JPH0153848B2 - - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to the use of novel compounds having immunostimulatory activity. The inventor has the general formula A compound represented by and a pharmacologically acceptable salt,
We also succeeded in producing a hydrate of the compound or a salt thereof. In the formula, R represents a hydrogen atom, a linear or branched lower alkyl group such as a methyl group, an ethyl group, or a propyl group. Furthermore, as a result of examining the usefulness of these compounds as medicines, it was discovered that this substance has an immunostimulatory effect. That is, the present invention provides the general formula (In the formula, R represents a hydrogen atom or a linear or branched lower alkyl group) and a pharmacologically acceptable salt thereof, and an immunostimulant containing the hydrate of the compound or a salt thereof as an active ingredient and immunosuppressive cancer drugs. Representative compounds of the present invention included in formula () are:
For example, 3-hydroxy-4-(hydroxymethyl)benzoic acid, 3-hydroxy-4-(hydroxymethyl)benzoic acid methyl ester, 3-hydroxy-4-(hydroxymethyl)benzoic acid ethyl ester, and pharmacologically acceptable and hydrates of the compound or salts thereof. Next, a method for producing the compound of the present invention will be described. The compound of the present invention has the general formula (In the formula, R ¹ is a straight-chain or branched lower alkyl group,
^R2 represents a hydrogen atom or a linear or branched lower alkyl group), by selectively reducing the carboxylic acid ester adjacent to the hydroxyl group to a hydroxymethyl group. Representative examples of this raw material compound () include hydroxyterephthalic acid dimethyl ester,
Hydroxyterephthalic acid diethyl ester is mentioned. The compound of formula () is prepared by reacting the starting compound () with sodium borohydride, sodium dihydrobis-(2-methoxyethoxy) under appropriate reaction conditions.
It is obtained by a reduction reaction using a complex metal hydride such as aluminate. As a typical example of this reaction, when sodium borohydride is used as a reducing agent, a lower alkanol such as methanol or ethanol is used as a solvent, and the reaction is carried out at room temperature or under heating. In addition, when using sodium dihydrobis-(2-methoxyethoxy)aluminate as a reducing agent, use an organic solvent that does not adversely affect the reaction, such as benzene or toluene, and conduct the reaction under ice cooling or at room temperature. Manufactured. The compound according to claim 1, in which R represents a hydrogen atom in formula (), that is, 3-hydroxy-4-(hydroxymethyl)benzoic acid,
When a compound in which R ¹ is a straight-chain or branched lower alkyl group and R ² is a hydrogen atom is used as a raw material compound in formula (), by reacting the complex metal hydride under appropriate conditions. However, when a compound in which R ¹ and R ² in formula () are both straight-chain or branched lower alkyl groups is used as a raw material compound, the complex metal hydride is first reacted under appropriate conditions. It can also be obtained by selectively reducing the ester adjacent to the hydrogen group to a hydroxymethyl group, and then hydrolyzing the remaining ester to a carboxylic acid. This hydrolysis reaction can be carried out by any known and commonly used method, but preferably in a mixed solvent of water and a water-miscible organic solvent such as methanol, or in an alkali such as sodium hydroxide or barium hydroxide. It is carried out using a metal or alkaline earth metal hydroxide at room temperature or under heating. general formula (In the formula, R represents a linear or branched lower alkyl group) The compound represented by the general formula (In the formula, R ¹ and R ² both represent a linear or branched lower alkyl group) When using a compound represented by the above formula, it is produced by selectively reducing the ester adjacent to the hydroxyl group. However, as a raw material compound, the following formula When a compound represented by is used, it can also be produced by esterifying a carboxylic acid. That is, the following equation By esterifying the carboxylic acid of the compound represented by the general formula A compound represented by the formula (wherein R represents a linear or branched lower alkyl group) can be produced. For this esterification, all known and commonly used methods can be used. The compound () of the present invention has an immunostimulatory effect, and its details will be described using a compound (BF-127) in which R represents a hydrogen atom as a representative example. (1) Effect of BF-127 on cell-mediated immunity Delayed-type hypersensitivity obtained by inoculating mouse footpads with sheep erythrocyte antigen and immunizing with it. We examined its effectiveness using DTH (abbreviated as DTH) as an indicator.
The DTH reaction was performed by Lagrange et al. (PH Lagrange, G.
B. Mackaness and TEMiller: J. Exp. Med.
139 1529-1539, 1974). ¹⁰⁸ sheep red blood cells were suspended in 0.05 ml of physiological saline and injected subcutaneously into one footpad of an ICR female, 6-week-old mouse, and each amount of BF-127 substance was injected into 0.25 ml of physiological saline. It was dissolved in water and injected intraperitoneally. Four days later, a DTH response was induced by subcutaneously injecting ¹⁰⁸ sheep red blood cells into the other footpad, and 24
After a period of time, the degree of swelling observed in the footpad (thickness of the footpad) was measured using calipers to determine the effect (Table 1). [Table] Footpad thickness of untreated mice As shown in Table 1, 1 mg to 0.1 μg/mouse
Administration of BF-127 substance enhanced the degree of DTH response by 58% to 134% compared to the control. Such a potentiating effect is also observed when this substance is administered orally, as well as by intravenous and subcutaneous injections; examples of oral administration are shown below. ICR, female, 6-week-old mice were immunized with sheep red blood cells as described above and treated with 1 mg, 100 μg, and
10 μg of BF-127 substance was dissolved in 0.25 ml of physiological saline and orally administered at the same time as immunization to examine its effect (second step). [Table] Footpad thickness of untreated mice As a result, as shown in Table 2, oral administration of BF-127 substance reduced the DTH response by 30-50% at any dose.
