[go: up one dir, main page]

WO2001028561A1 - Composes antibacteriens selectifs contre les bacteries gram positif, compositions contenant ces composes et techniques de traitement - Google Patents

Composes antibacteriens selectifs contre les bacteries gram positif, compositions contenant ces composes et techniques de traitement Download PDF

Info

Publication number
WO2001028561A1
WO2001028561A1 PCT/US2000/028786 US0028786W WO0128561A1 WO 2001028561 A1 WO2001028561 A1 WO 2001028561A1 US 0028786 W US0028786 W US 0028786W WO 0128561 A1 WO0128561 A1 WO 0128561A1
Authority
WO
WIPO (PCT)
Prior art keywords
aryl
alkyl
heteroaryl
groups
optionally substituted
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Ceased
Application number
PCT/US2000/028786
Other languages
English (en)
Inventor
Amjad Ali
Gayle E. Taylor
Donald W. Graham
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Merck and Co Inc
Original Assignee
Merck and Co Inc
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Merck and Co Inc filed Critical Merck and Co Inc
Priority to AU12125/01A priority Critical patent/AU1212501A/en
Publication of WO2001028561A1 publication Critical patent/WO2001028561A1/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D239/00—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings
    • C07D239/02—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings not condensed with other rings
    • C07D239/24—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings not condensed with other rings having three or more double bonds between ring members or between ring members and non-ring members
    • C07D239/28—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings not condensed with other rings having three or more double bonds between ring members or between ring members and non-ring members with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, directly attached to ring carbon atoms
    • C07D239/46—Two or more oxygen, sulphur or nitrogen atoms
    • C07D239/48—Two nitrogen atoms
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A61K31/506—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim not condensed and containing further heterocyclic rings
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D403/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00
    • C07D403/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing two hetero rings
    • C07D403/12—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing two hetero rings linked by a chain containing hetero atoms as chain links
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D405/00—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom
    • C07D405/02—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom containing two hetero rings
    • C07D405/12—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom containing two hetero rings linked by a chain containing hetero atoms as chain links
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D417/00—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00
    • C07D417/02—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00 containing two hetero rings
    • C07D417/12—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00 containing two hetero rings linked by a chain containing hetero atoms as chain links

