WO2003041500A1 - Formulations topiques de natamycine/pimaricine - Google Patents

Formulations topiques de natamycine/pimaricine Download PDF

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WO2003041500A1
WO2003041500A1 PCT/US2002/036616 US0236616W WO03041500A1 WO 2003041500 A1 WO2003041500 A1 WO 2003041500A1 US 0236616 W US0236616 W US 0236616W WO 03041500 A1 WO03041500 A1 WO 03041500A1
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composition
natamycin
solvent
acid
infection
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PCT/US2002/036616
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English (en)
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Borje S. Andersson
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Board Of Regents, The University Of Texas System
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/0012—Galenical forms characterised by the site of application
    • A61K9/0014—Skin, i.e. galenical aspects of topical compositions
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N43/00—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds
    • A01N43/90—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds having two or more relevant hetero rings, condensed among themselves or with a common carbocyclic ring system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
    • A61K47/08—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite containing oxygen, e.g. ethers, acetals, ketones, quinones, aldehydes, peroxides
    • A61K47/10—Alcohols; Phenols; Salts thereof, e.g. glycerol; Polyethylene glycols [PEG]; Poloxamers; PEG/POE alkyl ethers
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
    • A61K47/08—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite containing oxygen, e.g. ethers, acetals, ketones, quinones, aldehydes, peroxides
    • A61K47/12—Carboxylic acids; Salts or anhydrides thereof
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
    • A61K47/20—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite containing sulfur, e.g. dimethyl sulfoxide [DMSO], docusate, sodium lauryl sulfate or aminosulfonic acids
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
    • A61K47/26—Carbohydrates, e.g. sugar alcohols, amino sugars, nucleic acids, mono-, di- or oligo-saccharides; Derivatives thereof, e.g. polysorbates, sorbitan fatty acid esters or glycyrrhizin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/0012—Galenical forms characterised by the site of application
    • A61K9/007—Pulmonary tract; Aromatherapy
    • A61K9/0073—Sprays or powders for inhalation; Aerolised or nebulised preparations generated by other means than thermal energy
    • A61K9/0078—Sprays or powders for inhalation; Aerolised or nebulised preparations generated by other means than thermal energy for inhalation via a nebulizer such as a jet nebulizer, ultrasonic nebulizer, e.g. in the form of aqueous drug solutions or dispersions

Definitions

  • the present invention relates generally to the fields of infectious conditions and drug formulations. More particularly, it concerns the development of a pharmaceutical formulation of Pimaricin (also called as Natamycin) that can be used to treat and prevent a wide variety of topical infections caused by fungal and microbial pathogens.
  • Pimaricin also called as Natamycin
  • Superficial fungal infections constitute one of the more difficult challenges for clinicians caring for patients with diabetes mellitus and/or immunocompromised states, such as those caused by cancer, cancer-therapy and HIN-infection (Anaissie et al, 1989). While the incidence of hematogenous candidiasis in immunocompromised patients has decreased significantly with the introduction of fluconazole, the occurrence of superficial opportunistic infections affecting the mucous membranes and the skin of immunocompromised patients and have persisted and pose a major clinical problem carrying also the risk of serving as an entry port for systemic infection. The most common of such infections is oral thrush and vaginal Candida infections.
  • AMB polyene antibiotic Amphotericin B
  • Pimaricin also called as Natamycin, is an effective anti-fungal agent, exerting significant activity against molds and yeast, particularly of the Candida, Trichomonas, Aspergillus and Fusarium species. Pimaricin is especially effective in molds and yeasts that are resistant to azole compounds and to Amphotericin B.
  • Pimaricin was first isolated in 1955 from a strain of Streptomyces (Strayk et al., 1957- 1958). Pimaricin exhibited a wide range of in vitro activity against fungi, yeast, and trichomonads (Struyk et al., 1957-1958; Korteweg et al., 1963; Raab, 1972). The drug was found to have little or no toxicity after oral administration, being virtually non-absorbable from the gastrointestinal tract (Korteweg et al., 1963; Raab, 1972).
  • pimaricin's prominent chemical stability paired with its apparent lack of intestinal absorption and systemic toxicity, formed the basis for its FDA-approved use in the food industry, where it is used to prevent the proliferation of aflatoxin-producing molds (Code of Federal Regulations, Food and Drugs, 1995).
  • Parenteral formulations of pimaricin are useful in treating systemic infections such as disseminated fungal infections that are often seen in patients with hematological cancers.
  • Such parenterally acceptable, nontoxic formulations of pimaricin are beneficial not only for cancer patients, but also for other groups of immunocompromised patients, e.g. those suffering from HIV and those having recently undergone open heart surgery, all of which are commonly targets for opportunistic systemic infections.
  • the present invention relates to drug formulations that are useful for the treatment, prevention and suppression of infections caused by infectious pathogens such as fungi and microbes.
  • Microbes are exemplified by organisms such as bacteria and viruses.
  • Some exemplary fungal pathogens include those of the Candida, Trichomonas, Aspergillus and Fusarium species.
  • the invention provides pharmaceutically acceptable topical antifungal and/or an anitmicrobial compositions that comprise: an amount of pimaricin or a salt thereof that is effective to inhibit the growth of an infectious pathogen in a mammal; a pharmaceutically acceptable dipolar aprotic solvent or an acid; and a pharmaceutically acceptable aqueous secondary solvent.
  • the final composition is substantially free of the aprotic solvent and/or the acid.
  • substantially free it is meant that the composition is about 80 to about 99% free of organic solvents. Therefore the composition may be about 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 98.1%, 98.2%, 98.3%, 98.4%, 98.5%, 98.6%, 98.7%, 98.8%, 98.9%, 99% 99.1%, 99.2%, 99.3%, 99.4% 99.5%, 99.6%, 99.7%o, 99.8% or about 99.9%, free of the aprotic solvent and/or the acid. Intermediate ranges such as 98.75% etc., are also contemplated.
  • the final composition is free ofthe aprotic solvent and/or the acid.
  • free ofthe aprotic solvent or acid it is meant that the composition has no detectable levels of dipolar aprotic solvent or solvent.
  • the composition further comprises an ointment and/or a cream base.
