JP3054741B2 - TAN-1313 and acyl derivatives thereof - Google Patents

Description

Description: TECHNICAL FIELD The present invention relates to a compound TAN-1313 (hereinafter referred to as “T”) which is useful as a therapeutic agent for human autoimmune diseases and rejection of transplanted organs.
AN-1313 ") and acyl derivatives thereof.

BACKGROUND ART Compounds having an immunosuppressive action derived from microorganisms include cyclic peptide compounds, cyclosporin A [JFBorel et al., Agents and Actions.
ns), 6, 468, 1976; A. Rgger et al., Helvetica Chimica Acta, 59.
Volume, 1075, 1979] and macrocyclic lactone compounds, FK
506 [T. Kino et al., The Journal of Antibiotics, 40, 124
9 1987; H.Tanaka et al., Journal of American Chemical Society
mical Society), 109, 5031, 1987].

Autoimmune diseases include a wide range of diseases such as systemic lupus erythematosus, rheumatoid arthritis, multiple sclerosis, pernicious anemia, and generate autoantibodies. Various drugs have been used as therapeutic agents for this disease, but no satisfactory effect has been obtained so far.

On the other hand, in the field of organ transplantation, an anti-rejection agent is used to help the transplanted organ survive. Recently, instead of the traditional combination therapy of azathioprine and prednisolone,
Cyclosporin A is frequently used and has been highly evaluated.
However, at the same time, frequent occurrence of renal disorders and generation of autoreactive lymphocytes associated with the use of cyclosporin A have become a new problem [Pharmacy, Vol. 39, p. 35, p. 49, 1988; Science ( Sience), Vol. 241, p. 1655, 1988].

In order to solve these problems, there is a need in the art for novel compounds having an immunomodulatory activity or intermediate materials for synthesizing those compounds.

Means for Solving the Problems In view of such a current situation, the present inventors have repeatedly searched for an immunomodulator from a new viewpoint. As a result, among a large number of microorganisms isolated from soil, certain microorganisms suppress the concanavalin A (hereinafter abbreviated as Con A) response of mouse spleen cells and a compound that suppresses the mixed lymphocyte reaction induced by alloantigen stimulation in the medium. Knowing that it accumulates in the body, we isolated this compound and examined its physicochemical and biological properties and found it to be a known compound, FK506 (T. Kino et al., The Journal of Antibioti
cs), Vol. 40, p. 1249, 1987), and also revealed that this producing strain is an actinomycete belonging to the genus Streptomyces. Furthermore, the present inventors have found that certain microorganisms belonging to the genus Streptomyces or Amycolatopsis have the ability to produce and accumulate a compound different from FK506 using this FK506 as a substrate. The compound is isolated and characterized for its physicochemical and biological properties, and the compound is represented by the formula (II) Is confirmed to be a compound represented by
I decided to call it. The present inventors have further studied based on these findings and completed the present invention.

That is, the present invention provides a compound represented by the general formula (I) [Wherein, R represents an acyl group. TAN-1
313 Acyl derivative, culture of Streptomyces or Amycolatopsis or a processed product thereof, and compound FK
An expression characterized by contacting with 506 Production method and general formula (II) of TAN-1313 represented by [In the formula, R ¹ , R ² and R ³ are the same or different and represent an organic residue via hydrogen, a hydroxyl group or oxygen. And dissolving the compound represented by the formula (1) in water with a cyclic polysaccharide.

Examples of the acyl group in the general formula (I) include an aliphatic acyl group derived from a carboxylic acid having 1 to 9 carbon atoms, an aromatic acyl group, and an aliphatic acyl group substituted with an aromatic group. Is mentioned.

Examples of the aliphatic acyl group include formyl, acetyl, propionyl, butyryl, pivaloyl, cyclohexanecarbonyl, and the like. Examples of the aromatic acyl group include benzoyl, toluoyl and the like, and an aliphatic group substituted with an aromatic group. Examples of the acyl group include phenylacetyl, phenylpropionyl and the like.

