CN103664972B

CN103664972B - Diamino dihydrogen triazine derivative, its salt, preparation method, composition and application

Info

Publication number: CN103664972B
Application number: CN201310729225.6A
Authority: CN
Inventors: 周晓天; 林快乐; 周伟澄
Original assignee: Shanghai Institute of Pharmaceutical Industry; China State Institute of Pharmaceutical Industry
Current assignee: Shanghai Institute of Pharmaceutical Industry; China State Institute of Pharmaceutical Industry
Priority date: 2013-12-25
Filing date: 2013-12-25
Publication date: 2016-02-10
Anticipated expiration: 2033-12-25
Also published as: CN103664972A

Abstract

The invention discloses diamino dihydrogen triazine derivative, its salt, preparation method, composition and application.The preparation method of this diamino dihydrogen triazine derivative or its salt comprises: method one: compound of Formula IV and compounds of formula V are reacted and prepare compound of Formula I; Method two: in acid condition, mixes Formula VIII compound with Compounds of formula II, carries out ring-closure reaction, obtains the compound shown in general formula I.Present invention also offers above-mentioned diamino dihydrogen triazine derivative or the application of its salt in preparation people dihydrofolate reductase inhibitor, control tumour medicine or control bacterial infection disease medicine.Present invention also offers a kind of pharmaceutical composition, it contains above-mentioned diamino dihydrogen triazine derivative and/or its salt for the treatment of significant quantity, and pharmaceutically acceptable carrier.Diamino dihydrogen triazine spiroheterocyclic derivatives of the present invention has excellent inhibit activities to people's Tetrahydrofolate dehydrogenase, tumour cell, bacterium.

Description

Diamino dihydrogen triazine derivative, its salt, preparation method, composition and application

Technical field

The invention belongs to pharmaceutical chemistry synthesis technical field, be specifically related to have the diamino dihydrogen triazine derivative of spirane structure, its salt, preparation method, containing its composition and application.

Background technology

Tetrahydrofolate dehydrogenase (DHFR) in folic acid biological metabolic process is an important drug target.Tetrahydrofolate dehydrogenase is the key enzyme in folic acid biological metabolic process, and its major function is that in catalysis biological body, dihydrofolate reduction becomes tetrahydrofolic acid (THFA), and tetrahydrofolic acid (THFA) is coenzyme required in DNA building-up process.The structure of DHFR inhibitor is similar to the substrate of Tetrahydrofolate dehydrogenase, can emulatively be combined with DHFR, hinder the combination of DHFR and normal substrates, suppress its catalytic reduction activity, dihydrofolic acid is made to be difficult to be converted into tetrahydrofolic acid (THFA), interference folate cycle process, then disturbs the biosynthesizing of the DNA of pathogenic agent (as tumour cell and bacterium), finally causes the death of pathogen cells.Therefore, the inhibitor researching and developing Tetrahydrofolate dehydrogenase is the important directions of a drug research always.

The classical DHFR inhibitor be widely used, as methotrexate (Methotrexate, MTX), according to beating Qu Sha (Edetvexate, ETX), pemetrexed (ALIMTA), usually containing glutaminic acid residue in structure, need to enter cell by reduction folate carrier albumen active transport, by leaf acyl polyglutamic acid synthetic enzyme (FPGS) catalysis polyglutamic acid, then play a role.Polyglutamic acidization can cause DHFR inhibitor longer in the Intracellular retention time, easily causes Normocellular toxicity excessive; On the other hand, because DHFR inhibitor is the substrate of FPGS, easily cause FPGS too to express, thus inducing tumor cell produce resistance.In order to reduce toxicity and the resistance of DHFR inhibitor, research and development containing glutaminic acid residue, the micromolecular compound of non-FPGS substrate, be one of study hotspot of current DHFR inhibitor.

The people such as Ma Xiang had once synthesized a series of diamino dihydrogen triazine compound (XiangMa with spirane structure; ReneeSer-PengWoon; AntiproliferativeActivityAgainstMCF-7BreastCancerCellsby Diamino-TriazaspirodieneAntifolates, Chem.Biol.DrugDes.2009; 74:322 – 326), and done corresponding biological activity test respectively, it relates to compound 1 as follows, compound 2, compound 3, but in above-mentioned report, do not studied the relevant nature of triazine spiro-heterocycle compound, also the research of diamino dihydrogen triazine spiro-heterocycle compound structure activity relationship is not reported in prior art, thus the structure of modification for this compounds is subject to certain restrictions all the time, be difficult to find out their impact of structure for activity, therefore, the research for this compounds need further deeply.

Summary of the invention

The present invention want technical solution problem to be, provide a kind of novel structure, there is good people's Tetrahydrofolate dehydrogenase (hDHFR), tumour cell, the diamino dihydrogen triazine derivative of bacteriostatic activity, its salt, preparation method, composition and application.The present invention is that research diamino dihydrogen triazine derivative provides a new research direction, widened Research Thinking, and a series of diamino dihydrogen triazine spiroheterocyclic derivatives of the present invention has excellent inhibit activities to people's Tetrahydrofolate dehydrogenase (hDHFR), tumour cell, bacterium.

The invention provides a kind of such as formula the diamino dihydrogen triazine derivative shown in I or its salt,

Wherein,

X is oxygen (O) or sulphur (S);

N is 0,1,2,3 or 4;

R is hydrogen, phenyl, or substituted or unsubstituted phenyl oxygen base;

When n is 0, R is not substituted or unsubstituted phenyl oxygen base;

In the phenyl oxygen base of described replacement, described in be substituted by monosubstituted or polysubstituted; Substituting group is halogen, C _{1 ~ 4}alkoxyl group, C _{1 ~ 4}alkyl, nitro, C _{1 ~ 4}one or more in acyl group and cyano group; When described be substituted by polysubstituted time, the multiple substituting groups on the phenyl oxygen base of described replacement are identical or different.

In the phenoxy group of described replacement, when described substituting group is halogen, preferred fluorine, chlorine or bromine.

In the phenoxy group of described replacement, described substituting group is C _{1 ~ 4}during alkoxyl group, preferred methoxyl group, oxyethyl group or isopropoxy.

In the phenoxy group of described replacement, described substituting group is C _{1 ~ 4}during alkyl, preferable methyl, ethyl or the tertiary butyl.

In the phenoxy group of described replacement, described substituting group is C _{1 ~ 4}during acyl group, preferred ethanoyl.

The preferred hydrogen of described R, phenyl, phenyl oxygen base,

Described such as formula the diamino dihydrogen triazine derivative shown in I, preferably,

When X=O, n=0, R is H;

When X=O, n=1, R is phenyl;

When X=O, n=2, R is phenyl, 4-chlorophenoxy, 2,4 dichloro benzene oxygen base or 4-bromine phenoxy group;

X=O, during n=3, R is phenoxy group, 2,4-dichlorophenoxy, 4-fluorophenoxy, 4-methoxyphenoxy, 4-tertiary butyl phenoxy group, 4-chlorophenoxy, 4-methylphenoxy, 2,4,5-Trichlorophenoxy, 4-nitrophenoxy, 4-bromine phenoxy group, 4-ethanoyl phenoxy group, 4-cyano-benzene oxygen, 2,4-difluoro phenoxy group, 3,4-dichlorophenoxy, 2,3-dichlorophenoxy, 2-methoxyphenoxy, 3,5-dimethyl phenoxies, 2-chlorophenoxy, 3-chlorophenoxy, 2-bromine phenoxy group or 3-bromine phenoxy group;

When X=O, n=4, R is phenyl, 4-chlorophenoxy, 2,4 dichloro benzene oxygen base, 4-bromine phenoxy group;

When X=S, n=1, R is phenyl;

X=S, during n=3, R is 2,4-dichlorophenoxy, 4-methoxyphenoxy, 4-tertiary butyl phenoxy group, 4-chlorophenoxy, 4-bromine phenoxy group, 3,4-dichlorophenoxy, 2,3-dichlorophenoxy, 2-chlorophenoxy, 3-chlorophenoxy, 2-bromine phenoxy group, 3-bromine phenoxy group or 2,4,5-Trichlorophenoxy.

The described salt preferably salt hydrochlorate such as formula the diamino dihydrogen triazine derivative shown in I or hydrobromate.

Present invention also offers the above-mentioned preparation method such as formula the diamino dihydrogen triazine derivative shown in I or its salt, it is following either method:

Method one comprises the steps: compound of Formula IV and compounds of formula V to react to prepare compound of Formula I;

Wherein, Y is halogen, mesyloxy, trifluoro-methanesulfonyl oxy or tolysulfonyl oxygen base (described Y is preferably chlorine or bromine), and the definition of n, X, R as previously mentioned;

Method two comprises the steps: in acid condition, to be mixed by Formula VIII compound with Compounds of formula II, carries out ring-closure reaction, obtains the compound shown in general formula I;

Wherein, the definition of n, X, R as previously mentioned.

In method one, the preparation method of described compound of Formula I can reference Ma, Xiang; Chui, Wai-Keung.BioorganicandMedicinalChemistry, 2010, Vol.18, Issue2:737-743 are prepared, and the present invention preferably includes following steps:

(1) compound shown in general formula I V is in alcoholic solvent, with alkali reaction;

(2) compound shown in reaction solution step (1) obtained and general formula V reacts in aprotic polar solvent, obtains compound of Formula I.

