Blood-based assay for detecting tauopathy or amyloid-forming disorders

Abstract

To provide a method for detecting tauopathy and / or amyloid-forming disorders. [Solution] The method comprises contacting a sample with a capture antibody directed against a p217+ tau epitope to form an antibody-peptide complex, and separately contacting the antibody-peptide complex with a detection antibody to bind the detection antibody to the antibody-peptide complex. The amount of p217+ tau is determined by detecting the detection antibody. The detected amount of p217+ tau is used to determine whether a subject has tauopathy or is at risk of developing tauopathy, or whether a subject has an amyloid-forming disease or is at risk of developing an amyloid-forming disease, if the amount of p217+ tau peptide exceeds a predetermined threshold. The method has improved sensitivity such that the predetermined threshold exceeds the lower limit of quantification and / or the lower limit of detection of the assay.

Claims

1. An assay method for detecting p217+ tau peptide in a target, Obtaining a plasma sample from the aforementioned subject, The plasma sample is then contacted with a capture antibody targeted against the p217+ tau epitope. Then, the capture antibody is bound to the p217+ tau peptide in the plasma sample, and the antibody - Forming peptide complexes, Washing the antibody-peptide complex, The antibody-peptide complex is brought into contact with the detection antibody, and the detection antibody is then converted to the antibody-peptide complex. To bind to the complex, The aforementioned detection antibody is detected to determine the amount of the p217+ tau peptide in the plasma sample. To determine The assay method, including the above.

2. The method according to claim 1, wherein the capture antibody is immobilized on a solid phase.

3. The method according to claim 2, wherein the solid phase is magnetic beads.

4. The aforementioned capture antibody contains an epitope containing amino acids 210-220 of human tau protein. The method according to claim 1, wherein the members are coupled together.

5. The detection antibody contains amino acids 7-20 or 116-127 of human tau protein. The method according to claim 1, wherein the pitope is attached.

6. The method according to claim 4, wherein the capture antibody is pT3.

7. The method according to claim 5, wherein the detected antibody is pT82.

8. The plasma sample is diluted with a sample diluent before contact with the capture antibody, The sample diluent contains a nonionic surfactant and tris(hydroxymethyl)aminomethane. The method according to claim 3, comprising at least one of the following.

9. A method for detecting tauopathy in a subject, Obtaining a plasma sample from the aforementioned subject, The amount of p217+ tau peptide in the plasma sample is detected using an assay. (Here, the assay uses a capture antibody targeted against the p217+ tau epitope.) By using this method, the capture antibody is bound to the p217+ tau peptide in the plasma sample, and the antibody A body-peptide complex is formed, and the detection antibody is used to convert the detection antibody into the antibody-peptide. (It is something that binds to the complex.) If the amount of the p217+ tau peptide exceeds a predetermined threshold, the target is To determine that you have tauopathy or are at risk of developing tauopathy (here And the predetermined threshold exceeds the lower limit of quantification (LLOQ) of the assay. The method, including the method described above.

10. Before the assay measures the amount of the p217+ tau peptide present, The claim is that the p217+ tau peptide derived from the plasma sample is not concentrated by sedimentation. The method described in 9.

11. The method according to claim 9, wherein the plasma sample is crude plasma.

12. The claim states that the LLOQ corresponds to the coefficient of variation (CV) of 15-25% of the assay. The method described in 9.

13. The method according to claim 12, wherein the LLOQ corresponds to 20% of the CV of the assay.

14. The method according to claim 9, wherein the predetermined threshold is at least three times the LLOQ. 。

15. Claim 9, wherein the predetermined threshold is at least 10 times the lower detection limit of the assay. Methods used.

16. The aforementioned tauopathies include familial Alzheimer's disease, sporadic Alzheimer's disease, and chromosome 17 Frontotemporal dementia with Parkinsonian syndrome (FTDP-17), progressive supranuclear Paralysis, corticobasal degeneration, Pick's disease, progressive subcortical gliosis, predominance of neurofibrillary tangles Type 1 dementia, diffuse neurofibrillary tangle disease with calcification, argyrophilic grain dementia, amyotrophic lateral sclerosis Parkinson's syndrome, dementia complex, Down syndrome, Gerstmann-Sträussler syndrome Jaicker's disease, Hallerforden-Spatz disease, inclusion body myositis, Creutzfeldt-Jacob's disease B's disease, multiple system atrophy, Niemann-Pick disease type C, prion protein, cerebral amyloid, blood vessels Disorders, subacute sclerosing panencephalitis, myotonic dystrophy, and non-guanine malformations due to neurofibrillary tangles. An motor neuron disease, post-encephalitis parkinsonian syndrome, chronic traumatic encephalopathy, and boxer cognition. The method according to claim 9, selected from the group consisting of diseases (boxer's disease).

