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 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. [Background technology]

[0002] Amylin is a member 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 implicated in a variety of metabolic diseases and disorders. Human amylin was first isolated, purified, and characterized as a major component of amyloid deposits in pancreatic islets of type 2 diabetic patients.

[0003] Natural human amylin has the formula H-KC()NTATC()ATQRLANFLVHSSNNFGAILSSTNVGSNTY-NH2 is a 37 amino acid peptide having the formula: In the formula, 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), -NH2 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.

[0004] Amylin is believed to be beneficial in the treatment of metabolic disorders such as diabetes and / or obesity. Amylin is believed to regulate gastric emptying and inhibit glucagon secretion and food intake, thereby regulating the rate of glucose release into the circulation. Amylin appears to complement the action of insulin. Compared to healthy adults, type 1 diabetic patients have no circulating amylin, and type 2 diabetic patients exhibit reduced postprandial amylin concentrations. In human studies, the amylin analogue known as pramlintide, which is described in WO 93 / 10146 and has 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, has also been shown to reduce body weight or reduce weight gain.

[0005] An alternative amylin analogue, designated IAPP-GI, that incorporates N-methylated residues and has a reduced tendency to fibrillate has been 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 less activity than native amylin.

[0006] WO2018 / 046719 describes amylin analogs having, inter alia, lactam bridges instead of disulfide bridges, N-methylated residues, and deletions corresponding to residues Asn21 and Asn22 of native human amylin. Such analogs have a much lower tendency to fibrillate than native amylin, while also having a higher potency than the analogs described in Yan et al. (supra). These are typically amenable to formulation at or near physiological pH. However, there is a need for the development of improved formulations of these analogs, in particular to provide stable formulations that can be stored for extended periods without excessive fibrillation or degradation of the active monomeric form of the peptide. [Prior art documents] [Patent documents]

[0007] [Patent Document 1] WO93 / 10146 [Patent Document 2] WO2006 / 042745 [Patent Document 3] WO2018 / 046719 [Non-patent literature]

[0008] [Non-Patent Document 1] Yan et al., PNAS, 103(7), pp. 2046-2051, 2006 [Non-Patent Document 2] Angew. Chem. Int. Ed. 2013, 52, pp. 10378~10383 [Non-Patent Document 3] Remington's Pharmaceutical Sciences, 17th edition, Alfonso R. Gennaro (ed.), Mack Publishing Company, Easton, PA, USA, 1985 [Non-Patent Document 4] Encyclopaedia of Pharmaceutical Technology [Non-Patent Document 5] Peptide and Protein Drug Delivery, pp. 247-301, edited by Vincent Lee, Marcel Dekker, Inc., New York, NY, Pubs. (1991) [Non-Patent Document 6] Jones, A. Adv. Drug Delivery Rev. 10: pp. 29-90 (1993) [Non-Patent Document 7] Groenning, M., J. Chem. Biol. 3(1) (2010), pp. 1-18 [Non-Patent Document 8] Groenning et al., J. Struct. Biol. 158 (2007) 358-369 [Non-Patent Document 9] Levine, H., III, Protein Sci. 2 (1993) pp. 404-410 Summary of the Invention [Means for solving the problem]

[0009] In general, the present invention is based on the studies described in the Examples, which have led to surprising findings regarding aqueous formulations of amylin analogues that are particularly suitable for long-term storage, in particular, low buffer concentrations and specific pH ranges have been found to be unexpectedly beneficial for optimizing stability.

[0010] The present invention relates to [19CD]-isoGlu-RD()GTATK()ATERLA-Aad-FLQRSSF-Gly(Me)-A-Ile(Me)-LSSTEVGSNT-Hyp-NH2 or a pharma- ceutically acceptable salt and / or derivative thereof. and providing a stable aqueous liquid pharmaceutical formulation comprising: The formulation is (a) an amylin analog at a concentration of about 0.4 mg / ml to about 25 mg / ml, and (b) a buffer having a concentration of about 0.5 mM to about 25 mM Including, The formulation has a pH of about 5.8 to about 6.9.

[0011] The amylin analog is present in 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 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.

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

[0015] Any suitable buffer may be used, 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. 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 is increased at relatively low buffer concentrations, i.e., 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 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, particularly (but not limited to) when the buffer is a phosphate, histidine, or citrate, it 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.

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

[0019] The pH of the formulation was also found to be significant for stability. Although the compounds described in WO2018 / 046719 were thought to be stable at physiological pH (e.g., about pH 7.4), it was surprisingly found that chemical stability was increased at lower pH values, while physical stability was maintained. Thus, the formulation may have 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.

[0020] The stable aqueous liquid formulation may include a tonicity adjuster. The tonicity adjuster may be ionic or non-ionic. Suitable ionic tonicity adjusters include alkali metal salts (e.g., halides) and earth metal salts (e.g., halides), such as NaCl, NaBr, NaI, KCl, KBr, KI, LiCl, CaCl2, and Na2SO4. Non-ionic tonicity adjusters include mannitol (e.g., D-mannitol), propylene glycol, sucrose, glycerol, sorbitol, and trehalose.

[0021] In some embodiments, mannitol (eg, D-mannitol), propylene glycol, and NaCl, especially mannitol and propylene glycol, may be preferred as tonicity adjusting agents.