Preferred 1:1 ~ the 1:4 of mol ratio of the compound shown in described general formula I V and the compound shown in general formula V, more preferably 1:1.2.

In step (1), the preferred mineral alkali of described alkali, as alkali-metal oxyhydroxide and/or alkali-metal carbonate.One or more in the preferred sodium hydroxide of described mineral alkali, potassium hydroxide, salt of wormwood and sodium carbonate, more preferably sodium hydroxide and/or potassium hydroxide.The mol ratio of described alkali and Formula VIII compound is preferably 1:1.

In step (1), described alcoholic solvent can be the alcoholic solvent that this area routine uses, preferred C ₁~ C ₄alcohol, more preferably methyl alcohol or ethanol.The consumption of described alcoholic solvent does not generally affect the carrying out of reaction, preferably 10 ~ 30ml/g formula IV compound.

In step (1), the temperature of described reaction preferably 50 ~ 80 DEG C.

In step (1), the process of described reaction is monitored by TLC or HPLC, take generally compound IV as unbound state, but not as the terminal of reaction during salify, preferred reaction of the present invention 30 minutes.

After step (1) reaction terminates, be further purified product by aftertreatment.Described aftertreatment preferably includes following steps: reaction system filtered while hot step (1) obtained, removes alcoholic solvent by filtrate, then carries out step (2) described reaction.

In step (2), the preferred acetonitrile of described aprotic polar solvent, DMF or methyl-sulphoxide, more preferably DMF.The consumption of described aprotic polar solvent does not generally affect the carrying out of reaction, preferably 3 ~ 8ml/g formula IV compound.

In step (2), the temperature of described reaction preferably 0 ~ 100 DEG C, more preferably 20 ~ 35 DEG C.

In step (2), the process of described reaction is monitored by TLC or HPLC, as the terminal of reaction when generally disappearing using compound IV (dissociating).

After step (2) reaction terminates, be further purified product by aftertreatment, described aftertreatment comprises the steps: the pH value of reaction system to be adjusted to 2 ~ 3, except desolventizing, and residuum recrystallization.Wherein, preferably mass concentration is adopted to be that 40% Hydrogen bromide carries out adjust ph.The described method except desolventizing preferably adopts underpressure distillation.The solvent of described recrystallization preferably adopts second alcohol and water, and its volume ratio is preferably 10:1 ~ 7:1(ethanol: water).

In method one, the preparation method of described compound of Formula IV can comprise the steps: that compound of formula III hydrogenation debenzylation obtains compound of Formula IV;

Wherein, the definition of X as previously mentioned.

The reaction conditions of the preparation method of described compound of Formula IV and step can refer to this type of popular response condition of reacting of this area and step is selected.The present invention is preferably as follows step:

The preparation method of described compound of Formula IV preferably includes: under palladium carbon catalytic condition, compound III mixed with aqueous ethanolic solution, under 0.2 ~ 2MPa hydrogen pressure, carries out hydrogenation debenzylation.

In the preparation method of described compound of Formula IV, described palladium carbon is preferably the palladium carbon of the palladium of 10% containing massfraction.The mass ratio of described palladium carbon and compound III is preferably 1% ~ 20%.

In the preparation method of described compound of Formula IV, described aqueous ethanolic solution preferred volume mark is the aqueous ethanolic solution of 40%.Described aqueous ethanolic solution is preferably 10 ~ 100mL/g compound III with the volume mass ratio of compound III.

In the preparation method of described compound of Formula IV, the temperature of reaction of described hydrogenation debenzylation preferably 10 DEG C ~ 50 DEG C.

In the preparation method of described compound of Formula IV, the process of described reaction is monitored by TLC or HPLC, as the terminal of reaction when generally disappearing using compound III.

After described hydrogenation debenzylation terminates, be further purified product by aftertreatment, described aftertreatment comprises the steps: suction filtration, concentrated filtrate, obtains solid recrystallization.The solvent that described recrystallization adopts is preferably second alcohol and water, and its volume ratio is preferably 5:1 ~ 10:1(ethanol: water).

In method one, the preparation method of described compound of formula III can comprise the steps: in acid condition, in solvent, benzyloxy semicarbazide hydrochloride, Dicyanodiamide and Compounds of formula II cyclization is obtained compound of formula III;

Wherein, the definition of X as previously mentioned.

The preparation method of described compound of formula III preferably includes following steps:

1, in solvent, by benzyloxy semicarbazide hydrochloride and Dicyanodiamide hybrid reaction;

2, in acid condition, mixture step 1 obtained and Compounds of formula II cyclization generate 2,4-diamino-5-benzyloxy-1,3,5-tri-azepine-9-oxa--spiral shell [5.5] undecane-1,3-bis-pinene hyhrochlorides shown in general formula III.

The physico-chemical property of 2,4-diamino-5-benzyloxy-1,3,5-tri-azepine-9-oxa--spiral shell [5.5] undecane-1,3-bis-pinene hyhrochlorides shown in general formula III is in table 1.

The reaction of described preparation compound of formula III can refer to document ZhouWeiCheng; XinZhiMing; ZhangXiuPing; ShenJie; QiuQiaoPing.ActaPharmaceuticaSinica, the preparation method of 1996, Vol.31, Issue11:823-830 is prepared, and the present invention is preferably as follows processing condition:

In the preparation method of described compound of formula III, the preferred 1:1 of mol ratio of described Dicyanodiamide and benzyloxy semicarbazide hydrochloride.

In the preparation method of described compound of formula III, the preferred 1:1 ~ 1:6 of mol ratio of described Compounds of formula II and benzyloxy semicarbazide hydrochloride.

In the preparation method of described compound of formula III, described acidic conditions preferably adopts concentrated hydrochloric acid to realize, and described concentrated hydrochloric acid refers generally to the aqueous hydrochloric acid that mass concentration is 36% ~ 38%.Preferred 1:1 ~ the 1.5:1 of mol ratio of described hydrochloric acid and benzyloxy semicarbazide hydrochloride.

In the preparation method of described compound of formula III, described solvent preferred alcohol.The volume mass of described solvent and benzyloxy semicarbazide hydrochloride is than preferred 4:1 ~ 10:1mL/g benzyloxy semicarbazide hydrochloride.

In the preparation method of described compound of formula III, in step 1, described reaction preferably includes following steps: in solvent, is mixed by benzyloxy semicarbazide hydrochloride with Dicyanodiamide, reacts 1 ~ 12 hour, then be placed to room temperature at 50 ~ 100 DEG C.

In the preparation method of described compound of formula III, in step 1, the temperature of described reaction preferably 78 ~ 80 DEG C.

In the preparation method of described compound of formula III, the reaction process of step 1 is monitored by TLC or HPLC, as the terminal of reaction when generally disappearing using benzyloxy semicarbazide hydrochloride.

After described step 1 reaction terminates, preferably directly carry out the reaction of step 2 without aftertreatment.

In the preparation method of described compound of formula III, in step 2, the temperature of described reaction preferably 10 ~ 35 DEG C.

In the preparation method of described compound of formula III, the reaction process of step 2 is monitored by TLC or HPLC, as the terminal of reaction when the biguanide compound generally generated using step 1 disappears, and preferably 3 ~ 10 days.

In the preparation method of described compound of formula III, be also further purified product by post-processing step after the reaction of described step 2 terminates, described post-processing step preferably includes: suction filtration, filter cake washing with alcohol, solid second alcohol and water recrystallization.In described recrystallization, the preferred 20:1 ~ 10:1 of volume ratio of second alcohol and water.

In method one, described compounds of formula V, except benzyl bromine, bromine ethylbenzene, 4-phenyl-1-butyl bromide are bought by business, all can prepare by following method: in acetonitrile, Compound I X and two halogen alkane substitutes, salt of wormwood hybrid reaction;

Wherein, Y is halogen, mesyloxy, trifluoro-methanesulfonyl oxy or tolysulfonyl oxygen base (described Y is preferably chlorine or bromine); As previously mentioned, and n is not 0 in the definition of n; R is substituted or unsubstituted phenyl oxygen base;

In compounds of formula V, Formula IX compound and compound of formula X, in the phenoxy group of described replacement, when described substituting group is halogen, preferred fluorine, chlorine or bromine.

In compounds of formula V, Formula IX compound and compound of formula X, in the phenoxy group of described replacement, described substituting group is C _{1 ~ 4}during alkoxyl group, preferred methoxyl group, oxyethyl group or isopropoxy.

In compounds of formula V, Formula IX compound and compound of formula X, in the phenoxy group of described replacement, described substituting group is C _{1 ~ 4}during alkyl, preferable methyl, ethyl or the tertiary butyl.

In compounds of formula V, Formula IX compound and compound of formula X, in the phenoxy group of described replacement, described substituting group is C _{1 ~ 4}during acyl group, preferred ethanoyl.

In compounds of formula V and Formula IX compound, described R preferred phenyl oxygen base,

In compounds of formula V, Formula IX compound and compound of formula X, preferably,

As n=2, R is phenyl, 4-chlorophenoxy, 2,4 dichloro benzene oxygen base, 4-bromine phenoxy group;

As n=3, R is phenoxy group, 2,4-dichlorophenoxy, 4-fluorophenoxy, 4-methoxyphenoxy, 4-tertiary butyl phenoxy group, 4-chlorophenoxy, 4-methylphenoxy, 2,4,5-Trichlorophenoxy, 4-nitrophenoxy, 4-bromine phenoxy group, 4-ethanoyl phenoxy group, 4-cyano group phenoxyl, 2,4-difluoro phenoxy group, 3,4-dichlorophenoxy, 2,3-dichlorophenoxy, 2-methoxyphenoxy, 3,5-dimethyl phenoxies, 2-chlorophenoxy, 3-chlorophenoxy, 2-bromine phenoxy group, 3-bromine phenoxy group;

As n=4, R is phenyl, 4-chlorophenoxy, 2,4 dichloro benzene oxygen base, 4-bromine phenoxy group.

