KR19980042868A - Inositol glycans with insulin-like action - Google Patents

Abstract

The present invention relates to compounds of the general formula (1). The compounds of formula (I) are suitable for the treatment of diabetes mellitus or non-insulin-dependent diabetes mellitus.

Formula 1

A-Z-R

In this formula,

A radical is HP (O) (OH) - , HP (S) (OH) -, HO-P (S) (OH) -, HS-P (S) (OH) -, (C 1 -C 4) -alkyl -P (O) (OH) - , (C 1 -C 4) - alkyl, -P (S) (OH) - , S (O) 2 (OR 1) -, S (O) (OR 1) _{-, NH 2 -C (O)} -, R 1 R 2 N-, R 1 R 2 NC (O) -NH-, R 1 O-SO 2 -NH-, (C 1 -C 4) - alkyl - SO ₂ -, (C ₁ -C ₄ ) -alkyl-S (O) - or R ¹ -S-,

Z is a substituted or unsubstituted sugar radical of 2 to 6,

R is substituted or unsubstituted inositol.

Description

Inositol glycans with insulin-like action

The present invention relates to an inositol glycan having insulin-like action, which is suitable for the treatment of diabetes mellitus.

It is known that the metabolism of insulin also induces the formation of low molecular weight compounds with insulin-like action (see U.S. Patent No. 4,446,064). A number of inositol glycan compounds having insulin-like action have already been proposed (WO 96/14075, JP 6/293790, JP 4/120089).

Diabetes Type II, non-insulin-dependent diabetes, involves insulin resistance in peripheral tissues such as muscle or adipose tissue. The reduced glucose utilization thereby is caused by glucose transport and insulin-stimulated deficiency of subsequent metabolic processes.

In an attempt to find another active compound having insulin-like action, the compounds according to the present invention have in vivo insulin-like action, exhibit excellent serum stability, and have insulin-like action in insulin-resistant tissues, Of the patients.

The present invention relates to an inositol glycan having an insulin-like action of the formula (1) and / or a physiologically acceptable salt thereof and / or a stereoisomer thereof.

A-Z-R

In this formula,

A is a radical 1) H-P (O) (OH) -,

2) H-P (S) (OH) -,

3) HO-P (S) (OH) -,

4) HS-P (S) (OH) -,

5) (C ₁ -C ₄ ) -alkyl-P (O) (OH) -,

6) (C ₁ -C ₄ ) -alkyl-P (S) (OH) -,

7) S (O) ₂ (OR &^lt; ^{1 >} ) -,

8) S (O) (OR &^lt; ¹ >) -,

9) NH ₂ -C (O) -,

10) R ¹ R ² N-,

11) R ¹ R ² NC (O) -NH-,

12) R ¹ O-SO ₂ -NH-,

13) (C ₁ -C ₄ ) -alkyl-SO ₂ -,

14) (C ₁ -C ₄ ) -alkyl-S (O) - or

15) R ¹ -S-, wherein R ¹ and R ² are independently of each other hydrogen or (C ₁ -C ₄ ) -alkyl,

Z is 1) 2 to 6 sugar radicals,

2) (2.1) methyl,

(2.2) per radical,

(2.3) disulfide radical,

(2.4) -SO ₂ -OH,

(2.5) -C (O) -NR < ¹ &^gt; R < ² &

(2.6) -C (O) - (C 1 -C 4) - alkyl,

(2.7) -P (O) (H) OH,

(2.8) -P (O) (OH) ₂ ,

(2.9) -P (S) (H) OH,

(2.10) -P (S) (OH) ₂ ,

(2.11) -P (S) (SH) (OH),

(2.12) -P (O) (OH) -O-CH ₂ -CH ₂ -NR ¹ R ² wherein R ¹ and R ² are independently of each other a hydrogen atom or (C ₁ -C ₄ ) ≪ / RTI > or < RTI ID = 0.0 >

(2.13) The glycoside bond between the 2 to 6 sugar radicals -CH ₂ - or a 1 a 2 to 6 sugar radicals optionally substituted 1-6 times by -S-,

R is 1) inositol,

2) Inositol phosphate,

3) Inositol thiophosphate,

4) Inositol cyclophosphate,

5) Inositol cyclothiophosphate,

6) (6.1) phosphate or

(6.2) a radical in the group defined under R 2) to R 5), which is monosubstituted or disubstituted independently of one another by thiophosphate,

7) (7.1) a cyclophosphate radical or

(7.2) a radical in the group defined under R 2) to R 5, which is monosubstituted by a cyclothiophosphate radical, or

8) Two adjacent OH groups

(8.1) -CH ₂ -SO ₂ -NH-.

