JPS61275299A - Deoxymuramyldipeptide derivative - Google Patents

Abstract

NEW MATERIAL:A compound shown by the formula I (Ala is alanine residue; R1, R2 and R4 are acryl; R3 is H or alkyl) and its salt. EXAMPLE:N-[2-O-(2-Acetamido-1, 5-anhydro-2, 3-dideoxy-6-O-tetradecanoyl-D-glu citol-3-yl)-D-lactoyl]-L-alanyl-D-(N'-tetradecanoy-l)-isoglut amine methyl ester. USE:A preventive and remedy for infection with microorganisms and an anticancer. Having improved adjuvant activity and low heat generation. PREPARATION:A raw material compound shown by the formula II is dissolved in a dried pyridine, etc., and reacted with a fatty acid chloride. The raw material compound is obtained by using a compound shown by the formula III as a starting raw material according to the reaction formula.

Description

Detailed Description of the Invention <Industrial Application Field> The present invention relates to novel muramyl dipeptide derivatives having excellent immune adjuvant activity and microbial infection preventive/therapeutic effects, more specifically, a novel muramyl dipeptide derivative having the general formula (I) (wherein A1m is Alanine residues are R1, R2, and R4
means an acyl group, and 几 means a hydrogen atom or an alkyl group. ) and its salts 0 <Problems to be solved by the invention> The present inventors have made extensive efforts to study compounds that have excellent preventive and therapeutic effects on microbial infections.

The compound of formula (1), which is synthesized by the minimum structural unit of the cell wall of Mycobacterium tuberculosis, BOG, other mycobacteria, and cell-parasitic bacteria, and further deoxylated at the 1-position of the sugar, has an excellent preventive and therapeutic effect on microbial infection and is an adjuvant. The present invention was completed by discovering that it has activity and significantly reduced exothermicity.

<Configuration of the Invention> That is, the present invention relates to the compound of formula (I) and its salt. Examples of the alkyl group include methyl, ethyl, isopropyl, and butyl groups.

Acyl groups include acetyl, propanoyl, butyryl,
Examples include octanoyl, nonanoyl, tridecamyl, hexadecamyl, heptadecanoyl, eicosanoyl, pivaloyl, imbutyryl and invaleryl groups, etc.
For R1 and ~, intermediate to higher acyl groups are generally preferred, and for general, lower to intermediate acyl groups are generally preferred.

The compound of formula (I) of the present invention can be produced by the method shown in the following reaction formula.

(In the formula, R2, Rs, R4 and Alm are the same as above.

) That is, the desired compound of formula (I) can be produced by dissolving the compound of formula (n) in a solvent such as dry pyridine and reacting it with a fatty acid chloride.

The compound of formula (n) as a raw material can be produced by the method shown in the reaction formula below.

(In the formula -B-z, Rs, R4 and Ala are the same as above. That is, the compound of formula 9) is dissolved in a solvent such as ethanol, and a catalyst such as Raneyi or Kel is added for catalytic reduction. A compound of the formula (pieces) can be obtained.

This is dissolved in a solvent such as 1,4-dioxane, and an alkali such as potassium hydroxide is added for hydrolysis.The obtained compound is dissolved in a solvent such as 1,4-dioxane to obtain N-hydroxysuccinimide. The compound of formula (v) can be produced by reacting with dicyclohexylcarbodiimide or the like to obtain an active ester, followed by reaction with alanyl isoglutamine trifluoroacetate and triethylamine. The compound of formula (6) can be produced by dissolving this in a solvent such as a mixture of anhydrous pyridine and anhydrous dichloromethane, adding dimethylaminopyridine, and reacting with fatty acid chloride. The desired raw material compound of formula (1) can be obtained by deketalizing the obtained compound in a solvent such as 80% aqueous acetic acid and then reacting with a fatty acid chloride in a solvent such as dry pyridine.

