Novel depsipeptides and uses thereof

Abstract

To provide novel depsipeptides and uses thereof.SOLUTION: The present invention relates generally to novel depsipeptides, to methods for the preparation of these novel depsipeptides, to pharmaceutical compositions comprising the novel depsipeptides; and to methods of using the novel depsipeptides to treat or inhibit various disorders. The present invention provides a compound of the present invention or an enantiomer, diastereomer, tautomer, or pharmaceutically acceptable salt thereof. In specific embodiments, the compound of the invention may be a natural product of bacterial isolate ISO1862.SELECTED DRAWING: None

Description

[Technical field]

[0001] Related Applications This application claims the benefit of U.S. Provisional Application No. 61 / 732,894, filed December 3, 2012, the entire contents of which are incorporated herein by reference. [Background technology]

[0002] 2. Background of the Invention One of the great achievements of medicine in recent years is the development and successful use of antibacterial agents against disease-causing microorganisms. Antibacterial agents have saved many lives and reduced the complications of many diseases and infectious diseases. However, currently available antibacterial agents are no longer as effective as they once were.

[0003] Over time, many microorganisms have developed to evade the antibacterial effects of known antibacterial agents, and in recent years, infections caused by microorganisms resistant to multiple antibacterial agents have increased worldwide. As global travel has become more accessible and easier, the rapid spread of drug-resistant microorganisms around the world has become a serious problem. In the community, microbial resistance can result from nosocomial infections of drug-resistant pathogens (e.g., methicillin-resistant Staphylococcus aureus (MRSA), vancomycin-resistant Enterococci (VRE)), resistance emergence due to antibiotic use in the community (e.g., penicillin-resistant and quinolone-resistant Neisseria gonorrheae), infection with resistant pathogens as a result of travel (e.g., antibiotic-resistant Shigella), or infection with resistant pathogens as a result of antibacterial use in animals and subsequent transmission of the resistant pathogens to humans (e.g., antibiotic-resistant Salmonella). Nosocomial antibiotic resistance, usually resulting from antibiotic overuse, is a serious problem for MRSA, VRE and multidrug-resistant Gram-negative bacilli (MDR-GNB) (e.g., Enterobacter, Klebsiella, Serratia, Citrobacter, Pseudomonas and E. coli). In particular, bacterial catheter-associated bloodstream infections and skin and soft tissue infections (SSTIs) are becoming a major problem.

[0004] Bacteria, viruses, fungi and parasites are all developing resistance to known antimicrobial agents. Resistance usually results from three mechanisms: (i) changes in the drug's target, resulting in poor binding of the antimicrobial agent and therefore reduced effectiveness in controlling infections; (ii) impaired drug penetration or active efflux, resulting in reduced access of the drug to its target; and (iii) enzymatic inactivation of the drug by enzymes produced by the microorganism. Antimicrobial resistance allows microorganisms to survive and thus to eliminate microbial infections from the body. This increased difficulty in fighting microbial infections has increased the risk of infectious disease outbreaks in hospitals and other practice settings. Diseases such as tuberculosis, malaria, gonorrhea and childhood ear infections are now more difficult to treat than they were just a few decades ago. Drug resistance is a major problem for hospitals with critically ill patients who would otherwise be susceptible to infection. Unfortunately, these patients are chosen to use large amounts of antibiotics to alter the microorganisms that develop drug resistance. These drug-resistant bacteria are resistant to the most powerful antibiotics and continue to take their toll on vulnerable hospitalized patients: it has been reported that 5 to 10 percent of hospitalized patients acquire an infection during their stay, and that this risk has been steadily increasing in recent decades.

[0005] Regarding these issues, novel drugs are being developed to combat the problems of microbial infections and increasing drug resistance. There is an increasing need for antibacterial drugs, and as pathogens develop resistance to available drugs, a renewed focus on antibacterial drug discovery is crucial.

[0006] So far, synthetic compounds have not been able to replace natural antibiotics, and even efforts combining combinatorial synthesis, high-throughput screening, advanced medicinal chemistry, genomics and proteomics, and rational drug design have not yet produced a novel class of broad-spectrum compounds. The problem in obtaining new synthetic antibiotics may be partly related to the fact that synthetic antibiotics are uniformly pumped through the bacterial outer membrane barrier by multidrug resistance pumps (MDR). The bacterial outer membrane is a barrier to amphipathic compounds (essentially all drugs are amphipathic compounds), and the MDR expels drugs through this barrier. Although evolution has produced antibiotics that are largely able to circumvent this dual barrier / expulsion mechanism, synthetic compounds have, for the most part, been uniformly unsuccessful. Summary of the Invention [Means for solving the problem]

[0007] This application is directed, at least in part, to novel depsipeptides that are useful in the treatment of a number of disorders, including microbial infections.

[0008] In one embodiment, the present invention provides an isolated compound of formula (I) [ka] or an enantiomer, diastereomer, tautomer, or pharma- ceutically acceptable salt thereof wherein each stereocenter (indicated with an "*") may be in either the R or S configuration. Regarding.

[0009] In some embodiments, the compound of formula (I) is a natural product of an isolated bacterial species. For example, in some embodiments, the compound of formula (I) is a natural product of an isolated bacterial isolate ISO18629. In some embodiments, the compound of formula (I) can be produced from a bacterial species. For example, in some embodiments, the compound of formula (I) can be produced from the bacterial isolate ISO18629.

[0010] In another embodiment, the present invention provides an isolated compound of formula (II) [ka] or a tautomer or a pharma- ceutically acceptable salt thereof.

[0011] In some embodiments, the compound of formula (II) is a natural product of an isolated bacterial species. For example, in some embodiments, the compound of formula (II) is a natural product of an isolated bacterial isolate ISO18629. In some embodiments, the compound of formula (II) can be produced from a bacterial species. For example, in some embodiments, the compound of formula (II) can be produced from the bacterial isolate ISO18629.

[0012] In some embodiments, the compound of formula (I) has at least 10 NMR spectroscopy images at chemical shifts (+ / - 0.2 ppm) in DMSO-d6 selected from 36.3 ppm, 36.5 ppm, 36.9 ppm, 37.4 ppm, 52.1 ppm, 52.2 ppm, 52.7 ppm, 53.5 ppm, 55.7 ppm, 56.1 ppm, 56.4 ppm, 56.7 ppm, 57.3 ppm, 57.8 ppm, 57.9 ppm, 61.8 ppm, and 71.1 ppm. 13 In some embodiments, the compound of formula (I) is characterized by a C nuclear magnetic resonance peak at chemical shifts (+ / - 0.2 ppm) in DMSO-d6 of 36.3 ppm, 36.5 ppm, 36.9 ppm, 37.4 ppm, 52.1 ppm, 52.2 ppm, 52.7 ppm, 53.5 ppm, 55.7 ppm, 56.1 ppm, 56.4 ppm, 56.7 ppm, 57.3 ppm, 57.8 ppm, 57.9 ppm, 61.8 ppm, and 71.1 ppm. 13 It is characterized by a C nuclear magnetic resonance peak.

[0013] In yet another embodiment, the present invention relates to a compound of formula (I) characterized by at least one of a molecular weight of about 1242.47 g / mol, a proton nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 1, a carbon-13 nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 2, a COSY nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 3, a DEPT-135 nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 4, an HSQC nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 5, or a HMBC nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 6. In yet another embodiment, the present teachings provide a method for the preparation of a compound of formula (I), characterized by a molecular weight of about 1242.47 g / mol, a proton nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 1, a carbon-13 nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 2, a COSY nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 3, a DEPT-135 nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 4, an HSQC nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 5, and a HMBC nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 6. Concerning compounds.

[0014] In one embodiment, the present invention provides an isolated compound of formula (III) [ka] or a tautomer or a pharma- ceutically acceptable salt thereof.

[0015] Each of R1 to R6 is independently hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, aryl, C(=O)R a and S(=O)2R b Each R a is independently hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, or aryl; each R b is independently alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, or aryl.

[0016] Also, any carbon or hydrogen can be 13 C or 2 It may be replaced by H.

[0017] In one embodiment, the present invention provides an isolated compound of formula (IV) [ka] wherein X is NH, O or S. or a tautomer or a pharma- ceutically acceptable salt thereof.

[0018] In yet another embodiment, the invention relates to a pharmaceutical composition comprising a compound described herein, e.g., a compound of Formula (I), (II), (III) or (IV), and a pharma- ceutically acceptable excipient, carrier or diluent. In some embodiments, a pharmaceutical composition comprising any of the compounds described herein can further comprise an agent selected from the group consisting of an antibiotic, an antifungal agent, an antiviral agent, an antiprotozoan agent, an antiparasitic agent, an antineoplastic agent, an immunomodulatory agent, an antihypercholesterolemic agent, and combinations thereof.

[0019] In yet another embodiment, the present invention provides a compound of formula (I) [ka] or an enantiomer, diastereomer, tautomer, or pharma- ceutically acceptable salt thereof, comprising culturing bacterial isolate ISO18629 in a culture comprising assimilable sources of carbon, nitrogen, and inorganic salts under aerobic conditions, thereby producing the compound of formula (I). In some embodiments, the method further comprises isolating the compound of formula (I) from the culture.

[0020] In yet another embodiment, the present invention relates to a compound of formula (I) prepared according to the methods described herein.

[0021] In yet another embodiment, the present invention relates to a compound of formula (II) [ka] or a tautomer or a pharma- ceutically acceptable salt thereof, comprising culturing bacterial isolate ISO18629 in a culture comprising assimilable sources of carbon, nitrogen and inorganic salts under aerobic conditions, thereby producing the compound of formula (II). In some embodiments, the method further comprises isolating the compound of formula (II) from the culture.

[0022] In yet another embodiment, the present invention relates to a compound of formula (II) prepared according to the methods described herein.

[0023] In yet another embodiment, the present invention relates to a compound of formula (III) [ka] or a tautomer or a pharma- ceutically acceptable salt thereof where the variables are described above. In some embodiments, the present invention relates to a method for producing a compound of formula (III), comprising culturing the bacterial isolate ISO18629 in a culture comprising assimilable sources of carbon, nitrogen and inorganic salts under aerobic conditions, thereby producing a compound of formula (III): 13 C or 2 H is under aerobic conditions. 13 C and / or 2 H-labeled carbon, nitrogen and inorganic salts The bacterial isolate ISO18629 was cultivated in a culture medium containing an assimilable source of 13 C or 2 The compound of formula (III) can be incorporated by forming a compound of formula (III) containing H. For example, 13 C and / or 2H-labeled amino acids may also be used. In some embodiments, the method further comprises isolating the compound of formula (III) from the culture.

[0024] In yet another embodiment, the present invention relates to a compound of formula (III) prepared according to the methods described herein.

[0025] In yet another embodiment, the present invention relates to a compound of formula (IV) [ka] or a tautomer or a pharma- ceutically acceptable salt thereof wherein X is NH, O or S. The present invention relates to a method for producing a compound of formula (IV), comprising culturing the bacterial isolate ISO18629 in a culture comprising assimilable sources of carbon, nitrogen and inorganic salts under aerobic conditions, thereby producing a compound of formula (IV). In some embodiments, the method further comprises isolating the compound of formula (IV) from the culture.

[0026] In yet another embodiment, the present invention relates to a compound of formula (IV) prepared according to the methods described herein.

[0027] In yet another embodiment, the present invention relates to an isolated culture comprising a bacterial species having an ISO 18629 identifying characteristic.

[0028] The present invention also relates to a method of treating a disorder in a subject, e.g., a human, in need of such treatment. The method comprises administering to the subject a therapeutically effective amount of a compound, e.g., a compound of formula (I), (II), (III) or (IV), as described herein, thereby treating the disorder in the subject. In some embodiments, the subject is a mammal, a human, an animal or a plant. In a particular embodiment, the subject is a human. In a particular embodiment, the disorder is caused by an external pathogen, such as, but not limited to, a bacterium, a fungus, a virus, a protozoan, a helminth, a parasite, and combinations thereof.

[0029] In certain embodiments, the infectious agent is a bacterium. In one embodiment, the bacterium is a gram-positive bacterium. Non-limiting examples of gram-positive bacteria include Streptococcus, Staphylococcus, Enterococcus, Corynebacteria, Listeria, Bacillus, Erysipelothrix and Actinomycetes.In some embodiments, the compounds of Formula (I), (II), (III) or (IV) are used to treat infections caused by one or more of the following: Helicobacter pylori, Legionella pneumophilia, Mycobacterium tuberculosis, Mycobacterium avium, Mycobacterium intracellulare, Mycobacterium kansaii, Mycobacterium gordonae, Mycobacteria sporozoites, Staphylococcus aureus, Staphylococcus epidermidis, Neisseria gonorrhoeae, Neisseria meningitidis, Listeria monocytogenes, Streptococcus pyogenes (Group A Streptococcus), Streptococcus agalactiae pyogenes (Group B Streptococcus), Streptococcus dysgalactia, Streptococcus faecalis, Streptococcus bovis, Streptococcus pneumoniae, pathogenic Campylobacter sporozoites, Enterococcus sporozoites, Haemophilus influenzae, Pseudomonas aeruginosa, Bacillus anthracis, Bacillus subtilis, Escherichia coli, Corynebacterium diphtheriae, Corynebacterium jeikeium, Corynebacterium sporozoites, Erysipelothrix rhusiopathiae, Clostridium perfringens, Clostridium tetani, Clostridium difficile, Enterobacter aerogenes, Klebsiella pneumoniae, Pasturella multocida, Bacteroides thetaiotamicron, Bacteroides uniformis, Bacteroides. vulgatus, Fusobacterium nucleatum, Streptobacillus moniliformis, Leptospira and Actinomyces israelli. In particular, the gram-positive bacterium is Bacillus anthracis.

[0030] In another embodiment, the bacterium is a gram-negative bacterium. Non-limiting examples of Gram-negative bacteria include Helicobacter pylori, Legionella pneumophilia, Neisseria gonorrhoeae, Neisseria meningitidis, pathogenic Campylobacter sporozoites, Haemophilus influenzae, Pseudomonas aeruginosa, Enterobacter aerogenes, Enterobacter cloacae, Klebsiella pneumoniae, Klebsiella oxytoca, Pasteurella multocida, Bacteroides sporozoites, Bacteriodes fragilis, Bacteriodes thetaiotaomicron, Bacteriodes uniformis, Bacteriodes vulgatus Fusobacterium nucleatum, Streptobacillus moniliformis, Leptospira, Escherichia coli, Salmonella enterica, Salmonella salamae, Salmonella arizonae, Salmonella diarizonae, Salmonella houtenae, Salmonella bongori, Salmonella indica, Salmonella Enteritidis, Salmonella typhi and Citrobacter freundii.

