Liquid formulations of amylin analogues

Abstract

The present invention relates to formulations of amylin analogues and their use in the treatment of metabolic disorders such as obesity and diabetes. In particular, the present invention relates to stable aqueous liquid formulations of amylin analogues.

Description

Technical Field

[0001] The present invention relates to formulations of amylin analogs and their use in the treatment of metabolic disorders such as obesity and diabetes. In particular, the present invention relates to stable aqueous liquid formulations of amylin analogs.

Background Art

[0002] Amylin is one of a family of peptide hormones that includes amylin, calcitonin, calcitonin gene-related peptide, adrenomedullin, and intermedin (intermedin is also known as AFP-6) and has been thought to be associated with various metabolic diseases and disorders. Human amylin was first isolated, purified, and characterized as the major component of amyloid deposits in the pancreatic islets of patients with type 2 diabetes.

[0003] Native human amylin is a 37-amino acid peptide having the formula H-KC()NTATC()ATQRLANFLVHSSNNFGAILSSTNVGSNTY-NH 2 wherein H at the N-terminus designates a hydrogen atom and corresponds to the presence of a free amino group on the N-terminal amino acid residue (i.e., the lysine (K) residue at sequence position 1 in the sequence shown above), and -NH at the C-terminus indicates that the C-terminal carboxyl group is in amide form, and the parentheses "()" associated with the two cysteine (C, Cys) residues at sequence positions 2 and 7 indicate the presence of an intramolecular disulfide bridge between the two Cys residues in question. 2

[0004] ​Amylin is thought to be beneficial in the treatment of metabolic disorders such as diabetes and / or obesity. Amylin is thought to regulate gastric emptying, suppress glucagon secretion and food intake, thereby regulating the rate of glucose release into the circulation. Amylin appears to complement the action of insulin. Type 1 diabetic patients have no circulating amylin, and type 2 diabetic patients exhibit a reduction in postprandial amylin concentration compared to healthy adults. In human studies, an amylin analog known as pramlintide, described in WO93 / 10146 and having the sequence Lys-Cys-Asn-Thr-Ala-Thr-Cys-Ala-Thr-Gln-Arg-Leu-Ala-Asn-Phe-Leu-Val-His-Ser-Ser-Asn-Asn-Phe-Gly-Pro-Ile-Leu-Pro-Pro-Thr-Asn-Val-Gly-Ser-Asn-Thr-Tyr, also has a disulfide bridge between the Cys residues at positions 2 and 7 and has been shown to reduce body weight or reduce weight gain.

[0005] An alternative amylin analog designated IAPP-GI, which incorporates N-methylated residues and has a reduced tendency to fibrillate, is described by Yan et al. (PNAS, 103(7), pp. 2046-2051, 2006; Angew. Chem. Int. Ed. 2013, 52, pp. 10378-10383; WO2006 / 042745). However, IAPP-GI appears to have lower activity than native amylin.

[0006] WO2018 / 046719 describes amylin analogs that have, among other things, lactam crosslinks instead of disulfide crosslinks, N-methylated residues, and deletions corresponding to the residues Asn21 and Asn22 of native human amylin. Such analogs have a considerably lower tendency towards fibrillation than native amylin while also having a higher potency than the analogs described by Yan et al. (supra). These are typically easy to formulate at or near physiological pH. However, there is a need for the development of improved formulations of these analogs, particularly for providing stable formulations that can be stored long-term without causing excessive fibrillation or degradation of the active monomeric form of the peptide.

Prior Art Documents

Patent Documents

[0007]

Patent Document 1

Patent Document 2

Patent Document 3

Non-Patent Documents

[0008]

Non-Patent Document 1

Non-Patent Document 2

Non-Patent Document 3

Non-Patent Document 4

Non-Patent Document 5

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Non - Patent Document 9

Summary of the Invention

Means for Solving the Problems

[0009] Generally, the present invention is based on the research described in the examples, and such research has led to surprising findings regarding aqueous formulations of amylin analogs that are particularly suitable for long - term storage. In particular, it has been found that low buffer concentrations and a specific pH range are unexpectedly significant for optimizing stability.

[0010] The present invention provides [19CD]-isoGlu - RD()GTATK()ATERLA - Aad - FLQRSSF - Gly(Me)-A - Ile(Me)-LSSTEVGSNT - Hyp - NH 2 a stable aqueous liquid pharmaceutical formulation comprising an amylin analog or a pharmaceutically acceptable salt and / or derivative thereof wherein the formulation comprises (a) an amylin analog at a concentration of about 0.4 mg / ml to about 25 mg / ml, and (b) a buffer at a concentration of about 0.5 mM to about 25 mM comprising the formulation has a pH of about 5.8 to about 6.9.

[0011] The amylin analog is present at a concentration of about 0.4 mg / ml to about 25 mg / ml.

[0012] In some embodiments, the amylin analog is present at a concentration of at least about 0.4 mg / ml, at least about 0.5 mg / ml, at least about 0.6 mg / ml, at least about 1.2 mg / ml, or at least about 2.5 mg / ml.

[0013] The amylin analog may be present up to about 25 mg / ml, up to about 20 mg / ml, up to about 15 mg / ml, or up to about 10 mg / ml.

[0014] The amylin analog may be present at a concentration of about 3 mg / ml to about 12 mg / ml, such as about 3 mg / ml to about 7 mg / ml, or about 8 mg / ml to about 12 mg / ml, such as about 5 mg / ml or about 10 mg / ml.

[0015] Any suitable buffer including phosphate buffer, histidine buffer, TRIS buffer, acetate buffer, arginine buffer, citrate buffer, bicarbonate buffer, diethanolamine buffer, lysine buffer, TAPS buffer, succinate buffer, malate buffer, and α-ketoglutarate buffer may be used. Particularly preferred buffers include phosphate buffer, histidine buffer, citrate buffer, and TRIS buffer, especially phosphate buffer and histidine buffer.

[0016] Surprisingly, it has been found that stability increases at relatively low buffer concentrations. That is, the buffer is present at a concentration of about 0.5 mM to about 25 mM.

[0017] The buffer may be present at a concentration of from about 0.5 mM to about 20 mM, such as from about 0.5 mM to about 15 mM, such as from about 1 mM to about 15 mM, such as from about 1 mM to about 12 mM. For example, particularly (but not limited to) when the buffer is phosphate, histidine, or citrate, it may be present at a concentration of from about 3 mM to about 7 mM, or from about 8 mM to about 12 mM, such as about 5 mM, or about 10 mM.

[0018] For TRIS buffer, since the buffering capacity of TRIS is lower than that of other buffers such as phosphate and histidine within the relevant pH range (for example), it may preferably be present at about 15 mM to about 25 mM, such as about 17 mM to about 23 mM, such as about 20 mM.

[0019] The pH of the formulation has also been found to be significant for stability. The compounds described in WO2018 / 046719 were thought to be stable at physiological pH (for example, approximately pH 7.4), but surprisingly, it has been found that the chemical stability increases at lower pH values while the physical stability is maintained. Thus, the formulation may have a pH of from about 5.8 to about 6.9, such as, for example, from about 6.2 to about 6.8, such as from about 6.4 to about 6.6, such as about 6.5.

[0020] The stable aqueous liquid formulation may contain a tonicity modifier. The tonicity modifier may be ionic or non-ionic. Suitable ionic tonicity modifiers include alkali metal salts (such as halides) and alkaline earth metal salts (such as halides), such as NaCl, NaBr, NaI, KCl, KBr, KI, LiCl, CaCl2, and Na 2 SO 4 is included. Non-ionic tonicity modifiers include mannitol (such as D-mannitol), propylene glycol, sucrose, glycerol, sorbitol, and trehalose.

[0021] In some embodiments, mannitol (such as D-mannitol), propylene glycol, and NaCl, particularly mannitol and propylene glycol, may be preferred as tonicity modifiers.

[0022] The aqueous liquid formulation is typically stable at 2 to 8 °C (e.g., 5 °C) for at least 6 months, such as at least 12 months, at least 18 months, or at least 24 months. Preferably, it is stable at 2 to 8 °C (e.g., 5 °C) for at least 18 months, such as at least 24 months.

[0023] For example, after storing the formulation at 2 to 8 °C (e.g., 5 °C) for at least 6 months, at least 12 months, at least 18 months, or at least 24 months, it does not substantially exhibit changes that are indicators of a decrease in physical stability (e.g., demonstrated by turbidity of the solution, or aggregation, fibrillation, or gelation of the amylin analog). Preferably, after storing the formulation at 2 to 8 °C (e.g., 5 °C) for at least 18 months, such as at least 24 months, it does not substantially exhibit such changes. As will be presented in detail below, it will be understood that accelerated conditions can also be employed as a surrogate for the evaluation of longer-term physical stability.

