WO2024112631A1

WO2024112631A1 - Streptogramin a monotherapy for treating or preventing bacterial infections

Info

Publication number: WO2024112631A1
Application number: PCT/US2023/080469
Authority: WO
Inventors: Kara S. Keedy; Laurene WANG
Original assignee: Aimmax Therapeutics, Inc.
Priority date: 2022-11-22
Filing date: 2023-11-20
Publication date: 2024-05-30

Abstract

Compositions, kits, packaged articles, methods and uses are described herein for treating a bacterial infection, or a disease associated with a bacterial infection, or a disease caused at least in part by a bacterial infection. The compositions, kits, packaged articles, methods and uses described herein include flopristin or a pharmaceutically acceptable salt thereof. Illustrative bacterial infections, diseases associated with a bacterial infection, diseases caused at least in part by bacterial infection include Staphylococcus spp., Streptococcus spp., Enterococcus spp., Haemophilus spp., Moraxella spp., Legionella spp., Chlamydia spp., Mycoplasma spp., Ureaplasma spp., Neisseria spp., Gardnerella spp., Bacillus spp., and Francisella spp. as well as other bacterial species infections.

Description

STREPTOGRAMIN A MONOTHERAPY FOR TREATING OR PREVENTING

BACTERIAL INFECTIONS

GOVERNMENT RIGHTS

[001] This invention was made with government support under R44AI170150 awarded by the National Institute of Allergy and Infectious Diseases. The government has certain rights in the invention.

CROSS-REFERENCE TO RELATED APPLICATIONS

[002] This application claims the benefit under 35 U.S.C. § 119(e) of United States Provisional Application Serial Number 63/427,433, filed November 22, the disclosure of which is incorporated herein by reference in its entirety.

TECHNICAL FIELD

[003] The invention described herein relates to the discovery of certain streptogramin A antibiotics that are useful in treating diseases without the necessary co-administration of a streptogramin B antibiotic. Illustrative diseases include those caused, at least in part by, Staphylococcus spp., Streptococcus spp., Enterococcus spp., Haemophilus spp., Moraxella spp., Legionella spp., Chlamydia spp., Mycoplasma spp., Ureaplasma spp., Neisseria spp., Gardnerella spp., Bacillus spp., and Francisella spp. as well as other bacterial species.

BACKGROUND

[004] The introduction of antibiotics once reduced human morbidity and mortality caused by infectious diseases dramatically. However, the widespread use of antibiotics has led to the emergence of antibiotic resistant pathogenic bacteria leading to loss or reduction in clinical efficacy and consequently the need for new antibiotic therapeutic options. Mechanisms of antibiotic resistance include efflux of antibiotics by transporters, prevention of interaction of antibiotics with target by mutation, modification and protection of target, and modification of antibiotics (Ogawara H “Comparison of Antibiotic Resistance Mechanisms in Antibiotic- Producing and Pathogenic Bacteria” Molecules 2019, vol. 24, no. 19, 3430).

[005] In 2019, the Centers for Disease Control and Prevention (CDC) released an update to their Antibiotic Resistance Threats in the United States report. The following pathogens were among those highlighted: drug-resistant Neisseria gonorrhoeae, methicillin-resistant Staphylococcus aureus (MRSA), drug-resistant Streptococcus pneumoniae, vancomycin- resistant Enterococci (VRE), erythromycin-resistant Streptococcus pyogenes, clindamycin- resistant Streptococcus agalactiae, and drug-resistant Mycoplasma genitalium (CDC: Antibiotic Resistance Threats in the United States, 2019. Atlanta, GA: CDC). N. gonorrhoeae, S. pneumoniae, MRSA, and VRE are also listed on the World Health Organizations (WHO) list of priority pathogens requiring new antibiotics (Tacconelli E, et al. “Discovery, research, and development of new antibiotics: the WHO priority list of antibiotic-resistant bacteria and tuberculosis” The Lancet Infectious Diseases 2018, vol. 18, no. 3, pages 318-327). Both of these lists serve to highlight public health priorities for the identification and development of new antibiotics.

[006] Streptogramins are a unique class of antibiotics remarkable for their antibacterial activity and their mechanism of action. These antibiotics are produced naturally as secondary metabolites by several Streptomyces species. They consist of two structurally different compound groups. Streptogramins A or M are macrolactones (polyunsaturated macrocyclic lactones) of which pristinamycin I IA (PIIA) is an example; while streptogramins B or S are cyclic hexadepsipeptides of which pristinamycin IA (PIA) is an example.

Pristinamycin IA (PIA) Pristinamycin I IA (PIIA) (Streptogramin B) (Streptogramin A)

[007] Streptogramins have demonstrated activity against gram-positive, certain gram-negative, and atypical bacteria as well as aerobic and anaerobic bacteria both in vitro and in vivo, including those with multi-drug resistance.

[008] Both groups of streptogramins bind to bacterial ribosomes and inhibit protein synthesis and they work synergistically against several bacterial species, and thus have been used in combination.

[009] Streptogramin A components inactivate the donor and acceptor sites of peptidyl transferases. They block two of the peptide chain elongation steps: aminoacyl-tRNA binding to the A site of ribosomes and peptide bond formation with peptidyl-tRNA at the P site.

[0010] Streptogramin B components have a mechanism of action that involves inhibition of peptide bond formation with release of incomplete peptide chains.

[0011 ] The synergy between the streptogramin A and B components is believed to result from conformational changes imposed upon the peptidyl transferase center by the binding of the streptogramin A component, and also by cooperative inhibition of both early and late stages of protein synthesis. The conformational change induced by the streptogramin A component increases ribosomal affinity for the streptogramin B component.

[0012] Only two streptogramin antibiotic combinations have been approved for use in humans for the treatment of bacterial infections. The first, pristinamycin, is a streptogramin antibiotic comprising pristinamycin IA and pristinamycin HA co-produced naturally by Streptomyces pristinaespiralis in a ratio by weight of about 30:70 and is used primarily in the treatment of staphylococcal infections, and to a lesser extent streptococcal infections. Pristinamycin is only available for use as a tablet for oral administration and due to its low oral bioavailability, it must be administered at high doses several times per day. Additionally, there is a need to improve efficacy of the composition as pristinamycin treatment failures have occurred.

[0013] Quinupristin-dalfopristin is the second streptogramin combination that has been approved for use in humans. The streptogramin B component, quinupristin, is a semi-synthetic derivative of pristinamycin IA. The streptogramin A component, dalfopristin, is a semi-synthetic derivative of pristinamycin IIA. Similar to pristinamycin, the quinupristin-dalfopristin components are combined in a B to A ratio by weight of approximately 30:70. It is available only in an intravenous injectable formulation with limited uses.

Quinupristin Dalfopristin

[0014] The streptogramin combination, linopristin-flopristin, was evaluated in a Phase 2 human clinical trial which demonstrated efficacy and safety for treating pneumonia infection; and was also evaluated in another clinical trial to treat bacterial skin infection. The streptogramin B component, linopristin, is another semisynthetic derivative of pristinamycin IA. The streptogramin A component, flopristin, is a semi-synthetic fluorinated derivative of pristinamycin I IB. However, the development of this streptogramin A+B combination did not continue past clinical Phase 2, and has not been approved for human use

Linopristin Flopristin

[0015] While streptogramins have been successful for treating infections under certain conditions, alternative, less complicated therapies that do not require the co-administration of both a streptogramin B and a streptogramin A compound are needed. In addition, new therapies are needed to address the constant threat and evolution of antibiotic resistance. Further, antibiotics that also have bactericidal activity are needed. Bactericidal activity is important to efficacy as a rapid elimination of the bacterial load during an infection reduces the potential for resistance development.

SUMMARY OF THE INVENTION

[0016] It has been unexpectedly discovered herein that flopristin is highly potent and efficacious in treating bacterial infections when used as a monotherapy; the co-administration of or combination with a streptogramin B is not necessary.

[0017] It has also been unexpectedly discovered that flopristin monotherapy is less likely to induce bacterial resistance than currently approved streptogramin cotherapy protocols. Streptogramin B included in currently approved cotherapies may lead to antibiotic pressure on or at the streptogramin B binding site of the bacterial ribosome resulting in bacterial resistance. Without being bound by theory, it is believed herein that resistance to streptogramin B may also negatively impact binding by other antibiotics, such as macrolides and lincosamides.

[0018] In one embodiment, a composition comprising, consisting essentially of, and/or consisting of flopristin or a pharmaceutically acceptable salt thereof is described.

[0019] In another embodiment, a pharmaceutical composition comprising, consisting essentially of, and/or consisting of flopristin or a pharmaceutically acceptable salt thereof, and one or more pharmaceutically acceptable carriers, diluents, or excipients, or a combination thereof, is described.

[0020] In another embodiment, a kit comprising, consisting essentially of, and/or consisting of (a) flopristin or a pharmaceutically acceptable salt thereof and one or more pharmaceutically acceptable carriers, diluents, or excipients, or a combination thereof, and (b) and instructions for administration to a subject to treat a bacterial infection is described.

[0021] In another embodiment, a packaged article comprising, consisting essentially of, and/or consisting of (a) flopristin or a pharmaceutically acceptable salt thereof and one or more pharmaceutically acceptable carriers, diluents, or excipients, or a combination thereof, and (b) and instructions for administration to a subject to treat a bacterial infection is described. Illustratively, the packaged article may be a blister pack.

[0022] In another embodiment, a method for treating or preventing a bacterial infection in a subject is described. The method includes administering a therapeutically effective amount of a composition or a pharmaceutical composition comprising, consisting essentially of, and/or consisting of flopristin or a pharmaceutically acceptable salt thereof to the subject.

[0023] In another embodiment, a method of treating or preventing a bacterial infection in a subject is described. The method includes administering to said subject a therapeutically effective amount of a pharmaceutical composition comprising flopristin or a pharmaceutically acceptable salt thereof. The infection or threat of infection may be caused by caused by one or more susceptible and/or resistant strains of gram-positive bacteria, gram-negative bacteria, atypical bacteria, aerobic bacteria, and/or anaerobic bacteria. [0024] In another embodiment, a method for treating or preventing a respiratory infection and/or disease in a subject is described. The method includes administering a therapeutically effective amount of a composition or a pharmaceutical composition comprising, consisting essentially of, and/or consisting of flopristin or a pharmaceutically acceptable salt thereof to the subject.

[0025] In another embodiment, a method for treating or preventing a sexually transmitted infection and/or a genitourinary tract infection in a subject is described. The method includes administering a therapeutically effective amount of a composition or a pharmaceutical composition comprising, consisting essentially of, and/or consisting of flopristin or a pharmaceutically acceptable salt thereof to the subject.

[0026] In another embodiment, a method for treating or preventing a skin and/or soft tissue infections and/or disease in a subject is described. The method includes administering a therapeutically effective amount of a composition or a pharmaceutical composition comprising, consisting essentially of, and/or consisting of flopristin or a pharmaceutically acceptable salt thereof to the subject.

[0027] In another embodiment, a method for treating or preventing an infection in a subject with cystic fibrosis is described. The method includes administering a therapeutically effective amount of a composition or a pharmaceutical composition comprising, consisting essentially of, and/or consisting of flopristin or a pharmaceutically acceptable salt thereof to the subject.

[0028] In another embodiment, a method for treating or preventing a bone and/or joint infection in a subject is described. The method includes administering a therapeutically effective amount of a composition or a pharmaceutical composition comprising, consisting essentially of, and/or consisting of flopristin or a pharmaceutically acceptable salt thereof to the subject.

[0029] In another embodiment, a method for treating or preventing endocarditis in a subject is described. The method includes administering a therapeutically effective amount of a composition or a pharmaceutical composition comprising, consisting essentially of, and/or consisting of flopristin or a pharmaceutically acceptable salt thereof to the subject.

[0030] In another embodiment, a method for treating or preventing bacteremia and/or sepsis in a subject is described. The method includes administering a therapeutically effective amount of a composition or a pharmaceutical composition comprising, consisting essentially of, and/or consisting of flopristin or a pharmaceutically acceptable salt thereof to the subject.

[0031] In another embodiment, a method for treating or preventing anthrax, tularemia, plague, glanders, and/or melioidosis in a subject is described. The method includes administering a therapeutically effective amount of a composition or a pharmaceutical composition comprising, consisting essentially of, and/or consisting of flopristin or a pharmaceutically acceptable salt thereof to the subject.

[0032] It is understood herein that each of the compounds, compositions, and pharmaceutical compositions are useful in the manufacture of one or more medicaments for treating and/or preventing any of the bacterial infections, disorders, and/or diseases described herein. BRIEF DESCRIPTION OF THE DRAWINGS

[0033] FIG. 1 . shows the time-to-kill kinetic activity of flopristin against multidrug-resistant A/. gonorrhoeae strains (A) WHO L, (B) WHO V, (C) WHO X, and (D) CDC 175 at 2X, 4X, and 8X the MIC. Bactericidal activity is defined as a >3-logio colony forming unit (CFU)/mL reduction from baseline.

[0034] FIG. 2. shows the intracellular activity of flopristin compared to ceftriaxone against three N. gonorrhoeae strains (A) ATCC 49226, (B) WHO L and (C) WHO X in an infected human cervical epithelial cell line when evaluated at 4X and 8X the MIC. Error bars represent standard deviation of three independent biological replicates. For WHO X, the lines showing the kill curve for flopristin at 4X and 8X the MIC are superimposable, indicating that both doses are bactericidal. Similarly, the lines for ceftriaxone at 4X and 8X the MIC against ATCC 49226 and flopristin at 4X and 8X the MIC against WHO L also overlap. Ceftriaxone is the current standard of care for gonorrhea.

