TW201602065A - Hydroxamic acid derivative - Google Patents

Abstract

The present invention addresses the problem of providing a novel compound which is useful as a pharmaceutical product, and which has high antibacterial activity with regard to gram-negative bacteria such as pseudomonas aeruginosa and with regard to the drug-resistant bacteria of gram-negative bacteria. The present invention provides a 2-[{4-[4-(1-aminocyclopropyl)buta-1,3-diyne-1-yl]benzoyl}(methyl)amino]-N-hydroxyl-N',2-dimethyl propane diamide represented by formula [1], or a pharmaceutically-acceptable salt.

Description

Hydroxamic acid derivative

The present invention relates to a novel hydroxamic acid derivative having antibacterial activity against Gram-negative bacteria and resistant bacteria thereof. Further, the present invention relates to a pharmaceutical composition or an antibacterial agent containing a hydroxamic acid derivative.

In the Gram-negative bacteria, there is a membrane comprising a lipid bilayer which is not present in Gram-positive bacteria. Therefore, in terms of the problem of drug permeability, Gram-negative bacteria have a tendency to be more resistant than Gram-positive bacteria. Further, it is known that Gram-negative bacteria have a plurality of drug excretion proteins. Non-Patent Document 1 discloses that drug excretion proteins are also associated with drug resistance. Further, lipopolysaccharide (LPS), which is one of the main constituent components of the outer membrane, is associated with toxicity as an endotoxin.

It is known that among Gram-negative bacteria, especially Pseudomonas aeruginosa, there is a strong tendency to exhibit natural resistance to various antibacterial agents. Pseudomonas aeruginosa is widely present in the natural environment or living environment, but is an attenuated bacterium that does not usually show pathogenicity to healthy people. However, patients with severe underlying diseases, patients with so-called immunosuppressive hosts who use immunosuppressants for transplantation, etc., and patients with medical procedures such as medical catheters or endotracheal intubation, and surgery are used. In other words, Pseudomonas aeruginosa is a pathogen causing severe acute infections such as sepsis. Therefore, Pseudomonas aeruginosa is one of the important causative bacteria of opportunistic infections or nosocomial infections.

In recent years, third-generation cephems, carbapenems, or amines that have been expected to be effective against Pseudomonas aeruginosa have been clinically isolated on the medical site. Pseudomonas aeruginosa which is resistant to a glucoside-based drug or the like (Non-Patent Document 2). In addition, the multidrug-resistant Pseudomonas aeruginosa which has been resistant to the above three types of drugs has been isolated (Non-Patent Document 3).

Since there is almost no useful drug when infecting multi-drug resistant Pseudomonas aeruginosa, multidrug-resistant Pseudomonas aeruginosa is a major problem in the world as a cause of refractory infectious diseases. Therefore, it is highly desirable to develop an agent having a novel mechanism of action.

UDP-3-O-mercapto-N-acetylglucosamine deacetylase (LpxC) is an enzyme that synthesizes a hydrophobic anchor, LPS, which is a constituent of the outer membrane. Lipid A biosynthesis involves a 10-stage reaction. LpxC catalyzes the second stage of the biosynthesis reaction, and cleaves the acetyl group of UDP-3-O-mercapto-N-acetylglucosamine (Non-Patent Document 4). Lipid A is a component necessary for the formation of an outer membrane, and as a result, it is necessary for the survival of Gram-negative bacteria (Non-Patent Document 5).

That is, LpxC is one of the important enzymes that limit the rate during lipid A biosynthesis, and is an enzyme necessary for lipid A biosynthesis. Therefore, since the agent which inhibits the activity of LpxC particularly has a mechanism different from that of the prior agent for Gram-negative bacteria including Pseudomonas aeruginosa, it is strongly expected to be an effective antibacterial agent against the drug-resistant Pseudomonas aeruginosa.

As an LpxC inhibitor, an inhibitor having a guanamine structure is disclosed in Patent Documents 1 to 11 and Non-Patent Documents 6 to 13.

Among these, as a compound having a propylene glycol skeleton, Patent Documents 5 and 9 disclose a compound having a propylene glycol skeleton and a diacetylene structure.

Patent Document 5 specifically discloses a compound 507 having a cyclopropyl ring at a terminal portion of a diacetylenic group.

Further, in Patent Document 9, specifically, a compound 233 is disclosed as a compound having a cyclopropyl ring. Patent Document 9 discloses a compound 197, a compound 209, and a compound 221, and a compound 8 discloses a synthesis method and an antibacterial activity.

[Previous Technical Literature] [Patent Literature]

[Patent Document 1] International Publication No. 2004/062601

[Patent Document 2] International Publication No. 2007/069020

[Patent Document 3] International Publication No. 2008/154642

[Patent Document 4] International Publication No. 2010/031750

[Patent Document 5] International Publication No. 2011/132712

[Patent Document 6] International Publication No. 2012/154204

[Patent Document 7] International Publication No. 2013/039947

[Patent Document 8] International Publication No. 2013/170030

[Patent Document 9] International Publication No. 2013/170165

[Patent Document 10] International Publication No. 2014/142298

[Patent Document 11] International Publication No. 2014/165075

[Non-patent literature]

[Non-Patent Document 1] Antimicrobial Resistance (2002) Mar 1, 34, p. 634-640.

[Non-Patent Document 2] J. Antimicrob. Chemother. (2003) Jan 14, 51, p. 347-352.

[Non-Patent Document 3] Jpn. J. Antibiotics (2006), 59(5), p. 355-363.

[Non-Patent Document 4] J. Biol. Chem. (1995) Dec 22, 270, p. 30384-30391.

[Non-Patent Document 5] J. Bacteriol. (1987), 169, p. 5408-5415

[Non-Patent Document 6] J. Med. Chem. (2002), 45, p3112-3129.

[Non-Patent Document 7] Proc. Natl. Acad. Sci. USA (2007), 104, p18433-18438.

[Non-Patent Document 8] Chem. Biol. (2011), 18, p38-47.

[Non-Patent Document 9] Bioorg. Med. Chem. (2011), 19, p852-860.

[Non-Patent Document 10] Bioorg. Med. Chem. Lett. (2011), 21, p1155- 1161.

[Non-Patent Document 11] Current Med. Chem. (2012), 19, p2038-2050.

