Monday, August 25

12:00 Conference Registration

The Future of Clinical Proteomics: Focusing on Cancer

1:15-1:20 Chairperson's Opening Remarks
Dr. Timothy Veenstra, Director, Analytical Chemistry Laboratory

1:20-1:50 Challenges and Opportunities for Clinical Proteomics
Dr. Emmanuel Petricoin
Clinical proteomics focuses on the translational application of new protein-based technologies for bedside applications. Serum proteomic pattern diagnostics, a new concept in multiplexed biomarker analysis, may offer a new and exciting way to predict disease earlier, more accurately and specifically than ever before. However, large technological challenges remain. Applying this technology in a clinical setting is very different from applying it at the bench. Protein microarrays may also have a dramatic impact on drug target discovery, patient-tailored therapy and disease monitoring. Use of protein arrays is limited, however, by sensitivity and specificity issues, since there is no PCR-like tool for proteins. Translational bedside applications of these new tools hold great promise and are beginning to be successfully employed in clinical trials, but significant challenges remain for routine use.

1:50-2:20 Promises and Challenges of Proteomics in Cancer Diagnostics
Dr. Sudhir Srivastava, Chief, Cancer Biomarkers Research Group, Division of Cancer Prevention, National Cancer Institute
Protein-based approaches to early detection and diagnosis of cancer have a clear advantage. For screening purposes, biomarkers detectable in sera or other body fluids are most useful. Proteomic technologies can be used to identify markers for cancer diagnosis, for monitoring of disease progression, and to identify therapeutic targets. Proteomics is also valuable in the discovery of biomarkers, as the proteome reflects both the intrinsic genetic program of the cell and the impact of its immediate environment. By integrating analysis methods like protein profiling, the ability to detect and measure levels of promising protein-based markers in body fluids is of significant importance.

2:20-2:50 The Role of Proteomics in Breast Cancer Research
Dr. William S. Hancock, The Barnett Institute and Department of Chemistry, Northeastern University
With a diversity of potential molecular markers it is of clear interest to develop a proteomic approach to the analysis of cancer tissue samples, with the ultimate goal of analysis of protein expression patterns in individual patients. One approach to proteomic studies is termed "shotgun sequencing," in which the components of a proteomic sample are digested with a suitable protease, and the resulting peptides are then resolved by chromatography and detected by mass spectrometry (LC/MS/MS). This presentation will identify a substantial number of potential tumor markers in a small number of cells from the breast cancer cell line, SKBR-3, isolated by laser capture microdissection. The total number of cells used in this study, (10,000), is compatible with relatively non-invasive tumor biopsy performed in individual patients, and thus opens the possibility of profiling cancer markers in patients during disease progression or treatment.

2:50-3:30 Refreshment Break with Poster and Exhibit Viewing

Toxicoproteomics

3:30-3:35 Chairperson's Opening Remarks
Dr. Sven Beushausen, Associate Director, Proteomics, Investigative Toxicology, Pfizer Inc.

3:35-4:05 Application of Toxicoproteomics to Safety Assessment: Opportunities and Challenges
Dr. James MacGregor, Deputy Director, Washington Office, National Center for Toxicological Research, U.S. Food and Drug Administration
Proteomic technologies promise to provide new biomarkers of cellular injury that can link laboratory findings directly to human outcomes, ("bridging biomarkers"), to facilitate identification of perturbations in functional molecular pathways, and to allow "global" monitoring of gene products and key signaling molecules. The opportunities are exciting, but introduction into routine regulatory practice will require establishing the relationship of the endpoints measured to health outcomes and validation of the performance of the analytical platforms employed-a significant challenge with multiplexed assays that may involve hundreds or thousands of endpoints. Thus, different technologies may be employed in screening and "discovery" of key biomarkers, with simpler formats employed for regulatory testing.

4:05-4:35 Toxicoproteomics: Approaches to Toxicity Biomarker Identification and Validation in Pre-Clinical Safety Studies
Dr. Sven Beushausen
This presentation has been designed to provide an overview of how proteomics tools are currently being used to approach safety biomarker identification and development. The discussion will focus on platform technologies, data acquisition, output analysis, biomarker and biomarker profile identification, assay development and validation. Specific examples will be provided, where appropriate, and promising new technologies will be addressed.

