JP2003507467A - Fatty acid synthase inhibitors - Google Patents

Abstract

  (57) [Summary] The present invention relates to the use of compounds as inhibitors of fatty acid synthase FabH.

Description

Detailed Description of the Invention

[0001] (Technical field)   The present invention relates to the use of compounds as inhibitors of fatty acid synthase FabH.

The biosynthetic pathways for saturated fatty acids are very similar in prokaryotes and eukaryotes.
However, although the chemical reaction remains the same, the mechanism of the biosynthetic organs is completely different. Vertebrate and yeast have type I fatty acid synthase (FAS), all of which enzymatic activity is encoded on one or two polypeptide chains, respectively. The acyl carrier protein (ACP) is an integral part of the complex. In contrast, in most bacteria and plants, each FAS (type II) in the reaction is catalyzed by a unique monofunctional enzyme, and ACP is a distinct protein. Mycobacteria are special in that they have both type I and type II FAS; the former are involved in basic fatty acid biosynthesis, whereas the latter are of complex cell envelope lipids such as mycolic acid. Involved in synthesis. Thus, there seems to be considerable potential for selective inhibition of bacterial systems using broad spectrum antibiotics (Jackowski, S. (1992) In Emerging Targets in
Antibacterial and Antifungal Chemotherapy. J. Sutcliffe & N. Georgopapad
akou, Chapman & Hall, New York; Jackowski, S. et al. (1989) J. Biol. Chem.
264, 7624-7629).

The first step in the biosynthetic cycle is malonyl-ACP and acetyl-C by FabH.
It is the condensation of oA. In a subsequent round, malonyl-ACP is condensed with growing chain acyl-ACP (FabB and FabF, synthases I and I, respectively).
I). The second step in the extension cycle is NADPH-dependent β-ketoacyl-AC
Ketoester reduction by P-reductase (FabG). Subsequent dehydration with β-hydroxyacyl-ACP dehydrase (either FabA or FabZ) results in trans-2-enoyl-ACP, which is then converted to acyl-ACP by NADH-dependent enoyl-ACP reductase (FabI). It In a further round of this cycle, which adds 2 carbon atoms per cycle, essentially palmitoyl-ACP is obtained, followed by palmitoyl-ACP.
Feedback inhibition of FabH and I by H. et al. Greatly stops the cycle (Heath et al. (1996) J. Biol. Chem. 271, 1833-1836). Therefore, FabH is a major biosynthetic enzyme that is also an important regulatory point throughout the synthetic pathway (Heath, RJ and Rock, CO (1996) J. Biol. Chem. 27.
1, 1833-1835; Heath, RJ and Rock, CO (1996) J. Biol. Ch.
em. 271, 10996-11000).

Thiolactomycin antibiotics have been shown to have broad spectrum antibiotic activity both in vivo and in vitro and specifically inhibit all three condensing enzymes. The antibiotic is non-toxic and does not inhibit mammalian FAS (Hayashi,
T. et al. (1984) J. Antibiotics 37, 1456-1461; Miyakawa, S. et al.
1982) J. Antibiotics 35, 411-419; Nawata, Y et al. (1989) Act.
a Cryst. C45, 978-979; Noto, T. et al. (1982) J. Antibiotics 3
5, 401-410; Oishi, H. et al. (1982) J. Antibiotics 35, 391-.
396). Similarly, cerulenin is a potent inhibitor of FabB & F and a bactericidal agent, but it competes with eukaryotes for the fatty-acyl binding site common to both types of FAS. It is toxic (D'Agnolo, G. et al. (1973) Biochim. Biophys. Acta. 326, 155-166). Extensive research using these inhibitors has revealed that these enzymes are essential for survival. Little research has been done on Gram-negative bacteria.

The need to develop a series of new antibiotic compounds that do not lend themselves to existing resistance mechanisms is still unmet. None of the commercially available antibiotics target fatty acid biosynthesis, and thus novel antibiotics of this type will not be rendered inactive by known antibiotic resistance mechanisms. Moreover, this class of antibiotics may be broad-spectrum targets. Therefore, FabH inhibitors would help meet this unmet need.

DISCLOSURE OF THE INVENTION The present invention provides cinnamic acid derivatives and pharmaceutical compositions containing these compounds and their Fas useful as antibiotics for treating infections of Gram-negative and positive bacteria.
Including use as a bH inhibitor. Further, the invention provides a method of treating Gram-negative or Gram-positive bacterial infections in animals, including humans, which method comprises administering to an animal in need of such treatment an effective amount of a compound of the invention. Constitute.

