Antibiotic combination therapy

Abstract

The present invention provides antibiotic combination therapies for treating Acinetobacter baumannii infections in a subject. These combination therapies comprise rifabutin and a second antibiotic such as colistin and cefiderocol.

Description

[0001] Related Applications

[0002] This application claims the benefit and priority of U.S. Provisional Patent Application Serial No. 62 / 902,019, filed on September 18, 2019; U.S. Provisional Patent Application Serial No. 62 / 899,257, filed on September 12, 2019; U.S. Provisional Patent Application Serial No. 62 / 941,160, filed on November 27, 2019; and U.S. Provisional Patent Application Serial No. 62 / 977,659, filed on February 17, 2020, the content of each of these U.S. Provisional Patent Applications being incorporated herein by reference in its entirety. Technical Field

[0003] The present invention generally relates to a rifabutin combination therapy for treating Acinetobacter baumannii infections. Background Art

[0004] The emergence of multi-drug resistant (MDR) or extensively drug resistant (XDR) bacterial strains over the past few decades has made bacterial infections an increasingly serious public health problem. One bacterial species that poses a major threat to health is Acinetobacter baumannii, which can cause pneumonia, meningitis, and infections of the blood, urinary tract, and skin. Since Acinetobacter baumannii cells can survive on artificial surfaces for long periods of time, the bacteria are easily spread in hospital settings, and most Acinetobacter baumannii infections are hospital-acquired. For example, many soldiers in the Middle East were infected with Acinetobacter baumannii while being treated for injuries sustained during combat, and multi-drug resistant strains of the bacteria represent a serious complication in the recovery process of injured soldiers.

[0005] Treatment of Acinetobacter baumannii infections is challenging. Through the use of transposable genetic elements, Acinetobacter baumannii strains have developed resistance to several different classes of antibiotics, including aminoglycosides, aminocyclitols, tetracyclines, chloramphenicol, and carbapenems. Polymyxins such as colistin are usually used as a last resort due to their severe side effects, but some Acinetobacter baumannii strains are also resistant to colistin (Zubair et al., 2015). Thus, existing tools for the treatment and prevention of diseases caused by this bacterium are insufficient for many patients. To find solutions for treating these nosocomial pathogens, significant efforts have been made, and one of these solutions is combination therapy (Levin et al., 1999; Wood et al., 2003). Combinations of two antibiotics have shown different effects on each other, and in many cases, the effect is synergistic or enhanced, but in some cases, antagonistic effects have been observed (Montero et al., 2004; Tripodi et al., 2007). Rifampicin (an antibiotic belonging to the rifamycin class of antibiotics such as rifabutin) targets the bacterial DNA-dependent RNA polymerase B subunit (rpoB) and is an antibiotic that is often used in combination with other antibiotics. Rifampicin (also known as rifampin) has been shown to have a synergistic effect with colistin against Acinetobacter baumannii; however, the outcome of this combination depends on the MIC of rifampicin (Giannouli et al., 2012). Specifically, no synergistic effect of rifampicin and colistin was observed in Acinetobacter baumannii isolates in which the elevated MIC of rifampicin was caused by mutations in the rpoB target gene. Summary of the Invention

[0006] The present invention provides combination therapies for treating Acinetobacter baumannii infections, which combination therapies comprise rifabutin and a second antibiotic, which second antibiotic is, for example, a polymyxin (e.g., colistin, polymyxin B, polymyxin B nonapeptide; a polymyxin analog exemplified by MRX-8), other cationic antimicrobial peptides (e.g., SPR741; a chimeric peptide mimetic antibiotic exemplified by POL7306; octapeptin cyclic peptides) or cefiderocol. The present invention is based on the discovery that rifabutin acts synergistically with certain other antibiotics to inhibit Acinetobacter baumannii cell growth. The combination of antibiotics shows synergistic activity against a wide range of Acinetobacter baumannii strains. This synergistic activity greatly increases the susceptibility of Acinetobacter baumannii cells to rifabutin and colistin, where for some strains, the sensitivity to the antibiotic shows an increase of more than 500-fold when the antibiotic is used in combination with other antibiotics as compared to when the antibiotic is used alone. Additionally, and unexpectedly, the combination of rifabutin and colistin acts synergistically to inhibit the growth of strains that are resistant to both antibiotics when the two antibiotics are provided alone, and thus renders these strains susceptible to the combination therapy, even if the elevated MIC of rifabutin and rifampicin is due to a mutation in the rpoB gene. This observation stands in sharp contrast to what is observed in the case of the combination of rifampin and colistin, where strains with the mutation remain resistant to the combination. Accordingly, the present invention unlocks the therapeutic potential of antibiotics in environments where they were previously ineffective and provides an effective therapy for treating severe Acinetobacter baumannii infections.