The preparation method of described compounds of formula V can refer to this area ordinary method and is prepared, as can refer to document (HidekiKubota, ToshihiroWatanabe.Synthesisandpharmacologicalevaluationo fN-acyl-1,2,3,4-tetrahydroisoquinolinederivativesasnovelspecificbradyc ardicagents.Bioorganic & MedicinalChemistry, 2004,12:871-882).

In the preparation method of described compounds of formula V, described two halogen alkane substitutes preferably 1,3-dibromopropane, glycol dibromide or Isosorbide-5-Nitrae-dibromobutane.

In the preparation method of described compounds of formula V, the preferred 1:3 ~ 1:6 of mol ratio of described Compound I X and two halogen alkane substitutes.

In the preparation method of described compounds of formula V, the preferred 1:1 ~ 2:1 of mol ratio of described salt of wormwood and Compound I X.

In the preparation method of described compounds of formula V, described acetonitrile compares preferably 10 ~ 40mL/g Compound I X with the volume mass of Compound I X.

In the preparation method of described compounds of formula V, described temperature of reaction preferably 0 ~ 100 DEG C, more preferably 75 ~ 80 DEG C.

In the preparation method of described compounds of formula V, reaction process is monitored by TLC or HPLC, as the terminal of reaction when generally disappearing using Compound I X, and preferably 5 ~ 12 hours.

After the reaction preparing compounds of formula V terminates, be also further purified product by aftertreatment, described aftertreatment preferably includes following steps: reaction system is placed to room temperature, suction filtration, and filtrate concentrates, and extraction is dry, underpressure distillation.Described underpressure distillation, preferably under 0.48mbar pressure, gets the cut of 76 ~ 78 DEG C.

The described preparation method such as formula the diamino dihydrogen triazine derivative shown in I or its salt, in method two, described ring-closure reaction preferably includes following steps: in acid condition, by compound VI II and Compound II per hybrid reaction.

In method two, described acidic conditions preferably adopts concentrated hydrochloric acid, and described concentrated hydrochloric acid refers generally to the aqueous hydrochloric acid that mass concentration is 36% ~ 38%.Preferred 1:1 ~ the 1.5:1 of mol ratio of described concentrated hydrochloric acid and compound VI II.

In method two, the preferred 1:1 ~ 6:1 of mol ratio of described Compounds of formula II and Formula VIII compound.

In method two, the preferred room temperature of temperature of reaction of described ring-closure reaction.

In method two, the process of described ring-closure reaction is monitored by TLC or HPLC, as the terminal of reaction when general Formula VIII compound disappears, and preferably 3 ~ 10 days.

In method two, be also further purified product by aftertreatment after described ring-closure reaction terminates, described aftertreatment preferably includes following steps: suction filtration, solid recrystallization.Described recrystallization can adopt second alcohol and water, and its volume ratio is preferably 20:1 ~ 10:1(ethanol: water).

In method two, the preparation method of the biguanide compound shown in described general formula VIII can comprise the steps: in solvent, by Formula VII compound and Dicyanodiamide hybrid reaction, forms the biguanide compound shown in general formula VIII;

Wherein, described in the definition ditto of n, R.

In method two, reactions steps and the condition of the preparation method of the biguanide compound shown in described general formula VIII can refer to document: Ma, Xiang; Tan, Soo-Tong; Khoo, Chai-Ling; Sim, Hong-May; Chan, Lai-Wah; Chui, Wai-Keung.BioorganicandMedicinalChemistryLetters, 2011, Vol.21, Issue18:5428 – 5431 selects.The present invention is following reaction conditions particularly preferably:

In method two, the preparation method of the biguanide compound shown in described general formula VIII preferably includes following steps: in solvent, mixed by Formula VII compound with Dicyanodiamide, reacts 1 ~ 24 hour at 50 ~ 100 DEG C, be placed to room temperature again, form the biguanide compound shown in general formula VIII.

In method two, in the preparation method of the biguanide compound shown in described general formula VIII, the mol ratio of described Formula VII compound and Dicyanodiamide is preferably 1:1.

In method two, in the preparation method of the biguanide compound shown in described general formula VIII, described solvent is preferably ethanol.Described solvent compares preferably 5 ~ 30mL/g Formula VII compound with the volume mass of described Formula VII compound.

In method two, in the preparation method of the biguanide compound shown in described general formula VIII, the temperature of described reaction preferably 50 ~ 100 DEG C.

In method two, preferably, described general formula VII aminated compounds and Dicyanodiamide are reacted and are generated biguanide compound VIII, biguanide compound without separation direct one kettle way and Compounds of formula II in acid condition cyclization generate the target compound shown in general formula I.

In method two, the preparation method of described Formula VII compound can comprise the steps: in acid condition, and compound of formula VI is sloughed benzoyl and obtained Formula VII compound;

Wherein, the definition of n, R as previously mentioned.

In method two, step and the condition of the preparation method of described Formula VII compound can refer to document: Bailey, Stuart; Harnden, MichaelR.; Jarvest, RichardL.; Parkin, Ann; Boyd, MalcolmR.JournalofMedicinalChemistry, 1991, Vol.34, Issue1:57 – 65 selects, preferably include following steps: in alcoholic solvent, the N-substituted benzamide compound shown in general formula VI and concentrated hydrochloric acid reaction, hydrolysis obtains the aminated compounds of general formula VII.

In method two, in the preparation method of described Formula VII compound, described concentrated hydrochloric acid refers generally to the aqueous hydrochloric acid that mass percent is 36% ~ 38%.Hydrogenchloride in described concentrated hydrochloric acid and the mol ratio of compound of formula VI are preferably 1:1.

In method two, in the preparation method of described Formula VII compound, described alcoholic solvent particular methanol or ethanol.Described alcoholic solvent compares preferably 5 ~ 20mL/g compound of formula VI with the volume mass of compound of formula VI.

In method two, in the preparation method of described Formula VII compound, temperature of reaction preferably 50 ~ 80 DEG C.

In method two, the reaction process of the preparation method of Formula VII compound is monitored by TLC or HPLC, as the terminal of reaction when generally disappearing using compound of formula VI.

In method two, preparation compound VI I reaction terminate after, be also further purified product by aftertreatment.Described aftertreatment preferably includes following steps: concentration response system, solid recrystallization.Described recrystallization can adopt ethyl acetate and sherwood oil, and its volume ratio is preferably 1:1 ~ 1:4.

In method two, the preparation method of described compound of formula VI can comprise the steps: that benzyl hydroximic acid and compounds of formula V are obtained by reacting compound of formula VI;

Wherein, described in the definition ditto of n, R, Y.

In method two, the preparation method of described compound of formula VI preferably includes following steps: in alcoholic solvent, and the reactant aqueous solution of benzyl hydroximic acid, compounds of formula V and alkali prepares the N-substituted benzamide compound shown in general formula VI.

In the preparation method of described compound of formula VI, described alcoholic solvent particular methanol or ethanol (being generally the ethanol that volumetric concentration is 95%).Described alcoholic solvent compares preferably 10 ~ 20mL/g benzyl hydroximic acid with the volume mass of benzyl hydroximic acid.

In the preparation method of described compound of formula VI, the preferred mineral alkali of described alkali, as alkali-metal oxyhydroxide and/or alkali-metal carbonate.One or more in the preferred sodium hydroxide of described mineral alkali, potassium hydroxide, salt of wormwood and sodium carbonate, more preferably sodium hydroxide and/or potassium hydroxide.Preferred 1:1 ~ the 1.2:1 of mol ratio of described alkali and benzyl hydroximic acid.

In the preparation method of described compound of formula VI, the temperature of reaction is preferred: 0 ~ 100 DEG C, be more preferably 60-79 DEG C.The molar ratio of described benzyl hydroximic acid and compounds of formula V is preferably 1:1.

In the preparation method of described compound of formula VI, the process of reaction is monitored by TLC or HPLC, as the terminal of reaction when generally disappearing using benzyl hydroximic acid.

After the reaction preparing compound of formula VI terminates, be further purified product by aftertreatment.Described aftertreatment preferably includes following steps: system is placed to room temperature, and concentrated, suction filtration, filter cake washes with water, solid recrystallization.Described recrystallization preferably adopts ethyl acetate and sherwood oil, and its volume ratio is preferably 5:1 ~ 10:1.

In method two, described compounds of formula V, except benzyl bromine, bromine ethylbenzene, 4-phenyl-1-butyl bromide are bought by business, all can prepare by following method: in acetonitrile, Compound I X and two halogen alkane substitutes, salt of wormwood hybrid reaction;

Described R preferred phenyl oxygen base,

Described compounds of formula V preferably,

As n=2, R is phenyl, 4-chlorophenoxy, 2,4 dichloro benzene oxygen base or 4-bromine phenoxy group;

As n=3, R is phenoxy group, 2,4-dichlorophenoxy, 4-fluorophenoxy, 4-methoxyphenoxy, 4-tertiary butyl phenoxy group, 4-chlorophenoxy, 4-methylphenoxy, 2,4,5-Trichlorophenoxy, 4-nitrophenoxy, 4-bromine phenoxy group, 4-ethanoyl phenoxy group, 4-cyano group phenoxyl, 2,4-difluoro phenoxy group, 3,4-dichlorophenoxy, 2,3-dichlorophenoxy, 2-methoxyphenoxy, 3,5-dimethyl phenoxies, 2-chlorophenoxy, 3-chlorophenoxy, 2-bromine phenoxy group or 3-bromine phenoxy group;

As n=4, R is phenyl, 4-chlorophenoxy, 2,4 dichloro benzene oxygen base or 4-bromine phenoxy group.

Present invention also offers a kind of pharmaceutical composition, it contains the above-mentioned such as formula the diamino dihydrogen triazine derivative shown in I and/or its salt for the treatment of significant quantity, and pharmaceutically acceptable carrier.

In the present invention, described " treatment effective dose " refers to (i) to prevent or treat the amount of the compounds of this invention of disease specific described in the application or illness, (ii) weaken, improve or eliminate the amount of the compounds of this invention of one or more symptoms of disease specific described in the application or illness, or the amount of the compounds of this invention of the (iii) outbreak of one or more symptoms of prevention or the disease specific postponed described in the application or illness.The dosage for the treatment of human patients can be 0.0001mg/kg ~ 50mg/kg, is generally 0.001mg/kg ~ 10mg/kg body weight most, such as, within the scope of 0.01mg/kg ~ 1mg/kg.Such dosage can give such as every day 1-5 time.

Described pharmaceutically acceptable carrier is the carrier that preparation conventional in this area adopts.

In the present invention, according to therapeutic purpose, pharmaceutical composition can be made various types of administration unit dosage, as tablet, pill, pulvis, liquid, suspension, emulsion, granule, capsule, suppository and injection (solution and suspension) etc., preferred liquid, suspension, emulsion, suppository and injection (solution and suspension) etc.

In order to make the pharmaceutical composition of tablet form be shaped, any known and widely used vehicle in this area can be used.Such as, carrier, as lactose, white sugar, sodium-chlor, glucose, urea, starch, calcium carbonate, kaolin, crystalline cellulose and silicic acid etc.; Tackiness agent, as water, ethanol, propyl alcohol, common syrup, glucose solution, starch solution, gelatin solution, carboxymethyl cellulose, lac, methylcellulose gum and potassiumphosphate, polyvinylpyrrolidone etc.; Disintegrating agent, as dry starch, sodiun alginate, agar powder and Kelp Powder, the fatty acid ester of sodium bicarbonate, calcium carbonate, polyethylene sorbitan, sodium lauryl sulphate, stearic acid monoglycerides, starch and lactose etc.; Disintegration inhibitor, as white sugar, tristearin, Oleum Cocois and winterized stearin; Adsorption enhancer, as quaternary amine alkali and sodium lauryl sulphate etc.; Wetting agent, as glycerine, starch etc.; Sorbent material, as starch, lactose, kaolin, wilkinite and colloid silicic acid etc.; And lubricant, as pure talcum, stearate, boric acid powder and polyoxyethylene glycol etc.Common painting stain material can also be selected as required to make sugar coated tablet, be coated with gelatin film tablet, enteric coated tablets, film coated tablets, duplicature tablet and multilayer tablet.

In order to make the pharmaceutical composition of pill be shaped, any known and widely used vehicle in this area can be used, such as, carrier, as lactose, starch, Oleum Cocois, hardened vegetable oils, kaolin and talcum powder etc.; Tackiness agent, as gum arabic powder, tragacanth gum powder, gelatin and ethanol etc.; Disintegrating agent, as agar and Kelp Powder etc.

In order to make the pharmaceutical composition of suppository form be shaped, any known and widely used excipient in this area can be used, such as, polyoxyethylene glycol, Oleum Cocois, higher alcohols, the ester of higher alcohols, gelatin and semisynthetic glyceryl ester etc.

In order to prepare the pharmaceutical composition of injection form, by after solution or suspension sterilization (preferably adding appropriate sodium-chlor, glucose or glycerine etc.), the isotonic injection with blood can be made.When preparing injection, any conventional carrier in this area also can be used.Such as, water, ethanol, propylene glycol, the isooctadecanol of ethoxylation, the isooctadecanol of polyoxy and the fatty acid ester etc. of polyethylene sorbitan.In addition, also common solvating agent, buffer reagent and pain killer etc. can be added.

In the present invention, the content of described composition in pharmaceutical composition, without particular restriction, can be selected, usually be can be 10 ~ 90% of mass percent in very wide scope, is preferably mass percent 30 ~ 80%.

In the present invention, the medication of the pharmaceutical composition of described composition is not particularly limited.According to patient age, sex and other condition and symptom, the preparation administration of various formulation can be selected.Such as, tablet, pill, solution, suspension, emulsion, granule or capsule oral administration; Injection can be individually dosed, or be mixed into row vein injection with injection conveying liquid (as glucose solution and amino acid solution); Suppository is for being administered into rectum.

Present invention also offers and above-mentionedly preparing the application in antitumor drug, resistance to bacteria infection medicine or people's Tetrahydrofolate dehydrogenase (hDHFR) inhibitor such as formula the diamino dihydrogen triazine derivative shown in I, its salt or described pharmaceutical composition.

Described tumour can be adenocarcinoma of lung, colorectal carcinoma, leukemia or lymphatic cancer.

Described bacterial infection can be pneumococcus infection, gamma streptococcus infection, infection of staphylococcus aureus or Staphylococcus albus and infects.

Described people's Tetrahydrofolate dehydrogenase (hDHFR) inhibitor preferably treats and/or prevents the medicine of autoimmune disorder.Described autoimmune disorder can be rheumatoid arthritis or lupus erythematosus.

Without prejudice to the field on the basis of common sense, above-mentioned each optimum condition, can arbitrary combination, obtains the preferred embodiments of the invention.

Agents useful for same of the present invention and raw material are all commercially.

Positive progressive effect of the present invention is: the present invention is that research diamino dihydrogen triazine derivative provides a new research direction, widened Research Thinking, and a series of diamino dihydrogen triazine spiroheterocyclic derivatives of the present invention has excellent inhibit activities to people's Tetrahydrofolate dehydrogenase (hDHFR), tumour cell, bacterium.

Embodiment

Mode below by embodiment further illustrates the present invention, but does not therefore limit the present invention among described scope of embodiments.The experimental technique of unreceipted actual conditions in the following example, conventionally and condition, or selects according to catalogue.

In the examples below, undefined abbreviation has its generally accepted implication, and unless stated otherwise, all room temperatures all refer to temperature 20 DEG C ~ 30 DEG C.

Benzyloxy semicarbazide hydrochloride, Tetrahydro-pyran-4-one are bought from Shanghai De Mo Pharmaceutical Technology Co., Ltd, purity >98%;

Tetrahydric thiapyran-4-ketone is bought from Shanghai De Mo Pharmaceutical Technology Co., Ltd, purity >98%.

Embodiment of the method 1

The bromo-3-phenoxypropane of 1-

Phenol (4.7g, 0.05mol), 1,3-dibromopropane (50g, 0.25mol), salt of wormwood (10g, 0.075mol) are mixed in 100ml acetonitrile, react 9 hours at 80 DEG C.When TLC monitoring phenol runs out of, stopped reaction, is placed to room temperature, and suction filtration removing solid, filtrate reduced in volume, adds 100ml chloroform, and the aqueous sodium hydroxide solution of chloroform layer 0.2N is washed twice (100ml*2), uses anhydrous magnesium sulfate drying.Underpressure distillation, under 0.48mbar pressure, gets the cut of 76 ~ 78 DEG C, obtains colorless oil 6.7g, yield 62.2%.Concrete grammar reference (HidekiKubota, ToshihiroWatanabe.Synthesisandpharmacologicalevaluationo fN-acyl-1,2,3,4-tetrahydroisoquinolinederivativesasnovelspecificbradyc ardicagents.Bioorganic & MedicinalChemistry, 2004,12:871-882).

Compounds of formula V, except benzyl bromine, bromine ethylbenzene, 4-phenyl-1-butyl bromide are bought by business, method prepares all thus.

Embodiment of the method 2

The bromo-3-(2 ' of 1-, 4 '-dichlorophenoxy) propane

2,4 dichloro phenol (8.2g, 0.05mol), 1,3-dibromopropane (50g, 0.25mol), salt of wormwood (10g, 0.075mol) are mixed in 100ml acetonitrile, react 8 hours at 80 DEG C.When TLC monitoring phenol runs out of, stopped reaction, is placed to room temperature, and suction filtration removing solid, filtrate reduced in volume, adds 100ml chloroform, and the aqueous sodium hydroxide solution of chloroform layer 0.2N is washed twice (100ml*2), uses anhydrous magnesium sulfate drying.Underpressure distillation, under 0.3mbar pressure, gets the cut of 106 ~ 108 DEG C, obtains colorless oil 8.5g, yield 60.5%.

