CN100536837C

CN100536837C - Application of scyllo inositol for preparation of diagnosing reagent

Info

Publication number: CN100536837C
Application number: CNB2004800113358A
Authority: CN
Inventors: 乔安妮·麦克劳林
Original assignee: Individual
Current assignee: Individual
Priority date: 2003-02-27
Filing date: 2004-02-27
Publication date: 2009-09-09
Anticipated expiration: 2024-02-27
Also published as: CN1780613A; CN101385722A

Abstract

Disclosed are methods of preventing, treating, or diagnosing in a subject a disorder in protein folding or aggregation., or amyloid formation., deposition, accumulation, or persistence consisting of administering to said subject a pharmaceutically effective amount of inositol stereoisomers, enantiomers or derivatives thereof.

Description

The purposes of Cocositol in the preparation diagnostic reagent

Related application

The application requires the priority of U.S. Provisional Patent Application series number 60/451,363,60/520,958 and 60/523,534, and above-mentioned application is respectively at submitting on February 27th, 2003, on November 17th, 2003 and on November 19th, 2003.

Invention field

The present invention relates to treat the method for Alzheimer and other amyloidosis; More particularly, the present invention relates in the therapeutic of Alzheimer and other amyloidosis gets involved, suppress and reduce the method that amyloid fibrils forms.

Description of related art

The neuropathological feature of Alzheimer is amyloid beta deposition thing, neurofibrillary tangle and selective neuronal loss.The main component of amyloid beta deposition thing is amyloid-β (A β), and it is the peptide of a kind of 39-43 residue.The A β of the soluble form that is produced by the amyloid precursor protein cracking is metabolic normal product.The importance outstanding behaviours of residue 1-42 in the Alzheimer (A β 42) causes A β 42 to increase than A β 1-40 output in codon 717 sudden changes of having found amyloid precursor protein gene, presenilin 1 and presenilin 2 genes.These with ripe speckle and dispersivity amyloid in exist the relevant result of A β 42 to derive following hypothesis: the material of the more amyloids of this generation may be crucial factor in speckle forms.This hypothesis has obtained the support of the following fact: A β 42 depositions have surpassed A β 40 depositions at the mongolism of PS1 sudden change and in the hereditary cerebral hemorrhage of amyloidosis.

Many in vitro studies are verified, and A β can be neurovirulent or can increase the susceptibility that neuron is subjected to exitotoxicity, metabolism or oxidative damage.At first, people only think that the A of fibril form is toxic to neuron, but more fully the sign of A beta structure have been confirmed that the dimerization of A β and little aggregation also are neurovirulent.These Notes of Key Datas prevent that A β oligomerization from will be the prevention neurodegenerative suitable strategy relevant with AD.Several studies is verified, utilize to increase the neurotoxicity that the chemical compound of neuron resistance can cause at external elimination A β, this chemical compound is by the targeting cellular pathways relevant with natural death of cerebral cells, after the A that destroys the path is beta induced, block downstream pathway, perhaps block A β oligomerization and final fibril and form and increase the neuron resistance.Illustrate to play though await and induce neurovirulent A β site, can block its toxic action by various medicament.

In the AD process, it can be early stage interfered with step that A β-fibril is accommodated on neuron and the glial cell film.The formation of amyloid plaque, neurotoxicity and inflammation may be the result that A and the interaction of molecules that contains glycosyl group directly or indirectly cause.Early stage research is verified, and A β and aminopolysaccharide interact and cause the A beta peptide aggregation may increase its insoluble and speckle persistency.Aminopolysaccharide is also relevant with the microgliacyte activation with neurotoxicity.In addition, the interaction with glycolipid such as ganglioside causes stabilisation and prevents that the Ab fibril from forming and A β produces the site.On the other hand, phosphatidylinositols family causes quickening fibriilar formation.A base of phosphatidylinositols is an inositol, and it is a kind of naturally occurring simple sugars, and is relevant with the control of lipid biosynthesis, signal transduction and osmotic pressure.

It should be noted that equally, multiple other human diseases is alleged occurrence amyloid beta deposition and be usually directed to general organ (promptly being positioned at the organ or tissue of central nervous system periphery) also, and these organs are with causing organ dysfunction or depleted amyloid to gather.For Alzheimer and " general " amyloid disease, do not cure at present or effective Therapeutic Method, and patient is dead in the 3-10 of morbidity usually.

United States Patent (USP) 4,847 discloses phytic acid, phytate, the isomer of phytic acid or the purposes that hydrolyzate is used for the treatment of Alzheimer No. 082.The isomer of phytic acid or phytate is also disclosed, its comprise six phosphoric acid myo-inositol esters, six phosphoric acid Cocositol esters, six phosphoric acid D-chirality-mesoinositols, six phosphoric acid L-chirality-mesoinositols, six phosphoric acid new-inositol (neo-inositol) ester and six phosphoric acid are sticking-inositol (muco-inositol) ester configuration.Phytic acid is inositol-six phosphate ester (IP6).

United States Patent (USP) 5,112 discloses phytic acid No. 814 and isomery is used for the treatment of parkinsonian purposes.With United States Patent (USP) 4,847, No. 082 is identical, the phytic acid isomer of this patent disclosure on six carbon inositol sugar, have six phosphate-based.

It should be noted that in follow-up open source literature, studied inositol-phosplate, inositol-1,4-bisphosphate and inositol-1,4, the 5-triguaiacyl phosphate suppresses the ability of amyloid-β peptide microfibre generation and finds that they are invalid (J.Mol.Biol.278:183-194,1998).

People such as Barak disclose the purposes that inositol is used for the treatment of Alzheimer (AD).(Prog?Neuro-psychopamacol?&?Biol?Psychiat.20：729-735，2000)。But this list of references does not disclose the purposes of inositol isomer.Between inositol treatment patient AD and placebo (glucose) treatment patient AD, aspect overall cognitive function mark (CAMCOG index), the patient of inositol treatment does not demonstrate evident difference, but exponential two the concrete inferior indexs of CAMCOG but demonstrate significant improvement the (orientation and language).

Levine J. has summarized people's such as above-mentioned Barak article and has particularly pointed out the inositol treatment and has been unfavorable for AD or the inductive cognitive impairment of ECT-(Eur Neuropsychoparm.1997; 7,147-155,1997).

With reference to people's such as above-mentioned Barak article, it is useless for Alzheimer that people such as Colodny further research and propose inositol, does not therefore have purposes (Altern Med Rev 3 (6): 432-47,1998) open or the isomer of suggestion inositol.

People such as McLaurin disclose the little micelle (J.Mol.Biol.278,183-194,1998) that myo-inositol can be stablized A β 42.In addition, people such as McLaurin discloses table-inositol and Cocositol but has not been that chirality-inositol can induce A β 42 to take place by random structural transformation to beta structure (the J BiolChem.6 month 16; 275 (24): 18495-502,2000; With J Struct Biol 130:259-270,2000).In addition, stereoisomer all can not be induced the structural transformation of A β 40.Ultramicroscope shows that inositol can be stablized the little accumulation of A β 42.The interaction that these lists of references also disclose inositol-A β has produced a kind of PC-12 cell and avirulent complex of former generation human neure culture to the nerve growth factor differentiation.

Alzheimer has been finished many work, but current known chemical compound or medicament seldom is used for the treatment of scheme and forms, deposits, gathers and/or retain to stop or to reverse the amyloid that takes place in Alzheimer or other amyloidosis.

Therefore, being starved of the noval chemical compound or the medicament that are used for the treatment of scheme forms, deposits, gathers and/or retains to stop or to reverse the amyloid that takes place in Alzheimer or other amyloidosis.

