IGFBP4

Gene Summary

Gene:IGFBP4; insulin like growth factor binding protein 4
Aliases: BP-4, IBP4, IGFBP-4, HT29-IGFBP
Location:17q21.2
Summary:This gene is a member of the insulin-like growth factor binding protein (IGFBP) family and encodes a protein with an IGFBP domain and a thyroglobulin type-I domain. The protein binds both insulin-like growth factors (IGFs) I and II and circulates in the plasma in both glycosylated and non-glycosylated forms. Binding of this protein prolongs the half-life of the IGFs and alters their interaction with cell surface receptors. [provided by RefSeq, Jul 2008]
Databases:OMIM, HGNC, Ensembl, GeneCard, Gene
Protein:insulin-like growth factor-binding protein 4
Source:NCBIAccessed: 30 August, 2019

Ontology:

What does this gene/protein do?
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Pathways:What pathways are this gene/protein implicaed in?
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Cancer Overview

Research Indicators

Publications Per Year (1994-2019)
Graph generated 31 August 2019 using data from PubMed using criteria.

Literature Analysis

Mouse over the terms for more detail; many indicate links which you can click for dedicated pages about the topic.

  • Immunohistochemistry
  • Ovarian Cancer
  • Messenger RNA
  • Antineoplastic Agents
  • Cell Movement
  • Cell Proliferation
  • Somatomedins
  • Insulin-Like Growth Factor Binding Protein 4
  • Neoplasm Proteins
  • Disease Progression
  • Epithelial-Mesenchymal Transition
  • Chromosome 17
  • Cancer Gene Expression Regulation
  • Cancer RNA
  • Infant
  • Apoptosis
  • Nucleic Acid Amplification Techniques
  • Bone Cancer
  • Breast Cancer
  • Signal Transduction
  • Chromosome Mapping
  • Gene Expression
  • Lung Cancer
  • Small Cell Lung Cancer
  • beta Catenin
  • Drug Resistance
  • Estrogen Receptors
  • Gene Expression Profiling
  • Valine
  • Gene Expression Regulation
  • Tissue Array Analysis
  • Staging
  • Tamoxifen
  • Estrogens
  • RTPCR
  • Transcription
  • Transcription Factors
  • Neoplastic Cell Transformation
  • Treatment Failure
  • Oligonucleotide Array Sequence Analysis
  • Fulvestrant
Tag cloud generated 30 August, 2019 using data from PubMed, MeSH and CancerIndex

Specific Cancers (5)

Data table showing topics related to specific cancers and associated disorders. Scope includes mutations and abnormal protein expression.

Note: list is not exhaustive. Number of papers are based on searches of PubMed (click on topic title for arbitrary criteria used).

Latest Publications: IGFBP4 (cancer-related)

Wang J, Luo XX, Tang YL, et al.
The prognostic values of insulin-like growth factor binding protein in breast cancer.
Medicine (Baltimore). 2019; 98(19):e15561 [PubMed] Free Access to Full Article Related Publications
Insulin-like growth factor binding proteins (IGFBPs) are a family of proteins binding to insulin-like growth factors, generally consisting 6 high-affinity IGFBPs, namely IGFBP1 through IGFBP6. IGFBP family members have been indicated to be involved in the development and progression of tumors and may be useful prognostic biomarkers in various malignancies. However, the prognostic role of individual IGFBPs, especially at the mRNA level in breast cancer patients remains elusive.We accessed the prognostic roles of IGFBPs family (IGFBP1-6) in breast cancer through the "Kaplan-Meier plotter" online database and OncoLnc database.Our results showed that the high expression of IGFBP1 mRNA was associated with favorable relapsed free survival (RFS) in all breast cancer patients. The high expression of IGFBP2 mRNA was associated with favorable overall survival (OS) and RFS in all breast cancer patients. The high expression of IGFBP3 mRNA was significantly correlated to worsen RFS in all breast cancer patients. The high expression of IGFBP4 mRNA was associated with favorable OS, RFS, distant metastasis-free survival, and post-progression survival in all breast cancer patients.Our results indicated that expression of IGFBPs mRNA may have prognostic values in breast cancer patients, and have a benefit for developing tools to predict the prognosis more accurately.

Choi HJ, Joo HS, Won HY, et al.
Role of RBP2-Induced ER and IGF1R-ErbB Signaling in Tamoxifen Resistance in Breast Cancer.
J Natl Cancer Inst. 2018; 110(4) [PubMed] Related Publications
Background: Despite the benefit of endocrine therapy, acquired resistance during or after treatment still remains a major challenge in estrogen receptor (ER)-positive breast cancer. We investigated the potential role of histone demethylase retinoblastoma-binding protein 2 (RBP2) in endocrine therapy resistance of breast cancer.
Methods: Survival of breast cancer patients according to RBP2 expression was analyzed in three different breast cancer cohorts including METABRIC (n = 1980) and KM plotter (n = 1764). RBP2-mediated tamoxifen resistance was confirmed by invitro sulforhodamine B (SRB) colorimetric, colony-forming assays, and invivo xenograft models (n = 8 per group). RNA-seq analysis and receptor tyrosine kinase assay were performed to identify the tamoxifen resistance mechanism by RBP2. All statistical tests were two-sided.
Results: RBP2 was associated with poor prognosis to tamoxifen therapy in ER-positive breast cancer (P = .04 in HYU cohort, P = .02 in KM plotter, P = .007 in METABRIC, log-rank test). Furthermore, RBP2 expression was elevated in patients with tamoxifen-resistant breast cancer (P = .04, chi-square test). Knockdown of RBP2 conferred tamoxifen sensitivity, whereas overexpression of RBP2 induced tamoxifen resistance invitro and invivo (MCF7 xenograft: tamoxifen-treated control, mean [SD] tumor volume = 70.8 [27.9] mm3, vs tamoxifen-treated RBP2, mean [SD] tumor volume = 387.9 [85.1] mm3, P < .001). Mechanistically, RBP2 cooperated with ER co-activators and corepressors and regulated several tamoxifen resistance-associated genes, including NRIP1, CCND1, and IGFBP4 and IGFBP5. Furthermore, epigenetic silencing of IGFBP4/5 by RBP2-ER-NRIP1-HDAC1 complex led to insulin-like growth factor-1 receptor (IGF1R) activation. RBP2 also increased IGF1R-ErbB crosstalk and subsequent PI3K-AKT activation via demethylase activity-independent ErbB protein stabilization. Combinational treatment with tamoxifen and PI3K inhibitor could overcome RBP2-mediated tamoxifen resistance (RBP2-overexpressing cells: % cell viability [SD], tamoxifen = 89.0 [3.8]%, vs tamoxifen with BKM120 = 41.3 [5.6]%, P < .001).
Conclusions: RBP2 activates ER-IGF1R-ErbB signaling cascade in multiple ways to induce tamoxifen resistance, suggesting that RBP2 is a potential therapeutic target for ER-driven cancer.

