KR101966867B1 - Bismuth-thiols as antiseptics for biomedical uses, including treatment of bacterial biofilms and other uses - Google Patents

Abstract

(BT) compound and a particular antibiotic, in order to provide a formulation comprising an antimicrobial agent, an antiseptic agent, an antiseptic agent, an antiseptic agent, Compositions and methods comprising a homogeneous particulate suspension are described. The described BT compounds and BT compounds, including the desired efficacy of certain such compositions for treating certain Gram-positive bacterial infections, and the apparent desirable efficacy of certain of such compositions for treating certain Gram-negative bacterial infections, And antimicrobial properties of antibiotic combinations that have not previously been anticipated are also described.

Description

BISMUTH-THIOLS AS ANTISEPTICS FOR BIOMEDICAL USES, INCLUDING TREATMENT OF BACTERIAL BIOFILMS AND OTHER USES As a preservative for biomedical uses, including treatment of bacterial bacterium and other uses,

[Before and after reference to related application]

This application claims the benefit of PCT Application No. PCT / US2010 / 023108, filed February 3, 2010, and U.S. Provisional Patent Application No. 61 / 373,188, filed August 12, 2010, The full text of which is incorporated herein by reference.

[TECHNICAL FIELD]

The presently disclosed embodiments of the invention relate to compositions and methods for the treatment of microbial infections. In particular, this aspect relates to the management of bacterial infections in epithelial tissues, including treatment of bacterial biofilms and other conditions, such as chronic wounds and wounds such as acute wounds, and in clinical, personal health care, and other contexts ≪ / RTI >

Description of Related Technology

A complex family of harmonized cell and molecular interactions that contribute to skin wound healing, respond to microbial infections, are resistant to microbial infections and / or contribute to the healing or maintenance of body tissues in general, For example, clinical treatments that may increase the risk of infection), topical or systemic administration of antibiotics (which may affect cell growth, migration or other infections and may also be selected for antibiotic-resistant microorganisms) The need for open-air exposure of the wound to accelerate healing, transient prosthetic matrix matrix or scaffold material, removal of dead tissue, and / or the need for infected or necrotic tissue And / or other factors such as repeated surgery to remove other factors There.

Thus, wound healing continues to be a tremendous challenge to clinicians around the world. Current treatments for intractable wounds are impractical and ineffective, and often require multiple operations to seal the wound. For example, Regranex ^® (Bekaplermin, manufacturer: Ortho-McNeil Pharmaceutical, Inc., Distributor: Ethicon, Inc., recombinant platelet-derived growth factor) is one of the few available therapies for chronic wounds But are expensive to produce and have limited clinical use.

Chronic and acute wounds and scar

Wounds may be transmitted between cells in a tissue or between tissues, for example, by physical, mechanical, biological, pathological and / or chemical forces (e.g., burns, skin infections, Respiratory disorders such as skin ulcers, radiation poisons, malignant cancers, gangrene, autoimmune diseases, immunodeficiency diseases, inhalation or infections, gastrointestinal injuries such as those caused by harmful ingestion or infection, circulation and blood disorders including coagulation defects ) Or by other traumatic injuries or the like.

The limited level of bacterial contamination or " colonization " of wounds may not necessarily interfere with the process of wound healing, while the presence of a sufficient number of bacteria to suppress the host immune defense may be due to the presence of bacterial membranes Existing wounds or acute wounds or chronic wounds, such as bacterial growth, can lead to wound infections that are detrimental to the host (Bryant and Nix, Acute and Chronic Wounds: Current Management Concepts, 2006 Mosby (Elsevier) , NY; Baronoski, Wound Care Essentials: Practical Principles (2nd Ed.), 2007 Lippincott, Williams and Wilkins, Philadelphia, PA]. For example, acute wounds can result from damage, trauma, surgical operations or other causes, which typically can quickly lack potential health problems and healing, but sometimes fail to do so due to the presence of the infection; Rapid formation of bacterial membranes has been described in acute wounds (e.g., WO / 2007/061942). Additional factors that may contribute to the progression of chronic wounds include mobility (e.g., producing sustained pressure applied to the wound site), lack of sensation or mental force, inaccessibility of the wound site (e.g., Gastrointestinal tract) and circulatory defect. Infection at the chronic wound site can be detected by skin congestion, edema, pus formation and / or unpleasant odor or other relevant, clinically acceptable criteria.

Thus, chronic wounds may be present in the upper organism (including humans and other mammals) when the immune system of the host is suppressed by a bacterial infection of the wound site (e.g., an acute wound) , But not limited to), the process can proceed to invade the tissue and further destroy it. Generally, chronic wounds are wounds that are not cured within three months, and instead of being smaller they tend to grow larger as bacterial infiltration progresses. Chronic wounds can be very painful and stressful to the patient if the nerve is damaged as the wound progresses (neuropathy). These injuries affect about four million Americans each year and cost about $ 9 billion in treatment costs. Individuals who are suffering are older than 60 years of age.

Chronic injuries may in some cases originate as acute injuries, and thus may be caused by, for example, gunshot or fragment wounds, burns, glans, nerve ulcers, pressure ulcers, diabetic ulcers, radiation intoxication, malignancies, skin infections, Diabetic foot ulcers, pressure ulcers, venous leg ulcers, infected and / or bacterium-containing non-healing surgical wounds, necrotizing papillomatosis, trauma wounds, acute arterial insufficiency, necrotizing fasciitis, osteomyelitis (bone infections) For example, it may include radiation necrosis and soft tissue necrosis, or other types of wounds. For example, venous ulcers may include poor circulation (e.g., ischemia), malfunctioning valve of the vein, or repeated physical trauma (e. G., Repeated injury). Pressure ulcers can be present, for example, when the local pressure exerted at or near the wound site is greater than the blood pressure, such that poor circulation, paralysis and / or pressure ulcers can contribute to or worsen chronic wounds. Diabetic ulcers may be caused by individuals with diabetes mellitus, such as uncontrolled hyperglycaemia, which contributes to the loss of sensation in the limbs, resulting in repeated impairment and / or neglecting individual attention to injury in some do. Factors that can be complicated or that can affect the outcome of clinical onset and chronic wounds include, but are not limited to, the immunological status of the subject (e.g., immunosuppression, pathologically (eg, HIV-AIDS) Or pharmacologically compromised immune system; age; stress); Skin aging (including photochemical aging), and the development and progression of the bacterium in the wound. In the case of epithelial tissue in the respiratory and / or gastrointestinal tract, difficulties in developing epithelial surface-clearing fluid forces or localized microenvironments that are helpful for microbial survival can lead to clinical complications .

Wound-related injuries may be accompanied by lost or compromised period function, shock, bleeding and / or thrombosis, apoptosis (e.g., necrosis and / or apoptosis), stress and / or microbial infection. Any or all of these phenomena, and in particular infection, can delay or prevent effective tissue repair processes involved in wound healing. Therefore, it is important to remove tissue that is not viable from the wound site in an individual who has sustained the wound, a process referred to as early tissue depletion as early as possible, and to remove any foreign material from the wound site, also referred to as wound cleansing It may be important to remove it.

Severe wounds, acute wounds, chronic wounds, burns, and ulcers can be beneficial from cell wound dressings. Apligraft ^® [manufactured by Novartis ^{(Norvartis)], Demagraft ®,} Biobrane ®, Transcyte ® [ manufactured by Advanced Tissue Sciences (Advance Tissue Science)], Integra ® Dermal Regeneration Template ® [ manufactured by Integra Life Scientific System Technology (Integra Life Sciences Technology), and OrCel ^® , are available for unhealed wounds or burns. However, these products are not designed to focus on the problem of bacterial tissue invasion and wound spread.

Unfortunately, systemic antibiotics are not effective in the treatment of chronic wounds and generally are not used unless an acute bacterial infection is present. Current trials involve the administration or application of antibiotics, but such therapeutic agents may promote the emergence of antibiotic-resistant bacterial strains and / or may not be effective against bacterial bacteria. Thus, it may be particularly important to use preservatives when drug resistant bacteria (e.g., methicillin resistant Staphylococcus aureus or MRSA) are detected. Although there are many preservatives widely used, established bacterial populations or sub-populations may not respond to these agents, or any other currently available treatments. Also, such preservatives are not suitable because many preservatives may be toxic to host cells at concentrations that may be required to be effective against established bacterial infections. The problem is solved by the use of a respiratory device such as a respiratory device (e.g., airway, coin and larynx, tracheal, lung, bronchus, bronchus, alveoli, etc.) or gastrointestinal ) Duct, or in an effort to clear an infection from a natural surface, including other epithelial surfaces.

Particularly problematic is the fact that cell-cell adhesion results in the release of single-celled (" cell-free " cells) into organized, multicellular populations with significantly different susceptibility to environmental agents including behavioral, gene expression, Is an infection composed of bacterial membranes, which is the organization of relatively recently recognized bacteria, which collects " planktonic " bacteria. Fungal membranes can operate biological defense mechanisms not found in planktonic bacteria, and this mechanism can protect bacterial populations against antibiotic and host immune responses. Established membranes can be blocked by a tissue-healing process.

Common microbial contamination sources that are persistent and potentially harmful are S. S. aureus, for example, MRSA [Methicillin Resistant Staphylococcus aureus], Enterococcus, E. coli. E. coli, blood. P. aeruginosa, Streptococcus, and Acinetobacter baumannii. ≪ / RTI > Some of these organisms exhibit the ability to survive on non-nutritional clinical surfaces to the oral cavity. s. S. aureus has been shown to survive for 4 weeks in dried glass and between 3 months and 6 months in dried blood and cotton fibers (Domenico et al., 1999 Infect. Immun. 67: 664-669). this. E. coli and p. Aeruginosa (P. aeruginosa) in dry blood and cotton fiber S. It has been found to survive much longer than Aureus.

Microbial membranes are associated with substantially increased resistance to both preservatives and antibiotics. Membrane morphology is generated when bacteria and / or fungi adhere to the surface. This attachment initiates altered transcription of the gene, resulting in secretion of the polysaccharide matrix, which is remarkably resilient and difficult to penetrate, thereby protecting the microorganism. In addition to their very substantial resistance to antibiotics, bacterial membranes are also highly resistant to the mammalian immune system. Preventing bacterial growth is a very important clinical priority, because once the membranes are established they are very difficult to eradicate. Recent investigations have shown that open wounds can be rapidly contaminated by the bacteria. These microbial membranes are believed to delay wound healing and are very closely related to the establishment of a serious wound infection.

Current guidance for protection for military wounds includes, for example, vigorous and thorough cleaning or irrigation and removal of dead tissue (see Blankenship CL, Guidelines for care of open combat casualty wounds, Fleet Operations and Support, US Bureau of Medicine and Surgery). Although such early intervention is important, it is not appropriate to prevent the progress of the infection. An additional therapeutic step is to reduce the chance of establishing wound infections and wound sclerosing membranes that are established by promoting healing and reducing microbial bio-burden after ingestion of dead tissue.

Due to the complex nature of the wounded wound, the potential for infection is large, especially when introducing external objects and introducing other environmental contaminants. Both the respiratory and clinical environment (including those within both of these environments) serve as an important source of potentially pathogenic microorganisms, especially for those suffering from open and / or complex wounds. Acute and chronic wounds, including surgical and rescue wounds, are already infected; It compromises the body's primary defense against the skin and the barrier. Thus, a wound exposes the interior of the body (humid and nutritive environment) to opportunistic and pathogenic infections. Many of these infections, particularly persistent wound infections, may be associated with bacterial formation, as evidenced in the case of chronic wounds (James et al., 2008). In hospitals, wound infections constitute one of the most common causes of infection in hospitals, and wounds in military and natural disaster environments are particularly susceptible to microbial contamination. Gunshot wounds are typically associated with tissue damage, tend to be extensive and profound, foreign bodies may be introduced to interfere with local blood supply, may be associated with fractures and burns, and may induce shock and compromised immune defense It is easy to get infected.

Skin structure and wound healing

Vascular epithelium that forms a barrier between the organism and its external environment, such as those found in the skin and also in the inner walls of the respiratory tract and gastrointestinal tract, Surface) is important for the health and survival of humans and other animals. Skin is the largest body organ that protects humans and other higher vertebrate animals (eg, mammals) from environmental damage through its barrier function, mechanical strength and impermeability to water. As an important environmental interface, the skin provides a protective body covering to maintain physiological equilibrium.

The skin structure is well known. In summary, the epidermis, the outer layer of the skin, is covered with the stratum corneum, the protective layer of dead epidermal cells (e. G., Keratinocytes) and extracellular connective tissue proteins. The epithelium undergoes a continuous process of exfoliating, replacing essential epithelial granule cells, spiny cells, and new substances that are pushed up from the basal cell layer, where continuous cell division and protein synthesis can lead to new skin cells and skin proteins For example, keratin, collagen). The dermis lies beneath the epithelium and is an elaboration of dermal fibroblasts of connective tissue proteins (eg collagen, elastin, etc.) that are assembled into a fibrous structure that imparts flexibility, strength and elasticity to the extracellular matrix and skin. ) Region. The skin also contains nerves, blood vessels, smooth muscle cells, hair follicles and skin oil spots.

As the body's first line of defense, skin is a major target for clinical damage, such as physical, mechanical, chemical, and biological (eg, biofluid, autoimmune) attacks that can alter its structure and function. Skin is also considered an important component of the immunological defense of organisms. In the skin, epithelial resin [Langerhans] cells with strong antigen-labeling activity that contribute to immune protection and migrating leukocyte cells (eg, lymphocytes, macrophages, mast cells) . Pigmented melanocytes in the basal layer absorb potentially harmful ultraviolet (UV) radiation. Skin destruction presents an undesirable risk to a subject, including those associated with opportunistic infections, incomplete or inappropriate tissue remodeling, scar formation, damaged mobility, pain and / or other complications. Like skin, other epithelial surfaces (eg, respiratory tract, gastrointestinal tract, and fossa lining) can have severe health hazards, such as infection or other destruction, by having defined structural properties at the time of health.

Damaged or torn skin can be generated, for example, from blemishes, abrasions, scratches, abrasions, burns, burns (including chemical burns), infections, extreme temperatures, incisions (eg, surgical incisions) have. Thus, effective skin recovery through wound healing can be clearly desirable in these and similar contexts.

Although skin may exhibit remarkable capacity for self-healing after many types of damage, skin healing does not occur rapidly enough and / or improper cytotoxic mechanisms result in integrity, barrier properties, mechanical strength, elasticity, flexibility, Or incompletely remodeled skin that may lack other desirable properties of undamaged skin. Thus, skin wound healing represents such an associated challenge with respect to chronic wounds, for example.

Wound healing occurs in three dynamic and overlapping phases, beginning with the formation of fibrin clotting. Such clotting provides a temporary storage and storage of growth factors that attack cells into the wound. It is also provided as a transient extracellular matrix (ECM) in which cells invade during recovery. It is inflammatory, mixed with clot formation, characterized by macrophage infiltration into the wound and neutrophils, which releases growth factors that amplify the early healing response, eliminating wound scrap and germs. The process of restoring the scraped area is initiated by the proliferation of healing and is induced by chemokines, cytokines, and proteases secreted from immune cells and concentrated in the coagulum. The keratinocytes are stimulated to proliferate and migrate to form a new layer of epithelium that covers the wound while wound angiogenesis delivers oxygen, nutrients and inflammatory cells to the injured site. The remodeling phase is the final phase of wound repair, which is performed by myofiber blasts that promote contraction of connective tissue, increase wound strength, and deposit ECMs that form scars.

Bismuth thiol- (BT) preservatives

Many natural products (e.g., antibiotics) and synthetic chemicals with antimicrobial and especially antimicrobial properties are known in the art and have chemical structure and antimicrobial effects, for example, the ability to kill microorganisms Colonizing or infecting a site, bacterial secretion of an exopolysaccharide, and / or the ability of an exopolysaccharide to secrete bacterial secretions and / By the ability to inhibit microbial function, such as the conversion of microorganisms into bacterial populations or expansion of bacterial formation. Antiseptics, antiseptics, etc. (including bismuth-thiol or antimicrobial agents), including factors affecting the selection and use of such compositions, including, for example, bactericidal or bacteriostatic efficacy, effective concentrations and toxicity risks to host tissues. BT compounds) are discussed, for example, in U.S. Patent No. 6,582,719.

Bismuth, a Group 5 metal, is an element like silver with antimicrobial properties. Bismuth itself may not be therapeutically useful and may exhibit certain undesirable properties, so it may instead be typically administered by delivery means using a complexing agent, carrier and / or other vehicle, the most common of which is Pepto Bismol ^® , Where the bismuth is coupled (chelated) with the subsalicylate. The previous studies have shown that the combination of certain thiol- (-SH, sulfhydryl) containing compounds such as ethanol dithiol with bismuth provides an exemplary bismuth thiol (BT) compound compared to other currently available bismuth formulations , And the improvement of the antimicrobial efficacy of bismuth was measured. There are a number of thiol compounds that can be used to produce BT (see, e.g., Domenico et al., 2001 Antimicrob. Agent. Chemotherap. 45 (5): 1417-1421, Domenico et al., 1997 Antimicrob. Agent. Chemother. 41 (8): 1697-1703, and US RE 37373, US 6,248,371, US 6,086,921, and US 6,380,248; See also, for example, U.S. Patent No. 6,582,719). Some of these preparations can inhibit the formation of a membrane.

The BT compound inhibits MRSA [methicillin resistance < RTI ID = 0.0 > S. S. aureus], MRSE [resistant to methicillin S. S. epidermidis], Mycobacterium tuberculosis, Mycobacterium avium, drug-resistant blood. A. aeruginosa, enterotoxin production. E. coli, intestinal hemorrhagic. For example, Klebsiella pneumoniae, Clostridium difficile, Heliobacter pylori, Legionella pneumophila, Enterococcus faecalis, , Enterobacter cloacae, Salmonella typhimurium, Proteus vulgaris, Yersinia enterocolitica, Vibrio cholerae, and Shigella plexa, The 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 BT role also has been shown to reduce bacterial pathogenicity, to inhibit or kill antibiotic-resistant microorganisms (gram-positive and gram-negative) in a broad spectrum, to prevent bacterial growth, to prevent septic shock, to treat sepsis, (See, for example, Domenico et al., 2001 Agents Chemother. ≪ RTI ID = 0.0 > et al. 45: 1417-1421; Domenico et al., 2000 Infect. Med. 17: 123-127; Domenico et al., 2003 Res. Adv. In Antimicrob. Agents & Chemother. 3: 79-85; 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; Veloira et al. , 2003 J Antimicrob. Chemother. 52: 915-919; Wu et al., 2002 Am J Respir Cell Mol Biol. 26: 731-738).

Despite the availability of BT compounds well over a decade, the effective selection of appropriate BT compounds for specific infectious disease applications remains an unattainable goal, where the behavior of the special BT for special microorganisms is unpredictable, Here, the synergistic activity of the special antibiotic and the special BT for the special microorganism can not be predicted. Here, the in vitro BT effect can not always predict the BT effect in vivo. Here, the planktonic (single-cell) microorganism The BT effect on the population can not predict the BT effect on the microbial community, such as bacterial organisms. In addition, limitations on solubility, tissue permeability, bioavailability, bio-distribution, etc., hinder the ability to deliver clinical benefits safely and effectively in the case of some BT compounds. The invention described herein focuses on these needs and provides other related advantages.

A brief summary

First, and without being bound by theory, as described herein, the bismuth-thiol (BT) compound is used as a preservative for the treatment of a wide variety of clinical infectious diseases and conditions and for use in personal healthcare, according to certain embodiments described herein While at the same time reducing the costs imposed on the treatment of such infections, including the savings realized by prevention or prevention, at least partially mediated by BT.

Also, in certain embodiments, as described herein, bacteria associated with bacterial formation, including one or more BT compounds and one or more antibiotic compounds (e.g., bacteria capable of forming a bacterium or otherwise capable of promoting a bacterium ), Or a formulation for treating tissues and / or surfaces containing bacteria bacterium, wherein a suitably selected combination of BT compound (s) and antibiotic (s) based on the present disclosure in accordance with the non-limiting theory, In anticipation of the unexpected enhancement effect for prevention, prevention and / or therapeutically effective treatment of microbial infections including the antibacterial (including anti-bacterial) effect of the formulation and / or infection containing bacteria bacterium, Provides unpredictable synergy

Also provided herein are unprecedented bismuth-thiol compositions comprising a substantially monodisperse microparticulate suspension, and methods and uses for their synthesis.

According to certain embodiments of the present invention described herein, a bismuth-thiol composition comprising a plurality of particulates comprising a bismuth-thiol (BT) compound having a volume mean diameter of substantially all of the particles of from about 0.4 microns to about 5 microns Wherein the BT compound comprises a bismuth or bismuth salt and a thiol-containing compound. In another embodiment substantially all of the particulates have a volume average diameter of from about 0.4 microns to about 5 microns, and (a) sufficient time and sufficient conditions to obtain a solution substantially free of solid precipitate, wherein (i) at least 50 mM (Ii) at least about 5%, 10%, 15%, 20%, 25%, or 30% by volume ethanol of a bismuth salt containing a bismuth salt, Mixing with an amount of ethanol sufficient to obtain the mixture comprising; And (b) the BT compound-containing ethanolic solution containing the thiol-containing compound present in the reaction solution at a molar ratio of about 1: 3 to about 3: 1 based on bismuth to the mixture of step (a) Thiourea composition is formed by a process comprising the steps of: adding a reaction solution under sufficient conditions and for a sufficient time to form a precipitate to form a bismuth-thiol composition. 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% and 22.5 wt% bismuth. In certain embodiments, the acidic aqueous solution comprises at least 0.5 wt%, 1 wt%, 1.5 wt%, 2 wt%, 2.5 wt%, 3 wt%, 3.5 wt%, 4 wt%, 4.5 wt% . In certain embodiments, the thiol-containing compound is selected from the group consisting of 1,2-ethanedithiol, 2,3-dimercaptopropanol, pyrithione, dithioerythritol, 3,4- dimercaptotoluene, 2,3- , 3-propanedithiol, 2-hydroxypropanethiol, 1-mercapto-2-propanol, dithioerythritol, alpha-lipoic acid and dithiothreitol.

(I) a bismuth salt comprising bismuth at a concentration of at least 50 mM, wherein the bismuth salt comprises a hydrophilic, polar or organic (Ii) mixing at least about 5% by volume, 10% by volume, 15% by volume, 20% by volume, 25% by volume or 30% by volume of ethanol with the solubilized acidic aqueous solution; And (b) adding to the mixture of step (a) an ethanolic solution comprising a thiol-containing compound in a molar ratio of about 1: 3 to about 3: 1 relative to bismuth in the reaction mixture, Bismuth thiol (BT), wherein the volume average diameter of substantially all of the fine particles is from about 0.4 탆 to about 5 탆, comprising the step of adding a sufficient amount of a sufficient amount of time to form a precipitate containing bismuth-thiol A method of preparing a bismuth-thiol composition comprising a plurality of fine particles comprising a compound is provided. In certain embodiments, the method further comprises recovering the precipitate to remove impurities. In a particular embodiment, the salt of bismuth 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 comprises at least 0.5 wt%, 1 wt%, 1.5 wt%, 2 wt%, 2.5 wt%, 3 wt%, 3.5 wt%, 4 wt%, 4.5 wt% . In certain embodiments, the thiol-containing compound is selected from the group consisting of 1,2-ethanedithiol, 2,3-dimercaptopropanol, pyrithione, dithioerythritol, 3,4- dimercaptotoluene, 2,3- (CH ₃ SH [m-mercaptan]), 3-propanediol, 3-propanediol, 2-hydroxypropanethiol, 1-mercapto-2-propanol, dithioerythritol, dithiothreitol, , ethanethiol (C ₂ H ₅ SH [e- mercaptan;), propane-1-thiol (C ₃ H ₇ SH [nP mercaptan;), 2-propane thiol _{(CH 3 CH (SH) CH} 3 [2C 3 mercaptans;), butane thiol _{(C 4 H 9 SH ([} n- butyl mercaptan]), tert-butyl mercaptan _{(C (CH 3) 3 SH} [t- butyl mercaptan]), pentane thiol (C ₅ H- ₁₁ SH [pentyl mercaptan]), coenzyme A, lipoamide, glutathione, cysteine, cystine, 2-mercaptoethanol, dithiothreitol, dithioerythritol, 2-mercaptoindole, transglutaminase , (11-mercaptoundecyl) hexa (ethylene glycol), (11-mercaptoundecyl) tetra (ethylene glycol), (11- (Ethylene glycol) functionalized gold nanoparticles, 1,1 ', 4', 1 "-terphenyl-4-thiol, 1,11-undecanedithiol, 1,16-hexadecanedithiol, 1,2- Ethanedithiol technical grade, 1,3-propanedithiol, 1,4-benzenedimethanethiol, 1,4-butanedithiol, 1,4-butanedithiol diacetate, 1,5-pentanedithiol, 1 1-butanethiol, 1-dodecanethiol, 1-heptanethiol, 1-heptanethiol, 1-heptanethiol, 1-heptanethiol, -Heptanethiol fumarate, 1-hexadecanethiol, 1-hexanethiol, 1-mercapto- (triethylene glycol), 1-mercapto- (triethylene glycol) methyl ether functionalized gold nanoparticles, 1-mercapto- Octanethiol, 1-octanethiol, 1-pentadecanethiol, 1-pentanethiol, 1-propanethiol, 1-tetradecanethiol, 1- Undecane thiol, 11- (1H-pyrrol-1-yl) undecane-1-thiol, 11-amino-1-undecanethiol hydrochloride, 1-undecanol, 11-mercaptoundecanoic acid, 11-mercaptundecanoic acid, 11-mercaptoundecyltrifluoro acetate, 11-mercapto-undecyl phosphoric acid, 12-mercapto Todo acid, 12-mercapto Todo acid, 15-mercapto-pentadecane acid, 16-mercapto-hexadecanoic acid, 16-mercapto-hexadecanoic acid, 1 H , 1 H , 2 H , 2 H -perfluorodecanethiol, 2,2 '- (ethylene dioxy) 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-hexanethiol , 3 (dimethoxymethylsilyl) propane-1-thiol, 3-chloro-propane-1-thiol, 3-mercapto-1-propanol, 3-mercapto-2-butanol, 3-mercapto - N - nonyl 3-mercaptopropionic acid, 3-mercaptopropyl-functionalized silica gel, 3-methyl-1-butanethiol, 4,4'-bis (mercaptomethyl) biphenyl, 4,4'- dimercapto Stilbene, 4- (6- Hexanoloxy) benzyl alcohol, 4-cyano-1-butanethiol, 4-mercapto-1-butanol, 6- (ferrocenyl) hexanethiol, 6-mercapto- Mercaptoacetic acid, 9-mercapto-1-nonanol, biphenyl-4,4'-dithiol, butyl 3-mercaptopropionate, copper I) 1-butanethiolate, cyclohexanethiol, cyclopentanethiol, decanthiol functionalized silver nanoparticles, dodecanthiol functionalized gold nanoparticles, dodecanthiol functionalized silver nanoparticles, hexa (ethylene glycol) mono-11 Mercapto-succinic acid, methyl 3-mercaptopropionate, nanoTether BPA-HH, NanoThinks ^TM 18, NanoThinks ^TM 8, NanoThinks ^TM ACID11, NanoThinks ^TM ACID16, NanoThinks ^TM ALCO11, NanoThinks ^TM TH108, octanethiol-functionalized gold nanoparticles, PEG dithiol, average Mn 8,000, PEG dithiol average molecular weight (mol wt) 1,500, PEG dithiol average molecular weight 3,400, S- (11-bromoundecyl) thioacetate, S- (4-cyanobutyl) thioacetate, thiophenol, triethylene glycol mono-11-mercapto undecyl ether, trimethylolpropane tris (3-mercaptopropionate), [11- (methylcarbonylthio) One or more agents selected from the group consisting of tetra (ethylene glycol), m -carborane-9-thiol, p -terphenyl-4,4'-dithiol, tert-dodecylmercaptan, .

(Ii) inhibiting the viability or cell growth of substantially all of the planktonic cells of a bacterial, fungal or viral pathogen; (iii) Inhibiting bacterial growth by bacteria, fungi or viral pathogens, and (iv) inhibiting bacterial growth of bacterial, fungal or viral pathogens or bacterial growth of substantially all bacterial-forming cells under conditions sufficient and sufficient time (BT) compound, wherein the volume average diameter of substantially all of the fine particles is from about 0.4 microns to about 5 microns, with an epithelial tissue surface comprising a plurality of microparticles comprising a bismuth-thiol , Bacterial pathogens, fungal pathogens, and viral pathogens, including biological tissue surfaces such as epithelial tissue surfaces Is provided a method of protecting natural surface. In certain embodiments, bacterial pathogens comprise (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. (S. aureus), MRSA (methicillin-resistant, S. aureus), Staphylococcus epidermidis, MRSE (methicillin-resistant S. epidermidis), Mai Mycobacterium tuberculosis, Mycobacterium avium, Pseudomonas aeruginosa, drug-resistant pemphigus, Mycobacterium tuberculosis, Mycobacterium tuberculosis, Mycobacterium tuberculosis, Mycobacterium avium, Pseudomonas aeruginosa, Escherichia coli, enterotoxin production. Coli, intestinal hemorrhagic. Klebsiella pneumoniae, Clostridium difficile, Heliobacter pylori, Legionella pneumophila, Enterococcus faecalis, Enterococcus faecalis, Enterococcus faecalis, Klebsiella pneumoniae, Clostridium difficile, Heliobacter pylori, Enterococcus faecalis, Enterobacter cloacae, Salmonella typhimurium, Proteus vulgaris, Yersinia enterocolitica, and the like. , Vibrio cholera, Shigella flexneri, vancomycin-resistant Enterococcus (VRE), Burkholderia cepacia complex, Francesella tularensis (Francisella tularensis), Bacillus anthracis, Yersinia pestis, Pseudomonas spp. The compounds of the present invention may be used in combination with other antibiotics such as Pseudomonas aeruginosa, vancomycin-resistant enterococci, Streptococcus pneumonia, Penicillin-resistant Streptococcus pneumonia, Escherichia coli, At least one of bacterial pathogens selected from the group consisting of Burkholderia cepacia, Bukholderia multivorans, Mycobacterium smegmatis and Acinetobacter baumannii . In certain embodiments, bacterial pathogens exhibit antibiotic resistance. In certain embodiments, the bacterial pathogen is resistant to an antibiotic selected from methicillin, vancomycin, napthyline, gentamicin, ampicillin, chloramphenicol, doxycycline and tobramycin.

In certain embodiments, the natural surface comprises an oral / intraoral surface. In a further embodiment, the natural surface includes biological surfaces such as bone, joints, muscles, ligaments or tendons.

In certain embodiments, the surface comprises an epithelial tissue surface comprising a tissue selected from the group consisting of epithelium, skin, respiratory tract, gastrointestinal tract, and fistula.

