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US 7,682,807 B2Granted
Alzheimer's disease diagnosis based on mitogen-activated protein kinase phosphorylation
AssigneeBlanchette Rockefeller Neurosciences Institute
InventorsWei-Qin Zhao, Daniel L. Alkon
Life of this patent
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Abstract
A method of diagnosing Alzheimer's disease in a patient comprises determining whether the phosphorylation level of an indicator protein in cells of the patient after stimulus with an activator compound is abnormally elevated as compared to a basal phosphorylation level, the indicator protein being e.g. Erk1/2 and the activator compound being e.g. bradykinin.
Claims62
A method comprising the steps of:
- (a) measuring a basal phosphorylation level of Erk1/2 protein in cells selected from the group consisting of fibroblast, buccal mucosal, neuron, and blood from a subject;
- (b) contacting said cells from the subject with a PKC activator;
- (c) measuring an activated phosphorylation level of Erk1/2 protein in said subject cells after step (b);
- and
- (d) calculating a ratio of the activated phosphorylation level determined in step (c) to the basal phosphorylation level of step (a);
- and
- (e) comparing the calculated ratio of step (d) to a calculated ratio of activated to basal phosphorylation levels of Erk1/2 measured from known Alzheimer's disease cells, or
- comparing the calculated ratio of step (d) to a calculated ratio of activated to basal phosphorylation levels of Erk1/2 measured from non-Alzheimer's disease cells;
- wherein said method is indicative of Alzheimer's disease
- if the calculated ratio of step (d) is not higher or lower than said calculated ratio of step (e) for known Alzheimer's disease cells or
- if the calculated ratio of step (d) is higher or lower than said calculated ratio of step (e) for non-Alzheimer's disease cells.
8 more elements
A method comprising the steps of:
- (a) measuring a background phosphorylation level of Erk1/2 protein in cells selected from the group consisting of fibroblast, buccal mucosal, neuron, and blood cells from a subject;
- (b) stimulating cells of the subject by contacting said cells with an IP3R agonist;
- (c) measuring an activated phosphorylation level of the Erk1/2 protein in the contacted cells, and
- (d) comparing the activated phosphorylation level of the Erk1/2 protein in the contacted cells to the background level and the activated phosphorylation level known for a cell from an Alzheimer's disease subject or a cell from a healthy subject;
- wherein said method is indicative of Alzheimer's disease
- if the activated phosphorylation level of said contacted cells is not higher or lower than the response level for a cell from an Alzheimer's disease subject or
- if the response phosphorylation level of said contacted cells is higher or lower than the response level known for a cell from a healthy subject.
4 more elements
A method comprising the steps of:
- (a) obtaining cells from a subject wherein said cells are selected from the group consisting of fibroblast, buccal mucosal, neuron, and blood;
- (b) measuring a basal phosphorylation level of Erk1/2 protein in said cells;
- (c) contacting said cells with a PKC activator;
- (d) measuring the phosphorylation level of Erk1/2 protein in said cells after said contacting step (c);
- and
- (e) calculating a first ratio of the phosphorylation level measured in step (d) to the phosphorylation level measured in step (b) and comparing said calculated first ratio to a calculated second ratio from known Alzheimer's disease cells and a calculated third ratio from known non-Alzheimer's disease cells;
- wherein said method is indicative of Alzheimer's disease
- (i) if the calculated first ratio of step (e) is not higher or lower than the second calculated ratio, or
- (ii) if the first ratio of step (e) is not higher or lower than the third calculated ratio.
6 more elements
A method comprising the steps of:
- contacting cells from a subject with a PKC activator wherein said cells are selected from the group consisting of fibroblast, buccal mucosal, neuron, and blood,
- measuring phosphorylation of Erk1/2 protein in said subject cells at one or more time points-after the contacting step, and
- comparing the measured phosphorylation of Erk1/2 protein in the subject cells to phosphorylation in non-Alzheimer's disease cells at the same time points after contacting the non-Alzheimer's disease cells with the PKC activator,
- wherein said method is indicative of a positive diagnosis of Alzheimer's disease
- if phosphorylation of Erk1/2 protein in the subject cells is increased compared to the non-Alzheimer's disease cells.
2 more elements
A method comprising the steps of:
- (a) incubating cells from a subject with a PKC activator in a diluent, wherein said cells are selected from the group consisting of fibroblast, buccal mucosal cell, neuron, and blood;
- (b) before, at the same time or after step (a) incubating cells of a same type from the subject with a control compound or with said diluent, thereby producing unstimulated control cells;
- (c) comparing a level of phosphorylated Erk1/2 protein in the cells of step (a) to a level of phosphorylated Erk1/2 protein in the unstimulated control cells;
- wherein said method is indicative of a positive diagnosis of Alzheimer's disease
- if the level of the phosphorylated Erk1/2 protein in the cells of step (a) is increased as compared to the phosphorylated Erk1/2 protein in the unstimulated control cells.
2 more elements
A method comprising:
- measuring phosphorylation of Erk1/2 protein in Alzheimer's disease cells and non-Alzheimer's disease control cells and selecting a compound that increases phosphorylation of Erk1/2 protein and/or increases duration of phosphorylation of Erk1/2 protein in Alzheimer's disease cells compared to non-Alzheimer's disease control cells after contacting said cells with said compound.
A method comprising the steps of:
- (a) contacting skin fibroblast cells from a subject and from a non-Alzheimer's disease control subject with an effective, phosphorylation-stimulating concentration of bradykinin,
- (b) measuring the amount of phosphorylated Erk1/2 in the subject cells at one or more time points selected from the group consisting of 2 minutes, 5 minutes, 10 minutes, 20 minutes, and 30 minutes comprising Western blotting using an antibody specific for phospho-Erk1/2;
- (c) measuring phosphorylated Erk1/2 in skin fibroblast cells from a non-Alzheimer's disease control subject at the same time point or points of step (b) comprising Western blotting using an antibody specific for phospho-Erk1/2;
- wherein the amount of phosphorylated Erk1/2 in steps (b) and (c) is normalized to protein in said cells;
- (d) comparing phosphorylated Erk1/2 in the subject cells with phosphorylated Erk1/2 in the non-Alzheimer's disease control cells at said time points,
- wherein said method is indicative of a positive diagnosis of Alzheimer's disease
- if phosphorylated Erk1/2 in the subject cells is increased compared to the non-Alzheimer's disease control cells at one or more of said time points.
4 more elements
A method comprising the steps of:
- a) stimulating cells from a subject with a PKC activator that increases phosphorylation of Erk1/2 protein wherein said cells are selected from the group consisting of fibroblast, buccal mucosal, neuron, and blood, and
- b) comparing a level of unphosphorylated Erk1/2 protein and phosphorylated Erk1/2 protein in the cells of step (a) to a level of unphosphorylated Erk1/2 protein and phosphorylated Erk1/2 protein in unstimulated control cells of the same type from said subject,
- wherein said method is indicative of a positive diagnosis of Alzheimer's disease
- if phosphorylated Erk1/2 protein in the cells of step (a) is increased-compared to unstimulated cells.
1 more element
Dependencies are inferred from the claim text.