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// Esther Ekpe-Adewuyi 1 , Ana Lopez-Campistrous 1 , Xiaoyun Tang 2 , David N. Brindley 2 , Todd P. W. McMullen 1, 3 1 Department of Surgery, University of Alberta, Edmonton, Canada 2 Signal Transduction Research Group, Department of Biochemistry, University of Alberta, Edmonton, Canada 3 Division of Surgical Oncology, Cross Cancer Institute, University of Alberta, Edmonton, Canada Correspondence to: Todd P. W. McMullen, email: todd.mcmullen@albertahealthservices.ca Keywords: papillary thyroid cancer (PTC), platelet derived growth factor receptor-alpha (PDGFRα), epithelial to mesenchymal transition (EMT), invadopodia, crenolanib Received: June 30, 2016 Accepted: October 22, 2016 Published: November 11, 2016 ABSTRACT Recently platelet derived growth factor receptor-alpha (PDGFRα) was recognized as a potential target to treat aggressive papillary thyroid cancer given its strong association with lymph node metastases. However, it is unclear how PDGFRα potentiates metastases and if it works through the canonical MAPK pathway traditionally linked to PTC oncogenesis. We explored the phenotypic changes driven by PDGFRα activation in human papillary thyroid cancer (PTC) cells and the downstream signalling cascades through which they are effected. We demonstrate that PDGFRα drives an impressive phenotypic change in PTC cell lines as documented by significant cytoskeletal rearrangement, increased migratory potential, and the formation of invadopodia. Cells lacking PDGFRα formed compact and dense spheroids, whereas cells expressing active PDGFRα exhibited invadopodia in three-dimensional culture. To achieve this, active PDGFRα provoked downstream activation of the MAPK/Erk, PI3K/Akt and STAT3 pathways. We further confirmed the role of PDGFRα as a transformative agent promoting the epithelial to mesenchymal transition of PTC cells, through the augmentation of Snail and Slug expression. Crenolanib, a small molecule inhibitor of PDGFRα, suppressed the levels of Snail and Slug and almost completely reversed all the phenotypic changes. We demonstrate that PDGFRα activation is an essential component that drives aggressiveness in PTC cells, and that the signaling pathways are complex, involving not only the MAPK/Erk but also the PI3K/Akt and STAT3 pathways. This argues for upstream targeting of the PDGFRα given the redundancy of oncogenic pathways in PTC, especially in patients whose tumors over-express this tyrosine kinase receptor.

Epithelial to mesenchymal transition (EMT) is a developmental process which is aberrantly activated during cancer invasion and metastasis. Elevated expression of EMT-inducers like ZEB1 enables tumor cells to detach from the primary tumor and invade into the surrounding tissue. The main antagonist of ZEB1 in controlling EMT is the microRNA-200 family that is reciprocally linked to ZEB1 in a double negative feedback loop. Here, we further elucidate how the ZEB1/miR-200 feedback loop controls invasion of tumor cells. The process of EMT is attended by major changes in the actin cytoskeleton. Via in silico screening of genes encoding for actin interacting proteins, we identified two novel targets of miR-200c - TKS5 and MYLK (MLCK). Co-expression of both genes with ZEB1 was observed in several cancer cell lines as well as in breast cancer patients and correlated with low miR-200c levels. Depletion of TKS5 or MYLK in breast cancer cells reduced their invasive potential and their ability to form invadopodia. Whereas TKS5 is known to be a major component, we could identify MYLK as a novel player in invadopodia formation. In summary, TKS5 and MYLK represent two mediators of invasive behavior of cancer cells that are regulated by the ZEB1/miR-200 feedback loop.

// Krystal D. Nolan 1, * , Jasmine Kaur 1, * , Jennifer S. Isaacs 1 1 Department of Cell and Molecular Pharmacology, Medical University of South Carolina, Hollings Cancer Center, Charleston, SC, USA * These authors have contributed equally to this work Correspondence to: Jennifer S. Isaacs, email: isaacsj@musc.edu Keywords: Hsp90, cancer stem cells, prostaspheres, epithelial to mesenchymal transition (EMT) Received: October 20, 2016 Accepted: December 05, 2016 Published: December 27, 2016 ABSTRACT Heat-shock protein 90 (Hsp90), a highly conserved molecular chaperone, is frequently upregulated in tumors, and remains an attractive anti-cancer target. Hsp90 is also found extracellularly, particularly in tumor models. Although extracellular Hsp90 (eHsp90) action is not well defined, eHsp90 targeting attenuates tumor invasion and metastasis, supporting its unique role in tumor progression. We herein investigated the potential role of eHsp90 as a modulator of cancer stem-like cells (CSCs) in prostate cancer (PCa). We report a novel function for eHsp90 as a facilitator of PCa stemness, determined by its ability to upregulate stem-like markers, promote self-renewal, and enhance prostasphere growth. Moreover, eHsp90 increased the side population typically correlated with the drug-resistant phenotype. Intriguingly, tumor cells with elevated surface eHsp90 exhibited a marked increase in stem-like markers coincident with increased expression of the epithelial to mesenchymal (EMT) effector Snail, indicating that surface eHsp90 may enrich for a unique CSC population. Our analysis of distinct effectors modulating the eHsp90-dependent CSC phenotyperevealed that eHsp90 is a likely facilitator of stem cell heterogeneity. Taken together, our findings provide unique functional insights into eHsp90 as a modulator of PCa plasticity, and provide a framework towards understanding its role as a driver of tumor progression.

The dynamic nature of cancer stem cells that underlie metastasis or their ability to switch between different cellular identities, as in EMT and MET, has profound implications for cancer therapy. The functional relationship between molecules involved in cancer cell stemness and metastasis is not clear. In this regard, our studies on hGBM tissue grade IV specimens showed significant expression of Twist1 and Sox2, known mesenchymal and stemness related markers, respectively, indicating their association with glial tumor genesis and metastasis. The glioma stem cells obtained from CD133+ cells demonstrated increased expression of Twist1 and Sox2 accompanied by significant increase in the mesenchymal markers such as N-cadherin, vimentin and β-catenin. Our studies on glioma stem cells treatment with human umbilical cord blood derived- mesenchymal stem cells, showed down regulation of Twist1 and Sox2 proteins, apart from other mesenchymal stem cell markers. Based on the in vitro experiments and in vivo intracranial xenograft mouse model studies, we elucidated the potential therapeutic role of hUCBSC in suppressing glioma cancer stemness by the induction of MET.