WO2008039641A2 - Composés phényléther-thiénopyridone substitués à activité antibactérienne - Google Patents

Composés phényléther-thiénopyridone substitués à activité antibactérienne Download PDF

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WO2008039641A2
WO2008039641A2 PCT/US2007/078161 US2007078161W WO2008039641A2 WO 2008039641 A2 WO2008039641 A2 WO 2008039641A2 US 2007078161 W US2007078161 W US 2007078161W WO 2008039641 A2 WO2008039641 A2 WO 2008039641A2
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propylamino
thieno
benzylamino
pyridine
dibromo
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PCT/US2007/078161
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WO2008039641A3 (fr
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Joseph Guiles
Xicheng Sun
Nebojsa Janjic
Sarah Strong
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Replidyne, Inc.
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D495/00—Heterocyclic compounds containing in the condensed system at least one hetero ring having sulfur atoms as the only ring hetero atoms
    • C07D495/02—Heterocyclic compounds containing in the condensed system at least one hetero ring having sulfur atoms as the only ring hetero atoms in which the condensed system contains two hetero rings
    • C07D495/04—Ortho-condensed systems

Definitions

  • the present invention relates to novel bicyclic heteroaromatic compounds having a left hand side phenylether and right hand side thienopyridone, and in particular to the use of these compounds as inhibitors of bacterial methionyl tRNA synthetases (MetRS), to processes for their preparation and to their uses in therapy as antibacterial agents, and in particular to their uses in therapy of Clostridium difficile based infections.
  • MetalRS bacterial methionyl tRNA synthetases
  • tRNA synthetases are involved in protein biosynthesis so that inhibition thereof may be expected to lead to a cessation of cell growth.
  • the compound mupirocin produced by the organism Pseudomonas fluorescens, is an antibacterial agent and is used as the active ingredient in the product Bactroban®, marketed by GlaxoSmithKline. Mupirocin has been shown to be an inhibitor of the isoleucyl tRNA synthetase.
  • Each tRNA synthetase represents a separate target for drug discovery.
  • tRNA synthetase inhibitors which are selective for bacterial cells over mammalian cells are of considerable therapeutic interest as they have the potential to be used as antibacterial agents.
  • C. difficile C. difficile is becoming a more prevalent infectious agent, where one to three percent of healthy individuals are carriers of the organism. (Bartlett & Perl, N. Engl. J Med., 353, 2503-2505, 2005; Clabots et al., J Infect. Dis., 166, 561-567, 1992; McFarland et al., N. Engl. J Med., 320, 204-210, 1989).
  • the risk of infection and disease becomes increasingly prevalent in the immunodeficient, elderly, and especially to the elderly in healthcare settings, e.g., nursing home, hospital, doctors office, etc. Few conventional antibacterial drugs have shown promise in the treatment of C. difficile, in fact only vancomycin is approved by the FDA for treatment of C. difficile associated diarrhea (CDAD).
  • CDAD C. difficile associated diarrhea
  • bicyclic heteroaromatic compounds that are potent inhibitors of bacterial MetRS. This new class of compounds is shown to have broad applicability as antibacterial agents for numerous Gram-positive and Gram-negative bacteria. MetRS inhibitors of the invention have superior activity against Gram-positive organisms, in particular C. difficile, as compared to Gram-negative organisms.
  • bicyclic heteroaromatic compounds of the invention have a left hand side (LHS) phenylether and a right hand side (RHS) thienopyridone.
  • R 1 is selected from the group consisting of aryl and heteroaryl groups, including but not limited to substituted and unsubstituted benzene, toluene, phenol, anisole, thiazole, thiazolidine and pyridine, alkenes, imines, and other like substituents;
  • R 2 is independently selected from halo, cyano, hydroxyl, (Ci- 6 )alkyl (optionally substituted by halo, hydroxyl, amino, carboxy, or (Cr 6 ) alkoxycarbonyl), (C 1 - ⁇ ) cycloalkyl, (Cr 6 ) alkoxy, amino, mono- or di-(C 1 - 6 )alkylamino, acylamino, carboxy, (Ci- 6 )alkoxycarbonyl, carboxy(Ci_ 6 )alkyloxy, (Ci- 6 )alkylthio, (C 1 - ⁇ )alkylsulphinyl, (Ci- 6 )alkylsulphonyl, sulphamoyl, mono- and di-(Ci- 6 )alkylsulphamoyl, carbamoyl, mono- and di-(Ci- 6 )alkylcarbamoyl, and hetero
  • R 3 is selected from a halo, (Ci ⁇ alkyl, (C 2 - 3 )alkenyl, (C 2 - 3 )alkynyl or other like substituents;
  • n is one, two or three;
  • m is 0, 1, 2 or 3.