[0024] Further or alternatively, at least 80%, more preferably at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% of the amylin analog remains in its original monomeric form in the formulation after storing at 2 to 8 °C (e.g., 5 °C) for at least 6 months, at least 12 months, at least 18 months, or at least 24 months. Preferably, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% of the amylin analog remains in its original monomeric form after storing at 2 to 8 °C (e.g., 5 °C) for at least 6 months, at least 12 months, at least 18 months, or at least 24 months. More preferably, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% of the amylin analog remains in its original monomeric form in the formulation after storing at 2 to 8 °C (e.g., 5 °C) for at least 18 months, such as at least 24 months.

[0025] The formulation may further comprise a preservative, especially if it is provided in a multiple-use format. Alternatively, especially if it is provided in a single-use format, the formulation may not contain or may substantially not contain a preservative.

[0026] The stable aqueous liquid formulation of the present invention is formulated for parenteral administration, and may be especially formulated for administration to a subject by injection, for example subcutaneous injection.

[0027] In accordance with normal pharmaceutical practice, the formulations of the present invention are sterile and / or do not contain (or substantially do not contain, for example contain no more than about 100 ppm) a reducing agent.

[0028] The individual dosages of the formulations of the present invention may be provided in any suitable volume. For example, the individual dosages may have a volume of 0.1 - 1.5 ml, for example 0.3 - 1.5 ml, for example 0.3 ml - 1 ml. Examples of specific individual dosage volumes include 0.5 ml and 1 ml (for example 1.0 ml). As described in more detail below, the individual dosages may be separately packaged for single use, for example in individual vials, cartridges, or syringes.

[0029] By way of example, a particular formulation may comprise or consist of the following.

[0030] An amylin analog at a concentration of about 0.8 mg / ml, a phosphate buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0031] An amylin analog at a concentration of about 1.2 mg / ml, a phosphate buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0032] An amylin analog at a concentration of about 2.4 mg / ml, a phosphate buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0033] An amylin analog at a concentration of about 4.8 mg / ml, a phosphate buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0034] An amylin analog at a concentration of about 6.0 mg / ml, a phosphate buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0035] An amylin analog at a concentration of about 9.0 mg / ml, a phosphate buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0036] An amylin analog at a concentration of about 18 mg / ml, a phosphate buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0037] An amylin analog at a concentration of about 0.8 mg / ml, a phosphate buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0038] An amylin analog at a concentration of about 1.2 mg / ml, a phosphate buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0039] An amylin analog at a concentration of about 2.4 mg / ml, a phosphate buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0040] An amylin analog at a concentration of about 4.8 mg / ml, a phosphate buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0041] An amylin analog at a concentration of about 6.0 mg / ml, a phosphate buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0042] An amylin analog at a concentration of about 9.0 mg / ml, a phosphate buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0043] An amylin analog at a concentration of about 18 mg / ml, a phosphate buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0044] An amylin analog at a concentration of about 0.8 mg / ml, a histidine buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0045] An amylin analog at a concentration of about 1.2 mg / ml, a histidine buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0046] An amylin analog at a concentration of about 2.4 mg / ml, a histidine buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0047] An amylin analog at a concentration of about 4.8 mg / ml, a histidine buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0048] An amylin analog at a concentration of about 6.0 mg / ml, a histidine buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0049] An amylin analog at a concentration of about 9.0 mg / ml, a histidine buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0050] An amylin analog at a concentration of about 18 mg / ml, a histidine buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0051] An amylin analog at a concentration of approximately 0.8 mg / ml, a histidine buffer at a concentration of approximately 10 mM, and mannitol at a concentration of approximately 250 - 260 mM; pH 6.5;

[0052] An amylin analog at a concentration of approximately 1.2 mg / ml, a histidine buffer at a concentration of approximately 10 mM, and mannitol at a concentration of approximately 250 - 260 mM; pH 6.5;

[0053] An amylin analog at a concentration of approximately 2.4 mg / ml, a histidine buffer at a concentration of approximately 10 mM, and mannitol at a concentration of approximately 250 - 260 mM; pH 6.5;

[0054] An amylin analog at a concentration of approximately 4.8 mg / ml, a histidine buffer at a concentration of approximately 10 mM, and mannitol at a concentration of approximately 250 - 260 mM; pH 6.5;

[0055] An amylin analog at a concentration of approximately 6.0 mg / ml, a histidine buffer at a concentration of approximately 10 mM, and mannitol at a concentration of approximately 250 - 260 mM; pH 6.5;

[0056] An amylin analog at a concentration of approximately 9.0 mg / ml, a histidine buffer at a concentration of approximately 10 mM, and mannitol at a concentration of approximately 250 - 260 mM; pH 6.5;

[0057] An amylin analog at a concentration of approximately 18 mg / ml, a histidine buffer at a concentration of approximately 10 mM, and mannitol at a concentration of approximately 250 - 260 mM; pH 6.5;

[0058] An amylin analog at a concentration of approximately 0.8 mg / ml, a citrate buffer at a concentration of approximately 5 mM, and mannitol at a concentration of approximately 250 - 260 mM; pH 6.5;

[0059] An amylin analog at a concentration of approximately 1.2 mg / ml, a citrate buffer at a concentration of approximately 5 mM, and mannitol at a concentration of approximately 250 - 260 mM; pH 6.5;

[0060] An amylin analog at a concentration of about 2.4 mg / ml, a citrate buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0061] An amylin analog at a concentration of about 4.8 mg / ml, a citrate buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0062] An amylin analog at a concentration of about 6.0 mg / ml, a citrate buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0063] An amylin analog at a concentration of about 9.0 mg / ml, a citrate buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0064] An amylin analog at a concentration of about 18 mg / ml, a citrate buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0065] An amylin analog at a concentration of about 0.8 mg / ml, a citrate buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0066] An amylin analog at a concentration of about 1.2 mg / ml, a citrate buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0067] An amylin analog at a concentration of about 2.4 mg / ml, a citrate buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0068] An amylin analog at a concentration of about 4.8 mg / ml, a citrate buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0069] An amylin analog at a concentration of about 6.0 mg / ml, a citrate buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0070] An amylin analog at a concentration of about 9.0 mg / ml, a citrate buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0071] An amylin analog at a concentration of about 18 mg / ml, a citrate buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0072] These formulations may be particularly suitable for a dosing volume of 0.5 ml, but it will be understood that they may also be used with other dosing volumes. Alternatively, the concentration of the amylin analog may be adjusted according to various dosing volumes.

[0073] Certain further formulations may comprise or consist of the following.

[0074] An amylin analog at a concentration of about 0.4 mg / ml, a phosphate buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0075] An amylin analog at a concentration of about 0.6 mg / ml, a phosphate buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0076] An amylin analog at a concentration of about 1.2 mg / ml, a phosphate buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0077] An amylin analog at a concentration of about 2.4 mg / ml, a phosphate buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0078] An amylin analog at a concentration of about 4.5 mg / ml, a phosphate buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0079] An amylin analog at a concentration of about 9 mg / ml, a phosphate buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0080] An amylin analog at a concentration of about 0.4 mg / ml, a phosphate buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0081] An amylin analog at a concentration of about 0.6 mg / ml, a phosphate buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0082] An amylin analog at a concentration of about 1.2 mg / ml, a phosphate buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0083] An amylin analog at a concentration of about 2.4 mg / ml, a phosphate buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0084] An amylin analog at a concentration of about 4.5 mg / ml, a phosphate buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0085] An amylin analog at a concentration of about 9 mg / ml, a phosphate buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0086] An amylin analog at a concentration of about 0.4 mg / ml, a histidine buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0087] An amylin analog at a concentration of about 0.6 mg / ml, a histidine buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0088] An amylin analog at a concentration of about 1.2 mg / ml, a histidine buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0089] An amylin analog at a concentration of about 2.4 mg / ml, a histidine buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0090] An amylin analog at a concentration of about 4.5 mg / ml, a histidine buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0091] An amylin analog at a concentration of about 9 mg / ml, a histidine buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0092] An amylin analog at a concentration of about 0.4 mg / ml, a histidine buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0093] An amylin analog at a concentration of about 0.6 mg / ml, a histidine buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0094] An amylin analog at a concentration of about 1.2 mg / ml, a histidine buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0095] An amylin analog at a concentration of about 2.4 mg / ml, a histidine buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0096] An amylin analog at a concentration of about 4.5 mg / ml, a histidine buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0097] An amylin analog at a concentration of about 9 mg / ml, a histidine buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0098] An amylin analog at a concentration of about 0.4 mg / ml, a citrate buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0099] An amylin analog at a concentration of about 0.6 mg / ml, a citrate buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0100] An amylin analog at a concentration of about 1.2 mg / ml, a citrate buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0101] An amylin analog at a concentration of about 2.4 mg / ml, a citrate buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0102] An amylin analog at a concentration of about 4.5 mg / ml, a citrate buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0103] An amylin analog at a concentration of about 9 mg / ml, a citrate buffer at a concentration of about 5 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0104] An amylin analog at a concentration of about 0.4 mg / ml, a citrate buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0105] An amylin analog at a concentration of about 0.6 mg / ml, a citrate buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0106] An amylin analog at a concentration of about 1.2 mg / ml, a citrate buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0107] An amylin analog at a concentration of about 2.4 mg / ml, a citrate buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0108] An amylin analog at a concentration of about 4.5 mg / ml, a citrate buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0109] An amylin analog at a concentration of about 9 mg / ml, a citrate buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250 - 260 mM; pH 6.5;

[0110] These formulations may be particularly suitable for a dosing volume of 1.0 ml, but it will be understood that they may be used with other dosing volumes. Alternatively, the concentration of the amylin analog may be adjusted according to various dosing volumes.

