ITMI951593A1 - NON-FLUORINATED QUINOLONE DERIVATIVES AND THEIR USE AS ANTIBACTERIAL AGENTS - Google Patents

Abstract

Non fluorinated quinolone derivatives having general formula (I) (FORMULA I) in which the meaning R1, and R2 and R3 will be specified in the text, having a high antibacterial activity particularly against gram positive microorganisms and fluoroquinolone resistant microorganisms.

Description

Description of the patent application for industrial invention entitled: Non-fluorinated quinoionic derivatives and their use as antibacterial agents

FRONT TECHNIQUE

Quinoions are synthetic antibacterial agents that have been known for about 30 years. Among the older quinoionic derivatives can be mentioned, for example, nalidixic acid, pipemidic acid and cinoxacin, which are used exclusively for the treatment of urinary infections.

A significant development in the field of antibacterial chemotherapy was recorded with the discovery of fluoroquinolones. The introduction of a fluorine atom in position 6 of the kinoionic structure allowed to obtain compounds with a broader spectrum of action, a higher potency and a better bioavailability.

Fluoroquinolones, particularly ciprofloxacin, are also widely used for the treatment of infections by gram-positive microorganisms, however they have the drawback that their use has induced the emergence of resistant microorganism strains {Antimicrob. Agents Chemother. 32 377 "383 (1993) and Antimicrob. Agents Chemother. 38. 163-169 (1994)).

Furthermore, fluoroquinolones exert important toxic effects particularly on the central nervous system (Antimicrob. Agents Chemother. 32 · 1764-1770 (1993)) ·

It is therefore desirable to have new antibacterial substances also effective in the treatment of infections sustained by strains resistant to fluoroquinolones and having reduced toxic effects.

Recent studies (J. MED. CHEM. 38. 973-982 (1995)) have shown that a good antibacterial activity can also be obtained by replacing the fluorine atom in position 6 of the fluoroquinolones with an amino group.

SUMMARY

We have now surprisingly found a new class of non-fluorinated quinoionic derivatives having high antibacterial activity which allow to overcome the drawbacks of the known art.

Said quinoionic derivatives have the general formula (I)

wherein the meaning of R1, R2 and R3 will be specified in the detailed description.

The quinoionic derivatives according to the present invention show a surprisingly high activity particularly towards gram-positive microorganisms and towards methicillin-resistant and fluoroquinolone-resistant strains.

Furthermore they show reduced toxic effects.

DETAILED DESCRIPTION OF THE INVENTION

The characteristics and advantages of the quinoionic derivatives according to the present invention and the process for their preparation will be described in detail in the following description.

Said quinoionic derivatives have general formula (I)

where R4 and R5 are selected from H, OH. alkyl from C1 to C4, aminoalkyl from to Cty and hydroxyalkyl from a

The invention also relates to the salts of the derivatives (I) with pharmacologically acceptable acids.

The preparation of the quinoionic derivatives having general formula (I) in which R1 = and R2 = H is carried out by the process represented by the following scheme.

This process allows to prepare the compounds of the present invention with high yield and is suitable for industrial production.

Step (a) is carried out by transforming 2-chloro-4-fluoro-3-methyl benzoic acid into the corresponding chloride by heating it in thionyl chloride. The crude chloride, dissolved in toluene, is reacted with ethyl 3_ dimethylaminoacrylate and Et3N at 90 ° C to obtain the enaminoketoester (1).

In step (b) the compound (1) dissolved in a mixture of ethanol and ethyl ether is treated with cyclopropylamine at room temperature. The intermediate obtained, dissolved in DMF, by cyclization with K2CO3 at a temperature of 100 ° C is transformed into quinolone (2).

In step (c) the compound (2) is reacted with HBF4 in aqueous solution at a temperature between 90 and 100 ° C, to prepare the difluoroborane (3).

In step (d) the compound (3) is reacted with an amine having the formula R3H in which R3 has the meaning defined above. The molar ratio between the amine R3H and the compound (3) is between 3: 1 and 10: 1. The reaction is carried out in DMSO at a temperature between 75 and 85 ° C and the reaction mixture, after cooling to room temperature, is poured into a mixture of water and ice to free the carboxylic acid from the borane.

