WO2004083186A1

WO2004083186A1 - Therapeutic agent for viral hepatitis and carcinostatic agent

Info

Publication number: WO2004083186A1
Application number: PCT/JP2004/003509
Authority: WO
Inventors: Kunitada Shimotohno; Naoki Tohdoh
Original assignee: Sumitomo Pharmaceuticals Co. Ltd.
Priority date: 2003-03-18
Filing date: 2004-03-16
Publication date: 2004-09-30
Also published as: JPWO2004083186A1

Abstract

A therapeutic agent for chronic hepatitis C which contains either a compound represented by the formula (1) (wherein R1 represents hydrogen or ribofuranosyl; and R2 represents hydrogen or a hydroxy-protecting group) or a salt thereof. It is administered in combination with interferon.

Description

Anti-diabetic hepatitis agent and anticancer drug

The present invention relates to a combination therapy with interferon for viral hepatitis or cancer disease. Background art

As a mainstream treatment method for hepatitis, for example, for chronic hepatitis C, treatment mainly for eliminating hepatitis C virus (HCV) and improving liver function is being promoted. In particular, the treatment of chronic hepatitis C with interferon, which has both HCV elimination and liver function improving effects, is regarded as the only treatment to eliminate the virus, but the complete elimination efficiency of HCV is 20 to 30. In recent years, interferon combination therapy with ribavirin pamantadine has been promoted. The effects of interferon are broadly classified into antiviral effects that act directly on infected cells and activation of host immunity.

The antiviral effect of interferon is caused by signal transduction via the interferon receptor, and promotes the degradation of HCV RA and the inhibition of translation by inducing the production of antiviral factors. In addition, activation of immunity by interferon is involved in activation and maintenance of the antiviral state of a living body by activating cellular immunity and humoral immunity. Interferon therapy for HCV has different therapeutic effects depending on patient characteristics such as HCV serotype, viral load, and histological progression. In particular, it is known that the improvement effect is low and the virus elimination rate is low in cases of ginotype I and hyperviremia. This low exclusion rate is thought to be related to viral factors and the patient's immunological background factors.

Improving liver function with immunomodulators, such as glycyrrhizin and interphenic acid, has been attempted for patients who show resistance to interferon, but liver function is promptly improved after glycyrrhizin administration is completed. At present, hepatocellular carcinogenesis has not been suppressed against interfering oral resistant chronic hepatitis C due to worsening conditions.

Currently, the most effective treatments are interferon and pegylated interferon Although combined with villin, the complete elimination rate of the virus is still about 50% even with 1 year of combined treatment with interferon and ribavirin, and its efficacy is still not complete (Nippon Shokakibyo Gakkai Zasshi. 1995; 92: 1929). —1936). In addition, a response rate of 50% or less can still be expected for HCV genotype 1 which is considered to be difficult to respond to the interface. The response rate for refractory high-virus cases infected with dienotype 1 is about 40%, which is still insufficient as a clinical therapeutic effect. Ribavirin has also been reported to frequently cause hemolytic anemia due to its high accumulation in the body (N. Engl. J. Med. 2002; 347: 975-982).

Interferon is also being promoted as a treatment for cancer, but is administered interferon alone as chronic myelogenous leukemia, multiple myeloma, non-Hodgkin's malignant lymphoma, renal cell carcinoma, etc. In combination with is limited to limited cancers such as hepatocellular carcinoma, stomach cancer, spleen cancer, prostate cancer, esophageal cancer, superficial bladder cancer (Am. J. Surg. 2000; 179: 367-371 Prostate 1998; 35: 56-62, Am. J. Clin. Oncol. 2002; 25: 391-397). The effect is thought to be due to the effects of interferon on cell growth suppression and the activation of cell-mediated immunity, but the therapeutic effect is not satisfactory.

Therefore, if a highly safe drug that activates cell-mediated immunity, enhances the antiviral effect of interferon, and has no problems such as accumulation in the body is found, it will be very useful not only for chronic hepatitis C However, it is likely to be an effective drug for cancer. Disclosure of the invention

It is an object of the present invention to provide a hepatitis virus elimination rate and a cancer cell elimination rate, especially a hepatitis C virus elimination rate and chronic myelogenous leukemia, multiple myeloma, myelodysplastic syndrome, and polycythemia in combination with interferon. It is intended to provide a compound which has an improved elimination rate of cancer cells such as non-Hodgkin's malignant lymphoma, hepatocellular carcinoma, and stomach cancer, and has no accumulation in the body and is highly safe.

The present inventors have conducted intensive studies to solve the above-mentioned problems, and as a result, have found that the compound represented by the formula (1) enhances the antiviral effect when used in combination with interferon. Furthermore, the optimal concentration range of the anti-Wilz effect enhancing action is The present inventors have newly found that the concentration corresponds to the concentration range in which interferon-γ production is induced (enhancement of production), and completed the present invention.

That is, the present invention relates to formula (1)

[In the formula, R ¹ represents a hydrogen atom, a nitrogen atom protecting group or a nitrogen atom substituent, and R ² represents a hydrogen atom or a hydroxyl group protecting group. ] Or a salt thereof as an active ingredient, which is a combination drug with interferon which is effective and safe against viral hepatitis and cancer diseases.

The gist of the present invention resides in the following points (1) to (49).

(1) Equation (1)

(Wherein R ¹ and R ² have the same meanings as described above) or a salt thereof as an active ingredient, in combination with interferon for viral hepatitis or cancer disease,

(2) The substituent of the nitrogen atom of R ¹ is a ribofuranosyl group, a 2-hydroxyethoxymethyl group, a 2-hydroxymethyl-1,3-oxathiolan-5-yl group, or a 4-hydroxymethyl-2-methylcyclopropyl Benzyl group, [2-hydroxy-11- (hydroxymethyl) ethoxy] methyl group, tetrahydroxy 2-furyl group, n-hexylcarbamoyl group, 3-hydroxymethyl-tetrahydro-2-furyl group, 3 —Hydroxyme Cyl-5-hydroxy-tetrahydro-2-furyl group, 3-hydroxymethyl-4,5-dihydroxy-1-tetrahydro-2-furyl group, 3-hydroxymethyl-4,5-dihydroxy-5-methinolatetetrahydro-2- The combination therapy with interferon according to (1), which is a furyl group or a 3-hydroxymethyl-4-azidotetrahydro-2-furyl group.

(3) The agent according to (1) or (2), wherein R ¹ is a hydrogen atom or a lipofuranosyl group.

(4) The agent according to any one of (1) to (3), wherein R ² is a chromium atom.

(5) the interferon is alpha-interferon α (1) to (4), the agent according to any of the above,

(6) The agent according to any one of (1) to (4), wherein the interferon is interferon-αηΐ.

