Subcutaneous formulations and methods for treating edema refractory to oral diuretics - Patent Application 20070229933

Abstract

The present invention features methods and compositions for the subcutaneous administration of bumetanide to treat subjects suffering from edema refractory to oral diuretics.

Description

[Technical field]

[0001] The present disclosure features methods and compositions for subcutaneous injection to treat edema refractory to oral diuretics. [Background technology]

[0002] Congestive heart failure (CHF) is a common cardiac disease. The incidence of congestive heart failure has increased in recent years, and its diagnosis is associated with significant morbidity and mortality. In fact, congestive heart failure is a highly fatal disease with an estimated 5-year mortality in the majority of affected individuals, both men and women. Congestive heart failure results from the loss or impairment of normal cardiac function. This loss or impairment reduces cardiac output. This results in a reduction in both blood flow and blood pressure in the kidneys. This reduction in blood flow and blood pressure causes a renin-angiotensin reaction that exacerbates congestive heart failure. Blood volume increases as angiotensin II stimulates the secretion of aldosterone from the adrenal cortex, which in turn causes increased salt and water retention in the kidneys. The increased blood volume and corresponding vasoconstriction cause an increase in blood pressure and thus fluid overload of the heart, further worsening the cardiac condition.

[0003] To treat CHF, doctors place patients on a strict low-sodium diet and monitor their fluid intake. Some patients are restricted to only one liter of fluid per day. The most important drugs doctors use to combat fluid overload are a class of drugs called diuretics. Diuretics affect kidney function in such a way that fluid reabsorption is inhibited. As a result, urine output increases, contrary to the neurohormonal cues the kidneys receive. Doctors may treat patients with drugs that improve the heart's pumping ability, increase blood pressure, and attempt to reactivate more normal behavior of the body's control system. This is generally effective in keeping many heart failure patients alive. However, in hundreds of thousands of patients, treatment with drugs and diet alone is unsuccessful. When patients are edematous and experiencing fluid overload, gastrointestinal absorption is impaired and thus oral diuretics may be of limited effectiveness. As a result, patients are often instructed to increase the dosage of oral diuretics, further straining the patient's kidneys. Eventually, oral diuretics become insufficient to remove excess fluid, and patients seek intravenous diuretics to avoid absorption by the gastrointestinal system. As a result, patients are often hospitalized for intensive care and iv diuretics, and are at risk of excessive diuresis (in each event) when gastrointestinal absorption is restored. Ultimately, excessive diuresis can lead to renal failure. Renal replacement therapy, such as hemofiltration or dialysis, is increasingly used as a method of removing fluid in acute CHF conditions when available treatments no longer provide sufficient fluid removal through existing renal function. Acute heart failure may be treated with continuous renal replacement therapy (also known as an artificial kidney or dialysis machine) in the hospital's ICU.

[0004] Thus, there remains a need for treatment options for heart failure patients in acute distress from fluid overload that reduce the risk of hospitalization and renal failure. Summary of the Invention

[0005] The present invention features subcutaneous compositions and methods for treating edema refractory to oral diuretics.

[0006] In a first aspect, the invention features a method of treating edema in a subject, the method comprising subcutaneously administering an effective amount of a pharmaceutical composition that includes: (i) an aqueous solution having a pH of about 5 to about 9 (e.g., a pH of 5±1, 6±1, 7±1, 8±1, or 9±1); (ii) about 4 mg / mL to about 20 mg / mL (e.g., 4±1, 5±1, 6±1, 7±1, 8±1, 9±1, 10±1, 11±1, 12±1, 13±1, 14±1, 15±1, 16±1, 17±1, 18±1, 19±1, or 20±1 mg / mL) of bumetanide arginine salt; and (iii) one or more pharma- ceutically acceptable excipients. In certain embodiments, the pharmaceutical composition comprises: (i) an aqueous solution having from about 5 mg / mL to about 12 mg / mL of bumetanide arginine salt; (ii) one or more pharma- ceutically acceptable excipients; and (iii) the aqueous solution has a pH of from about 6 to about 8.

[0007] In some embodiments, the pharma- ceutically acceptable excipient comprises a tonicity agent (e.g., any of the tonicity agents described herein) or a buffer (e.g., a citrate buffer, a phosphate buffer, or any other buffer described herein). In certain embodiments, the aqueous solution does not contain a buffer other than the buffer formed by bumetanide free acid combined with arginine.

[0008] In a first aspect, the invention features a method of treating edema in a subject, the method including subcutaneously administering an effective amount of a pharmaceutical composition including: (i) an aqueous solution having a pH of about 5 to about 9 (e.g., a pH of 5±1, 6±1, 7±1, 8±1, or 9±1); (ii) about 4 mg / mL to about 20 mg / mL (e.g., 4±1, 5±1, 6±1, 7±1, 8±1, 9±1, 10±1, 11±1, 12±1, 13±1, 14±1, 15±1, 16±1, 17±1, 18±1, 19±1, or 20±1 mg / mL) of bumetanide potassium salt; and (iii) one or more pharma- ceutically acceptable excipients. In certain embodiments, the pharmaceutical composition comprises: (i) an aqueous solution having from about 5 mg / mL to about 12 mg / mL of bumetanide potassium salt; (ii) one or more pharma- ceutically acceptable excipients; and (iii) the aqueous solution has a pH of from about 6 to about 8.

[0009] In some embodiments, the pharma- ceutically acceptable excipient comprises a tonicity agent (e.g., any of the tonicity agents described herein) or a buffer (e.g., a citrate buffer, a phosphate buffer, or any other buffer described herein). In certain embodiments, the aqueous solution does not contain a buffer other than the buffer formed by bumetanide free acid combined with potassium hydroxide.

[0010] In some embodiments, the aqueous solution has a pH of about 6 to about 8. In some embodiments, the one or more pharma- ceutically acceptable excipients include a sugar or sugar alcohol (e.g., sucrose and / or mannitol). In some embodiments, the pharma-ceutically acceptable excipients include a preservative (e.g., benzyl alcohol).

[0011] The invention further features a method of treating edema in a subject in need thereof, comprising administering to the subject an effective amount of a pharmaceutical composition of the invention. In some embodiments, the pharmaceutical composition is administered in a dosage volume of 0.5 mL or less. The administration can include delivering a dosage volume of 25 μl to 500 μl (e.g., 35±10, 50±10, 75±25, 125±25, 150±25, 200±50 μl, 300±100 μl, or 400±100 μl) of the pharmaceutical composition subcutaneously to the subject. In certain embodiments, the doses are delivered no more than 1-4 times over a 6 hour period. In certain embodiments, 4 doses of about 400 μL each are delivered over a 1 hour period. In certain embodiments, 2 doses of about 400 μL each are delivered to the subject, followed by 2 more doses of about 400 μL each about 30-60 minutes later. In other embodiments, the subject suffers from edema refractory to oral diuretics. In certain embodiments, the subject has congestive heart failure. In certain embodiments, the subject suffers from pulmonary edema (e.g., pulmonary edema). In some embodiments, the pharmaceutical composition is administered subcutaneously to the subject in an outpatient setting. In certain embodiments, the pharmaceutical composition is self-administered.

[0012] In one embodiment of any of the above methods, the pharmaceutical composition has a bioavailability of at least 90% (e.g., 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100%) or 95% (e.g., 96%, 97%, 98%, 99%, or 100%) when administered subcutaneously to a subject.

[0013] In certain embodiments, the pharmaceutical composition comprises a buffer selected from a citrate buffer and a phosphate buffer.

[0014] In some embodiments, the pharmaceutical composition is administered subcutaneously to the subject in an outpatient setting or the pharmaceutical composition is self-administered. In certain embodiments, the subject is experiencing reduced intestinal motility prior to administration.

[0015] This method may be particularly beneficial when the subject has failed to achieve diuresis with oral diuretic therapy before subcutaneous administration.The subject to be treated may experience leg swelling, shortness of breath, difficulty in breathing, or chest pain that does not resolve with oral diuretic therapy before subcutaneous administration.In some embodiments, the subject experiences reduced intestinal motility before subcutaneous administration.

[0016] In a related aspect, the present invention provides a method of treating edema refractory to oral diuretics in a subject with congestive heart failure, comprising administering to the subject an effective amount of a pharmaceutical composition comprising any one of the pharmaceutical compositions described herein.

[0017] In one embodiment of any of the above methods, the subject has been treated with at least one dose of an oral diuretic within the last 24 hours prior to administration. The at least one oral diuretic may be selected from a loop diuretic, such as bumetanide, furosemide, or torsemide, or a potassium-sparing diuretic, such as amiloride or spironolactone.

[0018] In some embodiments of any of the above methods, the patient does not receive more than about 10 mg total of bumetanide salt in a 12 hour period. In some embodiments, a patient receiving more than 10 mg total of bumetanide salt in a 12 hour period consults a physician.

[0019] In some embodiments of any of the above methods, the subject's risk of hospitalization due to complications associated with edema is reduced.

[0020] In one aspect of any of the above methods, the pharmaceutical composition is administered in 1, 2, 3, or 4 doses over a 12 hour period. In some embodiments, the pharmaceutical composition is administered in a single dose. In some embodiments, a bumetanide salt is administered in 1, 2, 3, or 4 doses of 1-2.5 mg over a 12 hour period. In some embodiments, a bumetanide salt is administered in 1, 2, 3, or 4 doses of 2-5 mg over a 12 hour period. In some embodiments, a bumetanide salt is administered in 1, 2, 3, or 4 doses of 3-7.5 mg over a 12 hour period. In some embodiments, a bumetanide salt is administered in 1, 2, 3, or 4 doses of 4-10 mg over a 12 hour period.

[0021] In a related aspect, the invention features a pharmaceutical composition that includes (i) an aqueous solution having a pH of about 6 to about 8 (e.g., a pH of 6±1, 6.5±1, or 7±1); (ii) a bumetanide arginine salt of about 5 mg / mL to about 12 mg / mL (e.g., 6±1, 7±1, 8±1, 9±1, 10±1, or 11±1 mg / mL); and (iii) a pharmaceutical excipient that includes a sugar or sugar alcohol. In certain embodiments, the bumetanide arginine has a concentration of about 8 mg / mL to about 10 mg / mL. In some embodiments, the aqueous solution has a pH of about 6.5 to about 7.5. In certain embodiments, the pharmaceutical composition includes benzyl alcohol (e.g., about 0.2% (w / w) to about 1% (w / w) benzyl alcohol). In certain embodiments, the pharmaceutical composition includes about 0.5% (w / w) benzyl alcohol.

