Bacterial efflux pump inhibitors and methods of use

Abstract

Disclosed herein are bacterial efflux inhibitor compounds and methods of using the compounds to treat bacterial infections.

Description

[Technical field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims priority to U.S. Provisional Patent Application No. 63 / 228,541, filed August 2, 2021. The contents of the aforementioned application are incorporated herein by reference in their entirety.

[0002] government support

[0002] This invention was made with Government support under Grant NIH R33 AI121365 awarded by the National Institutes of Health. The Government has certain rights in the invention. [Background technology]

[0003] background

[0003] The rise of antibiotic-resistant bacteria has emphasized the need for novel antibacterial drugs. Historical approaches to antibiotic discovery have yielded many important treatments, but recent attempts to identify new drugs have lagged far behind the spread of resistance. In the golden age of antibiotic discovery in the 1940s and 1950s, actinomycete extracts were screened for growth inhibition of pathogenic bacteria. This empirical platform identified the major classes of antibiotics in use today. This broth-based strategy yielded inhibitors that targeted core growth processes: translation, DNA replication, and cell wall synthesis. Although highly effective, targeting essential processes leads to strong selection for resistance. To focus the search for antibacterial compounds on targets that are less likely to lead to resistance, the field has shifted to screening for pathogenicity-specific processes, aided by the advent of genomics and the concomitant identification of pathogenicity-associated targets. Pharmaceutical companies have invested in high-throughput screening of synthetic chemical libraries for inhibitory activity against validated molecular targets in biochemical assays. Over the past 30 years, target-based approaches have failed to yield any antibiotics for systemic use due to a combination of few identified hits through screening and a widespread lack of antibacterial activity across all bacteria.

[0004]

[0004] The discrepancy between biochemical inhibition and antibacterial activity is due to insufficient intracellular accumulation of small molecules in bacteria. Gram-negative bacteria, in particular, contain a cell membrane, a cell wall, and an outer membrane. This cell envelope limits the penetration of amphipathic and hydrophilic substances into the cytoplasm, which is a major challenge for antibiotics.

[0005]

[0005] Bacteria also use efflux pumps as a mechanism to defend against antibiotics. Efflux pumps span the periplasm between the inner and outer membranes and capture antibiotics and host antimicrobial peptides (AMPs) and excrete them in an energy-dependent manner. When bacteria are faced with toxic molecules such as antibiotics, they respond by expressing higher levels of efflux pumps. Efflux pumps capture and excrete antibiotics, and most antimicrobial resistant (AMR) clinical isolates acquire additional copies of efflux pumps and / or express them at high levels. Thus, bacterial efflux pumps are a major contributor to the increased antibiotic resistance of Gam-negative bacteria.

[0006]

[0006] Furthermore, bacteria that live within host cells (e.g., Salmonella enterica, Listeria monocytogenes, Staphylococcus aureus, Mycobacterium tuberculosis) are further protected by the host cell membrane; some pathogens that live within vesicles are also shielded by the phagosomal membrane. Thus, even conventional antibiotics that are useful against extracellular pathogens are ineffective against intracellular microorganisms. For example, aminoglycosides and β-lactams accumulate poorly within host cells and are usually ineffective. Fluoroquinolones are primarily localized in the cytosol of host cells and therefore are weakly active against pathogens within the phagosome. Macrolides, although highly concentrated within cells, are usually ineffective against vesicular microorganisms due to inactivation at low pH of the phagolysosome, as biochemical approaches essentially ignore cell permeability during initial screening. Thus, poor cell permeability represents a significant pitfall for antibacterial agents targeting pathogenicity.

[0007]

[0007] The present disclosure addresses these needs. Summary of the Invention [Means for solving the problem]

[0008] overview In an embodiment, the present disclosure provides a compound of formula (I): [ka] (In the formula: n is 1, 2, or 3; p is 1, 2, 3, 4, or 5; m is 1, 2, 3, 4, or 5; Each R 1 is independently halo, alkyl, or haloalkyl; R 2 is -H, alkyl, alkenyl, or alkynyl; R 3 is -H, alkyl, alkenyl, or alkynyl; R 4 is alkyl, alkenyl, alkynyl, cycloalkyl, or heterocyclyl, each of which optionally has one or more R 5 or R 3 and R 4 taken together form a heterocyclyl, which optionally has one or more R 5 Replaced with; R 5 is aryl, heteroaryl, alkyl, NH2, NHR A , or NR A R B or alkyl-NH; R A is alkyl, alkenyl, or alkynyl, each of which is optionally substituted with -OH or alkoxy; and R B is alkyl, alkenyl, or alkynyl) or a stereoisomer or a pharma- ceutically acceptable salt thereof.

[0009] In an embodiment of formula (I), R 3 is -H or alkyl. In an embodiment of formula (I), R 4 is one R 5 alkyl or heterocyclyl substituted with; and R 5 is NH2. In embodiments, the compound of formula (I) has the following structure: [ka] or a pharma- ceutically acceptable salt thereof.

[0010] In an embodiment of formula (I), R 3 and R 4 taken together form a heterocyclyl, which optionally has one or more R 5In an embodiment, the compound of formula (I) is substituted with formula (II): [ka] (In the formula: n is 1, 2, or 3; p is 1, 2, 3, 4, or 5; m is 1, 2, 3, 4, or 5; q is 1, 2, or 3; Each R 1 is independently halo, alkyl, or haloalkyl; R 2 is hydrogen, C 1-6 Alkyl, C 1-6 Alkenyl, or C 1-6 is alkynyl; The A ring is heterocyclyl; R 5 is aryl, heteroaryl, alky, NH2, NHR A , or NR A R B and; R A is C 1-6 Alkyl, C 1-6 Alkenyl, or C 1-6 alkynyl, each of which is optionally —OH or C 1-6 substituted with alkoxy; and R B is C 1-6 Alkyl, C 1-6 Alkenyl, or C 1-6 alkynyl, each of which is optionally —OH or C 1-6 substituted with alkoxy) or a stereoisomer or a pharma- ceutically acceptable salt thereof.

[0011] In an embodiment of formula (II), the A ring is a 5-8 membered heterocyclyl, optionally having 1, 2, or 3 heteroatoms selected from N, O, and S in addition to the ring N shown in formula (II). 5Heteroaryl, NH2, NHR A , or NR A R B ;R A Optionally, C 1-6 Alkoxy-substituted C 1-6 alkyl; and R B is C 1-6 It is an alkyl.

[0012] In an embodiment of formula (I) or formula (II), the compound has formula (III): [ka] (In the formula: n is 1 or 2; p is 1, 2, or 3; m is 1, 2, or 3; q is 1; Each R 1 is independently halo or haloalkyl; The A ring is a 5- or 6-membered heterocyclyl; R 5 Heteroaryl, C substituted with -NH 1-6 Alkyl, -NH2, -NHR A , or -NR A R B and; R A Optionally, C 1-6 Alkoxy-substituted C 1-6 is alkyl; and R B is C 1-6 (It is alkyl) or a stereoisomer or a pharma- ceutically acceptable salt thereof.

[0013] In embodiments of formula (I), (II), or (III), the compound has formula (III-1) or (III-2): [ka] (In the formula, n, p, m, q, R 1 , ring A, and R 5 is as defined above) or a stereoisomer or a pharma- ceutically acceptable salt thereof.

[0014] In an embodiment, n is 1 or 2. In an embodiment, n is 2.

[0015] In an embodiment, p is 1, 2, or 3. In an embodiment, p is 1.

[0016] In an embodiment, m is 1, 2, or 3. In an embodiment, m is 1.

[0017] In an embodiment, q is 1.

[0018] In an embodiment, each R 1 is independently halo or haloalkyl. In embodiments, each R 1 is independently halo. In embodiments, n is 2 and each R 1 is independently halo. In embodiments, each R 1 In embodiments, n is 2 and each R 1 is -Cl.

[0019] In embodiments, the A ring is a 5- or 6-membered heterocyclyl.

[0020]

[0020] R 5 is heteroaryl, C 1-6 Alkyl-NH2, -NH2, -NHR A , or -NR A R B In an embodiment, R A Optionally, C 1-6 Alkoxy-substituted C 1-6 In embodiments, R B is C 1-6 It is an alkyl.

[0021] In embodiments of formula (I), (II), or (III), the compound has formula (IV): [ka] (In the formula: n is 1 or 2; q is 1; Each R 1 is Cl or fluoroalkyl; R 5 is alkyl substituted with -NH2, or -NH2, -NHR A , or NR A R B and; R A is C 1-6 Alkyl, C 1-6 Alkyl is optionally C 1-6 substituted with alkoxy; R B is C 1-6 (It is alkyl) or a stereoisomer or a pharma- ceutically acceptable salt thereof.

[0022] In an embodiment, the compound of formula (I), (II), (III), or (IV) has formula (IV-1) or (IV-2): [ka] (In the formula, n, q, R 1 , and R 5 is as defined above) or a stereoisomer or a pharma- ceutically acceptable salt thereof.

[0023] In an embodiment, R 5 is alkyl substituted with -NH2, or -NH2. In an embodiment, the compound of formula (IV) has the following structure: [ka] or a stereoisomer or a pharma- ceutically acceptable salt thereof. In an embodiment, the compound of formula (IV) has the following structure: [ka] or a pharma- ceutically acceptable salt thereof.

[0024] In embodiments of formula (I), (II), or (III), the compound has formula (V): [ka] (In the formula: n is 1 or 2; q is 1; Each R 1 is Cl or fluoroalkyl; R 5 Heteroaryl, NH2, NHR A , or NR A R B and; R A Optionally, C 1-6 Alkoxy-substituted C 1-6 is alkyl; R B is C 1-6 (It is alkyl) or a stereoisomer or a pharma- ceutically acceptable salt thereof.

[0025] In an embodiment of formula (V), R 5 is a 5-7 membered heteroaryl having 1, 2, or 3 heteroatoms selected from N and S. In an embodiment, R 5 is a 5-membered heteroaryl having 1 or 2 N heteroatoms. In embodiments, R 5 is imidazolyl. In embodiments, the compound of formula (V) has the following structure: [ka] or a pharma- ceutically acceptable salt thereof.

[0026]

[0026] In an embodiment, the present disclosure provides a pharmaceutical composition comprising a therapeutically effective amount of a compound of formula (I), (II), (III), (IV), or (V) and one or more pharma- ceutically acceptable excipients.

[0027] In an embodiment, the present disclosure provides a method of treating a bacterial infection in a subject in need thereof, the method comprising administering to the subject a pharma- ceutically acceptable amount of a compound of formula (I), (II), (III), (IV), or (V). In an embodiment, the bacterial infection is caused by an intracellular pathogen. In an embodiment, the bacterial infection is caused by a gram-negative bacterium. In embodiments, the bacterial infection is one or more of Salmonella sp., Acinetobacter sp., Actinobacillus sp., Aeromonas sp., Bacteroide sp., Bordetella sp., Brucella sp., Burkholderia sp., Prevotella sp., Porphyromonas sp., Campylobacter sp., Citrobacter sp., Edwarsiella sp., Eikenella sp., Enterobacter sp., Escherichia sp., sp., Francisella sp., Haemophilus sp., Helicobacter sp., Kingella sp., Klebsiella sp., Legionella sp., Moraxella sp., Morganella sp., Neisseria sp., Pasteurella sp., Plesiomonas sp., Proteus sp., Providencia sp., Pseudomonas sp., Salmonella sp., Serratia sp., Shigella sp.), Stenotrophomonas sp., Streptobacillus sp., Vibrio sp., Yersinia sp., Chlamydophila sp., Rickettsia sp., Coxiella sp., Ehrlichia sp., or Bartonella sp. In an embodiment, the bacterial infection is caused by one or more Salmonella species. In an embodiment, the Salmonella species is food-borne toxic Salmonella: S. enterica serovar Typhimurium. In embodiments, the bacterial infection is caused by E. coli, Klebsiella pneumonia, or Enterobacter cloacea. In embodiments, the bacterial infection is resistant to one or more antibiotics. In embodiments, the methods of the disclosure further comprise administering one or more antibiotics. In embodiments, the antibiotic is a macrolide, tetracycline, fluoroquinolone, penicillin, cephalosporin, aminoglycoside, sulfonamide, β-lactam, tetracycline, trimethoprim-sulfamethoxazole, chloramphenicol, or lincosamide.

