Pharmaceutical formulations comprising a cyclodextrin, a glucagon-like peptide-1 receptor agonist and a glucagon receptor agonist

Abstract

Disclosed herein is a preserved liquid pharmaceutical formulation comprising a amylin receptor agonist, a GLP-1 receptor agonist, a hydroxypropyl-substituted cyclodextrin, and one or more preservatives. The co-formulation can be used for medical treatment of subjects with: overweight or obesity, with or without associated co-morbidities; diabetes, with or without associated co-morbidities; and cardiovascular disease.

Description

Technical Field

[0001] This invention relates to a pharmaceutical formulation, which is a co-formulation of a GLP-1 receptor agonist and an amylin receptor agonist. The pharmaceutical formulation may be used for the medical treatment of subjects suffering from: overweight or obesity with or without one or more related comorbidities; diabetes with or without one or more related comorbidities; and / or one or more cardiovascular diseases. Background Technology

[0002] Smegglutide is a glucagon-like peptide-1 (GLP-1) receptor agonist and the active pharmaceutical ingredient in Ozempic®. Ozempic® is indicated for (i) as an adjunct to diet and exercise to improve glycemic control in adults with type 2 diabetes and (ii) to reduce the risk of major adverse cardiovascular events in adults with type 2 diabetes and a history of cardiovascular disease.

[0003] Smegglutinin is also the active pharmaceutical ingredient in Wegovy®. Wegovy® is suitable for individuals with an initial body mass index (BMI) greater than or equal to 30 kg / m². 2 or greater than 27 kg / m 2 In adult patients with at least one weight-related comorbidity, it is used as an adjunct to low-calorie diets and increased physical activity for long-term weight management.

[0004] Ozempic® and Wegovy® are liquid pharmaceutical formulations containing 8 mM phosphate and with a pH of approximately 7.4.

[0005] Fixed-dose combinations of the amylin receptor agonist canagliptin and the GLP-1 receptor agonist smegglutide, used to treat overweight and obesity, have been investigated (Lancet 2021; 397:1736–48). The pharmaceutical products investigated are single liquid formulations containing either canagliptin or smegglutide for subcutaneous use.

[0006] To date, it has been considered impossible to co-formulate semaglutide and canagliflozin due to the different physicochemical properties of these active pharmaceutical ingredients. The isoelectric point of the GLP-1 receptor agonist semaglutide is incompatible with the optimal pH of the amylin receptor agonist canagliflozin. Semaglutide is optimally stable at pH 7.4 and is therefore prepared beforehand in a neutral to weakly alkaline solution at pH 7–8 to ensure acceptable chemical and physical stability in aqueous solution. Canagliflozin is optimally stable at pH 4.0 and is therefore prepared in an acidic solution, as higher pH levels accelerate its chemical degradation. The different physicochemical properties of canagliflozin and semaglutide prevent the formation of a simple mixture of these two peptides. The same applies to other combinations of GLP-1 receptor agonists and amylin receptor agonists when they have incompatible optimal pH ranges.

[0007] There is still a need in the art for simple means of co-administering GLP-1 receptor agonists such as smegglutide and amylin receptor agonists such as canagliflozin.

[0008] Similarly, there is still a need in the field for such means that are suitable for repeated use. Summary of the Invention

[0009] This document discloses a method for co-formulating an amylin receptor agonist and a GLP-1 receptor agonist. A pharmaceutical formulation is disclosed comprising an amylin receptor agonist, a GLP-1 receptor agonist, a cyclodextrin comprising a hydrophilic chemical substitute such as hydroxypropyl substituted molecule, and at least one preservative and / or stabilizer. The cyclodextrin may be a hydroxypropyl-substituted α-cyclodextrin comprising six cyclically arranged glucose units. The cyclodextrin may be a hydroxypropyl-substituted β-cyclodextrin comprising seven cyclically arranged glucose units. The at least one preservative may be m-cresol and / or phenol and / or EDTA. The at least one stabilizer may be EDTA. The pharmaceutical formulation may further comprise a buffer, such as histidine or citrate; a tensioning agent, such as sorbitol and / or propylene glycol; and / or a surfactant, such as polysorbate 20 and / or 80. The pharmaceutical formulation may be a liquid formulation. The pH of the liquid pharmaceutical formulation is in the range of 5.6-6.4, preferably 5.8-6.2. The pharmaceutical preparations disclosed herein can be administered via parenteral injection, preferably subcutaneous injection. The pharmaceutical preparations disclosed herein are suitable for multiple uses.

[0010] The pharmaceutical formulation of the present invention has several advantages. It allows for the co-formulation of an amylin receptor agonist and a GLP-1 receptor agonist, is suitable for multiple administrations, exhibits antimicrobial growth efficacy, and is well tolerated when administered subcutaneously.

[0011] The pharmaceutical formulations disclosed herein can be used for the medical treatment of subjects suffering from: overweight or obesity with or without one or more related comorbidities, and / or diabetes with or without one or more related comorbidities. The pharmaceutical formulations disclosed herein can improve convenience, treatment adherence, and ultimately clinical outcomes for such patients. Detailed Implementation

[0012] This document discloses a pharmaceutical formulation comprising an amylin receptor agonist, a GLP-1 receptor agonist, a hydroxypropyl-substituted cyclodextrin, and at least one preservative and / or stabilizer. This document also discloses a liquid pharmaceutical formulation comprising an amylin receptor agonist, a GLP-1 receptor agonist, a hydroxypropyl-substituted cyclodextrin, and at least one preservative and / or stabilizer.

[0013] This article discloses methods for co-formulating an amylin receptor agonist and a GLP-1 receptor agonist, wherein the isoelectric point of the GLP-1 receptor agonist prevents co-formulation within a pH range that enables the amylin receptor agonist to possess chemical stability. This article discloses methods for co-forming a GLP-1 receptor agonist with an isoelectric point (pI) less than about 5.0, such as 3.0-5.0, such as 3.8-4.9, such as less than about 4.5, such as 3.0-4.5, such as 3.5-4.5, such as 4.0-4.5, with an amylin receptor agonist.

[0014] The optimal pH for an amylin receptor agonist is the pH at which it is most chemically and physically stable. Those skilled in the art can readily determine the optimal pH of an amylin receptor agonist by testing its chemical and physical stability over a wide pH range in an aqueous solution consisting essentially of the amylin receptor agonist, a buffer, and water for injection.

[0015] The optimal pH of a GLP-1 receptor agonist is the pH at which it is most chemically and physically stable. Those skilled in the art can readily determine the optimal pH of a GLP-1 receptor agonist by testing its chemical and physical stability over a wide pH range in an aqueous solution consisting essentially of the GLP-1 receptor agonist, a buffer, and water for injection. The physical stability of a GLP-1 receptor agonist can reflect its isoelectric point, which may coincide with the pH at which its physical stability is expected to be at its worst.

[0016] Those skilled in the art will understand that the chemical stability and purity of any GLP-1 receptor agonist and / or any amylin receptor agonist in any liquid formulation can be quantified, for example, by reversed-phase (ultra) high-performance liquid chromatography (RP-UHPLC or RP-HPLC) and / or by measuring the percentage (%HMWP) of high molecular weight proteins using, for example, size exclusion chromatography (SEC).

[0017] Those skilled in the art will understand that the physical stability of any GLP-1 receptor agonist and / or any amylin receptor agonist in any liquid formulation can be quantified by measuring particle formation and / or fibrillation using microfluidic imaging (MFI) or thioflavin T (ThT) fluorescence stress assays, respectively.

[0018] This article discloses methods for formulating amylin receptor agonists and GLP-1 receptor agonists with an optimal pH difference of at least about 2 pH units, such as 2-5 pH units, 2-4 pH units, or 3-5 pH units.

[0019] The isoelectric point (pI) of the GLP-1 receptor agonist may be less than 5.0 or less than 4.5, such as 3.0-5.0, 3.5-5.0, 3.5-4.5, 3.8-4.9, or 4.0-4.5. The GLP-1 receptor agonist may be smegglutinin. The GLP-1 receptor agonist may be tirzepatide.

[0020] The isoelectric point (pI) of the amylin receptor agonist may be 7.6-9.4 or 8-9. The amylin receptor agonist may be a human amylin-derived peptide derivative with an isoelectric point (pI) of 7.6-9.4 or 8-9. The amylin receptor agonist may be canagliflozin or its bioactive metabolites or degradation products. The amylin receptor agonist may be a peptide derivative disclosed in WO2013 / 156594, such as the peptide derivative of Example 52.

[0021] The formulations disclosed herein maintain and / or enhance the chemical and physical stability of the active pharmaceutical ingredient, even when co-formulated at pH 5.6–6.4; maintain the pharmacokinetic profile of the active pharmaceutical ingredient in terms of its bioavailability and exposure; and exhibit acceptable local tolerability upon subcutaneous injection.

[0022] The terms “pharmaceutical formulation,” “co-formulation,” and “pharmaceutical product” are used interchangeably in this document and refer to a liquid pharmaceutical formulation containing a GLP-1 receptor agonist and an amylin receptor agonist.

[0023] The drug formulation disclosed in this article is suitable for parenteral injection, preferably subcutaneous injection.

[0024] The pharmaceutical formulation disclosed in this article is suitable for multiple uses.

[0025] amylin The term "amylin" in this article refers to a polypeptide that has the same amino acid sequence as endogenous amylin, such as human amylin.

[0026] amylin receptor Amylin receptor agonists can bind to and activate calcitonin receptor (CTR) and / or amylin receptor (AMYR). The latter consists of a heterodimer of two components: the calcitonin receptor (CTR) and one of three receptor activity-altering proteins (RAMP1–3), resulting in three possible complexes, AMYR1–3.

[0027] amylin receptor agonists The pharmaceutical formulations disclosed herein comprise amylin receptor agonists. An "amylin receptor agonist" can be defined as a chemical entity capable of binding to and activating or "stimulating" an amylin receptor. In the context of this invention, the amylin receptor agonist is capable of binding to and activating at least amylin receptor 3 (AMYR3). The amylin receptor agonist may also be capable of stimulating calcitonin receptor, amylin receptor 1 (AMYR1), and / or amylin receptor 2 (AMYR2).

[0028] Examples of endogenous amylin receptor agonists are human amylin and human calcitonin. Examples of exogenous amylin receptor agonists are canagliflozin and pramlintide (the active pharmaceutical ingredient in Symlin®).

[0029] The concentration of the amylin receptor agonist in the pharmaceutical formulations disclosed herein may be at least about 0.25 mg / ml. The concentration of the amylin receptor agonist in the pharmaceutical formulations disclosed herein may be at most about 22 mg / ml. The concentration of the amylin receptor agonist in the pharmaceutical formulations disclosed herein may be such that any of the doses specified herein is provided.

[0030] Amylin receptor agonists can activate amylin receptors; in other words, they are "effective" against amylin receptors. The in vitro potency of amylin receptor agonists against amylin receptors can be measured as described in assay 2 of WO2022 / 129526. The potency of a compound can be determined by its EC50. 50 The value is used to describe it, where EC 50 This represents the compound concentration at which 50% of its maximum effect was observed. EC 50 The lower the value, the more effective the compound.

[0031] When tested as described in determination 2 of WO2022 / 129526, the EC50 of the amylin receptor agonists disclosed herein... 50 The value can be less than 300 pM, such as less than 200 pM, such as less than 150 pM, preferably less than 100 pM, such as less than 75 pM, preferably less than 50 pM, such as less than 40 pM, such as less than 30 pM, such as less than 20 pM, such as less than 10 pM.

[0032] Canagliflozin The amylin receptor agonist in the pharmaceutical formulation disclosed herein may be canagliflozin or its bioactive metabolites or degradation products.

[0033] Canagliflozin, also known as AM833, is the compound of Example 53 in WO2012 / 168432: N-α-[(S)-4-carboxy-4-(19-carboxy-nonadecanoylamino)butyryl]-[Glu14,Arg17,Pro37]-pralamlinide. Canagliflozin can be prepared as described on pages 153-155 of WO2012 / 168432.

[0034] Canagliflozin may be in the form of a salt, preferably a pharmaceutically acceptable salt.

[0035] The bioactive metabolites or degradation products of canagliflozin may contain aspartic acid (Asp) at position 21 or 22. The bioactive metabolites or degradation products of canagliflozin may contain isoaspartic acid (iso-Asp) at position 21 or 22.

[0036] When the potency of canagliflozin was tested using the procedure described in assay 2 of WO2022129526, the EC50 of canagliflozin was... 50 The value is approximately 11 pM (WO2022 / 129526, Tables 4b and 4c).

[0037] The concentration of canagliflozin in the pharmaceutical formulation disclosed herein can be from about 0.25 mg / ml to about 22 mg / ml.

[0038] The pharmaceutical formulations disclosed herein may contain concentrations of approximately 0.33-18 mg / ml; such as 0.25-0.5 mg / ml, such as approximately 0.33 mg / ml; such as 0.5-1.0 mg / ml, such as approximately 0.67 mg / ml; such as 1.0-1.5 mg / ml, such as approximately 1.33 mg / ml; such as 1.5-2.0 mg / ml, such as approximately 1.5 mg / ml; such as 2.0-2.5 mg / ml; such as 2.5-3.0 mg / ml; such as 3.0-3.5 mg / ml; such as approximately 3.2 mg / ml; such as 3.5-4.0 mg / ml; such as 4.0-5.0 mg / ml; such as 5.0-6.0 mg / ml; such as 6.0-7.0 mg / ml; such as 7.0-8.0 mg / ml; such as 8.0-9.0 mg / ml; such as 9.0-10.0 mg / ml, such as approximately 9.6 mg / ml. mg / ml; such as 10-11 mg / ml, such as 11.0-12.0 mg / ml, such as 11-13 mg / ml; such as 13-22 mg / ml, such as about 18 mg / ml; such as about 20-22 mg / ml canagliflozin.

[0039] The concentration of canagliflozin in the pharmaceutical formulations disclosed herein may be at least about 0.25 mg / ml. The pharmaceutical formulations disclosed herein may contain not more than 22 mg / ml of canagliflozin. The pharmaceutical formulations disclosed herein may contain not more than 12 mg / ml of canagliflozin.

[0040] GLP-1 The term “GLP-1” or “natural GLP-1” in this article refers to human glucagon-like peptide-1 (GLP-1(7-37)).

[0041] GLP-1 receptor agonists The pharmaceutical formulations disclosed herein include GLP-1 receptor agonists. A “GLP-1 receptor agonist” can be defined as a ligand capable of binding to the GLP-1 receptor and producing a biological response similar to that of the endogenous ligand, glucagon-like peptide-1 (GLP-1(7-37)). A “complete” GLP-1 receptor agonist can be defined as a GLP-1 receptor agonist capable of eliciting a biological response of the same magnitude as GLP-1(7-37).

[0042] Examples of known exogenous GLP-1 receptor agonists include exenatide (the active pharmaceutical ingredient in Byetta®), liraglutide (the active pharmaceutical ingredient in Victoza® and Saxenda®, first disclosed in Example 37 of WO98 / 08871), lixisenatide (the active pharmaceutical ingredient in Lyxumia®, disclosed in WO01 / 04156), retatrutide (disclosed in Example 12 of WO2019 / 125938), smegglutide (the active pharmaceutical ingredient in Ozempic®, Rybelsus®, and Wegovy®), and tirzepatide (the active pharmaceutical ingredient in Mounjaro® / Zepbound®, disclosed in Example 1 of WO2016 / 111971 and U.S. Patent No. 9,474,780).

[0043] GLP-1 receptor agonists can be GLP-1 (7-37) peptide derivatives, such as smegglutinin or liraglutinin.

[0044] GLP-1 receptor agonists can be found in compounds that can also activate one or more other receptors; for example, dual or triple receptor agonists, such as telpolide. A GLP-1 receptor agonist can be telpolide.

[0045] The concentration of the GLP-1 receptor agonist in the pharmaceutical formulations disclosed herein can be at least about 0.25 mg / ml. The concentration of the GLP-1 receptor agonist in the pharmaceutical formulations disclosed herein can be at most about 30 mg / ml. The concentration of the GLP-1 receptor agonist in the pharmaceutical formulations disclosed herein can be from about 0.25 mg / ml to about 30 mg / ml. The concentration of the GLP-1 receptor agonist in the pharmaceutical formulations disclosed herein can be from about 0.25 mg / ml to about 22 mg / ml. The concentration of the GLP-1 receptor agonist in the pharmaceutical formulations disclosed herein can be from about 5 mg / ml to about 30 mg / ml. The concentration of the GLP-1 receptor agonist in the pharmaceutical formulations disclosed herein can be such that any of the doses specified herein is provided.

[0046] GLP-1 receptor agonists can bind to and activate or “excite” the GLP-1 receptor; in other words, they are “effective” against the GLP-1 receptor. The in vitro potency of GLP-1 receptor agonists against the GLP-1 receptor can be measured as described in Assay 1 of WO / 2022 / 129526. The potency of a compound can be determined by its EC50. 50 The value is used to describe it, where EC 50 This represents the compound concentration at which 50% of its maximum effect was observed. EC 50 The lower the value, the more effective the compound.

[0047] When tested as described in Determination 1 of WO / 2022 / 129526, the EC50 of the GLP-1 receptor agonists disclosed herein... 50 The value can be less than 300 pM, such as less than 200 pM, such as less than 150 pM, preferably less than 100 pM, such as less than 75 pM, even more preferably less than 50 pM, such as less than 40 pM, such as less than 30 pM, such as less than 20 pM, such as less than 10 pM.

[0048] Smegglutide Smegglutinin is a GLP-1 receptor agonist, also known as N... 6,26 -{18-[N-(17-carboxyheptadecyl)-L-γ-glutamyl]-10-oxo-3,6,12,15-tetraoxa-9,18-diazaoctadecyl}-[8-(2-amino-2-propionic acid),34-L-arginine]human glucagon-like peptide-1(7-37). Smegglutinin is listed in WO2006 / 097537 and J. Med. Chem As described in 2015, 58, 18, 7370–7380, it can be prepared using methods known to those skilled in the art, such as the method briefly described in Example 4 of WO2006 / 097537.

[0049] Smegglutide may be present in this pharmaceutical preparation in its fully or partially ionized form; for example, one or more carboxylic acid groups (-COOH) may be deprotonated to carboxylate groups (-COO). - And / or one or more amino groups (-NH2) can be protonated to -NH3. + Group.

[0050] Smegglutide can be in the form of a salt, preferably a pharmaceutically acceptable salt.

[0051] When the potency of semaglutide was tested according to the procedure described in Determination 1 of WO / 2022 / 129526, the EC50 of semaglutide was... 50 The value is approximately 5.5 pM (see WO / 2022 / 129526, Tables 4b and 4c).

[0052] The concentration of smegglutide in the pharmaceutical formulation disclosed herein can be from about 0.25 mg / ml to about 22 mg / ml.

[0053] The pharmaceutical preparation may contain concentrations of approximately 0.33-18 mg / ml; such as 0.25-0.5 mg / ml, such as approximately 0.33 mg / ml; such as 0.5-1.0 mg / ml, such as approximately 0.67 mg / ml; such as 1.0-1.5 mg / ml, such as approximately 1.33 mg / ml; such as 1.5-2.0 mg / ml, such as approximately 1.5 mg / ml; such as 2.0-2.5 mg / ml, such as approximately 2.2 mg / ml; such as 2.5-3.0 mg / ml; such as 3.0-3.5 mg / ml, such as approximately 3.2 mg / ml; such as 3.5-4.0 mg / ml; such as 4.0-5.0 mg / ml, such as approximately 4.8 mg / ml; such as 5.0-6.0 mg / ml; such as 6.0-7.0 mg / ml, such as approximately 6.4 mg / ml; such as 7.0-8.0 mg / ml, such as approximately 8.0 mg / ml. mg / ml; such as 8.0-9.0 mg / ml, such as 9.0-10.0 mg / ml, such as about 9.6 mg / ml; such as 10-11 mg / ml, such as about 10.7 mg / ml; such as 11.0-12.0 mg / ml, such as 11-13 mg / ml; such as about 12.8 mg / ml; such as 13-22 mg / ml, such as about 16 mg / ml; such as about 18 mg / ml; such as about 20-22 mg / ml of smegglutide.

