° C) δ = 164.7, 160.9, 157.9, 150.8, 149.0 (2C), 139.0, 134.6 , 134.1, 130.9, 126.8, 126.5, 113.6, 107.3, 106.1, 105.9, 101.1, 56.5, 56.4; HRMS (ESI) m / z calcd for C ₁₉ H ₁₈ N ₅ O ₂ [M + H] ⁺ 348.1455, found 348.1454

Compound 8
48.0 mg of a brown solid compound 8 was obtained, and the yield was 5%. IR (neat, cm- ¹ ) 3337, 2925, 1684, 1591, 1558, 1457, 1419, 1284, 1229, 1127, 1107, 801; ¹ H NMR (500 MHz, CD ₃ OD, 25 ° C) δ = 8.75 ( d, J = 5.0 Hz, 1H), 8.32 (d, J = 8.0 Hz, 1H), 8.24 (d, J = 5.0 Hz, 1H), 7.99 (dd, J = 8.0, 8.0 Hz, 1H), 7.47 ( dd, J = 8.0, 5.0 Hz, 1H), 7.11 (d, J = 5.0 Hz, 1H), 6.85 (s, 2H), 3.89 (s, 6H), 3.86 (s, 3H); ^13C NMR (125 MHz, CD ₃ OD, 25 ° C) δ = 157.9, 156.7, 155.8, 150.2, 144.1, 143.6, 140.9, 140.0, 139.1, 138.3, 130.6, 126.1, 124.6, 105.8, 105.7, 61.2, 56.9; HRMS (ESI) m / z calcd for C ₂₀ H ₂₀ N ₅ O ₃ [M + H] ⁺ 378.1561, found 378.1562

Compound 9
9.8 mg of compound 9 as a brown solid was obtained, and the yield was 11%. IR (neat, cm ^-1 ) 3116, 1675, 1559, 1517, 1280, 1132, 799; ¹ H NMR (500 MHz, CD ₃ OD, 25 ° C.) δ = 9.22 (d, J = 8.0 Hz, 1H), 8.85 (d, J = 4.5 Hz, 1H), 8.28 (d, J = 6.0 Hz, 1H), 8.07 (dd, J = 8.0, 8.0 Hz, 1H), 7.58 (dd, J = 8.0, 4.5 Hz, 1H ), 7.41 (ddd, J = 9.5, 2.5, 2.5 Hz, 2H), 7.27 (d, J = 6.0 Hz, 1H), 7.09 (ddd, J = 9.5, 2.5, 2.5 Hz, 2H); ^13C NMR ( 125 MHz, CD ₃ OD, 25 ° C) δ = 161.0, 160.6, 150.8, 148.9, 148.8, 139.0, 138.8, 134.8, 134.2, 129.7, 126.8, 126.5, 124.2, 118.3, 105.8; HRMS (ESI) m / z calcd for C ₁₇ H ₁₄ N ₅ O [M + H] ⁺ 304.1193, found 304.1193

Compound 10
8.4 mg of compound 10 as a brown solid was obtained, and the yield was 8%. IR (neat, cm ^-1 ) 3392, 2955, 1733, 1652, 1559, 1539, 1516, 1419, 1247, 1110; ¹ H NMR (500 MHz, CD ₃ OD, 25 ° C.) δ = 8.75 (d, J = 4.0 Hz, 1H), 8.35 (d, J = 8.0 Hz, 1H), 8.23 (d, J = 5.5 Hz, 1H), 7.94 (dd, J = 8.0, 8.0 Hz, 1H), 7.73 (d, J = 8.5 Hz, 1H), 7.49-7.46 (m, 3H), 7.04 (d, J = 5.5 Hz, 1H), 1.42 (s, 9H); ¹³ C NMR (125 MHz, CD ₃ OD, 25 ° C.) δ = 158.0, 156.5, 154.2, 150.2, 144.1, 143.4, 140.7, 139.1, 138.4, 132.3, 128.7, 127.6, 126.1, 124.7, 105.6, 35.8, 31.7; HRMS (ESI) m / z calculated for C ₂₁ H ₂₂ N ₅ [ M + H] ⁺ 344.1870, found 344.1874