[0035] FIG. 3. shows the in vivo efficacy of flopristin in a murine vaginal model of gonorrhea infection with N. gonorrhoeae strain FA0190. Flopristin was administered to groups of 5 ovariectomized and estradiol-treated mice as an intramuscular injection 2 hours post-infection either once at 2 hours post-infection or twice (BID) at 2 and 5 hours post-infection with 10 or 30 mg/kg. Two control groups were included, a 2-hour post-infection baseline control and a vehicle treated control arm. The bacterial load in vaginal lavage fluid was measured 26 hours postinfection and compared to the 2-hour baseline control. Error bars are standard error of the mean. Thresholds for 1-logw CFU/mL and 2-logw CFU/mL reduction from baseline are indicated along with the limit of detection for the assay. Treatments showing a statistically significant difference from the baseline control (p<0.05) are marked with

DETAILED DESCRIPTION

[0036] Several illustrative embodiments of the invention are described by the following clauses: [0037] A composition comprising flopristin or a pharmaceutically acceptable salt thereof for use in treating a bacterial infection, or a disease associated with a bacterial infection, or a disease caused at least in part by a bacterial infection.

[0038] A composition consisting essentially of flopristin or a pharmaceutically acceptable salt thereof for use in treating a bacterial infection, or a disease associated with a bacterial infection, or a disease caused at least in part by a bacterial infection.

[0039] A composition comprising a pharmaceutically active ingredient for use in treating a bacterial infection, or a disease associated with a bacterial infection, or a disease caused at least in part by a bacterial infection; wherein the pharmaceutically active ingredient consists of flopristin or a pharmaceutically acceptable salt thereof.

[0040] A kit or packaged article comprising flopristin or a pharmaceutically acceptable salt thereof, optionally formulated as a pharmaceutical composition, and a set of instructions for administering the flopristin or the pharmaceutically acceptable salt thereof in the treatment of a bacterial infection, or a disease associated with a bacterial infection, or a disease caused at least in part by a bacterial infection.

[0041] The composition, kit, or packaged article of any one of the preceding clauses wherein the compound or composition further comprises a pharmaceutically acceptable excipient, binder, solvent or carrier; optionally, where the composition is a solid, a semi-solid, a liquid, a solution, a suspension, or an aerosol; or optionally where the composition is a powder, granule, tablet, geltab, lozenge, rapid dissolving strip or tablet, capsule (hard or soft), caplet, syrup, elixir, suspension, emulsion, sachet, sprinkle, pellet, patch, spray, cream, ointment, gel, lotion or foam.

[0042] The composition, kit, or packaged article of any one of the preceding clauses wherein the composition is a sustained or controlled release dosage form, and/or configured for a prolonged and/or timed, pulsed administration at a predetermined rate.

[0043] A method for treating or preventing a bacterial infection in a subject, the method comprising administering to the subject a composition of any one of the preceding clauses. [0044] The composition, kit, packaged article, or method of any one of the preceding clauses, wherein the bacterial infection is an infection caused by one or more susceptible and/or resistant gram-positive bacteria, gram-negative bacteria, atypical bacteria, aerobic bacteria, and/or anaerobic bacteria, or a combination thereof..

[0045] The composition, kit, packaged article, or method of any one of the preceding clauses, wherein the bacterial infection is a sexually transmitted infection or a genitourinary tract infection.

[0046] The composition, kit, packaged article, or method of any one of the preceding clauses, wherein the bacteria comprise one or more susceptible and/or resistant strains of Neisseria gonorrhoeae, Chlamydia trachomatis, Mycoplasma genitalium or Gardnerella vaginalis, or a combination thereof.

[0047] The composition, kit, packaged article, or method of any one of the preceding clauses, wherein the bacteria comprise one or more resistant strains of Neisseria gonorrhoeae, Chlamydia trachomatis, Mycoplasma genitalium or Gardnerella vaginalis, or a combination thereof.

[0048] The composition, kit, packaged article, or method of any one of the preceding clauses, wherein the subject is a human, a host animal, a companion animal, or a livestock animal.

[0049] The composition, kit, packaged article, or method of any one of the preceding clauses, wherein the route of administration is oral, rectal, transmucosal, inhalation, intranasal, topical (including dermal, ophthalmic, otic), vaginal, intestinal, buccal, or sublingual administration; parenteral delivery, including intramuscular, subcutaneous, intravenous, or intramedullary injections, as well as intrathecal, epidural, direct intraventricular, intraperitoneal, intraamniotic, intranasal, or intraocular injections or injection into the joints.

[0050] The composition, kit, packaged article, or method of any one of the preceding clauses, wherein the bacterial infection is a respiratory infection.

[0051] The composition, kit, packaged article, or method of any one of the preceding clauses, wherein the bacterial infection is a respiratory infection selected from the group consisting of a community acquired pneumonia, healthcare associated pneumonia, chronic bronchitis, sinusitis, acute maxillary sinusitis, acute exacerbations of chronic bronchitis, pharyngitis, chronic obstructive pulmonary disease and or non-tuberculosis mycobacteria infection caused by or associated with bacteria.

[0052] The composition, kit, packaged article, or method of any one of the preceding clauses, wherein the bacteria comprise one or more susceptible and/or resistant strains of Streptococcus pneumoniae, Streptococcus pyogenes, Staphylococcus aureus, Haemophilus influenzae, Haemophilus parainfluenzae, Moraxella catarrhalis, Legionella spp., Chlamydia pneumoniae, Mycoplasma pneumoniae, Bordetella pertussis, Neisseria gonorrhoeae, Corynebacterium diphtheriae, Coxiella burnettii, Peptostreptococcus spp., Fusobacterium spp., Bacteroides spp., Prevotella spp., Nocardia spp., or Mycobacterium spp., or a combination thereof.

[0053] The composition, kit, packaged article, or method of any one of the preceding clauses, wherein the bacterial infection is a sexually transmitted infection or genitourinary tract infection. [0054] The composition, kit, packaged article, or method of any one of the preceding clauses, wherein the bacteria comprise one or more susceptible and/or resistant strains of Neisseria gonorrhoeae, Chlamydia trachomatis, Mycoplasma genitalium, Mycoplasma horn inis, Ureaplasma urealyticum, Ureaplasma parvum, or Gardnerella vaginalis, or a combination thereof.

[0055] The composition, kit, packaged article, or method of any one of the preceding clauses, wherein the bacterial infection is an infection of the skin and/or soft tissues.

[0056] The composition, kit, packaged article, or method of any one of the preceding clauses, wherein the bacteria comprise one or more susceptible and/or resistant strains of Staphylococcus aureus, Staphylococcus haemolyticus, Staphylococcus lugdunensis, Streptococcus agalactiae, viridans group streptococci, Streptococcus pyogenes, Enterococcus faecalis, Enterococcus faecium, Corynebacterium spp., or Propionibacterium acnes, or a combination thereof.

[0057] The composition, kit, packaged article, or method of any one of the preceding clauses, wherein the bacterial infection is a bone and/or joint infection.

[0058] The composition, kit, packaged article, or method of any one of the preceding clauses, wherein the bacteria comprise one or more susceptible and/or resistant strains of Staphylococcus aureus, Staphylococcus haemolytic! us, Staphylococcus lugdunensis, Streptococcus agalactiae, Streptococcus pyogenes, Streptococcus pneumoniae, viridans group streptococci, Salmonella spp., Shigella spp., Campylobacter spp., Yersinia spp., Enterococcus faecalis, Enterococcus faecium, Kingella kingae, Corynebacterium spp., Propionibacterium acnes, Chlamydia trachomatis or Neisseria gonorrhoeae, or a combination thereof. [0059] The composition, kit, packaged article, or method of any one of the preceding clauses, wherein the subject has endocarditis caused by or associated with the bacterial infection.

[0060] The composition, kit, packaged article, or method of any one of the preceding clauses, wherein the bacteria comprise one or more susceptible and/or resistant strains of Staphylococcus aureus, Staphylococcus haemolyticius, Staphylococcus lugdunensis, Streptococcus gallolyticus, viridans group streptococci, Enterococcus faecalis, Enterococcus faecium, Haemophilus spp., Kingella spp., Legionella spp., or Corynebacterium spp., or a combination thereof.

[0061] The composition, kit, packaged article, or method of any one of the preceding clauses, wherein the subject has bacteremia or sepsis caused by or associated with the bacterial infection.

[0062] The composition, kit, packaged article, or method of any one of the preceding clauses, wherein the subject has anthrax, tularemia, plague, glanders or melioidosis caused by or associated with the bacterial infection.

[0063] The composition, kit, packaged article, or method of any one of the preceding clauses, wherein the bacteria comprise one or more susceptible and/or resistant strains of Bacillus anthracis, Francisella tularensis, Yersinia pestis, Burkholderia mallei, or Burkholderia pseudomallei, or a combination thereof.

ABBREVIATIONS AND DEFINITIONS

[0064] All terms used herein generally have their ordinary meanings in the art, within the context of the invention, and in the specific context where each term is used, unless otherwise indicated. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains.

[0065] The term “minimum inhibitory concentration” (MIC) generally refers to the lowest concentration of a drug substance or active pharmaceutical ingredient that prevents visible growth of microorganisms including bacteria.

[0066] The term “minimum bactericidal concentration” (MBC) generally refers to the lowest concentration of a drug substance or active pharmaceutical ingredient that reduces the viability of the initial bacterial inoculum by >99.9%. MBC can be determined from broth dilution minimum inhibitory concentration (MIC) tests by subculturing to agar plates that do not contain the test agent.

[0067] The terms “treating” or “treatment” generally refer to administration of an effective amount of a therapeutic agent to a subject in need thereof to cure, alleviate, relieve, remedy, and/or ameliorate a disease. Such a subject can be identified by a health care professional based on results from any suitable diagnostic method. Treating or treatment also includes any approach for obtaining beneficial or desired results in a subject’s condition, including clinical results. Beneficial or desired clinical results can include, but are not limited to, reducing or eliminating in a subject a clinical symptom of bacterial infection; or delaying or preventing in a subject the onset of a clinical symptom of a bacterial infection, alleviation or amelioration of one or more symptoms or conditions, diminishment of the extent of a disease, stabilizing (i.e., not worsening) the state of disease, delay or slowing of disease progression, amelioration or palliation of the disease state, diminishment of the reoccurrence of disease, and remission, whether partial or total and whether detectable or undetectable.

[0068] The terms “prevent,” “preventative,” “preventive,” or “prophylactic” generally refer to a treatment that decreases the occurrence and/or severity of a disease or disease symptoms (e.g., infectious disease symptoms) in a subject. Such decreases may be complete (no detectable symptoms) or partial, such that fewer symptoms are observed than would likely occur absence of a preventive treatment. Preventive treatment also refers to pretreating a subject prior to exposure to an infectious agent.

[0069] The term “susceptible” bacteria strain generally refers to bacteria that are susceptible, meaning there is a high likelihood of therapeutic success using a dosing regimen of a conventional agent.

[0070] The term “resistant” bacteria strain generally refers to bacteria that are resistant, meaning there is a high likelihood of therapeutic failure using a dosing regimen of a conventional agent.

[0071] The terms “administer,” “administering”, “administration,” and the like generally refer to the methods that may be used to enable delivery of agents or compositions to the desired site of biological action.

[0072] The terms “host animal” and “subject” each generally refers to an animal, such as a mammal, that is the object of treatment, observation or experiment. Illustrative mammals include mice, rats, rodents, hamsters, gerbils, rabbits, guinea pigs, dogs, cats, sheep, goats, cows, horses, giraffes, elephants, tigers, lions, bears, platypuses, primates, such as monkeys, chimpanzees, apes, and humans. An animal can also be a bird, such as but not limited to chickens, turkeys, and other bird species.

[0073] The term “pharmaceutically acceptable” is employed herein to generally refers to those compounds, materials, compositions, formulations and/or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of animals, such as human beings, without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit/risk ratio.

[0074] The term “pharmaceutical composition” generally refers to a mixture of a compound or compounds disclosed herein with other chemical components such as diluents, binders, excipients or carriers. Pharmaceutical compositions may be used to make a dosage form. The pharmaceutical composition facilitates administration of the compound(s) to a subject.

[0075] The term “about”when used with numerical values or limits generally means that the number is approximate and that, as recited, it is understood to include a range of values. For example, a real number that is recited with a single significant figure, would by definition include a so-called rounding range; the number about 5 would at the very least include the range 4.5- 5.4, as each of those values rounds to 5. The same is to be understood for real numbers expressed with additional significant figures, where the corresponding rounding range applies to the last significant figure. Integers are to be understood to at least include the values ±1 for single-digit numbers, ±10 for two-digit numbers, etc. Depending upon the context and the variable recited, the term “about’’ is also interpreted to contemplate a range based on a percentage of the recited number, such as about 5 construed to include 5 ±10% or 5 ±20%. Notwithstanding the foregoing, it is understood that the range of values, unless otherwise indicated, should not be interpreted to include a negative range for a positively recited number, and vice-versa. In addition, depending up on the context, the recited number, unless otherwise indicated, should not be interpreted to include a value of zero when used in conjunction with an added component.

[0076] It is to be understood that in every instance disclosed herein, the recitation of a range of integers for any variable describes the recited range, every individual member in the range, and every possible subrange for that variable. For example, the recitation that n is an integer from 0 to 8, describes that range, the individual and selectable values of 0, 1 , 2, 3, 4, 5, 6, 7, and 8, such as n is 0, or n is 1 , or n is 2, etc. In addition, the recitation that n is an integer from 0 to 8 also describes each and every subrange, each of which may for the basis of a further embodiment, such as n is an integer from 1 to 8, from 1 to 7, from 1 to 6, from 2 to 8, from 2 to 7, from 1 to 3, from 2 to 4, etc.

