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The Eph (erythropoietin-producing human hepatocellular) receptor family, the largest subclass of receptor tyrosine kinases (RTKs), plays essential roles in embryonic development and neurogenesis. The intracellular Sterile Alpha Motif (SAM) domain presents a critical structural feature that distinguishes Eph receptors from other RTKs and participates in recruiting and binding downstream molecules. This study identified SASH1 (SAM and SH3 domain containing 1) as a novel Eph receptor-binding partner through SAM-SAM domain interactions. Our comprehensive biochemical analyses revealed that SASH1 selectively interacts with Eph receptors via its SAM1 domain, displaying the highest affinity for EphA8. The high-resolution crystal structure of the EphA8-SASH1 complex provided insights into the specific intermolecular interactions between these proteins. Cellular assays confirmed that EphA8 and SASH1 co-localize and co-precipitate in mammalian cells, with cancer mutations (EphA8 R942H or G978D) impairing this interaction. We demonstrated that SAM-SAM interaction is critical for SASH1-mediated regulation of EphA8 kinase activity, shedding new light on the Eph signaling pathway and expanding our understanding of the molecular basis of the tumor suppressor gene SASH1., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Background: Eph receptors tyrosine kinase (RTK) were identified in 1987 from hepatocellular carcinoma cell lines and were the largest known subfamily of RTK. Eph receptors can be divided into two categories, EphA and EphB, based on their structure and receptor-ligand specificity. EphA can be divided into 10 species (EphA 1-10) and EphB into 6 species (EphB1-6). Similarly, the ligands of Eph receptors are Ephrins. Ephrins also can be divided into Ephrin A and Ephrin B, of which there are five species(Ephrin-A1-5) and three species(Ephrin-B1-3). Among the Eph receptors, EphA1 has been the least studied so far. As far as we know, Eph receptors are involved in multiple pathologies, including cancer progression, tumor angiogenesis, intestinal environmental stability, the lymph node system, neurological disease, and inhibition of nerve regeneration after injury. There is a link between EphA1, integrin and ECM- related signal pathways. Ephrin-A1 is a ligand of the EphA1 receptor. EphA1 and ephrin-A1 functions are related to tumor angiogenesis. EphA1 and ephrin-A1 also play roles in gynecological diseases. Ephrin-A1 and EphA1 receptors regulate the follicular formation, ovulation, embryo transport, implantation and placental formation, which are of great significance for the occurrence of gynecological tumor diseases. EphA1 has been identified as an oncoprotein in various tumors and has been associated with the prognosis of various tumors in recent years. EphA1 is considered a driver gene in tumor genomics. There are significant differences in EphA1 expression levels in different types of normal tissues and tumors and even in different stages of tumor development, suggesting its functional diversity. Changes at the gene level in cell biology are often used as biological indicators of cancer, known as biomarkers, which can be used to provide diagnostic or prognostic information and are valuable for improving the detection, monitoring and treatment of tumors. However, few prognostic markers can selectively predict clinically significant tumors with poor prognosis. These malignancies are more likely to progress and lead to death, requiring more aggressive treatment. Currently available treatments for advanced cancer are often ineffective, and treatment options are mainly palliative. Therefore, early identification and treatment of those at risk of developing malignant tumors are crucial. Although pieces of evidence have shown the role of EphA1 in tumorigenesis and development, its specific mechanism is still unknown to a great extent., Objective: This review reveals the changes and roles of EphA1 in many tumors and cancers. The change of EphA1 expression can be used as a biological marker of cancer, which is valuable for improving tumor detection, monitoring and treatment and can be applied to imaging. Studies have shown that structural modification of EphA1 could make it an effective new drug. EphA1 is unique in that it can be considered a prognostic marker in many tumors and is of important meaning for clinical diagnosis and operative treatment. At the same time, the study of the specific mechanism of EphA1 in tumors can provide a new way for targeted therapy., Methods: Relevant studies were retrieved and collected through the PubMed system. After determining EphA1 as the research object, by analyzing research articles on EphA1 in the PubMed system in recent 10 years, we found that EphA1 was closely connected with the occurrence and development of tumors and further determined the references according to the influencing factors for review and analysis., Results: EphA1 has been identified as a cancer protein in various tumors, such as hepatocellular carcinoma, nasopharyngeal carcinoma, ovarian cancer, gastric cancer, colorectal cancer, clear cell renal cell carcinoma, esophageal squamous cell carcinoma, breast cancer, prostate cancer and uveal melanoma. EphA1 is abnormally expressed in these tumor cells, which mainly plays a role in cancer progression, tumor angiogenesis, intestinal environmental stability, the lymph node system, nervous system diseases and gynecological diseases. In a narrow sense, EphA1 is especially effective in breast cancer in terms of gynecological diseases. However, the specific mechanism of EphA1 leading to the change of cancer cells in some tumors is not clear, which needs further research and exploration., Conclusion: RTK EphA1 can be used as a biomarker for tumor diagnosis (especially a prognostic marker), an indispensable therapeutic target for new anti-tumor therapies, and a novel anti-tumor drug., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

While the typical role of receptor tyrosine kinases is to receive and transmit signals at the cell surface, in some cellular contexts (particularly transformed cells) they may also act as nuclear proteins. Aberrant nuclear localization of receptor tyrosine kinases associated with transformation often enhances the transformed phenotype (i.e. nuclear ErbBs promote tumor progression in breast cancer). Rhabdomyosarcoma (RMS), the most common soft tissue tumor in children, develops to resemble immature skeletal muscle and has been proposed to derive from muscle stem/progenitor cells (satellite cells). It is an aggressive cancer with a 5-year survival rate of 33% if it has metastasized. Eph receptor tyrosine kinases have been implicated in the development and progression of many other tumor types, but there are only two published studies of Ephs localizing to the nucleus of any cell type and to date no nuclear RTKs have been identified in RMS. In a screen for protein expression of Ephs in canine RMS primary tumors as well as mouse and human RMS cell lines, we noted strong expression of EphA1 in the nucleus of interphase cells in tumors from all three species. This localization pattern changes in dividing cells, with EphA1 localizing to the nucleus or the cytoplasm depending on the phase of the cell cycle. These data represent the first case of a nuclear RTK in RMS, and the first time that EphA1 has been detected in the nucleus of any cell type., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2022. Published by The Company of Biologists Ltd.)

MicroRNAs (miRNAs) have critical roles in colorectal cancer (CRC) tumorigenesis and development. It has been reported that Eph receptor A7 (EphA7) was a potential target of miR-944 which is transcriptionally activated in cancer. The aim of this study was to explore the expression profile of miR-944 and its target gene EPHA7 in the serum of Egyptian CRC patients. 150 CRC patients, 50 adenomatous polyps (AP) patients, and 100 healthy controls were included. Serum miR-944 was downregulated (0.304 ± 0.0512) while serum EPHA7 was upregulated (3.163 ± 0.610) in CRC and AP patients versus controls and discriminated aganst these groups by Receiver operating characteristic curve (ROC) analysis. miR-944 presented the highest diagnostic accuracy for CRC patients from control (AUC = 0.90). Moreover obvious prognostic power in distinguishing AP from CRC (AUC = 0.87). In conclusion, miR-944 and EPHA7 are potential genetic markers of CRC predisposition and novel potential non-invasive diagnostic biomarkers for CRC.

Autonomous heavy-chain variable (V H ) domains are the smallest functional antibody fragments, and they possess unique features, including small size and convex paratopes, which provide enhanced targeting of concave epitopes that are difficult to access with larger conventional antibodies. However, human V H domains have evolved to fold and function with a light chain partner, and alone, they typically suffer from low stability and high aggregation propensity. Development of autonomous human V H domains, in which aggregation propensity is reduced without compromising antigen recognition, has proven challenging. Here, we used an autonomous human V H domain as a scaffold to construct phage-displayed synthetic libraries in which aspartate was systematically incorporated at different paratope positions. In selections, the library yielded many anti-EphA1 receptor V H domains, which were characterized in detail. Structural analyses of a parental anti-EphA1 V H domain and an improved variant provided insights into the effects of aspartate and other substitutions on preventing aggregation while retaining function. Our naïve libraries and in vitro selection procedures offer a systematic approach to generating highly functional autonomous human V H domains that resist aggregation and could be used for basic research and biomedical applications., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Gastric cancer (GC) is a common cancer and a leading cause of cancer-related deaths worldwide. Recent studies have supported the important role of long non-coding RNAs (lncRNAs) in GC progression. This study identified functional significance of X inactive specific transcript (XIST) in GC. The expression of XIST and EPHA1 in GC tissues and cells was measured. Then, dual luciferase reporter gene assay, RNA immunoprecipitation (RIP) assay and Chromatin Immunoprecipitation (ChIP) assay were performed to explore the interaction among XIST, EPHA1 and HNF4A. The effects of XIST on GG progression were evaluated by determining expression of proliferation- and invasion-related proteins (Ki67, PCNA, MMP-2, and MMP-9). Further, the functional role of XIST in GC with the involvement of NFκB pathway was also analyzed. Subsequently, the tumor growth in nude mice was evaluated. High expression of XIST and EPHA1 was observed in GC. XIST elevated EPHA1 expression by recruiting HNF4A. In addition, silencing of XIST inhibited GC progression in vitro and in vivo. Overexpressed XIST and EPHA1 yielded a reversed effect on cell proliferation and invasion. SN50 treatment (inhibitor of NFκB pathway) counteracted the promotive effect on GC cell proliferation and invasion mediated by XIST. The present study unveils that XIST increases the enrichment of HNF4A in the promoter region of EPHA1, thus promoting the deterioration of GC.