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

[0023] For example, the formulation does not substantially exhibit changes indicative of reduced physical stability (e.g., as demonstrated by solution turbidity, or aggregation, fibrillation, or gelation of the amylin analogue) after storage 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, the formulation does not substantially exhibit such changes after storage at 2-8° C. (e.g., at 5° C.) for at least 18 months, such as at least 24 months. It will be appreciated that accelerated conditions can also be employed as a surrogate for the assessment of longer term physical stability, as provided in more detail below.

[0024] Additionally 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 analogue remains in its original monomeric form in the formulation after storage 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 analogue remains in its original monomeric form in the formulation after storage 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 analogue remains in its original monomeric form in the formulation after storage at 2-8° C. (e.g., at 5° C.) for at least 18 months, such as at least 24 months.

[0025] The formulation may further comprise a preservative, particularly if it is provided in a multi-use format, or alternatively, the formulation may be free or substantially free of a preservative, particularly if it is provided in a single-use format.

[0026] The stable aqueous liquid formulations of the present invention may be formulated for parenteral administration, in particular 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 free (or substantially free, e.g., greater than about 100 ppm) of reducing agents.

[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, such as 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 (e.g. 1.0 ml). As described in more detail below, the individual dosages may be packaged separately, for example in individual vials, cartridges, or syringes, for single use.

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

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

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

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

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

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

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

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

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

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

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

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

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

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

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

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

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

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

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

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

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

[0050] amylin analog at a concentration of about 18 mg / ml, 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 about 0.8 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;

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

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

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

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

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

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

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

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

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

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

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

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

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

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

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

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

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

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

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

[0071] amylin analog at a concentration of about 18 mg / ml, 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 dosage volume of 0.5 ml, although it will be appreciated that they may be used for other dosage volumes, or the concentration of the amylin analogue may be adjusted to suit various dosage volumes.

[0073] Further certain formulations may comprise or consist of the following:

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

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

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

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

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

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

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

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

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

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

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

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

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

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

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

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

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

[0091] amylin analog at a concentration of about 9 mg / ml, 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] amylin analog at a concentration of about 1.2 mg / ml, histidine buffer at a concentration of about 10 mM, and mannitol at a concentration of about 250-260 mM; pH 6.5;

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

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

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

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

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

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

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

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

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

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

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

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

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

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

[0109] amylin analog at a concentration of about 9 mg / ml, 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 1.0 ml dosage volume, although it will be appreciated that they may be used for other dosage volumes, or the concentration of the amylin analogue may be adjusted for different dosage volumes.

[0111] In these formulations as well as elsewhere, the amylin analog may be, for example, a compound having the following formula: ([19CD]-isoGlu-RD()GTATK()ATERLA-Aad-FLQRSSF-Gly(Me)-A-Ile(Me)-LSSTEVGSNT-Hyp-NH2), x(Cl) where x is 1.0 to 2.0.

[0112] The present invention further provides a container or delivery device that contains the stable aqueous liquid formulation described herein.For example, suitable containers and delivery devices include sealed vials, pre-filled syringes, or cartridges for injection devices such as injection pens, adjustable dose auto-injectors, disposable auto-injectors, wearable injectors, or infusion pumps.Delivery devices can be single-use devices such as pre-filled syringes.

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

[0114] The present invention further provides the stable aqueous liquid formulations described for use in methods 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 to treat various conditions, diseases, or disorders in a subject, including, but not limited to, obesity and various conditions, diseases, or disorders related to obesity, such as diabetes (e.g., type 2 diabetes), hypertension, dyslipidemia, sleep apnea, and cardiovascular disease. The subject may suffer from at least one weight-related comorbidity condition, such as obesity associated with diabetes (e.g., type 2 diabetes), hypertension, dyslipidemia, sleep apnea, and cardiovascular disease. It will thus be understood that the formulation may be administered to a subject suffering from 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 may be used to treat a combination of the conditions described.

[0116] That is, the invention provides a formulation of the 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 control of appetite, feeding, food intake, caloric intake, and / or energy expenditure.

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

[0118] The invention also provides formulations of the invention for use in a method of prevention or treatment of Alzheimer's disease, diabetes, type 1 diabetes, type 2 diabetes, prediabetes, insulin resistance syndrome, impaired glucose tolerance (IGT), disease states associated with high blood glucose levels, metabolic diseases including metabolic syndrome, hyperglycemia, hypertension, atherogenic dyslipidemia, hepatic steatosis (per se "fatty liver" including non-alcoholic fatty liver disease (NAFLD) including non-alcoholic steatohepatitis (NASH)), renal failure, arteriosclerosis (e.g. atherosclerosis), macrovascular disease, small vessel 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 invention also provides a formulation of the invention for use in a method of lowering circulating LDL levels and / or increasing the HDL / LDL ratio.

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

[0121] The invention further provides the use of a formulation of the 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 medicaments for treating obesity and related diseases, disorders, and conditions, including but not limited to morbid obesity, pre-surgical obesity, obesity-related inflammation, obesity-related gallbladder disease, obesity-induced sleep apnea and breathing disorders, cartilage degeneration, osteoarthritis, and reproductive health complications such as infertility due to obesity or overweight.The subject may suffer from at least one weight-related comorbidity condition, such as obesity associated with diabetes (e.g., type 2 diabetes), hypertension, dyslipidemia, sleep apnea, and cardiovascular disease.

