Methods for diagnosis of Alzheimer's Disease in blood samples

• US 7,598,049 B2
• Filed: 11/19/2004
• Issued: 10/06/2009
• Est. Priority Date: 11/19/2003
• Status: Expired due to Fees

• Current Assignee: Rules-Based Medicine, Inc., Board of Trustees of the Leland Stanford Junior University (Stanford Management Co.), U.S. Government Represented By The Department Of Veterans Affairs
• Original Assignee: US Department of Veterans Affairs (Government of the United States of America), Satoris, Inc., Board of Trustees of the Leland Stanford Junior University (Stanford Management Co.)
• Inventors: Wyss-Coray, Anton, Ray, Sandip
• Primary Examiner(s): Chernyshev; Olga N
• Assistant Examiner(s): MacFarlane; Stacey
• Application Number:
  US10/993,813
• Publication Number:
  US 20050221348A1
• Time in Patent Office: 1,782 Days
• Field of Search: None
• US Class Current: 435/7.21
• CPC Class Codes: G01N 2333/4745 Insulin-like growth factor binding protein
• CPC Class Codes: G01N 2800/2821 Alzheimer
• CPC Class Codes: G01N 33/6896 Neurological disorders, e.g. Alzheimer's disease

Abstract

The inventors have discovered a collection of proteinaceous biomarkers (“AD biomarkers) which can be measured in peripheral biological fluid samples to aid in the diagnosis of neurodegenerative disorders, particularly Alzheimer'"'"'s disease and mild cognitive impairment (MCI). The invention further provides methods of identifying candidate agents for the treatment of Alzheimer'"'"'s disease by testing prospective agents for activity in modulating AD biomarker levels.

Claims

1. A method of aiding diagnosis of Alzheimer'"'"'s disease (“

AD”

), comprising comparing normalized measured levels of at least forty-six AD diagnosis biomarkers in a blood sample from a human individual seeking a diagnosis for AD to reference levels for the at least forty-six biomarkers in the blood sample,

wherein the human individual has a Mini Mental State Exam (MMSE) score of 14-26,

wherein the reference levels are obtained from normalized measured values of the at least forty-six biomarkers from samples in the blood of human individuals without AD,

wherein the at least forty-six AD diagnosis biomarkers comprise;

GCSF (granulocyte-colony stimulating factor);

IFN-g (interferon-gamma);

IGFBP-1 (insulin-like growth factor binding protein

     1);

BMP-6 (bone morphogenetic protein

     6);

BMP-4 (bone morphogenetic protein

     4);

Eotaxin-2;

IGFBP-2 (insulin-like growth factor binding protein

     2);

TARC (thymus and activation-regulated chemokine);

RANTES;

ANG (angiogenin);

PARC (pulmonary and activation-regulated chemokine);

Acrp30 (adipocyte complement-related protein of 30 kDa);

AgRP(ART) (agouti-related protein (agouti-related transcript));

TIMP-1 (tissue inhibitor of metalloproteinase

     1);

TIMP-2 (tissue inhibitor of metalloproteinase

     2);

ICAM-1 (intercellular adhesion molecule

     1);

TRAIL R3 (tumor necrosis factor-related apoptosis-inducing ligand receptor

     3);

uPAR (urokinase-type plasminogen activator receptor);

IGFBP-4 (insulin-like growth factor binding protein

     4);

LEPTIN(OB);

PDGF-BB (platelet-derived growth factor BB);

EGF (epidermal growth factor);

BDNF (brain-derived neurotrophic factor);

NT-3 (neurotrophin

     3);

NAP-2(neutrophil-activating peptide

     2);

IL-1ra (interleukin 1 receptor antagonist);

MSP-a (macrophage stimulating protein alpha);

SCF (stem cell factor);

TGF-b3 (transforming growth factor, beta

     3);

TNF-b (tumor necrosis factor beta);

MIP-1d;

IL-3 (interleukin

     3);

FGF-6 (fibroblast growth factor

     6);

IL-6 R (interleukin-6 receptor);

sTNF RII (soluble tumor necrosis factor receptor II);

AXL;

bFGF (basic fibroblast growth factor);

FGF-4 (fibroblast growth factor

     4);