[0054] The concentration of semaglutide in the pharmaceutical formulations disclosed herein may be at least about 0.25 mg / ml. The pharmaceutical formulations disclosed herein may contain not more than 22 mg / ml of semaglutide. The pharmaceutical formulations disclosed herein may contain not more than 12 mg / ml of semaglutide.

[0055] Preparation method For example, GLP-1 receptor agonists and / or amylin receptor agonists in the pharmaceutical formulations disclosed herein can be produced by classical peptide synthesis, such as solid-phase peptide synthesis using t-Boc or Fmoc chemistry, or by other established techniques, such as those described in Greene and Wuts, “Protective Groups in Organic Synthesis”, JohnWiley & Sons, 1999; Florencio Zaragoza Dörwald, “Organic Synthesis on Solid Phase”, Wiley-VCH Verlag GmbH, 2000; and “Fmoc Solid Phase Peptide Synthesis”, edited by WC Chan and PD White, Oxford University Press, 2000.

[0056] Alternatively, GLP-1 receptor agonists and / or amylin receptor agonists can be produced via recombinant expression techniques, for example, by culturing host cells containing DNA sequences encoding peptide sequences and capable of expressing those peptides in suitable nutrient media under conditions that allow for peptide expression. Non-limiting examples of host cells suitable for expressing these peptides are *Escherichia coli* (E. coli). Escherichia coli ), brewer's yeast ( Saccharomyces cerevisiae ) and mammalian BHK or CHO cell lines.

[0057] Alternatively, a combination of recombinant expression techniques and chemical peptide synthesis, as described in WO2009 / 083549, can be used to semi-synthetically produce GLP-1 receptor agonists containing one or more non-protein amino acids. Compounds containing one or more non-natural amino acids and / or covalently linked N-terminal monopeptides or dipeptide mimics can also be produced, as described by Hodgson et al. in "The synthesis of peptides and proteins containing non-natural amino acids". Chemical Society Reviews It is produced as described in , vol. 33, no. 7 (2004), pp. 422-430.

[0058] Once the GLP-1 receptor agonist and the amylin receptor agonist (active pharmaceutical ingredient) have been prepared and purified, the pharmaceutical formulation (pharmaceutical product) disclosed herein can be prepared using the method described in WO2023 / 187067.

[0059] Alternatively, or subsequently, the composition comprising the active pharmaceutical ingredient and cyclodextrin, such as a pharmaceutical product, may be freeze-dried or spray-dried using methods known to those skilled in the art. Ohtake, S., Izutsu, KI, and Lechuga-Ballesteros, D. (Editors) "Drying technologies for biotechnology and pharmaceutical applications Such a method is described in John Wiley & Sons (2020). The dried composition containing the active pharmaceutical ingredient and cyclodextrin may further contain surfactants such as polysorbate 20 and / or 80.

[0060] If the composition has undergone intermediate freeze-drying or spray drying, the dry formulation is dissolved or “reconstituted” in an aqueous solution before use. The aqueous solution may be contained in a vial. The aqueous solution may contain a predetermined amount of water for injection or consist thereof. The aqueous solution may contain a predetermined amount of water for injection and one or more preservatives or consist thereof. The aqueous solution may contain a predetermined amount of water for injection and phenol and / or m-cresol and / or EDTA or consist thereof. The aqueous solution may contain a predetermined amount of water for injection, phenol and / or EDTA or consist thereof. The aqueous solution may contain a predetermined amount of water for injection and m-cresol and / or consist thereof. The aqueous solution may contain a predetermined amount of water for injection and phenol or consist thereof. The aqueous solution may contain a predetermined amount of water for injection and m-cresol or consist thereof. The aqueous solution may contain a predetermined amount of water for injection and EDTA or consist thereof. The aqueous solution may further contain a buffer having at least one pKa value of about 5.0-7.0, such as histidine or citrate. The aqueous solution may further contain sorbitol.

[0061] Regardless of whether the formulation undergoes intermediate freeze-drying or spray-drying, the liquid pharmaceutical formulation ultimately administered to the patient has the composition disclosed herein.

[0062] isoelectric point The isoelectric point (pI) of a molecule is the pH at which the molecule carries no net charge. The pI of a peptide, such as the peptide backbone of a GLP-1 receptor agonist or an amylin receptor agonist, can be theoretically calculated based on the pK values ​​of the peptide's amino acids and the terminal amine and carboxyl groups, and can be used to predict the peptide's solubility at a given pH.

[0063] The total net charge of a protein or peptide is related to the solution pH and can be approximated using the Henderson-Hasselbach equation (Po HN, Senozan NM. The Henderson-Hasselbach Equation: Its History and Limitations. J Chem Educ. 2001;78(11):1499). To derive the isoelectric point of a peptide or find its charge at a specific pH, the pKa values ​​of its amino acids must be considered. At pH 7.0, the carboxyl group is negative (-1), while the amino group is positive (+1). The net charge of a peptide at a given pH is the sum of the charges of the ionizable groups in the peptide at that given pH. The total charge of the peptide can be determined by summing the charges of all ionizable groups.

[0064] Several software tools can predict the charge state of peptides and help understand their behavior under different pH conditions [IPC - Isoelectric Point Calculator, Kozlowski, Lukasz P., Biology Direct (2016), 11, 55 / 1-55 / 16]. Because they employ algorithms that consider the pKa values ​​of amino acids, the predictions are considered to be free of significant deviations [Principles of Biochemistry, AL Lehninger and MM Cox, 1982].

[0065] When peptides are chemically modified, these modifications can affect the pI; however, these modifications can also be accounted for in pI predictions using the Henderson-Hasselback equation by evaluating the pKa of the ionizable groups in relation to the modifications. One example of this type of "chemical modification" is an elongated portion, such as a side chain containing a fatty acid that is covalently bound to the peptide.

[0066] The charge of a peptide affects its solubility. Peptides with a net charge, whether positive or negative, exhibit enhanced solubility in water because the charged groups can interact with water molecules [Pace CN, Grimsley GR, Scholtz JM. Protein ionizable groups: pK values ​​and their contribution to protein stability and solubility. J Biol. Chem. 2009;284(20):13285–13289], [The effect of net charge on the solubility, activity, and stability of ribonuclease Sa, Shaw, Kevin L.; Grimsley, Gerald R.; Yakovlev, Gennady I.; Makarov, Alexander A.; Pace, C. Nick, Protein Science (2001), 10 (6), 1206-1215]. Therefore, good solubility is generally obtained at formulation pH far from the isoelectric point, where the peptide carries a charge (positive or negative) sufficient to provide solubility.

[0067] In the context of this invention, the theoretically calculated or predicted isoelectric point of a GLP-1 receptor agonist may be equal to or less than about 5.0, preferably less than about 4.5. The theoretically calculated / predicted isoelectric point of a GLP-1 receptor agonist may be in the range of 3.5-5.0, such as 3.8-4.9, such as 3.5-4.5, for example, about 4.0-4.5. For example, the theoretically calculated / predicted isoelectric point of semaglutide is about 4.37. The theoretically calculated / predicted isoelectric point of liraglutide is about 4.47. The theoretically calculated / predicted isoelectric point of telposide is about 4.03. The theoretically calculated / predicted isoelectric point of retaglutide is about 3.93.

[0068] There are other GLP-1 receptor agonists with theoretically calculated isoelectric points higher than 4.5.

[0069] Theoretically calculated isoelectric points (pI) of amylin receptor agonists can be in the range of 7.6–9.4 or 8–9. Theoretically calculated isoelectric point of canagliflozin is approximately 8.56.

[0070] There are other amylin receptor agonists with theoretically calculated isoelectric points higher than 9.4.

[0071] auxiliary materials Cyclodextrin The pharmaceutical formulation disclosed herein contains cyclodextrin with hydroxypropyl substitution.

[0072] The pharmaceutical formulation may contain more than 10% w / v of hydroxypropyl-substituted cyclodextrin. The pharmaceutical formulation may contain less than 22% w / v of hydroxypropyl-substituted cyclodextrin. The pharmaceutical formulation may contain approximately 10-20% w / v, approximately 12-18% w / v, approximately 10-17.5% w / v, approximately 11.25-15%, for example, approximately 15% w / v of hydroxypropyl-substituted cyclodextrin.

[0073] Cyclodextrins are oligosaccharide starch derivatives composed of 6, 7, or 8 α-(1,4)-linked pyranose (glucose) units arranged in a cyclic pattern, representing α, β, or γ types, respectively. Cyclodextrins have a wide range of applications, especially as pharmaceutical excipients [P. Breen & S. S. Jambhekar, Cyclodextrins in pharmaceutical formulations II: solubilization, binding constant, and complexation efficiency, Drug Discovery Today, Vol. 21, No. 2, February 2016]. The European Medicines Agency has described guidance on their use as pharmaceutical excipients [Background review for cyclodextrins used as excipients, 2014, EMA / CHMP / 333892 / 2013, Committee for Human Medicinal Products (CHMP)], [Cyclodextrins used as excipients, 2017, EMA / CHMP / 333892 / 2013, Committee for Human Medicinal Products (CHMP)]. Cyclodextrins without hydrophilic substitutions have poor solubility and are rarely used in parenteral pharmaceutical products.

[0074] To improve the solubility of cyclodextrins, the hydroxyl groups of the glucose units can be replaced by varying numbers of hydrophilic chemical substituents, such as hydroxypropyl groups, resulting in differences in the degree of substitution. The degree of substitution can be described as the average number of hydroxypropyl groups per cyclodextrin molecule (abbreviated as DS) or the molar degree of substitution (abbreviated as MS) corresponding to the average number of hydroxypropyl groups per glucose unit present in the cyclodextrin. The hydroxypropyl value of each cyclodextrin can be obtained by multiplying the molar degree of substitution by the number of glucose units contained in the cyclodextrin. Differences in the degree of substitution can lead to changes in physicochemical properties such as surface activity and complexing ability. The hydroxyl groups can also be chemically substituted with sulfobutyl ether groups. These are mostly hydrophilic modifications that produce cyclodextrin derivatives that are well-suited for parenteral administration [Cyclodextrins used as excipients, 2017, EMA / CHMP / 333892 / 2013, Committee for Human Medicinal Products (CHMP)]. Cyclodextrins containing hydroxypropyl substitution are usually abbreviated as HP-CD, while cyclodextrins containing sulfobutyl ether substitution are abbreviated as SBE-CD.

[0075] Cyclodextrins containing hydrophilic substitutions take on a shape that can be described as conical, with a hydrophobic interior and a hydrophilic outer surface formed by numerous hydrophilic substituents capable of forming hydrogen bonds with neighboring water molecules, thereby improving water solubility [T. Loftsson, Cyclodextrins in Parenteral Formulations, Journal of Pharmaceutical Sciences, 2020, 1-11].

[0076] The hydrophobic microenvironment within the cavities of these conical structures enables them to form drug-cyclodextrin complexes primarily through hydrophobic interactions [T. Loftsson, Cyclodextrins in Parenteral Formulations, Journal of Pharmaceutical Sciences, 2020, 1-11]. When cyclodextrins form complexes with drug molecules carrying one or more hydrophobic regions, these hydrophobic regions, along with the hydrophobic cavities of the cyclodextrin, are shielded from water, thereby increasing the solubility of the complex compared to the solubility of the individual components. Furthermore, once cyclodextrins form complexes with peptide molecules, they weaken intermolecular interactions that would normally lead to aggregation [T. Loftsson, Cyclodextrins in Parenteral Formulations, Journal of Pharmaceutical Sciences, 2020, 1-11].

[0077] The pharmaceutical formulations disclosed herein preferably comprise hydroxypropyl-substituted α-cyclodextrin and / or hydroxypropyl-substituted β-cyclodextrin.

[0078] Surprisingly, this type of cyclodextrin with hydroxypropyl substitution was found to be superior to the same type of cyclodextrin with sulfobutyl ether substitution in its ability to stabilize co-formulations of canagliflozin and smegglutinin.

[0079] The pharmaceutical formulations disclosed herein may comprise hydroxypropyl-substituted α-cyclodextrins, which contain six cyclically arranged glucose units. The abbreviation for hydroxypropyl-substituted α-cyclodextrin is HP-A-CD. Hydroxypropyl-α-cyclodextrin (CAS: 128446-33-3 / 99241-24-4) is commercially available with an average molar degree of substitution (MS) of 0.8 and a range of 0.5–0.9.

[0080] The pharmaceutical formulations disclosed herein may comprise hydroxypropyl-α-cyclodextrin having at least about 0.4 hydroxypropyl groups per glucose unit. The pharmaceutical formulations disclosed herein may comprise hydroxypropyl-α-cyclodextrin having at most about 1.2 hydroxypropyl groups per glucose unit.

[0081] The pharmaceutical formulations disclosed herein may comprise hydroxypropyl-α-cyclodextrin with a molar substitution degree ranging from 0.5 to 0.9 hydroxypropyl groups per glucose unit. The pharmaceutical formulations disclosed herein may comprise hydroxypropyl-α-cyclodextrin with an average molar substitution degree (MS) of approximately 0.8 hydroxypropyl groups per glucose unit.

[0082] The pharmaceutical formulations disclosed herein may contain hydroxypropyl-substituted β-cyclodextrins, which contain seven cyclically arranged glucose units.

[0083] Hydroxypropyl-substituted β-cyclodextrin is abbreviated as HP-B-CD.

[0084] Hydroxypropyl-β-cyclodextrin is a well-known pharmaceutical excipient, commonly used in small molecule drug formulations, primarily to increase solubility and bioavailability [T. Loftsson, Cyclodextrins in Parenteral Formulations, Journal of Pharmaceutical Sciences, 2020, 1-11]. To date, the use of cyclodextrins and substituted cyclodextrin derivatives in protein and peptide drug formulations has been limited.

[0085] According to the European and American Pharmacopoes [USP 38 NF 33, Pharm Eur 8, estimated by the method described in USP<761> / Pharm. Eur.2.2.33], the degree of hydroxypropyl substitution (DS) of commercially available hydroxypropyl-β-cyclodextrins as pharmaceutical excipients ranges from 2.8 to 10.5, corresponding to 0.4–1.5 hydroxypropyl groups (MS) per glucose unit. Commercially available cyclodextrins, such as hydroxypropyl-β-cyclodextrin, are typically described by the average molar substitution (MS) within their molar substitution range.

[0086] Hydroxypropyl-β-cyclodextrin (CAS: 128446-35-5 / 94035-02-6) is commercially available for use as an excipient, with the following average molar degree of substitution (MS): MS 0.62, ranging from 0.58 to 0.68; MS 0.67, ranging from 0.6 to 0.9; MS 0.68, ranging from 0.58 to 0.72; MS 0.84, ranging from 0.8 to 1.0; MS 0.92, ranging from 0.81 to 0.99; MS 1.08, ranging from 0.86 to 1.14; each value describes the number of hydroxypropyl groups per glucose unit.

[0087] The pharmaceutical formulations disclosed herein may comprise hydroxypropyl-β-cyclodextrin having at least about 0.4 hydroxypropyl groups per glucose unit. The pharmaceutical formulations disclosed herein may comprise hydroxypropyl-β-cyclodextrin having at most about 1.2 hydroxypropyl groups per glucose unit.

[0088] The pharmaceutical formulations disclosed herein may comprise hydroxypropyl-β-cyclodextrin with a molar degree of substitution ranging from 0.58 to 1.0 hydroxypropyl groups per glucose unit. The pharmaceutical formulations disclosed herein may comprise hydroxypropyl-β-cyclodextrin with an average molar degree of substitution (MS) ranging from about 0.62 to 0.92 hydroxypropyl groups per glucose unit.

[0089] The pharmaceutical formulations disclosed herein may contain hydroxypropyl-β-cyclodextrin with an average molar degree of substitution (MS) of about 0.62-0.84 hydroxypropyl groups per glucose unit.

[0090] The pharmaceutical formulations disclosed herein may comprise hydroxypropyl-β-cyclodextrin with an average molar degree of substitution (MS) of approximately 0.62. The pharmaceutical formulations disclosed herein may comprise hydroxypropyl-β-cyclodextrin having approximately 0.58-0.68 hydroxypropyl groups per glucose unit.

[0091] The pharmaceutical formulations disclosed herein may comprise hydroxypropyl-β-cyclodextrin with an average molar degree of substitution (MS) of approximately 0.68. The pharmaceutical formulations disclosed herein may comprise hydroxypropyl-β-cyclodextrin having approximately 0.58-0.72 hydroxypropyl groups per glucose unit.

[0092] The pharmaceutical formulations disclosed herein may comprise hydroxypropyl-β-cyclodextrin with an average molar degree of substitution (MS) of approximately 0.67. The pharmaceutical formulations disclosed herein may comprise hydroxypropyl-β-cyclodextrin having approximately 0.6-0.9 hydroxypropyl groups per glucose unit.

[0093] The pharmaceutical formulations disclosed herein may comprise hydroxypropyl-β-cyclodextrin with an average molar degree of substitution (MS) of approximately 0.84. The pharmaceutical formulations disclosed herein may comprise hydroxypropyl-β-cyclodextrin having approximately 0.8-1.0 hydroxypropyl groups per glucose unit.

[0094] The pharmaceutical formulations disclosed herein may comprise hydroxypropyl-β-cyclodextrin with an average molar degree of substitution (MS) of approximately 0.92. The pharmaceutical formulations disclosed herein may comprise hydroxypropyl-β-cyclodextrin having approximately 0.81-0.99 hydroxypropyl groups per glucose unit.

[0095] The pharmaceutical formulations disclosed herein may contain more than 10% w / v and less than 22% w / v, such as about 10-20% w / v, such as about 12-18% w / v, such as about 10-17.5% w / v, such as about 11.25-15%, such as about 15% w / v of hydroxypropyl-β-cyclodextrin having at least about 0.4 hydroxypropyl groups per glucose unit and at most about 1.2 hydroxypropyl groups per glucose unit; such as an average of 0.62-0.92 hydroxypropyl groups per glucose unit, such as about 0.75 hydroxypropyl groups per glucose unit; such as an average of 0.62-0.84 hydroxypropyl groups per glucose unit; such as an average of 0.62 hydroxypropyl groups per glucose unit, such as about 0.58-0.68 hydroxypropyl groups per glucose unit.

[0096] Cyclodextrins bind to the hydrophobic portion of the molecule and form a complex.

[0097] The interactions between several common pharmaceutical preservatives and cyclodextrins are well documented and constitute a major obstacle in the development of preservative-containing liquid pharmaceutical formulations (Loftsson, T., Stefansdottir, O., Friôriksdóttir, H., & Guômundsson, Ö. (1992)). Drug development and industrial pharmacy (18(13), 1477-1484). First, the preservative loses its antimicrobial activity when the preservative-cyclodextrin complex is formed. Second, the preservative competes with and displaces the drug in the drug-cyclodextrin complex, thereby significantly reducing the chemical and physical stability of the formulation. The pharmaceutical formulations disclosed herein address these well-known obstacles.

[0098] Other auxiliary materials The pharmaceutical formulations disclosed herein may contain one or more preservatives. Preservatives used in pharmaceutical formulations are well known to those skilled in the art. For convenience, see Remington: The Science and Practice of Pharmacy , 21st edition, 2006. The concentration of the one or more preservatives must be such that regulatory requirements regarding the antimicrobial efficacy of reusable pharmaceutical preparations are met. In other words, the purpose of the one or more preservatives is to inhibit microbial growth once the pharmaceutical preparation is no longer in sterile packaging. The most widely used pharmaceutical antimicrobial preservatives are aromatic organic compounds such as benzyl alcohol, phenol, m-cresol, and parabens.

[0099] One or more preservatives in the pharmaceutical formulations disclosed herein may be m-cresol and / or phenol and / or EDTA.

[0100] One of the preservatives in the pharmaceutical formulations disclosed herein may be m-cresol at a concentration of 9-40 mM.

[0101] One of the preservatives in the pharmaceutical preparations disclosed herein may be phenol at a concentration of 18-65 mM.

