Siderophore cephalosporin conjugate and its use

Abstract

Compounds and uses thereof according to a compound of formula (I), wherein X is selected from the group consisting of CH, CCl and N, Z is selected from the group consisting of CH2COOH, CH(CH3)COOH, C(CH3)2COOH and CH2F, D is a single bond connecting A and Ar or is selected from the group consisting of CO, NHCO and N(C 0‑6 )alkyl-CO, A is selected from (C 1‑6 )alkanediyl and (C 3‑6 )cycloalkanediyl, or, if D is N(C0)alkyl-CO, A forms a 4- to 7-membered aliphatic heterocyclic ring with the nitrogen atom of N(C0)alkyl-CO in D, and Ar is a 6-membered aromatic ring having a first hydroxyl group at the para position of D, a second hydroxyl group at the meta position of D, and at least one electron withdrawing element. #imgabs0#

Description

[0001] The present invention relates to organic compounds, namely, siderophore cephalosporin conjugates and uses thereof.

[0002] Cephalosporins are a class of β-lactam antibiotics with a common core structure, the cephem ring system 8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid. The bacterial spectrum (i.e. the bactericidal activity of cephalosporins against different bacteria) varies according to the substitution pattern. Cephalosporin compounds containing a CH=N moiety in the side chain at position 3 of the cephem ring are described, for example, in WO 96 / 35692, wherein oxime-type CH=NO compounds are exemplified in Examples 3, 4 and 6.

[0003] Siderophores are iron chelating compounds, and siderophore drug conjugates exploit the iron uptake machinery in bacteria. Some examples of siderophore drug conjugates are summarized, for example, in Ji, C. et al., Future Med. Chem. (2012) 4(3), 297-313.

[0004] Siderophore conjugated cephalosporins are described, for example, in WO2010 / 050468, WO2011 / 136268, WO2012 / 134184 and WO2012 / 147773, which cover derivatives with alkyl or alkenyl groups and quaternary ammonium groups in the side chain at position 3 of the cephem ring. In, for example, WO2010 / 050468, cefiderocol is exemplified, which is a catechol-type siderophore cephalosporin conjugate that represents the most advanced siderophore drug conjugate and is currently in the preregistration stage for potential intravenous treatment of multidrug resistant (MDR) Gram-negative bacterial infections.

[0005]

[0006] Strong iron chelating properties are important for siderophore drug conjugates to be able to compete with natural siderophores secreted by microorganisms. Summary of the invention

[0007] The present invention provides compounds according to formula (I):

[0008]

[0009] in

[0010] X is selected from the group consisting of CH, CCl and N,

[0011] Z is selected from the group consisting of CH2COOH, CH(CH3)COOH, C(CH3)2COOH and CH2F,

[0012] D is a single bond connecting A and Ar or is selected from CO, NHCO and N(C0-6 ) alkyl-CO group,

[0013] A Select from the group consisting of

[0014] -(C 1-6 )alkanediyl, and

[0015] -(C 3-6 )cycloalkanediyl, or

[0016] If D is N(C0)alkyl-CO, A forms a 4- to 7-membered aliphatic heterocyclic ring with the nitrogen atom of N(C0)alkyl-CO in D, and

[0017] Ar is a 6-membered aromatic ring having a first hydroxyl group at a para position relative to D, a second hydroxyl group at a meta position relative to D, and having at least one electron withdrawing element.

[0018] In the compounds according to the invention, the siderophore group (Ar) is linked to the cephalosporin nucleus via a bond comprising an oxime ether at position 3 of the cephem ring. The resulting oxime-type siderophore conjugate cephalosporins have not been described before. The inventors surprisingly found that oxime-type siderophore conjugate cephalosporins, i.e. compounds according to formula (I), have interesting antibacterial activity and unexpectedly strong iron chelating properties. The oxime bond of the siderophore group (Ar) is associated with the surprisingly strong iron chelating properties. Example 4 differs from the compound cefiderocol mainly in the linker, which shows an iron chelating activity that is one order of magnitude higher than that measured for cefiderocol (see Table 1 below).

[0019] In the compound according to formula (I), Ar is considered to be primarily responsible for the iron chelating properties of the compound. Ar is a 6-membered aromatic ring with two hydroxyl groups. Two hydroxyl groups are mutually ortho-substituted (catechol type), which is known to allow iron atom complexation (i.e. iron chelation). The first hydroxyl group occupies a position (i.e. para position) relative to the position connected to the rest of the compound. The second hydroxyl group occupies a position on a carbon atom adjacent to the carbon atom to which the first hydroxyl group is combined (i.e., meta position of the position connected to the rest of the compound).

[0020] In the compounds according to the invention, the siderophore group Ar also has at least one electron-withdrawing element.

[0021] Electron withdrawal is a concept known in organic chemistry and describes the polarization or electrostatic forces relative to the reaction center or, in the present case, relative to the electron system in the aromatic ring. The electron withdrawing element can be, for example, an electron withdrawing substituent on the aromatic ring. If D is selected from CO, NHCO and N(C 0-6 )alkyl-CO, the aromatic ring Ar is substituted with a carbonyl group known to have an electron-withdrawing effect. However, according to the present invention, Ar has at least one additional electron-withdrawing element.

[0022] Other electron-withdrawing elements can be substituents located at any free position of the aromatic ring. Electron-withdrawing substituents include, for example, halogen, nitrile, carbonyl (particularly C1-C6 carbonyl, such as formyl, acetyl) or haloalkoxy.

[0023] In a preferred embodiment, the electron-withdrawing substituent is a substituent selected from the group consisting of F, Cl and OCF3. In particular, the electron-withdrawing substituent is a substituent in a free position of the aromatic ring, preferably in an ortho position to D.

[0024] Alternatively or additionally, the electron withdrawing element may be a heteroatom in the aromatic ring Ar.

