JPS62273992A - Novel sialic acid derivative - Google Patents

Abstract

NEW MATERIAL:A compound expressed by formula I [AC is acetyl; X is H, OH or phenoxythiocarbonyl; R1 and R2 are carboxyl, lower alkoxycarbonyl, formula II (R3 is acetyl; R4 is carboxyl or lower alkoxycarbonyl, etc.)] and salt thereof. EXAMPLE:A compound expressed by formula III. USE:Useful as a synthetic intermediate for sialic acid-containing physiologically active substance, e.g. ganglioside, etc., or physiologically active substance. PREPARATION:An N-acetylneuraminic acid donor expressed by formula IV (Hal is halogen; R' is lower alkyl) is glycosylated with a galactose derivative in the presence of a neutralizing agent consisting of a dialkali phosphate in solvent, e.g. toluene, etc., using trifluoromethanesulfonic acid as a catalyst, preferably at -15 deg.C-room temperature for 5min-1hr to afford the aimed compound expressed by formula I.

Description

DETAILED DESCRIPTION OF THE INVENTION (Field of Industrial Application) The present invention relates to a novel sialic acid derivative useful as a synthetic intermediate for sialic acid-containing physiologically active substances such as gangliosides, and as a physiologically active substance.

(Prior Art) Ganglioside is a type of glycosphingolipid that is a major glycolipid in the organs of higher animals, is a general term for substances containing sialic acid, and is a group of substances with a rich diversity of molecular species. In recent years, it has been gaining importance as a unique physiologically active substance that is involved not only in nerve function, but also in cell differentiation, proliferation, canceration, viral infection, inflammation and immunity, and receptors for hormones and toxins.

For example, a type of ganglioside represented by the structure shown below, G Q + b, is known to induce cell differentiation and cell division in cultured neuroblastoma cells and to have nerve growth effects.

(Cal is galactose, Glu is glucose, Ne
(uAc stands for N-acetylneuraminic acid) Since ganglioside is only contained in trace amounts in biological tissue, attempts have been made to produce it by organic synthesis, but the simplest synthesis of hematocysts has been reported. It is only
Complex types of gangliosides, such as GQ,b, have not yet been successfully developed, and this is particularly due to the extremely difficult formation of the bond between the 2- and 8-positions of the sialic acid.

(Problems to be Solved by the Invention) An object of the present invention is to develop novel sialic acid derivatives that are constituents of sialic acid-containing physiologically active substances such as gangliosides and are useful as synthetic intermediates when producing these. It is about providing.

(Means for Solving the Problems) The present inventors focused on ganglioside as a physiologically active substance, and as a result of conducting intensive research on its production method, it was possible to bond a hydroxyl group with low reactivity to sialic acid. We discovered a new useful glycosylation reaction and succeeded in producing a new sialic acid y:'jX form, which is a component of gangliosides.
The invention has been completed.

The compound of the present invention is a novel sialic acid derivative represented by the following general formula.

(In the formula, Ac is an acetyl group; lower alkoxycarbonyl group, R5 is hydrogen, lower R6 and R7 are each the same or different, hydrogen or benzyl group,
R8 represents hydrogen, a lower alkyl group, a lower alkenyl group or a benzyl group) The lower alkoxycarbonyl groups of R1, R2 and R4 in the above general formula (1) are methoxycarbonyl, ethoxycarbonyl, n-propoxycarbonyl, 1so- propoxycarbonyl, n-butoxycarbonyl, 1so
-butoxycarbonyl, 5ec-butoxycarbonyl,
1 to 4 carbon atoms such as ter t-butoxycarbonyl group
means an alkoxycarbonyl group, preferably a methoxycarbonyl or ethoxycarbonyl group.

The lower alkyl groups of R5 and R8 are methyl, ethyl, n
-propyl, 1so-propyl, n-butyl, 1so-
It means an alkyl group having 1 to 4 carbon atoms such as butyl, 5ec-butyl, tert-butyl group, and methyl and ethyl groups are preferable.

Furthermore, the lower alkenyl group for R8 is vinyl.

Examples include lower alkenyl groups having 2 to 4 carbon atoms such as allyl, 1-propenyl, isopropenyl, and butenyl groups.

However, either R1 or R2 represents a carboxyl group or a lower alkoxycarbonyl group (Itl represents a group other than a carboxyl group or a lower alkoxycarbonyl group).

