CN107789342A - The medical usage of sugared spore ketone compounds - Google Patents

Abstract

The invention belongs to field of pharmaceutical chemistry technology, the medical usage of sugared spore ketone compounds is related in particular to.Such compound is used for the disease for preventing or treating the tryptophan metabolism associated pathology feature as caused by IDO and/or TDO, including but not limited to tumor immune escape, neurological conditions, mental illness and angiocardiopathy etc..

Description

The medical usage of sugared spore ketone compounds

Technical field

The invention belongs to field of pharmaceutical chemistry technology, the medical usage of sugared spore ketone compounds is related in particular to.

Background technology

Heme-containing protein has different functions, and the transport, storage and single electron transfer including oxygen activate Deng.Indoleamine 2,3 dioxygenases（IDO）With tryptophan 2,3- dioxygenases（TDO）Belong to heme-containing protein family （Sedlmayr. et al, Frontiers in Immunology 2014;5:230）.TDO has found at first, in 1930 Year or so, TDO is typically considered to be only limited to express in the liver of mammal, but research confirms that it is present in eucaryote In exist in prokaryotes.What IDO was found in 1967, separated from the intestines of rabbit, and research confirms IDO Exist in many other eucaryotes（Ball HJ. et al, Front Immunol 2014;5:485）.

TDO and IDO active function is catalysis tryptophan oxidation generation formylkynurenine.TDO and IDO can pass through activation Oxygen molecule, it is catalyzed and inserts it into tryptophan, ultimately generate formylkynurenine.This reaction is Kynurenine metabolism way The first step and the rate-limiting reaction step in footpath, therefore, TDO and IDO can be crossed and consume tryptophan by catalytic reaction, and generate dog urinary ammonia The metabolite of acid metabolic approach.Tryptophan, be important amino acid for mammal, it be by diet regimen, Then it is metabolized by kynurenine metabolism pathway.Because tryptophan can be used for synthetic protein and neurochemical 5- hydroxyls Tryptamines, therefore TDO and IDO can regulate and control tryptophan levels by regulating and controlling kynurenine metabolism pathway, so as to suppress cell propagation With the synthesis of regulation and control serotonin.In addition, the metabolite of kynurenine metabolism pathway can also be NAD⁺Source, TDO are provided It can also be influenceed with IDO by regulating and controlling kynurenine metabolism pathway and regulate and control NAD⁺It is horizontal.Therefore, TDO and IDO is considered as swollen All there are important bioactive functions in a variety of diseases such as knurl, immune and nervous system.It is to be particularly noted that although IDO It is same reaction with TDO catalysis, but the sequence similarity of two enzymes is very low, and structure is also not quite similar（Badawy Abdulla A-B. et al, Bioscience Reports 2015;35:e00261）.

In tumor disease, TDO and IDO can be by controlling tryptophan consumption to adjust tryptophan levels, so as to press down The propagation and reduction T cell of T cell processed are to tumour cell immune response.Research shows, in the tumour cell of people, IDO can be through , also can be via GCN2 kinase pathways by the consumption of mTOR and GCN2 kinase pathways control tryptophan so as to suppress the propagation of T cell Treg cells are generated so as to reduce the immune response of tumour cell with kynurenin activation AhR approach（Munn DH. Et al, Trends in immunology 2013;34:137-43）.TDO also has expression in the tumour cell of many people, for example, TDO The immune response of tumour can be reduced in glioma by activating AhR（Opitz CA. et al, Nature 2011;478:197-203）.Therefore, TDO and IDO is considered as the important function target spot that tumour immunity suppresses.

In mental illness and in the nervous system disease, because tryptophan serotonin is to the pathology molecule machine of mental illness Adjustment effect is formed with, therefore IDO and psychotic disorder are closely related, such as insane, anxiety, depression, mental handicape etc..Separately Outside, caused other metabolites in kynurenine metabolism pathway, such as kynurenic acid, hydroxykynurenine and quinolinic acid, and The acting factor of nerve is stimulated, and it is relevant with the Pathological Physiology of many inflammation related neurological diseases（Kanai M. et al, Molecular Brain 2009;2:8）.Stages alzheimer's disease（AD）, Huntington disease（HD）And Parkinson's（PD）All with TDO is closely related with IDO, and wherein IDO is studied most, it is considered to be the therapy target of alzheimer and Huntington disease, TDO The potential research target spot of Huntington disease is also considered as later.Research discovery in the zoopery of stages alzheimer's disease, In APP/PS1 mouse models, intravenous injection IDO inhibitor can protect the loss of neuron, prevent the shape of amyloid patch Into raising cognitive ability.