Reinforced. The cell-mediated immunity-enhancing effect of the BF-127 substance was also observed in other strains of mice of different ages. CDF ₁ (DBA/2×BALB/CF ₁ ) male, 9
Week-old mice were immunized as described above,
Each of the BF-127 substances was administered intraperitoneally or orally, and the results were examined (Table 3). [Table] Footpad thickness of untreated mice As shown in Table 3, enhancement was observed in both intraperitoneal injection and oral administration. As described above, it has been revealed that the substance BF-127 acts to enhance cell-mediated immunity, and this suggests that it can be used for immunochemotherapy of cancer. Below, the effect of substance BF-127 on multiple animal tumors was investigated. (2) Effect on multiple animal tumors (a) Effect on Gardner lymphoma Once a week to female, 10-week-old C3H/He mice.
10 ⁵ Gardner lymphoma cells that have been transplanted and passaged were subcutaneously transplanted into the inguinal region of 5 syngeneic mice per group, and after 7 days, they were transplanted once a day for 5 days.
The BF-127 substance was injected intraperitoneally at 10 μg/mouse, 1 μg/mouse, and 0.1 μg/mouse, and the tumor was excised and weighed to determine the effect (Table 4). [Table] As a result, administration of 10 μg/mouse of BF-127 substance inhibited tumor growth compared to the control, and the inhibition rate was 56.5%. (b) Effect on IMC Carcinoma IMC Carcinoma is an ascites-type tumor that occurs naturally in CDF ₁ mice and is transplanted and subcultured into syngeneic mice at the Institute of Microbial Chemistry, and this tumor is an epithelial undifferentiated tumor. This was revealed by pathological examination. IMC
5 x 10 ⁶ tumor cells were subcutaneously transplanted into the inguinal region of 5 CDF ₁ mice per group, and 8 days later, BF-127 treatment was performed.
100 μg to 1 μg/mouse was administered once a day for 5 days, the tumor was excised on the 30th day after transplantation, and the results were examined. At the same time, the anticancer substances mitomycin c 20 μg/mouse and bleomycin 100 μg/mouse were used as controls (Table 5-A). As shown in Table 5-A, administration of the BF-127 substance at 10 μg to 1 μg/mouse showed the same suppressive effect as the anticancer substances bleomycin and mitomycin C. In addition, 1 × 10 ⁶ cells were transplanted, and 10 μg of BF-127 substance was added in the same manner.
~1 μg/mouse was administered intraperitoneally or orally to examine the effects (Table 5-B). Table 5-
As shown in B, both intraperitoneal injection and oral administration showed even better suppressive effects. [Table] [Table] Since the BF-127 substance is a non-cytotoxic substance, the inhibitory effect of this substance on various tumors is thought to be due to a reaction mediated by the host. From the above examples, it is clear that the BF-127 substance is useful for immunodeficiency in cancer-bearing organisms, cancer immunity and chemotherapy, and prevention of metastasis. Furthermore, diseases thought to be caused by immunodeficiency, such as rheumatoid arthritis, multiple sclerosis, collagen disease,
It can be used for systemic lupus erythematosus. Furthermore, the compound of general formula () of the present invention is a low-toxic compound, and as a representative compound thereof, for example, when BF-127 is orally administered to 6-week-old mice, the LD ₅₀ is 1.0 g/Kg or more. .
Therefore, the compound of the present invention can be used as an immunostimulant. The immunostimulant containing the compound (2) of the present invention as a main component contains either the compound (2) or a pharmacologically acceptable salt thereof, a hydrate of the compound, or a salt of the hydrate. It may be hot. The dosage form may be oral, injection, or direct suppository. When preparing an injection, a PH adjuster, buffer, stabilizer, excipient, etc. may be added to the above-mentioned main drug, and Freeze-dried injections can be prepared by freeze-drying in a conventional manner, and by adding PH regulators, buffers, stabilizers, isotonic agents, topical narcotics, etc. Internal and intravenous injections can also be made. When preparing oral solid preparations, add excipients to the main drug, and if necessary, binders, disintegrants, lubricants, coloring agents, flavoring agents, and flavoring agents, and then prepare tablets by conventional methods. Coated tablets, granules, powders, capsules, etc. can be made. When preparing a liquid preparation for oral use, a syrup can be prepared by a conventional method by adding a flavoring agent, a buffering agent, a stabilizer, a flavoring agent, etc. to the main drug. When preparing a direct suppository, a diluent and, if necessary, a surfactant are added to the main drug, and then the suppository can be prepared by a conventional method. The dosage of compound () varies depending on the symptoms, but the usual dosage for adults is 0.02 mg to 200 mg of compound () once a day.