More preferred compounds of formula (I) are those wherein A is 1) H-P (O) (OH) -,

2) S (O) ₂ (OR &^lt; ^{1 >} ) - or

_{And, - 3) NH 2 -C (} O)

Z is 1) (1.1) nose,

(1.2) Glucose,

(1.3) gluconic acid,

(1.4) galactonic acid,

(1.5) Nonsan,

(1.6) Glucose amine,

(1.7) fructose or

(1.8) 2 to 6 radicals selected from the group consisting of galactose,

2) selected from the group defined under Z (1.1) to (1.8)

(2.1) methyl,

(2.2) Mannose,

(2.3) Glucose amine,

(2.4) dianose or

(2.5) The glycoside bond between two sugar monosaccharides and glucose amines is monosubstituted to monosubstituted or monosubstituted, independently of one another, by mannose-glucose amines present between the two carbon atoms of carbon atoms 1-3, 1-2 or 1-6. ≪ / RTI > are substituted 2 to 6 sugar radicals,

R is 1) inositol,

2) Inositol phosphate,

3) Inositol thiophosphate,

4) Inositol cyclothiophosphate or

5) inositol cyclophosphate.

Particularly preferred compounds of formula (I) are those wherein A is H-P (O) (OH) -,

Z is 1) (1.1) only nose or

(1.2) 2 to 4 sugar radicals originating from the group consisting of glucose amines,

2) (2.1) Mannose or

(2.2) glucamine, and is 2 to 4 sugar radicals that are monosubstituted by a mannose,

R is 1) inositol,

2) inositol phosphate or

3) inositol cyclophosphate.

The sugar radical is understood to mean a compound derived from aldose and ketose having 3 to 7 carbon atoms and containing a D or L configuration, including amino acids or uronic acids. Examples which may be mentioned are glucose, mannose, fructose, galactose, ribose, erythrose, glyceraldehyde, sedoheptolulose, glucoseamine, galactoseamine, glucuronic acid, galacturonic acid, gluconic acid, Or nitric acid.

With respect to the disaccharide, the saccharide means two sugar units. Disaccharides, saccharides, pentoses or hexoses are formed by acetal-like bonds of 3 to 6 sugars. In this case, the bond may occur in the a- or b-form. The bond between sugars preferably occurs through carbon atoms 1 and 6 of each sugar, carbon atoms 1 and 2, and carbon atoms 1 and 4. The bond between sugars is preferably in the? -Form.

The bond of A to Z is generated, for example, by one of the oxygen atoms of Z or one of the carbon atoms of Z, preferably the carbon atom of the CH ₂ group of Z. A 10) to 12) is preferably generated by a carbon atom of Z, and the other bond of A is preferably generated by an oxygen atom of Z.

The bond of R to Z similarly occurs similarly to the binding of a disaccharide, a trisaccharide, a saccharide, a saccharide or a saccharide. In addition, the combination of R to Z are also -CH ₂ - may be substituted one or more times with a or -S-.

When the sugar is substituted, the substitution preferably occurs on the hydrogen atom of the sugar OH group.

The term insulin-resistant tissue is understood to mean, for example, rat adipocytes that do not contain any insulin receptor.

The compounds according to the invention may contain one or more phosphate groups which can be further derivatized by a phosphate protecting group. The phosphate protecting group is, for example, phenyl, benzyl or hydroxypropylnitrile (Houben Weyl, Methoden der Organischen Chemie [Methods of Organic Chemistry], Volume 12/1 or Volume 12/2; Teilheimer, Synthetic Methods of Organic Chemistry, Vol. 45].

Physiologically acceptable salts of the compounds of formula (I) are understood to mean especially pharmaceutically usable or non-toxic salts. Such salts may be formed, for example, from compounds of formula (I) containing acidic groups, such as phosphates or sulphates, with alkali metal or alkaline earth metals, such as Na, K, Mg and Ca and also physiologically acceptable organic amines , For example triethylamine and tris (2-hydroxyethyl) amine. The compounds of formula (I) containing basic groups, for example amino groups, can be prepared by reacting an inorganic acid such as hydrochloric acid, sulfuric acid or phosphoric acid and an organic carboxylic acid or a sulfonic acid such as acetic acid, to form a salt with p-toluenesulfonic acid. Compounds in which the basic and acidic groups are present in the same number form internal salts and are independent of the third salt component.