(7) The interferon is Sumiferon, Intron II, Adpaferon, Equinintron or Egasis (1) to (4)

(8) Interferon is 1/3 of interferon (1) to (4)

(9) The agent according to any one of (1) to (8), wherein the dose of the agent is 10 mg to 300 mg, and is administered one to three times a day.

(10) The agent according to any one of (1) to (8), wherein the dose of the agent is 20 mg to: 100 mg, and is administered 1 to 3 times a day. ,

(11) The agent according to any one of (1) to (8), wherein the agent has a characteristic of exhibiting activity at a concentration of 10 μΜ or less.

(1 2) The agent according to any one of (1) to (8), wherein the agent has a characteristic of exhibiting activity at a concentration of 1 μΜ to 8 μΜ.

(13) The agent is characterized in that it exhibits activity at a blood concentration of 10 μΜ or less.

The agent according to any one of (1) to (8),

(14) The agent according to any one of (1) to (8), wherein the agent exhibits activity at a blood concentration of 1 μm to 8 μm.

(15) The viral hepatitis is chronic hepatitis C (1) to (14). Agent

(16) The cancer is chronic myelogenous leukemia, multiple myeloma, myelodysplastic syndrome, polycythemia vera, non-Hodgkin's malignant lymphoma, dry cell cancer, gastric cancer, superficial bladder cancer, knee cancer, renal cell cancer , Prostate cancer or esophageal cancer (1) to (14)

(17) A compound represented by the formula (1) (wherein R ¹ and R ² are as defined above.

. ) Or an anti-hepatitis virus agent comprising a salt thereof as an active ingredient, which is used in combination with interferon;

(18) the agent according to (17), wherein the hepatitis virus is hepatitis C virus;

(19) The substituent for the nitrogen atom of R ¹ is a ribofuranosyl group, 2-hydroxyethoxymethyl group, 2-hydroxymethyl-1,3-oxathiolane-15-group, 4-hydroxymethyl-2-cyclopropenyl group , [2-hydroxy-1- (hydroxymethyl) ethoxy] methyl group, tetrahydroxy 2-furyl group, n-hexylcarbamoyl group, 3-hydroxymethyl-tetrahydroxy 2-furyl group, 3-hydroxy Doxymethyl-5-hydroxy-1-tetrahydro-2-furinole group, 3-hydroxymethynole-4,5-dihydroxy-1-tetrahydroxy 2-furyl group, 3-hydroxymethyl-14,5-dihydroxy-1-5-methyl-tetrahydro _ (2-) furyl or 3-hydroxymethyl-4-azidotetrahydro-12-furyl, (17) or (18) The agent according to the above,

(20) The agent according to any one of (17) to (19), wherein R ¹ is a hydrogen atom or a lipofuranosyl group.

(21) The agent according to any one of the above (17) to (20), wherein R ² is a hydrogen atom,

(22) The agent according to any one of (17) to (21), wherein the interferon is interferon-a.

(23) The agent according to any one of (17) to (21), wherein the interferon is interferon-αηΐ.

(24) The agent according to any one of (17) to (21), wherein the interferon is Sumiferon, Intron II, Adpaferon, Pug Intron or Pegasis

(25) The interferon is interferon _] 3 (17)-(21) The agent described in the

(26) The agent according to any one of (17) to (25), wherein the agent has a single dose of 10 mg to 300 mg and is administered one to three times a day. ,

(27) The agent according to any one of (17) to (25), wherein the dose of the agent is 20 mg to 100 mg, and the agent is administered 1 to 3 times. ,

(28) The agent according to any one of (17) to (25), wherein the agent has a characteristic of exhibiting activity at a concentration of 10 μΜ or less.

(29) The agent according to any one of (17) to (25), wherein the agent has a characteristic of exhibiting activity at a concentration of 1 μΜ to 8 μΜ.

(30) The agent is characterized in that it exhibits activity at a blood concentration of 10 μΜ or less.

, (17)-(25)

(31) the agent according to any one of (17) to (25), wherein the agent has an activity at a blood concentration of 1 μ 濃度 to 8 μΜ;

(32) A method for treating viral hepatitis, which comprises administering interferon to a patient with viral hepatitis and using any one of the above-mentioned (17) to (31) anti-hepatitis virus agents in combination.

(33) The method according to the above (32), wherein the viral hepatitis is hepatitis C.

(34) The treatment method according to (32) or (33), wherein the compound of any one of (17) to (31) is orally administered, and interferon is administered subcutaneously.

(35) an immune activator comprising a compound represented by the formula (1) (wherein R ¹ and R ² are as defined above) or a salt thereof as an active ingredient:

(36) the agent according to (35), which is characterized by activation of cellular immunity;

(37) The agent according to (35) or (36), which is characterized by inducing interferon-y production (enhancing production).

(38) The substituent of the nitrogen atom of R ¹ is a lipofuranosyl group, a 2-hydroxyethoxymethinole group, a 2-hydroxymethyl-1,3-oxathiolan-1-yl group, or a 4-hydroxymethyl-2-cyclopropyl group. Group, [2-hydroxy-1- (hydroxymethyl) ethoxy] methyl group, tetrahydro-2-furyl group, n-hexylcarba Moyl group, 3-hydroxymethyl-tetrahydroxy 2-furyl group, 3-hydroxymethylinole 5-hydroxytetrahydro-1-furinole group, 3-hydroxymethylinoleate 4, 5-dihydroxy Tetrahydroxy 2-furyl group, 3-hydroxymethylenolate 4, 5-dihydroxy-15-methyl-tetrahydro-2-furyl group or 3-hydroxymethyl-4-furazide-tetrahydro-12-furyl group , (35)-(37) any force, the agent described,

(39) The agent according to any one of (35) to (38), wherein R ¹ is a hydrogen atom or a ribofuranosyl group.

(40) The agent according to any one of the above (35) to (39), wherein R ² is a hydrogen atom,

(41) The agent according to any one of (35) to (40), wherein the agent has a characteristic of exhibiting activity at a concentration of 10 μΜ or less.

(42) The agent according to any one of (35) to (40), wherein the agent has a characteristic of exhibiting activity at a concentration of 1 μΜ to 8 μΜ.

(43) The agent is characterized in that it exhibits activity at a blood concentration of 10 μΜ or less.

The agent according to any one of (35) to (40),

(44) The agent according to any one of (35) to (40), wherein the agent has a characteristic of exhibiting active 1 "activity at a blood concentration of 1 μΜ to 8 μΜ.

(45) The preparation according to any one of (35) to (40), which is administered so that the blood concentration after administration is 10 μΜ or less,

(46) The preparation for oral administration according to any of (35) to (40), which is orally administered so that the blood concentration after administration is 1 μΜ to 8 μΜ.

(47) A method for treating viral hepatitis, which comprises administering the agent described in any one of (35) to (46) to a patient with viral hepatitis.

(48) The method of (47) above, wherein the diarrhea hepatitis is hepatitis C.