[0102] The preservatives disclosed in this article may be m-cresol at a concentration of 9-40 mM and phenol at a concentration of 18-65 mM.

[0103] One of the preservatives in the pharmaceutical formulations disclosed herein may be EDTA at a concentration of 0.5-5.0 mg / ml.

[0104] The preservatives disclosed in this article may be m-cresol at a concentration of 9-40 mM, phenol at a concentration of 18-65 mM, and EDTA at a concentration of 0.5-5.0 mg / ml.

[0105] The pharmaceutical formulation may contain stabilizers. The use of stabilizers in pharmaceutical formulations is well known to those skilled in the art. For convenience, see Remington: The Science and Practice of Pharmacy , 21st edition, 2006. The stabilizer can be EDTA. The stabilizer can be EDTA at a concentration of 0.5-5.0 mg / ml.

[0106] Therefore, in the context of the formulations disclosed herein, EDTA can act as a preservative and / or stabilizer.

[0107] The pharmaceutical formulation may contain a buffer. The use of buffers in pharmaceutical formulations is well known to those skilled in the art. For convenience, see Remington: The Science and Practice of Pharmacy , 21st edition, 2006.

[0108] pH can be measured at “room temperature”, which is typically defined as 15-25°C or 15-20°C. pH is preferably measured at about 20°C.

[0109] The pharmaceutical formulation disclosed herein may contain a buffer with a pKa close to the desired pH of the solution. The pharmaceutical formulation may contain a buffer having at least one pKa value of approximately 5.0-7.0. The pharmaceutical formulation may contain a buffer with a pKa of approximately 5.0-7.0. The pharmaceutical formulation may contain a buffer selected from histidine, citrate, and / or phosphate. The buffer may be citrate at a concentration of 3-30 mM. The buffer may be histidine at a concentration of 3-30 mM. The buffer may be phosphate at a concentration of 3-30 mM.

[0110] The pharmaceutical preparation may further contain one or more reagents for adjusting pH, such as NaOH and / or HCl.

[0111] The desired pH of the pharmaceutical preparation may be about 5.6-6.4, for example about 5.8-6.2. The pH may be about 5.6, for example about 5.7, for example about pH 5.8, for example about 5.9, for example about 6.0, for example about 6.1, for example about 6.2, for example about 6.3, for example about 6.4. The pH is preferably about 6.0.

[0112] The pharmaceutical formulation may contain a tensile agent. The use of tensile agents in pharmaceutical formulations is well known to those skilled in the art. For convenience, see Remington: The Science and Practice of Pharmacy , 21st edition, 2006.

[0113] The purpose of a turbinate is to protect living tissue when the formulation is injected into the body. A turbinate can be selected from glycerol, mannitol, propylene glycol, sorbitol, or trehalose, or a combination thereof. A turbinate can be glycerol. A turbinate can be mannitol. A turbinate can be propylene glycol. A turbinate can be sorbitol. A turbinate can be trehalose.

[0114] The concentration of the osmotic agent makes the preparation isotonic. When the osmotic agent is glycerol, it can be present at a concentration of 2.5-18 mg / ml. When the osmotic agent is mannitol, it can be present at a concentration of 16.5-37.5 mg / ml, for example, about 20 mg / ml. When the osmotic agent is propylene glycol, it can be present at a concentration of about 2-15 mg / ml. When the osmotic agent is sorbitol, it can be present at a concentration of about 5-35 mg / ml; for example, about 10-30 mg / ml; for example, about 16-28 mg / ml; for example, about 16.5-25 mg / ml; for example, about 16-26 mg / ml; for example, about 16-24 mg / ml; for example, about 26 mg / ml; for example, about 24 mg / ml; for example, about 22 mg / ml; for example, about 20 mg / ml; for example, about 18 mg / ml; for example, about 16 mg / ml; for example, about 12 mg / ml. When the tonic is trehalose, it can be present at a concentration of 33-75 mg / ml, for example, about 38 mg / ml.

[0115] The pharmaceutical formulation may contain surfactants. Surfactants can further increase the physical stability and robustness of the formulation during its preparation, storage, and use as a medicine; for example, by maintaining its stability when the formulation is exposed to air within the container. The use of surfactants in pharmaceutical formulations is well known to those skilled in the art. For convenience, see Remington: The Science and Practice of Pharmacy , 21st edition, 2006.

[0116] The surfactant may be selected from polysorbate 20 and / or polysorbate 80. The surfactant may be polysorbate 20. The surfactant is preferably polysorbate 80.

[0117] The pharmaceutical formulation may contain 0.01 mg / ml or more of polysorbate 20, and up to 2.0 mg / ml, such as up to 1.5 mg / ml of polysorbate 20.

[0118] The pharmaceutical formulation may contain about 0.01-1.0 mg / ml of polysorbate 20, for example, about 0.05 mg / ml of polysorbate 20.

[0119] When contained in vials, the pharmaceutical formulation may contain more than about 0.1 mg / ml and less than about 0.2 mg / ml of polysorbate 20.

[0120] The pharmaceutical formulation may contain 0.01 mg / ml or more of polysorbate 80, and up to 2.0 mg / ml, such as up to 1.5 mg / ml of polysorbate 80.

[0121] The pharmaceutical formulation may contain 0.01-0.1 mg / ml of polysorbate 80; preferably about 0.05 mg / ml of polysorbate 80.

[0122] When contained in vials, the pharmaceutical formulation may contain more than about 0.1 mg / ml and less than about 0.2 mg / ml of polysorbate 80.

[0123] The pharmaceutical formulation may contain water for injection (WFI). The pharmaceutical formulation may contain more than 75% w / w water, such as 80% w / w water, for example about 85% w / w water, or up to 90% w / w water.

[0124] Medical Applications The pharmaceutical preparations disclosed in this article can be used for medical purposes.

[0125] The drug formulations disclosed herein can be administered via parenteral injection. The drug formulations disclosed herein can also be administered via subcutaneous injection.

[0126] As used herein, the term "treatment" refers to medical treatment of any human or other vertebrate subject in need. The subject is expected to have undergone a physical examination by a licensed veterinarian who has provided a preliminary or definitive diagnosis indicating that the use of the specific treatment will be beneficial to the health of the human or other vertebrate. The timing and purpose of the treatment may vary from individual to individual, depending on the subject's current health condition. Therefore, the treatment may be preventative, palliative, symptomatic, and / or curative.

[0127] The drug formulations disclosed in this article can be administered to human subjects.

[0128] The pharmaceutical formulations disclosed in this article can be used for: (i) Prevention and / or treatment of any form of diabetes and related symptoms, such as hyperglycemia, type 2 diabetes, impaired glucose tolerance, type 1 diabetes, non-insulin-dependent diabetes, juvenile mature-onset diabetes (MODY), gestational diabetes, and / or for reducing HbA1c; (ii) To delay or prevent the progression of diabetes, such as the progression of type 2 diabetes, to delay the progression of impaired glucose tolerance (IGT) to type 2 diabetes requiring insulin, and / or to delay the onset and / or progression of type 2 diabetes that does not require insulin. (iii) Prevention and / or treatment of certain eating disorders, overweight and / or obesity; for example, by reducing food intake, suppressing appetite, inducing satiety, and reducing weight; treatment or prevention of binge eating disorder, eating impulses, bulimia nervosa and / or obesity induced by medications such as antipsychotics or steroids; reduction of gastric motility; and / or delay of gastric emptying; (iv) Prevention and / or treatment of cardiovascular diseases, such as delaying or reducing the development of major adverse cardiovascular events (MACE) selected from cardiovascular death, nonfatal myocardial infarction, nonfatal stroke, revascularization, hospitalization due to unstable angina, and hospitalization due to heart failure; (v) Prevention and / or treatment of non-alcoholic fatty liver disease (NAFLD) and / or non-alcoholic steatohepatitis (NASH), also known as metabolic dysfunction-associated steatohepatitis (MASH), and / or alcoholic liver disease (ALD); (vi) Prevention and / or treatment of chronic kidney disease; (vii) Prevention and / or treatment of obstructive sleep apnea; (viii) Prevention and / or treatment of cognitive impairments, such as Alzheimer's disease.

[0129] The indication may be (i). The indication may be (ii). The indication may be (iii). The indication may be (iv). The indication may be (v). The indication may be (vi). The indication may be (vii). The indication may be (viii). The indication may be overweight or obesity. The indication may be type 2 diabetes.

[0130] Typically, all participants with obesity are also considered overweight. Participants with obesity can be humans, such as adults or children, where "children" includes infants and adolescents.

[0131] The World Health Organization (WHO) defines obesity as the abnormal or excessive accumulation of fat that may impair health, and considers body mass index (BMI) to be the most convenient population-level measure of overweight and obesity. The formula for calculating BMI is weight in kilograms (kg) divided by weight in square meters (m²). 2 ( ) is the square of height in units of .

[0132] For adults, the WHO defines overweight and obesity as follows: overweight is defined as a BMI greater than or equal to 25; obesity is defined as a BMI greater than or equal to 30.

[0133] For children, the WHO considers age when defining overweight and obesity.

[0134] For children under five years old, overweight is defined as weight per height exceeding the WHO median growth rate for children by more than two standard deviations; while obesity is defined as weight per height exceeding the WHO median growth rate for children by more than three standard deviations.

[0135] For children aged 5 to 19, overweight and obesity are defined as follows: overweight is defined as an age-related BMI that is more than one standard deviation above the WHO growth reference median; while obesity is defined as an age-related BMI that is more than two standard deviations above the WHO growth reference median.

[0136] However, the diagnostic criteria for underweight, normal range, pre-obesity / overweight and obesity may vary between different countries / populations, as shown in Table (i) below for adults.

[0137] Table (i): Definitions of underweight, normal range, pre-obesity / overweight, and obesity in adults

[0138] Misra A et al. published guidelines for Asian populations in J Assoc Physicians India. 2009; 57:163-70.

[0139] The 2006 edition of the "Guidelines for the Prevention and Control of Overweight and Obesity in Chinese Adults," compiled by the China Obesity Working Committee, included guidelines for the Chinese population.

[0140] In 2016, the Japan Association for the Study of Obesity (JASSO) held a meeting... Guidelines for the management of obesity disease The guidelines for the Japanese population were published in the (Obesity Management Guidelines).

[0141] Therefore, adult subjects suffering from obesity may have a weight of 25 kg / m². 2 Or higher, 27 kg / m 2 Or higher, 28 kg / m 2 Or higher or 30 kg / m 2 Or a higher BMI; the subject may also be classified as obese. Obesity can be classified as Grade I, II, III, or IV obesity. Overweight adult subjects may have a BMI of 24 kg / m². 2 Or higher, 25 kg / m 2 Or higher or 27 kg / m 2 Or a higher BMI. In some implementations, the BMI of overweight human subjects is between 24 and <27 kg / m². 2Within the range of 24 to <28 kg / m 2 Within the range of 25 to <30 kg / m 2 Within the range, or between 27 and <30 kg / m 2 Within the range.

[0142] A BMI higher than normal increases an individual's risk of developing any of a number of other diseases or comorbidities. Weight-related comorbidities can be any one of the diseases mentioned in (i), (ii), (iv), (v), and (vii) above, or a combination thereof.

[0143] The pharmaceutical formulations disclosed herein can be used to treat or prevent overweight, where the patient may have at least one weight-related comorbidity. The pharmaceutical formulations disclosed herein can be used to treat or prevent obesity, where the patient may have at least one weight-related comorbidity.

[0144] The pharmaceutical formulation disclosed in this article can be used for long-term weight management in obese subjects at the start of treatment as an adjunct to a low-calorie diet and increased physical activity.

[0145] The pharmaceutical formulation disclosed herein can be used for long-term weight management in subjects who are overweight at the start of treatment and have at least one weight-related comorbidity, as an adjunct to a low-calorie diet and increased physical activity.

[0146] The pharmaceutical formulation disclosed in this article can be used at an initial body mass index (BMI) of 30 kg / m². 2 Or higher, 28 kg / m 2 Or higher, 27 kg / m 2 Or higher or 25 kg / m 2 In adults or older, it is used as an adjunct to a low-calorie diet and increased physical activity for long-term weight management.

[0147] The pharmaceutical formulation disclosed in this article can be used at an initial body mass index (BMI) of 25 kg / m². 2 Or higher or 24 kg / m 2 In adults of advanced age and with at least one weight-related comorbidity, it is used as an adjunct to a low-calorie diet and increased physical activity for long-term weight management.

[0148] Administration of the drug formulation disclosed herein can result in >15% weight loss, such as >20% weight loss, such as >25% weight loss, such as >30% weight loss, such as approximately 15-40% weight loss, such as approximately 20-35% weight loss, such as approximately 25-30% weight loss within 26 weeks of the start of treatment.

[0149] Administration of the drug formulation disclosed herein can result in >15% weight loss, such as >20% weight loss, such as >25% weight loss, such as >30% weight loss, such as approximately 15-40% weight loss, such as approximately 20-35% weight loss, such as approximately 25-30% weight loss within 26 weeks of the start of treatment.

[0150] The pharmaceutical preparations disclosed herein may be used to treat or prevent diabetes, as described in (i) or (ii) above. The pharmaceutical preparations disclosed herein may be used to treat or prevent diabetes and at least one comorbidity associated with diabetes, as described in (vi) above.

[0151] Compared to treatments using GLP-1 receptor agonists or amylin receptor agonists as the sole active ingredient, administration of the pharmaceutical formulation disclosed herein results in a higher percentage of HbA1c. 1c reduce.

[0152] Compared to treatments using semaglutide or canagliflozin as the sole active ingredient, administration of the semaglutide and canagliflozin formulations disclosed herein results in higher HbA1c levels on a percentage basis. 1c reduce.

[0153] dose The pharmaceutical formulation of the present invention comprises a specific concentration of an amylin receptor agonist and a specific concentration of a GLP-1 receptor agonist. For example, as described above, the pharmaceutical formulation may comprise 0.25 to 22 mg / ml of canagliflozin and 0.25 to 22 mg / ml of smegglutinin. The dosage of the GLP receptor agonist and amylin receptor agonist administered in a single injection depends on the concentration of these active ingredients in the pharmaceutical formulation and the volume of the pharmaceutical formulation administered.

[0154] The pharmaceutical formulation of the present invention can be administered in single doses at predetermined time intervals.

[0155] The pharmaceutical formulations disclosed herein may contain, in a single dose, any of the following doses: amylin receptor agonists such as canagliflozin and GLP-1 receptor agonists such as smegglutinin.

[0156] Effective doses of amylin receptor agonists such as canagliflozin and GLP-1 receptor agonists such as smegglutinin can be administered to subjects who require them.

[0157] The dose can be administered approximately once a week. The interval between two fixed doses can be approximately 4 days, 5 days, 6 days, 7 days, 8 days, 9 days, or 10 days. In a preferred embodiment, a fixed maintenance dose is administered approximately every 7 days (once a week).

[0158] The dose may be administered to an individual suffering from any of the diseases or comorbidities listed above, or a combination thereof. In some preferred embodiments, the dose may be administered to an individual suffering from obesity (body mass index [BMI] ≥ 30 kg / m²). 2 Subjects who are overweight (BMI ≥ 27 kg / m²). In some preferred embodiments, the dose is administered to overweight subjects (BMI ≥ 27 kg / m²). 2 –<30 kg / m 2 Individuals who also have at least one weight-related comorbidity (such as hypertension, type 2 diabetes, or dyslipidemia).

[0159] In some implementations, weekly treatment resulted in statistically significant, dose-dependent weight loss.

[0160] In some preferred embodiments, the dosage is administered as an adjunct to diet and exercise to improve glycemic control in adults with type 2 diabetes.

[0161] After treatment initiation, it may be beneficial to administer escalating doses of amylin receptor agonists such as canagliflozin and GLP-1 receptor agonists such as smegglutide to individuals in need. Once an individual has adapted to the treatment, it may be beneficial to administer maintenance doses of amylin receptor agonists such as canagliflozin and GLP-1 receptor agonists such as smegglutide to individuals in need.

[0162] Treatment can be administered weekly, with a dose escalation period of up to 16 weeks.

[0163] Treatment can be administered weekly, with dose escalation performed approximately once a week.

[0164] Treatment can be given once a week, with the dosage increased approximately every other week.

[0165] Treatment can be given weekly, with dose escalation occurring approximately every three weeks.

[0166] Treatment can be given weekly, with doses escalating approximately every four weeks.

[0167] The dose of the amylin receptor agonist administered can be about 0.25-16 mg, for example about 0.25-9.0 mg, for example about 0.25-4.5 mg, for example about 0.25-2.4 mg.

[0168] The dosage of canagliflozin can be about 0.25-16 mg, for example about 0.25-9.0 mg, for example about 0.25-4.5 mg, for example about 0.25-2.4 mg.

[0169] The dosage of canagliflozin administered can be approximately 0.25 mg.

[0170] The dosage of canagliflozin administered can be approximately 0.5 mg.

[0171] The dosage of canagliflozin administered can be approximately 1.0 mg.

[0172] The dosage of canagliflozin administered can be approximately 1.5 mg.

[0173] The dosage of canagliflozin administered can be approximately 1.7 mg.

[0174] The dosage of canagliflozin administered can be approximately 2.4 mg.

[0175] The dosage of canagliflozin administered can be approximately 3.4 mg.

[0176] The dosage of canagliflozin administered can be approximately 3.6 mg.

[0177] The dosage of canagliflozin administered can be approximately 4.5 mg.

[0178] The dosage of canagliflozin administered can be approximately 7.2 mg.

[0179] The dosage of canagliflozin administered can be approximately 8.0 mg.

[0180] The dosage of canagliflozin administered can be approximately 9.0 mg.

[0181] The dosage of canagliflozin administered can be approximately 16.0 mg.

[0182] The dose of the GLP-1 receptor agonist can be about 0.25-16 mg, for example about 0.25-9.0 mg, for example about 0.25-4.5 mg, for example about 0.25-2.4 mg.

[0183] The dosage of smegglutide can be about 0.25-16 mg, for example about 0.25-9.0 mg, for example about 0.25-4.5 mg, for example about 0.25-2.4 mg.

[0184] The dosage of smegglutinin administered can be approximately 0.25 mg.

[0185] The dosage of smegglutinin administered can be approximately 0.5 mg.

[0186] The dosage of smegglutinin administered can be approximately 1.0 mg.

[0187] The dosage of smegglutinin administered can be approximately 1.5 mg.

[0188] The dosage of smegglutinin administered can be approximately 1.7 mg.

[0189] The dosage of smegglutinin administered can be approximately 2.4 mg.

[0190] The dosage of smegglutinin administered can be approximately 3.6 mg.

[0191] The dosage of smegglutinin administered can be approximately 4.5 mg.

[0192] The dosage of smegglutinin administered can be approximately 4.8 mg.

[0193] The dosage of smegglutinin administered can be approximately 6.0 mg.

[0194] The dosage of smegglutinin administered can be approximately 6.9 mg.

[0195] The dosage of smegglutinin administered can be approximately 7.2 mg.

[0196] The dosage of smegglutinin administered can be approximately 8.0 mg.

[0197] The dosage of smegglutinin administered can be approximately 9.0 mg.

[0198] The dosage of smegglutinin administered can be approximately 12 mg.

[0199] The dosage of semaglutide administered may be approximately 16.0 mg.

[0200] The ratio of amylin receptor agonist to GLP-1 receptor agonist can be approximately 1:2. The ratio of canagliflozin to smegglutinin can also be approximately 1:2.

[0201] The dosage of canagliflozin can be approximately 0.125 mg, and the dosage of smegglutinin can be approximately 0.25 mg.

[0202] The dosage of canagliflozin can be approximately 0.25 mg, and the dosage of smegglutinin can be approximately 0.5 mg.

[0203] The dosage of canagliflozin can be about 0.5 mg, and the dosage of smegglutinin can be about 1.0 mg.

[0204] The dosage of canagliflozin can be approximately 0.75 mg, and the dosage of smegglutinin can be approximately 1.5 mg.

[0205] The dosage of canagliflozin can be approximately 0.85 mg, and the dosage of smegglutinin can be approximately 1.7 mg.