  • R 1 is selected from the group consisting of substituted and unsubstituted benzene,such as toluene, phenol, anisole, substituted and unsubstituted thiazole, such as methylthiazole, substituted and unsubstituted pyridine, and alkenyl groups ethanimine, and (C 1 -
  • R 2 is one or more halogen substituents (preferably bromine, iodine and/or chlorine), one or more sulfane substituents, e.g., methylsulfane, or a combination of a halogen and sulfane substituents;
  • halogen substituents preferably bromine, iodine and/or chlorine
  • sulfane substituents e.g., methylsulfane, or a combination of a halogen and sulfane substituents
  • n is one, two or three;
  • m is 0, 1, 2, or 3.
  • Salts may be formed from inorganic and organic acids.
  • suitable inorganic and organic acids from which pharmaceutically acceptable salts of compounds of formula (I) or formula (IA) may be formed include maleic, fumaric, benzoic, ascorbic, pamoic, succinic, bismethylene-salicylic, methanesulfonic, ethanedisulfonic, acetic, propionic, tartaric, salicylic, citric, gluconic, aspartic, stearic, palmitic, itaconic, glycolic, p- aminobenzoic, glutamic, benzenesulfonic, hydrochloric, hydrobromic, sulfuric, cyclohexylsulfamic, phosphoric and nitric acids.
  • alkyl and similar terms such as “alkoxy” includes all straight chain and branched isomers. Representative examples thereof include methyl, ethyl,
  • alkenyl and alkynyl include all straight chain and branched isomers. Representative examples thereof include vinyl, ethynyl and 1-propynyl.
  • Preferred substituents for alkyl and alkenyl groups include, for example, and unless otherwise defined, halogen, cyano, azido, nitro, carboxy, (C 1 _g)alkoxycarbonyl, carbamoyl, mono- or di-(Ci _g)alkylcarbamoyl, sulpho, sulphamoyl, mono- or di-(Ci .
  • alkylsulphamoyl amino, mono- or di-(C] ⁇ .g)alkylamino, acylamino, ureido, (C ⁇ .g)alkoxycarbonylamino, 2,2,2-trichloroethoxycarbonylamino, aryl, heterocyclyl, hydroxy, (Ci .g)alkoxy, acyloxy, oxo, acyl, 2-thienoyl, (Ci _g)alkylthio, (Ci _g)alkylsulphinyl, (C 1.
  • aryl includes, unless otherwise defined, phenyl or naphthyl optionally substituted with up to five, preferably up to three substituents.
  • an aryl group may have up to three substituents.
  • Preferred substituents for an aryl group include, for example, and unless otherwise defined, halogen, cyano, (Ci -g)alkyl, mono to perfluoro(Ci -3)alkyl, (C3"7)cycloalkyl, (C2-6)alkenyl,
  • heteroaryl includes single or fused rings comprising up to four hetero-atoms in the ring selected from oxygen, nitrogen and sulphur.
  • the heteroaryl ring comprises from 4 to 7, preferably 5 to 6, ring atoms.
  • a fused heteroaryl ring system may include carbocyclic rings and need only include one heterocyclic ring.
  • heterocyclyl includes aromatic and non-aromatic single or fused rings comprising up to four hetero-atoms in the ring selected from oxygen, nitrogen and sulphur.
  • the heterocyclic ring comprises from 4 to 7, preferably 5 to 6, ring atoms.
  • a fused heterocyclic ring system may include carbocyclic rings and need only include one heterocyclic ring.
  • a heteroaryl or a heterocyclyl group may have up to three substituents.
  • Preferred substituents include those previously mentioned for an aryl group as well as oxo.
  • halogen and “halo” include fluorine, chlorine, bromine, and iodine and fluoro, chloro, bromo, and iodo, respectively.
  • the compounds of the present invention are suitably provided in substantially pure form, for example at least 50% pure, suitably at least 60% pure, advantageously at least
  • All impure or less pure forms of a compound according to the invention may, for example, be used in the preparation of more pure forms of the same compound or of a related compound (for example a corresponding derivative) suitable for pharmaceutical use.
