WO2022196614A1 - Agent for treating or preventing chagas disease - Google Patents

Agent for treating or preventing chagas disease Download PDF

Info

Publication number
WO2022196614A1
WO2022196614A1 PCT/JP2022/011217 JP2022011217W WO2022196614A1 WO 2022196614 A1 WO2022196614 A1 WO 2022196614A1 JP 2022011217 W JP2022011217 W JP 2022011217W WO 2022196614 A1 WO2022196614 A1 WO 2022196614A1
Authority
WO
WIPO (PCT)
Prior art keywords
group
coptisine
compound
chagas disease
same
Prior art date
Application number
PCT/JP2022/011217
Other languages
French (fr)
Japanese (ja)
Inventor
謙二 平山
雄基 田山
修作 水上
かつ子 小松
一文 當銘
Original Assignee
国立大学法人 長崎大学
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by 国立大学法人 長崎大学 filed Critical 国立大学法人 長崎大学
Publication of WO2022196614A1 publication Critical patent/WO2022196614A1/en

Links

Images

Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/47—Quinolines; Isoquinolines
    • A61K31/4738—Quinolines; Isoquinolines ortho- or peri-condensed with heterocyclic ring systems
    • A61K31/4745—Quinolines; Isoquinolines ortho- or peri-condensed with heterocyclic ring systems condensed with ring systems having nitrogen as a ring hetero atom, e.g. phenantrolines
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P33/00—Antiparasitic agents
    • A61P33/02—Antiprotozoals, e.g. for leishmaniasis, trichomoniasis, toxoplasmosis

Definitions

  • the present invention relates to a compound effective in treating or preventing Chagas disease, preferably a compound effective in treating or preventing acute and/or chronic Chagas disease. Furthermore, it relates to a therapeutic or prophylactic agent for Chagas disease containing the compound.
  • Chagas disease caused by the protozoan Trypanosoma cruzi, is a tropical disease (infectious disease) transmitted by the triatomine bug, which is widespread in Latin America and the Caribbean.
  • the disease stage is roughly divided into an acute stage and a chronic stage.
  • An object of the present invention is to provide a therapeutic or preventive agent for Chagas disease containing a compound effective for the treatment or prevention of Chagas disease.
  • the compound is one or more selected from the group consisting of coptisine and dehydrocorridarin or a salt thereof, or a compound represented by general formula (I) or a salt thereof, [1] or [ 2].
  • the agent according to any one of [1] to [5] which is further characterized by being effective for the treatment or prevention of acute and chronic Chagas disease.
  • a compound effective for the treatment or prevention of Chagas disease and an agent for the treatment or prevention of Chagas disease containing the compound can be provided.
  • FIG. 2 is a diagram showing the results of evaluation of antitrypanosomal activity (crude drug extract/crude drug-derived compound), which is primary screening.
  • FIG. 2 is a diagram showing the results of a cytotoxicity test (herbal drug extract and herbal drug-derived compound), which is a secondary screening.
  • FIG. 2 shows the results of measuring the antitrypanosomal activity of the compounds of the present invention.
  • FIG. 3 shows the results of IC 50 measurements on intracellular amastigotes for the compounds of the present invention.
  • FIG. 2 shows the results of measurement of cytotoxicity against NBMH cells for compounds of the present invention.
  • FIG. 2 shows the results of measuring the cytotoxicity of the compounds of the present invention against HuH28 cells.
  • A Images of in vivo luminescence intensity after intraperitoneal administration of coptisine to mice.
  • Chaagas disease is a tropical disease (infectious disease) caused by Trypanosoma cruzi protozoa and transmitted by an insect called assassin bug.
  • acute phase Chagas disease usually refers to a period of several weeks to several months immediately after infection, and refers to asymptomatic symptoms or symptoms such as fever, fatigue, itching, headache, and diarrhea.
  • red swelling and lumps appearing on the skin where the protozoa have invaded chagoma
  • Romagna sign due to bites by kissing bugs or contact with the feces of kissing bugs.
  • This symptom usually disappears spontaneously within a few weeks, but the protozoa may remain in the body and enter a long incubation period.
  • the state in which trypomastigotes are present in the blood and the state in which amastigotes are present in the cells may be referred to as the "acute phase” or a state simulating the "acute phase.” .
  • the “chronic phase” of Chagas disease refers to cardiac complications (cardiac hypertrophy, heart failure, changes in heart rate, arrhythmia, apical artery aneurysm/thrombosis, cardiopulmonary arrest (sudden death), etc.), intestinal complications (enlargement of the esophagus or colon (megaesophagus/megacolon), difficulty eating or excretion due to these) refers to the state. These symptoms may be seen in about 10 to 30% of infected people.
  • the state in which amastigotes proliferate in cells may be referred to as a "chronic phase” or a state simulating a "chronic phase.”
  • the therapeutic or preventive agent (composition) for Chagas disease of the present invention is described in detail below.
  • the therapeutic or preventive agent (composition) for Chagas disease of the present invention is a compound or a salt thereof selected from the group consisting of coptisine, berberine, palmatine, epiberberine, berberrubine and ditetrahydrocoptisine, or the following general formula:
  • Compound represented by (I) in this specification, may be referred to as "analogous compound” or a salt thereof (wherein R 1 , R 2 , R 3 and R 4 may be the same or different, hydrogen atom, C 1-6 alkyl group, C 2-6 alkenyl group, C 2-6 alkynyl group, C 1 -6 haloalkyl group, C 3-6 cycloalkyl group, preferably hydrogen atom, C 1-6 alkyl group or C 3-6 cycloalkyl group, more preferably hydrogen atom or C 1-6 is an alkyl group.
  • R 1 may form a 5- or 6-membered ring together with R 2 and the adjacent oxygen atom
  • R 3 may form a 5- or 6-membered ring together with R 4 and the adjacent oxygen atom
  • R 5 , R 6 , R 7 and R 8 may be the same or different and are C 1-6 alkyl group, C 2-6 alkenyl group, C 2-6 alkynyl group, halogen atom, C 1-6 haloalkyl group , a C 3-6 cycloalkyl group, a C 1 -C 6 alkoxy group, a carboxyl group or an amino group, preferably a C 1-6 alkyl group, a halogen atom or a C 3-6 cycloalkyl group.
  • the nitrogen atom is a C 1-6 alkyl group or a halogen atom.
  • the therapeutic or preventive agent (composition) for Chagas disease of the present invention is a compound or a salt thereof selected from the group consisting of coptisine, berberine, palmatine, epiberberine, berberrubine and ditetrahydrocoptisine, or the above
  • the compound represented by general formula (I) or a salt thereof is contained as an active ingredient.
  • the "C 1 -C 6 alkyl group” includes, for example, methyl group, ethyl group, propyl group, isopropyl group, butyl group, isobutyl group, secondary butyl group, tertiary butyl group, pentyl group and isopentyl group. , a tertiary pentyl group, a neopentyl group, a 2,3-dimethylpropyl group, and other linear or branched alkyl groups, but are not limited thereto.
  • the "C 2 -C 6 alkenyl group” includes, for example, linear groups such as vinyl group, allyl group, isopropenyl group, 1-butenyl group, 2-butenyl group, 2-methyl-2-propenyl group, and the like. or branched alkenyl groups, but are not limited thereto.
  • the "C 2 -C 6 alkynyl group” includes, for example, ethynyl group, 1-propynyl group, 2-propynyl group, 1-butynyl group, 2-butynyl group, 3-butynyl group, 3-methyl- Linear or branched alkynyl groups such as 1-propynyl group and phenylethynyl group may be mentioned, but not limited thereto.
  • halo means "halogen atom"
  • the halogen atom indicates a chlorine atom, a bromine atom, an iodine atom or a fluorine atom.
  • C 3 -C 6 cycloalkyl group includes, but is not limited to, cyclic alkyl groups such as cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl groups.
  • the "C 1 -C 6 alkoxy group” includes, for example, methoxy group, ethoxy group, propoxy group, isopropoxy group, butoxy group, secondary butoxy group, tertiary butoxy group, pentyloxy group, isopentyloxy linear or branched alkoxy groups such as neopentyloxy groups, etc., but are not limited thereto.
  • Preferred salts for use in the present invention include, for example, inorganic acid salts such as hydrochlorides, sulfates, nitrates and phosphates, acetates, fumarates, maleates, oxalates, methanesulfonates, and benzenesulfones.
  • Salts, organic acid salts such as p-toluenesulfonate, and salts with inorganic or organic bases such as sodium ion, potassium ion, calcium ion, and trimethylammonium can be exemplified, but are not limited to these.
  • the agent for treating or preventing Chagas disease of the present invention contains a compound or a salt thereof selected from the group consisting of coptisine, berberine and palmatine, or a compound represented by the general formula (I) or a salt thereof. More preferably, the agent for treating or preventing Chagas disease of the present invention contains coptisine or a salt thereof, more preferably coptisine hydrochloride, but is not limited thereto.
  • the therapeutic or prophylactic agent for Chagas disease of the present invention includes, but is not limited to, dehydrocorridarin nitrate or palmatine hydrochloride.
  • Another preferred embodiment of the agent for treating or preventing Chagas disease of the present invention includes, but is not limited to, one or more selected from the following six compounds or salts thereof.
  • a compound or a salt thereof selected from the group consisting of coptisine, berberine, palmatine, epiberberine, berberrubine and ditetrahydrocoptisine, or the general formula (I)
  • a plurality of represented compounds or salts thereof eg, 1 or more, 2 or more, 3 or more, 1 to 5, or 2 to 4, etc. may be included.
  • the agent for treating or preventing Chagas disease of the present invention is useful not only for the treatment or prevention of acute phase Chagas disease, but also for the treatment or prevention of chronic phase Chagas disease.
  • the agent of the present invention is effective in treating or preventing acute phase symptoms of Chagas disease, and more preferably, the agent of the present invention is effective in treating or preventing acute phase symptoms and chronic phase symptoms.
  • the content of the above compound in the agent or composition for treating or preventing Chagas disease of the present invention is not particularly limited, but in 100% by mass of the agent or composition, for example, in the range of about 50% to about 100% by mass. It may be present, preferably in the range of about 75% to about 100% by weight, more preferably about 90% to about 100% by weight.
  • the agent or composition of the present invention can be formulated by appropriately blending the above-described compound of the present invention or a salt thereof and, if desired, pharmaceutically acceptable carriers, additives and the like.
  • Formulation methods and techniques for this purpose have been well established in the past, and may be followed.
  • pharmaceuticals specifically oral agents such as tablets, coated tablets, pills, powders, granules, capsules, liquids, suspensions, emulsions, injections, infusions, suppositories, ointments
  • Parenteral agents such as patches can be used, but are not limited to these.
  • the mixing ratio of the carrier or additive may be set as appropriate based on the range normally employed in the field of pharmaceuticals and the like.
  • Pharmaceutically acceptable carriers or additives are not particularly limited, but examples of carriers include various carriers such as aqueous or oily bases, and examples of aqueous carriers include water, physiological saline, and ethanol. , glycerin, polyethylene glycol, propylene glycol, methyl cellulose, hydroxypropylmethyl cellulose, hydroxypropyl cellulose, polyvinylpyrrolidone, polyacrylic acid, polysaccharide gum-based natural polymers, etc.
  • oily carriers include vaseline, squalane, Examples include, but are not limited to, oils such as paraffin and waxes.
  • additives examples include enzymes, pH adjusters, preservatives, bactericides, excipients, binders, disintegrants, lubricants, antioxidants, brighteners, flavors, sweeteners, acidulants, seasonings. , bittering agents, emulsifiers, thickeners, stabilizers, gelling agents, coloring agents, flavoring agents, flavoring agents, and the like. Techniques for these additives are well established in the prior art, and may be followed in the present invention. Furthermore, the composition of the present invention may contain any ingredient known in the fields of medicine, pharmacy, veterinary medicine, food, etc., as long as the effects of the present invention are not lost.
  • the compound of the present invention can be administered orally or parenterally.
  • the administration form include oral administration, topical administration to the eye, intravenous administration, transdermal administration, etc., and if necessary, pharmaceutically acceptable additives are appropriately selected and used, and a dosage form suitable for the administration form is used.
  • can be formulated into For example, when the compound of the present invention or a salt thereof is administered orally or by injection, it can be determined depending on the age and body weight of the ingestor, symptoms, administration time, dosage form, administration method, drug combination, and the like.
  • the intake is preferably about 60 to 600 mg/mL per adult (about 60 kg) per day. If administered by injection, it is preferably set to inject up to about 60 mg/mL.
  • the amount of the compound or its salt to be applied can be appropriately selected according to the skin area to be applied. is preferably about 0.01-5 mg, more preferably about 0.05-1 mg. It is preferable to administer or apply the above administration dose once or in several divided doses per day.
  • the compound of the present invention e.g., a compound or a salt thereof selected from the group consisting of coptisine, berberine, palmatine, epiberberine, berberrubine and ditetrahydrocoptisine, or the above Compounds represented by general formula (I), etc.
  • the compound of the present invention e.g., a compound or a salt thereof selected from the group consisting of coptisine, berberine, palmatine, epiberberine, berberrubine and ditetrahydrocoptisine, or the above Compounds represented by general formula (I), etc.
  • Methods for encapsulating compounds are well established in the art and may be followed.
  • Cukabit[n]uril (usually n is 5 to 10) is a barrel-shaped compound (see below) with n glycourils, which are structural units, arranged in a ring and having a hydrophobic vacancy inside. be.
  • Cukabit [n] uril is known to selectively show a very high interaction with compounds having a naphthalene structure, a cholesterol structure, etc. compounds selected from the group consisting of epiberberine, berberrubine and ditetrahydrocoptisine or salts thereof).
  • a method for producing such a cukabit[n]uril is described, for example, in JP-A-2012-246239, and the method may be followed. Alternatively, a commercially available product (Merck) may be used.
  • n is preferably 6-9, more preferably 7-8.
  • Cucurbit[n]uril is synthesized by the method described in JP-A-2012-246239.
  • the compounds of the present invention can be incorporated into cukabit[n]urils by mixing, for example, 2:1 to 1:2 in a solvent suitable for inclusion (such as an organic solvent). . Water solubility is improved by inclusion of the compound of the present invention.
  • Cukabit is sometimes written as cucurbit, but it is the same thing.
  • Another embodiment of the compound preferably encompassing the compound of the present invention includes dextrins such as ⁇ -cyclodextrin, ⁇ -cyclodextrin, ⁇ -cyclodextrin, and cluster dextrin, as well as crown ethers (e.g., 12-crown -4, 15-crown-5, 18-crown-6, etc.), fullerene (e.g., C60 fullerene, C70 fullerene, etc.), calix[m]arene (m: 3 to 20, preferably n: 4 to 8 ), pillar arenes (eg, 1,4-dimethoxy pillar[5]arene), and the like, but are not limited thereto.
  • the effect of the present invention treatment of acute and / or chronic Chagas disease or preventive effect, etc.
  • the effect of the present invention treatment of acute and / or chronic Chagas disease or preventive effect, etc.
  • ⁇ Compound, Reagent> Crude drug-derived compounds and crude drug extracts: those provided by the Institute of Japanese and Chinese Medicine, University of Toyama as a Japanese and Chinese drug library were used.
  • New Born Mouse Heart (NBMH) cells those provided by Nekken Bioresource Center, Nagasaki University were used.
  • HuH28 cells those provided by Chulabhorn International School of Medicine, Thammasat University were used.
  • D-luciferin (MB000102-R70170-2) Syd Labs
  • Coptisine chloride (C685500) Toronto Research Chemicals
  • Isoflurane (095-06573) Wako Pure Chemical
  • Cyclophosphamide monohydrate (C0768) SIGMA-ALDRICH.
  • Example 1 Primary screening using antitrypanosomal activity as an index 1-1 Experimental method Trypomastigotes ( 3 x 104 cells/well) in which luciferase gene was incorporated in a 96-well white plate and fibers isolated from mouse heart Blast cells (NBMH cells) (5 ⁇ 10 4 cells/well) were seeded. Either the herbal drug-derived compound (20 ⁇ M) or herbal drug extract (20 ⁇ g/mL) described in Tables 1 and 2 above was added to each well and cultured for 72 hours. NP40 was added as a positive control, and only the solvent in which the drug was dissolved was added to the negative control. In addition, wells were provided to add benznidazole, an existing drug. The volume per well was 100 ⁇ l.
  • Example 1 After culturing for 72 hours, 100 ⁇ l of 0.6% NP40 was added to disrupt the cells. The disrupted cells and medium were transferred to a 1.5 mL tube and centrifuged (12,000 rpm, 15 minutes). After centrifugation, 100 ⁇ l of luciferin (Picagene) was added to 50 ⁇ l of the supernatant, and the luminescence intensity (544 nm/590 nm) was measured with a plate reader (measurement time: 1 second). Taking the luminescence suppression rate of the negative control as 0% and the luminescence suppression rate of the positive control as 100%, the luminescence suppression rate of each compound or extract was calculated. The experimental method of Example 1 was based on the method described in Bettiol et al. (PLoS Neglected Tropical Diseases, 2009, 3(2): e384.) effect on amastigotes can be confirmed.
  • Example 2 Secondary Screening Using Cytotoxicity as Index 2-1 Experimental Method NBMH cells (1 ⁇ 10 4 /well) and HuH28 cells (1 ⁇ 10 4 /well) were seeded in a 96-well black plate. Either a crude drug-derived compound (20 ⁇ M) or a crude drug extract (20 ⁇ g/mL) was added to each well and cultured for 72 hours. As a positive control, NP40 was added, and as a negative control, only solvent in which the drug was dissolved was added. In addition, wells were provided to add benznidazole, an existing drug. The volume per well was 100 ⁇ l. After culturing for 72 hours, 10 ⁇ l of Alamar Blue reagent was added, and the cells were further incubated for 4 hours. After that, the fluorescence intensity was measured with a plate reader (measurement time 0.1 sec).
  • FIG. (A) is the result of the crude drug extract
  • (B) is the result of the crude drug-derived compound.
  • the compounds or extracts that showed a decrease in luminescence intensity of 80% or more in the primary screening three of them, Coptisine Chloride, Dehydrocorydaline Nitrate, and Palmatine Chloride, showed fluorescence. No decrease in strength was observed, indicating no cytotoxicity. Since these three compounds had a common structure, three more analogues (berberrubine, epiberberine, and di-tetrahydrocoptisine) were added. Six compounds were tested for activity against trypanosomes.
  • Example 3 Measurement of IC50 of Antitrypanosoma Activity 3-1 Experimental Method The antitrypanosoma activity of the above six compounds was measured by the same experimental method as in Example 1.
  • the concentrations of coptisine, dehydrocorridarin and palmatine were 40 ⁇ M, 33 ⁇ M, 20 ⁇ M, 16.5 ⁇ M, 10 ⁇ M, 8.25 ⁇ M, 5.0 ⁇ M, 4.13 ⁇ M, 2.5 ⁇ M, 2.06 ⁇ M and 1.25 ⁇ M.
  • Berberrubine, epiberberine and ditetrahydrocoptisine concentrations were 50 ⁇ M, 25 ⁇ M, 12.5 ⁇ M, 6.25 ⁇ M, 3.125 ⁇ M and 1.6 ⁇ M.
  • Example 4 Measurement of IC50 of Activity against Intracellular Amastigote 4-1 Experimental Method Trypomastigotes expressing luciferase gene and NBMH cells were seeded in a 25 cm 2 flask at a ratio of 2:1. The medium was MEM+1% NBCS to enhance intracellular infection. After 24 hours, the cells were washed twice with PBS, added with MEM+10% NBCS, and further cultured in an incubator for 24 hours. After that, trypsin treatment was performed to detach NBMH cells containing Trypanosoma cruzi (T. cruzi). NBMH cells (5 ⁇ 10 4 /well) containing T. cruzi were seeded in a 96-well white plate.
  • T. cruzi Trypanosoma cruzi
  • One of the above six compounds was added to each well and cultured for 72 hours.
  • the concentrations of coptisine, dehydrocorridarin and palmatine were 40 ⁇ M, 33 ⁇ M, 20 ⁇ M, 16.5 ⁇ M, 10 ⁇ M, 8.25 ⁇ M, 5.0 ⁇ M, 4.13 ⁇ M, 2.5 ⁇ M, 2.06 ⁇ M and 1.25 ⁇ M.
  • Berberrubine, epiberberine and ditetrahydrocoptisine concentrations were 50 ⁇ M, 25 ⁇ M, 12.5 ⁇ M, 6.25 ⁇ M, 3.125 ⁇ M and 1.6 ⁇ M.
  • NP40 was added, and as a negative control, only solvent in which the drug was dissolved was added.
  • Benznidazole an existing drug.
  • the volume per well was 100 ⁇ l.
  • the concentrations of coptisine hydrochloride, dehydrocorridarin nitrate and palmatine hydrochloride are the same as in Example 3.
  • the concentrations of epiberberine, berberrubine and ditetrahydrocoptisine were 100 ⁇ M, 50 ⁇ M, 25 ⁇ M, 12.5 ⁇ M, 6.25 ⁇ M and 3.125 ⁇ M.
  • 100 ⁇ l of 0.6% NP40 was added to disrupt the cells. The disrupted cells and medium were transferred to a 1.5 ml tube and centrifuged (12,000 rpm, 15 minutes).
  • Example 4 100 ⁇ l of luciferin (Picagene) was added to 50 ⁇ l of the supernatant, and the luminescence intensity (544 nm/590 nm) was measured with a plate reader (measurement time: 1 second). Taking the luminescence suppression rate of the negative control as 0% and the luminescence suppression rate of the positive control as 100%, the luminescence suppression rate of each compound or extract was calculated.
  • the experimental method of Example 4 was described by Rycker et al. (PLoS Negl Trop Dis. 2016 Apr 15;10(4):e0004584) and Alonso-Padilla et al. ) was used as a reference. This experimental system can confirm the effect of trypanosome-infected cells on proliferating amastigotes.
  • Example 5 Evaluation of cytotoxicity against NBMH cells 5-1 Experimental method For the above six compounds, the same experimental method as in Example 2 except that the cells used in the experimental method of Example 2 were changed to one type of NBMH cells. was evaluated for cytotoxicity against NBMH cells. The concentration of each compound is the same as in Example 4.
  • Example 6 Evaluation of cytotoxicity against HuH28 cells 6-1 Experimental method For the above six compounds, the same experimental method as in Example 2 except that the cells used in the experimental method of Example 2 were changed to one type of HuH28 cells. was evaluated for cytotoxicity against HuH28 cells. The concentration of each compound is the same as in Example 4.
  • Example 7 Effect of coptisine on mice 7-1
  • Experimental method 1 ⁇ 10 4 cells of T. cruzi (trypomastigotes type, Tulahuen strain) were intraperitoneally administered to 7- to 8-week-old female BALB/c mice. The drug was administered for 5 consecutive days from day 4 to day 8 of infection.
  • Coptisine was administered intraperitoneally twice daily at a dose of 30 mg/kg. Thereafter, immunosuppression was performed by intraperitoneal administration of 200 mg/kg of cyclophosphamide monohydrate on days 31, 34, and 37 of infection.
  • Benznidazole 100 mg/kg was orally administered once a day as a positive control.
  • As a negative control 1% PBS was orally administered once a day.
  • Luminescence intensity was measured on day 14 of infection, day 28 of infection, and day 40 of infection after immunosuppression, including the day before the start of drug administration (day 3 of infection) and the day after the end of drug administration (day 9 of infection). I went there a total of 5 times. Luminescence intensity was measured by intraperitoneally administering D-luciferin at 150 mg/kg, inhaling isoflurane for 10 minutes, and using the luminescence in vivo imaging system IVIS Lumina II.
  • FIG. Figure 7-1 is an image of the in vivo luminescence intensity of each group of mice
  • Figure 7-2 is the result of quantification of the luminescence intensity.
  • Intraperitoneal administration of coptisine to trypanosome-infected mice decreased the luminescence intensity.
  • coptisine attenuated the fluorescence intensity to the same level as or more than that of benznidazole on day 40 of infection after immunosuppression, which is a chronic phase model. This result indicates that coptisine is effective in the acute and chronic stages of trypanosomal infection.
  • Example 8 (improvement of water solubility by inclusion of compound) Copticine hydrochloride or berberine hydrochloride was added to cukabit[7]uril by mixing cukabit[7]uril and coptisine hydrochloride or berberine hydrochloride purchased from Merck & Co. 1:1 in water. subsumed. Inclusion improved the water solubility of coptisine hydrochloride or berberine hydrochloride. Coptisine hydrochloride or berberine hydrochloride with such improved water solubility is suitable for, for example, in vivo administration (oral administration, injection administration), and thus is useful.