Genome-wide association studies (GWAS) have identified several susceptibility loci of Alzheimer's disease (AD), which were mainly located in noncoding regions of the genome. Meanwhile, the putative biological mechanisms underlying AD susceptibility loci were still unclear. At present, identifying the functional variants of AD pathogenesis remains a major challenge. Herein, we first used summary data-based Mendelian randomization (SMR) with AD GWAS summary and expression quantitative trait loci (eQTL) data to identify variants who affects expression levels of nearby genes and contributed to the risk of AD. Using the SMR integrative analysis, we totally identified 14 SNPs significantly affected the expression level of 16 nearby genes in blood or brain tissues and contributed to the AD risk. Then, to confirm the results, we replicated the GWAS and eQTL results across multiple samples. Totally, four risk SNP (rs11682128, rs601945, rs3935067, and rs679515) were validated to be associated with AD and affected the expression level of nearby genes (BIN1, HLA-DRA, EPHA1-AS1, and CR1). Besides, our differential expression analysis showed that the BIN1 gene was significantly downregulated in the hippocampus (P = 2.0 × 10 -3 ) and survived after multiple comparisons. These convergent lines of evidence suggest that the BIN1 gene identified by SMR has potential roles in the pathogenesis of AD. Further investigation of the roles of the BIN1 gene in the pathogenesis of AD is warranted.

Background: Ephrin type-A receptors (EPHA) are members of family of receptor tyrosine kinases and are related to tumor immunogenicity and immune microenvironment, however, the association between EPHA mutation ( EPHA mut ) and efficacy of immune checkpoint inhibitors (ICIs) has not been investigated in non-small cell lung cancer (NSCLC)., Methods: Multiple cohorts were used to assess the immunotherapeutic predictive performance of EPHA mut , including one discovery cohort (n=79) and two public validation cohort (cohort 1: NSCLC, n=165; cohort 2: pan-cancer, n=1662). The Cancer Genome Atlas cohort was used for prognostic analysis and mechanism exploration., Results: In the discovery cohort, patients with EPHA mut had superior disease control rate (72.2% vs 36.1%, p=0.01) and progression-free survival (PFS) (HR 0.38; 95% CI 0.21 to 0.68; p<0.001) compared with those with wide-type EPHA ( EPHA wt ) in NSCLC. The association between EPHA mut and immunotherapy outcomes in NSCLC was consistently observed in the validation cohorts by multivariable models (cohort 1, PFS HR 0.59; 95% CI 0.37 to 0.96; p=0.03; cohort 2, overall survival (OS) HR 0.63; 95% CI 0.41 to 0.98; p=0.04). Further pooled estimates of the discovery and validation cohorts showed that patients with EPHA mut exhibited a significantly longer PFS and OS in lung adenocarcinoma (LUAD) while not squamous cell lung cancer (LUSC). Consistently, mechanism analysis revealed that patients with EPHA mut was associated with increased T cell signatures and downregulated transforming growth factor-β signaling compared with patients with EPHA wt in LUAD while not LUSC., Conclusions: Our results demonstrated that EPHA mut is an independent classifier that could stratify patients with LUAD for ICIs therapy. Further prospective studies are warranted., Trial Registration Number: NCC2016JZ-03, NCC2018-092., Competing Interests: Competing interests: CL, YX, GW and SC is the employee of Burning Rock Biotech. WX and ZZ is the employee of 3D Medicines., (© Author(s) (or their employer(s)) 2020. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

One of the reasons for recurrence following treatment of high grade serous ovarian carcinoma (HGSOC) is the persistence of residual cancer stem cells (CSCs). There has been variability between laboratories in the identification of CSC markers for HGSOC. We have identified new surface markers (CD24, CD9 and EPHA1) in addition to those previously known (CD44, CD117 and CD133) using a bioinformatics approach. The expression of these surface markers was evaluated in ovarian cancer cell lines, primary malignant cells (PMCs), normal ovary and HGSOC. There was no preferential expression of any of the markers or a combination. All the markers were expressed at variable levels in ovarian cancer cell lines and PMCs. Only CD117 and CD9 were expressed in the normal ovarian surface epithelium and fallopian tube. Both ALDEFLUOR (ALDH1A1) and side population assays identified a small proportion of cells (<3%) separately that did not overlap with little variability in cell lines and PMCs. All surface markers were co-expressed in ALDH1A1+ cells without preference for one combination. The cell cycle analysis of ALDH1A1+ cells alone revealed that majority of them reside in G0/G1 phase of cell cycle. Further separation of G0 and G1 phases showed that ALDH1A1+ cells reside in G1 phase of the cell cycle. Xenograft assays showed that the combinations of ALDH1A1 + cells co-expressing CD9, CD24 or EPHA1 were more tumorigenic and aggressive with respect to ALDH1A1-cells. These data suggest that a combined approach could be more useful in identifying CSCs in HGSOC., Competing Interests: Declaration of competing interest The authors declare that they have no conflict of interest., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Large-scale genome-wide studies have revealed the role of several genes and their respective single-nucleotide polymorphisms (SNPs) in the pathophysiology of late-onset Alzheimer's disease (LOAD). Here, the frequencies of ABCA7 SNPs rs3764650 and rs4147929 and EphA1 SNP rs11771145 were assessed and compared in LOAD patients and healthy subjects. In a case-control study, 110 patients with LOAD (case) and 88 healthy unrelated age- and gender-matched individuals (control), both from Azeri descent, were enrolled. DNA was extracted from blood samples using the salting out method, and the genotyping was performed by RFLP-PCR for rs3764650, rs4147929, and rs11771145 polymorphisms. Electrophoresis was carried out on agarose gel. Sequencing was utilized for confirmation of the results. No differences were found in the frequencies of ABCA7 SNP rs3764650 and EphA1 SNP rs11771145 between healthy subjects and LOAD patients. However, a significant difference was revealed in the frequencies of AA (p = 0.042, OR = 0.150; 95%CI = 0.005-1.410) and GG (p = 0.009, OR = 1.716; 95%CI = 0.918-3.218) genotypes of ABCA7 SNP rs4147929 between the mentioned groups. This study showed that ABCA7 SNP rs4147929 might be a predisposing factor for LOAD. However, such an association was not found between ABCA7 SNP rs3764650 as well as EphA1 SNP rs11771145 and LOAD. These results must be confirmed in other ethnic groups.

Aims: Although converging evidence from experimental and epidemiological studies indicates Alzheimer's disease (AD) and ischemic stroke (IS) are related, the genetic basis underlying their links is less well characterized. Traditional SNP-based genome-wide association studies (GWAS) have failed to uncover shared susceptibility variants of AD and IS. Therefore, this study was designed to investigate whether pleiotropic genes existed between AD and IS to account for their phenotypic association, although this was not reported in previous studies., Methods: Taking advantage of large-scale GWAS summary statistics of AD (17,008 AD cases and 37,154 controls) and IS (10,307 IS cases and 19,326 controls), we performed gene-based analysis implemented in VEGAS2 and Fisher's meta-analysis of the set of overlapped genes of nominal significance in both diseases. Subsequently, gene expression analysis in AD- or IS-associated expression datasets was conducted to explore the transcriptional alterations of pleiotropic genes identified., Results: 16 AD-IS pleiotropic genes surpassed the cutoff for Bonferroni-corrected significance. Notably, MS4A4A and TREM2, two established AD-susceptibility genes showed remarkable alterations in the spleens and brains afflicted by IS, respectively. Among the prioritized genes identified by virtue of literature-based knowledge, most are immune-relevant genes (EPHA1, MS4A4A, UBE2L3 and TREM2), implicating crucial roles of the immune system in the pathogenesis of AD and IS., Conclusions: The observation that AD and IS had shared disease-associated genes offered mechanistic insights into their common pathogenesis, predominantly involving the immune system. More importantly, our findings have important implications for future research directions, which are encouraged to verify the involvement of these candidates in AD and IS and interpret the exact molecular mechanisms of action., (© 2019 The Authors. CNS Neuroscience & Therapeutics Published by John Wiley & Sons Ltd.)