In angiocardiopathy, because IDO is relevant with many fields of Vascular Biology, therefore studies and find, by endothelium Kynurenin caused by upper IDO effects can play vasorelaxation action as vascular relaxing factors, and cause septic shock mould The low blood pressure of type mouse, and IDO inhibitor improves Septic Shock Model mouse and deposited by improving the blood pressure of inflammation mouse Motility rate（Wang Y. et al, Nature medicine 2010;16:279-85）.Mouse experiment confirms, by artery congee IDO is induced in the focus of sample hardening, therapeutic action of the omega-3 aliphatic acid to rat aorta atherosis focus can be destroyed （Nakajima K, et al, Arteriosclerosis, thrombosis and vascular biology 2011; 31:1963-1972）.

At present, the highly efficient depressor using IDO and/or TDO as target spot has become the study hotspot of drug development in recent years. Wherein IDO inhibitor makes much progress in preclinical and clinical research, IDO inhibitor 1- methyl-D-tryptophan, Be referred to as indoximod, after testing its on tumor disease the effect of and security.In addition, what is carried out simultaneously also has second For IDO inhibitor oral medicine, INCB024360 and NLG919.Other IDO inhibitors also include 1-methyl-L- Tryptophan, Methylthiohydantoin tryptophan, Brassinin, Annulin B and its derivative and Exiguamine A and its derivative etc., these IDO inhibitors also have certain progress in clinical studies（Vacchelli E. et al, Oncoimmunology 2014;3:e957994）.

Thailand scientific research personnel in 2013 saves soil actinomycete Saccharopolyspora BCC 21906 from Thailand Jian Zhuwen In once isolated sugared spore ketone A and B, its activity research is also seldom, only report its to plasmodium falciparum K1 have suppress live Property, IC50 is respectively 4.1 and 3.9 μM；Have to tumor cell line KB, MCF-7 and NCI-H187 and benign Vero cells Inhibitory activity, IC50 [Boonlarppradab C, et al., The Journal of between 3.4 ~ 12.3 μM Antibiotics, 2013, 66(6): 305-309].Have no sugared spore ketone compounds in tumor immune escape, nervous system The report of activity in terms of illness, mental illness and angiocardiopathy.

The content of the invention

Present invention finds a kind of Novel IDO and TDO inhibitor with formula, the present inventor it has been investigated that, Formula compound suppresses IDO and TDO activity well in molecule and cellular water average energy, and completes its preparation and medicine Research in terms of purposes, therefore complete the present invention.

The present invention provides following formula: compound

（ I ）

R is H or OH,

（ II ）,

The purposes in medicine in the disease for preparing treatment, preventing or slowing down the related pathological characteristicses of tryptophan metabolism.This Invention provides compound, its isomeric forms, its analogue, its pharmaceutically acceptable salt, is preparing prevention, treatment Or slow down purposes in medicine in the disease related to IDO/TDO activity inhibitors.

Structural formula of the present invention（I）The compound of representative, when R is H, referred to as compound A；When R is OH, referred to as change Compound B.

Structural formula of the present invention（II）The compound of representative, when R is H, referred to as compound C.

The present invention also provides pharmaceutical composition, wherein comprising pharmaceutical carrier and one or more the compounds of this invention, its is same Divide isomeric form, its analogue, its pharmaceutically acceptable salt.

The present invention provides compound its isomeric forms, its analogue, its pharmaceutically acceptable salt and can be used for controlling Treat and/or prevent a variety of diseases or illness related to IDO/TDO activities inhibitor.

The present invention, which provides compound, its isomeric forms, its analogue, its pharmaceutically acceptable salt, individually to be made Used with or with one or more other pharmaceutical agent combinations.

Purposes of the present invention, wherein the disease is tumor immune escape, the nervous system disease, mental illness or painstaking effort Pipe disease, including but not limited to above-mentioned disease.