Yang B, Zhang L, Cao Y, et al.
Overexpression of lncRNA IGFBP4-1 reprograms energy metabolism to promote lung cancer progression.
Mol Cancer. 2017; 16(1):154 [PubMed] Free Access to Full Article Related Publications
BACKGROUND: Reprogrammed energy metabolism as an emerging hallmark of cancer has recently drawn special attention since it facilitate cell growth and proliferation. Recently, long noncoding RNAs (lncRNAs) have been served as key regulators implicated in tumor development and progression by promoting proliferation, invasion and metastasis. However, the associations of lncRNAs with cellular energy metabolism in lung cancer (LC) need to be clarified.
METHODS: Here, we conducted bioinformatics analysis and found insulin-like growth factor binding protein 4-1 (IGFBP4-1) as a new candidate lncRNA located in the upstream region of IGFBP4 gene. The expression levels of lnc-IGFBP4-1, mRNA levels of IGFBP4 in 159 paired lung cancer samples and adjacent, histological normal tissues by qRT-PCR. Over-expression and RNA interference (RNAi) approaches were adopted to investigate the biological functions of lnc-IGFBP4-1. The intracellular ATP level was measured using the Cell Titer-Glo Luminescent Cell Viability Assay kit, and changes in metabolic enzymes were examined in cancer cells and normal pulmonary epithelial cells with qRT-PCR.
RESULTS: Our results showed that lnc-IGFBP4-1 was significantly up-regulated in LC tissues compared with corresponding non-tumor tissues (P < 0.01), and its expression level was significantly correlated with TNM stage (P < 0.01) and lymph node metastasis (P < 0.05). Further investigation showed that overexpression of lnc-IGFBP4-1 significantly promoted LC cell proliferation in vitro and in vivo, while downregulation of endogenous lnc-IGFBP4-1 could inhibited cell proliferation and induce apoptosis. Moreover, we found lnc-IGFBP4-1 could influences ATP production levels and expression of enzymes including HK2, PDK1 and LDHA, in addition, decline in both ATP production and these enzymes in response to 2-DG and 2-DG-combined Rho123, respectively, was observed in lnc-IGFBP4-1-overespressing LC cells, indicative of an enhanced aerobic glycolysis rate. Finally, lnc-IGFBP4-1 was observed to negatively correlate with gene IGFBP4, and lower expression level of IGFPB4 was found after lnc-IGFBP4-1-overexpression was transfected into PC9 cells, higher expression level of IGFPB4 was also found after lnc-IGFBP4-1-downregulation was transfected into GLC-82 cells, which indicates that IGFBP4 may exert its targeting function regulated by lnc-IGFBP4-1.
CONCLUSIONS: Taken together, these findings provide the first evidence that lnc-IGFBP4-1 is significantly up-regulated in LC tissues and plays a positive role in cell proliferation and metastasis through possible mechanism of reprogramming tumor cell energy metabolism, which suggests that lnc-IGFBP4-1 may be a promising biomarker in LC development and progression and as a potential therapeutic target for LC intervention.

Lee KW, Lee SS, Hwang JE, et al.
Development and Validation of a Six-Gene Recurrence Risk Score Assay for Gastric Cancer.
Clin Cancer Res. 2016; 22(24):6228-6235 [PubMed] Free Access to Full Article Related Publications
PURPOSE: This study was aimed at developing and validating a quantitative multigene assay for predicting tumor recurrence after gastric cancer surgery.
EXPERIMENTAL DESIGN: Gene expression data were generated from tumor tissues of patients who underwent surgery for gastric cancer (n = 267, training cohort). Genes whose expression was significantly associated with activation of YAP1 (a frequently activated oncogene in gastrointestinal cancer), 5-year recurrence-free survival, and 5-year overall survival were first identified as candidates for prognostic genes (156 genes, P < 0.001). We developed the recurrence risk score (RRS) by using quantitative RT-PCR to identify genes whose expression levels were significantly associated with YAP1 activation and patient survival in the training cohort.
RESULTS: We based the RRS assay on 6 genes, IGFBP4, SFRP4, SPOCK1, SULF1, THBS, and GADD45B, whose expression levels were significantly associated with YAP1 activation and prognosis in the training cohort. The RRS assay was further validated in an independent cohort of 317 patients. In multivariate analysis, the RRS was an independent predictor of recurrence [HR, 1.6; 95% confidence interval (CI), 1.02-2.4; P = 0.03]. In patients with stage II disease, the RRS had an HR of 2.9 (95% CI, 1.1-7.9; P = 0.03) and was the only significant independent predictor of recurrence.
CONCLUSIONS: The RRS assay was a valid predictor of recurrence in the two cohorts of patients with gastric cancer. Independent prospective studies to assess the clinical utility of this assay are warranted. Clin Cancer Res; 22(24); 6228-35. ©2016 AACR.

Yamaoka T, Ohmori T, Ohba M, et al.
Acquired Resistance Mechanisms to Combination Met-TKI/EGFR-TKI Exposure in Met-Amplified EGFR-TKI-Resistant Lung Adenocarcinoma Harboring an Activating EGFR Mutation.
Mol Cancer Ther. 2016; 15(12):3040-3054 [PubMed] Related Publications
Met-amplified EGFR-tyrosine kinase inhibitor (TKI)-resistant non-small cell lung cancer (NSCLC) harboring an activating EGFR mutation is responsive to concurrent EGFR-TKI and Met-TKI treatment in a preclinical model. Here, we determined that Met-amplified gefitinib-resistant cells acquire dual resistance to inhibition of EGFR and Met tyrosine kinase activities. PC-9 lung adenocarcinoma cells harboring 15-bp deletions (Del E746_A750) in EGFR exon 19 were treated with increasing concentrations of the Met-TKI PHA665752 and 1 μmol/L gefitinib for 1 year; three resistant clones were established via Met amplification. The three dual-resistance cell lines (PC-9DR2, PC-9DR4, and PC-9DR6, designated as DR2, DR4, and DR6, respectively) exhibited different mechanisms for evading both EGFR and Met inhibition. None of the clones harbored a secondary mutation of EGFR T790M or a Met mutation. Insulin-like growth factor (IGF)/IGF1 receptor activation in DR2 and DR4 cells acted as a bypass signaling pathway. Met expression was attenuated to a greater extent in DR2 than in PC-9 cells, but was maintained in DR4 cells by overexpression of IGF-binding protein 3. In DR6 cells, Met was further amplified by association with HSP90, which protected Met from degradation and induced SET and MYND domain-containing 3 (SMYD3)-mediated Met transcription. This is the first report describing the acquisition of dual resistance mechanisms in NSCLC harboring an activating EGFR mutation to Met-TKI and EGFR-TKI following previous EGFR-TKI treatment. These results might inform the development of more effective therapeutic strategies for NSCLC treatment. Mol Cancer Ther; 15(12); 3040-54. ©2016 AACR.

Minchenko DO, Kharkova AP, Halkin OV, et al.
Effect of hypoxia on the expression of genes encoding insulin-like growth factors and some related proteins in U87 glioma cells without IRE1 function.
Endocr Regul. 2016; 50(2):43-54 [PubMed] Related Publications
OBJECTIVE: The aim of the present study was to investigate the effect of hypoxia on the expression of genes encoding insulin-like growth factors (IGF1 and IGF2), their receptor (IGF1R), binding protein-4 (IGFBP4), and stanniocalcin 2 (STC2) in U87 glioma cells in relation to inhibition of endoplasmic reticulum stress signaling mediated by IRE1 (inositol requiring enzyme 1) for evaluation of their possible significance in the control of tumor growth.
METHODS: The expression of IGF1, IGF2, IGF1R, IGFBP4, and STC2 genes in U87 glioma cells transfected by empty vector pcDNA3.1 (control) and cells without IRE1 signaling enzyme function (transfected by dnIRE1) upon hypoxia was studied by qPCR.
RESULTS: The expression of IGF1 and IGF2 genes is down-regulated in glioma cells without IRE1 signaling enzyme function in comparison with the control cells. At the same time, the expression of IGF1R, IGFBP4, and STC2 genes was up-regulated in glioma cells upon inhibition of IRE1, with more significant changes for IGFBP4 and STC2 genes. We also showed that hypoxia does not change significantly the expression of IGF1, IGF2, and IGF1R genes but up-regulated IGFBP4 and STC2 genes expression in control glioma cells. Moreover, the inhibition of both enzymatic activities (kinase and endoribonuclease) of IRE1 in glioma cells does not change significantly the effect of hypoxia on the expression of IGF1, IGF1R, and IGFBP4 genes but introduces sensitivity of IGF2 gene to hypoxic condition. Thus, the expression of IGF2 gene is resistant to hypoxia only in control glioma cells and significantly down-regulated in cells without functional activity of IRE1 signaling enzyme, which is central mediator of the unfolded protein response and an important component of the tumor growth as well as metabolic diseases.
CONCLUSIONS: Results of this study demonstrate that the expression of IGF1 and IGF1R genes is resistant to hypoxic condition both in control U87 glioma cells and cells without IRE1 signaling enzyme function. However, hypoxia significantly up-regulates the expression of IGFBP4 gene independently on the inhibition of IRE1 enzyme. These data show that proteins encoded by these genes are resistant to hypoxia except IGFBP4 and participate in the regulation of metabolic and proliferative processes through IRE1 signaling.