[0007] (Best Mode for Carrying Out the Invention)   The compounds of the present invention have the formula (I):

[Chemical 2]

[0008] [In the formula;   R₁Is C_1-10Alkyl, C_1-3Arylalkyl, C_1-3Hetero ant
Alkylalkyl, heteroaryl, C_1-3Alkyl-C_3-6Cycloa
Rukiru and C_3-6Selected from the group consisting of cycloalkyl;   R_TwoIs O (CH_Two)_mAryl, O (CH_Two)_mHeteroaryl, N (R_Three) (CH
_Two)_mAryl, N (R_Three) (CH_Two)_mHeteroaryl, N (R_Four) CO aryl,
N (R_Four) CO heteroaryl, N (R_Four) SO_TwoAryl and N (R_Four) SO_TwoF
Teloaryl SO_TwoSelected from the group consisting of heteroaryl, wherein R₁and
R_TwoThe aryl and heteroaryl groups of are one or more CH_Three, CF_Three,
OCF_Three, OH, OCH_Three, NH_Two, NHCH_Three, N (CH_Three)_Two, SCH_Three, S
OCH_Three, SO_TwoCH_Three, Halogen, CO_TwoH, CO_TwoCH_Three, CONH_Two, C
ON (CH_Three)_Two, NHCOH, NHCOCH_Three, NHSO_TwoCH_Three, Methylenedi
Optionally substituted with oxy;   R_ThreeIs H, C_1-8Alkyl, C_1-3Alkyl-aryl, CO (C_1-8)
Selected from the group consisting of alkyl and COaryl;   R_FourIs H, C_1-8Alkyl good C_1-3A group of alkyl-aryl
Selected; and   m means an integer of 1-3] Or a pharmaceutically acceptable salt thereof.

Also included in the invention is a complex of pharmaceutically acceptable salts. As used herein, “C _1-10 alkyl” or “alkyl” means a straight or branched chain having 1 to 10 carbon atoms, unless the chain length is specifically limited.
Examples include, but are not limited to, methyl, ethyl, n-propyl, iso-propyl, n-butyl, sec-butyl, iso-butyl, tert-butyl, n-pentyl and the like. The alkyl may have a substituent such as hydroxy, carboxy and alkoxy. The term "cycloalkyl" as used herein refers to a cyclic ring, preferably, but not limited to, cyclopropyl, cyclopentyl, cyclohexyl, etc., preferably a cyclic ring having 3 to 8 carbon atoms. Used to do. The term "arylalkyl" or "heteroarylalkyl" or "heterocyclic alkyl" as used herein, unless otherwise stated, the above-mentioned C _1-10 bound aryl as above, heteroaryl or heterocyclic group Used to mean alkyl.

As used herein, “aryl” means phenyl and naphthyl and aryl substituted with hydroxy, carboxy, halo, alkoxy, methylenedioxy and the like. As used herein, "heteroaryl" means 1 such as pyrrole, pyrazole, furan, thiophene, quinoline, isoquinoline, quinazolinyl, pyridine, pyrimidine, oxazole, thiazole, thiadiazole, triazole, imidazole, benzotriazole or benzimidazole. Or a 5-10 membered aromatic ring system in which one or more rings contain one or more heteroatoms selected from the group consisting of N, O or S, but is not limited thereto. Absent. Preferred aryl substituents as used herein are halo, including chloro, fluoro, bromo and iodo, in any combination; C _1-10 alkyl, C _1-10 alkoxy, aryloxy or heteroaryl. Includes oxy. The compounds of the present invention may contain one or more asymmetric carbon atoms and may exist in racemic and optically active forms. All of these compounds and diastereomers are included within the scope of the invention.

Some of the compounds of the present invention can be crystallized or recrystallized from solvents such as organic solvents. In such cases, solvates may be formed. The present invention includes within its scope stoichiometric solvates including hydrates as well as compounds containing variable amounts of water, which can be prepared by methods such as lyophilization procedures. Because the antibiotic compounds of the present invention are intended for use in pharmaceutical compositions, the compounds may each be in a substantially pure form, eg, at least 60% pure, more suitably at least 75% pure. It will be readily understood that it is supplied with a purity of at least 85%, and preferably at least 85%, especially at least 95%, especially at least 98% (% is weight / weight based). Impure preparations of the compounds can also be used to prepare more pure forms for use in pharmaceutical compositions; less pure preparations of these compounds are at least 1%, more suitably at least 5%. , Preferably 10 to 49% of a compound of formula (I) or a salt thereof.