Definitions

  • DNA polymerase III (DNA pol III) enzyme has been shown to be crucial in the replicative DNA synthesis of Gram-positive bacteria. Since DNA pol III shows little homology to mammalian or gram-negative bacterial DNA polymerases it represents an attractive target for inhibition in the discovery of new Gram-positive selective antibacterial agents.
  • the present invention relates to DNA polymerase III inhibitors which are useful against gram positive microorganisms, especially methicillin resistant Staphylococcus aureus (MRSA), methicillin resistant Staphylococcus epidermidis
  • MRSE methicillin resistant coagulase negative Staphylococci
  • MRCNS methicillin resistant coagulase negative Staphylococci
  • the compounds represent a novel class of DNA pol DI inhibitors which have advantages with respect to efficacy, toxicity and/or metabolism.
  • the antibacterial compounds of the present invention thus comprise an important contribution to therapy for treating infections caused by these difficult to control pathogens.
  • MRSA/MRCNS agents effective against such pathogens
  • R! represents H, Ci-io alkyl, C2-10 alkenyl, C2-10 alkynyl, C3-10 alicyclic, C3-10 heterocyclic, (CH2) n C5-10 aryl, (CH2) n C5-10 heteroaryl, NH(CH2) n C5-10aryl, (CH ) n diaryl, halo, CN, -O(C 5 -10 aryl), O(CH 2 ) n 2 , O(HNCO(CH 2 ) n R 2 ), S(CH 2 ) n R 2 , (CH 2 )nOR 2 , (CH 2 ) m CO 2 R 2 , (CH 2 ) n N(R2) 2 , (CH 2 ) n CON(R2) 2 ,
  • R2 represents H, C5-ioaryl or Ci-i 0 alkyl; said aryl optionally substituted with 1 to 3 groups of R a ;
  • R4 represents H or Ci-i 0 alkyl
  • R is C3-ioalicyclic, C3-ioheterocyclic, (CH2)nC5-10aryl,
  • R a represents Cl-10 alkyl, C3-8 cycloalkyl, C3-ioalicyclic, C3-ioheterocyclic, halo, C1 -6 alkyl-OH, OH, Ci-6 alkylhalo, CF3, Ci-6 alkylNH 2 , -SH, -SC 1-4 alkyl, -CN, -COC ⁇ -8 alkyl, -(CH n )nC 5-10 aryl, -COC 5- , 0 aryl, -C 5-10 heteroaryl, -COC 5 - 10 heteroaryl, CONR 6 R 7 , CONR 6 C 5 - ⁇ o aryl, CON(R 6 ) 2 , NHCOR 2 , OCOR 6 , -NR 6 (CO)R 7 , NO 2 , NR 6 (CO)NHR 7 , SO 2 R 6 , SR 6 , NHSO 2 R 6 , OR 6 , CO 2 R 6 , (CH )
  • R is H, Ci-6 alkyl, Ci-6 alkyl-OH, halo, Cj-6 alkylhalo, OR 2 , CN, CF3, N(R 2 )2, CO2R 2 , NO2, -O-, NHCOR 2 , SO2R 2 or SR 2 ;
  • R 6 & R 7 independently represent H, Ci-8 alkyl, C3-ioalicyclic, C3-i()heterocyclic, C3-8 cycloalkyl, or -C 5 - ⁇ o aryl.
  • alkyl refers to a monovalent alkane (hydrocarbon) derived radical containing from 1 to 15 carbon atoms unless otherwise defined. It may be straight or branched, saturated or unsaturated. Preferred alkyl groups include methyl, ethyl, propyl, hexyl, isopropyl, butyl and t-butyl. When substituted, alkyl groups may be substituted with up to 3 substituent groups, selected from R a as defined, at any available point of attachment. When the alkyl group is said to be substituted with an alkyl group, this is used interchangeably with "branched alkyl group”.
  • Cycloalkyl is a specie of alkyl containing from 3 to 15 carbon atoms, without alternating or resonating double bonds between carbon atoms. It may contain from 1 to 4 rings which are fused.
  • Preferred cycloalkyl groups are cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl. When substituted, cycloalkyl groups may be substituted with up to 3 substituents selected from R a as defined.
  • unsaturated alkyl refers to "alkenyl” or "alkynyl”.
  • alkenyl refers to an unsaturated alkyl such as a hydrocarbon radical, straight or branched containing from 2 to 6 carbon atoms and at least one carbon to carbon double bond.
  • Preferred alkenyl groups include propenyl, hexenyl and butenyl.
  • alkynyl refers to an unsaturated alkyl such as a hydrocarbon radical straight or branched, containing from 2 to 16 carbon atoms and at least one carbon to carbon triple bond.
  • Preferred alkynyl groups include propynyl, hexenyl and butynyl.
  • alicyclic refers to non-aromatic monocyclic or bicyclic C3- C10 hydrocarbons, including unsaturated, which can be substituted with 0-3 groups of R.
  • groups include cycloalkyls such as cyclohexyl, cyclopentyl, bicyclo[2.2.1 ]heptyl, bicyclo[2.2.1 ]hepta-2,5-dienyl, bicyclo[2.2.2]octyl, bicyclo[2.2.2]octa-2,5-dienyl.
  • heterocyclic refers to a monocyclic non-aromatic moiety containing 3-8 ring atoms or a bicyclic non-aromatic moiety containing 6-10 ring atoms, at least one of which ring atoms is a heteroatom selected from nitrogen, oxygen and sulfur and where one additional ring atom may be oxygen or sulfur.
  • heterocyclic groups are furanyl, pyranyl, morpholinyl, dioxanyl and quinuclidinyl.
  • Aryl refers to aromatic rings e.g., phenyl, substituted phenyl and the like, as well as rings which are fused, e.g., naphthyl, phenanthrenyl and the like.
  • An aryl group thus contains at least one ring having at least 6 atoms, with up to five such rings being present, containing up to 22 atoms therein, with alternating (resonating) double bonds between adjacent carbon atoms.
  • the preferred aryl groups are phenyl, naphthyl and phenanthrenyl.
  • Aryl groups may likewise be substituted as defined.
  • Preferred substituted aryls include phenyl and naphthyl.
  • heteroaryl refers to a monocyclic aromatic hydrocarbon group having 5 or 6 ring atoms, or a bicyclic aromatic group having 8 to 10 atoms, containing at least one heteroatom, O, S or N, in which a carbon or nitrogen atom is the point of attachment, and in which one or two additional carbon atoms is optionally replaced by a heteroatom selected from O or S, and in which from 1 to 3 additional carbon atoms are optionally replaced by nitrogen heteroatoms, said heteroaryl group being optionally substituted as described herein. Examples of this type are pyrrole, pyridine, oxazole, thiazole benzimidazolyl, and oxazine.