Summary of the invention

The invention provides a kind of treatment or prevention experimenter with protein folding or gather disorder or amyloid forms, deposits, gathers or retain the method for relevant central or peripheral nervous system or general organ disease, it comprises the chemical compound that is selected from following structure to the medical effective dose of described experimenter's administration:

Wherein, each R ₁, R _{1 '}, R ₂, R _{2 '}, R ₃, R _{3 '}, R ₄, R _{4 '}, R ₅, R _{5 '}, R ₆And R _{6 '}Be independently selected from following group:

(a) hydrogen atom;

(b) NHR ₇, wherein said R ₇Be selected from hydrogen; C ₂-C ₁₀Acyl group and C ₁-C ₁₀Alkyl;

(c) NR ₈R ₉, wherein said R ₈Be C ₂-C ₁₀Acyl group or C ₁-C ₁₀Alkyl, and described R ₉Be C ₂-C ₁₀Acyl group or C ₁-C ₁₀Alkyl;

(d) OR ₁₀, wherein said R ₁₀Be selected from no group, hydrogen, C ₂-C ₁₀Acyl group, C ₁-C ₁₀Alkyl and SO ₃H;

(e) C ₅-C ₇Glycosyl;

(f) C ₃-C ₈Cycloalkyl, it randomly is selected from following substituent group and is replaced: hydrogen, OH, NH ₂, SH, OSO ₃H and OPO ₃H ₂

(g) SR ₁₁, R wherein ₁₁Be selected from hydrogen, C ₁-C ₁₀Alkyl and O ₃H;

(h) randomly be selected from hydrogen, OR ₁₀, NHR ₇, NR ₈R ₉And SR ₁₁The C that replaces of substituent group ₁-C ₁₀Alkyl; With

(i) C ₃-C ₈Cycloalkyl, it randomly is selected from following substituent group and is replaced: hydrogen, OR ₁₀, NHR ₇, NR ₈R ₉And SR ₁₁,

Condition is that this chemical compound is not a myo-inositol.

The present invention provides also that a kind of prevention experimenter abnormal protein is folding, abnormal protein gathers, amyloid forms, has deposited, gathers or retain or the interactional method of amyloid lipid, and it comprises the chemical compound that is selected from following structure to the medical effective dose of described experimenter's administration:

(a) hydrogen atom;

(e) C ₅-C ₇Glycosyl;

Condition is that this chemical compound is not a myo-inositol.

The present invention also provides a kind of protein dissociation of the experimenter's of making abnormal accumulation and/or has made amyloid fibril or the amyloid dissolving or the disruptive method of pre-formation or pre-deposition, and it comprises the chemical compound that is selected from following structure to described experimenter's administration medicine effective dose:

(a) hydrogen atom;

(e) C ₅-C ₇Glycosyl;

Condition is that this chemical compound is not a myo-inositol.

The method that the present invention also provides the unusual folding or protein that gathers of a kind of experimenter of diagnosis and/or amyloid fibril or amyloid to exist, it comprises: (a) to described experimenter's administration radioactive compound or labelling the chemical compound that is allowing the material of emission detectable signal under described chemical compound and unusual folding or the protein that gathers and/or fibril or the bonded condition of amyloid of q.s, if the folding unusually or protein that gathers and/or fibril or amyloid exist; (b) detect from unusual folding or the protein that gathers and/or the radioactivity or the signal of fibril or the bonded chemical compound of amyloid, diagnose described experimenter unusual folding or the protein that gathers and/or the existence of amyloid fibril or amyloid thus, wherein said chemical compound has following structure:

(a) hydrogen atom;

(e) C ₅-C ₇Glycosyl;

Condition is that this chemical compound is not a myo-inositol.

The method that the present invention also provides the unusual folding or protein that gathers of a kind of experimenter of diagnosis and/or amyloid fibril or amyloid to exist, it comprises: (a) collect from described experimenter's sample; (b) made described sample and radioactive compound or labelling and contacted, if protein and/or amyloid fibril or amyloid existence folding unusually or that gather at the chemical compound that allows the material of emission detectable signal under described chemical compound and unusual folding or the protein that gathers and/or amyloid fibril or the bonded condition of amyloid; (c) detect from unusual folding or the protein that gathers and/or the radioactivity or the signal of fibril or the bonded chemical compound of amyloid, diagnose described experimenter unusual folding or the protein that gathers and/or the existence of amyloid fibril or amyloid thus, wherein said chemical compound has following structure:

(a) hydrogen atom;

(e) C ₅-C ₇Glycosyl;

Condition is that this chemical compound is not a myo-inositol.

Brief description

Figure 1A has provided the structure of myo-inositol, table-inositol and Cocositol, and Figure 1B-1H has shown the georeferencing memory models of the Mo Lisi water maze test that carries out at the TgCRND8 mice simultaneously.Myo-inositol treatment change cognitive function (1B).When 6 monthly ages, untreated TgCRND8 (n=10) demonstrates cognitive impairment (every group of n=10, not treatment group is than treatment group p＜0.02) with respect to non--Tg matched group (1C) and Cocositol (1D) treatment mice.The performance of the TgCRND8 mice of table-inositol treatment is compared still damage to some extent with the brood mice of non--Tg (1E), but the performance close (1F) of the performance of Cocositol TgCRND8 mice and non--brood mice of Tg.The behavior of non--brood mice of Tg is not subjected to the influence of table-inositol (1G) or Cocositol (1H) treatment.Vertical bar is represented standard error.

The speckle of the TgCRND8 of table-inositol and Cocositol treatment stockpiled and astrocytosis when Fig. 2 A-2I showed for 6 monthly ages.Control animal has high speckle load and astrocytosis in hippocampus (2A) and cerebral cortex (2B).Higher enlarged drawing confirm the star-shaped glial cell activation not only with speckle load relevant (2C).The treatment of table-inositol stockpiles the effect of moderate to amyloid, and has reduced astrocytosis (2D, 2E and 2F).Cocositol treatment has reduced significantly that amyloid stockpiles and gliosis (2G, 2H and 2I).Higher enlarged drawing demonstrates Cocositol treatment mice and has less average speckle size (2I).Also use anti--A β antibody to identify that speckle stockpiles with anti--GFAP antibody labeling star-shaped glial cell.The calibration lines are 450 microns (A, B, D, E, G, H) and 94 microns (C, F, I).

Fig. 3 A-3D has shown that the degree of A beta substance 1-42,1-40 and 1-38 and APP progress (3B) is difficult to differentiation in the TgCRND8 mice of contrast and treatment.In serial sagittal plane section, the blood vessel amyloid is stockpiled quantification with treatment and untreated TgCRND8 mice.The TgCRND8 mice has significant blood vessel amyloid and stockpiles, and this is with little relevant with medium sized blood vessel, the loading of the TgCRND8 mice of Cocositol treatment descend (3A).With do not treat with show-the TgCRND8 mice of inositol treatment compares (3C), the Cocositol treatment has reduced total blood vessel loading significantly.Obviously reduce (3D) of average speckle size illustrated that Cocositol has reduced plaque deposition.

Fig. 4 has shown the georeferencing memory models that uses the Mo Lisi water maze, and water is to the effect of TgCRND8 and non-Tg mice cognitive function in 3 days test examples.

Fig. 5 has shown the georeferencing memory models that uses the Mo Lisi water maze, and Cocositol is to the effect of TgCRND8 and non-Tg mice cognitive function in 3 days test examples.

Fig. 6 showed the georeferencing memory models that uses the Mo Lisi water maze, tested the effect of example invading the exterior-inositol to TgCRND8 and non-Tg mice cognitive function at 3 days.

Fig. 7 has shown the georeferencing memory models that uses the Mo Lisi water maze, and myo-inositol is to the effect of TgCRND8 and non-Tg mice cognitive function in 3 days test examples.

Fig. 8 has shown the georeferencing memory models that uses the Mo Lisi water maze, and Cocositol, table-inositol and myo-inositol compare to the effect of TgCRND8 cognitive function (learning period and memory test) and with wild-type mice in 3 days test examples.

Fig. 9 has shown the brain area percent that the mice of untreated TgCRND8 mice and Cocositol, table-inositol or myo-inositol treatment is relatively covered by speckle.

Figure 10 A and 10B have shown the survival rate relatively (10A) or compare (10B) with the survival rate of the TgCRND8 mice of Cocositol treatment of survival rate and the TgCRND8 mice of table-inositol or myo-inositol treatment of the TgCRND8 mice of water treatment.