  • the ointment base can comprises one or more of petrolatum, mineral oil, ceresin, lanolin alcohol, panthenol, glycerin, bisabolol, cocoa butter and the like.
  • the ointment or cream may be any commonly known commercially available ointments or creams such as AquaphorTM or EucerinTM.
  • the acid is a carboxylic acid and is exemplified by acetic acid, hi other embodiments, the acid may be hydrochloric acid.
  • compositions of Natamycin dissolved in acetic acid as a primary solvent provide concentrations ranging from about 3mg/ml to more than about 500 mg/ml ofthe active drug which is a broad and effective range for topical administration of doses necessary to yield active antibiotic concentrations that are effective to eradicate localized infections sensitive to this drug.
  • the primary solvent is a aprotic solvent such as N,N- dimethylacetamide (DMA) and dimethyl sulfoxide (DMSO), polyethylene glycol (PEG) or polypropylene glycol (PE).
  • DMA N,N- dimethylacetamide
  • DMSO dimethyl sulfoxide
  • PEG polyethylene glycol
  • PE polypropylene glycol
  • the aqueous secondary solvent is a surfactant.
  • surfactants are well known in the art and are organic lipid compounds that are normally produced by the lung tissue and help with the opening of the alveolar constrictions during breathing. Surfactants are also commercially available.
  • the aqueous secondary solvent is an aqueous lipid emulsion.
  • the aqueous lipid emulsion can comprise a lipid component that includes at least one vegetable oil and at least one fatty acid.
  • a lipid component can comprise at least about 5% by weight soybean oil and at least about 50% by weight fatty acids.
  • the lipids in the composition are preferably present in a form other than liposomes, for example, at least about 50%o by weight of the lipid is not in the form of liposomes, more preferably at least about 75%, and most preferably at least about 95%.
  • the secondary solvent can be water, a saline solution, or a dextrose solution.
  • the invention also provides methods of preventing or treating a localized infection in a mammal, comprising administering to a mammal any of the pharmaceutically acceptable compositions described above.
  • the method administering can be topical by application of an ointment to the locally infected area or may be by inhalation or nebuhzation of the respiratory pathways in the case of respiratory tract infections.
  • the composition may be administered by aerosolization to an affected mucosal membrane.
  • any local infection that is caused by a pathogen sensitive to Natamycin may be treated by the methods of the invention.
  • Non-limiting examples of such infections are vaginal infections, perirectal infections, skin infections, infections of the ingunal area, mucosal infections, intertriginous infections, scalp infections, infections of the outer ear canal, or respiratory tract infections.
  • compositions of the present invention can further include additional pharmaceutically acceptable carriers, adjuvants, and/or biologically active substances.
  • Compositions ofthe present invention as described above, can be used in methods for treatment or prophylaxis of infections in mammals, particularly in humans. The methods involve administering to a mammal an amount of the compositions effective to prevent, eliminate, or control the infection. The administering step can suitably be topically.
  • the compositions can also be administered intranasally as an aerosol. Such administration is preferably repeated on a timed schedule, and may be used in conjunction with other forms of therapy or prophylaxis, including methods involving administration of different biologically active agents to the subject.
  • the dose administered of a composition in accordance with the present invention is preferably between approximately lmg/ml and 500 mg/ml.
  • the invention also provides antifungal and/or an antimicrobial composition pharmaceutically acceptable for local administration, comprising: an amount of pimaricin or a salt thereof effective to inhibit the growth of a pathogen in a mammal; acetic acid; and an aqueous secondary solvent.
  • the acetic acid may be further removed to obtain compositions that are either substantially free or completely free of acetic acid.
  • Some examples of methods for removing acetic acid include lyophilization and/or neutralization using a base. However, it is contemplated that any other method known in the art to remove acetic acid may be used.
  • the aqueous secondary solvent may be a surfactant.
  • the aqueous secondary solvent a lipid emulsion that comprises at least one vegetable oil and at least one fatty acid.
  • the composition can further comprises an ointment and/or a cream base.
  • the invention also provides methods of preventing or treating a local infection in a mammal, comprising the step of administering topically to a mammal a composition as described above.
  • the invention also provides pharmaceutically acceptable antifungal and/or antimicrobial compositions that are substantially free of aprotic solvents or acids prepared by the method comprising: obtaining pimaricin or a salt thereof; dissolving the pimaricin in a pharmaceutically acceptable dipolar aprotic solvent or an acid; further dissolving the composition in a pharmaceutically acceptable aqueous secondary solvent; and removing the dipolar aprotic solvent or the acid from the composition.
  • the composition is made by a method that further comprises adding to the composition an ointment and/or a cream base.
  • the acid is acetic acid.
  • the acid is hydrochloric acid
  • the aprotic solvent is selected from the group comprising DMSO, DMA, PE, or PEG.
  • the aqueous secondary solvent is a surfactant.
  • surfactants are well known in the art and are organic lipid compounds that are normally produced by the lung tissue and help with the opening of the alveolar constrictions during breathing. Surfactants are also commercially available.
  • the aqueous secondary solvent is an aqueous lipid emulsion.
  • the aqueous lipid emulsion can comprises a lipid component that includes at least one vegetable oil and at least one fatty acid.
  • a lipid component can comprise at least about 5% by weight soybean oil and at least about 50% by weight fatty acids.
  • the secondary solvent can be water, a saline solution, or a dextrose solution.
  • the present invention is especially useful for preventing or treating systemic fungal infections, it can also be used for prevention and treatment of systemic infections caused by other infectious agents that are sensitive to pimaricin in vivo, such as viruses.
  • FIG. 1 Chemical structure of pimaricin as free drug.
  • AMB Amphotericin B. AquaphorTM: Comprises Petrolatum, Mineral Oil, Ceresin, Lanolin Alcohol, Panthenol, Glycerin, Bisabolol.
  • DMA Anhydrous N,N,-dimethylacetamide.
  • HAc Glacial acetic acid.
  • HL-60 Human myeloid leukemia cell line.
  • EMDM Iscove's modified Dulbecco Medium (GIBCO, Grand Island, New York, N.Y.).
  • IntralipidTM Brand name of an aqueous lipid emulsion, made from soy bean oil, and marketed for parenteral nutrition by Clintec.
