WO2013027694A1

WO2013027694A1 - Molecular imaging probes for diagnosing conformation disease

Info

Publication number: WO2013027694A1
Application number: PCT/JP2012/070978
Authority: WO
Inventors: 英郎佐治; 小野　正博; 孟超崔
Original assignee: 国立大学法人京都大学
Priority date: 2011-08-24
Filing date: 2012-08-20
Publication date: 2013-02-28
Also published as: JPWO2013027694A1

Abstract

The purpose of the present invention is to provide a near infrared light-emitting compound which has an ability to highly specifically binding to Aβ, a high permeability through the blood-brain barrier, an ability to quickly disappear from the brain and a high biopermeability. Provided are a compound having a benzoxazole skeleton and an aromatic compound having a dimethylamino group, a dicyanovinyl group, etc. In formula (I), either R1 or R2 is an electron donating group and the other is an electron withdrawing group; X and Y are the same or different and each represents a carbon atom, a nitrogen atom, an oxygen atom or a sulfur atom; and m is an integer of 0 to 5.

Description

Molecular imaging probe for conformation disease diagnosis

The present invention relates to a near-infrared fluorescent compound and a benzoxazole derivative. These compounds can be used for diagnosis of conformational diseases.

With the recent rapid aging of society, an increase in dementia diseases such as Alzheimer's disease (AD) has become one of the major social problems. Currently, there are Hasegawa's method, ADAS, and MMSE as clinical diagnostic methods for AD, and all of them generally use a method for quantitatively evaluating the decline in cognitive function of an individual suspected of having AD. Other diagnostic imaging methods (MRI, CT, etc.) are used as supplementary methods, but these diagnostic methods are not sufficient for definitive diagnosis of AD. For definitive diagnosis, biopsy of the brain before birth, postmortem brain In histopathological examination, it is necessary to confirm the appearance of senile plaques and neurofibrils. Therefore, with current diagnostic methods, it is difficult to diagnose AD at an early stage before extensive brain damage occurs. To date, there have been several reports as biological diagnostic markers for AD, but clinically practical ones have not yet been developed. Under such circumstances, there is a high social demand for early diagnosis of AD, and its rapid development is strongly desired.

Senile plaque is the most characteristic brain lesion of AD, and its main component is amyloid β protein (Aβ) having a β sheet structure. Imaging of senile plaques from outside the body is thought to lead to the establishment of an effective diagnostic method for AD, but imaging requires an amyloid imaging probe that specifically binds to Aβ. Known amyloid imaging probes include radioactive probes used in PET and SPECT (Patent Document 1) and fluorescent probes using donor-acceptor type fluorescent molecules (Patent Document 2).

Special table 2008-546804 International Publication 2010/125907

A compound used as an amyloid imaging probe is required to have high binding specificity for Aβ, high blood-brain barrier permeability, and rapid disappearance from the brain. Furthermore, the compound used as a fluorescent probe is also required to emit near-infrared light excellent in biological penetration.

The object of the present invention is to provide a novel compound having the above properties.

As a result of intensive studies to solve the above problems, the present inventor has found that a compound having a benzoxazole skeleton has high binding specificity for Aβ, high permeability of the blood brain barrier, and rapid disappearance from the brain. In addition, the present inventors have found that an aromatic ring compound having a dimethylamino group and a dicyanovinyl group emits near-infrared fluorescence, and completed the present invention.

That is, the present invention provides the following (1) to (6).
(1) General formula (I), general formula (II), or general formula (IV)

[In the formula, any _one of R ₁ and R ₂ is an electron donating group, the other is an electron withdrawing group, X and Y are the same or different, and are a carbon atom, a nitrogen atom, an oxygen atom or a sulfur atom. And Z is an oxygen atom or a sulfur atom, and m represents an integer of 0 to 5. ]
Or a pharmaceutically acceptable salt thereof.
(2) The electron donating group is a hydroxyl group, methoxy group, methyl group, amino group, methylamino group, dimethylamino group, methylaminophenyl group, or dimethylaminophenyl group, and the electron withdrawing group is a nitro group, a nitrovinyl group, The compound or a pharmaceutically acceptable salt thereof according to (1), which is a cyano group, a dicyanovinyl group, a tricyanovinyl group, or a formyl group.
(3) General formula (III)

[Wherein R ₃ represents a hydroxy group, a C _1-3 alkoxy group, a C _1-3 alkoxy group containing a radioactive carbon atom, a halogen atom, a radioactive halogen atom, a chelate moiety bonded to a radioactive metal, or — (CH ₂ CH ₂ O) _i -A [wherein _i represents an integer of 1 to 10, and A represents a chelate moiety bonded to a halogen atom, a radioactive halogen atom, or a radioactive metal. R ₄ represents a hydroxy group, a methoxy group, — (CH ₂ CH ₂ O) _j —OH [wherein _j represents an integer of 1 to 10. A group represented by the formula: — (CH ₂ CH ₂ O) _k —F [wherein, k represents an integer of 1 to 10. Or —NRaRb [wherein Ra and Rb each independently represents a hydrogen atom or a C _1-3 alkyl group. And n represents an integer of 0 to 5. ]
Or a pharmaceutically acceptable salt thereof.
(4) The compound or pharmaceutically acceptable salt thereof according to (3), wherein the radioactive halogen atom is ¹⁸ F, ¹²³ I, ¹²⁴ I or ¹²⁵ I.
(5) The compound or pharmaceutically acceptable salt thereof according to (3), wherein the chelate moiety bound to the radioactive metal is a chelate moiety that binds to ^99m Tc or ⁶⁸ Ga.
(6) A composition for diagnosing conformation disease, comprising the compound according to any one of (1) to (5).

Since the compound of the present invention has a high binding specificity for Aβ, it is useful for early and accurate diagnosis of conformational diseases such as AD.

The synthesis route of the near-infrared fluorescent compound of Example 1. The fluorescence spectrum of the near-infrared fluorescent compound of Example 1. The upper curve was measured in the presence of Aβ, and the lower curve was measured in the absence of Aβ. The figure which shows the result of the saturation binding assay to the A (beta) 42 aggregate of the near-infrared fluorescent compound of Example 1. FIG. Fluorescent staining of a Tg model mouse brain tissue section using the near-infrared fluorescent compound of Example 1. Fluorescence imaging of Tg model mice using KR-5. Fluorescent staining of Tg model mouse brain tissue sections using KR-5. The synthesis route of the near-infrared fluorescent compound of Example 2 (hereinafter sometimes referred to as “DANIR”). Fluorescence spectra of near-infrared fluorescent compounds of Example 2 (A and B, DANIR-1; ＤC and D, DANIR-2; E and F, DANIR-3; G and H, DANIR-4; left, EX right, EM). When the upper curve is measured with the addition of Aβ1-42 fibers, and the lower curve is measured in PBS. Fluorescent staining of DANIR 2-4 on a section of brain tissue from AD human (93 years old,) female). DANIR-2 (A, Tx-red); DANIR-3 (C, Cy5); DANIR-4 (E, Cy5.5). A, C, E: Aβ plaque; B, D, F: NFTs (neurofibrillary tangles). Fluorescent staining of DANIR 1-4 (A, C, E, G) of brain tissue sections obtained from Tg model mouse (C57-APP / PS1, 12 months old, male). Labeled plaques were confirmed by staining of adjacent sections with thioflavin S (B, D, F, H). The figure which shows the result of the binding assay using aggregation A (beta) _1-42 in a solution. The figure which shows the result of the saturation binding assay of each DANIR to A (beta) _1-42 aggregate. Brain fluorescence imaging of DANIR-3 in vivo in nude mice. Brain fluorescence imaging of DANIR-3 in APP vivo and wild-type mice in vivo. DANIR-3 time intensity curves in APP and wild type mice. [ ¹⁸ F] Compound 7 synthesis route. The numbers in the figure correspond to the compound numbers in Example 3. Reagents and conditions: (a) Pd / C, MeOH, rt; (b) 4-methylaminobenzoic acid, PPA, 180 ° C; (c) BBr ₃ (1 M in CH ₂ Cl ₂ ), CH ₂ Cl ₂ , -78 ℃ - rt; (d) 2- [2- (2- chloroethoxy) ethoxy] _{_{ethanol, K 2 CO 3, DMF,}} 110 ℃; (e) TBMSCl, _{_{imidazole, CH 2 Cl 2, rt;}} (f ) DAST, CH ₂ Cl ₂ , -78 ℃-rt; (g) (Boc) ₂ O, THF, reflux; (h) TBAF (1 M in THF), THF, rt; (i) MsCl, Et ₃ N , CH ₂ Cl ₂ , rt; (j) ¹⁸ F-, K ₂ CO ₃ , Kryptofix-2.2.2, acetonitrile, 120 ° C .; (k) HCl (1 M), 120 ° C. Synthesis route for [ ¹⁸ F] compound 15. The numbers in the figure correspond to the compound numbers in Example 3. Reagents and conditions: (a) Pd / C, MeOH, rt; (c) BBr ₃ (1 M in CH ₂ Cl ₂ ), CH ₂ Cl ₂ , -78 ° C- rt; (d) 2- [2- ( 2-chloroethoxy) ethoxy] ethanol, K ₂ CO ₃ , DMF, 110 ° C; (l) 4- (dimethylamido) benzoic acid, PPA, 180 ° C; (m) TsCl, pyridine, rt; (n) TBAF ( 1 M in THF), THF, reflux. HPLC profiles of Compound 7 (left) and [ ¹⁸ F] Compound 7 (right). HPLC conditions: Cosmosil C18 column (Nakalai Tesque, 5C ₁₈ -AR-II, 4.6 mm × 150 mm), CH ₃ CN / H ₂ O = 50/50, 1 mL / min, UV, 254 nm. HPLC profiles of Compound 15 (left) and [ ¹⁸ F] Compound 15 (right). HPLC conditions: Cosmosil C18 column (Nakalai Tesque, 5C ₁₈ -AR-II, 4.6 mm × 150 mm), CH ₃ CN / H ₂ O = 60/40, 1 mL / min, UV, 254 nm. Inhibition curve for binding of [ ¹²⁵ I] IMPY to Aβ1-42 aggregates. Mouse brain slice (A: Tg mouse, C57-APP / PS1, 12 months old, male; C: Wild-type, C57, 12 months old, male) and human brain slice (E: AD, 93 years old, female; G: Normal, 71 years old, male) [ ¹⁸ F] Compound 7 in vitro autoradiography. The presence and distribution of plaques in the sections were confirmed by immunohistochemical staining using thioflavin S (B, D) and monoclonal Aβ antibody (F, H). Mouse brain slice (A: Tg mouse, C57-APP / PS1, 12 months old, male; C: Wild-type, C57, 12 months old, male) and human brain slice (E: AD, 93 years old, female; G: Normal, 71 years old, male) [ ¹⁸ F] Compound 15 in vitro autoradiography. The presence and distribution of plaques in the sections were confirmed by immunohistochemical staining using thioflavin S (B, D) and monoclonal Aβ antibody (F, H). Ex vivo autoradiography of [ ¹⁸ F] Compound 7 (top) and [ ¹⁸ F] Compound 15 (bottom) using 18 month old transgenic mice (APPswTg2576, C57BL6) and wild-type cotton roll (C57BL6, 18 month). Plaque was also confirmed by staining the same section with thioflavin S. In vivo micro PET study of [ ¹⁸ F] Compound 7 using 31 month old transgenic mice (APPswTg2576, C57BL6) (right) and wild type control (C57BL6, 31 month) (left). [ ¹⁸ F] 7 time radioactivity curves in the brains of transgenic mice (APPswTg2576, C57BL6) (right) and wild type controls (C57BL6, 31 month) (left). The synthesis route of the near-infrared fluorescent compound of Example 4. The numbers in the figure correspond to the compound numbers in Example 4. Reagents: (a) dimethylamine, H ₂ O; (b) malononitrile, pyridine, 2-propanol; (c) (1,3-dioxalan-2-ylmethyl) triphenylphosphonium bromide, NaH, 18-crown-6, THF. Fluorescent staining of brain sections of Tg2576 mice. A, C, E, G, I, and K are images stained with DTA-0, DTA-1, DFA-1, DTA-2, DFA-2, and DTA-3, respectively. B, D, F, H, J, and L are images obtained by staining adjacent sections of A, C, E, G, I, and K with thioflavin S, respectively.