Examples of the organic residue in the organic residue via oxygen in the above general formula (II) include alkyl, aralkyl, asil, cycloalkyl and aminocarbonyl which may be substituted, and the substituent is halogen. Atom, lower alkyl group having 1 to 3 carbon atoms, 1 to 3 oxygen atoms
And an alkoxy group and an amino group.

In the above, examples of the cyclic polysaccharide include α-, β
-And γ-cyclodextrin, among which β-cyclodextrin is preferred.

The strain that produces TAN-1313 used in the present invention may be any microorganism that belongs to the genus Streptomyces or Amycolatopsis and has the ability to produce TAN-1313 using FK506 as a substrate. . Examples include the known bacterium Streptomyces trypofols
omyces tolypophorus) IFO 13151 and Amycolatopsis mediterranei IF
O 13415. IFO 13151 and IFO 1 above
3415 is described in the List of Cultures 1988 edition of the Institute of Fermentation. The microorganisms listed in these lists can be obtained from the fermentation laboratory.

As with other actinomycetes, these microorganisms can change their properties easily, such as ultraviolet rays, X-rays,
Many mutant strains obtained by irradiating with radiation, single spore separation, various mutation treatments, and other mutations or naturally-occurring mutant strains are not substantially different from each other. Any substance having the property of producing the substance using FK506 as a substrate can be used in the method of the present invention.

The medium used for culturing the microorganism may be liquid or solid as long as it contains a nutrient source that can be used by the microorganism, but it is more appropriate to use a liquid medium when treating a large amount. The medium is appropriately mixed with a carbon source that can be assimilated by the microorganism, a nitrogen source that can be assimilated, inorganic substances, and micronutrients. Examples of carbon sources include glucose, lactose,
Sucrose, maltose, dextrin, starch, glycerin, mannitol, sorbitol, fats and oils (eg, soybean oil, lard oil, chicken oil, etc.), n-paraffin and the like. Nitrogen sources include, for example, meat extract, yeast extract, dried yeast,
Soy flour, corn steep liquor, peptone, cottonseed powder, molasses, urea, ammonium salts (eg, ammonium sulfate, ammonium chloride, ammonium nitrate, ammonium acetate, etc.) and the like are used. Further, salts containing sodium, potassium, calcium, magnesium, etc., metal salts such as iron, manganese, zinc, cobalt and nickel, salts such as phosphoric acid and boric acid, and salts of organic acids such as acetic acid and propionic acid are appropriately used. . Other amino acids (eg, glutamic acid, aspartic acid, alanine,
Lysine, methionine, proline, etc.), peptides (eg,
Dipeptides, tripeptides, etc.), vitamins (eg,
B ₁ , B ₂ nicotinic acid, B ₁₂ , C, etc.), nucleic acids (eg, purines,
Pyrimidine, derivatives thereof, etc.).
Of course, an inorganic or organic acid, alkali, buffer or the like may be added for the purpose of adjusting the pH of the medium, or an appropriate amount of an oil or fat, a surfactant or the like may be added for the purpose of defoaming. During liquid culture, the pH of the medium should be around neutral, especially pH
6 to 8 are preferred. The culture temperature is preferably about 24 ° to 30 ° C., and the culture time is preferably about 24 hours to 96 hours.

The “culture” used in the present invention refers to those obtained by the above culture.

The "treated material" used in the present invention means that the culture obtained above is subjected to physicochemical treatment such as filtration, centrifugation, ultrasonic treatment, French press treatment, alumina grinding, lytic enzyme treatment, surfactant or organic compound. It refers to bacterial cells obtained by solvent treatment or the like and crushed bacterial cells containing demethylase. Further, a demethylase obtained by purification by a known method or a cell or a demethylase immobilized by a known method can also be used.