[0111] As in other places in these formulations, the amylin analog may be, for example, of the following formula: ([19CD]-isoGlu-RD()GTATK()ATERLA-Aad-FLQRSSF-Gly(Me)-A-Ile(Me)-LSSTEVGSNT-Hyp-NH 2 ), x(Cl) and may be provided as a chloride salt where x is from 1.0 to 2.0.

[0112] The present invention further provides a container or delivery device comprising the stable aqueous liquid formulation described herein. By way of example, suitable containers and delivery devices include sealed vials, prefilled syringes, or cartridges for injection devices such as injection pens, dose-adjustable autoinjectors, disposable autoinjectors, wearable syringes, or infusion pumps. The delivery device may be a single-use device such as a prefilled syringe.

[0113] In a further aspect, the present invention provides a manufactured article or kit comprising a container holding the stable aqueous liquid pharmaceutical composition of the present invention.

[0114] The present invention further provides the described stable aqueous liquid formulation for use in a method of medical treatment.

[0115] The formulation is particularly useful in reducing food intake, promoting weight loss, and preventing or reducing weight gain. As a result, the formulation may be used for the treatment of various conditions, diseases, or disorders in a subject, including but not limited to obesity and various conditions, diseases, or disorders associated with obesity, such as diabetes (e.g., type 2 diabetes), hypertension, dyslipidemia, sleep apnea, and cardiovascular diseases. The subject may be afflicted with obesity associated with at least one weight-related co-morbid condition, such as diabetes (e.g., type 2 diabetes), hypertension, dyslipidemia, sleep apnea, and cardiovascular diseases. Thus, it will be understood that the formulation may be administered to a subject afflicted with a condition characterized by inappropriate appetite control or other overeating, such as binge eating disorder and Prader-Willi syndrome. It will be apparent that the formulation can be used for the treatment of combinations of the described conditions.

[0116] That is, the present invention provides the formulation of the present invention for use in a method of treating, preventing, or reducing weight gain, promoting weight loss, and / or reducing overweight. Treatment can be achieved, for example, by controlling appetite, feeding, food intake, calorie intake, and / or energy consumption.

[0117] The present invention also provides a formulation of the present invention for use in a method of treating obesity and related diseases, disorders, and health conditions, including but not limited to morbid obesity, pre-surgical obesity, obesity-related inflammation, obesity-related gallbladder disease, sleep apnea and respiratory disorders induced by obesity, cartilage degeneration, osteoarthritis, and reproductive health complications such as infertility due to obesity or overweight. The subject may be afflicted with obesity associated with at least one weight-related co-morbid condition, such as diabetes (e.g., type 2 diabetes), hypertension, dyslipidemia, sleep apnea, and cardiovascular disease.

[0118] The present invention also provides a formulation of the present invention for use in a method of preventing or treating Alzheimer's disease, diabetes, type 1 diabetes, type 2 diabetes, prediabetes, insulin resistance syndrome, impaired glucose tolerance (IGT), disease states associated with elevated blood glucose levels, metabolic diseases including metabolic syndrome, hyperglycemia, hypertension, atherogenic dyslipidemia, hepatic steatosis (including "fatty liver" including non-alcoholic fatty liver disease (NAFLD) including non-alcoholic steatohepatitis (NASH)), renal insufficiency, arteriosclerosis (e.g., atherosclerosis), macrovascular disease, microvascular disease, diabetic heart disease (including diabetic cardiomyopathy and heart failure as a complication of diabetes), coronary heart disease, peripheral arterial disease or stroke, and combinations thereof.

[0119] The present invention also provides a formulation of the present invention for use in a method of reducing circulating LDL levels and / or increasing the HDL / LDL ratio.

[0120] The effect of the formulation on these conditions may be mediated, in whole or in part, through or independent of an effect on body weight.

[0121] The present invention further provides the use of a formulation of the present invention in the manufacture of a medicament for treating, preventing, or reducing weight gain, promoting weight loss, and / or reducing overweight.

[0122] The present invention also provides the use of the formulations of the present invention in the manufacture of a medicament for treating obesity and related diseases, disorders, and health conditions, including but not limited to morbid obesity, pre-surgical obesity, obesity-related inflammation, obesity-related gallbladder disease, sleep apnea and respiratory disorders induced by obesity, cartilage degeneration, osteoarthritis, and reproductive health complications such as infertility due to obesity or overweight. The subject may be afflicted with obesity associated with at least one weight-related co-morbid condition, such as diabetes (e.g., type 2 diabetes), hypertension, dyslipidemia, sleep apnea, and cardiovascular disease.

[0123] The present invention also provides the use of the formulations of the present invention in the manufacture of a medicament for preventing or treating Alzheimer's disease, diabetes, type 1 diabetes, type 2 diabetes, prediabetes, insulin resistance syndrome, impaired glucose tolerance (IGT), disease states associated with elevated blood glucose levels, metabolic diseases including metabolic syndrome, hyperglycemia, hypertension, atherogenic dyslipidemia, hepatic steatosis (including "fatty liver" including non-alcoholic fatty liver disease (NAFLD) including non-alcoholic steatohepatitis (NASH)), renal insufficiency, atherosclerosis (e.g., atherosclerotic arteriosclerosis), macrovascular disease, microvascular disease, diabetic heart disease (including diabetic cardiomyopathy and heart failure as a complication of diabetes), coronary heart disease, peripheral arterial disease or stroke, and combinations thereof.

[0124] The present invention also provides the use of the formulations of the present invention in the manufacture of a medicament for reducing circulating LDL levels and / or increasing the HDL / LDL ratio.

[0125] The present invention further provides a method for treating, preventing, or reducing weight gain, promoting weight loss, and / or reducing overweight in a subject, comprising the step of administering to the subject a therapeutically effective amount of the formulation of the present invention.

[0126] The present invention also provides a method for treating obesity and related diseases, disorders, and health conditions, including but not limited to morbid obesity, pre-surgical obesity, obesity-related inflammation, obesity-related gallbladder diseases, sleep apnea and respiratory disorders induced by obesity, cartilage degeneration, osteoarthritis, and reproductive health complications such as infertility due to obesity or overweight, in a subject, comprising the step of administering to the subject a therapeutically effective amount of a formulation of the present invention. The subject may be afflicted with obesity associated with at least one weight-related co-disease state, such as diabetes (e.g., type 2 diabetes), hypertension, dyslipidemia, sleep apnea, and cardiovascular diseases.

[0127] The present invention also provides a method for preventing or treating Alzheimer's disease, diabetes, type 1 diabetes, type 2 diabetes, prediabetes, insulin resistance syndrome, impaired glucose tolerance (IGT), disease states related to high blood glucose levels, metabolic diseases including metabolic syndrome, hyperglycemia, hypertension, atherogenic dyslipidemia, hepatic steatosis (''fatty liver'' including non-alcoholic fatty liver disease (NAFLD) including non-alcoholic steatohepatitis (NASH) per se), renal insufficiency, arteriosclerosis (e.g., atherosclerosis), macrovascular diseases, microvascular diseases, diabetic heart diseases (including diabetic cardiomyopathy and heart failure as a complication of diabetes), coronary heart diseases, peripheral arterial diseases or attacks, and combinations thereof in a subject, comprising the step of administering to the subject a therapeutically effective amount of a formulation of the present invention.

[0128] The present invention further provides a method for reducing circulating LDL levels and / or increasing the HDL / LDL ratio in a subject, comprising the step of administering to the subject a therapeutically effective amount of a formulation of the present invention.

[0129] The present invention further provides the use of the above-mentioned formulation in a cosmetic (i.e., non-therapeutic) method of weight loss.