[0022] In a related aspect, the invention features a pharmaceutical composition that includes (i) an aqueous solution having a pH of about 6 to about 8 (e.g., a pH of 6±1, 6.5±1, or 7±1); (ii) about 5 mg / mL to about 12 mg / mL (e.g., 6±1, 7±1, 8±1, 9±1, 10±1, or 11±1 mg / mL) of bumetanide potassium salt; and (iii) a pharmaceutical excipient that includes a sugar or sugar alcohol. In certain embodiments, the bumetanide potassium has a concentration of about 8 mg / mL to about 10 mg / mL. In some embodiments, the aqueous solution has a pH of about 6.5 to about 7.5. In certain embodiments, the pharmaceutical composition includes benzyl alcohol (e.g., about 0.2% (w / w) to about 1% (w / w) benzyl alcohol). In certain embodiments, the pharmaceutical composition includes about 0.5% (w / w) benzyl alcohol.

[0023] In one embodiment of any of the above methods, the pharmaceutical composition comprises mannitol (e.g., about 2.5% (w / w) to about 5% (w / w) mannitol, or about 3±0.5%, 3.5±0.5%, 4±0.5%, or 4.5%±0.5% (w / w) mannitol).

[0024] In one embodiment of any of the above methods, the pharmaceutical composition comprises sucrose (e.g., 1% (w / w) to about 3% (w / w) sucrose, or about 1.5±0.5%, 2.0±0.5%, or 2.5%±0.5% (w / w) sucrose). [Brief description of the drawings]

[0025] [Figure 1] 1 is a graph showing the solubility (mg / mL) of various bumetanide salt forms compared to the weight (mg) of excipients added per 20 mg of bumetanide. [Diagram 2]FIG. 13 shows the concentration of bumetanide (ng / mL) in dog serum over 0-240 minutes following IV or SC administration to compare the concentration of bumetanide achieved in serum using IV administration of 1.0 mg of commercially available bumetanide to the concentration of bumetanide achieved by SC administration of 1.01 mg of the FS-3 and FS-4 bumetanide formulations, 0.97 mg of the FS-5 bumetanide formulation, or 1.0 mg of the FS-6 bumetanide formulation. [Diagram 3] FIG. 13 is a graph showing the concentration of bumetanide (ng / mL) in dog serum over 0-60 minutes following IV or SC administration to compare the concentration of bumetanide achieved in serum using IV administration of 1.0 mg of commercially available bumetanide to the concentration of bumetanide achieved by SC administration of 1.01 mg of FS-3 and FS-4 bumetanide formulations, 0.97 mg of FS-5 bumetanide formulation, or 1.0 mg of FS-6 bumetanide formulation. [Figure 4] FIG. 1 shows the area under the curve (AUC) of the concentration of bumetanide (ng*h / mL) in dog plasma over 2 to 240 minutes following IV administration of approximately 1.0 commercial bumetanide, subcutaneous administration of approximately 1.01 mg FS-4, approximately 1.01 mg FS-4 formulation, approximately 0.97 mg FS-5 formulation, approximately 1.0 mg FS-6 formulation, and intranasal administration of approximately 1.0 mg F82 formulation. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0026] definition As used herein, the term "about" refers to values ​​that are within 10% above or below the stated value. For example, the expression "about 6 mg / mL" refers to a value between 5.4 and 6.6 mg / mL.

[0027] As used herein, the term "effective amount" refers to an amount of a pharmaceutical composition that is sufficient to produce a beneficial or desired result, such as a clinical result, when administered to a subject, e.g., a human subject. For example, in the context of edema as described herein, this term refers to an amount of a composition that is sufficient to achieve a reduction in symptoms of edema when compared to the response obtained without administration of the composition. The amount of a given composition described herein that corresponds to such an amount depends on various factors, such as a given drug, pharmaceutical formulation, route of administration, identity of the subject being treated (e.g., age, sex, weight), etc.

[0028] As used herein, "emulsion" refers to a two-phase colloidal system, such as a mixture of two or more immiscible liquids, that may be added as an excipient to a pharmaceutical composition. A liquid emulsion is one in which both the dispersed and continuous phases are liquid. To form an emulsion, energy input is usually required by shaking, stirring, homogenization, or spraying processes. For example, an emulsion may include an aqueous phase and a non-aqueous phase, may include a self-emulsifying system, or may be a nanoparticulate (e.g., nanoemulsion or microemulsion) that includes an aqueous phase and a non-aqueous phase. "Nanoemulsion or microemulsion" refers to a clear, stable, isotropic liquid mixture of oil, water, and surfactant, optionally in combination with a cosurfactant. The aqueous phase may include salt(s) and / or other ingredients in addition to the biologically active agent. Unlike regular emulsions, microemulsions are formed by simple mixing of the ingredients and do not require the high shear conditions typically used in the formation of regular emulsions. The two basic types of microemulsions are normal phase (oil dispersed in water, o / w) and inverse phase (water dispersed in oil, w / o).

[0029] As used herein, the term "pulmonary edema" or "pulmonary edema" refers to a condition in which a patient has excess fluid in the lungs, resulting in difficulty breathing. Pulmonary edema can be caused by conditions related to heart failure, pneumonia, trauma, an allergic reaction, or another cause.

[0030] As used herein, the term "oral diuretic-refractory edema" refers to edema that does not respond to oral diuretic treatment such that diuresis is not achieved and excess fluid remains, resulting in a persistent edematous state.

[0031] As used herein, the term "failure to achieve diuresis" refers to the failure of administration of oral diuretics to increase fluid excretion from a patient, resulting in a persistent edematous state.

[0032] As used herein, the term "loop diuretic" refers to drugs used to reduce the symptoms of hypertension and edema in patients with congestive heart failure or renal insufficiency. Loop diuretics belong to a class of diuretics that decrease the reabsorption of sodium and chloride by the kidneys, thereby increasing urine production.

[0033] As used herein, the term "low viscosity sodium carboxymethylcellulose" or "low viscosity sodium CMC" refers to sodium carboxymethylcellulose having a viscosity of 30 cP to 45 cP (e.g., 30±1 cP, 31±1 cP, 32±1 cP, 33±1 cP, 34±1 cP, 35±1 cP, 36±1 cP, 37±1 cP, 38±1 cP, 39±1 cP, 40±1 cP, 41±1 cP, 42±1 cP, 43±1 cP, 44±1 cP, and 45±1 cP) in a 2% aqueous solution at 25° C., or 50 to 200 cP in a 4% aqueous solution at 25° C. The low viscosity sodium carboxymethylcellulose may have a molecular weight of approximately 90 kDa.

[0034] As used herein, the term "pharmacologically acceptable" refers to compounds, substances, compositions, and / or dosage forms that are suitable for contact with the tissues of a subject, such as a mammal (e.g., a human), without undue toxicity, irritation, allergic response, and other problematic adverse effects, commensurate with a reasonable benefit / risk ratio.

[0035] As used herein, the term "potassium-sparing diuretics" refers to a class of diuretics that do not promote the secretion of potassium into the urine, thus increasing fluid excretion. These diuretics may be used alone or in combination with loop or thiazide diuretics.

[0036] As used herein, the term "reduced intestinal motility" refers to the slowing down of the activity of the gastrointestinal tract of a subject.One of the effects of slowing down of activity can be reduced absorption, thus preventing effective absorption of pharmaceutical compositions.For example, this reduced intestinal motility can be caused by edema (such as fluid overload resulting from congestive heart failure), which prevents sufficient absorption of oral diuretics required for treating edema.

[0037] As used herein, the term "hospitalization risk" refers to the potential likelihood that a patient will be hospitalized for the treatment of edema, rather than effectively treating edema without hospitalization by self-administering bumetanide using the methods of the present invention. Reduction in hospitalization risk is assessed by comparing the rate at which patients who self-administer bumetanide are hospitalized for the treatment of edema with patients who rely solely on oral diuretics for the treatment of edema for a given disease population of a particular severity (e.g., patients suffering from congestive heart failure). Using the bumetanide methods of the present invention, the rate of hospitalization for the treatment of edema in a patient population can be reduced by at least 10%, 20%, 30%, or 50%, thus reducing the risk of hospitalization for individual patients who use bumetanide.

[0038] As used herein, "treatment" and "treating" refer to therapy for a subject in need of diuresis, such as therapy to ameliorate one or more symptoms of edema in a subject suffering from edema or prophylactically reduce the risk of one or more symptoms of edema in a subject at risk for edema.

[0039] As used herein, the term "unit dose" or "dosage" when used in reference to a therapeutic composition refers to physically discrete units suitable as unitary administration for subjects, each unit containing a predetermined quantity of active material calculated to produce a desired therapeutic effect, in association with the required diluent, i.e., carrier, or vehicle.

[0040] Detailed Description The present invention features pharmaceutical compositions and methods for treating edema refractory to oral diuretics. Bumetanide can be formulated for subcutaneous delivery in a salt form that allows a therapeutically effective amount of bumetanide to be delivered in a small volume (approximately 100-150 μL) that is particularly suitable for subcutaneous delivery. The concentration of bumetanide is optimally 5 mg / mL or 10 mg / mL so that an ideal dosage can be realized in this small volume. To ensure suitable shelf-life stability of a pharmaceutical product, changes in solubility as a result of changes in temperature and storage conditions must be considered, so that the concentration of bumetanide salt in solution approaches its saturation limit. This further highlights the need addressed by the present invention for a salt form capable of stability of at least 5 mg / mL.

[0041] Using these bumetanide salt concentrations, subcutaneous administration of bumetanide to patients experiencing edema refractory to oral diuretics due to edematous conditions (e.g., occurrences seen in patients with reduced bowel motility associated with edema) can alleviate the edema and restore the effectiveness of orally administered diuretics without the need for hospitalization. Subjects may self-administer the pharmaceutical composition based on their symptoms, thus obviating the need for hospitalization, or the pharmaceutical composition may be administered by a medical professional (e.g., a physician, paramedic, or nurse) when the subject is experiencing acute distress.

