WO2000063215A2

WO2000063215A2 - Indazole derivatives, preparation and therapeutic application thereof

Info

Publication number: WO2000063215A2
Application number: PCT/FR2000/000923
Authority: WO
Inventors: Lydia Zard; Serge Perard
Original assignee: Sanofi-Synthelabo
Priority date: 1999-04-16
Filing date: 2000-04-11
Publication date: 2000-10-26
Also published as: FR2792318A1; AU3971500A; WO2000063215A3; FR2792318B1

Abstract

The invention concerns a compound of general formula (I) wherein: R1 represents a hydrogen atom, a halogen, a C1-C4 alkoxyl, methyl, hydroxy or amino group; R2 represents -(CH2)2-OH or-CH2-C(O)OH when R3 represents a hydrogen atom, or R2 represents a hydrogen atom when R3 represents (CH2)2-OH or -CH2-C(O)OH, or R2 and R3 are bound to form one of the cycles represented by (a) and (b); X represents an oxygen atom, a -NH- or- N(C1-C4 alkyl)-group; and A is a group of formula (B) or (C). The invention is applicable in therapeutics.

Description

INDAZOLE DERIVATIVES, THEIR PREPARATION AND THEIR APPLICATION

THERAPEUTIC

The subject of the present invention is indazole derivatives, their preparation and their therapeutic application, more particularly as antagonists of the 5-HT ₃ /5-HT ₄ receptors.

Consequently, the first subject of the present invention is a compound of general formula (I):

in which: Ri represents a hydrogen atom, a halogen, a Cι- ₄ alkoxy, methyl, hydroxy or amino group, R ₂ represents - (CH ₂ ) ₂ -OH or -CH ₂ -C (0) OH when R ₃ represents a hydrogen atom, or R ₂ represents a hydrogen atom when R ₃ represents- (CH ₂ ) ₂ -OH or -CH ₂ -C (0) OH, or R ₂ and R ₃ are linked to f below :

X represents an oxygen atom, a group -NH- or -N (Cχ-alkyl) -, and A is a gr

BC in which:

Y represents a nitrogen atom or a methylene group, R ₄ represents a hydrogen atom, a Cι_ ₄ alkyl group, or a benzyl, n is equal to 0 or 1, and m is equal to 0 or 1. Another subject of the invention is represented by the compounds of formula (II) _

in which: A, X, m, n and Ri are as defined above and

R ₅ and Rβr, identical or different, represent a hydrogen atom, a Cι_ ₄ alkyl group, a phenyl or a benzyl.

The preferred compounds according to the invention are the compounds of formula (I) for which A represents a group of formula C, more particularly those for which A represents a group of formula C and X represents an oxygen atom. Among the latter, the compounds for which m = 0 and n = 0 are preferred and more particularly those for which R ₄ represents a Cι_ alkyl group methyl. Among these, the compounds of formula (I) in which R ₂ represents - (CH ₂ ) ₂ -OH or -CH ₂ -C (0) OH, more particularly -CH ₂ -C (0) OH, when R ₃ represents a hydrogen atom, are preferred.

Mention may more particularly be made of the following preferred compounds:

- 8-hydroxy-7,8-dihydropyrazolo [1,5,4-de] [1,4] benzoxazine-2-carboxylate of endo-8-methyl-8-azabicyclo [3.2.1] oct-3-yl;

- 7-hydroxy-1- (2-hydroxyethyl) indazole-3-carboxylate of endo-8-methyl-8-azabicyclo [3.2.1] oct-3-yl;

- 3- (endo-8-methyl-8-azabicyclo [3.2.1] oct-3-yl-oxy-carbonyl) -7-hydroxyindazole-1-acetic acid;

- 7- (2-hydroxy-1-ethyloxy) indazole-3-carboxylate of endo-8-methyl-8-azabicyclo [3.2.1] oct-3-yl; - endo-8-methyl-8-azabicyclo [3.2.1] oct-3-yl 7-carboxymethoxyindazole-3-carboxylate; and

- 7, 8-dihydropyrazolo [1, 5, 4-de] [1, 4] benzoxazine-2-carboxylate of endo-8-methyl-N-oxide-8-azabicyclo [3.2.1] oct-3-yl. In this application:

- Cι_4 alkyl represents a carbon chain which can have from 1 to 4 carbon atoms, such as for example a methyl, ethyl, propyl, isopropyl, butyl, isobutyl or tert-butyl,

the term alkyl or alkoxy represents respectively an alkyl or alkoxy with a linear or branched carbon chain, and

- halogen represents an iodine, bromine, chlorine or fluorine atom.