Preferred compounds of the invention are: DL-2- (6-chloropiperonyl) -3- [4- (2,6-dichlorobenzyloxy) phenyl] propionic acid; DL-2- (6-chloropiperonyl) -3 -[4- (3,4-dichloroanilino) phenyl] propionic acid; and DL-3- [4- (3,4-dichlorobenzyloxy) phenyl] -2-phenylpropionic acid.

Compounds of formula (I) wherein R ₂ is benzyloxy are prepared by the method described in Scheme 1.

[Chemical 3] a) NaH, DMF; b) acetic acid, piperidine, benzene; c) NaBH ₄ , EtO
H; d) 6-chloropiperonyl chloride, NaH, DMF; e) KOH, EtOH

4-hydroxybenzaldehyde (1-Scheme-1) and benzyl halide (such as 2,6-dichlorobenzyl bromide) are reacted with a base (such as sodium hydride) in a solvent (such as DMF) and stirred. (6 to 30 hours, preferably 16 hours
Time) to obtain 3-Scheme-1. Benzyloxybenzaldehyde (3-Scheme-4) [4- (2,6-dichlorobenzyloxy) benzaldehyde, etc.]
-Scheme-1 and a catalyst (such as piperidine and acetic acid) in a solvent (such as benzene)
While removing water by azeotropic mixing at reflux temperature with Knoevena gel (Knoevena
gel) condensation to give 5-Scheme-1. 5-Scheme-1 is reduced with a hydride reducing agent (such as sodium borohydride) in a solvent (such as ethanol) to give 6-Scheme-1. This is a base (sodium hydride etc.) in a solvent (DMF etc.)
Alkylation with benzyl halide such as 6-chloropiperonyl chloride to give 7-Scheme-1. 7-Scheme-1 was saponified with a base (potassium hydroxide, etc.) in a solvent (ethanol, etc.) and simultaneously decarboxylated,
Scheme-1 is obtained.

Compounds of formula (I) wherein R ₂ is anilino are prepared by the method described in Scheme 2.

[Chemical 4] a) NaH, DMF; b) KOH, EtOH; c) acetic acid, piperidine, benzene;
d) Cu (Oac) ₂ , TEA, 4A sieve, CH ₂ Cl ₂ ; e) SmI ₂ , CH ₃
OH; f) KOH, EtOH

Diethyl malonate (2-Scheme-2) was treated with a base (such as sodium hydride) in a solvent (such as DMF) and a benzyl halide (6-chloropiperonyl chloride (
1-Scheme-2) etc.) and stirred (1 hour to 10 hours, preferably 3 hours). The resulting diester was treated with a base (
Mono-saponification with (for example, potassium hydroxide) and stirring (4 to 30 hours, preferably 18 hours) gives 3-Scheme-2. 3-Scheme-2 to 4-Scheme-
2 (prepared by reacting arylboronic acid with ethylene glycol and simultaneously removing water) is azeotropically mixed at a reflux temperature with a catalyst (such as piperidine and acetic acid) in a solvent (such as benzene) to remove water. While undergoing Knoevenagel condensation, the boronic acid (5-Scheme-2) is obtained. This compound is used in a solvent (such as methylene chloride) with a cupric salt (such as copper (II) acetate) and a base (such as TEA) together with a molecular sieve to give a substituted phenol or aniline (such as 3,4-dichloroaniline). ) To give 6-Scheme-2. 6-Scheme-2 is reacted with a reducing agent (such as samarium iodide) to give the hydrocinnamic acid ester (7-Scheme-2).
Saponification of 7-Scheme-2 with a base (such as potassium hydroxide) in a solvent (such as ethanol) gives 8-Scheme-2.

Compounds of formula (I) wherein R ₁ is aryl or heteroaryl are prepared by the method described in Scheme 3.

[Chemical 5] a) LDA; b) _Ac2 O, DBU; c) KOH; d) SmI ₂

The optionally substituted benzaldehyde (1-Scheme-3) is condensed with arylacetic acid (2-Scheme-3) using a base such as LDA, followed by acetic anhydride and DB.
Acylation with a base such as U, followed by saponification with a base such as KOH, followed by an elimination reaction, gives the substituted cinnamic acid (3-Scheme-3). Reduction with a reducing agent such as samarium iodide gives 4-Scheme-3.

Synthetic Examples The present invention will be described using the following examples. These examples are exemplary only,
It does not limit the scope of the present invention in any way. Unless otherwise noted, all temperatures are degrees Celsius and solvents are the purest available.

