8:30 Chairperson's Opening Remarks
Dr. James L. Wittliff, Professor of Biochemistry & Molecular Biology, James Graham Brown Cancer Center, University of Louisville

8:40 Challenges in the Preparation of Clinical Samples for Genomic and Proteomic Analysis 
Dr. Scott D. Patterson, Senior Director, Medical Sciences, Amgen Inc.
Obtaining samples from clinical trials for subsequent genomic and proteomic-based studies can be challenging. Assays that perform well in the research laboratory setting can pose problems when translated to the clinical setting. It may be that a portion of the assay has to be conducted at the clinical site and the remainder in your own laboratory. This presentation will discuss some examples of such assays and how they were conducted successfully.

9:20 Effective BioBank Management in an Age of Uncertainty
Dr. Richard Hegele, Professor and Head, Department of Pathology and Laboratory Medicine, James Hogg iCAPTURE Centre, Vancouver
Biobanks and other types of biological repositories are a tremendous resource not only for current research but for future research in areas we cannot even begin to imagine at present. At the management, operations and regulatory levels, this presents a number of challenges for continued use of previously obtained specimens, in light of new privacy legislation and other developments. This presentation will review how the James Hogg iCAPTURE Centre has addressed these challenges, by harmonizing its separate cardiovascular and pulmonary tissue registries, its consent processes in light of new legislation, and activities in quality assurance.

8:30 Chairperson's Opening Remarks

8:40 Triage for Predicting Clinical Outcome using Molecular Profiling of Human Carcinoma Cells Procured by LCM 
Dr. James L. Wittliff, Professor of Biochemistry & Molecular Biology, James Graham Brown Cancer Center, University of Louisville
Forecasting clinical behavior and therapeutic response of human cancer currently utilizes a few tumor markers and disease characteristics. Medically relevant genomic and proteomic test development requires a troika consisting of human tissue specimens collected and stored under specialized conditions with annotated clinical records, isolation of distinct cell types with technologies such as laser capture microscopy and analyses of macromolecules with protease and RNase-free protocols to generate gene and protein expression profiles. Using an extensive Biorepository of frozen and formalin-fixed, paraffin-embedded tissue specimens with long-term clinical follow-up, pure populations of cells were procured by LCM for RNA extraction, amplification and microarray. Zinc-finger binding proteins, analyzed as models for proteomics, were identified using 32P-labeled hormone response elements and band-shift assays. Genes associated with estrogen receptor-alpha (ER) status dominated expression profiles suggesting a larger hormone-related network than previously appreciated. Other prominent gene expression patterns divided ER+ carcinomas into two subgroups, each associated with significantly different clinical outcomes. Another gene expression signature was identified strongly correlated with breast cancer recurrence using supervised learning methods. This large retrospective investigation of pure carcinoma cell populations identified molecular signatures highly associated with clinical outcome suggesting promise for development of novel prognostic tests for breast cancer management and identification of new molecular targets for drug design.

9:20 Cells, Microchips, and Clinical and Biological Information
Dr. Mehmet Toner, Professor of Biomedical Engineering, Harvard Medical School, Massachusetts General Hospital
Microfabrication techniques that have revolutionized the electronics industry, are now poised to revolutionize the pharmaceutical, biotechnology, and biomedical device industries. The two leading applications of microfabrication in biology include "genes-on-a-chip" to monitor the expression level of potentially all genes in humans or various model systems and organisms simultaneously, and "lab-on-a-chip" type devices to perform biochemistry in microchambers. Equally exciting is the fact that biomedical application of microfabricated devices is no longer limited to non-living systems such as genes-on-a-chip or lab-on-a-chip. Recent advances in the understanding of cellular behavior in microenvironments have started to pave the way towards living micro-devices. The ability to integrate cells with microdevices is important for controlling cellular interactions on a sub-cellular level, for obtaining highly parallel, statistically meaningful readouts over large cell populations, and for miniaturizing instrumentation towards minimally-invasive, portable, fast, inexpensive devices. The rich assortment of living microdevices that can be built using BioMEMS techniques have important applications in fundamental cell biological studies, tissue engineering, cell separation and culture devices, diagnostics, and high-throughput drug screening tools. This presentation will provide an overview of some of the key advances and remaining challenges related to the combination of microfabrication technology and living cells and a sampling of a broad spectrum of exciting opportunities and promises in biology and medicine.

