AU2013206401A1 - Antibiotic formulations, unit doses, kits, and methods - Google Patents

Abstract

An aqueous or powder composition includes anti-gram-negative antibiotic or salt thereof being present at an amount ranging from about 100 mg/ml to about 200 mg/ml. Another aqueous or powder composition includes anti-gram-positive antibiotic or salt thereof being present at a concentration ranging from about 0.6 to about 0.9 of the water solubility limit, at 25'C and 1.0 atmosphere, of the anti-gram-positive antibiotic or salt thereof Other embodiments include unit doses, kits, and methods. 4435277_2 (GHMatters) P77144.AU.1 ~ I 3 050:-:0 - - 1 .0 _ Fig. 15 0). 4 - -a F25o/60%RH - 1-- - d-4-2400C/75% RH 2 -TRS Spec NMT 10.0% 0 ... I .... I I I T=O 1M 3M 5M 6M 9M 12M iM Time points Fig. 16

Description

Antibiotic Formulations, Unit Doses, Kits, and Methods BACKGROUND OF THE INVENTION BACKGROUND [0011 The present application claims priority under 35 U.S.C. Q119(e) of U.S. Provisional Application No. 60/722,564, filed September 29, 2005, which is incorporated herein by reference in its entirety. Field of the Invention [0023 The present invention relates to anti-infective, such as antibiotic formulations, unit doses, kits, and methods, and in particular to aminoglycoside formulations, unit doses, kits, and methods Background of The Invention [003] The need for effective therapeutic treatment of patients has resulted in the development of a variety of phannaceutical formulation delivery techniques. One traditional technique involves the oral delivery of a phannaceutical formulation in the form of a pill, capsule, elixir, or the like. However, oral delivery can in some cases be undesirable, For example, many pharmaceutical formulations may be degraded in the digestive tract before the body can effect vely absorb them, Inhaleable drug delivery, where a patient orally or nasally inhales an aerosolized pharmaceutical formulation to deliver the formulation to the patient's respiratory tract, may also be effective and/or desirable, in one inhalation technique, an aerosolized pharmaceutical formulation provides local therapeutic treatment and/or propirylaxis to a portion of the respiratory tract, such as the lungs, to treat respiratory diseases such as asthma and emphysema and/or to treat local long infections, such as fungal infections and cystic fibrosis, In another inhalation technique, a pharmaceutical formulation is delivered deep within a patient's lungs where it may be absorbed into the bloodstream for systemic delivery of the formulation throughout the body, Many types of aerosolization devices exist including devices comprising a pharmaceutical formulation stored in or with a propellant, devices that aerosolize a powder, devices which use a compressed gas or other mechanism to aerosolize a liquid pharmaceutical formulation, and similar devices.

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[004] One known aerosolization device is commonly referred to as a nebulizer. A nebulizer imparts energy into a liquid pharmaceutical formulation to aerosolize the liquid, and to allow delivery to the pulmonary system, e.g. the lungs, of a patient. A nebulizer comprises a liquid delivery system, such as a container having a reservoir that contains a liquid phannaceutical formulation. The liquid pharmaceutical formulation generally comprises an active agent that is either in solution or suspended within a liquid medium. In one type of nebulizer, generally referred to as a jet nebulizer, compressed gas is forced through an orifice in the container. The compressed gas forces liquid to be withdrawn through a nozzle, and the withdrawn liquid mixes with the flowing gas to form aerosol droplets. A cloud of droplets is then administered to the patient's respiratory tract. In another type of nebulizer, generally referred to as a vibrating mesh nebulizer, energy, such as mechanical energy, vibrates a mesh. This vibration of the mesh aerosolizes the liquid pharmaceutical formulation to create an aerosol cloud that is administered to the patient's lungs. In still another type of nebulizer, ultrasonic waves are generated to directly vibrate and aerosolize the pharmaceutical formulation, [005] Nebulizers are often used to deliver (1) an aerosolized pharmaceutical formulation to a hospitalized or non-ambulatory patient; (2) large doses of aerosolized active agent and/or (3) an aerosolized pharmaceutical formulation to a child or other patient unable to receive a dry powder or propellant based pharmaceutical formulation. [006] Nebulizers are useful for delivering an aerosolized Pharmaceutical formulation to the respiratory tract of a patient who is breathing under the assistance of a ventilator. But there are problems associated with the introduction of aerosolized pharmaceutical formulation into ventilator circuits. For example, by introducing the aerosolized phannaceutical formulation into the inspiratory line of the ventilator, significant residence volume exists between the point of introduction and the patient's lungs. Accordingly, large amounts of aerosolized pharmaceutical formulation are needed and much of the fonnulation is lost to the exhalation line. This problem is exacerbated when the nebulizer is used in conjunction with ventilators having continual bias flows. In addition, the large residence volume in the ventilator line may dilute the aerosolized pharmaceutical formulation to an extent where the amount delivered to the patient is difficult to reproduce consistently. 2 [007] U.SA Published Application Nos. 2004/0011358, 2004/0035490, and 2004/0035413, which are incorporated herein by reference in their entireties, disclose methods, devices, and formulations for targeted endobronchiai therapy, Aerosolized antibiotics are delivered into a ventilator circuit, 'he aerosol generator, e.g., nebulizer, may be placed in the lower part of a Y-piece, for eample, distal to the Y, to be proximal to a patient airway and/or endotracheal tube. [008] U.S. Patent Nos. 5,508,269 and 6,890,907, which are incorporated herein by reference in their entireties, disclose aminoglycoside solutions for nebulization. . The '269 patent discloses that if the solution approaches the solubility of tobramycin, 160 mg/ml, precipitation on storage is expected. The '269 patent also discloses that a higher concentration of tobramycin than is clinically needed is economically disadvantageous. Further the '269 patent discloses that a more concentrated solution will increase the osrnolarity of the solution, thus decreasing the output of the formulation with both jet and ultrasonic nebulizers. The '269 patent discloses that the alternative of a more concentrated solution in a smaller total volume is also disadvantageous. The '269 patent further discloses that most nebulizers have a dead space volume of I ml, i.e, that of the last 1 ml of solution is wasted because the nebulizer is not performing. Therefore, while for example, a 2 nil solution would have 50% wastage, the 5 ml solution (the capacity of the nebulizer) has only 20% wastage. Additionally, the '269 patent discloses that since there is no sufficient aerosolization of the drug into the small particles, the drug in large particles or as a solution is deposited in the upper airways and induces cough and may also cause bronchospasm. According to the '269 patent, large aerosol particles also limit the drug delivery [009] There remains, however, a need for improved antibiotic formulations, such as antibiotic formulations for nebulization. There also remains a need for improved unit doses and kits of antibiotic formulations. Accordingly, there also remains a need for improved methods of making and/or using such antibiotic formulations. SUMMARY OF THE INVENTION [010] Accordingly, one or more embodiments of the present invention satisfies one or more of these needs. Thus the present invention provides antibiotic formulations, such as 3 antibiotic formulations for nebulization. The present invention also provides unit doses and kits of antibiotic fonnulations. The present invention further provides methods of making and/or using such antibiotic formulations. Other features and advantages of the present invention will be set forth in the description of invention that follows, and will be apparent, in part, from the description or may be learned by practice of the invention. The invention will be realized and attained by the devices and methods particularly pointed out in the written description and claims hereof. [011] In one aspect, one or more embodiments are directed to an aqueous composition, comprising an antibiotic or salt thereof being present at a therapeutic-effective (including prophylatic- effective) amount. In one or more embodiments, the therepautic effective amount is based upon aerosolized administration to the pulmonary system. [012] In one aspect, one or more embodiments are directed to an aqueous composition, comprising anti-gram-negative antibiotic or salt thereof being present at an amount ranging from about 90 mg/ml to about 300 mg/ml [0131 In another aspect, an aqueous composition comprises anti-gran-negative antibiotic or salt thereof, and optionally, a bronchodilator. [014] In still another aspect, an aqueous composition comprises anti-gram-positive antibiotic or salt thereof being present at a concentration ranging from about 0.6 to about 0.9 of the water solubility limit, at 250C and 1.0 atmosphere, of the anti-gram-positive antibiotic or salt thereof, [0151 In yet another aspect, a unit dose comprises a container and an aqueous composition, comprising anti-gram-negative antibiotic or salt thereof being present at a concentration ranging from about 100 mg/ml to about 200 mg/mL. [016] In still another aspect, a kit comprises a first container containing a first aqueous solution comprising an anti grai-negative antibiotic or salt thereof; and a second container containing a second aqueous solution comprising an anti gram-negative antibiotic or salt thereof. A concentration, or an amount, or both of the first aqueous solution is different from a concentration, or an amount, or both, of the second aqueous solution. [017] in yet another aspect, a kit comprises a first container containing a first aqueous solution comprising anti gram-negative antibot ic or salt thereof, and a second 4 container containing a second aqueous solution comprising anti gram-positive antibiotic or salt thereof [018] In another aspect, a unit dose comprises a container and a powder comprising an antibiotic or salt thereof, wherein the powder is present in an amount ranging from about 550 mg to about 900 mg. [019] In still another aspect, a unit dose comprises a container; and a powder comprising an antibiotic or salt thereof, wherein the powder is present in an amount ranging from about 150 mg to about 450 mg. [020] in yet another aspect, a kit comprises a first container containing a first composition comprising anti-gram-positive or an anti gram-negative antibiotic or salt thereof and a second container containing a second composition comprising water. The first composition and/or the second composition comprises osmolaiity adjuster. [0211 In another aspect, a kit comprises a first container containing a powder comprising anti-gram-positive antibiotic or salt thereof and a second container containing a powder comprising anti gram-positive antibiotic or salt thereof A concentration, or an amount, or both, of the anti gram-positive antibiotic or salt thereof in the first container is different from a concentration, or an amount, or both, of the anti gram-positive antibiotic or salt thereof in the second container. [022] In a further aspect, a kit comprises a first container containing a solution comprising anti gram-negative antibiotic or salt thereof and a second container containing a powder comprising anti gram-positive antibiotic or salt thereof [023] In still another aspect, a method of administering an antibiotic formulation to a patient in need thereof comprises aerosolizing an antibiotic formulation to administer the antibiotic formulation to the lungs of the patient. The antibiotic formulation has a concentration of antibiotic or salt thereof ranging from about 90 mg/ml to about 300 mg/mil. (024] In another aspect, a method of administering an antibiotic formulation to a patient in need thereof comprises inserting a tube into a trachea of a patient. The method also comprises aerosolizing an antibiotic formiulation to administer the antibiotic fonnulation to the lungs of the patient. The antibiotic formulation consists essentially of anti-gram-negative antibiotic or salt thereof and water. 5 [025] In yet another aspect, a method of administering an antibiotic formulation to a patient in need thereof comprises aerosolizing an antibiotic formulation to administer the antibiotic formulation to the lungs of the patient. The antibiotic formulation comprises an antibiotic or salt thereof at a concentration ranging from about 0.7 to about 0.9 of the water solubility limit, at 254C and 1.0 atmosphere, of the antibiotic or salt thereof, [026] In a further aspect, a method of administering an antibiotic fonnulation to a patient in need thereof comprises dissolving an antibiotic or salt thereof in a solvent to form an antibiotic formulation, wherein the antibiotic or salt thereof is present at a concentration ranging from about 0.6 to about 0.9 of the water solubility limit, at 25*C and 1.0 atmosphere, of the antibiotic or salt thereof The method also includes aerosolizing the antibiotic formulation to administer the antibiotic formulation to the lungs of the patient. [027] In yet another aspect, a method of administering an antibiotic formulation to a patient in need thereof comprises dissolving an antibiotic or salt thereof in a solvent to form an antibiotic formulation. The method also includes aerosolizing the antibiotic formulation to administer the antibiotic formulation to the lungs of the patient, wherein the aerosolizing is conducted within about 16 hours of the dissolving, (028] In another aspect, a method involves fonning a powder comprising an antibiotic or salt thereof The method includes dissolving an antibiotic or salt thereof in a solvent to forn a solution having a concentration ranging from about 60 mg/ml to about 120 mg/ml. The method also includes lyophilizing the solution to form the powder. [029] In another aspect, a method involves forming a powder comprising an antibiotic or salt thereof Thlre method comprises dissolving an antibiotic or salt thereof in a solvent to form a solution having a volume ranging from about 4.5 ml to about 5.5 ml. The method also includes lyophilizing the solution to form the dry powder, [030] In another aspect, any method which comprises forming a powder may also include a method of reconstituting the powder to form a liquid, Similarly any method which comprises forming a liquid comprising an antibiotic (such as a solution) may also include a method of removing the liquid to yield a powder. [0311 In another aspect, any two or more of any of the foregoing features, aspects versions or embodiments are combined. 6 BRIEF DESCRIPTION OF THE DRAWINGS [032] The present invention is further described in the description of invention that follows, in reference to the noted plurality of non-limiting drawings, wherein: [033] Fig, 1A illustrates components of a pulmonary drug delivery system according to embodiments of the present invention. [034] Fig, 1B shows an embodiment of a device that can be used in a pulmonary drug delivery system according to embodiments of the invention. [035] Fig. 2A shows an exemplary off-ventilator configuration of a pulmonary drug delivery system according to embodiments of the invention. [036] Fig. 2B is a schematic view of an pharmaceutical delivery device of one or more embodiments of the present invention, useful for delivery of aerosolized medicaments. [037] Fig. 3 shows total drug recovered (nebulizer + filters) for gentamicin as a function of fill mass and solution strength. [038] Figs. 4a-b show emitted dose of gentamicin as a function of solution strength and fill volume, after nebulization (Fig. 4a) for 15 minutes, and (Fig. 4) 30 minutes. [039] Fig. 5 shows gentarnicin residual dose retained in a nebulizer as a function of fill volume and solution strength, [040] Fig. 6 shows distribution of nebulized vancomycin (60 mg/ml solution in normal saline) as a function of fill volume. [041] Fig. 7 shows emitted dose as a function of solution strength and fill volume, for the case of vancomycin solution in 0.45% saline. [042] Fig, 8 shows emitted dose as a function of solution strength and fill volume, for the case of vancomycin solution in water for injection (WFI). [043] Fig. 9 shows volume median diameter for nebulized gentamicin as a function of solution strength and fill volume. [044] Fig. 10 shows cumulative particle size distributions for gentamicin at different solution strengths and nebulizer fill volumes. [045] Fig. 11 shows volume median diameter for nebulized vancomycin (solution in WFI) as a function of solution strength and fill volume. 7 [046] Fig. 12 shows cumulative particle size distributions for nebulized vancomycin (solution in WF) at different solution strengths and nebulizer fill volumes [047] Fig. 13 shows volume median diameter for nebulized vancomycin (60 mg/ml solution in normal saline) as a function of nebulizer fill volume. [048] Fig. 14 shows volume median diameter for nebulized vancomycin (solution in 0.45% saline) as a function of solution strength and fill volume. [049] Fig. 15 shows volume median diameter for antibiotic drug and placebo solutions. [050] Fig 16 is a graph showing amikacin stability over time (as % related substance) for a formulation according to one or more embodiments of the present invention, wherein the formulation was stored at three different storage conditions. DESCRIPTION OF THE INVENTION [051] Unless otherwise stated, a reference to a compound or component includes the compound or component by itself, as well as in combination with other compounds or components, such as mixtures of compounds, [052] As used herein, the singular forms "a," "an," and "the" include the plural reference unless the con t ext clearly dictates otherwise. [053] Reference herein to "one embodment", "one version" or "one aspect" shall include one or more such embodiments, versions or aspects, unless otherwise clear from the context. [054] "Mass median diameter" or "MMD" is a measure of mean particle size, since the powders of the invention are generally polydisperse (i.e., consist of a range of particle sizes). MMD values as reported herein are determined by centrifugal sedimentation, although any number of commonly employed techniques ca. be used for measuring mean particle size. [055] "Mass median aerodynamic diameter" or "MMAD" is a measure of the aerodynamic size of a dispersed particle. The aerodynamic diameter is used to describe an aerosolized powder in terms of its settling behavior, and is the diameter of a unit density sphere having the same settling velocity, generally in air, as the particle. The aerodynamic diameter encompasses particle shape, density and physical size of a particle. As used herein, 8 MMAD refers to the midpoint or median of the aerodynamic particle size distribution of an aerosolized powder determined by cascade impaction. [056] Anti-gram negative, and gram-negative antibiotic are used interchangeably to refer to antibiotic active agents (and formulations comprising such active agents) which have effectiveness against gram negative bacteria, Similarly, anti-gram positive, and gram positive antibiotic are used interchangeably to refer to antibiotic active agents (and formulations comprising such active agents) which have effectiveness against gram positive bacteria. [057] "Antibiotic" moreover includes anti-infectives, such as antivirals and antifungals, as well as antibiotics, unless the context indicates otherwise. [058] "Pharmtaceutic formulation" and "composition" may be sometimes used interchangeably to refer to a formulation comprising an antibiotic. [059] As an overview, in one or more embodiments, an aqueous composition comprises anti-gram-negative and/or anti-gram positive antibiotic or salt thereof being present at an amount ranging from about 100 mg/ml to about 200 mg/mI. [060] In one or more embodiments, an aqueous composition comprises an antibiotic or salt thereof, and bronchodilator. 1061] In one or more embodiments, an aqueous composition comprises an antibiotic or salt thereof being present at a concentration ranging from about 0.6 to about 0.9 of the water solubility limit, at 250C and 1.0 atmosphere, of the antibiotic or salt thereof. [062] In one or more embodiments, a unit dose comprises a container and an aqueous composition, comprising an anti-gram-negative antibiotic or salt thereof at a concentration ranging from about 100 mg/mil to about 200 mg/ml.