Figure 11 A-D shown the TgCRND8 mice at 6 monthly ages through the result of the georeferencing memory models of mannitol treatment or untreated Mo Lisi water maze test (A, B).The TgCRND8 mice of mannitol treatment and untreated TgCRND8 mice do not have tangible difference (p=0.89; A).The performance of the TgCRND8 mice of mannitol treatment obviously is different from the brood mice (p=0.05 of non--Tg of mannitol treatment; B).Use the quantitative image analysis method analyzing proteins amount of stockpiling (C).When stockpiling quantitative determination as total speckle with the speckle counting, the TgCRND8 mice of mannitol treatment and untreated TgCRND8 mice as broad as long (p=0.87).Vertical bar is represented standard error.Drawn the Kaplan-Meier accumulation survival rate (D) of mannitol treatment or untreated TgCRND8 mice.Drawing between two treated animals (every group of n=35) by the Tarone-Ware statistical test does not have significant difference, p=0.87.

Figure 12 A and B have shown the treatment result of study of the georeferencing memory test that has carried out 3 days test examples.The performance of the TgCRND8 mice of Cocositol treatment can be comparable to the brood mice (p=0.38 of non--Tg of Cocositol treatment; A).As one man, treat after 2 months, the TgCRND8 mice of Cocositol treatment still with the as broad as long (p=0.67 of the brood mice of non--Tg; B).

Figure 13 A and B have shown to the TgCRND8 at 5 monthly ages mice and have been administered once the Cocositol of various dosage every day after 1 month, the A β content among the CNS.Under all dosage, soluble A β 42 content descend and obviously different with untreated matched group (A).On the contrary, under all conditions, insoluble A β 42 not significant different (B).Vertical bar is represented standard error.

Figure 14 is administered once the Cocositol of various dosage every day after 1 month to the TgCRND8 mice, analyzes the content of brain A β 40.Under all proof loads, different between solvable A β 40 content (A) of the TgCRND8 mice that does not detect in untreated and Cocositol treatment and soluble A β 40 content (B).

Figure 15 has shown with the brood mice of non--transgenic and has compared, the cognitive function of the TgCRND8 mice at 6 monthly ages of different-inositol treatment.

Figure 16 A-D has shown the mice of Cocositol treatment when 2 monthly ages, and the speckle amount of stockpiling in TgPS1 * APP mice decreases.Control animal has high speckle load in hippocampus (A) and cerebral cortex (B).The Cocositol treatment has reduced amyloid significantly and has stockpiled (C, D).Use anti--A β antibody (brown) to identify that speckle stockpiles.The calibration lines are 300 μ m.

Figure 17 A-C has shown quantitative that speckle stockpiles in Cocositol treatment back TgPS1 * APP mice.The speckle brain covers percent (A), average speckle size (B) and speckle counting (C) significantly to be reduced.Vertical bar is represented standard error.

Detailed Description Of The Invention

The invention discloses that some inositol stereoisomer has with amyloid related The novelty that the treatment of disease such as Alzheimer's is relevant, unpredictalbe and unexpected Character.

Be surprised to find, some stereoisomer of inositol and relevant compound can be blocked Carrying out property cognitive decline and the brain amyloid plaque pathology of A β-induce, and deposit at A β The newborn stage, can improve when giving the transgene mouse model of people's Alzheimer's with it Survival rate.

Different such as some (being not whole) inositol solids of above-mentioned disclosed, the former Notes of Key Data The structure body may act on to some extent to the amyloid aggregation of the neuronal cell of in vitro culture (people such as McLaurin, J.Biol.Chem.275 (24): 18495-18502 (2000)). That The stereoisomer that a little investigation do not provide any method to come predict what to study (myo-inositol, Epi-cyclohexanehexol, scyllitol and chiro-cyclohexanehexol) will have this effect, do not predict any yet Other will have this effect. And whether those researchs can not predict any flesh Alkanol isomer can act on amyloid beta deposition, cognitive defect or life-span in vivo to some extent. This Following uncertain result has been described in invention: the animal of the disease relevant with amyloid In the model, only there are some inositol stereoisomer, particularly scyllitol and different-inositol to reduce starch Sample albuminous plasue burden is improved cognition and is increased the life-span, and other the isomers that is studied is not then Has this effect.

Some inositol stereoisomer (for example epi-cyclohexanehexol and shark flesh are also only pointed out in research in the past Alcohol) can suppress external newborn amyloid aggregation. The invention describes following unpredictable The result: scyllitol suppresses established brain amyloid beta deposition, and in the brain of living also Like this. Previously disclosed vitro data is to this hint not, and these data are thought only effect In some neuronal cell type of cultivating, but not the complex organization of the brain of living, and only point out flesh Alcohol can suppress newborn and assemble, thus with established disease independent.

Vitro data in the past also points out administration epi-cyclohexanehexol and scyllitol also to affect amyloid A β 40 content and A β 42 content. Dose study has disclosed following unexpected in the body of the present invention The result: administration is different-inositol or scyllitol especially reduce A β 42 content, however to insoluble A β 42 With solvable or insoluble not impact of A β 40 content.

Investigation of the present invention demonstrates the variation of the active and inflammation of neuroglia, this be novelty and Surprising, and can not by data prediction in the previously disclosed body to.

Investigation of the present invention confirms that scyllitol improves the life-span of transgenic models animal, and this also is new Grain husk and surprising, because there is not the medicine of Alzheimer's to demonstrate in the body in the past Can increase survival rate and life-saving.

Preferably, compound of the present invention is 1,2,3,4,5,6-inositol, more preferably be selected from suitable-, Table-, different-, sticking-, new-, shark-, D-chirality-and L-chiro-cyclohexanehexol.

Also preferred

compound

1,2,3,4,5-quercitol (quercitol), more preferably be selected from table-, vibo-, Shark-, different-, Ta Luo-, gala-, suitable-, sticking-, new-, former-quercitol and enantiomter thereof.

Also preferred those be selected from cyclohexanetetraol, phloroglucite, cyclohexanetetraol stereoisomer, The mapping of the stereoisomer of phloroglucite, the enantiomter of cyclohexanetetraol and phloroglucite is different The compound of structure body.

These compounds can also be penta hydroxy group cyclohexanone or its stereoisomer or enantiomerism Body.

Also preferred, these compounds are to be selected from scyllitol single ketones, L-chiro-cyclohexanehexol single ketones-1 and L-The inosose of epi-cyclohexanehexol single ketones.

Also preferred, these compounds are that trihydroxy cyclohexanone or its stereoisomer or mapping are different The structure body. More preferably (-)-1-deoxidation-scyllitol single ketones.

Also preferred, these compounds are that penta hydroxy group cyclohexanone (inosose) or its solid are different Structure body or enantiomter more preferably are selected from scyllitol single ketones, L-chiro-cyclohexanehexol single ketones-1 and L-The epi-cyclohexanehexol single ketones.

Randomly, these compounds are trihydroxy cyclohexanone or its stereoisomer or enantiomerism Body is such as (-)-1-deoxidation-scyllitol single ketones.

Also preferred, these compounds are O-monomethyl-inositol or its stereoisomer or right Reflect isomers, more preferably be selected from D-pine camphor, L-bornesitol and D-bornesitol.

In addition, these compounds can be selected from an amino quercitol (inosamine), diaminourea Cyclohexanetetraol (inositol diamines), diaminourea phloroglucite, its stereoisomer and mapping thereof Isomers and its officinal salt, for example L-new-inosamine, D, L-epi-cyclohexanehexol amine-2, strepto-Amine and deoxystreptamine.

Also preferred, these compounds are single sulfydryl-quercitol or its stereoisomer or mapping Isomers, more preferably 1L-1-deoxidation-1-sulfydryl-8-O-methyl-chiro-cyclohexanehexol.