  • Additional nitrogen atoms may be present together with the first nitrogen and oxygen or sulfur, giving, e.g., thiadiazole.
  • heteroatom means O, S or N, selected on an independent basis.
  • Halogen and "halo" refer to bromine, chlorine, fluorine and iodine.
  • protecting groups for the compounds of the present invention will be recognized from the present application taking into account the level of skill in the art, and with reference to standard textbooks, such as Greene, T. W. et al. Protective Groups in Organic Synthesis Wiley, New York (1991). Examples of suitable protecting groups are contained throughout the specification.
  • protecting groups consist of groups which are used to protectively block the hydroxyl or carboxyl group during the synthesis procedures described herein. These conventional blocking groups are readily removable, i.e., they can be removed, if desired, by procedures which will not cause cleavage or other disruption of the remaining portions of the molecule. Such procedures include chemical and enzymatic hydrolysis, treatment with chemical reducing or oxidizing agents under mild conditions, treatment with a transition metal catalyst and a nucleophile and catalytic hydrogenation.
  • carboxyl protecting groups include allyl, benzhydryl, 2-naphthylmethyl, benzyl, silyl such as t-butyldimethylsilyl (TBDMS), phenacyl, p-methoxybenzyl, o-nitrobenzyl, p-methoxyphenyl, p-nitrobenzyl, 4-pyridylmethyl and t-butyl.
  • Suitable hydroxy protecting groups include triethylsilyl, t- butyldimethylsilyl, o-nitrobenzyloxycarbonyl, p-nitrobenzyloxycarbonyl, benzyloxycarbonyl, allyloxycarbonyl, t-butyloxycarbonyl, 2,2,2- trichloroethyloxycarbonyl and the like.
  • the compounds of the present invention are useful per se and in their pharmaceutically acceptable salt and ester forms for the treatment of bacterial infections in animal and human subjects.
  • pharmaceutically acceptable ester, salt or hydrate refers to those salts, esters and hydrated forms of the compounds of the present invention which would be apparent to the pharmaceutical chemist, i.e., those which are substantially non-toxic and which may favorably affect the pharmacokinetic properties of said compounds, such as palatability, absorption, distribution, metabolism and excretion.
  • Other factors, more practical in nature, which are also important in the selection are cost of the raw materials, ease of crystallization, yield, stability, solubility, hygroscopicity and flowability of the resulting bulk drug.
  • pharmaceutical compositions may be prepared from the active ingredients in combination with pharmaceutically acceptable carriers.
  • the present invention is also concerned with pharmaceutical compositions and methods of treating bacterial infections utilizing as an active ingredient the novel carbapenem compounds.
  • the pharmaceutically acceptable salts referred to above may take the form of a negative charge, which is balanced by a counterion, e.g., an alkali metal cation such as sodium or potassium.
  • a counterion e.g., an alkali metal cation such as sodium or potassium.
  • Other pharmaceutically acceptable counterions may be calcium, magnesium, zinc, ammonium, or alkylammonium cations such as tetramethylammonium, tetrabutylammonium, choline, triethylhydroammonium, meglumine, triethanolhydroammonium, etc.
  • the pharmaceutically acceptable salts referred to above also include acid addition salts.
  • the Formula I compounds can be used in the form of salts derived from inorganic or organic acids. Included among such salts are the following: acetate, adipatc, alginatc, aspartate, benzoate, benzenesulfonate, bisulfate, butyrate, citrate, camphorate, camphorsulfonate, cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate, fumarate, glucoheptanoate, glycerophosphate, hemisulfate, heptanoate, hexanoate, hydrochloride, hydrobromide, hydroiodide, 2- hydroxyethanesulfonate, lactate, maleatc, methanesulfonate, 2-naphthalenesulfonate, nicotinate, oxalate,
  • Acid addition salts of the compounds of formula I include compounds that contain a protonated, basic moiety.
  • Compounds containing a basic moiety are capable of protonation in aqueous media near pH 7, so that the basic moiety can exist as an equilibrium mixture of its neutral form and acid addition (protonated) form. The more basic the group, the greater the degree of protonation near pH 7. All such compounds are included in the present invention.
  • the pharmaceutically acceptable esters are such as would be readily apparent to a medicinal chemist, and include, for example, those described in detail in U.S. Pat. No. 4,309,438. Included within such pharmaceutically acceptable esters are those which are hydrolyzed under physiological conditions, such as pivaloyloxymethyl, acetoxymethyl, phthalidyl, indanyl and methoxymethyl, and others described in detail in U.S. Pat. No. 4,479,947. These are also referred to as "biolabile esters".
  • Biolabile esters are biologically hydrolizable, and may be suitable for oral administration, due to good absorption through the stomach or intenstinal mucosa, resistance to gastric acid degradation and other factors.
  • biolabile esters include compounds in which M represents an alkoxyalkyl, alkylcarbonyloxyalkyl, alkoxycarbonyloxyalkyl, cycloalkoxyalkyl, alkenyloxyalkyl, aryloxyalkyl, alkoxyaryl, alkylthioalkyl, cycloalkylthioalkyl, alkenylthioalkyl, arylthioalkyl or alkylthioaryl group.
  • M species are examples of biolabile ester forming moieties.: acetoxymethyl, 1-acetoxyethyl, 1-acetoxypropyl, pivaloyloxymethyl, 1 -isopropyloxycarbonyloxyethyl, 1-cyclohexyloxycarbonyl- oxyethyl, phthalidyl and (2-oxo-5-methyl-l,3-dioxolen-4-yl)methyl.