[0028] In an embodiment, the disclosure provides a method of inhibiting a bacterial efflux pump in a subject having a bacterial infection, the method comprising administering to the subject a pharma- ceutically acceptable amount of a compound of formula (I), (II), (III), (IV), or (V). In an embodiment, the bacterial infection is caused by an intracellular pathogen. In an embodiment, the bacterial infection is caused by a gram-negative bacterium. In embodiments, the bacterial infection is one or more of Salmonella sp., Acinetobacter sp., Actinobacillus sp., Aeromonas sp., Bacteroide sp., Bordetella sp., Brucella sp., Burkholderia sp., Prevotella sp., Porphyromonas sp., Campylobacter sp., Citrobacter sp., Edwarsiella sp., Eikenella sp., Enterobacter sp., Escherichia sp., sp., Francisella sp., Haemophilus sp., Helicobacter sp., Kingella sp., Klebsiella sp., Legionella sp., Moraxella sp., Morganella sp., Neisseria sp., Pasteurella sp., Plesiomonas sp., Proteus sp., Providencia sp., Pseudomonas sp., Salmonella sp., Serratia sp., Shigella sp.), Stenotrophomonas sp., Streptobacillus sp., Vibrio sp., Yersinia sp., Chlamydophila sp., Rickettsia sp., Coxiella sp., Ehrlichia sp., or Bartonella sp. In an embodiment, the bacterial infection is caused by one or more Salmonella species. In an embodiment, the Salmonella species is food-borne toxic Salmonella: S. enterica serovar Typhimurium. In embodiments, the bacterial infection is caused by E. coli, Klebsiella pneumonia, or Enterobacter cloacea. In embodiments, the bacterial infection is resistant to one or more antibiotics. In embodiments, the methods of the disclosure further comprise administering one or more antibiotics. In embodiments, the antibiotic is a macrolide, tetracycline, fluoroquinolone, penicillin, cephalosporin, aminoglycoside, sulfonamide, beta-lactam, tetracycline, trimethoprim-sulfamethoxazole, chloramphenicol, or lincosamide.

[0029]

[0029] In an embodiment, the present disclosure provides a method for increasing the susceptibility of Gram-negative bacteria to an antibiotic, the method comprising administering a compound of formula (I), (II), (III), (IV), or (V) in combination with an antibiotic.

[0030]

[0030] In an embodiment, the present disclosure provides a method for reversing or reducing antibiotic resistance in antibiotic-resistant Gram-negative bacteria, the method comprising administering a compound of formula (I), (II), (III), (IV), or (V). DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0031] Detailed Description definition

[0031] The term "pharmaceutical acceptable salt" includes both acid addition salts and base addition salts. Pharmaceutically acceptable salts include those obtained by reacting an active compound that functions as a base with an inorganic or organic acid to form a salt, such as salts of hydrochloric acid, sulfuric acid, phosphoric acid, methanesulfonic acid, camphorsulfonic acid, oxalic acid, maleic acid, succinic acid, citric acid, formic acid, hydrobromic acid, benzoic acid, tartaric acid, fumaric acid, salicylic acid, mandelic acid, carbonic acid, etc. Those skilled in the art will further recognize that acid addition salts can be prepared by reacting a compound with a suitable inorganic or organic acid through any of several known methods.

[0032] The term "treatment" refers to one or more of alleviating, relieving, delaying, relieving, improving, or managing at least one symptom of a condition in a subject. The term "treatment" can also refer to preventing, delaying the onset of one or more (i.e., the period before clinical symptoms of a condition), or reducing the risk of development or worsening of a condition.

[0033] The compounds of the present disclosure or their pharma- ceutically acceptable salts have at least one asymmetric center. The compounds of the present disclosure having one asymmetric center give rise to enantiomers whose absolute stereochemistry can be represented as (R)- and (S)- or (+) and (-). When the compounds of the present disclosure have three or more asymmetric centers, the compounds can exist as diastereomers or other stereoisomeric forms. The present disclosure is intended to include all such possible isomers, as well as their racemic and optically pure forms, whether or not they are specifically depicted herein. Optically active (+) and (-) or (R)- and (S)-isomers can be prepared using chiral synthons or chiral reagents or separated using conventional techniques, such as chromatography and fractional crystallization. Conventional techniques for preparing / separating individual enantiomers include chiral synthesis from suitable optically pure precursors or separation of the racemate (or racemate of a salt or derivative) using, for example, chiral high pressure liquid chromatography (HPLC). If the compounds described herein contain olefinic double bonds or other centers of geometric asymmetry, and unless otherwise specified, the compounds are intended to include both E and Z geometric isomers. Likewise, all tautomeric forms are intended to be included.

[0034]

[0034] "Stereoisomer" refers to a compound in which the same atoms are bonded by the same bonds but have different, incompatible three-dimensional structures. The present disclosure contemplates various stereoisomers and mixtures thereof, and includes "enantiomers," which refers to two stereoisomers whose molecules are non-superimposable mirror images of one another.

[0035]

[0035] The term "therapeutically effective" as applied to a dose or amount refers to a sufficient quantity of a compound or pharmaceutical formulation to result in a desired clinical benefit following administration to a patient in need thereof.

[0036]

[0036] The term "halo" refers to halogen. In particular, this term refers to fluorine, chlorine, bromine, and iodine.

[0037]

[0037] "Alkyl" or "alkyl group" refers to a fully saturated straight or branched hydrocarbon chain radical that is attached to the remainder of the molecule by a single bond. Included are alkyls containing any number of carbon atoms, including but not limited to 1-12. Alkyl groups containing up to 12 carbon atoms include C1-C 12 Alkyl, where alkyl has up to 10 carbon atoms, is C1-C 10 Alkyl, with alkyl having up to 6 carbon atoms being C1-C6 alkyl, and with alkyl having up to 5 carbon atoms being C1-C5 alkyl. C1-C5 alkyl includes C5 alkyl, C4 alkyl, C3 alkyl, C2 alkyl, and C1 alkyl (i.e., methyl). C1-C6 alkyl includes all of the moieties listed above for C1-C5 alkyl, but also includes C6 alkyl. C1-C 10 Alkyl includes all of the moieties described above for C1-C5 alkyl and C1-C6 alkyl, but also includes C7, C8, C9, and C 10 Also includes alkyl. Similarly, C1-C 12 Alkyl includes all of the above moieties, except that C 11 and C 12 Includes alkyl. C1-C 12 Non-limiting examples of alkyl include methyl, ethyl, n-propyl, i-propyl, sec-propyl, n-butyl, i-butyl, sec-butyl, t-butyl, n-pentyl, t-amyl, n-hexyl, n-heptyl, n-octyl, n-nonyl, n-decyl, n-undecyl, and n-dodecyl. Unless otherwise specified, alkyl groups can be optionally substituted.

[0038]

[0038] "Alkenyl" or "alkenyl group" refers to a straight or branched hydrocarbon chain having from 2 to 12 carbon atoms and having one or more carbon-carbon double bonds. Each alkenyl group is attached to the remainder of the molecule by a single bond. Alkenyl groups containing any number of carbon atoms from 2 to 12 are included. Alkenyl groups containing up to 12 carbon atoms include C2-C 12 Alkenyl, with up to 10 carbon atoms in the alkenyl group being C2-C10 An alkenyl group having up to 6 carbon atoms is C2-C6 alkenyl, and an alkenyl group having up to 5 carbon atoms is C2-C5 alkenyl. C2-C5 alkenyl includes C5 alkenyl, C4 alkenyl, C3 alkenyl, and C2 alkenyl. C2-C6 alkenyl includes all of the moieties listed above for C2-C5 alkenyl, but also includes C6 alkenyl. C2-C 10 Alkenyl includes all of the moieties listed above for C2-C5 alkenyl and C2-C6 alkenyl, but also includes C7, C8, C9, and C 10 Also includes alkenyl. Similarly, C2-C 12 Alkenyl includes all of the above moieties, except that C 11 and C 12 Includes alkenyl. C2-C 12 Non-limiting examples of alkenyl include ethenyl (vinyl), 1-propenyl, 2-propenyl (allyl), isopropenyl, 2-methyl-1-propenyl, 1-butenyl, 2-butenyl, 3-butenyl, 1-pentenyl, 2-pentenyl, 3-pentenyl, 4-pentenyl, 1-hexenyl, 2-hexenyl, 3-hexenyl, 4-hexenyl, 5-hexenyl, 1-heptenyl, 2-heptenyl, 3-heptenyl, 4-heptenyl, 5-heptenyl, 6-heptenyl, 1-octenyl, 2-octenyl, 3-octenyl, 4-octenyl, 5-octenyl, 6-octenyl, 7-octenyl, 1-nonenyl, 2-nonenyl, and 3-nonenyl. , 4-nonenyl, 5-nonenyl, 6-nonenyl, 7-nonenyl, 8-nonenyl, 1-decenyl, 2-decenyl, 3-decenyl, 4-decenyl, 5-decenyl, 6-decenyl, 7-decenyl, 8-decenyl, 9-decenyl, 1-undecenyl, 2-undecenyl, 3-undecenyl, 4-undecenyl, 5-undecenyl, 6-undecenyl, 7-undecenyl, 8-undecenyl, 9-undecenyl, 10-undecenyl, 1-dodecenyl, 2-dodecenyl, 3-dodecenyl, 4-dodecenyl, 5-dodecenyl, 6-dodecenyl, 7-dodecenyl, 8-dodecenyl, 9-dodecenyl, 10-dodecenyl, and 11-dodecenyl. Unless otherwise specified, an alkyl group can be optionally substituted.

[0039]

[0039] "Alkynyl" or "alkynyl group" refers to a straight or branched hydrocarbon chain having from 2 to 12 carbon atoms and having one or more carbon-carbon triple bonds. Each alkynyl group is attached to the remainder of the molecule by a single bond. Alkynyl groups containing any number of carbon atoms from 2 to 12 are included. Alkynyl groups containing up to 12 carbon atoms include those having C2-C 12 Alkynyl, up to 10 carbon atoms, is C2-C 10 An alkynyl group having up to 6 carbon atoms is C2-C6 alkynyl, and an alkynyl group having up to 5 carbon atoms is C2-C5 alkynyl. C2-C5 alkynyl includes C5 alkynyl, C4 alkynyl, C3 alkynyl, and C2 alkynyl. C2-C6 alkynyl includes all of the moieties listed above for C2-C5 alkynyl, but also includes C6 alkynyl. C2-C 10 Alkynyl includes all of the moieties listed above for C2-C5 alkynyl and C2-C6 alkynyl, but also includes C7, C8, C9, and C 10 Alkynyl is also included. Similarly, C2-C 12 Alkynyl includes all of the above moieties, except that C 11 and C 12 Alkynyl is also included. C2-C 12 Non-limiting examples of alkenyls include ethynyl, propynyl, butynyl, pentynyl, etc. Unless otherwise specified, alkyl groups can be optionally substituted.

[0040] "Alkoxy" refers to a group of the formula -OR a R refers to the group a is an alkyl, alkenyl, or alkynyl as defined above containing 1 to 12 carbon atoms. Unless otherwise specified, an alkoxy group can be optionally substituted.

[0041]

[0041] "Cycloalkyl" refers to a stable non-aromatic monocyclic or polycyclic fully saturated hydrocarbon group consisting only of carbon and hydrogen atoms, which may include fused or bridged ring systems connected to the remainder of the molecule by a single bond, having 3 to 20 carbon atoms, preferably 3 to 10 carbon atoms. Monocyclic cycloalkyl groups include, for example, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl. Polycyclic cycloalkyl groups include, for example, adamantyl, norbornyl, decalinyl, 7,7-dimethyl-bicyclo[2.2.1]heptanyl, and the like. Unless otherwise specified in the specification, cycloalkyl groups can be optionally substituted.