<Effects of the Invention> It was confirmed that the compound of the present invention exhibited excellent infection prevention effects and adjuvant activity, and was significantly attenuated in its 9 pyrogenic effects. Therefore, the compounds of the present invention can be used as agents for preventing and treating microorganism infections such as bacteria and viruses, and as anticancer agents for diseases caused by decreased immune function, and can also be used as adjuvants for vaccines.

<Examples> The present invention will be further explained in detail by reference examples, examples, and test examples, but these are not intended to limit the present invention.

Reference Example 1 (1) 2-acetamido-1,5-anhydro-2-deoxy-4,6-Q-isopropylidene-8-〇-(D
-1-(methoxycarbonyl)ethyl]-D-glucitol 2-acetamido-1-8-acetyl-2-deoxy-
4,6-0-isopropylidene-δ-〇-(D-1-(
Dissolve 1 g of (methoxycarboxol)ethyl)-1-thio-β-kuglucobylanose in 10-ethanol, add Raneyi Kel-W-21,02, and stir while introducing hydrogen gas for 5 hours. Column chromatography (Wako Gel 0200 (manufactured by Wako Pure Chemical Industries, Ltd.) > 209.
Effluent; a: chloroform, b: chloroform-methanol-ye2o: 1)) to obtain the title compound (compound A) 550119.

Calculated value 054.37. H7, 61, N4.28 analysis value 054.29. H7,60,N4.35(2)N-(
2-0-(2-acetamido-1,5-anhydro-2
,8-dideoxy-4,6-0-inpropylidene-D
-glucido-8-yl)-D-lactoyk]-L-
7 Raniru D-inglutamine methyl ester compound A100r11g 1,4-dioxane 5tLt
and 0.2N potassium hydroxide aqueous solution 5- was added.

React for 5 minutes at room temperature. after the reaction is complete)
IRC! After neutralizing with -50 (H") and separating the resin, 1.
Wash with 4-dioxane. 1.
50 mg N-hydroxysuccinimide and 1 dicyclohexylcarbodiimide dissolved in 4-dioxane 5-
00119 was added and stirred for 80 minutes to form an active ester, followed by 180 μl of L-alanyl-D-isoglutamine trifluoroacetate and 0.0 μl of triethylamine.
Add 06- and react for 300 minutes. After the reaction is completed, the dicyclohexylurea produced is separated and washed with 1,4-dioxane, and the P solution and washing solution are combined and concentrated under reduced pressure. The obtained silica powder is dissolved in chloroform, washed with water, dehydrated with sodium sulfate, and concentrated. The product was purified by column chromatography (Wako Gel 0-200209 effluent TFL;
a: Chloroform-methanol (100:1), b
: chlorotetraforsimethanol (50:l), 0:chloroform-methanol (30:1)), and the title compound (compound B) la was obtained from the effluent C.

■ is obtained as a syllabary.

[α) +15° (C zero 3.2. Chloroform) Elemental analysis 0.3H and N40. . Calculated value: 052.0? , H7, 22, N10.56 analysis value 052.21. H7, 18, N10.82 (3
)N-(2-0-(2-acetamido-1,5-anhydro-2,3-dideoxy-4,6-0-isopropylidene-D-glucido-8-yl)-D-lactoyl)-L -Alanyl-D-(N'-decanoyl)-isoglutamine methyl ester compound B420■ was dissolved in anhydrous pyridine 2.8- and anhydrous dichloromethane 12-, dimethylaminopyridine 481119 was added, and the reaction solution was heated to -30°C. While stirring, 4595 g of decanoyl chloride was added to 5 fI of dichloromethane.
7! A solution dissolved in is added dropwise and stirred at room temperature for 12 hours. The silica obtained by concentration under reduced pressure was subjected to column chromatography (Wakogel 0-200 209, effluent; a:
The title compound (compound C) was obtained from the effluent C.

277119 is obtained as Shirapu.