The present invention also relates to a process for the synthesis of an inositol glycan from protected saccharides and inositol precursors in a stepwise fashion, adding radical A, removing one or more protective groups that are transiently introduced for protection of other functional groups from the resulting compound, And converting the resulting compound of formula (I) into a physiologically acceptable salt thereof.

Synthesis of disaccharides or polysaccharides can be carried out by known methods (H. Paulsen, Angew. Chem. Int. Ed. 21 (1982) p. 155]. A trichloroacetimidation method is preferably used to synthesize oligosaccharides (R. R. Schmidt, Angew. Chem. Int. Ed. 25 (1986) 212-235; T. Ogawa, Tetrahedron Lett. 31 (1990) 2439-2442].

The synthesis of the phosphate is carried out using H-phosphate and phosphoramidite methods (W. Bannwath et al., Helvetica Chemica Acta, 70 (1987), pages 175-186; L. A. Slotin, Synthesis (1977), pages 737-752].

Substantially possible protecting groups for the hydroxyl groups of the sugars are: benzyl, acetyl, benzoyl, pivaloyl, trityl, tert-butyldimethylsilyl, benzylidene, cyclohexylidene or isopropylidene protecting groups.

The compounds of formula (I) and their physiologically acceptable salts are mainly used as active compounds in the manufacture of medicaments for the treatment of diabetes mellitus or non-insulin-dependent diabetes mellitus.

Accordingly, the present invention also relates to a pharmaceutical composition comprising at least one compound of formula 1 and / or one or more physiologically acceptable salts thereof in dissolved, amorphous and / or crystalline form, preferably in amorphous and / ≪ / RTI >

The pharmaceutical preparations are preferably injectable solutions or suspensions at a pH of from about 3.0 to 9.0, preferably from about 5.0 to 8.5, which may contain suitable isotonic agents, suitable preservatives and, where appropriate, suitable buffers, and, if necessary, It also contains a dropping agent. All formulation components independent of the active ingredient form the excipient. Suitable isotonic agents are, for example, for glycerol, glucose, mannitol, NaCl, calcium or magnesium compound, for example, CaCl ₂ or MgCl _2. Suitable preservatives are, for example, phenol, m-cresol, benzyl alcohol and / or p-hydroxybenzoic acid esters.

In particular, the buffer material that can be used to adjust the pH to about 5.0 to 8.5 is, for example, sodium acetate, sodium citrate or sodium phosphate. In addition, physiologically acceptable dilute acids (typically HCl) or alkalis (typically NaOH) are also suitable for adjusting the pH.

In order to differentiate the action profile of the preparations according to the invention, the modified (EP-B 132 769 and EP 132 132 770) and / or unmodified insulin, preferably bovine, porcine or human insulin, Insulin can also be mixed.

The pharmaceutical preparations can be prepared by mixing one or more compounds of formula (I) and / or one or more of its physiologically acceptable salts with physiologically acceptable excipients and, where appropriate, suitable additives and auxiliaries, if necessary, with modified and / or unmodified insulin Or an insulin derivative.

Now, the present invention will be described in more detail by the following examples.

Example 1

Preparation of Compound B

The synthesis procedure can be deduced from the reaction scheme in the late stage of Example 1.

Synthesis of Compound 3

Compound 1 60 g (94 mmol) (TG Mayer, B. Kratzer, RR Schmidt, Angew. Chem. 106 (1994) 2289-93] and 21.2 g (42.4 mmol) of compound 2 (A. Termin, RR Schmidt, Liebigs Ann. Chem. (1989) 789-795] is dissolved in 200 ml of anhydrous methylene chloride and 400 ml of anhydrous n-heptane. After 70 g of the dried molecular sieve (0.4 nm) was added, the mixture was stirred at room temperature for 15 minutes. After 15 minutes, 300 ml of n-heptane / ethyl acetate (1: 1) was added and the mixture was filtered through silica gel Filter. The silica gel is washed with n-heptane / ethyl acetate (1: 1), then the filtrate is concentrated. After purification by flash chromatography, 41.0 g (99%) of compound 3 is obtained as a colorless oil. TLC: n-heptane / ethyl acetate (1: 1), _Rf = 0.6, MS: (M + Li) ⁺ = 981.1, calculated for C ₅₅ H ₆₇ N ₃ O ₁₁ Si, M = 974.21.

