Compositions of clofazimine, combinations containing same, processes for preparing same, uses and methods containing same

Abstract

To provide a pharmaceutical composition containing clofazimine, a combination medicine containing the composition which is in the form of aerosol for inhalation, a system to be used in imparting antibacterial activity in treating or preventing pulmonary infection, and a method of treating or preventing pulmonary infection.SOLUTION: A pharmaceutical composition contains: a) clofazimine of a therapeutically effective dose, or a pharmaceutically acceptable derivative or salt thereof; b) a nonionic surfactant having a hydrophilic-lipophilic balance value exceeding 10; c) an aqueous liquid carrier selected from at least one of water, isotonic physiological saline, buffered physiological saline, and electrolytic aqueous solution; and d) a mucolytic agent and / or biofilm destroying agent, where the clofazimine or pharmaceutically acceptable derivative or salt thereof is provided in the form of particles in suspension, and the central diameter of the particles of the clofazimine or pharmaceutically acceptable derivative or salt thereof is less than 5 μm, and D90 is less than 6 μm.SELECTED DRAWING: None

Description

Technical Field

[0001] This application claims priority to U.S. Provisional Application No. 62 / 722,048, filed on Aug. 23, 2018, and this application also claims priority to U.S. Provisional Application No. 62 / 796,322, filed on Jan. 25, 2019, the contents of both of which are hereby incorporated herein by reference as part of this specification.

[0002] The present invention relates to an inhalation pharmaceutical composition comprising a therapeutically effective dose of clofazimine, wherein the clofazimine is provided in the form of a suspension; a process for preparing the same; and uses and treatment methods comprising the same. Further, the present invention provides a combination medicament comprising clofazimine in the form of an aerosol for pulmonary inhalation.

[0003]

Background Art

[0004] Clofazimine is a highly hydrophobic (LogP = 7.66) antibiotic, luminophenazine, which has antimycobacterial and anti-inflammatory activities, and was first described in 1957. The structural formula is shown below.

Chemical Formula

[0005] The exact mechanism by which clofazimine exerts its antibacterial action is not known. However, , which preferentially binds to mycobacterial DNA, thereby inhibiting DNA replication and cell proliferation It is known that it inhibits the oxidative stress response. Other proposed mechanisms of action include membrane damage / anxiety. stabilization, production of membrane-destabilizing lysophospholipids, potassium transport and / or intracellular redox In vitro, clofazimine has been shown to inhibit the Mycobacterium tuberculosis bacillus, including multidrug-resistant strains. (Mycobacterium tuberculosis While it has shown excellent activity against mycobacterial tuberculosis (MTB), until recently, it was not used to treat pulmonary tuberculosis. The general consensus was that it was not effective (e.g., Cholo M et al., J Antimicrob Chemother,2012 Feb,67(2):290 -8).

[0006] Clofazimine is an antibacterial agent that inhibits the leprosy bacteria (Mycobacterium leprae) It is used by the World Health Organization to treat leprosy, a disease caused by Bacillus subtilis leprae. The three types of dietary supplements recommended by the World Health Organization It is one of the main drugs for the treatment of mycobacteria caused by nontuberculous mycobacteria (NTM) in recent years. It is increasingly being used to treat other mycobacterial infections, such as drug-resistant tuberculosis.

[0007] Clofazimine is practically insoluble in water and exhibits high membrane permeability, making it a promising biopharmaceutical agent. Biopharmaceutics Classification System It is classified as a Class II drug in the Brain Stem Cell System (BCS).

[0008] Overcoming problems associated with poor oral absorption and poor bioavailability of drugs In order to optimize the oral dosage form, zation, recrystallization using supercritical fluid re-crystallization, spray freeze drying in liquid eeze drying into liquid), solid dispersions, and solid solutions. strategies have been used.

[0009] Clofazimine is classified as a BCS Class II drug and is therefore usually administered orally. Formulating solid dispersions to improve ioavailability is an ideal option. (e.g., Bhusnure et al. IJRPC 2014, 4 (4), 906-918).

[0010] Accordingly, clofazimine, which has lipophilic properties, was developed to improve oral absorption. It is usually administered as a microcrystalline suspension in an oil-wax base. The rate of absorption by the human body after oral administration varies considerably (45-62%). Adverse effects of Fazimin are dose-related and primarily affect the skin, eyes, and gastrointestinal tract. The QT interval prolongation side effect is accompanied by reddish-brown discoloration of the skin and conjunctiva, which slowly resolves with drug discontinuation. This is due to long-term retention in the whole body.

[0011] The genus Mycobacterium is a member of the phylum Actinobacteria. It is a genus of the Mycobacteria family. Mycobacteria are a distinct family of bacteria (Mycobacteria ae). They are rod-shaped and have a waxy outer membrane. do.

[0012] As such, mycobacteria can be divided into three groups: Mycobacterium tuberculosis (Mycobacterium tuberculosis) tuberculosis group - the causative agent of tuberculosis Mycobacterium leprae ))-Causing agent of leprosy ·Nontuberculous mycobacteria (NT M)-Mycobacterium tuberculosis (M. tuberculosis) and Mycobacterium leprae ( Includes all mycobacteria except M. leprae. Mycobacterium abscessus complex (Mycobacterium abscessus complex) Mycobacterium avium complex (MABSC) Mycobacterium avium complex (MAC), etc.

[0013] Tuberculosis (TB) is caused by Mycobacterium tuberculosis complex bacteria. TB is an infectious disease caused by the bacteria TB complex. It is one of the oldest documented infectious human pathogens and continues to cause mortality worldwide. The number of new TB infections in 2015 was 10 400,000 people are estimated to be infected with TB, and 1.4 million people die from active TB (e.g., World Health Organization (WHO) Global Tuberculosis Report In addition to the high morbidity and mortality rates, There is growing concern about the emergence of MDR-TB, with 580,000 new cases reported in 2015. It has been shown that 100% of patients were infected with drug-resistant TB. Co-morbidities such as TB, HIV, and TBV complicate treatment and were responsible for 1.2 million TB cases in 2015. was.

[0014] To treat multidrug-resistant (MDR) infections, WHO recommends the use of second-line anti-TB drugs. The authors have recommended a 9- to 12-month treatment regimen that includes Such regimens as the Bangladesh regimen In the study, gatifloxacin, ethambutol, pyrazinamide, and chloramphenicol were used for MDR-TB. Combination treatment with lofazimine was administered, and 87.9% of patients achieved recurrence-free cure (e.g. Sotgiu, G, et al., “Applicability of the sh orter'Bangladesh regimen'in high multidr. ug-resistant tuberculosis settings”,Inte National Journal of Infectious Diseases (2017) 56 190-193).

[0015] Other studies investigating shortening the duration of TB treatment have shown that clofazimine is administered orally for 2 weeks. It has been shown that there is no clinical benefit after administration (e.g., Diacon, AH, e t al.,“Bactericidal Activity of Pyrazina mide and Clofazimine Alone and in Combin ations with Pretomanid and Bedaquiline”, American Journal of Respiratory and Criticism ical Care Medicine(2015), 191(8), 943-953 It is theorized that the drug binds with high affinity to circulating serum proteins. This lack of efficacy may be due to poor bioavailability of the drug, Clofazimine is thought to be a potent inhibitor of MDR-TB and extremely drug-resistant TB (XDR-TB). Although empirical evidence has demonstrated that short-term treatment with systemic injections is effective in treating Its biological activity appears to be limited by its poor bioavailability after administration (e.g., Swanson, RV, et al., “Pharmacokinetics an d Pharmacodynamics of Clofazimine in a M ouse Model of Tuberculosis”,Antimicrobia l Agents and Chemotherapy(2015),59(6),30 (See 42-3051.)

[0016] Treating lung infections with inhaled antibiotics results in higher drug concentrations in the lungs and , and reduced side effects compared to systemic delivery (e.g., Touw, DJ, et al., “Inhalation of antibiotics in cystic fib European Respiratory Journal (1995) ), 8, 1594-1604), resulting in increased biological activity and efficacy. It is known (e.g., Hickey, AJ, "Inhaled drug trials eating for tuberculosis:Past progress a nd future prospects”,Journal of Controll ed Release, (2016), 240, 127-134). TB infection model When clofazimine was administered as an aerosol, it was found to be more effective than oral clofazimine. In comparison, the rate of bacilli elimination was 10% after just 28 days of treatment. Significant improvements in the efficacy of GABA have been demonstrated in in vivo mouse models (e.g., Verma et al., ,RK,et al.,“Inhaled microparticles con taining clofazimine are efficacious in t reatment of experimental Tuberculosis in Mice”, Antimicrobial Agents and Chemothe (see rapy (2013), 57(2), 1050-1052). The improved efficacy was due to the direct delivery of clofazimine to the lungs, resulting in the development of pulmonary tuberculous granulomas. This is likely due to higher clofazimine concentrations in intracellular macrophages.

[0017] Therefore, clofazimine was administered by aerosol to patients with MDR-TB or XDR-TB infection. Administration of the drug in a more concentrated form should improve patient outcomes and may be an effective treatment option for current treatment regimens. The period may be shortened.

[0018] Formerly called atypical or ubiquitous mycobacteria The nontuberculous mycobacteria (NTM) group includes more than 150 species. They are widespread and diverse in the soil, ground and drinking water, as well as in low temperature They can also be found in foods such as pasteurized milk and cheese. NTM is generally considered to be of low pathogenicity. However, these are considered to be harmful to humans, especially those with weakened immune systems or those who already have lung disease. NTM can cause serious illness in affected humans. They are classified according to their growth rate, and are divided into slow growing (SGM) and rapid growing (RGM) mycobacteria. They are divided into terriers.

[0019] Slow-growing Mycobacterium avium complex (Mycobacterium Mycobacterium avium complex (MAC) includes Mycobacterium avium cobacterium avium, Mycobacterium chimaera (Mycoba Mycobacterium chimaera, and Mycobacterium intracellulare ( These include the Mycobacterium intracellulare species. is the most important and most common pathogenic NTM. Mycobacterium kansasii ( Mycobacterium kansasii, Mycobacterium marmoense (Mycobaceterium malmoense), Mycobacterium xenopi (Mycobacterium xenopi), Mycobacterium simiae (Myc obacterium.simiae), Mycobacterium abscessus (Myco bacterium abscessus, Mycobacterium gordonii (Myco bacterium gordonae, Mycobacterium fortuitum (M Mycobacterium fortuitum, and Mycobacterium chelonae Most of these, like Mycobacterium chelonae, are pulmonary It causes infection. Mycobacterium marinum arinum is a skin tumor that resembles an aquarium granuloma. and soft tissue infections.

[0020] In particular, RGM causes severe, life-threatening chronic lung disease and is disseminated, often It is responsible for fatal infections. The infections are usually caused by catheter-related contaminated materials (co non-sterile materials) and invasive procedures, non-sterile surgical procedures or injections Contact with shower heads and jacuzzis also poses a risk of infection. NTM is usually associated with chronic obstructive pulmonary disease (COPD) and cystic fibrosis. It causes opportunistic infections in patients with chronic lung diseases such as CF and other immune-compromised patients.

[0021] In recent years, rapid growing type (RGM) Mycobacterium abscessus Mycobacterium abscessus group (subspecies Mycobacterium abscessus Mycobacterium abscessus subsp. abscessus bscessus) (Maabscessus (M·a·abscessus)), Myco Bacterium abcessus boleti (Mycobacterium abcessus bolletii.), and Mycobacterium abscessus massiliense (My Mycobacterium abscessus massiliense Bacterium abscessus complex (Mycobacterium abscessus complex) The importance of the MABSC (Mammalian essus complex) as a human pathogen has become clear. It has been associated with a much higher mortality rate than any other RGM.

[0022] CF patients are susceptible to Mycobacterium abscessus. cessus), lung destruction spreads and is often untreatable. This is especially problematic because the rate of success is high, at 60-66%. (See, for example, Obregon-He nao A et al, Antimicrobial Agents and Che motherapy, November 2015, Vol 59, No 11, p. 6904-6912;Qvist, T., Pressler, T., Hoiby, Na nd Katzenstein, T.L., “Shifting paradigms o f nontuberculous mycobacteria in cystic fibrosis”,Respiratory Research(2014),15( 1):pp.41-47).