  • KBM-7/B5 Human myeloid leukemia cell line.
  • PEG Polyethylene glycol-400.
  • PG Polypropylene glycol/l,2-propylene diol.
  • SDS Sodium dodecyl sulphate.
  • Pimaricin or Natamycin is a drug with a wide-spectrum of antibiotic effects.
  • its insolubility makes it largely unavailable for medical utility, h U.S. Patent 6,045,815, the inventors have described solubilized compositions of Pimaricin which are suitable for parenteral use, methods for solubilization of Pimaricin and methods for using solubilized Pimaricin to treat systemic infections.
  • the inventors have developed pimaricin compositions suitable for topical use where pimaricin or a salt thereof is dissolved using a dipolar aprotic solvent or an acid as the primary vehicle and then further dissolved in one or more secondary cosolvent(s) to increase the drug's aqueous solubility and stablity.
  • this composition can be combined with ointments or cream bases to allow topical applicability.
  • the composition can be in the form of an aerosol or a spray for other forms of topical administration.
  • the dipolar aprotic solvent and/or acid which are often agents that are not pharmaceutically acceptable in large doses, can be removed completely or substantially to pharmaceutically acceptable levels.
  • Pimaricin formulations are stable for many weeks at room temperature, and that they retain full antifungal activity when applied topically to immunocompromized, infected animals.
  • animal models of topical infection can be used to demonstrate the efficacy of the formulations ofthe invention.
  • the topical Natamycin formulations ofthe invention permit what has heretofore been impossible, safe topical administration of fully solubilized drug that is highly effective against pathogens and has negligible toxicity and yields negligible Natamycin plasma concentrations.
  • the present invention therefore, provides a solubilized topical Natamycin formulation that makes available high concentrations of the active drug.
  • the formulation of the invention may be further mixed in an ointment or cream base.
  • This composition is also highly shelf-stable.
  • the invention provides a formulation of Natamycin that can be used effectively against a wide variety of pathogens, including, azole resistant yeast species, as well as a variety of molds that cause infections such as, localized Aspergillus, Fusarium, and mucor infections that are inherently resistant to commonly available alternative treatments, i.e., azoles and AMB.
  • the invention provides methods for the treatment of a wide variety of pathogens including microbes and fungi.
  • Some examples of fungal pathogens that are susceptible to Natamycin include molds and yeasts.
  • Specially localized skin and mucosal infections may be treated and/or prevented by the methods of the invention.
  • topical treatments for the localized infections with the formulation ofthe invention the inventors also contemplate that the formulation may be administered by aerosolization or nebuhzation so that the composition may be inhaled to treat respiratory track infections.
  • Natamycin has antibiotic activity against Fusarium species as well as Aspergillosis sepcies use of the formulation will significantly improve the existing therapeutic armamentarium against opportunistic fungal pathogens that first infect the skin or mucosal surfaces and eventually gain entry into an organism causing wide-spread systemic infections.
  • Natamycin is known to have low local organ toxicity, broad antifungal spectrurm activity, predictable lack of systemic absorption after topical administration and activity against numerous pathogens. Natamycin is recognized by the FDA as a GRAS compound (Generally Regarded As Safe). Therefore, the invention provides formulation of Natamycin that are physiologically compatible with topical application in man and domestic animals.
  • the present invention involves methodology for solubilization of Natamycin in pharmaceutically acceptable, chemically diverse vehicles, such that the drug remains chemically stable and can be administered topically without undue toxicity from undissolved drug and/or from the solvents at drug doses predicted necessary to obtain clinically significant antibiotic effects.
  • the present invention involves solubilization of pimaricin in pharmaceutically acceptable vehicles, such that the drug remains chemically stable and can be further formulated into an ointment or cream and administered topically without undue toxicity from the solvents at drug doses necessary to obtain clinically significant antibiotic effects.
  • Pimaricin is available from Gist-Brocades N.V. (Netherlands) and Sigma Chemical Co. (Saint Louis, Mo.). Pimaricin optionally can be used in compositions ofthe present invention in the form of one of its antifungal derivatives, such as a salt of pimaricin (e.g., an alkali salt or an alkaline earth salt).
  • a salt of pimaricin e.g., an alkali salt or an alkaline earth salt.
  • the present inventors have formulated topical formulations of pimaricin which are lipid based, i.e., in the form of creams, ointments, gels, which are suitable for topical treatments.
  • the inventors have successfully used acetic acid as a primary solvent to dissolve the pimaricin.
  • Other primary solvents are DMSO, glycerol, 1,2,-propylene-diol, (PG), and polyethylene glycol- 400 (PEG).
  • the Natamycin formulation is further miscible in secondary solvents, examples of which are normal saline, dextrose in water (5% or 10%), and an aqueous soy bean lipid emulsion (IntralipidTM).
  • These solvents are examples of vehicles in which pimaricin can be suitably solubilized, yet be safe for human administration, alone or in combinations with other drugs.
  • the solubility of pimaricin in individual solvent vehicles is shown in Table 1 below.
  • the described vehicles can be utilized to dissolve Natamycin in concentrations ranging from about lmg/ml to more than about 100 mg/ml.
  • an enhanced solubility of >300 mg/ml was achieved by lowering the pH drastically by dissolving the drug in glacial acetic acid, and thereafter mixing it with a secondary solvent, such as a soybean lipid emulsion exemplified by IntralipidTM.
  • This range should cover the administration of doses necessary to yield active antibiotic concentrations in vivo to eradicate superficial infections sensitive to this drug.
  • the invention will therefore allow the introduction of Natamycin in clinical practice for the therapy of localized infections such as vaginal and/or dermatological infections.
  • the present invention provides a method to administer Natamycin topically to achieve very high local concentrations without achieving any systemic concentration of the drug.
  • the topical application approach will provide an opportunity to investigate the outstanding therapeutic potential of this agent. These have previously been hampered by the drug's perceived lack of solubility in clinically acceptable solvents for both systemic and topical administration.
  • the objective of this invention includes the topical administration of Natamycin to improve the control of localized infections that are sensitive to this agent. Infections by various Candida species are a paradigm for such infections. Topical use of a fully solubilized Natamycin formulation has not been previously investigated in the practice of medicine, although the drug has well documented anti-fungal properties in vitro, and some antifungal activity has been reported in crystalline suspensions of the drug (Struyk et al, 1957-1958; Korteweg et al, 1963; Raab, 1972).
