DD294707A5 - NEW BIOLOGICAL ACTIVE- (ALKYL) -P-AMINO-METHYLBENZOESAEURE PHENYLESTER - Google Patents

Abstract

The invention relates to new substances from the series of v-Aminomethylbenzoesaeureester (see formula I), in which unexpected biological effects have been found; The substances are effective in model experiments against inflammation, pain and a tumor model. They abolish the action of histamine and inhibit lipoxygenase, meaning: {new pharmaceuticals; Aminomethylbenzoesaeureester; pharmacological effect; Manufacturing facilities; anti-inflammatory drugs; analgesics; antihistamines; antitumor agents; lipoxygenase inhibitors; antiallergic}

Description

Field of application of the invention

These substances have not been described so far. Surprisingly, they show interesting pharmacological effects. According to their biological activity in model experiments, they can be used as antiphlogistic, analgesic, antiproliforative, antiallergic and mediator antagonistic drugs in human and veterinary medicine.

Characteristic of the known state of the art

For the treatment of inflammation and pain, mainly cyclooxygonase inhibitors or phenolic antioxidants have been used. Although the lipoxygenase inhibitors hitherto known have antiphlogistic, antiallergic and antiproliferative activity, their practical use is severely restricted because of their side effects. Strong antiproliferative compounds (including cancer chemotherapeutics) also have significant side effects that limit their use.

Object of the invention

The object of the invention is to use new substances for the treatment of inflammation, pain, allergies and hyperproliferations.

Explanation of the essence of the invention

The object underlying the invention is the discovery and production of new, hitherto unknown substances which are suitable for the treatment of inflammation, pain, hyperproliferations and allergies with low side effects.

Surprisingly, it has been found that substances of the above composition (see formula I) have very good therapeutic effects in the indications mentioned and are largely sparing of side effects.

) H CH, - (()) COO <("l> ^Λ

R ₂

where:

R i. R j = H, lower alkyl, aralkyl; R ₃ = alkyl (C ₁ to C ₇ , straight-chain and branched)

and their salts with R ₄ X, where:

R "= H, alkyl, aralkyl; X = halogen, toluenesulfonyl. These compounds have not previously been described. They are prepared by known methods, such as. In Houben Weyl VI / 3,53 and VIII, 340,508 and XI / 1,650 or detailed by analogy to Beilstein. According to the invention, the p-aminomethylbenzoic acid is converted into the N, N-dimethyl compound according to Leuckart-Wallach,

_HCDOH _{C ||}

H ^ N -CH ~ -; (V: - COOH fr- ^Ä : W -CH ~ - <i I) - COOH

^{2 2} - HCHO CH ₃ ² -

where temperatures up to 14O ⁰ C are applied.

This N, N-dimethylaminobenzoic acid is converted by means of thionyl chloride in the acid chloride, which after reaction with

tert-Butylphenol at 160 ⁰ C, the NN-dimethylaminobenzoic acid ^ '- tert-butylphenyl ester hydrochloride results.

The preparation of the quaternary salts with alkyl halides is carried out by reaction with the free base in nitromethane. The compounds thus synthesized, not previously described, were prepared according to generally accepted pharmacological Methods to investigate their biological activity. Surprisingly, a good antiphlogistic and analgesic effect was found. Likewise, representatives of these show

new substance series a very good inhibition of Ehrlich ascites carcinoma. In the model test will continue the

Histamine effect abolished. Finally, various üpoxygenasen are inhibited in vitro. Extensive pharmacological studies revealed no further biological activity (here

not detailed); no effect on the nervous or cardiovascular system could be detected. For example, special CNS tests, etc. Conducted: behavioral test, orienting toxicity (LD), electroconvulsive and

Pentetrazolkrampf. In all experiments and in an orienting toxicological test occurred at maximum administrable Doses no toxicity or other than the biological activities mentioned. Thus, the compounds surprisingly show a new spectrum of activity, which is characterized by particularly low Side effects is characterized. An antiphlogistic and analgesic effect with simultaneous antiallergic Active components suggests the absence of typical tolerability problems of classical anti-inflammatory drugs / analgesics. Mucosal irritations and z.T. severe pseudoallergic reactions, conclude. This should, in contrast to the

known drugs, such substances may also be suitable for the treatment of inflammation and pain with sufficiently strong action in allergic persons or for pseudoallergic reactions of predisposed persons.