Most preferred compound of the present invention is different-inositol and scyllitol, wherein scyllitol most preferably. As implied above, inositol stereoisomer of the present invention do not comprise myo-inositol and do not comprise table-Inositol.

Even administration after the amyloid pathology has formed the several months, these compounds also can Effectively reversing brain A β gathers and the amyloid pathology.

Therefore, find these compounds be useful on the treatment or the prevention experimenter in protein folding Or gather disorder or amyloid and form, deposit, gather or retain relevant maincenter or outer The illness of all nervous systems or general organ. Find that also these compounds can be used for preventing tested Person's abnormal protein is folding, abnormal protein gathers, amyloid forms, deposit, gather Or retain or the interaction of amyloid lipid, and the albumen that makes experimenter's abnormal accumulation Matter is dissociated and/or the amyloid fibrillation of pre-formation or pre-deposition or amyloid is dissolved Or destroy.

Preferably, the illness of described central or peripheral nervous system or general organ causes albumen Matter, protein fragments and peptide are with the form precipitation of beta-pleated sheet and/or fibrillation and/or aggregation. More preferably, the illness of described central or peripheral nervous system or general organ is selected from: A Erci The sea Mo's disease, senilism and involution form; Amyloid blood vessel disease; Mild cognitive goes down; With Ah The dementia that the Er Cihai Mo's disease is relevant; The tau disease; The alpha-synapse nucleoprotein disease; Parkinson's; Flesh The atrophic lateral spinal sclerosis; Motor neuron disease; Spastic paraplegia (paraplagia); Prosperous Prolong a disease; Spinocebellar ataxia; Friedreich ataxia; With in the cell and/or Neuron internal protein and polyglutamine, polyalanine or by three in the corresponding gene or four nuclears The expansion of the pathology of thuja acid unit and the relevant neurodegeneration disease of gathering of other repeat body of causing Sick; Cranial vascular disease; Down syndrome; Create with the head that amyloid beta after the wound gathers Hinder; The disease relevant with prion; Familial Britain dementia; Familial Denmark dementia; With The alzheimer's disease of spasmodic ataxia; Britain's type cerebral amyloid angiopathy; With spasm The ataxic alzheimer's disease of property; The danish type cerebral amyloid angiopathy; Having neurofilament presses down The familial encephalopathia of albumen inclusion body (FENIB); Amyloid polyneuropathy; By amyloid The inclusion body myositis that peptide causes; The familial amyloidosis Finnish type; With the Huppert's disease phase The SA that closes; Familial Mediterranean fever; Chronic infection and inflammation; And with The relevant type ii diabetes of IAPP (IAPP).

Also preferred, the dementia relevant with Alzheimer's is that vascular or Alzheimer are silly Slow-witted, and the tau disease be selected from argentaffine grannles dementia (argyrophilic grain dementia), Cortex substrate degeneration, dementia pugilistica, with dispersivity NFT, the companion of calcification The frontotemporal dementia of parkinson's syndrome, disease, the Halle Wo Deng-Shi Paci relevant with prion are arranged Disease, myotonia atrophica, C type Niemann-Pick disease, have the non-of NFT Ba Jinsen syndrome, prion protein brain after Guam motor neuron disease, pik disease, the encephalitis Amyloid angiopathy, progressive subcortical gliosis, stein-leventhal syndrome, subacute Sclerosing panencephalitis and only entanglement type dementia (tangle only dementia).

Also preferred, the alpha-synapse nucleoprotein disease be selected from the corps ronds with centrality multilayer dementia, MSA, shy-Drager syndrome, corpus straitum with Deiter's cells matter inclusion are black Qualitative change, olvopontocerebellar atrophy, has neurodegeneration, sense of smell that I type brain iron gathers Unusually and amyotrophic lateral sclerosis.

Also preferred, motor neuron disease and neurofilament and/or superoxides thread or the gathering shape Mutase albumen is relevant, and the function of spastic paraplegia and chaperone and/or three A albumen lacks Decrease relevantly, and spinocebellar ataxia is DRPLA or Ma-Yue disease.

Also preferred, the disease relevant with prion is selected from Creutzfeldt-Jakob disease, Gerstmann-

-Scheinker disease and mutation Creutz Fil spy-Jacob Disease and amyloid polyneuropathy are old amyloid polyneuropathy or SA.

More preferably, the illness of described central or peripheral nervous system or general organ is to comprise house The Parkinson's of family's property or non-familial. Most preferably, described central or peripheral nervous system or complete The illness of body sexual organ is Alzheimer's.

Preferably, with The compounds of this invention with the about 1g/kg experimenter's body weight of about 1mg-, preferred 1mg-About 200mg/kg experimenter's body weight, more preferably from about the about 100mg/kg experimenter's body weight of 10mg-, Most preferably from about the dosage of the about 70mg/kg experimenter's body weight of 30mg-is to experimenter's administration. Administration can Finish by all the whole bag of tricks described as follows: oral (oral administration pills, oral liquid or suspension), In the intravenous, intramuscular, peritonaeum, intracutaneous, in skin, subcutaneous, nose, hypogloeeis, rectal suppository Or suck oral administration most preferably wherein. But the multiple time of The compounds of this invention interval carry out to Medicine, for example once a day, twice of every day, weekly, January once or continue medication.

Preferably, The compounds of this invention and other therapeutic agent administering drug combinations, these therapeutic agents for example are Beta-secretase inhibitor, inhibitors of gamma-secretase (APP-specificity or non-specific), ε-minute Secrete enzyme inhibitor (APP-specificity or non-specific), β-sheet gathering/microfibre generation/ADDL Generate (for example Alzhemed) other inhibitor, nmda antagonist (for example Memantine), Nonsteroidal anti-inflammatory compound (for example brufen, Celebrex), antioxidant (vitamin for example E), hormone (for example estrogen), nutritional agents and food supplement (for example ginkgo); Acetyl Anticholinesterase (for example donezepil), poison

Alkali activator (AF102B (west for example Dimension Merrill Lynch, EVOXAC), AF150 (S) and AF267B), antipsychotic drug (fluorine for example Send pyridine alcohol, Clozapine, Olanzapine); Antidepressants comprise tricyclic antidepressants and thrombocytin Reuptake inhibitor (for example Sertraline and Citalopram Hbr), based on just regulating neprilysin Gene therapy and/or the medicine of (enzyme of a kind of A β that degrades); Based on just regulating the insulin degraded The immunity of the gene therapy of enzyme (enzyme of a kind of A of degraded β) and/or medicine, vaccine, A β is controlled Treat and antibody (for example ELAN AN-1792), statins and other cholesterol-lowering drug (for example Lovastatin and Simvastatin), stem cell and other therapeutic agent based on cell, phosphorus The kinases of acidifying TAU (CDK5, GSK3 α, GSK3 β) inhibitor (chlorine for example Change lithium) or (GSK3 α, GSK3 β, Rho/ROCK swash to regulate the kinases that A β generates Enzyme) inhibitor (for example lithium chloride and brufen).

We think that these other therapeutic agents work by different mechanism, and to the present invention Has summation action/synergy. In addition, many other therapeutic agents may have based on mechanism Effect and/or other side effect, this meeting dose limitation or tolerance at this moment can be independent with them Administration.

Because The compounds of this invention has hereinafter discussed in detail in vivo in conjunction with starch The ability of sample albumen is diagnosed tested so The compounds of this invention also is useful on following method Unusual folding or the protein that gathers and/or amyloid fibrils or amyloid deposits among the person , the method comprises: to described experimenter's administration radioactive compound or mark q.s Allowing described compound and protein and/or fibrillation or amyloid egg unusually folding or that gather If the compound of the material of emission detectable signal under the condition of white combination is unusually folding or long-pending The words that poly-protein and/or fibrillation or amyloid exist; With detect from unusual folding The radioactivity of the compound of folded or the protein that gathers and/or fibrillation or amyloid combination or Signal is diagnosed protein and/or amyloid fibrillation or shallow lake unusually folding or that gather thus The existence of powder sample albumen.