(49) A method for treating a cancer disease, which comprises administering the agent described in any one of (35) to (46) to a cancer disease patient. BRIEF DESCRIPTION OF THE FIGURES FIG. 1 shows that mizoribine induces interferon-γ. Splenocytes are prepared from naive mice and the culture supernatant contains sperm antigen

Staphylococcal enterotoxin B (SEB) was added at a concentration of 400 pg / ml. Stimulation was added by adding mizoribine to the culture solution, and after culturing for 48 hours, cytodynamics in the culture supernatant was measured by ELISA.

Figure 2 shows the pharmacokinetics when mizoribine was administered.

(A) shows the pharmacokinetics at the time of administration of 50 mg at a time, three times a day, and (B) shows the pharmacokinetics at the time of administration of 100 _mg at a time, three times daily.

FIG. 3 shows the results of measurement of RNA derived from HCV replicon by HLBI injection.

FIG. 4 shows the suppressive effect of replicon thighs by the combined use of Ribavirin (RBV) and natural interferon.

FIG. 5 shows the suppressive effect of replicon RA by the combined use of Mizoribine (MIZ) and natural interferon-α.

FIG. 6 shows the inhibitory effect of replicon RA by the combined use of SM-108 and natural interferon-1α.

FIG. 7 shows the inhibitory effect of replicon RNA by the combined use of natural interferon-α. The average value of the copy number measured twice by the real-time PCR method and the reduction (%) of the replicon RNA are shown. Ribavirin and mizoribine show the results of two experiments, and SM-108 shows the results of one experiment.

FIG. 8 shows the effect of HLBI and ribavirin in combination to eliminate replicon RNA. Samples from each lane were: 1; # 50-1 cells, 2-7; HLBI 3 IU / ml + ribavirin (2; 0 μΜ, 3; 1 μΜ, 4; 3 μΜ, 5; 5 μΜ, 6; 10 μΜ, 7; 20 μΜ).

(Α) is a diagram showing a migration pattern by 1% agarose gel electrophoresis. (Β) Northern hybridization using a probe that detects HCV thighs (arrow indicates replicon RNA). (C) Northern hybridization using G3PDH mRNA detection probe. (The arrow indicates G3PDH mRNA).

FIG. 9 shows the elimination effect of replicon RNA by the combined use of HLBI and mizoribine. Samples in each lane were: 1; # 50-1 cells, 2-7; HLBI 3 IU / ml + mizoribine (2; 0 μΜ, 3; 1 μΜ, 4; 3 μΜ, 5; 5 μΜ, 6; 10 μΜ, 7; 20 μΜ). (A) is a view showing a migration pattern by 1% agarose gel electrophoresis. (B) The result of Northern hybridization using a probe for detecting HCV RNA (the arrow indicates that the replicon is awake). (C) Northern hybridization using a probe for detecting GSPDH mRNA (arrows indicate G3PDH mRNA).

FIG. 10 shows the effect of HLBI and SM-108 in combination to eliminate replicon RNA. Samples in each lane were NC; Huh-7 cells, 1; # 50-1 cells, 2-7; HLBI 3 IU / ral + SM-108 (2; 0 μΜ, 3; 1 μΜ ヽ 4; 3 μΜ, 5; 5 μΜ, 6; 10 μΜ, 7; 20 μΜ). (Α) is a diagram showing a migration pattern by 1% agarose gel electrophoresis. (Β) Northern hybridization results using a probe that detects HCV RA (arrows indicate replicon RNA). (C) Northern hybridization using a probe for detecting G3PDH mRNA (arrows indicate GI3PDH mRNA).

FIG. 11 shows the disappearance effect of replicon RA by combined use of HLBI and ribavirin by Northern hybridization angle analysis. The ratio of replicon RNA to the amount of G3PDH gene expression at each amount of ribavirin added is shown.

FIG. 12 shows the elimination effect of replicon RA by the combined use of HLBI and mizoribine by Northern hybridization angle-harvesting. FIG. 4 is a graph showing the ratio of replicon RA to the amount of G3PDH gene expression at each amount of mizoribine added.

FIG. 13 shows the effect of elimination of replicon RA by the combined use of HLBI and SM-108 by Northern hybridization analysis. FIG. 3 is a view showing the ratio of replicon RNA to the amount of G3PDH gene expression in each amount of added SM-108.

Fig. 14 shows the anti-BVDV activity when sumiferon, mizoribine, and SM-108 were used alone or in combination (the vertical axis indicates the degree of cell proliferation, and the higher the value, the more effective the cell treatment with the drug. Indicating that they are growing).

(A) Sumiferon 30 IU / ml, mizoribine and SM-108 0.12 μΜ, (Β) Sumiferon 10 IU / ml, mizoribine SM-108 0.08 μΜ, used alone or in combination The result of the case is shown.

FIG. 15 shows the anti-BVDV activity when Sumiferon, Adpaferon or Intron II was used in combination with Mizoribine or SM-108.

(A) is 1 IU / ml of sumeferon or adpaferon, 1.88 μΜ of mizoribine, (B) shows the results obtained when using sumiferone or adpaferon at 10 IU / ml and mizoribine 1.88 μΜ alone or in combination. (C) shows the results when using interferon-α with 10 IU / ml each of sumiferon, intron II or adpaferon, and 7.5 μΜ of SM-108 alone or in combination. BEST MODE FOR CARRYING OUT THE INVENTION

A first aspect of the present invention is a combination therapy with interferon for the compound represented by the general formula (1) or a salt thereof.

[In the formula, R ¹ represents a hydrogen atom, a nitrogen atom protecting group or a nitrogen atom substituent, and R ² represents a hydrogen atom or a hydroxyl group protecting group. ]

Examples of the substituent represented by R ¹ in the compound of the present invention include a hydrogen atom, a lipofuranosyl group, a 2-hydroxyethoxymethyl group, a 2-hydroxymethyl-1,3-oxathiolan-1-inole group, and a 4-hydroxymethylino. Leh 2-cyclopropininole, [2-hydroxy-11- (hydroxymethyl) ethoxy] methyl, tetrahydro-2-furyl, η-hexylcarbamoyl, 3-hydroxymethyl-tetrahydryl 2-furyl And a 3-hydroxymethyl-4-azidotetrahydro-2-furyl group. Among them, a hydrogen atom and a ribofuranosyl group are preferable.

Examples of the protective group represented by R ¹ in the compound of the present invention include a substituted or unsubstituted alkoxycanoleboninole group such as a methoxycarbonyl group, an ethoxycarbonyl group, a t-butoxycarbol group, etc., for example, p-etrobenzi ^ / reoxycanolevo And substituted or unsubstituted aryloxycarbonyl groups such as phenolic groups, benzylinoxycarbonyl groups, and p-methoxybenzyloxycarbonyl groups. Also, protecting groups that are deprotected in vivo It can be used and can be used depending on the situation.

