LEVOTHYROXINE FORMULATIONS

CROSS REFERENCE TO PRIOR APPLICATIONS

This application claims priority under the Paris Convention to US provisional Patent Application Serial No. 62/538,912, filed July 31 , 2017, which is incorporated herein by reference as if set forth in its entirety.

FIELD OF THE DISCLOSURE

The present description relates generally to the field of levothyroxine. More particularly, the description relates to pharmaceutical formulations containing levothyroxine.

BACKGROUND OF THE DISCLOSURE

Thyroid hormones (THs) mediate important physiological processes such as development, growth, and metabolism in many tissues of the body. There are two major THs secreted by the thyroid gland, levothyroxine (T4) and tri-iodothyronine (T3), with the latter serving as the more biologically active form. The concentrations of T4 and T3 in the blood are regulated by the hypothalamic/pituitary/thyroid (HPT) axis. The hypothalamus secrets thyrotropin-releasing hormone (TRH) which is transported via the hypothalamic-hypophyseal portal system to the anterior pituitary gland where it binds to TRH receptors. This induces thyroid-stimulating hormone (TSH) release from pituitary thyrotroph cells. After TSH is released into the circulation, it eventually binds to TSH receptors located primarily in the thyroid gland. The activation of TSH receptors leads to thyrocyte proliferation, thyroglobulin and sodium-iodide symporter gene transcription, and stimulation of TH synthesis and secretion.

The majority of TH secreted by the thyroid gland is T4. TH synthesis in the thyroid gland requires several steps which includes the uptake of iodide by active transport, thyroglobulin (Tg) biosynthesis, oxidation and binding of iodide to Tg, and oxidative coupling of two iodotyrosines into iodothyronines. The synthesis of TH in thyroid gland is unilateral as evidenced by the iodide uptake, which leads to a 30-fold increase in intracellular iodide concentration in thyrocyte vs. serum. The release of TH from the thyroid gland is stimulated by TSH and followed by intracellular proteolysis and hydrolysis.

THs are transported by specific carrier proteins via the circulation to tissues throughout the body and also must pass the blood/brain barrier for delivery to the Central Nervous System (CNS). Approximately 0.03% of the total serum T4 and 0.3% of the total serum T3 are present in free or unbound form in human. TH binding to these carrier proteins ensures an even distribution and delivery of hormone throughout the body. Intracellular uptake of TH occurs by specific TH transporters, and the intracellular concentration of TH is further regulated by intracellular deiodinases that convert T4 to T3 to increase the TH activity or transform the THs to inert metabolites to reduce it. Intracellular THs then bind to nuclear thyroid hormone receptors (TRs), members of the nuclear receptor superfamily. TRs activate the gene transcription and synthesis of messenger RNA and cytoplasmic proteins. The physiological actions of THs are produced predominately by T3, the majority of which is derived from T4 by deiodination in peripheral tissues.

TH activity is an important determinant of development and growth, and in adults plays a critical role in the regulation of the function and metabolism of virtually every organ system. Hypothyroid patients are deficient in endogenously produced THs. There are many causes to hypothyroidism, including but not limited to, autoimmune disease, e.g. Hashimoto's thyroiditis and atrophic thyroiditis; surgical removal of part or all of the thyroid gland; radiation treatment; congenital hypothyroidism; and antithyroid medicines. Synthetic levothyroxine is a recommended replacement therapy for acute and chronic cases of hypothyroidism, providing patients long term control of their symptoms with a favourable side effect profile and a long serum half-life.

For less severe hypothyroidism patients, oral administration of levothyroxine sodium tablet or solution can restore the steady-state levels of T4 and TSH within 6 weeks. For severe hypothyroid patients or patients with myxedema coma requiring hospitalization, intravenous levothyroxine is administered initially with a loading dose followed by a daily maintenance dose until the patient's thyroid levels and symptoms are controlled where they are then transitioned to oral levothyroxine replacement therapy.

Levothyroxine sodium for injection is a sterile, lyophilized product for parental administration of levothyroxine sodium for thyroid replacement therapy in primary, secondary and tertiary hypothyroidism. Levothyroxine sodium for injection is particularly useful when thyroid replacement is needed on an emergency basis, for short term thyroid replacement, and/or when oral administration is not possible.

Conventional formulations of levothyroxine sodium for injection are preservative-free lyophilized powders containing synthetic crystalline levothyroxine sodium and the excipients mannitol, tribasic sodium phosphate or dibasic sodium phosphate and sodium hydroxide. In some conventional lyophilized formulations contain 10 mg of mannitol, 700 μg of tribasic sodium phosphate and either 200 μg or 500 μg of levothyroxine sodium. In some conventional lyophilized formulations contain 1 to 5 mg of mannitol, 400 to 600 μg of dibasic sodium phosphate and 100 to 500 μg of levothyroxine sodium. Administration of the conventional lyophilized formulation involves reconstitution of the lyophilized powder in 5 mL of 0.9% sodium chloride injection (USP), to provide injectable solutions having levothyroxine sodium concentrations of 20 μg/mL, 40 μg/mL or 100 μg/mL for the 10Omcg/vial,

200mcg/vial and 500mcg/vial presentations, respectively. Levothyroxine sodium is not a very stable compound. It is very hygroscopic and degrades rapidly under conditions of high humidity or in the presence of other moisture sources or light and under conditions of high temperature, especially in the presence of moisture or other pharmaceutical excipients such as certain

carbohydrates. Won CM et al. studied the kinetics of the levothyroxine degradation and concluded that levothyroxine sodium in solution degraded by deiodination. The degradation was pH dependent and followed first order kinetics. The log k - pH profile of deiodination of levothyroxine sodium showed a plateau in the acidic pH region, dropped off sigmoidally in the neutral pH region and showed another plateau in the alkaline region. The authors concluded that the kinetics of deiodination include proton attack on the anion and dianion in acidic solution and water attack on the anion and dianion in basic solution. Won CM et al. ("Kinetics of degradation of levothyroxine in aqueous solution and in solid state". Pharm Res. 1992;9: 131 -7) further concluded that in solid state, the degradation of levothyroxine sodium indicated a deamination reaction following bi-phasic degradation pattern.

Levothyroxine sodium degraded at a faster rate as the temperature increased and showed bi-phasic degradation kinetics. There was little or no degradation observed at 50°C, above which temperature, levothyroxine sodium degradation was observed. In another example, Wortsman J et al. ("Thermal inactivation of L-thyroxin". Clin Chem. 1989; 35:90-2) concluded that upon heating levothyroxine sodium rapidly degrades at >90°C, and fully decomposes at melting point (148.81 °C). Kazemiford et al. (2001 . "Identification and quantitation of sodium-thyroxine and its degradation products by LC using electrochemical and MS detection". J. Pharm. Biomed. Anal., 25, 697-71 1 ) studied the photo-degradation of levothyroxine sodium tablets from three manufacturers. The extracted levothyroxine sodium solution was irradiated with a 500W Xenon lamp at 320 nm for 2 hours. The observed degradation products were Liothyroxine, Diiodothyronine, lodothyronine, Diiodotyrosine, lodotyrosine and Tyrosine. It was concluded that levothyroxine sodium is photosensitive.

There are no current marketed solutions of levothyroxine in the US and Canadian Market. Previous liquid formulation inventions have been developed that focus on obtaining sufficient solubility of levothyroxine in oral solution through reliance of solubility enhances such as cyclodextrins, complexing agents or co-polymers. There is a need for improved and stable formulations of levothyroxine.

SUMMARY OF THE INVENTION

The details of one or more embodiments are set forth in the accompanying description below. Other features and advantages will become apparent from the description, and the claims.

In an aspect there is provided an aqueous parenteral formulation comprising levothyroxine or a pharmaceutically acceptable salt thereof and one or more pharmaceutically acceptable excipients.