In addition, collect suspection from the experimenter and contain protein and/or starch unusually folding or that gather The sample of sample fibrillation or amyloid, and make described sample and radioactive compound or mark Allowing described compound and unusual folding or the protein and/or the amyloid fibril that gather The compound contact of the material of emission detectable signal under the condition of dimension or amyloid combination, If the unusually folding or protein that gathers and/or amyloid fibrillation or amyloid are deposited Words; And detect subsequently from unusual folding or protein and/or fibrillation or the shallow lake gathered Radioactivity or the signal of the protein bound compound of powder sample diagnose described experimenter unusually to roll over thus The protein of folding or gathering and/or the existence of amyloid fibrillation or amyloid.

Preferably, described detectable signal is fluorescence signal or enzyme linked immunosorbent assay (ELISA) signal, with And described sample is whole blood (comprising all cell components) or blood plasma.

Shown in hereinafter, in the transgene mouse model of Alzheimer's, in " symptom Late period before taking place " stage, mouse occur before obvious cognitive defect and the amyloidotic neuropathy reason, Give The compounds of this invention, can eliminate that IC A β gathers, the brain amyloid plaque Deposition and cognitive decline. In addition, even cognitive defect and amyloid plaque neuropathy are occurring Give these compounds after the reason, they also can reverse amyloid beta deposition and nerve effectively Pathology. Importantly, regulate the set of A beta monomers according to these compounds and be neurovirulent widow Polymers and/or fibriilar ability are guided out reasonably by these function of chemical compound mechanism and establish Meter.

Other advantage of The compounds of this invention comprises that they pass through two kinds of known transporters and passive Diffusion is transported to CNS, thereby good CNS bioavilability is provided. The second, these Compound is metabolised to glucose. The 3rd, as a class material, these compounds have usually Hypotoxicity spectrum, and for different purposes was given in them some in the past to the mankind Medicine.

Embodiment 1-makes up the method for Alzheimers mouse model and administration The compounds of this invention

As described in people such as Janus (Nature 408:979-982 (2000)), the TgCRND8 mice is the strong catastrophic model of Alzheimer.They are expressing human amyloid precursor protein (APP695) transgenic under the regulation and control of the hamster,syrian of C3H/B6 outbreeding background and Protein virus promoter.People APP695 transgenic has two catastrophe points, and it causes the mankind to suffer from AD (K670N/M671L and V717F).From about 3 monthly ages, carrying out property space learning defective appears in the TgCRND8 mice, and follow brain A β content to raise and the outer amyloid plaque quantity increase of brain cell, these all similar (people such as C.Janus, Nature 408:979-982 (2000)) to situation about arriving seen in the human brain of suffering from AD.

With age and sex-matched TgCRND8 mice group and not genetically modified brood mice group (every group of n=35) or do not treat, or begin by following amount administration The compounds of this invention with 30mg/ days/Mus from about 6 all ages.When 4 monthly ages and 6 monthly ages, measure cognitive function, brain A β content, brain pathology and the survival rate of these mices subsequently.

The Prevention Research method

Mice-give test group TgCRND8 mouse feeding myo-inositol, table-inositol and Cocositol with the amount in 30mg/ Mus/sky.6 weeks during ages two groups enter research and analysis result during at 4 monthly ages and 6 monthly ages.Behavior in monitoring body weight, fur feature and the cage.The animal of all experimental evidence Canada committee is taken care of guide and carries out.

4 tests are carried out in performance testing-after the pre-training of non-space, mice was carried out the orientation discrimination training 5 days every day.Use medicine or genotype and training period to analyze behavioral data as the mixed model of the factor variance analysis (ANOVA) of replication factor.

Brain amyloid burden-taking-up brain and a brain hemisphere are fixed in 4% paraformaldehyde and along central sagittal plane (mid saggital plane) imbeds paraffin.In order to produce group system section at random uniformly, the whole hemisphere of crosscut is collected 5 μ m serial section.The section group of 50mm is used for analyzing (10-14 sheet/group) at interval.Carry out identifying speckle after the antigen retrieval with formic acid, and with it with former anti-amyloid beta antibodies (Dako M-0872), use secondary antigen (Dako StreptABCcomplex/horseradish kit) to hatch then.End-product carries out DAB through hematoxylin and can observe dying.Use the Leco IA-3001 image analysis software evaluation starch sample albuminous plasue that is connected with Hitachi KIP-MU ccd video camera with the Lycra microscope to bear.Analyze the blood vessel burden similarly, and use the detacher measurement to be attacked the diameter of blood vessel.

Blood plasma and brain A β content-in the sucrose buffer is brain hemisphere sample homogenization, and then or adding 0.4% diethylamine/100mM NaCl is used to dissolve A β content, or adding cold formic acid is used to separate total A β.After the neutralization, dilute sample also uses the test kit (BIOSOURCE International) that is purchased to analyze A β 40 and A β 42.Each hemisphere is analyzed three times, the record mean+SD.Carry out the plaque of protein point analysis to analyze the A beta substance in the enterprising enforcement of all parts with urea gel.The chemiluminescence (Amersham) of using 6E10 (BIOSOURCEInternational) and strengthening detects A β.

APP-in the analysis brain is with the homogenate in 20mM Tris pH7.4,0.25M sucrose, 1mM EDTA and 1mM EGTA and protease inhibitor mixed liquor of mouse brain hemisphere sample, mix and with 109 the 000Xg rotation with 0.4%DEA (diethylamine)/100mM NaCl.Use the analytically APPs content in the clear liquid of mAb 22C11 by Western blotting, use mAb C1/6.1 to analyze flaky precipitate to measure the APP whole protein.

Quantitatively gliosis-from the paraformaldehyde of treatment and control mice fix and refrigerated hemisphere on select evenly spaced five sagittal slices at random.To cut into slices with anti-Mus GFAP IgG2a (Dako; 1:50 dilution) comes immune labeled star-shaped glial cell and with anti-Mus CD68IgG2b (Dako; The 1:50 dilution) comes immune labeled microgliacyte.Use is installed in Coolsnap digital camera (Photometrics, Tuscon, Arizona) the collection digital picture on the Zeiss Axioscope 2 Plus microscopes.Use Openlab 3.08 image softwares (Improvision, Lexington MA) analysis image.

The survival rate investigation-by Kaplan-Meier technology evaluation survival odds, computational analysis survival odds when death occurring, thus make its suitable small sample amount.Each treatment group uses 35 mices to be used for the viability analysis.Use Tarone-Ware to detect the comparison of putting down in writing between each treatment group.

The prevention of the cognitive disappearance of embodiment 2-

Use the georeferencing memory models of Mo Lisi water maze, use the cognitive function of 5 days test example assessment TgCRND8 mices.(untreated with treatment, table-inositol or Cocositol) and genotype (TgCRND8 and non--Tg) as " between the experimenter " factor, use blended variance analysis (ANOVA) model analyze by the data that obtain with untreated TgCRND8 mice of treatment and by treatment with untreated non--data that mice obtains that Tg is brood are (for all combinations, n=10).Through the performance of the TgCRND8 mice of table-inositol or Cocositol treatment significantly than good (p＜0.02 of untreated TgCRND8 mice; Fig. 1 C and D).With treatment with untreated non--the brood mice of Tg compares, in first three day of training, the TgCRND8 mice that table-inositol is treated shows slower a little learning curve.But, after training 4 days, no statistical difference (Fig. 2 E) between the TgCRND8 mice of table-inositol treatment and the brood mice of its non--Tg.On the contrary, in all these days, indifference between the TgCRND8 mice of Cocositol treatment and the brood mice of non--Tg.Therefore, these two kinds of stereoisomers all suppress the development of cognitive defect, and in fact, and the cognitive disappearance of Cocositol prevention has reached a kind of like this degree of indifference between the TgCRND8 mice that makes the Cocositol treatment and the normal mouse.This improved performance is not owing to the nonspecific action to behavior system, motor system or consciousness system, because table-inositol and Cocositol treatment are to the not effect (Fig. 2 G and 2H) of performance of non--Tg mice.This improved performance neither be owing to nutrition or heat effect, because do not having difference aspect body weight, activity and the fur condition between treatment group and the not treatment group.In addition, with mannitol (a kind of sugar of similar molecular weight) treatment behavior is not acted on.The influence of sex not significantly (p=0.85) between any treatment group.