In certain embodiments, the contacting step is performed one or more times. In certain embodiments, at least one step of contact includes one of spraying, perfusing, dipping, and painting a natural surface. In certain embodiments, at least one contacting step comprises one of inhalation, digestion, and oral perfusion. In certain embodiments, the at least one contacting step is administered topically, intraperitoneally, orally, parenterally, intravenously, intraarterially, percutaneously, sublingually, subcutaneously, intracutaneously, intramuscularly, intranasally, Intravenous, intraperitoneal, intrathecal, subcutaneous, intrathecal, intraarticular, and intrathecally. In a particular embodiment, the BT composition is selected from the group consisting of BisBAL, BisEDT, bis-dimer caproyl, bis-DTT, bis-2-mercaptoethanol, bis-DTE, bis-Pyr, bis- -PDT, bis-Pyr / Bal, bis-Pyr / BDT, bis-Pyr / EDT, bis-Pyr / PDT, bis-Pyr / Tol, bis- And bis-EDT / 2-hydroxy-1-propanethiol.

In certain embodiments, bacterial pathogens exhibit antibiotic resistance. In certain other embodiments, the methods described above further comprise contacting the natural surface in any order with respect to the step of contacting the surface with the BT composition, either simultaneously or sequentially with the synergistic antibiotic and / or the enhancing antibiotic do. In certain embodiments, the synergistic and / or enhancing antibiotic is selected from the group consisting of an aminoglycoside antibiotic, a carbapenem antibiotic, a cephalosporin antibiotic, a fluoroquinolone antibiotic, a glycopeptide antibiotic, a rinkosamid antibiotic, a penicillinase-resistant penicillin antibiotic, ≪ / RTI > aminophenicil antibiotics. In certain embodiments, the synergistic and / or enhancing antibiotic is selected from the group consisting of amikacin, arbekacin, gentamicin, kanamycin, neomycin, nethymicin, paromomycin, rhodostryptomycin, streptomycin, tobramycin, 0.0 > aminoglycoside < / RTI > antibiotics.

In another aspect of the invention described herein, there is provided a method of treating a bacterial pathogen, comprising: (i) preventing infection of a surface by a bacterial pathogen, (ii) inhibiting cell viability or cell growth of substantially all of the planktonic cells of the bacterial pathogen, (iii) , And (iv) inhibiting the bacterial growth of bacterial pathogens or the growth of bacterial growth of substantially all of the bacterial-forming cells under conditions sufficient for sufficient time and for at least one time that the volume average diameter of substantially all of the microparticles (1) at least one bismuth-thiol (BT) composition, and (2) at least one BT composition, comprising a plurality of microparticles comprising a bismuth-thiol Comprising contacting an effective amount of at least one antibiotic that can act synergistically and / or ameliorate with the surface simultaneously or sequentially and in any order, A method of overcoming antibiotic resistance on a natural surface on which the agent-resistant bacterial pathogen exists (e.g., a method of overcoming the antimicrobial effect of at least one antibiotic known to have an anti-bacterial effect on bacteria of the same bacterial species) In the case of bacterial pathogens that are resistant to antibiotics, such pathogens are susceptible to antibiotics. In certain embodiments, bacterial pathogens comprise (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 Disco), Mycobacterium tuberculosis, Mycobacterium avium, Pseudomonas aeruginosa, drug-resistant blood. Escherichia coli, enterotoxin production. Coli, intestinal hemorrhagic. E. coli, Enterococcus faecalis, Enterobacter cloacae, Salmonella typhimurium, Enterococcus faecalis, Enterococcus faecalis, Enterococcus faecalis, Enterococcus faecalis, Enterococcus faecalis, , V. vulnificus enterococcus (VRE), brucolierea sephacia complex, francicella tularensis, bacillus anthracis, yersinia, bacteriocin-resistant Enterococcus, Resistant Streptococcus pneumoniae, Penicillin-resistant Streptococcus pneumoniae, Escherichia coli, Bourcolieria sephacia, Bacolieria juvenile, Staphylococcus aureus, Staphylococcus aureus, Staphylococcus aureus, At least one of the bacterial pathogens selected from < RTI ID = 0.0 > M. < / RTI > .

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

In certain embodiments, the natural surface comprises an oral / intraoral surface. In a further embodiment, the natural surface includes biological surfaces such as bone, joints, muscles, ligaments or tendons.

In certain embodiments, the surface comprises a tissue selected from the group consisting of epithelium, skin, respiratory tract, gastrointestinal tract, and the inner lining layer. In certain embodiments, the contacting step is performed one or more times. In certain embodiments, at least one step of contacting includes one of spraying, perfusing, dipping and painting a natural surface. In certain other embodiments, the at least one contacting step comprises one of inhalation, digestion, and oral perfusion. In certain embodiments, the at least one contacting step is administered topically, intraperitoneally, orally, parenterally, intravenously, intraarterially, percutaneously, sublingually, subcutaneously, intramuscularly, intrathecally, intranasally, Intravenous, intraperitoneal, intrathecal, subcutaneous, intrathecal, intraarticular, and intrathecally. In a particular embodiment, the BT composition is selected from the group consisting of BisBAL, BisEDT, bis-dimer caproyl, bis-DTT, bis-2-mercaptoethanol, bis-DTE, bis-Pyr, bis- - PDT, Bis-Pyr / Bal, Bis-Pyr / BDT, Bis-Pyr / EDT, Bis- Pyr / PDT, Bis- Pyr / Tol, Bis- Pyr / Ery, And bis-EDT / 2-hydroxy-1-propanethiol. In certain embodiments, the synergistic and / or enhancing antibiotic is selected from the group consisting of clindamycin, gatifloxacin, aminoglycoside antibiotics, capbepenem antibiotics, cephalosporin antibiotics, fluoroquinolone antibiotics, glycopeptide antibiotics, Penicillinase-resistant penicillin antibiotics, and aminophenicillin antibiotics. In certain embodiments, the synergistic and / or enhancing antibiotic is selected from the group consisting of amikacin, arbekacin, gentamicin, kanamycin, neomycin, nethymicin, paromomycin, rhodostryptomycin, streptomycin, tobramycin, 0.0 > aminoglycoside < / RTI > antibiotics.

Returning to another aspect, there is provided a kit comprising: (a) at least one BT compound; (b) at least one antibiotic compound that can be enhanced and / or act synergistically with the BT compound; And (c) a pharmaceutically acceptable carrier or excipient comprising a topical carrier. In certain embodiments, the BT compound is selected from the group consisting of BisBAL, BisEDT, Bis-Dimercaprol, Bis-DTT, Bis-2-mercaptoethanol, Bis-DTE, Bis- Pyr, Bis- -PDT, bis-Pyr / Bal, bis-Pyr / BDT, bis-Pyr / EDT, bis-Pyr / PDT, bis-Pyr / Tol, bis- And bis-EDT / 2-hydroxy-1-propanethiol. In a particular embodiment, the BT composition comprises a plurality of fine particles comprising a bismuth-thiol (BT) compound, wherein substantially all of the fine particles have a volume average diameter of from about 0.4 microns to about 5 microns. In certain embodiments, the BT compound is selected from BisEDT and BisBAL. In certain embodiments, the antibiotic compound comprises an antibiotic selected from the group consisting of methicillin, vancomycin, napisylin, gentamicin, ampicillin, chloramphenicol, doxycycline, tobramycin, clindamycin, gatifloxacin and aminoglycoside antibiotics. In certain embodiments, the aminoglycoside antibiotic is selected from amikacin, arbekacin, gentamicin, kanamycin, neomycin, nethymicin, paromomycin, rhodostreptomycin, streptomycin, tobramycin and apramycin. In certain embodiments, the aminoglycoside antibiotic is amikacin.

Confirming that in certain other embodiments (a) it comprises one of (i) a gram-positive bacterium, (ii) a gram negative bacterium, and (iii) both (i) and (ii) either on or on the surface; And (b) treating the surface by administering to the surface a formulation comprising at least one bismuth thiol (BT) composition, wherein a (i) ) When the bacterial infection comprises gram-positive bacteria, the formulation comprises a therapeutically effective amount of at least one BT compound and at least one antibiotic that is rifamycin, and (ii) when the bacterial infection comprises gram negative bacteria, Comprises a therapeutically effective amount of at least one BT compound and amikacin, and (iii) when the bacterial infection comprises both gram-positive and gram-negative bacteria, the formulation comprises a therapeutically effective amount of one or more BT compounds, Mucin and amikacin.

In certain embodiments, the bacterium comprises one or more antibiotic-resistant bacteria. In certain embodiments, the treatment of the surface may include (i) eradicating the bacteria germ layer; (ii) reducing the bacteria germ layer, and (iii) damaging the growth of the bacterial germ layer. In certain embodiments, the BT composition comprises a plurality of particulates comprising a bismuth-thiol (BT) compound, wherein the volume average diameter of substantially all of the particulates is from about 0.4 microns to about 5 microns.

These and other aspects of the present disclosure as described in aspects of the present invention will become apparent with reference to the following detailed description and accompanying drawings. Such as those described and / or referred to in the application bibliography, including U. S. Patent No. RE37,793, U.S. Patent No. 6,248,371, U.S. Patent No. 6,086,921, and U.S. Patent No. 6,380,248, All of the publications, U.S. Patents, foreign patents, foreign patent sources, and non-patent publications are incorporated herein by reference in their entirety, such as when each is incorporated separately. Aspects and aspects of the present invention may, in some instances, be modified to use the concepts of various patents, patent applications, and publications, still providing additional aspects.

details

A particular aspect of the invention described herein is that certain bismuth-thiol (BT) compounds, such as those provided herein, having a volume average diameter of less than < RTI ID = 0.0 & A strong antiseptic agent against a particular bacterium, including bacteria associated with a number of clinically significant infections, including infections that may include bacterial fungal membranes, in certain embodiments including BT microparticles of about 0.4 [mu] m to about 5 [ , Anti-bacterial and / or anti-bacterial activity.

Unexpectedly, not all BT compounds are uniformly effective against these bacteria in a predictable fashion, but instead showed different efficacy depending on the target bacterial species. Particularly, and particularly as described herein, certain BT compounds (preferably including BT particulates having a volume mean diameter of about 0.4 탆 to about 5 탆) exhibit higher efficacy for Gram-negative bacteria, Other BT compounds (including BT microparts, preferably having a volume mean diameter of about 0.4 microns to about 5 microns), can be used in accordance with a non-limiting theory to produce a clinically relevant strategy for the management of bacterial infections, including bacterial bacterial infection It has been found that in a manner that can be provided for the first time, it exhibits greater efficacy against Gram-positive bacteria.

Also, and as described in more detail below, certain aspects of the invention described herein relate to particle size (e.g., volume mean diameter is about 0.4 microns to about 5 microns), as described herein To produce a formulation comprising a plurality of BT microparticles that are substantially monodisperse. In certain of these and related embodiments, the particulate BT is not provided as a component of a liquid follicle or liposome, such as multilamellar phospholipocholine-cholesterol liposomes or other multilamellar or monolayer lamellar liposomal vesicles.

In connection with certain embodiments, as also described herein, antibiotic and anti-bacterial efficacy of certain antibiotics, which are antibiotics that have already been found to lack strong therapeutic effects against such bacterial infections, (I. E., Increased in a statistically significant manner) by treatment with the above and selected BT compounds, simultaneously or sequentially and in any order. In a manner not previously anticipated by the present disclosure, the particular BT compound may be combined with a particular antibiotic to provide an elevated level of anti-bacterial and / or anti-bacterial activity against a particular bacterial species or bacterial strain, as provided herein Action or combination of actions. As described in more detail below, the unexpected characteristic of this combination is that certain BT / antibiotic combinations exhibit synergistic or synergistic effects on certain bacteria, but certain other BT / Or failed to exhibit enhanced anti-bacterial and / or anti-bacterial activity.

According to these and related embodiments, antibiotics and BT compounds may be administered simultaneously or sequentially and in any order and may be used for the treatment of special infections (e. G., Bacteria formed by gram-negative or gram-positive bacteria) As described herein, the specific synergistic or improved combination of one or more antibiotics and one or more BT compounds does not exhibit predictable (e.g., only additive) activity, but instead indicates that the selected antibiotic, the selected BT compound and the specifically identified target bacteria It is noteworthy that it functions as an unexpected synergistic or enhanced [eg, super-additive] mode as a function of

For example, in connection with various natural surfaces that have been practically or efficaciously infected with microorganisms, and in conjunction with the improved substantially monodispersed particulate BT formulations, the specific antibiotic The combination of both the antibiotic compound and the BT compound, when used alone or in combination with a particular bacterial strain or species, may exhibit a limited anti-bacterial effect, (With statistical significance), the antibiotic efficacy of BT and / or BT in antibiotic efficacy according to a non-limiting theory, because the effect is bacterial and the effect is larger than the mere sum of the effects of each compound when used alone Antibiotic-BT synergistic effect (e.g., FICI? 0.5) or an improving effect (e.g., 0.5 <FICI ≤ 1.0). Thus, antibiotic-BT synergistic effects and BT-antibiotic enhancement are generally predictable, because not all, but not all, BT compounds can be elevated or reversible with all antibiotics, and not all, but antibiotics are elevated or enhanced with all BT compounds not. Instead, and according to certain embodiments described herein, the particular combination of synergistic or enhanced antibiotic and BT compounds surprisingly includes a special environment, such as a natural surface, as described herein, Which further imparts an anti-bacterial effect to the microorganism.

That is, certain BT-synergistic antibiotics are described herein and include antibiotics that can act synergistically (FICI? 0.5) with at least one BT composition comprising at least one BT compound as provided herein , Wherein this synergistic effect is greater in magnitude than the effect that antibiotics are present but in the absence of the BT compound and / or in the presence of the BT compound but in the absence of the antibiotic (i.e., statistically In a significant way). Similarly, certain BT-antibiotic combinations exhibit an enhancing action (0.5 < FICI &lt; 1.0), wherein such an enhancement occurs when antibiotics are present but the BT compound is absent and / (I. E., In a statistically significant manner with respect to the appropriate control condition) as a detectable effect. &Lt; / RTI &gt;

Examples of such detectable effects include, but are not limited to, (i) prevention of infection by bacterial pathogens, (ii) inhibition of cell growth or cell growth of substantially all planktonic cells of bacterial pathogens, (iii) Inhibition of bacterial growth, and (iv) inhibition of bacteriological viability of bacterial pathogens or bacterial growth of substantially all bacterium-forming cells, but since the present invention is not intended to be so limited, in other contemplated embodiments, -BT synergistic effect may be determined by one or more of the other clinically significant parameters, for example, the degree of resistance or sensitivity of the bacterial pathogen to one or more antibiotics or other drugs or chemicals, one or more chemical, (E.g., pH, ionic strength, temperature, pressure), and / or the degree of resistance or sensitivity of the bacterial pathogen to one or more biological agents For example, viruses, other bacteria, biologically active polynucleotides, immune cells, or free radicals such as antibodies, cytokines, chemokines, enzymes including degrading enzymes, membrane-breaking proteins, reactive oxygen species, etc.) (E. G., A statistically significant increase or decrease) of the degree of resistance or sensitivity of the substrate.

Those skilled in the art will appreciate that the effect of a special formulation on the structure, function and / or activity of a bacterial population is that provided herein that the effect of a synergistic or enhanced antibiotic-BT combination is such that one component of the combination is absent Can be measured for the purpose of ascertaining the antibiotic-BT synergistic or enhancing action present if the simple sum of the observed effects is exceeded (see, for example, Coico et al. (Eds.), Current Protocols in Microbiology, 2008, John Wiley & Sons, Hoboken, NJ; Schwalbe et al., Antimicrobial Susceptibility Testing Protocols, 2007, CRC Press, Boca Raton, FL] and various other criteria.

For example, in certain embodiments, synergism may be determined by measuring the antimicrobial efficacy, such as those described herein, using a variety of concentrations of the candidate agents (e.g., individually and in combination BT and antibiotics) Eliopoulos et al. (Eliopoulos and Moellering, (1996) Antimicrobial combinations. In Antibiotics in Laboratory Medicine (Lorian, V., Ed.), Pp. (FICI) and fractional saline bacterial concentration index (FBCI) can be calculated and measured according to the manufacturer's instructions, 330-96, Williams and Wilkins, Baltimore, MD, USA. Synergy can be defined as an FICI or FBCI index of &lt; = 0.5, and an antagonism of> 4 (see, for example, Odds, FC (2003) Synergy, antagonism, and what the chequerboard puts between them. Journal of Antimicrobial Chemotherapy 52: 1]. Synergism is also typically &lt; RTI ID = 0.0 &gt; a &lt; / RTI &gt; 4 times reduction in antibiotic concentration or alternatively, for example, in Hollander et al. (1998 Antimicrob. Agents Chemother. 42: 744.) In certain embodiments, the synergistic action is from a combination of two drugs (e.g., antibiotics and BT compositions) Where the effect of the combination is greater than the effect when the concentration of the second drug is replaced by the first drug (e.g., in a statistically significant manner).

Thus, as described herein and in certain preferred embodiments, combinations of BT and antibiotics will be understood to be synergistic when an FICI value of 0.5 or less is observed (see Odds, 2003). As further described herein, according to certain other preferred and non-limiting theories, certain BT-antibiotic combinations may exhibit an FICI value of 0.5 to 1.0, indicating a high efficacy for such synergy , Which can be observed using a non-optimal concentration of at least one BT and at least one antibiotic exhibiting synergistic antimicrobial efficacy alone or in combination. This effect may also be referred to herein as " enhanced " antibiotic activity or " enhanced " BT activity.

The enhanced antibiotic and / or BT activity is at least (i) at a concentration that is less than the characteristic minimum inhibitory concentration (MIC) of BT for the provided target microorganism (e.g., the provided bacterial species or strain) (in a statistically significant manner) One BT, and (ii) a characteristic IC ₅₀ (concentration that inhibits 50% growth of the microbial population; for example, Soothill et al., 1992 J Antimicrob Chemother 29 (2): 137] The presence of both of the at least one antibiotic at a concentration that is less than the bactericide-preventive concentration (BPC) of the antibiotic against the target microorganism provided, and / or the presence of both of the antimicrobial agent (e.g., BT or antibiotics (In a statistically significant manner) of the BT-antibiotic combination compared to the antimicrobial effect that can be observed when the same antimicrobial agent is used at the same concentration in the absence of another antimicrobial agent (e.g., antibiotic or BT) When creating a can be detected in accordance with a particular embodiment. In preferred embodiments, the "enhanced" antibiotics and / BT is active, if there is more than 1.0 and less than 0.5 FICI value, as measured.

As will be appreciated by those skilled in the art based on the present disclosure, in certain embodiments, the synergistic or enhanced antibiotic and / or BT activity may be measured using a Loewe additivity-based model (e.g., FIC index, Greco model), or Bliss independence based model (e.g., non-parametric and semi-parametric models) or those described herein and known in the art For example, Meletiadis et al., 2005 Medical Mycology 43: 133-152) using methods known in the art. Thus, an exemplary method for measuring synergistic or enhanced antibiotic and / or BT activity is described, for example, in Meletiadis et al., 2002 Rev Med Microbiol 13: 101-117; White et al., 1996 Antimicrob Agents Chemother 40: 1914-1918; Mouton et al., 1999 Antimicrob Agents Chemother 43: 2473-2478.

Certain other embodiments include methods wherein the BT compound (s) and the antibiotic (s) do not exhibit predictable (e.g., simply additive) activity in vivo, but instead comprise the selected antibiotic, the selected BT compound, One or more functions of the specifically identified target bacterial species may be used in an unexpected synergistic or enhancing manner [e.g., two or more such agents may be used in combination with the effect (E. G., Gram-negative or gram-positive &lt; / RTI &gt; One or more antibiotics as described herein that can exhibit a synergistic or enhanced effect in vivo for the treatment of one or more &lt; RTI ID = 0.0 &gt; Consider the transfer of the combination. Thus, in accordance with these and related aspects, the FICI or FBCI value (which is measured in vitro) in a particular in vivo situation may not be readily available, but instead the BT-antibiotic elevation or enhancement effect may be detected in a quantifiable matrix Lt; / RTI &gt;

For example, in one embodiment, such as in an open in vivo open fracture Rattus norvegicus femur critical defect model, as described in Example 11, an antibiotic treatment or a BT compound alone A statistically significant reduction in the observed post-treatment bacterial counts for the combined BT-antibiotic combination is an index of the enhancement or enhancement effect. Statistical significance can be determined using methods well known to those skilled in the art. In certain other embodiments, at least 5%, 10%, 20%, 30%, 40%, or 50% of the number of bacteria observed in post-treatment damage to the BT- antibiotic combination compared to the antibiotic treatment or BT compound alone, The observed reduction in these or other in vivo models is considered as an index of the enhancement or enhancement effect.

Other exemplary indices of in vivo infections can be measured according to established methodologies developed for the quantification of severity of infection, such as various wound scoring systems known to those skilled in the art (see, for example, European Wound Management Association (EWMA), Position Document: Identifying criteria for wound infection. London: MEP Ltd, Scoring System Considered in 2005]. An illustrative wound scoring system that can be used to assess the synergistic or enhancing activity of a BT-antibiotic combination as described herein can be found in ASEPSIS (Wilson AP, J Hosp Infect 1995; 29 (2): 81-86; Wilson et al., Lancet 1986; 1: 311-13), the Southampton Wound Assessment Scale (Bailey IS, Karran SE, Toyn K, et al BMJ 1992; 304: 469-71) Horan TC, Gaynes P, Martone WJ, et al., 1992 Infect Control Hosp Epidemiol 1992; 13: 606-08. In addition, a recognized clinical index of wound healing known to the skilled artisan can be determined, for example, wound size, depth, granular tissue condition, infection, etc., with or without a BT compound and / or antibiotic. Thus, and based on the present disclosure, it will be appreciated by those skilled in the art that the BT composition-antibiotic combination can be used to determine whether it alters in vivo wound healing (e.g., whether it increases or decreases in a statistically significant manner Will be readily appreciated.

In terms of these and related aspects, an effective amount (e. G., An effective amount) of a composition or formulation comprising one or more BT compounds and, optionally, one or more antibiotic compounds such as one or more synergistic antibiotics, A variety of methods are provided herein for treating microbial infected natural surfaces, such as surfaces that support or contain germ strains in a therapeutically effective amount in certain embodiments. Based on the present disclosure, it will be apparent to the present inventors that certain antibiotics may be contemplated for use in the treatment of infection types provided, wherein such antibiotics are administered to a person familiar with the art with the same type of infection It has already been considered to be ineffective for.

Thus, certain embodiments contemplate compositions comprising one or more BT compounds for use as preservatives. Preservatives are substances that kill or prevent microbial growth and are typically applied to living tissues to distinguish these classes from disinfectants that are generally applied to inanimate objects (Goodman and Gilman ' s " The Pharmacological Basis of Therapeutics "Goodman and Gilman", pp. 959-960). Typical examples of preservatives are the tinctures of ethyl alcohol and iodine (see, for example, Gilman et al., Eds, 1985, Macmillan Publishing Co. tincture. Germicides include preservatives that kill microorganisms, such as microbial pathogens.

Certain embodiments described herein may encompass compositions comprising one or more BT compounds and one or more antibiotic compounds (e.g., synergistic antibiotics and / or enhanced antibiotics as provided herein). Antibiotics are well known in the art and typically include drugs made from compounds produced by one species of microorganism to kill other species of microorganisms or synthetic products with the same or similar chemical structure and mechanism of action, , And drugs that destroy microorganisms in or on the body of a living organism, including such drugs when applied locally. Among the embodiments described herein are those in which antibiotics can belong to one of the following classes: aminoglycosides, carbapenems, cephalosporins, fluoroquinolones, glycopeptide antibiotics, rincosamides (e.g., Superstition), penicillinase-resistant penicillin, and aminopenicillin. Thus, the antibiotic may be selected from the group consisting of oxacillin, piperacillin, cefuroxime, cefaclor, cephapim, imipenem, aztreonam, streptomycin, tobramycin, tetracycline, minocycline, ciprofloxacin, levofloxacin, erythromycin, But are not limited to, but are not limited to, but are not limited to, formazin, capreomycin, isoniazid, ansamycin, carbamazepam, monobactam, nitrofuran, penicillin, quinolone, sulfonamide, clofazimine, , Pyrazinamide, rifampicin, rifampin, rifabutin, ripapentin, streptomycin, arsupenamine, chloramphenicol, phosphomycin, fosidic acid, linezolide, metronidazole, mulipyrosine, plethendimycin, quinupristin, , Rifaximin, thiamphenicol, tidinazole, aminoglycoside, beta-lactam, penicillin, cephalosporin, car But are not limited to, buprenorphine, bupenem, fluroquinolone, ketolide, rincozamide, macrolide, oxazolidinone, streptoglamine, sulfonamide, tetracycline, glycylcycline, methicillin, vancomycin, napthylline, gentamicin, ampicillin, chloramphenicol , Doxycycline, tobramycin, amikacin, arbekacin, gentamicin, kanamycin, neomycin, nethymicin, paromomycin, rhodostryptomycin, streptomycin, tobramycin, apramycin, Gatifloxacin, aminopenicillin, and others known in the art. A summary of these and other clinically useful antibiotics is available in the art and is known to those familiar with the art (see, for example, Washington University School of Medicine, The Washington Manual of Medical Therapeutics (32nd Ed.). , &Lt; / RTI &gt; 2007 Lippincott, Williams and Wilkins, Philadelphia, PA; Hauser, AL, Antibiotic Basics for Clinicians, 2007 Lippincott, Williams and Wilkins, Philadelphia, PA].

An exemplary class of antibiotics for use with one or more BT compounds in certain of the embodiments described herein is described in Edson RS, Terrell CL. The aminoglycoside. Mayo Clin Proc. 1999 May; 74 (5): 519-28). This class of antibiotics inhibits bacterial growth by damaging bacterial protein synthesis through the binding and inactivation of bacterial ribosomal subunits. In addition to these bacteriostatic properties, aminoglycosides also exhibit a bacteriostatic effect through the destruction of cell walls in Gram-negative bacteria.

Aminoglycoside antibiotics include gentamicin, amikacin, streptomycin, and others, and even though a class of resistant strains are reported, they are generally useful for the treatment of Gram-negative bacteria, mycobacteria and other microbial pathogens . Aminoglycosides are not normally absorbed through the digestive tract and are therefore not considered to be treatable with oral formulations. For example, amikacin, which is often effective against gentamicin-resistant bacterial strains, is typically administered intravenously or intramuscularly, which can cause pain to the patient. In addition, toxicity associated with aminoglycoside antibiotics such as amikacin can lead to kidney damage and / or irreversible hearing loss.

Notwithstanding these characteristics, certain embodiments described herein may be practiced in combination with an elevated BT / antibiotic combination (e. G., For example, for the treatment of epithelial tissue surfaces at one or more locations along the gastrointestinal / , Antibiotics need not be limited to aminoglycosides). Also, in certain other embodiments, the use of the compositions and methods described herein as disinfectants, which refers to agents that kill or block the growth of microorganisms on the outer surface of inanimate objects, may be contemplated.

As further described herein, the BT compound may be a bismuth or bismuth salt and a thiol- (e.g., -SH, or sulfhydryl) containing compound such as those described in Domenico et al., 1997 Antimicrob. Agent. Chemother. 41 (8): 1697-1703, Domenico et al., 2001 Antimicob. Agent. Chemother. 45 (5): 1417-1421, and U.S. Pat. No. RE37,793, U.S. Patent No. 6,248,371, U.S. Patent No. 6,086,921, and U.S. Patent No. 6,380,248; It may also be a composition comprising those described in U.S. Patent No. 6,582,719 (including methods for their production), for example. However, the specific embodiments are not limited, and other BT compounds including bismuth or bismuth salts and thiol-containing compounds can be considered. The thiol-containing compound may contain 1, 2, 3, 4, 5, 6 or more thiol (e.g., -SH) groups. In a preferred embodiment, the BT compound comprises bismuth associated with the thiol-containing compound as ion coordination and / or as a coordination complex, but in some other embodiments, bismuth may be present in the organometallic compound via a covalent bond Thiol-containing compounds. However, certain contemplated embodiments specifically exclude BT compounds, which are organometallic compounds, such as those found in covalent bonds to organic residues, of bismuth.

Exemplary BT compounds are shown in Table 1:

[Table 1]

Exemplary BT compounds ^*

* Represents the atomic ratio to a single bismuth atom for comparison, based on the known tendency of bismuth to form a ternary complex with the sulfur-containing compound and the stereochemical ratio of the reactants used. The indicated atomic ratios can not be the correct molecular formulations for all species in the formulation provided. The number in parentheses is the ratio of bismuth to "CPD", the compound, of one or more thiols (eg, Bi: thiol 1 / thiol 2).

BT compounds for use in certain presently described embodiments may be prepared using established procedures [see, for example, RE 37,793, US 6,248,371, US 6,086,921 and US 6,380,248; Domenico et al., 1997 Antimicrob. Agent. Chemother. 41 (8): 1697-1703, Domenico et al., 2001 Antimicob. Chemother. 45 (5): 1417-1421), and in certain other embodiments the BT compound may also be prepared according to the methods described herein. Thus, certain preferred embodiments contemplate the synthesis methods described herein for preparing BT compounds and in particular for obtaining BT compounds in the form of substantially monodispersed particulate, wherein the concentration of dissolved bismuth is at least 50 mM, at least 100 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 at least 1 M The acidic bismuth aqueous solution lacking the hydrophobic polarity or the organic solubilizing agent is mixed with ethanol to obtain a first ethanolic solution and the mixture is stirred under conditions and for a time sufficient to form a precipitate containing fine particles containing the BT compound , Concentration, solvent strength, temperature, pH, mixing, and / or pressure to be recognized by those skilled in the art based on the present disclosure as described herein Condition), a reaction solution is obtained in which the thiol-containing compound is present in the reaction solution in a molar ratio of about 1: 3 to about 3: 1 with respect to bismuth .

Thus, exemplary BT comprises compound 1B-1, bis-EDT (bismuth-1,2-ethanedithiol, reactant at 1: 1); 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-dimercaptrol, 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-butane dithiol, 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 / Ery (1: 1 / 0.5); Compound 15, bis-EDT / 2-hydroxy, propanethiol (1: 1/1) (see, for example, Table 1).

Without wishing to be bound by theory, it is believed that the method of making the presently described BT compounds, including producing or obtaining an acidic aqueous solution comprising bismuth, such as an aqueous nitric acid solution comprising bismuth nitrate in certain preferred embodiments, , Wherein such compositions are useful for mass production ease, improved product purity, uniformity or consistency (including uniformity in particle size), or for the manufacture and / or administration of the topical formulations Have different characteristics.

In a particular embodiment, the BT compositions prepared according to the methods described herein are substantially single dispersed suspensions of particulates having a volume mean diameter (VMD) of from about 0.4 microns to about 5 microns, according to certain currently preferred embodiments, associated with their occurrence And exhibits favorable homogeneity. Measurements of particle size can be referred to as volume mean diameter (VMD), mass median diameter (MMD), or mass median aerodynamic diameter (MMAD). These measurements can be made, for example, of impact (MMD and MMAD) or laser (VMD) characterization. For liquid particles, VMD, MMD and MMAD may be the same if environmental conditions are maintained, for example, standard humidity. However, if the humidity is not maintained, the MMD and MMAD measurements will be less than the VMD due to dehydration during impactor measurements. For purposes of this discussion, the VMD, MMD, and MMAD measurements will be compared by considering the VMD, MMD, and MMAD measurements as being under standard conditions. Similarly, a dry powder particle size measurement in MMD and MMAD is also considered to be comparable.

