HK1023943B - Medicament for the treatment of diabetes mellitus as well as its subsequent disorders - Google Patents

Description

The invention relates to medicinal products for the treatment of late complications and complications of type I and type II diabetes mellitus or subclinical insulin resistance and their late complications and complications.

R- ((+) -α-lipoic acid is the physiologically occurring enantiomer of 1,2-dithiocyclopentan-3-valerian acid. R-(+) -α-lipoic acid is a ubiquitous coenzyme of α-ketose dehydrogenases (pyruvate dehydrogenase, α-ketoglutarate dehydrogenase, etc.) in plants and animals, acting at a key site of sugar and energy metabolism in the cell.

Racemat, a 50/50 mixture of R- ((+) -α-lipoic acid and S- ((-) -α-lipoic acid, is used to treat diabetic and alcoholic polyneuropathy, tuber leaf fungal poisoning and chronic and alcoholic liver disease.

It is known that the enantiomers of α-lipoic acid differ in various pharmacological properties, e.g. in anti-inflammatory and analgesic effects. The Commission has decided to initiate proceedings against the applicant. It is further described in the literature that R,S- ((+,-) -α-lipoic acid has a blood glucose lowering effect in alloxan-induced diabetes in animal models, although it is unclear whether this effect is due to an influence on insulin secretion or directly to the activation of pyruvate dehydrogenase (Natraj C.V. et al. J. Biosci. Vol. 6 ((1), 37-46, 1984). Metabolic abnormalities in diabetes such as hyperglycaemia, ketonemia, ketonuria, reduced tissue glycogen and decreased fatty acid synthesis in the liver are corrected by administration of lipoic acid in animal experiments.

It is also known that oxidative stress is a promoter of late-onset diabetic complications and that adjuvant antioxidant therapy (with thiotcacid) can lead to regression of late-onset diabetic complications.

In vitro experiments with thioctic acid (material of the family Calbiochem ((Racemat)) have shown that it increases glucose uptake by the muscle. Time-tested studies have shown that, unlike the stimulating effect of insulin on glucose uptake, the effect of thioctic acid on rat diaphragms in vitro is only apparent after prolonged incubation. According to Haugaard, the mechanism of action of thioctic acid does not seem to be similar to that of insulin. Its effectiveness is additive to that of desulins. (N. and E.S. Haugaard, Biochim. Biophys. Acta 222, ((1970) 583-586). However, no information on a differentiated effect of thioctic acid or S-acet is found in the literature. Diabetes mellitus is a disease characterised by insulin deficiency or resistance to insulin action (decompensated insulin resistance).As a result, even with compensated insulin resistance (decrease in insulin activity without clinically manifest type II diabetes), numerous metabolic disorders, particularly carbohydrate and fat metabolism, occur. These disorders can lead to coma and death in the long term. Both insulin resistance and elevated blood sugar and impaired fat metabolism are involved in the development of complications and complications (e.g. cataracts, neuropathies, nephropathy). A fundamental disorder in diabetes and insulin resistance is glucose uptake by muscle cells.In this context, it is particularly important in the context of insulin resistance to treat glucose uptake by means other than insulin administration and insulin release stimulating drugs, but by mechanisms independent of these (Häring H.U., Mehnert H. Diabetologica 36, 176-182,

The metabolism of glucose required after cellular uptake in the mitochondrial energy metabolism is another necessary step, especially in the case of impaired glucose utilization in the context of insulin resistance.

Diabetics show increased glycosylation and oxidation of proteins with corresponding negative consequences for the patient (Makita Z. et al., Science 258, 651-653, 1992).

The finding that R- ((+) -α-lipoic acid is specifically suitable for the treatment of diabetes mellitus and insulin resistance, whereas 5- ((-) -α-lipoic acid is not practical, was new and unexpected and not derivable for the expert.

This is of clinical relevance in pathological disorders of blood glucose regulation such as type I and type II diabetes mellitus and in disorders of insulin sensitivity of the tissues (insulin resistant). This is true in monotherapy and in combination with other medicinal products for the treatment of diabetes mellitus and insulin resistance, e.g. oral insulin therapy and especially insulin therapy, and may also be used as a treatment for diabetes mellitus and other complications of diabetes mellitus.