Synthesis of Date 4

41 g (42.0 mmol) of Compound 3 are dissolved in 400 ml of tetrahydrofuran (THF) and 9 ml of acetic acid. 70 ml of 1 M TBAF / THF solution is added and the mixture is allowed to stand at room temperature for 8 hours. Acetic acid is removed by freezing (-30 < 0 > C for 16 h), and after concentrating the filtrate is purified by flash chromatography. 34.1 g (94%) of the deprotected compound are obtained. This is dissolved in 300 ml of anhydrous methylene chloride. After adding 50 ml of trichloroacetonitrile and 20 g of potassium carbonate, the mixture is stirred at room temperature for 4 hours. Filter through silica gel, wash the silica gel with n-heptane / ethyl acetate (1: 1) and concentrate the filtrate. Crude yield: 42.1 g. TLC: n-heptane / ethyl acetate (2: 1), _Rf = 0.5. ¹ H-NMR (CDCl ₃ ): characteristic signal for imidate; Σ = 8.78 for NH, 5.63 for anomer (β-imidate).

Synthesis of trisodium 6

33.2 g (33.0 mmol) of Compound 4 and 13.3 g of Compound 5 (R. Aneja, SG Aneja, A. Parra, Tetrahedron, Asymmetry 6 (1995) 17-18; (1990) 8243-54] are dissolved in 120 ml of anhydrous methylene chloride and 360 ml of anhydrous n-heptane. After addition of 100 g of molecular sieves, the mixture is stirred at room temperature for 15 minutes. Cooled to -20 < 0 > C under argon, and then treated with 20 ml of catalyst solution. After 30 minutes, the mixture is warmed to room temperature. For work-up, filter through silica gel and wash the silica gel with n-heptane / ethyl acetate (1: 1). The filtrate is concentrated and the crude product (46.2 g) is dissolved in 150 ml of methylene chloride. 400 ml of methanol and 15 ml of a 1 M NaOMe / MeOH solution are added and the mixture is allowed to stand at room temperature for 16 hours. The solution is treated with 1 ml of water and concentrated. The oil obtained is dissolved in 50 ml of ethyl acetate and diluted with 200 ml of n-heptane / ethyl acetate (1: 1) and then filtered through silica gel. After concentration, the residue is purified by flash chromatography. 32.5 g (97%) of a white foam are obtained as an anomer mixture. TLC: n-heptane / ethyl acetate (2: 1), _Rf = 0.5. MS: (M + Li) < ⁺ > = 1336.7; Calculated for C ₈₀ H ₈₇ N ₃ O ₁₅ , M = 1330.59.

Synthesis of trisaccharide 7

The anomeric mixture 6 can be easily separated by chromatography as the derivative 7. 32.4 g (24.4 mmol) of Compound 6 are dissolved in 200 ml of methylene chloride. 500 ml of 0.5 M HCl / MeOH (from 17.5 ml of AcCl in 500 ml of MeOH) and 20 ml of ethylene glycol are added and the mixture is allowed to stand at room temperature for 17 hours. After concentration, the residue is purified by flash chromatography. The obtained product (26.9 g, 88%) is dissolved in 300 ml of methylene chloride. 3.0 g of imidazole and 4.6 g of TBDMSCl are added. After 16 hours at room temperature, the mixture is diluted with n-heptane / ethyl acetate (2: 1) and filtered through silica gel. The silica gel is washed with n-heptane / ethyl acetate (2: 1) and the filtrate is concentrated. The obtained crude product is purified by flash chromatography. 21.1 g (72%) of compound 7 and 6.3 g (22%) of alpha product are obtained. TLC: _Rf = 0.5 for n-heptane / ethyl acetate (2: 1), compound 7, _Rf = 0.3 for alpha product. MS: (M + Li) < ⁺ > = 1370.6; Calculated for C ₈₀ H ₉₃ N ₃ O ₁₅ Si, M = 1364.71.

Synthesis of trisaccharide 8

21.2 g (15.5 mmol) of Compound 7 are dissolved in 60 ml of methylene chloride and 180 ml of dimethoxypropane. 250 mg of TsOH are added and the mixture is allowed to stand at room temperature for 1 hour. After adding 2 ml of trimethylamine, it is concentrated to obtain 23.1 g of a crude product. This is dissolved in 150 ml THF and treated with 30 ml 1 M TBAF / THF solution. After 16 h, it is concentrated and purified by flash chromatography. Yield: 20.0 g (99%) of Compound 8 as a white foam. TLC: n-heptane / ethyl acetate (2: 1), _Rf = 0.4. MS: (M + Li) < ⁺ > = 1296.7; Calculated for C ₇₇ H ₈₃ N ₃ O ₁₅ , M = 1290.51.

