WO2007140974A1 - Matrine-3 utilisée comme biomarqueur de la maladie d'alzheimer - Google Patents

Matrine-3 utilisée comme biomarqueur de la maladie d'alzheimer Download PDF

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Publication number
WO2007140974A1
WO2007140974A1 PCT/EP2007/004947 EP2007004947W WO2007140974A1 WO 2007140974 A1 WO2007140974 A1 WO 2007140974A1 EP 2007004947 W EP2007004947 W EP 2007004947W WO 2007140974 A1 WO2007140974 A1 WO 2007140974A1
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protein
disease
alzheimer
matrin
peptide
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PCT/EP2007/004947
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English (en)
Inventor
Nikolaos Berntenis
Bernd Bohrmann
Andreas Guentert
Bernd Mueller
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F. Hoffmann-La Roche Ag
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Publication of WO2007140974A1 publication Critical patent/WO2007140974A1/fr

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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
    • G01N33/6893—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids related to diseases not provided for elsewhere
    • G01N33/6896—Neurological disorders, e.g. Alzheimer's disease
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2800/00—Detection or diagnosis of diseases
    • G01N2800/28—Neurological disorders
    • G01N2800/2814—Dementia; Cognitive disorders
    • G01N2800/2821—Alzheimer

Definitions

  • Matrin-3 as biomarker for Alzheimer's Disease
  • AD Alzheimer's disease
  • a ⁇ amyloid- protein
  • CSF cerebrospinal fluid
  • the CSF is, compared to the brain, more easily accessible and is in direct contact with the extracellular space of the central nervous system where biochemical changes in the brain could potentially be reflected. Therefore, a number of putative biological markers of AD have been suggested, like CSF total tau (Andreasen, N., Vanmechelen, E., Van de Voorde, A., Davidsson, P., Hesse, C, Tarvonen, S., Raiha, L, Sourander, L., Winblad, B., and Blennow, K.
  • AD (1996) J Biol Chem 271, 22908-22914) or a combination thereof (Shoji, M., Matsubara, E., Kanai, M., Watanabe, M., Nakamura, T., Tomidokoro, Y., Shizuka, M., Wakabayashi, K., Igeta, Y., Ikeda, Y., Mizushima, K., Amari, M., Ishiguro, K., Kawarabayasbi, T., Harigaya, Y., Okamoto, K., and Hirai, S. (1998) J Neurol Sci 158, 134-140). Yet, a definitive diagnosis of AD can still only be made by a postmortem assessment of brain neuropathology and there is clearly a need for a reliable biomarker for AD.
  • the present invention provides a method for assessing Alzheimer's Disease in vitro comprising measuring in a body fluid sample the level of Matrin-3 or a variant thereof, wherein a decreased level of Matrin-3 or a variant thereof is indicative that said individual suffers from Alzheimer's Disease.
  • the protein of interest may be detected in the cerebrospinal fluid or blood.
  • variants in this context relates to proteins or peptides substantially similar to said proteins.
  • substantially similar is well understood by the person skilled in the art.
  • a variant may be an isoform or allele which shows amino
  • KM/11.05.2007 acid exchanges compared to the amino acid sequence of the most prevalent peptide isoform in the human population.
  • a substantially similar peptide has a sequence similarity to the most prevalent isoform of the protein or peptide of at least 80%, preferably at least 85%, more preferably at least 90%, most preferably at least 95%.
  • Substantially similar are also degradation products, e.g. proteolytic degradation products, which are still recognized by the diagnostic means or by ligands directed against the respective full-length protein or peptide.
  • variants is also meant to relate to splice variants.
  • variant also relates to a post-translationally modified protein such as glycosylated protein.
  • a “variant” is also a peptide which has been modified after collection of the sample, for example by covalent or non-covalent attachment of a label, particularly a radioactive or fluorescent label, to the protein.
  • protein(s) of interest refers to Matrin-3 or a variant thereof.
  • Matrin-3 (SEQ. ID NO: 1) is not known.
  • the present invention further provides the use of Matrin-3 or a variant thereof as biomarker for Alzheimer's disease.
  • biomarker refers to molecules in an individual which are differentially present (i.e. present in increased or decreased levels) depending on presence or absence of a certain condition, disease, or complication.
  • biochemical markers are gene expression products which are differentially present (e.g. through increased or decreased level of expression or turnover) in presence or absence of a certain condition, disease, or complication.
  • a biomarker of the invention is biochemical marker.
  • a biochemical marker is a protein, polypeptide or peptide.
  • the level of a suitable biomarker can indicate the presence or absence of a particular condition, disease, or risk, and thus allow diagnosis or determination of the condition, disease or risk.
  • level relates to amount or concentration of a peptide or polypeptide in an individual or a sample taken from an individual.
  • amount also relates to concentration. It is evident, that from the total amount of a substance of interest in a sample of known size, the concentration of the substance can be calculated, and vice versa.