Compound 11
9.6 mg of compound 11 as a brown solid was obtained, and the yield was 9%. IR (neat, cm ^-1 ) 2919, 2357, 1733, 1700, 1684, 1559, 1539; ¹ H NMR (500 MHz, CD ₃ OD, 25 ° C.) δ = 8.75 (d, J = 5.0 Hz, 1H), 8.35 (d, J = 8.0 Hz, 1H), 8.26 (d, J = 6.0 Hz, 1H), 8.02-7.97 (m, 3H), 7.79 (d, J = 8.0 Hz, 2H), 7.47 (dd, J = 7.5, 5.0 Hz, 1 H), 7.10 (d, J = 6.0 Hz, 1 H); ¹³ C NMR (125 MHz, CD ₃ OD, 25 ° C) δ = 157.5, 156.7, 150.2, 144.0, 143.5, 141.2, 139.3 , 138.7, 138.3, 132.3 (q, J = 33.4 Hz), 128.9, 128.8 (q, J = 3.6 Hz), 126.1, 125.3 (q, J = 269.4 Hz), 124.7, 105.4; HRMS (ESI) m / z calcd for C ₁₈ H ₁₃ F ₃ N ₅ [M + H] ⁺ 356.1118, found 356.1118

Compound 12
3.2 mg of compound 12 was obtained, and the yield was 25%. HRMS (ESI) m / z calcd for C ₁₇ H ₁₁ N ₇ O ₄ [M + H] ⁺ 378.0945, found 378.0945

Compound 13
7.4 mg of compound 13 as a brown solid was obtained, and the yield was 7%. IR (neat, cm ⁻¹ ) 3324, 2360, 1684, 1653, 1559, 1553, 1203, 1133, 799; ¹ H NMR (500 MHz, CD ₃ OD, 25 ° C.) δ = 9.37 (d, J = 8.0 Hz) , 1H), 8.89 (d, J = 5.0 Hz, 1H), 8.28 (dd, J = 7.0, 2.5 Hz, 1H), 8.25 (d, J = 6.5 Hz, 1H), 8.17-8.12 (m, 2H) , 7.82-7.78 (m, 2H), 7.68 (dd, J = 7.0, 7.0 Hz, 1H), 7.64-7.59 (m, 2H), 7.38 (d, J = 8.0 Hz, 1H), 7.06 (d, J ¹³ C NMR (125 MHz, CD ₃ OD, 25 ° C.) δ = 161.1, 150.9, 149.4, 148.7, 140.0, 139.1, 136.5, 134.5, 134.3, 132.9, 130.8, 130.2, 129.5, 129.3 , 128.6, 128.3, 127.2, 127.0, 126.9, 122.5, 105.9; HRMS (ESI) m / z calcd for C ₂₁ H ₁₆ N ₅ [M + H] ⁺ 338.1400, found 338.1402

Compound 14
6.9 mg of compound 14 as a brown solid was obtained, and the yield was 7%. IR (neat, cm ⁻¹ ) 3239, 2360, 1734, 1684, 1653, 1559, 1540, 1203, 1135; ¹ H NMR (500 MHz, CD ₃ OD, 25 ° C.) δ = 9.30 (d, J = 8.0 Hz) , 1H), 8.87 (d, J = 5.0 Hz, 1H), 8.28 (d, J = 5.5 Hz, 1H), 8.10 (dd, J = 8.0, 8.0 Hz, 1H), 7.88 (d, J = 8.5 Hz , 1H), 7.65 (s, 1H), 7.60 (dd, J = 8.0, 5.0 Hz, 1H), 7.50 (d, J = 3.0 Hz, 1H), 7.30 (d, J = 5.5 Hz, 1H), 7.16 (Dd, J = 8.5, 1.5 Hz, 1H), 6.66 (d, J = 3.0 Hz, 1H); ¹³ C NMR (125 MHz, CD ₃ OD, 25 ° C.) δ = 161.0, 150.8, 150.7, 149.2, 149.0 , 139.1, 137.7, 134.6, 134.2, 131.2, 128.9, 126.8, 126.7, 126.3, 123.4, 118.3, 111.4, 105.9, 103.0; HRMS (ESI) m / z calcd for C ₁₉ H ₁₅ N ₆ [M + H] ⁺ 327.1353, found 327.1352

Compound 15
9.1 mg of compound 15 was obtained, and the yield was 9%. HRMS (ESI) m / z calcd for C ₂₁ H ₁₅ N ₅ + H [M + H] ⁺ 338.1400, found 338.1401.