[0077] It is also to be understood that unless otherwise indicated the recitation of a numerical value necessarily reflects the relative precision of the numerical value. For example, the recitation of a number with a specified precision based on significant figures necessarily includes a range of values that would match that number after appropriate rounding. For example, the recitation of the number 1 with a single significant figure is understood to properly refer to a range of values from 0.5 to 1.4. Similarly, the recitation of the number 1 .0 with two significant figures is understood to properly refer to a range of values from 0.95 to 1 .04. The relative precision of the numerical value can be further indicated by modifying with the term “about” to indicate that the modified number has lower precision.

[0078] The transitional phrase “consisting essentially of’ means that the scope of the corresponding composition, unit dose, method or use is understood to encompass the specified compounds or recited steps, and those that do not materially affect the basic and novel characteristics of the invention described herein. For example, a method described herein that consists essentially of administering a single listed compound is understood to represent a monotherapy for the recited disease. Though the monotherapy may include co-administration of one or more carriers, vehicles, diluents, adjuvants, excipients, and the like, and combinations thereof, and/or include co-administration of one or more additional active pharmaceutical ingredients, those latter additional active pharmaceutical ingredients are to be understood to be for treating diseases and/or symptoms distinct from treating the underlying conditions described herein, such as the treatment of the bacterial infection itself. Illustrative additional active pharmaceutical ingredients may include, for example, active ingredients for treating pain, inflammation, cough, congestion, and the like.

[0079] As used herein, the amount, dose, dosage or concentration of any pharmaceutical composition described herein is the sum of the amounts or concentrations of the corresponding free base of flopristin.

COMPOSITION

[0080] In an illustrative embodiment of the invention, a therapeutically effective amount or prophylactically effective amount of any pharmaceutical composition disclosed herein can be administered in a range from about 10 ng/kg body weight/day to about 200 mg/kg body weight/day whether by one or more administrations.

[0081] In some embodiments, a therapeutically effective amount or prophylactically effective amount of any pharmaceutical composition disclosed herein is administered in the range from about 100 ng to about 10,000 mg per individual administration.

[0082] In some embodiments, the unit dosage form may include at least 5 mg, at least 10 mg, at least 15 mg, at least 20 mg, at least 25 mg, at least 30 mg, at least 35 mg, at least 40 mg, at least 45 mg, at least 50 mg, at least 55 mg, at least 60 mg, at least 65 mg, at least 70 mg, at least 75 mg, at least 80 mg, at least 85 mg, at least 90 mg, at least 95 mg, at least 100 mg, at least 125 mg, at least 150 mg, at least 175 mg, at least 200 mg, at least 250 mg, at least 300 mg, at least 350 mg, at least 400 mg, at least 450 mg, at least 500 mg, at least 550 mg, at least 600 mg, at least 650 mg, at least 700 mg, at least 750 mg, at least 800 mg, at least 850 mg, at least 900 mg, at least 950 mg, at least 1 ,000 mg, at least 1 ,100 mg, at least 1 ,200 mg, at least 1 ,300 mg, at least 1 ,400 mg, at least 1 ,500 mg, at least 1 ,600 mg, at least 1 ,700 mg, at least 1 ,800 mg, at least 1 ,900 mg, or at least 2,000 mg of flopristin or a pharmaceutically acceptable salt thereof.

[0083] In some embodiments, a therapeutically effective amount or prophylactically effective amount of flopristin or pharmaceutically acceptable salt thereof in any pharmaceutical composition disclosed herein may be in the range of, e.g., about 5 mg to about 2,000 mg.

[0084] In some embodiments, a therapeutically effective amount or prophylactically effective amount of flopristin or pharmaceutically acceptable salt thereof in any pharmaceutical composition disclosed herein may be in the range of, e.g., about 0.5 mg to about 150 mg per kg body weight per day.

[0085] In liquid, solution, suspension, and semi-solid formulations, a concentration of any therapeutic composition disclosed herein typically may be between about 0.01 mg/mL to about 1 ,000 mg/mL.

[0086] In liquid, solution, suspension, and semi-solid formulations, a concentration of any therapeutic composition disclosed herein typically may be between about 1 mg/mL to about 800 mg/mL.

[0087] A therapeutically effective amount or prophylactical ly effective amount of any pharmaceutical composition disclosed herein may comprise a solvent, emulsion, suspending agent, vehicle, carrier or a diluent in an amount of, e.g., less than about 90% (v/v), less than about 85% (v/v), less than about 80% (v/v), less than about 75% (v/v), less than about 70% (v/v), less than about 65% (v/v), less than about 60% (v/v), less than about 55% (v/v), less than about 50% (v/v), less than about 45% (v/v), less than about 40% (v/v), less than about 35% (v/v), less than about 30% (v/v), less than about 25% (v/v), less than about 20% (v/v), less than about 15% (v/v), less than about 10% (v/v), less than about 5% (v/v), or less than about 1% (v/v).

[0088] A therapeutically effective amount or prophylactically effective amount of any pharmaceutical composition disclosed herein may comprise a solvent, emulsion, suspending agent, vehicle, carrier or other diluent in an amount in a range of, e.g., about 1% (v/v) to 90% (v/v).

[0089] The final concentration of any therapeutically effective or prophylactically effective pharmaceutical composition disclosed herein may be of any concentration desired. In an aspect of this embodiment, the final concentration of a pharmaceutical composition comprising, consisting essentially, or consisting of flopristin or pharmaceutically acceptable salt thereof may be therapeutically effective. The final concentration of the active ingredient in any pharmaceutical composition disclosed herein may be, e.g., at least 0.01 mg/mL, at least 0.1 mg/mL, at least 1 mg/mL, at least 10 mg/mL, at least 25 mg/mL, at least 50 mg/mL, at least 100 mg/mL, at least 200 mg/mL at least 500 mg/mL or at least 1000 mg/mL. In other aspects of this embodiment, the final concentration of a pharmaceutical composition in a pharmaceutical composition may be in a range of, e.g., about 0.01 mg/mL to about 1 ,000 mg/mL, about 0.1 mg/mL to about 1 ,000 mg/mL, about 1 mg/mL to about 1 ,000 mg/mL, about 10 mg/mL to about 1000 mg/mL, about 25 mg/mL to about 1000 mg/mL, about 50 mg/mL to about 1000 mg/mL, about 100 mg/mL to about 1 ,000 mg/mL, about 250 mg/mL to about 1000 mg/mL, about 500 mg/mL to about 1 ,000 mg/mL, about 750 mg/mL to about 1 ,000 mg/mL, about 0.01 mg/mL to about 750 mg/mL, about 0.1 mg/mL to about 750 mg/mL, about 1 mg/mL to about 750 mg/mL, about 10 mg/mL to about 750 mg/mL, about 25 mg/mL to about 750 mg/mL, about 50 mg/mL to about 750 mg/mL, about 100 mg/mL to about 750 mg/mL, about 250 mg/mL to about 750 mg/mL, about 500 mg/mL to about 750 mg/mL, about 0.01 mg/mL to about 500 mg/mL, about 0.1 mg/mL to about 500 mg/mL, about 1 mg/mL to about 500 mg/mL, about 10 mg/mL to about 500 mg/mL, about 25 mg/mL to about 500 mg/mL, about 50 mg/mL to about 500 mg/mL, about 100 mg/mL to about 500 mg/mL, about 250 mg/mL to about 500 mg/mL, about 0.01 mg/mL to about 250 mg/mL, about 0.1 mg/mL to about 250 mg/mL, about 1 mg/mL to about 250 mg/mL, about 10 mg/mL to about 250 mg/mL, about 25 mg/mL to about 250 mg/mL, about 50 mg/mL to about 250 mg/mL, about 100 mg/mL to about 250 mg/mL, about 0.01 mg/mL to about 100 mg/mL, about 0.1 mg/mL to about 100 mg/mL, about 1 mg/mL to about 100 mg/mL, about 10 mg/mL to about 100 mg/mL, about 25 mg/mL to about 100 mg/mL, about 50 mg/mL to about 100 mg/mL, about 0.01 mg/mL to about 50 mg/mL, about 0.1 mg/mL to about 50 mg/mL, about 1 mg/mL to about 50 mg/mL, about 10 mg/mL to about 50 mg/mL, about 25 mg/mL to about 50 mg/mL, about 0.01 mg/mL to about 25 mg/mL, about 0.1 mg/mL to about 25 mg/mL, about 1 mg/mL to about 25 mg/mL, about 10 mg/mL to about 25 mg/mL, about 0.01 mg/mL to about 10 mg/mL, about 0.1 mg/mL to about 10 mg/mL, about 1 mg/mL to about 10 mg/mL, about 0.01 mg/mL to about 1 mg/mL, about 0.1 mg/mL to about 1 mg/mL, or about 0.01 mg/mL to about 0.1 mg/mL.

[0090] A therapeutically effective amount of any pharmaceutical composition disclosed herein is capable of reducing the number of bacterial cells or severity of a bacterial infection in a subject suffering from a bacterial infection as compared to a subject not receiving the same treatment. [0091] In some embodiments, any pharmaceutical composition disclosed herein reduces an infection or prevents the occurrence of an infection by, e.g., at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95% or at least 99%.

[0092] In yet other aspects of this embodiment, any pharmaceutical composition disclosed herein reduces an infection by a range of, e.g., about 5% to about 100%, about 10% to about 100%, about 20% to about 100%, about 30% to about 100%, about 40% to about 100%, about 50% to about 100%, about 60% to about 100%, about 70% to about 100%, about 80% to about 100%.

[0093] Dosing can be single dosage or cumulative (serial dosing), and can be readily determined by one skilled in the art. For instance, treatment of a bacterial infection may comprise a one-time administration of an effective dose of any pharmaceutical composition disclosed herein.

[0094] In another embodiment, a one-time administration of a therapeutically effective amount of any pharmaceutical composition disclosed herein is described for treating sexually transmitted infections, including but not limited to /V. gonorrhea, Chlamydia, or Mycoplasma infections, and combinations thereof.

[0095] In another embodiment, a one-time administration of a therapeutically effective amount of any pharmaceutical composition disclosed herein is described for treating a N. gonorrhea infection. Illustrative therapeutic doses for the treating N. gonorrhea infection may be in the range from about 1 to about 120 mg per kg body weight per day.

[0096] In another embodiment, a one-time administration of a therapeutically effective amount of any pharmaceutical composition disclosed herein is described for treating a Chlamydia infection. Illustrative therapeutic doses for the treating a Chlamydia infection may be in the range from about 1 to about 120 mg per kg body weight per day.

[0097] In another embodiment, a one-time administration of a therapeutically effective amount of any pharmaceutical composition disclosed herein is described for treating a Mycoplasma infection. Illustrative therapeutic doses for the treating Mycoplasma infection may be in the range from about 1 to about 120 mg per kg body weight per day.

[0098] Alternatively, treatment of a bacterial infection may comprise multiple administrations of an effective dose of any pharmaceutical composition disclosed herein carried out over a range of frequency or duration, such as, e.g., daily, or every 2, 3, 4, 5, 6 or 7 days, or every 1 , 2, 3 or 4 weeks, or every 1 , 2, 3 or 4 months. It is to be understood that the initial dose or doses may be larger than a subsequent sustaining or maintenance dose or doses.

[0099] A therapeutically effective amount of any pharmaceutical composition disclosed herein may be administered daily every about 0.5 hour, about 1 hour, about 2 hours, about 3 hours, about 4 hours, about 5 hours, about 6 hours, about 7 hours, about 8 hours, about 9 hours, about 10 hours, about 11 hours, about 12 hours, about 13 hours, about 14 hours, about 15 hours, about 16 hours, about 17 hours, about 18 hours, about 19 hours, about 20 hours, about 21 hours, about 22 hours, or about 23 hours.

[00100] A therapeutically effective amount of any pharmaceutical composition disclosed herein may be administered, such as by injection or infusion, over a period of about 5 minutes, about 0.25 hour, about 0.5 hour, about 1 hour, about 2 hours, about 3 hours, about 4 hours, about 5 hours, about 6 hours, about 8 hours, about 10 hours, about 12 hours, about 16 hours, about 18 hours, about 20 hours, about 22 hours or about 24 hours, each over a range of frequency, such as, e.g., once daily, twice daily, thrice daily, four times daily, six times daily, eight times daily, and/or every 1 , 2, 3, 4, 5, 6 or 7 days, or every 1 , 2, 3 or 4 weeks. It is understood herein that in certain embodiments, the dosing protocol may consist of a single dose for treating certain bacterial infections.

[00101] The course of treatment of a therapeutically effective amount of any pharmaceutical composition disclosed herein may be for 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days, 10 days, 11 days, 12 days, 13 days, 14 days, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks, 9 weeks, 10 weeks, 11 weeks, 12 weeks, 4 months, 5 months, 6 months, 7 months, 8 months, 9 months, 10 months, 11 months, 12 months, or more. In a further embodiment, a period of during which administration is stopped is for 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days, 10 days, 11 days, 12 days, 13 days, 14 days, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, 8 weeks, 9 weeks, 10 weeks, 11 weeks, 12 weeks, 4 months, 5 months, 6 months, 7 months, 8 months, 9 months, 10 months, 11 months, 12 months, or more.

[00102] The therapeutic dose of any pharmaceutical composition disclosed herein may be administered by the route of oral, rectal, transmucosal, inhalation, intranasal, topical (including dermal, ophthalmic, otic), vaginal, intestinal, buccal, or sublingual administration; parenteral delivery, including intramuscular, subcutaneous, intravenous, or intramedullary injections, as well as intrathecal, epidural, direct intraventricular, intraperitoneal, intraamniotic, or intranasal administration, or by intraocular injections or injection into the joints.

[00103] It is to be understood that the administration dose, frequency and duration of the pharmaceutical compositions disclosed herein are determined by the type of active ingredients, together with various factors such as the disease to be treated, the specific bacteria involved in the infection, administration route, the types of excipients, the concentration of active agent, the types of formulation or composition, the bioavailability of the pharmaceutical compositions used, the subject's age, gender, and body weight, ethnicity, dietary habits, social economic status, personal hygiene, the disease severity or severity of the condition, other disease complications or any combination thereof.