[0023] Human infections with NTM are increasingly being linked to outbreaks of human acquired immunodeficiency syndrome. Mycobacterium avium complex (Mycobacterium a Mycobacteria belonging to the Mycobacterium virum complex (MAC) are responsible for the development of human immunodeficiency viruses. It has been identified as a major cause of opportunistic infections in patients infected with HIV.

[0024] Some NTM species are known to form biofilms. Microcolonies are small colonies of bacteria contained within extracellular matrix and are involved in the human immune system. In recent years, some NTM species have been treated with disinfectants and antibiotics. It has been shown that it forms biofilms that increase resistance to antimicrobial agents. The assembly of bacteria can be mediated by reversible adhesion, irreversible adhesion, bacterial aggregation to form biofilms, organization, and This process proceeds through several steps, including endothelial cell death, signaling, and ultimately dispersal. During migration, bacteria attach to matrix, which contains extracellular polymeric substances (EPS) such as polysaccharides, lipids, and nucleic acids. They then assemble the complex into three-dimensional structures (e.g., Sousa S. et al. ,International Journal of Mycobacteriolo gy 4(2015), 36-43). Specifically, mycobacteria produce exopolysaccharides Mycobacterial EPS is different from other biofilms because it does not produce For example, Zambrano MM, Kolter R. Mycobacterial b iofilms:a greasy way to hold it together Mycobacterial biofilms vary among species, but Cholic acid, glycopeptide lipids, mycolyl-diacylglycerol, lipooligosaccharide, lipopeptide tides, and extracellular DNA (Rose SJ, Babrak LM, Bermu dez LE(2015)Mycobacterium avium Possesse s Extracellular DNA that Contributes to Biofilm Formation,Structural Integrity,a nd Tolerance to Antibiotics. Abstract from PLoS ONE (Thesis and original research). Aggregations within biofilms are known to increase resistance to antibacterial agents. (For example, Faria S. et al., Journal of Pathology ogens, Vol 2015, Article ID 809014).

[0025] Aerosolized liposomal ampicillin as a novel approach to treat NTM pulmonary infections Delivery of amikacin / inhalation of amikacin solution nebulized by jet nebulizer (Rose et al. S.et al,2014,PLoS ONE,Volume 9,Issue 9,e 108703 and Olivier K. et al., Ann Am Thorac So c Vol 11, No 1, pp.30-35), as well as the development of a dried anti-TB drug for pulmonary delivery. Inhalation of powder particles (Cholo M et al., J Antimicrob Che mother.2012 Feb;67(2):290-8 and Fourie B and Nettey O.,2015 Inhalation Magazine , Verma 2013 Antimicrob Agents Chemother ) has been proposed.

[0026] In the treatment of NTM lung disease, parenteral aminoglycosides, tigecycline, and linecycline are Other promising oral antibacterials, such as Zolid, delamanid, and bedaquiline, as well as antimicrobials for selected indications In this case, initial treatment was surgical intervention followed by multidrug therapy with inhaled amikacin. Combination therapy regimens have shown promising results (Lu Ryu et al. al.,Tuberc Respir Dis 2016;79:74-84). However, the incidence and prevalence of NTM infections, especially NTM lung disease, is increasing and treatment is becoming more difficult. Given the limited treatment options, currently used antibiotics such as clofazimine are There is a need to develop new dosage forms / pharmaceutical formulations that improve bioavailability. Inhalation has been shown to increase efficacy and reduce side effects compared to oral and parenteral treatments. There is a possibility that.

[0027] The combination of clofazimine and amikacin inhibits the proliferation of Mycobacterium abscessus (M ycobacterium abscessus) and Mycobacterium avium ( Mycobacterium avium) It has been shown in vitro (e.g., van Ingen, J., et al., “In Vitro Synergy between Clofazimine and A mikacin in Treatment of Nontuberculous M ycobacterial disease”,Antimicrobial Agen TS and Chemotherapy 56(12),6324-6327(201 2) In addition, Mycobacterium tuberculosis (Mycobacterium tuberculosis) Clofazimine and bedaquiline combination for erium tuberculosis Synergistic effects have also been shown by combining the two (e.g., Cokol, M. et al., "E fficient measurement and factorization o f high-order drug interactions in Mycoba cterium tuberculosis”,Sciences Advances 2017:3:e170881,11 October 2017). Clofazimine / bedakimine was also used against the mycobacterium Mycobacterium abscessus. A synergistic effect has been shown when phosphorus is used in combination (Ruth, MM et al., "AB edaquiline / Clofazimine Combination Regim en Might Add Activity to the Treatment o f Clinically Relevant Non-Tuberculous My cobacteria”, Journal of Antimicrobial Che motherapy(2019),doi.org / 10.1093 / jac / dky5 26).

[0028] Pathogenic fungi are emerging as a major cause of human mortality. Current estimates are In the current study, the number of deaths caused by deep mycoses is comparable to that caused by more well-known infectious diseases such as tuberculosis. It has been suggested that Candida albicans (Candida albicans ns), Cryptococcus neoformans mans, and Aspergillis fumigatus tus) are the most common pathogenic fungi in humans. Each of these is responsible for hundreds of thousands of infections each year, but diagnostic methods are scarce and treatment options are limited. The mortality rate is unacceptably high because of the combination of clofazimine and It has been shown to be effective against multiple species of fungi (e.g., Robbins, N., et al., “An Antifungal Combination Matrix Identifies a Rich Pool of Adjuvant Molec ules that Enhance Drug Activity against Diverse Fungal Pathogens”,Cell Reports 1 3, 1481-1492, November 17, 2015). The fungus is a cystic fibrosis. In fibrosis, they act as commensals, colonizers, and / or pathogens (e.g., C. hotirmall,SHand McElvaney,NG,“Fungi in the cystic fibrosis lung: Bystanders o r pathogens?”,The International Journal of Biochemistry & Cell Biology 52(2014), (see 161-173). Summary of the Invention [Problem to be solved by the invention]

[0029] Clofazimine is poorly water soluble, leading to poor oral bioavailability and resistance to the drug. The lower respiratory tract has high microbial resistance. A nebulizer is used to deposit aerosol particles onto the The drug is solubilized and formulated in a liquid aqueous carrier for aerosolization by aerosolization, etc. Special techniques for the synthesis and stabilization of cellulose are also required. [Means for solving the problem]

[0030] According to one embodiment of the present invention: (a) a therapeutically effective dose of clofazimine or a pharma- ceutical acceptable derivative or salt thereof; (b) a nonionic surfactant having a hydrophilic-lipophilic balance value greater than 10; (c) an aqueous liquid selected from water, isotonic saline, buffered saline, and aqueous electrolyte solutions; With carrier; A pharmaceutical composition comprising: The clofazimine or a pharma- ceutical acceptable derivative or salt thereof is in the form of particles in suspension. Provided in and The median diameter of the particles of clofazimine or its pharma- ceutical acceptable derivatives or salts is less than 5 μm. and a D90 of less than 6 μm.

[0031] According to another embodiment of the present invention, clofazimine or a pharma- ceutical acceptable derivative thereof is Alternatively, the salt particles have an average diameter of less than 2 μm and a D90 of less than 3 μm.

[0032] In another embodiment of the present invention: (a) a therapeutically effective dose of clofazimine; (b) a nonionic surfactant having a hydrophilic-lipophilic balance value greater than 10; (c) an aqueous liquid selected from water, isotonic saline, buffered saline, and aqueous electrolyte solutions; With carrier; A pharmaceutical composition comprising: The clofazimine is provided in the form of particles in a suspension, and The clofazimine particles have a median diameter of less than 5 μm and a D90 of less than 6 μm. Pharmaceutical compositions are provided.

[0033] In another embodiment of the invention, the particles of clofazimine have a median diameter of less than 2 μm; And D90 is less than 3 μm.

[0034] The compositions of the present invention can be aerosolized with a suitable nebulizer to produce aerosolized creatine. Lofazimine is effective in preventing and treating the lower respiratory system (i.e., bronchi, bronchioles, and middle and peripheral lungs). delivery to the alveoli (the pulmonary alveoli in the pulmonary septum and lower peripheral lung) This significantly improves the efficacy of the treatment.

[0035] More preferably, the inhaler is adapted to be axially aligned for uniform deposition in the lower respiratory system. The aerosol should be further adapted for localized delivery to the lungs with a suitable particle size distribution. be.

[0036] Thus, the present invention provides an aerosol having a particle size that facilitates delivery to the alveoli and bronchioles. The optimal aerodynamic particle size for targeting the alveoli and bronchioles is between 1 and 5 μm. Larger particles are absorbed by the upper lung, i.e. the bronchi. and is preferentially deposited in the trachea and mouth and pharynx, i.e., the oropharyngeal region. The device has a mass median aerodynamic diameter (MMAD) in the range of about 1 to about 5 μm, preferably is adapted to generate an aerosol in the range of about 1 to about 3 μm. In this case, the particle size distribution is narrow, with a geometric standard deviation (GSD) of less than about 2.5.

[0037] The present invention provides a method for administering clofazimine in the form of a suspension by aerosol to the lungs, thereby preventing hypoxia. (i.e., deep) respiratory system, thereby providing a highly Increased bioavailability of water-insoluble BCS Class II active substances, resulting in The unexpected discovery was that this significantly improved the therapeutic effect while reducing systemic side effects. It is based on.

[0038] In another embodiment, the discovery provides a method for the detection of inflammatory bowel diseases caused by mycobacteria and gram-positive bacteria. Infections caused by NTM, particularly pulmonary infections, such as CF, COPD, and immune system disorders. This will improve antibiotic treatment of opportunistic infections in patients with reduced immune function (such as HIV patients).

[0039] The present invention further relates to established methods for treating pulmonary infections caused by Gram-positive bacteria, in particular pulmonary TB and NTM infections. Not only does it eliminate the systemic side effects of conventional oral treatment regimens, but it also reduces the dose and The aim is to reduce the time and duration of treatment.

[0040] It will be understood by those skilled in the art that the present application also discloses any and all combinations of the individual features disclosed herein. Be better understood. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0041] definition The term "pharmaceutical acceptable salts" refers to salts that retain the biological effectiveness and properties of the compounds of the present invention. It refers to salts that have a biological or other undesirable effect and are not biologically or otherwise undesirable. The compounds of the present invention are those which have an amino group and / or a carboxyl group or a group similar thereto. By adding 1,000 to 1,000 sucrose, the salt of the acid and / or base can be formed. Addition salts can be formed with inorganic and organic acids. Examples of organic acids include hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, and phosphoric acid. Examples of organic acids that can be introduced include acetic acid, propionic acid, naphthoic acid, oleic acid, and the like. acid, palmitic acid, pamoic (embonic) acid, stearic acid, glycolic acid, pyruvic acid, Oxalic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, ascorbic acid acid, glucoheptonic acid, glucuronic acid, lactic acid, lactobionic acid, tartaric acid, benzoic acid, cinnamic acid cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, Licylic acid and the like.

[0042] Pharmaceutically acceptable base addition salts can be formed with inorganic and organic bases. Inorganic bases from which salts can be derived include, for example, sodium, potassium, lithium , ammonium, calcium, magnesium, iron, zinc, copper, manganese, aluminum, etc. Ammonium, potassium, sodium, calcium, and magnesium Salts are particularly preferred. Organic bases from which salts can be derived include, for example, primary, secondary, and tertiary amines, naturally occurring substituted amines and other substituted amines, cyclic amines, basic ion exchange resins, etc. Specific examples of the amines include isopropylamine, trimethylamine, diethylamine, trimethylamine, and triethylamine. Triethylamine, tripropylamine, histidine, arginine, lysine, benethamine, N-methyl-glucamine, and ethanolamine. Other acids include dodecyl sulfate, Acid, naphthalene-1,5-disulfonic acid, naphthalene-2-sulfonic acid, and saccharin Examples include:

[0043] According to the present invention, in addition to the free base, methanesulfonic acid, maleic acid, isonicotine Acids, nicotinic acid, malonic acid, and salicylates, especially clofazimine methanesulfonate It is preferred to use:

[0044] As used herein, the term "pharmaceutical acceptable derivative" refers to any medicament that is, for example, as described in the U.S. Pat. It means the compounds disclosed in U.S. Pat. No. 9,540,336. The entire disclosure of No. 40,336 is incorporated herein by reference. In addition, the meaning of derivatives is given by Lu, Y., Zhen, M., Wang, B., Fu ,L.,Zhao,W.,Li,P.,Xu,J.,Zhu,H.,Jin,H.,Yi n, D., Huang, H., Upton, A. M., and Ma, Z., “Clofaz imine Analogs with Efficacy against expe rimental Tuberculosis and reduced Potent ial for Accumulation”Antimicrobial Agent s and Chemotherapy(2011),55(11):pp.5185- 5193. In addition, the term "pharmaceutical acceptable derivatives" of the compounds are For example, the compound may be a prodrug of the compound. Generally, a prodrug is a compound that is released after administration. Derivatives of the compound are capable of providing the active form of the compound. Derivatives of this type include, for example, For example, esters or amides of carboxyl groups, carboxyl esters of hydroxyl groups The phosphate ester may be an alkyl, aryl, or hydroxyl group.