Surprisingly, R- ((+) -α-lipoic acid has now been found to be preferentially used for the treatment of secondary and late complications of type I and II diabetes mellitus, such as cataracts and nephropathy, and for the treatment of subclinically and clinically manifested insulin resistance and its sequelae.

Pharmaceutical examples 1.Pyruvate dehydrogenase activity after chronic administration in various tissues of the spontaneous diabetic rat Results

Tendency after double dose: decrease by S- ((-) -α-lipoic acid, increase by R- ((+) -α-lipoic acid

Test description

Spontaneous diabetic rats (BB-Wol BB, Moellegard, Denmark, n=10/group) were administered 0,3 ml of neutral 0,12 M (corresponding to 50 mg/kg body weight) R-(+) -α-lipoic acid or S-(-) -α-lipoic acid daily to the tail vein for seven days after the onset of diabetes. A control group was given saline. After seven days the animals were killed. Pyruvate hydrogenases activity was determined from the heart muscle. The tissue was homogenised.

Measurement of pyruvate dehydrogenase activity

Other The test method is to use a test chemical that is a mixture of the following:

The extinction of the reduced coenzyme is measured at 339 nm in vats with a Shimadzu UV 210 detector at 37 °C. The isolation of the enzyme complex (Köplin R. Ph.D. Thesis, University of Tübingen, FRG, 1988; Stanley C.J., Perham R.N. Biochem. J. 191, 147-154, 1980) and the enzyme assay (Lowry O.H. et al.. J. Biol. Chem. 256, 815-822, 1951) are described.

2.Glucose uptake by muscle cells under insulin (clip) Results Glucose uptake

The R-enantiomer (2.5 mM) stimulates glucose uptake by more than a factor of 2 compared to the S-enantiomer, while the S-enantiomer shows less efficacy at the same concentration. Other Glukoseaufnahme in Muskelzellen

15	15,1 ± 0.4	16,7 ± 0,6	16,3 ± 0,3
30	12,1 ± 0	15,9 ± 0,9	14,8 ± 0,7
60	16,5 ± 0,4	26,1 ± 0,9	21.6 ± 0,4
120	15,7 ± 0,6	27.0 ± 0,4	20,5 ± 0,8

Glukoseaufnahme Muskelzellen in Verbindung mit Insulin (200 nM) R-(+)-α-Liponsäure (2,5 mM)

15	20,0 ± 0,9	23,2 ± 0,5	24,7 ± 0,9	25,1 ± 0,6
30	18,1 ± 0,6	21,1 ± 0,4	21,6 ± 0,4	21,1 ± 0,2
60	18,0 ± 0,6	25,7 ± 0,5	23,7 ± 0,5	26,2 ± 0,7

The effect of the R-enantiomer is similar to that of insulin (200 nM), but is not additive. Other Glukoseaufnahme Muskelzellen in Verbindung mit Insulin (200 nM) S-(-)-α-Liponsäure (2,5 mM)

15	14.5 ± 0,3	14,8 ± 0,4	17,7 ± 0,3	16,0 ± 0,4
30	13,8 ± 0,5	13,3 ± 0,4	16,3 ± 0.5	15,7 ± 0,3
60	15,6 ± 0,5	16,0 ± 0,2	22,3 ± 0,5	19,8 ± 1,1

Test description

The tissue muscle cells (L6-myotubes) were placed and differentiated in 24-hole plates. After incubation with the test substances, an assay was performed for hexose uptake (3H-2-deoxyglucose, 10 μM, 10 min). Insulin was added at a concentration of 200 nM, the α-lipoic acid enantiomers at a concentration of 2.5 mM. After washing the cells, cellulose with NaOH, the uptake radioactivity was measured on the counter. Parallel test approaches with cytochalasin were performed to determine the glucose transporter-dependent glucose transduction. The results can be expressed in pmol/min x mg protein. The tests were performed according to the described biochemical method (Koivisto et al., J.-M., 1991; 26-262 U.M., 26-156).