[0130] It should be understood that references to the therapeutic use of the formulation and methods comprising administration of the formulation are to be construed equally as encompassing cosmetic use and administration.

[0131] The present invention [19CD]-isoGlu-RD()GTATK()ATERLA-Aad-FLQRSSF-Gly(Me)-A-Ile(Me)-LSSTEVGSNT-Hyp-NH 2 an amylin analog or a pharmaceutically acceptable salt and / or derivative thereof further provides a method for producing a stable aqueous pharmaceutical formulation comprising the method comprising (a) an amylin analog, salt, or derivative at a concentration of from about 0.4 mg / ml to about 25 mg / ml, and (b) a buffer at a concentration of from about 0.5 mM to about 25 mM formulating to produce a stable aqueous liquid formulation, the formulation having a pH of from about 5.8 to about 6.9.

[0132] It will be apparent that the method is applicable to any of the formulations of the invention described herein.

[0133] That is, for example, the method may further comprise formulating an amylin analog and a buffer with a tonicity modifier as described.

[0134] The method may include adjusting the pH to a desired value. When the tonicity modifier is a chloride salt, e.g., NaCl, it may be desirable to adjust the pH to the desired value before adding the tonicity modifier to the formulation. This is especially so when the amylin analog is provided itself in the form of a chloride salt, since the presence of chloride ions above a certain level may promote precipitation or fibrillation of the amylin analog at pH values near the pI of the peptide (pI = 4.36).

[0135] Also provided are formulations produced by the methods described.

[0136] Unless the context otherwise indicates, the descriptions and definitions of the features presented above are not limited to any particular embodiment or aspect of the invention and apply equally to all embodiments and aspects described.

[0137] Throughout this specification, the term "about" with respect to a numerical value is optional and means, for example, + / - 10%.

Mode for Carrying Out the Invention

[0138] Throughout this specification and the claims, the conventional one-letter or three-letter codes for natural amino acids are used. Unless otherwise specified, all amino acids are in the L-configuration.

[0139] Amylin analog The amylin analog (Compound 1; "CPD 1") present in the formulations of the present invention is described in WO2018 / 046719 and has the formula: [19CD]-isoGlu-RD()GTATK()ATERLA-Aad-FLQRSSF-Gly(Me)-A-Ile(Me)-LSSTEVGSNT-Hyp-NH 2 having the following structure. Gly(Me): N-methylglycine [also known as sarcosine (Sar)] Ile(Me): N-methylisoleucine Aad: 2-aminoadipic acid, for example (2S)-2-aminoadipic acid [(2S)-2-aminohexanedioic acid], also known as homoglutamic acid. [19CD]-: 19-carboxy-nonadecanoyl-

[0140] The parentheses "()" shown after the symbol of a particular amino acid residue indicate the residue whose side chain contributes to the intramolecular lactam bridge. That is, the amylin analog compound present in the formulations of the present invention has an intramolecular lactam bridge formed between the side chains of the residues shown in parentheses (aspartic acid and lysine, respectively).

[0141] The term "isoGlu" is used to mean a glutamic acid residue that contributes to the bond via the carboxyl group of its side chain and its alpha amino group. This may also be designated as γ-Glu.

[0142] That is, [19CD]-isoGlu- represents a 19-carboxy-nonadecanoyl group covalently bonded via an amide linkage to the alpha-amino group of the Glu linker. The side-chain carboxyl group of the Glu linker is linked via an amide linkage to the backbone alpha-amino group of the arginine residue next at the N-terminus of the peptide chain of the amylin analog.

[0143] The following formula shows this configuration including the side-chain of the N-terminal arginine residue and the carbonyl oxygen.

[0144]

Chemical formula

[0145] It should be understood that the amylin analog (active substance) can also be provided in the form of a salt or other derivative. The salt is typically a "pharmaceutically acceptable" salt, meaning a salt that is not harmful to the patient or the subject being treated thereby in the context of the present invention. Such salts are generally acid addition salts or basic salts. Acid addition salts include salts of inorganic acids and salts of organic acids. Non-limiting examples of suitable acid addition salts include hydrochloride, phosphate, formate, acetate, trifluoroacetate, and citrate. Examples of basic salts include those where the cation is an alkali metal ion such as sodium and potassium, an alkaline earth metal ion such as calcium, and for example NR(R') 3 + type where R and R' are independently optionally substituted C 1-6 alkyl, optionally substituted C 2-6It includes salts selected from alkenyl, optionally substituted aryl, or optionally substituted heteroaryl and the specified substituted ammonium ions. Other examples of pharmaceutically acceptable salts are described in Remington's Pharmaceutical Sciences, 17th Edition, Alfonso R. Gennaro (ed.), Mack Publishing Company, Easton, PA, U.S.A., 1985 and more recent editions, as well as in Encyclopaedia of Pharmaceutical Technology.

[0146] In particular, the salt may be a chloride salt. In some embodiments, the amylin analog has the formula ([19CD]-isoGlu-RD()GTATK()ATERLA-Aad-FLQRSSF-Gly(Me)-A-Ile(Me)-LSSTEVGSNT-Hyp-NH 2 ), x(Cl) where x is from 1.0 to 2.0.

[0147] Other derivatives of the amylin analog include coordination complexes with metal ions such as Mn 2+ and Zn 2+ esters such as esters of in vivo hydrolyzable esters, free acids or bases, hydrates, prodrugs, or lipids. Esters can be formed between the hydroxyl groups or carboxylic acid groups present in the compound and the reaction partners of appropriate carboxylic acids or alcohols using techniques well known in the art. Derivatives that are prodrugs can be converted to one of the parent compounds in vivo or in vitro. Typically, at least one biological activity of the compound is reduced in the prodrug form of the compound and can be activated by the conversion of the prodrug to release the compound or its metabolite. Examples of prodrugs include the use of protecting groups that are removed in situ to release the active compound or that act to inhibit the elimination of the drug in vivo.

[0148] The amylin analog is an agonist of the amylin receptor and has agonist activity at the receptors hAMYR1, hAMYR2, and hAMYR3 (human amylin receptors 1, 2, and 3), as described in WO2018 / 046719. Binding to the preferred receptor induces intracellular signaling, such as the production of cyclic AMP. In vivo, the amylin analog has (inter alia) the biological activities of reducing food intake, promoting weight loss, and / or preventing or reducing weight gain. This may be employed for various therapeutic uses, as described elsewhere in this specification and in WO2018 / 046719.

[0149] Aqueous liquid formulation Preferably, the formulations of the present invention are substantially free of organic solvents. In certain preferred embodiments, water is the only solvent used to make the aqueous liquid formulation.

[0150] The formulations of the present invention are aqueous liquid formulations, i.e., formulations containing water in the form of an aqueous solution. In the context of the present invention, the term "aqueous formulation" generally means a formulation containing at least 50% by weight (50% w / w) of water as a solvent, more preferably at least 75% w / w of water, more preferably at least 80% w / w of water, more preferably at least 85% w / w of water, more preferably at least 90% w / w of water, and most preferably at least 95% w / w of water. In certain embodiments, the aqueous formulations of the present invention are substantially free of organic solvents, such as aprotic polar solvents like dimethyl sulfoxide (DMSO). "Substantially" in this context means that the aqueous formulation contains less than 5% by volume of organic solvent, more preferably less than 2% by volume (v / v) of organic solvent, even more preferably less than 1% by volume (v / v) of organic solvent. In a preferred embodiment, no organic solvent is present. That is, typically, water is the only solvent used to make the aqueous liquid formulation.

[0151] The components and amounts of the liquid formulation of the present invention are selected to provide a formulation having a pH of about 5.8 to about 6.9, such as about 6.2 to about 6.8, such as about 6.4 to about 6.6, such as about 6.5. To avoid doubt, the pH is measured at 25°C.

[0152] The formulations of the present invention may further contain a preservative, especially when provided in a multiple-use format. Alternatively, especially when provided in a single-use format, the formulation may not contain or may substantially not contain a preservative. When a preservative is present, it will be employed in an amount sufficient to kill the types of bacteria within the range required for regulatory approval. Suitable preservatives include metacresol, phenol, methylparaben, ethylparaben, propylparaben, butylparaben, chlorobutanol, benzyl alcohol, phenylmercuric nitrate, thimerosal, sorbic acid, potassium sorbate, benzoic acid, chlorocresol, and benzalkonium chloride. The preservative will be present in an amount effective to kill the types of bacteria within the range required for regulatory approval.

[0153] The formulations of the present invention are typically suitable for parenteral administration by injection. That is, administration may be, for example, by subcutaneous, intramuscular, intraperitoneal, or intravenous injection, although subcutaneous administration may be preferred in some cases.