[0031] In other embodiments, the depsipeptide compounds described herein may be useful in treating viral disorders. Non-limiting examples of infectious viruses that can be treated by the compounds of Formula (I), (II), (III) or (IV) include Retroviridae (e.g., human immunodeficiency viruses, such as HIV-1 (also called HTLV-III, LAV or HTLV-III / LAV) or HIV-III; and other isolates, such as HIV-LP; Picornaviridae (e.g., poliovirus, Hepatitis A virus, enterovirus, human coxsackievirus, rhinovirus, echovirus); Caliciviridae (e.g., strains that cause gastroenteritis); Togaviridae (e.g., equine encephalitis virus, rubella virus); Flaviviridae (e.g., dengue virus, encephalitis virus, yellow fever virus); Coronaviridae (e.g., coronavirus, severe acute respiratory syndrome (SARS) virus); Rhabdoviridae (e.g., vesicular stomatitis virus, rabies virus); Filoviridae (e.g., Ebola virus); Paramyxoviridae (e.g., Parainfluenza virus, mumps virus, measles virus, respiratory syncytial virus; Orthomyxoviridae (e.g. influenza virus); Bunyaviridae (e.g. hantavirus, bungavirus, phlebovirus and nairovirus); Arenaviridae (hemorrhagic fever viruses); Reoviridae (e.g. reovirus, orbiviurses and rotavirus); Birnaviridae; Hepadnaviridae (e.g. hepatitis B virus); Parvoviridae (parvovirus); Papovaviridae (papillomavirus, polyomavirus); Adenoviridae (most adenoviruses); Herpesviridae (e.g. herpes simplex virus (HSV) 1 and 2, varicella zoster virus, cytomegalovirus (CMV), herpes virus); Poxviridae (e.g. smallpox virus, vaccinia virus, pox virus); and Iridoviridae (e.g. African swine fever virus);and unclassified viruses (e.g., agents of spongiform encephalopathies, agents of delta hepatitis (thought to be a defective satellite of hepatitis B virus), non-A, non-B hepatitis agents (Class 1 = endogenous, Class 2 = parentally transmitted, i.e., hepatitis C); Norwalk and related viruses, and astroviruses). In certain embodiments, the compounds of formula (I), (II), (III) or (IV) are used to treat influenza virus, human immunodeficiency virus, or herpes simplex virus;

[0032] In yet another embodiment, the depsipeptide compounds described herein are useful for treating disorders caused by protozoa.Non-limiting examples of protozoa that can be inhibited by the compounds of formula (I), (II), (III) or (IV) include, but are not limited to, Trichomonas vaginalis, Giardia lamblia, Entamoeba histolytica, Balantidium coli, Cryptosporidium parvum and Isospora belli, Trypansoma cruzi, Trypanosoma gambiense, Leishmania donovani and Naegleria fowleri.

[0033] In certain embodiments, the depsipeptide compounds described herein are useful for treating disorders caused by helminths. Non-limiting examples of helminths that can be inhibited by the compounds of formula (I), (II), (III) or (IV) include, but are not limited to, Schistosoma mansoni, Schistosoma cercariae, Schistosoma japonicum, Schistosoma mekongi, Schistosoma hematobium, Ascaris lumbricoides, Strongyloides stercoralis, Echinococcus granulosus, Echinococcus mu, and Schistosoma granulosus. ltilocularis, Angiostrongylus cantonensis, Angiostrongylus constaricensis, Fasciolopis buski, Capillaria philippinensis, Paragonimus westermani, Ancylostoma dudodenale, Necator americanus, Trichinella spiralis, Wuchereria bancrofti, Brugia malayi and Brugia timori, Toxocara canis, Toxocara cati, Toxocara vitulorum, Caenorhabiditis elegans and Anisakis species.

[0034] In some embodiments, the depsipeptide compounds described herein are useful for treating disorders caused by parasites. Non-limiting examples of parasites that can be inhibited by the compounds of formula (I), (II), (III) or (IV) include, but are not limited to, Plasmodium falciparum, Plasmodium yoelli, Hymenolepis nana, Clonorchis sinensis, Loa loa, Paragonimus westermani, Fasciola hepatica, and Toxoplasma gondii. In a particular embodiment, the parasite is a malaria parasite.

[0035] In some embodiments, the depsipeptide compounds of formula (I), (II), (III) or (IV) are useful for treating disorders caused by fungi. Non-limiting examples of fungi that can be inhibited by the compounds of formula (I), (II), (III) or (IV) include, but are not limited to, Cryptococcus neoformans, Histoplasma capsulatum, Coccidioides immitis, Blastomyces dermatitidis, Chlamydia trachomatis, Candida albicans, Candida tropicalis, Candida glabrata, Candida krusei, Candida parapsilosis, Candida dubliniensis, Candida lusitaniae, Epidermophyton floccosum, Microsporum audouinii, Microsporum canis, Microsporum canis var. distortum, Microsporum cookei, Microsporum equinum, Microsporum ferrugineum, Microsporum fulvum, Microsporum gallinae, Microsporum gypseum, Microsporum nanum, Microsporum persicolor, Trichophyton ajelloi, Trichophyton concentricum, Trichophyton equinum, Trichophyton flavescens, Trichophyton gloriae, Trichophyton megnini, Trichophyton mentagrophytes var. erinacei, Trichophyton mentagrophytes var. interdigitale, Trichophyton phaseoliforme, Trichophyton rubrum, Trichophyton rubrum downy strain, Trichophyton rubrum granular strain, Trichophyton schoenleinii, Trichophyton simii, Trichophyton soudanense, Trichophyton terrestre, Trichophyton tonsurans, Trichophyton vanbreuseghemii, Trichophyton verrucosum, Trichophyton violaceum, Trichophyton yaoundei, Aspergillus fumigatus, Aspergillus flav us and Aspergillus clavatus. In yet another embodiment, the present invention relates to a method of inhibiting the growth of an infectious agent, comprising contacting the agent with a compound described herein, e.g., a compound of Formula (I), (II), (III) or (IV), thereby inhibiting the growth of the infectious agent.

[0036] In one particular embodiment, the infectious agent is cultured in vitro.

[0037] The present invention is further illustrated by the following detailed description and figures. In certain embodiments, for example, the following are provided: (Item 1) Isolated Compound of Formula (I) [ka] or an enantiomer, diastereomer, tautomer, or pharma- ceutically acceptable salt thereof [wherein each stereocenter (indicated with an "*") may be in either the R or S configuration]. (Item 2) 2. The compound according to item 1, which is a natural product of the bacterial isolate ISO18629. (Item 3) 2. The compound according to item 1, which is producible from the bacterial isolate ISO18629. (Item 4) Isolated Compound of Formula (II) [ka] or a tautomer or a pharma- ceutically acceptable salt thereof. (Item 5) At least 10 chemical shifts in DMSO-d6 selected from 36.3 ppm, 36.5 ppm, 36.9 ppm, 37.4 ppm, 52.1 ppm, 52.2 ppm, 52.7 ppm, 53.5 ppm, 55.7 ppm, 56.1 ppm, 56.4 ppm, 56.7 ppm, 57.3 ppm, 57.8 ppm, 57.9 ppm, 61.8 ppm, and 71.1 ppm 13 Item 2. The compound according to item 1, characterized by a C nuclear magnetic resonance peak. (Item 6) Chemical shifts in DMSO-d6 at 36.3 ppm, 36.5 ppm, 36.9 ppm, 37.4 ppm, 52.1 ppm, 52.2 ppm, 52.7 ppm, 53.5 ppm, 55.7 ppm, 56.1 ppm, 56.4 ppm, 56.7 ppm, 57.3 ppm, 57.8 ppm, 57.9 ppm, 61.8 ppm and 71.1 ppm 13 Item 2. The compound according to item 1, characterized by a C nuclear magnetic resonance peak. (Item 7) (a) a molecular weight of about 1242.47 g / mol; (b) A proton nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 1; (c) Carbon-13 nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 2; (d) A COSY nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 3; (e) a DEPT-135 nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 4; (f) an HSQC nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 5; and (e) HMBC nuclear magnetic resonance spectrum substantially identical to that shown in Figure 6. The compound according to item 1, characterized by at least one of the following: (Item 8) (a) a molecular weight of about 1242.47 g / mol; (b) A proton nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 1; (c) Carbon-13 nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 2; (d) A COSY nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 3; (e) a DEPT-135 nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 4; (f) an HSQC nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 5; and (e) HMBC nuclear magnetic resonance spectrum substantially identical to that shown in Figure 6. The compound according to item 1, characterized in that (Item 9) Isolated Compound of Formula (II) [ka] or a tautomer or a pharma- ceutically acceptable salt thereof. (Item 10) Isolated Compound of Formula (III) [ka] or a tautomer or a pharma- ceutically acceptable salt thereof [In the formula, each of R1 to R6 is independently hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, aryl, C(═O)R a and S(=O)2R b Each R a is independently hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, or aryl; each R b is independently alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, or aryl. (Item 11) Isolated Compound of Formula (IV) [ka] wherein X is NH, O or S. or a tautomer or a pharma- ceutically acceptable salt thereof. (Item 12) 12. A pharmaceutical composition comprising a compound according to items 1, 4, 10 or 11, and a pharma- ceutically acceptable excipient, carrier or diluent. (Item 13) 13. The pharmaceutical composition according to item 12, further comprising an agent selected from the group consisting of antibiotics, antifungals, antivirals, antiprotozoans, anthelmintics, antineoplastic agents, immunomodulators, antihypercholesterolemic agents and combinations thereof. (Item 14) Compounds of formula (I) [ka] or an enantiomer, diastereomer, tautomer, or pharma- ceutically acceptable salt thereof, comprising the steps of: The bacterial isolate ISO18629 was cultivated under aerobic conditions in a medium containing assimilable sources of carbon, nitrogen and inorganic salts. thereby producing a compound of formula (I). (Item 15) Compound of formula (II) [ka] or a tautomer or a pharma- ceutically acceptable salt thereof, comprising the steps of: A method comprising culturing the bacterial isolate ISO18629 under aerobic conditions in a medium containing assimilable sources of carbon, nitrogen and inorganic salts, thereby producing the compound of formula (II). (Item 16) Compound of formula (III) [ka] or a tautomer or a pharma- ceutically acceptable salt thereof [In the formula, each of R1 to R6 is independently hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, aryl, C(═O)R a and S(=O)2R b Each R a is independently hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, or aryl; each R b is independently alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, or aryl. 1. A method for producing The bacterial isolate ISO18629 was cultivated under aerobic conditions in a medium containing assimilable sources of carbon, nitrogen and inorganic salts. thereby producing a compound of formula (III). (Item 17) Compound of formula (IV) [ka] wherein X is NH, O or S. or a tautomer or a pharma- ceutically acceptable salt thereof, comprising the steps of: The bacterial isolate ISO18629 was cultivated under aerobic conditions in a medium containing assimilable sources of carbon, nitrogen and inorganic salts. thereby producing a compound of formula (IV). (Item 18) 18. The method according to item 14, 15, 16 or 17, further comprising a step of isolating said compound of formula (I), (II), (III) or (IV). (Item 19) A compound of formula (I), (II), (III) or (IV) prepared according to the method described in items 14, 15, 16 or 17. (Item 20) A method of treating a disorder in a subject in need thereof, comprising administering to the subject an effective amount of a compound of formula (I). [ka] or an enantiomer, diastereomer, tautomer, or pharma- ceutically acceptable salt thereof wherein each stereocenter (denoted with an "*") can be in either the R or S configuration, thereby treating said disorder in said subject. (Item 21) A method of treating a disorder in a subject in need of such treatment, comprising administering to the subject an effective amount of a compound of formula (II) [ka] or a tautomer or a pharma- ceutically acceptable salt thereof, thereby treating said disorder in said subject. (Item 22) A method of treating a disorder in a subject in need thereof, comprising administering to the subject an effective amount of a compound of formula (III) [ka] or a tautomer or a pharma- ceutically acceptable salt thereof, wherein each of R1 to R6 is independently hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, aryl, C(=O)Ra, and S(=O)R b Each R a is independently hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, or aryl; each R b is independently alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, or aryl, thereby treating said disorder in said subject. (Item 23) A method of treating a disorder in a subject in need of such treatment, comprising administering to the subject an effective amount of a compound of formula (IV) [ka] wherein X is NH, O, or S, or a tautomer or pharma- ceutically acceptable salt thereof, thereby treating said disorder in said subject. (Item 24) Item 20. The subject is selected from the group consisting of mammals, humans, animals and plants. 24. The method according to claim 21, 22 or 23. (Item 25) 24. The method of item 20, 21, 22 or 23, wherein the disorder is caused by a pathogen selected from the group consisting of bacteria, fungi, viruses, protozoans, helminths, parasites, and combinations thereof. (Item 26) 26. The method of claim 25, wherein the pathogen is a bacterium. (Item 27) 27. The method of claim 26, wherein the bacterium is a gram-positive bacterium. (Item 28) 28. The method of claim 27, wherein the Gram-positive bacteria is selected from the group consisting of Streptococcus, Staphylococcus, Enterococcus, Corynebacteria, Listeria, Bacillus, Erysipelothrix, Mycobacterium, Clostridium, and Actinomycetales. (Item 29) The gram-positive bacteria include methicillin-susceptible and methicillin-resistant staphylococci (including Staphylococcus aureus, Staphylococcus epidermidis, Staphylococcus haemolyticus, Staphylococcus hominis, Staphylococcus saprophyticus, and coagulase-negative staphylococci), glycopeptide-intermediately susceptible Staphylococcus aureus (GISA), penicillin-susceptible and penicillin-resistant streptococci (including Streptococcus pneumoniae, Streptococcus pyogenes, Streptococcus agalactiae, Streptococcus dysgalactiae, Streptococcus avium, Streptococcus bovis, Streptococcus lactis, Streptococcus sangius, Streptococcus anginosus, Streptococcus intermedius, Streptococcus Enterococci (including vancomycin-susceptible and vancomycin-resistant strains, e.g., Enterococcus constellatus and group C streptococci, group G streptococci, and Viridans streptococci), Enterococci (including vancomycin-susceptible and vancomycin-resistant strains, e.g., Enterococcus faecalis and Enterococcus faecium), Clostridium difficile, Clostridium clostridiiforme, Clostridium innocuum, Clostridium perfringens, Clostridium tetani, Mycobacterium tuberculosis, Mycobacterium avium, Mycobacterium intracellulare, Mycobacterium kansaii, Mycobacterium gordonae, Mycobacteria sporozoites, Listeria monocytogenes, Bacillus subtilis, Bacillus anthracis, Corynebacterium diphtheriae, Corynebacterium28. The method according to item 27, wherein the bacterium is selected from the group consisting of Bacillus subtilis, Corynebacterium sporozoites, Erysipelothrix rhusiopathiae, and Actinomyces israelli. (Item 30) 26. The method according to item 25, wherein the disorder is caused by infection with Bacillus anthracis. (Item 31) 27. The method of claim 26, wherein the bacterium is a gram-negative bacterium. (Item 32) The Gram-negative bacteria include Helicobacter pylori, Legionella pneumophilia, Neisseria gonorrhoeae, Neisseria meningitidis, pathogenic Campylobacter sporozoites, Haemophilus influenzae, Pseudomonas aeruginosa, Enterobacter aerogenes, Enterobacter cloacae, Klebsiella pneumoniae, Klebsiella oxytoca, Pasteurella multocida, Bacteroides sporozoites, Bacteriodes fragilis, Bacteriodes thetaiotaomicron, Bacteriodes uniformis, Bacteriodes vulgatus Fusobacterium nucleatum, Streptobacillus moniliformis, Leptospira, Escherichia coli, Salmonella enterica, Salmonella salamae, Salmonella arizonae, Salmonella 32. The method according to claim 31, wherein the bacterial strain is selected from the group consisting of Salmonella diarizonae, Salmonella houtenae, Salmonella bongori, Salmonella indica, Salmonella Enteritidis, Salmonella typhi and Citrobacter freundii. (Item 33) 26. The method of claim 25, wherein the pathogen is a virus. (Item 34) 34. The method of claim 33, wherein the virus is selected from the group consisting of Retroviridae, Picornaviridae, Caliciviridae, Togaviridae, Flaviviridae, Coronaviridae, Rhabdoviridae, Filoviridae, Paramyxoviridae, Orthomyxoviridae, Bunyaviridae, Arenaviridae, Reoviridae, Birnaviridae, Hepadnaviridae, Parvoviridae, Papovaviridae, Adenoviridae, Herpesviridae, Poxviridae, and Iridoviridae. (Item 35) 34. The method of claim 33, wherein the virus is selected from the group consisting of influenza virus, human immunodeficiency virus, and herpes simplex virus. (Item 36) 26. The method of claim 25, wherein the pathogen is a protozoan. (Item 37) The protozoa is Trichomonas vaginalis, Giardia lamblia, Entamoeba histolytica, Balantidium coli, Cryptosporidium parvum and Isospora belli, Trypansoma cruzi, Trypanosoma gambiense, Leishmania donovani and Naegleria fowleri. (Item 38) 26. The method of claim 25, wherein the pathogen is a helminth. (Item 39) The helminths include Schistosoma mansoni, Schistosoma cercariae, Schistosoma japonicum, Schistosoma mekongi, Schistosoma hematobium, Ascaris lumbricoides, Strongyloides stercoralis, Echinococcus granulosus, Echinococcus multilocularis, Angiostrongylus cantonensis, Angiostrongylus constaricensis, Fasciolopis. buski, Capillaria philippinensis, Par 39. The method according to item 38, wherein the bacterium is selected from the group consisting of: agonimus westermani, Ancylostoma dudodenale, Necator americanus, Trichinella spiralis, Wuchereria bancrofti, Brugia malayi and Brugia timori, Toxocara canis, Toxocara cati, Toxocara vitulorum, Caenorhabiditis elegans and Anisakis species. (Item 40) 26. The method of claim 25, wherein the pathogen is a parasite. (Item 41) 41. The method of claim 40, wherein the parasite is selected from the group consisting of Plasmodium falciparum, Plasmodium yoelli, Hymenolepis nana, Clonorchis sinensis, Loa loa, Paragonimus westermani, Fasciola hepatica and Toxoplasma gondii. (Item 42) 41. The method of claim 40, wherein the parasite is a malarial parasite. (Item 43) 26. The method of claim 25, wherein the pathogen is a fungus. (Item 44) The aforementioned fungi are Cryptococcus neoformans, Histoplasma capsulatum, Coccidioides immitis, Blastomyces dermatitidis, Chlamydia trachomatis, Candida albicans, Candida tropicalis, Candida glabrata, Candida krusei, Candida parapsilosis, Candida dubliniensis, Candida lusitaniae, Epidermophyton floccosum, Microsporum audouinii, Microsporum canis, Microsporum canis var. distortum Microsporum cookei, Microsporum equinum, Microsporum ferrugineum, Microsporum fulvum, Microsporum gallinae, Microsporum gypseum, Microsporum nanum, Microsporum persicolor, Trichophyton ajelloi, Trichophyton concentricum, Trichophyton equinum, Trichophyton flavescens, Trichophyton gloriae, Trichophyton megnini, Trichophyton mentagrophytes var. erinacei, Trichophyton mentagrophytes var. interdigitale, Trichophyton phaseoliforme, Trichophyton rubrum, Trichophyton rubrum downy strain, Trichophyton rubrum granular strain, Trichophyton schoenleinii, Trichophyton simii, Trichophyton soudanense, Trichophyton terrestre, Trichophyton tonsurans, Trichophyton vanbreuseghemii, Trichophyton verrucosum, Trichophyton violaceum, Trichophyton yaoundei, Aspergillus fumigatus, Aspergillus flavus and Aspergillus clavatus. (Item 45) An isolated culture containing a bacterial species with identifying characteristics of an ISO 18629 isolate. (Item 46) A method of inhibiting the growth of an infectious agent, comprising administering to the agent a compound of formula (I) [ka] or an enantiomer, diastereomer, tautomer, or pharma- ceutically acceptable salt thereof wherein each stereocenter (indicated with an "*") may be in either the R or S configuration. thereby inhibiting the growth of the infectious agent. (Item 47) 47. The method of claim 46, wherein the infectious agent is cultured in vitro. (Item 48) A method of inhibiting the growth of an infectious agent, comprising administering to the agent a compound of formula (II) [ka] or a tautomer or pharma- ceutically acceptable salt thereof, thereby inhibiting the growth of the infectious agent. (Item 49) 49. The method of claim 48, wherein the infectious agent is cultured in vitro. (Item 50) A method of inhibiting the growth of an infectious agent, comprising administering to the agent a compound of formula (III) [ka] or a tautomer or a pharma- ceutically acceptable salt thereof [In the formula, each of R1 to R6 is independently hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, aryl, C(═O)R a and S(=O)2R b Each R a is independently hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, or aryl; each R b is independently alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, or aryl. thereby inhibiting the growth of the infectious agent. (Item 51) 51. The method of claim 50, wherein the infectious agent is cultured in vitro. (Item 52) A method of inhibiting the growth of an infectious agent, comprising administering to the agent a compound of formula (IV) [ka] [wherein X is NH, O or S] or a tautomer or a pharma- ceutically acceptable with a salt thereof, thereby inhibiting the growth of the infectious agent. (Item 53) 53. The method of claim 52, wherein the infectious agent is cultured in vitro.