Hereinafter, the present invention will be described in detail.
(A) Compound represented by general formula (I) R ₁ and R ₂ in general formula (I) may be either one of an electron donating group and the other an electron withdrawing group, but R ₁ is an electron In the donor group, R ₂ is preferably an electron withdrawing group.

R ₁ in the general formula (I) may be in any position on the heterocyclic ring, but is preferably in the 6-position (position assuming that the heterocyclic ring is benzothiazole).

The electron donating group in formula (I) is preferably a hydroxyl group, a methoxy group, a methyl group, an amino group, a methylamino group, a dimethylamino group, a methylaminophenyl group, or a dimethylaminophenyl group, and more preferably Is a dimethylamino group.

The electron withdrawing group in the general formula (I) is preferably a nitro group, a nitrovinyl group, a cyano group, a dicyanovinyl group, a tricyanovinyl group, or a formyl group, and more preferably a dicyanovinyl group.

X in the general formula (I) is preferably nitrogen.

Y in the general formula (I) is preferably sulfur.

M in the general formula (I) may be an integer of 0 to 5, and is preferably 1 or 2. In addition, when m is 0, it means that R ₂ is directly bonded to the heterocyclic ring.

Typical compounds among the compounds represented by the general formula (I) are shown in the following table. In the table, “Me” represents a methyl group, “Dicyano” represents a dicyanovinyl group, “Tricyano” represents a tricyanovinyl group, and “6-” represents a 6-position (complex of general formula (I)). It represents that the ring is a group at the position (assuming that it is benzothiazole).

Among the above compounds, preferred compounds include I-23 (KR-5), I-35 (KR-7), and I-47 (KR-9).

The compound represented by the general formula (I) can be synthesized by bonding an electron donating group and an electron withdrawing group directly or via an unsaturated carbon chain to a heterocyclic compound such as benzothiazole.
(B) the general formula (II) R ₁ and R ₂ in the compound the general formula (II) represented by the in either one electron donating group, but the other may be an electron withdrawing group, R ₁ is an electron In the donor group, R ₂ is preferably an electron withdrawing group.

R ₁ in the general formula (II) may be in any position on the benzene ring, but is preferably in the para position.

The electron donating group in the general formula (II) is preferably a hydroxyl group, a methoxy group, a methyl group, an amino group, a methylamino group, a dimethylamino group, a methylaminophenyl group, or a dimethylaminophenyl group, and more preferably Is a dimethylamino group.

The electron withdrawing group in the general formula (II) is preferably a nitro group, a nitrovinyl group, a cyano group, a dicyanovinyl group, a tricyanovinyl group, or a formyl group, and more preferably a dicyanovinyl group.

M in the general formula (II) may be an integer of 0 to 5, but is preferably 2 or 3. In addition, when m is 0, it means that R ₂ is directly bonded to the benzene ring.

Typical compounds among the compounds represented by the general formula (II) are shown in the following table. In the table, “Me” represents a methyl group, “Dicyano” represents a dicyanovinyl group, “Tricyano” represents a tricyanovinyl group, and “p-” represents a para-position group.

Among the above compounds, preferred compounds include II-11 (DANIR-1), II-23 (DANIR-2), II-35 (DANIR-3), and II-47 (DANIR-4).

The compound represented by the general formula (II) can be synthesized by bonding an electron donating group and an electron withdrawing group to a compound having a benzene ring directly or via an unsaturated carbon chain.
(C) Compound represented by the general formula (III) In the general formula (III), the “C _1-3 alkoxy group” is, for example, a methoxy group, an ethoxy group, an n-propoxy group, or an isopropoxy group. .

In the general formula (III), the “C _1-3 alkyl group” is, for example, a methyl group, an ethyl group, an n-propyl group, or an isopropyl group.

In the general formula (III), the “halogen atom” is, for example, F, Cl, Br, or I.

In the general formula (III), the “radiohalogen atom” is, for example, ¹⁸ F, ¹³¹ I, ¹²⁵ I, ¹²³ I, ¹²⁴ I, ⁷⁷ Br, or ⁷⁶ Br.

In the general formula (III), the “radioactive carbon atom” is, for example, ¹¹ C.

In the general formula (III), the “radioactive metal” is, for example, ^99m Tc, ⁶² Cu, ⁶⁷ Ga, or ⁶⁸ Ga. As these radioactive metals, ^99m Tc and ⁶⁸ Ga are suitable.

In the general formula (III), “a chelate moiety bonded to a radioactive metal” is, for example, an N 2 S 2 type metal chelate moiety bonded to the radioactive metal.

R ₃ in the general formula (III) is preferably a radioactive halogen atom, and more preferably ¹⁸ F, ¹²³ I, ¹²⁴ I or ¹²⁵ I.

R ₄ in the general formula (III) is preferably a group represented by —NRaRb, and more preferably a dimethylamino group or a methylamino group.

N in the general formula (III) may be an integer of 0 to 5, but is preferably 1, 2, or 3. In addition, when n is 0, it means that R ₃ is directly bonded to the benzoxazole ring.

In the formula: — (CH ₂ CH ₂ O) _i —A, i is preferably 1, 2, or 3.

_J in the formula: — (CH ₂ CH ₂ O) _j —OH is preferably 1, 2, or 3.

_K in the formula: — (CH ₂ CH ₂ O) _k —F is preferably 1, 2, or 3.

Typical compounds among the compounds represented by the general formula (III) are shown in the following table. In the table, “Me” represents a methyl group.

Among the above compounds, preferred compounds include III-26 ([ ¹⁸ F] compound 7) and III-27 ([ ¹⁸ F] compound 15).

The compound represented by the general formula (III) can be synthesized according to the method described in the examples or according to a method in which those methods are appropriately modified or modified with reference to the description.
R ₁ and R ₂ (D) a compound represented by the general formula (IV) formula in (IV) is a one of an electron donating group, but the other may be an electron withdrawing group, R ₁ is an electron In the donor group, R ₂ is preferably an electron withdrawing group.

R ₁ in the general formula (IV) may be in any position on the heterocycle, but is preferably in the 5-position (position when the heterocycle is assumed to be thiophene).

The electron donating group in the general formula (IV) is preferably a hydroxyl group, a methoxy group, a methyl group, an amino group, a methylamino group, a dimethylamino group, a methylaminophenyl group, or a dimethylaminophenyl group, and more preferably Is a dimethylamino group.

The electron withdrawing group in the general formula (IV) is preferably a nitro group, a nitrovinyl group, a cyano group, a dicyanovinyl group, a tricyanovinyl group, or a formyl group, and more preferably a dicyanovinyl group.

M in the general formula (IV) may be an integer of 0 to 5, and is preferably 1 or 2. In addition, when m is 0, it means that R ₂ is directly bonded to a thiophene ring or the like.

Typical compounds among the compounds represented by the general formula (IV) are shown in the following table. In the table, “Me” represents a methyl group, “Dicyano” represents a dicyanovinyl group, “Tricyano” represents a tricyanovinyl group, and “5-” represents the 5-position (complex of general formula (IV)). It represents that the ring is a group at the position when it is assumed to be thiophene.

Among the above compounds, preferred compounds are IV-23 (DTA-1), IV-35 (DTA-2), IV-47 (DTA-3), IV-71 (DFA-1), IV-83 (DFA -2).

The compound represented by the general formula (IV) can be synthesized by bonding an electron donating group and an electron withdrawing group directly or via an unsaturated carbon chain to a compound having a thiophene ring or a furan ring.
(E) Composition for diagnosing conformation disease The composition containing the compounds represented by the general formulas (I) to (IV) described above can be used for diagnosis of conformation disease.

Also, pharmaceutically acceptable salts can be used in place of the compounds represented by the general formulas (I) to (IV). Examples of pharmaceutically acceptable salts include alkali metal salts (sodium salt, potassium salt, lithium salt), alkaline earth metal salts (calcium salt, magnesium salt), sulfate, hydrochloride, nitrate, phosphate, etc. it can.

“Conformation disease” means a group of diseases caused by proteins that have become abnormal due to conformational transformation such as Aβ, tau protein, prion, etc. In addition to AD, hereditary cerebral dysemia with Down syndrome and Dutch amyloidosis (hereditary) cerebral hemorrhage with amyloidosis-Dutch type: HCHWA-D), Creutzfeldt-Jakob disease (CJD), bovine spongiform encephalopathy (BSE), and type 2 diabetes. In addition, diseases to be diagnosed include precursor symptoms of diseases that are generally not recognized as “diseases”. Examples of prodromal symptoms of such diseases include mild cognitive impairment (MCI) seen before the onset of AD.

In the diagnosis of conformation disease using the above composition, this composition is usually administered to a subject to be diagnosed or a laboratory animal, and then a brain image is taken and represented by the general formulas (I) to (IV) in the image. Based on the state (amount, distribution, etc.) of the compound to be produced. The method of administration of the composition is not particularly limited and can be appropriately determined according to the type of compound, the type of labeling substance, etc., but is usually intradermal, intraperitoneal, intravenous, arterial or spinal fluid injection or Administer by infusion. The dose of the composition is not particularly limited, and can be appropriately determined according to the type of compound, the type of labeling substance, etc. In the case of adults, the compound represented by general formulas (I) to (IV) is 1 It is preferable to administer 10 ⁻¹⁰ to 10 ⁻³ mg per day, and more preferably 10 ⁻⁸ to 10 ⁻⁵ mg.

As described above, since this composition is usually administered by injection or infusion, it may contain components usually contained in injection solutions or infusion solutions. Such components include liquid carriers (for example, potassium phosphate buffer, physiological saline, Ringer's solution, distilled water, polyethylene glycol, vegetable oils, ethanol, glycerin, dimethyl sulfoxide, propylene glycol, etc.), antibacterial agents And local anesthetics (eg, procaine hydrochloride, dibucaine hydrochloride, etc.), buffer solutions (eg, Tris-HCl buffer solution, Hepes buffer solution, etc.), osmotic pressure regulators (eg, glucose, sorbitol, sodium chloride, etc.) .

Hereinafter, the present invention will be described in more detail with reference to examples.

[Example 1] Near-infrared fluorescent compound having a benzothiazole ring (1) Synthesis of 1,3-benzothiazol-6-amine A mixture of 6-nitrobenzothiazole (5.0 g, 27.8 mmol) and cHCl (3.86 mL) 80% EtOH (126 mL) was added thereto, and iron powder (7.4 g) was added thereto, followed by stirring under reflux for 1 hour. The mixture was cooled to room temperature, filtered and extracted three times with EtOAc and H ₂ O. The organic layer was dehydrated and dried with Na ₂ SO ₄ , the solvent was distilled off under reduced pressure, and the resulting solid was purified by silica gel chromatography (CHCl ₃ : CH ₃ OH = 49: 1) (3.24 g, 77.6%). ¹ H NMR (400 MHz, CDCl ₃ ) d 8.70 (s, 1H), 7.89 (d, J = 8.8 Hz, 1H), 7.17 (d, J = 2.4 Hz, 1H), 6.87 (dd, J = 8.8, 2.4Hz, 1H), 3.85 (br, s, 2H).