[0045] "Therapeutically effective amount ) and "therapeutically effective dose" e)" or "pharmaceutical effective amount" "Clofazimine or a pharma- ceutical acceptable amount thereof" is used herein. The therapeutically effective dose of clofazimine is defined as the amount of the salt or derivative that is therapeutically effective. Thus, as used herein, a therapeutically effective amount is defined as an amount determined by clinical trial results. Judgment based on results and / or infection studies using animal models "Clofazimine" means the amount of clofazimine administered that produces the desired therapeutic effect.

[0046] The amount and daily dose of clofazimine can be routinely determined by one of skill in the art and may depend on a number of factors. This amount will vary depending on the organism, e.g., the particular microbial strain involved. The therapeutically effective amount for prophylactic treatment may also depend on the individual's height, weight, sex, age, and medical history. is an amount effective to prevent microbial infection.

[0047] A "therapeutic effect" means relief to some extent of one or more symptoms of an infection. This includes curing an infection. "Cure" means that the symptoms of an active infection have disappeared. Eliminated means that the excess of live bacteria responsible for the infection has been completely eliminated. This includes the complete or substantial disappearance of the substance below the detection threshold of conventional measurements. Certain long-term or permanent effects of an infection may exist even after a cure has been achieved. As used herein, a "therapeutic effect" refers to the reduction in the bacterial load of the host. A statistically significant decrease in the level of serotonin, development of resistance, or a statistically significant decrease in the level of serotonin as determined by human clinical results or animal studies. The treatment is defined as an improvement in symptoms of infection as determined by the patient.

[0048] As used herein, "treat" and "treatment" refer to "Treatment" or "treating" refers to the use of a pharmaceutical composition / combination for the prevention or treatment of a disease. and / or administration for therapeutic purposes.

[0049] The term "prophylactic treatment" is still used Patients who are not infected but are susceptible or otherwise at risk for a particular infection "Therapeutic treatment" refers to treating The term "treatment of patients who already have an infectious disease" refers to the administration of treatment to patients who already have an infectious disease. In a preferred embodiment, treatment involves treating a mammal (either for therapeutic or prophylactic purposes). The aim is to administer an effective amount of clofazimine.

[0050] Unless otherwise specified herein, the term "inhalation" means It means inhalation through the lungs.

[0051] Unless otherwise specified herein, the term "infection" as used herein The term is meant to refer to a lung infection.

[0052] Unless otherwise specified, the term "substantially" when used to refer to the purity of a compound means This indicates that the purity of a compound is greater than 95%.

[0053] Unless otherwise specified, the term "appropriate particle size" refers to the The particle size of the clofazimine or composition in the composition that provides the desired therapeutic effect when administered. Refers to...

[0054] Unless otherwise specified, the term "appropriate concentration" refers to a pharma- ceutical acceptable composition or combination. The term "composition" refers to the concentration of ingredients in a composition or combination that provides a beneficial effect.

[0055] Pharmaceutical Compositions and Combination Drugs The following water grades are particularly suitable for the present invention: sterile purified water, sterile injection water. Water for injection, sterile water for irrigation , sterile water for inhalation (US P) and, for example, the European Pharmacopoeia or Equivalent to water quality grades compliant with the National Formulary Something that is done.

[0056] The aqueous electrolyte solution used as the aqueous liquid carrier according to the present invention may further comprise sodium chloride, Potassium chloride, lithium chloride, magnesium chloride, calcium chloride, or mixtures thereof may include.

[0057] The aqueous liquid carrier is preferably isotonic saline (about 150 mM NaCl, preferably 0.9% NaCl, which corresponds to 154 mM NaCl).

[0058] Clofazimine has been shown to exist in at least four polymorphic forms (e.g., B annigan, et al., “Investigation into the S Solid and Solution Properties of Known an d Novel Polymorphs of the Antimicrobial Molecule Clofazimine”,Cryst.Growth Des.2 016,16(12),pp.7240-7250. Clofazimine is a triclinic It can exist in the FI form, the monoclinic FII form, and the orthorhombic FIII form. Some FIVs are also only seen at higher temperatures.

[0059] Thus, in a further embodiment of the invention: (a) a therapeutically effective dose of clofazimine; (b) a nonionic surfactant having a hydrophilic-lipophilic balance value greater than 10; (c) an aqueous liquid selected from water, isotonic saline, buffered saline, and aqueous electrolyte solutions; With carrier; A pharmaceutical composition comprising: The clofazimine is provided in the form of particles in a suspension, and The median particle size of clofazimine is less than 5 μm and the D90 is less than 6 μm, preferably Preferably, the median diameter is less than 2 μm and the D90 is less than 3 μm, and clofazimine The triclinic FI type, monoclinic FII type, and orthorhombic FIII type, as well as other types of this type Pharmaceutical compositions are provided in one or more polymorphs selected from the mixture. In an embodiment, clofazimine is provided in substantially orthorhombic Form FIII.

[0060] In further embodiments of the present invention, any of the composition embodiments described herein may be The pharmaceutical composition is provided by the method according to the present invention, wherein the non-ionic surfactant is polysorbate 20 (e.g., T Tween® 20, polysorbate 60 (e.g., Tween® 60 ), polysorbate 80 (e.g., Tween® 80), stearyl alcohol Hydrogenated castor oil with a hydrophilic-lipophilic balance value of 14-16 is used as a polyethylene glycol. derivatives (e.g., Cremophor® RH40), hydrophilic-lipophilic balan Hydrogenated castor oil polyethylene glycol derivatives with a Cre value of 15 to 17 (e.g., mophor® RH60), sorbitan monolaurate (e.g., Span( sorbitan monopalmitate (e.g., Span® 40); , sorbitan monostearate (e.g., Span® 60), polyoxyethylene (20) oleyl ether (e.g., Brij® 020), polyoxyethylene (20) cetyl ether (e.g., Brij® 58), polyoxyethylene (10) Cetyl Ether (e.g., Brij® C10), Polyoxyethylene (10) oleyl ether (e.g., Brij® O10), polyoxyethylene ethylene (100) stearyl ether (e.g., Brij® S100), poly Oxyethylene (10) stearyl ether (e.g., Brij® S10), Polyoxyethylene (20) stearyl ether (e.g., Brij® S20 ), polyoxyethylene (4) lauryl ether (e.g., Brij® L4) , polyoxyethylene (20) cetyl ether (e.g., Brij® 93), Polyoxyethylene (2) cetyl ether (e.g., Brij® S2), Caprylic / capric triglyceride Lylocaproyl polyoxyl-8 glyceride (e.g., Labrasol®) , polyethylene glycol (20) stearate (e.g., Myrj(TM) 49), poly Triethylene glycol (40) stearate (e.g., Myrj(TM) S40), poly Ethylene glycol (100) stearate (e.g., Myrj™ S100), poly Triethylene glycol (8) stearate (e.g., Myrj™ S8), and poly Oxyl 40 stearate (e.g., Myrj® 52), and mixtures thereof. are selected.

[0061] In another embodiment of the present invention, a composition according to any of the composition embodiments described herein is A pharmaceutical composition is provided, wherein the non-ionic surfactant is polysorbate 80, and the aqueous liquid carrier The solution is distilled water, hypertonic saline, or isotonic saline. In the case of the hypertonic saline, a pharmaceutical composition is provided, and the hypertonic saline is a solution of 1% to 7% (w / v) sodium chloride. In a further embodiment of the invention, a pharmaceutical composition is provided, comprising a non-ionic surfactant. The antibacterial agent was ultra-pure polysorbate 80 (e.g., NOF Corporation Po lysorbate 80 (Hx2)) and the aqueous liquid carrier is isotonic saline.

[0062] In another embodiment of the present invention, any one of the composition embodiments described herein may be The pharmaceutical composition according to the present invention has an osmolality of 200 to 700 mOsm / kg. In a further embodiment, the osmolality of the composition is in the range of 300 to 400 m in the Osm / kg range.

[0063] In a further embodiment of the invention, any one of the composition embodiments described herein The pharmaceutical composition according to the present invention is provided, and the nonionic surfactant is present in an amount of 0.001% to 5% of the total composition. % (v / v), and the amount of clofazimine ranges from 0.1% to 20% (w / v) of the total composition. / v).

[0064] In other embodiments of the present invention, any one of the composition embodiments described herein The pharmaceutical composition is provided by the method comprising the steps of: (1) A suspension of clofazimine, a nonionic surfactant, and water is mixed into a suitable particle size pellet of clofazimine. homogenizing to obtain a suspension containing azimine; (2) Adjust the pH of the suspension obtained from (1) to a pH between pH 5.5 and pH 7.5. and (3) adjusting the sodium chloride concentration to an appropriate concentration; (4) adjusting the osmolality to an appropriate level; It is prepared by a process comprising:

[0065] In a further embodiment, the pH is adjusted to 7.4 and the sodium chloride concentration is 154 mM In another embodiment, the homogenization in step (1) is carried out by high pressure This can be achieved by homogenization, high shear homogenization, wet milling, ultrasonic homogenization, or a combination of such processes. In another embodiment, the homogenization of clofazimine is carried out by multi-stage homogenization. In another embodiment, the clofazimine of suitable particle size has an average diameter of less than 5 μm. In a further embodiment, the particles are of suitable particle size. Azimin is a particle having a mean diameter of less than 2 μm and a D90 of less than 3 μm.

[0066] In a further embodiment of the invention, any one of the composition embodiments described herein The pharmaceutical composition is provided by the method comprising the steps of: (1) A suspension of clofazimine and a non-aqueous liquid is prepared by dissolving clofazimine in a suspension of suitable particle size. homogenizing to obtain a turbidity; (2) isolating clofazimine; and (3) adding clofazimine to a non-ionic surfactant and water; (4) Adjust the pH of the suspension obtained from (3) to a pH between pH 5.5 and pH 7.5. Steps and (5) adjusting the sodium chloride concentration to an appropriate concentration; It is prepared by a process comprising:

[0067] In a further embodiment, the pH is adjusted to 7.4 and the sodium chloride concentration is In a further embodiment, the homogenized solution in step (1) is adjusted to 154 mM thorium. The homogenization may be performed by high pressure homogenization, high shear homogenization, wet milling, ultrasonic homogenization, or a combination of such processes. In another embodiment, the homogenization of clofazimine is carried out by a multi-stage homogenization process. In another embodiment, the clofazimine having the appropriate particle size is The particles have an average diameter of less than 5 μm and a D90 of less than 6 μm. Therefore, clofazimine having a suitable particle size has a mean diameter of less than 2 μm and a D90 of 3. Particles smaller than μm.

[0068] In further embodiments, any one of the composition embodiments described herein. A pharmaceutical composition is provided, the composition comprising the steps of: (1) Micronizing clofazimine to obtain clofazimine of a suitable particle size and, (2) adding clofazimine to a non-ionic surfactant and water; (3) Adjust the pH of the suspension obtained from (2) to a pH between pH 5.5 and pH 7.5. and (4) adjusting the sodium chloride concentration to an appropriate concentration; It is prepared by a process comprising:

[0069] In a further embodiment, the pH is adjusted to 7.4 and the sodium chloride concentration is 154 mM Adjusted to sodium chloride.