  • certain compounds of the present invention may comprise one or more chiral centers so that compounds may exist as stereoisomers, including diastereoisomers and enantiomers.
  • Embodiments of the invention cover all such stereoisomers, and mixtures thereof, including racemates and mixtures having an enantiomeric excess of one of the enantiomers.
  • the compounds of formula (I)-(XV) may be prepared by methods described herein (see Example 1) or by methods described in the prior art that are incorporated by reference herein below.
  • a compound of formula (I) is generally prepared by reductive amination of 5-(3-aminopropylamino)thieno[3,2-b]pyridin-7-one (formula XVIII) with a substituted phenylether aldehyde (formula XVII) (see Example 1, Synthesis Scheme 1).
  • the compounds of this invention are active against a range of important pathogenic bacteria, including Gram positive organisms, such as Staphylococci, for instance S. aureus Oxford and coagulase negative strains of Staphylococci such as S. epidermidis; Streptococci, for instance S. pyogenes ATCC19615 and S.
  • compounds of this invention are also active against gram negative organisms, such as Haemophilus, for instance H. influenzae Ql; Moraxella, for instance M. catarrhalis 1502; Escherichia, for instance E. coli DCO; and Helicobacter, for instance H. pylori ATCC 700824.
  • the most preferred compounds of the present invention will be active against the organisms C. difficile, S. aureus, S. pneumoniae, E. faecalis, E. faecium, H. influenzae, M. catarrhalis, and H. pylori.
  • compounds of this invention are active against Staphylococci organisms such as S. aureus and coagulase negative strains of Staphylocci such as S. epidermidis which are resistant (including multiply-resistant) to other antibacterial agents, for instance, ⁇ - lactam antibiotics such as, for example, methicillin, macrolides, aminoglycosides, oxazolidinones, and lincosamides.
  • Staphylococci organisms such as S. aureus and coagulase negative strains of Staphylocci such as S. epidermidis which are resistant (including multiply-resistant) to other antibacterial agents, for instance, ⁇ - lactam antibiotics such as, for example, methicillin, macrolides, aminoglycosides, oxazolidinones, and lincosamides.
  • Compounds of the present invention are also active against strains of E. faecalis including vancomycin resistant strains and therefore of use in treating infections associated with
  • VRE organisms VRE organisms. Furthermore, compounds of the present invention are useful in the treatment of
  • Staphylococci organisms which are resistant to mupirocin.
  • Compounds of the present invention have particularly potent activity against strains of Clostridium including C. difficile. Therefore, compounds of the invention can be used to treat infections associated with C. difficile, e.g., pseudomembraneous colitis, toxic megacolin, and other antibiotic associated diarrheas (AAD).
  • infections associated with C. difficile e.g., pseudomembraneous colitis, toxic megacolin, and other antibiotic associated diarrheas (AAD).
  • Compounds of the present invention show little or no activity against mammalian cells. This provides an optimal combination of high activity against pathogenic bacteria and low or no activity against mammalian cells, allowing for the use of the compounds of the invention in human treatments.
  • Bacterial infections which may be treated include gastrointestinal tract infections, respiratory tract infections, otitis media, meningitis, endocarditis, skin and soft tissue infections in man, mastitis in cattle, and also respiratory infections in farm animals such as pigs and cattle.
  • the present invention provides a method of treating bacterial infection in human or non-human animals, which method comprises administering a therapeutically effective amount of a compound of formula (I)-(XV) as hereinbefore defined, to a human or non-human animal in need of such therapy.
  • a compound of the present invention which has a broad spectrum of antibacterial activity, including activity against both Gram positive and Gram negative bacteria will be of general use in the community for the empiric treatment of community acquired infections.
  • a compound of the present invention with a more limited spectrum, for instance activity against Gram positive bacteria is more likely to be used in circumstances where the causative pathogenic organism has been identified..
  • the present invention provides a pharmaceutical composition
  • a pharmaceutical composition comprising a compound of formula (I)-(XV) together with a pharmaceutically acceptable carrier or excipient.
  • the present invention further provides pharmaceutical compositions comprising combinations of compounds of formula (I)-(XV) together with a pharmaceutically acceptable carrier or excipient.
  • a pharmaceutical composition of the invention can include a compound of formula (III) and a compound of formula (VI) in combination with the carrier or excipient.
  • the present invention also provides a method of treating bacterial infections in mammals, especially in humans and in domesticated animals, which comprises administering a compound of the invention, or a composition according to the invention, to a patient in need thereof.
  • the invention further provides the use of compounds of the invention in the preparation of a medicament composition for use in the treatment of bacterial infections.
  • compositions according to the invention may be formulated for administration in any convenient way for use in human or veterinary medicine, by analogy with other antibiotics.
  • compositions according to the invention may be formulated for administration by any route, for example oral, topical, parenteral, or rectal.
  • the compositions may, for example, be made up in the form of tablets, capsules, powders, granules, lozenges, creams, suppositories, ointments, gels, lotions, syrups, or liquid preparations, for example solutions or suspensions, which may be formulated for oral use or in sterile form for parenteral administration by injection or infusion.
  • Tablets and capsules for oral administration may be in unit dosage form, and may contain conventional excipients including, for example, binding agents, for example, syrup, acacia, gelatin, sorbitol, tragacanth, or polyvinylpyrrollidone; fillers, for example lactose, sucrose, maize-starch, calcium phosphate, sorbitol or glycine; tabletting lubricants, for example magnesium stearate, talc, polyethylene glycol or silica; disintegrants, for example potato starch; and pharmaceutically acceptable wetting agents, for example sodium lauryl sulphate.
  • the tablets may be coated according to methods well known in normal pharmaceutical practice.
  • Oral liquid preparations may be in the form of, for example, aqueous or oily suspensions, solutions, emulsions, syrups or elixirs, or may be presented as a dry product for reconstitution with water or another suitable vehicle before use.
  • Such liquid preparations may contain conventional additives, including, for example, suspending agents, for example sorbitol, methyl cellulose, glucose syrup, gelatin, hydroxyethyl cellulose, carboxymethyl cellulose, aluminium stearate gel or hydrogenated edible fats; emulsifying agents, for example lecithin, sorbitan monooleate or acacia; non-aqueous vehicles (which may include edible oils), for example almond oil, oily esters (for example glycerine), propylene glycol, or ethyl alcohol; preservatives, for example methyl or propyl p-hydroxybenzoate or sorbic acid; and, if desired, conventional flavoring and color agents.
  • suspending agents for example sorbitol, methyl cellulose, glucose syrup, gelatin, hydroxyethyl cellulose, carboxymethyl cellulose, aluminium stearate gel or hydrogenated edible fats
  • emulsifying agents for example lecithin, sorbitan monooleate
  • compositions according to the invention intended for topical administration may, for example, be in the form of ointments, creams, lotions, eye ointments, eye drops, ear drops, impregnated dressings, and aerosols, and may contain appropriate conventional additives, including, for example, preservatives, and solvents to assist drug penetration, and emollients in ointments, gels, and creams.
  • Such topical formulations may also contain compatible conventional carriers, for example cream or ointment bases, and ethanol or oleyl alcohol for lotions.
  • Such carriers may constitute from about 1% to about 98% by weight of the formulation; more usually they will constitute up to about 80% by weight of the formulation.
  • compositions according to the invention may be formulated as suppositories, which may contain conventional suppository bases, for example cocoa-butter or other glycerides.
  • Compositions according to the invention intended for parenteral administration may conveniently be in fluid unit dosage forms, which may be prepared utilizing the compound and a sterile vehicle, water being preferred. The compound, depending on the vehicle and concentration used, may be either suspended or dissolved in the vehicle. In preparing solutions, the compound may be dissolved in water for injection and filter-sterilized before being filled into a suitable vial or ampoule, which is then sealed.
  • conventional additives including, for example, local anesthetics, preservatives, and buffering agents can be dissolved in the vehicle.
  • the composition may be frozen after being filled into the vial, and the water removed under vacuum; the resulting dry lyophilized powder may then be sealed in the vial and a accompanying vial of water for injection may be supplied to reconstitute the liquid prior to use.
  • Parenteral suspensions may be prepared in substantially the same manner except that the compound is suspended in the vehicle instead of being dissolved and sterilization cannot be accomplished by filtration. The compound may instead be sterilized by exposure to ethylene oxide before being suspended in the sterile vehicle.
  • a surfactant or wetting agent is included in such suspensions in order to facilitate uniform distribution of the compound.
  • a compound or composition according to the invention may suitably be administered to the patient in an antibacterially effective amount.
  • a composition according to the invention may suitably contain from 0.1% by weight, preferably from 10 to 60% by weight, of a compound according to the invention (based on the total weight of the composition), depending on the method of administration. Note that where a composition includes two or more compounds of the invention, the total weight/weight of the inhibitors is equal to from 0.1% to about 60%, i.e., compound (II) contributes 2% and compound (III) contributes 2% for a total of 4%.
  • the compounds according to the invention may suitably be administered to the patient at a daily dosage of from 1.0 to 100 mg/kg of body weight.
  • a daily dosage of from 1.0 to 100 mg/kg of body weight.
  • an adult human of approximately 70 kg body weight
  • the dosage for adult humans is from 5 to 40 mg/kg per day. Higher or lower dosages may, however, be used in accordance with normal clinical practice.
  • each unit dose may suitably comprise from 25 to 1000 mg, preferably from 50 to 500 mg, of a compound according to the invention.
  • Examples 20-24 below illustrate the potent antibacterial activity of the compounds of the present invention.
  • Example 21, 23 and 24 illustrate the surprisingly potent antibacterial activity of compounds of the invention against C. difficile, an organism that has proven to be difficult to treat using conventional antibiotic therapies.
  • the resultant protected aldehyde (XXIII) was dissolved in anhydrous DMF (60 mL) and mixed with alkyl mesylate or alky bromide (2 to 3 equivalents), potassium iodide (1.5 equivalents) and potassium carbonate (1.5 equivalents). The reaction was stirred at 40 0 C over 48 h. Then, the reaction mixture was diluted with water (200 mL), and the aqueous layer was extracted with EtOAc (80 mL X 4). The combined organic layers were partitioned with water and brine, and dried over anhydrous MgSO 4 , filtered, and solvent removed in vacuo. Purification by silica gel column chromatography, the product eluted with EtOAc/hexane (10% to 40%), afforded the desired protected aldehyde (XXIV) as a solid.
  • aldehyde intermediate XVII was prepared via method A, followed by reductive amination (method B) with XVIII to give 5-(3- ⁇ 3,5-Dibromo-2-[2-(4-methyl-thiazol-5- yl)-ethoxy]-benzylamino ⁇ -propylamino)-4H-thieno[3,2-b]pyridin-7-one.
  • aldehyde intermediate XVII was prepared via method A, followed by reductive amination (method B) with XVIII to give 5- ⁇ 3-[3,5-Dibromo-2-(2-pyridin-3-yl- ethoxy)-benzylamino]-propylamino ⁇ -4H-thieno[3,2-b]pyridin-7-one.
  • aldehyde intermediate XVII was prepared via method A, followed by reductive amination (method B) with XVIII to give 5-(3- ⁇ 3,5-Dibromo-2-[2-(4,5-dimethyl- thiazol-2-yl)-ethoxy]-benzylamino ⁇ -propylamino)-4H-thieno[3,2-b]pyridin-7-one.
  • aldehyde intermediate XVII was prepared via method A, followed by reductive amination (method B) with XVIII to give 5-[3-(3,5-Dibromo-2-phenethyloxy- benzylamino)-propylamino]-4H-thieno[3,2-b]pyridin-7-one.
  • 1 H NMR 400 MHz, CD 3 OD: ⁇ 7.69(d, IH), 7.65 (d, IH), 7.50 (d, IH), 7.29-7.20
  • aldehyde intermediate XVII was prepared via method A, followed by reductive amination (method B) with XVIII to give 5- ⁇ 3-[3,5-Dibromo-2-(3-pyridin-3-yl- propoxy)-benzylamino]-propylamino ⁇ -4H-thieno[3,2-b]pyridin-7-one.
  • aldehyde intermediate XVII was prepared via method A, followed by reductive amination (method B) with XVIII to give 5-(3- ⁇ 3,5-Dibromo-2-[2-(3,4-dichloro- phenyl)-ethoxy]-benzylamino ⁇ -propylamino)-4H-thieno[3,2-b]pyridin-7-one.
  • aldehyde intermediate XVII was prepared via method A, followed by reductive amination (method B) with XVIII to give 5-(3- ⁇ 3,5-Dibromo-2-[2-(4-methoxy- phenyl)-ethoxy]-benzylamino ⁇ -propylamino)-4H-thieno[3,2-b]pyridin-7-one.
