JP2013518895A - Bismuth-thiol as a disinfectant for biomedical use, including treatment of bacterial biofilms and other uses - Google Patents

Abstract

Formulations, including disinfecting formulations, for treating natural surfaces containing bacterial biofilms, including unexpected synergistic or enhancing effects between bismuth-thiol (BT) compounds and certain antibiotics A composition and method comprising a novel homogeneous microparticle suspension is described. Disclosed, including the appropriate efficacy of certain such compositions to treat certain gram-positive bacterial infections, and the different appropriate efficacy of certain such compositions to treat certain gram-negative bacterial infections The antibacterial and anti-biofilm properties of the BT compound and BT compound + antibiotic combination, which were previously unforeseen, are also described.
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Description

CROSS REFERENCE TO RELATED APPLICATIONS This application claims the benefit of PCT application PCT / US2010 / 023108 filed February 3, 2010, and US provisional application 61 / 373,188 filed August 12, 2010. Each of which is incorporated herein by reference in its entirety.

BACKGROUND OF THE INVENTION Embodiments of the invention disclosed herein relate to compositions and methods for the treatment of microbial infections. In particular, embodiments of the invention include in the treatment of bacterial biofilms and other conditions, in wounds such as chronic and acute wounds, and in epithelial tissues in clinical personal health care and other aspects. It relates to improved treatments for managing bacterial infections.

Description of Related Art A complex series of coordinated intramolecular and intermolecular interactions that contribute to skin wound healing and response to and resistance to microbial infections and / or healing or retention of body tissues are generally e.g. opportunistic infections and nosocomial Infection (eg, clinical regimen that may increase the risk of infection), local or systemic administration of antibiotics (may affect cell growth, migration or other functions, and may be selected for antibiotic-resistant microorganisms Possible), frequent wound dressing exchange, exposure to the outside air of the wound to accelerate healing, use of temporary artificial structural support matrix or scaffold material, removal of necrotic tissue and / or repeated surgery to remove infected or necrotic tissue Where it is adversely affected by a variety of external factors, such as the potential need for and / or other factors There is a match.

  Wound healing is thus a long-standing challenge for clinical personnel around the world. Current treatments for refractory wounds are impractical and ineffective and often require multiple surgeries to close the wound. For example, Regranex® (becaprelmine, available from Ortho-McNeil Pharmaceutical, Inc., Ethicon, Inc., recombinant platelet-derived growth factor) is one of the few available treatments for chronic wounds. It is expensive to manufacture and has limited clinical utility.

Chronic and acute wounds and wound biofilms Continuity between cells within a tissue, or continuity between tissues, is, for example, physical, mechanical, biological, pathological and / or chemical force (eg, burn, Dermal infection, stab wound, wound with handgun or shrapnel, skin ulcer, radiotoxicity, malignant tumor, gangrene, autoimmune disease, immune deficiency disease, respiratory injury due to inhalation or infection, ingestion or infection of dangerous goods, etc. Wounds occur when disrupted by gastrointestinal injury, circulatory and hematopoietic disorders such as inability to clot), or other traumatic injuries.

A slight level of bacterial contamination in the wound, or “colonization” of the wound, does not necessarily interfere with the process of wound healing, but if there is a sufficient number of bacteria to overwhelm the host immune defense, acute wounds or It can result in chronic wounds or wounds where bacterial biofilms are present, such as wound infections in which host damage proceeds with bacterial growth. Bryant and Nix, Acute and Chronic Wounds :;: (. 2 nd Ed) Current Management Concepts, 2006 Mosby (Elsevier), NY Baronski, Wound Care Essentials Practical Principles, 2007 Lippincott, Williams and Wilkins, Philadelphia, PA). For example, acute wounds that can arise due to injury, trauma, surgical intervention, or other causes typically heal rapidly without underlying health defects, but in some cases infection is present In some cases, the rapid formation of bacterial biofilms has been described in acute wounds (eg WO / 2007/061942). Additional factors that can cause chronic wound development include lack of mobility (eg, it applies continuous pressure to the wound site), loss of sensation or mental ability, inaccessibility of the wound site (eg, Respiratory or gastrointestinal tract), and cardiovascular defects. Infection at a chronic wound site may be detected by skin redness, edema, pus formation and / or unpleasant odor, or other relevant clinically accepted criteria.

  Thus, when the host immune system is overwhelmed by bacterial infection at the wound site (eg, acute wound) and an acceptable condition arises for the bacteria to enter and further destroy the tissue (non-limiting) In humans and other mammals), there may be acute wounds that cannot be healed properly, and thus chronic damage may occur. In general, chronic wounds are wounds that do not heal within 3 months and tend to grow larger as bacterial invasion progresses instead of the wound becoming smaller. Chronic wounds may increase patient pain and stress when nearby nerves become damaged (neuropathy) as the wound progresses. These wounds affect 4 million Americans each year and cost approximately $ 9 billion for treatment. Most affected individuals are over 60 years old.

  Chronic wounds can sometimes develop as acute wounds, i.e. wounds with handguns or grenades, burns, stab wounds, venous ulcers, pressure ulcers, diabetic ulcers, radioactive poisoning, malignant tumors, dermal infections , Gangrene, surgical wound, diabetic leg ulcer, pressure ulcer, venous leg ulcer, intractable surgical wound containing infection and / or biofilm, gangrene pyoderma, traumatic wound, acute arterial failure, necrotizing fasciitis, Mention may be made of osteomyelitis (bone infection), radioactive injuries such as radiation osteonecrosis and soft tissue radiation necrosis, or other types of wounds. For example, venous ulcers most often occur in the lower limbs due to restricted circulation (eg, ischemia), venous valve dysfunction, or repeated physical trauma (eg, repeated injury). For example, if the local pressure applied at or near the wound site is greater than the blood pressure, pressure ulcers may result, resulting in circulatory inhibition, paralysis, and / or bed slipping that causes chronic injury Or make it worse. Diabetic ulcers can occur in diabetic patients, for example, loss of limb sensation due to uncontrollable hyperglycemia, repetitive damage to the patient's body, and / or failure to notice the damage. Factors that complicate or otherwise affect the clinical signs and outcomes of chronic wounds include the patient's immune status (eg, immunosuppression, pathological (eg, HIV-AIDS), radiation therapy, or Immune system damaged by drugs; age; stress); aging of the skin (such as photochemical aging), and the development and progression of biofilms within the wound. In the case of epithelial tissue in the respiratory and / or gastrointestinal tract, inaccessibility, obstruction, difficulty in generating fluid forces to wash the epithelial surface, or creation of a local microenvironment that induces microbial survival, May cause clinical problems.

  Wound-related damage may involve loss or impairment of organ function, shock, bleeding and / or thrombus, cell death (eg, necrosis and / or apoptosis), stress and / or microbial infection. Any or all of these events, particularly infection, can delay or interfere with an effective tissue repair process involved in wound healing. For this reason, in individuals with persistent wounds, the removal of non-viable tissue from the wound site (a process called necrotic tissue removal) and the removal of any foreign material from the wound site (also known as wound cleansing) is as early as possible. Called) may be important.

  Severe wounds, acute wounds, chronic wounds, burns, and ulcers can benefit from cell wound dressings. A number of artificial skin products, for example, Aplygraft (R) (Norvartis), Demograft (R), Bioblane (R), Transcyt (R) (Advanced Tissue Science), Integra (R) Demolregulation (R) ) (Integra Life Sciences Technology), and OrCel® can be used for refractory or burn wounds. However, these products are not designed to address the issues of bacterial tissue invasion and wound enlargement.

  Unfortunately, systemic antibiotics are not effective in treating chronic wounds and are generally not used unless an acute bacterial infection is present. Current approaches include the administration or application of antibiotics, but such remedies may facilitate the emergence of antibiotic resistant strains and / or may be ineffective for bacterial biofilms . Therefore, it may be particularly important to use a disinfectant when drug resistant bacteria (eg, methicillin resistant Staphylococcus aureus or MRSA) are detected. Although there are many widely used antiseptics, established bacterial populations or subpopulations may not respond to these agents, or any other currently available treatment. In addition, disinfectants can be toxic to host cells at concentrations required to be effective against established bacterial infections, and as such, such disinfectants are inappropriate. This problem can occur in body epithelial surfaces such as respiratory tracts (eg, airways, nasopharynx and larynx, trachea, lungs, bronchi, bronchial tops, alveoli, etc.) or gastrointestinal tracts (eg, oral cavity, esophagus, stomach, intestines, It can be particularly serious in the case of attempts to cleanse infections from natural surfaces including rectal, anal, etc.) or other epithelial surfaces.

  Of particular concern are infectious organisms composed of bacterial biofilms, a relatively recently recognized bacterial tissue, which is caused by suspension unicellular (plankton) bacteria, which have markedly different behavioral patterns and gene expression due to cell-cell adhesion. And organized multicellular communities (biofilms) that are sensitive to environmental drugs such as antibiotics. Biofilms may deploy a biological defense mechanism not found in planktonic bacteria, which can protect the biofilm community from antibiotic and host immune responses. The established biofilm can stop the tissue healing process.

  Common microbiological contaminants lurking in persistent and potentially toxic infections include S. cerevisiae including MRSA (methicillin resistant Staphylococcus aureus). Aureus, Enterococcus, E. Coli, P.A. Examples include aeruginosa, Streptococcus, and Acinetobacter baumannii. Some of these organisms show the ability to survive on non-nutritive clinical surfaces for months. S. Aureus has been shown to be viable on dry glass for 4 weeks and on dry blood and cotton fibers for 3-6 months (Domenico et al., 1999 Infect. Immun. 67: 664-669). . E. Coli and P. Both aeruginosa are S. cerevisiae on dry blood and cotton fibers. It was shown to survive longer than Aureus (previous work).

  Microbial biofilms are associated with a substantial increase in resistance to both fungicides and antibiotics. Biofilm morphology occurs when bacteria and / or fungi attach to the surface. This attachment causes changes in gene transcription, resulting in secretions that are remarkably active and difficult to penetrate the polysaccharide matrix, and protect the microorganisms. In addition to having very substantial resistance to antibiotics, biofilms are very resistant to the mammal's immune system. Since biofilms become very difficult to eradicate once established, preventing the formation of biofilms is a very important clinical priority. Recent studies have shown that open wounds can be rapidly contaminated by biofilm. These microbial biofilms are thought to delay wound healing and are likely to be associated with the establishment of serious wound infections.

  For example, current military wound care guidelines specify vigorous and complete irrigation and necrotic tissue removal (Blankenship CL, Guidelines for open of comba calsound wounds and Fleet Operations and. (Bureau of Medicine and Surgery). This initial intervention is important but not sufficient to prevent the onset of infection. Additional treatment steps are required to undergo subsequent debridement to promote healing and reduce microbial bioburden, thereby reducing the chance of wound infection and wound biofilm colonization.

  Due to the complex nature of military trauma wounds, the potential for infection is high and the introduction of foreign objects and other environmental pollutants is particularly considered. Both military and clinical environments, including humans present in both of these environments, are particularly important for potentially pathogenic microorganisms, especially those that have suffered from open and / or complex wounds Act as a good source. Acute and chronic wounds, including surgical wounds and military wounds, already hurt the skin, which is the body's primary defense and barrier to infection. The wound thus exposes the body's interior (wet and nutritive environment) to opportunistic and pathogenic infections. Many of these infections, particularly persistent wound infections, can be involved in biofilm formation, as shown in the case of chronic wounds (James et al., 2008). Wound infections in hospital facilities are one of the most common causes of nosocomial infections, and wounds obtained in military and natural disaster environments are particularly susceptible to microbial contamination. Military wounds are typically associated with tissue damage, tend to be extensive and deep, may introduce foreign bodies and interfere with local blood supply, may be related to fractures and burns, shocks Military wounds are susceptible to infections because they can cause damage to immune defenses.

Skin structure and wound healing Skin and other epithelial tissues that are intact and functioning (eg, generally found in avascular epithelial surfaces that form a barrier between an organism and its external environment, such as skin, breathing Maintenance of organs and gastrointestinal tracts, as well as those found in the inner layers of glandular tissues, etc. are critical to the health and survival of humans and other animals. The skin is the largest body organ of humans and other higher vertebrates (eg, mammals) and protects against environmental injury through barrier function, mechanical strength and water impermeability. As a critical environmental interface, the skin provides a protective body covering that allows maintenance of physiological balance.

  The structure of the skin is well known. Briefly, the epidermis, or outer layer of the skin, is covered with a stratum corneum, a protective layer of inactive epidermal skin cells (eg, keratinocytes) and extracellular connective tissue proteins. The epidermis undergoes a continuous process of molting when exchanged by new material pushed up from the underlying epidermal granule cells, spinous cells, and basal cell layers, and through continuous cell division and protein synthesis new skin cells and skin proteins ( For example, keratin, collagen) is produced. The dermis resides beneath the epidermis and assimilates connective tissue proteins (eg, collagen, elastin, etc.) with dermal fibroblasts and assembles the extracellular matrix and fibrous structure to give the skin flexibility, strength and elasticity. It is a site to be given. Within the dermis are also nerves, blood vessels, smooth muscle cells, hair follicles and fatty glands.

  As the body's primary line of defense, the skin is an important target for clinical injuries such as physical, mechanical, chemical and biological (eg, xenobiotic, autoimmune) attacks that can alter structure and function is there. The skin is also considered an important component of the organism's immune defenses. In the skin, migratory and resident leukocytes (eg, lymphocytes, macrophages, mast cells) and epidermal dendritic (Langerhans) cells that have potent antigen-presenting activity and contribute to immunological defense can be found. Colored melanocytes in the basal layer absorb potentially harmful ultraviolet (UV) radiation. Skin destruction poses inappropriate risks to subjects, such as opportunistic infections, incomplete and inappropriate tissue remodeling, scarring, mobility impairment, risks associated with pain and / or other disorders. Like skin, other epithelial surfaces (eg, respiratory tract, gastrointestinal tract and gland lining) have defined structural attributes in normal times, so infection or other destruction poses a significant health risk There is.

  For example, wounds such as cuts, abrasions, abrasions, stabs, burns (including chemical burns), infections, extreme temperatures, incisions (eg, surgical incisions), trauma and other injuries can damage or destroy skin There is a case. Therefore, efficient skin repair by wound healing is clearly desirable in these and similar aspects.

  Skin naturally exhibits significant self-healing ability after many types of damage, but skin healing does not occur quickly enough and / or is incompletely remodeled by improper cellular tissue repair mechanisms There are still many aspects in which damaged skin can result and lack integrity, barrier properties, mechanical strength, elasticity, flexibility, or other properties that are desirable for intact skin. Thus, skin wound healing presents such associated challenges, for example, in the aspect of chronic wounds.

  Wound healing occurs at three dynamic and overlapping periods and begins with the formation of fibrin clots. The fibrin clot provides a temporary shield and reservoir of growth factors to attract cells into the wound. It also acts as an interim extracellular matrix (ECM) into which cells enter during repair. Overlapping with fibrin clot formation is the inflammatory phase, which infiltrates the wound with phagocytes and neutrophils that release growth factors that amplify the initial healing response while cleaning wound debris and bacteria It is characterized by doing. The process of regenerating the skinned area is initiated in the proliferative phase of healing and is manipulated by chemokines, cytokines and proteases secreted from immune cells, which are concentrated within the fibrin clot. The keratinocytes are stimulated to proliferate and migrate to form a new epithelial layer covering the wound, while wound angiogenesis delivers oxygen, nutrients, and inflammatory cells to the wound area. The remodeling phase is the final phase of wound repair, performed by myofibroblasts, which promote connective tissue contraction, increase wound strength and attach ECM to form scars (Martin, P. Wound Healing-Aiming for Perfect Skin Generation. Science 1997; 4: 75-80).

Disinfectants based on bismuth thiol (BT) Anti-microbial, in particular, multiple natural products (eg, antibiotics) and synthetic chemicals that have antibacterial properties are known in the art and are Antimicrobial effects, eg ability to kill microorganisms ("killing" effects such as bactericidal), ability to stop or damage the growth of microorganisms ("static" effects such as bacteriostatic), or microbial function, eg at the site At least in part by the ability to interfere with colony formation or infection of bacteria, secretion of exopolysaccharides by bacteria, and / or the conversion of planktonic populations to biofilm populations or the expansion of biofilm formation. Antibiotics, bactericides, disinfectants, etc. (including bismuth-thiol), including factors that affect the selection and use of such compositions, such as bactericidal or bacteriostatic capacity, effective concentration, and risk of toxicity to host tissues Or a BT compound) includes, for example, U.S. Pat. S. Discussed in US Pat. No. 6,582,719.

  Bismuth is a Group V metal and is an antimicrobial element (similar to silver). Bismuth alone may not be therapeutically useful and may exhibit certain inappropriate properties, so it may instead be administered by delivery with complexing agents, carriers, and / or other vehicles, The most common example is Pepto Bismol® in which bismuth is combined (chelated) with hyposalicylate. Combining certain thiol (-SH, sulfhydryl) -containing compounds, such as ethanedithiol and bismuth, to provide exemplary bismuth thiol (BT) compounds, compared to other currently available bismuth preparations, It has been determined in previous studies that antimicrobial activity is improved. There are many thiol compounds that can be used to produce BT (eg, Domenico et al., 2001 Antimicrob. Agent. Chemotherap. 45 (5): 1417-1421, Domenico et al., 1997 Antimicrob. 41 (8): 1697-1703, and U.S. RE 37,793, U.S. 6,248,371, 6,086,921, and 6,380,248. And see, for example, US 6,582,719), several of these preparations can inhibit biofilm formation.

  BT compounds are MRSA (methicillin resistant S. aureus), MRSE (methicillin resistant S. epidermidis), mycobacterium tuberculosis, mycobacterium abium, drug resistant P. cerevisiae. Aeruginosa, enterotoxigenic E. coli E. coli, enterohemorrhagic E. coli Cori, Klebsiella pneumoniae, Clostridium difficile, Helicobacter pylori, Legionella pneumophila, Enterococcus faecalis, Enterobacter cloacae, Salmonella typhimurium, Proteus bulgaris, Yersinia enterocolitica, Vibrio cholera, and Activity against Shigella flexneri has been demonstrated (Domenico et al., 1997 Antimicrob. Agents Chemother. 41: 1697-1703). There is also evidence of activity against cytomegalovirus, herpes simplex virus type 1 (HSV-1) and HSV-2, and yeast and fungi such as Candida albicans. The role of BT is to reduce bacterial virulence, inhibit or kill broad-spectrum antibiotic-resistant microorganisms (Gram-positive and Gram-negative), prevent biofilm formation, and prevent septic shock Have also been demonstrated in treating sepsis and increasing bacterial susceptibility to previously resistant antibiotics (eg, Domenico et al., 2001 Agent. Chemother. 45: 1417-1421; Domenico et al. , 2000 Infect.Med 17: 123-127 ;, Antimicrob Agents & Chemother.3: 79-85, Domenico et al., 2003 Res.Adv .; Microb. Agent. Chemother. 41 (8): 1697-1703; Domenico et al., 1999 Infect.Immun.67: 664-669; Huang et al.1999 J Antimicrob.Chemother. , 2003 J Antimicrob. Chemother. 52: 915-919; Wu et al., 2002 Am J Respir Cell Mol Biol.

  Despite the availability of BT compounds for well over a period of 10 years, the effective selection of appropriate BT compounds for indications of specific infectious diseases remains an elusive goal, and is specific to specific microorganisms. The behavior of BT cannot be predicted, the synergistic activity of a specific BT and a specific antibiotic against a specific microorganism cannot be predicted, and the in vitro BT effect can usually be predicted from the in vitro BT effect. First, the BT effect on a planktonic (single cell) microbial population cannot be a predictor of the BT effect on a microbial community, for example, a bacterium organized into a biofilm. In addition, limitations such as solubility, tissue permeability, bioavailability, and biodistribution may interfere with the ability to deliver clinical benefits safely and effectively in the case of some BT compounds. There is. The embodiments of the invention disclosed herein address these needs and present other related benefits.

BRIEF SUMMARY Although initially disclosed herein and not wishing to be bound by theory, according to certain embodiments described herein, a bismuth-thiol (BT) compound is: It can be used as a disinfectant used in a wide variety of clinical infectious diseases and conditions, as well as in personal health care, such as savings carried out by prevention or protection mediated at least in part by BT It also reduces the cost of treating the infection.

  Also in certain embodiments, a bacterial biofilm or a bacterium associated with biofilm formation (eg, forming or forming a biofilm) comprising one or more BT compounds and one or more antibiotic compounds described herein. Formulations for treating tissues and / or surfaces containing bacteria that can be promoted in other ways are contemplated, and according to non-limiting theory, BT compounds appropriately selected based on this disclosure and Combinations with antibiotics are synergistic previously unforeseen in the antimicrobial (including anti-biofilm) effects of such formulations, and / or prevention, protection and prevention of microbial infections, including infections involving bacterial biofilms Providing an unexpected enhancement effect for a therapeutically effective treatment.

  Also provided for the first time herein is an unprecedented bismuth-thiol composition comprising a substantially monodispersed particulate suspension and methods of synthesizing and using it.

In accordance with certain embodiments of the invention described herein, there is thus provided a bismuth-thiol composition comprising a plurality of microparticles comprising a bismuth-thiol (BT) compound, wherein substantially all of the microparticles are The BT compound comprises a bismuth or bismuth salt and a thiol-containing compound. In another embodiment, a bismuth-thiol composition comprising a plurality of microparticles comprising a bismuth-thiol (BT) compound is provided, wherein substantially all of the microparticles have a volume average diameter of about 0.4 μm to about 5 μm. Having (a) a bismuth salt containing bismuth at a concentration of at least 50 mM, under conditions and time sufficient to obtain a solution substantially free of solid precipitates, hydrophilic, polar or organic An acidic aqueous solution free of soluble solubilizer is sufficient to obtain an admixture comprising (ii) at least about 5%, 10%, 15%, 20%, 25% or 30% ethanol by volume (B) at a molar ratio of about 1: 3 to about 3: 1 with respect to bismuth, under conditions and time sufficient to form a precipitate containing fine particles containing the BT compound. The ethanolic solution containing the thiol-containing compounds present in 応溶 solution was added to the blend of (a), to obtain a reaction solution, it is formed by a method comprising. In certain embodiments, the bismuth salt is Bi (NO ₃ ) ₃ . In certain embodiments, the acidic aqueous solution comprises at least 5 wt%, 10 wt%, 15 wt%, 20 wt%, 22 wt% or 22.5 wt% bismuth. In certain embodiments, the acidic aqueous solution is at least 0.5 wt%, 1 wt%, 1.5 wt%, 2 wt%, 2.5 wt%, 3 wt%. Contains 3.5 wt%, 4 wt%, 4.5 wt% or 5 wt% nitric acid. In certain embodiments, the thiol-containing compound is 1,2-ethanedithiol, 2,3-dimercaptopropanol, pyrithione, dithioerythritol, 3,4-dimercaptotoluene, 2,3-butanedithiol, 1,3- One or more agents selected from propanedithiol, 2-hydroxypropanethiol, 1-mercapto-2-propanol, α-lipoic acid, and dithiothreitol are included.

In another embodiment, a method of preparing a bismuth-thiol composition comprising a plurality of microparticles comprising a bismuth-thiol (BT) compound is provided, wherein substantially all of the microparticles are from about 0.4 μm to about 5 μm. A bismuth salt having a volume average diameter, wherein the method comprises (a) conditions and time sufficient to obtain a solution substantially free of solid precipitates, and (i) a bismuth salt having a concentration of at least 50 mM. An acidic aqueous solution containing no hydrophilic, polar or organic solubilizers, (ii) at least about 5% by volume, 10% by volume, 15% by volume, 20% by volume, 25% by volume or 30% by volume ethanol. Admixing with a sufficient amount of ethanol to obtain an admixture comprising; and (b) bismuth for conditions and time sufficient to form a precipitate comprising microparticles comprising a BT compound. About 1: 3 to about 3: ethanolic solution containing the thiol-containing compound present in the reaction solution in a molar ratio was added to the admixture of (a), comprising the step of obtaining a reaction solution. In certain embodiments, the method further includes recovering the precipitate to remove impurities. In certain embodiments, the bismuth salt is Bi (NO ₃ ) ₃ . In certain embodiments, the acidic aqueous solution comprises at least 5 wt%, 10 wt%, 15 wt%, 20 wt%, 22 wt% or 22.5 wt% bismuth. In certain embodiments, the acidic aqueous solution is at least 0.5 wt%, 1 wt%, 1.5 wt%, 2 wt%, 2.5 wt%, 3 wt%. Contains 3.5 wt%, 4 wt%, 4.5 wt% or 5 wt% nitric acid. In certain embodiments, the thiol-containing compound is 1,2-ethanedithiol, 2,3-dimercaptopropanol, pyrithione, dithioerythritol, 3,4-dimercaptotoluene, 2,3-butanedithiol, 1,3- Propane dithiol, 2-hydroxypropanethiol, 1-mercapto-2-propanol, dithiothreitol, α-lipoic acid, methanethiol (CH ₃ SH [m-mercaptan]), ethanethiol (C ₂ H ₅ SH [e- mercaptans]), 1- propanethiol _{(C 3 H 7 SH [n} -P mercaptans]), 2- propanethiol _{(CH 3 CH (SH) CH} 3 [2C 3 mercaptan]), butanethiol _{(C 4} H 9 SH [N-butyl mercaptan]), tert-butyl mercaptan (C (CH ₃ ) ₃ SH [t-butyl mercaptan]), pentanethiol (C ₅ H ₁₁ SH [pentyl mercaptan]), coenzyme A, lipoamide, glutathione, cysteine, cystine, 2-mercaptoethanol, dithiothreitol, 2-mercaptoindole, Transglutaminase, (11-mercaptoundecyl) hexa (ethylene glycol), (11-mercaptoundecyl) tetra (ethylene glycol), (11-mercaptoundecyl) tetra (ethylene glycol) functionalized gold nanoparticles, 1, 1 ′, 4 ′, 1 ″ -terphenyl-4-thiol, 1,11-undecanedithiol, 1,16-hexadecanedithiol, technical grade 1,2-ethanedithiol, 1,3-propanedithiol, 1,4-benzenedimethane All, 1,4-butanedithiol, 1,4-butanedithiol diacetate, 1,5-pentanedithiol, 1,6-hexanedithiol, 1,8-octanedithiol, 1,9-nonanedithiol, adamantanethiol, 1-butanethiol, 1-decanethiol, 1-dodecanethiol, 1-heptanethiol, 1-heptanethiolpurum, 1-hexadecanethiol, 1-hexanethiol, 1-mercapto (triethylene glycol), 1- Mercapto (triethylene glycol) methyl ether functionalized gold nanoparticles, 1-mercapto-2-propanol, 1-nonanethiol, 1-octadecanethiol, 1-octanethiol, 1-pentadecanethiol, 1-pentanethiol, 1- Propanethiol, 1- Tetradecanethiol, 1-tetradecanethiolpurum, 1-undecanethiol, 11- (1H-pyrrol-1-yl) undecane-1-thiol, 11-amino-1-undecanethiol hydrochloride, 11-bromo-1-undecanethiol 11-mercapto-1-undecanol, 11-mercaptoundecanoic acid, 11-mercaptoundecyl trifluoroacetate, 11-mercaptoundecyl phosphate, 12-mercaptododecanoic acid, 15-mercaptopentadecanoic acid, 16-mercaptohexadecanoic acid 1H, 1H, 2H, 2H-perfluorodecanethiol, 2,2 ′-(ethylenedioxy) diethanethiol, 2,3-butanedithiol, 2-butanethiol, 2-ethylhexanethiol, 2-methyl-1 -Propanethiol, 2 -Methyl-2-propanethiol, 2-phenylethanethiol, 3,3,4,4,5,5,6,6,6-nonafluoro-1-hexanethiolpurum, 3- (dimethoxymethylsilyl) -1- Propanethiol, 3-chloro-1-propanethiol, 3-mercapto-1-propanol, 3-mercapto-2-butanol, 3-mercapto-N-nonylpropionamide, 3-mercaptopropionic acid, 3-mercaptopropyl functional group Silica gel, 3-methyl-1-butanethiol, 4,4′-bis (mercaptomethyl) biphenyl, 4,4′-dimercaptostilbene, 4- (6-mercaptohexyloxy) benzyl alcohol, 4-cyano-1 -Butanethiol, 4-mercapto-1-butanol, 6- (ferrocenyl) hexane , 6-mercapto-1-hexanol, 6-mercaptohexanoic acid, 8-mercapto-1-octanol, 8-mercaptooctanoic acid, 9-mercapto-1-nonanol, biphenyl-4,4′-dithiol, 3- Mercaptopropionate butyl, 1-butanethiolate copper (I), cyclohexanethiol, cyclopentanethiol, decanethiol functionalized silver nanoparticles, dodecanethiol functionalized gold nanoparticles, dodecanethiol functionalized silver nanoparticles, hexa (Ethylene glycol) mono-11- (acetylthio) undecyl ether, mercaptosuccinic acid, methyl 3-mercaptopropionate, NanoTether BPA-HH, NanoThinks ™ 18, NanoThinks ™ 8, NanoThinks ™ ACID11, NanoThinks (TM) ACID16, NanoThinks (TM) ALCO11, NanoThinks (TM) THIO8, octane thiol functionalized gold nanoparticles, PEG dithiol average _M n 8,000, PEG dithiol average molecular weight 1,500, PEG dithiol average molecular weight 3,400, S- (11-bromoundecyl) thioacetate, S- (4-cyanobutyl) thioacetate, thiophenol, triethylene glycol mono-11-mercaptoundecyl ether, trimethylolpropane tris (3-mercaptopropionate) ), [11- (methylcarbonylthio) undecyl] tetra (ethylene glycol), m-carborane-9-thiol, p-terphenyl-4,4 ''-dithiol, ter - dodecyl mercaptan, one or more agents selected from the group consisting of tert- nonyl mercaptan.

  In another embodiment, (i) prevention of infection of natural surfaces by bacterial, fungal or viral pathogens, (ii) inhibition of cell viability or cell growth of substantially all planktonic cells of bacterial, fungal or viral pathogens. (Iii) inhibition of biofilm formation by bacterial, fungal or viral pathogens, and (iv) inhibition of biofilm viability or biofilm development of virtually all biofilm-type cells of bacterial, fungal or viral pathogens Contacting the epithelial tissue surface with an effective amount of the BT composition at conditions and for a time sufficient for one or more of the particles, wherein the BT composition comprises a plurality of microparticles comprising a bismuth-thiol (BT) compound. And substantially all of the microparticles have a volume average diameter of about 0.4 μm to about 5 μm of bacterial, fungal and viral pathogens Chino against one or more, a method of protecting natural surface, including biological tissue surface, such as epithelial tissue surface is provided. In certain embodiments, the bacterial pathogen is (i) one or more gram negative bacteria; (ii) one or more gram positive bacteria; (iii) one or more antibiotic sensitive bacteria; (iv) one or more bacteria. (V) Staphylococcus aureus (S. aureus), MRSA (methicillin resistant S. aureus), Staphylococcus epidermidis, MRSE (methicillin resistant S. epidermidis), Mycobacterium tuberculosis Cis, Mycobacterium avium, Pseudomonas aeruginosa, drug resistance Aeruginosa, Escherichia coli, enterotoxigenic E. coli E. coli, enterohemorrhagic E. coli E. coli, Klebsiella pneumoniae, Clostridium difficile, Helicobacter pylori, Legionella pneumophila, Enterococcus faecalis, Methicillin-susceptible enterococcus fecaris, Enterobacter cloacae, Salmonella typhimurium, Proteus bulgaris, Yersinia enterico , Vibrio cholere, Shigella flexneri, vancomycin-resistant enterococcus (VRE), Burkholderia cepacia group, Francisella larensis, Bacillus anthracis, Yersinia pestis, Pseudomonas aeruginosa, vancomycin-resistant enterococcus genus, Streptococcus nippon Resistant Streptococcus pneumoniae, Escherichia Coli, including Burkholderia cepacia, Burkholderia multi borane scan, bacterial pathogens selected from Mycobacterium smegmatis and Acinetobacter Baumani, at least one of. In certain embodiments, the bacterial pathogen exhibits antibiotic resistance. In certain embodiments, the bacterial pathogen is resistant to an antibiotic selected from methicillin, vancomycin, nafcillin, gentamicin, ampicillin, chloramphenicol, doxycycline and tobramycin.

  In certain embodiments, the natural surface comprises an intraoral / oral surface. In further embodiments, the natural surface comprises a biological surface, such as a bone, joint, muscle, ligament, or tendon.

  In certain embodiments, the surface comprises an epithelial tissue surface comprising tissue selected from the epidermis, dermis, respiratory tract, gastrointestinal tract and glandular lining.

  In certain embodiments, the contacting step is performed one or more times. In certain embodiments, the at least one contacting step comprises one of spraying, ligating, dipping and applying a natural surface. In certain embodiments, the at least one contacting step includes one of inhalation, ingestion and oral ligation. In certain embodiments, the at least one contact step comprises topical, intraperitoneal, oral, parenteral, intravenous, intraarterial, transdermal, sublingual, subcutaneous, intramuscular, buccal, intranasal, inhalation, ocular Administration by a route selected from internal, intraauricular, intraventricular, intrafat, intraarticular and intrathecal. In certain embodiments, the BT composition comprises BisBAL, BisEDT, Bis-dimercaprol, Bis-DTT, Bis-2-mercaptoethanol, Bis-DTE, Bis-Pyr, Bis-Ery, Bis-Tol, Bis- BDT, Bis-PDT, Bis-Pyr / Bal, Bis-Pyr / BDT, Bis-Pyr / EDT, Bis-Pyr / PDT, Bis-Pyr / Tol, Bis-Pyr / Ery, Bismuth-1-mercapto-2- One or more BT compounds selected from the group consisting of propanol and Bis-EDT / 2-hydroxy-1-propanethiol are included.

  In certain embodiments, the bacterial pathogen exhibits antibiotic resistance. In other specific embodiments, the method comprises synergizing antibiotics and / or potentiating the natural surface simultaneously or sequentially in any order with respect to contacting the natural surface with the BT composition. It further comprises contacting with an antibiotic. In certain embodiments, the synergistic and / or potentiating antibiotics are aminoglycoside antibiotics, carbapenem antibiotics, cephalosporin antibiotics, fluoroquinolone antibiotics, glycopeptide antibiotics, lincosamides An antibiotic selected from antibiotics, penicillinase resistant penicillin antibiotics, and aminopenicillin antibiotics. In certain embodiments, the synergistic and / or potentiating antibiotic is an aminoglycoside antibiotic selected from amikacin, arbekacin, gentamicin, kanamycin, neomycin, netilmycin, paromomycin, rhodostreptomycin, streptomycin, tobramycin and apramycin It is.

  In another embodiment of the invention described herein, (i) prevention of infection by a bacterial pathogen on a natural surface where an antibiotic resistant pathogen is present, (ii) substantially all plankton of the bacterial pathogen Inhibition of cell viability or cell growth of sex cells, (iii) inhibition of biofilm formation by bacterial pathogens, and (iv) biofilm viability or biofilm growth of virtually all biofilm-type cells of bacterial pathogenesis In a condition and time sufficient for one or more of the inhibitions, the natural surface on which the antibiotic-resistant bacterial pathogen is present is converted to an effective amount of (1) at least one bismuth-thiol (BT) composition and ( 2) optional at the same time or sequentially with at least one antibiotic that is enhanced by and / or capable of acting synergistically with at least one BT composition The BT composition comprises a plurality of microparticles comprising a bismuth-thiol (BT) compound, wherein substantially all of the microparticles have a volume average diameter of about 0.4 μm to about 5 μm. Overcoming antibiotic resistance on the surface of epithelial tissues, overcoming antibiotic resistance on natural surfaces where antibiotic-resistant bacterial pathogens exist (for example, having antibacterial effects against bacteria of the same species) For bacterial pathogens that are resistant to at least one antimicrobial effect of at least one antibiotic known in the art, a method of making such pathogens susceptible to antibiotics is provided. In certain embodiments, the bacterial pathogen is (i) one or more gram negative bacteria; (ii) one or more gram positive bacteria; (iii) one or more antibiotic sensitive bacteria; (iv) one or more bacteria. (V) Staphylococcus aureus (S. aureus), MRSA (methicillin resistant S. aureus), Staphylococcus epidermidis, MRSE (methicillin resistant S. epidermidis), Mycobacterium tuberculosis Cis, Mycobacterium avium, Pseudomonas aeruginosa, drug resistance Aeruginosa, Escherichia coli, enterotoxigenic E. coli E. coli, enterohemorrhagic E. coli E. coli, Klebsiella pneumoniae, Clostridium difficile, Helicobacter pylori, Legionella pneumophila, Enterococcus faecalis, Methicillin-susceptible enterococcus fecaris, Enterobacter cloacae, Salmonella typhimurium, Proteus bulgaris, Yersinia enterico , Vibrio cholere, Shigella flexneri, vancomycin-resistant enterococcus (VRE), Burkholderia cepacia group, Francisella larensis, Bacillus anthracis, Yersinia pestis, Pseudomonas aeruginosa, vancomycin-resistant enterococcus genus, Streptococcus nippon Resistant Streptococcus pneumoniae, Escherichia Coli, including Burkholderia cepacia, Burkholderia multi borane scan, bacterial pathogens selected from Mycobacterium smegmatis and Acinetobacter Baumani, at least one of.

  In certain embodiments, the bacterial pathogen is resistant to an antibiotic selected from methicillin, vancomycin, nafcillin, gentamicin, ampicillin, chloramphenicol, doxycycline, tobramycin, clindamycin and gatifloxacin.

  In certain embodiments, the natural surface comprises an intraoral / oral surface. In further embodiments, the natural surface comprises a biological surface, such as a bone, joint, muscle, ligament, or tendon.

  In certain embodiments, the surface comprises tissue selected from the group consisting of the epidermis, dermis, respiratory tract, gastrointestinal tract, and glandular lining. In certain embodiments, the contacting step is performed one or more times. In certain embodiments, the at least one contacting step includes one of spraying, ligating, dipping and applying the surface. In other specific embodiments, the at least one contacting step comprises one of inhalation, ingestion and oral ligation. In certain embodiments, the at least one contact step comprises topical, intraperitoneal, oral, parenteral, intravenous, intraarterial, transdermal, sublingual, subcutaneous, intramuscular, buccal, intranasal, inhalation, ocular Administration by a route selected from internal, intraauricular, intraventricular, intrafat, intraarticular and intrathecal. In certain embodiments, the BT composition comprises BisBAL, BisEDT, Bis-dimercaprol, Bis-DTT, Bis-2-mercaptoethanol, Bis-DTE, Bis-Pyr, Bis-Ery, Bis-Tol, Bis- BDT, Bis-PDT, Bis-Pyr / Bal, Bis-Pyr / BDT, Bis-Pyr / EDT, Bis-Pyr / PDT, Bis-Pyr / Tol, Bis-Pyr / Ery, Bismuth-1-mercapto-2- One or more BT compounds selected from the group consisting of propanol and Bis-EDT / 2-hydroxy-1-propanethiol are included. In certain embodiments, the synergistic and / or potentiating antibiotics are clindamycin, gatifloxacin, aminoglycoside antibiotics, carbapenem antibiotics, cephalosporin antibiotics, fluoroquinolone antibiotics, An antibiotic selected from glycopeptide antibiotics, lincosamide antibiotics, penicillinase-resistant penicillin antibiotics, and aminopenicillin antibiotics. In certain embodiments, the synergistic and / or potentiating antibiotic is an aminoglycoside antibiotic selected from amikacin, arbekacin, gentamicin, kanamycin, neomycin, netilmycin, paromomycin, rhodostreptomycin, streptomycin, tobramycin and apramycin It is.