  • concentration preferably semi -quantitatively or quantitatively. Measuring can be done directly or indirectly. Indirect measuring includes measuring of cellular responses, bound ligands, labels, or enzymatic reaction products.
  • Measuring can be done according to any method known in the art. Preferred methods are described in the following
  • the method for measuring the level of a protein of interest comprises the steps of (a) contacting a cell capable of a cellular response to the protein with the protein for an adequate period of time, (b) measuring the cellular response.
  • the method for measuring the level of a protein of interest comprises the steps of (a) contacting a protein with a specifically binding ligand, (b) (optionally) removing non-bound ligand, (c) measuring the amount of bound ligand.
  • the protein is contained in a body fluid sample, and the amount of the protein in the sample is measured.
  • a body fluid may be blood, blood serum, blood plasma and cerebral liquor such as cerebrospinal fluid.
  • body fluids include cerebrospinal fluid.
  • One important example is the measurement in CSF. Samples of body fluids can be obtained by any method known in the art.
  • the samples may be further processed.
  • the protein maybe purified from the sample according to methods known in the art, including filtration, centrifugation, or extraction methods such as chloroform/phenol extraction.
  • the sample or processed sample is added to a cell culture and an internal or external cellular response is measured.
  • the cellular response may include the expression of a reporter gene or the secretion of a substance, e.g. a protein, peptide, polypeptide, or a small molecule.
  • Binding according to the present invention includes both covalent and non-covalent binding.
  • a ligand according to the present invention can be any protein, peptide, polypeptide, nucleic acid, or other substance binding to the protein of interest. It is well known that proteins, if obtained or purified from the human or animal body, can be modified, e.g. by glycosylation. A suitable ligand according to the present invention may bind the protein exclusively or additionally via such sites.
  • the ligand should bind specifically to the protein to be measured.
  • “Specific binding” means that the ligand should not bind substantially to ("cross-react” with) another protein or substance present in the sample investigated.
  • the specifically bound protein or isoform should be bound with at least 3 times higher, more preferably at least 10 times higher and even more preferably at least 50 times higher affinity than any other relevant protein, peptide or polypeptide.
  • Non-specific binding may be tolerable, particularly if the investigated protein can still be distinguished and measured unequivocally, e.g. according to its size (such as on a Western Blot), or by its relatively higher abundance in the sample.
  • Binding of the ligand can be measured by any method known in the art.
  • the method is semi-quantitative or quantitative. Suitable methods are described in the following.
  • binding of a ligand may be measured directly, e.g. by NMR or surface plasmon resonance.
  • the ligand may be coupled covalently or non-covalently to a label allowing detection and measurement of the ligand.
  • Labelling may be done by direct or indirect methods.
  • Direct labelling involves coupling of the label directly (covalently or non-covalently) to the ligand.
  • Indirect labelling involves binding (covalently or non-covalently) of a secondary ligand to the first ligand.
  • the secondary ligand should specifically bind to the first ligand.
  • Said secondary ligand may be coupled with a suitable label and/or be the target (receptor) of tertiary ligand binding to the secondary ligand.
  • the use of secondary, tertiary or even higher order ligands is often used to increase the signal.
  • Suitable secondary and higher order ligands may include antibodies, secondary antibodies, and the well-known streptavidin- biotin system (Vector Laboratories, Inc.)
  • the ligand may also be "tagged" with one or more tags as known in the art.
  • tags may then be targets for higher order ligands.
  • Suitable tags include biotin, digoxygenin, His-Tag, Glutathion-S-Transferase, FLAG, GFP, myc-tag, influenza A virus haemagglutinin (HA), maltose binding protein, and the like.
  • the tag is preferably at the N-terminus and/or C-terminus.
  • Suitable labels are any labels detectable by an appropriate detection method. Typical labels include gold particles, latex beads, acridan ester, luminol, ruthenium, enzymatically active labels, radioactive labels, magnetic labels ("e.g. magnetic beads", including paramagnetic and superparamagnetic labels), and fluorescent labels.
  • Enzymatically active labels include e.g. horseradish peroxidase, alkaline phosphatase, beta-Galactosidase, Luciferase, and derivatives thereof.
  • Suitable substrates for detection include di-amino-benzidine (DAB), 3,3'-5,5'-tetramethylbenzidine, NBT- BCIP (4-nitro blue tetrazolium chloride and 5-bromo-4-chloro-3-indolyl-phosphate, available as readymade stock solution from Roche Diagnostics), CDP-IO StarTM (Amersham Biosciences), ECFTM (Amersham Biosciences).