As described herein, a preferred embodiment relates to a substantially single dispersion suspension of BT-containing microparticles. Generation of a defined BT particle size with a limited geometric standard deviation (GSD) can be achieved, for example, by BT deposition, access to the desired target site in or on the natural surface and / or tolerability by the subject to which BT microparticles are administered Can be optimized. The narrow GSD limits the number of particles beyond the desired VDM or MMAD size range.

In one embodiment, there is provided a liquid or aerosol suspension of microparticles containing at least one BT compound described herein wherein the VMD is from about 0.5 microns to about 5 microns. In another embodiment, a liquid or aerosol suspension having from about 0.7 microns to about 4.0 microns VMP or MMAD is provided. In another embodiment, a liquid or aerosol suspension having a VMD or MMAD of about 1.0 to about 3.0 microns is provided. In certain other preferred embodiments, about 0.1 to about 5.0 microns VMD, or about 0.1, about 0.2, about 0.3, about 0.4, about 0.5, about 0.6, about 0.7, about 0.8 or about 0.9 microns to about 1.0, 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 of a BT compound as described herein There is provided a liquid suspension comprising one or more of the BT compound particles comprising.

Thus, and in certain preferred embodiments, a BT composition comprising a " substantially " monodispersed BT formulation as first described herein, for example, a BT compound in particulate form, wherein " substantially " 85%, 90%, 91%, 92%, 93%, or 94% of the particles have a mean diameter (VMD) within a defined range of, for example, , More preferably at least 95%, 96%, 97%, 98%, 99% or more of which have a VMD within the stated size range.

These and related properties of the BT compositions prepared according to the synthetic methods described herein enable lower cost and manufacturability than previously described BT and compliance with one or more pharmaceutical, pharmaceutical and cosmetic cosmetic standards Including the uniformity within the composition, which can permit its characterization in a manner that is well known to those skilled in the art.

Additionally or alternatively, the substantially monodisperse BT microparticles described herein can be used without the need for micronization, i.e., with other devices and processes typically used to produce microparticles or with expensive, labor-intensive grinding or supercritical Can be advantageously produced in the absence of fluid processing (see, for example, Martin et al. 2008 Adv. Drug Deliv. Rev. 60 (3): 339; Moribe et al., 2008 Adv. Drug Deliv. Rev. 60 (3): 328; Cape et al., 2008 Pharm. Res. 25 (9): 1967; Rasenack et al. 2004 Pharm. Dev. Technol. 9 (1): 1-13]. Thus, this embodiment includes improved and substantially uniform solubilization characteristics, suitability for the desired dosage form such as oral, inhalation or skin / topical skin topical formulations, increased bioavailability and other beneficial properties, But is not limited to, the advantageous effect of a substantially uniform particulate formulation.

BT compound microparticle suspensions may be formulated in aqueous form as suspensions or solutions in aqueous and also organic solvents, including halogenated hydrocarbon propellants, as anhydrous powders, or as suspensions or solutions in organic solvents, for example, to keep individual microparticles in suspension, May be administered in other forms, as described below, including additives, such as may be known, or other ingredients, wetting agents, surface active agents, or preparations containing mineral oil. The aqueous formulation may be aerosolized, for example, with a liquid nebulizer using hydraulic or ultrasonic granulation. The propellant-based system may use a suitable pressure distributor. A dry powder dispersing apparatus capable of effectively dispersing the BT-containing fine particles can be used. The desired particle size and distribution can be obtained by selecting an appropriate apparatus.

As also shown above, it is contemplated that, depending on the particular embodiment, the amount of the BT compound, including substantially all of these particulates having a volume mean diameter (VMD) of from about 0.1 to about 8 microns, and in certain preferred embodiments, from about 0.4 microns to about 5 microns A method of making a bismuth-thiol (BT) composition comprising a plurality of particulates is also provided herein.

In a general aspect, the method comprises the steps of: (a) providing a solution containing (i) a bismuth salt comprising (i) a concentration of bismuth of at least 50 mM, under conditions sufficient to obtain a solution substantially free of solid precipitate, (Ii) at least about 5% by volume, 10% by volume, 15% by volume, 20% by volume, 25% by volume or 30% by volume and preferably about 25% by volume of ethanol With an amount of ethanol sufficient to obtain a mixture comprising: And (b) adding to the mixture of step (a) an ethanolic solution comprising a thiol-containing compound under conditions sufficient to form a precipitate comprising a BT compound and for a sufficient time to form a thiol- In a molar ratio of about 1: 3 to about 3: 1 with respect to bismuth.

In certain preferred embodiments, the bismuth salt may be Bi (NO ₃ ) ₃ , but it will be appreciated that, in accordance with the present disclosure, bismuth may also 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 800 mM, at least 900 mM, or 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. In certain preferred embodiments, the acidic aqueous solution comprises at least 5 wt% nitric acid, and in certain other embodiments, the acidic aqueous solution contains at least 0.5 wt%, at least 1 wt%, at least 1.5 wt%, at least 2 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.

The thiol-containing compound may be any thiol-containing compound as described herein, and in a particular embodiment, 1,2-ethanedithiol, 2,3-dimercaptopropanol, pyrithione, dithioerythritol, At least one of 4-dimercaptotoluene, 2,3-butanedithiol, 1,3-propanedithiol, 2-hydroxypropanethiol, 1-mercapto-2-propanol, dithioerythritol and dithiothreitol . &Lt; / RTI &gt; Other exemplary thiol-containing compound is alpha-lipoic acid, methanethiol (CH ₃ SH [m- mercaptan;), ethanethiol (C ₂ H ₅ SH [e- mercaptan;), propane-1-thiol (C ₃ H ₇ SH [nP mercaptan;), 2-propane thiol _{(CH 3 CH (SH) CH} 3 [2C 3 mercaptan;), butane thiol _{(C 4 H 9 SH ([} n- butyl mercaptan]), tert- butyl mercaptan _{(C (CH 3) 3 SH} [t- butyl mercaptan]), thiol pentane (C ₅ H ₁₁ SH [cyclopentyl mercaptan;), coenzyme A, lipoic amide, glutathione, cysteine, cystine, 2-Merced The following thiol compounds available from Sigma-Aldrich (St. Louis, Mo.) can be used in the present invention, such as, for example, thiourea, mercaptoethanol, dithiothreitol, dithioerythritol, 2- mercaptoindole, transglutaminase and Sigma-Aldrich (Ethylene glycol), (11-mercaptoundecyl) tetra (ethylene glycol) functionalized gold nano (ethylene glycol), Particles, 1,1 ', 4', 1 "-terphenyl-4-thiol, 1,11-unde Dithiol, 1,16-hexadecanedithiol, 1,2-ethanedithiol technical grade, 1,3-propanedithiol, 1,4-benzenedimethanethiol, 1,4-butanedithiol, 1,4 Butanedithiol, 1-butanedithiol, 1-butanedithiol, 1,6-hexanedithiol, 1,8-octanedithiol, 1-heptanethiol, 1-heptanethiol, 1-hexadecanethiol, 1-hexanethiol, 1-mercapto- (triethylene glycol), 1-mercapto- Ethylene glycol) methyl ether functionalized gold nanoparticles, 1-mercapto-2-propanol, 1-nonantiol, 1-octadecanethiol, 1-octanethiol, 1-undecane thiol, 1-undecane thiol, 11- (1H-pyrrol-1-yl) undecane-1-thiol, 11-amino- 11-mercapto-decanoic acid, 11-mercapto-undecanoic acid, 11-mercapto-decanoic acid, 11-mercapto- Mercapto undecanoic acid, 11-mercaptundecanoic acid, 11-mercaptundecanoic acid, 11-mercaptoundecyl trifluoroacetate, 11-mercaptodecylphosphoric acid, 12-mercaptododecanoic acid, Hexanedioic acid, 16-mercaptohexadecanoic acid, 1H, 1H, 2H, 2H-perfluorodecanethiol, 2,2 '- (ethylene dioxy) diethanethiol, Butanethiol, 2-ethylhexanethiol, 2-methyl-1-propanediol, 2-methyl-2-propanethiol, 2-phenylethanethiol, 3,3,4,4,5,5,6,6,6 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) Phenyl, 4,4'-dimercaptostilben, 4- (6-mercaptohexyloxy) benzyl alcohol, 4-cyano-1-butanethiol, 4- Hexanethiol 6-mercaptohexanoic acid, 8-mercapto-1-octanol, 8-mercaptoonanoic acid, 9-mercapto-1-nonanol, biphenyl- Dithiol, butyl 3-mercaptopropionate, copper (I) 1-butanethiolate, cyclohexanethiol, cyclopentanethiol, decanthiol functionalized silver nanoparticles, dodecanethiol- (Ethylene glycol) mono-11- (acetylthio) undecyl ether, mercaptosuccinic acid, methyl 3-mercaptopropionate, nanoteter BPA-HH, NanoThinks ^TM 18, NanoThinks ^TM 8, NanoThinks ^TM , ACID 11, NanoThinks ^TM ACID16, NanoThinks ^TM ALC011, NanoThinks ^TM THIO8, octane thiol functionalized gold nanoparticles, PEG dithiol average Mn 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-mercapto undecyl ether, trimethylolpropane tris (3-mercaptopropionate), [11- (methylcarbonylthio) Silane] tetra (ethylene glycol), m-carboran-9-thiol, p-terphenyl-4,4'-dithiol, tert-dodecylmercaptan and tet-nonylmercaptan.

Exemplary reaction conditions are described herein, including temperature, pH, reaction time, stirring or agitation to dissolve the solute, and procedures for collecting and washing the precipitate, and techniques commonly known in the art are used do.

In contrast to the previously described method for producing BT compounds, according to the present method for making BT, the BT product has a VMD of about 0.4 to about 5 microns in certain preferred embodiments and generally about &lt; RTI ID = 0.0 &gt; Is provided as a particulate suspension having substantially all of the particulates of from 0.1 micron to about 8 microns. Further, unlike the previous attempts, according to this embodiment, bismuth has a bismuth salt concentration of at least about 50 mM to about 1 M, and at least about 0.5% to about 5% (w / w) / Weight) of nitric acid and is free of hydrophilic, polar or organic solubilizing agents.

In this regard, the process of the present invention is based on the discovery that bismuth is not 50-Meyer water-soluble (e. G., RE37793), bismuth is unstable in water (see, e.g., Kuvshinova et al. 2009 Russ. J Inorg. Chem 54 (11): 1816), bismuth provides surprising and unpredictable advantages in terms of generally acceptable teaching in the art, which is unstable in nitric acid solutions when there is no hydrophilic, polar or organic solubilizing agent. For example, a technique that defines a BT formulation method (see, for example, Domenico et al., 1997 Antimicrob. Agents. Chemother. 41: 1697; U.S. Patent 6,380,248; U.S. Re-registered Patent RE37793; No. 6,248,371), the hydrophilic solubilizing agent propylene glycol is required to dissolve the bismuth nitrate, and the bismuth concentration of the solution prepared for the reaction with the thiol is less than 15 mM, so that the available production pattern for the BT compound Limit.

In contrast, in accordance with the teachings of the present invention, there is no need for hydrophilic, polar or organic solubilizing agents to dissolve bismuth, but higher concentrations are surprisingly achieved. Hydrophilic, polar or organic solubilizing agents include propylene glycol (PG) and ethylene glycol (EG) and also include dioxane and dimethylsulfoxide (DMSO), polyols such as PG and EG and also polyethylene glycol ), Polar solvents such as polypropylene glycol (PPG), pentaerythritol, and others, polyhydric alcohols such as glycerol and mannitol, and other known solubility enhancers. Other water-miscible organic materials of high polarity include dimethylsulfoxide (DMSO), dimethylformamide (DMF) and NMP (N-methyl-2-pyrrolidone).

Thus, for example, if the solvent, including those commonly used as hydrophilic, polar or organic solubilizers as provided herein, has a water polarity (SPP) value of 0.962, an SPP value of toluene of 0.655, A system of Catalan et al., In which the SPP value of 2-propanol is 0.848 (cf., for example, 1995 Liebigs Ann. 241; (SPP) scale values using a system of the following reference: Catalan, 2001 In: Handbook of Solvents, Wypych (Ed.), Andrew Publ., NY and references cited therein. . Determination of the SPP value of the solvent based on UV-measurement of the 2-N, N-dimethyl-7-nitrofluorene / 2-fluoro-7-nitrofluorene probe / homomorph pair (See Catalan et al., 1995).

A solvent having a desired SPP value based on solubility (solubility) characteristics of a particular BT composition (whether it is a pure single component solvent or a solvent mixture of two, three, or four or more solvents; See, for example, Godfrey 1972 Chem. Technol. 2: 359), according to certain preferred embodiments of the synthetic method steps described herein, as shown above, a hydrophilic, polar or organic solubilizing agent dissolves bismuth , But may be readily ascertained by one of ordinary skill in the art in light of the present disclosure.

The solubility parameter may also be determined using the interaction parameter C, the Hildebrand solubility parameter d, or the partial [Hansen (Hansen) solubility parameter describing the polarity, hydrogen bonding potential, )] Availability parameters: δp, δh and δd. In certain embodiments, the maximum value for the solubility parameter describing the solvent or co-solvent system in which the bismuth salt containing bismuth is to be dissolved can be determined, for example, by adding the bismuth salt according to the methods currently described for preparing the particulate BT composition To provide a restriction on the aqueous solution containing it. For example, higher 5 delta values will have greater hydrogen-binding ability and therefore have greater affinity for solvent molecules such as water. Thus, a higher value of the maximum observed 6 delta for the solvent may be desirable in situations where a more hydrophilic environment is desired.

By way of non-limiting example, BisEDT having the structure of formula I shown below can be prepared according to the following scheme:

Briefly, and as a non-limiting illustrative example, an excess of (11.4 L) of 5% aqueous HNO ₃ was added to a solution of 0.331 L (about 0.575 moles) aqueous acidic bismuth solution, such as Bi (NO ₃ ) ₃ solution _{[e.g., 43% Bi (NO 3)} 3 (w / w), 5% nitric acid (w / w), 52% water (w / w), OH Cincinnati material Shepherd Chemical Company (Shepherd of Chemical Co.) is slowly added with stirring, and then anhydrous ethanol (4 L) can be slowly added. An ethanolic solution (1.56 L) of a thiol compound, for example 1,2-ethanedithiol [-0.55 M], is added to 1.5 L of anhydrous ethanol, 72.19 mL (0.863 moles) of 1,2- Followed by stirring in a 60-mL syringe and stirring for 5 minutes. 1,2-ethanedithiol (CAS 540-63-6) and other thiol compounds are commercially available, for example, from Sigma-Aldrich, St. Louis, Missouri. The ethanolic solution of the thiol compound is added to the Bi (NO ₃ ) ₃ / HNO ₃ aqueous solution with stirring overnight to form the 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 formed product is precipitated as a precipitate containing particulates as described herein, then collected by filtration and washed successively with ethanol, water and acetone to give BisEDT as a yellow amorphous powder solid. The crude product is redissolved in anhydrous ethanol with stirring, filtered, washed several times with ethanol, and washed several times with acetone. The washed powder is polished in 1M NaOH (500 mL), filtered, and washed successively with water, ethanol and acetone to obtain purified particulate BisEDT.

According to non-limiting theory, bismuth inhibits the ability of bacteria to produce extracellular polymeric substances (EPS) such as exopolysaccharides, which inhibition results in the formation of damaged membranes. Bacteria are believed to use glue-like EPS for bacterial adherence. Depending on the nature of the infection, bacterial formation and EPS synthesis can not contribute to bacterial pathogenicity, such as interfering with wound healing. However, bismuth alone is not therapeutically useful as an agent and is typically administered as part of a complex such as BT. Thus, bismuth-thiol (BT) is a family of compositions comprising compounds that result from the chelation of bismuth and a thiol compound and that includes compounds that exhibit a significant improvement in the antimicrobial therapeutic efficacy of bismuth. BT exhibits significant anti-infective, anti-bacterial, and immunomodulatory effects. Bismuth-thiol (BT) is effective against broad spectrum microorganisms and is typically not affected by antibiotic-resistance. BT can prevent bacterial formation at a significantly low (sub-inhibitory concentration) concentration, prevent many pathogenic features of common wound pathogens at sub-inhibitory concentrations, prevent septic shock in animal models, It can be synergistic.

As described herein, this synergistic effect in the antifungal effect of one or more of the defined BTs when combined with one or more defined antibiotic compounds is facilitated by a separate antibiotic against the specific bacterial type and on the profile of the BT effect Antibiotic combinations in the context of a particular bacterial population, including, but not limited to, confirmation of the presence of unexpected but surprisingly Gram-negative or Gram-positive (or both) bacteria. For example, as described herein, antibiotics that are synergistic with certain BTs include amikacin, ampicillin, aztreonam, cephazoline, cephapim, chloramphenicol, ciprofloxacin, clindamycin (or other rincoamidic antibiotics) the neoplasm? azithromycin (Cubicin ^®), doxycycline, gatifloxacin, gentamicin myth, already fetched s, levofloxacin, linezolid (Zyvox ^®), minocycline, naphthyl cylinder, wave our erythromycin, rifampin, sulfamic methoxy Sasol, tetracycline, Sat. &Lt; / RTI &gt; bramacin, and vancomycin. In vitro studies have shown that MRSA, which is poor or not entirely susceptible to, for example, gentamicin, cephazoline, cefepime, sulfamethoxazole, imipenim or levofloxacin, is exposed to antibiotics in the presence of the BT compound BisEDT Lt; RTI ID = 0.0 &gt; antibiotics &lt; / RTI &gt; Thus, certain embodiments contemplated herein include BT compound and, amikacin, ampicillin, Sefar sleepy, three pepim, chloramphenicol, ciprofloxacin, clean the myth (or other Linkoping four mid antibiotic), the neoplasm? Azithromycin (Cubicin ^®), doxycycline , gatifloxacin, gentamicin myth, already fetched s, levofloxacin, linezolid (Zyvox ^®), minocycline, naphthyl cylinder, wave our erythromycin, rifampin, sulfamic methoxy Sasol, tetracycline, soil bra one or more antibiotics selected from erythromycin and vancomycin Certain other aspects contemplated herein are contemplated to include compounds and / or methods that may include combinations of BT compounds and amikacin, ampicillin, cephazoline, and the like, wherein the one or more antibiotics are expressively excluded pepim, chloramphenicol, ciprofloxacin, clean the myth (or other Linkoping four mid antibiotic), the neoplasm? azithromycin (Cubicin ^®), doxycycline, Gatti Rock reaper, gentamicin myth, already fetched s, levofloxacin, linezolid (Zyvox ^®), minocycline, naphthyl cylinder, wave our erythromycin, rifampin, sulfamic methoxy Sasol, tetracycline, tobramycin, and combinations of one or more antibiotic selected from vancomycin Compositions and / or methods that may be included. In this connection, it is noted that gentamicin and tobramycin belong to the aminoglycoside family of antibiotics. Also, Domenico et al., 2001 Agents Chemother. 45: 1417-1421; Domenico et al., 2000 Infect. Med. 17: 123-127; Domenico et al., 2003 Res. Adv. In Antimicrob. Agents &amp; Chemother. 3: 79-85; 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; Veloira 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. Pharmaceut. 373: 141-146] is expressively excluded from certain considered embodiments; It should be noted here that none of these publications teach or suggest the monodisperse particulate BT compositions described herein.

Thus, and in certain preferred embodiments, as described herein, compositions for treating a subject with a composition comprising the particulate BT described herein and optionally and in certain other embodiments also synergistic and / or enhancing antibiotics And methods are provided. Those familiar with the relevant field will be able, based on the contents of the present invention, to determine the appropriate clinical context and circumstances in which such treatment may be desirable, in particular, for example, surgery, surgeons, dermatology, trauma medicine, (Including, for example, diabetes, obesity, etc.), infection and inflammation (including other epithelial tissue surfaces such as in the epithelial layer of the respiratory tract or gastrointestinal tract or in the gland tissue), and other related medical and subspecialities Will recognize established standards in the medical field.

Thus, in certain embodiments as described herein and as known in the art, it will be appreciated that the promotion of skin tissue repair (or other tissue repair, such as epithelial tissue, bone, joints, muscle tendons, or joint restoration) is contemplated. In certain embodiments, the promotion of dermal tissue or other epithelial tissue repair is accomplished by (i) epithelial cells (e.g. keratinocytes) or dermal fibroblast migration, (ii) epithelial cells (e. G., Keratinocytes) Growth, (iii) down regulation of epithelial cells (e.g., keratinocytes) or dermal fibroblast collagenase, gelatinase or matrix, (iv) dermal fibroblast extracellular matrix protein deposition, and (v) Stimulating or depressing one or more cytotoxic remediation activities selected from the group consisting of inducing or enhancing the formation of cells. Methods of identifying and characterizing such cytotoxic activity include, but are not limited to, the effects of compositions comprising the wound tissue repair-promoting compounds described herein, for example, a BT agent as described herein, And can be measured easily and without undue experimentation based on the contents of the description. For example, compositions and methods relating to art recognized models for wound repair based on keratinocyte wound sutures after scratched scar are disclosed.

Preferred compositions for treating a microbial infection on or within a natural surface for use in accordance with the embodiments described herein comprise a bismuth-thiol (BT) compound as described herein in certain embodiments, and certain specific but related In embodiments, it may include compositions comprising other compounds known in the art, such as one or more antibiotic compounds as described herein. BT compounds and methods for their preparation are described herein and are described, for example, in Domenico et al. 45 (5) 1417-1421) and RE37,793 (US), US Patent No. 6,248,371, United States Patent No. 6,248,371 Patent No. 6,086,921, and U.S. Patent No. 6,380,248. As also noted above, certain preferred BT compounds are those that are ionically bonded to a thiol-containing compound, such as a composition comprising bismuth chelated to a thiol-containing compound, or that contain a bismuth or bismuth salt that is a coordination complex thereto , Certain other preferred BT compounds are those containing a bismuth or bismuth salt covalently linked to a thiol-containing compound. Also, a substantially monodisperse particulate BT composition as described herein is preferred. From previous efforts to treat bacterial infections, or from other features of any of the compounds described herein for the first time in a composition and method for promoting the treatment of natural surfaces as described herein, , It was not possible to predict whether the present methods using these compounds would have the beneficial effects described herein.

Accordingly, in accordance with a preferred embodiment, a method is provided for treating a natural surface, comprising administering to the surface at least one particulate BT compound described herein. In certain embodiments, the methods may also be synergistic antibiotics as described herein in certain preferred embodiments, and in certain other preferred embodiments, at least one antibiotic compound that may be an enhancing antibiotic as described herein Concurrently or sequentially and in any order. The antibiotic compound may be an aminoglycoside antibiotic, a carbapenem antibiotic, a cephalosporin antibiotic, a fluoroquinolone antibiotic, a glycopeptide antibiotic, a rincozyme antibiotic, a penicillinase-resistant penicillin antibiotic, or an aminopenicillin antibiotic. Clinically useful antibiotics and are also discussed herein, for example, in, for example, Washington University School of Medicine, The Washington Manual of Medical Therapeutics (. 32 ^nd Ed), 2007 Lippincott, Williams and Wilkins , Philadelphia, PA; and in Hauser, AL, Antibiotic Basics for Clinicians, 2007 Lippincott, Williams and Wilkins, Philadelphia, PA.

As described herein, certain embodiments relate to a method of treating a bacterial infection comprising a Gram-positive bacterium, wherein the therapeutically effective formulation comprises a BT compound (such as BisEDT, bismuth: 1,2-ethanedithiol; BisPyr, bismuth: pyrithione; BisEDT / Pyr, bismuth: 1,2-ethanedithiol / pyrithione) and rifamycin, or BT compounds and doped Toma, who [e.g., Cubicin ^® manufactured by Pharma-ku Beast Bucharest Carlsbad (Cubist Pharmaceuticals ), Massachusetts Lexington material], or BT compounds and linezolid [Zyvox ^®, manufactured by Pfizer,, Inc. (Pfizer, Inc.), New York, NY], or BT compounds (e. g. , BisEDT, bismuth: 1,2-ethanedithiol, BisPyr, bismuth: pyrithione, BisEDT / Pyr, bismuth: 1,2-ethanedithiol / pyrithione) and at least one ampicillin, cephazoline, cefepime, chloramphenicol, Clindamycin (or other Rincomade antibiotic), Daportomycin One of Cubicin ^® , doxycycline, gatifloxacin, gentamicin, imipheline, levoflox, Zyvox ^® , napsilin, paromomycin, rifampin, sulfamethoxazole, tobramycin and vancomycin From the unexpected discovery that it may contain more than one.

As also described herein, certain embodiments are derived from unpredictable discoveries that in the case of bacterial infections involving gram negative bacteria, the preferred therapeutically effective formulations may include BT compounds and amikacin. Certain related embodiments contemplate treating an infection comprising a gram-negative bacterium with a BT compound and, in certain embodiments, other antibiotics, such as other aminoglycoside antibiotics, other than gentamicin or tobramycin. Accordingly, and in aspects of these aspects, another related aspect is a method for selecting appropriate antibiotic compound (s) for inclusion in a formulation to be administered in accordance with the present method, comprising the steps of: Considering identifying one or more bacterial populations or subgroups within or on a natural surface by well-known criteria of gram negative.

The compositions and methods described herein are typically prepared by mixing a compound described herein (e.g., with one or more particulates BT alone or with one or more synergistic and / or enhancing antibiotics as described herein) (For example, bacteria, viruses, yeast, fungi and other fungi, microbial parasites, etc.) in a wide range of contents by applying or administering them to microbial sites such as bacteria, Such natural surfaces include, but are not limited to, mammalian tissues such as skin, scalp, gastrointestinal lining, ball lumen, etc. Cells and tissue membranes such as endothelium, peritoneal membrane, pericardium, pleura, periosteum, And other mammalian tissues such as teeth, bones, joints, tendons, ligaments, muscles, heart, lungs, kidneys, liver, spleen, gallbladder, pancreas, bladder, nerves and the like.

The microparticide antimicrobial agents described herein can be used to inhibit microbial growth, reduce microbial contamination, reduce bacterial membranes, prevent the conversion of bacteria into bacteria, prevent or inhibit microbial infection, It can be used for applications. These agents may also be used for the prevention of viral infection by herpes viruses such as herpes viruses, for example, cytomegalovirus, herpes simplex virus type 1, and herpes simplex virus type 2, and / or infection by other viruses Or &lt; RTI ID = 0.0 &gt; inhibition. &Lt; / RTI &gt; In this regard, the preparations can be used for the treatment of various diseases such as Japanese chain RNA virus, Japanese chain DNA virus, Rous sarcoma virus (RSV), hepatitis A virus, hepatitis B virus (HBV), hepatitis C virus (HCV) Viruses, West nile virus (WNV), Epstein-Barr virus (EBV), eastern equine encephalitis virus (EEEV), severe acute respiratory virus And is useful for preventing or inhibiting viral infection by various viruses such as SARS, human immunodeficiency virus (HIV), human papilloma virus (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, yeast and fungi (e.g. Candida (e.g. Candida albicans, Candida glabrata, , Fungi such as C. parapsilosis, C. tropicalis, and C. dubliniensis or Cryptococcus or other fungi infections, Infection, Helicobacter pylori infection, and the treatment or prevention of digestive ulcer disease. In one embodiment, the agent is not generally fatal to the bacterium, but is nonetheless natural Is used at a dose sufficient to reduce the protective polysaccharide coating that is able to withstand the immune response of the human body. Thus, It is believed that the secondary mediated elimination-up of microbial infection if not harmful to the immune system (for example, a bunch of normal intestinal bacteria, etc.).

In a non-limiting and enumerated manner, certain considered aspects are now described.

In certain embodiments, the particulate BT compound (or composition comprising a particulate BT compound) described herein can be combined with at least one anti-mycotic agent to regulate bacterial development, disrupt the bacterial membrane, or reduce the amount of bacterial membrane have. As will be appreciated in the art, interspecies quorum sensing is associated with membrane formation. Certain agents that increase LuxS-dependent pathways or interspecies quorum sensing signals (see, e.g., U.S. Patent No. 7,427,408) contribute to regulating the development and / or proliferation of the myocardial membrane. Exemplary formulations include, by way of example and without limitation, combinations or combinations of N- (3-oxododecanoyl) -L-homoserine lactone (OdDHL) blocking compound and N-butyryl-L-homoserine lactone (BHL) (See, for example, U.S. Patent No. 6,455,031). Oral hygiene compositions comprising a particulate BT compound and at least one anti-fungal agent can be delivered topically for the destruction and inhibition of bacterial membranes and for the treatment of periodontal disease (see, e.g., U.S. Patent No. 6,726,898 number).

Compositions comprising particulate bismuth- thiols and uses for oral hygiene and for the treatment of inflammation and oral infections .

In another embodiment, a composition comprising a particulate BT compound is formulated orally and can be used in a method for preventing or reducing microbial growth in the mouth and for preventing and / or treating microbial infection and inflammation in the mouth. These compositions are therefore useful for the prevention or treatment of plaque, mouth odor, oral inflammation, periodontal disease, gingivitis, and other infections of the mouth (i.e., plaque, mouth odor, oral inflammation, periodontal disease, gingivitis, Or to reduce the likelihood of occurrence or recurrence thereof). Oral compositions comprising particulate BT compounds can also be used to prevent and / or regulate (i.e., slow down, slow down, or inhibit) bacterial development, to destroy bacterial membranes, or to inhibit the growth of bacterial membranes May be useful in reducing the amount.

Trapped food particles, poor oral hygiene and poor oral health, and improper cleaning of the dentures can promote microbial growth on the teeth, around the gums, and on the tongue. The presence of continuous microbial growth and cavities can produce bad breath, plaque (ie, bacteria formed by colonization of microorganisms), gingivitis, and inflammation. In the absence of proper oral care (e.g., toothbrushing, dental flossing), more severe infections such as periodontal disease and infection of the lower jaw may follow.

Good oral hygiene is important for oral health as well as prevention of serious chronic conditions. Control of bacterial growth in the oral cavity can help to reduce the risk of heart disease, preserve memory, and reduce the risk of infection and inflammation in other parts of the body. Because people with diabetes are at greater risk of developing serious gum problems, reducing the risk of gingivitis by maintaining good oral health can help control blood sugar. Pregnant women may be more likely to experience gingivitis, and some researchers suggest a relationship between gum disease and the delivery of premature, low birth weight babies in pregnant women.

Bacteria are the main pathogen in periodontal disease. More than 500 bacterial strains can be found in the plaque (Kroes et al., Proc. Natl. Acad. Sci. USA 96: 14547-52 (1999)). Bacteria have evolved to survive as membranes in the environment of the tooth surface, gingival epithelium, and oral cavity, which contributes to the difficulty in treating periodontitis. Current bactericides and antibiotics currently used to treat these infections do not kill all the organisms in question. The use of ineffective preparations for certain bacterial species may result in the proliferation of resistant bacterial species. Moreover, these agents can cause unpleasant side effects such as allergic reactions, inflammation and tooth discoloration.

Bacterial plaque (plaque) is a persistent adherent to oral surfaces, restorations and prostheses. The primary means for controlling the germs in the mouth is through mechanical cleaning (i.e., brushing, dental flossing, etc.). Within the first two days after this cleansing is not taken, the surface of the teeth is mainly colonized by gram-positive facultative cocci, which are mainly Streptococcus spp. The bacterium secretes an extracellular mucus layer that helps immobilize the bacteria on the surface and provides protection against adhered bacteria. Fine colonization is initiated when the surface of the oral cavity is covered with attached bacteria. Fungal membranes are covered by cell division of mainly attached bacteria, not through attachment of new bacteria. The doubling time of the plaque forming bacteria is faster in early development and slower in more mature bacteria.

