EphB3 receptor suppressor invasion, migration and proliferation in glioma by inhibiting EGFR-PI3K/AKT signaling pathway.

Eph receptors are the largest subfamily of receptor tyrosine kinases, and they have been shown to play a crucial role in glioma. The EphB3 receptor is a member of this family, and its effect on the invasion, migration and proliferation of glioma cells was examined in this study. It was found that the expression of EphB3 was decreased in glioma specimens with increasing tumor grade. Additionally, the U87MG and U251 cell lines showed low levels of EphB3 expression. This finding was consistent with the negative correlation between EphB3 expression in glioma tissues and tumor grade. Depletion of EphB3 gene in U87MG and U251 cell lines resulted in a substantial enhancement of their invasion, migration, and proliferation capacities in vitro. Furthermore, the knockdown of EphB3 led to an upregulation of EGFR, p-PI3K, and p-AKT protein levels. On the other hand, EphB3 overexpression reduced the invasiveness, proliferative capacity and migration rate of U87MG and U251 cells, and downregulated EGFR, p-PI3K and p-AKT. These findings indicate that EphB3 functions as a tumor suppressor in glioma, and its downregulation enhances the malignant potential of glioma cells by activating the EGFR-PI3K/AKT pathway. Thus, EphB3 is a promising diagnostic marker for glioma, and the EphB3-EGFR-PI3K / AKT axis deserves further investigation as a potential therapeutic target.

Super interactive promoters provide insight into cell type-specific regulatory networks in blood lineage cell types.

Existing studies of chromatin conformation have primarily focused on potential enhancers interacting with gene promoters. By contrast, the interactivity of promoters per se, while equally critical to understanding transcriptional control, has been largely unexplored, particularly in a cell type-specific manner for blood lineage cell types. In this study, we leverage promoter capture Hi-C data across a compendium of blood lineage cell types to identify and characterize cell type-specific super-interactive promoters (SIPs). Notably, promoter-interacting regions (PIRs) of SIPs are more likely to overlap with cell type-specific ATAC-seq peaks and GWAS variants for relevant blood cell traits than PIRs of non-SIPs. Moreover, PIRs of cell-type-specific SIPs show enriched heritability of relevant blood cell trait (s), and are more enriched with GWAS variants associated with blood cell traits compared to PIRs of non-SIPs. Further, SIP genes tend to express at a higher level in the corresponding cell type. Importantly, SIP subnetworks incorporating cell-type-specific SIPs and ATAC-seq peaks help interpret GWAS variants. Examples include GWAS variants associated with platelet count near the megakaryocyte SIP gene EPHB3 and variants associated lymphocyte count near the native CD4 T-Cell SIP gene ETS1. Interestingly, around 25.7% ~ 39.6% blood cell traits GWAS variants residing in SIP PIR regions disrupt transcription factor binding motifs. Importantly, our analysis shows the potential of using promoter-centric analyses of chromatin spatial organization data to identify biologically important genes and their regulatory regions.

Investigating the effects of compound paralogous EPHB receptor mutations on mouse facial development.

BACKGROUND: Variation in facial shape may arise from the combinatorial or overlapping actions of paralogous genes. Given its many members, and overlapping expression and functions, the EPH receptor family is a compelling candidate source of craniofacial morphological variation. We performed a detailed morphometric analysis of an allelic series of E14.5 Ephb1-3 receptor mutants to determine the effect of each paralogous receptor gene on craniofacial morphology. RESULTS: We found that Ephb1, Ephb2, and Ephb3 genotypes significantly influenced facial shape, but Ephb1 effects were weaker than Ephb2 and Ephb3 effects. Ephb2-/- and Ephb3-/- mutations affected similar aspects of facial morphology, but Ephb3-/- mutants had additional facial shape effects. Craniofacial differences across the allelic series were largely consistent with predicted additive genetic effects. However, we identified a potentially important nonadditive effect where Ephb1 mutants displayed different morphologies depending on the combination of other Ephb paralogs present, where Ephb1+/- , Ephb1-/- , and Ephb1-/- ; Ephb3-/- mutants exhibited a consistent deviation from their predicted facial shapes. CONCLUSIONS: This study provides a detailed assessment of the effects of Ephb receptor gene paralogs on E14.5 mouse facial morphology and demonstrates how the loss of specific receptors contributes to facial dysmorphology.