The compounds of general formula (I) or (II) may contain one or more asymmetric carbon atoms. They can therefore exist in the form of enantiomers or diastereoisomers. These enantiomers, diastereoisomers, as well as their mixtures, including racemic mixtures, form part of the invention.

The compounds of general formula (I) can be in the form of free base or of addition salts with acids, which also form part of the invention. These salts, according to the present invention, include those with mineral or organic acids which allow proper separation or crystallization of the compounds of formula (I), such as picric acid, oxalic acid or an optically active acid, by example a tartaric acid, a dibenzoyltartaric acid, a mandelic acid or a camphosulfonic acid, and those which form physiologically acceptable salts, such as the hydrochloride, the hydrobromide, the citrate, the sulfate, the hydrogen sulfate, the dihydrogen phosphate, the maleate, fumarate, pamoate, 2-naphthalenesulfonate, paratoluenesulfonate. Although the pharmaceutically acceptable salts are preferred, the other salts are part of the present invention. These salts can be prepared, according to methods known to a person skilled in the art, for example, by reaction of the compound of formula (I) in base form with the acid in an appropriate solvent, such as an alcoholic solution or a organic solvent, then separation of the medium which contains it by evaporation of the solvent or by filtration. The compounds of general formula (I) can also be in the form of N-oxide derivatives which form part of the present invention. These derivatives are obtained by oxidation reaction of the compound of formula (I) according to methods known to those skilled in the art.

A second subject of the present invention is processes for preparing the compounds of formula (I).

Thus, these compounds can be prepared by methods, illustrated in the diagrams which follow, the operating conditions of which are conventional for a person skilled in the art.

The compounds of the invention can be prepared by biotransformation of the compounds of formula (III)

in which Ri, R ₃ , Rβ, A and X are as defined for the compound of formula (I) or (II). More particularly, the compounds of formula (I) are prepared by biotransformation of the compounds of formula (III) in which R ₅ and R ₆ each represent a hydrogen atom.

Thus, the compounds of formula (I) or (II) can be prepared by biotransformation of the compounds of formula (III) in vi tro by incubation with hepatocytes in primary culture according to methods known to those skilled in the art.

Alternatively, the compounds of formula (I) in which R ₂ represents -CH ₂ -C (0) 0H when R ₃ represents a hydrogen atom, can be prepared by microbiological oxidation according to methods known to those skilled in the art , for example, by enzymatic oxidation in the presence of cytochrome P450 of filamentous fungi.

The compounds of formula (III) are described in patent application PCT / FR98 / 02221.

The following examples illustrate the methods and techniques suitable for the preparation of this invention, without however limiting the scope of the claim. Mass spectra, NMR and / or microanalyses confirm the structures. EXAMPLE 1:

1.1 Preparation of hepatocytes

Mouse and rat hepatocytes were prepared according to the procedure after dissociation with collagenase. Human hepatocytes were prepared from biopsies obtained by the Biochemistry Laboratory of the Paul Brousse Hospital in Villejuif in agreement with the ethics committee of the Hospital. The cryopreserved dog and monkey hepatocytes were obtained from Bioprédic (Liverbeads) (Rennes, France).

1.1 Preparation of the compounds according to the invention The hydrochloride of 7, 8-dihydropyrazolo [1, 5, 4- of] [1, 4] benzoxazine-2-carboxylate of endo-8-methyl-8-azabicyclo [3.2.1] oct-3-yl is incubated with hepatocytes of the selected species at a concentration of 20 μM. After 24 h of incubation, the compounds of formula (I) or (II) were isolated by HPLC (Kromasil C ₈ 5μ column, eluent: 15mM ammonium acetate, pH4 / acetonitrile) from the extracellular supernatant and identified by LC / ESI-MS (Electrospray, HP / MSD, Hewlett Packard) and by LC / ESI-MS / MS (Quattro I, VG). (see tables)