The compound (I) is thus obtained in which R1 = and R2 = H.

For illustrative purposes, the following examples of preparation of quinoionic derivatives according to the present invention are reported.

EXAMPLE 1

a) Preparation of the compound (1)

fi <)>

A mixture consisting of 2-chloro-4-fluoro-3-methyl benzoic acid (26 g, 0.13 mmol) and thionyl chloride (20 g) is heated under reflux for 3 hours. The excess thionyl chloride is removed by distillation under vacuum, and the residue thus obtained is solubilized in anhydrous toluene. To this solution is added ethyl 3 ~ dimethylaminoacrylate (18.6 g, 0.13 mmoles) obtained as described in J. Med. Chem. 18.44, (1975) and anhydrous Et3N (28 g. 0.27mmol). The mixture is heated at 90 ° C for 2 hours. After cooling, the formed triethylamine hydrochloride is filtered off, the filtrate is treated with water to eliminate the residual triethylamine hydrochloride, the solution is dehydrated with Na2SO4. and the solvent is evaporated. The residue treated with cyclohexane yields 30.3 g of ethyl 2- {2-chloro-4-fluoro-3-methylbenzoyl) -3 "(dimethylamino) acrylate (1). Yield 70%, m.p. 110-ll4 <e> C. ;;;; b) Preparation of compound (2) ;;;;;; (2); The solution of compound (1) (20 g, 63.9 mmol) in absolute ethanol (200 ml) and ethyl ether (150 ml), is treated drop by drop with cyclopropylamine (3.65 ml, 64 mmoles) and kept under stirring for 15 minutes at room temperature. The solvent is evaporated and the residue thus obtained is dissolved in anhydrous DMF (50 ml) and added with K2C03 (8.82 g, 63.9 mmoles). The mixture is heated to 100 ° C for 3 hours and after cooling it is poured in a mixture of H20 and ice. The precipitate obtained is filtered, washed with H 2 0 and dried to give 15 g of ethyl 1-cyclopropyl-7-fluor-8-methyl-4-oxo-1,4-dihydro-3-quinolynolecarboxylate (2). Yield 81%, m.p. 164-166 * 0.

c) Preparation of the compound (3)

(3)

The mixture consisting of compound (2) (8 g, 27 mmoles) and HBF4 at 50% in water (100 ml), is heated to 90-100 ° C for 8 hours. After cooling, the mixture is poured into H20 and the precipitate formed is collected by filtration, washed several times with H20 and dried. 7 g of (1-cyclopropyl-7-fluoro-8-methyl-4-oxo-1,4-dihydro-quinolin-3-yl) carbonyl difluoroborane (3) are obtained as a white solid. Yield 84%,

m.p. 276-278 ° C.

d) Preparation of the compound (4)

The mixture consisting of compound (3) (0.4 g, 1.3 mmol) and 4-methylpiperazine (Ο.65 g, 6.5 mmol) in anhydrous DMSO (4 ml), is heated for 20 hours at 80 " C. After cooling, the mixture is poured into a mixture of H20 and ice and the precipitate formed is collected by filtration, washed several times with H20 and dried. The solid is then solubilized in CHCl3 and the mixture is treated several times with saturated H20. with NaCl; the organic phase is dehydrated on Na2SO4 and evaporated. The residue obtained treated with EtOAc gives a solid which is filtered and dried to give 0.177 g of 1-cyclopropyl-8-methyl-7- (4-raethyl-1- piperazinyl) -1,4-dihydro-4-oxo-3-quinolynecarboxylic (4). Yield 40%,

M.P. 273-274 C.

EXAMPLE 2

Operating with the methods described in example 1 starting from intermediate (3) and using 4-hydroxypiperidine as amine, 1-cyclopropyl-8-methyl-7- (4-hydroxy-1-piperidinyl) acid was obtained -1, 4-dihydro-4-oxo-3-quinolincarboxylic (5) with a yield of 60%.

m.p. 226-229 ° C.

