JPS6239523A - Antienzyme - Google Patents

Abstract

PURPOSE:To provide an antienzyme containing 3,5-di-t-butyl-4-hydroxy-styrene derivative or its salt as an active component, effective to inhibit tyrosine kinase participating in the control of cell proliferation, and useful as a carcinostatic agent and anticarcinogenetic agent having low toxicity. CONSTITUTION:A tyrosine kinase inhibiting agent can be produced by using the compound of formula (R<1> is carboxy or carbamoyl; R<2> is 1-4C alkyl, CN, etc.) or its salt as an active component. The concrete example of the compound of formula is 3,5-di-t-butyl-4-hydroxy-alpha-hydroxyethylcinnamic acid, etc. The compound of formula is effective to inhibit tyrosine kinase participating in the proliferation and canceration of cell. It can be administered by oral, transintestinal or parenteral administration at a dose of preferably 20mg-5g/ day. The compound of formula can be produced by reacting 3,5-di-t-butyl-4- hydroxybenzaldehyde with the compound of formula II (Ar is aryl).

Description

[Detailed description of the invention] [Industrial application field] The present invention relates to tyrosine kinase inhibitors.

More specifically, 3 represented by the general formula (1) (where ILl represents a carboxyl group or a carbamoyi group, and R2 represents a C1 to C4 alkyl/I/ group, a cyano group, or a hydroxyethyl group) , 5-ditertiary-butyA/-4-hydroxystyrene derivative or a physiologically acceptable salt thereof as an active ingredient.

The present inventors have discovered various 3,5-ditertiary-buty A/-
When the physiological activity of 4-hydroxystyrene derivatives was investigated, it was discovered that the compound represented by the above-mentioned general formula (1) exhibits excellent physiological activity, particularly tyrosine kinase inhibitory activity, and the present invention was completed.

[Conventional technology]

Known tyrosine kinase inhibitors include 1, e.g.
There is Quercetin, but there are no reports of such inhibitory activity for 8,5-ditertiary-butytv-4-hydroxystyrene derivatives.

[Problem that the invention seeks to solve]

Recently, it has been discovered that a group of cell growth factor receptors that control cell proliferation have tyrosine kinase activity, and that oncogenic arrester genes encode proteins similar to cell growth factor receptors. It has become clear that there is a group of products that have tyrosine kinase activity. From the above, tyrosine kinases are deeply involved in the control of cell proliferation, and canceration, which is the unrestricted proliferation of cells, is a state in which tyrosine kinases are activated and cells are unable to enter the stationary phase of proliferation. It is said that Therefore, inhibiting tyrosine kinase in cells in such a state will lead to cancer control or prevention of carcinogenesis. pv-4-Hydroxystyrene derivatives have fewer side effects The tyrosine kinase inhibitor, anticancer agent, and anticancer agent according to the present invention is a 3,5-ditertiary-butyl-4-hydroxystyrene derivative represented by the general formula (1) or its The active ingredient is a physiologically acceptable salt, and specific examples of compounds represented by formula (1) include the following.

8.5-ditertiary-butyl-4-hydroxy-α-
Hydroxyethylcinnamic acid (hereinafter abbreviated as compound I) 8.5-ditertiary-butyl-4-hydroxy-α-
Methycinnamic acid (hereinafter abbreviated as compound ■) α-cyano-8,5-ditertiary-butyl-1v-4-hydroxycinnamic acid amide (hereinafter abbreviated as compound ■) Represented by general formula (1) The compound represented by formula (1) can form a salt with a base, and the base can be any base that can form a salt with the compound represented by formula (1). Specific examples of salts include (1) metal salts, especially alkali metals and alkaline earth metals;

(2) salts with aluminum, (3) salts with amines, especially salts with methylamine, ethylamine, diethylamine, triethylamine, pyrrolidine, piperidine, morpholine, hexamethyleneimine, pyridine, aniline, etc. , tyrosine kinase inhibitor, anticancer drug,
And as the anti-carcinogenic agent, physiologically acceptable salts may be selected from among these salts.

Synthesis of Compound The compound represented by the above general formula (1) can be synthesized by the following method.