[0042]

[0042] "Haloalkyl" refers to an alkyl group, as defined above, substituted by one or more halo groups, as defined above, such as trifluoromethyl, difluoromethyl, trichloromethyl, 2,2,2-trifluoroethyl, 1,2-difluoroethyl, 3-bromo-2-fluoropropyl, 1,2-dibromoethyl, etc. Unless otherwise specified, a haloalkyl group may be optionally substituted.

[0043]

[0043] "Aryl" refers to a hydrocarbon ring system containing hydrogen, 6 to 18 carbon atoms, and at least one aromatic ring. For purposes of this disclosure, an aryl group may be a monocyclic, bicyclic, tricyclic, or tetracyclic ring system, which may include fused or bridged ring systems. Aryl groups include, but are not limited to, aryl groups derived from aceanthrylene, acenaphthylene, acephenanthrylene, anthracene, azulene, benzene, chrysene, fluoranthene, fluorene, as-indacene, s-indacene, indane, indene, naphthalene, phenalene, phenanthrene, pleiadene, pyrene, and triphenylene. Unless otherwise indicated, the term "aryl" is meant to include aryl groups that are optionally substituted.

[0044]

[0044] "Heterocyclyl", "heterocyclic ring" or "heterocycle" refers to a stable 3- to 20-membered ring group consisting of 2 to 12 carbon atoms and 1 to 6 heteroatoms selected from the group consisting of nitrogen, oxygen, and sulfur. Heterocyclyl groups can be monocyclic, bicyclic, tricyclic, or tetracyclic ring systems, which can include fused or bridged ring systems. The nitrogen, carbon, or sulfur atoms in a heterocyclyl group can be optionally oxidized and the nitrogen atoms can be optionally quaternized. Heterocyclyl groups can be partially or fully saturated. Examples of such heterocyclyl groups include, but are not limited to, dioxolanyl, decahydroisoquinolyl, imidazolinyl, imidazolidinyl, isoxazolidinyl, morpholinyl, octahydroindolyl, octahydroisoindolyl, 2-oxopiperazinyl, 2-oxopiperidinyl, 2-oxopyrrolidinyl, oxazolidinyl, piperidinyl, piperazinyl, 4-piperidonyl, pyrrolidinyl, pyrazolidinyl, quinuclidinyl, thiazolidinyl, tetrahydrofuryl, trithianyl, tetrahydropyranyl, thiomorpholinyl, thiamorpholinyl, 1-oxo-thiomorpholinyl, and 1,1-dioxo-thiomorpholinyl. Unless otherwise specified, heterocyclyl groups may be optionally substituted.

[0045]

[0045] "Heteroaryl" refers to a 5-20 membered ring system containing a hydrogen atom, 1-13 carbon atoms, 1-6 heteroatoms (selected from the group consisting of nitrogen, oxygen, and sulfur), and at least one aromatic ring. For purposes of this disclosure, a heteroaryl group can be a monocyclic, bicyclic, tricyclic, or tetracyclic ring system, which can include fused or bridged ring systems; and the nitrogen, carbon, or sulfur atoms in the heteroaryl group can be optionally oxidized; and the nitrogen atom can be optionally quaternized. Examples include, but are not limited to, azepinyl, acridinyl, benzimidazolyl, benzothiazolyl, benzindolyl, benzodioxolyl, benzofuranyl, benzoxazolyl, benzothiazolyl, benzothiadiazolyl, benzo[b][1,4]dioxepinyl, 1,4-benzodioxanyl, benzonaphthofuranyl, benzoxazolyl, benzodioxolyl, benzodioxinyl, benzopyranyl, benzopyranonyl, benzofuranyl, benzofuranonyl, benzothienyl (benzothiophenyl), benzotriazolyl, benzo[4,6]imidazo[1,2-a]pyridinyl, carbazolyl, cinnolinyl, dibenzofuranyl, dibenzothiophenyl, furanyl, furanonyl, isothiazolyl, imidazolyl, indazolyl, indophenyl, and aryl, indazolyl, isoindolyl, indolinyl, isoindolinyl, isoquinolyl, indolizinyl, isoxazolyl, naphthyridinyl, oxadiazolyl, 2-oxoazepinyl, oxazolyl, oxiranyl, 1-oxidopyridinyl, 1-oxidopyrimidinyl, 1-oxidopyrazinyl, 1-oxidopyridazinyl, 1-phenyl-1H-pyrrolyl, phenazinyl, phenothiazinyl, phenoxazinyl, phthalazinyl, pteridinyl, purinyl, pyrrolyl, pyrazolyl, pyridinyl, pyrazinyl, pyrimidinyl, pyridazinyl, quinazolinyl, quinoxalinyl, quinolinyl, quinuclidinyl, isoquinolinyl, tetrahydroquinolinyl, thiazolyl, thiadiazolyl, triazolyl, tetrazolyl, triazinyl, and thiophenyl (i.e., thienyl). Unless otherwise specified, a heteroaryl group can be optionally substituted.

[0046]

[0046] As used herein, the term "substituted" means any of the above groups in which at least one hydrogen atom is replaced by a bond to a non-hydrogen atom, such as, but not limited to: a halogen atom, such as F, Cl, Br, and I; an oxygen atom in groups such as hydroxyl, alkoxy, and ester groups; a sulfur atom in groups such as thiol, thioalkyl, sulfone, sulfonyl, and sulfoxide groups; a nitrogen atom in groups such as amines, amides, alkylamines, dialkylamines, arylamines, alkylarylamines, diarylamines, N-oxides, imides, and enamines; a silicon atom in groups such as trialkylsilyl, dialkylarylsilyl, alkyldiarylsilyl, and triarylsilyl groups; and other heteroatoms in various other groups. "Substituted" also refers to any of the above groups in which one or more hydrogen atoms are replaced by a higher bond (e.g., a double bond or a triple bond) to a heteroatom, such as oxygen in oxo, carbonyl, carboxyl, and ester groups; and nitrogen in groups such as imines, oximes, hydrazones, and nitriles. For example, "substituted" refers to any of the above groups in which one or more hydrogen atoms are replaced by a higher bond (e.g., a double bond or a triple bond) to a heteroatom, such as oxygen in oxo, carbonyl, carboxyl, and ester groups; and nitrogen in groups such as imines, oximes, hydrazones, and nitriles. g R h , -NR g C(=O)R h , -NR g C(=O)NR g R h , -NR g C(=O)OR h , -NR g SO2R h , -OC(=O)NR g R h , -OR g , -SR g , -SOR g , -SO2R g , -OSO2R g , -SO2OR g , =NSO2R g , and -SO2NR g R h Substitution also includes any of the above groups substituted with one or more hydrogen atoms. g , -C(=O)ORg , -C(=O)NR g R h , -CH2SO2R g , -CH2SO2NR g R h In the above, R g and R h are the same or different and are independently hydrogen, alkyl, alkenyl, alkynyl, alkoxy, alkylamino, thioalkyl, aryl, aralkyl, cycloalkyl, cycloalkenyl, cycloalkynyl, cycloalkylalkyl, haloalkyl, haloalkenyl, haloalkynyl, heterocyclyl, N-heterocyclyl, heterocyclylalkyl, heteroaryl, N-heteroaryl, and / or heteroarylalkyl. "Substituted" further refers to any of the above groups in which one or more hydrogen atoms have been replaced by a bond to an amino, cyano, hydroxyl, imino, nitro, oxo, thioxo, halo, alkyl, alkenyl, alkynyl, alkoxy, alkylamino, thioalkyl, aryl, aralkyl, cycloalkyl, cycloalkenyl, cycloalkynyl, cycloalkylalkyl, haloalkyl, haloalkenyl, haloalkynyl, heterocyclyl, N-heterocyclyl, heterocyclylalkyl, heteroaryl, N-heteroaryl, and / or heteroarylalkyl group. In addition, each of the foregoing groups can also be optionally substituted with one or more of the above groups.

[0047] compound

[0047] In an embodiment, the present disclosure provides a compound that can be used to treat bacterial infection. In an embodiment, the compound inhibits efflux pumps, which reduces the ability of bacteria to excrete antibiotics. Thus, by inhibiting efflux pumps, the compounds of the present disclosure also increase bacterial sensitivity to antibiotics. Thus, in an embodiment, the compounds can be used in combination with antibiotics to treat antibiotic-resistant bacteria, reduce the ability of bacteria to acquire resistance to antibiotics, or increase the sensitivity of bacteria to antibiotics.

[0048] In some embodiments, the present disclosure provides a compound of formula (I): [ka] or a stereoisomer or a pharma- ceutically acceptable salt thereof.

[0049] In embodiments, n is 1, 2, or 3.

[0050] In embodiments, p is 1, 2, 3, 4, or 5.

[0051] In embodiments, m is 1, 2, 3, 4, or 5.

[0052] In an embodiment, each R 1 is independently halo, alkyl, or haloalkyl. In embodiments, the alkyl is substituted. In embodiments, the alkyl is unsubstituted.

[0053] In an embodiment, R 2 is -H, alkyl, alkenyl, or alkynyl. In embodiments, the alkyl is substituted. In embodiments, the alkyl is unsubstituted. In embodiments, the alkenyl is substituted. In embodiments, the alkenyl is unsubstituted. In embodiments, the alkynyl is substituted. In embodiments, the alkynyl is unsubstituted.

[0054] In an embodiment, R 3 is -H, alkyl, alkenyl, or alkynyl. In embodiments, the alkyl is substituted. In embodiments, the alkyl is unsubstituted. In embodiments, the alkenyl is substituted. In embodiments, the alkenyl is unsubstituted. In embodiments, the alkynyl is substituted. In embodiments, the alkynyl is unsubstituted.

[0055] In an embodiment, R 4is alkyl, alkenyl, alkynyl, cycloalkyl, or heterocyclyl, each of which optionally has one or more R 5 In embodiments, the alkyl is substituted. In embodiments, the alkyl is unsubstituted. In embodiments, the alkenyl is substituted. In embodiments, the alkenyl is unsubstituted. In embodiments, the alkynyl is substituted. In embodiments, the alkynyl is unsubstituted. In embodiments, the cycloalkyl is substituted. In embodiments, the cycloalkyl is unsubstituted. In embodiments, the heterocyclyl is substituted. In embodiments, the heterocyclyl is unsubstituted.

[0056] In embodiments, R 3 and R 4 taken together form a heterocyclyl, which optionally has one or more R 5 is replaced by.

[0057] In embodiments, R 5 is aryl, heteroaryl, alkyl, NH2, NHR A , or NR A R B or alkyl substituted with -NH. In embodiments, R A is alkyl, alkenyl, or alkynyl, each of which is optionally substituted with -OH or alkoxy. B is alkyl, alkenyl, or alkynyl. In embodiments, the alkyl is substituted. In embodiments, the alkyl is unsubstituted. In embodiments, the alkenyl is substituted. In embodiments, the alkenyl is unsubstituted. In embodiments, the alkynyl is substituted. In embodiments, the alkynyl is unsubstituted.

[0058] In embodiments, n is 1, 2, or 3. In embodiments, n is 1 or 2. In embodiments, n is 1. In embodiments, n is 2. In embodiments, each R 1 is independently halo, alkyl, or haloalkyl. In embodiments, each R1 is independently halo or haloalkyl. In embodiments, n is 2 and each R 1 is independently halo. In an embodiment, n is 1 and R 1 is haloalkyl. In embodiments, haloalkyl is C1-C6 alkyl substituted with one, two, three or more fluorines. In embodiments, haloalkyl is -CF3. In embodiments, p is 1, 2, 3, 4, or 5. In embodiments, p is 1.

[0059] In an embodiment, m is 1, 2, 3, 4, or 5. In an embodiment, m is 1.

[0060] In an embodiment, R 2 is -H, alkyl, alkenyl, or alkynyl. 2 is -H.

[0061] In an embodiment, R 3 is -H, alkyl, alkenyl, or alkynyl. 3 is -H or alkyl. In embodiments, R 3 is -H. In embodiments, R 3 is alkyl. In embodiments, R 3 is C1-C6 alkyl. In embodiments, R 3 is methyl or ethyl. In embodiments, R 3 is methyl.