Onstenk W, Sieuwerts AM, Mostert B, et al.
Molecular characteristics of circulating tumor cells resemble the liver metastasis more closely than the primary tumor in metastatic colorectal cancer.
Oncotarget. 2016; 7(37):59058-59069 [PubMed] Free Access to Full Article Related Publications
BACKGROUND: CTCs are a promising alternative for metastatic tissue biopsies for use in precision medicine approaches. We investigated to what extent the molecular characteristics of circulating tumor cells (CTCs) resemble the liver metastasis and/or the primary tumor from patients with metastatic colorectal cancer (mCRC).
RESULTS: The CTC profiles were concordant with the liver metastasis in 17/23 patients (74%) and with the primary tumor in 13 patients (57%). The CTCs better resembled the liver metastasis in 13 patients (57%), and the primary tumor in five patients (22%). The strength of the correlations was not associated with clinical parameters. Nine genes (CDH1, CDH17, CDX1, CEACAM5, FABP1, FCGBP, IGFBP3, IGFBP4, and MAPT) displayed significant differential expressions, all of which were downregulated, in CTCs compared to the tissues in the 23 patients.
PATIENTS AND METHODS: Patients were retrospectively selected from a prospective study. Using the CellSearch System, CTCs were enumerated and isolated just prior to liver metastasectomy. A panel of 25 CTC-specific genes was measured by RT-qPCR in matching CTCs, primary tumors, and liver metastases. Spearman correlation coefficients were calculated and considered as continuous variables with r=1 representing absolute concordance and r=-1 representing absolute discordance. A cut-off of r>0.1 was applied in order to consider profiles to be concordant.
CONCLUSIONS: In the majority of the patients, CTCs reflected the molecular characteristics of metastatic cells better than the primary tumors. Genes involved in cell adhesion and epithelial-to-mesenchymal transition were downregulated in the CTCs. Our results support the use of CTC characterization as a liquid biopsy for precision medicine.

Roque DR, Makowski L, Chen TH, et al.
Association between differential gene expression and body mass index among endometrial cancers from The Cancer Genome Atlas Project.
Gynecol Oncol. 2016; 142(2):317-22 [PubMed] Free Access to Full Article Related Publications
OBJECTIVE: The Cancer Genome Atlas (TCGA) identified four integrated clusters for endometrial cancer (EC): POLE, MSI, CNL and CNH. We evaluated differences in gene expression profiles of obese and non-obese women with EC and examined the association of body mass index (BMI) within the clusters identified in TCGA.
METHODS: TCGA RNAseq data was used to identify genes related to increasing BMI among ECs. The POLE, MSI and CNL clusters were composed mostly of endometrioid EC. Patient BMI was compared between these three clusters with one-way ANOVA. Association between gene expression and BMI was also assessed while adjusting for confounding effects of potential confounding factors. p-Values testing the association between gene expression and BMI were adjusted for multiple hypothesis testing over the 20,531 genes considered.
RESULTS: Mean BMI was statistically different between the ECs in the CNL (35.8) versus POLE (29.8) cluster (p=0.006) and approached significance for the MSI (33.0) versus CNL (35.8) cluster (p=0.05). 181 genes were significantly up- or down-regulated with increasing BMI in endometrioid EC (q-value<0.01), including LPL, IRS-1, IGFBP4, IGFBP7 and the progesterone receptor. DAVID functional annotation analysis revealed significant enrichment in "cell cycle" (adjusted p-value=1.5E-5) and "DNA metabolic processes" (adjusted p-value=1E-3) for the identified genes.
CONCLUSIONS: Obesity related genes were found to be upregulated with increasing BMI among endometrioid ECs. Patients with POLE tumors have the lowest median BMI when compared to MSI and CNL. Given the heterogeneity among endometrioid EC, consideration should be given to abandoning the Type I and II classification of EC tumors.

West DC, Pan D, Tonsing-Carter EY, et al.
GR and ER Coactivation Alters the Expression of Differentiation Genes and Associates with Improved ER+ Breast Cancer Outcome.
Mol Cancer Res. 2016; 14(8):707-19 [PubMed] Free Access to Full Article Related Publications
UNLABELLED: In estrogen receptor (ER)-negative breast cancer, high tumor glucocorticoid receptor (GR) expression has been associated with a relatively poor outcome. In contrast, using a meta-analysis of several genomic datasets, here we find that tumor GR mRNA expression is associated with improved ER(+) relapse-free survival (RFS; independently of progesterone receptor expression). To understand the mechanism by which GR expression is associated with a better ER(+) breast cancer outcome, the global effect of GR-mediated transcriptional activation in ER(+) breast cancer cells was studied. Analysis of GR chromatin immunoprecipitation followed by high-throughput sequencing in ER(+)/GR(+) MCF-7 cells revealed that upon coactivation of GR and ER, GR chromatin association became enriched at proximal promoter regions. Furthermore, following ER activation, increased GR chromatin association was observed at ER, FOXO, and AP1 response elements. In addition, ER associated with GR response elements, suggesting that ER and GR interact in a complex. Coactivation of GR and ER resulted in increased expression (relative to ER activation alone) of transcripts that encode proteins promoting cellular differentiation (e.g., KDM4B, VDR) and inhibiting the Wnt signaling pathway (IGFBP4). Finally, expression of these individual prodifferentiation genes was associated with significantly improved RFS in ER(+) breast cancer patients. Together, these data suggest that the coexpression and subsequent activity of tumor cell GR and ER contribute to the less aggressive natural history of early-stage breast cancer by coordinating the altered expression of genes favoring differentiation.
IMPLICATIONS: The interaction between ER and GR activity highlights the importance of context-dependent nuclear receptor function in cancer. Mol Cancer Res; 14(8); 707-19. ©2016 AACR.

Ricci F, Fratelli M, Guffanti F, et al.
Patient-derived ovarian cancer xenografts re-growing after a cisplatinum treatment are less responsive to a second drug re-challenge: a new experimental setting to study response to therapy.
Oncotarget. 2017; 8(5):7441-7451 [PubMed] Free Access to Full Article Related Publications
Even if ovarian cancer patients are very responsive to a cisplatinum-based therapy, most will relapse with a resistant disease. New experimental animal models are needed to explore the mechanisms of resistance, to better tailor treatment and improve patient prognosis. To address these aims, seven patient-derived high-grade serous/endometrioid ovarian cancer xenografts were characterized for the antitumor response after one and two cycles of cisplatinum and classified as Very Responsive, Responsive, and Low Responsive to drug treatment. Xenografts re-growing after the first drug cycle were much less responsive to the second one. The expression of epithelial-mesenchymal transition (EMT) and cancer stem cells (CSCs) genes was investigated in cisplatinum-treated and not-treated tumors. We found that different EMT (TCF3, CAMK2N1, EGFR, and IGFBP4) and CSCs (SMO, DLL1, STAT3, and ITGA6) genes were expressed at higher levels in Low Responsive than in Responsive and Very Responsive xenografts. The expression of STAT3 was found to be associated with lower survival (HR = 13.7; p = 0.013) in the TCGA patient data set. MMP9, CD44, DLL4, FOXP1, MERTK, and PTPRC genes were found more expressed in tumors re-growing after cisplatinum treatment than in untreated tumors. We here describe a new in vivo ovarian carcinoma experimental setting that will be instrumental for specific trials of combination therapy to counteract cisplatinum resistance in order to improve the prognosis of ovarian patients.