  In another embodiment, (a) at least one BT compound; (b) at least one antibiotic compound that is potentiated and / or capable of acting synergistically with the BT compound; and (C) A disinfecting composition is provided comprising a pharmaceutically acceptable excipient or carrier, including a carrier for topical use. In certain embodiments, the BT compound is BisBAL, BisEDT, Bis-dimercaprol, Bis-DTT, Bis-2-mercaptoethanol, Bis-DTE, Bis-Pyr, Bis-Ery, Bis-Tol, Bis-BDT. Bis-PDT, Bis-Pyr / Bal, Bis-Pyr / BDT, Bis-Pyr / EDT, Bis-Pyr / PDT, Bis-Pyr / Tol, Bis-Pyr / Ery, Bismuth-1-mercapto-2-propanol And Bis-EDT / 2-hydroxy-1-propanethiol. In certain embodiments, the BT composition comprises a plurality of microparticles comprising a bismuth-thiol (BT) compound, and substantially all of the microparticles have a volume average diameter of about 0.4 μm to about 5 μm. In certain embodiments, the BT compound is selected from BisEDT and BisBAL. In certain embodiments, the antibiotic compound comprises an antibiotic selected from methicillin, vancomycin, nafcillin, gentamicin, ampicillin, chloramphenicol, doxycycline, tobramycin, clindamycin, gatifloxacin and aminoglycoside antibiotics. . In certain embodiments, the aminoglycoside antibiotic is selected from amikacin, arbekacin, gentamicin, kanamycin, neomycin, netilmycin, paromomycin, rhodostreptomycin, streptomycin, tobramycin and apramycin. In certain embodiments, the aminoglycoside antibiotic is amikacin.

  In other specific embodiments, (a) bacterial infections on or in the surface are (i) gram positive bacteria, (ii) gram negative bacteria, and (iii) both (i) and (ii), And b) administering to the surface a treatment comprising one or more bismuth thiol (BT) compositions, thereby treating the surface, and A method is provided for treating a natural surface that supports or contains a film, wherein (i) when the bacterial infection comprises a Gram-positive bacterium, the combination is at least one BT compound and at least one rifamycin. (Ii) if the bacterial infection comprises a Gram negative bacterium, the combination comprises a therapeutically effective amount of at least one BT compound and amikacin, (Iii) Bacterial infection There comprising When including both Gram-positive and Gram-negative bacteria, compounding agents BT compounds, one or more therapeutically effective amount of a rifamycin and amikacin.

  In certain embodiments, the biofilm comprises one or more antibiotic resistant bacteria. In certain embodiments, treating the surface comprises (i) eradicating bacterial biofilm, (ii) reducing bacterial biofilm, and (iii) inhibiting bacterial biofilm growth. Including at least one of them. In certain embodiments, the BT composition comprises a plurality of microparticles comprising a bismuth-thiol (BT) compound, and substantially all of the microparticles have a volume average diameter of about 0.4 μm to about 5 μm.

  These and other aspects of the embodiments of the invention described herein will become apparent by reference to the following detailed description and accompanying drawings. U. S. RE37,793, U.S. Pat. S. No. 6,248,371, U.S. Pat. S. No. 6,086,921, and U.S. Pat. S. U.S. Patents and U.S. Patent Application Publications, U.S. Patent Applications, Foreign Patents, Foreign Patent Applications, Non-Patent Issuances Referenced herein and / or listed in the Application Data Sheet, including 6,380,248 All of the objects are hereby incorporated by reference in their entirety so that each is individually incorporated. Aspects and embodiments of the present invention can be modified as needed to provide further embodiments using the concepts of various patents, applications and publications.

Brief description of multiple appearances of drawings
Pseudomonas aeruginosa colony biofilm survival (log CFU; colony forming units) obtained after growth for 24 hours at 37 ° C. on 10% trypsin soy agar (TSA) followed by the indicated treatment for 18 hours Indicates. The indicated antibiotic treatments are: TOB, tobramycin 10 × MIC; AMK, amikacin, 100 × MIC; IPM, imipenem 10 × MIC; CEF, cefepime 10 × MIC; CIP, ciprofloxacin, 100 × MIC; Cpd 2B Compound 2B (Bis-BAL, 1: 1.5). (MIC; minimum inhibitory concentration, eg, lowest concentration to prevent bacterial growth). The number of surviving staphylococcus aureus colony biofilms (logCFU) obtained after growth on 10% trypsin soy agar for 24 hours and subsequent treatment as indicated is shown. Antibiotic treatments indicated were: rifamycin, RIF 100 × MIC; daptomycin, DAP 320 × MIC; minocycline, MIN 100 × MIC; ampicillin, AMP 10 × MIC; vancomycin, VAN 10 × MIC; Cpd 2B, compound 2B ( Bis-BAL, 1: 1.5), Cpd8-2, compound 8-2 (Bis-Pyr / BDT, 1: 1 / 0.5). Figure 2 shows scratch closure over time of keratinocytes exposed to biofilm. ( ^* ) Significantly different from control (P <0.001). Figure 2 shows subinhibited BisEDT that suppresses antibiotic resistance to multiple antibiotics. Figure 6 shows the effect of antibiotics with or without BisEDT on the MRSA (methicillin resistant S. aureus) flora. Panel A shows only a standard antibiotic immersion disc. [GM = gentamicin, CZ = cefazoline, FEP = cefepime, IPM = imipenem, SAM = ampicillin / sulbactam], LVX = levofloxacin. Figure 2 shows subinhibited BisEDT that suppresses antibiotic resistance to multiple antibiotics. Figure 6 shows the effect of antibiotics with or without BisEDT on the MRSA (methicillin resistant S. aureus) flora. Panel B shows a disc blended with BisEDT (BE). [GM = gentamicin, CZ = cefazoline, FEP = cefepime, IPM = imipenem, SAM = ampicillin / sulbactam], LVX = levofloxacin. Figure 2 shows the effect of BisEDT and antibiotics on biofilm formation. S. Epidermidis was grown for 48 hours at 37 ° C. in TSB + 2% glucose in polystyrene plates. Gatifloxacin (GF), clindamycin (CM), minocycline (MC), gentamicin (GM), vancomycin (VM), cefazolin (CZ), nafcillin (NC), and rifampicin (RP). The results were expressed as the average rate of variation of BPC (2-fold serial dilution step) with 0.25 μM BisEDT (n = 3). Grown in TSB + 2% glucose at 37 ° C. for 48 hours. Figure 2 shows the effect of BisEDT and antibiotics on the growth of epidermidis. Results are expressed as mean MIC (dilution step) variation with increasing BisEDT (n = 3). See the legend of FIG. 5 for the definition of antibiotic. In addition to the three BT formulations, Bis-EDT, MB-11 and MB-8-2, detected in bone and device samples of open fractures in in vivo rat models with or without cefazolin antibiotic treatment S. It is a bar graph which shows an Aureus average level. The standard error of the average value is shown as an error bar. Animals that were euthanized early were not excluded from the analysis, but a sample of one animal in group 2 was excluded due to significant contamination.

  Particular embodiments of the present invention disclosed herein may include specific bismuth-thiol (BT) compounds provided herein having BT microparticles having a volume average diameter of about 0.4 μm to about 5 μm. Potent disinfection, antibacterial and / or anti-bio against specific bacteria, including bacteria associated with multiple clinically significant infections, including infections that may include bacterial biofilms) It is based on the surprising discovery that it exhibits film activity but no other specific BT compounds (even if provided as microparticles).

  Contrary to expectations, not all BT compounds are uniformly effective in a predictable manner against such bacteria, but instead show different efficacy depending on the target species. In particular, as described herein, a specific BT compound can be applied in a manner that can for the first time provide clinically relevant strategies for the management of bacterial infections, including bacterial biofilm infections, by non-limiting theory. Although preferably found to contain higher efficacy against gram-negative bacteria, including BT microparticles having a volume average diameter of about 0.4 μm to about 5 μm, other specific BT compounds (preferably (Including BT microparticles having a volume average diameter of about 0.4 μm to about 5 μm) was found to show greater efficacy against gram positive bacteria.

  In addition, as described in more detail below, certain embodiments of the invention described herein may have particle sizes (eg, from about 0.4 μm to about 5 μm as disclosed herein). It relates to the surprising advantages provided by a novel bismuth-thiol (BT) composition that can be produced in a preparation comprising a plurality of BT microparticles that are substantially monodisperse with respect to (having a volume average diameter). In certain of these and related embodiments, the particulate BT is not provided as a component of lipid vesicles or liposomes, such as multilamellar phosphocholine-cholesterol liposomes or other multilamellar or unilamellar liposome vesicles.

  Also disclosed herein, certain antibiotic antibacterial and anti-biofilms previously found to lack a strong therapeutic effect against such bacterial infections with respect to certain embodiments Efficacy by treating the infection with either one or more of these antibiotics and the selected BT compound, either simultaneously, or sequentially, in either order (eg, an infection site such as a natural surface) Has been found to be significantly enhanced (eg, can be increased in a statistically significant manner). In an approach that could not have been predicted prior to the disclosure of the present invention, certain BT compounds may be used herein in combination with certain antibiotics for antibacterial and / or anti-biofilm activity against certain bacterial species or strains. Provided synergistic or enhancing combinations can be provided. Certain BT / antibiotics / combinations act synergistically or potentiate against a particular bacterium, while other specific BT / antibiotics / combinations have such synergistic or potentiating antibacterial and The observation that the anti-biofilm activity could not be demonstrated demonstrates the unexpected nature of such combinations described in more detail below.

  According to these and related embodiments, the antibiotic and BT compound may be administered simultaneously or sequentially in either order and may be formed by a particular infectious agent (eg, gram-negative or gram-positive). Specific synergistic or potentiating combinations of one or more antibiotics disclosed herein and one or more BT compounds to treat a biofilm) are predictable (eg, simply Instead of exhibiting (additive) activity, it is instead unexpectedly synergistic or potent (eg, superphase) as a function of the selected antibiotic, the selected BT compound and the specifically identified target bacteria It is noteworthy that it worked in an (additive) manner.

  For example, by way of example and not limitation, the specification herein in terms of a large variety of natural surfaces that are actually or potentially microbially infected, and further in terms of improved substantially monodisperse particulate BT formulations. Either or both of the specific antibiotic compound and the specific BT compound disclosed in the above may exhibit a slight antibacterial effect when used alone against a specific strain or strain, The combination of both compound and BT compound is stronger (with statistical significance) on a larger scale than the simple sum of the effects of each compound when used alone against the same strain or species. Antibacterial effect, thereby, according to non-limiting theory, the antibiotic-BT synergistic effect of BT on the efficacy of antibiotics and / or of antibiotics on the efficacy of BT (E.g., FICI ≦ 0.5) is believed to reflect or enhancing effect (e.g. 0.5 <FICI ≦ 1.0). Thus, each BT compound is not synergistic or potent with respect to each antibiotic, and each antibiotic is not synergistic or potent with respect to each BT compound. -BT synergy and BT-antibiotic potentiation are generally not predictable. Instead, according to certain embodiments disclosed herein, certain combinations that synergize or enhance antibiotics and BT compounds are surprisingly specific, such as the natural surfaces described herein. A strong antibacterial effect on specific bacteria, such as an antibacterial effect on biofilms formed by specific bacteria in certain environments, and even in specific situations.

  That is, certain BTs comprising antibiotics that can act synergistically with at least one BT composition comprising at least one BT compound provided herein (FICI ≦ 0.5). Antibiotics that synergize with are described herein, and such synergy is in the presence of antibiotics but no BT compounds and / or BT compounds but no antibiotics. In some cases, it manifests as a detectable effect on a larger scale than the detectable effect (ie, in a statistically significant manner with respect to the appropriate control conditions). Similarly, certain BT-antibiotic combinations show potentiating properties (0.5 <FICI <1.0), such potentiating properties in the presence of antibiotics but no BT compounds, and When a BT compound is present but no antibiotic is present, it develops as a detectable effect on a larger scale than that which can be detected (ie, in a statistically significant manner with respect to the appropriate control conditions).

  Examples of such detectable effects include, in certain embodiments, (i) prevention of infection by bacterial pathogens, (ii) inhibition of cell viability or cell growth of substantially all planktonic cells of bacterial pathogens. , (Iii) inhibition of biofilm formation by bacterial pathogenesis, and (iv) inhibition of biofilm viability or biofilm development of substantially all biofilm type cells of bacterial pathogenesis, In other contemplated embodiments, without limitation, antibiotic-BT synergy is one or more other clinically important parameters, such as one or more antibiotics or other drugs or chemistry. Degree of resistance or susceptibility of bacterial pathogens to agents, resistance of bacterial pathogens to one or more chemical, physical or mechanical conditions (eg pH, ionic strength, temperature, pressure) Degree of sex and sensitivity, and / or one or more biological agents (eg, viruses, other bacteria, biologically active polynucleotides, immune cells or immune cell products such as antibodies, cytokines, chemokines, degrading enzymes, etc. One or more detectable that may include a change (eg, statistically significant increase or decrease) in the resistance or susceptibility of bacterial pathogens to the enzyme, membrane-disrupting protein, free radicals such as reactive oxygen species, etc. You may express as an effect.

  As provided herein, an antibiotic present when the effect of a synergistic or potentiating antibiotic-BT combination exceeds the simple sum of the effects observed in the absence of one component of the combination. Those of skill in the art will understand these and various other criteria that can determine the effect of a particular agent on structure, function, and / or activity of a bacterial population in order to confirm substance-BT synergy or potentiation. Wax (e.g., Coico et al. (Eds.), Current Protocols in Microbiology, 2008, John Wiley & Sons, Hoboken, NJ; Schwalbe et al. 007, CRC Press, Boca Raton, FL).

  For example, in certain embodiments, antibacterial effects as described herein are measured using various concentrations of candidate agents (eg, BT and antibiotics alone or in combination) to determine Eliopoulos et al. (Antibiotics). in Laboratory Medicine (Lorian, V., Ed.), pp. 330-96, Williams and Willkins, Baltimore, MD, Eliopoulos and Moellering, (1996) Anticoin concentration in the USA. Synergy may be determined by calculating the bactericidal concentration index (FBCI). Synergy may be defined as a FICI or FBCI index of less than 0.5 and antagonism greater than 4 (see, eg, Odds, FC (2003) Synergy, antagonists, and what the chemboard puts between ther. Antimicrobiological Chemotherapy 52: 1). Synergy is conventionally defined as a four-fold decrease in antibiotic concentration or using partial inhibitory concentration (FIC) as described, for example, by Hollander et al. (1998 Antimicrob. Agents Chemother. 42: 744). May be. In certain embodiments, synergy is obtained with a combination of two drugs (eg, antibiotic and BT composition) and is greater than if the concentration of the second drug is replaced with the first drug ( It may be defined as a combined effect (for example, in a statistically significant manner).

  Thus, as described herein, it will be appreciated that in certain preferred embodiments, the combination of BT and antibiotics synergizes when a FICI value of 0.05 or less is observed (Odds, 2003). Also as described herein, in other specific preferred embodiments, and according to non-limiting theory, a specific BT-antibiotic combination may represent such a highly synergistic possibility. It is disclosed that it can exhibit FICI values between 5 and 1.0, the synergy of which is non-optimal of at least one BT and at least one antibiotic exhibiting unilateral or mutually enhanced cooperative antimicrobial efficacy May be observed using concentration. Such an effect may be referred to herein as “enhanced” antibiotic activity or “enhanced” BT activity.

According to certain embodiments, (i) a concentration (in a statistically significant manner) that is lower than the characteristic minimum inhibitory concentration (MIC) of BT for a given target microorganism (eg, a given strain or strain) Lower than (ii) a characteristic IC ₅₀ (concentration that inhibits 50% development of the microbial population; see, eg, Sothill et al., 1992 J Antimicrob Chemother 29 (2): 137) ( The presence of both at least one antibiotic in a statistically significant manner) and / or at a concentration lower than the biofilm prophylactic concentration (BPC) of the antibiotic for a given target microorganism, either One antimicrobial agent (eg, BT or antibiotic) is used at the same concentration in the absence of the other antibacterial agent (eg, antibiotic or BT). For anti-microbial effect is, when the result in high (in a statistically significant technique) BT- antibiotic antimicrobial efficacy of substances and combinations, enhancing antibiotics and / or BT activity can be detected. In preferred embodiments, “enhanced” antibiotic and / or BT activity is present when a FICI value of less than 1.0 and greater than 0.5 is measured.

  As will be appreciated by those of skill in the art based on the present disclosure, in certain embodiments, synergistic or potentiating antibiotics and / or BT activity may be generated by methods known in the art, for example, to the addition of Loewwe. Models based on (eg, FIC index, Greco model), or models based on Bliss independence (eg, nonparametric and semiparametric models) or other methods known in the art described herein (eg, Melediadis et al., 2005 Medical Mycology 43: 133-152). Exemplary methods for measuring synergistic or potentiating antibiotics and / or BT activity in this way are described, for example, by Melediadis et al. , 2005 Medical Mycology 43: 133-152 and references cited therein (also Melediadis et al., 2002 Rev Med Microbial 13: 101-117; White et al., 1996 AntimicrobAg: 40 centimeters). 1914-1918; see Mouton et al., 1999 Antimicrob Agents Chemother 43: 2473-3478).

  Other specific embodiments show that BT compounds and antibiotics do not exhibit predictable (simply additive) activity in vivo, instead of selecting selected antibiotics, selected BT compounds, and infectious agents. Unexpectedly synergistic or potentiating (eg, superadditive; or where two or more such agents are present in combination as a function of one or more of the specifically identified target species including Even when acting in a manner that provides a greater effect (eg, in a statistically significant manner) than would be obtained if the concentration of the second drug was replaced with the first drug. One or more antibiotics disclosed herein that may exhibit a synergistic or potentiating effect in vivo for the treatment of infectious agents (eg, biofilms formed by Gram negative or Gram positive bacteria) Substance and one or more It contemplates specific combination of the BT compound. Therefore, according to these and related embodiments, in certain in vivo situations, FICI or FBCI values (measured in vitro) may not be readily available and instead synergistic or potentiating effects of BT-antibiotics It will be appreciated that may be determined in a manner obtained by a quantifiable measure of the infectious agent.

  For example, in one embodiment such as the rat femur critical defect model of in vivo open fracture described in Example 11, observed after treatment with BT-antibiotic combination, compared to antibiotic treatment or BT compound alone. A statistically significant decrease in the number of bacteria is an indication of a synergistic or potentiating effect. Statistical significance can be determined using methods well known to those skilled in the art. In other specific embodiments, at least 5%, 10%, 20%, 30% of the number of bacteria observed in the injury after treatment with the BT-antibiotic combination, compared to antibiotic treatment or the BT compound alone, A decrease observed in this or other in vivo models of 40% or 50% is considered an indicator of synergistic or enhancing effects.

  Other exemplary indicators of in vivo infection include established methodologies developed to quantify the severity of infection, such as various wound scoring systems known to those skilled in the art (eg, European Wound Management Association (EWMA), Position Document: Identifying criteria for Wound Infection. London: See scoring system reviewed in MEP Ltd, 2005). Exemplary wound scoring systems that can be used in assessing the synergistic or potentiating activity of the BT-antibiotic combinations described herein include ASEPSIS (Wilson AP, J Hosp Infect 1995; 29 (2). Wilson et al., Lancet 1986; 1: 311-13), the Southampton Wound Assessment Scale (Baley IS, Karlan SE, Toyn K, et al. BMJ 1992; 304: 1992; 304: 1992; . Horan TC, Gaynes P, Martone WJ, et al. 1992 Infect Control Hosp Epidemiol 1992; 13: 606-08. In addition, recognized clinical indicators of wound healing known to skilled clinicians such as wound size, depth, granulation tissue status, infection, etc. in the presence or absence of BT compounds and / or antibiotics You may measure with. Thus, based on the present disclosure, to determine whether a BT composition-antibiotic combination changes in vivo wound healing (eg, increases or decreases in a statistically significant manner relative to the appropriate control) Various methods will be readily appreciated by those skilled in the art.

  In view of these and related embodiments, a microbially infected natural surface, such as a surface that supports or contains a bacterial biofilm, is treated with one or more BT compounds and optionally one or more antibiotic compounds, such as the present specification. Treatment with an effective amount (eg, in certain embodiments, a therapeutically effective amount) of a composition or combination comprising one or more synergistic antibiotics or one or more potentiating antibiotics provided in a written document A great variety of ways to do this are provided herein. Based on this disclosure, it will be appreciated that certain antibiotics previously considered ineffective for a given type of infection by those skilled in the art are contemplated herein for use in the treatment of the same type of infection. I want.

  Thus, certain embodiments contemplate compositions that include one or more BT compounds that are used as disinfectants. A disinfectant is a substance that kills or interferes with the growth of microorganisms and is typically applied to living tissue to distinguish classification from those typically applied to inanimate objects. (Goodman and Gilman's “The Pharmacological Basis of Therapeutics”, Seventh Edition, Gilman et al., Editorial, 1985, Book of G. G. Common examples of disinfectants are ethyl alcohol and iodine tincture. Disinfectants include disinfectants that kill microorganisms such as microbial pathogens.

Certain embodiments described herein include one or more BT compounds and one or more antibiotic compounds (eg, synergistic and / or potentiating antibiotics provided herein). A composition comprising it may be contemplated. Antibiotics are known in the art and are typically drugs produced by compounds that are produced by one species of microorganism and kill another species of microorganism, or the same or similar chemical structure and mechanism of action. For example, drugs that destroy microorganisms in or on the surface of living organisms when applied topically. In particular, embodiments disclosed herein are those in which the antibiotic may belong to one of the following classes: aminoglycosides, carbapenems, cephalosporins, fluoroquinolones, glycopeptide antibiotics, lincosamides (eg , Clindamycin), penicillinase resistant penicillin, and aminopenicillin. In other words, antibiotics need not be limited, but oxacillin, piperacillin, cefuroxime, cefotaxime, cefepime, imipenem, aztreonam, streptomycin, tobramycin, tetracycline, minocycline, ciprofloxacin, levofloxacin, erythromycin, linezolid, capreomycin, , Isoniazid, ansamycin, carbacephem, monobactam, nitrofuran, penicillin, quinolone, sulfonamide, clofazimine, dapsone, capreomycin, cycloserine, ethambutol, etionamide, isoniazid, pyrazinamide, rifamupicin, rifamupine, rifabutin, streptomycin, streptomycin, streptomycin Namin, Chloramphenicol, Phospho Isin, fusidic acid, linezolid, metronidazole, mupirocin, platencimycin, quinupristin, darfopristin, rifaximin, thiamphenicol, tinidazole, aminoglycoside, β-lactam, penicillin, cephalosporin, carbapenem, fluroquinolone, ketolide, lincosamide, macrolide Oxazolidinone, streptogramin, sulfonamide, tetracycline, glycylcycline, methicillin, vancomycin, nafcillin, gentamicin, ampicillin, chloramphenicol, doxycycline, tobramycin, amikacin, arbekacin, gentamicin, kanamycin, neomycin, netilmycin, paromomycin, rhodomycin Streptomycin, Tobrama Mention may be made of isine, apramycin, clindamycin, gatigloxacin, aminopenicillin, and others known in the art. An overview of these and other clinically useful antibiotics is available and known to those of skill in the art (see, eg, Washington University School of Medicine, The Washington Manual of Medicine Therapeutics E. 32 ^nd . Williams and Wilkins, Philadelphia, PA; Hauser, AL, Antibiotic Basics for Clinicians, 2007 Lippincott, Williams and Wilkins, Philadelphia, PA).

  An exemplary class of antibiotics used with one or more BT compounds in certain embodiments disclosed herein can be found in Edson RS, Terrell CL. The aminoglycosides. Mayo Clin Proc. 1999 May; 74 (5): 519-28, an aminoglycoside antibiotic. This class of antibiotics inhibits bacterial growth by damaging bacterial protein synthesis through binding and inactivation of bacterial ribosomal subunits. In addition to such bacteriostatic properties, aminoglycosides also exhibit bactericidal effects through the disruption of cell walls of gram-negative bacteria.

  Aminoglycoside antibiotics include gentamicin, amikacin, streptomycin and the like and are generally considered useful for the treatment of Gram-negative bacteria, mycobacteria and other microbial pathogens, but the classification of resistant strains has been reported. Aminoglycosides are not absorbed from the gastrointestinal tract and are therefore generally not considered applicable to oral formulations. Amikacin, for example, is often effective against gentamicin-resistant strains, but is typically administered intravenously or intramuscularly and can induce pain in patients. In addition, the toxicity associated with aminoglycoside antibiotics such as amikacin can lead to kidney damage and / or irreversible hearing damage.

  Despite these properties, certain embodiments disclosed herein are synergistic to treat epithelial tissue surfaces at one or more locations, eg, along the oral cavity, gastrointestinal / gastrointestinal tract. Contemplates oral administration of the BT / antibiotic combination (eg, the antibiotic need not be limited to aminoglycosides). Also contemplated in other specific embodiments are described herein as bactericides that refer to preparations that kill or block the growth of microorganisms on the outer surface of an inanimate object. Use of the prepared compositions and methods.

  As also described elsewhere herein, BT compounds are described in Domenico et al. , 1997 Antimicrob. Agent. Chemother. 41 (8): 1697-1703, Domenico et al. , 2001 Antimicrob. Agent. Chemother. 45 (5): 1417-1421, and U.S. Pat. S RE 37,793, U.S. Pat. S. 6,248,371, 6,086,921, and 6,380,248 (see also, for example, US 6,582,719) (including preparation methods) It may be a composition comprising a bismuth or bismuth salt and a thiol (eg, —SH, or sulfhydryl) -containing compound. However, certain embodiments are not so limited and other BT compounds including bismuth or bismuth salts and thiol-containing compounds may be contemplated. The thiol-containing compound may include 1, 2, 3, 4, 5, 6 or more thiol (eg, -SH) groups. In preferred embodiments, the BT compound comprises bismuth associated with a thiol-containing compound via ionic bonds and / or as a coordination complex, while in some other embodiments, bismuth is present in the organometallic compound. May be associated with the thiol-containing compound via a covalent bond. However, certain contemplated embodiments explicitly exclude organometallic compounds, such as BT compounds, where bismuth is a compound found in a covalent linkage to an organic moiety.

＊ 硫黄含有化合物と三価錯体を形成するために用いられる反応体の化学量論比およびビスマスの公知の傾向に基づき、比較のために単一ビスマス原子に対する原子比を示す。示された原子比は、所定の調製物中の全ての化学種の正確な分子式でない場合がある。カッコ内の数字は、１種（またはそれを超える）チオール剤に対するビスマスの比である（例えば、Ｂｉ：チオール１／チオール２）。「ＣＰＤ」：化合物。 Exemplary BT compounds are shown in Table 1.

* Based on the stoichiometric ratio of the reactants used to form the trivalent complex with the sulfur-containing compound and the known tendency of bismuth, the atomic ratio for a single bismuth atom is shown for comparison. The indicated atomic ratios may not be the exact molecular formula of all chemical species in a given preparation. The numbers in parentheses are the ratio of bismuth to one (or more) thiol agent (eg, Bi: thiol 1 / thiol 2). “CPD”: Compound.

  The BT compounds used in certain of the embodiments disclosed herein are well-established procedures (eg, US RE 37,793, US 6,248,371, 6,086, 921, and 6,380,248; Domenico et al., 1997 Antimicrob. Agent. Chemother. 41 (8): 1697-1703, Domenico et al., 2001 Antimicrob. Agent. 1417-1421), and in other specific embodiments, BT compounds may be prepared according to the methodology described herein. That is, certain preferred embodiments may be used under conditions and times sufficient to form a precipitate comprising microparticles comprising a BT compound (eg, concentrations, solvents, as described herein and understood by those of skill in the art based on this disclosure). (Under conditions such as strength, temperature, pH, mixing and / or pressure), the dissolved bismuth is at least 50 mM, at least 100 mM, at least 150 mM, at least 200 mM, at least 250 mM, at least 300 mM, at least 350 mM, at least 400 mM, at least 500 mM, at least An acidic bismuth aqueous solution containing 600 mM, at least 700 mM, at least 800 mM, at least 900 mM, or at least 1 M and containing no hydrophilic, polar or organic solubilizer is mixed with ethanol to form a first ethanol. BT is obtained and reacted with a second ethanolic solution containing a thiol-containing compound present in the reaction solution in a molar ratio of about 1: 3 to about 3: 1 with respect to bismuth to obtain a reaction solution. The synthetic methods described herein for preparing compounds, and in particular for obtaining BT compounds in substantially monodisperse particulate form, are contemplated.

  Thus, exemplary BTs include Compound 1B-1, Bis-EDT (1: 1 bismuth-1,2-ethanedithiol reactant); Compound 1B-2, Bis-EDT (1: 1.5); Compound 1B -3, Bis-EDT (1: 1.5); Compound 1C, Bis-EDT (soluble Bi preparation, 1: 1.5); Compound 2A, Bis-Bal (bismuth-British anti-Lewisite (bismuth-di) Mercaprol, bismuth-2,3-dimercaptopropanol), 1: 1); Compound 2B, Bis-Bal (1: 1.5); Compound 3A Bis-Pyr (bismuth pyrithione, 1: 1.5); Compound 3B, Bis-Pyr (1: 3); Compound 4, Bis-Ery (bismuth dithioerythritol, 1: 1.5); Compound 5, Bis-Tol (bismuth- 3,4-dimercaptotoluene, 1: 1.5); Compound 6, Bis-BDT (bismuth-2,3-butanediol, 1: 1.5); Compound 7, Bis-PDT (bismuth-1,3) -Propanedithiol, 1: 1.5); Compound 8-1 Bis-Pyr / BDT (1: 1/1); Compound 8-2, Bis-Pyr / BDT (1: 1 / 0.5); Compound 9 Bis-2-hydroxy, propanethiol (bismuth-1-mercapto-2-propanol, 1: 3); Compound 10, Bis-Pyr / Bal (1: 1 / 0.5); Compound 11, Bis-Pyr / EDT (1: 1 / 0.5); Compound 12 Bis-Pyr / Tol (1: 1 / 0.5); Compound 13, Bis-Pyr / PDT (1: 1 / 0.5); Compound 14 Bis- Pyr / ry (1: 1 / 0.5); compound 15, Bis-EDT / 2- hydroxy propane thiol (1: 1/1) and the like (e.g., see Table 1).

  While not wishing to be bound by theory, in certain preferred embodiments, the BT may comprise preparing or obtaining an acidic aqueous liquid solution containing bismuth, such as an aqueous nitric acid solution containing bismuth nitrate. The disclosed methods for preparing compounds can be used, if desired, for ease of mass production, improved product purity, homogeneity or consistency (including particle size uniformity), or topical formulations of the present invention. It is contemplated that a composition comprising a BT compound can be obtained having one or more desired properties, including other properties useful for the preparation and / or administration of

  In certain embodiments, the BT compositions prepared by the methods described herein for the first time each have a volume average diameter (VMD) of about 0.4 μm to about 5 μm, according to certain presently preferred embodiments. It has been discovered that it exhibits advantageous homogeneity in relation to their appearance as a substantially monodispersed particulate suspension. The measured particle diameter can be referred to as the volume average diameter (VMD), mass median diameter (MMD), or aerodynamic mass median diameter (MMAD). These measurements may be performed, for example, by characterization with impaction (MMD and MMAD) or laser (VMD). In the case of liquid particles, VMD, MMD, and MMAD can be the same if environmental conditions are maintained, eg, standard humidity. However, if the humidity is not maintained, the MMD and MMAD measurements will be smaller than the VMD due to dehydration during the impactor measurement. For purposes of this specification, VMD, MMD and MMAD measurements will be considered under standard conditions, so the description of VMD, MMD and MMAD would be equivalent. Similarly, dry powder particle size measurements with MMD and MMAD are considered equivalent.

  As described herein, a preferred embodiment relates to a substantially monodispersed suspension of BT-containing microparticles. Generating a defined BT particle size with a slight geometric standard deviation (GSD) can, for example, attach BT, accessibility to a desired target site in or on a natural surface, and / or BT microparticles Tolerability may be optimized by the subject to whom is administered. A narrow GSD will limit the number of particles outside the desired VMD or MMAD size range.

  In one embodiment, a liquid or aerosol suspension of microparticles having a VMD of about 0.5 microns to about 5 microns containing one or more BT compounds disclosed herein is provided. In another embodiment, a liquid or aerosol suspension having a VMD or MMAD of about 0.7 microns to about 4.0 microns is provided. In another embodiment, a liquid or aerosol suspension having a VMD or MMAD of about 1.0 microns to about 3.0 microns is provided. In other specific embodiments, the VMD is about 0.1 to about 5.0 microns, or about 0.1, about 0.2, about 0.3, about 0.4, about 0.5, about 0.0. 6, about 0.7, about 0.8, or about 0.9 to about 1.0, about 1.5, about 2.0, about 2.5, about 3.0, about 3.5, about 4. 0, about 4.5, about 5.0, about 5.5, about 6.0, about 6.5, about 7.0, about 7.5, or about 8.0 microns described herein A liquid suspension comprising one or more BT compound particles prepared as described above is provided.

  Thus, in certain preferred embodiments, “substantially” all of the BT preparations first described herein that are “substantially” monodisperse, eg, “substantially” all of the microparticles are within a specific range (eg, about 0 A BT composition comprising a BT compound in particulate form having a volume average diameter (VMD) of .4 μm to about 5 μm) is at least 80%, 85%, 90%, 91%, 92%, 93%, or 94 of the particles. %, More preferably at least 95%, 96%, 97%, 98%, 99% or more of those compositions having VMD within the quoted size range.

  These and related properties of BT compositions prepared by the synthetic methods described herein enable characterization in a manner that promotes regulatory compliance in accordance with one or more pharmaceutical, prescription and medicinal cosmetic standards. Presents unprecedented advantages over previously described BT, such as lower manufacturing costs and ease of manufacture, and homogeneity within the composition.

  In addition or alternatively, the substantially monodispersed BT microparticles described herein advantageously do not require micronization, i.e. expensive and laborious milling or supercritical fluid processing, or microparticles May be produced without using other equipment and procedures typically used to generate (eg, Martin et al. 2008 Adv. Drug Deliv. Rev. 60 (3): 339; Moribe et al. Rev. 60 (3): 328; Cape et al., 2008 Pharm.Res. 25 (9): 1967; Rasenack et al. 2004 Pharm.Dev.Technol. 1-13). Thus, embodiments of the present invention include, but are not limited to, enhanced and substantially homogeneous solubilizing properties, suitability for desired dosage forms such as oral, inhalation or topical forms on dermatological / skin wounds, It presents the beneficial effects of a substantially homogeneous microparticle preparation, including a high degree of bioavailability and other beneficial properties.

  BT compound particulate suspensions are, for example, wetting agents, surfactants, mineral oils, or other ingredients or additives that would be known to those skilled in the formulation to retain individual particulates in the suspension. As an aqueous formulation, as a suspension or solution in an aqueous and organic solvent such as a halogenated hydrocarbon propellant, as a dry powder, or in other forms detailed below be able to. Aqueous formulations may be aerosolized with a liquid nebulizer, for example, using either a hydraulic or ultrasonic spray. A propellant-based system may use a suitable pressurized dispenser. As the dry powder, a dry powder dispersion device capable of effectively dispersing the BT-containing fine particles may be used. The desired particle size and distribution may be obtained by selecting an appropriate device.

  As previously mentioned, also provided herein by certain embodiments is a method of preparing a BT composition comprising a plurality of microparticles comprising a bismuth-thiol (BT) compound, such as Substantially all of the fine particles have a volume average diameter (VMD) of about 0.1 to about 8 microns, and in certain preferred embodiments from about 0.4 microns to about 5 microns.

  In general conditions, the method comprises (a) a bismuth salt containing bismuth at a concentration of at least 50 mM, under conditions and time sufficient to obtain a solution substantially free of solid precipitate, (Ii) admixed with a sufficient amount of ethanol to mix at least about 5% by volume, 10% by volume, 15% by volume, 20% by volume, 25% by volume. Or obtaining an admixture comprising 30% by volume, preferably about 25% by volume ethanol; and (b) about 1: 3 to bismuth with conditions and time sufficient to form a precipitate comprising the BT compound. Adding an ethanolic solution containing a thiol-containing compound present in the reaction solution in a molar ratio of about 3: 1 to the blend of (a) to obtain a reaction solution.

In certain preferred embodiments, the bismuth salt may be Bi (NO ₃ ) ₃ , but it will be understood by the present disclosure that bismuth can be provided in other forms. In certain embodiments, the bismuth concentration in the acidic aqueous solution is at least 100 mM, at least 150 mM, at least 200 mM, at least 250 mM, at least 300 mM, at least 350 mM, at least 400 mM, at least 500 mM, at least 600 mM, at least 700 mM, at least 800 mM, at least 900 mM or It may be at least 1M. In certain embodiments, the acidic aqueous solution comprises at least 5 wt%, 10 wt%, 15 wt%, 20 wt%, 22 wt% or 22.5 wt% bismuth. The acidic aqueous solution may comprise at least 5 wt% or more nitric acid in certain preferred embodiments, and in other specific embodiments, the acidic aqueous solution is at least 0.5 wt%, at least 1 wt%. At least 1.5 wt%, at least 2 wt%, at least 2.5 wt%, at least 3 wt%, at least 3.5 wt%, at least 4 wt%, at least 4.5 wt% or at least 5 wt% nitric acid May be included.