[0038] The above and other objects of the present invention, its various features, as well as the invention itself, may be more fully understood from the following description read in conjunction with the accompanying drawings, in which: [Brief description of the drawings]

[0039] [Figure 1] FIG. 1 is a schematic diagram of the proton nuclear magnetic resonance spectrum (in DMSO-d6) of a compound isolated from a grown strain of ISO18629. [Diagram 2] FIG. 2 is a schematic representation of the carbon-13 nuclear magnetic resonance spectrum (in DMSO-d6) of a compound isolated from a grown strain of ISO18629. [Diagram 3] FIG. 3 is a schematic diagram of the COSY nuclear magnetic resonance spectrum (in DMSO-d6) of a compound isolated from a grown strain of ISO18629. [Figure 4] FIG. 4 is a schematic representation of the DEPT-135 nuclear magnetic resonance spectrum (in DMSO-d6) of a compound isolated from a grown strain of ISO18629. [Diagram 5] FIG. 5 is a schematic diagram of the HSQC nuclear magnetic resonance spectrum (in DMSO-d6) of a compound isolated from a grown strain of ISO18629. [Figure 6]FIG. 6 is a schematic diagram of the HMBC nuclear magnetic resonance spectrum (in DMSO-d6) of a compound isolated from a grown strain of ISO18629. [Figure 7] FIG. 7 is a graph showing the effect of the compound of formula (II) on the synthesis of a polymer. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0040] Detailed Description of the Invention The present invention generally relates to novel depsipeptides, methods for preparing these novel depsipeptides, pharmaceutical compositions containing the novel depsipeptides, and methods of using the novel depsipeptides to treat or inhibit various disorders, such as bacterial infections. The present invention relates to novel antibiotics that have broad activity against many bacterial pathogens, particularly gram-positive pathogens, including strains resistant to other antibiotics. The compounds disclosed herein have favorable bioavailability and low toxicity.

[0041] definition For convenience, some terms used in the present specification, examples and the appended claims are summarized here.Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art to which this invention belongs.The initial definition provided for a group or term in this specification applies to that group or term throughout this specification, individually or as part of another group, unless otherwise specified.

[0042] The articles "a" and "an" are used herein to refer to one or to more than one (i.e., to at least one) of the grammatical object of the article. For example, "an element" means one element or more than one element.

[0043] The term "or" is used herein to refer to, and is used interchangeably with, the term "and / or," unless otherwise clearly indicated by context.

[0044] The term "substantially the same" is used herein to mean that the two objects being compared share at least 90% of certain common characteristics. In certain embodiments, the two objects share at least 95% of certain common characteristics. In certain other embodiments, the two objects share at least 99% of certain common characteristics. do.

[0045] The term "isolated" is used herein to refer to a compound of formula (I), (II), (III) or (IV) that is substantially free of other materials associated with it in its natural environment. For example, an isolated compound may be substantially free of contaminants (e.g., cellular material, contaminants from the cell from which the compound is derived, chemical precursors or other chemicals if chemically synthesized). Substantially free of other materials generally refers to, for example, less than about 30%, or 20%, or 15%, or 10%, or 5%, or 2% (by dry weight) of impurities. In some embodiments, an isolated compound is substantially pure. In some embodiments, a preparation of a compound having less than about 10% (by dry weight) of contaminants or chemical precursors from the cell is considered to be substantially pure. In other embodiments, a preparation of a compound having less than about 5%, less than about 4%, less than about 3%, less than about 2%, or less than about 1% (by dry weight) of contaminants or chemical precursors from the cell is considered to be substantially pure.

[0046] Unless otherwise specified, any heteroatom with unsatisfied valences is presumed to have sufficient hydrogen atoms to satisfy the valences.

[0047] The term "heating" includes, but is not limited to, warming by conventional heating (eg, electrical heating, steam heating, gas heating, etc.), as well as microwave heating.

[0048] The term "pharmaceutically acceptable excipient, carrier or diluent" as used herein means a pharma- ceutically acceptable material, composition or vehicle, such as a liquid or solid filler, diluent, excipient, solvent or encapsulating material, that is involved in carrying or transporting a subject pharmaceutical agent from one organ or part of the body to another organ or part of the body. Each carrier must be "acceptable" in the sense of being compatible with the other ingredients of the formulation and not injurious to the patient.

[0049] The term "treating" with respect to a subject refers to improving at least one symptom of a disorder in the subject. Treating can be curing or ameliorating the disorder or condition.

[0050] The term "disorder" is used herein to refer to, and is used interchangeably with, the terms disease, condition or illness, unless the context clearly indicates otherwise.

[0051] The term "microorganism" is used herein to denote an organism such as a bacterium, virus, protozoa or fungus, particularly an organism that transmits disease.

[0052] The phrase "pharmacologically acceptable" is used herein to refer to compounds, materials, compositions and / or dosage forms that are, within the scope of good medical judgment, suitable for use in contact with the tissues of human, animals and plants without undue toxicity, irritation, allergic response, or other problem or complication, and are commensurate with a reasonable benefit / risk ratio.

[0053] Throughout the specification, groups and substituents thereof may be chosen to provide stable moieties and compounds.

[0054] Compounds of the Invention In one embodiment, the present invention provides an isolated compound of formula (I) [ka] and enantiomers, diastereomers, tautomers, or pharma- ceutically acceptable salts thereof. wherein each stereocenter (indicated with an "*") may be in either the R or S configuration. Thus, when the phrase "compound of formula (I)" is used herein, it is meant to include its enantiomers, diastereomers, tautomers, and pharma- ceutically acceptable salts. The depsipeptide compounds of the present invention and their salts may exist in their tautomeric forms (e.g., as amides or guanidines). All such tautomeric forms are contemplated herein as part of the present invention. The compound of formula (I), for example, the compound isolated from bacterial isolate ISO18629, may also be referred to as "NOVO26". In some embodiments, the teachings of the present invention relate to mixtures of stereoisomers. In other embodiments, the teachings of the present invention relate specifically to a single stereoisomer of the compound of formula (I).

[0055] In one particular embodiment, the present invention relates in particular to a compound of formula (II) [ka] or a tautomer or a pharma- ceutically acceptable salt thereof.

[0056] All stereoisomers of the depsipeptide compounds of the present invention, including enantiomeric and diastereomeric forms (e.g., those that may exist due to asymmetric carbons on various substituents), are intended to be within the scope of the present invention. In one embodiment, the compound of formula (I) is a mixture. Alternatively, however, the compound of formula (I) is a single stereoisomer that is substantially free of other stereoisomers (e.g., as a pure or substantially pure optical isomer having a particular activity). In a particular embodiment, the compound of formula (I) is a single stereoisomer represented by formula (II). In some embodiments, the present invention relates to a single stereoisomer in the presence of less than about 10% (by dry weight) of other isomers. In some embodiments, the present invention relates to a single stereoisomer in the presence of less than about 5% (by dry weight) of other isomers. In some embodiments, the present invention relates to a single stereoisomer in the presence of less than about 2% (by dry weight) of other isomers. In some embodiments, the present invention relates to a single stereoisomer in the presence of less than about 1% (by dry weight) of other isomers. In some embodiments, the invention relates to a single stereoisomer in the presence of about 5% (by dry weight) to about 10% (by dry weight) of other isomers. In some embodiments, the invention relates to a single stereoisomer in the presence of about 1% (by dry weight) to about 5% (by dry weight) of other isomers. In some embodiments, the invention relates to a single stereoisomer in the presence of about 1% (by dry weight) to about 10% (by dry weight) of other isomers. In some embodiments, the invention relates to a single stereoisomer in the presence of about 5% (by dry weight) to about 10% (by dry weight) of other isomers. Unrelated compounds constitute less than 2% (by dry weight). In other embodiments, the teachings of the invention are directed to mixtures of stereoisomers. The chiral centers of the compounds of formula (I) can have the S or R configuration as defined by the IUPAC 1974 Recommendations. Stereoisomeric forms can be resolved by physical methods, such as, for example, fractional crystallization, separation or crystallization of diastereomeric derivatives or separation by chiral column chromatography.The individual optical isomers can be obtained from the mixture of stereoisomers by any suitable method, including but not limited to conventional methods such as, for example, salt formation with an optically active acid followed by crystallization.

[0057] The depsipeptide compounds of the present invention, after their preparation, can be, for example, isolated and purified to obtain a composition containing an amount by weight of 90% or more (by dry weight), which is then used or formulated as described herein. The depsipeptide compounds of the present invention, after their preparation, can be, for example, isolated and purified to obtain a composition containing an amount by weight of 95% or more (by dry weight), which is then used or formulated as described herein. The depsipeptide compounds of the present invention, after their preparation, can be, for example, isolated and purified to obtain a composition containing an amount by weight of about 90% (by dry weight) to about 95% (by dry weight), which is then used or formulated as described herein. The depsipeptide compounds of the present invention, after their preparation, can be, for example, isolated and purified to obtain a composition containing an amount by weight of about 85% (by dry weight) to about 95% (by dry weight), which is then used or formulated as described herein. After they are prepared, the depsipeptide compounds of the invention can be, for example, isolated and purified to obtain a composition containing about 95% (by dry weight) to about 99% (by dry weight) by weight, which composition can then be used or formulated as described herein.

[0058] All configurational isomers of the compounds of the present invention are contemplated, either in admixture or in pure or substantially pure form.

[0059] In one embodiment, the present invention provides an isolated compound of formula (III) [ka] or a tautomer or a pharma- ceutically acceptable salt thereof.

[0060] Each of R1 to R6 is independently hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, aryl, C(=O)R a and S(=O)2R b Each R a is independently hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, or aryl; each R b are independently alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, or aryl. Also, any carbon or hydrogen may be 13 C or 2 H. In one embodiment, the present invention relates to an isolated compound of formula (III), or an enantiomer, diasteromer, tautomer, or pharma- ceutically acceptable salt thereof.

[0061] The term "alkyl" refers to a straight or branched chain alkane (hydrocarbon) radical containing 1 to 12 carbon atoms, e.g., 1 to 6 carbon atoms. Exemplary "alkyl" groups include methyl, ethyl, propyl, isopropyl, n-butyl, t-butyl, isobutyl pentyl, hexyl, isohexyl, heptyl, 4,4-dimethylpentyl, octyl, 2,2,4-trimethylpentyl, nonyl, decyl, undecyl, dodecyl, and the like.

[0062] The term "alkenyl" refers to a straight or branched chain hydrocarbon radical containing from 2 to 12 carbon atoms and at least one carbon-carbon double bond. Exemplary such groups include ethenyl or allyl.

[0063] The term "alkynyl" refers to a straight or branched chain hydrocarbon radical containing from 2 to 12 carbon atoms and at least one carbon-carbon triple bond. Exemplary such groups include ethynyl.

[0064] The term "cycloalkyl" refers to a fully saturated cyclic hydrocarbon group containing from 1 to 4 rings and from 3 to 8 carbons per ring. Exemplary such groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and the like.

