Blood-based Biomarkers for Detection
of early stage Alzheimer’s Disease:
A Successful Multi-Analyte
Profiling Approach
Ralph L. McDade, Ph.D.
Strategic Development Officer
The Myriad RBM Approach
The Platform and Validation
Success Stories
Alzheimer’s Disease story
Multi-Analyte Profile (MAP)
The Myriad RBM
Approach
The Approach
Pre-Clinical & Exploratory
Phase II & Beyond
Cast a wide net
Target key markers
1
Start with a large
Multi-Analyte
Profile (MAP)
biomarker panel
2
3
Identify key
biomarker patterns
Develop a focused,
custom panel
One, validated, highly automated platform throughout
drug development
The Platform
Industrialized form of
Luminex xMAP
All liquid handling steps
automated using the Tecan
Evo platform
Proprietary blockers to
handle most matrix effects
Validated to clinical lab
standards
GLP and CLIA certified
56 successful regulatory
compliance audits
The object is to find a robust biomarker pattern
These 13 analytes
were found to
discriminate
responders from
non-responders.
Custom MAP
Custom MAP 13-plex
These 13 analytes
were used to help the
clinicians stratify
clinical trial
participants
Simponi (golimumab )
1. Adiponectin
2. EGF
3. Eotaxin
4. ICAM-1
5. IL-6
6. IL-10
7. IL-15
8. MCP-1
9. MMP-9
10. PAP
11. TNF-a
12. VEGF
13. von Willebrand Factor
Core Competency – Immunoassay
development in a multiplexed environment
Cytokines
AcutePhase
Reactants
Metabolic
Markers
Hormones Inflammatory
markers
CardioVascular
Cancer Autoimmunity
Markers
Assay Development Contracts
Merck Germany
Merck U.S.
Celgene
AstraZeneca
Novartis
Eli Lilly
Pfizer
Amgen
Centocor
NIH
Satoris
BMS
EU/IMI
NCI
Psynova
Genentech
Validation Parameters
Lowest Detectable Dose / LLOQ
Normal Range
Dynamic Range
Imprecision
Spiked Recovery
Linearity
Correlation
Cross-reactivity
Matrix Interferences
Stability – Short term storage / Freeze-thaw
Just some of the over 400 users of this
biomarker approach that have publicly
acknowledged RBM success
MRBM Bibliography: Publications Citing MAP
Services
Publications By Therapeutic Indication
Bone
Autoimmune Disease
Disease/Metabolism,
and Arthritis, 6%
1%
Kidney/Tox,
8%
Cancer, 14%
Neurological Disease,
18%
Cardiovascular, 11%
Miscellaneous, 9%
Diabetes and
Metabolic Markers,
7%
Endocrine, 1%
Inflammation and
Immune Response,
37%
Gastrointestinal, 1%
A Few Success Stories
Schizophrenia
Pulmonary Fibrosis
Myelofibrosis
Bone Metastasis
Kidney Disease
COPD
Ocular Inflammation
Alzheimer’s Disease
Alcohol Abuse
Expertise in Biomarker Research for Neuroscience
Psychiatric vs Neurodegenerative
Schizophrenia, BD,
and MDD
SZP: 18 years
BD: 21 years
AD, PD, and OD
AD: >65 years
BD: 60 years
OD: >65 years
MDD: 25 years
“The blood based biomarker patterns of disease in younger
people are easier to see as there are fewer confounders
such as underlying diseases like CVD and diabetes. In
addition, an average 65 year old in the US is on a regimen of
at least five different drugs.”
We understand much
about the terminal
pathology
We understand very
little about the etiology
of AD
What
are
the
project’s
goals?
Early Dx for MCI/AD (blood test for >50 years)
Identify rapid MCI to AD converters (20%)
Differentiate AD from other forms of dementia
Identify responders in drug trials
Tony Wyss-Coray’s group at Stanford Med
Ray Biotech 2-D slide-based array 100+ analytes
First suggestion in literature that a signal for MCI/AD was
present in the plasma proteome
“We found 18 signaling proteins in blood plasma that can
be used to classify blinded samples from Alzheimer's and
control subjects with close to 90% accuracy and to
identify patients who had mild cognitive impairment that
progressed to Alzheimer's disease 2–6 years”
Ray et al. Heat Map of the 18 markers
EDTA Plasma from 19 AD/22 Controls
HumanMAP v 1.6 (90 analytes)
Attempt to reproduce the Ray, et al findings
Mentions early Rotterdam data with their 1,200 member
cohort and our later 152 analyte MAP.
“Furthermore, utilization of other analytes from the 90analyte panel did show a diagnostic accuracy of approximately
70%”
“
“
CSF from 62 AD; 33 Controls and 25 OD
Pre-DiscoveryMAP (152 analytes)
MAP data added with tau, P-tau181 & Aβ42 >90% accuracy in
AD/OD diagnosis
17 MAP analytes by Random Forest; 32 by PAM
“Two categories of biomarkers were identified: (1) analytes
that specifically distinguished AD (especially CSF Aβ42 levels)
from cognitively normal subjects and other disorders; and (2)
analytes altered in multiple diseases, but not in cognitively
normal subjects ”
Serum from 197 AD; 203 Controls (TARC Cohort)
Pre-DiscoveryMAP (152 analytes)
MAP data + clinical data+ ApoE4 genotype = >95% AUC AD vs.
normals
30 MAP analytes by Random Forest; 25 by SAM (minimal
overlap with Ray et al: Ang 2 and TNFα)
“The identification of blood-based biomarker profiles with
good diagnostic accuracy would have a profound impact
worldwide and requires further validation.”