The thiol-containing compound may be any thiol-containing compound described herein, and in certain embodiments, 1,2-ethanedithiol, 2,3-dimercaptopropanol, pyrithione, dithioerythritol, Including one or more of 3,4-dimercaptotoluene, 2,3-butanedithiol, 1,3-propanedithiol, 2-hydroxypropanethiol, 1-mercapto-2-propanol, and dithiothreitol Also good. Other exemplary thiol-containing compounds include α-lipoic acid, methanethiol (CH ₃ SH [m-mercaptan]), ethanethiol (C ₂ H ₅ SH [e-mercaptan]), 1-propanethiol (C ₃ H ₇ SH [n-P mercaptans]), 2- propanethiol _{(CH 3 CH (SH) CH} 3 [2C 3 mercaptan]), butanethiol _{(C 4} H 9 SH [n- butyl mercaptan]), tert Butyl mercaptan (C (CH ₃ ) ₃ SH [t-butyl mercaptan]), pentanethiol (C ₅ H ₁₁ SH [pentyl mercaptan]), coenzyme A, lipoamide, glutathione, cysteine, cystine, 2-mercaptoethanol, dithiotray Tall, dithioerythritol, 2-mercaptoindole, trans Rutaminase, and any of the following thiol compounds available from Sigma-Aldrich (St. Louis, MO): (11-mercaptoundecyl) hexa (ethylene glycol), (11-mercaptoundecyl) tetra (ethylene glycol) , (11-mercaptoundecyl) tetra (ethylene glycol) functionalized gold nanoparticles, 1,1 ′, 4 ′, 1 ″ -terphenyl-4-thiol, 1,11-undecanedithiol, 1,16- Hexadecanedithiol, technical grade 1,2-ethanedithiol, 1,3-propanedithiol, 1,4-benzenedimethanethiol, 1,4-butanedithiol, 1,4-butanedithioldiacetate, 1, 5-pentanedithiol, 1,6-hexanedithiol, 1,8 Octanedithiol, 1,9-nonanedithiol, adamantanethiol, 1-butanethiol, 1-decanethiol, 1-dodecanethiol, 1-heptanethiol, 1-heptanethiol plum, 1-hexadecanethiol, 1-hexanethiol, 1 -Mercapto (triethylene glycol), 1-mercapto (triethylene glycol) methyl ether functionalized gold nanoparticles, 1-mercapto-2-propanol, 1-nonanethiol, 1-octadecanethiol, 1-octanethiol, 1- Pentadecanethiol, 1-pentanethiol, 1-propanethiol, 1-tetradecanethiol, 1-tetradecanethiol plum, 1-undecanethiol, 11- (1H-pyrrol-1-yl) undecane-1-thiol, 11-ami No-1-undecanethiol hydrochloride, 11-bromo-1-undecanethiol, 11-mercapto-1-undecanol, 11-mercaptoundecanoic acid, 11-mercaptoundecyltrifluoroacetate, 11-mercaptoundecyl phosphate, 12-mercaptododecanoic acid, 15-mercaptopentadecanoic acid, 16-mercaptohexadecanoic acid, 1H, 1H, 2H, 2H-perfluorodecanethiol, 2,2 ′-(ethylenedioxy) diethanethiol, 2,3-butanedithiol 2-butanethiol, 2-ethylhexanethiol, 2-methyl-1-propanethiol, 2-methyl-2-propanethiol, 2-phenylethanethiol, 3,3,4,4,5,5,6, 6,6-nonafluoro-1-hexanethiolpurum, 3- (di Methoxymethylsilyl) -1-propanethiol, 3-chloro-1-propanethiol, 3-mercapto-1-propanol, 3-mercapto-2-butanol, 3-mercapto-N-nonylpropionamide, 3-mercaptopropionic acid , 3-mercaptopropyl functionalized silica gel, 3-methyl-1-butanethiol, 4,4′-bis (mercaptomethyl) biphenyl, 4,4′-dimercaptostilbene, 4- (6-mercaptohexyloxy) benzyl Alcohol, 4-cyano-1-butanethiol, 4-mercapto-1-butanol, 6- (ferrocenyl) hexanethiol, 6-mercapto-1-hexanol, 6-mercaptohexanoic acid, 8-mercapto-1-octanol, 8 -Mercaptooctanoic acid, 9-mercapto-1 Nonanol, biphenyl-4,4′-dithiol, butyl 3-mercaptopropionate, copper (I) 1-butanethiolate, cyclohexanethiol, cyclopentanethiol, decanethiol functionalized silver nanoparticles, dodecanethiol functionalized gold Nanoparticles, dodecanethiol functionalized silver nanoparticles, hexa (ethylene glycol) mono-11- (acetylthio) undecyl ether, mercaptosuccinic acid, methyl 3-mercaptopropionate, NanoTether BPA-HH, NanoThink ™ 18, NanoThinks (TM) 8, NanoThinks (TM) ACID11, NanoThinks (TM) ACID16, NanoThinks (TM) ALCO11, NanoThinks (TM) THIO8, Octane All functionalized gold nanoparticles, PEG dithiol average _M n 8,000, PEG dithiol average molecular weight 1,500, PEG dithiol average molecular weight 3,400, S- (11- bromo undecyl) thioacetate, S- (4-cyanobutyl ) Thioacetate, thiophenol, triethylene glycol mono-11-mercaptoundecyl ether, trimethylolpropane tris (3-mercaptopropionate), [11- (methylcarbonylthio) undecyl] tetra (ethylene glycol), m-carborane -9-thiol, p-terphenyl-4,4 ''-dithiol, tert-dodecyl mercaptan and tert-nonyl mercaptan.

  Exemplary reaction conditions such as temperature, pH, reaction time, use of stirring or agitation to dissolve solutes, and procedures for recovering and washing the precipitate are described herein and are generally described in the art. A known technique is used.

  Unlike previously described methodologies for producing BT compounds, according to the method of the present invention for preparing BT, the BT product, in certain preferred embodiments, is about 0.4 microns to about Provided as a microparticle suspension with substantially all microparticles having a VMD of 5 microns, and according to other particular embodiments, generally from about 0.1 microns to about 8 microns. Further, unlike previous approaches, according to embodiments of the present invention, bismuth is a bismuth salt at a concentration of at least about 50 mM to about 1 M, and at least about 0.5% (w / w) to about 5% (w / W), preferably in an acidic aqueous solution containing nitric acid in an amount of less than 5% (w / w) and free of hydrophilic, polar or organic solubilizers.

  In this regard, the method of the present invention is such that bismuth is 50 μM and is not water soluble (eg, US RE 37793) and bismuth is unstable in water (eg, Kuvshinova et al., 2009 Russ. J). Inorg. Chem 54 (11): 1816), surprising and expected in view of the generally recognized art that bismuth is also unstable in aqueous nitric acid unless hydrophilic, polar or organic solubilizers are present. Present outside profits. For example, in all of the most reliable descriptions of BT preparation methodologies (eg, Domenico et al., 1997 Antimicrob. Agents. Chemother. 41: 1697; US 6,380,248; US RE 37,793). No., US 6,248,371), the hydrophilic solubilizing agent propylene glycol is required to dissolve bismuth nitrate and the bismuth concentration of the solution prepared for reaction with thiol is 15 mM. Much lower, thereby limiting the available modes of production of BT compounds.

  In contrast, according to the present disclosure, a hydrophilic, polar or organic solubilizer is not required to dissolve bismuth, and surprisingly higher concentrations are achieved. Hydrophilic, polar or organic solubilizers include propylene glycol (PG) and ethylene glycol (EG), polar solvents such as dioxane and dimethylene sulfoxide (DMSO), polyols (eg polyethylene glycol (PEG), polypropylene) (Including glycols (PPG), pentaerythritol, etc.), polyhydric alcohols such as glycerol and mannitol, and any of a number of known solubility enhancers, including other drugs. Other highly polar water-miscible organic materials include dimethyl sulfoxide (DMSO), dimethylformamide (DMF) and NMP (N-methyl-2-pyrrolidone).

  Thus, solvents such as those commonly used as hydrophilic, polar or organic solubilizers provided herein include, for example, Catalan et al. (Eg, 1995 Liebigs Ann. 241; Handbook of Solvent, Wypych (Ed.), May also be selected based on the solvent polarity / polarizability (SPP) scale values using the system of Andrew Publ., Catalan in NY, 2001, and references cited therein, For example, those skilled in the art will appreciate that water has an SPP value of 0.962, toluene has an SPP value of 0.655, and 2-propanol has an SPP value of 0.848. A method for measuring the SPP value of a solvent based on UV measurement of 2-N, N-dimethyl-7-nitrofluorene / 2-fluoro-7-nitrofluorene probe / homophoric pair has been described ( Catalan et al., 1995).

  Solvents with the desired SPP value based on the solubility characteristics of the particular BT composition (either as a pure single component solvent or a solvent mixture of two, three, four or more solvents; solvent miscibility Regarding gender, see, for example, Godfrey 1972 Chem. Technol. According to certain preferred embodiments relating to the method steps, hydrophilic, polar or organic solubilizers are not required to dissolve bismuth.

  Solubility parameters may include interaction parameter C, Hildebrand solubility parameter d, or partial (Hansen) solubility parameters: δp, δh, and δd, representing solvent polarity, hydrogen bonding ability, and dispersive force interaction ability, respectively. . In certain embodiments, the highest solubility parameter describing the solvent or co-solvent system in which the bismuth salt comprising bismuth dissolves is obtained, for example, by the method of preparing the particulate BT composition described herein. Limits of aqueous solution containing may be provided. For example, a higher δh value has a greater hydrogen bonding capacity and thus a greater affinity for solvent molecules such as water. Therefore, in situations where a more hydrophilic environment is desired, it may be preferred that the observed solvent maximum δh be a higher value.

By way of non-limiting example, BisEDT having the structure shown in Formula I below may be prepared by the following reaction scheme:

Briefly, by way of non-limiting example, an excess of 5% aqueous HNO ₃ at room temperature (11.4 L) is added to a Bi (NO ₃ ) ₃ solution (eg, 43% Bi (NO ₃ ) ₃ (w / w ) While stirring 0.331 L (about 0.575 mol) of an aqueous acidic bismuth solution such as 5% nitric acid (w / w), 52% water (w / w), obtained from Shepherd Chemical Co., Cincinnati, OH). It may be added slowly and absolute ethanol (4 L) may be added slowly. Separately, 72.19 mL (0.863 mol) of 1,2-ethanedithiol was added to 1.5 L of absolute ethanol using a 60 mL syringe, followed by stirring for 5 minutes, thereby allowing 1,2-ethanedithiol [˜0. An ethanolic solution (1.56 L) of a thiol compound such as .55M] may be prepared. 1,2-ethanedithiol (CAS 540-63-6) and other thiol compounds are described, for example, in Sigma-Aldrich, St. Available from Louis, MO. Thereafter, an ethanolic solution of the thiol compound may be slowly added to the aqueous Bi (NO ₃ ) ₃ / HNO ₃ solution and stirred overnight to form a reaction solution. The thiol-containing compound may be present in the reaction solution in a molar ratio of about 1: 3 to about 3: 1 with respect to bismuth, according to certain preferred embodiments. The product formed is precipitated as a precipitate containing the particulates described herein, then collected by filtration and subsequently washed with ethanol, water and acetone to give BisEDT as a yellow amorphous powdered solid. obtain. The crude product may be redissolved in absolute alcohol with stirring, then filtered and subsequently washed several times with ethanol and then several times with acetone. The washed powder may be triturated in 1M NaOH (500 mL), filtered and subsequently washed with water, ethanol and acetone to obtain purified microparticle BisEDT.

  According to non-limiting theory, bismuth inhibits the ability of bacteria to produce extracellular polymeric substances (EPS), such as extracellular polysaccharides, which leads to a loss of biofilm formation. Bacteria are thought to use glue-like EPS for biofilm adhesion. Depending on the nature of the infectious agent, biofilm formation and EPS assimilation may contribute to bacterial pathogenicity such as interfering with wound healing. However, bismuth alone is not therapeutically useful as an intervention agent and is instead typically administered as part of a complex such as BT. Thus, bismuth-thiol (BT) is a family of compositions comprising compounds obtained from chelation of bismuth with thiol compounds and exhibiting dramatic improvements in the antimicrobial therapeutic efficacy of bismuth. BT exhibits a pronounced anti-infective effect, anti-biofilm effect, and immune modulating effect. Bismuth-thiol is effective against a wide range of microorganisms and is typically not affected by antibiotic resistance. BT can prevent biofilm formation at significantly lower (sub-inhibitory) concentrations, prevent many pathogenic features of common wound pathogens at the same sub-inhibitory level, and prevent septic shock in animal models; Can be synergistic with many antibiotics.

  As described herein, such synergy in the antibacterial effect of one or more specific BTs when combined with one or more specific antibiotic compounds is a distinct antibiotic for a particular bacterial type. And a BT effect profile that cannot be predicted immediately, but surprisingly, certain specific bacterial populations, such as confirming the presence of Gram negative bacteria or Gram positive bacteria (or both), are considered. It may be obtained by selecting a BT-antibiotic / combination. For example, as disclosed herein, antibiotics that synergize with a particular BT include amikacin, ampicillin, aztreonam, cefazolin, cefepime, chloramphenicol, ciprofloxacin, clindamycin (or other lincosamide antibiotics) Substance), daptomycin (Cubicin®), doxycycline, gatifloxacin, gentamicin, imipenim, levofloxacin, linezolid (Zyvox®), minocycline, nafcillin, paromomycin, rifampin, sulfamethoxazole, tetracycline, bramycin And one or more of vancomycin may be included. For example, MRSA, which is individually or not at all sensitive to gentamicin, cefazoline, cefepime, sfamethoxazole, imipenim or levofloxacin, is exposed to these antibiotics in the presence of the BT compound BisEDT. It has been shown in an in vitro test that it exhibits a marked sensitivity to one or the other. Thus, specific embodiments contemplated herein include BT compounds and amikacin, ampicillin, cefazolin, cefepime, chloramphenicol, ciprofloxacin, clindamycin (or other lincosamide antibiotics), daptomycin (Cubicin®), doxycycline, gatifloxacin, gentamicin, imipenim, levofloxacin, linezolid (Zyvox®), minocycline, nafcillin, paromomycin, rifampine, sulfamethoxazole, tobramycin and vancomycin Explicitly contemplated compositions and / or methods that may include combinations with one or more antibiotics, other specific embodiments contemplated herein include BT compounds, amikacin, Picillin, aztreonam, cefazolin, cefepime, chloramphenicol, ciprofloxacin, clindamycin (or other lincosamides), daptomycin (Cubicin®), doxycycline, gatifloxacin, gentamicin, imipenim, levofloxacin, One or more antibiotics from which one or more antibiotics selected from linezolid (Zyvox®), minocycline, nafcillin, paromomycin, rifampine, sulfamethoxazole, tobramycin and vancomycin can be explicitly excluded; Compositions and / or methods that may include combinations of these are contemplated. It should be noted herein that gentamicin and tobramycin belong to the antibiotic aminoglycosides. Also explicitly excluded from certain contemplated embodiments are Domenico et al. , 2001 Agent. Chemother. 45 (5): 1417-1421; Domenico et al. 2000 Infect. Med. 17: 123-127; Antimicrob. Agents & Chemother. 3: 79-85 in Domenico et al. , 2003 Res. Adv. Domenico et al. , 1997 Antimicrob. Agents Chemother. 41 (8): 1697-1703; Domenico et al. 1999 Infect. Immun. 67: 664-669; Huang et al. 1999 J Antimicrob. Chemother. 44: 601-605; Veloirah et al. , 2003 J Antimicrob. Chemother. 52: 915-919; Wu et al. , 2002 Am J Respir Cell Mol Biol. 26: 731-738; Halwani et al. , 2008 Int. J Pharmaceut. 358: 278; Halwani et al. , 2009 Int. J. et al. Pharmaceut. It is noted that none of these publications teach or suggest the monodisperse particulate BT composition disclosed herein. I want.

  Thus, as described herein, in certain preferred embodiments, the microparticles BT described herein are included, and optionally in other specific embodiments, synergistic and / or potentiating antibiotics are also included. Compositions and methods for treating a subject are provided. Those of ordinary skill in the relevant arts will be aware of appropriate clinical aspects and situations in which such treatment may be desirable based on the disclosure of the present invention, the criteria of which are, for example, surgical, military surgical, among others. Dermatological, traumatic, gerontological, cardiovascular, metabolic disease (eg, diabetes, obesity, etc.), infection and inflammation (respiratory or gastrointestinal lining, or glandular tissue, etc. As well as related medical specialties and sub-specialties.

  Thus, in certain embodiments disclosed herein and known in the art, promoting skin tissue repair (or repair of other tissues, such as epithelial tissue, bone, joints, muscles, tendons, or ligaments). Is contemplated. In certain embodiments, the promotion of skin tissue or other epithelial tissue repair comprises (i) migration of epithelial cells (eg, keratinocytes) or dermal fibroblasts, (ii) epithelial cells (eg, keratinocytes). Or development of dermal fibroblasts, (iii) downregulation of collagenase, gelatinase or matrix metalloproteinase activity in epithelial cells (eg keratinocytes) or dermal fibroblasts, (iv) extracellular matrix of dermal fibroblasts Stimulating or derepressing one or more intracellular wound repair activities selected from protein deposition and (v) induction or enhancement of dermal angiogenesis. Since a methodology for identifying and characterizing such intracellular wound repair activity has been described, it includes a wound tissue repair-promoting compound disclosed herein, eg, a BT agent described herein. The effects of the compositions on these and related activities can be determined immediately and without undue experimentation based on the present disclosure. For example, disclosed herein are compositions and methods relating to an art accepted model for wound repair based on keratinocyte wound closure following a scratch.

  Preferred compositions for treating microbial infections on or within natural surfaces used according to embodiments described herein, in certain embodiments, include bismuth-thiol (BT) compounds described herein. Which may comprise other compounds known in the art, such as one or more antibiotic compounds described herein, in certain different related embodiments. Also good. BT compounds and methods for making them are disclosed herein, see, for example, Domenico et al. (1997 Antimicrob. Agent. Chemother. 41 (8): 1697-1703; 2001 Antimicrob. Agent. Chemother. 45 (5): 1417-1421) and U.S. Pat. S RE 37,793, U.S. Pat. S. 6,248,371, 6,086,921, and 6,380,248. As also mentioned earlier, certain preferred BT compounds include those that are ionically bound to a thiol-containing compound or that contain bismuth or a bismuth salt in a coordination complex therewith, such as bismuth chelated to a thiol-containing compound. Other particularly preferred BT compounds are those containing bismuth or a bismuth salt in a covalent linkage to a thiol-containing compound. Also preferred are the substantially monodispersed particulate BT compositions described herein. Any of the compounds described herein for the first time having use in compositions and methods for promoting past treatments of bacterial infections, or natural surfaces described herein. The characterization in other aspects also did not predict that the methods of the invention using such compounds would have the beneficial effects described herein.

Thus, according to a preferred embodiment, there is provided a method of treating a natural surface comprising administering at least one particulate BT compound described herein to the natural surface. In certain embodiments, the method may be a synergistic antibiotic described herein in certain preferred embodiments, and may be described herein in other particular embodiments. The method further includes administering at least one antibiotic compound, which may be a potent antibiotic, simultaneously or sequentially in either order. Antibiotic compounds include aminoglycoside antibiotics, carbapenem antibiotics, cephalosporin antibiotics, fluoroquinolone antibiotics, glycopeptide antibiotics, lincosamide antibiotics, penicillinase-resistant penicillin antibiotics, or aminopenicillin antibiotics It may be an antibiotic. Clinically useful antibiotics are discussed elsewhere herein, for example, Washington University School of Medicine, The Washington Manual of Medical Therapeutics (32 nd Ed.), 2007 Lippincott, Williams and Wilkins, Also described in Philadelphia, PA; Hauser, AL, Antibiotic Basics for Clinicians, 2007 Lippincott, Williams and Wilkins, Philadelphia, PA.

  As described herein, certain embodiments show that in the case of bacterial infections, including gram positive bacteria, the preferred therapeutically effective combination is a BT compound (eg, BisEDT, bismuth; 1,2- Ethanedithiol; BisPyr, bismuth: pyrithione; BisEDT / Pyr, bismuth: 1,2-ethanedithiol / pyrithione) and rifamycin, or BT compound and daptomycin (Cubicin®, Cubist Pharmaceuticals, Lxinton, MA), or BT Compound and linezolid (Zyvox®, Pfizer, Inc., NY, NY), or BT compound (eg, BisEDT, bismuth; 1,2-ethanedithiol; BisPyr, bismuth: pyrithione; BisEDT / P r, bismuth: 1,2-ethanedithiol / pyrithione) and ampicillin, cefazolin, cefepime, chloramphenicol, clindamycin (or other lincosamide antibiotics), daptomycin (Cubicin®), doxycycline, gatiflo The unexpected discovery that it may include one or more of xacin, gentamicin, imipenim, levofloxacin, linezolid (Zyvox®), nafcillin, paromomycin, rifampine, sulfamethoxazole, tobramycin and vancomycin Derived from.

  As also described herein, certain embodiments are unexpected in that in the case of bacterial infections including gram-negative bacteria, a preferred therapeutically effective combination can include a BT compound and amikacin. Derived from discovery. Certain related embodiments contemplate treating an infectious agent comprising a Gram-negative bacterium with a BT compound and another antibiotic, such as another aminoglycoside antibiotic that is not gentamicin or tobramycin in certain embodiments. Thus, in view of these embodiments, other related embodiments are described in the medical methodology arts as a step of selecting an appropriate antibiotic compound to be included in a combination administered by a method of the invention. By methodology well known to those skilled in the art, it is contemplated to identify one or more bacterial populations or subpopulations within or on a natural surface by well-known criteria that are Gram positive or Gram negative.

  The compositions and methods described herein typically involve the compounds described herein (eg, one or more microparticles BT alone, or one or more synergisticities disclosed herein). And / or in combination with potentiating antibiotics) by applying or administering to microbial sites such as the presence of microorganisms on or within natural surfaces, such as microorganisms (eg bacteria, viruses, yeasts, molds and Applications may be found in the treatment of other fungi, microbial parasites, etc.). Such natural surfaces include, but are not limited to, mammalian tissues (eg, epithelium such as skin, scalp, gastrointestinal lining, oral cavity; endothelium, cells and tissue membranes such as peritoneum, pericardium, capsule, periosteum, marrow Cornea, sclera, mucous membrane, etc .; and other mammalian tissues such as teeth, bones, joints, tendons, ligaments, muscles, heart, lung, kidney, liver, spleen, gallbladder, pancreas, Bladder, nerve, etc.).

  The microparticulate antimicrobial agent described herein inhibits microbial growth, reduces microbial infestation, reduces biofilm, prevents bacterial to biofilm conversion, microbial infection May be used to prevent or inhibit and for any other use described herein. These agents have multiple antiviral purposes, including the prevention or inhibition of viral infection by herpes viruses such as cytomegalovirus, herpes simplex type 1 and herpes simplex type 2 and / or infection by other viruses. Also useful. In this context, the drugs are various viruses such as single stranded RNA viruses, single stranded DNA viruses, Rous sarcoma virus (RSA), hepatitis A virus, hepatitis B virus (HBV), hepatitis C. Virus (HCV), influenza virus, West Nile virus (WNV), Epstein-Barr virus (EBV), Eastern equine encephalitis virus (EEEV), severe acute respiratory virus (SARS), human immunodeficiency virus (HIV), human papilloma It is useful for the prevention or inhibition of viral infection by viruses (HPV) and human T cell lymphoma virus (HTLV).

  Other internal and external pharmaceutical uses of the antimicrobial agents described herein include, but are not limited to, bacterial infections, tuberculosis, fungal infections such as yeast and mold infections (eg, Candida (eg, Candida Treatment of genus (eg Candida albicans, Candida glabrata, C. parapsilosis, C. tropicalis, and C. dubriniensis) or Cryptococcus or other fungi), Helicobacter pylori infection, and peptic ulcer disease or In one embodiment, the agent is at a dosage that is generally not lethal to bacteria but is still sufficient to reduce a protective polysaccharide coating that otherwise resists the natural immune response. Thus, this technology is used for human symbiotic microorganisms (eg normal gut microbiota) Without damage to the extent that can be a case of using antibiotics is believed to assist in eradication of bacterial infection through the immune system.

  Illustrative, but not limiting, specific contemplated embodiments are described herein.

  In certain embodiments, the particulate BT compounds (or compositions comprising particulate BT compounds) described herein can be used to control biofilm generation, disrupt biofilm, or reduce the amount of biofilm. In combination with at least one or more anti-biofilm agents. As understood in the art, interspecies quorum sensing is involved in biofilm formation. Certain agents that increase the LuxS-dependent pathway or interspecies quorum sensing signal (see, eg, US Pat. No. 7,427,408) contribute to the control of biofilm development and / or proliferation. Exemplary agents include, for example, combined or separate N- (3-oxododecanoyl) -L-homoserine lactone (OdDHL) blocking compounds and N-butyryl-L-homoserine lactone (BHL) analogs. (See, eg, US Pat. No. 6,455,031). An oral hygiene composition comprising a particulate BT compound and at least one anti-biofilm agent can be delivered locally for disintegration and inhibition of bacterial biofilms and for the treatment of periodontal disease (eg, the United States No. 6,726,898).

Composition comprising particulate bismuth-thiol and use for oral hygiene and to treat oral inflammation and infection In another embodiment, a composition comprising particulate BT compound is formulated for oral use, It may be used in methods for preventing or reducing microbial growth in the mouth and for preventing and / or treating oral microbial infections and inflammation. Therefore, these compositions are used to prevent or treat plaque, bad breath, periodontal disease, gingivitis, and other infections of the mouth (ie, reduce or inhibit their onset, and develop or reoccurrence). Useful for reducing likelihood). An oral composition comprising a particulate BT compound prevents and / or controls the development of biofilm (ie slows, delays, inhibits), disrupts biofilm, or the oral surface, in particular teeth or It can also be useful to reduce the amount of biofilm present on the gums.

  Entrained food particles, poor oral hygiene and poor oral health, and inadequate cleaning of complete dentures can promote microbial growth on teeth, gum periphery, and tongue. The continued growth of microorganisms and the presence of caries can lead to bad breath, plaque (ie, a biofilm formed by microbial colonization), gingivitis, and inflammation. If proper oral care (eg, brushing, flossing) is not performed, more severe infections such as periodontal disease and jaw infection can result.

  Good oral hygiene is important not only for oral health, but also for the prevention of multiple chronic conditions. Controlling oral bacterial growth can help reduce the risk of heart disease, maintain memory, and reduce the risk of infection and inflammation in other areas of the body. People with diabetes have a high risk of developing severe gum problems, and reducing the risk of gingivitis by maintaining good oral health can help control blood sugar levels. Pregnant women are more likely to suffer from gingivitis, and several studies have suggested an association between gum disease in pregnant women and the delivery of preterm and low birth weight infants.

  Bacteria are a major etiological agent in periodontal disease. More than 500 strains may be found in plaque (Kroes et al., Proc. Natl. Acad. Sci. USA 96: 14547-52 (1999)). Bacteria have evolved to survive as biofilms on the tooth surface, gingival epithelium, and oral environment, which makes it difficult to treat periodontitis. Bactericides and antibiotics currently used to treat such infections often do not kill all of the problematic organisms. Use of a drug that is ineffective against a particular strain may result in the growth of resistant strains. In addition, these agents can induce undesirable side effects, such allergic reactions, inflammation, and tooth discoloration.

  Dental bacterial plaque is a biofilm that adheres stubbornly to tooth surfaces, restorations, and prostheses. The main means of controlling the biofilm in the mouth is by mechanical cleaning (ie tooth brushing, flossing, etc.). In the absence of such cleaning, within 2 days thereafter, the tooth surface is mainly colored by facultative Gram-positive cocci, which are mainly Streptococcus. Bacteria secrete an extracellular mucus layer that helps fix bacteria to the surface and protect the attached bacteria. When the tooth surface is covered with attached bacteria, microcolony formation begins. Biofilms grow primarily through adherent bacterial cell division rather than the attachment of new bacteria. The doubling time of plaque formed by bacteria is rapid during early development and is slower in more mature biofilms.

  Co-aggregation occurs when engraftment continues to adhere to bacteria already attached to the membrane. As a result of co-aggregation, a complex array of different bacteria connected to each other is formed. A few days after plaque formation without interruption, the gingival margin becomes inflamed and swollen. Inflammation may form deep gingival crevice. The biofilm expands into this subgingival area and grows in this protected environment, forming a mature subgingival plaque biofilm. Gingival inflammation is not observed until the biofilm is changed from one consisting mostly of gram positive bacteria to one containing gram negative anaerobic bacteria. Subgingival bacterial microcolonies composed mainly of gram-negative anaerobic bacteria become established in the gingival crevice between 3-12 weeks after plaque formation on the gingival margin begins. Currently, most bacterial species suspected of periodontal pathogens are anaerobic gram-negative bacteria.

  Bacterial microcolonies protected within the biofilm are typically resistant to antibiotics (systemic administration), disinfectants or bactericides (local administration), and immune defenses. For example, the antibiotic dose that kills airborne bacteria needs to be increased by 1500 times the dose that kills biofilm bacteria. At this high concentration, these antimicrobial agents tend to be toxic to patients (see, eg, Coghlan 1996, New Scientist 2045: 32-6; Elder et al., 1995, Eye 9: 102-9). .

  Careful and frequent physical removal of bacterial plaque biofilm is the most effective means of eliminating and controlling plaque. However, subgingival plaque in the pocket cannot be reached by brush, floss, or mouth rinse. Therefore, frequent periodontal debridement of the subgingival root surface by a dental hygienist or dentist is an essential element in the prevention and treatment of periodontitis.

  In certain embodiments, the particulate BT compound is a mouth hygiene composition that can be routinely used by any subject, such as, but not limited to, toothpaste, mouthwash (ie, mouth rinse), oral gel, toothpaste, oral It may be incorporated into a spray (such as a spray dispersed by a mouth inhaler), an edible film, a chewing gum, a mouth slurry, a complete denture cleaning solution, a complete denture preservation solution, and dental floss. The particulate BT compounds may be incorporated into oral hygiene compositions used primarily by dental care professionals, including, for example, liquid fluorine treatments, cleaning compositions, buffing compositions, oral rinses, and dental floss. Embodiments of the present invention replace antimicrobial agents formulated with oral hygiene compositions described in the art with the particulate BT compounds described herein to provide antimicrobial activity, solubility and biological utilization. It is intended to provide the advantages disclosed herein, including a range of degrees, anti-biofilm effects, non-toxicity, enhanced antibiotic efficacy, as well as other properties described herein.

  The particulate BT compound is used to prevent or treat caries and / or inflammation by administering the particulate BT compound to the tooth surface (ie, to reduce the likelihood of occurrence or recurrence of caries and / or inflammation, respectively). May be used). The composition comprising the particulate BT compound may be a mucoadhesive composition applied to the surface of the teeth and / or gums or oral mucosa, any form that adheres to the surface to some extent, or a pharmaceutically effective amount. In any form that delivers the active ingredient to the desired surface. The particulate BT compound can also be formulated so as to be slowly released from the composition applied to the teeth. For example, the composition may be a gel (eg, a hydrogel, thiomer, aerogel, or organogel) or a liquid. The organogel may include an organic solvent, lipoic acid, vegetable oil, or mineral oil. Such gel or liquid coating formulations may be applied to the interior or exterior of an amalgam or composite or other restorative composition. The sustained release composition provides a pharmaceutically effective amount of the particulate BT compound for 1, 2, 3, 4, 5, 6, or 7 days (1 week), or 2, 3, 4, 5, 6, 7 It may be delivered weekly or for 1, 2, 3, 4, 5, or 6 months. Such compositions can be prepared by one skilled in the art using any number of methods known in the art.

  In other specific embodiments, as described herein, an antimicrobial composition comprising a particulate BT compound and one or more additional antimicrobial compounds or agents is provided for oral use. Particularly useful are compositions comprising a second antimicrobial agent that when administered in combination have a potent or synergistic antimicrobial effect as described herein. As an example, an enhanced antimicrobial effect may be observed when a particulate BT compound is administered with an antimicrobial agent that chelates iron. In other specific embodiments, the particulate BT compound is formulated with an anti-inflammatory agent, compound, small molecule, or macromolecule (eg, a peptide or polypeptide).

  Any of the particulate BT compounds described herein may be formulated for oral use. In certain embodiments, a particulate BT compound prepared with a hydrophobic thiol (eg, thiochlorophenol) may be used, which has the ability to adhere to tooth and mouth tissues more than a less hydrophobic BT compound. May be shown greatly. BT compounds having a net negative charge, such as those having a molar ratio of 1: 2 (bismuth to thiol), may have suitable adhesion.

  The oral hygiene composition comprising the particulate BT compound may further comprise one or more active ingredients and / or one or more orally suitable excipients or carriers. In one embodiment, the oral hygiene composition may further comprise baking soda or other alkaline compound or substance. Due to the chemical and physical properties of baking soda, it has a wide range of applications including washing, deodorizing, and buffering. Baking soda chemically neutralizes rather than masks or absorbs odors. Baking soda can be combined with the particulate BT compound as a powder mixture, or can be dissolved or suspended in any one of the toothpastes, gels, pastes, and liquids described herein. In other embodiments, the particulate BT compound is combined with other alkali metal bicarbonate or carbonate materials (eg, potassium bicarbonate or calcium carbonate) that help maintain the desired alkaline pH and also have cleaning and deodorizing properties. be able to.

The oral hygiene composition containing the particulate BT compound may further contain one or more of the following components.
Antimicrobial agents: eg chlorhexidine: sanguinarine extract, metronidazole, quaternary ammonium compounds (eg cetylpyridinium chloride); bisguanides (eg chlorhexidine digluconate, hexetidine, octenidine, alexidine); halogenated bisphenol compounds (eg , 2,2'-methylenebis- (4-chloro-6-bromophenol) or other phenolic antibacterial compounds; alkyl hydroxybenzoates; cationic antimicrobial peptides; aminoglycosides; quinolones; lincosamides; Tetracycline; other antibiotics known in the art; Coleus forskori essential oil; silver or silver colloid antimicrobial agent; tin or copper based Antimicrobial agents; manuka oil; oregano; thyme; rosemary; or other herbal extracts; and grapefruit seed extract.
Anti-inflammatory or antioxidant: for example, ibuprofen, flurbiprofen, aspirin, indomethacin, aloe vera, turmeric, olive leaf extract, clove, panthenol, retinol, ω-3 fatty acid, γ-linolenic acid (GLA), green tea , Ginger, grape seed, etc. Caries prevention agents: for example, sodium fluoride and stannous fluoride, amine fluoride, sodium monofluorophosphate, sodium trimetaphosphate, zinc citrate or other zinc agents, and casein. Plaque buffer: for example, urea, calcium lactate, calcium glycerophosphate, and strontium polyacrylate. Vitamins: For example, vitamins A, C and E. Plant extract. Desensitisers: For example, potassium citrate, potassium chloride, potassium tartrate, potassium bicarbonate, potassium oxalate, potassium nitrate, and strontium salts. Anticalculus agents: for example, alkali metal pyrophosphates, hypophosphite-containing polymers, organic phosphonates and phosphocitrates. Biomolecule: For example, bacteriocin, bacteriophage, antibody, enzyme and the like. Flavoring agents: for example, peppermint and spearmint oil, fennel, cinnamon. Proteinaceous material: for example, collagen. Preservative. Opacifier. Coloring agent. pH adjuster. Sweetener. Pharmaceutically acceptable carrier: for example starch, sucrose, water or water / alcohol system. Surfactants: For example, anionic, nonionic, cationic and zwitterionic or amphoteric surfactants, saponins from plant materials (see, eg, US Pat. No. 6,485,711). Particle abrasive: for example, silica, alumina, calcium carbonate, dicalcium phosphate, calcium pyrophosphate, hydroxyapatite, trimetaphosphate, insoluble hexametaphosphate, agglomerated particle abrasive, chalk, finely pulverized natural chalk and the like. Hygroscopic agent: for example, glycerol, sorbitol, propylene glycol, xylitol, lactitol and the like. Binders and thickeners: for example sodium carboxymethylcellulose, hydroxyethylcellulose (Natrosol®), xanthan gum, gum arabic, synthetic polymers (for example polyacrylates and carboxyvinyl polymers such as Carbopol®). Polymeric compounds that improve the delivery of active ingredients such as antimicrobial agents. Buffers and salts that buffer the pH and ionic strength of the oral care composition. Bleach: For example, peroxy compounds (eg, potassium peroxydiphosphate). Firing system: for example, sodium bicarbonate / citric acid system. Discoloration system. In certain embodiments, the abrasive is silica or finely ground natural chalk.

  An oral hygiene composition comprising a particulate BT compound formulated for use as a toothpaste further comprises a hygroscopic agent (eg, glycerol or sorbitol), a surfactant, a binder, and / or a flavoring agent. Also good. Toothpaste may contain sweeteners, whitening agents, preservatives, and antimicrobial agents. The pH of the composition for toothpaste and other oral uses is typically between pH 5.5 and 8.5. In certain embodiments, the oral hygiene composition, including toothpaste, has a pH of 7-7.5, 7.5-8, 8-8.5, or 8.5-9, and the particulate BT The antimicrobial activity of the compound can be enhanced. The toothpaste compositions described herein include chalk, dicalcium phosphate dihydrate, sorbitol, water, hydrated aluminum oxide, precipitated silica, sodium lauryl sulfate, sodium carboxymethylcellulose, flavoring agent, monoolein Acid sorbitan, sodium saccharin, tetrasodium pyrophosphate, methyl paraben, propyl paraben. If desired, one or more colorants such as FD & C Blue can be used. Other suitable ingredients that may be included in the toothpaste formulation are described in the art, for example, in US Pat. No. 5,560,517.

  In one particular embodiment, the oral hygiene composition is a mouse spray and includes a particulate BT compound, an alkaline buffer (eg, potassium bicarbonate), an alcohol, a sweetening ingredient, and a flavoring system. The flavoring system may have one or more of the following: flavoring agents, hygroscopic agents, surfactants, sweeteners, and colorants (see, eg, US Pat. No. 6,579,513). ). The surfactants described herein and known in the art for use in oral hygiene compositions may be anionic, nonionic or amphoteric.

  In another embodiment, particulate BT-containing oral hygiene compositions have been described in the art as being useful for inclusion in toothpastes, toothpaste gels, mouthwashes, etc. to treat severe infections. It may be combined with further active ingredients such as taurolidine and taurultam (see, for example, British Patent Application GB 1557163, US Pat. No. 6,488,912). As described herein, microparticles BT can also be combined with one or more additional antimicrobial agents, when combined with microparticles BT, the combination has an additive or synergistic effect.

  In yet another specific embodiment, the oral hygiene composition described herein is at least one for controlling biofilm generation, for disrupting biofilm, or for reducing the amount of biofilm. Further or more anti-biofilm agents may be included. As understood in the art, interspecies quorum sensing is involved in biofilm formation. Certain agents that increase the LuxS-dependent pathway or interspecies quorum sensing signal (see, eg, US Pat. No. 7,427,408) contribute to the control of biofilm development and / or proliferation. Exemplary agents include, for example, combined or separate N- (3-oxododecanoyl) -L-homoserine lactone (OdDHL) blocking compounds and N-butyryl-L-homoserine lactone (BHL) analogs. (See, eg, US Pat. No. 6,455,031). An oral hygiene composition comprising a particulate BT compound and at least one anti-biofilm agent can be delivered topically for disintegration and inhibition of bacterial biofilm and treatment of periodontal disease (eg, US, for example) No. 6,726,898).

  The oral hygiene compositions described herein may contain a sufficient amount of particulate BT compound to perform substantial antimicrobial action at the time required for normal tooth brushing, mouth rinsing, or flossing. Good. As described herein, the particulate BT compound may be retained on the oral surface (eg, teeth, amalgam, composites, mucous membranes, gums). For example, particulate BT compounds retained on teeth and gums after brushing, rinsing, and flossing may continue to provide long-term anti-biofilm and anti-inflammatory effects.

  In another embodiment, the particulate BT compound is slowly released from the mucoadhesive polymer or other agent that contributes to the retention of the particulate BT compound on the mucosa or tooth surface. The particulate BT compound may be added to a stable viscous mucoadhesive aqueous composition for the prevention and treatment of mucosal ulcerous, inflammatory and / or erosive disorders and / or topical treatment. It may be used for delivery of pharmaceutically active compounds to the mucosal surface or transfer to the systemic circulation (see, eg, US Pat. No. 7,547,433).