[Non-Patent Document 12] Bioorg. Med. Chem. Lett. (2013), 23, p2362-2367.

[Non-Patent Document 13] J. Med. Chem. (2013), 56, p6954-6966.

However, in the compound 507 disclosed in Patent Document 5, the carbon atom adjacent to the hydroxamic acid of the skeleton portion does not have a methyl group. Further, Patent Document 5 does not disclose a compound having an unsubstituted amino group as a substituent in a terminal portion of a diacetylenic group.

Further, the compounds disclosed in Patent Document 9 are all compounds having a hydroxymethyl group as a substituent on a cyclopropyl ring, and have a chiral terminal structure. Further, Patent Document 9 does not disclose a compound having an amine group as a substituent at a terminal portion of a diacetylenic group.

An object of the present invention is to provide a novel compound which is useful as a pharmaceutical product for having a strong antibacterial activity against Gram-negative bacteria represented by Pseudomonas aeruginosa and its resistant bacteria.

As a result of intensive studies, the present inventors have found that 2-[{4-[4- represented by the following formula [1]) having a 1-aminocyclopropyl group as an achiral substituent in the terminal portion of the diacetylenic group. (1-Aminocyclopropyl)butan-1,3-diyn-1-yl]benzhydryl}(methyl)amino]-N-hydroxy-N',2-dimethylpropanedifluoride The amine can solve the above problems, thereby completing the present invention.

[Chemical 1]

The invention is as follows.

(1)

A compound or a pharmaceutically acceptable salt thereof, which is represented by the formula [1], 2-[{4-[4-(1-aminocyclopropyl)butan-1,3-diyn-1-yl) Benzyl hydrazino}(methyl)amino]-N-hydroxy-N',2-dimethylpropanediamine,

(2)

A compound or a pharmaceutically acceptable salt thereof, which is represented by the formula [2]: (2S)-2-[{4-[4-(1-aminocyclopropyl)butane-1,3-diyne -1-yl]benzhydryl}(methyl)amino]-N-hydroxy-N',2-dimethylpropanediamine, [Chemical 3]

(3)

A pharmaceutical composition comprising the compound of the above (1) or (2) or a pharmaceutically acceptable salt thereof.

(4)

An antibacterial agent comprising the compound of the above (1) or (2) or a pharmaceutically acceptable salt thereof.

The novel hydroxamic acid derivative of the present invention has strong antibacterial activity against Gram-negative bacteria represented by Pseudomonas aeruginosa and resistant bacteria thereof.

In the present invention, the term "pharmaceutically acceptable salt" means a salt used in chemotherapy and prevention of bacterial infection.

Examples of the "pharmaceutically acceptable salt" include acetic acid, propionic acid, butyric acid, formic acid, trifluoroacetic acid, maleic acid, tartaric acid, citric acid, stearic acid, succinic acid, and ethyl amber. Acid, malonic acid, lactobionic acid, gluconic acid, glucoheptonic acid, benzoic acid, methanesulfonic acid, ethanesulfonic acid, 2-hydroxyethanesulfonic acid, benzenesulfonic acid, tosic acid, lauryl Sulfuric acid, malic acid, aspartic acid, glutamic acid, adipic acid, cysteine, N-acetylcysteine, hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid, hydroiodic acid, nicotinic acid, oxalic acid, Salts of acid such as picric acid, thiocyanic acid, undecanoic acid, acrylic acid polymer and carboxy vinyl polymer, salts with organic amines such as morpholine and piperidine, and salts with amino acids Wait.

As a "pharmaceutically acceptable salt", it may be a salt with an inorganic base. Examples of the salt with the inorganic base include a lithium salt, a sodium salt, a potassium salt, a magnesium salt, and a calcium salt.

The compound of the present invention (hereinafter referred to as "the compound of the present invention" is used in the sense of the case where the compound represented by the formula [1], the compound represented by the formula [2] or the pharmaceutically acceptable salt thereof is contained. When used, a hydrate can be formed, and a solvate can also be formed. Where a compound of the invention forms a hydrate or solvate, such are also included within the scope of the invention.

The term "solvent" in the case where the compound of the present invention forms a "solvate" means, for example, a polar solvent (for example, an alcohol such as methanol, ethanol, 1-propanol, 2-propanol or butanol, unless otherwise specified). a solvent, ethyl acetate, etc.), an inert solvent (for example, a halogenated hydrocarbon solvent such as chloroform or dichloromethane, diethyl ether, diisopropyl ether, tetrahydrofuran, and An ether solvent such as an alkane; a guanamine solvent such as dimethylformamide or dimethylacetamide; an aprotic solvent such as dimethyl hydrazine or acetonitrile; an aromatic hydrocarbon such as toluene; and hexane and cyclohexane A hydrocarbon such as an alkane or the like, 2-butanone or acetone.

The "solvent" may also be a mixed solvent of the solvent exemplified herein.

In the present invention, the term "antibacterial agent" means a substance having an ability to inhibit growth or kill a bacterium such as a Gram-positive bacterium or a Gram-negative bacterium. It can also be reduced in number, such as inhibiting bacterial growth or killing a portion of the bacteria.

Examples of the Gram-positive bacteria include Staphylococcus (S. aureus and Staphylococcus epidermidis), Streptococcus (Streptococcus pyogenes, Streptococcus B and Streptococcus pneumoniae), Enterococcus (feces) Enterococcus, Enterococcus faecium, etc.).

Examples of the Gram-negative bacteria include Pseudomonas (Pseudomonas aeruginosa, etc.), Escherichia coli (Escherichia coli, etc.), and Klebsiella (K. pneumoniae and Klebsiella oxysporum). (Klebsiella oxytoca), etc., Haemophilus (influenza virus and parainfluenza virus, etc.), typhus (pertussis and bronchial septicemia, etc.), Serratia (viscous shale) Serratia marcescens, etc., Proteus (Proteus vulgaris and Proteus mirabilis, etc.), Enterobacter (Enterobacter aerogenes) and Enterobacter cloacae ), etc.), Aspergillus (C. jejuni, etc.), Citrobacter f. (Citrobacter freundii, etc.), Providencia (Providencia) (Providencia) Stuartii), etc., Stenotrophomonas (Stenotrophomonas maltophilia, etc.), Vibrio (Vibrio cholerae and cholera, etc.), Morganella (Morgan's) Bacteria (Morganella morganii), etc., Salmonella (S. typhimurium and Salmonella paratyphimurium), Shigella (erythrobacteria, etc.), Acinetobacter baumannii (Acinetobacter baumannii) and acetic acid Acinetobacter baumannii (Aci Netobacter calcoaceticus), Legionella (Legionella pneumophilia, etc.), Bacteroides (Bacteroides fragilis, etc.), Neisseria (Glutococcus and meningitis), Mo Lactobacillus (Moraxella catarrhalis, etc.), Chlamydia (Chlamydia trachomatis and Chlamydia psittaci), and Helicobacter (Helicobacter pylori ( Helicobacter pylori), etc.). The compound of the present invention can be preferably used as an antibacterial agent against Gram-negative bacteria.