Abstract

The present invention provides an agent for treating or preventing Chagas disease, characterized by containing one or more compounds selected from the group consisting of coptisin, berberine, palmatine, epiberberine, berberrubine, and ditetrahydrocoptisin, or a salt thereof, or a compound represented by general formula (I) or a salt thereof.

Description

シャーガス病治療又は予防剤Chagas disease therapeutic or prophylactic agent
　本発明は、シャーガス病の治療又は予防に有効な化合物、好ましくは、急性期及び／又は慢性期のシャーガス病治療又は予防に有効な化合物に関する。さらに、該化合物を含むシャーガス病治療又は予防剤に関する。 The present invention relates to a compound effective in treating or preventing Chagas disease, preferably a compound effective in treating or preventing acute and/or chronic Chagas disease. Furthermore, it relates to a therapeutic or prophylactic agent for Chagas disease containing the compound.
　クルーズトリパノソーマ原虫によって引き起こされるシャーガス病は、ラテンアメリカやカリブ諸国で蔓延するサシガメという昆虫に媒介される熱帯病（感染症）である。病期は大きく急性期と慢性期に分かれる。現在使用されている治療薬であるニフルチモックス、ベンズニダゾールは様々な副作用があり、慢性期には十分な効果が得られない。そのため、副作用が少なく、慢性期にも効果を示す新規な薬剤が求められている。 Chagas disease, caused by the protozoan Trypanosoma cruzi, is a tropical disease (infectious disease) transmitted by the triatomine bug, which is widespread in Latin America and the Caribbean. The disease stage is roughly divided into an acute stage and a chronic stage. Currently used therapeutic agents, nifurtimox and benznidazole, have various side effects and are not sufficiently effective in the chronic phase. Therefore, there is a demand for a novel drug that has few side effects and is effective even in the chronic stage.
　また、オウバク（黄柏）、オウレン（黄連）に含まれることが知られるコプチシン、ベルベリン、パルマチン等の化合物は、マラリアやトキソプラズマ症の治療に有効なことが知られている（特許文献１）。しかしながら、これらの化合物がシャーガス病治療に有効なこと、特に、急性期及び／又は慢性期のシャーガス病治療に有効であることは全く知られていない。 In addition, compounds such as coptisine, berberine, and palmatine, which are known to be contained in Phellodendron bark and Japanese coptis, are known to be effective in treating malaria and toxoplasmosis (Patent Document 1). However, it is completely unknown that these compounds are effective in treating Chagas' disease, particularly in treating acute and/or chronic Chagas' disease.
特開２０１８－１７７７３３号公報JP 2018-177733 A
　本発明はシャーガス病の治療又は予防に有効な化合物を含むシャーガス病治療又は予防剤を提供することを目的とする。 An object of the present invention is to provide a therapeutic or preventive agent for Chagas disease containing a compound effective for the treatment or prevention of Chagas disease.
　本発明者等は、上記課題を解決するために鋭意研究を重ねた結果、これまでマラリアやトキソプラズマ症の治療に有効なことが知られていたコプチシン、ベルベリン、パルマチン等の化合物又はその類縁化合物が、驚くべきことに、シャーガス病の治療又は予防に有効であること、さらに、これらの化合物が急性期及び／又は慢性期のシャーガス病治療又は予防にも有効であることを見出し、さらに研究を続け、本発明を完成させるに至った。 As a result of intensive studies to solve the above problems, the present inventors have found that compounds such as coptisine, berberine, and palmatine, which have been known to be effective in the treatment of malaria and toxoplasmosis, or analogous compounds thereof, Surprisingly, it was found that these compounds are effective in treating or preventing Chagas disease, and that these compounds are also effective in treating or preventing acute and/or chronic Chagas disease. , have completed the present invention.
　すなわち、本発明は以下に関する。
［１］　コプチシン、ベルベリン、パルマチン、エピベルベリン、ベルベルルビン及びジテトラハイドロコプチシンからなる群から選択される１種以上の化合物若しくはその塩、又は
　下記の一般式（Ｉ）で表される化合物若しくはその塩
Figure JPOXMLDOC01-appb-C000003
 （式中、Ｒ１、Ｒ２、Ｒ３及びＲ４は、同一又は異なってもよく、水素原子、Ｃ１－６アルキル基、Ｃ２－６アルケニル基、Ｃ２－６アルキニル基、Ｃ１－６ハロアルキル基、Ｃ３－６シクロアルキル基であり、
　Ｒ１はＲ２及び隣接する酸素原子と一緒に５又は６員環を形成してもよく、
　Ｒ３はＲ４及び隣接する酸素原子と一緒に５又は６員環を形成してもよく、
　Ｒ５、Ｒ６、Ｒ７及びＲ８は、同一又は異なってもよく、Ｃ１－６アルキル基、Ｃ２－６アルケニル基、Ｃ２－６アルキニル基、ハロゲン原子、Ｃ１－６ハロアルキル基、Ｃ３－６シクロアルキル基、Ｃ１－Ｃ６アルコキシ基、カルボキシル基又はアミノ基であり、
　３箇所の点線は、それぞれ、同一又は異なってもよく、二重結合（＝）又は単結合（－）であり、
　ｍ、ｎ、ｐ及びｑは、同一又は異なってもよく、０、１又は２である）
　を含有することを特徴とする、シャーガス病治療又は予防剤。
［２］　化合物が、コプチシン、デヒドロコリダリン及びパルマチンからなる群から選択される１種以上若しくはその塩又は一般式（Ｉ）で表される化合物若しくはその塩から選択されることを特徴とする、［１］に記載の剤。
［３］　化合物がコプチシン及びデヒドロコリダリンからなる群から選択される１種以上若しくはその塩又は一般式（Ｉ）で表される化合物若しくはその塩であることを特徴とする、［１］又は［２］に記載の剤。
［４］　化合物がコプチシン又はその塩であることを特徴とする、［１］～［３］のいずれかに記載の剤。
［５］　急性期のシャーガス病の治療又は予防に有効であることをさらに特徴とする、［１］～［４］のいずれかに記載の剤。
［６］　急性期及び慢性期のシャーガス病の治療又は予防に有効であることをさらに特徴とする、［１］～［５］のいずれかに記載の剤。
［７］　コプチシン、ベルベリン、パルマチン、エピベルベリン、ベルベルルビン及びジテトラハイドロコプチシンからなる群から選択される１種以上の化合物若しくはその塩又は
　下記の一般式（Ｉ）で表される化合物若しくはその塩
Figure JPOXMLDOC01-appb-C000004
 （式中、Ｒ１、Ｒ２、Ｒ３及びＲ４は、同一又は異なってもよく、水素原子、Ｃ１－６アルキル基、Ｃ２－６アルケニル基、Ｃ２－６アルキニル基、Ｃ１－６ハロアルキル基、Ｃ３－６シクロアルキル基であり、
　Ｒ１はＲ２及び隣接する酸素原子と一緒に５又は６員環を形成してもよく、
　Ｒ３はＲ４及び隣接する酸素原子と一緒に５又は６員環を形成してもよく、
　Ｒ５、Ｒ６、Ｒ７及びＲ８は、同一又は異なってもよく、Ｃ１－６アルキル基、Ｃ２－６アルケニル基、Ｃ２－６アルキニル基、ハロゲン原子、Ｃ１－６ハロアルキル基、Ｃ３－６シクロアルキル基、カルボキシル基又はアミノ基であり、
　３箇所の点線は、それぞれ、同一又は異なってもよく、二重結合（＝）又は単結合（－）であり、
　ｍ、ｎ、ｐ及びｑは、同一又は異なってもよく、０、１又は２である）
　を含有することを特徴とする、シャーガス病治療又は予防用組成物。 That is, the present invention relates to the following.
[1] One or more compounds or salts thereof selected from the group consisting of coptisine, berberine, palmatine, epiberberine, berberrubine and ditetrahydrocoptisine, or compounds represented by the following general formula (I) or its salt
Figure JPOXMLDOC01-appb-C000003
 (wherein R 1 , R 2 , R 3 and R 4 may be the same or different, hydrogen atom, C 1-6 alkyl group, C 2-6 alkenyl group, C 2-6 alkynyl group, C 1 -6 haloalkyl group, C 3-6 cycloalkyl group,
R 1 may form a 5- or 6-membered ring together with R 2 and the adjacent oxygen atom;
R 3 may form a 5- or 6-membered ring together with R 4 and the adjacent oxygen atom;
R 5 , R 6 , R 7 and R 8 may be the same or different and are C 1-6 alkyl group, C 2-6 alkenyl group, C 2-6 alkynyl group, halogen atom, C 1-6 haloalkyl group , a C 3-6 cycloalkyl group, a C 1 -C 6 alkoxy group, a carboxyl group or an amino group,
The three dotted lines, which may be the same or different, are double bonds (=) or single bonds (-),
m, n, p and q may be the same or different and are 0, 1 or 2)
An agent for treating or preventing Chagas disease, characterized by containing
[2] The compound is selected from one or more selected from the group consisting of coptisine, dehydrocorridarin and palmatine or a salt thereof, or a compound represented by general formula (I) or a salt thereof, The agent according to [1].
[3] The compound is one or more selected from the group consisting of coptisine and dehydrocorridarin or a salt thereof, or a compound represented by general formula (I) or a salt thereof, [1] or [ 2].
[4] The agent according to any one of [1] to [3], wherein the compound is coptisine or a salt thereof.
[5] The agent according to any one of [1] to [4], which is further characterized by being effective for the treatment or prevention of acute Chagas disease.
[6] The agent according to any one of [1] to [5], which is further characterized by being effective for the treatment or prevention of acute and chronic Chagas disease.
[7] One or more compounds or salts thereof selected from the group consisting of coptisine, berberine, palmatine, epiberberine, berberrubine and ditetrahydrocoptisine, or a compound represented by the following general formula (I) or the salt
Figure JPOXMLDOC01-appb-C000004
 (wherein R 1 , R 2 , R 3 and R 4 may be the same or different, hydrogen atom, C 1-6 alkyl group, C 2-6 alkenyl group, C 2-6 alkynyl group, C 1 -6 haloalkyl group, C 3-6 cycloalkyl group,
R 1 may form a 5- or 6-membered ring together with R 2 and the adjacent oxygen atom;
R 3 may form a 5- or 6-membered ring together with R 4 and the adjacent oxygen atom;
R 5 , R 6 , R 7 and R 8 may be the same or different and are C 1-6 alkyl group, C 2-6 alkenyl group, C 2-6 alkynyl group, halogen atom, C 1-6 haloalkyl group , a C 3-6 cycloalkyl group, a carboxyl group or an amino group,
The three dotted lines, which may be the same or different, are double bonds (=) or single bonds (-),
m, n, p and q may be the same or different and are 0, 1 or 2)
A composition for treating or preventing Chagas disease, characterized by containing
　本発明によれば、シャーガス病の治療又は予防に有効な化合物及び該化合物を含むシャーガス病治療又は予防剤を提供することができる。 According to the present invention, a compound effective for the treatment or prevention of Chagas disease and an agent for the treatment or prevention of Chagas disease containing the compound can be provided.
一次スクリーニングである抗トリパノソーマ活性評価した結果（生薬エキス剤・生薬由来化合物）を示す図である。FIG. 2 is a diagram showing the results of evaluation of antitrypanosomal activity (crude drug extract/crude drug-derived compound), which is primary screening. 二次スクリーニングである細胞傷害性試験（生薬エキス剤・生薬由来化合物）の結果を示す図である。FIG. 2 is a diagram showing the results of a cytotoxicity test (herbal drug extract and herbal drug-derived compound), which is a secondary screening. 本発明の化合物について、抗トリパノソーマ活性を測定した結果を示す図である。FIG. 2 shows the results of measuring the antitrypanosomal activity of the compounds of the present invention. 本発明の化合物について、細胞内アマスチゴートにおけるIC50の測定結果を示す図である。FIG. 3 shows the results of IC 50 measurements on intracellular amastigotes for the compounds of the present invention. 本発明の化合物について、NBMH細胞に対する細胞傷害性の測定結果を示す図である。FIG. 2 shows the results of measurement of cytotoxicity against NBMH cells for compounds of the present invention. 本発明の化合物について、HuH28細胞に対する細胞傷害性の測定結果を示す図である。FIG. 2 shows the results of measuring the cytotoxicity of the compounds of the present invention against HuH28 cells. （Ａ）：マウスに対し、コプチシンを腹腔内投与したin vivo 発光強度の画像である。(A): Images of in vivo luminescence intensity after intraperitoneal administration of coptisine to mice. （Ｂ）：マウスに対し、コプチシンを腹腔内投与したin vivo 発光強度を定量した結果である。(B): Results of quantifying in vivo luminescence intensity after intraperitoneal administration of coptisine to mice.
　本発明において「シャーガス病」とは、クルーズトリパノソーマ原虫によって引き起こされるサシガメという昆虫に媒介される熱帯病（感染症）である。
　本発明において「急性期」のシャーガス病とは、通常、感染直後の数週間～数カ月の期間を指し、無症状又は、熱、疲労感、かゆみ、頭痛、下痢が見られる症状を指す。また、原虫が侵入した部分に赤い腫脹やしこりが出る皮膚病変（シャゴーマ）や、サシガメに咬まれたり、眼などにサシガメの糞が入ることにより、眼のまぶたが腫れる症状（ロマーニャ徴候）が見られる場合も含まれる。この症状は、通常、数週間程度で自然に消滅するが、原虫は体内に留まり、長い潜伏期に入ることがある。
　なお、本明細書においては、血中にトリポマスチゴート（trypomastigotes）、細胞内にアマスチゴート（amastigotes）が存在する状態を、「急性期」又は「急性期」を模した状態とすることがある。
　本発明において「慢性期」のシャーガス病とは、上記の急性期経過後、通常、数十年ほどの潜伏期を経て、心臓合併症（心臓肥大、心不全、心拍数の変化や不整脈、心尖部動脈瘤・血栓形成、心肺停止（突然死）など）、腸管合併症（食道又は結腸の肥大（巨大食道・巨大結腸）、これらを原因とした食事困難や排せつ困難など）の重篤な症状を呈する状態を指す。１～３割程度の感染者においてこれらの症状が見られることがある。
　なお、本明細書においては、細胞内でアマスチゴート（amastigotes）が増殖している状態を、「慢性期」又は「慢性期」を模した状態とすることがある。 In the present invention, “Chagas disease” is a tropical disease (infectious disease) caused by Trypanosoma cruzi protozoa and transmitted by an insect called assassin bug.