  In another embodiment, the oral hygiene composition comprising the particulate BT compound further comprises olive oil that can enhance plaque removal. The use of olive oil in products intended for oral hygiene, such as toothpastes, mouthwashes, sprays, oral inhalers, or chewing gum, eliminates or reduces (decreases) bacterial plaque and / or Contributes to the elimination or reduction (reduction) of the number of bacteria present, thereby achieving a reduction in the incidence of dental diseases (eg, caries, periodontal disease) and bad breath (eg, US Pat. No. 7,074) 391).

  In other embodiments, the oral hygiene composition comprising the particulate BT compound may further comprise a mucosal bactericidal preparation for topical application in the mouth. The oral hygiene composition may further comprise an aqueous slurry useful for washing the tongue and throat (eg, US Pat. No. 6,861,049). In yet another embodiment, the oral hygiene composition comprising the particulate BT compound is at least used to prevent the formation of cavities (ie, reduce the likelihood of occurrence) or reduce the number of cavities. One mint may further be included. One such mint called CaviStat® (Ortek Therapeutics, Inc., Roslyn Heights, NY) contains arginine and calcium, helps neutralize acidic pH, and adheres to calcium on enamel surfaces Promote. Inclusion of mint in the oral hygiene composition containing the particulate BT compound can thus increase the pH and enhance the adhesion of the particulate BT compound to the oral surface.

Compositions comprising particulate bismuth-thiol formulated for orthopedic use In certain embodiments, orthopedic procedures (eg, orthopedic surgery, orthopedic treatment, arthroplasty (including two-stage arthroplasty) ), A method of using a composition comprising a particulate BT compound for preventing and / or treating microbial infection and inflammation caused by orthodontic treatment). Compositions comprising the particulate BT compounds described herein therefore prevent and / or treat microbial infections of the skeletal and supporting structures (ie, bones, joints, muscles, ligaments, tendons) such as osteomyelitis (Ie, reduce or inhibit its onset, reduce the likelihood of its occurrence or recurrence). A composition described herein comprising a particulate BT compound prevents and / or controls biofilm development (ie slows, delays, inhibits), disrupts biofilm, or within a joint, or It can also be useful to reduce the amount of biofilm present on the surface of bones, ligaments, tendons or teeth.

  The compositions described herein for orthopedic use comprising a particulate BT compound may further comprise one or more additional antimicrobial compounds or agents. Particularly useful are compositions comprising a particulate BT compound and a second antimicrobial agent that when administered in combination have a potent or synergistic antimicrobial effect as described herein. As an additional example, an enhanced antimicrobial effect can be observed when the particulate BT compound is administered with an antimicrobial agent that chelates iron. In other specific embodiments, the particulate BT compound is formulated with an anti-inflammatory agent, compound, small molecule, or macromolecule (eg, a peptide or polypeptide).

  A composition comprising a particulate BT compound has an enhanced or synergistic antimicrobial effect (ie, greater than an additive effect) when administered in combination with at least one other antimicrobial agent (ie, second, You may combine with the antimicrobial agent of 3rd, 4th etc.). As an example, an enhanced microbial effect can be observed when a particulate BT compound is administered with an antimicrobial agent that chelates iron. In certain embodiments, the composition comprising the particulate BT compound may be combined with at least one other antimicrobial and / or anti-inflammatory agent selected from: antimicrobial agents: for example, chlorhexidine : Sanguinarine extract, metronidazole, quaternary ammonium compounds (for example, cetylpyridinium chloride); bisguanides (for example, chlorhexidine digluconate, hexetidine, octenidine, alexidine); halogenated bisphenol compounds (for example, 2,2′-methylenebis) -(4-Chloro-6-bromophenol) or other phenolic antibacterial compounds; alkyl hydroxybenzoates; cationic antimicrobial peptides; aminoglycosides; quinolones; lincosamides; penicillins: cephalosporins; Tetracycline; other antibiotics known in the art; Coleus-Forscoli essential oil; silver or silver colloid antimicrobial agent; antimicrobial agent based on tin or copper; manuka oil; oregano; thyme; Or grapefruit seed extract Anti-inflammatory or antioxidant: eg ibuprofen, flurbiprofen, aspirin, indomethacin, aloe vera, turmeric, olive leaf extract, clove, panthenol, retinol, ω- 3 fatty acids, γ-linolenic acid (GLA), green tea, ginger, grape seed, etc. In certain embodiments, the composition comprising particulate BT compound comprises clindamycin, vancomycin, daptomycin, cefazolin, gentamicin, tobu Antibiotics selected from mycin, metronidazole, cefaclor, ciprofloxacin, or other antimicrobial agents such as quaternary ammonium compounds (eg benzalkonium chloride, cetylpyridinium chloride), antimicrobial zeolites, alkalis The composition may further comprise a metal hydroxide, or an alkaline earth metal oxide, the composition comprising one or more pharmaceutically suitable carriers (ie, excipients) as described herein, Surfactants, buffers, diluents, and salts, and optionally bleaching agents may optionally be included, and these and certain of the related embodiments disclosed herein may include one or more microparticles. BT may be included, optionally further including antibiotics such as synergistic or potentiating antibiotics described herein. Particulate BT compositions disclosed herein contemplates the inclusion and processes to such products.

Biological, Biomedical and Other Uses of Particulate BT Other specific embodiments are BT alone or bismuth moieties substituted with different group V metals such as antimony (Sb) or arsenic (As) BT, and / or such BT in combination with one or more antibiotics that exhibit synergistic or potentiated antimicrobial activity in nutritional formulas where BT is orally ingested as described herein. Any of the following is contemplated for use of the particulate BT described herein.

  According to non-limiting theory, with vitamins, minerals, amino acids, carbohydrates containing carbohydrates, fatty acids, oils, phytonutrients, tea, herbs or herbal extracts, and / or other ingredients such as other nutritional or food products The inclusion of particulate BT in such a combination may increase the bioavailability of BT and optionally antibiotics and / or additional nutrients to the host gastrointestinal tract in certain embodiments ( For example, techniques that promote (in a statistically significant manner relative to the appropriate control) can result in blockage or delay of nutrient uptake by the microbial population in the gastrointestinal tract. In other specific embodiments, certain vitamins, minerals, amino acids, carbohydrates containing carbohydrates, fatty acids, oils, phytonutrients, teas, herbs or herbal extracts included in the oral particulate BT (or AsT or SbT) formulation, And / or by changing other nutritional or food products, the bioavailability of the BT and optionally antibiotics and / or additional nutrients to the host gastrointestinal tract can be reduced ( For example, in a statistically significant manner relative to the appropriate control).

  For example, in the presence of a pathological gastrointestinal (GI) tract infection, the BT compound can be maintained so as to retain bioavailability in the GI tract to exert an antimicrobial effect against infectious pathogens. It may be desirable to administer a particulate BT formulation that interferes with intestinal absorption. As those skilled in the art are aware of multiple vitamins, minerals, amino acids, carbohydrates containing carbohydrates, fatty acids, oils, phytonutrients, teas, herbs and / or herbal extracts that promote or interfere with the GI tract absorption of nutrients, Formulations that increase or decrease the presence of one or more components of the GI tract (eg, a particulate BT compound, an antibiotic, or one or more specific nutrients) are disclosed in the particulate BT (or AsT or SbT).

  Other specific embodiments provided herein include oral delivery compositions to reduce stool or digestive gas odors, for example, in patients undergoing colostomy, or the presence of local microorganisms Aside from this, it is contemplated that other topical delivery compositions contain the particulate BT compounds disclosed herein to reduce foot or other body odor. Multiple skin and GI tract microbial populations, including planktonic bacteria and biofilm bacteria, when present with the potentiating or synergistic antibiotics described herein, have the particulate BT described herein. Sensitive to low concentrations of such particulate BT compounds, including compounds.

  Thus, certain embodiments reduce the population of GI or skin resident bacteria in a manner that reduces or reduces the problem of inappropriate odor (eg, statistically significant relative to appropriate controls). In particular, a particulate BT formulation for oral and topical delivery is contemplated. Oral and topical pharmaceutical formulations are described below, and these and related embodiments provide advantages associated with the particulate BT formulations of the present invention, such as compatible bioavailability and solubility characteristics, and Other factors that exhibit low toxicity and can affect the selection of antimicrobial compositions are described elsewhere in this specification, for example, US Pat. S. It can also be found in 6,582,719.

  An exemplary BT compound, BisEDT, was shown to neutralize body odor for 2-3 days when applied to the axilla area in human test subjects (50 μL of a 1 mg / mL solution in DMSO). The BisEDT mixture in talc powder applied to the human test foot substantially reduced leg odor. Experimental mice that were orally ingested 1 mg / kg BisEDT twice daily for 5 days showed a 90% reduction in fecal flora count. Any thiol-containing solution that emits odors, including particulate BT preparations described herein that can be produced using excess bismuth and added to the thiol-containing solution as an odor remover (eg, soot Commonly useful deodorants are also contemplated for fish oils such as The resulting mixture retains the antimicrobial properties of the particulate BT. Other contemplated applications include other biological source oils or solvents such as butter, such as hemp oil, tea tree oil, shea butter, linseed oil, fish oil, and in certain embodiments, Oils that may have independent or synergistic anti-inflammatory and / or pain-reducing properties and / or other beneficial physiological effects.

Pharmaceutical Compositions and Administration Certain embodiments also relate to pharmaceutical compositions containing the particulate BT compounds disclosed herein, and in certain such embodiments, the pharmaceutical composition comprises a BT compound as defined herein. It may further comprise one or more antibiotics, such as those that exhibit a synergistic or potentiating effect as described in the document. In one embodiment, one or more such particulate BT compounds when administered to an animal, preferably a mammal, most preferably a human patient, are pharmaceutically acceptable as disclosed herein. Compositions containing pharmaceutical amounts in a carrier, excipient or diluent are provided.

  Administration of the particulate BT compound or pharmaceutically acceptable salt in pure form or in a suitable pharmaceutical composition should be carried out via any of the accepted modes of administration of drugs suitable for similar uses. Can do. The pharmaceutical composition can be prepared by combining the particulate BT compound with a suitable pharmaceutically acceptable carrier, diluent or excipient, such as tablets, capsules, powders, granules, ointments, solutions, suppositories, Formulations may be formulated in solid, semi-solid, liquid or gaseous forms such as injection solutions, inhalants, gels, microspheres, and aerosols. Typical routes of administering such pharmaceutical compositions include, but are not limited to, oral, topical, transdermal, inhalation, parenteral, sublingual, rectal, vaginal, and intranasal. The term parenteral as used herein includes subcutaneous injections, intravenous, intramuscular, intrathecal injection or infusion methods. The pharmaceutical composition is formulated so that the active ingredient contained is bioavailable when the composition is administered to a patient. A composition to be administered to a subject or patient is in the form of one or more dosage units, for example, a tablet may be a single dosage unit, and a container of compounds in aerosol form may be in multiple dosage units. May be held. Actual methods of preparing such dosage forms are known or will be apparent to those skilled in the art, see, eg, The Science and Practice of Pharmacy, 20th Edition (Philadelphia College of Pharmacy and Science, 2000). I want. The composition to be administered will contain, in any event, a pharmaceutically effective amount of the compound or pharmaceutically acceptable salt thereof for the treatment of the disease or condition of interest according to the teachings herein.

  The pharmaceutical compositions useful herein also contain a pharmaceutically acceptable carrier, including any diluents or excipients, which are themselves detrimental to the individual to whom the composition is administered. Any pharmaceutical agent that does not produce antibodies and can be administered without undue toxicity is included. Pharmaceutically acceptable carriers include, but are not limited to, liquids such as water, saline, glycerol and ethanol. A thorough discussion of pharmaceutically acceptable carriers, diluents, and other excipients is given in REMINGTON'S PHARMACEUTICAL SCIENCES (Mack Pub. Co., NJ, latest edition).

  The pharmaceutical composition may be in solid or liquid form. In one embodiment, the composition is in the form of a tablet or powder, for example, since the carrier is a particle. The carrier can be a liquid and the composition can be, for example, an oral syrup, an injectable solution, or an aerosol useful, for example, for inhalation administration.

  When intended for oral administration, the pharmaceutical composition is preferably either in solid or liquid form, and semi-solid, semi-liquid, suspension and gel forms are herein described as either solid or liquid. Included in the forms considered in the book

  The pharmaceutical composition as a solid composition for oral administration may be formulated in the form of powder, granule, compressed tablet, pill, capsule, chewing gum, wafer, and the like. Such solid compositions typically contain one or more inert diluents or edible carriers. In addition, one or more of the following may be present: binders such as carboxymethylcellulose, ethylcellulose, microcrystalline cellulose, tragacanth gum or gelatin; excipients such as starch, lactose or dextrin; disintegrants such as alginic acid, Lubricants such as magnesium stearate or Sterotex; Lubricants such as colloidal silicon dioxide; Sweeteners such as honey, sucrose or saccharin; Flavoring agents such as peppermint, methyl salicylate or orange flavors; And colorants.

  When the pharmaceutical composition is in the form of a capsule, such as a gelatin capsule, it may contain a liquid carrier, such as polyethylene glycol or oil, in addition to the types of materials described above.

  The pharmaceutical composition may be in liquid form, for example in the form of elixirs, syrups, solutions, emulsions or suspensions. The liquid may be for oral administration or for delivery by injection, by way of example. When intended for oral administration, preferred compositions contain, in addition to the present compounds, one or more of a sweetening agent, preservatives, dye / colorant and flavor enhancer. Compositions intended for administration by injection may contain one or more of surfactants, preservatives, wetting agents, dispersing agents, suspending agents, buffering agents, stabilizing agents and isotonic agents.

  Liquid pharmaceutical compositions may include one or more of the following adjuvants, whether in the form of solutions, suspensions, etc .: sterile diluents such as water for injection, saline solution, Preferably, saline, Ringer's solution, isotonic sodium chloride, non-volatile oils such as synthetic mono- or diglycerides, polyethylene glycol, glycerin, propylene glycol or other solvents that can act as solvents or suspending media; antimicrobial agents such as Benzyl alcohol or methyl paraben; antioxidants such as ascorbic acid or sodium bisulfite; chelating agents such as ethylenediaminetetraacetic acid; buffers such as acetate, citrate or phosphate, and tonicity modifiers such as sodium chloride Or dextrose. The parenteral preparation can be enclosed in ampoules, disposable syringes or multiple dose vials made of glass or plastic. Physiological saline is a preferred adjuvant. The injectable pharmaceutical composition is preferably sterile.

  Liquid pharmaceutical compositions intended for either parenteral or oral administration must contain an amount of particulate BT compound so that an appropriate dosage is obtained. Typically this amount is at least 0.01% of the particulate BT compound in the composition. If intended for oral administration, this amount may vary from 0.1 to about 70% by weight of the composition. Preferred oral pharmaceutical compositions contain about 4% to about 50% of the BT compound. Preferred pharmaceutical compositions and preparations according to the present invention are prepared so that a parenteral dosage unit contains 0.01 to 10% by weight of the particulate BT compound prior to dilution.

  The pharmaceutical composition may be intended for topical administration, in which case the carrier may optionally comprise a solution, emulsion, ointment or gel base. The base may include, for example, one or more of the following: petrolatum, lanolin, polyethylene glycol, beeswax, mineral oil, shea butter, tea tree oil, linseed oil, hemp oil, or anti-inflammatory And / or having other vegetable oils or vegetable oils, including those known to have anti-pain or other beneficial effects, oil of shark, or anti-inflammatory and / or anti-pain or other beneficial effects Other fish oils, including those known in the art, diluents such as water and alcohols, and emulsifiers and stabilizers. A thickening agent may be present in a pharmaceutical composition for topical administration. If intended for transdermal administration, the composition may include a transdermal patch or iontophoretic device. The topical formulation may contain a particulate BT compound concentration of about 0.1 to about 10% w / v (weight per unit volume).

  The pharmaceutical composition may be intended for rectal administration, for example in the form of a suppository, which melts in the rectum to release the drug. Compositions for rectal administration may contain an oleaginous base as a suitable nonirritating excipient. Such bases include, without limitation, lanolin, cocoa butter, and polyethylene glycol.

  The pharmaceutical composition may include various materials that improve the physical form of a solid or liquid dosage unit. For example, the composition may include materials that form a coating shell around the active ingredients. The material forming the coating shell is typically inert and may be selected from, for example, sugar, shellac, and other enteric coating agents. Alternatively, the active ingredient may be enclosed in a gelatin capsule.

  A pharmaceutical composition in solid or liquid form may include an agent that is bound to the particulate BT compound, thereby assisting in the delivery of the compound. Suitable agents that can act in this capacity include monoclonal or polyclonal antibodies, proteins or liposomes. However, certain contemplated embodiments explicitly exclude the inclusion of liposomes within the pharmaceutical composition.

  The pharmaceutical composition may consist of dosage units that can be administered as an aerosol. The term aerosol is used to denote a variety of systems, from colloidal to systems consisting of pressurized vessels. Delivery may be by a liquefied or compressed gas that dispenses the active ingredients, or by a suitable pump system. The aerosol of particulate BT compound may be delivered in a single-phase, two-phase, or three-phase system to deliver the active ingredient. The delivery of the aerosol includes the necessary containers, activators, valves, subcontainers, etc., which may be combined to form a kit. One skilled in the art may determine a preferred aerosol without undue experimentation.

  The pharmaceutical composition may be prepared by methodology well known in the pharmaceutical industry. For example, a pharmaceutical composition intended for administration by injection can be prepared by mixing a compound of the present invention with sterile distilled water to form a solution. A surfactant may be added to facilitate the formation of a homogeneous solution or suspension. A surfactant is a compound that interacts non-covalently with a compound of the invention to facilitate dissolution or homogeneous suspension of the compound in an aqueous delivery system.

  The particulate BT compound described herein, or a pharmaceutically acceptable salt thereof, is administered in a therapeutically effective amount, which depends on the activity of the specific compound used; metabolic stability of the compound Gender and duration of action; patient age, weight, general health, sex and diet; mode of administration and time; excretion rate; combination of drugs; severity of a particular disorder or condition; and treatment the subject is receiving It varies depending on various factors including the beginning. Generally, a therapeutically effective daily dose is (for a 70 kg mammal) about 0.001 mg / kg (ie 0.07 mg) to about 100 mg / kg (ie 7.0 g); preferably therapeutically effective The amount is about 0.01 mg / kg (ie 7 mg) to about 50 mg / kg (ie 3.5 g) (for a 70 kg mammal); more preferably a therapeutically effective dose is (70 kg In mammals, it is about 1 mg / kg (ie 70 mg) to about 25 mg / kg (ie 1.75 g).

Effective dose ranges provided herein are not limiting and represent preferred dose ranges. However, the most preferred dosage will be understood by one of ordinary skill in the related art, as can be determined, so that adapt to each subject (e.g., Berkow et al., Eds, The Merk Manual 16 th edition, Merk and Co., Rahway, N.J., 1992 ;. Goodmanetna eds, Goodman and Cilman's The Pharmacological Basis of Therapeutics, 10 th edition, Pergamon Press, Inc., Elmsford, N.Y, (2001);. Avery 's Drug Treatment: Principles and Practice of Clinical Pharmacology and Therapeutics, 3rd edition, ADIS Press, LTD., Williams and Wilkins, Baltimore, MD. ^{Pharmaceutical Sciences, 18 th edition, Mack} Publishing Co., Easton, PA (1990); Katzung, see Basic and Clinical Pharmacology, Appleton and Lange , Norwalk, CT (1992)).

  If desired, the total dose required for each treatment can be administered by repeated or single administration within a day. Generally, treatment is initiated with smaller dosages which are less than the optimum dose of the compound. Thereafter, the dosage is increased by small increments until the optimum effect under the circumstances is reached. The diagnostic pharmaceutical compound or composition can be administered alone or together with other diagnostic agents and / or pharmaceuticals for the pathogenesis or other symptoms of the pathogen. The recipient of administration of the particulate BT compound and / or composition may be any vertebrate, such as a mammal. Preferred recipients of mammals are primates (humans, apes, monkeys, etc.), cloven-hoofed animals (horses, goats, cows, sheep, pigs, etc.), rodents (mouses, rats, rabbits, hamsters, etc.) , And mammals (such as cats and dogs). Of the birds, preferred recipients are turkeys, chickens and other animals of the same eye. The most preferred recipient is a human.

  For topical application, it is preferred to administer an effective amount of a pharmaceutical composition containing particulate BT to a target area, such as the skin surface, mucous membrane, and the like. This amount is generally about 0.0001 mg to BT compound per application, depending on the area treated whether it is diagnosed, prevented or treated, the severity of the symptoms, and the nature of the topical vehicle used. It will be in the range of about 1 g. A preferred topical preparation is an ointment, in which about 0.001 to about 50 mg of active ingredient is used per cc of ointment base. The pharmaceutical composition can be formulated as a transdermal composition or transdermal delivery device (“patch”). Such compositions include, for example, a support, a reservoir of active compound, a control membrane, a liner and a contact adhesive. Such transdermal patches may be used to provide continuous, pulsatile or on-demand delivery of the compounds of the invention if desired.

  The particulate BT composition can be formulated to provide rapid, sustained or delayed release of the active ingredient after administration to a patient by using procedures known in the art. Controlled release drug delivery systems include osmotic pump systems and dissolution systems containing polymer-coated reservoirs or drug polymer matrix formulations. Examples of controlled release systems include US Pat. Nos. 3,845,770 and 4,326,525, and J. et al. Kuzma et al, Regional Anesthesia 22 (6): 543-551 (1997), all of which are incorporated herein by reference.

  The particulate BT composition can also be delivered via intranasal drug delivery systems for topical, systemic, and nose-to-brain medical therapies. Controlled Particle Dispersion (CPD) (TM) technology, traditional nasal spray bottles, inhalers or nebulizers can provide effective local and systemic delivery of drugs by targeting the olfactory region and sinuses Are known to those skilled in the art.

  The invention also relates, in certain embodiments, to an intravaginal shell or core type drug delivery device suitable for administration to human or animal females. The device may be composed of a pharmaceutically active ingredient in a polymer matrix and surrounded by a sheath, as described in WO 98/50016, in a number of times similar to the device used to apply testosterone. , It may be possible to release the compound in a substantially zero order pattern.

  Current intraocular delivery methods include topical administration (eye drops), subconjunctival injection, periocular injection, intravitreal injection, surgical implantation and iontophoresis (using small currents into body tissues and body tissues Transport ionized drug via Those skilled in the art will combine the best suitable excipients with compounds for safe and effective intraocular administration.

  The most suitable route will depend on the nature and severity of the condition being treated. Those skilled in the art are also familiar with determining methods of administration (oral, intravenous, inhalation, subcutaneous, rectal, etc.), dosage forms, appropriate pharmaceutical excipients, and other matters related to delivering the compound to the subject in need. Will be.

  The compositions and methods described herein treat MRSA skin infections to prevent chronic wounds, for example, as a cream for burns, as a topical agent to treat existing wounds as described herein. For use in the treatment of acute and chronic wounds and wound biofilms, for example, and for other related indications disclosed herein and apparent to those of skill in the art in view of the present disclosure. Also good.

  Non-limiting examples of bacteria in which the compositions and methods described herein may find beneficial use according to certain embodiments described herein include Staphylococcus aureus (S. aureus). MRSA (methicillin resistant S. aureus), Staphylococcus epidermidis, MRSE (methicillin resistant S. epidermidis), Mycobacterium tuberculosis, Mycobacterium abium, Pseudomonas aeruginosa, drug resistant P. aureus Aeruginosa, Escherichia coli, enterotoxigenic E. coli E. coli, enterohemorrhagic E. coli E. coli, Klebsiella pneumoniae, Clostridium difficile, Helicobacter pylori, Legionella pneumophila, Enterococcus faecalis, Methicillin-susceptible enterococcus fecaris, Enterobacter cloacae, Salmonella typhimurium, Proteus bulgaris, Yersinia enterico Vibrio cholere, Shigella flexneri, vancomycin-resistant enterococcus (VRE), Burkholderia cepacia group, Francisella larensis, Bacillus anthracis, Yersinia pestis, Pseudomonas aeruginosa, vancomycin-sensitive and vancomycin-resistant enterococcus (e.g. E. faecalis, E. faecium), methicillin sensitivity and Syrin-resistant Staphylococcus spp. (Eg S. aureus, S. epidermidis) and Acinetobacter baumani, Staphylococcus haemolyticus, Staphylococcus hominis, Enterococcus pecium, Streptococcus pyogenes, Streptococcus agalanthia Klebsiella pneumoniae, Proteus mirabilis, Proteus bulgaris, Yersinia enterocolitica, Stenotrohomomonas maltophilia, Streptococcus pneumoniae, Penicillin-resistant Streptococcus pneumoniae, Burkholderia cepacia, Burkholderia multivolance Mycobacterium smegmatis and E. coli Black frog is listed.

  The practice of certain embodiments of the invention, unless specified to the contrary, is conventional in the art of microbiology, molecular biology, biochemistry, cell biology, virology and immunology, which are within the skill of the art, In addition, for reference purposes, the following reference materials are used. Such techniques are explained fully in the literature. For example, Sambrook, et al. , Molecular Cloning: A Laboratory Manual (2nd Edition, 1989): Maniatis et al. Molecular Cloning: A Laboratory Manual (1982); DNA Cloning: A Practical Approach, vol. I & II (D. Glover, ed.); Oligonucleotide Synthesis (N. Gait ed., 1984); Nucleic Acid Hybridization (B. Hames & S. Higgins ed., 1985); eds., 1984); Animal Cell Culture (R. Freshney ed., 1986); Perbal, A Practical Guide to Molecular Cloning (1984).

  Unless the context requires otherwise, throughout this specification and the claims, the language “comprise” and variations thereof, such as “comprises” and “including” It is an open and comprehensive meaning and should be interpreted as “including but not limited to”.

  Reference throughout this specification to “one embodiment” or “an embodiment” or “aspect” is intended to mean that a particular feature, structure, or characteristic described in connection with that embodiment is at least one of the present invention It is meant to be included in the embodiment. The appearances of the phrase “one embodiment” or “an embodiment” in various places throughout this specification are not necessarily all referring to the embodiment. Furthermore, the particular features, structures, or characteristics may be combined in any suitable manner in one or more embodiments.

Certain embodiments may include promoting skin tissue repair in a subject, one or more cells for treating an acute or chronic wound or wound biofilm in a subject, or in a cell or cells It relates to methods, compositions and kits for altering wound repair activity. A cell generally represents a single cell, but multiple cells represent more than one cell. A cell may comprise a tissue, organ or whole organism. Further, the cell or cells may be placed in vivo, in vitro, or ex vivo. Maintaining cell, tissue and organ cultures is a routine procedure for those skilled in the art, and the conditions and media can be readily ascertained (eg, Freshney, Culture of Animal Cells: A Manual of Basic Techniques). ^{, Wiley-Liss 5 th Ed (} 2005);. Davis, see Basic Cell Culture, Oxford University Press 2 nd Ed (2002))..

  As disclosed herein, certain embodiments relate to a method of treating an acute or chronic wound or wound biofilm in a subject, the method being described herein for use in such a method. Antibiotics described herein comprising a BT compound (eg, provided in the form of a plurality of substantially monodispersed microparticles), and optionally in certain further embodiments, used in such methods. Administering a composition comprising a substance compound to a subject in a therapeutically effective amount, for example, BT compound is BisEDT or BisBAL, or other compounds shown in Table 1 herein, or Domenico et al. 1997 Antimicrob.Agent.Chemother.41: 1697; 2001 Antimicrob.Age .. T Chemother 45: 1421) and / or U. S RE 37,793, U.S. Pat. S. With any other BT agent described in US Pat. Nos. 6,248,371, 6,086,921, and 6,380,248 and / or prepared by the methods disclosed herein is there. Another specific embodiment relates to a method comprising contacting any natural surface with a composition comprising one or more of the particulate BT compounds described herein, wherein the step of contacting directly Application, coating, dipping, irrigation, spraying, application, or one or more other methods of bringing the BT composition into contact with the natural surface.

  The step of administering to a subject, such as a human or other mammalian subject, can be performed by any means known in the art, eg, locally (skin or any epithelial tissue surface, such as glandular tissue or respiratory tract and / or gastrointestinal tract). Including direct administration to surfaces that may be present in the tube), intravaginal, intraperitoneal, oral, parenteral, intravenous, intraarterial, transdermal, sublingual, subcutaneous, intramuscular, buccal, intranasal, inhalation , Intraocular, intrafacial, intraarticular or intrathecal.

  In preferred embodiments, administration may be performed locally and pharmaceutical excipients or carriers for topical use are described herein and are known in the art.

  As previously mentioned, certain embodiments of the invention described herein relate to topical formulations of the described BT compounds (eg, BisEDT and / or BisBAL), which formulations are specific In further embodiments, one or more antibiotic compounds described herein, such as amikacin, ampicillin, cefazolin, cefepime, chloramphenicol, ciprofloxacin, clindamycin (or other lincosamide systems) Antibiotics), daptomycin (Cubicin®), doxycycline, gatifloxacin, gentamicin, imipenim, levofloxacin, linezolid (Zyvox®), minocycline, nafcillin, paromomycin, rifampin, sulfamethoxazole, tobramycin Or carbapenem antibiotics, cephalosporin antibiotics, fluoroquinolone antibiotics, glycopeptide antibiotics, lincosamide antibiotics, penicillinase resistant penicillin antibiotics, and / or aminopenicillin antibiotics and / or Or aminoglycoside antibiotics such as amikacin, arbekacin, gentamicin, kanamycin, neomycin, netilmycin, paromomycin, rhodostreptomycin, streptomycin, tobramycin or apramycin, and / or lipopeptide antibiotics such as daptomycin (Cubicin®), Alternatively, an oxazolidinone antibiotic such as linezolid (Zyvox®) may be included. These and related formulations may relate to animals, preferably mammals, most preferably humans, in particularly preferred embodiments humans with acute or chronic wounds, or biofilms (eg, biofilm formation). When administered locally to a human with a wound containing a bacterial infection, or a wound containing a bacterial infection such as a biofilm or the presence of bacteria A BT compound (and optionally one or more antibiotics) in therapeutic amounts in a pharmaceutically acceptable carrier, excipient or diluent as disclosed herein. Also good.

  Topical administration of a BT compound described herein or a pharmaceutically acceptable salt thereof in pure form or in a suitable pharmaceutical composition may be any of the acceptable modes of topical administration of drugs suitable for similar uses. Can be implemented through. Topical application or administration of the composition, in a preferred embodiment, applies the composition (eg, topical formulation) to the skin and / or other epithelial tissue surface (eg, respiratory tract, gastrointestinal tract and / or subject) to be treated. Or may be one or more localized or widely distributed skin and / or other epithelial tissue surface sites, typically topical formulations May be referred to as an acute or chronic wound site surrounded by, but not limited to, an intact stratum corneum or epidermis layer, for example, certain embodiments may be used as a topical application for damage, scratching or injury. The topical formulation described herein is intended to be administered to skin subjected to surgery or to the skin of a subject who has undergone surgery, wherein contact of the topical formulation is the stratum corneum or epidermal layer Not only in skin granule cells, spinous cells and / or basal cell layers, eg with certain types of wound repair or wound healing or other skin tissue remodeling, and / or in dermis or underlying tissue It may be broken.

  Such skin tissue repair is therefore in certain preferred embodiments, for example preventing or ameliorating acute or chronic wounds or wound biofilms, or as another example preventing or ameliorating the dehiscence of skin wounds. Or may include dermal wound healing that may be desirable in improving, promoting or otherwise enhancing dermal wound healing when acute or chronic wounds and / or skin wound tears may be present. Other specific embodiments contemplated for local administration to the surface of the epithelial tissue residing in the respiratory tract, gastrointestinal tract and / or glandular epithelium also provide the topical preparation provided herein to the respiratory tract. Present in ducts (eg, respiratory tract, nasopharyngeal and laryngeal routes, trachea, lung, bronchi, bronchiole, alveoli, etc.) and / or gastrointestinal tract (eg, oral cavity, esophagus, stomach, intestine, rectum, anus, etc.) 1 Administration of topical formulations by any suitable route known in the art for delivery to one or more epithelial tissues and / or other epithelial surfaces may be included.

  According to certain contemplated embodiments, topical administration may include direct application to an open wound. For example, open fractures or other open wounds may include lacerations in the skin that can expose additional underlying tissue to the external environment in a manner that sensitizes microbial infections. Such a situation is not uncommon in certain types of acute trauma military wounds including, for example, type III (severe) open fractures. According to these and related embodiments, topical administration may be achieved by applying the BT compositions described herein to such impaired skin and / or other epithelial surfaces and / or other tissues, such as muscle, Even by direct contact with connective tissues such as ligaments, tendons, bones, circulatory tissues such as blood vessels, related neural tissues, and any other organs that may be exposed within such open wounds Good. Examples of other tissues that may be exposed, i.e., such direct contact may be exposed to kidney, bladder, liver, pancreas, and opportunistic infections associated with open wounds in such deleterious conditions Any other tissue or organ may be mentioned.

  Topical formulations (e.g., pharmaceutical compositions) include the described BT compounds (e.g., RE 37,793, US 6,248,371, 6,086,921, and / or 6, 380,248, and / or compounds prepared according to the present disclosure, such as the particulate BT suspensions described herein) In one embodiment, one or more desired antibiotics (eg, aminoglycoside antibiotics such as amikacin) are combined with a BT compound separately or together with a suitable pharmaceutically acceptable for use in topical formulation preparations. May be prepared by combining with possible carriers, diluents or excipients, solids, semi-solids, gels, creams, colloids, suspensions or liquids, or other Where application forms, such as powders, granules, ointments, solutions, wash, gels, pastes, plasters, liniments, bioadhesives, microsphere suspensions, and the like in an aerosol spray, may be incorporated into the preparation.

  The pharmaceutical compositions of these and related embodiments are formulated such that the active ingredient can be included in the composition, and in certain preferred embodiments, the BT compounds described herein alone or in one form. Desired antibiotics (for example, carbapenem antibiotics, cephalosporin antibiotics, fluoroquinolone antibiotics, glycopeptide antibiotics, lincosamide antibiotics, penicillinase-resistant penicillin antibiotics, and aminopenicillin antibiotics) Substances, or aminoglycoside antibiotics such as amikacin, or rifamycin), which are subjects, eg mammals such as humans, in certain preferred embodiments, human patients with acute or chronic wounds, or Acute or chronic wound or wound biofilm Or a combination preparation containing a BT compound and / or an antibiotic compound in an acute or chronic wound and optionally into the surrounding skin of a high-risk human patient (eg obese and / or diabetic individual) May be applied in either order simultaneously or sequentially so as to be bioavailable upon topical administration. While certain embodiments disclosed herein contemplate topical administration of BT compounds and antibiotics, such as administration that may be in either order simultaneously or sequentially, the invention is not limited thereto. Rather, in other embodiments, distinct BT compound routes of administration are explicitly contemplated with respect to antibiotic routes of administration. That is, the antibiotic may be administered orally, intravenously, or by any other route of administration described herein, and the BT compound is administered by a route that is independent of the route used for antibiotics. May be. As a non-limiting example, the BT compound may be administered topically as provided herein, and the antibiotics may be administered simultaneously or sequentially (and in any order) by different routes, eg, oral, Administration may be by intravenous, transdermal, subcutaneous, intramuscular, and / or any other route of administration.

  The topical formulations described herein can be applied by spraying, ligating, dipping and / or applying a therapeutically effective amount of a disinfectant or wound healing agent (and optionally an antibiotic) to the wound site, For example, delivery to skin cells such as dermal fibroblasts. A preferred formulation may be contacted with a desired site, such as a local wound site, chronic wound, epithelial tissue surface or other site intended for administration, and such a formulation may therefore be permeated through the skin of the drug composition. It may indicate immediate skin penetration as determined by any of a number of established methodologies known in the art for testing sex (eg, Wagner et al., 2002 J. Invest. Dermatol. 118: 540 and references cited therein; Bronaugh et al., 1985 J. Pharm. Sci. 74:64; al., 1996 J. Pharm. Med. Anal. 1996 14: 1015-23; Bonferoni et al., 1999 Pharm. Dev. Technol., 4: 45-53; 59 in Frantz, Instrumentation and methodology for in vitro skin diffusion cells, methodology for skin abduction, YM, Transformal Controlled Systems Y el Dekker, New York, 1987, Tojo in the 127-158, Design and calibration of in vitro permeation apparatus; Dermatological Formulations: Percutaneous absorption, Marcel Dekker, New York, 1983, Barry in the 234-295, Methods for studying percutaneous absorption reference).

  Compositions to be administered to the subject's or patient's skin, and formulations containing such compositions, in certain embodiments, may take the form of one or more dosage units, such as liquid filled capsules or ampoules. A single dosage unit may be included, and a container of the topical formulation described herein in aerosol form may hold multiple dosage units. Actual methods of preparing such dosage forms are known or will be apparent to those skilled in the art, see, eg, The Science and Practice of Pharmacy, 20th Edition (Philadelphia College of Pharmacy and Science, 2000). I want. The composition or formulation to be administered is, in any event, an antiseptic and / or healing-promoting compound (eg, a BT compound) provided herein in accordance with the teachings of the present invention, or a pharmaceutically acceptable salt thereof Contains a therapeutically effective amount of salt.

  As mentioned above, the topical formulations of the present invention may take any of a wide variety of forms, including creams, lotions, solutions, sprays, gels, ointments, pastes, and / or liposomes. , And may be prepared to contain micelles and / or microspheres. See, for example, US Pat. No. 7,205,003. For example, creams are mucus or semi-solid emulsions, either oil-in-water or water-in-oil, as is well known in the pharmaceutical and cosmetic formulation arts. The cream base is washable and contains an oil phase, an emulsifier and an aqueous phase. The oil phase, also called the “inner” phase, is generally composed of petrolatum and fatty alcohols such as cetyl alcohol or stearyl alcohol. Usually, though not necessarily, the aqueous phase has a volume that exceeds the oil phase and generally contains a hygroscopic agent. Emulsifiers in cream formulations are generally nonionic, anionic, cationic or amphoteric surfactants.

  Preferred lotions for cosmetic delivery are preparations that are applied to the skin surface without rubbing, typically liquid or semi-liquid preparations in which solid particles containing the active agent are present in a water or alcohol base. It is. Lotions are usually solid suspensions, preferably including oil-in-water liquid oil emulsions. Lotion is the preferred formulation herein for treating large body surface areas because of the ease of applying a highly flowable composition. It is generally preferred that the insoluble material in the lotion is ultrafine. Lotions typically contain a suspending agent and, in addition to the suspending agent that produces a better dispersion, are useful for localizing the active ingredient and keeping it in contact with the skin. Contains compounds such as methylcellulose, sodium carboxymethylcellulose and the like.

  A solution is a homogeneous mixture prepared by dissolving one or more chemical substances (solutes) in a liquid and dispersing the molecules of the dissolved substance in the molecules of the solvent. The solution may contain other pharmaceutically acceptable and / or cosmetically acceptable chemicals to buffer, stabilize or store the solute. Common examples of solvents used to prepare the solution are ethanol, water, propylene glycol or any other pharmaceutically acceptable and / or cosmetically acceptable vehicle.