[0025] In a preferred embodiment, the electron withdrawing element is a nitrogen atom or a substituted nitrogen atom in the aromatic ring Ar. Thus, the resulting aromatic ring Ar is a pyridine-based aromatic ring. For example, the aromatic ring can be selected from pyridine, N-(C 1-6 )alkyl-pyridinium and pyridine-N-oxide. Preferably, the pyridine-based aromatic ring is substituted with two hydroxyl groups at the 3rd and 4th positions (or the 4th and 5th positions) relative to the nitrogen atom and is attached to D (or the rest of the molecule) at the 2nd position (or the 6th position) relative to the nitrogen atom. Illustratively, Ar is selected from the group consisting of 3,4-dihydroxy-2-pyridyl, 4,5-dihydroxy-2-pyridyl, 3,4-dihydroxy-1-(C1-C6)alkyl-2-pyridinium, 4,5-dihydroxy-1-(C1-C6)alkyl-2-pyridinium, 1,3-dihydroxy-4-oxo-2-pyridyl and 1,5-dihydroxy-4-oxo-2-pyridyl, preferably 4,5-dihydroxy-2-pyridyl, 4,5-dihydroxy-1-(C1-C6)alkyl-2-pyridinium and 1,5-dihydroxy-4-oxo-2-pyridyl.

[0026] In another preferred embodiment, the compound according to the invention is a compound according to formula (II)

[0027]

[0028] wherein X, Z, D and A are as defined above, and

[0029] in

[0030] E is selected from CH, N, N + – CH3 and N + –O - The group composed of

[0031] G is H or an electron-withdrawing substituent,

[0032] Provided that if E is CH, then D is selected from CO, NHCO and N(C 0-6)alkyl-CO and G is an electron withdrawing substituent.

[0033] In these embodiments, it is ensured that if E is CH, i.e. if the aromatic ring Ar is a phenyl-based ring, the substituents G and D are selected to contain electron withdrawing. This means that if E is CH, G is an electron withdrawing substituent and D includes a carbonyl functionality.

[0034] In this case, preferably, G is selected from the group consisting of F, Cl and OCF3.

[0035] In one embodiment, the linker element D is selected from the group consisting of CO, NHCO and N(C 0-6 )alkyl-CO, more preferably selected from the group consisting of NHCO and N(C 0-6 )alkyl-CO, and in particular the group consisting of NHCO and N(C0)alkyl-CO. If D is N(C0)alkyl-CO, A forms a 4- to 7-membered aliphatic heterocyclic ring with the nitrogen atom of N(C0)alkyl-CO in D. In this case, A forms two substituents of the nitrogen and no further alkyl substituents are present, i.e., it is defined as (C0)alkyl.

[0036] The linker element A (together with D) affects the geometric arrangement (e.g., distance, direction) of Ar relative to the cephem ring. Typically, A is selected independently of D. However, in some embodiments, the definition of A is related to D. Based on preliminary data, it is believed that the O of the oxime group is separated from the nearest atom of Ar by at least one atom (e.g., in Example 1), but preferably no more than six atoms (e.g., five atoms) is beneficial, especially for iron chelating activity. The distance can be described by the number of covalent bonds in the shortest connection path within the molecule (or within a portion of the molecule). This is a very simplified representation that ignores conformational / three-dimensional aspects, but allows for the definition of some preferences for the linker according to the present invention. Those of ordinary skill in the art will appreciate that covalent bonds including heteroatoms, involving sp 2 Covalent (CC) bonds of hybridized C atoms as well as cyclic (CC) bonds (due to geometric constraints within the ring system) are shorter than acyclic covalent (CC) single bonds.

[0037] In this context, the shortest connection path in A is defined by the number of bonds extending between O and D of the oxime group, excluding the bond to O of the oxime group (i.e., excluding the bond OA in formula (I) or (II)) and excluding the bond to D or the single bond of D (i.e., excluding the bond AD in formula (I) or (II)). Starting from the term "shortest", in the case of a forked or cyclic linker, the path with the fewest bonds should be considered.

[0038] In one embodiment, A is selected such that the shortest connection path in A is shorter than or equal to 3 covalent (CC) single bonds, in particular shorter than 3 acyclic covalent (CC) single bonds, and preferably shorter than or equal to 2 covalent (CC) single bonds, in particular shorter than 2 acyclic covalent (CC) single bonds.

[0039] In a preferred embodiment, the shortest connection path in A extends through zero single (CC) bonds, one cyclic or acyclic (CC) single bond, or two cyclic covalent (CC) bonds.

[0040] Thus, in a preferred embodiment, A is selected from the group consisting of methanediyl, ethane-1,2-diyl, propane-1,3-diyl, propane-1,2-diyl, cyclobutane-1,3-diyl, cyclopentane-1,3-diyl, cyclohexane-1,3-diyl, azetidine-1,3-diyl, pyrrolidine-1,3-diyl and piperidine-1,3-diyl, wherein in azetidine-1,3-diyl, pyrrolidine-1,3-diyl and piperidine-1,3-diyl, D is N(C0)alkyl-CO and the nitrogen atom of N(C0)alkyl-CO in D is a nitrogen in a heterocyclic ring,

[0041] Preferably selected from the group consisting of methanediyl, ethane-1,2-diyl, cyclobutane-1,3-diyl and pyrrolidine-1,3-diyl, wherein D is N(C0)alkyl-CO and the nitrogen atom of N(C0)alkyl-CO in D is the nitrogen in the pyrrolidine ring.

[0042] In another embodiment, the compound according to the invention is a compound according to formula (II),

[0043] wherein X and Z are defined as above, and

[0044] D is NHCO or N(C 0-6 )alkyl-CO,

[0045] A is

[0046] -(C 1-6 )alkanediyl, preferably (C 1-4 )alkanediyl, for example in particular (C2)alkanediylethane-1,2-diyl,

[0047] -(C 3-6 )cycloalkanediyl, preferably (C 4-5 )cycloalkanediyl, for example in particular (C4)cycloalkanediylcyclobutane-1,3-diyl, or

[0048] A and the nitrogen atom of N(C0)alkyl-CO in D form a 4- to 7-membered aliphatic heterocyclic ring, preferably a 5-membered aliphatic heterocyclic ring, such as, in particular, a 5-membered heterocyclic pyrrolidine-1,3-diyl, wherein the nitrogen atom of N(C0)alkyl-CO in D is the nitrogen in pyrrolidine, E is CH,

[0049] G is Cl.

[0050] Alternatively, if E is N, N + – CH3 and N + -O, i.e. if the aromatic ring Ar is a pyridyl-based ring, no such requirement is included for G and D. The heteroatom N is an electron withdrawing element in Ar. Thus, G may also be hydrogen and / or D may be a single bond (i.e. D does not exist as a separate moiety).