(2) Synthesis of N, N-dimethyl-1,3-benzothiazol-6-amine 1 (2.41 g, 16.0 mmol) was dissolved in THF (65 mL), and HCHO (11.9 mL), 30% H ₂ SO ₄ (13.3 mL, 49.2 mmol) was added, iron powder (7.14 g, 128.5 mmol) was added, and the mixture was vigorously stirred for 1.5 hours. The mixture was filtered and washed twice with EtOAc. The mixture was basified with 2N NaOH and separated with EtOAc (50 ml × 2). The organic layer was dehydrated and dried with Na ₂ SO ₄ and the solvent was distilled off under reduced pressure. The resulting viscous residue was purified by silica gel chromatography (CHCl ₃ 100%) (766.3 mg, 26.9%). ¹ H NMR (400 MHz, CDCl ₃ ) d 8.66 (s, 1H), 7.94 (d, J = 8.8 Hz, 1H), 7.14 (d, J = 2.4 Hz, 1H), 6.99 (dd, J = 8.8, 2.4Hz, 1H), 3.02 (s, 6H).

(3) Synthesis of 6- (dimethylamino) -1,3-benzothiazole-2-carbaldehyde Dehydrated THF (5.8 mL) was added to n-BuLi (0.5 mL, 1.3 mmol) at -78 ° C under N ₂ filling. And N, N-dimethyl-1,3-benzothiazol-6-amine (2.22 g, 1.22 mmol) was added slowly with vigorous stirring. After stirring for 2 hours, the temperature was returned to room temperature, and dehydrated DMF (0.38 mL) was slowly added, followed by stirring for 1.5 hours. After neutralizing with saturated NH ₄ Cl and extracting twice with EtOAc, the organic layer was dehydrated and dried over Na ₂ SO ₄ , and the solvent was distilled off under reduced pressure. The resulting material was purified by silica gel chromatography (Hexane: EtOAc = 2: 1) (153 mg, 60.8%). ¹ H NMR (400MHz, CDCl ₃ ) δ 10.05 (s, 1H), 8.01 (d, J = 10.0 Hz, 1H), 7.06-7.04 (m, 2H), 3.1 (s, 6H).

(4) Synthesis of 3- (6- (dimethylamino) benzo [d] thiazol-2-yl) acrylaldehyde 6- (dimethylamino) -1,3-benzothiazole-2-carbaldehyde under N ₂ filling Put (78.9 mg, 0.38 mmol), (1,3-dioxolan-2-ylmethyl) triphenylphosphonium bromide (325.3 mg, 0.76 mmol), 18-crown-6 (ca 5 mg), dehydrated THF (23 mL) Was added. To this, NaH was slowly added, and after stirring for 1 hour, H ₂ O was added with ice cooling to stop the reaction. After extraction twice with EtOAc, the organic layer was dried over Na ₂ SO ₄ and the solvent was distilled off under reduced pressure. The obtained substance was dissolved in THF, 10% oxalic acid dihydrate was added, and the mixture was stirred for 2 hr. Saturated NaHCO ₃ aqueous solution was added to neutralize to basic, and then THF was distilled off under reduced pressure and extracted with EtOAc. The organic layer was washed with saturated brine, dehydrated and dried over Na ₂ SO ₄ , and evaporated under reduced pressure. The obtained material was purified by silica gel chromatography (CHCl ₃ : MeOH = 49: 1) (36.7 mg, 41.6%). ¹ H NMR (400 MHz, CDCl ₃ ) δ 9.76 (d, J = 8.0 Hz, 1H), 7.91 (d, J = 9.2 Hz, 1H), 7.68 (d, J = 16.0 Hz, 1H), 7.04-6.97 (m, 2H), 6.82-6.76 (m, 1H), 3.09 (s, 6H).

(5) Synthesis of 2- (3- (6- (dimethylamino) benzo [d] thiazol-2-yl) arylidene) malononitrile (KR-5) 3- (6- (dimethylamino) benzo [d] thiazole- 2-yl) acrylaldehyde (25.0 mg, 0.11 mmol), malononitrile (10.9 mg, 0.165 mmol) and pyridine (0.10 mL) were placed in 2-propanol and stirred at reflux for 30 minutes. The mixture was extracted twice with EtOAc, dehydrated and dried over Na ₂ SO ₄ , and the solvent was evaporated under reduced pressure. The obtained substance was purified by silica gel chromatography (hexane: EtOAc = 1: 1) (15.6 mg, 50.6%). ¹ H NMR (400 MHz, CDCl ₃ ) δ 7.91 (d, J = 9.2 Hz, 1H), 7.59 (d, J = 12.0 Hz, 1H), 7.47 (d, J = 14.8 Hz, 1H), 7.37-7.34 (m, 1H), 7.06-7.01 (m, 2H), 3.12 (s, 6H). HRMS (EI): m / z calcd for C ₁₅ H ₁₂ N ₄ S 280.0782; found 280.0775.

(6) Synthesis of 5- (6- (dimethylamino) benzo [d] thiazol-2-yl) penta-2,4-dienal 3- (6- (dimethylamino) benzo [d] thiazol-2-yl) Add acrylic aldehyde (210.7 mg, 0.92 mmol), (1,3-dioxolan-2-ylmethyl) triphenylphosphonium bromide (780.0 mg, 1.82 mmol), 18-crown-6 (ca 5 mg), THF (50 mL) Was added. Furthermore, NaH (91 mg, 3.8 mmol) was added, and after stirring for 1.5 hours, H ₂ O was added with ice cooling to stop the reaction. After separation with EtOAc (50 mL × 1, 30 mL × 2), the organic layer was dried over Na ₂ SO ₄ and the solvent was distilled off under reduced pressure. The resulting material was dissolved in THF (100 mL), 10% oxalic acid dihydrate was added and stirred for 45 minutes. Saturated aqueous NaHCO ₃ solution was added to neutralize to basic, and then THF was distilled off under reduced pressure, followed by separation with EtOAc (100 mL × 2). The organic layer was washed with saturated brine, dehydrated and dried over Na ₂ SO ₄ , and evaporated under reduced pressure. The obtained material was purified by silica gel chromatography (hexane: EtOAc = 65: 35) (100.4 mg, 42.3%). ¹ H NMR (400 MHz, CDCl ₃ ) δ 9.66 (d, J = 8.0 Hz, 1H), 7.85 (d, J = 9.2 Hz, 1H), 7.25-7.05 (m, 4H), 6.96 (dd, J = 9.2, 2.4 Hz, 1H), 6.38-6.33 (m, 1H), 3.07 (s, 6H).

(7) Synthesis of 2- (5- (6- (dimethylamino) benzo [d] thiazol-2-yl) penta-2,4-diene-1-ylidene) malononitrile (KR-7) 5- (6- (Dimethylamino) benzo [d] thiazol-2-yl) penta-2,4-dienal (22.3 mg, 0.086 mmol), malononitrile (11.6 mg, 0.18 mmol), pyridine (0.10 mL) in 2-propanol, Stir at reflux for 1 hour. The mixture was partitioned twice with EtOAc, dehydrated and dried over Na ₂ SO ₄ , and the solvent was evaporated under reduced pressure. The obtained material was purified by silica gel chromatography (CHCl ₃ : MeOH = 49: 1) (7.9 mg, 28.5%). ¹ H NMR (400 MHz, CDCl ₃ ) δ 7.85 (d, J = 8.8 Hz, 1H), 7.46 (d, J = 11.6 Hz, 1H), 7.22 (d, J = 14.8 Hz, 1H), 7.13-7.02 (m, 3H), 6.97 (dd, J = 9.2, 2.4Hz, 1H), 6.91-6.85 (m, 1H), 3.09 (s, 6H) .HRMS (EI): m / z calcd for C ₁₇ H ₁₄ N ₄ S 306.0939; found 306.0930.

(8) Synthesis of 7- (6- (dimethylamino) benzo [d] thiazol-2-yl) hepta-2,4,6-trienal 5- (6- (dimethylamino) benzo [d] thiazole-2- Yl) penta-2,4-dienal (93.8 mg, 0.36 mmol), (1,3-dioxolan-2-ylmethyl) triphenylphosphonium bromide (298.5 mg, 0.72 mmol), 18-crown-6 (ca 5 mg) And THF (25 mL) was added. Further, NaH (35.6 mg, 1.48 mmol) was added and stirred for 1.5 hours, and then the same amount of NaH was added again. After stirring for 2 hours, H ₂ O was added with ice cooling to stop the reaction. After separation with EtOAc (50 mL × 2), the organic layer was dried over Na ₂ SO ₄ and the solvent was distilled off under reduced pressure. The obtained material was dissolved in THF (50 mL), 10% oxalic acid dihydrate was added, and the mixture was stirred for 30 min. Saturated aqueous NaHCO ₃ solution was added to make the solution neutral to basic, and then THF was distilled off under reduced pressure, followed by separation with EtOAc (50 mL × 2). The organic layer was washed with saturated brine, dehydrated and dried over Na ₂ SO ₄ , and evaporated under reduced pressure. The obtained substance was purified by silica gel chromatography (hexane: EtOAc = 60: 40) (75.8 mg, 74.0%). ¹ H NMR (400 MHz, CDCl ₃ ) δ 10.21-9.58 (m, 1H), 7.81 (d, J = 9.2 Hz, 1H), 7.43-6.6 (m, 7H), 6.24-5.91 (m, 1H), 3.04 (s, 6H).

(9) Synthesis of 2- (7- (6- (dimethylamino) benzo [d] thiazol-2-yl) hepta-2,4,6-trien-1-ylidene) malononitrile (KR-9) 7- ( 6- (Dimethylamino) benzo [d] thiazol-2-yl) hepta-2,4,6-trienal (72.3 mg, 0.254 mmol), malononitrile (25.2 mg, 0.381 mmol), pyridine (0.30 mL) The mixture was placed in propanol and stirred at reflux for 1.75 hours. More malononitrile (21.4 mg, 0.32 mmol) was added and stirred overnight. The mixture was partitioned with EtOAc (150 mL × 1, 50 mL × 1), dried over Na ₂ SO ₄ and dried under reduced pressure. The obtained material was purified by silica gel chromatography (CHCl ₃ : MeOH = 49: 1) (38.1 mg, 45.1%). ¹ H NMR (400 MHz, CDCl ₃ ) δ 7.82 (d, J = 9.6 Hz, 1H), 7.45 (d, J = 11.6 Hz, 1H), 7.10-6.93 (m, 5H), 6.89-6.75 (m, 2H), 6.67-6.6 (m, 1H), 3.07 (s, 6H) .HRMS (EI): m / z calcd for C ₁₉ H ₁₆ N ₄ S 332.1096; found 332.1090.

(10) a 100 [mu] L that the study sample was dissolved in CHCl ₃ fluorescence properties, further put CHCl ₃ 400 [mu] L to the cell, absorption, fluorescence was measured with excitation wavelength. The quantum yield was measured by adding 200 μL of a sample dissolved in CHCl ₃ and further 800 μL of CHCl ₃ in a dedicated cuvette (Table 6).

All the compounds showed an absorption wavelength and an excitation wavelength in the range of 530 to 550 nm, while the fluorescence wavelength became longer with the elongation of the vinyl group and showed a maximum in the range of 611 to 716 nm. The fluorescence quantum yield was 45.5% for the previously reported PP-BTA-1 [2-((6- (dimethylamino) benzo [d] thiazol-2-yl) methylene) malononitrile], whereas The value decreased with the elongation of the vinyl group, reaching 0.7% for KR-9.