[0206] The dosage of canagliflozin can be approximately 1.2 mg, and the dosage of smegglutinin can be approximately 2.4 mg.

[0207] The dosage of canagliflozin can be approximately 2.25 mg, and the dosage of smegglutinin can be approximately 4.5 mg.

[0208] The dose of canagliflozin administered may be approximately 3.6 mg, and the dose of smegglutinin may be approximately 7.2 mg.

[0209] The dosage of canagliflozin can be approximately 4.0 mg, and the dosage of smegglutinin can be approximately 8.0 mg.

[0210] The dosage of canagliflozin can be approximately 7.2 mg, and the dosage of smegglutinin can be approximately 14.4 mg.

[0211] The dosage of canagliflozin can be approximately 8.0 mg, and the dosage of smegglutinin can be approximately 16.0 mg.

[0212] The maintenance dose of canagliflozin can be about 1.2 mg, and the maintenance dose of smegglutinin can be about 2.4 mg.

[0213] The maintenance dose of canagliflozin can be approximately 2.25 mg, and the maintenance dose of smegglutinin can be approximately 4.5 mg.

[0214] The maintenance dose of canagliflozin can be approximately 4.0 mg, and the maintenance dose of smegglutinin can be approximately 8.0 mg.

[0215] The maintenance dose of canagliflozin can be approximately 8.0 mg, and the maintenance dose of smegglutinin can be approximately 16.0 mg.

[0216] The ratio of amylin receptor agonist to GLP-1 receptor agonist can be approximately 1:1. The ratio of canagliflozin to smegglutinin can also be approximately 1:1.

[0217] The dosage of canagliflozin can be approximately 0.25 mg, and the dosage of smegglutinin can be approximately 0.25 mg.

[0218] The dosage of canagliflozin can be approximately 0.5 mg, and the dosage of smegglutinin can be approximately 0.5 mg.

[0219] The dosage of canagliflozin can be approximately 1.0 mg, and the dosage of smegglutinin can be approximately 1.0 mg.

[0220] The maintenance dose of canagliflozin may be approximately 1.0 mg, and the maintenance dose of smegglutinin may be approximately 1.0 mg. The dose of canagliflozin may be approximately 1.7 mg, and the dose of smegglutinin may be approximately 1.7 mg.

[0221] The maintenance dose of canagliflozin can be approximately 1.7, and the maintenance dose of smegglutinin can be approximately 1.7.

[0222] The dosage of canagliflozin can be approximately 2.4 mg, and the dosage of smegglutinin can be approximately 2.4 mg.

[0223] The maintenance dose of canagliflozin can be approximately 2.4 mg, and the maintenance dose of smegglutinin can be approximately 2.4 mg.

[0224] The dosage of canagliflozin can be approximately 4.5 mg, and the dosage of smegglutinin can be approximately 4.5 mg.

[0225] The dosage of canagliflozin can be approximately 8.0 mg, and the dosage of smegglutinin can be approximately 8.0 mg.

[0226] The dosage of canagliflozin can be approximately 16.0 mg, and the dosage of smegglutinin can be approximately 16.0 mg.

[0227] The ratio of amylin receptor agonist to GLP-1 receptor agonist can be from 1:1 to 1:7.

[0228] The dosage of canagliflozin can be about 2.4 mg, and the dosage of smegglutinin can be from about 2.4 mg to 16.0 mg.

[0229] The dosage of canagliflozin can be about 2.4 mg, and the dosage of smegglutinin can be from about 3.6 mg to 16.0 mg.

[0230] The dosage of canagliflozin can be about 2.4 mg, and the dosage of smegglutinin can be from about 2.4 mg to 13.5 mg.

[0231] The dosage of canagliflozin can be about 2.4 mg, and the dosage of smegglutinin can be from about 3.6 mg to 13.5 mg.

[0232] The dosage of canagliflozin can be approximately 2.4 mg, and the dosage of smegglutinin can be approximately 3.6 mg.

[0233] The dosage of canagliflozin can be approximately 2.4 mg, and the dosage of smegglutinin can be approximately 4.8 mg.

[0234] The dosage of canagliflozin can be approximately 2.4 mg, and the dosage of smegglutinin can be approximately 6.0 mg.

[0235] The dosage of canagliflozin can be approximately 2.4 mg, and the dosage of smegglutinin can be approximately 6.9 mg.

[0236] The dosage of canagliflozin can be approximately 2.4 mg, and the dosage of smegglutinin can be approximately 7.2 mg.

[0237] The dosage of canagliflozin can be approximately 2.4 mg, and the dosage of smegglutinin can be approximately 8.0 mg.

[0238] The dosage of canagliflozin can be approximately 2.4 mg, and the dosage of smegglutinin can be approximately 12 mg.

[0239] The dosage of canagliflozin can be approximately 3.4 mg, and the dosage of smegglutinin can be approximately 13.5 mg.

[0240] Canagliflozin and smegglutinin can be administered once weekly at an initial dose of 0.25 mg, followed by increasing doses to 0.5 mg, 1.0 mg, and 1.7 mg until a target / maintenance dose of 2.4 mg once weekly is reached.

[0241] Canagliflozin and smegglutinin can be administered once weekly, with subsequent doses increasing to 0.5 mg, 1.0 mg, and 1.7 mg every four weeks until the target / maintenance dose of 2.4 mg once weekly is reached.

[0242] Canagliflozin and smegglutinin can be administered once weekly, with subsequent doses increasing to 0.5 mg, 1.0 mg, and 1.7 mg every four weeks until the target / maintenance dose of 2.4 mg once weekly is reached.

[0243] 0.25 mg canagliflozin and 0.25 mg smegglutide can be administered once weekly for four weeks (weeks 0-3), and every four weeks the dose can be increased to 0.5 mg canagliflozin and 0.5 mg smegglutide (weeks 4-7), 1.0 mg canagliflozin and 1.0 mg smegglutide (weeks 8-11), and 1.7 mg canagliflozin and 1.7 mg smegglutide (weeks 12-15) until the target / maintenance dose of 2.4 mg canagliflozin and 2.4 mg smegglutide once weekly (week 16 and thereafter) is reached.

[0244] Canagliflozin and smegglutinin can be administered once weekly at an initial dose of 0.25 mg, followed by increasing doses to 0.5 mg, 1.0 mg, 1.7 mg and 2.4 mg, until a target / maintenance dose of 4.5 mg once weekly is reached.

[0245] Canagliflozin and smegglutinin can be administered once weekly at an initial dose of 0.25 mg, followed by increasing doses of 0.5 mg, 1.0 mg, 1.7 mg, 2.4 mg, 3.6 mg and 4.5 mg until a target / maintenance dose of 7.2 mg once weekly is reached.

[0246] Canagliflozin and smegglutinin can be administered once weekly at an initial dose of 0.25 mg, followed by increasing doses of 0.5 mg, 1.0 mg, 1.7 mg, 2.4 mg, 3.6 mg, 4.5 mg and 7.2 mg until a target / maintenance dose of 8.0 mg once weekly is reached.

[0247] Canagliflozin and smegglutinin can be administered once weekly at an initial dose of 0.25 mg, followed by increasing doses of 0.5 mg, 1.0 mg, 1.7 mg, 2.4 mg, 3.6 mg, 4.5 mg, 7.2 mg and 8.0 mg until a target / maintenance dose of 16.0 mg once weekly is reached.

[0248] In this article, specific values ​​given as numbers or ranges can be interpreted as that specific value or an approximation (e.g., when certain quantities are available by weight, a specific value plus or minus 10%, 15%, or 20%; for example, when measuring pH, plus or minus 0.4).

[0249] Reagent test kit The pharmaceutical formulations disclosed herein may be presented in the form of a kit containing the pharmaceutical formulation and instructions for use. These instructions for use may include inserts from the pharmaceutical packaging.

[0250] The pharmaceutical formulations disclosed herein can be liquid formulations contained in an injection device. This injection device can be selected from a durable injection pen, a pre-filled injection pen, or a pre-filled syringe.

[0251] The pharmaceutical preparations disclosed herein may be liquid preparations contained in vials or cartridges.

[0252] The kit may include a dried formulation in a first vial, an aqueous solution in a second vial, and instructions for use, wherein the dried formulation is reconstituted in the aqueous solution to obtain the liquid pharmaceutical formulation disclosed herein.

[0253] The following is a non-limiting list of embodiments of the present invention.

[0254] Implementation Plan 1. A pharmaceutical formulation comprising an amylin receptor agonist, a GLP-1 receptor agonist, a cyclodextrin containing a hydroxypropyl substituted group, and at least one preservative and / or stabilizer.

[0255] 2. The pharmaceutical formulation according to embodiment 1, wherein the isoelectric point of the GLP-1 receptor agonist is incompatible with the optimal pH of the amylin receptor agonist.

[0256] 3. The pharmaceutical formulation according to any one of the foregoing embodiments, wherein the optimal pH of the GLP-1 receptor agonist and the optimal pH of the amylin receptor agonist differ by at least about 2 pH units, such as 2-5 pH units, such as 2-4 pH units, such as 3-5 pH units.

[0257] 4. The pharmaceutical formulation according to any one of the foregoing embodiments, wherein the optimal pH of the amylin receptor agonist is 3.5-4.5, for example, about 4.0.

[0258] 5. The pharmaceutical formulation according to any one of the foregoing embodiments, wherein the amylin receptor agonist is canagliflozin.

[0259] 6. The pharmaceutical formulation according to any one of the foregoing embodiments, wherein the isoelectric point of the GLP-1 receptor agonist is equal to or less than 5.0, for example equal to or less than 4.5; for example in the range of 3.0-5.0, for example in the range of 3.5-5.0, for example in the range of 3.5-4.5, for example in the range of 3.8-4.9, for example in the range of 4.0-4.5.

[0260] 7. The pharmaceutical formulation according to any one of the foregoing embodiments, wherein the GLP-1 receptor agonist comprises a GLP-1(7-37) derived peptide with an isoelectric point equal to or less than 5.0, for example less than 4.5, for example in the range of 3.0-5.0, such as 3.5-5.0, such as 3.5-4.5, such as 3.8-4.9, such as 4.0-4.5.

[0261] 8. The pharmaceutical formulation according to embodiment 1, wherein the theoretically calculated isoelectric point (pI) of the amylin receptor agonist is in the range of 7.6-9.4 or 8-9.

[0262] 9. The pharmaceutical formulation according to embodiment 1, wherein the amylin receptor agonist comprises a human amylin-derived peptide with a theoretically calculated isoelectric point (pI) in the range of 7.6-9.4 or 8-9.

[0263] 10. The pharmaceutical formulation according to any one of the foregoing embodiments, wherein the GLP-1 receptor agonist is semaglutide or telpolide.

[0264] 11. The pharmaceutical formulation according to any one of the foregoing embodiments, wherein the GLP-1 receptor agonist is semaglutide.

[0265] 12. The pharmaceutical formulation according to any one of the foregoing embodiments, wherein the cyclodextrin is an α-cyclodextrin comprising six cyclic glucose units and / or a β-cyclodextrin comprising seven cyclic glucose units.

[0266] 13. The pharmaceutical formulation according to any one of the foregoing embodiments, wherein the cyclodextrin is a hydroxypropyl-substituted α-cyclodextrin comprising six cyclically arranged glucose units.

[0267] 14. The pharmaceutical formulation according to any one of the foregoing embodiments, wherein the cyclodextrin is a hydroxypropyl-substituted β-cyclodextrin comprising seven cyclically arranged glucose units.

[0268] 15. The pharmaceutical formulation according to any one of the foregoing embodiments, wherein the cyclodextrin comprises up to about 1.2 hydroxypropyl groups per glucose unit.

[0269] 16. The pharmaceutical formulation according to any one of the foregoing embodiments, wherein the cyclodextrin comprises up to about 0.92 hydroxypropyl groups per glucose unit.

[0270] 17. The pharmaceutical formulation according to any one of the foregoing embodiments, wherein the cyclodextrin comprises up to about 0.75 hydroxypropyl groups per glucose unit.

[0271] 18. The pharmaceutical formulation according to any one of the foregoing embodiments, wherein the cyclodextrin comprises up to about 0.68 hydroxypropyl groups per glucose unit.

[0272] 19. The pharmaceutical preparation according to any one of the foregoing embodiments, wherein the cyclodextrin comprises at least about 0.4 hydroxypropyl groups per glucose unit.

[0273] 20. The pharmaceutical formulation according to embodiment 19, wherein the cyclodextrin comprises at least about 0.58 hydroxypropyl groups per glucose unit.

[0274] 21. The pharmaceutical formulation according to embodiment 20, wherein the cyclodextrin comprises about 0.58-1.0 hydroxypropyl groups per glucose unit.

[0275] 22. The pharmaceutical formulation according to embodiment 21, wherein the cyclodextrin comprises an average (MS) 0.62-0.92 hydroxypropyl groups per glucose unit.

[0276] 23. The pharmaceutical formulation according to embodiment 21, wherein the cyclodextrin comprises an average (MS) of about 0.62-0.84 hydroxypropyl groups per glucose unit.

[0277] 24. The pharmaceutical formulation according to embodiment 21, wherein the cyclodextrin comprises an average of about 0.62 hydroxypropyl groups per glucose unit (MS).

[0278] 25. The pharmaceutical preparation according to any one of the foregoing embodiments is a liquid preparation.

[0279] 26. The pharmaceutical formulation according to embodiment 25, wherein the pH is about 5.6-6.4, for example about 5.8-6.2, for example about 5.6, for example about 5.7, for example about pH 5.8, for example about 5.9, for example about 6.0, for example about 6.1, for example about 6.2, for example about 6.3, for example about 6.4, preferably 5.8-6.2.

[0280] 27. The pharmaceutical preparation according to any one of embodiments 25-26, comprising more than 10% w / v cyclodextrin.

[0281] 28. The pharmaceutical preparation according to any one of embodiments 25-27, comprising less than 22% w / v cyclodextrin.

[0282] 29. The pharmaceutical preparation according to any one of embodiments 25-28, comprising less than 20% w / v cyclodextrin.

[0283] 30. The pharmaceutical preparation according to any one of embodiments 25-28, comprising about 10-20% w / v of the cyclodextrin.

[0284] 31. The pharmaceutical preparation according to any one of embodiments 25-30, comprising about 10-17.5% w / v cyclodextrin.

[0285] 32. The pharmaceutical preparation according to any one of embodiments 25-31, comprising about 12-18% w / v cyclodextrin.

[0286] 33. The pharmaceutical preparation according to any one of embodiments 25-32, comprising about 11.25-15% w / v cyclodextrin.

[0287] 34. The pharmaceutical preparation according to any one of embodiments 25-32, comprising about 15% w / v cyclodextrin.

[0288] 35. A pharmaceutical formulation according to any one of embodiments 25-34, comprising at least about 1 mg / ml of the GLP-1 receptor agonist.

[0289] 36. A pharmaceutical formulation according to any one of embodiments 25-35, comprising up to about 22 mg / ml of the GLP-1 receptor agonist.

[0290] 37. The pharmaceutical formulation according to any one of embodiments 25-36, comprising about 1-12 mg / ml of a GLP-1 receptor agonist.

[0291] 38. The pharmaceutical formulation according to any one of embodiments 25-37, comprising at least about 1 mg / ml of the said amylin receptor agonist.

[0292] 39. The pharmaceutical formulation according to any one of embodiments 25-38, comprising up to about 30 mg / ml of an amylin receptor agonist.

[0293] 40. The pharmaceutical formulation according to any one of embodiments 25-39, comprising up to about 22 mg / ml of an amylin receptor agonist.

[0294] 41. The pharmaceutical formulation according to any one of embodiments 25-40, comprising about 1-12 mg / ml of an amylin receptor agonist.

[0295] 42. The pharmaceutical preparation according to any one of embodiments 25-41, comprising 0.25-22 mg / ml of canagliflozin.

[0296] 43. The pharmaceutical preparation according to any one of embodiments 25-42, comprising 0.25-22 mg / ml of smegglutide.

[0297] 44. The pharmaceutical preparation according to any one of embodiments 25-42, comprising 5-30 mg / ml of telpolide.

[0298] 45. A pharmaceutical preparation according to any one of embodiments 25-43, comprising 0.25-22 mg / ml canagliflozin and 0.25-22 mg / ml smegglutinin.

[0299] 46. ​​The pharmaceutical preparation according to any one of embodiments 25-41, comprising an effective amount of canagliflozin and smegglutinin or telpolide.

[0300] 47. The pharmaceutical formulation according to any one of the foregoing embodiments, wherein the at least one preservative is phenol and / or m-cresol and / or EDTA.

[0301] 48. The pharmaceutical preparation according to any one of embodiments 25-47, comprising m-cresol at a concentration of 9-40 mM.

[0302] 49. The pharmaceutical preparation according to any one of embodiments 25-48, comprising phenol at a concentration of 18-65 mM.

[0303] 50. The pharmaceutical preparation according to any one of embodiments 25-49, comprising m-cresol at a concentration of 9-40 mM and phenol at a concentration of 18-65 mM.

[0304] 51. The pharmaceutical formulation according to any one of the foregoing embodiments, wherein the at least one stabilizer is EDTA.

[0305] 52. The pharmaceutical preparation according to any one of embodiments 25-51, comprising EDTA at a concentration of 0.5-5.0 mg / ml.

[0306] 53. A pharmaceutical preparation comprising canagliflozin, smegglutinin, hydroxypropyl-substituted α- and / or β-cyclodextrin, m-cresol, phenol, and EDTA; and having a pH of 5.6-6.4, preferably 5.8-6.2.

[0307] 54. A pharmaceutical formulation comprising canagliflozin, smegglutinin, hydroxypropyl-substituted α- and / or β-cyclodextrin and EDTA; and having a pH of 5.6-6.4, preferably 5.8-6.2.

[0308] 55. The pharmaceutical preparation according to any one of the foregoing embodiments further comprises a tensile agent, provided that the tensile agent is not sodium chloride.

[0309] 56. The pharmaceutical formulation according to the foregoing embodiments, wherein the tonic agent is glycerol, mannitol, propylene glycol, sorbitol or trehalose, or a combination thereof.

[0310] 57. The pharmaceutical formulation according to embodiment 56, wherein the tonic agent is glycerol.

[0311] 58. The pharmaceutical preparation according to any one of embodiments 25-57, comprising glycerol at a concentration of about 2.5-18 mg / ml.

[0312] 59. The pharmaceutical formulation according to embodiment 56, wherein the tonic agent is mannitol.

[0313] 60. A pharmaceutical preparation according to any one of embodiments 25-59, comprising mannitol at a concentration of about 16.5-37.5 mg / ml, for example about 20 mg / ml.

[0314] 61. The pharmaceutical formulation according to embodiment 56, wherein the tonic agent is propylene glycol.

[0315] 62. The pharmaceutical preparation according to any one of embodiments 25-61, comprising propylene glycol at a concentration of about 2-15 mg / ml.

[0316] 63. The pharmaceutical formulation according to embodiment 56, wherein the tonic agent is sorbitol.

[0317] 64. A pharmaceutical preparation according to any one of embodiments 25-63, comprising sorbitol at a concentration of about 5-35 mg / ml, for example about 10-30 mg / ml, for example about 16-28 mg / ml, for example about 16.5-25 mg / ml, for example about 16-24 mg / ml, for example about 24 mg / ml, for example about 20 mg / ml, for example about 16 mg / ml, for example about 12 mg / ml.

[0318] 65. The pharmaceutical formulation according to embodiment 56, wherein the tonic agent is trehalose.

[0319] 66. The pharmaceutical preparation according to any one of embodiments 25-65, comprising trehalose at a concentration of about 33-75 mg / ml, for example about 33-45 mg / ml, for example about 38 mg / ml.

[0320] 67. The pharmaceutical formulation according to any one of the foregoing embodiments further comprises a buffer having at least one pKa of about 5.0-7.0.

[0321] 68. The pharmaceutical formulation according to any one of the foregoing embodiments further comprises a buffer selected from histidine, citrate and / or phosphate.

[0322] 69. The pharmaceutical preparation according to any one of embodiments 25-68, comprising a buffer of up to 30 mM.

