Combination of an Insulin and a GLP-1 Agonist

Abstract

The invention relates to a drug comprising at least one insulin and at least one GLP-1 receptor agonist.

Claims

1. A pharmaceutical formulation comprising a combination of: (a) insulin glargine or a pharmaceutically acceptable salt thereof, and (b) desPro.sup.36exendin-4(1-39)-Lys.sub.6-NH.sub.2 (AVE0010) or a pharmaceutically acceptable salt thereof; wherein the concentration of insulin glargine is about 100 units/mL, and the concentration of AVE0010 is about 33 μg/mL; and wherein the dosage of insulin glargine is between about 15 units and about 80 units, and the dosage of AVE0010 is between about 5 μg and about 20 μg.

2. The pharmaceutical formulation of claim 1, wherein the dosage of insulin glargine is between about 15 units and about 60 units, and the dosage of AVE0010 is between about 5 μg and about 20 μg.

Description

DETAILED DESCRIPTION OF THE DRAWINGS

[0168] FIG. 1: Study design for oral glucose tolerance test

[0169] FIG. 2: OGTT in the dog: Effect of insulin glargine relative to placebo.

[0170] FIG. 3: OGTT in the dog: Effect of AVE0010 relative to placebo.

[0171] FIG. 4: OGTT in the dog: Effect of an AVE0010/insulin glargine combination on blood glucose level.

[0172] FIG. 5A: OGTT in the dog: Effect of an AVE0010/insulin glargine combination on plasma insulin.

[0173] FIG. 5B: OGTT in the dog: Effect of an AVE0010/insulin glargine combination on the c-peptide level.

[0174] FIG. 6: OGTT in the dog: Effect of a dose lowering of AVE0010 with different proportions relative to insulin glargine in the combined formulation.

[0175] FIG. 7: Effect of an AVE0010/insulin glargine combination on blood glucose in the diabetic db/db mouse.

[0176] FIG. 8: Effect of an AVE0010/insulin glargine combination in the oral glucose tolerance test in the diabetic db/db mouse.

[0177] FIG. 9A: Effect of an AVE0010/insulin glargine combination on cytokine-induced-cell apoptosis in vitro.

[0178] FIG. 9B: Effect of an AVE0010/insulin glargine combination on lipotoxicity-induced cell apoptosis in vitro.

[0179] FIG. 10: The “3 pens cover all” system.

EXAMPLES

Example 1

[0180] Model: Oral glucose tolerance test (OGTT) in healthy dogs: Comparison of the insulin glargine/AVE0010 combination with the two individual active compounds.

Animals

[0181] Male normoglycemic beagles [0182] Bodyweight: ˜15 kg [0183] Number per group: n=6

Study Design (See FIG. 1)

[0184] Individual subcutaneous injections of placebo or test formulation at time 0 [0185] 2 oral administrations of glucose, at 2 g of glucose/kg of bodyweight, at times 30 min and 5 h [0186] Blood samples are taken to determine blood glucose, plasma insulin, and c-peptide
Group Division (n=6) [0187] Placebo (Lantus placebo formulation without API) [0188] Insulin glargine (0.3 μl/kg s.c., equivalent to 1.8 nmol/kg). Insulin glargine is Gly(A21)-Arg(B31)-Arg(B32) human insulin. [0189] AVE0010 (10 μg/kg s.c. in Lantus placebo formulation, equivalent to 2 nmol/kg).

[0190] AVE0010 is des Pro.sup.36exendin-4(1-39)-Lys.sub.6-NH.sub.2. [0191] AVE0010/insulin glargine combination (10 μg/kg AVE0010/0.3 μl/kg insulin glargine s.c.)

Example 2

[0192] OGTT in the dog: Effect of insulin glargine relative to placebo

[0193] The experiment was carried out in accordance with the protocol described in example 1. [0194] repeated OGTT (2 g/kg p.o.) [0195] male beagle, n=6 [0196] mean±Sem [0197] placebo=Lantus placebo [0198] insulin glargine (0.3 U/kg s.c.)