[0062] In embodiments, R 4 is alkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, each of which optionally has one or more R 5 In embodiments, R 4 is alkyl or heterocyclyl, each of which optionally has one or more R 5 In embodiments, R 4 is C1-C6 alkyl. In embodiments, R4 is methyl, ethyl, propyl (e.g., n-propyl, i-propyl, sec-propyl) or butyl (e.g., n-butyl, i-butyl, sec-butyl, t-butyl). 4 is propyl. In embodiments, R 4 is butyl. In embodiments, R 5 is alkyl-NH. In embodiments, R 5 is a C1-C6 alkyl (i.e., methyl, ethyl, n-propyl, i-propyl, sec-propyl, n-butyl, i-butyl, sec-butyl, t-butyl, n-pentyl, t-amyl, or n-hexyl) substituted with -NH2. 5 is C4 alkyl-NH2. In embodiments, R 5 is C1-NH2.

[0063] In an embodiment, R 4 is a 5-7 membered heterocyclyl having 1, 2 or 3 heteroatoms selected from O, N, or S, and optionally R 5 In embodiments, R 4 is 5-6 membered heterocyclyl. In an embodiment, R 4 is a 5-6 membered bicyclic heterocyclyl. In an embodiment, R 4 is a 6-membered heterocyclyl having one N heteroatom. In embodiments, R 4 is a 5,3 fused heterocyclyl having one N heteroatom.

[0064] In an embodiment, R 3 is -H or alkyl (e.g., C 1-3 alkyl), and R 4 is alkyl or heterocyclyl, and the alkyl or heterocyclyl is optionally represented by one R 5 In embodiments, R 5 is -NH2. In embodiments, the compound has the following structure: [ka] or a pharma- ceutically acceptable salt thereof.

[0065] In an embodiment, R 3 and R 4 are taken together to form a heterocyclyl. In embodiments, the heterocyclyl can be a 5- to 8-membered heterocyclyl, which can optionally be one or more R 5 In embodiments, R 3 and R 4 are taken together to form a 5-membered heterocyclyl, which is optionally substituted with -NH or alkyl-NH. In embodiments, R 3 and R 4 are taken together to form a 5-membered heterocyclyl, which is substituted with -NH or C-C alkyl-NH (e.g., -CH-NH). 3 and R 4 taken together form a 6-membered heterocyclyl, which may optionally be joined by one or more R 5 In embodiments, R 3 and R 4 taken together form a 6-membered heterocyclyl, which is further comprised of one or more R 5 is replaced by R 5 is alkyl, -NH2, or -NR A R B and R A is alkyl optionally substituted with alkoxy; R B is alkyl. In embodiments, R 3 and R 4 taken together form a six-membered heterocyclyl, R 5 is C1-C6 alkyl 、 -NH2 or -NR A R B and R A is C1-C6 alkyl substituted with C1-C6 alkoxy; R B is C1-C6 alkyl. In embodiments, R 3 and R4 are taken together to form a 6-membered heterocyclyl, which is represented by R 5 is replaced by R 5 is -NH2 or -NR A R B and R A is a C2-C6 alkyl substituted with a C1-C3 alkoxy; R B is C1-C3 alkyl.

[0066] In embodiments, R 5 is aryl, heteroaryl, alkyl, -NH2, -NHR A , or -NR A R B or alkyl-NH. In embodiments, R 5 is a 6- or 8-membered aryl, a 5- to 8-membered heteroaryl, a C1-C6 alkyl, -NH2, -NHR A , or -NR A R B or C1-C6 alkyl substituted with -NH2. 5 is a 5-membered heteroaryl. In embodiments, R 5 is imidazolyl. In embodiments, R 3 and R 4 taken together form a six-membered heterocyclyl, R 5 is a 5-membered heteroaryl (eg, imidazolyl).

[0067] In an embodiment, R A is alkyl, alkenyl, alkynyl, each of which is optionally substituted with -OH or alkoxy. A is C1-C6 alkyl. In embodiments, R 4 is methyl. In embodiments, R 4 is ethyl. In embodiments, R 4 is propyl. A is C1-C6 alkyl substituted with C1-C6 alkyloxy. 4is C1-C6 alkyl substituted with methoxy. In embodiments, R 4 is propyl substituted with methoxy.

[0068] In an embodiment, R B is alkyl, alkenyl, alkynyl, each of which is optionally substituted with -OH or alkoxy. B is C1-C6 alkyl. In embodiments, R B is methyl.

[0069] In an embodiment, the compound of formula (I) has the formula (II): [ka] or a stereoisomer or a pharma- ceutically acceptable salt thereof.

[0070] In an embodiment, n is 1, 2, or 3. In an embodiment, n is 1. In an embodiment, n is 2.

[0071] In embodiments, p is 1, 2, 3, 4, or 5. In embodiments, p is 1. In embodiments, m is 1, 2, 3, 4, or 5. In embodiments, m is 1.

[0072] In embodiments, q is 1, 2, or 3. In embodiments, q is 1. In embodiments, q is 2.

[0073] In an embodiment, each R 1 is independently halo, alkyl, or haloalkyl. In embodiments, each R 1 is independently halo or haloalkyl. In embodiments, n is 2 and each R 1 is independently halo. In embodiments, n is 2 and each R 1 is independently -Cl. In an embodiment, n is 1 and R 1is haloalkyl. In embodiments, haloalkyl is C1-C6 alkyl substituted with one, two, three or more fluorines. In embodiments, haloalkyl is -CF3.

[0074] In embodiments, R 2 is hydrogen, C 1-6 Alkyl, C 1-6 Alkenyl, or C 1-6 In embodiments, R 2 is hydrogen.

[0075] In an embodiment, the A ring is heterocyclyl. In an embodiment, the A ring is a 5-8 membered heterocyclyl, optionally having 1, 2, or 3 heteroatoms selected from N, O, or S in addition to the ring N shown in formula (II). In an embodiment, the A ring is a 5-8 membered heterocyclyl.

[0076] In embodiments, R 5 is aryl, heteroaryl, alky, NH2, NHR A , or NR A R B In an embodiment, R 5 is NH2. In embodiments, R 5 is methyl, ethyl, or propyl (n-propyl, i-propyl, sec-propyl). 5 is methyl.

[0077] In embodiments, R A is C 1-6 Alkyl, C 1-6 Alkenyl, C 1-6 alkynyl, each of which is optionally halo, OH, C 1-6 In embodiments, R is substituted with alkoxy. B is C 1-6 Alkyl, C 1-6 Alkenyl, C 1-6 alkynyl, each of which is optionally halo, OH, C 1-6In embodiments, R is substituted with alkoxy. 5 Heteroaryl, NH2, NHR A , or NR A R B In an embodiment, R A Optionally, C 1-6 Alkoxy-substituted C 1-6 In embodiments, R A is propyl substituted with methoxy. In embodiments, R B is C 1-6 In embodiments, R B is methyl or ethyl. In embodiments, R B is methyl.

[0078] In an embodiment, the compound of formula (I) or (II) has formula (III): [ka] or a stereoisomer or a pharma- ceutically acceptable salt thereof.

[0079] In an embodiment, the compound of formula (I), (II), or (III) has formula (III-1) or (III-2): [ka] or a stereoisomer or a pharma- ceutically acceptable salt thereof.

[0080] In an embodiment, n is 1 or 2. In an embodiment, n is 2.

[0081] In an embodiment, p is 1, 2, or 3. In an embodiment, p is 1.

[0082] In an embodiment, m is 1, 2, or 3. In an embodiment, m is 1.

[0083] In an embodiment, q is 1.

[0084] In an embodiment, each R 1 is independently halo or haloalkyl. In embodiments, each R 1 is independently halo. In embodiments, n is 2 and each R 1 are independently halo. 1 In embodiments, n is 2 and each R 1 are independently -Cl.

[0085] In embodiments, the A ring is a 5- or 6-membered heterocyclyl.

[0086] In embodiments, R 5 is heteroaryl, C 1-6 Alkyl, C 1-6 Alkyl-NH2, -NH2, -NHR A , or -NR A R B In an embodiment, R 5 is -NH2. In embodiments, R 5 is C 1-6 alkyl (e.g., -CH) and -NH. In embodiments, R A Optionally, C 1-6 Alkoxy-substituted C 1-6 In embodiments, R A is propyl substituted with methoxy. In embodiments, R B is C 1-6 In embodiments, R B is methyl.

[0087] In embodiments, the compound of formula (I), (II), or (III) may be represented by formula (IV): [ka] or a stereoisomer or a pharma- ceutically acceptable salt thereof.

[0088] In embodiments, the compound of formula (I), (II), (III), or (IV) has formula (IV-1) or (IV-2): [ka] or a stereoisomer or a pharma- ceutically acceptable salt thereof.

[0089] In embodiments, n is 1 or 2. In embodiments, each R 1 is Cl or fluoroalkyl. In embodiments, n is 2 and each R 1 is Cl. In an embodiment, n is 1 and R 1 are independently -CF3.

[0090] In an embodiment, q is 1. In an embodiment, R 5 is alkyl substituted with -NH2, or -NH2, -NHR A , or NR A R B In an embodiment, R A Optionally, C 1-6 Alkoxy-substituted C 1-6 In embodiments, R B is C 1-6 In embodiments, R 5 is alkyl substituted with -NH2, or -NH2.

[0091] In an embodiment, the compound of formula (IV) has formula (IV.A) or (IV.B): [ka] It has the structure:

[0092] In embodiments, the compound of formula (IV) has the following structure: [ka] or a stereoisomer or a pharma- ceutically acceptable salt thereof.

[0093] In embodiments, the compound of formula (IV) has the following structure: [ka] or a pharma- ceutically acceptable salt thereof.

[0094] In embodiments, the compound of formula (I), (II), or (III) has formula (V): [ka] or a stereoisomer or a pharma- ceutically acceptable salt thereof.

[0095] In embodiments, n is 1 or 2. In embodiments, each R 1 is Cl or fluoroalkyl. In embodiments, n is 2 and each R 1 is Cl. In an embodiment, n is 1 and R 1 is fluoroalkyl. In embodiments, fluoroalkyl is C1-C6 alkyl substituted with one, two, three, or more fluorines. In embodiments, fluoroalkyl is -CF3. In embodiments, q is 1. In embodiments, R 5 Heteroaryl, NH2, NHR A , or NR A R B In an embodiment, R A Optionally, C 1-6 Alkoxy-substituted C 1-6 It is an alkyl group. B is C 1-6 It is an alkyl group. A is C 1-6 Alkoxy-substituted C 1-6 In embodiments, R A is a methoxy-substituted C 1-6 In embodiments, R Ais propyl or butyl substituted with -OCH. In embodiments, R B is C 1-6 In embodiments, R B is -CH3.

[0096] In embodiments, R 5 is a 5-7 membered heteroaryl having 1, 2, or 3 heteroatoms selected from N and S. In an embodiment, R 5 is a 5-membered heteroaryl having 1 or 2 N heteroatoms. In embodiments, R 5 is imidazolyl.

[0097] In an embodiment, the compound of formula (V) has the formula (VA)-(VF): [ka] It has the structure:

[0098] In embodiments, the compound of formula (V) has the following structure: [ka] or a pharma- ceutically acceptable salt thereof.

[0099] General Synthesis

[0099] The compounds disclosed herein, including their salts, can be prepared using known organic synthesis techniques and can be synthesized according to any of a number of possible synthetic routes. Those skilled in the art of organic synthetic chemistry will understand that the choice of starting materials and reagents will depend in part on the desired product and / or the reagents used, for example, various mechanisms require primary or secondary alcohols.

[0100]

[0100] The reaction for preparing the compounds disclosed herein can be carried out in a suitable solvent that can be easily selected by those skilled in the art of organic synthesis. A suitable solvent can be substantially non-reactive with the starting material, intermediate, or product at the temperature at which the reaction is carried out, which can range from room temperature to the boiling temperature of the solvent. The selection of a suitable protecting group can be easily determined by those skilled in the art. A given reaction can be carried out in one solvent or a mixture of solvents.