[00104] It is to be understood that the timing of administration can vary from subject to subject, depending upon such factors as the severity of a subject's symptoms. For example, an effective dose of any pharmaceutical composition disclosed herein can be administered to a subject one or more times per day for an indefinite period of time, or until the subject no longer requires therapy. A person of ordinary skill in the art will recognize that the condition of the subject can be monitored throughout the course of treatment and that the effective amount of any pharmaceutical composition disclosed herein that is administered can be adjusted accordingly. [00105] Any pharmaceutical composition disclosed herein may reduce the frequency of a symptom of a disorder associated with a bacterial infection incurred over a given time period. Any pharmaceutical composition disclosed herein may reduce the frequency of a symptom of a disorder associated with a bacterial infection incurred over a given time period by, e.g., at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least

45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least

80%, at least 85%, at least 90%, at least 95%, or at least 100%.

[00106] Certain compounds disclosed herein can exist in unsolvated forms and solvated forms, including hydrated forms. In general, the solvated forms are equivalent to unsolvated forms and are encompassed within the scope of the present disclosure. Certain compounds disclosed herein may exist in multiple crystalline or amorphous forms. In general, all physical forms are equivalent for the uses contemplated by the present disclosure and are intended to be within the scope of the present disclosure.

PHARMACEUTICAL COMPOSITIONS AND FORMULATIONS

[00107] The term "pharmaceutically acceptable salts" generally refers to salts of the active compounds that are prepared with relatively nontoxic acids or bases, depending on the particular substituents found on the compounds described herein. When compounds disclosed herein contain sufficiently acidic functionalities, base addition salts can be obtained by contacting the neutral form of such compounds with a sufficient amount of the desired base, either neat or in a suitable inert solvent. When compounds of the present disclosure contain sufficiently basic functionalities, acid addition salts can be obtained by contacting the neutral form of such compounds with a sufficient amount of the desired acid, either neat or in a suitable inert solvent. Certain specific compounds disclosed herein may contain both basic and acidic functionalities that allow the compounds to be converted into either base or acid addition salts. [00108] The terms “pharmaceutically acceptable excipient" and "pharmaceutically acceptable carrier" refer to a substance that aids the administration of an active agent to and absorption by a subject or can facilitate the distribution or delivery of the active ingredients to the site of action and can be included in the compositions disclosed herein without causing a significant adverse toxicological effect on the subject. One of skill in the art will recognize that other pharmaceutical excipients are useful in the present disclosure.

[00109] The pharmaceutical compositions disclosed herein may further include a pharmaceutically acceptable carrier. For oral administration, the carrier may include, but is not limited to, a binder, a lubricant, a disintegrant, an excipient, a solubilizer, a dispersing agent, a stabilizer, an antioxidant, a suspending agent, a complexation agent, a surfactant, an emulsifying agent, a penetration enhancer, a thickening agent, a colorant, and/or a flavorant. For injectable preparations, the carrier may include a buffering agent, a preserving agent, an analgesic, a solubilizer, a co-solvent, an oil, a lipid, a fatty acid, an emulsifying agent, an isotonic agent, a biodegradable polymer, a hydrogel, viscosity modifier and/or a stabilizer. For preparations for topical administration, the carrier may include a base, an excipient, a lubricant, an emollient, a gel, an oil, a fatty acid, an ointment, a cream, an emulsion, a foam, viscosity modifier, a spray and/or a preserving agent.

[00110] The pharmaceutical compositions disclosed herein may be formulated into a variety of dosage forms in combination with the aforementioned pharmaceutically acceptable carriers. For example, for oral administration, the pharmaceutical composition may be formulated into tablets, troches, capsules (hard or soft), caplets, granules, powders, sachets, sprinkles, lozenges, rapid dissolving strips or tablets, elixirs, emulsion, suspensions, syrups or wafers. The pharmaceutical composition may be formulated into suppositories. For injectable preparations, the pharmaceutical composition may be formulated into an ampule or a pre-filled syringe as a single dosage form or a multidose container.

[00111] The pharmaceutical compositions disclosed herein may be formulated into a single dosage form suitable for the patient's body.

[00112] The pharmaceutical compositions disclosed herein may be used by blending with a variety of pharmaceutically acceptable carriers such as physiological saline or organic solvents, polymers, oils, lipids or fatty acids, surfactants or emulsifying agents, or hydrogel. In order to increase the stability or absorptivity, carbohydrates such as glucose, sucrose or dextrans, surfactants, antioxidants such as ascorbic acid or glutathione, chelating agents, low molecular weight proteins, polymers, or other stabilizers may be used.

[00113] In various embodiments, the pharmaceutical compositions disclosed herein can include, one or more preservatives and/or additives known in the art. Similarly, a pharmaceutical composition can further be formulated, without limitation, into any of various known delivery formulations. For example, in an embodiment, a pharmaceutical composition can include, surfactants, adjuvant, biodegradable polymers, hydrogels, etc., such optional components, their chemical and functional characteristics are known in the art. Similarly known in the art are pharmaceutical compositions that facilitate rapid, sustained or delayed release of the bioactive agents after administration. A formulation as described can be produced to include these or other formulation components known in the art.

[00114] Illustrative pharmaceutical formulations for parenteral administration, e.g., by bolus injection or continuous infusion, include aqueous solutions of the active compounds in water soluble form.

[00115] Illustrative pharmaceutical compositions for topical administration may be formulated as is known in the art for direct application to a target area. Forms conditioned for topical application may take the form, for example, of creams, milks, gels, powders, dispersion or microemulsions, eye drops, ear drops, nose drops, lotions thickened to a greater or lesser extent, impregnated pads, transdermal patches, ointments or sticks, aerosol formulations (e.g. sprays or foams), soaps, detergents, lotions, emollients, or cakes of soap. The pharmaceutical composition may further be formulated for topical administration in the mouth or throat. For example, the active ingredients may be formulated as a lozenge further comprising a flavored base, usually sucrose and acacia or tragacanth; pastilles comprising the composition in an inert base such as gelatin and glycerin or sucrose and acacia; and mouthwashes comprising the composition of the present invention in a suitable liquid carrier.

[00116] Illustrative formulations for oral administration include combining the active compounds with pharmaceutically acceptable carriers. Such carriers enable the compounds of the invention to be formulated as tablets, pills, dragees, capsules (soft or hard), caplets, liquids, solutions, gels, films, syrups, elixirs, slurries, sachets, powder, sprinkle, pellets, rapid dissolving strips or tablets, suspensions, emulsions and the like, for oral ingestion by a patient to be treated. Illustrative formulations for oral use can also be obtained by combining the active compounds with solid excipients, optionally grinding a resulting mixture, and processing the mixture of granules, after adding suitable auxiliaries, if desired, to obtain tablets or dragee cores. If desired, disintegrating agents may be added.

[00117] Additionally, the pharmaceutical composition may be formulated as sustained release, delayed release, and/or targeted release dosage forms and the like. The formulations may release the active agents, for example, in a particular part of the intestinal or respiratory tract, possibly over a period of time. Coatings, envelopes, and protective matrices may be made, for example, from polymeric substances, to provide sustained, delayed, and/or targeted release dosage forms.

[00118] The pharmaceutical compositions described herein can also be administered to the respiratory tract. For administration by inhalation or insufflation, the composition may be in the form of a nebulizing liquid or dry powder, for example, a powder mix of the therapeutic agent and a suitable powder base such as lactose or starch. Pharmaceutical compositions of the present invention can also be administered in an aqueous solution or nanoparticle suspensions when administered in an aerosol or inhaled form. Thus, other aerosol pharmaceutical formulations may comprise, for example, a physiologically acceptable buffered saline solution or suspension containing e.g., about 0.01 mg/mL to about 1 ,000 mg/mL of the active agents of the present invention specific for the indication or disease to be treated.

[00119] Drops or liquid spays, such as eye drops/sprays, ear drops/sprays, or nose drops/sprays, may be formulated with the active agents of the present invention in an aqueous or non-aqueous base also comprising one or more dispersing agents, solubilizing agents or suspending agents. Liquid sprays can be pumped, or are conveniently delivered from pressurized packs or containers. Drops can be delivered via a simple eye dropper-capped bottle, via a plastic bottle adapted to deliver liquid contents drop-wise, or via a specially shaped closure.

[00120] In an illustrative embodiment, the pH of any pharmaceutical composition described herein may be at least about 3, 3.5, 3.75, 4, 4.25, 4.5, 4.75, 5, 5.25, 5.5, 5.75, 6, 6.25, 6.5, 6.75, 7, 7.25, 7.5, 7.75, 8, 8.25, 8.5, 8.75, 9, 9.25, 9.5, 9.75, or 10. In another illustrative embodiment, the pH of any pharmaceutical composition described herein is in the range of pH from about 4 to about 10, from about 4.5 to about 9.5, from about 5 to about 9, from about 5.5 to about 8.5, from about 5.5 to about 8, from about 5.5 to about 7.5, from about 5.5 to about 7, from about 6 to about 7.5, or from about 6.5 to about 8.

[00121] In an embodiment, the pH of any pharmaceutical composition disclosed herein is from about 3.5 to about 9, about 3.5 to about 8, about 3.5 to about 7, about 3.5 to about 6, about 3.5 to about 5, about 3.5 to about 4, about 4 to about 9, about 4 to about 8, about 4 to about 7, about 4 to about 6, about 4 to about 5, about 5 to about 9, about 5 to about 8, about 5 to about 7, about 5 to about 6, about 6 to about 9, about 6 to about 8, about 6 to about 7, about 7 to about 9, about 7 to about 8, or about 8 to about 9.

METHOD OF TREATMENT AND PREVENTION

[00122] A therapeutically effective amount or prophylactically effective amount of flopristin or a pharmaceutically acceptable salt thereof, either within a single pharmaceutical composition (dosage form) or separately in divided pharmaceutical compositions (divided dosage forms), e.g., in a kit or packaged article, can be used in methods for treating or preventing a bacterial infection or a condition caused by a bacterial infection caused by, for example, one or more susceptible and/or resistant strains of gram-positive bacteria, gram-negative bacteria, atypical bacteria, aerobic bacteria, and/or anaerobic bacteria. Any of the pharmaceutical compositions, kits, or packaged articles described herein can be used in any of the methods.

[00123] In another embodiment, a method for treating or preventing a bacterial infection in a subject comprising, consisting essentially of, or consisting of administering to the subject a therapeutically effective amount of flopristin, or a pharmaceutical composition thereof, is described, wherein the bacterial infection is caused by one or more susceptible and/or resistant strains of gram-positive bacteria, gram-negative bacteria, atypical bacteria, aerobic bacteria, and/or anaerobic bacteria, such as, but not limited to, Bacillus spp., Bacteroides spp., Bordetella spp., Borrelia spp., Brucella spp., Burkholderia spp., Campylobacter spp., Chlamydia spp., Clostridium spp., Corynebacterium spp., Coxiella spp., Ehrlichia spp., Enterococcus spp., Francisella spp., Fusobacterium spp., Gardnerella spp., Haemophilus spp., Kingella spp., Legionella spp., Listeria spp., Moraxella spp., Mycoplasma spp., Mycobacterium spp., Neisseria spp., Nocardia spp., Peptostreptococcus spp., Porphyromonas spp., Propionibacterium spp., Prevotella spp., Rickettsia spp., Salmonella spp., Shigella spp., Staphylococcus spp., Streptococcus spp., Ureaplasma spp., Vibrio spp., or Yersinia spp.

[00124] In another embodiment, a method for treating or preventing a respiratory infection in a subject comprising, consisting essentially of, or consisting of administering to the subject a therapeutically effective amount of flopristin, or a pharmaceutical composition thereof, is described, wherein the respiratory infections and/or diseases may comprise community acquired pneumonia or healthcare associated pneumonia, chronic bronchitis, sinusitis, acute maxillary sinusitis, acute exacerbations of chronic bronchitis, pharyngitis, chronic obstructive pulmonary disease, and non-tuberculosis mycobacteria infection caused by or associated with bacteria, such as, but not limited to, one or more strains of susceptible and/or resistant Streptococcus pneumoniae, Streptococcus pyogenes, Staphylococcus aureus, Haemophilus influenzae, Haemophilus parainfluenzae, Moraxella catarrhalis, Legionella spp., Chlamydia pneumoniae, Mycoplasma pneumoniae, Bordetella pertussis, Neisseria gonorrhoeae, Corynebacterium diphtheriae, Coxiella burnettii, Peptostreptococcus spp., Fusobacterium spp., Bacteroides spp., Prevotella spp., Nocardia spp., or Mycobacterium spp.

[00125] In another embodiment, a method for treating or preventing a respiratory infection in a subject comprising, consisting essentially of, or consisting of administering to the subject a therapeutically effective amount of flopristin, or a pharmaceutical composition thereof, is described, wherein the respiratory infection may be caused by or associated with one or more strains of susceptible and/or resistant Moraxella catarrhalis and/or Haemophilus influenzae. [00126] In another embodiment, a method for treating or preventing a respiratory infection in a subject comprising, consisting essentially of, or consisting of administering to the subject a therapeutically effective amount of flopristin, or a pharmaceutical composition thereof, is described, wherein the respiratory infection may be caused by or associated with one or more strains of susceptible and/or resistant Legionella spp., Chlamydia pneumoniae, and/or Mycoplasma pneumoniae.

[00127] In another embodiment, a method for treating or preventing a sexually transmitted infection or genitourinary tract infection in a subject comprising, consisting essentially of, or consisting of administering to the subject a therapeutically effective amount of flopristin, or a pharmaceutical composition thereof, is described, wherein the sexually transmitted infection or genitourinary tract infection may be caused by or associated with one or more strains of susceptible and/or resistant Neisseria gonorrhoeae, Chlamydia trachomatis, Mycoplasma genitalium, Mycoplasma hominis, L/reaplasma urealyticum, Ureaplasma parvum, or Gardnerella vaginalis.