[0042] Furthermore, the method of the present invention can reduce the risk of renal failure in certain patients, such as those suffering from congestive heart failure. When such patients are in an edematous state and experience fluid overload, gastrointestinal absorption is impaired, and thus the effectiveness of oral diuretics may be limited. As a result, patients are often instructed to increase the dosage of oral diuretics, which further burdens the kidneys of the patients. Eventually, oral diuretics become insufficient to remove excess fluid, and the patients are hospitalized to bypass the gastrointestinal system and receive intravenous diuretics. As a result, the patients are at risk of excessive diuresis when gastrointestinal absorption is restored. This excessive diuresis may lead to renal failure.

[0043] Pharmaceutical Compositions The bumetanide formulation of the present invention may be a solution or a suspension. The formulation may include antioxidants, pH adjusters (e.g., acids or bases), buffers, preservatives, tonicity agents, and / or viscosity enhancers (e.g., carboxymethylcellulose). The formulation may be administered as an aqueous solution or in the form of an emulsion, including nanoemulsions and microemulsions. The formulation may be provided in a single dose or multidose form. The formulation may be administered, for example, subcutaneously, via a preloaded syringe.

[0044] Viscosity improver Viscosity enhancer may be used in the pharmaceutical composition of the present invention that is formulated for subcutaneous administration.The viscosity enhancer that may be used according to the present invention includes, but is not limited to, cellulose derivatives, Carbopol, gum, and hyaluronic acid (hyaluronate), dextran, polyvinyl alcohol, polyacrylic acid, povidone, polyethylene glycol, polyol (e.g., glycerol), propylene glycol, and chitosan, and for cellulose derivatives, particularly preferred are one or more of carboxymethylcellulose ("CMC") high molecular weight blend, CMC low molecular weight blend, CMC medium molecular weight blend, CMC sodium (low viscosity), methylcellulose, methylcellulose 4000, hydroxymethylcellulose, hydroxypropylcellulose ("HPC"), hydroxypropylmethylcellulose high molecular weight blend ("HPMC"), hydroxypropylmethylcellulose 2906, carboxypropylmethylcellulose high molecular weight blend ("CPMC"), hydroxyethylcellulose, or hydroxyethylcellulose, and hyaluronic acid. In certain embodiments, the viscosity enhancing agent is sodium CMC, preferably in combination with a polyol selected from the group consisting of mannitol, xylitol, sorbitol, isosorbide, erythritol, glycerol, maltitol, and combinations thereof.

[0045] Isotonic agent In the pharmaceutical composition of the present invention, a tonicity agent may be used to adjust the tonicity of the liquid pharmaceutical composition. Tonicity generally relates to the osmotic pressure of a solution, and is typically evaluated relative to that of human serum. A pharmaceutical composition (e.g., pharmaceutical dosage form) may contain a tonicity agent to increase osmolality. Non-limiting examples of tonicity agents include substantially neutral buffers (e.g., phosphate buffered saline, Tris buffer, or artificial perilymph), dextrose, mannitol, trehalose, sucrose, sorbitol, glycerin (also known as glycerol), potassium chloride, and sodium chloride (e.g., as hypertonic, isotonic, or hypotonic saline), and amino acids (e.g., arginine, glycine, ornithine, lysine, histidine, glutamic acid, aspartic acid, isoleucine, leucine, alanine, phenylalanine, tyrosine, tryptophan, methionine, serine, and proline). A pharmaceutical composition (e.g., pharmaceutical dosage form) includes a sufficient amount of a tonicity agent to administer a hypertonic pharmaceutical dosage form to a subject.

[0046] Buffer In some embodiments, the pharmaceutical composition includes a buffering agent. A buffering agent may be included in the pharmaceutical composition to increase the stability of the composition by maintaining a certain pH range. The buffering agent may improve the ability of the composition to be absorbed by the subject. Additionally, the buffering agent may act to increase the solubility of the composition. The buffering agent may increase the solubility of the drug by maintaining high solubility of the drug at a certain pH range, preferably pH 6-8. In certain embodiments, the buffering agent maintains a pH in the range of 6.5-6.7. Buffering agents that may be used in accordance with the present invention include, but are not limited to, potassium hydroxide, arginine, and lysine. One or more components of the pharmaceutical composition (e.g., potassium hydroxide, arginine, and lysine) may act as counterion species of the buffer, and bumetanide also functions as a buffering agent. The buffering agent may be citric acid, malic acid, succinic acid, potassium chloride, or ethylenediaminetetraacetic acid. For example, one or more buffering agents may be included in the pharmaceutical composition in an amount of about 5 mM to about 10 mM (e.g., 5±0.5 mM, 5.5±0.5 mM, 6±0.5 mM, 6.5±0.5 mM, 7±0.5 mM, 7.5±0.5 mM, 8±0.5 mM, 8.5±0.5 mM, 9±0.5 mM, 9.5±0.5 mM, or 10±0.5 mM). Buffering agents may be added to the pharmaceutical composition depending on whether the method of administration requires maintenance of a particular pH range.

[0047] Preservatives In some embodiments of the pharmaceutical composition, a preservative is included as part of the pharmaceutical composition to increase the stability and / or shelf life of the pharmaceutical composition. The preservative may be in the form of, for example, an antioxidant, an antimicrobial, or a chelating agent. An antioxidant may be added to the pharmaceutical composition to prevent oxidation of other components in the composition that may be oxidation sensitive in the presence of oxygen or sunlight. An antimicrobial agent may be included in the pharmaceutical composition to inhibit contamination of the pharmaceutical composition with microorganisms. A chelating agent may be added to the pharmaceutical composition to bind and protect the pharma- ceutical active ingredient from degradation and increase stability. Preservatives that may be used in accordance with the present invention include, but are not limited to, benzyl alcohol, benzoic acid, and ethylenediaminetetraacetic acid (EDTA).

[0048] Medication Regimen The dosing regimen used in the treatment methods described herein may vary depending on many factors, such as the age, health, and weight of the recipient, the nature and severity of the edema, and the frequency and type of concomitant treatment, if any. One of skill in the art can determine the appropriate dosage based on the above factors. The compounds used in the methods described herein may be administered initially at a suitable dosage, which may be adjusted, if necessary, depending on the individual response. In general, a suitable dosage of bumetanide according to the present invention is within the range of 0.5-10 mg of bumetanide (e.g., 0.5, 1.0, 2.0, and 5 mg) in 1-4 doses over a period of 1-4 hours until the patient is no longer edematous. Four doses of the pharmaceutical composition at 100 μL per dose may be delivered to the subject in less than an hour. Alternatively, two doses at about 100 μL per dose may be delivered to the subject, followed by two more doses of about 100 μL each about 30-60 minutes later. Bumetanide may be administered to the patient upon the onset of symptoms of edema. Alternatively, bumetanide is administered to a patient after a previously administered oral diuretic has failed to treat the edema and symptoms persist. If administration of an oral diuretic has been ineffective, the patient may be restricted to an initial dose of less than 5 mg bumetanide for at least 1 or 2 hours to further reduce the risk of excessive diuresis, which may manifest as symptoms of dehydration and hypotension in subjects suffering from excessive diuresis.

[0049] For sustained release formulations administered as subcutaneous depots, suitable doses of bumetanide according to the invention are in the range of 0.75-10 mg bumetanide (e.g., 1.5±0.5, 2.5±0.5, 3.5±0.5, 4.5±0.5, 5.5±0.5, 6.5±0.5, 7.5±0.5, 8.5±0.5, and 9.5±0.5 mg) in 1-2 doses over 1-5 days. Subcutaneous depots of bumetanide may be administered to patients to prophylactically treat symptoms of edema and reduce the risk of rehospitalization.

[0050] Oral diuretics The present disclosure features methods and compositions for treating edema refractory to oral diuretics. Oral diuretics have long been used to relieve fluid retention, a hallmark of congestive heart failure. Aggressive use of diuretics can reduce hospitalizations and improve exercise capacity, even in people using ACE inhibitors. Diuretics act on the kidneys to remove excess salt and water from the body, reducing fluid accumulation in the legs, abdomen, and lungs, lowering blood pressure, and improving the efficiency of circulation. Side effects of diuretics include low blood pressure, dehydration, and kidney dysfunction. They can also cause gout, increase blood glucose and triglycerides, LDL, and overall cholesterol levels, and deplete the B vitamin thiamine. Many diuretics are available, which are generally classified as thiazide and loop diuretics and are used with or without potassium-sparing agents. It is important to note that recent studies have found an increased incidence of hospitalization in patients who were taking nonsteroidal anti-inflammatory drugs (NTHEs) in combination with diuretics. Common NSAIDS include aspirin, ibuprofen, and naproxen. Thiazides such as hydrochlorothiazide (HydroDIURIL, Esidrix), chlorothiazide (Diuril), metolazone (Zaroxolyn), and chlorthalidone (Hygroton) are typically prescribed for patients with mild heart failure and good kidney function.

[0051] Loop diuretics, such as furosemide (Lasix), bumetanide (BumeX), and ethacrynic acid (Edecrine), are commonly used for more severe heart failure, especially when kidney function is impaired. Loop diuretics are used intravenously to treat pulmonary edema and acute congestive heart failure, and thiazides and loop diuretics may be administered simultaneously. Even after standard treatment of congestive failure, fluid may remain in the lungs, limiting the patient's ability to function normally. One study treated patients with this condition very aggressively with furosemide to further reduce fluid, but no improvement was observed. Alternative methods using filtration techniques worked better.

[0052] Potassium loss is a major problem with diuretic use. Doctors may recommend potassium supplements or the use of potassium-sparing diuretics such as spironolactone (Aldactone), amiloride (Midamor), and triamterene (Dyrenium) in combination with a thiazide or loop diuretic unless the patient is also taking an ACE inhibitor that raises potassium levels. All patients receiving diuretics, with or without a potassium-sparing drug, should have their blood potassium levels tested periodically.

[0053] The method of the present invention may include the bumetanide therapy described herein, which is used to manage the patient's edema in combination with an oral diuretic, such as a loop diuretic, a potassium-sparing diuretic, a thiazide, or another oral diuretic. Typically, subcutaneously administered bumetanide is administered to a patient who has previously taken an oral diuretic, for example within the past hour, two hours, or four hours, for edema that is known to be refractory to oral diuretics. For example, the patient may be taking oral diuretics such as potent loop diuretics (e.g., furosemide, ethacrynic acid, torsemide, and bumetanide), thiazides (e.g., hydrochlorothiazide acid), carbonic anhydrase inhibitors (e.g., acetazolamide and methazolamide), potassium-sparing diuretics (e.g., the aldosterone antagonist spironolactone, and the epithelial sodium channel blockers amiloride and triamterene), and / or calcium-sparing diuretics. After a patient has been successfully treated for an edematous condition using the methods of the present invention, the patient may continue on their usual regimen of oral diuretics.