  • aldehyde intermediate XVII was prepared via method A, followed by reductive amination (method B) with XVIII to give5- ⁇ 3-[3,5-Dibromo-2-(2-p-tolyl-ethoxy)- benzylamino]-propylamino ⁇ -4H-thieno[3,2-b]pyridin-7-one.
  • aldehyde intermediate XVII was prepared via method A, followed by reductive amination (method B) with XVIII to give 5-(3- ⁇ 3,5-Dibromo-2-[2-(3-fluoro-phenyl)- ethoxy]-benzylamino ⁇ -propylamino)-4H-thieno[3,2-b]pyridin-7-one.
  • aldehyde intermediate XVII was prepared via method A, followed by reductive amination (method B) with XVIII to give 5-(3- ⁇ 3,5-Dibromo-2-[2-(4-chloro-phenyl)- ethoxy]-benzylamino ⁇ -propylamino)-4H-thieno[3,2-b]pyridin-7-one.
  • MetRS was amplified and cloned into pETcoco-2.
  • the following primers were used to amplify DNA from genomic DNA: 5 1 -CTGCAGAGCTAGCAAACCGAGTTTTTATGTAAC-3 1 (forward) (SEQ ID NO:1), 5'-CTTTCTAAGCTTCTACTAACGAACCTCGGATCC-S' (reverse) (SEQ ID NO:2).
  • Amplified DNA was treated with Sphl and HindIII restriction endonucleases, which were heat-inactivated after digestion. The fragment was ethanol- precipitated and combined with pETcoco-2 vector (Novagen) that had been treated with the same enzymes plus shrimp alkaline phophatase.
  • the fragments were ligated and the ligation mixture transformed into competent DHlO E. coli.
  • Transformants were plated on F-medium plus glucose with 50 ug/ml ampicillin. Growth in glucose maintains the repressed state of the pBAD promoter driving expression of the replicator Trf A, thus maintaining low copy number.
  • the resulting expression clone, pETcoco-Cdiff-MRS, was confirmed by sequencing of the insert in both directions.
  • Fraction I Fraction I was loaded onto a 15 mL Ni-NTA column which was equilibrated in Load Buffer (50 mM Tris-HCl, pH 7.5, 10% glycerol, 40 mM KCl, 10 mM Imidazole, pH 6.8, and 7 mM beta mercaptoethanol).
  • Load Buffer 50 mM Tris-HCl, pH 7.5, 10% glycerol, 40 mM KCl, 10 mM Imidazole, pH 6.8, and 7 mM beta mercaptoethanol.
  • the column was washed with 10 column volumes of Wash Buffer (50 mM Tris-HCl, pH 7.5, 10% glycerol, 800 mM KCl, 20 mM Imidazole, pH 6.8, and 7 mM beta mercaptoethanol).
  • the protein was eluted in 10 column volume gradient from Wash Buffer to Elution Buffer (50 mM Tris-HCl, pH 7.5, 10% glycerol, 40 mM KCl, 250 mM Imidazole, pH 6.8, and 7 mM beta mercaptoethanol) at 0.5 niL/min collecting 3 mL fractions. Fractions were collected and analyzed for protein by SDS-PAGE. Fractions were assayed in the C.
  • Fraction II Fraction II (60 mg at 1.3 mg/ml). Fraction II had a specific activity of 3.2 x 10 5 units per mg. The purity was estimated at greater than 97% based on densitometry of an SDS-PAGE gel stained with Coomassie blue.
  • Example 20 Compounds of the Present Invention Have Potent Enzyme Activity Against MetRS
  • MetRS inhibitor compounds are competitive inhibitors of methionine and uncompetitive inhibitors of ATP.
  • Example 21 Compounds of the Present Invention Have Potent Antibacterial Activity Against C. difficile
  • Organisms All compounds were tested for antibacterial activity against a collection of non-repeat clinical isolates of C. difficile. The organisms were stored frozen in Brucella broth supplemented with 20% glycerol. The organisms were retrieved from the freezer and subcultured twice onto CDC agar to ensure purity and growth. The plates were incubated under anaerobic conditions for at least 24 hours. Bacterial colonies were examined for morphology; yellow color, ground glass texture and characteristic odor. The control organism tested was Bacteroides fragilis ATCC 25285.