  • a suitable enzyme-substrate combination may result in a colored reaction product, fluorescence or chemoluminescence, which can be measured according to methods known in the art (e.g. using a light-sensitive film or a suitable camera system).
  • Typical fluorescent labels include fluorescent proteins (such as GFP and its derivatives), Cy3, Cy5, Texas Red, Fluorescein, and the Alexa dyes (e.g. Alexa 568).
  • fluorescent labels are available e.g. from Molcular Probes (Oregon). Also the use of quantum dots as fluorescent labels is contemplated.
  • Typical radioactive labels include 35S, 1251, 32P, 33P and the like.
  • a radioactive label can be detected by any method known and appropriate, e.g. a light-sensitive film or a phosphor imager.
  • Suitable measurement methods according the present invention also include precipitation (particularly immunoprecipitation), electrochemiluminescence (electro- generated chemiluminescence), RIA (radioimmunoassay), ELISA (enzyme-linked immunosorbent assay), sandwich enzyme immune tests, electrochemiluminescence sandwich immunoassays (ECLIA), dissociation-enhanced lanthanide fluoro immuno assay (DELFIA), scintillation proximity assay (SPA), turbidimetry, nephelometry, latex- enhanced turbidimetry or nephelometry, solid phase immune tests, and mass spectrometry such as SELDI-TOF, MALDI-TOF, or capillary electrophoresis-mass spectrometry (CEMS).
  • precipitation particularly immunoprecipitation
  • electrochemiluminescence electrochemiluminescence (electro- generated chemiluminescence)
  • RIA radioimmunoa
  • Preferred ligands include antibodies, nucleic acids, proteins, peptides or polypeptides, and aptamers, e.g. nucleic acid or peptide aptamers.
  • Methods to such ligands are well-known in the art. For example, identification and production of suitable antibodies or aptamers is also offered by commercial suppliers. The person skilled in the art is familiar with methods to develop derivatives of such ligands with higher affinity or specificity. For example, random mutations can be introduced into the nucleic acids, proteins, peptides or polypeptides. These derivatives can then be tested for binding according to screening procedures known in the art, e.g. phage display.
  • Aptamers are chemically synthesized (usually short) strands of oligonucleotides (DNA or RNA) that can adopt highly specific three-dimensional conformations. Aptamers are designed to have appropriate binding affinities and specificities towards certain target molecules.
  • antibody includes both polyclonal and monoclonal antibodies, as well as any modifications or fragments thereof, such as Fv, Fab and F(ab)2 fragments that are capable of binding antigen or hapten.
  • the present invention also relates to a kit comprising a means or an agent for measuring Matrin-3 or a variant thereof.
  • Such means or agent may be any suitable means or agent known to the person skilled in the art. Examples for such means or agents as well as methods for their use have been given in this specification.
  • a suitable agent maybe any kind of ligand or antibody specific for measuring said biomarkers.
  • the kit may also comprise any other components deemed appropriate in the context of measuring the level(s) of the respective biomarkers, such as suitable buffers, filters, etc.
  • the kit may additionally comprise a user's manual for interpreting the results of any measurement(s) with respect to determining whether an individual suffers from Alzheimer's Disease.
  • a user's manual for interpreting the results of any measurement(s) with respect to determining whether an individual suffers from Alzheimer's Disease.
  • such manual may include information about what measured level corresponds to a decreased level.
  • the present invention also relates to the use of said kit for assessing Alzheimer's Disease in an individual.
  • the present invention also relates to the use of said kit in any of the methods according to the present invention for assessing Alzheimer's Disease in an individual.
  • normal level refers to the normal range of the level of protein of interest in a body fluid sample of a control.
  • a control is one or more individuals not suffering from Alzheimer's Disease. Preferably, the number of individuals is higher than 100, more preferably more than 500, most preferably more than 1000.
  • the normal range is determined by methods well known to the skilled person in the art. A preferred method is for example to determine the range of the values between quantile 2.5 and quantile 97.5, which leaves 5% of "normal" values outside the normal range or in other words, it covers 95% of all values of the control.
  • the pathological status is defied as deviation from the normal status. According to the invention this pathological status is indicated by a decreased or increased level of a biomarker.
  • the term "decreased level” as used herein refers to the level of protein of interest in a body fluid sample which is significantly lower than the normal level. Significantly lower means that the level is lower and that the difference to the normal level is statistically relevant (p ⁇ 0.05, preferably, p ⁇ 0.01).
  • the term “increased level” or “elevated” as used herein refers to the level of protein of interest in a body fluid sample which is significantly higher than the normal level. Significantly higher means that the level is higher and that the difference to the normal level is statistically relevant (p ⁇ 0.05, preferably, p ⁇ 0.01).
