JPH0551567B2 - - Google Patents

Description

[Detailed description of the invention]

[Industrial Field of Application] The present invention relates to an antiretroviral agent effective in treating various viral diseases caused by retroviruses. [Prior Art and Problems] As research on viruses continues, treatments for viral diseases are gradually being established. HIV (Human
Immuno deficiency virus), HTLV-I (adult T-cell leukemia virus), and the like are known as retroviruses. Retroviruses contain RNA-dependent DNA synthase (hereinafter referred to as reverse transcriptase) within their virus particles.
It is a virus containing After infecting a host cell, RNA is first transcribed into DNA by reverse transcriptase. This DNA is integrated into the host cell chromosome,
then by host cell RNA synthase
mRNA is synthesized. Various viral proteins are produced from this mRNA. There has been a desire to develop a drug that has an epoch-making therapeutic effect on human diseases caused by this retrovirus. [Means for Solving the Problem] As a result of conducting research on the retrovirus growth inhibiting effect of various herbal medicines, the present inventors discovered that hesperetin extracted and isolated from the herbal medicine Chinpi has a retrovirus growth inhibiting effect. Ta. The present invention is based on this knowledge and is an antiretroviral agent containing hesperetin as an active ingredient. Hesperetin is known to have anti-inflammatory, anti-allergic, and antihypertensive effects. It is also known that hesperetin inhibits the proliferation of DNA viruses, but until now it has no antiretroviral effect against retroviruses that contain reverse transcriptase and propagate by this reverse transcriptase. This was something that was unknown. Hesperetin is a compound shown by the chemical structure shown below, and is commercially available from Tokyo Kasei Kogyo Co., Ltd. [Effects of the Invention] The fact that the antiretroviral agent of the present invention has an antiretroviral effect will be explained with reference to experimental examples. Experimental Example 1 Cells infected with murine leukemia virus (Raussier strain) were cultured, and the method of Nakajima et al. [CONPARATIVE
LEUKEMIA RESEARCH 1973,
LEUKEMOGENESIS，ED.Y.ITO.AND RM
DUTCHER, UNIV.OF TOKYO PRESS
TOKYO/KARGER, BASEL, PP.603−605
(1975)], reverse transcriptase was isolated and purified. Next, a reaction mixture having the following composition was prepared. Γ Reverse transcriptase 1 unit/ml Γ Polyadenylic acid/oligothymidylic acid complex as a template/primer complex [polyadenylic acid (manufactured by Pharmacia): oligothymidylic acid (manufactured by Pharmacia) = 4:1] 2 μg/ml Γ Tris-HCl (pH8.0) 50mM Γ Dithiothreitol 5mM Γ Potassium chloride 50mM Γ Manganese chloride 0.2mM Γ [ ^3H ]deoxythymidine triphosphate 0.01mM (hereinafter abbreviated as [ ^3H ]dTTP)
(400cpm/pmol) Γ Glycerol 15 (v/v)% Γ Purified water Appropriate amount 〓1 unit means that reverse transcriptase is
It is a unit of specific activity that consumes 1 nmol of dNTP (deoxynucleic acid triphosphate). Add 20μ of this reaction mixture to
Add hesperetin and purified water to make 50μ,
The incorporation of radioactivity into the acid-insoluble fraction of [ ³ H]dTTP was measured using a Beckman scintillation counter to determine reverse transcriptase activity, and the inhibition rate at each concentration was calculated. The results are shown in Table 1.