The production of TAN-1313 according to the present invention is carried out by bringing FK506 into contact with a culture of the above microorganism or a processed product thereof. The concentration of FK506 in the reaction solution is suitably from 0.01 to 100 mg / ml. The reaction temperature is suitably from 15 ° to 50 ° C. and the pH is from 5 to 10, but the temperature is preferably from 24 ° to 30 ° C. and the pH is from 6 to 8. The reaction time is suitably from 10 minutes to 100 hours, more preferably from 24 to 96 hours. The reaction may be carried out at rest or under shaking, aeration or stirring conditions, but shaking, aeration or stirring is better. A buffer is added to the reaction solution as needed. Any buffer may be used as long as it can be added to the reaction solution to generate TAN-1313 using FK506 as a substrate. In addition, various organic substances and inorganic substances are appropriately added to the reaction solution in order to increase the reaction efficiency as desired. The detection of the active substance thus obtained is carried out for Candida parapsilosis.
sis) TLC-bioautography using IFO 0640 or HPLC using silica gel is effective.

In order to collect the desired compound TAN-1313 from the culture, since TAN-1313 is a neutral and fat-soluble compound, it can be collected by a general means utilizing this property.

Since the culture TAN-1313 is contained in the liquid, the culture
After adjusting the pH to H2 to 10, preferably pH 3 to 8, the mixture is passed through, and an organic solvent immiscible with water, for example, dichloromethane, ethyl acetate, methyl isobutyl ketone or isobutanol is added to extract the active substance. . To purify the active substance in the extract, an adsorbent resin such as Amberlite XAD-II (manufactured by Rohm and Haas Co., USA), Diaion HP-20 or SP-207
(Mitsubishi Kasei Co., Ltd.) is advantageously used. In order to elute the target active substance from the column using the adsorptive resin, a hydrated solvent, such as hydrated methanol or hydrated acetone, is advantageously used. The extract or eluate thus obtained is concentrated under reduced pressure to obtain a crude substance containing TAN-1313.

Various chromatographic methods are advantageously used to further purify the crude material to obtain pure TAN-1313. Carriers are silica gel, crystalline cellulose, Sephadex
LH-20 (manufactured by Formasia, Sweden) or the like is used, and these are usually performed by column chromatography. To elute the active substance from the column, a suitable organic solvent such as n-hexane, chloroform, toluene, ethyl acetate, dichloroethane, acetone, methanol or the like, alone or in a mixed solvent is used. The eluate is concentrated, and an appropriate solvent such as petroleum benzene, n-hexane, diethyl ether or a mixture thereof is added to the concentrated residue, and the mixture is allowed to stand in a cool place to obtain a crystalline powder of TAN-1313.

The physicochemical properties of TAN-1313 obtained in Example 2 described below are shown below.

(1) Appearance: white powder (2) Optical rotation: -90.4 ° (c 0.55, methanol) (3) Molecular weight: m / z 790 (M + H) ⁺ , (from SI mass spectrum) (4) Elemental analysis value: (%) Found: C, 65.39; H, 8.87; N, 1.70 Calculated: C, 65.38; H, 8.55; N, 1.77; O, 24.30 (5) Molecular formula: C ₄₃ H ₆₇ NO ₁₂ (6) Purple External absorption (UV) spectrum: In methanol, terminal absorption (7) Infrared absorption (IR) spectrum: In KBr tablet, (Figure-
1) Main absorption (fraction, cm ^-1 ) 3450, 2950, 1730 (bro
ad), 1650,1450,1380,1280,1200,1170,1090 (broad), 1
050,990,930,860,780,650,550 (8) Nuclear magnetic resonance (NMR) spectrum: (Table 1) (9) Color reaction Positive: phosphomolybdic acid, concentrated sulfuric acid, potassium permanganate Negative: Dragendorff, ninhydrin, Ehrlich reaction (10) High speed Liquid chromatography (HPLC): Carrier: YMC-Pack A-012 Sil Mobile phase: n-hexane: isopropanol (3: 1) Flow rate: 2 ml / min Detection method: UV absorption (214 nm) Elution time: 5.5 min (11) Thin layer chromatography (TLC): Carrier: Silica gel 60F254 (E. Merck, West Germany) Developing solvent: chloroform: methanol (19: 1) Rf value: 0.36 (12) Property: fat-soluble neutral substance (13) Structure Formula: as follows Various analysis of the NMR spectra by comparing chemical structure of TAN-1313 is a FK506, i.e. COSY method ⁽¹ H- ¹ H, ¹³ C-
It was determined by using a ¹ H) and COLOC method.