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0 6 3 ] In one or more embodiments, a kit comprises a first Container containing a first aqueous solution comprising anti-gram-negative antibiotic or salt thereof; and a second container containing a second aqueous solution comprising anti-gram-negative antibiotic or salt thereof. A concentration, or an amount, or both, of the first aqueous solution is different from a concentration, or an amount, or both, of the second aqueous solution. [064] In one or more embodiments, a kit comprises a first container containing a first aqueous solution comprising anti-gram-negative antibiotic or salt thereof, and a second 9 container containing a second aqueous solution comprising anti-gram-positive antibiotic or salt thereof [065] In one or more embodiments, a unit dose comprises a container and a powder comprising an antibiotic or sat thereof, wherein the powder is present n an amount ranging from about 550 mg to about 900 mg. [066] In one or more embodiments, a unit dose comprises a container; and a powder comprising an antibiotic or salt thereof, wherein the powder is present in an amount ranging from about 150 ng to about 450 mg. [067] In one or more embodiments, a kit comprises a first container containing a first composition comprising an anti-gram-positive or an anti gran-negative antibiotic or salt thereof and a second container containing a second composition comprising water. The first composition and/or the second composition comprises an osmolality adjuster. [068] In one or more embodiments, a kit comprises a first container containing a powder comprising an anti-gram-positive antibiotic or salt thereof and a second container containing a powder comprising an anti-grarn-positive antibiotic or salt thereof. A concentration, or an amount, or both, of the anti-gram-positive antibiotic or salt thereof in the first container is different from a concentration, or an amount, or both, of the anti-gram positive antibiotic or salt thereof in the second container. [069] In one or more embodiments, a kit comprises a first container containing a solution comprising an anti-gram-negative antibiotic or salt thereof and a second container containing a powder comprising anti-gram-positive antibiotic or salt thereof [070] in one or more embodiments, a method of administering an antibiotic formulation to a patient in need thereof comprises aerosolizing an antibiotic formulation to administer the antibiotic formulation to the pulmonary system of the patient. The antibiotic fornalation has a concentration of anti-gram-negative antibiotic or salt thereof ranging from about 100 mg/ml to about 200 mg/ml. [071] In one or more embodiments, a method of administering an antibiotic formulation to a patient in need thereof comprises inserting a tube into a trachea of a patient, The method also comprises aerosolizing an antibiotic formulation to administer the antibiotic formulation to the pulmonary system of the patient. The antibiotic formulation consists essentially of an anti-gram-negative antibiotic or salt thereof and water. 10 [072] In one or more embodiments, a method of administering an antibiotic formiulation to a patient in need thereof comprises aerosolizing an antibiotic formulation to administer the antibiotic formulation to the pulmonary system of the patient. The antibiotic formulation comprises an anti-gram-positive antibiotic or salt thereof at a concentration ranging from about 0.7 to about 0.9 of the water solubility limit, at 25*C and 1.0 atmosphere, of the anti-gram-positive antibiotic or salt thereof. [073] In one or more embodiments, a method of administering an antibiotic formulation to a patient in need thereof comprises aerosolizing an antibiotic fonnulation using a vibrating mesh nebulizer, and administering the antibiotic formulation to the pulmonary system of the patient via an endotracheal tube, wherein the nebulizer is positioned in close proximity to the endotracheal tube. [074] In one or more embodiments, a method of administering an antibiotic formulation to a patient in need thereof comprises dissolving an anti-grain-positive antibiotic or salt thereof in a solvent to form an antibiotic formulation, wherein the anti-gram-positive antibiotic or salt thereof is present at a concentration ranging from about 0.6 to about 0,9 of the water solubility limit, at 25"C and 1.0 atmosphere, of the anti-grain-positive antibiotic or salt thereof The method also includes aerosolizing the antibiotic formulation to administer the antibiotic formulation to the pulmonary system of the patient. [0751 In one or more embodiments, a method of administering an antibiotic formulation to a patient in need thereof comprises dissolving an antibiotic or salt thereof in a solvent to form an antibiotic formulation. The method also includes aerosolizing the antibiotic formulation to administer the antibiotic formulation to the pulmonary system of the patient, wherein the aerosolizing is conducted within about 16 hours of the dissolving. [076] In one or more embodiments, a method involves forming a powder comprising an antibiotic or salt thereof The method includes dissoi\ ng an antibiotic or salt thereof in a solvent to form a solution having a concentration ranging from about 60 mg/hl to about 120 mg/mI, The method also includes lyophilizing the solution to form the powder, [077] In one or more embodiments, a method involves forming a powder comprising an antibiotic or salt thereof The method comnprises dissolving an antibiotic or salt thereof in a solvent to forn a solution having a volume ranging from about 4,5 ml to about 5.5 ml. The method also includes lyophilizing the solution to form the dry powder. 11 [078] Therefore, in one or more embodiments, the present invention involves concentrated antibiotic formulations. The antibiotic formulations may comprise an aqueous composition of antibiotic or salt thereof being present at a concentration ranging from about 0.6 to about 0,9, such as about 03 to about 0.8, of the water solubility limit, at 250C and 1.0 atmosphere, of the antibiotic or salt thereof [079] The concentration of the antibiotic, corrected for potency, in one or more embodiments, may range fnom about 40 mg/ml to about 200 mg/ml, such as about 60 mg/mi to about 140 mg/ml, or about 80 mg/ml to about 120 mg/mL. For example, in the case of anti gram-negative antibiotics or salts thereof, the concentration as corrected for potency may range from about 40 mg/ml to about 200 mg/ml, such as from about 90 mg/ml to about 200 mg/ml, about 110 mg/mil to about 150) mg/mil, or about 120 mg/il to about 140 ng/ml. As another example, in the case of anti-gram-posiYive antibiotics or salts thereof the concentration as corrected for potency may range from about 60 mg/mi to about 140 mg/ml, such as about 80 mg/ml to about 120 mg/ml. [0801 The aqueous compositions typically have a pH that is compatible with physiological administration, such as pulmonary administration. For example, the aqueous composition may have a pH ranging from about 3 to about 7, such as about 4 to about 6, [081] In addition, the aqueous compositions typically have an osmolality that is compatible with physiological administration, such as pulmonary administration. in one or more embodiments, the aqueous composition may have an osmolality ranging from about 90 mOsmnol/kg to about 500 mnOsmol/kg, such as 120 mOsmol/kg to about 500 mOsmoi/kg, or about 150 msmol/kg to about 300 mOsmolkg. [082] In one or more embodiments, the aqueous compositions are stable. For instance, in some cases, no precipitate forms in the aqueous composition when the aqueous composition is stored for 1 year, or even 2 years, at 25 *C. [083) The potency of the antibiotic or salt thereof may range from about 500 pg/mg to about 1100 pg/mg. In one or more embodiments, the potency of anti-gram-negative antibiotics or salts thereof, such as gentanicin, typically ranges from about 500 g/ng to about 1100 pg/mg, such as about 600 g/mg to about 1000 pg/mg, or about 700 pg/mg to about 800 pg/mg. The potency of anti-gram-positive antibiotics or salts thereof, such as 12 vancomycin, typically ranges from about 900 ug/mg to about 1100 pg/rg, such as from about 950 pg/mg to about 1050 kg/mg, [084] The chromatographic purity level of the antibiotic or salt thereof typically greater than about 80%, such as greater than about 85%, greater than about 90%, or greater than about 95%. In this regard, there is generally no major impurity greater than about 10%, such as no greater than about 5% or no greater than about 2%. For instance, the amount of heavy metals is typically less than about 0.005 wt%, such as less than about 0.004 wt%, less than about 0.003 wt%, less than about 0,002 wt%, or less than about 0.001 wt%. [085] In the case of gentamicin, the compositions typically have a gentamicin C content ranging from about 25% to about 50%, such as about 30% to about 55%, about 35% to about 50%, or about 40% to about 45%, based on the total amount of gentamicin. The compositions typically have a gentamicin C 1 1 content ranging from about 10% to about 35%, such as about 15% to about 30%, about 20% to about 25%, based on the total amount of gentamicin. The compositions typically have a gentamicin C 2 and Cr content ranging from about 25 wt% to about 55 wt%, such as about 30% to about 50%, about 30% to about 45%, or about 35% to about 40%, based on the total amount of gentamicin. [0861 In embodiments of the present invention comprising anikacin, the compositions typically have an amikacin content ranging from about 25% to about 50%, such as about 30% to about 55%, about 35% to about 50%, or about 40% to about 45%, based on the total amount of amikacin. [087] Nearly any anti-gram-negative, anti-gram-positive antibiotic, or combinations thereof may be used. Additionally, antibiotics may comprise those having broad spectrum effectiveness, or mixed spectrum effectiveness, Antifungals, such as polyene materials, in particular, amphotericin B are also suitable for use herein. Examples of anti-gram-negativc antibiotics or salts thereof include, but are not limited to, aminoglycosides or salts thereof. Examples of aminoglycosides or salts thereof include gentamicin, amikacin, kanamycin, streptomycin, neomycin, netimicin, paramecin, tobramycin, salts thereof, and combinations thereof. For instance, gentamicin sulfate is the sulfate salt, or a mixture of such salts, of the antibiotic substances produced by the growth of sicrnomonosporapurpurea. Gentamicin sulfate, USP, may be obtained from Fujian Fukang Pharmaceutical Co., LTD, Fuzhou, China. 13 Amikacin is typically supplied as a sulfate salt, and can be obtained, for example, from Bristol-Myers Squibb, Amikacin may include related substances such as kanamicin. [088] Examples of anti-gram-positive antibiotics or salts thereof include, but are not limited to, macrolides or salts thereof. Examples of macrolides or salts thereof include, but are not limited to, vancomycin, erythromycin, clarithromycin, azithromycin, salts thereof, and combinations thereof. For instance, vancomycin hydrochloride is a hydrochloride salt of vancomycin, an antibiotic produced by certain strains of Amycolatopsis orientalis, previously designated Streptonyc.es orientalis. Vancomycin hydrochloride is a mixture of related substances consisting principally of the monohydrochloride of vancomycin B, Like all glycopeptide antibiotics, vancomycin hydrochloride contains a central core heptapeptide. Vancomycin hydrochloride, USP, may be obtained from Alpharma, Copenhagen, Denmark. [089] In some embodiments, the composition comprises an antibiotic and one or more additional active agents. The additional active agent described herein includes an agent, drug, or compound, which provides some pharmacologic, often beneficial, effect This includes foods, food supplements, nutrients, drugs, vaccines, vitamins, and other beneficial agents. As used herein, the tennis further include any physiologically or pharmacologically active substance that produces a localized or systemic effect in a patient An active agent for incorporation in the pharmaceutical formulation described herein may be an inorganic or an organic compound, including, without limitation, drugs which act on: the peripheral nerves, adrenergic receptors, cholinergic receptors, the skeletal muscles, the cardiovascular system, smooth muscles, the blood circulatory system, synoptic sites, neuroeffector junctional sites, endocrine and hormone systems, the immunological system, the reproductive system, the skeletal system, autacoid systems, the alimentary and excretory systems, the histamine system, and the central nervous system, [090] Examples of additional active agents inchde, but are not limited to, anti inflaxnmatory agents, bronchodilators, and combinations thereof, [091] Examples of bronchodilators include, but are not limited to, P-agonists, anti muscarinic agents, steroids, and combinations thereof For instance, the steroid may comprise albuterol, such as albuterol sulfate. [092] Active agents may comprise, for example, hypnotics and sedatives, psychic energizers, tranquilizers, respiratory drugs, anticonvulsants, muscle relaxants, antiparkinson 14 agents (dopamine antagnonists), analgesics, anti-inflammatories, antianxiety drags (anxiolytics), appetite suppressants, antimigraine agents, muscle contractants, additional anti infectives (antivirals, antifungals, vaccines) antiarthritics, antimalarials, antiernetics, anepileptics, cytokines, growth factors, anti-cancer agents, antithrombotic agents, antihypertensives, cardiovascular drugs, antiarrhythmics, antioxicants, anti-asthma agents, hormonal agents including contraceptives, sympathomimetics, diuretics, lipid regulating agents, antiandrogenic agents, antiparasitics, anticoagulants, neoplastics, antineoplastics, hypoglycemics, nutritional agents and supplements, growth ments, ents, antienteritis agents, vaccines, antibodies, diagnostic agents, and contrasting agents. The active agent, when administered by inhalation, may act locally or systemically, [093] The active agent may fall into one of a number of structural classes, including but not limited to small molecules, peptides, polypeptides, proteins, polysaccharides, steroids, proteins capable of eliciting physiological effects, nucleotides, oligonucleotides, polynucleotides, fats, electrolytes, and the like. [094] Examples of active agents suitable for use in this invention include but are not limited to one or more of calcitonin, amphotericin B, erythropoietin (EPO), Factor VIII, Factor IX, ceredase, cerezyme, cyclosporin, granulocyte colony stimulating factor (GCSF), thrombopoietin (TPO), alpha-I proteinase inhibitor, elcatonin, granulocyte macrophage colony stimulating factor (GMCSF), growth hormone, human growth hormone (HGH), growth hormone releasing hormone (GHRH), heparin, low molecular weight heparin (LMWAH), interferon alpha, interferon beta, interferon gamma, interleukin-l receptor, interleukin-2, interleukin-I receptor antagonist, interleukin-3, interleuidn-4, interleukin-6, luteinizing hormone releasing hormone (LHRH), factor X, insulin, pro-insulin, insulin analogues (e.g., mono-acylated insulin as described in U.S. Patent No. 5,922,675, which is incorporated herein by reference in its entirety), amylin, C-peptide, somatostatin, somatostatin analogs including octreotide, vasopressin, follicle stimulating hormone (FSH), insulin-like growth factor (IGF), insulintropin, macrophage colony stimulating factor (M CSF), nerve growth factor (NOF), tissue growth factors, keratinocyte growth factor (KGF), glial growth factor (GOF), tumor necrosis factor (TNF), endothelial growth factors, parathyroid hormone (PTH), glucagon-like peptide thymosin alpha 1, Ub/,ha inhibitor, alpha-1 antitrypsin, phosphodiesterase (PDE) compounds, VLA-4 inhibitors, bisphosponates, 15 respiratory syncytial virus antibody, cystic fibrosis traismembrane regulator (CFTR) gene, deoxyreibonuclease (Dnase), bactericidal/penneability increasing protein (BPI), anti-CMV antibody, 13-cis retinoic acid, oleandomycin, troleandomycin, roxitbromycin, clarithromycin, davercin, azithromycin, flurithromycin, dirithromycin, josanycin, spironycin, midecanycin, leucomycin, miocamycin, rokitamycin, andazithronycin, and swinolide A; fluoroquinolones such as ciprofloxacin, ofloxacin, levofloxacin, trovafloxacin, alatrofloxacin, moxifloxicin, norfloxacin, enoxacin, grepafloxacin, gatifloxacin, lomefloxacin, sparfloxacin, tenafloxacin, pefioxacin, amifloxacin, fleroxacin, tosufloxacin, prulifloxacin, irloxacin, pazufloxacin, clinafioxacin, and sitafloxacin, teicoplanin, rampolanin, mnideplanin, colistn, daptomycin, gramicidin, colistimethate, polymixins such as polynixin B, capreomycin, bacitracin, penems; penicillins including penicllinase-sensitive agents like penicillin G, penicillin V, penicillinase-resistant agents like methicillin, oxacillin, cloxacillin, dioloxacillin, floxacillin, nafeillin; gram negative microorganism active agents like ampicillin, amoxicillin, and hetacillin, cillin, and galanpicillin; antipseudornonal penicillins like carbenicillin, ticarcillin, azlocillin, meziocillin, and piperacillin; cephalosporins like cefyodoxine, cefprozil, ceftbuten, ceftizoxime, ceftriaxone, cephalothin, cephapirin, cephalexin, cephradrine, cefoxitin, cefamandole, cefazolin, cephaloridine, cefaclor, cefadroxil, cephaloglycin, cefuroxime, ceforanide, cefotaxime, cefatrizine, cephacetrile, cefepime, cefixime, cefonicid, cefoperazone, cefotetan, cefinetazole, ceftazidime, loracarbef and moxalactan, monobactans like aztreonam; and carbapenerns such as imipenem, ineropenern, pentanidine isethiouate, lidocaine, metaproterenol sulfate, beclomethasone diprepionate, triamncinolone acetamide, budesonide acetonide, fluticasone, ipratropiun bromide, flunisolide, cromolyn sodium, ergotamine tartrate and where applicable, analogues, agonists, antagonists, inhibitors, and pharmaceutically acceptable salt forms of the above. In reference to peptides and proteins, the invention is intended to encompass synthetic, native, glycosylated, unglycosylated, pegylated fonns, and biologically active fragments, derivatives, and analogs thereof. [095] Active agents for use in the invention further include nucleic acids, as bare nucleic acid molecules, vectors, associated viral particles, plasmid DNA or RNA or other nucleic acid constructions of a type suitable for transfection or transformation of cells, i.e., suitable for gene therapy incuding antisense. Further, an active agent may comprise live 16 attenuated or killed viruses suitable for use as vaccines. Other useful drugs include those listed within the Physician's Desk Reference (most recent edition), which is incorporated herein by reference in its entirety. {096) The amount of antibiotic or other active agent in the pharmaceutical formulation will be that amount necessary to deliver a therapeutically or prophylactically effective amount of the active agent per unit dose to achieve the desired result. In practice, this will vary widely depending upon the particular agent, its activity, the severity of the condition to be treated, the patient population, dosing requirements, and the desired therapeutic effect. The composition will generally contain anywhere from about I wt% to about 99 wt%, such as from about 2 wt% to about 95 wt%, or from about 5 wt% to 85 wt%, of the active agent, and will also depend upon the relative amounts of additives contained in the composition. The compositions of the invention are particularly useful for active agents that are delivered in doses of ftom 0.001 ig/day to 100 mg/day, such as in doses from 0,01 mg/day to 75 mg/day, or in doses from 0.10 mg/day to 50 mg/day, It is to be understood that more than one active agent may be incorporated into the formulations described herein and that the use of the term "agent" in no way excludes the use of two or more such agents. [097] Generally, the compositions are free of excessive excipients. In one or more embodiments, the aqueous composition consists essentially of the anti-gram-negative antibiotic, such as amikacin, or gentarnicin or both, and/or salts thereof and water, [098] Further, in one or more embodiments, the aqueous composition is preservative-free. In this regard, the aqueous composition may be methylparaben-free and/or propylparaben-free. Still further, the aqueous composition may be saline-free. [099] In one or more embodiments, the compositions comprise an anti-infective and an excipient. The compositions may comprise a pharmaceutically acceptable excipient or carrier which may be taken into the lungs with no significant adverse toxicological effects to the subject, and particularly to the lungs of the subject. In addition to the active agent, a pharmaceutical formulation may optionally include one or more pharmaceutical excipients which are suitable for pulmonary administration. These excipients, if present, are generally present in the composition in amounts sufficient to perform their intended function, such as stability, surface modification, enhancing effectiveness or delivery of the composition or the like, Thus if present,excipient may range from about 0.01 wt% to about 95 wt%, such as 17 from about 0,5 wt% to about 80 wt%, from about I wt% to about 60 w%. Preferably, such excipients will, in part, serve to further improve the features of the acti-ve agent composition, for example by providing more efficient and reproducible delivery of the active agent and/or facilitating manufacturing. One or more excipients may also be provided to serve as bulking agents when it is desired to reduce the concentration of active agent in the formulation. [0100] For instance, the compositions may include one or more osmolality adjuster, such as sodium chloride, For instance, sodium chloride may be added to solutions of vancomycin hydrochloride to adjust the osmolality of the solution. In one or more embodiments, an aqueous composition consists essentially of the anti-gram-positive antibiotic, such as vancomycin hydrochloride, the osimolality adjuster, and water. [0101] Pharmaceutical excipients and additives useful in the present phannaceutical formulation include but are not limited to amino acids, peptides, proteins, non-biological polymers, biological polymers, carbohydrates, such as sugars, derivatized sugars such as alditols, aldonic acids, esterified sugars, and sugar polymers, which may be present singly or in combination. 1 01021 Exemplary protein excipients include albumins such as human serum albumin (HSA), recombinant human albumin @(-HA), gelatin, casein, hemoglobin, and the like, Suitable amino acids (outside of the dileucyl-peptides of the invention), which may also function in a buffering capacity, include alanine, glycne., arginine, betaine, histidine, glutamic acid, aspartic acid, cysteine, lysine, leucine, isoleucine, valine, methionine, phenylalanine, aspartame, tyrosine, tryptophan, and the like. Preferred are amino acids and polypeptides that function as dispersing agents, Amnino acids falling into this category include hydrophobic amino acids such as leucine, valine, isoleucine, tryptophan, alanine, methionine, phenylalanine, tyrosine, histidine, and proline. [0103) Carbohydrate excipients suitable for use in the invention include, for example, monosaccharides such as fructose, maltose, galactose, glucose, D-mannose, sorbose, and the like; disaccharides, such as lactose, sucrose, trehalose, cellobiose, and the like; polysaccharides, such as raffinose, melezitose, maltodextrins, dextrans, starches, and the like; and alditols, such as mannitol, xyitol, maltitol, lactitol, xylitol sorbitol (glucitol), pyTanosyl sorbitol, myoinositol and the like. 18 [01043 The pharmaceutical formulation may also comprise a buffer or a pH adjusting agent, typically a salt prepared from an organic acid or base. Representative buffers comprise organic acid salts of citric acid, ascorbic acid, gluconic acid, carbonic acid, tartaric acid, succinic acid, acetic acid, or phthalic acid, Tris, tromethamine hydrochloride, or phosphate buffers. [0105] The pharmaceutical formulation may also include polymeric excipients/additives, e.g., polyvinyipyrrolidones, celluloses and derivatized celluloses such as hydroxymethyleellulose, hydroxyethyicellulose, and hydroxvpropylmethycellulose, Ficolls (a polymeric sugar), hydroxyethylstarch, dextrates (e.g., cyclodextrins, such as 2 hydroxypropylf-cyclodextrin and sulfobutylether-p-cyclodextrin), polyethylene glycols, and pectin. [0106] The pharmaceutical formulation may further include flavoring agents, taste msking agents, inorganic salts (for example sodium chloride), antimicrobial agents (for example benzalkonium chloride), sweeteners, antioxidants, antistatic agents, surfactants (for example polysorbates such as "TVEEN 20" and "TWEEN 80"), sorbitan esters, lipids (for example phospholipids such as lecithin and other phosphatidylcholines. phospIatidylethanolaimines), fatty acids and fatty esters, steroids (for example cholesterol), and chelating agents (for example EDTA, zinc and other such suitable cations). Other pharmaceutical excipients and/or additives suitable for use in the compositions according to the invention are listed in "Remington: The Science & Practice of Pharmacy", 19 ed,, Williams & Williams, (1995), and in the "Physician's Desk Reference", 52") ed., Medical Economics, Montvale, NJ (1998), both of which are incorporated herein by reference in their entireties, [0107] For MDI applications, the pharmaceutical fornuiation may also be treated so that it has high stability. Several attempts have dealt with improving suspension stability by increasing the solubility of surface-active agents in the FA propellants. To this end U.S. Patent No. 5,118,494, WO 91/11173 and WO 92/00107 disclose the use of HFA soluble fluorinated surfactants to improve suspension stability. Mixtures of HFA propellants with other perfluorinated cosolvents have also been disclosed as in WO 91/04011. Other attempts at stabilization involved the inclusion of nonfluorinated surfactants. In this respect, U.S. Patent No, 5,492,688 discloses that some hydrophilic surfactants (with a 19 hydrophilic/lipophilic balance greater than or equal to 9.6) have sufficient solubility.. in HFAs to stabilize medicament suspensions. Increases in the solubility of conventional nonfluorinated MDI surfactants (e.g. oleic acid, lecithin) can also reportedly be achieved with the use of co-solvents such as alcohols, as set forth in U.S. Pat, Nos. 5,683,677 and 5,605,674, as well as in WO 95/17195. A particularly useful class of MDIs are those which use bydrofluoroalkane (HFA) propellants. The HFA propellants are further particularly well suited to be used with stabilized dispersions of an active agent such as fornulations and composition of aminoglycoside antibiotics, Suitable propellants, fonnulations, dispersions, methods, devices and systems comprise those disclosed in US 6,309,623, the disclosure of which is incorporated by reference in its entirety. All of the aforementioned references being incorporated herein by reference in their entireties. [0108] In one or more embodiments, the compositions comprise an aerosol having a particle or droplet size selected to permit penetration into the alveoli of the lungs, Sich as a mass median aerodynamic diameter, less than about 10 tm, less than about 7,5 rum, less than about 5 rn, and usually being in the range of about 0,1 n to about 5 pm. [0109] The compositions of the present invention may be made by any of the various methods and techniques known and available to those skilled in the art. In this regard, procedures such as lyophilizing antibiotics to make powders and/or dissolving antibiotics in solvents are known in the art, 0110] For instance, a solution of antibiotic, e.g., amikacin sulfate or gentamicin sulfate, may be made using the following procedure. Typically, manufacturing equipment is sterilized before use. A portion of the final volume, e,g., 70%, of solvent, e.g, water for injection, may be added into a suitable container. Antibiotic or salt thereof may then be added. The antibiotic or salt thereof may be mixed until dissolved. Additional solvent may be added to make up the final batch volume. The batch may be filtered, e.g., through a 0.2 pm filter into a sterilized receiving vessel. Filling components may be sterilized before use in filling the batch into vials, e.g, 10 ml vials. [0111] As an example, the above-noted sterilizing may include the following. A 5 liter type I glass bottle and lid may be placed in an autoclave bag and sterilized at elevated temperature, e.g., 1214C for 15 minutes, using an autoclave. Similarly, vials may be placed into suitable racks, inserted into an autoclave bag, and sterilized at elevated temperature, e.g., 20 121C for 15 minutes, using an autoclave. Also similarly, stoppers may be placed in an autoclave bag and sterilized at elevated temperature, e.g., 121*C for 15 minutes, using an autoclave. Before sterilization, sterilizing filters may be attached to tubing, e,g., a 2 mm length of 7 mm x 13 mm silicone tubing. A filling line may be prepared by placed in an autoclave bag and sterilized at elevated temperature, e.g., 121'C for 15 minutes, using an autoclave. [0112] The above-noted filtration may involve filtration into a laminar flow work area. The receiving bottle and filters may be set up in the laminar flow work area. [0113] The above-noted filling may also be conducted under laminar flow protection. The filling line may be unvrapped and placed into the receiving bottle, The sterilized vials and stoppers may be unwrapped under laminar flow protection. Each vial may be filled, e.g., to a target fill of 5.940 g, and stoppered. A flip off collar may be applied to each vial. The sealed vials may be inspected for vial leakage, correct overseals, and cracks, [0114] As another example, one or more antibiotics, e.g., vancomycin, gentamicin or amikacin, and/or a salt thereof, may be prepared by lyophilizing the antibiotic to form a powder for storage. The powder is then reconstituted prior to use. This technique may be used when the antibiotic is unstable in solution. [0115] In one or more embodiments, the powder making process may begin with forming a solution to be lyophilized, For example, an antibiotic or salt thereof, such as amikacin, gentamicin or vancomycin and/or salts thereof, may be dissolved in a solvent to form a solution having an antibiotic concentration ranging from about 80 mg/ml to about 150 mg/ml, such as about 90 mg/ml to about 130 mg/mi, or about 100 mg/mI to about 124 mg/ml. The solution to be lyophilized may have a volume ranging from about 4,5 ml to about 5.5 ml, such as about 5 ml [0116] In other embodiments, the powder making process may begin with forming a solution of an anti-gramn negative antibiotic or salt