  • R a represents Ci-10 alkyl, C3-ioalicyclic, C3_ lOheterocyclic, halo, -(CH2)nC - ⁇ o aryl, -COC 5- io aryl, -(CH2)nC - ⁇ o heteroaryl, -COCs-io heteroaryl, CONR 6 C 5-!
  • aryl CON(R 6 ) 2 , OR 6 , CO 2 R 6 , COC, -8 alkyl or (CH 2 )nCO R 2 C - ⁇ 0 heteroaryl, said alkyl, aryl or heteroaryl optionally substituted with 1 to 3 groups of R.
  • composition comprised of a compound in accordance with formula I in combination with a pharmaceutically acceptable carrier.
  • Also included in this invention is a method of treating a bacterial infection comprising administering to a mammalian patient in need of such treatment a compound of formula I in an amount which is effective for treating a bacterial infection.
  • Ri and R 3 are defined with respect to the compounds of Formula I.
  • the compounds of the present invention wherein Ri is 3-methylphenoxy and R 3 is 3-iodo-4-methyl phenyl can be prepared by reacting 2- amino-4,6-dichloropyrimidine with 3-iodo-4-methyl aniline in solvents such as ethanol to afford compounds of type 1. Further reaction with 3-iodo-4-methyl aniline in the presence of cesium carbonate and dimethylformamide as the solvent affords the desired compounds of general formula 2.
  • solvents such as ethanol
  • R 1 is Cl
  • R 3 is 3,4- dimethyl phenyl
  • R 4 is ethyl
  • R 1 is Cl
  • R 3 is 3,4- dimethyl phenyl
  • R 4 is ethyl
  • 2,4,6-trichloropyrimidine with ethylamine in ethanol to afford 3.
  • Further reaction with 3,4-dimethyl aniline and caesium carbonate in DMF affords the desired compounds of general formula 4.
  • the synthesis of the target compound is completed by removing any protecting groups which are present in the penultimate intermediate using standard techniques which are well known to those skilled in the art.
  • the deprotected final product is then purified, as necessary, using standard techniques such as ion exchange chromatography, HPLC on reverse phase silica gel, MPLC on reverse phase polystyrene gel, and the like or by recrystallization.
  • the final product may be characterized structurally by standard techniques such as NMR, IR, MS, and UV.
  • the final product if not crystalline, may be lyophilized from water to afford an amorphous, easily handled solid.
  • the compounds of the present invention are valuable antibacterial agents active against various Gram-positive and to a lesser extent Gram-negative bacteria, and accordingly find utility in human and veterinary medicine. Many of compounds of the present invention are biologically active against MRSA/MRCNS. In vitro antibacterial activity is predictive of in vivo activity when the compounds are administered to a mammal infected with a susceptible bacterial organism. Using standard susceptibility tests, the compounds of the invention are determined to be active against MRSA.
  • the compounds of the invention can be formulated in pharmaceutical compositions by combining the compound with a pharmaceutically acceptable carrier. Examples of such carriers are set forth below.
  • the compounds may be employed in powder or crystalline form, in liquid solution, or in suspension. They may be administered by a variety of means; those of principal interest include: topically, orally and parenterally by injection (intravenously or intramuscularly).
  • Compositions for injection a preferred route of delivery, may be prepared in unit dosage form in ampules, or in multidose containers.
  • the injectable compositions may take such forms as suspensions, solutions, or emulsions in oily or aqueous vehicles, and may contain various formulating agents.
  • the active ingredient may be in powder (lyophillized or non- lyophillized) form for reconstitution at the time of delivery with a suitable vehicle, such as sterile water.
  • the carrier is typically comprised of sterile water, saline or another injectable liquid, e.g., peanut oil for intramuscular injections.
  • sterile water e.g., saline
  • another injectable liquid e.g., peanut oil for intramuscular injections.
  • various buffering agents, preservatives and the like can be included.
  • Topical applications may be formulated in carriers such as hydrophobic or hydrophilic bases to form ointments, creams, lotions, in aqueous, oleaginous or alcoholic liquids to form paints or in dry diluents to form powders.
  • Oral compositions may take such forms as tablets, capsules, oral suspensions and oral solutions.
  • the oral composions may utilize carriers such as conventional formulating agents, and may include sustained release properties as well as rapid delivery forms.
  • the dosage to be administered depends to a large extent upon the condition and size of the subject being treated, the route and frequency of administration, the sensitivity of the pathogen to the particular compound selected, the virulence of the infection and other factors. Such matters, however, are left to the routine discretion of the physician according to principles of treatment well known in the antibacterial arts. Another factor influencing the precise dosage regimen, apart from the nature of the infection and peculiar identity of the individual being treated, is the molecular weight of the compound.
  • the compositions for human delivery per unit dosage, whether liquid or solid, may contain from about 0.01% to as high as about 99% of active material, the preferred range being from about 10-60%.
  • composition will generally contain from about 15 mg to about 2.5 g of the active ingredient; however, in general, it is preferable to employ dosage amounts in the range of from about 250 mg to 1000 mg.
  • unit dosage will typically include the pure compound in sterile water solution or in the form of a soluble powder intended for solution, which can be adjusted to neutral pH and isotonic.