Option 1:
Image Analysis

2:00 Chairperson's Remarks 

2:05 The Use of Tissue Microarrays in the Human Proteome Resource Project
Dr. Caroline Kampf, Senior Scientist, Department of Genetics and Pathology, Rudbeck Laboratory, Uppsala University
Antibody-based proteomics provides a powerful approach for functional studies of the human proteome. We have developed a strategy for systematic generation of protein-specific affinity reagents (mono-specific antibodies) and utilized these to create a comprehensive atlas containing protein expression patterns from 48 normal human tissues and 20 different types of cancer (www.proteinatlas.org). A set of standardized tissue microarrays (TMAs), including a total of 708 spots from 372 different individual patients, was produced to allow for rapid tissue and cell profiling using immunohistochemistry. By combining immunohistochemically stained TMAs with image analysis, objective and reproducible data including fraction of positive cells, staining intensity and subcellular localization can be obtained. Image analysis also provides possibilities for a relative quantification of protein expression in defined cell populations, allowing for a comparison with transcription levels from cDNA array data. 

2:35 The Importance of Automated Image Capture and Analysis in High- Throughput Gene Expression Projects
Dr. Tony Warford, Senior Project Leader, Atlas of Protein Expression Project, The Wellcome Trust Sanger Institute
Several high throughput gene expression projects are underway that will provide a wealth of morphological information. One of these is the Atlas of Protein Expression Project (http://www.sanger.ac.uk/Teams/Team86/ ). Antibodies are selected on protein fragments using phage display technology and then screened on TMA and tissue sections to provide comprehensive landscaping of protein distribution at the cellular level. To create this, all image data is automatically captured and analysed using the Ariol® automated imaging and image analysis system by Applied Imaging Corp. Images of chromogenic and fluorescence slides are captured whilst quantitative and qualitative analysis tools provide objective data and aid interpretation. Up to 60 slides can be processed in 24h at a resolution of 0.377m per pixel when using the x20 objective. This provides for an output of up to 500,000 TMA core images per month. Annotation is undertaken within the Atlas Oracle database using drop down menus that are governed by ontology tables. Given the prestigious amount of image data being produced by this and other gene expression projects there is now an urgent need to automate annotation. Whilst technically feasible the development of algorithms will require the concerted international effort of morphologists.

3:05 Image Analysis: A High Throughput Solution for Biomarker Expression and Genetic Alteration Analysis using Tissue Microarrays
Dr. George J. Netto, Staff Pathologist, Division of Surgical Pathology, Johns Hopkins Medical Institutions
Accurate quantitative evaluation of a particular biomarker expression in a high density TMA's can be a daunting effort.  Although direct real time microscopic evaluation of tissue spots in smaller TMA's can be performed, measuring extent and intensity of immunohistochemical expression is subject to inter and intraobserver variations. Current image analysis technology offers a great solution for image capturing, image archiving as well as automated quantitation of biomarker expression. The technology can further lend itself to the evaluation of genetic alterations by FISH on TMA sections. Acquired data can be exported to existing databases allowing for correlation with clinical and or pathologic parameters. 

Option 2:
Preparing Genomic Samples

2:00 Chairperson's Remarks

2:05 Quantitative Gene Expression Results from Fixed Tissue without Specialized Sample Preparation 
Dr. Bruce Seligmann, CEO and President, HTG
The measurement of gene expression from fixed tissue has been at best problematic, time consuming, and cumbersome. However, using gene expression assay technology, fixed tissue can be dissolved in lysis reagent and tested using the same simple protocol that is used for cells or frozen tissues, and it provides the same quantitative measurements of gene expression levels. Data from matched frozen and fixed tissue will be presented, including experiments where tissue fixed years earlier is compared to freshly fixed tissue from the matched frozen block. The multiplexed format of the technology permitting up to 16 genes to be measured in each well of a microplate, and the throughput of both the qNPA sample processing and the measurement, means that extensive retrospective studies are now possible to validate gene signatures, targets for drug discovery, and biomarkers for diagnostic applications. The impact of this breakthrough on the target validation and drug discovery process will be discussed.