Example 1 Preparation of DL-2- (6-chloropiperonyl) -3- [4- (2,6-dichlorobenzyloxy) phenyl] -propanoic acid a) 4- (2,6-dichlorobenzyloxy) Benzaldehyde A solution of 4-hydroxybenzaldehyde (3.59 g, 24.39 mmol) and 2,6-dichlorobenzyl bromide (7.05 g, 29.39 mmol) in dimethylformamide (20 mL) at 0 ° C., 60 % Sodium hydride (1.176
g, 29.39 mmol) was added. After stirring for 16 hours at ambient temperature, the reaction was partitioned between ethyl acetate and water. The organic layer is washed with aqueous sodium chloride and dried (
MgSO ₄₎ was. Purify by flash column chromatography (silica gel, hexane / ethyl acetate) to give the title compound as an off-white solid (7.38 g, 89
%).

B) Diethyl 4- (2,6-dichlorobenzyloxy) benzylidene] malonate 4- (2,6-Dichlorobenzyloxy) benzaldehyde (5.62 g, 20 mmol) and diethyl malonate (3.52 g) , 22 mmol) of benzene (75
To a solution in (mL) was added piperidine (90 μL) and glacial acetic acid (0.25 mL). After refluxing for 24 hours while azeotropically mixing to remove water, the cooled reaction was concentrated under reduced pressure and partitioned between ether and water. The organic extracts 1N hydrochloric acid, saturated sodium bicarbonate solution, water, and washed successively with brine, dried (MgSO _4). The volatiles were removed under reduced pressure to give the crude title compound (8.35 g, 97%), which was used in the next step without further purification.

C) Diethyl [4- (2,6-dichlorobenzyloxy) benzyl] malonate [4- (2,6-Dichlorobenzyloxy) benzylidene] diethyl malonate (8.3
5 g, 19.2 mmol) under argon at 0 ° C. in a stirred solution of ethanol,
Sodium borohydride was added in small portions. After the addition is complete, p.
Adjusted to H6. The mixture was filtered, concentrated under reduced pressure and then partitioned between ether and water. The organic extracts 1N hydrochloric acid, saturated sodium bicarbonate solution, water, and washed successively with brine, dried (MgSO _4). Remove volatiles under reduced pressure,
The title compound (7.47 g, 88%) was obtained.

D) diethyl 2- [4- (2,6-dichlorobenzyloxy) benzyl] -2- (6-chloropiperonyl) malonate [4- (2,6-dichlorobenzyloxy) benzyl] diethyl malonate ( 4.38g
(10.3 mmol) in an aqueous solution in DMF stirred at 0 ° C. under argon to give 9
5% Sodium hydride (260 mg, 10.3 mmol) was added. 20 at 0 ° C
After stirring for 1 minute, 6-chloropiperonyl chloride (2.1 g, 10.3 mmol) D was added.
A solution in MF (5 mL) was added. The solution was warmed to room temperature and stirred for a further 3 hours. The reaction mixture was partitioned between ethyl acetate and 3N HCl and the organic extract was washed with water then brine and dried (Na ₂ SO ₄ ). The solvent was removed under reduced pressure,
Subsequent crystallization from ethanol gave the title compound (5.22 g, 85%).

E) DL-2- (6-chloropiperonyl) -3- [4- (2,6-dichlorobenzyloxy)
Phenyl] propanoic acid diethyl 2- [4- (2,6-dichlorobenzyloxy) benzyl] -2- (6-chloropiperonyl) malonate (1.0 g, 1.68 mmol) and potassium hydroxide (650 mg) in ethanol. Solution (10.1 mmol) was refluxed under argon for 24 hours. The solvent was removed under reduced pressure and the residue was dissolved in water. The aqueous solution is filtered and then 10%
Acidified with HCl. The precipitate was collected by filtration, washed with water and dried under vacuum at 65 ° C. Purify using chromatography (silica gel) to give the title compound (
0.83 g, 100%) was obtained. Mass spectrum: ES +; [M + H] ⁺ .