Agglutination occurs when a bacterial colonizer subsequently attaches to bacteria already attached to the membrane. The result of flocculation is the formation of a complex array of different bacteria linked together. A few days after unblocked plaque formation, the gums become inflamed and swollen. Inflammation can lead to the creation of deep gingival fissures. The membranes expand into these gingival areas and proliferate in this protected environment, leading to the formation of mature gingival plaque membranes. Gingival inflammation does not occur until the germ layer changes from being composed mainly of gram-positive bacteria to containing gram-negative anaerobes. A fine gingival microflora consisting mainly of gram-negative anaerobic bacteria is established in the gingival margin between 3 and 12 weeks. Currently, most of the suspected bacterial species of periodontal disease are anaerobic, gram-negative bacteria.

Bacterial microfluidics protected within the bacteria are typically resistant to antibiotics (systemically administered), preservatives or disinfectants (topically administered), and immune defense. Antibiotic doses that kill free-floating bacteria need to be increased 1,500 times, for example, to kill bacterial bacteria. At these high concentrations, these antimicrobial agents also tend to be toxic to patients (see, for example, Coghlan 1996, New Scientist 2045: 32-6; Elder et al., 1995, Eye 9: 102-9).

Diligent and frequent physical removal of bacterial plaque membranes is the most effective means of removing plaque and regulating it. However, the gingival sulcus plaque in the gum bag can not be reached by brushing, flossing, or mouthwash. Therefore, frequent removal of periodontal tissue from dental hygienist or dentist gingival root surfaces is an essential requirement for the prevention and treatment of periodontitis.

In a particular embodiment, the particulate particulate BT compound is administered to a patient in need thereof, such as a toothpaste, a mouthwash (i.e., mouthwash), an oral gel, a toothpaste powder, a mouthspray (including a spray dispersed by an oral inhaler) But are not limited to, edible films, chewing gums, oral slurries, dentifrice detergents, denture stock solutions, and dental floss. The particulate BT compound may be incorporated into oral care compositions that are of interest to dentists, including, for example, fluoride liquid therapy, cleaning compositions, buffer compositions, oral cleaning solutions, and floss. This aspect may be used to provide the advantages described herein, including broad anti-microbial activity, solubility and bioavailability, anti-bacterial effects, non-toxicity, improved antibiotic efficacy, and other properties as described herein The antimicrobial agent formulated with the oral care composition described in WO 00/58705 is contemplated to be replaced by the particulate BT compound described herein.

The particulate BT compound may also be used to prevent or treat cavities and / or inflammation (i. E., Reduce the incidence or recurrence tendency of each of the cavities and / or irritations) by administering a particulate BT compound to the surface of the teeth. A composition comprising a particulate BT compound, which may be a mucosal adhesive composition applied to the surface of teeth and / or gums or oral mucous membranes, may be applied to the surface to some extent or to deliver a pharmaceutically effective amount of the active ingredient (s) It can be a specific form. The particulate BT compound may also be formulated to release slowly from the composition applied to the teeth. For example, the composition can be a gel (e. G., A hydrogel, a thimer, an aerogel or an organogel) or a liquid. The organogel can include organic solvents, lipoic acid, vegetable oils or mineral oils. Such gel or liquid coating formulations may be applied internally or externally to amalgam or composite or other prosthetic compositions. The sustained release composition may contain an effective amount of a particulate BT compound for 1, 2, 3, 4, 5, 6, or 7 (one week) or for 2, 3, 4, 5, 6, 7 weeks, 3, 4, 5, or 6 months. Such compositions may be prepared by those skilled in the art using any method known in the art.

In certain other embodiments and as described herein, the antimicrobial composition is provided for oral use comprising a particulate BT compound and one or more additional antimicrobial compounds or agents. s and a second antimicrobial agent having an enhanced or synergistic antimicrobial effect as described herein when administered together are particularly useful. As an example, an improved antimicrobial effect can be observed when the particulate BT compound is administered with an antimicrobial agent that chelates iron. In another particular embodiment, the particulate BT compound is formulated with an anti-inflammatory agent, a compound, a small molecule or a macromolecule (e.g., 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 (e.g., thiochlorophenol) may be used, which may exhibit greater capacity than a hydrophobic BT compound that hardly adheres to the teeth and tissues of the oral cavity. BT compounds having net negative charge such that the molar ratio is 1: 2 (bismuth to thiol) may also have good adhesion properties.

Oral hygiene compositions comprising a 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 care 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 cleaning, deodorization and buffering. Baking soda chemically neutralizes the odor rather than shielding or absorbing it. Baking soda may be combined as a mixture of particulate BT compound and powder or may be dissolved or suspended in any of the toothpaste powders, gels, pastes, and liquids described herein. In another embodiment, the particulate BT compound may be combined with other alkali metal bicarbonates or carbonate materials (e.g., potassium bicarbonate or calcium carbonate) to maintain the desired alkaline pH and also have cleaning and deodorizing properties.

Oral hygiene compositions comprising a particulate BT compound may also include one or more of the following ingredients.

Antimicrobial agents such as chlorhexidine; Sanguinarin extract; Metronidazole; Quaternary ammonium compounds (e. G., Cetylpyridinium chloride); Bis-guanides (e.g., chlorhexidine digluconate, hexetidine, octenidine, alexidine); A halogenated bisphenolic compound such as 2,2'methylene bis- (4-chloro-6-bromophenol) or other phenolic antibiotic compound, alkylhydroxybenzoate, a cationic antimicrobial peptide, aminoglyco Coleus forskohlii essential oil; silver or colloid; colloidal silica; colloidal silica; colloidal silica; colloidal silica; colloidal silica; The antioxidants may be selected from the group consisting of antimicrobial agents, tin- or copper-based antimicrobial agents, Manuka oil, oregano, thyme, rosemary or other herbal extracts and grape seed extract.

Anti-inflammatory agents or antioxidants such as ibuprofen, flurbiprofen, aspirin, indomethacin, aloe vera, tourmalic, olive leaf extract, cloves, panthenol, retinol, omega-3 fatty acids, gamma-linolenic acid (GLA), green tea, ginger and grape seed.

Anti- cavities : for example sodium and tin fluoride, amine fluoride, sodium monofluorophosphate, sodium monofluorophosphate, sodium trimetaphosphate, zinc citrate or other zinc preparations, and casein.

Plaque buffers : for example, urea, calcium lactate, calcium glycerophosphate, and strontium polyacrylate.

Vitamins : For example, vitamins A, C, and E.

Plant extracts. Desensitizing agents : for example, potassium citrate, potassium chloride, potassium tartrate, potassium bicarbonate, potassium oxalate, potassium nitrate, and strontium salts.

Anti- stones agents : for example, alkali-metal pyrophosphates, hypophosphite-containing polymers, organic phosphonates and phosphocates, and the like.

Biomolecules : for example, bacteriocins, bacteriophages, antibodies, bacteria and the like.

Flavoring agents : for example, peppermint and spearmint oil, fennel, cinnamon and the like.

Protein substances : for example, collagen.

Preservative .

Milky matter . coloring agent. pH-adjusting agent. Sweetener .

Pharmaceutically acceptable carriers : for example, starch, sucrose, water or water / alcohol systems.

Surfactants : for example, anionic, nonionic, cationic and zwitterionic or amphoteric surfactants, saponins from plant materials (see, for example, U.S. Patent No. 6,485,711).

Particulate abrasives : for example, silica, alumina, calcium carbonate, calcium phosphate, calcium pyrophosphate, hydroxyapatite, trimetaphosphate, insoluble hexametaphosphate, agglomerated particulate abrasive, chalk, .

Wetting agents : for example, glycerol, sorbitol, propylene glycol, xylitol, lactitol, and the like.

Binders and thickening agents may be, for example, example sodium carboxymethylcellulose, hydroxyethyl cellulose (Natrosol ^®), xanthan gum, gum arabic, synthetic polymers (e.g., polyacrylates and carboxyvinyl polymers, such as, Carbopol ^® ).

Polymeric compounds that enhance the delivery of active ingredients such as antimicrobial agents.

Buffers and salts to buffer the pH and ionic strength of the mouth guard composition.

Bleaching agents : for example, peroxy compounds (e. G., Potassium peroxydiphosphate).

Boiling system: for example sodium bicarbonate / citric acid system.

Color change system .

In a particular embodiment, the abrasive is silica or finely divided natural chalk.

Oral hygiene compositions comprising particulate BT compounds that are formulated for use as toothpastes may also contain a wetting agent (e.g., glycerol or sorbitol), a surface active agent, a binder, and / or a flavoring agent. The toothpastes may also contain sweeteners, whitening agents, preservatives and antimicrobial agents. The pH of toothpastes and other oral compositions is typically between pH 5.5 and 8.5. In certain embodiments, the oral care composition comprising toothpaste has a pH between 7 and 7.5, between 7.5 and 8, between 8 and 8.5, or between 8.5 and 9, which can improve the antimicrobial activity of the particulate BT compound. The dentifrice compositions described herein may be used in combination with one or more of chalk, calcium dihydrogen phosphate, sorbitol, water, hydrated aluminum oxide, precipitated silica, sodium lauryl sulfate, sodium carboxymethylcellulose, flavor, sorbitan monooleate, , Sodium pyrosene, methylparaben, and propylparaben. One or more coloring agents, for example FD & C Blue, may be used if desired. Other suitable ingredients that may be included in the dentifrice composition are described in the art, for example, in U.S. Patent No. 5,560,517.

In another particular embodiment, the oral care composition is a mouthwash and comprises a particulate BT compound, an alkaline buffer (e.g., potassium bicarbonate), an alcohol, a sweetener component, and a flavor system. The flavor system may also have one or more of the following ingredients: flavors, wetting agents, surface active agents, sweeteners and colorants (see, for example, U.S. Patent No. 6,579,513). The surface-roughening agents described herein and known in the art for use in oral care compositions may be anionic, nonionic or amphoteric.

In another embodiment, the particulate BT-containing oral care composition is useful for incorporation into toothpastes, tooth gels, and mouthwashes for the treatment of serious infections, and may be combined with additional active ingredients such as taurolidine and taurultam as described in the art (See GB Patent No. 1557163, U.S. Patent No. 6,488,912). As described herein, particulate BT may also be combined with one or more additional antimicrobial agents wherein the combination has additional or synergistic effects when combined with particulate BT.

In still other particular embodiments, the oral care composition described herein may further comprise at least one anti-bacterial agent for regulating bacterial growth, destroying the bacterial membrane, or reducing the amount of the bacterial membrane. As will be understood in the art, interspecies quorum sensing is associated with membrane formation. Certain agents that increase the LuxS-dependent pathway or interspecies quorum sensing signal (see, e.g., U.S. Patent No. 7,427,408) contribute to regulating the development and / or proliferation of the bacterium. Exemplary formulations include, by way of example, the N- (3-oxododecanoyl) -L-homoserine lactone (OdDHL) blocking compound and the N-butyryl-L-homoserine lactone (BHL) analogue either alone or in combination See, for example, U.S. Patent No. 6455031). Oral hygiene compositions comprising a particulate BT compound and at least one anti-bacterial agent can be delivered locally for the destruction and inhibition of bacterial membranes and for the treatment of periodontal disease (see, e.g., U.S. Patent No. 6,726,898).

The oral care composition described herein may contain an amount of the particulate BT compound sufficient to exhibit substantial antimicrobial action during the time required for normal brushing, oral cleaning or dentistry. As described herein, the particulate BT compound may be retained on the oral surface (e.g., teeth, amalgam, complex, mucosa, gums). The particulate BT compounds retained in the teeth and gums after brushing, washing, and completion of dental floss can, for example, continue to provide extended anti-bacterial and anti-inflammatory effects.

In another embodiment, the particulate BT compound is released slowly from the mucosa-adherent polymer or other agent that contributes to retention of the particulate BT compound at the mucosal and tooth surfaces. The particulate BT compound may be used in combination with a pharmaceutically acceptable carrier which may be used for delivery to the mucosal surface or delivery to the systemic circulation for the prevention and / or treatment of erosive, inflammatory and / or mucosal erosive disorders and / , May be added to a viscous, viscous, aqueous composition (see, for example, U.S. Patent No. 7,547,433).

In another embodiment, the oral hygiene composition comprising the particulate BT compound further comprises an olive oil that can enhance plaque removal. The use of olive oil in products intended for oral hygiene, such as toothpaste, mouthwash, spray, oral inhaler or chewing gum, eliminates or reduces (reduces) bacterial plaque and / or eliminates (For example, tooth decay, periodontal disease) and reduction in halitosis outbreaks (see, for example, U.S. Patent No. 7,074,391).

In another embodiment, the oral care composition comprising a particulate BT compound may further comprise a mucoadhesive agent formulation for topical application in the mouth. The oral care composition may further comprise an aqueous slurry useful for cleaning the tongue and larynx (see, e.g., U.S. Patent No. 6,861,049). In still other embodiments, the oral hygiene composition comprising a particulate BT compound may also comprise at least one mint used to prevent (i. E., Reduce the occurrence tendency) or &lt; / RTI & (mint). &lt; / RTI &gt; One such mint, called CaviStat ^® (from Orthotactic Products, Incorporation), Roslin Heights, NY, is a combination of arginine and calcium, which helps promote the attachment of calcium to enamel surfaces and neutralize acidic pH . Thus, incorporation of mint in an oral care composition comprising a particulate BT compound can improve adhesion of the particulate BT compound to the oral surface and increase the pH.

Compositions comprising particulate bismuth- thiols formulated for orthopedic applications .

In particular aspects, there is provided a method of preventing and / or treating microbial infections and inflammation resulting from orthopedic surgery (e.g., orthopedic surgery, orthopedic therapies, arthroplasty (including two-stage arthroplasty) There is provided a method of using a composition comprising a particulate BT compound. Compositions comprising particulate BT compounds as described herein can be used to prevent and / or treat microbial infections of the skeleton and supporting structures (i.e., bone, joints, muscles, ligaments, tendons), such as osteomyelitis Or inhibiting, or decreasing the tendency of, or recurrence thereof). Compositions described herein, including particulate BT compounds, can also be used to prevent and / or modulate (i.e., slow down, slow down, or inhibit) bacterial growth, destroy bacterial membranes, It may be useful to reduce the amount of the bacterium present on the surface of the tooth.

Compositions described herein for orthopedic applications, including particulate BT compounds, 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 having enhanced or synergistic antimicrobial effects as described herein when administered together. As a further example, an improved antimicrobial effect can be observed when the particulate BT compound is administered with an antimicrobial agent that chelates iron. In another particular embodiment, the particulate BT compound is formulated with an anti-inflammatory agent, a compound, a small molecule, or a macromolecule (e.g., a peptide or polypeptide).

Compositions comprising a particulate BT compound may be formulated with at least one other antimicrobial agent (i.e., an antimicrobial agent such as a second, third, fourth, etc.) having an enhanced or synergistic antimicrobial effect (i. &Lt; / RTI &gt; As an example, the improved antimicrobial effect can be observed when the particulate BT compound is administered with an antimicrobial agent that chelates iron. In a particular embodiment, the composition comprising the particulate BT compound can be combined with at least one other antimicrobial and / or anti-inflammatory agent selected from:

Antimicrobial agents such as chlorhexidine; Sanguinarin extract; Metronidazole; Quaternary ammonium compounds (e. G., Cetylpyridinium chloride); Bis-guanides (e. G., Chlorhexidine digluconate, hexetidine, octenidine, alexidine); A halogenated bisphenolic compound such as 2,2'methylene bis- (4-chloro-6-bromophenol) or other phenolic antibiotic compound, alkylhydroxybenzoate, a cationic antimicrobial peptide, aminoglyco Or antibiotics known in the art; cholecystorhcic acid essential oils; silver or colloidal silver microbes; tin- or copper-based antimicrobials; Antimicrobial agents, manuka oil, oregano, thyme, rosemary, or other herbal extracts and grape seed extract.

Anti-inflammatory agents or antioxidants may be, for example, ibuprofen, FLOURE beeping pens, aspirin, indomethacin, aloe vera, Tour numeric, olive leaf extract, cloves, panthenol, retinol, omega-3 fatty acid, gamma-linolenic acid (GLA) , Green tea, ginger, grape seed, etc. In a particular embodiment, the composition comprising a particulate BT compound may also comprise a compound selected from the group consisting of clindamycin, vancomycin, dapotomycin, cephazoline, gentamicin, tobramycin, metronidazole, cepharchlor, ciprofloxacin, or quaternary ammonium compounds Benzalkonium chloride, cetylpyridinium chloride), anti-microbial zeolites, alkali metal hydroxides or alkaline earth metal oxides. The composition may optionally comprise one or more of the pharmaceutically acceptable carriers (i. E. Excipients), surface active agents, buffers, diluents, and salts, and bleaching agents described herein. Thus, these and certain related embodiments described herein may include one or more of the particulate BTs, and may optionally further include antibiotics such as synergistic or enhanced antibiotics as described herein, Of these products and processes.

Biological, biomedical and other uses for particulate BT .

Certain other aspects relate to the use of BT as a BT in which individual BT or bismuth moieties are substituted with different Group V metals such as antimony (Sb) or arsenic (As) within the nutritional formula taken orally, and / In addition, the use of the particulate BT described herein, regardless of whether BT is combined with one or more antibiotics exhibiting synergistic or improved antimicrobial activity, is contemplated.

In accordance with a non-limiting theory, it is also possible to combine these formulations with other ingredients such as hydrocarbons, fatty acids, oils, plant nutrients, tea, herbal or herbal extracts containing vitamins, minerals, amino acids, carbohydrates, and / or other nutrients or foodstuffs (E.g., in a statistically significant manner for an appropriate control) of BT and optionally antibiotic and / or additional nutrient (s) to the host alimentary canal, in certain embodiments, In a manner that promotes, it blocks or delays the uptake of nutrients by microbial populations within the gastrointestinal tract. In certain other embodiments, hydrocarbons, fatty acids, oils, plant nutrients, tea, herbal or herbal extracts and / or other nutrients, including special vitamins, minerals, amino acids, carbohydrates, contained in oral microparticle BT (or AsT or SbT) (E.g., in a statistically significant manner for an appropriate control) of the BT and optionally the antibiotic and / or additional nutrient (s) to the host alimentary tract by altering the nutrient content of the host.

For example, administration of a particulate BT formulation that inhibits intestinal absorption of BT compounds in the presence of pathological gastrointestinal (GI) tract infections would make them bioavailable in the GI tract, thereby causing the antimicrobial effect to infectious pathogens Lt; / RTI &gt; Those skilled in the art will appreciate that a variety of vitamins, minerals, amino acids, carbohydrates, hydrocarbons, fatty acids, oils, plant nutrients, tea, herbal or herbal extracts and / or other nutrients or foodstuffs that promote or prevent GI tract absorption of nutrients (Or AsT or SbT) for the formulation to increase or decrease the presence of GI tracts of one or more components (e. G., Particulate BT compounds, antibiotics, or one or more specialty nutrients) You can do it.

Certain other aspects provided herein include, for example, compositions for oral delivery to reduce fecal or digestive gas odors in patients who have undergone colostomy, and methods for reducing side effects, foot or other body odors associated with the presence of topical microorganisms Lt; RTI ID = 0.0 &gt; BT &lt; / RTI &gt; compounds presently present in other compositions for topical delivery. A number of skin and GI tract microbial populations, including planktonic and bacterial bacterium, may be used in combination with a lower concentration of the particulate BT compound described herein, including such BT compounds, when present with an enhancing or synergistic antibiotic as described herein .

Thus, certain embodiments relate to oral delivery (e.g., by oral administration) to lower the population of GI-persistent or skin-persistent bacteria in a manner that reduces or alleviates unwanted odor problems (e.g., in a statistically significant manner for an appropriate control) And topically delivered particulate BT formulations. Oral and topical pharmaceutical formulations have the advantage that these and related aspects relate to the current particulate formulations of BT, such as miscible bioavailability and solubility characteristics and low toxicity; Are described below to affect the selection of the antimicrobial composition described herein also and to provide other factors, such as those found in U.S. Patent No. 6,582,719.

An exemplary BT compound, BisEDT, was found to be applied to the armpit site of a human test subject (50 μL of a 1 mg / mL solution in DMSO) and to neutralize the body odor for a few days. A mixture of BisEDT as a talc powder applied to the feet of a human test subject substantially reduced foot odor. Laboratory mice fed 1 mg / kg BisEDT twice a day orally 5 days showed a 90% reduction in the number of stool flocks. A related aspect is also a process for the preparation of any odor-emitting thiol-containing solution comprising a particulate BT formulation as described herein which is prepared with an excess of bismuth and which can be added to the thiol-containing solution as an odor quenching agent. (For example, fish oil such as salmon oil). The resulting mixture retains the antimicrobial properties of the particulate BT. Other contemplated applications include the use of other biological source oils or butter, such as hemp oil, tea tree oil, shea butter, flaxseed oil, solvents such as fish oil, and independent or synergistic anti-inflammatory and / Or pain-relieving and / or other beneficial physiological effects, and thus includes such oils in certain embodiments.

Pharmaceutical composition and administration

Certain embodiments are also directed to pharmaceutical compositions containing the particulate BT compounds described herein; In certain such embodiments, the pharmaceutical composition may further comprise one or more antibiotics, such as antibiotics, wherein the BT compound exhibits synergistic or enhanced effects as described herein. In one embodiment, when administered to an animal, preferably a mammal, most preferably a human patient, one or more of these microparticles in a therapeutically effective amount as described herein and in a pharmaceutically acceptable carrier, excipient or diluent A composition comprising a BT compound is provided.

Administration of the particulate BT compound, or a pharmaceutically acceptable salt thereof, in pure form or as a suitable pharmaceutical composition may be carried out through any of the accepted models of administration of the agent to provide similar uses. The pharmaceutical composition may be prepared by combining the particulate BT compound with a suitable pharmaceutically acceptable carrier, diluent or excipient and may be in the form of a solid, semi-solid, liquid or gaseous form such as tablets, capsules, powders, Granules, ointments, liquid preparations, suppositories, injections, inhalants, gels, microspheres and aerosol preparations. Representative routes of administration of such pharmaceutical compositions include, but are not limited to, oral, topical, transdermal, inhalation, parenteral, sublingual, rectal, intrathecal, and intranasal. As used herein, the term parenteral includes subcutaneous, intravenous, intramuscular, intrasternal injection or infusion techniques. The pharmaceutical composition is formulated such that the active ingredient contained therein is bioavailable upon administration to the patient. The composition to be administered to a subject or patient takes the form of one or more dosage units, for example, the tablet may be a single dose unit, and the container of the aerosol-type compound may hold a plurality of dosage units. Actual methods of preparing such dosage forms are known or will be familiar to those skilled in the art (see, for example, The Science and Practice of Pharmacy, 20th Edition, Philadelphia College of Pharmacy and Science, 2000). The composition to be administered will in any event contain a therapeutically effective amount of a compound, or a pharmaceutically acceptable salt thereof, for the treatment of a desired disease or condition according to the teachings herein. Pharmaceutical compositions useful herein also include any suitable diluent or excipient that can be administered without undue toxicity, including any pharmaceutical agent that itself does not induce the production of antibodies that are deleterious to the individual receiving the composition , And a pharmaceutically acceptable carrier. Pharmaceutically acceptable carriers include, but are not limited to, water, saline, glycerol and liquids such as ethanol and the like. A complete discussion of pharmaceutically acceptable carriers, diluents and other excipients is provided in REMINGTON'S PHARMACEUTICAL SCIENCES (Mack Pub. Co., N. J. current edition).

The pharmaceutical composition may be in the form of a solid or a liquid. In one aspect, the carrier (s) is granulated so that the composition is in the form of, for example, tablets or powders. The carrier (s) may be, for example, a liquid which is useful for inhalation administration, for example a composition that is an oral syrup, injectable liquid or aerosol.

When intended for oral administration, the pharmaceutical composition is preferably in a solid or liquid form, wherein the semi-solid, semi-liquid, suspension and gel forms are included in the forms contemplated herein as solids or liquids.

As a solid composition for oral administration, the pharmaceutical composition may be formulated into powders, granules, compressed tablets, pills, capsules, chewing gums, wafers or the like. Such solid compositions will 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, gum tragacanth or gelatin; Excipients such as starch, lactose or dextrin, disintegrants such as alginic acid, sodium alginate, Primogel, corn starch and the like; Lubricants such as magnesium stearate or Sterotex; Bow topics such as colloidal silicon dioxide; Sweeteners such as honey, sugar or saccharin; Flavors such as peppermint, methyl salicylate or orange flavor; And a colorant.

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

The pharmaceutical composition may be in the form of a liquid, for example, in the form of elixir, syrup, solution, emulsion or suspension. The liquid may be, for example, for oral administration or for delivery by injection. When intended for oral administration, the preferred compositions comprise, in addition to the composition of the present invention, at least one of a sweetener, an antiseptic, a dye / colorant and a flavor enhancer. In the composition intended to be administered by injection, one or more of a surface active agent, an antiseptic, a wetting agent, a dispersing agent, a suspending agent, a buffering agent, a stabilizing agent and an isotonic agent may be included.

Liquid pharmaceutical compositions may contain one or more of the following adjuvants, irrespective of whether they are liquid solutions, suspensions or other similar forms: sterile diluents such as water for injection, saline solutions, preferably physiological saline, Ringer's solution, Sodium chloride, a fixed oil such as synthetic mono- or diglycerides which may be provided as a solvent or suspending medium, polyethylene glycols, glycerin, propylene glycol or other solvents; Antibacterial 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 isotonicity controlling agents such as sodium chloride or dextrose. Parenteral formulations may be enclosed in ampoules, disposable syringes or multi-dose vials made of glass or plastic. Physiological saline is the preferred excipient. The injectable pharmaceutical composition is preferably sterile.

Liquid pharmaceutical compositions intended for parenteral or oral administration contain an amount of a particulate BT compound that will result in a suitable dosage. Typically, the amount is at least 0.01% of the particulate BT compound in the composition. When intended for oral administration, the amount may vary from 0.1% to about 70% by weight of the composition. Preferred oral pharmaceutical compositions contain from about 4% to about 50% by weight of the BT compound. Preferred pharmaceutical compositions and formulations according to the present invention are prepared so that the parenteral dosage unit contains from 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 suitably comprise a liquid, a suspending, an ointment or a gel base. For example, the substrate may include one or more of the following: petroleum, lanolin, polyethylene glycol, beeswax, mineral oil, shea butter, tea tree oil, flaxseed oil, Other plant oils or vegetable oils, including those known to have anti-pain or other beneficial effects, salmon oil or other fish oils, including water and alcohol, including those known to have anti-inflammatory and / or anti- , And emulsifiers and stabilizers. Thickening agents may be present in topical pharmaceutical compositions. When intended for transdermal administration, the composition may comprise a transdermal patch or an iontophoresis device. The topical formulation may contain a concentration of the particulate BT compound of from about 0.1 to about 10% w / v (weight per unit dose).

The pharmaceutical composition may be intended for rectal administration in the form of suppositories, for example, which will melt in the rectum and release the drug. Compositions for rectal administration may contain a retentate substrate as a suitable non-irritating excipient. Such substrates include, but are not limited to, lanolin, cocoa butter, and polyethylene glycols.

Pharmaceutical compositions may include various materials that modify the physical form of a solid or liquid dosage unit. For example, the composition may comprise a material that forms a coating shell around the active ingredient. The material forming the coated shell is typically inert and may be selected from, for example, sugars, shellac, and other enteric coating agents. Alternatively, the active ingredient may be encapsulated in a gelatin capsule.

A pharmaceutical composition in solid or liquid form may comprise an agent that assists in the delivery of the compound by binding to the particulate BT compound. Suitable formulations which may serve at such doses include monoclonal or polyclonal antibodies, proteins or liposomes. However, certain contemplated embodiments exclude the incorporation of liposomes into pharmaceutical compositions.

The pharmaceutical composition may be in dosage units that can be administered as an aerosol. The term aerosol is used to denote a variety of system-wide systems of pressurized packages from those of a colloidal nature. Delivery may be by liquefied or squeezed gas or by a suitable pump system for dispersing the active ingredient. Aerosol formulations of particulate BT compounds can be delivered to a single phase, two-phase or three-phase system to deliver the active ingredient (s). The delivery of the aerosol includes the necessary vessels, activators, valves, subcontainers, etc., which together can form a kit. Skilled artisans will be able to determine preferred aerosols without undue experimentation.

Pharmaceutical compositions may be prepared by methods well known in the art of pharmacy. For example, pharmaceutical compositions intended for administration by injection may be prepared by combining the compounds of the present invention with sterile, distilled water to form a liquid. Surfactants may be added to promote the formation of homogeneous liquids or suspensions. Surfactants are compounds that promote dissolution or homogeneous suspension of a compound in an aqueous delivery system by non-covalent interaction with the compounds of the present invention.

The particulate BT compound, or a pharmaceutically acceptable salt thereof, as described herein may be prepared by the action of the specific compound employed; Metabolic stability and working length of the compound; Age, weight, general health, sex and diet of the patient; Mode and time of administration; Discharge rate; Drug combination; The severity of the particular disease or condition; And the subject being treated. The amount of &lt; RTI ID = 0.0 &gt; Generally, a therapeutically effective daily dose is from about 0.001 mg / kg (i.e., 0.07 mg) to about 100 mg / kg (i.e., 7.0 g) for a 70 kg mammal; Preferably, the therapeutically effective dose is about 0.01 mg / kg (i.e., 7 mg) to about 50 mg / kg (i.e., 3.5 g) (for a 70 kg mammal); More preferably, the therapeutic dose is about 1 mg / kg (i.e., 70 mg) to about 25 mg / kg (i.e., 1.75 g) (for a 70 kg mammal).

The range of effective doses provided herein is not intended to be limited by the preferred dosage range. However, the most preferred dosages will be adjusted according to the individual subject, as can be appreciated and measured by those skilled in the relevant art (see, for example, Berkow et al., Eds., The Merck Manual, 16th edition, Merck and Co.). , Rahway, NJ, 1992; Goodman et al., Eds., Goodman and Oilman ' s Pharmacological Basis of Therapeutics, 10th 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. (1987), Ebadi, Pharmacology, Little, Brown and Co., Boston, (1985); Osolci al., Eds., Remington ' s Pharmaceutical Sciences, 18th edition, Mack Publishing Co., Easton, PA (1990); Katzung, Basic and Clinical Pharmacology, Appleton and Lange, Norwalk, CT (1992)].

The total dose required for each treatment may be administered in multiple doses or single doses, if desired, over one day. In general, treatment is initiated with a smaller dose, less than the optimal dose of the compound. Thereafter, the capacity is increased by a small amount until the optimum effect is achieved under the circumstances. The diagnostic pharmaceutical compound or composition may be administered alone or in conjunction with other agents directed at other diagnostic agents and / or pathologies, or in other conditions of pathology. The recipient of the administration of the particulate BT compound and / or composition may be any vertebrate animal, such as a mammal. Among mammals, preferred recipients include, but are not limited to, primates (including humans, apes and monkeys), supplements (including horses, goats, cows, sheep, pigs), rodents (including mice, rats, rabbits and hamsters), and carnivores ). Of the birds, the preferred recipients are turkeys, chickens and many other identical throats. The most desirable prisoner is a person.

