Gene expression profiling of microdissected stroma from normal and malignant ovary identifies a role for CTGF in ovarian tumor progression.

A75

It is estimated that in 2007 there will be approximately 22,430 new cases and 15,280 deaths due to ovarian cancer in the US. This high fatality rate is due in part to late diagnosis of advanced-stage disease which has already spread beyond the ovary. A majority of ovarian tumors arise from the surface epithelium of the ovary. While cell signaling between ovarian surface epithelial cells and their supporting stromal cells is required to maintain normal function of the ovary, the signaling events between ovarian tumor epithelial cells and their surrounding stromal cells remain unclear. The aim of this study was to investigate alterations in gene expression that occur in tumor-derived versus normal stromal cells. We analyzed gene expression of 51 laser-capture microdissected tumor and 10 normal stromal samples by Affymetrix microarrays. BRB-ArrayTools software (NCI) was used to analyze the data. Unsupervised hierarchical clustering analysis clearly distinguished tumor-associated stroma from normal ovarian stroma. Supervised class comparison analysis identified 2,703 probe sets (p<0.001) that were differentially expressed between tumor-associated and normal stroma. Permutation analysis ensured that the resulting gene list contained no more than 10 false discoveries at a 95% confidence interval. To validate the array results, 9 differentially expressed genes were selected for real-time PCR analysis across all of the stromal samples. Eight of the 9 were found to be significantly up-regulated in tumor-associated stroma by 2.1- to 34.4-fold over normal stroma (89% validation rate). Analysis of the entire differentially expressed gene list identified the α_vβ₃ integrin pathway as activated in the tumor-associated stromal isolates. Biological studies of over-expression of CTGF, a member of the α_vβ₃ integrin pathway, in the ovarian cancer cell line A547 revealed increased activity of processes involved in tumor progression. Compared to controls, CTGF-transfected clones demonstrated higher proliferation rates (32.0 ± 5.5 versus 42.8 ± 14.0 hrs), anchorage-independent growth (20.7 ± 11.6 versus 5.6 ± 1.8 colonies), migration (71.8 ± 38.7 versus 6.2 ± 3.5 cells) and invasion (212.4 ± 7.2 versus 1.9 ± 1.5 cells). Our study is the first to measure gene expression in microdissected stromal ovarian cancer samples and demonstrates that stroma undergoes changes in gene expression in response to the epithelial tumor microenvironment, and that these changes contribute to tumor progression.