The following formula [1] of the present invention [Chemical 4]

2-[{4-[4-(1-Aminocyclopropyl)butan-1,3-diyn-1-yl]benzylidenyl}(methyl)amino]-N-hydroxyl -N',2-dimethylpropaneamine may be present as an optical isomer, and the compound represented by the formula [1] includes a mixture of the optical isomers and optical isomers. The pharmaceutical composition or antibacterial agent may contain specific optical isomers, and may also contain mixtures of optical isomers, especially racemates.

The present invention includes the compound represented by the formula [1] and a pharmaceutically acceptable salt thereof, and also includes a polymorph of the compound represented by the formula [1] and a pharmaceutically acceptable salt thereof.

The preferred optical isomer of the present invention is the following formula [2]

(2S)-2-[{4-[4-(1-Aminocyclopropyl)butan-1,3-diyn-1-yl]benzimidyl}(methyl)amino group] -N-Hydroxy-N',2-dimethylpropanediamine.

The present invention includes the compound represented by the formula [2] and a pharmaceutically acceptable salt thereof, and also includes a polymorph of the compound represented by the formula [2] and a pharmaceutically acceptable salt thereof.

The compound of the present invention can be combined with one or two or more pharmaceutically acceptable carriers, excipients or diluents to form a pharmaceutical composition.

Examples of the carrier, excipient, and diluent include water, lactose, dextrose, fructose, sucrose, sorbitol, mannitol, polyethylene glycol, propylene glycol, starch, rubber, gelatin, and alginic acid. Salt, calcium citrate, calcium phosphate, cellulose, liquid syrup, hydroxypropyl cellulose, hydroxypropyl methylcellulose, methyl cellulose, polyvinylpyrrolidone, alkyl p-hydroxybenzosorbide , talc, magnesium stearate, stearic acid, glycerin and various oils (sesame oil, olive oil and soybean oil, etc.).

As the pharmaceutical composition, an additive such as a bulking agent, a binding agent, a disintegrating agent, a pH adjusting agent, a solubilizing agent, and a flavoring agent which are generally used may be mixed with the above-mentioned carrier, excipient or diluent.

The pharmaceutical composition can be prepared into tablets, pills, capsules, granules, powders, liquids, emulsions, suspensions, ointments, injections (including intramuscular injections and intravenous injections) by conventional preparation techniques. Oral or non-oral medicine such as intravenous drip and skin patch.

With respect to the compound of the present invention, 30 to 3000 mg, preferably 100 to 1500 mg, may be administered to an adult patient once or several times a day, orally or orally. Preferably, the administration form is intravenous drip injection or intravenous injection, and the preferred administration form is intravenous drip injection. The dose can be appropriately increased or decreased depending on the type of the disease to be treated, the age, weight, and symptoms of the patient. Further, the compound of the present invention can also be used in combination with other agents.

[Examples]

Hereinafter, the present invention will be described in more detail by way of examples and test examples. The invention is not limited by the examples. In particular, the method for synthesizing the compound 1 in the present invention is not limited to the following methods, and the synthesis may be carried out by a method known in the art of changing the order of each step and protecting/deprotecting the functional group.

MS (mass spectrometry) was measured by LCMS-IT-TOF (Liquid Chromatography Mass Spectrometry-Ion Trap-Time Of Flight) (Shimadzu) apparatus. As the ionization method, an ESI (Electrospray Ionization) method or a double ionization method using ESI and APCI (Atmospheric Pressure Chemical Ionization) method is employed. The data is recorded as the found value. A molecular ion peak is usually observed, but in the case of a compound having a hydroxyl group (-OH), a peak of H ₂ O detachment as a fragment peak is sometimes observed. In the case of salt, a molecular ion peak or a fragment ion peak of the free body is usually observed.

High performance liquid chromatography-mass spectrometry (LCMS) was carried out under the following conditions.

Measuring machinery: Agilent 1290 from Agilent and Agilent 6130 from Agilent

Column: Waters Acquity UPLC ^® CSH ^TM C18 1.7μm 2.1×50mm

Ionization: Electron Impact Ionization (ESI)

Solvent: Liquid A is an aqueous solution containing 0.1% formic acid, and liquid B is acetonitrile containing 0.1% formic acid.

(Condition 1)

Flow rate: 0.8mL/min

Gradient: 0 minutes (A solution / B solution = 80 / 20), 1.2 minutes (A liquid / B liquid = 80 / 20), 1.4 minutes (A liquid / B liquid = 1 / 99)

(Condition 2)

Flow rate: 0.8mL/min (0 minutes to 1.2 minutes), 1.0mL/min (1.2 minutes to 1.38 minutes)

Gradient: 0 minutes (A liquid / B liquid = 95/5), 1.2 minutes (A liquid / B liquid = 50 / 50), 1.38 minutes (A liquid / B liquid = 3 / 97)

The NMR (nuclear magnetic resonance) spectrum indicates proton NMR, using tetramethyl decane as an internal reference, and the δ value is expressed in ppm.

Elemental analysis was carried out using a device of a vario MICRO cube (elementar).

OH NH type silica gel chromatography and chromatography on silica gel type carrier systems used in the manufacture of Grace Japan Co. REVELERIS ^TM packed column and the like. The phase separator (Phase Separator) is manufactured by Biotage Co., Ltd.

The abbreviations in the examples are shown below.