Synthesis of Safflower 10

20.0 g (15.4 mmol) of Compound 8 and 15.0 g (23.5 mmol) of Compound 9 (TG Mayer, B. Kratzer, RR Schmidt, Angew. Chem. 106 (1994) 2289-93] is reacted analogously to the method for compound 6 to give 10 20.3 g (76%) of the cis-saccharate as a white foam. TLC: calculated for n-heptane / ethyl acetate (2: 1), _Rf = 0.6, MS: (M + Li) ⁺ = 1770, C ₁₀₇ H ₁₁₅ N ₃ O ₂₀ , M = 1763.09.

Synthesis of octahydrate 12

20.3 g (11.8 mmol) of Compound 10 and 12.0 g (20.3 mmol) of Compound 11 (TG Mayer, B. Kratzer, RR Schmidt, Angew. Chem. 106 (1994) 2289-93] is reacted analogously to the method for compound 6 to give 19.4 g (80%) of the deacylated product. This product is dissolved in 200 ml of methylene chloride and treated with 3.4 g (50.0 mmol) of imidazole. 6.0 g (40.0 mmol) of TBDMSCl are added and the mixture is stirred at room temperature for 6 hours. After adding 5 ml of methanol, it is allowed to stand for 10 minutes, then diluted with 200 ml of n-heptane / ethyl acetate (1: 1) and filtered through silica gel. The silica gel is further washed with 200 ml of n-heptane / ethyl acetate (1: 1) and the filtrate is concentrated and purified by flash chromatography. Yield: 19.4 g (95%) of Compound 12 as a white foam. TLC: n-heptane / ethyl acetate (2: 1), _Rf = 0.7. MS: (M + Li) ^< ⁺ > = 2226; Calculated for C ₁₃₃ H ₁₅₁ N ₃ O ₂₅ Si, M = 2219.75.

Synthesis of hexose 14

19.4 g (8.9 mmol) of Compound 12 and 14.0 g (20.0 mmol) of Compound 13 (TG Mayer, B. Kratzer, RR Schmidt, Angew. Chem. 106 (1994) 2289-93] is dissolved in 100 ml of anhydrous methylene chloride and 300 ml of anhydrous n-heptane. After addition of 40 g of molecular sieve (0.4 nm), the mixture is stirred at room temperature for 15 minutes. 10 ml of the catalyst solution are added and the mixture is stirred for an additional 15 minutes. For work-up, filter through silica gel and wash the silica gel with n-heptane / ethyl acetate (1: 1). The filtrate is concentrated to give 33 g of crude product. This is dissolved in 200 ml of methylene chloride and treated with 500 ml of 0.5 M HCl in methanol. After 2 hours at room temperature, the mixture is concentrated and washed several times with methylene chloride. The resulting intermediate is dissolved in 200 ml of methylene chloride and treated with 3.4 g (50 mmol) of imidazole and 6.0 g (40 mmol) of TBDMSCl. After 16 h, the mixture is worked up analogously to compound 12 to give 20.2 g (85%) of compound 14 as a white foam. TLC: n-heptane / ethyl acetate (2: 1), _Rf = 0.3. MS: (M + Li) ^< ⁺ > = 2669; Calculated for C ₁₆₁ H ₁₇₇ N ₃ O ₃₀ Si, M = 2662.26.

Synthesis of Compound (15)

30.0 g of triazole is dissolved in 800 ml of anhydrous THF. 13.5 ml of phosphorus oxychloride is added dropwise at 10 占 폚. 60 ml of triethylamine are then added dropwise and the mixture is stirred for 15 minutes at room temperature. The precipitate is filtered off and washed with a small amount of anhydrous THF. The filtrate is added to 19.1 g (7.2 mmol) of compound 14. The solution is concentrated to 100 ml. After 15 minutes, dilute with 500 ml of ethyl acetate and wash twice with 100 ml of water. The organic phase is dried over magnesium sulfate, filtered and concentrated. After flash chromatography, 19.0 g (97%) of the cyclic phosphate derivative is obtained as a white foam. TLC: methylene chloride / methanol _{/ 33% NH 3 (100/7/1)} , R f = 0.3. MS: (M + 2 Li-H) < ⁺ > = 2737; Calculated for C ₁₆₁ H ₁₇₄ N ₃ O ₃₂ PSi, M = 2724.22. The cyclic phosphate is dissolved in 350 ml of THF and 100 ml of TBAF (1M in THF) is added. After 20 hours, the mixture is concentrated and the residue is purified by flash chromatography. 18.1 g (99%) of compound 15 as a white foam are obtained. TLC: methylene chloride / methanol _{/ 33% NH 3 (100/7/1)} , R f = 0.3 ( works the same as the starting material) MS: (M + 2Li- H) + = 2622; Calculated for C ₁₅₅ H ₁₆₂ N ₃ O ₃₂ P, M = 2609.96.