The minimizing of embodiment 3-brain A β burden and amyloidotic neuropathy reason

During 4 monthly ages, untreated TgCRND8 mice gives expression to a large amount of A β 40 and A β 42 (table 1) mice.When 4 monthly ages, as described in embodiment 1, (solvable and soluble storehouse has reduced 43 ± 2% to A β 40 content of table-inositol treatment group; P≤0.05) and A β 42 content (solvable storehouse has reduced 69%, p=0.005; Soluble storehouse has reduced 28%, p=0.02) has all reduced.But these improve not lasting, when 6 monthly ages brain A β content be elevated to do not treating the TgCRND8 mice in the similar content (table 1) observed.

On the contrary, when 4 monthly ages, total brain A β 40 of Cocositol treatment group 62% (p=0.0002) and total brain A β 42 22% (p=0.0096 that descended that descended; Table 1).When 6 monthly ages, to compare with untreated TgCRND8 mice, the Cocositol treatment causes A β 40 content to reduce by 32% (p=0.04) and A β 40 content have reduced by 20% (p=0.02).

Because detected A β lowering of concentration after the inositol treatment may be taken place to change and cause, so detected the A β-β content (table 1) in the blood plasma when 4 monthly ages and 6 monthly ages by the excessive inflow blood plasma of A β.The TgCRND8 mice had high plasma A β concentration and keep constant when 6 monthly age when 4 monthly ages, even when CNS speckle loading still rose at 6 monthly ages (table 1).Do not compare with treating the TgCRND8 mice, table-inositol or Cocositol are treated plasma A β content all without any effect (p=0.89).To this observation phenomenon the most economical explanation is that inositol has optionally changed the A betaization among the CNS, but to β-or gamma-secretase activity or A β be scavenged into the not influence of normal mechanism of blood plasma.But for two reasons, this observation phenomenon is significant.At first, in untreated patient AD, detect among blood plasma and the CSF A β content usually and reduce as clinical course progress (people such as Mayeux, Ann.Neurol 46,412,2001).Secondly, taken place the powerful antibody reaction and obviously the AN1792 immune Research patient of clinical response do not have plasma A β-β content people such as (, Neuron 38,5472003) Hock of change.Therefore, these results show in order to obtain the unnecessary change plasma A of effective therapeutic outcome β content.

In order to confirm of expression or the proteolysis processing not influence of inositol stereoisomer, in inositol treatment and untreated TgCRND8 mouse brain, detected the content of APP whole protein, sAPP-α and various A beta substances to APP.Data consistent (people such as McLaurin with report before us, Nat.Med.8,1263,2002), A β 42, A β 40 and A β 38 are the main matter (Fig. 3 A) in the TgCRND8 mouse brain, and no matter whether treat, the CNS content of the CNS content of immaturity and sophisticated glycolysis APP (Fig. 3 B) and sAPP-α all is difficult to distinguish.Integrating, these results show-inositol and Cocositol plays directly for A β oligomerization and optionally effect, but inoperative to APP processing.

The change of A β-β peptide loading is accompanied by the remarkable reduction (table 1 of speckle burden; Fig. 2 A-2I).The TgCRND8 mice of table-inositol treatment is not compared with treating the TgCRND8 mice, at 4 monthly ages but not during 6 monthly ages, average speckle diameter occurs significantly descending (being respectively 95 ± 4.3 μ m ²Contrast 136 ± 15 μ m ², p=0.04; 370 ± 9 μ m ²Contrast 423 ± 22 μ m ², p=0.06).In a single day these results show that when moderate A β content, table-inositol prevents A β oligomerization, but higher A β concentration occurs, table-inositol then can not suppress microfibre and generate.When 4 monthly ages, the average speckle diameter of Cocositol treatment group is by 136 ± 15 μ m ²Reduce to 103 ± 4 μ m ²(p=0.01).When 6 monthly ages, the A β peptide content of Cocositol treatment TgCRND8 mice group reduces, and is attended by speckle quantity minimizing 20% (p=0.005), and brain area minimizing 35% (p=0.015) and the average speckle size that are covered by speckle reduce (339 ± 10 μ m ²Contrast 423 ± 21 μ m ², p=0.009).By every test, these results confirm that after the Cocositol treatment, the speckle burden has descended.

The minimizing of embodiment 4-neuroglia activity and inflammation

Astroglia and microglial reaction are europathology feature (Irizarry etc., J Neuropathol ExpNeurol.56,965,1997 that people AD and all amyloid mouse models all have; K.D.Bornemann etc., Ann N Y Acad Sci.908,260,2000).Therefore, studied table-inositol and Cocositol astrocytosis and the outgrowth therapeutical effect of microglia (Fig. 3 A-3D) to the TgCRND8 mouse brain.A series of sagittal slices are come quantitatively with star-shaped glial cell label glial fibrillary acidic (GFAP) dyeing and with the percent of the brain area that astrocytosis was covered.The TgCRND8 mice has high basic astrocytosis (0.459 ± 0.048%) when 4 monthly ages, marginal increase (0.584 ± 0.089%) is arranged during to 6 monthly ages, and is not subject to plaque area (Fig. 2 A-2C).When 6 monthly ages, table-inositol is reduced to 0.388 ± 0.039% (p=0.04 with the proliferation response of astroglia; Fig. 2 D-F) on the other hand, Cocositol more effectively is reduced to 0.269 ± 0.028% (p=0.006 with the proliferation response of astroglia; Fig. 2 G-I).When with the age with sex-matchedly do not treat TgCRND8 mice (0.31 ± 0.01%; P＜0.001) when comparing, the TgCRND8 mice of Cocositol treatment has also weakened microglial activation (0.20 ± 0.008% brain area) significantly.But, confirmed that the microglia activation of the mice of table-inositol treatment when 6 monthly ages does not significantly reduce (0.248 ± 0.02%; P=NS).These data combine and show, Cocositol treatment minimizing A β among the CNS-inductive inflammatory reaction.

Embodiment 5-blood vessel amyloid loading

Alzheimer is characterised in that and has parenchyma and blood vessel amyloid beta deposition.In the TgCRND8 at 6 monthly ages mice, nearly 0.03% brain area and blood vessel are amyloid related.In the table-inositol treatment group at 6 monthly ages, do not observe the difference (Fig. 3 C) of blood vessel amyloid burden.On the contrary, the blood vessel amyloid of Cocositol treatment group burden has reduced (p=0.05) (Fig. 3 C) significantly, and amyloid beta deposition mainly is confined to less blood vessel, and diameter is less than 25m ²(56 ± 2% to 70 ± 8%, in the little blood vessel of untreated TgCRND8 mice).Compare with untreated mice, the average-size of the cerebrovascular speckle in the mice of Cocositol treatment obviously reduce (154 ± 16 to 363 ± 34, p=0.008; Fig. 3 D).

Embodiment 6-survival rate is improved

The TgCRND8 mice is 50% the 175th day survival rate, brings up to 72% (every group of n=35, Cocositol than matched group p＜0.02, Figure 10 B) after Cocositol treatment.Use the myo-inositol treatment not influence total survival rate (Figure 10 A) significantly.Controlled trial has confirmed that the growth of the survival rate of the mice that Cocositol is treated is not is the indirect action that the heat intake increases.Thereby, with Cocositol wild-type mice to be treated, it is to not effect of behavior in survival rate or other parameter such as body weight, coat condition or the cage.In addition, in body weight, coat condition or the cage of the TgCRND8 mice that inositol is treated, there is not difference between behavior and the untreated TgCRND8 mice.Carry out similar test with mannose (a kind of monosaccharide of similar molecular weight), it is to the also not effect of survival rate of TgCRND8 mice.