Embodiment of the method 3

1-bromo-3-(4 '-methoxyphenoxy) propane

4-methoxyphenol (6.2g, 0.05mol), 1,3-dibromopropane (50g, 0.25mol), salt of wormwood (10g, 0.075mol) are mixed in 100ml acetonitrile, react 9 hours at 80 DEG C.When TLC monitoring phenol runs out of, stopped reaction, is placed to room temperature, and suction filtration removing solid, filtrate reduced in volume, adds 100ml chloroform, and the aqueous sodium hydroxide solution of chloroform layer 0.2N is washed twice (100ml*2), uses anhydrous magnesium sulfate drying.Underpressure distillation, under 0.5mbar pressure, gets the cut of 98 ~ 102 DEG C, obtains colorless oil 8.1g, yield 66.1%.

Embodiment of the method 4

The bromo-3-(4 ' of 1--tertiary butyl phenoxy group) propane

4-TBP (7.5g, 0.05mol), 1,3-dibromopropane (50g, 0.25mol), salt of wormwood (10g, 0.075mol) are mixed in 100ml acetonitrile, react 8 hours at 80 DEG C.When TLC monitoring phenol runs out of, stopped reaction, is placed to room temperature, and suction filtration removing solid, filtrate reduced in volume, adds 100ml chloroform, and the aqueous sodium hydroxide solution of chloroform layer 0.2N is washed twice (100ml*2), uses anhydrous magnesium sulfate drying.Underpressure distillation, under 0.5mbar pressure, gets the cut of 110 ~ 114 DEG C, obtains colorless oil 9.4g, yield 70.5%.

Embodiment of the method 5

1-bromo-3-(4 '-chlorophenoxy) propane

4-chlorophenol (6.5g, 0.05mol), 1,3-dibromopropane (50g, 0.25mol), salt of wormwood (10g, 0.075mol) are mixed in 100ml acetonitrile, react 8 hours at 80 DEG C.When TLC monitoring phenol runs out of, stopped reaction, is placed to room temperature, and suction filtration removing solid, filtrate reduced in volume, adds 100ml chloroform, and the aqueous sodium hydroxide solution of chloroform layer 0.2N is washed twice (100ml*2), uses anhydrous magnesium sulfate drying.Underpressure distillation, under 0.3mbar pressure, gets the cut of 78 ~ 82 DEG C, obtains colorless oil 8.6g, yield 69.3%.

Embodiment of the method 6

The bromo-3-(2 ' of 1-, 4 ', 5 '-chlorophenoxy) propane

2 ', 4 ', 5 '-chlorophenol (9.85g, 0.05mol), 1,3-dibromopropane (50g, 0.25mol), salt of wormwood (10g, 0.075mol) are mixed in 100ml acetonitrile, react 8 hours at 80 DEG C.When TLC monitoring phenol runs out of, stopped reaction, is placed to room temperature, and suction filtration removing solid, filtrate reduced in volume, adds 100ml chloroform, and the aqueous sodium hydroxide solution of chloroform layer 0.2N is washed twice (100ml*2), uses anhydrous magnesium sulfate drying.Underpressure distillation, under 0.5mbar pressure, gets the cut of 122 ~ 124 DEG C, obtains colorless oil 10g, yield 63.2%.

Embodiment of the method 7

The bromo-3-(4 ' of 1--bromine phenoxy group) propane

4-bromophenol (8.6g, 0.05mol), 1,3-dibromopropane (50g, 0.25mol), salt of wormwood (10g, 0.075mol) are mixed in 100ml acetonitrile, react 8 hours at 80 DEG C.When TLC monitoring phenol runs out of, stopped reaction, is placed to room temperature, and suction filtration removing solid, filtrate reduced in volume, adds 100ml chloroform, and the aqueous sodium hydroxide solution of chloroform layer 0.2N is washed twice (100ml*2), uses anhydrous magnesium sulfate drying.Underpressure distillation, under 0.4mbar pressure, gets the cut of 116 ~ 120 DEG C, obtains colorless oil 9.4g, yield 64.8%.

Embodiment of the method 8

The bromo-3-(3 ' of 1-, 4 '-dichlorophenoxy) propane

3,4-chlorophenesic acid (8.2g, 0.05mol), 1,3-dibromopropane (50g, 0.25mol), salt of wormwood (10g, 0.075mol) are mixed in 100ml acetonitrile, react 8 hours at 80 DEG C.When TLC monitoring phenol runs out of, stopped reaction, is placed to room temperature, and suction filtration removing solid, filtrate reduced in volume, adds 100ml chloroform, and the aqueous sodium hydroxide solution of chloroform layer 0.2N is washed twice (100ml*2), uses anhydrous magnesium sulfate drying.Underpressure distillation, under 0.4mbar pressure, gets the cut of 121 ~ 123 DEG C, obtains colorless oil 8.5g, yield 69.7%.

Embodiment of the method 9

The bromo-2-(2 ' of 1-, 4 '-dichlorophenoxy) ethane

2,4 dichloro phenol (8.2g, 0.05mol), glycol dibromide (48g, 0.25mol), salt of wormwood (10g, 0.075mol) are mixed in 100ml acetonitrile, react 8 hours at 80 DEG C.When TLC monitoring phenol runs out of, stopped reaction, is placed to room temperature, and suction filtration removing solid, filtrate reduced in volume, adds 100ml chloroform, and the aqueous sodium hydroxide solution of chloroform layer 0.2N is washed twice (100ml*2), uses anhydrous magnesium sulfate drying.Underpressure distillation, under 0.55mbar pressure, gets the cut of 99 ~ 101 DEG C, obtains colorless oil 9.5g, yield 71.1%.

Embodiment of the method 10

The bromo-4-(2 ' of 1-, 4 '-dichlorophenoxy) butane

2,4 dichloro phenol (8.2g, 0.05mol), Isosorbide-5-Nitrae-dibromobutane (52g, 0.25mol), salt of wormwood (10g, 0.075mol) are mixed in 100ml acetonitrile, react 8 hours at 80 DEG C.When TLC monitoring phenol runs out of, stopped reaction, is placed to room temperature, and suction filtration removing solid, filtrate reduced in volume, adds 100ml chloroform, and the aqueous sodium hydroxide solution of chloroform layer 0.2N is washed twice (100ml*2), uses anhydrous magnesium sulfate drying.Underpressure distillation, under 0.65mbar pressure, gets the cut of 129 ~ 130 DEG C, obtains colorless oil 11.1g, yield 74.5%.

Embodiment of the method 11

2,4-diamino-5-benzyloxy-1,3,5-tri-azepine-9-oxa--spiral shell [5.5] certain herbaceous plants with big flowers alkane-1,3-bis-pinene hyhrochloride (A32)

Benzyloxy semicarbazide hydrochloride (2.45g, 0.015mol), Dicyanodiamide (1.3g, 0.015mol) are mixed in there-necked flask, add 15ml ethanol, and reflux 2 hours, is placed into room temperature.Adding Tetrahydro-pyran-4-one (3.8g, 0.038mol), concentrated hydrochloric acid (1.5g, 0.015mol), there is white precipitate in stirring at room temperature 5 days.Suction filtration, a small amount of cold ethanol of filter cake is washed, gained white solid ethanol: water=10:1(volume ratio) recrystallization, obtain white crystal 3.01g, yield 60.3%.The physico-chemical property of A32 in table 3, A32's ¹hNMR data are in table 4.

Embodiment of the method 12

2,4-diamino-5-hydroxyl-1,3,5-tri-azepine-9-oxa--spiral shell [5.5] undecane-1,3-bis-pinene hyhrochloride (A0)

2,4-diamino-5-benzyl-1,3,5-tri-azepine-9-oxa--spiral shell [5.5] undecane-1,3-bis-pinene hyhrochloride (2g, 0.006mol), 10% Pb/C(0.2g), 40% aqueous ethanolic solution 100ml is suspended in hydriding reactor, pass into hydrogen, under 1MPa pressure, stirring at room temperature.When TLC monitoring raw material consumption is complete, stopped reaction, suction filtration, the concentrated institute of filtrate obtains white solid ethanol: water=8:1(volume ratio) recrystallization, obtain white crystal 0.9g, yield 64.3%.The physico-chemical property of A0 in table 3, A0's ¹hNMR data are in table 4.

Embodiment of the method 13

2,4-diamino-5-(3 '-phenoxy-propoxy)-1,3,5-tri-azepine-9-oxa--spiral shell [5.5] undecane-1,3-diene hydrobromate (A1)

2,4-diamino-5-hydroxyl-1,3,5-tri-azepine-9-oxa--spiral shell [5.5] undecane-1,3-bis-pinene hyhrochloride (0.47g, 0.002mol), sodium hydroxide (0.08g, 0.002mol), methyl alcohol 10ml is mixed in round-bottomed bottle, reflux 30 minutes, heat filter, filtrate concentrates.Add the bromo-3-phenoxypropane (0.52g, 0.0024mol) of 1-and DMF2ml, stirring at room temperature.When TLC monitoring raw material consumption is complete, instills a small amount of concentrated hydrobromic acid and regulate pH value to be 2 ~ 3.Removal of solvent under reduced pressure, obtains white solid, with ethanol: water=10:1(volume ratio) recrystallization, obtain white crystal 0.26g, yield 31.7%, HPLC records purity 99.6%.

The synthetic method of A2 ~ A31 is identical with A1, the physico-chemical property of A1 ~ A31, mass spectrum ESI data in table 3, A1 ~ A31's ¹hNMR data are in table 4.

