Abstract C2: Reactivation of Embryonic Stem Cell-Like Program in Human Cancer Cells by Diverse Manners

Abstract Embryonic stem cell (ESC)-like gene expression signature has been identified in multiple types of cancers and associated with high grade tumors and poor prognosis. The ESC signature is further linked with cancer stem/tumor initiating cells. However, it is unclear whether the ES signature reflects inheritance of a stem cell-of-origin or reactivation during the course of tumor progression. One recent study implicates the latter because c-Myc, but not the other tested oncogenes in the report, is sufficient to reactivate the ESC signature in normal and cancer cells. We aim to induce aggressive cancer phenotype in two well-differentiated human cancer cell lines, A549 of lung adenocarcinoma origin, and MCF-7 of breast cancer origin. In A549 cells, we generated two variants in which one was transduced with a backbone retroviral vector (A549neo) and the other was a clone overexpressing histone deacetylase 6 (HDAC6), a mediator of TGF-β1-induced EMT, upon retroviral transduction (A549HD6). In MCF-7 cells, we established a clone that acquired resistance to TNF-α-induced apoptosis upon chronic exposure (MCF-7TNR). We observed striking similarity between A549HD6 and MCF-7TNR whose appearance was drastically different from their respective parental lines. The appearance of A549HD6 and MCF-7TNR culture resembled ESC culture which features compact cell colonies and high nucleus/cytoplasm ratio. A549HD6 and MCF-7TNR exhibited alteration in cell cycle progression that was highlighted by low percentage of cycling cells in G-1 phase, a signature of ESC. Analysis of gene expression revealed loss of E-cadherin, an epithelial marker. Moreover, profiling stemness and pluropotency markers (TaqMan Human Stem Cell Pluropotency Array, Applied Biosystems) revealed elevated expression of several ESC stemness markers, such as Nanog and UTF1, in A549HD6 and MCF-7TNR. Fourteen out of twenty-two genes that were aberrantly activated in both A549HD6 and MCF-7TNR were targets of Polycomb Repressive Complex 2 (PRC2). Majority of the activated PRC2 targets were either markers of adult stem cells (GATA family members), or promoters or markers of cell types other than the origins of A549HD6 and MCF-7TNR. A549HD6 and MCF-7TNR exhibited greater tumorigenic property in vitro (soft agar assays) and in vivo (xenograft assays) than their counterparts. Furthermore, A549HD6 cell derived tumors displayed disorganized cell mass which was histologcially distinct from A549neo cell derived tumors that features glandular architecture, a signature histology of well-differentiated adenocarcinomas. In summary, we demonstrate reactivation of ESC-like program in well-differentiated human cancer cells. The ESC-like phenotype correlates with greater tumorigenic potential. Because ESC-like reprogramming occurs upon overexpression of EMT mediator or acquisition of apoptosis resistance, our results warrant further investigation of a link between ESC signature and EMT/apoptosis resistance in cancer. Citation Information: Cancer Res 2009;69(23 Suppl):C2.