EXAMPLE 3

Operating with the modalities described in Example 1 starting from the intermediate (3) and using piperidine as amine, 1-cyclopropyl-8-methyl-7- (1-piperidinyl) -1,4-dihydro acid was obtained -4-oxo-3_quinolincarboxylic (6) with a yield of 65? M.p. 208-210 ° C.

EXAMPLE 4

Operating with the modalities described in Example 1 starting from the intermediate (3) and using 4-methylpiperidine as amine, 1-cyclopropyl-8-methyl-7- (4-methyl-1-piperidinyl) acid was obtained -1,4-dihydro-4-oxo-3-quinolincarboxylic (7) with a yield of 60%.

m.p. 249-250 ° C.

EXAMPLE 5

Operating with the modalities described in Example 1 starting from the intermediate (3) and using 2,6dimethylmorpholine as amine, 1-cyclopropyl-8-methyl-7- (2,6-dimethyl-4-) acid was obtained morpholinyl) -1,4-dihydro-4-oxo-3-quinolincarboxylic (8) with a yield of 50%.

m.p. 269-272 ° 0.

EXAMPLE 6

Preparation of the mixture (9)

(9)

The mixture consists of the compound (3) (0.6 g, 1.9 mmol) and 1, 2, 3, 4-tetrahydroisoquinoline (1.8 g, 13 mmol) in anhydrous DMSO (5 ml) and anhydrous Et3N (1.3 g, 13 mmol) is heated to 70 ° C for 30 hours. After cooling, the mixture is poured into H2O, alkalized with 10% NaOH and treated with CHCl3. The organic extracts are treated with diluted HC1, dehydrated on Na2SO4 and evaporated. The residue is purified on a silica gel column eluting with CHCl3. The fractions containing the product are combined and evaporated; the residue is further purified by treatment with Et20. 0.210 g of 1-cyclopropyl-8-methyl-7- (1.2,3.4, tetrahydroisoquinolin-2-yl) -1, 4-dihydro-4-oxo-3 <_> quinolecarboxylic acid (9) are obtained after filtration light yellow solid. Yield 30%,

mp 215-217 ° 0.

EXAMPLE 7

Operating with the modalities described in Example 1 starting from intermediate (3) and using thiomorpholine as amine, 1-cyclopropyl-8-methyl-7- (4-thiomorpholinyl) -1, 4-dihydro acid was obtained -4-oxo-3-quinolincarboxylic (10) with a yield of 65%.

m.p. 250-252 ° C.

EXAMPLE 8

a) Preparation of the compound (11)

Operating with the modalities described in Example 1 starting from intermediate (3) and using 3-acetamido pyrrolidine as amine, 1-cyclopropyl-8-methyl-7- {~ 3_ acetamido-1-pyrrolidinyl acid was obtained } -1,4-dihydro-4-oxo-3-quinolincarboxylic (11) with a yield of 60%.

m.p. 260-262'C.

b) Preparation of the compound (12)

The mixture consisting of compound (11) (0.100 g, 0.27 mmol), 20% NaOH (2 ml) and EtOH (1 ml), is heated under reflux for 14 hours. After cooling, the solution is acidified with AcOH up to pH 6.5 and the precipitate thus obtained is filtered, washed with H20 and then with Et20 and finally dried to give 0.08 g of 1-cyclopropyl-8-methyl-7- acid. {3-amino-1-pyrrolidinyl) -1, 4-dihydro-4-oxo-3-quinolynecarboxylic (12). Yield 88%,

M.p. 275-277 "C.

EXAMPLE 9

Preparation of the mixture (13)

Operating in the manner described in Example 8 a) and b), starting from the intermediate (3) and using the 3 ~ (S) -acetamido pyrrolidine as amine and by final treatment with acetic acid up to pH 4, the 1-cyclopropyl-8-methyl-7- (3- (S) -amino-1-pyrrolidinyl) -1, 4-dihydro-4-oxo-3-quinolincarboxylic acid (13) acetate with a yield of 35% -p.f. 212-213 ° C.