O’Bryant et al. 2010 – Arch Neurol. 67(9): 1077-1081
TARCC Analyses
• Restricted to only top 30 markers
• Added clinical lab values
• Total cholesterol, triglycerides, high density lipoproteins, low
density lipoproteins, lipoprotein-associated phospholipase [LpPLA2], homocysteine, C-peptide)
• Retained demographic factors
O'Bryant et al 2011a
TARCC Analyses
AUC (95% CI)
Sensitivity (95% CI)
Specificity (95% CI)
Demographic data
0.80(0.74-0.86)
0.71(0.62-0.79)
0.78(0.69-0.85)
Clinical variables
0.81(0.75-0.87)
0.74(0.65-0.82)
0.76(0.66-0.84)
Biomarker alone
0.91(0.87-0.95)
0.88(0.80-0.93)
0.82(0.73-0.88)
Combined
0.94(0.91-0.97)
0.89(0.81-0.94)
0.85(0.76-0.91)
O'Bryant et al 2011a
TARCC Analyses
O'Bryant et al 2011a
TARCC Analyses
• Need to cross-validate screener in an independent cohort
• Alzheimer’s Disease Neuroimaging Initiative (ADNI)
• Large-scale study of AD and Mild Cognitive Impairment (MCI)
• Has same biomarker panel on subset of AD cases and controls
O'Bryant et al 2011b
TARCC Analyses
• Problem – ADNI has plasma based proteins while TARCC
has serum
• There is no consensus as to what blood fraction to look at for AD
biomarkers
• Many groups look at both serum and plasma, even using same
markers
• Markers may or may not behave consistently across media
O'Bryant et al 2011b
TARCC Analyses
• TARCC has plasma-based proteins on 40 AD cases
• Looked at serum and plasma results to identify
• Proteins that behave consistently across serum and plasma R2>0.75
• Significant (p<0.05) relation to AD status
• Identified 11 proteins that met criteria
• CRP, adiponectin, pancreatic polypeptide, fatty acid binding protein, IL18,
beta 2 microglobulin, tenascin C, I.309, factor VIII, VCAM1, MCP1
O'Bryant et al 2011b
TARCC Analyses
• Created RF biomarker risk score based on the 11 proteins
using the TARCC serum data
• Applied the algorithm (protein risk score, demographics,
clinical labs) to the ADNI plasma data
O'Bryant et al 2011b
TARCC Analyses
AUC (95% CI)
Sensitivity (95% CI)
Specificity (95% CI)
Biomarker alone
0.70(0.62-0.78)
0.54(0.45-0.63)
0.78(0.65-0.87)
Biomarker +
clinical +
demographics
CSF tau/Aβ ratio
0.88(0.83-0.93)
0.79(0.71-0.86)
0.87(0.75-0.93)
0.92(0.87-0.96)
0.84(0.76-0.90)
1.0(0.93-1.0)
O'Bryant et al 2011b
Serum vs. Plasma?
“
Performance evaluation of a multiplex assay for future use in biomarker
discovery efforts to predict body composition
Clin Chem Lab Med 2011
”
Beam J., Wright, N., Thompson, P., Hu, C., Guerra, S., and Chen, Z.
Thoroughly compared serum and plasma from the same
donor and bleed with HumanMAP v. 1.6 (90 analytes)
70 “useful” analytes in “healthy, normal” samples
29 analytes had a concordance >0.8 and 41 had a
concordance of <0.8 between serum and plasma with
24<0.5
“Serum showed a slight advantage…..”
MRBM recommends serum for any MCI/AD diagnostic
ADNI Cohort of 566 individuals tested for 190 analytes
Focused on 54 controls and 163 MCI to AD converters
11 analyte signature with APOE
Meta-analysis produced an 8 feature signature with 86%
SN and 87% SP
By adding longitudinal data this was improved to over
90% for both SN and SP
Products Currently in
Development:
NeurodegenerativeMAP™
CSF MAP
MCI/AD Dx and Prognostic (identify
rapid converters)
AD vs OD Differential
NeurodegenerativeMAP™
Goal : Develop and validate blood-based
biomarkers for Alzheimer's Disease and
other neurodegenerative disorders
Processed thousands of samples on our
DiscoveryMAP panel from groups including:
Meta-analysis of datasets and publications
Condensed to the most robust assays
NeurodegenerativeMAP
1.
Adiponectin
16. BDNF
2.
ACT
17. CD40
31. Interleukin-1 Receptor
Antagonist
44. Super Oxide Dismutase
45. Stem Cell Factor
32. Interleukin-8
3.
Alpha 1 Antitrypsin
18. CEA
4.