[0054] Selecting a target The methods and compositions of the present invention may be used in patients who are generally at risk for edema. The edema may be, for example, paw edema, peripheral edema, pulmonary edema (e.g., pulmonary edema). The methods and compositions of the present invention may be used when the subject is a mammal. In particular, the subject may be a dog, if the dog is in need of veterinary care. Furthermore, the subject may be a human. The subjects that may be treated using the methods described herein are those with a diagnosis of congestive heart failure. Congestive heart failure (CHF) is characterized by the inability of the heart to generate sufficient cardiac output to meet the body's demands. Patients with CHF experience signs and symptoms of intravascular and interstitial volume overload, such as shortness of breath, tachycardia, fluid accumulation in the lungs, and edema, along with indicators of inadequate tissue perfusion, such as fatigue and / or poor exercise tolerance. Subjects who may be treated include those with heart failure diagnosed by standard and routine diagnostic procedures known in the art (e.g., electrocardiography (ejection fraction), radionuclide imaging, magnetic resonance imaging, computed tomography imaging, cardiac catheterization and angiography, myocardial biopsy, and / or evaluation of blood levels of atrial natriuretic peptide (ANP) and / or B-type natriuretic peptide (BNP)).

[0055] Subjects who may be treated are currently experiencing symptoms known to be associated with congestive heart failure, such as shortness of breath (e.g., respiratory distress), fatigue, weakness, peripheral edema, or fluid overload. The subject may be taking a daily dose of an oral diuretic, such as a loop diuretic, a potassium-sparing diuretic, or a thiazide, to reduce fluid overload and edema along with any one of the symptoms associated with congestive heart failure.

[0056] The subject that can be treated using the methods described herein is experiencing symptoms of renal insufficiency caused by reduced blood flow, such as reduced urine output, swelling of the legs, ankles, feet, or abdomen, shortness of breath, or fatigue, which are caused by fluid overload as a result of renal insufficiency.The subject may experience fluid overload to a level where intestinal motility is impaired, preventing normal absorption of ingested nutrients, leading to insufficient gastrointestinal absorption and contributing to reduced bioavailability of orally administered pharmaceutical compositions.

[0057] Subjects who may be treated using the methods described herein may experience symptoms of refractory edema as a result of congestive heart failure, as well as reduced intestinal motility that prevents the subject's oral diuretics from effectively causing diuresis to remove excess fluid. As a result of persistent symptoms of edema, such as fatigue, shortness of breath, and swelling of the limbs and abdomen, the subject may seek hospitalization to receive intravenous diuretics, thus placing the subject at risk for renal failure.

[0058] Subcutaneous administration of sustained release depot formulations can be used to prophylactically treat patients and allow them to receive reliable diuretic benefits for several days without complications. Sustained release therapy can be used to treat patients at risk of re-hospitalization. For example, after hospitalization for treatment of congestive heart failure, patients can be treated after discharge to achieve diuresis for an extended period (e.g., several days) and relieve symptoms such as shortness of breath, fatigue, and edema. EXAMPLES

[0059] The following examples are presented to provide one of ordinary skill in the art with an illustration of how the compositions and methods described herein can be used, made, and evaluated, and are intended to be purely illustrative of the disclosure and are not intended to limit the scope of what the inventors regard as their disclosure.

[0060] Example 1. Aqueous solubility of bumetanide in the presence and absence of surfactants 10 mg of bumetanide in free acid form was added to 1 mL of deionized water and mixed using a vortex and sonicator. The solution was filtered using a 0.22 μm nylon syringe filter. 100 μL of the filtered bumetanide solution was added to 900 μL of deionized water, and then the concentration of bumetanide in solution was analyzed using HPLC. A standard curve generated by bumetanide solutions of known concentrations was used to determine the concentration of bumetanide. Most of the free acid form of bumetanide was not water soluble. Therefore, 10 mg of bumetanide was added to 1 mL of deionized water containing 1.0%, 0.5%, or 0.1% of the surfactant tetradecyl-β-D-maltoside. Bumetanide was then solubilized using a vortex and sonicator, and then filtered using a 0.22 μm nylon syringe filter. 100 μL of the filtered bumetanide solution was added to 900 μL of deionized water, and then the solution concentration of bumetanide was analyzed using HPLC. The resulting bumetanide concentrations are recorded in Table 1.

[0061] [Table 1]

[0062] Although the addition of surfactant improved the amount of bumetanide that could be solubilized in water, these results clearly show that bumetanide, when not in salt form, has very low water solubility despite the presence of surfactant.

[0063] Example 2. Solubility of Bumetanide Salts A low pH solution of bumetanide was prepared by weighing 20 mg of bumetanide in acid form and dissolving it in 1 mL of deionized water by vortex mixing. A determined amount of base was added to this low pH bumetanide solution and mixed using a vortex for 2 minutes to produce a solution with a suitable pH for administration. As the pH increased, the bumetanide salts formed in situ partially precipitated, resulting in a saturated solution of the salt form. The saturated solution was then filtered through a 0.22 μm filter to remove any precipitated bumetanide salts. The filtrate was collected and its pH was measured. An aliquot of the saturated solution was assayed for bumetanide by HPLC to determine the maximum concentration achievable for the salt form in water at near neutral pH.

[0064] This procedure was repeated for a variety of bases, including arginine, lysine, potassium, sodium, glycine, histidine, and the cationic lipophilic surfactants soybean lecithin, distearoylglycerol-3-phosphatidylamine, L-α-phosphatidylethanolamine, and bis(2-ethylhexyl)amine, to determine the maximum solubility of each bumetanide salt solution. To determine the maximum solubility of each bumetanide salt, the concentration of bumetanide in solution for each bumetanide salt form is recorded in Table 2.

[0065] [Table 2]

[0066] The solubility data clearly demonstrates that the highest bumetanide salt concentrations in solution are achieved when either arginine or potassium is acting as the base, with maximum concentrations measured at 12.6 mg / mL and 20.19 mg / mL, respectively. The solubility of the bumetanide salt is significantly increased when potassium hydroxide is acting as the base, as compared to another base such as sodium hydroxide, where a solubility of 6.59 mg / mL was measured, as shown in FIG. 1. Surprisingly, the solubility of the bumetanide salt is significantly increased when arginine is acting as the base, as compared to another amino acid base such as lysine, where a solubility of 7.08 mg / mL was measured. As shown in FIG. 1, the potassium salt of bumetanide provides the lowest mass loading into the transmucosal formulation, the highest solubility at a pharma- ceutical desirable pH (e.g., pH 6-8), and a concentration that can provide PK performance similar to that provided by IV injection (as shown in animal studies) without the complications and risks associated with IV administration.

[0067] Example 3. Solubility of Bumetanide in Different pH Buffers The solubility of bumetanide was evaluated in various pH buffers. 10 mg of bumetanide was weighed into a 3 mL glass vial. To this vial, 2 mL of buffer with the desired pH was added. The solution was then sonicated for 2 hours at room temperature (25 °C) and then rotated overnight. The solution was then filtered through a 0.2 μm nylon syringe filter and the first 0.5 mL was discarded. The filtrate was then diluted 10-fold and at least 0.5 mL was aliquoted into HPLC vials. 1 M sodium citrate was used for F1-F4 and 1 M sodium phosphate buffer was used for F5-F9. The resulting bumetanide concentration in solution for each pH buffer was measured and is summarized in Table 3.

[0068] [Table 3]

[0069] 1 g of Kolliphor RH40 and 9 g of ethanol were added to a 15 mL Falcon tube. The solution was mixed and vortexed as necessary. 50 mg of bumetanide salt was weighed and transferred into the solution and mixed, vortexed, or sonicated as necessary to dissolve the solids to create a 5 mg / g stock solution. When all the solids had dissolved, an additional 10 mg of bumetanide salt was added. 1.0 g of this stock solution was added to a 10 mL glass vial using a 0.22 μm nylon syringe filter. The pH of the solution was measured and adjusted with 0.1 N HCl or NaOH to within ±0.1 of the target pH. If there was no precipitate in the vial, 0.5 mL of each formulation was aliquoted into a 1.5 mL HPLC vial for a total of 20 vials. If there was precipitate, the solution with the precipitate was transferred into a 10 mL syringe and filtered through a 0.45 μm filter. The filtrate was collected and aliquoted into 1.5 mL HPLC vials containing 0.5-1.0 mL of each formulation for a total of 15 vials. Each vial was provided with a crimp seal. Four vials were placed at 40°C, four vials at 50°C, and four vials at 60°C. Two vials were placed at -80°C for backup. The last vial was used for the T0 assay analysis. The pH and concentration of the various formulations were measured on the day, 3, 7, and 10 of storage at the indicated temperatures. The results are summarized in Table 4. The recovery of bumetanide was also measured on these days. The results are summarized in Table 5.

[0070] [Table 4]

[0071] [Table 5]

[0072] Example 4. Emulsion Formulation of Bumetanide Salts An emulsion formulation of bumetanide salt was developed. Glycerol (2.25 wt%), medium chain triglyceride (MCT) (10 wt%), and lecithin (E-80) (1.2 wt%) were weighed and dissolved by sonication for 30 minutes, followed by addition of 86 wt% deionized water and further mixing until a clear, colorless solution was obtained to produce the FEV-1 product described in Table 6. FEV-2 was produced by weighing MCT and lecithin into a tared 50 mL Falcon tube and sonicating for approximately 30 minutes at 50° C. to ensure all solids were dissolved. Deionized water was mixed with the solution to produce a white emulsion, as described in Table 6.

[0073] [Table 6]

[0074] Emulsions FE-1 through FE-7 were prepared using the FE-V1 product with various emulsifiers. The FE-8 emulsion was prepared using the FE-V2 product. These emulsions were prepared by weighing bumetanide, emulsifier (e.g., polysorbate 80, polysorbate 20, or PEG 400), and, in the case of FE-3 through FE-6, benzyl alcohol, as described in Table 7. Each emulsion formulation was made up to 1000 mg with FEV-1 solution and BB for 30 seconds per formulation. The appearance was visually inspected and the pH was measured. For FE-6, the pH was adjusted to 5.7 using 1N NaOH, and for FE-7, the pH was adjusted to 6.5 using 1N NaOH. Each emulsion formulation was then filtered using a 0.22 μm filter syringe. The resulting filtrates were analyzed for their respective bumetanide concentrations as well as impurities using HPLC.