2:35 Nanoparticle-Based DNA Purification for PCR
Dr. Michael Hogan, Managing Director, Scientific Affairs, Argylla Technologies LLC
We describe here a simple, inexpensive and highly efficient form of batch chromatography, based on chemically coated,sub-micron ceramic particles (nanoparticles) for the capture, purification and concentration of DNA for PCR-based genomics. Because of their sub-micron size, coated ARGYLLA nanoparticles present an enormous surface area to volume ratio (100cm2/uL) which generates a high DNA binding capacity (2mg DNA/uL). The nanoparticles are small enough that they form a stable suspension at 1g (where they can bind DNA effectively) but because they are dense, they are readily harvested by low speed centrifugation (2000g) to produce a discrete, white, microliter pellet, from which captured, purified DNA may be released at room temperature into a volume as small as 5uL: as a PCR-ready solution. Applications are described where chemically coated Argylla nanoparticles are used to isolate DNA for high-value applications: forensic database building (cheek wipes) crime scene analysis (surface wipes) population genetics (dried blood on FTA paper) and clinical genetics (fresh blood, paraffin-embedded thin sections). As assessed by quantitative PCR, both DNA yield and quality are found to equal or exceed that obtained by the use of industry-standard spin columns or magnetic bead products.

3:05 The Collection, Archiving and Isolation of DNA from Dried Clinical Samples for Molecular Analysis
Dr. Michael Harvey, Principal Scientist, R&D, Whatman
The collection and preparation of clinical samples for molecular analysis is important for expanding applications in drug discovery, predictive medicine and pharmacogenomics. Whatman has developed a novel technology, FTA and FTA Elute, for dried clinical sample collection, archiving and, most importantly, DNA preparation for amplification. With this technology sample preparation from samples collected on treated samples is simple, rapid and automatable. Blood and buccal cell samples collected on FTA® and FTA® Elute can be stored for long periods of time under ambient conditions. This presentation will review studies that will demonstrate the use of DNA from the aforementioned types of samples for forensic identification, real time PCR, DNA sequencing and allele specific hybridization methods.

Featured Presentations

4:15 Chairperson's Remarks

4:20 Molecular Maps of Prostate Cancer: Tissue Print Micropeel Profiles of Human Surgical and Biopsy Specimens
Dr. Sandra Gaston, Principal Investigator, Department of Surgery, Beth Israel Deaconess Medical Center
Recent advances in molecular technologies have led to acceleration in the identification potential biomarkers for human cancers. To realize the full potential of this wealth of new biomarkers in the development of targeted therapies for solid tumors, it is essential that we develop new strategies for profiling human tissue specimens that are compatible with the demands and restrictions of the clinical setting. My laboratory has developed a set of novel tissue print micropeel techniques that allow us to map a series of molecular marker profiles from an extended area of a human tissue sample without damaging the specimen (Gaston et al. Tissue-Print and Print-Phoresis as Platform Technologies for the Molecular Analysis of Human Surgical Specimens: Mapping Tumor Invasion of the Prostate Capsule. Nat Med. 2005;11(1):95-101). Because our tissue print platform supports both proteomic analysis and PCR-based DNA and mRNA profiling techniques, it can be used to generate detailed molecular cross-sections that can be layered directly over corresponding histological and radiological images. Such extensively annotated maps of human tissue and tumor specimens can be used for target-based tumor classification and for monitoring molecular responses to target-based therapies. I propose, in my talk, to present an overview of our tissue print micropeel technologies, with specific examples of the application of these technologies in the molecular assessment of human tissue biopsy and surgical specimens.

4:55 RNA Quality and Yields from Frozen Tissues: Analysis of Samples from a Large International Repository
Dr. James Eliason, Chief Scientific Officer, Research and Development, Asterand, Inc.
Frozen tissue repositories are an important resource for genomics and proteomics research. When repositories contain samples from multiple sites, it is important to have ways to assess the quality of the samples. RNA is highly sensitive to handling and freezing conditions. We have routinely performed an RNA quality control test for every sample we have received. We have compared several different measures for RNA quality including the RNA Integrity Number proposed by Agilent. We have also examined several factors that affect RNA quality and yields.