Gene expression and functional evidence of epithelial-to-mesenchymal transition in papillary thyroid carcinoma invasion

Abstract

Papillary thyroid carcinomas (PTCs) that invade into local structures are associated with a poor prognosis, but the mechanisms for PTC invasion are incompletely defined, limiting the development of new therapies. To characterize biological processes involved in PTC invasion, we analyzed the gene expression profiles of microscopically dissected intratumoral samples from central and invasive regions of seven widely invasive PTCs and normal thyroid tissue by oligonucleotide microarray and performed confirmatory expression and functional studies. In comparison with the central regions of primary PTCs, the invasive fronts overexpressed TGF ␤, NFB and integrin pathway members, and regulators of small G proteins and CDC42. Moreover, reduced levels of mRNAs encoding proteins involved in cell-cell adhesion and communication were identified, consistent with epithelial-to-mesenchymal transition (EMT). To confirm that aggressive PTCs were characterized by EMT, 34 additional PTCs were examined for expression of vimentin, a hallmark of EMT. Overexpression of vimentin was associated with PTC invasion and nodal metastasis. Functional, in vitro studies demonstrated that vimentin was required both for the development and maintenance of a mesenchymal morphology and invasiveness in thyroid cancer cells. We conclude that EMT is common in PTC invasion and that vimentin regulates thyroid cancer EMT in vitro.cdc42 ͉ runx2 ͉ thyroid cancer ͉ vimentin T hyroid carcinoma is the most common classical endocrine malignancy, and its incidence is rising rapidly, due almost entirely to an increase in papillary thyroid carcinoma (PTC) diagnoses (1). Patients diagnosed with PTC at an early stage have an excellent prognosis; however, individuals with large, invasive tumors and/or distant metastases have a 5-year survival rate of Ϸ40% (2, 3). Thus, there is a need to better understand the molecular causes of thyroid cancer progression to develop new treatment options.The genetic defects believed to be responsible for PTC initiation have been identified in the majority of cases; these include genetic rearrangements involving the tyrosine kinase domain of RET and activating mutations of BRAF and RAS (3-5). Although some correlation studies support an association between specific genetic alterations and aggressive cancer behavior (6-9), there are a number of events that are found nearly exclusively in aggressive PTCs, including mutations of P53 (10, 11), dysregulated ␤-catenin signaling (12), up-regulation of cyclin D1 (13), and overexpression of metastasis-promoting, angiogenic, and/or cell adhesion-related genes (14-20). We have determined that invasive regions of primary PTCs are frequently characterized by enhanced Akt activity and cytosolic p27 localization (21, 22). We, and others, have also demonstrated functional roles for PI3 kinase, Akt, and p27 in PTC cell invasion in vitro (16,23,24). However, the correlation between increased Akt activity and invasion was not found for PTCs with activating BRAF mutations. Most importantly, these focused s...

TIMP3 regulates migration, invasion and in vivo tumorigenicity of thyroid tumor cells

“…In the data sets of both He et al (2005) and Jarzab et al (2005), a statistically significant reduced (Po0.01) TIMP3 expression was displayed by PTC when compared with unaffected thyroid tissues from the same patients (Figures 1c and d). Although not statistically significant, in the data set of Vasko et al (2007), both microscopically dissected central and invasive regions of PTC of three out of four patients displayed reduced expression of TIMP3 transcript in comparison with the corresponding normal region (Figure 1e). Overall, this meta-analysis suggests that TIMP3 downregulation was found to be a general event in PTC carcinogenesis.…”

“…As control, the expression of E-cadherin in human primary thyrocytes is shown (Figure 7b). Vimentin is involved in invasion and its expression is upregulated in several tumors, including PTC (Vasko et al, 2007). As shown in Figure 7c (top), the vimentin level in NIM1-TIMP3 clones was reduced with respect to NIM1-EV cells.…”

Molecular profiling related to poor prognosis in thyroid carcinoma. Combining gene expression data and biological information

“…The TGFb signaling pathway has been associated with the epithelial-to-mesenchymal (EMT) transition in later stages of cancer (Siegel and Massague, 2003) and more recently, with papillary thyroid tumor invasion (Vasko et al, 2007). EMT is a multi-step process involving dissolution of tight and adherent junctions, cytoskeletal reorganization, loss of cell polarity, repression of epithelial markers and upregulation of mesenchymal proteins (Grunert et al, 2003).…”

Snail overexpression induces an epithelial to mesenchymal transition and cancer stem cell-like properties in SCC9 cells

“…The whole-cell lysates were collected and their concentrations determined using the Bradford assay. Protein (20 mg) 24 was separated by SDS-PAGE and transferred to PVDF membranes (Millipore, Billerica, MA, USA). The blots were then probed with antibodies, such as anti-b-actin (1:1000, Santa Cruz Biotechnology, Santa Cruz, CA, USA), anti-E-cadherin (1:400, Santa Cruz Biotechnology), anti-b-catenin (1:500, Santa Cruz Biotechnology), antivimentin (1:500, Santa Cruz Biotechnology), and anti-Snail (1:500, Abcam, Cambridge, MA, USA).…”