AcOEt: ethyl acetate

BSA: Bovine Serum Albumin (bovine serum albumin)

CuI: copper iodide

DMAPO: 4-(dimethylamino)pyridine 1-oxide

HEPES: 2-[4-(2-Hydroxyethyl)-1-piperazinyl]ethanesulfonic acid (2-[4-(2-hydroxyethyl)-1-) Ethyl sulfonate

IPE: diisopropyl ether

MeOH: methanol

MNBA: 2-methyl-6-nitrobenzoic anhydride

NBS: N-bromosuccinimide

PdCl ₂ (PPh ₃ ) ₂ : bis(triphenylphosphine)palladium(II) dichloride

Supers-table Pd: Tris[tris[3,5-bis(trifluoromethyl)phenyl]phosphine}palladium

p-TsOH‧H ₂ O: p-toluenesulfonic acid monohydrate

TEA: Triethylamine

THF: tetrahydrofuran

s: single peak

Br.s.: wide single peak (wide single peak)

d: double peak

Dd: double double peak

m: multiple peak

t: triple peak

q: Quadruple peak

Example 1

(2S)-2-[{4-[4-(1-Aminocyclopropyl)butan-1,3-diyn-1-yl]benzimidyl}(methyl)amino]-N- Hydroxy-N',2-dimethylpropanediamine (Compound 1)

(1) (2S)-2-[(4-iodobenzylidene)(methyl)amino group obtained by the method described in Patent Document 5 (International Publication No. 2011/132712) under a nitrogen atmosphere. Add a CuI at room temperature in a suspension of -N,2-dimethyl-N'-(tetrahydro-2H-pyran-2-yloxy)propanediamine (19.2 g) in THF (200 mL) (0.299 g), PdCl ₂ (PPh ₃ ) ₂ (0.551 g) and TEA (10.9 mL). Thereafter, trimethyldecyl acetylene (5.01 g) was added over 5 minutes, and the mixture was stirred at room temperature for 2 hours. The reaction mixture was filtered and concentrated using a EtOAc (EtOAc/MeOH = 95/5), and the obtained residue was subjected to OH type spur column chromatography (hexane/AcOEt=100/0→0/100). Purification, the obtained pale orange solid was washed with IPE and dried to obtain (2S)-N,2-dimethyl-2-(methyl{4-[(trimethyldecyl)ethynyl] Benzomidine}amino)-N'-(tetrahydro-2H-pyran-2-yloxy)propanediamine (17.6 g, colorless solid, 97%).

MS (ESI/APCI dual): m/z = 482 (M+Na) ⁺ , 458 (MH) ^-

¹ H NMR (600MHz, CHLOROFORM -d) δ ppm 0.26 (9H, s), 1.48-1.90 (6Hm), [1.80], 1.81 (3H, s), 2.82-2.87 (3H, m), [3.13], 3.15(3H,s), 3.50-3.69(1H,m),3.82-4.05(1H,m),4.92-5.01(1H,m),7.41-7.48(2H,m),7.48- 7.54(2H,m ), [6.99], 7.62 (1H, br.s.), [10.06], 10.47 (1H, s)

(2) To a solution of the compound (17.1 g) obtained in Example 1-(1) in MeOH (250 mL). Stir to room temperature and stir for 1 hour. The reaction mixture was added to a mixture of saturated aqueous ammonium chloride (1.0L) and chloroform (1.0L), and the organic layer was separated, and the aqueous layer was extracted with chloroform (0.5 L). The collected organic layer was dried (magnesium sulfate), filtered, and concentrated, and the obtained crude product was purified by OH-type silica gel column chromatography (hexane/AcOEt=80/20→0/100). The obtained pale orange solid was washed with IPE and dried to obtain (2S)-2-[(4-ethynylbenzylidenyl)(methyl)amino]-N,2-dimethyl- N'-(Tetrahydro-2H-pyran-2-yloxy)propanediamine (12.4 g, colorless solid, 86%).

MS (ESI/APCI dual): m/z = 410 (M+Na) ⁺ , 386 (MH) ^-

¹ H NMR (600MHz, CHLOROFORM -d) δ ppm 1.49-1.91 (6H, m), [1.80], 1.81 (3H, s), 2.83-2.87 (3H, m), 3.12-3.19 (1H, m), [3.14], 3.16 (3H, s), 3.53-3.69 (1H, m), 3.83-4.05 (1H, m), 4.93-5.01 (1H, m), 7.44-7.51 (2H, m), 7.52-7.57 (2H,m),[6.98],7.62(1H,br.s.),[10.06],10.47(1H,s)

[化8]

(3) Acetone (12.0 g) of the compound obtained in Example 1-(2) was added dropwise to a mixture of NBS (6.62 g) and a mixture 150 mL) of the mixture was stirred at the same temperature for 1.5 hours. After the reaction mixture was added to a saturated aqueous solution of sodium hydrogencarbonate (600 mL) and stirred for 0.5 hour, chloroform (600 mL) was added and the insoluble material was filtered, and the organic layer was separated. The aqueous layer was extracted with chloroform (100 mL). The collected organic layer was dried (anhydrous magnesium sulfate), filtered, and concentrated. The suspension obtained by adding water (500 mL) to the obtained residue was stirred for 1.5 hours, and then filtered and washed (water, 500 mL). The obtained pale orange solid was dissolved in chloroform (500 mL). The obtained chloroform solution was washed with water (200 mL), dried (anhydrous magnesium sulfate), filtered, and concentrated. The obtained pale orange solid was purified by OH type rubber column chromatography (chloroform / MeOH = 100 / 0 → 90/10) and then recrystallized (AcOEt / IPE) to obtain (2S) - 2 { [4-(Bromoethynyl)benzylidene](methyl)amino}-N,2-dimethyl-N'-(tetrahydro-2H-pyran-2-yloxy)propanedifluoride Amine (8.02 g, 56%).