Compound 16
7.0 mg of a brown solid compound 16 was obtained, and the yield was 6%. IR (neat, cm ^-1 ) 3311, 2357, 1774, 1653, 1559, 1539, 1457, 1205, 1132; ¹ H NMR (500 MHz, CD ₃ OD, 25 ° C.) δ = 9.56 (d, J = 8.0 Hz) , 1H), 8.38-8.35 (m, 2H), 8.03 (d, J = 8.5 Hz, 1H), 7.39 (d, J = 5.5 Hz, 1H), 7.25 (d, J = 8.5 Hz, 1H), 7.20 (D, J = 2.5 Hz, 1H), 7.16 (dd, J = 8.5, 2.5 Hz, 1H), 3.96 (s, 3H), 3.90 (s, 3H); ¹³ C NMR (125 MHz, CD ₃ OD, 25 ° C) δ = 161.2, 152.4, 152.1, 149.7, 149.5, 149.1 (q, J = 35.8 Hz), 141.3, 140.3, 135.3, 132.2, 129.5, 125.6, 123.1, 122.7 (q, J = 271.8 Hz), 121.0,113.7,111.7,106.2,56.7 (2C); HRMS (ESI) m / z calcd for C 20 H 17 F 3 N 5 O 2 [M + H] + 416.1329, found 416.1331

Compound 17
2.7 mg of compound 17 as a brown solid was obtained, and the yield was 13%. IR (neat, cm ^-1 ) 3347, 2360, 1684, 1653, 1559, 1539, 1507, 1204, 1136; ¹ H NMR (500 MHz, CD ₃ OD, 25 ° C) δ = 8.06 (d, J = 6.5 Hz , 1H), 7.28 (s, 1H), 7.22 (d, J = 8.5 Hz, 1H), 7.16-7.15 (m, 2H), 7.11 (dd, J = 8.5, 2.5 Hz, 1H), 3.95 (s, 3H), 3.89 (s, 3H); ¹³ C NMR (125 MHz, CD ₃ OD, 25 ° C.) δ = 160.5, 152.3, 152.0, 147.3, 137.1, 132.9, 129.3, 125.8, 125.7, 120.8, 115.4, 113.7, 111.6, 108.2, 104.2, 102.6, 56.7 (2C); HRMS (ESI) m / z calcd for C ₁₈ H ₁₆ Br ₂ N ₅ O ₂ [M + H] ⁺ 491.9665, found 491.9665

Compound 18
4.5 mg of Compound 18 was obtained as a brown solid, and the yield was 3%. IR (neat, cm ⁻¹ ) 3281, 2360, 1734, 1700, 1684, 1653, 1559, 1506, 1457, 1204, 1126; ¹ H NMR (600 MHz, CD ₃ OD, 25 ° C.) δ = 8.03 (d, J = 6.0 Hz, 1H), 7.37 (d, J = 9.0 Hz, 1H), 7.27 (s, 1H), 7.10 (d, J = 6.0 Hz, 1H), 7.07 (d, J = 9.0 Hz, 1H) ¹³ C NMR (150 MHz, CD ₃ OD, 25 ° C.) δ = 160.9, 160.6, 147.4, 137.0, 132.9, 129.6, 125.7, 124.4, 118.3, 115.4, 108.2, 106.5, 104.1, 102.6; HRMS (ESI) m / z calcd for C ₁₆ H ₁₂ Br ₂ N ₅ O [M + H] ⁺ 447.9403, found 447.9404

  The following methods were used to evaluate the effects of the compound of the present invention on neuronal differentiation-inducing activity, cytotoxicity, Dyrk1A kinase inhibitory activity, and astrocytes. In these evaluations, a β-carboline derivative was reported to have a strong neuronal cell differentiation promoting action via Dyrk1A kinase inhibitory activity. Harmin (Mazur-Kolecka B et al., J. Neurosci. Res. 2012, 90, 999-1010) as a positive control. The statistical test was performed using Dunn's multiple comparison test using the statistical analysis software JMP (SAS Institute Inc., USA) with the test compound non-added group as a control group, and p <0.05 was determined to be significant ( post-hoc test).