EXAMPLE 2

240 mg of 7,8-dihydropyrazolo hydrochloride [1,5,5-4 of] [1,4] benzoxazine-2-carboxylate of endo-8-methyl-8-azabicyclo [3.2.1] oct-3-yl are incubated in the presence of a strain of filamentous fungi Cunninghamella echinula ta NRRL 3655 for one month at 27 ° C, with orbital shaking. The biomass is then removed by filtration, and the partially evaporated filtrate is extracted with ethyl acetate. The aqueous phase containing the 7- (carboxymethoxy) -1H-indazole-3-carboxylate

8-methyl-8-azabicyclo [3.2.1] oct-3-yl is evaporated to dryness and then washed with a dichloromethane / methanol mixture. The insoluble material is drained and the concentrated filtrate is purified by chromatography on silica gel (elution solvent: dichloromethane 80% / methanol 20%). The fractions containing 8-methyl-8-azabicyclo [3.2.1] oct-3-yl 7- (carboxymethoxy) -lH-indazole-3-carboxylate are repurified by preparative HPLC (Kromasil C8, 5 μ) to give 15 mg 7 - (carboxymethoxy) -1H-indazole-3-carboxylate of endo-8-methyl-8-azabicyclo [3.2.1] oct-3-yl MS (API, ES ⁺ ): m / z 360 (M + 1); 301; 124 Elementary composition: m / z = 360.1536 (wedge 360.1524) for CιsH ₂ ιN ₃ 0 RMN ^λ R (in ppm): 7.7 (d, IH); 7.2 (t, 1H); 6.8 (d, 1H); 5.3

(m, 1H); 4.55 (s, 2H); 2.5 (s, 3H); 2-2.5 (m, 8H).

¹³ C NMR (in ppm): 68.6 (t); 70 (d); 107 (d); 114.3 (d); 125.6

(d); 137 (s); 147 (s); 166 (s); 178 (s).

EXAMPLE 3

By applying the methods described according to Example 1 or 2, other compounds according to the invention can be synthesized.

The tables which follow illustrate the compounds obtained according to example 1 or 2.

Table 1

In column A "C" indicates that compound C is in endo form.

Table 2

(II) n ° Ri R ₅ Re R ₄ X mn MS (API, ES ⁺ ) (M + l)

6 HHH CH ₃ 0 0 0 344 The compounds of formula (I) according to the invention have been the subject of pharmacological tests which have shown their interest as active substances in therapy.

In particular, they have been tested for their inhibitory effects on the binding of [ ³ H] - (S) -zacopride with serotonergic receptors of the 5-HT _{3 type} of the rat cortex, according to the method described by NM Barnes et al. , in J. Pharm. Pharmacol. , 40, 548-551 (1988). Male Sprague-Dawley rats (OFA, Iffa credo) weighing 200 to 250 g are euthanized and their brains removed. The cortex is dissected and homogenized using a Polytron ^® mill (position 7.20 s) in 20 volumes of 25 mM Tris buffer (pH = 7.4, 22 ° C). The homogenate is centrifuged for 10 min at 45000xg (in a SORVALL centrifuge fitted with a rotor

SS34), then the pellet is resuspended in 10 volumes of Tris buffer and incubated at 37 ° C for 10 min with shaking. The suspension is then diluted to 20 volumes using Tris buffer and centrifuged under the same conditions as above. The pellet obtained is resuspended in 5 volumes of Tris buffer and then distributed into aliquots of 5 ml which are frozen at -80 ° C. On the day of the experiment, the preparation is thawed at 4 ° C. and then diluted 1.2 times with the Tris-NaCl incubation buffer (25 mM Tris, 150 mM NaCl, pH = 7.4, 22 ° VS) .

The membrane suspension (100 μl, 1 mg of proteins) is incubated at 25 ° C for 25 min in the presence of 0.5 nM of [ ³ H] - (S) -zacopride (specific activity: 75-85 Ci / mmol, Amersham, Little Chalfont, United Kingdom) in a final volume of 500 μl of Tris-NaCl buffer, in the absence or in the presence of the compound to be tested.

Incubation was stopped by filtration using Whatman GF / B ^® pretreated with polyethylenimine (0.1%). Each reaction tube is prediluted with 4 ml of Tris-NaCl buffer and then rinsed 3 times with 4.5 ml of Tris-NaCl buffer.

