JPH064671B2 - Sialic acid derivative and method for producing the same - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to an immunologically active sialic acid derivative and a method for producing the same. Sialic acid is known to exist as a sialo complex (glycoprotein, glycolipid, oligosaccharide, and polysaccharide) on the cell surface of the animal kingdom or some bacteria.

In recent years, this compound has been medically and pharmaceutically important, such as nerve function, cancer, inflammation, immunity, virus infection, differentiation, hormone receptor, and the like, and is attracting attention as a specific active molecule localized on the cell surface. However, the role of sialic acid in the sialo complex is still speculative.

This compound has also been studied by many natural product organic chemists and has already led to various simple derivatives. However, a remarkable bioactive derivative is not yet known. Therefore, the present invention is to obtain a novel compound having excellent physiological activity.

Furthermore, in recent years, diversification of treatments for various cancer diseases, collagen diseases, etc., including malignant tumors of hematopoietic organs, has certainly brought about a life prolonging effect. On the other hand, however, it is unavoidable to increase the frequency of use of drugs used, such as corticosteroids and immunosuppressants, and as a result, so-called immunity is reduced / decreased and many side effects are occurring. The present inventors have focused their attention on sialic acid, which is an indigenous component of living organisms, and have conducted diligent research on an immunomodulator that has few side effects due to its chemical modification and that has a regulating action on the immune surveillance mechanism. Thus, a novel compound of the present invention having a so-called immunomodulatory effect of activating erythrocytes and suppressing B cell immunoglobulin production has been reached.

That is, the present invention has the general formula (In the formula, _one of R ₁ and R ₂ is a carboxyl group or a methoxycarbonyl group, and the other is a formula And R ³ is hydrogen or an acetyl group).

Further, the present invention has the general formula The present invention relates to a method for producing a sialic acid derivative represented by the above formula [I], which comprises reacting a compound represented by the formula (1) with cholesterol and then hydrolyzing the compound if necessary.

The sialic acid derivative of the general formula [I] is produced as shown by the following chemical formula.

The compound of the formula [II] used in the present invention is a known compound and can be easily obtained commercially.

In the above production method, the compound of formula [II] is added to Koenigs-Kn
Approximately 20 cholesterol at normal pressure in the presence of catalyst for orr reaction
The reaction is carried out at a temperature of -25 ° C for about 2-7 days. At that time, it is used in an amount of about 2 to 5 times the mole of the cholesterol compound [II].

As the catalyst, mercuric bromide, mercuric cyanide, silver perchlorate, silver trifluoromethanesulfonate, and silver trifluoroacetate sugar can be used.

The above catalyst is about 1.0 equivalent to 1.2 equivalent to 1 equivalent of the compound [II].
Use within the equivalent range.

Examples of the solvent include acetonitrile, nitromethane, acetone, benzene, tetrahydrofuran, dichloromethane, methylene chloride and the like. Above all, benzene, tetrahydrofuran and dichloromethane are preferred solvents.

The reaction product thus obtained is isolated and purified by a conventional method such as column chromatography.

Further, the above product can be hydrolyzed if necessary to convert a methoxycarbonyl group into a carboxyl group and an acetyl group into hydrogen. Hydrolysis can be carried out by a conventional method, for example, about 1 to 3N.
Can be carried out at a temperature of about 15 to 25 ° C. for about 5 to 15 hours.

In the present invention, the compounds represented by the general formula [I] each have a remarkable immunomodulating action.

The immunomodulatory effects could be confirmed by the following methods.

Effect of ConA on activation of mouse spleen lymphocytes: T cells are non-specifically activated by ConA. The effect was examined by adding the sialic acid derivative of the present invention to this reaction system. That is, splenic lymphocytes obtained from BALB / C mice (S
To PC), ConA and the compound of the general formula [I] are added respectively, and 20% is given on a microplate at 37 ° C. while giving 5% CO _2.
Cultured for several hours. To this, thymidine labeled with tritium was added, and after further culturing at 37 ° C. for 10 hours, SPC was collected and the amount of ³ H-thymidine incorporated into SPC was measured by a scintillation counter.

With respect to the compound represented by the general formula [I], promotion / enhancement of ³ H-thymidine uptake was observed, and there was an enhancing effect on T cell activation by ConA.

Effect of mouse spleen lymphocytes on immunoglobulin production: The sialic acid derivative of the present invention, which showed T cell activating effect in the above-mentioned experiment, was further examined for its effect on immunoglobulin production by measuring the number of plaque-forming cells. . First, sheep red blood cells (SRBC) and each compound of the general formula [I] were added to SPC and cultured at 37 ° C. for 5 days. SRBC and complement were added again to the resulting sensitized SPC. 37 ° C 3-12 in Cunningham chamber
PFCs (plaque forming cells) were counted after the time of culture.

Since a decrease in PFC was observed and the cell viability was similar to that of the control standard, it was confirmed that the inhibitory effect on immunoglobulin production was enhanced.

The compounds of the invention showed activity in the above two tests. That is, it is considered that immunoglobulin production was suppressed by activation of suppressor T cells.

Conventionally, in autoimmune diseases such as collagen disease, functional depression of suppressor T cells has been recognized. Therefore, the suppression T
The sialic acid derivative of the present invention having a cell activating effect is expected to be useful in clinical applications as an immunomodulator.