Synthesis of Compound 16

14 g of phosphoric acid are concentrated 4 times with pyridine and then dissolved in 200 ml of anhydrous pyridine. 16 ml of pivaloyl chloride is added dropwise at 10 占 폚. The reaction solution is allowed to stand at room temperature for 15 minutes. 18.1 g (6.9 mmol) of compound 15 are introduced into the above-mentioned reaction solution. After 1 hour, diluted with 200ml of toluene and methylene chloride / methanol _{/ 33% NH 3 (30/10/3)} 150ml. After concentration, the residual pyridine is further distilled three times with toluene. The residue is suspended in 200 ml of methylene chloride / methanol (20: 1). The insoluble components are filtered off and washed twice with 50 ml of methylene chloride / methanol (20: 1). The filtrate is concentrated and purified by flash chromatography. 16.9 g (91%) of the protected final product are obtained. TLC: methylene chloride / methanol _{/ 33% NH 3 (100/7/1)} , R f = 0.25. MS: (M + 3Li-2H) < ⁺ > = 2691; Calculated for C ₁₅₅ H ₁₆₃ N ₃ O ₃₄ P ₂ , M = 2673.94. 600 ml of ammonia are condensed at -78 < 0 > C for deprotection. 4.7 g (204 mmol) of sodium are dissolved therein. This solution is diluted with 300 ml of anhydrous THF and then 16.9 g (6.3 mmol) of the protected final product, dissolved in 100 ml of anhydrous THF, is slowly added dropwise at a reaction temperature of -78 < 0 > C. After 15 min of reaction (the blue color should not disappear), the mixture is carefully treated with 10 g of ammonium chloride. When the blue color disappears, the mixture is carefully diluted with 100 ml of water and 300 ml of methanol. After dissolution, it is concentrated to about 150 ml. This solution is diluted with 2 L of methylene chloride / methanol / 33% NH ₃ (3/3/1) and added to a flash silica gel column (silica gel 700 ml). Eluting with ₃ L of methylene chloride / methanol / 33% NH ₃ (3/3/2) followed by 3 L (3 / 3.5 / 3). When the product is eluted by chromatography the mixture with n- butanol / ethanol / water / 33% NH ₃ (2/2/2/1) to obtain a compound Photography 16 5.5g (78%) as a white solid. TLC: (2/2/2/1), _Rf = 0.4. MS: (M + H) < ⁺ & gt ^; = 1116.5; Calculated for C ₃₆ H ₆₃ NO ₃₄ P ₂ , M = 1115.83. ³¹ P-NMR (D ₂ O)? = 16, 3 ppm for cyclic phosphate and 7.9 for H-phosphate.

Example 2

Compounds A, B, C, D, E, F, G, H and I are prepared analogously to the method according to Example 1. Table 1 shows structural formula, empirical formula and mass spectrum.

Example 3

Pharmacological activity

The biological activity of the compounds of formula (1) according to the invention is determined using adipocytes isolated from rats.

The procedure for producing adipocytes from rats is as follows:

The white adipose tissue (Wistar rat, 160 to 180 g, without food restriction) of the solid body was digested with collagenase, the undegraded tissue was filtered to separate the separated adipose tissue, and then the Kreps-Ringer-Henseleit buffer Buffer solution) for several times.

A) Fatty formation

This test measures the insulin-stimulating conversion of glucose to a toluene-soluble product (triglyceride, phospholipid, fatty acid), which includes glucose transport and triglycerides including insulin signal transmission cascade ) / Promotes phospholipid / fatty acid synthesis. At the glucose concentration of 2.5 mM in the test, esterification (and not glycerol-3-P synthesis involving glucose transfer) is a rate-measure for stimulation of lipogenesis.

In the presence or absence of KRH buffer in the rat adipocytes 200㎕ (3 × 10 ⁵ cells / ml) of insulin for 90 min in a 37 ℃ (10ng / ml) or compound of formula I according to the invention (final volume 1ml) And incubated with 100 μl of D- [3- ³ H] glucose (25 mM, 0.4 μCi). The cells are blocked by the addition of toluene-soluble scintillation cocktail (10 ml), and the lipids are separated from the water-soluble product and the incubation medium. After phase separation, the radioactivity incorporated into the lipid product is measured directly without scraping the aqueous phase by scintillation counting ([ ³ H] lipid [dpm × 10 ^-3 ]). Control values (incubation under the same conditions without cells) are excluded from the above radiation activity values. Fat formation rate is linear until 120 minutes. The maximum stimulation rate (ie, the ratio of the results of the incubation with insulin to the results of the incubation without insulin) is taken as 100%. Percent data under% Ins _max is part of the maximum stimulus rate defined in this way. The term EC ₅₀ represents the concentration of the compound of formula (1) wherein 50% of the maximum stimulation achieved by each compound of formula (1) is observed.