Examples of the substituent represented by R ² in the compound of the present invention include a protecting group for a hydrogen atom or a hydroxyl group. Here, the hydroxyl-protecting group includes, for example, those described in The Chemical Society of Japan, Experimental Chemistry Course (Maruzen), and refers to a protecting group that can be chemically or biologically cleaved to return to a hydroxyl group. For example, acetyl groups such as acetyl group and benzoyl group can be mentioned.

Furthermore, it can be protected by condensing with a compound having a carboxyl group having antiviral activity to form an ester bond. For example, the compound having a carboxyl group may be an HCV RNA polymerase inhibitor such as JTK-003 or BILN-2061.

Examples of the salt include pharmaceutically acceptable salts, for example, a salt with an inorganic base, a salt with an organic base, a salt with an inorganic acid, a salt with an organic acid, a salt with a basic or acidic amino acid And the like. Preferable examples of the salt with an inorganic base include, for example, alkali metal salts such as sodium salt and potassium salt; alkaline earth metal salts such as calcium salt and magnesium salt; and aluminum salt, ammonium salt and the like. Can be Preferable examples of the salt with an organic base include, for example, trimethylamine, triethylamine, pyridine, picoline, ethanolanolamine, diethanolamine, triethanolamine, dicyclohexylamine, N, N′-dipentinoleethylene. Salts with diamine and the like. Preferable examples of the salt with an inorganic acid include salts with hydrochloric acid, hydrobromic acid, nitric acid, sulfuric acid, phosphoric acid and the like. Preferred examples of salts with organic acids include, for example, formic acid, acetic acid, trifluoroacetic acid, fumaric acid, oxalic acid, tartaric acid, maleic acid, citric acid, succinic acid, malic acid, methanesulfonic acid, benzenesulfonic acid, : — Salts with tonoren sulfonic acid and the like. Preferred examples of the salt with a basic amino acid include, for example, salts with arginine, lysine, orditin and the like. Preferred examples of the salt with an acidic amino acid include, for example, salts with aspartic acid, glutamic acid, etc. Are listed.

In the present invention, a compound in which the substituent represented by R in the above general formula (1) is a lipofuranosyl group is known as mizoribine, and a compound in which the substituent is a hydrogen atom is known as SM-108. Mizoribine is marketed as a drug for renal rejection and rheumatoid arthritis, but serious side effects such as hemolytic anemia have hardly appeared clinically. Mizoribine is It has an antiproliferative effect on specific cancer cell cells (Cancer Res. 1975; 35: 1643-1648), has an antiviral effect (Exp. Opin. Invest. Drug 2000; 9: 221-235, Antiviral Chemi. Chemother. 1994; 5: 366-371; J Infect Dis 1993; 168: 641-64). As an effect on immunity, the stimulatory response of immunity is maintained in vitro (Transplant. Pro 1992; 24: 2845-2846, J. Clin.

Invest. 1991; 87: 940-948), and is known to suppress the proliferation of T cells and B cells (J. Clin. Invest. 1991; 87: 940-948; J. Immunol. 1995; 155: 5175-5183, Clin. Exp. Immunol. 2000; 120: 448-453), and its effect on activating immunity at low concentrations is not known. In addition, it has been reported that the addition of mizoribine induces the expression of IMPDH-2 type gene (Endocrinol. 2001; 142: 193-204). Mizoribine is reported to have a blood half-life of about 2 hours and is a non-accumulative drug (Rheumatology 1991; 5: 287-300).

SM-108 is a prodrug of the active form of mizoribine, a compound that is converted to mizoribine by adenine-phosphoribosyltransferase (Cancer Res. 1982; 42: 1098-1102). It has been shown to be clinically effective when administered as a single agent against hematopoietic tumors such as chronic myelogenous leukemia, myelodysplastic syndrome, and polycythemia vera and lung cancer, and is reported to be an extremely safe drug as an anticancer drug. (Cancer and Chemotherapy 1989; 16: 123-130; Cancer and Chemotherapy 1989; 16: 113-121).

Mizoribine and SM-108 have not been administered to chronic hepatitis C as to whether they have an antiviral effect on HCV or the ability to eliminate virus-infected cells, and their efficacy is unknown. Nor has it been studied in combination with interferon.

The viral hepatitis in the present invention refers to, for example, hepatitis due to viral infections such as hepatitis C virus (HCV) and hepatitis B virus (丽), that is, hepatitis C, hepatitis B and the like. The viral hepatitis may be acute hepatitis, fulminant hepatitis or chronic hepatitis, and chronic hepatitis may be associated with cirrhosis. Viral hepatitis preferably includes chronic hepatitis C and chronic active hepatitis B, and more preferably chronic hepatitis C.

Hepatitis C virus includes, for example, the dienotypes la, lb, 2a, 2b, 3a, There are genotypes such as 3b, but are not limited to these HCV genotypes.

The cancer diseases according to the present invention include chronic myelogenous leukemia, multiple myeloma, myelodysplastic syndrome group, polycythemia vera, non-Hodgkin's malignant lymphoma, hairy cell leukemia, and hematopoietic tumors represented by cutaneous T lymphoma. Solid cancers such as hepatocellular carcinoma, gastric carcinoma, superficial bladder carcinoma, renal carcinoma, renal cell carcinoma, prostate carcinoma, esophageal carcinoma, melanoma, and lipopositive sarcoma associated with AIDS can be mentioned. Chronic myelogenous leukemia, multiple myeloma, myelodysplastic syndrome group, polycythemia vera, non-Hodgkin's malignant lymphoma, hepatocellular carcinoma, gastric cancer, superficial bladder cancer, knee cancer, renal cell cancer, prostate cancer and esophagus It is cancer.

The interferon (IFN) in the present invention is a natural IFN or a recombinant IFN. The natural IFN may be derived from lymphoblasts, leukocytes, or the like. IFN-O; IFN-O; -] 3 and IFN-ω. IFN was screened by a DNA shuffling method, or the like, in which one or more amino acids in the amino acid sequence were substituted, deleted and / or modified, as long as they had substantially the same action as the natural IFN. The modified product may be a modified product in which the sugar chain is substituted, deleted, Z- or added, or similarly, a sustained interferon agent such as PEGylated IFN or albumin-forced UIFN, tissue absorption or tissue transfer. It may be a DDS formulation of IFN with enhanced properties. Preferable examples include natural or recombinant IFN-a, natural or recombinant IFN-consensus IFN, and PEGylated IFN. As the IFN, a known commercially available IFN can be used.

Interferon-a (IFN-a) includes, for example, natural IFN-α and recombinant IFN-α.

