TWIST1, a transcription factor, plays a pivotal role in cancer initiation and progression. Anaplastic thyroid carcinoma (ATC) is one of the deadliest human malignancies; TWIST1 is overexpressed in ATC and increases thyroid cancer cell survival, migration and invasion. The molecular mechanisms underlying the effects of TWIST1 are partially known. Here, using miRNome profiling of papillary thyroid cancer cells (TPC-1) ectopically expressing TWIST1, we identified miR-584. We showed that TWIST1 directly binds miR-584 using chromatin immunoprecipitation. Importantly, miR-584 was up-regulated in human ATC compared to papillary thyroid carcinoma (PTC) and normal thyroid samples. Overexpression of miR-584 in TPC cells induced resistance to apoptosis, whereas stable transfection of anti-miR-584 in TPC-TWIST1 and 8505C cells increased the sensitivity to apoptosis. Using bioinformatics programs, we identified TUSC2 (tumor suppressor candidate 2) as a novel target of miR-584. TUSC2 mRNA and protein levels were decreased in TPC miR-584 and increased in TPC-TWIST1 anti-miR-584 cells. Luciferase assays demonstrated direct targeting. Restored expression of TUSC2 rescued the inhibition of apoptosis induced by miR-584. Finally, qRT-PCR and immunohistochemical analysis showed that TUSC2 was down-regulated in ATC and PTC samples compared to normal thyroids. In conclusion, our study identified a novel TWIST1/miR-584/TUSC2 pathway that plays a role in resistance to apoptosis of thyroid cancer cells.

We identified a set of genes that mediates Twist1 biological effects in thyroid cancer cells.

Purpose Aberrant expression of miRNAs is crucial in several tissues tumorigenesis including thyroid. Recent studies demonstrated that miR-650 plays different role depending on the cancer type. Herein, we investigated the role of miR-650 in thyroid carcinoma. Methods The expression of miR-650 was analyzed in human thyroid tissues by q-RT-PCR. Anaplastic (8505C, CAL62, SW1736) and papillary (TPC-1) thyroid cancer cell lines were used to dissect the role of miR-650 on malignant hallmarks of transformation. Label-free proteomic analysis was exploited to unravel the targets of miR-650, while luciferase reporter assay and functional experiments were performed to confirm a selected target. Spearman's rank correlation test was used to assess the association between miR-650 and its target in human thyroid cancer tissues. Results miR-650 is over-expressed in anaplastic (ATC) thyroid carcinoma where it enhances cell migration and invasion. Proteomic label-free and bioinformatics analysis revealed that the serine-threonine protein phosphatase 2 catalytic subunit alpha (PPP2CA) is a target of miR-650; these finding were confirmed by luciferase assay. Restoration of PPP2CA mRNA, deprived of its 3′UTR, is able to revert the malignant phenotype induced by miR-650 in HEK-293 cells. Importantly, PPP2CA is down-regulated in ATC tissues and is inversely correlated with miR-650. Conclusions miR-650 displayed oncogenic activity in ATC cells through targeting PPP2CA phosphatase. These results suggest that miR-650/PPP2CA axis could be modulated to interfere with motile ability of thyroid carcinoma cells.

Background Aberrant expression of microRNAs (miR) has been proposed as non-invasive biomarkers for breast cancers. The aim of this study was to analyse the miR-622 level in the plasma and in tissues of breast cancer patients and to explore the role of miR-622 and its target, the NUAK1 kinase, in this context. Methods miR-622 expression was analysed in plasma and in tissues samples of breast cancer patients by q-RT-PCR. Bioinformatics programs, luciferase assay, public dataset analysis and functional experiments were used to uncover the role of miR-622 and its target in breast cancer cells. Results miR-622 is downregulated in plasma and in tissues of breast cancer patients respect to healthy controls and its downregulation is significantly associated with advanced grade and high Ki67 level. Modulation of miR-622 affects the motility phenotype of breast cancer cells. NUAK1 kinase is a functional target of miR-622, it is associated with poor clinical outcomes of breast cancer patients and is inversely correlated with miR-622 level. Conclusions miR-622/NUAK1 axis is deregulated in breast cancer patients and affects the motility phenotype of breast cancer cells. Importantly, miR-622 and NUAK1 hold promises as biomarkers and as targets for breast cancers.