  • the protein of interest may also be used as target. Therefore, the present invention provides a method of screening for a compound which interacts with Matrin-3 or a variant thereof. Such methods are well known in the art.
  • a suitable method is for example the method of screening for a compound which interacts with Matrin-3 or a variant thereof comprising a) contacting said protein with a compound or a plurality of compounds under conditions which allow interaction of said compound or a plurality of compounds with said protein; and b) detecting the interaction between said compound or plurality of compounds with said protein.
  • the protein of interest maybe immobilized prior step a) or between step a) and step b).
  • Figure 1 shows an overview of the sample preparation procedure for LC/MS/MS analysis of brain tissue.
  • Figure 2 shows a SDS gel of solubilized protein sample of an AD case stained with Coomassie Blue.
  • the sample lane was cut into 14 pieces that were subjected to trypsin digestion and LC/MS/MS analysis.
  • the left lane shows the molecular weight marker, the gel excision pattern is shown on the right.
  • Brains - Human brains from AD patients were obtained from the Sun Health Research Institute through Dr. Thomas G. Beach (Sun City, Arizona, USA). The brain tissue was collected approximately two hours after death and was immediately frozen at -80 0 C. Brains were neuropathologically staged according to Braak and Braak (Neuropathological stageing of Alzheimer-related changes. Acta Neuropathol (Berl), 1991. 82, 239-259). Brain tissue was isolated from the occipital association gyrus of 2 different AD patients (80 and 78 years old) with Braak stage VI and a control case (88 years old) with Braak stage II. The patient with Braak stage II showed no plaque load and was used as a control patient, the patients with Braak stage VI showed heavy plaque load.
  • Sample preparation for LC-MS/MS - The brain tissue was isolated using a sterile surgical blade and measured areas of 30 and 35 mm2 (AD) and 32 and 36 mm2 (control) were collected. Exclusively brain tissue from grey matter areas was gathered. Brain tissue was solubilized by addition of 2x 50 ⁇ l of 2% SDS, 66 mM Na2CO3, 2% ⁇ -mercapto- ethanol and 10% saccharose with intermediate heating for 10 min at 70 0 C. Supernatants were combined and protein was precipitated as previously described (16). Briefly, 400 ⁇ l methanol were added to the supernatant and mixed thoroughly. 200 ⁇ l of chloroform were added.
  • the sample was incubated on ice for 5 min and centrifuged at 10000 rpm for 2 minutes until a clear phase separation was achieved.
  • the denatured proteins resided in the interphase and were isolated by carefully removing the hydrophilic phase and the addition of 300 ⁇ l methanol.
  • the proteins were sedimented by centrifugation for 5 min at 10000 rpm. The supernatant was removed and the protein pellet was dried at room temperature. Proteins in each sample were separated on a 4-12% SDS gel (1 mm thick) using 3-(N-morpholino) propanesulphonic acid (MOPS) running buffer for Ih 30 min at 120 volts.
  • MOPS 3-(N-morpholino) propanesulphonic acid
  • the samples were alkylated with 0.1M NH4HCO3 / 55 mM iodoacetamide in the dark at room temperature for 30 min.
  • the gel pieces were digested after two additional washing steps with NH4HCO3 and ACN by incubation with 30 ⁇ l trypsin (0.02 ⁇ g / ml, Seq. Grade modified trypsin, Promega, Madison, WI, USA) overnight at room temperature.
  • the peptides were eluted by addition of 250 ⁇ l 25mM NH4HCO3 and 5% formic acid for 15 min at 37°C. The eluates were combined and evacuated to dryness.
  • Antibodies The correlation of selected candidates detected by LC-MS/MS with immunohistochemistry or Western blotting was largely dependent on the availability of antibodies. As primary antibodies, the following were tested: anti-gelsolin (1:1000, Sigma, clone GS-2C4), anti-synaptotagmin (Synaptic Systems GmbH, Goettingen, Germany, clone 41.1), anti-synaptogyrin (Synaptic Systems, clone 80.1), anti-reticulon 1 (1:500, Santa Cruz Biotechnologies, Santa Cruz, CA, USA, clone Mon 160), anti-syntaxin (1:2000, Chemicon International, Temecula, CA, USA, clone MAB336), anti-A ⁇ (1:2000, The Genetics Company, Zuerich, Switzerland, clone WO-2), anti-ubiquitin (1:100, Dako, Glostrup, Denmark), anti-phosphotau (1:1000, kindly provided by Dr.
  • Immunohistochemistry - Brain tissue slices were rehydrated for 5 min with phosphate buffered saline (PBS, Gibco, 14200-067) containing protease inhibitors (Roche Diagnostics GmbH, 1836145, 1 tablet per 50 ml) and fixed on the polyethylene foil covered glass slide by applying 70% acetone (0 0 C) for approximately 1 min. Sections were washed twice for 2 min with 1 ml PBS and for doublestaining, thioflavine S was applied as a 1% aqueous solution for 3-5 min followed by a differentiation step with 70% ethanol for 3-5 min.
  • PBS phosphate buffered saline
  • protease inhibitors Roche Diagnostics GmbH, 1836145, 1 tablet per 50 ml