Prithviraj P, Anaka M, McKeown SJ, et al.
Pregnancy associated plasma protein-A links pregnancy and melanoma progression by promoting cellular migration and invasion.
Oncotarget. 2015; 6(18):15953-65 [PubMed] Free Access to Full Article Related Publications
Melanoma is the most common cancer diagnosed in pregnant women and an aggressive course with poorer outcomes is commonly described during pregnancy or shortly after childbirth. The underlying mechanisms for this are not understood. Here, we report that melanoma migration, invasiveness and progression are promoted by Pregnancy-Associated Plasma Protein-A (PAPPA), a pregnancy-associated metalloproteinase produced by the placenta that increases the bioavailability of IGF1 by cleaving it from a circulating complex formed with IGFBP4. We show that PAPPA is widely expressed by metastatic melanoma tumors and is elevated in melanoma cells exhibiting mesenchymal, invasive and label-retaining phenotypes. Notably, inhibition of PAPPA significantly reduced invasion and migration of melanoma cells in vitro and in vivo within the embryonic chicken neural tube. PAPPA-enriched pregnancy serum treatment enhanced melanoma motility in vitro. Furthermore, we report that IGF1 can induce the phenotypic and functional effects of epithelial-to-mesenchymal transition (EMT) in melanoma cells. In this study, we establish a clear relationship between a pregnancy-associated protein PAPPA, melanoma and functional effects mediated through IGF1 that provides a plausible mechanism for accelerated melanoma progression during pregnancy. This opens the possibility of targeting the PAPPA/IGF1 axis therapeutically.

Meng G, Li Y, Lv Y, et al.
Preliminary screening of differentially expressed genes involved in methyl-CpG-binding protein 2 gene-mediated proliferation in human osteosarcoma cells.
Tumour Biol. 2015; 36(4):3009-15 [PubMed] Related Publications
Methyl-CpG-binding protein 2 (MeCP2) is essential in human brain development and has been linked to several cancer types and neuro-developmental disorders. This study aims to screen the MeCP2 related differentially expressed genes and discover the therapeutic targets for osteosarcoma. CCK8 assay was used to detect the proliferation and SaOS2 and U2OS cells. Apoptosis of cells was detected by flow cytometry analysis that monitored Annexin V-APC/7-DD binding and 7-ADD uptake simultaneously. Denaturing formaldehyde agarose gel electrophoresis was employed to examine the quality of total RNA 18S and 28S units. Gene chip technique was utilized to discover the differentially expressed genes correlated with MeCP2 gene. Differential gene screening criteria were used to screen the changed genes. The gene up-regulation or down-regulation more than 1.5 times was regarded as significant differential expression genes. The CCK8 results indicated that the cell proliferation of MeCP2 silencing cells (LV-MeCP2-RNAi) was significantly decreased compared to non-silenced cells (LV-MeCP2-RNAi-CN) (P < 0.05). MeCP2 silencing could also induce significant apoptosis compared to non-silenced cells (P < 0.05); 107 expression changed genes were screened from a total of 49,395 transcripts. Among the total 107 transcripts, 34 transcripts were up-regulated and 73 transcripts were down-regulated. There were five significant differentially expressed genes, including IGFBP4, HOXC8, LMO4, MDK, and CTGF, which correlated with the MeCP2 gene. The methylation frequency of CpG in IGFBP4 gene could achieve 55%. In conclusion, the differentially expressed IGFBP4, HOXC8, LMO4, MDK, and CTGF genes may be involved in MeCP2 gene-mediated proliferation and apoptosis in osteosarcoma cells.

Ueda J, Ho JC, Lee KL, et al.
The hypoxia-inducible epigenetic regulators Jmjd1a and G9a provide a mechanistic link between angiogenesis and tumor growth.
Mol Cell Biol. 2014; 34(19):3702-20 [PubMed] Free Access to Full Article Related Publications
Hypoxia promotes stem cell maintenance and tumor progression, but it remains unclear how it regulates long-term adaptation toward these processes. We reveal a striking downregulation of the hypoxia-inducible histone H3 lysine 9 (H3K9) demethylase JMJD1A as a hallmark of clinical human germ cell-derived tumors, such as seminomas, yolk sac tumors, and embryonal carcinomas. Jmjd1a was not essential for stem cell self-renewal but played a crucial role as a tumor suppressor in opposition to the hypoxia-regulated oncogenic H3K9 methyltransferase G9a. Importantly, loss of Jmjd1a resulted in increased tumor growth, whereas loss of G9a produced smaller tumors. Pharmacological inhibition of G9a also resulted in attenuation of tumor growth, offering a novel therapeutic strategy for germ cell-derived tumors. Finally, Jmjd1a and G9a drive mutually opposing expression of the antiangiogenic factor genes Robo4, Igfbp4, Notch4, and Tfpi accompanied by changes in H3K9 methylation status. Thus, we demonstrate a novel mechanistic link whereby hypoxia-regulated epigenetic changes are instrumental for the control of tumor growth through coordinated dysregulation of antiangiogenic gene expression.

Praveen Kumar VR, Sehgal P, Thota B, et al.
Insulin like growth factor binding protein 4 promotes GBM progression and regulates key factors involved in EMT and invasion.
J Neurooncol. 2014; 116(3):455-64 [PubMed] Related Publications
Insulin like growth factor binding protein 4 (IGFBP4) regulates growth and development of tissues and organs by negatively regulating IGF signaling. Among most cancers, IGFBP4 has growth inhibitory role and reported as a down-regulated gene, except for renal cell carcinoma, wherein IGFBP4 promotes tumor progression. IGFBP4 expression has been shown to be higher in increasing grades of astrocytoma. However, the functional role of IGFBP4 in gliomas has not been explored. Surgical biopsies of 20 normal brain and 198 astrocytoma samples were analyzed for IGFBP4 expression by qRT-PCR. Highest expression of IGFBP4 mRNA was seen in GBM tumors compared to control brain tissues (median log2 of 2.035, p < 0.0001). Immunohistochemical analysis of 53 tissue samples revealed predominant nuclear staining of IGFBP4, seen maximally in GBMs when compared to DA and AA tumors (median LI = 29.12 ± 16.943, p < 0.001). Over expression of IGFBP4 in U343 glioma cells resulted in up-regulation of molecules involved in tumor growth, EMT and invasion such as pAkt, pErk, Vimentin, and N-cadherin and down-regulation of E-cadherin. Functionally, IGFBP4 over expression in these cells resulted in increased proliferation, migration and invasion as assessed by MTT, transwell migration, and Matrigel invasion assays. These findings were confirmed upon IGFBP4 knockdown in U251 glioma cells. Our data suggest a pro-tumorigenic role for IGFBP4 in glioma.