[0101]

[0101] In embodiments, the compounds disclosed herein can be prepared according to the following Schemes I-IV. Scheme I: Synthesis of epoxide intermediates [ka] During the ceremony: X is H or a cation; LG is a leaving group such as a halide; n is 1, 2, or 3; p is 1, 2, 3, 4, or 5; Each R 1 is independently halo, alkyl, or haloalkyl. Scheme II: General synthesis of compounds [ka] During the ceremony: n is 1, 2, or 3; p is 1, 2, 3, 4, or 5; m is 1, 2, or 3; q is 1 or 2; and Each R 1 is independently halo, alkyl, or haloalkyl; R 5 is aryl, heteroaryl, alky, NH2, NHR A , or NR A R; R A each of which is optionally substituted with -OH 1-6 Alkyl, C1-6 Alkenyl or C 1-6 Alkynyl, or C 1-6 is alkoxy; R B each of which is optionally substituted with -OH 1-6 Alkyl, C 1-6 Alkenyl or C 1-6 Alkynyl, or C 1-6 It is an alkoxy. Scheme III: General synthesis of compounds [ka] During the ceremony: n is 1, 2, or 3; p is 1, 2, 3, 4, or 5; q is 1 or 2; and Each R 1 is independently halo, alkyl, or haloalkyl; R 3 is -H, alkyl, alkenyl, or alkynyl; R 4 is alkyl, alkenyl, alkynyl, cycloalkyl, or heterocyclyl, each of which optionally has one or more R 5 or R 3 and R 4 taken together form a heterocyclyl, which optionally has one or more R 5 is replaced by R 5 is aryl, heteroaryl, alkyl, NH2, NHR A , or NR A R B or alkyl substituted with -NH2; R A is alkyl, alkenyl, alkynyl, each of which is optionally substituted with -OH or alkoxy; and R B is alkyl, alkenyl, or alkynyl. Scheme IV: Deprotection of the amine [ka] During the ceremony, n is 1, 2, or 3; m is 1, 2, 3, 4, or 5; q is 1 or 2; and Each R 1 is independently halo, alkyl, or haloalkyl.

[0102] Treatment method In an embodiment, the disclosure provides a method of treating a bacterial infection in a subject in need thereof, the method comprising administering to the subject a pharma- ceutically acceptable amount of one or more compounds of formula (I), (II), (III), (IV), (V) or stereoisomers or salts thereof. In an embodiment, the disclosure provides a method of inhibiting a bacterial efflux pump in a subject having a bacterial infection, the method comprising administering to the subject a pharma- ceutically acceptable amount of one or more compounds of formula (I), (II), (III), (IV), (V) or stereoisomers or pharma- ceutical acceptable salts thereof.

[0103]

[0103] In an embodiment, the bacterial infection is caused by a gram-negative bacterium. In an embodiment, the gram-negative bacterium may be an intracellular pathogen.

[0104] In embodiments, the bacterial infection is caused by one or more of Salmonella sp., Acinetobacter sp., Actinobacillus sp., Aeromonas sp., Bacteroide sp., Bordetella sp., Brucella sp., Burkholderia sp., Prevotella sp., Porphyromonas sp., Campylobacter sp., Citrobacter sp., Edwarsiella sp., Eikenella sp., Enterobacter sp., or any combination thereof. sp., Escherichia sp., Francisella sp., Haemophilus sp., Helicobacter sp., Kingella sp., Klebsiella sp., Legionella sp., Moraxella sp., Morganella sp., Neisseria sp., Pasteurella sp., Plesiomonas sp., Proteus sp., Providencia sp., Pseudomonas sp., Salmonella sp., Serratia sp., Shigella sp. sp., Stenotrophomonas sp., Streptobacillus sp., Vibrio sp., Yersinia sp., Chlamydophila sp., Rickettsia sp.), Coxiella sp., Ehrlichia sp., or Bartonella sp. .

[0105]

[0105] Acinetobacter baumannii, Acinetobacter haemolyticus, Actinobacillus actinomycetemcomitans, Aeromonas hydrophila, Bacteroides fragilis, Bacteroides theataioatamides theataioatamides theataioatides distasonis, Bacteroides ovatus, Bacteroides vulgatus, Bordetella pertussis pertussis, Brucella melitensis, Burkholderia cepacia, Burkholderia pseudomallei, Burkholderia mallei, Prevotella corporis, Prevotella intermedia, Prevotella endodontalis, Porphyromonas asaccharolytica, Campylobacter jejuni, Campylobacter coli, Campylobacter fetus, Citrobacter freundii freundii, Citrobacter koseri, Edwarsiella tarda, Eikenella corrodenscorrodens, Enterobacter cloacae, Enterobacter aerogenes, Enterobacter agglomerans, Escherichia coli, Francisella tularensis, Haemophilus influenzae, Haemophilus ducreyi, Helicobacter pylori, Kingella kingae, Klebsiella pella pneumonia ella rhinoscleromatis, Klebsiella ozaenae, Legionella pneumophila penumophila, Moraxella catarrhalis, Morganella morganii, Neisseria gonorrhoeae, Neisseria meningitidis, Pasteurella multocida, Plesiomonas shigelloides, Proteus mirabilis, Proteus vulgaris, Proteus penneri, Proteus myxofaciens, Providencia stuartii, Providencia rettgeri, Providencia alcalifaciens alcalifaciens, Pseudomonas aeruginosa, Pseudomonas fluorescens, Salmonella typhityphi, S. enterica, Salmonella paratyphi, Serratia marcescens, Shigella flexneri, Shigella boydii, Shigella sonnei, Shigella dysenteriae, Stenotrophomonas maltophilia, Streptobacillus moniliformis, Vibrio cholerae, Vibrio parahaemolyticus, Vibrio vulnificus, Vibrio alginolyticus, Yersinia enterocolitica enterocolitica, Yersinia pestis, Yersinia pseudotuberculosis, Chlamydophila pneumoniae, Chlamydophila trachomatis, Rickettsia prowazekii, Coxiella burnetii, Ehrlichia chaffeensis, or Bartonella hensenae.

[0106] In an embodiment, the bacterial infection is caused by one or more Salmonella sp. In an embodiment, the Salmonella sp. is S. enterica serovar Typhimurium. In an embodiment, the bacterial infection is caused by E. coli. In an embodiment, the bacterial infection is caused by Klebsiella pneumonia. In an embodiment, the bacterial infection is caused by Enterobacter cloacae.

[0107] In embodiments, the compounds of the present disclosure can be administered in combination (separately, simultaneously, e.g., as part of the same composition in a combination product, or sequentially) with one or more antibiotics. In embodiments, the antibiotic is a macrolide, tetracycline, fluoroquinolone, penicillin, cephalosporin, aminoglycoside, sulfonamide, β-lactam, tetracycline, trimethoprim-sulfamethoxazole, chloramphenicol, or lincosamide.

[0108]

[0108] In embodiments, the antibiotic is penicillin G, penicillin V, methicillin, oxacillin, cloxacillin, dicloxacillin, nafcillin, ampicillin, amoxicillin, carbenicillin, ticarcillin, mezlocillin, piperacillin, azlocillin, temocillin, cepalothin, cephapirin, cephradine, cephaloridine, cefazolin, cefamandole, cefuroxime, cephalexin, cefprozil, cefaclor, loracarbef, cefoxitin, cefmatozole, cefotaxime, cephalosporin ... ftizoxime, ceftriaxone, cefoperazone, ceftazidime, cefixime, cefpodoxime, ceftibuten, cefdinir, cefpirome, cefepime, BAL5788, BAL9141, imipenem, ertapenem, meropenem, astreonam, clavulanic acid, sulbactam, tazobactam, streptomycin, neomycin, kanamycin, paromycin, gentamicin, tobramycin, amikacin, netilmicin, spectinomycin, sisomicin, dibekalin, isepamicin, Tetracycline, chlortetracycline, demeclocycline, minocycline, oxytetracycline, methacycline, doxycycline, erythromycin, azithromycin, clarithromycin, telithromycin, ABT-773, lincomycin, clindamycin, vancomycin, oritavancin, dalbavancin, teicoplanin, quinupristin and dalfopristin, sulfanilamide, para-aminobenzoic acid, sulfadiazine, sulfisoxazole, sulfamethoxazole, sulfatariidine alidine), linezolid, nalidixic acid, oxolinic acid, norfloxacin, perfloxacin, enoxacin, ofloxacin, ciprofloxacin, temafloxacin, lomefloxacin, fleroxacin, grepafloxacin, sparfloxacin, trovafloxacin, clinafloxacin, gatifloxacin, moxifloxacin, gemifloxacin, sitafloxacin, metronidazole, daptomycin, garenoxacin, ramoplanin, faropenem, polymyxin, tigecycline, AZD2563, or trimethoprim.

[0109] In embodiments, the compounds disclosed herein are used to treat gram-negative bacterial infections that have acquired resistance to antibiotics. The terms "resistance" and "antimicrobial resistance" refer to bacteria that can withstand exposure to one or more antibiotics. In embodiments, the bacteria can withstand exposure to aminoglycoside antibiotics (e.g., amikacin, gentamicin, kanamycin, neomycin, netilmicin, tobramycin, paromomycin, spectinomycin), ansamycin antibiotics (e.g., rifaximin, streptomycin), carbapenem antibiotics (e.g., ertapenem, doripenem, imipenem / cilastatin, meropenem), cephalosoprin antibiotics (e.g., cefadroxil, cefazolin, cefadroxil ... Cefaxolin, cefatolin, cephalexin, cefaclor, cefamandole, cefoxitin, cefprozil, cefuroxime, cefisime, cefdinir, cefditoren, cefoperazone, cefotaxime, cefpodoxime, ceftazidime, certibuten, ceftizoxime, ceftriaxone, cefepime, ceftaroline fosamil, ceftobiprole), glycopeptide antibiotics (e.g., teicoplanin, vancomycin, ampicillin, telavancin), lincosamide antibiotics (e.g., clindamycin, lincomycin), daptomycin, macrolide antibiotics (e.g., azithromycin, clarithromycin, dirithromycin, erythromycin, roxithromycin, troleandomycin, telithromycin, spiramycin), aztreonam, furazolidone, nitrofurantoin, oxazolidinone antibiotics (e.g., linezolid, posizolid, radezolid, dezolid, torezolid), penicillin antibiotics (e.g., amoxacillin, ampicillin, azlocillin, carbenicillin, cloxacillin, dicloxacillin, flucloxacillin, mezlocillin, methicillin, nafcillin, oxacillin, penicillin, piperacillin, temocillin, ticarcillin), amoxicillin / clavulante, ampicillin / sulbactam, piperacillin / tazobactam, ticarcillin / clavulanic acid,Quinolone antibiotics (e.g., ciprofloxacin, enoxacin, gatifloxacin, gemifloxacin, levofloxacin, lomefloxacin, moxifloxacin, nalidixic acid, norfloxacin, ofloxacin, trovafloxin, grepafloxacin, sparfloxacin, temafloxacin), suflonamide antibiotics (e.g., mafenide, sulfacetamide, sulfadiazine, sulfadiazine Resistant to one or more of silver, sulfadimethoxine, sulfamethizole, sulfamethoxazole, sulfanilimide, sulfasalazine, sulfisoxazole, trimethoprim / sulfamethoxazole-TMP-SMX), and tetracycline antibiotics (e.g., demeclocycline, doxycycline, minocycline, oxytetracycline, tetracycline, tigeclycline).

[0110] In embodiments in which a compound of the present disclosure is used in combination with an antibiotic, the compound reduces the MIC of the antibiotic (e.g., as measured in Example 2) by about 1.5-fold, about 2-fold, about 2.5-fold, about 3-fold, about 3.5-fold, about 4-fold, about 4.5-fold, about 5-fold, about 5.5-fold, about 6-fold, about 6.5-fold, about 7-fold, about 7.5-fold, about 8-fold, about 8.5-fold, about 9-fold, about 9.5-fold, or about 10-fold (including all values ​​and ranges therebetween). In embodiments where a compound of the present disclosure is used in combination with an antibiotic, the dose of the antibiotic can be reduced by about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, or more (including all values ​​and ranges therebetween) (compared to the dose of the antibiotic administered in the absence of a disclosed compound). In embodiments, a compound of the present disclosure can allow the antibiotic to be effective at the same dose as when the antibiotic was administered in the absence of one of the disclosed compounds and was ineffective.