Pygo1 regulates pathological cardiac hypertrophy via a ß-catenin-dependent mechanism.

Wnt/ß-catenin signaling plays a key role in pathological cardiac remodeling in adults. The identification of a tissue-specific Wnt/ß-catenin interaction factor may provide a tissue-specific clinical targeting strategy. Drosophila Pygo encodes the core interaction factor of Wnt/ß-catenin. Two Pygo homologs (Pygo1 and Pygo2) have been identified in mammals. Different from the ubiquitous expression profile of Pygo2, Pygo1 is enriched in cardiac tissue. However, the role of Pygo1 in mammalian cardiac disease is yet to be elucidated. In this study, we found that Pygo1 was upregulated in human cardiac tissues with pathological hypertrophy. Cardiac-specific overexpression of Pygo1 in mice spontaneously led to cardiac hypertrophy accompanied by declined cardiac function, increased heart weight/body weight and heart weight/tibial length ratios, and increased cell size. The canonical ß-catenin/T-cell transcription factor 4 (TCF4) complex was abundant in Pygo1-overexpressing transgenic (Pygo1-TG) cardiac tissue, and the downstream genes of Wnt signaling, that is, Axin2, Ephb3, and c-Myc, were upregulated. A tail vein injection of ß-catenin inhibitor effectively rescued the phenotype of cardiac failure and pathological myocardial remodeling in Pygo1-TG mice. Furthermore, in vivo downregulated pygo1 during cardiac hypertrophic condition antagonized agonist-induced cardiac hypertrophy. Therefore, our study is the first to present in vivo evidence demonstrating that Pygo1 regulates pathological cardiac hypertrophy in a canonical Wnt/ß-catenin-dependent manner, which may provide new clues for tissue-specific clinical treatment via targeting this pathway.NEW & NOTEWORTHY In this study, we found that Pygo1 is associated with human pathological hypertrophy. Cardiac-specific overexpression of Pygo1 in mice spontaneously led to cardiac hypertrophy. Meanwhile, cardiac function was improved when expression of Pygo1 was interfered in hypertrophy-model mice. Our study is the first to present in vivo evidence demonstrating that Pygo1 regulates pathological cardiac hypertrophy in a canonical Wnt/ß-catenin-dependent manner, which may provide new clues for a tissue-specific clinical treatment targeting this pathway.

Thymus aging in mice deficient in either EphB2 or EphB3, two master regulators of thymic epithelium development.

BACKGROUND: The epithelial microenvironment is involved in thymus aging, but the possible role of EphB receptors that govern the thymic epithelium development has not been investigated. Herein, we study the changes undergone by the thymus of EphB-deficient mice throughout their life. RESULTS: Immune alterations occurring throughout life were more severe in mutant than in wild-type (WT) mice. Mutant thymuses exhibit lower cellularity than WT ones, as well as lower proportions of early thymic progenitors cells and double-positive (CD4+ CD8+ ) thymocytes, but higher of double-negative (CD4- CD8- ) and single-positive (CD4+ CD8- , CD4- CD8+ ) cells. Throughout life, CD4+ naïve cells decreased particularly in mutant mice. In correlation, memory T cells, largely CD8+ cells, increased. Aged thymic epithelium undergoes changes including appearance of big epithelial free areas, decrease of K8+ K5- areas, which, however, contain higher proportions of Ly51+ UEA1- cortical epithelial cells, in correlation with reduced Aire+ medullary epithelial cells. Also, aged thymuses particularly those derived from mutant mice exhibited increased collagen IV, fat-storing cells, and connective cells. CONCLUSIONS: The absence of EphB accelerates the alterations undergone throughout life by both thymic epithelium and thymocytes, and the proportions of peripheral naïve and memory T cells, all of which are hallmarks of immune aging.

Expression Profile and Prognostic Significance of EPHB3 in Colorectal Cancer.