<Nerve differentiation inducing activity of compounds 1 to 22>
Neuronal cell differentiation-inducing activity (1) Method for measuring neuronal cell differentiation-inducing activity The neural cell differentiation-inducing activity was measured using mouse ES cell-derived neural stem cells. The cryopreserved neural stem cells were seeded at 1 × 10 ⁴ cells / well in a 96-well plate. At this time, as a culture solution, 2% -B-27 (gibco), 1% penicillin / streptomycin (Gibco, Thermo Fisher Scientific Inc. (USA)), 20 ng / mL rhFGF-2 (R & D Systems Inc. (USA) ), Using a Neurobasal Medium (Gibco) containing 20 ng / mL rh EGF (R & D Systems Inc.) in a CO ₂ incubator at 37 ° C with 5% CO ₂ . Two days later, the medium was replaced, and the next day, the medium was changed to a Neurobasal Medium containing 2% -B-27, 1% penicillin / streptomycin, and at that time, at a concentration of 0.001, 0.01, 0.1, and 1 μM. The compound of the present invention was added. Further, two days later, the medium was replaced, and the next day, immunostaining described below was performed.

  After removing the culture medium from each well and washing twice with PBS (Takara Bio Inc. (Shiga)), add 4% paraformaldehyde (Wako Pure Chemical Industries, Ltd. (Osaka)) and let stand at 4 ° C for 30 minutes. Placed and fixed. After removing paraformaldehyde, the plate was washed three times with PBS, and PBS containing 0.2% Triton X-100 (Alfa Aesar, Johnson Massey (UK)) and 5% Skim milk (Wako Pure Chemical Industries, Ltd.) was added, followed by 4 ° C. For 30 minutes. A PBS solution containing 5% skim milk containing the primary antibody was added, and the mixture was allowed to stand at 4 ° C. overnight to perform an antigen-antibody reaction. Rabbit monoclonal anti-MAP-2 antibody (Merck Millipore (USA)) was diluted 200-fold and used as the primary antibody.

  The PBS solution containing 5% Skim milk containing the primary antibody was removed, and the plate was washed three times with a PBS solution containing 5% Skim milk. After removing the solution, a PBS solution containing 5% skim milk containing a fluorescently labeled secondary antibody was added, and the mixture was allowed to stand at room temperature for 30 minutes. As a secondary antibody, a PE-labeled donkey polyclonal anti-rabbit antibody (Abcam (UK)) was used at a 100-fold dilution.

  After removing the PBS solution containing 5% Skim milk containing the secondary antibody and washing three times with a PBS solution containing 0.2% Triton X-100, increase the size of Hoechst33342 (Dojindo Research Laboratories, Kumamoto) by 1000 times. The solution was added at a dilution, and a fluorescence microscope (Olympus Industries, Ltd., Tokyo) was used for observation.

  The number of Hoechst33342-positive cells and the number of MAP-2 positive cells in each well were counted, and (number of MAP-2 positive cells) / (number of Hoechst33342-positive cells) was calculated as a neural cell differentiation rate.

(2) Results of measurement of nerve cell differentiation-inducing activity Compared to the control group (test compound-free group), Compounds 1 to 18 showed a regulatory effect on nerve cell differentiation in a concentration range of 0.001 to 1 μM. In many of them, the differentiation rate increased (individual experimental results: not shown).

<Concentration-dependent test on the neuronal differentiation-inducing activity of compounds 1 to 3>
(1) Experimental Method The concentration dependency of the compounds 1 to 3 on the nerve cell differentiation promoting action was evaluated basically in the same manner as in Example 2. The concentration of the compound was evaluated at four concentrations of 0.001, 0.01, 0.1 and 1.0 μM.

(2) Experimental Results Compound 1 having a basic skeleton exhibited a promoting activity at 0.1 and 1 μM, and showed concentration dependency. Compounds 2 and 3 exhibited promoting activity at 0.01, 0.1 and 1 μM and showed concentration dependence (FIG. 1).