[0111] In embodiments, the present disclosure provides methods of increasing the susceptibility of bacteria to antibiotic treatment by administering a compound of the present disclosure and one or more antibiotics.

[0112] In an embodiment, the method comprises administering to a subject in need thereof a pharma- ceutically acceptable amount of one or more compounds of formula (I), (II), (III), (IV), (V) or stereoisomers or pharma- ceutically acceptable salts thereof and one or more antibiotics. In an embodiment, the method increases the susceptibility of Gram-negative bacteria to antibiotics. In an embodiment, the compounds of the present disclosure can render antibiotic-resistant strains susceptible to antibiotics to which they are otherwise resistant. In an embodiment, the antibiotic is a macrolide, a tetracycline, a fluoroquinolone, a penicillin, a cephalosporin, an aminoglycoside, a sulfonamide, a β-lactam, a tetracycline, a trimethoprim-sulfamethoxazole, a chloramphenicol, or a lincosamide.

[0113] In embodiments where a compound of the present disclosure is used in combination with an antibiotic, the compound increases antibiotic susceptibility by reducing the MIC of the antibiotic by about 1.1-fold, about 1.2-fold, about 1.3-fold, about 1.4-fold, about 1.5-fold, about 2-fold, about 2.5-fold, about 3-fold, about 3.5-fold, about 4-fold, about 4.5-fold, about 5-fold, about 5.5-fold, about 6-fold, about 6.5-fold, about 7-fold, about 7.5-fold, about 8-fold, about 8.5-fold, about 9-fold, about 9.5-fold, or about 10-fold (including all values ​​and ranges therebetween). In embodiments where a compound of the present disclosure is used in combination with an antibiotic, the compound increases antibiotic susceptibility by reducing the dose and / or IC 50by about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, or more (including all values ​​and ranges therebetween) (compared to the dose of antibiotic administered in the absence of a disclosed compound). In embodiments, the disclosed compounds can allow an antibiotic to be effective at the same dose as when the antibiotic was administered in the absence of one of the disclosed compounds and was ineffective.

[0114] In an embodiment, the present disclosure also provides a method for reversing or reducing antibiotic resistance of antibiotic-resistant Gram-negative bacteria. In an embodiment, the method comprises administering to a subject in need thereof a pharma- ceutically acceptable amount of one or more compounds of formula (I), (II), (III), (IV), (V) or stereoisomers or pharma- ceutically acceptable salts thereof and one or more antibiotics. In an embodiment, the antibiotic is a macrolide, a tetracycline, a fluoroquinolone, a penicillin, a cephalosporin, an aminoglycoside, a sulfonamide, a β-lactam, a tetracycline, a trimethoprim-sulfamethoxazole, a chloramphenicol, or a lincosamide.

[0115] In embodiments, the reversal or reduction of antibiotic resistance is determined by assessing the reduction in the growth rate of antibiotic-resistant strains in the presence of the compound and one or more antibiotics. In embodiments, the compounds of the present disclosure administered in combination with antibiotics reverse or reduce antibiotic resistance by reducing bacterial virulence, inhibiting or killing antibiotic-resistant bacteria, preventing biofilm formation, preventing septic shock, treating sepsis, and / or increasing bacterial susceptibility to antibiotics to which they were previously resistant. In embodiments where the compounds of the present disclosure are used in combination with antibiotics, the virulence or growth rate of antibiotic-resistant bacteria can be reduced by about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, or more (including all values ​​and ranges therebetween) (compared to the dose of antibiotic administered in the absence of the disclosed compounds). In embodiments in which a compound of the disclosure is used in combination with an antibiotic, the compound reverses or reduces antibiotic resistance by reducing the MIC of the antibiotic by about 1.1-fold, about 1.2-fold, about 1.3-fold, about 1.4-fold, about 1.5-fold, about 2-fold, about 2.5-fold, about 3-fold, about 3.5-fold, about 4-fold, about 4.5-fold, about 5-fold, about 5.5-fold, about 6-fold, about 6.5-fold, about 7-fold, about 7.5-fold, about 8-fold, about 8.5-fold, about 9-fold, about 9.5-fold, or about 10-fold (including all values ​​and ranges therebetween).

[0116] Pharmaceutical Compositions

[0116] In some embodiments of the present disclosure, the pharmaceutical composition comprises a therapeutically effective amount of one or more compounds of formula (I), (II), (III), (IV), or (V) or a stereoisomer or pharma- ceutically acceptable salt thereof.

[0117] In an embodiment, a pharmaceutical composition is provided that includes one or more compounds disclosed herein or their stereoisomers or pharma- ceutically acceptable salts, and a pharma- ceutically acceptable excipient or adjuvant. Pharmaceutically acceptable excipients and adjuvants are added to compositions or formulations for various purposes. In an embodiment, the pharmaceutical composition includes a pharma- ceutical acceptable carrier, binder, and / or diluent. In an embodiment, the pharmaceutical composition may include additional materials useful for physically formulating various dosage forms of the compositions of the present disclosure, such as dyes, flavoring agents, preservatives, antioxidants, opacifiers, thickeners, stabilizers, lubricants, wetting agents, emulsifiers, salts that affect osmotic pressure, buffers, colorants, flavors, and / or aromatic substances.

[0118]

[0118] In certain embodiments, the pharmaceutical composition of the present disclosure may additionally contain other auxiliary ingredients conventionally found in pharmaceutical compositions at their art-established usage levels.Thus, for example, the pharmaceutical composition may contain additional compatible pharma- ceutical active materials such as antipruritic agents, astringents, local anesthetics, or anti-inflammatory agents.

[0119] The compounds of the present disclosure can be formulated into preparations that contain pharma- ceutically acceptable carriers, adjuvants, and vehicles for administration by various means, including oral and parenteral.The term parenteral as used herein includes subcutaneous injection, intravenous injection, intramuscular injection, and intraarterial injection by various infusion techniques.Intraarterial injection and intravenous injection as used herein include administration via catheter.

[0120] The compounds disclosed herein can be formulated according to routine procedures that are compatible with desired administration routes.Accordingly, the compounds disclosed herein can take the form of suspension, solution, or emulsion in oily or aqueous vehicles, and can contain suspending, stabilizing, and / or dispersing agents.The compounds disclosed herein can also be formulated as preparations for injection.

[0121]

[0121] In certain embodiments, the pharmaceutical compositions of the present disclosure are prepared using known techniques including, but not limited to, mixing, dissolving, granulating, dragee-making, pulverizing, emulsifying, encapsulating, entrapping, or tabletting processes.

[0122] In embodiments, the pharmaceutical composition may be a solid, powder, liquid, and gel. In embodiments, the medicament is a solid (e.g., a powder, tablet, capsule, granule, and / or aggregate). In certain such embodiments, the solid pharmaceutical composition includes one or more excipients known in the art, including, but not limited to, starch, sugar, diluent, granulating agent, lubricant, binder, and disintegrant.

[0123]

[0123] Suitable solid carriers for use in this application include, but are not limited to, sugars and sugar alcohols (such as lactose, glucose, and mannitol), starch, methylcellulose, magnesium stearate, dicalcium phosphate, calcium phosphate, magnesium stearate, talc, sugars, dextrin, starch, gelatin, cellulose, and polyvinylpyrrolidine.Solid carriers may further include one or more substances that act as flavoring agents, lubricants, solubilizers, suspending agents, fillers, glidants, compression aids, binders, or tablet disintegrants.Tablets can be produced by compression or molding, optionally with one or more accessory ingredients. Compressed tablets can be prepared by compressing in a suitable machine the active ingredient in a free-flowing form such as powder or granules, optionally mixed with a binder (e.g., povidone, gelatin, hydroxypropyl methylcellulose), lubricant, inert diluent, preservative, and / or disintegrant (e.g., sodium starch glycolate, cross-linked povidone, cross-linked sodium carboxymethylcellulose). Molded tablets can be made by molding in a suitable machine a mixture of the powdered compound moistened with an inert liquid diluent.

[0124] In an embodiment, the pharmaceutical composition is formulated as a liquid.The liquid pharmaceutical composition suitable for use in the present disclosure includes solutions, suspensions, emulsions, syrups, elixirs, and pressurized compounds.The active ingredient can be dissolved or suspended in a pharmaceutically acceptable liquid carrier, such as water, an organic solvent, a mixture of both, or a pharmaceutically acceptable oil or fat.In an embodiment, the liquid solution can be an aqueous solution or a non-aqueous solution.

[0125]

[0125] Examples of non-aqueous carriers include, but are not limited to, propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable organic esters.

[0126]

[0126] Aqueous carriers suitable for use in the present application include, but are not limited to, water, ethanol, alcoholic / aqueous solutions, glycerol, emulsions or suspensions, including saline and buffered media.

[0127]

[0127] Parenteral carriers suitable for use in this application include, but are not limited to, sodium chloride solution, Ringer's dextrose, dextrose and sodium chloride, lactated Ringer's, and fixed oils. Intravenous carriers include fluid and nutrient replenishers, electrolyte replenishers, such as those based on Ringer's dextrose. Preservatives and other additives may also be present, such as, for example, antibacterial agents, antioxidants, chelating agents, inert gases, and the like. Liquid carriers may include other suitable pharmaceutical additives, such as solubilizers, emulsifiers, buffers, preservatives, sweeteners, flavoring agents, suspending agents, thickeners, coloring agents, viscosity adjusting agents, stabilizers, or osmolality adjusting agents.

[0128] In an embodiment, the amount of a compound disclosed herein or a stereoisomer or a pharma- ceutically acceptable salt thereof can be administered at about 0.001 mg / kg to about 100 mg / kg body weight (e.g., about 0.01 mg / kg to about 10 mg / kg or about 0.1 mg / kg to about 5 mg / kg). In an embodiment, the amount of a compound disclosed herein or a stereoisomer or a pharma- ceutically acceptable salt thereof can be administered at about 0.1 mg to about 1,000 mg (e.g., about 0.1 mg to about 500 mg / kg or about 0.1 mg / kg to about 100 mg / kg). EXAMPLES

[0129] Working Example The following experiments were carried out using the SAFIRE assay described in US Patent Publication No. 2020 / 0022961, the entire contents of which are incorporated herein by reference for all purposes. SAFIRE (IntracellulaR Enterobacteriaceae Fluorescence Microscopy Anti-Infective Screening) uses cell culture as a surrogate to identify compounds that prevent intracellular replication of model Gram-negative human pathogens. The method identifies compounds that inhibit bacterial growth in broth cultures and compounds that reduce Gram-negative bacterial loads in macrophages at concentrations of 10 μM or less, but do not affect bacterial growth in standard Mueller Hinton Broth (MHB) at 100 μM. Thus, SAFIRE allows the discovery of compounds that are effective against Gram-negative bacteria in host cells and whole animals, regardless of whether they are antibacterial in broth.

[0130] Example 1. Inhibition curve Bacterial strains

[0131] Wild-type S. enterica serovar Typhimurium strain SL1344 was first isolated from the blood of an infected calf. For macrophage screening and validation, SL1344 sifB::gfp was grown overnight to saturation in Luria-Bertani broth (LB) containing 30 μg / ml streptomycin and 30 μg / ml kanamycin, diluted to an OD of 0.001, and frozen in 100 μL aliquots in 20% glycerol at -80°C. Prior to infection, aliquots containing 30 μg / ml streptomycin and 30 μg / ml kanamycin were grown in 5 mL LB medium with aeration at 37°C for 18 hours. Bacterial strains were routinely grown in LB containing the antibiotics: 30 μg / ml streptomycin, 30 μg / ml kanamycin, 50 μg / ml ampicillin, 10 μg / ml tetracycline, and / or 1.15 μg / ml meropenem. acrAB::kan and macAB::kan strains were constructed according to published methods. S. enterica subsp. enterica, serovar Typhimuriums strain S10801, NR-22067 is a multidrug-resistant isolate from a septic calf. This and other strains shown were obtained from BEI resources, NIAID, NIH.