Alpha 1 Microglobulin
19. Clusterin
5.
Angiopoietin 2
20. Complement C3
46. Tenascin C
33. Interleukin-10
34. Lipoprotein (a)
47. Thyroxine Binding
Globulin
35. Macrophage Migration
Inhibitory Factor
48. Tissue Inhibitor of
Metalloproteinases 1
49. TRAIL-R3
6.
Angiotensinogen
21. Complement Factor H
7.
Apolipoprotein A1
22. Cortisol
8.
Apolipoprotein A2
23. EGFR
36. Macrophage
Inflammatory Protein-1
alpha
9.
Apolipoprotein B
24. Factor VII
37. MMP-2
10. Apolipoprotein C-III
25. FAS Ligand
38. MMP-9
11. Apolipoprotein E
26. Ferritin
39. Myeloperoxidase
12. Apolipoprotein H
27. Haptoglobin
40. Pancreatic Polypeptide
13. Anti-thrombin III
28. HB-EGF
41. RANTES
14. BLC
29. IgM
42. Resistin
15. Beta 2 microglobulin
30. IGFBP 2
43. Sortilin
50. Tumor Necrosis Factor
Receptor 2
51. Vascular Cell Adhesion
Molecule 1
52. Vascular Endothelial
Growth Factor
53. Vitamin D Binding Protein
54. Von Willebrand Factor
CSF MAP
Goal : Develop and validate CSF-based
biomarkers for Alzheimer's Disease and
other neurodegenerative disorders
Processed hundreds of CSF samples on
DiscoveryMAP® and other panels
Meta-analysis of datasets and publications
Condensed to the most robust assays
CSF MAP
1. Amyloid Beta 40
15. MMP-10
2. Amyloid Beta 42
16. NCAM
3. ACT
17. NT-proBNP
4. Alpha 1 Antitrypsin
18. P-Tau 181
5. Alpha 2 Macroglobulin
19. Placental Growth Factor
6. Apolipoprotein H
20. Stem Cell Factor
7. AXL
21. Super Oxide Dismutase 1
8. EGFR
22. Stem Cell Factor
9. FAS Ligand
23. Tau
10. Ferritin
24. TRAIL-R3
11. Fetuin A
25. Transforming Growth Factor Alpha
12. HB-EGF
26. Vascular Cell Adhesion Molecule 1
13. Interleukin-1 Receptor Antagonist
27. Vascular Endothelial Growth Factor
14. Interleukin-8
Myriad RBM’s Collaboration with the Spinal
Muscular Atrophy Foundation
The Spinal Muscular Atrophy Foundation Announces a Biomarker Panel to Guide SMA
Therapeutic Development
NEW YORK, NY – April 3, 2012 – The Spinal Muscular Atrophy (SMA) Foundation
announced today the launch of a biomarker assay panel for SMA using Myriad RBM’s MultiAnalyte Profiling (MAP) technology platform. The SMA-MAP panel is designed to evaluate
the severity of SMA and disease progression and can be used to assess drug efficacy and
shorten the duration of clinical trials for SMA therapeutics.
Study Design and Results
129 plasma samples from 18 clinical sites
LC/MS + 267 biomarkers from OncologyMAP® and
DiscoveryMAP ®
27 biomarker panel developed, including 7 new immunoassays
The Myriad RBM
Advantages
Myriad RBM’s Consultative Services
MRBM has built an extensive database through years of
careful sample procurement and collaboration with
leading institutions
Successful partnerships with major biopharma
companies to develop improved drugs and diagnostics
Data generated using MRBM’s services have been
featured in over 330 peer-reviewed journal articles
Myriad RBM’s Companion Diagnostic
Programs
Neurodegenerative disease
Psychiatric disorders
Infectious disease
Oncology
Inflammatory disease
Impact of Subpopulation Response
Myriad RBM’s Companion Diagnostic
Program with Roche Pharma
Roche Poster, SIRS 2012
Schizophrenia International Research Society, April 14-18, 2012
Myriad RBM Advantages
Cost Effective
Low Sample Volume
Requirements
Extensive Biomarker
Menu
Assay Precision &
Reproducibility
Biomarker
Expertise/Database
Myriad RBM’s Product and Services Portfolio
DiscoveryMAP™ 250+ (264 analytes)
RodentMAP® v2.0 (58 analytes)
DiscoveryMAP™ (189 analytes)
Rat MetabolicMAP™ (21 analytes)
OncologyMAP™ (101 analytes)
Rat KidneyMAP™ (12 analytes)
HumanMAP® v1.6 (88 analytes)
Mouse CytokineMAP A, B & C
PsyMAP™ v 1.0 (51 analytes)
CustomMAP
CardiovascularMAP™ (50 analytes)
InflammationMAP™ (46 analytes)
MetabolicMAP™ (21 analytes)
TruCulture™ Tubes
KidneyMAP™ (16 analytes)
TruCulture™ MAP (46 analytes)
CustomMAP (any combination of
assays from our menu)
For more information on Myriad RBM’s Companion
Diagnostic services, please contact us at
sbs@rulesbasedmedicine.com