Cancer-initiating cells derived from established cervical cell lines exhibit stem-cell markers and increased radioresistance

Abstract

BackgroundCancer-initiating cells (CICs) are proposed to be responsible for the generation of metastasis and resistance to therapy. Accumulating evidences indicates CICs are found among different human cancers and cell lines derived from them. Few studies address the characteristics of CICs in cervical cancer. We identify biological features of CICs from four of the best-know human cell lines from uterine cervix tumors. (HeLa, SiHa, Ca Ski, C-4 I).MethodsCells were cultured as spheres under stem-cell conditions. Flow cytometry was used to detect expression of CD34, CD49f and CD133 antigens and Hoechst 33342 staining to identify side population (SP). Magnetic and fluorescence-activated cell sorting was applied to enrich and purify populations used to evaluate tumorigenicity in nude mice. cDNA microarray analysis and in vitro radioresistance assay were carried out under standard conditions.ResultsCICs, enriched as spheroids, were capable to generate reproducible tumor phenotypes in nu-nu mice and serial propagation. Injection of 1 × 103 dissociated spheroid cells induced tumors in the majority of animals, whereas injection of 1 × 105 monolayer cells remained nontumorigenic. Sphere-derived CICs expressed CD49f surface marker. Gene profiling analysis of HeLa and SiHa spheroid cells showed up-regulation of CICs markers characteristic of the female reproductive system. Importantly, epithelial to mesenchymal (EMT) transition-associated markers were found highly expressed in spheroid cells. More importantly, gene expression analysis indicated that genes required for radioresistance were also up-regulated, including components of the double-strand break (DSB) DNA repair machinery and the metabolism of reactive oxygen species (ROS). Dose-dependent radiation assay indicated indeed that CICs-enriched populations exhibit an increased resistance to ionizing radiation (IR).ConclusionsWe characterized a self-renewing subpopulation of CICs found among four well known human cancer-derived cell lines (HeLa, SiHa, Ca Ski and C-4 I) and found that they express characteristic markers of stem cell, EMT and radioresistance. The fact that CICs demonstrated a higher degree of resistance to radiation than differentiated cells suggests that specific detection and targeting of CICs could be highly valuable for the therapy of tumors from the uterine cervix.

Cervical Cancer Stem Cells Selectively Overexpress HPV Oncoprotein E6 that Controls Stemness and Self-Renewal through Upregulation of HES1

“…The identification of uterine cervical epithelial stem cells has been reported in cervical lesions and cancer cell lines (9,10); these studies, however, did not elucidate the role of HPV, an essential etiologic agent for the development of cervical cancer, particularly in exacerbating malignant progression and stemness properties.…”

“…Reports suggest preferential physical interaction of the virus with squamocolumnar junction (SCJ) in TZ that forms basal/reserve cells with stem cell properties (8). These observations along with technological advancements in identification of CSCs using established phenotypic and functional markers have facilitated identification and characterization of cervical CSCs from primary tumors (7,9) and cervical cancer cell lines (10). The cell lines, SiHa (HPV16 þ ) and HeLa (HPV18 þ ) constitutively express markers that closely resemble SCJ cells (8) thus making them a suitable experimental model for investigation of cervical CSCs in greater detail.…”

Human Embryonic Stem Cells - a Potential Vaccine for Ovarian Cancer

“…Unfortunately, targeting one antigen alone is unlikely to generate effective antitumor immune responses to mediate tumor rejection because of rapid appearance of escape mutants and the general inefficiency of monovalent cancer vaccines (Buonaguro et al, 2011;Durrant et al, 2011). Interestingly, it was found that cancer stem cells (CSCs) and embryonic stem cells (ESCs) shared similar cell surface markers and antigens not presented by adult tissues, which played a part in metastasis, angiogenesis and increased chemoradiotherapy resistance in cancer (Field et al, 2010;Kee et al, 2012;Lopez et al, 2012), so immune response against ESCs…”

IL-24 sensitizes tumor cells to TLR3-mediated apoptosis

“…Tumor spheres were prepared as previously described. 51 In brief, cells were detached with EDTA (Life Technologies, Carlsbad, CA, USA), washed with PBS and placed in serum-free medium mixed with 20 ng/ ml epidermal growth factor (EGF PHG0311L, Life Technologies), 20 ng/ml basic fibroblast growth factor (bFGF PHG0021, Life Technologies) and 0.4% bovine serum albumin (BSA, Sigma-Aldrich) at a density of 1 Â 10 3 cells/ml and cultured in ultra-low attachment plates (CLS3473 Costar, Sigma-Aldrich). The sphere culture medium was changed every 48 h until the majority of spheres reached 300-400 mm in diameter.…”