Gap junctions are intercellular conduits that permit the passage of ions, small metabolites, and signaling molecules between cells. Connexin32 (Cx32) is a major gap junction protein in the liver and brain. Phosphorylation is integral to regulating connexin assembly, degradation, and electrical and metabolic coupling, as well as to interactions with molecular partners. Cx32 contains two intracellular tyrosine residues, and tyrosine phosphorylation of Cx32 has been detected after activation of the epidermal growth factor receptor; however, the specific tyrosine residue and the functional implication of this phosphorylation remain unknown. To address the limited available information on Cx32 regulation by tyrosine kinases, here we used the Cx32 C-terminal (CT) domain in an in vitro kinase-screening assay, which identified ephrin (Eph) receptor family members as tyrosine kinases that phosphorylate Cx32. We found that EphB1 and EphA1 phosphorylate the Cx32CT domain residue Tyr 243 Unlike for Cx43, the tyrosine phosphorylation of the Cx32CT increased gap junction intercellular communication. We also demonstrated that T-cell protein-tyrosine phosphatase dephosphorylates pTyr 243 The data presented above along with additional examples throughout the literature of gap junction regulation by kinases, indicate that one cannot extrapolate the effect of a kinase on one connexin to another., (© 2019 Trease et al.)

Primary Objective: Eph/Ephrin signaling is inhibitory for developing axons and blocking Eph pathways enhances regeneration after spinal cord injury. It was hypothesized that inhibition of Eph signaling promotes cellular and behavioral recovery after traumatic brain injury (TBI). Research design: Lateral fluid percussion (LFP) injury was performed on wildtype (WT) and EphA6 knockout (KO) mice. EphA6-Fc, Ephrin-A5-Fc fusion proteins, and sodium orthovanadate were used to alter the signaling pathway. Immunohistochemistry and tissue explants revealed cellular changes. Rotarod tests demonstrated vestibulomotor function. Outcomes: The EphA6 receptor expression is upregulated following LFP. Uninjured EphA6 KO mice exhibit greater neurite density and clustered Ephrin-A5-Fc causes growth cone collapse in vitro. After LFP, EphA6 KO mice demonstrate longer neurites and decreased neuronal cell death and astrocytosis compared to WT mice. Blocking EphA signaling by soluble EphA6-Fc fusion protein reduces cell death and improves motor function following LFP whereas clustered Ephrin-A5-Fc exacerbates cell death and neurodegeneration. Sodium orthovanadate rescues growth cone collapse in vitro as well as cell death and neurodegeneration in vivo. Conclusions: Eph/Ephrin signaling plays an inhibitory role following TBI. Targeting the Eph signaling pathway with Fc fusion proteins and pharmacological agents can be a novel strategy to counter the damaging effects of TBI. Abbreviations: LFP: lateral fluid percussion; TBI: traumatic brain injury; KO: knockout; WT: wildtype; PTP2: protein phosphotyrosine phosphatase 2; Tg: transgenic; YFP: yellow fluorescent protein; ATM: atmospheres; RT-qPCR: Real-time-quantitative PCR; dpi: days post injury; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; DAPI: 4',6-diamidino-2-phenylindole; PBS: phosphate buffered saline; GFAP: glial fibrillary acidic protein; FLJC: fluorojade C; CA: cornu ammonis; SEM: standard error of the mean; ANOVA: analysis of variance; PLSD: posthoc least significant difference.

Mild behavioral impairment (MBI) describes the emergence of later-life neuropsychiatric symptoms (NPS) as an at-risk state for incident cognitive decline and dementia, and for some as a potential manifestation of prodromal dementia. How NPS mechanistically link to the development of mild cognitive impairment and Alzheimer's disease (AD) is not fully understood, with potential mechanisms including shared risk factors related to both NPS and cognitive impairment, or AD pathology promoting NPS. This is the first exploratory study to examine whether AD genetic loci as a genetic risk score (GRS), or individually, are a shared risk factor with MBI. Participants were 1,226 older adults (aged 72-79; 738 males; 763 normal cognition) from the Personality and Total Health Through Life project. MBI was approximated in accordance with Criterion 1 of the ISTAART-AA diagnostic criteria using a transformation algorithm for the neuropsychiatric inventory. A GRS was constructed from 25 AD risk loci. Binomial logistic regression adjusting for age, gender, and education examined the association between GRS and MBI. A higher GRS and APOE*ε4 were associated with increased likelihood of affective dysregulation. Nominally significant associations were observed between MS4A4A-rs4938933*C and MS4A6A-rs610932*G with a reduced likelihood of affective dysregulation; ZCWPW1-rs1476679*C with a reduced likelihood of social inappropriateness and abnormal perception/thought content; BIN1-rs744373*G and EPHA1-rs11767557*C with higher likelihood of abnormal perception/thought content; NME8-rs2718058*G with a reduced likelihood of decreased motivation. These preliminary findings suggest a common genetic etiology between MBI and traditionally recognized cognitive problems observed in AD and improve our understanding of the pathophysiological features underlying MBI., (© 2018 Wiley Periodicals, Inc.)

The objective of this study was to ascertain whether mRNA and protein expressions of implantation-related genes (erythropoietin-producing hepatocellular receptor-ligand A1, Eph-ephrin A1 and leptin receptor-leptin, LEPR-LEP) differed between pigs with high and low number of embryos, and whether these differences in gene expression might affect embryo implantation. Experimental pig groups (n = 24) for high and low number of embryos were prepared by altering the number of eggs ovulated in pre-pubertal gilts treated with 1.5 × (High) or 1.0 × (Low) PG600 ([400 IU PMSG + 200 IU hCG]/dose, AKZO-NOBEL). Gilts expressing oestrus were artificially inseminated twice and maintained in breeding and gestation until the reproductive tract was collected on day 22 of pregnancy. At slaughter, the reproductive tracts from each pregnant gilt from each treatment were immediately processed to collect samples for RNA and protein analysis. Within each gilt, three conceptus points were sampled, one from each horn and then a random conceptus within the tract. At each conceptus point, endometrial attachment site, chorion-allantois and embryo were collected and immediately frozen in liquid nitrogen. Number of corpus luteum (CL) (35.4 vs. 12.6) and total embryo number (18.8 vs. 10.2) were greater in the high-embryo compared to the low-embryo group, respectively (p < .05). Real-time qPCR results showed that Eph-ephrin A1 mRNA expression was less in the high-embryo (p < .05) compared to the low-embryo group. In addition, Western blotting analysis indicated that Eph-ephrin A1 and LEP protein expression at endometrial attachment site in high-embryo was less (p < .05) compared to low-embryo group. It was also noted that mRNA expression of Eph-ephrin A1 and LEPR-LEP was greater in pregnant than non-pregnant gilts (p < .05). Moreover, mRNA expression of Eph-ephrin A1 (p < .05) and LEPR-LEP was greatest at endometrial attachment site among all three tissues. There was a positive correlation between expressions of Eph-ephrin A1, LEPR-LEP and embryo length with the correlation coefficient 0.31-0.59. For Eph-ephrin A1, the highest correlation coefficient appeared between Eph A1 expression and normal embryo number, between ephrin A1 expression and embryo length. For LEPR-LEP, the highest correlation coefficient appeared between LEPR-LEP expression and ovary weight (0.79 for both, p < .05), followed by embryo length and weight. The results of this study suggest that low expression of Eph-ephrin A1 and LEPR-LEP is somehow related to increased embryo number during implantation and that endometrial attachment site might be the main target tissue of these gene products. Yet, the increased expression of Eph-ephrin A1 and LEPR-LEP appeared associated with increased embryo growth (length and weight) and ovary weight, Eph-ephrin A1 and LEPR-LEP might play roles in the regulation of embryo implantation in pigs., (© 2018 Blackwell Verlag GmbH.)