[0007] In one aspect, the present invention provides a method of treating an Acinetobacter baumannii infection in a subject by administering rifabutin and a second antibiotic to the subject infected with Acinetobacter baumannii.

[0008] The second antibiotic can be a polymyxin (e.g., colistin, polymyxin B, polymyxin B nonapeptide) or cefiderocol.

[0009] The subject may be infected with Acinetobacter baumannii strains resistant to one or more antibiotics. The strain may be resistant to one or more of the following: aminocyclitol, aminoglycoside, β-lactam, β-lactamase inhibitor, carbapenem, cephalosporin, polymyxin, quinolone, rifamycin, sulfonamide, minocycline, eravacycline, sulbactam, and tetracycline. The strain may be resistant to one or more of the following: amikacin, trimethoprim-sulfamethoxazole, cefepime, cefiderocol, ceftazidime, chloramphenicol, ciprofloxacin, colistin, polymyxin B, doripenem, gentamicin, imipenem, levofloxacin, meropenem, penicillin, piperacillin, rifabutin, rifampicin, tazobactam, and tigecycline.

[0010] Each antibiotic can be administered by a separate route of administration. Two or more of these antibiotics can be administered by the same route of administration. Each antibiotic can be administered independently intravenously, orally, parenterally, subcutaneously, by inhalation, by injection, and / or by infusion.

[0011] Each antibiotic can be administered in the form of a separate formulation. Two or more of these antibiotics can be administered in the form of a single formulation. These antibiotics can be administered according to the same dosing regimen, or two or more antibiotics can be administered according to different dosing regimens. The dosing regimen can include one or more of dose, dose frequency, or the interval between doses.

[0012] The subject can be a human. The subject can be a pediatric, neonatal, infant, toddler, child, adolescent, preteen, young, adult, or elderly subject. The subject can be in emergency care, intensive care, neonatal intensive care, pediatric intensive care, coronary care, cardiothoracic care, surgical intensive care, medical intensive care, long-term intensive care, operating room, ambulance, field hospital, out-of-hospital scene environment.

[0013] The method can include providing one or more antibiotics in addition to the first two antibiotics (e.g., rifabutin and colistin or cefiderocol). The one or more additional antibiotics can be aminocyclitols, aminoglycosides, β-lactams, β-lactamase inhibitors, carbapenems, cephalosporins, polymyxins, quinolones, rifamycins, sulfonamides, minocycline, eravacycline, sulbactam, and tetracyclines. The one or more additional antibiotics can be amikacin, trimethoprim-sulfamethoxazole, cefepime, cefiderocol, ceftazidime, chloramphenicol, ciprofloxacin, colistin, doripenem, gentamicin, imipenem, levofloxacin, meropenem, penicillin, piperacillin, polymyxin B, rifabutin, rifampicin, tazobactam, and tigecycline.

[0014] In another aspect, the present invention provides a combination therapy comprising a therapeutically effective amount of rifabutin and a second antibiotic for treating Acinetobacter baumannii infection in a subject. The second antibiotic can be a polymyxin (e.g., colistin, polymyxin B, polymyxin B nonapeptide) or cefiderocol.

[0015] The subject may be infected with an Acinetobacter baumannii strain that is resistant to one or more antibiotics such as any of the antibiotics described above.

[0016] Each antibiotic can be administered by a separate route of administration. Two or more of these antibiotics can be administered by the same route of administration. Each antibiotic can independently be administered intravenously, orally, parenterally, subcutaneously, by inhalation, by injection, and / or by infusion.

[0017] Each antibiotic can be administered in the form of a separate formulation. Two or more of these antibiotics can be administered in the form of a single formulation. These antibiotics can be administered according to the same dosing regimen, or two or more antibiotics can be administered according to different dosing regimens. The dosing regimen can include one or more of dose, dose frequency, or interval between doses.