The protein tyrosine kinase Ephrin type-B receptor 3 (EPHB3) is expressed in cells at the base of intestinal crypts, acting as a cellular guide in the maintenance of intestinal crypt architecture. We aimed to investigate the expression profile of EPHB3 in colorectal precancerous lesions and colorectal cancers (CRCs), and assess its prognostic value. EPHB3 expression was higher in CRCs than in normal mucosa and was associated with the intestinal stem cell markers EPHB2, OLFM4, LRIG1, and a proposed cancer stem cell marker, CD44. Enhanced EPHB3 expression significantly declined during the transformation from adenoma to carcinoma and as the tumor invaded into deeper tissue layers. Namely, a substantial reduction of EPHB3 expression was observed in the budding cancer cells at the invasive tumor fronts, which was more extensive than E-cadherin downregulation. In an azoxymethane/dextran sulfate sodium-induced, colitis-associated, CRC model, EPHB3 expression increased along with tumor development. In a large cohort of CRC patients, EPHB3 positivity was observed in 24% of 610 CRCs and was negatively correlated with tumor differentiation, lympho-vascular invasion, and tumor, node, and metastasis stages. EPHB3 was positively associated with microsatellite instability but was associated with neither CpG island methylation, nor with KRAS and BRAF mutations. Notably, EPHB3 positivity was associated with better clinical outcomes, although it was not an independent prognostic marker. Overexpression of EPHB3 in the colon cancer cell line, DLD1, led to decreased cell growth and migration and reduced mitogen-activated protein kinase signaling. Taken together, our data demonstrate the suppressive role of EPHB3 in CRC progression.

Gradual differentiation and confinement of the cardiac conduction system as indicated by marker gene expression.

The components of the cardiac conduction system, responsible for coordinated activation of the heart chambers, are well defined and their cells differ in gene expression profile and phenotype from those of the surrounding working myocardium. Yet, when and on what basis the myocardium of each of the conduction system components become distinguishable from other myocardium during heart development has not been well established. To identify and assess cell type-specific expression profiles and differentiation markers, we performed transcriptome analysis on fluorescence activated cell sorted purified conduction system (Venus+) and chamber myocardial cells (Katushka+) of Tbx3+/Venus;TgNppb(Katushka) double transgenic mouse fetuses. We found that transcripts associated with nervous system development and ion channel activity were enriched in Tbx3+ conduction system cells, whereas transcripts associated with mitochondrial function, muscle contraction and fatty acid metabolism were enriched in the Nppb+ working myocardium. We analyzed spatio-temporal expression patterns of several candidate markers (Cacna2d2, Cacna1g, Ephb3, Tnni1), reviewed those of established conduction system markers (Tbx3, Hcn4, Gja5, Cntn2), and placed the patterns in the context of conduction system development. The overview indicates that different properties of conduction system components develop gradually and at different developmental stages, and that chamber myocardium gradually differentiates and diverges from conduction system myocardium until after birth.

Sonic hedgehog pathway activation is associated with cetuximab resistance and EPHB3 receptor induction in colorectal cancer.

A major problem of colorectal cancer (CRC) targeted therapies is relapse caused by drug resistance. In most cases of CRC, patients develop resistance to anticancer drugs. Cetuximab does not show many of the side effects of other anticancer drugs and improves the survival of patients with metastatic CRC. However, the molecular mechanism of cetuximab resistance is not fully understood. Methods: EPHB3-mediated cetuximab resistance was confirmed by in vitro western blotting, colony-forming assays, WST-1 colorimetric assay, and in vivo xenograft models (n = 7 per group). RNA-seq analysis and receptor tyrosine kinase assays were performed to identify the cetuximab resistance mechanism of EPHB3. All statistical tests were two-sided. Results: The expression of EFNB3, which upregulates the EPHB3 receptor, was shown to be increased via microarray analysis. When resistance to cetuximab was acquired, EPHB3 protein levels increased. Hedgehog signaling, cancer stemness, and epithelial-mesenchymal transition signaling proteins were also increased in the cetuximab-resistant human colon cancer cell line SW48R. Despite cells acquiring resistance to cetuximab, STAT3 was still responsive to EGF and cetuximab treatment. Moreover, inhibition of EPHB3 was associated with decreased STAT3 activity. Co-immunoprecipitation confirmed that EGFR and EPHB3 bind to each other and this binding increases upon resistance acquisition, suggesting that STAT3 is activated by the binding between EGFR and EPHB3. Protein levels of GLI-1, SOX2, and Vimentin, which are affected by STAT3, also increased. Similar results were obtained in samples from patients with CRC. Conclusion: EPHB3 expression is associated with anticancer drug resistance.