The sialic acid derivatives of the present invention include pharmaceutically acceptable salts of the compound represented by the general formula (1), such as alkali metals such as sodium and potassium, alkaline earth metals such as calcium and magnesium, Other examples include metal salts such as aluminum, and salts with organic bases such as ammonia.

These salts can be produced from the free sialic acid derivative of the present invention by known methods, or can be converted into each other.

Next, a method for producing the compound of the present invention will be described.

The compound of the present invention can be produced by a fr-regular glycosylation reaction using a sialic acid derivative represented by the following general formula (II) as a donor.

Ac (where Ac is acetyl 15, Hal is halogen, R'
represents a lower furkyl group similar to R5 above, the general formula ()
In I[), halogens include fluorine, chlorine, bromine,
Examples include iodine, but chlorine and bromine are preferred.

The compound represented by the above general formula (■) can be produced according to the following method.

2-chloro-N-acetylneuraminic acid compound represented by the following formula: Ac (R, Kuhn et al-+ Chew, Bar
, 99+611 (1966)) in benzene,
1,8-diazabicyclo(5,4,0
By treatment with 3-7-undecene (DBU), the following compound can be obtained with a yield of 80% or more.

Ac (2) Next, the above compound was mixed with N-bromosuccinimide (
After treatment with NBS), fractional purification is performed using a silica gel column to obtain bromohydrin with siaxial trans addition. By treating this bromohydrin with DBU in anhydrous acetonitrile at room temperature for several minutes, 90%
The following epoxy compound was obtained with the above yield. By reacting for ten minutes to several hours, the compound represented by the general formula (n) can be obtained in a yield of 95% or more.

The sialic acid derivative represented by the general formula (I[) obtained in this way has a reactivity that either does not proceed in conventional glycosylation reactions or results in very low yields even if the reaction occurs. It is very useful as a donor in glycosylation reactions because it reacts well with very low acceptor.

Therefore, extremely low reactivity acceptor sites such as the hydroxyl group at the 8-position and 9-position of neuraminic acid and the 3-position of galactose and the N-7
Binding to the 2-position of the cetylneuraminic acid donor became possible, and the compound of the present invention was successfully synthesized.

The method of this glycosylation reaction will be explained below.

The galactose derivatives and neuraminic acid derivatives to be reacted with the N-acetylneuraminic acid donor represented by the general formula (■) above can be used to convert other hydroxyl groups, carboxyl groups, etc. at the desired bonding site into acetyl groups, alkyl groups, etc. It is necessary to protect the compound appropriately with a group such as a benzyl group or a benzyl group.

As a solvent in the glycosylation reaction, toluene, benzene, or a mixed solvent of toluene and 1,2-dichloroethane can be used, the catalyst is silver trifluoromethanesulfonate, and the neutralizing agent is disodium phosphate, dipotassium phosphate, etc. Dialkali phosphate is preferred.

As for the reaction temperature, the production ratio of β-isomer is high at room temperature, but
The ratio of α-isomer increases as the temperature is lowered, so it can be set appropriately depending on the target product. Since decomposition etc. will occur, about 5 minutes to 1 hour is sufficient.

In order to reduce the 3-position hydroxyl group of the non-reducing terminal neuraminic acid of the compound of the present invention produced by the above glycosylation reaction, for example, the hydroxyl group is phenoxythiocarbonylated,
A method of reducing with tri-n-butylstannane, etc. can be used.

That is, by treating the product in acetonitrile at room temperature or with appropriate heating with phenylchlorothionocarbonate in the presence of dimethylaminopyridine for 1 hour to 1 day, the thiocarbonate form can be obtained in a yield of 80% or more. Obtainable.

Next, this thionocarbonate compound is heated under reflux for several tens of minutes with tri-n-butyl stannane in the presence of 2.2°-azobisisobutyronitrile (AIBN) in a solvent such as toluene or tetrahydrofuran. , the phenylthiocarbonyl group can be reduced with a yield of 95% or more, and the objective can be achieved.

Removal of the protecting group can be carried out according to conventional methods.
For example, debenzylation can be carried out in a suitable solvent such as methanol.
Catalytic reduction is carried out in the presence of palladium on carbon, and removal of acetyl groups is carried out using a basic catalyst such as an alkali metal alkoxide such as potassium t-butoxide or sodium methoxide in an appropriate solvent such as methanol or ethanol. can be carried out in the presence of

The compound of the present invention obtained as described above is purified by conventional means such as distillation, chromatography, recrystallization, etc.
LC1 elemental analysis, melting point measurement, specific rotation, IR, NMR,
Identification was performed using UV, mass spectra, etc.