B) Glucose transport

Rat adipocytes (5 x 10 ⁵ cells / ml of the activity) in 100 μl of KRH buffer are incubated with insulin or the compound of formula (1) according to the invention for 15 min at 37 ° C with gentle shaking. 15 μl of 2- [ ³ H] -deoxyglucose (0.3 mM, 0.2 μCi) was added and the mixture was incubated at room temperature. After a certain period of time (0 to 20 minutes), 100 占 퐇 of the test batch was removed, and 250 占 퐇 of dinonyl phthalate was transferred to a reaction vessel (content 400 占 퐇) into which was initially introduced. After centrifugation (15,000 x g, 1 min), the cells are removed from the oil layer of the incubation medium below the oil layer by cutting the tube at the height of the oil layer and transferring it to the scintillation vessel. After addition of 5 ml of water-soluble scintillation fluid, the radiation activity is measured. This whole cell-associated radiation activity is passively dispersed into cells and corrected for [< ³ > H] -deoxyglucose contained in the intercellular space (control temperature in the presence of the glucose transport inhibitor cytochalasin B) . The initial (stimulated) glucose transport rate is linear until 15 minutes. The maximum stimulation rate (ie, the ratio of the results of the incubation with insulin to the results of the incubation without insulin) is taken as 100%.

The terms% Ins _max and EC ₅₀ are as defined in A) Fatty formation.

The compound Fat formation Glucose transport % Ins _max EC ₅₀ [[mu] M] % Ins _max EC ₅₀ [[mu] M] HO-PO (H) O-6Manα (Manα1-2) -2Manα1-6Manα1-4GluNβ1-6 (L) Inositol-1,2- (cyclic) -phosphate A 59 7 28 8 HO-PO (H) O-6Manα1 (Manα1-2) -2Manα1-6Manα1-4GluNβ1-6 (D) Inositol-1,2- (cyclic) -phosphate B 77 8.2 41 8.1 HO-PO (H) O-6Manα1 (Manα1-2) -2Manα1-6Manα1-4GluNβ1-6 (D) Inositol-1,2- (cyclic) -thiophosphate C 64 7 29 10 HO-PO (H) O-6Manα1 (Manα1-2) -2Manα1-6Manα1-3GluNβ1-6 (L) Inositol-3-phosphate D 33 20 18 18 (2-EtNP) -4GluN? 1-6 (D) Inositol-1, 2- (cyclic) -phosphate E 48 60 34 30 HO-PO (H) O-6Manα1 (Manα1-2) -2Manα1-6Manα1-4GluNα1-6 (L) Inositol-2-phosphate F 53 15 26 25 HO-PO (H) O-6Manα1-4GluNα1-6 (L) Inositol-1,2- (cyclic) -phosphate G 25 15 9 15 HO-PO (H) O-6Manα1-4GluNα1-6 (L) Inositol H 15 25 2 10 HO-PO (H) O-6Manα1-6-Manα1-6Manα1-6Manα1-3GluNβ1-6 (L) Inositol-3-phosphate I 18 20 14 50

The compounds according to the present invention are suitable for the treatment of diabetes mellitus because they have in vivo insulin-like action and exhibit excellent serum stability and insulin-like action in insulin-resistant tissues.

Claims (9)

