Identification of Circulating Tumor Cell Phenotype in Differentiated Thyroid Carcinoma

Objective: Circulating tumor cells (CTCs) have been considered as the origin of tumor metastasis and recurrence, which always indicate a poor prognosis. There are three phenotypes of CTCs according on different epithelial-to-mesenchymal transition (EMT) markers, including epithelial, mesenchymal, and epithelial/mesenchymal (mixed phenotypic) CTCs. We intended to explore the relationship among CTC phenotypes and the clinicopathological characteristics of patients with differentiated thyroid carcinoma (DTC). Methods: Peripheral blood samples from 58 patients with DTC were collected, and CTCs were isolated by cell sizes. To identify phenotypes of CTCs, branched DNA signal amplification technology was adopted to capture and amplify target sequences, and then multiplex RNA-in situ hybridization (RNA-ISH) assay was used to identify CTC phenotypes depended on epithelial-mesenchymal transition (EMT) markers. Results: The positive rate of CTCs was 77.59% in 58 DTC patients. Totally, 488 CTCs with detective phenotype were found. Among them, there were 121 (24.80%) epithelial CTCs, 67 (13.72%) mesenchymal CTCs, and 300 (61.48%) mixed phenotypic CTCs. An obvious increased epithelial CTCs was observed in male patients compared with female. Notably, CTCs were more prevailing in younger male patients with ETI and bilateral focus. Conclusions: The CTCs are common in DTC patients, and mixed phenotypic is the major phenotype, indicating that EMT is prevalent in DTC even though its prognosis was better than other epithelial tumors. Detection of CTC and its phenotypes might independently predict the prognosis of DTC.

MicroRNA-517c Functions as a Tumor Suppressor in Hepatocellular Carcinoma via Downregulation of KPNA2 and Inhibition of PI3K/AKT Pathway

Objective. Hepatocellular carcinoma (HCC) is a kind of solid and highly aggressive malignant tumor with poor prognosis. MicroRNA (miRNA/miR) has been confirmed to be involved in HCC development. The current study focused on the functions and mechanisms of miR-517c in HCC. Methods. Expressions of miR-517c and Karyopherin α2 (KPNA2) mRNA in HCC cell lines and tissue samples were examined using quantitative real-time polymerase chain reaction (qRT-PCR). Western blot was conducted for detections of epithelial-to-mesenchymal transition (EMT) and PI3K/AKT markers. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) and Transwell assays were utilized to investigate the influence of miR-517c on HCC cell proliferation, invasion, and migration. TargetScan and luciferase reporter assay were performed to search for the potential target gene of miR-517c. Results. We demonstrated that miR-517c expressions were decreased in HCC tissues and cells. Moreover, the clinical analysis showed that decreased miR-517c expressions in HCC tissues correlated with shorter overall survival and malignant clinicopathologic features of HCC patients. MTT assay showed that miR-517c upregulation prominently repressed HCC cell proliferation. In addition, miR-517c restoration could significantly suppress HCC cell invasion and migration as demonstrated by Transwell assays. We also found that miR-517c directly targeted KPNA2 and regulated the PI3K/AKT pathway and EMT, exerting prohibitory functions in HCC. Conclusion. Taken together, this study stated that miR-517c inhibited HCC progression via regulating the PI3K/AKT pathway and EMT and targeting KPNA2 in HCC, providing a novel insight into HCC treatment.

Oncogenic Role of Exosomal Circular and Long Noncoding RNAs in Gastrointestinal Cancers

Circular RNAs (circRNAs) and long noncoding RNAs (lncRNAs) are differentially expressed in gastrointestinal cancers. These noncoding RNAs (ncRNAs) regulate a variety of cellular activities by physically interacting with microRNAs and proteins and altering their activity. It has also been suggested that exosomes encapsulate circRNAs and lncRNAs in cancer cells. Exosomes are then discharged into the extracellular environment, where they are taken up by other cells. As a result, exosomal ncRNA cargo is critical for cell–cell communication within the cancer microenvironment. Exosomal ncRNAs can regulate a range of events, such as angiogenesis, metastasis, immune evasion, drug resistance, and epithelial-to-mesenchymal transition. To set the groundwork for developing novel therapeutic strategies against gastrointestinal malignancies, a thorough understanding of circRNAs and lncRNAs is required. In this review, we discuss the function and intrinsic features of oncogenic circRNAs and lncRNAs that are enriched within exosomes.