Example 2 DL-2- (6-chloropiperonyl) -3- [4- (3,4-dichloroanilino) phenyl]-
Preparation of propionic acid a) Diethyl 2- (6-chloropiperonyl) malonate Diethyl malonate (9.44 g, 59 mmol) stirred at 0 ° C. under argon.
To an aqueous solution of in DMF (25 mL) was added 60% sodium hydride as a slurry in DMF (10 mL) (2.4 g, 60 mmol, oil removed by pentane washing). After stirring at 0 ° C for 20 minutes, 6-chloropiperonyl chloride (11.
A solution of 0 g, 53.7 mmol) in DMF (10 mL) was added. The solution was warmed to room temperature and stirred for another 13 hours. The reaction mixture was partitioned between ethyl acetate and 3N hydrochloric acid and the organic extract was washed with water then brine and dried (Na ₂ S).
O ₄ ). The solvent was removed under reduced pressure, followed by crystallization from ether / hexane to give the title compound (13.14 g, 74.5%).

B) 2- (6-Chloropiperonyl) malonic acid monoethyl ester 2- (6-chloropiperonyl) malonic acid diethyl (10.7 g, 32.6 mmol)
Was dissolved in ethanol (110 mL) and stirred with potassium hydroxide (1.84 g, 32.55 mmol) at room temperature for 18 hours. The reaction mixture was diluted with water (100 mL) and then concentrated under reduced pressure to half the original volume. The resulting aqueous solution was washed with ether then acidified with 3N HCl and the product extracted into ethyl acetate.
The organic extract was washed with water, then brine and dried (Na ₂ SO ₄ ). The solvent was removed under reduced pressure to give the title compound (9.84 g, 92%).

C) (E) -2 ′-(6-Chloropiperonyl) -4-dihydroxyboryl ethyl cinnamate 4- (1,3,2-dioxaborolan-2-yl) benzaldehyde (azeotropic removal of water While refluxing 4-dihydroxyborylbenzaldehyde and ethylene glycol in benzene) (3.04 g, 17.3 mmol) and 2
To a solution of-(6-chloropiperonyl) malonic acid monoethyl ester (6.5 g, 21.7 mmol) in benzene (75 mL) was added piperidine (86 μL) and glacial acetic acid (
247 μL) was added. After refluxing for 23 hours while azeotropically mixing and removing water,
The cooled reaction was diluted with ethyl acetate. The solution was washed successively with 3N hydrochloric acid, water, brine and dried (Na ₂ SO ₄ ). Crystallization from ethyl acetate / hexane gave the title compound as a white solid (5.8g, 86%).

D) Ethyl (E) -2 ′-(6-chloropiperonyl) -4-dihydroxyborylcinnamate (E) -2 ′-(6-chloropiperonyl) -4- (3,4-dichloroanilino) cinnamate Ethyl acid (20
0 mg, 0.51 mmol), 3,4-dichloroaniline (100 mg, 0.61 mmol), copper (II) acetate (122 mg, 0.61 mmol), triethylamine (430 μL, 3.1 mmol) and 4A powder sieve. A slurry of (1.2 g) in methylene chloride (6 mL) was stirred at room temperature for 2 h with open air.
The mixture was filtered and all volatiles were removed under reduced pressure. Purification by chromatography (silica gel, hexane / ethyl acetate) gave the title compound as a white solid (142 mg, 46%).

E) DL-2- (6-chloropiperonyl) -3- [4- (3,4-dichloroanilino) phenyl] propionate ethyl (E) -2 ′-(6-chloropiperonyl) -4- ( To a solution of ethyl 3,4-dichloroanilino) cinnamate (30 mg, 0.06 mmol) in methanol (0.5 mL) was added samarium iodide (about 0.4 mmol in 14 mL THF). This was stirred for 10 minutes then partitioned between water and ethyl acetate. The organic extract was washed successively with 3N hydrochloric acid, water, brine, dried (Na ₂ SO ₄ ) and all volatiles removed under reduced pressure. The product obtained was used in the next step without further purification.

F) DL-2- (6-chloropiperonyl) -3- [4- (3,4-dichloroanilino) phenyl] propionic acid DL-2- (6-chloropiperonyl) -3- [4- (3 , 4-Dichloroanilino) phenyl]
Ethyl propionate (crude product above) and potassium hydroxide (35 mg, 0
A solution of (0.6 mmol) in ethanol (2 mL) was stirred at reflux temperature for 1 hour. The solution was diluted with water and the aqueous solution was washed with ether. The aqueous solution was acidified with 3N hydrochloric acid and the product was extracted into ethyl acetate. The organic extract was washed with water, brine and dried (Na ₂ SO ₄ ). Remove all volatiles under reduced pressure, followed by preparative HPL
Purification by C afforded the title compound (15.4 mg, 54% for 2 steps). Mass spectrum: ES +; 478.0 [M + H] ⁺ .