[0018] The subject can be a human or a class of humans such as any of those described above.

[0019] The combination therapy can include providing one or more additional antibiotics in a therapeutically effective amount such as any of the antibiotics described above.

[0020] In another aspect, the present invention provides the use of a combination comprising rifabutin and a second antibiotic for the preparation of one or more medicaments for treating Acinetobacter baumannii infection in a subject. The second antibiotic can be colistin or cefiderocol.

[0021] In an embodiment of this use, the subject may become infected with an Acinetobacter baumannii strain that is resistant to one or more antibiotics such as any of those antibiotics described above.

[0022] In an embodiment of this use, each antibiotic is administered by a separate route of administration. In an embodiment of this use, two or more of these antibiotics are administered by the same route of administration. In an embodiment of this use, each antibiotic is independently administered intravenously, orally, parenterally, subcutaneously, by inhalation, by injection, and / or by infusion.

[0023] In an embodiment of this use, each antibiotic is administered in the form of a separate formulation. In an embodiment of this use, two or more of these antibiotics are administered in the form of a single formulation. In an embodiment of this use, these antibiotics are administered according to the same dosing schedule. In an embodiment of this use, two or more of these antibiotics are administered according to different dosing schedules. The dosing schedule may include one or more of dose, dose frequency, or the interval between doses.

[0024] In an embodiment of this use, the subject can be a human or a member of a class of humans, such as any of those humans described above.

[0025] In an embodiment of this use, the combination includes one or more additional antibiotics such as any of those antibiotics described above. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] Figure 1 is an image of a 96-well plate checkerboard of Acinetobacter baumannii cells cultured with various concentrations of rifabutin and colistin.

[0027] Figure 2 is an image of a 96-well plate checkerboard of Acinetobacter baumannii cells cultured with various concentrations of rifabutin and cefiderocol. DETAILED DESCRIPTION

[0028] The present invention provides combination therapies for treating Acinetobacter baumannii infections in a subject. These combination therapies are based on the discovery that rifabutin acts synergistically with antibiotics such as colistin and cefiderocol to inhibit the growth of Acinetobacter baumannii cells. Thus, in treating Acinetobacter baumannii infections, rifabutin is used in combination with colistin or cefiderocol more effectively than either of these antibiotics alone. Additionally, and unexpectedly, the combination of rifabutin and colistin is effective even against Acinetobacter baumannii strains that are resistant to both of these antibiotics when administered alone but become susceptible to treatment when administered in combination.

[0029] Combination Therapies

[0030] The combination therapy of the present invention comprises two antibiotics that act synergistically to inhibit the growth of Acinetobacter baumannii cells. The synergistic effect between antibiotics such as rifabutin and colistin can be determined by any suitable method. One method comprises determining the minimum inhibitory concentration (MIC) of each antibiotic alone and in combination and calculating the fractional inhibitory concentration index (FICI) as follows:

[0031]

[0032] Based on the following criteria, the pair of antibiotics is characterized as acting synergistically or not based on the FICI: synergy (FICI ≤ 0.5); no interaction (FICI > 0.50 and ≤ 4); antagonism (FICI > 4). The determination of the synergistic effect of antibiotics based on the FICI is described, for example, in Jenkins, S.G. and Schuetz, A.N. Current Concepts in Laboratory Testing to Guide Antimicrobial Therapy. Mayo Clin. Proc. 87, 290 - 308 (2012), the content of which is incorporated herein by reference.

[0033] In the combination therapy of the present invention, one of these antibiotics is rifabutin. These combination therapies comprise a second antibiotic that acts synergistically with rifabutin. The second antibiotic can be a polymyxin such as colistin or a cephalosporin such as cefiderocol. Colistin can be provided in the form of colistimethate sodium or colistin sulfate. These combination therapies can comprise additional antibiotics. For example, these combination therapies can comprise 3, 4, 5 or more different antibiotics. Each antibiotic can independently be an aminocyclitol, aminoglycoside, β - lactam, β - lactamase inhibitor, carbapenem, cephalosporin, polymyxin, quinolone, rifamycin, sulfonamide, minocycline, eravacycline, sulbactam or tetracycline. Each antibiotic can independently be amikacin, trimethoprim - sulfamethoxazole, cefepime, cefiderocol, ceftazidime, chloramphenicol, ciprofloxacin, colistin, doripenem, gentamicin, imipenem, levofloxacin, meropenem, penicillin, piperacillin, polymyxin B, rifabutin, rifampicin, tazobactam or tigecycline.