PHARMACOLOGICAL TESTS

The antibacterial activity of the quinoionic derivatives according to the invention was tested against gram-positive and gram-negative microorganisms against microorganisms resistant to methicillin (S. aureus Methi-r) and to fluoroquinolones (S. aureus Cipro-r and E. coli Cyprus-r). The minimal inhibitory concentration (MIC) was determined using the broth microdilution method. The comparison was performed with ciprofloxacin (CYPRUS) which is a drug with a fluoroquinolone structure and is widely used in therapy thanks to its high antibacterial activity.

The microorganisms used in the experiment were stored in a 15-20% solution of glycerol in water at -80 ° C and the concentration of the frozen cultures was approximately 1-2 x 10 ° CFU / ml.

The Muller-Hinton Broth (MHB) medium was used to determine the MIC. In the case of fastidious microorganisms, the broth was integrated as suggested by the guidelines of the National Committee for Clinical Laboratory Standards (NCCLS Document M7-A3, vol. 13. n. 25).

For the growth of S. pyogenes and S. pneumoniae 5% horse blood was added to the broth and bovine haematin, NAD and yeast extract were added for H. influenzae.

On the day of the test, a vial of bacterial culture was thawed and diluted 1: 100 with MHB, integrating as required in the case of fastidious microorganisms.

The reference compounds and those to be tested were dissolved in dimethyl sulfoxide (DMSO) at a concentration of 1.6 mg / ml and the solution obtained was diluted to a concentration of 32 pg / ml with MHB.

Test microplates were filled with 0.05 mL MHB per well.

0.05 ml of a 32 Mg / ml solution in MHB of the test compound was added to the first well of each row and serially diluted 11 times. 0.05 of the inoculum suspension was then dispensed into each well and the plates were incubated at 35 ° C for 24 hours.

For each row the first well without growth was recorded and the corresponding drug concentration calculated. MIC is defined as the minimum concentration of the antibacterial agent that visually inhibits the growth of the microorganism.

The results obtained are shown in table 1.

The results reported in Table 1 show that all tested substances exert a surprisingly high antibacterial activity against gram-positive and resistant microorganisms. In particular, compounds 5, 6, 7 and 10 are at least 10 times more active than ciprofloxacin towards the methicillin-resistant strains and 4-6 times towards the fluoroquinolon-resistant ones. This activity indicates a use of the compounds of the invention particularly for respiratory tract infections and for infections caused by resistant microorganisms.

Another important characteristic of the quinoionic derivatives according to the present invention consists in a marked decrease of the toxic effects with respect to the 6-fluoroquinolones. Thanks to their characteristics, the quinoionic derivatives of the present invention can be successfully used in the preparation of pharmaceutical compositions comprising an effective amount of at least one of said derivatives in admixture with pharmacologically acceptable excipients and diluents, for the treatment of bacterial infections.

For the formulation in pharmaceutical compositions the compounds of general formula (I) can be salified with pharmacologically acceptable acids selected from the group comprising p-toluenesulfonic acid, citric acid, tartaric acid, maleic acid, salicylic acid, fumaric acid, succinic acid, oxalic acid , etc.

Said compositions can be administered orally or parenterally at a dose ranging from 100 to 800 mg / day and between 25 and 250 mg / day, respectively.

Claims (6)

CLAIMS 1. Non-fluorinated quinoionic derivatives suitable for use as antibacterials having the following general formula and salts of derivatives (I) with pharmacologically acceptable acids, wherein: where R4 and R5 are selected from H. OH, alkyl from a, aminoalkyl from a and hydroxyalkyl from C1 to C4. 2. Process for the preparation of quinoionic derivatives according to claim 1, characterized in that, when R1 = and R2 = H. it is carried out by means of the following reaction steps: 3. Process according to claim 2, characterized in that step (d) is carried out by reacting compound (3) with an R3H amine in DMSO at a temperature between 75 and 85 ° C and with a molar ratio between amine R3H and compound (3) between 3: 1 and 10: 1. 4. Pharmaceutical compositions comprising as active substance - an effective amount of at least one of the compounds according to claim 1. in mixture with excipients and pharmacologically acceptable, suitable for the treatment of bacterial infections. 5. Compositions according to claim 4, in form suitable for oral administration. 6. Compositions according to claim 4, in form suitable for parenteral administration.