Dual role of Ovol2 on the germ cell lineage segregation during gastrulation in mouse embryogenesis

In mammals, primordial germ cells (PGCs), the origin of the germ line, are specified from the epiblast at the posterior region where gastrulation simultaneously occurs, yet the functional relationship between PGC specification and gastrulation remains unclear. Here, we show that Ovol2, a transcription factor conserved across the animal kingdom, balances these major developmental processes by repressing the epithelial-to-mesenchymal transition (EMT) driving gastrulation and the upregulation of genes associated with PGC specification. Ovol2a, a splice variant encoding a repressor domain, directly regulates EMT-related genes and consequently induces re-acquisition of potential pluripotency during PGC specification, whereas Ovol2b, another splice variant missing the repressor domain, directly upregulates genes associated with PGC specification. Taken together, these results elucidate the molecular mechanism underlying allocation of the germ line among epiblast cells differentiating into somatic cells through gastrulation.

A comprehensive framework for analysis of microRNA sequencing data in metastatic colorectal cancer

ABSTRACT Although microRNAs (miRNAs) contribute to all hallmarks of cancer, miRNA dysregulation in metastasis remains poorly understood. The aim of this work was to reliably identify miRNAs associated with metastatic progression of colorectal cancer (CRC) using novel and previously published next-generation sequencing (NGS) datasets generated from 268 samples of primary (pCRC) and metastatic CRC (mCRC; liver, lung and peritoneal metastases) and tumor adjacent tissues. Differential expression analysis was performed using a meticulous bioinformatics pipeline, including only bona fide miRNAs, and utilizing miRNA-tailored quality control and processing. Five miRNAs were identified as up-regulated at multiple metastatic sites Mir-210_3p, Mir-191_5p, Mir-8-P1b_3p [mir-141–3p], Mir-1307_5p and Mir-155_5p. Several have previously been implicated in metastasis through involvement in epithelial-to-mesenchymal transition and hypoxia, while other identified miRNAs represent novel findings. The use of a publicly available pipeline facilitates reproducibility and allows new datasets to be added as they become available. The set of miRNAs identified here provides a reliable starting-point for further research into the role of miRNAs in metastatic progression.

MicroRNA-92a-3p Enhances Cisplatin Resistance by Regulating Krüppel-Like Factor 4-Mediated Cell Apoptosis and Epithelial-to-Mesenchymal Transition in Cervical Cancer

Recent studies have confirmed the existence and key roles of microRNA (miRNAs) in cancer drug resistance, including cervical cancer (CC). The present study aims to establish a novel role for miR-92a-3p and its associated gene networks in cisplatin (DDP) resistance of CC. First, the disparities in miRNA expression between CC tissues and adjacent normal tissues were screened based on GSE19611 microarray data that retrieved from Gene Expression Omnibus (GEO), and we identified several miRs that were significantly downregulated or upregulated in CC tissues including miR-92a-3p. Moreover, miR-92a-3p was significantly up-regulated in DDP-resistant cells and was the most differently expressed miRNA. Functionally, knockdown of miR-92a-3p increased the sensitivity of DDP-resistant cells to DDP via inhibiting cell proliferation, migration and invasion, and promoting apoptosis. Conversely, overexpression of miR-92a-3p significantly induced DDP resistance in CC parental cells including HeLa and SiHa cells. Moreover, Krüppel-like factor 4 (KLF4) was identified as a direct target of miR-92a-3p, and an obvious inverse correlation was observed between the expression of miR-92a-3p and KLF4 in 40 pairs of cancer tissues. Furthermore, KLF4 knockdown reversed the promoting effect of miR-92a-3p inhibition on DDP sensitivity in DDP-resistant CC cells. Besides, high expression of miR-92a-3p was associated with DDP resistance, as well as a short overall survival in clinic. Taken together, these findings provide important evidence that miR-92a-3p targets KLF4 and is significant in DDP resistance in CC, indicating that miR-92a-3p may be an attractive target to increase DDP sensitivity in clinical CC treatment.

Expression of Fibroblast Activation Protein Is Enriched in Neuroendocrine Prostate Cancer and Predicts Worse Survival

Background: Advanced prostate cancer (PC) may accumulate genomic alterations that hallmark lineage plasticity and transdifferentiation to a neuroendocrine (NE) phenotype. Fibroblast activation protein (FAP) is a key player in epithelial-to-mesenchymal transition (EMT). However, its clinical value and role in NE differentiation in advanced PC has not been fully investigated. Methods: Two hundred and eight patients from a multicenter, prospective cohort of patients with metastatic castration-resistant prostate cancer (CRPC) with available RNA sequencing data were analyzed for tumor FAP mRNA expression, and its association with overall survival (OS) and NE tumor features was investigated. Results: Twenty-one patients (10%) were found to have high FAP mRNA expression. Compared to the rest, this subset had a proportionally higher exposure to taxanes and AR signaling inhibitors (abiraterone or enzalutamide) and was characterized by active NE signaling, evidenced by high NEPC- and low AR-gene expression scores. These patients with high tumor mRNA FAP expression had a more aggressive clinical course and significantly shorter survival (12 months) compared to those without altered FAP expression (28 months, log-rank p = 0.016). Conclusions: FAP expression may serve as a valuable NE marker indicating a worse prognosis in patients with metastatic CRPC.