MS (ESI/APCI dual): m/z = 488 (M+Na) ⁺ , 464 (MH) ^-

¹ H NMR (600MHz, CHLOROFORM -d) δ ppm 1.46-1.90 (6H, m), [1.80], 1.81 (3H, s), 2.82-2.87 (3H, m), [3.14], 3.16 (3H, s ), 3.52-3.68 (1H, m), 3.82-4.04 (1H, m), 4.92-5.01 (1H, m), 7.42-7.53 (4H, m), [6.97], 7.61 (1H, br.s. ), [10.05], 10.47 (1H, s)

[Chemistry 9]

(4) Acetonitrile (50 mL) of 1-ethynylcyclopropylamine hydrochloride (1.59 g) synthesized by the method described in the literature (Synthesis, 2010, Vol. 23, pp. 3967-3973) TEA (4.35g), ultra-stable palladium (547mg) and CuI (49mg) were added to the solution, and a solution of the compound obtained in Example 1-(3) in acetonitrile (10 mL) was added dropwise over 10 minutes under nitrogen atmosphere. The reaction was carried out for 1 hour at room temperature. After filtering the reaction mixture, the filtrate was concentrated under reduced pressure, and the residue obtained was purified by EtOAc (EtOAc/EtOAc/EtOAc/EtOAc/EtOAc -[{4-[4-(1-Aminocyclopropyl)butan-1,3-diyn-1-yl]benzhydryl}(methyl)amino]-N,2-dimethyl -N'-(tetrahydro-2H-pyran-2-yloxy)propanediamine (2.42 g, pale yellow foam, 60%).

LCMS retention time: 0.81 minutes (condition 2)

MS (ESI): m/z = 465 (MH) ^-

¹ H NMR (600MHz, CHLOROFORM -d) δ ppm 1.00-1.02 (2H, m), 1.06-1.08 (2H, m), 1.39-2.13 (6H, m), [1.79], 1.80 (3H, s), 2.83-2.85(3H,m),[3.13],3.16(3H,s),3.55-3.66(1H,m),3.84-4.02(1H,m),4.89-5.00(1H,m),7.41-7.55 (4H,m),[6.97],7.61(1H,br.s.),[10.07],10.48(1H,br.s.)

(5) To a MeOH solution (50 mL) of Compound ( ₂ . A saturated sodium hydrogencarbonate solution was added to the reaction mixture, and the mixture was extracted with chloroform, and then the organic layer was washed with brine. After drying the organic layer with anhydrous sodium sulfate, the desiccant was separated by filtration, and the solvent was evaporated under reduced pressure. The residue obtained was purified by OH type gel chromatography (chloroform / MeOH = 100 / 0 → 85 / 15) to obtain (2S)-2-[{4-[4-(1-amino ring) Propyl)butyl-1,3-diyn-1-yl]benzhydryl}(methyl)amino]-N-hydroxy-N',2-dimethylpropanediamine (Compound 1, 1.69) g, pale yellow solid, 86%).

LCMS retention time: 0.53 minutes (condition 2)

MS (ESI): m/z =381 (MH) ^-

¹ H NMR (600 MHz, CD ₃ OD) δ ppm 0.95-1.00 (2H, m), 1.02-1.06 (2H, m), 1.76 (3H, s), 2.78 (3H, s), 3.13 (3H, s) , 7.47-7.60 (4H, m)

Compound 1 can also be synthesized by the method shown below.

(6) After adding 122 g of cesium carbonate to a solution of diethyl 2-((t-butoxycarbonyl)amino)malonate (25.7 g) in dimethylformamide (250 mL), it was dropped in a water bath Methyl iodide (23.2 mL). After completion of the dropwise addition, the reaction system was allowed to stand in a closed system and stirred at room temperature for 5 days. Water was added to the reaction mixture, and a solution of n-hexane/AcOEt=4/1 (700 mL) was added thereto for extraction. Then, the organic layer was washed successively with a saturated aqueous solution of ammonium chloride and brine. Anhydrous magnesium sulfate and activated carbon (2 g) were added to the organic layer and stirred for 1 hour, and filtered by Celite (registered trademark). The filtrate was concentrated under reduced pressure to give 2-((t-butoxycarbonyl)(methyl)amino)-2- Diethyl methylmalonate (25.0 g, pale yellow oil, 88%).

MS (ESI/APCI dual): m/z = 326 (M+Na) ⁺

¹ H NMR (600MHz, CHLOROFORM -d) δ ppm 1.27 (6H, t, J = 7.0Hz), 1.38-1.47 (9H, m), 1.68 (3H, s), 2.87 (3H, s) 4.22 (4H, q, J=7.0Hz)

(7) A phosphate buffered aqueous solution (680 mL) was added to the compound (22.8 g) obtained in Example 1-(6), and PLE (Pig Liver Esterase, pig liver esterase) (342 mg) was added thereto at room temperature. Stir under 26 hours. The phosphate buffer aqueous solution was obtained by diluting a mixture of 0.2 M aqueous sodium dihydrogen phosphate solution (65 mL) and 0.2 M aqueous sodium hydrogen phosphate solution (435 mL) with water to 1000 mL. After adding 1 mol/L of sodium hydroxide aqueous solution (75 mL) to the reaction liquid, the pH was adjusted to 8-9, and extraction was performed using toluene (0.5 L). At this time, the mixed solution was bubbled, and thus Celite (registered trademark) filtration was again performed. Phosphoric acid (20 mL) was added to the aqueous layer obtained after the extraction, and the pH was adjusted to 2 to 3, and extraction was carried out using AcOEt (1 L). At this time, the mixed solution was also in the form of a bubble, and thus Celite (registered trademark) was filtered. After adding anhydrous magnesium sulfate to the obtained organic layer and drying it, the desiccant was separated by filtration, and the solvent was distilled off under reduced pressure. The residue was dissolved in AcOEt (0.2 L), activated carbon (1.8 g) was added and stirred for 1 hour. The activated carbon was separated by filtration and the solvent was evaporated under reduced pressure to give (R)-2-((t-butoxycarbonyl)(methyl)amino)-3-ethoxy-2-methyl-3. - oxopropionic acid (18.0 g, pale yellow syrup, 87%, ee > 99%).

MS (ESI/APCI dual): m/z = 298 (M+Na) ⁺

¹ H NMR (600 MHz, DMSO- d ₆ ) δ ppm 1.11-1.19 (3H, m), 1.32 (9H, br.s.), 1.54 (3H, s), 2.73 (3H, s), 4.02-4.13 ( 2H,m)

The chirality analysis of the compound obtained in Example 1-(7) was carried out in the following manner.

The measuring machine used was a high performance liquid chromatograph manufactured by Shimadzu Corporation. The model number of each machine is as follows.

Pump: LC-30AD, autosampler: SIL-30AC, column oven: CTO-20AC, LED array detector: SPD-M20A, deaerator: DGO-20A5R.