Feierabend D, Walter J, Grube S, et al.
Methylation-specific multiplex ligation-dependent probe amplification and its impact on clinical findings in medulloblastoma.
J Neurooncol. 2014; 116(2):213-20 [PubMed] Related Publications
Gain of (proto-)oncogenes and loss or promoter hypermethylation of tumor suppressor genes (TSGs) play essential roles in tumorigenesis. Methylation-specific multiplex ligation-dependent probe amplification (MS-MLPA) allows simultaneous detection of both these alterations. MS-MLPA was performed on 20 medulloblastoma samples (n = 12 cryoconserved; n = 8 formalin-fixed paraffin-embedded, FFPE) in order to screen for copy number changes in 77 unselected TSGs and (proto-)oncogenes as well as for promoter hypermethylation in a subset of 33 TSGs. In all specimens, determination of promoter methylation status was possible, whereas robust data concerning copy number changes could be obtained on cryopreserved material only. We found a median of 1.5 deletions and 6.5 amplifications in the 12 cryopreserved medulloblastoma and a median of 5 promoter hypermethylation per tumor. Frequent copy number changes included amplification of ASC on 16p12 (5/12) and amplification of several adjacent genes on 17q (3/12) including IGFBP4. Hypermethylation of MSH6 on 2p16 was found in 16 samples. MS-MLPA findings were also correlated with clinical and histological characteristics. The number of promoter hypermethylation was significantly associated with presence of necrosis (p = 0.004). Tumors which recurred within 1 year were more likely to show amplification of the GATA5 gene (p = 0.038), while hypermethylation of CASP8 was associated with a lower tumor recurrence rate (p = 0.036). There was also a trend towards a correlation between total number of aberrations and CSF dissemination (p = 0.055). Our findings confirm frequent presence of certain aberrations and reveal novel candidates for improving prognosis based on genetic and epigenetic tumor features. A medulloblastoma-specific MS-MLPA probe set seems a potentially valuable tool for further investigations on larger sample series.

Yeh TS, Wang F, Chen TC, et al.
Expression profile of microRNA-200 family in hepatocellular carcinoma with bile duct tumor thrombus.
Ann Surg. 2014; 259(2):346-54 [PubMed] Related Publications
OBJECTIVE: The aim of this study was to assess the role of the miR-200 family in the pathogenesis of hepatocellular carcinoma with bile duct tumor thrombus (HCC-BDTT).
BACKGROUND: Hepatocellular carcinoma with bile duct tumor thrombus is a challenging condition because of its rarity and dismal prognosis. Epithelial-to-mesenchymal transition (EMT) is considered a critical step in the progression and metastasis of HCC and is regulated by the microRNA-200 (miR-200) family.
METHODS: Thirty patients with HCC-BDTT were enrolled and 1240 patients with conventional HCC (cHCC) served as clinicopathologic controls. Sixty age- and sex-matched cHCC patients were selected to compare the miR-200 family expression profile and immunohistochemical characteristics. Gain- and loss-of-function studies of the miR-200 family were conducted using the hepatoma cell lines.
RESULTS: Although the mean size of HCC-BDTT was smaller than that of cHCC, the former had a higher incidence of vascular invasion and a poorer long-term survival. The expressions of miR-200c and miR-141 were downregulated in HCC-BDTT (4.5- and 4.8-fold decrease, respectively). Downregulation of both miR-200c and miR-141 independently predicted disease-free survival. The HCC-BDTT, but not cHCC, exhibited overexpression of ZEB1, Twist, transforming growth factor-β receptor type II, and vimentin, and aberrant E-cadherin expression, indicating EMT. The HCC-BDTT demonstrated increased expression in IL-6 and stemness factor Bmi1, but reduced level of metastasis-suppressive protein, insulin-like growth factor-binding protein 4. The invasive ability of the highly aggressive Mahlavu cell was attenuated by pre-miR-200c+141, whereas the invasive ability of the less aggressive Huh7 cell was enhanced by anti-miR-200c+141.
CONCLUSIONS: Simultaneous silencing of miR-200c and miR-141 was likely to be responsible for the development of HCC-BDTT via ZEB1-directed EMT activation and Sec23a-mediated secretome.

Huang J, Tabata S, Kakiuchi S, et al.
Identification of pregnancy-associated plasma protein A as a migration-promoting gene in malignant pleural mesothelioma cells: a potential therapeutic target.
Oncotarget. 2013; 4(8):1172-84 [PubMed] Free Access to Full Article Related Publications
Despite recent advances in treatment, malignant pleural mesothelioma (MPM) remains a deadly disease. Targeted therapy generated broad interests and is highly expected for the treatment of MPM, yet promising preclinical results have not been translated into substantial clinical benefits for the patients. In this study, we tried to identify the genes which play functional roles in cell migration as well as to test whether they can be used as novel targets for molecular targeted therapy for MPM in preclinical model. In our study, pregnancy-associated plasma protein A (PAPPA) was identified as a gene whose expression level is correlated with MPM cell migration by correlation analysis combining MPM cell migration ability and their gene expression profiles. Highly migratory cells were selected from MPM cell lines, MSTO-211H, NCI-H290 and EHMES-1 in vitro and up-regulation of PAPPA in these cells were confirmed. In vitro, PAPPA was demonstrated to stimulate the MPM cell migration via cleavage of insulin-like growth factor-binding protein-4 and subsequent release of IGF-1. Gene silencing of PAPPA in MPM cells led to reduced migration, invasion and proliferation. Furthermore, PAPPA shRNA transfected NCI-H290 when orthotopically inoculated into pleural cavity of severe combined immunodeficiency recipient mice, failed to develop tumors and produce bloody pleural effusion as control shRNA transfected cells did. Our study suggests that PAPPA plays a functional role in promoting MPM cell migration and it might serve as a potential therapeutic target for the treatment of MPM.

Fellenberg J, Sähr H, Liu L, et al.
Rescue of silenced UCHL1 and IGFBP4 expression suppresses clonogenicity of giant cell tumor-derived stromal cells.
Cancer Lett. 2013; 336(1):61-7 [PubMed] Related Publications
Giant cell tumor (GCT) of bone is a generally benign tumor with a locally aggressive behavior. Histologically, GCTs consist of multinucleated giant cells, mononuclear histiocytes and the neoplastic fibroblast-like stromal cells (GCTSC). Growing evidence exists that GCTSCs develop from mesenchymal stem cells (MSCs), but little is known about the underlying molecular mechanisms. In previous studies we observed inactivation of the ubiquitin carboxyl-terminal hydrolase L1 (UCHL1) gene in primary GCTSC due to strong DNA hypermethylation, indicating that epigenetic silencing might be involved in neoplastic transformation of MSCs. Here we investigated further candidate genes and identified strong hypermethylation of the insulin-like growth factor binding protein 4 (IGFBP4) promoter, resulting in IGFBP4 downregulation in GCTs compared to MSCs. Overexpression of UCHL1 and IGFBP4 by stable transfection of GCTSC did not influence cell viability, proliferation, migration and chemosensitivity compared to parental cells. However, colony-formation was significantly decreased suggesting that rescue of UCHL1 and IFGBP4 suppresses clonogenicity of GCT stromal cells. The observation of reduced expression of the stem-cell-specific transcription factors OCT4 and SOX2 in these cell lines further supported our findings. Epigenetic silencing of UCHL1 and IGFBP4 in GCTs might thus be a crucial event during the malignant transformation of MSCs in the context of GCT development and represent promising targets for the development of new diagnostic and therapeutic strategies.

