9:00 Chair’s Remarks

9:05 Dynamic Arrays for High Efficiency Real-time qPCR 
Mike Lucero, EVP Sales and Marketing, Fluidigm Corporation
We have introduced dynamic arrays for real-time qPCR validation of microarray data against 100s to 1000s of samples. These nanofluidic chips automatically assemble 2,304 TaqMan assays in parallel and provide the same level of flexibility as microplates. Dynamic arrays also reduce time and running costs associated with validation studies.

9:35 Strategies for Patenting Gene Expression Data
John Thallemer, Senior Patent Attorney, Sandoz, Inc.
This talk will discuss the various types of gene expression data that are patentable, and methods for identifying valuable IP rights in the genetic data obtained using microarrays and chips. Strategies for obtaining patent coverage on gene profiles, SNPs and other genetic biomarkers will be presented. The patent strategies will focus on the use of gene expression data as markers for the presence, stage, origin and other factors associated with disease, therapeutic efficacy, and identifying responders and non-responders to a drug.

10:05 qRT-PCR Evaluation of Diagnostic Transcription Markers of Cervical Neoplasia: A Clinical Study
Martin Steinau, Ph.D., Research Scientist, VEHB, Centers for Disease Control and Prevention The presentation would illustrate how transcription biomarkers for the diagnosis of high grade cervical intraepithelial neoplasia in 179 clinical cytology samples were evaluated by qRT-PCR in a population based study. It will be detailed how controls for various aspects of experimental errors were applied, i.e. verification for contaminating DNA in cDNA prepared from total nucleic acid (RNA and DNA) samples, monitoring of RT efficiency differences through an external control, empirical determination of a reference for normalization and detection of unspecific signals. Finally it will be shown how quality controls were set and how the raw data were analyzed by ROC to determine the diagnostic value of transcription levels in case and control groups.

10:35 Coffee Break, Poster and Exhibit Viewing

11:15 Effective Application of Microarray Expression Profiling and Quantitative PCR Assays to Understand Drug Action Mechanisms in Animal Models
Herbert Auer, MS, Director of the Functional Genomics Core, Columbus Children’s Research Institute
Microarray expression profiles, quantitative PCR assays, histopathological data and clinical scores are often difficult to combine and interpret. A major source of difficulty is noisy and biased molecular data. Here we describe a successful combined analysis of molecular and clinical data sets generated from an inflammatory bowl disease model for the protective effects of a novel drug. Guidelines are presented for the effective performance of microarray expression profiling, data analysis and quantitative PCR assays to obtain a coherent characterization of drug action mechanisms. 

11:45 Functional Validation of Gene Dysregulation in Insulin Resistance
Margaret C. Cam, Ph.D., Director, Genomics Core Laboratory, NIDDK, NIH 
The discovery of key regulatory mechanisms underlying a complex disease phenotype is highly dependent on the accurate and quantitative profiling of the functional genome. We have used a variety of algorithms in an attempt to extract the most accurate set of genes showing moderate changes in expression in adipose cells derived from insulin-resistant subjects with a strong family history of Type 2 diabetes. We have found that both gene selection and relative enrichment of known gene ontology categories are algorithm-dependent, and have validated selected genes by RT-PCR. The plethora of algorithms for the Affymetrix platform are a source for the instability of gene expression results and must be considered carefully especially when sample size is small.

12:15 Lunch on Your Own (Technology Workshop Sponsorship Available)

1:45 Recognizing Apples from Oranges: Strategies for Validating Transcript Data in Biopharmaceutical Development
Eric R. Fedyk, Ph.D., Senior Scientist II, Drug Safety Evaluation, Millennium Pharmaceuticals, Inc.
Valuable biomarkers identified in discovery efforts often fail to impact downstream clinical programs. A number of variables can prevent translation of biomarkers from discovery to utilization, some of which are controllable, whereas others are uncontrollable. A critical activity for successful translation is to identify these variables, control them as best possible, annotate those that are uncontrollable, and analyze data accordingly.

2:15 Measuring Oncogene Amplification in FFPE Tumor Samples by TaqMan Real-Time PCR
Jason Yang, MD, Ph.D., Senior Principal Scientist, Cancer and Immunology Biomarkers, Pfizer, Inc.
Recurrent gene amplification is a hallmark of cancer. This genetic alteration causes gene copy gain of oncogenes that results in increased level of the respective mRNA and protein in cells, leading to tumorigenesis and cancer progression. Although FISH has been widely used in detecting oncogene amplification there are several limitations for FISH technology including difficulties in reproducible detection and objective scoring of low or very high levels of amplification, and requirement for preservation of histological details. In addition, FISH is not feasible for interrogating amplification status of multiple genes because of limited availability of FFPE tissue slides. We have developed an accurate assay to measure copy changes of multiple genes from limited amounts of genomic DNA derived from FFPE tumor slides using TaqMan real-time PCR technology.

2:45 Analytical Validation of RNA External Standard qRT-PCR Assays for Use In Clinical Studies
Michael Burczynski, Ph.D., Principal Scientist, Clinical Translational Medicine, Wyeth
With the increasing number of biomarkers identified in transcriptional profiling studies, there is an accompanying demand for translation of these findings into highly quantitative assays that can be used to assess these biomarkers in collected clinical samples. The present talk will summarize an approach to the analytical validation process for RNA transcripts that will be evaluated as pharmacodynamic biomarkers in clinical studies. Analytical aspects of qRT-PCR validation that will be covered include determination of assay specificity, sensitivity, linearity, accuracy, efficiency, dynamic range, and the establishment of upper and lower limits of quantitation for the target gene(s) of interest. In addition, the importance of characterizing reference ranges, implementing quality controls and conducting storage stability studies will also be highlighted.

3:55 Enabling CROs to Provide Quality-Controlled Data That Support Development of New Pharmaceuticals
James C. Willey, MD, Professor of Medicine and Pathology, Medical College of Ohio, Chief Scientific and Medical Consultant to Gene Express, Inc.
After biotechnology or pharmaceutical companies discover promising pharmaceuticals, the candidates must be taken through several stages of testing. Much of this testing is now outsourced to contract research organizations (CROs). In order to support regulatory review of a New Drug Application (NDA), the CRO must generate data on an analytical platform that can demonstrate several performance characteristics specified by the FDA. Gene Express, Inc. has focused on developing a platform technology, StaRT-PCR, that has each of these FDA specified performance characteristics. CRO implementation of StaRT-PCR in Standardized Expression Measurement (SEM) Centers will be discussed.

4:25 Development of qRT-PCR Assays for use in Monitoring Viremia in a Clinical Trial Setting
Linda Starr-Spires, Ph.D., Deputy Director, Nucleic Acid Methods Platform, Assay Development and Clinical Testing, Global Clinical Immunology Department, sanofi pasteur
Assays intended for use in clinical trial monitoring are bound by restrictions unique to the setting. In developing qRT-PCR assays for such applications, it is necessary to consider not only typical development issues, but also take into account such things as workflow restrictions, increasing sample numbers, short turn around times, and regulatory requirements. Additionally, the assays are frequently directed at targets for which there are no commercial or compendial reagents available, requiring in-house manufacture and qualification of reagents by the development group. Approaches and solutions to these issues will be discussed. 

4:55 Panel Discussion with Afternoon Speakers