[0034] Rifabutin is a dark red - purple powder with the molecular formula C 46 H 62 NO 11 , a molecular weight of 847.02 and having the following structure:

[0035]

[0036] Rifabutin has broad-spectrum antimicrobial activity. Rifabutin has much higher activity against MAC, Mycobacterium tuberculosis (M. tuberculosis), and Mycobacterium leprae (M. leprae) compared to rifampicin. Rifabutin also has activity against most atypical mycobacteria, including Mycobacterium kansasii; however, Mycobacterium chelonae is relatively resistant. Rifabutin has activity against staphylococci, group A streptococci, Neisseria gonorrhoeae (N. gonorrhoeae), Neisseria meningitidis (N. meningitidis), Haemophilus influenzae (H. influenzae), Haemophilus ducreyi (H. ducreyi), Campylobacter jejuni (C. jejuni), Helicobacter pylori (H. pylori), Chlamydia trachomatis (C. trachomatis), Toxoplasma gondii (T. gondii), and Acinetobacter baumannii.

[0037] Each antibiotic can be administered by any suitable route of administration. For example, and without limitation, each antibiotic can independently be administered intravenously, orally, parenterally, subcutaneously, by inhalation, by injection, and / or by infusion.

[0038] One or more of the antibiotics in these combination therapies can be administered relative to the same dosing regimen. One or more of the antibiotics can be administered according to different dosing regimens. A dosing regimen can include a dose, a schedule, or both administration and. The dose can be described by the absolute amount of the drug (e.g., mg) or by the relative amount of the drug relative to the subject (e.g., mg / kg). The schedule of administration can be described by the interval between doses. For example, and without limitation, the interval between doses can be about one hour, about 2 hours, about 3 hours, about 4 hours, about 6 hours, about 8 hours, about 12 hours, about 24 hours, about 36 hours, about 48 hours, about 3 days, about 4 days, about 5 days, about 6 days, about 7 days, or longer.

[0039] Formulations

[0040] One or more of these antibiotics can be provided in the form of a single formulation. One or more of the antibiotics can be provided in separate formulations. Each formulation can be prepared for delivery by a specific route of administration, such as intravenously, orally, parenterally, subcutaneously, by inhalation, by injection, and / or by infusion.

[0041] These antibiotics can be provided in the form of pharmaceutically acceptable salts, such as non-toxic acid addition salts, which are salts of amino groups formed using inorganic acids such as, but not limited to, hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid, and perchloric acid, or using organic acids such as, but not limited to, acetic acid, maleic acid, tartaric acid, citric acid, succinic acid, methanesulfonic acid, glucuronic acid, malic acid, gluconic acid, lactic acid, aspartic acid, or malonic acid.

[0042] The formulation can be administered by injection, infusion, implantation (intravenous, intramuscular, subcutaneous, etc.), or by inhalation of a dosage form, formulation, or by a suitable delivery device or implant containing conventional non-toxic pharmaceutically acceptable carriers, solvents, diluents, and adjuvants. The formulations and preparations of such compositions are well known to those skilled in the art of pharmaceutical formulations.

[0043] Formulations for parenteral use can be provided in unit dosage forms (e.g., in single-dose ampoules and vials), in vials containing several doses and to which a suitable preservative (see below) can be added, in pre-filled syringes, or in pre-filled IV bags.

[0044] The pharmaceutical compositions described herein can be in a form suitable for sterile injection.

[0045] The formulation can comprise a solution containing rifabutin. The rifabutin solution and methods for preparing the rifabutin solution are described in co-owned, co-pending U.S. Application No. 62 / 902,019, the content of which is incorporated herein by reference.

[0046] Depending on the needs of the patient and the clinical condition, administration of the composition by IV may be preferred over oral administration because IV administration allows for the rapid introduction of the antibiotic into the systemic circulation, provides complete bioavailability, allows for better control of the pharmacokinetic parameters driving pharmacological efficacy, and avoids issues of stability and absorption in the gastrointestinal tract.

[0047] A typical dose of rifabutin is a dose that can achieve a plasma or local level of rifabutin C max > 2 mg / L but < 50 mg / L and an AUC of 10 mg*h / L < 200 mg*h / L.