[0123] The invention also provides use of a formulation of the invention in the manufacture of a medicament for the prevention or treatment of Alzheimer's disease, diabetes, type 1 diabetes, type 2 diabetes, prediabetes, insulin resistance syndrome, impaired glucose tolerance (IGT), disease states associated with high blood glucose levels, metabolic diseases including metabolic syndrome, hyperglycemia, hypertension, atherogenic dyslipidemia, hepatic steatosis (per se "fatty liver" including non-alcoholic fatty liver disease (NAFLD) including non-alcoholic steatohepatitis (NASH)), renal failure, arteriosclerosis (e.g. atherosclerosis), macrovascular disease, small vessel 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 invention also provides the use of a formulation of the invention in the manufacture of a medicament for lowering circulating LDL levels and / or increasing the HDL / LDL ratio.

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

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

[0127] The invention also provides methods 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 high blood glucose levels, metabolic diseases including metabolic syndrome, hyperglycemia, hypertension, atherogenic dyslipidemia, hepatic steatosis (per se "fatty liver" including non-alcoholic fatty liver disease (NAFLD) including non-alcoholic steatohepatitis (NASH)), renal failure, arteriosclerosis (e.g. atherosclerosis), macrovascular disease, small vessel 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 in a subject, comprising the step of administering to the subject a therapeutically effective amount of a formulation of the invention.

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

[0129] The present invention further provides the use of the above formulations in a method for cosmetic (ie non-therapeutic) weight loss.

[0130] It will be understood that references to therapeutic uses of the formulations and methods involving administration of the formulations should be equally interpreted as embracing cosmetic uses and administration.

[0131] The present invention relates to [19CD]-isoGlu-RD()GTATK()ATERLA-Aad-FLQRSSF-Gly(Me)-A-Ile(Me)-LSSTEVGSNT-Hyp-NH2 or a pharma- ceutically acceptable salt and / or derivative thereof. The present invention further provides a method for producing a stable aqueous pharmaceutical formulation comprising: The method comprises: (a) an amylin analog, salt, or derivative at a concentration of about 0.4 mg / ml to about 25 mg / ml, and (b) a buffer having a concentration of about 0.5 mM to about 25 mM to produce a stable aqueous liquid formulation, wherein the formulation has a pH of about 5.8 to about 6.9.

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

[0133] Thus, for example, the method may further comprise the step of formulating the amylin analogue and buffer with a tonicity adjusting agent as described.

[0134] The method may include a step of adjusting the pH to a desired value. If the tonicity adjusting agent is a chloride salt, such as NaCl, it may be desirable to adjust the pH to a desired value before adding the tonicity adjusting agent to the formulation. This is particularly the case when the amylin analogue is itself provided 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 analogue at pH values ​​near the pI of the peptide (pI=4.36).

[0135] Formulations produced by the described methods are also provided.

[0136] Unless the context dictates otherwise, the feature descriptions and definitions provided above are not limited to any particular aspect or embodiment of the present invention, but apply equally to all aspects and embodiments described.

[0137] Throughout this specification the term "about" in reference to numerical values ​​is optional and means, for example, + / - 10%. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

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

[0139] Amylin Analogues The amylin analogue present in the formulations of the invention (Compound 1; "CPD 1") is described in WO2018 / 046719 and has the formula: [19CD]-isoGlu-RD()GTATK()ATERLA-Aad-FLQRSSF-Gly(Me)-A-Ile(Me)-LSSTEVGSNT-Hyp-NH2 has. Gly(Me): N-methylglycine [also known as sarcosine (Sar)] Ile(Me): N-methylisoleucine Aad: 2-aminoadipic acid, e.g. (2S)-2-aminoadipic acid [(2S)-2-aminohexanedioic acid], also known as homoglutamic acid. [19CD]-: 19-carboxy-nonadecanoyl-

[0140] The parentheses "()" following the symbols of particular amino acid residues indicate the residue whose side chain contributes to an intramolecular lactam bridge, i.e., the amylin analog compounds present in the formulations of the invention have an intramolecular lactam bridge formed between the side chains of the residues indicated in parentheses (aspartic acid and lysine, respectively).

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

[0142] Thus, [19CD]-isoGlu- denotes a 19-carboxy-nonadecanoyl group covalently attached via an amide linkage to the alpha amino group of a Glu linker, the side chain carboxyl group of which is in turn linked via an amide linkage to the backbone alpha amino group of an arginine residue at the N-terminus of the peptide chain of the amylin analog.

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

[0144] [ka]

[0145] It should be understood that the amylin analog (active agent) may also be provided in the form of a salt or other derivative. A salt is typically a "pharmaceutically acceptable" salt, meaning in the context of the present invention that is not harmful to the patient or subject being treated therewith. Such salts are generally acid addition salts or base 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 base salts include salts in which the cation is an alkali metal ion, such as sodium and potassium, an alkaline earth metal ion, such as calcium, and an ion such as NR(R')3. + C, wherein R and R' are independently optionally substituted 1-6 Alkyl, optionally substituted C 2-6 Included are salts selected from substituted ammonium ions designated alkenyl, optionally substituted aryl, or optionally substituted heteroaryl. Other examples of pharma- ceutically acceptable salts are described in Remington's Pharmaceutical Sciences, 17th Edition, Alfonso R. Gennaro (ed.), Mack Publishing Company, Easton, PA, USA, 1985 and more recent editions, and in the 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-NH2), x(Cl) where x is between 1.0 and 2.0.