According to the results of the model tests, a particularly good compatibility is to be expected. This is done by the Absence of allergenic reactions in guinea pigs, which after a standard procedure several times a 1% lge Substance solution was injected, further affirmed. Also noteworthy is the antihistaminic effect together with the Lipoxvgenasehernmung in the absence of CNS side effects. This is in practice an anti-allergic effect without impairments or good Compatibility expected. Favorable is the additional existing pain inhibition. Classic antiallergic drugs are common CNS side effects; Many previously known lipoxygenase inhibitors have no entry into the therapy because of their toxicity

found.

The inhibition of tumor growth is in the range of classical chemotherapeutic drugs, without approximating their general toxicity

to reach.

Ausführungsbelsplele Preparation Examples The chemical substances used for the investigations are prepared by known methods. You can

For example, obtained by the following methods:

Preparation of ^ (N, N-dimethylaminomethyll-benzoic acid, C'-tert-butylphenyl ester hydrochloride (Substance A) To 1 Z ml of formic acid are added with stirring at 0 to 5 ⁰ C 15.1 g of 4-aminomethylbenzoic acid. Subsequently

21 ml of formalin solution are added. The clear, colorless solution is slowly heated, resulting in carbon dioxide cleavage. The temperature rises to 85 ⁰ C by itself; then it is heated to 14O ⁰ C for 7 hours. After cooling it is with

Hydrochloric acid made congo and concentrated in vacuo to dryness. 20-21.5g of white crystals (= 92-99%) of Fp.256-263 ⁰ C. Without further purification, the product is C with the same amount of thionyl chloride for 1 hour at 6O ⁰ and

Heated to 75 ⁰ C for 3 hours. After stripping off the excess thionyl chloride, 4- (N ₁ N- ₎ is obtained in quantitative yield.

DimethylaminomethyD-benzoyl chloride as a yellowish-white solid, which is further processed without further purification. For this purpose, 5.5 g of this product with 3.9 g p-tert.-butylphenol at 160 ⁰ C are reacted and, after cooling, with ether

and then recrystallized from ethanol.

There are'ca. 6g 4- (N, N-dimethylaminomethyl) benzoic acid 4'-tert-butylphenyl ester hydrochloride as white crystals from Mp. 232-238 ⁰ C received. Preparation of quaternary salts of 4- (N, N-dimethylaminomethyl) benzoic acid 4'-tert-butylphenyl ester The free base is made from the Hydrochloric! by precipitation with caustic soda as a white solid

receive.

The quaternization is carried out by known methods, most conveniently in nitromethane, virtually quantitatively the

desired products are obtained, which are precipitated for purification from an acetone / ethanol solution with water. It was produced:

4- (N, N-Dimethylaminomethyl) benzoic acid 4'-tert-butylphenyl ester methobromide (Substance B) m.p.230-232 ° C

4- (N, N-dimethylaminomethyl) benzoic acid 4'-tert.biitvlphenylester-methiodide (substance C). M.p. 218-220 ⁰ C

4- (N, N-Dimethylaminomethyl) benzoic acid 4'-tert-butylphenyl ester ethoiodide (Substance D)

The elemental analyzes of these products corresponded to the expected values; DC studies showed no By-products. IR and NMR spectra confirmed the composition of the compounds. Methods for testing biological activity Kaolin edema of the rat paw Male or female Wistar rats (from Falcke / Barby) weighing between 100 and 200 g are used. The animals are deprived of food on the afternoon before the trial, so that they are fasted for about 16-18 hours. They receive drinking water ad libitum. Each experimental group comprises 10 animals. The control group receives the corresponding amount of water instead of the test substance

orally.