  • the membranes were incubated with a horseradish peroxidase linked secondary antibody (Amersham) for Ih at room temperature. After 2 x 15 min washing in PBS-T (or TBS-T), the membranes were developed for 2 min at room temperature with ECLTM (Amersham) solution according to the protocol of the manufacturer. Visualization was done with conventional X-Ray films (Amersham) and band intensities were compared. For the image analysis, GeneTools software (Syngene, Cambridge, United Kingdom) was used.
  • High resolution confocal immunofluoroescence images were recorded using HCX PL APO 63x / 1.3 Glyc Corr objective. The three dimensional image were reconstructed and processed using Imaris Software.
  • Peptides were dissolved in 1% formic acid (buffer A: 0.5% Acetic acid, 0.012% Heptafluorobutyric acid) and concentrated and desalted online on a C18 PepMap 100 micro precolumn (5 ⁇ m particle size, 300 ⁇ m x 1 mm; Dionex Corporation, Sunnyvale, CA, USA) that was coupled to a self packed (7 cm, 3 ⁇ m; ProntoSil C18-ACE-EPS, Bischoff Chromatography, Atlanta, GA, USA) and pulled (P-2000 laser puller, Sutter Instrument, Novato, CA, USA) fused silica capillary (100 ⁇ m i.d. X 365 ⁇ m o.d.). The chromatographic separation was then performed by a 100 minute nonlinear gradient from 5 to 55% buffer B (80% ACN/0.5% Acetic Acid/0.012% HFBA) with a constant flow rate of0.20 ⁇ l/min.
  • buffer A 0.5% Acetic acid, 0.012% Heptafluoro
  • the mass spectrometric data acquisition was performed with a survey scan followed by 7 data dependent MS2 scans with a repeat count of 2.
  • the collision energy was set to 35%.
  • Database searching for protein identification - SEQUEST was used to for searching the HumanGP database for peptide sequence and protein identification.
  • HumanGP is a protein sequence database that is derived by assembling in sequences the results of Blast searches against the human chromosomes of a non-redundant protein set from Swissprot and Trembl (from Human, Mouse and other major organisms). Search parameters included differential mass modification to methionine due to possible oxidation and static mass modification to cysteine due to alkylation by iodoacetamide. Furthermore one missed cleavage of trypsin was accepted.
  • Protein quantification by mass spectrometry Protein (semi-)quantification was based on the comparison of peptide counts, which reflect the number of peptides identified for a given protein, in samples derived from AD patients with samples from age-matched controls. This number is indicative for the relative abundance of a protein in a sample. The comparison was performed with in-house developed software (MSPresso).
  • Criteria for peptide and protein identification For the comparisons of peptide counts, the single datasets from the analysis of the respective samples were combined to final sets.
  • Peptides identified by SEQUEST may have three different charge states (+1, +2, or
  • the peptide counts of the AD patients were normalized. For this reason, the ratios of several abundant proteins (tubulin, actin, actinin and GAPDH) in the samples were averaged and the peptide counts in the AD patients were standardized against this ratio. Therefore, the peptide counts of the first AD patient were normalized with 0.93, the second AD patient with 1.44. The comparison of protein levels was based on the normalized peptide counts.
  • proteins where the measured peptide ratio in the respective pair of samples was in a range of ⁇ 0.3, were accepted.
  • protein X was found in the first measurement with a ratio of 0.6 between AD patient and control and in the second series with a ratio of 0.8. In this case, the average was taken (0.7) and the protein was accepted for the analysis and evaluation. Some exceptions were made, but only when the ratios belonged into the same classification group (see below). Proteins consistently present in only the AD or the control sample, were classified as well.
  • the peptide count is indicative for protein levels actually present in the brain.
  • the peptide count reflects the number of peptides identified for a given protein after said protein was digested with an enzyme. As not all peptides produced by the digestion may be measured, the approach using peptide count is not quantitative, but semi quantitative, therefore it is only possible to describe a tendency and to assign the proteins into classification groups.
  • a protein with an averaged peptide count from the two series of 1.3 ⁇ x was considered to be increased.
  • a protein with an averaged ratio of 0.7 ⁇ x ⁇ 1.3 was regarded as comparable, whereas a ratio being x ⁇ 0.7 was accepted as decreased.
  • AD Alzheimer's disease .
  • proteins related to AD we found increased levels of A ⁇ , tau, apolipoprotein E, Ubiquitin, several heat shock proteins (e.g. Hsp90), members of the complement pathway (complement C4 and C3), as well as GFAP.
  • Other proteins known to be associated with AD e.g. antichymotrypsin, serum amyloid P component or C-reactive protein
  • Matrin-3 was not reported before in relation to Alzheimer's Disease and showed significantly decreased level of expression in the AD patients.
  • Table 1 Peptide counts of the protein of invention. The peptide counts were normalized: with 0.93 for the peptide counts of the first AD-patient (white) and with 1.44 for the peptide counts of the second AD-patient (grey). Average peptide counts of x ⁇ 0.7 means that the expression level in AD patient is decreased compared to a control.
  • AD Patient suffering from Alzheimer's Disease
  • r range: ⁇ 0.3
  • av. average of the two patients.