[0147] Other derivatives of amylin analogues include Mn 2+ and Zn 2+ The prodrug derivatives include coordination complexes with metal ions such as aryl ions, esters such as in vivo hydrolyzable esters, free acids or bases, hydrates, prodrugs, or lipids. Esters can be formed between hydroxyl or carboxylic acid groups present in the compound and suitable carboxylic acid or alcohol reaction partners using techniques well known in the art. Prodrug derivatives 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 conversion of the prodrug to release the compound or its metabolites. Examples of prodrugs include the use of protective groups that are removed in situ to release the active compound or act to inhibit the elimination of the drug in vivo.

[0148] Amylin analogs are agonists of amylin receptors and have agonist activity at the receptors hAMYR1, hAMYR2, and hAMYR3 (human amylin receptors 1, 2, and 3), as described in WO2018 / 046719. Binding to a suitable receptor induces intracellular signaling, such as the production of cyclic AMP. In vivo, amylin analogs have (among other things) the biological activity of reducing food intake, promoting weight loss, and / or preventing or reducing weight gain. This may be employed in a variety of therapeutic applications, as described elsewhere herein and in WO2018 / 046719.

[0149] Aqueous Liquid Formulations Preferably, the formulations of the present invention are substantially free of organic solvents, hi 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 that contain water in the form of an aqueous solution. In the context of the present invention, the term "aqueous formulation" generally means a formulation that contains at least 50% by weight (50% w / w) water as a solvent, more preferably at least 75% w / w water, more preferably at least 80% w / w water, more preferably at least 85% w / w water, more preferably at least 90% w / w water, and most preferably at least 95% w / w water. In certain embodiments, the aqueous formulations of the present invention are substantially free of organic solvents, such as aprotic polar solvents, such as dimethylsulfoxide (DMSO). "Substantially" in this sense means that the aqueous formulation contains less than 5% by volume of organic solvent, more preferably less than 2% by volume (v / v), and even more preferably less than 1% by volume (v / v). 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 ingredients and amounts of the liquid formulations of the present invention are selected to provide the formulation with 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, for example about 6.5. For the avoidance of doubt, pH is measured at 25°C.

[0152] The formulations of the present invention may further comprise a preservative, particularly when provided in a multi-use format. Alternatively, the formulations may be free or substantially free of preservatives, particularly when provided in a single-use format. If a preservative is present, it will be employed in an amount sufficient to kill the range of bacterial types 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 range of bacterial types required for regulatory approval.

[0153] Formulations of the present invention are typically suitable for parenteral administration by injection, i.e. administration may be by injection, for example subcutaneous, intramuscular, intraperitoneal or intravenous, although subcutaneous administration may be preferred.

[0154] The formulations of the 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 use.

[0155] Examples include sealed vials, pre-filled syringes, and injection devices such as injection pens, adjustable-dose auto-injectors, disposable auto-injectors, wearable injectors, 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. A variety of analytical techniques for measuring protein stability are available in the art and are reviewed, for example, in Peptide and Protein Drug Delivery, pp. 247-301, Vincent Lee, ed., Marcel Dekker, Inc., New York, NY, Pubs. (1991) and Jones, A. Adv. Drug Delivery Rev. 10: pp. 29-90 (1993). The chemical and physical stability of the amylin analog in any given formulation can be determined according to the assays outlined in the Examples below.

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

[0158] It is also possible to test stability under accelerated or extreme conditions, where an increased storage temperature is typically used to assess 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 assess stability over a short period of time, such as 1, 2, 3, or 4 weeks, 1 month, 2 months, 3 months, or 6 months.

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

[0160] The physical stability (e.g., turbidity of the solution, or aggregation, fibrillation, or gelation of the amylin analogue) can be qualitatively and / or quantitatively assessed in a variety of different ways, e.g., by visual inspection, by size-exclusion chromatography, UV light scattering, dynamic light scattering (DLS), circular dichroism, by measuring turbidity, or using spectroscopic probes (such as Thioflavin T) that indicate the conformational state of the peptide. The probes are preferably small molecules that preferentially bind to non-native conformers of the peptide. Thioflavin T (ThT) is a fluorescent dye that has been widely used to detect amyloid fibrils. In the presence of fibrils and possibly other peptide configurations, Thioflavin T produces a new excitation maximum at about 450 nm and enhanced emission at 482 nm when bound to the fibrillar form of the peptide. Unbound Thioflavin T is essentially non-fluorescent at the wavelengths of interest.

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

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

[0163] The term "chemical stability" as used herein refers to the stability of the peptide with respect to covalent / chiral chemical alterations of the peptide structure that result in the formation of chemical degradation products and / or covalently linked oligomers with potentially reduced biological potency compared to the intact monomeric peptide structure. The primary degradation products from the amylin analogues present in the formulations of the present invention are believed to result from deamidation of the asparagine side chain and / or the C-terminal amide group to form free carboxylic acid groups. Covalently linked oligomers are also believed to form in suboptimal formulations.

