WO2008024285A2 - Composé antibiotique - Google Patents

Composé antibiotique Download PDF

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Publication number
WO2008024285A2
WO2008024285A2 PCT/US2007/018288 US2007018288W WO2008024285A2 WO 2008024285 A2 WO2008024285 A2 WO 2008024285A2 US 2007018288 W US2007018288 W US 2007018288W WO 2008024285 A2 WO2008024285 A2 WO 2008024285A2
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WO
WIPO (PCT)
Prior art keywords
compound
actinoplanes
fermentation
compounds
formula
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/US2007/018288
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English (en)
Other versions
WO2008024285A3 (fr
Inventor
Angela Basilio Carnicer
Olga Genilloud
Sheo B. Singh
Francisca Vicente
Chaowei Zhang
Deborah L. Zink
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 Sharp and Dohme de Espana SA
Merck and Co Inc
Original Assignee
Merck Sharp and Dohme de Espana SA
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 Sharp and Dohme de Espana SA, Merck and Co Inc filed Critical Merck Sharp and Dohme de Espana SA
Publication of WO2008024285A2 publication Critical patent/WO2008024285A2/fr
Publication of WO2008024285A3 publication Critical patent/WO2008024285A3/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

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Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K9/00—Peptides having up to 20 amino acids, containing saccharide radicals and having a fully defined sequence; Derivatives thereof
    • C07K9/006—Peptides having up to 20 amino acids, containing saccharide radicals and having a fully defined sequence; Derivatives thereof the peptide sequence being part of a ring structure
    • C07K9/008—Peptides having up to 20 amino acids, containing saccharide radicals and having a fully defined sequence; Derivatives thereof the peptide sequence being part of a ring structure directly attached to a hetero atom of the saccharide radical, e.g. actaplanin, avoparcin, ristomycin, vancomycin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides

Definitions

  • the present invention relates to broad spectrum thiazolyl-peptide antibiotic compounds that are useful in treating bacterial infections.
  • Infections caused by bacteria are a growing medical concern as many of these bacteria are resistant to various antibiotics.
  • Such microbes include Staphylococcus aureus, Staphylococcus hemolyticus, Pediococcus spp., and Streptococcus pyogenes, Streptococcus pneumoniae, Pseudomonas aeruginosa, Vibrio cholerae, Vibrio parahemolyticus, Actinobacter calcoaeticus, Stenotrophomonas maltophilia.
  • the antibiotic of this invention thus comprises an important contribution to therapy for treating infections which are resistant to various known antibiotics.
  • thiazolyl-peptide antibiotics are produced from an Actinoplanes sp. fermentation and possess antibacterial activity against bacterial infections that are sensitive and resistance to currently available antibiotics.
  • This invention is concerned with a novel thiazolyl-peptide antibiotic of the formula I:
  • R represents:
  • the invention is also concerned with a process for the production of the 10 compounds of formula I by fermentation with a Actinoplanes sp.
  • the invention is also concerned with a process for isolating the compound of formula I from the fermentation broth. 2205 IY
  • This invention is concerned with the compounds of formula I in particular compounds of structural formula Ia:
  • the pharmaceutically acceptable salts of the compounds of this invention include the conventional non-toxic salts, as formed from non-toxic inorganic or organic bases.
  • such conventional non-toxic salts include those derived from inorganic bases such as an alkali or alkaline earth metal hydroxide, e.g., potassium, sodium, lithium, calcium, or magnesium, and the like: and the salts prepared from organic bases such as an amine, e.g., dibenzylethylene-diamine, trimethylamine, piperidine, pyrrolidine, benzylamine and the like, or a quaternary ammonium hydroxide such as tetramethylammonium hydroxide and the like.
  • an amine e.g., dibenzylethylene-diamine, trimethylamine, piperidine, pyrrolidine, benzylamine and the like
  • a quaternary ammonium hydroxide such as tetramethylammonium hydroxide and the like.
  • the pharmaceutically acceptable salts can be synthesized from the compounds of this invention by conventional chemical methods. Generally, the salts are prepared by reacting the free acid with stoichiometric amounts or with an excess of the desired salt-forming inorganic or organic base in a suitable solvent or various combinations of solvents.
  • the compounds of this invention are broad spectrum antibiotics useful in the
  • the minimum inhibitory concentration (MlC) values range from 0.0001 to less than 200 ug/mL for test strains such as Staphylococcus aureus, Staphylococcus hemolyticus, Streptococcus pyogenes, Streptococcus pneumoniae, and E. faecalis.
  • the compounds of the invention can be formulated in pharmaceutical compositions by combining the 5 compounds 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
  • compositions for injection 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
  • the carrier is typically comprised of sterile water, saline or another injectable liquid, e.g., peanut oil for intramuscular injections.
  • a suitable vehicle such as sterile water, hi injectable compositions
  • the carrier is typically comprised of sterile water, saline or 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 compositions may utilize carriers such as conventional formulating agents, and may include sustained release properties as well as rapid delivery
  • 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 Compound, the virulence of the infection and other factors. Such matters, however, are left to the routine discretion of the physician according to principles of
  • compositions for administration to humans per unit dosage may contain from about 0.001% to as high as about 99% of Compound I, one embodiment of the range being from about 10-60%.
  • the composition will generally contain from about 2 mg to about 2.5 g of Compound I, one embodiment of this range being from
  • the unit dosage will typically include pure Compound I in sterile water solution or in the form of a soluble powder intended for solution, which can be adjusted to neutral pH and isotonicity. 2205 IY
  • the invention described herein also includes a method of treating a bacterial infection in a mammal in need of such, treatment comprising the administration of the compound of formula I to the mammal in an amount effective to treat the infection.
  • One embodiment of the methods of administration of a compound of formula I 5 includes oral and parenteral methods, e.g., i.v. infusion, i.v. bolus and i.m. injection.
  • a compound of formula I per kg of body weight given one to four times daily is preferred.
  • the preferred dosage is 2 mg to 1000 mg of the antibacterial given one to four times per day. More specifically, for mild infections a dose of about 5-200 mg two or three times daily is recommended. For moderate infections against
  • a dose of about 20-1000 mg three or four times daily is recommended.
  • a dose of about 100-2000 mg three to four times daily may be recommended.
  • a dose of about 0.1-50 mg of a compound of formula I per kg of body weight given one to four times daily is typically recommended.
  • 15 aspect of this invention is realized when the dosage is 2 mg to 1000 mg of the antibacterial given one to four times per day.
  • Another aspect of this invention is the process for producing the compounds of formula I which comprises cultivating a Actinoplanes sp. microorganism (identified herein as either MA 7347/P-000503667 or ATCC PTA 7551) in a suitable nutrient medium and then
  • the compounds of structural formula I are produced by the aerobic fermentation of a suitable medium under controlled conditions via inoculation with a culture of the MA
  • the suitable medium is preferably aqueous and contains sources of assimilable carbon, nitrogen, and inorganic salts.
  • the medium employed for fermentation non-streptomyces actinomycete is primarily the well-known Difco Tryptic Soy Broth, either alone or with added nutrients, which is commonly used by those skilled in the art.
  • the fermentation is conducted at temperatures ranging from about 10 0 C to about 40 0 C; however for optimum results it is preferred to conduct the fermentation at about 26-30 0 C.
  • the pH of the nutrient medium during the fermentation can be about 5.5 to about 7.5.
  • the invention is not limited to the use of the particular Actinoplanes sp. with ATCC PTA 7551. It is especially desired and intended that there be included in the scope of this invention the use of other natural or artificial mutants produced or derived from the described cultures, or other variants or species of the Actinoplanes genus insofar as they can produce the compound of this invention.
  • the artificial production of mutant species or strains of 10 Actinoplanes from ATCC PTA 7551 may be achieved by conventional, physical or chemical mutagens, for example, ultraviolet irradiation of the described culture, or nitrosoguanidine treatment and the like. Recombinant DNA techniques such as protoplast fusion, plasmid incorporation, chromosome fragment incorporation and the like also may prove useful.
  • CAMHB cation adjusted Mueller-Hinton broth
  • DMSO dimethyl sulfoxide
  • HRFTMS high resolution fourier transform mass spectrometry
  • i.m. intramuscular
  • i.v. intravenous
  • MIC minimum inhibitory concentration
  • MOPS 3-(N-morpholino) propanesulfonic acid