  • the composite diluent vehicles rapidly solubilize Natamycin without destroying its antifungal properties.
  • the preferred vehicles are nontoxic and safe for administration in large as well as small animals, and should be acceptable for human administration in the proposed concentrations and total doses to be utilized; indeed, HAc, DMSO, and PG have previously been used for solubilization of various pharmacologically active agents used in man (U.S. Dept. of Health and Human Services, 1984; Weiss et al, 1962; Kim, 1988).
  • DMSO is also extensively used as a cryoprotective agent for low-temperature storage of human bone manow and peripheral blood derived hematopoietic stem cell preparations to be used for transplantation after high-dose chemotherapy (McGann, 1978; Gorin, 1986; Davis and Rowley, 1990; Gorin, 1992). No serious adverse effects have been experienced from the use of these agents.
  • the clinical use of normal saline, dextrose in water (70%), and aqueous lipid emulsions as well as various ointments are well established means to alter dermal elasticity, promote wound healing and to combat infections.
  • Natamycin formulations allows reproducible topical treatment of localized infections and provides superior infection control without systemic side effects.
  • the plasma concentrations of Natamycin clearly remained in the undetectable range whereas the antifungal activity remained at its peak for prolonged time periods.
  • the fungicidal range was established by in vitro studies of antifungal activity against Candida species, Aspergillus species, and Fusarium species.
  • the novel Natamycin/HAc/lipid solution is chemically stable and simple to handle at room temperature (RT) and can be further mixed with an ointment base such as AquaforTM.
  • the formulations of the invention provide reliable and easily controlled dosing with a retained and highly effective antifungal effect.
  • the addition of a lyophilization step substantially eliminates the DMA, HAc, or other dipolar aprotic solvent or acids, from the final clinical "working solution", thereby abolish the potential for adverse reactions related to the drastically lowered pH attained with the use of HAc as the primary solvent. This added step should therefore assist in maximizing patient safety after drug administration.
  • the composition is made using Hac, it may be of some advantage to retain some of the HAc in the Natamycin formulations such that the lower pH corresponds to the vaginal pH.
  • the skilled artisan can decide how much, if any, ofthe acid or dipolar aprotic solvent is acceptable in a final composition.
  • Natamycin allows the investigation of its clinical usefulness against localized infections.
  • the access to topical Natamycin might be particularly important, since their possible assimilation of infectious agents through impaired mucosal surfaces might predispose them for secondary systemic fungal infections.
  • the availability of Natamycin for effective and reliable topical administration will for the first time make it possible to clinically compare the activity of Natamycin against that of "the gold standard MonistatTM", for the treatment of vaginal yeast infections.
  • Natamycin may be the only effective drug for the treatment of azole resistant yeasts, which are becoming gradually more widespread with the rapidly increasing indiscriminant use of azoletherpay (primarily ftuconazole).
  • a nontoxic, pharmaceutically acceptable, water miscible, topical formulation of Natamycin eliminates the risk of treatment failure from the suboptimal bioavailability of incompletely solublized drug.
  • the addition of a lyophilization step in the preparative procedure will create a Natamycin solvate with minimal acid content. This will reduce the risk of adverse effects related to the vehicle's acid component.
  • compositions Natamycin for clinical applications will entail preparing compositions that are essentially free of pyrogens, as well as other impurities that could be harmful to humans or animals.
  • phrases "pharmaceutically or pharmacologically acceptable” refer to molecular entities and compositions that do not produce adverse, allergic, or other untoward reactions when administered to an animal or a human.
  • pharmaceutically acceptable carrier includes any and all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents and the like. The use of such media and agents for pharmaceutically active substances is well know in the art. Except insofar as any conventional media or agent is incompatible with formulations of the present invention, its use in therapeutic compositions is contemplated. Supplementary active ingredients also can be incorporated into the compositions.
  • compositions according to the present invention will be via any common topical route so long as the target tissue is available via that route.
  • Solutions ofthe active compounds as free base or pharmacologically acceptable salts can be prepared in water suitably mixed with a surfactant, such as hydroxypropylcellulose. Dispersions can also be prepared in glycerol, liquid polyethylene glycols, and mixtures thereof and in oils. Under ordinary conditions of storage and use, these preparations contain a preservative to prevent the growth of microorganisms.
  • compositions of the present invention may be formulated in a neutral or salt form.
  • Pharmaceutically-acceptable salts include the acid addition salts (formed with the free amino groups of the protein) and which are formed with inorganic acids such as, for example, hydrochloric or phosphoric acids, or such organic acids as acetic, oxalic, tartaric, mandelic, and the like. Salts formed with the free carboxyl groups can also be derived from inorganic bases such as, for example, sodium, potassium, ammonium, calcium, or ferric hydroxides, and such organic bases as isopropylamine, trimethylamine, histidine, procaine and the like.
  • the composition may be formulated as a "unit dose.”
  • one unit dose could be dissolved in 1 ml of isotonic NaCl solution and either added to 1000 ml of hypodermoclysis fluid or injected at the proposed site of infusion, (see for example, "Remington's Pharmaceutical Sciences” 15th Edition, pages 1035-1038 and 1570-1580).
  • Some variation in dosage will necessarily occur depending on the condition of the subject being treated.
  • the person responsible for administration will, in any event, determine the appropriate dose for the individual subject.
  • preparations should meet sterility, pyrogenicity, general safety and purity standards as required by FDA Office of Biologies standards.
  • the sensitivity data are displayed in Tables II and III; the activity of Natamycin was similar to that of AMB. Most ofthe Aspergillus and Fusarium spp. were sensitive to Natamycin, independent of the solvent system.
  • the HAc/IntralipidTM formulation that was lyophilized and reconstituted with distilled water, retained full and stable anti-fungal efficacy, when assayed both after 3 days and after having been stored for more than 2 months at 4°C. All the Aspergillus strains, except for A. flavus, had Natamycin MIC values in the 2-4 ⁇ g/ml (2.1-4.2 M) range. The tested A.
