Meeting Report

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Meeting Report

• 2nd Annual Conference of US HUPO (Human
  Proteomics Organization)
• March 11-15, 2006 Boston

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Topics

• Serum Proteomics (HUPO plasma proteomics project)
• Disease Biomarker Discovery
• Protein Chips

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HUPO Plasma Proteome Project (PPP)

• Reference samples prepared by BD Diagnostics
• 18 Labs with diff. separation and MS platforms
• Evaluated technologies on depletion,
  fractionation, and MS technology platforms
• compared specimens of human serum and EDTA,
  heparin, and citrate-anticoagulated plasma
• www.bioinformatics.med.umich.edu/hupo/ppp
• www.ebi.ac.uk/pride

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PPP Findings

• 3020 proteins with gt 2 peptide ID
• IPI 2.2.1 and Sequest cut off score
• 49 proteins with quantitation by immunoassay (12
  in ng/ml range)

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Alternative search algorithms of ID

• PPP Jamboree Workshop, June 2004
• One raw file from PNNL LCQ-MS/MS data on serum
  with IgG depletion
• MSACOT, SEQUEST, Sonar, Spectrum Mill, X!Tandem
• Experts with the software plus manual inspection

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Kapp et al. An evaluation, comparison, and
accurate benchmarking of several publicly
available MS/MS search algorithms Sensitivity
and specificity analysis. PROTEOMICS Volume 5,
Issue 13 2005
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Independent analyses of raw spectra or peak list

• PepMiner software with raw spectra from 8 labs
• 2895 based on gt2 pep shared 865 with 2868 PPP
  3020
• ISB Peptide Prophet/ProteinProphet
• 960 proteins with pgt0.9 shared 479 with PPP 3020

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PPP suggestions

• Mass-spec screening is not efficient in biomarker
  discovery.
• Depletion of abundant proteins followed by
  fractionation are required to identify proteins
  with moderate abundance
• Sample preparation
• Plasma with EDTA or citrate for coagulation

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Lee Hartwell on Biomarker Discovery

• FDA approval of protein based-diagnostics low
• Issues with Plasma proteomics
• Sample collection
• Dynamic Range and complexity of proteins
• Under-sampling by ms/ms
• Bioinformatics algorithms

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Improve Biomarker Discovery

• Work at large scale
• Establish data standard
• Open informatics platform
• Share standard agents and samples

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SessionHuman Disease Proteomics

• Steven A. Carr, Broad Institute
• Not start with blood, start with cancer tissues
• Fluid close to the disease loci has high
  concentration of biomarkers
• Use patient as control
• Tissue-gtminced in PBS-gtms-gtsecretome

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Session post-tran modification

• Donald F. Hunt, University of Virginia
  comparative analysis of post-trans modified
  proteins
• MHC class I present peptide from self-antigens
  and tumor antigens
• Isolation of MHC I bound peptides
• IMAC enrichment for phospho-peptides
• MS characterization

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Session New proteomics technologies

• Richard M. Caprioli, Vanderbelt Univ.
• Tissue imaging with MALDI-MS
• FNA-gtslides-gtfix-gtstain-gtadd matrix-gtMALDI

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Session Cancer Proteomics

• Paul Tempst, MSKCC Differential protease
  activities confer tumor specific serum peptidome
  patterns. J Clin Invest. 2006 Jan116
• MALDI-MS to study peptides in serum from bladder,
  breast, or prostate Cancer patients

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Serum Peptide Signatures
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Session Functional Genomics

• Niro Ramachandran, Harvard Institute of
  Proteomics
• Self-assembling protein microarrays.Science.
  2004 Jul 2305(5680)86-90.

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Session Functional Proteomics

• Geeta Devgan, Yale Univ. Global Analysis of
  Protein Phosphorylation in Yeast. Nature. 2005
  Dec 1438679-84

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Nature. 2005 Dec 1438(7068)679-84.
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Mathias Uhlen on Antibody proteomics (
http//www.proteinatlas.org/ )

• 718 Antibodies again 718 Genes through QA
• Tissue microarray for 48 human tissues and 20
  different type of cancer
• 413,568 images
• Antibodies against gt 10,000 human proteins by
  2009

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Study lymphoid malignancies with antibody arrays

• Mol Cell Proteomics. 2006, From gene expression
  analysis to tissue microarrays - a rational
  approach to identify therapeutic and diagnostic
  targets in lymphoid malignancies
• 19 patients diagnosed with MCL
• Affymetrix U95v2 for gene expression profiling
• 83 antibodies against 57 gene products associated
  with MCL

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Thank you
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MS machines

• High resolution machines
• FTMS
• Orbitrap
• Electron transfer dissociation (ETD) vs CID
• "softer" mode of fragmentation
• Preserve labile modifications eg phoshporylation
• c- and z-series ions

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