[0070] In another embodiment, the micronization of clofazimine is carried out by jet milling, spray drying, boiling, or the like. In another embodiment, the clofa is treated with a cycloaliphatic solvent, such as cycloaliphatic solvent, by milling, or by supercritical fluid processing. Micronization of the dimine is carried out by multi-stage micronization. In another embodiment, the appropriate particle size The clofazimine particles have a mean diameter of less than 5 μm and a D90 of less than 6 μm. In a further embodiment, the suitable particle size of clofazimine is less than 2 μm in average diameter. and the particles have a D90 of less than 3 μm.

[0071] In further embodiments, any one of the composition embodiments described herein. A pharmaceutical composition is provided, the composition comprising clofazimine in a non-ionic surfactant, suitable In water containing a suitable concentration of sodium chloride and adjusted to a pH between 5.5 and 7.5 The suspension is subjected to a process including homogenization to obtain clofazimine of a suitable particle size. In a further embodiment, the pH is adjusted to 7.4 and sodium chloride is added. The sodium concentration is adjusted to 154 mM sodium chloride. can be achieved by high pressure homogenization, high shear homogenization, wet milling, ultrasonic homogenization, or a combination of such processes. In another embodiment, the homogenization of clofazimine is carried out by multi-stage homogenization. In another embodiment, the clofazimine of suitable particle size is obtained by dissolving the clofazimine in a suspension having a mean particle size of about 100 mm. In a further embodiment, the diameter of the pores is less than 5 μm and the D90 is less than 6 μm. The particle size of clofazimine is less than 2 μm in mean diameter and less than 3 μm in D90. be.

[0072] In other embodiments, a pharmaceutical composition according to any of the composition embodiments described herein. 1. A process for preparing a composition comprising the steps of: (1) A suspension of clofazimine, a nonionic surfactant, and water is mixed into a suitable particle size pellet of clofazimine. homogenizing to obtain a suspension containing azimine; (2) Adjust the pH of the suspension obtained from (1) to a pH between pH 5.5 and pH 7.5. and (3) adjusting the sodium chloride concentration to an appropriate concentration; (4) adjusting the osmolality to an appropriate level; A process is provided that includes:

[0073] In another embodiment, the pH is adjusted to 7.4 and the sodium chloride concentration is 154 mM sodium chloride. In a further embodiment, the homogenization is performed by high pressure homogenization, wet milling, , ultrasonic homogenization, or a combination of such treatments. In a further embodiment, the homogenization of clofazimine is carried out by multi-stage homogenization. Therefore, suitable particle size of clofazimine is a mean diameter of less than 5 μm and a D90 of less than 6 μm. In another embodiment, the clofazimine of suitable particle size has an average diameter of less than 2 μm. m, and D90 is less than 3 μm.

[0074] In other embodiments, for preparing the pharmaceutical composition embodiments described herein, A process comprising: (1) A suspension of clofazimine and a non-aqueous liquid is prepared by dissolving clofazimine in a suspension of suitable particle size. homogenizing to obtain a turbidity; (2) isolating clofazimine; and (3) adding clofazimine to a non-ionic surfactant and water; (4) Adjust the pH of the suspension obtained from (3) to a value between pH 5.5 and pH 7.5. and adjusting the (5) adjusting the sodium chloride concentration to an appropriate concentration; A process is provided that includes:

[0075] In another embodiment, the pH is adjusted to 7.4 and the sodium chloride concentration is 154 mM sodium chloride. In a further embodiment, the homogenization is performed by high pressure homogenization, wet milling, , ultrasonic homogenization, or a combination of such treatments. In a further embodiment, the homogenization of clofazimine is carried out by multi-stage homogenization. Therefore, suitable particle size of clofazimine is a mean diameter of less than 5 μm and a D90 of less than 6 μm. In another embodiment, the clofazimine of suitable particle size has an average diameter of less than 2 μm. m and D90 is less than 3 μm.

[0076] In further embodiments, any one of the pharmaceutical composition embodiments described herein A process for preparing a pharmaceutical composition according to the present invention, comprising the steps of: (1) Micronizing clofazimine to obtain clofazimine of a suitable particle size and, (2) adding clofazimine to a non-ionic surfactant and water; (3) Adjust the pH of the suspension obtained from (2) to a pH between pH 5.5 and pH 7.5. and (4) adjusting the sodium chloride concentration to an appropriate concentration; A process is provided that includes:

[0077] In another embodiment, the pH is adjusted to 7.4 and the sodium chloride concentration is 154 mM In a further embodiment, the micronization of clofazimine is adjusted to sodium chloride. This can be accomplished by wet milling, spray drying, ball milling, or supercritical fluid processing. In another embodiment, the micronization of clofazimine is carried out by multi-stage micronization. In a further embodiment, the suitable particle size of clofazimine is less than 5 μm in average diameter; In another embodiment, the clofazidazole of suitable particle size is used. Min is a particle with a mean diameter of 2 μm and a D90 of less than 3 μm.

[0078] In other embodiments, any one of the pharmaceutical composition embodiments described herein is 2. A process for preparing a pharmaceutical composition comprising: It contains a stimulant, an appropriate concentration of sodium chloride, and the pH is adjusted to between pH 5.5 and pH 7.5. The suspension in water is homogenized to obtain the appropriate particle size of clofazimine. In another embodiment, the pH is 7.4 and sodium chloride is A suitable concentration of sodium is 154 mM sodium chloride. The homogenization is carried out by high pressure homogenization, wet grinding, ultrasonic homogenization, or a combination of such processes. In a further embodiment, the homogenization of clofazimine is carried out by multi-stage homogenization. In a further embodiment, the suitable particle size of clofazimine is less than 5 μm in average diameter. In another embodiment, the particles are of suitable particle size. Fazimin is a particle having a mean diameter of 2 μm and a D90 of less than 3 μm.

[0079] In further embodiments, a pharmaceutical composition according to any one of the composition embodiments described herein is provided. A process for preparing a pharmaceutical composition comprising the steps of: (a) administering clofazimine , a non-ionic surfactant, and water suspension to form a suspension containing clofazimine of suitable particle size. (b) homogenizing the resulting suspension to obtain a pH of 5.5 to 7.5; (c) adjusting the sodium chloride concentration to an appropriate concentration; (d) adjusting the osmolality to an appropriate level; Steps (b), (c) and (d) are: (b), (c), (d); (b), (d), (c );(c), (b), (d);(c), (d), (b);(d), (b), (c); or A process is provided that can be performed in the order of (d), (c), and (b).

[0080] In other embodiments, a medicament according to any one of the composition embodiments described herein. A process for preparing a composition comprising the steps of: (a) combining clofazimine and The suspension in the non-aqueous liquid is homogenized to obtain a suspension containing clofazimine of appropriate particle size. (b) isolating clofazimine; and (c) isolating clofazimine. (d) adjusting the pH of the resulting suspension to pH 6.0 or higher; (e) adjusting the pH to between pH 5.5 and pH 7.5; and and adjusting the concentration of the cellulose to a suitable concentration; and steps (d) and (e) are ), (e); or (e), (d).

[0081] In other embodiments, a pharmaceutical composition according to any one of the composition embodiments described herein. A process for preparing a pharmaceutical composition comprising the steps of: (a) administering clofazimine (b) micronizing the clofazimine to obtain a suitable particle size; and The solution contains a non-ionic surfactant, an appropriate concentration of sodium chloride, and a pH of 5.5. and adding to water adjusted to between 7.5 and 7.5°C.

[0082] In another embodiment of the present invention, a pharmaceutical combination in the form of an inhalable aerosol is A composition according to any one of the composition embodiments described herein is administered using an ultrasonic nebulizer. -Electron spray nebulizer, vibrating membrane nebulizer, A nebulizer selected from a jet nebulizer and a mechanical soft mist metered dose inhaler ( It is prepared by aerosolization using a nebulizing device. A combination medicament is provided, and the aerosol particles generated by the nebulizer device are In a further embodiment, the inhalable aerosol is administered to the lower respiratory tract. In another embodiment, the spray device has a spray velocity of 0.1 to 1.5. In another embodiment, the total inhalation volume is between 1 ml and 5 ml. do.

[0083] In other embodiments, a pharmaceutical composition according to any one of the composition embodiments described herein. A pharmaceutical composition comprising nebulized 4-7% hypertonic saline, metaperiodate, dodecyl Sodium sulfate, sodium bicarbonate, tromethamine, silver nanoparticles, bismuth-thiol , ethylenediaminetetraacetic acid, gentamicin-loaded phosphatidylcholine-modified gold nanoparticles ( gentamicin loaded phosphatidylcholine-de Coated gold nanoparticles), chelating agent, cis-2-de D-enantiomers, D-amino acids, and peptides containing D-amino acid residues c peptide), gallium mesoporphyrin IX, gallium protoporphyrin I X, curcumin, patulin, penicillic acid, baicalein, naringenin, ursolic acid , asiatic acid, corosolic acid, fatty acids, host defense peptides, and antimicrobial peptides. Selected agents for dispersing and / or disrupting biofilms, mucolytic agents and / or or in combination with a mucoactive agent and / or an agent that reduces biofilm formation. In another embodiment, a pharmaceutical composition for this use is provided. The substance is bedaquiline or a pharma- ceutical acceptable salt or derivative thereof, cefoxitin (cefoxitin foxitine), amikacin, clarithromycin, pyrazinamide, rifampin , moxifloxacin, levofloxacin, and para-aminosalicylates, and The compound is administered before, simultaneously with, or after the administration of an agent selected from the mixture of compounds listed above.

[0084] In other embodiments, a combination therapy according to any of the combination embodiments described herein. The drug is aerosolized 4-7% hypertonic saline, metaperiodate, sodium dodecyl sulfate, Thorium, sodium bicarbonate, tromethamine, silver nanoparticles, bismuth-thiol, ethyl Diaminetetraacetic acid, gentamicin-loaded phosphatidylcholine modified gold nanoparticles, chiral cis-2-decenoic acid, D-amino acids, peptides containing D-amino acid residues, gallium Mesoporphyrin IX, Gallium Protoporphyrin IX, Curcumin, Patulin, Penicillin Syrinic acid, baicalein, naringenin, ursolic acid, asiatic acid, corosolic acid, The biofilm-dissolving peptides are selected from fatty acids, host defense peptides, and antimicrobial peptides. Dispersing and / or destroying agents, mucolytic and / or mucoactive agents, and / or A combination pharmaceutical for use in combination with an agent for reducing biofilm formation is provided. In another embodiment, the combination for this use comprises bedaquiline or a pharma- ceutical acceptable salt thereof. Acceptable salts or derivatives, cefoxitine, amikacin, Larithromycin, pyrazinamide, rifampin, moxifloxacin, levofloxacin and administering an agent selected from para-aminosalicylate, para-aminosalicylate, and mixtures thereof. It is used before, simultaneously with, or after administering the composition of the present invention. The composition comprises bedaquiline or a pharma- ceutical acceptable salt or derivative thereof and amine. The present invention relates to a method for treating a pulmonary circulation disorder, the method for treating a pulmonary circulation disorder, and the method for treating a pulmonary circulation disorder. In a further embodiment, the composition comprises bedaquiline or a pharma- ceutical acceptable salt thereof. or a derivative thereof.

[0085] In other embodiments, a pharmaceutical composition according to any one of the composition embodiments described herein. The pharmaceutical composition is for treating pulmonary infections caused by mycobacteria or other gram-positive bacteria. In a further embodiment, the method is provided for use in the treatment and / or prophylaxis of an infection. The microbial composition of nontuberculous mycobacteria and Mycobacterium tuberculosis (Mycobacterium tuberculosis) and combinations thereof. Mycobacterium species In a further embodiment, the nontuberculous acid-fast bacterium is a mycobacterium. Mycobacterium avium, Mycobacterium in Mycobacterium intracellulare, Mycobacterium Mycobacterium abscessus, and Mycobacterium leprae e)), and combinations thereof. MAC pulmonary disease and nontuberculous disease in patients with cystic fibrosis, chronic obstructive pulmonary disease, or acquired immune deficiency syndrome Selected from nontuberculous infections In another embodiment, the infection is a non-cancerous infection in a cystic fibrosis patient. In another embodiment, the composition for this use is bedaquiline or its pharma- ceutical acceptable salts or derivatives, cefoxitin (cef oxitine), amikacin, clarithromycin, pyrazinamide, rifampin, Moxifloxacin, levofloxacin, and para-aminosalicylates, and In another embodiment, the compound is administered before, simultaneously with, or after administration of an agent selected from the following mixture: The composition comprises bedaquiline or a pharma- ceutical acceptable salt or derivative thereof, and The drug is administered before, simultaneously with, or after administration of an agent selected from the group consisting of mikacin, and mixtures thereof. In a further embodiment, the composition comprises bedaquiline or a pharma- ceutical acceptable salt thereof. It may be administered before, simultaneously with, or after administration of a salt or derivative.