thereof, such as amnikacin or salt thereof The antibiotic and/or salt may be dissolved in a solvent to form a solution having a concentration ranging from about 80 mg/ml to about 130 mg/ml, such as about 90 mg/ml to about 120 mg/mi, or about 100 mg/mi to about 110 mg/ml. The solution to be lyophilized may have a volume ranging from about 4.5 ml to about 5.5 ml, such as about 5 ml. 21 [0117] The solvent for the solution to be lyophilized may comprise water. The solution may be excipient-free. For instance, the solution may be cryoprotectant-free, [0118] In one or more embodiments, a suitable amount (e.g, 120 g per liter of final solution) of drug substance (for example vancomycin hydrochloride) may be dissolved, e.g., in about the 75% of the theoretical total amount of water for injection under nitrogen bubbling. The dissolution time may be recorded and appearance may be evaluated, [01193 Then, the dilution to the final volume with WFI may be carried out. Final volume may be checked. Density, pH, endotoxin, bioburden, and content by UV may be measured both before and after sterile filtration. [01201 The solution may be filtered before lyophilizing. For instance, a double 0.22 pan filtration may be performed before filling, The filters may be tested for integrity and bubble point befo re and after the filtration. [0121] Pre-washed and autoclaved via's may be aseptically filled using an automatic filling line to a target of 5 ml per vial and then partially stoppered. In process check for fill volumes may be done by checking the fill weight every 15 minutes. [0122] The lyophilizing is generally conducted within about 72 hours, such as within about 8 hours, or within about 4 hours, of the dissolving. [0123] In one or more embodiments, the lyophilizing comprises freezing the solution to form a frozen solution. The frozen solution is typically held at an initial temperature ranging from about - 40"C to about -50'C, such as about -45*C, During the initial temperature period, the pressure around the frozen solution is typically atmospheric pressure. The initial temperature period typically ranges from about 1 hour to about 4 hours, such about 1.5 hours to about 3 hours, or about 2 hours, [0124] The lyophilizing may further comprise raising a temperature of the frozen solution to a first predetermined temperature, which may range from about 10 C to about 201C, such as about 15'C. The time for the heat ramp from the initial temperature to the first predetermined temperature generally ranges from about 6 hours to about 10 hours, such as about 7 hours to about 9 hours. [0125] During the first predetermined temperature period, the pressure around the solution typically ranges from about 100 pbar to about 250 pbar, such as about 150 pbar to 22 about 225 jLbar. The solution may be held at the first predetermined temperature for a period ra nging from about 20 hours to about 30 hours, such as from about 24 hours, [0126] The lyophilizing may still further comprise raising a temerature of the solution to a second predetermined temperature, which may range from about 250C to about 354C, such as about 30*C. During the second predetermined temperature period, the pressure around the frozen solution typically ranges from about 100 phar to about 250 bar, such as about 150 pbar to about 225 pbar. The solution may be held at the second predetermined temperature for a period ranging from about 10 hours to about 20 hours, [0127] In view of the above, the lyophilization cycle may comprise a freezing ramp, e.g,, from 200C to - 451C in 65 minutes, followed by a freeze soak, e.g, at - 450C for 2 hours, Primary drying may be accomplished with a heating ramp, e.g., from - 45*C to 15*C in 8 hours, followed by a temperature hold, e.g., at 150C for 24 hours at a pressure of 200 pbar, Secondary drying may be accomplished with a heating ramp, e.g., from 15'C to 300C in 15 minutes, followed by a temperature hold at 30*C for 15 hours at a pressure of 200 jsbar. At the end of the lyophilization cycle, the vacuum may be broken with sterile nitrogen, and the vials may be automatically stoppered. £01281 The water content of the powder e.g., vancomycin powder, or amikacin powder, is typically less than about 7 wt%, such as iess than about 5 wt %, less than about 4 wt%, less than about 3 wt%, or less than about 2 or 1 wt%. [0129] The chromatographic purity level of the powder, e.g., vancomycin powder, or anikacin powder, typically greater than about 80%, such as greater than about 90%, greater than about 95%, or greater than about 97%. In this regard, there is generally no major impurity greater than about 10%, such as no greater than about 7% or no greater than about 5%. For instance, the amount of heavy metals is typically less than about 0.005 wt%, such as less than about 0.004 wt%, less than about 0.003 vt%, less than about 0.002 wt%, or less than about 0.001 wt%. [01301 The powder is capable of being reconstituted with water at 25*C and 1.0 atmosphere and with manual agitation, in less than about 60 seconds, such as less than about 30 seconds, less than about 15 seconds, or less than about 10 seconds, 23 [01313 The powder typically has a large specific surface rea that facilitates reconstitution. The specific surface area typically ranges from about 5 m2/g to about 20 m 2 such as about 8 m 2 /g to 15 m 2 /g, or about 10 m 2 /g to 12 m2/g [0132 Upon reconstitution with water, the antibiotic solution (such as vancomycin or amikacin) typically has a pH that ranges from about 2.5 to about 7, such as about 3 to about 6. Amikacin in particular may have a pH of about 5.5 to about 6.3. [0133] In addition to use formulations for nebulization, the formulations of the present invention may be administered other routes, e.g., parenteral administration, [0134] One or more embodiments involve methods for treating or preventing pulmonary infections, including nosocomial infections, in animals, including, especially, humans, The method generally comprises administering to an animal subject or human patient in need thereof, as an aerosol, a therapeutically or prophylactically effective amount of the antibiotic or salt thereof, Several antibiotics may be delivered in combination according to the invention, or in seriatim, In one or more embodiments, the amounts delivered to the airways, if delivered systemically in such arnounts, would not be sufficient to be therapeutically effective and would certainly not be enough to induce toxicity. At the same time, in such embodiments, such amounts can result in sputum levels of antibiotic of more than about 10-100 times the minimum inhibitory concentration ("MIC"). [0135] In one particular embodiment, the pharmaceutical formulation comprises an antibiotic for administration to a ventilated patient to treat or prevent ventilator associated pneumonia (VAP) and/or hospital-acquired pneumonia (HAP) and/or community acquired pneumonia (CAP) as well as other forms of pneumonia, and other respiratory infections or conditions, Such administration is described in U.S. Patent Application Nos. 10/430,658; 10/430,765; and 10/991,092, and in U.S. Provisional Application Nos. 60/378,475; 60/380,783; 60/420,429; 60/439,894; 60/442,785; 60/682,099, and in U.S. Patent Application Publication No. 2005/021766, all of which are incorporated herein by reference in their entireties. [01363 In one aspect, the aerosolized particles are prevented from undergoing significant hygroscopic enlargement, since particles enrobed in water will tend to condense on the walls. For instance, the method may involve reducing humidity in the ventilator circuit by a predetermined amount before nebulization begins, In this embodiment, the 24 humidity may facilitate an MMAD of less than about 3 pm or less than about 1.5 pm, In another embodiment, each aerosol particle is delivered enrobed in a substantially anhygroscopic envelope. [0137} Of course, embodiments can be used where diameters are greater, Moreover, in some cases, the present invention contemplates adjustments to the surface electrical charges on the particles or the walIls, For example, assuming surface charge on the device is important, the present invention contemplates embodiments wherein the components of the device connectors are made of metal (or at least coated with metal). Alternatively, the components can be treated with agents (e.g. wetting agents, detergents, soaps) to adjust surface charge. [0138] In one aspect, the method comprises inserting an aerosol delivery end of the device within said patient's trachea to create a positioned device, The antibiotic composition is aerosolized under conditions such that the composition is delivered through said aerosol delivery end of the device to the patient, Yherein the aerosol first contacts the patient's trachea (thereby bypassing the oro-pharynx), The method may involve administering a mixture of antibiotics and is particularly appropriate for intubated patients. [0139] In another aspect, a method of administering commises administering to free breathing patients by way of an aerosol generator device and/or system for administration of aerosolized medicaments such as those disclosed in U.S. Patent Application Publication Nos, 20050235987, 20050211253, 20050211245, 20040035413, and 20040011358, the disclosures of which are incorporated herein by reference in their entirities. [0140] Such devices may deliver medicament phasically or non-phasically. Additionally or alternatively, such devices may incorporate a chamber or reservoir to accumulate and periodically dispense the aerosolized medicament, In one or more embodiments, an aerosolized nedicanent comprises amikacin, [0141] In one or more embodiments, the method of administering an antibiotic formulation involves dissolving an antibiotic or salt thereof in a solvent to fonn an antibiotic formulation. The aerosolizing is conducted within about 16 hours, such as with about 12 hours, or within about 8 hours, of the dissolving. 25 [01421 In another aspect, particular with respect to "constant-flow" ventilators, the present invention contemplates limiting the delivery event to the inspiratory phase of the ventilator cycle and, if possible, at a reduced flow-rate, Thus, in one embodiment, aerosolization is actuated during (or in fixed realtion to) the inspiration phase of the breathing cycle. [0143] It is not intended that the present invention be limited to particular dosages. On the other hand, the efficiency of the aerosol systems and methods described herein permit amounts to be delivered that are too low to be generally effective if administered systemically, but are nonetheless effective amounts when administered in a suitable and pharmaceutically acceptable formulation directly to the airway. Importantly, while efficiencies can be increased, in some embodiments efficiencies are not increased at the expense of control over the dose. Thus, lower efficiencies are contemplated as preferred when delivery is more reproducible. [0144) It is not intended that the present invention be limited to antimicrobials that only kill particular organisms. The present invention contemplates drugs and drug combinations that will address a wide variety of organisms. In one or more embodiments, the present invention contemplates drugs or drug combinations effective in the treatment of infections caused by P. aeruginosa, S. aureus, H inrfuenza, and & pneunoniae and/or antibiotic-resistant strains of bacteria such as methicillin-resistant S. aureus, and Acetinobacter species, among others. [0145] Moreover, while certain embodiments of the present invention are presented in the context of the intubated patient, other patients at risk for infection are contemplated as treatable with the compositions, methods, and devices of the present invention. For example, the elderly (particularly those in nursing homes), horses, dogs and cats in competitions (show and racing animals), animals that frequently travel (e.g., circus animals), animals in close quarters (e.g., zoos or farms), humans and animals in general are at risk for lung infections, The present invention contemplates delivery of aerosols to the trachea and/or deep lung for such individuals-both prophylactically (i.e., before symptoms) and under acute conditions (i.e., after symptoms)--wherein said aerosols comprise antimicrobials, and in particular, the antibiotic mixtures described above. 26 [01461 In one embodiment, the present invention contemplates administering the appropriate medication to a patient diagnosed with ARDS or chronic obstructive pulmonary disease (COPD), [0147] One or more embodiments are directed to unit doses comprising a container and the compositions. [0148] Examples of the container include, but are not limited to, vials, syringes, ampoules, and blow fill seal, For instance, the vial may be a colorless Type I borosilicate glass ISO 6R 10 mL vial with a chlorobutyl rubber siliconized stopper, and rip-off type aluminum cap with colored plastic cover. [0149] The amount of the composition in the unit dose typically ranges from about 2 ml to about 15 ml, such as from about 3 mi to about 10 ml, about 4 m1 to about 8 ml, or about 5 ml to about 6 ml. [0150] The amount of the antibiotic in the unit dose, adjusted for potency, typically ranges from about 150 mg to about 900 mg, such as about 400 mg to about 750 mg. For instance, an amount of the anti-gram-negative antibiotic or salt thereof may range from about 400 mg to about 750 mg. As another example, the amount of anti-gram-positive antibiotic or salt thereof may range from about 150 mg to about 450 mg, or from about 550 mug to about 900 Mg. [01511 One or more embodiments are directed to kits. For instance, the kit may includes a first container containing a first aqueous solution comprising anti-gram-negative antibiotic or salt thereof and a second container containing a second aqueous solution comprising nUti-gram-negative antibiotic or salt thereof A concentration, or an amount, or both, of the first aqueous solution is different from a concentration, or an amount, or both, of the second aqueous solution. For instance, the amount of the first aqueous solution may range from about 2 ml to about 5 ml, and the amount of the second aqueous solution may range from about 5 ml to about 8 ml. [0152] In one or more enbodiments, the kit includes a first container containing a first aqueous solution comprising anti-gram-negative antibiotc or salt thereof A second container contains a second aqueous solution comprising anti-gram-positive antibiotic or salt thereof The concentrations and/or amounts of the anti-gram-negative antibiotic or salt and the anti-gram-positive antibiotic or salt may be the same or different. 27 [0153] In one or more embodiments, a kit includes a first container containing a first composition comprising an antibiotic or salt thereof A second container contains a second composition comprising water. The first composition and/or the second composition comprises ai osmolality adjuster, [0154] In one or more embodiments, a kit includes a first container containing a powder comprising anti-gram-positive antibiotic or salt thereof A second container contains a powder comprising anti-gram-positive antibiotic or salt thereof A concentration, or an amount, or both of the anti-gram-positive antibiotic or salt thereof in the first container is different from a concentration, or an amount, or both of the anti-gram-positive antibiotic or salt thereof in the second container. [0155] For instance, the amount of the anti-gram-positive antibiotic or salt thereof in the first container may range from about 400 mg to 600 mg. The amount of the anti-gram positive antibiotic or salt thereof in the second container may range from about 600 mg to about 800 mg. [01561 In another aspect, a kit may include a first container containing a solution comprising anti-gram-negative antibiotic or salt thereofE A second container may contain a powder comprising anti-gram-positive antibiotic or salt thereof Alternatively, the anti-gram negative antibiotic or salt thereof may be a powder, and the anti-grain-positive antibiotic or salt thereof may be a solution or dispersion. An amount of the anti-gram-positive antibiotic or salt thereof generally ranges from about 150 mg to about 900 mg. [0157] The kits may farther comprise a package, such as a bag, that contains the first container and the second container. [0158] The kits may further comprise an aerosolization apparatus. The aerosolization apparatus may be of any type that is capable of producing respirable particles or droplets. Alternatively, the antibiotic may be dissolved in or suspended in a liquid propellant, as described in U.S. Patent Nos. 5,225,183; 5,681,545; 5,683,677; 5,474,759; 5,508,023; 6,309,623; or 5,655,520, all of which are incorporated herein by reference in their entireties, in such cases, the aerosolization apparatus may comprise a metered dose inhaler (MDI). [0159] Alternatively or additionally, the pharmaceutical formulation may be in a liquid form, and may be aerosolized using a nebulizer as described in WO 2004/071368, which is herein incorporated by reference in its entirety, as well as US. Published 28 Application Nos, 2004/0011358 and 2004/0035413, which are both herein incorporated by reference in their entireties. Other examples of nebulizers include, but are not limited to, the Aeroneb@Go or Aeroneb@Pro nebulizers, available from Aerogen, Inc. of Mountain View, CA; the PARI eFlow and other PARI nebulizers available from PARI Respiratory Equipment, Inc. of Midlothian, VA; the Lumiscope@ Nebulizer 6600 or 6610 available from Lumiscope Company, Inc. of East Brunswick, NJ; and the Omron NE-U22 available from Omron Healthcare, Inc. of Kyoto, Japan, [0160] It has been found that a nebulizer of the vibrating mesh type, such as one that that forms droplets without the use of compressed gas, such as the Aeroneb@ Pro provides unexpected improvement in dosing efficiency and consistency. By generating fine droplets by using a vibrating perforated or unperforated membrane, rather than by introducing compressed air, the aerosolized pharmaceutical formulation can be introduced into the ventilator circuit without substantially affecting the flow characteristics within the circuit and without requiring a substantial re-selection of the ventilator settings. In addition, the generated droplets when using a nebulizer of this type are introduced at a low velocity, thereby decreasing the likelihood of the droplets being driven to an undesired region of the ventilator circuit. Furthermore, the combination of a droplet forming nebulizer and an aerosol introducer as described is beneficial in that there is a reduction in the variability of dosing when the ventilator uses different tidal volumes, thus making the system more universal. [0161] Using an adaptor, device or system as disclosed in U.S, Application No. 10/991,092 and/or U.S. Provisional Application No, 60/682,099, and/or U.S. Application Publication No. 2005/0217666, all of which are incorporated herein by reference in their entireties, in connection with the administration of aerosolized antibiotics offers substantial benefits. For example, when using such adaptors, substantially less pharmaceutical formulation is lost to the environment vhich results in a reduction in bacterial resistance against the antibiotic. In addition, the adaptors, devices or syaters are able to deliver a more consistent dose which is particularly useful for antibiotic therapy. [0162] FIG, IA shows an embodiment of an adapter or system for aerosol delivery of medicaments, comprising a pulmonary drug delivery system ("PDDS") 100 siutable for use with the present invention. 'The PDDS 100 may include a nebulizer 102 (also called an 29 aerosolizer), which aerosolizes a liquid medicament stored in reservoir 104. The aerosol exiting nebulizer 102 may first enter the 7-adaptor 106 that couples the nebulizer 102 to the ventilator circuit. The T-adaptor 106 is also coupled to the circuit wye 108 that has branching ventilator limbs 110 and 112. 101631 Coupled to one of the ventilator limbs 110 or 112 may be an air pressure feedback unit 114, which equalizes the pressure in the limb with- the air pressure feedback tubing 116 connected to the control module 118. In the embodiment shown, feedback unit 114 has a female connection end (e.g., an ISo 22 mm female fitting) operable to receive ventilator limb 112, and a male connection end (e.g., an ISO 22 mm male fitting) facing opposite, and operable to be inserted into the ventilator. The feedback unit may also be operable to receive a filter 115 that can trap particulates and bacteria attempting to travel between the ventilator circuit and tubing 116. [01641 The control module 118 may monitor the pres sure in the ventilator limb via tubing 116, and use the information to control the nebulizer 102 through system cable 120. In other embodiments (not shown) the control module 118 may control aerosol generation by transmitting wireless signals to a wireless control module on the nebulizer 102, [0165] During the inhalation phase of the patient's breathing cycle, aerosolized medicament entering T-adaptor 106 may be mixed with the respiratory gases from the inspiratory ventilator limb 112 flowing to the patients nose and/or lungs. In the embodiment shown, the aerosol and respiratory gases flow through nose piece 122 and into the nasai passages of the patients respiratory tract. [0166] Other embodiments of the circuit wye 108 shown in FIG. IA are also contemplated in embodiments of the invention, [0167] Referring to FIG iB, a nebulizer 85, which may have a top portion 93 through which liquid may be provided may be incorporated into a ventilator breathing circuit of a ventilated patient. The breathing circuit may comprise a "Y" connector 88, which may in turn have an inlet portion 89, an endotracheal tube portion 90 and an outlet portion 91. The inlet portion 89 carries air provided from the ventilator 92 toward the patient. The endotracheal tube portion 90 of the Y connector 88 carries the incoming air to the patient's respiratory tract; this direction is represented by arow . The endotracheal tube portion 90 also carries the patient's exhalation to the outlet portion 91 of the Y connector 88, and the outlet portion 30 may lead to an exhaust, represented by arrow "b", to remove the patient's exhalation from the system. The nebulizer 85 of the present invention aerosolization element generates an aerosol cloud 94 that remains substantially within the inlet portion 89 of the Y connector 88 when there is no inspiratory air flowing through the inlet portion, by virtue of the aerosolization element, as described above, producing a low velocity mist. In this manner, aerosol that is generated when there is no inhalation air being provided will not be carried out through the outlet portion 91 of the Y connector and lost to the ambient environment. Accordingly, a dose of aerosolized medication may be preloaded, i.e., produced and placed substantially within the inlet portion 89 prior to an inhalation phase being sent by the ventilator 92. In this manner, such medication can be swept into a patient's respiratory system at the very start of the inhalation cycle. This may be of particular benefit in the case of neonatal patients and in other instances in which only the initial blast of inhalation phase will reach the target portion of the respiratory system. In alternate embodiments, the ventilator may generate a continuous bias flow of gas through the ventilator circuit. The bias flow may push some of the aerosolized medicament through the outlet portion 91, but there is still an overall benefit from having the aerosolized medicament preloaded through the ventilator circuit. [01 68] Referring now to FIG. 2A, an embodiment of an off-ventilator configuration of an adapter and/or system for pulmonary delivery is shown. In FIG, 2A, the adapter 400 is intended for off-ventilator use, and includes an endpiece 402 that is coupled to a nebulizer 404 and wye 406, The nebulizer 404 may include reservoir 408, which supplies the liquid medicament that is aerosolized into connector 410, The connector 410 can provide a conduit for the aerosolized medicament and gases to travel from the wye 406 to endpiece 402, and then into the patient's mouth and/or nose. The first wye limb 412 may be connected to a pump or source of pressurized respiratory gases (not shown), which flow through the wye limb 412 to the endpiece 402. A one-way valve 413 may also be placed in the limb 412 to prevent respired gases from flowing back into the pump or gas source. The limb 412 may also include a pressure feedback port 414 that may be connected to a gas pressure feedback unit (not shown). In the embodiment shown, a feedback filter 416 may be coupled between the port 414 and feedback unit. [0169] The off-ventilator adapter 400 may also include a second wye limb 420, which includes a filter 422 and one-way valve 424, through which gases may pass during an 31 exhalation cycle. The filter 422 may filter out aerosolized medicament and infectious agents exhaled by the patient to prevent these materials from escaping into the surrounding atmosphere, The one-way valve 424 can prevent ambient air from flowing back into the adapter 400. [0170) A general form of an aerosolized composition delivery system 1100 is shown in Fig, 2B. The aerosolized composition delivery system 1100 delivers an aerosolized composition to a portion of a user's respiratory tract, such as the user's lungs. The aerosolized composition delivery system 1100 is useful in delivering the aerosolized composition to a patient whose breathing is being assisted by a ventilator 1105 but may also be configured to be used to deliver a composition to a non-ventilated patient. The ventilator circuit 110 is shown diagrammatcally in Fig. 23, Extending from the ventilator 1105 is an inhalation line 1115 and an exhalation line 1120. The inhalation line 1115 and the exhalation line 1120 are both composed of tubing having an airflow lumen extending therethrough. The inhalation line 1115 and the exhalation line 1120 meet at an adaptor 1145 remote from the ventilator 1105. At the adapter 1145 the lumen of the inhalation line 1115 is in communication with the lumen from the exhalation line 1120, and both lumens are in communication with a patient line 1130. The patient line 1130 comprises a lumen that extends to the lumen of an endotracheal or tracheostomy tube 1135, which is inserted into a patient. The tube 1135 has an opposite end that may extend into or near the lungs of the user. Accordingly, in use, oxygenated air is introduced into the inhalation line 1115 by the ventilator 1105. Thme oxygenated air passes through the lumen of the inhalation line 1115, into the patient line 1130, through the lumen of the tube 1135, and into the lungs of the patient. The patient fhen exhales, either naturally or by applying negative pressure from the ventilator, and the exhaled air passes through the tube 1135, through the patient line 1130, and through the exhalation line 1120 to the ventilator 1105. The cycle is continuously repeated to assist the patient's breathing or to entirely control the breathing of the patient. [017 1] The adapter 1145 introduces aerosolized composition into the ventilator circuit 1110. The aerosol that is introduced by the adapter 1145 is generated by an acrosolization apparatus 1150, which comprises a reservoir for containing a composition. Thus, in one or more embodiments, aerosolization energy is supplied to the aerosolization device by an energy source 1160 to generate the aerosolized composition. The aerosolized pharmaceutical 32 formulation passes through a passage 1165 to the adapter 1145 where it may be introduced into the ventilator circuit 1110. The aerosolization apparatus 1150 may be, for example, a jet nebulizer where the energy source is compressed air, a vibrating mesh nebulizer where the energy source is wave energy, an ultrasonic nebulizer, or a metered dose inhaler where the energy source is a propellant that boils under ambient conditions, [0172] Examples of the adaptor 1145 for introducing the aerosol-zed pharmaceutical formulation are disclosed in U.S. Application No. 10/991,092, filed November 17, 2004 and U.S. Provisional Application No. 60/682,099, which applications are herein incorporated by reference in their entirety. [01731 The introduction of the aerosolized pharmaceutical formulation at the adapter 1145 is advantageous in many respects over systems where the aerosol is introduced into the inhalation line 1115 or within the ventilator 1105. For example, by introducing the aerosolized pharmaceutical formulation at the adapter 1145, the ventilator circuit volume from the point of introduction to the patient's lungs is substantially reduced. Accordingly, the aerosolized pharmaceutical formulation is more concentrated and is less diffused throughout the ventilator circuit 1110. In addition, if the fonnulation is added in the inhalation line 1115, much of the formulation is drawn into the exhalation line 1120, further limiting the efficiency of the administration. Because of this diffusion and reduced efficiency, the consistency of dosing is difficult to control in known systems. Also, the presence of high quantities of the aerosolized pharmaceutical formulation that are not administered to the lungs of the patient may be undesirable in that much of the aerosol may be introduced into the environment where it may be inhaled by healthcare workers or others, [0174] Therefore, the adaptor 1145 of the invention has been designed to introduce the aerosolized pharmaceutical formulation in an improved mamner to increase the efficiency and/or the consistency of the dosing. The adaptor 1145 serves to reduce the amount of aerosolized pharmaceutical fornmlation that is drawn into the exhalation line 1120 of the ventilator-circuit 1120. [0175] The adaptors of the present invention when used in a ventilator circuit are often able to reproducibly and efficiently deliver pharmaceutical formulation. For instance, the pre ent invention is typically able to reproduce the delivered dose within about - 10%, :h 8%, 6z 6%, - 4%, =' 2%, or i 1%, of the total nominal dose. The present invention is often 33 able to achieve a delivered efficiency of at least about 30%, such as at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 80%, or at least about 90%. [0176] The adaptor of the present invention typically has minimal impact on the patient to ventilator interface, The minimal impact allows the ventilator to react more efficiently to the patient. The adaptor and valves are arranged so that at an air flow rate of 60 L/min, the pressure drop between the first end and the second end of the adaptor is often less than about 50 cm H20, such as less than about 30 cm H20, less than about 5 cm 1