Purposes of the present invention, wherein tumor immune escape are the cancer of the brain, cutaneum carcinoma, carcinoma of urinary bladder, oophoroma, breast cancer, stomach It is cancer, cancer of pancreas, prostate cancer, colon cancer, leukemia, lung cancer, kidney, liver cancer, osteocarcinoma, incidence cancer, lymthoma, leukaemia, black Melanoma, including but not limited to above-mentioned tumor disease.

Purposes of the present invention, wherein the nervous system disease are stages alzheimer's disease, Huntington disease or Parkinson's, including but It is not limited to above-mentioned the nervous system disease.

Purposes of the present invention, wherein mental illness be anxiety, depression, mental handicape, insomnia, manic disorder, obsession, Neurosis, schizophrenia, including but not limited to above-mentioned mental illness.

Purposes of the present invention, its cardiovascular disease are atherosclerosis, angina pectoris, miocardial infarction, rhythm of the heart mistake Often, low blood pressure, cerebral apoplexy, ischemic cardiomyopathy caused by infecting, including but not limited to above-mentioned angiocardiopathy.

One or more other drugs or treatment method, such as other anticancers, immunopotentiator, immunodepressant, put Penetrate therapy, antitumor immunotherapy, cytokine therapy, optionally with the compounds of this invention, its isomeric forms, its knot Structure analog, its pharmaceutically acceptable salt, which combine, to be used to treat IDO and/or TDO relevant diseases or illness, or can be used as independent agent Type is simultaneously or sequentially administered.

Also include with other anticancers that compound of the present invention is applied in combination:For example, Alkylators, including it is but unlimited In nitrogen Jie, ethylenimine derivatives, alkylsulfonate, nitroso ureas and triazine, such as uracil mustard, nitrogen Jie, endoxan, Ifosfamide, American and French human relations, benzenebutanoic acid nitrogen Jie, the life of arc blood, triethylene-melamine, triethylene D2EHDTPA amine, disappear in vain Peace, BCNU, lomustine, streptozotocin, Dacarbazine and Temozolomide；For example, antimetabolite, includes but is not limited to Antifol, pyrimidine analogue, such as methopterin, 5-fluor-uracil, floxuridine, cytarabine, 6- thioguanines, phosphoric acid Fluorine reaches his shore, Pentostatin and gemcitabine；For example, some natural products and its derivative, including but not limited to Changchun peanut Alkaloids, anti-tumour antibody, enzyme, lymphokine, such as vinblastine, vincristine, eldisine, bleomycin, actinomyces D, Daunomycin, adriamycin, epirubicin, idarubicin, cytarabine, taxol, docetaxel, mithramycin, mitomycin- C, L- asparagus ferns amine acyl, interferon, Etoposide and Teniposide；Other anticancers also include, and adjust antibody, such as CTLA- altogether 4th, 4-1BB, PD-L1 and PD-1；Cell factor, such as IL-10, TGF-β etc.；Prevent immune cell migration, chemokine receptors Antagonist, such as CCR2, CCR4 and CCR6；Anti-cancer vaccine, including BMDC, synthetic peptide, DNA vaccination and recombinant virus.

The present invention provides compound, its isomeric forms, its analogue, its pharmaceutically acceptable salt and medicine group Closing can utilize various methods of administration or mode to discharge to patient.Suitable method of administration includes but is not limited to suction, transdermal, mouth Clothes, rectum, transmucosal, enteral and parenteral, parenteral include intramuscular, subcutaneous and intravenous injection.

Term " administration " used herein can be covered all directly with discharging compound indirectly to its predictive role position Means.

Reactive compound of the present invention can form administration itself, or be administered with pharmaceutical compositions, its Middle reactive compound is mixed with one or more pharmaceutically acceptable carriers, excipient or diluent.According to the present invention What the pharmaceutical composition used was typically prepared in the usual way, use one or more physiologically acceptable carriers, bag Containing excipient and auxiliary agent, they are advantageous to for reactive compound to be processed into the preparation that can pharmaceutically use.Appropriate preparation Depending on selected method of administration.