[0199] Result: The data are shown in FIG. 2. The single administration of insulin glargine does not prevent the OGTT-induced increase in blood glucose. Insulin glargine reinforces the expected delayed lowering of blood glucose concentration in the postabsorptive phase.

Example 3

[0200] OGTT in the dog: Effect of AVE0010 relative to placebo

[0201] The experiment was carried out in accordance with the protocol described in example 1. [0202] repeated OGTT (2 g/kg p.o.) [0203] male beagle, n=6 [0204] mean±Sem [0205] placebo=Lantus placebo [0206] AVE0010 (10 μg/kg s.c.)

[0207] Result: The data are shown in FIG. 3. AVE0010 prevents the OGTT-induced postprandial increase in blood glucose almost completely. There is no effect on the glucose concentration in the postabsorptive phase. This example shows that the effect of AVE0010 on the OGTT-induced postprandial increase in blood glucose is complementary to the blood sugar-lowering effect of insulin glargine in the postabsorptive phase.

Example 4

[0208] OGTT in the dog: Effect of an AVE0010/insulin glargine combination on the blood glucose level.

[0209] The experiment was carried out in accordance with the protocol described in example 1 [0210] repeated OGTT (2 g/kg p.o.) [0211] male beagle, n=6 [0212] mean±Sem [0213] placebo=Lantus placebo [0214] AVE0010 (10 μg/kg s.c.) [0215] Insulin glargine (0.3 U/kg s.c.) [0216] AVE+Lan (=premix of 10 μg/kg of AVE0010 and 0.3 U/kg of insulin glargine in one formulation)

[0217] Result: The data are shown in FIG. 4. The combination has the same action on the postprandial glucose increase as AVE0010 (cf. example 3). The hypoglycemic effect of insulin glargine In the postabsorptive phase is likewise present, but attenuated (cf. example 2). This is a synergistic effect of insulin glargine and AVE0010, since AVE0010 alone has no effect on the level of glucose, which has fallen again following administration of glucose, and insulin glargine on its own has no effect on the postprandial glucose level.

Example 5

[0218] OGTT in the dog: Effect of an AVE0010/insulin glargine combination on the plasma insulin and the c-peptide level.

[0219] The experiment was carried out in accordance with the protocol described in example 1. [0220] repeated OGTT (2 g/kg p.o.) [0221] male beagle, n=6 [0222] mean±Sem [0223] placebo=Lantus placebo [0224] AVE0010 (10 μg/kg s.c.) [0225] Insulin glargine (0.3 U/kg s.c.) [0226] AVE+Lan (=premix of 10 μg/kg of AVE0010 and 0.3 U/kg of insulin glargine in one formulation)

[0227] The C-peptide is released in the course of the conversion of proinsulin to insulin, and serves as a marker for the secretion of insulin by the pancreatic β-cells. In a glucose loading test, the c-peptide can be used to determine the response capacity of the pancreas.

[0228] Result: The data are shown in FIG. 5A and FIG. 5B. In the combination group, the postprandial reduction in insulin is followed by an increased postabsorptive insulin glargine level C-peptide levels for the combination correspond to the insulin curve of AVE0010 during the prandial phases, and of insulin glargine during the postabsorptive phase.

Example 6

[0229] OGTT in the dog: Effect of a dose lowering of AVE0010 with different proportions to insulin glargine in the combined formulation.

[0230] The experiment was carried out in accordance with the protocol described in example 1. [0231] repeated OGTT (2 g/kg p.o.) [0232] male beagle, n=11/6/6/6 [0233] mean±Sem [0234] control=Lantus placebo [0235] AVE+Lan (=premix of 0.15 to 1.0 μg/kg of AVE0010 and 0.3 U/kg of insulin glargine in one formulation). In examples 2 to 5, AVE0010 concentrations of 10 μg/kg were used.