[0048] The formulation can be formulated for parenteral administration, such as by injection or infusion. The injection or infusion can be subcutaneous or intravenous.

[0049] Treatment of Acinetobacter baumannii infections

[0050] The combination therapy of the present invention can be used to treat Acinetobacter baumannii infections in a subject. The subject can be a human. The subject can be a pediatric, neonatal, infant, toddler, child, adolescent, preteen, young adult, adult, or elderly subject. The subject can be in emergency care, intensive care, neonatal intensive care, pediatric intensive care, coronary care, cardiothoracic care, surgical intensive care, medical intensive care, long-term intensive care, operating room, ambulance, field hospital, out-of-hospital scene environment.

[0051] The subject may have an Acinetobacter baumannii infection that is resistant to antibiotics. For example and without limitation, the Acinetobacter baumannii infection can be resistant to one or more of the following: aminocyclitol, aminoglycoside, β-lactam, β-lactamase inhibitor, carbapenem, cephalosporin, polymyxin, quinolone, rifamycin, sulfonamide, tetracycline, amikacin, trimethoprim-sulfamethoxazole, cefepime, cefiderocol, ceftazidime, chloramphenicol, ciprofloxacin, colistin, doripenem, gentamicin, imipenem, levofloxacin, meropenem, penicillin, piperacillin, rifabutin, rifampicin, tazobactam, and tigecycline. The Acinetobacter baumannii infection can be resistant to rifabutin, colistin, or both. The Acinetobacter baumannii infection can be resistant to rifabutin, cefiderocol, or both.

[0052] The antibiotics in the combination therapy can be administered simultaneously or sequentially. Sequential or alternating administration can include exclusively providing each antibiotic for a certain period of time. Sequential administration can include an overlapping period during which both an IV formulation containing rifabutin and a formulation containing another therapeutic agent are provided to the subject. The exclusive period and the overlapping period can independently be 1 hour, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 1 week, or 2 weeks.

[0053] Example

[0054] Overview

[0055] The goal of this study was to identify antibiotics that synergize with rifabutin against the standard of care (SoC) for Acinetobacter baumannii. Synergy was evaluated by checkerboard minimum inhibitory concentration (MIC) against multiple clinical isolates of Acinetobacter baumannii. First, the synergy of rifabutin with colistin and cefiderocol was tested and identified against the LAC-4 strain. When tested against a panel of strains, rifabutin / cefiderocol synergy was observed on 100% of the strains, with a ≥4-fold decrease in the MIC of cefiderocol. Rifabutin / colistin synergy was observed in 100% of the strains, and this synergy was strong and independent of the initial resistance levels to rifabutin or colistin. When combined with colistin, rifabutin was more active than rifampicin, because, as described in the literature, the synergy with rifampicin depends on the initial resistance level to rifampicin and the presence of rpoB mutations. Unexpectedly, the rifabutin / colistin combination was active against strains resistant to rifabutin (including isolates with rpoB mutations) and / or strains resistant to colistin, indicating that this combination overcomes both resistances.

[0056] In summary, rifabutin has the ability to enhance the antibacterial activity of cefiderocol and colistin against Acinetobacter baumannii strains.

[0057] Antimicrobial agents

[0058] BV-015-3219-001-02 (rifabutin (batch number 17008MR89D)) was manufactured by Olon S.p.A. and a 10 g / L stock solution was prepared in DMSO. Stock solutions of rifampicin (Sigma R3501) and cefiderocol (Synnovator SYNNAAX397783) were prepared at 10 mg / mL in DMSO. Stock solutions of colistin sulfate (Sigma C4461), meropenem (Sigma M2578), cefotaxime (Acros 45495), tobramycin (Sigma T1783), eravacycline (MedChemExpress HY-16980A), minocycline (Sigma M9511), and SPR741 (Spero Therapeutics, FullReg P0271508-1) were prepared at 10 mg / mL in water. Finally, a stock solution of ciprofloxacin (Sigma 17850) was prepared at 10 mg / ml in 0.1 N NaOH. The stock solutions were stored at -20 °C until use.

[0059] Bacterial strains

[0060] The Acinetobacter baumannii clinical isolates used in this study were from the BioVersys strain collection. These strains were stored at -80 °C as 20% (v / v) glycerol stock cultures.