On being administered orally, compound is combined with pharmaceutically acceptable carrier well known in the art.This kind of carrier makes this Invention compound can prepare piece agent, pill, lozenge, capsule, liquid, gel, syrup, slurries, suspension etc., for quilt The patient to be treated is oral.Mouth can be obtained so with pharmaceutical preparation, be mixed with solid excipient, and alternatively grinding gained is mixed Compound, processes the mixture of particle, adds suitable auxiliary agent if desired, obtains tablet or lozenge core.Suitable figuration Agent is precisely that filler, such as sugar, including lactose, sucrose or mannitol；Cellulose preparations, for example, it is cornstarch, small Wheat starch, rice starch, farina, gelatin, bassora gum, methylcellulose, hydroxypropyl methyl cellulose, carboxymethyl cellulose Plain sodium and/or polyvinylpyrrolidone（PVP）.If desired, disintegrant, such as crosslinked polyethylene pyrrolidines can be added Ketone, agar or alginic acid or its salt, such as mosanom.

Pharmaceutical preparation that can be oral includes the capsule of sucking fit, is made up of gelatin, and soft fluid sealant wafer, It is made up of gelatin and a kind of plasticizer, such as glycerine.The capsule of sucking fit can contain active component and following ingredients Mixture：Filler, such as lactose；Adhesive, such as starch；And/or lubricant, such as talcum powder or magnesium stearate or micro mist Silica gel；With optional stabilizer.In soft capsule, reactive compound can be dissolved or suspended in suitable liquid, Such as fat oil, atoleine or liquid macrogol.Furthermore it is possible to add stabilizer.All oral Preparations all should It is suitable for the dosage of this kind of administration.

On by inhalation, used according to the present invention compound it is suitable in aerosol from pressurized package or Discharged in atomizer, wherein utilizing suitable propellant, such as dicholorodifluoromethane, trichlorofluoromethane, dichloro-tetrafluoro Ethane, carbon dioxide or other suitable gases.In the case of a pressurized aerosol, the valve for release being measured by providing can be with Determine dosage unit.Gelatine capsule and cartridge case in inhalator or insufflator can be configured to containing compound be adapted to The mixture of powders of powdered substrate, such as lactose or starch.

Compound can be formulated for parenteral, pass through injection, such as hitch injection or continuous infusion.Injection system Agent can be unit dosage forms, such as in ampoule or multi-dose container, wherein adding preservative.Composition can be taken in oiliness Or the form of suspension in aqueous carrier, solution or emulsion, and preparation reagent can be contained, such as suspending agent, stabilizer And/or dispersant, such as PVPP, agar or alginic acid or its salt, such as mosanom.

Parenteral includes the aqueous solution of water soluble active compound with pharmaceutical preparation.Active ingredient can be prepared as one sees fit The oily injection suspension of thing.Suitable lipophilic solvent or carrier include fat oil, such as sesame oil, or the aliphatic acid of synthesis Ester, such as ethyl oleate or triglycerides, or liposome, Aqueous injection suspensions can contain the material of increase suspension viscosity, example Such as sodium carboxymethylcellulose glucan.Alternatively, suspension can also contain suitable stabilizer or increase compound solubility Reagent, to prepare highly enriched solution.

Or active component can be powder type, suitable carrier regeneration is used before use, for example, it is sterile pyrogen-free Water.

Compound can also be configured to rectal compositions, such as suppository or enema,retention, such as the suppository containing routine Matrix, such as cocoa butter or other glyceride.

In addition to the formulations described previously, compound can also be configured to Drug Storage preparation.This kind of durative action preparation can pass through implantation Or percutaneous release（Such as subcutaneously or intramuscularly）, intramuscular injection or transdermal patch delivery.Thus, for example, compound can be with being adapted to Polymerization or hydrophobic material（Such as the emulsion in acceptable oil）Or ion exchange resin is prepared together, or it is configured to Microsolubility derivative, such as slightly soluble salt.

Pharmaceutical composition can also include suitable solid or gel phase carriers or excipient.This kind of carrier or excipient Example includes but is not limited to calcium carbonate, calcium phosphate, various sugar, starch, cellulose derivative, gelatin and polymer, such as poly- second Glycol.

The selection of administering mode and effective dose will be different in particular according to the disease treated.Administering mode and dosage Selection is in the limit of power of those skilled in the art.

Embodiment

Present disclosure is further proved with specific embodiment below, but is not meaned in any way to this hair It is bright to be limited.Method therefor is conventional method unless otherwise instructed in the following example.

The specifications and models of instrument and equipment and part raw material involved by the following example are as follows：

Ultraviolet spectrometer：The Pro types of Pharmacia companies Ultrospec 2100.

NMR：Bruker companies Bruker 500MHz（TMS is internal standard）.