2a and recombinant IFN-a 2b and the like. Any subtype may be used as long as it is IFN-α. Specifically, for example, the natural forms of IFN-0 !, IFN-αηΐ, such as Sumiferon (Sumitomo Pharmaceutical), OIF (Otsuka Pharmaceutical) and IFN-α Mochida (Mochida Pharmaceutical), and Ito-Katari IFN-a2a Examples include, but are not limited to, certain canferon II (Takeda Pharmaceutical), mouth feron A (Chugai Pharmaceutical), and intron II (Schering-Plough), which is a recombinant IFN-α2b.

Interferon-1] (IFN-β) includes, for example, natural IFN-β and recombinant IFN-β, and any type of IFN-i3 may be used. Specifically, for example, iFN i3 Mochida (Mochida Pharmaceutical), Feron (Toray), mm For example, beta-feron (Nippon Schering), which is IFN-lb, can be mentioned, but is not limited thereto.

Consensus interferon (consensus IFN) is a newly developed IFN based on the consensus sequence theory and includes, but is not limited to, interferon-analfacon-1. Adfaferon (Yamanouchi Pharmaceutical Co., Ltd.) is an example of the interface / refacon-1. A long-acting interferon is an interferon formulation that uses PEG (polyethylene glycol), microparticles and nanoparticles, atelocollagen, silicon, and other sustained-release carriers to improve its sustainability. IFN is PEG (polyethylene glycol) bound to an IFN protein molecule. What is a long-acting interferon?

For example, PEGylated IFN-α 2a and PEGylated IFN-hi 2b can be mentioned. The types of IFN are not limited to these. Specific examples include Vegasis (Roche), which is PEGylated IFN-hi 2a, and Peguintron (Schering'Brow), which is PEGylated IFN-α2b.

Interferon DDS preparations with enhanced tissue absorption include, for example, pulmonary or nasal administration using microparticles, nanoparticles, or other fine particle carriers.

, Interferen preparations with improved absorption efficiency in the nasal mucosa, etc.

1 Interferon DDS preparations with enhanced tissue absorption may be used, and are not limited to these.

Interferon DDS preparations with improved tissue transport include, for example, interferon preparations that have improved affinity for the liver by modification, such as glycation of sugar chains, etc. It is sufficient if it is a DDS preparation of interferon, and it is not limited to these.

As the interferon (IFN) in the present invention, preferably, Sumiferon, Intron A, Adpaferon, Pguintron and Pegasis, particularly preferably Sumiferon, Intron A and Adpaferon, more preferably Sumiferon are used.

The administration in the present invention includes, but is not limited to, oral administration, parenteral administration (including rectal administration, subcutaneous injection, intravenous injection, intramuscular injection, intraperitoneal injection, etc.). No. Administration is more preferably oral administration. The administration can be carried out by appropriately adjusting the dose and the number of administrations according to the administration form, age, weight, symptoms, side effects, etc. of the patient.

The single dose is not particularly limited as long as it is not more than an amount that causes side effects, but is, for example, 10 mg to 500 mg, preferably 10 mg to 300 mg, and more preferably 20 mg to: LOO mg. But not limited to this.

The number of administrations is, for example, 1 to 3 times a day, but is not limited thereto. The administration may be once every other day or several days. It may be administered several times daily at the beginning of administration, and then continuously several times or once daily or once every several days.

The concentration in the present invention refers to the blood concentration of the agent after administration of the agent. The blood concentration is the concentration of the agent in the blood in the body after administration of the agent, and is not particularly limited as long as it is lower than the concentration that causes side effects and is an effective concentration. The blood concentration is, for example, 10 μΜ or less, and preferably 1 μΜ to 8 μΜ.

The activity in the present invention is an activity of activating an antiviral action or immunity, and refers to an action of enhancing an antiviral action and an action of enhancing the ability to eliminate virus-infected cells when used in combination with interferon. .

A second aspect of the present invention is an immunoactivator comprising a compound represented by the general formula (1) or a salt thereof as an active ingredient.

The immunity activator in the present invention is an agent having an action of activating immunity, and the cell immunity activator is an agent having an action of activating cell immunity. . Cell-mediated immunity is an immune mechanism in which the immune cells themselves attack and eliminate foreign substances, and are responsible for defense against intracellular parasitic microorganisms, such as killing against intracellular parasites, fungi, and protozoa. It is involved in the elimination of ruptures and rupture of tumor cells, and has natural immunity and acquired immunity.

A third aspect of the present invention is an interferon-γ production inducer (production enhancer) comprising a compound represented by the general formula (1) or a salt thereof as an active ingredient.

The agent for inducing interferon-γ production (enhancing production) in the present invention is an agent for increasing the concentration of interferon-γ (IFN-γ) in blood or / and in living tissue. IFN-γ is an HLA protein involved in surface antigen presentation recognized by immune cells. Ras I antigen, HLA class II antigen,] 32_ Microglobulin, TAP, LMP, etc., induces antigen-specific activation of cytotoxic T lymphocytes (CTL), expression of immunoglobulin Fc receptor It has the effect of activating antigen-specific acquired immunity by effector cells by induction, activating NK cells, and activating macula phage. Therefore, this agent is effective as an agent showing a killing action against intracellular parasite microorganisms such as intracellular parasite bacteria, fungi and protozoa, and a destruction / elimination action of virus-infected cells and tumor cells.

The compound represented by the formula (1) or a salt thereof according to the present invention has a short half-life in the body, does not accumulate in the body, and enhances the antiviral effect of interferon on HCV by combination with interferon. It is useful as a therapeutic agent for viral hepatitis or cancer disease in mammals. Furthermore, the compound represented by the formula (1) or a salt thereof according to the present invention has an action of enhancing an immune response, and thus is useful as an immunoactivating agent.

The compound represented by the general formula (1) or a salt thereof may be used as it is or as a pharmaceutically acceptable carrier known per se (including excipients, bulking agents, binders, lubricants, etc.), Pharmaceutical compositions or formulations (e.g., powders, granules, tablets, pills, capsules, powders, syrups, injections, drops, topical preparations, suppositories, emulsions, enxyls) Agent) can be used. Example

Next, the present invention will be described more specifically with reference to examples. However, the present invention is not limited to these examples.

Example 1

Analysis of immunomodulatory effect of mizoribine

Prepare spleen cells from a mouse (Nippon Charl. Riva Co.) and add mizoribine to the culture solution.

(Sigma) was added for stimulation, and after culturing for 48 hours, cytokines in the culture supernatant were measured by ELISA. Ribavirin (Sigma) stimulation was performed in the same manner as a control. Experimental Method The procedure was performed according to the method of Tam et al. (Antimicrob. Agent. Cherao. 2000; 44: 1276-1283). (1) Preparation of mouse splenocytes

Splenocytes were prepared from naive mice. The mouse spleen was removed, transferred to a 60-ml petri dish containing 5 ml of RPMI 1640 medium, crushed with the blasted part of a slide glass, and collected in a 50 ml centrifuge tube through a cell strainer (FALCON 70 μιτι Nylon). .