Note that α-form and β-form were mainly distinguished and identified using NMR. Further, all specific optical rotations were measured using sodium D line.

The invention will be explained in further detail by the following examples.

(Example) Example 1゜Ac 7.7 g of the above compound was dissolved in dry benzene 70-g,
2.7-DBU was added, and the mixture was stirred at room temperature for 1 hour under an argon atmosphere. After washing with water and saturated brine, it was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The obtained crude product was purified with a silica gel column to obtain a syrup-like substance. This was crystallized from hexane-ethyl acetate to obtain compound i as white crystals.

Yield: 81% Melting point: 126-127°C (α) z+: +79.9° (C-1,3)
IR (Kllr): 3270. 1738. 1
645. 1565 cm-'FAn-Mass:
474 (M+1l) (21200w of compound i was dissolved in a mixed solvent of acetonitrile and water, 0.48+*
mol of NBS was added at room temperature and stirred for 12 minutes at 80°C. The residue obtained by concentration and drying was applied to a silica gel column, the percentage of bromohydrin in the siaxialtrans adduct eluted after the diequatorial adduct was collected, and the bromohydrin was recrystallized from hexane-ethyl acetate to give 7% of the bromohydrin.
Obtained with a yield of 3%.

Dissolve 500 mW of this bromohydrin in 4-molecular anhydrous acetonitrile, add 0.16 mW of DBU, and
Stir for a minute. The crude product obtained by concentration under reduced pressure was purified with a silica gel column, and then recrystallized from hexane-ethyl acetate to obtain compound ii as white crystals.

Ac Yield: 92% Melting point: 17? -178℃ ((r) ”: -10,0°(C-1,3)
IR (Kllr): 3420. 1740. 1
650. 1572 cs-'FAB-Mass:
Compound 11 of 490 (M+Il)+31200■
In anhydrous 1,2-dichloroethane solution containing 3-, 0.4
5s+wol of titanium tetrabromide was added and stirred at room temperature for 10 minutes. The residue obtained by condensation under reduced pressure was dissolved in ethyl acetate, washed with saturated sodium sulfate solution, 5% sodium bicarbonate, and saturated brine, and then dried over anhydrous sodium sulfate. The syrupy crude product obtained by concentration under pressure was purified with a silica gel column to obtain compound iii.

AC Yield: 98% Melting point: Oil [α)”: -91,1@(C・0.6)I R
(KBr): 3420.1?42.1660.15
40 cm-'FAR-Mass: 570.57
2 (M+II) Example 2゜(Compound iii of 1140011f, the following compound iv of 330* and 360■ of anhydrous sodium hydrogen phosphate were dissolved in a mixed solvent of anhydrous 1,2-dichloroethane/toluene, and 180W of trifluoromethane was dissolved. Three anhydrous toluene solutions containing silver sulfonate were added to the solution at 0°C under an argon atmosphere. The reaction solution was stirred at 0°C for 10 minutes, then at room temperature for 10 minutes, and filtered to remove insoluble matter. Washed with chloroform. After distilling off the solvent from the filtrate, the resulting syrupy product was applied to a silica gel column and separated and purified into two fractions. Analyzed by NMR ratio, the latter eluted fraction was separated into α-form ( Compound 1α-■), and the first half elution fraction was identified as the β form (compound lβ-■).

~Compound 1α-■~ Yield: 42% Melting point: 137-139°C [α] -Coni+29.9'' (C,1,2)
IR (Br): 3430. 1?40. 166
0. 1540 elm-'FAB-Mass:
921 (M+II) ~ Compound lβ-Φ ~ Yield: 21% Melting point: Oil [α)'': +23.9° (C ~ 1,2) IR (KB
r):3420.1745.1660.1540c
m-'FAB-Mass: 921 (M+II) (
2) Using the following compound: Ac instead of compound iv, the same operation as in (1) above was performed, and further separation and purification by high performance liquid chromatography was performed to obtain compound 2.
α-■ and compound 2β-■ were obtained.