  Gels are semi-solid suspension type systems. Single-phase gels are typically aqueous, but preferably contain organic macromolecules distributed substantially homogeneously throughout the carrier liquid, which also contains alcohol and any oil. Preferred “organic macromolecules”, ie gelling agents, may be chemically cross-linked polymers, such as cross-linked acrylic acid polymers, such as polymer “carbomers”, such as carboxypolyalkylenes, which are Carbopol®. (Trademark) may be obtained commercially. Also preferred in certain embodiments are hydrophilic polymers such as polyethylene oxide, polyoxyethylene-polyoxypropylene copolymers, and polyvinyl alcohol; cellulosic polymers such as hydroxypropylcellulose, hydroxyethylcellulose, hydroxypropylmethylcellulose, hydroxy It may be propyl methylcellulose phthalate and methylcellulose; gums such as tragacanth gum and xanthan gum; sodium alginate; and gelatin. To prepare a homogeneous gel, a dispersing agent such as alcohol or glycerin can be added, and the gelling agent can be dispersed by grinding, mechanical mixing or stirring, or a combination thereof.

  Ointments, which are also well known in the art, are typically semisolid preparations based on petrolatum or other petroleum derivatives. The specific ointment base used is one that provides a plurality of desired characteristics, such as emollient, as understood by those skilled in the art. As with other carriers or vehicles, an ointment base must be inert, stable, nonirritating, and nonsensitizing. Remington: The Science and Practice of Pharmacy, 19th Ed (Easton, Pa .: Mack Publishing Co., 1995), p1399-1404, ointment bases are classified into four categories: oil-based bases; Agents; emulsion bases; and water-soluble bases. Oily ointment bases include, for example, vegetable oils, fats obtained from animals, and semisolid hydrocarbons obtained from petroleum. Emulsifying ointment bases, also known as absorbent ointment bases, contain little or no water and include, for example, hydroxystearic sulfate, anhydrous lanolin, and hydrophilic petrolatum. Emulsion ointment bases are either water-in-oil (W / O) emulsions or oil-in-water (O / W) emulsions and include, for example, cetyl alcohol, glyceryl monostearate, lanolin, and stearic acid. Preferred water-soluble ointment bases are prepared from polyethylene glycols of various molecular weights (see for example Remington's previous work).

  The paste is a semi-solid dosage form, and the active agent is suspended in a suitable base. Depending on the nature of the base, the paste is fractionated into a fat paste or a paste produced with a single phase aqueous gel. The base in the fat paste is generally petrolatum or hydrophilic petrolatum. A paste produced from a single-phase aqueous gel generally incorporates carboxymethyl cellulose or the like as a base.

  The formulation may be prepared using liposomes, micelles and microspheres. Liposomes are microscopic vesicles having one (unilamellar) or multiple (multilamellar) lipid walls containing lipid bilayers, and are used herein as one of the topical formulations described herein. The above ingredients, such as antiseptics, wound healing / skin tissue / epithelial tissue repair-promoting compounds (eg, particulate BT compounds, optionally with one or more antibiotics) or specific carriers or excipients, lipid membranes It may be encapsulated on the surface and / or adsorbed on the lipid membrane surface. The liposomal preparations herein include cationic (positive charge), anionic (negative charge), and neutral preparations. Anionic liposomes are readily available. For example, N [1- (2,3-dioleyloxy) propyl] -N, N, N-triethylammonium (DOTMA) liposomes are commercially available under the trade name Lipofectin® (GIBCO BRL, Grand Island, NY). ). Similarly, anionic and neutral liposomes are readily available, eg, from Avanti Polar Lipids (Birmingham, AL) or can be readily prepared using readily available materials. Such materials include, among others, phosphatidylcholine, cholesterol, phosphatidylethanolamine, dioleylphosphatidylcholine (DOPC), dioleylphosphatidylglycerol (DOPG), and dioleylphosphatidylethanolamine (DOPE). These materials can also be mixed with DOTMA in appropriate ratios. Methods for producing liposomes using these materials are well known in the art.

  Micelles are composed of surfactant molecules arranged so that the polar head group forms an outer spherical shell and the hydrophobic hydrocarbon chain is positioned towards the center of the sphere to form the core. Is known in the art. Since micelles are formed in an aqueous solution containing a sufficiently high concentration of surfactant, micelles are naturally obtained. Surfactants useful for forming micelles include, but are not limited to, potassium laurate, sodium octane sulfonate, sodium decane sulfonate, sodium dodecane sulfonate, sodium lauryl sulfate, sodium doxate, decyltrimethylammonium bromide , Dodecyltrimethylammonium bromide, tetradecyltrimethylammonium bromide, tetradecyltrimethylammonium chloride, dodecylammonium chloride, polyoxy-8 dodecyl ether, polyoxy-12 dodecyl ether, nonoxynol 10, and nonoxynol 30.

  Similarly, microspheres may be incorporated into the topical formulations described herein. Like liposomes and micelles, microspheres essentially encapsulate one or more components of the combination of the present invention. In general, but not necessarily, they are formed from lipids, preferably charged lipids such as phospholipids. The preparation of lipidic microspheres is well known in the art.

Various additives known to those skilled in the art may be included in the topical formulation. For example, a solvent containing a relatively small amount of alcohol may be used to solubilize certain compounding ingredients. In certain topical formulations, or in particular in the case of severe skin damage, such as post-operative acute or chronic wounds or post-operative dermal wound dehiscence, including a skin penetration enhancer added in the topical formulation May be desirable. Examples of suitable enhancers include, but are not limited to, ethers such as diethylene glycol monoethyl ether (commercially available as Transcutol®) and diethylene glycol monomethyl ether; surfactants such as sodium laurate, sodium lauryl sulfate, cetyltrimethyl Ammonium bromide, benzalkonium chloride, Poloxamer® (231, 182, 184), Tween® (20, 40, 60, 80), and lecithin (US Pat. No. 4,783,450); Alcohols such as ethanol, propanol, octanol, benzyl alcohol, etc .; polyethylene glycol and its esters such as polyethylene glycol monolaurate (PEGML; eg US Pat. No. 4 568,343); amides and other nitrogen compounds such as urea, dimethylacetamide (DMA), dimethylformamide (DMF), 2-pyrrolidone, 1-methyl-2-pyrrolidone, ethanolamine, diethanolamine, and triethanolamine Terpenes; alkanones; and organic acids, especially citric acid and succinic acid. Azone® and sulfoxides such as DMSO and C ₁₀ MSO may be used but are less preferred.

The most preferred skin penetration enhancers are their lipophilic co-enhancers, typically referred to as “plasticization” enhancers, ie a molecular weight in the range of about 150-1000 daltons, about 1% by weight. Less than, preferably less than about 0.5 wt.%, Most preferably less than about 0.2 wt. The Hildebrand solubility parameter of the plasticization enhancer is in the range of about 2.5 to about 10, preferably in the range of about 5 to about 10. Preferred lipophilic enhancers are fatty esters, fatty alcohols, and fatty ethers. Examples of specific and most preferred fatty acid esters include methyl laurate, ethyl oleate, propylene glycol monolaurate, propylene glycerol dilaurate, glycerol monolaurate, glycerol monooleate, isopropyl n-decanoate, and octyldodecyl Miri start is mentioned. Examples of fatty alcohols include stearyl alcohol and oleyl alcohol, and examples of fatty ethers include diols or triols, preferably C ₂ -C ₄ alkanediols or triols substituted with 1 or 2 fatty ether substituents. Compounds. Additional skin penetration enhancers are known to those skilled in the art of topical drug delivery and / or are described in the relevant literature. For example, Percutaneous Penetration Enhancements eds. Smith et al. (CRC Press, Boca Raton, FL, 1995).

In addition to those previously identified, a variety of other additives may be included in the topical formulations according to certain embodiments of the invention. These include, but are not limited to, antioxidants, astringents, fragrances, preservatives, emollients, pigments, dyes, hygroscopic agents, propellants, and sunscreens, as well as cosmetics, drugs, Or other types of materials desirable in other respects. Typical examples of optional additives included in the formulations of certain embodiments of the invention are as follows: preservatives such as sorbates; solvents such as isopropanol and propylene glycol; astringents such as Menthol and ethanol; emollients such as polyalkylene methyl glucoside; hygroscopic agents such as glycerine; emulsifiers such as glycerol stearate, PEG-100 stearate, polyglyceryl-3 hydroxylauryl ether, and polysorbate 60; sorbitol and other polyhydroxy alcohols For example, polyethylene glycol; sunscreens such as octyl methoxycinnamate (commercially available as Parsol MCX) and butylmethoxybenzoylmethane (commercially available under the trade name Parsol 1789) Antioxidants such as ascorbic acid (vitamin C), alpha-tocopherol (vitamin E), beta-tocopherol, .gamma.-tocopherol, .delta.-tocopherol, .epsilon.-tocopherol, zeta ₁ - tocopherol, zeta ₂ - tocopherol, .eta. tocopherol, And retinol (vitamin A); essential oils, ceramides, essential fatty acids, mineral oils, wetting agents and other surfactants such as the PLURONIC® series of hydrophilic polymers available from BASF (Mt Olive, NJ), vegetable oils ( Eg, soybean oil, palm oil, liquid fraction of shea butter, sunflower oil), animal oil (eg, perhydrosqualene), mineral oil, synthetic oil, silicone oil or wax (eg, cyclomethicone and dimethicone), fluorinated Oil (generally Perf Olopolyethers), fatty alcohols (eg, cetyl alcohol), and waxes (eg, beeswax, carnauba wax, and paraffin wax); skin-feel modifiers; and thickeners and structures ), For example, swellable clays and bridged carboxypolyalkylenes which may be obtained commercially under the trade name Carbopol®.

  Other additives include materials that condition the skin (especially the top layer of the skin in the stratum corneum) to maintain flexibility and / or protect the skin by delaying skin moisture loss Useful drugs are listed. Such conditioners and humectants include, by way of example, pyrrolidine carboxylic acids and amino acids; organic antimicrobial agents such as 2,4,4′-trichloro-2-hydroxydiphenyl ether (triclosan) and benzoic acid; anti-inflammatory agents such as Acetylsalicylic acid and glycyrrhetinic acid; antiseborrheic agents such as retinoic acid; vasodilators such as nicotinic acid; melanogenesis inhibitors such as kojic acid; and mixtures thereof. Other cosmetically active agents that are advantageously included include, for example, α-hydroxy acids, α-keto acids, polymeric hydroxy acids, moisturizers, collagen, marine extracts, and antioxidants such as ascorbic acid (vitamin C ), Α-tocopherol (vitamin E) or other tocopherols, such as those described above, and retinol (vitamin A), and / or cosmetically acceptable salts, esters, amides, or other derivatives thereof May be. Additional cosmetic agents include those that can improve the oxygen supply in the skin tissue, as described, for example, in WO 94/00098 and WO 94/00109. Sunscreen agents may be included.

  Other embodiments may include various non-carcinogenic, non-irritating healing materials that facilitate treatment with the combination of certain embodiments of the invention. Such healing materials include nutrients, minerals, vitamins, electrolytes, enzymes, herbs, plant extracts, honey, gland or animal extracts, or safe therapeutics that can be added to a combination to promote dermal healing. May be. The amounts of these various additives are those conventionally used in the cosmetics field, for example in the range of about 0.01% to about 20% of the total weight of the topical formulation.

  Formulations of certain embodiments of the present invention may contain conventional additives such as opacifiers, fragrances, colorants, gelling agents, thickeners, stabilizers, surfactants and the like. Other agents, such as antimicrobial agents, may be added to prevent spoilage during storage, ie to inhibit the growth of microorganisms such as yeast and mold. Suitable antimicrobial agents are typically selected from methyl and propyl esters of p-hydroxybenzoic acid (eg, methyl and propylparaben), sodium benzoate, sorbic acid, imidourea, and combinations thereof. The combination is an anti-irritant that minimizes or eliminates the possibility of skin irritation or skin damage arising from the administered anti-infective acute or chronic wound healing and skin tissue repair-promoting compounds or from other components of the composition An additive may be contained. Suitable stimulant mitigating additives include α-tocopherols; monoamine oxidase inhibitors, in particular phenyl alcohols such as 2-phenyl-1-ethanol; glycerine; salicylates; ascorbates; ionophores such as monensin; Amines; ammonium chloride; N-acetylcysteine; capsaicin; and chloroquine. An irritation additive, if present, is effective to alleviate irritation or skin damage, typically representing no more than about 20% by weight of the formulation, more typically no more than about 5% by weight. It may be incorporated into topical formulations at a concentration.

  The topical formulation may be a disinfectant / wound healing / anti-biofilm / skin tissue repair-promoting compound (eg, a BT compound, preferably as substantially homogeneous microparticles provided herein and optionally herein). In addition to the described synergistic antibiotic (s), it may contain a therapeutically effective amount of one or more additional pharmacologically active agents suitable for topical administration. Such agents include, for example, phospholipids capable of improving the oxygen supply in skin tissue and oxygenated fluorocarbons or fluorocarbon compounds as described in International Patent Publication Nos. WO94 / 00098 and WO94 / 00109. An asymmetric lamellar aggregate composed of the mixture may be included.

  Suitable pharmacologically active agents that can be incorporated into, ie applied topically, the topical formulations of the present invention may include, but are not limited to: colored or uncolored aging spots, horns Agents that improve or cure wrinkles and wrinkles; antimicrobial agents; antibacterial agents, anti-itch and anti-drying agents; anti-inflammatory agents; local anesthetics and analgesics; corticosteroids; retinoids (eg retinoic acid; vitamins, Hormones, and antimetabolites). Some examples of topical pharmacologically active agents include acyclovir, amphotericin, chlorhexidine, clotrimazole, ketoconazole, econazole, miconazole, metronidazole, minocycline, nystatin, neomycin, kanamycin, phenytoin, p-aminobenzoate Octyl methoxycinnamate, octyl salicylate, oxybenzone, dioxybenzone, tocopherol, tocopheryl acetate, selenium sulfide, pyrithione zinc, diphenhydramine, paramoxine, lidocaine, procaine, erythromycin, tetracycline, clindamycin, crotamiton, monoquinone and benzyl ether , Naproxen, ibuprofen, cromolyn, retinoic acid, retinol Retinyl palmitate, retinyl acetate, coal tar, griseofulvin, estradiol, hydrocortisone, hydrocortisone 21-acetic acid, hydrocortisone 17-valeric acid, hydrocortisone 17-butyric acid, progesterone, betamethasone valeric acid, betamethasone dipropionic acid, triamcinolone acetonide, fluocinozide Examples include propionic acid, minoxidil, dipyridamole, diphenylhydantoin, benzoyl peroxide, and 5-fluorouracil. As also previously mentioned, certain embodiments include antibiotics such as carbapenem antibiotics, cephalosporin antibiotics, fluoroquinolone antibiotics, glycopeptide antibiotics, lincosamide antibiotics, penicillinase resistant penicillin. It is intended to be included in a combination drug such as an antibiotic, an aminopenicillin antibiotic, or an aminoglycoside antibiotic such as amikacin.

  A pharmacologically acceptable carrier may be incorporated into the topical formulation of certain embodiments of the present invention or any carrier conventionally used in the art. Examples include water, lower alcohols, higher alcohols, honey, polyhydric alcohols, monosaccharides, disaccharides, polysaccharides, sugar alcohols such as glycol (2-carbon), glycerol (3-carbon), erythritol and threitol (4-carbon), arabitol, xylitol and ribitol (5-carbon), mannitol, sorbitol, dulcitol and iditol (6-carbon), isomaltol, maltitol, lactitol and polyglycitol, hydrocarbon oil, fat, wax, Examples include fatty acids, silicone oils, nonionic surfactants, ionic surfactants, silicone surfactants, and aqueous and emulsion based mixtures of such carriers.

  Embodiments of the topical formulations of the present invention can be affected by any acute or chronic wound site (eg, the wound itself and surrounding tissue, eg, infection) that requires treatment at the frequency and amount necessary to achieve the desired result. Regularly applied to the skin area or other epithelial tissue surface (eg, gastrointestinal tract, respiratory tract, glandular tissue) or other normal or healthy surrounding tissues Also good. The frequency of treatment depends on the condition of the skin (or epithelial tissue) (eg, acute or chronic wounds, or other skin wounds that may be found in dehiscence caused by surgical excision, or other types of skin wounds), skin ( Or the degree of injury or deterioration of the other tissue), the responsiveness of the user's skin (or other tissue), the strength of the active ingredient in certain embodiments (eg, wound healing / disinfection / Anti-biofilm / skin tissue repair-promoting compounds such as BT compounds and optionally one or more additional pharmaceutically active ingredients such as antibiotics such as amikacin or other antibiotics, skin (or other epithelial surface containing The efficacy of the vehicle used to deliver the active ingredient to the appropriate layer of tissue) is removed by the physical contact with the bandage or other dressing or closing. Ease of being removed by perspiration or other intrinsic or extrinsic fluids, as well as the ease of the activity level or lifestyle of a subject or patient.

Typical concentrations of active agents, such as the BT compounds described herein, disinfecting / anti-biofilm / wound healing / skin tissue repair-promoting compositions are typically 0.001-30 based on the total weight of the composition, for example. It may be in the range of% by weight to about 0.01 to 5.0%, more preferably about 0.1 to 2.0%. As a representative example, the compositions of these embodiments of the present invention are applied to acute or chronic wounds and / or skin at a rate of about 1.0 mg / cm ² skin to about 20.0 mg / cm ² skin. Also good. Representative examples of topical formulations include, but are not limited to, aerosols, alcohols, anhydrous bases (eg, lipsticks and powders), aqueous solutions, creams, emulsions (including either water-in-oil or oil-in-water emulsions). Fats, foams, gels, hydroalcoholic solutions, liposomes, lotions, microemulsions, ointments, oils, organic solvents, polyols, polymers, powders, salts, silicone derivatives, and waxes. Topical formulations may include, for example, chelating agents, conditioning agents, emollients, excipients, hygroscopic agents, protective agents, thickeners, or UV absorbers. Those skilled in the art will appreciate that other formulations than those listed can be used in embodiments of the present invention.

A chelating agent may optionally be included in the topical formulation and may be selected from any agent suitable for use in a cosmetic composition, such as Ca ²⁺ , Mn ²⁺ , or Mg ^2+. Any natural or synthetic chemicals that have the ability to bind to the valent cationic metal may be included. Examples of chelating agents include, but are not limited to, EDTA, disodium EDTA, EGTA, citric acid, and dicarboxylic acids.

Conditioning agents may optionally be included in the topical formulation. Examples of skin conditioning agents include, but are not limited to, acetylcysteine, N-acetyldihydrosphingosine, acrylate / behenyl acrylate / dimethicone acrylate copolymer, adenosine, cyclic adenosine monophosphate, adenosine monophosphate, adenosine triphosphate Phosphate, alanine, albutene, seaweed extract, allantoin and derivatives, aloe barbadensis extract, aluminum PCA, amyloglucosidase, arbutin, arginine, azulene, bromelain, powdered buttermilk, butylene glycol, caffeine, calcium gluconate, Capsaicin, carbocysteine, carnosine, β-carotene, casein, catalase, cephalin, ceramide, chamomile / recutita (matricaria) flower extract, cholecalci Errol, cholesteryl ester, cocobetaine, coenzyme A, processed corn starch, crystallin, cycloethoxymethicone, cysteine DNA, cytochrome C, daltside, dextran sulfate, dimethicone copolyol, dimethylsilanol hyaluronate, DNA, elastin, elastin amino acid, epithelium Growth factor, ergocalciferol, ergosterol, ethylhexyl PCA, fibronectin, folic acid, gelatin, gliadin, β-glucan, glucose, glycine, glycogen, glycolipid, glycoprotein, glycosaminoglycan, glycosphingolipid, horseradish peroxidase, Hydrogenated proteins, protein hydrolysates, jojoba oil, keratin, keratin amino acids, and kinetin, lactoferri Lanosterol, lauryl PCA, lecithin, linoleic acid, linolenic acid, lipase, lysine, lysozyme, malt extract, maltodextrin, melanin, methionine, inorganic salt, niacin, niacinamide, oat amino acid, oryzanol, palmitoyl hydrolyzed protein, bread Creatine, papain, PEG, pepsin, phospholipid, phytosterol, placental enzyme, placental lipid, pyridoxal 5-phosphate, quercetin, resorcinol acetate, riboflavin, RNA, Saccharomyces lysate extract, silk amino acid, sphingolipid, stearamide propyl Betaine, Stearyl Palmitate, Tocopherol, Tocopheryl Acetate, Tocopheryl Linoleate, Ubiquinone, Vitis Vinhera (Grape) Seed Oil, Wheat Ami Acid, xanthan gum, and zinc gluconate. As can be readily appreciated by those skilled in the art, skin conditioning agents other than those listed above may be combined with the disclosed compositions or the preparations provided thereby.

The topical formulation may optionally include one or more emollients, examples include but are not limited to acetylated lanolin, acetylated lanolin alcohol, acrylate / C _10-30 alkyl acrylate crosspolymer, Acrylate copolymer, alanine, seaweed extract, aloe barbadensis extract or gel, Altea officinalis extract, starch aluminum octenyl succinate, aluminum stearate, apricot (Prunus armenia) kernel oil, arginine, arginine aspartate, arnica Montana Extract, Ascorbic Acid, Ascorbyl Palmitate, Aspartic Acid, Avocado (Persia Gratissima) Oil, Barium Sulfate, Barrier Sphingolipid, Butyl Alcohol, Beeswax, Behe Nyl alcohol, β-sitosterol, BHT, birch (Betula alba) bark extract, borage (borago officinalis) extract, 2-bromo-2-nitropropane-1,3-diol, nagyida (Luscus acuretus) extract, Butylene glycol, Calendula officinalis extract, Calendula officinalis oil, Calendula officinalis wax, Canola oil, Canola oil, Caprylic acid / Capric acid triglyceride, Cardamom (Eretteria cardamomum) oil, Carnauba (Copernicia cerifera) wax, Carrageenan (Kondras Crispas), carrot (Daucus carrotta sativa) oil, castor (Licinus communis) oil, ceramide, ceresin, ceteareth-5, ceteareth-12, ceteareth-20, Cetaryl allate, ceteth-20, ceteth-24, cetyl acetate, cetyl octanoate, cetyl palmitate, chamomile (antemis nobilis) oil, cholesterol, cholesterol ester, cholesteryl hydroxystearate, citric acid, salvia (salvia sclare) Oil, cocoa (theobroma cacao) butter, coco-caprylic / capric acid, coconut (cocos nucifera) oil, collagen, collagen amino acid, corn oil, fatty acid, decyl oleate, dextrin, diazolidinyl urea, dimethi Corn copolyol, dimethiconol, dioctyl adipate, dioctyl succinate, hexacaprylic acid / dipentaerythrityl hexacaprate, DMDM hydantoin, DNA, erythritol, ethoxydiglyco , Ethyl linoleate, Eucalyptus globulus oil, evening primrose (Oenothera biennis) oil, fatty acid, fructose, gelatin, wild geranium oil, glucosamine, glutamic acid glucose, glutamic acid, glyceres-26, glycerin, glycerol, glyceryl distearate, hydroxystearin Glyceryl acid, glyceryl laurate, glyceryl linoleate, glyceryl myristate, glyceryl oleate, glyceryl stearate, glyceryl stearate SE, glycine, glycol stearate, glycol stearate SE, glycosaminoglycan, grape (Vitis vinifera) Seed oil, hazel (Corillas americana) nut oil, hazel (Corillas averana) nut oil, hexylene glycol, honey, hyal Acid, hybrid safflower (Casamas tinctorias) oil, hydrogenated castor oil, hydrogenated cocoglyceride, hydrogenated coconut oil, hydrogenated lanolin, hydrogenated lecithin, hydrogenated palm glyceride, hydrogenated palm kernel oil, hydrogenated large Bean oil, hydrogenated tallow glyceride, hydrogenated vegetable oil, hydrolyzed collagen, hydrolyzed elastin, hydrolyzed glycosaminoglycan, hydrolyzed keratin, hydrolyzed soy protein, hydroxylated lanolin, hydroxyproline, imidazolidinyl urea, iodopropynyl butyl carbamate , Isocetyl stearate, Isocetyl stearoyl stearate, Isodecyl oleate, Isopropyl isostearate, Isopropyl lanolinate, Isopropyl myristate, Isopropyl palmitate, Isopropyl stearate , Isostearamide DEA, isostearic acid, isostearyl lactate, isostearyl neopentanoate, jasmine (jasminum officinale) oil, jojoba (baxus chinensis) oil, kelp, kukui (aleurites molkana) nut oil, lactamide MEA, lanes- 16, lanes-10 acetate, lanolin, lanolin acid, lanolin alcohol, lanolin oil, lanolin wax, lavender (Labandula angstifolia) oil, lecithin, lemon (citrus medica limonam) oil, linoleic acid, linolenic acid, macadamia・ Ternifolia nut oil, magnesium stearate, magnesium sulfate, maltitol, chamomile (camomile / recuta) oil, methyl glucose sesquistearate, methylsilanol PCA, microcrystalline wax, mineral Oil, Mink oil, Mortierella oil, Myristyl lactate, Myristyl myristate, Myristyl propionate, Dicaprylic acid / Neopentyl glycol dicaprate, Octyl decanol, Octyl dodecyl myristate, Octyl dodecyl stearate stearate, Octyl hydroxy stearate, Palmitin Octyl acid, octyl salicylate, octyl stearate, oleic acid, olive (olea europaea) oil, orange (citrus aurantium durcis) oil, coconut (elaeis guiinensis) oil, palmitic acid, panthetin, panthenol, panthenyl Ethyl ether, paraffin, PCA, peach (Prunus persica) kernel oil, peanut (Arakis hippogae) oil, PEG-8 C12 18 ester, PEG-15 coca PEG-150 distearic acid, PEG-60 glyceryl isostearate, PEG-5 glyceryl stearate, PEG-30 glyceryl stearate, PEG-7 hydrogenated castor oil, PEG-40 hydrogenated castor oil, PEG-60 hydrogenated Castor oil, PEG-20 methyl sesquistearate, PEG-40 sorbitan peroleate, PEG-5 soy sterol, PEG-10 soy sterol, PEG-2 stearate, PEG-8 stearate, PEG-20 stearate, stearin Acid PEG-32, stearic acid PEG-40, stearic acid PEG-50, stearic acid PEG-100, stearic acid PEG-150, pentadecalactone, peppermint (menta piperita) oil, petrolatum, phospholipid, polyamino sugar condensate , Polyglyceryl-3 stearate, polyquaternium-24, polysorbate 20, polysorbate 40, polysorbate 60, polysorbate 80, polysorbate 85, potassium myristate, potassium palmitate, potassium sorbate, potassium stearate, propylene glycol, dicaprylic acid / propylene dicaprate Glycol, propylene glycol dioctanoate, propylene glycol dipelargonate, propylene glycol laurate, propylene glycol stearate, SE propylene glycol stearate, PVP, pyridoxine dipalmitate, quaternium-15, quaternium-18 hectorite, quaternium-22, retinol , Retinyl palmitate, rice (Oryza sativa) nuka oil, RNA, Rosemary (Rosmarinus officinalis) oil, rose oil, safflower (Casamas tinctorias) oil, sage (Salvia officinalis) oil, salicylic acid, sandalwood (Santaram album) oil, serine, serum protein, sesame (sesamum Indicam) oil, shea butter (butyrospermum parky), silk powder, sodium chondroitin sulfate, DNA sodium, sodium hyaluronate, sodium lactate, sodium palmitate, sodium PCA, sodium polyglutamate, sodium stearate, water-soluble collagen, sorbin Acid, sorbitan laurate, sorbitan oleate, sorbitan palmitate, sorbitan sesquioleate, sorbitan stearate, sorbitol, soybean (glycine soya) oil, sphingolipid Squalane, squalene, stearamide stearate MEA, stearic acid, stearoxydimethicone, stearoxytrimethylsilane, stearyl alcohol, stearyl glycyrrhetinate, stearyl heptanoate, stearyl stearate, sunflower (Helianthus anas) seed oil, sweet almond (Prunus・ Amygdalus durcis) oil, synthetic beeswax, tocopherol, tocopheryl acetate, tocopheryl linolenate, tribehenine, tridecyl neopentanoate, tridecyl stearate, triethanolamine, tristearin, urea, vegetable oil, water, wax, wheat (Triticum vulgare ) Germ oil and ylang ylang (kananga odorata) oil.

  In some embodiments, the topical formulation may contain suitable excipients, which typically have a high affinity for the skin, are well tolerated, It must produce a consistency that is stable and readily available. Suitable topical excipients and vehicles are described by those skilled in the art, particularly in Remington's Pharmaceutical Sciences, Vol. 18, Mack Publishing Co. Easton, Pa. (1990), particularly Chapter 87, can be routinely selected for a particular application with reference to one of many standard textbooks in the art. Optionally, one or more hygroscopic agents are also included in the topical formulation. Examples of hygroscopic agents include, but are not limited to, amino acids, chondroitin sulfate, diglycerin, erythritol, fructose, glucose, glycerin, glycerol, glycol, 1,2,6-hexanetriol, honey, hyaluronic acid, hydrogenated honey, Hydrogenated starch hydrolyzate, inositol, lactitol, maltitol, maltose, mannitol, natural moisturizing factor, PEG-15 butanediol, polyglyceryl sorbitol, pyrrolidone carboxylate, potassium PCA, propylene glycol, sodium glucuronate, sodium PCA, sorbitol, Sucrose, trehalose, urea, and xylitol.

  Certain embodiments contemplate topical formulations containing one or more additional skin protectants. Examples of skin protectants include, but are not limited to, seaweed extract, allantoin, aluminum hydroxide, aluminum sulfate, betaine, camelia sinensis leaf extract, cerebroside, dimethicone, glucuronolactone, glycerin, kaolin, lanolin, malt extract, mineral Mention may be made of oil, petroleum jelly, potassium gluconate and talc. It will be readily appreciated by those skilled in the art that skin protectants other than those listed above may be combined with the disclosed compositions of the invention or the preparations provided thereby.

  Surfactants may also be included in certain topical formulations contemplated herein, if desired, cationic, anionic, zwitterionic or nonionic surfactants, or Can be selected from any natural or synthetic surfactant suitable for use in cosmetic compositions (such as Rosen, M., “Surfactants and Interfacial Phenomena,” Second Edition, John Wiley & Sons, New York, NY) , 1988, Chapter 1p431). Examples of cationic surfactants include, but are not limited to, DMDAO or other amine oxides, long chain primary amines, diamines, polyamines and their salts, quaternary ammonium salts, polyoxyethylenated Mention may be made of long chain amines, as well as quaternized polyoxyethylenated long chain amines. Examples of anionic surfactants include, but are not limited to, SDS; carboxylic acid salts (eg, soaps); sulfonic acid salts, sulfuric acid salts, phosphoric acid and polyphosphate esters; alkyl phosphates; Mention may be made of the salts of perfluorocarboxylic acids. Examples of zwitterionic surfactants include cocamidopropyl hydroxysultain (CAPHS) and others that are pH sensitive and require special care in designing the optimum pH of the formulation (i.e. , Alkylaminopropionic acid, imidazoline carboxylate, and betaine) or those not pH sensitive (eg, sulfobetaine, sultaine). Examples of nonionic surfactants include, but are not limited to, alkylphenol ethoxylates, alcohol ethoxylates, polyoxyethylenated polyoxypropylene glycols, polyoxyethylenated mercaptans, long chain carboxylic acid esters, alkanols Mention may be made of amides, tertiary acetylene glycols, polyoxyethylenated silicones, N-alkylpyrrolidones, and alkyl polyglycosidases. Wetting agents, mineral oils or other surfactants such as non-ionic detergents or agents such as one or more of the PLURONIC® series (BASF, Mt Olive, NJ) can also be used, for example, according to non-limiting theory: It may be contained in order to prevent aggregation of BT fine particles in the fine particle suspension. Any combination of surfactants is acceptable. Certain embodiments are at least one anionic surfactant and at least one cationic surfactant that are compatible, ie, do not form a appreciable precipitate upon mixing, or It may comprise at least one cationic surfactant and at least one zwitterionic surfactant.

  Examples of thickeners that may also be present in certain topical formulations include, but are not limited to, acrylamide copolymer, agarose, amylopectin, bentonite, calcium alginate, carboxymethylcellulose calcium, carbomer, carboxymethylchitin, cellulose gum, dextrin , Gelatin, hydrogenated tallow, hydroxyethylcellulose, hydroxypropylcellulose, hydroxypropyl starch, magnesium alginate, methylcellulose, microcrystalline cellulose, pectin, various PEGs, polyacrylic acid, polymethacrylic acid, polyvinyl alcohol, various PPGs, sodium acrylate copolymer, carrageenan sodium, xanthan gum, and yeast β-glucan It is below. Thickeners other than those listed above may be used in embodiments of the present invention.

  According to certain embodiments contemplated herein, the topical formulation may include one or more sunscreens or UV absorbers. Where ultraviolet (UVA and UVB) absorption properties are desired, such agents include, for example, benzophenone, benzophenone-1, benzophenone-2, benzophenone-3, benzophenone-4, benzophenone-5, benzophenone-6, benzophenone-7, Benzophenone-8, benzophenone-9, benzophenone-10, benzophenone-11, benzophenone-12, benzyl salicylate, butyl PABA, cinnamate, cinoxalate, DEA-methoxycinnamate, diisopropyl methylcinnamate, ethyl dihydroxypropyl PABA, diisopropylcinnamic acid Ethyl, ethyl methoxycinnamate, ethyl PABA, ethyl urocanate, dimethoxycinnamate glyceryl octanoate, glyceryl PABA, salicylic acid glyco , Homosalate, p- methoxycinnamic acid isoamyl, titanium, zinc, zirconium, silicon, oxides of manganese and cerium, PABA, PABA esters, Parsol 1789, and isopropylbenzyl salicylate benzyl, and may include mixtures thereof. Those skilled in the art will appreciate that sunscreens and UV absorbers or protectants other than those listed may be used in certain embodiments of the invention.

  The topical formulations disclosed herein are typically effective at pH values between about 2.5 and about 10.0. Preferably, the pH of the composition is in the following pH range or approximate values thereof: about pH 5.5 to about pH 8.5, about pH 5 to about pH 10, about pH 5 to about pH 9, about pH 5 to about pH 8, about pH 3 To about pH 10, about pH 3 to about pH 9, about pH 3 to about pH 8, and about pH 3 to about pH 8.5. Most preferably, the pH is from about pH 7 to about pH 8. One skilled in the art may add appropriate pH adjusting ingredients to the compositions of the present invention to adjust the pH to an acceptable range. If it is recognized that the pH can vary from the original value depending on the composition and storage conditions of the formulation, the specific pH with “about” is the actual measured pH at any given time. One skilled in the art will appreciate that the formulation includes a formulation that can be raised or lowered by no more than 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, or 0.1 pH units above the desired value. I will.

  Creams, lotions, gels, ointments, pastes, etc. may be spread over the affected surface and gently rubbed. The solution may be applied in the same manner, but more typically it will be applied using a dropper, swab, etc. and will be carefully applied to the affected area. This application regimen will depend on many factors that can be readily determined, such as the severity of the wound and responsiveness to the initial treatment, but will usually involve more than one application daily. Let's go. One skilled in the art can readily determine the appropriate amount of formulation to be administered, the method of administration and the rate of repetition. In general, the formulations of these and related embodiments of the present invention are applied in a range from 1 or 2 or more times per week to 1, 2, 3, 4 or more times per day. Is contemplated.

  As also previously discussed, topical formulations useful herein thus also contain a pharmaceutically acceptable carrier, including any suitable diluent or excipient, and as such Any pharmaceutical agent that is not harmful to the subject to which the composition is administered can be administered without undue toxicity. Pharmaceutically acceptable carriers include, but are not limited to, liquids such as water, saline, glycerol and ethanol, and viscosity enhancers (eg, balsam fir resin) or film formers such as collodion or nitrocellulose solutions. Can also be included. A thorough discussion of pharmaceutically acceptable carriers, diluents, and other excipients is given in REMINGTON'S PHARMACEUTICAL SCIENCES (Mack Pub. Co., NJ, latest edition).

  Where the topical formulation is in the form of a gel- or liquid-filled capsule, such as a gelatin capsule, it may contain a liquid carrier such as polyethylene glycol or oil in addition to the previous type of material. The liquid pharmaceutical composition of certain embodiments of the present invention, whether in solution, suspension or other similar form, may include one or more of the following: a sterile diluent such as water for injection Saline solution, preferably physiological saline, Ringer's solution, isotonic sodium chloride, non-volatile oils such as synthetic monoglycerides or diglycerides which can serve as solvent or suspending medium, polyethylene glycol, glycerin, propylene glycol Or other solvents; antibacterial agents such as benzyl alcohol or methyl paraben; additional antioxidants such as ascorbic acid or sodium bisulfite; chelating agents such as ethylenediaminetetraacetic acid (EDTA); buffering agents such as acetate, citrate Or phosphate, and tonicity adjusting agents such as sodium chloride Or dextrose.

  For topical administration, the carrier may suitably comprise a solution, emulsion, ointment or gel base. The base may contain, for example, one or more of the following: petrolatum, lanolin, polyethylene glycol, beeswax, mineral oil, diluents such as water and alcohols, and emulsifiers and stabilizers. Thickeners may be present in pharmaceutical or cosmetic compositions for topical administration. If intended for transdermal administration, the composition may include a transdermal patch or iontophoresis device. The topical formulation may comprise a concentration of the compound of certain embodiments of the invention from about 0.1 to about 10% w / v (weight / unit volume). Topical formulations may be provided in the form of creams, lotions, solutions, sprays, gels, ointments, pastes, etc. and / or contain liposomes, micelles, microspheres and / or other particulate or nanoparticle delivery elements. It may be. Topical formulations are of time-release or sustained-release particles or pellets that can be administered directly to the wound site, such as sustained-release ethylene vinyl acetate polymer (eg, Elvax® 40, Aldrich, Milwaukee, WI) pellets. It may be provided in the form.

  The topical formulation may include an agent that binds to the skin tissue repair promoting compound, thereby assisting delivery to skin epithelial cells (eg, keratinocytes) and / or fibroblasts. Suitable drugs that can act in this capacity include inclusion agents such as cyclodextrins; other drugs may include proteins or liposomes.

  The topical formulations of certain embodiments of the invention may be provided in the form of dosage units that can be administered as an aerosol. The term aerosol is used to denote a variety of systems ranging from colloidal to systems consisting of pressurized packaging. Delivery may be by a liquefied or compressed gas or by a suitable pump system that disperses the active ingredients. Aerosols of the compounds of certain embodiments of the invention may be delivered in a single phase, two phase, or three phase system to deliver the active ingredient (s). The delivery of the aerosol includes the necessary containers, activators, valves, sub-containers, etc., which may be combined to form a kit. One skilled in the art may determine a preferred aerosol for delivering the topical formulation to the skin or wound site without undue experimentation.

