EP2097755A1 - Procédé in vitro de diagnostic et de diagnostic précoce de maladies neurodégénératives - Google Patents

Procédé in vitro de diagnostic et de diagnostic précoce de maladies neurodégénératives

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Publication number
EP2097755A1
EP2097755A1 EP07856296A EP07856296A EP2097755A1 EP 2097755 A1 EP2097755 A1 EP 2097755A1 EP 07856296 A EP07856296 A EP 07856296A EP 07856296 A EP07856296 A EP 07856296A EP 2097755 A1 EP2097755 A1 EP 2097755A1
Authority
EP
European Patent Office
Prior art keywords
disease
determination
dementia
diagnosis
alzheimer
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Ceased
Application number
EP07856296A
Other languages
German (de)
English (en)
Inventor
Andreas Bergmann
Andrea Ernst
Harald Hampel
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
BRAHMS GmbH
Original Assignee
BRAHMS GmbH
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by BRAHMS GmbH filed Critical BRAHMS GmbH
Publication of EP2097755A1 publication Critical patent/EP2097755A1/fr
Ceased legal-status Critical Current

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Classifications

    • 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

  • the present invention relates to a novel in vitro method for the diagnosis and, in particular, early diagnosis of neurodegenerative diseases, in particular dementias such as Alzheimer's disease and its precursors.
  • diagnosis is used as a generic term for medical determinations which, depending on the clinical condition of the patient in which the determination is carried out, can be based on different questions and those of recognition and, in the present case, especially serve the early detection, the determination of severity and the course assessment, including the therapy-accompanying course assessment, and the prognosis of the future course of a disease.
  • a diagnosis can also be a negative diagnosis in which, for example, the presence of a particular disease is rendered unlikely on account of the fact that a certain disease-typical concentration of biomarker in a blood sample of a patient can not be detected.
  • the diseases to be diagnosed in the present invention are rather slow-developing, chronic neurodegenerative diseases of noninfectious etiology, especially dementia disorders.
  • Dementia diseases are generally referred to diseases for which a common feature is that acquired intellectual ability, v. A. the memory, and the normal personality level are impaired as a result of brain damage. Dementias are usually relatively slow-developing diseases of chronic nature. If dementia occurs before old age in middle age, it is referred to as pre-senile dementia. In the case of dementia patients, a distinction is made between the following diseases or groups of diseases on the basis of the symptoms typical for them and changes in the brain pathology:
  • Alzheimer's disease (Alzheimer's disease) is the most common neurodegenerative dementia disorder, accounts for 2/3 of all cases of dementia and is also the most important therapeutic area for the present invention
  • Post mortem pathological features that can be ascertained with certainty: the formation of amyloid plaques and neurofibrillary tangles and the loss of nerve cells (for review see (1)), references in the description in the form of numbers refer to the description following bibliography).
  • Amyloid plaques consist of extraneuronal aggregates of the amyloid ⁇ -protein, while the neurofibrillary tangles mainly contain tau protein and neurofilaments. It is believed that plaque and neurofibrillary formation is the cause of nerve cell death.
  • AD Alzheimer's disease
  • Dementia with Lewy bodies is the second leading cause of dementia after Alzheimer's disease.
  • DLB is characterized by the appearance of so-called Lewy bodies in the brainstem and in the cortex. These Lewy bodies consist predominantly of aggregates of the presynaptic protein ⁇ -synuclein and ubiquitin. Lewy body pathology may be associated to varying degrees with Alzheimer's and Parkinson's-type neuropathological changes.
  • DLB also causes the formation of beta-amyloid and senile plaques, but not neurofibrillary tangles (for review see (6)).
  • Lewy bodies are also present in the brain of patients with Parkinson's disease, albeit in a different distribution.
  • DLB Frontotemporal dementia
  • tauopathies which are characterized by an over- or underexpression of a subtype of subtypes or by the expression of a mutated tau protein.
  • vascular dementia refers to diseases in which dementia is triggered by circulatory disorders in the brain.
  • VAD multi-infarct dementia
  • subcortical VAD also referred to as Binswanger's disease
  • Binswanger's disease is a slowly progressive dementia that is pathologically characterized by cerebrovascular lesions in the cerebral white matter. Clinically, this results in behavioral problems such as agitation, irritability, depression and euphoria as well as a mild memory disorder.
  • the MuIti infarction dementia gradually develops as a result of several small strokes, also known as transient ischemic attacks (TIA), which led to the destruction of brain tissue in the cortex and / or subcortical areas.
  • TIA transient ischemic attacks
  • the strokes may have gone completely unnoticed, the dementia is the first noticeable in this case Episode.
