COMPOSITIONS COMPRISING AT LEAST AN AMYLIN RECEPTOR AGONIST AND A GLP-1 RECEPTOR AGONIST

Abstract

An invention relates to therapies for treating obesity, overweight and/or diabetes, in particular type 2 diabetes. The invention relates to a composition including at least one amylin receptor agonist, such as pramlintide, and at least one GLP-1 receptor agonist, such as exenatide or lixisenatide. The invention also relates to a composition for use in the method for the treatment of obesity, overweight and/or diabetes, in particular type 2 diabetes, wherein it reduces the slowing effect of gastric emptying of amylin RA as compared to the same composition without the corresponding GLP-1 RA.

Claims

1. A composition comprising an amylin receptor agonist and a GLP-1 receptor agonist (GLP-1 RA).

2. The composition according to claim 1 comprising pramlintide and a GLP-1 receptor agonist (GLP-1 RA).

3. The composition according to claim 1, wherein the GLP-1 receptor agonist (GLP-1 RA) is chosen amongst lixisenatide and exenatide.

4. The composition according to claim 1, wherein the GLP-1 receptor agonist (GLP-1 RA) is exenatide.

5. The composition according to claim 1, wherein it is in the form of an injectable aqueous formulation.

6. The composition according to claim 5, wherein its pH is ranging from 3.5 to 4.4.

7. The composition according to claim 1, wherein it comprises a pramlintide at a concentration of ranging from 0.5 to 10 mg/ml.

8. The composition according to claim 1, wherein it comprises a exenatide at a concentration of ranging from 20 to 500 μg/ml.

9. The composition according to claim 1, wherein ratio pramlintide/exenatide (weight/weight) is ranging from 2 to 50.

10. The composition according to claim 1, for use in the method for the treatment of obesity, overweight and/or diabetes, in particular type 2 diabetes, wherein it reduces the slowing effect of gastric emptying of amylin RA as compared to the same composition without the corresponding GLP-1 RA.

11. The composition according to claim 1, for the treatment of obesity and/or diabetes, in particular Type 2 Diabetes, wherein the daily dose of pramlintide is ranging from 150 to 2000 μg/day

12. The compositions according to claim 1, wherein the daily dose of exenatide is ranging from 10 to 250 μg.

Description

EXAMPLES

Compositions

Example C1: 250 μg/mL Exenatide Solution (Byetta®)

[0503] This solution is a commercial solution of exenatide marketed by the company NOVO NORDISK under the name of Byetta®. The excipients in this composition C1 are detailed in Table 1.

Example C1′: Preparation of a 2 mg/mL Exenatide Solution at pH 4 (Composition C1′)

[0504] Exenatide powder (obtained by peptide synthesis) is dissolved with a 10 mM HCl solution. An acetate buffer solution is then added to give the composition C1′. The composition is detailed in Table 1.

Example C2: Preparation of a 250 μg/mL Pramlintide Solution at pH 4 (Composition C2)

[0505] A 5 mg/mL concentrated solution of pramlintide (obtained by peptide synthesis) and excipients is prepared via a stepwise addition in an empty container in the following order: m-cresol, acetate buffer, mannitol and pramlintide. Sufficient quantity of water is then added and the pH is adjusted to 4 to obtain composition C2 which is detailed in Table 1.

Example C2′: Preparation of a 10 mg/mL Pramlintide Solution at pH 4 (Composition C2′)

[0506] Pramlintide powder (obtained by peptide synthesis) is dissolved with a 10 mM HCl solution. An acetate buffer solution is then added to give the composition C2′. The composition is detailed in Table 1.

Example C3: Preparation of a 3 mg/mL Lixisenatide Solution at pH 4 (Composition C3)

[0507] Lixisenatide powder (obtained by peptide synthesis) is dissolved with a 10 mM HCl solution. An acetate buffer solution is then added to give the composition C3. The composition is detailed in Table 1.

Example C4: Preparation of a Stock Solution of Excipients (Matrix) at pH 4 (Composition C4)

[0508] A concentrated solution of excipients is prepared via a stepwise addition in an empty container in the following order: m-cresol, acetate buffer and mannitol. Sufficient quantity of water is then added and the pH is adjusted to 4 to obtain Composition C4. This composition is detailed in Table 1.

Example C5: Preparation of a Dilution Solution at pH 4 (Composition C5)

[0509] A concentrated solution of excipients is prepared via a stepwise addition in an empty container in the following order: BSA (Bovine Serum Albumine), acetate buffer, NaCl and polysorbate 20. Sufficient quantity of water is then added and the pH is adjusted to 4 to obtain Composition C5.

Example C6: Preparation of a Solution of a Vehicle (Composition C6)

[0510] To a volume VC4 of the composition C4, as prepared in example C4, is added a VC5 of the dilution solution C5, as prepared in example C5 to obtain the final solution C6 which composition is detailed in Table 1.