[0236] Result: The data are shown in FIG. 6. A reduction in the AVE0010 dose from 10 μg/kg (cf. in particular example 4) to 1 μg/kg (i.e., by a factor of 10), and the resultant increase in the proportion of insulin glargine to AVE0010, has no effect on the synergistic activity of the combination of AVE0010 with insulin glargine (cf. in particular example 4). Only at significantly smaller AVE0010 doses does the effect of the combination approach the effect of insulin glargine alone (ct in particular FIG. 2). The AVE0010 dose may therefore be varied at least within one order of magnitude (i.e., by a factor of at least 10) without loss of the synergistic effect.

Example 7

[0237] Model: Diabetic, insulin-resistant db/db mouse: Comparison of the insulin glargine/AVE0010 combination with the two individual active compounds.

[0238] Animals [0239] Female db/db mouse [0240] Age: 10-11 weeks [0241] Number per group: n=10

Study Design

[0242] Individual subcutaneous injection of placebo or test formulation [0243] Taking of blood samples to determine blood glucose

Group Division

[0244] Placebo (=Lantus placebo formulation without API) [0245] AVE0010 (10 μg/kg s.c.) [0246] Insulin glargine (5 IU/s.c.) [0247] AVE0010/insulin glargine combination (premix of 10 μg/kg of AVE0010 plus 5 IU/kg of insulin glargine s.c.)

Example 8

[0248] Effect of an AVE0010/insulin glargine combination on blood glucose in the diabetic db/db mouse

[0249] The experiment was carried out in accordance with the protocol described in example 7. [0250] Female db/db mouse, 10 weeks [0251] n=10, mean±Sem [0252] Vehicle=Lantus placebo [0253] AVE0010 (10 μg/kg sc) [0254] Lantus (5 U/kg sc) [0255] AVE0010/insulin glargine (=premix of AVE0010 10 μg/kg and insulin glargine 5 U/kg in one formulation)

[0256] Result: The data are shown in FIG. 7. In diabetic db/db mice, the AVE0010/insulin glargine combination produced a more rapid and more pronounced decrease in the blood glucose concentration as compared with the two individual active compounds. Consequently the combination takes diabetic db/db mice closer to normoglycemia than either of the two active compounds alone.

Example 9

[0257] Effect of an AVE0010/insulin glargine combination in the oral glucose tolerance test in the diabetic db/db mouse

[0258] The experiment was carried out in accordance with the protocol described in example 7. Additionally an OGTT (2 g/kg p.o. @ 30 min) was carried out. [0259] Female db/db mouse, 11 weeks [0260] n=10, mean±Sem [0261] Control=Lantus placebo [0262] AVE0010 (10 μg/kg sc) [0263] Insulin glargine (5 U/kg sc) [0264] AVE0010/insulin glargine (=premix of AVE0010 10 μg/kg and insulin glargine 5 U/kg in one formulation)

[0265] Result The data are shown in FIG. 8. The AVE0010/insulin glargine combination leads to significantly improved glucose tolerance and lower postabsorptive glucose levels.

Example 10

[0266] Effect of the AVEOO 10/insulin glargine combination on cytokine- and lipotoxicity-induced β-cell apoptosis in vitro. [0267] Insulinoma cell line INS-1, rat [0268] Incubation with the test compound for 5 h [0269] Further incubation with a cytokine mix for 22 h (1 ng/mL IFN-γ+4 ng/mL IL-1β) or [0270] Further incubation with 0.5 mM FFA for 18 h (palmitates: BSA 3:1)

[0271] The measures used for the apoptosis are the caspase-3 activity and the fragmentation of the cell nuclei, which correlate with apoptosis.

[0272] Result: The data are shown in FIG. 9A and FIG. 9B. AVE0010 or insulin glargine (glargine, Glar) alone prevent the apoptosis by ˜40-50%. The AVE0010 and insulin glargine combination prevents apoptosis significantly better. On the basis of this synergistic effect, the combination brings about increased protection against cytokine- and lipotoxicity-induced apoptosis.

