Differential gene expression in ovarian carcinoma: identification of potential biomarkers

Abstract

Ovarian cancer remains the fifth leading cause of cancer death for women in the United States. In this study, the gene expression of 20 ovarian carcinomas, 17 ovarian carcinomas metastatic to the omentum, and 50 normal ovaries was determined by Gene Logic Inc. using Affymetrix GeneChip HU_95 arrays containing approximately 12,000 known genes. Differences in gene expression were quantified as fold changes in gene expression in ovarian carcinomas compared to normal ovaries and ovarian carcinoma metastases. Genes up-regulated in ovarian carcinoma tissue samples compared to more than 300 other normal and diseased tissue samples were identified. Seven genes were selected for further screening by immunohistochemistry to determine the presence and localization of the proteins. These seven genes were: the beta8 integrin subunit, bone morphogenetic protein-7, claudin-4, collagen type IX alpha2, cellular retinoic acid binding protein-1, forkhead box J1, and S100 calcium-binding protein A1. Statistical analyses showed that the beta8 integrin subunit, claudin-4, and S100A1 provided the best distinction between ovarian carcinoma and normal ovary tissues, and may serve as the best candidate tumor markers among the seven genes studied. These results suggest that further exploration into other up-regulated genes may identify novel diagnostic, therapeutic, and/or prognostic biomarkers in ovarian carcinoma.

Figure 1

Dependence of the number of gene fragments present in all samples on the number of samples analyzed. The number of fragments present in all samples of a sample set is shown as a function of the number of samples. Normal ovaries (top), serous papillary ovarian carcinoma (middle), and serous papillary ovarian carcinoma metastatic to the omentum (bottom).

Figure 2

Differentially expressed genes in ovarian carcinoma, normal ovaries, and 20 other tissues. Eisen Cluster software was used to graphically display the intensity of gene expression values for each of the 40 genes listed in Table 2 for the 391 different tissue samples. The color of each square represents the ratio of the gene expression in the indicated sample relative to the average signal of expression of all genes examined. Red indicates gene expression above the median; black, equal to the median; and green, below the median. The intensity of the color reflects the magnitude of divergence from the median. Columns represent individual cDNAs for the 40 genes listed in Table 2, and rows represent the indicated tissue samples, as described in the Material and Methods section.

Figure 3

E-Northern analysis of differentially expressed gene fragments in human tissues. The expression of each indicated gene fragment was examined in 20 serous papillary ovarian carcinomas, 17 serous papillary ovarian carcinomas metastatic to the omentum, and 50 normal ovaries. The bar graph on the left depicts the percentage of samples that express detectable levels of the indicated gene fragment. The intensity of gene expression in each sample is plotted as average expression value on a linear scale on the right; median ± 2 SD of expression values are shown.

Figure 4

Immunohistochemical staining of differentially expressed gene products. Normal ovaries (A, D, G, J, M), serous papillary ovarian carcinoma (B, E, H, K, N), and serous papillary ovarian carcinoma metastatic to the omentum (C, F, I, L, O) tissues were stained with a mAb against the β1 integrin subunit (A–C), normal mouse IgG (D–F), and antibodies against: the β8 integrin subunit (G–I), BMP-7 (J–L), and claudin-4 (M–O). Original magnifications, ×60.

Figure 5

Immunohistochemical staining of differentially expressed gene products. Normal ovaries (A, D, G, J), serous papillary ovarian carcinoma (B, E, H, K), and serous papillary ovarian carcinoma metastatic to the omentum (C, F, I, L) tissues were stained with antibodies against: COL IX α2 (A–C), CRABP-1 (D–F), FOX J1 (G–I), and S100A1 (J–L). Original magnifications, ×60.