Example C7: Preparation of Solutions of Pramlintide Alone or in Combination with Exenatide or Lixisenatide at pH 4

[0511] The dilution solution (Composition C5) is used to dilute the solution of pramlintide (Composition C2) and of exenatide (Composition C1—Byetta) to prepare the solutions C7-1 to C7-8 as shown in Table 1 that will be injected to the animals during the in vivo assays.

[0512] The compositions of pramlintide and exenatide C7-9 to C7-33 are obtained by mixing a VC4 of composition C4, a volume VC1′ or VC1 of the composition of exenatide C1′ or C1 and a VC2′ of the composition of pramlintide C2′.

[0513] The compositions of pramlintide and lixisenatide C8-1 to C8-24 are obtained by mixing a VC4 of composition C4, a volume VC3 of the composition of lixisenatide C3 and a VC2′ of the composition of pramlintide C2′.

TABLE-US-00004 TABLE 1 Stock solutions C1 to C4, dilute solution C5, vehicle solution C6, diluted solution of pramlintide C7-1 and C7-2 and compositions according to the invention C7-3 to C7-33 and C8-1 to C8-24 prepared to be injected to the animals in the in vivo assays. Pramlin- Compo- tide Exenatide Lixisenatide sition (mg/mL) (μg/mL) (μg/mL) Excipients C1 — 250 — m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C1′ — 2000 — Acetate buffer (20 mM) C2 0.250 — — m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C2′ 10 — — Acetate buffer (20 mM) C3 — — 3000 Acetate buffer (20 mM) C4 — — — m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C5 — — — Acetate buffer (20 mM) BSA (1 mM) NaCl (130 mM) Polysorbate 20 (0.5 mM) C6 — — — m-cresol (3.3 mM) Acetate buffer (20 mM) Mannitol (42 mM) BSA (0.83 mM) NaCl (108 mM) Polysorbate 20 (0.42 mM) C7-1 0.0042 — — m-cresol (3.3 mM) Acetate buffer (20 mM) Mannitol (42 mM) BSA (0.83 mM) NaCl (108 mM) Polysorbate 20 (0.42 mM) C7-2 0.042 — — m-cresol (3.3 mM) Acetate buffer (20 mM) Mannitol (42 mM) BSA (0.83 mM) NaCl (108 mM) Polysorbate 20 (0.42 mM) C7-3 0.0042 0.42 — m-cresol (3.4 mM) Acetate buffer (20 mM) Mannitol (42 mM) BSA (0.83 mM) NaCl (108 mM) Polysorbate 20 (0.42 mM) C7-4 0.042 0.42 — m-cresol (3.4 mM) Acetate buffer (20 mM) Mannitol (42 mM) BSA (0.83 mM) NaCl (108 mM) Polysorbate 20 (0.42 mM) C7-5 0.042 4.2 — m-cresol (3.4 mM) Acetate buffer (20 mM) Mannitol (42 mM) BSA (0.83 mM) NaCl (108 mM) Polysorbate 20 (0.42 mM) C7-6 0.042 0.83 — m-cresol (3.4 mM) Acetate buffer (20 mM) Mannitol (43 mM) BSA (0.83 mM) NaCl (108 mM) Polysorbate 20 (0.42 mM) C7-7 0.042 3.3 — m-cresol (3.6 mM) Acetate buffer (20 mM) Mannitol (45 mM) BSA (0.82 mM) NaCl (107 mM) Polysorbate 20 (0.41 mM) C7-8 0.042 8.3 — m-cresol (4.0 mM) Acetate buffer (17 mM) Mannitol (50 mM) BSA (0.67 mM) NaCl (87 mM) Polysorbate 20 (0.33 mM) C7-9 0.6 24 — m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C7-10 0.6 36 — m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C7-11 0.6 48 — m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C7-12 0.6 60 — m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C7-13 0.6 72 — m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C7-14 1 40 — m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C7-15 1 60 — m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C7-16 1 80 — m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C7-17 1 100 — m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C7-18 1 120 — m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C7-19 2 80 — m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C7-20 2 120 — m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C7-21 2 160 — m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C7-22 2 200 — m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C7-23 2 240 — m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C7-24 3 120 — m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C7-25 3 180 — m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C7-26 3 240 — m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C7-27 3 300 — m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C7-28 3 360 — m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C7-29 4 160 — m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C7-30 4 240 — m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C7-31 4 320 — m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C7-32 4 400 — m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C7-33 4 460 — m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C8-1 0.6 — 60 m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C8-1 0.6 — 90 m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C8-2 0.6 — 120 m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C8-3 0.6 — 150 m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C8-4 0.6 — 180 m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C8-5 1 — 100 m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C8-6 1 — 150 m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C8-7 1 — 200 m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C8-8 1 — 250 m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C8-9 1 — 300 m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C8-10 2 — 200 m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C8-11 2 — 300 m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C8-12 2 — 400 m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C8-13 2 — 500 m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C8-14 2 — 600 m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C8-15 3 — 300 m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C8-16 3 — 450 m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C8-17 3 — 600 m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C8-18 3 — 750 m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C8-19 3 — 900 m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C8-20 4 — 400 m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C8-21 4 — 600 m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C8-22 4 — 800 m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C8-23 4 — 1000 m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM) C8-24 4 — 1200 m-cresol (20 mM) Acetate buffer (20 mM) Mannitol (250 mM)

In Vivo Studies

[0514] Effects of Pramlintide and Pramlintide with Exenatide Compositions on Gastric Emptying

[0515] The aim of this study was to evaluate the effects of pramlintide compositions C7-1 and C7-2 and pramlintide combined with exenatide compositions (C7-3 to C7-5) on gastro-intestinal transit (gastric emptying) in the mouse.

Example D1: General Procedure for In Vivo Studies of Compositions of Pramlintide Alone and Pramlintide in Combination with Exenatide

[0516] Studies were conducted on naïve male Swiss mice (Janvier Lab) as no gender specific effect were expected. Body weight range of 25-32 g were included in the analysis.

[0517] Mice were approximately fasted for 18 hours (˜T0−18 h). The gastric emptying (i.e. stomach content) was measured via the administration of a suspension of charcoal at T0 (activated charcoal (0.4 ml/mouse, ˜0.4 mg/mouse; Sigma Reference number C9157), suspended 10% with 2.5% arabic gum in distilled water). The studied composition or a vehicle (as a standard control) was administered subcutaneously using Hamilton syringes 20 minutes before charcoal (T0−20 min), then 50 minutes later (T0+50 min), they were sacrificed, and the stomach was removed. The stomach was excised from the abdomen and weighed with its contents (full stomach weight). Afterwards, the stomach was opened, the contents washed out and the weight recorded again (empty stomach weight).

[0518] The results are expressed as the variation of gastric content with the administration compositions comprising pramlintide compared with the gastric content with administration the vehicle (weight of gastric content with pramlintide/weight of gastric content with vehicle).

Example D2: Gastric Emptying Measurements with Compositions Comprising Pramlintide Alone

[0519] Measurements of the gastric emptying for compositions C7-2 were obtained according to the general procedure. Five different doses of pramlintide were studied. Each dose was tested on 4 mice. The results of this study are presented in table 2.

TABLE-US-00005 TABLE 2 Effect of compositions of pramlintide alone on the gastric content Dose of Concentration Variability of the Compo- pramlintide of pramlintide stomach content weight sition (μg/kg) (μg/mL) compared to the vehicle C6 —* 0 — (Vehicle) C7-2 50 42 +209% (±10%) C7-2 80 42 +166% (±25%) C7-2 150 42 +233% (±20%) C7-2 250 42 +144% (±20%) C7-2 350 42 +205% (±30%) *Vehicle administered

[0520] These examples demonstrate that subcutaneous injection of a composition comprising pramlintide alone increases the stomach content weight i.e. slows down the gastric emptying.

1

Example D3: Gastric Emptying Measurements of Compositions Comprising a Combination of Pramlintide and Exenatide at Different Doses Compared to Compositions of Pramlintide Alone

[0521] Measurements of the gastric emptying for compositions C6 and C7-1 to C7-5 were obtained according to the general procedure described in example D1. Two sets of experiments were carried out. Each dose was tested on 8 mice apart for composition C7-2 that was tested on 10 mice. The results are presented in table 3.

TABLE-US-00006 TABLE 3 Effect of compositions of pramlintide/exenatide on the gastric content Variability of the stomach content Variability of weight compared to the stomach the content weight Pramlin- content weight of the composition Compo- tide Exenatide compared to of pramlintide alone sition (μg/kg) (μg/kg) the vehicle at the same dose C6 —* — — C7-1 12.5 — +158% (+10%) — C7-3 12.5 0.125 +100% (±10%) −22% C7-2 25 — +118% (±15%) — C7-4 25 0.25  +7.5% (±15%) −49% C7-4 50 0.5 +107% (±20%) — C6 —* — — — C7-2 25 —  +75% (±15%) C7-5 25 5  −18% (±10%) −53% C7-5 25 2  −24% (±20%) −57% C7-4 25 0.5 +8% −38% *Vehicle administered

[0522] These results show that pramlintide alone increases the gastric content weight i.e. slows down the gastric emptying and that the addition of exenatide to pramlintide decreases the gastric content weight in comparison with the same composition without exenatide i.e. increase the gastric emptying.