[0065] The term "cycloalkenyl" refers to a partially unsaturated cyclic hydrocarbon group containing 1 to 4 rings and 3 to 8 carbons per ring. Exemplary such groups include cyclobutenyl, cyclopentenyl, cyclohexenyl, and the like.

[0066] The term "aryl" refers to a cyclic aromatic hydrocarbon group having from 1 to 5 aromatic rings, particularly a monocyclic or bicyclic group such as phenyl, biphenyl, or naphthyl. If the aromatic rings of an aryl group contain more than one aromatic ring (e.g., bicyclic), they may be linked at a single point (e.g., biphenyl) or fused (e.g., naphthyl, phenanthrenyl, etc.).

[0067] In one embodiment, the present invention provides an isolated compound of formula (IV) [ka] wherein X is NH, O or S. or a tautomer or a pharma- ceutically acceptable salt thereof. In one particular embodiment, X is NH. Alternatively, X is O. Alternatively, X is S. In one embodiment, the invention relates to an isolated compound of formula (IV), or an enantiomer, diastereomer, tautomer, or a pharma- ceutically acceptable salt thereof.

[0068] In some embodiments, the compound of formula (I), (II), (III) or (IV) is a natural product of a bacterial species. In some embodiments, the compound of formula (I), (II), (III) or (IV) is a natural product isolated from a bacterial species. In some embodiments, the compound of formula (I), (II), (III) or (IV) is a natural product of bacterial isolate ISO18629. For example, in some embodiments, the compound of formula (I), (II), (III) or (IV) is derived from a bacterial culture of bacterial isolate ISO18629. In some embodiments, the compound of formula (I), (II), (III) or (IV) is a natural product isolated from bacterial isolate ISO18629.

[0069] In some embodiments, the compound of formula (I) has at least five chromatic adenosine triphosphate (ADP) at chemical shifts (+ / - 0.2 ppm) in DMSO-d6 selected from 36.3 ppm, 36.5 ppm, 36.9 ppm, 37.4 ppm, 52.1 ppm, 52.2 ppm, 52.7 ppm, 53.5 ppm, 55.7 ppm, 56.1 ppm, 56.4 ppm, 56.7 ppm, 57.3 ppm, 57.8 ppm, 57.9 ppm, 61.8 ppm, and 71.1 ppm. 13 In some embodiments, the compound of formula (I) is characterized by a C nuclear magnetic resonance peak. In some embodiments, the compound of formula (I) is characterized by a C nuclear magnetic resonance peak. In some embodiments, the compound of formula (I) is characterized by a C nuclear magnetic resonance peak. In some embodiments, the compound of formula (I) is characterized by a C nuclear magnetic resonance peak. In some embodiments, the compound of formula (I) is characterized by a C nuclear magnetic resonance peak. In some embodiments, the compound of formula (I) is characterized by a C nuclear magnetic resonance peak. In some embodiments, the compound of formula (I) is characterized by a C nuclear magnetic resonance peak. In some embodiments, the compound of formula (I) is characterized by a C nuclear magnetic resonance peak. 13 Characterized by C nuclear magnetic resonance peaks In some embodiments, the compound of formula (I) has at least 10 chromatic aberrations at chemical shifts (+ / - 0.2 ppm) in DMSO-d6 selected from 36.3 ppm, 36.5 ppm, 36.9 ppm, 37.4 ppm, 52.1 ppm, 52.2 ppm, 52.7 ppm, 53.5 ppm, 55.7 ppm, 56.1 ppm, 56.4 ppm, 56.7 ppm, 57.3 ppm, 57.8 ppm, 57.9 ppm, 61.8 ppm, and 71.1 ppm. 13 In some embodiments, the compound of formula (I) is characterized by a C nuclear magnetic resonance peak. In some embodiments, the compound of formula (I) is characterized by a C nuclear magnetic resonance peak. In some embodiments, the compound of formula (I) is characterized by a C nuclear magnetic resonance peak. In some embodiments, the compound of formula (I) is characterized by a C nuclear magnetic resonance peak. In some embodiments, the compound of formula (I) is characterized by a C nuclear magnetic resonance peak. In some embodiments, the compound of formula (I) is characterized by a C nuclear magnetic resonance peak. In some embodiments, the compound of formula (I) is characterized by a C nuclear magnetic resonance peak. In some embodiments, the compound of formula (I) is characterized by a C nuclear magnetic resonance peak. 13 In some embodiments, the compound of formula (I) is characterized by a C nuclear magnetic resonance peak at chemical shifts (+ / - 0.2 ppm) in DMSO-d6 of 36.3 ppm, 36.5 ppm, 36.9 ppm, 37.4 ppm, 52.1 ppm, 52.2 ppm, 52.7 ppm, 53.5 ppm, 55.7 ppm, 56.1 ppm, 56.4 ppm, 56.7 ppm, 57.3 ppm, 57.8 ppm, 57.9 ppm, 61.8 ppm, and 71.1 ppm. 13 C nuclear magnetic resonance peak. In certain embodiments, the chemical shifts are recorded from a nuclear magnetic resonance spectrum obtained using a 500 MHz magnet. Chemical shifts are reported herein within an error of + / - 0.2 ppm.

[0070] In a further embodiment, the present invention provides a compound having a molecular weight of about 1242.47 g / mol, a proton nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 1, a proton nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 133, a COSY nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 3, a DEPT-135 nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 4, an HSQC nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 5, or an HMBC nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 6. In another embodiment, the invention relates to a compound of formula (I) characterized by at least two, at least three, at least four, at least five, or at least six of the above features. In yet a further embodiment, the invention relates to a compound of formula (I) characterized by at least two, at least three, at least four, at least five, or at least six of the above features. In yet a further embodiment, the invention relates to a compound of formula (I) characterized by at least two, at least three, at least four, at least five, or at least six of the above features. 13 C nuclear magnetic resonance spectrum, a COSY nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 3, a DEPT-135 nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 4, an HSQC nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 5, and an HMBC nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 6.

[0071] In some embodiments, the present invention provides a method for determining the molecular weight of a compound having a molecular weight of 36.3 ppm, 36.5 ppm, 36.9 ppm, 37.4 ppm, 52.1 ppm, 52.2 ppm, 52.7 ppm, 53.5 ppm, 55.7 ppm, 56.1 ppm, 56.4 ppm, 56.7 ppm, 57.3 ppm, 57.8 ppm, 57.9 ppm, 61.8 ppm, and 71.1 ppm in DMSO-d6 (+ / - 0.2 ppm) chemical shifts. 13 The present invention relates to an isolated natural product of the bacterial isolate ISO18629, characterized by a molecular weight of about 1242.47 g / mol, a proton nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 133 , a COSY nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 3 , a DEPT-135 nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 4 , an HSQC nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 5 , and / or an HMBC nuclear magnetic resonance spectrum substantially the same as that shown in FIG. 6 .

[0072] In another embodiment, the present invention relates to a pharmaceutical composition comprising a compound of the present invention as described herein and a pharma- ceutically acceptable excipient, carrier, or diluent, hi certain embodiments, the composition further comprises an agent selected from an antibiotic, an antifungal, an antiviral, an antiprotozoan, an antiparasitic, an antineoplastic, an immunomodulatory agent, an antihypercholesterolemic agent, and combinations thereof.

[0073] The depsipeptide compounds of the present invention can form salts, which are also within the scope of the present invention. Reference to the compounds of the present invention herein is understood to include reference to salts of the compounds, unless otherwise specified. The term "salt(s)" as used herein refers to acid salts formed with inorganic and / or organic acids. Pharmaceutically acceptable (i.e., non-toxic, physiologically acceptable) salts are preferred, although other salts are also useful, for example, in isolation or purification steps that may be used during preparation. Salts of the compounds of the present invention can be formed by reacting a compound of formula (I), (II), (III) or (IV) with an amount (e.g., equivalent amount) of acid in a medium (e.g., a medium in which the salt precipitates) or in an aqueous medium or an aqueous and organic medium, followed by lyophilization.

[0074] The depsipeptide compounds of the invention that contain a basic moiety, including, but not limited to, an amine or guanidine, can form salts with a variety of organic and inorganic acids. Exemplary acid addition salts include acetate (such as those formed with acetic acid or a trihaloacetic acid, e.g., trifluoroacetic acid), adipate, alginate, ascorbate, aspartate, benzoate, benzenesulfonate, bisulfate, borate, butyrate, citrate, camphorate, camphorsulfonate, cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate, fumarate, glucoheptanoate, glycerophosphate, hemisulfate, heptanoate, hexanoate, hydrochloride, hydrobromide, hydroiodide, hydroxyethanesulfate, and the like. Examples of suitable salts include sulfonates (e.g., 2-hydroxyethanesulfonate), lactate, maleate, methanesulfonate, naphthalenesulfonate (e.g., 2-naphthalenesulfonate), nicotinate, nitrate, oxalate, pectinate, persulfate, phenylpropionate (e.g., 3-phenylpropionate), phosphate, picrate, pivalate, propionate, salicylate, succinate, sulfate (such as formed with sulfuric acid), sulfonate, tartrate, thiocyanate, toluenesulfonate, e.g., tosylate, undecanoate, and the like.

[0075] Methods for Preparing the Compounds of the Invention In yet another embodiment, the present invention provides a compound of formula (I) [ka] or a method for producing an enantiomer, diastereomer, tautomer, or pharma- ceutically acceptable salt thereof.

[0076] In yet another embodiment, the present invention relates to a compound of formula (II) [ka] or a tautomer or a pharma- ceutically acceptable salt thereof.

[0077] In one embodiment, the present invention provides a compound of formula (III) [ka] or a tautomer or a pharma- ceutically acceptable salt thereof.

[0078] Each of R1 to R6 is independently hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, aryl, C(=O)R a and S(=O)2R b Each R a is independently hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, or aryl; each R b are independently alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, or aryl. Also, any carbon or hydrogen may be 13 C or 2 It may be replaced by H.

[0079] In yet another embodiment, the present invention relates to a compound of formula (IV) [ka] wherein X is NH, O or S. or a tautomer or a pharma- ceutically acceptable salt thereof.

[0080] The method comprises cultivating or culturing the bacterial isolate ISO18629 under aerobic conditions in a medium containing assimilable sources of carbon, nitrogen and inorganic salts, so as to produce an assayable amount of a compound of formula (I), (II), (III) or (IV). Specific conditions for culturing the bacterial isolate ISO 18629 are described in the Examples section below.

[0081] In certain embodiments, the method further comprises isolating the compound of formula (I), (II), (III) or (IV). The compound of formula (I), (II), (III) or (IV) may be isolated by centrifuging the fermentation broth (e.g., centrifugation at 10,000 rpm for 20 minutes). The supernatant from the centrifugation is then adsorbed onto a reverse-phase polymeric resin (e.g., HP-20, Mitsubishi). After washing the column with water and 80% aqueous acetone, the compound is eluted with acetone. This acetone extract is then dried (e.g., under reduced pressure) to leave a dark orange oil. Hexane is then added to the oil, and the mixture is then sonicated and centrifuged. The supernatant is then decanted, and the remaining residue is dried under reduced pressure. This residue is dissolved in DMSO and purified. Purification can be achieved by RP-HPLC on a preparative C-18 column (HO / AcN (with 0.1% TFA), 10-100% over 35 minutes). The compound-containing fractions may then be lyophilized. Specific conditions for isolating compounds of Formula (I), (II), (III) or (IV) are described in the Examples section below.

[0082] In some embodiments, the method further comprises isolating the compound of formula (I), (II), (III) or (IV) to a purity of at least about 75% (by dry weight). In other embodiments, the process further comprises isolating the compound of formula (I), (II), (III) or (IV) to a purity of at least about 80% (by dry weight). In other embodiments, the process further comprises isolating the compound of formula (I), (II), (III) or (IV) to a purity of at least about 85% (by dry weight). In other embodiments, the process further comprises isolating the compound of formula (I), (II), (III) or (IV) to a purity of at least about 90% (by dry weight). In other embodiments, the process further comprises isolating the compound of formula (I), (II), (III) or (IV) to a purity of at least about 95% (by dry weight). In other embodiments, the process further comprises isolating the compound of formula (I), (II), (III) or (IV) to a purity of at least about 97% (by dry weight). In other embodiments, the process further comprises isolating the compound of formula (I), (II), (III) or (IV) to a purity of at least about 99% (by dry weight).

[0083] In yet another embodiment, the present invention relates to a compound of Formula (I), (II), (III) or (IV) prepared according to the methods described herein.

[0084] Compounds of formula (II), such as NOVO26, are produced by the ISO18629 isolate. Bacterial isolate ISO18629 was deposited at the Agricultural Research Service Culture Collection (NRRL), National Center for Agricultural Utilization Research, Agricultural Research Service, USDepartment of Agriculture, 1815 North University Street, Peoria, Illinois 61604, on September 6, 2013, and was assigned NRRL Accession No. B-50868. This deposit is maintained under the terms of the Budapest Treaty on the International Recognition of the Deposit of Microorganisms for the Purposes of Patent Procedure. This deposit is deemed merely a convenience to those skilled in the art and is not an admission that it is required under 35 U.S.C. § 112.

[0085] ISO 18629 was isolated from terrestrial soil in Maine using a technique for isolating "unculturable" microorganisms described in U.S. Patent No. 7,011,957. They utilize a growth chamber that is sealed with a permeable membrane and thus impermeable to microbial cells but permeable to the diffusion of components from the environment. The nucleotide sequence of the 16S rDNA gene of ISO18629 was shown to be most closely related (97.4% similarity) to the Burkholderiales bacterium YT0099 (accession no. AB362826-1) when compared to the GenBank nucleotide collection by BLAST alignment.

[0086] The growth chamber is designed to allow the growth, isolation into pure culture, and characterization of microorganisms that are "unculturable" at the time of the present invention. This desired result can be achieved because the conditions within the chamber are quite similar, if not identical, to the microorganism's natural environment. One version of such a chamber is formed from a solid substrate, such as a glass or silicon slide or a nylon or stainless steel washer, onto which is attached an orifice sandwiched between two sturdy membranes, such as polycarbonate or other inert materials. The membranes have pore sizes, such as 0.025 μm to 0.03 μm, small enough to retain all the microorganisms within the chamber, but large enough to allow components from the environment to diffuse into the chamber and waste products to diffuse out of the chamber. After sealing the substrate on the bottom with a membrane, the chamber is partially filled with a cell suspension in an appropriate growth medium.

[0087] The compounds of the present invention can be isolated from bacterial isolate ISO18629, for example, using the following method. The bacterial isolate can be grown on a suitable support. The colonies can then be homogenized and used to inoculate a seed broth. After fermentation, the seed culture can then be used to inoculate a production broth. After optional further fermentation, the compound of formula (I), (II), (III) or (IV) can be recovered from the broth, for example, using centrifugation and preparative HPLC. Specific exemplary methods for producing the compound of formula (II) are discussed in more detail in the Examples section below.

[0088] Alternatively, the compounds of the present disclosure can be synthesized. For example, Scheme 1 shows one non-limiting method for synthesizing the guanidine-substituted compounds of formula (III). [ka] Scheme 2 is a non-limiting representative example of the synthesis of amine-substituted compounds of formula (III). [ka] Scheme 3 provides a non-limiting method for synthesizing alcohol-substituted compounds of formula (III). [ka]

[0089] Scheme 4 is an exemplary method for synthesizing compounds of formula (IV). [ka]

[0090] Methods of Treatment with Compounds of the Invention The present invention also provides a method of inhibiting the growth of a pathogen. The method includes contacting the pathogen with an effective amount of one or more depsipeptide compounds of formula (I), (II), (III) or (IV), thereby inhibiting the growth of the pathogen compared to the growth of the pathogen not treated with the compound. In certain embodiments, the method reduces the growth of the pathogen compared to the growth of the pathogen not treated with the compound. In other cases, the treatment results in the death of the pathogen. Non-limiting examples of pathogens include, but are not limited to, bacteria, fungi, viruses, protozoa, helminths, parasites and combinations thereof. These methods can be performed in vivo or in vitro.