■ The compound represented by the general formula (1) (R1 and R2 are the same as above) is 8
．． Obtained by reacting 5-ditertiary-butyA/-4-hydroxybenza V dehyde with a ylide compound represented by general formula (2) (Mr represents an aryl group, R1 and R2 are the same as above) I can do things.

■ Among the compounds represented by the general formula (1), the general formula (
R1 is the same as above; R8 is a C1-C4 alkyl group or a cyano group), and the S compound is 8,5-ditertiary-buty-A/-4-hydroxybenzaldehyde and the general formula (4)% A compound represented by formula % (4) (R1 and R8 are the same as above), or general formula (5) % formula % (R4 is a C1-C4 alkyl group, R6 is OH or NH
2) in the absence of a catalyst or with an acid or base as a catalyst, and optionally an acid anhydride represented by the general formula (6) (R4 is the same as above). It can be obtained by adding something to cause a reaction.

Examples of acids that can be used as catalysts include protonic acids such as sulfuric acid, benzenesulfonic acid, and 1)-)A/enesulfonic acid; Lewis acids such as boron trifluoride. Bases that can be used as catalysts include monoethanolamine, heptalporine, pyridine, 1.8-
Organic bases such as azabicycloC5,4,0)undec-7-2; acetic acid, alkali metal salts of organic acids such as potassium acetate; alkali metal hydroxides such as sodium hydroxide and potassium hydroxide; lithium Examples include alkali metal amides such as diisopropyl chloride; alkali metal acholates such as sodium methylate and sodium ethylate; and alkali metal hydrides such as sodium hydride and potassium hydride. Further, the hydroxyl group at the 4-position of 3,6-ditertiary-butyl-4-hydroxybenzaldehyde is an alkyl group, a benzyl group, or an acyl group.

After protection with a trialkylsilyl group or the like, the reaction may be performed, and after the reaction is completed or during the reaction, these protecting groups may be removed by a known method.

■ The compound represented by the above-mentioned general formula (1) has 8.5
-Ditertiary-butyl-4-hydroxybenzaldehyde and a compound represented by the general formula (7) % formula % (7) (R2 is the same as above, R6 represents a C1 to C4 alkyl group) are combined with sodium hydride, sodium In the presence of a basic catalyst such as amide or sodium ethylate, or with the general formula (M1 and R11 are the same as above)
The general formula (9) obtained by reacting with the ylide represented by
) (R2 and R6 are the same as above) by hydrolysis with a base such as sodium hydroxide or an acid such as sulfuric acid, or by ammonolysis with ammonia.

In addition, in the first step reaction, the hydroxyl group of 8,5-ditertiary-butyl-4-hydroxybenzaldehyde is protected with an alkyl group, an acyl group, a trialkylsilyl group, etc., and then the reaction is carried out. , these protecting groups may be removed by known methods.

@ Among the compounds represented by general formula (1), general formula α
The compound represented by O (R" is the same as above) is 8.6-
Obtained by reacting ditertiary-buty Iv-4-hydroxybenza V dehyde and γ-butyrolactone in the presence of a base catalyst such as sodium hydride, sodium amide, sodium ethylate, or with a ylide represented by , formula hydroxide) can be obtained by hydrolysis with a base such as IJium or by ammonolysis with ammonia. In addition, when reacting with γ-butyrolactone in the first stage reaction, the hydroxyl group of 8,5-ditertiary-butypv-4-hydroxybenzaldehyde is protected with an alkyl group, an acyl group, or a trialkylsilyl group, and then the reaction is performed.
During or after the reaction, these protecting groups may be removed by a known method.

Enzyme Inhibitory Effect The tyrosine kinase inhibitor according to the present invention tζ contains a compound represented by the above general formula (1) or a salt thereof as an active ingredient. The 1118 inhibitory effect and toxicity of these compounds are as shown in the experimental examples below. Specifically, the tyrosine kinase inhibitory effect was measured by the method shown below.

