Research article
Generation of tumor-initiating cells by exogenous delivery of OCT4 transcription factor
-
* Corresponding author: Pilar Blancafort pilar_blancafort@med.unc.edu
1 Department of Pharmacology, The University of North Carolina at Chapel Hill, 120 Mason Farm Road, Chapel Hill, NC 27599, USA
2 UNC Lineberger Comprehensive Cancer Center, The University of North Carolina at Chapel Hill, 450 West Drive, Chapel Hill, NC 27599, USA
3 Department of Biochemistry and Biophysics, The University of North Carolina at Chapel Hill, 120 Mason Farm Road, Chapel Hill, NC 27599, USA
4 State Key Laboratory of Molecular Oncology, Cancer Hospital/Institute, Chinese Academy of Medical Sciences, Pan Jia Yuan Nan Li 17, Chaoyang District, Beijing, 100021, P.R. China
5 Department of Pathology and Laboratory Medicine, The University of North Carolina at Chapel Hill, 515 Brinkhous-Bullitt Building, Chapel Hill, NC 27599-7365, USA
Breast Cancer Research 2011, 13:R94 doi:10.1186/bcr3019
Published: 27 September 2011Abstract
Introduction
Tumor-initiating cells (TIC) are being extensively studied for their role in tumor etiology, maintenance and resistance to treatment. The isolation of TICs has been limited by the scarcity of this population in the tissue of origin and because the molecular signatures that characterize these cells are not well understood. Herein, we describe the generation of TIC-like cell lines by ectopic expression of the OCT4 transcription factor (TF) in primary breast cell preparations.
Methods
OCT4 cDNA was over-expressed in four different primary human mammary epithelial (HMEC) breast cell preparations from reduction mammoplasty donors. OCT4-transduced breast cells (OTBCs) generated colonies (frequency ~0.01%) in self-renewal conditions (feeder cultures in human embryonic stem cell media). Differentiation assays, immunofluorescence, immunohistochemistry, and flow cytometry were performed to investigate the cell of origin of OTBCs. Serial dilutions of OTBCs were injected in nude mice to address their tumorigenic capabilities. Gene expression microarrays were performed in OTBCs, and the role of downstream targets of OCT4 in maintaining self-renewal was investigated by knock-down experiments.
Results
OTBCs overcame senescence, overexpressed telomerase, and down-regulated p16INK4A. In differentiation conditions, OTBCs generated populations of both myoepithelial and luminal cells at low frequency, suggesting that the cell of origin of some OTBCs was a bi-potent stem cell. Injection of OTBCs in nude mice generated poorly differentiated breast carcinomas with colonization capabilities. Gene expression microarrays of OTBC lines revealed a gene signature that was over-represented in the claudin-low molecular subtype of breast cancer. Lastly, siRNA-mediated knockdown of OCT4 or downstream embryonic targets of OCT4, such as NANOG and ZIC1, suppressed the ability of OTBCs to self-renew.
Conclusions
Transduction of OCT4 in normal breast preparations led to the generation of cell lines possessing tumor-initiating and colonization capabilities. These cells developed high-grade, poorly differentiated breast carcinomas in nude mice. Genome-wide analysis of OTBCs outlined an embryonic TF circuitry that could be operative in TICs, resulting in up-regulation of oncogenes and loss of tumor suppressive functions. These OTBCs represent a patient-specific model system for the discovery of novel oncogenic targets in claudin-low tumors.