CN1795861A - Medication composition of containing nucleotide of 1,3- oxathipentane and preparation method - Google Patents
Medication composition of containing nucleotide of 1,3- oxathipentane and preparation method Download PDFInfo
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- CN1795861A CN1795861A CN 200410065885 CN200410065885A CN1795861A CN 1795861 A CN1795861 A CN 1795861A CN 200410065885 CN200410065885 CN 200410065885 CN 200410065885 A CN200410065885 A CN 200410065885A CN 1795861 A CN1795861 A CN 1795861A
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Abstract
An enantiomer mixture of cis-2-hydroxymethyl-5-(5'-fluorocytosine-1'-)-1, 3-oxythiapentance containing (-) cis-2-hydroxymethyl-5-(5'-fluorocytosine-1'-)- 1,3-oxythiapentane whose content is more than 50%, its pharmacologically acceptable derivative or salt, its preparing process and its application in preparing antiviral medicine for suppressing HIV and HBV are disclosed.
Description
Technical field
The present invention relates to suitable-2-methylol-5-(5 '-flucytosine-1 '-)-1, (-) of 3-oxygen thia pentane (hereinafter to be referred as FTC), (+) mixture of enantiomers, the application in the preparation antiviral drugs of its pharmaceutically acceptable derivates and its pharmaceutically acceptable salt.The invention still further relates to (-) with FTC, (+) mixture of enantiomers, its pharmaceutically acceptable derivates or its pharmaceutically acceptable salt are as pharmaceutical composition and this preparation of drug combination method of active ingredient.
Background of invention
1,3-oxygen Thiophane class nucleoside is the newfound in recent years active and active nucleoside analog of anti-hepatitis B virus (HBV) of excellent anti AIDS virus (HIV) that has, and the main flow medicine lamivudine that is used for the treatment of acquired immune deficiency syndrome (AIDS) and hepatitis B virus just belongs to this compounds.Up-to-date in this a compounds medicine that is approved for the AIDS virus resisting treatment is suitable-2R-methylol-5-(5 '-flucytosine-1 '-)-1 of commodity Emtriva by name, 3-oxygen thia pentane (hereinafter to be referred as (-) FTC), commodity are called suitable-methylol-5-(5 '-flucytosine-1 '-)-1 of Racivir in addition, and 3-oxygen thia pentane racemic modification (hereinafter to be referred as (±) FTC) is in the clinical research stage.
(±) FTC is 1: 1 mixture of (-) FTC of being shown below and (+) FTC.
United States Patent (USP) 5,210,085 disclosed have HIV (human immunodeficiency virus)-resistant activity suitable-2-methylol-5-(5 '-flucytosine-1 '-)-1, the preparation method of 3-oxygen thia pentane ((±) FTC), this patent have disclosed the HIV (human immunodeficiency virus)-resistant activity of (±) FTC and this chemical compound simultaneously in the treatment of acquired immune deficiency syndrome (AIDS) and the application in the prevention.United States Patent (USP) 5,210,085 methods that disclose can only prepare suitable-2-methylol-5-(5 '-flucytosine-1 '-)-1, the racemic mixture of 3-oxygen thia pentane, can't obtain single enantiomer, and the HIV (human immunodeficiency virus)-resistant activity of this Patent publish also is the data of racemic mixture (±) FTC.
Chinese patent 95118741.4 has disclosed to split with enzyme and has prepared suitable-2-methylol-5-(5 '-flucytosine-1 '-)-1, single enantiomer (+) FTC of 3-oxygen thia pentane and the method for (-) FTC, and find that (-) FTC has similar low cytotoxicity than (+) though FTC has stronger antiviral activity both.This patent has provided the HIV (human immunodeficiency virus)-resistant activity data of pure (-) FTC and pure (+) FTC, and the active ingredient of having emphasized to be used for pharmaceutical compositions should be pure (-) FTC or (+) FTC content less than 5% (-) FTC and (+) FTC mixture.
United States Patent (USP) 6703396 and United States Patent (USP) 6642245 have also disclosed by splitting the method that (±) FTC obtains (-) FTC and (+) FTC, point out that simultaneously (+) FTC content can be used to prepare antiviral drugs less than 5% (-) FTC and (+) FTC mixture.
United States Patent (USP) 6114343 and United States Patent (USP) 6069252 have disclosed the preparation method of (+) FTC, and find that (+) FTC also has antiviral activity preferably, and point out that (+) FTC and (-) FTC content can be used for pharmaceutical compositions less than 5% (+) FTC and (-) FTC mixture.
In all disclosed reports, though pure (-) FTC and the antiviral activity data of (+) FTC are known, but except ratio was mixture (±) FTC of (-) FTC of 1: 1 and (+) FTC, the antiviral activity when these two kinds of chemical compounds use jointly with arbitrary proportion was unknown.Because the interaction between medicine, when two kinds of antiviral compounds during common the use, their antiviral activity can add up, and is collaborative or antagonism.AAC " antimicrobial medicament and chemotherapy " 1992,2423 has reported (±) FTC, (-) FTC and (+) FTC anti-hiv activity at the different virus strain in various cell line.In the PBM cell, (±) FTC, (-) FTC and (+) FTC are at the effective concentration 50 (EC of HIV-1
50) be respectively 0.03,0.008 and 0.84 μ M, in cem cell, (±) FTC, (-) FTC and (+) FTC are at the effective concentration 50 (EC of HIV-1
50) being respectively 0.08,0.009 and 1.4 μ M, AAC " antimicrobial medicament and chemotherapy " 1992,2689 has reported (±) FTC, the effective concentration 50 (EC of (-) FTC and the anti-hepatitis virus of (+) FTC in HepG2 2.2.15 cell line
50) be respectively 0.083 ± 0.05,0.01 ± 0.005 and 0.96 ± 0.5 μ M.Analyzing all these activity datas can find, the EC of (±) FTC
50All, also be higher than (-) FTC EC far away far above with 1: 1 (-) FTC and the resulting data of the active stack of (+) FTC
50Twice, illustrate that under 1: 1 concentration conditions the antivirus action of (-) FTC and (+) C has antagonism, the existence of (+) FTC has suppressed the antiviral activity of (-) FTC.
The antiviral activity of the mixture of (-) FTC of arbitrary proportion and (+) FTC did not appear in the newspapers up to now, (-) FTC of arbitrary proportion and (+) FTC interaction in vivo were not studied yet, simultaneously, (-), the trend that the antiviral activity of (+) FTC mixture of enantiomers changes along with the change of the ratio of enantiomer in the mixture and the change of interaction pattern also are unknown.
In sum, prior art clearly emphasizes to have only pure (-) FTC, pure (+) FTC, racemic mixture (±) FTC and (+) FTC content can be used in pharmaceutical compositions less than (-) FTC of 5% and (+) FTC mixture and (-) FTC content less than 5% (-) FTC and (+) FTC mixture.Up to now, it is known having only the antiviral activity of mixture (±) FTC of the antiviral activity of pure (-) FTC and (+) FTC and (-) FTC that ratio is 1: 1 and (+) FTC, and (-) FTC and (+) FTC do not appear in the newspapers with the antiviral activity that other arbitrary proportions mix when using.Existing data show, under 1: 1 concentration conditions, the antiviral activity of (-) FTC and (+) FTC is mutual antagonism, the interaction when but (-) FTC of other arbitrary proportions and (+) FTC use jointly also yet there are no, and it also is unknown that its antivirus action is subjected to interactional influence of these two chemical compounds.
Though use pure (-) FTC as antiviral drugs clinically at present, owing to (-) FTC need prepare by the method for enzyme fractionation or chiral column chromatography, complex process, production cost is very high.And (±) though its production cost of FTC is lower, because its antiviral activity is also lower, the dosage of clinical use is very big, and the maximum dose level that uses in the clinical trial reaches 600 milligrams of every days, has increased the burden of patient body metabolic drug greatly.Because the acquired immune deficiency syndrome (AIDS) patient need take medicine throughout one's life, the treatment cycle of hepatitis patient is also very long, so that antiviral drugs need have a cost is low, and the characteristics low that drug effect is high with toxicity.Provide efficiently 1 under low-cost condition, 3-oxathiolane nucleosides class antiviral drugs still is a great challenge.
Summary of the invention
The invention provides a kind of preparation (-) FTC content and surpass 50% (-) FTC and the short-cut method cheaply of (+) FTC mixture, and be surprised to find that racemic mixture (±) FTC with respect to FTC, this mixture has more excellent antiviral activity, and its antiviral activity does not have to reduce because of the interaction of (-) FTC and (+) FTC.The present invention also provides a kind of and has surpassed (-) FTC of 50% and (+) FTC mixture as the pharmaceutical composition that is used for antiviral therapy of active ingredient and the method for preparing said pharmaceutical composition with (-) FTC content.
Those of ordinary skill in the art should know that the base on the FTC can form salt with organic acid or mineral acid, and 5 ' of FTC-OH can form derivants such as carboxylate or phosphate ester.Above-mentioned salt or derivant can change into the metabolite that antiviral activity is arranged of parent compound FTC or FTC in vivo directly or indirectly.Therefore (-) FTC content is greater than 50% (-), and the pharmaceutical salts and the medicinal derivative of (+) FTC mixture are also included within the scope of the present invention.
When utilizing antiviral active substance provided by the invention to treat, though directly administration of active substance, normally with the pharmaceutical dosage forms administration that contains effective ingredient.The present invention also provides and has thought that it is the pharmaceutical preparation of effective ingredient that (-) FTC content surpasses (-) FTC of 50% and (+) FTC mixture or its pharmaceutical salts or medicinal derivative, and the present invention simultaneously also provides the method for preparing these preparations.
Detailed Description Of The Invention:
The preparation of active ingredient:
The present invention at first provides the method for the mixture of (-) FTC of a kind of preparation (-) FTC enrichment and (+) FTC.
The structure of the simulation of FTC sugar loop section can be represented with following formula, its numbering as shown in the figure:
In this structure, 2-position and 5-position are two chiral centres, and the key of whole synthesis technique and difficult point are exactly optionally to make up this two chiral centres.Make up existing a lot of bibliographical information about the chiral centre of 5-position, and the chiral centre of 2-position to make up be the key issue of FTC stereoisomer in synthetic always.
We have adopted as next bar reaction scheme FTC mixture of enantiomers of (-) FTC of having synthesized (-) FTC or enrichment;
Each step reaction is described below:
Steps A: in aromatic hydrocarbon solvent (for example toluene), pivaloyl chloride and L-type mandelic acid are reacted the special pentyl ester of the 2-hydroxyl that generates L-type mandelic acid, and then add dimethyl sulfoxine or oxalyl chloride converts it into acyl chlorides, the acyl chlorides that obtains is with 1, and the reaction of 4-butylene glycol generates the diester (I) shown in structural formula.
Step B: the diester (I) that generates in the steps A is dissolved in the alcohols solvent, in this temperature range of room temperature, feed ozone at-78 ℃, after the diester full consumption is intact, stop to feed ozone, obtain aldehyde shown in structural formula (II) with suitable Reducing agent (for example thiourea, dimethyl sulfide) reduction ozone addition product.
Ozonization need carry out under extremely low temperature (being lower than-70 ℃) usually, but solvent that we are selected and reactant systems make reaction to carry out in this temperature range of room temperature at 0 ℃, greatly reduce requirement, can be applicable to suitability for industrialized production reaction unit.
Reaction process
Step C: the glycolaldehyde derivant (II) that obtains among the step B is reacted with TGA under Bronsted acid or lewis acidic catalysis, and reaction is chosen in the aromatic hydrocarbon solvent to be carried out, and by heating azeotropic removal of water branch.With this understanding, the reaction high productivity generate 2-position configuration shown in structural formula (III) different 1, the mixture of the diastereomer of 3-oxygen thia pentane-5-ketone.
Step D: with obtain among the step 3C 1, mixture fractional crystallization in the mixed solvent of being made up of toluene and petroleum ether of the diastereomer of 3-oxygen thia pentane-5-ketone obtains the mixture of two diastereomers of the III of the individual isomer (IIIA) of 2-L configuration or IIIA enrichment.Behind the general recrystallization for the first time, the content of IIIA can reach about 75%, and the content of IIIA can reach about 90% behind twice recrystallization, and the content of IIIA reaches a platform then, and recrystallization of every increase can improve the content 1-2% of IIIA.
Step e: with the mixture of two diastereomers of the III of the individual isomer (IIIA) of the 2-L configuration that obtains among the step 3D or IIIA enrichment with Reducing agent (preferred three tert-butoxy lithium aluminium hydride reductions) reduction; add acetic anhydride acylation again and obtain the 5-acetoxyl group-1 of corresponding 2-L configuration; two IV of 5-acetoxyl group-1,3 oxygen thia pentane (IV) enrichment of 3-oxygen thia pentane (IV) or 2-L configuration are at the mixture of 2 diastereomer.
Step F: with the 5-acetoxyl group-1 that obtains in the step e, two IV of 3-oxygen thia pentane (IV) or IV enrichment obtain the mixture of four stereoisomers of a pair of cis-trans-isomer V that the 2-position is the L configuration or the enrichment of 2-L configuration of compound in the mixture of 2 diastereomer and 5-flurocytosine condensation under the catalysis of Iodotrimethylsilane or Iodotrimethylsilane base triflate with the HMDS protection.
Step G: the mixture of four stereoisomers of a pair of cis-trans-isomer that obtains in the step e or the enrichment of 2-L configuration of compound is carried out fractional crystallization in methanol.Transisomer at room temperature crystallization is separated out, and obtains the mixture of two cis-isomers of 2-L cis-isomer (VI) or VI enrichment after removing by filter.
Step H: promptly obtain (+) FTC with obtaining removing the mandelic acid residue with the n-butylamine hydrolysis after chemical compound (VI) is dissolved in the methanol among the step 3G, the mixture of two cis-isomers of VI enrichment is removed the mandelic acid residue at n-butylamine and is promptly obtained (-) FTC of (-) FTC enrichment and the mixture of (+) FTC.
It is pointed out that 2 configuration can not change the process from step e to step H, the content of (-) FTC is the content decision of the IIIA that obtained by step D in (-) FTC that finally obtains and the mixture of (+) FTC.
Though use above-mentioned method can obtain pure (-) FTC single enantiomer, this need separate with the method for recrystallization repeatedly and obtains pure compound III A.Pure IIIA compares with preparation, the IIIA of preparation IIIA content about 90% and the non-enantiomer mixture of its 2-D configuration then will be easy many, because the minimizing of recrystallization number of times, greatly reduce the use amount of solvent, reduced the pollution that discharging brought of solvent slop, productive rate improves exponentially simultaneously.Therefore; (-) FTC of (if-) FTC enrichment and the antiviral activity that mixture had of (+) FTC and approaching with pure (-) FTC, the mixture that then utilizes (-) FTC of (-) FTC enrichment and (+) FTC as antiviral agent be economically can save and also also be favourable in environmental conservation.
Utilize the HepG2 2.2.15 cell that produces HBV, measure (-) FTC content and be respectively 100%, 95%, 90% and 85%, anti-hepatitis B virus activity with (-) FTC of 50% and (+) FTC mixture of enantiomers, we find above-mentioned (-) FTC content greater than the anti-HBV activity of 50%FTC mixture far above racemic FTC, and approach pure (-) FTC.This presentation of results (-) FTC content can be used as the antiviral therapy agent greater than the 50%FTC mixture, and its curative effect should approach pure (-) FTC and be higher than (±) FTC of racemization far away.
Though one aspect of the present invention provides preparation (-) FTC content to surpass (-) FTC of 50% and a method of (+) FTC mixture, also identical antiviral activity should be arranged but (-) FTC content that obtains with method for distinguishing surpasses (-) FTC of 50% with (+) FTC mixture, prepare application on the antiviral drugs also within the scope at this patent so (-) FTC content that obtains with method for distinguishing surpasses (-) FTC of 50% and (+) FTC mixture.
Second aspect of the present invention provides a kind of pharmaceutical composition, and this pharmaceutical composition contains (-) FTC content and surpasses 50% (-) FTC and (+) FTC mixture or its pharmaceutical salts or medicinal derivative and one or more pharmaceutically acceptable carriers.
Said pharmaceutical composition can comprise oral by any known mode administration; Muscle, subcutaneous and intravenous injection; By the oral cavity, Sublingual, nose, the modes such as topical of rectum and vagina.
Being suitable for oral pharmaceutical composition can be solid preparation such as tablet, capsule, and powder, granule also can be liquid preparation such as emulsion, suspension, solution etc.Oral tablet, capsule can contain conventional excipient such as binding agent, lubricant, disintegrating agent, filler or wetting agent.Oral solid preparation can also be made slow release formulation.
Preferred dosage form is an oral formulations, and preferred oral formulations comprises tablet and capsule.
Following examples can help to illustrate better the present invention, but these embodiment do not limit the scope of the invention.
Embodiment 1
1,4-two (2 '-pivaloyl oxygen base-L-almond acyloxy)-2-butylene (I)
L-mandelic acid 100 grams reacted 8 hours at 50 ℃ under the situation that feeds nitrogen in 200 milliliters of toluene for 100 milliliters with pivaloyl chloride.To add 10 milliliters of DMF behind the reactant liquor cool to room temperature that obtain, slow then dripping thionyl chloride.Be reflected at 35 ℃ and continue three hours concentrated oily liquids that obtain of final vacuum.This oily liquids slowly joined after with 500 milliliters of dilutions of dichloromethane be equipped with 1,4-butylene glycol 28ml, triethylamine 110ml is in the there-necked flask of DMAP0.4g and dichloromethane 1600ml, continue in the process that drips to stir, and keep reaction temperature below 10 ℃.Dropwise and in reactant liquor, add saturated potassium hydrogen carbonate aqueous solution 40ml after half an hour is stirred in the back, after continuing to stir half an hour, hydrochloric acid with 5% (1000ml) is washed 3 times, 0.1M sodium carbonate (1000ml) is washed 1 time, after water (1000ml) was washed 1 time, rotary evaporation was drained solvent and is got yellow oily liquid 180ml.
1H-NMR(CDCl
3)1.28(S，9H)，4.62(t，J＝1.6Hz，2H)，5.61(t，J＝4Hz，1H)，5.86(s，1H)，7.27-7.47(m，5H)。
Embodiment 2
2-(2 '-pivaloyl oxygen base-L-almond acyloxy) acetaldehyde (II)
In having three mouthfuls of round-bottomed flasks of device for absorbing tail gas, add 1,2000 milliliters of 4-two (2 '-pivaloyl oxygen base-L-almond acyloxy)-180 milliliters of 2-butylene (I) and methanol.Bathing the cooling reactant liquor with cryosel feeds ozone and keeps reaction temperature below 10 ℃ after-5 ℃.Feed ozone after 8 hours thin layer chromatography (silica gel plate use ethyl acetate: normal hexane=launch at 1: 2,1% sulphuric acid methanol solution colour developing) demonstration reaction raw materials disappear.After feeding nitrogen removing in 10 minutes ozone, start and stir, the speed with per minute 1 gram adds thiourea 12.6g after 5 ℃ to being cooled to reactant liquor with ice-water bath, after adding thiourea, reaction is stirred and is spent the night, and after the solids removed by filtration, filtrate adds dichloromethane 500ml after concentrating and removing methanol again under vacuum, after water 500ml washing is removed residual thiourea 2 times, drain to such an extent that yellow oily liquid 150 restrains.
1H-NMR(CDCl
3)：1.29(s，9H)，4.62(s，2H)，6.01(s，1H)，7.27-7.55(m，5H)，9.52(s，1H)。
Embodiment 3
2-(2 '-pivaloyl oxygen base-L-almond acyl-oxygen methyl)-1,3-oxygen thia pentane-5-ketone (III)
With 2-(2 '-pivaloyl oxygen base-L-almond acyloxy) acetaldehyde (II) 150 gram, TGA 35.8ml, p-methyl benzenesulfonic acid 1 gram and toluene 800ml join in one 2000 milliliters the round-bottomed flask.Load onto the water knockout drum afterreaction that has condensing tube and be heated to 120 ℃ of backflows two hours, have 18 ml waters therebetween and told with oil bath.Thin layer chromatography (silica gel plate is used ethyl acetate: normal hexane=launch at 1: 2,1% sulphuric acid methanol solution colour developing) shows that reaction is complete.To wash 3 times with 1M sodium carbonate 1000ml behind the reactant liquor cool to room temperature, rotary evaporation obtains 200 milliliters of yellow oily liquid after removing and desolvating, be heated and be dissolved in 1000 milliliters of isopropyl alcohols: in 6: 1 the mixed solvent of water, being positioned over 5 ℃ of 12 hours white crystals then separates out, filtration obtains solid 180 grams, and nuclear-magnetism hydrogen is dived and shown that this solid is 1: 1 mixture of two diastereomers.
Embodiment 4
2-L-(2 '-pivaloyl oxygen base-L-almond acyl-oxygen methyl)-1,3-oxygen thia pentane-5-ketone (IIIA)
With 2-(2 '-pivaloyl oxygen base-L-almond acyl-oxygen methyl)-1,3-oxygen thia pentane-5-ketone (III) 180 grams are dissolved in 600 milliliters of toluene at 60 ℃: in the mixed solvent of petroleum ether=2: 1, solution is positioned over 8 hours after-filtration of room temperature and collects the crystal of separating out and obtain 120 gram white solids
1H-NMR shows that the content of IIIA is about 75%.Repeated recrystallization obtains white crystal 78 grams after once under similarity condition,
1H-NMR shows that the content of IIIA is about 90%.Repeat above-mentioned recrystallization process up to
1H-NMR shows that the content of IIIA obtains 26 gram white solids after 97%.
1H-NMR(CDCl
3)：1.29(s，9H)，3.54(s，2H)，4.28-4.43(m，2H)，5.52-5.55(m，1H)，5.90(s，1H)，7.38-7.49(m，5H)。
Embodiment 5
2-L-(2 '-pivaloyl oxygen base-L-almond acyl-oxygen methyl)-5-acetoxyl group-1,3-oxygen thia pentane (IV)
Add lithium aluminium hydride 6.0g in the there-necked flask of a 2L, 228 milliliters of anhydrous tetrahydro furans drip the mixture of tert-butyl alcohol 37ml and oxolane 82ml under 0 ℃ and nitrogen protection.Rate of addition is per minute 2ml.Remove ice bath after dropwising, the temperature of question response system rises to the water-bath that again reaction unit is placed 30 degree after the room temperature, continues stirring reaction, stops to discharge bubble in reaction system.Change ice-water bath, in addition funnel, add 2-L-(2 '-pivaloyl oxygen base-L-almond acyl-oxygen methyl)-1, the mixture of 3-oxygen thia pentane-5-ketone (IIIA) 26 grams and THF75ml, with each second 1 speed add this mixture, add the back and continued stirring reaction 2 hours, thin layer chromatography (silica gel plate, ethyl acetate: normal hexane=launch at 1: 4,1% vitriolic methanol solution colour developing) shows that raw material has consumed fully.Add acetic anhydride 120ml in addition funnel, join in the reactant with the speed that each second, 2-3 dripped in ice-water bath, continue stirring reaction and spend the night, thin layer chromatography shows that reaction finishes.Add kieselguhr 100 grams, stirring reaction half an hour, mixture filters, and filtrate is with saturated potassium hydrogen carbonate (500ml) washing 3 times, behind the anhydrous sodium sulfate drying, after the solvent removed in vacuo 30 milliliters of light green oily liquids.
Embodiment 6
Suitable-2-L-(2 '-pivaloyl oxygen base-L-almond acyl-oxygen methyl)-5-(5 '-flucytosine-1 '-) 1,3-oxygen thia pentane (V)
With 5-flurocytosine 10g, hexamethyldisilane 50ml, ammonium sulfate solids a little and anhydrous 1,2-dichloroethanes 260ml reflux under nitrogen protection after about four hours white solid disappear and form settled solution.Add 2-L-(2 '-pivaloyl oxygen base-L-almond acyl-oxygen methyl)-5-acetoxyl group-1,30 milliliters of 3-oxygen thia pentanes (3-IIA) after this solution is cooled to 0 ℃.Under nitrogen protection, add 10.8 milliliters of Iodotrimethylsilanes.Reactant liquor stirred spend the night and allow the temperature of reactant liquor be raised to room temperature gradually.Obtain the brown residue of 50 gram syrupy shapes with under vacuum, removing behind 200 milliliters of washing reaction liquid of aqueous solution of sodium bisulfite to desolvate.This residue heating is dissolved in 100 ml methanol postcooling to room temperature, and placing had solid to separate out after 12 hours.After the solids removed by filtration mother solution is concentrated 1/3rd, being positioned over room temperature 12 hours has solid to separate out after again.Solids removed by filtration, mother solution is removed methanol under vacuum, and residue is positioned over-20 ℃ of freezing and crystallizings after being dissolved in 100 milliliters of ethyl acetate, filters to obtain white solid 14 grams, and yield is 43%, fusing point: 56-58 ℃, [α]
D 20+ 97.8 ° (c0.45, dichloromethane).
1H-NMR?(CDCl
3)：1.29(s，9H)，3.19(d，J＝12Hz，1H)，3.43(dd，J＝5.2，12.8Hz，1H)，4.25(ddd，J＝4，12.4，16.4Hz，2H)，5.46(br?s，NH，1H)，5.90(S，1H)，6.27(d，J＝4.8Hz，1H)，7.18(br?s，NH，1H)，7.27-7.51(m，5H)。
Embodiment 7
Suitable-2-L-5-(5 '-flucytosine-1 '-)-1,3-oxygen thia pentane (-) FTC.
With suitable-2-L-(2 '-pivaloyl oxygen base-L-almond acyl-oxygen methyl)-5-(5 '-flucytosine-1 '-) 1,3-oxygen thia pentane (V) 14 grams are dissolved in 100 methanol, add 10 milliliters of n-butylamines then, stirring at room reaction 2 hours.Solvent removed in vacuo, with the gained residue with petroleum ether after, the gained gray solid.Use recrystallizing methanol, obtain suitable-2-L-5-(5 '-flucytosine-1 '-) 1,3-oxygen thia pentane (FTC) 5.8 grams, yield is 89%.Fusing point: 184-186 ℃, optical rotation " α]
D 20-119.8 ° (c 0.98, methanol).Its enantiomeric purity of chirality HPLC assay determination is 98.5%.The HPLC analytical column is Chiral Pak AS (Dajcel chemical industries Ltd), and mobile phase is isopropyl alcohol, flow velocity 0.5ml/min, and the retention time of (-) FTC is 9.2min, the retention time of (+) FTC is 16.5min.
Embodiment 8:(-) FTC content is the mixture of (-) FTC of 91.3% and (+) FTC
The IIIA content that obtains among the embodiment 4 is about 90% mixture, 26 grams and obtains solid 5.3 with the identical preparation method described in the embodiment 5-7 and restrain optical rotation [α]
D 20-96.8 ° (c0.83, methanol), its enantiomeric purity of chirality HPLC assay determination is 91.3%.The HPLC analytical column is Chiral Pak AS (Daicel chemical industries Ltd), and mobile phase is isopropyl alcohol, flow velocity 0.5ml/min, and the retention time of (-) FTC is 9.2min, the retention time of (+) FTC is 16.5min.
Embodiment 9 (-) FTC content surpasses 50% (-) FTC and the anti-HBV activity of (+) FTC mixture
(-) FTC content is that 95%, 90% and 85 % (-) FTC and (+) FTC mixture make with embodiment 8 described methods.
The HepG2 2.2.15 cell line that produces HBV is maintained in the essential medium of minimum, under the conditions of air that contains 5% CO2 and 95% of 37 ℃ of humidities, cultivate, with the 2.2.15 cell inoculation in the culture dish of 25cm2.(-) FTC content is 95%, 90% and 85 % (-) FTC and (+) FTC mixture and pure (-) FTC and (±) FTC 3 are added in the medium greatly after inoculation, every kind of effective ingredient repeats three groups, cell was grown 12 days under the condition that medicine exists, medium changed once in per 3 days, after cultivation finishes, and the centrifugal 2000g10 of medium minute, and to add PEG (Mr, 8000) ultimate density to supernatant be 10%.Virus is by centrifugation, precipitate is suspended among the TNE of original volume 1% again, float adds 1% SDS and E.C. 3.4.21.64 0.5mg/ml, electrophoretic separation on 0.8% agarose gel then, transfer on the hybond membrane afterwards, HBV dna probe with the 32P labelling was hybridized 30 minutes, and automatic again radiography is transferred the scanning density instrument quantitative assay of the intensity of band from radiography.In repeated experiments independently, the amount of the DNA that HBV is special is similar, EC
50Be defined as suppressing HBV DNA and produce 50% concentration in medium, this value obtains with the drug level mapping by suppressing percent.The EC of institute's test agent
50List in the table one:
Table one
Effective ingredient EC50 (μ M)
(-)FTC 21.5±5.6
95％(-)FTC 18.3±6.5
90％(-)FTC 28.6±4.1
85％(-)FTC 23.8±11.3
(±)FTC 115.7±12.1
Embodiment 10
Capsule
Pharmaceutical formulation:
Active component 200mg
Microcrystalline Cellulose 25mg
Polyvinylpyrrolidone 75mg
Magnesium stearate 5mg
Each composition in the above-mentioned prescription mixed to be filled into making capsule in the hard gelatin capsule, wherein active component be (-) FTC content greater than 50% (-) FTC and the mixture of (+) FTC.
Embodiment 15
Tablet
Pharmaceutical formulation:
Active component 200mg
Microcrystalline Cellulose 75mg
Glycolic acid Starch Sodium 25mg
Magnesium stearate 5mg
Colloidal silica 2mg
Preparation method:
With active component 2.0kg, microcrystalline Cellulose 0.75kg, mix after glycolic acid Starch Sodium 0.25kg and colloidal silica 20g sieve respectively, add magnesium stearate 50g again after 30 minutes, continue to mix and use the rotary tablet machine dry-pressing to granulate after 5 minutes with suitable blender mixing.
Wherein said active component is (-) FTC content greater than 50% (-) FTC and the mixture of (+) FTC.
Claims (10)
1. suitable-2-methylol-5-(5 '-flucytosine-1 '-)-1, the mixture of enantiomers of 3-oxygen thia pentane, the application in the preparation antiviral drugs of its pharmaceutically acceptable derivates or its pharmaceutically acceptable salt, said suitable-2-methylol-5-(5 '-flucytosine-1 '-)-1, the mixture of enantiomers of 3-oxygen thia pentane be (-) suitable-2-methylol-5-(5 '-flucytosine-1 '-)-1,3-oxygen thia pentane and (+) be suitable-2-methylol-5-(5 '-flucytosine-1 '-)-1, the mixture of 3-oxygen thia pentane, it is characterized in that (-) suitable-2-methylol-5-(5 '-flucytosine-1 '-)-1, the content of 3-oxygen thia pentane is no less than 50% and no more than 95%.
2. according to claim 1, said suitable-2-methylol-5-(5 '-flucytosine-1 '-)-1, in the 3-oxygen thia pentane (-) suitable-2-methylol-5-(5 '-flucytosine-1 '-)-1, the content of 3-oxygen thia pentane is no less than 75%.
3. according to claim 1, said suitable-2-methylol-5-(5 '-flucytosine-1 '-)-1, in the 3-oxygen thia pentane (-) suitable-2-methylol-5-(5 '-flucytosine-1 '-)-1, the content of 3-oxygen thia pentane is no less than 90%.
4. according to claim 1, said suitable-2-methylol-5-(5 '-flucytosine-1 '-)-1, in the 3-oxygen thia pentane (-) suitable-2-methylol-5-(5 '-flucytosine-1 '-)-1, the content of 3-oxygen thia pentane is no less than 93%.
5. one kind with suitable-2-methylol-5-(5 '-flucytosine-1 '-)-1,3-oxygen thia pentane, its pharmaceutically acceptable derivates or its pharmaceutically acceptable salt are as the pharmaceutical composition of active ingredient, said suitable-2-methylol-5-(5 '-flucytosine-1 '-)-1,3-oxygen thia pentane be (-) suitable-2-methylol-5-(5 '-flucytosine-1 '-)-1,3-oxygen thia pentane and (+) be suitable-2-methylol-5-(5 '-flucytosine-1 '-)-1, the mixture of 3-oxygen thia pentane, it is characterized in that (-) suitable-2-methylol-5-(5 '-flucytosine-1 '-)-1, the content of 3-oxygen thia pentane is no less than 50% and no more than 95%.
6. the pharmaceutical composition described in claim 5, wherein said suitable-2-methylol-5-(5 '-flucytosine-1 '-)-1, in the 3-oxygen thia pentane (-) suitable-2-methylol-5-(5 '-flucytosine-1 '-)-1, the content of 3-oxygen thia pentane is no less than 75%.
7. the pharmaceutical composition described in claim 5, wherein said suitable-2-methylol-5-(5 '-flucytosine-1 '-)-1, in the 3-oxygen thia pentane (-) suitable-2-methylol-5-(5 '-flucytosine-1 '-)-1, the content of 3-oxygen thia pentane is no less than 90%.
8. the pharmaceutical composition described in claim 5, wherein said suitable-2-methylol-5-(5 '-flucytosine-1 '-)-1, in the 3-oxygen thia pentane (-) suitable-2-methylol-5-(5 '-flucytosine-1 '-)-1, the content of 3-oxygen thia pentane is no less than 93%.
9. the pharmaceutical composition described in claim 5, described pharmaceutical composition contains suitable-2-methylol-5-(5 '-flucytosine-1 '-)-1,3-oxygen thia pentane is at pharmaceutically acceptable salt.
10.-plant prepare claim 5-9 and appoint-described in the method for pharmaceutical composition, this method comprises the suitable-2-methylol-5-(5 '-flucytosine-1 '-)-1 as active ingredient, 3-oxygen thia pentane, its pharmaceutically acceptable derivates or its pharmaceutically acceptable salt mix with one or more pharmaceutically acceptable carriers.
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| Publication number | Priority date | Publication date | Assignee | Title |
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| WO2019176732A1 (en) * | 2018-03-16 | 2019-09-19 | 国立大学法人東京農工大学 | Production method for tetrahydroisoquinoline ring-containing compound |
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| WO2019176732A1 (en) * | 2018-03-16 | 2019-09-19 | 国立大学法人東京農工大学 | Production method for tetrahydroisoquinoline ring-containing compound |
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