[0164] The chemical stability of a formulation can be assessed by measuring the amount of intact monomeric peptide, specific chemical degradation products, or covalent oligomers remaining 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 intact monomeric peptide remaining can be a preferred indicator of chemical stability. Separation is usually performed based on molecular size and / or charge using various chromatographic techniques such as size exclusion chromatography (SEC) or reversed-phase (RP) chromatography, typically using 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 storage 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 storage 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 analogue remains in its original monomeric form in the formulation after storage at 2-8°C (e.g., at 5°C) for at least 18 months, such as at least 24 months.

[0166] Buffer As used herein, the term "buffering agent" refers to a pharma- ceutically acceptable agent that stabilizes the pH of a pharmaceutical formulation. Suitable buffering agents are well known in the art and can be found in the literature.

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

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

[0169] The buffering agent 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, for example, about 1 mM to about 12 mM. For example, the buffering agent 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, this is preferably present at about 15 mM to about 25 mM, such as about 17 mM to about 23 mM, e.g., about 20 mM, since the buffering capacity of TRIS is lower than that of other buffers such as (for example) phosphate, histidine, and citrate.

[0171] It is possible for a formulation to contain more than one buffer, for example, two or more buffers. In such cases, the concentrations cited 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 a total of 5 mM buffer.

[0172] Tonicity Adjustment Agent The term "tonicity adjusting agent" as used herein means a pharma- ceutically acceptable tonicity agent used to adjust the tonicity of a formulation. The terms "tonicity adjusting agent" and "tonicity agent" may be used interchangeably. The formulations of the present invention are preferably isotonic, i.e., they have an osmolarity substantially the same as human serum, e.g., about 300+ / -80 mOsm, e.g., 300+ / -60 mOsm, as measured by an osmometer.

[0173] The tonicity adjuster may be ionic or non-ionic. Suitable ionic tonicity adjusters include alkali metal salts (e.g., halides) and earth metal salts (e.g., halides), such as NaCl, NaBr, NaI, KCl, KBr, KI, LiCl, CaCl2, and Na2SO4. Non-ionic tonicity adjusters include mannitol (e.g., D-mannitol), propylene glycol, sucrose, glycerol, sorbitol, trehalose, and dextrose.

[0174] If the tonicity modifier is a chloride salt such as NaCl, it may be desirable to add the tonicity modifier to the formulation only after the pH has been adjusted to the desired value. This is particularly true if the amylin analog is itself provided 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).

[0175] In some embodiments, mannitol (eg, D-mannitol), propylene glycol, and NaCl, especially mannitol (eg, D-mannitol) and propylene glycol, may be preferred as tonicity adjusting agents.

[0176] It will be understood that the concentration of the tonicity adjusting agent will depend on the concentrations of the other components of the formulation, particularly if the formulation is intended to be isotonic. One skilled in the art will be able to establish the appropriate concentration for any given agent for a particular formulation.

[0177] For example, it may be appropriate to use mannitol at a concentration of 200 to 300 mM, for example, 230 to 260 mM. It may be appropriate to use propylene glycol at a concentration of 200 to 250 mM, for example, 224 mM. It may be appropriate to use NaCl at a concentration of 100 to 150 mM, for example, 118 to 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. Thus, the formulations may provide attractive treatment options, inter alia, for metabolic disorders caused by, characterized by, or associated with obesity and overweight.

[0179] That is, the formulations may be used in methods to treat, prevent or reduce weight gain, promote weight loss, reduce food intake, and / or reduce overweight. Treatment may be accomplished, for example, by control of appetite, feeding, food intake, calorie intake, and / or energy expenditure.

[0180] The formulations may be used in methods for treating obesity and related diseases, disorders, and conditions, including but not limited to, morbid obesity, pre-surgical obesity, obesity-related inflammation, obesity-related gallbladder disease, obesity-induced sleep apnea and breathing disorders, cartilage degeneration, osteoarthritis, and reproductive health complications such as infertility due to obesity or overweight.

[0181] The formulations may be used in methods of prevention or treatment of Alzheimer's disease, diabetes, type 1 diabetes, type 2 diabetes, prediabetes, insulin resistance syndrome, impaired glucose tolerance (IGT), disease states associated with high blood glucose levels, metabolic diseases including metabolic syndrome, hyperglycemia, hypertension, atherogenic dyslipidemia, hepatic steatosis (per se "fatty liver", including non-alcoholic fatty liver disease (NAFLD), including non-alcoholic steatohepatitis (NASH)), renal failure, arteriosclerosis (e.g. atherosclerosis), macrovascular disease, small vessel disease, diabetic heart disease (including diabetic cardiomyopathy and heart failure as a complication of diabetes), coronary heart disease, peripheral arterial disease or stroke.

[0182] The formulations may also be useful in lowering 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 effects on body weight.

[0184] Metabolic syndrome is characterized by a group of metabolic risk factors in a person. The metabolic risk factors include abdominal obesity (excess 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 plaque in arterial walls), elevated blood pressure (hypertension), insulin resistance and glucose intolerance, prothrombotic states (e.g. high fibrinogen or plasminogen activator inhibitor-1 in the blood), and proinflammatory states (e.g. elevated C-reactive protein in the blood). Individuals with metabolic syndrome are at increased risk for other diseases (e.g. stroke and peripheral vascular disease) associated with coronary heart disease and other manifestations of atherosclerosis. The major risk factor underlying this syndrome appears to be abdominal obesity.