4:35-5:05 Identification of Protein Biomarkers of Idiosyncratic Liver Toxicity
Dr. Gregory Opiteck, Senior Research Investigator, Clinical Discovery, Bristol-Myers Squibb
Since drug-induced liver injury is a great concern to the pharmaceutical industry, we are seeking better biomarkers to aid in its diagnosis and prevention. We are using proteomics to identify extra cellular protein biomarkers in cell culture systems, following treatment with various compounds known to be toxic or non-toxic. In this way, we can correlate the appearance of certain proteins in the cellular supernatants with a drug's clinical safety profile, and then later use assays to these proteins as diagnostics for drug-induced liver injury.

5:05-5:35 An Overview of Protein-Based Biomarkers of Safety with Emphasis on Nephrotoxicity
Dr. Rakesh Dixit, Senior Research Scientist, Safety Assessment, Merck & Co., Inc.
Protein-based safety biomarkers in accessible body fluids and/or certain accessible tissues can significantly aid in the monitoring of drug safety in preclinical and clinical studies. The presentation will provide an overview of currently used or potential safety biomarkers of liver, muscle, cardiovascular injury and nephrotoxicity. Special focus will be on the biomarkers of site-specific nephrotoxicity, including glomerulus injury, proximal and distal tube injury, and renal papillary necrosis. The application of newer technologies in discovery and development of protein-based safety biomarkers will be discussed.

Tuesday, August 26th

7:30-8:15 Technology Workshop (sponsorship available)

8:30-8:55 Recap of Roundtable Discussions

Technologies for Biomarker Discovery, Validation, and Profiling

8:55-9:00 Chairperson's Opening Remarks
Dr. Timothy D. Veenstra, Director, Analytical Chemistry Laboratory, National Cancer Institute, SAIC-Frederick

9:00-9:30 Multiple High-Resolution Serum Proteomic Features for Ovarian Cancer Detection
Dr. Timothy D. Veenstra
Serum proteomic pattern diagnostics is an emerging clinical paradigm that typically utilizes low-resolution mass spectrometry and generates a single set of biomarker classifiers. In the present study, we utilized a well-controlled serum study set from women being followed and evaluated for the presence of ovarian cancer to extend serum proteomic pattern analysis to a higher resolution instrument platform, in order to explore the existence of multiple distinct and highly accurate diagnostic sets of features present in the same mass spectrum. Using high-resolution mass spectral data, at least 56 different patterns were discovered that achieve greater than 85% sensitivity and specificity in testing and validation. Four of those feature sets exhibited 100% sensitivity and specificity in blinded validation tests. No models generated from data acquired from the same samples analyzed with a more common low-resolution mass spectrometer were validated as 100% sensitive and specific. The sensitivity and specificity of diagnostic models generated from high-resolution mass spectral data were dramatically superior than those generated from low-resolution mass spectral data.

9:30-10:00 Technology Integration for Proteomics Delivery
Dr. John Rogers, Senior Group Leader, Proteomics, Abbott Laboratories
Proteomics technologies are being applied in Abbott's pharmaceutical discovery efforts for the discovery and identification of protein markers of mechanism, efficacy, and toxicity. These technologies consist of a combination of robust sample preparation and fractionation methods, analysis by multiple gel and MS technologies, and informatics and automation improvements. Several case studies will be used to summarize the challenges and utility of integrating these components.