-

1 0 , less than about 4 cm 1-120, less than about 3 cm H2O, less than about 2 1-120, less than about i cm H20, less than about M.5 cm 120, or less than about 0.1 cm H20, and may range from about 0,05 cm 1-120 to about 10 cm H20, about I cm H20 to about 5 cm H20, or about 2 cm H20 to about 4 cm B20. At an air flow rate of 30 /min, the pressure drop between the first end and the second end of the adaptor is typically ranges from about 1 cm H2O to about 2 can 120. [0177] The adaptor may be made of a transparent, translucent, or opaque material. Using a transparent material is advantageous because the user can visu ally inspect the functioning of the adaptor. Examples of materials for the adaptor include, but are not limited, to polymers, such as polypropylene, SAN (styrene acrylonitrile copolymer), ABS (acrylonitriie-butadiene-styrene, polycarbon ate, acrylic polysulfone, K-resin® styrene butadiene-copolymer (available from Chevron Phillips Chemical), polyethylene, PVC (polyvinyl chloride), polystyrene, and the like. [0178] For vibrating mesh nebulizers, such as the Aeroneb Pro and the PARI eFlow, reproducible administrations can result from smaller first channel volumes. It has been determined, for example, that the first channel volume for an adaptor 1145 used with a vibrating mesh nebulizer may be any volume greater than about 10 ml, such as from about 10 mil to about 1000 ml, about 50 ml to about 200 ml, or about 90 ml. Both the stored volume and valving affect the performance of the present invention. [0179] Additional examples of devices and methods are disclosed in US. Patent Application No. 11/436,329, "Valves, Devices, and Methods for Endobronchial Trapy," filed May 18, 2006, which is incorporated herein by reference in its entirety. [0180] The present invention is not limited to any precise desired outcome when using the above-described compositions, devices, and methods. However, it is believed that 34 the compositions, devices, and methods of the present invention may result in a reduction in mortality rates of intubated patients, a decrease in the incidence of resistance (or at least no increase in resistance) because of the reduced systemic antibiotic exposure and elevated exposure at the targeted mucosal surface of the lung caused by local administration. As noted above, it is contemplated that the compositions, devices, and methods of the present invention are useful in the treatment of pneumonia (and may be more effective than systemic treatment -or at the very least, a useful adjunct). It is believed that related infections may also be prevented or reduced (e.g., prevention of sepsis, suppression of urinary tract infections, etc.) [0181] Of course, a reduced use of systemic antibiotics because of the efficacy of the compositions, devices, and methods of the present invention may result in reduced cost, reduced time on IV lines, and/or reduced time on central lines). Moreover, such a reduction should reduce antibiotic toxicity (as measured by reduced incidence of diarrhea and C dgficile infection, better nutrition, etc.) ['0182] It is believed that the compositions, devices, and methods of the present invention will locally result in a reduction of the ET/Trach tube biofilm. This should, in turn, get rid of secretions, decrease airway resistance, and/or decrease the work of breathing. The latter should ease the process of weaning the patient off of the ventilator. [0183] The present invention contemplates specific embodinents that can replace commonly used elements of a ventilator system. In one or more embodiments, the present invention contemplates an adapter attachable to a ventilator circuit and to an endotracheal tube, wherein the adaptor comprises an aerosol generator. While not limited to any precise desired outcome, it is contemplated that the adapter with integral generator will reduce the effects of the ventilator on all conventional aerosol systems (et, ultrasonic and MDI), and at the same time enhance the positive qualities of a device like the AerogenTM pro. Again, while not limited to any precise desired outcome, it is contemplated that the adapter with integral generator wil (I) reduce variability in delivery (reduced effects of humidification, bias flow, continuous vs breath-actuated) so as to achieve the same delivery (no matter what commercial ventilator system is used); (2) allow for maximal effects of breath actuation; and (3) allow for maximal effect to enhanced nebulizer efficiency using nebulizers having no dead volume. 35 [0184 The present invention is not limited to the precise configuration or nature of the circuit. In one embodiment, said circuit is a closed circuit. In another embodiment, said circuit is an open circuit. [0185] Again, the present invention is not limited to particular vent configurations, In one embodiment, said inspiratory and said expiratory lines are connected to a mechanical ventilator. In one embodiment, said mechanical ventilator controls a breathing cycle, said cycle comprising an inspiration phase. In one embodiment, the aerosol is administered during the inspiration phase of the breathing cycle. [0186] Although the present invention has been described in considerable detail with regard to certain versions thereof, other versions are possible, and alterations, permutations and equivalents of the version shown will become apparent to those skilled in the art upon a reading of the specification and study of the drawings. For example, the relative positions of the elements in the aerosolization device may be changed, and flexible parts may be replaced by more rigid parts that are hinged, or otherwise movable, to mimic the action of the flexible part. in addition, the passageways need not necessarily be substantially linear, as shown in the drawings, but may be curved or angled, for example. Also, the various features of the versions herein can be combined in various ways to provide additional versions of the present invention. Furthermore, certain terminology has been used for the purposes of descriptive clarity, and not to limit the present invention, Therefore, any appended claims should not be limited to the description of the preferred versions contained herein and should include all such alterations, permutations, and equivalents as fall within the true spirit and scope of the present invention. [0187] The foregoing description will be more fully understood with reference to the following Examples. Such Examples, are, however, merely representative of methods of practicing one or more embodiments of the present invention and should not be read as limiting the scope of the invention. Example 1 [0188) This Example involves determining the solubility of gentarnicin sulfate in water and saline. The required strengths were initially set at 20 mg/ml, 40 mg/mi, and up to 200 mg/mi. 36 WATER SOLUBILITY DETERMINATIONS [0189] Solubility in water was determined via visual assessment. Osmolality and pH were also determined. [0190] The batch size of all the solutions manufactured for the solubility determination studies was 10 ml,. The method of manufacture consisted of weighing the appropriate amount of gentamicin sulfate and then taking to final volume with water. It was noted that especially for higher concentrations, the solution was first shaken by hand and then placed on a magnetic stirrer to ensure complete dissolution. {0191] Table I lists the pH and osmolality values obtained for solutions of gentamicin sulfate in water for injection (WfI) with concentrations ranging from 20 mg/ni to 400 mg/ml, 37 Table 1. Test Matrix for Gentamicin Solution in WI Active Weight of Gentamicin Sulfate pH Osmolahty Concentration Dispensed' (mg/mi) (mOsmol/kg) m