[00128] In another embodiment, a method for treating or preventing a sexually transmitted infection or genitourinary tract infection in a subject comprising, consisting essentially of, or consisting of administering to the subject a therapeutically effective amount of flopristin, or a pharmaceutical composition thereof, is described, wherein the sexually transmitted infection or genitourinary tract infection may be caused by or associated with one or more strains of susceptible and/or resistant Neisseria gonorrhoeae.

[00129] In another embodiment, a method for treating or preventing a sexually transmitted infection or genitourinary tract infection in a subject comprising, consisting essentially of, or consisting of administering to the subject a therapeutically effective amount of flopristin, or a pharmaceutical composition thereof, is described, wherein the sexually transmitted infection or genitourinary tract infection may be caused by or associated with one or more strains of susceptible and/or resistant Chlamydia trachomatis and/or Mycoplasma genitalium.

[00130] In another embodiment, a method for treating or preventing a skin and/or soft tissue infection in a subject comprising, consisting essentially of, or consisting of administering to the subject a therapeutically effective amount of flopristin, or a pharmaceutical composition thereof, is described, wherein the skin and/or soft tissue infections and/or disease may be caused by bacteria, such as, but not limited to, one or more strains of susceptible and/or resistant Staphylococcus aureus, Staphylococcus haemolyticus, Staphylococcus lugdunensis, Streptococcus agalactiae, viridans group streptococci, Streptococcus pyogenes, Enterococcus faecalis, Enterococcus faecium, Corynebacterium spp., or Propionibacterium acnes.

[00131] In another embodiment, a method for treating or preventing cystic fibrosis, and symptoms and complications thereof, in a subject comprising, consisting essentially of, or consisting of administering to the subject a therapeutically effective amount of flopristin, or a pharmaceutical composition thereof, is described, wherein the infection or threat of infection may be caused by or associated with bacteria, such as, but not limited to, one or more strains of susceptible and/or resistant Staphylococcus aureus, Haemophilus influenzae, or Mycobacterium spp. [00132] In another embodiment, a method for treating or preventing a bone and/or joint infection in a subject comprising, consisting essentially of, or consisting of administering to the subject a therapeutically effective amount of flopristin, or a pharmaceutical composition thereof, is described, wherein the infection or threat of infection may be caused by bacteria, such as, but not limited to, one or more strains of susceptible and/or resistant Staphylococcus aureus, Staphylococcus haemolyticius, Staphylococcus lugdunensis, Streptococcus agalactiae, Streptococcus pyogenes, Streptococcus pneumoniae, viridans group streptococci, Salmonella spp., Shigella spp., Campylobacter spp., Yersinia spp., Enterococcus faecalis, Enterococcus faecium, Kingella kingae, Corynebacterium spp., Propionibacterium acnes, Chlamydia trachomatis or Neisseria gonorrhoeae.

[00133] In another embodiment, a method for treating or preventing endocarditis in a subject comprising, consisting essentially of, or consisting of administering to the subject a therapeutically effective amount of flopristin, or a pharmaceutical composition thereof, is described, wherein the endocarditis or threat of endocarditis may be caused by bacteria, such as, but not limited to, one or more strains of susceptible and/or resistant Staphylococcus aureus, Staphylococcus haemolyticius, Staphylococcus lugdunensis, Streptococcus gallolyticus, viridans group streptococci, Enterococcus faecalis, Enterococcus faecium, Haemophilus spp., Kingella spp., Legionella spp., or Corynebacterium spp.

[00134] In another embodiment, a method for treating or preventing bacteremia or sepsis in a subject comprising, consisting essentially of, or consisting of administering to the subject a therapeutically effective amount of flopristin, or a pharmaceutical composition thereof, is described, wherein the infection or threat of infection may be caused by one or more strains of susceptible and/or resistant bacteria.

[00135] In another embodiment, a method for treating or preventing anthrax, tularemia, plague, glanders, or melioidosis in a subject comprising, consisting essentially of, or consisting of administering to the subject a therapeutically effective amount of flopristin, or a pharmaceutical composition thereof, is described, wherein the infection or threat of infection may be caused by bacteria, such as, but not limited to, one or more strains of susceptible and/or resistant Bacillus anthracis, Francisella tularensis, Yersinia pestis, Burkholderia mallei, or Burkholderia pseudomallei.

[00136] In another embodiment, a method for treating or preventing tularemia in a subject comprising, consisting essentially of, or consisting of administering to the subject a therapeutically effective amount of flopristin, or a pharmaceutical composition thereof, is described, wherein the respiratory infection may be caused by or associated with one or more strains of susceptible and/or resistant Francisella tualrensis.

[00137] In any of the methods for treating or preventing a bacterial infection, the subject can be a host animal, such as a companion animal, livestock or human. Illustrative companion animals include but are not limited to dogs, cats, birds, reptiles, and the like. Illustrative livestock include but are not limited to cows, sheep, goats, pigs, geese, turkeys, chickens, ducks, fish, and the like.

[00138] Illustratively, the administering step may consist of a single administration or may include a series of administrations. The dose, frequency and length of the treatment period depends on a variety of factors, such as the severity of the disease or condition, the age of the patient, the route of administration, the concentration of active agent, the bioavailability of the compositions and formulation used in the treatment, the subject’s age, gender/sex and body weight, the complication of the disease or a combination thereof. It will also be appreciated that the effective dosage of an agent used for the treatment may increase or decrease over the course of a particular treatment regimen. Changes in dosage may result and become apparent by standard diagnostic assays known in the art. In some instances, chronic administration may be required.

[00139] Preventive or prophylactic treatment methods include administering to a subject a therapeutically effective amount of an active agent or a combination of active agents. The administering step may consist of a single administration or may include a series of administrations. The dose, frequency and length of the treatment period depends on a variety of factors, such as severity of the disease or condition, the age of the patient, the route of administration, the concentration of active agent, the bioavailability of the compositions and formulation used in the treatment, the subject’s age, gender/sex and body weight, the complication of the disease. It will also be appreciated that the effective dosage of an agent used for the preventive or prophylaxis may increase or decrease over the course of a particular preventive or prophylaxis treatment regimen. Changes in dosage may result and become apparent by standard diagnostic assays known in the art. In some instances, chronic administration may be required.

[00140] An "effective amount" is an amount sufficient for a compound to accomplish a stated purpose relative to the absence of the compound (e.g. achieve the effect for which it is administered, treat a disease, reduce enzyme activity, increase enzyme activity, reduce binding to a target or receptor, reduce or block a signaling pathway, or reduce one or more symptoms of a disease or condition). An example of an "effective amount" is an amount sufficient to contribute to the treatment, prevention, or reduction of bacterial load, or symptoms of a disease, which could also be referred to as a "therapeutically effective amount".

[00141] The therapeutically effective amount can be initially determined from cell culture assays or pharmacokinetic studies. Target concentrations will be those concentrations of active compound(s) that are capable of achieving the methods of treatment of prevention described herein, as measured using the methods described herein or known in the art.

[00142] As is well known in the art, therapeutically effective amounts for use in humans can also be determined from animal models. For example, a dose for humans can be formulated to achieve a concentration that has been found to be effective in animals. The dosage in humans can be adjusted by monitoring compounds effectiveness and/or plasma concentrations and adjusting the dosage upwards or downwards, as described above. Adjusting the dose to achieve maximal efficacy in humans based on the methods described above and other methods is well within the capabilities of the ordinarily skilled artisan.

[00143] Dosages may be varied depending upon the requirements of the subject and the compound being employed. The dose administered to a subject, in the context of the present disclosure should be sufficient to affect a beneficial therapeutic response in the patient over time. The size of the dose also will be determined by the existence, nature, and extent of any adverse side-effects. Determination of the proper dosage for a particular situation is within the skill of the practitioner. T reatment may be initiated with smaller dosages which are less than the optimum dose of the compound. Thereafter, the dosage may be increased by small increments until the optimum effect under circumstances is reached. Dosage amounts and intervals can be adjusted individually to provide levels of the administered compound effective for the clinical indication being treated. This will provide a therapeutic regimen that is commensurate with the severity of the subject's disease state.

ROUTE OF ADMINISTRATION

[00144] Any of the pharmaceutical compositions or kits described herein can be administered in any of the routes of administration.

[00145] In an embodiment, a pharmaceutical composition as described herein can be administered by any suitable route, including, but not limited to oral, buccal, sublingual, parenteral, topical route, intravenous, intramuscular, intra-arterial, intramedullary, intramedullary, intraventricular, intrathecal, epidural, pulmonary, inhalation, transdermal, subcutaneous, intraperitoneal, intraamniotic, intranasal, intracolonic, ocular, intraocular, sublingual, vaginal, rectal administration, parental administration into a joint, and the like. It is understood that the route will vary with the condition and age of the recipient, and the disease being treated. Methods of determining the most effective means and dosage of administration are known to those of skill in the art and will vary, without limitation, with the pharmaceutical composition used for therapy, the purpose of the therapy, and the subject being treated. In an embodiment, for oral, rectal, vaginal, parenteral, pulmonary, sublingual and/or intranasal delivery formulations, tablets can be made by compression or molding, optionally with one or more accessory ingredients or additives. In an embodiment, compressed tablets are prepared, for example, by compressing in a suitable tableting machine, the therapeutic compounds in a free-flowing form such as a powder or granules, optionally mixed with a binder, lubricant, inert diluent, preservative, surfactants, disintegrant and/or surface-active or dispersing agent.

[00146] In an embodiment, molded tablets are made, for example, without limitation, by molding in a suitable tableting machine, a mixture of powdered compounds moistened with an inert liquid diluent. In an embodiment, the tablets may optionally be coated or scored, and may be formulated so as to provide slow or controlled release of the active ingredients. In an embodiment, tablets may optionally be provided with a coating, without limitation, such as a thin film, sugar coating, or an enteric coating to provide release in parts of the gut other than the stomach. In an embodiment, processes, equipment, and toll manufacturers for tablet and capsule making are well-known in the art.

[00147] In an embodiment, capsule pharmaceutical composition can utilize either hard or soft capsules, including, without limitation, gelatin capsules or vegetarian capsules. In an embodiment, a type of capsule is a gelatin capsule. In an embodiment, capsules may be filled using a capsule filling machine such as, without limitation, those available from commercial suppliers such as Miranda International or employing capsule manufacturing techniques well- known in the industry.

[00148] Packaging and instruments for administration may be determined by a variety of considerations, such as, without limitation, the volume of material to be administered, the conditions for storage, whether skilled healthcare practitioners will administer or patient selfcompliance, the dosage regime, the geopolitical environment (e.g., exposure to extreme conditions of temperature for developing nations), and other practical considerations.

[00149] Injection devices include pen injectors, auto injectors, safety syringes, injection pumps, infusion pumps, glass prefilled syringes, plastic prefilled syringes and needle free injectors syringes may be prefilled with liquid, or may be dual chambered, for example, for use with lyophilized material. Administration by injection may be, without limitation intravenous, intramuscular, intraperitoneal, or subcutaneous, as appropriate. Administrations by noninjection route may be, without limitation, nasal, oral, ocular, dermal, or pulmonary, as appropriate.

[00150] In certain embodiments, kits can comprise, without limitation, one or more single or multi-chambered syringes (e.g., liquid syringes and lyosyringes) for administering one or more pharmaceutical composition described herein. In various embodiments, the kit can comprise pharmaceutical composition for parenteral, subcutaneous, intramuscular or IV administration, sealed in a vial under partial vacuum in a form ready for loading into a syringe and administration to a subject. In this regard, the pharmaceutical composition can be disposed therein under partial vacuum. In all of these embodiments and others, the kits can contain one or more vials in accordance with any of the foregoing, wherein each vial contains a single unit dose for administration to a subject.

[00151] The kits can comprise lyophilizates, disposed as herein, that upon reconstitution provide pharmaceutical compositions in accordance therewith. In various embodiment the kits can contain a lyophilizate and a sterile diluent for reconstituting the lyophilizate.

[00152] The kit can comprise a flopristin or a pharmaceutically acceptable salt thereof. The kit can further comprise a package within which the pharmaceutical composition is contained with one or more of instructions, labels, containers and other items necessary for the use of the pharmaceutical composition to treat an infection and/or disease.

COMBINATION THERAPY

[00153] The term "combination therapy" generally refers to a composition or treatment protocol that includes administration of more than one active pharmaceutical agent, either simultaneously, contemporaneously, sequentially, or according to a predetermined pattern of dosing. The compounds in the combination therapy provided herein can be administered separately or can be coadministered together to the subject. Coadministration is meant to include simultaneous or sequential administration of the compounds individually or in fixed combination (more than one compound together).

[00154] It is understood herein that the combined amount of the two or more active components is the therapeutically effective dose. The total daily dose should be determined through appropriate medical judgment by a physician and administered once or several times. The specific therapeutically effective dose level, dosing frequency and duration for any particular subject may vary depending on various factors well known in the medical art, including the kind and degree of the response to be achieved, pharmaceutical compositions according to whether other agents are used therewith or not, the type for the formulation, the bioavailability of the formulation, the patient’s age, body weight, health condition, gender, and diet, the time and route of administration, the elimination rate of the pharmaceutical composition, the time period of therapy, other drugs used in combination or coincident with any of the pharmaceutical compositions disclosed herein, and like factors well known in the medical arts.