(11) Examination of fluorescence characteristics in the presence and absence of Aβ Each sample was dissolved in 50% EtOH so that the concentration was 12.5 μM. 90 μL of this solution, 504 μL of PBS, and 396 μL of Aβ42 aggregate (57 nM) were mixed and allowed to stand for 30 minutes, and then the fluorescence wavelength was measured. In addition, a test solution in which Aβ42 aggregates were not mixed was also prepared, and the fluorescence wavelength was similarly measured (FIG. 2).

In any compound, a new fluorescence peak was observed in a wavelength region that was not detected in the absence of Aβ in the presence of Aβ. These results indicate that these compounds emit fluorescence having a new wavelength when bound to Aβ, and thus may be applicable to Aβ-specific imaging.

(12) In vitro binding experiments using Aβ42 aggregates 30 μL of 10% ethanol solution of PP-BTA-1, KR-5, KR-7, KR-9 prepared at various concentrations, 168 μL of PBS, Aβ42 coagulation The collected (57 nM) 132 μL was mixed and allowed to stand for 30 minutes. In addition, a test solution was prepared by mixing 30 μL of each sample and 300 μL of PBS without adding Aβ42 aggregates. The fluorescence intensity of each was measured, and the dissociation constant (Kd) of each compound was calculated from the saturation curve (FIG. 3).

All the compounds showed binding ability to Aβ aggregates in the order of nM, and the binding properties improved in the order of PP-BTA-1 <KR-5 <KR-9 <KR-7.

(13) Examination of in vitro fluorescent staining using Tg2567 mouse brain sections In order to evaluate the binding of each compound to mouse senile plaques, fluorescent staining experiments using frozen brain sections of 32-month-old Tg2576 mice were performed. 150 μL of a 50% ethanol solution of each compound prepared to a concentration of 100 μM was stained with a brain section, allowed to stand for 10 minutes, washed twice with 50% EtOH for 1 minute, and observed with a fluorescence microscope. Further, adjacent brain sections were stained with thioflavin S (100 μM, 150 μL) and observed with a fluorescence microscope (FIG. 4).

In any compound, a large number of fluorescent images were observed on Tg2576 mouse brain sections, and these fluorescent images were consistent with the stained images of thioflavin S on adjacent brain sections. From these results, it was shown that all of these compounds have binding properties to mouse senile plaques.

(14) In Vivo Imaging Experiment Using Tg2576 Mice 50 μL of KR-5 solution (1.6 × 10 ⁻² mg / mouse, 20% DMSO, 80% propylene glycol) was added to Tg2576 mice (24 months old) and wild mice (5 Week). Thereafter, imaging was performed at an excitation wavelength of 535 nm and a fluorescence wavelength of 660 nm using a fluorescence imaging apparatus (IVIS Spectrum) at 5 min, 30 min, and 60 min (FIG. 5). In addition, after 1 hour, the mice were sacrificed and the brains were removed and imaged by IVIS (FIG. 5). Furthermore, after freezing the brains of Tg2576 mice, sections were prepared with a thickness of 20 μm, and the sections were observed with a fluorescence microscope (FIG. 6). After washing this section with 50% EtOH for 20 minutes, it was confirmed that the fluorescence image of KR-5 was completely washed, and reacted with 150 μL of thioflavin S (100 μM) for 2 minutes. Thereafter, it was washed with 50% EtOH and observed with a fluorescence microscope (FIG. 6).

Fluorescence accumulation in the mouse brain was observed after administration of KR-5 to mice, suggesting that KR-5 exhibits brain migration. Thereafter, the disappearance of fluorescence from the brain was observed over time, suggesting that there was little non-specific binding of the compounds. Furthermore, when the brain was removed and compared with the wild-type mouse brain using a fluorescence imaging apparatus, remarkable fluorescence accumulation was confirmed in the Tg2576 mouse brain compared with the wild-type mouse brain. After imaging, frozen sections were prepared and observed with a fluorescence microscope. As a result, a fluorescent image derived from a compound almost corresponding to the fluorescent image of thioflavin S was observed in the same section. From these results, it became clear that KR-5 has the ability to migrate to the mouse brain and has binding properties to senile plaques in the brain.

[Example 2] Near-infrared fluorescent compound having a benzene ring (1) Synthesis of (2E, 4E) -5- (4- (dimethylamino) phenyl) penta-2,4-dienal 175 mg of 4-N, 20 mL containing N-dimethylcinnamaldehyde (1.0 mmol), 53 mg 18-crown-6 (0.2 mmol) and 132 mg (1,3-dioxolan-2-yl) methyltriphenylphosphonium bromide (1.0 mmol) To a THF solution of 80 mg NaH (2.0 mmol) was added. The reaction mixture was stirred at room temperature for 3 hours, then 5 mL of concentrated hydrochloric acid was added, stirred for an additional hour at room temperature, and then neutralized with K ₂ CO ₃ . The mixture was extracted with CHCl ₃ (30 mL). After the solvent was removed, the residue was purified by silica gel chromatography to give 84.4 mg of product. The yield was 42% (cis-trans isomer).

(2) Synthesis of (2E, 4E, 6E) -7- (4- (dimethylamino) phenyl) hepta-2,4,6-trienal (2E, 4E) -5- (4- (dimethylamino) phenyl) The same reaction described for penta-2,4-dienal was used. A brown oil was obtained. The yield was 33%.

(3) Synthesis of 2- (4- (dimethylamino) benzylidene) malononitrile (DANIR-1) 20 mg containing 300 mg 4-N, N-dimethylcinnamaldehyde (2.0 mmol) and 132 mg malononitrile (2.0 mmol) To mL of EtOH solution, 200 μL piperazine was added. The reaction mixture was stirred at room temperature for 10 minutes and yellow crystals (362 mg) were formed. The yield was 92%. ¹ H NMR (400 MHz, CDCL ₃ ) δ 7.82 (d, J = 9.0 Hz, 2H), 7.47 (s, 1H), 6.69 (d, J = 9.1 Hz, 2H), 3.14 (s, 6H). ¹³ C-NMR (125 MHz, CDCl ₃ ): δ 157.98, 154.19, 133.73, 119.19, 115.96, 114.89, 111.54, 71.65, 40.02.HRMS (EI): m / z calcd for C ₁₂ H ₁₁ N ₃ 197.0953; found 197.0948 Anal.Calcd: C 73.07, H 5.62, N 21.30; Found: C 73.02, H 5.68, N 21.25.

(4) Synthesis of (E) -2- (3- (4- (dimethylamino) phenyl) arylidene) malononitrile (DANIR-2) The same reaction described for DANIR-1 was used. Dark red crystals were obtained. The yield was 90%. ¹ H NMR (400 MHz, CDCL ₃ ) δ 7.48 (m, 3H), 7.17 (d, J = 14.8 Hz, 1H), 7.02 (dd, J = 14.8, 11.7 Hz, 1H), 6.68 (d, J = . 9.0 Hz, 2H), 3.10 (s, 6H) 13 C-NMR (125 MHz, CDCl 3):. δ 160.44, 153.02, 151.53, 131.57, 121.76, 117.16, 114.98, 113.07, 111.89, 40.04 HRMS (EI) : m / z calcd .for C ₁₄ H ₁₃ N ₃ 223.1109; found 223.1115. Anal.Calcd: C 75.31, H 5.87, N 18.82; Found: C 75.08, H 5.94, N 18.76.

(5) Synthesis of 2-((2E, 4E) -5- (4- (dimethylamino) phenyl) penta-2,4-dien-1-ylidene) malononitrile (DANIR-3) Same as described for DANIR-1 The reaction was used. A purple solid was obtained. The yield was 69.0%. ¹ H NMR (400 MHz, CDCL ₃ ) δ 7.42 (m, 3H), 7.06 (dd, J = 14.4, 11.2 Hz, 1H), 6.97 (d, J = 14.8 Hz, 1H), 6.80 (dd, J = 15.2, 11.2 Hz, 1H), 6.72 (d, J = 14.6 Hz, 1H), 6.68 (d, J = 9.2 Hz, 2H), 3.06 (s, 6H). ¹³ C-NMR (125 MHz, CDCl ₃ ) : δ 159.52, 152.10, 151.88, 146.40, 130.00, 123.36, 123.09, 121.99, 114.73, 112.72, 111.94, 40.06.HRMS (EI): m / z calcd for C ₁₆ H ₁₅ N ₃ 249.1266; found 249.1259. Anal. Calcd : C 77.08, H 6.06, N 16.85; Found: C 76.95, H 6.04, N 16.73.

(6) Synthesis of 2-((2E, 4E, 6E) -7- (4- (dimethylamino) phenyl) hepta-2,4,6-trien-1-ylidene) malononitrile (DANIR-4) DANIR-1 The same reaction described for was used. A black solid was obtained. The yield was 31%. ¹ H NMR (400 MHz, CDCL ₃ ) δ 7.37 (d, J = 11.0 Hz, 1H), 7.36 (d, J = 9.2 Hz, 2H), 6.94 (dd, J = 14.3, 11.5 Hz, 1H), 6.89 -6.58 (m, 6H), 6.41 (dd, J = 13.6, 11.8 Hz, 1H), 3.17-2.68 (m, 6H) .HRMS (EI): m / z calcd for C ₁₈ H ₁₇ N ₃ 275.1417; found 275.1421.

(7) Fluorescent staining of Aβ plaques using Tg mice and AD human brain (FIGS. 9 and 10)
Paraffin-embedded brain tissue from an AD animal model (the C57-APP / PS1 mouse, 12 months old) and AD human (93 years old, female) was used for in vitro fluorescent staining. Brain slices were washed 2 x 20 min in xylene, 2 x 5 min 100% ethanol, 5 min 90% ethanol / H ₂ O, 5 min 80% ethanol / H ₂ O Deparaffinized by washing in water, 5 minutes in 60% ethanol / H ₂ O, and 10 minutes in running tap water, then in PBS (0.2 M, pH = 7.4) for 30 minutes Incubated. They were then incubated with a 10% solution of probe in ethanol (1 μM) for 10 minutes. Plaque location was confirmed by staining of adjacent sections with Thioflavin S (1 μM).

(8) Binding assay using aggregated Aβ _{1-42 in} solution (FIG. 11)
100 μL of aggregated Aβ fiber (concentration in final assay mixture: 60 nM), 100 μL of radioligand ([ ¹²⁵ I] IMPY) at appropriate concentration, 10 μL of inhibitor (concentration in ethanol of 10 ⁻⁵ -10 ^{− 10} M.) and 790 μL of PBS (0.2 M, pH = 7.4) and added to a mixture with a final volume of 1 mL. Nonspecific binding was defined in the presence of 1 μM IMPY. The mixture was incubated for 2 hours at 37 ° C. with constant rocking. The bound and free radioactivity was then separated by vacuum filtration through a borosilicate glass fiber filter (Whatman GF / B) using an M-24 cell harvester (Brandel, Gaithersburg, MD). The radioactivity of the filter containing bound ¹²⁵ I ligand was measured with a γ-counter (WALLAC / Wizard 1470, USA) with an efficiency of 70%. Under the assay conditions, the specific binding fraction accounted for 10% of the total radioactivity. The half-maximal inhibitory concentration (IC ₅₀ ) was determined using GraphPad Prism 4.0, and the inhibition constant (K _i ) was determined using the Cheng-Prusoff equation: K _i = IC ₅₀ / (1 + [L] / K _d ) It was calculated.

(9) Saturation binding assay of DANIRs to Aβ _1-42 aggregates (Figure 12, Table 7)
100 μL of aggregated Aβ fiber (10 μg in the final assay mixture) contains 100 μL DANIRs (10 ^-5.5 -10 ^-10 nM in EtOH) and 800 μL PBS (0.2 M, pH = 7.4) Added to the mixture with a volume of 1 mL. Non-specific binding was defined without DANIRs. The mixture was incubated for 30 minutes at room temperature, then the solution was transferred to a 96-well plate and the fluorescence intensity was measured by a UV microplate reader. The results of saturation experiments were calculated using GraphPad Prism 4.0.