Matise LA, Palmer TD, Ashby WJ, et al.
Lack of transforming growth factor-β signaling promotes collective cancer cell invasion through tumor-stromal crosstalk.
Breast Cancer Res. 2012; 14(4):R98 [PubMed] Free Access to Full Article Related Publications
INTRODUCTION: Transforming growth factor beta (TGF-β) has a dual role during tumor progression, initially as a suppressor and then as a promoter. Epithelial TGF-β signaling regulates fibroblast recruitment and activation. Concurrently, TGF-β signaling in stromal fibroblasts suppresses tumorigenesis in adjacent epithelia, while its ablation potentiates tumor formation. Much is known about the contribution of TGF-β signaling to tumorigenesis, yet the role of TGF-β in epithelial-stromal migration during tumor progression is poorly understood. We hypothesize that TGF-β is a critical regulator of tumor-stromal interactions that promote mammary tumor cell migration and invasion.
METHODS: Fluorescently labeled murine mammary carcinoma cells, isolated from either MMTV-PyVmT transforming growth factor-beta receptor II knockout (TβRII KO) or TβRIIfl/fl control mice, were combined with mammary fibroblasts and xenografted onto the chicken embryo chorioallantoic membrane. These combinatorial xenografts were used as a model to study epithelial-stromal crosstalk. Intravital imaging of migration was monitored ex ovo, and metastasis was investigated in ovo. Epithelial RNA from in ovo tumors was isolated by laser capture microdissection and analyzed to identify gene expression changes in response to TGF-β signaling loss.
RESULTS: Intravital microscopy of xenografts revealed that mammary fibroblasts promoted two migratory phenotypes dependent on epithelial TGF-β signaling: single cell/strand migration or collective migration. At epithelial-stromal boundaries, single cell/strand migration of TβRIIfl/fl carcinoma cells was characterized by expression of α-smooth muscle actin and vimentin, while collective migration of TβRII KO carcinoma cells was identified by E-cadherin+/p120+/β-catenin+ clusters. TβRII KO tumors also exhibited a twofold greater metastasis than TβRIIfl/fl tumors, attributed to enhanced extravasation ability. In TβRII KO tumor epithelium compared with TβRIIfl/fl epithelium, Igfbp4 and Tspan13 expression was upregulated while Col1α2, Bmp7, Gng11, Vcan, Tmeff1, and Dsc2 expression was downregulated. Immunoblotting and quantitative PCR analyses on cultured cells validated these targets and correlated Tmeff1 expression with disease progression of TGF-β-insensitive mammary cancer.
CONCLUSION: Fibroblast-stimulated carcinoma cells utilize TGF-β signaling to drive single cell/strand migration but migrate collectively in the absence of TGF-β signaling. These migration patterns involve the signaling regulation of several epithelial-to-mesenchymal transition pathways. Our findings concerning TGF-β signaling in epithelial-stromal interactions are important in identifying migratory mechanisms that can be targeted as recourse for breast cancer treatment.

Becker MA, Hou X, Harrington SC, et al.
IGFBP ratio confers resistance to IGF targeting and correlates with increased invasion and poor outcome in breast tumors.
Clin Cancer Res. 2012; 18(6):1808-17 [PubMed] Free Access to Full Article Related Publications
PURPOSE: To improve the significance of insulin-like growth factor-binding protein 5 (IGFBP-5) as a prognostic and potentially predictive marker in patients with breast cancer.
EXPERIMENTAL DESIGN: Increased IGFBP-5 expression was identified in MCF-7 cells resistant (MCF-7R4) to the IGF-1R/insulin receptor (InsR) inhibitor BMS-536924 and its role examined by targeted knockdown and overexpression in multiple experimental models. Protein expression of IGFBP-5 was measured by immunohistochemistry in a cohort of 76 patients with breast cancer to examine correlative associations with invasive tumor fraction and outcome. The use of a combined IGFBP-5/IGFBP-4 (BPR) expression ratio was applied to predict anti-IGF-1R/InsR response in a panel of breast cancer lines and outcome in multiple breast tumor cohorts.
RESULTS: IGFBP-5 knockdown decreased BMS-536924 resistance in MCF-7R4 cells, whereas IGFBP-5 overexpression in MCF-7 cells conferred resistance. When compared with pathologically normal reduction mammoplasty tissue, IGFBP-5 expression levels were upregulated in both invasive and histologically normal adjacent breast cancer tissue. In both univariate and multivariate modeling, metastasis-free survival, recurrence free survival (RFS), and overall survival (OS) were significantly associated with high IGFBP-5 expression. Prognostic power of IGFBP-5 was further increased with the addition of IGFBP-4 where tumors were ranked based upon IGFBP-5/IGFBP-4 expression ratio (BPR). Multiple breast cancer cohorts confirm that BPR (high vs. low) was a strong predictor of RFS and OS.
CONCLUSION: IGFBP-5 expression is a marker of poor outcome in patients with breast cancer. An IGFBP-5/IGFBP-4 expression ratio may serve as a surrogate biomarker of IGF pathway activation and predict sensitivity to anti-IGF-1R targeting.

Korpal M, Ell BJ, Buffa FM, et al.
Direct targeting of Sec23a by miR-200s influences cancer cell secretome and promotes metastatic colonization.
Nat Med. 2011; 17(9):1101-8 [PubMed] Free Access to Full Article Related Publications
Although the role of miR-200s in regulating E-cadherin expression and epithelial-to-mesenchymal transition is well established, their influence on metastatic colonization remains controversial. Here we have used clinical and experimental models of breast cancer metastasis to discover a pro-metastatic role of miR-200s that goes beyond their regulation of E-cadherin and epithelial phenotype. Overexpression of miR-200s is associated with increased risk of metastasis in breast cancer and promotes metastatic colonization in mouse models, phenotypes that cannot be recapitulated by E-cadherin expression alone. Genomic and proteomic analyses revealed global shifts in gene expression upon miR-200 overexpression toward that of highly metastatic cells. miR-200s promote metastatic colonization partly through direct targeting of Sec23a, which mediates secretion of metastasis-suppressive proteins, including Igfbp4 and Tinagl1, as validated by functional and clinical correlation studies. Overall, these findings suggest a pleiotropic role of miR-200s in promoting metastatic colonization by influencing E-cadherin-dependent epithelial traits and Sec23a-mediated tumor cell secretome.

Rossi E, Klersy C, Manca R, et al.
Correlation between genomic alterations assessed by array comparative genomic hybridization, prognostically informative histologic subtype, stage, and patient survival in gastric cancer.
Hum Pathol. 2011; 42(12):1937-45 [PubMed] Related Publications
It is difficult to evaluate the prognostic value of histologic criteria in gastric cancer because of the high variability of morphologic patterns. Recently, histologic subtypes of low, intermediate, or high malignant potential have been identified, providing the basis for a prognostically informative grading system. Because array comparative genomic hybridization systems allow systematic analysis of chromosome alterations, which may be prognostically and pathogenetically informative, we applied high-resolution genome-wide array comparative genomic hybridization to archival material from 81 gastric cancer cases followed for a median of 150 months after surgery. The DNA extracted from paraffin sections gave useful results in 49 tumors, 18 of which were of low-grade, 24 of intermediate, and 7 of high-grade histotypes. Based on the number of chromosome aberrations and the presence/absence of amplifications, 3 tumor clusters of increasing genomic lesion severity were constructed, which proved to correlate significantly with histologic grade and stage as well as with patient survival. Further investigation documented the lower number and severity of genomic alterations in tumors with microsatellite DNA instability and high CD8-rich lymphoid response; the close association of 8p23.1 amplification with cardial cancer; the frequent amplification of genes involved in cell renewal (CDC6, HER2, GRB7, IGFBP4) at 17q12-q21.1, with close histochemical correlation with HER2 membranous expression; and more sporadic amplification of chromosome regions harboring important oncogenes like MYC, KRAS, NRAS, CRKL, CCNE1, or ZNF217. We conclude that genome-wide array comparative genomic hybridization of gastric cancer contributes prognostically relevant information providing a genetic background for histologic grading.