EZH2-mediated epigenetic suppression of EphB3 inhibits gastric cancer proliferation and metastasis by affecting E-cadherin and vimentin expression.

EphB3 is a member of the EPH family of receptors and has been found to play a role in the carcinogenesis of some human cancers. However, its expression and clinical significance in gastric cancer (GC) have not been well documented. In the present study, we detected the expression of EphB3 in GC and adjacent noncancerous tissues and explored its relationships with the clinicopathological features and prognosis of GC patients. It was found that EphB3 silenced GC cells epigenetically by direct transcriptional repression of GC cells via polycomb group protein EZH2 mediation. EphB3 was downregulated in GC cells and tissues, and EphB3 depletion promoted GC cell growth and invasion, while ectopic overexpression of EphB3 produced a significant anti-tumor effect. EphB3 was found to be involved in epithelial-mesenchymal transition by regulating E-cadherin and vimentin expression. In addition, patients with reduced EphB3 expression had shorter disease-free survival (DFS), indicating that EphB3 may prove to be a biomarker for prognosis of GC. These results demonstrated that EphB3 functioned as a tumor-suppressor and prognostic biomarker in GC.

Screening for therapeutic targets of tumor angiogenesis signatures in 31 cancer types and potential insights.

Tumor vessel normalization can increase pericyte coverage, perfusion efficiency and immune infiltration, while reducing hypoxia, vessel leakage, CTC and metastasis. In this study, we systemically presented the expression pattern of tumor angiogenesis gene signatures in 31 cancer types and its association with immune infiltration and cancer metastasis. Specifically, READ, COAD etc. have relatively similar expression patterns with low GPAGs and high PPAGs. Patients with this expression pattern may benefit from tumor vessel normalization. COAD was selected for further investigation and we found GPAG CXCL12 was downregulated while PPAG EPHB3 was overexpressed in COAD, which were further validated using two independent colon cancer dataset. Further study indicated that CXCL12 expression was positively correlated innate inflammation pathways such as NFκB and negatively correlated with metastasis, while EPHB3 had a reverse result. Moreover, CXCL12 was positively correlated with cancer immune infiltration while EPHB3 was negatively correlated with cancer immune infiltration. Besides, the association between CXCL12/EPHB3 and mutation/CNA landscape were also explored. We also discussed the potential application of gut microbiota in cancer treatment. In summary, blood vessel normalization could promote immune infiltration and repress cancer metastasis while immune cell infiltration can promote blood vessel normalization through a positive feedback loop.

Role of EphB3 Receptor in Mediating Head and Neck Tumor Growth, Cell Migration, and Response to PI3K Inhibitor.

Eph proteins have emerged as critical drivers affecting tumor growth and progression in human malignancies. Our The Cancer Genome Atlas (TCGA) data analysis showed that EphB3, a receptor tyrosine kinase, is frequently coamplified with PIK3CA in head and neck squamous cell carcinoma (HNSCC). We therefore hypothesized that EphB3 amplification plays a protumorigenic role in HNSCC and that EphB3 and PIK3CA are cooperating oncogenes that contribute toward its pathogenesis. This hypothesis was not experimentally supported, because EphB3 knockdown failed to alter HNSCC tumor cell growth in vitro or in vivo with an orthotopic model. However, responsiveness of EphB3 knockdown tumors to the PI3K inhibitor, BKM120, was significantly decreased in terms of both tumor growth delay and survival. This is correlated with an increase in prosurvival proteins, S6 and BcL-XL, in the EphB3 shRNA tumors treated with BKM120 compared with controls. We further observed that EphB3 knockdown resulted in increased migration in vitro and increased EMT gene signature in vivo To explain these results, we examined EphB3 phosphorylation levels in HNSCC at baseline. Although total EphB3 levels were high, we found low phospho-EphB3 levels in HNSCCs. Forced EphB3 phosphorylation with an ephrin-B2-Fc fusion protein resulted in decreased HNSCC migration and cell growth, and enhanced response to BKM120 in vitro These data collectively indicate that progression of HNSCC selects for low/inhibited EphB3 activity to enhance their survival and migratory abilities and decrease response to PI3K signaling. Therefore, strategies focused on activating EphB3 might be helpful to inhibit tumor growth and enhance sensitivity to PI3K inhibitors in HNSCC. Mol Cancer Ther; 17(9); 2049-59. ©2018 AACR.