(10) Brain fluorescence imaging of DANIR-3 in vivo in nude mice (Figure 13)
50 μL of DANIR-3 (0.4 mg / kg, 20% DMSO, 80% propylene glycol) was injected into nude mice in vivo. Fluorescent signals from the brain were recorded by Xenogen's IVIS 200 Imaging station at 2, 5, 10, 30, 60, and 240 minutes after intravenous injection of the probe, and the data was analyzed by Living Image Software.

(11) Brain fluorescence imaging of DANIR-3 in vivo in normal and Tg mice (FIGS. 14 and 15)
50 μL of DANIR-3 (0.4 mg / kg, 20% DMSO, 80% propylene glycol) was injected into normal C57-BL6 and Tg-2576 (20 months) mice in vivo. Fluorescent signals from the brain were recorded by the Xenogen's IVIS 200 Imaging station at various times after injection and the data analyzed by Living Image Software. The ROI was drawn around the brain area and a time intensity curve was drawn.

Example 3 Benzoxazole Derivatives (1) General Review Unless otherwise indicated, all reagents were obtained commercially and used without further purification. Using a JEOL JNM-AL400 NMR spectrometer at 400 MHz, a ¹ H-NMR spectrum was obtained in CDCl ₃ solution using TMS as an internal standard. Chemical shifts are reported as δ values relative to the internal standard TMS. Coupling constants are reported in Hertz. Multiplicity is defined by s (single line), d (double line), t (triple line), and m (multiple line). Mass spectra were obtained with Shimadzu GC MS-QP2010 Plus (APCI). Shimadzu system (a LC-20AT pump with a SPD) using acetonitrile / water and Cosmosil C18 column (Nakalai Tesque, 5C ₁₈ AR II, 4.6 mm x 150 mm) as the mobile phase sent at a flow rate of 1.0 mL / min. HPLC was performed with a -20A UV detector, λ = 254 nm). Observation was performed with a fluorescence microscope (KEYENCE BioRevo BZ-9000) equipped with a GFP filter set. The binding assay was performed on a cell harvester (Brandel, M-24, Gaithersburg, MD).

(2) Synthesis of 2-amino-4-methoxyphenol (compound 2) A mixture of compound 1 (5.1 g, 40 mmol) and Pd / C (10%, 0.5 g) was mixed with anhydrous methanol under a hydrogen atmosphere (balloon). (200 mL) was stirred vigorously at room temperature for 24 hours. The mixture was filtered and the filtrate was washed with methanol (20 mL) and concentrated under reduced pressure to give compound 2 (3.72 g, 89.2%).

(3) Synthesis of 4- (5-methoxybenzo [d] oxazol-2-yl) -N-methylaniline (Compound 3) Compound 2 (1.39 g, 10 mmol) and 4-methylaminobenzoic acid (1.51 g, 10 mmol) ) Was added polyphosphoric acid (10 g). The mixture was stirred for 30 minutes at 180 ° C., ice water (400 mL) was added and neutralized with K ₂ CO ₃ . The mixture was extracted with ethyl acetate (200 mL) and the organic phase was separated and dried over Na ₂ SO ₄ . After removing the solvent, the residue was purified by silica gel chromatography to give compound 3 (783 mg, 30.8%). ¹ H NMR (400 MHz, CD ₃ OD) δ 7.94 (d, J = 9.0 Hz, 2H), 7.44 (d, J = 8.7 Hz, 1H), 7.12 (d, J = 2.4 Hz, 1H), 6.89 ( dd, J = 8.9, 2.6 Hz, 1H), 6.69 (d, J = 8.9 Hz, 2H), 3.84 (s, 3H), 2.85 (s, 3H). MS (ESI): m / z calcd for C ₁₅ H ₁₄ N ₂ O ₂ 254.11; found 255.10 (M + H ⁺ ).

(4) Synthesis of 2- (4- (methylamino) phenyl) benzo [d] oxazol-5-ol (compound 4) A droplet of BBr ₃ (1 M in CH ₂ Cl ₂ , 10 mL) was added to dry ice. To a solution of compound 3 (510 mg, 2 mmol) in an acetone bath. The reaction mixture was stirred at −78 ° C. for 1 hour and then at room temperature for 12 hours, then ice water (50 mL) was added and neutralized with K ₂ CO ₃ . The mixture was extracted with chloroform (100 mL) and the organic phase was separated and dried over Na ₂ SO ₄ . Filtration and purification of the residue by silica gel chromatography gave Compound 4 (437 mg, 91.1%). ¹ H NMR (400 MHz, CD ₃ OD) δ 7.93 (d, J = 8.8 Hz, 2H), 7.37 (d, J = 8.8 Hz, 1H), 6.99 (d, J = 2.1 Hz, 1H), 6.77 ( dd, J = 8.7, 2.4 Hz, 1H), 6.69 (d, J = 8.8 Hz, 2H), 2.85 (s, 3H) .MS (ESI): m / z calcd for C ₁₄ H ₁₂ N ₂ O ₂ 240.09 ; found 241.10 (M + H ⁺ ).

(5) Synthesis of 2- (2- (2-((2- (4- (methylamino) phenyl) benzo [d] oxazol-5-yl) oxy) ethoxy) ethoxy) ethanol (Compound 5) Compound 4 ( 480 mg, 2 mmol) and 2- [2- (2-chloroethoxy) -ethoxy] ethanol (50 μL, 0.66 mmol) in DMF (15 mL) with anhydrous K ₂ CO ₃ (830 mg, 6 mmol ) Was added. The reaction mixture was stirred at 105 ° C. for 4 hours, poured into water and extracted with chloroform. The organic layers were combined and dried over Na ₂ SO ₄ . The solvent was evaporated to give a residue that was purified by silica gel chromatography to give compound 5 (550 mg, 73.9%). ¹ H NMR (400 MHz, CDCL ₃ ) δ 8.05 (d, J = 8.3 Hz, 2H), 7.38 (d, J = 8.8 Hz, 1H), 7.28 (d, J = 2.4 Hz, 1H), 6.90 (dd , J = 8.8, 2.5 Hz, 1H), 6.67 (d, J = 8.4 Hz, 2H), 4.24-4.14 (m, 2H), 3.91-3.86 (m, 2H), 3.79-3.68 (m, 6H), 3.67-3.61 (m, 2H), 2.92 (s, 3H). MS (ESI): m / z calcd for C ₂₀ H ₂₄ N ₂ O ₅ 372.17; found 373.15 (M + H ⁺ ).

(6) N-methyl-4- (5-((2,2,3,3-tetramethyl-4,7,10-trioxa-3-siladodecan-12-yl) oxy) benzo [d] oxazole-2 Synthesis of -yl) aniline (compound 6) A solution of compound 5 (372 mg, 1 mmol), imidazole (140 mg, 2 mmol) in TBDMSCl (226 mg, 1.5 mmol) and dichloromethane (10 ml) at room temperature 3 Stir for hours. A white solid was formed and filtered. After evaporating the filtrate, the residue was purified by silica gel column chromatography to obtain compound 6 (302 mg, 62.1%). ¹ H NMR (400 MHz, CDCL ₃ ) δ 7.96 (d, J = 8.6 Hz, 2H), 7.30 (d, J = 8.8 Hz, 1H), 7.14 (d, J = 2.5 Hz, 1H), 6.82 (dd , J = 8.8, 2.4 Hz, 1H), 6.58 (d, J = 8.7 Hz, 2H), 4.22 (s, 1H), 4.13-4.04 (m, 2H), 3.84-3.79 (m, 2H), 3.71 ( t, J = 5.4 Hz, 2H), 3.69-3.61 (m, 4H), 3.51 (t, J = 5.4 Hz, 2H), 2.81 (s, 3H), 0.83 (s, 9H). MS (ESI): m / z calcd for C ₂₆ H ₃₈ N ₂ O ₅ Si 486.25; found 487.30 (M + H +).

(7) Synthesis of 4- (5- (2- (2- (2-fluoroethoxy) ethoxy) ethoxy) benzo [d] oxazol-2-yl) -N-methylaniline (Compound 7) Compound 5 (74 mg , 0.2 mmol) in CHCl ₃ (10 mL) was added DAST (100 mg, 0.6 mmol) in a dry ice acetone bath. The reaction mixture was stirred for 2 hours at room temperature, then poured into saturated NaHSO ₃ solution and extracted with chloroform. The organic phase was separated, dried over MgSO ₄ and filtered. The residue was purified by silica gel column chromatography to obtain compound 7 (7.1 mg, 9.6%). ¹ H NMR (400 MHz, CDCL ₃ ) δ 8.04 (d, J = 8.5 Hz, 2H), 7.38 (d, J = 8.8 Hz, 1H), 7.21 (d, J = 2.5 Hz, 1H), 6.89 (dd , J = 8.8, 2.5 Hz, 1H), 6.67 (d, J = 8.6 Hz, 2H), 4.63 (dd, J = 4.5, 3.9 Hz, 1H), 4.51 (dd, J = 6.0, 2.3 Hz, 1H) , 4.23-4.13 (m, 2H), 3.95-3.86 (m, 2H), 3.84-3.79 (m, 2H), 3.78-3.71 (m, 4H), 2.92 (s, 3H) .HRMS (EI): m / z calcd for C ₂₀ H ₂₃ FN ₂ O ₄ 374.1642; found 374.1637 (M + H ⁺ ).

(8) tert-butylmethyl (4- (5-((2,2,3,3-tetramethyl-4,7,10-trioxa-3-siladodecan-12-yl) oxy) benzo [d] oxazole-2 Synthesis of -yl) phenyl) carbamate (Compound 8) Under nitrogen atmosphere, Compound 6 (300 mg, 0.62 mmol) was dissolved in anhydrous THF (20 ml) and then in Boc anhydride (1.35 g, 6.2 mmol). It was. The solution was refluxed for 48 hours. After completion of the reaction, the solvent was removed and the residue was purified by silica gel column chromatography to obtain compound 8 (273 mg, 74.7%). ¹ H NMR (400 MHz, CDCL ₃ ) δ 8.11 (d, J = 8.4 Hz, 2H), 7.37 (d, J = 8.8 Hz, 1H), 7.35 (d, J = 8.5 Hz, 2H), 7.19 (d , J = 2.5 Hz, 1H), 6.91 (dd, J = 8.8, 2.5 Hz, 1H), 4.19-4.09 (m, 2H), 3.88-3.80 (m, 2H), 3.71 (t, J = 5.4 Hz, 2H), 3.69-3.61 (m, 4H), 3.51 (dd, J = 5.6, 5.1 Hz, 2H), 3.26 (s, 3H), 1.42 (s, 9H), 0.83 (s, 9H). MS (ESI ): m / z calcd for C ₃₁ H ₄₆ N ₂ O ₇ Si 586.31; found 587.40 (M + H ⁺ ).

(9) Synthesis of tert-butyl (4- (5- (2- (2- (2-hydroxyethoxy) ethoxy) ethoxy) benzo [d] oxazol-2-yl) phenyl) (methyl) carbamate (Compound 9) To a solution of compound 8 (270 mg, 0.46 mmol) in dry THF (10 mL) was added anhydrous TBAF (1.0 mL, 1 M in THF). The solution was stirred at room temperature for 2 hours. After removal of THF, the residue was purified by silica gel column chromatography to obtain compound 9 (281 mg, 75.7%). ¹ H NMR (400 MHz, CDCL ₃ ) δ 8.09 (d, J = 8.8 Hz, 2H), 7.39-7.30 (m, 3H), 7.22 (s, 1H), 6.90 (d, J = 8.8 Hz, 1H) , 4.11 (dd, J = 6.2, 2.9 Hz, 2H), 3.88-3.75 (m, 2H), 3.70-3.60 (m, 6H), 3.58-3.51 (m, 2H), 3.24 (s, 3H), 1.41 (s, 9H). MS (ESI): m / z calcd for C ₂₅ H ₃₂ N ₂ O ₇ 472.22; found 473.30 (M + H ⁺ ).