In the present invention, "acute phase" Chagas disease usually refers to a period of several weeks to several months immediately after infection, and refers to asymptomatic symptoms or symptoms such as fever, fatigue, itching, headache, and diarrhea. In addition, red swelling and lumps appearing on the skin where the protozoa have invaded (chagoma), and swelling of the eyelids (Romagna sign) due to bites by kissing bugs or contact with the feces of kissing bugs. It also includes cases where This symptom usually disappears spontaneously within a few weeks, but the protozoa may remain in the body and enter a long incubation period.
In the present specification, the state in which trypomastigotes are present in the blood and the state in which amastigotes are present in the cells may be referred to as the "acute phase" or a state simulating the "acute phase." .
In the present invention, the “chronic phase” of Chagas disease refers to cardiac complications (cardiac hypertrophy, heart failure, changes in heart rate, arrhythmia, apical artery aneurysm/thrombosis, cardiopulmonary arrest (sudden death), etc.), intestinal complications (enlargement of the esophagus or colon (megaesophagus/megacolon), difficulty eating or excretion due to these) refers to the state. These symptoms may be seen in about 10 to 30% of infected people.
In the present specification, the state in which amastigotes proliferate in cells may be referred to as a "chronic phase" or a state simulating a "chronic phase."
　以下、本発明のシャーガス病治療又は予防剤（組成物）について詳しく説明する。
　本発明のシャーガス病治療又は予防剤（組成物）は、コプチシン、ベルベリン、パルマチン、エピベルベリン、ベルベルルビン及びジテトラハイドロコプチシンからなる群から選択される化合物若しくはその塩、又は下記の一般式（Ｉ）で表される化合物（本明細書において、「類縁化合物」と呼ぶこともある）若しくはその塩
Figure JPOXMLDOC01-appb-C000005
 （式中、Ｒ１、Ｒ２、Ｒ３及びＲ４は、同一又は異なってもよく、水素原子、Ｃ１－６アルキル基、Ｃ２－６アルケニル基、Ｃ２－６アルキニル基、Ｃ１－６ハロアルキル基、Ｃ３－６シクロアルキル基であり、好ましくは、水素原子、Ｃ１－６アルキル基又はＣ３－６シクロアルキル基である。より好ましくは、水素原子か、Ｃ１－６アルキル基である。
　Ｒ１はＲ２及び隣接する酸素原子と一緒に５又は６員環を形成してもよく、
　Ｒ３はＲ４及び隣接する酸素原子と一緒に５又は６員環を形成してもよく、
　Ｒ５、Ｒ６、Ｒ７及びＲ８は、同一又は異なってもよく、Ｃ１－６アルキル基、Ｃ２－６アルケニル基、Ｃ２－６アルキニル基、ハロゲン原子、Ｃ１－６ハロアルキル基、Ｃ３－６シクロアルキル基、Ｃ１－Ｃ６アルコキシ基、カルボキシル基又はアミノ基であり、好ましくは、Ｃ１－６アルキル基、ハロゲン原子又はＣ３－６シクロアルキル基である。より好ましくは、Ｃ１－６アルキル基又はハロゲン原子である。
　３箇所の点線は、それぞれ、同一又は異なってもよく、二重結合（＝）又は単結合（－）であり（但し、窒素原子の下の結合が二重結合となる場合は、窒素原子はイオン（Ｎ＋）である）、
　ｍ、ｎ、ｐ及びｑは、同一又は異なってもよく、０、１又は２である）
　を含有することを特徴とする。 The therapeutic or preventive agent (composition) for Chagas disease of the present invention is described in detail below.
The therapeutic or preventive agent (composition) for Chagas disease of the present invention is a compound or a salt thereof selected from the group consisting of coptisine, berberine, palmatine, epiberberine, berberrubine and ditetrahydrocoptisine, or the following general formula: Compound represented by (I) (in this specification, may be referred to as "analogous compound") or a salt thereof
Figure JPOXMLDOC01-appb-C000005
 (wherein R 1 , R 2 , R 3 and R 4 may be the same or different, hydrogen atom, C 1-6 alkyl group, C 2-6 alkenyl group, C 2-6 alkynyl group, C 1 -6 haloalkyl group, C 3-6 cycloalkyl group, preferably hydrogen atom, C 1-6 alkyl group or C 3-6 cycloalkyl group, more preferably hydrogen atom or C 1-6 is an alkyl group.
R 1 may form a 5- or 6-membered ring together with R 2 and the adjacent oxygen atom;
R 3 may form a 5- or 6-membered ring together with R 4 and the adjacent oxygen atom;
R 5 , R 6 , R 7 and R 8 may be the same or different and are C 1-6 alkyl group, C 2-6 alkenyl group, C 2-6 alkynyl group, halogen atom, C 1-6 haloalkyl group , a C 3-6 cycloalkyl group, a C 1 -C 6 alkoxy group, a carboxyl group or an amino group, preferably a C 1-6 alkyl group, a halogen atom or a C 3-6 cycloalkyl group. More preferably, it is a C 1-6 alkyl group or a halogen atom.
The three dotted lines, which may be the same or different, are double bonds (=) or single bonds (-) (however, if the bond under the nitrogen atom is a double bond, the nitrogen atom is ion (N + )),
m, n, p and q may be the same or different and are 0, 1 or 2)
It is characterized by containing
　好ましくは、本発明のシャーガス病治療又は予防剤（組成物）は、コプチシン、ベルベリン、パルマチン、エピベルベリン、ベルベルルビン及びジテトラハイドロコプチシンからなる群から選択される化合物若しくはその塩、又は上記した一般式（Ｉ）で表される化合物若しくはその塩を、有効成分として含有する。 Preferably, the therapeutic or preventive agent (composition) for Chagas disease of the present invention is a compound or a salt thereof selected from the group consisting of coptisine, berberine, palmatine, epiberberine, berberrubine and ditetrahydrocoptisine, or the above The compound represented by general formula (I) or a salt thereof is contained as an active ingredient.
　本発明において、「Ｃ１－Ｃ６アルキル基」としては、例えば、メチル基、エチル基、プロピル基、イソプロピル基、ブチル基、イソブチル基、セカンダリーブチル基、ターシャリーブチル基、ペンチル基、イソペンチル基、ターシャリーペンチル基、ネオペンチル基、２，３－ジメチルプロピル基等の直鎖又は分岐鎖状のアルキル基が挙げられるが、これらに限定されない。 In the present invention, the "C 1 -C 6 alkyl group" includes, for example, methyl group, ethyl group, propyl group, isopropyl group, butyl group, isobutyl group, secondary butyl group, tertiary butyl group, pentyl group and isopentyl group. , a tertiary pentyl group, a neopentyl group, a 2,3-dimethylpropyl group, and other linear or branched alkyl groups, but are not limited thereto.
　本発明において、「Ｃ２－Ｃ６アルケニル基」としては、例えば、ビニル基、アリル基、イソプロペニル基、１－ブテニル基、２－ブテニル基、２－メチル－２－プロペニル基等の直鎖又は分鎖状のアルケニル基が挙げられるが、これらに限定されない。 In the present invention, the "C 2 -C 6 alkenyl group" includes, for example, linear groups such as vinyl group, allyl group, isopropenyl group, 1-butenyl group, 2-butenyl group, 2-methyl-2-propenyl group, and the like. or branched alkenyl groups, but are not limited thereto.
　本発明において、「Ｃ２－Ｃ６アルキニル基」としては、例えば、エチニル基、１－プロピニル基、２－プロピニル基、１－ブチニル基、２－ブチニル基、３－ブチニル基、３－メチル－１－プロピニル基、フェニルエチニル基等の直鎖又は分鎖状のアルキニル基が挙げられるが、これらに限定されない。 In the present invention, the "C 2 -C 6 alkynyl group" includes, for example, ethynyl group, 1-propynyl group, 2-propynyl group, 1-butynyl group, 2-butynyl group, 3-butynyl group, 3-methyl- Linear or branched alkynyl groups such as 1-propynyl group and phenylethynyl group may be mentioned, but not limited thereto.
　本発明において、「ハロ」とは「ハロゲン原子」を意味し、ハロゲン原子は、塩素原子、臭素原子、ヨウ素原子又はフッ素原子を示す。 In the present invention, "halo" means "halogen atom", and the halogen atom indicates a chlorine atom, a bromine atom, an iodine atom or a fluorine atom.
　本発明において、「Ｃ３－Ｃ６シクロアルキル基」としては、例えば、シクロプロピル基、シクロブチル基、シクロペンチル基、シクロヘキシル基等の環状のアルキル基が挙げられるが、これらに限定されない。 In the present invention, the “C 3 -C 6 cycloalkyl group” includes, but is not limited to, cyclic alkyl groups such as cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl groups.
　本発明において、「Ｃ１－Ｃ６アルコキシ基」としては、例えば、メトキシ基、エトキシ基、プロポキシ基、イソプロポキシ基、ブトキシ基、セカンダリーブトキシ基、ターシャリーブトキシ基、ペンチルオキシ基、イソペンチルオキシ基、ネオペンチルオキシ基等の直鎖又は分岐鎖状のアルコキシ基等が挙げられるが、これらに限定されない In the present invention, the "C 1 -C 6 alkoxy group" includes, for example, methoxy group, ethoxy group, propoxy group, isopropoxy group, butoxy group, secondary butoxy group, tertiary butoxy group, pentyloxy group, isopentyloxy linear or branched alkoxy groups such as neopentyloxy groups, etc., but are not limited thereto.
　本発明に用いられる好ましい塩としては、例えば、塩酸塩、硫酸塩、硝酸塩、燐酸塩等の無機酸塩、酢酸塩、フマル酸塩、マレイン酸塩、シュウ酸塩、メタンスルホン酸塩、ベンゼンスルホン酸塩、パラトルエンスルホン酸塩等の有機酸塩、ナトリウムイオン、カリウムイオン、カルシウムイオン、トリメチルアンモニウム等の無機又は有機の塩基との塩を例示することができるが、これらに限定されない。 Preferred salts for use in the present invention include, for example, inorganic acid salts such as hydrochlorides, sulfates, nitrates and phosphates, acetates, fumarates, maleates, oxalates, methanesulfonates, and benzenesulfones. Salts, organic acid salts such as p-toluenesulfonate, and salts with inorganic or organic bases such as sodium ion, potassium ion, calcium ion, and trimethylammonium can be exemplified, but are not limited to these.
　好ましくは、本発明のシャーガス病治療又は予防剤は、コプチシン、ベルベリン及びパルマチンからなる群から選択される化合物若しくはその塩、又は上記一般式（Ｉ）で表される化合物若しくはその塩を含む。より好ましくは、本発明のシャーガス病治療又は予防剤は、コプチシン又はその塩を含み、さらに好ましくは、コプチシン塩酸塩を含むが、これらに限定されない。また、別の本発明の好ましい例としては、本発明のシャーガス病治療又は予防剤は、デヒドロコリダリン硝酸塩又はパルマチン塩酸塩を含むが、これらに限定されない。 Preferably, the agent for treating or preventing Chagas disease of the present invention contains a compound or a salt thereof selected from the group consisting of coptisine, berberine and palmatine, or a compound represented by the general formula (I) or a salt thereof. More preferably, the agent for treating or preventing Chagas disease of the present invention contains coptisine or a salt thereof, more preferably coptisine hydrochloride, but is not limited thereto. As another preferred example of the present invention, the therapeutic or prophylactic agent for Chagas disease of the present invention includes, but is not limited to, dehydrocorridarin nitrate or palmatine hydrochloride.
　さらに、本発明の別の好ましいシャーガス病治療又は予防剤の態様は、下記の６つの化合物又はその塩から選択される１種以上を含むが、これらに限定されない。
Figure JPOXMLDOC01-appb-C000006
 Furthermore, another preferred embodiment of the agent for treating or preventing Chagas disease of the present invention includes, but is not limited to, one or more selected from the following six compounds or salts thereof.
Figure JPOXMLDOC01-appb-C000006
　本発明のシャーガス病治療又は予防剤においては、コプチシン、ベルベリン、パルマチン、エピベルベリン、ベルベルルビン及びジテトラハイドロコプチシンからなる群から選択される化合物若しくはその塩、又は上記一般式（Ｉ）で表される化合物若しくはその塩が複数（例えば、１種以上、２種以上、３種以上、１～５種、又は２～４種など）含まれていてもよい。 In the agent for treating or preventing Chagas disease of the present invention, a compound or a salt thereof selected from the group consisting of coptisine, berberine, palmatine, epiberberine, berberrubine and ditetrahydrocoptisine, or the general formula (I) A plurality of represented compounds or salts thereof (eg, 1 or more, 2 or more, 3 or more, 1 to 5, or 2 to 4, etc.) may be included.
　本発明のシャーガス病治療又は予防剤は、急性期のみならず、慢性期のシャーガス病治療又は予防にも有効であるため、有用である。好ましくは、本発明の剤は、シャーガス病の急性期症状の治療又は予防に有効であり、より好ましくは、本発明の剤は、急性期症状及び慢性期症状の治療又は予防に有効である。 The agent for treating or preventing Chagas disease of the present invention is useful not only for the treatment or prevention of acute phase Chagas disease, but also for the treatment or prevention of chronic phase Chagas disease. Preferably, the agent of the present invention is effective in treating or preventing acute phase symptoms of Chagas disease, and more preferably, the agent of the present invention is effective in treating or preventing acute phase symptoms and chronic phase symptoms.