[00155] In various embodiments, the pharmaceutical compositions disclosed herein also can include, without limitation, two or more different therapeutic compounds for a single or multiple conditions. Use of multiple therapeutic compounds in a formulation can be directed to, for example, the same or different indications. Similarly, in another embodiment, multiple therapeutic compounds can be used in a formulation to treat, for example, both a pathological condition and one or more side effects caused by the primary treatment. In a further embodiment, multiple therapeutic compounds also can be included, without limitation, in a pharmaceutical composition as described herein to accomplish different medical purposes including, for example, simultaneous treatment and monitoring of the progression of the pathological condition. In an additional embodiment, multiple, concurrent therapies such as those exemplified herein as well as other combinations well known in the art are particularly useful for patient compliance because a single pharmaceutical composition can be sufficient for some or all suggested treatments and/or diagnoses. Those skilled in the art will know those therapeutic compounds that can be admixed for a wide range of combination therapies.

Similarly, in various embodiments, a first therapeutic compound can be used with a second or more therapeutic compound and combinations of one or more therapeutic compounds together with one or more other therapeutic compounds, including a small molecule (e.g., another antibiotic) or an antibody pharmaceutical. Therefore, in various embodiments a formulation is provided containing 1 , 2, 3, 4, 5 or 6 or more different therapeutic compounds, as well as, for one or more therapeutic compounds combined with one or more other therapeutic compounds. [00156] It is to be understood that such combination therapies include more than one active pharmaceutical ingredient for treating a bacterial infection, and/or additional active ingredients for treating comorbid diseases, symptoms, side effects, and the like accompanying bacterial infections.

EXAMPLES

Example 1. Antibacterial Profile of Flopristin Across Species including Drug Resistant Strains [00157] The antibacterial activity of flopristin was evaluated against various gram-positive, gramnegative, and atypical bacterial species including C. trachomatis, G. vaginalis, H. influenzae, M. catarrhalis, M. genitalium, N. gonorrhoeae, S. aureus, and S. pneumoniae.

[00158] Stock solutions of antibiotics were prepared in DMSO at 101X the final concentration. Eleven serial two-fold dilutions of test articles were made in DMSO and added to wells of a standard 96-well microdilution plate containing medium appropriate for the species being tested. Each row of the microtiter plate contained the two-fold dilution series in Columns 1 through 11. The final DMSO concentration was < 1%.

[00159] For C. trachomatis, which are obligate intracellular bacteria, in vitro susceptibility testing was performed using an intracellular based assay as previously described (Kohlhoff SA, Huband MD, Hammerschlag MR. In vitro activity of AZD0914, a novel DNA gyrase inhibitor, against Chlamydia trachomatis and Chlamydia pneumoniae. Antimicrobial agents and chemotherapy. 2014 Dec;58(12):7595-6). After incubation at 35°C for 72 h, cultures were fixed and stained for inclusions with fluorescein-conjugated antibody to the chlamydial lipopolysaccharide genus-specific antigen. The MIC was the lowest antimicrobial concentration at which no inclusions were seen.

[00160] For G. vaginalis, which are facultative anaerobes, minimum inhibitory concentrations (MICs) were determined by agar dilution as recommended by the Clinical and Laboratory Standards Institute (CLSI) for susceptibility testing of anaerobic bacteria (CLSI “Methods for Antimicrobial Susceptibility Testing of Anaerobic Bacteria, Approved Standard - Eleventh Edition” CLSI document M11 , 11^th Ed. CLSI, 950 West Valley Road, Suite 2500, Wayne, Pennsylvania 19087 USA, 2018). Supplemented Brucella agar plates containing 5% (v/v) laked sheep blood were utilized for susceptibility testing and plates were incubated in a Bactron 600 anaerobe chamber (Sheldon Manufacturing, Cornelius, Oregon) containing an atmosphere of 5% carbon dioxide, 5% hydrogen, and 90% nitrogen. MIC endpoints were determined at 72 hours for all G. vaginalis isolates with the MIC defined as the minimum concentration at which a marked reduction occurs in the appearance of growth on the test plate as compared to that of growth on the anaerobic control plate.

[00161] For testing of M. genitalium, CLSI methods for the susceptibility testing of mycoplasmas were followed (CLSI “Methods for antimicrobial susceptibility testing of human mycoplasmas” Approved Guideline M43-A. CLSI, 950 West Valley Road, Suite 2500, Wayne, Pennsylvania 19087 USA, 201 1) using SP4 broth prepared in the University of Alabama - Birmingham Diagnostic Mycoplasma Laboratory. Bacteria in microdilution plates were incubated at 37°C in an ambient air incubator and MIC values were read as the concentration of antimicrobial agent inhibiting visible color change in broth medium wells at the time when the organism control well first shows color change.

[00162] For H. influenzae and M. catarrhalis, MICs were determined following the CLSI broth microdilution procedure for fastidious bacteria (CLSI “Methods for Antimicrobial Dilution and Disk Susceptibility Testing of Infrequently Isolated or Fastidious Bacteria. 3^rd ed. CLSI Guideline M45. CLSI, 950 West Valley Road, Suite 2500, Wayne, Pennsylvania 19087 USA, 2016). After incubation for approximately 16 - 24 hours at 35°C in ambient atmosphere, plates were viewed from the bottom using a plate viewer. MIC values were read as the lowest concentration of test article where visible growth of the organism was completely inhibited.

[00163] For S. aureus, S. pneumoniae, and N. gonorrhoeae, MICs were determined using the CLSI broth microdilution procedure (CLSI “Performance Standards for Antimicrobial Susceptibility Testing. 28th ed.” CLSI supplement M100. CLSI, 950 West Valley Road, Suite 2500, Wayne, Pennsylvania 19087 USA, 2018; CLSI “Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria That Grow Aerobically; Approved Standard — 1 1th Edition” CLSI standard M07. CLSI, 950 West Valley Road, Suite 2500, Wayne, Pennsylvania 19087 USA, 2018). For testing of N. gonorrhoeae, a modified medium described by the American Type Culture Collection (ATCC) capable of supporting growth was used. This medium contains 15 g Oxoid Special Peptone, 1 g corn starch, 5 g NaCI, 4 g K₂HPO₄ and 1 g KH₂PO₄ per liter and 1% IsoVitaleX enrichment (Becton Dickinson). After incubation for approximately 20 hours at 35°C in ambient atmosphere for S. aureus and S. pneumoniae and in an atmosphere of 5% CO₂ for N. gonorrhoeae, plates were viewed from the bottom using a plate viewer. MIC values were read as the lowest concentration of test article where visible growth of the organism was completely inhibited.

[00164] The C. trachomatis tested included 7 reference strains from the ATCC and 3 clinical isolates, 2 that were obtained from cervical sites and 1 obtained from an infant eye.

[00165] The G. vaginalis tested included 30 clinical isolates collected during 1992-2020 from the United States and Italy.

[00166] The H. influenzae, M. catarrhalis, S. aureus, and S. pneumoniae tested included reference strains from the ATCC and clinical isolates from the Micromyx repository.

[00167] The M. genitalium tested included reference strains from the ATCC and clinical isolates from University of Alabama - Birmingham Diagnostic Mycoplasma Laboratory.

[00168] The N. gonorrhoeae tested included reference strains from the ATCC, the World Health Organization (WHO), and Centers for Disease Control and Prevention (CDC) Antimicrobial Resistance Isolate Bank collections as well as a clinical isolate from the Micromyx repository. [00169] The majority of strains selected for inclusion in the testing panels were antibiotic resistant.

[00170] The panel of H. influenzae included 8 strains that were beta-lactamase positive and resistant to ampicillin and 9 strains that were beta-lactamase negative, but ampicillin-resistant. Three strains were non-susceptible to the pleuromutilin, lefamulin, which binds to a region of the bacterial ribosome near to the flopristin binding site and one was non-susceptible to the macrolide, azithromycin.

[00171] Breakpoints have not been established for M. genitalium. However, among the 12 M. genitalium strains and isolates tested, 3 isolates had elevated MICs for azithromycin, moxifloxacin, and doxycycline. These isolates were recovered from 2 patients who had failed multiple treatments with azithromycin, moxifloxacin, and doxycycline, thus they are clinically considered to be resistant. The two isolates from the same patient were recovered 2 years apart.

[00172] The panel of N. gonorrhoeae strains selected were predominantly multi-drug resistant or extensively-drug resistant strains. These included isolates with resistance to penicillin, fluoroquinolones, tetracyclines, macrolides, and extended-spectrum cephalosporins.

[00173] Among the 15 S. aureus strains, 11 were resistant to methicillin, 10 were resistant to macrolides, and 2 were resistant to linezolid.

[00174] Among the 5 S. pneumoniae strains, 4 were resistant to macrolides and one of these was also resistant to penicillin and tetracycline.

[00175] Table 1 shows the antibacterial activity of flopristin against C. trachomatis, H. influenzae, G. vaginalis, M. catarrhalis, M. genitalium, N. gonorrhoeae, S. aureus, and S. pneumoniae. MICso (minimum concentration at which 50% of isolates are inhibited), MIC90 (minimum concentration at which 90% of isolates are inhibited) and MIC ranges were determined and presented in the table. Note that the assays used H. influenzae, M. genitalium, N. gonorrhoeae, S. aureus, and S. pneumoniae collections enriched for antibiotic resistant strains. Flopristin demonstrated antibacterial activity against all species with MIC values comparable to antibiotics marketed for treating various bacterial infections. Some of the most potent activity was observed against the sexually transmitted pathogens C. trachomatis, M. genitalium and N. gonorrhoeae, with MICgo values of 0.12 and 0.25 ng/ml_, respectively. In contrast, the streptogramin combination pristinamycin, which includes both streptogramin A and streptogramin B components, had MIC₉₀ values of 0.5 and 1 ng/mL, respectively, against the same panels.

Thus, flopristin alone is 4-fold more potent against these 2 species than pristinamycin.

Table 1 . Antibacterial Activity of Flopristin Across Bacterial Species

MICso: minimum concentration at which 50% of isolates are inhibited; MICgo: minimum concentration at which 90% of isolates are inhibited a. The MIC50 and MICg₀ values for S. pneumoniae were not calculated (NC) due to number of strains.

[00176] The sexually transmitted pathogens M. genitalium and C. trachomatis are neither grampositive nor gram-negative, but are considered “atypical” bacteria because they lack a cell wall. Due to the absence of the cell wall, antibiotics such as the beta-lactams, which target the bacterial cell wall, are inactive against atypical bacteria and thus the number of classes of antibiotics that have activity against M. genitalium and C. trachomatis are greatly reduced. [00177] The individual MIC data for flopristin against 12 M. genitalium are presented in Table 2 alongside comparators azithromycin, moxifloxacin, doxycycline, and pristinamycin. The 12 M. genitalium include ATCC reference strains and clinical isolates collected during 1980-2021 from the United Kingdom, Denmark, Japan, or the United States.

[00178] Flopristin exhibited potent activity against each isolate, with MICs ranging from 0.015 - 0.5 pg/mL and retained activity against 3 isolates with elevated MICs for azithromycin, moxifloxacin, and doxycycline (isolates 75956, 84634, and 84211), which were recovered from patients who had failed multiple treatments with doxycycline, azithromycin, and moxifloxacin. Against each isolate, flopristin alone was 2- to 33-fold more potent than the streptogramin combination pristinamycin, which consists of both streptogramin A and streptogramin B components.

Table 2. Activity of Flopristin against Mycoplasma genitalium

MIC: minimum inhibitory concentration

Example 2. Bactericidal Activity

[00179] Minimum bactericidal concentrations (MBCs) of flopristin were determined for the collection of 20 /V. gonorrhoeae strains summarized in Example 1. Following incubation of the 96-well plates and recording the broth microdilution MIC values, duplicate 10 pL aliquots from the MIC well and three wells above the MIC were spotted and dragged across the surface of a Chocolate agar plate (BD; Catalog No. 221267; Lot No. 1057195). Plates were left to dry before inverting and incubating as described above. The number of colonies per plate were counted manually. The sums of the counts for the two spotted aliquots were compared to the values in the appropriate table of rejection values published by CLSI (CLSI “Methods for determining bactericidal activity of antimicrobial agents; approved guideline. Document M26-A” CLSI, 950 West Valley Road, Suite 2500, Wayne, Pennsylvania 19087 USA, 1999). These values were based upon the cell density of the inoculum and the target viable count reduction of 99.9%; if the sum of the colonies was less than or equal to the value in the table, the concentration of drug in the sampled well was considered to be bactericidal. The MBC was then defined as the lowest concentration of agent to demonstrate a bactericidal effect. MBC:MIC ratios were determined and MBC:MIC ratios < 4 are indicative of bactericidal activity; ratios > 8 which are indicative of bacteriostatic activity.

[00180] In MBC assays, flopristin is bactericidal against N. gonorrhoeae, as determined by MBC:MIC ratios, which are all 1 or 2 against all (20 out of the 20) N. gonorrhoeae strains (Table 3).

Table 3. Bactericidal Activity of Flopristin against Neisseria gonorrhoeae

PEN: penicillin; TET: tetracycline; CIP: ciprofloxacin; AZM: azithromycin; CFM: cefixime; CRO: ceftriaxone; SPT: spectinomycin; R: resistant; I: Intermediate; MIC: minimum inhibitory concentration; MBC: minimum bactericidal concentration a. Resistance profile for WHO strains as published in Unemo et. al 2016 (Unemo, M, D Golparian, L Sanchez-Buso, Y Grad, S Jacobsson, M Ohnishi, MM Lahra, A Limnios, AE Sikora, T Wi, SR Harris. 2016. The novel 2016 WHO Neisseria gonorrhoeae reference strains for global quality assurance of laboratory investigations: phenotypic, genetic and reference genome characterization. The Journal of antimicrobial chemotherapy 71(11): p. 3096-3108). Resistance profile for CDC strains as reported by CDC. Resistance profile for MMX 6921 as reported by the testing laboratory (Micromyx, Inc.) b. MBC:MIC ratios < 4 are indicative of bactericidal activity.