  Topical formulations may be prepared by methodology well known in the pharmaceutical art. For example, a pharmaceutical composition that is to be administered as a spray, wash or rinse to a wound site or skin comprises a BT disinfection / wound healing / anti-biofilm / skin tissue repair-promoting compound as described herein in a solution. It can be prepared by mixing with sterile distilled water to form. A surfactant may be added to facilitate the formation of a homogeneous solution or suspension. A surfactant is a compound that interacts non-covalently with an antioxidant active compound to facilitate dissolution or homogeneous suspension of the compound in an aqueous delivery system.

  The BT disinfection / wound healing / anti-biofilm / skin tissue repair-promoting compound, or pharmaceutically acceptable salt thereof used in the topical formulation is administered in a therapeutically effective amount, The nature of the wound site (if relevant), the activity of the specific BT compound used (such as with or without an aminoglycoside antibiotic, eg a combination of antibiotics such as amikacin); metabolic stability and action of the compound Duration; subject age, weight, general health, gender, skin type, immune status and diet; mode of administration and time; excretion rate; drug combination; severity of specific skin wounds where skin tissue repair is desired; and It varies depending on various factors, including the treatment the subject is receiving. Generally, a therapeutically effective daily dose is (for a 70 kg mammal) about 0.001 mg / kg (ie 0.07 mg) to about 100 mg / kg (ie 7.0 g); preferably therapeutically effective A dose of about 0.01 mg / kg (ie 7 mg) to about 50 mg / kg (ie 3.5 g) (for a 70 kg mammal); more preferably a therapeutically effective dose (of 70 kg In mammals, it is about 1 mg / kg (ie 70 mg) to about 25 mg / kg (ie 1.75 g).

Effective dose ranges provided herein are not limiting and represent preferred dose ranges. However, the most preferred dosage will be understood by one of ordinary skill in the related art, as may be determined, will be adapted to each target (e.g., Berkow et al., Eds. , The Merk Manual 16 th edition, Merk and Co., Rahway, N.J., 1992 ;... Goodman et al, eds, Goodman and Gilman's The Pharmacological Basis of Therapeutics, 10 th edition, Pergamon Press, Inc., Elmsford, N.Y, ( 2001); Avery's Drug Treatment: Principles and Practice of Clinical Pharmacology a d Therapeutics, 3rd edition, ADIS Press, Ltd., Williams and Wilkins, Baltimore, MD. ^{s Pharmaceutical Sciences, 18 th edition,} Mack Publishing Co., Easton, PA (1990); Katzung, Basic and Clinical Pharmacology, Appleton and Lange, Norwalk, CT (1992) reference).

  If desired, the total dose required for each treatment can be administered during the day by repeated or single administration. Certain preferred embodiments contemplate applying a single topical formulation per day. In general, in obvious embodiments, treatment may be initiated with smaller dosages which are less than the optimum dose of the compound. Thereafter, the dosage is increased by small increments until the optimum effect under the circumstances is reached.

  The topical formulation can be administered alone or in conjunction with other treatments and / or medications directed to the skin wound or directed to other related symptoms or etiological factors. For example, as mentioned above, the topical formulation may further comprise retinoic acid. As another example, the topical formulation may include one or more skin tissue repair promoting compounds described herein, or includes two or more such compounds having different cellular wound repair activities. May be.

  The topical formulation recipient described herein may be any vertebrate, such as a mammal. Preferred recipients of mammals are primates (humans, apes, monkeys, etc.), cloven-hoofed animals (horses, goats, cows, sheep, pigs, etc.), rodents (mouses, rats, rabbits, hamsters, etc.) , And mammals (such as cats and dogs). Of the birds, preferred recipients are turkeys, chickens and other animals of the same eye. The most preferred recipient is a human, and particularly preferred is a human having an acute or chronic wound, or a wound comprising a biofilm.

  For topical application, according to embodiments described herein, an effective amount of a pharmaceutical composition comprising a BT compound disinfection / wound healing / anti-biofilm / skin tissue repair-promoting compound is applied to a target area, eg, It is preferred to administer to skin wounds such as acute or chronic wounds and / or areas at risk of skin (eg wound dehiscence). This amount generally depends on the particular embodiment of the invention per application, depending on the area to be treated, the severity of symptoms (or past or intended surgical resection), and the nature of the topical vehicle used. Of the compound will range from about 0.0001 mg to about 1 g. Preferred topical preparations are ointments or sustained release pellets, in which about 0.001 to about 50 mg of active ingredient is used per cc of ointment base or pellet suspension. The pharmaceutical composition can be formulated as a transdermal composition or transdermal delivery device (“patch”). Such compositions include, for example, a support, an active compound reservoir, a control membrane, a liner and a contact adhesive. Such transdermal patches may be used to provide continuous, pulsatile or on-demand delivery of the compounds of the invention if desired.

  Certain embodiments of the compositions can be formulated to provide rapid, sustained, or delayed release of the active ingredient after administration to a patient using procedures known in the art. Controlled release drug delivery systems include osmotic pump systems and dissolution systems containing polymer-coated reservoirs or drug polymer matrix formulations. Examples of controlled release systems include US Pat. Nos. 3,845,770 and 4,326,525, and J. et al. Kuzma et al, Regional Anesthesia 22 (3): 543-551 (1997), all of which are incorporated herein by reference.

  The most appropriate route will depend on the nature and severity of the condition being treated. Those skilled in the art will know how to administer topical methods (spray, cream, open application, closed dressing, soaking, washing, etc.), dosage forms, suitable pharmaceutical excipients, and compounds to subjects in need thereof. They are also familiar with determining other things related to delivery.

  Throughout this specification, unless stated otherwise, the languages “comprise”, “comprises”, and “including” include the stated step or element, or step or element group. However, it is to be understood that it does not exclude any other steps or elements, or steps or elements. “Consisting” means including the matter following the phrase “consisting of” and is limited thereto. In other words, the phrase “consisting of” indicates that the listed elements are necessary or required, and that no other elements can exist. “Consisting essentially of” is meant to include any element listed after the phrase, other that does not interfere with or contribute to the activity or action specified in the disclosure relating to the listed element Limited to elements. In other words, the phrase “consisting essentially of” requires or lists the listed elements, but does not require other elements, depending on whether it affects the activity or action of the listed elements. , May or may not be present.

  In this specification and the appended claims, the singular forms “a”, “an”, and “the” include plural references unless indicated otherwise. The term “about” or “approximately” as used in certain embodiments herein, when present before a numerical value, plus or minus 5%, 6%, 7%, 8% or 9% Indicates the range. In other embodiments, the term “about” or “approximately”, when present before a numerical value, indicates that value plus or minus 10%, 11%, 12%, 13%, or 14% of the range. In yet another embodiment, the term “about” or “approximately”, when present before a number, ranges from that value plus or minus 15%, 16%, 17%, 18%, 19%, or 20%. Indicates.

  The following examples are given by way of illustration and are not meant to be limiting.

Example Example 1
Preparation of BT compounds The following BT compounds were prepared according to the method of Domenico et al. (US RE 37,793, US 6,248,371, 6,086,921, 6,380,248). Or as microparticles according to the synthesis protocol described below for BisEDT. Shown is the atomic ratio for a single bismuth atom as a comparison, based on the theoretical ratio of the reactants used to form the trivalent complex with the sulfur-containing compound and the known propensity of bismuth. The numbers in parentheses are the ratio of bismuth to one (or several) thiol agent (eg Bi: thiol 1 / thiol 2; see also Table 1).
1) CPD 1B-1 Bis-EDT (1: 1) BiC ₂ H ₄ S ₂
2) CPD 1B-2 Bis-EDT (1: 1.5) BiC ₃ H ₆ S ₃
3) CPD 1B-3 Bis-EDT (1: 1.5) BiC ₃ H ₆ S ₃
4) CPD 1C Bis-EDT (soluble Bi preparation) (1: 1.5) BiC ₃ H ₆ S ₃
5) CPD 2A Bis-Bal (1: 1) BiC ₃ H ₆ S ₂ O
6) CPD 2B Bis-Bal (1: 1.5) BiC _4.5 H ₉ O _1.5 S ₃
7) CPD 3A Bis-Pyr (1: 1.5) BiC _7.5 H ₆ N _1.5 O _1.5 S _1.5
8) CPD 3B Bis-Pyr (1: 3) BiC ₁₅ H ₁₂ N ₃ O ₃ S ₃
9) CPD 4 Bis-Ery (1: 1.5) BiC ₆ H ₁₂ O ₃ S ₃
10) CPD 5 Bis-Tol (1: 1.5) BiC _10.5 H ₉ S ₃
11) CPD 6 Bis-BDT (1: 1.5) BiC ₆ H ₁₂ S ₃
12) CPD 7 Bis-PDT (1: 1.5) BiC _4.5 H ₉ S ₃
13) CPD 8-1 Bis-Pyr / BDT (1: 1/1)
14) CPD 8-2 Bis-Pyr / BDT (1: 1 / 0.5)
15) CPD 9 Bis-2 hydroxy, propanethiol (1: 3)
16) CPD 10 Bis-Pyr / Bal (1: 1 / 0.5)
17) CPD 11 Bis-Pyr / EDT (1: 1 / 0.5)
18) CPD 12 Bis-Pyr / Tol (1: 1 / 0.5)
19) CPD 13 Bis-Pyr / PDT (1: 1 / 0.5)
20) CPD 14 Bis-Pyr / Ery (1: 1 / 0.5)
21) CPD 15 Bis-EDT / 2 hydroxy, propanethiol (1: 1/1)

  Particulate bismuth-1,2-ethanedithiol (Bis-EDT, a soluble bismuth preparation) was prepared as follows:

To an excess of room temperature 5% aqueous HNO ₃ (11.4 L) in a 15 L polypropylene carboy, add aqueous Bi (NO ₃ ) ₃ solution (43% Bi (NO ₃ ) ₃ (w / w), 5% nitric acid ( w / w), 52% (w / w) water, Shepherd Chemical Co., Cincinnati, OH, product number 2362; δ˜1.6 g / mL) 0.331 L (˜0.575 mol) with stirring Was added dropwise and absolute ethanol (4 L) was added slowly. A small amount of white precipitate formed but dissolved upon continuous stirring. Separately, 72.19 mL (0.863 mol) of 1,2-ethanedithiol (CAS 540-63-6) was added to 1.5 L of absolute ethanol using a 60 mL syringe, followed by stirring for 5 minutes. An ethanolic solution of 1,2-ethanedithiol (˜1.56 L, ˜0.55 M) was prepared. The 1,2-ethanedithiol / EtOH reagent was then slowly added dropwise over 5 hours to the aqueous Bi (NO ₃ ) ₃ / HNO ₃ solution and continuously stirred overnight. The formed product was allowed to settle as a precipitate for approximately 15 minutes, after which the filtrate was removed at 300 mL / min using a peristaltic pump. The product was then recovered by filtration through a filter paper in a 15 cm diameter Buchner funnel and subsequently washed three times with 500 mL volumes each of ethanol, USP water and acetone to give BisEDT (694.51 g / mol) as a yellow amorphous. Obtained as a fine powder solid. The product was placed in a 500 mL brown glass bottle and dehydrated with CaCl ₂ under high vacuum for 48 hours. The recovered material (yield ~ 200 g) released an odor characteristic of thiols. The crude product was redissolved in 750 mL absolute ethanol, stirred for 30 minutes, then filtered, washed 3 times with 50 mL ethanol, 2 times 50 mL acetone, and washed again with 500 mL acetone. The rewashed powder was triturated in 1M NaOH (500 mL), filtered and washed 3 times with 220 ml of water, 2 times with 50 mL of ethanol and 1 time with 400 mL of acetone to give 156.74 g of purified BisEDT. . The next batch, prepared in essentially the same way, yielded a yield of about 78.91%.

The product was previously shown by analysis of data from ¹ H and ¹³ C nuclear magnetic resonance (NMR), infrared spectroscopy (IR), ultraviolet spectroscopy (UV), mass spectrometry (MS) and elemental analysis. Having the structure of formula I. An HPLC method was developed to determine the chemical purity of BisEDT, thereby preparing a sample in DMSO (0.5 mg / mL). Λ _max was measured by scanning a solution of BisEDT in DMSO at 190-600 nm. Waters (Millipore Corp., Milford, Mass.) With a UV detector monitoring at 265 nm (λ _max ) and equipped with a YMC Pack PVC Sil NP (inner diameter 250 × 4.6 mm) analytical column (Waters) with an injection volume of 2 μL. A model 2695 chromatograph was used to perform a gradient HPLC elution at 1 mL / min at ambient temperature with 0.1% formic acid in acetone: water (9: 1) as the mobile phase to detect a single peak. The chemical purity was 100 ± 01%. Elemental analysis was consistent with the structure of formula (I).

  The dry particulate material was characterized to evaluate particle size characteristics. Briefly, the microparticles are resuspended in 2% Pluronic® F-68 (BASF, Mt. Olive, NJ) and the suspension is sonicated for 10 minutes in a standard water bath sonicator. And then analyzed using a Nanosizer / Zetasizer Nano-S particle analyzer (model ZEN 1600 (without zeta potential measurement capability), Malverm Instruments, Worcestershire, UK) according to the manufacturer's recommendations. From the combined data of the two measurements, the microparticles show a unimodal distribution and all detectable events occur between volume mean diameter (VMD) 0.6 microns to 4 microns, peaking at about 1.3 microns Had VMD. On the other hand, when BisEDT was prepared by prior art methods (Domenico et al., 1997 Antimicrob. Agents Chemother. 41 (8): 1697-1703), the majority of the particles were heterodispersed and significantly larger in size. Characterization based on is no longer possible.

Example 2
Colony Biofilm Model of Chronic Wound Infection: Inhibition by BT Compounds Bacteria present in chronic wounds essentially follow the method described to adopt a biofilm lifestyle (Anderl et al., 2003 Antimicrob Agents Chemother). 47: 1251-56; Walters et al., 2003 Antimicrob Agents Chemother 47: 317; Wentland et al., 1996 Biotnol. Prog. 12: 316; Zheng et al., 2002 Antim. Using biofilm, BT can be compared with biofilm for the effect on bacterial cell survival. And inspected.

  Briefly, colony biofilms were grown on 10% trypsin soy agar for 24 hours and transferred to treatment containing Mueller Hinton plates. After the treatment, the biofilm was dispersed in peptone water containing 2% w / v glutathione (BT neutralization), serially diluted with peptone water, and then spotted on a counting plate. Two bacteria isolated from chronic wounds were used separately in generating a colony biofilm for testing. These were Pseudomonas aeruginosa Gram-negative strains and Gram-positive bacteria, methicillin-resistant Staphylococcus aureus (MRSA).

  Bacterial biofilm colonies were obtained essentially as described (Anderl et al., 2003 Antimicrob Agents Chemother 47: 1251-56; Walters et al., 2003 Antimicrob Agents Chemel 47.317 et al .; Prog. 12: 316; Zheng et al., 2002 Antimicrob Agents Chemother 46: 900), grown on microporous membranes on agar plates. Colony biofilms have shown many of the familiar properties of other biofilm models, for example, they were composed of cells that were densely aggregated in a highly hydrated matrix. As reported elsewhere (Brown et al., J Surg Res 56: 562; Millward et al., 1989 Microbios 58: 155; Such et al., 1995 J Pharm Pharmacol 47: 1094; Thrower et al. Med Microbiol 46;: 425), it was observed that the bacteria in the colony biofilm also showed significantly reduced antimicrobial susceptibility, as quantified in the higher performance in vitro biofilm reactor. The majority of colony biofilms occurred immediately and reproducibly. According to a non-limiting theory, this colony biofilm model shares some of the characteristics of infected wounds, and the bacteria are in the air with nutrients supplied from below the biofilm and a minimum amount of fluid. Developed at the interface. Various nutrient sources, such as blood agar, thought to mimic in vivo nutrient conditions, were used to culture the colony biofilm.

Colony biofilms were prepared by inoculating a 25 mm diameter polycarbonate filter membrane with a 5 μl spot of planktonic bacterial liquid medium. Prior to inoculation, the membrane was sterilized by exposing it to UV light for 10 minutes per side. The inoculum was grown overnight in 37 ° C. bacterial medium, diluted with fresh medium to an optical density of 0.1 at 600 nm, and then attached to the membrane. The membrane was then placed on an agar plate containing a growth medium. Thereafter, the plate was covered and inverted in a 37 ° C. incubator. Every 24 hours, membranes and colony biofilms were transferred to new plates using sterile tweezers. Colony biofilms were typically used for experiments 48 hours after development, at which time there were approximately 10 ⁹ bacteria per membrane. The colony biofilm method has been successfully used to culture a wide variety of single and mixed species biofilms.

To determine susceptibility to antimicrobial agents (eg, BT compounds such as BT compound combinations; antibiotics; and BT compounds-antibiotic combinations), colony biofilms are supplemented with candidate antimicrobial treatments Transferred to agar plates. When the duration of exposure to antimicrobial treatment exceeded 24 hours, the colony biofilm was transferred daily to a new treatment plate. At the end of the treatment period, the colony biofilm was placed in a test tube containing 10 ml of buffer and vortexed for 1-2 minutes to disperse the biofilm. In some instances, the sample had to be easily treated with a tissue homogenizer to break up cell aggregates. The resulting cell suspension was then serially diluted and allowed to stand and the surviving bacteria were counted and reported as colony forming units (CFU) per unit area. Survival data was analyzed using log ₁₀ transformation.

  For each type of bacterial biofilm colony culture (Pseudomonas aeruginosa, PA; methicillin resistant Staphylococcus aureus, MRSA or SA), 5 antibiotics and 13 BT compounds were tested. Antimicrobial agents tested against PA include BisEDT and BT referred to herein as compounds 2B, 4, 5, 6, 8-2, 9, 10, 11 and 15 (see Table 1) and antibiotics And tobramycin, amikacin, imipenim, cefazolin, and ciprofloxane. Antimicrobial agents tested against SA include BisEDT and BT called compounds 2B, 4, 5, 6, 8-2, 9, 10 and 11 (see Table 1) and antibiotics rifampicin, daptomycin, minocycline, Ampicillin and vancomycin were included. Antibiotics were tested at concentrations approximately 10-400 times the minimum inhibitory concentration (MIC) according to established microbiological methodology, as described previously in the “Brief Description of the Drawings”.

  Seven BT compounds showed significant effects on PA bacterial survival at the tested concentrations, and two BT compounds demonstrated significant effects on MRSA survival at the tested concentrations; bacterial survival Representative results showing the effect of BT on BD are shown in FIG. 1 for BisEDT and BT compound 2B (tested against PA) and in FIG. 2 for BT compounds 2B and 8-2 (tested against SA) Both are relative to the effects of the indicated antibiotics. As also shown in FIGS. 1 and 2, inclusion of the indicated BT compound in combination with the indicated antibiotic provides a synergistic effect, whereby the ability to reduce bacterial survival is either antibiotic alone or BT compound It was enhanced compared to any single antimicrobial effect. Effect obtained with the combination of 1600 μg / mL AMK + 80 μg / mL BisEDT on Compound 15 (BisEDT / 2-hydroxy, propanethiol (1: 1/1)) at a concentration of 80 μg / mL in the PA survival assay (FIG. The effect comparable to 1) (not shown) was shown.

Example 3
Inhibition of chronic wound infection by drip flow biofilm model BT compounds Drip flow biofilms are recognized and reliable models for forming bacterial biofilms and testing the effects of candidate antimicrobial compounds on them. is there. The drip flow biofilm is produced on a coupon (substrate) placed in the drip flow reactor groove. Many different types of materials, such as frosted glass microscope slides, can be used as substrates for bacterial biofilm formation. The nutrient liquid medium is dropped into the cell chamber of the drip flow bioreactor near the top to place the medium into the chamber and then run at a 10 ° C. ramp over the length of the coupon.

Biofilms are grown in a drip flow bioreactor, BT compounds alone or in combination with antibiotic compounds and / or antibiotic compounds alone or in combination with other antibacterial agents including BT compounds Or other conventional or candidate treatments for chronic wounds. Thus, the BT compound is characterized for its effect on bacterial biofilm in the drip flow reactor. Biofilms in drip flow reactors are prepared according to established methodologies (eg, Stewart et al., 2001 J Appl Microbiol. 91: 525; Xu et al., 1998 Appl. Environ. Microbiol. 64: 4035). . This design involves culturing the biofilm on a polystyrene coupon tilted in a covered chamber. An exemplary medium is 5% v / v adult donor bovine serum (pH 6.8) that mimics conditions of high serum protein and very low iron concentrations, similar to in vivo biofilm growth conditions such as chronic wounds. Contains supplemented 1 g / l glucose, 0.5 g / l NH ₄ NO ₃ , 0.25 g / l KCl, 0.25 g / l KH ₂ PO ₄ , 0.25 g / l MgSO ₄ .7H ₂ O Yes. The medium is added dropwise (5 ml / hour) onto four coupons contained in four separate parallel chambers, each 10 cm × 1.9 cm and 1.9 cm deep. A reactor divided into chambers was assembled with polysulfone plastic. Each chamber is fitted with an individual removable plastic lid that can be tightly closed. The biofilm reactor is placed in a 37 ° C. incubator and warmed by passing the bacterial cell medium through an aluminum heating bath held in the incubator. This method mimics the antibiotic-resistant phenotype observed in certain biofilms, mimicking the low fluid shear environment and the proximity to the air interface characteristics of chronic wounds, while It is compatible with numerous analytical methods that provide continuous supplementation and characterize and monitor the effectiveness of the introduced candidate antimicrobial regimen. The drip flow reactor has been successfully used to culture a wide variety of pure and mixed species biofilms. Biofilms are typically grown for 2-5 days before an antimicrobial agent is applied.

  To measure the effect of an anti-biofilm agent on a biofilm grown in a drip flow reactor, the fluid flow passing over the biofilm is mixed with the desired treatment formulation (eg, one or more BT compounds and / or Or modified or supplemented with one or more antibiotics, or controls, and / or other candidate agents). The flow is continued for a specific treatment period. The treated biofilm coupon is then briefly removed from the reactor and scraped into a beaker containing 10 ml of buffer. Samples are briefly treated with a tissue homogenizer (typically 30 seconds to 1 minute) to disperse bacterial aggregates. The suspension is serially diluted to enumerate viable microorganisms according to standard microbiological methodology.

Example 4
Inhibition of keratinocyte scratch repair by wound biofilm: Biofilm suppression by BT compounds This example modifies an established in vitro keratinocyte scratch model of wound healing to provide biofilm-related wound pathology And relevance to wound healing, in particular to reach a model with relevance to acute or chronic wounds or wound-containing biofilms as described herein. With the keratinocyte scratch model for the effects of chronic wound biofilms, the culture of mammalian (eg, human) keratinocytes and bacterial biofilm populations proceeds in separate chambers in fluid contact with each other, Allows assessment of the effect of conditions that affect the effect of soluble components assimilated by the biofilm on keratinocyte wound healing events.

  Neonatal human foreskin cells are cultured as a monolayer in a treated plastic dish in which controlled “wounds” or scratches are formed by mechanical means (eg, sterile scalpel, razor, By means of a monophasic physical disruption, such as scratching an essentially straight cell-free zone between regions of the monolayer using a suitable tool such as a cell scraper, tweezers, or other instrument). In vitro keratinocyte monolayer model systems are known to undergo cellular structural and functional processes in response to wound events in a manner that mimics wound healing in vivo. According to the embodiments disclosed herein, the effect of the presence of bacterial biofilm on such a process, for example on the healing time of the scratch, was observed and selected in these and related embodiments. The effect of the presence of candidate antimicrobial (eg, antibacterial and antibiofilm) treatments is also evaluated.

  Wounded keratinocyte monolayers cultured in the presence of biofilms are investigated by morphological, biochemical, molecular genetics, cell physiology and other parameters, and the introduction of BT compounds is Decide whether to alter the distracting effects of the film (eg, increase or decrease in a statistically significant manner relative to the appropriate control). In order to evaluate the effect of such treatment on the impact of biofilm on the model wound healing process after examining the toxicity of each BT compound treatment, the wound was first designed with each BT compound alone and BT compound. Expose to combination.

  In an exemplary embodiment, a membrane (eg, a TransWell membrane insert) retained in a previous tissue culture well and in fluid communication with a keratinocyte monolayer that is scratched to initiate the wound healing process. Above, the third day biofilm is cultured. Biofilms cultured from genuine acute or chronic wounds are contemplated for use in these and related embodiments.

  Thus, an in vitro system has been developed to evaluate the effect of soluble biofilm components on human keratinocyte migration and proliferation. The system separates biofilm and keratinocytes using a dialysis membrane. Keratinocytes are cultured from neonatal foreskin as previously described (Flekman et al., 1997 J Invest. Dermatol. 109: 36; Pieporn et al., 1987 J Invest. Dermatol. 88: 215-219) Grow as a confluent monolayer on a glass cover slip. The monolayer of keratinocytes can then be scratched to create a uniform width “wound” and then the cell repair process can be monitored (eg, Tao et al., 2007 PLoS ONE 2: e697; Buth et al., 2007 Eur. J Cell Biol. 86: 747; Phan et al., 2000 Ann. Acad. Med. Singapore 29:27). Then, using aseptic technique, the artificial wound is placed in the bottom of a sterile double-sided chamber and the chamber is assembled. A keratinocyte growth medium (EpiLife) with or without antibiotics and / or bismuth-thiol is filled on both sides of the chamber. An uninoculated system is used as a control.

  Bacteria isolated from the wound are inoculated into the system and incubated for 2 hours in static conditions to allow the bacteria to adhere to the surface in the upper chamber. Following the attachment period, liquid medium flow is initiated in the upper chamber to remove unattached cells. Thereafter, the flow of the medium is continued at a rate that minimizes the growth of the plactonic cells in the upper chamber by washing out unattached cells. After an incubation time in the range of 6 to 48 hours, the system (keratinocyte monolayer on the coverslip and bacterial biofilm on the membrane substrate) is disassembled and the coverslip is removed and analyzed. In a related embodiment, after the mature biofilm is grown in the upper chamber, the chamber is assembled. In other related embodiments, changing at least one indicator of scratch healing (e.g., statistically relative to an appropriate control), such as the time that elapses during wound repair or other signs of wound repair. Candidate agents such as BT compounds or potentially synergistic BT compounds + antibiotic combinations for keratinocyte scratch repair (to increase or decrease in a significant manner). For example, BT compounds provided herein, such as BT provided in particulate form, as well as amikacin, ampicillin, cefazoline, cefepime, chloramphenicol, ciprofloxacin, clindamycin (or other lincosamide antibiotics ), Daptomycin (Cubicin®), doxycycline, gatifloxacin Biofilm and scraping to determine the effects of gentamicin, imipenim, levofloxacin, linezolid (Zyvox®), minocycline, nafcillin, paromomycin, rifampine, sulfamethoxazole, tobramycin and vancomycin. Separate co-cultures of wounded keratinocyte monolayers are performed in the absence or presence of one or more BT compounds, optionally with or without one or more antibiotics (eg, Tao et al., 2007 PLoS ONE 2: e697; Buth et al., 2007 Eur.J Cell Biol.86: 747; Phan et al., 2000 Ann. Acad. Med. Singapore 29:27).

Example 5
Inhibition of keratinocyte scratch repair by wound biofilm Isolated human keratinocytes were cultured on glass coverslips to create scratches according to the methodology described in Example 4 above. The wounded culture was maintained on a membrane support in fluid communication with the keratinocyte culture, under culture conditions alone or in the presence of a co-cultured biofilm. The time interval between scratch closure was then measured while cell growth and / or migration of keratinocytes re-established the keratinocyte monolayer throughout the scratch zone. FIG. 3 shows the effect of biofilm fluid communication (but not in direct contact) on the healing time of a scratched keratinocyte monolayer.

  Thus, in certain embodiments, scratched cell (eg, keratinocytes or fibroblasts) monolayers are cultured in the presence of a bacterial biofilm with or without a candidate anti-biofilm agent. And assessing an indication of healing of a scratched cell monolayer in the absence and presence of a candidate anti-biofilm agent, wherein the at least one indication of healing is promoted Drugs (eg, BT compounds such as the substantially monodispersed BT microparticle suspensions described herein alone or antibiotics such as amikacin, ampicillin, cefazoline, cefepime, chloramphenicol, ciprofl Loxacin, clindamycin, daptomycin (Cubicin®), doxycycline, gatifloxacin, gentamicin , Imipenim, levofloxacin, linezolid (Zyvox®), minocycline, nafcillin, paromomycin, rifampine, sulfamethoxazole, tobramycin and vancomycin in combination with one or more acute or chronic wounds Or a method of identifying an agent that treats a chronic wound, identified as an appropriate agent to treat a wound comprising a biofilm.

Example 6
Synergistic Bismuth-Thiol (BT) Antibiotic Combination This example comprises one or more bismuth-thiol compounds and one or more antibiotics for various bacterial species and strains, such as multiple antibiotic-resistant bacteria. An example of the demonstrated synergistic effect of the combination is shown.

  Materials and Methods susceptibility testing, NCCLS protocol (National Committee for Clinical Laboratory Standards (1997) Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria that Grow Aerobically:... Approved Standard M7-A2 and Informational Supplement M100-S10 NCCLS, Wayne, PA USA), and was performed by broth dilution in 96-well tissue culture plates (Nalge Nunc International, Denmark).

Briefly, overnight bacterial cultures were utilized to prepare a 0.5 McFarland standard suspension that was further 1:50 in cation-conditioned Mueller-Hinton broth medium (BBL, Cockeysville, MD, USA). Diluted (˜2 × 10 ⁶ cfu / ml). BT (prepared as described above) and antibiotics were added at increasing concentrations to keep the final volume constant at 0.2 mL. Cultures were incubated at 37 ° C. for 24 hours and turbidity was assessed by absorbance at 630 nm using an ELISA plate reader (Biotek Instruments, Winooski, VT, USA) according to manufacturer's recommendations. The minimum inhibitory concentration (MIC) was expressed as the lowest drug concentration that prevented growth for 24 hours. Viable bacterial counts (cfu / mL) were determined by standard plating on nutrient agar. The minimum bactericidal concentration (MBC) was expressed as the drug concentration that reduced initial viability by 99.9% at 24 hours after inoculation.

  The activity of the antimicrobial combination was evaluated using the checkerboard method. Partial Inhibitory Concentration Index (FICI) and Partial Bactericidal Concentration Index (FBCI) were determined according to Eliopoulos et al. (Antibiotics in Laboratory Medicine (Lorian, V., Ed.), Pp. 330-96, Williams and Wilkins, Balm US, Balm US, Of Eliopoulos and Meollering, (1996) Antimicrobial combina- tions. Synergy is defined as a FICI or FBCI index of 0.5 or less, 0.5 to 4 or less, no interaction, 4 or more defined as antagonistic (Odds, FC (2003) Synergy, antagonistism) , And what the chequebord puts between them.Journal of Antibiotic Chemotherapy 52: 1). As before, a reduction of more than 4 times the antibiotic concentration was also defined as synergy.

ＢＥ＝０．２μｇ／ｍｌ ＢｉｓＥＤＴ；菌株は、Ｗｉｎｔｈｒｏｐ−Ｕｎｉｖｅｒｓｉｔｙ Ｈｏｓｐｉｔａｌ， Ｍｉｎｅｏｌａ， ＮＹのＣｌｉｎｉｃａｌ Ｍｉｃｒｏｂｉｏｌｏｇｙ Ｌａｂｏｒａｔｏｒｙから得た。ナフシリンは、Ｓｉｇｍａ（Ｓｔ． Ｌｏｕｉｓ， ＭＯ）から得た。

ＢＥ＝０．２μｇ／ｍｌ ＢｉｓＥＤＴ；菌株は、Ｗｉｎｔｈｒｏｐ−Ｕｎｉｖｅｒｓｉｔｙ Ｈｏｓｐｉｔａｌ， Ｍｉｎｅｏｌａ， ＮＹのＣｌｉｎｉｃａｌ Ｍｉｃｒｏｂｉｏｌｏｇｙ Ｌａｂｏｒａｔｏｒｙから得た。ナフシリンは、Ｓｉｇｍａから得た。

ＢＥ＝０．２μｇ／ｍｌ ＢｉｓＥＤＴ；菌株Ｓ２４４６−３は、Ｗｉｎｔｈｒｏｐ−Ｕｎｉｖｅｒｓｉｔｙ Ｈｏｓｐｉｔａｌ， Ｍｉｎｅｏｌａ， ＮＹのＣｌｉｎｉｃａｌ Ｍｉｃｒｏｂｉｏｌｏｇｙ Ｌａｂｏｒａｔｏｒｙから得た。抗体は、Ｓｉｇｍａから得た。

ＧＭ＝ゲンタマイシン；菌株Ｓ２４００−１は、Ｗｉｎｔｈｒｏｐ−Ｕｎｉｖｅｒｓｉｔｙ Ｈｏｓｐｉｔａｌ， Ｍｉｎｅｏｌａ， ＮＹのＣｌｉｎｉｃａｌ Ｍｉｃｒｏｂｉｏｌｏｇｙ Ｌａｂｏｒａｔｏｒｙから得た。ゲンタマイシンは、ＷｉｎｔｈｒｏｐのＰｈａｒｍａｃｙ Ｄｅｐａｒｔｍｅｎｔから得た。太字は相乗性。

μｇ／ｍｌでのデータ；菌株Ｓ２４００−１は、Ｗｉｎｔｈｒｏｐ−Ｕｎｉｖｅｒｓｉｔｙ Ｈｏｓｐｉｔａｌ， Ｍｉｎｅｏｌａ， ＮＹのＣｌｉｎｉｃａｌ Ｍｉｃｒｏｂｉｏｌｏｇｙ Ｌａｂｏｒａｔｏｒｙから得た。抗生物質は、ＷｉｎｔｈｒｏｐのＰｈａｒｍａｃｙ Ｄｅｐａｒｔｍｅｎｔから得た。

μｇ／ｍｌでのデータ；菌株Ｓ２４００−１は、Ｗｉｎｔｈｒｏｐ−Ｕｎｉｖｅｒｓｉｔｙ Ｈｏｓｐｉｔａｌ， Ｍｉｎｅｏｌａ， ＮＹのＣｌｉｎｉｃａｌ Ｍｉｃｒｏｂｉｏｌｏｇｙ Ｌａｂｏｒａｔｏｒｙから得た。抗生物質は、ＷｉｎｔｈｒｏｐのＰｈａｒｍａｃｙ Ｄｅｐａｒｔｍｅｎｔから得た。

μｇ／ｍｌでのデータ；菌株Ｓ２４００−１は、Ｗｉｎｔｈｒｏｐ−Ｕｎｉｖｅｒｓｉｔｙ Ｈｏｓｐｉｔａｌ， Ｍｉｎｅｏｌａ， ＮＹのＣｌｉｎｉｃａｌ Ｍｉｃｒｏｂｉｏｌｏｇｙ Ｌａｂｏｒａｔｏｒｙから得た。抗生物質は、ＷｉｎｔｈｒｏｐのＰｈａｒｍａｃｙ Ｄｅｐａｒｔｍｅｎｔから得た。

μｇ／ｍｌでのデータ；抗生物質は、Ｗｉｎｔｈｒｏｐ−Ｕｎｉｖｅｒｓｉｔｙ Ｈｏｓｐｉｔａｌ， Ｍｉｎｅｏｌａ， ＮＹのＰｈａｒｍａｃｙ Ｄｅｐａｒｔｍｅｎｔから得た。

ＡＢ＝抗生物質；ＣＭ＝クロラムフェニコール；ＡＭ＝アンピシリン；ＢＥ＝０．３μｇ／ｍｌのＢｉｓＥＤＴ、菌株は、Ｄｅｐａｒｔｍｅｎｔ ｏｆ Ｍｏｌｅｃｕｌａｒ Ｇｅｎｅｔｉｃｓ ａｎｄ Ｃｅｌｌ Ｂｉｏｌｏｇｙ，Ｔｈｅ Ｕｎｉｖｅｒｓｉｉｔｙ ｏｆ Ｃｈｉｃａｇｏ， Ｃｈｉｃａｇｏ，ＩＬのＤｒ． ＭＪ Ｃａｓａｄａｂａｎの研究室から得た。抗生物質は、Ｗｉｎｔｈｒｏｐ−Ｕｎｉｖｅｒｓｉｔｙ Ｈｏｓｐｉｔａｌ， Ｍｉｎｅｏｌａ， ＮＹのＰｈａｒｍａｃｙ Ｄｅｐａｒｔｍｅｎｔから得た。

ＤＯＸ＝ドキシサイクリン；ＢＥ＝０．３μｇ／ｍｌのＢｉｓＥＤＴ；菌株は、Ｄｅｐａｒｔｍｅｎｔ ｏｆ Ｂａｃｔｅｒｉｏｌｏｇｙ， Ｔｈｅ Ｕｎｉｖｅｒｓｉｔｙ ｏｆ Ｂｒｉｓｔｏｌ， Ｂｒｉｓｔｏｌ， ＵＫのＤｒ．Ｉ Ｃｈｏｐｒａの研究室から得た。抗生物質は、Ｗｉｎｔｈｒｏｐ−Ｕｎｉｖｅｒｓｉｔｙ Ｈｏｓｐｉｔａｌ， Ｍｉｎｅｏｌａ， ＮＹのＰｈａｒｍａｃｙ Ｄｅｐａｒｔｍｅｎｔから得た。

Ａｇｒ＝アミノグリコシド耐性；ＮＮ＝トブラマイシン；ＰＡ＝シュードモナス・エルギノーサ；ＢＥ＝ＢｉｓＥＤＴ、０．３μｇ／ｍｌ；菌株は、Ｄｅｐａｒｔｍｅｎｔ ｏｆ Ｍｉｃｒｏｂｉｏｌｏｇｙ ａｎｄ Ｉｍｍｕｎｏｌｏｇｙ， Ｑｕｅｅｎｓ Ｕｎｉｖｅｒｓｉｔｙ， Ｏｎｔａｒｉｏ， ＣＮのＤｒ． Ｋ． Ｐｏｏｌｅの研究室から得た。トブラマイシンは、Ｗｉｎｔｈｒｏｐ−Ｕｎｉｖｅｒｓｉｔｙ Ｈｏｓｐｉｔａｌ， Ｍｉｎｅｏｌａ， ＮＹのＰｈａｒｍａｃｙ Ｄｅｐａｒｔｍｅｎｔから得た。

ＮＮ＝トブラマイシン；ＢＥ＝ＢｉｓＥＤＴ、０．４μｇ／ｍｌ；菌株は、Ｄｅｐａｒｔｍｅｎｔ ｏｆ Ｐｅｄｉａｔｒｉｃｓ ａｎｄ Ｃｏｍｍｕｎｉｃａｂｌｅ Ｄｉｓｅａｓｅ， Ｕｎｉｖｅｒｓｉｔｙ ｏｆ Ｍｉｃｈｉｇａｎ， Ａｎｎ Ａｒｂｏｒ， ＭＩのＤｒ． Ｊ． Ｊ． ＬｉＰｕｍａの研究室から；そして同じくＶｅｌｏｉｒａ．他，２００３により得た。トブラマイシンは、Ｗｉｎｔｈｒｏｐ−Ｕｎｉｖｅｒｓｉｔｙ Ｈｏｓｐｉｔａｌ， Ｍｉｎｅｏｌａ， ＮＹのＰｈａｒｍａｃｙ Ｄｅｐａｒｔｍｅｎｔから得た。

ＮＮ＝トブラマイシン；ＢＥ＝ＢｉｓＥＤＴ、０．４μｇ／ｍｌ；菌株は、Ｄｅｐａｒｔｍｅｎｔ ｏｆ Ｐｅｄｉａｔｒｉｃｓ ａｎｄ Ｃｏｍｍｕｎｉｃａｂｌｅ Ｄｉｓｅａｓｅ， Ｕｎｉｖｅｒｓｉｔｙ ｏｆ Ｍｉｃｈｉｇａｎ， Ａｎｎ Ａｒｂｏｒ， ＭＩのＤｒ． Ｊ． Ｊ． ＬｉＰｕｍａの研究室から；そして同じくＶｅｌｏｉｒａ．他，２００３により得た。トブラマイシンは、Ｗｉｎｔｈｒｏｐ−Ｕｎｉｖｅｒｓｉｔｙ Ｈｏｓｐｉｔａｌ， Ｍｉｎｅｏｌａ， ＮＹのＰｈａｒｍａｃｙ Ｄｅｐａｒｔｍｅｎｔから得た。

ＮＮ＝トブラマイシン；ＢＥ＝ＢｉｓＥＤＴ、０．８μｇ／ｍｌ；Ｌｉｐｏ−ＢＥ−ＮＮ＝リポソームＢＥ−ＮＮ；菌株は、Ｄｅｐａｒｔｍｅｎｔ ｏｆ Ｃｈｅｍｉｓｔｒｙ ａｎｄ Ｂｉｏｃｈｅｍｉｓｔｒｙ， Ｌａｕｒｅｎｔｉａｎ Ｕｎｉｖｅｒｓｉｔｙ， Ｏｎｔａｒｉｏ， ＣＮのＤｒ． Ａ． Ｏｍｒｉの研究室から得た（Ｍ株は、粘液様Ｂ．セパシア；ＰＡ＝Ｐ．エルギノーサ；ＳＡ＝Ｓ．アウレウス）。トブラマイシンは、Ｗｉｎｔｈｒｏｐ−Ｕｎｉｖｅｒｓｉｔｙ Ｈｏｓｐｉｔａｌ， Ｍｉｎｅｏｌａ， ＮＹのＰｈａｒｍａｃｙ Ｄｅｐａｒｔｍｅｎｔから得た。

ＮＮ＝トブラマイシン；ＢＥ＝ＢｉｓＥＤＴ、０．８μｇ／ｍｌ；Ｌｉｐｏ−ＢＥ−ＮＮ＝リポソームＢＥ−ＮＮ；菌株は、Ｄｅｐａｒｔｍｅｎｔ ｏｆ Ｃｈｅｍｉｓｔｒｙ ａｎｄ Ｂｉｏｃｈｅｍｉｓｔｒｙ， Ｌａｕｒｅｎｔｉａｎ Ｕｎｉｖｅｒｓｉｔｙ， Ｏｎｔａｒｉｏ， ＣＮのＤｒ． Ａ． Ｏｍｒｉの研究室から得た（Ｍ株は、粘液様Ｂ．セパシア；ＰＡ＝Ｐ．エルギノーサ；ＳＡ＝Ｓ．アウレウス）。トブラマイシンは、Ｗｉｎｔｈｒｏｐ−Ｕｎｉｖｅｒｓｉｔｙ Ｈｏｓｐｉｔａｌ， Ｍｉｎｅｏｌａ， ＮＹのＰｈａｒｍａｃｙ Ｄｅｐａｒｔｍｅｎｔから得た。

ＢＥ＝ＢｉｓＥＤＴ；ＮａＰＹＲ＝ナトリウムピリチオン；化学薬品は、Ｓｉｇｍａ Ａｌｄｒｉｃｈから得た。太字は相乗性。示された菌株は、Ａｍｅｒｉｃａｎ Ｔｙｐｅ Ｃｕｌｔｕｒｅ Ｃｏｌｌｅｃｔｉｏｎ （ＡＴＣＣ， Ｍａｎａｓｓａｓ， ＶＡ）からのものであった。 The results are shown in Tables 2-17.