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20 34 4.81 61 40 68 4.72 101 80 136 4.92 197 20 204 4.93 275 200 340 5.01 524 250 425 5,06 1178 300 510 5.13 2013 350 55, 2 NR 400 680 5.26 NR Activity of gentamicin sulfate = 58,8%, so conversion factor = 1.701 NR: No result, sample did not freeze [0192] As seen in Table 1, all the solutions had a pH that was higher than 4, which is considered to be acceptable for drug delivery to the lungs, However, with regard to osmolality readings, doses greater than 200 mg/ml exceeded the targeted range. GENTAMICIN SULFATE IN 0.9% SALINE SOLUTION [0193] The solubility, pH, and osmoLlity of gentamicin sulfate solutions prepared with 0.9% saline solution were determined, The solubility was determined by visual assessment. Only three concentrations of gentamnicin were investigated (20, 40, and 80 mg/ml). [0194] Table 2 lists the parameters measured for gentamicin solutions and the observations recorded during manufacture. Table 2, Osmolality and p1i of Gentamicin Sulfate in 0.9% Saline 38 Active Weight of pH Osmnolality Concentration Gentamicin Sulfate (mOsmol/kg) (mng/mni) Dispensed (mg/mi) _________ ___ 20 34 4.94 318 40 68 4.72 353 80 136 4.82 445 Example 2 This Example involves developing the freeze-drying cycle for the clinical manufacture of the Vanconycin HC lyophilisate. A 120 mg, 240 mg, and 480 mg of Vancomycin HC / vial strength were investigated. MATERIALS/EQUIPMENT Materials a Vancomycin hydrochloride, USP, Alpharma - Denmark o ISO 6R clear type I glass vials, Nuova Ompi - Italy * 20 rn freeze-drying stoppers, West Pharmaceutical Service-USA a 20 mm flip-off caps, Capsulit S.p.A. - Italy * 13 mm freeze-drying stoppers, West Pharmaceutical Service-USA * 13 rm flip-off caps, West Pharmaceutical Service-USA Equipment * Glassware for Vancomycin solution before and after filtration (bottles). * Pressure vessel, Sartorius - Germany * Balance to check the filling weight (10 mug sensitivity), Sartorius - Germany * Digital pH meter, Mettler Toledo - Switzerland * Karl Fischer automatic titrator DL38, Mettler'Toledo -- Switzerland. o 0,22 um sterilizing PVDF filter, Pall * Manual doser, Hirschmann 39 * Isolator, E.Co.Tec - Italy * Lyophilizer, BOC Edwards Lyoflex 04 (or Minifast 8000) with the following characteristics: 0.4 m2 (or 0,8 m.2) shelf surface; temperature range -50*C to 50*C; PT 100 temperature probes; Pirani gauge for vacuum monitoring; coil condenser with ice capacity of 8 kg; condenser coil inlet temperature to -60vC, stainless steel trays with a thickness of about 2 nm; semiautomatic crimLpiug machine (Flexseal - Denmark) * DSC Pyris Diamond - USA COMPOSITION Solubility Study [01951 The solubility of the Vancomycin HCl has been evaluated in order to establish a suitable formulation to obtain a final Jyophilised product which matches all the criteria required by its use as phannaceutical form. [0196] The solubility coupled with a pH evaluation of Vancomycin HCl solutions at different concentration was the first step to focus the suitable final fornulation for a better development of the lyophilization cycle. [0197] A saturated solution of Vancomycin HCl in water for injection was prepared by adding under stirring the active agent to the solvent. [0198] At first, the solution was clear with the solid suspended as an agglomerate; after the solid worked as crystallization nucleus and a new precipitation occurred; so the solutions became white and more viscous because the solid partially swells. [0199] Suspension was stirred for 48 h in order to reach the equilibrium conditions for the dissolution. [0200] Suspensions was filtered first through a paper filter and then through a 0,45 gim PVDF filter discarding the first drops of solution which could have been diluted because of the binding of the product to the membrane, [0201] The resulting solution obtained after the two filtrations was stored at 2-84C in order to evaluate if precipitation of the solid occurs. [0202] The filtrated solutions of Vancomycin HCI in water coming from the respective saturated solutions, was diluted to reach a final concentration which gave an Abs value at )=280 nm included into the calibration curve. 40 [L0203] Each diluted solution was analysed in triplicate with UV at ,= 280 nm. For each solution the absorbances have been mediated and the final value has been substituted in the respective calibration curve equation to calculate the concentration. [0204] The maximum solubility of Vancomycin HC in water is 140,9 mg/mL, pH of Vancomycin HCI Solution in Water [0205] Besides the solubility evaluation it was also measured the pH and density of Vancomycin HC solutions at different concentrations which could have been taken into account for the development of the formulation and of the lyophilization cycle. Solution Concentration PH Density (mg/mL) (g/mL) 140. 34-3.5 046 130.5 5 3.532 - 1.042 12026 3.6-38 1.037 110.16 3.7 - 3,9 1.034 -t ---------- 100.12 38- 4 .1 1.027 [0206] The pH varied within a restricted range for each concentration and the overall pH within 140 mg/mL and 100 mg/mL was stable around the acid value. FORMULATION 0207] Vancomycin hydrochloride was dissolved in water for injection to form 100 mg/mil formulations in 1.00 ml and 1 .20 ml amounts, as shown below, -- Quantity In redients Amount / m Amount / Unit Vancomycin HC10 100.00 120.00 n,, Water for direction to 100 nl to 1.20 ml [0208] Vancomycin hydrochloride was also dissolved in water for injection to form 120 mg/ml fornulations in 1.00 ml, 2.00 nil, and 4.00 ml amounts, as shown below, Quantity 41 Ingredients Amount /Unit . .... I Amount I ml i20 mg/vial 240 mg/vial . 480 mg/vial vanCornyoin: 1 1.20.0 120,00 mg 240.00 mg 480.00 mg Water for to 100 ml to 1.00 ml to 2.00 ml to 4.00 ml DSC STUDIES {0209] DSC was performed on the ready to fill solution with a concentration of 100 mg/ml and 120 mg/miL [0210] The DSC runs were performed by cooling the samples to -50'C at a cooling rate of I 4C/min, and by heating them back to 20*C at different scan rates after a period of few minutes of isothermal step. [0211] Samples amount ranged approximately from 1 to 3 mg, [0212] All the peaks corresponding to the detected thernal events were calculated as onset temperature. [0213] The DSC studies showed that there was a main event of crystallization during freezing and that there is no evidence of smaller crystallization events. These phenomena seem to indicate an absence of amorphous phase during freezing and a complete retention of crystalline structure by vancomycin, as confirmed by the lack of glass transitions events during the heating steps in all cases. [0214] As expected, crystallization peak was displaced to lower temperatures when increasing the weight of the sample or the concentration of the solution. [0215] However no significant difference was detected among the different concentrations. [0216] Detected differences are more linked to the internal variability of samples, [0217] A freezing end temperature of -45*C as well as a freezing rate of I4C / min was chosen to ensure a full crystalline state offthe Vancomycin H1CI during freezing. [0218] Since the maximum allowable product temperature during initial primary drying was -25 0C, the pressure during primary drying was within % to % of the vapor 42 pressure of ice at -25 C, Vapor of ice at -25 0C is 630 tbar, The average of the thresholds, 230 bar, was selected as the maximum allowed chamber pressure for primary drying. MANUFACTURING PROCESS [0219] Water for injection was weighed out in a glass container on calibrated balances, [0220} Vancomycin HC1 was added under stirring; the solution was agitated until vancomycin was completely dissolved and the dissolution time 'as recorded. [0221] Then, water for injection was added until the required final amount was reached. [0222] On the final solution, pH and density were measured and appearance was evaluated. [0223] The solution was filtered through a 0.22 um PVDF membrane. [0224] The vials were washed with distilled water and dried in an oven at 120*C for 2h, [0225] The filling was performed by mass and the in process controls were carried out by weighing the filled vials every 20 vials. [0226] After lyophilization the following analyses were performed on the final product: water content by Karl Fischer titration; appearance of the cake, reconstitution time, appearance/clarity, pH after reconstitution. [0227] RP-HPLC was run to confirm processing did not influence purity of vancomycin, [0228] Twenty (20) ml of reconstituted drug product were passed through the sterility testing membrane to confirm fonnulation compatibility. Example 2A 43 [0229] After evaluation of the DSC results, the following lyophilization cycle nominal parameters were planned for use on the 100 mg/ml solution: Step N 0 Description Temperature (*C) Pressure rime (fih:m I Load 20 Atmospheric NA 2 Product freezing 20-+-45 Atmospheric 01:05 3 Freeze soak time -45 Atmospheric 03:00 4 Evacuation 45 100 gbar 00:01 5 Primary drying -45-> 10 100 pbar 08:00 6 - arydri-g10100 ubar 14:00 6 Prima dry rymg 10 - ---------- -_=66 . Secondary drying 10-+440 100 pbar 00:30 8 Secondary drying 40 100 pbar 09:00 - - - ------ - 9 Pre-aeration 0.95 bar NA

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10 Stoppering 0.95 bar NA LI Aeration Atmospheric NA _____________ I ________________________-

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______________ _______ Total length 35:36 J-__----------r~ length ------- [023 0] The freezing soak and primary drying times were shortened with respect to the set lyophilization program. [0231] Actually, the product reached -45*C after 80 minutes of the freezing soak step. It was been kept at -45*C one hour more and then the vacuum was pulled in the chamber to start primary drying. [0232] During step 6 (primary drying), all the product temperature probes reached the temperature of the shelves (10 C) after 450 minutes, [02331 The product was left at 10C for 1 hour; afterwards several pressure raise tests were performed to evaluate the sublimation rate, The positive results of these tests allowed to start heating to 404C for secondary drying. Step 6 lasted 510 minutes instead of 840 minutes. [02341 Total length of the cycle was 29 hours, [0235] The cake had a cohesive structure that prevented loss of friable material from the container during sublimation; lyophilised product was not really elegant because of some cracks in the cake (see the picture 1), 44 Example 2B [02363 In this Example involving 100 mg/mil solution. 6R vials were used, In this regard, twenty (20) mn neck vials enable a faster sublimation than the 13 in neck vials, [0237] An intermediate step at 0*C during the primary drying was inserted to have slower water vapor flow during sublimation. In this way less cracks in the lyophilization cake were observed. [0238] The secondary drying temperature was reduced from 40*C to 30*C according a client's request. [0239] Final primary drying temperature was increased from I (0C to 15*C to try to maintain the total length of the cycle to about 29 hours, [02403 The nominal lyophilization parameters for this Example were: Load 20 Atmospheric NA Product freezing 20-45 Atmospheric 01:05 3 Freeze soak time -45 Atmospheric 03:00 4 Evacuation -45 100 [pbar 00:01 5 Primary drying -45-+ 0 100 pbar 04:00 6 Pnimary drying 100 pbar 02 00 7 Primary drying 0-+ 15 100 pbar 02:00 8 Primary drying 15 100 itbar 10:00 9 Secondary drying 15- 30 100 pbar 00:15 10 Secondary drying 30 100 gbar 09:00 11 Pre-aeration 0,95 bar NA 12 Stoppering 095 bar NA 13 Aeration Atmospheric NA 45 [0241] The lower secondary drying temperature did not allow the product to maintain a relatively low residual moisture, The overall average value was 3.61 wt%, while the average moisture content of previous batch was 1.71 wt%. Example 2C {0242] In this Example involving 100 mg/ml solution, the pressure in the chamber was increased from 100 pbar to 200 pbar; a higher pressure will favor the thermal exchanges at the gas/product interface and the thermal conductivity from the shelf to the tray. The bigger amount of heat transported to the product should result in a rise of product temperature and consequently in a faster ice sublination. [0243] Furthermore, after the evaluation of the lyophilization printout, 4 hours were cut from the primary drying and added the secondary drying step. [0244] This Example involved the following nominal parameters: Step N, Description Temperature ('C) Pressure Time (hh:mnm) 1 Load 20 l Atmospheric NA Product freezing 20+45 Atmospheric 01:05 Freeze soak time -45 Atmospheric 03:00 4 Evacuation -45 200 bar 00:01 - - --- - 5 Primary drying -45-+ 0 200 pbar 04 00 6 Primary drying 0 200 pbar 02:00 Primary dryng 0 15 200 gbar 02:00 Primary drying 15 200 bar 06:00 9 Secondary drying 15 - 30 200 sLbar 00:15 10 Secondary drying 30 200 bar 13:00 11 Pre-aeration 0.95 bar NA 12 Stoppering 0.95 bar NA 13 Aeration in.0Atmospheric N A Total length 31:21 46 [0245] Secondary drying was shortened from the programmed 780 minutes to 450 minutes. Actually, the product temperature matched the shelf temperature very soon due to the better heat exchange by drying at 200 Vbar. [0246] The total length of the cycle was 24.5 hours. [0247] Average moisture content was I82 wt%. [0248] The lyophilization product still showed cracks in the cake. Example 2D [0249] This Example involves a filling solution of 120mg/mi to allow doses of 120 mg, 240 mg, and 480 mg per vial. [0250] All three fill volumes were lyophilized using the cycle for the larger fill sample without paying attention to a possible over drying of the lower fill volume samples. [0251] The vancomycin 120 mg/mL filling solution was investigated by perfonning a scansion with the differential calorimeter, and it has been verified that the main thermal events were very close to the ones detected on the 100 mg/n filling solution. [0252] This meant that the same lyophilization cycle conditions were used for the 100 mg/mL could be applied to the 120 mg/mL. [0253] New holding time studies were also perfonned on the 120 rmg/mL concentration. The new cycle was tested on the 480 mg/vial presentation that had the higher fill volume: 4 mL/vial. (0254] The following nominal parameters were tested: Step N' Descriptiori Temperature (*C) Presure Time (hhnm) 1 Load 20 Atmospheric NA 2 Product freezing 20-+-45 AtmospheriG 01:05 3 Freeze soak time -45 Atnospheric 02:00 4 Evacuation -45 20 bar 00:01 -5 P d 45-0-200-bar -0: 0-------0 5 Primary drying 450 200 libar 04:00 7 Primary drying 0 15 200 pba 02:00 47 8 Primary drying 15 200 pbar 24:00 9 Secondary drying 15 -30 200 jibar 00:15 10 Secondary drying 30 200 pa 1 15:00 - i -re-e rt------- - - - - - - - - ------ a -A -i- Pre-aeration 0,95 bar NA 12 Stoppering 0.95 bar NA 13 Aeration Atmospheric NA Total length 50:21 [0255] An overall average moisture content value of 0.97 wt% was found by Karl Fisher titration. Example 2E [0256] Following the evaluation of the product temperature profile versus the shelves temperature, the following run was cut three to four hours in the primary drying step and four hours in the secondary drying. [0257] The 120 mg and the 240 rmg units were placed in the Iyophilizer during the 480 mg cycle to check if over drying will affect the chemical stability of the 120 mg and 240 mg vials. [025 81 The average residual moisture was 0.97 wt% for the 4 ml fill, 1.23 wt% for the 2 ml fill, and 1 34 &t% for the I ml. [0259] The lyophilization cycle had a total length of nearly 42 hours, Step N Descriptiot Temerature (0C) :ressure Tie (bh:mf) I Load 20 Atmospheric NA 2 Product freezing 20-*-45 Atmospheric 01:05 3 Freeze soak time -45 Atmospheric 02:00 4 Evacuation -45 200 pbar 00:01 1---- - --------------- ____ _ ___ _ ---- _ - ----- -

-----

5 Primary drying __-45-_ 0 200 bar 04:00 6 Primary drying 0 200 paar 02:00 Primary drying J 0- 05 20par 02:00 Primary drying 15 200 pLbar 20:00 48 9 Secondary drying 15-+30 200 bar 00:15 10 Secondary drying 30 200 pbar 11:00 I--............ --------------- _____ ___________ -. ________ ___ --- I 11 Pre-aeration 0.95 bar NA 12 Stoppering 0.95 bar NA 13 Aeration Atmospherie NA Total length 42:21 [0260] All three presentations had cake with a very cohesive structure even if some cracks were present. ANALYTICAL RESULTS In Process Controls Results Process step Analytical Test - 2A 2B 2C 2DE PH 3.910 3.86 3,83 3.69 3.72 Formulated ------ Bulk solution density (g/mL) 1027 1,028 1.030 1.0394 1.0389 49 Tests on Freeze-dried drug product t" Results Process step Analytical Test --- 0-2- 2A 2B2C 2D 2B Water content See by KF [% w/w] below Visual aspect or Conform Conform Conform Conform I Conform constitution 301 30" ~ " 5" See below Appearance of Final reconstituted Conform Conform Conform Conform Conform Lyophilizate solution (water, 50mg/ml) See pH 3,54 3.53 3.54 3.30 1 ee % Vancomycin 93,0 92,9 92,9 92,0 B IHCi [HPLC] below [ See % impurities 7.0 7.2 6.9 8,0 beo Moisture Content (K.F,) [wt%] Results Sample R.2E 2A 2B 20 2D 12ig240 mg 4 80 mng. Front sample 2,06 3 54el 192 1 021 ,4 1.26 0.96 Middle sample 1.40 3.12 1,80 1 ,01 1.27 1 ,15 .97 Back sample 1.68 4,17 174 0.92 1 127 1.27 0.97 Average 1.71 3.61 1.82 0.97 1.34 1.23 0.9 pH 50 Sample - - - ----- -e213 2A 2B3 2C 2D 10g20m 40m Frntamle 39 3.54 3.56 3,32 3,36 3,9 3,31 Front _ tapl 3 A__9 339 _ _ Middle sample 3.57 3,52 3'5 2 3.29 IM.3 3.38 3.33 Back sample 358 1 3,52 3.5 3,30__ 3,3-__ 339 3.1 I Average 3,54 M.3 3.54 3.30 13.36 39 .2 51 % Content Vancomycin B Hydrochloride (%VMB) Results Sample Reference S 2A 2B 2C 2E 2D VancomycI - 120mg (91.3 Frontsampie 92.9 92.9 929 93,7 240 mg 91.6 480 mg 92.5 120,21g 91.8 Middle sample 93.0 93.0 93,0 92, 1 240mg 91. 480mg 92.1 ----- ---- 20 92i Back sample 93.1 030 930 919 240 mg 91.2 480 mg 91 8 9120omg 91.7 Average 0 92.9 92.9 92.0 240 ing 91.5 480mg 92.1 -0ng 0,415 Standard Dev 104 0.0327 0.0327 0.108 240 mg 0261 480mg 0.343

--

120mg 0453 % RSD 0.112 0,0352 0.0352 0,117 -J40 mg 0235 80mg0.372 52 Related substances (% impurities) Results Sample Reference2C 2D 2E vancomyci ______~ -__ _ - i --- ---------- _ _ _ T 1o0mg 858 Front sample 71 72 69 63 m 8.4j 480 mg 7,5 120ming 8.2 Middle sample 7.1 7.3 6.9 9 240 mg 8.3 480 mng 7.9 ._ _ .. __ ._ __ _ ..... _ _ 7,9 Back sample 6,9 71 7.0 8.1 240mg. 8.8 480 mg 1 8,3 1 3120 mg 8.3 Average 7.0 7.2 6.9 8.0 240mng 85 480 mg 79 120 mg 0, 4-2 Standard Ue 0.12 0.09 0.013 0.11 240,mg 0.26 480mg 042 1.9 'dg 5.0 RSD1 1,2 0,23 1.3 240.g 3.1 I .48Orag 5.3 RECONSTITUTION TIME [0261] Reconstitution time measurement was carried out adding: 1.0 mL of WFI to the 120 mg/vial strength - 2,0 mL of WFI to the 240 mg/vial strength - 4.0 mL of WFI to the 480 mg/vial strength [0262] The observed reconstitution time on the product of Example 2E was quite short relative to all the tested vials; about 10 seconds were needed to completely reconstitute the 120 mg freeze-dried drug product; 10 to 15 seconds were needed to completely 53 reconstitute the 240 mg / vial presentation, while about 20 seconds were needed to completely reconstitute the 480 mg units. [0263] The reconstituted solution had a clear, light pinkish appearance and was particle free. COMPAT3ILITY WITH STERILITY TESTING MEMBRANE [0264] 20 mL of reconstituted drug product were passed through the sterility testing membrane to confirm the foru nation compatibility. (0265] The solution passed through the filter membrane, and 17 ml of the 20 ml were collected below the membrane. Example 3 SUMMARY [0266] This Example involves a freeze-drying cycle for a 600 mg of Vancomycin HCl / vial strength, MATERIALS / EQUIPMENT Materials e Vancomycin hydrochloride, USP, Alphanna - Denmark 0 ISO 6R clear type I glass vials, Nuova Ornpi - Italy 0 20 rum freeze-drying stoppers, West Pharmaceutical Service-USA * 20 mm flip-off caps, Capsulit S.p.A, - Italy * 13 mm freeze-drying stoppers, West Pharmaceutical Service-USA * 13 mm flip-off caps, West Pharmaceutical Service-USA 54 Equipment 0 Glassware for Vancomycin solution before and after ilt-ration (bottles), a Pressure vessel, Sartorius - Germany o Balance to check the filling weight (1 0 mg sensitivity), Sartorius - Germany 0 Digital pH meter, Mettler Toledo - Switzerland * Karl Fischer automatic titrator DL3S, Mettler Toledo - Switzerland 0 0,22 pm sterilizing PVDF filter, Pall * Semiautomatic filling machine, Flexicon PF6 - Denmark * Isolator, E.Co.Tec - Italy " Lyophilizer, BOC Edwards Lyoflex 04 (or Minifast 8000) with the following characteristics: 0A m 2 (or0.8 ir?) shelf surface, shelf temperature range was -504C to + 5002, PT 100 temperature probes, Pirani gauge for vacuum monitoring, coil condenser with ice capacity of 8 kg, condenser coil inlet, temperature anives to 6002, stainless steel trys with a thickness of about 2 mm " Semiautomatic crimping machine, Flexseal - Denmark FORMULATION [0267] Vancomycin hydrochloride was dissolved in water for injection to form a 120 mg/ml formulation, as shown below. Quantiy Ingredients Amount / m Amount / Unit Vancomycin H-C1 120.00 600.00 mg Water for injection to L Mi to 5.00 m1 MANUFACTURING PROCESS 0268] Water for injection was weighed out in a glass container on calibrated balances. 55 (0269] Vancomycin HC was added under stirring; the solution was agitated until vancomycin was completely dissolved and the dissolution time was recorded. [0270] Then, water for injection was added until the required final amount was reached. [02711 On the final solution, pH and density were measured and appearance was evaluated, [0272] The solution was filtered through a 0,22 urn PVDF membrane. [0273] The vials were washed with distilled water and dried in an oven at 120"C for 2h. [0274] The filling was performed by mass and the in process controls were carried out by weighing the filled vials every 20 vials. [0275] After lyophilization the following analyses were performed on the final product: - water content by Karl Fischer titration; - appearance of the cake, - reconstitution time, appearance/clarity, - pH after reconstitution. [0276] RP-HPLC was run to confirm processing didn't influence purity of Vancomycin, LYOPHILIZATION CYCLE [0277] The product was freeze-dried according the following nominal lyophilization cycle parameters: ---- -.-------------