For the chiral column, 4.6 x 150 mm manufactured by Daicel Co., Ltd. and AD3 of 4.6 x 250 mm were connected in series. The developing solvent was n-hexane/ethanol = 98/2, and the flow rate was 1.0 mL/min.

Detection by absorption wavelength at 210 nm for racemization of 2-((t-butoxycarbonyl)(methyl)amino)-3-ethoxy-2-methyl-3-oxopropionic acid The peak hold time of the body was 26.5 minutes in the (S) body and 37.9 minutes in the (R) body.

The analysis of the compound obtained in Example 1-(7) was carried out under the above conditions, and as a result, only the (R) body was detected, and the (S) body was below the detection limit. The enantiomeric excess (ee) was >99%.

(8) To a solution of the compound (13.3 g) obtained in Example 1-(7) in toluene (121 mL), EtOAc (10.5 mL) hour. The reaction mixture was concentrated under reduced pressure to give (S)-3-chloro-2-methyl- as a crude product. Ethyl 2-(methylamino)-3-oxopropanoate (9.37 g, brown solid).

¹ H NMR (600 MHz, chloroform-d) δ ppm 1.30 (3H, t, J = 7.8 Hz), 1.72 (3H, s), 2.93 (3H, s), 4.23-4.33 (2H, m)

Simulation calculation for C ₇ H ₁₂ ClNO ₃ : C, 43.42; H, 6.25; N, 7.23;

Found: C, 45.49; H, 6.09; N, 6.75;

(9) After O-(tetrahydro-2H-pyran-2-yl)hydroxylamine (1.00 g) was azeotropically dried twice in toluene, TEA (3.57 mL) was added to a solution toluene (5.0 mL). A mixture of the compound obtained in Example 1-(8) (1.95 g) in EtOAc (EtOAc) The suspension obtained by adding AcOEt (10 mL) to the reaction mixture was filtered, and the filtrate was concentrated, and the obtained crude product was purified by OH type gel chromatography (hexane/AcOEt=50/50→0/100). And (2R)-2-methyl-2-(methylamino)-3-oxo-3-((tetrahydro-2H-pyran-2-yl)oxy)amino)propyl Ethyl acetate (1.78 g, light brown oil, 76%).

MS (ESI/APCI dual): m/z = 275 (M+H) ⁺ , 273 (MH) ^-

¹ H NMR (600 MHz, chloroform - d ) δ ppm 1.25-1.30 (3H, m), 1.43-1.90 (6H, m), [1.529] 1.532 (3H, s), 2.30 (3H, s), 3.58-3.67 (1H,m), 3.88-4.00(1H,m),4.13-4.34(2H,m),4.82-5.02(1H,m),9.68(1H,br.s.)

[化15]

(10) A 40% methylamine methanol solution (6.0 mL) was added to the compound obtained in Example 1-(9) (540 mg), and stirred for 49 hours. The reaction mixture was concentrated under reduced pressure, and the obtained residue was purified by EtOAc (EtOAc/EtOAc/EtOAc/EtOAc/EtOAc/EtOAc -N,2-dimethyl-2-(methylamino)-N'-((tetrahydro-2H-pyran-2-yl)oxy)propanediamine (433 mg, pale yellow oil) , 85%).

LCMS retention time: 0.28 minutes (condition 2)

MS (ESI): m/z = 260 (M+H) ⁺

¹ H NMR (600MHz, CHLOROFORM -d) δ ppm 1.48-1.94 (6H, m), [1.529] 1.532 (3H, s), 2.29 (3H, d, J = 4.5Hz), 2.84 (3H, dd, J =5.0, 1.2 Hz), 3.62-3.70 (1H, m), 3.93-4.06 (1H, m), 4.93-5.03 (1H, m), 7.60-7.91 (1H, m), 10.72 (1H, br.s .)

(11) To a solution of 1-ethynylcyclopropylamine hydrochloride (10.0 g) in MeOH (100 mL) Ethyl trifluoroacetate (11.2 mL) was added to the reaction mixture at room temperature and stirred overnight. The reaction solution was concentrated under reduced pressure, and water was added. It was extracted twice with AcOEt. After the organic layer obtained was combined and washed with a saturated aqueous solution of ammonium chloride, anhydrous magnesium sulfate was added to the obtained organic layer to dryness, and then the desiccant was separated by filtration, and the solvent was evaporated under reduced pressure. N-(1-ethynylcyclopropyl)-2,2,2-trifluoroacetamide (14.9 g, pale yellow solid, 99%) was obtained from residue.

MS (ESI/APCI dual): m/z = 200 (M+Na) ⁺

¹ H NMR (600 MHz, chloroform - d ) δ ppm 1.19-1.23 (2H, m), 1.35-1.43 (2H, m), 2.23 (1H, s), 6.74 (1H, br.s.)

(12) Copper (I) chloride (44 mg) was dissolved in a 30% aqueous solution of butylamine (74.1 mL), and hydroxylamine hydrochloride (36.7 mg) was added. After adding N-(1-ethynylcyclopropyl)-2,2,2-trifluoroacetamide (4.3 g) to the reaction mixture, the reaction solution was cooled in an ice bath and stirred for 5 minutes. After adding 4-bromoethynylbenzoic acid (5.0 g) to the reaction mixture and warming to room temperature, hydroxylamine hydrochloride (22.0 mg) was added and stirred for 30 minutes. A saturated aqueous ammonium chloride solution was added to the reaction mixture, and extracted twice with AcOEt. After adding anhydrous magnesium sulfate to the obtained organic layer and drying it, the desiccant was separated by filtration, and the solvent was distilled off under reduced pressure. The obtained pale yellow solid was washed with IPE and dried to obtain 4-((1-(2,2,2-trifluoroacetamide)cyclopropyl)butane-1,3-diyne-1. -yl)benzoic acid (1.8 g, lavender solid, 25%).