Eph receptor‑A1 (EphA1) was the first member of the erythropoietin producing hepatocellular carcinoma (Eph) family of receptor tyrosine kinases. Although the roles of EphA1 in the tumorigenesis of various human cancers have been investigated, few studies have focused on ovarian carcinoma. The present study aimed to explore the profile of EphA1 expression in ovarian carcinomas, to analyzed the association between EphA1 expression and clinicopathologic parameters, and to investigate the roles of overexpressed EphA1 in ovarian cancer cells. EphA1 protein was detected in ovarian cancer cell lines and in a set of formalin‑fixed tissues, including normal fallopian tube, ovarian benign serous cystadenoma, borderline serous tumors and serous carcinoma. Ovarian cancer cell lines HO8910 and A2780 were transiently transfected with EphA1‑pCMV6‑GFP plasmid, and the proliferation and apoptosis of cells were measured. The association between EphA1 expression and clinicopathological parameters was statistically analyzed. EphA1 expression was negative in HO8910 and weakly positive in A2780 cells. The proliferation rate was significantly reduced in ovarian cancer cells after transfection with EphA1 plasmid compared with cells transfected with mock plasmid or untreated cells, but no obvious alteration in apoptosis was detected among these groups. EphA1 expression was positively detected in all normal fallopian tubes (10/10, 100%) and ovarian benign serous cystadenomas (12/12, 100%) as well as in some borderline serous tumors (9/15, 60%) and ovarian serous carcinomas (33/76, 43.42%). EphA1 expression was associated with grade of ovarian serous carcinomas, with loss of EphA1 more often observed in high‑grade tumors (P=0.016) and high Ki67 index tumors (P=0.007). These data suggest that EphA1 might be a useful marker for distinguishing low grade from high‑grade ovarian serous carcinoma.

Background/aim: Erythropoietin-producing hepatocellular carcinoma receptor A (EphA) is associated with angiogenesis and invasive tumor progression. In this study, we evaluated the EphA1-4 expression levels in advanced gastric cancer., Patients and Methods: Tumor tissues obtained from 114 patients with advanced gastric adenocarcinoma who underwent gastrectomy were analyzed. In addition, the impact of EPHA 1-4 mRNA expression on survival was analyzed using the Kaplan-Meier plotter database on the website., Results: High EphA 1, 2, and 4 expression levels were significantly related to recurrence (p<0.01, p=0.04, and p<0.01). Both high EphA 1 and 4 expression levels were independent predictors of relapse-free interval (hazard ratio [HR]=2.0, p=0.03; HR=2.4, p=0.03) and disease-specific survival (HR=2.0, 95% p=0.03; HR=2.5, p=0.02) on multivariate analysis. In the Kaplan-Meier plotter database, high EPHA2 mRNA expression was significantly associated with poor survival in patients with gastric cancer (p=0.0098), and high expression levels of EPHA1 and 4 tended to be associated with poor survival (p=0.050, p=0.052)., Conclusion: EphA 1, 2, and 4 may play key roles in recurrence and survival in patients with advanced gastric cancer., (Copyright© 2018, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.)

Large-scale genome-wide association studies have reported EPHA1 rs11767557 variant to be associated with Alzheimer's disease (AD) risk in the European population. However, it is still unclear how this variant functionally contributes to the underlying disease pathogenesis. The rs11767557 variant is located approximately 3 kb upstream of EPHA1 gene. We think that rs11767557 may modify the expression of nearby genes such as EPHA1 and further cause AD risk. Until now, the potential association between rs11767557 and the expression of nearby genes has not been reported in previous studies. Here, we evaluate the potential expression association between rs11767557 and EPHA1 using multiple large-scale eQTLs datasets in human brain tissues and the whole blood. The results show that rs11767557 variant could significantly regulate EPHA1 gene expression specifically in human whole blood. These findings may further provide important supplementary information about the regulating mechanisms of rs11767557 variant in AD risk.

Background/aim: Overexpression of erythropoietin-producing hepatocellular A1 (EPHA1), a member of the EPH super family, is frequently observed in various cancer types. The dysregulated interaction of EPHA1 with its ligand Ephrin A1 has been linked to the progression of ovarian cancer (OC). However, the contribution of EPHA1 in the regulation of the aggressive properties of OC cells remains unknown., Materials and Methods: In this study we investigated the differential expression of EPHA1 in human OC cells. The EPHA1 gene was knocked-down using the CRISPR/Cas9 technique to evaluate its effect on the progressive properties of OC cells., Results: After EPHA1 was knocked-down using a CRISPR/CAS9 genomic editing system in OC cells (SKOV3 and COV504), we observed cell-cycle arrest at the G 0 /G 1 phases in both OC cell lines. Knockdown of EPHA1 in the two OC cells inhibited their aggressive traits, including proliferation, invasion and migration, as well as improving their attachment to extracellular matrix. EPHA1 may play a role in OC through its regulation of multiple signaling pathways, such as matrix metalloproteinase-2 (MMP2), extracellular signal-regulated kinase 2 (ERK2) and proto-oncogene c-MYC., Conclusion: EPHA1 may promote the aggression of some OC cells and, thus, be considered a potential therapeutic target for the treatment of malignant OC., (Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.)

Proper function of the neural network results from the precise connections between axons and dendrites of presynaptic and postsynaptic neurons, respectively. In the Drosophila olfactory system, the dendrites of projection neurons (PNs) stereotypically target one of ∼50 glomeruli in the antennal lobe (AL), the primary olfactory center in the brain, and form synapses with the axons of olfactory receptor neurons (ORNs). Here, we show that Eph and Ephrin, the well-known axon guidance molecules, instruct the dendrodendritic segregation during the discrete olfactory map formation. The Eph receptor tyrosine kinase is highly expressed and localized in the glomeruli related to reproductive behavior in the developing AL. In one of the pheromone-sensing glomeruli (DA1), the Eph cell-autonomously regulates its dendrites to reside in a single glomerulus by interacting with Ephrins expressed in adjacent PN dendrites. Our data demonstrate that the trans interaction between dendritic Eph and Ephrin is essential for the PN dendritic boundary formation in the DA1 olfactory circuit, potentially enabling strict segregation of odor detection between pheromones and the other odors., (© 2017 Anzo et al.; Published by Cold Spring Harbor Laboratory Press.)

The Eph family of receptor tyrosine kinases serves as key modulators of various cellular functions, including inflammation, hypertrophy and fibrosis. Recent analyses have revealed that a member of the Eph family, EphA1, plays a pivotal role in regulating insulin metabolism and kidney injury. However, the importance of EphA1 in diabetic nephropathy has not been recognized. We established a diabetic nephropathy mouse model using a high-fat diet and streptozotocin (STZ) injection. Then, the recombinant adeno-associated virus type 9 (AAV9) overexpressing EphA1 or a negative control was injected locally into the kidney. Metabolite testing and histopathological analyses of kidney fibrosis, pancreatic islet function and signaling pathways were evaluated. Our study showed that hyperglycemia, insulin resistance, and renal fibrosis accompanied the deterioration of kidney function in diabetic mice. The overexpression of EphA1 in the kidney attenuated renal fibrosis and improved kidney function but did not affect systemic glucose metabolism and pancreatic islet function. Furthermore, the overexpression of EphA1 decreased the phosphorylation of ERK1/2, JNK and MYPT1 (a substrate of Rho kinase). The overexpression of EphA1 can be therapeutically targeted to inhibit diabetic renal fibrosis, which suggests that the EphA1-Epha receptor axis may be a novel therapy target for diabetic nephropathy. Mechanistically, the overexpression of EphA1 could inhibit MAPK and the Rho pathway in diabetic kidneys., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Background: Erythropoietin-producing hepatocellular A1 (EPHA1) is the first member of the EPH superfamily. Its abnormal expression has been reported in various cancer types. However, the contribution of EPHA1 to the regulation of colorectal cancer cell behaviour remains unknown., Materials and Methods: In this study, we investigated the expression profile of EPHA1 in human colorectal cancer and its effect on the adhesion and motility of colorectal cancer cells. We used human colorectal cancer specimens and the colorectal adenocarcinoma cell line HRT18 for this purpose., Results: Our cohort screening data showed that in patients with colorectal cancer, low expression of EPHA1 gene is correlated with a remarkably reduced survival. After EPHA1 is knocked-down in colorectal cancer cells using a clustered regularly interspaced short palindromic repeats-associated nuclease 9 (CRISPR-CAS9) genomic editing system, we observed an increase in the spreading and adhesion of HRT18 cells. Moreover, protein array data indicated that the extracellular-regulated kinase (ERK) and c-Jun NH2-terminal kinase (JNK) signaling pathways were activated as a consequence. Inhibition of ERK and JNK proteins with specific inhibitors led to suppression of migration of the colorectal cancer cells., Conclusion: EPHA1 suppresses spreading and adhesion of HRT18 colorectal cancer cells through deactivation of ERK and JNK signaling pathways., (Copyright© 2016 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.)