[0075] [Table 7]

[0076] Measurement of the concentration of bumetanide showed that the highest concentration was obtained in emulsion formulation FE-6, which had a bumetanide concentration of 6.55 mg / g, as shown in Table 8.

[0077] [Table 8]

[0078] To test the stability of FE-9 at various temperatures, FE-9 was filled into spray vials and placed in a stability chamber at 2-8°C, vials were placed in a stability chamber at 25°C, and vials were placed at -20°C. The remaining vials were used to measure the initial concentration and impurities. After 1 week, 2 weeks, and 4 weeks, samples were removed from the stability chamber and visually inspected for their appearance. The samples were then prepared for HPLC for assay and impurity analysis. At this time, the pH and osmolality of the formulation were measured. The osmolality at T0 was 264 Osm and the viscosity was 1.3 cp run at 10 rpm. The results of the stability study are summarized in Table 9.

[0079] [Table 9]

[0080] Particle size distribution analysis was performed using dynamic light scattering on a Malvern Panalytical Zetasizer to determine that the mean particle size of the FE-9 bumetanide emulsion was less than 200 nm. FE-9 samples were prepared for analysis by passing the emulsion twice through a microfluidizer prior to particle size distribution analysis. If the measured mean particle size was greater than 200 nm, the process was repeated. The mean particle size of FE-9 was 112-120 nm when stored at both room temperature and 2-8°C over a 4-week period, as reported in Table 11.

[0081] [Table 10]

[0082] To prepare FE-8, weigh each excipient into a tared 15 mL Falcon tube (see Table 7) and qs to FEV-2. Adjust pH to 6.0 with 1 N NaOH and vortex to ensure no solids in the mixture. Use sonication if necessary. Final pH was kept at 6.0, inspected for visual appearance, measured pH, and measured osmolality. Filter solution through 0.22 μm filter, collect filtrate, aliquot 1.0 mL into 3 mL glass vials, and crimp to seal vials for a total of 8 vials.

[0083] To test the stability at various temperatures, three vials were placed in a stability chamber at 2-8°C, three vials in a stability chamber at 25°C, and two vials at -20°C. The remaining vials were used to measure the initial concentration and impurities. After four weeks, the samples were removed from the stability chamber and their appearance was visually inspected. The samples were then prepared for HPLC for assay and impurity analysis. At this time, the pH and osmolality of the formulation were measured. Before measuring the osmolality, 0.2 mL of the formulation was diluted with 1.8 mL of deionized water. The concentration and recovery of bumetanide obtained after two weeks for the samples stored at various temperatures are summarized in Table 9. The osmolality was measured to be 5 mOsm at T0 with a dilution factor of 10.

[0084] [Table 11]

[0085] Example 5. Nanoemulsion Formulation of Bumetanide Salts To improve transmucosal absorption, a nanoemulsion formulation of bumetanide salt was developed. To prepare the nanoemulsion, glycocholic acid (46.8 mg) was first weighed and suspended in sterile water for injection (SWFI) (600 mg) using a vortex to generate the FNEV-1 formulation, as listed in Table 12. To this solution, 10 N NaOH was added (10 μL) and mixed until the solution was clear. Soy lecithin (PL90G) and benzyl alcohol were added as listed in Table 12 and completely dissolved using a vortex and sonicator as necessary. The solution was brought to final weight with deionized water and mixed, as listed in Table 12. The pH of this solution was measured and adjusted to a final pH of 7.0, after which the solution was placed on a shaker overnight to generate the FNEV-1 product.

[0086] [Table 12]

[0087] FNEV-1 products were used to prepare FNE-1 nanoemulsions of various pH values. Bumetanide nanoemulsions were prepared by weighing bumetanide, dissolving it in FNEV-1 solution using a vortex, and adjusting the pH to 5.0 or 6.0 using 1.0 N NaOH or 1.0 N HCl, as described in Table 13. The solution was left overnight, after which the pH was measured. The solution was filtered using a 0.22 μm centrifuge tube filter. The filtrate was then diluted by a factor of 20, such that 0.25 mL of the filtrate was diluted to a final volume of 5 mL, and mixed using a vortex, after which the solution was transferred to the HPLC step.

[0088] [Table 13]

[0089] The concentrations of bumetanide in FNE-1 solutions with final pH values ​​of 5.3, 5.6, and 6.1 were measured using HPLC by comparison with solutions of known bumetanide concentrations, as shown in Table 14. The FNE-1 formulation with a pH of 6.1 exhibited the highest solution concentration of bumetanide with a calculated concentration of 10.1 mg / mL. The results show that the solution concentration of bumetanide more than doubled when the pH of the nanoemulsion was changed from 5.3 to 6.1, thus indicating that a pH of about 6 is optimal for solution concentrations of 5 mg / mL or higher of bumetanide.

[0090] [Table 14]

[0091] Particle size distribution analysis was performed using dynamic light scattering on a Malvern Panalytical Zetasizer to determine the mean particle size of FNE-1 bumetanide nanoemulsion. The mean particle size of FNE-1 at pH 6.0 was 9-11 nm when stored at both room temperature and 2-8°C over a 4 week period, as shown in Table 15.

[0092] [Table 15]

[0093] To test the stability at various temperatures, the filled spray vials were placed in a stability chamber at 2-8°C, vials in a stability chamber at 25°C, and vials at -20°C. The remaining vials were used to determine the initial concentration and impurities. After 1 week, 2 weeks, and 4 weeks, samples were removed from the stability chamber and their appearance was visually inspected. The samples were then prepared for HPLC for assay and impurity analysis, at which time the pH was also measured. After 2-fold dilution, the osmolality of the formulation was measured to be 140 Osm, and the viscosity was 6 cp run at 10 rpm. The concentrations and recoveries of bumetanide obtained for samples stored at various temperatures over a period of 4 weeks are summarized in Table 16.

[0094] [Table 16]

[0095] Example 6. Stability of Bumetanide Salts The stability of arginine and bumetanide potassium salts stored at either 2-8° C. or 25° C. for four weeks was monitored over the relevant time periods. The bumetanide salt solutions were evaluated based on pH, bumetanide concentration in solution, assay recovery, and the presence of impurities (see Tables 17 and 18).

[0096] [Table 17]

[0097] [Table 18]

[0098] For bumetanide arginine salt, the initial pH was 7.0 and after 4 weeks it was 7.0 and 7.1 for samples stored at 5° C. and 25° C., respectively. The bumetanide concentration in solution was determined to be 5.07 mg / mL initially and 4.89 mg / mL and 4.97 mg / mL for samples stored at 5° C. and 25° C., respectively, after 4 weeks. Assay recoveries were 100% initially and 97.8% after 4 weeks for both samples stored at 5° C. and 25° C. Finally, no impurities were detected either initially or after 4 weeks for any samples subjected to HPLC detection.

[0099] These data show that the bumetanide arginine salt is stable for up to four weeks at either 5° C. or 25° C., as shown in Table 17. Additionally, the data show that the bumetanide potassium salt is stable for up to four weeks at either 5° C. or 25° C., as shown in Table 18.

[0100] Example 7. Methods for preparing pharmaceutical formulations of the arginine salt of bumetanide, the potassium salt of bumetanide, and the lysine salt of bumetanide To prepare a pharmaceutical formulation of the arginine salt of bumetanide for subcutaneous administration to patients suffering from congestive heart failure, first, 0.1 g of sodium carboxymethylcellulose was dissolved in 100 mL of deionized water and mixed to make a 0.1% sodium carboxymethylcellulose solution, which was filtered through a 0.2 μm nylon filter. 4 g, 0.5 g, and 0.5 g of mannitol, benzyl alcohol, and 98.3% bumetanide were weighed and mixed, respectively. To this mixture, about 16 g of 0.1% sodium carboxymethylcellulose solution was added. To this solution, 0.45 g of L-arginine was added and mixed using a vortex until the solution was clear. The pH of the solution was then adjusted using 1.0 N HCl to reach a final pH of 7.0. The formulations of the arginine salt of bumetanide in both 1 g and 20 g scales were recorded in Table 19.

[0101] [Table 19]

[0102] [Table 20]

[0103] [Table 21]

[0104] 10 g batches of bumetanide arginine salt (F42), bumetanide potassium salt (F43), and bumetanide lysine salt (F44) for subcutaneous administration to patients suffering from congestive heart failure were formulated according to Table 20. 0.1 g of CMC sodium was added to a 250 mL beaker using a stir bar, 100 mL of deionized water was added, and mixed well to obtain a 0.1% CMC-Na solution. Bumetanide and edetate disodium dihydrate were weighed and placed into a tared 15 mL Falcon tube. Approximately 8 g of 0.1% CMC-Na solution was added to the tube and mixed. To this solution, the amount of arginine, KOH, or lysine listed in Table 20 was added and vortexed until the solution was clear. The appropriate amount of base was added and vortexed until a clear solution was obtained. At this point, the pH was measured and adjusted to pH 7.0 if necessary. The 0.1% CMC-Na solution is brought to a total weight of 10 g and mixed well. Approximately 1 mL of the solution was used to test the osmolality. If necessary, the osmolality was adjusted to approximately 290±10 mOsm / kg with either NaCl or mannitol. After adjusting the osmolality, assay and impurity analyses were performed and the viscosity and sprayability of the solution were measured. The solubility, pH, and osmolality measurements are summarized in Table 21.

[0105] [Table 22]

[0106] 100 g batches of the arginine salt of bumetanide (F69) and the potassium salt of bumetanide (F70) for subcutaneous administration to patients suffering from congestive heart failure were prepared according to Table 22. To a 500 mL beaker, 0.3 g of CMC sodium was added with a stir bar. 300 mL of deionized water was added and mixed well to obtain a 0.1% CMC-Na solution. The mixture was then filtered through a 0.2 μm nylon filter.

[0107] For the arginine salt of bumetanide (F69), bumetanide, benzyl alcohol, and mannitol were weighed into a tared 125 mL Erlenmeyer flask using a stir bar. Approximately 75 g of 0.1% CMC-Na solution was added to the flask and mixed. The appropriate amount of L-arginine was added and the solution was vortex mixed until the solution was clear. At this point the pH was measured and adjusted to pH 7.0 if necessary. The appropriate amount of 0.1% CMC-Na solution was added to bring the total weight to 100 g and the solution was mixed well.