  • flavus was also sensitive to the drug, but with a slightly higher MIC value of 16 ⁇ g/ml (17 ⁇ M). All the tested strains of Fusarium and Candida spp. were sensitive to Natamycin in the range of 2-4 ⁇ g/ml (Tables II and III).
  • Lipid+DMA refers to freshly mixed DMA and Intralipid (1:10, v/v), which exerts no antifungal activity by itself.
  • ⁇ ata-lipid 1 and " ⁇ ata-lipid 2,” pimaricin was dissolved in DMA to 100 mg/ml then diluted with 20% Intralipid to 10 mg/ml "use-formulation.”
  • ⁇ ata-lipid 1 refers to a formulation where pimaricin was dissolved as above, and after dilution to 10 mg/ml using Intralipid, it was lyophilized. The lyophilized material was refrigerated for 4 months, then reconstituted in normal saline to 10 mg/ml and tested for antifungal activity.
  • ⁇ ata- lipid 2 refers to a formulation where the pimaricin/DMA/Intralipid formulation was prepared as for ⁇ ata-lipid 1 and lyophilized immediately, and was reconstituted and tested for antifungal activity three days later.
  • AMP+DMSO refers to a formulation of Amphotericin B dissolved immediately prior to use in DMSO, to serve as a positive control.
  • Natamycin solubility was determined in several individual vehicles (see Table I). Briefly, a known amount of the drug, as a powder (different lots of purified drug were obtained from Gist-Brocades N.N., Netherlands, and from Sigma Chemical Company, St Louis, MO), was equilibrated in the respective solvent at RT (22°C) over 5 min to 4 hours. An aliquot was then removed and diluted in MeOH prior to HPLC at predetermined times. Based on the Natamycin solubility in these particular vehicles, the inventors attempted to enhance the solubility by mixing different solvents according to the principle of cosolvency (Spiegel and Noseworthy, 1963; Yalkowsky and Roseman, 1981).
  • the examined solvents included DMA, DMSO, PEG, and PG, and HAc, NS, 5% dextrose in water and further with an aqueous soy bean emulsion (IntralipidTM) as secondary solvent.
  • HAc and DMA were the best primary solvents, providing a solubility in excess of 10 mg/ml.
  • there is a major difference in solubility between DMA and HAc in that maximum solubility of >300 mg/ml was reached in HAc within a few minutes whereas maximum solubility in DMA was reached after several hours at RT.
  • Natamycin could be dissolved in HAc and DMA to at least 100 mg/ml, it started degrading already within a few hours in solution.
  • the problem of stabilizing Natamycin once dissolved in HAc or DMA was addressed with the following approach based on the cosolvency principle. First the dissolved Natamycin was stabilized by the addition of an aqueous soy bean lipid emulsion (Intralipid), and the HAc was then removed by lyophilization or through neutralization ofthe acid by addition of a base in equal amount to a pH in the low-nonnal range (pH 5.0-6.0).
  • Intralipid aqueous soy bean lipid emulsion
  • a composition suitable for topical administration was obtained.
  • a natamycin ointment was made with HAc as the primary solvent, IntralipidTM as a secondary solvent, followed by mixing in an ointment base exemplified by Aquaphor.
  • a liquid chromatographic system equipped with an LDC 4000 multi-solvent delivery systemTM and a WatersTM system 717plus AutoinjectorTM was used to detect low concentrations of Natamycin.
  • the absorbance detector was a LDC 3100 variable wave length detector in sequence with an LDC model CI 4100 fully computerized integrator.
  • the column used was a Whatman EQC 10 ⁇ l 125ATM C18 column (4.6 mm i.d. x 21.6 cm) (Whatman Inc. Clifton, NJ).
  • the mobile phase system was an isocratic mixture of MeOH (47% v/v), tetrahydrofuran (2% v/v), and NH 4 -acetate (0.1% w/v) made up to 100% with double-distilled water. All chemicals were HPLC grade unless otherwise indicated.
  • the flow rate was 1.5 ml/min and the recorder's chart speed was 5 rnm/min.
  • HPLC Assay Two examples of authentic Natamycin chromatograms from the HPLC assay were obtained.
  • the drug was (a) analyzed in the aqueous DMA- tralipid solvent, and (b) was extracted from human plasma that had been spiked with 5 ⁇ g/ml of natamycin prior to extraction as described above.
  • the retention time under the above conditions was 9.8-10.8 min, and the assay was linear from 100 ng/ml to 25 ng/ml in protein-free solutions, i.e. the various solvent systems utilized in the formulation-feasibility and formulation-stability studies, and from about 50 ng/ml to 1 mg/ml for protein-containing solutions (plasma samples).
  • This assay consistently yielded high recovery, accuracy and a lower sensitivity limit of about 10 ng/ml.
  • the technique was standardized and used without modifications for the studies of both stability and pharmacokinetics.
  • the objective of the present example was to; (1) design stable Natamycin formulations that are suitable for topical administration; (2) establish the chemical and physical stability of Natamycin in the novel vehicles; (3) establish the solubility of Natamycin in these vehicles when mixed with IntralipidTM; and (4) investigate the in vitro properties of these formulations; i.e. parameters such as their osmolarity, hemolytic potential, and cytotoxicity, to show that they are non-toxic, non-irritating and therefore appropriate for the intended topical treatment purposes.
  • Natamycin was dissolved at a concentration of 100 - 300 mg/ml in HAc ("stock solution”) and incubated at 4°C, at 22°C and at 40°C. The drug concentration was analyzed by HPLC in samples taken immediately after solubilization and after gradually increasing time intervals of up to 48 hours.
  • the Natamycin-HAc stock solution was diluted with PEG/water (1:1:1, v:v:v, HAc:PEG:water), or PG/DMSO (1:1:1, v:v:v), or PG DMSO/PEG (1:1:1:1, v:v:v:v), or aqueous lipid emulsion (1:10 and 1:100, v:v, HAc:IntralipidTM), to yield Natamycin concentrations ranging from 1-50 mg/ml.
  • HAc-Natamycin mixture was diluted in NS or 5% dextrose to a drug concentration of 1 mg/ml.
  • HAc-Natamycin mixture was blended with DMSO and IntralipidTM, or directly in IntralipidTM.