1, 200 and centrifuged RPRA _s 5 min, cells were collected. ACK buffer for hemolysis in pellets of splenocytes _{(0. 15M NH 4 C1, ΙΟηιΜ} KHC0 3, 0. IraM EDTA, pH7 2 -. 7. 4) was added and after Shizu匱room temperature, 5 minutes, 20 ml of the medium was added, and collected in a 50 ml centrifuge tube through a cell strainer.

(2) Stimulation by superantigen

Staphylococcal enterotoxin B (SEB; Toxin technology), a superantigen, was added to the culture supernatant at a concentration of 400 pg / ml. After adding 2.5 ral of PBS to 1 mg of SEB, the solution was sterilized by filtration using a 0.2 μπι filter to prepare a concentrated solution of 400 ng / ml. To 0.1 ml of this solution, 100 ml RPMI 1640 medium (1% L-glutaraine, 10% Penicil lin-Streptomycin, 10% FCS) was added, and the spleen-derived cell pellet was washed twice with 10 ml medium for 1,200 minutes. rpm, 5 minutes, after centrifugation washing 4 ° C, were suspended in RPMI 1640 5 ml of medium containing SEB, cell number determined, suspended in RPMI medium containing SEB to be 2 X 10 ⁶ cells / ml The solution became turbid and 1 ml was added to a 24-well plate.

(3) Examination of immune response by mizoribine

To each well, 1 raM mizoribine solution was directly added to the medium, and the cells were cultured at 37 ° C for 48 hours, and then IFN-γ in the culture supernatant was measured. The mizoribine stimulation concentration was 0, 1, 2, 5, 10, 15, 20 μΜ. Dissolve 10.368 mg of mizoribine (molecular weight 259.2) in 400 μ PBS of PBS, dissolve 12.21 mg of linobilin (molecular weight 244.2) in 500 μΐ of PBS, and filter with a 0.2 μιη filter. , 100 mM solution, diluted 10 times or 100 times with PBS, and used for experiments (4) Measurement of IFN-T /

For IFN-γ, use 10 μΐ of the culture supernatant, and use Genztme / Techne's AN'ALYZA (registered trademark).

The measurement was performed using an Immunoassay system mouse IFN-γ kit. The absorbance was measured at 450 nm by an endpoint method using a microplate reader (BioRad, model 3550 UV). Perform 4-parameter regression using SoftMax (Molecular Device) The concentration was calculated. Fig. 1 shows the measurement results.

From the results shown in FIG. 1, it was found that the interferon-γ-inducing action was suitable at a 1 μΜ concentration and a mizoribine concentration of 8 μΜ. This concentration corresponds to approximately 0.25 g / ral to 2 | ig / ml, and according to the literature on pharmacokinetics (Rheumatology 1991; 5: 287-300), a single dose is 50 mg to 100 mg. Is good. Figure 2 shows the pharmacokinetics of mizoribine described in the literature. As shown in Fig. 2, mizoribine disappears at a high rate (Τ _1/2 = 2.2 hours), and no drug accumulation is observed. Repeated doses maintain drug responsiveness. In addition, since there is no accumulation of the drug, the frequency of administration may be once to three times a day.

In addition to its antiviral effect, interferon-1 α /] 3 promotes differentiation of Th1 cells, induces proliferation of CD8 + T cells with antigen-specific cytotoxicity, promotes differentiation of NK cells, surface antigen presenting molecule It has various immunomodulatory activities, such as promotion of expression of. The cell-mediated immunity-activating effect of mizoribine and SM-108 is not only effective for the treatment of chronic hepatitis C in combination with interferon-α /] 3, but also at the same dose to eliminate cancer cells. It was shown to be effective.

Example 2

Effects of natural interferon-α on HCV replicon cells

In the presence of high doses of interferon-Q7 J3, it is difficult to examine the enhancement of antiviral activity by adding concomitant drugs. Therefore, the optimal concentration for reducing the amount of HCV-rebricon-derived RA to about 10% was examined.

(1) Construction and culture of HCV replicon cells

The construction of HCV replicon, introduction into cultured cells (Huh-7), establishment of established cells (# 50-1), and cell culture were performed according to the literature (Biochem. Biophys. Res. Comm. 2002; 293: 993-999, J. Virol. 2001; 75: 1437-1449).

(2) Examination of residual amount of HCV replicon-derived RNA by natural interferon # 1 For cell culture, # 50-1 cells were prepared so that 2-3 × 10 ⁴ cells were contained in each well of a 12-well plate. Make a solution of 30, 100, 300, 1000, 3000, 10000 IU / ml of natural interferon-a (Smiferon; HLBI: Sumitomo Pharmaceutical), add ΙΟμΙ per 1 ml (1 ml), and add 0.3, 1 , 3, 10, 30, and 100 IU / ml.

After culturing for one week, the cells are collected and separated with Sepasol RNA 1 Super (Nacalai Tesque, Inc.). RNA was extracted according to the instructions. RNA was quantified using a spectrophotometer and adjusted to a concentration of 50 ng / μΐ.

(3) Measurement of RNA derived from HCV replicon by real-time PCR

The instrument used was PE Applied Biosystems (7700 Sequence Detector). Reaction conditions for real-time PCR are described in the literature (Biochem. Biophys. Res. Comm. 2002;

293: 993-999), 2 μΐ each of the above RNA was used as a template. Of the primer sequences used, bases 130 to 146 of the replicon RNA were used as forward primers, and bases 290 to 272 of the replicon RNA were used as revers primers.

In addition, as a probe labeled for detection with TaqMan, an oligonucleotide (DNA) consisting of bases 148 to 168 of the replicon RNA was labeled with TaqMan Kit (Kuchisha). The primers and probes used were the base sequence shown in Seq. ID No. 1 for the forward primer, SEQ ID NO. 2 for the reverse primer, and SEQ ID NO. 3 for the Probe.

The measurement by real-time PCR was performed twice for each sample, and the average value was obtained. Figure 3 shows the average measured values and the measurement results. The residual amount of RA derived from HCV replicon was about 10%, that is, an HLBI concentration of 3 IU / ml was considered to be appropriate for the evaluation of concomitant drugs.

(4) Antiviral effect of concomitant drug in the presence of interferon

Ribavirin, mizoribine, SM- 108 dissolved each in PBS of, 0. 5, 1, 2, 5, 10 mM solution was prepared, 10 to 2 x l0 ⁴ each Ueru cells / well and the 2 4 well plates μΐ Drugs were added in increments of 0, 5, 10, 20, and 20 µl. HLBI was added so that the addition amount was 0, 3 IU / ml. One week after culturing in the same manner as in Example 2, the amount of residual HCV replicon-derived RNA was measured by real-time PCR. Experiments with ribavirin and mizoribine in combination with HLBI were performed twice each, and experiments with SM-108 in combination with HLBI were performed once.The measurement results for each combination experiment are shown in Figs. 4, 5, and 6, respectively. Was.