  • NCCLS National Committee on Clinical Laboratory Standards
  • RPMI-1640 Roswell Park Memorial Hospital 1640 culture medium
  • ID NMR one-dimensional NMR
  • 2D NMR two-dimensional 20 NMR
  • TFA trifluoroacetic acid.
  • a first seed culture of the strain was prepared by inoculating 10 ml of -seed medium in a 50 ml tube with frozen agar plugs of the producing strain and incubating the tube at 28 0 C with shaking 10 220 rpm for about 96 hours.
  • a second seed culture was prepared by inoculating 50 mL of seed medium in a 250 mL flask with 2ml of the first seed.
  • the IL broth was extracted with IL Acetone by shaking for 2 hours. 50% Acetone extract was filtered in a sintered glass funnel packed with celite. The filtrate was 20 concentrated to 15% acetone in H2O, which was subjected to Amberchrome chromatography
  • Compound I was determined by the use of mass spectroscopy, lH NMR and 13C NMR.
  • the complete 16S rDNA sequence was determined from the 1500 bp PCR fragment obtained using primers 27f and 1525r (Lane, 1991, Nucleic acid techniques in bacterial systematics. E. Stackebrandt and M. Goodfellow (Eds). The PCR product was used as template
  • PA UP Parsimony Analysis program version 4.0. (Swofford D. L. (1993). PA UP: Phylogenetic analysis using parsimony, version 3.1.1. Washington D.C.: Laboratory of Molecular Systematics, Smithsonian Institute).
  • the strain was obtained from a soil collected in Namaqualand, South Africa and associated to the rhizosphere of a specimen of Dinteranthus microspermus. The strain was isolated after high 2205 IY
  • centrifiigation speed enrichment method and plating on NZ amine A based agar supplemented with 20 ug/ml nalidixic acid.
  • the strain grows well on a range of agar media such as Yeast Malt Extract, Oatmeal, Glycerol Asparagine, Inorganic Salts Starch and Tyrosine agars at 28 0 C.
  • agar media such as Yeast Malt Extract, Oatmeal, Glycerol Asparagine, Inorganic Salts Starch and Tyrosine agars at 28 0 C.
  • the gross colonial morphology is typical of Micromonosporaceae and its growth characteristics, sporulation structures, substrate mycelial pigmentation and the production of different pigments were recorded in different agar media (Table 1).
  • strain P-000503667 contains meso-diamino pimelic acid in whole-cell hydrolysates, a characteristic of Micromonosporaceae, and galactose as major cell wall sugar.
  • the strain cultivated in Trypticase Soy Agar (TSBA) for 13 days at 28°C is rich in saturated and 25 unsaturated fatty acids and whole-cell methanoly sates contain the predominant fatty acids 15:0 iso (9.78%),15:0 (10.61%), 16:0 iso (22.76%), 16:1 cis9 (6.71%) and 17:1 cis9 (30.84%).
  • TSBA Trypticase Soy Agar
  • Strain P-000503667 presents the following carbon utilization pattern: good utilization of D-glucose " , D-mannitol, rhamnose, L-arabinose, D-xylose, D-fructose, I- inositol and sucrose; no utilization of cellulose and raf ⁇ inose.
  • strain P-000503667 belongs to the genus Actinoplanes.
  • the general description, both colony morphology and micro morphology is also consistent with the genus Actinoplanes. Based on the data presented herein, P-000503667 should be classified and identified as a species in the genus Actinoplanes.
  • strain P-000503667 is associated in the phylogenetic tree to the strain Actinoplanes philippinensis DSM 43019. This close relationship is highly supported by the
  • Table 2 Major fatty acids found in strain P-000503667.
  • HTM Haemophilus Test Medium
  • TLB Trypticase Soy Broth
  • Microbank Beads (Kramer Scientific) 30 MIC 2000 Microtiter plate inoculator.
  • BBL Cation- Adjusted Mueller Hinton Broth
  • Haemophilus Test Medium Received prepared from manufacturer. Filter-sterilized 20 before use using a Corning 0.45 Tm cellulose acetate filter.
  • ESCHERICHI CATION-ADJUSTED MUELLER HINTON (CAMHB; BBL);
  • PNEUMONIAE INCUBATION CONDITIONS, 35°C; MICS READ AT 22-24
  • INFLUENZAE HOURS
  • the strains used are isolates from either the Merck Culture Collection, the Merck Clinical Culture Collection or from Clinical Trials.
  • the strain of Haemophilus influenzae is a mouse pathogen used for in vivo testing at Merck.
  • the Escherichia coli strain is a cell wall permeable strain.
  • the Candida albicans strain is used as a control. These culture are maintained as frozen stocks at -8O 0 C in a) Microbank beads; b) 2X Skim Milk; or c) in 2X Trypticase Soy Broth + 15% glycerol/50% horse serum (Haemophilus and Streptococcus pneumoniae).
  • Selected isolates are sub-cultured onto either Chocolate Agar Plates (Haemophilus influenzae), onto Trypticase Soy + 5% Sheep Blood Agar Plates (Streptococcus pneumoniae, Staphylococcus aureus, Escherichia coli, Enterococcus, Bacillus) or onto Sabouraud Dextrose
  • Colonies are selected from plates and used to prepare an inoculum equivalent to a 0.5 McFarland standard in Trypticase Soy Broth.
  • An inoculum with a density equivalent to a 1.0 5 McFarland standard is prepared for Streptococcus pneumoniae.
  • the inoculum density for all cultures is ⁇ 10 8 CFU/mL in TSB.
  • This TSB inoculum is diluted 1 : 10 in sterile saline (4 mL inoculum + 36 mL saline; equivalent to ⁇ 10 7 CFU/mL) and kept on ice until used to inoculate microtiter plates.
  • Haemophilus test media plates are prepared to test Haemophilus influenzae; Cation- Adjusted Mueller Hinton + 5% Lysed Horse Blood plates are prepared to test Streptococcus pneumoniae; Cation-Adjusted Mueller Hinton Broth plates are prepared to test Enterococcus, Staphylococcus aureus, Escherichia coli and Bacillus subtilis.
  • RPMI 1640 is used to test Candida. The MICs against S. aureus Smith are determined in Cation-adjusted Mueller Hinton and in Cation-
  • Adjusted Mueller Hinton + 50% Human Serum to determine if the compound is inactivated by some component in serum (indicated by an increase in the MIC). Filled plates are wrapped in plastic bags (to minimize evaporation), stored frozen and thawed before use.
  • DMSO fetal sulfate
  • the last row contains a growth control of 3.2% DMSO. With each assay, controls are run. They are Penicillin G and chloramphenicol, prepared in the same manner as the compounds. Ertapenem is included as a control for the serum protein binding assay.
  • microliter plates are inoculated with (saline-diluted) culture using the MIC 2000 System, an automated plate inoculating device which delivers an inoculum of 1.5 mL per well. Plates are incubated at 35 0 C in ambient air. An uninoculated plate is also incubated as a sterility check. Results are recorded after 22-24-hours' incubation. Plates were read to no 10 growth. The MIC is defined as the lowest antimicrobial level which resulted in no growth after 22-24-hours' incubation.
  • Compound I demonstrates antibacterial activity against various strains of S. aureus, E. faecalis, E. faecium, B. subtilus and S. pneumoniae.
  • Compounds Ia and Ib also demonstrate antibacterial activity against various strains of S. aureus, E. faecalis, E. faecium, B. subtilus and S. pneumoniae.
  • Compounds Ia and Ib also demonstrate antibacterial activity against various strains of S. aureus, E. faecalis, E. faecium, B. subtilus and S. pneumoniae.
  • Compounds Ia and Ib also demonstrate antibacterial activity against various strains of S. aureus, E. faecalis, E. faecium, B. subtilus and S. pneumoniae.
  • Compounds Ia and Ib also demonstrate antibacterial activity against various strains of S. aureus, E. faecalis, E. faecium, B
  • MRSA methicillin-resistant S. aureus
  • VRE vancomycin-resistant Enterococcus sp.
  • MIC minimum inhibitory concentration