[0185] The term "treatment" (as well as "treating" and other grammatical variations) employed in the context of the present invention refers to an approach for obtaining beneficial or desired clinical outcomes. For purposes of the present invention, beneficial or desired clinical outcomes include, but are not limited to, detectable or undetectable alleviation of symptoms, attenuation of the extent of the disease, stabilization of the disease state (i.e., not worsening), delay or slowing of disease progression, improvement or mitigation of the disease state, and remission (partial or complete). "Treatment" can also mean prolonging survival compared to expected survival in the absence of treatment. "Treatment" is an intervention performed with the intent of preventing the progression of a disorder or altering its pathology. Thus, "treatment" refers to both therapeutic treatment and prophylactic or preventative measures. When used in the context of prophylactic or preventative measures, the pharmaceutical preparation may not completely prevent the progression of the disease or disorder. Those in need of treatment include those already suffering from the disorder as well as those in whom the progression of the disorder is to be prevented. "Treatment" also means preventing or reducing the increase in a disease state or symptom (e.g., weight gain or hypoglycemia) compared to the absence of 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 invention and are not intended to limit the scope of the invention.

[0187] method Preparation of Compound 1 (CPD 1) Formulations (Method I) Compound 1 (described in 2018 / 046719) was prepared as the chloride salt. Except where noted below, formulations of CPD 1 were prepared by dissolving the CPD 1 peptide in alkaline MilliQ water (MQW) (pH adjusted with NaOH) to generate a clear drug substance (DS) stock solution of approximately pH 7 at a concentration relevant to the target end product concentration. Relevant excipients were dissolved in MQW at the appropriate pH to form excipient solutions. The two solutions were mixed and the pH was measured and adjusted as necessary with diluted NaOH / HCl to reach the desired pH. Finally, the appropriate amount of MQW was added to reach the final product concentration. The formulations were sterile 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 methods.

[0189] Determination of Purity (%) by RP-HPLC (Assay IIa) A Dionex Ultimate 3000 HPLC system was used for the analysis with a flow rate of 0.3 mL / min. The mobile phase 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 amount was 4 μg of peptide. The column used for the HPLC analysis was a Waters Acquity UPLC BEH C18, 150 × 2.1 mm, 1.7 μm particle size. The run time was 35–37 min depending on the reconditioning 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 expressed herein as the relative purity of the peptide peak (i.e. the main peptide peak) normalized to the absolute purity of the peptide peak (i.e. the main peptide peak) at day zero (day 0 (D0)), which is set to 100% purity, and therefore the chemical stability is expressed in % of CPD 1.

[0191] Purity is a measure of the remaining intact compound in the stressed solution after incubation, measured on day 0 (D0, day zero) and expressed as % normalized purity of CPD 1.

[0192] Determination of the formation of covalent oligomers by size exclusion chromatography (SEC) (assay IIb) A Dionex Ultimate 3000 HPLC system was used for the analysis with a flow rate of 0.5 mL / min (isocratic elution). The mobile phase 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 amount was 4 μg of peptide. The column used for the HPLC analysis was a Tosoh BioScience TSKgel SuperSW2000, 30 cm × 4.6 mm, 4 μm particle size. The run time was 25 min.

[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 main intact monomer peak of CPD-1 and is expressed as % oligomer.

[0194] Methods for assessing physical stability (Assay III) To assess the physical stability of CPD 1 in the formulation, the following method was used.

[0195] Visual Inspection of Solutions (Assay IIIa) Samples were provided in transparent containers to allow visual inspection of the formulation by trained and experienced technicians. Visual inspection was performed manually and recorded by an experienced technician. Samples were classified as "clear", "suspended", "gel", or "particulate" depending on their appearance.

[0196] UV absorbance at 325 nm (Assay IIIb) Turbidity was measured by UV absorbance analysis. 100 μL of each sample was loaded into a Corning clear 96-well UV-compatible plate. Samples were read at 325 nm on a SpectraMax 190 Elisa reader.

[0197] Dynamic Light Scattering (DLS) (Assay IIIc) Peptide particle size distribution was determined by dynamic light scattering analysis. Standard DLS experiments were performed in a Nunc 96-well plate with a sample volume of 165 μL and measured with a DynaPro Plate reader II instrument. Typical parameters used were: temperature 25 °C, acquisition time 2 s, 50 acquisitions per sample, detection: hydrodynamic radius Rh, correlation function cutoff value [μs]: 0.5–1 × 10 5 , peak radius cutoff [nm]: 0.5-1000.

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

[0199] A 1 mM ThT stock solution was prepared and 12 μL was mixed with 288 μL of CPD 1 formulation. 2×150 μL of sample was loaded in duplicate into a 96-well black fluorescent plate (clear bottom). Data were collected at fixed intervals of 10 min up to 60 h at 40° C. on a Biotek Synergy™ H4 Hybrid Multi-Mode fluorescent plate reader after 300 s of automated mixing (agitation) each. Physical stability was expressed as the lag time (in hours) for fibril formation and was defined as the intersection of two linear regressions representing the initial stable and growth phases.