After subplantar injection of 0.1 ml of a 10% aqueous kaolin suspension (Bolus alba DAB7-DDR) in a Rat hindpaw develops an acute inflammation, which reached its maximum after approx. 7 hours. The test substances are given orally immediately after the inflammation has been triggered (1 ml / 100g BW). Test substances which are not water-soluble are used as aqueous suspensions with 2 drops of Tween 80 (Sorbimacrogolum

oleinicum, DAB7-DDR) per 5ml.

With a plethysmometer, the volume of the paw is before the start of the test and 3 hours after the inflammation

certainly.

The percentage difference in paw volume per animal becomes the mean difference in the standard deviation group

calculated.

Esslgsfiure-Writhlng Ten animals (male or female) are used per experimental group or control group. The animals will be 15 hours before Start of the experiment fasted; they get water ad libitum. The administration of the test substances is carried out p. o. (0.4 ml / 20g KM). 20 minutes after application, the 0.8% acetic acid i. p. (0.2 ml /

20g KM), after another three minutes, the writhings (abdominal cramps) are counted over 12 minutes.

The control group receives tap water or solubilizer instead of the test substance. The average number of Abdominal cramps must be at least 30 in the control group. For a smaller number, the mice are for this test

not suitable.

The number of writhings in the control group is assumed to be 100% effective. This results in the calculation of the Inhibition following equation:

% Inhibition = - 100

b '

a - Number of writhings of the experimental group

b - number of writhings of the control group

To calculate the ED ₆₀ , the dose-response curve is recorded with 3 to 4 doses in the linear range. Solid Ehrlich ascites carcinoma (EAC) 10 female ICR mice (18 to 22g, vomVEB Tierzuchtkombinat Schönwalde) with 0.2 ml of the group are used per group EAC cell suspension (6 million cells / ml) inoculated intracutaneously. From the next day, the substances are administered orally daily. The control group receives the appropriate amount

Water.

On the 8th day the animals are killed, the body weight is determined to control, and it becomes the solid ulcer of the Removed the abdominal skin and weighed immediately. The mean percentage difference of the tumor weight with standard deviation in relation to the Control group. The internal standard is a group that has been treated with cyclophosphamide. Hlstamlnlyse in vitro The guinea pig urchin prepared in Tyrode (Boomers, colorful, male, 250 ... 35Og KM) will be 15 to 30 minutes in the Organ bath (air bubbled, load 1 g, Tyrode, pH 7.38 to 7.42, 37 ° C) adapted. The histamine-induced contractions become isotonic via a transmitter system to laboratory writers

transfer. It is the effect of the substances against the submaximal histamine concentration (5 10 ~ ⁷ mol / l

Bath concentration, histamine dihydrochloride). As soon as a plateau of histamine contraction has formed, becomes

add one (further) dose of the test substance (accumulating) to the organ bath.

For each effective concentration, the mean percent inhibition is based on the 100% submaximal Histamine concentration, calculated. Inhibition of lipoxygenases Both the preparation of the peas · and the Retikulozytenlipoxygenase and the determination of the enzyme activity takes place

according to the literature (R.GRUPE, T.ZISKA, A. RICHTER, B.BÖHME and E.GÖRES, Biomed., Biochim, Acta [1988], 12, 955-963) based on methods of RAPOPORT and SCHEWE (T.GCHEWE, R WIESNER, SM RAPOPORT,