  • the inventors investigated lyophilization as part of the preparation of a complete Natamycin/HAc/aqueous lipid-solvate vehicle.
  • the lyophilization step can create a composite solvate containing Natamycin and lipid, but only trace amounts of HAc.
  • the reconstitution of this solvate can be achieved by the addition of distilled water or a buffer only, and the low HAc content and/or the neutral pH of the final solvent system yields a low- acceptable hemolytic potential, and therefore also low inflammatory potential, of the final use- formulation.
  • HL-60 or KBM-7/B5 cells in Iscove's modified Dulbecco medium (EVIDM) supplemented with 10% fetal bovine serum were incubated for 60 min at 37°C with the complete vehicles (a: HAc/IntralipidTM, 1:10, v/v, or c: HAc/IntralipidTM, 2:6:3, v/v) at increasing concentrations of the vehicle (0.5%, 1.0%, 2.0%, 3.0%, and 10%, v/v) . with or without Natamycin.
  • the cells were washed in ice- cold PBS and resuspended in BVIDM with 10% fetal bovine serum at 37°C.
  • MTT solution (5 mg/ml) (Sigma Chemicals, St. Louis, MO) was added to each sample, and following an additional 2 hours of incubation at 37°C, 100 ⁇ l extraction buffer was added (extraction buffer: 20% (w/v) SDS dissolved to saturation at 37°C in a solution of DMF and deionized water (1 :1); pH 4.7). After incubation overnight at 37°C, the optical . densities were measured at 570 nm using a Titer-TechTM 96 well multi-scannerTM, against extraction buffer as the calibrating blank.
  • cytotoxicity was determined as the colorimetric difference between the samples exposed to solvent (Natamycin as above and the background reactivity of cells that had been incubated in parallel in PBS alone. All determinations were performed in triplicate (Hansen et al,; Andersson et al, 1996).
  • Parthasarathy et al to examine the hemolytic potential of a few selected preparations (Parthasarathy et al, 1994), and the LD 50 values of the various formulations were constructed as described. Briefly, heparinized blood was mixed with an equal volume of Alsever's solution. This mixture was washed twice in PBS, and a 10% (v/v) erythrocyte/PBS solution was then prepared and mixed with increasing amounts of the complete solvent system with or without the addition of Natamycin. These mixtures were then incubated for 4 hours at 37°C.
  • the cells were pelleted at 10,000 x g in an EppendorffTM centrifuge, and the release of hemoglobin in the supernatant (i.e. hemolysis) was spectrophotometrically determined at 550 nm. Maximum lysis was measured against a reference solution of erythrocytes that had been completely lysed by hypotonic shock.
  • the hemolytic potential of three of the complete formulations was evaluated as described (Parthasarathy et al, 1994), and the data were plotted as the fraction of healthy cells versus the total volume percent. Total volume percent was defined as the volume percent of the vehicle in the mixture after dilution with blood.
  • the temperature-dependent stability of solubilized Natamycin in the different solvent systems was studied as follows: The drug was dissolved in HAc at 100 mg/ml, and different aliquots were stored at 4°C, at 22°C, and at 40°C. Immediately after solubilization and at various intervals up to 48 hours later, aliquots from the different samples were analyzed by HPLC. The drug samples stored at 4°C and at 22°C degraded slower than those stored at higher temperatures: at 40°C the Natamycin started degrading within 1 hour after the start of incubation, and at RT there was a loss of 5-10% in the first 4 hours.
  • HAc and water have similar boiling points, (116°C and 100°C, respectively) it was envisioned that the HAc could be effectively removed by lyophilization of the Natamycin/HAc/aqueous lipid complex to create a solvate that was stable yet easily reconstituted by adding only double-distilled water under gentle agitation without any appreciable loss of antifungal efficacy, and this assumption proved correct.
  • the drug was reconstituted at 10 - 50 mg/ml, without remaining amounts of HAc.
  • This reconstituted Natamycin formulation retained an anti-fungal efficacy that was equivalent to that of the freshly prepared HAc/aqueous lipid formulation when assayed in vitro (see below).
  • This reconstituted formulation was also stable at RT for more than 4 weeks when mixed with an ointment base, AquaphorTM.
  • the lyophilized Natamycin formulation remained stable (by HPLC) for more than four months at 4°C. This preparation could still be readily reconstituted to 10 mg/ml within a few minutes with distilled water, with retention of full anti-fungal activity in vitro (see Table II).
  • the inventors further simulated a final clinical use- formulation with a Natamycin of 5 mg/ml by diluting the 50 mg/ml-HAc formulations (prepared fresh with HAc/Intralipid or after lyophilization reconstitution respectively) with different ointment bases.
  • the "use-formulations" are stabile at RT.
  • HAc and DMSO were used as the primary solvent system prior to mixing with Intralipid and followed by lyophilization, the majority ofthe organic solvent, here DMSO, was removed and the result was a stable lipid-based solvate was obtained, that could be easily reconstituted to 10 mg/ml under gentle agitation after the addition of distilled water.
  • This reconstituted formulation was also stable for more than 24 hours at RT assessed by HPLC.
  • Osmotic Pressure It is desirable that a parenteral formulation of a pharmacologically active agent be isosmotic to blood in order not to cause secondary irritation/inflammation on an already irritated (from infection) mucosal surface or skin.
  • a hypertonic delivery system should therefoOre be avoided if at all possible.
  • the osmotic pressure of a few ofthe tested formulations are shown in Table III.
  • a DMA-based formulation with or without Natamycin was very hypertonic and therfore omitted from further testing.
  • the HAc/Intralipid preparation was closer to isosmotic, both when prepared fresh and when reconstituted after lyophilization (Table III).
  • the lyophilized/reconstituted HAc/DMSO/IntralipidTM vehicle was also close to isosmotic. Adding Natamycin to the respective vehicles did not appreciably change their osmolarity (P > 0.05).
  • P > 0.05 In Vitro Cytotoxicity of Natamycin.
  • the HL-60 and KBM-7/B5 myeloid cells were exposed to the selected vehicles at increasing volume ratios with or without the addition of increasing drug concentrations. The cytotoxicity of each formulation was then assayed in the MTT assay (Hansen et al, 1989; Andersson et al, 1996). None ofthe examined solvent systems exerted any detectable toxicity against the cells in this assay.