Background: Erythropoietin-producing hepatocellular (Eph) receptors are the largest subfamily of receptor tyrosine kinases that have been investigated as a possible target for molecular targeted therapy of various cancers., Methods: Patients (n = 222) who underwent gastrectomy for primary gastric cancer were enrolled in this study. Tumor protein expression of EphA1 and EphB6 in surgically resected specimen was investigated using immunohistochemistry. The associations between expression of EphA1 and EphB6 and clinicopathological factors and prognosis were analyzed., Results: High expression of EphA1 was associated with undifferentiated histology (P = 0.002), depth of tumor (P < 0.001), lymph node metastasis (P = 0.001), venous invasion (P = 0.015), stage (P = 0.001), and remote metastasis or recurrence (P < 0.001). In univariate analysis, patients with high expression of EphA1 had significantly poorer overall survival and relapse-free survival compared with patients with low EphA1 expression. The expression level of EphB6 was not associated with any clinicopathological factors and patient survival. Multivariate analysis indicated that depth of tumor [hazard ratio (HR) 9.26, 95 % confidence interval (CI) 0.03-0.46, P = 0.003], lymph node metastasis (HR 9.26, 95 % CI 0.07-0.39, P < 0.001), and high expression of EphA1 (HR 1.86, 95 % CI 0.29-0.99, P = 0.048) are independent prognostic factors for relapse-free survival., Conclusions: EphA1 is a possible target of molecular targeted therapy of gastric cancer.

Ephs and FGFRs belong to a superfamily of receptor tyrosine kinases, playing important roles in stem cell biology. We previously reported that EphA4 and FGFR form a heterodimer following stimulation with ligands, trans-activating each other and signaling through a docking protein, FRS2α, that binds to both receptors. Here, we investigated whether the interaction between EphA4 and FGFRs can be generalized to other Ephs and FGFRs, and, in addition, examined the downstream signal mediating their function in embryonic neural stem/progenitor cells. We revealed that various Ephs and FGFRs interact with each other through similar molecular domains. When neural stem/progenitor cells were stimulated with FGF2 and ephrin-A1, the signal transduced from the EphA4/FGFR/FRS2α complex enhanced self-renewal, while stimulation with ephrin-A1 alone induced neuronal differentiation. The downstream signal required for neuronal differentiation appears to be MAP kinase mainly linked to the Ras family of G proteins. MAP kinase activation was delayed and sustained, distinct from the transient activation induced by FGF2. Interestingly, this effect on neuronal differentiation required the presence of FGFRs. Specific FGFR inhibitor almost completely abolished the function of ephrin-A1 stimulation. These findings suggest that the ternary complex of EphA, FGFR and FRS2α formed by ligand stimulation regulates self-renewal and differentiation of mouse embryonic neural stem/progenitor cells by ligand-specific fine tuning of the downstream signal via FRS2α.

Background: NR4A nuclear receptors are a conserved, functionally diverse group of nuclear receptors that regulate multiple cellular processes including proliferation and differentiation. The gene nhr-6 encodes the sole Caenorhabditis elegans NR4A nuclear receptor homolog with an essential role in reproduction by regulating morphogenesis of the spermatheca, a somatic gonad organ involved in ovulation and fertilization., Results: Here, we identify the spermatheca cell lineage defects that occur in nhr-6 mutants. Utilizing cell marker analysis, we find that nhr-6 is required for cell cycle progression and that the cell proliferation phenotype is not due to premature cell cycle exit. We also show that loss of the negative cell cycle regulators fzr-1 and lin-35 suppresses the cell proliferation defects. We further demonstrate that NHR-6 activity intersects with Eph receptor signaling during spermatheca cell proliferation., Conclusions: NHR-6 has an essential function in promoting cell cycle progression during G1 phase in a specific spermatheca cell lineage. Genetic suppression of the proliferation phenotype does not affect the differentiation phenotypes observed in nhr-6 mutants, indicating a dualistic role for nhr-6 in regulating cell proliferation and cell differentiation during spermatheca organogenesis., (© 2014 Wiley Periodicals, Inc.)

We tested association of nine late-onset Alzheimer's disease (LOAD) risk variants from genome-wide association studies (GWAS) with memory and progression to mild cognitive impairment (MCI) or LOAD (MCI/LOAD) in older Caucasians, cognitively normal at baseline and longitudinally evaluated at Mayo Clinic Rochester and Jacksonville (n>2000). Each variant was tested both individually and collectively using a weighted risk score. APOE-e4 associated with worse baseline memory and increased decline with highly significant overall effect on memory. CLU-rs11136000-G associated with worse baseline memory and incident MCI/LOAD. MS4A6A-rs610932-C associated with increased incident MCI/LOAD and suggestively with lower baseline memory. ABCA7-rs3764650-C and EPHA1-rs11767557-A associated with increased rates of memory decline in subjects with a final diagnosis of MCI/LOAD. PICALM-rs3851179-G had an unexpected protective effect on incident MCI/LOAD. Only APOE-inclusive risk scores associated with worse memory and incident MCI/LOAD. The collective influence of the nine top LOAD GWAS variants on memory decline and progression to MCI/LOAD appears limited. Discovery of biologically functional variants at these loci may uncover stronger effects on memory and incident disease., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Ephrin type-A receptor 1 (EPHA1) (11771145) was documented to be one of the most strongly associated locus with Alzheimer's disease (AD) in a recent meta-analysis of five genome wide association studies. However, its contribution to the pathogenesis of AD remains unclear to date. Here, we addressed the role of EPHA1 in AD by investigating the influence of EPHA1 on cerebrospinal fluid and neuroimaging biomarkers in three clinical stages from the Alzheimer's Disease Neuroimaging Initiative (ADNI) database. We did not detect significant association of EPHA1 with amyloid-β deposition or tau protein. However, the A-allele in the mild cognitive impairment group remarkably prevented hippocampal atrophy (partial correlation coefficient 2.812, 95% CI 0.651 to 4.973) at two-year follow-up. Additionally, AD subjects with the A-allele displayed less atrophy and greater cerebral metabolic rate for glucose (CMRgl) in the right lateral occipitotemporal gyrus (volume: partial correlation coefficient 540.10, 95% CI 247.26 to 832.95; CMRgl: partial correlation coefficient 0.056, 95% CI 0.024 to 0.087) and inferior temporal gyrus (volume: partial correlation coefficient 327.98, 95% CI 11.65 to 644.31; CMRgl: partial correlation coefficient 0.055, 95% CI 0.019 to 0.091) at baseline. This study suggests EPHA1 (rs11771145) interferes with the pathological alteration of the hippocampus and the lateral occipitotemporal and inferior temporal gyri throughout the AD process, leading to a lower risk of AD. However, the limited sample size and follow-up as well as the diversity across ethnicities precluded explanation of these findings.

The formation of straight compartment boundaries separating groups of cells with distinct fates and functions is an evolutionarily conserved strategy during animal development. The physical mechanisms that shape compartment boundaries have recently been further elucidated, however, the molecular mechanisms that underlie compartment boundary formation and maintenance remain poorly understood. Here, we report on the outcome of an RNA interference screen aimed at identifying novel genes involved in maintaining the straight shape of the anteroposterior compartment boundary in Drosophila wing imaginal discs. Out of screening 3114 transgenic RNA interference lines targeting a total of 2863 genes, we identified a single novel candidate that interfered with the formation of a straight anteroposterior compartment boundary. Interestingly, the targeted gene encodes for the Eph receptor tyrosine kinase, an evolutionarily conserved family of signal transducers that has previously been shown to be important for maintaining straight compartment boundaries in vertebrate embryos. Our results identify a hitherto unknown role of the Eph receptor tyrosine kinase in Drosophila and suggest that Eph receptors have important functions in shaping compartment boundaries in both vertebrate and insect development.

Hypoxia regulates the expression of genes that promote tumor growth, angiogenesis and invasion. We previously studied hypoxic tumor cells in vitro and from hepatic metastases of colorectal cancer and determined several potential prognostic factors for hepatocellular carcinoma (HCC). In this study, we evaluated the prognostic impact of the expression of ephrin-A1 (EFNA1) and its receptor, EPHA2, in patients with HCC after curative resection. Samples from a total of 139 HCC patients were analyzed by either microarray alone (n=86) or by microarray and quantitative PCR (n=53). There was no correlation between EFNA1 expression and clinicopathological factors. EPHA2 expression was not significantly correlated with any clinicopathological factors, except for microscopic portal invasion. EFNA1 was an independent prognostic factor for HCC (p=0.0277). These findings suggest that EFNA1 expression may be a useful marker for predicting high risk of recurrence in patients who have undergone curative resection for HCC.