(10) 2- (2- (2-((2- (4-((tert-butoxycarbonyl) (methyl) amino) phenyl) benzo [d] oxazol-5-yl) oxy) ethoxy) ethoxy) ethylmethane) Synthesis of Sulfonic Acid (Compound 10) To a solution of Compound 9 (281 mg, 0.6 mmol) in methanesulfonic acid (10 ml) was added triethylamine (121 mg, 1.2 mmol). Methanesulfonyl chloride (137 mg, 1.2 mmol) was then added via syringe. The solution was stirred at room temperature for 2 hours. Then 20 mL of water was added and extracted with CHCl ₃ . The organic layer was dried with MgSO ₄ . After the solvent was removed, the residue was purified by silica gel chromatography to give compound 10 (173 mg, 52.4%). ¹ H NMR (400 MHz, CDCL ₃ ) δ 8.17 (d, J = 8.7 Hz, 2H), 7.45 (d, J = 8.9 Hz, 1H), 7.42 (d, J = 8.7 Hz, 2H), 7.25 (d , J = 2.5 Hz, 1H), 6.97 (dd, J = 8.9, 2.5 Hz, 1H), 4.42-4.35 (m, 2H), 4.21-4.14 (m, 2H), 3.88 (dd, J = 5.3, 4.0 Hz, 2H), 3.81-3.77 (m, 2H), 3.76-3.69 (m, 4H), 3.33 (s, 3H), 3.06 (s, 3H), 1.49 (s, 9H). MS (ESI): m / z calcd for C ₂₆ H ₃₄ N ₂ O ₉ S 550.20; found 551.25 (M + H ⁺ ).

(11) Synthesis of 4- (5-methoxybenzo [d] oxazol-2-yl) -N, N-dimethylaniline (Compound 11) Using the above reaction for preparing Compound 3, Compound 11 was obtained in a yield of 25.1%. Got in. ¹ H NMR (400 MHz, CDCL ₃ ) δ 8.08 (d, J = 8.7 Hz, 2H), 7.39 (d, J = 8.8 Hz, 1H), 7.21 (d, J = 2.5 Hz, 1H), 6.86 (dd , J = 8.8, 2.5 Hz, 1H), 6.77 (d, J = 8.9 Hz, 2H), 3.86 (s, 3H), 3.07 (s, 3H). MS (ESI): m / z calcd for C ₁₆ H ₁₆ N ₂ O ₂ 268.12; found 269.15 (M + H ⁺ ).

(12) Synthesis of 2- (4- (dimethylamino) phenyl) benzo [d] oxazol-5-ol (Compound 12) Using the above reaction for preparing Compound 4, Compound 12 was obtained in a yield of 88.0%. . ¹ H NMR (400 MHz, CD ₃ OD) δ 7.99 (d, J = 8.9 Hz, 2H), 7.38 (d, J = 8.7 Hz, 1H), 7.00 (d, J = 2.4 Hz, 1H), 6.84 ( d, J = 8.9 Hz, 2H), 6.78 (ddd, J = 8.8, 2.4, 0.8 Hz, 1H), 3.07 (s, 6H) .MS (ESI): m / z calcd for C ₁₅ H ₁₄ N ₂ O ₂ 254.11; found 255.10 (M + H ⁺ ).

(13) Synthesis of 2- (2- (2-((2- (4- (dimethylamino) phenyl) benzo [d] oxazol-5-yl) oxy) ethoxy) ethoxy) ethanol (compound 13) Compound 13 was obtained in a yield of 43.1% using the prepared reaction. ¹ H NMR (400 MHz, CDCL ₃ ) δ 8.05 (d, J = 7.9 Hz, 2H), 7.37 (d, J = 8.8 Hz, 1H), 7.24 (d, J = 2.3 Hz, 1H), 6.88 (dd , J = 8.8, 2.5 Hz, 1H), 6.72 (d, J = 8.2 Hz, 2H), 4.17 (dd, J = 4.9, 3.8 Hz, 2H), 3.86 (dd, J = 4.9, 3.8 Hz, 2H) , 3.78-3.42 (m, 8H), 3.02 (s, 6H) .MS (ESI): m / z calcd for C ₂₁ H ₂₆ N ₂ O ₅ 386.18; found 387.27 (M + H ⁺ ).

(14) 2- (2- (2-((2- (4- (dimethylamino) phenyl) benzo [d] oxazol-5-yl) oxy) ethoxy) ethoxy) ethyl 4-methylbenzenesulfonic acid (compound 14 Tosyl chloride (248 mg, 1.30 mmol) was added to a solution of compound 13 (250 mg, 0.65 mmol) in pyridine (10 mL). The reaction mixture was stirred at room temperature for 3 hours, 50 mL of water was added, and the mixture was extracted with CHCl ₃ . The organic layer was dried over Na ₂ SO ₄ and the residue was obtained by evaporation of the solvent. The residue was purified by silica gel chromatography to give compound 14 (173 mg, 52.4%). ¹ H NMR (400 MHz, CDCL ₃ ) δ 8.06 (d, J = 9.0 Hz, 2H), 7.79 (d, J = 8.3 Hz, 2H), 7.37 (d, J = 8.8 Hz, 1H), 7.31 (d , J = 8.0 Hz, 2H), 7.19 (d, J = 2.5 Hz, 1H), 6.87 (dd, J = 8.8, 2.5 Hz, 1H), 6.74 (d, J = 9.1 Hz, 2H), 4.20-4.11 (m, 4H), 3.88-3.81 (m, 2H), 3.74-3.56 (m, 6H), 3.04 (s, 6H), 2.40 (s, 3H). MS (ESI): m / z calcd for C ₂₈ H ₃₂ N ₂ O ₇ S 540.19; found 541.29 (M + H ⁺ ).

(15) Synthesis of 4- (5- (2- (2- (2-fluoroethoxy) ethoxy) ethoxy) benzo [d] oxazol-2-yl) -N, N-dimethylaniline (Compound 15) To compound 14 (54 mg, 0.1 mmol) in dry tetrahydrofuran (THF) was added anhydrous TBAF (300 μL, 1 M in THF). The reaction mixture was refluxed for 3 hours. After removal of THF, the residue was purified by silica gel chromatography to give compound 15 (22 mg, 56.7%). ¹ H NMR (400 MHz, CDCL ₃ ) δ 8.08 (d, J = 9.0 Hz, 2H), 7.38 (d, J = 8.8 Hz, 1H), 7.21 (d, J = 2.5 Hz, 1H), 6.89 (dd , J = 8.8, 2.5 Hz, 1H), 6.77 (d, J = 9.1 Hz, 2H), 4.67-4.59 (m, 1H), 4.55-4.45 (m, 1H), 4.24-4.14 (m, 2H), 3.93-3.85 (m, 2H), 3.84-3.65 (m, 6H), 3.07 (s, 6H) .HRMS (EI): m / z calcd for C ₂₁ H ₂₅ FN ₂ O ₄ 388.1798; found 388.1805 (M + H ⁺ ).

(16) ^[18 F] 7 and ^[18 F] 15 procedure ^[18 F] labeled fluoride is produced by ^{^{18 O (p, n) 18}} F reaction by JSW TypeBC3015 cyclotron, ¹⁸ O enriched water The solution was passed through a Sep-Pak Light QMA cartridge (Waters). The cartridge was dried in a stream of air and the ¹⁸ F ^- activity was eluted with K ₂ CO ₃ solution (33 mM). Kryptofix ₂₂₂ (6-8 mg) was dissolved in [ ¹⁸ F] fluoride aqueous solution. The solvent was removed at 120 ° C. under a nitrogen stream. The residue was azeotroped twice with 0.3 mL anhydrous acetonitrile at 120 ° C. under a stream of nitrogen.

For [ ¹⁸ F] 7, a solution of mesylate precursor 10 (1.0 mg) in CH ₃ CN (0.2 mL) was added to the reactor containing ¹⁸ F ^- activity. The mixture was heated at 120 ° C. for 6 minutes. After the solution was cooled to room temperature, HCl (1 M aqueous solution, 0.2 mL) was added and the mixture was heated again at 120 ° C. for 5 min. An aqueous K ₂ CO ₃ solution was added to adjust the pH to basic (pH 8-9). Water (5 mL) was added and the mixture passed through a preconditioned Sep-Pak Plus PS-2 cartridge (Waters). The cartridge was washed with 20 mL water and the labeled compound was eluted with 3 mL acetonitrile. After the solvent was removed, the residue was dissolved in CH ₃ CN and subjected to HPLC for purification (Nakalai Tesque, 5C18-AR-II, 4.6 mm x 150 mm, CH ₃ CN / water = 1 / 1; flow rate = 1 mL / min). The retention time of [ ¹⁸ F] 7 in this HPLC system was 8.22 minutes (FIG. 18). The preparation took 60 minutes and the radiochemical yield was 30% (corrected for attenuation).

For [ ¹⁸ F] 15, a solution of tosylate precursor 14 in CH ₃ CN (0.2 mL) was added to the reactor containing ¹⁸ F ^- activity. The mixture was heated at 120 ° C. for 6 minutes. Water (5 mL) was added and the mixture passed through a preconditioned Sep-Pak Plus PS-2 cartridge (Waters). The cartridge was washed with 20 mL water and the labeled compound was eluted with 3 mL acetonitrile. The eluted compound was purified by HPLC (Nakalai Tesque, 5C18-AR-II, 4.6 mm × 150 mm). The retention time of [ ¹⁸ F] 15 in this HPLC system (CH ₃ CN / water = 6/4; flow rate = 1 mL / min) was 7.55 minutes (FIG. 19). Preparation took 40 minutes and the radiochemical yield was 60% (corrected for attenuation). The radiochemical purity of both tracers was over 98%.

(17) Measurement of partition coefficient Radiolabeled compound ([ ¹⁸ F] compound 7 or [ ¹⁸ F] compound 15) (10 μCi) was prepared by adding 3 g of n-octanol and 3 g of PBS (0.05 M, pH = 7.4). Added to the premixed suspension in the containing test tube. The tubes were vortexed for 3 minutes at room temperature and centrifuged for 5 minutes at 3000 rpm. Two weighted samples from the n-octanol (100 μL) layer and the buffer (500 μL) layer were measured. The partition coefficient was expressed as the logarithm of the ratio of counts per gram from n-octanol to PBS. Samples from the n-octanol layer were redistributed until consistent partition coefficient values were obtained. Measurements were made in triplicate and repeated three times.

(18) In vitro binding assay using Aβ aggregates (Figure 20, Table 8)
Inhibition experiments were performed on 12 × 75 mm borosilicate glass tubes according to a procedure modified from several previously described procedures. Briefly, 100 μL of aggregated Aβ fibers (final assay mixture concentration: 60 nM), 100 μL of radioligand ([ ¹²⁵ I] IMPY) at the appropriate concentration, 10 μL of inhibitor (concentration in ethanol of 10 ^-5 -10 ^-10 M.) and 790 μL PBS (0.2 M, pH = 7.4) and added to a mixture with a final volume of 1 mL. Nonspecific binding was defined in the presence of 1 μM IMPY. The mixture was incubated for 2 hours at 37 degrees with constant rocking. The bound and free radioactive fractions were then separated by vacuum filtration using a M-24 cell harvester (Brandel, Gaithersburg, MD) through a borosilicate glass fiber filter (Whatman GF / B). Radioactivity from the filter containing bound ¹²⁵ I ligand was measured with a γ-counter (WALLAC / Wizard 1470, USA) with 70% efficiency. Under the assay conditions, the specific binding fraction accounted for 10% of the total radioactivity. The half-maximal inhibitory concentration (IC ₅₀ ) was determined using GraphPad Prism 4.0, and the inhibition constant (K _i ) was determined using the Cheng-Prusoff equation: K _i = IC ₅₀ / (1 + [L] / K _d ) It was calculated.

(19) In vitro autoradiography using Tg mouse and human AD brain (FIGS. 21 and 22)
Paraffin-embedded mouse brain section (Tg: C57-APP / PS1, 12 months old, male; Wild-type: C57, 12 months old, male) and human brain section (AD: 93 years old, female; Normal: 71 years old, male) is 2 x 20 minutes in xylene, 2 x 5 minutes in 100% ethanol, 5 minutes in 90% ethanol / H ₂ O, 5 minutes in 80% ethanol / H washing in ₂ O, washed with 5 minutes 60% ethanol / H ₂ O in, and deparaffinized by washing in running water for 10 minutes in tap, then, PBS (0.2 M, pH = 7.4) in Incubated for 30 minutes. Sections were incubated with radiolabeled tracer (10 μCi / 100 μL) for 1 hour at room temperature, then washed with 50% ethanol for 1 hour and rinsed with water for 1 minute. After drying, the labeled sections were exposed to a Fujifilm imaging plate overnight. In vitro autoradiographic images were obtained using the BAS5000 scanner system (Fuji Film). The presence and location of plaques in the sections were confirmed by fluorescent staining with thioflavin S or immunohistochemical staining with BC05 (Wako), a monoclonal Aβ _1-42 antibody.

(20) In vivo biodistribution experiment using normal ddY mice (Tables 9 and 10)
Biodistribution experiments were performed in normal male ddY mice (5 weeks, 20-22 g, n = 5) and approved by the animal care committee of Kyoto University. Saline containing radiolabeled tracer (10 μCi / 100 μL saline, 5% EtOH) was injected directly into the tail. Mice were sacrificed at various times after injection. The target organ was removed, weighed, and the radioactivity was measured with an automatic γ counter (WALLAC / Wizard 1470, USA). The percent dose per gram of wet tissue was calculated by comparing tissue counts to appropriately diluted doses of injected material.

(21) using AD model mice ^[18 F] Compound 7 and ^[18 F] compounds 15 ex vivo autoradiography (Figure 23)
Autoradiography was performed using Tg2576 transgenic mice (female, 18-month-old) and wild-type mice (female, 18-month-old). 1.1 mCi of [ ¹⁸ F] 7 or [ ¹⁸ F] 15 saline (100 μL, 5% EtOH) was infused through the femoral vein in vivo, and the animals were sacrificed by decapitation 30 minutes later. The brain was removed and immediately frozen in powdered dry ice. 20 μm sections were cut and exposed to BAS imaging plates (Fuji Film, Tokyo, Japan) for 8 hours. Autoradiographic images were obtained using a BAS5000 scanner system (Fuji Film). After autoradiographic examination, the presence of amyloid plaques was confirmed by staining the same section with thioflavin S in vitro.

(22) [ ¹⁸ F] 7 micro-PET study on AD model mice and wild-type mice (FIGS. 24 and 25)
Female Tg2576 mice (C57BL6, 31-month-old) and female wild-type mice (C57BL6, 31-month-old) were anesthetized with isoflurane (1.5-2.5%) in oxygen at a flow rate of 1-2 L / min. 1.1 mCi [ ¹⁸ F] 7 (100 μL, 5% EtOH) was injected through the tail vessel, and dynamic microPET images were obtained in 60 minutes using filtered back projection with a RAMP filter. The data was analyzed using PMOD software and the desired quantities (VOIs) were defined for the summarized images and time activity curves (TACs) were drawn.

[Example 4] Near-infrared fluorescent compound having a thiophene ring or a furan ring (1) Synthesis of 5-dimethylamino-2-thiophenecarboxaldehyde (Compound 101) 5-Bromo-2-thiophenecarboxaldehyde (1 g, 5.23 mmol) ) And dimethylamine (1.64 mL, 15.69 mmol) were mixed, and water (3 mL) was added. The mixture was heated under reflux for 1 hour, extracted with chloroform, dehydrated by adding anhydrous sodium sulfate, and the solvent was evaporated under reduced pressure. The residue was purified by medium pressure preparative chromatography using ethyl acetate: hexane = 1: 1 as an elution solvent to obtain 203 mg (yield: 25.0%) of Compound 101. ¹ H NMR (400 MHz, CDCl ₃ ) δ3.09 (s, 6H), 5.93 (d, J = 4.6 Hz, 1H), 7.48 (d, J = 4.6 Hz, 1H), 9.49 (s, 1H).

(2) Synthesis of 5-dimethylamino-2-furaldehyde (Compound 102) 5-Bromo-2-furaldehyde (915 mg, 5.23 mmol) and dimethylamine (1.64 mL, 15.69 mmol) are mixed, and water (3 mL ) Was added. The mixture was heated under reflux for 1 hour, extracted with chloroform, dehydrated by adding anhydrous sodium sulfate, and the solvent was evaporated under reduced pressure. The residue was purified by medium pressure preparative chromatography using ethyl acetate: hexane = 3: 2 as an elution solvent to obtain 322 mg (yield: 44.3%) of Compound 102. ¹ H NMR (400 MHz, CDCl ₃ ) δ3.07 (s, 6H), 5.24 (d, J = 4.6 Hz, 1H), 7.20 (s, 1H), 8.97 (s, 1H).

(3) Synthesis of 2-((5- (dimethylamino) thiophen-2-yl) methylene) malononitrile (Compound 103, DTA-0) Compound 101 (118 mg, 0.76 mmol) was dissolved in 2-propanol to give malononitrile. (50 mg, 0.76 mmol) and pyridine (91 μL) were added, and the mixture was heated to reflux for 4 hours. After completion of the reaction, the reaction solution was cooled to room temperature, and the precipitated solid was collected by suction to obtain 28 mg (yield: 18.1%) of Compound 103. MS m / z 203 (M ⁺ ).

(4) Synthesis of (E) -3- (5- (dimethylamino) thiophen-2-yl) acrylaldehyde (Compound 104) Compound 102 (360 mg, 2.32 mmol) and (1,3-dioxalan-2-ylmethyl) ) Triphenylphosphonium bromide (1919 mg, 4.47 mmol) was dissolved in THF (10 mL), NaH (230 mg, 9.6 mmol) and 18-crown-6 (5 mg) were added, and the mixture was stirred at room temperature for 12 hours. Concentrated hydrochloric acid (1 mL) was slowly added, followed by neutralization with saturated NaHCO ₃ solution. After extraction with ethyl acetate and dehydration by adding sodium sulfate, the solvent was distilled off under reduced pressure. The residue was purified by medium pressure preparative chromatography using ethyl acetate: hexane = 1: 1 as an elution solvent to obtain 315 mg (yield: 74.9%) of Compound 104. ¹ H NMR (400 MHz, CDCl ₃ ) δ3.06 (s, 6H), 5.86 (d, J = 4.0 Hz, 1H), 6.47 (dd, J = 8.0, 15.0 Hz, 1H), 7.11 (d, J = 4.0 Hz, 1H), 7.40 (d, J = 15.2 Hz, 1H), 9.45 (d, J = 8.0 Hz, 1H).

(5) Synthesis of (E) -3- (5- (dimethylamino) furan-2-yl) acrylaldehyde (Compound 105) Compound 103 (287 mg, 2.06 mmol) and (1,3-dioxalan-2-ylmethyl) ) Triphenylphosphonium bromide (1704 mg, 3.97 mmol) was dissolved in THF (10 mL), NaH (205 mg, 8.51 mmol) and 18-crown-6 (5 mg) were added, and the mixture was stirred at room temperature for 12 hours. Concentrated hydrochloric acid (1 mL) was slowly added, followed by neutralization with saturated NaHCO ₃ solution. After extraction with ethyl acetate and dehydration by adding sodium sulfate, the solvent was distilled off under reduced pressure. The residue was purified by medium pressure preparative chromatography using chloroform: methanol = 99: 1 as an elution solvent to obtain 103 mg (yield: 30.2%) of Compound 105. ¹ H NMR (400 MHz, CDCl ₃ ) δ3.00 (s, 6H), 5.21 (d, J = 4.1 Hz, 1H), 6.22 (dd, J = 8.2, 14.7 Hz, 1H), 6.78 (d, J = 3.7 Hz, 1H), 6.96 (d, J = 15.1 Hz, 1H), 9.44 (d, J = 8.2 Hz, 1H).

(6) Synthesis of 2-((E) -3- (5- (dimethylamino) thiophen-2-yl) arylidene) malononitrile (Compound 106, DTA-1) Compound 104 (111 mg, 0.61 mmol) was converted to 2- Dissolved in propanol, malononitrile (40 mg, 0.61 mmol) and pyridine (73 μL) were added, and the mixture was heated to reflux for 17 hours. After completion of the reaction, water was added, the mixture was extracted with ethyl acetate, dehydrated with sodium sulfate, and the solvent was distilled off under reduced pressure. The residue was purified by medium pressure preparative chromatography using chloroform as an eluting solvent to obtain 93 mg (yield: 66.2%) of Compound 106. ¹ H NMR (400 MHz, CDCl ₃ ) δ3.15 (s, 6H), 5.96 (d, J = 4.6 Hz, 1H), 6.08 (dd, J = 12.4, 14.0 Hz, 1H), 7.18-7.21 (m , 2H), 7.33 (d, J = 11.9 Hz, 1H) .MS m / z 229 (M ⁺ ).

(7) Synthesis of 2-((E) -3- (5- (dimethylamino) furan-2-yl) arylidene) malononitrile (Compound 107, DFA-1) Compound 104 (38 mg, 0.23 mmol) was converted to 2- Dissolved in propanol, malononitrile (15 mg, 0.23 mmol) and pyridine (28 μL) were added, and the mixture was heated to reflux for 1 hour. After completion of the reaction, water was added, the mixture was extracted with ethyl acetate, dehydrated with sodium sulfate, and the solvent was distilled off under reduced pressure. The residue was purified by medium pressure preparative chromatography using chloroform: methanol = 99: 1 as an elution solvent to obtain 15 mg (yield: 30.6%) of Compound 107. ¹ H NMR (400 MHz, CDCl ₃ ) δ3.12 (s, 6H), 5.40 (d, J = 4.1 Hz, 1H), 6.55-6.70 (m, 2H), 6.93 (s, 1H), 7.29 (d , J = 11.9 Hz, 1H) .MS m / z 213 (M ⁺ ).

(8) Synthesis of (2E, 4E) -5- (5- (dimethylamino) thiophen-2-yl) penta-2,4-dienal (Compound 108) Compound 105 (315 mg, 1.73 mmol) and (1, 3-Dioxalan-2-ylmethyl) triphenylphosphonium bromide (1430 mg, 3.33 mmol) was dissolved in THF (10 mL), NaH (170 mg, 7.1 mmol) and 18-crown-6 (5 mg) were added, Stir at room temperature for 23 hours. Concentrated hydrochloric acid (1 mL) was slowly added, followed by neutralization with saturated NaHCO ₃ solution. After extraction with ethyl acetate and dehydration by adding sodium sulfate, the solvent was distilled off under reduced pressure. The residue was purified by medium pressure preparative chromatography using ethyl acetate: hexane = 1: 1 as an elution solvent to obtain 335 mg (yield: 92.9%) of Compound 108. ¹ H NMR (400 MHz, CDCl ₃ ) δ3.02 (s, 6H), 5.78 (d, J = 4.0 Hz, 1H), 6.08 (dd, J = 8.4, 14.6 Hz, 1H), 6.36 (dd, J = 10.8, 15.0 Hz, 1H), 6.92 (d, J = 4.0 Hz, 1H), 7.01 (d, J = 14.8 Hz, 1H), 7.16 (dd, J = 11.2, 15.0 Hz, 1H), 9.49 (d , J = 8.4 Hz, 1H).