Low Resolution Mass Spectra instrument：Waters LCMS ZQ 2000 (ESI mode).

High-pressure liquid phase system（Analysis）：Waters companies, 2 515 type pumps, 996 detectors.

Analytic type chromatographic column：Chromasil ODS column (4.6×250 mm, 10 µm).

High-pressure liquid phase system（Prepare）：It is prepared by Beijing Chuangxin Tongheng Science and Technology Co., Ltd. LC3000 type binary geopressure gradient half System.

Preparative scale chromatography post：Suzhou Na Wei companies Unisil 10-120 C18,10um, 21.2*250mm.

ELIASA：The Multilabel Counter of Perkin Elmer company Victor2 1420.

Chromatographic silica gel：300 ~ 400 mesh, Haiyang Chemical Plant, Qingdao.

Hplc grade methanol and acetonitrile are purchased from Honeywell companies, and other reagents are that analysis is pure, purchased from Tianjin big chemistry forever Reagent Company.

Sugared spore ketone compounds A and B can use any prior art to prepare in example below, and preparation method is in document [Boonlarppradab C, et al., The Journal of Antibiotics, 2013, 66(6): 305-309] In be described

Inventor's profit is prepared for compound A-C with the following method, and actinomyces N04W-1645 separates from the soil for pick up from Yunnan Arrive, be preserved in DSMZ of North China Pharmacuetical Group New Drug Research ＆ Development Co., Ltd, the public can be from North China pharmacy collection New Drug Research Co., Ltd of group obtains.

It is prepared by the extraction of 1 compound of the present invention of embodiment

Prepare N04W-1645 seed liquor

Bacterial strain N04W-1645 slant culture or spore liquid are inoculated in seed culture medium, 28 DEG C, the rpm of rotating speed 200 cultures 48 h obtain seed liquor；

Wherein described seed culture medium is made by the following method：20.0 grams of cornstarch, 4.0 grams of glucose, peptone 3.0 Gram, 4.0 grams of beef extract, 2.0 grams of yeast extract, add water to be settled to 1000 mL；7.0,121 DEG C of 30 min of sterilizing of pH value；

Prepare N04W-1645 zymotic fluid

Above-mentioned seed liquor is inoculated in fermentation medium, shaking flask or fermentation cylinder for fermentation with the % of percent by volume 8 inoculum concentration, sent out 27 DEG C of ferment temperature, the h of fermentation time 120；

Wherein described fermentation medium is made by the following method：30.0 grams of glucose, 5.0 grams of peptone, yeast extract 5.0 Gram, 2.0 grams of defatted soy flour, 1.0 grams of NaCl, CaCO₃6.0 grams, water is added to be settled to 1000 mL, 7.5,121 DEG C of sterilizings of pH value 30 min。

Tunning（Liquid fermentation）About 18L, centrifugation（4000 revs/min, 10 minutes）, it is divided into supernatant thalline.Thalline 6L, equal-volume acetone immersion is added, is centrifuged after 2h, acetone is removed in concentration, obtains thalline crude extract.The isometric ethyl acetate of supernatant Extraction, ethyl acetate phase evaporated under reduced pressure, obtains supernatant crude extract.Merge above-mentioned two parts extract, obtain the common 15g of paste.

Above-mentioned paste is dissolved with a small amount of methanol, silica gel mixed sample, in ODS compression leg (3.5 × 50 cm) chromatograph, acetonitrile and Water makees mobile phase, the elution of acetonitrile 10% ~ 100% (60min) linear concentration gradient, flow velocity 50mL/min, 245nm detection, is received by peak Collection, is numbered from first peak, is followed successively by 1-13 components, is concentrated to dryness, obtain 13 components（It is denoted as 1645-1 ~ 13））.

Prepared by the thick component 1.2g of 1645-7, methanol dissolving, liquid phase, prepare post to receive micro- Unisil 10-120 C18, 10um, 21.2*250mm, mobile phase CH₃CN-H₂O(27:73, V:V), flow velocity：18mL/min, UV245nm are detected, and obtain chemical combination Thing B（Rt=22min ,120mg）, compound C（Rt=30min ,8.7mg）；

Prepared by the thick component 0.7g of 1645-9, methanol dissolving, liquid phase, prepare post to receive micro- Unisil 10-120 C18,10um, 21.2*250mm mobile phase CH3CN-H2O (46:54, V:V), flow velocity：18mL/min, UV=245nm, obtain compound A（Rt =16min, 172mg）；

Compound A-C is identified respectively as sugared spore ketone A by MS and NMR data（Saccharosporone A）, sugared spore ketone B （Saccharosporone B）, sugared spore ketone D（Saccharosporone D）.