Altered Maturation of Medullary TEC in EphB-Deficient Thymi Is Recovered by RANK Signaling Stimulation.

In the present study, the relevance of EphB2 and EphB3 tyrosine kinase receptors for the maturation of medullary thymic epithelial cells (TECs) is analyzed. The absence of both molecules, but particularly that of EphB2, courses with altered maturation of medullary Cld3,4hiSSEA1+ epithelial progenitor cells, mature medulla epithelial cells, defined by the expression of specific cell markers, including UEA1, MHCII, CD40, CD80, and AIRE, and reduced expansion of medullary islets. In vivo assays demonstrate that these changes are a consequence of the absence of EphBs in both TECs and thymocytes. On the other hand, the changes, that remains in the adult thymus, correlated well with reduced proportions of E15.5 Vγ5+RANKL+ cells in EphB-deficient thymi that could result in decreased stimulation of RANK+ medullary TECs to mature, a fact that was confirmed by recovering of proportions of both CD40hiCD80+ and MHCIIhiUEA1+ mature medullary TECs of mutant E14.5 alymphoid thymic lobes by agonist anti-RANK antibody treatment. Accordingly, the effects of EphB deficiency on medullary TECs maturation are recovered by RANK stimulation.

Upregulation of the long noncoding RNA FOXD2-AS1 promotes carcinogenesis by epigenetically silencing EphB3 through EZH2 and LSD1, and predicts poor prognosis in gastric cancer.

Accumulating data indicate that long noncoding RNAs (lncRNAs) serve as important modulators in biological processes and are dysregulated in diverse tumors. The function of FOXD2-AS1 in gastric cancer (GC) progression and related biological mechanisms remain undefined. A comprehensive analysis identified that FOXD2-AS1 enrichment was upregulated markedly in GC and positively correlated with a large tumor size, a later pathologic stage, and a poor prognosis. Gene-set enrichment analysis (GSEA) in GEO datasets uncovered that cell cycle and DNA replication associated genes were enriched in patients with high FOXD2-AS1 expression. Loss of FOXD2-AS1 function inhibited cell growth via inhibiting the cell cycle in GC, whereas upregulation of FOXD2-AS1 expression promoted cancer progression. The enhancer of zeste homolog 2 (EZH2) and lysine (K)-specific demethylase 1A (LSD1) proteins were found to serve as binding partners of FOXD2-AS1 and mediators of FOXD2-AS1 function. Mechanically, FOXD2-AS1 promoted GC tumorigenesis partly through EZH2 and LSD1 mediated EphB3 downregulation. The present results revealed that FOXD2-AS1 acted as a tumor inducer in GC partly through EphB3 inhibition by direct interaction with EZH2 and LSD1, and may prove to be a potential biomarker of carcinogenesis.

Formononetin inhibits colon carcinoma cell growth and invasion by microRNA­149­mediated EphB3 downregulation and inhibition of PI3K/AKT and STAT3 signaling pathways.