[Table] Experimental example 2 MT-2 cells were prepared at a concentration of 3 x ¹⁰ cells/ml,
It was cultured in RPMI1640 (manufactured by Gibco) medium containing 10% FCS (manufactured by Gibco). 2 days later, MT-
2 After infecting 1 HIV per 200 cells,
The effects of drug addition were observed by dividing into a group to which hesperetin was added at 5 μg/ml and a group to which no hesperetin was added (control group). One week later, the number of viable cells was counted, and it was confirmed that the antiretroviral agent of the present invention suppressed MT-2 cell degeneration caused by HIV by 90% or more. This result confirmed an excellent antiretroviral effect. As described above, the antiretroviral agent of the present invention
Since it has the effect of suppressing the proliferation of retroviruses by inhibiting the reverse transcriptase activity necessary for their proliferation, it can be applied to any retrovirus. Specific examples of retroviruses include leukemia virus, sarcoma virus, breast cancer virus, bisvirus, maedi virus, HIV, HTLV-I, and the like. Next, we conducted an acute toxicity test of oral administration of hesperetin in ddY male mice and Wistar.
When this study was carried out using a strain of male rats, there were no cases of death even when hesperetin was orally administered at a dose of 1 g/Kg. Thus, hesperetin has extremely low toxicity and high safety. Next, the dosage and formulation of hesperetin will be explained. Hesperetin can be administered to animals and humans either neat or with conventional pharmaceutical carriers. The dosage form is not particularly limited and may be selected and used as necessary, including tablets, capsules,
Examples include oral preparations such as granules, fine granules, and powders, and parenteral preparations such as injections and suppositories. In order to exert the desired effect as an oral agent,
Although it varies depending on the patient's age, weight, and severity of the disease,
Usually 100 to 6000 as the weight of hesperetin in adults
It seems appropriate to take mg in divided doses several times a day. In the present invention, oral preparations such as tablets, capsules, and granules are manufactured according to conventional methods using, for example, starch, lactose, sucrose, mannitol, carboxymethyl cellulose, cornstarch, inorganic salts, and the like. In addition to the excipients mentioned above, binders, disintegrants, surfactants, lubricants, fluidity promoters, flavoring agents, coloring agents, fragrances, and the like can be used in this type of preparation as appropriate. Specific examples of each are shown below. [Binder] Starch, dextrin, gum arabic powder, gelatin, hydroxypropyl starch, methylcellulose, sodium carboxymethylcellulose, hydroxypropylcellulose, crystalline cellulose, ethylcellulose, polyvinylpyrrolidone, macrogol. [Disintegrant] Starch, hydroxypropyl starch, sodium carboxymethyl cellulose, calcium carboxymethyl cellulose, carboxymethyl cellulose, low substituted hydroxypropyl cellulose. [Surfactant] Sodium lauryl sulfate, soybean lecithin, sucrose fatty acid ester, polysorbate 80. [Lubricant] Talc, waxes, hydrogenated vegetable oil, sucrose fatty acid ester, magnesium stearate, calcium stearate, aluminum stearate,
Polyethylene glycol. [Fluidity promoter] Light anhydrous silicic acid, dry aluminum hydroxide gel, synthetic aluminum silicate, magnesium silicate. Hesperetin can also be administered as a suspension, emulsion, syrup, or elixir, and these various dosage forms may contain flavoring agents and coloring agents. In order to exert the desired effect as a parenteral agent, it depends on the age, weight, and severity of the disease of the patient, but in general, adults should take 1 dose of hesperetin per day.
Intravenous injections, intravenous drips, subcutaneous injections, and intramuscular injections of up to ~100 mg are considered appropriate. This parenteral preparation is manufactured according to a conventional method, and diluents generally include distilled water for injection, physiological saline, aqueous glucose solution, vegetable oil for injection, sesame oil, mustard oil, soybean oil, corn oil, propylene glycol, polyethylene glycol, etc. can be used. Furthermore, a bactericide, a preservative, and a stabilizer may be added as necessary. In addition, from the standpoint of stability, this parenteral preparation is frozen after being filled into vials, etc.
The liquid formulation can also be reconstituted from the lyophilizate immediately prior to use by removing moisture by conventional lyophilization techniques. Furthermore, isotonizing agents, stabilizers, preservatives, soothing agents, etc. may be added as appropriate. Other parenteral preparations include liquid preparations for external use, liniments such as ointments, and suppositories for intrarectal administration, which are manufactured according to conventional methods. Example 1 Crystalline cellulose 84.5g Magnesium stearate 0.5g Carboxymethyl cellulose calcium 5g Hesperetin 10g Total 100g According to the above recipe, a part of and was mixed uniformly, compressed and molded, and then crushed, and the remaining amount of and was The mixture was added, mixed, and compressed using a tablet machine to obtain tablets each weighing 200 mg. One tablet of this tablet contains 20 mg of hesperetin, and adults should take 5 to 7 tablets a day in several doses. Example 2 Corn starch 84g Magnesium stearate 0.5g Calcium carboxymethyl cellulose 5g Light anhydrous silicic acid 0.5g Hesperetin 10g Total 100g Mix ~ uniformly according to the above recipe, compress and mold with a compression molding machine, and then crush with a crusher death,
Granules were obtained by sieving. 1 g of this granule contains 100 mg of hesperetin, and adults should take 1 to 2.5 g per day in several doses. Example 3 Cornstarch 89.5g Light anhydrous silicic acid 0.5g Hesperetin 10g Total 100g Mix ~ uniformly according to the above recipe,
200 mg was filled into No. 2 capsules. Each capsule contains hesperetin.
It contains 20 mg, and adults should take 5 to 7 capsules a day in several doses. Example 4 Distilled water for injection Appropriate amount Glucose 200mg Hesperetin 10mg Total amount 15ml After dissolving and in distilled water for injection, 5
The mixture was injected into a ml ampoule and autoclaved at 121°C for 15 minutes to obtain an injection.

Claims (1)

[Claims] 1.An antiretroviral agent containing hesperetin as an active ingredient.