Example 3 DL-3- [4- (3,4-Dichlorobenzyloxy) phenyl] -2-phenylpropionic acid a) 4- (3,4-dichlorobenzyloxy) -2-phenyldiisopropyl cinnamate Amine (232 mg, 2.3 mmol) TH at -78 ° C.
To a solution in F (3 mL) under argon, with stirring, n-butyllithium (1.5
3 mL, 1.5 M in hexane, 2.3 mmol) was added. It was warmed to room temperature over 15 minutes and then cooled to -78 ° C. Methyl phenylacetate (30
0 mg, 2 mmol) was added and the solution was again warmed to room temperature over 15 minutes and then cooled to -78 ° C. A solution of (3,4-dichlorobenzyloxy) benzaldehyde (562 mg, 2 mmol) in THF (3 mL) was added and the reaction mixture was allowed to warm to room temperature over 1 hour. Acetic anhydride (0.5 mL) and DBU (760 mg, 5 mmol) were added to the resulting solution with stirring for 1 hour. The reaction mixture was partitioned between ethyl acetate and water. The organic extract was washed with brine, dried (sodium sulfate) and the volatiles removed under reduced pressure. The residue was refluxed in ethanol with excess KOH for 4 hours, then cooled and diluted with water. The aqueous solution was washed with ether, then acidified with 3N hydrochloric acid and extracted with ethyl acetate. The organic extract was washed with water, brine, then dried (sodium sulfate) and the volatiles removed under reduced pressure. Preparative HPLC gave the title compound as a white solid (
220 mg, 27.5%). Mass spectrum: ES +; 399 [M + H] ⁺ .

B) DL-3- [4- (3,4-dichlorobenzyloxy) phenyl] -2-phenylpropionic acid 4- (3,4-dichlorobenzyloxy) -2-phenylcinnamic acid (100 mg, 0 .2
To a solution of (5 mmol) in methanol (1 mL) was added samarium iodide (about 1 mmol in 10 mL THF). This was stirred for 10 minutes then partitioned between water and ethyl acetate. The organic extract was washed with water, brine and dried (Na ₂ SO ₄ ).
And all volatiles were removed under reduced pressure. The resulting residue was purified by preparative HPLC to give the title compound as a white solid (32mg, 32%). Mass spectrum: ES +; 401 [M + H] ⁺ .

[0033]

Biological assay: his-tagged S. aureus FabD and Sigma
FabH was assayed in a binding format using an acyl carrier protein (ACP) purchased from Lyophilized ACP was treated with β in phosphate buffer.
-Reduced with mercaptoethanol. Malonyl-CoA and FabD were added to the reduced ACP, thus producing malonyl-ACP. After the FabD reaction reached equilibrium, [ ¹⁴ C] acetyl-CoA and the inhibitor were added to the FabD.
The reaction was started by the addition of H 2. [ ¹⁴ C] using TCA precipitation and filtration
The acetyl-CoA substrate was separated from the [ ¹⁴ C] acetoacetyl-ACP product.

The reproducibility, sensitivity, throughput and analytical power of the second and third screens suitable for developing the first screen hits have been characterized, confirmed and are currently in use. A compound, a purified mammalian fatty acid biosynthetic enzyme, E. coli (E.
coli) FabH, FabB and human lung cell cytotoxicity assays.

In addition, whole-cell antibacterial activity was reduced to a series of clinically relevant wild-type and efflux (
efflux impaired) bacteria using standard and novel fluorescence-based techniques. The FabH assay has been completely characterized kinetically and its reaction mechanism has been proposed. Detailed studies yielded new data on the mechanism of inhibition by instrumental compounds, including thiolactomycin. The screen of use is directly related to the therapeutic goal-eradication of bacteria from the site of infection (cure). At present, several types of bacterially infected animal experiments are available, which are meaningful and are currently used in the present invention and in numerous other studies in SB. Extensive prior experiments with known antibacterial agents have confirmed that bacterial death in vitro and in animal studies is a good indicator of bacterial death and cure of infections in vivo.

The present invention also provides a pharmaceutical composition comprising a compound of formula (I) or a pharmaceutically acceptable salt or in vivo hydrolysable ester thereof and a pharmaceutically acceptable carrier. . The compositions of the invention, including those in a form adapted for oral, topical or parenteral use, may be used to treat bacterial infections in mammals, including humans. The antibiotic compounds of the present invention, as well as other antibiotics, may be formulated for administration in any convenient way for use in human or veterinary medicine. The composition may be formulated for administration by any route, such as oral, topical or parenteral, especially oral. The composition may be in the form of tablets, capsules, powders, granules, lozenges, creams or liquid preparations such as oral or sterile parenteral solutions or suspensions.