[0091] The antibacterial activity of the depsipeptide compounds of formula (I), (II), (III) or (IV) against a particular bacterium can be assessed by in vitro assays such as monitoring the zone of inhibition and minimum inhibitory concentration (MIC) assays. The antifungal activity of the depsipeptide compounds of formula (I), (II), (III) or (IV) ... those described, for example, in Sanati et al., A new triazole, voriconazole (UK-109,496), The antiviral properties of the depsipeptide compounds of formula (I), (II), (III) or (IV) can be determined, for example, by monitoring the inhibition of influenza neuraminidase or as described in Tisdale M., Monitoring of viral The antiprotozoal activity of the depsipeptide compounds of formula (I), (II), (III) or (IV) can be determined by determining the viability of parasitic protozoans such as Trichomonas vaginalis and Giardia lamblia as described in Katiyar et al., Antiprotozoal activities of benzimidazoles and correlations with beta-tubulin sequence, Antimicrob. Agents Chemother., 1994 September; 38(9): 2086-2090. The anthelmintic activity of the depsipeptide compounds of formula (I), (II), (III) or (IV) can be determined by determining the effect of said compounds on the survival rate of nematodes such as Schistosoma mansoni, Schistosoma cercariae and Caenorhabditis elegans, for example as described in Molgaard P. et al., Traditional herbal remedies used for the treatment of urinary schistosomiasis in Zimbabwe, J. Ethnopharmacol., 1994 Apr; 42(2):125-32.

[0092] In other embodiments, the present invention is directed to methods of treating a disorder (e.g., a pathogen infection) in a subject in need of such treatment by administering to the subject an effective amount of one or more depsipeptide compounds described herein. In certain embodiments, the disorder is caused by a pathogen, such as, but not limited to, a bacterium, a fungus, a virus, a protozoan, a helminth, a parasite, or a combination thereof.

[0093] In some embodiments, the disorder is caused by bacteria. The depsipeptide compounds described herein may be useful against both gram-positive and gram-negative bacteria. In particular, the disorder is caused by gram-positive bacteria. Alternatively, the disorder is caused by gram-negative bacteria. Non-limiting examples of gram-positive bacteria include Streptococcus, Staphylococcus, Enterococcus, Corynebacteria, Listeria, Bacillus, Erysipelothrix and Actinomycetes. In some embodiments, the compounds of formula (I), (II), (III) or (IV) are used to treat infections caused by one or more of the following: Helicobacter pylori, Legionella pneumophilia, Mycobacterium tuberculosis, Mycobacterium avium, Mycobacterium intracellulare, Mycobacterium kansaii, Mycobacterium gordonae, Mycobacteria sporozoites, Staphylococcus aureus, Staphylococcus epidermidis, Neisseria gonorrhoeae, Neisseria meningitidis, Listeria monocytogenes, Pyogenic Streptococcus (Group A Streptococcus), Pyogenic Streptococcus agalactiae (Group B Streptococcus), Streptococcus dysgalactia, Streptococcus faecalis, Streptococcus bovis, Streptococcus pneumoniae, Pathogenic Campylobacter sporozoites, Enterococcus sporozoites, Haemophilus influenzae, Pseudomonas aeruginosa, Bacillus anthracis, Bacillus subtilis, Escherichia coli, Corynebac terium diphtheriae, Corynebacterium jeikeium, Corynebacterium sporozoite, Erysipelothrix rhusiopathiae, Clostridium perfringens, Clostridium tetani, Clostridium difficile, Enterobacter aerogenes, Klebsiella pneumoniae, Pasteurella multocida, Bacteroides thetaiotamicron, Bacteroides uniformis, Bacteroides vulgatus, Fusobacterium nucleatum, Streptobacillus moniliformis, Leptospira and Actinomyces israelli. In certain embodiments, the compounds described herein are useful for treating infections caused by methicillin-resistant Staphylococcus aureus (MRSA) or vancomycin-resistant Enterococci (VRE). MRSA causes approximately 19,000 deaths per year in the United States. Most of these deaths are due to hospital-acquired MRSA (HA-MRSA), but it is known that community-acquired MRSA (CA-MRSA) is in fact more virulent and can be fatal to still healthy individuals. The virulence of CA-MRSA is due in part to the expression of phenol-soluble modulin or PSM peptides. Thus, to treat CA-MRSA, compounds of formula (I), (II), (III) or (IV) can be used in combination with agents that modulate the expression and / or activity of virulence factors, such as, but not limited to, PSM peptides. In certain embodiments, depsipeptide compounds of formula (I), (II), (III) or (IV) can be used to treat spirochetes such as Borelia burgdorferi, Treponema pallidium and Treponema pertenue.

[0094] In a particular embodiment, the gram-positive bacteria is selected from Staphylococcus (including, for example, S. aureus, S. epidermidis, S. warneri and S. haemolyticus), Streptococcus (including, for example, S. viridans, S. pneumoniae, S. agalactiae and S. pyogenes), Bacillus (including, for example, B. anthracis and B. subtilis), Clostridium (including, for example, C. difficile), Propionibacterium (including, for example, P. acnes), Enterococcus (including, for example, E. faecium, E. faecalis, vancomycin-resistant E. faecium and vancomycin-resistant E. faecalis), and Mycobacterium (including, for example, M. smegmatis and M. tuberculosis). The compounds described herein are useful for treating disorders caused by these bacteria. Examples of such disorders include acute bacterial skin and skin structure infections, diarrhea associated with C. difficile, anthrax, sepsis, botulism, urinary tract infections, bacteremia, bacterial endocarditis, diverticulitis, meningitis, pneumonia, and tuberculosis.

[0095] In a particular embodiment, the gram-negative bacteria is selected from the group consisting of Haemophilus (including, for example, H. influenzae species), Klebsiella (including, for example, K. pneumoniae species), Pseudomonas (including, for example, P. aeruginosa species), Escherichia (including, for example, E. coli species), Yersinia (including, for example, Y. pestis species), Neisseria (including, for example, N. gonorrhoeae species), Bacteriodes (including, for example, B. fragilis species), Proteus spp. teus (including, for example, P. mirabilis and P. vulgaris species), Enterobacter (including, for example, E. cloacae and E. aerogenes species), Serratia (including, for example, S. marcescens species), Aci In one particular embodiment, the gram-negative bacteria is selected from Haemophilus, particularly H. influenzae, or Moraxella, particularly M. catarrhalis. The compounds described herein are useful for treating disorders caused by these bacteria. Examples of such disorders include influenza, bacteremia, pneumonia, acute bacterial meningitis, gonorrhea, urinary tract infections, respiratory tract infections, catheter-associated bacteremia, wound infections, otitis media, bronchitis, sinusitis, and laryngitis. In other embodiments, the depsipeptide compounds described herein may be useful for treating viral disorders. Non-limiting examples of infectious viruses that can be treated by the compounds of Formula (I), (II), (III) or (IV) include Retroviridae (e.g., human immunodeficiency viruses, such as HIV-1 (also called HTLV-III, LAV or HTLV-III / LAV) or HIV-III; and other isolates, such as HIV-LP; Picornaviridae (e.g., poliovirus, Hepatitis A virus, enterovirus, human coxsackievirus, rhinovirus, echovirus); Calciviridae (e.g., strains that cause gastroenteritis); Togaviridae (e.g., equine encephalitis virus, rubella virus); Flaviviridae (e.g., avian encephalitis virus ... ridae) (e.g., dengue virus, encephalitis virus, yellow fever virus); Coronaviridae (e.g., coronavirus, severe acute respiratory syndrome (SARS) virus); Rhabdoviridae (e.g., vesicular stomatitis virus, rabies virus); Filoviridae (e.g., Ebola virus); Paramyxoviridae (e.g., parainfluenza virus, mumps virus, measles virus, respiratory syncytial virus); Orthomyxoviridae (e.g., influenza virus); Bunyaviridae (e.g., Hantavirus, Bungavirus, Phlebovirus, and Nairovirus); Arenaviridae (hemorrhagic fever viruses);Reoviridae (e.g., reoviruses, orbiviurses, and rotaviruses); Birnaviridae; Hepadnaviridae (e.g., hepatitis B virus); Parvoviridae (parvoviruses); Papovaviridae (papillomaviruses, polyomaviruses); Adenoviridae (most adenoviruses); Herpesviridae (e.g., herpes simplex virus (HSV) 1 and 2, varicella zoster virus, cytomegalovirus (CMV), herpes virus, viridae (e.g., variola virus, vaccinia virus, pox virus); and iridoviridae (e.g., African swine fever virus); and unclassified viruses (e.g., pathogens of spongiform encephalopathies, pathogens of delta hepatitis (thought to be a defective satellite of hepatitis B virus), pathogens of non-A, non-B hepatitis (Class 1=endogenous, Class 2=parentally transmitted, i.e., hepatitis C); Norwalk and related viruses, and astroviruses). In certain embodiments, the compounds of formula (I), (II), (III) or (IV) are used to treat influenza virus, human immunodeficiency virus, and herpes simplex virus.;

[0096] In some embodiments, the depsipeptide compounds of formula (I), (II), (III) or (IV) may be useful for treating disorders caused by fungi. Non-limiting examples of fungi that may be inhibited by the compounds of formula (I), (II), (III) or (IV) include, but are not limited to, Cryptococcus neoformans, Histoplasma capsulatum, Coccidioides immitis, Blastomyces dermatitidis, Chlamydia trachomatis, Candida albicans, Candida tropicalis, Candida glabrata, Candida krusei, Candida parapsilosis, Candida dubliniensis, Candida lusitaniae, Epidermophyton floccosum, Microsporum audouinii, Mic rosporum canis, Microsporum canis var. distortum Microsporum cookei, Microsporum equinum, Microsporum ferrugineum, Microsporum fulvum, Microsporum gallinae, Microsporum gypseum, Microsporum nanum, Microsporum persicolor, Trichophyton ajelloi, Trichophyton concentricum, Trichophyton equinum, Trichophyton flavescens, Trichophyton gloriae, Trichophyton megnini, Trichophyton mentagrophytes var. erinacei, Trichophyton mentagrophytes var. interdigitale, Trichophyton phaseoliforme, Trichophyton rubrum, Trichophyton rubrum downy strain, Trichophyton rubrum granular strain, Trichophyton schoenleinii, Trichophyton simii, Trichophyton soudanense, Trichophyton terrestre, Trichophyton tonsurans, Trichophyton vanbreuseghemii, Trichophyton verrucosum, Trichophyton violaceum, Trichophyton yaoundei, Aspergillus fumigatus, Aspergillus flavus and Aspergillus clavatus.

[0097] In yet another embodiment, the depsipeptide compounds described herein are useful for treating disorders caused by protozoa.Non-limiting examples of protozoa that can be inhibited by the compounds of formula (I), (II), (III) or (IV) include, but are not limited to, Trichomonas vaginalis, Giardia lamblia, Entamoeba histolytica, Balantidium coli, Cryptosporidium parvum and Isospora belli, Trypansoma cruzi, Trypanosoma gambiense, Leishmania donovani and Naegleria fowleri.

[0098] In certain embodiments, the depsipeptide compounds described herein are useful for treating disorders caused by helminths. Non-limiting examples of helminths that may be inhibited by the compounds of formula (I), (II), (III) or (IV) include, but are not limited to, Schistosoma mansoni, Schistosoma cercariae, Schistosoma japonicum, Schistosoma mekongi, Schistosoma hematobium, Ascaris lumbricoides, Strongyloides stercoralis, Echinococcus granulosus, Echinococcus multilocularis, Angiostrongylus cantonensis, Angiostrongylus constaricensis, Fasciolopis buski, Capillaria philippinensis, Paragonimus westermani, Ancylostoma dudodenale, Necator americanus, Trichinella spiralis, Wuchereria bancrofti, Brugia malayi and Brugia timori, Toxocara canis, Toxocara cati, Toxocara vitulorum, Caenorhabiditis elegans and Anisakis species.

[0099] In some embodiments, the depsipeptide compounds described herein are useful for treating disorders caused by parasites. Non-limiting examples of parasites that can be inhibited by the compounds of formula (I), (II), (III) or (IV) include, but are not limited to, Plasmodium falciparum, Plasmodium yoelli, Hymenolepis nana, Clonorchis sinensis, Loa loa, Paragonimus westermani, Fasciola hepatica, and Toxoplasma gondii. In a particular embodiment, the parasite is a malaria parasite.

[0100] In other embodiments, the depsipeptide compounds are used to inhibit the growth of an infectious agent compared to the growth of the infectious agent without treatment with the compound. Non-limiting examples of infectious agents include, but are not limited to, bacteria, fungi, viruses, protozoa, helminths, parasites, and combinations thereof. The depsipeptide compounds can be used to inhibit the infectious agent in vivo or in vitro.

[0101] Pharmaceutical Compositions Containing Compounds of the Invention The present invention also provides pharmaceutical compositions comprising at least one depsipeptide compound of formula (I), (II), (III) or (IV) (or an enantiomer, diastereomer, tautomer, or a pharma- ceutically acceptable salt or solvate thereof) and a pharma- ceutically acceptable carrier. These depsipeptide compositions are suitable for administration to a subject (e.g., a mammal, such as a human). The pharmaceutical compositions can be used to treat disorders. Non-limiting examples of such disorders are provided above and include infection by a pathogen (e.g., bacterial).

[0102] In one embodiment, the depsipeptide compound is administered in a pharma- ceutically acceptable carrier. Any suitable carrier known in the art can be used. A carrier that effectively solubilizes the compound of the present invention is preferred. Carriers include, but are not limited to, solids, liquids, or mixtures of solids and liquids. The carriers can take the form of capsules, tablets, pills, powders, lozenges, suspensions, emulsions, or syrups. The carriers can include substances that act as flavoring agents, lubricants, solubilizers, suspending agents, binders, stabilizers, tablet disintegrants, and encapsulating materials. The phrase "pharma- ceutically acceptable" is used herein to refer to compounds, materials, compositions, and / or dosage forms that, within the scope of good medical judgment, are suitable for use in contact with the tissues of human beings and animals without undue toxicity, irritation, allergic response, or other problems or complications, and are commensurate with a reasonable benefit / risk ratio.

[0103] Non-limiting examples of materials that can serve as pharma- ceutically acceptable carriers include: (1) sugars, such as lactose, glucose, and sucrose, (2) starches, such as corn starch and potato starch, (3) cellulose and its derivatives, such as sodium carboxymethylcellulose, ethylcellulose, and cellulose acetate, (4) powdered tragacanth, (5) malt, (6) gelatin, (7) talc, (8) excipients, such as cocoa butter and suppository wax, (9) oils, such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil, and soybean oil, and (10) glycols. , for example, propylene glycol, (11) polyols, such as glycerin, sorbitol, mannitol, and polyethylene glycol, (12) esters, such as ethyl oleate and ethyl laurate, (13) agar, (14) buffers, such as magnesium hydroxide and aluminum hydroxide, (15) alginic acid, (16) pyrogen-free water, (17) isotonic saline, (18) Ringer's solution, (19) ethyl alcohol, (20) phosphate buffers, and (21) other compatible non-toxic substances used in pharmaceutical formulations.