[0051] Preferably, in this case, the compound according to the invention is a compound according to formula (II),

[0052] wherein X and Z are defined as above, and

[0053] Where D is a single bond,

[0054] A is

[0055] -(C 1-6 )alkanediyl, preferably (C 1-4 ) alkanediyl, for example methanediyl,

[0056] -(C 3-6 )cycloalkanediyl, or

[0057] E is for N + –O - ,

[0058] G is H.

[0059] In another embodiment, preferably, —AD—Ar in formula (I) is selected from the group consisting of:

[0060]

[0061] Preferably selected from the group consisting of

[0062]

[0063] More preferably selected from the group consisting of:

[0064]

[0065] The compounds of the present invention include a thiazole ring (X is CH or X is CCl) or 1,3,4-thiadiazole (X is N).

[0066] In a preferred embodiment, X is CH or N.

[0067] The oxime in the 7-position substitution is O-substituted with a substituted alkyl group Z selected from the group consisting of CH2COOH, CH(CH3)COOH, C(CH3)2COOH and CH2F.

[0068] In a preferred embodiment, Z is C(CH3)2COOH or CH2F.

[0069] In another aspect, the present invention provides a composition comprising a compound according to the present invention and at least one pharmaceutically acceptable excipient.

[0070] In one embodiment, the composition optionally further comprises at least one pharmaceutically active agent.

[0071] In another aspect, the present invention provides a compound or composition according to the present invention for use as a medicament.

[0072] The compound or composition according to the present invention can be used in a method for treating or preventing a disease, wherein the compound according to the present invention or a composition comprising the compound is administered to a subject in need thereof.

[0073] It has been found that the compounds according to the invention have bactericidal activity. They are therefore suitable for the treatment and prevention of infectious diseases.

[0074] Thus, in one embodiment, the present invention provides a compound or composition according to the invention for use in the treatment and prevention of infectious diseases mediated by microorganisms or bacteria, preferably Gram-negative bacteria.

[0075] In a preferred embodiment, the infectious disease is selected from the group consisting of:

[0076] - Respiratory infections of the upper and / or lower respiratory tract, including cystic fibrosis and bronchiectasis,

[0077] - Urinary tract infection,

[0078] - Intra-abdominal infection,

[0079] - Systemic infection,

[0080] - Prosthetic joint infection,

[0081] - Gastrointestinal infections, and

[0082] - Skin and / or soft tissue infection.

[0083] In another embodiment, the infectious disease is mediated by a bacterium selected from the group consisting of Escherichia coli, Salmonella typhimurium, Citrobacter freundii, Klebsiella pneumoniae, Enterobacter cloacae, Enterobacter aerogenes, Moraxella catarrhalis, Proteus mirabilis, Proteus vulgaris, Providencia stuartii, Serratia marcescens, Acinetobacter baumannii, Acinetobacter lwoffi, Pseudomonas aeruginosa, aeruginosa), Haemophilus influenzae, Burkholderia cepacia, Burkholderia cenocepatia, Burkholderia vietnamensis, Streptococcus spp. and Streptococcus pneumoniae, preferably selected from the group consisting of Escherichia coli, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa and Burkholderia cepacia.

[0084] In another aspect, the present invention provides a method for treating an infectious disease in a subject in need thereof, comprising administering to the subject an effective amount of a compound according to formula (I), wherein in the compound according to formula (I), X, Z, A, D and Ar are defined as shown above.

[0085] In one embodiment, the compound is administered in the form of a pharmaceutical composition. Treatment includes treatment and prevention. For antimicrobial and cystic fibrosis treatment, a suitable dosage (effective amount) will of course vary according to, for example, the chemical properties and pharmacokinetic data of the compounds of the invention used, individual hosts, modes of administration, and the nature and severity of the disease being treated. However, in general, in order to obtain satisfactory results in large mammals such as humans, the indicated daily dosage is within the range of about 0.5 mg to 3 g of the compounds of the invention, which is conveniently administered, for example, in divided doses up to four times a day. DETAILED DESCRIPTION

[0086] According to the compound of formula (I) or (II), (C X-Y The definition of a ) group includes any group having from X to Y carbon atoms. For example, (C 1-5 (C1-C6)alkyl or (C1-C6)alkanediyl include, for example, methyl, ethyl, propyl, prop-2-yl, butyl, but-2-yl, trimethylmethyl, 2-methylprop-1-yl, pentyl, pent-2-yl, pent-3-yl, 2-methylbutyl, 3-methylbutyl, 3-methylbut-2-yl. Similarly, (C1-C6)alkyl or (C1-C6)alkanediyl also include straight chain and branched alkane groups.

[0087] Any group defined as part of a compound according to the invention may be unsubstituted or substituted, for example mono- or poly-.

[0088] (C1-C6)alkyl, (C1-C6)alkanediyl, cycloalkyl, (C1-C6)alkyl 3-6 )cycloalkanediyl, 4 to 7 membered aliphatic heterocyclic ring and 6 membered aromatic ring include unsubstituted or substituted (C1-C6)alkyl, (C1-C6)alkanediyl, cycloalkyl, (C 3-6 )cycloalkanediyl, 4 to 7 membered aliphatic heterocyclic ring and 6 membered aromatic ring. These groups may be substituted by conventional groups in organic chemistry, for example. Substituents include, for example, (C1-C6) alkyl, (C2-C6) alkenyl, (C1-C6) alkoxy, hydroxy, oxo, carboxyl, (C1-C6) alkylcarbonyl, amido, urea, guanidinyl, thiourea, amino, halogen.

[0089] Group A and D (if D is not a single bond) are linking groups with two free valences for forming a key with the other parts of the compound. If not otherwise specified, linking group A or D are arranged from left to right in formula (I) or (II). For example, if D in formula (I) is NHCO, the nitrogen-atom of amides is connected to A, and the carbon atom is connected to Ar.