The topical formulations of the present invention may be provided as, for example, ointments, creams or lotions, eye ointments and eye drops or ear drops, impregnated pharmaceutical surgical materials and aerosols, preservatives, drug penetration It may contain solvents to assist and suitable conventional additives such as emollients in ointments and creams. The formulations may also contain compatible conventional carriers, such as cream or ointment bases and ethanol or oleyl alcohol for lotions. Such carriers may be present as from about 1% to about 98% of the formulation. More commonly, they form up to about 80% of the formulation.

Tablets and capsules for oral administration may be in unit dosage form, with binders such as syrup, gum arabic, gelatin, sorbitol, gum tragacanth or polyvinylpyrrolidone; fillers such as lactose, sugar, Corn starch, calcium phosphate, sorbitol or glycine; tableting lubricants such as magnesium stearate, talc, polyethylene glycol or silica; disintegrants such as potato starch; or acceptable wetting agents,
For example, it may contain a conventional excipient such as sodium lauryl sulfate. The tablets may be coated according to methods well known in normal pharmaceutical practice. Oral liquid formulations include, for example, aqueous or oily suspensions, solutions, emulsions,
It may be in the form of a syrup or elixir, or it may be presented as a dry product for constitution with water or other suitable vehicle for reconstitution before use. Such liquid formulations include suspending agents such as sorbitol, methylcellulose, glucose syrup, gelatin, hydroxyethylcellulose, carboxymethylcellulose, aluminum stearate gel or hardened edible fats, emulsifiers such as lecithin, sorbitan monooleate, or gum arabic. Non-aqueous vehicles (which may include edible oils) such as almond oil, oily esters such as glycerin, propylene glycol or ethyl alcohol; preservatives such as p-
It may contain methyl or propyl hydroxybenzoate or sorbic acid and, if desired, conventional additives such as flavoring agents or colorants.

Suppositories will contain conventional suppository bases, eg cocoa-butter or other glyceride. For parenteral administration, fluid unit dosage forms are prepared using the compound and a sterile vehicle, water being preferred. The compound, depending on the vehicle and concentration used, can either be suspended or dissolved in the vehicle. In preparing solutions, the compound is dissolved in water for injection and filter sterilized before filling into a suitable vial or ampoule and sealing. The solution preferably contains a buffer (eg, phosphate) to maintain the pH in the range of about 3.5-7. In addition, DMSO or an alcoholic solvent (0.01-1) may be added to aid solubility and permeability of the compound of formula (I).
May be present) (at a concentration such as 0 mL / liter). Advantageously, agents such as local anesthetics, preservatives and buffering agents can be dissolved in the vehicle.
To improve stability, the composition can be frozen after filling into vials and the water removed under vacuum. The dried lyophilized powder is then sealed in a vial,
An accompanying vial of water for injection may be supplied to reconstitute the liquid prior to use. Parenteral suspensions are prepared in substantially the same manner, except that the compound is suspended in the vehicle instead of being dissolved and sterilization cannot be achieved by filtration. The compound can be sterilized by exposure to ethylene oxide before suspension in a sterile vehicle. Advantageously, a surfactant or wetting agent is included in the composition to facilitate uniform distribution of the compound.

The composition may contain from 0.1% by weight, preferably from 10-60% by weight, of the active material, depending on the method of administration. When the composition comprises dosage units, each unit will preferably contain 50-500 mg of active ingredient. The dose used for the treatment of adults depends on the route and frequency of administration, but is preferably 1 to 140.
It will be in the mg / kg body weight range. β-ketoacyl-ACP synthase (FabH
The inhibitors of) can be administered intravenously, intramuscularly, intraperitoneally by injection in solution, or orally. The solution preferably contains a buffer (eg, phosphate) to maintain the pH in the range of about 3.5-7. Also, DMSO or alcoholic solvents may be present (at concentrations such as 0.01-10 mL / liter) to aid in the solubility and permeability of the β-ketoacyl-ACP synthase (FabH) inhibitors.

No unacceptable toxicological effects are expected when a compound of formula (Ia) or a pharmaceutically acceptable salt or in vivo hydrolysable ester thereof is administered in the above dosage range. The compound of formula (I) may be the only therapeutic agent in the composition of the present invention, or in combination with other antibiotics or compounds which enhance the antibacterial activity of the compound of formula (I). Good.