Hereinafter, the present invention will be described with reference to examples. These examples are merely illustrative of the invention and are, of course, not intended to limit the invention.

Example 1 Methyl 5-acetamido-4,7,8,9-tetra-O
-Acetyl-2-O- (5-cholesten-3-β-yl) -3,5-dideoxy-α-D-glycero-D-galacto-2-nonulopyranosonate synthesis Cholesterol 3.87 g (10 mmole) 20m of dry benzene
0.829 g ( ₄ mmole) of AgClO ₄ was added thereto, 2 g of drylite was further added, and the mixture was stirred at room temperature for 1.5 hours. Then, a 50 m solution of the compound of the formula [II] in methylene chloride was added, and the mixture was reacted at room temperature for 7 days.

The reaction solution was filtered through Celite, and the filtrate was dried under reduced pressure. The residue was dissolved in ethyl acetate, washed thoroughly with saturated brine, and washed with Na ₂
After drying with SO ₄ , ethyl acetate was distilled off under reduced pressure. Brown powder
4.78 g was obtained.

This was separated and purified by column chromatography (silica gel) to give the title compound α-form 244 mg (16.0%) and β-form 253 mg (16.6%).

Also, methyl 5-acetamido-4,7,8,9-tetra-O-acetyl-2,6-anhydro-3,5-dideoxy-D-glycero-D-galacto-non-2-ene-oneate 106 mg ( 11.2%).

Physical constant of the title compound Mass (EI) m / z859 (M ⁺ ), 800 (M ⁺ -59) ¹ H NMR CDCl ₃ δ (TMS) 90MHz 0.68 3H, s, CH ₃ -18 0.84 and 0.87 6H, CH ₃ -26, CH ₃ = 27 0.90 3H.d, J = 4.5Hz, CH ₃ -21 0.99 3H, s, CH ₃ -19 1.90 3H, s, NAc 2.03,2.12,2.15 12H, 3s, OAc × 4 2.60 1H, dd, J = 4.5 and 12.6Hz, 3-Heq 3.78 3H, s, COOMe 4.86 1H, m, 4-H Physical constant of β body Mass (FD) m / z860 (M ⁺ +1) ¹ H NMR CDCl ₃ δ (TMS) 90MHz 0.66 3H, s, CH ₃ -18 0.83 and 0.87 6H, CH ₃ -26, CH ₃ -27 0.90 3H, d, J = 4.5Hz, CH ₃ -21 1.00 3H, s, CH ₃ -19 1.86 3H, s, NAc 2.00,2.03,2.06 and 2.11 12H, 4s, OAc × 4 2.53 1H, dd, J = 4.5 and 12.6Hz, 3-Heq 3.78 3H, s, COOMe Example 2 5-acetamido-2-O-acetyl- (5-cholesten-3-β-yl) -3,5-dideoxy-α-D-glycero-D-galacto-2-nonulopyranosonic acid Synthesis The compound (α-form) obtained in Example 1 was added with 2 m of methanol.
Dissolved in, add 3m IN caustic soda solution to it,
Stir overnight at room temperature. 2 m of water was added to the reaction solution, neutralized with Dowe-X 50, a trace amount of precipitate was removed by filtration, and the filtrate was dried under reduced pressure. 31.4 mg (79.7%) of the title compound as a white powder.
Got

In the same manner as above, from the β-form obtained in Example 1, 30.0 mg (76.1%) of the β-form which was an isomer of the title compound was obtained.

Physical constant of the title compound ¹ H NMR CDCl ₃ δ (TMS) 90MHz 0.71 3H, s, CH ₃ -18 0.84 and 0.91 6H, CH ₃ -26 and CH ₃ -27 0.95 3H.d, J = 4.5Hz, CH ₃ -21 1.00 3H, s, CH ₃ -19 2.01 3H, s, NAc 2.43 1H, dd, J = 4.5 and 12.6Hz, 3-Heq Physical constant of β body ¹ H NMR CD ₃ OD δ (TMS) 90MHz 0.71 3H, s 0.86 and 0.92 6H, CH ₃ -26 and CH ₃ -27 0.95 3H, d, J = 4.5Hz, CH ₃ -21 1.00 3H, s, CH ₃ -19 2.00 3H, s, NAc 2.39 1H, dd, J = 4.5 and 12.6Hz, 3-Heq

 ─────────────────────────────────────────────────── ─── Continuation of front page (72) Inventor Yoshiho Shitori 3-5-24-408 Nakamachi, Musashino-shi, Tokyo (56) Reference JP-A-57-193496 (JP, A)

Claims (2)

[Claims] 1. A sialic acid derivative represented by the following general formula [I]. In the formula, _one of R ₁ and R ₂ is a carboxyl group or a methoxycarbonyl group, and the other is the following formula And R ₃ is hydrogen or an acetyl group. 2. General formula [II] A method for producing a sialic acid derivative represented by the following general formula [I], which comprises reacting a compound represented by the formula (1) with cholesterol and then hydrolyzing the compound if necessary. In the formula, _one of R ₁ and R ₂ is a carboxyl group or a methoxycarbonyl group, and the other is the following formula And R ₃ is hydrogen or an acetyl group.