According to these results, a transient decrease in HCV replicon-derived RNA was observed for all drugs when a relatively low dose of about 5 μ 添加 was added in the presence of 3 IU / ml HLBI. The RNA loss rate was 18-35% with ribavirin, 53-61% with mizoribine, and 59% with SM-108.The combination of interferon with mizoribine or SM-108 was higher than that with ribavirin. Was. Therefore, when the blood concentration of the drug is 5 μΜ, the effect on HCV in the combination administration of mizoribine or SM-108 is not less than that in the combination administration of ribavirin, and Since the blood concentration of ribavirin in chronic hepatitis C exceeds 10 μΜ, it is concluded that mizoribine and SM-108 are more useful than ribavirin as a concomitant drug with interferon in chronic hepatitis C. Was done.

(5) Northern blot analysis of antiviral effects of concomitant drugs in the presence of interferon-a

For cell culture, # 50-1 cells were prepared in each well of a 6-well plate so that I x lO ⁵ cells were obtained. HLBI at 3 IU / ml, ribavirin, mizoribine, and SM-108 were each added to the mixture to obtain 0, 1, 3, 5, 10, 20, and 20 μ 20. After culturing for one week, the cells were collected, and RNA extraction was performed using Sepasol RNA 1 Super. The obtained RNA was quantified with a spectrophotometer and subjected to 1% agarose gel electrophoresis using MOPS buffer. According to a conventional method, RNA is transferred to a membrane filter, a probe is prepared using a DNA fragment obtained from a plasmid containing an HCV replicon and a DNA fragment of a Glyceraldehyde 3-phosphate dehydrogenase (G3PDH) gene, and hybridization is performed. Was.

As a specific method for preparing a probe, the plasmid pNNRz2 DNA (Biochera. Biophys. Res. Comm. 2002; 293: 993-999) into which the HCV genome was integrated and digested with the restriction enzyme EcoT22I was used as a primer type XN7- A single-stranded DNA probe (about 300 bp upstream of NS5B) can be prepared using 4R (SEQ ID NO: 4) and ³² P-dCTP. The probe used is shown in SEQ ID NO: 5.

In addition, a probe for detecting the G3PDH gene was prepared by using a PCR primer of about 250 bp amplified using primers GAPDH S (SEQ ID NO: 6) and GAPDH R (SEQ ID NO: 7) to ³² P- Label with dCTP. Using an ULTRAhyb solution (Ambion) containing one of the above probes, heat at -42 ° C, perform hybridization, wash the filter, and use the bio-imaging analyzer BAS-5000 (Fujifilm). The analysis was performed by.

Figure 8 shows the electrophoresis pattern of agarose gel when ribavirin, mizoribine and SM-108 were added in the presence of HLBI, and the results of hybridization using HCV and G3PDH as probes. 9, shown in Figure 10. The intensity of the detected band was measured with a densitometer, and the remaining amount of replicon RNA was quantified based on the expression of the constantly expressed G3PDH gene. Mizoribine is shown in Table 2 and Figure 12, and SM-108 is shown in Table 3 and Figure 13. As shown in Table 2, Table 3, Figures 12 and 13, both low doses of mizoribine and SM-108 synergistically reduce replicon RNA in the presence of 3 IU / ml of interferon-α. This was confirmed.

Table 2

Table 3

When the natural interferon-α, Sumiferon (HLBI; Sumitomo Pharmaceutical) is administered at 3 to 6 MIU, the blood Cmax becomes approximately 50 to 80 IU / ml 6 to 7 hours after administration, and several IU / ml 2 days after administration. To fall. In the present invention, although usefulness was found in the presence of HLBI at 3 IU / ml, it was considered that the synergistic effect of the concomitant drug was imparted even by administration of high-dose interferon-α. In the case of mizoribine, achieving a blood concentration of approximately 10 μΜ (approximately 2.5 μg / ml) or less would enhance the antiviral effect of interferon. According to FIG. 2, the single dose may range from 50 tng to 100 mg, and the dose was consistent with the blood concentration (1-8 μΜ) at which immune activation was expected. In the case of SM-108, too, the blood concentration was 10 μΜ or less, which was considered to enhance the antiviral effect of Sumiferon. In addition, since the activation of mizoribine immune activity and the enhancement of antiviral effect were achieved at almost the same concentration, the blood concentration at which immunostimulation of SM-108, a prodrug of mizoribine monophosphate, is expected to increase. , Was determined to be 10 μΜ or less.

Also, in cancer treatment, this dose was considered to be effective in eliminating cancer cells.

The dose of mizoribine shown in this study is a dose that has been confirmed to have no clinically significant side effects, and does not cause the hemolytic anemia that is problematic with ribavirin administration It was considered.

In the case of SM-108, the administration of 400 rag / ra ² (about 520 rag) per day slightly reduced the hemoglobin level. The dose at which the blood concentration of SM-108 is less than 10 μΜ is considered to be approximately 20 mg / m ^{2 to} 60 mg / ra ² (clinical cancer: 1985; 31: 757-766). Since this corresponds to a single dose of approximately 26 to 78 mg, it was determined that hemolytic anemia would not occur.

The dose should also be considered to be influenced by factors such as the patient's age, gender, weight, and excretion rate. In the case of SM-108, it has been shown that the blood concentration tends to be longer at higher doses (clinical cancer: 1985; 31: 757-766). It is expected that they will be eliminated from the blood one day later. Therefore, administration of high doses of SM-108 may maintain the concentration range in which synergistic effects can be produced in combination with interferon over a long period of time.

Therefore, SM-108 used in combination with interferon is effective even at a high dose, and it is considered that high dose SM-108 can be administered once a day or every other day. It is thought that the same applies to mizoribine. Therefore, the dose may be any effective concentration that is lower than the concentration that induces side effects, and may be 100 mg or more.

Example 3

ゥ Against Civil Virus Diarrhea Virus (BVDV: Bovine Viral Diarrhea Virus) Multiplication control effect

We investigated the effect of interferon and concomitant drugs on the growth inhibition of bovine viral diarrhea virus (BVDV).

96-well plate in © Shi kidney cells: (MDBK Madin-Darby Bovine Kidney cells, manufactured by Dainippon agents purchased from) were seeded at ^{8xl0 3 cells / Veil, 37 °} C, 5% C0 24 hours at ₂ presence did . Except for the medium (Ham's F-12 medium containing 10% FBS (anti-BVDV antibody negative)), wash the cells with 0.1 ml / well of new medium, and then dilute the BVDV virus (BVDV KS-WS strain, inoculated with Sumitomo Pharma at preparation) was 0. 1 ml / well, incubated for 2 hours at 37 ° C, 5% C0 ₂ presence. Sumiphelon (Sumitomo Pharmaceutical, Lot. F-018), Mizoribine (SIGMA, Lot. 047H4087) and SM-108 (Sumitomo Pharmaceutical, Lot. , Lot. B307E), Adopaferon (inner pharmaceutical Noriyuki mountains, Lot. L008Y01), Bae Gashisu (Roche, Lot. K0013Yl), was added portionwise to 0. 05 m 1 / well, 37 ° C, 5% C0 2 Cultured for 4 days in the presence.