　本発明のシャーガス病治療又は予防剤又は組成物中の、上記化合物の含有量は、特に限定されないが、剤又は組成物１００質量％中、例えば、約５０質量％～約１００質量％の範囲であってもよく、好ましくは約７５質量％～約１００質量％、より好ましくは、約９０質量％～約１００質量％の範囲であってよい。 The content of the above compound in the agent or composition for treating or preventing Chagas disease of the present invention is not particularly limited, but in 100% by mass of the agent or composition, for example, in the range of about 50% to about 100% by mass. It may be present, preferably in the range of about 75% to about 100% by weight, more preferably about 90% to about 100% by weight.
　本発明の剤又は組成物は、上記した本発明の化合物又はその塩と、所望により、薬学的に許容される担体、添加剤等とを適宜配合して製造することにより、製剤化できる。そのための製剤化方法や技術は従来十分に確立されているので、それに従ってよい。
　例えば、医薬品用の場合、具体的には、錠剤、被覆錠剤、丸剤、散剤、顆粒剤、カプセル剤、液剤、懸濁剤、乳剤等の経口剤、注射剤、輸液、坐剤、軟膏、パッチ剤等の非経口剤とすることができるが、これらに限定されない。担体又は添加剤の配合割合については、医薬品等の分野において通常採用されている範囲に基づいて適宜設定すればよい。 The agent or composition of the present invention can be formulated by appropriately blending the above-described compound of the present invention or a salt thereof and, if desired, pharmaceutically acceptable carriers, additives and the like. Formulation methods and techniques for this purpose have been well established in the past, and may be followed.
For example, in the case of pharmaceuticals, specifically oral agents such as tablets, coated tablets, pills, powders, granules, capsules, liquids, suspensions, emulsions, injections, infusions, suppositories, ointments, Parenteral agents such as patches can be used, but are not limited to these. The mixing ratio of the carrier or additive may be set as appropriate based on the range normally employed in the field of pharmaceuticals and the like.
　薬学的に許容される担体又は添加剤は特に制限されないが、担体の例としては、水性または油性基剤等の各種担体が挙げられ、水性の担体としては、例えば、水、生理食塩水、エタノール、グリセリン、ポリエチレングリコール、プロピレングリコール、メチルセルロース、ヒドロキシプロピルメチルセルロース、ヒドロキシプロピルセルロース、ポリビニルピロリドン、ポリアクリル酸、多糖ガム系天然高分子類等が挙げられ、油性の担体としては、例えば、ワセリン、スクワラン、パラフィン等の油類やワックス類等が挙げられるが、これらに限定されない。
　添加剤の例としては、酵素、ｐＨ調整剤、保存剤、殺菌剤、賦形剤、結合剤、崩壊剤、滑沢剤、酸化防止剤、光沢剤、香料、甘味料、酸味料、調味料、苦味料、乳化剤、増粘剤、安定剤、ゲル化剤、着色剤、矯味剤、香料等が挙げられるが、これらに限定されない。これら添加剤に関する技術は、従来十分に確立されているので、本発明において、それらに従ってよい。
　さらに、本発明の組成物には、本発明の効果を失しない限り、例えば、医学、薬学、獣医学又は食品等の分野で知られる任意の成分が含有されていてもよい。 Pharmaceutically acceptable carriers or additives are not particularly limited, but examples of carriers include various carriers such as aqueous or oily bases, and examples of aqueous carriers include water, physiological saline, and ethanol. , glycerin, polyethylene glycol, propylene glycol, methyl cellulose, hydroxypropylmethyl cellulose, hydroxypropyl cellulose, polyvinylpyrrolidone, polyacrylic acid, polysaccharide gum-based natural polymers, etc. Examples of oily carriers include vaseline, squalane, Examples include, but are not limited to, oils such as paraffin and waxes.
Examples of additives include enzymes, pH adjusters, preservatives, bactericides, excipients, binders, disintegrants, lubricants, antioxidants, brighteners, flavors, sweeteners, acidulants, seasonings. , bittering agents, emulsifiers, thickeners, stabilizers, gelling agents, coloring agents, flavoring agents, flavoring agents, and the like. Techniques for these additives are well established in the prior art, and may be followed in the present invention.
Furthermore, the composition of the present invention may contain any ingredient known in the fields of medicine, pharmacy, veterinary medicine, food, etc., as long as the effects of the present invention are not lost.
　なお、本発明の化合物は経口でも、非経口でも投与することができる。投与形態としては、経口投与、眼局所投与、静脈内投与、経皮投与等が挙げられ、必要に応じて医薬として許容される添加剤を適宜選択して使用し、投与形態に適した剤型に製剤化することができる。
　例えば、本発明の化合物又はその塩が、経口又は注射投与される場合は、摂取者の年齢及び体重、症状、投与時間、剤形、投与方法、薬剤の組み合わせ等に依存して決定できる。本発明の化合物が、好ましくは、後述するように包摂等される場合で、例えば、液体で、経口投与される場合、成人１人（約６０ｋｇ）１日当たり、好ましくは約６０～６００ｍｇ／ｍＬ摂取されるように設定するのが好ましく、注射により投与される場合は、最大約６０ｍｇ／ｍＬが注射されるように設定するのが好ましい。 The compound of the present invention can be administered orally or parenterally. Examples of the administration form include oral administration, topical administration to the eye, intravenous administration, transdermal administration, etc., and if necessary, pharmaceutically acceptable additives are appropriately selected and used, and a dosage form suitable for the administration form is used. can be formulated into
For example, when the compound of the present invention or a salt thereof is administered orally or by injection, it can be determined depending on the age and body weight of the ingestor, symptoms, administration time, dosage form, administration method, drug combination, and the like. When the compound of the present invention is preferably subsumed as described below, for example, when administered orally in liquid form, the intake is preferably about 60 to 600 mg/mL per adult (about 60 kg) per day. If administered by injection, it is preferably set to inject up to about 60 mg/mL.
　外用塗布の場合は、適用する皮膚面積に応じて、化合物又はその塩の塗布量を適宜選択することができるが、通常、当該塗布量は、適用部位の面積約１０ｃｍ２に対して、１日につき、好ましくは約０．０１～５ｍｇ、より好ましくは約０．０５～１ｍｇである。前記の投与用量を、１日あたり、１回又は数回に分けて投与ないし適用するとよい。 In the case of external application, the amount of the compound or its salt to be applied can be appropriately selected according to the skin area to be applied. is preferably about 0.01-5 mg, more preferably about 0.05-1 mg. It is preferable to administer or apply the above administration dose once or in several divided doses per day.
化合物の包摂
　本発明の好ましい態様としては、本発明の化合物（例えば、コプチシン、ベルベリン、パルマチン、エピベルベリン、ベルベルルビン及びジテトラハイドロコプチシンからなる群から選択される化合物若しくはその塩、あるいは上記一般式（Ｉ）で表される化合物など）が、水溶性の向上水溶液中での安定性、生体への吸収性等の観点から、包摂されている場合が挙げられる。化合物を包摂する方法については、従来十分に確立されているので、そのような方法に従ってよい。 Inclusion of compounds In a preferred embodiment of the present invention, the compound of the present invention (e.g., a compound or a salt thereof selected from the group consisting of coptisine, berberine, palmatine, epiberberine, berberrubine and ditetrahydrocoptisine, or the above Compounds represented by general formula (I), etc.) are included from the viewpoint of improving water solubility, stability in an aqueous solution, bioabsorbability, and the like. Methods for encapsulating compounds are well established in the art and may be followed.
　例えば、このような、本発明の化合物を好ましく包摂する化合物の具体例としては、特に限定されないが、キュカービット［ｎ］ウリルが挙げられる。キュカービット［ｎ］ウリル（通常、ｎは５～１０）は、構成単位であるグライコウリルがｎ個環状に連なり、内部に疎水性を有する空孔を備えた樽状の化合物（下記参照）である。
Figure JPOXMLDOC01-appb-C000007
 For example, specific examples of such compounds that preferably include compounds of the present invention include, but are not limited to, cukabit[n]uril. Cukabit[n]uril (usually n is 5 to 10) is a barrel-shaped compound (see below) with n glycourils, which are structural units, arranged in a ring and having a hydrophobic vacancy inside. be.
Figure JPOXMLDOC01-appb-C000007
　キュカービット［ｎ］ウリルは、ナフタレン構造やコレステロール構造等を有する化合物と、選択的に非常に高い相互作用を示すことが知られており、本発明の化合物（例えば、コプチシン、ベルベリン、パルマチン、エピベルベリン、ベルベルルビン及びジテトラハイドロコプチシンからなる群から選択される化合物若しくはその塩など）と相性がよい。なお、このようなキュカービット［ｎ］ウリルの製造方法は、例えば、特開2012-246239号に記載されているので、当該方法に従ってよい。あるいは市販品（メルク社）を使用してもよい。なお、本発明において、好ましくは、キュカービット［ｎ］ウリルは、ｎが６～９であり、より好ましくは、ｎが７～８である。 Cukabit [n] uril is known to selectively show a very high interaction with compounds having a naphthalene structure, a cholesterol structure, etc. compounds selected from the group consisting of epiberberine, berberrubine and ditetrahydrocoptisine or salts thereof). A method for producing such a cukabit[n]uril is described, for example, in JP-A-2012-246239, and the method may be followed. Alternatively, a commercially available product (Merck) may be used. In the present invention, in the cukabit[n]uril, n is preferably 6-9, more preferably 7-8.
　キュカービット［ｎ］ウリルは、特開2012-246239号に記載の方法により、合成する場合、キュカービット［ｎ］ウリル（例えば、ｎ：７～９）と、本発明の化合物とを水中又は包摂に適した溶媒（例えば、有機溶媒等）中で、例えば、２：１～１：２等で混合することにより、本発明の化合物をキュカービット［ｎ］ウリルに包摂させることが出来る。本発明の化合物が包摂されることにより、水溶性が向上する。なお、キュカービットは、ククルビットと表記されることもあるが同じものである。
　別の本発明の化合物を好ましく包摂する化合物の態様としては、例えば、α‐シクロデキストリン、β‐シクロデキストリン、γ‐シクロデキストリン、クラスターデキストリンなどのデキストリン類の他、クラウンエーテル（例えば、12－クラウン－4、15－クラウン－5、18－クラウン－6など）、フラーレン（例えば、Ｃ６０フラーレンや、Ｃ７０フラーレンなど）、カリックス［ｍ］アレーン（ｍ：３～２０、好ましくは、ｎ：４～８）、ピラーアレーン（例えば、1,4-ジメトキシピラー[5]アレーン）等を挙げることができるが、これらに限定されない。
　なお、本発明の化合物が上記の包摂のための化合物により包摂され、水溶性が向上している場合には、より一層、本発明の効果（急性期及び／又は慢性期のシャーガス病の治療又は予防効果など）が好ましく得られる。 Cucurbit[n]uril is synthesized by the method described in JP-A-2012-246239. Alternatively, the compounds of the present invention can be incorporated into cukabit[n]urils by mixing, for example, 2:1 to 1:2 in a solvent suitable for inclusion (such as an organic solvent). . Water solubility is improved by inclusion of the compound of the present invention. Cukabit is sometimes written as cucurbit, but it is the same thing.
Another embodiment of the compound preferably encompassing the compound of the present invention includes dextrins such as α-cyclodextrin, β-cyclodextrin, γ-cyclodextrin, and cluster dextrin, as well as crown ethers (e.g., 12-crown -4, 15-crown-5, 18-crown-6, etc.), fullerene (e.g., C60 fullerene, C70 fullerene, etc.), calix[m]arene (m: 3 to 20, preferably n: 4 to 8 ), pillar arenes (eg, 1,4-dimethoxy pillar[5]arene), and the like, but are not limited thereto.
In addition, when the compound of the present invention is subsumed by the above compound for inclusion and has improved water solubility, the effect of the present invention (treatment of acute and / or chronic Chagas disease or preventive effect, etc.) can be preferably obtained.
　次に、実験例、実施例を挙げて本発明をさらに具体的に説明するが、本発明はこれらの実施例により何ら限定されるものではなく、多くの変形が本発明の技術的思想内で当分野において通常の知識を有する者により可能である。
　なお、本発明に使用する実施例、比較例及び参考例の化合物は市販されたものを容易に入手することができ、それを使用することができる。 Next, the present invention will be described in more detail with reference to experimental examples and examples, but the present invention is not limited by these examples, and many modifications can be made within the technical concept of the present invention. It can be done by a person having ordinary knowledge in the field.
The compounds of Examples, Comparative Examples, and Reference Examples used in the present invention are commercially available and can be used.
＜化合物、試薬＞
　生薬由来化合物および生薬エキス：富山大学和漢医薬学総合研究所が和漢薬ライブラリーとして提供されたものを用いた。
　ベンズニダゾール (419656)：SIGMA-ALDRICH、
　ピッカジーン (309-04321)：東洋ビーネット(株)、
　NP40 (145-09701)：和光純薬、
　MEMメディウム (135-15175)：和光純薬、
　New Born Calf Serum (16010159)：Thermo Fisher Scientific、