[0090] The compound according to the present invention has two oxime ether functional groups, also referred to as oxygen-imine functional groups. For these functional groups, different stereochemical orientations are possible based on the E / Z configuration at the double bond (C=N bond) with the nitrogen atom. It should be understood that the diagram of the direction in the formula should not exclude alternative configurations. Therefore, in the compound according to formula (I) or (II), if stereochemistry is not clearly indicated (i.e., in the case of a chiral carbon atom on the cephem ring), all configuration isomers should be understood to be included. In particular, for two oxime functional groups independent of each other, both configurations are included in formula (I) or (II), so four diastereoisomeric forms are also covered. However, preferably, each oxime functional group is independently in a configuration visualized by a diagram of formula (I) or (II), more preferably both are in a configuration visualized by a diagram of formula (I) or (II).

[0091] Pyridine-N-oxides, pyridones or pyridiniums may exist in tautomeric forms:

[0092]

[0093] The present invention includes all tautomeric forms regardless of the tautomer chosen for the diagram or nomenclature used for the corresponding formula.

[0094] The compounds of the present invention include compounds in any form, such as in free form / inner salt / zwitterion form, in salt form, in solvate form, or in salt and solvate form. Salts preferably include pharmaceutically acceptable salts, although any salt is included, such as for preparation / isolation / purification purposes.

[0095] Methods for preparing the compounds of the invention are known or can be prepared according to (eg analogously to) conventional methods or the methods specified in the following Synthesis Scheme 1:

[0096] Solution 1

[0097]

[0098] In the compounds of formula (I), formula (A) and formula (D), X, Z, A, D and Ar are as defined above.

[0099] In the preparation of the compounds of the invention, the intermediates or starting materials (A), (B) and (D), optionally in protected form or in salt form or both, are known or can be prepared according to (eg analogously to) conventional methods.

[0100] The intermediates or starting materials of formula (B) and (C) contain an aldehyde which, together with the proximal carboxylic acid, may exist as a hemi-acylal:

[0101]

[0102] The compounds of the invention may be used (or employed in a method) for treatment or prophylaxis (ie prevention) wherein the compounds of the invention are applied to a subject, ie administered.

[0103] The compounds of the invention can be administered by any conventional route, for example enterally, for example including nasal, buccal, oral administration; parenterally, for example including intravenous, intramuscular, subcutaneous administration; or topically, for example including inhalation, epidermal, intranasal, intratracheal, intraaural, ophthalmic administration, for example in the form of coated or uncoated tablets, capsules, injectable solutions or suspensions, for example in the form of ampoules, vials, in the form of creams, gels, pastes, inhalation liquids, inhalation powders, sprays, foams, tinctures or drops.

[0104] Preferably, the compounds according to the invention are not administered in pure form but formulated as compositions.

[0105] Therefore, in another aspect, the present invention provides a pharmaceutical composition comprising a compound of the present invention and at least one pharmaceutical excipient, such as a carrier or a diluent, wherein the compound according to the present invention may be in free form or in a pharmaceutically acceptable salt form and / or solvate form.

[0106] Depending on the route of administration, the composition is formulated according to the prior art of pharmaceutical galenics. A person skilled in the art may select at least one pharmaceutical excipient from known excipients, including, for example, fillers, binders, disintegrants, flow regulators, lubricants, sugars and sweeteners, spices, preservatives, stabilizers, wetting agents and / or emulsifiers, solubilizers, salts for regulating the osmotic pressure and / or buffers.

[0107] The compounds of the present invention can be used alone or in combination with one or more other pharmaceutically active agents for (or in a method for) treatment or prevention. Such other pharmaceutically active agents include, for example, β-lactamase inhibitors, potentiators and other antibiotics, and if the compounds of the present invention are used to treat cystic fibrosis, other pharmaceutically active agents also include agents commonly used to treat cystic fibrosis.

[0108] The other pharmaceutically active agent may be provided in a composition comprising a compound according to the invention, or the other pharmaceutically active agent may be administered alone.

[0109] The uses and methods of the invention are primarily directed to the treatment and prevention of human subjects, but may also be used for the treatment and prevention of other mammalian subjects, i.e. animals and pets such as poultry, pigs, cattle, dogs, cats, i.e. for veterinary purposes and for diluting liquids for artificial insemination and egg dipping techniques.

[0110] The compounds of the present invention can be used for (or used in methods for) treating or preventing infectious diseases. Infectious diseases can be diagnosed by those skilled in the art, who associate the symptoms of a subject with a microbial or bacterial infection, or the infectious disease can be diagnosed by detecting the microorganism or bacteria in a subject or in a sample taken from the subject.

[0111] Compounds according to the present invention show pharmacological activity, and therefore can be used as medicines. For example, compounds of the present invention show antibacterial activity against: Gram-negative bacteria, such as Escherichia coli, Salmonella typhimurium, Citrobacter freundii, Klebsiella pneumoniae, Enterobacter cloacae, Enterobacter aerogenes, Moraxella catarrhalis, Proteus mirabilis, Proteus vulgaris, Providencia stuartii, Serratia marcescens, Acinetobacter baumannii, Acinetobacter lofeti, Pseudomonas aeruginosa, Haemophilus influenzae, Burkholderia cepacia, Burkholderia cenocepacia, Burkholderia vietnamiensis, and Gram-positive bacteria, such as Streptococcus and Streptococcus pneumoniae.

[0112] Infectious diseases mediated by Gram-negative bacteria can, for example, be mediated by a bacterium selected from the group consisting of: Escherichia coli, Salmonella typhimurium, Citrobacter freundii, Klebsiella pneumoniae, Enterobacter cloacae, Enterobacter aerogenes, Moraxella catarrhalis, Proteus mirabilis, Proteus vulgaris, Providencia stuartii, Serratia marcescens, Acinetobacter baumannii, Acinetobacter lofy, Pseudomonas aeruginosa, Haemophilus influenzae, Burkholderia cepacia, Burkholderia cenocepacia, and Burkholderia vietnam.

[0113] Therefore, the compounds according to the present invention are suitable for treating and preventing diseases mediated by microorganisms (e.g., bacteria). Based on the antibacterial activity to gram-positive and gram-negative bacteria, the compounds according to the present invention are suitable for treating and preventing infectious diseases. Diseases that can be treated with the compounds according to the present invention include, for example, upper and lower respiratory tract infections, including cystic fibrosis and bronchiectasis, as well as urinary tract infections, intra-abdominal infections, systemic infections, prosthetic joint infections, gastrointestinal infections, and soft tissue infections.

