02 October 2009 CECON II. Budapest 1 Application of proteomics methods Application of proteomics methods in the identification of in the identification of biomarkers, suitable for studying biomarkers, suitable for studying obesity and obesity related obesity and obesity related diseases diseases O B E K O N O B E K O N Consortium OBECON Consortium
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02 October 2009CECON II. Budapest1 Application of proteomics methods in the identification of biomarkers, suitable for studying obesity and obesity related.
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02 October 2009 CECON II. Budapest 1
Application of proteomics methods in the Application of proteomics methods in the identification of biomarkers, suitable for studying identification of biomarkers, suitable for studying
obesity and obesity related diseasesobesity and obesity related diseases
O B E K O NO B E K O NConsortium
OBECON Consortium
2
Identifying clinically relevant biomarkers
Select patients to increase likelihood of clinical trial success
Select the best treatment/drug for each patient
Biomarkers as “early killers” or as approved surrogate markers
Improve patient compliance in the absence of early clinical improvement
Differentiate efficacy or safety of a drug within the same class
Select the best treatment/drug for each patient
Biomarkers as “early killers” or used to exclude certain patient groups from clinical trials
Monitor and avoid potential toxic effects
Patient recruitment for clinical trials
Early disease detection, early treatment
Patient recruitment for clinical trials
Predict likely course of disease
Disease managementDisease management
Stratification markers
Stratification markers
Efficacy biomarkersEfficacy biomarkers
Differentiation markers
Differentiation markers
Toxicity biomarkersToxicity biomarkers
Screening markersScreening markers
Prognostic markersPrognostic markers
Drug DevelopmentDrug DevelopmentBiomarker applicationsBiomarker
applications
02 October 2009 CECON II. Budapest
Multivariate panels versus single biomarkers
New tests are likely to increasingly focus on multivariate panels
Multivariate patterns appear more robust than single analyte tests (less sensitive to individual difference and assay noise)
The rational behind is well known in the clinical field (physicians generally integrate multiple test results to arrive to the diagnosis)
Identification of the proteins/analytes in the panel is desirable but not critical
Regulations are currently being put in place (MammaPrint™, FDA regulations on In Vitro Diagnostic Multivariate Index Assays)
02 October 2009 CECON II. Budapest 3
Genome vs. proteomeHuman Genome = 20 - 30,000 genes
Human Proteome = 300,000 to 1,200,000 protein variants
Genome – static; proteome - dinamic; „… there is only a 0.4 correlation between global mRNA and protein
•apolipoprotein a-ii•chorein 1d•collagen alpha 1(xvii) chain•dorsal neural-tube nuclear protein•fam91a1 protein•filip1l protein•g protein-coupled receptor 45•homeobox protein cux-2•hypothetical protein fp972•inter-alpha-trypsin inhibitor heavy chain 1•kiaa0357•paternally expressed gene 3 isoform 2•scavenger receptor class a, member 3•transcription intermediary factor 1•uncharacterized protein dkfzp667f0711•unnamed protein product 3•vacuolar protein sorting-associated protein 8 homolog•zonadhesin variant 3
•abhydrolase domain-containing protein fam108b1•alpha-2-antiplasmin•anti-oxldl immunoglobulin•complement factor properdin•gelsolin•giant protein p619•heat repeat containing 2•kiaa1662•kininogen•lim and calponin homology domains-containing protein 1•nsp5•nucleoporin nup160•tchp protein•titin•transmembrane protein 108•tubulin•u3 small nucleolar rna-associated protein 15•unnamed protein product 1•unnamed protein product 4
02 October 2009 CECON II. Budapest
Proteomics strategy of OBECON
02 October 2009 CECON II. Budapest 17
Plasma collection; Represent. sampling
Depletion
Fraction generatio
n
Shotgun MS
OGE-IEF, PGC -LC
mAB-productio
n
MS/MS Prot.,N-glyc
Selection differentia
ls
Result verification
on individual samples
Epitope ID
Protein ID
Fractionation of plasma samples by OFFGEL isoelectrophoresis
02 October 2009 CECON II. Budapest 18
M ND D ND D ND D ND D OGE Fr# 5 5 6 6 7 7 8 8
Analysis of complex oligosaccharides using graphitized carbon LC/MS
Glycosilation – PTM;Changes properties of proteins;
Congenital disordersAffects cell-cell communication;Divers and its effect pattern dependent;
Comparison of Average VmaxN values (and corresponding SD) between six experiments done with a single tracer, either on the same day or on different days
Average VmaxN 6 Experiments Same tracer Pool Different days(SD of the VmaxN from the 6 experiments)
1BioSystems International (BSI) Ltd., Debrecen; 2BSI SAS, Evry, France; 3Gedeon Richter Ltd., Budapest; 4Hungarian Academy of Sciences, Chemical Research Center, Budapest; 5TargetEx Ltd., Dunakeszi;