[0104] The above formulations can be conveniently presented in unit dosage form and can be prepared by any method well known in the art of pharmacy. The amount of active ingredient that can be combined with carrier materials to produce a single dosage form will vary depending on the subject being treated, the particular mode of administration, the particular condition being treated, among others. The amount of active ingredient that can be combined with carrier materials to produce a single dosage form is generally the amount of compound that produces a therapeutic effect. Generally, this amount ranges from about 1 percent to about 99 percent of active ingredient out of 100%, preferably from about 5 percent to about 70 percent, most preferably from about 10 percent to about 30 percent.

[0105] Methods of preparing these formulations or compositions include the step of bringing into association a compound of formula (I), (II), (III) or (IV) with the carriers and, optionally, one or more accessory ingredients. In general, the formulations are prepared by uniformly and intimately bringing into association a depsipeptide compound of formula (I), (II), (III) or (IV) with liquid carriers, or suitably divided solid carriers, or both, and then, if necessary, shaping the product.

[0106] In solid dosage forms for oral administration (e.g., capsules, tablets, pills, dragees, powders, granules, etc.), the active ingredient is mixed with one or more additional ingredients, such as sodium citrate or dicalcium phosphate, and / or any of the following ingredients: fillers or extenders, such as, but not limited to, starch, lactose, sucrose, glucose, mannitol, and / or silicic acid; binders, such as, but not limited to, carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidone, sucrose, and / or gum acacia; humectants, such as, but not limited to, glycerol; disintegrants, such as, but not limited to, glycerol. Examples of suitable additives include, but are not limited to, agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates and sodium carbonate; solution retarders, such as, but not limited to, paraffin; absorption accelerators, such as, but not limited to, quaternary ammonium compounds; wetting agents, such as, but not limited to, cetyl alcohol and glycerol monostearate; absorbents, such as, but not limited to, kaolin and bentonite clay; lubricants, such as, but not limited to, talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate and mixtures thereof; and coloring agents. In the case of capsules, tablets and pills, the pharmaceutical compositions may also contain buffering agents. Similar types of solid compositions may also be used as fillers in soft and hard filled gelatin capsules, using excipients such as lactose or milk sugar and high molecular weight polyethylene glycols.

[0107] In powder, the carrier is a finely divided solid, which is mixed with an effective amount of finely divided medicine.Powder and spray can contain excipients such as lactose, talc, silicic acid, aluminum hydroxide, calcium silicate and polyamide powder, or mixtures of these substances, in addition to the compound of formula (I), (II), (III) or (IV).Spray can further contain customary propellants such as chlorofluorohydrocarbons, and volatile unsubstituted hydrocarbons such as butane and propane.

[0108] Tablets for systemic oral administration may contain one or more excipients known in the art, such as, for example, calcium carbonate, sodium carbonate, sugars (e.g., lactose, sucrose, mannitol, sorbitol), celluloses (e.g., methylcellulose, sodium carboxymethylcellulose), gums (e.g., gum arabic, tragacanth), one or more disintegrating agents (e.g., corn, starch or alginic acid, binders such as gelatin, collagen or gum acacia), lubricants (e.g., magnesium stearate, stearic acid or talc), inert diluents, preservatives, disintegrants, etc. (eg, sodium starch glycolate), together with a surfactant and / or a dispersing agent. A tablet may be made by compression or molding, optionally with one or more accessory ingredients.

[0109] In solutions, suspensions, emulsions or syrups, an effective amount of the depsipeptide compound is dissolved or suspended in a carrier, such as sterile water or an organic solvent, for example, aqueous propylene glycol. Other compositions can also be produced by dispersing the compound in aqueous starch or sodium carboxymethylcellulose, or in a suitable oil known in the art. The liquid dosage form can contain, for example, inert diluents commonly used in the art, such as water or other solvents, solubilizers and emulsifiers, for example, but not limited to, ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol, oils (especially cottonseed oil, peanut oil, corn oil, germ oil, olive oil, castor oil and sesame oil), glycerol, tetrahydrofuryl alcohol, polyethylene glycol and fatty acid esters of sorbitan, and mixtures thereof.

[0110] Besides inert diluents, the oral compositions can also include adjuvants such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, coloring, perfuming, and preservative agents.

[0111] Suspensions may contain, in addition to the active compound, suspending agents such as, for example, ethoxylated isostearyl alcohol, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum metahydroxide, bentonite, agar-agar and tragacanth, and mixtures thereof.

[0112] The formulation of the pharmaceutical composition for rectal or vaginal administration can be presented as a suppository, which can be prepared by mixing one or more compounds of formula (I), (II), (III) or (IV) with one or more suitable non-irritating excipients or carriers, including, for example, cocoa butter, polyethylene glycol, suppository wax or salicylates, and the suppository is also solid at room temperature and becomes liquid at body temperature, thus melting in the rectum or vaginal cavity and releasing the drug.Formulations suitable for vaginal administration also include pessaries, tampons, creams, gels, pastes, foams or sprays, containing carriers known in the art to be suitable.

[0113] Dosage forms for topical or transdermal administration of a compound of formula (I), (II), (III) or (IV) include powders, sprays, ointments, pastes, creams, lotions, gels, solutions, drops, patches and inhalants. The active depsipeptide compound can be mixed under sterile conditions with a pharma- ceutically acceptable carrier, and with any preservatives, buffers or propellants, as may be required.

[0114] The ointments, pastes, creams and gels may contain, in addition to the active compound, excipients such as animal and vegetable fats, oils, waxes, paraffins, starch, tragacanth, cellulose derivatives, polyethylene glycols, silicones, bentonite, silicic acid, talc and zinc oxide, or mixtures thereof.

[0115] Transdermal patches have the added advantage of providing controlled delivery of a compound of formula (I), (II), (III) or (IV) to the body. Such dosage forms can be produced by dissolving or dispersing the agent in a suitable medium. Absorption enhancers can also be used to increase the flux of the agent across the skin. The rate of such flux can be controlled by either providing a rate-controlling membrane or dispersing the depsipeptide compound in a polymer matrix or gel.

[0116] The depsipeptide compound is administered in an effective amount to a subject in need of such treatment. The phrase "effective amount" as used herein means an amount of a compound of formula (I), (II), (III) or (IV) or a composition containing a compound of formula (I), (II), (III) or (IV) effective to produce some desired effect in an animal. When a drug is used to achieve a therapeutic effect, it is recognized that the actual dose, including the "effective amount", will vary depending on several conditions, including but not limited to the particular condition being treated, the severity of the disease, the size and health of the patient, and the route of administration. A skilled medical practitioner can easily determine the appropriate dose using methods well known in the medical field. In some embodiments, the effective amount is an amount effective to treat a disorder in a subject in need of such treatment. Moreover, those skilled in the art will appreciate that the effective amount of the depsipeptide compound can be reduced or increased by fine-tuning and / or administering two or more depsipeptide compounds, or by administering the depsipeptide compound together with a second agent (e.g., antibiotics, antifungals, antivirals, NSAIDs, DMARDS, steroids, etc.). The effective amount can be determined, for example, empirically, by starting with a relatively low amount and gradually increasing it while evaluating the beneficial effect (e.g., reduction in symptoms). The actual effective amount is established by dose / response assays using methods standard in the art (Johnson et al., Diabetes, 42:1179 (1993)). As known to those skilled in the art, the effective amount depends on the bioavailability, bioactivity, and biodegradability of the depsipeptide compound.

[0117] In some embodiments, an effective amount is an amount that can reduce symptoms of a disorder in a subject. Thus, this amount can vary for each subject receiving treatment. For example, an effective amount of the depsipeptide compound can be about 1 μg to about 100 mg per kg of body weight. In one embodiment, an effective amount of the compound is about 1 μg to about 50 mg per kg of body weight. In a further embodiment, an effective amount of the compound is about 10 μg to about 10 mg per kg of body weight. When combined with a carrier, one or more depsipeptide compounds or agents can be present in an amount of about 1 weight percent to about 99 weight percent, with the remainder consisting of a pharma- ceutically acceptable carrier. In some embodiments, an effective amount is about 1 mg to about 10 g per dose, for example, about 10 mg to about 1 g per dose. The range values ​​and intermediate values ​​of the ranges recited above are intended to be encompassed by the teachings of the present invention.

[0118] Administration of the depsipeptide compound can be hourly, daily, weekly, monthly, yearly, or can be a single event. Furthermore, administration can be for a period of one day to one year or more. In some embodiments, administration refers to daily administration for a period of time, such as about one week, two weeks, three weeks, one month, three months, six months, or one year. In some embodiments, administration refers to weekly administration for a period of time, such as about one month, three months, six months, one year, or more.

[0119] The present invention also provides a kit comprising at least one depsipeptide compound of formula (I), (II), (III) or (IV). The kit may contain at least one container and may also include instructions directing the use of these materials. In another embodiment, the kit may include a drug used to treat the disorder in question, with or without the above materials that may be present to determine whether a subject has an inflammatory disease.

[0120] Administration of the Pharmaceutical Composition of the Invention Depsipeptidation of Formula (I), (II), (III) or (IV) as described herein The method of administration of the formulations of the present invention, including the combination, can be carried out by any of several methods well known in the art. These methods include local or systemic administration. Exemplary routes of administration include oral, parenteral, transdermal, intradermal, intramuscular, intraperitoneal, intravenous, subcutaneous, intranasal (e.g., nebulizer, inhaler, aerosol dispenser), intraocular (e.g., for treating conjunctivitis), intraaural (e.g., for treating ear infections), colorectal, rectal, vaginal, and any combination thereof. In addition, it may be desirable to introduce the pharmaceutical composition of the present invention into the central nervous system by any suitable route, including intraventricular and intrathecal injection. Intraventricular injection may be facilitated by an intraventricular catheter attached to a reservoir, such as an Ommaya reservoir. The method of introduction may also be provided by a rechargeable or biodegradable device, such as a depot. It is further contemplated that administration may also be carried out by coating a device, implant, stent, or prosthesis. The compounds of formula (I), (II), (III) or (IV) may also be used to coat catheters in any situation where a catheter is inserted into the body.

[0121] In another embodiment, the subject depsipeptide compound can be administered as part of a combination therapy with other drugs.Combination therapy refers to any administration form that combines two or more different therapeutic compounds, such that the second compound is administered while the previously administered therapeutic compound is still effective in the body (e.g., when two compounds are effective in a patient at the same time, the synergistic effect of the two compounds may be included).For example, different therapeutic compounds can be administered simultaneously or sequentially, either in the same formulation or in separate formulations.Thus, an individual who undergoes such treatment can have the combined effect of different therapeutic compounds.

[0122] For example, the depsipeptide compounds can be used in combination with other known antibiotics. The depsipeptide compounds of formula (I), (II), (III) or (IV) can be administered sequentially or substantially simultaneously. Altering the antibiotics can be beneficial to reduce the pathogen's ability to develop resistance to drugs. Non-limiting examples of antibiotics include penicillins (e.g., natural penicillins, penicillinase-resistant penicillins, antipseudomonas penicillins, aminopenicillins), tetracyclines, microlides (e.g., erythromycin), lincosamides (e.g., clindamycin), streptogramins (e.g., Synercid), aminoglycosides, and sulfonamides. In some embodiments, the depsipeptide compounds of formula (I), (II), (III) or (IV) are used in combination with compounds that target virulence factors, such as, but not limited to, phenol-soluble modulins. In some embodiments, the depsipeptide compounds of Formula (I), (II), (III) or (IV) are used in combination with compounds that target efflux pumps of said pathogens.

[0123] Kits and Articles of Manufacture Containing Pharmaceutical Compositions of the Compounds of the Invention Also included within the scope of the present invention is a kit comprising a compound of formula (I), (II), (III) or (IV), or a pharma- ceutically acceptable salt thereof, and instructions for its use. The term "kit" as used herein refers to a packaged product containing components to be administered together with a pharmaceutical composition comprising a compound of formula (I), (II), (III) or (IV), or a pharma- ceutically acceptable salt thereof, of the present invention for treating a disorder caused by a pathogen. The kit preferably comprises a box or container holding the components of the kit. The box or container is accompanied by a label or a Food and Drug Administration approved protocol. The box or container preferably holds the components of the present invention contained in a plastic, polyethylene, polypropylene, ethylene, or propylene container. The container can be a tube or bottle with a cap, or a bottle containing a dropper suitable for administering a solution containing the compound of the present invention dropwise, for example, into the ear or eye of a subject. The kit can also comprise a compound of formula (I), (II), (III) or (IV), or a pharma- ceutically acceptable salt thereof. In one particular embodiment, the kit can include (a) a pharmaceutical composition comprising a compound of Formula (I), (II) or (III) or a pharma- ceutically acceptable salt thereof, in a container, and (b) instructions describing how to use the pharmaceutical composition.

[0124] The kits may further contain one or more additional reagents, such as antibiotics, antifungals, antivirals, antiprotozoans, antiparasitics, antineoplastics, immunomodulators, antihypercholesterolemic agents, and combinations thereof. The kits typically include a label indicating the intended use of the contents of the kit. The term label includes any writing or recorded material supplied on or with the kit, or which otherwise accompanies the kit.

[0125] The present invention also provides a pharmaceutical pack or kit comprising one or more containers filled with a liquid formulation of a pharmaceutical composition comprising a compound of formula (I), (II), (III) or (IV), or a pharma- ceutically acceptable salt thereof. In one embodiment, the container filled with the liquid formulation of the present invention is a pre-filled syringe. In a particular embodiment, the formulation of the present invention is formulated as a sterile solution in a single-dose vial. For example, the formulation can be provided in a 3cc USP Type I borosilicate amber vial (West Pharmaceutical Services-Part No. 6800-0675) with a target volume of 1.2mL. Such container(s) can be accompanied by a notice in the form prescribed by a governmental agency that regulates the manufacture, use or sale of pharmaceutical or biological products, reflecting approval by the agency for manufacture, use or sale for administration to humans, as required.

[0126] Any pre-filled syringe known to those skilled in the art can be used in combination with the liquid formulation of the present invention. Pre-filled syringes that can be used include, but are not limited to, those described in PCT Publication Nos. WO05032627, WO08094984, WO9945985, WO03077976, and WO03077976. No. 5,607,400, US5,893,842, US7,081,107, US7,041,087, US5,989,227, US6,807,797, US6,142,976, US5,899,889, US20070161961A1, US20050075611A1, US20070092487A1, US20040267194A1, US20060129108A1. The prefilled syringe can be manufactured from various materials. In one embodiment, the prefilled syringe is a glass syringe. In another embodiment, the prefilled syringe is a plastic syringe. Those skilled in the art will understand that the nature and / or quality of the material used in the manufacture of the syringe can affect the stability of the compound formulation stored in the syringe. For example, it is understood that the silicon-based lubricant deposited on the inner surface of the syringe chamber can affect the formation of particles in the compound formulation. In one embodiment, the pre-filled syringe comprises a silicone-based lubricant. In one embodiment, the pre-filled syringe comprises a silicone-based content (comprises baked on silicone). In another embodiment, the pre-filled syringe does not comprise a silicone-based lubricant. It is also understood by those skilled in the art that a small amount of contaminant elements leaching from the syringe barrel, syringe tip cap, plunger or stopper into the formulation can affect the stability of the formulation. For example, it is understood that tungsten introduced during the manufacturing process can have a detrimental effect on the stability of the formulation. In one embodiment, the pre-filled syringe can contain tungsten at a level of more than 500 ppb. In another embodiment, the pre-filled syringe is a low-tungsten syringe. In another embodiment, the pre-filled syringe can contain tungsten at levels of about 500 ppb to about 10 ppb, about 400 ppb to about 10 ppb, about 300 ppb to about 10 ppb, about 200 ppb to about 10 ppb, about 100 ppb to about 10 ppb, about 50 ppb to about 10 ppb, or about 25 ppb to about 10 ppb.