[0114] According to the Clinical and Laboratory Standards Institute CLSI file (antimicrobial susceptibility test performance standard) M100Ed29E (2019) and (dilution method for antimicrobial susceptibility test of aerobic growth bacteria) M07Ed11 (2018), the in vitro activity against bacteria was determined by standard broth microdilution. The data of the compound with iron chelating activity were obtained using cation-regulated MuellerHinton broth medium (CAMHB) and additional iron-depleted CAMHB medium (ID-CAMHB). The results of embodiments 1 to 8 and comparative compounds are summarized in Table 1.

[0115] The iron chelating properties were determined by the chromeazurol assay according to Schwyn B. et al., Universal Chemical Assay for the Detection and Determination of Siderophores, 1987. Analytical Biochemistry 160: 47-56. The assay uses the affinity of chromeazurol S / iron (III) / hexadecyltrimethylammonium bromide for iron (III). When a strong chelating agent removes iron from the dye, its color changes from blue to orange, and the absorbance measured at 630 nm indicates the chelating properties of the siderophore. The results of Examples 1 to 8 and comparative compounds are summarized in Table 1.

[0116] Thus, in one embodiment, the compounds according to the invention are 50 The defined iron chelating activity is ≤ 40 μM, preferably < 20 μM, wherein the iron chelating activity is determined by the chrome azuro assay.

[0117] Table 1

[0118]

[0119]

[0120] Abbreviations in Table 1:

[0121] CI 95 LL-UL 95% confidence interval, from lower limit to upper limit

[0122] EC Escherichia coli

[0123] KP Klebsiella pneumoniae

[0124] AB Acinetobacter baumannii

[0125] PA Pseudomonas aeruginosa

[0126] BC Burkholderia cepacia

[0127] CF Cystic Fibrosis

[0128] nd Not determined

[0129] Example

[0130]

[0066] Throughout this document, including in the Examples and Reaction Schemes, the following abbreviations are used:

[0131] 1 H-NMR Proton Nuclear Magnetic Resonance Spectroscopy

[0132] ℃ Celsius

[0133] BOC tert-Butyloxycarbonyl

[0134] BSA N,O-Bis(trimethylsilyl)-acetamide

[0135] DMSO Dimethyl sulfoxide

[0136] EDC N-(3-Dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride

[0137] eq equivalent

[0138] h hour

[0139] HOBT 1-Hydroxybenzotriazole

[0140] HPLC High Performance Liquid Chromatography

[0141] MS

[0142] N Normality

[0143] NMR Nuclear Magnetic Resonance Spectroscopy

[0144] m / e mass-to-charge ratio

[0145] THF Tetrahydrofuran

[0146] Example 1: (6R, 7R)-7-[[(2Z)-2-(2-aminothiazol-4-yl)-2-(1-carboxy-1-methyl-ethoxy)imino-acetyl]amino]-3-[(1,5-dihydroxy-4-oxo-2-pyridyl)methoxyiminomethyl]-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid

[0147] Solution 2

[0148]

[0149] Step 1: Amide formation

[0150] Compound A1 (US4500526) (3.8 g, 1 eq) in acetonitrile (20 mL) suspension was added to an ice-cooled solution of compound B (WO95 / 29182) (1.47 g, 6.44 mmol) and BSA (3.15 mL, 2 eq) in acetonitrile (10 mL). The resulting reaction mixture was stirred for 1 hour under ice cooling, stirred at room temperature for 1 hour, and then slowly added to ice water. The resulting precipitate was filtered and dried under reduced pressure to obtain compound C1 as a yellow solid, which was directly used in the next step.

[0151] 1H-NMR (400 MHz, DMSO-d6, δ, ppm): 9.46 (d, 1H, CONH, J = 8.2 Hz), 7.43-7.18 (m, 15H, trityl-H), 6.71 (s, 1H, thiazole-H), 6.30, 6.22 (2xs, 1H, O-CH-O), 5.90 (dd, 1H, H-6, J = 5.2 Hz, 8.2 Hz), 5.16 (d, 1H, H-7, J = 5.2 Hz), 3.76 (d, 1H, H-2a, J = 18.6 Hz), 3.58 (d, 1H, H-2b, J = 18.6 Hz), 1.41 (s, 6H, 2xCH3), 1.37 (s, 9H, COO t Bu).

[0152] MS m / e:782[M+H]

[0153] Step 2: Oxime formation

[0154] To a suspension of compound C1 (Example 1, step 1) and compound D1 (EP0430286) (1.11 g, 1 eq) in acetonitrile (15 mL) was added 1N hydrochloric acid (19.3 mL, 3 eq) under ice cooling. The resulting mixture was warmed to room temperature, stirred for 3 hours, and then slowly added to ice water. The resulting precipitate was filtered out and dried under reduced pressure to obtain compound I1 in a protected form as a solid, which was used directly in the next step.

[0155] 1 H-NMR (400MHz, DMSO-d6, δ, ppm): 9.41 (d, 1H, CONH, J = 8.1Hz),), 8.36 (s, 1H, azomethine), 7.80 (s, 1H, dihydropyridine H-6), 7.38-7.17 (m, 15H, trityl-H), 6.74 (s, 1H, dihydropyridine H-3), 6.68 (s, 1H, thiazole-H), 5 .79(dd,1H,H-6,J=5.0Hz,8.1Hz),5.23(d,1H,H-7,J=5.0Hz),5.15(s,2H,O-CH2),3.85(d ,1H,H-2a,J=18.0Hz),3.58(d,1H,H-2b,J=18.0Hz),1.38(bs,6H.2xCH3),1.35(s,9H,COO t Bu).

[0156] MS m / e:936[M+H] +

[0157] Step 3: Deprotect

[0158] The protected form of compound I1 (Example 1, step 2) was dissolved in trifluoroacetic acid (20 mL) under ice cooling. The resulting mixture was stirred for 4 hours under ice cooling and then slowly added to cold ether. The resulting precipitate was filtered out and dried under reduced pressure to obtain compound I1 as a solid in the form of a trifluoroacetate salt.