[0323] 70. The pharmaceutical preparation according to any one of embodiments 25-69, comprising about 3-30 mM of citrate.

[0324] 71. The pharmaceutical preparation according to any one of embodiments 25-70, comprising about 3-30 mM histidine, such as 3-15 mM histidine, such as 3-10 mM histidine, for example about 6 mM histidine.

[0325] 72. The pharmaceutical preparation according to any one of embodiments 25-71, comprising about 3-30 mM of phosphate.

[0326] 73. The pharmaceutical formulation according to any one of the foregoing embodiments further comprises a surfactant.

[0327] 74. The pharmaceutical formulation according to the foregoing embodiments, wherein the surfactant is polysorbate 20 and / or polysorbate 80.

[0328] 75. The pharmaceutical formulation according to any one of embodiments 25-74, comprising up to about 2.0 mg / ml of polysorbate 20 and / or polysorbate 80.

[0329] 76. The pharmaceutical formulation according to embodiment 75, comprising 0.01-0.1 mg / ml, for example about 0.05 mg / ml, of polysorbate 20 and / or polysorbate 80; or more than 0.1 mg / ml and less than 0.2 mg / ml of polysorbate 20 and / or polysorbate 80.

[0330] 77. The pharmaceutical formulation according to any one of embodiments 73-76, wherein the surfactant is polysorbate 80.

[0331] 78. The pharmaceutical formulation according to the foregoing embodiments comprises 0.01-0.1 mg / ml of polysorbate 80, preferably 0.05 mg / ml of polysorbate 80.

[0332] 79. A pharmaceutical preparation according to any one of embodiments 25-78, comprising at least 75% w / w water, for example about 80% w / w water, for example about 85% w / w water, for example up to about 90% w / w water.

[0333] 80. The pharmaceutical formulation according to any one of the foregoing embodiments is substantially composed of the following components: effective amounts of canagliflozin and smegglutinin; hydroxypropyl-substituted α- and / or β-cyclodextrins, said cyclodextrins comprising 0.58-1.0 hydroxypropyl groups per glucose unit; phenol and / or m-cresol; EDTA; histidine; sorbitol; polysorbate 20 and / or 80; and about 75-90% w / w water; and a pH of 5.6-6.4, preferably about 5.8-6.2.

[0334] 81. The pharmaceutical formulation according to any one of the foregoing embodiments is substantially composed of the following components: effective amounts of canagliflozin and smegglutinin; hydroxypropyl-substituted α- and / or β-cyclodextrins, said cyclodextrins comprising an average of 0.62-0.92 hydroxypropyl groups per glucose unit; phenol and / or m-cresol; EDTA; histidine; sorbitol; polysorbate 20 and / or 80; and about 75-90% w / w water; and a pH of 5.6-6.4, preferably about 5.8-6.2.

[0335] 82. A pharmaceutical preparation comprising: -0.25-22 mg / ml canagliflozin, Smegglutide at concentrations of -0.25 to 22 mg / ml - Hydroxypropyl-substituted α- and / or β-cyclodextrins, such as 10-20% w / v, exceeding 10% w / v and less than 22% w / v, wherein the cyclodextrin contains 0.58-1.0 hydroxypropyl groups per glucose unit. - Approximately 18-65 mM phenol and / or 9-40 mM m-cresol and / or 0.5-5.0 mg / ml EDTA, - A buffer having at least one pKa value of approximately 5.0-7.0. - Approximately 5-35 mg / ml of sorbitol, - Up to 1.0 mg / ml of polysorbate 20 and / or 80, such as 0.01-0.1 mg / ml, for example about 0.05 mg / ml of polysorbate 20 and / or 80, or more than 0.1 mg / ml but less than 0.2 mg / ml of polysorbate 20 and / or 80. - Approximately 75-90% w / w of water And the pH is 5.6-6.4, such as 5.7-6.4, preferably 5.8-6.2.

[0336] 83. A pharmaceutical preparation comprising: -0.25-22 mg / ml canagliflozin, Smegglutide at concentrations of -0.25 to 22 mg / ml - Hydroxypropyl-substituted α- and / or β-cyclodextrins, such as 10-20% w / v, exceeding 10% w / v and less than 22% w / v, wherein the cyclodextrin contains 0.58-1.0 hydroxypropyl groups per glucose unit. - Approximately 18-65 mM phenol and / or 9-40 mM m-cresol and / or 0.5-5.0 mg / ml EDTA, - Approximately 3-30 mM of histidine, - Approximately 5-35 mg / ml of sorbitol, - Up to 1.0 mg / ml of polysorbate 20 and / or 80, such as 0.01-0.1 mg / ml, for example about 0.05 mg / ml of polysorbate 20 and / or 80, or more than 0.1 mg / ml but less than 0.2 mg / ml of polysorbate 20 and / or 80. - Approximately 75-90% w / w of water And the pH is 5.6-6.4, for example about 5.7-6.4, preferably about 5.8-6.2.

[0337] 84. A pharmaceutical preparation comprising: -0.25-22 mg / ml canagliflozin, Smegglutide at concentrations of -0.25 to 22 mg / ml - Hydroxypropyl-substituted α- and / or β-cyclodextrins, such as 10-20% w / v, exceeding 10% w / v and less than 22% w / v, wherein the cyclodextrin contains 0.58-1.0 hydroxypropyl groups per glucose unit. - Approximately 18-65 mM phenol and / or 9-40 mM m-cresol and / or 0.5-5.0 mg / ml EDTA, - Approximately 3-30 mM citrate, - Approximately 5-35 mg / ml of sorbitol, - Up to 1.0 mg / ml of polysorbate 20 and / or 80, such as 0.01-0.1 mg / ml, such as 0.05 mg / ml of polysorbate 20 and / or 80, or more than 0.1 mg / ml but less than 0.2 mg / ml of polysorbate 20 and / or 80. - Approximately 75-90% w / w of water, And the pH is 5.6-6.4, for example about 5.7-6.4, preferably about 5.8-6.2.

[0338] 85. The pharmaceutical preparation according to any one of the foregoing embodiments is substantially composed of the following components: - Effective amounts of canagliflozin and smegglutinin - Hydroxypropyl-substituted α- and / or β-cyclodextrins, such as 10-20% w / v, exceeding 10% w / v and less than 22% w / v, wherein the cyclodextrin contains 0.58-1.0 hydroxypropyl groups per glucose unit. - Approximately 3-30 mM of histidine, - Approximately 5-35 mg / ml of sorbitol, - Up to 1.0 mg / ml of polysorbate 20 and / or 80, such as 0.01-0.1 mg / ml, such as 0.05 mg / ml of polysorbate 20 and / or 80, or more than 0.1 mg / ml but less than 0.2 mg / ml of polysorbate 20 and / or 80. - The pH is 5.6-6.4, for example about 5.7-6.4, preferably about 5.8-6.2. - Water for injection.

[0339] 86. The pharmaceutical preparation according to any one of the foregoing embodiments is substantially composed of the following components: -0.25-22 mg / ml canagliflozin, Smegglutide at concentrations of -0.25 to 22 mg / ml - Hydroxypropyl-substituted α- and / or β-cyclodextrins, such as 10-20% w / v, exceeding 10% w / v and less than 22% w / v, wherein the cyclodextrin contains 0.58-1.0 hydroxypropyl groups per glucose unit. - Approximately 3-30 mM, preferably approximately 6 mM of histidine. - Approximately 5-35 mg / ml of sorbitol, - Up to 1.0 mg / ml of polysorbate 20 and / or 80; such as 0.01-0.1 mg / ml, such as 0.05 mg / ml; or more than 0.1 mg / ml but less than 0.2 mg / ml or about 0.1-0.2 mg / ml of polysorbate 20 and / or 80. - The pH is 5.6-6.4, for example about 5.7-6.4, preferably about 5.8-6.2. - Water for injection.

[0340] 87. The pharmaceutical preparation according to any one of the foregoing embodiments, which is used as a medicine.

[0341] 88. A pharmaceutical preparation according to any one of embodiments 1-86, used for the treatment or prevention of obesity.

[0342] 89. A pharmaceutical preparation according to any one of embodiments 1-86, used for treating or preventing overweight in a subject suffering from at least one weight-related comorbidity.

[0343] 90. A pharmaceutical preparation according to any one of embodiments 1-86, for the treatment or prevention of at least one weight-related comorbidity in a subject who is overweight or obese at the start of treatment.

[0344] 91. A pharmaceutical preparation according to any one of embodiments 1-86, used for treating or preventing obesity in a subject, said subject having an initial body mass index (BMI) of 25 kg / m². 2 Or higher, 27 kg / m 2 Or higher, 28 kg / m 2 Or higher or 30 kg / m 2 Or higher.

[0345] 92. A pharmaceutical preparation according to any one of embodiments 1-86, used to treat overweight in a subject, said subject having an initial body mass index (BMI) of 24 kg / m². 2 Or higher, 25 kg / m 2Or higher or 27 kg / m 2 Or higher, and have at least one weight-related comorbidity.

[0346] 93. The pharmaceutical preparation according to any one of embodiments 1-86, wherein the initial body mass index (BMI) is 25 kg / m². 2 27 kg / m 2 28 kg / m 2 Or 30 kg / m 2 Or higher (obese) or an initial body mass index (BMI) of 24 kg / m² 2 25 kg / m 2 Or 27 kg / m 2 In adult subjects who are overweight or higher and have at least one weight-related comorbidity, it is used as an adjunct to a low-calorie diet and increased physical activity for the treatment or prevention of overweight or obesity.

[0347] 94. The pharmaceutical preparation according to any one of embodiments 1-86, used as an adjunct to a low-calorie diet and increased physical activity for individuals with an initial body mass index (BMI) of 30 kg / m². 2 Or higher (obese) or 27 kg / m 2 Long-term weight management in adult subjects who are overweight or higher and have at least one weight-related comorbidity.

[0348] 95. Use of a liquid pharmaceutical preparation according to any one of embodiments 87-94, wherein the at least one comorbidity is diabetes and / or one or more cardiovascular diseases (CVD) and / or non-alcoholic steatohepatitis (NASH) or metabolic dysfunction-associated steatohepatitis (MASH) and / or alcoholic liver disease (ALD) and / or obstructive sleep apnea.

[0349] 96. A pharmaceutical preparation according to any one of embodiments 1-86, used to treat a subject suffering from diabetes, such as type II diabetes.

[0350] 97. A pharmaceutical preparation according to any one of embodiments 1-86, for treating a subject suffering from diabetes such as type 2 diabetes and at least one comorbidity associated with diabetes.

[0351] 98. A pharmaceutical preparation according to any one of embodiments 1-86, used as an adjunct to diet and exercise to improve glycemic control in adults with type 2 diabetes.

[0352] 99. A pharmaceutical preparation according to any one of embodiments 1-86, for the treatment and / or prevention of chronic kidney disease (CKD) in a subject with diabetes.

[0353] 100. A pharmaceutical preparation according to any one of embodiments 1-86, for the treatment and / or prevention of one or more cardiovascular diseases (CVD).

[0354] 101. A pharmaceutical preparation according to any one of embodiments 1-86, for the treatment and / or prevention of non-alcoholic steatohepatitis (NASH), metabolic dysfunction-associated steatohepatitis (MASH), and / or alcoholic liver disease (ALD).

[0355] 102. A pharmaceutical preparation according to any one of embodiments 1-86, for the treatment and / or prevention of obstructive sleep apnea in subjects who are overweight or obese at the start of treatment.

[0356] 103. A pharmaceutical preparation according to any one of embodiments 1-86, used for the treatment and / or prevention of cognitive impairment, such as cognitive impairment caused by Alzheimer's disease.

[0357] 104. A pharmaceutical preparation according to any one of embodiments 1-86 for use according to any one of embodiments 87-103, characterized in that the preparation is administered by parenteral injection.

[0358] 105. A pharmaceutical preparation according to any one of embodiments 1-86 for use according to any one of embodiments 87-103, characterized in that the preparation is administered by subcutaneous injection.

[0359] 106. A pharmaceutical preparation according to any one of embodiments 1-86 for use according to any one of embodiments 87-103, characterized in that the preparation is applied approximately once a week.

[0360] 107. A pharmaceutical preparation according to any one of embodiments 1-86 for use according to any one of embodiments 87-106, wherein the ratio of the dose of canagliflozin administered to the dose of smegglutinin administered is approximately 1:1.

[0361] 108. A pharmaceutical preparation according to any one of embodiments 1-86 for use according to any one of embodiments 87-106, wherein the ratio of the dose of canagliflozin administered to the dose of smegglutinin administered is 1:1 to 1:7.

[0362] 109. A pharmaceutical preparation according to any one of embodiments 1-86 for use according to any one of embodiments 87-106, wherein the ratio of the dose of canagliflozin administered to the dose of smegglutinin administered is approximately 1:2.

[0363] 110. A kit comprising a pharmaceutical composition as defined in any one of embodiments 1-86 and instructions for use.

[0364] 111. A kit comprising a vial containing a pharmaceutical preparation according to any one of embodiments 1-86, and instructions for use.

[0365] 112. The kit according to embodiment 110, wherein the pharmaceutical preparation is a liquid pharmaceutical preparation.

[0366] 113. The kit according to any one of embodiments 110-112, wherein the liquid pharmaceutical formulation comprises: -0.25-22 mg / ml canagliflozin, Smegglutide at concentrations of -0.25 to 22 mg / ml - Hydroxypropyl-substituted α- and / or β-cyclodextrins, such as 10-20% w / v, exceeding 10% w / v and less than 22% w / v, wherein the cyclodextrin contains 0.58-1.0 hydroxypropyl groups per glucose unit. - Approximately 18-65 mM phenol and / or 9-40 mM m-cresol and / or 0.5-5.0 mg / ml EDTA, - Buffers with at least one pKa value of about 5.0-7.0, such as 3-30 mM histidine or citrate. - Approximately 5-35 mg / ml of sorbitol, - Polysorbate 20 and / or 80 at concentrations greater than 0.1 mg / ml and less than 0.2 mg / ml - Approximately 75-90% w / w of water, and The pH is 5.6-6.4, for example about 5.7-6.4, preferably about 5.8-6.2.

[0367] 114. The kit according to embodiment 110, comprising: - The freeze-dried or spray-dried formulation in the first vial -The aqueous solution in the second vial, and -Instruction manual The mixture of the contents of the two vials provides a liquid pharmaceutical preparation according to any one of embodiments 25-86.

[0368] 115. A kit comprising a freeze-dried or spray-dried formulation in a first vial, an aqueous solution in a second vial, and instructions for use. The first vial contains the following formulation: -Cagglitazone, -Smigratide, - Hydroxypropyl-substituted α- and / or β-cyclodextrins, wherein the cyclodextrin comprises 0.58-1.0 hydroxypropyl groups per glucose unit, -Optional, EDTA -Optionally, a buffer having at least one pKa value of about 5.0-7.0, -Optional, sorbitol - Polysorbate 20 and / or 80; And the aqueous solution in the second vial contains -Water for injection -Optionally, phenol and / or m-cresol -Optional, EDTA - Optionally, a buffer having at least one pKa value of about 5.0-7.0, such as histidine or citrate, -Optional, sorbitol.

[0369] 116. The kit according to embodiment 115, wherein the freeze-dried or spray-dried formulation is dissolved in the aqueous solution to obtain a liquid pharmaceutical formulation comprising the following components: -0.25-22 mg / ml canagliflozin, Smegglutide at concentrations of -0.25 to 22 mg / ml - Hydroxypropyl-substituted α- and / or β-cyclodextrins, such as 10-20% w / v, exceeding 10% w / v and less than 22% w / v, wherein the cyclodextrin contains 0.58-1.0 hydroxypropyl groups per glucose unit. - Approximately 18-65 mM phenol and / or 9-40 mM m-cresol and / or 0.5-5.0 mg / ml EDTA, - Buffers with at least one pKa value of about 5.0-7.0, such as histidine or citrate. - Approximately 5-35 mg / ml of sorbitol, - Polysorbate 20 and / or 80 at concentrations greater than 0.1 mg / ml and less than 0.2 mg / ml - Approximately 75-90% w / w of water, and The pH is 5.6-6.4, for example about 5.7-6.4, preferably about 5.8-6.2.

[0370] 117. A kit comprising a pharmaceutical composition as defined in any one of embodiments 1-86 and an injection device for administering the pharmaceutical composition to a subject, wherein the injection device is selected from durable injection pens, pre-filled injection pens, and pre-filled syringes.

[0371] Example Example 1: Effect of hydroxypropyl-β-cyclodextrin (HP-B-CD) on the chemical stability of canagliflozin This embodiment demonstrates the ability of HP-B-CD to chemically stabilize canagliflozin, with chemical stability measured from the perspective of canagliflozin purity and canagliflozin-associated high molecular weight protein (HMWP).

[0372] Canagliflozin exhibits optimal stability at pH 4.0, and its chemical degradation rate typically increases with increasing pH. Surprisingly, when canagliflozin is formulated with HP-B-CD, a stable canagliflozin formulation was obtained at pH 6.

[0373] composition The compositions of canagliflozin formulations 1, 2 and 3 are shown in Table 1.

[0374] Table 1 Composition of canagliflozin formulations 1, 2 and 3

[0375] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. 2 Different buffer concentrations are used between formulations to ensure buffering at different pH levels. Preparation process Each canagliflozin formulation was prepared as follows: First, the excipients were dissolved in water, and then the canagliflozin active pharmaceutical ingredient was dissolved in the excipient solution. The pH of the solution was adjusted, and water was added to reach the final desired volume. The solution was then filtered through a 0.22 μm sterile filter for sterilization. After filtration, the formulation was filled into 1 ml pre-filled syringes.

[0376] method Samples were stored at 37°C for up to 21 days. After 14 and 21 days, the samples were analyzed to determine the purity levels of HMWP and canagliflozin.

[0377] The levels of covalently bound HMWPs were quantified using size exclusion chromatography (SEC). A Waters HMWP column (7.8 x 300 mm) was used, with isocratic elution consisting of 500 mM sodium chloride, 10 mM sodium dihydrogen phosphate monohydrate, 5 mM orthophosphate, and 50% (v / v) isopropanol. Chromatographic analysis was performed at 50 °C using a 10 μL injection volume and a flow rate of 0.5 mL / min, with UV detection (215 nm). HMWPs were quantified by dividing the area of ​​all components eluted before the main peak by the area of ​​the main peak multiplied by 100%.

[0378] The purity of canagliflozin was determined by reversed-phase ultra-high performance liquid chromatography (RP-UHPLC). A Kinetex C18 1.7 μm, 100 Å column (2.1 x 150 mm) was used for analysis. Sedimentation was performed using eluent A and eluent B, consisting of 90% v / v 0.09 M phosphate solution (pH 3.6) and 10% v / v acetonitrile, while eluent B consisted of 60% v / v acetonitrile and 20% v / v isopropanol. Chromatographic analysis was performed at 60 °C using an injection volume of 2–7.5 μl and a flow rate of 0.25 ml / min, with UV detection (215 nm). Purity was assessed by dividing the area of ​​the main peak by the area of ​​all peaks multiplied by 100%.

[0379] Table 2. Chemical purity (%) of canagliflozin at pH 4.0 and 6.0 in the presence and absence of HP-B-CD.

[0380] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. Concluding remarks Table 2 shows that when canagliflozin was stored at 37°C and pH 4.0, very little HMWP was formed, and only a slight decrease in canagliflozin purity was observed. In contrast, at pH 6.0, the rate of HMWP formation and the decrease in canagliflozin purity accelerated. Surprisingly, this rapid chemical degradation was counteracted by adding HP-B-CD to the formulation, allowing canagliflozin to be formulated at pH 6.

[0381] Example 2: Effect of HP-B-CD on the physical stability of smegglutinin This example demonstrates the ability of HP-B-CD to physically stabilize semaglutide, which has a tendency to form peptide fibrils. This effect is significant when semaglutide is formulated at a suboptimal pH.

[0382] composition The compositions of smegglutide formulations 1, 2 and 3 are shown in Table 3.

[0383] Table 3 Composition of Smegglutide formulations 1, 2 and 3

[0384] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. 2 Different buffer concentrations are used between formulations to ensure buffering at different pH levels. Preparation process The formulation was prepared as described in Example 1.