[0154] The formulations of the present invention may be provided in a container or delivery device. In some embodiments, the container or delivery device is for single use. In other embodiments, the container or delivery device is for multiple uses.

[0155] Examples include sealed vials, prefilled syringes, and injection devices such as injection pens, dose-adjustable autoinjectors, disposable autoinjectors, wearable syringes, or infusion pumps.

[0156] Stability A "stable" formulation is one in which the peptide therein essentially retains its physical and / or chemical and / or biological stability upon storage. Preferably, the formulation essentially retains its physical and chemical stability, as well as its biological stability, upon storage. The storage period is generally selected based on the intended shelf life of the formulation. Various analytical techniques for measuring the stability of proteins are available in the art, for example, there are reviews in Peptide and Protein Drug Delivery, pages 247 - 301, edited by Vincent Lee, Marcel Dekker, Inc., New York, N.Y., Pubs. (1991) and Jones, A. Adv. Drug Delivery Rev. 10: 29 - 90 (1993). The chemical and physical stability of an amylin analog in any given formulation can be determined according to the assays outlined in the following examples.

[0157] An aqueous liquid formulation is typically stable at 2 - 8°C (e.g., 5°C) for at least 6 months, e.g., at least 12 months, at least 18 months, or at least 24 months. Preferably, it is stable at 2 - 8°C (e.g., 5°C) for at least 18 months, e.g., at least 24 months.

[0158] It is also possible to test stability under accelerated or extreme conditions, in which case generally an elevated storage temperature is used to evaluate stability over a reduced time period. For example, storage at 25°C (sometimes referred to as "accelerated conditions at 25°C" or simply "accelerated conditions") may be used to evaluate stability over a short period, e.g., 1, 2, 3, or 4 weeks, 1 month, 2 months, 3 months, or 6 months.

[0159] As used herein, the term "physical stability" means a measure of the tendency of an amylin analog to form soluble or insoluble aggregates, such as fibrils.

[0160] Physical stability (e.g., turbidity of a solution, or aggregation, fibrillation, or gelation of an amylin analog) can be evaluated qualitatively and / or quantitatively by a variety of different methods, such as visual inspection, size exclusion chromatography, UV light scattering, dynamic light scattering (DLS), circular dichroism, measurement of turbidity, or using a spectroscopic probe (such as thioflavin T) that indicates the conformational state of the peptide. The probe is preferably a small molecule that preferentially binds to non-native conformers of the peptide. Thioflavin T (ThT) is a fluorescent dye that has been widely used for the detection of amyloid fibrils. In the presence of fibrils and presumably other peptide assemblies, thioflavin T gives rise to a new excitation maximum at about 450 nm and enhanced emission at 482 nm when bound to the fibril form of the peptide. Unbound thioflavin T is essentially non-fluorescent at the wavelengths in question.

[0161] In some embodiments, the formulation exhibits substantially no change that is an indicator of a decrease in physical stability (e.g., as indicated by turbidity of a solution, or aggregation, fibrillation, or gelation of an amylin analog) after storage at 2 - 8 °C (e.g., 5 °C) for at least 6 months, at least 12 months, at least 18 months, or at least 24 months. Preferably, the formulation exhibits substantially no such change after storage at 2 - 8 °C (e.g., 5 °C) for at least 18 months, e.g., at least 24 months.

[0162] Alternatively or in addition, the formulation exhibits substantially no such change after storage under accelerated conditions (25 °C) for a period of 1, 2, 3, or 4 weeks, 1 month, 2 months, 3 months, or 6 months.

[0163] As used herein, the term "chemical stability" refers to the stability of a peptide with respect to covalent / chiral chemical changes in the peptide structure that result in the formation of chemical degradation products and / or covalent oligomers having potentially lower biological activity compared to the native monomeric peptide structure. The primary degradation products derived from amylin analogs present in the formulations of the invention are thought to be due to deamidation of asparagine side chains and / or C-terminal amide groups that form free carboxylic acid groups. Covalently linked oligomers are also thought to form in sub-optimal formulations.

[0164] The chemical stability of a formulation can be evaluated by measuring the amount of the remaining native monomeric peptide, specific chemical degradation products, or covalent oligomers at various time points after exposure to various environmental conditions. The amount of each individual degradation product can be determined by separating the degradation products to generate a "degradation profile". However, the amount of the remaining native monomeric peptide can be a preferred indicator of chemical stability. Separation is typically carried out using various chromatography techniques based on molecular size and / or charge, such as size exclusion chromatography (SEC) or reverse phase (RP) chromatography, typically high performance liquid chromatography (HPLC), e.g., SEC-HPLC (size exclusion chromatography HPLC) and / or RP-HPLC (reverse phase HPLC).

[0165] In some embodiments, at least 80%, more preferably at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% of the amylin analog remains in its original monomeric form in the formulation after being stored at 2-8 °C (e.g., at 5 °C) for at least 6 months, at least 12 months, at least 18 months, or at least 24 months. Preferably, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% of the amylin analog remains in its original monomeric form in the formulation after being stored at 2-8 °C (e.g., at 5 °C) for at least 6 months, at least 12 months, at least 18 months, or at least 24 months. More preferably, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% of the amylin analog remains in its original monomeric form in the formulation after being stored at 2-8 °C (e.g., at 5 °C) for at least 18 months, e.g., at least 24 months.

[0166] Buffer As used herein, the term "buffer" means a pharmaceutically acceptable agent that stabilizes the pH of a pharmaceutical formulation. Suitable buffers are well known in the art and can be found in the literature.

[0167] Any suitable buffer may be used, including phosphate buffers, histidine buffers, TRIS (tris(hydroxymethyl)aminomethane) buffers, acetate buffers, arginine buffers, citrate buffers, bicarbonate buffers, diethanolamine buffers, lysine buffers, and TAPS ([tris(hydroxymethyl)methylamino]propanesulfonic acid) buffers. Particularly preferred buffers include phosphate buffers, histidine buffers, citrate buffers, and TRIS buffers, especially phosphate buffers, histidine buffers, and citrate buffers.

[0168] Surprisingly, it has been found that stability increases at relatively low buffer concentrations. That is, the buffer is present at a concentration of about 0.5 mM to about 25 mM.

[0169] The buffer may be present at a concentration of about 0.5 mM to about 20 mM, such as about 0.5 mM to about 15 mM, such as about 1 mM to about 15 mM, such as about 1 mM to about 12 mM. For example, the buffer may be present at a concentration of about 3 mM to about 7 mM, or about 8 mM to about 12 mM, such as about 5 mM or about 10 mM.

[0170] For TRIS buffer, it is preferably present at about 15 mM to about 25 mM, such as about 17 mM to about 23 mM, such as about 20 mM. This is because the buffering capacity of TRIS is lower than that of other buffers such as phosphate, histidine, and citrate.

[0171] It is also possible for the formulation to contain two or more buffers, such as two buffers or more. In such cases, the cited concentrations may represent the sum of the concentrations of the individual buffers. For example, a formulation containing 2.5 mM phosphate buffer and 2.5 mM histidine buffer may be considered to contain 5 mM buffer in total.

[0172] Tonicity adjuster As used herein, the term "tonicity adjuster" means a pharmaceutically acceptable tonicity agent used to adjust the tonicity of a formulation. The terms "tonicity adjuster" and "tonicity agent" may be used interchangeably. The formulations of the present invention are preferably isotonic, i.e., they have an osmotic pressure that, when measured with an osmometer, is substantially the same as that of human serum, such as about 300 + / - 80 mOsm, such as 300 + / - 60 mOsm.

[0173] The tonicity adjuster may be ionic or non-ionic. Suitable ionic tonicity adjusters include alkali metal salts (such as halides), and alkaline earth metal salts (such as halides), such as NaCl, NaBr, NaI, KCl, KBr, KI, LiCl, CaCl 2 and Na 2 SO 4It is included. Non-ionic tonicity regulators include mannitol (e.g., D-mannitol), propylene glycol, sucrose, glycerol, sorbitol, trehalose, and dextrose.

[0174] When the tonicity regulator is a chloride salt such as NaCl, it may be desirable to add the tonicity regulator to the formulation only after the pH has been adjusted to the desired value. This is especially so when the amylin analog is provided in the form of a chloride salt itself. This is because the presence of chloride ions above a certain level may promote precipitation or fibrillation of the amylin analog at pH values near the pI of the peptide (pI = 4.36).

[0175] In some embodiments, mannitol (e.g., D-mannitol), propylene glycol, and NaCl, particularly mannitol (e.g., D-mannitol) and propylene glycol, may be preferred as tonicity regulators.

[0176] It will be understood that the concentration of the tonicity regulator will depend on the concentration of the other components of the formulation, particularly when the formulation is intended to be isotonic. One of ordinary skill in the art can establish the appropriate concentration for any given agent with respect to a particular formulation.