4.Effects on the translocation of glucose transporters Results

R- ((+) -α-lipoic acid stimulates the translocation of glucose transporters (Glut 1 and GLUT 4) from the cytosol to the plasma membrane, which is equivalent to activation. S- ((-) -α-lipoic acid has no or no inhibitory effect and appears to decrease the total glucose transporter content in the cell (GLUT4). Other Einfluß von Enantiomeren der α-Liponsäure (2,5 mM) auf die Translokation von GLUT1 Glukosetransportern in L6-Mytubes

Kontrolle	1,00	1,00
R-(+)-Lipoat	1,56 ± 0,25	0,46 ± 0,06
S-(-)-Lipoat	0.93 ± 0,37	0,38 ± 0,09
Insulin	1,07 ± 0,14	0,68 ± 0,10

Einfluß von Ennatiomeren der α-Liponsäure (2,5 mM) auf die Translokation von GLUT4 Glukosetransportern in L6-Myotubes

Kontrolle	1,00	1,00
R-(+)-Lipoat	1,40 ± 0,08	0,59 ± 0,04
S-(-)-Lipoat	0,84 ± 0,37	0,71 ± 0,11
Insulin	1,38 ± 0,09	0,75 ± 0,11

Test description

L6-myotubes are used in 15-cm shells (n=4-5) and incubated for one hour with 2.5 mM lipoate in MEM with 5 mM glucose and 2% fetal bovine serum. The cells are removed, homogenised and processed into fractions (4°C). The processing is done in a HEPES buffer with a defined protease inhibitor additive. The cell fractions are obtained in 6 defined zenrifugation steps. The fractions are added to a 10 % polyacrylamide Western blot gel. The glucose transporters are analyzed with anti-GLUT1 and anti-GLUT4 antibodies using iodine-labelled protein A and autoradiographic detection.

5. Influence on the cellular content of glucose transporters Results

R- ((+) -α-lipoic acid increases the cellular content of glucose transporters GLUT1 and GLUT4 after 4 hours incubation. Other Einfluß von Liponsäure-Enantiomeren (2,5 mM) nach 4 Stunden Inkubation auf den Gehalt von Glukosetransportern in L6-Myotubes

Kontrolle	1,00	1,00
R-(+)-Lipoat	1,81 ± 0,01	1,55 ± 0,24
S-(-)-Lipoat	1.08 ± 0,01	0,79 ± 0,47

Test description

L6 myotubes are incubated for 4 hours with 2.5 mM lipoic acid enantiomers in MEM medium with 2% fetal calf serum and 5 mM glucose.

6.Tissue damage related to diabetes Results

In an animal model of diabetes (streptozotocin-induced diabetes), it was now found, surprisingly, that R-thioctic acid corrected numerous pathologically altered parameters (glycosylated haemoglobin, protein oxidation), while the S-enantiomer showed little or no effect. Other Glykosiliertes Hämoglobin

Kontrolle	9,7 ± 1,5 (n=8)
R-Thioctsäure Diät	8,4 ± 1,3 (n=11)
S-Thioctsäure Diät	10,7 ± 2,1 (n=6)

Protein-carbonylbildung in Linse und Leber

Kontrolle	0,513 ± 0,015 (n=3)	100 ± 8,9 (n=6)
R-Thioctsäure-Diät	0,429 ± 0,063 (n=3)	73,2 ± 17,8 (n=6)
S-Thioctsäure-Diät	0,554 ± 0,022 (n=3)	90,3 ± 10,7 (n=6)

Todesrate der Streptozotocin-behandelten Ratten

Kontrolle	33,3
R-Thioctsäure Diät	8,3
S-Thioctsäure Diät	50,0

Test description

Female Wistar rats (n=3-6/ group) were administered thioctic acid enantiomers per dose with feed (1.65 g/ kg feed) in separate groups for 14 weeks. At eight weeks, the animals were induced with streptoscopic diabetes, and six weeks after the induction of diabetes, the surviving animals were killed, and tissue was removed and analyzed.

R-(+) -α-lipoic acid can therefore be considered as a highly specific, effective medicine for the treatment of cataracts and nephropathy as consequential diseases and late complications of type I and type II diabetes mellitus or in tissue insulin sensitivity disorders (insulin resistance).

R- ((+) dihydroliponic acid, the metabolites e.g. bisnorilic and tetranorliponic acid and their salts, esters and amides, may also be used.

The following are therefore considered as indications for the use of medicinal products containing the substances mentioned: Subclinical and clinically manifest insulin resistance and associated diseases (compensated and decompensated insulin resistance) Cataract Nephropathies

The production of R- ((+) -α-lipoic acid, R- ((-) -dihydrolipic acid or their metabolites (e.g. bisnoro- or teranorlipic acid) and their salts, esters, amides is known (see also, for example, DE-OS 41 37 773).

The invention relates to the use of medicinal products for the treatment of the above diseases containing optically pure R- ((+) -α-lipoic acid, R- ((-) -dihydrolipic acid or its metabolites and their salts, esters, amides.

Galenic examples

The weight values given in the patent application refer to the pure optical isomer, not to the salts.

The salts are produced by a known method (see also patent No EP-A 901213405). Pharmaceutical preparations generally contain 5 mg to 3 g of the compounds of the invention in single doses. It is given as tablets, chewable tablets, aspirators, pills, capsules, granules, dredges, lozenges, lozenges, ready-to-drink solutions, liquid forms for parenteral use and aerosols.

For example, tablets containing between 10 mg and 2 g and solutions containing between 1 mg and 200 mg/ ml liquid active substance are preferred.

For example, the following are to be considered as single doses of the active substance: a. Oral forms: 10 mg to 3 g. Parenteral forms (intravenous or intramuscular): 10 mg to 12 g. Inhalation: 10 mg to 2 g.

For example, doses a) to c) may be given 1 to 6 times a day or as a continuous infusion.

Examples of implementations: Example 1: Tablets containing 100 mg of R- ((+) -α-lipoic acid

250 g R- ((+) -α-lipoic acid is mixed evenly with 750 g microcrystalline cellulose, 250 g starch (Starch 1500/Colorcon), 732.5 g lactose, 15 g magnesium stearate and 2.5 g highly dispersible silicon dioxide are added after the mixture has been agitated and the mixture is dispersed into 800,0 mg tablets. One tablet contains 100 mg R- ((+) -α-lipoic acid, which can be coated with a gastric-juice-soluble or gastric-juice-permeable film, as appropriate.

Example 2: Ampoules containing 250 mg of R- ((+) -α-lipoic acid as the salt of tromethamol in 10 ml

250 g R- ((+) -α-lipoic acid is reconstituted with 352.3 g of trometamol (2-amino-2- ((hydroxymethyl) -1.3-propandiol) in a mixture of 9 litres of water for injections and 200 g of 1,2-propylene glycol, stirred, the solution is made up to 10 litres of water for injections and then filtered through a membrane filter with a pore diameter of 0.2 μm with a glass fibre pre-filter. One ampoule contains 250 mg R- ((+) -α-lipoic acid in 10 ml solution for injection as a salt of tromethamol.

Example 3: Ampules containing 250 mg R-dihydrolipic acid in 10 ml solution for injection

60 mg of trometamol and 1 g of ethylene diamine tetracyclic acid, disodium salt, are dissolved in 1.8 litres of water for injections. The solution is soaked with nitrogen for 30 minutes. Subsequent soaking with nitrogen dissolves 2 g of sodium disulphite and then 50 g of R- ((-) dihydroliponic acid in the mixture. The solution is filled with water for injections, soaked with nitrogen, to a volume of 2 litres. After careful mixing, the solution is filtered through a membrane filter with a pore size of 0.2 μm and the filtrate is filled in 10 ml filters under aseptic conditions and pre- and post-soaked with nitrogen. One ampoule contains 250 mg R- ((-) dihydroliponic acid in 10 ml of solution as a salt of tromethamol.

Other

The following is a list of the most important and important scientific publications published by the German Academy of Sciences and Sciences:

Claims (3)

1. Use of R-(+)-α-lipoic acid, R-(-)dihydrolipoic acid or the metabolites as well as their salts, esters, amides for the preparation of a medicament for the treatment of cataract or nephropathy as sequelae or late complications of diabetes mellitus and insulin resistence.
2. Use according to claim 1, comprising a combination of R-(+)-α-lipoic acid, R-(-)dihydrolipoic acid or the metabolites as well as their salts, esters, amides with at least one other antidiabetic.
3. Use according to claim 2, wherein the other antidiabetic is insulin.