The antibiotic compounds of the present invention include Gram-negative microorganisms such as Escherichia coli and Klebsiella pneumoniae, including isolates resistant to existing antibiotics and Klebsiella pneumoniae. Staphylococcus aureus, Streptococcus pneumoniae, Enterococcus faecalis
rococcus faecalis) and Enterococcus faecalis
It is active against a wide range of microorganisms including Gram-positive microorganisms such as ecium).

All publications cited herein, including but not limited to patents and patent applications, are each individual publication individually and individually, as if fully set forth. Are incorporated herein by reference, as if they were indicated to be incorporated herein by citation. The above description fully discloses the invention including preferred embodiments thereof. Modifications and improvements of the embodiments specifically disclosed herein are within the scope of the following claims. Without further elaboration, one of ordinary skill in the art, using the above description, believes that the present invention can be utilized to its full extent. Therefore, the examples herein should be construed as merely illustrative, and not limiting the scope of the invention in any way. Specific embodiments of the invention in which an exclusive property or privilege is claimed are set forth above.

─────────────────────────────────────────────────── ─── Continued front page    (81) Designated countries EP (AT, BE, CH, CY, DE, DK, ES, FI, FR, GB, GR, IE, I T, LU, MC, NL, PT, SE), OA (BF, BJ , CF, CG, CI, CM, GA, GN, GW, ML, MR, NE, SN, TD, TG), AP (GH, GM, K E, LS, MW, MZ, SD, SL, SZ, TZ, UG , ZW), EA (AM, AZ, BY, KG, KZ, MD, RU, TJ, TM), AE, AL, AU, BA, BB, BG, BR, CA, CN, CZ, DZ, EE, GE, G H, GM, HR, HU, ID, IL, IN, IS, JP , KP, KR, LC, LK, LR, LT, LV, MA, MG, MK, MN, MX, NO, NZ, PL, RO, S G, SI, SK, SL, TR, TT, TZ, UA, US , UZ, VN, YU, ZA (72) Inventor Siegfried Be Christensen             The Force             United States 19103 Hu, Pennsylvania             Philadelphia, Number 4-M, Che             Lee Street 2301, Rivers Et             The (72) Inventor Robert A. Danes             United States 19446 La Pennsylvania             N'sdale, Cold Spring Low             No. 2587 (72) Inventor Jack De Laver             United States 18901 Do Pennsylvania             Illestown, Pine Run Road 403             Turn (72) Inventor Joseph Waynestock             United States 19087 U, Pennsylvania             Wayne, Painter's Rain No. 10 F-term (reference) 4C086 AA01 AA02 AA03 BA12 MA01                       MA04 NA14 ZB35 ZC20

Claims (4)

[Claims] 1. Formula (I): [Chemical 1] [In the formula;   R₁Is C_1-10Alkyl, C_1-3Arylalkyl, C_1-3Hetero ant
Alkylalkyl, heteroaryl, C_1-3Alkyl-C_3-6Cycloa
Rukiru and C_3-6Selected from the group consisting of cycloalkyl;   R_TwoIs O (CH_Two)_mAryl, O (CH_Two)_mHeteroaryl, N (R_Three) (CH
_Two)_mAryl, N (R_Three) (CH_Two)_mHeteroaryl, N (R_Four) CO aryl,
N (R_Four) CO heteroaryl, N (R_Four) SO_TwoAryl and N (R_Four) SO_TwoF
Selected from the group consisting of teloaryl;   R_ThreeIs H, C_1-8Alkyl, C_1-3Alkyl-aryl, CO (C_1-8)
Selected from the group consisting of alkyl and COaryl;   R_FourIs H, C_1-8Alkyl and C_1-3A group of alkyl-aryl
Selected; and   m is an integer from 1 to 3] Or a pharmaceutically acceptable salt thereof or a pharmaceutically acceptable salt thereof
Salt complex. 2. DL-2- (6-chloropiperonyl) -3- [4- (2,6-dichlorobenzyloxy) phenyl] propanoic acid; and DL-2- (6-chloropiperonyl) -3- [4-- The compound of claim 1 selected from the group consisting of (3,4-dichloroanilino) phenyl] propionic acid. 3. A method of treating a bacterial infection comprising administering to a patient in need of treatment of the bacterial infection an effective amount of a compound of formula (I) according to claim 1. 4. A compound of formula (I) is DL-2- (6-chloropiperonyl) -3- [4- (2,6-dichlorobenzyloxy) phenyl] propanoic acid; and DL-2- (6- The treatment method according to claim 3, which is selected from the group consisting of chloropiperonyl) -3- [4- (3,4-dichloroanilino) phenyl] propionic acid.