After removing the medium, the cells were washed with the same medium at 0.1 ral / well. After adding a new medium, the cells were added with Alla Marble I (Wako Pure Chemical, Lot. 141937SA). 37 ° C, 5% C0 ₂ was cultured for 4 hours in the presence, fluorometer (Full O b Scan Ascent Type374: Labsystems, Inc.) fluorescence intensity using a (; i ex = 544 nm, λ em = 590 nm) Was measured.

As the BVDV virus grows in cells, the extent of growth of infected cells is reduced. Therefore, as an index of the anti-BVDV activity, the degree of cell growth of the test cells to which the drug and the BVDV virus were added was determined using the control cells without the BVDV virus as a control.

The experimental results are shown in FIGS. 14 and 15. As shown in Figure 14, at the concentration at which mizoribine or SM-108 shows only low anti-BVDV activity with the drug alone, when used in combination with sumiferon, the activity of anti-BVDV activity is more than doubled compared to the activity of sumiferone alone. Sumiferon was found to show a strong combination effect when combined with mizoribine or SM-108. In addition, as shown in FIG. 15, this combined effect was observed not only with Sumiferon, but also with other recombinant interferon-α, Intron A or Advbhaferon.

These results indicate that mizoribine or SM-108 and interferon It was confirmed that the combined effect of is also not only limited to Sumiferon but also to other interferons currently used for the treatment of hepatitis C. Industrial applicability

According to the present invention, a concomitant drug with interferon for diarrhea hepatitis or cancer disease is provided. This drug has no accumulation in the body, activates cell-mediated immunity, and has a synergistic effect when used in combination with interferon. Hepatitis c, chronic myelogenous leukemia, polymyeloma myeloma, myelodysplastic syndrome, polycythemia vera, non-Hodgkin's malignant lymphoma, hepatocellular carcinoma, gastric cancer, superficial bladder, for which interferon is effective It can be a very effective combination for cancer, knee, renal cell, prostate and esophageal cancer. Sequence listing free text

The base sequence described in SEQ ID NO: 1 is a forward primer.

The base sequence described in SEQ ID NO: 2 is Reverse priraer.

The nucleotide sequence set forth in SEQ ID NO: 3 is Probe.

The nucleotide sequence set forth in SEQ ID NO: 4 is primer XN7-4R.

The base sequence described in SEQ ID NO: 5 is Probe.

The nucleotide sequence set forth in SEQ ID NO: 6 is a primer GAPDH S _c

The nucleotide sequence set forth in SEQ ID NO: 7 is a primer GAPDH R _t

Claims

Claim formula (1)

[In the formula, R ¹ represents a hydrogen atom, a nitrogen atom protecting group or a nitrogen atom substituent, and R ² represents a hydrogen atom or a hydroxyl group protecting group. ] A therapeutic agent for co-administration of viral hepatitis or cancer disease with interferon, comprising a compound represented by the formula: or a salt thereof as an active ingredient.

2. The substituent of the nitrogen atom of R ¹ is ribofuranosyl group, 2-hydroxyethoxymethyl group, 2-hydroxymethyl-1,1,3-oxathiolan-15-yl group, 4-hydroxymethylenol-2-cyclopropyl Ninole group, [2-hydroxy-11- (hydroxymethyl) ethoxy] methyl group, tetrahydro-2-furyl group, n-hexylcarbamoyl group, 3-hydroxymethyl-tetrahydro-2-furyl group, 3-hydroxymethylenol 5-hydroxy Seatetrahydro-2-furinole, 3-hydroxymethinole-1,5-dihydroxytetrahydro-2-furinole, 3-hydroxymethyl-4,5-dihydroxy-1-5-methyl-1-tetrahydro-12-frigure or 3- Hydroxymethyl_4-azido-1-tetrahydro-12-furyl group, characterized by the above-mentioned. Combination therapy with interferon described 1.

3. The method according to claim ¹ , wherein R ¹ is a hydrogen atom or a ribofuranosyl group.

The agent according to 1 or 2.

4. The agent according to any one of claims 1 to 3, wherein the interface is interferon- _α .

5. The agent according to any one of claims 1 to 4, wherein the interferon is Sumiferon, Intron II, Adpaferon, Pug Intron or Pegasis.

6. The agent according to any one of claims 1 to 3, wherein the interface is interferon- /.

7. The agent according to claim 1, wherein the viral hepatitis is chronic hepatitis C.

8. The cancer is chronic myelogenous leukemia, multiple myeloma, myelodysplastic syndrome, polycythemia vera, non-Hodgkin's malignant lymphoma, hepatocellular carcinoma, stomach cancer, superficial bladder cancer, knee cancer, renal cell cancer The agent according to any one of claims 1 to 6, wherein the agent is prostate cancer or esophageal cancer.

9. An immunostimulator comprising a compound represented by the formula (1) (wherein R 1 and R 2 are as defined above) or a salt thereof as an active ingredient.

10. The agent according to claim 9, characterized in that it activates cellular immunity.

11. The agent according to claim 9 or 10, which is characterized by inducing interferon-γ production (enhancing production).

12. The substituent of the nitrogen atom of R ¹ is a ribofuranosyl group, a 2-hydroxyethoxymethyl group, a 2-hydroxymethyl-1,3-oxathiolan-1-yl group, or a 4-hydroxymethyl-12-cyclopropenyl group. , [2-Hydroxy_1- (hydroxymethyl) ethoxy] methyl group, Tetrahydro_2-furyl group, η-Hexylcanole vamoyl group, 3-Hydroxymethyl-tetrahydroxy 2-Furinole group, 3-Hydroxy group Cimethyl-5-hydroxy-1-tetrahydro-2-furinole group, 3-hydroxymethinole-4,5-dihydroxy-tetrahydro-2-furinole group, 3-hydroxymethinolate 4,5-dihydroxy-15-methyl-1-tetrahydro-2-furyl Or a 3-hydroxymethyl-4-azidotetrahydro-12-furyl group. Item 9-1.

13. The agent according to claim 9, wherein R ¹ is a hydrogen atom or a lipofuranosyl group.

14. The agent according to claim 9, wherein R ² is a hydrogen atom.

15. The preparation according to any one of claims 10 to 14, which is administered so that the blood concentration after administration is 10 µΜ or less.

16 6. It is specially administered orally so that the blood concentration after administration is 1 μΜ to 8 μΜ. 16. The preparation for oral administration according to claim 15, which is a feature.