[0131]

[0132] cell culture

[0133] Mouse macrophage-like RAW264.7 cells and HeLa human epithelial cells were obtained from the American Type Tissue Collection. BMDMs were isolated as previously described. Briefly, bone marrow was flushed from femurs of 1- to 4-month-old 129SvEvTac mice (Taconic Laboratories) bred in-house. Mononuclear cells were isolated using Histopaque-1083 (Sigma), washed, and directly seeded into assay plates at 1 × 105 cells / ml in complete medium supplemented with 35% conditioned medium from 3T3 cells expressing MCSF. After 3 days, the medium was refreshed. After 1 week, the medium was replaced with 100 μL of fresh medium, and cells were infected as described below. All three cell types were grown in DMEM high glucose (Sigma) supplemented with 10% fetal bovine serum, 2 mM L-glutamine, 1 mM sodium pyruvate, 10 mM HEPES, and 50 μM β-mercaptoethanol. Cells were maintained in a 5% CO2 humidified atmosphere at 37° C. For screening, frozen aliquots of RAW264.7 were thawed and grown for 3 days before seeding; other experiments were performed on cultures between passages 4 and 20.

[0132]

[0134] Bacterial infection for SAFIRE and CFU plating

[0135] SAFIRE-RAW264.7 macrophages (7x103 in 40 μL or 5x104 in 100 μL) were seeded in 384-well or 96-well black-walled glass-bottom plates (Brooks Automation), respectively. 24 hours after seeding, bacteria in 20 μL or 50 μL PBS were added to a final concentration of 1x107 CFU / mL, conditions that resulted in approximately 70% of the macrophages being infected 18 hours after infection with minimal toxicity to the macrophages. A sifB::gfp bacterial reporter strain was used to minimize the green signal from extracellular bacteria. 45 minutes after bacterial addition, 20 μL or 50 μL of gentamicin was added to a final concentration of 40 μg / mL. This did not affect intracellular infection but inhibited extracellular bacterial replication. Two hours after infection, 200 nL or 500 nL of compound was added using a pin tool (CyBio) to a final concentration of 25 μM. Each assay plate included rifampicin and DMSO controls. In some experiments, the medium was removed and replaced with fresh medium containing 40 μg / mL gentamicin and increasing concentrations of the disclosed compound. 17.5 hours after infection, Mito Tracker Red CMXRos (LifeTechnologies) in PBS was added to a final concentration of 300 nM or 100 nM to 384- or 96-well plates, respectively. After 30 minutes, 16% paraformaldehyde was added to a final concentration of 1 ± 2% and incubated for 15 minutes at room temperature. Wells were washed twice with PBS and stained with 1 μM DAPI for 20 minutes; wells were washed twice and stored in 90% glycerol in PBS until imaging. The Z' coefficients for the screening platform were 0.59 and 0.48 for 96-well and 384-well plates, respectively, which are within the published range for complex cell-based screens.

[0133]

[0136] Infection of HeLa cells with S. Typhimurium was performed as above, except that 1 x 104 cells were seeded and cells were infected with S. Typhimurium constitutively expressing GFP from the rpsM locus, as sifB::gfp is poorly expressed in HeLa cells, and plates were spun at 500 x g for 5 min after bacteria were added to promote infection.

[0134]

[0137] CFU-infection was performed as described above, except that cells were seeded into 96-well tissue culture-coated plates (Greiner). Eighteen hours after infection, wells were washed three times with PBS and lysed with 30 μL of 0.1% TritonX-100, diluted, and plated to determine CFU.

[0135]

[0138] The IC50 values ​​of compounds of the present disclosure were determined using SAFIRE at at least eight concentrations using two-fold dilutions ranging from 50 to 0.001 μM and are shown in Table A below.

[0136]

[0139] In addition, toxicity was measured according to the following procedures: (1) analysis of macrophage cell morphology; and (2) counting cells using MATLAB (cells that lift or float after treatment are considered dead as they are not in the proper Z-plane to be counted). Compounds are considered non-toxic if 70% of cells are attached (measured by comparing cell counts before and after treatment). Compounds that pass #1 and #2 above are considered to have a toxicity of "greater than 50uM". Compounds that do not pass either #1 or #2 are considered to have a toxicity of "less than 25uM". The data are shown in Table A below.

[0137] [Table 1]

[0138] [Table 2]

[0139] [Table 3]

[0140] [Table 4]

[0141] Example 2. Spectrum of broth activity To evaluate the ability of the disclosed compounds to susceptible bacteria to antibiotics, the disclosed compounds were combined with the known antibiotics doxycycline, ciprofloxacin, and chloramphenicol in several bacterial cell lines, including carbapenem-resistant Enterobacteriaceae, to measure minimum inhibitor concentrations (MICs). The MICs of the antibiotics alone (i.e., in the absence of the disclosed compounds) are shown in the first row marked "none." The comparative compounds EPI35 and PaβN, which have been reported to inhibit bacterial efflux pumps, and the disclosed compounds were added at 50 μg / mL, and the antibiotic MICs were re-evaluated in each cell line. These results are shown in Tables B-D. The disclosed compounds lowered the MICs of these antibiotics when used at a concentration of 50 μg / mL. At this concentration (50 μg / mL), the disclosed compounds lacked intrinsic antibacterial activity, yet significant improvements in efficacy were still observed. Thus, the disclosed compounds can be used to enhance the activity of antibiotics that have lost efficacy due to efflux.

[0142] [Table 5]

[0143] [Table 6]

[0144] [Table 7]

[0145] Example 3. Method for producing compounds

[0141] The compounds disclosed herein can be prepared according to general synthetic schemes I-IV as shown below, and / or by any other suitable method. The compounds can be characterized according to any suitable method known in the art, such as NMR, UV, HPLC, LC-MS, and TLC. Scheme I: Synthesis of epoxide intermediates [ka] During the ceremony: X is H or a cation; LG is a leaving group such as a halide; n is 1, 2, or 3; p is 1, 2, 3, 4, or 5; and Each R 1 is independently halo, alkyl, or haloalkyl.

[0146] A stirred mixture of 1.0 g of 3,4-fluorophenol, 1.5 equivalents of epichlorohydrin, and 2.0 equivalents of Cs2CO3 was heated at 80° C. overnight. The reaction was quenched with water, and the precipitate was purified in good yield by normal phase column chromatography using ethyl acetate and hexane. The product was used in the next step. The product was characterized by LC-MS. The desired mass was observed. Scheme II: General synthesis of compounds [ka] During the ceremony: n is 1, 2, or 3; p is 1, 2, 3, 4, or 5; m is 1, 2, or 3; q is 1 or 2; and Each R 1 is independently halo, alkyl, or haloalkyl; R 5 is aryl, heteroaryl, alky, NH2, NHR A , or NRA R B and; R A teeth, -6 Alkyl, C 1-6 Alkenyl, or C 1-6 alkynyl, each of which is optionally —OH or C 1-6 substituted with alkoxy; and R B is C 1-6 Alkyl, C 1-6 Alkenyl, or C 1-6 alkynyl, each of which is optionally —OH or C 1-6 Substituted with alkoxy.

[0147] The two starting materials (0.16 mMol) were mixed in 1 mL of deionized water in a 1:1 molar ratio and heated to 140° C. in a microwave for 10 minutes. The resulting product was purified by reversed-phase column chromatography to obtain the product. The product was characterized by LC-MS. The desired mass was observed as shown in Table E. Scheme III: General synthesis of compounds [ka] During the ceremony: n is 1, 2, or 3; p is 1, 2, 3, 4, or 5; q is 1 or 2; and Each R 1 is independently halo, alkyl, or haloalkyl; R 3 is -H, alkyl, alkenyl, or alkynyl; R 4 is alkyl, alkenyl, alkynyl, cycloalkyl, or heterocyclyl, each of which optionally has one or more R 5 or R 3 and R 4 taken together form a heterocyclyl, which optionally has one or more R 5is replaced by R 5 is aryl, heteroaryl, alkyl, NH2, NHR A , or NR A R B or alkyl substituted with -NH2; R A is alkyl, alkenyl, alkynyl, each of which is optionally substituted with -OH or alkoxy; and R B is alkyl, alkenyl, or alkynyl.

[0148] The two starting materials (0.16 mMol) were mixed in 1 mL of deionized water in a 1:1 molar ratio and heated to 140° C. in a microwave for 10 minutes. The resulting product was purified by reversed-phase column chromatography to obtain the product. The product was characterized by LC-MS. The desired mass was observed as shown in Table E. Scheme IV: Deprotection of the amine [ka] During the ceremony: n is 1, 2, or 3; m is 1, 2, 3, 4, or 5; q is 1 or 2; and Each R 1 is independently halo, alkyl, or haloalkyl.

[0149] The starting material (1.11 mMol) was dissolved in 3 mL of 4N hydrochloric acid in dioxane and stirred overnight. The solvent was removed by evaporation and the final product was precipitated from diethyl ether, filtered and dried to obtain the product as the di-hydrochloride salt. The product was characterized by LC-MS. The desired mass was observed as shown in Table E.

[0150] [Table 8]

[0151]

Table 9

[0152]

Table 10

[0153]

Table 11

Claims

1. Formula (I) 【Chemical 1】 (wherein: n is 1, 2, or 3; p is 1, 2, 3, 4, or 5; m is 1, 2, 3, 4, or 5; Each R １ is independently halo, alkyl, or haloalkyl; R ２ is -H, alkyl, alkenyl, or alkynyl; R ３ is -H, alkyl, alkenyl, or alkynyl; R ４ is alkyl, alkenyl, alkynyl, cycloalkyl, or heterocyclyl, each of which is optionally substituted with one or more R ５ groups; or R ３ and R ４ together form a heterocyclyl, said heterocyclyl being optionally substituted with one or more R ５ groups, R ５ is aryl, heteroaryl, alkyl, NH ２ , NHR Ａ , or NR Ａ R Ｂ , or alkyl-NH ２ ; R Ａ is alkyl, alkenyl, or alkynyl, each of which is optionally substituted with -OH or alkoxy; and R Ｂ is alkyl, alkenyl, or alkynyl) ) of the compound or its stereoisomer or pharmaceutically acceptable salt.

2. R ３ and R ４ together form a heterocyclyl, said heterocyclyl optionally being substituted with one or more R ５ groups, a compound according to claim 1.

3. Formula (II): 【Chemical 2】 (wherein: n is 1, 2, or 3; p is 1, 2, 3, 4, or 5; m is 1, 2, 3, 4, or 5; q is 1, 2, or 3; Each R １ is independently halo, alkyl, or haloalkyl; R ２ is hydrogen, C １－６ alkyl, C １－６ alkenyl, or C １－６ alkynyl; Ring A is heterocyclyl; R ５ is aryl, heteroaryl, alky, NH ２ , NHR Ａ , or NR Ａ R Ｂ ; R Ａ is C １－６ alkyl, C １－６ alkenyl, or C １－６ alkynyl, each of which is optionally substituted with -OH or C １－６ alkoxy; and R Ｂ is C １－６ alkyl, C １－６ alkenyl, or C １－６ alkynyl, each of which is optionally substituted with -OH or C １－６ alkoxy). ) having the structure of the compound according to claim 1 or 2 or its stereoisomer or pharmaceutically acceptable salt.

4. The Ring A is 5- to 8-membered heterocyclyl, optionally having 1, 2, or 3 heteroatoms selected from N, O, or S in addition to the ring N shown in formula (II), the compound according to claim 3.

5. The Ring A is 5- to 6-membered heterocyclyl, the compound according to claim 3.

6. R ５ is heteroaryl, NH ２ , NHR Ａ , or NR Ａ R Ｂ ; R Ａ is optionally C １－６ alkyl substituted with C １－６ alkoxy; and R Ｂ is C １－６ alkyl, and the compound according to claim 3.