MS (ESI/APCI dual): m/z = 344 (M+Na) ⁺

¹ H NMR (600 MHz, DMSO-d ₆ ) δ ppm 1.18-1.29 (2H, m), 1.32-1.43 (2H, m), 7.62 (2H, d, J = 8.3 Hz), 7.83-7.97 (2H, m ), 10.23 (1H, s)

(13) (2S)-N,2-dimethyl-2-(methylamino)-N'-((tetrahydro-2H-pyran-2) obtained in Example 1-(10) 4-((1-(2,2,2-trifluoroethyl) obtained in Example 1-(12) was added to a solution of chloro)propanamide (45.0 mg) in chloroform (3.5 mL) Hydrazine)cyclopropyl)butan-1,3-diyn-1-yl)benzoic acid (50.2 mg), TEA (72.6 μL), DMAPO (4.8 mg) and MNBA (77.7 mg), stirred at room temperature 4 days. A saturated aqueous solution of sodium hydrogencarbonate was added to the mixture and the mixture was extracted three times with chloroform. The obtained organic layer was combined and dried using a phase separator, and then distilled under reduced pressure. The residue obtained was purified by OH type gel chromatography (chloroform / MeOH = 98/2 → 80 / 20) to obtain (2S)-N,2-dimethyl-2-(N-methyl-4). -((1-(2,2,2-trifluoroacetamide)cyclopropyl)butan-1,3-diyn-1-yl)benzamide)-N'-((tetrahydro-2H) -pyran-2-yl)oxy)propanediamine (light yellow solid) (33.0 mg, 34%).

MS (ESI/APCI dual): m/z = 585 (M+Na) ⁺

¹ H NMR (600MHz, CHLOROFORM -d) δ ppm 1.27-1.34 (2H, m), 1.41-1.51 (2H, m), 1.49-1.90 (9H, m), 2.84 (3H, m), 3.15 (3H, m), 3.50-3.74 (1H, m), 3.83-4.09 (1H, m), 4.86-5.06 (1H, m), 6.94-7.62 (6H, m), 9.98-10.57 (1H, m)

By (2S)-N,2-dimethyl-2-(N-methyl-4-((1-(2,2,2-trifluoro)) obtained in Example 1-(13) Indoleamine)cyclopropyl)butan-1,3-diyn-1-yl)benzamide)-N'-((tetrahydro-2H- Deprotection of the trifluoroethenyl group of pyran-2-yl)oxy)propanediamine gives the same compound as the compound obtained in Example 1-(4). Further, Compound 1 can be obtained by deprotecting the tetrahydropyranyl group by the method of Example 1-(5). As a method of deprotecting a trifluoroethane group, for example, a method described in P.G.M. Watts et al., Protective Groups in Organic Synthesis, 4th edition, 2006, John Wiley & Sons, INC.

The protecting group for the amine group of 1-ethynylcyclopropylamine hydrochloride is not limited to the trifluoroethenyl group described in Example 1-(11), and the above-mentioned "protecting group in organic synthesis" may also be used. The protecting groups of the commonly known amine groups described in the above are protected. Examples of the protecting group for the amine group include a urethane-based protecting group such as a third butoxycarbonyl group or an ethoxycarbonyl group, or a trityl group. For example, by using trityl to protect the amine group of 1-ethynylcyclopropylamine hydrochloride to obtain 1-ethynyl-N-tritylcyclopropylamine, the same as in Example 1-( 12) In the same manner as in the case of 1-(13), and using the appropriate deprotection method as described in the above "Protective Group in Organic Synthesis", the trityl group and the tetrahydropyranyl group are sequentially or simultaneously Deprotection, whereby Compound 1 can be obtained.

The effects of the compounds of the present invention were confirmed by the following tests.

Test Example 1 Antibacterial activity evaluation test

The minimum developmental inhibition concentration (MIC) was determined according to the CLSI (American Clinical Laboratory Standardization Association) standard method using the micro liquid dilution method shown below.

For the Legionella pneumophila ATCC33152, the test cells obtained by culturing for 72 hours in BCYE (Buffered charcoal yeast extract) agar medium were suspended at a concentration equivalent to 0.5 McFarland standard. The obtained suspension was diluted to 10 times as an inoculum. 0.005 mL of the inoculum was inoculated into BYE (Buffered yeast extract) α medium containing the test compound, and cultured at 35 ° C for 72 hours.

About Haemophilus influenza ATCC43095, scraped the cultured cells obtained by culturing in chocolate II agar medium for 24 hours, so that Equivalent to a concentration of 0.5 McFarland standard suspension, the obtained suspension was diluted to 10 times as an inoculum. 0.005 mL of the inoculum was inoculated into a HTM (Haemophilus Test Medium) medium containing the test compound, and cultured at 35 ° C for 22 hours.

For the strain other than the above strain, the test cells obtained by culturing for 1 night in a heart infusion agar medium were suspended in a concentration equivalent to 0.5 McFarland standard, and the obtained suspension was diluted to 10 times as an inoculum. 0.005 mL of the inoculum was inoculated to the cation-adjusted Mueller Hinton Agar medium containing the test compound, or the human serum albumin (HSA) was added in such a manner that the final concentration was 5%. The cations were adjusted in Ma-Xin's agar medium and incubated at 35 ° C for 18 hours. The minimum drug concentration at which bacterial development cannot be visually confirmed is taken as the MIC.

The test results of Compound 1 are shown in Table 1.

Test Example 2 Receptive Distribution Test

The minimum developmental inhibitory concentration (MIC) of the clinical isolates of 30 strains of Pseudomonas aeruginosa and 27 strains of Klebsiella pneumoniae was determined, and the MIC which successfully prevented the development of 90% of the strains was calculated and recorded as MIC ₉₀ . The MIC measurement of each clinical isolate was carried out in the same manner as in Test Example 1.

The MIC ₉₀ for Compound 1 was 2 μg/mL in the case of Pseudomonas aeruginosa and 2 μg/mL in the case of Klebsiella pneumoniae.

Test Example 3 Pharmacological effect test on infected animals

Pseudomonas aeruginosa TS88 strain (clinical isolate) was used as a bacterium. The cells obtained by culturing for one night in heart agar extract agar medium were suspended in physiological saline solution and prepared to have a concentration equivalent to 3.5 McFarland standard. The obtained suspension was diluted with a physiological saline solution containing 3 w/v% of mucin so as to be about 6 × 10 ⁵ CFU (Colony-Forming Units) / mL as an inoculum. Mice (ICR (Institute of Cancer Research), male, 4.5 weeks old) were intraperitoneally inoculated with 0.5 mL of inoculum to infect them, and 1 hour after inoculation, Compound 1 was administered intravenously. (6.25 mg/kg) or medium (11 w/v% β-cyclodextrin sulfobutyl ether sodium salt (registered trademark: CAPTISOL, Ligand)).

The survival rate after 3 days of inoculation in Compound 1 group was 100% (8 of 8 patients survived), and the survival rate in the vehicle administration group was 0% (8 patients all died).

Test Example 4 Solubility Test

0.5 mL of physiological saline was added to the excess of Compound 1, and the pH was adjusted to 4 by adding hydrochloric acid, followed by shaking for 24 hours. The solubility after shaking was measured by HPLC (High Performance Liquid Chromatography), and the solubility was measured. This test was carried out at 25 °C.

The solubility of Compound 1 was 15.6 mg/mL.

Test Example 5 Serum protein binding test

Serum protein binding rates were assessed using equilibrium dialysis.

A dimethyl sulfoxide solution of Compound 1 was added to human serum and mouse serum to prepare a 1 μg/mL serum sample. Various serum samples were added to one side of the membrane of the equilibrium dialysis apparatus, and phosphate buffer (pH 7.4) was added to the opposite side, and culture was carried out at 37 ° C for 4 hours. After the completion of the culture, the serum sample and the phosphate buffer were sampled, and acetonitrile/MeOH (9:1) was added thereto, followed by centrifugation. The concentration of Compound 1 in the supernatant was measured by LC-MS/MS to calculate the serum protein binding rate.

The serum protein binding rates of Compound 1 in humans and mice were 59.3% and 61.8%, respectively.

Test Example 6 Rat Maximum Durability (MTD) Test

A total of 10 (3 mL/kg, 3 mL/min) of Compound 10 (control), 400, 600 and 800 mg/kg were administered to SD male rats of 1~3 cases/group for their maximum durability (MTD). . 11% CAPTISOL (registered trademark, Ligand) (pH 4) was administered in the same manner to the control group.

One in 1 case of 800 mg/kg group confirmed death. Regarding the death case, red tears, distorted body and barking behavior were confirmed during the administration, and died shortly after the end of the administration. The cause of death is not clear.

Regarding the survival example, red tears, twisted body and squeaking behavior, and tail discoloration (cyan) were confirmed in the 400 and 600 mg/kg group, and the prone position was confirmed in the 600 mg/kg group.

No changes worth mentioning were identified in the anatomical examination.

No significant change was confirmed with respect to the control group.

When a single intravenous administration of Compound 1 to male rats was confirmed to be death at 800 mg/kg, it was estimated that the MTD was 600 mg/kg.

Test Example 7 hERG channel binding assay

Human type hERG (Human ether) diluted with binding assay buffer [final concentration of 10 mM HEPES (pH 7.4), 71.5 mM NaCl, 60 mM KCl, 2 mM MgCl ₂ , 1 mM CaCl ₂ , 0.1% BSA] -a-go-go-related gene, a human ether-a-go-go related gene) channel stably exhibits membrane components prepared by cells. A membrane fraction (final concentration: 30 μg/mL), [ ³⁵ S]MK-499 (final concentration: 0.5 nM) was added to a 96-well plate in which the test compound was previously dispensed, and allowed to react at room temperature for 75 minutes. The reaction solution was suction-filtered on a GF/C filter plate, and washed with a washing buffer [HEPES (pH 7.4) having a final concentration of 10 mM, 131.5 mM NaCl, 2 mM MgCl ₂ , 1 mM CaCl ₂ ] for 5 times. Dry at 45 ° C. Micro Scint-O was added to the dried filter plate, and radioactivity was measured using a microplate luminescence analyzer NXT.

In the above reaction, the difference in the amount of [ ³⁵ S]MK-499 binding obtained in the presence of the test compound is recorded as the specific binding amount, and [ ³⁵ S]MK-499 obtained in the presence of MK-499. The amount of binding is recorded as the amount of non-specific binding. The S50 analysis of the concentration response curve was carried out based on the specific binding amount in the presence of the test compound in the absence of the specific binding amount of the test compound, and the IC ₅₀ value was calculated.

Compound 1 has an IC ₅₀ value of 30 μM or more.

Test Example 8 Safety pharmacological test of guinea pig cardiovascular system under anesthesia

The guinea pigs were intravenously administered with 10 mg/kg (control) and 70 mg/kg (5 mL/kg/30 min) intravenously under anesthesia for 30 minutes to study the effect on the electrocardiogram. For the control group, 11% CAPTISOL (registered trademark, Ligand) (pH 4) was administered in the same manner. Under artificial respiration and pentobarbital anesthesia, the electrocardiogram was used to measure the standard induction (II-induced) electrocardiogram of the extremities, and the analysis was performed every 5 minutes from 15 minutes before the start of the test substance administration to 30 minutes after the end of the administration. . Use Bazett style (QTcB=QT/ RR) The corrected QTcB is obtained, and the rate of change (ΔQTcB) from the reference line value [the average value of the three time points (-15, -10, and -5 minutes before the administration of the test substance)] is calculated.

The ΔQTcB at the end of the administration of Compound 1 was -0.1%, and the maximum was +0.2% (20 minutes and 45 minutes after the start of administration) (control group: -4.6 to -1.0%), and it was not confirmed that it was administered by administration. Substance and significant impact on QTcB. It was confirmed that Compound 1 is also excellent in safety.

[Industrial availability]

The novel hydroxamic acid derivative of the present invention has strong antibacterial activity against Gram-negative bacteria represented by Pseudomonas aeruginosa and its resistant bacteria, and can be used as a pharmaceutical.

Claims (4)

A compound or a pharmaceutically acceptable salt thereof, which is represented by the formula [1], 2-[{4-[4-(1-aminocyclopropyl)butan-1,3-diyn-1-yl) Benzyl hydrazino}(methyl)amino]-N-hydroxy-N',2-dimethylpropanediamine, A compound or a pharmaceutically acceptable salt thereof, which is represented by the formula [2]: (2S)-2-[{4-[4-(1-aminocyclopropyl)butane-1,3-diyne -1-yl]benzhydryl}(methyl)amino]-N-hydroxy-N',2-dimethylpropanediamine, A pharmaceutical composition comprising a compound of claim 1 or 2, or a pharmaceutically acceptable salt thereof. An antibacterial agent comprising a compound of claim 1 or 2, or a pharmaceutically acceptable salt thereof.