For topical application, it is preferred to administer an effective amount of a particulate BT-containing pharmaceutical composition to a target site, such as a skin surface, mucosa, or the like. The amount will generally range from about 0.0001 mg to about 1 g per application BT compound, depending on the site to be treated, the severity of the symptoms, and the nature of the topical vehicle used, regardless of whether the use is diagnostic, prophylactic or therapeutic. A preferred topical agent is an ointment, wherein from about 0.001 to about 50 mg of the active ingredient is used per cc of ointment substrate. The pharmaceutical composition may be formulated as a transdermal composition or transdermal delivery device (" patch "). Such compositions include, for example, backings, active ingredient reservoirs, regulating membranes, liners and contact adhesives. These transdermal patches may be used to provide continuous beating, or, if desired, to deliver a compound of the present invention.

The particulate BT composition may be formulated to provide rapid, sustained or delayed release of the active ingredient after administration to the patient by using procedures known in the art. Modulated 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 are provided in U.S. Patent Nos. 3,845,770 and 4,326,525 and PJ Kuzma et al, Regional Anesthesia 22 (6): 543-551 (1997), all of which are incorporated herein by reference &Lt; / RTI &gt;

The particulate BT compositions may also be delivered via intranasal drug delivery systems for topical, systemic and nose-to-brain medical therapies. Provides a controlled particle distribution (CPD) ^TM technology, traditional nasal spray bottles, inhalers or atomizer (nebulizer) is effective local and systemic delivery of drugs by targeting the olfactory hierarchical region and paranasal sinuses (paranasal sinuses).

The invention also relates to a vaginal shell or core drug delivery device suitable for administration to a human or animal woman in certain embodiments. The device can be composed of active pharmaceutical ingredients in a polymer matrix surrounded by a coating and can be prepared by mixing the compound with a compound of formula &lt; RTI ID = 0.0 &gt; . &Lt; / RTI &gt;

Currently, ocular delivery methods include local administration (eye drop), subconjunctival injection, periocular injection, intravitreal injection, surgical implantation and ion transfer (using a small current to transfer and transport ionized drug into the body tissue) . One skilled in the art can combine the most suitable excipients with safe and effective intra-ocular administration compounds.

The most appropriate route will depend on the nature and severity of the condition being treated. Those of skill in the art will also be familiar with the administration methods (oral, intravenous, inhalation, subcutaneous, rectal, etc.), dosage forms, suitable pharmaceutical excipients, and others related to the delivery of the compound to a subject in need thereof.

The presently described compositions and methods are also useful for the treatment of existing wounds, including those described herein, for the prevention of chronic wounds, for the treatment of MRSA skin infections, as described, for example, And other related prescription topical agents familiar to the skilled person in terms of the present disclosure.

Non-limiting examples of bacteria in which the compositions and methods described herein may find advantageous use according to certain embodiments as described herein include Staphylococcus aureus (S. aureus), MRSA (methicillin - resistant, S. aureus), Staphylococcus epidermidis, MRSE (methicillin-resistant S. epidermidis), M. tuberculosis, Maocobacillium absium, Pseudomonas aeruginosa Luginosa, drug-resistant blood. Escherichia coli, enterotoxin production. Coli, intestinal hemorrhagic. E. coli, Enterococcus faecalis, Enterobacter cloacae, Salmonella typhimurium, Enterococcus faecalis, Enterococcus faecalis, Enterococcus faecalis, Enterococcus faecalis, Enterococcus faecalis, , V. vulnificus enterococcus (VRE), brucolierea sephacia complex, francicella tularensis, bacillus anthracis, yersinia, bacteriocin-resistant Enterococcus, Festis, pseudomonas aeruginosa, vancomycin-sensitive enterococci [for example, this. E. faecalis, Lee. E. faecium], methicillin-sensitive and methicillin-resistant Staphylococci [e.g. Aureus, S. S. epidermidis] and Acanthoptera baumannii, Staphylococcus haemolyticus, Staphylococcus hominis, Enterococcus faecium, Enterococcus faecium, Streptococcus pyogenes, Streptococcus agalactiae, Bacillus anthracis, Klebsiella pneumonia, Proteus spp.), Streptococcus pyogenes, Streptococcus agalactiae, Bacillus anthracis, Klebsiella pneumonia, , such as Staphylococcus aureus, mirabilis, Proteus vulgaris, Yersinia enterocolytica, Stenotrophomonas maltophilia, Streptococcus pneumonia, Penicillin-resistant Streptococcus Burkholderia cepacia, Bukholderia multivorans, Mycobacterium squirrel, Tees (Mycobacterium smegmatis) and the. And E. cloacae.

The practice of certain embodiments of the present invention will employ conventional methods of microbiology, molecular biology, biochemistry, cell biology, virology and immunology techniques within the skill of the art, and reference to some of them, unless otherwise specifically indicated, For purposes of illustration. Such techniques are fully described in the literature (see, for example, Sambrook, et al. Molecular Cloning: A Laboratory Manual (2nd Edition, 1989); Maniatis et al. Molecular Cloning: A Laboratory Manual (1982); DAW Cloning: A Practical Approach, vol. I & II (D. Glover, ed.); Oligonucleotide Synthesis (N. Gait, ed., 1984); Nucleic Acid Hybridization (B. Hames & S. Higgins, eds., 1985); Transcription and Translation (B. Hames & S. Higgins, 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, variations such as "including" and "including" and "including" And should be considered in its broader meaning.

Throughout this specification, "an aspect" or "aspect" or "aspect" means that the particular features, structures, and characteristics described in connection with the aspects are included in at least one aspect of the present invention. Thus, the appearances of the phrase " in one embodiment " or " in an aspect, " which may vary widely throughout the specification, are not necessarily all referring to the same aspect. In addition, a particular feature, structure, or characteristic may be combined in any suitable manner in one or more aspects.

Certain embodiments include methods, compositions, and methods for treating acute or chronic scarring or wound scarring in a subject, or modifying one or more cytotoxic scarring activity in a cell or a plurality of cells, To a kit. A cell generally represents a single cell, but many cells represent more than one cell. The cell may comprise a tissue, organ or whole organism. In addition, the cells or cells may be located in vivo, in vitro or in vivo. It is an ordinary course of the skilled person in the art to maintain cell, tissue and organ culture, conditions and medium for this can be easily estimated (see, for example, Freshney, Culture of Animal Cells: A Manual of Basic Technique, Wiley-Liss 5th Ed. (2005); Davis, Basic Cell Culture, Oxford University Press 2nd Ed. (2002).

As described herein, certain embodiments relate to a BT compound as described herein for use in a method of treating an acute or chronic wound or wound fungal membrane, such as one provided in the form of a plurality of substantially single-dispersible microparticles ), And optionally antibiotic compounds such as those described herein for use in such methods in certain additional embodiments, such as BT compounds such as BisEDT or BisBAL, or other compounds shown herein in Table 1, Domenico et al. Chemother. 45: 1421) and / or US RE 37373, US 6,248,371, US 6,086,921, and US &lt; RTI ID = Comprising administering to a subject a therapeutically effective amount of a composition that further comprises any other BT agent, such as those described in US Patent No. 6,380,248, and / or any other BT agent as prepared according to the methods described herein, Or a method for treating a wound fungal membrane. Certain other embodiments relate to a method comprising contacting any native surface with a composition comprising at least one of the particulate BT compounds described herein, wherein such contacting step includes direct application, coating, dipping, irrigating, Spraying, painting, or otherwise contacting the BT composition with the natural surface.

Administration to a subject, such as a human or other mammalian subject, can be by any means known in the art, for example, locally (such as in the skin or glandular tissue or in the respiratory tract and / Intraperitoneally, orally, parenterally, intravenously, intraarterially, percutaneously, sublingually, subcutaneously, intramuscularly, intraperitoneally, intraperitoneally, intramuscularly, intraperitoneally, intramuscularly, intramuscularly, intraperitoneally, Intravenously, subcutaneously, intrathecally, intrathecally, or intramodularly. The term &quot; intraperitoneal &quot;

In a preferred embodiment, administration can be performed topically, where topical pharmaceutical excipients or carriers are described herein and are known in the art.

As indicated above, certain aspects of the invention described herein relate to topical formulations of the BT compounds described (e. G., BisEDT and / or BisBAL), which formulations are described herein in certain additional embodiments (Or other rinsosamide antibiotic), diphotomycin (Cubicin ^® ), doxycycline, caffeicin, ciprofloxacin, ciprofloxacin, ciprofloxacin, Zygomycin, imiphecine, levofloxacin, zyvox ( ^R ), minocycline, napsilin, paromomycin, rifampin, sulfamethoxazole, tobramycin and vancomycin; Or a carbapenem antibiotic, a cephalosporin antibiotic, a fluoroquinolone antibiotic, a glycopeptide antibiotic, a rincozyme antibiotic, a penicillinase-resistant penicillin antibiotic, and / or an aminopenicillin antibiotic, and / or an aminoglycoside antibiotic, , Amikacin, arbekacin, gentamicin, ganamycin, neomycin, nethymicin, paromomycin, rhodophorptomycin, streptomycin, tobramycin or apramycin, and / or lipopeptide antibiotics, For example, it may include Dapotomycin ( ^R ), or oxazolidinone antibiotics such as Zyvox ( ^R ). These and related formulations, when topically administered to an animal, preferably a mammal, and most preferably a human, comprise a BT compound (s) (and optionally one or more compounds) in a pharmaceutically acceptable carrier, excipient or diluent as described herein And in a particularly preferred embodiment, humans are capable of producing a bacterial-related (e.g., a bacterial, which may be present but capable of promoting bacterial growth, Have a wound that contains a bacterial infection or contains a bacterial infection such as a bacterium or the presence of other bacteria, or an acute or chronic wound.

Local administration of the BT compounds, or pharmaceutically acceptable salts thereof, as described herein in pure form or as a suitable pharmaceutical composition can be accomplished by any of the accepted modes of topical administration of the agents to provide similar uses. Topical application or administration of the composition may, in a preferred embodiment, include administering the composition (e.g., topical formulation) to the skin and / or other epithelial tissue surfaces (e.g., the respiratory tract, gastrointestinal tract and / Which may be present in one or more localized or widely distributed skin and / or other epithelial surface areas and is generally surrounded by a complete stratum corneum or epidermis, but need not be limited thereto Refers to contact with acute or chronic wound sites; For example, certain embodiments may not be such that contact of topical formulations can occur only in the stratum corneum or epithelium, by administering topical formulations as described herein to the skin of a damaged, peeled, or injured skin, May also occur in skin granule cells, ocular cells and / or basal cell layers, and / or in the skin or underlying tissue, as they may be associated with certain types of wound healing or wound healing or other skin tissue remodeling, .

Thus, such dermal tissue repair can be used in certain preferred embodiments, for example, in preventing or alleviating acute chronic scarring or wound scarring, or as another example, in preventing or alleviating skin scarring dehiscence, Or skin wound healing, as it may be desirable to improve, accelerate, or improve skin wound healing when chronic wounds and / or skin wounds may be present. Certain other embodiments that contemplate topical administration to the epithelial tissue surface present in the respiratory tract may likewise include administering topical agents as provided herein to the respiratory tract (e. G., Airway, coin tract and larynx pathway, For delivery to one or more epithelial tissue surfaces and / or other epithelial surfaces present in the stomach (e.g., bronchial, bronchial, alveolar, etc.) and / or gastrointestinal (e.g., ball, esophagus, stomach, Administration of topical formulations by any suitable route known in the art.

According to a particular contemplated embodiment, topical administration may include direct application into an open wound. For example, open fractures or other open wounds may include ruptures in the skin that may expose additional underlying tissue to the external environment in a manner that makes them susceptible to microbial infection. This situation is common in certain types of acute traumatic injury, including, for example, Type III (severe) open fractures. According to these and related embodiments, topical administration may be achieved by administering the BT compositions described herein to such damaged skin and / or other epithelial surfaces and / or other tissues such as circulatory tissues such as muscles, ligaments, tendons, Direct contact with the associated nerve tissue, and any other organ that may be exposed to such an open wound. Examples of other tissues that can be exposed and thus to which such direct contact can be considered include kidney, bladder, liver, pancreas and any other tissue or organ that may be exposed to an opportunistic infection in connection with the open wound .

Topical formulations (e. G., Pharmaceutical compositions) may be prepared using the BT compounds described (e. G., US RE 37,793, US 6,248,371, US 6,086,921, and / or US 6,380,248 Including, for example, compounds prepared according to the present disclosure, such as the particulate BT suspensions described and / or described herein, and in certain related embodiments may be combined with one or more of the desired antibiotics (e. G., Amino Glycoside antibiotics), either alone or in combination with a BT compound, may be prepared in combination with suitable pharmaceutically acceptable carriers, diluents or excipients for use in topical formulations, including solid, semi-solid, gel, cream, Colloids, suspensions or liquids or other topically applied forms such as powders, granules, ointments, solutions, cleansers, gels, pastes, plasters, , Bioadhesives, microspheres, suspensions, and aerosol spray formulations.

These and related embodiments of the pharmaceutical compositions can be prepared by mixing the active ingredients contained therein and the BT compound (s) described herein in a particularly preferred embodiment, alone or in combination with one or more of the desired antibiotics Such as carbapenem antibiotics, cephalosporin antibiotics, fluoroquinolone antibiotics, glycopeptide antibiotics, rinkosamid antibiotics, penicillinase-resistant penicillin antibiotics, and aminopenicillin antibiotics, such as amikacin or rifamycin (S) and / or antibiotic composition (s), and optionally a pharmaceutically acceptable carrier, such as a mammal, including a human, and a particular preferred &lt; RTI ID = In an embodiment, the composition has an acute or chronic wound, or an acute or chronic wound, To ensure bioavailability when topically applied to the surrounding skin of a person at increased risk of having a wound (e.g., diabetic and / or diabetic individual). While certain embodiments described herein contemplate topical administration of BT compounds and antibiotics, including administration that can be administered simultaneously or sequentially and in any order, the present invention is not intended to be so limited, and in other embodiments administration of antibiotics Expression of a clear route of administration of the BT compound to the pathway is expressly considered. Thus, the antibiotic may be administered orally, intravenously, or by any other route of administration as described herein, but the BT compound may be administered by a route that is separate from the route used for antibiotics. By way of a non-limiting, illustrative example, the BT compound may be administered topically as provided herein, while the antibiotic may be administered by a clear route, such as by oral, intravenous, transdermal, subcutaneous, intramuscular, and / (And in any order) by a &lt; / RTI &gt;

The topical formulations described herein deliver a therapeutically effective amount of a therapeutically effective amount of an antiseptic or wound-healing agent (s) (and optionally an antibiotic (s)) to a skin cell, such as a skin fibroblast, at a wound site . Preferred formulations can be contacted with a targeted site, such as a local wound site, a chronic wound, epithelial tissue surface or other intended site of administration by spray, perfusion dip and / or painting; Such formulations may be prepared by a number of established methods known in the art to test skin penetration of drug compositions (see, for example, Wagner et al., 2002 J. Invest. Dermatol. 118: 540, and references cited therein; Bronaugh et al., 1985 J. Pharm. Sci. 74:64; Bosnian et al., 1998 J. Pharm. Biomed. Anal. 17: 493-499; Bosnian et al., 1996 J. Pharm Biomed Anal. 1996 14: 1015-23; Bonferoni et al., 1999 Pharm. Dev. Technol. 4: 45-53; Frantz, Instrumentation and methodology for in vitro skin diffusion cells in methodology for skin absorption. In: Methods for Skin Absorption (Kemppainen & Reifenrath, Eds), CRC Press, Florida, 1990, pp. 35-59; Tojo, Design and calibration of in vitro permeation apparatus. In: Transdermal Controlled Systemic Medications (Chien YW, Ed), Marcel Dekker, New York, 1987, 127-158; Barry, Methods for studying percutaneous absorption. In: Dermatological Formulations: Percutaneous absorption, Marcel Dekker, New York, 1983, 234-295], and may exhibit easy penetration into the skin.

Compositions to be administered to a subject or patient &apos; s skin, and formulations comprising such compositions, may take the form of one or more dosage units in certain embodiments, wherein the liquid filled capsules or ampoules may contain a single dosage unit , Topically-formulated containers such as those described herein in the form of aerosols may hold a plurality of dosage units. Actual methods of producing such dosage forms will be known or apparent to those skilled in the art (see, e. G., The Science and Practice of Pharmacy, 20th Edition, Philadelphia College of Pharmacy and Science, 2000). The composition or formulation to be administered is in any case provided as provided herein with a therapeutically effective amount of a preservative and / or a wound healing stimulating compound (e.g., a BT compound), or a pharmaceutically acceptable salt thereof, in accordance with the teachings of the present invention something to do.

As noted above, the topical formulations of the present invention can take any of a wide variety of forms and include, for example, creams, lotions, liquids, sprays, gels, ointments, pastes, Micelles, and / or microspheres (see, e.g., U.S. Patent No. 7,205,003). For example, as is well-known in the art of pharmaceutical and medicinal cosmetic formulations, the cream formulations may be formulated as a creamy, oil-in-water or oil-in-water viscous liquid Or semi-solid emulsion. The cream base is water-washable and contains an oil phase, an emulsifier and an aqueous phase. The oil phase, also referred to as the " inner " phase, is generally composed of petroleum and fatty acids such as cetyl or stearyl alcohol. The aqueous phase generally, although not necessarily, exceeds the oil phase in volume, and generally contains a wetting agent. Emulsifiers in a cream formulation are generally nonionic, anionic, cationic or amphoteric surfactants.

Lotion preparations which are suitable for the delivery of cosmetic preparations are preparations to be applied to the skin surface without friction and are typically liquid or semi-liquid preparations in which the solid particles comprising the active agent are present in water or in an alcoholic base. Lotions are generally suspensions of solids, preferably liquid-in-oil emulsions of the water-in-oil type. Lotion preparations are the preferred formulations of the present invention for treating large body parts due to the ease of application of the compositions being more fluid. It is generally preferred that the insoluble material in the lotion formulation is finely divided. Lotion preparations will typically contain suspending agents to obtain a better dispersion and compounds useful for localizing and maintaining the active agent upon contact with the skin, such as methylcellulose, sodium carboxymethyl-cellulose, and the like.

Liquids are homogeneous mixtures prepared by dissolving one or more chemicals (solutes) in a liquid such that the molecules of the dissolved material are dispersed in the molecules of the solvent. The liquid agent may contain other pharmaceutically acceptable and / or medicinal cosmetically acceptable chemicals for buffering, stabilizing or preserving the solute. Common examples of solvents used to make the liquid solutions are ethanol, water, propylene glycol or any other pharmaceutically acceptable and / or pharmaceutically acceptable cosmetically acceptable vehicle.

The gel is a semi-solid, suspension-type system. The single-phase gel typically contains an organic macromolecule that is aqueous but also preferably substantially homogeneously dispersed throughout the carrier liquid containing an alcohol, and optionally an oil. Preferred "organic macromolecules," i.e., gelling agents are, for chemical cross-linked acrylic acid polymers such as such as an acrylic acid polymer cross-linked, for the "carbomer" family, for example, of a polymer, commercially available under the Carbopol ^® trade name Carboxypolyalkylene. &Lt; / RTI &gt; Also, in certain embodiments, hydrophilic polymers such as polyethylene oxide, polyoxyethylene-polyoxypropylene copolymer and polyvinyl alcohol; Polymers such as hydroxypropylcellulose, hydroxyethylcellulose, hydroxypropylmethylcellulose, hydroxypropylmethylcellulose phthalate and methylcellulose; Gums such as tragacanth and xanthan gum; Sodium alginate; And gelatin. To prepare a single-form gel, a dispersing agent such as alcohol or glycerin may be added, or the gelling agent may be dispersed by grinding, mechanical mixing or stirring, or a combination thereof.

Ointments well known in the art are typically semi-solid preparations based on petroleum or other petroleum derivatives. As will be familiar to those skilled in the art, the particular ointment substrate to be used will provide a number of desirable features, such as, for example, emolliency. By using other carriers or vehicles, the ointment substrate should be inert, stable, non-irritating and nonsensitizing. Remington: The Science and Practice of Pharmacy, 19th Ed. (Easton, Pa .: Mack Publishing Co., 1995), at pages 1399-1404, the ointment substrate may be divided into four classes: a retentive substrate; An emulsifiable substrate; A latex substrate; And water-soluble substrates. Retentive ointment bases include, for example, vegetable oils, fats obtained from animals, and semisolid hydrocarbons obtained from petroleum. Emulsifiable ointment bases, also known as adsorptive ointment bases, contain little or no water, including, for example, hydroxystearin sulfate, anhydrous lanolin, and hydrophilic oils. Latex ointment bases include oil-in-water (W / O) emulsions or water-in-oil emulsions such as 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, Id.).

The pastes are semi-solid dosage forms in which the active agent is suspended in a suitable substrate. Depending on the properties of the substrate, the paste is dispensed between those made from an oleaginous paste or a single-phase aqueous gel. The substrate in the fatty paste is generally petroleum or hydrophilic oil. Pastes prepared from single-aqueous gels generally incorporate carboxymethylcellulose or the like as a base.

Formulations may also be prepared with liposomes, micelles, and microspheres. Liposomes are microscopic vesicles having one (single lamella) or a plurality (majority lamella) of liquid layers comprising a lipid bilayer and in this context on their liquid membrane surface, one or more components of the topical formulations described herein, For example, antiseptic, wound healing / dermal tissue / epithelial tissue repair-promoting compounds (eg, particulate BT compounds optionally with one or more antibiotics) or a particular carrier or excipient may be encapsulated and / or adsorbed. The liposome formulations herein include cationic (positively charged), anionic (negatively charged), and neutral preparations. Cationic liposomes are readily available. N, N, N-triethylammonium (DOTMA) liposomes are commercially available under the trade names Lipofectin ^® [ Grand Island of Iceland]. Similarly, anionic and neutral liposomes can also be prepared using materials readily available or readily available, for example, from Avanti Polar Lipids (Birmingham, Ala.). Such materials include, among others, phosphatidylcholine, cholesterol, phosphatidylethanolamine, dioloylphosphatidylcholine (DOPC), dioloylphosphatidylglycerol (DOPG), and dioloylphosphatidylethanolamine (DOPE). These materials can also be mixed with DOTMA at an appropriate ratio. Methods for making liposomes using these materials are well known in the art.

Micelles are composed of ordered surfactant molecules so that their polar headgroups form an outer spherical shell while the hydrophobic hydrocarbon chains are oriented towards the center of the sphere to form the core, Lt; / RTI &gt; Micelles are formed in an aqueous solution containing a surfactant at a sufficiently high concentration, whereby micelles are naturally produced. Surfactants useful for forming micelles include but are not limited to potassium laurate, sodium octanesulfonate, sodium decane sulfonate, sodium dodecane sulfonate, sodium lauryl sulfate, doxate sodium decyltrimethylammonium bromide, dodecyltrimethylammonium bromide, But are not limited to, ammonium bromide, tetradecyltrimethylammonium chloride, dodecylammonium chloride, polyoxyl-8 dodecyl ether, polyoxyl-12 dodecyl ether, nonoxynol 10, and noroxynol 30.

Similarly, microspheres can be incorporated into the topical formulations described herein. Similar to liposomes and micelles, microspheres essentially encapsulate one or more of the present formulations. These are generally, but not necessarily, formed from charged lipids such as lipids, preferably phospholipids. The preparation of lipid microspheres is well known in the art.

Various additives as known to those skilled in the art may also be included in topical formulations. For example, a solvent comprising a relatively small amount of alcohol may be used to dissolve a particular formulation component. In a topical formulation, it may be desirable to include the added skin penetration enhancer in the formulation in the case of a particular topical formulation or, in particular, severe skin damage such as post-surgical acute or chronic wound or post-surgical skin wound. Examples of suitable enhancers include ethers such as diethylene glycol monoethyl ether (commercially available as Transcutol ^® ) and diethylene glycol monomethyl ether; Sodium lauryl sulfate, cetyltrimethylammonium bromide, benzalkonium chloride, Poloxamer ^® (231, 182, 184), Tween ^® (20,40,60,80), and lecithin (U.S. Patent No. 4,783,450) Surface active agents such as; Alcohols such as ethanol, propanol, octanol, benzyl alcohol and the like; Polyethylene glycols and esters thereof, such as polyethylene glycol monolaurate (PEGML; see, for example, U.S. Patent 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; Terpene; Alcanon; And organic acids, especially citric acid and succinic acid. Azone, such as DMSO and C ₁₀ MSO ^® and sulfoxide side, but can also be used, which is not substantially preferred.

The most preferred skin penetration enhancers are the lipophilic co-enhancements typically referred to as " plasticizers " enhancers, i.e. those having a molecular weight of about 150 to 1000 daltons and having a water solubility of less than about 1% Less than 0.5% by weight, and most preferably less than about 0.2% by weight. The Hildebrand solubility parameter of the plasticizer is in the range of from about 2.5 to about 10, preferably from 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 are methyl laurate, ethyl laurate, propylene glycol monolaurate, propylene glycerol dilaurate, glycerol monolaurate, glycerol monooleate, isopropyl n-decanoate, The thread contains a state in advance. The fatty alcohols include, for example, stearyl alcohols and oleyl alcohols, while the fatty ethers include diols or triols, preferably C ₂ -C ₄ alkanediols or triols, with one or two fatty ether substituents &Lt; / RTI &gt; Additional skin penetration enhancers will be known to those of ordinary skill in the art of topical drug delivery and / or may be found in the pertinent literature (see, e.g., Percutaneous Penetration Enhancers, eds. Smith et al. (CRC Press, Boca Raton, FL, 1995).

Various other additives may be included in topical formulations according to certain embodiments of the present invention, in addition to those defined above. They may also contain other classes of substances which may be cosmetically, medically present or even desirable such as antioxidants, astringents, perfumes, preservatives, softeners, pigments, dyes, moisturizers, propellants and sunscreen agents, But is not limited thereto. Representative examples of optional additives for incorporation into formulations of certain embodiments of the present invention are as follows: preservatives such as sorbate; A solvent such as isopropanol and propylene glycol; Astringents such as menthol and ethanol; A softening agent such as polyalkylene methyl glucoside; Moisturizers such as glycerin; Emulsifiers such as glycerol stearate, PEG-100 stearate, polyglyceryl-3hydroxy lauryl ether, and polysorbate 60; Sorbitol and other polyhydroxy alcohols such as polyethylene glycol; Sunscreen agents such as octyl methoxycinnamate (commercially available as Parsol MCX) and butyl methoxybenzoylmethane (available under the trade name Parsol 1789); Ascorbic acid (vitamin C), α- tocopherol (vitamin E), tocopherol, β-, γ- tocopherol, tocopherol δ-, ε- tocopherol, ζ ₁ - tocopherol, ζ ₂ - tocopherol, η- tocopherol, and retinol (vitamin A Antioxidants such as; Essential oils, ceramides, essential fatty acids, mineral oil, wetting agents, and, other surface active agents, for example, BASF (BASF), PLURONIC ^® series, vegetable oil of a hydrophilic polymer, available from (NJ Mount Olive material) (e.g., (E.g., soybean oil, palm oil, liquid fraction of cedar butter, sunflower oil), animal oils (e.g., hydro hydroqualone), mineral oils, synthetic oils, silicone oils or waxes (e.g., cyclomethicone and dimethicone) , Fluorinated oils (typically perfluoropolyethers), fatty alcohols (e.g., cetyl alcohol), and waxes (e.g., waxes, carnauba waxes, and paraffin waxes); Skin-feel modifier; And a water-swelling clay and the crosslinked Carbopol ^®, which may be commercially obtained under the trade as - thickening and structuring agents such as a combination carboxy polyalkylene (structurant).

Other additives include beneficial agents, such as substances that keep it soft and / or protect the skin by delaying the conditioning and reducing the water content of the skin (especially the upper layer of the skin in the stratum corneum). Such conditioners and wetting agents include, by way of example, pyrrolidinecarboxylic 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; Anti-balloons such as retinoic acid; Vasodilators such as nicotinic acid; Melanogenesis inhibitors such as kojic acid; And mixtures thereof. Hydroxycarboxylic acids, polymeric hydroxy acids, wetting agents, collagen, marine extracts and ascorbic acid (vitamin C), as well as other medicinally active agents, such as, for example, There may be an antioxidant such as alpha -tocopherol (vitamin E) or other tocopherols such as those described above, and retinol (vitamin A), and / or a cosmetically acceptable salt, ester, amide, or other derivative thereof . Additional cosmetic preparations include, for example, those that can enhance the oxygen supply to the skin tissue, as described in WO 94/00098 and WO 94/00109. A sunscreen agent may also be included.

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

Formulations of certain embodiments of the present invention may also include conventional additives such as emulsifiers, perfumes, colorants, gelling agents, thickeners, stabilizers, surface active agents and the like. Other agents such as antimicrobial agents may also be added to prevent spoilage during storage, i.e., to inhibit the growth of microorganisms such as yeast and fungi. Suitable antimicrobial agents are typically selected from p-hydroxybenzoic acid (e. G., Methyl and propyl paraben), sodium benzoate, sorbic acid, imidurea, and combinations thereof. The formulations may also be used to minimize or eliminate the possibility of skin irritation or skin damage resulting from an anti-infective acute or chronic wound healing and dermal tissue recovery-promoting compound to be administered by containing a stimulus-relieving additive, can do. Suitable stimulation-relieving additives include, for example,? -Tocopherol; Monoamine oxidase inhibitors, especially phenyl alcohols such as 2-phenyl-1-ethanol; glycerin; Salicylate; Ascorbate; An ion-permeable carrier such as monensin; Amphoteric amine; Ammonium chloride; N-acetylcysteine; Capsaicin; And a key queen. Stimulation-relieving additives, if present, can be incorporated at concentrations effective to relieve irritation or skin damage in topical formulations, typically up to about 20% by weight of the formulation, more typically up to about 5% by weight.

Topical formulations may also contain one or more synergistic antibiotics, such as the antimicrobial / wound healing / anti-bacterial / dermal tissue repair-promoting compounds (e. G., Preferably as substantially homogeneous microparticles provided herein, In combination with one or more additional pharmacologically active agents suitable for topical administration in therapeutically effective amounts. Such formulations may be, for example, phospholipids and oxygen-loaded fluorocarbons, such as those described in International Patent Publications WO 94/00098 and WO 94/00109, Asymmetric lamellar aggregates comprised of a mixture of fluorocarbon compounds.