BE = 0.2 μg / ml BisEDT; The strain was obtained from the Clinical Microbiology Laboratory of Winthrop-University Hospital, Mineola, NY. Nafcillin was obtained from Sigma (St. Louis, MO).

BE = 0.2 μg / ml BisEDT; The strain was obtained from the Clinical Microbiology Laboratory of Winthrop-University Hospital, Mineola, NY. Nafcillin was obtained from Sigma.

BE = 0.2 μg / ml BisEDT; Strain S2446-3 was obtained from Clinical Microbiology Laboratory of Winthrop-University Hospital, Minnea, NY. The antibody was obtained from Sigma.

GM = gentamicin; Strain S2400-1 was obtained from the Clinical Microbiology Laboratory of Wintrop-University Hospital, Mineola, NY. Gentamicin was obtained from Pharmatrop Department of Winthrop. Bold is synergistic.

Data in μg / ml; Strain S2400-1 was obtained from the Clinical Microbiology Laboratory of Wintrop-University Hospital, Mineola, NY. Antibiotics were obtained from Winthrop's Pharmacy Department.

Data in μg / ml; Strain S2400-1 was obtained from the Clinical Microbiology Laboratory of Wintrop-University Hospital, Mineola, NY. Antibiotics were obtained from Winthrop's Pharmacy Department.

Data in μg / ml; Strain S2400-1 was obtained from the Clinical Microbiology Laboratory of Wintrop-University Hospital, Mineola, NY. Antibiotics were obtained from Winthrop's Pharmacy Department.

Data in μg / ml; Antibiotics were obtained from Pharmacy Department of Winthrop-University Hospital, Mineola, NY.

AB = antibiotics; CM = chloramphenicol; AM = ampicillin; BE = 0.3 μg / ml BisEDT, strains are from Department of Molecular Genetics and Cell Biology, The University of Chicago, IL, IL. Obtained from the laboratory of MJ Casadaban. Antibiotics were obtained from Pharmacy Department of Winthrop-University Hospital, Mineola, NY.

DOX = doxycycline; BE = 0.3 μg / ml BisEDT; strains are from Department of Bacteriology, The University of Bristol, Bristol, UK Obtained from I Chopra's laboratory. Antibiotics were obtained from Pharmacy Department of Winthrop-University Hospital, Mineola, NY.

Agr = aminoglycoside resistance; NN = tobramycin; PA = Pseudomonas aeruginosa; BE = BisEDT, 0.3 μg / ml; strains are from Department of Microbiology and Immunology, Queens University, Ontario, K. Obtained from Poole laboratory. Tobramycin was obtained from Pharmacy Department of Winthrop-University Hospital, Mineola, NY.

NN = tobramycin; BE = BisEDT, 0.4 μg / ml; strains are from Dr. of Department of Pediatrics and Communicable Disease, University of Michigan, Ann Arbor, MI. J. et al. J. et al. From LiPuma's laboratory; and also Veloira. Et al., 2003. Tobramycin was obtained from Pharmacy Department of Winthrop-University Hospital, Mineola, NY.

NN = tobramycin; BE = BisEDT, 0.4 μg / ml; strains are from Dr. of Department of Pediatrics and Communicable Disease, University of Michigan, Ann Arbor, MI. J. et al. J. et al. From LiPuma's laboratory; and also Veloira. Et al., 2003. Tobramycin was obtained from Pharmacy Department of Winthrop-University Hospital, Mineola, NY.

NN = tobramycin; BE = BisEDT, 0.8 μg / ml; Lipo-BE-NN = liposome BE-NN; strains are from Department of Chemistry and Biochemistry, Laurentian University, Ontario, N. A. Obtained from Omri's laboratory (strain M is mucus-like B. cepacia; PA = P. Aeruginosa; SA = S. Aureus). Tobramycin was obtained from Pharmacy Department of Winthrop-University Hospital, Mineola, NY.

NN = tobramycin; BE = BisEDT, 0.8 μg / ml; Lipo-BE-NN = liposome BE-NN; strains are from Department of Chemistry and Biochemistry, Laurentian University, Ontario, N. A. Obtained from Omri's laboratory (strain M is mucus-like B. cepacia; PA = P. Aeruginosa; SA = S. Aureus). Tobramycin was obtained from Pharmacy Department of Winthrop-University Hospital, Mineola, NY.

BE = BisEDT; NaPYR = sodium pyrithione; chemicals were obtained from Sigma Aldrich. Bold is synergistic. The strains shown were from the American Type Culture Collection (ATCC, Manassas, Va.).

Example 7
Comparison of the effects of bismuth-thiol (BT) and antibiotics against Gram-positive and Gram-negative bacteria, including antibiotic-resistant strains. In this example, the in vitro activity of BisEDT and a comparison agent was compared to Gram-positive and responsible for skin and soft tissue infections. Evaluated against multiple clinical isolates of negative bacteria.

Materials and Methods Test compounds and test concentration ranges are as follows: BisEDT (Domenico et al 1997; Domenico et al., Antimicrob. Agents. Chemother. 45 (5): 1414-1421, and Example 1), 16-0.015 μg / mL; linezolid (ChemPacifica Inc., # 35710), 64-0.06 μg / mL; daptomycin (Cubist Pharmaceuticals # MCB2007), 32-0.03 μg / ml and 16-0.015 μg / mL; Vancomycin (Sigma-Aldrich, St. Louis, MO, # V2002), 64-0.06 μg / mL; ceftazidime (Sigma # C3809), 6 4-0.06 μg / mL and 32-0.03 μg / mL; imipenem (US Pharmacopeia, NJ, # 1337809), 16-0.015 μg / mL and 8-0.008 μg / mL; ciprofloxacin (US Pharmacopoeia, # IOC265), 32-0.03 μg / mL and 4-0.004 μg / mL; Gentamicin (Sigma # G3632) 32-0.03 μg / mL and 16-0.015 μg / mL. All test substances except gentamicin were dissolved in DMSO and gentamicin was dissolved in water. Stock solutions were prepared at 40 times the highest concentration in the test plate. The final concentration of DMSO in the inspection system was 2.5%.

Organisms Examination organisms were obtained from the following clinical laboratories: CHP, Clarian Health Partners, Indianapolis, IN; UCLA, University of California, Ros Angel Medical Center, Los U. Center, Public Health Institute Tuberculosis Center, New York, NY; ATCC, American Type Culture Collection, Manassas, VA; Mt Sinais Hp. , Mount Sinai Hospital, New York, NY; UCSF, University of California San Francisco General Hospital, San Francisco, CA; Bronson Hospital, Bronson Methodist Hospital, Kalamazoo, MI; isolates for quality management, American Type Culture Collection (ATCC , Manassas, VA). The organisms were streaked for isolation on an agar medium appropriate for each organism. Colonies were removed from the isolation plate by swab and placed in suspension in an appropriate broth containing cryoprotectant. The suspension was aliquoted into cold storage vials and kept at -80 ° C. Abbreviations: BisEDT, bismuth-1,2-ethanedithiol; LZD, linezolid; DAP, daptomycin; VA, vancomycin; CAZ, ceftazidime; IPM, imipenem; CIP, ciprofloxacin; GM, gentamicin; MSSA, methicillin sensitive staphylo CLSI QC, quality control strain of Clinical and Laboratory Standards Institute; MRSA, methicillin-resistant Staphylococcus aureus; CA-MRSA, community-acquired methicillin-resistant Staphylococcus aureus; MSSE, methicillin susceptibility Epidermidis; MRSE, methicillin-resistant Staphylococcus epidermidis; VSE, vancomycin sensitivity Enterococcus.

Isolates can be obtained from frozen vials in appropriate media (Trypticase Soy Agar (Becton-Dickinson, Sparks, MD) for most organisms, or Trypticase Soy Agar for Streptococcus + 5% sheep blood (Clevelandfic). Streaked up. Plates were incubated overnight at 35 ° C. Quality control organisms were included. The medium used for the MIC assay was Mueller Hinton II Broth (MHB II-Becton Dickinson, # 212322) for most organisms. MHBII was supplemented with 2% dissolved horse blood (Cleveland Scientific Lot # H13913) and adapted to the development of Streptococcus pyogenes and Streptococcus agalactia. The medium was prepared at 102.5% of normal weight to offset the dilution caused by adding 5 μL of drug solution to each well of the microdilution panel. In addition, testing with daptomycin supplemented the medium with an additional 25 mg / L Ca ²⁺ .

  MIC assay, Clinical and Laboratory Standards Institute (Clinical and Laboratory Standards Institute Methods for Dilution Antimicrobial Susceptibyity Tests for Bacteria That Grow Aerobically;.. Approved Standard-Seventh Edition Clinical and Laboratory Standards Institute document M7-A7 [ISBN 1-56238- 587-9] Clinical and Laboratory Standards Institute Ute, 940 West Valley Road, Suite 1400, Wayne, Pennsylvania 19087-1898 USA, 2006), and serial dilution and liquid transfer were performed using an automatic liquid handler. Automatic liquid handlers included Multidrop 384 (Labsystems, Helsinki, Finland), Biomek 2000 and Multimek 96 (Beckman Coulter, Fullerton CA). The wells in rows 2-12 of a standard 96 well microdilution plate (Falcon 3918) were filled with Multidrop 384 with 150 μL of water for DMSO or gentamicin. Drug (300 μL) was dispensed into the first column of the appropriate row in these plates. These became mother plates when preparing inspection plates (daughter plates). Biomek 2000 completed the series movement up to the eleventh row of the mother plate. The wells in the twelfth row contained no drug and were organism growth control wells on the daughter plate. 185 μL of the appropriate test medium (described above) was added to the daughter plate using a Multidrop 384. Daughter plates were prepared in a Multimek 96 instrument by transferring 5 μL of the drug solution from each well of the mother plate to each corresponding well of each daughter plate in one step.

  A standard inoculum for each organism was prepared by the CLSI method (ISBN 1-56238-587-9, cited above). A suspension was prepared in MHB to equal the turbidity of a 0.5 McFarland standard. The suspension was diluted 1: 9 with broth appropriate for the organism. The inoculum of each organism was dispensed into vertically separated sterile reservoirs (Beckman Coulter) and inoculated into plates using Biomek 2000. The daughter plate was placed on the working surface of the Biomek 2000, which was reversed so that inoculation was performed from a low drug concentration to a high drug concentration. Biomek 2000 delivered 10 μL of the standardized inoculum to each well. This resulted in a final cell concentration of the daughter plate of approximately 5 × 10 5 colony forming units / mL. The daughter plate wells thus finally contained 185 μL broth, 5 μL drug solution, and 10 μL bacterial inoculum. Plates were stacked in triplicate, the top plate was covered, placed in a plastic bag, and most of the isolates were incubated at 37 ° C. for approximately 18 hours. Streptococcus plates were read after 20 hours of incubation. The microplate was viewed from the bottom using a plate viewer. In each of the test media, an uninoculated soluble control plate was observed for evidence of drug precipitation. The MIC was read and recorded as the lowest drug concentration that inhibited the visual development of the organism.

Results All commercially available drugs were soluble at all test concentrations in both media. BisEDT showed a small amount of precipitate at 32 μg / mL but did not affect the MIC reading because the inhibitory concentration of all tested organisms was well below that concentration. On each assay day, the appropriate quality control strain was included in the MIC assay. MIC values obtained from these strains, as appropriate, published for each drug (Clinical and Laboratory Standards Institute-Performance Standards for Bioscience Sustainable Stift. 18-National Institute of Biomedical Sustainability. -653-0] Clinical and Laboratory Standards Institute, 940 West Valley Road, Suite 1400, Wayne, Pennsylvania 19087-1898 USA, 2008).

  On each assay day, the appropriate quality control strain was included in the MIC assay. MIC values obtained from these strains, as appropriate, published for each drug (Clinical and Laboratory Standards Institute-Performance Standards for Bioscience Sustainable Stift. 18-National Institute of Biomedical Sustainability. -653-0]). Of the 141 types of quality control strains for which the quality control range was announced, 140 types (99.3%) were within the specified range. One type of exception is S.M. Imipenem for Aureus strain 29213, one numerical value obtained in one run was one dilution lower than the published QC range (0.008 μg / mL or less). All other quality control results during the implementation were within the specified quality control scope.

  BisEDT is a methicillin sensitive Staphylococcus aureus (MSSA), methicillin resistant S. cerevisiae. Demonstrates potent activity against Aureus (MRSA) and community-acquired MRSA (CA-MRSA), inhibits all strains tested below 1 μg / mL and has a MIC90 value of 0 in all three organism groups 0.5 μg / mL. BisEDT showed greater activity than linezolid and vancomycin, and equivalent activity to daptomycin. Imipenem was more potent than BisEDT against MSSA (MIC90 = 0.03 μg / mL). However, MRSA and CAMRSA were resistant to imipenem, while BisEDT demonstrated activity comparable to that shown for MSSA. BisEDT was highly active against methicillin sensitive and methicillin resistant Staphylococcus epidermidis (MSSE and MRSE) with MIC90 of 0.12 and 025 μg / mL, respectively. BisEDT was more active against MSSE than any of the other drugs tested other than imipenem. BisEDT was the most active drug tested against MRSE.

  BisEDT demonstrated activity comparable to daptomycin, vancomycin, and imipenem against vancomycin-sensitive Enterococcus faecalis (VSEfc) with a MIC90 value of 2 μg / mL. Importantly, BisEDT is the most active drug tested against vancomycin-resistant Enterococcus faecalis (VREfc) with a MIC90 value of 1 μg / mL.

  BisEDT was very active against vancomycin-sensitive Enterococcus faecium (VSEfm) with a MIC90 value of 2 μg / mL, which was similar or similar to daptomycin and was one dilution higher than vancomycin . BisEDT and linezolid were the most active drugs tested against vancomycin-resistant Enterococcus faecium (VREfm), each demonstrating a MIC90 value of 2 μg / mL. The activity of BisEDT against Streptococcus pyogenes (MIC90 value of 0.5 μg / mL) was comparable to vancomycin, greater than linezolid, and slightly lower than daptomycin and ceftazidime. The compound inhibited all strains tested at 0.5 μg / mL or less. In these studies, the bacterial species that is at least sensitive to BisEDT was Streptococcus agalactia, and the observed MIC90 value was 16 μg / mL. BisEDT was less active than all drugs tested except for gentamicin.

  The activity of BisEDT and comparative substances against the gram negative bacteria included demonstrated the efficacy of BisEDT against Acinetobacter baumannii (MIC 90 value 2 μg / mL), making BisEDT the most active compound tested. A significant number of test isolates for comparative agents have increased MICs, and these drugs have MIC90 values outside the measurement range. BisEDT was a potent inhibitor of Escherichia coli and inhibited all strains at 2 μg / mL or less (MIC90 = 2 μg / mL). The compound was less active than imipenem, but more active than ceftazidime, ciprofloxacin and gentamicin. BisEDT also demonstrated activity against Klebsiella pneumoniae, with a MIC90 value of 8 μg / mL, equivalent to imipenem. The relatively high MIC90 values exhibited by imipenem, ceftazidime, ciprofloxacin, and gentamicin indicated that this is a group of organisms with high antibiotic resistance. BisEDT is the most active compound tested against P. aeruginosa with a MIC90 value of 4 μg / mL. There was a high level of resistance to the comparator in this group of test isolates.

  In summary, BisEDT displays a broad spectrum capability for multiple clinical isolates representing multiple species, including those commonly involved in acute and chronic skin and skin structure infections in humans. The activities of BisEDT and important comparators were evaluated against 723 clinical isolates of gram positive and gram negative bacteria. BT compounds demonstrated broad spectrum activity, and for a number of test organisms in this study, BisEDT was the most active compound tested for antimicrobial activity. BisEDT is available from MSSA, MRSA, CA-MRSA, MSSE, MRSE and S.M. It was most active against Piogenes, and the MIC90 value was 0.5 μg / mL or less. Strong activity is observed in VSEfc, VREfc, VSEfm, VREfm, A. Baumani, E. Coli, and P. a. As demonstrated in Elginosa, MIC90 values were in the range of 1-4 μg / mL. The MIC90 value is Pneumoniae (MIC90 = 8 μg / mL) and S. pneumoniae. Observed with agalactia (MIC90 = 16 μg / mL).

Example 8
Enhancement and synergistic activity of particulate BT-antibiotics This example shows that particulate bismuth-thiol (BT) promotes antibiotic activity through potentiating and / or synergistic interactions.

  A major complication in treating infection is the emergence of bacterial antibiotic resistance. S. Epidermidis (MRSE) and S. Methicillin resistance in Aureus (MRSA) actually represents multidrug resistance, which makes it very difficult to eradicate these pathogens. However, hundreds of strains of Staphylococcus tested did not show resistance to BT. Furthermore, sub-inhibition (subMIC) concentrations of BT reduced resistance to several important antibiotics.

  Providing a graphical representation of the antibiotic-resensitizing effects of bismuth ethanedithiol (BisEDT) of subMIC against Staphylococcus aureus MRSA (FIG. 4), gentamicin, cefazolin, cefepime, imipenem, sulfa It shows potentiating antibiotic action with several classes of antibiotics including methoxazole and levofloxacin. That is, BisEDT nonspecifically enhanced the activity of most antibiotics.

１２のＭＲＳＡ臨床単離物をＢＨＩＧ／Ｘ中で発育させて、０〜０．１μｇ／ｍＬ ＢｉｓＥＤＴの存在下で抗生物質の系列希釈物に暴露した。μｇ／ｍＬで計算されたＭＩＣおよびＢＰＣは、少なくとも３回のトライアルから得た平均±標準偏差である。右側の列は、抗生物質感受性（Ｓ）および耐性（Ｒ）に関する標準ＭＩＣを列挙している。 Antimicrobial susceptibility testing by broth dilution was performed on 12 MRSA strains using multiple antibiotics in combination with subMIC levels of BisEDT. Both biofilm preventive concentration (BPC) and minimum inhibitory concentration (MIC) were measured with a special biofilm medium (BHIG / X). The MIC and BPC of gentamicin and cefazolin were reduced by subMIC BisEDT (BisEDT MIC, 0.2-0.4 μg / mL), but not below the sensitive breakpoint. The subMIC BisEDT enhanced the sensitivity of MRSA to gatifloxacin and cefepime to near the breakpoint of sensitivity. These strains were already sensitive to vancomycin, but became more pronounced in the presence of subMIC BisEDT. In general, MIC and BPC were reduced 2-5 fold by subMIC BisEDT.

Twelve MRSA clinical isolates were grown in BHIG / X and exposed to serial dilutions of antibiotics in the presence of 0-0.1 μg / mL BisEDT. The MIC and BPC calculated in μg / mL are the mean ± standard deviation obtained from at least 3 trials. The right column lists standard MICs for antibiotic susceptibility (S) and resistance (R).

１２種のセフェピム耐性ＭＲＳＡを、ｓｕｂＭＩＣのＢｉｓＥＤＴと組み合わせたセフェピムへの感受性について、ポリスチレンプレート内のＢＨＩＧ／Ｘ培地で３７℃で４８時間検査した。 The broth dilution test of cefepime resistant MRSA isolates is shown in Table 19. 0.1 μg / mL BisEDT significantly enhanced the inhibitory activity of cefepime in 11 out of 12 isolates. In this particular study, the data showed synergy between BisEDT and cefepime (FIC <0.5), with many of the isolates at sensitive breakpoints.

Twelve cefepime resistant MRSA were tested for 48 hours at 37 ° C. in BHIG / X medium in polystyrene plates for sensitivity to cefepime in combination with subMIC BisEDT.

ＮＡＦまたはＧＭはμｇ／ｍＬを表わし；ＢＥは０．２μｇ／ｍＬ The results of the combination test with nafcillin or gentamicin are shown in Table 20. BisEDT (0.2 μg / mL) in combination with nafcillin reduced nafcillin MIC90 more than 4-fold relative to MRSA (FIC, 0.74). BisEDT in combination with gentamicin reduced gentamicin's MIC90 more than 10-fold over MRSA (FIC, 0.6). BT suppressed the resistance of all four gentamicin resistant isolates tested to clinically relevant concentrations [Domenico et al. , 2002]. The MIC of these antimicrobial agents was substantially reduced, especially with gentamicin. The broth used in these studies was Trypticase Soy Broth (TSB) containing 2% glucose, showing similar results to those observed in Mueller-Hinton II broth supplemented with 1% sheep blood.

NAF or GM represents μg / mL; BE is 0.2 μg / mL

  Staphylococcus epidermidis The activity of most classes of antibiotics was enhanced in the presence of BisEDT. For BPC, clindamycin and gatifloxacin are S. cerevisiae when combined with BisEDT. It showed significantly higher anti-biofilm activity against epidermidis (FIG. 5). Stated differently, clindamycin, gatifloxacin and gentamicin BPC were reduced by 50-fold, 10-fold and 4-fold in the presence of subMIC BisEDT, respectively.

  A slight decrease in biofilm preventive concentration (BPC) was observed with minocycline, vancomycin, and cefazolin, but rifampicin and nafcillin still showed no effect at 0.05 μg / mL BisEDT. With 0.1 μg / mL BisEDT, no biofilm was detected regardless of the antibiotic used, indicating that no antagonism occurred. This BisEDT concentration is determined by S. It approximated the epidermidis MIC [Domenico et al. , 2003] (see FIG. 5).

  Of the 8 antibiotics tested for growth inhibition, 7 It was significantly enhanced against epidermidis in the presence of 0.1 μg / mL (0.5 μM) BisEDT (FIG. 6). The MIC variation was most pronounced with clindamycin and gentamicin, followed by vancomycin, cefazolin, minocycline, gatifloxacin and nafcillin, and rifampicin had no effect. These strains were resistant to (NC, CZ, GM, CM) antibiotics, and only cefazolin resistance was suppressed to clinically relevant levels by BisEDT.

  S. The minimum bactericidal concentration (MBC) of most antibiotics tested against epidermidis was slightly reduced by subMIC BisEDT. Gentamicin shows the greatest reduction in MBC (4- to 16-fold), followed by cefazolin (4- to 5-fold), vancomycin and nafcillin (3- to 4-fold), minocycline and gatifloxacin (2- to 3-fold) ) And clindamycin and rifampicin MBC were still not significantly affected. It is explained that clindamycin is a bacteriostatic agent and has no bactericidal activity. Cefazolin resistance was suppressed for MBC [Domenico et al. , 2003]. These effects were additive.

  Antimicrobial efficacy has also been demonstrated in an in vivo rat graft infection model (Table 21). With a low level of BisEDT of 0.1 μg / mL, 7 days resistance The prevention of epidermidis biofilm could be promoted.

^ａ 各群は動物１５匹であった。ＭＳ、メチシリン感受性Ｓ．エピデルミディス；ＭＲ、メチシリン耐性Ｓ．エピデルミディス
^ｂ ダクロン移植片に０．１ｍｇ／ｌのＢＴ、１０ｍｇ／ｌのＲＩＰ、１０ｍｇ／ｌのリファンピンを含浸させた。
^ｃ 対照群のＭＳおよびＭＲと比較した場合に統計学的に有意
^ｄ ＭＳ３群と比較した場合に統計学的に有意
^ｅ ＭＲ１群、ＭＲ２群およびＭＲ３群と比較した場合に統計学的に有意 As summarized in Table 21, implants impregnated with 0.1 μg / mL BisEDT, 10 μg / mL RIP and 10 μg / mL rifampin alone or in combination were implanted subcutaneously into rats. A physiological solution (1 ml) containing 2 × 10 ⁷ cfu / ml of MS and MR strain was inoculated onto the surface of the graft using a tuberculin syringe. All grafts were removed 7 days after transplantation and sonicated in sterile saline for 5 minutes to remove attached bacteria. Quantification of viable bacteria was obtained by culturing the dilutions on blood agar plates. The detection limit was approximately 10 cfu / cm ² .

^a Each group had 15 animals. MS, methicillin sensitive Epidermidis; MR, methicillin resistant Epidermidis
^b Dacron grafts were impregnated with 0.1 mg / l BT, 10 mg / l RIP, 10 mg / l rifampin.
^c Statistically significant when compared to MS and MR in control group
^d Statistically significant when compared to MS3 group
^e Statistically significant when compared to MR1, MR2 and MR3 groups

Ｐ．エルギノーサの耐性株を、トブラマイシン（ＮＮ）およびＢｉｓＥＤＴ（ＢＥ；０．３３μｇ／ｍｌ）の存在下、３７℃のＭｕｅｌｌｅｒ−Ｈｉｎｔｏｎ ＩＩブロス中で培養した。ＭＩＣを、２４±１時間の発育を阻害する抗生物質濃度として測定した。 The activity of tobramycin against gram-negative bacteria-resistant Pseudomonas aeruginosa was enhanced several fold by subMIC BisEDT (Table 22). In these trials, the MIC was more accurately defined as IC ₂₄ .

P. A resistant strain of aeruginosa was cultured in Mueller-Hinton II broth at 37 ° C. in the presence of tobramycin (NN) and BisEDT (BE; 0.33 μg / ml). MIC was measured as the concentration of antibiotic that inhibits growth for 24 ± 1 hour.

^ａ ０．４μｇ／ｍｌのＢｉｓＥＤＴにより阻害された３種の菌株は、更なる試験から除外した。
０．５以下のＦＩＣ指数は、相乗性を示し、０．５を超え１．０未満のＦＩＣは、増強性を示す。 For tobramycin-resistant Burkholderia cepacia, 0.4 μg / mL BisEDT made 7 of 10 isolates tobramycin sensitive (mean FIC; 0.48) and reduced MIC ₉₀ 10-fold. (Table 23). Both tobramycin MIC and MBC were significantly reduced by subMIC BisEDT to a level that was feasible for 50 clinical Burkholderia cepacia isolates [Veloria et al. , 2003]. Liposomal forms of BisEDT and tobramycin are described in P. Has a high synergy demonstrated against Elginosa (Halwani et al., 2008; Halwani et al., 2009).

^a Three strains inhibited by 0.4 μg / ml BisEDT were excluded from further testing.
An FIC index of 0.5 or less indicates synergy, and an FIC greater than 0.5 and less than 1.0 indicates enhancement.

Ｅ．コリの耐性株を、単独または組み合わせでのクロラムフェニコール（ＣＭ）またはアンピシリン（ＡＭＰ）およびＢｉｓＥＤＴの存在下、３７℃のＭｕｅｌｌｅｒ−Ｈｉｎｔｏｎ ＩＩブロス中で培養した（ＢＥ：０．３３μｇ／ｍｌ）。ＭＩＣを、２４±１時間の発育を阻害した抗生物質濃度として測定した。 Chloramphenicol and ampicillin-resistant Escherichia coli became sensitive to these drugs with the addition of subMIC BisEDT (Table 24).

E. Resistant strains of E. coli were cultured in Mueller-Hinton II broth at 37 ° C. in the presence of chloramphenicol (CM) or ampicillin (AMP) and BisEDT, alone or in combination (BE: 0.33 μg / ml) . MIC was measured as the concentration of antibiotic that inhibited growth for 24 ± 1 hour.

Ｅ．コリの耐性株を、単独または組み合わせでのドキシサイクリン（ＤＯＸ）およびＢｉｓＥＤＴの存在下、３７℃のＭｕｅｌｌｅｒ−Ｈｉｎｔｏｎ ＩＩブロス中で培養した（ＢＥ：０．３３μｇ／ｍｌ）。ＭＩＣを、２４±１時間の発育を阻害した抗生物質濃度として測定した。 Tetracycline resistant Escherichia coli became sensitive to tetracycline by the addition of subMIC BisEDT (Table 25). The combination was synergistic for TET M and TET D strains (FIC ≦ 0.5) and had an additive effect on TET A and TET B strains.

E. Resistant strains of E. coli were cultured in Mueller-Hinton II broth at 37 ° C. (BE: 0.33 μg / ml) in the presence of doxycycline (DOX) and BisEDT alone or in combination. MIC was measured as the concentration of antibiotic that inhibited growth for 24 ± 1 hour.

References Domenico P, R O'Leary, BA Cunha. 1992. Differential effect of bismuth and salicylate compounds on antibiotic sensitivity of Pseudomanas aeruginosa. Eur J Clin Microbiol Infect Dis 11: 170-175; Domenico P, D Parikh, BA Cunha. 1994. Bismuth modulation of antibacterial activity, against gastrointestinal bacterial pathogens. Med Microbiol Lett 3: 114-119; Domenico P, Kazzaz JA, Davis JM, Niederman MS. 2002. Subinhibit bimusmu etherethiol (BisEDT) sensitives resist Staphylococcus aureus to nafcillin or gentamicin. Annual Meeting, ASM, Salt Lake City, UT; Domenico P, Kazzaz JA, Davis JM. 2003. Combining antibiotic resistance with bismuth-thiol. Research Advances in Antimicrob Agents Chemother 3: 79-85; 2004. BisEDT and RIP act in synergy to prevent graft effects by resist staphylococci. Peptides 25: 2047-2053; Halwani M, Bloome S, Suntres ZE, Alipore M, Azghani AO, Kumar A, Omri A. 2008. Liposomal bismuth-ethanediolformation enhancements antimicrobial activity of tobramcin. Intl J Pharmaceut 358: 278-84; Halwani M, Hebert S, Suntres ZE, Lafrénie RM, Azghani AO, Omri A. et al. 2009. Bismuth-thiol enhancement enhancements bioactives of liposomal tobramcincin aginest biofilm and quorum sensation moleculare. Int J Pharmaceut 373: 141-6; Veloira WG, Gurzenda EM, Domenico P, Davis JM, Kazzaz JA. 2003. Synergy of tobramycin and bismuth thiols against Burkholderia cepacia. J Antimicrob Chemother 52: 915-919.

ＳＡ、スタフィロコッカス・アウレウス；ＭＲＳＡ、メチシリン耐性スタフィロコッカス・アウレウス；ＥＦｃ、エンテロコッカス・フェカーリス；ＳＰ、ストレプトコッカス・ニューモニエ；ＰＲＳＰ、ペニシリン耐性ストレプトコッカス・ニューモニエ；ＥＣ、エシェリヒア・コリ；ＫＰ、クレブシエラ・ニューモニエ；ＰＡ、シュードモナス・エルギノーサ；Ｂｃｅｐ、バークホルデリア・セパシア；Ｂｍｕｌｔ、バークホルデリア・マルチボランス；Ａｂａｕ、アシネトバクター・バウマニー；Ｍｓｍｅｇ、マイコバクテリウム・スメグマチス Example 9
Particulate BT-Antibiotic Enhancement and Synergistic Activity This example shows that BisEDT, a particulate bismuth thiol, has antibiotic activity through enhanced and / or synergistic interactions with specific antibiotics against specific microbial target organisms. To promote. One-point data for each combination shown in Table 26 was created essentially according to the method used in Example 8.

SA, Staphylococcus aureus; MRSA, methicillin-resistant Staphylococcus aureus; EFc, Enterococcus faecalis; SP, Streptococcus pneumoniae; PRSP, Penicillin-resistant Streptococcus pneumoniae; EC, Escherichia coli; KP, Klebsiella; PA, Pseudomonas aeruginosa; Bcep, Burkholderia cepacia; Bmult, Burkholderia multiborance; Abau, Acinetobacter baumannii; Msmeg, Mycobacterium smegmatis

Example 10
Enhanced and synergistic activity of microparticulate BT-antibiotics To test the effect of a combination of microparticulate BisEDT and four BisEDT analogs prepared as described above with other agents on representative strains of multiple Gram-negative pathogens did. Utilizing a variation of the general test method, the partial inhibitory concentration (FIC) and the FIC index (FICI) used were synergistic (FICI ≦ 0.5), enhancing (0.5 <FICI ≦ 1.0) ), Antagonistic (FICI> 4.0) and insignificant (1.0 <FICI ≦ 4.0) were determined (Antibiotics in Laboratory Medicine, edited by V Lorian. Williams and Wikins, Baltimore, MD). Edition, pp. 432-492, Elipoulos G and R Moellering., 1991. Antimicrobial combinations .; Odds, 2003 J. Antimicrob.Chemother.52 (1): 1). The checkerboard technique was used to determine the FIC index and used for this test.

  Stock solutions of all test substances were prepared 40 times the final target concentration with an appropriate solvent. All test substances were in solution under these conditions. Unless the strain is totally resistant to the test agent, the final drug concentration in the FIC assay plate was set so that the MIC values of each drug for each test organism were collected. The tested density range is shown in Table 27. Test organisms were initially obtained from clinical sources or from the American Type Culture Collection. Upon acquisition, the isolate was streaked on Tryptic Soy Agar II (TSA). Colonies were collected from these plates and cell suspensions were prepared in an appropriate broth growth medium containing a cryoprotectant. Thereafter, aliquots were frozen at -80 ° C. Frozen seeds of organisms to be tested in a given assay were thawed and streaked onto TSA plates for incubation and incubated at 35 ° C. All organisms were examined with Mueller-Hinton II Broth (Beckton Dickinson, Lot No. 9044411). Broth was prepared at 1.05 times the normal weight / volume to offset the 5% volume of drug in the final test plate.

  The final inhibitory concentration (MIC) value was measured as before using the broth microdilution method for aerobic bacteria (Clinical and Laboratory Standards Institute (CLSI). Applied Standard-Eight Edition. CLSI document M07-A8 [ISBN 1-56238-689-1] .Clinical and Laboratory Standards Institute, 940 West Valley Valley, 1940. e, Pennsylvania 19087-1898 USA, 2009).

  FIC values were measured using the broth microdilution method described previously (Sweeney et al., 2003 Antimicrob. Agents Chemother. 47 (6): 1902-1906). To prepare the test plate, serial dilution and liquid transfer were performed using an automated liquid handler (Multidrop 384, Labsystems, Helsinki, Finland; Biomek 2000 and Multimek 96, Beckman Coulter, Fullerton CA).

  Using Multidrop 384, 150 μL of the appropriate solvent was loaded into the second through twelfth rows in the appropriate wells of a standard 96 well microdilution plate (Falcon 3918). 300 μL of each second test agent was added to each well in the first row of the plate. These plates were used to prepare drug “mother plates” that provide serial dilutions for drug combination plates. Using Biomek 2000, 150 μL of each second drug solution (40 ×) was transferred from the first row of wells of the mother plate to generate 11 2 × dilution series. Bis-EDT (and analog) mother plates were serially diluted from top to bottom using a multichannel pipette. Combine two mother plates, one for each second drug and one for Bis-EDT (or analog), and transfer (using a multichannel pipette) the same volume to the drug combination plate As a result, a “checkerboard” pattern was formed. Row H and column 12 each contained serial dilutions of one of the drugs for which the MIC was measured.

180 μL of test medium was added to the “daughter plate” using Multidrop 384. Thereafter, using Multimek 96, 10 μL of the drug solution was transferred from each well of the drug combination mother plate to each corresponding well of the daughter plate in one step. Finally, the test organism was inoculated on the daughter plate. A standardized inoculum for each organism was prepared according to published guidelines (CLSI, 2009). For all isolates, the inoculum of each organism was dispensed into a vertically-sterilized sterile reservoir (Beckman Coulter) and inoculated into the plate using a Biomek 2000. The instrument delivered 10 μL of the standardized inoculum to each well to give a final cell concentration in the daughter plate of approximately 5 × 10 ⁵ colony forming units / mL.