~Compound 2α-■~ Melting point: Oil [α)”: +27.4° (C・1.1) E R
(KBr): 3430.1?40.1660.154
0 cll-'FAIL-Mass: 921 (M+
ll) ~Compound 2β-■~ Yield: 28% Melting point: Oil [α)'': +20.3° (C=1.6) I R
(KrJr): 3420.1745.1660.1
538 cm-'FAB-Mass: 921 (
M+l1) (3) Using the following compound instead of compound iv, perform the same operation as in (2) above to obtain compound 3α-
(2) and compound 3β-■ were obtained.

~Compound 3α-■~ Yield: 37% Melting point 2 Oil #R product [α)”: -19,7° (C-1,0) IR (
to [lr): 3430.1740.1660.15
40 ell-'FAII-Mass: 865
(M+Il) ~ Compound 4β-Φ ~ Yield: 15% Melting point: Oil (α) ”: -23,3° (C, 1,2) I R
ffflr) knee 3430.1740.1660.1
540 cts-'F All-Mass: 86
5 (M+II) (4) Using the following compound instead of compound iv, the same operation as in +11 above was performed to obtain compound 4α-■ and compound 4β-■.

~Compound 4α-■~ Yield: 18% Melting point - Oil [α) + 3. -44°(C,0,9)I R(Kfl
r): 3425.1?47.1660.1540
on-'FAfl-Mass: 1323 (
Mill) ~ Compound 3β-Φ ~ Yield: 31% Melting point: Oil [α)'': -5,8° (C-1,1) I R (KB
r): 3440.1?45.1665.1540
am-'FAR-Mass: 1323 (Mil
l) Example 3 Compound 1α-■ of 75■, 4.4-dimethylaminopyridine of 40■ and phenylchlorothionocarbonate of 23μE were dissolved in 1.0-hydroacetonitrile, and the mixture was dissolved under argon atmosphere. After stirring at 50° C. for 2.5 hours, the reaction solution was concentrated to dryness. Water and ethyl acetate were added to the residue for extraction. The ethyl acetate layer was separated, washed with saturated brine, dried over anhydrous sodium sulfate, and the solvent was distilled off. The residue was applied to a silica gel column, and the obtained crude crystals were recrystallized from ethanol/acetone to obtain compound 1α-■
A compound 1α-■ in which the hydroxyl group at the 3-position was phenoxythiocarbonylated was obtained as white crystals.

~Compound 1α-■~ Yield: 91% Melting point: 207-209 °C [α)”? +35.3 ° (C-1
, 3) I R (WBr): 3420.1745.1
660.1540 cs-”FAR-Mass:
1057 (M+H) In the same manner as above, the following compounds were obtained by phenoxythiocarbonylating the 3-hydroxyl group of Compound 1β-■, Compound 2α-■, and Compound 2β-■.

~Compound lβ-■~ Yield: 83% Melting point 2 Oil [α)'': +35.4° (C 1.3) I
R (KBr): 3420. 1750. 166
0. 1540 cm-heavy FAB-Mass:
1057 (M+H)~Compound 2α-■~ Yield: 83% Melting point 2 Oil (α)'': +35.4° (C-1,3)IR(
KBr): 3430.1?43.1660.154
0 cm-'FA [l-Mass: 1057 (
Mill) ~ Compound 2β-■ ~ Yield: 80% Melting point 2 Oil (α) ”: +22.9° (C-0,8) I R (
X[lr): 3420.174B, 1660.1
530 cm-'FAB-Mass: 1057
(M 10 amounts ■) Example 3 Compound 1α-■ of 70* was dissolved in a mixed solvent of anhydrous toluene 1.5- and tetrahydrofuran 1.0-
A reaction solution containing 1 liter of tri-n-butylstannane and a catalytic amount of AtBN was heated under reflux at 110° C. for 10 minutes, and then concentrated. The resulting syrup-like product was purified with a silica gel column, then crystallized from hexane/ethyl acetate, and washed with hexane to obtain Compound 1α-■, which is a reduction at the 3-position of Compound 1α-■, as a white powder. .

~Compound 1α-■~ Yield: 97% Melting point: Oil (α)”: +17.8° (C-2,8)t R(
KBr): 3430.1?44.1660.154
0 am-'F An-Mass: 905 (M
÷11) Similarly, compound lβ-■, compound 2α-■
The following compound was obtained by reducing the 3-hydroxyl group of compound 2β-■.