[0177] For example, it may be appropriate to employ mannitol at a concentration of 200-300 mM, such as 230-260 mM. It may be appropriate to employ propylene glycol at a concentration of 200-250 mM, such as 224 mM. It may be appropriate to employ NaCl at a concentration of 100-150 mM, such as 118-137 mM.

[0178] Therapeutic Use The formulations of the present invention are useful, inter alia, in reducing food intake, promoting weight loss, and preventing or reducing weight gain. Accordingly, the formulations may provide an attractive treatment option for metabolic diseases caused by, characterized by, or associated with obesity and overweight.

[0179] That is, the present formulation may be used in a method for treating, preventing, or reducing weight gain, promoting weight loss, reducing food intake, and / or reducing overweight. The treatment can be achieved, for example, by controlling appetite, feeding, food intake, calorie intake, and / or energy consumption.

[0180] The present formulation can be used in a method for treating obesity and related diseases, disorders, and health conditions, including but not limited to morbid obesity, pre-surgical obesity, obesity-related inflammation, obesity-related gallbladder diseases, sleep apnea and respiratory disorders induced by obesity, cartilage degeneration, osteoarthritis, and reproductive health complications such as infertility due to obesity or overweight.

[0181] The present formulation may also be used in a method for preventing or treating Alzheimer's disease, diabetes, type 1 diabetes, type 2 diabetes, prediabetes, insulin resistance syndrome, impaired glucose tolerance (IGT), disease states related to high blood glucose levels, metabolic diseases including metabolic syndrome, hyperglycemia, hypertension, atherogenic dyslipidemia, hepatic steatosis (''fatty liver'' including non-alcoholic fatty liver disease (NAFLD) including non-alcoholic steatohepatitis (NASH)), renal insufficiency, arteriosclerosis (e.g., atherosclerosis), macrovascular diseases, microvascular diseases, diabetic heart diseases (including diabetic cardiomyopathy and heart failure as a complication of diabetes), coronary heart disease, peripheral arterial disease, or stroke.

[0182] The present formulation may also be useful in reducing circulating LDL levels and / or increasing the HDL / LDL ratio.

[0183] These effects may be mediated, in whole or in part, through or independent of the effect on body weight.

[0184] Metabolic syndrome is characterized by a cluster of metabolic risk factors in an individual. Metabolic risk factors include abdominal obesity (excessive adipose tissue around the abdominal viscera), atherogenic dyslipidemia (blood lipid disorders including high triglycerides, low HDL cholesterol and / or high LDL cholesterol, which promote the accumulation of plaques in the arterial wall), elevated blood pressure (hypertension), insulin resistance and glucose intolerance, a prothrombotic state (e.g., high fibrinogen or plasminogen activator inhibitor-1 in the blood), and a proinflammatory state (e.g., elevated blood C-reactive protein). Individuals with metabolic syndrome are at increased risk of coronary heart disease and other diseases associated with other signs of atherosclerosis (e.g., stroke and peripheral vascular disease). The main risk factor underlying this syndrome appears to be abdominal obesity.

[0185] As used in the context of the present invention, the term "treatment" (and "treating" and other grammatical variations thereof) means an approach for obtaining a beneficial or desired clinical outcome. For the purposes of the present invention, beneficial or desired clinical outcomes include, but are not limited to, alleviation of symptoms, whether detectable or undetectable, attenuation of the degree of the disease, stabilization of the state of the disease (i.e., not getting worse), delay or retardation of the progression of the disease, improvement or alleviation of the state of the disease, and remission (partial or complete). "Treatment" can also mean an extension of the survival period compared to the survival period expected in the absence of treatment. "Treatment" is an intervention that is intended to prevent the progression of a disorder or to change its pathological condition. Thus, "treatment" means both therapeutic treatment and prophylactic or preventive means. In the context of prophylactic or preventive means, the pharmaceutical formulation need not completely prevent the progression of the disease or disorder. Persons in need of treatment include those already suffering from the disorder and those in whom progression of the disorder should be prevented. "Treatment" also means the prevention or reduction of an increase in pathological condition or symptoms (e.g., weight gain or hypoglycemia) compared to the situation without treatment, and does not necessarily imply a complete cessation of the associated condition.

Examples

[0186] The following examples are provided to illustrate preferred embodiments of the present invention and are not intended to limit the scope of the present invention.

[0187] Method Preparation of Compound 1 (CPD 1) Formulation (Method I) Compound 1 (described in 2018 / 046719) was prepared as a chloride salt. Except as noted below, the formulation of CPD 1 was prepared by dissolving the CPD 1 peptide in alkaline MilliQ water (MQW) (pH adjusted with NaOH) to form a clear drug substance (DS) stock solution at a concentration related to the target final product concentration and at approximately pH 7. The relevant excipients were dissolved in MQW at the appropriate pH to form an excipient solution. The two solutions were mixed, the pH was measured, and if necessary, diluted NaOH / HCl was used to adjust it to the desired pH as needed. Finally, an appropriate amount of MQW was added to reach the concentration of the final product. The formulation was aseptically filtered through a 0.22 μm filter and filled into suitable containers.

[0188] Method for Determining Chemical Stability (Assay II) The chemical stability of CPD 1 was evaluated using the following method.

[0189] Determination of Purity (%) by RP-HPLC (Assay IIa) For the analysis, a Dionex Ultimate 3000 HPLC system was used at a flow rate of 0.3 mL / min. The mobile phase components consisted of a gradient elution of 20 mM phosphate buffer pH 7.0 in acetonitrile (ACN). A wavelength of 220 nm was used for detection. The injection volume was 4 μg of the peptide. The column used for HPLC analysis was a Waters Acquity UPLC BEH C18, 150 × 2.1 mm, particle size 1.7 μm. The runtime was 35 - 37 minutes depending on the re-equilibration time.

[0190] The results shown in the examples are measured by RP-HPLC after incubation under stress conditions. The chemical stability of CPD 1 (peptide) is determined by HPLC at a given time point and is represented herein as the relative purity of the peptide peak (i.e., the major peptide peak) normalized to the absolute purity of the peptide peak at day zero (day 0 (D0)), i.e., the major peptide peak. The peptide peak at D0 is set to 100% purity, and thus the chemical stability is expressed as a percentage of CPD 1.

[0191] Purity is a measure of the remaining intact compound in the stressed solution after incubation, expressed as a percentage relative to the purity measured at day zero (D0, day zero) and normalized to the purity of CPD 1.

[0192] Determination of covalent oligomer formation by size exclusion chromatography (SEC) (assay IIb) For the analysis, a Dionex Ultimate 3000 HPLC system was used at a flow rate of 0.5 mL / min (isocratic elution). The mobile phase components consisted of 45% acetonitrile (ACN) and 0.1% trifluoroacetic acid (TFA) in 55% MQW. A wavelength of 215 nm (+280 nm) was used for detection. The injection volume was 4 μg of peptide. The column used for HPLC analysis was a TSKgel SuperSW2000 from Tosoh BioScience, 30 cm × 4.6 mm, particle size 4 μm. The runtime was 25 minutes.

[0193] The results shown in the examples are measured by HPLC after incubation under stress conditions. The level of covalent oligomers is calculated as the peak area of the covalent oligomers relative to the intact monomer main peak of CPD-1 and is expressed as % oligomers.

[0194] Method for evaluating physical stability (assay III) To evaluate the physical stability of CPD 1 in the formulation, the following method was used.

[0195] Visual inspection of the solution (Assay IIIa) The samples were provided in transparent containers, and it was possible for trained and experienced technicians to visually inspect the formulations. The visual inspection was carried out manually and recorded by experienced technicians. The samples were classified as "clear", "suspended", "gel", or "particulate" according to their appearance.

[0196] UV absorbance at 325 nm (Assay IIIb) Turbidity was measured by UV absorbance analysis. 100 μL of each sample was loaded onto a transparent 96-well UV-compatible plate from Corning. The samples were measured at 325 nm using a SpectraMax 190 Elisa reader.

[0197] Dynamic light scattering (DLS) (Assay IIIc) The particle size distribution of the peptide was determined by dynamic light scattering analysis. Using a 96-well plate from nunc, a standard DLS experiment was carried out with a sample volume of 165 μL and measured by a DynaPro Plate reader II device. Typically used parameters were temperature 25 °C, acquisition time 2 s, 50 acquisitions per sample, detection: hydrodynamic radius Rh, correlation function cut-off value [μs]: 0.5~1×10 5 , peak radius cut-off [nm]: 0.5~1000.

[0198] Evaluation of physical stability by ThT stress test (Assay IIId) Aggregation in the form of fibril formation was detected using thioflavin T (ThT), an amyloid-specific dye. This is often employed to demonstrate the presence of fibrils in solution (see, for example, Groenning, M., J. Chem. Biol. 3(1) (2010), pp. 1~18; Groenning et al., J. Struct. Biol. 158 (2007) 358~369; and Levine, H., III, Protein Sci. 2 (1993) 404~410).