The EPH receptor tyrosine kinases and their cell-bound ligands, the ephrins, have been shown to be associated with cancer development and progression. In this study, mRNA and protein expression of the receptors EPHA1 and EPHA2 as well as of their ligand EFNA1 and their prognostic relevance in clear cell renal cell carcinoma was evaluated. Gene expression was measured in 75 cryo-preserved primary tumors and matched non-malignant renal specimens by quantitative PCR. Protein expression was analyzed by immunohistochemistry on tissue microarrays comprising non-malignant, primary tumors and metastatic renal tissues of 241 patients. Gene and protein expression of all three factors was altered in tumor specimens with EPHA1 and EPHA2 being generally diminished in tumors compared to normal renal tissue, whereas EFNA1 was commonly elevated. A positive EPHA1 and EPHA2 protein staining as well as a low EFNA1 protein level were significantly linked to more aggressive tumor features, but only a positive EPHA1 immunoreactivity was significantly associated with poor survival. In subgroup analyses, EPHA1 and EPHA2 protein levels were significantly higher in metastatic than in primary lesions. Patients with EPHA1/EPHA2-positive tumors or with tumors with positive EPHA1 and low EFNA1 immunoreactivity had the shortest survival rates compared to the respective other combinations. In a multivariate model, EPHA1 was an independent prognostic marker for different survival endpoints. In conclusion, an impaired EPH-ephrin signaling could contribute to the pathogenesis and progression of clear cell renal cell carcinoma.

Background: Down's syndrome (DS) is caused by either complete or partial triplication of chromosome 21, affecting approximately 1/1000 live births, and it is widely accepted that individuals with DS are more likely to develop dementia of Alzheimer's disease (DAD) compared with the general population. Recent collaborative genome-wide association studies of large case control data sets of individuals with and without Alzhemier's disease (AD) have revealed new risk variants for dementia, as well as confirming previously identified risk variants. In this study, nine AD-derived SNPs, near or within the CR1 (rs3818361), BIN1 (rs744373), CD2AP (rs9349407), EPHA1 (rs11767557), CLU (rs1532278), MS4A6A/4A (rs610932), PICALM (rs561655), ABCA7 (rs3764650) and CD33 (rs3865444) genes were genotyped in 295 individuals with DS., Results: There were no significant associations between these nine GWAS-derived SNPs and DAD in British Caucasian individuals with DS. Interestingly the CR1 rs3818361 variant appeared to be associated with mortality in our cohort, particularly in the subjects without dementia. To our knowledge, this is the first time that this variant has been implicated as a determinant of mortality and the finding warrants further investigation in other cohorts with DS., Conclusions: This study shows negative associations of nine AD-derived SNPs with DAD in DS. This may be due to the modest size of our cohort, which may indicate that our study is insufficiently powered to pick up such associations. We cannot conclusively exclude a role for these SNPs in DAD in DS. Clearly, efforts to investigate genetic variants with small effects on disease risk require a much larger cohort of individuals with DS. In fact, we hypothesize that a sample size of 4465 individuals with DS would be needed to determine the role in DAD in DS of the nine AD-derived SNPs investigated in this study. We therefore recommend that all national and international clinics with access to individuals with DS should contribute DNA samples to form DS consortia.

Eph receptor A10 (EphA10) is a valuable breast cancer marker that is highly expressed in breast cancer tissues by comparison with normal breast tissues, as we previously reported. However, the role of EphA10 expression in breast cancer is not well understood. Here, we have analyzed the expression of EphA10 at the mRNA- and protein-level in clinical breast cancer tissues and then evaluated the relationship with clinicopathological parameters for each sample. EphA10 mRNA expression was quantified by real-time polymerase chain reaction using complimentary DNA (cDNA) samples derived from breast cancer patients. Lymph node (LN) metastasis and stage progression were significantly correlated with EphA10 expression at the mRNA level (P = 0.0091 and P = 0.034, respectively). Furthermore, immunohistochemistry (IHC) staining of breast cancer tissue microarrays (TMAs) revealed that EphA10 expression at the protein level was also associated with LN metastasis and stage progression (P = 0.016 and P = 0.011, respectively). These results indicate that EphA10 expression might play a role in tumor progression and metastasis. Our findings will help elucidate the role of EphA10 in clinical breast cancer progression., (© 2013 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.)

The Eph receptors are the largest of the RTK families. Like other RTKs, they transduce signals from the cell exterior to the interior through ligand-induced activation of their kinase domain. However, the Eph receptors also have distinctive features. Instead of binding soluble ligands, they generally mediate contact-dependent cell-cell communication by interacting with surface-associated ligands-the ephrins-on neighboring cells. Eph receptor-ephrin complexes emanate bidirectional signals that affect both receptor- and ephrin-expressing cells. Intriguingly, ephrins can also attenuate signaling by Eph receptors coexpressed in the same cell. Additionally, Eph receptors can modulate cell behavior independently of ephrin binding and kinase activity. The Eph/ephrin system regulates many developmental processes and adult tissue homeostasis. Its abnormal function has been implicated in various diseases, including cancer. Thus, Eph receptors represent promising therapeutic targets. However, more research is needed to better understand the many aspects of their complex biology that remain mysterious.

The erythropoietin-producing hepatocellular (Eph) family of receptor tyrosine kinases regulates a multitude of physiological and pathological processes. EphA1 is the first member of Eph superfamily and is involved in carcinogenesis. The aim of this study was to investigate the expression of EphA1 in prostate cancers cell lines and the tissues, then explore the correlation with the clinicopathologic parameters. The EphA1 transcript expression in prostate cancer cell lines was detected by Quantitative real-time PCR. The expression of EphA1 protein in 138 prostate cancer tissue samples and 21 benign prostate hyperplasia samples were checked by using immunohistochemical staining. EphA1 mRNA was high expressed in LNCap, moderately expressed in 22RV1 and Du145, and lost in PC3. Loss of expression of EphA1 transcript was related to hypermethylation of CpG island around the translation start site. EphA1 protein was differentially expressed in prostate cancers and hyperplasia. Increased expression of EphA1 protein was more frequently detected in prostate cancers than in hyperplasia (P = 0.02), and more often detected in prostate cancer with high Gleason score (P < 0.001). Our data indicate that EphA1 receptor may have roles in carcinogenesis and progression of prostate cancer, and can be a potentially useful target for prognostic and therapeutic application.

Eph receptors play important roles in the development of cancer. The present study aimed to investigate the expression of EphA1 and its clinicopathologic significance in esophageal squamous cell carcinoma (ESCC). The expression levels of the EphA1 transcript and protein were compared between ESCC and matched normal mucosa in the same patient. High expression levels of the EphA1 transcript and protein were detected in 25.6% (21/82) and 23.2% (19/82) of tumors compared with matched normal mucosa. The up-regulation of EphA1 transcript in tumors was positively associated with differentiation (p = 0.002), disease stage (p = 0.034), and lymph node metastasis (p = 0.042). Increased expression of EphA1 protein in tumors was more often observed in patients with well-differentiated tumors (p = 0.004), lymph node metastasis (p = 0.028), and advanced tumor stage (p = 0.008). EphA1 protein expression was detected in intraepithelial neoplasias as well. Decreased expression of EphA1 protein was detected in 65.2% (15/23) of samples with low-grade neoplasia and in 85.3% (8/15) of samples with high-grade intraepithelial neoplasia. Increased staining of EphA1 protein was not detected in low-grade intraepithelial neoplasia samples, but was detected in 21.3% (2/15) of high-grade intraepithelial neoplasia samples. Taken together, our results show that EphA1 appears to be a differentiation marker for esophageal squamous cells, and its increased expression is positively associated with lymph node metastasis and advanced disease stage., (© 2012 The Authors APMIS © 2012 APMIS.)

Recent genome-wide association studies have identified a number of susceptibility loci for Alzheimer disease (AD). To understand the functional consequences and potential interactions of the associated loci, we explored large-scale data sets interrogating the human genome for evidence of positive natural selection. Our findings provide significant evidence for signatures of recent positive selection acting on several haplotypes carrying AD susceptibility alleles; interestingly, the genes found in these selected haplotypes can be assembled, independently, into a molecular complex via a protein-protein interaction (PPI) network approach. These results suggest a possible coevolution of genes encoding physically-interacting proteins that underlie AD susceptibility and are coexpressed in different tissues. In particular, PICALM, BIN1, CD2AP, and EPHA1 are interconnected through multiple interacting proteins and appear to have coordinated evidence of selection in the same human population, suggesting that they may be involved in the execution of a shared molecular function. This observation may be AD-specific, as the 12 loci associated with Parkinson disease do not demonstrate excess evidence of natural selection. The context for selection is probably unrelated to AD itself; it is likely that these genes interact in another context, such as in immune cells, where we observe cis-regulatory effects at several of the selected AD loci., (Copyright © 2012 The American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.)