[α]D+14° (0 day 0.4.Chloroform) Elemental analysis Calculated value as 0saH5IIN4On 057.
88. H8, 24, N8.18 analysis value 057.99
．． H8,36,N8゜01(4)N-(2-0-(
2-acetamido-1,5-anhydro-2,3-dideoxy-D-glucido-3-ylto-D-lactoyl)-L-alanyl-D-(N'-decanoyl)-isoglutamine methyl ester compound 0160■ Dissolved in 80% acetic acid aqueous solution 10-
Heat at 45°C for 8 hours. After the reaction was completed, it was concentrated under reduced pressure and crystallized from ether to obtain the title compound (compound D) 15.
0■ is obtained quantitatively.

Melting point 145-146°C [α] D + L9° (G7 = 1.0. Chloroform-methanol (1:1)) Elemental analysis Calculated value as 030H, N, O, 055
．． 89. H8,13,N8.69 analysis value C54,0
5, H8,23, H8,63 Reference Example 2 (1) N -(2-0-(2-acetamido-1,5
-anhydro-2,3-dideoxy-4,6-0-inpropylidene-D-glucido-3-yl)-D-lactoyl]-L-alanyl-D-(N'-tetradecanoyl)-iso Glutamine methyl ester compound E 4 s o Tn 9 and tetradecanoyl chloride 600''9 are reacted in the same manner as in Reference Example 1 (3) to obtain 300'' of the title compound (compound B) as a silica.

[α) +a, o' (c=o, a, chloroform) Elemental analysis Calculated value as C5tHa+N+0++ 0
59.98. H8°71. H7,56 analysis value 059
．． 69. H8,95,H7,40(2) N-(2
-0-(2-acetamido-1,5-anhydro-2,
3-dideoxy-D-glucido-3-yl)-D-
Lactoyl)-L-alanyl-D-(N'-tetradecanoyl)-isoglutamine methyl ester compound E15
0■ was reacted in the same manner as in Reference Example 1 (4) to quantitatively obtain 140 m9 of the title compound (compound F). Melting point 149-1
51℃ [α) + & 9° (0 = 1.4.Chloroform-methanol-
(1et3) Elemental analysis Calculated value as CuH1+oNaOo 058
．． 27. H8, 63, H7, 99 analysis value 058.1
? , H8,8B, H7,69 Reference Example 3 (1) N-(2-0-(2-acetamido-1,5-anhydro-2,8-dideoxy-4,6-0-isopropylidene-D-glucide) -8-yl)-D-lactoyl]-L-alanyl D-(N'-octadecanoyl)-isoglutamine methyl ester compound B410 and octadecanoyl chloride 700
$119 was reacted in the same manner as in Reference Example 1 (3) to obtain the title compound (compound G) 322.

[α) D+15.5° (0=0.6.Chloroform) Elemental analysis 041HT! Calculated value as N40!1 061
．． 78. H9, 19, H7, 03 analysis value 061.5
8. H9,32,H6,88(2) N-(2-0
-(2-acetamido-1,5-anhydro-2,3-
Dideoxy-D-glucido-3-yl)-D-lactoyl)-L-alanyl-D-(N'-octadecanoyl)-isoglutamine methyl ester compound 0120!
'19 was reacted in the same manner as in Reference Example 1 (4) to produce the title compound (
Compound H) 1101119 is obtained quantitatively. melting point 14
5-146°C [α) +27.8° (0-0,9, rioo form)
Elemental analysis Calculated value as 0saH6aNcOn 060
．． 29. H9,05, H7,40 analysis value C60,2
5,H9,12,H7,41 Example I N-(2-0-(2-acetamido-1,5-anhydro-6-0-decamyl-2,3-dideoxy-D-glucid-8-yl )-D-lactoyl]-L-alanyl-D-(N'-decamyl)-isoglutamine methyl ester Compound D 51.7 ml was dissolved in anhydrous pyridine 3-.

A solution prepared by dissolving 18ff19 of decanoyl chloride in 1-dichloromethane was added dropwise at -25°C, and the mixture was stirred at room temperature for 12 hours. The reaction solution was concentrated under reduced pressure and subjected to separation using thin layer chromatography (Kiesel Gel T+ 6 F 254 (manufactured by Merck & Co.)).
Separation and purification using chloroform-methanol (5:l) yielded 42.3m9 of the title compound as a silica.