[0199] A 1 mM ThT stock solution was prepared, and 12 μL was mixed with 288 μL of CPD 1 formulation. Two 150 μL samples were loaded in duplicate into a 96-well black fluorescence plate (transparent bottom). Data were collected at 40 °C at fixed 10-minute intervals up to 60 hours using a Biotek Synergy™ H4 Hybrid Multi-Mode fluorescence plate reader after 300 seconds of automatic mixing (stirring) each. Physical stability was expressed as the lag time (in hours) of fibril formation and was defined as the intersection of two linear regressions representing the initial stable and growth phases.

[0200] Evaluation of physical stability by the ThT single-point method (assay IIIe) Aggregation in the form of fibril formation was detected using the amyloid-specific dye thioflavin T (ThT). This is often employed to demonstrate the presence of fibrils in solution (see, for example, Groenning, M., J. Chem. Biol. 3(1) (2010), pp. 1–18; Groenning et al., J. Struct. Biol. 158 (2007) 358–369; and Levine, H., III, Protein Sci. 2 (1993) 404–410).

[0201] A 170 μL sample of the CPD 1 formulation was loaded into a black 96-well plate with an optical bottom from Nunc. An additional 7 μL of 1 mM ThT was added and gently pipetted up and down to mix, ensuring uniform dispersion of ThT. The 96-well plate was placed in a Fluorescence Plate reader Biotek Synergy™ H4 Hybrid Multi-Mode or a Fluorescence Plate reader Biotek Synergy™ MX. Excitation 450 nm (bandwidth 9 nm), emission 485 nm (bandwidth 9 nm), and excitation 295 nm (bandwidth 9 nm), emission 355 nm (bandwidth 9 nm) and emission 330 nm (bandwidth 9 nm).

[0202] (Example 1) Accelerated Chemical and Physical Stability of the Formulation at 25°C in the pH Range of 6.1 - 7.8 The formulations described in Table 1 were prepared as described in Method I, except as described in the table.

[0203]

Table 1

[0204] The prepared formulations were filled into 1 - ml type 1 glass vials, stored at 25°C in the dark for the indicated period, and then analyzed.

[0205]

Table 2

[0206] There is no obvious influence of pH on physical stability, but the purity increases with the decrease of pH. See Table 2.

[0207] (Example 2) Effect of Formulation pH on Physical Stability CPD 1 pharmaceutical biologic formulations containing 1 mg / ml of CPD 1, 20 mM of TRIS (buffer), and 118 mM of NaCl (tonicity agent) were prepared at pH 5.8, 6.0, 6.3, 6.8, and 7.0 as described in Method I.

[0208] Physical stability was evaluated in terms of fibril formation by ThT stress test measured over 60 hours at 40°C. The initial peptide particle size (z - average) was also determined by DLS.

[0209]

Table 3

[0210] Table 3 shows that in the pH range of 5.8 - 7.0, CPD 1 is stable against fibrillation and aggregation that affect the peptide particle size.

[0211] (Example 3) Accelerated chemical and physical stability at 25 °C of formulations with buffer concentrations of 2 mM to 20 mM The formulations described in Table 4 (Table 4) were prepared as described in Method I. The prepared formulations were filled into 1 ml type 1 glass vials, stored at 25 °C in the dark for the indicated period, and then analyzed.

[0212]

Table 4A

[0213]

Table 4B

[0214] As exemplified by Formulations 1 to 4, the purity is higher than 20 mM concentration in a buffer of 5 mM concentration of histidine and phosphate. For buffer concentrations of 5 mM and 20 mM of histidine and phosphate, no or limited effects on visual appearance, physical stability, and covalent oligomers were observed.

[0215]

[0216] (Example 4) Accelerated physical stability in the pH range of 6.1 to 6.9 CPD 1 biopharmaceutical formulations #1 to 3 of Table 1 (Table 1) (Example 1) were filled into 1 ml type 1 glass vials and subjected to physical stress in the form of a steady rotation at approximately 15 rpm (revolutions per minute) at room temperature on a multi-rotator installed in a standard laboratory where sunlight from the window hits (no light shielding) for the indicated period. Stability was determined as shown in the table.

[0217]

Table 5

[0218] The formulations within the pH interval of 6.1 - 6.9 are physically stable during 4 weeks of rotation at room temperature.

[0219] (Example 5) Chemical stability or solubility of formulations with CPD 1 concentration of 0.4 mg / mL - 50 mg / mL A 4 mg / ml formulation of CPD 1 containing 20 mM of TRIS and 118 mM of NaCl was prepared as described in Method I. The formulation was diluted with placebo buffer to reach a concentration of 0.4 mg / ml and analyzed after storage at 25 °C for 24 hours.

[0220]

Table 6

[0221] After storage at a low concentration of 0.4 mg / ml, there was no decrease in purity. See Table 7 (Table 6).

[0222] To test solubility, CPD 1 was dissolved in alkaline MQW by stirring (pH was adjusted with NaOH).

[0223]

Table 7

[0224] CPD 1 could be dissolved into a clear solution at concentrations up to 50 mg / ml in MQW. See Table 8 (Table 7).

[0225] (Example 6) Accelerated chemical stability at 40 °C of formulations at pH 6 and buffer concentration of 0.5 - 40 mM Formulations in Tables A1 - A3 (Table A1 - A3) were prepared according to Method I and filled into 1.5 ml LowBind Eppendorf tubes. Tonicity agents were not included.

[0226] Stability was tested according to Assay II, with the modification (Assay IIb) that results in the formation of covalent oligomers (SEC) reported as the difference in total oligomers between the initial value and the 4-week time point after storage at 40°C for 4 weeks.

[0227] [Table 8]

[0228] [Table 9]

[0229] [Table 10]

[0230] At pH 6, the decrease in purity is reduced as the buffer concentration decreases (Table 9). The buffer concentration has no significant effect on the formation of covalent oligomers (Table 10).

[0231] The total buffer concentration appears to be the most significant factor. Switching from a single buffer at 5 mM to a mixture of two buffers at 2.5 mM each does not appear to have a significant effect (compare Formulations 29 - 32 in Table 11 and Tables 9 and 10).

[0232] (Example 7) Accelerated Chemical and Physical Stability at 25°C of Formulations Containing Various Concentrations of Compound 1 Formulations in Table 12 were prepared according to Method I and filled into 1-ml Type 1 glass cartridges that had been siliconized and fitted with bromobutyl plungers. After storage at 25°C for the indicated periods, stability was tested according to Assay II and Assays IIIa, IIIb, and IIIc.

[0233]

Table 11

[0234] The change in the concentration of CPD 1 in the range of 1 - 10 mg / ml tested did not affect the chemical and physical stability.

[0235] (Example 8) Accelerated chemical stability at 25°C of formulations containing various tonicity agents The formulations in Table 13 (Table 12) were prepared according to Method I and filled into 1 - ml type 1 glass vials. After storage at 25°C for the indicated period, the stability was tested according to Assay II.

[0236]

Table 12

[0237] The decrease in purity seems to be lower for formulations containing mannitol and propylene glycol as tonicity agents than for formulations containing glycerol and NaCl as tonicity agents. The formation of covalent oligomers seems to be higher for formulations containing glycerol as tonicity agent than for formulations containing other tonicity agents.