[0108] For the potassium salt of bumetanide (F70), sodium CMC was weighed and placed in a suitable beaker with stirring, to which a certain amount of deionized water was added and the solution was mixed well to produce a 0.1% CMC-Na solution. This solution was then filtered through a 0.2 μm nylon filter. Bumetanide, benzyl alcohol, and mannitol were then weighed and added to the primary formulation container with stirring. To this container, approximately 3 / 4 of the 0.1% CMC-Na was added and mixed or stirred. To this solution, an appropriate amount of 1N KOH was slowly added dropwise and mixed by stirring until the solution was clear. The clear solution was allowed to equilibrate until a stable pH was achieved in the pH range of 6.3 to 7.3. The pH was then measured and readjusted to 7.0 using 1N HCl if necessary. Sufficient 0.1% CMC-Na solution was added and mixed well to bring the solution to the appropriate weight. The osmolality was measured to ensure it was within the specified range.

[0109] Both F69 and F70 were filtered through a 0.2 μm nylon filter. One vial from each formulation was used for appearance, pH, assay / impurity, osmolality, and working dose testing. The results are summarized in Table 23.

[0110] [Table 23]

[0111] Example 8. Analysis of lead formulation over 5 months The stability of bumetanide arginine salt (F69), bumetanide potassium salt (F70), bumetanide emulsion (FE-9), and bumetanide nanoemulsion (FNE-1) was investigated over a 5-month period when stored at 25° C. One sample of F70 was opened for stability testing and observed for 10 months. The appearance of each sample was recorded and is summarized in Table 24.

[0112] [Table 24]

[0113] The concentrations of each of these formulations after 5 months at 25° C. and one sample of F69 stored for 10 months were measured using HPLC analysis. The concentrations obtained are summarized in Table 25. In addition, the assay recoveries at 5 and 10 months were calculated (Table 26).

[0114] [Table 25]

[0115] [Table 26]

[0116] The composition of the bumetanide potassium salt formulations is summarized in Table 27, along with the measured pH, appearance, osmolality, and assay recovery of each formulation.

[0117] [Table 27]

[0118] [Table 28]

[0119] [Table 29]

[0120] Particle size distribution analysis was performed using dynamic light scattering on a Malvern Panalytical Zetasizer to determine the mean particle size of FE-9 and FNE-1 bumetanide emulsions. The mean particle sizes of FE-9 and FNE-1 stored at room temperature for 5 months are listed in Table 28.

[0121] [Table 30]

[0122] At 25°C storage conditions, the assay of F69 after 5 months showed a concentration of 5.13mg / g, which was close to the TO assay, which was 4.98mg / g. The assay of F70 after 5 months showed a concentration of 5.16mg / g, which was close to the TO assay, which was 4.89mg / g. After 10 months, the assay of F70 showed a concentration of 5.21mg / g, which was close to the TO assay, which was 4.89mg / g. After 10 months, the assay recovery of F70 was about 106.5% compared to TO. This could be due to the fact that the samples were sampled from an open container (used), which could be the cause of the loss of water over time when stored at 25°C. The value of the claim rate (%) after 10 months (10M) storage at room temperature was 104%. The assay for FE9 after 5 months measured a concentration of 4.67 mg / g, which was close to the TO assay, which was 4.59 mg / g. The assay for FNE-1 measured 4.83 mg / g, which was close to the TO assay, which was 5.09 mg / g. No impurities were observed in all four formulations when stored at 25° C. for 5 months. The assay recoveries for F69, F70, FE-9, and FNE-1 formulations relative to TO were all greater than 94% at 25° C. for 5 months. No particles, precipitates, or phase separation were observed in all four formulations. These data indicate that all four formulations were stable for 5 months when stored at 25° C. Furthermore, after 5 months, the particle size distributions for both FE-9 and FNE-1 were close to those at TO or T=1W.

[0123] Example 9. Subcutaneous administration of the arginine or potassium salts of bumetanide to patients suffering from congestive heart failure with severe edema due to insufficient gastrointestinal absorption According to the methods described herein, a physician in the art may treat a patient, such as a human patient, to reduce or alleviate symptoms of edema resulting from congestive heart failure. To this end, a physician in the art may have the patient self-administer the potassium or arginine salt of bumetanide. The potassium or arginine salt of bumetanide is administered by a patient experiencing symptoms of congestive heart failure, such as shortness of breath, fatigue, or edema that have not been reduced by the patient's typical daily dose of oral diuretics. The patient then subcutaneously administers the potassium or arginine salt of bumetanide. A typical dosage is administered based on body weight and is within the range of about 0.5-10 mg of the potassium or arginine salt of bumetanide in 12 hours, but not to exceed 10 mg of the potassium or arginine salt of bumetanide in 12 hours without consulting a physician in the art.

[0124] The potassium or arginine salt of bumetanide is administered in 1, 2, 3, or 4 doses over a 4 hour period. Each dose comprises 100-150 μL of buffer solution containing the potassium or arginine salt of bumetanide at a concentration of about 5-25 mg / mL. Each unit dose contains about 0.5-2.5 mg of the potassium or arginine salt of bumetanide, not to exceed 10 mg of pharmaceutical composition for four doses. The potassium or arginine salt of bumetanide is administered to the patient in an amount sufficient to treat symptoms of congestive heart failure according to the patient's self-assessment, including reduced swelling, increased urinary output, and reduced shortness of breath.

[0125] Example 10. Sustained release formulation for subcutaneous administration. The methods and compositions described herein can provide a subcutaneously administered formulation that releases bumetanide to a patient over an extended period of time as a long-acting diuretic (e.g., 8-hour, 12-hour, 24-hour, 36-hour, 48-hour, 3-day, or 5-day diuresis). The sustained release formulation is used to treat patients who are at risk of being re-admitted after treatment for congestive heart failure. The use of a long-acting diuretic allows such patients to be treated on an outpatient basis rather than being re-admitted for edema-related symptoms. The sustained release formulation is administered as a subcutaneous depot (e.g., a depot formed by emulsion) and released from the subcutaneous depot over an extended period of time.

[0126] The sustained release formulation is formed from an aqueous emulsion of bumetanide arginine salt at a concentration of about 7-15 mg / mL bumetanide. Each dose consists of about 100-300 μL of aqueous emulsion, and the dosage level may be determined by the patient's weight and health condition. The emulsion is a mixture formed from glycerol (about 2.25% by weight), medium chain triglycerides (MCT) (about 10% by weight), and lecithin (E-80) (about 1.2% by weight).

[0127] Subcutaneous administration of the depot formulation allows patients to receive the benefits of reliable diuresis over several days without complications. As a result of the sustained release subcutaneous formulation of bumetanide arginine emulsion, patients who are at risk of being re-hospitalized after hospitalization for treatment of congestive heart failure receive treatment after discharge that achieves diuresis over an extended period of time and relieves symptoms such as shortness of breath, fatigue, and edema.

[0128] Example 11. Preparation of bumetanide salts To prepare the bumetanide arginine salt, 1.00 g of bumetanide and 0.53 g of arginine were added to a mortar. Approximately 15 mL of 200 proof ethanol diluted to 50% with deionized water was added to the arginine and bumetanide in the mortar. A slurry was made by manually granulating with a pestle for approximately 30 minutes. The resulting slurry was transferred to a tared beaker. The slurry was then vacuum dried overnight at -20°C. After more than 24 hours of vacuum drying, a loose, fluffy white solid was obtained with a molar ratio of bumetanide to arginine of 1:1.1, as listed in Table 30. All chemicals used to produce the bumetanide arginine salt are listed in Table 29.

[0129] [Table 31]

[0130] [Table 32]

[0131] To prepare bumetanide potassium salt, 1.0 g of bumetanide was added to a 20 mL glass vial. To this glass vial, 5 mL of 200 proof ethanol solution diluted 50% with deionized water was added and mixed by vortexing for approximately 1 minute. To the wet bumetanide, 3.0 mL of 1 N KOH solution was added and mixed with a vortex or spatula for approximately 10 minutes to obtain a white slurry. This mixture was frozen at -20 °C for approximately 1 hour. The mixture was then vacuum dried overnight at -20 °C. Once dry, the white fluffy soft solid bumetanide potassium (the molar ratio of bumetanide to potassium was 1:1.1 as listed in Table 32) was transferred to a clean glass vial. This bumetanide potassium salt was set aside at 2-8 °C for formulation preparation. All chemicals used in the production of bumetanide potassium salt are listed in Table 31.

[0132] [Table 33]

[0133] [Table 34]

[0134] Example 12. Analysis of subcutaneous formulations of potassium and arginine salts of bumetanide Approximately 3.7 g of sterile water for injection (SWFI) was weighed and added to two tared 15 mL centrifuge tubes. One tube was labeled FS-1 and the other was labeled FS-3. Bumetanide was added to each tube as described in Table 33. To each solution, the appropriate base was added (arginine to FS-1 and KOH to FS-3). The solutions were then mixed to dissolve the bumetanide. If the bumetanide did not dissolve, more base was added slowly and the amount added was recorded. To each solution, benzyl alcohol and mannitol were added and brought to 5.0 g with SWFI. The solutions were vortexed to mix and the initial pH was measured. 2.5 g of FS-1 was dispensed into a 15 mL Falcon tube and labeled FS-2. 2.5 g of FS-3 was dispensed into a 15 mL Falcon tube and labeled FS-4. The pH was adjusted for each formulation as listed in Table 33 using 1N HCl and the final pH was recorded.

[0135] [Table 35]

[0136] [Table 36]

[0137] The various formulations were analyzed for appearance, pH, osmolality, and bumetanide concentration as summarized in Table 34. HPLC assay was used to measure bumetanide concentration using the conditions listed in Table 35. HPLC assay was performed by preparing 2 L of methanol:distilled water:tetrahydrofuran:glacial acetic acid in the ratio of 1:45:5:2 (v / v / v / v) and filtering the solution through a 0.8 μM nylon membrane filter. Bumetanide standard with a concentration of 0.125 mg / mL was prepared by weighing 12.5 mg of bumetanide into a 100 mL volumetric flask and dissolving in 100 mL of stock diluent made by adding 10 mL of tetrahydrofuran with 4 mL of glacial acetic acid to a 100 mL volumetric flask and bringing to 100 mL with methanol. Sample solution was prepared by weighing 125 mg of sample into a 10 mL volumetric flask and bringing to 10 mL with distilled water.