Suitable pharmacologically active agents that can be topically applied by incorporation into the topical formulations of the invention include, but are not limited to, those that improve or eradicate pigmented or non-pigmented black mold, keratosis, and wrinkles Formulation; Antimicrobial agents; Antibacterial agents; Anti-itch and anti-dryness agents; Antiinflammatory agents; Local anesthetics and analgesics; Corticosteroids; Retinol (e. G., Retinoic acid); vitamin; hormone; And antimetabolites, but are not limited thereto. Some examples of topical pharmacologically active agents are acyl chlorvir, amphotericin, chlorhexidine, clotrimazole, ketoconazole, econazole, myconazole, metronidazole, minocycline, nystatin, neomycin, kanamycin, But are not limited to, aminobenzoic acid ester, octyl methoxycinnamate, octyl salicylate, oxybenzone, dioxybenzone, tocopherol, tocopheryl acetate, selenium sulfide, zinc pyrithione, diphenhydramine, pramosin, lidocaine, procaine, erythromycin, And the like. Examples of the antioxidant include tetracycline, clinodimin, crotamiton, hydroquinone and its monomethyl and benzyl ethers, naltoxene, ibuprofen, cromolyn, retinoic acid, retinol, retinyl palmitate, retinyl acetate, coal tar, Ceflobin, estradiol, hydrocortisone, hydrocortisone 21-acetate, hydrocortisone 17-valerate But are not limited to, hydrocortisone 17-butyrate, progesterone, betamethasone valerate, betamethasone dipropionate, triamcinolone acetonide, fluorocinonide, clobetasol propionate, minoxidil, dipyridamole, diphenylhydantoin, benzoyl peroxide , And 5-fluorouracil. As also shown above, certain embodiments include a combination of a carbapenem antibiotic, a cephalosporin antibiotic, a fluoroquinolone antibiotic, a glycopeptide antibiotic, a rincozyme antibiotic, a penicillinase-resistant penicillin antibiotic, an aminopenicillin antibiotic, Consider incorporating into the formulation of antibiotics such as aminoglycoside antibiotics.

The pharmacologically acceptable carrier may also be incorporated into the topical formulations of this particular embodiment and may be the particular carrier conventionally used in the art. Examples include water, lower alcohols, higher alcohols, honey, polyhydric alcohols, monosaccharides, disaccharides, polysaccharides, sugar alcohols such as glycols (2- carbon), glycerol (3- carbon), erythritol, Carbon), arabitol, xylitol and ribitol (5-carbon), mannitol, sorbitol, dititol and iditol (6-carbon), isomaltol, maltitol, lactinol and polyglycitol, And oils, waxes, fatty acids, silicone oils, nonionic surfactants, ionic surfactants, silicone surfactants, and water-based and emulsion-based mixtures of such carriers.

The topical formulation of the present invention may be applied regularly to acute or chronic wound sites (e. G., Surrounding tissues including normal or healthy peripheral tissues that are considered to be unaffected by the wound itself or infection) The epithelial tissue surfaces (e.g., the gastrointestinal tract, respiratory tract, and gland tissue) require treatment with the number and volume needed to achieve the desired result. The number of treatments may depend on the nature of the skin (or other epithelial tissue) condition (e.g., acute or chronic wound, or other skin wound or other type of skin wound, such as may be found in a defect resulting from a surgical incision) The activity of the active ingredient in the particular aspect of the skin (or other tissue) (e.g., the BT compound and optionally one or more additional pharmaceutically active ingredients such as antibiotics, such as amikacin or other antibiotics, The effectiveness of the vehicle used to deliver the active ingredient (s) to the appropriate layer of skin (or other epithelial surface-containing tissue), the effectiveness of the formulation Ease of removal by physical contact with bandages or other dressings or garments, or removal thereof by sweat or other inherent or external fluid, Depends on the convenience of the activity level or lifestyle of the patient.

Representative concentrations of the active substance, such as the BT compound antiseptic / anti-bacterial / wound-healing / dermal tissue restoration-promoting composition described herein, can be, for example, from about 0.001 to 30% by weight, based on the total weight of the composition, To 0.01% to 5.0%, and more preferably to about 0.1% to 2.0%. As a representative example, the composition of this embodiment of the present invention may be applied to acute or chronic wounds and / or skin at the same rate as about 1.0 mg / cm ^{2 of} skin to about 20.0 mg / cm ² of skin. Representative examples of topical formulations include, but are not limited to, aerosols, alcohols, anhydrous substrates (e.g., lipsticks or powders), aqueous solutions, creams, emulsions (oil-in-water or water- But are not limited to, gels, hydro-alcoholic solutions, liposomes, lotions, microemulsions, ointments, oil agents, organic solvents, polyols, polymer preparations, powders, silicone derivatives, and waxes Do not. Topical formulations may include, for example, chelating agents, conditioning agents, softening agents, excipients, wetting agents, protectants, thickeners, or UV absorbers. Those skilled in the art will recognize that formulations other than those listed may be used in embodiments of the present invention.

Chelating agents may optionally be included in the topical formulations, and may be selected from any agent suitable for use in cosmetic ^{compositions, Ca 2 +, Mn 2 +} , or 2, such as Mg ^{2 +} some with the ability to bind to the cationic metal Natural or synthetic chemicals. Examples of chelating agents include, but are not limited to, EDTA, disodium EDTA, EGTA, citric acid, and dicarboxylic acids.

Conditioning agents may also optionally be included in topical formulations. Examples of skin conditioners include, but are not limited to, acetylcysteine, N-acetyldichlorospingosine, acrylate / behenyl acrylate / dimethiconacrylate copolymer, adenosine, adenosine cyclic phosphate, adenosine phosphate, adenosine triphosphate, alanine, But are not limited to, algae extract, allantoin and derivatives, aloe verbadensis extract, aluminum PCA, amyloglucosidase, arbutin, arginine, azulene, bromelain, buttermilk powder, butylene glycol, caffeine, calcium gluconate, (S) selected from the group consisting of carboxy stearin, carnosine, beta- carotene, casein, catalase, cephalin, ceramides, chamomilla recutits (matricaria) flower extract, cholecalciferol, cholesteryl ester, -Betaine, coenzyme A, modified corn starch, crystals, cycloethoxymethicone, cysteine DNA, cytochrome C, Elastin, amino acid, elastin, epithelial growth factor, ergocalciferol, ergosterol, ethylhexyl PCA, fibronectin, folic acid, gelatin, liposomes, liposomes, lactose, dextran sulfate, dimethicone copolyol, dimethylsilanol hyaluronate, Glycogen, glycogen, glycine, glycolipid, glycoprotein, glycosaminoglycan, glycoprotein glyphide, horseradish peroxidase, hydrogenated protein, hydrolyzed protein, jojoba oil, Keratin, keratin amino acids, and kinetin, lactoferrin, lanosterol, lauryl PCA, lecithin, linoleic acid, linolenic acid, lipase, lysine, lysozyme, malt extract, maltodextrin, melanin, methionine, inorganic salts, niacin, Orzanol, palmitoyl hydrolyzed protein, pancreatin, papain, PEG, pepsin, phospholipid, phytosterol, placental enzyme, Rocobacillus acetate, Tocopherol acetate, Tocopherol acetate, Tocopherol acetate, Tocopherol acetate, Tocopherol acetate, Tocopherol acetate, Tocopherol acetate, Tocopherol acetate, But are not limited to, peryllinolate, ubiquinone, Vitiviniferra (grape) seed oil, wheat amino acid, xanthan gum, and zinc gluconate. Skin conditioning agents other than those listed above may be combined with the compositions or formulations described herein provided herein as would be readily appreciated by those skilled in the art.

Topical formulations may also optionally comprise one or more softening agents, examples of which are acetylated lanolin, acetylated lanolin alcohol, acrylate / _C10-30 alkyl acrylate crosspolymer, acrylate copolymer, alanine, algae extract , Aloe barbadensis extract or gel, althea officinalis extract, aluminum starch octenyl succinate, aluminum stearate, apricot (prunus armeniaca) seed oil, arginine, arginine asparagine Ascorbic acid, aspartic acid, aspartic acid, avocado [persea gratissima] oil, barium sulphate, barrier sphingolipids, fatty alcohols such as alcohols, such as arnica montana extract, ascorbic acid, ascorbyl palmitate, , Beeshenyl alcohol, beta-sitosterol, BHT, birch [betula alba] bark extract, 2-bromo-2-nitropropane-1,3-diol, butcherbroom (ruscus aculeatus) extract, butylene glycol , Calendula officinalis extract, calendula porphinificinil oil, candeleria [euphorbia cerifera] wax, canola oil, caprylic / capric triglyceride, cardamone [ (elettaria cardamomum) oil, carnauba [copernicia cerifera] wax, carrageenan [chondrus crispus], carrot [daucus carota sativa] Oil, carter [ricinus communis] oil, ceramide, ceresin, ceteareth-5, ceteareth-12, ceteareth-20, ceteoryl octanoate, Ceteth-24, cetyl acetate, cetyl octanoate, cetyl palmitate Cholesterol, cholesterol ester, cholesteryl hydroxystearate, citric acid, clary [salvia sclarea] oil, cocoa [the ovary cacao (theobroma cacao) butter, coco-caprylate / caprate, coconut (cocos nucifera) oil, collagen amino acid, corn [zea mays] oil, fatty acid, decyl oleate, dextrin, Dimethicone copolyol, dimethiconol, dioctyl adipate, dioctyl succinate, dipentaerythritol hexa caprylate / hexacaprate, DMDM hydantoin, DNA, erythritol, ethoxydi Eucalyptus globulus oil, evening primrose oil (oenothera biennis) oil, fatty acid, truckose, gelatin, Geranium &lt; RTI ID = 0.0 & Glyceryl myristate, glyceryl myristate, glyceryl myristate, glycerin, glycerin, glycerin, glyceryl distearate, glyceryl hydroxystearate, glyceryl laurate, glyceryl linoleate, glyceryl myristate, Glyceryl stearate SE, glycine, glycol stearate, glycol stearate SE, glycosaminoglycan, grape [vitis vinifera] seed oil, hazelnut [Coriolus americana corylus americana] nuts oil, western hazel [corylus avellana] nuts oil, hexylene glycol, honey, hyaluronic acid, safflower [carthamus tinctorius] oil, hydrogenation Hydrogenated coconut oil, hydrogenated coconut oil, hydrogenated coconut oil, hydrogenated lanolin, hydrogenated lecithin, Hydrogenated palm kernel oil, hydrogenated soybean oil, hydrogenated resin glyceride, hydrogenated vegetable oil, hydrolyzed collagen, hydrolyzed elastin, hydrolyzed glycosaminoglycan, hydrolyzed palm kernel oil, hydrolyzed palm kernel oil, Keratin, hydrolyzed soy protein, hydroxylated lanolin, hydroxyproline, imidazolidinyl urea, iodopropynyl butyl carbamate, isocetyl stearate, isocetyl stearoyl stearate, isodecyl oleate , Isopropyl isostearate, isopropyl lanolate, isopropyl myristate, isopropyl palmitate, isopropyl stearate, isostearamide DEA, isostearic acid, isostearyl lactate, isostearyl neopentanoate, Jasmine (jasminum officinale) oil, jojoba [buxus (buxus) chinensis] oil, kelp, kukui [aleurites moluccana] nuts oil, lactamide MEA, lanes-16, lanes-10 acetate, lanolin, lanolin, lanolin alcohol , Lanolin oil, lanolin wax, lavender [lavandula angustifolia] oil, lecithin, lemon citrus medica limonum oil, linoleic acid, linolenic acid, macadamia ternifolia ) Nuts oil, magnesium stearate, magnesium sulfate, maltitol, matricaria [chamomilla recutita] oil, methylglucose sequestrate, methylsilanol PCA, microcrystalline wax, mineral oil, mink oil , Mortierella oil, myristyl lactate, myristyl myristate, myristyl propionate, neopentyl glycol dicaprylate / dicaprate, octyldodecanol, octyl Octyldodecyl stearoyl stearate, octyl hydroxystearate, octyl palmitate, octyl salicylate, octyl stearate, oleic acid, olea olea europaea oil, orange [ Citrus aurantium dulcis] oil, palm [elaeis guineensis] oil, palmitic acid, pantethine, panthenol, panthenyl ethyl ether, paraffin, PCA, peach [prunus Arachis hypogaea] oil, PEG-8 C1218 ester, PEG-15 cocamine, PEG-150 distearate, PEG-60 glyceryl PEG-20 hydrogenated castor oil, PEG-40 hydrogenated castor oil, PEG-5 hydrogenated castor oil, PEG-30 hydrogenated castor oil, PEG-40 hydrogenated castor oil, Quinostearate, P EG-40 sorbitan peroleate, PEG-5 soybean sterol, PEG-10 soy sterol, PEG-2 stearate, PEG-8 stearate, PEG-20 stearate, PEG-32 stearate, PEG- , PEG-50 stearate, PEG-100 stearate, PEG-150 stearate, pentadecaractone, peppermint [mentha piperita] oil, petroleum, phospholipids, polyamine sugar condensates, polyglyceryl- Isostearate, polyquantnium-24, polysorbate 20, polysorbate 40, polysorbate 60, polysorbate 80, polysorbate 85, potassium myristate, potassium palmitate, potassium sorbate, potassium stearate , Propylene glycol, propylene glycol dicaprylate / dicaprate, propylene glycol dioctanoate, propylene glycol dipelargonate, propylene glycol laurate, propylene glycol stearate, (Oryza sativa), rice paddy root, oryzae sativa SE, PVP, pyridoxine dipalmitate, quaternium-15, quaternium-18 hectorite, quaternium-22, retinol, retinyl palmitate, The present invention relates to barn oil, RNA, rosemary rosmarinus officinalis oil, rose oil, safflower [carthamus tinctorius] oil, sage [salvia officinalis salvia officinalis oil, salicylic acid, petroleum jelly (santalum album) oil, serine, sesame protein, Sesame sesamum indicum oil, shea butter [butyrospermum sodium stearate, soluble collagen, sorbic acid, sorbitan laurate, sorbitan monolaurate, sorbitan monolaurate, sorbitan monolaurate, sorbitan monolaurate, sorbitan monolaurate, sorbitan monolaurate, Sorbitan oleate, sorbitan palmitate, sorbitan sesquioleate, sorbitan stearate, sorbitol soybean [glycine soja] oil, sphingolipids, squalane, squalane, stearamide MEA-stearate, Stearyl stearate, sunflower [helianthus annuus] seed oil, dextrin (stearyl glyceryl stearate, stearyl stearate, stearyl stearate, (Prunus amygdalus dulcis) oil, synthetic beeswax, tocopherol, tocopheryl acetate, tocopheryl linoleate, tribehenin, tridecyl neopentanoate, tridecyl stearate, triethanolamine , Tristearin, urea, vegetable oil, water, wax, wheat (triticum vulgare) sprout oil, and ylang ylang) [cananga odorata] oil.

In some embodiments, topical formulations may contain suitable excipients that are highly compatible with the skin, have good resistance, are stable, and can provide consistency that is readily exploited. Suitable topical excipients and vehicles are well known in the art for special applications by those skilled in the art, and in particular those described in Remington ' s Pharmaceutical Sciences, Vol. 18, Mack Publishing Co., Easton, Pa. (1990), in particular Chapter 87], which is one of many standard textbooks in the field. Optionally, at least one wetting agent is also included in the topical formulation. Examples of wetting agents include, but are not limited to, amino acids, chondroitin sulfate, diglycerin, erythritol, fructose, glucose, glycerin, glycerol, glycols, 1,2,6- hexanetriol, honey, hyaluronic acid, hydrogenated honey, hydrogenated starch hydrolyzate, PEG-15 butanediol, polyglyceryl sorbitol, salts of pyrrolidonecarboxylic acid, potassium PCA, propylene glycol, sodium gluconate, sodium PCA, sorbitol, sorbitol, sorbitol, But are not limited to, cross, trehalose, urea, and xylitol.

Certain embodiments contemplate topical formulations containing one or more additional skin protectants. Examples of skin protectants are algae extract, allantoin, aluminum hydroxide, aluminum sulfate, betaine, camellia leaf extract, cervulocide, dimethicone, glucuronolactone, glycerin, kaolin, lanolin, malt extract, , Potassium gluconate, and talc.

Surfactants may also be included in the specific topical formulations contemplated herein and may be any natural or synthetic surfactant suitable for use in cosmetic compositions such as cationic, anionic, zwitterionic, or non-ionic surfactants, or mixtures thereof, Surfactants and Interfacial Phenomena, " Second Edition, John Wiley &amp; Sons, New York, 1988, Chapter 1, pages 4 31]. Examples of cationic surface active agents are DMDAO or other amine oxides, long chain primary amines, diamines and polyamines and their salts, quaternary ammonium salts, polyoxyethylenated long chain amines, and quaternized polyoxyethylenated long chain amines But is not limited thereto. Examples of anionic surfactants include SDS; Salts of carboxylic acids (e.g., soaps); Salts of sulfonic acid, salts of sulfuric acid, phosphoric acid and polyphosphoric acid esters; Alkyl phosphates; Monoalkyl phosphate (MAP); And salts of perfluorocarboxylic acids. Examples of zwitterionic surface active agents include, but are not limited to, comoamidopropylhydroxybutyrate (CAPHS) and others that are pH-sensitive and require special attention in designing the proper pH of the formulation But are not limited to, thiocarboxylic acids, thiocarboxylic acids, thiocarboxylic acids, thiocarboxylic acids, thiocarboxylic acids, thiocarboxylic acids, Examples of non-ionic surfactants are alkylphenol ethoxylates, alcohol ethoxylates, polyoxyethylenated polyoxypropylene glycols, polyoxyethylenated mercaptans, long chain carboxylic acid esters, alkanolamides, , Polyoxyethylene silicones, N-alkylpyrrolidones, and alkyl polyglycosidases. Ionic detergent or, PLURONICS ^® Series [manufactured - a wetting agent, a mineral oil or non-: The other surface-active agent such as a preparation such as one or more members of BASF (BASF), NJ, Mount Olive material) In addition, for example and non-limiting May be included to prevent agglomeration of the BT microparticles in the particulate suspension according to theory. Any combination of surface active agents is suitable. Certain embodiments include at least one anionic and one cationic surfactant or at least one cationic and one zwitterionic surfactant that is miscible, i.e., does not form a noticeably precipitated complex upon mixing .

Examples of thickeners that may also be present in a particular topical formulation include, but are not limited to, acrylamide copolymers, agarose, amylopectin, betonite, calcium alginate, calcium carboxymethyl cellulose, carbomer, carboxymethyl chitin, cellulose gum, dextrin, gelatin, Hydroxypropylcellulose, hydroxypropyl starch, magnesium alginate, methylcellulose, microcrystalline cellulose, pectin, various PEG, polyacrylic acid, polymethacrylic acid, polyvinyl alcohol, various PPG , Sodium acrylate copolymer, sodium carrageenan, xanthan gum, and yeast beta-glucan. Thickening agents other than those listed above may also be used in embodiments of the present invention.

According to a particular embodiment contemplated herein, topical formulations may comprise one or more sunscreens or UV absorbers. When ultraviolet- (UVA and UVB) absorption characteristics are required, these agents may be used in combination with, for example, benzophenone, benzophenone-1, benzophenone-2, benzophenone-3, benzophenone- Benzophenone-11, benzophenone-12, benzyl salicylate, butyl PABA, cinnamate ester, synoxate, DEA - methoxycinnamate, diisopropylmethyl cinnamate, ethyl dihydroxypropyl PABA, ethyldiisopropyl cinnamate, ethyl methoxy cinnamate, ethyl PABA, ethyl urocanate, glyceryl octanoate dimethoxycinnamate , PABA, PABA ester, Parsol 1789, Palladium, Palladium, Palladium, Palladium, Palladium, Palladium, And isopropylbenzyl salicylate, and mixtures thereof . Those skilled in the art will recognize that besides what is listed, a line screening and UV absorbing or protective agent can be used in certain embodiments of the present invention.

The topical formulations described herein are typically effective at pH values between about 2.5 and about 10.0. Preferably, the pH of the composition is in the vicinity of or near the following pH range: 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 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 can adjust the pH to an acceptable range by adding an appropriate pH adjusting component to the composition of the present invention. The " about " stated pH includes formulations in which the actual measured pH at any time provided to a person skilled in the art may be less than or greater than a specified value below 0.7, 0.6, 0.5, 0.4, 0.3, , Where formulation and storage conditions can be created by shifting the pH from the original value.

Creams, lotions, gels, ointments, pastes and the like may be sprayed onto or gently rubbed against the affected surface. Liquids can be applied in the same way, but will more typically be applied with a dropper, a swab or the like, and will be carefully applied to the affected area. The application regimen will depend on a number of factors that can be readily measured, such as the severity of the wound and its responsiveness to the initial treatment, but will generally include one or more applications per day on a progression basis. One of ordinary skill in the art can readily determine the optimum amount, mode of administration, and repetition rate of the formulation to be administered. In general, the formulations of these and related aspects of the invention will be applied in the range of once, twice, three times, more than four times daily, more than once a week or more than once a week.

As discussed further above, the topical formulations useful herein are those that are pharmaceutically acceptable, including any suitable diluent or excipient, including any pharmaceutical agent that is not detrimental to the recipient to which the composition is provided and can be administered without undue toxicity It contains an acceptable carrier. Pharmaceutically acceptable carriers include, but are not limited to, liquids such as water, saline, glycerol, ethanol and the like, and also include film-forming agents such as viscosity enhancers (e.g., balsam fir resin) or colloidion or nitrocellulose solutions . &Lt; / RTI &gt; A complete discussion of pharmaceutically acceptable carriers, diluents and other excipients is set forth in REMINGTON'S PHARMACEUTICAL SCIENCES (Mack Pub. Co., N. J. current edition).

Where the topical formulation is in the form of a gel- or liquid-filled capsule, for example a gelatin capsule, it may contain, in addition to the above-mentioned materials, a liquid carrier such as polyethylene glycol or oil. Liquid pharmaceutical compositions of certain embodiments of the present invention may include one or more of the following, irrespective of whether they are liquid solutions, suspensions or other similar forms: injection water, saline solution, preferably physiological saline, Ringer's solution, Isotonic sodium chloride, sterile diluents such as synthetic mono- or diglycerides, polyethylene glycols, glycerin, propylene glycol or other solvents which may be provided as a solvent or suspending medium; 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); Buffers such as acetate, citrate or phosphate, and anisotropic agents such as sodium chloride or dextrose.

For topical administration, the carrier may suitably comprise a solution, emulsion, ointment or gel base. For example, such substrates may include one or more of the following: diluents such as petroleum, lanolin, polyethylene glycol, wax, inorganic oils, water and alcohols, and emulsifiers and stabilizers. The thickening agent may be present in a pharmaceutical or medicinal cosmetic composition for topical administration. When intended for transdermal administration, the composition may comprise a transdermal patch or an ion transfer device. Topical formulations may contain a compound of certain embodiments of the present invention in a concentration from about 0.1 to about 10% w / v (weight per unit volume). Topical formulations may be provided in the form of creams, lotions, solutions, sprayers, gels, ointments, pastes, and / or may contain liposomes, micelles, microspheres and / or other particulates or nanoparticle delivery components. Topical formulations are also time which can be directly administered to the wound-release or sustained-release particles or pellets, e.g., sustained release ethylene vinyl acetate polymer [e.g., Elvax ^® 40, manufactured by Aldrich (Aldrich), Wisconsin, Milwaukee, WI].

Topical formulations may include agents that bind to a dermal tissue repair-promoting compound and thus aid in the delivery thereof to skin epithelial cells (e. G. Keratinocytes) and / or fibroblasts. Suitable formulations that may work in such doses include clathrating agents such as cyclodextrins and other agents may include proteins or liposomes.

Topical formulations of certain embodiments of the present invention may also be presented in the form of dosage units that may be administered as aerosols. The term aerosol is used to denote various systems ranging from those of a colloidal character to systems consisting of a pressurized package. The delivery may be by liquefied or squeezed gas or by an appropriate pump system for dispensing the active ingredient. Aerosols of the compounds of certain embodiments of the present invention can be delivered to a single-phase, two-phase, or three-phase system to deliver the active ingredient (s). The delivery of the aerosol includes the necessary vessels, activators, valves, small vessels, etc., which together form a kit. One of ordinary skill in the art can determine the preferred aerosol formulation for delivery of the topical formulation to the skin or wound site without undue experimentation.

Topical formulations may be prepared by methods well known in the art of pharmacy. For example, a pharmaceutical composition intended to be administered by spray, wash, or rinse to a wound site or skin may be prepared by dissolving the BT preservative / wound-healing / anti-bacterium / skin tissue repair-promoting compound as described herein in sterile distilled water To form a solution. Surfactants may be added to promote the formation of a homogeneous solution or suspension. Surfactants facilitate dissolution or homogeneous suspension of compounds in aqueous delivery systems by non-covalent interaction with antioxidant active compounds.

The BT preservative / wound-healing / anti-bacterial / dermal tissue repair-promoting compound for use in topical formulations or pharmaceutically acceptable salts thereof is administered in a therapeutically effective amount, , The activity of the specific BT compound used (incorporation or absence of aminoglycoside antibiotics, for example from the formation of antibiotics such as amikacin); Metabolic safety and action length of the compound; Age, weight, general health, sex, skin type, immune status and diet of the subject; Mode and time of administration; Discharge rate; Drug combination; Severity of special skin wounds requiring restoration of skin tissue; And the therapeutic regimen the subject is undergoing. Generally, a therapeutically effective daily dose is from about 0.001 mg / kg (i.e., 0.07 mg) to about 100 mg / kg (i.e., 7.0 g) for a 70 kg mammal; Preferably, the therapeutically effective dose is about 0.01 mg / kg (i.e., 7 mg) to about 50 mg / kg (i.e., 3.5 g) (for a 70 kg mammal); More preferably, the therapeutically effective dose is about 1 mg / kg (i.e., 70 mg) to about 25 mg / kg (i.e., 1.75 g) (for a 70 kg mammal).

The range of effective doses provided herein is intended to be representative of the preferred dosage range and is not intended to be limited thereto. However, the most preferred dosages will be adjusted according to the individual subject, as can be understood and measured by those skilled in the relevant art (see, for example, Berkow et al., Eds., The Merck Manual, 16th edition, Merck and Co., Rahway, NJ. , 1992; Goodman et al., Eds., Goodman and Gilman ' s Pharmacological Basis of Therapeutics, 10th 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. (1987); Ebadi, Pharmacology, Little, Brown and Co., Boston, (1985); Osolci al., Eds., Remington ' s Pharmaceutical Sciences, 18th edition, Mack Publishing Co., Easton, PA (1990); Katzung, Basic and Clinical Pharmacology, Appleton and Lange, Norwalk, CT (1992)].

The total dose required for each treatment may, as the case may be, be administered in multiple doses or single doses throughout the day. Certain preferred embodiments contemplate single application of topical formulations per day. In general, and in specific embodiments, the treatment can be initiated with less dose, less than the optimal dose of the compound. Thereafter, the capacity is increased in small increments until reaching the optimum effect under the circumstances.

Topical formulations may be administered alone or in conjunction with other treatments and / or agents directed against skin wounds or indicated for other related symptoms or etiological factors. For example, and as indicated above, topical formulations may additionally comprise retinoic acid. As another example, topical formulations may comprise one or more of the dermal tissue repair-promoting compounds described herein, or may comprise two or more such compounds having different cytotoxic activity.

The recipients of the topical formulations described herein may be any vertebrate animal, such as a mammal. Among mammals, preferred recipients include, but are not limited to, primates (including humans, apes and monkeys), supplements (including horses, goats, cows, sheep, pigs), rodents (including mice, rats, rabbits and hamsters), and carnivores ). Of the birds, the preferred recipients are turkeys, chickens and many other identical throats. The most preferred prisoner is a person, a person who has a bacterium-containing wound, or one or more acute or chronic wounds.

For topical application, an effective amount of a pharmaceutical composition comprising a BT compound antiseptic / wound-healing / anti-bacterial / dermal tissue repair-promoting compound according to embodiments described herein may be administered to a target site, for example, an acute or chronic wound , And / or a dangerous part of the skin (for example, a wound of the wound). The amount will generally range from about 0.0001 mg to about 1 g of a compound of the present invention per application, depending on the site to be treated, the severity of the wound (or past or considered surgical incision), and the nature of the topical vehicle used Lt; / RTI &gt; Preferred topical agents are ointments or sustained release pellets wherein from about 0.001 to about 50 mg of the active ingredient is used per ounce base or cc of the pellet suspension. The pharmaceutical composition may be formulated as a transdermal composition or transdermal delivery device (" patch &quot;). Such compositions include, for example, backings, active compound reservoirs, regulating membranes, liners and contact adhesives. These transdermal patches may be used to provide continuous beating, or, if desired, to deliver a compound of the present invention.

Certain embodiments of the compositions 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. Modulated 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 are provided in U.S. Patent Nos. 3,845,770 and 4,326,525 and PJ Kuzma et al, Regional Anesthesia 22 (6): 543-551 (1997), all of which are incorporated herein by reference &Lt; / RTI &gt;

The most appropriate route will depend on the nature and severity of the condition being treated. Those skilled in the art will also be able to determine other methods of administering a compound to a subject in need thereof, such as topical administration methods (spray, cream, open application, closed dressing, be used to.

Throughout the specification, unless the content requires otherwise, the words " comprise, " and " comprises " embrace the inclusion of a stated step or element or group of steps or elements, Or components or steps or groups of components. &Quot; comprising " means including, but not limited to, " consisting of " Thus, the phrase " consisting of " indicates that the listed components may be required or mandatory, and other components may not be present. &Quot; Essentially " means to include any ingredient listed after the phrase, and is limited to other ingredients that do not interfere with or contribute to the activity or action specified in the description of the ingredients listed. Thus, " consisting essentially of " as used herein, means that the listed ingredients are required or mandatory, but other ingredients are not required and may or may not be present depending on whether they affect the activity or action of the listed ingredients .

In the present specification and the appended claims, the singular forms " a, " an, " and " the " include plural references unless the content clearly dictates otherwise. As used herein, in particular aspects, the term "about" or "approximately" refers to values ± 5%, 6%, 7%, 8%, or 9% In another aspect, the term "about" or "approximately" indicates a range of values ± 10%, 11%, 12%, 13%, or 14% Approximately "indicates a range of ± 15%, 16%, 17%, 18%, 19%, or 20% when present in front of a number.

The following examples are presented for illustration and not limitation.

The bismuth-thiol composition according to the present invention comprising a plurality of microparticles comprising substantially all microparticles having a volume average diameter of from about 0.4 microns to about 5 microns comprises bismuth-thiol (BT) compounds according to the present invention exhibit biofilm formation, , Bacterial growth, bacterial cell viability or bacterial cell growth.