[0385] method The tendency of smegglutinin to aggregate and form peptide fibrils was measured using thioflavin T (ThT) fluorescence stress assays; these are parameters used to quantify physical stability. Analysis of the presence of peptide fibrils was based on the fluorescence properties of the ThT probe, which exhibits low fluorescence in its unbound / native peptide-bound state but high fluorescence when bound to peptide fibrils, with a redshift at the wavelength of maximum fluorescence when bound to fibrils.

[0386] Two samples were combined, and 1400 μl of the sample was added to 28 μl of 1 mM ThT stock solution. Then, 200 μl of this solution was transferred to six different wells of a 96-well microtiter plate containing glass beads. The experiment was run for 169 hours at 40°C with dual-track shaking at 300 rpm on a BMG CLARIOstar fluorescence reader equipped with monochromators for excitation and emission at 450 nm and 480 nm, respectively. The hysteresis time from the start of the experiment until fibril formation was measured, showing an increase in ThT fluorescence.

[0387] Table 4. Physical stability of smegglutinin at pH 6.0 and 7.4

[0388] 1 The result is the average of 6 replicates. 2 No fibril formation was observed in any of the six replicates during the 169-hour experimental period. 3 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. Concluding remarks The semagraviride formulations were subjected to induced shear stress, and the tendency of semagraviride to form peptide fibrils was measured. Surprisingly, the presence of HP-B-CD was found to inhibit semagraviride fibril formation. When semagraviride was formulated at pH 6 in the absence of HP-B-CD (semagraviride formulation 1), fibril formation occurred after 2.35 hours; that is, semagraviride is physically unstable. However, when semagraviride was formulated at pH 6 in the presence of HP-B-CD (semagraviride formulation 2), no fibril formation was observed throughout the entire experimental period; that is, semagraviride is physically stable. Furthermore, the physical stability of semagraviride formulated at pH 6 in the presence of HP-B-CD (semagraviride formulation 2) was found to be comparable to that of semagraviride formulated in the absence of HP-B-CD but under its optimal formulation conditions (pH 7.4) (semagraviride formulation 3).

[0389] Example 3: Effect of HP-B-CD on the chemical stability of smegglutinin This embodiment demonstrates the ability of HP-B-CD to chemically stabilize semaglutide, with chemical stability measured from the perspective of semaglutide purity and semaglutide-associated high molecular weight protein (HMWP).

[0390] composition The same formulation as in Example 2 was used.

[0391] Preparation process The formulation was prepared as described in Example 1.

[0392] method The purity levels of HMWP and smegglutinin were determined after storage at 37°C for 0, 14, and 21 days.

[0393] The purity of smegglutide was determined by reversed-phase high-performance liquid chromatography (RP-HPLC) using a Kinetex C18 2.6 μm column (4.6 x 150 mm). Sequencing was employed with eluent A and eluent B, consisting of 90% v / v 0.09 M phosphate solution (pH 3.6) and 10% v / v acetonitrile, and eluent B consisting of 60% v / v acetonitrile and 20% v / v isopropanol. Chromatographic analysis was performed at 30 °C using an injection volume of 10–100 μL and a flow rate of 0.7 mL / min, with UV detection (210 nm). Purity was quantified by dividing the area of ​​the main peak by the area of ​​all peaks multiplied by 100%.

[0394] The levels of covalently bound HMWPs were determined using size exclusion chromatography (SEC). Samples were analyzed using a Waters SEC 1.7 μm column (4.6 x 150 mm) with isocratic elution consisting of 300 mM sodium chloride, 10 mM sodium dihydrogen phosphate, 5 mM orthophosphate, and 50% v / v 2-propanol. Chromatographic analysis was performed at 50 °C using UV detection (280 nm) with injection volumes of 1–10 μL and a flow rate of 0.3 mL / min. HMWPs were quantified by dividing the area of ​​all components eluted before the main peak by the area of ​​the main peak multiplied by 100%.

[0395] Table 5. Purity of semaglutide at suboptimal and optimal pH values.

[0396] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. Concluding remarks The results in Table 5 show that the chemical purity of semaglutide decreased over time. When semaglutide was formulated at pH 6.0 (semaglutide formulation 1), its chemical purity decreased more rapidly than when it was formulated at its optimal pH 7.4 (semaglutide formulation 3). Surprisingly, when formulated at pH 6.0 (semaglutide formulation 2), HP-B-CD improved the chemical stability of semaglutide (in terms of purity decrease and HMWP formation).

[0397] Example 4: Effect of hydroxypropyl-β-cyclodextrin concentration on the chemical stability of smegglutinin This example demonstrates the concentration-dependent effect of HP-B-CD on the chemical stability of smegglutinin.

[0398] composition The composition of co-preparations containing different concentrations of HP-B-CD is shown in Table 6.

[0399] Table 6 contains the composition of co-formulations containing different concentrations of HP-B-CD.

[0400] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. 2 Because the HP-B-CD concentrations tested are different, different sorbitol concentrations are required to obtain isotonicity. preparation The formulation was prepared as described in Example 1.

[0401] method The samples were stored at 37°C for 28 days, and analyzed after 14, 21, and 28 days to determine the chemical purity of smegglutinin.

[0402] The purity of smegglutinin was determined by reversed-phase ultra-high performance liquid chromatography (RP-UHPLC). A Waters Acquity phenyl-hexyl 1.7 μm column (2.1 x 150 mm) was used for analysis with gradient elution using eluent A and eluent B. Eluent A consisted of 0.09% TFA in MQ water, while eluent B consisted of 0.09% TFA in 80% acetonitrile. Chromatographic analysis was performed at 62 °C using an injection volume of 2–14 μL and a flow rate of 0.25 mL / min, with UV detection (215 nm). Purity was assessed by dividing the area of ​​the smegglutinin main peak by the area of ​​all relevant peaks multiplied by 100%.

[0403] Please note that the same method was used to determine the purity of canagliflozin in other experiments.

[0404] Table 7. Chemical purity (%) of smegglutinin at different HP-B-CD concentrations

[0405] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. Concluding remarks The results in Table 7 show that the chemical stability of semaglutide, and therefore its purity, depends on the concentration of HP-B-CD. Semaglutide remains chemically stable in all co-formulations (containing 11.25–15% w / v HP-B-CD). However, when the co-formulation contains 15% w / v HP-B-CD, semaglutide exhibits the highest chemical stability and therefore the highest purity.

[0406] Example 5: Effect of different tensioning agents on the physical stability of co-formulations This example demonstrates the stabilizing effect of different tensioning agents on the physical stability of a co-formulation of canagliflozin and smegglutinin that are otherwise identical.

[0407] composition The composition of co-formulations containing different tensile agents is shown in Table 8.

[0408] Table 8 contains the composition of co-formulations with different tensile agents.

[0409] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. Preparation process The formulation was prepared as described in Example 1.

[0410] method All samples were stored under stress conditions, which were defined as follows: Duration: 18 days Temperature: 37℃ • Stress conditions: During storage, the samples are inverted 360° to simulate patient use outside of refrigerated storage. This is performed 100 times five days a week.

[0411] The number of sub-visible particles present quantified the physical stability of the combined canagliflozin and smegglutinin, and was obtained by microfluidic imaging (MFI; for the principles of MFI, see, for example, Sharma, DK et al., AAPS J. (2010), 12: 455-464). The following procedure was used for each syringe sample analyzed: The experiment was conducted at ambient temperature. The plunger was first removed, and then liquid was pipetted into the sample container, thereby removing liquid from each syringe. The sample was transferred to a 96-well plate, which was then inserted into the sample processing unit (Bot1) of a Protein Simple MFI™ 5200 instrument equipped with a standard Protein Simple MFI™ 100 μm flow cell. Sample analysis was performed using the standard MFI system setup, which means that liquid was pipetted into a reservoir connected to the flow cell, the liquid was illuminated with a 10 LED light source (470 nm), and a digital camera (via magnifying optics) recorded the contents of the flow cell as a bright-field image throughout the experiment. Data acquisition was performed using Protein Simple MVSS software. The image stream recorded throughout the run was processed by the validated Novo Nordisk proprietary software MFI Data Validator to obtain the number of individual particles (normalized to a count per ml of analyzed liquid) and presented by size; >5 µm, >10 µm, and >25 µm, which are the standard size ranges for subvisible particles. Note that the number of particles >5 μm includes all particles with a diameter greater than 5 μm (>5 μm, >10 μm, and >25 μm), while the number of particles >10 μm includes all particles with a diameter greater than 10 μm (>10 μm and >25 μm). Particle size was defined as equivalent circle diameter (ECD).

[0412] Table 9. Effects of different tensile agents on the physical stability of co-formulations

[0413] The result is the average of two replicates, rounded to the nearest integer. (-) No sampling performed 1 For co-preparation 10 containing NaCl, sampling was stopped earlier than for other preparations due to the rapid increase in subvisible particle count.

[0414] Concluding remarks The results in Table 9 show that the subvisible particle count increased most rapidly in the co-preparation containing NaCl as a tensile agent (co-preparation 10). After 7 days, the particle count far exceeded that measured in other co-preparations. Therefore, sampling for subvisible particle count analysis of the NaCl-containing co-preparation was discontinued after 7 days.

[0415] After 14 days, an increase in subvisible particle count was observed in the co-formulations containing glycerol and sucrose, and the two co-formulations were considered comparable in terms of physical stability. In the co-formulations containing mannitol, sorbitol, or trehalose, the particle count remained the lowest. In these co-formulations, almost no increase in subvisible particle count was observed during 18 days of storage under stress conditions.

[0416] Among the co-formulations tested, those containing mannitol, sorbitol, or trehalose as tensile agents remained the most stable over time.

[0417] Example 6: Effect of different surfactants on the physical stability of co-formulations This example demonstrates the effect of different surfactants on the physical stability of a co-formulation of canagliflozin and smegglutinin that are otherwise identical.

[0418] composition The composition of co-formulations containing different types of surfactants is shown in Table 10.

[0419] Table 10 contains the composition of co-formulations with different surfactants.

[0420] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. Preparation process The formulation was prepared as described in Example 1.

[0421] method All samples were stored under stress conditions, which were defined as follows: Duration: 17 days Temperature: 37℃ ± 2℃ • Stress conditions: During storage, the samples are inverted 360° to simulate patient use outside of refrigerated storage. This is performed 100 times five days a week.

[0422] The number of subvisible particles was quantified as described in Example 5.

[0423] Table 11 Effects of different surfactants on the physical stability of co-formulations

[0424] The result is the average of two replicates, rounded to the nearest integer. 1 Only one repetition was performed. Concluding remarks Co-formulation 12 contained the fewest subvisible particles after 17 days of storage under stress. In co-formulation 11 containing polysorbate 20, an increase in subvisible particles was observed after 14 days, while in co-formulation 13 containing poloxamer 188, subvisible particles formed after 7 days under stress. Clearly, the co-formulation containing polysorbate 80 was the most stable, and the co-formulation containing polysorbate 20 also exhibited acceptable stability.

[0425] Example 7: Effects of different buffering substances on the physical stability of co-prepared formulations This embodiment demonstrates that buffering substances affect the physical stability of the co-formulation of canagliflozin and smegglutinin, which are otherwise identical.

[0426] composition The compositions of co-formulation 1 and co-formulation 14 are shown in Table 12.

[0427] Table 12 Composition of Co-formulation 1 and Co-formulation 14

[0428] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. Preparation process The formulation was prepared as described in Example 1.

[0429] method All samples were stored under stress conditions, which were defined as follows: Duration: 21 days Temperature: 37℃ ± 2℃ Stress conditions: During storage, the samples were inverted 360° to simulate patient use outside of refrigerated storage. This was repeated 100 times five days a week.

[0430] The number of subvisible particles was quantified as described in Example 5.

[0431] Table 13 Effects of buffering substances on the physical stability of co-formulations

[0432] The result is the average of two replicates, rounded to the nearest integer. Concluding remarks Up to day 14 of storage under stress conditions, the physical stability of the two co-formulations was similar and acceptable. However, after 18 days, the number of sub-visible particles in the citrate-buffered co-formulation (co-formulation 1) was significantly greater than that in the histidine-buffered co-formulation (co-formulation 14). Histidine-buffered co-formulation 14 was the most stable.

[0433] Example 8: Effect of different buffer concentrations on the chemical stability of co-prepared formulations This example demonstrates the effect of buffer concentration on the chemical stability of co-prepared formulations that are otherwise identical.

[0434] composition The composition of co-preparations containing different concentrations of buffer is shown in Table 14.

[0435] Table 14 Composition of co-preparations with different buffer concentrations

[0436] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. Preparation process The formulation was prepared as described in Example 1.

[0437] method The samples were stored at 30°C for 21 days and analyzed after 7, 14, and 21 days to determine the chemical purity of canagliflozin. The purity of canagliflozin was determined as described in Example 4 (for smegglutinin).

[0438] Table 15 Effect of buffer concentration on the chemical stability of canagliflozin in co-formulation

[0439] Concluding remarks The results in Table 15 show that both co-formulations are stable. However, canagliflozin exhibits the highest chemical purity among co-formulations 15. At a histidine concentration of 20 mM, the purity of canagliflozin decreases more rapidly over time.

[0440] Example 9: Effect of different buffer concentrations on the physical stability of co-prepared formulations This example demonstrates the effect of histidine buffer concentration on the physical stability of the co-formulation.

[0441] composition The composition of the tested co-formulations is shown in Table 14.

[0442] Preparation process The formulation was prepared as described in Example 1.

[0443] method All samples were stored under stress conditions, which were defined as follows: Duration: 18 days Temperature: 37℃ ± 2℃ • Stress conditions: During storage, the samples are inverted 360° to simulate patient use outside of refrigerated storage. This is performed 100 times five days a week.

[0444] The number of subvisible particles was quantified as described in Example 5.

[0445] Table 16 Effect of buffer concentration on the physical stability of co-preparation

[0446] The result is the average of two repetitions, rounded to the nearest integer.

[0447] Concluding remarks The difference in physical stability between co-formulations 15 and 16 became most pronounced after 14 days. Data in Table 16 show that the number of sub-visible particles observed in co-formulation 16 (containing 20 mM histidine) was greater than that observed in co-formulation 15 (containing 6 mM histidine). In other words, the co-formulation containing 6 mM histidine was the most physically stable.

[0448] Example 10: Effect of HP-B-CD concentration on subcutaneous tolerance after subcutaneous injection This example demonstrates the concentration-dependent effect of HP-B-CD on subcutaneous tissue after subcutaneous injection.

[0449] composition The compositions of the co-prepared carriers prepared with different HP-B-CD concentrations are shown in Table 17.

[0450] Table 17 Composition of co-formulation carriers containing different concentrations of HP-B-CD

[0451] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. 2 The sorbitol concentration varies with the HP-B-CD concentration to maintain isotonic conditions.

[0452] Preparation process Except for the absence of any active pharmaceutical ingredient, the formulation was prepared as described in Example 1.

[0453] method Local (subcutaneous) tolerability of HP-B-CD-containing formulations following subcutaneous administration was investigated in five live Landrace×Yorkshire×Duroc (LYD) pigs. The method involved assessing skin lesions 6 days after subcutaneous administration of 600 µl using a syringe equipped with a 25G needle and a 5 mm stopper. At necropsy, 2 x 2 cm skin samples were collected, fixed in neutral-buffered formalin, trimmed with multiple scalpels, embedded in paraffin, cut into 4 µm thin sections, mounted on glass slides, and subsequently stained with hematoxylin and eosin (HE). The level of subcutaneous tissue necrosis was assessed using an optical microscope and scored using a numerical scale, where code 1 represented “no necrosis” and code 4 represented “moderate necrosis”. A total of five skin samples were tested for each co-loadant. However, due to variations in the methods of sectioning subcutaneous tissue for successful assessment of necrosis, not all injection sites could be assigned scores. 1. No necrosis 2. Minimal necrosis 3. Mild necrosis 4. Moderate necrosis The level of subcutaneous necrosis following subcutaneous injection was assessed for isotonic co-conjugate formulations containing 10% w / v to 20% w / v HP-B-CD. The results are presented in Table 20.

[0454] Table 18 Necrosis scores of subcutaneous tissue necrosis observed 6 days after injection of co-loaders with different percentages of HP-B-CD.

[0455] Concluding remarks A correlation was observed between increased HP-B-CD concentration in the co-loading agent and necrosis at the injection site. In one example, a co-loading agent containing 20% ​​w / v HP-B-CD resulted in moderate subcutaneous necrosis at the injection site. Formulations containing less than 20% w / v HP-B-CD resulted in only mild or minimal subcutaneous necrosis at the injection site. All co-loading agents containing 10-20% w / v HP-B-CD showed acceptable tolerability, with those containing 10-17.5% w / v HP-B-CD being preferred.

[0456] Example 11: Effects of different tensile agents on subcutaneous tolerance after subcutaneous injection This example demonstrates the effect of any one of three different tensioning agents (sorbitol, mannitol, and trehalose) on local tolerance in otherwise identical isotonic co-conjugation carriers.

[0457] composition The composition of the co-loaders tested is shown in Table 19.

[0458] Table 19 Composition of isotonic co-constitution carriers prepared with different tensile agents

[0459] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. Preparation process Except for the absence of any active pharmaceutical ingredient, the formulation was prepared as described in Example 1.

[0460] method Local tolerance to subcutaneous administration of isotonic loading products containing HP-B-CD and three different tensile agents was investigated in two live Landrace×Yorkshire×Duroc (LYD) pigs to assess skin reactions induced by subcutaneous administration of 600 µl of the loading product. The products were injected using a syringe equipped with a 25G needle and a 5mm stopper. Autopsy was performed approximately 24 hours post-injection. 2x2 cm skin samples were fixed in neutral-buffered formalin, trimmed to 4 µm sections using a multi-blade technique, embedded in paraffin, and subsequently stained with hematoxylin and eosin (HE). For both samples, the severity of subcutaneous tissue necrosis, inflammatory cell infiltration, and hemorrhage distribution was assessed using optical microscopy by a trained toxicological pathologist and scored using a numerical scale, where code 1 represented “no abnormalities” and code 3 represented “mild severity”. 1. No abnormalities 2. Minimum severity 3. Mild to severe Table 20 Severity scores of subcutaneous tissue necrosis, inflammatory cell infiltration, and hemorrhage distribution 24 hours after subcutaneous injection of a co-prescription carrier containing three types of tensile agents. Concluding remarks The data presented in Table 20 show that, overall, sorbitol was the tonic agent that caused the least severity of necrosis, inflammatory cell infiltration, and hemorrhage. These observations confirm that sorbitol is the preferred tonic agent for obtaining good and acceptable subcutaneous tolerability in co-formulations containing active pharmaceutical ingredients.

[0462] Example 12: Confirmation of the effects of the type of tension agent in histidine buffer formulations and citrate buffer formulations on subcutaneous tolerance after subcutaneous injection. This experiment examined: (1) The effect of the type of tonic agent on the local tolerance profile of other similar histidine-buffered co-preparations after subcutaneous injection; and (2) The effect of co-conjugates containing citrate buffers and without tension agents on local tolerance profiles after subcutaneous injection.

[0463] composition The composition of the evaluated co-formulations is described in Tables 21 and 22.

[0464] Table 21 Composition of the isotonic co-prepared histidine buffer

[0465] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. Table 22 Composition of the isotonic co-carrier for citrate buffer

[0466] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. Preparation process Co-formulations 17 and 18 were prepared as described in Example 1. Co-formulation carrier 9 was prepared as described in Example 1, except that no active pharmaceutical ingredient was added.

[0467] method Local tolerance of the co-preparations described in Tables 21 and 22 following subcutaneous administration was investigated in eight live miniature pigs. The resulting skin lesions were assessed 6 days after subcutaneous administration of 750 µl of the sample using a syringe fitted with a 25G needle and a 5 mm stopper (post-mortem). At post-mortem, 2 x 2 cm skin samples were collected, fixed in neutral-buffered formalin, trimmed with multiple scalpels, embedded in paraffin, cut into 4 µm thin sections, mounted on glass slides, and subsequently stained with hematoxylin and eosin (HE). For each sample, the severity of subcutaneous tissue necrosis was assessed using an optical microscope by a trained toxicology pathologist and scored on a numerical scale, where code 1 represented “no abnormalities” and code 5 represented “significant severity”. 1. No abnormalities 2. Minimum severity 3. Mild to severe 4. Moderate severity 5. Significant severity The necrosis score results are shown in Tables 23 and 24.

[0468] Table 23 Severity scores of subcutaneous tissue necrosis 6 days after injection of histidine-buffered co-preparation.

[0469] Table 24 Severity score of subcutaneous tissue necrosis 6 days after injection of citrate-buffered co-loader

[0470] Concluding remarks The results presented in Table 23 show that the type of tonic agent contained in the formulation affects its local tolerability in vivo. A correlation exists between the tonic agent and subcutaneous necrosis observed at the injection site. Subcutaneous injection of co-formulation 17 containing trehalose resulted in two mild necrosis events (score 3). Subcutaneous injection of co-formulation 18 containing sorbitol resulted in only the lowest degree of necrosis (score 2), which is a better result. These results confirm that, in the case of otherwise identical co-formulation carriers, co-formulations containing 15% w / v HP-B-CD (mean MS: 0.62) and sorbitol are superior to co-formulations containing 15% w / v HP-B-CD (mean MS: 0.62) and trehalose.

[0471] The results presented in Table 24 show that three significant necrosis events (score of 5) were observed using a co-formulation containing 25% w / v HP-B-CD (mean MS: 0.92), citrate, and no tensioning agent, confirming that this particular co-formulation is not suitable for subcutaneous administration.

[0472] Example 13: Effects of different types of hydroxypropyl-substituted cyclodextrins on the physical and chemical stability of canagliflozin and smegglutinin co-formulation This example demonstrates the effects of hydroxypropyl-α-cyclodextrin (HP-A-CD), hydroxypropyl-β-cyclodextrin (HP-B-CD), and hydroxypropyl-γ-cyclodextrin (HP-G-CD) on the formation of subvisible particles and the chemical degradation of canagliflozin in a co-formulation of canagliflozin and smegglutinin that are otherwise identical.

[0473] composition The compositions of co-formulations 20, 21 and 23 are shown in Table 25.

[0474] Table 25 Composition of co-preparations with different types of hydroxypropyl cyclodextrins

[0475] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. Preparation process The formulation was prepared as described in Example 1.

[0476] method The sample used to determine the subvisible particle count is stored under stress conditions, which are defined as follows: Duration: 42 days Temperature: 30℃ ± 2℃ - Stress conditions: During storage, the sample was inverted 360° to simulate patient use outside of refrigerated storage. 20 rotations were performed three days a week, and 40 rotations were performed two days a week.

[0477] The number of subvisible particles was determined as described in Example 5.

[0478] Samples used for determining canagliflozin purity were stored at 37°C for up to 42 days. Canagliflozin purity was determined using the following reversed-phase high-performance liquid chromatography (RP-HPLC) method, employing a Kinetex C18 2.6 μm column (4.6 x 150 mm). Sequencing was performed using eluent A and eluent B, consisting of 90% v / v 0.09 M phosphate solution (pH 3.6) and 10% v / v acetonitrile, and eluent B consisting of 60% v / v acetonitrile and 20% v / v isopropanol. Chromatographic analysis was performed at 30°C using an injection volume of 10–100 μl and a flow rate of 0.7 ml / min, with UV detection (210 nm). Canagliflozin purity was quantified by dividing the area of ​​the main peak by the area of ​​all relevant peaks multiplied by 100%.

[0479] The purity of smegglutinin was determined using the same method in other experiments.

[0480] Table 26 Physical stability of canagliflozin and smegglutinin co-formulations prepared with different types of hydroxypropyl cyclodextrin

[0481] The subvisible particle number result is the average of three replicates, rounded to the nearest integer. (-) No sampling performed 1 For co-formulation 22 containing HP-G-CD, sampling was stopped earlier than for other formulations due to the rapid increase in particle count.

[0482] Table 27 Chemical purity (%) of canagliflozin in co-formulations of canagliflozin and smegglutinin prepared with different types of hydroxypropyl cyclodextrin

[0483] Concluding remarks The results presented in Table 26 show that a large number of subvisible particles were observed at time zero in co-formulation 22 (HP-G-CD), which prevented the use of HP-G-CD for co-formulation of canagliflozin and smegglutinin. For co-formulation 22 containing HP-G-CD, sampling for analyzing subvisible particle counts was stopped after the initial analysis at time zero. For co-formulation 20 (HP-A-CD) and co-formulation 21 (HP-B-CD), almost no increase in the number of subvisible particles was observed.

[0484] The results presented in Table 27 regarding the chemical purity of canagliflozin in the presence of HP-A-CD or HP-B-CD show that the purity of canagliflozin in co-formulation 20 containing HP-A-CD decreased at a slightly faster rate than that in co-formulation 21 containing HP-B-CD.

[0485] Based on the results in Table 26, HP-A-CD or HP-B-CD is acceptable for co-formulations of canagliflozin and smegglutinin. However, based on the results in Table 27, HP-B-CD is preferred over HP-A-CD for co-formulations of canagliflozin and smegglutinin because canagliflozin has better purity when formulated with HP-B-CD.

[0486] Example 14: Effect of β-cyclodextrin substitution type on the physical stability of co-formulations This example demonstrates the effect of sulfobutyl ether-β-cyclodextrin (SBE-B-CD) and hydroxypropyl-β-cyclodextrin on the physical stability of a co-formulation of canagliflozin and smegglutinin that are otherwise identical.

[0487] composition The composition of co-formulations containing HP-B-CD or SBE-B-CD is shown in Table 28.

[0488] Table 28 Composition of co-preparations containing HP-B-CD or SBE-B-CD

[0489] 1 MS: Molar degree of substitution, corresponding to sulfobutyl ether / hydroxypropyl per glucose unit. Preparation process The formulation was prepared as described in Example 1.

[0490] method The sample used to determine the number of subvisible particles was stored under stress conditions, which were defined as follows: Duration: 35 days Temperature: 30℃ ± 2℃ - Stress conditions: During storage, the sample was inverted 360° to simulate patient use outside of refrigerated storage. 20 rotations were performed three days a week, and 40 rotations were performed two days a week.

[0491] The number of subvisible particles was quantified as described in Example 5.

[0492] Table 29. Levels of subvisible particles in co-formulations of canagliflozin and smegglutinin containing HP-B-CD or SBE-B-CD.

[0493] The subvisible particle number result is the average of three replicates, rounded to the nearest integer. Concluding remarks The results in Table 29 show that when canagliflozin and smegglutinin were co-formulated using SBE-B-CD, a significant increase in the number of subvisible particles was observed after 14 days; that is, the co-formulation is physically unstable. In contrast, when HP-B-CD was used, almost no increase was observed during the 35-day study period; that is, the co-formulation is physically stable.

[0494] These results demonstrate, compared to our results for hydroxypropyl-β-cyclodextrin, that sulfobutyl ether-β-cyclodextrin (SBE-B-CD) is not a suitable cyclodextrin for co-formulation of canagliflozin and smegglutinin.

[0495] Example 15: Effect of the concentration ratio of canagliflozin and smegglutinin on the physical stability of the co-formulation This example demonstrates the effect of different concentration ratios of canagliflozin and smegglutinin on the level of subvisible particles observed in the co-formulation.

[0496] composition The compositions of histidine buffer co-formulations 25 to 29 are shown in Table 30, while the compositions of histidine buffer co-formulations 30 to 40 are shown in Table 31.

[0497] Table 30 Composition of histidine buffer co-formulations with different canagliflozin and smegglutinin concentration ratios

[0498] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. Table 31 Composition of co-formulations with different canagliflozin and smegglutinin concentration ratios and modified histidine buffers

[0499] 1MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. Preparation process The formulation was prepared as described in Example 1.

[0500] method The sample used to determine the number of subvisible particles was stored under stress conditions, which were defined as follows: Duration: 28 days Temperature: 30℃ ± 2℃ - Stress conditions: During storage, the sample was inverted 360° to simulate patient use outside of refrigerated storage. 20 rotations were performed three days a week, and 40 rotations were performed two days a week.

[0501] The number of subvisible particles was quantified as described in Example 5.

[0502] Table 32 shows the levels of subvisible particles in co-formulations containing different concentration ratios of canagliflozin and smegglutinin.

[0503] The subvisible particle number result is the average of three replicates, rounded to the nearest integer. Concluding remarks The results presented in Table 32 show that, after 21 days, almost no increase in subvisible particle count was observed in co-formulations 25-39 containing 3.2 mg / ml canagliflozin and up to 12 mg / ml smegglutinin.

[0504] After 14 days, an increase in subvisible particle count was observed in the co-formulation containing 3.2 mg / ml canagliflozin and 16 mg / ml smegglutinin.

[0505] Co-formulations containing 3.2 mg / ml canagliflozin and up to 16 mg / ml smegglutinin with histidine buffers 25 to 40 are physically stable.

[0506] Example 16: Effect of HP-B-CD concentration on the physical stability of the co-formulation This example demonstrates the effect of HP-B-CD concentration on the physical stability of the canagliflozin and smegglutinin co-formulation when the co-formulation is exposed to physical stress.

[0507] composition The compositions of co-formulations 41 to 44 containing histidine buffers are shown in Table 32.

[0508] Table 32 Composition of co-formulations with different HP-B-CD concentrations

[0509] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. Preparation process The formulation was prepared as described in Example 1.

[0510] method The tendency of canagliflozin and smegglutinin in the co-formulation to aggregate and form peptide fibrils was measured using the thioflavin T (ThT) fluorescence stress assay described in Example 2.

[0511] Table 33 Physical stability of canagliflozin and smegglutinin co-formulations with different HP-B-CD concentrations

[0512] 1 The result is the average of 6 replicates. 2 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. Concluding remarks The results presented in Table 33 show that the physical stability of the canagliflozin and smegglutinin co-formulation depends on the concentration of HP-B-CD; lower concentrations result in shorter hysteresis times before fibrillation. The co-formulation containing 7.5% w / v HP-B-CD is the least stable, while the co-formulation containing 15% w / v HP-B-CD is the most stable.

[0513] Example 17: Local tolerance of pigs to subcutaneously injected carrier formulations with different HP-B-CD contents, molar substitution degrees, and total buffer compositions. This experiment examined: (1) Effects of HP-B-CD concentration and mean MS (0.62 vs. 0.92) on local tolerance profile after subcutaneous injection.

[0514] (2) The effect of the formulation carrier on the local tolerance spectrum after subcutaneous injection in the presence of HP-B-CD.

[0515] composition The composition of the co-loaders tested is shown in Table 34.

[0516] Table 34 Composition of co-loadors with different HP-B-CD contents, molar substitution degrees, and total buffer compositions.

[0517] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. Preparation process Except for the absence of any active pharmaceutical ingredient, the formulation was prepared as described in Example 1.

[0518] method Local (subcutaneous) tolerability of a formulation containing HP-B-CD following subcutaneous administration was investigated in four live Landrace×Yorkshire×Duroc (LYD) pigs. The method involved assessing the resulting skin lesions five days after subcutaneous administration of 200 µl of NovoPen 4 with a NovoFine Plus needle (32 G / 4 mm) at necropsy. At necropsy, 2 x 2 cm skin samples were collected, fixed in neutral-buffered formalin, trimmed with multiple scalpels, embedded in paraffin, cut into 4 µm thin sections, mounted on glass slides, and subsequently stained with hematoxylin and eosin (HE). For all four samples, the severity of subcutaneous tissue necrosis and inflammatory cell infiltration was assessed using optical microscopy by a trained toxicology pathologist and scored on a numerical scale, where code 1 represented “no abnormalities” and code 5 represented “significant severity”. 1. No abnormalities detected. 2. Minimum severity 3. Mild to severe 4. Moderate severity 5. Significant severity The levels of subcutaneous tissue necrosis and inflammatory cell infiltration induced by the co-loader after subcutaneous injection were assessed. The results are presented in Table 35.

[0519] Table 35 Severity Scoring: Severity of subcutaneous tissue necrosis and inflammatory cell infiltration 5 days post-injection for co-loaders with different HP-B-CD content, molar substitution degree, and total buffer composition.

[0520] Concluding remarks The results in Table 35 show that in vivo local subcutaneous tolerability depends on the concentration of HP-B-CD and the total buffer composition. Co-loadors containing histidine and sorbitol showed better tolerability than co-loadors containing citrate.

[0521] Co-formulations containing 20% ​​w / v or less HP-B-CD did not cause, or mainly caused, minimal necrosis or inflammatory cell infiltration (score 1 or 2), with one instance of mild inflammatory cell infiltration observed (score 3). Co-formulations containing 22% w / v or more HP-B-CD all caused minimal to moderate necrosis and inflammatory cell infiltration (score up to 4). Based on these results, co-formulations containing less than 22% HP-B-CD appear suitable for subcutaneous application.

[0522] Co-loadors containing 20% ​​w / v and 22% w / v HP-B-CD and citrate (co-loadors 15 and 16) resulted in significant necrosis and inflammatory cell infiltration (scores up to 5). Surprisingly, co-loadors containing 20% ​​w / v and 22% w / v HP-B-CD, histidine, and sorbitol (co-loadors 11 and 12) were better tolerable, resulting in moderate necrosis and inflammatory cell infiltration (scores up to 4).

[0523] The co-loaded formulation containing m-cresol, phenol, and EDTA, as well as 15% w / v or 20% w / v HP-B-CD, was well tolerated, resulting in mild necrosis and inflammatory cell infiltration (scores up to 3).

[0524] Example 18: Effects of different pharmaceutical preservatives on the antiseptic efficacy of canagliflozin and smegglutinin co-preparation against Staphylococcus aureus This example demonstrates the effects of different antimicrobial preservatives in the co-preparation of canagliflozin and smegglutinin on Staphylococcus aureus after 24 hours. Staphylococcus aureus The impact on growth.

[0525] composition The compositions of co-formulations 45 to 56 are shown in Table 36.

[0526] Table 36 Composition of co-preservatives with different types of preservatives

[0527] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. preparation The co-formulation was prepared as follows: a solution of excipients containing the active pharmaceutical ingredient was prepared, and then adjusted to achieve the final pH and volume. The co-formulation was filtered to remove bacteria and then filled into 3 ml cartridges.

[0528] method The preservative efficacy test was conducted in accordance with the European Pharmacopoeia (5.1.3), except that only Staphylococcus aureus was tested, and the preservative efficacy was calculated only after 24 hours based on the logarithmic decrease in the number of viable microorganisms compared to the zero-point value obtained from the inoculum.

[0529] Table 37 Effects of different preservatives on the logarithmic reduction after 24 hours

[0530] Concluding remarks The highest reduction in Staphylococcus aureus growth was achieved after 24 hours in co-formulations 49, 50, and 51, which contained potassium metabisulfite or sodium sulfite. Co-formulation 48, containing m-cresol and phenol, showed a low logarithmic reduction, while the remaining co-formulations did not show any ability to inhibit Staphylococcus aureus growth after 24 hours.

[0531] Example 19: Effects of sodium sulfite and potassium metabisulfite on the chemical stability of the co-formulation of canagliflozin and smegglutinin The following examples demonstrate the effect of sodium sulfite and potassium metabisulfite, two preservatives, on the chemical stability of canagliflozin and smegglutinin in co-formulations that are otherwise identical, with respect to the chemical purity.

[0532] composition The compositions of co-formulations 57 to 63 are shown in Table 38.

[0533] Table 38 Composition of co-preparations containing sodium sulfite or potassium metabisulfite, 57 to 63

[0534] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. preparation The co-formulation was prepared as described in Example 18.

[0535] method Samples used to determine the purity of canagliflozin and smegglutinin were stored at 37°C for up to 21 days.

[0536] The purity of canagliflozin was determined using the reversed-phase high-performance liquid chromatography (RP-HPLC) method described in Example 1.

[0537] The purity of smegglutinin was determined using the reversed-phase high-performance liquid chromatography (RP-HPLC) method described in Example 3.

[0538] Table 39 Chemical purity (%) of canagliflozin in co-preparations containing sodium sulfite or potassium metabisulfite

[0539] Table 40 Chemical purity (%) of smegglutide in co-preparations containing sodium sulfite or potassium metabisulfite

[0540] Concluding remarks The results presented in Tables 39 and 40 show that, compared to the preservative-free co-formulation 57, the purity of canagliflozin and smegglutinin decreased rapidly when sodium sulfite or potassium metabisulfite was added to the co-formulation. The higher the concentration of the preservative, the faster the purity loss.

[0541] Example 20: Effect of different pharmaceutical preservatives on the preservative efficacy of canagliflozin and smegglutinin co-preparation after 7 days This example demonstrates the effects of different antimicrobial preservatives in the co-preparation of canagliflozin and smegglutinin on Staphylococcus aureus and Candida albicans after 7 days. Candida albicans ) and Aspergillus brasiliensis ( Aspergillus brasiliensis The impact on microbial growth.

[0542] composition The compositions of co-formulations 45 to 48 and 52 to 56 are as shown in Example 18.

[0543] preparation The co-formulation was prepared as described in Example 18.

[0544] method The preservative efficacy test was conducted in accordance with the United States Pharmacopeia (Chapter 51), except that only three microorganisms were tested: Staphylococcus aureus, Candida albicans, and Aspergillus brasiliensis, and the preservative efficacy was calculated only after 7 days based on the logarithmic reduction in the number of viable microorganisms compared to the zero-point value obtained from the inoculum.

[0545] Table 41 Effects of different preservatives on the logarithmic reduction after 7 days

[0546] Concluding remarks The results presented in Table 7 show that for Staphylococcus aureus, co-formulations 47, 52, and 53, containing m-cresol and phenol or benzalkonium chloride, achieved the highest log reduction. For Candida albicans, no growth was observed in co-formulations 46 and 53, which contained EDTA. For Aspergillus brasiliensis, no differences were observed among the tested co-formulations.

[0547] Example 21: Effects of different concentrations of benzalkonium chloride and EDTA on the preservative efficacy of canagliflozin and smegglutinin co-preparation after 7 days This example demonstrates the effect of a co-formulation of canagliflozin and smegglutinin containing different concentrations of benzalkonium chloride and EDTA disodium dihydrate on the growth of Staphylococcus aureus, Candida albicans, and Aspergillus brasiliensis after 7 days.

[0548] composition The compositions of co-formulations 64 to 67 are shown in Table 42.

[0549] Table 42 Composition of co-preparations containing different concentrations of benzalkonium chloride and EDTA

[0550] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. preparation The co-formulation was prepared as described in Example 18.

[0551] method The preservative efficacy of the co-preservative formulation was tested against three microorganisms: Staphylococcus aureus, Candida albicans, and Aspergillus brasiliensis. The preservative efficacy was calculated based on the logarithmic decrease in the number of viable microorganisms 7 days after the time zero value was obtained from the inoculum.

[0552] Table 43 Effects of different preservatives on the logarithmic reduction after 7 days

[0553] Concluding remarks The results presented in Table 43 show that higher concentrations of benzalkonium chloride resulted in a greater reduction in the logarithmic number of Staphylococcus aureus. Increasing the concentration of EDTA led to a greater reduction in the logarithmic number of Candida albicans, while no difference was observed among the different co-preparations for Aspergillus brasiliensis.

[0554] Example 22: Effects of different concentrations of benzalkonium chloride and EDTA on the chemical stability of the co-formulation of canagliflozin and smegglutinin The following examples demonstrate the effect of different concentrations of benzalkonium chloride and EDTA on the chemical stability of canagliflozin and smegglutinin in terms of their chemical purity.

[0555] composition The compositions of co-formulations 64 to 68 are shown in Table 44.

[0556] Table 44 Composition of co-preparations containing benzalkonium chloride and EDTA 64 to 68

[0557] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. preparation The co-formulation was prepared as described in Example 18.

[0558] method Samples used to determine the purity of canagliflozin and smegglutinin were stored at 37°C for 6 weeks.