The filters are precut before drying in the oven (120 ° C, 5 min). The radioactivity retained on the filters is measured by liquid scintigraphy. Nonspecific binding is determined in the presence of 10 μM of MDL 72222 (ligand described in the article NM Barnes et al. referenced above).

For each concentration of compound studied, the percentage of inhibition of the specific binding of [ ³ H] - (S) -zacopride is determined, then the concentration of the compound inhibiting 50 ′ of the specific binding of [3H] - (S) -zacopride (CI ₅₀ ). The IC50 values of the compounds of the invention are between 0.5 nM and 3 μM.

The compounds of the invention were studied as to their antagonistic effects with respect to the 5-HT ₃ receptors of the smooth muscle of the descending colon isolated from guinea pigs, according to the method described by Grossman et al. in Br. J. Pharmacol. (1989) 97,451.

Serotonin (0.1-100 μM), after blocking of receptors of 5-HTι and 5-HT ₂ types (Methysergide 0.1 μM) and desensitization of 5-HT ₄ receptors (5-methoxytryptamine 10 μM) causes a contraction , dependent on the concentration, of the smooth muscle part of the descending guinea-pig colon, by stimulation of the 5-HT ₃ receptors. Contractions are recorded in isometry. The antagonistic effect of a compound on the 5-HT ₃ serotonergic receptors is quantified by measuring the displacement of a control effect-concentration curve of serotonin (successive increasing concentrations not accumulated), at concentrations between 1 nM and 0 , 1 μM, with an incubation of 30 min.

The compounds of the invention have also been studied as regards their affinity with respect to 5-HT ₄ receptors in the guinea pig striatum, according to the method described by Grossman et al., In Br. J. Pharmacol. , 109, 618-624 (1993). Guinea pigs (Hartley, Charles River) weighing 300 to 400 g are euthanized and their brains removed. The striata are excised and frozen at -80 ° C. The day of the experiment, the tissue is thawed at + 4 ° C. in 33 volumes of 50 mM Hepes-NaOH buffer (pH = 7.4 at 20 ° C.) and it is homogenized using a Polytron mill. ®. Centrifuge the homogenate for 10 min at 48000xg, recover the pellet is resuspended and centrifuged again under the same conditions. The final pellet is suspended in Hepes-NaOH buffer (30 mg of fresh tissue / ml). This membrane suspension is used as it is. 100 μl of the membrane suspension are incubated at 0 ° C for 120 min, in the presence of 0.1 nM of [ ³ H] GR113808 (specific activity: 80-85 Ci / mmol), in a final volume of 1 ml of buffer Hepes-NaOH (50 mM, pH = 7.4), in the absence or in the presence of the test compound. The incubation is stopped by filtration through Whatman GF / B® filters, previously treated with 0.1% polyethyleneimine, each tube is rinsed with 4 ml of buffer at 0 ° C. and filtered again. The radioactivity retained on the filters is measured by liquid scintigraphy. The non-specific binding is determined in the presence of 30 μM serotonin. The specific binding represents 90% of the total radioactivity recovered on the filter. For each concentration of compound studied, the percentage of inhibition of the specific binding of [ ³ H] GR113808 is determined, then the concentration of the compound tested which inhibits 50% of the specific binding (IC50) •

The IC 50 values of the compounds of the invention are between 1 nM and 2 μM.

Finally, the compounds of the invention were studied as to their agonist or antagonist effects with respect to the 5-HT ₄ receptors in the rat esophagus according to the method described by Baxter et al., In Naunyn Schmied Arch. Pharmacol. (1991), 343, 439.

Male Sprague Dawley rats weighing 300 to 450 g are used. We quickly remove a fragment of about 1.5 cm from the terminal part of the esophagus, we eliminate the muscular layer, we open the inner muscular mucous membrane longitudinally, we mount it in a tank with isolated organ containing a Krebs solution -Henseleit at 32 ° C oxygenated by a carbogenic current (95% 0 ₂ and 5% C0 ₂ ), and it is connected to an isometric transducer under a basal voltage of 0.5 g.