A brief description of some aspects of the drawing
Figure 1 shows the survival counts (log CFU) from colonies of Pseudomonas aeruginosa cultured for 24 hours at 37 ° C on 10% tryptic soy agar (TSA) Forming unit). The indicated antibiotic treatments were TOB, tobramycin 10X MIC; AMK, Amikacin 100X MIC; IPM, Imipenem 10X MIC; CEF, Cephalem 10X MIC; CIP, ciprofloxacin 100X MIC; Cpd 2B, compound 2B (bis-BAL, 1: 1.5). (MIC; minimum inhibitory concentration, e. G., Lowest concentration to prevent bacterial growth).
Figure 2 shows the survival counts (log CFU) from the Staphylococcus aureus colony bacterium subjected to the treatments after 24 hours of growth on 10% Trim Shin soybeans. The indicated antibiotic treatment was rifampicin, RIF 100X MIC; Daportomycin, DAP 320X MIC; Minocycline, MIN 100X MIC; Ampicillin, AMC 10X MIC; VANCOMAIN, VAN 10X MIC; Cpd 2B, Compound 2B (bis-BAL, 1: 1.5), Cpd 8-2, and Compound 8-2 (bis-Pyr / BDT (1: 1 / 0.5)).
Figure 3 shows scratch closure over time of keratinocytes exposed to the bacterium. (*) Was significantly different from the control group (P <0.001).
Figures 4A and 4B show the bacteriostatic BisEDT reverse antibiotic-resistance to several antibiotics. The effect of antibiotics is shown in the lawn of MRSA (methicillin-resistant S. aureus) in the presence and absence of BisEDT (0.05 [mu] g / ml). Panel A shows only standard antibiotic-immersed discs and Panel B shows discs combined with BisEDT (BE). [GM = gentamicin, CZ = cephazoline, FEP = cephalim, IPM = imipheine, SAM = ampicillin / sulphamant, LVX = levofloxacin.
Figure 5 shows the effect of BisEDT and antibiotics in the formation of bacteria. s. Epidermidis was grown in a polystyrene plate at 37 占 폚 for 48 hours in TSB + 2% Glucose. (GF), clindamycin (CM), minocycline (MC), gentamicin (GM), vancomycin (VM), cephazoline (CZ), napsilin (NC), and rifampicin (RP). The results are expressed as the mean change in BPC (series of twofold dilution steps) at 0.25 [mu] M BisEDT (n = 3).
Fig. 6 is a graph showing the results of S &lt; 1 &gt; The effect of BisEDT and antibiotics on the growth of epidermidis. Results are expressed as mean changes in MIC (dilution phase) with increasing BisEDT (n = 3). Refer to the legend in Figure 5 for antibiotic definitions.
Figure 7 shows bone and hardware samples from open fractures in vivo rat models after treatment with three BT formulations, bis-EDT, MB-11 and MB-8-2, with or without cephazoline antibiotic therapy. ). A bar graph showing the level of Aureus bacteria. The standard error of the mean is indicated by the error bars. Early euthanasia animals were not excluded from the analysis, but samples from one animal in group 2 were excluded due to significant contamination.

Example

Example  1: Preparation of BT compound

The following BT compounds were prepared according to the methods of Domenico et al. (See RE 37,793, US 6,248,371, US 6,086,921, US 6,380,248) &Lt; / RTI &gt; The atomic ratio to a single bismuth atom for comparison, based on the known tendency of bismuth to form a ternary complex with the sulfur-containing compound and the stoichiometric ratio of the reactants used. The numbers in parentheses are the ratios of bismuth to one (or more) thiols (for example, 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 (available from Bi Bi) (1: 1.5) BiC ₃ H ₆ S ₃

5) CPD 2A Bis-Bal (1: 1) BiC ₃ H ₆ S ₂ 0

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-Erry (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)

Fine particulate bismuth-1,2-ethanedithiol (bis-EDT, soluble bismuth formulation) was prepared as follows:

A solution of 0.331 L (~ 0.575 moles) of Bi (NO ₃ ) ₃ solution [43% Bi (NO ₃ ) ₃ (w) in room temperature was added to 5% aqueous HNO ₃ in excess of 11 L of carboy in 15 L of polypropylene carboy / w), 5% nitric acid (w / w), 52% water (w / w), product number 2362 from Shepherd Chemical Co., Cincinnati, OH;隆 to 1.6 g / mL] was added dropwise with stirring, and then anhydrous ethanol (4 L) was slowly added. Some formed white precipitate was dissolved by continuous stirring. Ethanolic solution (~ 1.56 L, ~ 0.55 M) of 1,2-ethanedithiol (CAS 540-63-6) was added to 1.5 L of absolute ethanol, 72.19 mL (0.863 moles) of 1,2-ethanedithiol Was added using a 60 mL syringe and then separately prepared by stirring for 5 minutes. Thereafter, the 1,2-ethanedithiol / EtOH the reagent was added slowly dropwise added with continuous stirring overnight, the _{Bi (NO 3) 3 / HNO} 3 aqueous solution over the course of 5 hours. The formed product was allowed to settle to a precipitate for about 15 minutes and the filtrate was then removed using a peristaltic pump at 300 mL / min. Thereafter, the product was collected by filtration on fine filter paper in a 15-cm diameter Buchner funnel and washed three times with 500-mL volumes of ethanol, USP water, and acetone, respectively, to yield a yellow amorphous powder solid BisEDT (694.51 gm / mole) was obtained. The product was placed in a 500 mL amber glass bottle and dried under high vacuum over CaCl ₂ for 48 hours. The recovered material (yield ~ 200 g) removed the thiol-characteristic odor. The crude product was redissolved in 750 mL of anhydrous ethanol, stirred for 30 min, then filtered, washed successively with 3 x 50 mL ethanol, 2 x 50 mL acetone, and again with 500 mL acetone. The re-washed powder was polished in 1M NaOH (500 mL), filtered and washed with 3 x 220 mL of water, 2 x 50 mL of ethanol, and 1 x 400 mL of acetone to give 156.74 gm of purified BisEDT Respectively. Subsequent batches produced essentially in the same manner yielded a yield of about 78 to 91%.

The product was characterized as having the structure shown above in formula I by data analysis from ¹ H and ³ C nuclear magnetic resonance (NMR), infrared spectroscopy (IR), ultraviolet spectroscopy (UV), mass spectrometry (MS) Respectively. HPLC method was developed to measure the chemical purity of the BisEDT in which the sample was prepared in DMSO (0.5 mg / mL). lambda _max was measured by scanning a solution of BisEDT in DMSO between 190 and 600 nm. 1 mL / min 265 nm at ambient temperature for isocratic HPLC eluted at (λ _max), 2μm using a UV detector for monitoring the injection volume, the YMC Pack PVC Sil NP, 5μm, 250X4.6 mm internal diameter analytical column (Waters ) Was carried out in a mobile phase of 0.1% formic acid in acetonitrile: water (9: 1) on a Model 2695 chromatograph equipped with a Waters (Waters, Milford, A single peak reflecting chemical purity was detected. The elemental analysis was consistent with the structural formula (I).

The dried particulate material was characterized to evaluate particle size characteristics. Briefly, the microparticles were resuspended in 2% Pluronic ^® F-68 (BASF, Mount Olive, NJ) and the suspension was poured into a Nanosizer for 10 minutes in a water bath sonicator, / Zetasizer Nano-S Particle Analyzer [Model ZEN 1600 (Zeta-Dislocation Measuring Capability), Malvern Instruments, Worcestershire, UK] Lt; / RTI &gt; From the edited data of the two measurements, the microparticles exhibited a single phase distribution with all detectable events between about 0.6 microns and 4 microns in volume mean diameter (VMD) and a peak VMD of about 1.3 microns. In contrast, when BisEDT was prepared according to the preceding method (see Domenico et al., 1997 Antimicrob. Agents Chemother. 41 (8): 1697-1703), most of the particles were heterodispersed, Which were significantly larger, excluding their characterization.

Example  2: chronic wound infection Colony (colony) Membrane  Model

Suppression by BT compound

Since bacteria present in chronic wounds adopt a bacterial lifestyle, BT has been shown to be effective as a method of treatment (Anderl et al., 2003 Antimicrob Agents Chemother 47: 125-56; Walters et al., 2003 Antimicrob Agents Chemother 47: 317; Wentland et al., 1996 Biotchnol. Prog. 12: 316; Zheng et al., 2002 Antimicrob Agents Chemother 46: 900], the bacterial membrane was tested for its effect on bacterial cell viability.

Briefly, the colonic bacteria membrane was grown for 24 hours on 10% tryptic soy agar and transferred to a Mueller Hinton plate containing the treatment agent. After treatment, the bacterium was dispersed in a number of peptones containing 2% w / v glutathione (neutralizing BT) and serially diluted into a number of peptones before spotting on plates for counting. Two bacteria isolated from chronic wounds were used separately for the production of test colonies. These were Pseudomonas aeruginosa, a gram-negative bacterial strain, and Staphylococcus aureus (MRSA), a gram-positive methicillin-resistant Staphylococcus aureus (MRSA).

Bacterial bacterial colonies were determined by the method of Anderl et al., 2003 Antimicrob Agents Chemother 47: 1251-56; Walters et al. , &Lt; / RTI &gt; 2003 Antimicrob Agents Chemother 47: 317; Wentland et al., 1996 Biotchnol. Prog. 12: 316; Zheng et al., 2002 Antimicrob Agents Chemother 46: 900]. Colonic bacteria membranes exhibited many familiar features of different membrane models, for example, they consisted of cells densely aggregated within a highly hydrated matrix. Other references (Brown et al., J Surg Res 56: 562; Millward et al, 1989 Microbios 58: 155; Sutch et al., 1995 J Pharm Pharmacol 47: 1094; As reported in Thrower et al., 1997 J Med Microbiol 46: 425, it has been observed that bacteria in the colon bacterium exhibited equally greatly reduced anti-microbial susceptibility, quantified in a more refined manner in the in vitro bacterial reactor . Colony-forming bacteria were easily and reproducibly produced in large numbers. According to a non-limiting theory, the colony-membrane model shared some of the characteristics of infected wounds; Bacteria grew at the interface with the nutrients supplied from the submerged and minimal fluid flow. A variety of nutrient sources were used to cultivate a colony bacterium containing blood agar that is believed to mimic in vivo nutritional conditions.

The colony bacterium was prepared by inoculating a spot of 5 μl of planktonic bacterial liquid culture into a 25 mm diameter polycarbonate filter membrane. Membranes were sterilized by exposure to ultraviolet light for 10 minutes per side before inoculation. The inoculum was grown overnight in bacterial medium at 37 ° C and diluted to an optical density of 0.1 at 600 nm in fresh medium before deposition on the membrane. Thereafter, the membrane was placed on the agar plate containing the growth medium. Thereafter, the plate was covered and placed inverted in a thermostat at 37 &lt; 0 &gt; C. Every 24 hours, membranes and colonic bacteria membranes were transferred to fresh plates using sterile forceps. The membranes were typically used for experiments after 48 h of growth, and there were about 10 ⁹ bacteria per membrane at that time. A wide range of single species and mixed species bacterium were cultured using the colony bacterium method successfully.

In order to determine the sensitivity to an antimicrobial agent (for example, a BT compound containing a combination of an antibiotic and a combination of a BT compound and an antibiotic), a colony bacterium membrane is immersed in a solution containing the antifungal antimicrobial agent (s) Plate. When the exposure period for antimicrobial treatment was more than 24 hours, the colon bacilli membrane was transferred daily to fresh treatment plates. At the end of the treatment period, the colonic bacteria membrane was placed in a tube containing 10 ml of buffer solution and vortexed for 1 to 2 minutes to disperse the bacteria membrane. In some cases, it was essential to simply process the sample with a tissue homogenizer to destroy cell aggregates. The resulting cell suspension is then serially diluted and plated to list the surviving bacteria and report it as colony forming units (CFU) per unit area. Survival data were analyzed using the log ₁₀ log transformed (log ₁₀ transformation).

Five antibiotics and thirteen BT compounds were tested for each type of bacterial culture colony (Shumomonasueruginosa, PA; methicillin resistant Staphylococcus aureus, MRSA or SA). The antimicrobial agents tested for PA are herein referred to as BisEDT and compounds 2B, 4, 5, 6, 8-2, 9, 10, 11 and 15 (see Table 1), and the antibiotics tobramycin, amikacin, Pem, cipazoline, and BT referred to as ciprofloxacin. The antimicrobial agents tested for SA include BT referred to herein as BisEDT and compounds 2B, 4, 5, 6, 8-2, 9, 10 and 11 (see Table 1) and antibiotics, rifampicin, Minocycline, ampicillin and vancomycin. The antibiotics were tested at a concentration of about 10 to 400 times the minimum inhibitory concentration (MIC) according to established microbiological methods, as described under "Brief Description of the Drawings" above.

Seven BT compounds showed significant effects on PA bacterial viability at the tested concentrations, and two BT compounds demonstrated a significant effect on MRSA survival at the tested concentrations; Representative results demonstrating the BT effect on bacterial viability are shown in Figure 1 for BisEDT and BT compound 2B (tested for PA) and in Figure 2 for BT compounds 2B and 8-2 (tested for SA) In both cases, the effect of the indicated antibiotics was shown in comparison. As also shown in FIGS. 1 and 2, inclusion of the BT compound shown with the indicated antibiotics produced a synergistic effect, whereby the decreased bacterial viability was improved compared to the anti-bacterial effect of the antibiotic alone or the BT compound alone. In the PA survival assay, compound 15 [bis-EDT / 2 hydroxy, propanethiol (1: 1/1)] was combined at a concentration of 80 μg / mL with 1600 μg / mL AMK and 80 μg / mL BisEDT (Not shown), which is comparable to the effect obtained by the use of the present invention (Fig. 1).

Example  3: chronic wound infection Dot  Drip flow model

Suppression by BT compound

Dotted flow-through membranes represent an art-accepted authentication model for forming and testing the effect of post-treasure anti-bacterial compounds on bacterial membranes. The dotted fluid film is produced in a coupon (substrate) placed in the channel of the drip flow reactor. Many different types of materials can be used as substrates for bacterial membrane formation, including translucent glass microscope slides. The nutrient liquid medium is introduced into the drip flow reactor cell chamber by dripping into the chamber near the top, followed by a tilt of 10 degrees to flow along the length of the coupon.

The bacterium grows in the point-of-view flow bioreactor and can be used for BT compounds either individually or together, and / or for antibiotic compounds individually or together with other antibiotics including BT compounds, or other conventional or candidate treatment of chronic wounds . Thus, BT compounds are characterized for their effect on bacterial bacteria in dot-flow reactors. A dot-flow reactor internal membrane is prepared according to established methods (eg, Stewart et al., 2001 J Appl Microbiol. 91: 525; Xu et al., 1998 Appl. Environ Microbiol. 64: 4035). The design involves culturing a germ layer on an inclined polystyrene coupon in a covered chamber. Exemplary culture media consisted of 1 g / l (as in chronic wounds) supplemented with 5% v / v adult donor bovine serum (pH 6.8), replicating in vitro, conditions rich in serum proteins similar to bacterial growth conditions in vivo Of glucose, 0.5 g / l NH ₄ NO ₃ , 0.25 g / l KCl, 0.25 g / l KH ₂ PO ₄ and 0.25 g / l MgS0 ₄ -7H ₂ O. The medium was flowed in dropwise fashion (50 ml / h) over four coupons contained in four separate parallel chambers each measuring a depth of 10 cm x 1.9 cm x 1.9 cm. The chambered reactor is made from polysulfone plastic. Each chamber is equipped with an individual removable plastic lid that can be tightly sealed. The bacterial reactor is contained in a thermostat at 37 ° C and the bacterial cell culture medium is warmed by passing it through an aluminum heat sink maintained in a thermostat. The method regenerates the antibiotic resistance phenotype observed in a specific bacterium and provides continuous supplementation of nutrients while simulating the proximity of the hyphe fluid shear environment and the chronic wounds to the air interface characteristics, And is compatible with a number of analytical methods for characterization and monitoring. A wide range of pure and mixed-species bacterial membranes were cultured using a dot-flow reactor successfully. The membrane is typically grown for 2 to 5 days before application of the antimicrobial agent.

In order to determine the effect of the anti-fungal agent on the membrane grown in the dot-flow reactor, the flow of the emulsion that passes over the membrane is compared with the desired therapeutic formulation (e. G., One or more BT compounds and / or one or more antibiotics, Control, and / or other retention agent). The flow is maintained for the prescribed treatment period. Thereafter, the treated bacterium coupon is simply removed from the reactor and the bacterium is scraped into a beaker containing 10 ml of buffer. The sample is simply processed using a tissue homogenizer (typically 30 seconds to 1 minute) to disperse bacterial aggregates. The suspension is serially diluted and plated to list viable microorganisms according to standard microbiological methods.

Example  4: wound of keratinocyte scratch recovery Membrane  control

Membrane inhibition by BT compound

This example demonstrates an established in vitro keratinocyte scratching model of wound healing to reach a bacterium-related wound pathology and wound healing, and in particular to reach a bacterium associated with an acute or chronic wound or wound containing a bacterium membrane as described herein Lt; / RTI &gt; According to the keratinocyte scratching model of the effect of chronic scarring, the cultures of mammalian (e. G., Human) keratinocytes and germ bacterial populations are advanced in separate chambers in contact with each other to produce keratinocyte wound healing , The conditions affecting the effect, and the effect of the soluble components elaborately generated by the membrane.

The neonatal human epidermal cells are cultured as a monolayer in a treated plastic dish where the " wound " or scratching in the monolayer is measured by mechanical means (e.g., sterilized scalpel, razor, cell scraper through physical destruction of the monolayer, such as by scraping essentially linear cell-free zones between areas of the monolayer using suitable tools such as scraper, forceps or other tools). The in vitro keratinocyte cell model system is known to simulate in vivo wound healing and undergoes cellular structural and functional processes in response to scarring phenomena. In accordance with an embodiment described herein, the effect of the presence of bacterial membranes in such processes, e. G., The time of healing of the scratches is observed, and in this and related aspects, the effect is also selected after the selected antimicrobial agent Bacteria and antimicrobial membrane) therapy.

The wounded keratinocyte monolayer cultured in the presence of a bacterium is tested according to morphological, biochemical, molecular genetic, cellular physiological and other parameters to determine whether the introduction of the BT compound alters the damage effect of the bacterium For example, increase or decrease in a statistically significant manner for a suitable control). The wound was first exposed to each of the BT compound alone, and to the considered combination of BT compounds, and the toxicity of each BT compound treatment was tested prior to assessing the effect of this treatment on the membrane wound healing process for the model wound healing process.

In a representative embodiment, a 3-day bacterium membrane is placed on a membrane (e.g., a TransWell membrane insert, etc.) maintained in the tissue culture well and through a scratched keratinocyte layer to initiate a wound healing process In fluid communication. Bacterial membranes cultured outside certified acute or chronic wounds are considered for use in these and related embodiments.

Therefore, an in vitro system was developed to evaluate the effect of soluble bacterial components on migration and proliferation of human keratinocytes. The system uses a dialysis membrane to separate the bacterium from the keratinocytes. The keratinocytes were cultured from neonatal epidermis as previously described (Fleckman et al., 1997 J Invest. Dermatol. 109: 36; Piepkorn et al., 1987 J Invest. Dermatol. 88: 215-219) And grown as a conformable monolayer on slip. Afterwards, scratched keratinocytes to obtain a " wound " with a uniform width, then the cell repair process can be monitored (e.g. 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). Subsequently, the artificial wound is placed on the bottom of a sterile double-sided chamber and the chamber is assembled using aseptic technique. Both sides of the chamber were filled with keratinocyte growth medium (EpiLife) in the presence or absence of antibiotics and / or bismuth-thiols. The uninoculated system was used as a control.

The system is inoculated with the bacteria isolated from the wound and incubated for 2 hours at rest to allow the bacteria to adhere to the surface of the upper chamber. After the attachment period, liquid medium flow is initiated in the upper chamber to remove unattached cells. Thereafter, the flow of the medium is maintained by washing of unattached cells at a rate that minimizes the growth of planktonic cells in the upper chamber. After a period of incubation ranging from 6 to 48 hours, the system (keratinocytes on the cover slip and bacterial bacteria on the membrane substrate) is disassembled and the cover slip is removed and analyzed. In a related embodiment, mature membranes are grown in the upper chamber before assembling the chamber. In another related embodiment, separate co-culture of the scar-scarred wound keratocyte monolayer with or without one or more BT compounds, optionally with or without one or more antibiotics, is carried out to produce keratinocyte recovery A BT compound, such as BT provided in particulate form, and a combination of one or more amikacin, ampicillin, or a pharmaceutically acceptable salt thereof, in a sustained release BT compound-and-antibiotic combination Sepharose sleepy, three pepim, chloramphenicol, ciprofloxacin, clean the myth (or other Linkoping four mid antibiotic), the neoplasm? azithromycin (Cubicin ^®), doxycycline, gatifloxacin, gentamicin myth, already fetched s, levofloxacin, linezolid (Zyvox ^®), minocycline, and naphthyl published, our wave erythromycin, rifampin, measure the effectiveness of sulfamic methoxy Sasol, tobramycin, and vancomycin], for example, be I have used a formulation that alters (e.g., increases or decreases in a statistically significant manner as compared to a suitable control) at least one index of scratch wound healing, such as the time elapsed for wound healing or other wound- Or combinations of agents (see, for example, Tao et al., 2007 PLoS ONE 2: e697; Buth et al. 2007 Eur. J Cell Biol. 86: 747; Phan et al. 2000 Ann. Singapore 29:27).

Example  5: scratching wound of keratinocytes Membrane  control

Isolated human keratinocytes were cultured on a glass cover slip and scratch-scarred according to the method described above in Example 4. Wounded cultures were maintained in the presence of co-cultured bacterium on membrane supports in fluid communication with keratinocyte cultures, or under single culture conditions. Scratch seal time intervals were measured while keratinocyte cell growth and / or migration was reestablishing keratinocytes on the scratch area. Figure 3 illustrates the effect of the presence of fluid communication (but not direct contact) of the bacterium on the healing time of scratched keratinocytes.

Thus, in certain embodiments, scratch-scarred cells (e. G. Keratinocytes or fibroblasts) monolayers are cultured in the presence of a bacterium microfilm in the presence or absence of a post-retention anti-bacterial membrane; There is contemplated a method of identifying an agent for treating chronic wounds, including assessing the index of healing of scratched-wounded cell monolayers in the absence and presence of a post-treasure anti-fungal agent, wherein at least one index of healing agents which promote [for example, a substantially monodisperse as described herein BT particle suspension alone or amikacin, ampicillin, Sefar sleepy, three pepim, chloramphenicol, ciprofloxacin, clean the myth, the neoplasm? azithromycin (Cubicin ^® ), doxycycline, gatifloxacin, gentamicin myth, already fetched s, levofloxacin, one or more of linezolid (Zyvox ^®), minocycline, naphthyl cylinder, wave our erythromycin, rifampin, sulfamic methoxy Sasol, tobramycin, and vancomycin; and Ascending combination with the same antibiotic] is identified as an agent suitable for treating acute or chronic scars or wounds containing a germ membrane.

Example  6: synergistic bismuth- Thiol  (BT) - Antibiotics Combination

This example demonstrates the proven synergistic effect of a combination of one or more bismuth-thiol compounds and one or more antibiotics against various bacterial species and bacterial strains including several antibiotic-resistant bacteria.

Materials and methods. Sensitivity studies were performed by broth dilution in a 96-well tissue culture plate (Nalge Nunc International, Denmark) using the NCCLS protocol (see 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).

Briefly, a 0.5 McFarland standard suspension was prepared using overnight bacterial cultures and seeded on a cation-regulated Müller-Hinton broth medium (BBL, Cockeysville, MD, USA) (2 x 10 &lt; ⁶ &gt; cfu / mL). BT (prepared as described above) and antibiotics were added at an incremental concentration while maintaining a fixed final volume of 0.2 mL. The cultures were incubated for 24 hours at 37 ° C and turbidity was assessed by absorption at 630 nm using an ELISA plate reader (Biotek Instruments, Winslow, Vt., USA). The minimum inhibitory concentration (MIC) was expressed as the lowest drug concentration that inhibited growth for 24 hours. The number of surviving bacteria (cfu / mL) was determined by standard plating on nutrient agar. The minimum bacterial concentration (MBC) was expressed as the concentration of drug that decreased initial viability up to 99.9% at 24 hours of incubation.

The activity of the antimicrobial combination was assessed using the checkerboard method. Fractional inhibitory concentration index (FICI) and fractionated bacterial concentration index (FBCI) were determined by Eliopoulos et al. (Eliopoulos and Moellering, (1996) Antimicrobial combinations. In Antibiotics in Laboratory Medicine (Lorian, V., Ed.), Pp. 330-96, Williams and Wilkins, Baltimore, MD, USA). The synergistic effect was defined as FICI or FBCI index of ≤ 0.5, no interaction at> 0.5 to 4 and antagonism at> 4 (Odds, FC (2003) Synergy, antagonism, and what the chequerboard puts between them. Journal of Antimicrobial Chemotherapy 52: 1]. Synergism was also defined as ≥4-fold reduction in antibiotic concentration.

The results are shown in Tables 2 to 17.

Example  7: Gram-positive and Gram-negative, including antibiotic-resistant bacterial strains Compared bismuth for bacteria - Thiol (BT) and antibiotic effect

Bacterial strain

In this example, the in vitro activity of BisEDT and comparative agents was evaluated against a number of clinical isolates of Gram-positive and Gram-negative bacteria involved in skin and soft tissue infections.

Materials and methods. Test compounds and test concentration ranges were as follows: BisEDT (Domenico et al., 1997; Domenico et al., Antimicrob. Agents Chemother. 45 (5): 1417-1421. And Example 1], 16-0.015 [mu] g / mL; Linezolid [ChemPacifica Inc., # 35710], 64-0.06 [mu] g / mL; 0.0 &gt; ug / mL &lt; / RTI &gt; and 16-0.015 pg / mL; &lt; RTI ID = 0.0 &gt; Cubist &lt; / RTI &gt; Vancomycin (Sigma-Aldrich, # L2002, St. Louis, Mo.), 64-0.06 ug / mL; Ceftazidime (Sigma # C3809), 64-0.06 [mu] g / mL and 32-0.03 [mu] g / mL; 16-0.015 占 퐂 / mL and 8-0.008 占 퐂 / mL of Imipenem [United States Pharmacopeia, NJ, # 1337809]; Ciprofloxacin (United States State Pharmacopoeia, # IOC265), 32 - 0.03 ug / mL and 4 - 0.004 / / mL; 32 - 0.03 / / mL and 16 - 0.015 / / mL of gentamicin (Sigma # G3632). All test products except gentamicin were dissolved in DMSO; Gentamicin was dissolved in water. The stock solution was prepared in a test plate at a 40-fold maximum concentration. The final concentration of DMSO in the test system was 2.5%.

organism. The test organisms were obtained from the following clinical laboratories: CHP, Clarian Health Partners, Indianapolis, Ind .; UCLA, University of California Los Angeles Medical Center, CA, Los Angeles, CA; GR Micro, London, England; PHRI TB Center, Public Health Research Institute Tuberculosis Center, New York, NY; ATCC, American Type Culture Collection, Manassas, Va .; Mt Sinai Hosp., Mount Sinai Hospital, New York, NY; UCSF, University of California San Francisco General Hospital, San Francisco, California; Branson Hospital, Branson Methodist Hospital, Kalamazoo, Michigan; Quality control isolates were obtained from the American Type Culture Collection (ATCC, Manassas, Va.). The organisms were streaked to separate on appropriate agar medium for each organism. The colonies were picked up from the separating plate with a cotton swab and placed into the supernatant in a suitable broth containing the cryoprotectant. The suspension was aliquoted into cryogenic vials and kept at -80 占 폚.

The abbreviations are: BisEDT, bismuth-1,2-ethanedithiol; LZD, linezolid; DAP, dapotomycin; VA, vancomycin; CAZ, ceftazidime; IPM, Imipenem; CIP, ciprofloxacin; GM, gentamicin; MSSA, methicillin-sensitive Staphylococcus aureus; CLSI QC, Clinical and Laboratory Standards Agency Quality control strain; MRSA, methicillin-resistant Staphylococcus aureus; CA-MRSA, co-acquired methicillin-resistant Staphylococcus aureus; MSSE, methicillin-sensitive staphylococcus epidermidis; MRSE, methicillin-resistant Staphylococcus epidermidis; VSE, vancomycin-sensitive enterococcus.

Isolates from frozen vials are grown in a suitable medium: trypticase soybean agar for most organisms (Becton-Dickinson, Sparks, MD) or trypticase soybean agar for streptococcus and 5 Sheep blood [manufacturer: Cleveland Scientific, Baths, Ohio]. Plates were incubated at 35 &lt; 0 &gt; C overnight. Quality control organisms were included. The medium used for the MIC assay was Mueller Hinton II Broth (MHB II - manufacturer: Becton Dickinson, # 212322) for most of the organisms: 2% degraded horse blood in MHB II (Cleveland Scientific Lot # H13913) was supplemented to give growth to Streptococcus pieogenes and Streptococcus agalactia. The medium was prepared at 102.5% normal weight and added to each well of a microdilution panel The dilution produced by addition of 5 μL of the drug solution was counteracted. Further, for the test with the adjuvant, the medium was supplemented with an additional 25 mg / L Ca ²⁺ .

The MIC assay method is followed by following the procedure described in the Clinical and Laboratory Standards Institute (see Clinical and Laboratory Standards Institute, Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria That Grow Aerobically; Approved Standard- Using Automated Liquid Handlers, the use of an automated liquid handler can be used to perform the &lt; RTI ID = 0.0 &gt; M7-A7 &lt; / RTI &gt; Serial dilution and liquid transfer were performed. Automated liquid handlers are available from Multidrop 384 (Labsystems, Helsinki, Finland), Biomek 2000 and Multimek 96 [Beckman Coulter, California Main puller tone material]. Wells of columns 2 to 12 of a standard 96-well microdilution plate (Falcon 3918) were filled with 150 [mu] L of DMSO or ZentamidCredit water on 384 multi-drop. The drug (300 [mu] L) was dispersed in column 1 of the appropriate column in the plate.

These come from the mother plate on which the test plate (daughter plate) is made. Biomec 2000 was delivered via column 11 in series delivery in the mother plate. The wells of column 12 contained no drug and were organic growth control wells in the daughter plate. The daughter plate was loaded with 185 μL of Multidrop 384 in an appropriate test medium (as described above). The daughter plate was prepared on a MultiMeg 96 device that transferred 5 μL of the drug solution from each well of the parent plate to each corresponding well of each daughter plate in one step.

Standardized inoculations of each organism were prepared per CLSI method (ISBN 1-56238-587-9, cited above). The suspension was prepared in the same manner as the turbidity of the 0.5 McFarland standard in MHB. The suspension was diluted 1: 9 in an appropriate broth for the organism. The inoculum for each organism was dispensed into length-divided sterile reservoirs (Beckman Coulter) and inoculated onto plates using Biomek 2000. The daughter plate was placed on the inverted Biomech 2000 work surface so that the inoculation consisted of low concentration and high concentration of drug. Biomec 2000 delivered 10 μL of the standardized inoculum into each well. This is about 5 x 10 ⁵ colonies were obtained, the daughter plates in a final cell density of the forming units / mL. Thus, the wells of the daughter plate ultimately contained 185 μL of broth, 5 μL of drug solution, and 10 μL of bacterial inoculum. Plates were stacked at three elevations, covered on top plate, placed in a plastic bag and incubated at 35 ° C for about 18 hours for most of the separates. Streptococcus plates were incubated for 20 hours and then read. The microplate was observed from the bottom using a plate observer. For each test medium, a non-inoculated soluble control plate was observed for evidence of drug precipitation. The MIC was read and recorded as the lowest concentration of drug that inhibited the visible growth of the organism.

result. All commercially available drugs were soluble both at the test concentrations in both media. BisEDT showed a small amount of precipitate at 32 ug / mL, but MIC readings were not affected as the inhibitory concentrations for all the organisms tested were below that concentration. On each assay day, the appropriate quality control strain (s) were included in the MIC assay. The MIC values derived for these strains are suitably determined by the quality control range published for each formulation (see CLSI document M100-S18 [ISBN 1 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 (s) were included in the MIC assay. The MIC values derived from these strains are suitably determined by the published quality control range for each formulation (see CLSI document M100-S18 [ISBN 1- 56238-653-0]). Of the 141 values for the quality control strains for which the quality control range was announced, 140 (99.3%) were within the defined range. One exception is S. Imipenem for Aureus 29213, which yielded one value (≤ 0.008 ug / mL) for a single run, which is one dilution below the published QC range. In this performance all other quality control results were within the defined quality control range.