A compound of formula (I), a physiologically acceptable salt thereof and a stereoisomer thereof. Formula 1 A-Z-R In this formula, A is a radical 1) H-P (O) (OH) -, 2) H-P (S) (OH) -, 3) HO-P (S) (OH) -, 4) HS-P (S) (OH) -, 5) (C ₁ -C ₄ ) -alkyl-P (O) (OH) -, 6) (C ₁ -C ₄ ) -alkyl-P (S) (OH) -, 7) S (O) ₂ (OR &^lt; ^{1 >} ) -, 8) S (O) (OR &^lt; ¹ >) -, 9) NH ₂ -C (O) -, 10) R ¹ R ² N-, 11) R ¹ R ² NC (O) -NH-, 12) R ¹ O-SO ₂ -NH-, 13) (C ₁ -C ₄ ) -alkyl-SO ₂ -, 14) (C ₁ -C ₄ ) -alkyl-S (O) - or 15) R ¹ -S-, wherein R ¹ and R ² are independently of each other hydrogen or (C ₁ -C ₄ ) -alkyl, Z is 1) 2 to 6 sugar radicals, 2) (2.1) methyl, (2.2) per radical, (2.3) disulfide radical, (2.4) -SO ₂ -OH, (2.5) -C (O) -NR < ¹ &^gt; R < ² & (2.6) -C (O) - (C 1 -C 4) - alkyl, (2.7) -P (O) (H) OH, (2.8) -P (O) (OH) ₂ , (2.9) -P (S) (H) OH, (2.10) -P (S) (OH) ₂ , (2.11) -P (S) (SH) (OH), (2.12) -P (O) (OH) -O-CH ₂ -CH ₂ -NR ¹ R ² wherein R ¹ and R ² are independently of each other a hydrogen atom or (C ₁ -C ₄ ) ≪ / RTI > or < RTI ID = 0.0 > (2.13) The glycoside bond between the 2 to 6 sugar radicals -CH ₂ - or a 1 a 2 to 6 sugar radicals optionally substituted 1-6 times by -S-, R is 1) inositol, 2) Inositol phosphate, 3) Inositol thiophosphate, 4) Inositol cyclophosphate, 5) Inositol cyclothiophosphate, 6) (6.1) phosphate or (6.2) a radical in the group defined under R 2) to R 5), which is monosubstituted or disubstituted independently of one another by thiophosphate, 7) (7.1) a cyclophosphate radical or (7.2) a radical in the group defined under R 2) to R 5, which is monosubstituted by a cyclothiophosphate radical, or 8) Two adjacent OH groups (8.1) -CH ₂ -SO ₂ -NH-. A compound according to claim 1, wherein A is 1) H-P (O) (OH) -, 2) S (O) ₂ (OR &^lt; ^{1 >} ) - or _{And, - 3) NH 2 -C (} O) Z is 1) (1.1) nose, (1.2) Glucose, (1.3) gluconic acid, (1.4) galactonic acid, (1.5) Nonsan, (1.6) Glucose amine, (1.7) Fructose or (1.8) galactose, 2 to 6 sugar radicals selected from the group consisting of < RTI ID = 0.0 > 2) selected from the group defined under Z (1.1) to (1.8) (2.1) methyl, (2.2) Mannose, (2.3) Glucose amine, (2.4) dianose or (2.5) The glycoside bond between two sugar monosaccharides and glucose amines is monosubstituted to monosubstituted or monosubstituted, independently of one another, by mannose-glucose amines present between the two carbon atoms of carbon atoms 1-3, 1-2 or 1-6. ≪ / RTI > are substituted 2 to 6 sugar radicals, R is 1) inositol, 2) Inositol phosphate, 3) Inositol thiophosphate, 4) Inositol cyclothiophosphate or 5) Inositol cyclophosphate. A compound according to claim 1, wherein A is H-P (O) (OH) -, Z is 1) (1.1) only nose or (1.2) 2 to 4 sugar radicals selected from the group consisting of glucose amines, 2) (2.1) Mannose or (2.2) glucamine, and is 2 to 4 sugar radicals that are monosubstituted by a mannose, R is 1) inositol, 2) inositol phosphate or 3) Inositol cyclophosphate. Stepwise synthesis of inositol glycans from protected sugars and inositol precursors, addition of radical A, removal of one or more transiently introduced protecting groups from the resulting compounds for protection of other functional groups and, if necessary, Lt; RTI ID = 0.0 > (I) < / RTI > to a physiologically acceptable salt thereof. A pharmaceutical formulation comprising an effective amount of at least one compound of formula (I) according to any one of claims 1 to 3 in dissolved, amorphous or crystalline form. 6. The pharmaceutical preparation according to claim 5, comprising an effective amount of at least one modified or unmodified insulin or insulin derivative. A pharmaceutical formulation according to claims 5 or 6, comprising preparing one or more compounds of formula (I) in a suitable dosage form using physiologically acceptable excipients and, where appropriate, suitable excipients and / or adjuvants, How to. 8. The method of claim 7, wherein at least one modified or unmodified insulin or insulin derivative is added. Use of a compound of formula (1) as claimed in any one of claims 1 to 3 for the manufacture of a pharmaceutical preparation for the treatment of diabetes mellitus or non-insulin-dependent diabetes mellitus or obtained according to the method according to claim 4.