  • Antimicrobial susceptibility testing was conducted by the agar dilution method on Brucella agar supplemented with vitamin K ⁇ hemin and 5% laked sheep blood in accordance with CLSI guidelines (CLSI, Ml 1-A2). The test compounds were serially diluted and added to molten supplemented Brucella agar. Drug free plates were inoculated before and after inoculation of each antimicrobial plate series and were used as growth controls. Anaerobic/aerobic growth controls were conducted on drug free plates after two sets of drug plates.
  • Results Data from the present example shows MIC 9 0 for formulas (II) to (XV) which ranged from 0.5 to >32 ⁇ g/ml. These results indicate the potent activity of the compounds of the present invention against C. difficile, typically around 1.0 ⁇ g/ml.
  • IC 50 data indicates that the compounds of the present invention are specific for C. difficile, showing little or no activity against mammalian MetRS. MetRS inhibitor compounds show potent activity against C. difficile and Gram-positive aerobic bacteria while sparing normal gut flora.
  • Example 22 Compounds of the Present Invention Have Potent Antibacterial Activity against Other Bacteria
  • Several compounds (formulas (II), (III) and (IV) + others (all were tested) of the present invention were tested for antibacterial activity against a panel of Gram-positive bacteria.
  • Compounds were tested against Gram-positive aerobic bacteria using the CLSI-reference broth microdilution method. Data was obtained against S. aureus, E. faecalis, E. faecium, S. pyogenes, S. epidermidis and S. haemolyticus .
  • the compounds tested demonstrated potent antibacterial activity against all isolates with a MIC range of ⁇ 0.008-8 ⁇ g/ml, including resistant strains of S.
  • Example 23 Compounds of the Present Invention Show Strong Therapeutic Utility During in vivo Trials:
  • MetRS inhibitors Animal studies were performed to determine the efficacy of MetRS inhibitors for treating C. difficile -infections.
  • the MetRS inhibitors tested were 5-[3-((R)-6,8-Dibromo- chroman-4-ylamino)-propylamino]-4H-thieno[3,2-b]pyridine-7-one (both racemic mixture and the R enantiomer), 5-[3-((R)-8-Bromo-6-chloro-chroman-4-ylamino)-propylamino]-4H- thieno[3,2-b]pyridin-7-one, and 2-(3- ⁇ 3,5-Dibromo-2-[2-(4-methyl-thiazol-5-yl)-ethoxy]- benzylamino ⁇ -propylamino)-lH-quinolin-4-one.
  • Results were compared to C. difficile-infected hamsters treated with the conventional antibiotic, vancomycin. Infected hamsters were treated with either a solution or suspension of a MetRS inhibitor at 5 to 50 mg/kg or vancomycin at 2.5, 5 or 25 mg/kg. There were eight hamsters per group with the final endpoint of the experiment being survival. Expired hamsters were examined for GI condition.
  • C. difficile The pathogenicity of C. difficile is associated with its ability to produce the extracellular toxins A and B. Hypertoxinogenic strains are responsible for recent outbreaks with high mortality. In contrast, isolates that do not produce toxins are non-pathogenic. Since toxin production requires active protein synthesis, inhibition of the protein synthesis machinery is expected to suppress de novo toxin production. Therefore, MetRS inhibitors were evaluated for their effect on C. difficile toxin production in vitro.
  • C. difficile strain ATCC43255 was grown and maintained anaerobically on CDC anaerobe agar (Remel, Lenexa, KS).
  • CDC anaerobe agar Remel, Lenexa, KS.
  • BHI brain heart infusion
  • BHI brain heart infusion
  • the cells were grown anaerobically for 24 h at 35°C in 96-well brain heart infusion (BHI) broth cultures.
  • Spent medium was then replaced with fresh broth containing MetRS inhibitors and control agents at a concentration range of 0.015 - 16 ⁇ g/mL. After 4 days, growth and cell viability were monitored by optical density measurements at 595 nm and by culture on CDC anaerobe agar, respectively. Culture supernatants were collected, and toxin A was detected by ELIFA (enzyme-linked immuno-flow assay) using an anti toxin A monoclonal antibody (Novus Biologicals, Centennial, CO). Results:
  • MetRS inhibitors demonstrate inhibitory effects on both growth and toxin production of C. difficile in broth cultures. Furthermore, toxin production was effectively blocked in stationary phase cultures. As a consequence of this suppression of toxin production by bacteriostatic MetRS inhibitors, C. difficile becomes essentially non-toxinogenic and thus non-pathogenic. This effect is unique to protein synthesis inhibitors, such as MetRS inhibitors, whose mode-of-action does not require that the bacteria are actively growing.