[0159] 1 H-NMR (400 MHz, MeOH-d4, δ, ppm): 8.46 (s, 1H, azomethine), 7.84 (s, 1H, dihydropyridine H-6), 6.94 (s, 1H, thiazole-H), 6.84 (s, 1H, dihydropyridine H-3), 5.87 (d, 1H, H-6, J = 5.1 Hz), 5.20 (s, 2H, O-CH2), 5.16 (d, 1H, H-7, J = 5.1 Hz), 3.89 (d, 1H, H-2a, J = 18.2 Hz), 3.50 (d, 1H, H-2b, J = 18.2 Hz), 1.52 (s, 3H, CH3), 1.51 (s, 3H, CH3).

[0160] MS m / e:638[M+H] +

[0161] Step 4a (optional): Salt exchange and chromatography

[0162] Compound I1 (1.6 g, 1.85 mmol) in the form of trifluoroacetate (Example 1, step 3) was suspended in water and 1N NaOH was slowly added until pH 6 to 6.5 was reached. 1N HCl was added to the resulting solution until pH 2 was reached, and the resulting precipitate was filtered and subjected to reverse phase chromatography to give Compound I1 (530 mg) as an off-white solid in the form of the hydrochloride after freeze drying.

[0163] 1 H-NMR (400 MHz, MeOH-d4, δ, ppm): 8.45 (s, 1H, azomethine), 7.77 (s, 1H, dihydropyridine H-6), 6.81 (s, 1H, thiazole-H), 6.77 (s, 1H, dihydropyridine H-3), 5.87 (d, 1H, H-6, J = 5.0), 5.18 (s, 2H, O-CH2), 5.14 (d, 1H, H-7, J = 5.0 Hz), 3.86 (d, 1H, H-2a, J = 18.0 Hz), 3.48 (d, 1H, H-2b, J = 18.0 Hz), 1.52 (s, 3H, CH3), 1.50 (s, 3H, CH3).

[0164] MS m / e:638[M+H] +

[0165] Compounds of formula (III) wherein R, X and Z are as defined in Table 2 (Examples 2 to 8) are prepared according to (e.g. analogously to) conventional methods or as described above in Example 1 steps 2 to 3 and optional step 4, but using appropriate starting materials.

[0166]

[0167] Table 2

[0168]

[0169]

[0170]

[0171]

[0172] Suitable starting materials C2 and C3 (WO98 / 43981) are prepared according to (eg analogously to) conventional methods or according to (eg analogously to) the method described in step 1 of example 1

[0173]

[0174] Suitable starting materials D2, D3 and D4 are prepared according to (eg analogously to) conventional methods or as described below:

[0175] Intermediate D2

[0176] Solution 3

[0177]

[0178] HOBT (316 mg, 1 eq), EDC (446 mg, 1 eq) and O-(2-aminoethyl)hydroxylamine hydrochloride (348 mg, 1 eq) were added to a solution of 2-chloro-3,4-bis[(4-methoxyphenyl)methoxy]benzoic acid (EP 0 416 410) (1 g, 2.33 mmol) in dichloromethane and stirred at room temperature for 1 hour. Triethylamine (976 μL, 3 eq.) was then added and the resulting reaction mixture was stirred at room temperature for another 2 hours. The resulting reaction mixture was concentrated to dryness and the residue was purified by chromatography (ethyl acetate / methanol=10 / 1) to obtain compound D2 (1.92 g) as a white solid.

[0179] 1H-NMR(400MHz,MeOH-d4,δ,ppm):7.41(m,2H,arom.),7.30(m,2H,arom.),7.22(d,1H,arom.,J=8.5Hz),7.12(d,1H,arom.,J=8.5Hz),6.96(m,2H,arom.) ,6.82(m,2H,arom.),5.11(s,2H,OCH2),4.94(s,2H,OCH2),3.83(s,3H,OCH3 ), 3.80 (t, NOCH2, J = 5.3Hz), 3.79 (s, 3H, OCH3), 3.58 (t, 2H, NCH2, J = 5.3Hz).

[0180] MS m / e:487[M+H] + ,531[M+HCOO - ]

[0181] Intermediate D3

[0182] Solution 4

[0183]

[0184] Step a

[0185] Potassium carbonate (1.55 g, 2.5 eq) and cis-tert-butyl N-(3-iodocyclobutyl)carbamate (1.99 g, 1.5 eq) were added to a DMSO solution of N-hydroxy-phthalimide (730 mg, 4.48 mmol). The resulting reaction mixture was stirred at 80 ° C for 3 hours and then slowly added to ice water. The resulting precipitate was filtered, dried under reduced pressure and purified by chromatography (dichloromethane / methanol=100 / 1) to give trans-tert-butyl N-[3-(1,3-dioxoisoindolin-2-yl)oxycyclobutyl]carbamate (290 mg) as a white solid.

[0186] MS m / e:350[M+NH4 + ]

[0187] Step b

[0188] To a solution of trans-tert-butyl N-[3-(1,3-dioxoisoindolin-2-yl)oxycyclobutyl]carbamate (2.26 g, 6.80 mmol) in dichloromethane was added hydrogen chloride (2M in diethyl ether, 12 mL) and stirred overnight at room temperature. The resulting reaction mixture was concentrated to dryness to give trans-2-(3-aminocyclobutyloxy)isoindoline-1,3-dione (1.84 g wet) as a light brown solid.

[0189] MS m / e:233[M+H] +

[0190] Step c

[0191] To a solution of 2-chloro-3,4-bis[(4-methoxyphenyl)methoxy]benzoic acid (EP 0 416 410) (3.40 g, 7.92 mmol) in dichloromethane was added HOBT (1.07 g, 1 eq), EDC (1.52 g, 1 eq) and trans-2-(3-aminocyclobutyloxy)isoindoline-1,3-dione (1.84 g wet, 6.80 mmol) as a light brown solid and stirred at room temperature for 1 hour. Then triethylamine (3.29 mL, 3 eq.) was added and the resulting reaction mixture was stirred at room temperature overnight. The obtained reaction mixture was concentrated to dryness, and the residue was purified by chromatography (dichloromethane / methanol / ammonia=100 / 5 / 0.5) to give trans-2-chloro-N-[3-(1,3-dioxoisoindolin-2-yl)oxycyclobutyl]-3,4-bis[(4-methoxyphenyl)methoxy]benzamide (2.49 g) as a pale yellow solid.