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  • Chemical & Material Sciences (AREA)
  • Organic Chemistry (AREA)
  • Genetics & Genomics (AREA)
  • Biochemistry (AREA)
  • Biophysics (AREA)
  • General Health & Medical Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Medicinal Chemistry (AREA)
  • Molecular Biology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
  • Preparation Of Compounds By Using Micro-Organisms (AREA)

Abstract

L'invention concerne un nouveau composé antibiotique à spectre large de formule structurale I : (I) ou un de ses sels, esters, énantiomères, diastéréoisomères ou mélanges pharmaceutiquement acceptables, ledit composé étant obtenu par fermentation d'un milieu nutritif avec un Actinoplanes sp.
PCT/US2007/018288 2006-08-23 2007-08-17 Composé antibiotique Ceased WO2008024285A2 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US83949706P 2006-08-23 2006-08-23
US60/839,497 2006-08-23

Publications (2)

Publication Number Publication Date
WO2008024285A2 true WO2008024285A2 (fr) 2008-02-28
WO2008024285A3 WO2008024285A3 (fr) 2008-10-09

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PCT/US2007/018288 Ceased WO2008024285A2 (fr) 2006-08-23 2007-08-17 Composé antibiotique

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Family Cites Families (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20020055465A1 (en) * 2000-08-14 2002-05-09 Wenying Li Nocathiacin antibiotics prepared by biotransformation or chemical methods

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