  • MID transient ischemic attacks
  • cognitive abilities associated with severe depression, mood swings, and epilepsy.
  • a diagnosis of dementia diseases today is predominantly based on neuropsychological examinations and the observation of disease development and its course, using exclusion criteria for certain forms of dementia. These studies provide very ambiguous results in many cases, which the o.g. Explain numbers for the underdiagnosed dementia forms or incorrectly diagnosed cases.
  • the disease-typical brain changes naturally can not be detected directly in living patients, and apparatus-medical examinations of brain functions by means of e.g. Computer or magnetic resonance tomography are complex and expensive.
  • the diagnostic sensitivity and the specificity of at least 80% should be given.
  • the disease-specific change of the biomarker should be manifested at the earliest possible stage of the disease in order to begin appropriate therapies (8).
  • biomarker in the circulation (i.e., in the blood or in blood preparations such as serum or plasma) of a patient that could be used in clinical practice to improve the early and differential diagnosis of AD and all the above. Criteria met.
  • inflammatory markers such as IL-6 and TNF ⁇
  • markers of oxidative stress such as 3-nitrotyrosine
  • markers associated with characteristic pathological changes in AD such as amyloid ⁇ , which is a major component of amyloid plaques
  • tau protein which is an essential component of the neurofibrillary tangles (see the overview in (7); (9)).
  • AD Alzheimer's disease
  • suitable substances in blood or plasma samples and in patients who are suspected of Existence of dementia, in particular of AD is supportive of an early positive diagnosis and / or suitable for a negative exclusion diagnosis.
  • the present invention provides such an assay method in the form of an in vitro method for detection and early detection, for determining the severity and for evaluating the course of disease and prognosis of neurodegenerative diseases, which comprises administering to a patient in a serum or plasma sample suffers from objectively detectable cognitive disorders, determines the Apo C I immunoreactivity using an immunodiagnostic assay, and based on the concentration for the occurrence of, or a typical early form of, neurodegenerative disease or the course of the disease and / or the success of efforts to alleviate or prevent it.
  • the present invention is based on the experimental finding that in the blood (serum, plasma) of patients with cognitive disorders up to the diagnosis "possibly Alzheimer's” or "probably Alzheimer's” in a characteristic manner reduced concentrations of the immunoreactive Apo CI are measured in comparison with control persons ,
  • analyte is defined as it using an immunoassay, in particular an immunoassay, as described in the experimental part of the present application or in German patent application DE 103 43 815 Al the applicant is described in more detail, can be determined.
  • hydrophobic molecular structures eg the octyl residues of a hydrophobic octyl sepharose chromatography material, from which it is under typical conditions for the elution of proteins , in particular with dilute acetic acid, can be eluted.
  • free apolipoprotein CI does not necessarily mean that the material which can be separated by hydrophobic interaction chromatography must be completely free in the original samples.
  • said immunoassay in the blood of cancer patients also recognizes an apo C-I fraction which does not bind to octylsepharose and whose occurrence appears to be characteristic of tumor diseases. Possibly. Therefore, in the context of the overall diagnosis for dementia disease or its precursors using the measured values obtained in the determination of the immunoreactive Apo C-I, it would be ruled out that the measured values in the respective patient would be falsified by a cancer. This can be done, for example, in the manner described in DE 103 43 815 A1 by checking whether the measured concentration values are obtained by contact of the sample with e.g. Octyl Sepharose change.
  • the immunoreactive Apo CI to be determined according to the present invention comprises, in addition to analytes which represent or contain the complete peptide Apo CI, also "Apo CI derivatives". These include in particular immunoreactive apo CI fragments and aggregates, in particular those which behave in the said sandwich immunoassay as the free protein apolipoprotein CI.
  • the "derivatives" may be, for example, apolipoprotein C-1 molecules shortened by individual amino acids or amino acid sequences, or also, for example, by aggregation, sterically or conformally altered complete apolipoprotein C-I molecules.
  • Apo C-1 derivatives should also be encompassed by the invention takes into account the known facts that immunoassays normally can not differentiate between different analytes if the differences are outside the molecular segments used for antibody binding, i. in the case of a sandwich immunoassay in end regions of the detected molecule outside the amino acid sequence delimited by the two antibody binding sites. Furthermore, the further known fact is to be taken into account that when peptidic analytes occur in the circulation, the appearance of their degradation products in the form of peptide fragments must always be expected.