7. p is 1 or 2, the compound according to claim 3.

8. m is 1 or 2, the compound according to claim 3.

9. q is 1, the compound according to claim 3.

10. Formula (III): 【Chemical 3】 (wherein: n is 1 or 2; p is 1, 2, or 3; m is 1, 2, or 3; q is 1; Each R １ is independently halo or haloalkyl; Ring A is 5-membered or 6-membered heterocyclyl; R ５ is a heteroaryl or C ２ alkyl substituted with -NH Ａ , -NHR Ａ , or -NR Ｂ R １－６ ; R Ａ is optionally C １－６ alkyl substituted with alkoxy; and １－６ is alkyl; and R Ｂ is C １－６ alkyl) ) having the structure of the compound according to claim 1 or its stereoisomer or pharmaceutically acceptable salt.

11. Each R １ is Cl, and the compound according to any one of claims 1, 2, and 10.

12. Formula (IV): 【Chemical Formula 4】 (wherein: n is 1 or 2; q is 1; Each R １ is Cl or fluoroalkyl; R ５ is alkyl, -NH ２ or -NH ２ , -NHR Ａ or NR Ａ R Ｂ ; R Ａ is optionally C １－６ alkyl substituted with C １－６ alkoxy; R Ｂ is C １－６ alkyl) ) having the structure of the compound according to any one of claims 1, 2, and 10 or its stereoisomer or pharmaceutically acceptable salt.

13. R ５ is alkyl-NH ２ or -NH ２ and is the compound according to claim 12.

14. The following structure: 【Chemical Formula 5】 having the structure of the compound according to claim 12 or its stereoisomer or pharmaceutically acceptable salt.

15. The following structure: 【Chemical Formula 6】 having the structure of the compound according to claim 14 or its pharmaceutically acceptable salt.

16. Formula (V): 【Chemical Formula 7】 (wherein: n is 1 or 2; q is 1 or 2; Each R １ is Cl or fluoroalkyl; R ５ is C １－６ alkyl, heteroaryl, NH ２ , NHR Ａ , or NR Ａ R Ｂ ; R Ａ is optionally C １－６ alkyl substituted with C １－６ alkoxy; R Ｂ is C １－６ alkyl) ) having the structure of the compound according to any one of claims 1, 2, and 10 or its stereoisomer or pharmaceutically acceptable salt.

17. R ５ The compound according to claim 16, wherein R is a 5- to 7-membered heteroaryl having one, two, or three heteroatoms selected from N or S.

18. R ５ The compound according to claim 16, wherein R is a 5-membered heteroaryl having one or two N heteroatoms.

19. R ５ is the compound according to claim 16, which is imidazolyl.

20. The following structure: 【Chemical Formula 8】 having the structure of the compound according to claim 16 or its pharmaceutically acceptable salt.

21. R ３ The compound according to claim 1, wherein R is -H or alkyl.

22. R ４ is alkyl substituted with R ５ and; and R ５ is NH ２ and is the compound according to claim 1 or 21.

23. R ４ The compound according to claim 1 or 21, wherein R is heterocyclyl.

24. R ４ is the compound according to claim 23, which is a 5,3-fused heterocyclyl.

25. R ２ The compound according to claim 21, wherein R is H.

26. n is 1 or 2, and R １ is halo, the compound according to claim 21.

27. The compound according to claim 21, wherein m is 1 or 2.

28. The compound according to claim 21, wherein p is 1 or 2.

29. The following structure: 【Chemical Formula 9】 The compound according to claim 21 or a pharmaceutically acceptable salt thereof having one of the above.

30. The following structure: 【Chemical 10】 【Chemical Formula 11】 The compound according to claim 1 or a pharmaceutically acceptable salt thereof having one of the above.

31. A pharmaceutical composition comprising a therapeutically effective amount of the compound according to any one of claims 1, 2, 10 and 30 and one or more pharmaceutically acceptable excipients.

32. A method for treating a bacterial infection in a subject in need thereof, comprising administering to the subject a pharmaceutically acceptable amount of the compound according to any one of claims 1, 2, 10 and 30.

33. The method according to claim 32, wherein the bacterial infection is caused by an intracellular pathogen.

34. The method according to claim 32, wherein the bacterial infection is caused by Gram-negative bacteria.

35. The bacterial infection is one or more Salmonella sp., Acinetobacter sp., Actinobacillus The method according to claim 34, caused by a species of Sphingomonas, Aeromonas sp., Bacteroides sp., Bordetella sp., Brucella sp., Burkholderia sp., Prevotella sp., Porphyromonas sp., Campylobacter sp., Citrobacter sp., Edwardsiella sp., Eikenella sp., Enterobacter sp., Escherichia sp., Francisella sp., Haemophilus sp., Helicobacter sp., Kingella sp., Klebsiella sp., Legionella sp., Moraxella sp., Morganella sp., Neisseria sp., Pasteurella sp., Plesiomonas sp., Proteus sp., Providencia sp., Pseudomonas sp., Salmonella sp., Serratia sp., Shigella sp., Stenotrophomonas sp., Streptobacillus sp., Vibrio sp., Yersinia sp., Chlamydophila sp., Rickettsia sp., Coxiella sp., Ehrlichia sp., or Bartonella sp.

36. The method according to claim 35, wherein the bacterial infection is caused by one or more Salmonella species.

37. The method according to claim 36, wherein the Salmonella sp. is S. enterica serovar Typhimurium.

38. The method according to claim 32, wherein the bacterial infection is caused by Escherichia coli, Klebsiella pneumonia, or Enterobacter cloacea.

39. The compound is: 【Chemical Formula 12】 【Chemical 13】 or a pharmaceutically acceptable salt thereof, according to the method of claim 32.

40. The method according to claim 32, wherein the bacterial infection is resistant to one or more antibiotics.

41. The method according to claim 32, further comprising administering one or more antibiotics.

42. The antibiotic is macrolide, tetracycline, fluoroquinolone, penicillin, cephalosporin, aminoglycoside, sulfonamide, β-lactam, tetracycline, trimethoprim-sulfamethoxazole, chloramphenicol, or lincosamide, according to the method of claim 40.

43. A method for treating a bacterial infection in a subject in need thereof, comprising administering to the subject a pharmaceutically acceptable amount of the compound according to any one of claims 1, 2, 10, and 30 and one or more antibiotics.

44. The method according to claim 43, wherein the bacterial infection is caused by an intracellular pathogen.

45. The method according to claim 43, wherein the bacterial infection is caused by Gram-negative bacteria.

46. The bacterial infection is one or more Salmonella sp., Acinetobacter sp., Actinobacillus The method according to claim 45, caused by a species of Sphingomonas, Aeromonas sp., Bacteroide sp., Bordetella sp., Brucella sp., Burkholderia sp., Prevotella sp., Porphyromonas sp., Campylobacter sp., Citrobacter sp., Edwarsiella sp., Eikenella sp., Enterobacter sp., Escherichia sp., Francisella sp., Haemophilus sp., Helicobacter sp., Kingella sp., Klebsiella sp., Legionella sp., Moraxella sp., Morganella sp., Neisseria sp., Pasteurella sp., Plesiomonas sp., Proteus sp., Providencia sp., Pseudomonas sp., Salmonella sp., Serratia sp., Shigella sp., Stenotrophomonas sp., Streptobacillus sp., Vibrio sp., Yersinia sp., Chlamydophila sp., Ricketsia sp., Coxiella sp., Ehrlichia sp., or Bartonella sp.

47. The method according to claim 46, wherein the bacterial infection is caused by one or more Salmonella species.

48. The method according to claim 47, wherein the Salmonella sp. is S. enterica serovar Typhimurium.

49. The method according to claim 45, wherein the bacterial infection is caused by Escherichia coli, Klebsiella pneumonia, or Enterobacter cloacea.

50. The compound is: 【Chemical 14】 【Chemical Formula 15】 The compound or a pharmaceutically acceptable salt thereof according to claim 42.

51. The method according to claim 42, wherein the antibiotic is a macrolide, tetracycline, fluoroquinolone, penicillin, cephalosporin, aminoglycoside, sulfonamide, β-lactam, tetracycline, trimethoprim-sulfamethoxazole, chloramphenicol, or lincosamide.

52. A method of inhibiting a bacterial efflux pump in a subject having a bacterial infection, comprising administering to the subject a pharmaceutically acceptable amount of the compound according to any one of claims 1, 2, 10, and 30.

53. The method according to claim 52, wherein the bacterial infection is caused by an intracellular pathogen.

54. The method according to claim 53, wherein the bacterial infection is caused by a Gram-negative bacterium.

55. The bacterial infection is one or more Salmonella sp., Acinetobacter sp., Actinobacillus sp. The method according to claim 54, caused by a species of Actinobacillus (Actinobacillus sp.), a species of Aeromonas (Aeromonas sp.), a species of Bacteroides (Bacteroide sp.), a species of Bordetella (Bordetella sp.), a species of Brucella (Brucella sp.), a species of Burkholderia (Burkholderia sp.), a species of Prevotella (Prevotella sp.), a species of Porphyromonas (Porphyromonas sp.), a species of Campylobacter (Campylobacter sp.), a species of Citrobacter (Citrobacter sp.), a species of Edwardsiella (Edwarsiella sp.), a species of Eikenella (Eikenella sp.), a species of Enterobacter (Enterobacter sp.), a species of Escherichia (Escherichia sp.), a species of Francisella (Francisella sp.), a species of Haemophilus (Haemophilus sp.), a species of Helicobacter (Helicobacter sp.), a species of Kingella (Kingella sp.), a species of Klebsiella (Klebsiella sp.), a species of Legionella (Legionella sp.), a species of Moraxella (Moraxella sp.), a species of Morganella (Morganella sp.), a species of Neisseria (Neisseria sp.), a species of Pasteurella (Pasteurella sp.), a species of Plesiomonas (Plesiomonas sp.), a species of Proteus (Proteus sp.), a species of Providencia (Providencia sp.), a species of Pseudomonas (Pseudomonas sp.), a species of Salmonella (Salmonella sp.), a species of Serratia (Serratia sp.), a species of Shigella (Shigella sp.), a species of Stenotrophomonas (Stenotrophomonas sp.), a species of Streptobacillus (Streptobacillus sp.), a species of Vibrio (Vibrio sp.), a species of Yersinia (Yersinia sp.), a species of Chlamydophila (Chlamydophila sp.), a species of Rickettsia (Ricketsia sp.), a species of Coxiella (Coxiella sp.), a species of Ehrlichia (Ehrlichia sp.), or a species of Bartonella (Bartonella sp.).

56. The method according to claim 55, wherein the bacterial infection is caused by one or more Salmonella species.

57. The method according to claim 56, wherein the Salmonella sp. is S. enterica serovar Typhimurium.

58. The compound is: 【Chemical 16】 【Chemical 17】 The compound or a pharmaceutically acceptable salt thereof according to claim 52.

59. The method according to claim 52, wherein the bacterial infection is resistant to one or more antibiotics.

60. The method according to claim 52, further comprising administering one or more antibiotics.

61. The method according to claim 59, wherein the antibiotic is macrolide, tetracycline, fluoroquinolone, penicillin, cephalosporin, aminoglycoside, sulfonamide, β-lactam, tetracycline, trimethoprim-sulfamethoxazole, chloramphenicol, or lincosamide.

62. A method for enhancing the susceptibility of Gram-negative bacteria to an antibiotic, comprising administering a compound according to any one of claims 1, 2, 10, and 30 in combination with the antibiotic.

63. A method for reversing or reducing the antibiotic resistance of antibiotic-resistant Gram-negative bacteria, comprising administering a compound according to any one of claims 1, 2, 10, and 30 in combination with the antibiotic.

64. The method according to claim 62, wherein the antibiotic is macrolide, tetracycline, fluoroquinolone, penicillin, cephalosporin, aminoglycoside, sulfonamide, β-lactam, tetracycline, trimethoprim-sulfamethoxazole, chloramphenicol, or lincosamide.

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