　アラマーブルー試薬 (OOSA11225)：フナコシ(株)、
　ルシフェラーゼ(Luc2)を発現する組み換えTrypanosoma cruzi (Tulahuen株)：長崎大学熱研バイオリソースセンターから提供されたものを用いた。
　New Born Mouse Heart (NBMH) 細胞：長崎大学熱研バイオリソースセンターから提供されたものを用いた。
　HuH28細胞：タマサート大学チュラポーン国際医学部から提供されたものを用いた。
　D-luciferin(MB000102-R70170-2)：Syd Labs、
　Coptisine chloride (C685500)：Toronto Research Chemicals、
　イソフルラン(095-06573)：和光純薬、
　シクロフォスファミド一水和物(C0768）:SIGMA-ALDRICH。 <Compound, Reagent>
Crude drug-derived compounds and crude drug extracts: those provided by the Institute of Japanese and Chinese Medicine, University of Toyama as a Japanese and Chinese drug library were used.
Benznidazole (419656): SIGMA-ALDRICH,
Picka Gene (309-04321): Toyo Benet Co., Ltd.,
NP40 (145-09701): Wako Pure Chemical,
MEM Medium (135-15175): Wako Pure Chemical,
New Born Calf Serum (16010159): Thermo Fisher Scientific,
Alamar Blue reagent (OOSA11225): Funakoshi Co., Ltd.,
Recombinant Trypanosoma cruzi (Tulahuen strain) expressing luciferase (Luc2): The one provided by Nekken Bioresource Center, Nagasaki University was used.
New Born Mouse Heart (NBMH) cells: those provided by Nekken Bioresource Center, Nagasaki University were used.
HuH28 cells: those provided by Chulabhorn International School of Medicine, Thammasat University were used.
D-luciferin (MB000102-R70170-2): Syd Labs,
Coptisine chloride (C685500): Toronto Research Chemicals,
Isoflurane (095-06573): Wako Pure Chemical,
Cyclophosphamide monohydrate (C0768): SIGMA-ALDRICH.
表１：使用生薬由来化合物一覧
Figure JPOXMLDOC01-appb-T000008
 Table 1: List of crude drug-derived compounds used
Figure JPOXMLDOC01-appb-T000008
表２：使用生薬エキス一覧
Figure JPOXMLDOC01-appb-T000009
 Table 2: List of crude drug extracts used
Figure JPOXMLDOC01-appb-T000009
Figure JPOXMLDOC01-appb-T000010
Figure JPOXMLDOC01-appb-T000010
実施例１：抗トリパノソーマ活性を指標にした一次スクリーニング
１－１　実験方法
　96 well 白プレートにルシフェラーゼ遺伝子を組み込んだトリポマスチゴート(3×104cell/well)とマウスの心臓から単離した線維芽細胞(NBMH細胞)(5×104cell/well)を播種した。そこに上記した表１及び表２に記載の生薬由来化合物(20 μM)又は生薬エキス(20 μg/mL)のいずれかを1 well当たり1種を加え、７２時間培養した。ポジティブコントロールとしてNP40、ネガティブコントロールには薬剤を溶かした溶媒のみを加えた。また、既存薬であるベンズニダゾールを加えるwellを設けた。1 well当たりの容量は100 μlとした。７２時間培養後に100μlの0.6% NP40を加え、細胞を破砕した。破砕した細胞及びメディウムを1.5 mLチューブに移し、遠心分離(12,000 rpm、15分間)を行った。遠心分離操作後、上清50μlに、ルシフェリン(ピッカジーン)を100μl加え、発光強度(544 nm/590 nm)をプレートリーダーにより測定した(測定時間：1秒)。ネガティブコントロールの発光抑制率を0％、ポジティブコントロールの発光抑制率を100％とし、各化合物またはエキスの発光抑制率を算出した。
　なお、実施例１の実験方法は、Bettiolら（PLoS Neglected Tropical Diseases, 2009, 3(2): e384に記載の方法を参考にした。この実験系は、培地中のトリポマスチゴートおよび細胞内のアマスチゴートに対する効果を確認することができる。 Example 1: Primary screening using antitrypanosomal activity as an index 1-1 Experimental method Trypomastigotes ( 3 x 104 cells/well) in which luciferase gene was incorporated in a 96-well white plate and fibers isolated from mouse heart Blast cells (NBMH cells) (5×10 4 cells/well) were seeded. Either the herbal drug-derived compound (20 μM) or herbal drug extract (20 μg/mL) described in Tables 1 and 2 above was added to each well and cultured for 72 hours. NP40 was added as a positive control, and only the solvent in which the drug was dissolved was added to the negative control. In addition, wells were provided to add benznidazole, an existing drug. The volume per well was 100 µl. After culturing for 72 hours, 100 µl of 0.6% NP40 was added to disrupt the cells. The disrupted cells and medium were transferred to a 1.5 mL tube and centrifuged (12,000 rpm, 15 minutes). After centrifugation, 100 μl of luciferin (Picagene) was added to 50 μl of the supernatant, and the luminescence intensity (544 nm/590 nm) was measured with a plate reader (measurement time: 1 second). Taking the luminescence suppression rate of the negative control as 0% and the luminescence suppression rate of the positive control as 100%, the luminescence suppression rate of each compound or extract was calculated.
The experimental method of Example 1 was based on the method described in Bettiol et al. (PLoS Neglected Tropical Diseases, 2009, 3(2): e384.) effect on amastigotes can be confirmed.
１－２　結果
　結果を図１に示した。（A）が生薬エキス剤の結果、（B）が生薬由来化合物の結果である。上記表１の９６種の生薬由来化合物のうち８種（4 (Alisol B), 5 (Alkannin), 17 (Berberine Chloride), 29 (Coptisine Chloride), 33 (Dehydrocorydaline Nitrate), 78 (Palmatine Chloride), 91 (Shikonin), 95 (Timosaponin A-III)）及び表２の１２０種の生薬エキスのうち２種（8: Phellodendron Bark、黄柏（オウバク）, 10: Coptis Rhizome、黄連（オウレン））において、８０％以上の発光強度の低下がみられた。 1-2 Results The results are shown in FIG. (A) is the result of the crude drug extract, and (B) is the result of the crude drug-derived compound. Of the 96 crude drug-derived compounds in Table 1 above, 8 (4 (Alisol B), 5 (Alkannin), 17 (Berberine Chloride), 29 (Coptisine Chloride), 33 (Dehydrocorydaline Nitrate), 78 (Palmatine Chloride), 91 (Shikonin), 95 (Timosaponin A-III)) and two of the 120 crude drug extracts in Table 2 (8: Phellodendron Bark, Phellodendron bark, 10: Coptis Rhizome, Huanglian), A decrease in emission intensity of 80% or more was observed.
実施例２：細胞傷害性を指標にした二次スクリーニング
２－１　実験方法
　96 well 黒プレートにNBMH細胞（1×104/well）およびHuH28細胞（1×104/well）を播種した。そこに生薬由来化合物(20 μM)又は生薬エキス(20 μg/mL)のいずれかを、1 well当たり1種を加え、72時間培養した。ポジティブコントロールとしてNP40、ネガティブコントロールとして薬剤を溶かした溶媒のみを加えた。また、既存薬であるベンズニダゾールを加えるwellを設けた。1 well当たりの容量は100 μlとした。72時間培養後にアラマーブルー試薬を10 μl加え、さらに4時間インキュベーションした。その後蛍光強度をプレートリーダーで測定した（測定時間0.1秒）。 Example 2: Secondary Screening Using Cytotoxicity as Index 2-1 Experimental Method NBMH cells (1×10 4 /well) and HuH28 cells (1×10 4 /well) were seeded in a 96-well black plate. Either a crude drug-derived compound (20 μM) or a crude drug extract (20 μg/mL) was added to each well and cultured for 72 hours. As a positive control, NP40 was added, and as a negative control, only solvent in which the drug was dissolved was added. In addition, wells were provided to add benznidazole, an existing drug. The volume per well was 100 μl. After culturing for 72 hours, 10 μl of Alamar Blue reagent was added, and the cells were further incubated for 4 hours. After that, the fluorescence intensity was measured with a plate reader (measurement time 0.1 sec).
２－２　結果
　結果を図２に示した。（A）が生薬エキス剤の結果、（B）が生薬由来化合物の結果である。一次スクリーニングにおいて80%以上の発光強度の低下がみられた化合物又はエキスにおいて、コプチシン塩酸塩（Coptisine Chloride）、デヒドロコリダリン硝酸塩（Dehydrocorydaline Nitrate）、パルマチン塩酸塩（Palmatine Chloride）の3種は、蛍光強度の低下が確認されず、細胞傷害性がないことが示された。
　これらの3種の化合物には共通の構造がみられたため、さらに3種の類縁体（ベルベルルビン（berberrubine）、エピベルベリン（epiberberine）、ジテトラハイドロコプチシン（di-tetrahydrocoptisine））を加えた６種の化合物について、トリパノソーマに対する活性を検証した。 2-2 Results The results are shown in FIG. (A) is the result of the crude drug extract, and (B) is the result of the crude drug-derived compound. Among the compounds or extracts that showed a decrease in luminescence intensity of 80% or more in the primary screening, three of them, Coptisine Chloride, Dehydrocorydaline Nitrate, and Palmatine Chloride, showed fluorescence. No decrease in strength was observed, indicating no cytotoxicity.
Since these three compounds had a common structure, three more analogues (berberrubine, epiberberine, and di-tetrahydrocoptisine) were added. Six compounds were tested for activity against trypanosomes.
実施例３：抗トリパノソーマ活性のIC50の測定
３－１　実験方法
　上記６種の化合物について、実施例１と同じ実験方法で抗トリパノソーマ活性を測定した。コプチシン、デヒドロコリダリンおよびパルマチンの濃度は、40μM、33μM、20μM、16.5μM、10μM、8.25μM、5.0μM、4.13μM、2.5μM、2.06μMおよび1.25μMとした。ベルベルルビン、エピベルベリンおよびジテトラハイドロコプチシンの濃度は、50μM、25μM、12.5μM、6.25μM、3.125μMおよび1.6μMとした。 Example 3 Measurement of IC50 of Antitrypanosoma Activity 3-1 Experimental Method The antitrypanosoma activity of the above six compounds was measured by the same experimental method as in Example 1. The concentrations of coptisine, dehydrocorridarin and palmatine were 40 μM, 33 μM, 20 μM, 16.5 μM, 10 μM, 8.25 μM, 5.0 μM, 4.13 μM, 2.5 μM, 2.06 μM and 1.25 μM. Berberrubine, epiberberine and ditetrahydrocoptisine concentrations were 50 μM, 25 μM, 12.5 μM, 6.25 μM, 3.125 μM and 1.6 μM.
３－２　結果
　結果を図３に示した。（A）がコプチシン塩酸塩（Coptisine Chloride）、（B）がデヒドロコリダリン硝酸塩（Dehydrocorydaline Nitrate）、（C）がパルマチン塩酸塩（Palmatine Chloride）、（D）がエピベルベリン（epiberberine）、（E）がベルベルルビン（berberrubine）、（F）がジテトラハイドロコプチシン（di-tetrahydrocoptisine）の結果である。コプチシン塩酸塩、デヒドロコリダリン硝酸塩、パルマチン塩酸塩において、ベンズニダゾールと同等の発光強度の減弱がみられた。ベルベルルビンの発光強度の減弱はベンズニダゾールより弱かった。エピベルベリン、ジテトラハイドロコプチシンは発光強度の減弱がみられなかった。
　この実験系は、血中にトリポマスチゴート、細胞内にアマスチゴートが存在する、いわゆる急性期の状態を模倣しているため、発光強度が減弱した化合物は、トリパノソーマ感染の急性期に効果があると考えられる。結果の詳細は表３を参照のこと。 3-2 Results The results are shown in FIG. (A) Coptisine Chloride, (B) Dehydrocorydaline Nitrate, (C) Palmatine Chloride, (D) epiberberine, (E) is for berberrubine and (F) is for di-tetrahydrocoptisine. In coptisine hydrochloride, dehydrocorridarin nitrate, and palmatine hydrochloride, attenuation of luminescence intensity equivalent to that of benznidazole was observed. Attenuation of luminescence intensity of berberrubine was weaker than that of benznidazole. Epiberberine and ditetrahydrocoptisine showed no decrease in luminescence intensity.
Since this experimental system mimics the so-called acute phase, in which trypomastigotes are present in blood and amastigotes are present in cells, compounds with reduced luminescence intensity are effective in the acute phase of trypanosoma infection. it is conceivable that. See Table 3 for details of the results.
実施例４：細胞内アマスチゴートに対する活性のIC50の測定
４－１　実験方法
　ルシフェラーゼ遺伝子を発現するトリポマスチゴートおよびNBMH細胞を2:1の割合で25cm2フラスコに播種した。細胞内への感染を強めるために培地をMEM+1%NBCSとした。24時間後PBSで2回洗浄した後MEM+10％NBCSを加え、さらに24時間インキュベーター内で培養した。その後、トリプシン処理を行いTrypanosoma cruzi（T. cruzi）を含むNBMH細胞を剥がした。96 well 白プレートにT. cruziを含むNBMH細胞（5×104/well）を播種した。そこに上記6種の化合物を1 well当たり1種を加え、72時間培養した。コプチシン、デヒドロコリダリンおよびパルマチンの濃度は、40μM、33μM、20μM、16.5μM、10μM、8.25μM、5.0μM、4.13μM、2.5μM、2.06μMおよび1.25μMとした。ベルベルルビン、エピベルベリンおよびジテトラハイドロコプチシンの濃度は、50μM、25μM、12.5μM、6.25μM、3.125μMおよび1.6μMとした。
　ポジティブコントロールとしてNP40、ネガティブコントロールとして薬剤を溶かした溶媒のみを加えた。また、既存薬であるベンズニダゾールを加えるwellを設けた。1 well当たりの容量は100 μlとした。コプチシン塩酸塩、デヒドロコリダリン硝酸塩およびパルマチン塩酸塩の濃度は、実施例３と同じである。エピベルベリン、ベルベルルビンおよびジテトラハイドロコプチシンの濃度は、100μM、50μM、25μM、12.5μM、6.25μMおよび3.125μMとした。72時間培養後に100 μlの0.6%NP40を加え、細胞を破砕した。破砕した細胞及びメディウムを1.5mlチューブに移し、遠心分離（12,000rpm、15分間）を行った。上清50 μlにルシフェリン（ピッカジーン）を100 μl加え発光強度（544 nm/590 nm）をプレートリーダーにより測定した（測定時間1秒）。ネガティブコントロールの発光抑制率を0％、ポジティブコントロールの発光抑制率を100％とし、各化合物またはエキスの発光抑制率を算出した。