The acylation of TAN-1313 is carried out by a method known per se. Generally, in an anhydrous organic solvent, TAN-1313 and a carboxylic acid capable of introducing the acyl group, for example, an acid halide, an acid anhydride, an active ester, etc. By reacting with an acylating agent such as The reaction may be carried out in the presence of a suitable organic or inorganic base, if desired,
The reaction temperature is suitably from 0 ° to 70 ° C and the reaction time is from 0.5 to 48 hours, but the temperature is preferably from 20 ° to 50 ° C and the time is from 1 to 20 hours. Acetylation of TAN-1313 using this method gave TAN-1313 triacetate, and FK506 gave diacetate by the same treatment, thus supporting the above structure. Tables 1 and 2 show ¹³ C NMR spectrum data of these compounds.

On the other hand, as shown in Table 3, TAN-1313 has clearly increased water solubility as compared with FK506, and in particular, further increased water solubility due to the effect of adding β-cyclodextrin. These properties contribute to the development of biological activity and are also considered to be advantageous properties in pharmaceutical preparations.

The solubility was measured as follows. Each sample (5 mg) was added to the respective solvent (5 ml) and stirred at room temperature for 5 minutes using a mixer. The solution was passed through a Millipore filter (<0.45 μm), the same amount of ethyl acetate was added to the solution to extract each compound, the extract was subjected to HPLC, and the solubility was measured.

 A, B, C, and D in the table represent the following.

 A: Water B: 1% α-cyclodextrin / water C: 1% β-cyclodextrin / water D: 1% γ-cyclodextrin / water Next, the biological activity of TAN-1313 will be described.

First, Table 4 shows the inhibitory effect of mouse spleen cells on the blastogenesis.

Assay method: BALB / c mouse spleen cells 1 × 10 ⁶ / ml, Con A (Sigma,
United States) 1 μg / ml or Escherichia coli-derived lipopolysaccharide (hereinafter abbreviated as LPS; manufactured by Difco, USA) 10 μg / ml, 5 μ
M 2-mercaptoethanol, 2 mM L-glutamine, 3
RP containing 0μg / ml gentamicin (Flow Laboratories, Scotland) and 10% fetal calf serum (Wittaka M.A. Bio Products, USA)
TAN-1313 is appropriately added to MI 1640 medium (Wittercar MA Bioproducts, USA) at 37 ° C, 5%
After culturing for 3 days under carbon dioxide, the MTT reduction method [Tada et al.
Journal of Immunological Methods (Jour
nal of Immunological Method) 93, 157, 1986]
The lymphocyte blastogenesis reaction was measured by the method.

Next, Table 5 shows the inhibitory effect of TAN-1313 on the mouse mixed lymphocyte reaction.

Measuring method: C3H / He mouse spleen cells were used as response cells, and DBA / 2 mouse spleen cells were prepared in advance using mantomycin C (Kyowa Hakko) 25
The cells that were allowed to stand at 37 ° C. and 5% CO 2 for 30 minutes at 37 μg / ml were used as stimulated cells. Both were suspended in the same RPMI 1640 medium as in Table 4, mixed to a cell concentration of 2.5 × 10 ⁶ / ml, TAN-1313 was added as appropriate, and the suspension was added at 37 ° C. under 5% CO 2 for 5 minutes.
After culturing for one day, the proliferation of responder cells was measured by the MTT reduction method.

Furthermore, mouse interleukin 2 of TAN-1313
(Hereinafter abbreviated as "IL-2").
BALB / c mouse spleen cells were cultured in RPMI 1640 medium
Suspend to a cell concentration of × 10 ⁶ / ml, and add Con A1
μg / ml and an appropriate concentration of TAN-1313, and added at 37 ° C., 5%
The cells were cultured under carbon dioxide for 18 hours. Next, IL of this culture supernatant was
-2 is an IL-2-dependent mouse T cell line CTLL-2 (ATCC
TIB214) was measured as an index. As a result, TAN
-1313 inhibited IL-2 production by 50% or more at a concentration of 1 ng / ml.