<Measurement of cytotoxicity of Compounds 1 to 18>
(1) Method of measuring cytotoxicity For the measurement of cytotoxicity, mouse ES cell-derived neural stem cells were used. The cryopreserved neural stem cells were seeded at 1 × 10 ⁴ cells / well in a 96-well plate. As this time cultures, 2% -B-27,1% penicillin / streptomycin, using Neurobasal Medium containing 20 ng / mL rhFGF-2,20 ng / mL rhEGF, 5% CO 2, 37 ℃ of CO ₂ The cells were cultured in an incubator. The next day, the compounds of the present invention were added at the concentrations of 0.01, 0.1, 1, and 10 μM, and the culture was continued for another 3 days.

  Thereafter, 25 μL of a 1 mg / mL MTT (Invitrogen) solution was added to 100 μL / well of the medium and incubated for 3 hours. After removing the medium, 150 μL of DMSO (Invitrogen) was added, and a microplate reader (PerkinElmer) was added. , Inc. (USA)), cell viability was calculated from the ratio of the absorbance of the control to the absorbance at the time of compound addition, and the cytotoxicity was evaluated.

(2) Measurement results of cytotoxicity The results are shown in Table 2 below. All of the compounds 1 to 18 showed no or only weak cytotoxicity, and had an IC ₅₀ of 5 μM or more. On the other hand, IC ₅₀ of harmine used as a comparative example was 4.0 [mu] M.

<Evaluation of Dyrk1A kinase inhibitory activity>
(1) Experimental method The inhibitory activity of a compound on Dyrk1A kinase was entrusted to Carna Biosciences Co., Ltd. (Kobe) and performed using the following off-chip mobility shift assay (MSA).

  The full-length human Dyrk1A protein expressing the GST fusion protein at the N-terminus was expressed using a baculovirus expression system, and purified using glutathione sepharose chromatography. The test compound was dissolved in DMSO at a concentration of 25 mM, and diluted to 1 mM with assay buffer (20 mM HEPES [pH 7.5], 0.01% Triton X-100, 2 mM DTT: dithiothreitol). Thereafter, 5 μL of the solution was mixed with 5 μL of ATP / substrate / metal solution (16 μM ATP, 1000 nM substrate peptide, 5 mM Mg), and 10 μL of 200 μg / mL Dyrk1A kinase solution was added. The mixture was reacted at room temperature for 1 hour, and the reaction was terminated using 60 μL of a reaction stop buffer (QuickScout Screening Assist MSA; Carna Biosciences). Phosphorylated peptides and non-phosphorylated peptides were separated and quantified using LabChip system (PerkinElmer, Inc.). The inhibition rate was calculated from the ratio of phosphorylated peptide.

Experimental Results The pyridine derivative (compound 1) and several compounds showed inhibitory activity, but the novel compounds (compounds 2 to 4) which showed remarkable inducing activity did not show inhibitory activity, and showed neuronal differentiation inducing activity and Dyrk1A kinase. No correlation was found with inhibitory activity (Table 3).

<Measurement of Astrocyte Differentiation Inducing Activity of Compounds 1 to 4>
(1) Experimental method Measurement of astrocyte differentiation-inducing activity used mouse ES cell-derived neural stem cells. The cryopreserved neural stem cells were seeded at 1 × 10 ⁴ cells / well in a 96-well plate. At this time, as a culture solution, 2% -B-27 (Gibco), 1% penicillin / streptomycin (Gibco), 20 ng / mL rhFGF-2 (R & D Systems Inc.), 20 ng / mL rh EGF (R & D Systems Inc.) Using a Neurobasal Medium (Gibco) containing (.), The cells were cultured in a 5% CO ₂ , 37 ° C. CO ₂ incubator. Two days later, the medium was exchanged. On the next day, the medium was changed to Astrocyte Conditioned Medium (ACM, Sumitomo Bakelite Co., Ltd. (Tokyo)). Compound was added. Further, two days later, the medium was replaced, and the next day, immunostaining described below was performed.

  After removing the medium from each well and washing twice with PBS (Takara Bio Inc.), 4% paraformaldehyde (Wako Pure Chemical Industries, Ltd.) was added, and the mixture was allowed to stand at 4 ° C. for 30 minutes and fixed. After removing paraformaldehyde, the plate was washed three times with PBS, and PBS containing 0.2% Triton X-100 (Alfa Aesar, Johnson Massey) and 5% Skim milk (Wako Pure Chemical Industries, Ltd.) was added. Blocking was performed. A PBS solution containing 5% skim milk containing a primary antibody was added, and the mixture was allowed to stand at 4 ° C. overnight to exhibit an antigen-antibody reaction. As a primary antibody, a mouse monoclonal anti-GFAP antibody (Merck Millipore) was diluted 500-fold and used.