In an embodiment of the formulation, the pharmaceutically acceptable salt of levothyroxine is levothyroxine sodium. In an embodiment of the formulation, the concentration of levothyroxine sodium in the formulation is between about 5 and about 500 μg/mL.

In an embodiment of the formulation, the formulation comprises: (i) levothyroxine sodium; (ii) one or more antioxidants; (iii) one or more chelating agents; (iv) one or more buffering agents; (v) one or more pH adjusting agents; and (vi) one or more solvents.

In an embodiment of the invention, the formulation comprises (i) levothyroxine sodium; (ii) one or more antioxidants; (iii) one or more chelating agents; (iv) one or more stabilizing agents; (v) one or more buffering agents; (vi) one or more pH adjusting agents; and (vii) one or more solvents.

In various embodiments of the formulation, the one or more antioxidants is selected from sodium sulfite, sodium bisulfite, sodium metabisulfite, potassium metabisulfite, alpha-tocopherol, acetone sodium bisulfite, ascorbic acid, sodium ascorbate, butylated hydroxyanisole, butylated hydroxytoluene, gentisic acid, gentisic ethanolamide, glutathione, methionine, monothioglycerol, and sodium formaldehyde sulfoxylate.

In various embodiments of the formulation, the one or more chelating agents is selected from edetate disodium, edetate disodium anhydrous, edetate sodium, edetate calcium disodium, edetate calcium disodium anhydrous, edetic acid, anhydrous citric acid, citric acid monohydrate, gluceptate sodium, pentasodium pentetate, pentetate calcium trisodium, and pentetic acid.

In various embodiments of the formulation, the one or more buffering agents is selected from sodium phosphate, dibasic, heptahydrate; sodium phosphate, dibasic; sodium phosphate, dibasic, anhydrous; sodium phosphate, dibasic dehydrate;

sodium phosphate, dibasic dodecahydrate; sodium phosphate; sodium phosphate dehydrate; sodium phosphate, monobasic, anhydrous; sodium phosphate, monobasic, dehydrate; sodium phosphate, monobasic, monohydrate; dibasic potassium phosphate; potassium phosphate, monobasic; sodium acetate; sodium acetate anhydrous; ammonium acetate; sodium citrate; disodium hydrogen citrate; anhydrous trisodium citrate; disodium citrate sesquihydrate; trisodium citrate dehydrate; sodium lactate; (L)-sodium lactate; sodium tartrate; ammonium sulfate; and ethanolamine hydrochloride.

In various embodiments of the formulation, the one or more pH adjusting agents is selected from sodium hydroxide; calcium hydroxide; potassium hydroxide; sodium bicarbonate; sodium carbonate; sodium carbonate decahydrate; sodium carbonate monohydrate; diethanolamine; meglumine; tromethamine; ammonia; hydrochloric acid; acetic acid; acetic anhydride; adipic acid; anhydrous citric acid;

benzenesulfonic acid; boric acid; citric acid monohydrate; lactic acid; (DL)-lactic acid; (L)-lactic acid; maleic acid; metaphosphoric acid; methanesulfonic acid; nitric acid; phosphoric acid; succinic acid; sulfuric acid; sulfurous acid; tartaric acid; (DL)-tartaric acid; and trifluoroacetic acid.

In various embodiments of the formulation, the one or more solvents is selected from water for injection (USP), propylene glycol, glycerin, and benzyl alcohol.

In various embodiments of the formulation, wherein the one or more stabilizing agents is selected from sodium iodide, potassium iodide, povidone, povidone K12, povidone K17, crospovidone, sorbitol, and sorbitol solution.

In various embodiment of the formulation, the formulation comprises (i) levothyroxine sodium; (ii) one or more antioxidants selected from sodium sulfite, sodium bisulfite, sodium metabisulfite, and potassium metabisulfite; (iii) one or more chelating agents selected from edetate disodium, edetate disodium anhydrous, edetate sodium, edetate calcium disodium, edetate calcium disodium anhydrous, and edetic acid; (iv) one or more buffering agents selected from sodium phosphate, dibasic,

heptahydrate; sodium phosphate, dibasic; sodium phosphate, dibasic, anhydrous; sodium phosphate, dibasic dehydrate; sodium phosphate, dibasic dodecahydrate; sodium phosphate; sodium phosphate dehydrate; sodium phosphate, monobasic, anhydrous; sodium phosphate, monobasic, dehydrate; sodium phosphate, monobasic, monohydrate; dibasic potassium phosphate; and potassium phosphate, monobasic; (v) one or more pH adjusting agents selected from sodium hydroxide; calcium hydroxide; potassium hydroxide; sodium bicarbonate; sodium carbonate; sodium carbonate decahydrate; sodium carbonate monohydrate; diethanolamine; meglumine; tromethamine; and ammonia; and (vi) one or more solvents selected from water for injection (USP), propylene glycol, glycerin, and benzyl alcohol.

In various embodiments of the formulation, the formulation comprises (i)

levothyroxine sodium; (ii) one or more antioxidants selected from sodium sulfite, sodium bisulfite, sodium metabisulfite, and potassium metabisulfite; (iii) one or more chelating agents selected from edetate disodium, edetate disodium anhydrous, edetate sodium, edetate calcium disodium, edetate calcium disodium anhydrous, edetic acid; (iv) one or more stabilizing agents selected from sodium iodide and potassium iodide; (v) one or more buffering agents selected from sodium phosphate, dibasic, heptahydrate; sodium phosphate, dibasic; sodium phosphate, dibasic, anhydrous; sodium phosphate, dibasic dehydrate; sodium phosphate, dibasic dodecahydrate; sodium phosphate; sodium phosphate dehydrate; sodium

phosphate, monobasic, anhydrous; sodium phosphate, monobasic, dehydrate;

sodium phosphate, monobasic, monohydrate; dibasic potassium phosphate; and potassium phosphate, monobasic; (vi) one or more pH adjusting agents selected from sodium hydroxide; calcium hydroxide; potassium hydroxide; sodium

bicarbonate; sodium carbonate; sodium carbonate decahydrate; sodium carbonate monohydrate; diethanolamine; meglumine; tromethamine; and ammonia; and (vii) one or more solvents selected from water for injection (USP), propylene glycol, glycerin, and benzyl alcohol.

In various embodiments of the formulation, the formulation comprises (i)

levothyroxine sodium; (ii) sodium sulfite; (iii) edetate disodium, edetate disodium anhydrous, or edetate sodium; (iv) sodium phosphate, dibasic, heptahydrate; sodium phosphate, dibasic; sodium phosphate, dibasic, anhydrous; sodium phosphate, dibasic dehydrate; sodium phosphate, dibasic dodecahydrate; sodium phosphate; sodium phosphate dehydrate; or dibasic potassium phosphate; (v) sodium

hydroxide; calcium hydroxide; potassium hydroxide; sodium bicarbonate; sodium carbonate; sodium carbonate decahydrate; sodium carbonate monohydrate;

diethanolamine; meglumine; tromethamine; or ammonia; and (vi) water for injection (USP).

In various embodiments of the formulation, the formulation comprises (i)

levothyroxine sodium; (ii) sodium sulfite; (iii) edetate disodium, edetate disodium anhydrous, or edetate sodium; (iv) sodium iodide or potassium iodide; (v) sodium phosphate, dibasic, heptahydrate; sodium phosphate, dibasic; sodium phosphate, dibasic, anhydrous; sodium phosphate, dibasic dehydrate; sodium phosphate, dibasic dodecahydrate; sodium phosphate; sodium phosphate dehydrate; or dibasic potassium phosphate; (vi) sodium hydroxide; calcium hydroxide; potassium hydroxide; sodium bicarbonate; sodium carbonate; sodium carbonate decahydrate; sodium carbonate monohydrate; diethanolamine; meglumine; tromethamine; and ammonia; and (vii) water for injection (USP).

In an embodiment of the formulation, the formulation comprises (i) levothyroxine sodium; (ii) sodium sulfite; (iii) edetate disodium; (iv) sodium phosphate, dibasic, heptahydrate; (v) sodium hydroxide; and (vi) water for injection (USP).

In an embodiment of the formulation, the formulation comprises (i) levothyroxine sodium; (ii) sodium sulfite; (iii) edetate disodium; (iv) sodium iodide; (v) sodium phosphate, dibasic, heptahydrate; (vi) sodium hydroxide; and (vii) water for injection (USP).

In an embodiment of the formulation, the formulation comprises between about 5 and about 500 μg/mL of levothyroxine sodium.

In an embodiment of the formulation, the formulation comprises between about 0.01 % and about 0.5% w/v of sodium sulfite. In an embodiment of the formulation, the formulation comprises between about 0.005% and about 0.5% w/v of edetate disodium.

In an embodiment of the formulation, the formulation comprises between about 0.01 % and about 5% w/v of sodium phosphate, dibasic, heptahydrate.

In an embodiment of the formulation, the formulation comprises between about 0.01 % and about 5% w/v of sodium iodide.

In an embodiment of the formulation, the pH of the formulation is between about 9.5 and about 1 1 .5.

In an embodiment of the formulation, when the formulation is stored at 60°C for a period of 4 weeks, at least 90% levothyroxine (e.g. , levothyroxine sodium) remains. In other embodiments, when the formulation is stored at 60°C for a period of 4 weeks, less than 0.15% liothyronine and/or less than <0.61 % total impurity levels are present in the formulation.

In an embodiment of the formulation, when the formulation is stored at 60°C for a period of 2 weeks, at least 90% levothyroxine (e.g., levothyroxine sodium) remains. In other embodiments, when the formulation is stored at 60°C for a period of 2 weeks, less than 0.1 1 % liothyronine and/or less than <0.18% total impurity levels are present in the formulation.

In an embodiment of the formulation, when the formulation is stored at 60°C for a period of 2 weeks, at least 90% levothyroxine (e.g. , levothyroxine sodium) remains. In other embodiments, when the formulation is stored at 60°C for a period of 2 weeks, less than 0.16% liothyronine and/or less than <0.32% total impurity levels are present in the formulation.

In an embodiment of the formulation, when the formulation is stored at 60°C for a period of 40 days, less than about 5% of levothyroxine (e.g., levothyroxine sodium) undergoes degradation.

In other embodiments of the formulation, when the formulation is stored at 25°C/60% RH conditions for a period of 24 months, less than 1 %, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 1 1 %, 12%, 13%, 14%, 15%, 20%, 25%, 30% or 35% of the

levothyroxine (e.g., levothyroxine sodium) undergoes degradation. In other embodiments of the formulation, the dosage strength of the formulation is 20 mcg/mL, 40 mcg/mL or 100 mcg/mL.

In an embodiment of the formulation, the formulation does not contain a cyclodextrin, such as hydroxypropyl- -cyclodextrin. In an embodiment of the formulation, the formulation does not contain tromethamine. In an aspect there is provided a method of treating myxedema coma in a subject by administering to the subject the aqueous parenteral formulation as disclosed herein.

In an aspect there is provided a method of treating primary, secondary or tertiary hypothyroidism in a subject by administering to the subject the aqueous parenteral formulation as disclosed herein. In an embodiment, the wherein the formulation is for use as replacement or supplemental therapy. In an embodiment of the method, the formulation is administered intravenously to provide an initial loading dose of about 300 to about 500 μg of levothyroxine or a pharmaceutically acceptable salt to the subject. In an embodiment of the method, the formulation is administered intravenously to provide a maintenance daily dose of about 50 to about 100 μg of levothyroxine to the subject. In an embodiment of the method, the levothyroxine is levothyroxine sodium.

Detailed Description of the Invention

As described herein the inventors have provided stable aqueous pharmaceutical formulations of levothyroxine or a pharmaceutically acceptable salt thereof. These are available in an aqueous solution with a sufficient shelf-life that is commercially viable for parenteral or oral use. The advantages of the formulations disclosed herein compared to conventional formulations include: a) the formulations do not need to undergo an expensive lyophilisation process that results in increased manufacturing cost per vial on an industrial scale; b) the formulations do not need to be reconstituted, a process that requires aseptic technique on the part of the health care practitioners, and which can lead to sterility compromise; c) the formulations are ready for administration, and do not require reconstitution, that can delay treatment in an emergency setting; d) the formulations reduce administration errors associated with levothyroxine because the formulations do not require reconstitution using aseptic technique in an emergency setting.

Surprisingly, the formulations of the disclosure provide for stabilization of

levothyroxine in an aqueous solution within the solubility profile of the active ingredient (levothyroxine). Such formulation provide for levothyroxine in ready-to- use solutions.

Definitions

As used in this specification and the appended claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise. Thus, for example, reference to "a container" includes one or more of such containers and reference to "the excipient" includes reference to one or more of such excipients. In describing and claiming the present invention, the following terminology will be used in accordance with the definitions set forth below.

As used herein, the term "levothyroxine" refers to a synthetic or endogenous thyroid hormone with the general formula of (S)-2-Amino-3-[4-(4-hydroxy-3,5- diiodophenoxy)-3,5-diiodophenyl]propanoic acid.

As used herein, the term "levothyroxine sodium" refers to the sodium salt of levothyroxine with the general formula of L-Tyrosine-0-(4-hydroxy-3,5-diiodophenyl)- 3,5-diiodo-monosodium salt.

As used herein, the term "subject" refers to a mammal. Examples of subjects include humans, and may also include other animals such as horses, pigs, cattle, dogs, cats, rats, rabbits, and aquatic mammals.

As used herein, "treat", "treating" and "treatment", means the treatment of a disease in a subject, for example, a human, and includes inhibiting the disease (e.g., decreasing its rate of progression); regressing the disease; relieving or decreasing the severity of one or more symptoms of the disease; and/or curing the disease.

As used herein, "prevent," "preventing," and "prevention" means the prevention of a disease in a subject, and includes inhibiting initiation of the disease; decreasing a predisposition toward the disease; and/or delaying the onset of at least one symptom of the disease.

As used herein, the term "about" is synonymous with "approximately" and is used to provide flexibility to a numerical value or range endpoint by providing that a given value may be "a little above" or "a little below" the value stated. "About" can mean, for example, within 3 or more than 3 standard deviations. "About" can mean within a percentage range of a given value. For example, the range can be ±1 %, ±5%, ±10%, ±20%, ±30%, ±40% or ±50% of a given value. "About" can mean with an order of magnitude of a given value, for example, within 2-fold, 3-fold, 4-fold or 5-fold of a value. However, it is to be understood that even when a numerical value is accompanied by the term "about" in this specification, that express support shall be provided at least for the exact numerical value as well as though the term "about" were not present. As used herein, "comprises," "comprising," "containing" and "having" and the like can have the meaning ascribed to them in patent law and can mean "includes,"

"including," and the like, and are generally interpreted to be open ended terms. The terms "consisting of" or "consists of" are closed terms, and include only the components, structures, steps, or the like specifically listed in conjunction with such terms, as well as that which is in accordance with patent law. "Consisting essentially of" or "consists essentially of" have the meaning generally ascribed to them by patent law. In particular, such terms are generally closed terms, with the exception of allowing inclusion of additional items, materials, components, steps, or elements, that do not materially affect the basic and novel characteristics or function of the item(s) used in connection therewith. For example, trace elements present in a composition, but not affecting the composition's nature or characteristics would be permissible if present under the "consisting essentially of" language, even though not expressly recited in a list of items following such terminology. When using an open ended term, like "comprising" or "including," it is understood that direct support should be afforded also to "consisting essentially of" language as well as "consisting of" language as if stated explicitly and vice versa. In essence, use of one of these terms in the specification provides support for all of the others.

As used herein, a plurality of items, structural elements, compositional elements, and/or materials may be presented in a common list for convenience. However, these lists should be construed as though each member of the list is individually identified as a separate and unique member. Thus, no individual member of such list should be construed as a de facto equivalent of any other member of the same list solely based on their presentation in a common group without indications to the contrary. Concentrations, amounts, and other numerical data may be expressed or presented herein in a range format. It is to be understood that such a range format is used merely for convenience and brevity and thus should be interpreted flexibly to include not only the numerical values explicitly recited as the limits of the range, but also to include all the individual numerical values or sub-ranges encompassed within that range as if each numerical value and sub-range is explicitly recited. As an illustration, a numerical range of "about 1 to about 5" should be interpreted to include not only the explicitly recited values of about 1 to about 5, but also include individual values and sub-ranges within the indicated range. Thus, included in this numerical range are individual values such as 2, 3, and 4 and sub-ranges such as from 1 -3, from 2-4, and from 3-5, etc. , as well as 1 , 2, 3, 4, and 5, individually. This same principle applies to ranges reciting only one numerical value as a minimum or a maximum. Furthermore, such an interpretation should apply regardless of the breadth of the range or the characteristics being described.

Abbreviations

The following abbreviations are used throughout the disclosure:

CMC - Chemistry, Manufacturing and Control

CNS - Central Nervous System

EDTA - edetate disodium

HPLC - High Performance Liquid Chromatography

HPT - hypothalami ic/pituitary/thyroid

ID - Identification

kg - kilogram

meg - microgram

mg - milligram

ml_ - milliliter

mm - millimeter

mOsm - milliosmoles

Na2S03 - Sodium sulfite

Nal - Sodium Iodide

NDA - New Drug Application

NLT - No Less Than

nm - nanometer

NMT - No More Than

Out of Range - Out of Range PG - Propylene Glycol

qs - quantum sufficit

RH - Relative Humidity

RNA - ribonucleic acid

RS - Reference Standard

T3 - tri-iodothyronine

T4 - levothyroxine

Tg - thyroglobulin

Th - Thyroid hormone

TR - nuclear thyroid hormone receptor

TRH - thyrotropin-releasing hormone

TSH - thyroid-stimulating hormone

μg - microgram

μιη - micrometer

USP - United States Pharmacopeia

UV - Ultra violet

w/v - weight by volume

Levothyroxine Formulations

As described herein the inventors have provided aqueous formulations of levothyroxine or a pharmaceutically acceptable salt thereof that are stable for extended storage in high temperature. These formulations are not only stable, but are formulated as, injectable and ready to administer aqueous compositions. Such formulation are also formulated as, oral and ready to administer aqueous compositions. In a preferred embodiment, the formulation is a parenteral formulation.

The aqueous formulations of the present invention comprise levothyroxine or a pharmaceutically acceptable salt thereof and one or more pharmaceutically acceptable excipients. In a preferred embodiment, the salt of levothyroxine is levothyroxine sodium. In one embodiment, the aqueous formulation comprises: (i) levothyroxine or a pharmaceutically acceptable salt thereof

(ii) one or more antioxidants (iii) chelating agents

(iv) one or more buffering agents

(v) one or more pH adjusting agents and

(vi) one or more solvents The levothyroxine may be the free base or may be a pharmaceutically acceptable salt thereof. In a preferred embodiment, the levothyroxine is levothyroxine sodium.

Antioxidants used in the formulations include, but are not limited to sodium sulfite, sodium bisulfite, sodium metabisulfite, potassium metabisulfite, alpha-tocopherol, acetone sodium bisulfite, ascorbic acid, sodium ascorbate, butylated hydroxyanisole, butylated hydroxytoluene, gentisic acid, gentisic ethanolamide, glutathione, methionine, monothioglycerol, and/or sodium formaldehyde sulfoxylate.

Chelating agents used in the formulations include, but are not limited to edetate disodium, edetate disodium anhydrous, edetate sodium, edetate calcium disodium, edetate calcium disodium anhydrous, edetic acid, anhydrous citric acid, citric acid monohydrate, gluceptate sodium, pentasodium pentetate, pentetate calcium trisodium, and/or pentetic acid.

Buffering agents used in the formulations include, but are not limited to sodium phosphate, dibasic, heptahydrate; sodium phosphate, dibasic; sodium phosphate, dibasic, anhydrous; sodium phosphate, dibasic dehydrate; sodium phosphate, dibasic dodecahydrate; sodium phosphate; sodium phosphate dehydrate; sodium phosphate, monobasic, anhydrous; sodium phosphate, monobasic, dehydrate;

sodium phosphate, monobasic, monohydrate; dibasic potassium phosphate;

potassium phosphate, monobasic; sodium acetate; sodium acetate anhydrous;

ammonium acetate; sodium citrate; disodium hydrogen citrate; anhydrous trisodium citrate; disodium citrate sesqui hydrate; trisodium citrate dehydrate; sodium lactate;

(L)-sodium lactate; sodium tartrate; ammonium sulfate; and/or ethanolamine hydrochloride.

The pH adjusting agents used in the formulations include, but are not limited to sodium hydroxide; calcium hydroxide; potassium hydroxide; sodium bicarbonate; sodium carbonate; sodium carbonate decahydrate; sodium carbonate monohydrate; diethanolamine; meglumine; tromethamine; ammonia; hydrochloric acid; acetic acid; acetic anhydride; adipic acid; anhydrous citric acid; benzenesulfonic acid; boric acid; citric acid monohydrate; lactic acid; (DL)-lactic acid; (L)-lactic acid; maleic acid;

metaphosphoric acid; methanesulfonic acid; nitric acid; phosphoric acid; succinic acid; sulfuric acid; sulfurous acid; tartaric acid; (DL)-tartaric acid; and/or

trifluoroacetic acid.

Solvents used in the formulations include, but are not limited to water for injection (USP), propylene glycol, glycerin, and/or benzyl alcohol.

In another embodiment, the aqueous formulation comprises: (i) levothyroxine sodium

(ii) one or more antioxidants

(iii) one or more chelating agents

(iv) one or more stabilizing agents

(v) one or more buffering agents (vi) one or more pH adjusting agents and

(vii) one or more solvents

The levothyroxine may be the free base or may be a pharmaceutically acceptable salt thereof. In a preferred embodiment, the levothyroxine is levothyroxine sodium.

Antioxidants used in the formulations include, but are not limited to sodium sulfite, sodium bisulfite, sodium metabisulfite, potassium metabisulfite, alpha-tocopherol, acetone sodium bisulfite, ascorbic acid, sodium ascorbate, butylated hydroxyanisole, butylated hydroxytoluene, gentisic acid, gentisic ethanolamide, glutathione, methionine, monothioglycerol, and/or sodium formaldehyde sulfoxylate.

Chelating agents used in the formulations include, but are not limited to edetate disodium, edetate disodium anhydrous, edetate sodium, edetate calcium disodium, edetate calcium disodium anhydrous, edetic acid, anhydrous citric acid, citric acid monohydrate, gluceptate sodium, pentasodium pentetate, pentetate calcium trisodium, and/or pentetic acid. Stabilizing agents used in the formulations include, but are not limited to sodium iodide, potassium iodide, povidone, povidone K12, povidone K17, crospovidone, sorbitol, and/or sorbitol solution.

Buffering agents used in the formulations include, but are not limited to sodium phosphate, dibasic, heptahydrate; sodium phosphate, dibasic; sodium phosphate, dibasic, anhydrous; sodium phosphate, dibasic dehydrate; sodium phosphate, dibasic dodecahydrate; sodium phosphate; sodium phosphate dehydrate; sodium phosphate, monobasic, anhydrous; sodium phosphate, monobasic, dehydrate;

sodium phosphate, monobasic, monohydrate; dibasic potassium phosphate;

potassium phosphate, monobasic; sodium acetate; sodium acetate anhydrous; ammonium acetate; sodium citrate; disodium hydrogen citrate; anhydrous trisodium citrate; disodium citrate sesquihydrate; trisodium citrate dehydrate; sodium lactate; (L)-sodium lactate; sodium tartrate; ammonium sulfate; and/or ethanolamine hydrochloride. The pH adjusting agents used in the formulations include, but are not limited to sodium hydroxide; calcium hydroxide; potassium hydroxide; sodium bicarbonate; sodium carbonate; sodium carbonate decahydrate; sodium carbonate monohydrate; diethanolamine; meglumine; tromethamine; ammonia; hydrochloric acid; acetic acid; acetic anhydride; adipic acid; anhydrous citric acid; benzenesulfonic acid; boric acid; citric acid monohydrate; lactic acid; (DL)-lactic acid; (L)-lactic acid; maleic acid; metaphosphoric acid; methanesulfonic acid; nitric acid; phosphoric acid; succinic acid; sulfuric acid; sulfurous acid; tartaric acid; (DL)-tartaric acid; and/or

trifluoroacetic acid.

Solvents used in the formulations include, but are not limited to water for injection (USP), propylene glycol, glycerin, and/or benzyl alcohol.

In a preferred embodiment, the formulation is a parenteral formulation.

In another aspect, one or all of the composition invention embodiments are sterile.

In another aspect, one or all of the composition invention embodiments are free of particulate matter. In another aspect, one or all of the composition invention embodiments are packaged in an amber glass container. In another aspect, one or all of the composition invention embodiments are aseptically packaged in an amber glass container.

In another aspect, one or all of the composition invention embodiments are packaged in an amber glass container and terminally sterilized. The formulations described here can be prepared using conventional techniques known to the person skilled in the art. For example, the formulations may be prepared by dissolving levothyroxine or a pharmaceutically acceptable salt thereof in water, dissolving one or more pharmaceutically acceptable excipients, optionally adjusting the pH of the solution. Alternatively, the formulations may be prepared by dissolving one or more pharmaceutically acceptable excipients prior to dissolving the levothyroxine or pharmaceutically acceptable salt thereof, optionally adjusting the pH of the solution. The composition can be aseptically filled into a container that facilitates ready to administration. Alternatively, the composition can be filled into a container that facilitates ready to administration, and sterilizing said container.

Examples of processes to prepare an aqueous formulation (e.g., parenteral or oral) of levothyroxine or a pharmaceutically acceptable salt thereof include, but are not limited to, those described herein.

In various embodiments, when the formulation is stored at 60°C for a period of 40 days, less than 1 %, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 1 1 %, 12%, 13%, 14%, 15%, 20%, 25%, 30% or 35% of the levothyroxine sodium undergoes degradation. In a preferred embodiment, when the formulation is stored at 60°C for a period of 40 days, less than 5%, of the levothyroxine undergoes degradation. In other various embodiments after having been terminally sterilized at about 121 °C for about 30 minutes less than 1 %, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 1 1 %, 12%, 13%, 14%, 15%, 20%, 25%, 30% or 35% of the levothyroxine has undergone degradation. In a preferred embodiment, after having been terminally sterilized at about 121 °C for about 30 minutes less than 5% of the levothyroxine has undergone degradation. Such stability characteristics allow the formulations to be stored as aqueous formulations for immediate administration to a patient in need thereof. Such formulations are particularly advantageous in emergency situations, because they are in a ready-to-administer parenteral or oral solution. Such solutions save time in emergency situations because the levothyroxine does not have to be reconstituted, there is no concern whether the drug has been reconstituted correctly and aseptically in the emergency situation. These characteristics improve the safety of patients in need of levothyroxine, particularly in emergency situations.

The formulations can be provided in various dosage strengths. In some

embodiments, the dosage strength is 10 mcg/mL, 15 mcg/ml_, 20 mcg/mL, 25 mcg/mL, 30mcg/ml_, 35 mcg/mL, 40 mcg/mL, 45 mcg/mL, 50 mcg/mL, 55 mcg/mL, 60 mcg/mL, 65 mcg/mL, 70 mcg/mL, 75 mcg/mL, 80 mcg/mL, 85 mcg/mL, 90 mcg/mL, 95 mcg/mL, 100 mcg/mL, 150 mcg/mL, or 200 mcg/mL of levothyroxine (e.g., levothyroxine sodium) in an IV direct inject or oral solution. Preferably, the formulation is provided at 20mcg/mL, 40mcg/mL, or 100mcg/mL, of levothyroxine (e.g., levothyroxine sodium) in an IV direct inject solution, which will provide a dosing regimen of 300-500 meg initial (e.g., loading) dose followed by 50- 1 10 meg daily (e.g. , maintenance) dose.

The formulations may be packaged in a storage container standardly used for packaging pharmaceuticals (e.g., sterile packaging), particularly liquid formulations. For example, the storage contain may be a glass or plastic vial or ampoule (e.g. , 2ml, 4mL or 6mL volume). In some embodiments, the packaging protects its contents from light. In a preferred embodiment, the vial is an amber glass vial.

Preferably the packaging includes a closure system that is compatible for storage and transport of the formulation.

Suitable caps include those from West Pharmaceutical (size 13mm; sample ID: I3FO LQ LGTE (6B) 767 RED MT STEAM RU/RP; Formula: IP, STM 13FO LNG TE (6B) 3767 RED MATTE). Suitable stoppers include those from West Pharmaceutical (size 13mm; sample ID: 13mm Serum Novapure V-35 4031/45 or RP S2-F451 4432/50 G; Formula: 13mm Serum NovaPure (Bromobutyl/Chlorobutyl) Stopper). Suitable vials include those from Schott (size: 2mL, 4mL, 6mL; sample ID: Fiolax amber Type I glass vial/ Schott; Formula: Type I Amber glass).

Methods of Treatment

The aqueous levothyroxine formulations disclosed herein may be used to treat a disease or condition that is treatable using other levothyroxine formulations. Such diseases include, but are not limited to, myxedema coma. To treat myxedema coma, an initial intravenous loading dose of the levothyroxine formulation may be about 30C^g to about 50C^g followed by once daily intravenous maintenance doses of between about 50 and about 100μg should be administered to the subject. The aqueous levothyroxine formulations disclosed herein may be used for replacement or supplemental therapy in primary, secondary or tertiary

hypothyroidism.

The aqueous levothyroxine formulations of the present invention may be used for severe hypothyroid subjects or subjects with myxedema coma requiring

hospitalization. The aqueous parenteral formulations of the present invention may be administered intravenously to provide an initial loading dose of about 300 to about 500 μg of levothyroxine sodium to the subject. The aqueous parenteral levothyroxine formulation may be administered intravenously to provide a maintenance daily dose of about 50 to about 100 μg of levothyroxine sodium to the subject. The dose(s) of the disclosed levothyroxine formulations for treating a subject with one or more of the above-described diseases or conditions will depend on the mode of delivery and is within the skill of the skilled person to determine.

When used orally, the formulation may comprise one or more flavoring, sweetening or taste-making agents, known to those skilled in the art. The correct dose to be administered to the subject should be aseptically withdrawn from the vial and inspected visually for particulate matter and discoloration prior to administration into the subject.

EXAMPLES

Example 1

Water for injection qs to 1 mL

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 10.5±0.5. Benzyl alcohol was added to the above solution and mixed. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 10.5±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers.

Example 2

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 10.5±0.5. Benzyl alcohol was added to the above solution and mixed. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 10.5±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers.

Example 3

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 10.5±0.5. Monothioglycerol was added to the above solution and mixed. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 10.5±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers.

Example 4

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 1 1 .0±0.5. Sodium sulfite was added to the above solution and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 1 1 .0±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers.

Example 5

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 12.0±0.5. Monothioglycerol was added to the above solution and mixed. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 12.0±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers.

Example 6

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 10.5±0.5. Sodium sulfite was added to the above solution and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 10.5±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers. Example 7

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 1 1 .0±0.5. Monothioglycerol was added to the above solution and mixed. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 1 1 .0±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers.