Wang YC, Yu SQ, Wang XH, et al.
Differences in phenotype and gene expression of prostate stromal cells from patients of varying ages and their influence on tumour formation by prostate epithelial cells.
Asian J Androl. 2011; 13(5):732-41 [PubMed] Free Access to Full Article Related Publications
Prostate cancer (PCa) is an age-related disease, and the stromal microenvironment plays an important role in prostatic malignant progression. However, the differences in prostate stromal cells present in young and old tissue are still obscure. We established primary cultured stromal cells from normal prostatic peripheral zone (PZ) of donors of varying ages and found that cultured stromal cells from old donors (PZ-old) were more enlarged and polygonal than those from young donors (PZ-young). Furthermore, based on immunocytochemical and ultrastructural analysis, the components of stromal cells changed from a majority of fibroblasts to a mixture of fibroblasts and myofibroblasts with increasing donor age. Using a three-dimensional in vitro culture system, we found that PZ-old stromal cells could enhance the proliferation, migration and invasion of cocultured benign BPH-1 and PC-3 cells. Using an in vivo tissue recombination system, we also found that PZ-old stromal cells are more effective than PZ-young cells in promoting tumour formation by BPH-1 cells of high passage (>100) and PC-3 cells. To probe the possible mechanism of these effects, we performed cDNA microarray analysis and profiled 509 upregulated genes and 188 downregulated genes in PZ-old cells. Among the changed genes, we found genes coding for a subset of paracrine factors that are capable of influencing adjacent epithelial cells; these include hepatocyte growth factor (HGF), fibroblast growth factor 5 (FGF5), insulin-like growth factor 2 (IGF2), insulin-like growth factor-binding protein 4 (IGFBP4), IGFBP5 and matrix metallopeptidase 1 (MMP1). Changes in the expression of these genes were further confirmed by quantitative real-time polymerase chain reaction (PCR), Western blotting and enzyme-linked immunosorbent assays. Overall, our findings indicate that stromal cells from prostate PZ of old donors are more active than similar cells from young donors in promoting the malignant process of adjacent epithelial cells. This finding hints at a new potential strategy for the prevention of PCa.

Luo F, Poulogiannis G, Ye H, et al.
Synergism between K-rasVal12 and mutant Apc accelerates murine large intestinal tumourigenesis.
Oncol Rep. 2011; 26(1):125-33 [PubMed] Related Publications
K-ras (KRAS) is mutated in 40-50% of human colorectal adenomas and carcinomas and plays key roles in cell proliferation, apoptosis, motility and differentiation, but its functional contribution to intestinal tumourigenesis in vivo remains incompletely understood. We have previously crossed K-rasVal12 transgenic mice with Ah-Cre mice to produce K-rasVal12/Cre offspring that inducibly express K-rasVal12 4A and 4B in the intestines, but this alone showed no significant effect on intestinal adenoma formation. Here, we crossed these mice with Min mice to evaluate the effect of K-rasVal12 and Apc mutation on intestinal tumourigenesis in vivo. The double mutant K-rasVal12/Cre/ApcMin/+ mice showed a moderate (1.86-fold) increase in adenomas in the small intestines, but a striking acceleration (6-fold increase) of large intestinal adenoma formation (P<0.01) and significantly reduced survival (by ~5 weeks) compared with control ApcMin/+ mice (P<0.01). There was recombination of the mutant K-rasVal12 transgene in 80% of large intestinal adenomas with expression of both K-rasVal12 4A and 4B isoform transcripts and expression of K-RasVal12 protein. The large intestinal adenomas showed immunohistochemical evidence of activation of MapK, Akt and Wnt signaling pathways and this was confirmed by quantitative RT-PCR analysis of relative transcript expression levels of target genes using a panel of 23 selected genes evaluated in both adenomas and non-tumour-bearing intestines. Several genes including Tiam1, Gastrin, CD44, uPA, Igfbp4, VEGF and Cox-2 that are known to be transcriptionally regulated by activation of the Wnt signaling pathway were found to be expressed at higher levels in the large intestinal adenomas from K-rasVal12/Cre/ApcMin/+ mice compared with those from controls, although other Wnt signaling pathway target genes remained unchanged. These data show that intestinal expression of K-rasVal12 accelerates Apc-initiated intestinal adenomagenesis in vivo with particularly striking tumour promotion in the large intestines and indicate synergistic effects between mutant K-ras and mutant Apc in this process.

Romero D, O'Neill C, Terzic A, et al.
Endoglin regulates cancer-stromal cell interactions in prostate tumors.
Cancer Res. 2011; 71(10):3482-93 [PubMed] Free Access to Full Article Related Publications
Endoglin is an accessory receptor for TGF-β that has been implicated in prostate cancer cell detachment, migration, and invasiveness. However, the pathophysiologic significance of endoglin with respect to prostate tumorigenesis has yet to be fully established. In this study, we addressed this question by investigation of endoglin-dependent prostate cancer progression in a TRAMP (transgenic adenocarcinoma mouse prostate) mouse model where endoglin was genetically deleted. In this model, endoglin was haploinsufficient such that its allelic deletion slightly increased the frequency of tumorigenesis, yet produced smaller, less vascularized, and less metastatic tumors than TRAMP control tumors. Most strikingly, TRAMP:eng(+/-)-derived tumors lacked the pronounced infiltration of carcinoma-associated fibroblasts (CAF) that characterize TRAMP prostate tumors. Studies in human primary prostate-derived stromal cells (PrSC) confirmed that suppressing endoglin expression decreased cell proliferation, the ability to recruit endothelial cells, and the ability to migrate in response to tumor cell-conditioned medium. We found increased levels of secreted insulin-like growth factor-binding proteins (IGFBP) in the conditioned medium from endoglin-deficient PrSCs and that endoglin-dependent regulation of IGFBP-4 secretion was crucial for stromal cell-conditioned media to stimulate prostate tumor cell growth. Together, our results firmly establish the pathophysiologic involvement of endoglin in prostate cancer progression; furthermore, they show how endoglin acts to support the viability of tumor-infiltrating CAFs in the tumor microenvironment to promote neovascularization and growth.

Lamy PJ, Fina F, Bascoul-Mollevi C, et al.
Quantification and clinical relevance of gene amplification at chromosome 17q12-q21 in human epidermal growth factor receptor 2-amplified breast cancers.
Breast Cancer Res. 2011; 13(1):R15 [PubMed] Free Access to Full Article Related Publications
INTRODUCTION: Human epidermal growth factor receptor 2 (HER2)-amplified breast cancers represent a tumor subtype with chromosome 17q rearrangements that lead to frequent gene amplifications. The aim of this study was to quantify the amplification of genes located on chromosome 17q and to analyze the relations between the pattern of gene amplifications and the patients' characteristics and survival.
METHODS: Patients with HER2-positive breast tumors (HER2 score of 3+ by immunohistochemistry or positive for HER2 amplification by fluorescence in situ hybridization (FISH)) (n = 86) and with HER2-negative breast tumors (n = 40) (negative controls) were included in this study. Using a quantitative polymerase chain reaction method and DNA extracted from frozen tumor specimens, 11 genes (MED1, STARD3, HER2, GRB7, THRA, RARA, TOP2A, IGFBP4, CCR7, KRT20, KRT19 and GAS), which are localized within Chr17q12-q21 and have a putative role in breast cancer development, were quantified. Relapse-free and overall survival rates were estimated from the date of surgery to the date of the event of interest (recurrence or death) using the Kaplan-Meier method.
RESULTS: Gene amplification was observed only in HER2-positive tumors, and the frequency of amplification decreased with the distance of the gene from HER2. HER2 presented the highest level of amplification. TOP2A was not included in the smallest region of amplification involving HER2. Amplification of RARA, KRT20 and KRT19 was significantly associated with node-positive breast cancer (P = 0.030, P = 0.002 and P = 0.033, respectively). During a median follow-up period of 55 months (range, 6 to 81 months), the subgroup of patients with hormone receptor-negative cancer and without TOP2A amplification showed the worst survival (relapse-free survival: hazard ratio (HR) = 0.29, 95% confidence interval (95% CI), 0.13 to 0.65, P = 0.001; and overall survival: HR = 0.28, 95% CI, 0.10 to 0.76, P = 0.008).
CONCLUSIONS: HER2 amplification seems to drive genomic instability along chromosome 17q, leading to different patterns of gene amplification. This study confirms the clinical importance of identifying, among patients with HER2-positive breast tumors, the subgroup of patients with hormone receptor-negative and nonamplified TOP2A cancers as they have the worst prognosis.