----

Step NI Description Temperature (4Pessue Time (hh:mm) I Load 20 Atmospheric NA ------ -- ------ 1 .---- 2 f 20--45 Atmosphenc 01:05 3 Freeze soak time 45 Atmospheric 02:00 56 __- -_---_-__-------- 4 Evacuation -45 200 pbar 00:01 5 Primary drying -45-> 0 200 Iar 04:00 6 Primary drying 0 200 bar 02:00 Primary drying 0-+ 15 200 pbar 02:00 Primary drying 15 200 pbar 24:00 - Secondary drying 15 - 30 200 gtbar 00:15 8 Secondary drying 30 200 pbar 15:00 9 Preaeration 0.95 bar NA 10 Stoppering 0.95 bar NA 11 Aeration Atmospheric NA Total length (without stoppering) 50:21 RESULTS [0278] An overall average moisture content value of 1,04 wt% was found by Karl Fisher titration. [0279] Cakes had a very cohesive structure even if some cracks were present. ANALYTICAL RESULTS In Process Controls Process step Analytical Test Results pH 3.69 Formulated Density (g/mL) 1,0384 Bulk solution ------ 4 Concentration (UV) 116.88 mg/mL Tests on Freeze-died Drug Product 57 Process step Analytical Test Results Water content by KF 1,04 % w!w Visual aspect of the cake Whitish solid compact mass Reconstitution time 30 seconds Final Appearance of Lyophilizate I reconstituted solution Clear colorless solution (water, 50mg/ml) pH 3.44 % Vancomycin B by 933 % RP-HPLC Moisture content (K..) Sam i Sample I (back) 1.07 % Sample 2(iddie 1,02% Sample 3 (front) 1.02 % I Overall average 1,04 % HPLC Assay (% Vancomycin B) Sample I (back) 933 /o Sample 2 (rniddle) 93.3 % Sample 3 (front) 933 % Overall average 933 % pH Sm Sample I (back) 3.45 Sample 2 (middle) 3.44 Sample 3 (front) 3.44 Overall av ge 3 .. 44 RECONSTITUTION TIME 58 [02801 About 30 seconds were needed to completely reconstitute the freeze-dried drug product with 5.0 mL of WIL The reconstituted solution had a clear, colorless appearance and was particle free. 59 Example 4 SUMMARY [0281] Nebulization characteristics of gentamicin and vancomycin solutions were evaluated as a function of solution strength, nebulizer fill volume, and saline concentration. Key aerosol attributes measured were emitted dose and particle size distributions, All experiments were performed using Aerotech II jet nebulizers operated Continuously at 8 LFM. For gentamicin solutions in WFI, the range of solution strengths varied from 40 to 120 mug/mi, and fill volumes ranged from 2 to 4 ml, The resulting aerosol dose emitted over 30 minutes of nebulization was found to vary from 40 mg to over 300 mg, with the dose increasing proportionally with increasing fill volume and solution strength. Emitted dose measurements for vancomycin were performed for solutions in normal saline, in 0.45% saline, and in water for injection, Tfhe range of solution concentrations tested ranged from 60 mg/nil to 140 mg/mi. The cumulative aerosol dose emitted for a 30 minute nebulization period varied from about 50 mg to over 300 mg, with the dose increasing proportionally with solution strength and fill mass. [0282) Particle size distributions were measured for the above drug solutions using a laser diffraction spectrometer. The median particle size for all solutions tested was in the range 2 -- 3 pIm, well within the respirable size range. Particle size distributions for these antibiotic drugs were found to be relatively insensitive to solution strength and fill volume. Follow-on measurements with drug and normal saline solutions indicated that the size distribution of nebulized antibiotics were comparable to that for the normal saline solution. [0283] Combined together, the above results indicate that a broad range of aerosol doses in the respirable range may be achieved for nebulized vancomycin and gentamicin by suitably selecting nebulizer fill volume and solution strengths. OBJECTIVES 60 [0284] To determine the amount of drug aerosol emitted during the nebulization of gentanioin and vancomycin solutions, as a function of nebulizer fill volume and solution strength. [0285] To determine the size distribution of aerosols produced during the nebulization of gentamicin, vancomycin, and saline solutions as a fRunction of nebulizer fill volume and solution strength. INTRODUCTION [0286] This Exarmple involves assessing nebulization characteristics such as the emitted dose and droplet size distribution for antibiotic drug solutions of different strengths and at different nebulizer fill volumes. The emitted dose information is useful in selecting solution strengths and fill masses to deliver a chosen target dose, The particle size information is useful in determining whether the aerodynamic size of the aerosol produced is in the range required for effective lung deposition (I - 5 pm). Results for a placebo solution (i.e., normal saline) are also reported for comparison. All of the experiments were performed using an Aerotech II jet nebulizer operated continuously at a nominal flow rate of 8 LPM. Aerosol emitted dose was estimated by using filters to collect the aerosol output generated by the nebulizer, and assaying the amount of drug deposited. Particle size distributions of the generated aerosol were measured using a Sympatec laser diffraction spectrometer. 6 1 STUDY DESIGN Characterization of Emitted Dose [028I] For the case of gentamicin solution in water, a full factorial experiment was performed to characterize emitted rmass of aerosol as a function of two factors, i.e. nebulizer fill volume and fill mass. The range of solution strengths and fill volume was chosen to provide a broad range of target doses achievable with a nebulization time of 30 minutes. {0288] The test matrix for this experiment is presented in Table 1. Gentamicin solution strength (based on mass of drug) was varied from 40 mg/mI to 120 mg/m1, while the nebulizer fill volmine was varied from 2 to 4 ml. Each of the 9 treatment combination was repeated twice, for a total of 18 runs. The gentamicin solutions were prepared in water for injection (WFI), and were preservative free. [0289] For the case of vancomycin, the emitted mass of aerosol was haracterized for following three cases: e Vancomycin in normal saline, solution strength of 60 mg/ml, nebulizer fill volume ranging from 2 - 4 ml, * Vancomycin in 0.45% saline, solution strength ranging from 60 - 90 mg/ml, nebulizer fill volume ranging from 2 - 4 ml. * Vancomycin in WFI, solution strength ranging from 60 - 140 mg/mi, nebulizer fill volume ranging from 2 - 4 ml. [02901 In the case of vancomycin, addition of salt to the formulation allows for tuning of solution properties such as osmolality, Test matrices for the above three experiments are presented in Tables 2 - 4, 62 Table 1. Test Matrix for Gentamicin Solution in WFI Pattem Fl Volume ml] s o srnh 13 1 2 120 31 4 40 12 3 80 2 so 2 40 21 3 40 21 3 40 13 2 j 120 23 3 120 E 31 4 4 40 33 4 120 11 2 40 22 ] 3 80 12 2 80 23 3 120 33 4 120 2 4 - 80 32 4 80 J Table 2. Test Matrix for Vancomycin Solution (60 mg/ml) in Normal Saline Vancomycin at 60 mg/r (in nonnat saline) ill volume 2 ml FilIvolume 3 ml Fili volume 2 ml L.Fil Volume 4 m! Fnvolume 4 ml Fi volume 4 ml -Fill volume 3 il Fill volume 3 ml Fill volume 4 ml Fill volume 2 mit [0291) The responses measured for all of the above experiments included: (i) the mass of drug delivered in 15 mins (ii) the cumulative mass of drug delivered in 30 mins, and (iii) the mass of drug remaining in the nebulizer after 30 mins of operation. 63 Table 3. Test Matrix for Vancomycin Solution in 0,45% Saline Fi Volume Solution Strength PatternIm 11 60 2 - ------- 90 3 2 90 2 60 32 4 75 23 3 9o T 2 76 3 60 23 3 90 32 4 75 21 3 60 33 4 90 1 2 75 33 4 g0 22 3 75 31 4 60 31 4 60 3 75 Table 4. Test Matrix for Vancomycin Solution in WFI 1 Fill Solution Pattern Volume Strength [m} 1 ~mg/m. 2 60 2 140 13 2 140 i 2 60 32 4 100 23 140 12 2 100 21 3 60 23 3 140 32 4 100 21 3 60 33 4 140 12 2 100 33 4 140 22 3 100 31 4 60 31 4 60 22 3 100 64 Characterization of Particle Size Distribution [02921 For the case of gentamicin solution in water, a -full factorial experiment experiment was performed to characterize the particle size distribution of aerosol as a function of two factors, i.e. nebulizer fill volume and fill mass. The test matrix for this experiment is presented in Table 5, Gentamicin solution strength (based on mass of drug) was varied from 40 mg/ml to 120 mg/ml, while the nebulizer fill volume was varied from 2 to 4 ml, The 9 treatment combinations were run in a random order. A fresh nebulizer was used for each run. The nebulizers in this experiment were prequalified using a flow rate test to minimize variability in the test results. Table 5. Test Matrix for Gentamicin Solution in WFI Solution F -I1 Volume Strength Run Pattem [mL] mg/mL] 131, 4 40 2 32 r g 20 3 v1 3 40 --- - 2]3_E_:' 'E 3 ''. 120 5 22 ----- 3 go 3 2 120 T2 40 3 _3 420 [0293] For the case of vancomayin, the emnitted mass of aerosol was characterized for fol w n h ee cases: a Vancomycin in nonna1 saline, solution strength of" 60 mg/m1, nebulizer -fill volume ranging from 2 - 4 ml. * Vanconycin in 0.45% saline, soltion strength ranging from 60 - 90 mg/m l, nebulizer fll volume ranging from 2 - 4 ml, SVanCOmycin in WFI, solution strength ranging from 60 - 140 mg/ml, nebulizer fill volume ranging from 2 - 4 ml. 65 [0294] The test matrices for the above three experiments are presented in Tables 6 8. A fresh nebulizer was used for each run. The nebulizers in these experiments were pre screened using a flow rate test to minimize variability in the test results. Table 6. Test Matrix for Vancomycin Solution (60 mg/mil) in Normal Saline Solution Fill Volume Strength Run Pattern [mLI [mjgrnL] 1 1 2 60 2 3 60 Table 7, Test Matrix for Vancomycin Solution in 0.45% Saline Solution I Fill Volume Strength Run Pattern [mL [m g/mLi 1 12 2 75 31 4 60 3 22 3 75 j39 513 2g9 6 11 2 60 32 4 75 21 3 60 .3 .39 66 Table 8. Test Matrix for Vancomycin Solution in WAFI Solution Fill Volume Strength Run Pattern [mL' [mgimL] 33 4 140 2 22 3 1-00 3 13 2 140 412 2 100 5 1 2 60 6 32 4 100 7 21 360 23 3 140 9____ 3 ~ 1 4 50 [0295] A follow on experiment was performed to characterize particle size distributions of aerosols generated using vancomycin and gentamicin solutions in water at a fixed solution strength of 120 mg/ml, and a fixed fill volume of 5 ml. Particle size distributions of drug aerosol were compared against those obtained by nebulizing nonnal saline solution at a fill volume of 5 nl. The test matrix for this follow on experiment is presented in Table 9. Each treatment was repeated 3 times. Table 9. Test Matrix for Evaluation of Drug and Placebo Solutions Fill Volume Run DrUg [mLI I Normal Saline 5 2 Vancomycin 5 3 Gehtamki ~ 5 4 Gentatol 5 Normal Saline 6 - Geitariscir- 5< Vancomyc-n-- 5 Vanoomycir 5 9 Normalisaline EQUIPMENT AND MATERIALS Equipment Sympatec HELOS Magic BFS laser diffraction spectrometer, Ser. No. 085 67 & Mass flow meter (TSI 4000 series) 0 Rotameter * Volumetric flow meter, Dry Cal 0 Pressure regulator * Flow regulating valve * Flow shut-off valve * Pipet Materials * Aerotech II Nebulizer * Tee connector and mouthpiece from H-Iaudson RCI MicroMist Nebulizer (Cat No. 1882) * Inspiratory filter (PARI electret filter) * Filter holder 0 One way valve 6 50 ml centrifuge tubes * HPLC water * HPLC water dispenser * Vancomycin HCI * Gentamycin Sulfate 68 PROCEDURE Characterization of Emitted Dose [0296] The nebulizer was connected to a standard "T" piece coupled to a filter holder on one end, and a flow inlet channel provided with a one-way valve on the other end. The filter holder supported a PARI electret filter used to collect the aerosol dose emitted by the nebulizer. [0297] The nebulizer was operated using using clean, dry compressed air from a source regulated to a pressure of about 50 psig. The flow rate of air through the nebulizer was controlled using a rotameter and set to a nominal flow rate 8 LPM. The drug laden air from the nebulizer passed through the collection filter into an exhaust line provided with a backup filter and a flow regulating valve, and connected to a vacuum source. The flow regulating valve was set so that the vacuum suction flow was slightly higher th an the nebulizer output flow. A small amount or clean make up air was allowed to enter through the one way valve to make up for the flow deficit, This arrangement enabled efficient collection of the nebulizer drug output by the filter. The emitted dose experiments were performed with the nebulizer operating continuously at 8 LPM for a total nebulization time of 30 minutes. The filter/filter holder were replaced with a fresh filter/filter holder at the 15 minute point, so that the accumulated drug output at 15 minutes and 30 minutes could be evaluated. The filter samples were placed in centrifuge tubes and rinsed with a pre-deternined amount of HPLC water (ranging from 30 - 40 ml). Residual drag from each filter holder was also rinsed into the corresponding centrifuge tube using some of the filter rinsate. The residual drug from the neLulizer was also rinsed into a 50 ml centrifuge tube using a pre-detenriined amount of HPLC water (ranging from 30 - 40 ml). The drug content of the filter and nebulizer samples were assessed by drug specific IPLC assays. Note that the measurement of filter and nebulizer samples permit a full mass balance to be performed for each run. Characterization of Particle Size Distribution 69 [0298] Droplet size distributions for aerosolized drug and placebo solutions were measured using the Sympatec HELOS laser diffraction spectrometer. in preparation for a run, the nebulizer was connected to the compressed air line, the flow turned on and the pressure regulator set to a driving pressure to generate a flow rate of 8 LPM through the nebulizer. The flow was then turned off by closing the flow shut-off valve. Next, the nebulizer was conrected to a "T" piece with one port plugged, and the other port coupled to a mouthpiece. The nebulizer was then filled with drug solution, and mounted so that nebulizer mouthpiece was aligned parallel to the nozzle of the Rodos dry powder disperser apparatus already installed in the spectrometer. The laser diffraction system, was setup to automatically trigger when it sensed the presence of the aerosol generated by the nebulizer, Measurernents were initiated by opening the shut-off valve to pressurize the nebulizer and generate the aerosoL A total of 6 particle size distribution scans were taken for each nebulizer run, and then averaged to provide representative size distribution results. RESULTS AND DISCUSSION Characterization of Emitted Dose [0299] Summarized dose delivery results for the case of gentamicin solutions are presented in Figs. 3-5. Fig. 3 is a bar graph showing the total drug recovered from the nebulizer and as a function of nebulizer fill volume and solution strength. Each recovery value is the average of two replicate runs (run order listed in Tablel). The drug recovery was very consistent across solution strengths and fill volumes, varying in the range 97,1% 101.2% of fill mass, indicating that a full mass balance was achieved from these measurements. [0300] Figs. 4a and 4b present the cumulative emitted dose of gentamicin, respectively at the 15 min and 30 mrin time points, as a function of fill volume and solution strength. Again, each value reported is the average of two replicate runs. The delivered dose was observed to increase with an increase in both fill volume and solution strength, consistent with expectation. A comparison of these two figures shows that the collected dose at 15 minutes was comparable to that at 30 minutes for 2 and 3 ml fill volumes, indicating that the dose emission at these fill volumes occured within 15 minutes. For the 4 mi fill volume, the collected dose at 30 mins was only slightly larger than the value at 15 minutes, indicating that 70 nebulization was largely completed within the 15 minute period. From this it can be concluded that fill volumes of up to 4 ml of gentamicin solution of strengths up to 120 mg/mi can be effectively nebulized within a duration of 30 minutes, Fig. 4b also indicates that a gentamicin aerosol doses spanning a factor of up to 7 can be delivered from the nebulizer by suitably tuning the solution strength and fill volume within the ranges tested. [0301] Fig. 5 presents the gentamicin dose retained by the nebulizer at the end of 30 minutes, as a function of solution strength and fill volume, The values reported are averages of two replicate runs. The retained dose was found to increase with increasing solution strength and fill volume, with a steeper increase observed with increasing solution strength. [0302] Similar trends in emitted dose as a function of solution strength and fill volume were obtained for the case of vanconycin, Illustrative emitted dose measurernents for vancomycin are presented in Figs. 6-8, [0303] For the case of 60 mg/m solution in normal saline (see Table 2), Fig. 6 plots 'he distribution of vancomycin drag after 30 minutes of nebulization as a function of fill volume, The plot shows the dose retained in the nebulizer and that collected at the 15 minute (filter I) and 30 minute (filter 2) timepoint, The reported values are averages calculated for 3 replicate runs, As with the case of gentamicin, dose emission was found to be largely completed within 15 minutes, and the accumulated dose (i,e, filter I + filter 2) at the end of 30 minutes was found to increase with fill volume, {0304] For the case of vancomycin solutions in 0.45% saline (see test matrix in Table 3) Fig, 7 plots the cumulative ernitted dose after 30 minutes of nebulization as a function of solution strength and fill volume. The delivered dose was observed to increase with increasing fill volume and solution strength, as expected. Fig, 8 plots similar results for the case of vancomycin solutions in WFI, obtained for the test matrix presented in Table 4. [0305] It is clear from Figs. 6 - 8 that aerosol doses of vancomycin spanning a six fold range can be obtained from the nebulizer by suitably tuning the fill volume and solution strength within the ranges tested, Characterization of Particle Size Distribution 71 [0306] Representative laser diffraction particle size measurements for the case of gentarnicin solutions (test matrix of Table 5) are summarized in Figs. 9 and 10. Fig. 9 plots the volume median diameter for aerosolized gentamicin as a function of fill volume and solution strength (test matrix in Table 5). Each reported value was obtained by averaging 6 replicate laser diffraction measurements for each nebulization run. The measured median particle size for all of the gentarnicin solutions varied slightly in the 2 - 3 pnm range, and appeared to be relatively insensitive to fill volume or solution strength. In all cases, the median particle diameter was well within the "respirable size range" considered to be suitable for pulomary drug delivery (I - 5 gim). Fig. 10 plots the cumulative volume weighted particle size distributions for gentamicin aerosol for all of the solution strengths and fill volumes tested. The size distributions obtained for these solutions were observed to vary within a narrow range over the fill volumes and solution strengths tested. Fig, 10 also provides a measure of the spread of the aerosol size distribution, and it was observed that a maor fraction of the aerosol was within the respirable size range. [0307] Representative particle sizing measurements for vancomycin solutions in WFI (test matrix in Fable 8) are presented in Fig. 11 and 12, and are roughly comparable to that obtained for gentamicin solutions. [03083 Fig. 11 indicates that the volume weighted median sizes for these vancomycin solutions were largely within the range of 2 - 3 prn, also well within the respirable range. The spreads of the aerosol size distribution, shown in Fig. 12, were similar to that obtained for nebulized gentamicin. [0309] Fig. 13 and 14 are plots of volume median diameter for the case of vancomycin solutions in normal saline (test matrix in Table 6), and 0.45% saline (test matrix in Table 7) respectively, obtained at different solution strengths and fill volumes. The size distributions were found to be comparable to that obtained for the vancomycin solutions in water. In general, the size distributions of vancomycin solutions were largely insensitive to fill volume, solution strength, and saline concentration. [0310] Finally, results from tie follow-on particle sizing study with the test matrix listed in Table 9 are presented in Fig. 15. This figure plots volume median diameters for solutions of vancomycin (120 mug/mi), gentamicin (120 mg/mi) and normal saline, all obtained for nebulizer fill volumes of 5 ml. For each solution, results from three nebulizer 72 runs are provided. It is seen from this plot that the median particle size for all three solutions were comparable and were in the 2 - 3 pm range, well within the respirable size range. CONCLUSIONS [0311] The emitted dose of nebulized gentamicin and vancomycin was measured as a function of solution strength, fill volume, and saline concentration. All experiments were peroirned using Aerotech II jet nebulizers operated continuously at 8 LPM. For gentamicin solutions in WFI, the range of solution strengths varied from 40 to 120 mg/mi, and fill volumes ranged from 2 to 4 ml, The resulting aerosol dose emitted over 30 minutes of nebulization was found to vary from 40 mg to over 300 mg, with the dose increasing with increasing fill volume and solution strength. Emitted dose measurements for vancomycin were performed for solutions in nomnal saline, in 0.45% saline, and in water for injection. The range of solutions tested ranged from 60 mg/ml to 140 mg/ml. The cumulative aerosol dose emitted over a 30 minute nebulization period varied from about 50 mg to over 300 mg, with the dose increasing with solution strength and fill mass. [0312] Particle size distributions were measured for the above drug solutions using a laser diffraction spectrome t er. The median particle size for all solutions tested was in the range 2 - 3 km, well within the respirable size range, Particle size distributions for these antibiotic drugs were found to be relatively insensitive to solution strength and fill volume. Follow-on measurements with drug and normal saline solutions indicated that the size distribution of nebulized antibiotics were comparable to that for the normal saline solution. [0313] Combined together, the above results demonstrate that a broad range of aerosol doses in the respirable range may be achieved for nebulized vancomycin and gentamicin by suitably selecting nebulizer fill volume and solution strengths. Example 5 [0314) This Example involved evaluating the potential toxicity and recovery resulting from a 14-consecutive day, nose-only inhalation administration of vancomycin hydrochloride (vancomycin) to CD rats. 73 [0315] With in 2 hours prior to usage, a vancomycin nebulizer solution having a concentration of 120 mg/ml (based on vancomycin potency of bulk material) was formed by dissolving vancomycin hydrochloride (available from Alpharma, Copenhagen, Denmark) in sterile water for injection USP (available from B. Braun Medical Inc., Bethlehem, PA), The solution was used to generate aerosolized vancomycin for all vancomycin exposure groups. [0316] Nose-only exposures were conducted in a "flow-past" cylindrical inhalation chamber placed inside a steel-framed Plexiglas secondary containment box, The chamber contained 48 animal ports, each compatible with a single nose-only exposure tube, aerosol concentration sampling device (e.g., filter), or oxygen monitor. [0317] The total air flow through the exposure system was balanced to achieve individual animal port flows of-500 mL/min (port flow approximated based on total chamber flow). Measured flows included sample flow rate, nebulizer flow rate, dilution flow rate (chamber make-up air), and chamber exhaust flow. The exposure chamber had a slightly higher exhaust flow rate than inlet flow rate. [0318] Vancomycin solution was aerosolized with two Aerotech II nebulizers operated at 20 psi driving pressure, The target aerosol Vancomycin concentration for all exposure levels was -1 .0 mg/L. [03191 Aerosolized vancomycin was administered to 3 groups of male and female CD rats (available from Charles River Laboratories, Kingston, NY) for durations of 30 min (Low), 90 min (Mid), and 180 min (High). A control group was exposed for 180 min to aerosols generated from a normal saline solution. Groups of rats from the Control and High level 14-day exposures were also studied following a 14-day recovery period. Endpoints included clinical observations, body weights, clinical pathology (hematology, clinical chemistry), urinalyses, organ weights, and histopathology. [03201 Vancomycin aerosol concentrations were 23 0,16, 1.25 i 0.12, and 1.23 a 0.08 mg/L for the Low, Mid, and High exposure levels, respectively. Mean particle size was determined to be in the inhalable size range for rodents (2.0-2.6 pm mass median aerodynamic diameter). Mean total inhaled doses were estimated as 23, 71 and 139 mg/kg, and mean doses deposited in lung were estimated as 3, 9, and 17 mg/kg for the Low, Mid, and High exposure levels, respectively. 