  • the HAc/IntralipidTM "fresh" formulation had a significantly lower hemolytic potential than any of the solvent systems containing organic solvent. Further, the hemolytic potential of the lyophilized HAc/Intralipid formulation was significantly lower than that of the freshly prepared HAc/aqueous lipid formulation for all tested Natamycin concentrations.
  • the inventors have shown that the drug can be recovered from the "final" topical use- formulation exemplified by HAc-Intralipid-Natamycin mixed in a suitable ointment base, intended for application to experimental animals or humans using a quantitative extraction technique and HPLC assay.
  • Natamycin in a complete use-formulation of HAc-fritralipid was mixed with a suitable ointment base, AquaphorTM or petroleum jelly, at Natamycin concentrations of 10-50 mg/ml.
  • the Natamycin was then extracted from the ointment using a slight modification of a method described in U.S. Patent 6,045,815, the contents of which are incorporated herein by reference. Briefly, 0.2 ml ointment was mixed with 0.2 N HCl in MeOH (1:1, v/v), and after thorough mixing by a vortex machine, the sample was extracted with three volumes of hexane.
  • the hexane was separated from the Natamycin by evaporation and the drug was reconstituted in 200 ⁇ l of MeOH prior to HPLC. Natamycin was spectrophotometrically detected in the HPLC analysis as described above. The Natamycin recovered from the ointment conesponded to a drug concentration of 10 mg/ml was calculated to be 86 ⁇ 5%. The assay was linear in the interval from 50 ng/ml to 1,000 ⁇ g/ml (data not shown).
  • lyophilization or neutralization is used in the methods of the invention to prepare natamycin formulations that are free or substantially free of the dipolar aprotic or acid components to arrive at pharmaceutically acceptable formulations, h this example, lyophilized formulations of natamycin were analyzed by gas-chromatography/mass- spectroscopy (GC/MS) to determine how much organic solvent (dipolar aprotic solvent or acid) is remaining in the formulation following lyophilization.
  • GC/MS gas-chromatography/mass- spectroscopy
  • Gas chromatographic studies such as the GC/MS methods described herein are typically used in the art as assays to detect the presence of residual organic solvents in compositions such as the instant solvent vehicles. These assays are extremely sensitive and are known to detect organic solvents as low as 10-100 ng/mL. For examples, see Mulligan et al, 1995; Camarasu et al, 1998; Li et al, 2002; that describe the use of gas-chromatography-mass spectroscopy as standard methods to detect the presence of residual solvents.
  • GC/MS was performed on samples of the instant compositions, exemplified by N'N- dimethlyacetamide (DMA) as a dipolar aprotic solvent and Intralipid (an aqueous lipid emulsion) as a secondary aqueous solvent with or without pimaricin.
  • DMA N'N- dimethlyacetamide
  • Intralipid an aqueous lipid emulsion
  • solvent vehicle with pimaricin was analyzed.
  • the standard curve for comparison was prepared using DMA and hexane.
  • Other characteristics ofthe GC/MS are as outlined below:
  • Oven temperature 40C, 2min; at 5C /min, to 70C.
  • the samples numbered 1, 2, 3, 4, 5, and 6 in Table N were prepared by mixing DMA and Intralipid (1ml and 5 ml), 2 ml of which was analyzed for DMA content by GC/MS prior to lyophilization. These samples are designated as having 100% DMA content.
  • samples numbered 7, 8 and 9 were prepared as described above for samples 1-6 by mixing DMA and Intralipid (1ml and 5 ml), 2 ml of which was then lyophilized for 36 hours, followed by reconstitution in 2ml of saline prior to GC/MS analysis.
  • the samples numbered 10, 11 and 12 were prepared by mixing the dipolar aprotic DMA (lml) with the drug pimaricin at a concentration of 10 mg/ml. This was followed by mixing Intralipid (5 ml). 2 ml of this was then lyophilized for 36 hours, followed by reconstitution in 2ml of saline prior to GC/MS analysis.
  • Samples 13- 16 were prepared by mixing 200 ⁇ l of DMA and 200 ⁇ l of hexane by vortexing for 2 minutes. The mixture was centrifuged for 5 minutes at 2500 rpm, 100 ⁇ l of the organic phase was obtained and 1.0 ⁇ l of this was injected into the GC/MS. Samples 13-15 were further diluted with hexane to obtain different DMA concentrations of 1.0, 2.5, and 5.0%> respectively. These samples were then subject to GC/MS to obtain a standard curve that indicates the linearity for detection of DMA by GC/MS.
  • samples that were not lyophilized. represented by sample numbers 1-6 have a peak area of about 16422300 to 18576262 (see column 4 of Table V) which is designated as having 100%> of DMA.
  • Samples that were sub ect to lyophilization, represented by sample numbers 7-12 have a peak area of about 0 in three cases out of six cases and peak areas of 298308, 199250 and 183498 in three other cases. This represents 0% of DMA content for sample numbers 7, 11 and 12 and 1.7% DMA for sample number 8, 1.1% DMA for sample number 9, and 1.0% DMA for sample number 10 respectively (see data in column 7, Table V).
  • the samples of solvent vehicle and drug samples numbered 10, 11 and 12 that were prepared by mixing the dipolar aprotic DMA (1ml) with the drug pimaricin at a concentration of 10 mg/ml were "free” of the DMA in the cases of samples number 11 and 12 with 0.0% being detected and “substantially free” of DMA in sample 10 with only 1% being detectable.
  • the DMA content was "virtually eliminated” in 3 out of six cases (with only 1.7%., 1.1% and 1.0% being detected in these samples) and “eliminated” in the other 3 samples where no detectable DMA was seen.
  • dipolar aprotic solvent or acid 98% or more ofthe dipolar aprotic solvent or acid is eliminated by these methods resulting in final compositioins of natamycin that are free or substantially free of organic solvents.
  • all of the dipolar aprotic solvent or acid can be removed if required by extending lyophilization times. However, removal of all the dipolar aprotic solvent or acid may not be necessary in some cases.