Ueno K, Hirata H, Majid S, et al.
IGFBP-4 activates the Wnt/beta-catenin signaling pathway and induces M-CAM expression in human renal cell carcinoma.
Int J Cancer. 2011; 129(10):2360-9 [PubMed] Related Publications
The Wnt/β-catenin signaling pathway is inactivated by Wnt antagonists in most cancers and IGFBP-4 is an antagonist of the Wnt/ β-catenin signaling pathway. However, the function of IGFBP-4 is not currently understood in renal cell carcinoma (RCC). We initially found that the expression of IGFBP-4 was significantly lower in primary RCC and higher in metastatic RCC compared to normal human kidney tissues. To assess the function of IGFBP4, we established IGFBP4 transfectants (primary renal cancer cell line) and performed functional analyses including Tcf reporter assays, cell viability, invasive capability, mortality, and in vivo tumor growth. Interestingly IGFBP-4 transfectants promoted cell growth (in vitro and in vivo), invasion, and motility in primary renal cancer. Tcf transcriptional activity was significantly increased in IGFBP-4 transfectants compared to mock cells and β-catenin expression was increased. Also the β-catenin downstream effector, MT1-MMP showed increased expression in IGFBP4 transfectants. Additionally IGFBP4 induced the expression of M-CAM, a marker of tumor progression. In order to assess the role of IGFBP4 in metastatic renal cancer, IGFBP-4 mRNA in a metastatic renal cancer cell lines (ACHN) was knocked-down using a siRNA technique. The cell growth and motility was decreased in si-IGFBP4 transfected ACHN cells compared to cells transfected with control siRNA. Tcf activity in ACHN cells was also decreased with si-IGFBP-4 transfection. This is a first report documenting that IGFBP-4 expression in RCC activates cell growth, metastasis, Wnt/beta-catenin signaling and may be involved in RCC metastasis.

Sato H, Sakaeda M, Ishii J, et al.
Insulin-like growth factor binding protein-4 gene silencing in lung adenocarcinomas.
Pathol Int. 2011; 61(1):19-27 [PubMed] Related Publications
Gene silencing by promoter hypermethylation plays an important role in molecular pathogenesis. We previously reported that insulin-like growth factor (IGF) binding protein-4 (IGFBP-4), which inhibits IGF-dependent growth, is expressed via early growth response-1 (EGR-1) and is often silenced in cultivated lung cancer cells. The purpose of the present study was to clarify clinicopathological factors associated with IGFBP-4 gene silencing in lung adenocarcinomas. Seventy-six surgically resected adenocarcinomas (20 well-, 35 moderately-, and 21 poorly-differentiated) were subjected to methylation-specific polymerase chain reaction (PCR) analysis for EGR-1-binding sites located in the IGFBP-4 promoter and immunohistochemistry for IGFBP-4, EGR-1, and Ki-67. Thirty-two adenocarcinomas (42%) revealed IGFBP-4 promoter hypermethylation, and the severity inversely correlated with the level of IGFBP-4 expression (P < 0.0001) and tumor differentiation (well versus poor, P = 0.0278; well/moderate versus poor, P = 0.0395). Furthermore, there was a negative correlation between Ki-67 labeling index and IGFBP-4 expression (P = 0.0361). These findings suggest that the expression of IGFBP-4 in adenocarcinoma cells in vivo is downregulated by epigenetic silencing in association with tumor differentiation, resulting in disruption of the mechanism of IGFBP-4-mediated growth inhibition.

Zhou GD, Richardson M, Fazili IS, et al.
Role of retinoic acid in the modulation of benzo(a)pyrene-DNA adducts in human hepatoma cells: implications for cancer prevention.
Toxicol Appl Pharmacol. 2010; 249(3):224-30 [PubMed] Free Access to Full Article Related Publications
Carcinogen-DNA adducts could lead to mutations in critical genes, eventually resulting in cancer. Many studies have shown that retinoic acid (RA) plays an important role in inducing cell apoptosis. Here we have tested the hypothesis that levels of carcinogen-DNA adducts can be diminished by DNA repair and/or by eliminating damaged cells through apoptosis. Our results showed that the levels of total DNA adducts in HepG2 cells treated with benzo(a)pyrene (BP, 2 μM)+RA (1 μM) were significantly reduced compared to those treated with BP only (P=0.038). In order to understand the mechanism of attenuation of DNA adducts, further experiments were performed. Cells were treated with BP (4 μM) for 24h to initiate DNA adduct formation, following which the medium containing BP was removed, and fresh medium containing 1 μM RA was added. The cells were harvested 24h after RA treatment. Interestingly, the levels of total DNA adducts were lower in the BP/RA group (390 ± 34) than those in the BP/DMSO group (544 ± 33), P=0.032. Analysis of cell apoptosis showed an increase in BP+RA group, compared to BP or RA only groups. Our results also indicated that attenuation of BP-DNA adducts by RA was not primarily due to its effects on CYP1A1 expression. In conclusion, our results suggest a mechanistic link between cellular apoptosis and DNA adduct formation, phenomena that play important roles in BP-mediated carcinogenesis. Furthermore, these results help understand the mechanisms of carcinogenesis, especially in relation to the chemopreventive properties of nutritional apoptosis inducers.

Xie HL, Li ZY, Gan RL, et al.
Differential gene and protein expression in primary gastric carcinomas and their lymph node metastases as revealed by combined cDNA microarray and tissue microarray analysis.
J Dig Dis. 2010; 11(3):167-75 [PubMed] Related Publications
OBJECTIVE: To gain insight into the molecular events of lymph node metastasis of human gastric carcinoma.
METHODS: The gene expression profile of five matched primary gastric carcinomas and their lymph node metastases was analyzed by complementary DNA (cDNA) microarray. Differential genes were identified in the metastatic and corresponding primary tumor pairs. Among the differentially expressed genes, carbonic anhydrase II (CAII) and insulin-like growth factor binding protein 4 (IGFBP 4) genes were detected by RT-PCR. CTTN protein expression was examined by tissue microarray.
RESULTS: There was a high expression (over twofold) of 44 genes and a low expression (under twofold) of 32 genes in lymph node metastasis compared with primary gastric carcinoma, respectively. CAII mRNA was downregulated and IGFBP 4 mRNA was upregulated in paired lymph node metastases of gastric carcinomas. The overexpression of CTTN protein was related to the lymph node metastasis and the clinical stage of gastric carcinomas.
CONCLUSION: This study showed that there is a low expression of genes relative to growth signal and immune response in lymph node metastases, and a high expression of genes relative to growth factor, cell cycle, cell motility and adhesion in lymph node metastases compared with primary gastric carcinomas. The expression of CTTN was related to the invasion and metastasis of gastric cancer.

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