Example 11

[0273] The “3 pens cover all” system (FIG. 10) [0274] 3 premix pens with 3 different predetermined proportions: [0275] Mix A: 100 U of insulin glargine+66.66 μg of AVE0010 per mL [0276] Mix B: 100 U of insulin glargine+40 μg of AVE0010 per mL [0277] Mix C: 100 U of insulin glargine+25 μg of AVE0010 per mL. [0278] Use of the 3 premix pens: The table in FIG. 10, representing an example, starts from a therapeutic range of 15 to 80 U per dose of insulin glargine and 10 to 20 μg of AVE0010. For a particular patient, a dose of insulin glargine to be administered is specified or predetermined. The predetermined dose is looked up in the left-hand column. Where the columns MIX A-MIX C specify a corresponding AVE0010 dose in the range between 10 and 20 μg, the corresponding MIX is selected, dosed, and administered. The ranges are overlapping: for example, in the case of a requirement of 26 to 30 U of insulin glargine, it will be possible to choose MIX A or MIX B (with a higher dose of AVE0010). The same applies to MIX B and C. If, for example, a dose of 50 U of insulin is intended, then 0.5 ml of MIX B or MIX C can be dosed. This dose contains 20 μg (MIX B) or 12.5 μg (MIX C) of AVE0010. [0279] Conclusion: On the assumption that a probable AVE0010 effect is obtained at between 10 and 15 μg, and a therapeutic effect between 15 and 22 μg, almost all patients who take insulin glargine doses of 15-80 U can also obtain therapeutic doses of AVE0010 of between 10 and 20 μg if they use one of the three premix pens which contain three different insulin glargine:AVE0010 ratios (Mix A, B or C). On the basis of the broad range of possible proportions of insulin glargine to AVE0010 (cf. example 6) with a synergistic effect, the proportions in the pens can be tailored such that for each dose of insulin glargine there is a synergistic dose of AVE0010 in at least one pen.

Example 12

[0280] This example shows how a combination of two or more active compounds can be formulated in such a way that, when two or more compositions are combined, both active compounds can be administered in any desired amounts and in any desired proportions to one another. It is taken into account here that at least one of the active compounds must not be diluted as a result of the combining (e.g., through mixing directly prior to administration).

[0281] In this example, the designations “active A” and “active B” stand for any desired active compounds. In particular, active A is an insulin and active B is a GLP-1 agonist. Active A can also be a GLP-1 agonist, and active B can also be an insulin.

1. Comparative Example

[0282] For a combination therapy with an active A (e.g., an insulin) and an active B (e.g., a GLP-1 agonist), a container 1 with a composition with active A at a concentration of a mg/ml, and a container 2 with a composition with active B at a concentration of b mg/ml, are provided.

[0283] For the administration of a combination of the two actives, a volume V.sub.1 ml from container 1 and a volume V.sub.2 ml from container 2 are mixed.

[0284] For the dosing of the two actives, at given concentrations a and b, the volumes V.sub.1 and V.sub.2 to be administered are selected in dependence on the amount of the actives A and B to be administered. The volumes V.sub.1 and V.sub.2 of the two actives are determined on the basis of the amount of active, as follows:

V.sub.1.Math.a mg  Amount of active A:

V.sub.2.Math.b mg  Amount of active B:

[0285] The concentrations of the actives A and B in the mixture of the two compositions are determined as follows.

x mg/mL=V.sub.1.Math.a/(V.sub.1+V.sub.2)  Active A:

y mg/mL=V.sub.2.Math.b/(V.sub.1+V.sub.2)  Active B:

[0286] V.sub.1+V.sub.2 is the total administered volume. This means that the two actives dilute one another. With this system, therefore, it is not possible to keep, for example, the concentration of the active A (e.g., of the insulin) at a predetermined level in the case of varying amounts of active B.

2. Inventive Example

[0287] In this example, for a combination therapy with an active A (e.g., an insulin) and an active B (e.g., a GLP-1 agonist), a container 1 with a composition with active A at a concentration of a mg/ml, and a container 2 with a composition with active A at a concentration of a mg/ml and with active B at a concentration of b mg/ml, are provided. The concentration of the active A is therefore the same in both compositions.