The Eph receptor tyrosine kinase family includes many members, which are often expressed together in various combinations and can promiscuously interact with multiple ephrin ligands, generating intricate networks of intracellular signals that control physiological and pathological processes. Knowing the entire repertoire of Eph receptors and ephrins expressed in a biological sample is important when studying their biological roles. Moreover, given the correlation between Eph receptor/ephrin expression and cancer pathogenesis, their expression patterns could serve important diagnostic and prognostic purposes. However, profiling Eph receptor and ephrin expression has been challenging. Here we describe a novel and straightforward approach to catalog the Eph receptors present in cultured cells and tissues. By measuring the binding of ephrin Fc fusion proteins to Eph receptors in ELISA and pull-down assays, we determined that a mixture of four ephrins is suitable for isolating both EphA and EphB receptors in a single pull-down. We then used mass spectrometry to identify the Eph receptors present in the pull-downs and estimate their relative levels. This approach was validated in cultured human cancer cell lines, human tumor xenograft tissue grown in mice, and mouse brain tissue. The new mass spectrometry approach we have developed represents a useful tool for the identification of the spectrum of Eph receptors present in a biological sample and could also be extended to profiling ephrin expression.

X-ray irradiation influences metastatic properties of tumor cells and, moreover, metastasis and cellular motility can be modified by members of the Eph receptor/ephrin family of receptor tyrosine kinases. We hypothesized that irradiation-induced changes in cellular properties relevant for metastasis in melanoma cells could be mediated by Eph receptor/ephrin signaling. In this pilot study, we analyzed one pre-metastatic (Mel-Juso) and three metastatic human melanoma (Mel-Juso-L3, A375, and A2058) cells lines and predominantly found anti-metastatic effects of X-ray irradiation with impaired cell growth, clonal growth and motility. Additionally, we observed an irradiation-induced increase in adhesion paralleled by a decrease in migration in Mel-Juso and Mel-Juso-L3 cells and, in part, also in A375 cells. We further demonstrate a decrease of EphA2 both in expression and activity at 7 d after irradiation paralleled by an upregulation of EphA3. Analyzing downstream signaling after irradiation, we detected decreased Src kinase phosphorylation, but unchanged focal adhesion kinase (FAK) phosphorylation, indicating, in part, irradiation-induced downregulation of signaling via the EphA2-Src-FAK axis in melanoma cells. However, to which extent this finding contributes to the modification of metastasis-relevant cellular properties remains to be elucidated.

Receptor tyrosine kinase EphB2 and autophagic machinery are known as tumor suppressors; however, the connection remains to be elucidated. Here, we show the link between EphB2 and autophagy. Sesamin, a major lignan in sesame oil, induced autophagy in the human colon cancer cell lines HT29 and LS180, as shown by electron microscopy, as well as Western blotting and immunofluorescence imaging using an anti-LC3 antibody. Receptor tyrosine kinase array analysis revealed that sesamin treatment increased the levels of unphosphorylated -EphA1 and -EphB2 in HT29 cells. Silencing of EphA1 and EphB2 blocked sesamin-induced autophagy as well as sesamin-induced loss of cell viability. These results show that EphA1 and EphB2 play a critical role in this process. The present study reveals a novel function for EphA1 and EphB2 in the induction of autophagy, suggesting a tumor suppressor role for these proteins in colorectal cancer.

We sought to identify new susceptibility loci for Alzheimer's disease through a staged association study (GERAD+) and by testing suggestive loci reported by the Alzheimer's Disease Genetic Consortium (ADGC) in a companion paper. We undertook a combined analysis of four genome-wide association datasets (stage 1) and identified ten newly associated variants with P ≤ 1 × 10(-5). We tested these variants for association in an independent sample (stage 2). Three SNPs at two loci replicated and showed evidence for association in a further sample (stage 3). Meta-analyses of all data provided compelling evidence that ABCA7 (rs3764650, meta P = 4.5 × 10(-17); including ADGC data, meta P = 5.0 × 10(-21)) and the MS4A gene cluster (rs610932, meta P = 1.8 × 10(-14); including ADGC data, meta P = 1.2 × 10(-16)) are new Alzheimer's disease susceptibility loci. We also found independent evidence for association for three loci reported by the ADGC, which, when combined, showed genome-wide significance: CD2AP (GERAD+, P = 8.0 × 10(-4); including ADGC data, meta P = 8.6 × 10(-9)), CD33 (GERAD+, P = 2.2 × 10(-4); including ADGC data, meta P = 1.6 × 10(-9)) and EPHA1 (GERAD+, P = 3.4 × 10(-4); including ADGC data, meta P = 6.0 × 10(-10)).

The Alzheimer Disease Genetics Consortium (ADGC) performed a genome-wide association study of late-onset Alzheimer disease using a three-stage design consisting of a discovery stage (stage 1) and two replication stages (stages 2 and 3). Both joint analysis and meta-analysis approaches were used. We obtained genome-wide significant results at MS4A4A (rs4938933; stages 1 and 2, meta-analysis P (P(M)) = 1.7 × 10(-9), joint analysis P (P(J)) = 1.7 × 10(-9); stages 1, 2 and 3, P(M) = 8.2 × 10(-12)), CD2AP (rs9349407; stages 1, 2 and 3, P(M) = 8.6 × 10(-9)), EPHA1 (rs11767557; stages 1, 2 and 3, P(M) = 6.0 × 10(-10)) and CD33 (rs3865444; stages 1, 2 and 3, P(M) = 1.6 × 10(-9)). We also replicated previous associations at CR1 (rs6701713; P(M) = 4.6 × 10(-10), P(J) = 5.2 × 10(-11)), CLU (rs1532278; P(M) = 8.3 × 10(-8), P(J) = 1.9 × 10(-8)), BIN1 (rs7561528; P(M) = 4.0 × 10(-14), P(J) = 5.2 × 10(-14)) and PICALM (rs561655; P(M) = 7.0 × 10(-11), P(J) = 1.0 × 10(-10)), but not at EXOC3L2, to late-onset Alzheimer's disease susceptibility.

Objective: To investigate the expression and prognostic significance of EphA2 and EphrinA-1 in ovarian serous carcinomas., Methods: Ninety five tumors from the patients with ovarian serous carcinomas and 2 ovarian cancer cell lines were recruited. The expressions of EphA2 and EphrinA-1 were examined by means of immunohistochemistry. The relationships among protein expression and clinicopathological features, survival of patients were analyzed. The mRNA and protein expressions of EphA2 and EphrinA-1 in ovarian cancer cell lines were measured with semiquantitative polymerase chain reaction and western blotting., Results: The protein expressions of EphA2 and EphrinA-1 in tumor were 92.6% (88/95)and 97.9% (93/95) respectively, while were 40%(8/20) and 30% (6/20) in adjacent ovarian tissue (P = 0.000). EphA2 immunohistochemical staining could be observed in both tumour and vascular endothelial cells, and EphrinA-1 mainly localized in the tumor cells. The expression of EphA2 was significantly associated with the expression of EphrinA-1 (r = 0.98, P = 0.02). There was no significant correlation between the expressions of EphA2/EphrinA-1 and age, FIGO stage, residual tumour size and histological grade. High levels of both EphA2 and EphrinA-1 protein expression were significantly associated with a shorter overall survival in multivariate analysis (P < 0.05). High levels of EphA2 and EphrinA-1 mRNA and protein were detected in OVCAR3 and SKOV3 cell lines., Conclusion: There are over-expression of EphA2 and EphrinA-1 in ovarian serous carcinomas and ovarian cancer cell lines. Tumours with higher expression levels of both EphA2 and EphrinA-1 significantly associated with poorer clinical outcome.