Example 8

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 12.0±0.5. Sodium sulfite was added to the above solution and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 12.0±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers. Example 9

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 10.5±0.5. Monothioglycerol was added to the above solution and mixed. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 10.5±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers.

Example 10

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 1 1 .0±0.5. Sodium sulfite was added to the above solution and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 1 1 .0±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers. Example 1 1

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 12.0±0.5. Monothioglycerol was added to the above solution and mixed. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 12.0±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers.

Example 12

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 10.5±0.5. Edetate disodium was added to the above solution and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 10.5±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers. Example 13

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 1 1 .0±0.5. Edetate disodium was added to the above solution and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 1 1 .0±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers.

Example 14

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 12.0±0.5. Edetate disodium was added to the above solution and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 12.0±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers. Example 15

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 1 1 .0±0.5. Sodium iodide was added to the above solution and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 1 1 .0±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers.

Example 16

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 12.0±0.5. Sodium iodide was added to the above solution and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 12.0±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers. Example 17

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 10.5±0.5. Sodium iodide was added to the above solution and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 10.5±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers.

Example 18

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 1 1 .0±0.5. Sodium iodide was added to the above solution and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 1 1 .0±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers. Example 19

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 10.5±0.5. Sodium iodide was added to the above solution and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 10.5±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers.

Example 20

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 10.5±0.5. Sodium iodide was added to the above solution and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 10.5±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers. Example 21

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 10.5±0.5. Propylene glycol was added to the above solution and mixed. Levothyroxine sodium was added to the above solution and mixed until dissolved. The solution was filtered and filled into containers.

Example 22

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 1 1 .0±0.5. Propylene glycol was added to the above solution and mixed. Levothyroxine sodium was added to the above solution and mixed until dissolved. The solution was filtered and filled into containers.

Example 23

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 12.0±0.5. Propylene glycol was added to the above solution and mixed. Levothyroxine sodium was added to the above solution and mixed until dissolved. The solution was filtered and filled into containers.

Example 24

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 1 1 .0±0.5. Glycerin was added to the above solution and mixed. Levothyroxine sodium was added to the above solution and mixed until dissolved. The solution was filtered and filled into containers. Example 25

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 12.0±0.5. Glycerin was added to the above solution and mixed. Levothyroxine sodium was added to the above solution and mixed until dissolved. The solution was filtered and filled into containers. Example 26

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 10.5±0.5. Glycerin was added to the above solution and mixed.

Levothyroxine sodium was added to the above solution and mixed until dissolved. The solution was filtered and filled into containers.

All samples of Examples 1 - 26 were tested for osmolarity, pH, appearance and potency of levothyroxine sodium. All samples of Examples 1 - 26 were placed in a 50°C oven, and potency of levothyroxine sodium was tested after 3 days (see Table 1 ).

To analyze the potency of levothyroxine sodium that can be present in an illustrative formulation of levothyroxine sodium, gradient assay HPLC method was employed to separate levothyroxine from formulation components. The HPLC column is Zorbax SB-C18, 4.6x150 mm, 5 μιη. Column temperature is ambient room temperature. Flow rate was 1 .0 mL/min. Levothyroxine was detected in the eluent using UV absorbance. UV Detector wavelength was set at 225 nm. Injection volume was 5 μί for assay determination.

The mobile phase used for the gradient assay HPLC method was a mixture of acetonitrile and sulfamic acid buffer adjusted to pH 2.0 with sodium hydroxide. The diluent for HPLC analysis had same composition as levothyroxine sodium

formulation and it was dibasic sodium phosphate heptahydrate having a pH of 10.5..

The levothyroxine stock standard solution was 0.1 mg/mL of USP levothyroxine reference standard in the diluent. The sample solutions were injected onto HPLC without further dilution or

preparation.

The concentration of levothyroxine in the sample was determined by the external standard calibration method, where the peak area of levothyroxine in sample injections was compared to the peak area of levothyroxine reference standards in a solution of known concentration.

To calculate the potency of levothyroxine sodium (or Assay %) in the sample solutions, the following formula was used:

Result = {ru/rs) * {Cs/Cu) * 100

Where:

ru = peak response of levothyroxine from the Sample solution

rs = peak response of levothyroxine from the Standard solution

Cs = concentration of USP Levothyroxine RS in the Standard solution ( Q/mL)

Cu = concentration of Levothyroxine Sodium in the Sample solution ^g/mL)

Table 1 : Examples 1 to 26 - Stability Results

10 241 10.89 Clear 94.0 100.1

1 1 338 10.00 Cloudy 93.3 NA

12 68 7.91 Cloudy 94.3 NA

13 104 10.82 Clear 94.6 95.3

14 232 1 1 .67 Clear 95.3 94.0

15 84 10.58 Clear 92.9 95.4

16 374 1 1 .71 Clear 93.0 NA

17 129 10.29 Cloudy 91 .2 98.3

18 404 10.65 Cloudy 77.3 NA

19 219 10.43 Cloudy 95.7 94.7

20 495 10.31 Cloudy 93.0 NA

Out of

21 NA Clear 96.3 98.2 range

Out of NA

22 Clear 95.9 98.3 range

Out of NA

23 Clear 89.9 98.9 range

24 635 NA Clear 101 .5 78.5

25 707 NA Clear 96.1 97.7

26 795 NA Clear 92.0 90.8

NA - Not tested

Primary observations from the above analyses include the following: Levothyroxine was light sensitive. Monothioglycerol caused the pH to drop in the formulation, resulting in precipitation of levothyroxine sodium. EDTA also caused the pH to drop and precipitation of levothyroxine. Sodium sulfite didn't change the pH or osmolality significantly an appeared to provide stabilizing activity to the formulation. Sodium iodide also did not change the pH and osmolality. Levothyroxine sodium did not degrade whenever Nal was present in the solution. Example 27

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 10.5±0.5. Sodium sulfite was added to the above solution and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 10.5±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers.

Example 28

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 12.0±0.5. Sodium sulfite was added to the above solution and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 12.0±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers. Example 29

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 10.5±0.5. Sodium sulfite was added to the above solution and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 10.5±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers.

Example 30

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 12.0±0.5. Sodium sulfite was added to the above solution and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 12.0±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers. Exam le 31

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 10.5±0.5. Sodium sulfite was added to the above solution and mixed until dissolved. Edetate disodium was added to the above solution and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 10.5±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers. Example 32

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 10.5±0.5. Sodium sulfite was added to the above solution and mixed until dissolved. Sodium iodide was added to the above solution and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 1 0.5±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers.

Example 33

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 12.0±0.5. Sodium sulfite was added to the above solution and mixed until dissolved. Edetate disodium was added to the above solution and mixed until dissolved. Sodium iodide was added to the above solution and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 12.0±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers.

Example 34

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 12.0±0.5. Sodium sulfite was added to the above solution and mixed until dissolved. Benzyl alcohol was added to the above solution and mixed.

Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 12.0±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers.

Example 35

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 10.5±0.5. Sodium sulfite was added to the above solution and mixed until dissolved. Edetate disodium was added to the above solution and mixed until dissolved. Sodium iodide was added to the above solution and mixed until dissolved Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 10.5±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers. Exam le 36

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 10.5±0.5. Sodium sulfite was added to the above solution and mixed until dissolved. Benzyl alcohol was added to the above solution and mixed.

Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 10.5±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers. Example 37

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 12.0±0.5. Sodium sulfite was added to the above solution and mixed until dissolved. Edetate disodium was added to the above solution and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 12.0±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers.

Example 38

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 12.0±0.5. Sodium sulfite was added to the above solution and mixed until dissolved. Sodium iodide was added to the above solution and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 12.0±0.5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers.

Example 39

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 10.5±0.5. Sodium sulfite was added to the above solution and mixed until dissolved. Edetate disodium was added to the above solution and mixed until dissolved. Propylene glycol was added to the above solution and mixed.