The tyrosine kinase inhibitory effect of the compound of the present invention is 8
．． Tyrosine kinase activity measurement method by C0hen et al. [The Journal of Biological Chemistry (
J, Biol, Chem, ), 2 mutual units, 1528
(1982)) was measured as #horse. That is, human cancer cell-derived established strain Mu-481 was treated with 10% fetal bovine serum, streptomycin (50 al/ml'), and penicillin G.
(50 international units/ml') and kanamycin (50 μI
/ml') in Dulbecco's modified Eagle medium [Hakusui Pharmaceutical ■] at 87°C and 5% CO! cultured under the following conditions.

The obtained cells were treated according to the method of Cohen et al. in 9 above to obtain a membrane preparation containing the epidermal growth factor receptor-tyrosine kinase complex (hereinafter abbreviated as membrane preparation).

This standard sample was used for the following measurements without solubilizing it.

N-2-hydroxyethylpiperazino-N-2-ethanesulfonic acid buffer (2omn, pu7.41Mn(3
Add the sample dissolved in dimethyl sulfoxide to 1g (1mM) and bovine serum albumin c7.5plχ membrane preparation (10μl as protein), and after incubating for 5 minutes at 0°C, add epidermal growth factor (hereinafter abbreviated as EGF) ( 10
011,F) and incubated for 16 minutes at 0°C. Next, [γ-32p] TP (80000i
/mmol, 0.1 μCi) to make a final 70μ
After further incubation at 0° C. for 15 minutes, 50 μl of the reaction solution was impregnated onto Watfu 8MMF paper, and the reaction was immediately stopped with a 10% trichloroacetic acid-10 mM sodium pyrophosphate aqueous solution. One paper was thoroughly washed with the same wave, then washed with ethanol, dried, and the radioactivity remaining on the P paper was measured using a liquid scintillation counter, and this value was designated as A. At the same time, as controls, a reaction without the addition of EGF, a reaction without the addition of the sample, and a reaction without the addition of EGF and the sample were performed, and the same measurements were performed for each B.
, O and D.

The tyrosine kinase inhibition rate was determined by the following formula.

Table 1 shows the tyrosine kinase inhibitory effects of the compounds according to the present invention. This result shows that the compound according to the present invention strongly inhibits tyrosine kinase.

Table 1: Acute toxicity IOR female mice (body weight 28-26I) were used, with 6 mice per group. 0.1 ml of compounds (I) to (In) suspended in a 2.5% gum arabic aqueous solution containing 062% Tween 80! /10, F body weight was administered orally. For two weeks after administration, general symptoms were observed to determine the number of deaths/tested cases, and the 50% lethal dose LD50 (ml
/Kll) was estimated. As a result, the compound C of the present invention
No deaths were observed for compounds (I) to (Ill) even after 1,000 days of administration, and the LD50 of compounds (I) to (m) was 1.
000 m, estimated to be over 9M, and was found to be of low toxicity.

Preparation and Dosage The tyrosine kinase inhibitor, anticancer agent, or anticancer agent according to the present invention can be formulated in any of oral, rectal, or parenteral administration. Specific formulations include tablets, capsules, fine granules, syrups, crude drugs,
Examples include ointments and injections. The carrier for the formulation of the tyrosine kinase inhibitor, anticancer agent, or anticancer agent according to the present invention may be an organic or inorganic solid or liquid, usually inert, suitable for oral, rectal, or other oral administration. A pharmaceutical carrier material is used. Specifically, for example, crystalline cellulose, gelatin, and lactose.

Starch, magnesium stearate, talc, vegetable and animal fats and oils, gums, polyalkylene glycols. The ratio of the tyrosine kinase inhibitor of the present invention, anticancer agent 1, and anticancer agent to the carrier in the formulation can be varied between 0.2 and 100%. Furthermore, the tyrosine kinase inhibitor, anticancer agent, and anticancer agent according to the present invention may be used with other compatible tyrosine kinase inhibitors, anticancer agents, anticancer agents,
Other medicaments may be included. In this case, it goes without saying that the tyrosine kinase inhibitor, anticancer agent, or anticancer agent of the present invention does not need to be the main ingredient in the preparation.

The tyrosine kinase inhibitor, anticancer agent, and anticancer agent according to the present invention is generally administered at a dosage that achieves the desired effect without side effects. The specific value should be determined by a doctor, but in general, it is 10 per day for adults.
m, p~10,9. Preferably, it will usually be administered at about 20m, p~51. The tyrosine kinase inhibitor, anticancer agent, and anticancer agent of the present invention can be administered as a pharmaceutical preparation with an active ingredient of 1 my to 511, preferably Bmy to II.