  • the invention described herein also includes a method of treating a bacterial infection in a mammal in need of such treatment comprising administering to said mammal a compound of formula I in an amount effective to treat said infection.
  • the preferred methods of administration of the formula I antibacterial compounds include oral and parenteral, e.g., i.v. infusion, i.v. bolus and i.m. injection.
  • the preferred dosage is 250 mg to 1000 mg of the antibacterial given one to four times per day. More specifically, for mild infections a dose of about 250 mg two or three times daily is recommended. For moderate infections against highly susceptible gram positive organisms a dose of about 500 mg three or four times daily is recommended. For severe, life-threatening infections against organisms at the upper limits of sensitivity to the antibiotic, a dose of about 1000-2000 mg three to four times daily may be recommended. For children, a dose of about 5-25 mg/kg of body weight given 2, 3, or 4 times per day is preferred; a dose of 10 mg/kg is typically recommended.
  • DNA polymerase in (Pol III) was cloned by PCR from S. aureus genomic DNA.
  • the Pol III gene was cloned into pET15.
  • Pol III/pET15 was transformed into BL21 cells.
  • the cells were grown at 29°C in M9ZB media containing ampicillin and chloramphenicol to an OD of 1.0.
  • the cells were induced by adding 1 mM IPTG and grown for three hours. The cells were centrifuged at 2500 x g for 10 minutes.
  • the cell pellet was resuspended in 50 mM Tris, pH 7.4, 0.1% Triton XI 00, 1 mM EDTA, 20% glycerol, and complete protease inhibitor cocktail (Boehringer Mannheim) and lysed by freeze/thawing three times in a dry ice and methanol bath. The lysate was centrifuged at 10,000 x g for one hour. The supernatant was loaded onto a HiTrapQ column that was equilibrated in 50 mM Tris, pH 7.4, 20% glycerol, and 1 mM EDTA. Following loading, the column was washed with 20 column volumes of buffer.
  • the Pol III was eluted using a linear NaCl gradient from 0-0.5 M.
  • the Pol m activity eluted between 0.3 and 0.4 M NaCl.
  • the peak of activity was loaded onto a HiTrap Blue column equilibrated in 50 mM Tris, Ph 7.4, 20% glycerol, lmM EDTA, and 1 mg/ml BSA.
  • the column was washed with 10 column volumes of equilibration buffer, followed by a wash with 20 column volumes of equilibration buffer containing 0.5 M NaCl.
  • the Pol III was eluted by washing the column with equilibration buffer containing 3 M NaCl.
  • the reaction mix consisted of 30 mM Tris HC1, pH 7.5, 20% glycerol, 4 mM DTT, 10 mM MgOAc, 0.003 mM dATP, 0.003 mM dGTP, 0.001 mM dCTP, 0.001 mM 3 H-dTTP, and 0.35 mg/ml 'activated' calf thymus DNA (Worthington Enzymes) in a final volume of 0.1 ml.
  • the assay was initiated by the addition of enzyme and incubated for 30 minutes at 30°C. The assay was stopped by the addition of cold 10%TCA/0.1% NaPPi.
  • Results are expressed as the MLC, that is, the minimum log concentration that gives inhibition of the growth of the indicated organism (ie >100, 100, 10, and ⁇ ). Any sample which was active on one or more strains was then tested in a more shallow dilution series to give a more specific measure of activity (see MIC below).
  • the range of activity for the compounds of Formulas I and II in this assay were about 10 to about 100 ⁇ M.
  • the procedures for the MIC assay were essentially as described above for the MLC assay, except that serial two-fold dilutions of the compounds were made from a final initial high concentration of 100 ⁇ M.
  • the effect of serum albumin on the activity of the compounds was examined by testing MICs against the MB2865 strain in the presence and absence of a final 4.3 mg/ml concentration of human serum fraction V (Calbiochem). Results are expressed as the minimum inhibitory concentration (MIC); the lowest two-fold dilution which gives inhibition of growth (>100, 100, 50, 25, 12.5 etc., in ⁇ M).
  • the range of activity for the compounds of Formulas I and II in this assay were about ⁇ 0.5 to about 50 ⁇ M.
  • MOA assay Compounds which were active in the MIC assay were tested for the mode of action (MOA) by whole-cell dose-response labeling. Each compound was tested versus five different labeled precursors to investigate the effect on DNA synthesis (6- 3 H Thymidine, 1 ⁇ Ci/ml final, plus uridine at 50 ⁇ g/ml final concentration), RNA synthesis (5,6- 3 H Uracil, 1 ⁇ Ci/ml final), protein synthesis (4,5-
  • test and control compounds were serially diluted either two-fold or three-fold (depending on potency) in the appropriate diluent for the compound (water or DMSO) and then 5 ⁇ l of each transferred to each of five assay plates.
  • a final assay volume of 100 ⁇ l also included precursor diluted into MH medium to obtain the final concentration indicated above and 70 ⁇ l of cells at OD 0.5. Addition of cells began the reaction allowed to proceed for 20 min. and stopped by addition of 50% TCA to a final concentration of 10%.
  • TCA precipitable counts were collected onto glass fiber filters (Wallac) using a Skatron Micro96 harvester. Filter mats were placed in bags with 10 ml scintillation fluid and loaded into cassettes for measurement in an LKB 1205 Betaplate counter (Wallac). Results are expressed as the percentage of control counts (vehicle alone, no drug) present in each sample well plotted as a function of percent of control versus concentration of compound on a semi-log plot. The concentration of compound giving 50% inhibition of each macromolecular synthesis (IC 50 ) is determined from this plot. The range of activity for the compounds of Formulas I and II in this assay were about 1.0 to about 25 ⁇ M.