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

[0201] 170 μL of CPD 1 formulation samples were loaded into a Nunc black 96-well plate with optical bottom. 7 μL of 1 mM ThT was further added and mixed by gently pipetting up and down to ensure uniform distribution of ThT. The 96-well plate was placed in a Fluorescence Plate reader Biotek Synergy™ H4 Hybrid Multi-Mode or 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 at 25°C of formulations in the pH range 6.1-7.8 The formulations listed in Table 1 were prepared as described in Method I except as noted in the table.

[0203] [Table 1]

[0204] The prepared formulations were filled into 1 ml type 1 glass vials and stored at 25° C., protected from light, for the indicated periods before analysis.

[0205] [Table 2]

[0206] There is no clear effect of pH on physical stability, but purity increases with decreasing pH, see Table 2.

[0207] Example 2 Effect of formulation pH on physical stability CPD 1 drug product formulations containing 1 mg / ml CPD 1, 20 mM TRIS (buffering agent) and 118 mM NaCl (tonicity agent) were prepared as described in Method I at pH 5.8, 6.0, 6.3, 6.8, and 7.0.

[0208] Physical stability was assessed in terms of fibril formation by ThT stress test measured over 60 h at 40° C. 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, which affects peptide particle size.

[0211] Example 3 Accelerated chemical and physical stability of formulations with buffer concentrations between 2mM and 20mM at 25°C. The formulations listed in Table 4 were prepared as described in Method I. The prepared formulations were filled into 1 ml type 1 glass vials and stored at 25° C., protected from light, for the indicated periods before analysis.

[0212] [Table 4A]

[0213] [Table 4B]

[0214] Purity is higher at 5 mM histidine and phosphate buffers than at 20 mM, as exemplified by formulations 1-4. No or limited effect on visual appearance, physical stability, and covalent oligomers was observed for the 5 mM and 20 mM histidine and phosphate buffer concentrations.

[0215] As exemplified by formulations 7-8, the beneficial effect of lowering the buffer concentration to 5 mM phosphate is maintained at 2 mM phosphate.

[0216] Example 4 Accelerated physical stability in the pH range 6.1-6.9 CPD 1 drug product bioformulations #1-3 from Table 1 (Example 1) were filled into 1 ml type 1 glass vials and subjected to physical stress in the form of constant rotation at approximately 15 rpm (revolutions per minute) at room temperature for the indicated periods on a multirotator located in a standard laboratory with window sunlight (no light shielding). Stability was determined as shown in the table.

[0217] [Table 5]

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

[0219] Example 5 CPD 1 Chemical stability or solubility of formulations with concentrations of 0.4mg / mL to 50mg / mL A 4 mg / ml formulation of CPD 1 containing 20 mM TRIS and 118 mM 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 24 hours of storage at 25° C.

[0220] [Table 6]

[0221] There was no loss of purity after storage at concentrations as low as 0.4 mg / ml, see Table 7.

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

[0223] [Table 7]

[0224] CPD 1 could be dissolved in MQW at concentrations up to 50 mg / ml to give a clear solution, see Table 8.

[0225] Example 6 Accelerated chemical stability of formulations at pH 6 and buffer concentrations of 0.5-40 mM at 40°C The formulations in Tables A1-A3 were prepared according to Method I and filled into 1.5 ml LowBind Eppendorf tubes. No tonicity agent was included.

[0226] After 4 weeks of storage at 40°C, stability was tested according to Assay II with a modification (Assay IIb) that results in the formation of covalently linked oligomers (SECs), reported as the difference in total oligomers between the initial value and the 4 week time point.

[0227] [Table 8]

[0228] [Table 9]

[0229] [Table 10]

[0230] At pH 6, the loss of purity is reduced with decreasing buffer concentration (Table 9). Buffer concentration has no significant effect on the formation of covalent oligomers (Table 10).

[0231] 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 Table 11 with Table 9 and Table 10, formulations 29-32).

[0232] Example 7 Accelerated chemical and physical stability of formulations containing various concentrations of Compound 1 at 25°C The formulations in Table 12 were prepared according to Method I and filled into 1 ml Type 1 glass cartridges fitted with siliconized bromobutyl plungers. After storage at 25° C. for the indicated periods, the stability was tested according to Assay II and Assays IIIa, IIIb, and IIIc.

[0233] [Table 11]

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

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

[0236] [Table 12]

[0237] The loss of purity appears to be less in formulations containing mannitol and propylene glycol as tonicity agents than in formulations containing glycerol and NaCl as tonicity agents. The formation of covalent oligomers appears to be greater in formulations containing glycerol as tonicity agent than in formulations containing other tonicity agents.