Formononetin (Form), a phytoestrogen extracted from the roots of Astragalus membranaceus, is one of the fundamental herbs used in traditional Chinese medicine because of its protective effects against certain malignant tumors. However, its role in colon carcinoma cells and the underlying molecular mechanisms have not been completely elucidated. The present study aimed to demonstrate that Form significantly inhibited the proliferation and invasion of the colon carcinoma cell lines SW1116 and HCT116. Mechanistic studies have suggested that Form suppresses colon carcinoma cell growth by downregulating cell cycle­associated protein (cyclin D1) expression and arresting the cell cycle at the G0­G1 checkpoint. Further studies revealed that treatment with Form inhibits matrix metalloproteinase (MMP)2 and MMP9 expression. Aditionally, the results demonstrated that Form significantly increased microRNA (miR)­149 expression. Following miR­149 overexpression in SW1116 and HCT116 cells using an miR­149 mimic, cell viability and Ephrin type­B receptor 3 (EphB3) levels decreased. Furthermore, the inhibitory effects of Form were associated with phosphatidylinositol 3­kinase (PI3K)/protein kinase B (AKT) and signal transducer and activator of transcription 3 (STAT3) signaling pathways. These results indicated the suppressive effect of Form on colon carcinoma cell proliferation and invasion, possibly via miR­149­induced EphB3 downregulation and the inhibition of the PI3K/AKT and STAT3 signaling pathways. Overall, Form may be used as a novel candidate for the clinical treatment of colorectal cancer in the future.

EphB3 signaling induces cortical endothelial cell death and disrupts the blood-brain barrier after traumatic brain injury.

Damage to the cerebrovascular network is a major contributor to dysfunction in patients suffering from traumatic brain injury (TBI). Vessels are composed of lumen-forming endothelial cells that associate closely with both glial and neuronal units to establish a functional blood-brain barrier (BBB). Under normal physiological conditions, these vascular units play important roles in central nervous system (CNS) homeostasis by delivering oxygen and nutrients while filtering out molecules and cells that could be harmful; however, after TBI this system is disrupted. Here, we describe a novel role for a class of receptors, called dependence receptors, in regulating vessel stability and BBB integrity after CCI injury in mice. Specifically, we identified that EphB3 receptors function as a pro-apoptotic dependence receptor in endothelial cells (ECs) that contributes to increased BBB damage after CCI injury. In the absence of EphB3, we observed increased endothelial cell survival, reduced BBB permeability and enhanced interactions of astrocyte-EC membranes. Interestingly, the brain's response to CCI injury is to reduce EphB3 levels and its ligand ephrinB3; however, the degree and timing of those reductions limit the protective response of the CNS. We conclude that EphB3 is a negative regulator of cell survival and BBB integrity that undermine tissue repair, and represents a protective therapeutic target for TBI patients.

EphB receptors, mainly EphB3, contribute to the proper development of cortical thymic epithelial cells.

EphB and their ligands ephrin-B are an important family of protein tyrosine kinase receptors involved in thymocyte-thymic epithelial cell interactions known to be key for the maturation of both thymic cell components. In the present study, we have analyzed the maturation of cortical thymic epithelium in EphB-deficient thymuses evaluating the relative relevance of EphB2 and EphB3 in the process. Results support a relationship between the epithelial hypocellularity of mutant thymuses and altered development of thymocytes, lower proportions of cycling thymic epithelial cells and increased epithelial cell apoptosis. Together, these factors induce delayed development of mutant cortical TECs, defined by the expression of different cell markers, i.e. Ly51, CD205, MHCII, CD40 and ß5t. Furthermore, although both EphB2 and EphB3 are necessary for cortical thymic epithelial maturation, the relevance of EphB3 is greater since EphB3-/- thymic cortex exhibits a more severe phenotype than that of EphB2-deficient thymuses.

EphB3-targeted regulation of miR-149 in the migration and invasion of human colonic carcinoma HCT116 and SW620 cells.

microRNAs play key roles during various crucial cell processes such as proliferation, migration, invasion and apoptosis. Also, microRNAs have been shown to possess oncogenic and tumor-suppressive functions in human cancers. Here, we describe the regulation and function of miR-149 in colorectal cancer cell lines. miR-149 expression patterns were detected in human colorectal cell lines and tissue samples, and then focused on its role in regulation of cell growth, migration, invasion, and its target gene identification. Furthermore, the function of the target gene of miR-149 was analyzed in vitro and in vivo. miR-149 expression was downregulated in human colorectal cancer HCT116 and SW620 cell lines compared to the normal colon epithelial NCM460 cell line using quantitative real-time polymerase chain reaction methods. Further studies indicated that introduction of miR-149 was able to suppress cell migration and invasion. Then, EphB3 was identified as a direct target gene of miR-149 in colorectal cancer cells. Moreover, experiments in vitro showed that knockdown expression of EphB3 could suppress cell proliferation and invasion, and ectopic expression of EphB3 restored the phenotypes of CRC cell lines transfected with miR149. In addition, silencing of EphB3 significantly affected cycle progression distribution and increased apoptosis in CRC cell lines. Finally, in vivo results demonstrated that knockdown of EphB3 by siRNA inhibited tumor growth. In conclusion,the important role of miR-149 in colorectal cancer progression suggesting that miR-149 may serve as a therapeutic target for colorectal cancer treatment.