Landscapes

  • Chemical & Material Sciences (AREA)
  • Organic Chemistry (AREA)
  • Health & Medical Sciences (AREA)
  • Medicinal Chemistry (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Epidemiology (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Animal Behavior & Ethology (AREA)
  • General Health & Medical Sciences (AREA)
  • Public Health (AREA)
  • Veterinary Medicine (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)

Abstract

La présente invention concerne les composés représentés par la formule (I). Cette formule comprend des sels pharmaceutiquement acceptables, des stéréo-isomères et des esters de ces éléments.
PCT/US2000/028786 1999-10-21 2000-10-18 Composes antibacteriens selectifs contre les bacteries gram positif, compositions contenant ces composes et techniques de traitement Ceased WO2001028561A1 (fr)

Priority Applications (1)

Application Number Priority Date Filing Date Title
AU12125/01A AU1212501A (en) 1999-10-21 2000-10-18 Gram-positive selective antibacterial compounds, compositions containing such compounds and methods of treatment

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US16081399P 1999-10-21 1999-10-21
US60/160,813 1999-10-21

Publications (1)

Publication Number Publication Date
WO2001028561A1 true WO2001028561A1 (fr) 2001-04-26

Family

ID=22578562

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/US2000/028786 Ceased WO2001028561A1 (fr) 1999-10-21 2000-10-18 Composes antibacteriens selectifs contre les bacteries gram positif, compositions contenant ces composes et techniques de traitement

Country Status (2)

Country Link
AU (1) AU1212501A (fr)
WO (1) WO2001028561A1 (fr)

Cited By (10)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2003062227A1 (fr) * 2002-01-23 2003-07-31 Bayer Pharmaceuticals Corporation Inhibiteurs de kinase rho
WO2003106450A1 (fr) * 2002-06-17 2003-12-24 Bayer Aktiengesellschaft Phenylaminopyrimidines et leur utilisation en tant qu'inhibiteurs de la rho-kinase
EP1406875A4 (fr) * 2001-06-26 2005-05-04 Bristol Myers Squibb Co Inhibiteurs n-heterocycliques d'expression tnf-alpha
US6924290B2 (en) 2002-01-23 2005-08-02 Bayer Pharmaceuticals Corporation Rho-kinase inhibitors
US6926763B2 (en) 2002-02-08 2005-08-09 Glsynthesis, Inc. Purine and isosteric antibacterial compounds
JP2006522143A (ja) * 2003-04-04 2006-09-28 アイアールエム・リミテッド・ライアビリティ・カンパニー プロテインキナーゼ阻害剤としての新規化合物および組成物
US7648986B2 (en) 2002-01-10 2010-01-19 Bayer Healthcare Llc Substituted thieno[3,2-D]pyrimidines as Rho kinase inhibitors
US7723347B2 (en) 2004-04-27 2010-05-25 Bayer Schering Pharma Aktiengesellschaft Substituted phenylamino-pyrimidines
WO2010038081A3 (fr) * 2008-10-03 2010-05-27 Astrazeneca Ab Dérivés hétérocycliques et procédés d’utilisation associés
US8796292B2 (en) 2009-09-11 2014-08-05 Glsynthesis Inc. Selective antibacterials for clostridium difficile infections