Alternatively, it is preferable to administer the drug more than once a day depending on the symptoms. Next, production examples and preparation examples of the compounds of the present invention will be explained with reference to Examples, but the present invention is not limited thereto. Example 1 10g of the compound () of the present invention was dissolved in distilled water.
The volume was made up to 1000 ml, and after sterilization using a conventional method, 2 ml each was dispensed into vials and freeze-dried. When using this drug, dilute it with distilled water and make an injection. Example 2 1 part of the compound of the present invention (200 parts), 200 parts of lactose, and 50 parts of cornstarch were mixed, granulated and dried with ethanol in a conventional manner, 0.5% magnesium stearate was added, and after mixing, 1 tablet by law
Take the form of a 3.6 mg tablet. Example 3 1 part of the compound of the present invention () and 900 parts of lactose are thoroughly mixed, and the mixture is sieved through a 50-mesh sieve to form a powder. Reference example 1 Hydroxyterephthalic acid dimethyl ester 580
mg was dissolved in 30 ml of anhydrous methanol, and 2.5 g of sodium borohydride was slowly added. After the addition was completed, the mixture was stirred at room temperature for 1 hour, and then the pH was adjusted to 1.95 using 6N hydrochloric acid under ice cooling. The residue obtained by extraction with butanol and distillation of the solvent was purified by silica gel column chromatography (benzene-ethyl acetate = 9:1 to 4:1) to obtain 3-hydroxy-4-(hydroxymethyl). 418 mg of benzoic acid methyl ester was obtained. Yield 83%. Crystallization from benzene-methanol and ethyl ether gave 320 mg of colorless crystals. Melting point 104.5-105℃ Infrared absorption spectrum γ ^KBr _nax (cm ^-1 ): 3430, 3200, 2920, 1708, 1695,
1612, 1590, 1517, 1440, 1420, 1370,
1360, 1310, 1295, 1280, 1255, 1235,
1220, 1190, 1178, 1110, 1095, 1030, 978,
950, 923, 880, 872, 840, 805, 790, 756 Nuclear magnetic resonance spectrum (60MHz, heavy methanol solution) δ ^TMS _ppn : 3.88 (3H.S), 4.72 (2H.S), 7.3-7.7 (
3H) Reference Example 2 Dissolve 316 mg of 3-hydroxy-4-(hydroxymethyl)benzoic acid methyl ester in 1 ml of methanol, add 5 ml of 1N caustic soda, and stir at room temperature.
The saponification reaction was carried out for 1.5 hours. After the reaction, the mixture was diluted with water, 5 ml of 1N hydrochloric acid was added, and extracted with ethyl acetate. Ethyl acetate was distilled off under reduced pressure to obtain 272 mg of -hydroxy-4-(hydroxymethyl)benzoic acid. Yield 93%. Crystallization was performed from a benzene-ethanol mixture to obtain 154 mg of colorless crystals. Melting point 173.5-174.5℃ (decomposed) Infrared absorption spectrum υ ^KBr _nax (cm ^-1 ): 3390, 3080, 2960, 2880, 2800,
2700, ~2150, 1660, 1615, 1588, 1520,
1425, 1395, 1365, 1303, 1248, 1195,
1125, 1098, 1033, 982, 940, 888, 850,
793, 770, 760, nuclear magnetic resonance absorption spectrum (60MHz), heavy dioxane + heavy water) δ ^TMS _ppn : 4.68 (2H.S), 7.35-7.6 (3H) Reference example 3 Using hydroxyterephthalic acid diethyl ester as a raw material compound, 3- by the same method as in Example 1.
Hydroxy-4-(hydroxymethyl)benzoic acid ethyl ester was obtained. Melting point 92-94℃ Elemental analysis Experimental value C: 61.14, H: 6.24 Theoretical value (as C ₁₀ H ₁₂ O ₄ ) C: 61.21, H:
6.17 Ultraviolet absorption spectrum λ ^Etoll _nax (nm): 213 (ε25000), 245 (ε10000
), 300
(ε3500) Infrared absorption spectrum ν ^KBr _nax (cm ^-1 ): 3430, 3140, 1675, 1585, 1420,
1290, 1235, 1090, 1035, 1010, 985, 945,
890, 825, 765 Nuclear magnetic resonance absorption spectrum (60MHz, heavy dimethyl sulfoxide) δ ^TMS _nax : 1.30 (3H.t), 4.26 (2H.q), 4.55 (2H.s
),
5.10 (1H.s), 7.44 (3H.s), 9.70 (1H.s)

Claims (1)

[Claims] 1 General formula () (In the formula, R represents a hydrogen atom or a linear or branched lower alkyl group) and a pharmacologically acceptable salt thereof, and an immunostimulant containing the hydrate of the compound or a salt thereof as an active ingredient . 2 General formula () (wherein R represents a hydrogen atom or a linear or branched lower alkyl group) and its pharmacologically acceptable salts, as well as hydrates of the compound or its salts, are used as immunocancer agents. An immunostimulant according to claim 1, characterized in that: