CN101622265B - Antiviral compounds - Google Patents
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- CN101622265B CN101622265B CN200880006407.8A CN200880006407A CN101622265B CN 101622265 B CN101622265 B CN 101622265B CN 200880006407 A CN200880006407 A CN 200880006407A CN 101622265 B CN101622265 B CN 101622265B
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- 0 CC(C1)(C2)C1C(*)*C2(*)C(O)=O Chemical compound CC(C1)(C2)C1C(*)*C2(*)C(O)=O 0.000 description 2
- QNEYULQCJWDXDP-UHFFFAOYSA-N CC1CN(CCN(C)C)CC1 Chemical compound CC1CN(CCN(C)C)CC1 QNEYULQCJWDXDP-UHFFFAOYSA-N 0.000 description 1
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Abstract
The invention provides a new class of compounds exhibiting an inhibitory effect on neuraminidase (NA), hemagglutinin (HA) and structural M2 protein bearing viruses. These compounds are also useful as inhibitors of the replication factors of hepatitis virus type C (HVC). The invention also describes pharmaceutical compositions containing the compounds of the invention either alone or in combination admixed with a suitable and pharmaceutically acceptable carrier.
Description
Technical field
The present invention relates to as neuraminidase (NA), erythrocyte agglutination element (HA), structural protein M
2the novel cpd of the inhibitor of (HEF) glycoprotein is merged with hemaglutinin esterase; Comprise the pharmaceutical composition of the described compound being used for the treatment of, preventing or improving virus infection; And use the method for described compound and composition.
Spheroid form and/or filament shape (Yoshinori Fujiyoshi et al. " Finestructure if influenza A virus observed by electroncryo-microscopy " that diameter is 80 to 120nm is demonstrated when having been found that influenza virus is observed under an electron microscope, The EMBO Journal 13 (2), p 318-26,1994).The most characteristic feature of viromembrane is, when there is radial ridge (radialprojections) [(Wilson I.A.et al. " Structure of thehaemoagglutinin membrane glycoprotein of influenza virus at corresponding to when the A type of erythrocyte agglutination element (HA) and neuraminidase (NA) and Type B virus
resolution ", Nature, 289, p 366-73,1981, Varghese J.N.et al. " Structure of the influenza virus glycoprotein antigenneuraminidase at
resolution ", Nature, 303, p 35-40,1983, Colman P.M.et al. " Structure of the catalytic and antigenicsites in influenza virus neuraminidase ", Nature, 303, p41-44, 1983)], and when corresponding to being called that hemaglutinin esterase produces three kinds of biologic activity when merging the C type influenza virus of (HEF) glycoprotein: be attached to acceptor (H), receptor deactivation (E) and fusion (F) (Herrier Georg, et al., " A synthetic sialic acid analogueis recognized by influenza Cvirus as a receptor but is resistantto the receptor-destroying enzyme ", J.Biol.Chem., 2567 (8), p 12501-12505, 1992).
First-generation anti-virus product (mainly adamantane derivative, similar amantadine, Rimantadine etc.) it is believed that the M by blocking influenza A
2-protein ion channel and working.Block H
+ion passes through M
2the inflow of-protein ion channel can suppress shelling and the release of free ribonucleoprotein in tenuigenin.This only finds in A C-type virus C, but does not occur in Type B virus.
After this, viral resistant strategies relates to the exploitation of s-generation antiviral agent such as zanamivir and oseltamivir, they suppress erythrocyte agglutination element (HA) or neuraminidase (NA), and it is present in the surface of A type and Type B influenza virus with mushroom ridge.These albumen are attached to the film surface of the target cell that will infect by the sialic acid moities of cracking sialoglycoprotein and glycolipid class.In addition, at the terminal of virus replication, neuraminidase is for making virus release be required from cracking sialic acid on acceptor.
By contrast, the strategy of current suppression hepatitis C virus (HVC) is included in the synthesis of monophosphate guanosine-and uses ribavirin (for monophosphate) to suppress by reducing Intracellular levels.In addition, ribavirin (for triphosphate) suppresses ARNm-guanilyltranspherase (guanilyltranspherase) by the synthesis reducing viral ARNm and ARN polysaccharase.
Much more more and more international application reports resistance strains of influenza viruses, and lasting sudden change (particularly A C-type virus C) and drug resistant variants or its combination are transmissible and are pathogenic completely.In this respect, in recent years, A type bird flu (H5N1) virus is described as the material risk that world pop disease is broken out by scientists.
Similarly, hepatitis C infection is very general in the whole world, therefore also represent a kind of serious pathogenic situation affecting patient numbers and increase.
Therefore being badly in need of the antiviral compound developing improvement, preferably there is the Multiple Combination mechanism to virus replication effect in it.
Background technology
Itzstein, M.von et al.; " Nature ", discloses the appropriate design of the inhibitors of influenza viruses replication based on sialidase in 363 (6428), p 418-423 (1993).Colman, P.M.et al.; WO 92/06691 (PCT/AU90/00501, publication date: on April 30th, 1992), Itzstein, L.M.von et al.; EP 0539204A1 (European application 92309684.6, publication date: on April 28th, 1993), and Itzstein, L.M.von et al.; Disclose the compound in conjunction with neuraminidase in WO 91/16320 (PCT/AU91/00161, publication date: on October 31st, 1991), and it is believed that and demonstrate interior resisting virus activity.Bischofberger N.W.et al.; US 5,952,375 (U. S. application 08/606,624, the applying date: on February 26th, 1996) discloses the new compound as neuraminidase inhibitor.
Babu Y.S., Chad P., Bantia S.et al.: " Discovery of a novel; highly potent; orally active; and selective influenzaneuraminidase inhibitor through structure-based drug design " .J Med Chem, 43 (19): 3482 (2000), international application WO 99/33781 (international application no PCT/US 98/26871, publication date: on July 8th, 1999) discloses new substituted compound as neuraminidase inhibitor and derivative.
In addition, infect to treat chronic HVC, being such as described in Martindale 33.rd Ed. (2002) p 639-43, common medical practice is combined administration purine nucleoside analogs such as ribavirin or Viramidine (trade(brand)name) and cytokine such as Interferon Alpha-2b (or glycol interferon alpha-2b) simultaneously in people experimenter.In fact, it is believed that, monophosphate ribavirin and this analog derivative suppress to concentrate in the synthesis of monophosphate bird (purine) nucleosides and cell, and triphosphate RNA interfering m-guanilyltranspherase.
Goal of the invention
An object of the present invention is to provide virus particularly infected by influenza and the effective inhibiting new compound of hepatitis virus display.This compounds is to the M in membranin, erythrocyte agglutination element (HA), structural protein such as virus
2, and glycolytic enzyme such as neuraminidase (NA) produce its combination and optionally restraining effect, work more particularly by disturbing with Viral nerve ammonia neuraminidase.These new compounds also produce inhibit activities to hepatitis C virus (HVC).That another object is to provide improvement and cheaper and cause the most commonly encountered diseases poison transmittance process inhibitor of severe viral infection, and with the antiviral agent used at present without any cross resistance.Further object is to provide the method for the reasonable combination using the known antiviral agent of new compound of the present invention or they and other.Another object is to provide the pharmaceutical composition for above object.
Consider according to the present invention's entirety to those skilled in the art, the object of these and other will become obvious.
Summary of the invention
In a first aspect of the present invention, there is provided herein general formula (I) compound:
Wherein:
X is-CH
2-,-O-,-CHF-,-CF
2-;
The C of singly-bound or double bond shack
2-C
3;
R
1represent-OH, halogen or-B/ part, condition is the C that there is double bond shack
2-C
3time R
1do not exist; And
R
2represent-OH ,-O-CH (C
2h
5)
2,-NH
2,-NHC (NH) NH
2or-NH-OH; And
R
3-NH
2,-NHCO-CH
3or-NH-CO-CH
2-OH; And
R
4expression-CHOH-CHOH-CH
2-OH ,-CHOH-CH
2-B/ part ,-CH
2-B/ part or-B/ part
Wherein-B/ part represents:
-B1/ part-B2/ part
Wherein:
R
5represent and connect functional group-NH-,-CH
2-NH-,-CH (CH
3)-NH-,-NH-CH (CH
3)-NH-,-C (CH
3)
2-CH
2-NH-,-NH-CO-CH
2-O-CH
2-CH
2-NH-or
and
R
6-H ,-CH
3or-C
2h
5; And
R
7-H ,-CH
3or-C
2h
5; And
R
8represent and connect functional group-NH-,-CO-NH-or-C (NH)-NH-;
And their C1-4 carboxyl list or polyester, additive salt, solvate, the enantiomorph of fractionation and the diastereomer of purifying.
The present invention also comprises pharmaceutical composition, and it is being applicable to contain independent the compounds of this invention in the Mammals pharmaceutically acceptable carrier that particularly people uses or combine with other promoting agent.
In another embodiment of the present invention, neuraminidase and/or albumen M
2the activity method that can be comprised the following steps suppress: suspect containing neuraminidase and/or albumen M by the compounds of this invention or compositions-treated
2sample.
The present invention provides on the other hand for treatment in host or the method for preventing the virus infection such as caused by influenza virus or hepatitis virus, and the method comprises by any suitable route of administration to the compounds of this invention disclosed herein of host's administering therapeutic effective dose.
In other embodiment of the present invention, additionally provide the novel method of synthesis the compounds of this invention.
Embodiment
The present invention relates to structural formula (I) compound of following configuration:
Wherein:
X is-CH
2-,-O-,-CHF-,-CF
2-;
The C of singly-bound or double bond shack
2-C
3; And
R
1represent-OH, halogen or-B/ part, condition is the C that there is double bond shack
2-C
3time R
1do not exist; And
R
2-OH ,-O-CH (C
2h
5)
2,-NH
2,-NHC (NH) NH
2or-NH-OH; And
R
3-NH
2,-NHCO-CH
3or-NH-CO-CH
2-OH; And
R
4expression-CHOH-CHOH-CH
2-OH ,-CHOH-CH
2-B/ part ,-CH
2-B/ part or-B/ part
Wherein-B/ part represents:
-B1/ part-B2/ part
Wherein:
R
5represent and connect functional group-NH-,-CH
2-NH-,-CH (CH
3)-NH-,-NH-CH (CH
3)-NH-,-C (CH
3)
2-CH
2-NH-,-NH-CO-CH
2-O-CH
2-CH
2-NH-or
and
R
6-H ,-CH
3or-C
2h
5; And
R
7-H ,-CH
3or-C
2h
5; And
R
8represent and connect functional group-NH-,-CO-NH-or-C (NH)-NH-.
In preferred embodiments, X is represented by-O-, and it is typical sialic acid ring, wherein at C
1on carboxylic acid keep do not replace, because it is considered to the sorption site of Viral nerve ammonia neuraminidase.In another preferred embodiment, the C of singly-bound or double bond shack
2-C
3, condition is, when there is double bond, and R
1do not exist.In another typical embodiment, at least R
1or R
4can be mono-substituted by-B/ part.Therefore, R
1preferred expression-OH, halogen or typical-B/ part, work as R
4expression-CHOH-CHOH-CH
2-OH or typical-CHOH-CH
2during-B/ part, should-B/ part can be wherein-B1/ part or-B2/ part.
When using typical-B1/ part, the specific inhibitory mechanisms of A and B strains of influenza viruses to neuraminidase is preferably induced, and M
2the blocking-up of-protein ion channel (existing only in A strain) is also allowed.-B2/ part also can be used for monosubstituted derivative, to realize similar restraining effect.In the combination that another is suitable, R
1and R
4can be identical or different dibasic.In preferred combination, R
1and R
4can all be replaced by-B2-part, gained compound it is believed that and more optionally suppresses HVC to copy.
Work as R
1replaced and R by-B1/ part or-B2/ part
4when being differently expressed as-B2/ part or-B1/ part, it is believed that gained restraining effect can affect A and B strains of influenza viruses, and can HVC be affected.
In preferably combining further, work as R
1or R
4be connected to-B1/ the part of main structure (I) when replacing, R simultaneously
2can be-OH and R
3can be-NHCO-CH
3.Another preferred embodiment connects functional group R
5can be-NH-or-CH (CH
3)-NH-, and R
6and R
7-H or-CH can be preferably
3.Work as R
1or R
4by be connected to main structure (I)-B2/ part represent time, connect functional group R8 can be preferably-CO-NH-.
The present invention also comprises the straight or branched C of the compounds of this invention
1-4carboxyl monoesters or polyester, additive salt, solvate, the enantiomorph of fractionation and the diastereomer of purifying.
In another embodiment of the present invention, neuraminidase and/or M
2the activity of ionic channel can by suspecting containing neuraminidase and/or M by the compounds of this invention or compositions-treated
2the viral sample of albumen suppresses or blocks.
Another aspect of the present invention to relate in mammalian hosts the method for the treatment of or prophylaxis of viral infections particularly influenza A and B virus strain or HVC, and it comprises by any suitable route of administration to the compounds of this invention of host's administering therapeutic effective dose.
In another embodiment, the novel method of synthesis the compounds of this invention is additionally provided.
The present invention also comprises pharmaceutical composition, and it is being applicable to containing independent the compounds of this invention in such as to the pharmaceutically acceptable carrier of administration, or with the combination of such as another compound of the present invention or one or more promoting agents.
The further aspect of the present invention comprises treatment or prevents the method for described virus disease or illness, the method is mammiferous treatment by combinationally using the compounds of this invention or pharmaceutical composition or its mixture and simultaneously or be used alternatingly the treatment of promoting agent of another treatment effective dose, and described promoting agent also can suppress this type of virus infection.
The present invention also comprises the method producing the compounds of this invention.Described compound is by applicable technology preparation any in organic synthesis, and they well known to a person skilled in the art.
For R
1be directly connected to the C of sialic acid ring
2(X=-O-) the generality preparation of preferred compound of the present invention, can preferred reductive amidation methodology, as by Tang et al.Biochem.Biophys.Res.Commun., the method that 132:474-80 (1985) describes in early days, and by Stoll et al., Biochem.J., 256:661-4 (1988) and by ScharzmannG.et al., Biochem., the method revised of 22:5041-9 (1983).In fact, by according to above method, the C of sialic acid ring can be produced
2and the covalency amine between the amino group of-B/ part is connected.
Similarly, for other preferred compound of the present invention (such as R
4be directly connected to the C of sialic acid ring
6(X=-O-) those compounds) general preparation method, the many known technologies described can be followed in following publication: Compendium of Organic SyntheticMethods (John Wiley & Sons, New York), Vol.1, Ian T; Harrisonand Shuye, Harrison, 1971; Vol.2, Ian T.Harrison and ShuyenHarrison, 1974; Vol.3, Louis S.Hegedus and Leroy Wade, 1977; Vol.4, Leroy G.Wade, jr., 1980; Vol.5, Leroy G.Wade, Jr.1984; And Vol.6, Michael B.Smith; And March, J., 2AdvancedOrganic Chemistry, 3rd edition, (John Wiley & Sons, New York, 1985), Comprehensive Organic Synthesis.Selectivity, Strategy & Efficacy in modern Organic Chemistry.In 9 Volumes, Barry M.Trost, Editor-in-Chief (Pergamon Press, New York, printing in 1993).
Provided below is many exemplary methods preparing the compounds of this invention, but they can not the scope of restricted application method.
Usually, reaction conditions such as temperature, reaction times, solvent, treatment process etc. are for the concrete reaction carried out being common those in this area.The detailed description of above-mentioned condition is comprised in the document material quoted and the material wherein quoted.The exemplarily method of property, below general step R used in the present invention
4the generality synthesis of the compound replaced.
But, it will be understood to those of skill in the art that other standard operation can be used for producing identical material.
The preparation of the intermediate 1 of the following structure of step 1-:
intermediate 1
Under reflux, in methyl alcohol, Dowex 50 (H is used
+) stir process sialic acid reaches time of 24-48 hour.Leach resin, filtrate is concentrated into drying, (intermediate 1) that the dimethyl obtaining needing replaces.
The preparation of the intermediate 2 of the following structure of step 2-:
intermediate 2
The solid aqueous sodium hydroxide solution obtained from step 1 restores, and at room temperature stirs, and usually continues 1-3 hour.Then by this mixture Dowex 50 (H
+) resin moderated to pH7.0-7.5.Filter, then make filtrate lyophilize, obtain corresponding non-methylated ester (intermediate 2).
The preparation of the intermediate 3 and 4 of the following structure of step 3-:
intermediate 3
intermediate 4
Make the intermediate obtained from step 2 in the dark react 1 hour from the sodium periodate aqueous solution with different mol ratios, produce intermediate 3 (lower ratio) or intermediate 4 (higher ratio).Then barium acetate is added in throw out, then passed through to filter excessive iodate and periodate.Then by filtrate lyophilize.Obtain micro-yellow solid (intermediate 3 or 4).
The preparation of the intermediate 5 and 6 of the following structure of step 4-:
intermediate 5
intermediate 6
Under agitation the intermediate 3 or 4 from step 3 gained is dissolved in aqueous formic acid respectively, then heats 1 hour at 80 DEG C.By the lyophilize separately of gained solution.Obtain intermediate 6 and 5 respectively.
Step 5-prepares final R according to following exemplary reaction scheme
4mono-substituted compound
Option A
Intermediate 5 produces the reaction product of-B1/ part
Compound 1
R of the present invention
4mono-substituted (-B1/ part) compound
Option b
Intermediate 6 produces the reaction product of-B2/ part
Compound 2
Another R of the present invention
4mono-substituted (-B2/ part) compound
Used by each intermediate obtained 5 or 6 from step 4 reaction product of-B/ part needed for distilled water and selected generation (-B1/ part or-B2/ part) to restore respectively, make reaction mixture keep hold over night at 4 DEG C.Then add sodium borohydride, and make reaction mixture at room temperature keep leave standstill reach 1 hour.Make each sample at Dowex 50 (H
+) deionization on post, then by elutriant lyophilize, obtain different finalization compounds of the present invention respectively.
Other typical R of the present invention
4mono-substituted (having different-B/ part) compound is such as expressed as follows, and describes in a further embodiment:
Compound 3
Compound 4
Compound 5
Compound 6
Compound 7
Compound 8
Compound 9
Compound 10
Another is preferably prepared embodiment and comprises by using following reactions steps to synthesize R
1mono-substituted compound:
Step 1-R
1the preparation of mono-substituted (-B/ part) compound
Sialic acid is dissolved in the mixture of the first alcohol and water of different ratios (preferably 9: 1), then under gentle agitation, adds the reaction product producing-B/ part.After adding sodium borohydride, this mixture is kept 2 hours under 60 DEG C and gentle agitation.Then make this mixture by Dowex50 (H
+) post, become boric acid to make reforming sodium borohydride.By elutriant lyophilize, restore with the methyl alcohol of aliquot, then pass through paper filter.Isolate resistates, make elutriant dry.Repeatedly operation below for several times (usually more than 5 times).Gained solid is finally dissolved in aliquot water, then lyophilize, obtains the R of the present invention needed
1mono-substituted (there is-B/ part) compound.
Other typical reaction scheme can be expressed as follows:
Scheme C
Sialic acid (intermediate) produces the reaction product of-B1/ part
R of the present invention
1mono-substituted (-B1/ part) compound (compound 11)
Scheme D
Sialic acid produces the reaction product of-B2/ part
Another R of the present invention
1mono-substituted (-B2/ part) compound
Another other preferably prepare embodiment and comprise by adopting following general reaction scheme synthesis R
1and R
4dibasic compound:
Step 1-R
1and R
4the preparation of dibasic (-B/ part) compound
By in conjunction with life side described above scheme (A, B, C and D), those skilled in the art can obtain disubstituted compound of the present invention.
Typical finally disubstituted (the having identical or different-B/ part) compound of the present invention is expressed as follows:
Typical R
1and R
4disubstituted (there is identical-B1/ part) compound
Another typical R
1and R
4disubstituted (there is identical-B2/ part) compound
It will be obvious to those skilled in the art that it provides the R as general formula (I) represents separately by using identical generation-B1/ part or the reaction product of-B2/ part
6, R
7and R
8other possible combination, this may obtain similar bisubstituted compound of the present invention.
The typical R of the present invention
1and R
4disubstituted (there is-B/ part of various combination) compound
Another typical R of the present invention
1and R
4disubstituted (there is-B/ part of various combination) compound
Similarly, by using the differential responses product of combination-B1/ part and-B2/ part, it provides the R as general formula (I) represents separately
6, R
7and R
8other possible combination, this may obtain the different bisubstituted compound of similar the present invention.
Be apparent that to those skilled in the art, by using classical Resorcinol-HC l method (Svennerholm L. " Quantitative estimation of sialic acids.II.A colorimetric resorcinol-hydrochloric acid method. " Biochem.Biophys.Acta, 24 (3),: 604-611,1957; Miettinen J.etal. " Use of butyl acetate in determination of sialic acid. ", Acta Chem.Scand., 13,856-858,1959) and TBA (2-thiobarbituricacidα-) method (Warren L. " The thiobarbituric acid assay of sialic acids. ", J.Biol.Chem., 234,1971-5,1959; Aminoff D. " Methods for thequantitative estimation of N-acetylneuraminic acid and theirapplication to hydrolysates of sialomucoids. ", Biochem.J., 81 (2), 384-392,1961) detect different because of following reason with quantifying sialic acid: Resorcinol-HCl method to be dissociated existence and all can detecting and quantitative assay with during other sugar such as sialic acid sugar compounds (sialoglycocompounds) conjugation at sialic acid and derivative thereof.By contrast, TBA method is only being connected to 2 (C
2) the hydroxyl-partly (R of carbon atom
1=-OH) be not substituted Shi Caike and detect and quantifying sialic is sour and derivative.
That the compounds of this invention is also included in the enrichment at any or all of asymmetric atom place or split optical isomer.Racemic mixture and non-enantiomer mixture, and isolated or synthesized, other enantiomorph essentially no to or the right single optical isomer of diastereomer, all within the scope of the present invention.By known technology racemic mixture is separated into they single, be essentially optically pure isomer, described technology such as, the diastereoisomeric salt formed as acid or alkali with optical activity auxiliary agents is separated, then transforms and be returned to optically active substance.In most of the cases, the optical isomer of needs is by synthesizing with stereospecificity reaction from the suitable steric isomer of required initial substance.
The present composition optionally comprises the salt of compound herein, and the especially acceptable non-toxic salt of pharmacy, it comprises such as inorganic or preferred organic acid or alkali.When needing the water-soluble salt of compound, salification is preferred methodology.
The present invention relates to the method suppressing neuraminidase activity on the other hand, and the method comprises suspects with the compounds of this invention process the step containing the neuraminidase such as sample of Viral nerve ammonia neuraminidase.It is believed that the compounds of this invention can be used as the inhibitor of neuraminidase, as the intermediate of this type of inhibitor, or there is other purposes described below.In fact, in the surface that described inhibitor can be attached to neuraminidase or inner chamber, these surfaces or inner chamber have geometry uniqueness for neuraminidase.But the compound in conjunction with neuraminidase can Reversible binding in various degree.The compound of these irreversible fixation is substantially the ideal candidate for the inventive method.Organism containing neuraminidase comprises bacterium (vibrio cholerae, clostridium perfringens, streptococcus pneumoniae and Arthrobacter sialophilus) and virus (especially orthomyxovirus or Paramyxo virus such as influenza virus A and B, parainfluenza virus, rhinovirus, coronavirus, coronavirus mutant and/or improvement coronavirus, mumps virus, Avian pneumo-encephalitis virus, ewcastle disease virus and Sendai virus).From the restraining effect of these organic neuraminidase activity that are that obtain any one or that find within the scope of the invention.The compounds of this invention is also used in animal or human treats or prevents above-mentioned infection, and described animal is duck, rodent or pig such as.
In further embodiment, the compounds of this invention is that the inhibit activities of routine techniques to its neuraminidase by evaluating enzymic activity screens.In the context of the present invention, first typical compound is screened for vitro inhibition neuraminidase.
The further aspect of the present invention relates to blocking-up H
+ion by the inflow of M2-protein ion channel, suppress shelling and free ribonucleoprotein to be discharged into cytoplasmic method, the method comprises the step suspecting the sample containing M2-albumen such as influenza virus A with the compounds of this invention process.In fact, also think that the compounds of this invention is the onset by blocking virus M2-protein function.
Another further aspect of the present invention relates to the method for the synthesis suppressing monophosphate guanosine-and RNAm-guanilyltranspherase, and the method comprises by synthesizing with the RNAm and RNA polymerase that block HVC by suspection sample with the compounds of this invention process.
In another embodiment, it is believed that disubstituted compound that the present invention is different simultaneously as the inhibitor onset of neuraminidase and M2-protein ion channel.
The compounds of this invention can be prepared with conventional carrier and vehicle, and these carriers and vehicle will be selected according to conventional practice.Tablet will containing vehicle, glidant, weighting agent, tackiness agent etc.Aqueous solution preparation is made into sterile preparation, and when needing to carry out administration by the mode except oral, they are normally isotonic.All preparations optionally will contain vehicle, such as, at known publication " Handbook of Pharmaceutical Excipients ", the 4th edition, describe in RoweR.C.et al., Pharmaceutical Press (2003) those.Vehicle comprises xitix and other oxidation inhibitor, sequestrant such as EDTA, carbohydrate such as dextrin, hydroxy alkyl cellulose, hydroxyalky methyl celluloses, stearic acid etc.
One or more compounds (being also called activeconstituents below) of the present invention can be used by any approach being suitable for illness to be treated.Suitable approach comprises mouth, rectum, nose, locally (comprise oral cavity and sublingual), vagina and parenteral (comprising in subcutaneous, intramuscular, vein, intracutaneous, sheath and epidural) etc.Be appreciated that preferred approach can change with the illness of such as experimenter.The advantage of the compounds of this invention is, they are that oral bio is effective, and can with oral Pharmaceutical dosage forms administration; Can but not must by lung or intra-nasal route give them.
Although it is possible that inventive compound is used separately, they preferably exist with pharmaceutical preparation.Invention formulation, it can for animals or people's use, comprises at least one activeconstituents of the present invention defined above or compound, and one or more acceptable carriers and other optional therapeutic component.Described carrier is " acceptable " necessarily, represents that other composition of itself and preparation is compatible and to its subject physiologic is nontoxic.
Preparation comprises those that be applicable to aforementioned route of administration.Preparation can exist expediently in a unit, and prepares by the known any method of pharmaceutical field.
Preparation technique is found in usually " Remington ' s Pharmaceutical Sciences " Mack Publishing Co., Easton, Pa., U.S.A..These class methods comprise makes activeconstituents and carrier-bound step, this carrier by a kind of or what time ancillary component form.Usually, said preparation, by making activeconstituents evenly and combining with both the solid carrier of liquid vehicle or dispersion and fining or this densely, then, if needed, makes product be configured as desired.
Be applicable to oral invention formulation and can be prepared as solid unit such as capsule, cachet or tablet, it contains the activeconstituents of predetermined amount separately; Powder or particle; Solution in waterborne liquid or non-aqueous liquid or suspension; Oil-in-water liquid emulsion or water-in-oil liquid emulsion.Activeconstituents can also exist with bolus, Electuary or paste.
Tablet is suppressed by optional and one or more ancillary components or is molded and prepares.Compressed tablets is prepared by a suitable machine activeconstituents being pressed into free-pouring form such as powder or particle, optional and tackiness agent, lubricant, inert diluents, sanitas, tensio-active agent or dispersant.Molded tablet is by preparing the mixture pressing mold of the powdered active ingredient by inert liquid diluent humidifying on a suitable machine.Tablet can optionally by dressing or indentation, and is optionally mixed with and provides activeconstituents from wherein slow releasing or Co ntrolled release.
For the infection of eye or other outside organization such as mouth and skin, said preparation can be preferably used as topical ointment or emulsifiable paste, the amount that described ointment or emulsifiable paste contain activeconstituents is that such as 0.075 to 20%w/w (comprises the activeconstituents that scope is 0.1% to 20%, increment is 0.1%w/w, such as 0.6%w/w, 0.7%w/w etc.), be preferably 0.2 to 15%w/w, most preferably be 0.5 to 10%w/w.When being mixed with ointment, activeconstituents can use with paraffin matrix or water dispersible ointment base.Or this activeconstituents can be mixed with emulsifiable paste together with Oil-in-water emulsifiable paste matrix.
If needed, the aqueous phase of emulsifiable paste matrix can comprise, such as, and at least polyvalent alcohol of 30%w/w, namely there is alcohol such as propylene glycol, 1,3 butylene glycol, N.F,USP MANNITOL, sorbyl alcohol, the glycerine and polyoxyethylene glycol (comprising PEG 400) and composition thereof of two or more hydroxyl.Topical formulations also desirably can comprise promotion activeconstituents is subject to invasion and attack region absorption or infiltration compound by skin or other.The example of this type of dermal osmosis accelerator comprises methyl-sulphoxide and analogue.The oil phase of emulsion of the present invention can be made up of in a known way principal component.Although this phase only can comprise emulsifying agent (being separately referred to as emulgent), its expect comprise at least one emulsifying agent with fat or oil or with fat and oil both mixture.Hydrophilic emulsifier preferably uses together with the lipophilic emulsifier as stablizer.Also preferably include both oil & fats., to have or emulsifying agent without stablizer is prepared into so-called emulsifying wax, and this wax is prepared into so-called emulsifying ointment base together with oil & fat, this emulsifying ointment base forms the oil dispersion phase of cream formulation meanwhile.
The emulgent and the emulsion stabilisers that are applicable to invention formulation comprise
60,
80, cetostearyl alcohol, phenylcarbinol, tetradecyl alcohol, Zerol and Sulfuric acid,monododecyl ester, sodium salt.
Carry out according to the cosmetic properties that will realize expecting for the suitable oil of said preparation or the selection of fat.This emulsifiable paste should be preferably non-greasy, non-contamination and the product that can wash and have suitable denseness to avoid leaking from pipe or other container.Can use the propylene glycol diesters of straight or branched, list or dialkyl such as two-different adipic acid ester, isocetyl stearate base ester, coconut fatty acid, isopropyl myristate, decyl oleate, Wickenol 111, butyl stearate, palmitinic acid 2-(ethyl hexyl) ester or be called the mixture of side chain/ester of Crodamol CAP, last three kinds is preferred ester.They can be used alone or use according to required combination of properties.Or, use high-melting-point lipid such as white vaseline.
Preparation for topical application to eye also comprises eye drops, wherein the activeconstituents appropriate carrier especially in aqueous solvent of being dissolved or being suspended in activeconstituents.Activeconstituents is preferably present in this type of preparation with the concentration of 0.5 to 20%, is advantageously 0.5 to 10%, particularly about 2.0%w/w.
Preparation for topical application to mouth comprises, lozenge, and it contains the activeconstituents be in flavoured base (being generally sucrose, gum arabic or tragacanth gum); Pastille, it contains the activeconstituents be in inert base (such as gelatin and glycerine, or sucrose and gum arabic); And mouth wash shua, it contains the activeconstituents be in suitable liquid vehicle.
Preparation for rectal administration can exist with the suppository with appropriate substrate, and this matrix comprises such as cocoa butter or salicylate.
For in lung or the preparation of the intranasal administration granularity with 0.1 to 500 micrometer range (comprise the granularity that scope is 0.1 to 500 micron, increment micron is such as 0.5,1,30 micron, 35 microns etc.), it is by sucking fast through nasal meatus or being sucked by per os thus arrive alveolar sac to use.Suitable preparation comprises water-based or the oily solution of activeconstituents.Be applicable to the preparation that aerosol or dry powder doses use can prepare according to conventional methods, and can administration together with other therapeutical agent, this other therapeutical agent be such as up to the present for compound that treatment as described below or flu-prevention A or B are infected.
Preparation for vaginal application can exist with vaginal suppository, tampon agent, ointment, gelifying agent, paste, foaming agent or sprays, and these preparations also contain applicable carrier known in the art except activeconstituents.
Be applicable to the preparation that parenteral uses and can comprise water-based and non-aqueous sterile injection solution, it can contain oxidation inhibitor, buffer reagent, fungistat and provides and expect the solute of the preparation that the blood of experimenter is isotonic; And water-based and the non-aqueous sterile suspensions of suspension agent and thickening material can be comprised.
Preparation can be present in single dose or multi-dose container, such as, in sealed ampoule bottle or phial, and can preserve under lyophilize (freeze-drying) condition, only need adding sterile liquid carrier (solvent) such as water for injection before use immediately.Facing and joining injection solution and suspension is prepare from the sterilized powder of aforesaid kind, particle or tablet.Preferred unit dose formulations be containing activeconstituents as herein every per daily dose recited above or unit sub-doses every day, or the formulation of its suitable number.
Should be appreciated that above except the composition mentioned especially, other reagent relevant with discussed preparation type that preparation of the present invention can comprise this area routine, such as, be suitable for those Orally administered reagent, comprise seasonings.
Invention further provides veterinary composition, it comprises at least one activeconstituents as defined above and Veterinary carriers.
Veterinary carriers is the material that can be used for using said composition object, can be solid, liquid or gaseous matter, its be inert or veterinary applications acceptable, and compatible with activeconstituents.These veterinary compositions can be used oral, parenteral, or are used by any approach that other is expected.
The compounds of this invention can be used for providing controlled-release pharmaceutical formulation, it contains one or more the compounds of this invention (" controlled release preparation ") as activeconstituents, in said preparation, the release of activeconstituents is controlled and regulates, to make administration frequency lower or improve pharmacokinetics or the toxicity of given activeconstituents.
The effective dose of activeconstituents at least depends on sanatory character, toxicity, no matter and this compound whether for prevention (comparatively low dosage) no matter or suppress active influenza infection, also medication and pharmaceutical preparation, and the effective dose of activeconstituents will use the research of routine dose increasing proportion to determine by clinician.
Expect for every day about 0.0001 is to about 100mg/kg body weight.Typically be every day about 0.01 to about 10mg/kg body weight.Be more typically every day about 0.01 to about 5mg/kg body weight.Be more typically every day about 0.05 to about 0.5mg/kg body weight.Such as inhalation, adult's recommended dose every day of about 70kg body weight will be 1mg to 1000mg, be preferably 5mg to > 500
Mg, and the form that can be single dose or multiple doses.When the pathologic condition of experimenter needs, the effective per daily dose of larger treatment also can be used.
Activeconstituents of the present invention (or compound) also can use with other active ingredient combinations.This type of combination is selected to be based on treated illness, the cross reactivity of each composition and the pharmaceutical properties of combination.Such as, when treat respiratory system virus infection particularly influenza infection time, composition of the present invention and antiviral agent (such as amantadine, Rimantadine and ribavirin), mucolytic, expectorant, bronchodilator, microbiotic, febrifuge or anodyne combine.Usually, microbiotic, febrifuge are used together with the compounds of this invention with anodyne.
The detailed description of the present invention is enough to make those skilled in the art can prepare and use the theme material of following examples.Obviously, some modification of the method and composition of following examples can be carried out in the scope of the invention and spirit.The present invention is further described with reference to following exemplary embodiment and appended Fig. 1-4.
Embodiment
Embodiment 1
The preparation of intermediate 1
The sialic acid (0.73mmol) of the 225mg be dissolved in the anhydrous methanol of 40ml is mixed into the Dowex 50 (H of 0.5g
+) in resin.This mixture is made to reflux under continuous stirring 48 hours.Measured from Resorcinol HCl and thiobarbituricacidα-(TBA) analysis, constantly little 24 and 48, there is the sialic acid of 85% and 97% to change into intermediate 1 respectively.Then by resin filter on general paper filter, and elutriant is concentrated into drying by rotatory evaporator, obtains the micro-yellow liquid of oily.Then by the ether of this oily liquids with a small amount of volume: the mixture of methyl alcohol (3: 1w/w) restores.At 4 DEG C, make this solution keep leaving standstill 24-48 hour, by collected by filtration throw out, then use P
2o
5dry.Obtain solid intermediate 1 (the M.W.337.4) (yield: 60.0%) of 145mg.
Gained intermediate 1 couple of Resorcinol-HCl reacts aobvious positive (identical with sialic acid intensity), and aobvious negative to TBA reaction.
Embodiment 2
The preparation of intermediate 2
The intermediate 1 (0.43mmol) of 145mg is dissolved in the 0.06M aqueous sodium hydroxide solution of 10.7ml, more at room temperature Keep agitation 2-3 hour.Then by this mixture Dowex 50 (H
+) resin moderated to pH7.0-7.5.Leach resin, by elutriant lyophilize, the solid of the look that obtains turning white.Obtain solid intermediate 2 (the M.W.323.3) (yield: 95.0%) of 132.6mg.
Gained intermediate 2 couples of Resorcinol-HCl react aobvious positive (identical with sialic acid intensity), and aobvious negative to TBA reaction.
Embodiment 3
The preparation of intermediate 3
The solid (intermediate 2) of the freeze-drying of 132.6mg (0.41mmo l) is dissolved in the distilled water of 4.3ml.Then the 0.038M sodium periodate aqueous solution (NaIO of every part of 10.7ml is added
4) (0.41mmol) (mol ratio=1: 1), then this solution is at room temperature in the dark kept Keep agitation 1 hour.The 0.1M barium acetate aqueous solution of 12.8ml is added in this mixture, to make excessive iodate and periodate precipitation.General paper filter is used to be filtered by this mixture.This elutriant is made to advertise into carbonic acid gas saturated, to make excessive barium acetate precipitation, then paper using frit.Elutriant lyophilize, obtains micro-yellow solid.Collect solid intermediate 3 (the M.W.291.3) (yield: 85.0%) of 101.9mg.
Gained intermediate 3 couples of Resorcinol-HCl react aobvious positive (identical with sialic acid intensity), and aobvious negative to TBA reaction.
Embodiment 4
The preparation of intermediate 4
The solid of the freeze-drying of 132.6mg (0.41mmol) (intermediate 2) is dissolved in the distilled water of 4.3ml.By the 0.2M sodium periodate (NaIO of 10.7ml
4) (mol ratio: 1: 5.24) add in this solution makes its at room temperature in the dark Keep agitation 1 hour to the aqueous solution (2.14mmol).
The 0.1M barium acetate aqueous solution of 12.8ml is added in this mixture, to make excessive iodate and periodate precipitation.General paper filter is used to be filtered by this mixture.Elutriant is advertised into carbonic acid gas saturated, to be settled out excessive barium acetate, then filter on paper filter.Elutriant lyophilize, obtains micro-yellow solid.Collect solid intermediate 4 (the M.W.261.23) (yield: 85.0%) of 91.4mg.
Gained intermediate 4 couples of Resorcinol-HCl react aobvious positive (identical with sialic acid intensity), and aobvious negative to TBA reaction.
Embodiment 5
The preparation of intermediate 5
The powder intermediate 4 of the freeze-drying of 91.43mg (0.35mmol) is dissolved in the 2.3mM aqueous formic acid of the 2.0ml of pH about 4.0.Make solution be heated to 80 DEG C and reach 1 hour.Then solution lyophilize is made.
Obtain intermediate 5 (the M.W.247.20) (yield: 84.1%) of 72.76mg.
Gained intermediate 5 couples of Resorcinol-HCl and TBA reaction is all aobvious positive.
Embodiment 6
The preparation of intermediate 6
The solid intermediate 3 of the freeze-drying of 101.96mg (0.35mmol) is dissolved in the 2.3mM aqueous formic acid of the 2.0ml of pH about 4.0.Make solution be heated to 80 DEG C and reach 1 hour.Then solution lyophilize is made.
Obtain intermediate 6 (the M.W.277.23) (yield: 96.7%) of 93.83mg.
Gained intermediate 6 couples of Resorcinol-HCl and TBA reaction is all aobvious positive.
Embodiment 7
The preparation of compound 1
The solid of the freeze-drying of 84.0mg (0.34mmol) (intermediate 5) is dissolved in the distilled water of 5.0ml.Then the amantadine (0.34mmol) of 51.442mg is added under slow stirring.Slowly stirring and at 4 DEG C, making the stirring of this mixture spend the night.After adding 40.0mg sodium borohydride, reaction mixture is made to keep 1 hour under stirring at room temperature.Then make this mixture by Dowex 50 (H
+) resin column, make excessive reforming sodium borohydride become boric acid.Make elutriant lyophilize.Then with aliquot methyl alcohol, cryodesiccated solid is restored, make elutriant dry.This operation is repeated at least 3 times.Dry solid is finally restored with aliquot water, and then lyophilize, obtains finalization compound 1.
Obtain compound 1 (the M.W.382.45) (yield: 85.0%) of 110.53mg.
The compound collected 1 couple of Resorcinol-HCl reacts and all aobvious positive of TBA reaction.FT-IR (Fourier transform infrared spectroscopy) spectrum (A) of compound 1 is plotted in Fig. 1, and (B) that contrast with it is starting intermediates amantadine, shows following reference value: a) acid amides I bands of a spectrum: 1640cm
-1; B) acid amides II bands of a spectrum: 1550cm
-1; C) primary amino group: 600-800cm
-1, 1590-650cm
-1, 3330-3380cm
-1; Parahelium group: 700-800cm
-1; 1615cm
-1; 3300cm
-1.
Embodiment 8
The preparation of compound 2
The solid of the freeze-drying of 94.26mg (0.34mmol) (intermediate 6) is dissolved in the distilled water of 5.0ml.Then the ribavirin (0.34mmol) of 82.7mg is added under slow stirring.This mixture is slowly stirred at 4 DEG C spend the night.After adding the sodium borohydride of 40.0mg, reaction mixture is kept 1 hour under stirring and room temperature.Then make this mixture by Dowex50 (H
+) resin column, become boric acid to make excessive reforming sodium borohydride.Make elutriant lyophilize.Then with aliquot methyl alcohol, lyophilize thing is restored, make elutriant again dry.This operation repeats at least 3 times.Make dry solid finally be dissolved in aliquot water, then lyophilize, obtains solid (compound 2).Collect solid chemical compound 2 (the M.W.505.4) (yield: 89.0%) of 94.02mg.
Compound 2 couples of Resorcinol-HCl collected and TBA reaction is all aobvious positive.
Embodiment 9
The preparation of compound 3
The solid of the freeze-drying of 94.26mg (0.34mmol) (intermediate 6) is dissolved in the distilled water of 5.0ml.Then the amantadine (0.34mmol) of 51.442mg is added under slow stirring.This mixture is slowly stirred at 4 DEG C spend the night.After adding the sodium borohydride of 40.0mg, reaction mixture is made at room temperature to stir 1 hour.Then make this mixture by Dowex 50 (H
+) post, to make excessive reforming sodium borohydride become boric acid.Make elutriant lyophilize.Then by cryodesiccated dissolution of solid in aliquot methyl alcohol, make filtrate dry.This operation repeats at least 3 times.Then by the dissolution of solid of drying in aliquot water, then lyophilize, obtains finalization compound 3.Collect solid chemical compound 3 (the M.W.412.5) (yield: 87.0%) of 122.02mg.
Compound 3 couples of Resorcinol-HCl collected and TBA reaction is all aobvious positive.
Embodiment 10
The preparation of compound 4
The solid of the freeze-drying of 84.0mg (0.34mmol) (intermediate 5) is dissolved in the distilled water of 5.0ml.Then the Rimantadine (0.34mmol) of 60.962mg is added under slow stirring.This mixture is slowly stirred at 4 DEG C spend the night.After adding the sodium borohydride of 40.0mg, reaction mixture is made at room temperature to stir 1 hour.Then make this mixture by Dowex 50 (H
+) post, become boric acid to make excessive reforming sodium borohydride.Make elutriant lyophilize.Then with aliquot methyl alcohol, cryodesiccated solid is restored, make elutriant dry.This operation repeats at least 3 times.Then by the powder of drying aliquot water dissolution, then lyophilize, obtains finalization compound 4.
Collect compound 4 (the M.W.410.5) (yield: 90.0%) of 125.61mg.
The compound collected 4 couples of Resorcinol-HCl react and all aobvious positive of TBA reaction.The FT-IR spectrum (A) of compound 4 is plotted in Fig. 2, and (B) that contrast with it is starting intermediates Rimantadine, shows following reference value: a) acid amides I bands of a spectrum: 1640cm
-1; B) acid amides II bands of a spectrum: 1550cm
-1; C) primary amino group: 600-800cm
-1, 1590-650cm
-1, 3330-3380cm
-1; Parahelium group: 700-800cm
-1; 1615cm
-1; 3300cm
-1.
Embodiment 11
The preparation of compound 5
The solid of the freeze-drying of 94.26mg (0.34mmol) (intermediate 6) is dissolved in the distilled water of 5.0ml.Then the Rimantadine (0.34mmol) of 60.962mg is added under slow stirring.This mixture is slowly stirred at 4 DEG C spend the night.After adding the sodium borohydride of 40.0mg, reaction mixture is made at room temperature to stir 1 hour.Then make this mixture by Dowex 50 (H
+) post, to make excessive reforming sodium borohydride become boric acid.Make elutriant lyophilize.Lyophilize thing is dissolved in aliquot methyl alcohol, makes elutriant dry.This operation repeats at least 3 times.Then make dry solid aliquot water dissolution, then lyophilize, obtains finalization compound 5.Collect final solid chemical compound 5 (the M.W.440.5) (yield: 92.0%) of 137.79mg.
Compound 5 couples of Resorcinol-HCl collected and TBA reaction is all aobvious positive.
Embodiment 12
The preparation of compound 6
The solid (intermediate 5) of the freeze-drying of 84.0mg (0.34mmol) is dissolved in the distilled water of 5.0ml.Then the somantadine (0.34mmol) of 70.5mg is added under slow stirring.This mixture is slowly stirred at 4 DEG C spend the night.After adding the sodium borohydride of 40.0mg, reaction mixture is made at room temperature to stir 1 hour.Then make this mixture by Dowex 50 (H
+) post, become boric acid to make excessive reforming sodium borohydride.Make elutriant lyophilize.Lyophilize thing is dissolved in aliquot methyl alcohol, makes filtrate dry.This operation repeats at least 3 times.Then by the powder of drying aliquot water dissolution, then lyophilize, obtains finalization compound 6.Collect solid chemical compound 6 (the M.W.438.6) (yield: 84.0%) of 125.26mg.
Compound 6 couples of Resorcinol-HCl collected and TBA reaction is all aobvious positive.
Embodiment 13
The preparation of compound 7
The solid of the freeze-drying of 94.26mg (0.34mmol) (intermediate 6) is dissolved in the distilled water of 5.0ml.Then the somantadine (0.34mmol) of 61.0mg is added under slow stirring.This mixture is slowly stirred at 4 DEG C spend the night.After adding the sodium borohydride of 40.0mg, reaction mixture is made at room temperature to stir 1 hour.Then make this mixture by Dowex 50 (H
+) post, become boric acid to make excessive reforming sodium borohydride.Then by elutriant lyophilize.Then make lyophilize thing be dissolved in aliquot methyl alcohol, make filtrate dry.This operation repeats at least 3 times.Then by the powder of drying aliquot water dissolution, then lyophilize, obtains finalization compound 7.Collect solid chemical compound 7 (the M.W.468.6) (yield: 85.0%) of 135.43mg.
The compound collected is all aobvious positive to Resorcinol-HCl and TBA reaction.
Embodiment 14
The preparation of compound 8
The solid of the freeze-drying of 84.0mg (0.34mmol) (intermediate 5) is dissolved in the distilled water of 5.0ml.Then the memantine (0.34mmol) of 61.0mg is added under slow stirring.This mixture is slowly stirred at 4 DEG C spend the night.After adding the sodium borohydride of 40.0mg, reaction mixture is made at room temperature to stir 1 hour.Then make this mixture by Dowex 50 (H
+) post, become boric acid to make excessive reforming sodium borohydride.Make elutriant lyophilize.Lyophilize thing is dissolved in aliquot methyl alcohol, makes filtrate dry.This operation repeats at least 3 times.Then by the powder of drying aliquot water dissolution, then lyophilize, obtains finalization compound 8.Collect solid chemical compound 8 (the M.W.410.5) (yield: 93.0%) of 129.80mg.
The compound collected is all aobvious positive to Resorcinol-HCl and TBA reaction.The FT-IR spectrum (A) of compound 8 is plotted in Fig. 3, and (B) that contrast with it is starting intermediates memantine, shows following reference value: a) acid amides I bands of a spectrum: 1640cm
-1; B) acid amides II bands of a spectrum: 1550cm
-1; C) primary amino group: 600-800cm
-1, 1590-650cm
-1, 3330-3380cm
-1; Parahelium group: 700-800cm
-1; 1615cm
-1; 3300cm
-1.
Embodiment 15
The preparation of compound 9
The solid of the freeze-drying of 94.26mg (0.34mmol) (intermediate 6) is dissolved in the distilled water of 5.0ml.Then the memantine (0.34mmol) of 61.0mg is added under slow stirring.This mixture is slowly stirred at 4 DEG C spend the night.After adding the sodium borohydride of 40.0mg, reaction mixture is made at room temperature to stir 1 hour.Then make this mixture by Dowex 50 (H
+) post, become boric acid to make excessive reforming sodium borohydride.Then by elutriant lyophilize.Then make lyophilize thing be dissolved in aliquot methyl alcohol, make filtrate dry.This operation repeats at least 3 times.Then by the powder of drying aliquot water dissolution, then lyophilize, obtains finalization compound 9.
Collect compound 9 (the M.W.440.5) (yield: 90.0%) of 134.79mg.
Compound 9 couples of Resorcinol-HCl collected and TBA reaction is all aobvious positive.
Embodiment 16
The preparation of compound 10
The solid of the freeze-drying of 84.0mg (0.34mmol) (intermediate 5) is dissolved in the distilled water of 5.0ml.Then the ribavirin (0.34mmol) of 82.7mg is added under slow stirring.This mixture is slowly stirred at 4 DEG C spend the night.After adding the sodium borohydride of 40.0mg, reaction mixture is made at room temperature to stir 1 hour.Then make this mixture by Dowex 50 (H
+) post, become boric acid to make excessive reforming sodium borohydride.Make elutriant lyophilize.Lyophilize thing is dissolved in aliquot methyl alcohol, makes filtrate dry.This operation repeats at least 3 times.Then by the powder of drying aliquot water dissolution, then lyophilize, obtains finalization compound 10.Collect compound 10 (the M.W.475.4) (yield: 87.0%) of 140.62mg.The compound collected is aobvious positive to Resorcinol-HCl and TBA reaction.The FT-IR spectrum (A) of compound 10 is plotted in Fig. 4, and (B) that contrast with it is starting intermediates ribavirin, shows following reference value: a) acid amides I bands of a spectrum: 1640cm
-1; B) acid amides II bands of a spectrum: 1550cm
-1; C) primary amino group: 600-800cm
-1, 1590-650cm
-1, 3330-3380cm
-1; Parahelium group: 700-800cm
-1; 1615cm
-1; 3300cm
-1.
Embodiment 17
The preparation of compound 11
The intermediate sialic acid of the freeze-drying of 102.5mg (0.34mmol) is made to be dissolved in the methyl alcohol of 50.0ml: in the mixture of distilled water (9: 1, v/v).Then the amantadine (0.34mmol) of 51.442mg is added under slow stirring.After adding the sodium borohydride of 100.0mg, reaction mixture is stirred 1 hour at 60 DEG C.Then make this mixture by Dowex 50 (H
+) post, to make excessive reforming sodium borohydride become boric acid.Make elutriant lyophilize.Lyophilize thing is dissolved in aliquot methyl alcohol, makes filtrate dry.This operation repeats at least 5 times.Then by the solid of drying aliquot water dissolution, then lyophilize, obtains finalization compound 11.Collect and amount to 133.90mg solid chemical compound 11 (M.W.442.5) (yield: 89.0%).The compound collected reacts aobvious positive to Resorcinol-HCl, and aobvious negative to TBA reaction.
Embodiment 18
The compounds of this invention suppresses the antiviral activity evaluation of influenza virus A and Type B strain.
Object and research project
Especially, compare with memantine with amantadine, have rated two kinds of Anti-viral activity in vitro having the compound of the antiviral activity of high value and compound 1 and compound 8 to suppress influenza virus A and Type B.In this research, the virus type B employing two kinds of influenza A virus separated He separate.
Materials and methods
The propagation of 1-influenza virus and titration
Make influenza virus A/H 3N2 (A/ Panama/2007/99), A/H1N1 (A/NewCaledonia/20/99) and B (B/ Shandong/7/97) bacterial strain propagates in embryo's egg.In brief, strains of influenza viruses is made to be inoculated in the fertilization egg of 11 days by allantoic route.Then make egg hatch 3 days at 37 DEG C, collect allantoic fluid, then measure titre.