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US3152112A (en) * 1964-10-06 Phenylazo- and phenylhydrazo-
DE3717480A1 (de) * 1987-05-23 1988-12-01 Shell Agrar Gmbh & Co Kg Neue herbizid und mikrobizid wirksame 2,6-diaminopyrimidine

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US3152112A (en) * 1964-10-06 Phenylazo- and phenylhydrazo-
DE3717480A1 (de) * 1987-05-23 1988-12-01 Shell Agrar Gmbh & Co Kg Neue herbizid und mikrobizid wirksame 2,6-diaminopyrimidine

Non-Patent Citations (9)

* Cited by examiner, † Cited by third party
Title
CHEMICAL ABSTRACTS, vol. 105, no. 19, 10 November 1986, Columbus, Ohio, US; abstract no. 172385G, SEN ET AL.: "Synthesis of 2,6-disubstituted 9-arylpyrines, 9-aryl-8-azapyrines and related 5-phenylazopyrimidines and their biological activity" page 740; XP002937995 *
CHEMICAL ABSTRACTS, vol. 108, no. 7, 15 February 1988, Columbus, Ohio, US; abstract no. 56043G, GHONEIM ET AL.: "Synthesis and evaluation of some 2-, 4- and 2,4-disubstituted-6-methylpyrimidine derivatives for antimicrobial activity" page 724; column 1; XP002937994 *
CHEMICAL ABSTRACTS, vol. 114, no. 7, 18 February 1991, Columbus, Ohio, US; abstract no. 62037Y, GHONEIM ET AL.: "Synthesis of some Mannich bases of 2- and 4-amino and 2,4-diamino-6-methylpyrimidines as potential biodynamic agents" column 2; XP002937993 *
CHEMICAL ABSTRACTS, vol. 124, no. 3, 15 January 1996, Columbus, Ohio, US; abstract no. 29691R, KANDEEL ET AL.: "Synthesis of certain 1,2,3-selenadiazole, 1,2,3-thiadiazole and 1,2-oxazoline derivatives of anticipated antibacterial activity" page 970; column 1; XP002937992 *
EGYPT. J. CHEM. 1987, vol. 30, no. 6, 1989, (ENG), pages 295 - 304 *
INDIAN J. CHEM., SECT. B, vol. 24B, no. 9, 1985, (ENG), pages 952 - 958 *
J. INDIAN CHEM. SOC., vol. 63, no. 10, 1986, (ENG), pages 914 - 917 *
TRANTOLO ET AL.: "Inhibitors of bacillus subtilis DNA polymerase III. Influence of modifications in the pyrimidine ring of anilino- and (benzylamino)pyrimidines", J. MED. CHEM., vol. 29, no. 5, May 1986 (1986-05-01), pages 676 - 681, XP002937991 *
ZAGAZIG J. PHARM. SCI., vol. 3, no. 3B, 1994, (ENG), pages 197 - 205 *

Cited By (14)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US7253174B2 (en) 2001-06-26 2007-08-07 Bristol-Myers Squibb Company N-heterocyclic inhibitors of TNF-α expression
US8044040B2 (en) 2001-06-26 2011-10-25 Bristol-Myers Squibb Company N-heterocyclic inhibitors of TNF-alpha expression
EP1406875A4 (fr) * 2001-06-26 2005-05-04 Bristol Myers Squibb Co Inhibiteurs n-heterocycliques d'expression tnf-alpha
AU2002316421B2 (en) * 2001-06-26 2008-05-15 Bristol-Myers Squibb Company N-heterocyclic inhibitors of TNF-ALPHA expression
US7648986B2 (en) 2002-01-10 2010-01-19 Bayer Healthcare Llc Substituted thieno[3,2-D]pyrimidines as Rho kinase inhibitors
US6924290B2 (en) 2002-01-23 2005-08-02 Bayer Pharmaceuticals Corporation Rho-kinase inhibitors
US6943172B2 (en) 2002-01-23 2005-09-13 Bayer Pharmaceuticals Corporation Rho-kinase inhibitors
WO2003062227A1 (fr) * 2002-01-23 2003-07-31 Bayer Pharmaceuticals Corporation Inhibiteurs de kinase rho
US6926763B2 (en) 2002-02-08 2005-08-09 Glsynthesis, Inc. Purine and isosteric antibacterial compounds
WO2003106450A1 (fr) * 2002-06-17 2003-12-24 Bayer Aktiengesellschaft Phenylaminopyrimidines et leur utilisation en tant qu'inhibiteurs de la rho-kinase
JP2006522143A (ja) * 2003-04-04 2006-09-28 アイアールエム・リミテッド・ライアビリティ・カンパニー プロテインキナーゼ阻害剤としての新規化合物および組成物
US7723347B2 (en) 2004-04-27 2010-05-25 Bayer Schering Pharma Aktiengesellschaft Substituted phenylamino-pyrimidines
WO2010038081A3 (fr) * 2008-10-03 2010-05-27 Astrazeneca Ab Dérivés hétérocycliques et procédés d’utilisation associés
US8796292B2 (en) 2009-09-11 2014-08-05 Glsynthesis Inc. Selective antibacterials for clostridium difficile infections