[0086] In other embodiments, a combination according to any of the combination embodiments described herein. The medicament is for treating lung infections caused by mycobacteria or other gram-positive bacteria. and / or for use in the prevention of infection. Nontuberculous mycobacteria and Mycobacterium tuberculosis (Mycobacterium tuberculosis) Mycobacterium tuberculosis) and combinations thereof. Triggered by species of the genus Mycobacterium In a further embodiment, the nontuberculous mycobacterium is Mycobacterium abioticum. Mycobacterium avium, Mycobacterium intracellulare Mycobacterium intracellulare, Mycobacterium Mycobacterium abscessus, and Mycobacterium leprae (Mycobacterium leprae), and In another embodiment, the infection is selected from cystic fibrosis. Pulmonary MAC disease and nontuberculous anti-tuberculous antibodies in patients with pulmonary tuberculosis, chronic obstructive pulmonary disease, or acquired immune deficiency syndrome In another embodiment, the infection is an opportunistic infection selected from mycobacterial infections. In another embodiment, the present invention relates to opportunistic infections caused by nontuberculous mycobacteria in patients with chronic fibrosis. The combination for this use comprises bedaquiline or a pharma- ceutical acceptable salt or derivative thereof. , cefoxitine, amikacin, clarithromycin, pyrazinamide Aminoside, rifampin, moxifloxacin, levofloxacin, and para-aminosalicylic acid The present invention is directed to a method for treating a patient suffering from atopic dermatitis, comprising administering to the patient a therapeutically effective amount of the present invention before, simultaneously with, or after administering an agent selected from the group consisting of rithyrate, ricinate, and mixtures thereof. In another embodiment, the combination for this use is The drugs are bedaquiline or a pharma- ceutical acceptable salt or derivative thereof, and amikacin, and administering the composition of the present invention before, simultaneously with, or after administering an agent selected from these mixtures to In another embodiment, the combination for this use comprises bedaquiline. or a pharma- ceutical acceptable salt or derivative thereof, It is used to administer the composition.

[0087] In another embodiment, the antibody is caused by mycobacteria or other gram-positive bacteria. A system is provided for use in providing antibacterial activity in treating or preventing pulmonary infections, The system is: 1) (a) a therapeutically effective dose of clofazimine; (b) a nonionic surfactant having a hydrophilic-lipophilic balance value greater than 10; (c) an aqueous liquid selected from water, isotonic saline, buffered saline, and aqueous electrolyte solutions; With carrier; an aerosolized combination pharmaceutical comprising: 2) a sprayer; Including, The clofazimine is present in the form of a suspension, The aerosol particles generated by this system have mass median aerodynamic diameters of 1-5 μm.

[0088] In further embodiments, the present invention relates to a pulmonary fungal infection or Clostridium difficile ( Treatment and / or prevention of Clostridium difficile (Clostridium difficile) or a combination of these A pharmaceutical composition according to any one of the composition embodiments described herein for use in the prevention of In another embodiment, a method for treating and / or preventing a pulmonary fungal infection is provided. There is provided a pharmaceutical composition according to any one of the composition embodiments described herein for In a further embodiment, the pulmonary fungal infection is Candida albicans. da albicans) or Aspergillus fumigatus fumigatus), or a combination thereof.

[0089] In further embodiments, the present invention relates to a pulmonary fungal infection or Clostridium difficile or and combinations thereof for use in the treatment and / or prevention of The pharmaceutical combination according to any one of the embodiments is provided. A combination therapy according to any one of the combination embodiments described herein for use in the prevention of In a further embodiment, the pulmonary fungal infection is Candida albicans ( candida albicans or Aspergillus fumigatus gilus fumigatus), or a combination thereof.

[0090] In another embodiment, a method for treating or preventing a pulmonary infection in a patient in need thereof. A method comprising inhaling a composition according to any one of the composition embodiments described herein. In another embodiment, the infection is The microbial composition of nontuberculous mycobacteria and Mycobacterium tuberculosis (Mycobacterium tuberculosis) and combinations thereof. Mycobacterium species In a further embodiment, the nontuberculous mycobacterium is caused by Mycobacterium tuberculosis. Mycobacterium avium, Mycobacterium intracellulare Mycobacterium intracellulare, Mycobacterium Mycobacterium abscessus, and Mycobacterium leprae (Mycobacterium leprae) In a further embodiment, the infection is selected from the group consisting of bursitis, Pulmonary MAC disease and pulmonary pulmonary disease in patients with cystic fibrosis, chronic obstructive pulmonary disease, or acquired immune deficiency syndrome and nontuberculous mycobacterial infections. Mycobacterium tuberculosis is an opportunistic infection caused by nontuberculous mycobacteria in patients with cystic fibrosis.

[0091] In a further embodiment, the present invention relates to the treatment of mycobacteria or other infections in a patient in need thereof. A method for treating or preventing a pulmonary infection caused by a gram-positive bacterium, comprising administering to a subject a pulmonary infection comprising administering to said subject a pulmonary infection a gram-positive bacterium. salts of phosphorus or its pharma- ceutically acceptable derivatives, cefoxitine , amikacin, clarithromycin, pyrazinamide, rifampin, moxifloxacin levofloxacin, and para-aminosalicylates, and mixtures thereof. any of the composition embodiments described herein before, simultaneously with, or after administration of the agent to be treated. Methods are provided that include administering by inhalation a composition according to one of the methods described above. In an embodiment, the agent is bedaquiline or amikacin. , this agent is bedaquiline.

[0092] Particle size and distribution The therapeutic effect of aerosol therapy depends on the dose deposited and its distribution. The particle size of the aerosol determines the deposition and distribution of the drug aerosol in the lungs. It is one of the important variables.

[0093] In general, inhaled aerosol particles are dispersed by two mechanisms: collisions (usually by larger aerosol particles); aerosols) and settling (dominant for smaller aerosol particles). Collisions occur when the momentum of an inhaled aerosol particle is transferred to the airstream, causing the particle to settle in the air. This occurs when the surface is large enough to not conform to the physiological surface. In contrast, sedimentation occurs when the water is carried by the intake air current. The result of gravity settling of very small aerosol particles, primarily in the lower respiratory system This occurs when the ion reaches the physiological surface as

[0094] Drug delivery to the lungs can be achieved by inhaling an aerosol through the oropharynx. Aerosol particles with an aerodynamic diameter greater than about 5 μm generally do not reach the lungs. Instead, they tend to impact the back of the throat and may be swallowed and absorbed orally. Aerosol particles with a diameter of about 3 to about 5 μm are distributed in the upper-middle airway region. Small enough to reach the mid-pulmonary region (conducting airways) However, they are too large to reach the alveoli. Aerosol particles can reach the alveolar region. Air particles smaller than about 0.5 μm in diameter During quiet breathing, osol particles tend to be expelled with breathing, but when breath-holding is performed, they are expelled from the alveolar region. It is also possible to deposit the

[0095] Aerosols used for pulmonary drug delivery consist of a wide range of aerosol particle sizes. Statistical descriptors are used to characterize the distribution of aerosols in pulmonary drug delivery. The mass median diameter (MMD) of the aerosol particles is expressed as the mass median diameter (MMD) of the aerosol particles. D and half of the mass contained in the aerosol particles is less than MMD. If the diameter is uniform, the volume median diameter (VMD) can be used interchangeably with the MMD. The VMD and MMD are determined using laser diffraction. The width of the distribution is the geometric standard deviation. On the other hand, deposition of aerosol particles in the respiratory tract is expressed as the aerodynamic velocity of the particles. The aerodynamic mass median diameter is usually used since it is more accurate to use the diameter. D measurements are performed by inertial impaction or time-of-flight measurements. For water particles, VMD, MMD , and MMAD should be equal. However, If humidity is not controlled, dehydration will occur and the MMAD reading will be lower than the MMD and VM readings. D. To use this notation, VMD, MMD, and MM Measurement of AD is based on comparable descriptions of VMD, MMD, and MMAD. ) is deemed to be carried out under controlled conditions.

[0096] However, for purposes of illustration, the aerosol particle size of the aerosol particles is determined according to the U.S. Pharmacopoeia. Compliant with the US Pharmacopeial Convention and next-generation Using the Next Generation Impactor (NGI) The MMAD will be given as the MMAD determined by measuring at room temperature (In Pr Ocess Revision <601> Aerosols, Nasal Spray ys, Metered-Dose Inhalers,and Dry Powder Inhalers,Pharmacopeial Forum(2003),Volu me Number 29, pages 1176-1210, Jolyon Mitch hell, Mark Nagel“Particle Size Analysis of Aerosols from Medicinal Inhalers”,KON A Powder and Particle Journal(2004),Volu (See also 22, pages 32-65.)

[0097] According to the present invention, the particle size of the aerosol is selected to maximize deposition of clofazimine at the site of infection. The aerosol particle size is determined by the aerodynamic mass It can be expressed in terms of median diameter (MMAD). Large particles (e.g., MMAD>5μ m) are too large to follow the bends of the airways and therefore tend to be deposited in the extrathoracic and upper airways. Deposition of large particles in the upper respiratory tract may result in intolerance symptoms (e.g. coughing and bronchospasm). There is a possibility that.

[0098] Therefore, according to a preferred embodiment, the MMAD of the aerosol should be less than about 5 μm. Preferably, the diameter is between about 1 and 5 μm, more preferably less than 3 μm (<3 μm). It is.

[0099] However, guided breathing maneuvers ) allows larger particles to pass through the extrathoracic cavity and upper airways, resulting in a greater Allows deep lung penetration, thereby distributing aerosols to the middle and lower respiratory tract This will increase deposition in the blood. It is also possible to slow breathing to 100 ml / min with guidance. Therefore, when using supervised breathing techniques, the preferred MMAD for aerosols is It should be less than about 10 μm.

[0100] Another factor that is equally important (besides the aerosol size) is the size and size distribution of the solid particles. The particle size and distribution of the solid particles in a given aerosol particle is The particle size of the aerosol must be smaller than that of the aerosol particles that contain it. Sol particles may contain one or more solid particles. Furthermore, dilute suspensions can be handled using In such cases, the majority of the aerosol particles may not contain solid particles.

[0101] Therefore, it is possible to use solid drug particles that are significantly smaller than the MMAD of the aerosol particles. For example, if the MMAD of the aerosol particles is 3 μm, the solid particles should be 1 μm or smaller. It would be preferable to lower

[0102] Other considerations include, for example, the use of vibrating mesh nets. In the case of using a blister, the formulation is delivered through holes in a plate, which The pores break up the suspension into small droplets. For this reason, the solid particles are then forced to pass through the pores. In addition, it is also necessary to make the thickness smaller than the hole.

[0103] The particle size of the solids in the suspension can be given by the average particle size and by the particle distribution. The D90 value indicates that 90% of the particles in a suspension are below the mean diameter. .