EphB3 Stimulates Cell Migration and Metastasis in a Kinase-dependent Manner through Vav2-Rho GTPase Axis in Papillary Thyroid Cancer.

Eph receptors, the largest subfamily of transmembrane tyrosine kinase receptors, have been increasingly implicated in various physiologic and pathologic processes, and the roles of the Eph family members during tumorigenesis have recently attracted growing attentions. In the present study, we explored the function of EphB3, one member of Eph family, in papillary thyroid cancer (PTC). We found that the expression of EphB3 was significantly elevated in PTC. Either overexpression of EphB3 or activation of EphB3 by EfnB1-Fc/EfnB2-Fc stimulated in vitro migration of PTC cells. In contrast, siRNA-mediated knockdown of EphB3 or EphB3-Fc treatment, which only blocked EphB3-mediated forward signaling, inhibited migration and metastasis of PTC cells. A mechanism study revealed that EphB3 knockdown led to suppressed activity of Rac1 and enhanced activity of RhoA. Moreover, we found that Vav2, an important regulator of Rho family GTPases, was activated by EphB3 in a kinase-dependent manner. Altogether, our work suggested that EphB3 acted as a tumor promoter in PTC by increasing the in vitro migration as well as the in vivo metastasis of PTC cells through regulating the activities of Vav2 and Rho GTPases in a kinase-dependent manner.

TCF7L1 Modulates Colorectal Cancer Growth by Inhibiting Expression of the Tumor-Suppressor Gene EPHB3.

Dysregulation of the Wnt pathway leading to accumulation of ß-catenin (CTNNB1) is a hallmark of colorectal cancer (CRC). Nuclear CTNNB1 acts as a transcriptional coactivator with TCF/LEF transcription factors, promoting expression of a broad set of target genes, some of which promote tumor growth. However, it remains poorly understood how CTNNB1 interacts with different transcription factors in different contexts to promote different outcomes. While some CTNNB1 target genes are oncogenic, others regulate differentiation. Here, we found that TCF7L1, a Wnt pathway repressor, buffers CTNNB1/TCF target gene expression to promote CRC growth. Loss of TCF7L1 impaired growth and colony formation of HCT116 CRC cells and reduced tumor growth in a mouse xenograft model. We identified a group of CTNNB1/TCF target genes that are activated in the absence of TCF7L1, including EPHB3, a marker of Paneth cell differentiation that has also been implicated as a tumor suppressor in CRC. Knockdown of EPHB3 partially restores growth and normal cell cycle progression of TCF7L1-Null cells. These findings suggest that while CTNNB1 accumulation is critical for CRC progression, activation of specific Wnt target genes in certain contexts may in fact inhibit tumor growth.

Identification of an Interfering Ligand Aptamer for EphB2/3 Receptors.

The Eph receptors are transmembrane proteins that belong to the receptor tyrosine kinases superfamily. Elevated Eph/ephrin expression levels have been associated with angiogenesis and tumor vasculature in many types of human cancers, including breast, lung, and prostate cancers, melanoma, and leukemia. In glioblastoma (GBM), the dysregulated expression of Eph receptors and of corresponding ephrin ligands has been associated with higher tumor grade and poor prognosis making them effective targets for therapeutic drugs. In this study, we describe the GL43.T, an anti-Eph aptamer, able to bind at high-affinity EphB3 and EphB2. Moreover, the GL43.T aptamer inhibits the glioma cell vitality and interferes with ephrine-B1 inhibition of chemotactic serum-stimulated cell migration. GL43.T aptamer represents a promising therapeutic molecule for EphB3-dependent cancers.