[0138] [Table 37]

[0139] Formulations FS-3 and FS-4 containing the potassium salt of bumetanide were clear and transparent at pH 7.4 and 6.7. HPLC results showed that the assay was within the expected target range for bumetanide (10 mg / mL) at both pH values. For FS-3, the osmolality was slightly higher than for FS-4. Formulations FS-1 and FS-2 containing the arginine salt of bumetanide were initially clear but crashed out as a white precipitate over time at both pH values ​​using these conditions.

[0140] In the bumetanide potassium formulations, mannitol was adjusted from 4% to 3% (w / w) in the formulation, resulting in an osmolality range of 280-360 mOSm / kg. The resulting osmolality results are listed in Table 36.

[0141] [Table 38]

[0142] Example 13. Formulation of Bumetanide Salts for Administration to Subjects To prepare the bumetanide formulation for administration, approximately 162.8 g of SWFI was weighed into two tared 250 mL Nalgene bottles with stir bars. One was labeled FS-1 and the other was labeled FS-3. Bumetanide was added to the bottles as described in Tables 37 and 38. Arginine base (FS-5) or KOH base (FS-3) was then added to the solution and mixed to dissolve the bumetanide. If the bumetanide did not dissolve, more base was added slowly and recorded. To the solution, benzyl alcohol and mannitol were added to 5.0 g with SWFI. The solution was vortexed to mix and the initial pH was measured. 110 g of FS-1 was divided into another 100 mL Nalgene bottle and labeled FS-2. 110 g of FS-3 was divided into another 100 mL Nalgene bottle and labeled FS-4. For FS-2 and FS-4, the pH was adjusted to approximately 6.7. For FS-1 and FS-3, the pH was adjusted to approximately 7.4. The solutions were filtered through a 0.22 μm filter in a biosafety hood. Appearance, bumetanide concentration, impurities, pH, and osmolality were evaluated.

[0143] [Table 39]

[0144] Two 15 mL Falcon tubes were tared and one was labeled FS-5 and the other FS-6 to produce the subcutaneous bumetanide arginine formulations, FS-5 and FS-6. To each tube, approximately 8 g of sterile water for injection (SWFI) was added and mixed well until the arginine was completely dissolved. To this solution, benzyl alcohol, bumetanide, and mannitol were added as described in Table 38. The solution was brought to 10 g with SWFI, vortexed to mix, and the initial pH was measured. Formulations FS-5 and FS-6 were adjusted to pH 7.0 and 7.2, respectively, with glacial acetic acid, and then vortexed to mix for 5-10 minutes. The solutions were left at room temperature overnight and protected from light. The solutions were then filtered through a 0.22 μm filter. The solutions were dispensed into crimp-sealed glass vials as 1.0 mL aliquots. Each formulation was evaluated for concentration, pH, and osmolality.

[0145] An intranasal formulation of bumetanide (F82) was made according to Table 39.

[0146] [Table 40]

[0147] [Table 41]

[0148] Example 14. Stability of Bumetanide Salt Formulations Stability of potassium salt formulations Formulations FS-3 and FS-4 were tested for stability at 25° C. and 40° C. for three months. The formulations were filled into Type 1 clear glass vials, stoppered and crimp sealed. The vials were then placed in storage conditions (protected from light) at 25° C. and 40° C. for stability testing. Each sample was unstoppered and tested according to Table 39 below. The results of the stability study are summarized in Tables 40-44.

[0149] [Table 42]

[0150] Panel A (T0 only): Appearance, pH, Osmolality, Injectability (27G-30G) per Annex Method, Density / Viscosity, HPLC Assay / Impurities, USP <788> Particulate matter by smear (visual inspection) was studied.

[0151] Panel B (0.5, 1, 2, 3 min): Appearance, pH, HPLC Assay / Impurities, Viscosity, USP <788> Particulate matter by smear (visual inspection) was studied.

[0152] [Table 43]

[0153] [Table 44]

[0154] [Table 45]

[0155] [Table 46]

[0156] [Table 47]

[0157] Stability of arginine salt formulations Formulations FS-5 and FS-6 were tested for stability at 25°C and 40°C for 9 months. The formulations were filled into Type 1 clear glass vials, stoppered and crimp sealed. The vials were then placed in storage conditions (protected from light) at 25°C and 40°C for stability testing. Each sample was unstoppered and tested according to Table 45 below. The results of the stability study are summarized in Tables 46-50.

[0158] [Table 48]

[0159] Panel A (T0 only): Appearance, pH, Osmolality, Injectability (27G-30G) per Annex Method, Density / Viscosity, HPLC Assay / Impurities, and USP <788> Particulate matter by visual inspection was studied.

[0160] Panel B (0.5, 1, 2, 3 min): Appearance, pH, HPLC Assay / Impurities, Viscosity, and USP <788> Particulate matter by visual inspection was studied.

[0161] [Table 49]

[0162] [Table 50]

[0163] [Table 51]

[0164] [Table 52]

[0165] [Table 53]

[0166] Stability of nanoemulsion formulations Formulation EF-6 was tested for stability at 25° C. and 40° C. for 3 months. Formulation FE-6 was filled into Type 1 clear glass vials, stoppered and crimp sealed. The vials were then placed in storage conditions (protected from light) at 25° C. and 40° C. for stability testing. Each sample was unstoppered and tested according to Table 51 below. The results of the stability study are summarized in Tables 52-54.

[0167] [Table 54]

[0168] Panel A (T0 only): Appearance, pH, osmolality, injectability (27G-30G) as per attached method, density / viscosity, and HPLC assay / impurities were studied.

[0169] Panel C (0.5, 1, 2, 3 min): Appearance, pH, Viscosity, HPLC Assay / Impurities, USP <788> The particulate matter and particle size distribution were studied by the method.

[0170] [Table 55]

[0171] [Table 56]

[0172] [Table 57]

[0173] Stability of CLEARSOL™ Formulations The FC-2 formulation was made according to the composition set forth in Table 55 below. The FC-2 formulation was made by adding 0.8 g of ClearSol™ F95V to a 2 mL centrifuge vial. The pH was checked and adjusted to about 8.0 with 1N NaOH. To this composition, 10 mg of bumetanide was added and mixed for about 30 seconds. The appearance as well as the pH was checked and the mass was brought to 1 g with ClearSol F95V. 0.75 mL of the composition was added to a spin-x with a 0.22 μm filter and centrifuged at 13,000 rpm for 2 minutes. The filtrate was then decanted, diluted, and analyzed by high performance liquid chromatography. The formulation was a clear, pale yellowish liquid.

[0174] [Table 58]

[0175] Formulation FC-2 was tested for stability at 25° C. and 40° C. for 3 months. Formulation FC-2 was filled into Type 1 clear glass vials, stoppered and crimp sealed. The vials were then placed in storage conditions (protected from light) at 25° C. and 40° C. for stability testing. Each sample was unstoppered and tested according to Table 56 below. The results of the stability study are summarized in Tables 57-59.

[0176] [Table 59]

[0177] Panel A (T0 only): Appearance, pH, Osmolality, Injectability (27G-30G) per Annex Method, Density / Viscosity, HPLC Assay / Impurities, and USP <788> Particulate matter was studied by the method.

[0178] Panel B (0.5, 1, 2, 3 min): Appearance, pH, Viscosity, HPLC Assay / Impurities, and USP <788> Particulate matter was studied by the method.

[0179] [Table 60]

[0180] [Table 61]

[0181] [Table 62]

[0182] Arginine salt formulations FS-5 and FS-6 showed good stability over 3 months at both room temperature and accelerated conditions (40°C). Potassium salt formulations FS-3 and FS-4 had needle-like precipitates and showed poor appearance after 1 month at room temperature. Nanoemulsion formulation FE-6 showed a significant increase in degradation impurities over 3 months at both room temperature and accelerated conditions (40°C). Clearsol formulation FC-2 showed color formation and a significant increase in degradation impurities after 3 months at accelerated conditions (40°C).

[0183] Example 15. Administration of Salts of Bumetanide The objective of this study was to evaluate the pharmacokinetic profiles of various bumetanide formulations, including bumetanide arginine salt (FS-5 and FS-6) and bumetanide potassium salt (FS-3 and FS-4) for subcutaneous administration, as well as the bumetanide intranasal formulation (F82), when administered as single subcutaneous or intranasal doses in different vehicles to male and female beagle dogs, as compared to a single IV dose, as described in Table 60.

[0184] A total of four dogs ranging from 1 to 3 years old were assigned to the study (three males, 9-13 kg, and one female, 7-10 kg).

[0185] The study involved 6 events in the same animals, separated by a washout period of at least 48 hours or more between events.

[0186] Subcutaneous administration of bumetanide formulations was performed by subcutaneous injection using a 25G hypodermic needle (100 μL / dog). After each application, successful dosing was confirmed by visual inspection of the dog at the injection site and 1 minute after dosing. Only successful dosing will be accepted.

[0187] For intranasal dosing, each dog was held in an upright position with its nose facing up. Test article (50 μL) was delivered to each nostril (total of 100 μL / dog). Nose was held upright for approximately 60 seconds after dosing. Successful dosing was confirmed by visual inspection of the pipette tip and the dog's nostril after each application.

[0188] For intravenous administration, a commercially available injection of bumetanide (Walgreens, 0.25 mg / mL) was used and given over 30 seconds via the cephalic vein (4.0 mL / dog, IV).

[0189] The body weight of each dog was measured before each dose on the dosing day and approximately 48 hours after dosing at each event. At some events, the body weight 48 hours after dosing was the same as the pre-dose body weight for the following dose. Diuresis was assessed by monitoring post-dose urine volume by observations at 30 minutes, 1 hour, 2 hours, 3 hours, and 4 hours after dosing. Urine volume was not measured. Blood (approximately 600 μL / time point, jugular or cephalic vein) was collected into red top tubes (Sarstedt, Inc-41.1392.105 or similar) with clot activator at the time points listed in Table 61 to obtain serum on each dosing day. Blood was centrifuged at 5000×g for 10 minutes at 4° C. If samples were collected outside of the collection period, they were not considered protocol violations. Serum was divided into two aliquots of approximately 150 μL each and stored frozen at −20° C. before being shipped to Climax Laboratories, Inc. on dry ice.

[0190] [Table 63]

[0191] [Table 64]

[0192] The individual serum concentrations and pharmacokinetic parameters of bumetanide following IV administration of commercial bumetanide to dogs are shown in Table 62 and graphically depicted in Figures 2-4. Bumetanide levels also declined rapidly following IV administration of bumetanide.