〜Compound lβ-■〜 Yield: 96% Melting point: Oil (α) ”: +35.1°(C-1,3)rR(K
Br): 3430.1742.1660.1540
am-'FAR-Mass: 905 (M
ill) ~Compound 2α-■~ Yield: 95% Melting point: Oil (α) ”: +22.3”(C,0,4)r
RO[Br): 3420.1?42.1660.1
538 co+-'FA [l-Mass: 905
(Mill) ~Compound 2β-■~ Yield: 96% Melting point: Oil (α)'': + 31.2' (C=1.2)
IR (Kllr): 3400. 1?45. 1
662. 1538 am-'FA [l-Mass:
905 (Mill) Compound 1α-■ of Example 4°45■ was added to 4.5. , i dissolved in anhydrous methanol and to which a catalytic amount of potassium t-butoxide was added under argon, the reaction solution was stirred at room temperature for 30 minutes, 0.6-IN sodium hydroxide solution was added, and The reaction solution stirred for 1 hour was cooled to -10°C and DOW
After acidification with EX 5011-X8, the resin was washed thoroughly with methanol/water. The solution was concentrated, and the resulting syrupy product was crystallized from chloroform/methanol to obtain compound 1α-■ as a white powder.

~Compound 1α-Φ~ JM Yield: 97% Melting point: 144-146°C [α)”: +12.7° (Cd, 4)r R(
KBr): 3400.1?19.1640.156
0 am-'FAIL-Mass: 583 (M
ill) Similarly, compound 1β-■, compound 2α-■
and preservation of compound 2β-■! The following compound was obtained by removing the i group.

~Compound 1β-■~ Yield: 96% Melting point: 168-172°C [α) ”: +6.6' (C,0,6)I
R (Kllr): 3390.1?20.1640.
1558 ctm-'FA [l-Mass: 58
3 (Mill) ~ Compound 2α-■ ~ I Yield: 97% Melting point: 163-166°C (α) ”: +35.7” (C
Fable 0.1) I R (Kllr): 2400.171
B, 1640.1560 cm-'FA[l-Ma
ss: 583 (Mill) ~ Compound 2β-■ ~ Yield: 93% Melting point: 232-235°C (α)”: +42.6° (C-1,1)I R
(K[lr): 3400. 1?20. 1630
．． 1570 ell-'FA[l-Mass:
583 (Mill) (Effects of the Invention) Compounds 2α-■ to 2α-■, compound 3α and compound 4α in the present invention are useful as intermediates for the synthesis of ganglioside components.

In addition to gangliosides, there are many substances containing sialic acid, such as polysancharide C of Neisseria meningitidis. Polysancalide C can prevent bacterial meningitis through sensitization through its administration.

The compounds of the present invention 1α-1 to 1 are the 2- and 9-positions of sialic acid.
It is a sialic acid dimer with a bonding mode of 1-position.

This is a component of polysaccharide C of Neisseria meningitidis and is useful as a synthetic intermediate for polysaccharide C.

Compounds of the present invention 1α-■, 1β-■, 2α-■, 2β-■
etc., there is a double bond between the 2- and 3-positions of the sialic acid on the reducing end side, so it passes through the epoxy form again and becomes a 2-halogeno-3-hydroxy form like compound Glycosylated with sialic acid etc.
Furthermore, the sialic acid derivative of the present invention can be used as an immunomodulatory agent, an autoimmune disease treatment agent, a meningitis treatment agent, and a platelet in view of the physiological activity of naturally occurring sialic acid-containing substances. It can be expected to be useful as an inhibitor of i-concentration, an anticancer agent, an antagonist of cholera toxin, an agent for restoring nervous tissue function, a cell surface marker, and the like.

Claims (1)

[Claims] (1) General formula (I): ▲Mathematical formulas, chemical formulas, tables, etc.▼(I) (In the formula, Ac is an acetyl group, X is hydrogen, hydroxyl group, or phenoxythiocarbonyl group, and R_1 and R_2 are each different from carboxyl R_3 represents hydrogen or acetyl R_4 is a carboxyl group or a lower alkoxycarbonyl group, R_5 is hydrogen, a lower alkyl group, a benzyl group, or ▲There are mathematical formulas, chemical formulas, tables, etc.▼, and R_6 and R_7 are each the same or different and represent hydrogen or benzyl. group, R_8 represents hydrogen, a lower alkyl group, a lower alkenyl group, or a benzyl group) and a pharmaceutically acceptable salt thereof.