Levothyroxine sodium was added to the above solution and mixed until dissolved. The solution was filtered and filled into containers.

Example 40

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 10.5±0.5. Sodium sulfite was added to the above solution and mixed until dissolved. Sodium iodide was added to the above solution and mixed until dissolved. Propylene glycol was added to the above solution and mixed.

Levothyroxine sodium was added to the above solution and mixed until dissolved. The solution was filtered and filled into containers.

Exam le 41

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 12.0±0.5. Sodium sulfite was added to the above solution and mixed until dissolved. Edetate disodium was added to the above solution and mixed until dissolved. Sodium iodide was added to the above solution and mixed until dissolved. Propylene glycol was added to the above solution and mixed. Levothyroxine sodium was added to the above solution and mixed until dissolved. The solution was filtered and filled into containers.

Example 42

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 12.0±0.5. Sodium sulfite was added to the above solution and mixed until dissolved. Propylene glycol was added to the above solution and mixed. Benzyl alcohol was added to the above solution and mixed. Levothyroxine sodium was added to the above solution and mixed until dissolved. The solution was filtered and filled into containers.

Example 43

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 10.5±0.5. Sodium sulfite was added to the above solution and mixed until dissolved. Edetate disodium was added to the above solution and mixed until dissolved. Sodium iodide was added to the above solution and mixed until dissolved. Propylene glycol was added to the above solution and mixed. Levothyroxine sodium was added to the above solution and mixed until dissolved. The solution was filtered and filled into containers. Exam le 44

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 10.5±0.5. Sodium sulfite was added to the above solution and mixed until dissolved. Propylene glycol was added to the above solution and mixed. Benzyl alcohol was added to the above solution and mixed. Levothyroxine sodium was added to the above solution and mixed until dissolved. The solution was filtered and filled into containers. Example 45

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 12.0±0.5. Sodium sulfite was added to the above solution and mixed until dissolved. Edetate disodium was added to the above solution and mixed until dissolved. Propylene glycol was added to the above solution and mixed.

Levothyroxine sodium was added to the above solution and mixed until dissolved. The solution was filtered and filled into containers.

Example 46

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 12.0±0.5. Sodium sulfite was added to the above solution and mixed until dissolved. Sodium iodide was added to the above solution and mixed until dissolved. Propylene glycol was added to the above solution and mixed.

Levothyroxine sodium was added to the above solution and mixed until dissolved. The solution was filtered and filled into containers. All samples of Examples 27 - 46 were tested for osmolarity, pH, appearance and potency of levothyroxine sodium (see Table 2). All samples of Examples 27 - 46 were placed in the following storage conditions: ambient room temperature under ambient light in clear vials; 2-8°C under refrigeration in amber vials; 60°C/ambient RH in amber vials. The potency of levothyroxine sodium (Assay %, described above) and appearance were tested at selected stability storage time point (see Tables 3 - 5). Table 2: Examples 27 - 46 Test Results at Initial Time

Table 3: Examples 27 - 46 Stability Results (ambient temperature under ambient light in clear vials)

NA - Not tested Table 4: Examples 27 - 46 Stability Results (2-8°C under refrigeration in amber vials)

Table 5: Examples 27 - 46 Stability Results (60°C/ambient RH in amber vials)

NA - Not tested

Based on the above analyses, a Prototype I study was initiated. Six formulations (see Examples 47-52) were prepared at 100 mL scale. Each sample was filled with 1 mL of fill volume of solution into 2 mL Type I Clear Glass Vials and then stoppered to represent a worst-case scenario with respect to the process of parameters (e.g., head space oxygen, light). Storage conditions were 60°C.

Example 47

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 9.5 - 1 1 .5. Sodium sulfite was added to the above buffer and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 9.5 - 1 1 .5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers. Example 48

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 9.5 - 1 1 .5. Sodium sulfite was added to the above buffer and mixed until dissolved. Propylene glycol was added to the above solution and mixed.

Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 9.5 - 1 1 .5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers.

Example 49

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 9.5 - 1 1 .5. Sodium sulfite and edetate disodium were added to the above buffer and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 9.5 - 1 1 .5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers.

Example 50

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 9.5 - 1 1 .5. Sodium sulfite, edetate disodium and sodium iodide were added to the above buffer and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 9.5 - 1 1 .5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers. Example 51

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 9.5 - 1 1 .5. Sodium sulfite and edetate disodium were added to the above buffer and mixed until dissolved. Propylene glycol was added to the above solution and mixed. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 9.5 - 1 1 .5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers.

Example 52

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 9.5 - 1 1 .5. Sodium sulfite, edetate disodium and sodium iodide were added to the above buffer and mixed until dissolved. Propylene glycol was added to the above solution and mixed. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 9.5 - 1 1 .5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers.

The samples of Examples 47 - 52 were placed in 121 °C oven for 30min, which simulated heat sterilization conditions, in both upright and inverted orientations. The potency (Assay %, described above) and impurity of levothyroxine sodium were tested (see Table 6). To analyze the impurities of levothyroxine sodium that can be present in an illustrative formulation of levothyroxine sodium, gradient HPLC method was employed to separate levothyroxine and impurities from formulation components. The gradient HPLC parameters and gradient table are depicted as follows:

The mobile phase A used for the gradient HPLC method was prepared by dissolving 9.7 g of sulfamic acid in 2000 mL of water, followed by adding 1 .5 g of sodium hydroxide and mixing to dissolve. Mobile phase A was adjusted to pH of 2.0 with 2N sodium hydroxide. Mobile phase B is 100% acetonitrile.

Diluent was dibasic sodium phosphate heptahydrate adjusted to pH 10.5. The levothyroxine stock standard solution was 0.1 mg/mL of USP levothyroxine reference standard in the Diluent. To prepare levothyroxine stock standard solution, accurately weighed and transferred about 1 1 mg of levothyroxine USP reference standard into a 100 mL volumetric flask. Dissolved and diluted to volume with Diluent and mixed well.

The levothyroxine working standard solution was 0.001 mg/mL of USP levothyroxine reference standard. To prepare levothyroxine working standard solution, pipetted 2.0 mL of the levothyroxine stock standard solution to a 200 mL volumetric flask and diluted to volume with Diluent. Mixed well.

The sensitivity and identification standard solution was 0.1 μg/mL of USP

levothyroxine reference standard and liothyronine reference standard.

The sample solutions were injected onto HPLC without further dilution or

preparation. Injection volume for both levothyroxine working standard solution and sample solution was 50 μί.

Diluent was injected as blank injection.

The concentration of levothyroxine in the sample was determined by the external standard calibration method, where the peak area of impurity peaks in sample injections was compared to the peak area of levothyroxine reference standard in a solution of known concentration.

To calculate the percentage of liothyronine sodium (T3 %) in the sample solutions, the following formula was used:

Result = (ru/rs) * (Cs/Cu) * * 100

Where:

ru = peak response of liothyronine from the Sample solution

rs = peak response of levothyroxine from the Standard solution

Cs = concentration of USP levothyroxine RS in the Standard solution (Mg/mL)

Cu = concentration of Levothyroxine Sodium in the Sample solution ( g/mL)

To calculate the percentage of any other impurity (Impurity %) in the sample solutions, the following formula was used:

Result = {ru/rs) * {Cs/Cu) * 100 Where:

ru = peak response of any impurity from the Sample solution

rs = peak response of levothyroxine from the Standard solution

Cs = concentration of USP Levothyroxine RS in the Standard solution

(MQ/mL)

Cu = concentration of Levothyroxine Sodium in the Sample solution ^g/mL)

Any peaks corresponding to those of the blank solution were disregarded. Any peaks corresponding to less than 0.03% were disregarded. To calculate the total impurity, added Impurity % from all impurity peaks.