[0288] For the administration of a combination of the two actives, a volume V.sub.3 ml from container 1 and a volume V.sub.2 ml from container 2 are mixed.

[0289] For the dosing of the two actives, at given concentrations a and b, the volumes V.sub.3 and V.sub.2 to be administered are selected in dependence on the amount of the actives A and B to be administered. The volumes V.sub.3 and V.sub.2 of the two actives are determined on the basis of the amount of active, as follows:

(V.sub.3.Math.a+V.sub.2).Math.a(mg))  Amount of active A:

V.sub.2.Math.b mg  Amount of active B:

[0290] The concentrations of the actives A and B are determined as follows.

a mg/mL=(V.sub.3.Math.a+V.sub.2.Math.a)/(V.sub.3+V.sub.2)  Active A:

z mg/mL=V.sub.2.Math.b/(V.sub.3+V.sub.2)  Active B:

[0291] V.sub.3+V.sub.2 is the total administered volume. From the above calculation it is evident that the concentration of the active A is always a mg/ml, i.e., is constant, irrespective of what volume ratio V.sub.3/V.sub.2 is being dosed.

[0292] Comparing the comparative example (see section 1) with the present inventive example, it is apparent that, for an equal dosing quantity of actives A and B, the total volume required in the inventive example is lower.

[0293] For a given dose (amount of active compound) of the active A, the figure in the comparative example is:

V.sub.1.Math.a mg

[0294] In the inventive example it is:

(V.sub.3.Math.a+V.sub.2.Math.a)mg

[0295] Since the amount of active compound is to be the same in both cases,

(V.sub.3.Math.a+V.sub.2.Math.a)=V.sub.1.Math.a

(V.sub.3+V.sub.2).Math.a=V.sub.1.Math.a

and V.sub.3+V.sub.2=V.sub.1

or V.sub.3=V.sub.1−V.sub.2

[0296] Here, the volume V.sub.2 in which the active B is administered is the same in both case

[0297] The total volume in the comparative example is V.sub.1+V.sub.2

[0298] The total volume in the inventive example is V.sub.3+V.sub.2

[0299] According to the above equation, for the inventive example it is the case that:

V.sub.3+V.sub.2=V.sub.1−V.sub.2+V.sub.2=V.sub.1

[0300] This volume V.sub.1 is smaller than the volume V.sub.1+V.sub.2 of the comparative example.

[0301] As a result of the mixing of the composition with actives A and B with the composition with active A, active B is diluted. This dilution is less than the dilution of the active B in the comparative example (i.e., the concentration b>concentration z>concentration

b>z

b>V.sub.2.Math.b/(V.sub.3+V.sub.2)

b>b V.sub.2/(V.sub.3+V.sub.2), where V.sub.2/(V.sub.3+V.sub.2) is <1, and

z>y

V.sub.2.Math.b/(V.sub.3+V.sub.2)>V.sub.2.Math.b/(V.sub.1+V.sub.2)

1/(V.sub.3+V.sub.2)>1/(V.sub.1+V.sub.2)

1/(V.sub.1−V.sub.2+V.sub.2)>1/(V.sub.1+V.sub.2)

1/V.sub.1>1/(V.sub.1+V.sub.2)

[0302] Hence the dosing system of the invention for administering variable doses of the actives A (e.g., an insulin) and B (e.g., a GLP-1 agonist) has three advantages over the comparative system: [0303] The concentration of active A (e.g., an insulin) can be kept constant at a predetermined level [0304] Where the doses of actives A and B to be administered are the same, the total volume to be administered is smaller. [0305] The dilution of active B (e.g., the GLP-1 agonist) is less than in the comparative experiment. Accordingly the concentration of active B can be held more easily within a predetermined range.

[0306] The present example can be readily extended to medicaments with three or more active compounds, the first active compound being present in all of the compositions (preferably in equal weight fractions) and there being at least one further active compound in each further composition. The first composition can be mixed with each further composition in the same proportion without the concentration of the active compound in the first composition becoming diluted.