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Abstract

La présente invention concerne un procédé pour déterminer la maladie d'Alzheimer in vitro, le procédé consistant à mesurer dans un échantillon de fluide corporel le niveau de Matrine-3 ou d'un variant de cette substance. Un niveau réduit de Matrine-3 ou d'un variant de cette substance indique que ledit individu est atteint de la maladie d'Alzheimer.
PCT/EP2007/004947 2006-06-06 2007-06-04 Matrine-3 utilisée comme biomarqueur de la maladie d'alzheimer WO2007140974A1 (fr)

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EP06115034.8 2006-06-06

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Non-Patent Citations (4)

* Cited by examiner, † Cited by third party
Title
FOUNTOULAKIS MICHAEL: "Application of proteomics technologies in the investigation of the brain.", MASS SPECTROMETRY REVIEWS 2004 JUL-AUG, vol. 23, no. 4, July 2004 (2004-07-01), pages 231 - 258, XP002447635, ISSN: 0277-7037 *
GÜNTERT A: "Characterization of Amyloid-beta and Other Proteins Related to Alzheimer's Disease, their Role in Neurodegeneration and Biomarker Discovery", INAUGURALDISSERTATION ZUR ERLANGUNG DER WUERDE EINES DOKTORS DER PHILOSOPHIE VORGELEGT DER PHILOSOPHISCHEN-NATURWISSENSCHAFTLICHENFAKU LTAET DER UNIVERSITAET BASEL, XX, XX, 4 July 2006 (2006-07-04), XP002445051 *
WANG GUOQING ET AL: "Preliminary studies on Alzheimer's disease using cDNA microarrays.", MECHANISMS OF AGEING AND DEVELOPMENT JAN 2003, vol. 124, no. 1, January 2003 (2003-01-01), pages 115 - 124, XP002447627, ISSN: 0047-6374 *
XU ET AL: "Differences in apolipoprotein E3/3 and E4/4 allele-specific gene expression in hippocampus in Alzheimer disease", NEUROBIOLOGY OF DISEASE, BLACKWELL SCIENCE, OXFORD, GB, vol. 21, no. 2, February 2006 (2006-02-01), pages 256 - 275, XP005243139, ISSN: 0969-9961 *

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