[0559] The purity of canagliflozin and smegglutinin was determined using reversed-phase high-performance liquid chromatography (RP-HPLC) as described in Example 13.

[0560] Table 45 Chemical purity (%) of canagliflozin in co-preparations containing different concentrations of benzalkonium chloride and EDTA.

[0561] Table 46 Chemical purity (%) of smegglutide in co-preparations containing different concentrations of benzalkonium chloride and EDTA.

[0562] Concluding remarks The results presented in Table 45 show that the chemical purity of canagliflozin in co-formulations 64 to 67, which contain different concentrations of benzalkonium chloride and EDTA, was significantly lower than that in co-formulation 68, which does not contain preservatives. The results in Table 46 show a slight positive effect on the chemical purity of smegglutinin.

[0563] Example 23: Effect of different tension agents on the preservative efficacy of the co-preservative formulation of canagliflozin and smegglutinin after 28 days This example demonstrates the effectiveness of comprehensive antiseptic efficacy testing (European Pharmacopoeia 5.1.3, USP). <51> In the test, the effects of two different tensioning agents, sorbitol or propylene glycol, together with m-cresol, phenol and EDTA, on the preservative efficacy of the co-formulation of canagliflozin and smegglutinin were investigated.

[0564] composition The compositions of co-formulations 69 and 70 are shown in Table 47.

[0565] Table 47 Composition of co-preparations containing m-cresol, phenol, EDTA, and sorbitol or propylene glycol

[0566] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. preparation The co-formulation was prepared as described in Example 18.

[0567] method According to USP ( <51> The co-preservative efficacy of the formulation was tested against the following microorganisms in the comprehensive antiseptic efficacy tests described in the European Pharmacopoeia (5.1.3.): Staphylococcus aureus, Pseudomonas aeruginosa (…). Pseudomonas aeruginosa *Escherichia coli*, *Candida albicans*, and *Aspergillus brasiliensis* were included. Preservative efficacy was calculated based on the logarithmic decrease in viable microbial count compared to the zero-point value obtained from the inoculum.

[0568] Table 48 Effects of m-cresol, phenol, and EDTA on the preservative efficacy of different tensile agents

[0569] Concluding remarks Overall, no difference in preservative efficacy was observed between co-formulations 69 and 70 containing sorbitol or propylene glycol as a tensile agent.

[0570] Example 24: Effect of different toning agents on the chemical stability of preservative-preserved canagliflozin and smegglutinin co-formulation This example demonstrates the effect of two different tensor agents, m-cresol, phenol, EDTA, and sorbitol or propylene glycol, on the chemical stability of a canagliflozin and smegglutinin co-formulation, as measured by the purity of canagliflozin and smegglutinin.

[0571] composition The compositions of co-formulations 69 and 70 are shown in Example 23.

[0572] preparation The co-formulation was prepared as described in Example 18.

[0573] method Samples used to determine the purity of canagliflozin and smegglutinin were stored at 37°C for 4 weeks.

[0574] The purity of canagliflozin and smegglutinin was determined using reversed-phase high-performance liquid chromatography (RP-HPLC) as described in Example 13.

[0575] Table 49 Chemical purity (%) of canagliflozin in co-formulations containing sorbitol or propylene glycol as a tensile agent.

[0576] Table 50 Chemical purity (%) of semaglutide in co-formulations containing sorbitol or propylene glycol as a tensile agent.

[0577] Concluding remarks The results presented in Tables 49 and 50 show that no difference in chemical purity was observed between canagliflozin and smegglutinin in co-preparations containing sorbitol or propylene glycol as a tensile agent and m-cresol, phenol and EDTA as preservatives.

[0578] Example 25: Effect of polysorbate 80 on the physical stability of a co-formulation of canagliflozin and smegglutinin preservatives of m-cresol, phenol and EDTA. This example demonstrates the effect of polysorbate 80 on the physical stability of a co-formulation of canagliflozin and smegglutinin with the same preservatives (m-cresol, phenol, and EDTA) in terms of subvisible particle count.

[0579] composition The compositions of co-formulations 71 to 72 are shown in Table 51.

[0580] Table 51 Composition of co-formulations containing and without polysorbate 80

[0581] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. preparation The co-formulation was prepared as described in Example 18.

[0582] method The sample used to determine the number of subvisible particles was stored under stress conditions, which were defined as follows: Duration: 28 days Temperature: 30℃ ± 2℃ - Stress conditions: During storage, the sample was inverted 360° to simulate patient use outside of refrigerated storage. 20 rotations were performed three days a week, and 40 rotations were performed two days a week.

[0583] The number of subvisible particles was quantified as described in Example 5.

[0584] Table 52 Levels of subvisible particles in co-formulations

[0585] The subvisible particle number result is the average of three replicates, rounded to the nearest integer. Concluding remarks The data in Table 52 show that the number of subvisible particles observed in co-formulation 71, which does not contain polysorbate 80, is greater than the number observed in co-formulation 72, which contains polysorbate 80. In other words, the co-formulation containing polysorbate 80 is the most physically stable.

[0586] Example 26: Effect of the molar substitution degree of HP-B-CD on the preservative efficacy of a co-preservative formulation of canagliflozin and smegglutinin with m-cresol, phenol, and EDTA as preservatives. This example demonstrates the effectiveness of comprehensive antiseptic efficacy testing (European Pharmacopoeia 5.1.3, USP). <51> The effect of HP-B-CD molar substitution degree on the preservative efficacy of canagliflozin and smegglutinin co-preservatives with m-cresol, phenol, and EDTA as preservatives was tested in the study.

[0587] composition The compositions of co-formulations 73 to 74 are shown in Table 53.

[0588] Table 53 Composition of co-formulations containing HP-B-CD with different molar degrees of hydroxypropyl substitution and preservatives of m-cresol, phenol, and EDTA.

[0589] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. preparation The co-formulation was prepared as described in Example 18.

[0590] method According to USP ( <51> The preservative efficacy of the co-preservative formulation was tested against the following microorganisms in the comprehensive preservative efficacy tests described in the European Pharmacopoeia (5.1.3.): Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli, Candida albicans, and Aspergillus brasiliensis. Preservative efficacy was calculated based on the logarithmic decrease in the viable microbial count compared to the zero-point value obtained from the inoculum.

[0591] Table 54 Effect of the molar substitution degree of HP-B-CD on the preservative efficacy of co-preservatives using m-cresol, phenol, and EDTA.

[0592] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. Concluding remarks Compared to co-formulation 73 (mean MS: 0.62), co-formulation 74 (mean MS: 0.92) showed an increased result in Table 54, as indicated by the logarithmic reduction in Staphylococcus aureus, Escherichia coli, and Aspergillus brasiliensis, while a greater logarithmic reduction in Candida albicans was observed for co-formulation 73 (mean MS: 0.62). No difference was observed between the two co-formulations for Pseudomonas aeruginosa.

[0593] Example 27: Effect of the molar substitution degree of HP-B-CD on the physical stability of the preservative-preserved canagliflozin and smegglutinin co-formulation This example demonstrates the effect of HP-B-CD molar substitution degree on the physical stability of otherwise identical preservative-preserved canagliflozin and smegglutinin co-formulations, with respect to subvisible particle count.

[0594] composition The compositions of co-formulations 73 to 74 are as shown in Example 26.

[0595] preparation The co-formulation was prepared as described in Example 18.

[0596] method The sample used to determine the number of subvisible particles was stored under stress conditions, which were defined as follows: Duration: 35 days Temperature: 30℃ ± 2℃ - Stress conditions: During storage, the sample was inverted 360° to simulate patient use outside of refrigerated storage. 20 rotations were performed three days a week, and 40 rotations were performed two days a week.

[0597] The number of subvisible particles was quantified as described in Example 5.

[0598] Table 55 Levels of subvisible particles in co-formulations

[0599] The subvisible particle number result is the average of three replicates, rounded to the nearest integer. Concluding remarks The results presented in Table 55 show that, compared with co-formulation 73 (mean MS: 0.62), the overall level of subvisible particles was lower in co-formulation 74 (mean MS: 0.92) after 35 days. In these otherwise identical semaglutide and canagliflozin co-formulations, co-formulation 74, containing HP-B-CD (mean MS: 0.92), was physically more stable than co-formulation 73, which also contained HP-B-CD (mean MS: 0.62).

[0600] Example 28: Effects of m-cresol, EDTA, and HP-B-CD concentrations on the preservative efficacy of a preservative-treated canagliflozin and smegglutinin co-preservative formulation This example demonstrates the effect of different concentrations of m-cresol, EDTA, and HP-B-CD on the preservative efficacy of a co-preserved canagliflozin and smegglutinin formulation during preservative efficacy testing.

[0601] composition The compositions of co-formulations 75 to 78 are shown in Table 56.

[0602] Table 56 Composition of co-formulations containing different concentrations of m-cresol, EDTA, and HP-B-CD

[0603] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. preparation The co-formulation was prepared as described in Example 18.

[0604] method The preservative efficacy of the co-preservative formulation was tested against three microorganisms: Staphylococcus aureus, Candida albicans, and Aspergillus brasiliensis. The preservative efficacy was calculated based on the logarithmic decrease in the number of viable microorganisms 7 days after the time zero value was obtained from the inoculum.

[0605] The methods used to test the preservative efficacy of the four co-preparations were performed in accordance with the United States Pharmacopeia (Chapter 51) and the European Pharmacopoeia (5.1.3), except that the preservative efficacy was tested only for the microorganism Candida albicans, and the preservative efficacy was calculated only after 7 days based on the logarithmic decrease in the number of viable microorganisms compared to the time zero value obtained from the inoculum.

[0606] Table 57 Effects of different concentrations of m-cresol, EDTA, and HP-B-CD on the preservative efficacy of Candida albicans.

[0607] Concluding remarks The results presented in Table 57 show no differences among co-formulations 75, 76, and 77, which contain different concentrations of HP-B-CD and m-cresol. For co-formulation 78, which contains a higher concentration of EDTA, a greater logarithmic reduction in *Candida albicans* was observed.

[0608] Example 29: Effect of the molar substitution degree of HP-B-CD on the physical stability of the preservative-preserved canagliflozin and smegglutinin co-formulation This example demonstrates the effect of HP-B-CD molar substitution degree on the physical stability of otherwise identical preservative-preserved canagliflozin and smegglutinin co-formulations, with respect to subvisible particle count.

[0609] composition The compositions of co-formulations 79 to 84 are shown in Table 58.

[0610] Table 58 contains the composition of co-formulations of HP-B-CD with different average molar degrees of substitution.

[0611] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. preparation The co-formulation was prepared as follows: An excipient solution containing HP-B-CD, histidine, and sorbitol was prepared, with the active pharmaceutical ingredient added. Then, polysorbate 80 and a preservative were added, followed by adjustments to achieve the final pH and volume. The co-formulation was filtered sterilized and filled into 3 ml cartridges.

[0612] method The sample used to determine the number of subvisible particles was stored under stress conditions, which were defined as follows: Duration: 28 days Temperature: 30℃ ± 2℃ - Stress conditions: During storage, the sample was inverted 360° to simulate patient use outside of refrigerated storage. 20 rotations were performed three days a week, and 40 rotations were performed two days a week.

[0613] The number of subvisible particles was quantified as described in Example 5.

[0614] Table 59 Levels of subvisible particles in co-formulations

[0615] The subvisible particle number result is the average of three replicates, rounded to the nearest integer. 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. Concluding remarks The results presented in Table 59 show that, in co-formulations of smegliflozin and canagliflozin that are otherwise identical, the number of subvisible particles counted after 28 days was unacceptably high in co-formulation 84 (containing HP-B-CD with an average molar substitution degree of 1.08) compared to other co-formulations with lower average molar substitution degrees. Co-formulations containing HP-B-CD with average molar substitution degrees of 0.62, 0.67, 0.68, 0.84, and 0.92 were physically stable.

[0616] Example 30: Effect of the molar substitution degree of HP-B-CD on the chemical stability of a preservative-preserved co-formulation of canagliflozin and smegglutinin This example demonstrates the effect of HP-B-CD molar substitution degree on the chemical stability, as measured by smegglutinin purity, of a preservative-preserved co-formulation of canagliflozin and smegglutinin that are otherwise identical.

[0617] composition The compositions of co-formulations 79 to 84 are as shown in Example 29.

[0618] preparation The co-formulation was prepared as described in Example 29.

[0619] method Samples used to determine the purity of canagliflozin and smegglutinin were stored at 37°C for 4 weeks.

[0620] The purity loss of smegglutinin was determined using the reversed-phase high-performance liquid chromatography (RP-HPLC) method described in Example 13.

[0621] Table 60 Chemical purity (%) of smegglutide in co-formulations containing HP-B-CD with different molar degrees of substitution

[0622] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. Concluding remarks The results presented in Table 60 show that the chemical purity of semaglutide is affected by the average molar degree of substitution of HP-B-CD. Among co-formulation 84 (average MS: 1.08), semaglutide showed the greatest loss of chemical purity. Co-formulations containing HP-B-CD with average molar degrees of substitution of 0.62, 0.67, 0.68, 0.84, or 0.92 were all chemically stable.

[0623] Example 31: Effect of pH on the physical stability of a preservative-preserved co-formulation of canagliflozin and smegglutinin This example demonstrates the effect of pH on the physical stability of otherwise identical, preservative-preserved co-formulations of canagliflozin and / or smegglutinin, with respect to their tendency to aggregate and form peptide fibrils.

[0624] composition The compositions of co-formulations 85 to 89 are shown in Table 61.

[0625] Table 61 Composition of co-preparations with different pH values

[0626] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. preparation The co-formulation was prepared as described in Example 29.

[0627] method The tendency of canagliflozin and / or smegglutinin to aggregate and form peptide fibrils—a parameter used to quantify physical stability—was quantified as described in Example 2.

[0628] Table 62 Physical stability of canagliflozin and smegglutinin co-formulations with different pH values

[0629] 1 The result is the average of 6 replicates. Concluding remarks The co-preparations in Table 62 were subjected to induced stress conditions, and the tendency of canagliflozin and / or smegglutinin to form peptide fibrils was quantified by measuring the time from the start of the assay until fibril formation occurred (“hysteresis time”). Longer hysteresis times indicated greater stability of the co-preparations. The results showed that co-preparation 87 at pH 6 had the longest hysteresis time before fibril formation and was therefore the most stable co-preparation. Co-preparations 86 (pH 5.8) and 88 (pH 6.2) were found to be quite stable. Of the co-preparations tested, co-preparation 85 (pH 5.6) had the shortest hysteresis time before fibril formation, meaning it was the least stable in terms of physical stability.

[0630] Example 32: Effect of HP-B-CD concentration on the physical stability of preservative-preserved canagliflozin and smegglutinin co-formulation This example demonstrates the effect of HP-B-CD concentration on the physical stability of a preservative-preserved canagliflozin and smegglutinin co-formulation, with respect to subvisible particle count.

[0631] composition The compositions of co-formulations 90 to 94 are shown in Table 64.

[0632] Table 64 Composition of co-preparations containing different concentrations of HP-B-CD

[0633] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. preparation The co-formulation was prepared as described in Example 29.

[0634] 1 MS: Molar degree of substitution, corresponding to the hydroxypropyl group per glucose unit. 2 Co-formulations 90 (0% w / v) and 91 (5% w / v) could not be prepared because the solution was not clear and precipitation was observed, resulting in no data being available for the affected batches.

[0635] 3 The result is the average of 6 replicates. Concluding remarks The results presented in Table 65 show that co-formulations of canagliflozin and smegglutinide containing less than 10% w / v HP-B-CD (mean MS: 0.92) could not be prepared due to precipitation. All co-formulations containing more than 10% w / v were considered physically stable. The longest pre-fibrillation hysteresis time was obtained in co-formulation 94 containing 15% w / v HP-B-CD, while the pre-fibrillation hysteresis time was longer in co-formulation 93 containing 12.5% ​​w / v than in co-formulation 92 containing 10% w / v. In other words, co-formulations of canagliflozin and smegglutinide containing higher levels of HP-B-CD (mean MS: 0.92) exhibit better physical stability.

[0636] While certain features of the invention have been set forth and described herein, many modifications, substitutions, alterations, and equivalents will now occur to those skilled in the art. Therefore, it should be understood that all such modifications and alterations falling within the true scope of the invention are intended to be covered by the appended claims.

Claims

1. A pharmaceutical formulation comprising an amylin receptor agonist, a GLP-1 receptor agonist, a hydroxypropyl-substituted α- and / or β-cyclodextrin, and at least one preservative; wherein the theoretically calculated isoelectric point of the GLP-1 receptor agonist is equal to or less than about 4.5, for example, about 4.0-4.5; and wherein the cyclodextrin comprises 0.58-1.0 hydroxypropyl groups per glucose unit.

2. The pharmaceutical formulation according to claim 1, wherein the theoretically calculated isoelectric point (pI) of the amylin receptor agonist is approximately 7.6-9.4 or 8-9.

3. A pharmaceutical formulation comprising canagliflozin, smegglutinin, hydroxypropyl-substituted α- and / or β-cyclodextrin, and at least one preservative, said cyclodextrin comprising 0.58-1.0 hydroxypropyl groups per glucose unit.

4. The pharmaceutical formulation according to any one of claims 1-3, wherein the cyclodextrin is a hydroxypropyl-substituted β-cyclodextrin.

5. The pharmaceutical formulation according to any one of claims 1-4, wherein the cyclodextrin has an average molar degree of substitution (MS) of about 0.92 hydroxypropyl groups per glucose unit.

6. The pharmaceutical formulation according to any one of claims 1-5, further comprising a buffer having at least one pKa of about 5.0-7.0, such as citrate and / or histidine.

7. The pharmaceutical formulation according to any one of claims 1-6, further comprising a tensile agent, provided that the tensile agent is not sodium chloride.

8. The pharmaceutical formulation according to any one of claims 1-7, further comprising a surfactant, such as polysorbate 20 and / or polysorbate 80; preferably polysorbate 80.

9. The pharmaceutical preparation according to any one of claims 1-3, wherein it is a liquid preparation.

10. The pharmaceutical formulation according to claim 9, wherein the pH is 5.6-6.4, such as 5.7-6.4, for example about 5.7, about 5.8, about 5.9, about 6.0, about 6.1, about 6.2, about 6.3 or about 6.4; preferably about 5.8-6.

2.

11. The pharmaceutical formulation according to any one of claims 9-10, comprising at least 10% w / v and less than 22% w / v of cyclodextrin, such as 10-20% w / v of cyclodextrin, for example about 15% w / v of cyclodextrin.

12. The pharmaceutical preparation according to any one of claims 1-11, wherein the at least one preservative is phenol and / or m-cresol and / or EDTA.

13. The pharmaceutical preparation according to any one of claims 9-12, comprising m-cresol at a concentration of 9-40 mM and / or phenol at a concentration of 18-65 mM and / or EDTA at a concentration of 0.5-5.0 mg / ml.

14. A pharmaceutical preparation comprising: - 0.25-22 mg / ml canagliflozin, - Smegglutide at a concentration of 0.25-22 mg / ml - More than 10% w / v and less than 22% w / v, such as 10-20% w / v hydroxypropyl-substituted α- and / or β-cyclodextrins, wherein the cyclodextrin contains 0.58-1.0 hydroxypropyl groups per glucose unit. - Approximately 18-65 mM of phenol and / or 9-40 mM of m-cresol and / or 0.5-5.0 mg / ml of EDTA, - A buffer of about 3-30 mM with at least one pKa value of about 5.0-7.0, such as histidine or citrate. - Approximately 5-35 mg / ml of sorbitol, - Up to 1.0 mg / ml of polysorbate 20 and / or 80; such as 0.01-0.1 mg / ml, such as 0.05 mg / ml; or more than 0.1 mg / ml but less than 0.2 mg / ml of polysorbate 20 and / or 80. - Approximately 75-90% w / w of water, and The pH is approximately 5.6-6.4, for example, approximately 5.8-6.

2.

15. The pharmaceutical preparation according to any one of claims 1-14, which is used as a medicine.

16. A kit comprising a pharmaceutical composition as defined in any one of claims 1-15 and instructions for use.

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