The treatment and the reverse of embodiment 7-amyloid beta deposition

In sum, Prevention Research has confirmed that Cocositol can suppress the amyloid beta deposition of parenchyma and blood vessel, thereby has improved the survival rate and the cognitive function of the TgCRND8 mouse model of Alzheimer.But, most of Alzheimer people probably only when symptom occurring and A β oligomerization, deposition, toxicity and speckle form when in CNS, having reached high level and just seek treatment.Therefore the TgCRND8 mice with 5 monthly ages begins to carry out the test of leading property.Compare with people's brain of suffering from AD, these mices have tangible A β and speckle stockpiles.

The treatment research method

Mice-give test group TgCRND8 mouse feeding myo-inositol, table-inositol and Cocositol with the amount in 30mg/ Mus/sky.One group begins to enter research and analysis result when 6 monthly ages when 5 monthly ages.Behavior in monitoring body weight, fur feature and the cage.The animal of all experimental evidence Canada committee is taken care of guide and carries out.

The survival rate investigation-by Kaplan-Meier technology evaluation survival odds, computational analysis survival odds when death occurring, thus make its suitable small sample amount.Each treatment group uses 35 mices to be used for survival rate analysis.Use the comparison between each treatment group of Tarone-Ware check record.

Performance testing-reverse research-without pre-training allows mice enter to have the Mo Lisi water maze test of hiding platform.Mice is accepted 6 training and reach 3 days every day.At the 4th day, from the pond, take platform away, and every mice is accepted one time 30 seconds swimming detection tests.In the end one day, once point out test to animal, to estimate swimming ability, the visual field and common cognitive power.In prompting test, platform is placed on and the flag of having tested used different signal area and labelling.Allow animal with finding platform in 60 seconds.Do not find the animal of platform to be not used in the final analysis of spatial memory.Use medicine or genotype and training period to analyze behavioral data as the mixed model of the factor variance analysis (ANOVA) of replication factor.

A brain amyloid burden-taking-up brain and brain hemisphere is fixed in 4% paraformaldehyde and along central sagittal plane (mid saggital plane) imbeds paraffin.In order to produce the section at random uniformly of a group system, the whole hemisphere of crosscut is to collect 5 μ m serial section.The section group of interval 50mm is used for analyzing (10-14 sheet/group).Carry out identifying speckle after the antigen retrieval with formic acid, and with it with former anti-amyloid beta antibodies (Dako M-0872), use secondary antigen (Dako StreptABCcomplex/horseradish kit) to hatch then.End-product carries out DAB through hematoxylin and can observe dying.Use the amount of load of the Leco IA-3001 image analysis software evaluation starch sample albuminous plasue that is connected with Hitachi KIP-MU ccd video camera with the Lycra microscope.

Blood plasma and brain A β content-in the sucrose buffer is brain hemisphere sample homogenization, and then or adding 0.4% diethylamine/100mM NaCl is used to dissolve A β content, or adding cold formic acid is used to separate total A β.After neutralization, dilute sample also uses the test kit (BIOSOURCE International) that is purchased to analyze A β 40 and A β 42.Each hemisphere is analyzed three times, record meansigma methods ± standard error of mean.

Result and significance-participation reverses the external sign that all animals of studying all survive and do not demonstrate worry or poisoning.Use the georeferencing memory models of Mo Lisi water maze, use the cognitive function (Fig. 4-8) of 3 days test example assessment TgCRND8 mices.(use myo-inositol, table-inositol or Cocositol with treatment, treatment) and genotype (TgCRND8 with non--Tg) as " between the experimenter " factor, the mixed model of user's difference analysis (ANOVA) analyze by the data that obtain with untreated TgCRND8 mice of treatment and by treat with untreated non--data (all combination n=10) of mice acquisition that Tg is brood.Compare with the brood mice of wild type, the ability of TgCRND8 mice obviously weakens (Fig. 4).On the contrary, in all these days, indifference (p=0.38 between the TgCRND8 mice of Cocositol treatment and the brood mice of non--Tg; Fig. 5).Compare the almost obviously different (p=0.07 of the TgCRND8 mice of table-inositol treatment with the brood mice of non--Tg of treatment; Fig. 6).Similarly, the also obviously different (p=0.05 of TgCRND8 mice of myo-inositol treatment with the brood mice of non--Tg of treatment; Fig. 7).This water maze test is during the learning period between the relatively treatment group, all mice behaviors similar " Fig. 8).On the contrary, indifference (Fig. 8) between Cocositol group and the non--Tg compatriot mice.Therefore, in fact, Cocositol reverses cognitive disappearance and has reached a kind of like this degree of indifference between the TgCRND8 mice that makes the Cocositol treatment and the normal mouse.This improved performance is not owing to the nonspecific action to behavior system, motor system or consciousness system, because table-inositol and Cocositol treatment are to the not effect of performance of non--Tg mice.This improved performance neither be owing to nutrition or heat effect, because do not have difference aspect body weight, activity and the fur condition between treatment and not treatment group.

Whether relevant with the minimizing of A β load for the cognition of determining to improve with the speckle amount of stockpiling, cerebral tissue has been carried out obduction.The corresponding change (Fig. 9 and table 2) of speckle amount of stockpiling and A β load has been followed in cognitive variation.The myo-inositol treatment is to speckle amount of stockpiling or the not influence (Fig. 9 and table 2) of A β load.The TgCRND8 mice of table-inositol treatment is compared with untreated TgCRND8 mice, and average speckle diameter does not reduce (Fig. 9) significantly, but A β load obviously reduces (table 2).These results show, when moderate A β content, table-inositol prevents A β oligomerization, but in case higher A β concentration occurs, table-inositol then can not suppress the microfibre generation fully.The speckle amount of stockpiling of Cocositol treatment group and A β load obviously reduce.By every test, these results confirm that the speckle amount of stockpiling has descended after the Cocositol treatment.These results are comparable to 6 months preventative research effect in effect aspect big or small, and have further supported the potentiality of Cocositol.

Because detected A β lowering of concentration after the inositol treatment may change take place owing to the excessive inflow blood plasma of A β, so we has detected the A β content (table 2) in the blood plasma.The TgCRND8 mice has high plasma A β concentration when 6 monthly ages.Compare with untreated TgCRND8 mice, myo-inositol, table-inositol or Cocositol are treated plasma A β content all without any effect (p=0.89).To this observation phenomenon the most economical explanation is that inositol has optionally changed the A betaization among the CNS, but to β-or gamma-secretase activity or A β be eliminated the not effect of normal mechanism that enters blood plasma.But for two reasons, this observation phenomenon is significant.At first, in untreated patient AD, detect A β content among blood plasma and the CSF usually and reduce progress as clinical course.Secondly, the plasma A β content that in the AN1792 immune Research patient that powerful antibody reaction and obvious clinical response take place, does not have change.Therefore, these results further show in order to obtain the unnecessary change plasma A of effective therapeutic outcome β content.

In sum, these data have disclosed in the transgene mouse model of Alzheimer, before significantly cognitive disappearance and amyloidotic neuropathy reason appearred in " presymptomatic late period " stage, mice, the Cocositol of selection can eliminate that IC A β gathers, brain amyloid plaque deposition and cognitive decline.In addition, even give Cocositol after cognitive disappearance and amyloid plaque europathology occurring, these chemical compounds also can reverse amyloid beta deposition, europathology and cognitive disappearance effectively.Therefore, these results show that Cocositol is effective in prevent disease with treating in the disease of having diagnosed patient's existence of suffering from AD.

Treatment researchs in two months of embodiment 8-Cocositol

In order to determine the more long-acting scope of Cocositol treatment disease, to the TgCRND8 at 5 monthly ages mouse feeding Cocositol or do not treat and reach 2 months (every group of n=10).With three days Mo Lisi water maze tests 7 months big are compared through the TgCRND8 mice of Cocositol treatment and the brood mice of non--Tg of untreated TgCRND8 mice and treatment.Use medicine and genotype to analyze behavioral data as the mixed model of the factor variance analysis (ANOVA) of experimenter's internal variable as variable between the experimenter and training period.Find out between the brood mice of non--Tg of 2 months TgCRND8 mice of Cocositol treatment and Cocositol treatment, do not having difference (Figure 12 B) by Cocositol treatment 1 month (Figure 12 A).For cognitive power and the pathology that will improve connect, A β 40 and A β 42 content (table 3) in the brain have been analyzed.After the Cocositol treatment, insoluble A β 40 and A β 42 content have all descended 20%.These results confirm that Cocositol acts on during disease progression lasting.

