The members of the Ovarian Cancer Demonstration Project include 10 investigators from 9 universities and institutes including the Dana Farber Cancer Institute, the Ontario Cancer Institute, Harvard, Johns Hopkins, MIT, Stanford, UCLA, University of Colorado and USC. The members of Dr. Dennis Slamon's laboratory will perform microarray based transcriptional analyses of 225 ovarian lesions including 175 tumor samples with clinical follow up information and correlate the results with clinical parameters and behavior. All of the potential relevant and/or informative alterations found using the Agilent transcriptome-wide analysis of the ovarian cancer cohort will be evaluated and scrutinized with data derived from the DNA array comparative genomic hybridization analyses designed to look for specific regions of gene amplification or deletion. These assays will be carried out in Dr. Mak's laboratory, which will perform analyses on DNA obtained from these exact same lesions so that an effective "across-platform" comparison can be performed. Dr. Baylin will utilize new high throughput techniques to examine the extent of histone modifications and DNA methylation patterns around promoter regions, since these epigenetic processes can lead to gene silencing within tumors. In addition, a panel of 35 established human ovarian epithelial cancer cell lines will undergo the same RNA and DNA analyses. This will allow the APCR investigators to determine if the spectrum of observed alterations (or any specific alteration) is recapitulated in any of these laboratory models.
These ovarian cancer cell lines will be simultaneously characterized for their in vitro (both on plastic and in soft agar) and in vivo growth characteristics in the laboratory of Dr. Brugge. Her lab will also develop organotypic, 3D models as well as cultures of tumors cells isolated from fluid exudates (e.g. ascites or pleural fluid). These cell culture models will be used to evaluate the role of genes implicated in ovarian carcinogenesis by the APCR ovarian tumor studies. The goal of these studies is to identify key components of the pathways that are critical for tumor cell proliferation and survival and those that confer resistance to known chemotherapeutics or those in the pipeline. Given the known high frequency of alterations in the PI3 kinase pathway, the group will focus significant efforts on this pathway. Dr. Velculescu will use advanced, high throughput, DNA sequencing methodologies to examine a panel of ovarian tumors for mutations in components of the PI3 kinase pathway. Pathway dissection will involve approaches to impose blocks (siRNA, dominant negative variants) or bypasses (constitutively active mutants) to pathway components in order to assess how the pathway operates, as well as which proteins are critical for the function of the pathway. Dr. Levy's studies map phosphoprotein responses to cytokines and examine basal phosphorylation states at the individual cell level. Flow cytometry will also reveal underlying dysregulated signaling nodes in primary human ovarian tumors and identify pathological tumor signaling profiles. This information will be closely evaluated by Drs. Brugge and Jacks to develop, utilize and scientifically exploit additional laboratory models for all of the APCR Demonstration Project investigators to use in the validation process of new translational therapeutic approaches. Any genes/pathways identified in these studies that may be of particular interest will be targeted by Dr. Press to develop a useful and effective molecular pathology tissue-based assay to detect key alterations in clinical material.
A comprehensive analysis of the relative expression levels of steroid hormone receptors, the known cofactors associated with their transcriptional activity, and the downstream effector genes/pathways which they activate will be performed by Dr. Press. These studies will be done using the cell lines, animal models and tissue resources, as well as the analytic platforms common to the Demonstration Project. Should the molecular analysis of these tissue specimens give a signal implicating any of the steroid hormone receptor pathways in a given subtype of ovarian cancer, he will work closely with Drs. Brugge and Jacks to attempt to develop and identify a relevant model for this subtype. The model(s) will then be used for preclinical exploratory studies involving blockade of the suspect steroid hormone receptor pathway. If the preclinical models demonstrate that blockade of a specific steroid hormone receptor pathway results in growth inhibition, assays suitable for identification and characterization of the critical molecular markers of these pathways will be developed and used to identify specific ovarian cancer patients (subgroups) whose tumors have a steroid receptor pathway signature associated with the potential for "responsiveness" to hormone receptor blockade.
Drs. Demetri and Bunn's studies will be designed to translate the molecular advances of the APCR-funded investigative efforts into rational and innovative clinical trials. They propose to implement a scientifically rigorous clinical trial with applicability to ovarian cancer based on a novel targeted therapeutic strategy and anticipate using the PI-3-Kinase and the IGF-IGF-R pathways as leading targets for future translational and clinical trials. Additionally, they will generate correlative scientific studies using tissue samples obtained from patients participating in the APCR-focused translational and clinical research.