■R,,fi1m,,-1 3300-3250(OH
, Nu) ax 2920 and 2850 (methyl, methylene) 1785 and 1240 (ester) 1600, 1540 and 15ao (amide) [α) D-
6,6° (0=0.7.Chloroform) Elemental analysis 0,
0R7ON, O,, calculated value 060.13. H
8,83,H7,01 analysis value 060.02. H8,
99, N? , 24 Example 2 N-(2-0-(2-acetamido-1,5-anhydro-2,3-dideoxy-6-〇-tetradecanoyl-D-glucido-3-yl ID-lactoyl) - L
-Alanyl-D-(N'-tetradecanoyl)-isoglutamine methyl ester Compound F70WI9 and tetradecanoyl chloride are reacted in the same manner as in Example 1 to obtain the title compound 55. Melting point 126-129℃ 2920 and 2840 (methyl, °methylene) 173
5 and 1240 (ester) 1655, 1540 and 1530 (amide) [α)-
8,2° (0=0.1.Chloroform) Elemental analysis 04
8H1i6N401! Calculated value as 063.27.
l-19,51,N6.15 analysis value 063.05.
H9,73,N6.08 Example 3 N-(2-0-(2-acetamido-1,5-anhydro-2,8-dideoxy-6-〇-octadecanoyl-D-glucide-3-yl )-D-lactoyl)-L
-Alanyl-D-(N'-octadecanoyl)-isoglutamine methyl ester compound H69,8! "9 and octadecanoyl chloride are reacted in the same manner as in Example 1 to obtain the title compound 51rn9. Melting point 54-56°C (decomposition) 2920 and 2850 (methyl, methylene) 1740 and 1250 (ester) 1660, 1550 and 1530 ( Amide] [α) −
12,0° (0=0.2 chloroform) elemental analysis
0511H+02 Calculated value as N4012 C! 65
．． 72. Hlo, 05. N5.47 analysis value 065
．． 45. HlO, 12, N5.45 Test Example The effect of the compound of the present invention was confirmed by the method described below.

(1) Effect on preventing microbial infection The compounds obtained in each example were suspended in phosphate buffered saline at a concentration of 0.45 cm/-. This was subcutaneously administered to 12 to 19 8td-ddY mice (body weight: 18.7 to 80.59.5 weeks old) per group, and 24 hours later, 5.8f of B. eoli 177156 strain was administered. ! Infect subcutaneously at the number of XIO' bacteria/the amount of inoculated bacteria per mouse. Efficacy is determined by mouse survival rate 7 days after infection (4)
Find from. The results are shown in Table 1. As is clear from Table 1, the compounds of the present invention exhibited excellent infection prevention effects.

Table I E, protective effect in colf-infected mice (2
) Immunological adjuvant activity The immunoadjuvant activity of the compound of the present invention is as follows: N-acetyltyrosine, -8-azobenzene-4 in 1 guinea pig
'-Arsonic acid (hereinafter referred to as AEA-N-Ae-Tyr)
This was confirmed by the effect of enhancing delayed allergic expression.

That is, ABA-N-Ae-Tyr (50 μg per animal)
) and the compounds obtained in each example (100
A water-in-oil (Wlo) emulsion is prepared by mixing 70 μg) with an incomplete adjuvant. Administer this to guinea pigs. After 2 weeks, ABA-N-Ac-T
1001tF, a conjugate of yr and bovine serum albumin, is injected into the skin, and the diameter of the skin reaction (erythema and induration) is measured 24 hours later. The size of this diameter is considered a measure of cell-mediated immunity.

The results are shown in Table 1 below.

Table 2 Adjuvant activity against delayed allergic manifestation □ (3) Pyrogenic substance test

Claims (1)

[Claims] Compounds represented by the general formula ▲ Numerical formulas, chemical formulas, tables, etc. ▼ and their salts (In the above formula, Ala represents an alanine residue, R_1, R_2,
and R_4 means an acyl group, and R_3 means a hydrogen atom or an alkyl group. )