[0061] Antimicrobial susceptibility and synergy testing

[0062] The synergy of rifabutin with SoC antibiotics was tested using the broth microdilution checkerboard method. The checkerboard method was performed according to the CLSI parameters used for microbroth dilution MIC5. CA-MHB, iron-depleted CA-MHB (ID-CA-MHB), or RPMI supplemented with 10% FCS medium was used as specified, and ID-CA-MHB was prepared according to CLSI guidelines. Rifabutin was serially diluted along the abscissa and the combination antibiotics were diluted along the ordinate. This setup allowed the combination of rifabutin and another antibiotic to be tested at increasing concentrations to provide a final classification of the combination based on the fractional inhibitory concentration (FIC) index (FICI) as follows: synergy (FICI ≤ 0.5); no interaction (FICI > 0.50 and ≤ 4); antagonism (FICI > 4). The FICI was calculated as follows:

[0063]

[0064] Thus, synergy was defined when the MIC of the antibiotics tested in combination was reduced by at least 4-fold compared to the MIC of the antibiotics tested alone.

[0065] Example 1

[0066] Rifabutin synergistically targeted Acinetobacter baumannii strain LAC-4 with colistin and cefiderocol. The synergy of rifabutin with SoC antibiotics was tested using a checkerboard assay against Acinetobacter baumannii strain LAC-4. The checkerboard method was performed for cefiderocol in CA-MHB or ID-CA-MHB. The results are shown in Table 1.

[0067] Table 1.

[0068] Second Antibiotic FICI Explanation Colistin 0.254 Synergy Meropenem 1 Irrelevant Cefotaxime 1.5 Irrelevant Ciprofloxacin 1.5 Irrelevant Tobramycin 1.5 Irrelevant Cefiderocol* 0.5 Synergy Eravacycline 1 Irrelevant Minocycline 1 Irrelevant

[0069] *Cefiderocol was tested in ID-CA-MHB

[0070] Among the 8 SoC antibiotics tested, only colistin and cefiderocol showed synergy with rifabutin, while the other 6 were non-interactive.

[0071] Figure 1An image of a 96-well plate checkerboard of Acinetobacter baumannii cells cultured with rifabutin and colistin at various concentrations. Wells used to determine the MIC of the individual antibiotics are circled in green, wells of the combined MIC are circled in blue, and wells used to calculate the FICI are circled in red.

[0072] Figure 2 An image of a 96-well plate checkerboard of Acinetobacter baumannii cells cultured with rifabutin and cefiderocol at various concentrations. Wells used to determine the MIC of the individual antibiotics are circled in green, wells of the combined MIC are circled in blue, and wells used to calculate the FICI are circled in red.

[0073] Example 2

[0074] Rifabutin decreased the MIC of cefiderocol against the tested Acinetobacter baumannii strains by ≥4-fold.

[0075] To further investigate the synergistic effect of rifabutin and cefiderocol against Acinetobacter baumannii, the checkerboard method was performed on a panel of 16 MDR clinical isolates of Acinetobacter baumannii, which included 5 isolates with elevated MICs (≥32 mg / L) of rifabutin (and rifampin) due to mutations in the rpoB gene. To more precisely describe the level of synergy, the MICs of rifabutin and cefiderocol alone and in combination, as well as the fold changes associated with these MICs, are presented in Table 2 for each strain. As expected, cefiderocol in combination with rifabutin had little or no effect on the activity of rifabutin against isolates with mutations in the rpoB gene. Unexpectedly, rifabutin decreased the MIC of cefiderocol against all tested Acinetobacter baumannii strains by at least 4-fold.

[0076] Table 2.

[0077]

[0078]

[0079] **Synergy was determined in ID-CA-MHB.

[0080] In summary, the data suggest that combining rifabutin with cefiderocol can improve the treatment outcome of Acinetobacter baumannii infections.

[0081] Example 3

[0082] Rifabutin exhibited strong synergy with colistin on 100% of the tested Acinetobacter baumannii strains.