　なお、実施例４の実験方法は、Ryckerら（PLoS Negl Trop Dis. 2016 Apr 15;10(4):e0004584）およびAlonso-Padillaら（PLoS Negl Trop Dis. 2015 Jan 23;9(1):e0003493）に記載の方法を参考にした。この実験系は、細胞に感染したトリパノソーマの増殖型アマスチゴートに対する効果を確認することができる。 Example 4 Measurement of IC50 of Activity Against Intracellular Amastigote 4-1 Experimental Method Trypomastigotes expressing luciferase gene and NBMH cells were seeded in a 25 cm 2 flask at a ratio of 2:1. The medium was MEM+1% NBCS to enhance intracellular infection. After 24 hours, the cells were washed twice with PBS, added with MEM+10% NBCS, and further cultured in an incubator for 24 hours. After that, trypsin treatment was performed to detach NBMH cells containing Trypanosoma cruzi (T. cruzi). NBMH cells (5×10 4 /well) containing T. cruzi were seeded in a 96-well white plate. One of the above six compounds was added to each well and cultured for 72 hours. The concentrations of coptisine, dehydrocorridarin and palmatine were 40 μM, 33 μM, 20 μM, 16.5 μM, 10 μM, 8.25 μM, 5.0 μM, 4.13 μM, 2.5 μM, 2.06 μM and 1.25 μM. Berberrubine, epiberberine and ditetrahydrocoptisine concentrations were 50 μM, 25 μM, 12.5 μM, 6.25 μM, 3.125 μM and 1.6 μM.
As a positive control, NP40 was added, and as a negative control, only solvent in which the drug was dissolved was added. In addition, wells were provided to add benznidazole, an existing drug. The volume per well was 100 μl. The concentrations of coptisine hydrochloride, dehydrocorridarin nitrate and palmatine hydrochloride are the same as in Example 3. The concentrations of epiberberine, berberrubine and ditetrahydrocoptisine were 100 μM, 50 μM, 25 μM, 12.5 μM, 6.25 μM and 3.125 μM. After culturing for 72 hours, 100 µl of 0.6% NP40 was added to disrupt the cells. The disrupted cells and medium were transferred to a 1.5 ml tube and centrifuged (12,000 rpm, 15 minutes). 100 µl of luciferin (Picagene) was added to 50 µl of the supernatant, and the luminescence intensity (544 nm/590 nm) was measured with a plate reader (measurement time: 1 second). Taking the luminescence suppression rate of the negative control as 0% and the luminescence suppression rate of the positive control as 100%, the luminescence suppression rate of each compound or extract was calculated.
In addition, the experimental method of Example 4 was described by Rycker et al. (PLoS Negl Trop Dis. 2016 Apr 15;10(4):e0004584) and Alonso-Padilla et al. ) was used as a reference. This experimental system can confirm the effect of trypanosome-infected cells on proliferating amastigotes.
４－２　結果
　結果を図４に示した。（A）がコプチシン塩酸塩（Coptisine Chloride）、（B）がデヒドロコリダリン硝酸塩（Dehydrocorydaline Nitrate）、（C）がパルマチン塩酸塩（Palmatine Chloride）、（D）がエピベルベリン（epiberberine）、（E）がベルベルルビン（berberrubine）、（F）がジテトラハイドロコプチシン（di-tetrahydrocoptisine）の結果である。コプチシンは細胞内アマスチゴートに対してベンズニダゾールと同等の発光強度の減弱がみられた。デヒドロコリダリン硝酸塩、パルマチン塩酸塩およびベルベルルビンの発光強度の減弱はベンズニダゾールより弱かった。エピベルベリンおよびジテトラハイドロコプチシンは発光強度の減弱がみられなかった。
　この実験系は、細胞内でアマスチゴートが増殖している、いわゆる慢性期の状態を模倣しているため、発光強度が減弱した化合物は、トリパノソーマ感染の慢性期に効果があると考えられる。結果の詳細は表３を参照のこと。 4-2 Results The results are shown in FIG. (A) Coptisine Chloride, (B) Dehydrocorydaline Nitrate, (C) Palmatine Chloride, (D) epiberberine, (E) is for berberrubine and (F) is for di-tetrahydrocoptisine. Coptisine reduced the luminescence intensity of intracellular amastigotes to the same extent as benznidazole. Dehydrocorridarin nitrate, palmatine hydrochloride and berberrubine attenuated the luminescence intensity weaker than that of benznidazole. Epiberberine and ditetrahydrocoptisine did not decrease the luminescence intensity.
Since this experimental system mimics the so-called chronic phase state in which amastigotes proliferate in cells, compounds with reduced luminescence intensity are considered to be effective in the chronic phase of trypanosome infection. See Table 3 for details of the results.
実施例５：NBMH細胞に対する細胞傷害性の評価
５－１　実験方法
　上記６種の化合物について、実施例２の実験方法において使用細胞をNBMH細胞1種類に変更した以外は実施例２と同じ実験方法でNBMH細胞に対する細胞傷害性を評価した。各化合物の濃度は実施例４と同じである。 Example 5: Evaluation of cytotoxicity against NBMH cells 5-1 Experimental method For the above six compounds, the same experimental method as in Example 2 except that the cells used in the experimental method of Example 2 were changed to one type of NBMH cells. was evaluated for cytotoxicity against NBMH cells. The concentration of each compound is the same as in Example 4.
５－２　結果
　結果を図５に示した、（A）がコプチシン塩酸塩（Coptisine Chloride）、（B）がデヒドロコリダリン硝酸塩（Dehydrocorydaline Nitrate）、（C）がパルマチン塩酸塩（Palmatine Chloride）、（D）がエピベルベリン（epiberberine）、（E）がベルベルルビン（berberrubine）、（F）がジテトラハイドロコプチシン（di-tetrahydrocoptisine）の結果である。コプチシン塩酸塩、デヒドロコリダリン硝酸塩、パルマチン塩酸塩、エピベルベリン、ジテトラハイドロコプチシンは、蛍光強度の減弱がみられず、NBMH細胞に対して細胞傷害性がないことが示された。ベルベルルビンにおいては蛍光強度の減弱が若干みられ、NBMH細胞に対して弱い細胞傷害性が示された。結果の詳細は表３を参照のこと。 5-2 Results The results are shown in Figure 5, (A) for Coptisine Chloride, (B) for Dehydrocorydaline Nitrate, (C) for Palmatine Chloride, ( D) is for epiberberine, (E) is for berberrubine, and (F) is for di-tetrahydrocoptisine. Coptisine hydrochloride, dehydrocorridarin nitrate, palmatine hydrochloride, epiberberine, and ditetrahydrocoptisine did not show any decrease in fluorescence intensity, indicating that they had no cytotoxicity to NBMH cells. Berberrubine slightly decreased fluorescence intensity, indicating weak cytotoxicity to NBMH cells. See Table 3 for details of the results.
実施例６：HuH28細胞に対する細胞傷害性の評価
６－１　実験方法
　上記６種の化合物について、実施例２の実験方法において使用細胞をHuH28細胞１種類に変更した以外は実施例２と同じ実験方法でHuH28細胞に対する細胞傷害性を評価した。各化合物の濃度は実施例４と同じである。 Example 6: Evaluation of cytotoxicity against HuH28 cells 6-1 Experimental method For the above six compounds, the same experimental method as in Example 2 except that the cells used in the experimental method of Example 2 were changed to one type of HuH28 cells. was evaluated for cytotoxicity against HuH28 cells. The concentration of each compound is the same as in Example 4.
６－２　結果
　結果を図６に示した、（A）がコプチシン塩酸塩（Coptisine Chloride）、（B）がデヒドロコリダリン硝酸塩（Dehydrocorydaline Nitrate）、（C）がパルマチン塩酸塩（Palmatine Chloride）、（D）がエピベルベリン（epiberberine）、（E）がベルベルルビン（berberrubine）、（F）がジテトラハイドロコプチシン（di-tetrahydrocoptisine）の結果である。コプチシン塩酸塩、デヒドロコリダリン硝酸塩、パルマチン塩酸塩、エピベルベリン、ベルベルルビン、ジテトラハイドロコプチシンは、いずれも蛍光強度の減弱がみられず、HuH28細胞に対して細胞傷害性がないことが示された。結果の詳細は表３を参照のこと。 6-2 Results The results are shown in Figure 6, (A) for Coptisine Chloride, (B) for Dehydrocorydaline Nitrate, (C) for Palmatine Chloride, ( D) is for epiberberine, (E) is for berberrubine, and (F) is for di-tetrahydrocoptisine. None of coptisine hydrochloride, dehydrocorridarine nitrate, palmatine hydrochloride, epiberberine, berberrubine, and ditetrahydrocoptisine showed a decrease in fluorescence intensity, indicating that they had no cytotoxicity to HuH28 cells. shown. See Table 3 for details of the results.
　実施例３～６の実験結果の詳細を以下の表３に示す。
Figure JPOXMLDOC01-appb-T000011
 Details of the experimental results for Examples 3-6 are provided in Table 3 below.
Figure JPOXMLDOC01-appb-T000011
実施例７：マウスに対するコプチシンの効果
７－１　実験方法
　7-8週齢の雌のBALB/cマウスに、T.cruzi（trypomastigotes型、Tulahuen 株）1×104cellを腹腔内投与した。感染4日目から8日目まで5日間連続で薬剤を投与した。コプチシンは1日2回、30 mg/kgの用量で腹腔内投与した。その後、感染31日目、感染34日目、感染37日目にシクロフォスファミド一水和物を200 mg/kgの用量で腹腔内投与することで免疫抑制を行った。ポジティブコントロールとしてベンズニダゾール（100 mg/kg）を1日1回経口投与した。ネガティブコントロールとして1%PBSを1日1回経口投与した。発光強度の測定は薬剤投与開始前日（感染3日目）と薬剤投与終了の翌日（感染9日目）を含め、感染14日目、感染28日目、更に免疫抑制後の感染40日目の計5回行った。発光強度はD-luciferinを150 mg/kgで腹腔内投与し、10分間イソフルラン吸入後、発光 in vivoイメージングシステムIVIS Lumina IIを使用して測定した。 Example 7: Effect of coptisine on mice 7-1 Experimental method 1×10 4 cells of T. cruzi (trypomastigotes type, Tulahuen strain) were intraperitoneally administered to 7- to 8-week-old female BALB/c mice. The drug was administered for 5 consecutive days from day 4 to day 8 of infection. Coptisine was administered intraperitoneally twice daily at a dose of 30 mg/kg. Thereafter, immunosuppression was performed by intraperitoneal administration of 200 mg/kg of cyclophosphamide monohydrate on days 31, 34, and 37 of infection. Benznidazole (100 mg/kg) was orally administered once a day as a positive control. As a negative control, 1% PBS was orally administered once a day. Luminescence intensity was measured on day 14 of infection, day 28 of infection, and day 40 of infection after immunosuppression, including the day before the start of drug administration (day 3 of infection) and the day after the end of drug administration (day 9 of infection). I went there a total of 5 times. Luminescence intensity was measured by intraperitoneally administering D-luciferin at 150 mg/kg, inhaling isoflurane for 10 minutes, and using the luminescence in vivo imaging system IVIS Lumina II.
７－２　結果
　結果を図７に示した。図７－１（A）が各群のマウスの in vivo 発光強度の画像であり、図７－２（B）が発光強度を定量した結果である。トリパノソーマに感染したマウスに対してコプチシンを腹腔内投与すると発光強度の減弱がみられた。特に慢性期モデルである免疫抑制後の感染40日目では、コプチシンはベンズニダゾールと同等以上に蛍光強度を減弱させた。この結果は、コプチシンがトリパノソーマ感染急性期および慢性期において有効であることを示すものである。 7-2 Results The results are shown in FIG. Figure 7-1 (A) is an image of the in vivo luminescence intensity of each group of mice, and Figure 7-2 (B) is the result of quantification of the luminescence intensity. Intraperitoneal administration of coptisine to trypanosome-infected mice decreased the luminescence intensity. In particular, coptisine attenuated the fluorescence intensity to the same level as or more than that of benznidazole on day 40 of infection after immunosuppression, which is a chronic phase model. This result indicates that coptisine is effective in the acute and chronic stages of trypanosomal infection.
実施例８（化合物の包摂による水溶性向上）
　メルク社から購入したキュカービット［7］ウリルとコプチシン塩酸塩又はベルベリン塩酸塩を水中で、１：１：で混合することにより、コプチシン塩酸塩又はベルベリン塩酸塩をキュカービット［7］ウリルに包摂させた。包摂することにより、コプチシン塩酸塩又はベルベリン塩酸塩の水溶性が向上した。このように水溶性が向上したコプチシン塩酸塩又はベルベリン塩酸塩は、例えば、vivo投与（経口投与、注射投与）等に適するため、有用である。 Example 8 (improvement of water solubility by inclusion of compound)
Copticine hydrochloride or berberine hydrochloride was added to cukabit[7]uril by mixing cukabit[7]uril and coptisine hydrochloride or berberine hydrochloride purchased from Merck & Co. 1:1 in water. subsumed. Inclusion improved the water solubility of coptisine hydrochloride or berberine hydrochloride. Coptisine hydrochloride or berberine hydrochloride with such improved water solubility is suitable for, for example, in vivo administration (oral administration, injection administration), and thus is useful.