[0238] The following paragraphs (paras.) present certain embodiments of the invention. 1. [19CD]-isoGlu-RD()GTATK()ATERLA-Aad-FLQRSSF-Gly(Me)-A-Ile(Me)-LSSTEVGSNT-Hyp-NH 2 An amylin analog or a pharmaceutically acceptable salt and / or derivative thereof comprising a stable aqueous liquid pharmaceutical formulation, (a) an amylin analog at a concentration of about 0.4 mg / ml to about 25 mg / ml, and (b) a buffer at a concentration of about 0.5 mM to about 25 mM comprising, a stable aqueous liquid formulation having a pH of about 5.8 to about 6.9. 2. The stable aqueous liquid formulation according to paragraph 1, wherein the amylin analog is present at a concentration of at least about 0.4 mg / ml, at least about 0.5 mg / ml, at least about 0.6 mg / ml, at least about 1.2 mg / ml, or at least about 2.5 mg / ml. 3. The stable aqueous liquid formulation according to paragraph 1 or 2, wherein the amylin analog is present at up to about 25 mg / ml, up to about 20 mg / ml, up to about 15 mg / ml, or up to about 10 mg / ml. 4. The stable aqueous liquid formulation according to any one of paragraphs 1 to 3, wherein the amylin analog is present at a concentration of about 3 mg / ml to about 12 mg / ml, such as about 3 mg / ml to about 7 mg / ml, or about 8 mg / ml to about 12 mg / ml, such as about 5 mg / ml or about 10 mg / ml. 5. The stable aqueous liquid formulation according to any one of paragraphs 1 to 4, wherein the buffer is present at a concentration of about 0.5 mM to about 20 mM, such as about 0.5 mM to about 15 mM, such as about 1 mM to 15 mM, such as about 1 mM to about 12 mM. 6. The stable aqueous liquid formulation according to any one of paragraphs 1 to 5, wherein the buffer is present at a concentration of about 3 mM to about 7 mM, or about 8 mM to about 12 mM, such as about 5 mM, or about 10 mM. 7. The stable aqueous liquid formulation according to any one of paragraphs 1 to 6, wherein the buffer is phosphate, histidine, or citrate. 8. The stable aqueous liquid formulation according to any one of paragraphs 1 to 6, wherein the buffer is a TRIS buffer and the buffer is present at about 15 mM to about 25 mM, such as about 17 mM to about 23 mM, such as about 20 mM. 9. The stable aqueous liquid formulation according to any one of paragraphs 1 to 8, wherein the pH is about 6.2 to about 6.8, such as about 6.4 to about 6.6, such as about 6.5. 10. The stable aqueous liquid formulation according to any one of paragraphs 1 to 9, further comprising a tonicity modifier. 11. A stable aqueous liquid formulation as described in any one of paragraphs 1 to 10, which is stable at 2 - 8°C for at least 6 months, at least 12 months, at least 18 months, or at least 24 months. 12. A stable aqueous liquid formulation as described in any one of paragraphs 1 to 11, which, after storage at 2 - 8°C for at least 6 months, at least 12 months, at least 18 months, or at least 24 months, does not substantially exhibit turbidity, or aggregation, fibrillation, or gelation of the amylin analog. 13. A stable aqueous liquid formulation as described in any one of paragraphs 1 to 12, in which at least 80%, more preferably at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% of the amylin analog remains in its original monomeric form in the formulation after storage at 2 - 8°C for at least 6 months, at least 12 months, at least 18 months, or at least 24 months. 14. A stable aqueous liquid formulation as described in any one of paragraphs 1 to 13, in which the amylin analog is provided as a chloride salt. 15. A stable aqueous liquid formulation as described in any one of paragraphs 1 to 14, in which water is the only solvent used to prepare the aqueous liquid formulation. 16. A stable aqueous liquid formulation as described in any one of paragraphs 1 to 15, which is formulated for administration to a subject by injection. 17. A stable aqueous liquid formulation as described in paragraph 16, in which the injection is a subcutaneous injection. 18. A stable aqueous liquid formulation as described in any one of paragraphs 1 to 17, which is sterile. 19. A container or delivery device containing the liquid formulation as described in any one of paragraphs 1 to 18. 20. A stable aqueous liquid formulation as described in any one of paragraphs 1 to 18, for use in a method of medical treatment. 21. A stable aqueous liquid formulation for use in a method of treating, preventing or reducing weight gain, promoting weight loss, and / or reducing overweight, as described in any one of paragraphs 1 to 18. 22. A stable aqueous liquid formulation for use in a method of treating obesity (e.g., morbid obesity, pre-surgical obesity), obesity-related inflammation, obesity-related gallbladder disease, sleep apnea induced by obesity, cartilage degeneration, osteoarthritis, or reproductive health complications due to obesity or overweight, as described in any one of paragraphs 1 to 18. 23. A stable aqueous liquid formulation for use in a method of preventing or treating Alzheimer's disease, diabetes (e.g., type 1 diabetes, type 2 diabetes), prediabetes, insulin resistance syndrome, impaired glucose tolerance (IGT), metabolic syndrome, hyperglycemia, hypertension, atherogenic dyslipidemia, hepatic steatosis ("fatty liver", e.g., non-alcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH)), renal insufficiency, arteriosclerosis (e.g., atherosclerotic arteriosclerosis), macrovascular disease, microvascular disease, diabetic heart disease (e.g., diabetic cardiomyopathy), coronary heart disease, peripheral arterial disease or stroke, as described in any one of paragraphs 1 to 18. 24. A stable aqueous liquid formulation for use in a method of reducing circulating LDL levels and / or increasing the HDL / LDL ratio, as described in any one of paragraphs 1 to 18. 25. [19CD]-isoGlu-RD()GTATK()ATERLA-Aad-FLQRSSF-Gly(Me)-A-Ile(Me)-LSSTEVGSNT-Hyp-NH 2 An amylin analog or a pharmaceutically acceptable salt and / or derivative thereof A method for producing a stable aqueous pharmaceutical formulation comprising (a) an amylin analog at a concentration of about 0.4 mg / ml to about 25 mg / ml, and (b) a buffer at a concentration of about 0.5 mM to about 25 mM Formulating to produce a stable aqueous liquid formulation, the formulation having a pH of about 5.8 to about 6.9. 26. A formulation produced by the method according to paragraph 25, which is a stable aqueous liquid formulation as described in any one of paragraphs 1 to 18.

Claims

1. [19CD]-isoGlu-RD()GTATK()ATERLA-Aad-FLQRSSF-Gly(Me)-A-Ile(Me)-LSSTEVGSNT-Hyp-NH 2 or a pharma- ceutically acceptable salt thereof. A stable aqueous liquid pharmaceutical formulation comprising: (a) an amylin analog at a concentration of 0.4 mg / ml to 25 mg / ml; and (b) Buffers with concentrations between 0.5 mM and 25 mM Including, A stable aqueous liquid formulation having a pH of 5.8 to 6.

9.

2. 2. The stable aqueous liquid formulation of claim 1, wherein the amylin analog is present in a concentration of at least 0.4 mg / ml, at least 0.5 mg / ml, at least 0.6 mg / ml, at least 1.2 mg / ml, or at least 2.5 mg / ml.

3. 2. The stable aqueous liquid formulation of claim 1, wherein the amylin analog is present at up to 25 mg / ml, up to 20 mg / ml, up to 15 mg / ml, or up to 10 mg / ml.

4. 2. The stable aqueous liquid formulation of claim 1, wherein the amylin analogue is present in a concentration of from 3 mg / ml to 12 mg / ml, such as from 3 mg / ml to 7 mg / ml, or from 8 mg / ml to 12 mg / ml, such as from 5 mg / ml or 10 mg / ml.

5. The stable aqueous liquid formulation of claim 1, wherein the amylin analog is present at a concentration of 1.2 mg / ml to 10 mg / ml.

6. The stable aqueous liquid formulation of claim 1, wherein the amylin analog is present at a concentration of 12 mg / ml.

7. The stable aqueous liquid formulation of claim 1, wherein the amylin analog is present at a concentration of 18 mg / ml.

8. 2. The stable aqueous liquid formulation of claim 1, wherein the buffering agent is present at a concentration of 0.5 mM to 20 mM, such as 0.5 mM to 15 mM, such as 1 mM to 15 mM, such as 1 mM to 12 mM.

9. 2. The stable aqueous liquid formulation of claim 1, wherein the buffering agent is present at a concentration of 3 mM to 7 mM, or 8 mM to 12 mM, such as 5 mM, or 10 mM.

10. 2. The stable aqueous liquid formulation of claim 1, wherein the buffer is phosphate, histidine, or citrate.

11. 2. The stable aqueous liquid formulation of claim 1, wherein the buffer is a TRIS buffer and the buffer is present at 15 mM to 25 mM, such as 17 mM to 23 mM, such as 20 mM.

12. 2. The stable aqueous liquid formulation according to claim 1, having a pH of 6.2 to 6.8, such as 6.4 to 6.6, such as 6.

5.

13. 10. The stable aqueous liquid formulation of claim 1, further comprising a tonicity adjuster.

14. 2. The stable aqueous liquid formulation of claim 1, which is stable at 2-8° C. for at least 6 months, at least 12 months, at least 18 months, or at least 24 months.

15. 2. The stable aqueous liquid formulation of claim 1, which exhibits substantially no turbidity, or aggregation, fibrillation, or gelation of the amylin analogue after storage at 2-8° C. for at least 6 months, at least 12 months, at least 18 months, or at least 24 months.

16. 2. The stable aqueous liquid formulation of claim 1, wherein at least 80% of the amylin analogue remains in its original monomeric form in the formulation after storage at 2-8°C for at least 6 months, at least 12 months, at least 18 months, or at least 24 months.

17. 2. The stable aqueous liquid formulation of claim 1, wherein the amylin analog is provided as a chloride salt.

18. 18. The stable aqueous liquid formulation of any one of claims 1 to 17, formulated for administration to a subject by injection, for example wherein the injection is a subcutaneous injection.

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