  • removal of dipolar aprotic solvent or acids to an acceptable amount may be required by a health regulating body. The removal of dipolar aprotic solvent or acid component minimizes the side effects from contaminant organic solvent in the final use pharmaceutical formulations.
  • the efficacy of the topical formulations of the invention can further be analyzed using animal models of infections.
  • animal models such as the rodent models.
  • rodent models Some examples of rodent models used in the art are described in Black et al, 1999; Ghelardi et al., 1998; Foldvari et al, 2000; Jansen et al., 1991; Larsen and Galask, 1984; the entire contents of which are incorporated here by reference. Described below is a rodent model of vaginal yeast infection which can be used.
  • Intravaginal Infection Model An in vivo assay to test the antifungal properties of a topical natamycin preparation, made by the methods of the invention, may be done on a rodent model of vaginal yeast infection. Rodents models are useful due to their high reproducibility, ability to be pharmacologically manipulated to be immunosuppressed, and their vaginal tract can be easily evaluated for the local adverse effects of polyenes and other antifungal agents. After immunosuppression with estrogen derivatives +/- progesterone would be administered subcutaneously prior to vaginal inoculation with yeast.
  • the Natamycin compositions of the invention exemplified in one instance by HAc/IntralipidTM mixed in an ointment base, e.g.
  • AquaphorTM would then be administered intravaginally in a small volume (not exceeding 100 ⁇ L). To assure reproducibility of the experimental conditions, the experiments would be staggered; one set of animals rats at each of several dose levels would be started on consecutive days, and repeat experiments would be performed after 2 weeks. The investigation would be performed in female rats weighing 150-250 g. Blood samples would be obtained for determination of Natamycin plasma concentrations at 1 and 4 hours after the intravaginal drug application. All animals would be allowed free access to food and water. The Natamycin administration would be repeated on day 3, and again blood samples would be accrued to determine possible drug adsorption from the mucosal surface into the blood stream with this mode of application.
  • the antifungal treatment effect would be evaluated as the fraction of animals clearing the infection versus untreated control animals within a 7-day period.
  • the blood samples would be centrifuged at 1,000 x g for 10 min, and the plasma separated and stored at -80°C until assayed by HPLC.
  • Natamycin in Plasma The drug extraction with hexane and MeOH from plasma is essential to avoid interference from endogenous plasma components and to recover the maximum amount of drug.
  • Authentic chromatograms from blank plasma, Natamycin-spiked plasma, and one example of that obtained after extraction of a plasma sample from the proposed study would be expected to demonstrate that there is negligible adsorption of Natamycin to the blood after topical/vaginal application.
  • the Natamycin retention time in this system is 11.1-12.0 min.
  • the recovery of Natamycin with the above described technique is 91 ⁇ 5% when human plasma is spiked in vitro with 10 ⁇ g/ml of drug.
  • the assay is linear after drug extraction from plasma samples in the range from 50 ng/ml to 1.0 mg/ml. When a standard curve was prepared in the concentration range from 100 ng/ml to 25 ⁇ g/ml, a good correlation has been obtained between the plasma Natamycin concentration and peak AUC value:
  • the in vitro data demonstrates that the successful design of pharmaceutically acceptable topical formulations of Natamycin can be accomplished, ones that are compatible with ointment bases available for topical application, i.e. on the skin, or intravaginally. These formulations should be accepted with good tolerance and negligible toxicity, as could be demonstrated using the intravaginal route in a rodent model.
  • the topical application of one of the preparations in this rodent model should not provide any detectable plasma drug concentrations over a 4-hour observation interval, and should maintain fungicidal Natamycin concentrations without discernible untoward effects on the animals' clinical performance as detected by assessment of their infectious state after a 7-day observation period.
  • Novel vehicles have been invented for the stable, pharmaceutically acceptable formulation of Natamycin to make it safe to administer this drug topically as treatment of localized fungal infections
  • the inventors first established a sensitive and specific HPLC assay, which allowed the reproducible quantitation of Natamycin concentrations as low as 10-20 ng/ml.
  • the inventors developed extraction techniques to recover Natamycin in a reproducible manner from topical formulations and from blood plasma samples.
  • the stable and non-toxic novel Natamycin formulations will be useful as topical treatment of fungal infections in an experimental setting or as a prophylactic agent that can be given either topically (on the skin or intravaginally) or through nebuhzation.
  • a prophylactic agent that can be given either topically (on the skin or intravaginally) or through nebuhzation.
  • clinical studies of Natamycin's efficacy against localized infections are envisioned.
  • the present invention allows one to investigate the beneficial effects of fully solubilized Natamycin against localized infectious agents with the potential for a major improvement in the outcome of such infections.

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Abstract

L'invention concerne des formulations de pimaricine (également appelée natamycine) utiles pour le traitement et la suppression d'infections topiques, telles que celles engendrées par différents pathogènes, notamment des moisissures et de la levure, résistant aux composés azole et à l'amphotéricine B. L'invention concerne également des méthodes de traitement d'infections.
PCT/US2002/036616 2001-11-14 2002-11-14 Formulations topiques de natamycine/pimaricine WO2003041500A1 (fr)

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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN105067734A (zh) * 2015-07-21 2015-11-18 中国农业大学 一种葡萄酒中纳他霉素含量的测定方法

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6045815A (en) * 1997-08-15 2000-04-04 Board Of Regents, The University Of Texas System Parenteral pimaricin as treatment of systemic infections
US6239113B1 (en) * 1999-03-31 2001-05-29 Insite Vision, Incorporated Topical treatment or prevention of ocular infections
US20020177562A1 (en) * 2000-12-21 2002-11-28 Michael Weickert Pulmonary delivery of polyene antifungal agents

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6045815A (en) * 1997-08-15 2000-04-04 Board Of Regents, The University Of Texas System Parenteral pimaricin as treatment of systemic infections
US6239113B1 (en) * 1999-03-31 2001-05-29 Insite Vision, Incorporated Topical treatment or prevention of ocular infections
US20020177562A1 (en) * 2000-12-21 2002-11-28 Michael Weickert Pulmonary delivery of polyene antifungal agents

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN105067734A (zh) * 2015-07-21 2015-11-18 中国农业大学 一种葡萄酒中纳他霉素含量的测定方法

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