BisEDT is a methicillin-sensitive Staphylococcus aureus (MSSA), methicillin-resistant S. (MRSA), and community-acquired MRSA (CA-MRSA), with a MIC90 value of 0.5 [mu] g / mL for all three organism groups of 1 [mu] g / mL and inhibited all strains tested. BisEDT was greater than the activity of linezolid and vancomycin and showed the same activity as that of dapotomycin. Imipenem was stronger than BisEDT for MSSA (MIC90 = 0.03 [mu] g / mL). However, MRSA and CAMRSA were resistant to imipenem, but BisEDT demonstrated the same activity as shown for MSSA. BisEDT was highly active, with values of 0.12 and 0.25 [mu] g / mL for methicillin-sensitive and methicillin-resistant Staphylococcus epidermidis (MSSE and MRSE), respectively. BisEDT was more active for MSSE than any of the other agents tested except imipenem. BisEDT was the most active agent (maximum active agent) tested for MRSE.

BisEDT demonstrated the same activity as that of dapotomycin, vancomycin and imipenem with a MIC90 value of 2 [mu] g / mL for vancomycin-sensitive Enterococcus faecalis (VSEfc). Significantly, BisEDT was the maximal active agent tested for vancomycin-resistant Enterococcus faecalis (VREfc) with a MIC90 value of 1 ug / mL.

BisEDT was highly active at an MIC90 value of 2 ug / mL for vancomycin-sensitive enterococcus faecium (VSEfm); Its activity was the same or similar to that of dapotomycin and was 1-fold higher than that of vancomycin. BisEDT and linezolid were the largest active agents tested for vancomycin-resistant Enterococcus faecium (VREfm), each demonstrating a MIC90 value of 2 ug / mL. The activity of BisEDT against Streptococcus piegensis (MIC90 value at 0.5 [mu] g / mL) was equal to that of vancomycin, greater than that of linzolide, slightly less than that of dapotomycin and ceftazidime. Compounds inhibited all tested strains below 0.5 ㎍ / mL. In these studies, the least susceptible species to BisEDT was Streptococcus agalactiae, where the observed MIC90 value was 16 ug / mL. BisEDT was less active than all of the tested agents except gentamicin.

The activity of BisEDT and comparative factors on the Gram-negative bacteria involved was evidenced by the BisEDT efficacy (MIC90 value of 2 ug / mL) versus the Acinetobacter baumannii, making it the most active compound tested by BisEDT. The elevated MIC of the significant number of test isolates for comparative agent formulations produced an off-scale MIC90 value for these agents. BisEDT was a potent inhibitor of Escherichia coli that inhibited all strains at less than 2 μg / mL (MIC90 = 2 ug / mL). The compound was less active than Imipenem, but more active than ceftazidime, ciprofloxacin, and gentamicin. BisEDT also demonstrated activity against Klebsiella pneumoniae with an MIC90 value of 8 [mu] g / mL which is the same as that of Imipenem. Relatively high MIC90 values were inhibited by imipenem, ceftazidime, ciprofloxacin, and gentamicin, indicating that this was a highly antibiotic-resistant group of organisms. BisEDT was the most active compound tested for Pseudomonas aeruginosa at an MIC90 value of 4 ug / mL. There was a high level of tolerance to the comparative agent formulations for this group of test isolates.

In summary, BisEDT has demonstrated broad spectrum efficacy for a number of clinical isolates representing a number of species, including species commonly involved in acute and chronic skin and skin structure infections in humans. The activity of BisEDT and the major comparator formulations was evaluated for 723 clinical isolates of Gram-positive and Gram-negative bacteria. BT compounds have demonstrated broad spectrum activity, and for many test organisms in the study, BisEDT was the most active compound tested in terms of anti-bacterial activity. BisEDT can be used for MSSA, MRSA, CA-MRSA, MSSE, MRSE, 0.0 &gt; MIC90 &lt; / RTI &gt; value was less than 0.5 [mu] g / mL. Strong active is also VSEfc, VREfc.VSEfm, VREfm, AA. Bau Maney, Lee. Coli, and blood. Which has been demonstrated for agar ginseng, wherein the MIC90 value was in the range of 1 to 4 [mu] g / mL. The observed MIC90 values are shown in Fig. (MIC90 = 8 [mu] g / mL), and S. In the case of S. agalactiae (MIC90 = 16 / / mL).

Example  8: Particulate BT-antibiotic enhancing and synergistic activity

This example demonstrates that the particulate bismuth thiol (BT) promotes antibiotic activity through improved and / or synergistic interactions.

The main complex factor in treating infections is the emergence of bacterial resistance to antibiotics. s. Epidermidis (MRSE) and S. In aureus (MRSA), methicillin resistance actually reflects multiple drug resistance and makes it very difficult to eradicate these pathogens. However, no staphylococcus from hundreds of strains tested showed resistance to BT. In addition, at subMIC concentrations, BT reduced resistance to several important antibiotics.

Staphylococcus Aureus . A graphical description of the antibiotic-re-sensitizing effect of subMIC bismuth ethanedithiol (BisEDT) against MRSA is provided (Figure 4), including gentamicin, cephazoline, cephapim, imiphecin, sulfamethoxazole and levofloxacin Of the antibiotics. Thus, BisEDT improves the activity of the best antibiotics nonspecifically.

Broth dilution antimicrobial susceptibility studies were performed on 12 MRSA strains using several antibiotics combined with subMIC-level BisEDT (Table 18). Both the bacterial-protective concentration (BPC) and the minimal inhibitory concentration (MIC) were measured in a special bacterium culture medium (BHIG / X). MIC and BPC for gentamicin and cephazoline were reduced by subMIC BisEDT (BisEDT MIC, 0.2-0.4 [mu] g / ml), but not below the breakpoint for sensitivity. subMIC BisEDT improved the susceptibility of MRSA to gatifloxacin and cephalim close to the breakpoint for sensitivity. These strains were already susceptible to vancomycin, but much more so in the presence of subMIC BisEDT. In general, MIC and BPC were reduced by 2 to 5 fold with subMIC BisEDT.

Twelve MRSA clinical isolates were grown in BHIG / X and exposed to serial dilutions of antibiotics in the presence of 0 to 0.1 μg / ml of BisEDT. The MIC and BPC, calculated as [mu] g / ml, are the mean ± standard deviation from at least three tests. The right column lists the standard MIC for antibiotic sensitivity (S) and resistance (R).

A broth dilution study of the ceftepim-resistant MRSA isolate is shown in Table 19. At 0.1 ㎍ / ml, BisEDT significantly enhanced the inhibitory activity of ceftepim in 11 out of 12 isolates. In this particular study, the data showed synergism between BisEDT and cefepime (FIC <0.5), along with many isolates at breakpoints for sensitivity.

Twelve cephapym-resistant MRSA were tested in a BHIG / X medium in polystyrene plates at 37 占 폚 for 48 hours for susceptibility to cePePIM combined with subMIC BisEDT.

The results for the combination studies using napsililin or gentamicin are shown in Table 20. &lt; tb &gt; &lt; TABLE &gt; When combined with nafcillin, BisEDT (0.2 μg / ml) reduced MIC90 to nafcillin by 4-fold over the MRSA (FIC, 0.74). When combined with gentamicin, BisEDT reduced the MIC90 to gentamicin by more than 10-fold relative to MRSA (FIC, 0.6). BT reversed the resistance of all four gentamycin-resistant isolates tested for clinically relevant concentrations (Domenico et al., 2002). The MICs for these antimicrobials were substantially reduced, especially for gentamicin. The broth used in these studies was tryptic soy broth (TSB) containing 2% glucose, which was similar to that observed in the Müller-Hinton II broth enhanced with 1% blood.

Staphylococcus Epidermidis . The activity of most classes of antibiotics was enhanced in the presence of BisEDT. In connection with BPC, clindamycin and gatifloxacin, when combined with BisEDT, And exhibited significantly more antibacterial membrane activity against epidermidis (Fig. 5). From a different point of view, BPCs for clinodimicin, gatifloxacin and gentamicin were reduced by 50, 10 and 4 fold, respectively, in the presence of subMIC BisEDT.

Only modest decreases in bactericidal concentration (BPC) were noted for minocycline, vancomycin and cephazoline, while rifampicin and nafcillin were unaffected by 0.05 ug / ml BisEDT. In 0.1 μg / ml BisEDT, no membrane was detected, regardless of the antibiotic used, indicating that no antagonism occurred. The concentration of the bisEDT was determined by the method described in S. (See, for example, Domenico et al., 2003) (see FIG. 5).

Regarding growth inhibition, seven of the eight antibiotics tested were S. Was significantly improved in the presence of 0.1 [mu] g / ml (0.5 [mu] M) BisEDT against epidermidis (Fig. 6). MIC changes were most prominent in the case of clinodimin and gentamicin, followed by vancomycin, cephazoline, minocycline, gatifloxacin and nafcillin, and rifampicin was not affected. Among the antibiotics, the strain was resistant to (NC, CZ, GM, CM) and only the cephazoline resistance was reversed to the clinically relevant level by BisEDT.

s. The minimum bacterial concentration (MBC) for most antibiotics tested against epidermidis was slightly reduced with subMIC BisEDT. Gentamicin exhibited the greatest reduction in MBC (4X to 16X), followed by cephazoline (4X to 5x), vancomycin and nafcillin (3X to 4X), minocycline and gatifloxacin To 3-fold). However, clinodimin and rifampicin MBC remained largely unaffected. Clindamycin is a bacteriostatic agent, which accounts for its deficiency in bacterial activity. Sepharolin resistance was reversed with respect to MBC (Domenico et al., 2003). The effect was additive.

The efficacy of the antimicrobial agent was also demonstrated in vivo in rat model of implant-infected rats (Table 21). BisEDT levels as low as 0.1 μg / ml are also resistant to S for 7 days. It was possible to promote the prevention of the epidermidis membrane.

As summarized in Table 21, implants implanted alone or in combination with 0.1 [mu] g / ml BisEDT, 10 [mu] g / ml RIP and 10 [mu] g / ml rifampin were subcutaneously transplanted into rats. Physiological solution (1 ml) containing MS and MR at 2x10 ⁷ cfu / ml was inoculated onto the graft surface using a tuberculin syringe. All grafts were explanted on the 7th day after transplantation and sonicated in sterile saline solution 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 S. Epidermidis; MR, methicillin-resistant S. Epidermidis

^b Dacron graft segment impregnated with 0.1 mg / l BT, 10 mg / l RIP, 10 mg / l rifampin,

^c Compared with the control group MS and MR, statistically significant

statistically significant when compared with the group ^d MS3

^e Compared with MR1, MR2, and MR3 groups, statistically significant

Gram-negative bacteria. The tobramycin activity against resistant Pseudomonas aeruginosa was improved several-fold by subMIC BisEDT (Table 22). In this trial, the MIC was defined in more detail as IC ₂₄ .

blood. Resistant strain of A. aeruginosa was cultured in the presence of tobramycin (NN) and BisEDT (BE; 0.33 / / ml) in a Mueller-Hillton II broth at 37 째 C. MIC was measured as antibiotic concentration inhibiting growth for 24 +/- 1 hour.

For tobramycin-resistant bursal isolate, 0.4 μg / ml BisEDT resulted in tobramycin sensitivity (mean FIC: 0.48) and seven-fold reduction in MIC ₉₀ (Table 7) . Both MIC and MBC of tobramycin were significantly reduced by subMIC BisEDT to a level achievable for 50 clinical isolates of Bruchelia sephacia (Veloira et al., 2003). BisEDT and tobramycin in the form of liposomes are p. (See Halwani et al., 2008; Halwani et al., 2009).

Chloramphenicol and ampicillin-resistant Escherichia coli were made sensitive to these drugs by the addition of subMIC BisEDT (Table 24).

this. Cola resistant strains were cultured in Mueller-Hinton II broth at 37 DEG C in the presence of chloramphenicol (CM) or ampicillin (AMP) and BisEDT alone or in combination (BE; 0.33 mu g / ml) Respectively. MIC was measured as antibiotic concentration inhibiting growth for 24 +/- 1 hour.

The tetracycline-resistant Escherichia coli subMIC BisEDT was added to make it resistant to doxycycline (Table 25). The combination exhibited a synergistic effect on the TET M and TET D strains (FIC ≤ 0.5) and showed additional effects on the TET A and TET B strains.

this. Cola resistant strains were cultured in the presence of doxycycline (DOX) and BisEDT alone or in combination (BE; 0.33 [mu] g / ml) at 37 DEG C in a Mueller-Hinton II broth. MIC was measured as antibiotic concentration inhibiting growth for 24 +/- 1 hour.

Reference literature:

Domenico P, R O'Leary, BA Cunha. 1992. Differential effect of bismuth and salicylate compounds on antibiotic sensitivity of Pseudomonas aeruginosa. Eur J Clin Microbiol Infec Dis 1 1: 170-175; Domenico P, D Parikh, BA Cunha. 1994. Bismuth modulation of antibiotic activity against gastrointestinal bacterial pathogens. Med Microbiol Lett 3: 1 14-119; Domenico P, Kazzaz JA, Davis JM, Niederman MS. 2002. Subinhibitory bismuth ethanthiol (BisEDT) sensitizes resistant Staphylococcus aureus to nafcillin or gentamicin. Annual Meeting, ASM, Salt Lake City, UT; Domenico P, Kazzaz JA, Davis JM. 2003. Combating antibiotic resistance with bismuth-thiols. Research Advances in Antimicrob Agents Chemother 3: 79-85; Domenico P, E Gurzenda, A Giacometti, O Cirioni, R Ghiselli, F Orlando, M Korem, V Saba, G Scalise, N Balaban. 2004. BisEDT and RIP act synergy to prevent graft infections by resistant staphylococci. Peptides 25: 2047-2053; Halwani M, Blomme S, Suntres ZE, Alipour M, Azghani AO, Kumara, Omri A. 2008. Liposomal bismuth-ethandithiol formulation enhances antimicrobial activity of tobramycin. Intl J Pharmaceut. 358: 278-84; Halwani M, Hebert S, Suntres ZE, Lafrenie RM, Azghani AO, Omri A. 2009. Bismuth-thiol incorporation enhancers biological activities of liposomal tobramycin against bacterial biofilm and quorum sensing molecules production by Pseudomonas aeruginosa. 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.

Example  9: Particulate BT-antibiotic enhancing and synergistic activity

This example demonstrates that the microparticulated bismuth thiol BisEDT promotes antibiotic activity through enhanced and / or synergistic interactions with specific antibiotics against a specific microbial target organism. Single-point data for each of the combinations shown in Table 26 was essentially generated according to the method used in Example 8.

SA, Staphylococcus aureus; MRSA, methicillin-resistant Staphylococcus aureus; E Fc, Enterococcus faecalis; SP, Streptococcus pneumoniae; PRSP, penicillin-resistant Streptococcus pneumoniae; EC, Escherichia coli; KP, Klebsiella pneumoniae; PA, Pseudomonas aeruginosa; Bcep, Bourcolieria sephacia; Bmult, Burkholderia Mt. Tiborance; Abau, Akhsineto Bertha Baumanny; Msmeg, Mycobacterium segmatis.

Example  10: Atomization  BT-antibiotic enhancement and synergistic activity

The effects of the microparticulated bis-EDT and the four bis-EDT analogs and other agents prepared as described above on representative strains of seven gram-negative pathogenic bacteria were tested. Using a variation of the usual experimental method, it is used as a synergistic (FICI ≤ 0.5), an enhancement (0.5 <FICI ≤ 1.0), antagonism (FICI> 4.0), FIC and FICI (1.0 < FICI &lt; = 4.0) (Eliopoulos G and R Moellering. 1991. Antimicrobial combination. In Antibiotics in Laboratory Medicine, Third Edition, edited by V Lorian. Williams and Wilkins, Baltimore, MD, pp. 432-492; Odds, 2003 J. Antimicrob. Chemother. 52 (1): 1]. The FIC index was measured using the checkerboard technique and used in the study.

Stock solutions of all test products were prepared at the final target concentration of 40X in an appropriate solvent. All test products were present in solution under these conditions. The final drug concentration in the FIC assay plate was set in parentheses for each test organism, as long as the strain was not resistant to the test agent as a whole. The concentration ranges tested are shown in Table 27. The test organism was provided from a clinical source, or from the American Type Culture Collection. Upon receipt, the isolate was streaked onto trypsin soy agar II (TSA). Colonies were collected from these plates and cell suspensions were prepared in a suitable broth growth medium containing a cryoprotectant. The aliquots were then frozen at -80 占 폚. Frozen seeds of the organisms to be tested in the provided assay were thawed and streaked for isolation on TSA plates and incubated at 35 ° C. All organisms were tested in a Mueller Hinton II broth (Becton Dickinson, Lot # 9044411). Broth was prepared at 1.05X normal weight / volume to offset 5 vol% of the drug in the final test plate.

The minimum inhibitory concentration (MIC) value was previously measured using Broth microdilution for aerobic bacteria (Clinical and Laboratory Standards Institute (CLSI). Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria That Grow Aerobically; Approved Standard-Eighth Edition. CLSI document M07-A8 [ISBN 1-56238-689-1]. Clinical and Laboratory Standards Institute, 940 West Valley Road, Suite 1400, Wayne, Pennsylvania 19087-1898 USA, 2009].

FIC values were determined using the broth microdilution method described above (Sweeney et al., 2003 Antimicrob. Agents Chemother. 47 (6): 1902-1906). To prepare the test plates, an automated liquid handler (Multidrop 384, Labsystems, Biomac 2000 and MultiMeg 96 from Beckman Coulter, Fullerton, Fla.) Was added Serial dilutions and liquid transfer were performed.

A suitable well of a standard 96-well microdilution plate (Falcon 3918) was filled with 150 μL of the appropriate solvent using a MultiGrop 384 in columns 2-12. 300 microliters of each second test drug was added to each well in column 1 of the plate. These plates were used to prepare the drug " parent plate " and provided it for a series of drug dilutions for the drug combination plate. Biomec 2000 was used to transfer 150 μL of each second drug solution (40X) from the wells in column 1 of the parent plate to produce 11 double serial dilutions. The parent plates of Bis-EDT (and analogs) were serially diluted by hand using a multi-channel pipette from top to bottom. One plate for each second drug and one for Bis-EDT (or analogue) are combined to deliver the equivalent volume (using a multi-channel pipette) to the drug combination plate, resulting in a " checker plate & . Column H and column 12 each contained a single dilution of one of the formulations for the measurement of MIC.

&Quot; daughter plate " was loaded with 180 [mu] L of test medium using Multidrop 384. Subsequently, 10 μL of the drug solution was transferred from each well of the drug combination parent plate to the corresponding well of each of the daughter plates in one step using Multimec 96. Finally, the daughter plate was inoculated with the test organism. Standardized inoculations of each organism were prepared according to a known guide (CLSI, 2009). For all isolates, the inoculum for each organism was dispensed into length-divided sterile storage (Beckman Coulter) and inoculated onto plates using Biomek 2000. The device delivered 10 μL of the standardized inoculum to each well to obtain a final cell concentration in a daughter plate of approximately 5 × 10 ⁵ colon-forming-units / mL.

The test format produced an 8 × 12 checker plate where each compound was tested alone (column 12 and column H) and combined at various drug concentrations. All organism plates were stacked three heights, covered on the top plate, placed in a plastic bag, and incubated at 35 DEG C for approximately 20 hours. After incubation, the microplate was removed from the thermostat and observed from the bottom using a ScienceWare plate viewer. The prepared reading sheet was labeled for the wall of drug 1 MIC (column H), drug 2 MIC (column 12) and growth-no growh interface.

The FIC was measured according to the following equation using the Excel program: (MIC of compound 1 / MIC of compound 1 alone) + (MIC of compound 2 / MIC of compound 2 alone). The FICI for the checker plate was calculated from the individual FIC from the formula: (FIC ₁ + FIC ₂ + ... FIC _n ) / n, where n is the number of individual wells per plate for which FIC is calculated. For example, if an off-scale MIC result is obtained by the agent alone, the following maximum concentration is used as the MIC value in the FIC calculation.

Both particulate Bis-EDT, four particulate BT analogs, and other agents (and combinations of agents) were available at all final test concentrations. The measured MIC and FICI values are shown in the following Tables.

Example  11: Ratous Norvegicus ( Rattus Norvegicus ) Major defects in the femur Bismuth during infection Thiol  effect

Current standards of protection for open fractures are perfusion, debridement, and antibiotics; This is intended to reduce the bacterial load in the wound to a point where no infection occurs. Despite these treatments, infection still complicates up to 75% of severe multiple open tibial fractures. Interestingly, although early infections are often caused by gram negative bacteria, end-stage infections associated with healing problems and amputation are due to gram-positive infections, often staphylococcus species (Johnson 2007).

s. One of the reasons that Aureus is resistant to standard therapy is their ability to form a bacterium. Bacterial germs may be resistant to concentrations of antimicrobial compounds capable of killing similar organisms in the culture medium (see Costerton 1987).

The aim of the study was to determine whether BT would reduce infection in itself or in open fracture models contaminated with antibiotics. The contaminated rat femur major defect model was a well-accepted model and was used for the experiments described in this example. The model provides a standardized model for comparing the effects of various possible treatments and infections in the reduction of healing and / or improvement of healing.

Although the compound (CPD) CPD-8-2 (bismuth pyrithione / butanediol; Table 1) and CPD-11 (bismuth pyrithione / ethanedithiol; Table 1) have different activity spectra than Bis- Two analogues of BIS-Bis that have been shown to be effective against bacterial secretory bacteria in vitro.

The three BT formulations, Bis-EDT, CPD-11, and CPD-8-2 (see Table 1) were used to determine the presence of tobramycin and vancomycin in a polymethylmethacrylate (PMMA) cement bead vehicle, If used under test in the absence of S. The inhibitory effect on Aureus strain was proved. Three formulations of particulate BT were produced in the form of clinically useful hydrogel gels as described herein. These BTs were tested in suspension in a gel at a concentration of 5 mg / ml &lt; ^-1 &gt; which was found to be a suitable concentration for gel delivery. The gel formulation was tailed to the wound contour and did not require removal after application.

Two treatment arms were used: in the first arm, BT alone was used; In the second arm, BT was used with systemic antibiotic (ABx).

(a) BT alone

s. After 6 hours of inoculation, the wound was removed from the dead spot, perfused with saline, and 1 ml of BT gel was inserted into the defect.

(b) BT with systemic antibody (ABx).

s. After 6 hours of inoculation with Aureus, the wound was removed from the dead spot, and 1 ml BT gel added with perfusion through brine was added to the defect. The antibiotic used was a dose of cephazoline equivalent to 5 mgKg- ¹ delivered via subcutaneous-injection twice daily for a total of 3 days after injury. The first dose was administered just before the dead spot removal. The previous data suggests that the dose will produce a reduction in bacterial levels of about 10 ⁶ to about 10 ⁴ , thus allowing the relative effectiveness of different BT to be still measured.

(c) Control

s. After 6 hours of inoculation with aureus, the wound was removed from the dead place and resuspended in brine. Control animals were also treated with cephazoline according to the regimen described above.

process:

The procedure for in vivo rat damage model is described in Chen et al. As described in 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 The rats were anesthetized and prepared for surgery.The frontal aspect of the femoral body was exposed through a 3-cm incision.The periosteum and attached muscles were stripped from the bone.Poly acetyl plates (27 x 4 x 4 mm ) Were placed on the femoral front surface of the femur.The plate was pre-drilled to accept a 0.9-mm diameter Kirschner wire.The base of these plates was formed to conform to the contour of the femoral body. Pilot holes were drilled through both the cortex of the thigh using a plate as a template and Kirschner wire was inserted through the plate and thighs. A notch 6 mm away from the plate was provided as a guide for bone removal (C) using a small vibrating top (oscillating saw) is cooled by a continuous infusion in an effort to avoid damage while the tissue heat to produce a defect.

1 x 10 &lt; ⁵ &gt; CFU of S of 10 groups each. Aureus was inoculated and treated with BT alone or with antibiotics 6 hours after inoculation as described above. The groups are as follows: Bis-EDT gel; MB-11 gel; MB-8-2 gel; Bis-EDT gel &Abx; MB-11 gel &Abx; MB-8-2 Gel &Abx; Control (Abx only).

Animals were euthanized 14 days post-surgery and bone and hardware were sent for microbiological analysis and the results are shown in FIG.

On the basis of force analysis, 10 animals per group were given 80% force to detect a 25% difference between the treatment group and the control group. This is done with an estimated standard deviation of 35% and an alpha of 0.05.

As shown in Fig. 7, along with Bis-EDT, MB-11 and MB-8-2, the cephazoline antibiotic activity was improved compared to either the cephazoline or the Bis compound alone. Aureus infection was reduced. Sepharzolin in combination with MB-11 and MB-8-2 showed enhanced antibiotic activity compared to cytosol alone, Aureus infection was reduced. Bis-EDT appeared to have no effect on cephazoline in this capacity.

Reference literature:

Costerton JW, Cheng KJ, Geesey GG, et al. Bacterial Biofilms in Nature and Disease. Ann Rev Microbiol. 1987; 41 : 435-64.

Domenico P, Baldassarri L, Schoch PE, Kaehler K, M Sasatsu M, Cunha BA. Activities of Bismuth Thiols against Staphylococci and Staphylococcal Biofilms. Antimicrob Agents and Chemother. 2001; 45 (5) : 1417-21.

Halwani M. Blomme S, Suntres ZE, et al. Liposomal bismuth-ethanedithol formulation enhances antimicrobial activity of tobramycin. Int J Pharm. 2008; 358 : 279-84.

Johnson EN, Burns TC, Hayda RA, Hospenthal DR, Murray CK. Infection complications of open type III tibial fractures among combat casualties. Clin Infect Dis. 2007; 45 (4) : 409-415.

Other citations and related literature

Domenico et al., Canadian J. Microbiol. 31: 472-78 (1985); Domenico et al., Reduction of capsular polysaccharides and potentiation of aminoglycoside inhibition in gram-negative bacteria with bismuth subsalicylate. 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. 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-dimercaprol 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 Angens Chemother 2001; 45: 1417-21; Domenico et al., Combating antibiotic resistance with bismuth-thiols. Research Advances in Antimicrob Agents Chemother 2003; 3: 79-85; Domenico et al., Reductionof capsular polysaccharides and otentiation of aminoglycoside inhibition in gram-negative bacterial with bismuth subsalicylate. J Antimicrob Chemo 1991; 28: 801-810; Domenico et al., BisEDT and RIP act in synergy to prevent graft infections by resistant staphylococci. Peptides 2004; 25: 2047-53; domenico et al., 2005. Pyrithione enhanced antimicrobial activity of bismuth. Antibiotics for Clinicians 9: 291-297; U.S. Patent No. 6,582,719; Re-registered Patent RE37,793; U.S. Patent No. 6,248,371; U.S. Patent No. 6,086,921; U.S. Patent No. 6,380,248; U.S. Patent No. 6,582,719; U.S. Patent No. 6,380,248; U.S. Patent No. 6,875,453.

The various aspects described above may be combined to provide additional aspects. All of the US patents, U.S. patent publications, U.S. patent sources, foreign patents, foreign patent sources, and non-patent publications mentioned in and / or listed in these specifications are incorporated herein by reference in their entirety. Aspects of the aspects may be modified as necessary to still provide additional aspects by using the concepts of various patents, patent applications, and publications.

These and other variations can be made in terms of aspects in terms of the above detailed description. Generally, in the following claims, the terms used should not be construed to limit the scope of the claims to the specific embodiments described in the specification and claims, but should be construed broadly to cover the full scope of equivalents But should be construed to include all possible embodiments. Accordingly, the claims are not limited by the context of the present disclosure.

Claims (36)

A composition comprising a therapeutically effective amount of a microparticle for use in treating or preventing a bacterial infection in or on an epithelial wound, said composition comprising (i) the microparticle comprises bismuth-ethanedithiol (BisEDT), (ii) Wherein the particulate has a volumetric mean diameter of from 0.4 micrometers to 5 micrometers. A composition comprising a therapeutically effective amount of a microparticle for use in treating or preventing a bacterial infection in or on a diabetic foot ulceration, the composition comprising: (i) the microparticle comprises BisEDT; (ii) &Lt; / RTI &gt; to 5 microns. A composition comprising a therapeutically effective amount of microparticles for use in treating or preventing a bacterial infection resulting from an orthopedic procedure, said composition comprising: (i) said microparticle comprises BisEDT; (ii) said microparticles have a mean particle size of from 0.4 m to 5 m And a volume average diameter. 4. The composition according to any one of claims 1 to 3, wherein the bacterial infection comprises a biofilm. 4. The composition according to any one of claims 1 to 3, wherein the bacterial infection comprises at least one species of Gram-negative bacteria. 4. The composition according to any one of claims 1 to 3, wherein the bacterial infection comprises at least one species of Gram-positive bacteria. 4. The composition according to any one of claims 1 to 3, wherein the bacterial infection comprises at least one species of an antibiotic-resistant bacterium. 8. The composition according to claim 7, wherein the antibiotic is selected from methicillin, vancomycin, napthyline, gentamicin, ampicillin, chloramphenicol, doxycycline and tobramycin. 4. The method according to any one of claims 1 to 3, wherein the bacterial infection is selected from the group consisting of Staphylococcus aureus , methicillin-resistant S. aureus , Kus Epidermidis , Staphylococcus epidermidis , methicillin-resistant S. epidermidis , Mycobacterium tuberculosis , Mycobacterium avium , Pseudomonas aeruginosa, Pseudomonas aeruginosa , drug-resistant P. aeruginosa , Escherichia coli , enterotoxin-producing E. coli , enterohemorrhagic E. coli , Klebsiella A bacterial strain such as Klebsiella pneumoniae , Clostridium difficile , Heliobacter pylori , Legionella pneumophila , Enterococcus faecalis , methicillin- Enterococcus faecalis, Enterobacter cloacae , Salmonella typhimurium &lt; RTI ID = 0.0 &gt; (Salmonella typhimurium), Proteus vulgaris (Proteus vulgaris), Yersinia Enterococcus coli urticae (Yersinia enterocolitica), Vibrio cholera (Vibrio cholera), Shigella flexneri , vancomycin-resistant Enterococcus , Burkholderia cepacia complex, Francisella tularensis , Bacillus anthracis ( Bacillus anthracis ), Bacillus anthracis ), Yersinia pestis , Streptococcus pneumonia , Penicillin-resistant Streptococcus pneumoniae , Burkholderia sephacia , Bukholderia multivorans , Wherein the composition comprises at least one of Mycobacterium smegmatis and Acinetobacter baumannii . 4. The composition according to any one of claims 1 to 3, wherein the treatment or prevention of the bacterial infection comprises the viability or growth inhibition of the bacteria. 2. The composition of claim 1, wherein the epithelial wound is a diabetic ulcer. 12. The composition of claim 11, wherein the diabetic ulcer is a diabetic foot ulcer. 4. The composition of claim 3, wherein the orthopedic procedure comprises orthopedic surgery, orthopedic surgery, arthroplasty or orthodontic therapy. 14. The composition of claim 13, wherein the orthopedic procedure is orthopedic surgery. 4. The composition according to any one of claims 1 to 3, wherein the treatment or prophylaxis comprises topical administration of the composition. The composition according to any one of claims 1 to 3, wherein the particulate has not been applied to micronized, ground or supercritical fluid processing.
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