[0104] sprayer A variety of nebulizers (small volume nebulizers) for aqueous and other non-pressurized liquid systems (including small volume nebulizers) to aerosolize the formulation. Compressor-driven atomizers incorporate jet atomization technology. These devices use compressed air to generate a liquid aerosol. ,Healthdyne Technologies,Inc.;Invacare,I nc.;Mountain Medical Equipment,Inc.;Pari Respiratory,Inc.;Mada Medical,Inc.;Puri tan-Bennet;Schuco,Inc.,DeVilbiss Health Ultrasonic nebulizers are available from Eppendorf Care, Inc. and Hospitalak, Inc. The pump uses mechanical energy in the form of vibrating piezoelectric crystals to generate inhalable droplets. For example, Omron Heathcare, Inc. and DeVil Commercially available from Biss Health Care, Inc. Vibrating mesh nebulizer The laser uses either piezoelectric or mechanical pulses to generate inhalable droplets. Other examples of nebulizers for use with clofazimine as described herein include: U.S. Patent No. 4,268,460; U.S. Patent No. 4,253,468; U.S. Patent No. Patent No. 4,046,146; U.S. Pat. No. 3,826,255; U.S. Pat. No. 4,649,911; U.S. Pat. No. 4,510,929; U.S. Pat. ,624,251; U.S. Pat. No. 5,164,740; U.S. Pat. No. 86,550; U.S. Pat. No. 5,758,637; U.S. Pat. No. 6,644 ,304; U.S. Pat. No. 6,338,443; U.S. Pat. No. 5,906,2 No. 02; U.S. Patent No. 5,934,272; U.S. Patent No. 5,960,792 No. 5,971,951; No. 6,070,575 Fine print; U.S. Patent No. 6,192,876; U.S. Patent No. 6,230,706 ;U.S. Patent No. 6,349,719;U.S. Patent No. 6,367,470;U.S. Patent No. US Patent No. 6,543,442; US Patent No. 6,584,971; US ​​Patent No. 6,601,581; U.S. Pat. No. 4,263,907; U.S. Pat. No. 5,709,202; U.S. Pat. No. 5,823,179; U.S. Pat. No. 192,876; U.S. Pat. No. 6,644,304; U.S. Pat. No. 5,54 No. 6,161, 536; U.S. Pat. No. 6,264,922; U.S. Pat. No. 6,557,54 No. 9; and U.S. Pat. No. 6,612,303, all of which are incorporated herein by reference. The entire contents of the above references are incorporated herein by reference. Examples of commercially available nebulizers that can be used with lofazimine compositions include Aerosols. Gen's Respirgard II®, Aeroneb®, and Ae Aeroneb® Pro, and Aeroneb® Go; Aradigm AERx® and AERx Essence® manufactured by I-neb Resp Ironics, Inc. Porta-Neb (registered trademark), Freeway Fre edom(TM), Sidestream, Ventstream; and PARI,G mbH PARI LCPlus (registered trademark), PARI LC-Star (registered trademark) Other non-limiting examples include U.S. Pat. This is disclosed in specification No. 9.

[0105] According to the present invention, the pharmaceutical composition is preferably administered by ultrasonic nebulizer, electrostatic nebulizer, etc. Nebulizer, vibrating membrane nebulizer, jet nebulizer, or mechanical soft mist metered dose aspiration The composition can be aerosolized using a nebulizer device selected from the group consisting of a nebulizer, aerosolizer, and a nebulizer.

[0106] It is preferred that the device controls the patient's inhalation rate either electrically or mechanically. I wish.

[0107] In a further preferred embodiment, the generation of the aerosol by the device is carried out by an AKITA device. Like the AKITA device, it is activated by the patient's inhalation.

[0108] A preferred (commercially available) example of the above-mentioned sprayer / instrument to be used according to the present invention is the Vectu ra fox, Pari eFlow, Pari Trek S, Philips In nospire mini, Philips InnoSpire Go, Medspr ay device, Aeroneb Go, Aerogen Ultra, Respi ronics Aeroneb, Akita, Medspray Ecomyst, and It's Respimat.

[0109] Therapeutic and / or prophylactic uses Pharmaceutical compositions and combination pharmaceuticals according to the present invention (aerosols, aerosolized formulations) and The system is designed to detect Mycobacteria or Staphylococcus aureus (Sta Phylococcus aureus (methicillin-resistant and vancomycin-intermediate resistant) strains), Streptococcus pneumoniae (Streptococcus cus pneumoniae), and enterococci (Enterococcus spp. Pulmonary infections caused by other clofazimine-susceptible bacteria, such as coccus spp. The pharmaceutical compositions of the present invention are intended for use in the treatment and / or prevention of infectious diseases. The pharmaceutical combination may also be used to treat and / or prevent pulmonary fungal infections.

[0110] Dosing of clofazimine According to the present invention, the pharmaceutical composition of the present invention is prepared by administering about 1 to 5 ml, preferably 1 to 2 ml of the pharmaceutical composition of the present invention. Delivered by nebulization of 2 ml.

[0111] Therefore, based on the concentration of clofazimine in the pharmaceutical composition being about 20 mg / ml, The target fill dose is equivalent to 20-100 mg of clofazimine. The amount is about 1 to 5 ml.

[0112] The daily lung dose of clofazimine administered according to the present invention is se (i.e., the amount of drug deposited in the lungs) was In the case of infection, the dose is about 5-10 mg, which is the nominal dose of 15-30 mg (the dose administered by the device). This corresponds to the amount of drug.

[0113] Those skilled in the art will appreciate that the pulmonary dose of clofazimine to be administered (and thus the loading dose / nominal dose) The dose / volume of spray was determined by the respective fractions of clofazimine, which are well established in the art. It is reasonable to expect that the formulation will be adjusted automatically based on the minimum inhibitory concentration (MIC) for the strain. Let it be understood.

[0114] Therefore, the daily pulmonary dose will be divided according to the frequency of administration, either once or twice daily. do.

[0115] According to the present invention, clofazimine results in a total pulmonary dose of about 5-10 mg per day. The dose is administered once or twice daily so as to

[0116] The amounts stated above relate to the free base of clofazimine; the dosages for derivatives and salts are It will be apparent to one skilled in the art that the MICs must be adjusted based on the MIC of each compound and strain. Wax.

[0117] Mucolytic / biofilm modifying agents ) Reduces sputum viscosity during aerosol treatment and destroys existing biofilms For this purpose, the treatment and / or prevention according to the present invention may be achieved by using mucolytic agents and / or biologics. An additional dose of a film disrupting agent may be included.

[0118] These agents may be formulated as a fixed combination or or simultaneously with or in combination with a pharmaceutical composition / aerosol containing clofazimine according to the present invention. This may be followed by administration.

[0119] Agents for dispersing / disrupting biofilms, mucolytic agents and / or or mucoactive agents and / or agents that reduce biofilm formation are administered by nebulizing the ~7% hypertonic saline, metaperiodate, sodium dodecyl sulfate, sodium bicarbonate , tromethamine, silver nanoparticles, bismuth-thiol, ethylenediaminetetraacetic acid, gentamicin Mycin-loaded phosphatidylcholine modified gold nanoparticles, chelating agent, cis-2-decenoic acid, D-amino acids, peptides containing D-amino acid residues, gallium mesoporphyrin IX, gallium Umprotoporphyrin IX, curcumin, patulin, penicillic acid, baicalein, Lingenin, ursolic acid, asiatic acid, corosolic acid, fatty acids, host defense peptides, and antimicrobial peptides.

[0120] Additionally, other pharma- ceutical active agents may be used in combination with the pharmaceutical composition / aerosol combination according to the present invention. Such an active agent may be bedaquiline or a pharma- ceutical acceptable salt or derivative thereof. Cefoxitine, amikacin, clarithromycin, pyrimidine Zinamide, rifampin, moxifloxacin, levofloxacin, and para-amino salicylates, and mixtures thereof.

[0121] These agents can be formulated as combinations or can be used in combination with the clotting agent of the present invention. It may also be administered before, simultaneously with, or after the pharmaceutical composition / aerosol combination containing Fazimin. Cut.

[0122] The following examples more fully illustrate the manner of using the above-described invention and further illustrate the invention. It serves to illustrate the best modes contemplated for carrying out various aspects of the present invention. Embodiments in accordance with the invention are encompassed within the claims set forth herein. EXAMPLES

[0123] test The following exemplary compositions and combinations were prepared according to the processes described herein.

[0124] Example 1 Clofazimine (as triclinic form I) 200 mg, sodium chloride 90 mg, and water 9.5 ml, homogenized in an Ultra-Turrax homogenizer at 10,000 rpm for 5 min. The mixture was mixed twice. 0.5 ml of polysorbate 80 (NOF Hx2) was added. The mixture was subjected to an ultrasonic probe (Bandelin S Branson Digital with Onoplus Probe MS73 The sample was processed 7 times (3 minutes each) at 70% amplitude using a Sonifier (trademark 250D). The suspension was then filtered through a VWR folded qualitative filter paper (303, particle retention capacity: 10 ml). The composition of Example 1 was obtained by filtering through a sieve with a filter (size: 150 mm) of 5 to 13 μm. The median particle size of clofazimine in the composition of Example 1 is 3.9 μm and the D90 is 6.7 μm The concentration of clofazimine was measured by UV / Vis spectroscopy at 280 nm and migration By calibrating with a 1 mg / ml stock solution of clofazimine diluted in phase 1, 7.16 mg / ml.

[0125] The composition of Example 1 is shown in Table 1.

[0126] [Table 1]

[0127] Preparation of orthorhombic form III of clofazimine. A slurry of clofazimine (10 g) in toluene (20 ml) was placed in a mag- netic oil bath at 40°C. The slurry was stirred at 800 rpm for 72 hours using a thermostatic stirrer. The mixture was filtered through a crucible and then placed in an oven at a maximum temperature of 40°C. This resulted in the production of substantially pure (≧98%) orthorhombic form III chromium. The yield of azimine was 8.64 g.

[0128] Example 2 Water containing 0.5% polysorbate 80 (NOF Hx2) and 0.6% sodium chloride A suspension containing 6 g of clofazimine in orthorhombic form III per 100 ml was prepared using Ultra-T The mixture was pre-micronized using Urrax® at 10,000 rpm for approximately 40 seconds. Preliminary preparation: 0.6% sodium chloride in water to a volume of 300 ml. 300 ml of this suspension was homogenized in a homogenizer, M-110EH-30 ml. icrofluidizer (Microfluidics, Westwood, MA, The suspension was then circulated in the H30Z chamber at 5,000 psi for 15 minutes. A pre-homogenization step was then performed by placing the second H10Z chamber in Attached in series to the first chamber, the suspension was homogenized at 25,000 psi for an additional 23 minutes. Particle size analysis was performed using HORIBA LA 950, and the median particle size was 0. UV / Vis spectroscopy showed a chromatic aberration of 0.83 μm and a D90 value of 1.17 μm. The measurement was performed at 280 nm and calibrated with a 1 mg / ml stock solution of clofazimine diluted in mobile phase. By this, the concentration of clofazimine was determined to be 16.05 mg / ml.

[0129] The composition of Example 2 is shown in Table 2.

[0130] [Table 2]

[0131] Example 3 Clofazimine (crystal modification: orthorhombic form III) was dissolved in water, sodium chloride, and polyisoprene. The suspension was suspended in a solution of Rubeto 80 and then filtered using the M-110EH-30 Microfluidic Using dizer (registered trademark) Processor (Chamber: H30Z and G10Z) The H30Z-G10Z configuration was operated at a pressure of 28,250 psi for 30 minutes. This produced the composition of Example 3. The resulting clofazimine particles had a median particle size of 1.28 μm. m and D90 was less than 2 μm.

[0132] The composition of Example 3 is shown in Table 3.

[0133] [Table 3]

[0134] Viscosity measurement The viscosity of the composition of Example 3 was measured by stress control using a STRESSTECH Rheometer. The measurements were performed in a controlled mode. A hollow cylindrical fixture (double gap geometry) was used. The spindle is then rotated continuously to measure the temperature at each point. The viscosity was measured at 0 The pressures were 0.01, 0.05, and 0.1 Pa, and the measurements were performed at 20°C, 25°C, and 30°C, respectively. The average of the viscosities obtained by performing two separate sample loadings. The values ​​are shown in Table 4 below.

[0135] [Table 4]

[0136] Animal Models and Efficacy Studies in lung tissue following direct respiratory delivery of clofazimine rather than systemic administration Clinical isolates were analyzed to obtain preliminary data for establishing concentration levels. al) The composition of the present invention increases the proliferation of NTM strains in an in vivo acute lung infection mouse model. Two strains were used to investigate the effect of NTM on lung infection by each species of bacteria of interest. Separate mouse models were used. To carry out the study, Mycobacterium avium (M Mycobacterium avium, )2285 strain and Mycobacterium avium Mycobacterium abscessus strain 103 was used ( The details of the strains are registered in the EMBL / GenBank / DDBJ database (September 2014). "Phylogenetic analysis of Mycobacterium terial species using whole genome sequence ces”.Hazbon MH,Riojas MA,Damon AM, Alalade R., Cantwell B.J., Monaco A., King These two species have already been described in the literature. It has been used as a model for NTM infection (Obregon-Henao et al. l.2015 Antimicrob Agents Chemother;and C han et al.Animal Models of Non-Tuberculo us Mycobacterial Infections,Mycobact Dis 2016).