A contraction of the tissue is induced by the addition of 0.5 μM of carbachol, we wait for the contraction to stabilize (15 min), then the preparation is exposed to the compound to be studied, in increasing cumulative concentration of 0.1 nM to 1 μM. Compounds that induce relaxation are characterized as 5-HT ₄ agonists. For compounds which do not induce relaxation, the preparation is exposed to serotonin in increasing cumulative concentrations, from 0.1 nM up to a concentration inducing maximum relaxation, and the relaxation curve due to serotonin, in the presence of the compound to be studied, is then compared to a control curve established in the absence of said compound. If its presence induces a shift of the curve to the right, the compound studied is characterized as a 5-HT ₄ antagonist.

The pK _b of the compounds according to the invention are between 5 and 10.

They can therefore be used for the preparation of medicaments intended to act as antagonists of the serotonergic receptors 5-HT ₃ and 5-HT.

Thus, the compounds of the invention of formula (I) are particularly useful for the treatment and / or prevention of functional intestinal disorders such as intestinal motor and secretory disorders, disorders of intestinal secretion, irritable colon, viscerosensitivity, intestinal pain, diarrhea, but also esophageal reflux; cystic fibrosis of the pancreas, carcinoid syndrome; megacolon urinary or intestinal incontinence.

On the other hand, the compounds of the invention, due to their affinity for 5-HT ₃ receptors can also be used for the treatment and / or prevention of nausea and vomiting, for example consecutive to treatment antitumor or administration of an anesthetic; esophageal spasm; central nervous system disorders such as schizophrenia, mania, anxiety and depression; cognitive impairment such as senile dementia such as Alzheimer's or dementia related to age, memory and attention deficits, cerebrovascular impairments, Parkinson's disease; psychoses; dyskinesia, pain, migraines and headaches; alcohol or drug dependence or withdrawal disorders; gastrointestinal function disorders such as dyspepsia, peptic ulcer, heartburn, flatulence; disorders of the cardiovascular system and respiratory disorders.

The use of the compounds according to the invention for the preparation of a medicament intended to treat or prevent a pathology where the inhibition of a 5-HT ₃ or 5-HT ₄ antagonist brings a therapeutic benefit.

The use of the compounds according to the invention for the preparation of a medicament intended to treat the pathologies mentioned above forms an integral part of the invention.

According to another of its aspects, the present invention relates to pharmaceutical compositions containing, as active principle, a compound according to the invention.

Thus, these pharmaceutical compositions contain an effective dose of a compound according to the invention or of a pharmaceutically acceptable salt, N-oxide or hydrate thereof, and one or more suitable pharmaceutical excipients.

Said excipients are chosen according to the pharmaceutical form and the desired mode of administration.

In the pharmaceutical compositions of the present invention for oral, sublingual, subcutaneous, intramuscular, intravenous, topical, intratracheal, intranasal, transdermal or rectal administration, the active principle of formula (I) above, its salt or hydrate, if appropriate, can be administered in unit administration form, in mixture with conventional pharmaceutical excipients, to animals and humans for the prophylaxis or treatment of disorders or above diseases. Suitable unit dosage forms include oral forms such as tablets, capsules, powders, granules and oral solutions or suspensions, sublingual, buccal, intratracheal, intranasal forms of administration. subcutaneous, intramuscular or intravenous administration and forms of rectal or vaginal administration. For topical application, the compounds according to the invention can be used in creams, ointments or lotions.

In order to obtain the desired prophylactic or therapeutic effect, the dose of active principle can vary between 0.1 mg and 50 mg per kg of body weight per day. Although these dosages are examples of an average situation, there may be special cases where higher or lower dosages are appropriate, such dosages also belong to the invention. According to usual practice, the appropriate dosage for each patient is determined by the doctor according to the method of administration, the weight and the response of said patient.

Each unit dose may contain from 0.1 to 1000 mg, preferably from 0.1 to 500 mg, of active ingredient in combination with one or more pharmaceutical excipients. This unit dose can be administered 1 to 5 times a day so as to administer a daily dosage of 0.5 to 5000 mg, preferably 0.5 to 2500 mg.