Claims (7)

  1. 　コプチシン、ベルベリン、パルマチン、エピベルベリン、ベルベルルビン及びジテトラハイドロコプチシンからなる群から選択される１種以上の化合物若しくはその塩、又は
    　下記の一般式（Ｉ）で表される化合物若しくはその塩
    Figure JPOXMLDOC01-appb-C000001
     （式中、Ｒ１、Ｒ２、Ｒ３及びＲ４は、同一又は異なってもよく、水素原子、Ｃ１－６アルキル基、Ｃ２－６アルケニル基、Ｃ２－６アルキニル基、Ｃ１－６ハロアルキル基、Ｃ３－６シクロアルキル基であり、
    　Ｒ１はＲ２及び隣接する酸素原子と一緒に５又は６員環を形成してもよく、
    　Ｒ３はＲ４及び隣接する酸素原子と一緒に５又は６員環を形成してもよく、
    　Ｒ５、Ｒ６、Ｒ７及びＲ８は、同一又は異なってもよく、Ｃ１－６アルキル基、Ｃ２－６アルケニル基、Ｃ２－６アルキニル基、ハロゲン原子、Ｃ１－６ハロアルキル基、Ｃ３－６シクロアルキル基、Ｃ１－Ｃ６アルコキシ基、カルボキシル基又はアミノ基であり、
    　３箇所の点線は、それぞれ、同一又は異なってもよく、二重結合（＝）又は単結合（－）であり、
    　ｍ、ｎ、ｐ及びｑは、同一又は異なってもよく、０、１又は２である）
    　を含有することを特徴とする、シャーガス病治療又は予防剤。     One or more compounds or salts thereof selected from the group consisting of coptisine, berberine, palmatine, epiberberine, berberrubine and ditetrahydrocoptisine, or compounds represented by the following general formula (I) or salts thereof
    Figure JPOXMLDOC01-appb-C000001
     (wherein R 1 , R 2 , R 3 and R 4 may be the same or different, hydrogen atom, C 1-6 alkyl group, C 2-6 alkenyl group, C 2-6 alkynyl group, C 1 -6 haloalkyl group, C 3-6 cycloalkyl group,
    R 1 may form a 5- or 6-membered ring together with R 2 and the adjacent oxygen atom;
    R 3 may form a 5- or 6-membered ring together with R 4 and the adjacent oxygen atom;
    R 5 , R 6 , R 7 and R 8 may be the same or different and are C 1-6 alkyl group, C 2-6 alkenyl group, C 2-6 alkynyl group, halogen atom, C 1-6 haloalkyl group , a C 3-6 cycloalkyl group, a C 1 -C 6 alkoxy group, a carboxyl group or an amino group,
    The three dotted lines, which may be the same or different, are double bonds (=) or single bonds (-),
    m, n, p and q may be the same or different and are 0, 1 or 2)
    An agent for treating or preventing Chagas disease, characterized by containing
  2. 　化合物が、コプチシン、デヒドロコリダリン及びパルマチンからなる群から選択される１種以上若しくはその塩又は一般式（Ｉ）で表される化合物若しくはその塩から選択されることを特徴とする、請求項１に記載の剤。 (1) wherein the compound is selected from one or more selected from the group consisting of coptisine, dehydrocorridarin and palmatine, salts thereof, or compounds represented by general formula (I) or salts thereof; The agent described in .
  3. 　化合物がコプチシン及びデヒドロコリダリンからなる群から選択される１種以上若しくはその塩又は一般式（Ｉ）で表される化合物若しくはその塩であることを特徴とする、請求項１又は２に記載の剤。 3. The compound according to claim 1 or 2, wherein the compound is one or more selected from the group consisting of coptisine and dehydrocorridarin or a salt thereof, or a compound represented by general formula (I) or a salt thereof. agent.
  4. 　化合物がコプチシン又はその塩であることを特徴とする、請求項１～３のいずれか１項に記載の剤。 The agent according to any one of claims 1 to 3, wherein the compound is coptisine or a salt thereof.
  5. 　急性期のシャーガス病の治療又は予防に有効であることをさらに特徴とする、請求項１～４のいずれか１項に記載の剤。 The agent according to any one of claims 1 to 4, which is further characterized by being effective for the treatment or prevention of acute phase Chagas disease.
  6. 　急性期及び慢性期のシャーガス病の治療又は予防に有効であることをさらに特徴とする、請求項１～５のいずれか１項に記載の剤。 The agent according to any one of claims 1 to 5, further characterized by being effective for the treatment or prevention of acute and chronic Chagas disease.
  7. 　コプチシン、ベルベリン、パルマチン、エピベルベリン、ベルベルルビン及びジテトラハイドロコプチシンからなる群から選択される１種以上の化合物若しくはその塩又は
    　下記の一般式（Ｉ）で表される化合物若しくはその塩
    Figure JPOXMLDOC01-appb-C000002
     （式中、Ｒ１、Ｒ２、Ｒ３及びＲ４は、同一又は異なってもよく、水素原子、Ｃ１－６アルキル基、Ｃ２－６アルケニル基、Ｃ２－６アルキニル基、Ｃ１－６ハロアルキル基、Ｃ３－６シクロアルキル基であり、
    　Ｒ１はＲ２及び隣接する酸素原子と一緒に５又は６員環を形成してもよく、
    　Ｒ３はＲ４及び隣接する酸素原子と一緒に５又は６員環を形成してもよく、
    　Ｒ５、Ｒ６、Ｒ７及びＲ８は、同一又は異なってもよく、Ｃ１－６アルキル基、Ｃ２－６アルケニル基、Ｃ２－６アルキニル基、ハロゲン原子、Ｃ１－６ハロアルキル基、Ｃ３－６シクロアルキル基、カルボキシル基又はアミノ基であり、
    　３箇所の点線は、それぞれ、同一又は異なってもよく、二重結合（＝）又は単結合（－）であり、
    　ｍ、ｎ、ｐ及びｑは、同一又は異なってもよく、０、１又は２である）
    　を含有することを特徴とする、シャーガス病治療又は予防用組成物。     One or more compounds selected from the group consisting of coptisine, berberine, palmatine, epiberberine, berberrubine and ditetrahydrocoptisine or a salt thereof, or a compound represented by the following general formula (I) or a salt thereof
    Figure JPOXMLDOC01-appb-C000002
     (wherein R 1 , R 2 , R 3 and R 4 may be the same or different, hydrogen atom, C 1-6 alkyl group, C 2-6 alkenyl group, C 2-6 alkynyl group, C 1 -6 haloalkyl group, C 3-6 cycloalkyl group,
    R 1 may form a 5- or 6-membered ring together with R 2 and the adjacent oxygen atom;
    R 3 may form a 5- or 6-membered ring together with R 4 and the adjacent oxygen atom;
    R 5 , R 6 , R 7 and R 8 may be the same or different and are C 1-6 alkyl group, C 2-6 alkenyl group, C 2-6 alkynyl group, halogen atom, C 1-6 haloalkyl group , a C 3-6 cycloalkyl group, a carboxyl group or an amino group,
    The three dotted lines, which may be the same or different, are double bonds (=) or single bonds (-),
    m, n, p and q may be the same or different and are 0, 1 or 2)
    A composition for treating or preventing Chagas disease, characterized by containing
PCT/JP2022/011217 2021-03-17 2022-03-14 Agent for treating or preventing chagas disease WO2022196614A1 (en)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
JP2021-044164 2021-03-17
JP2021044164 2021-03-17