〔Example〕

Next, the present invention will be specifically explained with reference to production examples and examples of the compounds of the present invention, but these examples are not intended to limit the present invention.

Production Example 1 Synthesis of Compound Cutting 8. 5-Ditertiary-butytv-4-hydroxybenzylidehyde 18F (!: α-TripheniM diphosphoranylidene-butyrolactone 27g and dimethyl sulfoxide (DM80) 150fnl! and reacted for 200 hours with stirring at 80°C on a hot water bath.After the reaction was completed, chloroform aoomI! was added to the cooled reaction solution.
The DM80 solvent was removed by washing 5 times with the same amount of water.

After separating the chloroform layer, it was concentrated to dryness under reduced pressure to remove chloroform. Ethanol was added to the residue to perform crystallization, followed by recrystallization from the same solvent to obtain α-(8,5-ditertiary-buty-4-hydroxybenzylidene)-γ.
-Butyrolactone 18! I got i. Next, 1.51 of the a-(8,5-ditertiary-butyl-4-hydroxybenzylidene)-r-butyrolactone obtained here was added to 0.
A hydrolysis reaction was carried out for 1 hour with stirring in an aqueous solution of IJ.

After the reaction was completed, 10% certified acid was added little by little to the cooled reaction solution I to make it acidic and to form a precipitate. The precipitate was separated by F and thoroughly washed with water. The precipitate was dissolved in benzene and crystallized to obtain 1.0 g of the target compound 1.

Production example 2 Compound! ! II [8.5-ditertiary-butylene/v
-4-hydroxybenza M dehyde 8.5,7.

Cyanoacetamide 1.8ji and piperidine 1mJ
was added and mixed, and the mixture was stirred at 40 to 50°C. After a while, the solution became homogeneous, and the reaction was continued for 4 hours with further stirring. After the reaction is complete.

Remove the solvent under reduced pressure and dissolve the residue in chloroform,
Washed 8 times with diluted hydrochloric acid. After separating the chloroform layer, chloroform was removed under reduced pressure, and the residue was dissolved in ethanol and crystallized. Yield 8.5, @.

Example 1 Compound 1100. A mixture of P, 55I lactose and 41P dry high lead seaweed starch was kneaded with 20ml of water, extruded through a 16 mesh screen, dried at 40°C and granulated. Next, it was mixed uniformly with magnesium stearate 4Ii and compressed into tablets by a conventional method, each having a size of 200 m
, q was obtained.

Example 2 Granules 196y obtained in exactly the same manner as in Example 1 were mixed with magnesium stearate 41, and then mixed with 200
Fill 2 Xiudan capsules with m, 9 each. A hard capsule containing Compound I in Loom, 9 was prepared.

Example 8 Compound I 10. O. p milk *
85. Ojjt Crystalline cellulose 4.5 g Magnesium stearate 0.5 g The above components were thoroughly mixed to obtain a powder containing too much of Compound I in 1i.

Patent applicant Makoto Asano, agent of Kanebuchi Kagaku Kogyo Co., Ltd., patent attorney - Procedural amendment (2) J-) March 27, Showa IT naa+69'p#tata〒 Application No. 179095 2.
Name of invention ψBudong! L plaster I address
3-2-4 Nakanoshima, Kita-ku, Osaka City
o q bell rr: + tHigaku Kogyo Co., Ltd. Representative Masato Niino 4, Agent name (693m) Patent attorney Makoto Asano -5
, date of correction order t=irbqζ

Claims (1)

[Claims] (1) 3,5-jiter represented by the following general formula (1)
A tyrosine kinase inhibitor containing a shari-butyl-4-hydroxystyrene derivative or a physiologically acceptable salt thereof as an active ingredient. ▲There are mathematical formulas, chemical formulas, tables, etc.▼ (1) (Here, R^1 represents a carboxyl group or carbamoyl group, and R^2 represents a C_1 to C_4 alkyl group, cyano group, or hydroxyethyl group.)