[0127] The invention also encompasses the finished pharmaceutical product, which is packaged and labeled. The article of manufacture comprises a suitable unit dosage form in a suitable container or vessel, such as a glass vial, a pre-filled syringe, or other container that is hermetically sealed.In one embodiment, the unit dosage form is provided as a particulate-free sterile solution suitable for parenteral administration, comprising a pharmaceutical composition comprising a compound of formula (I), (II), (III) or (IV), or a pharmaceutically acceptable salt thereof.In another embodiment, the unit dosage form is provided as a sterile lyophilized powder suitable for reconstitution, comprising a pharmaceutical composition comprising a compound of formula (I), (II), (III) or (IV), or a pharmaceutically acceptable salt thereof.

[0128] In one embodiment, the unit dosage form is suitable for intravenous, intramuscular, intranasal, oral, topical or subcutaneous delivery. Thus, the present invention encompasses sterile solutions suitable for each delivery route. The present invention further encompasses sterile lyophilized powders suitable for reconstitution.

[0129] As with any pharmaceutical product, the packaging materials and containers are designed to protect the stability of the product during storage and shipping. Additionally, the products of the present invention include instructions for use or other informational material that informs the physician, technician, or patient how to properly prevent or treat the disease or disorder in question, as well as how and how often to administer the pharmaceutical product. In other words, the article of manufacture includes instruction means that indicate or suggest a dosing regimen, including, but not limited to, the actual dose, monitoring procedures, and other monitoring information.

[0130] Specifically, the present invention provides an article of manufacture comprising packaging material, such as a box, bottle, tube, vial, container, pre-filled syringe, nebulizer, inhaler, intravenous (iv) bag, envelope, etc., and at least one unit dosage form of a compound of the present invention contained within the packaging material, wherein the compound comprises a liquid formulation containing an antibiotic. The packaging material includes an instruction means indicating how the compound may be used to prevent, treat and / or manage one or more symptoms associated with a disease or disorder.

[0131] The present invention is further illustrated by the following examples, which should not be construed as limiting in any manner.Throughout this application, various patent documents, patent application documents, and publications are referenced.The entire disclosures of these patent documents, patent application documents, and publications are incorporated herein by reference.In the event that any inconsistency exists between the patent documents, patent application documents, and publications and this disclosure, this disclosure shall prevail. EXAMPLES

[0132] Example 1: Preparation of compound of formula (II) Bacterial isolates (ISO 18629) were cultured in a 10% R4 medium containing 0.125 g / L casein digest, 0.1 g / L potato starch, 1 g / L casamino acids, and 10% ethanol (100% R4 is 10 g / L glucose, 1 g / L yeast extract, 0.1 g / L casamino acids, 3 g / L L-proline, 10 g / L MgCl2.6H2, 4 g / L CaCl2 2H2O, 0.2 g / L K2SO4, 5.6 g / L TES). Grown on 2% Bacto agar supplemented with 0.56%, 1 mL of all trace elements made up to 1 liter with tap water, and adjusted to pH 7.0 with 1.0 M NaOH), one colony was homogenized and used to inoculate 40 mL of BP inoculum broth (15 g / L glucose, 10 g / L malt extract, 10 g / L soluble starch, 5 g / L yeast extract, 10 g / L casamino acids, 0.05 g / L calcium carbonate made up to 1 liter with tap water) in a 250 mL flask. After 4 days of fermentation at 28°C (250 rpm), 12.5 mL of the seed culture was used to inoculate 0.5 L of R4 production broth at 2.5% inoculum (v / v). Fermentation was continued for 7 days at 28°C (180 rpm) before harvesting.

[0133] The fermentation broth was centrifuged at 10,000 rpm for 20 minutes. The supernatant was washed with reversed phase polymerase The compound was adsorbed onto a resin (HP-20, Mitsubishi). After washing the column with water and 80% aqueous acetone, the compound was eluted with acetone. The acetone extract was dried under reduced pressure to leave a dark orange oil. Hexane was then added to the oil, and the mixture was then sonicated and centrifuged. The supernatant was then decanted, and the remaining residue was dried under reduced pressure. The residue was dissolved in DMSO and purified by RP-HPLC on a preparative C-18 column (HO / AcN (with 0.1% TFA), 10-100% can over 35 minutes). The fractions containing the compound of formula (I) or (II) were lyophilized to leave a white powder.

[0134] The two-dimensional structure of the compound of formula (II) is1 H, 13 The structures were determined by C, COSY, DEPT-135, HSQC and HMBC NMR experimental methods. They can be seen in Figures 1-6, respectively. 13 The C attribution is as follows: [Table 1]

[0135] The structure was further confirmed by MS / MS.

[0136] [Table 2]

[0137] All fragments in Table 2 are consistent with the proposed two-dimensional structure based on NMR data.

[0138] Standard Marfey analysis was used to determine the three-dimensional structure of the compounds. The Marfey standard was prepared using the conditions shown in Scheme 5, Bhusan, R.; Brueckner, H. This was achieved by reacting all amino acids possible to be present in the structure of the compound of formula II with L-FDLA in an array manner, as derived from Amino Acids 2004, Vol. 27, p. 247, the teachings of which are incorporated herein by reference. [ka]

[0139] The amino acids used were L-isoleucine, D-isoleucine, L-allo-isoleucine, D-allo-isoleucine, L-serine, D-serine, L-glutamic acid, D-glutamic acid, N-methyl-L-phenylalanine, N-methyl-D-phenylalanine, L-alanine, D-alanine, L-threonine, D-threonine, L-allo-threonine, D-allo-threonine, and all four diastereomers of enduracididine. In addition, L- and L-allo-enduracididine were synthesized as follows: Marfey standards were prepared of two separate mixtures of enduracidin and D- and D-allo-enduracidin. Hydrolysis of the compound of formula (II) was followed by isolation of peptide fragments, allowing further Marfey analysis to determine the stereochemistry and position of the isoleucine and serine.

[0140] A 6:1 mixture of (2S,4R) as the major diastereomer and (2S,4S) as the minor diastereomer. [ka]

[0141] To (2S,4R)-2-tert-butoxycarbonylamino-4-hydroxy-5-nitro-pentanoic acid tert-butyl ester (2.3 g, 6.9 mmol) in anhydrous methanol (30 mL) was added ammonium formate (4.4 g, 69 mmol) and 10% palladium on carbon (735 mg, 0.7 mmol) at room temperature under nitrogen atmosphere. The reaction was heated to 40° C. for 1 h, cooled and filtered through Celite. The filtrate was concentrated in vacuo to give an oily residue, which was dissolved in ethyl acetate. The organic layer was washed with a saturated aqueous solution of sodium bicarbonate, which was back-extracted with ethyl acetate. The combined ethyl acetate layers were combined, dried through a hydrophobic frit and concentrated in vacuo. The resulting oil was immediately dissolved in anhydrous acetonitrile and 1-H-(pyrazole)-1-N-tertbutoxycarbonylcarboxamidine (2.1 g, 10.4 mmol) was added at room temperature under nitrogen atmosphere. The reaction was stirred for 20 hours and then concentrated in vacuo. The residue was purified by silica gel chromatography using isohexane / ethyl acetate (5 / 1) as eluent to give the title compound as a viscous oil (2.1 g, 56%, 3 steps from Boc-Asp-OtBu). LCMS (m / z) = 547.3 [M+H], Tr = 3.45 min. 1 H NMR (300 MHz, CDCl3) δ 1.39 - 1.55 (m, 36H), 1.81 - 2.01 (m, 2H), 3.37 - 3.49 (m, 1H), 3.56 - 3.66 (m, 1H), 3.88 - 4.01 (m, 1H), 4.19 - 4.30 (m, 1H), 5.19 - 5.30 (m, 1H), 5.38 - 5.52 (m, 1H), 8.67 - 8.78 (m, 1H), 11.45 (s, 1H).

[0142] (S)-5-((S)-2-tert-butoxycarbonyl-2-tert-butoxycarbonylamino-ethyl)-2-[(E)-tert-butoxycarbonylimino]-imidazolidine-1-carboxylic acid tert-butyl ester (6:1 mixture of the major diastereomer and (2S,4R) as the minor diastereomer) [ka]

[0143] (2S,4R)-2-tert-butoxycarbonylamino-4-hydroxy-5-nitro A solution of tropentanoic acid tert-butyl ester (900 mg, 1.7 mmol) in anhydrous dichloromethane (63 mL) was cooled to -78°C and N-N'-diisopropylethylamine (1.5 mL, 8.2 mmol) was added under a nitrogen atmosphere. This was followed by the dropwise addition of trifluoromethanesulfonic anhydride (0.3 mL, 1.8 mmol) while maintaining the temperature below -76°C. After the addition, the reaction was stirred at -78°C for 4 hours, then warmed to 0°C and quenched with a saturated aqueous solution of sodium bicarbonate. The organic layer was separated, washed with 1M HCl (x1), dried through a hydrophobic frit and concentrated in vacuo. The residue was purified by silica gel chromatography using isohexane / ethyl acetate (1 / 2 then 0 / 1) as eluent to give the title compound as a viscous oil (740 mg, 85%). LCMS (m / z) = 529.5 [M+H], Tr = 2.38 min. 1 H NMR (300 MHz, CDCl3) δ 1.38 - 1.66 (m, 36H), 1.87 - 2.02 (m, 1H), 2.14 - 2.34 (m, 1H), 3.60 - 3.84 (m, 1H), 3.91 - 4.09 (m, 1H), 4.21 - 4.35 (m, 1H), 4.38 - 4.60 (m, 1H), 5.09 - 5.29 (m, 1H).

[0144] (2S,4S)-Enduracididine trifluoroacetate (6:1 mixture of the major diastereomer with (2S,4R) as the minor diastereomer) [ka]

[0145] A solution of (S)-5-((S)-2-tert-butoxycarbonyl-2-tert-butoxycarbonylamino-ethyl)-2-[(E)-tert-butoxycarbonylimino]-imidazolidine-1-carboxylic acid tert-butyl ester (740 mg, 1.4 mmol) in trifluoroacetic acid (20 mL) and water (7 mL) was stirred at room temperature for 20 h. The reaction was concentrated in vacuo and the resulting residue was co-evaporated from toluene (3 times). The viscous oil was dried under vacuum for 24 h and the resulting solid was triturated in diethyl ether. The solid was filtered and dried to give the title compound as an off-white solid (600 mg, 83%). LCMS (m / z) = 173.3 [M+H], Tr = 0.22 min. 1 H NMR (300 MHz, D2O) δ 1.99 - 2.13 (m, 1H), 2.20 - 2.33 (m, 1H), 3.34 - 3.43 (m, 1H), 3.83 (t, J = 9.8 Hz, 1H), 3.96 (t, J = 6.9 Hz, 1H), 4.17 - 4.31 (m, 1H).

[0146] (2R,4R)-enduracididine trifluoroacetate (6:1 mixture of the major diastereomer with (2R,4S) as the minor diastereomer) [ka]

[0147] A 6:1 mix of (2R,4R)-enduracididine trifluoroacetate as the major diastereomer and the minor (2R,4S) diastereomer was prepared in a similar manner to (2S,4S)-enduracididine trifluoroacetate.

[0148] (Example 2) Antibacterial spectrum The compound of formula (II) prepared according to Example 1 was subjected to a panel of bacterial species. The antibacterial spectrum of the compound is shown in Table 3. The addition of 10% serum had no effect on the minimum inhibitory concentration (MIC). As shown by the data presented in Table 3, the compound has excellent activity against Gram-positive pathogens, including resistant strains. The compound also has low potency against most Gram-negative bacteria, but good activity against H. influenzae, which opens the possibility of treating respiratory tract infections.

[0149] [Table 3]

[0150] Moreover, to date, no mutants of S. aureus resistant to the compounds of formula (II) have been identified. Data indicate that the resistance frequency is <10 -10 (Tested at 4× the MIC). After 17 days of serial passaging, no detectable resistance was observed.

[0151] Compounds of formula (II) were then tested (at Micromyx, Kalamazoo, Missouri) against 255 contemporary clinical isolates, including linezolid-, vancomycin-, and daptomycin-resistant / intermediately susceptible strains. The results of these experiments are summarized below in Tables 4-6.

[0152] [Table 4-1] [Table 4-2] [Table 4-3]

[0153] [Table 5]

[0154] [Table 6]

[0155] Compounds of formula (II) were tested against ten B. anthracis isolates in the BL-3 facility at the Southern Research Institute. Screening was accomplished using a microdilution (96-well) broth-based minimum inhibitory concentration (MIC) assay and included the following isolates of Bacillus anthracis: NR-36, NR-38, NR-41, NR-46, NR-411, NR-412, NR-413, NR-414, NR-415, and NR-3838. First, a first Mueller-Hinton broth containing 0.05% Tween-80 was added to each well. Test compounds diluted in Mueller-Hinton broth containing 0.05% Tween-80 were then added to the appropriate wells (0.1 mL / well) at twice the desired starting concentration and serially diluted 2-fold across the plate for a total of 12 concentrations (128-0.06 μg / mL). The plate was then dosed with 1.0×10 200 µl of the target concentration diluted in Mueller-Hinton broth containing 0.05% Tween-80. 6 CFU / mL of pathogen was inoculated (0.1 ml / well) and incubated overnight (16-20 hours). After incubation, plates were read visually and individual wells were scored for turbidity as either partially clearing or completely clearing. Tests were performed in duplicate and each test plate contained the following controls: i) medium only (sterile control), ii) organisms in medium (negative control), and iii) doxycycline (positive control). The MIC is recorded as the lowest concentration (μg / mL) of drug that visually inhibited the growth of the organism (Table 3). The above compounds were highly active and had broth MICs of 100 μg / mL. 90=0.125 μg / mL to ≦0.06 μg / mL, which was better than the comparative drug doxycycline (see Table 7).

[0156] [Table 7]

[0157] (Example 3) Cytotoxic activity, hemolytic activity, DNA binding, hERG inhibition, cytochrome P450 inhibitory activity, genotoxicity, plasma stability, protein binding Cytotoxic activity Cultures of NIH3T3 mouse embryonic fibroblasts (ATCC CRL-1658, in Dulbecco's modified Eagle's medium supplemented with 10% fetal bovine serum) and Hep-G2 (ATCC HB-8065TM, in Dulbecco's modified Eagle's medium supplemented with 10% fetal bovine serum) were grown for 3 days in a humidified atmosphere of 5% CO2 in air at 37°C. Cultures were trypsinized, counted, and seeded into wells of sterile 96-well flat-bottom plates (1500 cells / well / 100 μL for NIH3T3 and 4000 cells / well / 100 μL for Hep-G2). After 24 hours, the medium in each well was replaced with 100 μL of fresh medium (containing two-fold serial dilutions of the compound of formula (II) from 0.8 μg / mL to 100 μg / mL prepared according to Example 1, and no compound control) and incubated for 2 hours at 37° C. in humidified conditions with 5% CO in air. After an additional 48 hours of incubation at 37° C. in humidified conditions with 5% CO in air, the reporter (Promega CellTiter 20 μL of 96® AQueous One Solution Cell Proliferation Assay was added to the cells. After 2 hours of incubation, the plates were read at 490 nm using a Spectramax Plus Spectrophotometer. Viable cells can reduce the reporter, whereas dead cells cannot. The results showed that the compound of formula (II) was tested at concentrations up to 100 μg / mL and after 2 days of incubation, there was no loss of viability in NIH3T3 and Hep-G2.