Table 6: Stability of Samples of Examples 47 - 52 in simulated heat sterilization condition

The samples of Examples 47 - 52 were placed in 60°C oven for about 36 days. The potency and impurity of levothyroxine sodium were tested (see Table 7). Table 7: Stability results of samples of Examples 47- 52 in 60°C oven

EDTA and Nal positively contributed to the stability of the formulation. Formulations containing propylene glycol showed more impurities compared to formulation the did not contain propylene glycol.

Based on the above studies, a Prototype II study was initiated, to further address qualitative and quantitative aspects of the formulation. Six formulations (see Examples 47-52) were prepared at 100 mL scale. Each sample was filled with 1 mL of fill volume of solution into 2 mL Type I Clear Glass Vials and then stoppered to represent a worst-case scenario with respect to the process of parameters (e.g., head space oxygen, light). Storage conditions were 60°C.

Example 53

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 9.5 - 1 1 .5. Sodium sulfite and edetate disodium were added to the above buffer and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 9.5 - 1 1 .5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers.

Example 54

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 9.5 - 1 1 .5. Sodium sulfite and sodium iodide were added to the above buffer and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 9.5 - 1 1 .5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers. Example 55

Water for injection qs to 1 ml_

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 9.5 - 1 1 .5. Sodium sulfite, edetate disodium and sodium iodide were added to the above buffer and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 9.5 - 1 1 .5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers.

Example 56

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 9.5 - 1 1 .5. Sodium sulfite, edetate disodium and sodium iodide were added to the above buffer and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 9.5 - 1 1 .5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers. Example 57

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 9.5 - 1 1 .5. Sodium sulfite, edetate disodium and sodium iodide were added to the above buffer and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 9.5 - 1 1 .5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers. Example 58

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 9.5 - 1 1 .5. Sodium sulfite, edetate disodium and sodium iodide were added to the above buffer and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 9.5 - 1 1 .5 by addition of sodium hydroxide if necessary. The solution was filtered and filled into containers.

The samples of Examples 53 - 58 were stored in 2ml_ Type I Clear Glass Vials and placed in 60°C oven for about 40 days. All storage conditions were void of light exposure. Vials were stored in both upright and inverted orientations. The osmolarity, pH, appearance, potency and impurity of levothyroxine sodium were tested (as described herein) at selected stability storage time point (see Table 8).

Table 8: Stability results of samples of Examples 53 - 58 in 60°C oven

Table 8: Stability results of samples of Examples 53 - 58 in 60°C oven (cont'd)

No impurities were observed from a placebo sample (containing 1 .8 mg sodium sulfite; 10 mg sodium iodide; 1 mg EDTA, 10.72 mg sodium phosphate, dibasic, heptahydrate; sodium (qs); and water for injection (qs to 1 ml_)). The presence of 0.1 % EDTA, without sodium iodide, and the associated stability profile as seen in Example 53 demonstrates that this excipient has a stabilizing effect alone or in combination with sodium sulfite.

Example 59 Multivariate Statistical Design:

Formulation development and R&D Scale-up data indicates there is an underlying interaction within the formulation with the three key excipients; sodium iodide, disodium edetate and sodium sulfite. A multivariate statistical design studies was carried out as two sets of experimental trials: 2 ³ Full Factorial DoE Study #1 and 2 ³ Full Factorial DoE Study #2.

All the individual formulations/trials of each of the DoE were packaged in 2ml_ USP Type I amber glass, stoppered with 'Flurotec' coated 'Chlorobutyl' rubber stoppers (for improved compatibility with a high pH of 10.5 as recommended from the supplier) and subjected to short-term stress stability at 60°C (Inverted orientation) for a period of 4 weeks (see Table 9).

2 ³ Full Factorial DoE Study #1:

DoE #1 was carried out to evaluate/identify if an interaction effect is present within the formulation variables.

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 9.5 - 1 1 .5. Sodium sulfite, edetate disodium and sodium iodide were added to the above buffer and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 9.5 - 1 1 .5 by addition of sodium hydroxide if necessary. The solution was filled into containers.

Table 9: Stability results of samples of Example 59 Initial and 60°C oven

Example 60

2 ³ Full Factorial DoE Study #2:

The goal of DoE study #2 was to further optimize the composition of sodium iodide, sodium sulfite and disodium edetate as per the results of DoE study #1 . The concentration levels of three excipients (sodium iodide, sodium sulfite and disodium edetate) were different from that of DoE #1 . Sodium iodide was evaluated at higher levels, while sodium sulfite and disodium edetate were evaluated at lower levels (see Table 10).

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 9.5 - 1 1 .5. Sodium sulfite, edetate disodium and sodium iodide were added to the above buffer and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 9.5 - 1 1 .5 by addition of sodium hydroxide if necessary. The solution was filled into containers.

Table 10: Stability results of samples of Example 60 Initial and 60°C oven

After 4 weeks of storing samples at 60°C stability conditions, all samples exhibited >90% assay, <0.15% liothyronine and <0.61 % total impurity levels. Example 61

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 9.5 - 1 1 .5. Sodium sulfite, edetate disodium and sodium iodide were added to the above buffer and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 9.5 - 1 1 .5 by addition of sodium hydroxide if necessary. The solution was filled into containers. Stability of the samples was evaluated initially and after 1 or 2 weeks storage in a 60°C oven (see Table 1 1 ).

Table 11 : Stability results of sample of Example 61 Initial and 60°C oven

Time

Strength Test

Initial 1Wk 2Wk

Assay % 101.7 100.7 99.3

40 μ9/πιΙ_ Liothyronine % ND 0.08 0.1 1

Total Impurities % ND 0.15 0.18

After 2 weeks of storing samples at 60°C stability conditions, all samples

exhibited >90% assay, <0.1 1 % liothyronine and <0.18% total impurity levels.

Example 62

To a processing vessel, water for injection was added. Disodium phosphate, heptahydrate and sodium hydroxide were added and dissolved. The pH was maintained at 9.5 - 1 1 .5. Sodium sulfite, edetate disodium and sodium iodide were added to the above buffer and mixed until dissolved. Levothyroxine sodium was added to the above solution and mixed until dissolved. The pH of the solution was adjusted to 9.5 - 1 1 .5 by addition of sodium hydroxide if necessary. The solution was filled into containers. Stability of the samples was evaluated initially and after 1 or 2 weeks storage in a 60°C oven (see Table 12).

Table 12: Stability results of sample of Example 62 Initial and 60°C oven

After 2 weeks of storing samples at 60 degrees stability conditions, all samples exhibited >90% assay, <0.16% liothyronine and <0.32% total impurity levels.

The results of the experiments described herein show, inter alia, that levothyroxine is stabilized with the use of formulations comprising sodium iodide, sodium sulfite and disodium edetate in combination and at certain ratios. Example 63

A formulation ("Control Formulation") containing: 20 mcg/mL Levothyroxine sodium, USP; 6.48 mg/mL sodium chloride; 0.14 mg/mL sodium iodide; 10 mg/ml

tromethamine, USP; sodium hydroxide (1 N) and hydrochloric acid as needed to adjust pH to 10-10.5 and purified water (q.s.). The formulation of Example 60, Trial 8 ("Formulation E60/T8") was also prepared. The solution was filled into containers. Stability of the samples was evaluated, as described herein, initially and after 4 weeks storage in a 60°C oven.

Table 13: Stability results of samples of Example 63 Initial and 60°C oven

Formulation E60/T8 exhibited superior stability compared to the Control Formulation, and had an Assay % within the USP acceptable range of 90% - 1 10% and low impurity levels at elevated temperatures of 60°C for 4 weeks. Even under these high stress conditions, Formulation E60/T8 is robust.

It is to be understood that the invention embodiments are not limited to the particular structures, process steps, or materials disclosed herein, but are extended to equivalents thereof as would be recognized by those ordinarily skilled in the relevant arts. It should also be understood that terminology employed herein is used for the purpose of describing particular embodiments only and is not intended to be limiting.