CNTF (ciliary neurotrophic factor);

MCP-1(monocyte chemoattractant protein

     1);

MIP-1b (macrophage inflammatory protein-1beta);

TPO (thrombopoietin);

VEGF-B (vascular endothelial growth factor B);

IL-8 (interleukin

     8);

FAS; and

EGF-R (epidermal growth factor receptor),

whereby the diagnosis of AD is aided by determining a difference between the normalized measured levels of the at least forty-six AD diagnosis biomarkers to the reference levels of the at least forty-six biomarkers from non-AD samples wherein the difference meets or exceeds a statistically significant difference between normalized measured values of the at least forty-six AD diagnosis biomarkers in the blood samples from individuals without AD and individuals with AD, wherein the statistically significant difference indicates a diagnosis of AD.

2. The method of claim 1, wherein said blood sample is serum or plasma.

3. The method of claim 1, wherein the blood sample is obtained from the human individual immediately prior to measuring the levels of said biomarkers.

4. The method of claim 1, wherein measured values are measured from the blood samples from individuals without AD and individuals with AD.

5. The method of claim 1, wherein the reference levels for the at least forty-six biomarkers are obtained by a method comprising:

determining normalized measured levels of the at least forty-six biomarkers in normal individuals with a Mini Mental State Examination (MMSE) score greater than 25, having a statistically significant difference from normalized measured levels of the at least forty-six biomarkers in AD subjects with MMSE score of 25 and below.

6. The method of claim 1, wherein the statistically significant difference in normalized measured values of the at least forty-six AD diagnosis biomarkers in blood samples from individuals with AD relative to samples from individuals without AD is determined by a method comprising Significance Analysis of Microarrays (SAM).

7. The method of claim 6, wherein the statistically significant difference in normalized measured values of the at least forty-six AD diagnosis biomarkers determined by SAM has a q-value range from about 0.0001 to about 0.05.

8. The method of claim 1, wherein the statistically significant difference in normalized measured values of the at least forty-six AD diagnosis biomarkers in blood samples from individuals with AD relative to samples from individuals without AD is determined by a method comprising a t test.

9. The method of claim 8, wherein the statistically significant difference is measured in terms of a p-value or a q-value.

10. The method of claim 9, wherein the statistically significant difference is measured in terms of a p-value, and wherein the p-value is less than about 0.0403.

11. The method of claim 1, wherein the normalized measured values are normalized relative to median values determined contemporaneously using a pool of samples from individuals with AD and individuals without AD which includes the sample from the individual.

12. The method of claim 1, wherein comparing the measured levels comprises a method selected from the group consisting of Significance Analysis of Microarrays, Tree Harvesting, CART, MARS, Self Organizing Maps, Frequent Item Set, and Bayesian networks.

13. The method of claim 1, wherein the aiding the diagnosis of AD further comprises clinical diagnostic methods comprising taking patient histories, administering memory tests, attributing a MMSE score, administering psychological tests, or ruling out temporary or permanent conditions that may explain memory loss.

14. The method of claim 1, wherein determining the statistically significant difference associated with a diagnosis of AD comprises:

determining a mean value of normalized measured values of each of the at least forty-six AD diagnosis biomarkers in the blood samples from a group of individuals with AD;

determining a mean value of normalized measured values of each of the at least forty-six AD diagnosis biomarkers in the blood samples from a group of individuals without AD; and

finding a statistically significant difference between the mean values of the normalized measured values of the at least forty-six AD diagnosis biomarkers in the blood samples between the two groups.

15. The method of claim 14, wherein the group of individuals with AD and group of individuals without AD are age-matched populations.

16. The method of claim 1, wherein determining the statistically significant difference associated with progression of AD comprises:

determining a mean value of normalized measured values of each of the at least forty-six AD diagnosis biomarkers in the blood samples from a group of individuals with AD;

determining a mean value of normalized measured values of each of the at least forty-six AD diagnosis biomarkers in the blood samples from a group of individuals without AD; and

finding a statistically significant difference between the mean values of the normalized measured values of the at least forty-six AD diagnosis biomarkers in the blood samples between the two groups.

17. The method of claim 16, wherein the group of individuals with AD and group of individuals without AD are age-matched populations.