Embodiment of the method 14

2,4-diamino-5-(3 '-(2 ' ', 4 ' '-dichlorophenoxy)-propoxy-)-1,3,5-tri-azepine-9-oxa--spiral shell [5.5] undecanes-1,3-diene hydrobromate (A2)

2,4-diamino-5-hydroxyl-1,3,5-tri-azepine-9-oxa--spiral shell [5.5] undecane-1,3-bis-pinene hyhrochloride (0.47g, 0.002mol), sodium hydroxide (0.08g, 0.002mol), methyl alcohol 10ml is mixed in round-bottomed bottle, reflux 30 minutes, heat filter, filtrate concentrates.Add the bromo-3-(2 ' of 1-, 4 '-dichlorophenoxy) propane (0.68g, 0.0024mol) and DMF2ml, stirring at room temperature.When TLC monitoring raw material consumption is complete, instills a small amount of concentrated hydrobromic acid and regulate pH value to be 2 ~ 3.Removal of solvent under reduced pressure, obtains white solid, with ethanol: water=10:1(volume ratio) recrystallization, obtain white crystal 0.41g, yield 42.5%, HPLC records purity 99.8%.

Embodiment of the method 15

2,4-diamino-5-(3 '-(4 ' '-methoxyphenoxy)-propoxy-)-1,3,5-tri-azepine-9-oxa--spiral shell [5.5] undecanes-1,3-diene hydrobromate (A4)

2,4-diamino-5-hydroxyl-1,3,5-tri-azepine-9-oxa--spiral shell [5.5] undecane-1,3-bis-pinene hyhrochloride (0.53g, 0.002mol), sodium hydroxide (0.08g, 0.002mol), methyl alcohol 10ml is mixed in round-bottomed bottle, reflux 30 minutes, heat filter, filtrate concentrates.Add 1-bromo-3-(4 '-methoxyphenoxy) propane (0.58g, 0.0024mol) and DMF2ml, stirring at room temperature.When TLC monitoring raw material consumption is complete, instills a small amount of concentrated hydrobromic acid and regulate pH value to be 2 ~ 3.Removal of solvent under reduced pressure, obtains white solid, with ethanol: water=10:1(volume ratio) recrystallization, obtain white crystal 0.42g, yield 62.9%, HPLC records purity 99.5%.

Embodiment of the method 16

2,4-diamino-5-(3 '-(4 ' '-tertiary butyl phenoxy group)-propoxy-)-1,3,5-tri-azepine-9-oxa--spiral shell [5.5] undecanes-1,3-diene hydrobromate (A5)

2,4-diamino-5-hydroxyl-1,3,5-tri-azepine-9-oxa--spiral shell [5.5] undecane-1,3-bis-pinene hyhrochloride (0.47g, 0.002mol), sodium hydroxide (0.08g, 0.002mol), methyl alcohol 10ml is mixed in round-bottomed bottle, reflux 30 minutes, heat filter, filtrate concentrates.Add the bromo-3-(4 ' of 1--tertiary butyl phenoxy group) propane (0.68g, 0.0024mol) and DMF2ml, stirring at room temperature.When TLC monitoring raw material consumption is complete, instills a small amount of concentrated hydrobromic acid and regulate pH value to be 2 ~ 3.Removal of solvent under reduced pressure, obtains white solid, with ethanol: water=10:1(volume ratio) recrystallization, obtain white crystal 0.39g, yield 42.0%, HPLC records purity 99.7%.

Embodiment of the method 17

2,4-diamino-5-(3 '-(4 ' '-chlorophenoxy)-propoxy-)-1,3,5-tri-azepine-9-oxa--spiral shell [5.5] undecanes-1,3-diene hydrobromate (A6)

2,4-diamino-5-hydroxyl-1,3,5-tri-azepine-9-oxa--spiral shell [5.5] undecane-1,3-bis-pinene hyhrochloride (0.47g, 0.002mol), sodium hydroxide (0.08g, 0.002mol), methyl alcohol 10ml is mixed in round-bottomed bottle, reflux 30 minutes, heat filter, filtrate concentrates.Add 1-bromo-3-(4 '-chlorophenoxy) propane (0.55g, 0.0024mol) and DMF2ml, stirring at room temperature.When TLC monitoring raw material consumption is complete, instills a small amount of concentrated hydrobromic acid and regulate pH value to be 2 ~ 3.Removal of solvent under reduced pressure, obtains white solid, with ethanol: water=10:1(volume ratio) recrystallization, obtain white crystal 0.42g, yield 47.5%, HPLC records purity 99.5%.

Embodiment of the method 18

2,4-diamino-5-(3 '-(2 ' ', 4 ' ', 5 ' ' and-Trichlorophenoxy)-propoxy-)-1,3,5-tri-azepine-9-oxa--spiral shell [5.5] undecanes-1,3-diene hydrobromate (A8)

2,4-diamino-5-hydroxyl-1,3,5-tri-azepine-9-oxa--spiral shell [5.5] undecane-1,3-bis-pinene hyhrochloride (0.47g, 0.002mol), sodium hydroxide (0.08g, 0.002mol), methyl alcohol 10ml is mixed in round-bottomed bottle, reflux 30 minutes, heat filter, filtrate concentrates.', 5 ' '-Trichlorophenoxy that adds the bromo-3-(2 ' ' of 1-, 4 ') propane (0.68g, 0.0024mol) and DMF2ml, stirring at room temperature.When TLC monitoring raw material consumption is complete, instills a small amount of concentrated hydrobromic acid and regulate pH value to be 2 ~ 3.Removal of solvent under reduced pressure, obtains white solid, with ethanol: water=10:1(volume ratio) recrystallization, obtain white crystal 0.31g, yield 29.5%, HPLC records purity 99.6%.

Embodiment of the method 19

2,4-diamino-5-(3 '-(4 ' '-bromine phenoxy group)-propoxy-)-1,3,5-tri-azepine-9-oxa--spiral shell [5.5] undecanes-1,3-diene hydrobromate (A10)

2,4-diamino-5-hydroxyl-1,3,5-tri-azepine-9-oxa--spiral shell [5.5] undecane-1,3-bis-pinene hyhrochloride (0.47g, 0.002mol), sodium hydroxide (0.08g, 0.002mol), methyl alcohol 10ml is mixed in round-bottomed bottle, reflux 30 minutes, heat filter, filtrate concentrates.Add the bromo-3-(4 ' of 1--bromine phenoxy group) propane (0.7g, 0.0024mol) and DMF2ml, stirring at room temperature.When TLC monitoring raw material consumption is complete, instills a small amount of concentrated hydrobromic acid and regulate pH value to be 2 ~ 3.Removal of solvent under reduced pressure, obtains white solid, with ethanol: water=10:1(volume ratio) recrystallization, obtain white crystal 0.4g, yield 40.8%, HPLC records purity 99.6%.

Embodiment of the method 20

2,4-diamino-5-(3 '-(3 ' ', 4 ' '-dichlorophenoxy)-propoxy-)-1,3,5-tri-azepine-9-oxa--spiral shell [5.5] undecanes-1,3-diene hydrobromate (A19)

2,4-diamino-5-hydroxyl-1,3,5-tri-azepine-9-oxa--spiral shell [5.5] undecane-1,3-bis-pinene hyhrochloride (0.47g, 0.002mol), sodium hydroxide (0.08g, 0.002mol), methyl alcohol 10ml is mixed in round-bottomed bottle, reflux 30 minutes, heat filter, filtrate concentrates.Add the bromo-3-(3 ' of 1-, 4 '-dichlorophenoxy) propane (0.62g, 0.0024mol) and DMF2ml, stirring at room temperature.When TLC monitoring raw material consumption is complete, instills a small amount of concentrated hydrobromic acid and regulate pH value to be 2 ~ 3.Removal of solvent under reduced pressure, obtains white solid, with ethanol: water=10:1(volume ratio) recrystallization, obtain white crystal 0.29g, yield 30.4%, HPLC records purity 99.7%.

Embodiment of the method 21

N-[1 '-bromo-3 '-(4 ' '-bromo-phenoxy group) propyl group oxygen base] benzamide

Benzyl hydroximic acid (11.7g, 0.085mol), sodium hydroxide (3.2g, 0.08mol, be dissolved in 10ml water), the bromo-3-(4 ' of 1--bromo-phenoxy group) propane (25g, 0.085mol), 95% ethanol 150ml is mixed in round-bottomed bottle, reflux, stopped reaction when TLC monitoring raw material consumption is complete, is placed as room temperature, concentrated, suction filtration, a large amount of washing of filter cake, obtains light red solid, by ethyl acetate: sherwood oil=7:1(volume ratio) recrystallization, obtain white solid 13.9g, yield 46.7%, MP=110 ~ 111 DEG C.

Compound of formula VI all thus method prepares.

Embodiment of the method 22

N-[1 '-bromo-3 '-(4 ' '-chloro-phenoxy group) propyl group oxygen base] benzamide

Benzyl hydroximic acid (11.7g, 0.085mol), sodium hydroxide (3.2g, 0.08mol, be dissolved in 10ml water), the chloro-phenoxy group of 1-bromo-3-(4 ’ –) propane (21g, 0.085mol), 95% ethanol 150ml is mixed in round-bottomed bottle, reflux, stopped reaction when TLC monitoring raw material consumption is complete, is placed as room temperature, concentrated, suction filtration, a large amount of washing of filter cake, obtains light red solid, by ethyl acetate: sherwood oil=7:1(volume ratio) recrystallization, obtain white solid 20.7g, yield 80.1%, MP=115 ~ 117 DEG C.