Also Published As

Publication number Publication date
AU1212501A (en) 2001-04-30

Similar Documents

Publication Publication Date Title
US4978659A (en) 2-(Heteroaryliumalkyl)phenyl carbapenem antibacterial agents
CZ286878B6 (en) Carbapenems, process of their preparation, intermediates for their preparation, those carbapenems used for treating infections and pharmaceutical preparations
US9643933B2 (en) Compounds useful as antibiotic tolerance inhibitors
DD156180A5 (de) Verfahren zur herstellung von 3-amino-beta-lactam-1-sulfonsaeuren und ihren salzen
JPH0853453A (ja) ２−［１−（１，３−チアゾリン−２−イル）アゼチジン−３−イル］チオ−カルバペネム化合物
US4550102A (en) Cephem compounds
WO2001029010A1 (fr) Composes antibacteriens a selectivite gram-positif, compositions contenant ces composes et methodes therapeutiques
EP0289801B1 (fr) Dérivés d'acides (1R,5S,6S)-6-[(R)-1-hydroxy-éthyl]-1-méthyl-carbapénème-3-carboxyliques substitués en position 2 par un groupe thio substitué
GB2533136A (en) Compounds
DE69619702T2 (de) Pyrido (2,3-b) pyrazinderivate
WO2001028561A1 (fr) Composes antibacteriens selectifs contre les bacteries gram positif, compositions contenant ces composes et techniques de traitement
US5336674A (en) 2-phenanthridinyl carbaphenem antibacterial agent
KR0170046B1 (ko) 신규한 세펨 유도체
JPH10508285A (ja) カルバペネム化合物、組成物および治療方法
WO2001029045A1 (fr) Composes antibacteriens gram-positifs selectifs, compositions contenant de tels composes et methodes de traitement
AU2020400770A1 (en) Monobactam compound and use thereof
JP3527003B2 (ja) 新規イミダゾ［５，１−ｂ］チアゾール誘導体
JPS6383082A (ja) テトラヒドロフランカルボン酸誘導体の製造法
US5258509A (en) Process for producing 2-carbon-substituted carbapenem derivatives
US5328904A (en) 2-phenanthridinyl carbapenem antibacterial agents
CA1150726A (fr) Cephalosporines, procede de preparation et produit pharmaceutiques les contenant
NO153298B (no) Analogifremgangsmaate ved fremstilling av terapeutisk aktive n-methylenderivater av thienamycin
US4103011A (en) Novel penicillin compounds and process for preparation thereof
AU732358B2 (en) Carbapenems with naphthosultam derivative linked via methylene
JPH09507222A (ja) フリルチアゾールおよびそれらのｈ▲下２▼受容体拮抗剤および抗微生物剤としての用途

Legal Events

Date Code Title Description
AK Designated states

Kind code of ref document: A1

Designated state(s): AE AG AL AM AT AU AZ BA BB BG BR BY BZ CA CH CN CR CU CZ DE DK DM DZ EE ES FI GB GD GE GH GM HR HU ID IL IN IS JP KE KG KR KZ LC LK LR LS LT LU LV MA MD MG MK MN MW MX MZ NO NZ PL PT RO RU SD SE SG SI SK SL TJ TM TR TT TZ UA UG US UZ VN YU ZA ZW

AL Designated countries for regional patents

Kind code of ref document: A1

Designated state(s): GH GM KE LS MW MZ SD SL SZ TZ UG ZW AM AZ BY KG KZ MD RU TJ TM AT BE CH CY DE DK ES FI FR GB GR IE IT LU MC NL PT SE BF BJ CF CG CI CM GA GN GW ML MR NE SN TD TG

121 Ep: the epo has been informed by wipo that ep was designated in this application
DFPE Request for preliminary examination filed prior to expiration of 19th month from priority date (pct application filed before 20040101)
WA Withdrawal of international application
REG Reference to national code

Ref country code: DE

Ref legal event code: 8642

NENP Non-entry into the national phase

Ref country code: JP