[0238] The following paragraphs present certain specific embodiments of the present invention. 1.[19CD]-isoGlu-RD()GTATK()ATERLA-Aad-FLQRSSF-Gly(Me)-A-Ile(Me)-LSSTEVGSNT-Hyp-NH2 or a pharma- ceutically acceptable salt and / or derivative thereof. A stable aqueous liquid 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 having a concentration of about 0.5 mM to about 25 mM Including, A stable aqueous liquid formulation having a pH of about 5.8 to about 6.9. 2. The stable aqueous liquid formulation of paragraph 1, wherein the amylin analog is present in 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 of 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. A stable aqueous liquid formulation according to any one of paragraphs 1 to 3, wherein the amylin analogue is present in a concentration of from about 3 mg / ml to about 12 mg / ml, for example from about 3 mg / ml to about 7 mg / ml, or from about 8 mg / ml to about 12 mg / ml, for example from 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 buffering agent is 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, for example from about 1 mM to about 15 mM, such as from about 1 mM to about 12 mM. 6. The stable aqueous liquid formulation of any one of paragraphs 1 to 5, wherein the buffering agent 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 of 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, for example about 20 mM. 9. The stable aqueous liquid formulation according to any one of paragraphs 1 to 8, wherein the pH is from about 6.2 to about 6.8, such as from about 6.4 to about 6.6, for example about 6.5. 10. The stable aqueous liquid formulation of any one of paragraphs 1 to 9, further comprising a tonicity adjuster. 11. The stable aqueous liquid formulation of 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. The stable aqueous liquid formulation of any one of paragraphs 1 to 11, wherein the formulation 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. 13. A stable aqueous liquid formulation according to any one of paragraphs 1 to 12, 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. 14. The stable aqueous liquid formulation of any one of paragraphs 1 to 13, wherein the amylin analog is provided as a chloride salt. 15. The stable aqueous liquid formulation of any one of paragraphs 1 to 14, wherein water is the only solvent used to make the aqueous liquid formulation. 16. The stable aqueous liquid formulation of any one of paragraphs 1 to 15, formulated for administration to a subject by injection. 17. The stable aqueous liquid formulation according to paragraph 16, wherein the injection is a subcutaneous injection. 18. The stable aqueous liquid formulation of any one of paragraphs 1 to 17, which is sterile. 19. A container or delivery device comprising a liquid formulation according to any one of paragraphs 1 to 18. 20. A stable aqueous liquid formulation according to any one of paragraphs 1 to 18 for use in a method of medical treatment. 21. The stable aqueous liquid formulation of any one of paragraphs 1 to 18 for use in a method for treating, preventing or reducing weight gain, promoting weight loss and / or reducing overweight. 22. The stable aqueous liquid formulation of any one of paragraphs 1 to 18 for use in a method for treating obesity (e.g. morbid obesity, pre-surgical obesity), obesity-related inflammation, obesity-related gallbladder disease, obesity-induced sleep apnea, cartilage degeneration, osteoarthritis, or reproductive health complications due to obesity or being overweight. 23. The stable aqueous liquid formulation of any one of paragraphs 1 to 18 for use in a method for the prevention or treatment of 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 fatty liver disease (NASH)), renal failure, arteriosclerosis (e.g. atherosclerosis), macrovascular disease, small vessel disease, diabetic heart disease (e.g. diabetic cardiomyopathy), coronary heart disease, peripheral arterial disease or stroke. 24. The stable aqueous liquid formulation of any one of paragraphs 1 to 18 for use in a method for lowering circulating LDL levels and / or increasing the HDL / LDL ratio. 25.[19CD]-isoGlu-RD()GTATK()ATERLA-Aad-FLQRSSF-Gly(Me)-A-Ile(Me)-LSSTEVGSNT-Hyp-NH2 or a pharma- ceutically acceptable salt and / or derivative thereof. 1. A method for producing a stable liquid 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 having a concentration of about 0.5 mM to about 25 mM to produce a stable aqueous liquid formulation, wherein the formulation has a pH of about 5.8 to about 6.9. 26. A formulation produced by the method of paragraph 25, which is a stable aqueous liquid formulation according to 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 and / or derivative thereof. A stable aqueous liquid pharmaceutical formulation comprising: (a) an amylin analog at a concentration of about 0.4 mg / ml to about 25 mg / ml, and (b) a buffering agent at a concentration of about 0.5 mM to about 25 mM Including, A stable aqueous liquid formulation having a pH of about 5.8 to about 6.

9.

2. 2. The stable aqueous liquid formulation of claim 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. 3. The stable aqueous liquid formulation of claim 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. 4. The stable aqueous liquid formulation of claim 1, wherein the amylin analogue is present in 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. 5. The stable aqueous liquid formulation according to any one of claims 1 to 4, wherein the buffering agent 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 about 15 mM, such as about 1 mM to about 12 mM.

6. 6. The stable aqueous liquid formulation of any one of claims 1 to 5, wherein the buffering agent 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. 7. The stable aqueous liquid formulation of claim 1, wherein the buffer is phosphate, histidine, or citrate.

8. 7. The stable aqueous liquid formulation of any one of claims 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. 9. The stable aqueous liquid formulation according to any one of claims 1 to 8, having a pH of about 6.2 to about 6.8, such as about 6.4 to about 6.6, such as about 6.

5.

10. 10. The stable aqueous liquid formulation of any one of claims 1 to 9, further comprising a tonicity adjuster.

11. 11. The stable aqueous liquid formulation of any one of claims 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. 12. The stable aqueous liquid formulation of any one of claims 1 to 11, 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.

13. 13. The stable aqueous liquid formulation of any one of claims 1 to 12, wherein 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 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.

14. 14. The stable aqueous liquid formulation of any one of claims 1 to 13, wherein the amylin analog is provided as a chloride salt.

15. 15. The stable aqueous liquid formulation of any one of claims 1 to 14, formulated for administration to a subject by injection, for example wherein the injection is a subcutaneous injection.

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