MiR-21 in Lung Transplant Recipients With Chronic Lung Allograft Dysfunction

Background: Micro-RNA-21 (miR-21) is a post-translational regulator involved in epithelial-to-mesenchymal transition (EMT). Since EMT is thought to contribute to chronic lung allograft dysfunction (CLAD), we aimed to characterize miR-21 expression and distinct EMT markers in CLAD.Methods: Expression of miR-21, vimentin, Notch intracellular domain (NICD) and SMAD 2/3 was investigated in explanted CLAD lungs of patients who underwent retransplantation. Circulating miR-21 was determined in collected serum samples of CLAD and matched stable recipients.Results: The frequency of miR-21 expression was higher in restrictive allograft syndrome (RAS) than in bronchiolitis obliterans syndrome (BOS) specimens (86 vs 30%, p = 0.01); Vimentin, NICD and p-SMAD 2/3 were positive in 17 (100%), 12 (71%), and 7 (42%) BOS patients and in 7 (100%), 4 (57%) and 4 (57%) RAS cases, respectively. All four markers were negative in control tissue from donor lungs. RAS patients showed a significant increase in serum concentration of miR-21 over time as compared to stable recipients (p = 0.040).Conclusion: To the best of our knowledge this is the first study highlighting the role miR-21 in CLAD. Further studies are necessary to investigate the involvement of miR-21 in the pathogenesis of CLAD and its potential as a therapeutic target.

Wnt5A and TGFβ1 Converges through YAP1 Activity and Integrin Alpha v Up-Regulation Promoting Epithelial to Mesenchymal Transition in Ovarian Cancer Cells and Mesothelial Cell Activation

In this paper, we investigate whether Wnt5A is associated with the TGF-β1/Smad2/3 and Hippo-YAP1/TAZ-TEAD pathways, implicated in epithelial to mesenchymal transition (EMT) in epithelial ovarian cancer. We used 3D and 2D cultures of human epithelial ovarian cancer cell lines SKOV-3, OVCAR-3, CAOV-4, and different subtypes of human serous ovarian cancer compared to normal ovary specimens. Wnt5A showed a positive correlation with TAZ and TGFβ1 in high- and low-grade serous ovarian cancer specimens compared to borderline serous and normal ovaries. Silencing Wnt5A by siRNAs significantly decreased Smad2/3 activation and YAP1 expression and nuclear shuttling in ovarian cancer (OvCa) cells. Furthermore, Wnt5A was required for TGFβ1-induced cell migration and invasion. In addition, inhibition of YAP1 transcriptional activity by Verteporfin (VP) altered OvCa cell migration and invasion through decreased Wnt5A expression and inhibition of Smad2/3 activation, which was reverted in the presence of exogenous Wnt5A. We found that the activation of TGFβ1 and YAP1 nuclear shuttling was promoted by Wnt5A-induced integrin alpha v. Lastly, Wnt5A was implicated in activating human primary omental mesothelial cells and subsequent invasion of ovarian cancer cells. Together, we propose that Wnt5A could be a critical mediator of EMT-associated pathways.

Analysis of the glyco-code in pancreatic ductal adenocarcinoma identifies glycan-mediated immune regulatory circuits

Pancreatic ductal adenocarcinoma (PDAC) remains one of the most aggressive malignancies with a 5-year survival rate of only 9%. Despite the fact that changes in glycosylation patterns during tumour progression have been reported, no systematic approach has been conducted to evaluate its potential for patient stratification. By analysing publicly available transcriptomic data of patient samples and cell lines, we identified here two specific glycan profiles in PDAC that correlated with progression, clinical outcome and epithelial to mesenchymal transition (EMT) status. These different glycan profiles, confirmed by glycomics, can be distinguished by the expression of O-glycan fucosylated structures, present only in epithelial cells and regulated by the expression of GALNT3. Moreover, these fucosylated glycans can serve as ligands for DC-SIGN positive tumour-associated macrophages, modulating their activation and inducing the production of IL-10. Our results show mechanisms by which the glyco-code contributes to the tolerogenic microenvironment in PDAC.