74 [0321] The vancomycin exposures were well-tolerated by all groups of rats. All rats survived to scheduled necropsy, and there were no vancomycin related effects noted on clinical observations. There were also no vancomycin treatment related effects on body weight. [0322] The ory organ weights to show consistent vancomycin related effects were lungs. Lung weiLhts were statistically significantly increased by an average approximately 8, 20, 19 % of control for the Low, Mid and High exposure levels respectively. [0323] Exposure related histopathologic findings were limited to the respiratory tract. Observations included minimal to mild nasal mucous cell hyperplasia and hypertrophy, minimal to mild pulmonary interstitial inflammation and alveolar macrophage hyperplasia with an apparent dose-response effect, lymphoid hyperplasia of the tracheobronchial and mediastinal lymph nodes, and slight laryngeal inflammation. There was substantial diminution of these findings after 14 days of recovery with pulmonary interstitial inflammation, alveolar macrophage hyperplasia, and nasal mucus cell hyperplasia persisting in the high dose group, but at a lesser severity overall than seen at the end of exposure. A threshold of response was not established although the effects in the low dose group were generally minimal. [0324] Clinical pathology findings were generally unremarkable. The only vancomycin related effect on hematology was a statistically significant increase in neutrophils at the Mid and High exposure levels. The only vancomycin related effect on clinical chemistry was a mild but statistically significant increase in aspartate aminotransferase (AST) values (~28-46%) at the Mid and High exposure levels. Neutrophil changes were diminished after the recovery period resolved. AST observations resolved after the recovery period. Both findings likely resulted from the minimal to mild pulmonary inflammation manifested in the histopathology findings. No vancomycin related changes were seen after examination of serum indicators of kidney function or urinalysis. [0325] To conclude, the findings indicate that exposure to vancomycin at the Mid level and High level exposures, predominantly, caused an irritant reaction in the respiratory tract manifested by minimal to moderate mucous cell changes in the nose and minimal to mild inflammation and macrophage hyperplasia in the lungs. Corresponding changes in neutrophil and AST vaines likely resulted from the pulmonary inflammatory findings. 75 Recovery from these effects was evident, but not entirely resolved after the 14-day observation period. A no observed effect level was not established. Example 6 [0326] This Example involved evaluating the potential toxicity and recovery resulting from a 14-consecutive day, face mask inhalation administration of vancomycin hydrochloride to beagle dogs. [03271 Within 2 hours prior to usage, a vancomycin nebulizer solution having a concentration of 120 mg/mil was formed by dissolving vancomycin hydrochloride (available from Alpharma, Copenhagen, Denmark) in sterile water for injection USP (available from B. Braun Medical Inc., Bethlehem, PA). The solution was used to generate aerosolized vancomycin for all vancomycin exposure groups. [0328] The exposure system consisted of a single, cylindrical, plexiglass inhalation chamber (volume of - 23 '7 L, 14.61-rcm radius, 35,56-cm height). The chamber was supplied with two Aerotech II nebulizers operated at ~ 40 psi. Nebulized test article and nebulizer air supply was diluted with ~ 10 L/min HEPA-filtered dilution air, The flow through the system was ~ 36 L/min, [0329] The aerosolized vancomycin was administered via a face mask to 3 groups of male and female beagle dogs for durations of 15 min (Low), 30 min (Mid), and 60 min (High). A control group was exposed for 60 min to aerosols generated from normal saline solution, i.e., 0.9% sodium chloride injection USP (available from B. Braun Medical Inc.), [0330] Groups of dogs from the Control and High level 14-day exposures were also studied following a 14-day recovery period. Endpoints for all groups of dogs included physical examinations, clinical observations, body weights, ophthalmology, cardiovascular EKG, clinical pathology (hematology, clinical chemistry) urinalysis, organ weights, histopathology, and toxicokinetics. [0331] Vancomycin aerosol concentrations were 1.39 i 0.20, 1,51 i 0.19, and 1,49:1 0.15 mg/L for the Low, Mid, and High exposure levels, respectively. Mean particle size was determined to be in the inhaleable size range for dogs (1.9-2.6 pm mass median aerodynamic diameter). Mean total inhaled doses were estimated as 10, 23, and 45 mg/kg, and mean doses 76 deposited in lung were estimated as 2, 5., and 9 mg/kg for the Low, Mid, and High exposure levels, respectively. [0332] The vancomycin exposures were well-tolerated by all groups of dogs. All dogs survived to scheduled necropsy, There were no vancomycin related effects noted on physical examinations, clinical observations, ophthalmology, cardiac ECG tracings, hematology, clinical chemistry, urinalyses, gross necropsy observations, and organ weights. [0333) Histopathology examinations of tissues revealed no effect of Vancomycin exposure in the organs and tissues examined outside of the respiratory tract. Likewise, there was an absence of microscopic alterations in the nasal cavity/turbinates, larynx, and trachea. The effects of Vancomycin exposure were limited to microscopic findings in the lung. Treatment-related increased incidence of minimal to mild chronic interstitial inflammation, alveolar histiocytosis, and bronchial lymph node lymphoreticular hyperplasia were observed. Among Control and High level animals in the Recovery groups there were no treatment related differences in the macroscopic and microscopic findings. [0334] In conclusion, effects of Vancomycin exposure were limited to minimal to mild pulmonary histopathology at the termination of exposure. Recovery of histopathological effects was complete after 14 days. The minimal to mild chronic interstitial inflammation was generally comparable with background inflarnmatory changes in beagle dogs. The alveolar histiocytosis was reflective of enhanced clearance that occurs without alveolar injury. The ymphoreticuilar hyperplasia was considered an adaptive response that facilitates lung clearance mechanisms. Since corresponding fibrosis and alveolar epithelial injury were not characteristic of the observed effects, the lung changes and related lymph node changes were not considered adverse effects. Based on these findings, the no observed adverse effect level (NOAEL) was the high exposure level corresponding to an inhaled dose of 45 mg/kg and a deposited hng dose of 9 mgkg. Example 7 [0335] Amikacin Sulfate sterile solution for inhalation, 125mg/ml was manufactured and characterized as follows, Approximately 13.5 L of sterile water for injection was added to a glass carboy fitted with a lightning labmaster mixer. Arnikacin sulfate was added to the carboy and the solution was stirred. The solution was mixed until the entire API was 77 dissolved. A sample of the solution was taken and pH measured, With continued stirring, pH was adjusted with 1 ON HC to be within 5,5-6.3 with a target pH of 5.9, After pH adjustment, sufficient quantity of sterile water for injection was added until the final weight of solution of 21,328 g. was reached. The pH of the final solution was verified to be within an acceptable range, The solution was then sparged with filtered nitrogen at a rate of 1.5L/min for 15 minutes. The solution was then filtered through the 0.22 micron sterile filter. [0336] Prior to filling the solution, each viai was purged with nitrogen, The solution was filled by weight using a Cazzoli filler/stopper machine into 5 ml amber vials to a target weight of 4.27 ± 0,08 g. The vials were stoppered with 20 mm Teflon-coated stoppers and secured with aluminum flip off seals. Filled vials were stored at 2 - 80C The composition is summarized in Table A below. Table A Ingredient g per batch Amikacin Sulfate 3525.0 Hydrochloric Acid qs to pH 5 9 NaOH qs_ ,opH5,9 Sterile Water for Injection qs to 21, 328 Nitrogen, NF QS [03371 Stability over time was assessed for as formulation made substantially as show in table A, with regard to total amikacin active, related substances, such as degradation products, appearance, pH, particulates and sterility, Thus samples were stored at 50C (Table B), at 25"/60% relative humidity (RH) (Table C), and at 404C/75%RH (Table D). In each case samples were stored in 5 mL amber glass vials, with 20 mm Teflon stoppers and 20 mm aluminum overseals. Results of each of these storage conditions are shown in Tables B, C and D, respectively. Table B 111t al s 5_ ec o - ----- I - - M:__ b MO o7s. 12 m" 1 Mos. 2M.0% - 110.0% Ls, 99S5 IOU. 104.0 104.3 10419 100M0 105.5 570c Meets Test' MAT MTf jT MT. JI WTITM 78 in @ rr Renor resuas V3 M4. 0 - 033 028 022 M Rnce A Popori results 06 010.46 ed @ nt eporq esIts 6g0 631 -3 , t 3.02 2 T90 229 6,27 d Report result 0.3 0,45 A5 023 .26 021 100% -22 8.74 620 40 4.07 3.89 3.10 5. - .01 telre Pariiles=10pm 50 3 7 24 I 36 25 NMT 6000 Particles=25pm 0 0 Q T-sI ----- ---- cnems NP _____ ______ NMT 600 N P N M -____ Meets USP oims N NP P TN Table C Attributes Spe mfations Initial i mo. 3 mos- -- 6 mos Assay 90.0%~110.0% 998 104.0 105.3 1001 __________t s. 99.8 ____ ______ Appearance ;Meets Test MT MT MT MT Kanamycin rrt Report results 0.73 0,81 0,1 0.44 032 0.64 Re. Substance A Report results 0,66 072 0.18 0.59 @ rit 086 0 60 IUnientified @ ri Report results 6.90 6.87 5128 3.55 0.61 627 Unidentified @ rrt Report results 053 0.50 0,49 028 0.67 .50 Total Related 10,% 8.82 8,90 5.66 4,86 Substances 8.0 5.5-6.3 5.6 5.6 Particulate Malte Particles = 1 Op m 1 18 33 NMIT 6000 Partiles=25pm 0 0 NMT 600 Sterlity Meets USP oniorms NP NP NP M e-ts - ------------------ Table D Attributes I Specifications iInitial 1 mc. 3 mos. 8 moes Assay 1 90.0%- 110% Is. 99.8 104,1 1047 106,3 99.8 - --- ---- ---- 79 Appearance Meets Test 1 MT T MT clear, faint yellow solution Kanamyci @ nir Report results 073 0.99 100 0.72 0.64 Ret Substance A @ Report results 0.66 0.71 0,16 0.60 rt0,86 0.60 Unidentied @ rr Report results 6,90 6.81 4.64 362 0.61 6.27 Unidentified @ irt Report results 0.53 0.54 0.60 0.61 0.67 0.50 TotalRelated 10.0% 88 9.05 646 573 Substances 6 8.01 ....... _ 5 6. 3 6 5.6 5.6 o.5 5,5 aticulate Matter Particles= 10pr5 32 20 27 NMT 6000 Partilkes=25pm 0 1 0 NMT 600 e ty Meets. U SP ConAorms NP NP NP [03381 Fig 16 is a graphical representation of certain of the stability data provided in Tables B, C and D. In the Fig., the line marked by diamonds represents the 51C storage condition, the line marked by the squares represents 250C/60% RH storage conditions and the line marked by the triangle represents 400C/75% RH storage conditions. The Fig shows that the percentage related substances, i.e. impurities, diminishes over storage time, It is thought that this is a function of detactability of the impurities. It is evident, however, that the compositions remain stable, with respect to impurities, over time, {0339i Having now fully described this invention, it will be understood to those of ordinary skill in the art that the methods of the present invention can be carried out with a wide and equivalent range of conditions, formulations, and other parameters without departing from the scope of the invention or any embodiments thereof. [0340] All patents and publications cited herein are hereby fully incorporated by reference in their entirety. The citation of any publication is for its disclosure prior to the filing date and should not be construed as an admission that such publication is prior art or that the present invention is not entitled to antedate such publication by virtue of prior invention, 80 [0341] Also disclosed is: 1. An aqueous composition for aerosolization comprising: anti-gram-negative antibiotic or salt thereof being present at an amount from about 100 mg/ml to about 200 mg/ml. 2. The aqueous composition of item 1, wherein the aqueous composition consists essentially of the anti-gram-negative antibiotic or salt thereof and water. 3. The aqueous composition of item 1, wherein the amount of the anti-gram negative antibiotic or salt thereof is from about 110 mg/ml to about 150 mg/ml, or a potency from about 500 p.g/mg to about 1100 p.g/mg, or both. 4. The aqueous composition of item 1, wherein the anti-gram-negative antibiotic or salt thereof comprises aminoglycoside or salt thereof 5. The aqueous composition of item 1, wherein the anti-gram-negative antibiotic or salt thereof comprises at least one member selected from gentamicin, amikacin, kanamycin, streptomycin, neomycin, netilmicin, paramecin, tobramycin, and salts thereof 6. The aqueous composition of item 1, wherein the anti-gram-negative antibiotic or salt thereof comprises amikacin or salt thereof 7. The aqueous composition of item 1, wherein the aqueous composition has a pH from about 4 to about 6. 8. The aqueous composition of item 1, wherein the aqueous composition has an osmolality ranging from about 90 mOsmol/kg to about 500 mOsmol/kg. 9. The aqueous composition of item 1, wherein the aqueous composition is preservative-free. 10. The aqueous composition of item 1, further comprising an additional active agent. 11. The aqueous composition of item 1, further comprising an additional active agent selected from an anti-inflammatory and a bronchodilator. 12. The aqueous composition of item 1, further comprising a bronchodilator selected from p-agonist, anti-muscarinic agent, and steroid. 13. The aqueous composition of item 1, further comprising albuterol. 14. The aqueous composition of item 1, further comprising an osmolality adjuster. 15. The aqueous composition of item 1, wherein no precipitate forms in the 81 4435277_2 (GHMatters) P77144.AU.1 aqueous composition when the aqueous composition is stored for 1 year at 25 'C. 16. The aqueous composition of item 1 and further including an anti-gram positive antibiotic. 17. The aqueous composition of item 1, wherein the anti-gram-negative antibiotic or salt thereof comprises gentamicin or amikacin or a salt thereof, wherein the aqueous composition consists essentially of the gentamicin or salt thereof and water, wherein the gentamicin or salt thereof is present in an amount from about 80 mg/ml to about 140 mg/ml, wherein the aqueous composition has a pH from about 3 to about 7, and wherein the aqueous composition has an osmolality from about 120 mOsmol/kg to about 300 mOsmol/kg. 18. An aqueous composition, consisting essentially of: An anti-gram-negative antibiotic or salt thereof; a bronchodilator; and water. 19. The aqueous composition of item 18, wherein the anti-gram-negative antibiotic or salt thereof is present in an amount ranging from about 100 mg/ml to about 200 mg/ml. 20. The aqueous composition of item 18, wherein the anti-gram-negative antibiotic or salt thereof comprises an aminoglycoside or salt thereof. 21. The aqueous composition of item 18, wherein the aqueous composition has a pH from about 3 to about 7, and an osmolality from about 90 mOsmol/kg to about 500 mOsmol/kg. 22. The aqueous composition of item 18, wherein the bronchodilator is present in an amount of at least about 1 mg/ml. 23. The aqueous composition of item 22, wherein the bronchodilator is selected from -agonist, anti-muscarinic agent, and steroid. 24. The aqueous composition of item 23, wherein the bronchodilator comprises albuterol or salt thereof 25. An aqueous composition, comprising: an anti gram-positive antibiotic or salt thereof being present at a concentration from about 0.6 to about 0.9 of the water solubility limit, at 25'C and 1.0 atmosphere, of the anti-gram-positive antibiotic or salt thereof 26. The aqueous composition of item 25, wherein the anti-gram-positive antibiotic 82 4435277_2 (GHMatters) P77144.AU.1 or salt thereof is present in an amount ranging from about 60 mg/ml to about 140 mg/ml, or an osmolality of 90 mOsmol/kg to about 500 mOsmol/kg, or both. 27. The aqueous composition of item 25, wherein the anti-gram-positive antibiotic or salt thereof comprises macrolide or salt thereof 28. The aqueous composition of item 25, wherein the anti-gram-positive antibiotic or salt thereof is selected from vancomycin, erythromycin, clarithromycin, azithromycin, and salts thereof 29. The aqueous composition of item 28, wherein the aqueous composition is preservative-free. 30. The aqueous composition of item 25, further comprising an additional active agent selected from anti-inflammatory, bronchodilator, osmolality adjuster, and combinations thereof. 31. The aqueous composition of item 30, further comprising bronchodilator selected from p-agonist, anti-muscarinic agent, and steroid. 32. The aqueous composition of item 25, further comprising albuterol or salt thereof. 33. The aqueous composition of item 25, wherein the aqueous composition is made from a powder that has a potency greater than about 900 tg/ml after the powder is stored for 1 year at 25 'C. 34. The aqueous composition of item 25, wherein the anti-gram-positive antibiotic or salt thereof comprises vancomycin or salt thereof, wherein the osmolality adjuster comprises sodium chloride, wherein the aqueous composition consists essentially of the vancomycin or salt thereof, the sodium chloride, and water, wherein the vancomycin or salt thereof is present in an amount ranging from about 60 mg/ml to about 140 mg/ml, wherein the aqueous composition has a pH ranging from about 3 to about 7, wherein the aqueous composition has an osmolality ranging from about 120 mOsmol/kg to about 300 mOsmol/kg. 35. A unit dose, comprising: a container; and an aqueous composition, comprising an antibiotic or salt thereof being present at a, and wherein the composition is preservative free. 36 The unit dose of item 35 wherein, the antibiotic comprises an anti gram 83 4435277_2 (GHMatters) P77144.AU.1 negative, and is present at a concentration from about 90 mg/ml to about 200 mg/ml. 37. The unit dose of item 36 wherein the antibiotic comprises amikacin. 38 The unit dose of item 35 wherein, the antibiotic comprises an anti gram positive, and is present at a concentration from about 550 mg/ml to about 900 mg/ml. 39. The unit dose of item 38, wherein the anti-gram-positive antibiotic or salt thereof comprises a macrolide or salt thereof 40. The unit dose of item 35, wherein the antibiotic comprises amphotericin B. 41. A kit, comprising: a first container containing a first aqueous solution comprising a first antibiotic or salt thereof; and a second container containing a second aqueous solution comprising a second antibiotic or salt thereof, wherein the first and second antibiotics are the same or different, and wherein a concentration, or an amount, or both of the first antibiotic is the same as, or different from an a concentration, or an amount, or both of the second antibiotic. 42. The kit of item 41, wherein an amount of the first aqueous solution is from about 2 ml to about 5 ml, an amount of the second aqueous solution is from about 5 ml to about 8 ml, a concentration of first antbiotic or salt thereof is from about 100 mg/ml to about 120 mg/ml, and a concentration of second antibiotic or salt thereof is from about 120 mg/ml to about 140 mg/ml. 43. The kit of item 41, wherein the anti-gram-negative antibiotic or salt thereof comprises aminoglycoside or salt thereof 44. The kit of item 43, wherein the anti-gram-negative antibiotic or salt thereof comprises amikacin or a salt thereof 45. The kit of item 41, further comprising an aerosol introducer. 46. The kit of item 41, further comprising a vibrating mesh nebulizer. 47. A kit, comprising: a first container containing a first aqueous solution comprising anti-gram negative antibiotic or salt thereof; and a second container containing a second aqueous solution comprising anti-gram negative antibiotic or salt thereof, 84 4435277_2 (GHMatters) P77144.AU.1 wherein a concentration, an amount, or both, of the anti-gram-negative antibiotic or salt thereof of the first aqueous solution is different from a concentration, or an amount, or both, of the anti-gram-negative antibiotic or salt thereof of the second aqueous solution. 48. The kit of item 47, further comprising an aerosol introducer. 49. The kit of item 48, further comprising a vibrating mesh nebulizer. 50. A unit dose, comprising: a container; and a powder comprising anti-gram-positive antibiotic or salt thereof, wherein the powder is present in an amount ranging from about 550 mg to about 900 mg. 51. The unit dose of item 50, wherein the anti-gram-positive antibiotic or salt thereof comprises a macrolide or salt thereof 52. The unit dose of item 50, wherein the anti-gram-positive antibiotic or salt thereof is selected from vancomycin, erythromycin, clarithromycin, azithromycin, and salts thereof. 53. The unit dose of item 50, wherein the powder has a specific surface area ranging from about 5 m 2 /g to about 20 m 2 /g. 54. A kit, comprising: a first container containing a first composition comprising anti-gram-positive antibiotic or salt thereof; and a second container containing a second composition comprising water, wherein the first composition and/or the second composition comprises osmolality adjuster. 55. The kit of item 54, wherein the first composition comprises powder. 56. The kit of item 54, wherein the first composition comprises the osmolality adjuster. 57. The kit of item 54, wherein the second composition comprises the osmolality adjuster. 58. The kit of item 54, further comprising an aerosol introducer. 59. The kit of item 58, further comprising a vibrating mesh nebulizer. 60. A kit, comprising: 85 4435277_2 (GHMatters) P77144.AU.1 a first container containing a first antibiotic or salt thereof; and a second container containing a second antibiotic or salt thereof, wherein the first and second antibiotics a re the same or different, and wherein a concentration, or an amount, or both of the first antibiotic or salt thereof in the first container is different from a concentration, or an amount, or both, of the second antibiotic or salt thereof in the second container. 61. The kit of item 60 wherein each of the first and second antibiotics comprise a powder. 62. The kit of item 60 wherein each of the first and second antibiotics comprise a liquid. 63. The kit of item 60 wherein one of the first or second antibiotics comprises a powder, and the other of the first or second antibiotics comprises a liquid. 64. The kit of item 60, wherein the solution is preservative-free. 65. The kit of item 60, further comprising an aerosol introducer. 66. The kit of item 65, further comprising a vibrating mesh nebulizer. 67. A method of administering an antibiotic formulation to a patient in need thereof, comprising: aerosolizing an antibiotic formulation to administer the antibiotic formulation to the pulmonary system of the patient, wherein the antibiotic formulation has a concentration of anti-gram-negative antibiotic or salt thereof ranging from about 100 mg/ml to about 200 mg/ml. 68. The method of item 67, wherein the antibiotic formulation consists essentially of anti-gram-negative antibiotic or salt thereof and water. 69. The method of item 67, wherein the aerosolizing comprises forming droplets having a mass median aerodynamic diameter of less than about 10 gim. 70. The method of item 69, wherein the aerosolizing comprises aerosolizing the antibiotic formulation in a vibrating mesh nebulizer. 71. The method of item 67, wherein the pulmonary administration comprises prophylactic treatment of ventilator associated pneumonia. 72. A method of administering an antibiotic formulation to a patient in need thereof, comprising: 86 4435277_2 (GHMatters) P77144.AU.1 inserting a tube into a trachea of a patient; and aerosolizing an antibiotic formulation to administer the antibiotic formulation to the lungs of the patient, wherein the antibiotic formulation consists essentially of anti-gram-negative antibiotic or salt thereof and water. 73. The method of item 72, wherein the pulmonary administration comprises prophylactic treatment of ventilator associated pneumonia, hospital associated pneumonia, community acquired pneumonia, or combinations thereof 74. The method of item 72, wherein the aerosolizing comprises aerosolizing the antibiotic formulation in a vibrating mesh nebulizer. 75. A method of administering an antibiotic formulation to a patient in need thereof, comprising: dissolving an anti-gram-positive antibiotic or salt thereof in a solvent to form an antibiotic formulation, wherein the anti-gram-positive antibiotic or salt thereof is present at a concentration ranging from about 0.6 to about 0.9 of the water solubility limit, at 25'C and 1.0 atmosphere, of the anti-gram-positive antibiotic or salt thereof; and aerosolizing the antibiotic formulation to administer the antibiotic formulation to the lungs of the patient. 76. A method of administering an antibiotic formulation to a patient in need thereof, comprising: dissolving an anti-gram-positive antibiotic or salt thereof in a solvent to form an antibiotic formulation; and aerosolizing the antibiotic formulation to administer the antibiotic formulation to the lungs of the patient, wherein the aerosolizing is conducted within about 16 hours of the dissolving. 77. A method of forming a powder comprising anti-gram-positive antibiotic or salt thereof, the method comprising: dissolving anti-gram-positive antibiotic or salt thereof in a solvent to form a solution having a concentration ranging from about 60 mg/ml to about 120 mg/ml; and lyophilizing the solution to form the powder. 78. The method of item 77, wherein the solvent comprises water. 87 4435277_2 (GHMatters) P77144.AU.1 79. The method of item 77, wherein the solution is excipient-free. 80. The method of item 77, wherein the solution is cryoprotectant-free. 81. The method of item 77, wherein the powder has a water content of less than about 5 wto. 82. The method of item 77, wherein the lyophilizing is conducted within about 8 hours of the dissolving. 83. The method of item 77, wherein the powder is capable of being reconstituted with water at 25'C and 1.0 atmosphere and with manual agitation, in less than about 60 seconds. 84. A method of forming a powder comprising anti-gram-positive antibiotic or salt thereof, the method comprising. dissolving anti-gram-positive antibiotic or salt thereof in a solvent to form a solution having a volume ranging from about 4.5 ml to about 5.5 ml; and lyophilizing the solution to form the dry powder. 85. The method of item 84, wherein a concentration of the anti-gram-positive antibiotic or salt thereof in the solution ranges from about 60 mg/ml to about 120 mg/ml. 86. The method of item 84, wherein the solution is excipient-free. 87. The method of item 84, wherein the solution is cryoprotectant-free. 88. The method of item 84, wherein the powder has a water content of less than about 5 wto. 89. The method of item 84, wherein the lyophilizing is conducted within about 8 hours of the dissolving. 90. The method of item 84, wherein the powder is capable of being reconstituted with water at 25'C and 1.0 atmosphere and with manual agitation, in less than about 60 seconds. 91. The method of item 84, wherein the lyophilizing comprises filtering the solution, and freezing the filtered solution to form a frozen solution. 88 4435277_2 (GHMatters) P77144.AU.1