Embodiment of the method 23

N-[1 '-bromo-3 '-(2 ' ', 4 ' '-dichloro-phenoxy) propyl group oxygen base] benzamide

Benzyl hydroximic acid (11.7g, 0.085mol), sodium hydroxide (3.2g, 0.08mol, be dissolved in 10ml water), the bromo-3-(2 ' of 1-, 4 ’ – dichloro-phenoxy) propane (24g, 0.085mol), 95% ethanol 150ml be mixed in round-bottomed bottle, reflux, stopped reaction when TLC monitoring raw material consumption is complete, be placed as room temperature, concentrated, suction filtration, the a large amount of washing of filter cake, obtain white solid, by ethyl acetate: sherwood oil=7:1(volume ratio) recrystallization, obtain white solid 18.9g, yield 65.5%, MP=120 ~ 122 DEG C.

Embodiment of the method 24

O-[3 '-(4 ' '-bromo-phenoxy group) propyl group oxygen base] hydroxylamine hydrochloride

N-[1 '-bromo-3 '-(4 ' '-bromo-phenoxy group) propyl group oxygen base] benzamide (7g, 0.02mol), concentrated hydrochloric acid (4g, 0.04mol) be dissolved in methyl alcohol 100ml, reflux, stopped reaction when TLC monitoring raw material consumption is complete, concentrated, obtain white solid, by ethyl acetate: sherwood oil=1:2(volume ratio) recrystallization, obtain white crystal 4.6g, yield 81.4%, MP=138 ~ 140 DEG C.

Formula VII compound all can according to said method prepare.

Embodiment of the method 25

O-[3 '-(4 ' '-chloro-phenoxy group) propyl group oxygen base] hydroxylamine hydrochloride

N-[1 '-bromo-3 '-(4 ' '-chloro-phenoxy group) propyl group oxygen base] benzamide (6.6g, 0.02mol), concentrated hydrochloric acid (4g, 0.04mol) be dissolved in methyl alcohol 100ml, reflux, stopped reaction when TLC monitoring raw material consumption is complete, concentrated, obtain white solid, by ethyl acetate: sherwood oil=1:2(volume ratio) recrystallization, obtain white crystal 4.2g, yield 90.5%, MP=132 ~ 135 DEG C.

Embodiment of the method 26

O-[3 '-(2 ' ', 4 ' '-dichloro-phenoxy) propyl group oxygen base] hydroxylamine hydrochloride

N-[1 '-bromo-3 '-(2 ' ', 4 ' '-dichloro-phenoxy) propyl group oxygen base] benzamide (6.8g, 0.02mol), concentrated hydrochloric acid (4g, 0.04mol) be dissolved in methyl alcohol 100ml, reflux, stopped reaction when TLC monitoring raw material consumption is complete, concentrated, obtain white solid, by ethyl acetate: sherwood oil=1:2(volume ratio) recrystallization, obtain white crystal 4.8g, yield 88.1%, MP=140 ~ 141 DEG C.

Embodiment of the method 27

2,4-diamino-5-benzyloxy-1,3,5-tri-azepine-9-thia-spiral shell [5.5] undecane-1,3-bis-pinene hyhrochloride (B13)

Benzyloxy semicarbazide hydrochloride (0.32g, 0.002mol), Dicyanodiamide (0.168g, 0.002mol), 15ml ethanol are mixed in round-bottomed bottle, and reflux 2 hours, is placed into room temperature.Adding tetrahydric thiapyran-4-ketone (0.9g, 0.008mol, 4 times of equivalents), concentrated hydrochloric acid (0.2g, 0.002mol), there is white precipitate in stirring at room temperature 5 days.Suction filtration, gained white solid ethanol: water=10:1(volume ratio) recrystallization, obtain white crystal 0.22g, yield 32.8%.The physico-chemical property of B13, mass spectrum ESI data in table 1, B13's ¹hNMR data are in table 2.

Embodiment of the method 28

2,4-diamino-5-(3 '-(4 ' '-bromo-phenoxy group)-propoxy-)-1,3,5-tri-azepine-9-thia-spiral shell [5.5] undecanes-1,3-bis-pinene hyhrochloride (B1)

O-[3 '-(4 ' '-bromo-phenoxy group) propyl group oxygen base] hydroxylamine hydrochloride (0.56g, 0.002mol), Dicyanodiamide (0.168g, 0.002mol), 15ml ethanol is mixed in round-bottomed bottle, and reflux 2 hours, is placed into room temperature.Adding tetrahydric thiapyran-4-ketone (0.22g, 0.002mol, 1 times of equivalent), concentrated hydrochloric acid (0.2g, 0.002mol), there is white precipitate in stirring at room temperature 5 days.Suction filtration, gained white solid ethanol: water=10:1(volume ratio) recrystallization, obtain white crystal 0.2g, yield 21.5%, HPLC purity 99.6%

The synthetic method of B2 ~ B12 is identical with B1, the physico-chemical property of B1, mass spectrum ESI data in table 1, B1's ¹hNMR data are in table 2.

Embodiment of the method 29

2,4-diamino-5-(3 '-(4 ' '-chloro-phenoxy group)-propoxy-)-1,3,5-tri-azepine-9-thia-spiral shell [5.5] undecanes-1,3-bis-pinene hyhrochloride (B2)

O-[3 '-(4 ' '-chloro-phenoxy group) propyl group oxygen base] hydroxylamine hydrochloride (0.5g, 0.002mol), Dicyanodiamide (0.168g, 0.002mol), 15ml ethanol is mixed in round-bottomed bottle, and reflux 2 hours, is placed into room temperature.Adding tetrahydric thiapyran-4-ketone (0.9g, 0.008mol, 4 times of equivalents), concentrated hydrochloric acid (0.3g, 0.003mol), there is white precipitate in stirring at room temperature 5 days.Suction filtration, gained white solid ethanol: water=10:1(volume ratio) recrystallization, obtain white crystal 0.25g, yield 27.0%, HPLC purity 99.7%.

Embodiment of the method 30

2,4-diamino-5-(3 '-(2 ' ', 4 ' '-dichloro-phenoxy)-propoxy-)-1,3,5-tri-azepine-9-thia-spiral shell [5.5] undecanes-1,3-bis-pinene hyhrochloride (B3)

O-[3 '-(2 ' ', 4 ' '-dichloro-phenoxy) propyl group oxygen base] hydroxylamine hydrochloride (0.62g, 0.002mol), Dicyanodiamide (0.168g, 0.002mol), 15ml ethanol is mixed in round-bottomed bottle, reflux 2 hours, is placed into room temperature.Adding tetrahydric thiapyran-4-ketone (1.3g, 0.012mol, 6 times of equivalents), concentrated hydrochloric acid (0.3g, 0.003mol), there is white precipitate in stirring at room temperature 5 days.Suction filtration, gained white solid ethanol: water=10:1(volume ratio) recrystallization, obtain white crystal 0.3g, yield 32.5%, HPLC purity 99.8%

Table 1,

2,4-diamino-5-replaces the physico-chemical property of-1,3,5-tri-azepine-9-oxa-or thia-spiral shell [5.5] undecane-1,3-diolefinic compounds

In table 1, " Recry " refers to recrystallization, EtOH:H in table 1 ₂the volume ratio of O is 10:1.

Table 2,

2,4-diamino-5-replaces-1,3,5-tri-azepine-9-oxa-or thia-spiral shell [5.5] undecane-1,3-diolefinic compounds ¹hNMR data

Effect example 1

To the inhibit activities determination experiment of people's Tetrahydrofolate dehydrogenase (hDHFR)

Experimental technique:

All enzymes and substrate are all bought from Sigma-Aldrich.All tests use the phosphoric acid buffer of pH=7.4.The NADPH aqueous solution of configured in advance 2M and with 0 DEG C at preserve.Test is at room temperature carried out, and uses HITACHIU-1900 spectrophotometer to carry out the absorption detecting at 340nm place.

The inhibiting rate of all samples under each concentration measures all according to following flow process: in 921 μ L phosphoric acid buffers (pH=7.4), add the DHFR solution (0.109U/ml is in the phosphoric acid buffer of pH=7.4) of sample 13.8 μ L successively, the testing sample of 10 μ L (is dissolved in DMSO; Blank group is 10 μ LDMSO), the DHF(2nM of 25 μ L is in the 2 mercapto ethanol solution of 0.25M), the NADPH solution (2nM is in the phosphoric acid buffer of pH=7.4) of 30 μ L.After jog mixing, every 30 seconds records 350nm place absorbed, until 6 minutes.M-absorbancy relation curve when making according to the data of record, the slope of 0.5-6min section is the speed of enzymatic reaction.All tests all do 3 groups, IC ₅₀get the mean value of 3 times.

It is preferably active to the vitro inhibition of people's Tetrahydrofolate dehydrogenase that result shows that compound of the present invention has, its inhibit activities close to or be better than positive control MTX(methotrexate), experimental result in table 3, wherein IC ₅₀-Ave refers to 3 mean values tested.

The inhibit activities experiment of table 3, people DHFR

Effect example 2

Sample segment carries out extracorporeal anti-tumor function test

The cell strain chosen: HCT116(human colon cancer cell), HL-60(human leukemia cell), A549(human lung adenocarcinoma cell).

Above cell strain is all bought from Chinese Academy of Sciences's cell bank, frozen and go down to posterity by Shanghai Institute of Pharmaceutical Industry's Pharmacology Lab.

Nutrient solution: RPMI1640+15%NBS+ is dual anti-.