17. The method according to claim 16, wherein the tauopathy is Alzheimer's disease.

18. The method according to claim 16, wherein the tauopathy is progressive supranuclear palsy.

19. A method for detecting amyloid formation disease in a subject, Obtaining a plasma sample from the aforementioned subject, The amount of p217+ tau peptide in the plasma sample is detected using an assay. (Here, the assay uses a capture antibody targeted against the p217+ tau epitope.) The capture antibody is then bound to the p217+ tau peptide in the plasma sample. , an antibody-peptide complex is formed, and the detection antibody is used to convert the detection antibody to the antibody-peptide complex. (It is something that binds to the cytoplasmic complex.) If the amount of the p217+ tau peptide exceeds a predetermined threshold, the target is To determine that you have an amyloid-forming disorder or are at risk of developing one. (Here, the predetermined threshold exceeds the lower limit of quantification (LLOQ) of the assay.) )and The method, including the method described above.

20. Before the assay measures the amount of the p217+ tau peptide present, The claim is that the p217+ tau peptide derived from the plasma sample is not concentrated by sedimentation. The method described in 19.

21. The method according to claim 19, wherein the plasma sample is crude plasma.

22. The claim states that the LLOQ corresponds to the coefficient of variation (CV) of 15-25% of the assay. The method described in 19.

23. The method according to claim 22, wherein the LLOQ corresponds to 20% of the CV of the assay.

24. The method according to claim 19, wherein the predetermined threshold is at least three times the LLOQ. Law.

25. Claim 1, wherein the predetermined threshold is at least 10 times the lower limit of quantification of the assay. The method described in 9.

26. The method according to claim 19, wherein the amyloid-forming disease is Alzheimer's disease.

27. A method for detecting or predicting tauopathy in a subject, The amount of p217+ tau peptide in the plasma sample is determined by the amount of p217+ tau peptide in the plasma sample. The capture antibody is brought into contact with the epitope, and the capture antibody is brought into contact with the plasma sample. By binding to the aforementioned p217+ tau peptide, an antibody-peptide complex is formed, and separately The antibody-peptide complex is brought into contact with the detection antibody, and the detection antibody is brought into contact with the antibody-peptide complex. By binding to the complex, detection is possible, To generate tau data corresponding to the amount of the detected p217+ tau peptide. and, Biomarker data corresponding to at least one biomarker detected from the subject To obtain data (where the biomarkers are NFL, adiponectin, and (Selected from the group containing leptin) Using a machine learning module, the tau data and further biomarker data are referenced. Compared with the set of illumination data, the subject has tauopathy or develops tauopathy. To determine or predict whether there is a risk of developing the disease. The method, including the method described above.

28. The aforementioned machine learning module includes a support vector machine module and a random forest module. Among the modules, logistic regression module, and gradient boosting module The method according to claim 27, comprising at least one of the following.

29. The aforementioned tauopathies include familial Alzheimer's disease, sporadic Alzheimer's disease, and chromosome 17 Frontotemporal dementia with Parkinsonian syndrome (FTDP-17), progressive supranuclear Paralysis, corticobasal degeneration, Pick's disease, progressive subcortical gliosis, predominance of neurofibrillary tangles Type 1 dementia, diffuse neurofibrillary tangle disease with calcification, argyrophilic granule dementia, amyotrophic lateral sclerosis Dementia, Parkinson's syndrome, dementia complex, Down syndrome, Gerstmann-Sträussler Scheinker's disease, Hallerforden-Spatz disease, inclusion body myositis, Creutzfeldt-Jackson's disease Cobb disease, multiple system atrophy, Niemann-Pick disease type C, prion protein cerebral amyloid blood Tube disorders, subacute sclerosing panencephalitis, myotonic dystrophy, non-guanine tangles due to neurofibrillary tangles Nian motor neuron disease, post-encephalitis-related Parkinsonian syndrome, chronic traumatic encephalopathy, and boxer's disease. The method according to claim 27, selected from the group consisting of cognitive impairment (boxer's disease).

30. The method according to claim 27, wherein the tauopathy is Alzheimer's disease.

31. The method according to claim 27, wherein the tauopathy is progressive supranuclear palsy.

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