  • concentrations of such fragments are usually approximately proportional to the concentration of the starting peptide, the differences in concentration differences in stability or different rates of "clearing" of the peptide or fragment by further proteolytic degradation or by discharge from the circulation, e.g. reflect on the kidneys.
  • the determination according to the invention of immunoreactive Apo C-I in the context of the diagnosis of neurodegenerative diseases is proposed in particular as a measurement which supplements the parallel determination of physiological or clinical and neuropsychological parameters and other biomarkers and refines them for differential diagnostic purposes.
  • the method is therefore preferably carried out within the scope of a multiparameter determination, in which at least one additional one for the respective clinical picture is simultaneously detected.
  • meaningful biochemical or physiological parameter is determined and in which a measurement result is obtained in the form of a set of at least two measured variables, which is evaluated for the fine diagnosis of the neurodegenerative disease.
  • the immunoreactive Apo C-I e.g. at least one other biochemical parameter selected from the groups of natriuretic peptides, inflammatory mediators, complement components, cytokines, chemokines, blood coagulants and fibrinolytic factors, acute phase proteins and free radical compounds.
  • the co-parameters to be mentioned in particular include those parameters which are described in the earlier German patent applications of the applicant with the file references 10 2005 034 174.8 (midregionales Proadrenomedullin), 10 2005 036 094.7 (Liquorodiagnostic determination of procalcitoline), 10 2006 027 818.6 and 10 2006 023 175.9 and the parameter CPS-I (carbamoyl phosphate synthetase 1), to which relevant discussions with regard to neurodegenerative diseases can be found in the applications EP 05023420.2 and DE 10 2006 021 406.4.
  • the method according to the invention can also be carried out as a simultaneous determination by means of a chip technology measuring device or an immunochromatographic measuring device and the evaluation of the complex measurement results possibly obtained can take place with the aid of a suitable computer program.
  • FIG. 1 shows the results of the measurement of the concentrations of the immunoreactive Apo CI determinable with the sandwich immunoassay described in the experimental section, namely in EDTA plasmas of 60 healthy control persons, and of 196 patients with cognitive disorders of various severity corresponding to the above-mentioned groups (b), (c) and (d), ie the groups 11 SKS “(50 patients),” LKS mgl AD “(46 patients) and” ws AD "(100 patients).
  • a sandwich-type immunoassay was constructed from the components described below:
  • a) Coated Tubes Polystyrene tubes (Greiner) were coated with a commercially available polyclonal affinity-purified antibody against apolipoprotein C-I (source: Acris Antibody, Bad Neuheim, Germany). The antibody was obtained according to the manufacturer by immunization of rabbits with human Apo C-I and purified on a Sepharose affinity column with human apolipoprotein C-I. For coating, 0.2 ⁇ g of antibody in 300 ⁇ l of PBS was bound to polystyrene tubes (Greiner, Germany) coated with sheep anti-rabbit IgG antibody (Sigma). The binding was stopped after 18 hours at room temperature.
  • the tubes were washed twice with 3 ml each of 0.5% bovine serum albumin (BSA) in PBS. After drying in vacuo, the tubes were used as the solid phase for the apolipoprotein C-I immunoassay.
  • BSA bovine serum albumin
  • Acridinium ester-labeled antibody 100 ⁇ g of another antibody to human apolipoprotein CI (from Rabbit, reference: Academy Bio-Medical Company, Texas, USA) in 100 ⁇ l of PBS was incubated with 10 ⁇ g of acridinium-NHS ester (in 10 ⁇ l of acetonitrile ) implemented. After a 10 minute incubation at room temperature, the purification was carried out of the labeled antibody with removal of unreacted constituents of the reaction mixture by HPLC on SW 300 (Waters).
  • Serum or plasma samples were diluted 1:10 000 with PBS, 0.5% BSA. Of these, 300 ⁇ l each were added to the o.g. coated with the immobilized antibody. Pipette and then incubated for 4 hours at room temperature with shaking (300 U / min on a Heidolph orbital shaker). The tube contents were then washed out with PBS (4 times filling and decanting with 1 ml PBS each), and apolipoprotein CI bound to the tube wall was incubated within 20 hours at room temperature and 300 rpm with 300 ⁇ l per tube of acridinium ester labeled anti -Apolipoprotein C-I antibody reacted. Thereafter, unbound labeled antibody was removed by washing 5 times with 1 ml PBS each time, and the remaining chemiluminescence was measured in a known manner in a Berthold Luminometer 952T.
  • concentrations of apo C-I which can be determined immunologically in plasmas, which can be identified by the values for the medians of the various patient groups, decrease with the severity of the symptoms in the direction:
  • apo CI is not a brain-specific parameter
  • the measurement of apo C-I immunoreactivity in the circulation is very well suited for the purpose of assisting AD early diagnosis, given the above-mentioned correlations.