[00181] The bactericidal activity of flopristin against the 10 C. trachomatis strains summarized in Example 1 was evaluated by determining MBCs using previously described methods (Kohlhoff SA, Huband MD, Hammerschlag MR. In vitro activity of AZD0914, a novel DNA gyrase inhibitor, against Chlamydia trachomatis and Chlamydia pneumoniae. Antimicrobial agents and chemotherapy. 2014 Dec;58(12):7595-6). After MICs were measured, the antibiotic-containing medium was aspirated. Cells were washed twice with phosphate-buffered saline and fresh antibiotic-free medium was added. The infected cells were then frozen at -70°C. A few days later, the cells were thawed, passed onto new cells, incubated at 35°C for 72 hours and then fixed and stained for inclusions with fluorescein-conjugated antibody to the chlamydial lipopolysaccharide genus-specific antigen. The MBC was defined as the lowest antibiotic concentration that resulted in no inclusions after passage.

[00182] In the MBC assays, flopristin demonstrated potent bactericidal activity with MBC values either the same or within one 2-fold dilution of the MIC for each strain.

Table 4. Bactericidal Activity of Flopristin against C. trachomatis

MBCso: minimum concentration at which 50% of isolates are killed; MBC₉₀: minimum concentration at which 90% of isolates are killed

Example 3. In vitro Time-Kill Kinetics

[00183] An in vitro time-kill assay allows a quantitative assessment of the bactericidal activity of an antibiotic at various timepoints following exposure to the antibiotic. The time-to-kill kinetics of the biological activity of flopristin and ceftriaxone was evaluated against multi-drug resistant N. gonorrhoeae strains WHO L, WHO V, WHO X, and CDC 175. WHO L and WHO X are both resistant to penicillin, tetracycline, and ciprofloxacin, and intermediate susceptible to azithromycin. WHO L has an elevated MIC for ceftriaxone, the only standard of care antibiotic for the treatment of gonorrhoeae, meeting the CDC’s Gonococcal Isolate Surveillance Project (GISP) “alert value” criteria. WHO X also has high level resistance to both ceftriaxone and cefixime. WHO V is resistant to penicillin, tetracycline, and ciprofloxacin and has high level resistance to azithromycin (MIC >256 pg/mL). CDC 175 is also resistant to azithromycin (MIC 16 pg/mL) and has intermediate susceptibility to penicillin and tetracycline.

[00184] For WHO L and X, baseline MIC values were measured in triplicate using the broth macrodilution (tube) methodology according to CLSI M07 guidelines (CLSI “Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria That Grow Aerobically; Approved Standard — 11th Edition” CLSI standard M07. CLSI, 950 West Valley Road, Suite 2500, Wayne, Pennsylvania 19087 USA, 2018). Fastidious broth was used as the testing media with a 4 mL total volume in each tube. Isolates were incubated for 24 hours at 35°C in a 5% CO₂ incubator for all compounds. MIC values corresponded to the lowest concentration tube with a lack of turbidity upon visual examination.

[00185] For WHO V and CDC 175, MIC values were obtained in triplicate using broth microdilution as described in Example 1.

[00186] Time-kill bactericidal activity assays were conducted according to methods described by CLSI (CLSI “Methods for determining bactericidal activity of antimicrobial agents; approved guideline. Document M26-A” CLSI, 950 West Valley Road, Suite 2500, Wayne, Pennsylvania 19087 USA, 1999). The strains were initially grown for 2 hours in in Fastidious Broth. Cultures were then diluted into tubes containing antimicrobial agents at 2X, 4X, and 8X the concentration of the modal baseline MIC in 4 mL of fastidious broth. Each testing event also included a growth control tube with no antimicrobial compound for each strain.

[00187] A sample of each tube was removed at 0, 2, 4, 8, and 24 hours of growth at 35°C after the addition of the compound. WHO V and CDC 175 had an additional timepoint at 1 hour. Each sample was serial diluted in saline and plated on GC agar with 1% Gonococcus-Haemophilus influenzae (GCHI) enrichment for CFU measurements.

[00188] Time-kill kinetic results for flopristin are shown in FIG. 1. Flopristin resulted in bactericidal activity (i.e., >3-logw CFU/mL) against all four multi-drug resistant N. gonorrhoeae strains at all concentrations evaluated including as low as 2X the MIC. Further, at 8X the MIC, flopristin was bactericidal within 8 hours of exposure against all 4 strains. Against WHO V, flopristin was bactericidal within 2 hours of exposure at 4X and 8X the MIC, and within 4 hours of exposure at 2X the MIC.

Example 4. Intracellular Activity

[00189] N. gonorrhoeae and C. trachomatis are intracellular pathogens. Although N. gonorrhoeae is also capable of surviving extracellularly in the human host, C. trachomatis is an obligate intracellular pathogen. Thus, the ability of an antibiotic to penetrate human cells and exert antibiotic activity against pathogens within the cells is an essential characteristic or criterion for an antibiotic to successfully treat a sexually transmitted infection caused by either /V. gonorrhoeae or C. trachomatis.

[00190] The intracellular activity of flopristin against N. gonorrhoeae in a human cervical epithelial cell line was evaluated in three N. gonorrhoeae strains (one susceptible strain, ATCC 49226, and two multidrug resistant strains, WHO L and WHO X) using an in vitro invasion assay. Ceftriaxone, the only first line treatment remaining for gonorrhea recommended by the CDC, was included as a comparator. Initial MICs were determined using the broth microdilution method as described in Example 1.

[00191] For cellular invasion assays, HeLa cells (Human cervix carcinoma, European Collection of Authenticated Cell Cultures (ECACC; London, UK, Lot No. 86090201) were seeded at approximately 2 x10⁵ cells/mL and were grown to 70 to 80% confluency in 24-well flat-bottom plates containing 1 mL/well of RPMI-glutamine medium supplemented with 10% (v/v) heat- inactivated fetal bovine serum (FBS; Gibco; Waltham, MA; Lot No. 2509971 RP). Enough wells were seeded to perform the assay in triplicate for each test concentration. Cells were incubated at 37°C with 5% CO₂ for 48 h, then washed three times in serum-free RPMI-1640 in advance of infection.

[00192] N. gonorrhoeae isolates for infection were grown on chocolate agar overnight, and organisms scraped and resuspended into 1 mL Dulbecco’s Phosphate Buffered Saline (DPBS; Sigma; Lot No. RNK9608). Bacteria were washed once by pelleting and resuspending in RPMI with 2% FBS at a concentration of 3.2 x 10⁷ CFU/mL. This bacterial suspension was then used to infect wells at a multiplicity of infection (MOI) of 100:1 in a volume of 1 mL per well. Plates were centrifuged for 5 minutes at 500 rpm to enhance bacteria-cell association, and then the cultures were incubated for 1 hour at 37°C with 5% CO₂. Cell-free controls and uninoculated controls were also included.

[00193] To remove non-adherent bacteria, cells were washed gently three times with serum-free RPMI, then 200 pg/mL gentamicin in RPMI-glutamine medium with 2% FBS was added, and cells were incubated for 1 h, at 37°C with 5% CO₂ to kill any remaining extracellular bacteria. Cells were washed gently three times with serum-free RPMI, then fresh RPMI-glutamine medium with 2% FBS was added containing the test articles at the desired concentrations (at 0, 4X and 8X the MIC) and incubated at 37°C with 5% CO₂ for 0, 3 and 24 hours.

[00194] Following incubation, cells were washed gently three times with 1 mL of DPBS, followed by addition of 1 mL of 1% saponin (Thermo Scientific; Ward Hill; MA, Lot No. 10229709) in PBS (Thermo; Lot No. 368065). The plates were incubated for 15 minutes at 37°C with 5% CO₂ to permeabilize and lyse the mammalian cells and the GC were diluted and plated onto chocolate II agar. Plates were incubated at 35°C in 5% CO₂ for 24 h to quantify viable intracellular bacteria. [00195] Flopristin demonstrated potent and rapid intracellular bactericidal activity against all 3 /V. gonorrhoeae strains (FIG. 2). Flopristin exhibited a rapid reduction in logw CFU/mL of all 3 strains of N. gonorrhoeae compared to baseline within as little as 3 hours of exposure. While both flopristin and ceftriaxone reduced the bacterial load of WHO X by ~1 logw CFU/mL at 3 hours, the reduction in bacterial load by flopristin against ATCC 49226 and WHO L was significantly greater than that by ceftriaxone at this early timepoint. Surprisingly, flopristin monotherapy is equally efficacious against the ceftriaxone susceptible strain ATCC 49226. Flopristin also demonstrates superior bactericidal activity at lower multiples of the MIC than ceftriaxone against the ceftriaxone-resistant strains WHO L and WHO X.

[00196] Following exposure to flopristin for 24 hours, the number of quantifiable bacterial colonies was below the limit of detection for the assay against all 3 strains at both 4X and 8X of the MIC. In contrast, while ceftriaxone exhibited intracellular activity against the susceptible ATCC strain at 24 hours, variable activity was observed with ceftriaxone against the multidrug resistant strains WHO-L and WHO-X at this late timepoint. WHO L possess a low-level resistance to ceftriaxone due to mutations in penicillin binding protein 2 (PBP2) with reduced binding affinity for ceftriaxone as well as an mtr^o mutation that leads an over-expressed MtrCDE efflux pump and WHO X possesses a high-level resistance to ceftriaxone due to a mosaic penA allele (Unemo M, Golparian D, Sanchez-Buso L, Grad Y, Jacobsson S, Ohnishi M, Lahra MM, Limnios A, Sikora AE, Wi T, Harris SR. The novel 2016 WHO Neisseria gonorrhoeae reference strains for global quality assurance of laboratory investigations: phenotypic, genetic and reference genome characterization. Journal of Antimicrobial Chemotherapy. 2016 Nov 1 ;71 (11):3096-108.).

[00197] The MICs and MBCs reported for C. trachomatis in Table 1 and Table 4, respectively, were obtained using an intracellular based susceptibility assay with an epithelial cell line (HEp-2 cells) as since C. trachomatis cannot survive in culture extracellularly. Thus, these data are also supportive of the ability of flopristin to enter into human cells and exert bactericidal antibacterial activity.

[00198] In summary, flopristin has potential to be an effective therapy in vivo in a subject infected with sexually transmitted pathogens such as N. gonorrhoeae and C. trachomatis inside the human cells.

Example 5. Lack of Potential for Resistance Development

[00199] Three N. gonorrhoeae strains, maintained as part of the World Health Organization (WHO) N. gonorrhoeae reference bank, were tested in this study (WHO-F, WHO-L, and WHO- X). WHO-F is susceptible to most antibiotics active against N. gonorrhoeae. WHO-L is ciprofloxacin, penicillin, and tetracycline resistant, intermediate susceptible to azithromycin, and possesses a low-level resistance to ceftriaxone. WHO-X is resistant to ciprofloxacin, penicillin, tetracycline, cefixime, and ceftriaxone, and has an intermediate susceptibility phenotype to azithromycin. [00200] Initial flopristin and azithromycin MICs were determined by testing each isolate in triplicate using agar dilution susceptibility testing methods as described in CLSI document M07 (CLSI “Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria That Grow Aerobically; Approved Standard — 11th Edition” CLSI standard M07. CLSI, 950 West Valley Road, Suite 2500, Wayne, Pennsylvania 19087 USA, 2018).

[00201] Agar plates were prepared with a GC agar base and 1% (v/v) IsoVitaleX containing 4X, 8X, and 16X the baseline agar dilution modal MIC for either flopristin or azithromycin. Direct colony inocula were prepared in sterile saline from isolated colonies on chocolate agar plates incubated for 18 hours at 35°C in 5% CO₂ The flopristin- and azithromycin-containing agar plates were inoculated with 0.1 mL and 1 mL aliquots of the inoculum suspension (targeting 1-2 x 10⁹ CFU/mL) and incubated at 35°C in 5% CO₂ for 48 hours. The starting inoculum concentration for each A/, gonorrhoeae strain were determined by performing serial 10-fold dilutions and plating on antimicrobial-free chocolate agar plates.

[00202] Following 48 hours of incubation, visible colonies were counted on each agar plate. The frequency of resistance was calculated for each N. gonorrhoeae isolate by dividing the number of viable colonies by the starting inoculum concentration. Presumptive resistant mutants were isolated and passaged twice on drug-free agar and subjected to confirmatory MIC testing.

Confirmatory MIC values were determined for all mutants and baseline isolates in parallel using the CLSI reference agar dilution methodology.

[00203] No resistant colonies were recovered on the flopristin plates at 4X, 8X, or 16X the baseline MIC, resulting in resistance frequencies at the limit of detection for the assay, or <5.62 x IO ¹⁰, among the 3 isolates of N. gonorrhoeae (Table 5). These data demonstrate that against N. gonorrhoeae, flopristin has a low potential for resistance development. In contrast, spontaneous resistant mutants were observed with all 3 strains following exposure to azithromycin.

[00204] On the azithromycin 4X MIC plates, 4 azithromycin-resistant colonies were observed for N. gonorrhoeae WHO F at a resistance frequency of 2.25 x 10’⁹. Mutant colonies were isolated and subjected to agar dilution MIC testing to confirm resistance. An 8-fold increase in the azithromycin MIC value was observed for all 4 selected resistant colonies.

[00205] N. gonorrhoeae WHO L azithromycin resistant colonies were observed on the 4X MIC plates (n=10; resistance frequency of 5.56 x 10’⁹). Mutant colonies were isolated and subjected to agar dilution MIC testing to confirm resistance. Four- to 32-fold increases of the azithromycin MIC values above baseline were observed among the 10 selected resistant colonies. In addition, on the 8X MIC plate (0.1 mL plated) for azithromycin, one resistant colony of N. gonorrhoeae WHO L was observed (resistance frequency of 1 .11 x 10’⁸). A 64-fold increase in azithromycin MIC was confirmed by the agar dilution method.