  The PBS solution containing 5% Skim milk containing the primary antibody was removed, and the plate was washed three times with a PBS solution containing 5% Skim milk. After removing the solution, a PBS solution containing 5% skim milk containing a fluorescently labeled secondary antibody was added, and the mixture was allowed to stand at room temperature for 30 minutes. As the secondary antibody, a Chromeo488-labeled goat polyclonal anti-mouse antibody (Active motif) was used at a dilution of 1: 200.

  After removing the PBS solution containing 5% Skim milk containing the secondary antibody and washing three times with a PBS solution containing 0.2% Triton X-100, increase the size of Hoechst33342 (Dojindo Laboratories, Kumamoto) by 1000 times. The solution was added at a dilution, and a fluorescence microscope (Olympus Industries, Ltd., Tokyo) was used for observation.

  The number of Hoechst33342-positive cells and the number of GFAP-positive cells in each well were counted, and (the number of GFAP-positive cells) / (the number of Hoechst33342-positive cells) was calculated as the astrocyte differentiation rate.

Experimental Results Compounds 1 to 3 did not show any effect on astrocyte differentiation induction. Harmin, on the other hand, showed a statistically significant inhibitory effect on astrocyte differentiation (FIG. 2).

  The compound of the present invention has an excellent neuronal cell differentiation promoting action, a pharmaceutical composition for preventing or treating a neurodegenerative disease, an agent for promoting differentiation of pluripotent stem cells into nerve cells, and an agent for regulating the differentiation of cultured cells It is useful for use in screening methods.

Claims (10)

A compound represented by the following formula 1, or a pharmaceutically acceptable salt or solvate thereof ;
R _1, R ₂ 及 beauty X is each independently, represent H,
R ₃ is selected from optionally substituted phenyl, optionally substituted naphthyl, optionally substituted ethyl, optionally substituted indole,
m is 0 ,
The ring A is represented by one ring selected from the following formulas 2 to 4;
Y, Y ′, Y ″ and Y ′ ″ each independently may be the same or different, and each independently represents H, halogen, or optionally substituted alkyl;
p is 0 or 1,
q is 0,
r is 0,
s represents 0 or 1. The compound according to claim 1, wherein the compound is represented by the following Formulas 6 to 22, or a pharmaceutically acceptable salt or solvate thereof .



A method for producing an imidazopyridine amine compound represented by the following reaction formula 1,
(i) Using the substituted amine derivative (1) as a raw material, guanidination (3) by reaction with a cyanamide compound (2),
(ii) Next, reacting with a conjugated dialdehyde compound (4) to form an imidazole compound (5),
(iii) Next, an imine is formed with the ammonium salt (6) and the aldehyde compound (7), and an imidazopyridine ring is formed by advancing the subsequent aza electron cyclization reaction to produce an imidazopyridine amine compound (8). A manufacturing method characterized in that:
However, R ₁ , R ₂ , R ₃ and ring A follow the definition in claim 1. The method according to claim 3 , wherein the manufacturing method is a one-pot method. A medicament comprising the compound according to claim 1 or 2 or a pharmaceutically acceptable salt or solvate thereof as an active ingredient. A pharmaceutical composition for preventing or treating a neurodegenerative disease, comprising the compound according to claim 1 or a pharmaceutically acceptable salt or solvate thereof as an active ingredient. The pharmaceutical composition according to claim 6 , wherein the neurodegenerative disease is selected from Alzheimer's disease, mild cognitive impairment, Parkinson's disease, Lewy body dementia, and multiple sclerosis. A nerve for promoting differentiation of a pluripotent stem cell into a nerve cell, comprising the compound according to claim 1 or a pharmaceutically acceptable salt or solvate thereof as an active ingredient. Cell differentiation promoter. A compound that regulates differentiation into nerve cells, comprising a step of adding the compound according to claim 1 or a pharmaceutically acceptable salt or solvate thereof to cultured cells in vitro. A method for screening. The method according to claim 9 , wherein the cultured cells are pluripotent stem cells.