Compound A（Sugared spore ketone A, Saccharosporone A）：Yellow solid powder, it is soluble in chloroform, methanol.UVλ max(CH₃CN)=245, 276, 414nm。ESI-MS m/z 329[M+H]⁺, 327 [M-H]^-.Molecular formula C₁₉H₂₀O₅,¹H NMR (500 MHz, Acetone-d6) δ 12.15 (s, 1H), 7.71 (t, J=8.0Hz, 1H), 7.58 (d, J = 7.5 Hz, 1H), 7.26(d, J = 8.5 Hz, 1H), 4.04(m, 1H), 2.88-2.90(m, 2H), 2.78(t, J=8.8Hz, 1H), 2.29(m, 1H), 2.06(m, 1H), 1.92(m, 2H), 1.70(m, 1H), 1.39(t, J= 13.0Hz, 1H), 0.89(d, J=6.5Hz, 3H)。¹³C NMR (125 MHz, acetone-d6) δ191.7, 184.4, 161.9, 148.2, 142.0, 137.1, 133.6, 124.2, 119.3, 115.7, 73.4, 70.0, 50.6, 43.1, 35.4, 31.6, 30.4, 29.3, 23.3。

Compound B（Sugared spore ketone B, Saccharosporone B）：Yellow solid powder, it is soluble in chloroform, methanol.UVλmax (MeOH)=245, 275, 421 nm。ESI-MS m/z 345[M+H]⁺, 343[M-H]^-.Molecular formula C₁₉H₂₀O₆,¹H NMR (500 MHz, Acetone-d6) δ 12.11(s, 1H), 7.73(dd, J=8.4, 7.5Hz), 7.59(dd, J=7.5, 1.0Hz), 7.28(dd, J=8.4, 1.0Hz), 4.49(s, 1H), 4.25(d, J=5.3Hz, 2H), 4.09(m, 1H), 3.04(d, J=22.4Hz), 2.83(dd, J=19.1, 1.8Hz, 1H), 2.65(dd, J=19.1, 3.3Hz, 1H),2.12-2.14(m, 1H), 2.09-2.11(m, 1H), 1.84(m, 1H), 1.64(t, J=12.5Hz), 1.17 (m, 1H), 1.02(d, J=6.4Hz, 3H),。¹³C NMR (125 MHz, acetone-d6) δ 190.7, 182.7, 161.2, 145.8, 143.7, 136.3, 132.5, 123.6, 118.5, 115.4, 83.5, 76.7, 73.7, 46.8, 45.4, 39.5, 31.7, 25.9, 20.5。

Compound C（Sugared spore ketone D, Saccharosporone D）：Faint yellow solid powder, it is soluble in chloroform, methanol.UV λmax(MeOH)=236, 275(sh), 361nm。ESI-MS m/z 363[M+H]⁺.Molecular formula C₁₉H₂₂O₇,¹H NMR (500 MHz, Acetone-d6) δ 11.86(s,1H), 7.81 (t, J=8.0Hz,1H), 7.35 (d, J=8.3Hz,1H), 7.62 (d, J=7.5Hz,1H), 5.93(s,1H), 5.36(s,1H), 4.00(s,1H), 3.68(s,1H), 2.83 (dd, J=14.6, 2.3Hz, 1H), 2.38(dd, J=14.5, 3.1Hz, 1H), 2.14(1H,m), 2.05(dd, J= 1.7, 2.6Hz, 1H), 1.96(1H, ddd, J=13.7, 4.2, 1.7Hz), 1.46(m, 2H), 1.21(t, J= 12.9Hz,1H),0.94(d, J=6.6Hz, 3H), 0.88 (m, 1H), 0.72 (m, 1H)。¹³C NMR (125 MHz, acetone-d6) δ199.94, 197.6, 162.4, 137.2, 135.2, 123.6, 117.4, 116.7, 83.0, 77.0, 76.2, 71.2, 51.3, 48.8, 34.7, 32.2, 28.4, 27.8, 22.2。