  The test format produced an 8 × 12 checkerboard that tested each compound alone (columns 12 and 8) and combined with drug concentrations in various ratios. All organism plates were stacked in triplicate, the top plate was covered, placed in a plastic bag and incubated at 35 ° C. for approximately 20 hours. After incubation, the microplate was removed from the incubator and viewed from the bottom using a ScienceWare plate viewer. The prepared reading sheet was filled with the MIC of Drug 1 (Row H), the MIC of Drug 2 (Column 12) and the well with and without growth.

Using the Excel program, FIC was measured by the formula: (MIC of Compound 1 in combination / MIC of Compound 1 alone) + (MIC of Compound 2 in combination / MIC of Compound 2 alone). The FICI for the checkerboard was calculated from each FIC by the formula: (FIC ₁ + FIC ₂ +... FIC _n ) / n, where n = number of each well per plate on which the FIC is calculated. If a single drug produced an MIC result outside the measurement range, the highest next concentration was used as the MIC value in the FIC calculation.

^１ＭＩＣ、最小阻止濃度
^２ＦＩＣＩ、部分阻止濃度指数

^１ＭＩＣ、最小阻止濃度
^２ＦＩＣＩ、部分阻止濃度指数

^１ＭＩＣ、最小阻止濃度
^２ＦＩＣＩ、部分阻止濃度指数

^１ＭＩＣ、最小阻止濃度
^２ＦＩＣＩ、部分阻止濃度指数

^１ＭＩＣ、最小阻止濃度
^２ＦＩＣＩ、部分阻止濃度指数

^１ＭＩＣ、最小阻止濃度
^２ＦＩＣＩ、部分阻止濃度指数

^１ＭＩＣ、最小阻止濃度
^２ＦＩＣＩ、部分阻止濃度指数

^１ＭＩＣ、最小阻止濃度
^２ＦＩＣＩ、部分阻止濃度指数

^１ＭＩＣ、最小阻止濃度
^２ＦＩＣＩ、部分阻止濃度指数

^１ＭＩＣ、最小阻止濃度
^２ＦＩＣＩ、部分阻止濃度指数

^１ＭＩＣ、最小阻止濃度
^２ＦＩＣＩ、部分阻止濃度指数

^１ＭＩＣ、最小阻止濃度
^２ＦＩＣＩ、部分阻止濃度指数

^１ＭＩＣ、最小阻止濃度
^２ＦＩＣＩ、部分阻止濃度指数

^１ＭＩＣ、最小阻止濃度
^２ＦＩＣＩ、部分阻止濃度指数

^１ＭＩＣ、最小阻止濃度
^２ＦＩＣＩ、部分阻止濃度指数

^１ＭＩＣ、最小阻止濃度
^２ＦＩＣＩ、部分阻止濃度指数

^１ＭＩＣ、最小阻止濃度
^２ＦＩＣＩ、部分阻止濃度指数

^１ＭＩＣ、最小阻止濃度
^２ＦＩＣＩ、部分阻止濃度指数

^１ＭＩＣ、最小阻止濃度
^２ＦＩＣＩ、部分阻止濃度指数

^１ＭＩＣ、最小阻止濃度
^２ＦＩＣＩ、部分阻止濃度指数 All of the microparticle Bis-EDT, the four microparticle BT analogs, and other drugs (and drug combinations) were soluble at all final test concentrations. The measured MIC and FICI values are shown in the table below.

¹ MIC, minimum inhibitory concentration
² FICI, partial inhibitory concentration index

¹ MIC, minimum inhibitory concentration
² FICI, partial inhibitory concentration index

¹ MIC, minimum inhibitory concentration
² FICI, partial inhibitory concentration index

¹ MIC, minimum inhibitory concentration
² FICI, partial inhibitory concentration index

¹ MIC, minimum inhibitory concentration
² FICI, partial inhibitory concentration index

¹ MIC, minimum inhibitory concentration
² FICI, partial inhibitory concentration index

¹ MIC, minimum inhibitory concentration
² FICI, partial inhibitory concentration index

¹ MIC, minimum inhibitory concentration
² FICI, partial inhibitory concentration index

¹ MIC, minimum inhibitory concentration
² FICI, partial inhibitory concentration index

¹ MIC, minimum inhibitory concentration
² FICI, partial inhibitory concentration index

¹ MIC, minimum inhibitory concentration
² FICI, partial inhibitory concentration index

¹ MIC, minimum inhibitory concentration
² FICI, partial inhibitory concentration index

¹ MIC, minimum inhibitory concentration
² FICI, partial inhibitory concentration index

¹ MIC, minimum inhibitory concentration
² FICI, partial inhibitory concentration index

¹ MIC, minimum inhibitory concentration
² FICI, partial inhibitory concentration index

¹ MIC, minimum inhibitory concentration
² FICI, partial inhibitory concentration index

¹ MIC, minimum inhibitory concentration
² FICI, partial inhibitory concentration index

¹ MIC, minimum inhibitory concentration
² FICI, partial inhibitory concentration index

¹ MIC, minimum inhibitory concentration
² FICI, partial inhibitory concentration index

¹ MIC, minimum inhibitory concentration
² FICI, partial inhibitory concentration index

Example 11
The effect of bismuth-thiol on infection in critical femurs of the rat rat Current care standard methods for open fractures are irrigation, necrotic tissue removal and antibiotics, which do not cause infection of bacterial load in wound It is intended to reduce to a certain extent. Despite these treatments, infection complicates up to 75% of severe open tibial fractures due to war. Interestingly, even though early infections are often caused by Gram-negative bacteria, late infections that are involved in healing problems and cleavage are Gram-positive infections, mostly due to Staphylococcus species (Johnson 2007).

  S. One reason Aureus is resistant to standard treatment is its ability to form biofilms. Bacteria in the biofilm can withstand concentrations of antimicrobial compounds that kill similar organisms in the medium (Costerton 1987).

  The purpose of this study was to determine whether BT alone or with antibiotics would reduce infection in a contaminated open fracture model. The contaminated rat femoral critical defect model is a well-accepted model and was used in the experiment described in this example. This model is a standardized model for comparing various potential treatments and their effects in terms of reducing infection and / or improving healing.

  Although the compounds (CPD) CPD-8-2 (bismuth-pyrithione / butanedithiol; Table 1) and CPD-11 (bismuth pyrithione / ethanedithiol; Table 1) have a different spectrum of activity than Bis-EDT. Rather, it is two BIS-Bis analogs that have shown the ability to biofilm-secreting bacteria in vitro.

Three BT formulations: Bis-EDT, CPD-11 and CPD-8-2 (see Table 1) with or without tobramycin and vancomycin in polymethylmethacrylate (PMMA) cement bead vehicle. When used, in vitro S. cerevisiae. Inhibitory effect on Aureus strain. Three formulations of particulate BT were produced in the clinically useful hydrogel form described herein. These BT were tested in suspension in the gel at a concentration of 5 mg / ml ⁻¹ found to be the appropriate concentration for gel delivery. The gel formulation conformed to the wound profile and did not need to be removed after application.

  Two treatment objectives were utilized, with BT used alone for the first objective and BT with systemic antibiotics (ABx) for the second objective.

(A) BT alone At 6 hours after inoculation with Aureus, the necrotic tissue of the wound was removed, irrigated with physiological saline, and 1 ml of BT gel was inserted into the defect.

(B) BT and systemic antibiotics (ABx)
S. At 6 hours after inoculation with Aureus, the debridement of the wound was removed, irrigated with physiological saline, and 1 ml of the added BT gel was inserted into the defect. The antibiotic used was a dose of cefazolin equal to 5 mg kg ⁻¹ delivered by subcutaneous injection twice daily for a total of 3 days after injury. The first dose was administered immediately before necrotic tissue removal. Past data suggested that this dose could reduce bacterial levels from about 10 ⁶ to about 10 ⁴ , and therefore measure the relative effects of different BT.

(C) Control Six hours after inoculation with Aureus, the wound necrotic tissue was removed and ligated with saline. Control animals were also treated with cefazolin with the regimen described previously.

procedure:
In vivo rat injury model procedures were performed as described by Chen et al. (2002 J. Orthop. Res. 20: 142; 2005 J. Orthop. Res. 23: 816; 2006 J. Bone Joint Surg. Am. 88. : 1510; 2007 J. Orthop. Trauma 21: 693). Rats were anesthetized and prepared for surgery. The anterolateral surface of the femoral shaft was exposed through a 3 cm incision. The periosteum and attached muscle were detached from the bone. A polyacetyl plate (27 × 4 × 4 mm) was placed on the anterior-lateral surface of the femur. The plate was pre-drilled and passed through a twisted Kirschner wire of 0.9 mm diameter. These plate substrates were shaped to fit the contours of the femoral shaft. A pilot hole was cut through both femoral cortical bones using a plate as a mold and a twisted Kirschner wire was inserted through the plate and femur. Indentations present at 6 mm intervals on the plate served as a guide for bone removal. In an attempt to prevent thermal damage, the defect was created using a small oscillating saw while cooling the tissue by continuous irrigation.

In each of a plurality of groups of 10 animals, Aureus 1 × 10 ⁵ CFU was inoculated and treated with BT alone or in combination with antibiotics 6 hours after inoculation as described above. The groups were as follows: Bis-EDT gel; MB-8-2 gel; Bis-EDT gel; Bis-EDT gel &Abx; MB-11 gel &Abx; MB-8-2 gel &Abx; ).

  The animals are euthanized 14 days after surgery and the bones and devices are sent for microbial analysis and the results are shown in FIG.

  Based on power analysis, 10 animals per group will give 80% power to detect a 25% difference between the treatment and control groups. In this case, the predicted standard deviation is 35% and α is 0.05.

  As shown in FIG. 7, when combined with Bis-EDT, MB-11 and MB-8-2, cefazolin's antibiotic activity was enhanced compared to cefazolin or any Bis compound alone, and S . Reduced Aureus infection. Cefazolin in combination with MB-11 and MB-8-2 showed high antibiotic activity compared to single cefazolin and was detected on the device. It has been shown to reduce Aureus infection. Bis-EDT was not thought to affect cefazolin activity in this capacity.

Reference material:
Costerton JW, Cheng KJ, Geesey GG, et al. Baceral Biofilms in Nature and Disease. Ann Rev Microbiol. 1987; 41: 435-64
Domenico P, Baldassari L, Schoch PE, Kaehler K, Sasatsu M, Cunha BA. Activities of Bismuth Thiols Against Staphylococci and Staphylococcal Biofilms. Antimicrob Agents and Chemother. 2001; 45 (5): 1417-21.
Halwani M, Bloome S, Suntres ZE, et al. Liposomal bismuth-ethanediol formation enhancement enhances activity of tobramcin. Int J Pharm. 2008; 358: 278-84.
Johnson EN, Burns TC, Hayda RA, Hospital DR, Murray CK. Infection complications of open type III timber fractures amon comb casualties. Clin Infect Dis. 2007; 45 (4): 409-415.

Other cited documents and related documents Domenico et al. , Canadian J. Microbiol. 31: 472-78 (1985); Domenico et al. , Reduction of capsular polysaccharides and potentiation of aminocosid inhibition in gram-negative bacteria with bismuth subsiculate. J Antimicrob Chemo 1991; 28: 801-810; Domenico et al. Infection 20: 66-72 (1992); Domenico et al. , Infect. Immun. 62: 4495-99 (1994); Domenico et al. , J. et al. Antimicrol. Chemother. 38: 1031-40 (1996); Domenico et al. , Enhancement of bismuth antibacterial activity with lipophilic thiol chelators. Antimicrob Agents Chemother 1997; 41: 1697-703; Domenico et al. , Surface antigen exposure by bismuth-dimercap suppression of Klebsiella pneumoniae capsular polysaccharide. Infect Immun 67: 664-669 (1999); Domenico et al. 2000. The potential of bismuth-thiols for treatment and prevention of infection. Infect Med 17: 123-127; Domenico et al. , Activities of bismuth thiols against staphylococci and staphylococcal biofilms. Antimicrob Agents Chemother 2001; 45: 1417-21; Domenico et al. , Combing antibiotic resistance with bismuth-thiols. Research Advances in Antimicrob Agents Chemother 2003; 3: 79-85; Domenico et al. , Reduction of capsular polysaccharides and potentiation of aminocosid inhibition in gram-negative bacteria with bismuth subsiculate. J Antimicrob Chemo 1991; 28: 801-810; Domenico et al. , BisEDT and RIP act in synergy to prevent graft effects by resist staphylococci. Peptides 2004. 25: 2047-53; Domenico et al. , 2005. Pyrithion enhanced antimicrobial activity of bismuth. Antibiotics for Clinicians 9: 291-297; US Pat. No. 6,582,719; US Pat. No. RE37,793, US Pat. Nos. 6,248,371, 6,086,921, 6,380,248 No. 6,582,719, No. 6,380,248, No. 6,875,453.

  The various embodiments described above can be combined to provide further embodiments. All of the U.S. patents, U.S. patent application publications, U.S. patent applications, foreign patents, foreign patent applications and non-patent publications referenced herein and / or listed in the application data sheet are hereby incorporated by reference in their entirety. Incorporated into the book. The aspects of the embodiments can be modified as needed to provide further embodiments using various patent, application and publication concepts.

These and other changes can be made to the embodiments in light of the above detailed description. In general, in the following claims, the terms used should not be construed as limiting the claims to the specific embodiments disclosed in the specification and the claims, and as such The claims should be construed to include all possible embodiments, along with the full scope of equivalents to which the claims are entitled.

Claims (47)

  A bismuth-thiol composition comprising a plurality of microparticles comprising a bismuth-thiol (BT) compound, wherein substantially all of the microparticles have a volume average diameter of about 0.4 μm to about 5 μm, wherein the BT compound is A bismuth-thiol composition comprising a bismuth or bismuth salt and a thiol-containing compound. A bismuth-thiol composition comprising a plurality of microparticles comprising a bismuth-thiol (BT) compound, wherein substantially all of the microparticles have a volume average diameter of about 0.4 μm to about 5 μm;
(A) under conditions and time sufficient to obtain a solution substantially free of solid precipitate, (i) comprising a bismuth salt containing bismuth at a concentration of at least 50 mM, and hydrophilic, polar or organic solubilization (Ii) mixing a sufficient amount of ethanol to obtain an admixture containing about 25% by volume of ethanol; and (b) forming a precipitate containing microparticles containing a BT compound. An ethanolic solution containing a thiol-containing compound present in the reaction solution in a molar ratio of about 1: 3 to about 3: 1 to bismuth under conditions and time sufficient to To obtain a reaction solution,
A bismuth-thiol composition formed by a method comprising: The bismuth-thiol composition according to claim 2, wherein the bismuth salt is Bi (NO ₃ ) ₃ .   The bismuth-thiol composition of claim 2, wherein the acidic aqueous solution comprises at least 5 wt%, 10 wt%, 15 wt%, 20 wt%, 22 wt% or 22.5 wt% bismuth.   Acidic aqueous solution at least 0.5 wt%, 1 wt%, 1.5 wt%, 2 wt%, 2.5 wt%, 3 wt%. The bismuth-thiol composition of claim 2 comprising 3.5 wt%, 4 wt%, 4.5 wt% or 5 wt% nitric acid. Thiol-containing compound is 1,2-ethanedithiol, 2,3-dimercaptopropanol, pyrithione, dithioerythritol, 3,4-dimercaptotoluene, 2,3-butanedithiol, 1,3-propanedithiol, 2-hydroxy Propanethiol, 1-mercapto-2-propanol, α-lipoic acid, dithiothreitol, methanethiol (CH ₃ SH [m-mercaptan]), ethanethiol (C ₂ H ₅ SH [e-mercaptan]), 1- propanethiol _{(C 3 H 7 SH [n} -P mercaptans]), 2- propanethiol _{(CH 3 CH (SH) CH} 3 [2C 3 mercaptan]), butanethiol _{(C 4} H 9 SH [n- butyl mercaptan] ), Tert-butyl mercaptan (C (CH ₃ ) ₃ SH [t-butyl mecaptan Lucaptan]), pentanethiol (C ₅ H ₁₁ SH [pentyl mercaptan]), coenzyme A, lipoamide, glutathione, cysteine, cystine, 2-mercaptoethanol, 2-mercaptoindole, transglutaminase, (11-mercaptoundecyl) hexa (Ethylene glycol), (11-mercaptoundecyl) tetra (ethylene glycol), (11-mercaptoundecyl) tetra (ethylene glycol) functionalized gold nanoparticles, 1,1 ′, 4 ′, 1 ″ -ter Phenyl-4-thiol, 1,11-undecanedithiol, 1,16-hexadecanedithiol, technical grade 1,2-ethanedithiol, 1,3-propanedithiol, 1,4-benzenedimethanethiol, 1, 4-butanedithiol, 1,4- Tandithiol diacetate, 1,5-pentanedithiol, 1,6-hexanedithiol, 1,8-octanedithiol, 1,9-nonanedithiol, adamantanethiol, 1-butanethiol, 1-decanethiol, 1-dodecane Thiol, 1-heptanethiol, 1-heptanethiolpurum, 1-hexadecanethiol, 1-hexanethiol, 1-mercapto (triethylene glycol), 1-mercapto (triethylene glycol) methyl ether functionalized gold nano Particles, 1-mercapto-2-propanol, 1-nonanethiol, 1-octadecanethiol, 1-octanethiol, 1-pentadecanethiol, 1-pentanethiol, 1-propanethiol, 1-tetradecanethiol, 1-tetradecanethiol Rupurum, 1-undecanthiol, 11- (1H-pyrrol-1-yl) undecan-1-thiol, 11-amino-1-undecanthiol hydrochloride, 11-bromo-1-undecanthiol, 11-mercapto-1- Undecanol, 11-mercaptoundecanoic acid, 11-mercaptoundecyl trifluoroacetate, 11-mercaptoundecyl phosphate, 12-mercaptododecanoic acid, 15-mercaptopentadecanoic acid, 16-mercaptohexadecanoic acid, 1H, 1H, 2H, 2H-perfluorodecanethiol, 2,2 ′-(ethylenedioxy) diethanethiol, 2,3-butanedithiol, 2-butanethiol, 2-ethylhexanethiol, 2-methyl-1-propanethiol, 2-methyl 2-propanethiol, 2-phenyl Ethanethiol, 3,3,4,4,5,5,6,6,6-nonafluoro-1-hexanethiolpurum, 3- (dimethoxymethylsilyl) -1-propanethiol, 3-chloro-1-propanethiol 3-mercapto-1-propanol, 3-mercapto-2-butanol, 3-mercapto-N-nonylpropionamide, 3-mercaptopropionic acid, 3-mercaptopropyl functionalized silica gel, 3-methyl-1-butanethiol 4,4′-bis (mercaptomethyl) biphenyl, 4,4′-dimercaptostilbene, 4- (6-mercaptohexyloxy) benzyl alcohol, 4-cyano-1-butanethiol, 4-mercapto-1-butanol 6- (ferrocenyl) hexanethiol, 6-mercapto-1-hexanol, 6 Mercaptohexanoic acid, 8-mercapto-1-octanol, 8-mercaptooctanoic acid, 9-mercapto-1-nonanol, biphenyl-4,4′-dithiol, butyl 3-mercaptopropionate, copper 1-butanethiolate (I ), Cyclohexanethiol, cyclopentanethiol, decanethiol functionalized silver nanoparticles, dodecanethiol functionalized gold nanoparticles, dodecanethiol functionalized silver nanoparticles, hexa (ethylene glycol) mono-11- (acetylthio) undecyl Ether, mercaptosuccinic acid, methyl 3-mercaptopropionate, NanoTether BPA-HH, NanoThinks ™ 18, NanoThinks ™ 8, NanoThinks ™ ACID11, NanoThinks ™ CID16, NanoThinks (TM) ALCO11, NanoThinks (TM) THIO8, octane thiol functionalized gold nanoparticles, PEG dithiol average _M n 8,000, PEG dithiol average molecular weight 1,500, PEG dithiol average molecular weight 3,400, S- (11-Bromoundecyl) thioacetate, S- (4-cyanobutyl) thioacetate, thiophenol, triethylene glycol mono-11-mercaptoundecyl ether, trimethylolpropane tris (3-mercaptopropionate), [11- ( Methylcarbonylthio) undecyl] tetra (ethylene glycol), m-carborane-9-thiol, p-terphenyl-4,4 ''-dithiol, tert-dodecyl mercaptan, and tert-no An agent selected from the group consisting of mercaptan containing 1 or more, bismuth according to claim 2 - thiol composition. A method of preparing a bismuth-thiol composition comprising a plurality of microparticles comprising a bismuth-thiol (BT) compound, wherein substantially all of the microparticles have a volume average diameter of about 0.4 μm to about 5 μm. ,
(A) under conditions and time sufficient to obtain a solution substantially free of solid precipitate, (i) comprising a bismuth salt containing bismuth at a concentration of at least 50 mM, and hydrophilic, polar or organic solubilization (Ii) admixing an acidic aqueous solution without agent with a sufficient amount of ethanol to obtain an admixture comprising about 25% by volume ethanol; and (b) forming a precipitate comprising microparticles comprising a BT compound. An ethanolic solution containing a thiol-containing compound present in the reaction solution in a molar ratio of about 1: 3 to about 3: 1 to bismuth under conditions and time sufficient to Obtaining a reaction solution.   8. The method of claim 7, further comprising recovering the precipitate to remove impurities. The method according to claim 7, wherein the bismuth salt is Bi (NO ₃ ) ₃ .   8. The method of claim 7, wherein the acidic aqueous solution comprises at least 5 wt%, 10 wt%, 15 wt%, 20 wt%, 22 wt% or 22.5 wt% bismuth.   Acidic aqueous solution at least 0.5 wt%, 1 wt%, 1.5 wt%, 2 wt%, 2.5 wt%, 3 wt%. 8. The method of claim 7, comprising 3.5%, 4%, 4.5% or 5% by weight nitric acid. Thiol-containing compound is 1,2-ethanedithiol, 2,3-dimercaptopropanol, pyrithione, dithioerythritol, 3,4-dimercaptotoluene, 2,3-butanedithiol, 1,3-propanedithiol, 2-hydroxy Propanethiol, 1-mercapto-2-propanol, dithiothreitol, α-lipoic acid, methanethiol (CH ₃ SH [m-mercaptan]), ethanethiol (C ₂ H ₅ SH [e-mercaptan]), 1- propanethiol _{(C 3 H 7 SH [n} -P mercaptans]), 2- propanethiol _{(CH 3 CH (SH) CH} 3 [2C 3 mercaptan]), butanethiol _{(C 4} H 9 SH [n- butyl mercaptan] ), Tert-butyl mercaptan (C (CH ₃ ) ₃ SH [t-butyl mecaptan Lucaptan]), pentanethiol (C ₅ H ₁₁ SH [pentyl mercaptan]), coenzyme A, lipoamide, glutathione, cysteine, cystine, 2-mercaptoethanol, 2-mercaptoindole, transglutaminase, (11-mercaptoundecyl) hexa (Ethylene glycol), (11-mercaptoundecyl) tetra (ethylene glycol), (11-mercaptoundecyl) tetra (ethylene glycol) functionalized gold nanoparticles, 1,1 ′, 4 ′, 1 ″ -ter Phenyl-4-thiol, 1,11-undecanedithiol, 1,16-hexadecanedithiol, technical grade 1,2-ethanedithiol, 1,3-propanedithiol, 1,4-benzenedimethanethiol, 1, 4-butanedithiol, 1,4- Tandithiol diacetate, 1,5-pentanedithiol, 1,6-hexanedithiol, 1,8-octanedithiol, 1,9-nonanedithiol, adamantanethiol, 1-butanethiol, 1-decanethiol, 1-dodecane Thiol, 1-heptanethiol, 1-heptanethiolpurum, 1-hexadecanethiol, 1-hexanethiol, 1-mercapto (triethylene glycol), 1-mercapto (triethylene glycol) methyl ether functionalized gold nanoparticles, 1 -Mercapto-2-propanol, 1-nonanethiol, 1-octadecanethiol, 1-octanethiol, 1-pentadecanethiol, 1-pentanethiol, 1-propanethiol, 1-tetradecanethiol, 1-tetradecanethiol plum, 1- Undecanethiol, 11- (1H-pyrrol-1-yl) undecane-1-thiol, 11-amino-1-undecanethiol hydrochloride, 11-bromo-1-undecanthiol, 11-mercapto-1-undecanol, 11- Mercaptoundecanoic acid, 11-mercaptoundecyl trifluoroacetate, 11-mercaptoundecyl phosphate, 12-mercaptododecanoic acid, 15-mercaptopentadecanoic acid, 16-mercaptohexadecanoic acid, 1H, 1H, 2H, 2H-perfluorodecane Thiol, 2,2 '-(ethylenedioxy) diethanethiol, 2,3-butanedithiol, 2-butanethiol, 2-ethylhexanethiol, 2-methyl-1-propanethiol, 2-methyl-2-propane Thiol, 2-phenylethanethiol 3,3,4,4,5,5,6,6,6-nonafluoro-1-hexanethiolpurum, 3- (dimethoxymethylsilyl) -1-propanethiol, 3-chloro-1-propanethiol, 3 -Mercapto-1-propanol, 3-mercapto-2-butanol, 3-mercapto-N-nonylpropionamide, 3-mercaptopropionic acid, 3-mercaptopropyl functionalized silica gel, 3-methyl-1-butanethiol, 4 , 4′-bis (mercaptomethyl) biphenyl, 4,4′-dimercaptostilbene, 4- (6-mercaptohexyloxy) benzyl alcohol, 4-cyano-1-butanethiol, 4-mercapto-1-butanol, 6 -(Ferrocenyl) hexanethiol, 6-mercapto-1-hexanol, 6-mercaptohe Sanic acid, 8-mercapto-1-octanol, 8-mercaptooctanoic acid, 9-mercapto-1-nonanol, biphenyl-4,4′-dithiol, butyl 3-mercaptopropionate, copper 1-butanethiolate (I) , Cyclohexanethiol, cyclopentanethiol, decanethiol functionalized silver nanoparticles, dodecanethiol functionalized gold nanoparticles, dodecanethiol functionalized silver nanoparticles, hexa (ethylene glycol) mono-11- (acetylthio) undecyl ether , Mercaptosuccinic acid, methyl 3-mercaptopropionate, NanoTether BPA-HH, NanoThinks ™ 18, NanoThinks ™ 8, NanoThinks ™ ACID11, NanoThinks ™ ACID16, AnoThinks (TM) ALCO11, NanoThinks (TM) THIO8, octane thiol functionalized gold nanoparticles, PEG dithiol average _M n 8,000, PEG dithiol average molecular weight 1,500, PEG dithiol average molecular weight 3,400, S- (11 -Bromoundecyl) thioacetate, S- (4-cyanobutyl) thioacetate, thiophenol, triethylene glycol mono-11-mercaptoundecyl ether, trimethylolpropane tris (3-mercaptopropionate), [11- (methylcarbonyl Thio) undecyl] tetra (ethylene glycol), m-carborane-9-thiol, p-terphenyl-4,4 ''-dithiol, tert-dodecyl mercaptan, and tert-nonyl mercapta From comprising one or more agents selected from the group consisting The method of claim 7. (I) prevention of infection of natural surfaces by bacterial, fungal or viral pathogens,
(Ii) inhibition of cell viability or cell growth of virtually all planktonic cells of bacterial, fungal or viral pathogenesis,
(Iii) inhibition of biofilm formation by bacterial, fungal or viral pathogens, and (iv) inhibition of biofilm viability or biofilm development of virtually all biofilm type cells of bacterial, fungal or viral pathogens,
Contacting the natural surface with an effective amount of the BT composition at conditions and for a time sufficient for one or more of
The BT composition comprises a plurality of microparticles comprising a bismuth-thiol (BT) compound, substantially all of the microparticles have a volume average diameter of about 0.4 μm to about 5 μm.
A method of protecting a natural surface against one or more of bacterial, fungal and viral pathogens. Bacterial pathogens
(I) one or more gram-negative bacteria;
(Ii) one or more gram-positive bacteria;
(Iii) one or more antibiotic-sensitive bacteria;
(Iv) one or more antibiotic-resistant bacteria;
(V) Staphylococcus aureus (S. aureus), MRSA (methicillin resistant S. aureus), Staphylococcus epidermidis, MRSE (methicillin resistant S. epidermidis), Mycobacterium tuberculosis, Mycobacterium Avium, Pseudomonas aeruginosa, drug resistance Aeruginosa, Escherichia coli, enterotoxigenic E. coli E. coli, enterohemorrhagic E. coli E. coli, Klebsiella pneumoniae, Clostridium difficile, Helicobacter pylori, Legionella pneumophila, Enterococcus faecalis, Methicillin-susceptible enterococcus fecaris, Enterobacter cloacae, Salmonella typhimurium, Proteus bulgaris, Yersinia enterico , Vibrio cholere, Shigella flexneri, vancomycin-resistant enterococcus (VRE), Burkholderia cepacia group, Francisella larensis, Bacillus anthracis, Yersinia pestis, Pseudomonas aeruginosa, Streptococcus pneumoniae, Escherichia coli, Burkholderia cepacia, ba Kuhoruderia multi borane scan, bacterial pathogens selected from the group consisting of Mycobacterium smegmatis and Acinetobacter Baumani,
14. The method of claim 13, comprising at least one of:   14. The method of claim 13, wherein the bacterial pathogen exhibits antibiotic resistance.   14. The method of claim 13, wherein the bacterial pathogen is resistant to an antibiotic selected from the group consisting of methicillin, vancomycin, nafcillin, gentamicin, ampicillin, chloramphenicol, doxycycline and tobramycin.   14. The method of claim 13, wherein the surface comprises an epithelial tissue surface selected from the group consisting of epidermis, dermis, respiratory tract, gastrointestinal tract, and glandular lining.   14. The method of claim 13, wherein the contacting step is performed one or more times.   The method of claim 18, wherein the at least one contacting step comprises one of spraying, ligating, dipping and applying the surface.   19. The method of claim 18, wherein the at least one contacting step comprises one of inhalation, ingestion and oral ligation.   At least one contact step can be local, intraperitoneal, oral, parenteral, intravenous, intraarterial, transdermal, sublingual, subcutaneous, intramuscular, buccal, intranasal, inhalation, intraocular, intraauricular, cardiac 19. The method of claim 18, comprising administering to the subject by a route selected from indoor, intrafacial, intraarticular and intrathecal.   BT composition is BisBAL, BisEDT, Bis-dimercaprol, Bis-DTT, Bis-2-mercaptoethanol, Bis-DTE, Bis-Pyr, Bis-Ery, Bis-Tol, Bis-BDT, Bis-PDT, Bis-Pyr / Bal, Bis-Pyr / BDT, Bis-Pyr / EDT, Bis-Pyr / PDT, Bis-Pyr / Tol, Bis-Pyr / Ery, Bismuth-1-mercapto-2-propanol, and Bis-EDT The method according to claim 13, comprising one or more BT compounds selected from the group consisting of / 2-hydroxy-1-propanethiol.   14. The method of claim 13, wherein the bacterial pathogen exhibits antibiotic resistance.   Antibiotics that enhance (i) synergizing antibiotics and (ii) coordinated antimicrobial efficacy, in any order, simultaneously or sequentially with respect to contacting the surface with the BT composition 24. The method of any one of claims 13 to 23, further comprising contacting with at least one of.   Synergistic antibiotics or antibiotics that enhance coordinated antimicrobial efficacy include aminoglycoside antibiotics, carbapenem antibiotics, cephalosporin antibiotics, fluoroquinolone antibiotics, glycopeptide antibiotics, lincosamide antibiotics 25. The method of claim 24, comprising an antibiotic selected from the group consisting of: penicillinase resistant penicillin antibiotics, and aminopenicillin antibiotics.   An aminoglycoside antibiotic selected from the group consisting of amikacin, arbekacin, gentamicin, kanamycin, neomycin, netilmycin, paromomycin, rhodostreptomycin, streptomycin, tobramycin and apramycin 26. The method of claim 25, wherein the method is a substance. (I) prevention of infection caused by bacterial pathogens on the surface of epithelial tissue,
(Ii) inhibition of cell viability or cell growth of virtually all planktonic cells of bacterial pathogenesis,
(Iii) inhibition of biofilm formation by bacterial pathogenesis, and (iv) inhibition of biofilm viability or biofilm development of virtually all biofilm-type cells of bacterial pathogenesis,
An epithelial tissue surface in an effective amount of (1) at least one bismuth-thiol (BT) composition and (2) at least one BT composition at conditions and time sufficient for one or more of The BT composition comprises a bismuth-thiol (contained in any order simultaneously with or sequentially with at least one antibiotic capable of being enhanced by or synergistically with BT) a plurality of microparticles comprising a compound, wherein substantially all of the microparticles have a volume average diameter of about 0.4 μm to about 5 μm, thereby overcoming antibiotic resistance at the surface of epithelial tissue A method to overcome antibiotic resistance on natural surfaces where substance-resistant bacterial pathogens exist.   Bacterial pathogens are Staphylococcus aureus (S. aureus), MRSA (methicillin resistant S. aureus), Staphylococcus epidermidis, MRSE (methicillin resistant S. epidermidis), Mycobacterium tuberculosis, Mycobacterium・ Abium, Pseudomonas aeruginosa, drug resistance Aeruginosa, Escherichia coli, enterotoxigenic E. coli E. coli, enterohemorrhagic E. coli E. coli, Klebsiella pneumoniae, Clostridium difficile, Helicobacter pylori, Legionella pneumophila, Enterococcus faecalis, Methicillin-susceptible enterococcus fecaris, Enterobacter cloacae, Salmonella typhimurium, Proteus bulgaris, Yersinia enterico , Vibrio cholere, Shigella flexneri, vancomycin-resistant enterococcus (VRE), Burkholderia cepacia group, Francisella larensis, Bacillus anthracis, Yersinia pestis, Pseudomonas aeruginosa, Streptococcus pneumoniae, Escherichia coli, Burkholderia cepacia, ba Kuhoruderia multi borane scan is selected from the group consisting of Mycobacterium smegmatis and Acinetobacter Baumani The method of claim 27.   The bacterial pathogen is resistant to an antibiotic selected from the group consisting of methicillin, vancomycin, nafcillin, gentamicin, ampicillin, chloramphenicol, doxycycline, tobramycin, clindamycin and gatifloxacin. The method described.   28. The method of claim 27, wherein the surface comprises an epithelial surface of tissue selected from the group consisting of the epidermis, dermis, respiratory tract, gastrointestinal tract, and glandular lining.   28. The method of claim 27, wherein the contacting step is performed one or more times.   32. The method of claim 31, wherein the at least one contacting step comprises one of spraying, irrigation, dip coating, and applying the surface.   32. The method of claim 31, wherein the at least one contacting step comprises one of inhalation, ingestion and oral ligation.   At least one contact step can be local, intraperitoneal, oral, parenteral, intravenous, intraarterial, transdermal, sublingual, subcutaneous, intramuscular, buccal, intranasal, inhalation, intraocular, intraauricular, cardiac 32. The method of claim 31, comprising administering to the subject by a route selected from indoor, intrafacial, intraarticular and intrathecal.   BT composition is BisBAL, BisEDT, Bis-dimercaprol, Bis-DTT, Bis-2-mercaptoethanol, Bis-DTE, Bis-Pyr, Bis-Ery, Bis-Tol, Bis-BDT, Bis-PDT, Bis-Pyr / Bal, Bis-Pyr / BDT, Bis-Pyr / EDT, Bis-Pyr / PDT, Bis-Pyr / Tol, Bis-Pyr / Ery, Bismuth-1-mercapto-2-propanol, and Bis-EDT 28. The method of claim 27, comprising one or more BT compounds selected from the group consisting of / 2-hydroxy-1-propanethiol.   Synergistic or potentiating antibiotics include clindamycin, gatifloxacin, aminoglycoside antibiotics, carbapenem antibiotics, cephalosporin antibiotics, fluoroquinolone antibiotics, penicillinase resistant penicillin antibiotics, and aminopenicillin 36. The method of claim 35, comprising an antibiotic selected from the group consisting of systemic antibiotics.   The synergistic or potentiating antibiotic is an aminoglycoside antibiotic selected from the group consisting of amikacin, arbekacin, gentamicin, kanamycin, neomycin, netilmycin, paromomycin, rhodostreptomycin, streptomycin, tobramycin and apramycin. The method described. (A) at least one BT composition comprising a plurality of microparticles comprising a bismuth-thiol (BT) compound, wherein substantially all of the microparticles have a volume average diameter of about 0.4 μm to about 5 μm;
(B) at least one antibiotic compound capable of acting synergistically or potentiating with the BT compound;
An antiseptic composition for treating a natural surface containing a bacterial biofilm comprising:   BT compounds are BisBAL, BisEDT, Bis-dimercaprol, Bis-DTT, Bis-2-mercaptoethanol, Bis-DTE, Bis-Pyr, Bis-Ery, Bis-Tol, Bis-BDT, Bis-PDT, Bis -Pyr / Bal, Bis-Pyr / BDT, Bis-Pyr / EDT, Bis-Pyr / PDT, Bis-Pyr / Tol, Bis-Pyr / Ery, Bismuth-1-mercapto-2-propanol, and Bis-EDT / 40. The composition of claim 38, selected from the group consisting of 2-hydroxy-1-propanethiol.   40. The composition of claim 38, wherein the BT compound is selected from the group consisting of BisEDT and BisBAL.   40. The antibiotic compound comprises an antibiotic selected from methicillin, vancomycin, nafcillin, gentamicin, ampicillin, chloramphenicol, doxycycline, tobramycin, clindamycin, gatifloxacin, cefazolin and aminoglycoside antibiotics. A composition according to 1.   42. The composition of claim 41, wherein the aminoglycoside antibiotic is selected from the group consisting of amikacin, arbekacin, gentamicin, kanamycin, neomycin, netilmycin, paromomycin, rhodostreptomycin, streptomycin, tobramycin and apramycin.   42. The composition of claim 41, wherein the aminoglycoside antibiotic is amikacin. (A) a bacterial infection on or within a natural surface comprising one of (i) gram positive bacteria, (ii) gram negative bacteria, and (iii) both (i) and (ii) And (b) administering a formulation comprising one or more bismuth thiol (BT) compositions to the surface, thereby treating the surface.
A method of treating a natural surface containing a bacterial biofilm, wherein (i) when the bacterial infection comprises gram positive bacteria, the combination comprises an effective amount of at least one BT compound and , And at least one antibiotic that is rifamycin,
(Ii) if the bacterial infection comprises gram negative bacteria, the combination comprises an effective amount of at least one BT compound and amikacin;
(Iii) The method wherein, when the bacterial infection comprises both gram positive and gram negative bacteria, the combination comprises an effective amount of one or more of BT compounds, rifamycin and amikacin.   45. The method of claim 44, wherein the bacterial infection comprises one or more antibiotic resistant bacteria.   Treating comprises at least one of (i) eradicating bacterial biofilm, (ii) reducing bacterial biofilm, and (iii) inhibiting bacterial biofilm growth, 46. The method of claim 45.   47. Any of claims 44 to 46, wherein the BT composition comprises a plurality of microparticles comprising a bismuth-thiol (BT) compound, and substantially all of the microparticles have a volume average diameter of about 0.4 μm to about 5 μm. The method according to claim 1.

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