[0083] The same procedure was performed for the synergistic effect of rifabutin and colistin against Acinetobacter baumannii. The checkerboard method was performed on 16 MDR clinical isolates of Acinetobacter baumannii, which included 5 isolates with elevated MICs (>32 mg / L) of rifabutin (and rifampicin) with mutations in the rpoB gene and 5 colistin-resistant strains (MIC > 4 mg / L). Synergism was tested in CA-MHB medium, which is the approved medium for testing colistin MIC. The results are shown in Table 3. Unexpectedly, colistin in combination with rifabutin had a significant effect on the activity of rifabutin against isolates with mutations in the rpoB gene. In these cases, the change in rifabutin MIC in combination with colistin was >32-fold.

[0084] Table 3.

[0085]

[0086]

[0087] Synergism between rifabutin and colistin was observed in 100% of the strains when tested in CA-MHB. Significantly, as exemplified in CA-MHB, the synergism was independent of the original resistance levels of rifabutin or colistin, where the colistin MIC was reduced by at least 16-fold in colistin-resistant strains, rendering all but one of the strains colistin-susceptible. These results suggest that combining rifabutin with colistin has the potential to overcome rifabutin and colistin resistance in clinical isolates of Acinetobacter baumannii.

[0088] These observations could suggest that colistin acts as a cell permeabilizer to synergize with rifabutin. To evaluate whether colistin only acts as a permeabilizer, the synergistic effect of rifabutin in combination with the colistin derivative SPR741, which retains permeabilizing activity but has lost antibacterial activity, was tested. The results are shown in Table 4.

[0089] Table 4.

[0090]

[0091] Synergism between rifabutin and SPR741 was observed in 88% of the strains. However, for most strains, the synergism was less pronounced compared to the synergism where the MICs in combination with colistin and rifabutin remained in the range of 0.125 - 2 mg / L. In addition, the concentration of SPR741 required to achieve synergism with rifabutin was at least 8-fold higher than the colistin concentration. These results suggest that the intrinsic antibacterial activity of colistin is required for strong synergism with rifabutin.

[0092] Example 4

[0093] Comparison of the Synergistic Effects of Rifampicin and Colistin against Acinetobacter baumannii Isolates

[0094] As a comparative example, the synergistic effects of rifampicin and colistin were determined for a set of Acinetobacter baumannii strains. The results are shown in Table 5.

[0095] Table 5.

[0096]

[0097] Synergistic effects were observed on 88% of the strains. Regarding rifabutin, as demonstrated by the synergistic effects against most colistin-resistant strains, the synergistic effects were independent of the colistin resistance level. However, in contrast to rifabutin and as expected from the literature, the synergistic effects of the rifampicin / colistin pair were weak for isolates with mutations in the rpoB gene. For these isolates, the rifampicin combination MIC remained high (≥32 mg / L).

[0098] Overall, it was demonstrated that colistin could enhance the activity of rifabutin against Acinetobacter baumannii strains with elevated MICs to rifabutin, colistin, or both, and vice versa. However, unexpectedly, in contrast to rifampicin, rifabutin in combination with colistin was active against isolates with mutations in the rpoB gene that were originally resistant to these antibiotics.

[0099] Incorporated by Reference

[0100] Throughout this disclosure, other documents have been referenced and cited, such as patents, patent applications, patent publications, magazines, books, papers, web page content. For all purposes, all such documents are hereby incorporated by reference in their entirety into this text.

[0101] Equivalents

[0102] Based on the entire content of this document, including the references to scientific and patent literature cited herein, various modifications of the present invention and many additional embodiments thereof will become apparent to those skilled in the art. The subject matter herein contains important information, examples, and guidance that can be suitable for practicing the present invention in various embodiments of the present invention and their equivalents.

Claims

1. Use of a combination of rifabutin and a second antibiotic, polymyxin or cefiderocol, in the manufacture of a medicament for treating Acinetobacter baumannii infections in a subject.

2. The use according to claim 1, wherein the subject is infected with an Acinetobacter baumannii strain resistant to rifabutin.

3. The use according to claim 1, wherein the subject is infected with an Acinetobacter baumannii strain resistant to the second antibiotic.

4. The use according to claim 1, wherein the rifabutin is administered intravenously.

5. The use according to claim 1, wherein the rifabutin is administered by inhalation.

6. The use according to claim 1, wherein the rifabutin is administered orally.

7. The use according to claim 1, wherein the rifabutin and the second antibiotic are provided in a single formulation.

8. The use according to claim 1, wherein the rifabutin and the second antibiotic are provided separately.

9. The use according to claim 1, wherein the Acinetobacter baumannii comprises an rpoB mutation.

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