Claims (50)

1. An aqueous composition for aerosolization comprising: anti-gram-negative antibiotic or salt thereof being present in an amount from about 100 mg/ml to about 200 mg/ml.

2. The aqueous composition of claim 1, wherein the aqueous composition consists essentially of the anti-gram-negative antibiotic or salt thereof and water.

3. The aqueous composition of claim 1, wherein the amount of the anti-gram-negative antibiotic or salt thereof is from about 110 mg/ml to about 150 mg/ml, or a potency from about 500 p.g/mg to about 1100 p.g/mg, or both.

4. The aqueous composition of claim 1, wherein the anti-gram-negative antibiotic or salt thereof comprises an aminoglycoside or salt thereof

5. The aqueous composition of claim 1, wherein the anti-gram-negative antibiotic or salt thereof comprises at least one member selected from gentamicin, amikacin, kanamycin, streptomycin, neomycin, netilmicin, paramecin, tobramycin, and salts thereof

6. The aqueous composition of claim 1, wherein the anti-gram-negative antibiotic or salt thereof comprises amikacin or salt thereof

7. The aqueous composition of claim 1, wherein the aqueous composition has a pH from about 4 to about 6.

8. The aqueous composition of claim 1, wherein the aqueous composition has an osmolality ranging from about 90 mOsmol/kg to about 500 mOsmol/kg.

9. The aqueous composition of claim 1, wherein the aqueous composition is preservative free.

10. The aqueous composition of claim 1, further comprising an additional active agent.

11. The aqueous composition of claim 1, further comprising an additional active agent selected from an anti-inflammatory and a bronchodilator.

12. The aqueous composition of claim 1, further comprising a bronchodilator selected from -agonist, anti-muscarinic agent, and steroid.

13. The aqueous composition of claim 1, further comprising albuterol.

14. The aqueous composition of claim 1, further comprising an osmolality adjuster.

15. The aqueous composition of claim 1, wherein no precipitate forms in the aqueous composition when the aqueous composition is stored for 1 year at 25'C. 89 4435277_2 (GHMatters) P77144.AU.1

16. The aqueous composition of claim 1, further comprising an anti gram positive antibiotic.

17. The aqueous composition of claim 1, wherein the anti-gram-negative antibiotic or salt thereof comprises gentamicin or amikacin or a salt thereof, wherein the aqueous composition consists essentially of the gentamicin or salt thereof and water, wherein the gentamicin or salt thereof is present in an amount from about 80 mg/ml to about 140 mg/ml, wherein the aqueous composition has a pH from about 3 to about 7, and wherein the aqueous composition has an osmolality from about 120 mOsmol/kg to about 300 mOsmol/kg.

18. An aqueous composition, consisting essentially of an anti-gram-negative antibiotic or salt thereof, a bronchodilator, and water.

19. The aqueous composition of claim 18, wherein the anti-gram-negative antibiotic or salt thereof is present in an amount ranging from about 100 mg/ml to about 200 mg/ml.

20. The aqueous composition of claim 18, wherein the anti-gram-negative antibiotic or salt thereof comprises an aminoglycoside or salt thereof

21. The aqueous composition of claim 18, wherein the aqueous composition has a pH from about 3 to about 7, and an osmolality from about 90 mOsmol/kg to about 500 mOsmol/kg.

22. The aqueous composition of claim 18, wherein the bronchodilator is present in an amount of at least about 1 mg/ml.

23. The aqueous composition of claim 22, wherein the bronchodilator is selected from a p agonist, anti-muscarinic agent, and steroid.

24. The aqueous composition of claim 22, wherein the bronchodilator comprises albuterol or salt thereof

25. A unit dose, comprising: a container; and an aqueous composition comprising an antibiotic or salt thereof, wherein the composition is preservative free.

26. The unit dose of claim 25, wherein the antibiotic comprises an anti gram-negative, and is present at a concentration from about 90 mg/ml to about 200 mg/ml.

27. The unit dose of claim 26, wherein the antibiotic comprises amikacin.

28. A kit, comprising: a first container containing a first aqueous solution comprising a first antibiotic or salt 90 4435277_2 (GHMatters) P77144.AU.1 thereof; and a second container containing a second aqueous solution comprising a second antibiotic or salt thereof, wherein the first and second antibiotics are the same or different, and wherein a concentration, or an amount, or both of the first antibiotic is the same as, or different from a concentration, or an amount or both of the second antibiotic.

29. The kit of claim 28, wherein an amount of the first aqueous solution is from about 2 ml to about 5 ml, an amount of the second aqueous solution is from about 5 ml to about 8 ml, a concentration of first antibiotic or salt thereof is from about 100 mg/ml to about 120 mg/ml, and a concentration of second antibiotic or salt thereof is from about 120 mg/ml to about 140 mg/ml.

30. The kit of claim 28, wherein the anti-gram-negative antibiotic or salt thereof comprises aminoglycoside or salt thereof

31. The kit of claim 30, wherein the anti-gram-negative antibiotic or salt thereof comprises amikacin or salt thereof

32. The kit of claim 28, further comprising an aerosol introducer.

33. The kit of claim 28, further comprising a vibrating mesh nebulizer.

34. A kit, comprising: a first container containing a first aqueous solution comprising anti-gram-negative antibiotic or salt thereof; and a second container containing a second aqueous solution comprising anti-gram negative antibiotic or salt thereof, wherein a concentration, an amount, or both, of the anti-gram-negative antibiotic or salt thereof of the first aqueous solution is different from a concentration, or an amount, or both, of the anti-gram-negative antibiotic of the second aqueous solution.

35. The kit of claim 34, further comprising an aerosol introducer.

36. The kit of claim 35, further comprising a vibrating mesh nebulizer.

37. A kit, comprising: a first container containing a first antibiotic or salt thereof; and a second container containing a second antibiotic or salt thereof, wherein the first and second antibiotics are the same or different, and wherein a 91 4435277_2 (GHMatters) P77144.AU.1 concentration, or an amount, or both of the first antibiotic in the first container is different from a concentration, or an amount, or both, of the second antibiotic or salt thereof in the second container.

38. The kit of claim 37, wherein each of the first and second antibiotics comprise a liquid.

39. The kit of claim 37, wherein one of the first or second antibiotics comprises a powder, and the other of the first or second antibiotics comprises a liquid.

40. The kit of claim 37, wherein the solution if preservative free.

41. The kit of claim 37, further comprising an aerosol introducer.

42. The kit of claim 41, further comprising a vibrating mesh nebulizer.

43. A method of administering an antibiotic formulation to a patient in need thereof, comprising: aerosolizing an antibiotic formulation to administer the antibiotic formulation to the pulmomary system of the patient, wherein the antibiotic formulation has a concentration of anti-gram-negative antibiotic or salt thereof ranging from about 100 mg/ml to about 200 mg/ml.

44. The method of claim 43, wherein the antibiotic formulation consists essentially of anti gram-negative antibiotic and water.

45. The method of claim 43, wherein the aerosolizing comprises forming droplets having a mass median aerodynamic diameter of less than about 10 gim.

46. The method of claim 43, wherein the aerosolizing comprises aerosolizing the antibiotic formulation in a vibrating mesh nebulizer.

47. The method of claim 35, wherein the pulmonary administration comprises prophylactic treatment of ventilator associated pneumonia.

48. A method of administering an antibiotic formulation to a patient in need thereof, comprising: inserting a tube into a trachea of a patient; and aerosolizing an antibiotic formulation to administer the antibiotic formulation to the lungs of the patient, wherein the antibiotic formulation consists essentially of anti-gram-negative antibiotic or salt thereof and water.

49. The method of claim 48, wherein the pulmonary administration comprises 92 4435277_2 (GHMatters) P77144.AU.1 prophylactic treatment of ventilator associated pneumonia, hospital associated pneumonia, community acquired pneumonia, or combinations thereof

50. The method of claim 48, wherein the aerosolizing comprises aerosolizing the antibiotic formulation in a vibrating mesh nebulizer. 93 4435277_2 (GHMatters) P77144.AU.1