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  • Life Sciences & Earth Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Biomedical Technology (AREA)
  • Hematology (AREA)
  • Chemical & Material Sciences (AREA)
  • Urology & Nephrology (AREA)
  • Molecular Biology (AREA)
  • Immunology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Medicinal Chemistry (AREA)
  • Microbiology (AREA)
  • Biotechnology (AREA)
  • Neurosurgery (AREA)
  • Neurology (AREA)
  • Food Science & Technology (AREA)
  • Cell Biology (AREA)
  • Physics & Mathematics (AREA)
  • Analytical Chemistry (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • General Physics & Mathematics (AREA)
  • Pathology (AREA)
  • Investigating Or Analysing Biological Materials (AREA)

Abstract

L'invention concerne un procédé in vitro de détection et de détection précoce, de détermination du degré de gravité et d'évaluation de l'évolution ainsi que de pronostic de maladies neurodégénératives, dans lequel on détermine à l'aide d'un procédé de détermination d'immunodiagnostic l'immunoréactivité de l'apolipoprotéine C-I (Apo C-I) dans un échantillon de sérum ou de plasma d'un patient qui souffre de troubles cognitifs ressentis objectivement ou subjectifs.
EP07856296A 2006-11-29 2007-11-28 Procédé in vitro de diagnostic et de diagnostic précoce de maladies neurodégénératives Ceased EP2097755A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
DE102006056337A DE102006056337A1 (de) 2006-11-29 2006-11-29 In vitro Verfahren zur Diagnose und Frühdiagnose von neurodegenerativen Erkrankungen
PCT/EP2007/010331 WO2008064883A1 (fr) 2006-11-29 2007-11-28 Procédé in vitro de diagnostic et de diagnostic précoce de maladies neurodégénératives

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EP2097755A1 true EP2097755A1 (fr) 2009-09-09

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EP07856296A Ceased EP2097755A1 (fr) 2006-11-29 2007-11-28 Procédé in vitro de diagnostic et de diagnostic précoce de maladies neurodégénératives

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US (1) US20100035286A1 (fr)
EP (1) EP2097755A1 (fr)
JP (1) JP2010511159A (fr)
DE (1) DE102006056337A1 (fr)
WO (1) WO2008064883A1 (fr)

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Publication number Priority date Publication date Assignee Title
GB201616691D0 (en) * 2016-09-30 2016-11-16 King S College London Assay
CN111650372A (zh) * 2019-03-04 2020-09-11 中国医学科学院药物研究所 载脂蛋白c1在作为胃癌诊断及预后评价生物标记物中的应用
WO2024004944A1 (fr) * 2022-06-28 2024-01-04 株式会社Mcbi Procédé de génération d'informations de prise en charge d'évaluation, système de génération d'informations de prise en charge d'évaluation et dispositif de traitement d'informations

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Publication number Priority date Publication date Assignee Title
DE69333461T2 (de) * 1992-10-13 2005-03-24 Duke University VERFAHREN ZUM nachweis der Alzheimer Krankheit
DE60225821T2 (de) * 2001-10-04 2009-04-30 Immuno-Biological Laboratories Co., Ltd., Takasaki Reagens zum Nachweis eines Risikofaktors der Alzheimer-Krankheit, Nachweiskit dafür und Verfahren zum Nachweis eines Risikofaktors der Alzheimer-Krankheit unter ihrer Verwendung
DE10343815A1 (de) 2003-09-22 2005-04-14 B.R.A.H.M.S Ag Verfahren zur Diagnose von Erkrankungen unter Bestimmung von Apolipoprotein C-I, und dessen Verwendung in der Therapie von Erkrankungen
EP1694816B1 (fr) * 2003-11-07 2013-08-28 Ciphergen Biosystems, Inc. Biomarqueurs pour la maladie d'alzheimer
EP1780287A1 (fr) * 2005-10-26 2007-05-02 B.R.A.H.M.S. Aktiengesellschaft Procédé in vitro destiné au diagnostic de maladies neurodégénératives

Non-Patent Citations (1)

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Title
See references of WO2008064883A1 *

Also Published As

Publication number Publication date
JP2010511159A (ja) 2010-04-08
DE102006056337A1 (de) 2008-06-05
WO2008064883A1 (fr) 2008-06-05
US20100035286A1 (en) 2010-02-11

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