(9) Synthesis of (2E, 4E) -5- (5- (dimethylamino) furan-2-yl) penta-2,4-dienal (Compound 109) Compound 106 (103 mg, 0.54 mmol) and (1, 3-Dioxalan-2-ylmethyl) triphenylphosphonium bromide (447 mg, 2.23 mmol) is dissolved in THF (10 mL), NaH (53.3 mg, 2.23 mmol) and 18-crown-6 (5 mg) are added, Stir at room temperature for 23 hours. Concentrated hydrochloric acid (1 mL) was slowly added, followed by neutralization with saturated NaHCO ₃ solution. After extraction with ethyl acetate and dehydration by adding sodium sulfate, the solvent was distilled off under reduced pressure. The residue was purified by medium pressure preparative chromatography using ethyl acetate: hexane = 1: 1 as an elution solvent to obtain 26 mg (yield: 21.8%) of Compound 109. ¹ H NMR (400 MHz, CDCl ₃ ) δ2.98 (s, 6H), 5.12 (d, J = 3.7 Hz, 1H), 6.12 (dd, J = 8.2, 14.7 Hz, 1H), 6.48-6.60 (m , 3H), 7.17 (dd, J = 10.5, 14.7 Hz, 1H), 9.50 (d, J = 7.8 Hz, 1H).

(10) Synthesis of 2-((2E, 4E) -5- (5- (dimethylamino) thiophen-2-yl) penta-2,4-dienylidene) malononitrile (compound 110, DTA-2) Compound 108 (83 mg, 0.4 mmol) was dissolved in 2-propanol, malononitrile (26.4 mg, 0.4 mmol) and pyridine (48 μL) were added, and the mixture was heated to reflux for 12 hours. After completion of the reaction, water was added, the mixture was extracted with ethyl acetate, dehydrated with sodium sulfate, and the solvent was distilled off under reduced pressure. The residue was purified by medium pressure preparative chromatography using chloroform as an elution solvent to obtain 62 mg (yield: 60.6%) of Compound 110. ¹ H NMR (400 MHz, CDCl ₃ ) δ3.09 (s, 6H), 5.87 (d, J = 4.4 Hz, 1H), 6.32 (dd, J = 11.6, 14.6 Hz, 1H), 6.57 (dd, J = 12.4, 13.7 Hz, 1H), 6.94-7.05 (m, 3H), 7.35 (d, J = 12.0 Hz, 1H). MS m / z 255 (M ⁺ ).

(11) Synthesis of 2-((2E, 4E) -5- (5- (dimethylamino) furan-2-yl) penta-2,4-dienylidene) malononitrile (Compound 111, DFA-2) Compound 109 (26 mg, 0.13 mmol) was dissolved in 2-propanol, malononitrile (8.5 mg, 0.13 mmol) and pyridine (16 μL) were added, and the mixture was heated to reflux for 1.5 hours. After completion of the reaction, water was added, the mixture was extracted with ethyl acetate, dehydrated with sodium sulfate, and the solvent was distilled off under reduced pressure. The residue was purified by medium pressure preparative chromatography using chloroform as an eluting solvent to obtain 12 mg (yield: 36.3%) of compound 111. ¹ H NMR (400 MHz, CDCl ₃ ) δ3.06 (s, 6H), 5.27 (d, J = 3.7 Hz, 1H), 6.47 (dd, J = 11.5, 14.2 Hz, 1H), 6.56-6.63 (m , 2H), 6.71 (d, J = 3.7 Hz, 1H), 6.98 (dd, J = 11.5, 13.7 Hz, 1H), 7.33 (d, J = 11.9 Hz, 1H) .MS m / z 239 (M ⁺ ).

(12) Synthesis of (2E, 4E, 6E) -7- (5- (dimethylamino) thiophen-2-yl) hepta-2,4,6-trienal (Compound 112) Compound 108 (335 mg, 1.62 mmol) And (1,3-dioxalan-2-ylmethyl) triphenylphosphonium bromide (1336 mg, 3.11 mmol) dissolved in THF (10 mL), NaH (159 mg, 6.6 mmol) and 18-crown-6 (5 mg ) Was added and stirred at room temperature for 23 hours. Concentrated hydrochloric acid (1 mL) was slowly added, followed by neutralization with saturated NaHCO ₃ solution. After extraction with ethyl acetate and dehydration by adding sodium sulfate, the solvent was distilled off under reduced pressure. The residue was purified by medium pressure preparative chromatography using ethyl acetate: hexane = 1: 1 as an eluting solvent to obtain 93 mg (yield: 24.7%) of Compound 112. ¹ H NMR (400 MHz, CDCl ₃ ) δ3.01 (s, 6H), 5.76 (d, J = 3.7 Hz, 1H), 6.09 (dd, J = 8.2, 15.1 Hz, 1H), 6.30 (dd, J = 11.0, 14.9 Hz, 1H), 6.38 (dd, J = 11.5, 14.7 Hz, 1H), 6.71-6.82 (m, 3H), 7.15 (dd, J = 11.5, 15.1 Hz, 1H), 9.52 (d, J = 7.8 Hz, 1H).

(13) Synthesis of 2-((2E, 4E, 6E) -7- (5- (dimethylamino) thiophen-2-yl) hepta-2,4,6-trienylidene) malononitrile (Compound 113, DTA-3) Compound 112 (93 mg, 0.40 mmol) was dissolved in 2-propanol, malononitrile (26 mg, 0.40 mmol) and pyridine (10 μL) were added, and the mixture was heated to reflux for 21 hours. After completion of the reaction, water was added, the mixture was extracted with ethyl acetate, dehydrated with sodium sulfate, and the solvent was distilled off under reduced pressure. The residue was purified by medium pressure preparative chromatography using chloroform as an eluting solvent to obtain 66 mg (yield: 53.5%) of Compound 113. ¹ H NMR (400 MHz, CDCl ₃ ) δ3.04 (s, 6H), 5.81 (d, J = 4.1 Hz, 1H), 6.30-6.37 (m, 1H), 6.60 (dd, J = 11.9, 14.0 Hz , 1H), 6.80 (dd, J = 11.5, 14.2 Hz, 1H), 6.87 (d, J = 15.1 Hz, 1H), 6.90 (d, J = 4.1 Hz, 1H), 6.96 (dd, J = 11.9, 14.2 Hz, 1H), 7.37 (d, J = 11.9 Hz, 1H) .MS m / z 281 (M ⁺ ).

(14) Measurement of fluorescence wavelength DTA and DFA derivatives were dissolved in chloroform so as to have a concentration of 10 µM, and the absorption, excitation, and fluorescence wavelength of each compound in chloroform were measured. The results are shown in the table below.

(15) Preparation of Aβ (1-42) aggregates PBS (pH 7.4) was used to prepare Aβ (1-42) at a concentration of 0.25 mg / mL. An Aβ (1-42) aggregate solution was prepared by incubating at 37 ° C. for 42 hours. The aggregate solution was stored at −80 ° C. until used for experiments.

(16) Fluorescence saturation experiment using Aβ aggregates: Calculation of K _d value PBS (168 μL) and sample solution (30 μL) diluted to 0-37.5 μM are mixed, and finally 25 μg / mL of Aβ aggregate is mixed. The collection solution (132 μL) was added and vortexed, and allowed to stand at room temperature for 30 minutes. Measurement was performed with a plate reader (Tecan) at excitation and fluorescence wavelengths suitable for each, and a saturation curve was created with Graph Pad Prism 4.0 to obtain a _Kd value. The results are shown in the table below.

DTA1-3 and DFA-1,2 had binding affinity for Aβ aggregates. On the other hand, DTA-0 showed no binding affinity for Aβ aggregates.

(17) Fluorescence staining using brain sections of Tg2576 mice Tg2576 mice (28 months old) were selected as model mice overexpressing amyloid precursor protein. DTA and DFA derivatives were diluted to 100 μM with 50% EtOH solution. After reacting with the compound solution (150 μL) for 5 minutes, it was washed twice with 50% EtOH for 1 minute × 2 and observed using a fluorescence microscope. Furthermore, the adjacent sections were reacted with thioflavin S solution (150 μL), washed and observed in the same manner as described above. The results are shown in FIG.

The five compounds except DTA-0 selectively bound to amyloid plaques accumulated on model mouse brain slices. Moreover, when it stained with thioflavin S using the adjacent section | slice, it corresponded with the dyeing | staining site | part of each compound. This result reflected the result of the fluorescence saturation experiment.

It can be used for the development of Aβ imaging reagents for the purpose of AD diagnosis, development support for therapeutic agents targeting Aβ, and disease state determination using Aβ accumulation in AD patients as an index.

This specification includes the contents described in the specification and / or drawings of the Japanese patent application (Japanese Patent Application No. 2011-182148) which is the basis of the priority of the present application. In addition, all publications, patents, and patent applications cited in this specification are incorporated herein by reference as they are.

Claims

General formula (I), general formula (II), or general formula (IV)

[In the formula, any one of R 1 and R 2 is an electron donating group, the other is an electron withdrawing group, X and Y are the same or different, and are a carbon atom, a nitrogen atom, an oxygen atom, or a sulfur atom. And Z is an oxygen atom or a sulfur atom, and m represents an integer of 0 to 5. ]
Or a pharmaceutically acceptable salt thereof.
The electron donating group is a hydroxyl group, a methoxy group, a methyl group, an amino group, a methylamino group, a dimethylamino group, a methylaminophenyl group, or a dimethylaminophenyl group, and the electron withdrawing group is a nitro group, a nitrovinyl group, a cyano group, The compound according to claim 1 or a pharmaceutically acceptable salt thereof, which is a dicyanovinyl group, a tricyanovinyl group, or a formyl group.
General formula (III)

[Wherein R 3 represents a hydroxy group, a C 1-3 alkoxy group, a C 1-3 alkoxy group containing a radioactive carbon atom, a halogen atom, a radioactive halogen atom, a chelate moiety bonded to a radioactive metal, or — (CH 2 CH 2 O) i -A [wherein i represents an integer of 1 to 10, and A represents a chelate moiety bonded to a halogen atom, a radioactive halogen atom, or a radioactive metal. R 4 represents a hydroxy group, a methoxy group, — (CH 2 CH 2 O) j —OH [wherein j represents an integer of 1 to 10. A group represented by the formula: — (CH 2 CH 2 O) k —F [wherein, k represents an integer of 1 to 10. Or —NRaRb [wherein Ra and Rb each independently represents a hydrogen atom or a C 1-3 alkyl group. And n represents an integer of 0 to 5. ]
Or a pharmaceutically acceptable salt thereof.
The compound or a pharmaceutically acceptable salt thereof according to claim 3, wherein the radioactive halogen atom is 18 F, 123 I, 124 I or 125 I.
The compound according to claim 3 or a pharmaceutically acceptable salt thereof, wherein the chelate moiety bound to the radioactive metal is a chelate moiety bound to 99m Tc or 68 Ga.
A composition for diagnosing conformation disease, comprising the compound according to any one of claims 1 to 5.