Publications (1)

Publication Number Publication Date
WO2022196614A1 true WO2022196614A1 (en) 2022-09-22

Family

ID=83320380

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/JP2022/011217 WO2022196614A1 (en) 2021-03-17 2022-03-14 Agent for treating or preventing chagas disease

Country Status (1)

Country Link
WO (1) WO2022196614A1 (en)

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPH07330623A (en) * 1994-06-10 1995-12-19 Tsumura & Co Suppressant for neuron death
JP2005281284A (en) * 2004-03-26 2005-10-13 Ivy Cosmetics Corp Skin care preparation for external use for promoting collagen synthesis
JP2018177733A (en) * 2017-04-19 2018-11-15 国立大学法人帯広畜産大学 Herbal medicine-derived compound having antiprotozoal action and herbal medicine extract

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPH07330623A (en) * 1994-06-10 1995-12-19 Tsumura & Co Suppressant for neuron death
JP2005281284A (en) * 2004-03-26 2005-10-13 Ivy Cosmetics Corp Skin care preparation for external use for promoting collagen synthesis
JP2018177733A (en) * 2017-04-19 2018-11-15 国立大学法人帯広畜産大学 Herbal medicine-derived compound having antiprotozoal action and herbal medicine extract

Non-Patent Citations (3)

* Cited by examiner, † Cited by third party
Title
EKINS SEAN, LAGE DE SIQUEIRA-NETO JAIR, MCCALL LAURA-ISOBEL, SARKER MALABIKA, YADAV MANEESH, PONDER ELIZABETH L., KALLEL E. ADAM, : "Machine Learning Models and Pathway Genome Data Base for Trypanosoma cruzi Drug Discovery", PLOS NEGLECTED TROPICAL DISEASES, vol. 9, no. 6, 26 June 2015 (2015-06-26), pages e0003878, XP055966948, DOI: 10.1371/journal.pntd.0003878 *
NKWENGOUA ERNESTINE, NGANTCHOU IGOR, NYASSE BARTHéLEMY, DENIER COLETTE, BLONSKI CASIMIR, SCHNEIDER BERND: "In vitro inhibitory effects of palmatine from Enantia chlorantha on Trypanosoma cruzi and Leishmania infantum", NATURAL PRODUCT RESEARCH, TAYLOR AND FRANCIS HEALTH SCIENCES, ABINGDON, GB, vol. 23, no. 12, 1 January 2009 (2009-01-01), GB , pages 1144 - 1150, XP009539704, ISSN: 1478-6419, DOI: 10.1080/14786410902726241 *
SCHINELLA, G.R.; TOURNIER, H.A.; PRIETO, J.M.; RIOS, J.L.; BUSCHIAZZO, H.; ZAIDENBERG, A.: "Inhibition of Trypanosoma cruzi growth by medical plant extracts", MEDICINAL & AROMATIC PLANTS ABSTRACTS, SCIENTIFIC PUBLISHERS, SCIENTIFIC PUBLISHERS, NEW DELHI - INDIA, vol. 25, no. 3, 1 June 2003 (2003-06-01), Scientific Publishers, New Delhi - India , XP018014552, ISSN: 0250-4367 *

Similar Documents

Publication Publication Date Title
US5153178A (en) Compositions and method of treatment for improving circulatory performance
US4761417A (en) Compounds, compositions and method of treatments for improving circulatory performance
US9895323B2 (en) Astaxanthin anti-inflammatory synergistic combinations
WO2021147235A1 (en) Application of favipiravir in treating coronavirus infection
JPH09505809A (en) Inhibition of smooth muscle migration and proliferation by hydroxycarbazole compounds
KR20210151816A (en) Cannabinoid acid ester compositions and uses thereof
JPH08253416A (en) Composition of phonofibrate and vitamin e and its application on medical treatment
JPH04504726A (en) Use of lipoxin A↓4 and its derivatives as antagonists against slow-reacting substances in anaphylaxis
JPS58134031A (en) Dihydrocyclosporin d for treating multiple screlosis
WO2021068802A1 (en) Application of nadh and salt and/or nmn thereof in preparation of drug or health care product for enhancing muscle or inhibiting muscle volume reduction
Hall et al. Human plasma and urine quinine levels following tablets, capsules, and intravenous infusion
US4152448A (en) Method and composition for the treatment of a condition of the gastro intestinal tract
EP0108147B1 (en) Use of a protoberberine alkaloid and composition containing same
AU2007341218A1 (en) Isosorbide mononitrate derivatives for the treatment of intestinal disorders
WO2022196614A1 (en) Agent for treating or preventing chagas disease
CN1477957A (en) Tanshinone compound containing dihydrofuran ring structure for curing hyperammonemia and hepatic encephalopathy
US10953004B2 (en) Combination therapy for proliferative diseases
JPS625913A (en) Dihydropyridine-containing medicine
JPH04295428A (en) Antiallergic agent
JPS6330416A (en) Agent for improving cerebral circulation and metabolism
WO2021242758A1 (en) Improved nitro-fatty acid oral dose regimens
BRPI0804764A2 (en) alpha-glycosity inhibitors, pharmaceutical compositions comprising them and process for their preparation
US3737552A (en) Method of treating gonorrhea
JP2019507116A (en) Novel compounds that activate the estrogen receptor and compositions and methods of use thereof
JP2016514142A (en) Methods and products for enhancing the bioavailability of drugs and nutritional supplements

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 22771360

Country of ref document: EP

Kind code of ref document: A1

NENP Non-entry into the national phase

Ref country code: DE

122 Ep: pct application non-entry in european phase

Ref document number: 22771360

Country of ref document: EP

Kind code of ref document: A1