2 compound of the present invention of embodiment determines to IDO or TDO inhibitory activity

With technique for gene engineering, the expression plasmid of IDO containing people and people's TDO genes is built respectively, weight is carried out in Escherichia coli Group expression, isolates and purifies recombined human IDO and TDO, and establish IDO or TDO activity monitor system.IDO or TDO activity methods Detection method：Using 96 well plate methods, detection buffer solution includes 40mM vitamin Cs, 400 μ g/ml hydrogen peroxide and 20 μM of methylenes The blue 50mM kaliumphosphate buffers of base（pH 6.5）.Appropriate IDO or TDO enzymes are added into detection buffer solution, are made into enzyme mixing Solution is added in 96 orifice plates, is then added substrate L-tryptophan, is reacted and 60min is carried out at 37 DEG C, adds 30%（w/v） Trichloroacetic acid makes reaction terminating.96 orifice plates are incubated 15min at 65 DEG C, are allowed to complete from formylkynurenine to kynurenin Conversion, 6000g centrifugation 5min, take out appropriate supernatant and be transferred in 96 new orifice plates, add 2%（w/v）To diformazan ammonia The acetic acid solution of benzaldehyde, kynurenin react the yellow color of production, and its OD value can determine under 490nm.Pressed down with IDO Preparation 1- methyl tryptophans（l-MT）As positive drug, compound is separately added into detection buffer solution, is made into compound concentration Difference 0.5,1,2,4,8,16,32,64ng/ml mixed liquor, 37 DEG C of insulation 5min, then add appropriate IDO into mixed liquor Or TDO enzymes, reacted according to above-mentioned condition, and OD values are detected under 490nm, data result is carried out using IC50 software for calculation Calculate.

Compound A-C of the present invention see the table below to IDO or TDO inhibitory activity.

Inhibitory activity of 3 compound of the present invention of embodiment to IDO in Hela cells

Hela cells are cultivated in 96 orifice plates, culture medium is penicillin containing 50U/ml, and 50U/ml streptomysins, 10%FBS's is high sugared DMEM, condition of culture are 37 DEG C, 5%CO₂.The gamma interferon that concentration is 200IU stimulates Hela cells, with IDO inhibitor 1- methyl tryptophans（l-MT）As positive drug, the positive drug and compound and 50 μM of substrate L- color ammonia of various concentrations are added Acid, after cultivating 48 hours, the supernatant of cell culture is taken out, adds 30%（w/v）Trichloroacetic acid, reaction system is in 65 DEG C of incubations 15min, is allowed to complete conversion from formylkynurenine to kynurenin, 12000rpm centrifugation 10min, take supernatant with it is isometric 2% （w/v）The acetic acid solution of paradime thylaminobenzaldehyde mixes reaction, detects OD values under 490nm, data result utilizes IC₅₀ Software for calculation is calculated.

Compound A-C of the present invention see the table below to the inhibitory action of IDO activity caused by interferon-induced Hela cells.

Claims (6)

1. following formula: compound

（ I ）

R is H or OH,

（ II ）,

The purposes in medicine in the disease for preparing treatment, preventing or slowing down the related pathological characteristicses of tryptophan metabolism.

2. purposes according to claim 1, wherein the disease is tumor immune escape, the nervous system disease, spiritual disease Disease or angiocardiopathy.

3. purposes according to claim 2, wherein tumor immune escape are the cancer of the brain, cutaneum carcinoma, carcinoma of urinary bladder, oophoroma, breast It is gland cancer, stomach cancer, cancer of pancreas, prostate cancer, colon cancer, leukemia, lung cancer, kidney, liver cancer, osteocarcinoma, incidence cancer, lymthoma, white Blood disease, melanoma.

4. purposes according to claim 2, wherein the nervous system disease be stages alzheimer's disease, Huntington disease, Parkinson's, Spinal muscular atrophy, ataxia telangiectasia.

5. purposes according to claim 2, wherein mental illness be anxiety, depression, mental handicape, insomnia, manic disorder, Obsession, neurosis, schizophrenia.

6. purposes according to claim 2, its cardiovascular disease is atherosclerosis, angina pectoris, miocardial infarction, the heart Low blood pressure, cerebral apoplexy, ischemic cardiomyopathy caused by restraining not normal, infection.