Exploring the Interplay of Telomerase Reverse Transcriptase and ß-Catenin in Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is one of the deadliest human cancers. Activating mutations in the telomerase reverse transcriptase (TERT) promoter (TERTp) and CTNNB1 gene encoding ß-catenin are widespread in HCC (~50% and ~30%, respectively). TERTp mutations are predicted to increase TERT transcription and telomerase activity. This review focuses on exploring the role of TERT and ß-catenin in HCC and the current findings regarding their interplay. TERT can have contradictory effects on tumorigenesis via both its canonical and non-canonical functions. As a critical regulator of proliferation and differentiation in progenitor and stem cells, activated ß-catenin drives HCC; however, inhibiting endogenous ß-catenin can also have pro-tumor effects. Clinical studies revealed a significant concordance between TERTp and CTNNB1 mutations in HCC. In stem cells, TERT acts as a co-factor in ß-catenin transcriptional complexes driving the expression of WNT/ß-catenin target genes, and ß-catenin can bind to the TERTp to drive its transcription. A few studies have examined potential interactions between TERT and ß-catenin in HCC in vivo, and their results suggest that the coexpression of these two genes promotes hepatocarcinogenesis. Further studies are required with vertebrate models to better understand how TERT and ß-catenin influence hepatocarcinogenesis.

Quantifying Liver Size in Larval Zebrafish Using Brightfield Microscopy.

In several transgenic zebrafish models of hepatocellular carcinoma (HCC), hepatomegaly can be observed during early larval stages. Quantifying larval liver size in zebrafish HCC models provides a means to rapidly assess the effects of drugs and other manipulations on an oncogene-related phenotype. Here we show how to fix zebrafish larvae, dissect the tissues surrounding the liver, photograph livers using bright-field microscopy, measure liver area, and analyze results. This protocol enables rapid, precise quantification of liver size. As this method involves measuring liver area, it may underestimate differences in liver volume, and complementary methodologies are required to differentiate between changes in cell size and changes in cell number. The dissection technique described herein is an excellent tool to visualize the liver, gut, and pancreas in their natural positions for myriad downstream applications including immunofluorescence staining and in situ hybridization. The described strategy for quantifying larval liver size is applicable to many aspects of liver development and regeneration.

Heterogeneous beta-catenin activation is sufficient to cause hepatocellular carcinoma in zebrafish.

Up to 41% of hepatocellular carcinomas (HCCs) result from activating mutations in the CTNNB1 gene encoding ß-catenin. HCC-associated CTNNB1 mutations stabilize the ß-catenin protein, leading to nuclear and/or cytoplasmic localization of ß-catenin and downstream activation of Wnt target genes. In patient HCC samples, ß-catenin nuclear and cytoplasmic localization are typically patchy, even among HCC with highly active CTNNB1 mutations. The functional and clinical relevance of this heterogeneity in ß-catenin activation are not well understood. To define mechanisms of ß-catenin-driven HCC initiation, we generated a Cre-lox system that enabled switching on activated ß-catenin in (1) a small number of hepatocytes in early development; or (2) the majority of hepatocytes in later development or adulthood. We discovered that switching on activated ß-catenin in a subset of larval hepatocytes was sufficient to drive HCC initiation. To determine the role of Wnt/ß-catenin signaling heterogeneity later in hepatocarcinogenesis, we performed RNA-seq analysis of zebrafish ß-catenin-driven HCC. At the single-cell level, 2.9% to 15.2% of hepatocytes from zebrafish ß-catenin-driven HCC expressed two or more of the Wnt target genes axin2, mtor, glula, myca and wif1, indicating focal activation of Wnt signaling in established tumors. Thus, heterogeneous ß-catenin activation drives HCC initiation and persists throughout hepatocarcinogenesis.

Events of Molecular Changes in Epithelial-Mesenchymal Transition.

EMT is the process by which epithelial cells, characterized by well-developed intercellular contacts, transdifferentiate into motile and invasive mesenchymal cells. This process is associated with the loss of transmembrane intercellular adhesion molecule E-cadherin and disruption of cell-cell junctions along with acquisition of migratory properties. EMT is integral in embryonic development, wound healing, and stem cell behavior; however, its aberrant activation by micro-environmental alterations and abnormal stimuli can lead to cancer progression. Here, we review the different molecular changes associated with EMT that are responsible for downregulation of epithelial genes. Increased knowledge of the EMT process is essential for therapeutic targeting of cancer cells.

Epithelial Mesenchymal Transition and Vascular Mimicry in Breast Cancer Stem Cells.

Vasculogenic mimicry (VM), a newly defined pattern of tumor microvascularization differs from angiogenesis and vasculogenesis in its noninvolvement of endothelial cells, by which highly aggressive tumor cells can form vessel-like structures themselves, because of their high plasticity. The presence of VM has been shown to be strongly associated with a poor prognosis in several types of cancer, but biological features of tumor cells that form VM remains unknown. Human breast cancer, characterized by a group of highly heterogeneous lesions, is the most common cancer in women and one of the leading causes of cancer-related deaths worldwide. The epithelialmesenchymal transition (EMT) state in breast cancer has been associated with cancer stem cell (CSC) properties, self-renewal capabilities, resistance to conventional therapies, and a tendency for posttreatment recurrence. With increasing knowledge about cancer stem cell phenotypes and functions, they are implicated in VM formation. Studies also indicate that EMT is relevant to the acquisition and maintenance of stem cell-like characteristics and is involved in VM. This review discusses the correlation between CSCs, EMT, and VM formation with a focus on breast cancer. Also, the signalling molecules and pathways involved in VM and some recently defined direct VM targeting strategies in breast cancer are reviewed here.

Breast cancer stem cells, EMT and therapeutic targets.

A small heterogeneous population of breast cancer cells acts as seeds to induce new tumor growth. These seeds or breast cancer stem cells (BCSCs) exhibit great phenotypical plasticity which allows them to undergo "epithelial to mesenchymal transition" (EMT) at the site of primary tumor and a future reverse transition. Apart from metastasis they are also responsible for maintaining the tumor and conferring it with drug and radiation resistance and a tendency for post-treatment relapse. Many of the signaling pathways involved in induction of EMT are involved in CSC generation and regulation. Here we are briefly reviewing the mechanism of TGF-ß, Wnt, Notch, TNF-α, NF-κB, RTK signalling pathways which are involved in EMT as well as BCSCs maintenance. Therapeutic targeting or inhibition of the key/accessory players of these pathways could control growth of BCSCs and hence malignant cancer. Additionally several miRNAs are dysregulated in cancer stem cells indicating their roles as oncogenes or tumor suppressors. This review also lists the miRNA interactions identified in BCSCs and discusses on some newly identified targets in the BCSC regulatory pathways like SHIP2, nicastrin, Pin 1, IGF-1R, pro-inflammatory cytokines and syndecan which can be targeted for therapeutic achievements.