Carry out analysis by plaque-forming assay (plaque assay, PA) to measure.In detail, the serial dilutions (10 of the virus making each be separated
-1-10
-11) be inoculated in MDCK (Madin-Darby canine kidney) cell of the fusion individual layer in 12 orifice plates.Hatch 1 hour at 37 DEG C after, removing virus inoculation thing, then add infective agents (MEM (minimum essential medium) containing 10 μ g/ml trypsinase, 2% agar).At 37 DEG C, 5%CO
2after 3 days are hatched in place, cell monolayer 5% glutaraldehyde solution is fixed, after removing agar, with the solution-dyed of 5% carbol fuchsin.
To plaque range estimation counting, this compartment analysis represents with the number of every ml plaque forming unit (PFU) (PFU/ml).
The evaluation of the antiviral activity of the compound 1 that 2-is relevant with amantadine and memantine and compound 8.
The preparation of the compound 2.1. will analyzed
Drying, after 72 hours, makes compound 1 and 8, and contrast compound amantadine and memantine suitably redissolve in sterile distilled water with the concentration of 1000 μMs.Then based on 10, carry out serial dilution to each test compounds, scope is 0.01 μM to 100 μMs.
2.2. plaque reduces analysis (PRA)
Make the fusion MDCK cell monolayers (1 × 10 being grown on 12 orifice plates
5cell/ml) infect with each isolated viral of about 50PFU/ml (A/ Panama/2007/99-H3N2, A/ New Caledonia/20/99-H1N1 and B/ Shandong/7/97).At 37 DEG C, 5%CO
2under hatch 1 hour with after making viruses adsorption, removing virus inoculation thing, washs 2 times by this cell monolayer MEM substratum.Add in each hole and cover substratum (10 μ g/ml trypsinase in MEM, 2% agar), this covering substratum contains the analysis of compounds (from 0.01 μM to 100 μMs) of serial dilution.Double is carried out in test, adds the reaction contrast not containing antiviral compound simultaneously.Make cell culture at 37 DEG C, 5%CO thus
2under hatch 3 days.Then, cell monolayer 5% glutaraldehyde solution is fixed, more at room temperature hatches at least 3 hours, to promote to penetrate in agar.After removing agar, with 5% carbolfuchsin solution, cell monolayer is dyeed.Range estimation counting plaque, calculates the degree of Plaque reduction assay relative to the contrast not containing test compounds.Measure the necessary compound concentration of plaque reduced number 50% relative to the contrast not containing the compounds of this invention thus.
Result
The antiviral activity of compound 1 and compound 8 and the antiviral activity of control compound amantadine and memantine reduce analysis (PRA) by plaque and use mdck cell to evaluate.Result is shown in table 1.
Compound 1 demonstrates the activity of the suppression A type H 3N2 virus similar to amantadine, but suppresses the activity of A type H1N1 virus low 10 times.Compound 1 suppresses the antiviral activity of influenza virus B type higher than this activity of amantadine (> 100 μMs) (> 10 times) (table 1).
Compound 8 is to the antiviral activity suppressing the influenza A (A/H3N2 and A/H1N1) analyzed to demonstrate 10 times magnitudes larger than memantine.Within the scope of test concentrations, compound 8 demonstrates the antiviral activity (table 1) suppressing Type B influenza virus.
Table 1
Conclusion:
The data presentation compound 1 provided, compared to amantadine, improves the antiviral activity suppressing Type B influenza virus.Needs are remembered, amantadine exists only in influenza A by blocking-up and is not present in the M2 albumen onset of the ionic channel in Type B influenza virus, and therefore this may be a kind of new mechanism of action, and this may be the target of future studies.
Compound 8 demonstrates the antiviral activity of the suppression influenza A of higher than memantine 10 times.
Accompanying drawing explanation
Fig. 1 is FT-I R (Fourier transform infrared spectroscopy) spectrum (A) of compound 1, in contrast starting intermediates amantadine (B),
Fig. 2 is the FT-IR spectrum (A) of compound 4, in contrast starting intermediates Rimantadine (B),
Fig. 3 is the FT-IR spectrum (A) of compound 8, in contrast starting intermediates memantine (B),
Fig. 4 is the FT-IR spectrum (A) of compound 10, in contrast starting intermediates ribavirin (B).
Claims (4)
1. the compound of structural formula (I) and additive salt thereof;
It comprises-B/ part; And
Wherein-B/ part is:
-B1/ part; And
Wherein:
X represents-O-;
The C of singly-bound shack
2-C
3;
R
1expression-OH; And
R
2-OH or-NH
2; And
R
3-NHCO-CH
3; And
R
4-CH
2-B/ part, and
R
5-NH-; And
R
6-H or-CH
3; And
R
7-H or-CH
3.
2. the compound of claim 1, it is be selected from following C
7the compound of single-B/ part-replacement:
3. a pharmaceutical composition, it is included in compound or its mixture of the claim 1 or 2 in pharmaceutically acceptable carrier.
4. claim 1 or 2 compound for the preparation for the treatment of by erythrocyte agglutination element and/or neuraminidase and/or containing M
2purposes in the medicine of the virus infection that the virus of albumen causes, wherein said infection is caused by A and Type B influenza virus or its mutant.
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| ITMI20070093 ITMI20070093A1 (en) | 2007-01-23 | 2007-01-23 | ANTIVIRAL COMPOUNDS |
| ITMI2007A000093 | 2007-01-23 | ||
| ITMI2007A002212 | 2007-11-22 | ||
| ITMI20072212 ITMI20072212A1 (en) | 2007-11-22 | 2007-11-22 | ANTIVIRAL COMPOUNDS |
| PCT/EP2008/050703 WO2008090151A1 (en) | 2007-01-23 | 2008-01-22 | Antiviral compounds |
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| CN101622265A CN101622265A (en) | 2010-01-06 |
| CN101622265B true CN101622265B (en) | 2015-06-10 |
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| IT (1) | ITMI20070093A1 (en) |
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| CN103509066B (en) * | 2012-06-25 | 2016-03-30 | 沈阳药科大学 | A kind of lipid derivate and application thereof containing sialic acid fragment |
| CN104031097B (en) * | 2013-03-04 | 2016-12-28 | 沈阳药科大学 | A kind of lipid derivate containing sialic acids groups and application thereof |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO1992006691A1 (en) * | 1990-10-19 | 1992-04-30 | Biota Scientific Management Pty. Ltd. | Anti-viral compounds that bind the active site of influenza neuramidase and display in vivo activity against orthomyxovirus and paramyxovirus |
| WO2000029419A1 (en) * | 1998-11-17 | 2000-05-25 | Tropix, Inc. | Chemiluminescent substrates for neuraminidase, assays for detection of neuraminidase and kits therefor |
-
2007
- 2007-01-23 IT ITMI20070093 patent/ITMI20070093A1/en unknown
-
2008
- 2008-01-22 CN CN200880006407.8A patent/CN101622265B/en not_active Expired - Fee Related
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO1992006691A1 (en) * | 1990-10-19 | 1992-04-30 | Biota Scientific Management Pty. Ltd. | Anti-viral compounds that bind the active site of influenza neuramidase and display in vivo activity against orthomyxovirus and paramyxovirus |
| WO2000029419A1 (en) * | 1998-11-17 | 2000-05-25 | Tropix, Inc. | Chemiluminescent substrates for neuraminidase, assays for detection of neuraminidase and kits therefor |
Non-Patent Citations (2)
| Title |
|---|
| Mark von Itzstein et al.Rational design of potent sialidase-based inhibitors of influenza virus replication.《NATURE》.1993,第363卷418-423. * |
| Sudhakar Babu Y et al.BCX-1812 (RWJ-270201): Discovery of a Novel, Highly Potent, Orally Active, and Selective Influenza Neuraminidase Inhibitor through Structure-Based Drug Design.《Journal of Medicinal Chemistry》.2000,第43卷3482-3486. * |
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|---|---|
| ITMI20070093A1 (en) | 2008-07-24 |
| CN101622265A (en) | 2010-01-06 |
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