Systematic analysis and validation of differential gene expression in ovarian serous adenocarcinomas and normal ovary

Abstract

Purpose: Cancer of the ovary confers the worst prognosis among women with gynecological malignancies, primarily because most ovarian cancers are diagnosed at late stage. Hence, there is a substantial need to develop new diagnostic biomarkers to enable detection of ovarian cancer at earlier stages, which would confer better prognosis. In addition, the identification of druggable targets is of substantial interest to find new therapeutic strategies for ovarian cancer.

Methods: The expression of 22,500 genes in a series of 67 serous papillary carcinomas was compared with 9 crudely enriched normal ovarian tissue samples by RNA hybridization on oligonucleotide microarrays. Multiple genes with near-uniformly expression were elevated in carcinomas of varying grade and malignant potential, including several previously described genes (e.g., MUC-1, CD9, CD24, claudin 3, and mesothelin). We performed immunohistochemical staining with antibodies against several of the proteins encoded by differentially expressed genes in an independent cohort of 71 cases of paraffin-embedded ovarian cancer samples.

Results: We found striking differences in EpCAM (p < 0.005), CD9 (p < 0.001), MUC-1 (p < 0.001), and claudin 3 proteins (p < 0.001) but not for mesothelin (p > 0.05) using the Mann-Whitney U test.

Conclusions: Protein expression of a majority of the differentially expressed genes tested was found to be elevated in ovarian carcinomas and, as such, define potential new biomarkers or targets.

Fig. 1

Visualization of the relative expression of genes predicted to be differentially expressed in normal and malignant ovarian tissue. Gene expression was normalized to mean of 0 and a variance of 1. C > N, cancer expression > normal expression; N > C: normal expression > cancer expression

Fig. 2

Box plot depiction of protein expression levels based on immunoscores (0–300); a mesothelin (p > 0.05) (n = 70), b CD-9 (p = 0.004) (n = 70), c Muc-1 (p < 0.001) (n = 71), d EpCAM (p < 0.001) (n = 69), e claudin 3 (p < 0.001) (n = 65). Black horizontal line mean value; gray rectangle 25th–75th percentile. ° outlier >1.5 inter-quartiles range, * outlier >3 inter-quartiles range, meso Mesothelin, muc Muc-1, clau Claudin 3

Fig. 3

Immunohistochemical staining of formalin-fixed and paraffin-embedded ovarian cancer tissue (tissue micro array) and normal ovarian tissue at 10- and 40-fold magnification

Fig. 4

Box plot depiction of protein expression levels based on immuno scores (0–300) within different histological subtypes; a CD9 (p = 0.029), b Muc-1 (p = 0.001), c EpCAM (p < 0.001), d claudin 3 (p = 0.003). Black horizontal line mean value; gray rectangle 25th–75th percentile. ° outlier >1.5 inter-quartiles range, * outlier >3 inter-quartiles range. Papillary serous ovarian cancer = papser (n = 51). Endometrial ovarian cancer = end (n = 10). Solid ovarian cancer = solid (n = 10). Normal ovarian tissue = NT (n = 6). meso Mesothelin, muc Muc-1, clau Claudin 3