[0192] Step d

[0193] To a solution of trans-2-chloro-N-[3-(1,3-dioxoisoindolin-2-yl)oxycyclobutyl]-3,4-bis[(4-methoxyphenyl)methoxy]benzamide (1.99 g, 3.09 mmol) in acetonitrile was added hydrazine hydrate (64-65% hydrazine, 0.30 mL, 2 eq) and stirred at room temperature for 4 hours. The resulting reaction mixture was filtered. The filtrate was diluted with aqueous ammonium solution and extracted twice with ethyl acetate. The organic phases were combined, dried over Na2SO4 and concentrated to give Compound D3 (0.98 g) as a white solid.

[0194] MS m / e:513[M+H] +

[0195] Intermediate D4

[0196] Solution 5

[0197]

[0198] Step a

[0199] 2-Chloro-3,4-bis[(4-methoxyphenyl)methoxy]benzoic acid (EP 0 416 410) (4.53 g, 1 eq), EDC (2.02 g, 1 eq) and HOBT (1.43 g, 1 eq) were added to a solution of (3R)-hydroxypyrrolidine (919 mg, 10.56 mmol) in dichloromethane and stirred overnight at room temperature. The resulting reaction mixture was diluted with dichloromethane and washed with water. The resulting organic phase was concentrated to dryness and purified by chromatography (dichloromethane / methanol / ammonia=100 / 5 / 0.5) to give [2-chloro-3,4-bis[(4-methoxyphenyl)methoxy]phenyl]-[(3R)-3-hydroxypyrrolidin-1-yl]methanone (3.93 g).

[0200] MS m / e:498[M+H] +

[0201] Step b

[0202] Triethylamine (1.59 mL, 1.5 eq) and methanesulfonyl chloride (0.71 mL, 1.2 eq) were added to a THF solution of [2-chloro-3,4-bis[(4-methoxyphenyl)methoxy]phenyl]-[(3R)-3-hydroxypyrrolidin-1-yl]methanone (3.77 g, 7.57 mmol) under ice cooling and stirred at room temperature for 2 days. The resulting reaction mixture was concentrated, distributed between ethyl acetate and water, and the resulting organic phase was washed with a saturated aqueous NaCl solution. The organic phase was separated again, dried over Na2SO4, and evaporated to dryness to give a crude product [(3R)-1-[2-chloro-3,4-bis[(4-methoxyphenyl)methoxy]benzoyl]pyrrolidin-3-yl]methanesulfonate (4.36 g wet).

[0203] MS m / e:573[M+H] +

[0204] Step c

[0205] Potassium carbonate (2.30 g, 2.5 eq) and N-hydroxy-phthalimide (2.00 g, 1 eq) were added to a DMSO solution of crude [(3R)-1-[2-chloro-3,4-bis[(4-methoxyphenyl)methoxy]benzoyl]pyrrolidin-3-yl]methanesulfonate (3.83 g, 6.65 mmol) and stirred at 80°C for 3 hours. The reaction mixture was partitioned between ethyl acetate and water, and the resulting organic phase was washed with a saturated aqueous NaCl solution. The organic phase was separated again, dried over Na2SO4 and concentrated to dryness to give crude 2-[(3S)-1-[2-chloro-3,4-bis[(4-methoxyphenyl)methoxy]benzoyl]pyrrolidin-3-yl]oxyisoindoline-1,3-dione (2.73 g wet).

[0206] MS m / e:643[M+H] +

[0207] Step d

[0208] To a solution of crude 2-[(3S)-1-[2-chloro-3,4-bis[(4-methoxyphenyl)methoxy]benzoyl]pyrrolidin-3-yl]oxyisoindoline-1,3-dione (2.96 g, 4.60 mmol) in acetonitrile (20 mL) was added hydrazine hydrate (64-65% hydrazine, 0.45 mL, 2 eq) and stirred at room temperature for 1 hour. The resulting reaction mixture was diluted with aqueous ammonium solution and extracted twice with ethyl acetate. The organic phases were combined, dried over Na2SO4 and concentrated to give crude [(3S)-3-aminooxypyrrolidin-1-yl]-[2-chloro-3,4-bis[(4-methoxyphenyl)methoxy]phenyl]methanone (1.83 g wet).

[0209] MS m / e:513[M+H] +

[0210] Step e

[0211] To a solution of crude [(3S)-3-aminooxypyrrolidin-1-yl]-[2-chloro-3,4-bis[(4-methoxyphenyl)methoxy]phenyl]methanone (1.01 g, 1.97 mmol) in dichloromethane (10 mL) was added trifluoroacetic acid (5 mL) and stirred at room temperature for 1 hour. The resulting reaction mixture was concentrated and hydrogen chloride (2M in diethyl ether, 1 eq) was added and evaporated to dryness again to give crude (3S)-3-aminooxypyrrolidin-1-yl]-(2-chloro-3,4-dihydroxy-phenyl)methanone hydrochloride (1.36 g).

[0212] MS m / e:273[MH] +

Claims

1. Compounds according to formula (II) in X is selected from the group consisting of CH, CCl and N, Z is selected from the group consisting of CH2COOH, CH(CH3)COOH, C(CH3)2COOH and CH2F, D is a single bond or D is selected from CO, NHCO and N(C 0-6 ) alkyl-CO group, A is -(C 1-6 )alkanediyl, -(C 3-6 )cycloalkanediyl, or If D is N(C0)alkyl-CO, A and the nitrogen atom of N(C0)alkyl-CO in D form a 4- to 7-membered aliphatic heterocyclic ring, E is selected from CH, N, N + – CH3 and N + –O - The group composed of G is H or an electron-withdrawing substituent, Provided that if E is CH, then D is selected from CO, NHCO and N(C 0-6 ) alkyl-CO and G is an electron withdrawing substituent, wherein the electron withdrawing substituent is selected from the group consisting of F, Cl and OCF3.

2. The compound according to claim 1, wherein the shortest connection path in A is defined by the number of bonds extending between O and D of the oxime group, excluding the bond to O of the oxime group and excluding the bond to D or a single bond to D, and A is selected such that the shortest connection path in A is shorter than or equal to 3 covalent (CC) single bonds.