[0193] [Table 65]

[0194] Individual serum concentrations and pharmacokinetic parameters of bumetanide following subcutaneous administration of the bumetanide potassium formulations are shown in Tables 63 and 64. When the bumetanide formulations were delivered subcutaneously, bumetanide potassium formulations FS-4 and FS-3 provided the greatest absorption of bumetanide, as shown in Figures 2-4. Subcutaneous administration of bumetanide potassium provided higher serum concentrations compared to IV administration of bumetanide, as shown in Figures 2 and 3. Subcutaneous administration provided rapid administration of FS-4 and FS-3, and more sustained levels over time compared to IV administration, as shown in Figure 3. Formulations FS-3 and FS-4 demonstrated significantly higher AUCs compared to the FS-5 and FS-6 arginine salt formulations and IV administration (Figure 4).

[0195] [Table 66]

[0196] [Table 67]

[0197] The individual serum concentrations and pharmacokinetic parameters of bumetanide following subcutaneous administration of the bumetanide arginine formulations are shown in Tables 65 and 66. When the bumetanide formulations were delivered subcutaneously, the bumetanide arginine formulations FS-5 and FS-6 resulted in lower absorption of bumetanide than the bumetanide potassium formulation, as shown in Figures 2-4.

[0198] [Table 68]

[0199] [Table 69]

[0200] The individual serum concentrations and pharmacokinetic parameters of bumetanide following intranasal administration of the bumetanide formulations are shown in Table 67. When the bumetanide formulations were delivered intranasally, the bumetanide formulation F82 provided similar absorption of bumetanide as the bumetanide arginine formulation, as shown in Figure 4.

[0201] [Table 70]

[0202] Pharmacokinetic data collected upon administration of bumetanide formulations to dogs are summarized in Table 68. These data indicate that the bioavailability and systemic exposure of bumetanide was high following subcutaneous and intranasal administration.

[0203] [Table 71]

[0204] Example 16. Subcutaneous administration of the potassium salt of bumetanide to patients suffering from congestive heart failure with severe edema due to insufficient gastrointestinal absorption According to the methods described herein, a physician of the art can treat a patient, such as a human patient, to reduce or alleviate symptoms of edema resulting from congestive heart failure. To this end, a physician of the art can have the patient self-administer the potassium salt of bumetanide or administer the potassium salt of bumetanide to the patient. The potassium salt of bumetanide includes formulations FS-3 or FS-4. The potassium salt of bumetanide is administered by or to a patient experiencing symptoms of congestive heart failure, such as shortness of breath, fatigue, or edema that have not been reduced by the patient's typical daily dose of oral diuretics. The patient then receives the potassium salt of bumetanide subcutaneously in an amount of 50 μl to 100 μl. A typical dosage is administered based on body weight and is in the range of about 0.5 to 10 mg of the potassium salt of bumetanide in 12 hours, but not to exceed 10 mg of the potassium salt of bumetanide in 12 hours without consulting a physician of the art. Subcutaneous formulations of potassium salts are administered in 1, 2, 3, 4, 5, or 6 doses over a 4-hour period.

[0205] Other embodiments Various modifications and variations of the disclosure described herein will be apparent to those skilled in the art without departing from the scope and spirit of the disclosure. Although the disclosure has been described in connection with a specific embodiment, it should be understood that the claimed disclosure should not be unduly limited to such specific embodiment. Indeed, various modifications of the described modes for carrying out the disclosure that are apparent to those skilled in the art are intended to be within the scope of the disclosure. This application claims the benefit of U.S. Provisional Application No. 63 / 208,863, filed June 9, 2021, which is incorporated herein by reference in its entirety.

[0206] Other embodiments are found in the claims.

Claims

1. A pharmaceutical composition for use in the treatment of edema in a subject, comprising (i) an aqueous solution having a pH of from about 5 to about 9, (ii) bumetanide arginine salt at a concentration of from about 4 mg / mL to about 20 mg / mL, and (iii) one or more pharmaceutically acceptable excipients, wherein the treatment comprises administering a therapeutically effective amount of the pharmaceutical composition subcutaneously.

2. The pharmaceutical composition according to claim 1, wherein the pharmaceutical composition comprises (i) an aqueous solution having bumetanide arginine salt at a concentration of from about 5 mg / mL to about 12 mg / mL, (ii) one or more pharmaceutically acceptable excipients, and (iii) the aqueous solution has a pH of from about 6 to about 8.

3. The pharmaceutical composition according to claim 1 or 2, wherein the pharmaceutical composition comprises a buffering agent.

4. The pharmaceutical composition according to claim 3, wherein the aqueous solution does not contain a buffering agent other than a buffer formed by bumetanide free acid combined with arginine.

5. A pharmaceutical composition for use in the treatment of edema in a subject, comprising (i) an aqueous solution having a pH of from about 5 to about 9, (ii) bumetanide potassium salt at a concentration of from about 4 mg / mL to about 20 mg / mL, and (iii) one or more pharmaceutically acceptable excipients, wherein the treatment comprises administering a therapeutically effective amount of the pharmaceutical composition subcutaneously.

6. The pharmaceutical composition according to claim 5, wherein the pharmaceutical composition comprises (i) an aqueous solution having bumetanide potassium salt at a concentration of from about 5 mg / mL to about 12 mg / mL, (ii) one or more pharmaceutically acceptable excipients, and (iii) the aqueous solution has a pH of from about 6 to about 8.

7. The pharmaceutical composition according to claim 5 or 6, wherein the pharmaceutical composition comprises a buffering agent.

8. The pharmaceutical composition according to claim 7, wherein the aqueous solution does not contain a buffering agent other than a buffer formed by bumetanide free acid combined with potassium hydroxide.

9. The pharmaceutical composition according to claim 1 or 5, wherein the pharmaceutically acceptable excipient contains an isotonic agent.

10. The pharmaceutical composition according to claim 1 or 5, wherein the aqueous solution has a pH of about 6 to about 8.

11. The pharmaceutical composition according to claim 1 or 5, wherein the one or more pharmaceutically acceptable excipients contain sugar or sugar alcohol.

12. The pharmaceutical composition according to claim 11, wherein the pharmaceutical composition contains sucrose and / or mannitol.

13. The pharmaceutical composition according to claim 1 or 5, wherein the one or more pharmaceutically acceptable excipients contain a preservative.

14. The pharmaceutical composition according to claim 13, wherein the pharmaceutical composition contains benzyl alcohol.

15. The pharmaceutical composition according to claim 1 or 5, wherein the pharmaceutical composition is administered with a dosing volume of 0.5 mL or less.

16. The pharmaceutical composition according to claim 15, wherein the pharmaceutical composition is administered with a dosing volume of 25 μl to 400 μl.

17. The pharmaceutical composition according to claim 1 or 5, wherein the pharmaceutical composition has at least 85% bioavailability when administered to the subject.

18. The pharmaceutical composition according to claim 17, wherein the pharmaceutical composition has at least 95% bioavailability when administered to the subject.

19. The pharmaceutical composition according to claim 1 or 5, wherein the edema is refractory to oral diuretics.

20. The pharmaceutical composition according to claim 1 or 5, wherein the subject has congestive heart failure.

21. The pharmaceutical composition according to claim 1 or 5, wherein the subject suffers from pulmonary edema.

22. The pharmaceutical composition according to claim 1 or 5, wherein the pharmaceutical composition is administered subcutaneously to the subject in an external environment or the pharmaceutical composition is self-administered.

23. The pharmaceutical composition according to claim 1 or 5, wherein the subject was unable to achieve diuresis with oral diuretic therapy prior to the administration.

24. The pharmaceutical composition according to claim 1 or 5, wherein the subject has experienced lower extremity swelling, shortness of breath, dyspnea, or chest pain prior to the administration that is not resolved by oral diuretic therapy.

25. The pharmaceutical composition according to claim 1 or 5, wherein the subject has experienced reduced bowel motility prior to the administration.

26. The pharmaceutical composition according to claim 1 or 5, wherein the one or more pharmaceutically acceptable excipients include a viscosity increasing agent.

27. The pharmaceutical composition according to claim 26, wherein the excipient is sodium carboxymethylcellulose of low viscosity.

28. A pharmaceutical composition comprising an aqueous solution of bumetanide arginine formulated for subcutaneous administration having a pH of about 6 to about 8 and a concentration of about 5 mg / mL to 12 mg / mL, and a pharmaceutical excipient comprising a sugar or a sugar alcohol.

29. The pharmaceutical composition according to claim 28, wherein the bumetanide arginine has a concentration of about 5 mg / mL to about 10 mg / mL.

30. A pharmaceutical composition comprising an aqueous solution of bumetanide potassium formulated for subcutaneous administration having a pH of about 6 to about 8 and a concentration of about 5 mg / mL to 12 mg / mL, and a pharmaceutical excipient comprising a sugar or a sugar alcohol.

31. The pharmaceutical composition according to claim 30, wherein the bumetanide potassium has a concentration of about 5 mg / mL to about 10 mg / mL.

32. The pharmaceutical composition according to any one of claims 28 to 31, wherein the aqueous solution has a pH of from about 6.5 to about 7.

5.

33. The pharmaceutical composition according to any one of claims 28 to 31, wherein the pharmaceutical composition contains benzyl alcohol.

34. The pharmaceutical composition according to claim 33, wherein the benzyl alcohol is in an amount of from about 0.2% (w / w) to about 1% (w / w).

35. The pharmaceutical composition according to claim 34, wherein the benzyl alcohol is in an amount of about 0.5% (w / w).

36. The pharmaceutical composition according to any one of claims 28 to 31, wherein the sugar alcohol is mannitol.

37. The pharmaceutical composition according to claim 36, wherein the mannitol is in an amount of from about 2.0% (w / w) to about 5% (w / w).

38. The pharmaceutical composition according to claim 37, wherein the mannitol is in an amount of from about 2.5% (w / w) to about 4% (w / w).

39. The pharmaceutical composition according to any one of claims 28 to 31, wherein the pharmaceutical composition contains a buffer selected from citrate buffer and phosphate buffer.

40. The pharmaceutical composition according to any one of claims 28 to 31, wherein the pharmaceutical composition contains low viscosity sodium carboxymethyl cellulose.

41. The pharmaceutical composition according to claim 40, wherein the low viscosity sodium carboxymethyl cellulose is in an amount of from about 0.05% (w / w) to about 0.1% (w / w).

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