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  • Heterocyclic Carbon Compounds Containing A Hetero Ring Having Oxygen Or Sulfur (AREA)

Abstract

L'invention concerne de nouveaux composés hétéroaromatiques bicycliques étant des inhibiteurs de la méthionyl-ARNt synthétase bactérienne (MetRS). Les composés selon l'invention ont un constituant phényléther côté gauche et un constituant thiénopyridone côté droit. L'invention concerne également des procédés de préparation de ces composés et leur utilisation en thérapie en tant qu'agents antibactériens, notamment en tant qu'agents anti-clostridium difficile.
PCT/US2007/078161 2006-09-26 2007-09-11 Composés phényléther-thiénopyridone substitués à activité antibactérienne WO2008039641A2 (fr)

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Cited By (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2008039641A3 (fr) * 2006-09-26 2008-12-04 Replidyne Inc Composés phényléther-thiénopyridone substitués à activité antibactérienne
WO2010083318A2 (fr) 2009-01-14 2010-07-22 Dow Agrosciences Llc Compositions fongicides incluant des dérivés d'hydrazone et du cuivre
US7973050B2 (en) 2006-09-26 2011-07-05 Crestone, Inc. Enantiomeric compounds with antibacterial activity
US7994192B2 (en) 2006-09-26 2011-08-09 Crestone, Inc. Substituted thienopyridone compounds with antibacterial activity
US8658670B2 (en) 2006-09-26 2014-02-25 Crestone, Inc. Methods and compounds for treatment of clostridium based infection
US8697720B2 (en) 2006-09-26 2014-04-15 Crestone, Inc. Substituted phenylether-thienopyridone compounds with antibacterial activity
CN115073489A (zh) * 2021-03-15 2022-09-20 同济大学 一种噻吩骈吡啶酮类化合物、制备方法及一种抗真菌药物

Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2008039642A2 (fr) * 2006-09-26 2008-04-03 Replidyne, Inc. Procédés et composés de traitement d'infections par clostridium

Family Cites Families (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2008039641A2 (fr) * 2006-09-26 2008-04-03 Replidyne, Inc. Composés phényléther-thiénopyridone substitués à activité antibactérienne

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2008039642A2 (fr) * 2006-09-26 2008-04-03 Replidyne, Inc. Procédés et composés de traitement d'infections par clostridium

Cited By (12)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2008039641A3 (fr) * 2006-09-26 2008-12-04 Replidyne Inc Composés phényléther-thiénopyridone substitués à activité antibactérienne
US7973050B2 (en) 2006-09-26 2011-07-05 Crestone, Inc. Enantiomeric compounds with antibacterial activity
US7994192B2 (en) 2006-09-26 2011-08-09 Crestone, Inc. Substituted thienopyridone compounds with antibacterial activity
US8658670B2 (en) 2006-09-26 2014-02-25 Crestone, Inc. Methods and compounds for treatment of clostridium based infection
US8697720B2 (en) 2006-09-26 2014-04-15 Crestone, Inc. Substituted phenylether-thienopyridone compounds with antibacterial activity
WO2010083318A2 (fr) 2009-01-14 2010-07-22 Dow Agrosciences Llc Compositions fongicides incluant des dérivés d'hydrazone et du cuivre
US8455394B2 (en) 2009-01-14 2013-06-04 Dow Agrosciences, Llc Fungicidal compositions including hydrazone derivatives and copper
US8461078B2 (en) 2009-01-14 2013-06-11 Dow Agrosciences Llc Fungicidal compositions including hydrazone derivatives and copper
US8476194B2 (en) 2009-01-14 2013-07-02 Dow Agrosciences, Llc Fungicidal compositions including hydrazone derivatives and copper
US8715745B2 (en) 2009-01-14 2014-05-06 Dow Agrosciences, Llc. Fungicidal compositions including hydrazone derivatives and copper
CN115073489A (zh) * 2021-03-15 2022-09-20 同济大学 一种噻吩骈吡啶酮类化合物、制备方法及一种抗真菌药物
CN115073489B (zh) * 2021-03-15 2023-11-28 同济大学 一种噻吩骈吡啶酮类化合物、制备方法及一种抗真菌药物

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