10:00-10:45 Coffee Break with Poster and Exhibit Viewing

10:45-11:15 Discovery and Quantification of Protein Biomarkers in Rheumatoid Arthritis Using Mass Spectrometry
Dr. Brad Guild, Director of Biomarker Discovery Proteomics, Millennium Pharmaceuticals Inc.
Working with our partner, Roche Diagnostics, we are using synovial fluid and serum samples as discovery media for the identification of new biomarkers in rheumatoid arthritis. Mass spectrometry has proven to be a powerful tool for differential protein analysis in these clinical samples. These protein-rich media are subjected to fractionation and enrichment strategies using multidimensional chromatography. Biochemically processed samples are trypsinized and further fractionated using online 2D/LC methods prior to analysis by ion trap mass spectrometry. The combination of methods yields high-resolution, high-throughput analysis of peptides that are used to catalog the protein composition of each sample. Prospective candidate markers are further assessed in serum using multiple reaction monitoring to provide a semi-quantitative measure. Thus, mass spectrometry is being applied in both the discovery and quantification of protein biomarkers of disease.

11:15-11:45 Discovery and Validation of Efficacy Biomarkers
Dr. Kevin Duffin, Director, Analytical Sciences, Pfizer Inc.
Efficacy biomarkers, including target-modulation biomarkers and disease biomarkers, are used at Pfizer to assess drug candidates in in vivo animal models and human patients. Discovery, assay development, and validation of novel biomarkers is coupled with use of commercially available biomarker assays. The strategies and technologies used for biomarker discovery/validation and their use in assessment of drug efficacy will be described in this presentation.

11:45-12:15 The Role of Systems Biology in Biomarker Discovery
Dr. Stephen Naylor, Chief Technology Officer, Beyond Genomics, Inc.
Understanding disease onset through progression, mechanisms of drug action, and monitoring such complex processes utilizing biomarkers, requires a perspective on how such systems behave and are modulated. In this regard, systems biology aims to simultaneously analyze and integrate data from multiple biological and clinical sources-including genes, proteins, and metabolites-to elucidate networks. The challenges of integrating and exploiting data from multiple sources has necessitated the development of new experimental design, bioanalytical methods, data integration, bioinformatics and knowledge-assembly protocols, all under the umbrella of Systems Biology. In the present work we describe how the integrated data from our systems biology correlation network affords new insight into biomarker discovery, and how such data can minimize the number of biomarkers needed to ensure maximum predictive effect in a complex disease process.

12:15-1:45 Lunch (on your own)

Plasma Biomarkers

1:55-2:00 Chairperson's Opening Remarks

2:00-2:30 The Diagnostic Potential of the Human Plasma Proteome
Dr. Leigh Anderson, CEO, Plasma Proteome Institute
The human plasma proteome holds the promise of a revolution in disease diagnosis and therapeutic monitoring, provided that major challenges in proteomics can be addressed. Plasma represents the largest and deepest version of the human proteome present in one sample: it contains all tissue proteins, (as leakage markers), plus very numerous immunoglobulin sequences, and it has an extraordinary dynamic range (more than 10 orders of magnitude in concentration, separating albumin from the rarest proteins now measured clinically). While almost 150 proteins are measured clinically in plasma, the rate of introduction of new protein tests approved by the FDA has paradoxically declined; only two new analytes have been approved in the last four years. The Plasma Proteome Institute has been formed to define and address the factors that currently impede translation of proteomics' promise into clinically useful measurements.

2:30-3:00 The HUPO Human Plasma Proteome Project
Prof. Gilbert Omenn, Professor, Medicine/Genetics, University of Michigan
The HUPO Plasma Proteome Project aims to comprehensively and quantitatively analyze serum and plasma proteins in large population cohorts, elucidating biological and pathological variation and discovering useful biomarkers. As of April 2003, 43 laboratories in 14 countries have committed to the Pilot Phase, utilizing HUPO reference specimens to evaluate a wide array of technology platforms.

3:00-3:30 Characterization of the Human Plasma Proteome
Dr. Joel Pounds, Senior Scientist, Biological Sciences, Pacific Northwest National Laboratory
The global characterization of blood plasma is complicated by the large dynamic range and protein diversity of blood plasma. In addition to chromatographic approaches, we have used Iodoacetyl PEO-biotin to reduce the complexity of trypsinized serum to cysteine-containing peptides. The sensitivity and high mass accuracy of the LC-ESI-FTICR-MS permits identification of greater than 100 serum proteins from a single microanalysis.

3:30-4:00 Refreshment Break with Poster and Exhibit Viewing

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