BackgroundThrough a transcriptome microarray analysis, we have isolated Anterior gradient protein 2 (AGR2) as a gene up-regulated in papillary thyroid carcinoma (PTC). AGR2 is a disulfide isomerase over-expressed in several human carcinomas and recently linked to endoplasmic reticulum (ER) stress. Here, we analyzed the expression of AGR2 in PTC and its functional role.MethodsExpression of AGR2 was studied by immunohistochemistry and real time PCR in normal thyroids and in PTC samples. The function of AGR2 was studied by knockdown in PTC cells and by ectopic expression in non-transformed thyroid cells. The role of AGR2 in the ER stress was analyzed upon treatment of cells, expressing or not AGR2, with Bortezomib and analyzing by Western blot the expression levels of GADD153.ResultsPTC over-expressed AGR2 at mRNA and protein levels. Knockdown of AGR2 in PTC cells induced apoptosis and decreased migration and invasion. Ectopic expression of AGR2 in non-transformed human thyroid cells increased migration and invasion and protected cells from ER stress induced by Bortezomib.ConclusionsAGR2 is a novel marker of PTC and plays a role in thyroid cancer cell survival, migration, invasion and protection from ER stress.

Thyroid carcinoma is the most common endocrine cancer and includes different forms. Among these, anaplastic thyroid carcinoma (ATC) is the rarest but the most lethal subtype, compared to papillary thyroid carcinoma (PTC) which shows an overall good prognosis. We have previously showed that Tumor Suppressor Candidate 2 (TUSC2), a known tumour suppressor gene, is downregulated in human PTC and ATC compared to normal thyroid samples. The aim of this study was to gain insight into the molecular mechanisms induced by TUSC2 in thyroid cancer cells. Here, we stably transfected TUSC2 in papillary (TPC-1) and in anaplastic (8505C) thyroid cancer cell lines and studied its effects on several biological processes, demonstrating that TUSC2 overexpression decreased thyroid cancer cell proliferation, migration and invasion. Through the proteome profiler apoptosis array, we observed that TUSC2 increased sensitivity to apoptosis by increasing the SMAC/DIABLO and CYTOCHROME C proteins. On the other hand, transient silencing of TUSC2, by siRNA, in an immortalized thyroid follicular epithelial cell line (Nthy-ori 3-1) showed the opposite effect. Finally modulation of SMAC/DIABLO partially rescued the biological effects of TUSC2. Thus, our data highlight a tumour suppressor role of TUSC2 in thyroid carcinogenesis, suggesting that it could be a promising target and biomarker for thyroid carcinoma.

Junctional adhesion molecule A (JAM‐A) is a transmembrane protein that contributes to different biological process, including the epithelial to mesenchymal transition (EMT). Through an EMT profiler array, we explored the molecular players associated with human thyroid cancer progression and identified JAM‐A as one of the genes mostly deregulated. The quantitative real‐time polymerase chain reaction and immunohistochemistry analyses showed that downregulation of JAM‐A occurred in anaplastic thyroid carcinoma (ATC) compared with normal thyroid (NT) and papillary thyroid carcinoma (PTC) tissues and correlated with extrathyroid infiltration, tumor size, and ATC histotype. In ATC cell lines, JAM‐A restoration suppressed malignant hallmarks of transformation including cell proliferation, motility, and transendothelial migration. Accordingly, knockdown of JAM‐A enhanced thyroid cancer cell proliferation and motility in PTC cells. Through the proteome profiler human phospho‐kinase array, we demonstrated that higher expression of JAM‐A was associated with a significant increased level of phosphorylation of p53 and GSK3 α/β proteins. In conclusion, our findings highlight a novel role of JAM‐A in thyroid cancer progression and suggest that JAM‐A restoration could have potential clinical relevance in thyroid cancer treatment.

This study demonstrated the technical feasibility of an HFUS-guided orthotopic mouse model of FTC. The HFUS-guided orthotopic model is easily reproducible and allows prolonged monitoring of the disease because the animals showed an increased survival rate.