Receptor tyrosine kinases of the Eph subfamily are involved in the development and the carcinogenesis of certain cancers. In this study we investigated expression of EphA1 in gastric carcinomas, and analyzed associations between EphA1 expression and clinicopathological parameters to investigate the role of EphA1 in gastric carcinoma. The level of EphA1 transcript expression in gastric carcinoma tissue and corresponding normal tissue was determined by quantitative real-time reverse transcriptase polymerase chain reaction (RT-PCR). The status of the CpG island associated with the promoter region of EphA1 was assessed by methylation-specific PCR (MSP), and EphA1 protein expression of was examined by immunohistochemical staining (IHC). Down-regulation of EphA1 transcripts was detected in 34% of the cases, up-regulation in 25% of the cases, and no difference in expression in 41% of the cases. The EphA1 transcript expression level was associated with tumor size (P=0.05), stage (P=0.001), and lymph node metastasis (P=0.011). Methylated EphA1 DNA was detected in most of the carcinomas with EphA1 down-regulation that were examined. EphA1 protein expression was associated with depth of wall invasion (P=0.069), stage (P<0.001), and lymph node metastasis (P=0.018). The survival analysis showed that patients whose tumor exhibited EphA1 up-regulation had a poorer outcome than those whose tumor exhibited down-regulation (P=0.005) or no difference in expression (P=0.003). EphA1 may have roles in invasion and metastasis by gastric carcinoma. The EphA1 expression level is a potential prognostic marker in gastric carcinoma, and the EphA1 gene may provide a novel target of therapy for gastric carcinoma.

Eph receptor tyrosine kinases and their ephrin ligands regulate cell navigation during normal and oncogenic development. Signaling of Ephs is initiated in a multistep process leading to the assembly of higher-order signaling clusters that set off bidirectional signaling in interacting cells. However, the structural and mechanistic details of this assembly remained undefined. Here we present high-resolution structures of the complete EphA2 ectodomain and complexes with ephrin-A1 and A5 as the base unit of an Eph cluster. The structures reveal an elongated architecture with novel Eph/Eph interactions, both within and outside of the Eph ligand-binding domain, that suggest the molecular mechanism underlying Eph/ephrin clustering. Structure-function analysis, by using site-directed mutagenesis and cell-based signaling assays, confirms the importance of the identified oligomerization interfaces for Eph clustering.

The Eph family of receptor tyrosine kinases has emerged as one of the pivotal regulators of tumor angiogenesis. EphA1, the first identified member of the Eph receptor family, has been found to be overexpressed in several types of human tumors. A recent report indicated that EphA1 was overexpressed in hepatocellular carcinoma (HCC) and that elevated expression of EphA1 can promote proliferation of HCC cells through stimulation by exogenous Ephrin-A1. To investigate the role of EphA1 in angiogenesis and progression of HCC, we down-regulated EphA1 by RNA interference (RNAi) technology, in an HCC-derived cell line with a high level of EphA1 expression. We established a stable knockdown clone named SiEphA1/Huh-7. The knockdown resulted in decreased proliferation of Huh-7 cells, as well as decreased motility and invasion capability in vitro. siRNA-based EphA1 knockdown also down-regulated the expression of vascular endothelial growth factor (VEGF) and matrix metalloproteinase (MMP)-2 and -9. Interestingly, the suppression of EphA1 expression in Huh-7 cells reduced their outgrowth when inoculated in the subcutaneous space in the flank of nude mice, presumably through angiogenesis inhibition since microvessel density was found to be inhibited.

Aberrant expression of Eph and ephrin proteins has well-established functions in oncogenesis and tumour progression. We describe EphA1 expression in 6 colorectal cancer (CRC) cell lines, 18 controls and 125 CRC specimens. In addition, a well-characterised cohort of 53 paired normal colon and CRCs was also assessed. Expression of EphA1 mRNA was assessed by quantitative real-time PCR and correlated with protein expression by flow cytometry, immunoprecipitation, western blotting and immunohistochemistry. Significant upregulation (2- to 10-fold) of EphA1 was seen in over 50% of cases (P=0.005) whereas many of the remainder showed downregulation of EphA1. Intriguingly, EphA1 over-expression was more prevalent in stage II compared to stage III CRCs (P=0.02). Low EphA1 expression significantly correlated with poor survival (P=0.02). Epigenetic silencing appeared to explain the loss of EphA1 expression as methylation of the EphA1 CpG island strongly correlated with low EphA1 expression (P<0.01). Furthermore, EphA1 re-expression could be induced by treatment with demethylating agents. Our findings identify EphA1 as a potential prognostic marker in CRC. Although therapies targeting high EphA1 expression seem plausible in CRC, the loss of expression in advanced disease suggests a potential risk that targeted therapy, by selecting for loss of expression, might contribute to disease progression.

The molecular mechanisms underlying the homeostatic modulation of presynaptic neurotransmitter release remain largely unknown. In a screen, we isolated mutations in Drosophila ephexin (Rho-type guanine nucleotide exchange factor) that disrupt the homeostatic enhancement of presynaptic release following impairment of postsynaptic glutamate receptor function at the Drosophila neuromuscular junction. We show that Ephexin is sufficient presynaptically for synaptic homeostasis and localizes in puncta throughout the nerve terminal. However, ephexin mutations do not alter other aspects of neuromuscular development, including morphology or active zone number. We then show that, during synaptic homeostasis, Ephexin functions primarily with Cdc42 in a signaling system that converges upon the presynaptic CaV2.1 calcium channel. Finally, we show that Ephexin binds the Drosophila Eph receptor (Eph) and Eph mutants disrupt synaptic homeostasis. Based on these data, we propose that Ephexin/Cdc42 couples synaptic Eph signaling to the modulation of presynaptic CaV2.1 channels during the homeostatic enhancement of presynaptic release.

The Eph-ephrin receptor-ligand system is implicated in cell behavior and morphology. EphA1 is the founding member of the Eph receptors, but little is known about its function. Here, we show that activation of EphA1 kinase inhibits cell spreading and migration in a RhoA-ROCK-dependent manner. We also describe a novel interaction between EphA1 and integrin-linked kinase (ILK), a mediator of interactions between integrin and the actin cytoskeleton. The C-terminal sterile alpha motif (SAM) domain of EphA1 is required and the ankyrin region of ILK is sufficient for the interaction between EphA1 and ILK. The interaction is independent of EphA1 kinase activity but dependent on stimulation of the EphA1 ligand ephrin-A1. Activation of EphA1 kinase resulted in a decrease of ILK activity. Finally, we demonstrated that expression of a kinase-active form of ILK (S343D) rescued the EphA1-mediated spreading defect, and attenuated RhoA activation. These results suggest that EphA1 regulates cell morphology and motility through the ILK-RhoA-ROCK pathway.

The Eph gene family has important roles in the developmental processes and may also be involved in the initiation, progression, and metastasis of certain types of cancers. In the present study, quantitative real-time reverse-transcriptase PCR was performed to detect the expression of EphA1 transcript in 5 colon cancer cell lines and 75 colorectal carcinomas. Immunohistochemical staining was used to check the expression of EphA1 protein in 20 colorectal adenomas and in 111 colorectal carcinomas specimens. EphA1 protein expression was not completely consistent with transcript expression. EphA1 protein was expressed in all adenomas and reduced in 54% colorectal cancers. Reduced expression of EphA1 protein occurred more often in male patients (P=0.028) and in patients with poor differentiation (P=0.027), greater depth of wall invasion (P=0.003), lymph node metastasis (P=0.034), and advanced tumor stage (P=0.003). Patients with reduced EphA1 expression had a poor overall survival (P=0.059). Reduced EphA1 expression in patients over 55 years or with rectal cancers and sigmoid colon cancers is associated with a poor overall survival (P=0.034 and 0.015, respectively). Our data indicate that the EphA1 may play different roles during the different stages of colorectal carcinoma progression.odern Pathology (2009) 22, 151-160; doi:10.1038/modpathol.2008.188; published online 14 November 2008.

Eph receptor tyrosine kinases (RTKs) are a highly conserved family of signaling proteins with functions in cellular migration, adhesion, apoptosis, and proliferation during both adult and embryonic life. Here, we describe a knock-in mouse in which EphA1 expression is disrupted via the insertion of an internal ribosome entry site (IRES)-human placental alkaline phosphatase (ALPP) reporter cassette into exon II of the EphA1 gene. This was shown to successfully knockout expression of endogenous EphA1 and enforce expression of the ALPP reporter by the EphA1 promoter. Staining for the ALPP reporter protein demonstrated an epithelially restricted expression pattern in mouse tissues. In EphA1 null mice, two separate phenotypes were identified: abnormal tail development manifesting as a kinky tail was found in approximately 80% of homozygous adults. A second, distinct abnormality present in approximately 18% of females was characterized by imperforate uterovaginal development with hydrometrocolpos and caused by a resistance of cells to apoptosis during reproductive tract canalization. These results indicate a possible role for EphA1 in tissue patterning and hormone-induced apoptotic processes., (Copyright 2008 Wiley-Liss, Inc.)