3. The compound according to claim 1, wherein the shortest connection path in A is defined by the number of bonds extending between O and D of the oxime group, excluding the bond to O of the oxime group and excluding the bond to D or a single bond to D, and A is selected such that the shortest connection path in A is shorter than three acyclic covalent (CC) single bonds.

4. The compound according to claim 1, wherein the shortest connection path in A is defined by the number of bonds extending between O and D of the oxime group, excluding the bond to O of the oxime group and excluding the bond to D or a single bond to D, and A is selected such that the shortest connection path in A is shorter than or equal to 2 covalent (CC) single bonds.

5. The compound of claim 2, wherein the shortest connection path in A extends with zero single (CC) bonds, one cyclic or acyclic (CC) single bond, or two cyclic covalent (CC) bonds.

6. The compound according to claim 1, wherein A is selected from the group consisting of methanediyl, ethane-1,2-diyl, propane-1,3-diyl, propane-1,2-diyl, cyclobutane-1,3-diyl, cyclopentane-1,3-diyl, cyclohexane-1,3-diyl, azetidine-1,3-diyl, pyrrolidine-1,3-diyl and piperidine-1,3-diyl, wherein in azetidine-1,3-diyl, pyrrolidine-1,3-diyl and piperidine-1,3-diyl, D is N(C0)alkyl-CO and the nitrogen atom of N(C0)alkyl-CO in D is a nitrogen in a heterocyclic ring.

7. The compound according to claim 1, wherein D is NHCO or N(C 0-6 )alkyl-CO, A is -(C 1-6 )alkanediyl, -(C 3-6 )cycloalkanediyl, or A and the nitrogen atom of the N(C0)alkyl-CO in D form a 4- to 7-membered aliphatic heterocyclic ring, E is CH, and G is Cl.

8. The compound according to claim 1, wherein D is a single bond, A is -(C 1-6 )alkanediyl, or -(C 3-6 )cycloalkanediyl, E is for N + –O - ,and G is H.

9. The compound according to claim 1, wherein Select from the group consisting of 10. The compound according to claim 1, wherein Selected from the group consisting of:

11. The compound of claim 1, wherein X is CH or N.

12. The compound of claim 1, wherein Z is C(CH3)2COOH or CH2F.

13. The compound according to claim 1, selected from the group consisting of: (6R,7R)-7-[[(2Z)-2-(2-aminothiazol-4-yl)-2-(1-carboxy-1-methyl-ethoxy)imino-acetyl]amino]-3-[(1,5-dihydroxy-4-oxo-2-pyridyl)methoxyiminomethyl]-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid, (6R,7R)-7-[[(2Z)-2-(5-amino-1,2,4-thiadiazol-3-yl)-2-(1-carboxy-1-methyl-ethoxy)imino-acetyl]amino]-3-[(1,5-dihydroxy-4-oxo-2-pyridyl)methoxyiminomethyl]-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid, (6R,7R)-7-[[(2Z)-2-(5-amino-1,2,4-thiadiazol-3-yl)-2-(fluoromethoxyimino)acetyl]amino]-3- [(1,5-dihydroxy-4-oxo-2-pyridyl)methoxyiminomethyl]-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid, 7-[[(2Z)-2-(2-aminothiazol-4-yl)-2-(1-carboxy-1-methyl-ethoxy)imino-acetyl]amino]-3-[2-[(2-chloro-3,4-dihydroxy-benzoyl)amino]ethoxyiminomethyl]-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid, 7-[[(2Z)-2-(5-amino-1,2,4-thiadiazol-3-yl)-2-(1-carboxy-1-methyl-ethoxy)imino-acetyl]amino]-3-[2-[(2-chloro-3,4-dihydroxy-benzoyl)amino]ethoxyiminomethyl]-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid, 7-[[(2Z)-2-(5-amino-1,2,4-thiadiazol-3-yl)-2-(fluoromethoxyimino)-acetyl]amino]-3-[2-[(2-chloro-3,4-dihydroxy-benzoyl)amino]ethoxyiminomethyl]-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid, 7-[[(2Z)-2-(2-aminothiazol-4-yl)-2-(1-carboxy-1-methyl-ethoxy)imino-acetyl]amino]-3-[[3-[(2-chloro-3,4-dihydroxy-benzoyl)amino]-trans-cyclobutyloxy]iminomethyl]-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid, and 7-[[(2Z)-2-(2-Aminothiazol-4-yl)-2-(1-carboxy-1-methyl-ethoxy)imino-acetyl]amino]-3-{[(3S)-1-(2-chloro-3,4-dihydroxy-benzoyl)pyrrolidin-3-yl]oxyiminomethyl}-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid.

14. A composition comprising a compound according to any one of claims 1 to 13 and at least one pharmaceutically acceptable excipient.

15. The composition of claim 14, further comprising at least one pharmaceutically active agent.

16. Use of a compound according to any one of claims 1 to 13 or a composition according to claim 14 or 15 in the preparation of a medicament for the treatment and prevention of infectious diseases mediated by bacteria, wherein the bacteria is selected from Escherichia coli, Salmonella typhimurium, Citrobacter freundii, Klebsiella pneumoniae, Enterobacter cloacae, Enterobacter aerogenes, Moraxella catarrhalis, Proteus mirabilis, Proteus vulgaris, Providencia stuartii, Serratia marcescens, marcescens, Acinetobacter baumannii, Acinetobacter lwoffi, Pseudomonas aeruginosa, Haemophilus influenzae, Burkholderia cepacia, Burkholderia cenocepatia, Burkholderia vietnamensis, Streptococcus spp., and Streptococcus pneumoniae.

17. Use of a compound according to any one of claims 1 to 13 or a composition according to claim 14 or 15 in the preparation of a medicament for treating and preventing infectious diseases mediated by Gram-negative bacteria.

18. The use according to claim 16, wherein the infectious disease is selected from the group consisting of respiratory infections of the lower and / or upper respiratory tract, urinary tract infections, intra-abdominal infections, systemic infections, prosthetic joint infections, gastrointestinal infections and skin and / or soft tissue infections, wherein upper and lower respiratory tract infections include cystic fibrosis and bronchiectasis.

19. The use according to claim 16, wherein the infectious disease is mediated by bacteria selected from the group consisting of Escherichia coli, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa and Burkholderia cepacia.

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