Full-automatic microplate reader model: WellscanMK-2, production firm: Labsystems

Test method:

1) sample preparation: by compound DMSO(Merck of the present invention) dissolve after, add PBS (-) (phosphate buffered saline buffer containing the pH7.4 of 0.05% tween 20) and be made into the solution of 1000 μ g/ml or uniform suspension, then dilute with the PBS (-) containing DMSO.

2) mtt assay.The 96 every holes of orifice plate add the cell suspension 100 μ l that concentration is 4 ~ 5 × 104/ml, put 37 DEG C, 5%CO ₂in incubator.After 24h, add sample liquid, 10 μ l/ holes, if duplicate hole, 37 DEG C, 5%CO ₂effect 72h.Every hole adds the MTT(tetrazolium bromide of 5mg/ml) solution 20l, add lysate after effect 4h, 100 μ l/ holes, put in incubator, survey 570nmOD value after dissolving by the full-automatic microplate reader of MK-2.

Result shows that compound of the present invention has the activity of good extracorporeal anti-tumor cell, and test-results is in table 4.

In table 4, general formula I, part of compounds is to the IC of tumour cell ₅₀

Effect example 3

The determination experiment of anti-microbial activity

Pharmacological test procedures:

Bacterial classification is originated: evaluated biological activity centralab of Shanghai Institute of Pharmaceutical Industry reference culture

Medicine: first dissolve with DMSO, then be diluted to 250 μ g/ml with sterilized water, then two-fold dilution successively.

Bacterial classification: 20 strain G+ and G-bacterium are seeded in 37 DEG C of overnight incubation in meat soup respectively.

Method: agar plate dilution method.Ou Nuo multiple spot inoculation instrument is quantitative, inoculates often some 105CFU.

18 hours observationss are cultivated in 37 DEG C of incubators.Survey minimum inhibitory concentration (MIC value), bacteriostatic activity is in table 5.

The bacteriostatic activity of table 5, compound of Formula I

Claims

1. such as formula the diamino dihydrogen triazine derivative shown in I or its salt,

Wherein,

X is oxygen or sulphur;

N is 0,1,2,3 or 4;

R is phenyl, or substituted or unsubstituted phenyl oxygen base;

When n is 0, R is not substituted or unsubstituted phenyl oxygen base;

2. diamino dihydrogen triazine derivative as claimed in claim 1 or its salt, is characterized in that:

In the phenoxy group of described replacement, when described substituting group is halogen, described halogen is fluorine, chlorine or bromine;

In the phenoxy group of described replacement, described substituting group is C _{1 ~ 4}during alkoxyl group, described C _{1 ~ 4}alkoxyl group is methoxyl group, oxyethyl group or isopropoxy;

In the phenoxy group of described replacement, described substituting group is C _{1 ~ 4}during alkyl, described C _{1 ~ 4}alkyl is methyl, ethyl or the tertiary butyl;

In the phenoxy group of described replacement, described substituting group is C _{1 ~ 4}during acyl group, described C _{1 ~ 4}acyl group is ethanoyl.

3. diamino dihydrogen triazine derivative as claimed in claim 1 or its salt, is characterized in that:

Described R be phenyl, phenyl oxygen base,

4. diamino dihydrogen triazine derivative as claimed in claim 1 or its salt, is characterized in that: described such as formula the diamino dihydrogen triazine derivative shown in I or its salt,

When X=O, n=1, R is phenyl;

When X=O, n=4, R is phenyl, 4-chlorophenoxy, 2,4 dichloro benzene oxygen base or 4-bromine phenoxy group;

When X=S, n=1, R is phenyl;

5. diamino dihydrogen triazine derivative as claimed in claim 1 or its salt, is characterized in that: the described salt such as formula the diamino dihydrogen triazine derivative shown in I is hydrochloride or hydrobromate.

6. the diamino dihydrogen triazine derivative as described in any one of Claims 1 to 5 or the preparation method of its salt, it is following either method:

Wherein, Y is halogen, mesyloxy, trifluoro-methanesulfonyl oxy or tolysulfonyl oxygen base, and the definition of n, X, R is as described in any one of Claims 1 to 5;

Wherein, the definition of n, X, R is as described in any one of Claims 1 to 5.

7. the preparation method of diamino dihydrogen triazine derivative as claimed in claim 6 or its salt, it is characterized in that: in method one, the preparation method of described compound of Formula I comprises the steps:

8. the preparation method of diamino dihydrogen triazine derivative as claimed in claim 6 or its salt, is characterized in that: in method two, and described acidic conditions adopts concentrated hydrochloric acid; The mol ratio of described concentrated hydrochloric acid and compound VI II is 1:1 ~ 1.5:1; The mol ratio of described Compounds of formula II and Formula VIII compound is 1:1 ~ 6:1.

9. the preparation method of diamino dihydrogen triazine derivative as claimed in claim 6 or its salt, it is characterized in that: method two comprises the steps, I) in solvent, by Formula VII compound and Dicyanodiamide hybrid reaction, form the biguanide compound shown in general formula VIII; II) in acid condition, by step I) Formula VIII compound that obtains mixes with Compounds of formula II, carries out ring-closure reaction, obtain the compound shown in general formula I;

10. the preparation method of diamino dihydrogen triazine derivative as claimed in claim 9 or its salt, is characterized in that: method two comprises the steps, I) in acid condition, compound of formula VI is sloughed benzoyl and is obtained Formula VII compound; II) in solvent, by step I) Formula VII compound that obtains and Dicyanodiamide hybrid reaction, form the biguanide compound shown in general formula VIII; III) in acid condition, by step II) Formula VIII compound that obtains mixes with Compounds of formula II, carries out ring-closure reaction, obtain the compound shown in general formula I;

The preparation method of 11. diamino dihydrogen triazine derivatives as claimed in claim 10 or its salt, is characterized in that: method two comprises the steps, I) benzyl hydroximic acid and compounds of formula V be obtained by reacting compound of formula VI; II) in acid condition, compound of formula VI is sloughed benzoyl and is obtained Formula VII compound; III) in solvent, by step II) Formula VII compound that obtains and Dicyanodiamide hybrid reaction, form the biguanide compound shown in general formula VIII; Iv) in acid condition, by step III) Formula VIII compound that obtains mixes with Compounds of formula II, carries out ring-closure reaction, obtain the compound shown in general formula I;

The preparation method of 12. diamino dihydrogen triazine derivatives as claimed in claim 11 or its salt, it is characterized in that: method two comprises the steps, I) in acetonitrile, Compound I X and two halogen alkane substitutes, salt of wormwood hybrid reaction, obtain compounds of formula V; II) benzyl hydroximic acid and step I) obtained compounds of formula V is obtained by reacting compound of formula VI; III) in acid condition, by step II) obtained compound of formula VI sloughs benzoyl and obtains Formula VII compound; Iv) in solvent, by step III) Formula VII compound that obtains and Dicyanodiamide hybrid reaction, form the biguanide compound shown in general formula VIII; V) in acid condition, by step I v) Formula VIII compound that obtains mixes with Compounds of formula II, carries out ring-closure reaction, obtain the compound shown in general formula I;

In compounds of formula V, Formula IX compound and compound of formula X, Y is halogen; N is 1,2,3 or 4; R is substituted or unsubstituted phenyl oxygen base;

In the phenyl oxygen base of described replacement, described in be substituted by monosubstituted or polysubstituted; Substituting group is halogen, C _{1 ~ 4}alkoxyl group, C _{1 ~ 4}alkyl, nitro, C _{1 ~ 4}one or more in acyl group and cyano group; When described be substituted by polysubstituted time, the multiple substituting groups on the phenyl oxygen base of described replacement are identical or different;

In other general formula compounds, the definition of each group is as described in any one of Claims 1 to 5.

The preparation method of 13. diamino dihydrogen triazine derivatives as claimed in claim 12 or its salt, is characterized in that:

In compounds of formula V, Formula IX compound and compound of formula X, in the phenoxy group of described replacement, when described substituting group is halogen, described halogen is fluorine, chlorine or bromine;

The preparation method of 14. diamino dihydrogen triazine derivatives as claimed in claim 12 or its salt, is characterized in that: in compounds of formula V and Formula IX compound, described R be phenyl oxygen base,

The preparation method of 15. diamino dihydrogen triazine derivatives as claimed in claim 12 or its salt, is characterized in that: in compounds of formula V and Formula IX compound,

16. 1 kinds of pharmaceutical compositions, its contain treatment significant quantity as described in any one of Claims 1 to 5 such as formula the diamino dihydrogen triazine derivative shown in I and/or its salt, and pharmaceutically acceptable carrier.

17. diamino dihydrogen triazine derivatives as described in any one of Claims 1 to 5 or its salt or as claimed in claim 16 pharmaceutical composition are preparing the application in antitumor drug, resistance to bacteria infection medicine or people's dihydrofolate reductase inhibitor; Described tumour is adenocarcinoma of lung, colorectal carcinoma or leukemia.

18. apply as claimed in claim 17, it is characterized in that:

Described bacterial infection is pneumococcus infection, gamma streptococcus infection, infection of staphylococcus aureus or Staphylococcus albus infection;

Described people's dihydrofolate reductase inhibitor is the medicine treating and/or preventing autoimmune disorder.

19. apply as claimed in claim 18, it is characterized in that: described autoimmune disorder is rheumatoid arthritis or lupus erythematosus.