[00206] N. gonorrhoeae WHO X azithromycin-resistant colonies were observed on the 4X MIC plates (n=4; resistance frequencies ranging from 1 .12 x 10^-8 to 1 .69 x 10⁹). Mutant colonies were isolated and subjected to agar dilution MIC testing to confirm resistance and 16- to 32-fold increases of the azithromycin MIC values above baseline were observed for these resistant colonies.

[00207] In summary, the fact that flopristin did not show any spontaneous resistant mutants developing at any of the concentration levels in all 3 strains confirms its unique and desirable characteristics as an effective antibiotic to treat A/, gonorrhoeae infection.

Table 5. Spontaneous Mutation Frequencies of Flopristin and Azithromycin in N. gonorrhoeae

Values of <5.62x1 O'¹⁰ indicate no resistant mutants were isolated

Example 5. In Vivo Efficacy in Gonorrhea Vaginal Infection Model

[00208] The efficacy of flopristin was assessed in an N. gonorrhoeae vaginal infection model in mice. Ovariectomy was performed at 4 weeks of age to BALB/c mice (N=5 per group), aged 5 to 6 weeks. A small dorsal midline incision was made in anesthetized mice, and the abdominal cavity was accessed by performing a blunt puncture through the abdominal wall. The left ovary and attached fat pad were individually pulled out of the abdominal cavity. The fallopian tube was ligated, following the dissection of the left ovary. The fat pad and tissue were placed back into the abdominal cavity upon completion of surgery. The process was repeated for the right ovary. The period of surgical recovery and acclimation was at least 7 days. Mice were subcutaneously injected with estradiol solution at 0.23 mg/mouse 2 days before infection (Day -2) and on the day of infection (Day 0). To minimize the indigenous vaginal bacteria, from Day -2 until the end of the study mice were treated twice daily (BID) with streptomycin (1 .2 mg/mouse) and vancomycin (0.6 mg/mouse) by intraperitoneal injection along with trimethoprim sulfate at 0.4 mg/mL supplied in the drinking water.

[00209] On Day 0, the vaginas of anesthetized mice were rinsed with 50 mM Hepes (pH 7.4, 30 L) and then inoculated with a suspension of N. gonorrhoeae strain FA1090 (ATCC 700825) at 8.7 x 10⁴ CFU/mouse. The vehicle was administered BID at 2 and 5 hours post-infection. Flopristin at 10 and 30 mg/kg, were intramuscularly administered once at 2 hours or BID at 2 and 5 hours post-infection. One infected but untreated control group was sacrificed at 2 hours after infection for the initial bacterial counts.

[00210] Mice from the vehicle control and flopristin groups were euthanized by CO₂ asphyxiation and sacrificed at 26 hours after infection. Vaginal lavage was performed twice with 30 pL lavage buffer (GC broth containing 0.05% saponin, pre-warmed to room temperature) to recover vaginal bacteria with ~10 times pipetting, then the lavage samples from each animal were pooled in a total volume of 500 L. The bacterial counts (CFU/mL) in vaginal lavage fluid were measured and the difference in bacterial density between the baseline group (2 hour initial counts) and the treatment group were determined.

[00211] Statistical significance (p < 0.05) was assessed with one-way ANOVA followed by Dunnett’s method using the GraphPad Prism software version 8. Treatments producing a 1- log CFU/mL reduction from baseline were considered to demonstrate bactericidal efficacy. [00212] A single intramuscular injection of flopristin at 30 mg/kg resulted in bactericidal efficacy against N. gonorrhoeae in a vaginal gonorrhea infection model with a statistically significant 1- logio CFU/mL decrease from baseline in just 24 hours after treatment (FIG. 3). Adding a second 30 mg/kg intramuscular injection 3 hours after the first dose, further decreased the bacterial burden (1.7-log₁₀ CFU/mL reduction from baseline) with two mice in this group having bacterial counts below the limit of detection. At 10 mg/kg flopristin dosed once or twice, there was little reduction in the CFU/mL count compared to baseline when flopristin was dosed at 10 mg/kg with a single injection or two injections separated by 3h, illustrating a dose-response relationship for flopristin in this model.

OTHER EMBODIMENTS

[00213] Certain embodiments of the present invention are described herein, including the best mode known to the inventors for carrying out the invention. Of course, variations on these described embodiments will become apparent to those of ordinary skill in the art upon reading the foregoing description. The inventor expects skilled artisans to employ such variations as appropriate, and the inventors intend for the present invention to be practiced otherwise than specifically described herein. Accordingly, this invention includes all modifications and equivalents of the subject matter recited in the claims appended hereto as permitted by applicable law. Moreover, any combination of the above-described embodiments in all possible variations thereof is encompassed by the invention unless otherwise indicated herein or otherwise clearly contradicted by context.

[00214] Unless otherwise indicated, all numbers expressing a characteristic, item, quantity, parameter, property, term, and so forth used in the present specification and claims are to be understood as being modified in all instances by the term “about.” Unless indicated to the contrary, the numerical parameters set forth in the specification and attached claims are approximations that may vary. At the very least, and not as an attempt to limit the application of the doctrine of equivalents to the scope of the claims, each numerical indication should at least be construed in light of the number of reported significant digits and by applying ordinary rounding techniques. Notwithstanding that the numerical ranges and values setting forth the broad scope of the invention are approximations, the numerical ranges and values set forth in the specific examples are reported as precisely as possible. Any numerical range or value, however, inherently contains certain errors necessarily resulting from the standard deviation found in their respective testing measurements. Recitation of numerical ranges of values herein is merely intended to serve as a shorthand method of referring individually to each separate numerical value falling within the range. Unless otherwise indicated herein, each individual value of a numerical range is incorporated into the present specification as if it were individually recited herein.

[00215] All patents, patent publications, and other publications referenced and identified in the present specification are individually and expressly incorporated herein by reference in their entirety for the purpose of describing and disclosing, for example, the compositions and methodologies described in such publications that might be used in connection with the present invention. These publications are provided solely for their disclosure prior to the filing date of the present application. Nothing in this regard should be construed as an admission that the inventors are not entitled to antedate such disclosure by virtue of prior invention or for any other reason. All statements as to the date or representation as to the contents of these documents is based on the information available to the applicants and does not constitute any admission as to the correctness of the dates or contents of these documents.

[00216] In closing, it is to be understood that although aspects of the present specification are highlighted by referring to specific embodiments, one skilled in the art will readily appreciate that these disclosed embodiments are only illustrative of the principles of the subject matter disclosed herein. Therefore, it should be understood that the disclosed subject matter is in no way limited to a particular methodology, protocol, and/or reagent, etc., described herein. As such, various modifications or changes to or alternative configurations of the disclosed subject matter can be made in accordance with the teachings herein without departing from the spirit of the present specification. Lastly, the terminology used herein is for the purpose of describing particular embodiments only and is not intended to limit the scope of the present invention, which is defined solely by the claims. Accordingly, the present invention is not limited to that precisely as shown and described.

Claims

WHAT IS CLAIMED IS:

1. A composition comprising flopristin or a pharmaceutically acceptable salt thereof for use in treating a bacterial infection, or a disease associated with a bacterial infection, or a disease caused at least in part by a bacterial infection.

2. A composition consisting essentially of flopristin or a pharmaceutically acceptable salt thereof for use in treating a bacterial infection, or a disease associated with a bacterial infection, or a disease caused at least in part by a bacterial infection.

3. A composition comprising a pharmaceutically active ingredient for use in treating a bacterial infection, or a disease associated with a bacterial infection, or a disease caused at least in part by a bacterial infection; wherein the pharmaceutically active ingredient consists essentially of flopristin or a pharmaceutically acceptable salt thereof.

4. A kit or packaged article comprising flopristin or a pharmaceutically acceptable salt thereof, optionally formulated as a pharmaceutical composition, and a set of instructions for administering the flopristin or the pharmaceutically acceptable salt thereof in the treatment of a bacterial infection, or a disease associated with a bacterial infection, or a disease caused at least in part by a bacterial infection.

5. A method for treating or preventing a bacterial infection in a subject, the method comprising administering to the subject a composition comprising flopristin or a pharmaceutically acceptable salt thereof.

6. The composition, kit, packaged article, or method of any one of claims 1 to 5, wherein the composition further comprises a pharmaceutically acceptable excipient, binder, solvent or carrier.

7. The composition, kit, packaged article, or method of any one of claims 1 to 5, wherein the composition is a sustained or controlled release dosage form.

8. The composition, kit, packaged article, or method of any one of claims 1 to 5, wherein the bacterial infection is an infection caused by one or more gram-positive bacteria, gramnegative bacteria, atypical bacteria, aerobic bacteria, or anaerobic bacteria, or a combination thereof.

9. The composition, kit, packaged article, or method of any one of claims 1 to 5, wherein the bacterial infection is a sexually transmitted infection or a genitourinary tract infection.

10. The composition, kit, packaged article, or method of any one of any one of claims 1 to 5, wherein the bacteria comprise one or more strains of Neisseria gonorrhoeae, Chlamydia trachomatis, Mycoplasma genitalium or Gardnerella vaginalis, or a combination thereof.

11 . The composition, kit, packaged article, or method of any one of any one of claims 1 to 5, wherein the subject is a human, a host animal, a companion animal, or a livestock animal. The composition, kit, packaged article, or method of any one of any one of claims 1 to 5, wherein the route of administration is oral, rectal, transmucosal, inhalation, intranasal, topical (including dermal, ophthalmic, otic), vaginal, intestinal, buccal, or sublingual administration; parenteral delivery, including intramuscular, subcutaneous, intravenous, or intramedullary injections, as well as intrathecal, epidural, direct intraventricular, intraperitoneal, intraamniotic, intranasal, or intraocular injections or injection into the joints. The composition, kit, packaged article, or method of any one of claims 1 to 5, wherein the bacterial infection is a respiratory infection. The composition, kit, packaged article, or method of any one of claims 1 to 5, wherein the bacterial infection is a respiratory infection selected from the group consisting of a community acquired pneumonia, healthcare associated pneumonia, chronic bronchitis, sinusitis, acute maxillary sinusitis, acute exacerbations of chronic bronchitis, pharyngitis, chronic obstructive pulmonary disease and or non-tuberculosis mycobacteria infection caused by or associated with bacteria. The composition, kit, packaged article, or method of claim 14, wherein the bacteria comprise one or more strains of Streptococcus pneumoniae, Streptococcus pyogenes, Staphylococcus aureus, Haemophilus influenzae, Haemophilus parainfluenzae, Moraxella catarrhalis, Legionella spp., Chlamydia pneumoniae, Mycoplasma pneumoniae, Bordetella pertussis, Neisseria gonorrhoeae, Corynebacterium diphtheriae, Coxiella burnettii, Peptostreptococcus spp., Fusobacterium spp., Bacteroides spp., Prevotella spp., Nocardia spp., or Mycobacterium spp., or a combination thereof. The composition, kit, packaged article, or method of any one of claims 1 to 5, wherein the bacterial infection is a sexually transmitted infection or genitourinary tract infection. The composition, kit, packaged article, or method of claim 16, wherein the bacteria comprise one or more strains of Neisseria gonorrhoeae, Chlamydia trachomatis, Mycoplasma genitalium, Mycoplasma hominis, Ureaplasma urealyticum, Ureaplasma parvum, or Gardnerella vaginalis, or a combination thereof. The composition, kit, packaged article, or method of claim 16, wherein the bacteria comprise one or more strains of resistant Neisseria gonorrhoeae, Chlamydia trachomatis, Mycoplasma genitalium, Mycoplasma hominis, Ureaplasma urealyticum, Ureaplasma parvum, or Gardnerella vaginalis, or a combination thereof. The composition, kit, packaged article, or method of any one of claims 1 to 5, wherein the bacterial infection is an infection of the skin and/or soft tissues. The composition, kit, packaged article, or method of claim 19, wherein the bacteria comprise one or more strains of Staphylococcus aureus, Staphylococcus haemolyticus, Staphylococcus lugdunensis, Streptococcus agalactiae, viridans group streptococci, Streptococcus pyogenes, Enterococcus faecalis, Enterococcus faecium, Corynebacterium spp., or Propionibacterium acnes, or a combination thereof. The composition, kit, packaged article, or method of any one of claims 1 to 5, wherein the bacterial infection is a bone and/or joint infection infection. The composition, kit, packaged article, or method of claim 21 , wherein the bacteria comprise one or more strains of Staphylococcus aureus, Staphylococcus haemolyticius, Staphylococcus lugdunensis, Streptococcus agalactiae, Streptococcus pyogenes, Streptococcus pneumoniae, viridans group streptococci, Salmonella spp., Shigella spp., Campylobacter spp., Yersinia spp., Enterococcus faecalis, Enterococcus faecium, Kingella kingae, Corynebacterium spp., Propionibacterium acnes, Chlamydia trachomatis or Neisseria gonorrhoeae, or a combination thereof. The composition, kit, packaged article, or method of any one of claims 1 to 5, wherein the subject has endocarditis caused by or associated with the bacterial infection. The composition, kit, packaged article, or method of claim 23, wherein the bacteria comprise one or more strains of Staphylococcus aureus, Staphylococcus haemolyticius, Staphylococcus lugdunensis, Streptococcus gallolyticus, viridans group streptococci, Enterococcus faecalis, Enterococcus faecium, Haemophilus spp., Kingella spp., Legionella spp., or Corynebacterium spp., or a combination thereof. The composition, kit, packaged article, or method of any one of claims 1 to 5, wherein the subject has bacteremia or sepsis caused by or associated with the bacterial infection. The composition, kit, packaged article, or method of any one of claims 1 to 5, wherein the subject has anthrax, tularemia, plague, glanders or melioidosis caused by or associated with the bacterial infection. The composition, kit, packaged article, or method of claim 26, wherein the bacteria comprise one or more strains of Bacillus anthracis, Francisella tularensis, Yersinia pestis, Burkholderia mallei, or Burkholderia pseudomallei, or a combination thereof.