[0137] In vivo safety study in Balb / C mice: To evaluate the in vivo safety and tolerability, 6- to 8-week-old Balb / C female mice were cultured in The drugs were obtained from Harles River. Mice were allowed to rest for one week before dosing. Three healthy mice were provided for each dose of zimin, and the drugs were administered three times, every other day. Mice were administered 10.0, 5.01, and 2.51 mg / kg of amine in the composition of Example 1. The compound was aerosolized using a microsprayer (registered trademark) in three healthy mice. The vaccine was administered intratracheally every other day for a total of three times using the same drug (Trademark).

[0138] Clofazimine has been shown to be safe at 20 mg / kg (oral gavage, 200 μl). The composition of Example 1 showed no toxicity even at the maximum dose tested (10.0 mg / kg). / kg; 0.2506 mg / dose in 35 μl, intratracheal). Thus, the composition of formula I is 0.0 mg / kg is considered safe and well tolerated.

[0139] Determination of minimum inhibitory concentration Minimum inhibitory concentration (MIC) tests were performed in Mueller-Hinton (MH) broth (cation-adjusted (Calcium and magnesium ion concentrations recommended by CLSI standard M7-A7) ) (Becton Dickinson) and the broth microdilution method The MIC test was performed using 7H9 broth. The compound was also analyzed by the broth microdilution method using Sigma-Aldrich. The rationale for using both MH and 7H9 broths for screening is that antimycobacterial It has been found that various antimicrobial compounds show different MIC activities depending on the broth used for the MIC test. This is because it is used.

[0140] M. abscessus was cultured on 7H11 agar plates (Sigma-Aldrich). in ambient air at 35-37°C (depending on the bacterial strain) on M. avium was grown on 7H11 agar plates ( Sigma-Aldrich) at 37°C in air for 21-30 days.

[0141] Colony forming units (CFU) were harvested from agar plates and incubated with 0.05% Tween-80. The mice were added to either MH broth or 7H9 broth supplemented with 100 μg / mL of M. abscessus (M .abscessus) or after the 12th day (M.avium) The density (OD) was measured by absorbance and was adjusted to an OD of 0.08 to 0.1 (McFarland turbidity standard). The bacteria were grown in air at 35-37°C until they reached a concentration of 0.5. Then, the bacteria were soaked in saline. The suspension is adjusted to an OD of 0.08-0.1 (McFarland turbidity standard solution no. 0.5). The compounds were suspended in DMSO at a concentration of 1.28 mg / ml. Stock solutions of the compounds were prepared by turbidity, with the test range being 64 to 0.062 μg / ml. Then, 180 μl of broth (either MH or 7H9) was added to a 96-well plate. 100 μl of broth was added to the first row of the plate plus the remaining rows of the 96-well plate. 20 μl of the compound stock solution was added to the first row of wells and serially diluted. 100 μl of the solution was added to all wells except the medium-only control wells. QC Reagents: 1) Bacteria only negative control 2) Media only negative control 3) Clarithromycin positive control Sex-drug control.

[0142] The OD of M. abscessus was measured after 3 days, and that of M. avium was measured after 3 days. (M. avium) was measured after 12 days. Following these measurements, Resaz Plates were measured using the urin microtiter assay plate method. Briefly, the method involves the synthesis of resazurin (7-hydroxy-3H-phenoxazine-3- Resazurin is blue in colour. It is a compound that emits weak fluorescence by itself, but when irreversibly reduced, it emits a strong pink-red light. This results in fluorescent resorufin, which determines bacterial cell viability in MIC measurements. It is used as a redox indicator for

[0143] Tests were performed in triplicate. Test #1 was performed after the composition of Example 1 was stored at 4°C for 2 months. Test #1 was conducted 4 months later and Test #3 was conducted 5 months later.

[0144] Minimum inhibitory concentration in the presence and absence of CF sputum Minimum inhibitory concentration determinations were performed as described above.

[0145] Cystic fibrosis (CF) patient sputum was treated with clofazimine (CFZ) and the composition of Example 1. To investigate the effect on bacterial activity, patients who had not received antibiotics within 48 hours were Sputum was collected from the mice and sterilized by exposing it to UV light to remove any endogenous bacteria. After sterilization, M. abscessus, M. avium ), M. intracellulare, and M. chimera (M. Chimaera) was incubated in 10% CF sputum and then subjected to MIC testing. The same CLSI protocol as described above was followed in the presence and absence of sputum from cystic fibrosis patients. The MICs of the composition of Example 1 were determined in the presence and absence of . All tests were performed in duplicate.

[0146] Table 1 shows the MIC values ​​of clofazimine and the composition of Example 1 in the presence and absence of sputum. Shown in 5.

[0147] [Table 5]

[0148] The results shown in Table 5 indicate that both clofazimine and the composition of Example 1 have a broad spectrum of non-tuberculous It can be seen that the compound shows stable MICs against mycobacterial species.

[0149] These data indicate that the composition of Example 1 is effective in preventing and treating M. abscessus It showed potent in vitro activity against both M. avium and M. , suggesting that it is stable for at least this period.

[0150] A mouse model of M. abscessus in SCID mice Six to eight week old SCID female mice were purchased from Charles River. were allowed to rest for 1 week before infection.

[0151] Test solution of M. abscessus strain 103 (working 1 ml of the stock was taken and stored at -80°C until use. The solution was thawed and injected into a 1 ml Luer lock syringe fitted with a 26 g needle for 20 min. The mixture was dissolved in 1× sterile PBS.

[0152] 1 x 10 in acutely infected SCID mouse model 6 CFU / animal (M. abscessus (M. abscessus) Lung infection was induced non-invasively by intratracheal instillation of 103 strains of B. scessus.

[0153] Three mice were sacrificed on day 1 postinfection and bacterial uptake was measured. The whole lung, spleen, and liver were removed and homogenized in 4.5 ml of 1x PBS. The homogenate was serially diluted at a dilution ratio of 1:10, and the dilutions (0-1-2- 3-4-5-6-7) was smeared on a 7H11 agar plate. The plate was then cooled in a dry air incubator at 32°C. The plates were then placed in an incubator (varies depending on the strain) for 7 days.

[0154] The composition of Example 1 was applied at a concentration of 10.0 mg / kg using a Microsprayer®. (35 μl) was administered by the pulmonary route, clofazimine (oral gavage), amikacin (subcutaneous ) was administered in a volume of 200 μl per mouse. These were administered starting on day 2 post-infection, This continued on alternate days for eight days.

[0155] Mice were sacrificed two days after the last dose of compound was administered. (untreated control, clofazimine (administered by oral gavage), the composition of Example 1, and amikacin The treated mice were sacrificed and the bacterial load was measured. -7, spleen 0-1-2-3-4-5-6-7, and liver 0-1-2-3-4-5-6 Plates were smeared at -7.

[0156] The log10 protection value is at least 0. 60 indicates that the activity is statistically significant. FU was transformed into logarithms, which were then evaluated by one-way ANOVA, followed by one-way The analysis was performed using a multiple comparison analysis with a two-way Tukey test (G (Data were analyzed using raphPad Prism analysis software). Differences were considered significant at a confidence level of 95%. .

[0157] Table 6 shows the results of infection of SCID mice with M. abscessus. Later Log 10 The mean and standard error of the mean (SEM) of the CFU data are shown. Total number of animals per group at time of sacrifice.

[0158] [Table 6]

[0159] The data in Table 6 show that treatment with the composition of Example 1 reduced the incidence of M. abscessus (M. abscessus). The amount of bacteria recovered from the lungs and spleens of animals infected with B. scessus was significantly reduced. This reduction in bacterial load was comparable to treatment with amikacin or oral clofazimine. It was statistically improved.

[0160] A mouse model in which beige mice were infected with M. avium Female Beige mice aged 6–8 weeks were purchased from Charles River. The mice were allowed to rest for 1 week before.

[0161] 1 x 10 in acutely infected Beige mouse model 8 Colony forming units (CFU) / ml (M Exposure to M. avium (rough type 2285 strain) via aerosol The lungs were infected non-invasively with M. avium strain 2285. 1 ml of the stock solution for the test was taken, frozen, and stored at -80°C until use. Thaw the aliquot and prepare a 1 ml Luer lock syringe fitted with a 26 g needle. The mixture was then dissolved 20 times using PBS and diluted with sterile 1× phosphate buffered saline.

[0162] Three mice were sacrificed on days 1 and 7 post-infection to measure bacterial uptake. The spleen and liver were removed, homogenized in 4.5 ml of 1x PBS, and diluted 1:10. The dilution solution (0-1-2-3-4-5-6-7) was added to 7H11 / OADC, TSA, and The bacteria are spread on a charcoal agar plate and incubated at 32°C (varies depending on the strain) in dry air. The plates were incubated at 4°C for 30 days.

[0163] The composition of Example 1 was applied at a concentration of 10.0 mg / kg using a Microsprayer®. and clofazimine (oral gavage) was administered via the pulmonary route (35 μl) per mouse. These were administered in a volume of 200 μl starting on day 7 post-infection and continued every other day for 10 days. Ta.

[0164] Mice were sacrificed 5 days after the administration of the final dose of compound. Six mice in every group (untreated control, clofazimine (gavage), and the composition of Example 1) were sacrificed and The amount of 0-1-2-3-4-5-6-7 in the lung homogenate and 0-1-2- 3-4-5-6-7 and liver 0-1-2-3-4-5-6-7 were smeared onto plates.

[0165] A protection value of at least 0.60, expressed as Log10, indicates that activity is statistically significant. Statistical analysis was performed by first converting CFU to logarithms, which were then After evaluation by e-way ANOVA, multiple ratios were evaluated using one-way Tukey test. The results were analyzed by analysis of variance (SigmaStat software program). Differences are considered significant at the 95% confidence level.

[0166] Table 7 shows the Log-10 ratios after infection of Beige mice with M. avium. 10The mean CFU data are shown.

[0167] [Table 7]

[0168] The data in Table 7 show that treatment with the composition of Example 1 reduced the incidence of M. avium m) showed a significant reduction in the amount of bacteria recovered from the lungs and spleens of animals infected with .

[0169] Chronically infected Beige mouse model Six to eight week old Beige mice were allowed to rest for one week before infection. On day 0, M. avium ( M. avium) 2285 rough type 1×10 8 Mice were infected with the CFUs via the lungs. Three mice were sacrificed on the 2nd day and six mice on the 27th day, and the bacterial uptake and bacterial load before treatment were measured. The whole lung, spleen, and liver were removed and homogenized in 4.5 ml of 1x PBS. (0-1-2-3-4-5-6-7) was spread on 7H11 agar plates and charcoal agar plates. The plates were placed in a dry air incubator at 37°C for 25 to 30 days.

[0170] The remaining infected Beige mice were treated every other day starting on day 28 for a total of 14 The animals received one of the following treatments: saline (Microspora gracilis) Rayer®, 35 μl); clofazimine (oral gavage, 20 mg / kg , 200 μl); the composition of Example 1 (IT, Microsprayer®, 1 0 mg / kg, 35 μl).

[0171] Mice were sacrificed on day 57, 2 days after the last treatment. The plates were then placed in a dry air incubator at 37°C. The mixture was placed in an incubator for 30 days.

[0172] Statistical analysis was performed by first transforming CFU into logarithms and then performing a one-way ANOV. After evaluation with A, a one-way Tukey test was used to perform multiple comparison analysis of variance. Differences were considered significant at the 95% confidence level.

[0173] Table 8 shows the L after chronic infection of Beige mice with M. avium. og 10 The mean CFU data are shown.