For example, when preparing a solid composition in the form of tablets, the main active ingredient is mixed with a pharmaceutical excipient, such as gelatin, starch, lactose, magnesium stearate, talc, gum arabic or the like. The tablets can be coated with sucrose, a cellulose derivative, or other materials. The tablets can be produced by different techniques, direct compression, dry granulation, wet granulation or hot melting. According to a second example, a preparation in capsules is obtained by mixing the active ingredient with a diluent and by pouring the mixture obtained into soft or hard capsules.

For parenteral administration, aqueous suspensions, isotonic saline solutions or sterile injectable solutions are used which contain pharmacologically compatible dispersing agents and / or wetting agents, for example propylene glycol or butylene glycol.

The present invention according to another of its aspects, also relates to a method of treatment of the pathologies indicated above which comprises the administration of a compound according to the invention or one of its salts, N-oxides or hydrates.

Claims

claims

1. Compound of formula (I)

in which :

Ri represents a hydrogen atom, a halogen, a Cι_ ₄ alkoxyl, methyl, hydroxy or amino group,

R ₂ represents - (CH ₂ ) ₂ -OH or -CH ₂ -C (0) 0H when R ₃ represents a hydrogen atom, or R ₂ represents a hydrogen atom when R ₃ represents- (CH ₂ ) ₂ -0H or -CH ₂ -C (0) 0H, or R ₂ and R ₃ are linked to form one of the cycles shown below:

X represents an oxygen atom, a group -NH- or -N (Cι_ alkyl) -, and

A is a group of formula B or C

B C in which:

Y represents a nitrogen atom or a methylene group,

R ₄ represents a hydrogen atom, a Cι_ ₄ alkyl group, or a benzyl, n is equal to 0 or 1, and m is equal to 0 or 1, and optionally in the form of N-oxides and / or salts of addition to pharmaceutically acceptable acids of one of these forms.

2. Compound of formula (I) according to claim 1 characterized in that A represents a group of formula C.

3. Compound of formula (I) according to claim 2 characterized in that X represents an oxygen atom, m = 0, n = 0.

4. Compound of formula (I) according to claim 3 characterized in that R4 represents a methyl.

5. Composed of fo

in which :

X, m, n and Ri, R ₄ are as defined in claim 1 and, R ₅ and R ₆ , identical or different, represent a hydrogen atom, a Cι_ ₄ alkyl group, a phenyl or a benzyl and optionally in the form of an enantiomer, a diastereoisomer, including a racemic mixture as well as the addition salts with pharmaceutically acceptable acids of one of these forms.

6. Compound of formula (I) characterized in that it consists of:

- 8-hydroxy-7,8-dihydropyrazolo [1,5,4-de] [1,4] benzoxazine-2-carboxylate of endo-8-methyl-8-azabicyclo [3.2.1] oct-3-yl; - 7-hydroxy-1- (2-hydroxyethyl) indazole-3-carboxylate of endo-8-methyl-8-azabicyclo [3.2.1] oct-3-yl;

- 7- (2-hydroxy-1-ethyloxy) indazole-3-carboxylate of endo-8-methyl-8-azabicyclo [3.2.1] oct-3-yl;

- endo-8-methyl-8-azabicyclo [3.2.1] oct-3-yl 7-carboxymethoxyindazole-3-carboxylate; or

- 7, 8-dihydropyrazolo [1,5, 4-de] [1, 4] benzoxazine-2-carboxylate of endo-8-methyl-N-oxide-8-azabicyclo [3.2.1] oct-3-yl.

7. Process for the preparation of a compound according to one of claims 1 to 6 by biotransformation of a compound of formula (III)

in which A, X, Ri, R ₅ and Rβ are as defined in claim 1 or 5.

8. Pharmaceutical composition characterized in that it contains a compound of formula (I) according to one of claims 1 to 6 in combination with one or more pharmaceutically acceptable excipients.

9. Use of a compound of formula (I) according to any one of claims 1 to 6 for the preparation of a medicament intended to treat or prevent a pathology where the inhibition of a 5-HT ₃ or 5 antagonist -HT provides a therapeutic benefit.

10. Use according to claim 9 characterized in that the pathology consists of intestinal functional disorders, disorders of intestinal secretion, irritable bowel, viscerosensitivity; intestinal pain, diarrhea, esophageal reflux, cystic fibrosis of the pancreas, carcinoid syndrome, urinary or intestinal megacolon incontinence.