Methods Enzymol. 71 [1981] 430-441; D. REGEL, T. SCHEWE, S. M. RAPOPORT, Biomed. Biochim. Acta44 (1985) 1411-1428; H. KÜHN, T. SCHEWE, S. M. RAPOPORT, Biomed. Biochim. Acta 43 [1984] 358-361). The enzymes are obtained by fractionated ammonium sulfate precipitation gc. Their activity towards linoleic acid in aqueous Solution is measured with a Clark electrode. Results of the biological tests

1. kaolin edema

Substance C was used as the test substance. The ED _M values are given in mol / kg po. Test substance 1.6 ~ 10 ⁴ (1.4 x 10 ~ ⁶ ... 1.9 x 10 ^"3) Diclofenac-Na 1.7 × 10 ^-6 (1.2 × 10 -6 × 4.7 × 10 ^-6 ) Piroxicam 2.0 x 10 " ^B (3.9 x 10" ^e ... 9.7 x 10 " ⁶ ) Phenylbutazone 2.9 x 10 "(" ⁴ 1.2 x 10 "* ... 7.0 x ΙΟ ⁴⁾

2. Acetic acid writhing

Substance A was used as test substance and 2,3-dimethyl-4-methylamino-1-phenyl-5-pyrazolone-N-methanesulfonic acid as standard. The ED _M values are given in mol / kg po.

Test substance 3.3 x 10 ^"4 (3.2 x 10" ⁴ ... 3.5 x 10 ^"4)

Standard 8.7 x 10 ^"6 (2.7 x 10- ^e ... 2.9 x 10" ⁴⁾

3. Ehrlich-Ascltes-Carclnom

Substance A was used as test substance and cyclophosphamide as standard.

substance dose effect in mol / kg p.o. in% inhibition test substance 1 · 10 " ⁴ -77 ± 7 1 · 10 " ⁶ -80 ± 7 1 · ΙΟ " ⁸ -73 + 17 default 1.5-10 " ⁴ -70 ± 10

4. Hlstamlnlyse

The effects of substance C are: Dose effect

inmol / l in% inhibition

2 · 10 " ⁴ 100

2 · 10 " ⁶ 100

2 · 10 "'30

5. Inhibition of pea lipoxygenase

As test substance 1, substance B was used, as test substance 2 substance D and as standard propyl gallate. The effects are reported as ICw values in mol / l.

Substance 2.3 · 10- ^e (1.9 · 10 " ^B ... 2.6 · 1 (T ⁶ )

Substanz2 3.8 x 1 <T ⁵ {2.5 x 10- ⁶ ... 5.7 x 10 ^"6)

Standard 2.8 x 10 " ⁶ (2.2 x 1 (T ⁶ ... 3.6 x 10" ⁶ )

6. Inhibition of Reticulocyte Oxygenase

Substance D was used as test substance and phenidone as standard. The effects are given as IC _M values in mol / l.

Test substance 1.6 x 10 ^"4 (1.4 x 10 ~ ⁴ ... 1.8 x 10" ⁴⁾

Standard 1.3 x 1 ^<Γ β (8.9 x 10 ^"7 ... 1.7 x 1 ^<Γ β)

Claims (3)

1. New cD-iAlkyD-p-aminomethylbenzoic acid phenyl esters according to the general formula I _# where: R1 / R2 = Hj-lower alkylCralkyl-; R ₃ = alkyl (C ₁ to C ₇ , straight-chain and branched) and their salts with R4X, wherein: R ₄ = H, alkyl, aryl, aralkyl; X = halogen, toluenesulfonyk 2. Agents for inflammation, pain, allergies and hyperproliferations, characterized in that it contains in addition to pharmaceutically customary carriers and excipients u> (alkyl) -paminomethylbenzoesäure-phenyl ester. 3. A process for the preparation of novel substances of general formula I, characterized in that according to Leuckart-gelding from p-aminomethylbenzoic the N, N-dimethyl compound is formed, as is converted with thionyl chloride in the acid chloride, after which the ester mittert by reaction Butylphenol un '' iequatemären salts with alkyl halides by reaction with the free base in nitromethane.