[0174] [Table 8]

[0175] These data support the finding that in established, "chronic" NTM-infected animal models This suggests that clofazimine is less likely to penetrate into the more established granuloma-like structures. The compositions of the present invention do not have this problem and exert their antimycobacterial activity only after the infection has become well established. It appears that the effect can be maintained even after

[0176] The integrity of the barrier function and the effect of the composition of Example 3 on lung epithelial cells after exposure in vitro Effects on inflammation Cell viability Three different cell types were used to assess cell viability of lung epithelial cells: Calu-3; hAELVi cells were used under two in vitro conditions. (i.e., in cell culture medium on Transwell™ plates) or in apical The cells were treated either in an "air-liquid interface" simulated condition (ALI) with the cell culture medium removed from the side of the plate. In the "immersion condition," Calu-3 cells were immersed in the composition of Example 3 at three doses (10%, The cells were exposed to 0% or 100% CO for 4 h. To assess cell viability, the cells were incubated with acrid Staining was performed using fluorescein orange / propidium iodide (AO / PI) staining to identify live / dead cells. Red fluorescence indicates dead cells.

[0177] Uptake by macrophages THP-1 cells were incubated with 124 ng of phorbol 12-myristate 13-acetate (PMA). The cells were incubated with 100 / ml of erythrocytes for 3 days to differentiate into macrophage-like cells. Once ripe, place in the composition of Example 3 (diluted 1:200 with Hank's Balanced Salt Solution (HBSS)) for 4 h. The cells were stained with AO / PI as described above to confirm the cell Survival rates were determined.

[0178] Transepithelial Electrical Resistance (TEER) Measurement 1 × 10 Calu-3 cells 5 Transwell (commercially available) was used to obtain 100 cells / well. The cells were seeded on a 3460 and grown for 12 days until they reached confluence. World Precision Instruments, Friedberg, Ge TEER measurements were performed using a Calu RT-MS according to the manufacturer's instructions. -3 cells were treated with saline (negative control) or the composition of Example 3 (concentration: 20 mg / ml, 10 After 2–4 h of exposure, cells were exposed to either 0.1 mg / ml or 0.2 mg / ml of erythrocytes. TEER was measured.

[0179] Inflammatory cytokine production Differentiated THP-1 cells (dTHP-1) were exposed to the composition of Example 3 for 4 hours or 24 hours. Exposure to HBSS alone was used as a negative control. As a positive control, lipopolysaccharide (LPS) (100 ng / ml) was administered.

[0180] After incubation for t=4 or 24 hours, the supernatant was removed from the cells and plated. Pooled supernatant samples were subjected to enzyme-linked immunosorbent assay (ELISA). Separate E. coli antibodies were used for TNF-α, IL-6, IL-8, and IL-10 according to the manufacturer's instructions. The LISA kit was used.

[0181] One-way analysis of variance (ANOVA) was performed, followed by Tukey post-hoc te Statistical analysis was performed using the st method. Probability values ​​<0.05 were considered statistically significant. Ta.

[0182] result After 4 hours of incubation under "immersion" conditions, the composition of Example 3 still maintained the same None of the concentrations visibly reduced cell viability.

[0183] Under “ALI” conditions, three different cell lines (Calu-3, A549, and HA ELVi cells) were examined three times at different time points (after 5 hours, 2 days, and 7 days). There was little or no cytotoxicity in any of the cells after 2 days in the case of Calu-3 cells. In A549 cells, the Calu-3 cells were not affected after 2 and 7 days. Some degree of toxicity was observed after 7 days in the control group. Due to technical limitations, quantification of dead cells was not possible. There wasn't.

[0184] For macrophage uptake, differentiated THP-1 cells were incubated at 1:200 HB The cells were incubated with SS for 4 hours, and the cell viability of the macrophages after exposure was determined. The composition of Example 3 did not induce cell death, but clofazimine was taken up by macrophages. It was clearly shown that

[0185] For TEER measurement, Calu-3 cells were cultured in HBSS or the mixture of Example 3 at three different concentrations. The animals were exposed to the composition for 4 hours and TEER measurements were taken at various time points during exposure. At this point, a decrease in TEER of ≥ 50% compared to the control indicates a significant decrease in barrier function integrity. It was deemed that.

[0186] When Calu-3 cells were exposed to the composition of Example 3, the barrier function was improved even after 1 hour of exposure. No effect was observed on the health of the rats. Exposure to 20 mg / ml significantly increased the At a concentration of 10 mg / ml, there was a slight decrease at any time point after 2 hours. A significant decrease (i.e., 25-35%) was observed in the 2 mg / ml dose group throughout the study period. No reduction in barrier function was observed.

[0187] Inflammatory cytokine production The positive control, LPS, behaved as expected in this model. Composition of Example 3 No significant changes in cytokines were observed at any of the time points examined.

[0188] The results are shown in Table 9.

[0189] [Table 9]

[0190] In vivo safety and tolerability The mice were administered the drug every other day for a total of three times to 6-8 week-old Balb / C female mice. The composition of Example 1 was administered at 10.0, 5.01, and 2.51 mg / kg. Aerosol was administered intratracheally at a volume of 35 μl per animal using a Microsprayer®. After injection, mice were observed at 10 min, 1, 2, and 4 h after injection. Thereafter, observations were made daily.

[0191] Table 10 shows the overall observations after administration. "BAR" indicates that the animals were bright and ), active and responsive. represent.

[0192] [Table 10] [Table 11]

[0193] These data indicate that the body weight did not show a statistically significant change over the 3 days of treatment. is doing.

[0194] These results indicate that the compositions of the present invention are well tolerated at the doses tested. is doing.

Claims

1. a) a therapeutically effective dose of clofazimine or a pharma- ceutical acceptable salt thereof; b) a nonionic surfactant having a hydrophilic-lipophilic balance value greater than 10; c) an aqueous liquid carrier selected from at least one of water, isotonic saline, buffered saline, and aqueous electrolyte solutions; d) a mucolytic and / or biofilm disrupting agent; A pharmaceutical composition comprising: The clofazimine or pharma- ceutically acceptable salt thereof is provided in the form of particles in a suspension; and A pharmaceutical composition for administration by inhalation, wherein said particles of said clofazimine or a pharma- ceutically acceptable salt thereof have a median diameter of less than 5 μm and a D90 of less than 6 μm.

2. 2. The pharmaceutical composition of claim 1, wherein the particles of clofazimine or a pharma- ceutically acceptable salt thereof have a mean diameter of less than 2 μm and a D90 of less than 3 μm.

3. 2. The pharmaceutical composition of claim 1, wherein the mucolytic and / or biofilm disrupting agent is selected from the group consisting of at least one of nebulized 4-7% hypertonic saline, metaperiodate, sodium dodecyl sulfate, sodium bicarbonate, tromethamine, silver nanoparticles, bismuth-thiol, ethylenediaminetetraacetic acid, gentamicin-loaded phosphatidylcholine modified gold nanoparticles, chelating agents, cis-2-decenoic acid, D-amino acids, peptides containing D-amino acid residues, gallium mesoporphyrin IX, gallium protoporphyrin IX, curcumin, patulin, penicillic acid, baicalein, naringenin, ursolic acid, asiatic acid, corosolic acid, fatty acids, host defense peptides, and antimicrobial peptides.

4. 2. The pharmaceutical composition of claim 1, wherein the mucolytic and / or biofilm disrupting agent is an active agent selected from bedaquiline or a pharma- ceutically acceptable salt thereof, cefoxitin, amikacin, clarithromycin, pyrazinamide, rifampin, moxifloxacin, levofloxacin, and para-aminosalicylate, and mixtures thereof.

5. The nonionic surfactants include polysorbate 20, polysorbate 60, polysorbate 80, stearyl alcohol, polyethylene glycol derivatives of hydrogenated castor oil having a hydrophilic-lipophilic balance value of 14 to 16, polyethylene glycol derivatives of hydrogenated castor oil having a hydrophilic-lipophilic balance value of 15 to 17, sorbitan monolaurate, sorbitan monopalmitate, sorbitan monostearate, polyoxyethylene (20) oleyl ether, polyoxyethylene (20) cetyl ether, polyoxyethylene (10) cetyl ether, polyoxyethylene (10) oleyl ether, polyoxyethylene (100 2. The pharmaceutical composition of claim 1, wherein the polyoxyethylene (4) lauryl ether, polyoxyethylene (20) cetyl ether, polyoxyethylene (2) cetyl ether, caprylocaproyl polyoxyl-8 glyceride, polyethylene glycol (20) monostearate, polyethylene glycol (40) stearate, polyethylene glycol (100) stearate, polyethylene glycol (8) stearate, and polyoxyl 40 stearate, and mixtures thereof.

6. The non-ionic surfactant is polysorbate 80, 2. The pharmaceutical composition of claim 1, wherein the aqueous liquid carrier is distilled water, hypertonic saline, or isotonic saline.

7. The pharmaceutical composition of claim 6, wherein the hypertonic saline is 1% to 7% (w / v) sodium chloride.

8. The non-ionic surfactant is ultra-pure polysorbate 80; 7. The pharmaceutical composition of claim 6, wherein the aqueous liquid carrier is an isotonic saline solution.

9. 2. The pharmaceutical composition of claim 1, wherein the osmolality of the composition is in the range of 200-700 mOsm / kg.

10. 2. The pharmaceutical composition of claim 1, wherein the osmolality of the composition is in the range of 300-400 mOsm / kg.

11. The non-ionic surfactant is present in the range of 0.001% to 5% (v / v) of the total composition; and 2. The pharmaceutical composition of claim 1, wherein the amount of clofazimine ranges from 0.1% to 20% (w / v) of the total composition.

12. A pharmaceutical composition in the form of an inhalable aerosol, comprising:

10. The pharmaceutical composition of claim 1, wherein the pharmaceutical composition is prepared by aerosolizing the pharmaceutical composition with a nebulizing device selected from an ultrasonic nebulizer, an electrostatically charged spray nebulizer, a vibrating membrane nebulizer, a jet nebulizer, and a mechanical soft mist metered dose inhaler; A pharmaceutical composition, wherein the aerosol particles produced by said nebulizer device have a mass median aerodynamic diameter of 1 to 5 μm.

13. 1. A system for use in providing antibacterial activity in treating or preventing pulmonary infections caused by mycobacteria or other gram-positive bacteria, the system comprising: １） a therapeutically effective dose of clofazimine; a nonionic surfactant having a hydrophilic-lipophilic balance value of greater than 10; an aqueous liquid carrier selected from water, isotonic saline, buffered saline, and aqueous electrolyte solutions; a mucolytic and / or biofilm disrupting agent; A pharmaceutical composition for administration by inhalation comprising: 2) a nebulizer for atomizing the pharmaceutical composition for administration by inhalation; Including, said clofazimine being present in the form of a suspension; The aerosol particles produced by the system have a mass median aerodynamic diameter of 1 to 5 μm; and The treatment or prevention comprises inhaling the pharmaceutical composition sprayed by the nebulizer. system.

14. The pharmaceutical composition of claim 1, 20. A pharmaceutical composition for use in a method for the treatment or prevention of a pulmonary infection in a patient in need thereof comprising administering said pharmaceutical composition by inhalation.

15. The pharmaceutical composition according to claim 14, wherein the infectious disease is caused by a species belonging to the genus Mycobacterium selected from the group consisting of nontuberculous mycobacteria and Mycobacterium tuberculosis and combinations thereof.

16. 16. The pharmaceutical composition of claim 15, wherein the nontuberculous mycobacteria is selected from Mycobacterium avium, Mycobacterium intracellulare, Mycobacterium abscessus, and Mycobacterium leprae, and combinations thereof.

17. 16. The pharmaceutical composition of claim 15, wherein the infection is an opportunistic infection selected from pulmonary MAC disease and nontuberculous mycobacterial disease in patients with cystic fibrosis, chronic obstructive pulmonary disease, or acquired immune deficiency syndrome.

18. 18. The pharmaceutical composition of claim 17, wherein the infection is an opportunistic nontuberculous mycobacterial infection in a cystic fibrosis patient.

19. The pharmaceutical composition of claim 1, 1. A pharmaceutical composition for use in a method for the treatment or prevention of pulmonary infections caused by mycobacteria or other gram-positive bacteria in a patient in need thereof comprising administering said pharmaceutical composition by inhalation before, simultaneously with, or after administration of an agent selected from bedaquiline or a pharma- ceutical acceptable salt thereof, cefoxitin, amikacin, clarithromycin, pyrazinamide, rifampin, moxifloxacin, levofloxacin, and para-aminosalicylate, and mixtures thereof.

20. 20. The pharmaceutical composition of claim 19, wherein the agent is bedaquiline or amikacin.

21. 21. The pharmaceutical composition of claim 20, wherein the agent is bedaquiline.

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