[0158] hemolytic activity Fresh human red blood cells were washed with phosphate buffered saline (PBS) until the upper phase was clear after centrifugation (no visible red color detectable). The pellet was resuspended in PBS to a final OD600 of 24 and added to wells of a sterile 96-well U-bottom plate (99 μL per well). Compounds of formula (II) were serially diluted 2-fold in water supplemented with 0.05% Tween and 1 μL of each concentration was added to wells to give final test concentrations of compounds of formula (II) ranging from 0.003 to 200 μg / mL. After 1 hour at 37°C, cells were centrifuged at 1000×g for 5 minutes at 4°C. 10 μL of the supernatant was removed and added to 90 μL / well of PBS (in a 96-well plate). The contents of the wells were ... Mix and read plates at 450 nm using a Spectramax Plus Spectrophotometer. Results showed that there was no detectable hemolysis after 1 hour incubation with compounds of formula (II) tested at concentrations up to 200 μg / mL.

[0159] DNA binding Compounds of formula (II) were serially diluted 2-fold in water supplemented with 0.05% Tween (concentrations from 2.5 mg / mL to 4.9 μg / mL). Serial concentrations of compounds of formula (II) in 3 μl volumes were added to (a) 6 μl sheared salmon sperm DNA (10 mg / mL, sheared by passing 20 times through a syringe with a 20-gauge needle) or (b) 6 μL water. Control compounds were treated similarly. Actinomycin D was used as a positive control compound since it is known to bind to DNA, and vancomycin was used as a negative compound since it is known not to bind to DNA. After thorough mixing, 7.5 μL volumes were spotted onto a lawn of grown Staphylococcus aureus NCTC8325 cells. The zone of growth inhibition around the spot is measured after 16 hours of growth at 37° C. A decrease in the size of the zone of inhibition indicates loss of antibacterial activity due to binding to DNA. For example, the activity of 1 μg of actinomycin D was eliminated by the presence of 50 μg of DNA, but there was no effect of DNA on the activity of vancomycin. The results showed that there was no loss of growth inhibition of the compound of formula (II) in the presence of DNA (6.6 μg / μL), indicating that the compound of formula (II) does not bind to DNA.

[0160] hERG inhibition The compound of formula (II) also exhibited negligible inhibition of hERG in an in vitro patch clamp assay [IC 50 >100 μg / mL (approximately 1,000 × MIC)].

[0161] Briefly, the experiments were performed on an IonWorks™ HT instrument (Molecular Devices Corporation) that automatically performs electrophysiological measurements of 48 single cells simultaneously in a dedicated 384-well plate (PatchPlate™). The cells used were Chinese Hamster Ovary (CHO) cells stably transfected with hERG. Single-cell suspensions were prepared in extracellular solution (Dulbecco's phosphate-buffered saline with calcium and magnesium, pH 7-7.2) and an aliquot was automatically added to each well of the PatchPlate™. Cells were then placed over a small hole at the bottom of each well by applying a vacuum under the plate to form an electrical seal. The vacuum was applied through a single compartment common to all wells, which was filled with intracellular solution (buffered to pH 7.2 with HEPES). The resistance of each seal was measured via a common ground electrode in the intracellular compartment, and individual electrodes were placed in each of the upper wells. Electrical access to the cells was achieved by circulating the perforating agent amphotericin under the PatchPlate™ and then measuring the hERG current before compound application. Electrodes were placed in the extracellular compartment and a holding potential of -80 mV was applied for 15 seconds. hERG channels were then activated by applying a depolarizing step to +40 mV for 5 seconds, which was then clamped at -50 mV for 4 seconds to elicit the hERG tail current, followed by a return to -80 mV for 0.3 seconds.

[0162] Test compounds were then added to the top wells of the PatchPlate™ at various concentrations (0.008, 0.04, 0.2, 1, 5 and 25 μM). Test compounds were left in contact with the cells for 300 seconds, after which currents were recorded using the same voltage step protocol as in the pre-compound scan. Quinidine, an established hERG inhibitor, was included as a positive control.

[0163] The current after compound application was expressed as a percentage of the current before compound application and was linearly correlated with the concentration for each compound. When concentration-dependent inhibition was observed, the data were fit to the following equation, IC 50 Calculate the value. y=y max -y min +y min 1+(x / x 50 )s In the formula, y = (current after compound application / current before compound application) x 100, x = concentration, x 50 = concentration required to inhibit current by 50% (IC 50 ), s = the slope of the graph.

[0164] [Table 8]

[0165] Cytochrome P450 inhibitory activity Compounds of formula (II) were then tested for cytochrome P450 inhibitory activity. Briefly, liver microsomes were incubated in triplicate with 10 μM and 30 μM of compound at 37° C. Control incubations containing vehicle or reference inhibitor were performed in parallel with the test agents. Final assays contained test agent, the indicated concentrations of probe substrate, 2 mM NADPH, 3 mM MgCl2 in 50 mM potassium phosphate buffer, pH 7.4. Final microsome concentration was 0.5 mg / mL. Maximum solvent concentration in the final assay was ≦0.5% to minimize inhibition of cytochromes by solvent. Final addition of NADPH was used to start the assay. At the end of the 10 min incubation, the assay was stopped by adding acetonitrile containing internal standard, samples were centrifuged, and the amount of probe metabolite in the supernatant was determined by LC / MS / MS. The assay was performed and validated using the following probe substrates, probe metabolites and control inhibitors (Table 9).

[0166] [Table 9]

[0167] The test results of the compound of formula (II) are shown in Table 10. [Table 10]

[0168] The control compounds performed as expected (Table 11). [Table 11]

[0169] In summary, the compound of formula (II) at 10 μM (approximately 40×MIC) exhibited less than 16% inhibition of five of the six CYP isoenzymes tested and 32.9% inhibition of CYP2D6.

[0170] Genotoxicity Compounds of formula (II) were then tested for genotoxic potential in a standard in vitro micronucleus assay. Briefly, the in vitro micronucleus assay uses fluorescent cell imaging to assess cytotoxicity and quantify micronuclei, a well-accepted model for testing genotoxic carcinogens. The assay was performed using CHO-K1 cells in the presence or absence of Aroclor-treated rat liver S9 fraction, and compounds were tested in duplicate over a 10-point concentration range (0.004, 0.01, 0.04, 0.1, 0.4, 1.0, 4.0, 10, 40, 100 μM). No genotoxicity was observed for compounds of formula (II) in the in vitro micronucleus assay up to 100 μM (approximately 400×MIC) (+ / −S9).

[0171] No genotoxic potential was observed for compounds of formula (II) when computationally evaluated against a set of predictive models consisting of the following endpoints: Salmonella mutagenicity, E. coli mutagenicity, mouse lymphoma, in vitro chromosomal aberrations, and in vivo micronuclei. The computational evaluation was based on quantitative structure-activity relationship predictions performed using the Leadscope® Model Applier and Leadscope® Toxicity Databases (Leadscope Inc., Columbus, Ohio). Briefly, the genotoxicity QSAR models were developed by the U.S. Food and Drug Administration (FDA) Center for Drug Evaluation and Research (CDER) at the Department of Informatics and The model was developed by the Computational Safety Analysis Staff (ICSAS). The QSAR models were constructed using the Data Miner (PDM) software with all default settings. Most of the QSAR models were constructed from publicly available information. We include training set structures and conclusions as part of the model description. QSAR models are trained on molecular descriptors, including structural features and eight calculated properties. The structural features include Leadscope default hierarchical features (Leadscope features) and predicted scaffolds created with default settings [4, 5]. The eight calculated properties are parent molecular weight, aLogP, polar surface area, hydrogen bond acceptors, hydrogen bond donors, number of rotatable bonds, and Lipinski score (rule violation count).

[0172] plasma stability Compounds of formula (II) were then tested for plasma stability. First, a sensitive bioanalytical method was developed to detect compounds of formula (II) in plasma using HPLC coupled to a triple quadrupole mass spectrometer. Plasma samples were spiked with various concentrations of compounds of formula (II), extracted with acetonitrile and analyzed by LC / MS / MS. The lower limit of quantification in plasma was 8.5 ng / mL. Compounds of formula (II) were then incubated in duplicate at 5 μM with different plasma samples (mouse, rat, human and dog) at 37° C. At several time points (0, 20, 40, 80, 120 min), aliquots were removed from each experimental reaction and mixed with 3 volumes of ice-cold stop solution (methanol containing propranolol, diclofenac or other internal standards). The stopped reactions were incubated on ice for at least 10 min. Samples were then centrifuged to remove precipitated proteins and the supernatants were analyzed by LC / MS / MS to quantify remaining parent. In summary, the compound of formula (II) exhibited a long (≧150 min) plasma half-life in the plasma of all species tested.

[0173] Protein binding Compounds of formula (II) were then tested for protein binding by equilibrium dialysis. Briefly, the compounds were first incubated with rat plasma at 5 μM. The mixture was then dialyzed against 5% glucose (D5W) according to the manufacturer's instructions with a RED Device (Pierce) (dialysis membrane with 8,000 MW cutoff) and incubated on a shaker. At the end of the incubation, aliquots were collected from both the plasma and D5W sides, an equal volume of D5W was added to the plasma sample, and an equal volume of plasma was added to the D5W sample. Methanol (3 volumes) containing an internal standard was added to precipitate proteins and release the drug. After centrifugation, the supernatant was transferred to a new plate and the compounds were analyzed by LC / MS / MS. In summary, the compounds exhibited 82-85% plasma protein binding.

[0174] Example 4: Microsome stability The metabolic stability of the compound of formula (II) was then measured in rat liver microsomes (Invitrogen / Life Technologies, CA) by monitoring the disappearance of the compound over a 2 hour period. The compound was incubated with the microsomes and samples were taken at 0, 0.5, 1 and 2 hours. Samples were tested by LC / MS for growth inhibition of S. aureus. After 2 hours, the compound was only slightly decreased, whereas verapamil, used as a positive control, degraded to its expected metabolites. No significant decrease in antibacterial activity or extensive metabolite formation was observed for the compound over a 2 hour period.

[0175] Example 5: Specificity of action To gain insight into the mechanism of action and specificity of the compound of formula (II) prepared according to Example 1, macromolecular synthesis studies were carried out using S. aureus. The incorporation of tritium-labeled thymidine, uridine, leucine, or N-acetyl-glucosamine into DNA, RNA, protein, or peptidoglycan, respectively, was measured and correlated with the above relative to the rate of biosynthesis. The effect of the compound was investigated, and the results showed that the compound of formula (II) inhibited peptidoglycan synthesis with a potency comparable to that of vancomycin (see FIG. 7).

[0176] Example 6: Mouse Sepsis Study Compounds of formula (II) were tested against the clinical isolate S. aureus MRSA ATCC 33591 in a mouse sepsis protection assay (Frimodt-Moller, N., Knudsen, JD and Espersen, F. (1999) The mouse peritonitis / sepsis model. In: Zak, O., Sande, MA eds. Handbook of animal model infection San Diego: Academic Press, pp. 127-136, the teachings of which are incorporated herein by reference in their entirety). ), its in vivo bioavailability and 50% survival rate of infected mice after 48 hours (PD50 CD-1 female mice were infected with a bacterial suspension (3.28 × 10 7 The mice were infected with 0.5 mL of 0.1 CFU / mouse by intraperitoneal injection. One hour after infection, the mice were treated with the above compounds by a single intravenous dose. The results are shown in Table 12. The compound of formula (II) showed an excellent PD at 0.19 mg / kg. 50 This indicates that vancomycin PD 50 It is very useful in comparison.

[0177] [Table 12]

[0178] In summary, the compound demonstrated a superior 50% protective dose of 0.19 mg / kg compared to vancomycin (approximately 2.75 mg / kg) in a mouse sepsis protection model. The compound provided complete protection at a dose of 0.5 mg / kg.

[0179] Example 7: In vivo efficacy: Mouse thigh infection model The compound of formula (II) prepared in Example 1 was then tested against MRSA in a thigh infection model in neutropenic mice. This model is an excellent system for measuring the effect of drugs against pathogens, in which most of the host immune system is eliminated. This model is widely used to determine PK / PD parameters and estimate dosage. This model is also a good model for acute bacterial skin and skin structure infections (ABSSSI), a potential clinical indication based on the antibacterial spectrum of the compound.

[0180] Female CD-1 mice were pretreated with cyclophosphamide to make them neutropenic, delivering two consecutive doses of 150 mg / kg and 100 mg / kg 4 days and 1 day before infection. Bacteria were resuspended in sterile saline and adjusted to an OD of 0.1 at 625 nm (infection inoculum: 2.8 × 10 5CFU / mouse). Mice were injected with inoculum in a volume of 0.1 mL into the right thigh. Two hours after infection, mice were treated with a single dose of 1, 2.5, 5, 10, or 20 mg / kg of a compound of formula (II) by intravenous injection. Four mice were treated per dose concentration. One group of infected mice was euthanized and the thighs were processed for CFU to serve as treatment time control (T=Rx). 26 hours after infection, mice were euthanized by CO2 inhalation. The right thighs were aseptically removed, weighed, homogenized, serially diluted, and plated on trypticase soy agar. Plates were incubated overnight at 37°C in 5% CO2, and CFU per gram of thigh was determined (see Table 13).

[0181] [Table 13]

[0182] The compound of formula (II) significantly reduced pathogen burden when administered at 2.5 mg / kg or greater by single IV injection.

[0183] Compounds of formula (II) were also tested in a thigh infection model (neutropenic mice) using MRSA as the infectious agent. Greater log CFU reductions from the starting infection load were observed when administered at 20, 10, 5, or 2.5 mg / kg (single iv dose in water). Infection was static at 1 mg / kg.

[0184] The above data demonstrate low resistance development, together with safety and an attractive mode of action, which make the compounds of formula (II) attractive candidates for therapeutic development.

[0185] Example 8. In vivo efficacy: Mouse pneumonia model Compounds of formula (II) were then evaluated against Streptococcus pneumoniae in an immunocompetent mouse pneumonia model to determine the compound's potential to treat acute respiratory infection. Briefly, CD-1 mice were infected intranasally with Streptococcus pneumoniae ATCC6301 (UNT012-2) (1.5×10 6 CFU / mouse). The compounds were delivered intravenously at doses ranging from 0.5 to 10 mg / kg / dose at 24 and 36 hours post-infection. At 48 hours post-infection, treated mice were euthanized and lungs were aseptically removed and processed for CFU titers.

[0186] As shown in Table 14, the compound of formula (II) was highly effective in this model, demonstrating a nearly 6 log CFU reduction at 48 hours at 10 mg / kg / dose. A dose-response relationship for the compound of formula (II) was observed across the range of concentrations evaluated. 1, 2 and 3 log 10 The amounts of compound of formula (II) required to achieve a reduction in CFU were calculated to be 0.27, 0.48 and 0.75 mg / kg / dose, respectively [using Graph Pad Prizm (ver. 5.0f)]. The static dose of compound of formula (II) was estimated to be <0.5 mg / kg / dose. [Table 14]

[0187] In follow-up studies, a dose-response relationship for the compound of formula (II) was observed across the range of concentrations evaluated (10, 5.0, 2.5, 1.0 and 0.5 mg / kg BID). 10 The doses of the compound of formula (II) required to achieve a reduction in CFU were determined to be 0.27, 0.48 and 0.75 mg / kg / dose, respectively. The static dose of the compound of formula (II) was estimated to be <0.5 mg / kg / dose.

[0188] Doctrine of Equivalents Those skilled in the art will be able to recognize, or be able to ascertain using no more than routine experimentation, numerous equivalents to the specific embodiments specifically described herein which equivalents are intended to be encompassed by the following claims.

Claims

【Claim 1】 Microbial infection.

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