Differential gene expression profiling of cultured neu-transformed versus spontaneously-transformed rat cholangiocytes and of corresponding cholangiocarcinomas

Previously, we described an orthotopic cholangiocarcinoma model based on bile duct inoculation of spontaneously-transformed low grade malignant rat BDE1 cholangiocytes (BDEsp cells) compared to high grade malignant erbB-2/neu- transformed BDE1 cholangiocytes (BDEneu cells) into the livers of syngeneic rats, which closely mimics clinical features of early versus advanced stages of the human cancer. We now used gene expression microarray together with quantitative real-time RT-PCR to profile genes differentially expressed in highly tumorigenic BDEneu cells and corresponding tumors compared to less aggressive tumorigenic BDEsp cells and tumors. Genes identified as being commonly overexpressed in parent BDEneu cells, tumors, and in a BDEneu tumor-derived cholangiocarcinoma cell line included Sox17, Krt20, Erbb2, and Sphk1 when respectively compared to BDEsp cells, tumors, and tumor-derived BDEsp cholangiocarcinoma cells. Muc1 was also prominently overexpressed in BDEneu cells and tumor-derived cholangiocarcinoma cells over that expressed in corresponding BDEsp cell lines. Periostin and tenascin-C, which were produced exclusively by cholangiocarcinoma-associated fibroblastic cells, were each significantly overexpressed in BDEneu tumors compared to BDEsp tumors. Interestingly, amphiregulin was representative of a gene found to be significantly underexpressed in vitro in BDEneu cells compared to BDEsp cells, but significantly overexpressed in BDEneu tumors compared to BDEsp tumors, and correlated with BDEneu cholangiocarcinoma progression in vivo. Our data support a unique animal model that recapitulates important molecular features of human cholangiocarcinoma progression, and may serve as a potentially powerful preclinical platform for identifying and rapidly testing novel molecular targeting strategies for cholangiocarcinoma therapy and/or prevention.

Research highlights
► Provides first comprehensive gene expression analysis of a unique orthotopic cholangiocarinoma rat syngeneic model that closely mimics pathological and clinical features of human cholangiocarcinoma progression.
► First demonstration of Sox17 (Sry-related HMG box gene-17) and Sphk1 ( sphingosine kinase 1) overexpression in cholangiocarcinoma cells correlated with increased hepatobiliary cancer cell malignancy.
► First to identify caveolin-1 as a putative molecular target with possible relevance to cholangiocarinoma progression.
► First to demonstrate Postn (periostin), a gene associated with the extracellular matrix gene ontogeny, to be significantly overexpressed in highly tumorigenic orthotopic rat cholangiocarcinomas versus tumors resulting from less malignant transformed rat cholangiocytes.
► Identifed select gene targets differentially expressed in cholangiocarcinoma cells of differing malignant potential, as well as in cancer-associated fibroblastic cells, which are suggested to be planning a role in cholangiocarcinoma progression, and which may serve as potential cancer and tumor stromal cell targets for cholangiocarcinoma therapy.