The content of the treatment reduction of table 3 inositol A β 40 and A β 42

Use the PLSD of Fisher to carry out variance analysis, ^*P＜0.05.

Embodiment 9-dosage is to the effect of the TgCRND8 mice pathological examination of suffering from disease

Dosage with 10mg/Kg, 30mg/Kg and 100mg/Kg is treated 5 months big TgCRND8 mices of Cocositol feeding soluble in water once a day or not.Treat and put to death animal after one month and analyze pathological examination.A β analysis on Content in the brain of all groups has been confirmed to compare with untreated TgCRND8 mice, and all drug doses all effectively reduce to identical degree soluble A β 42 content and (descend 20%, F _3,15=3.1, p=0.07; Figure 13 A).Single dosage analytical proof 10mg/Kg and 30mg/Kg dosage group be different from significantly and do not treat matched group (being respectively p=0.03 and p=0.02).Selected dosage does not have significant difference (F each other _2,11=0.6, p=0.57; Figure 13 A).Feeding dosage is for insoluble A β 42 (F _3,15=0.69, p=0.58; Figure 13 B) or soluble and insoluble A β 40 (be respectively F _3,15=0.04, p=0.99 and F _3,15=0.36, p=0.79; Figure 14 A and 14B) have no significant effect.

Embodiment 10-is different-and inositol is to the therapeutical effect of the TgCRND8 mice that suffers from disease

Whether also can effectively prevent and further develop and/or part reverses established AD-sample phenotype in order to assess different-inositol, Cai begin treatment TgCRND8 mice when being deferred to for 5 monthly ages.TgCRND8 mice group and the brood mice of non--transgenic are organized or treated 28 days with scyllitol, or do not treat.In these trials, the dosage of chemical compound carries out with the neuro chemistry assessment employing mode identical with above-mentioned therapeutic test with oral administration and behavior.

The TgCRND8 mice group of different-inositol treatment at 6 monthly ages is than the untreated TgCRND8 mice (F that puts up a good show significantly _1,13=0.45, p=0.05; Data do not provide).The cognitive performance of the TgCRND8 mice of the different-inositol treatment at 6 monthly ages also is different from the brood mice of non--transgenic significantly.This beneficial effect of inositol treatment is not owing to the nonspecific action to behavior system, motor system or consciousness system, because the cognitive performance that inositol is treated non--Tg mice does not act on (F _1,12=0.98; P=0.49).Treatment and the brain A β content of untreated TgCRND8 mice are analyzed, whether changed relevant (table 4) with A β with the behavior of determining to improve.Different-inositol treatment has reduced soluble A β 42 (descending 20%, p＜0.05), and it is similar that this effect and Cocositol are observed.Different-inositol does not change insoluble A β 42 or A β 40 (solvable and soluble storehouse) significantly.A kind of possible explanation that A β 42 reduces is the removing of periphery A β 42, causes the increase of plasma A β 42 subsequently.In different-inositol treatment back blood plasma in A β 42 content and the untreated TgCRND8 blood plasma content do not have difference (table 5).Consistent with other inositol stereoisomer, the influence of different-inositol treatment that these results have proved that plasma A β content is not subjected to.

Table 4. is different-inositol treatment reduction A β 42 content

Use Fisher ' s PLSD to carry out variance analysis, ^*P＜0.05.

Table 5 haemobiochemistry-Cocositol dose study

The treatment of embodiment 11-inositol does not influence hematochemistry

In order to get rid of any illeffects of inositol treatment, after 1 month blood has been carried out analyzing (table 5 and 6) at Cocositol and different-inositol treatment to hematochemistry and organ dysfunction.Between the treatment group or and untreated TgCRND8 mice between, gross protein, albumin, globulin, bilirubin, alkaline phosphatase, glucose, carbamide and creatinine all do not have notable difference.All content all dropped in the normal range of being measured by non--transgenic wild-type mice.Except haemolysis, also have jaundice and lipidemia also all normal.These results suggest are different-and inositol and Cocositol do not demonstrate deleterious effects to hematochemistry and organ dysfunction.

The treatment research of table 6 haemobiochemistry-1 month

Embodiment 12-is at the double transgenic mouse model of Cocositol to Alzheimer The pathological effect of prevention AD-sample among PS1 * APP

Tg PS1 * APP mice is the reinforcement model of Alzheimer, and it expresses the mutant human PS1 transgenic of two familial form sudden changes of coding (M146L and L286V) and the people APP transgenic of coding Indiana and Sweden's familial sudden change.Through 30-45 days ages, these animals were developed the expression and the amyloid beta deposition of strong brain A β level.In preventive trial, begin to treat TgPS1 * APP mice from wean, and when 2 monthly ages, estimate neuro pathology's effect (Figure 16 and 17) with Cocositol.Compare with untreated TgPS1 * APP mice, TgPS1 * APP the mice of Cocositol treatment demonstrates significant minimizing (brain area %=0.157 ± 0.007 ratio=.065 ± 0.016 that speckle covers, p＜0.001 in all tests of speckle burden when 2 monthly ages; Average speckle size=177 ± 8 μ m ²Than 149 ± 5 μ m ², p＜0.05; Speckle counting 3054 ± 324 to 1514 ± 510, p＜0.01; Figure 17).These results show that Cocositol has prevented amyloid beta deposition in two strong Alzheimer models.

The heat that embodiment 13-increases is taken in the influence to the TgCRND8 mice

For the heat of getting rid of increase is taken in or the contribution of nonspecific action, with the monosaccharide-mannitol treatment TgCRND8 mice of similar molecular weight.When 6 monthly ages, the TgCRND8 mice of mannitol treatment and untreated TgCRND8 mice do not have difference (Figure 11 A), and with the brood mice of non--Tg of mannitol treatment between significantly different (Figure 11 B) are arranged.Mannitol is to not effect of the behavior of non--Tg mice, be because the brood mice of non--Tg of mannitol treatment and untreated non--the Tg mice do not have difference.These results relevant with pathological study show that mannitol does not change the speckle load (Figure 11 C) of TgCRND8 mice.Detection to survival rate has simultaneously proved the survival rate not effect (Figure 11 D) of mannitol to the TgCRND8 mice.

Though invention has been described with its concrete embodiment, to those skilled in the art, numerous other changes and improvements and other purposes are conspicuous.Therefore, the present invention is not limited to the concrete disclosed content of this paper, only is subject to appended claims.

Claims

1. there is purposes in the reagent that uses in the method for unusual folding or the protein that gathers and/or amyloid fibril or amyloid in Cocositol in preparation in order to the diagnosis experimenter, and wherein said method comprises the steps:

(a) to described experimenter's administration radioactivity Cocositol or labelling q.s and Cocositol that allowing the material of emission detectable signal under described Cocositol and unusual folding or the protein that gathers and/or fibril or the bonded condition of amyloid, if the folding unusually or protein that gathers and/or fibril or amyloid exist; With

(b) detect from unusual folding or the protein that gathers and/or the signal radioactivity or the detectable signal of fibril or the bonded Cocositol of amyloid, diagnose unusual folding or the protein that gathers and/or the existence of amyloid fibril or amyloid in described experimenter thus.

2. the purposes of claim 1, wherein said detectable signal is the enzyme-linked immunosorbent assay signal.

3. the purposes of claim 1, wherein said detectable signal is a fluorescence signal.

4. the purposes of claim 1, wherein said detectable signal is a radiated signal.

5. claim 1,2,3 or 4 purposes, wherein said reagent is in order to alzheimer's disease diagnosis.