In addition, the triacetate of TAN-1313 suppressed mouse lymphocyte blastogenesis by Con A at a concentration of 20 ng / ml by 50% or more.

As is apparent from the above, TAN-1313 and its acyl derivative are useful substances for treating autoimmune diseases in humans or as drugs for preventing rejection of transplanted organs, because they exhibit an inhibitory effect on lymphocyte function.

In order to investigate the acute toxicity of TAN-1313, a test was performed by intraperitoneal administration using mice, and as a result, the dose was 200 mg / kg.
No deaths were observed.

The compound (I) is used parenterally or parenterally by injection or topically by topical application to the skin, mucous membranes, vagina, rectum and the like, in addition to oral administration.

The dose of Compound (I) when used in humans may vary depending on the disease to be treated, the administration route, the age of the individual patient to be treated and the degree of the disease.
200 mg, especially 0.1-50 mg, is used for the treatment of disease.

For oral administration, the preparation is in the form of capsules, tablets, granules, syrups, powders, etc., together with compound (I), additives such as excipients, binders, disintegrants, lubricants, coloring agents. ,
Flavoring agents, stabilizers and the like may be included. Among them are starch, sucrose, fructose, lactose, glucose, mannitol, sorbitol, precipitated calcium carbonate, crystalline cellulose, carboxymethylcellulose, dextrin,
Gelatin, gum arabic, magnesium stearate,
Talc, hydroxypropyl methylcellulose and the like.

For parenteral administration, the active ingredient can be dissolved or suspended in conventional diluents (aqueous or non-aqueous carriers) and used in solutions, emulsions, suspensions, suppositories and other suitable dosage forms. it can. Examples of the diluent include physiological saline, Ringer's solution, aqueous glucose solution, alcohol, glycols, glycerin, aliphatic glycerides, oils derived from plants and animals, paraffins and the like. In addition, pharmaceutical preparations include additives such as emulsifiers, suspending agents, solubilizers,
Stabilizers, preservatives, soothing agents, tonicity agents, buffers, pH adjusters, coloring agents, coatings and the like may be included. In addition, these preparations can be manufactured by a usual method.

Examples Hereinafter, the present invention will be described more specifically with reference to Examples, but the present invention is not limited thereto. The percentage (%) in the medium represents weight / volume percent unless otherwise specified.

Example 1 Yeast extract / malt extract A Streptomyces trypofols IFO 13151 strain cultured on a slope agar medium was mixed with 40 ml of a 200 ml Erlenmeyer flask containing 1% glucose, 1% tryptone and 0.6% yeast extract (pH 7.0) in a 200 ml Erlenmeyer flask. Inoculate the medium, 28
Culturing was performed on a rotary shaker at 48 ° C. for 48 hours to obtain a seed culture solution.
1 ml of the resulting seed culture was transferred to 40 ml of the above medium in a 200 ml Erlenmeyer flask, and cultured on a rotary shaker at 28 ° C. for 48 hours. In the main culture, at 24 hours, 1 ml of a methanol solution of FK506 (10 mg / ml) was added per flask.

Example 2 After adjusting the culture solution (20) obtained in Example 1 to pH 7,
Extracted twice with ethyl acetate (10). Extract the water with water (5
) And concentrated. Residue (5.5 g) is silica gel 60
(70-230 mesh, 50 g, Merck, West Germany) column chromatography, n-hexane: ethyl acetate (1: 1,600 ml), n-hexane: ethyl acetate (1: 2,600 ml).
And ethyl acetate (600 ml). n-
Concentration of the first half eluate in the elution section of hexane: ethyl acetate (1: 2) yielded FK506 (850 mg) as a raw material. Collect the latter half of the above eluate and the activity category of the ethyl acetate elution part,
After concentration, the residue (970 mg) was again subjected to column chromatography on silica gel 60 (35 g). Chloroform (100m
l), then wash with chloroform: methanol (49: 1,280m
l) and (19: 1,385 ml). The fraction containing TAN-1313 was concentrated, and the residue (315 mg) was further purified with silica gel 6
The column was subjected to 0 (11 g) column chromatography. After washing with n-hexane: ethyl acetate (1: 2,150 ml), elution and fractionation with n-hexane: ethyl acetate (1: 4,150 ml) was performed, and TAN-13 was removed.
The fraction mainly containing 13 was concentrated to dryness. The residue was subjected to column chromatography on Sephadex LH-20 (100 ml, manufactured by Pharmacia, Sweden), and fractionated by elution with methanol. The active fractions were collected, concentrated to dryness and pulverized from n-hexane to give a white powder of TAN-1313 (220 mg)
I got

Example 3 Amycolatopsis mediterranei IFO 13415 cultured on a yeast agar / malt extract slant agar medium was mixed with 500 ml of a medium containing 1% glucose, 1% tryptone and 0.6% yeast extract (pH 7.0) in a 2 L Sakaguchi flask. At 28 ℃,
The cells were cultured on a reciprocating shaker for 24 hours to obtain a seed culture solution. 25 ml of this seed culture, 500 ml of the above medium in a 2 L Sakaguchi flask
and cultured on a reciprocating shaker at 28 ° C. for 20 hours. The obtained main culture solution was centrifuged (8000 × g), the resulting precipitate was collected, suspended in water, and then centrifuged again. Water was again added to the obtained precipitate, and the same operation was repeated twice to obtain washed cells. Next, the washed cells were treated with 50 mM
3- (N-morpholino) propanesulfonic acid [3-
(N-morpholino) propansulfonic acid] buffer (pH
The mixture was suspended in 7.0), adjusted to twice the amount of the original culture (1), and 20 ml of a methanol solution of FK506 (10 mg / ml) was added to a final concentration of 200 μg / ml. This suspension 500
A 2 L Sakaguchi flask containing 20 ml (20 bottles) was stirred on a reciprocating shaker at 24 ° C. for 20 hours to obtain a reaction solution.

Example 4 The reaction solution (10) obtained in Example 3 was kept at pH 6.8.
Extracted twice with ethyl acetate (5). Extract the water with water (3
) And concentrated. Residue (2.3 g) is silica gel 60
(70-230 mesh, 50 g) column chromatography, hexane: ethyl acetate (1: 1,300 ml), hexane: ethyl acetate (1: 2,600 ml) and ethyl acetate (600 m
The fractions were eluted sequentially in l). Hexane: ethyl acetate (1: 2)
The eluate was concentrated to recover the raw material FK506 (1440 mg). Collect and concentrate the active fractions of the ethyl acetate elution part,
The residue (210 mg) was again subjected to column chromatography on silica gel 60 (25 g). After washing with chloroform (100 ml), the mixture was eluted and fractionated with chloroform: methanol (49: 1,280 ml) and (19: 1,385 ml). The fraction containing TAN-1313 was concentrated, and the residue (168 mg) was further purified with silica gel 60 (10 g).
Column chromatography. After washing with hexane: ethyl acetate (1: 2,150 ml), elution and fractionation were carried out with hexane: ethyl acetate (1: 4,150 ml) and ethyl acetate (50 ml), and the fraction containing TAN-1313 was concentrated to dryness. The residue was triturated from hexane to give a white powder of TAN-1313 (97 mg).

Example 5 Crude oil of TAN-1313 (about 80 mg, 27 mg as TAN-1313)
(Contained, determined by HPLC)) was dissolved in acetic anhydride (0.3 ml) and pyridine (0.6 ml) and stirred at room temperature for 7 hours.
The solvent was removed from the reaction solution under reduced pressure.
Dissolve in hexane mixture (2: 1, 15 ml) and add 0.05N hydrochloric acid (10m
l) twice, 2% aqueous sodium hydrogen carbonate solution (10 ml),
The extract was washed successively with water (10 ml) and saturated saline (10 ml), dried over an anhydrous sodium sulfate column, and concentrated under reduced pressure to give a colorless oil (52 mg). This is silica gel 60
And eluted with a mixed solution of ethyl acetate-hexane (1: 4, 1: 2, 2: 3, 30 ml each), and fractionated into 5 ml portions. Each fraction was analyzed by TLC on silica gel (mobile phase: ethyl acetate-hexane 1: 1). Fractions showing a single spot of the target substance were collected and concentrated under reduced pressure to give a TAN-1313 triacetyl white powder. (25 mg) was obtained.

TLC: Silica gel 60F254, developing phase: ethyl acetate-hexane (1: 1), Rf value: 0.56 (TAN-1313: 0.03) Elemental analysis: C ₄₉ H ₇₃ NO ₁₅ (%) Calculated value, C: 64.24, H : 8.03, N: 1.53 actual value, C: 64.14, H: 8.17, N: 1.48 IR (KBr, cm ^-1 ); 3440,2940,1740,1650,1440,1370,1240,
1165,1095,1030,900. ¹³ C NMR spectrum (75 MHz, in deuterated chloroform, δpp
m); Main peak / Sub peak 207.99 / 207.57 (Q), 196.58 / 192.30 (Q), 170.58
(Q), 170.53 / 170.43 (Q), 169.83 / 169.97 (Q), 16
9.17 / 168.89 (Q), 164.85 / 166.12 (Q), 138.48 / 139.3
4 (Q), 135.46 / 135.76 (CH), 132.02 / 130.03 (CH), 13
1.80 / 132.26 (Q), 123.13 / 122.72 (CH), 116.65 / 116.3
_{9 (CH 2), 98.54 /} 97.33 (Q), 79.89 / 78.67 (CH), 75.28
/76.55 (CH), 73.65 / 73.79 (CH), 73.59 (CH), 73.23 / 7
3.12 (CH), 72.98 / 72.28 (CH), 71.45 / 71.64 (CH), 57.
18 / 57.41 (CH ₃ ), 56.40 / 52.76 (CH), 56.31 / 56.20 (C
_{H 3), 53.48 / 52.83 (} CH), 48.99 / 48.13 (CH 2), 43.18 / 4
3.92 (CH ₂ ), 39.17 / 41.87 (CH ₂ ), 37.75 / 37.09 (CH), 3
6.39 / 36.51 (CH ₂ ), 35.58 / 34.96 (CH ₂ ), 34.67 / 33.54
(CH), 34.46 (CH) , 33.98 (CH 2), 32.65 (CH 2), 29.98
_{/30.16(CH 2), 29.57 (CH 2} ), 27.55 / 26.26 (CH 2), 25.9
6 / 26.43 (CH), 24.28 / 24.49 (CH ₂ ), 21.11 / 21.00 (CH ₃
× 3), 20.90 / 20.72 (CH ₂ ), 19.85 / 19.50 (CH ₃ ), 16.12
/16.04 (CH ₃ ), 15.71 / 16.36 (CH ₃ ), 12.50 / 13.52 (C
_{H 3), 10.33 / 10.20 (} CH 3). The symbols in the parentheses indicate the following.

Q: quaternary carbon, CH: methine, CH ₂ : methylene, CH ₃ : methyl Effect of the Invention The TAN-1313 and its acyl derivative of the present invention are compounds having an immunosuppressive action and are useful as pharmaceuticals.

[Brief description of the drawings]

 Figure 1 shows the IR absorption (in KBr) of TAN-1313.

──────────────────────────────────────────────────の Continued on the front page (51) Int.Cl. ⁷ identification code FI C12R 1: 465) (C12P 17/18 C12R 1:01) (58) Investigated field (Int.Cl. ⁷ , DB name) C07D 498/18 A61P 37/02 A61P 37/06 C12P 17/18 BIOSIS (DIALOG) CA (STN) REGISTRY (STN) WPI (DIALOG)

Claims (3)

(57) [Claims] (1) General formula [Wherein, R represents an acyl group. TAN-1
313 acyl derivative. 2. A formula comprising contacting a culture of Streptomyces or Amycolatopsis or a treated product thereof with the compound FK506. A method for producing TAN-1313 represented by the formula: 3. The general formula [In the formula, R ¹ , R ² and R ³ are the same or different and represent an organic residue via hydrogen, a hydroxyl group or oxygen. A method for producing a water-soluble preparation of a compound represented by the formula: