Research IndicatorsGraph generated 26 August 2015 using data from PubMed using criteria.
Mouse over the terms for more detail; many indicate links which you can click for dedicated pages about the topic. Tag cloud generated 26 August, 2015 using data from PubMed, MeSH and CancerIndex
Specific Cancers (3)
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).
OMIM, Johns Hopkin University
Referenced article focusing on the relationship between phenotype and genotype.
Cancer Genome Anatomy Project, NCI
COSMIC, Sanger Institute
Somatic mutation information and related details
Search the Epigenomics database and view relevant gene tracks of samples.
Latest Publications: TGFB3 (cancer-related)
Lam S, Wiercinska E, Teunisse AF, et al.Wild-type p53 inhibits pro-invasive properties of TGF-β3 in breast cancer, in part through regulation of EPHB2, a new TGF-β target gene.
Breast Cancer Res Treat. 2014; 148(1):7-18 [PubMed
] Related Publications
The p53 tumor suppressor protein is primarily known for its important role in tumor suppression. In addition, p53 affects tumor cell migration, invasion, and epithelial-mesenchymal transition (EMT); processes also regulated by the transforming growth factor-β (TGF-β) signaling pathway. Here, we investigated the role of p53 in breast tumor cell invasion, migration, and EMT and examined the interplay of p53 with TGF-β3 in these processes. MCF-10A1 and MCF-10CA1a breast cancer cells were treated with Nutlin-3 and TGF-β3, and the effects on tumor cell migration and invasion were studied in transwell and 3D spheroid invasion assays. The effects of Nutlin-3 and TGF-β3 on EMT were examined in NMuMG cells. To identify genes involved in TGF-β-induced invasion that are modulated by p53, a Human Tumor Metastasis-specific RT-PCR array was performed. Verification of EPHB2 regulation by TGF-β3 and p53 was performed on breast cancer tumor cell lines. We demonstrate that p53 inhibits basal and TGF-β3-induced invasion, migration, and EMT in normal breast epithelial and breast cancer cells. Pharmacological activation of p53 inhibited induction of several TGF-β3 targets involved in TGF-β3-induced tumor cell invasion, i.e., matrix metallo proteinase (MMP)2, MMP9, and integrin β 3 . The ephrin-type B receptor 2 (EPHB2) gene was identified as a new TGF-β target important for TGF-β3-mediated invasion and migration, whose transcriptional activation by TGF-β3 is also inhibited by p53. The results show an intricate interplay between p53 and TGF-β3 whereby p53 inhibits the TGF-β3-induced expression of genes, e.g., EPHB2, to impede tumor cell invasion and migration.
Oktem G, Sercan O, Guven U, et al.Cancer stem cell differentiation: TGFβ1 and versican may trigger molecules for the organization of tumor spheroids.
Oncol Rep. 2014; 32(2):641-9 [PubMed
] Related Publications
Cancer stem cells (CSCs) have the ability to self-renew similar to normal stem cells. This process is linked with metastasis and resistance to chemotherapy and radiotherapy. In the present study, we constructed an in vitro differentiation model for CSCs. CSCs isolated and proliferated for one passage were maintained as monolayers or spheroid-forming cells with serum included media for differentiation process. Differentiation of adhesion molecules and cellular ultrastructural properties were investigated and compared in both monolayer and spheroid cultures. CD133+/CD44+ cancer-initiating cells were isolated from DU-145 human prostate cancer cell line monolayer cultures and propagated as tumor spheroids and compared with the remaining heterogeneous cancer cell bulk population. Microarray-based gene expression analysis was applied to determine genes with differential expression and protein expression levels of candidates were analyzed by immunohistochemistry. Electron microscopy showed detailed analysis of morphology. TGFβ1 was found to be significantly upregulated in monolayer CSCs. High expression levels of VCAN, COL7A1, ITGβ3, MMP16, RPL13A, COL4A2 and TIMP1 and low expression levels of THBS1, MMP1 and MMP14 were detected when CSCs were maintained as serum-grown prostate CSC spheroids. Immunohistochemistry supported increased immunoreactivity of TGFβ1 in monolayer cultures and VCAN in spheroids. CSCs were found to possess multipotential differentiation capabilities through upregulation and/or downregulation of their markers. TGFβ1 is a triggering molecule, it stimulates versican, Col7A1, ITGβ3 and, most importantly, the upregulation of versican was only detected in CSCs. Our data support a model where CSCs must be engaged by one or more signaling cascades to differentiate and initiate tumor formation. This mechanism occurs with intracellular and extracellular signals and it is possible that CSCc themselves may be a source for extracellular signaling. These molecules functioning in tumor progression and differentiation may help develop targeted therapy.
BACKGROUND: Transforming Growth Factor beta (TGF-β) acts as a tumor suppressor early in carcinogenesis but turns into tumor promoter in later disease stages. In fact, TGF-β is a known inducer of integrin expression by tumor cells which contributes to cancer metastatic spread and TGF-β inhibition has been shown to attenuate metastasis in mouse models. However, carcinoma cells often become refractory to TGF-β-mediated growth inhibition. Therefore identifying patients that may benefit from anti-TGF-β therapy requires careful selection.
METHODS: We performed in vitro analysis of the effects of exposure to TGF-β in NSCLC cell chemotaxis and adhesion to lymphatic endothelial cells. We also studied in an orthotopic model of NSCLC the incidence of metastases to the lymph nodes after inhibition of TGF-β signaling, β3 integrin expression or both.
RESULTS: We offer evidences of increased β3-integrin dependent NSCLC adhesion to lymphatic endothelium after TGF-β exposure. In vivo experiments show that targeting of TGF-β and β3 integrin significantly reduces the incidence of lymph node metastasis. Even more, blockade of β3 integrin expression in tumors that did not respond to TGF-β inhibition severely impaired the ability of the tumor to metastasize towards the lymph nodes.
CONCLUSION: These findings suggest that lung cancer tumors refractory to TGF-β monotherapy can be effectively treated using dual therapy that combines the inhibition of tumor cell adhesion to lymphatic vessels with stromal TGF-β inhibition.
Levy G, Malik M, Britten J, et al.Liarozole inhibits transforming growth factor-β3--mediated extracellular matrix formation in human three-dimensional leiomyoma cultures.
Fertil Steril. 2014; 102(1):272-281.e2 [PubMed
] Free Access to Full Article Related Publications
OBJECTIVE: To investigate the impact of liarozole on transforming growth factor-β3 (TGF-β3) expression, TGF-β3 controlled profibrotic cytokines, and extracellular matrix formation in a three-dimensional (3D) leiomyoma model system.
DESIGN: Molecular and immunohistochemical analysis in a cell line evaluated in a three-dimensional culture.
SETTING: Laboratory study.
INTERVENTION(S): Treatment of leiomyoma and myometrial cells with liarozole and TGF-β3 in a three-dimensional culture system.
MAIN OUTCOME MEASURE(S): Quantitative real-time reverse-transcriptase polymerase chain reaction and Western blotting to assess fold gene and protein expression of TGF-β3 and TGF-β3 regulated fibrotic cytokines: collagen 1A1 (COL1A1), fibronectin, and versican before and after treatment with liarozole, and confirmatory immunohistochemical stains of treated three-dimensional cultures.
RESULT(S): Both TGF-β3 gene and protein expression were elevated in leiomyoma cells compared with myometrium in two-dimensional and 3D cultures. Treatment with liarozole decreased TGF-β3 gene and protein expression. Extracellular matrix components versican, COL1A1, and fibronectin were also decreased by liarozole treatment in 3D cultures. Treatment of 3D cultures with TGF-β3 increased gene expression and protein production of COL1A1, fibronectin, and versican.
CONCLUSION(S): Liarozole decreased TGF-β3 and TGF-β3-mediated extracellular matrix expression in a 3D uterine leiomyoma culture system.
The presence of regulatory T cells (Treg) in solid tumors is known to play a role in patient survival in ovarian cancer and other malignancies. We assessed inherited genetic variations via 749 tag single-nucleotide polymorphisms (SNP) in 25 Treg-associated genes (CD28, CTLA4, FOXP3, IDO1, IL10, IL10RA, IL15, 1L17RA, IL23A, IL23R, IL2RA, IL6, IL6R, IL8, LGALS1, LGALS9, MAP3K8, STAT5A, STAT5B, TGFB1, TGFB2, TGFB3, TGFBR1, TGRBR2, and TGFBR3) in relation to ovarian cancer survival. We analyzed genotype and overall survival in 10,084 women with invasive epithelial ovarian cancer, including 5,248 high-grade serous, 1,452 endometrioid, 795 clear cell, and 661 mucinous carcinoma cases of European descent across 28 studies from the Ovarian Cancer Association Consortium (OCAC). The strongest associations were found for endometrioid carcinoma and IL2RA SNPs rs11256497 [HR, 1.42; 95% confidence interval (CI), 1.22-1.64; P = 5.7 × 10(-6)], rs791587 (HR, 1.36; 95% CI, 1.17-1.57; P = 6.2 × 10(-5)), rs2476491 (HR, = 1.40; 95% CI, 1.19-1.64; P = 5.6 × 10(-5)), and rs10795763 (HR, 1.35; 95% CI, 1.17-1.57; P = 7.9 × 10(-5)), and for clear cell carcinoma and CTLA4 SNP rs231775 (HR, 0.67; 95% CI, 0.54-0.82; P = 9.3 × 10(-5)) after adjustment for age, study site, population stratification, stage, grade, and oral contraceptive use. The rs231775 allele associated with improved survival in our study also results in an amino acid change in CTLA4 and previously has been reported to be associated with autoimmune conditions. Thus, we found evidence that SNPs in genes related to Tregs seem to play a role in ovarian cancer survival, particularly in patients with clear cell and endometrioid epithelial ovarian cancer.
A growing body of evidence supports that the epithelial-to-mesenchymal transition (EMT), which occurs during cancer development and progression, has a crucial role in metastasis by enhancing the motility of tumor cells. Transforming growth factor-β (TGF-β) is known to induce EMT in a number of cancer cell types; however, the mechanism underlying this transition process is not fully understood. In this study we have demonstrated that TGF-β upregulates the expression of tumor suppressor protein Par-4 (prostate apoptosis response-4) concomitant with the induction of EMT. Mechanistic investigations revealed that exogenous treatment with each TGF-β isoform upregulates Par-4 mRNA and protein levels in parallel levels of phosphorylated Smad2 and IκB-α increase. Disruption of TGF-β signaling by using ALK5 inhibitor, neutralizing TGF-β antibody or phosphoinositide 3-kinase inhibitor reduces endogenous Par-4 levels, suggesting that both Smad and NF-κB pathways are involved in TGF-β-mediated Par-4 upregulation. NF-κB-binding sites in Par-4 promoter have previously been reported; however, using chromatin immunoprecipitation assay we showed that Par-4 promoter region also contains Smad4-binding site. Furthermore, TGF-β promotes nuclear localization of Par-4. Prolonged TGF-β3 treatment disrupts epithelial cell morphology, promotes cell motility and induces upregulation of Snail, vimentin, zinc-finger E-box binding homeobox 1 and N-Cadherin and downregulation of Claudin-1 and E-Cadherin. Forced expression of Par-4, results in the upregulation of vimentin and Snail expression together with increase in cell migration. In contrast, small interfering RNA-mediated silencing of Par-4 expression results in decrease of vimentin and Snail expression and prevents TGF-β-induced EMT. We have also uncovered a role of X-linked inhibitor of apoptosis protein in the regulation of endogenous Par-4 levels through inhibition of caspase-mediated cleavage. In conclusion, our findings suggest that Par-4 is a novel and essential downstream target of TGF-β signaling and acts as an important factor during TGF-β-induced EMT.
BACKGROUND: Transforming growth factor (TGF)-β signaling pathway, may act both as a tumor suppressor and as a tumor promoter in pancreatic cancer, depending on tumor stage and cellular context. TGF-β pathway has been under intensive investigation as a potential therapeutic target in the treatment of cancer. We hypothesized a correlation between TGF-βR2/SMAD4 expression in the tumor, plasma TGF-β1 ligand level, genetic variation in TGF-B pathway and prognosis of pancreatic cancer.
METHOD: We examined TGF-βR2 and SMAD4 protein expression in biopsy or surgical samples from 91 patients with pancreatic ductal adenocarcinoma (PDAC) using immunohistochemistry. Plasma level of TGF-β1 was measured in 644 patients with PDAC using ELISA. Twenty-eight single nucleotide polymorphisms (SNP) of the TGF-β1, TGF-β2, TGF-β3, TGF-βR1, TGF-βR2, and SMAD4 genes were determined in 1636 patients with PDAC using the Sequenom method. Correlation between protein expression in the tumor, plasma TGF-β1 level, and genotypes with overall survival (OS) was evaluated with Cox proportional regression models.
RESULTS: The expression level of TGF-βR2 and SMAD4 as an independent marker was not associated with OS. However, patients with both low nuclear staining of TGF-βR2 and high nuclear staining of SMAD4 may have better survival (P = 0.06). The mean and median level of TGF-β1 was 15.44 (SD: 10.99) and 12.61 (interquartile range: 8.31 to 19.04) ng/ml respectively. Patients with advanced disease and in the upper quartile range of TGF-β1 level had significantly reduced survival than those with low levels (P = 0.02). A significant association of SMAD4 SNP rs113545983 with overall survival was observed (P<0.0001).
CONCLUSION: Our data provides valuable baseline information regarding the TGF-β pathway in pancreatic cancer, which can be utilized in targeted therapy clinical trials. High TGF-β1 plasma level, SMAD4 SNP or TGF-βR2/SMAD4 tumor protein expression may suggest a dependence on this pathway in patients with advanced pancreatic cancer.
Shaikhibrahim Z, Menon R, Braun M, et al.MED15, encoding a subunit of the mediator complex, is overexpressed at high frequency in castration-resistant prostate cancer.
Int J Cancer. 2014; 135(1):19-26 [PubMed
] Related Publications
The mediator complex is an evolutionary conserved key regulator of transcription of protein-coding genes and an integrative hub for diverse signaling pathways. In this study, we investigated whether the mediator subunit MED15 is implicated in castration-resistant prostate cancer (CRPC). MED15 expression and copy number/rearrangement status were assessed by immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH), respectively on 718 prostate cancer (PCa) specimens and sequenced by Sanger on a subset. Furthermore, SMAD3 phosphorylation, androgen receptor (AR) and proliferation markers were evaluated by IHC. In PCa cells, siRNA/shRNA knockdown of MED15 was followed by proliferation assays with/without dihydrotestosterone (DHT), and treatments with recombinant TGF-β3. Our results show that MED15 is overexpressed in 76% of distant metastatic CRPC (CRPC(MET) ) and 70% of local-recurrent CRPC (CRPC(LOC) ), in contrast to low frequencies in androgen-sensitive PCa, and no expression in benign prostatic tissue. Furthermore, MED15 overexpression correlates with worse clinical outcome thus defining a highly lethal phenotype. Moreover, TGF-β signaling activation associates with MED15 overexpression in PCa tissues, and leads to increased expression of MED15 in PCa cells. MED15 knockdown effects phosphorylation and shuttling of p-SMAD3 to the nucleus as well as TGF-β-enhanced proliferation. In PCa tissues, MED15 overexpression associates with AR overexpression/amplification and correlates with high proliferative activity. MED15 knockdown decreases both androgen-dependent and -independent proliferation in PCa cells. Taken together, these findings implicate MED15 in CRPC, and as MED15 is evolutionary conserved, it is likely to emerge as a lethal phenotype in other therapeutic-resistant diseases, and not restricted to our disease model.
BACKGROUND: Cancer-associated fibroblasts (CAFs) activated by tumour cells are the predominant type of stromal cells in breast cancer tissue. The reciprocal effect of CAFs on breast cancer cells and the underlying molecular mechanisms are not fully characterised.
METHODS: Stromal fibroblasts were isolated from invasive breast cancer tissues and the conditioned medium of cultured CAFs (CAF-CM) was collected to culture the breast cancer cell lines MCF-7, T47D and MDA-MB-231. Neutralising antibody and small-molecule inhibitor were used to block the transforming growth factor-β (TGF-β) signalling derived from CAF-CM, which effect on breast cancer cells.
RESULTS: The stromal fibroblasts isolated from breast cancer tissues showed CAF characteristics with high expression levels of α-smooth muscle actin and SDF1/CXCL12. The CAF-CM transformed breast cancer cell lines into more aggressive phenotypes, including enhanced cell-extracellular matrix adhesion, migration and invasion, and promoted epithelial-mesenchymal transition (EMT). Cancer-associated fibroblasts secreted more TGF-β1 than TGF-β2 and TGF-β3, and activated the TGF-β/Smad signalling pathway in breast cancer cells. The EMT phenotype of breast cancer cells induced by CAF-CM was reversed by blocking TGF-β1 signalling.
CONCLUSION: Cancer-associated fibroblasts promoted aggressive phenotypes of breast cancer cells through EMT induced by paracrine TGF-β1. This might be a common mechanism for acquiring metastatic potential in breast cancer cells with different biological characteristics.
BACKGROUND: Colorectal cancer (CRC) is characterised by hypoxia, which activates gene transcription through hypoxia-inducible factors (HIF), as well as by expression of epidermal growth factor (EGF) and EGF receptors, targeting of which has been demonstrated to provide therapeutic benefit in CRC. Although EGF has been demonstrated to induce expression of angiogenic mediators, potential interactions in CRC between EGF-mediated signalling and the hypoxia/HIF pathway remain uncharacterised.
METHODS: PCR-based profiling was applied to identify angiogenic genes in Caco-2 CRC cells regulated by hypoxia, the hypoxia mimetic dimethyloxallylglycine (DMOG) and/or EGF. Western blotting was used to determine the role of HIF-1alpha, HIF-2alpha and MAPK cell signalling in mediating the angiogenic responses.
RESULTS: We identified a total of 9 angiogenic genes, including angiopoietin-like (ANGPTL) 4, ephrin (EFNA) 3, transforming growth factor (TGF) β1 and vascular endothelial growth factor (VEGF), to be upregulated in a HIF dependent manner in Caco-2 CRC cells in response to both hypoxia and the hypoxia mimetic dimethyloxallylglycine (DMOG). Stimulation with EGF resulted in EGFR tyrosine autophosphorylation, activation of p42/p44 MAP kinases and stabilisation of HIF-1α and HIF-2α proteins. However, expression of 84 angiogenic genes remained unchanged in response to EGF alone. Crucially, addition of DMOG in combination with EGF significantly increased expression of a further 11 genes (in addition to the 9 genes upregulated in response to either DMOG alone or hypoxia alone). These additional genes included chemokines (CCL-11/eotaxin-1 and interleukin-8), collagen type IV α3 chain, integrin β3 chain, TGFα and VEGF receptor KDR.
CONCLUSION: These findings suggest that although EGFR phosphorylation activates the MAP kinase signalling and promotes HIF stabilisation in CRC, this alone is not sufficient to induce angiogenic gene expression. In contrast, HIF activation downstream of hypoxia/DMOG drives expression of genes such as ANGPTL4, EFNA3, TGFβ1 and VEGF. Finally, HIF activation synergises with EGF-mediated signalling to additionally induce a unique sub-group of candidate angiogenic genes. Our data highlight the complex interrelationship between tumour hypoxia, EGF and angiogenesis in the pathogenesis of CRC.
Liu Y, Song H, Pan J, Zhao JComprehensive gene expression analysis reveals multiple signal pathways associated with prostate cancer.
J Appl Genet. 2014; 55(1):117-24 [PubMed
] Related Publications
Prostate cancer (PC) depends on androgenic signaling for growth and survival. To data, the exact molecular mechanism of hormone controlling proliferation and tumorigenesis in the PC remains unclear. Therefore, in this study, we explored the differentially expressed genes (DEGs) and identified featured genes related to hormone stimulus from PC. Two sets of gene expression data, including PC and normal control sample, were downloaded from Gene Expression Omnibus (GEO) database. The t-test was used to identify DEGs between PC and controls. Gene ontology (GO) functional annotation was applied to analyze the function of DEGs and screen hormone-related DEGs. Then these hormone-related DEGs were further analyzed in constructed cancer network and Human Protein Reference Database to screen important signaling pathways they participated in. A total of 912 DEGs were obtained which included 326 up-regulated genes and 586 down-regulated genes. GO functional enrichment analysis identified 50 hormone-related DEGs associated with PC. After pathway and PPI network analysis, we found these hormone-related DEGs participated in several important signaling pathways including TGF-β (TGFB2, TGFB3 and TGFBR2), MAPK (TGFB2, TGFB3 and TGFBR2), insulin (PIK3R3, SHC1 and EIF4EBP1), and p53 signaling pathways (CCND2 and CDKN1A). In addition, a total of five hormone-related DEGs (SHC1, CAV1, RXRA, CDKN1A and SRF) were located in the center of PPI network and 12 hormone-related DEGs formed six protein modules. These important signal pathways and hormone-related DEGs may provide potential therapeutic targets for PC.
BACKGROUND: Lung cancer is a common cancer and the leading cause of cancer-related death worldwide. SIX3 is a human homologue of the highly conserved sine oculis gene family essential during embryonic development in vertebrates, and encodes a homeo-domain containing transcription factor. Little is known about the role of SIX3 in human tumorigenesis. This study is to assess the expression/function of SIX3 and the significance of SIX3 as a prognostic biomarker in lung adenocarcinoma.
METHODS: Quantitative real-time RT-PCR was used to analyze SIX3 mRNA expression and quantitative methylation specific PCR (MSP) was used to examine promoter methylation. MTS and colony formation assays were performed to examine cell proliferation. Wound healing assays were used to assess cell migration, and microarrays were utilized to examine genes regulated by SIX3 in lung cancer cells. Association of SIX3 expression levels with clinical outcomes of patients with lung adenocarcinoma was evaluated using the Kaplan-Meier method and a multivariate Cox proportional hazards regression model.
RESULTS: SIX3 was down-regulated in lung adenocarcinoma tissues compared to their matched adjacent normal tissues, and this down-regulation was associated with methylation of the SIX3 promoter. SIX3 was also methylation-silenced in lung cancer cell lines. Restoration of SIX3 in lung cancer cells lacking endogenous SIX3 suppressed cell proliferation and migration, and downregulated a number of genes involved in proliferation and metastasis such as S100P, TGFB3, GINS3 and BAG1. Moreover, SIX3 mRNA expression was associated with significantly improved overall survival (OS) and progression-free survival (PFS) in adenocarcinoma patients and patients with bronchioloalveolar carcinoma (BAC) features.
CONCLUSIONS: SIX3 may play an important role as a novel suppressor in human lung cancer. SIX3 has potential as a novel prognostic biomarker for patients with lung adenocarcinomas.
Van der Woude syndrome is the most common form of syndromic orofacial clefting, accounting for 1-2% of all patients with cleft lip and/or cleft palate. Van der Woude and popliteal pterygium syndromes are caused by mutations in IRF6, but phenotypic variability within and among families with either syndrome suggests that other genetic factors contribute to the phenotypes. The aim of this study was to identify common variants acting as genetic modifiers of IRF6 as well as genotype-phenotype correlations based on mutation type and location. We identified an association between mutations in the DNA-binding domain of IRF6 and limb defects (including pterygia). Although we did not detect formally significant associations with the genes tested, borderline associations suggest several genes that could modify the VWS phenotype, including FOXE1, TGFB3, and TFAP2A. Some of these genes are hypothesized to be part of the IRF6 gene regulatory network and may suggest additional genes for future study when larger sample sizes are also available. We also show that families with the Van de Woude phenotype but in whom no mutations have been identified have a lower frequency of cleft lip, suggesting there may be locus and/or mutation class differences in Van de Woude syndrome.
Stepanenko AA, Vassetzky YS, Kavsan VMAntagonistic functional duality of cancer genes.
Gene. 2013; 529(2):199-207 [PubMed
] Related Publications
Cancer evolution is a stochastic process both at the genome and gene levels. Most of tumors contain multiple genetic subclones, evolving in either succession or in parallel, either in a linear or branching manner, with heterogeneous genome and gene alterations, extensively rewired signaling networks, and addicted to multiple oncogenes easily switching with each other during cancer progression and medical intervention. Hundreds of discovered cancer genes are classified according to whether they function in a dominant (oncogenes) or recessive (tumor suppressor genes) manner in a cancer cell. However, there are many cancer "gene-chameleons", which behave distinctly in opposite way in the different experimental settings showing antagonistic duality. In contrast to the widely accepted view that mutant NADP(+)-dependent isocitrate dehydrogenases 1/2 (IDH1/2) and associated metabolite 2-hydroxyglutarate (R)-enantiomer are intrinsically "the drivers" of tumourigenesis, mutant IDH1/2 inhibited, promoted or had no effect on cell proliferation, growth and tumorigenicity in diverse experiments. Similar behavior was evidenced for dozens of cancer genes. Gene function is dependent on genetic network, which is defined by the genome context. The overall changes in karyotype can result in alterations of the role and function of the same genes and pathways. The diverse cell lines and tumor samples have been used in experiments for proving gene tumor promoting/suppressive activity. They all display heterogeneous individual karyotypes and disturbed signaling networks. Consequently, the effect and function of gene under investigation can be opposite and versatile in cells with different genomes that may explain antagonistic duality of cancer genes and the cell type- or the cellular genetic/context-dependent response to the same protein. Antagonistic duality of cancer genes might contribute to failure of chemotherapy. Instructive examples of unexpected activity of cancer genes and "paradoxical" effects of different anticancer drugs depending on the cellular genetic context/signaling network are discussed.
Naber HP, Drabsch Y, Snaar-Jagalska BE, et al.Snail and Slug, key regulators of TGF-β-induced EMT, are sufficient for the induction of single-cell invasion.
Biochem Biophys Res Commun. 2013; 435(1):58-63 [PubMed
] Related Publications
TGF-β plays a dual role in cancer; in early stages it inhibits tumor growth, whereas later it promotes invasion and metastasis. TGF-β is thought to be pro-invasive by inducing epithelial-to-mesenchymal transition (EMT) via induction of transcriptional repressors, including Slug and Snail. In this study, we investigated the role of Snail and Slug in TGF-β-induced invasion in an in vitro invasion assay and in an embryonic zebrafish xenograft model. Ectopic expression of Slug or Snail promoted invasion of single, rounded amoeboid cells in vitro. In an embryonic zebrafish xenograft model, forced expression of Slug and Snail promoted single cell invasion and metastasis. Slug and Snail are sufficient for the induction of single-cell invasion in an in vitro invasion assay and in an embryonic zebrafish xenograft model.
Kapral M, Wawszczyk J, Hollek A, Weglarz LInduction of the expression of genes encoding TGF-beta isoforms and their receptors by inositol hexaphosphate in human colon cancer cells.
Acta Pol Pharm. 2013 Mar-Apr; 70(2):357-63 [PubMed
] Related Publications
Transforming growth factors-beta (TGF-beta) are multifunctional cytokines involved in the regulation of cell development, differentiation, survival and apoptosis. They are also potent anticancer agents that inhibit uncontrolled proliferation of cells. Incorrect TGF-beta regulation has been implicated in the pathogenesis of many diseases including inflammation and cancer. In humans, the TGF-beta family consists of three members (TGF-beta1, 2, 3) that show high similarity and homology. TGF-betas exert biological activities on various cell types including neoplastic cells via their specific receptors. Inositol hexaphosphate (phytic acid, IP6), a phytochemical has been reported to possess various health benefits. The aim of this study was to examine the effect of IP6 on the expression of genes encoding TGF-beta1, TGF-beta2, TGF-beta3 isoforms and their receptors TbetaRI, TbetaRII, TbetaRIII in human colorectal cancer cell line Caco-2. The cells were treated with 0.5, 1 and 2.5 mM IP6 for 3, 6 and 12 h. The untreated Caco-2 cells were used as the control. Quantification of genes expression was performed by real time QRT-PCR technique with a SYBR Green I chemistry. The experimental data revealed that the TGF-beta1 mRNA was the predominant isoform in Caco-2 cells and that IP6 enhanced transcriptional activity of genes of all three TGF-beta isoforms and their receptors TbetaRI, TbetaRII TbetaRIII in these cells. At concentrations up to 1 mM, IP6 over-expressed the genes in 6 h lasting cultures, and its higher dose (2.5 mM) caused successively increasing transcript level of TGF-beta isoforms and receptors with the duration of experiment up to 12 h. The findings of this study indicate that one of anti-cancer abilities of IP6 can be realized by enhancing the gene expression of TGF-beta isoforms and their receptors at the transcriptional level.
Roth P, Silginer M, Goodman SL, et al.Integrin control of the transforming growth factor-β pathway in glioblastoma.
Brain. 2013; 136(Pt 2):564-76 [PubMed
] Related Publications
Transforming growth factor-β is a central mediator of the malignant phenotype of glioblastoma, the most common and malignant form of intrinsic brain tumours. Transforming growth factor-β promotes invasiveness and angiogenesis, maintains cancer cell stemness and induces profound immunosuppression in the host. Integrins regulate cellular adhesion and transmit signals important for cell survival, proliferation, differentiation and motility, and may be involved in the activation of transforming growth factor-β. We report that αvβ3, αvβ5 and αvβ8 integrins are broadly expressed not only in glioblastoma blood vessels but also in tumour cells. Exposure to αv, β3 or β5 neutralizing antibodies, RNA interference-mediated integrin gene silencing or pharmacological integrin inhibition using the cyclic RGD peptide EMD 121974 (cilengitide) results in reduced phosphorylation of Smad2 in most glioma cell lines, including glioma-initiating cell lines and reduced transforming growth factor-β-mediated reporter gene activity, coinciding with reduced transforming growth factor-β protein levels in the supernatant. Time course experiments indicated that the loss of transforming growth factor-β bioactivity due to integrin inhibition likely results from two distinct mechanisms: an early effect on activation of preformed inactive protein, and second, major effect on transforming growth factor-β gene transcription as confirmed by decreased activity of the transforming growth factor-β gene promoter and decreased transforming growth factor-β(1) and transforming growth factor-β(2) messenger RNA expression levels. In vivo, EMD 121974 (cilengitide), which is currently in late clinical development as an antiangiogenic agent in newly diagnosed glioblastoma, was a weak antagonist of pSmad2 phosphorylation. These results validate integrin inhibition as a promising strategy not only to inhibit angiogenesis, but also to block transforming growth factor-β-controlled features of malignancy including invasiveness, stemness and immunosuppression in human glioblastoma.
Although imatinib mesylate (IM) has transformed the treatment of gastrointestinal stromal tumors (GIST), many patients experience primary/secondary drug resistance. In a previous study, we identified a gene signature, consisting mainly of Kruppel-associated box (KRAB) domain containing zinc finger (ZNF) transcriptional repressors that predict short-term response to IM. To determine if these genes have functional significance, a siRNA library targeting these genes was constructed and applied to GIST cells in vitro. These screens identified seventeen "IM sensitizing genes" in GIST cells (sensitization index (SI) <0.85 ratio of drug/vehicle) with a false discovery rate (FDR) <15%, including twelve ZNF genes, the majority of which are located within the HSA19p12-13.1 locus. These genes were shown to be highly specific to IM and another tyrosine kinase inhibitor (TKI), sunitinib, in GIST cells. In order to determine mechanistically how these ZNFs might be modulating response to IM, RNAi approaches were used to individually silence genes within the predictive signature in GIST cells and expression profiling was performed. Knockdown of the 14 IM-sensitizing genes (10 ZNFs) universally led to downregulation of six genes, including TGFb3, periostin, and NEDD9. These studies implicate a role of KRAB-ZNFs in modulating response to TKIs in GIST.
Oh S, Kim E, Kang D, et al.Transforming growth factor-β gene silencing using adenovirus expressing TGF-β1 or TGF-β2 shRNA.
Cancer Gene Ther. 2013; 20(2):94-100 [PubMed
] Related Publications
Tumor cells secrete a variety of cytokines to outgrow and evade host immune surveillance. In this context, transforming growth factor-β1 (TGF-β1) is an extremely interesting cytokine because it has biphasic effects in cancer cells and normal cells. TGF-β1 acts as a growth inhibitor in normal cells, whereas it promotes tumor growth and progression in tumor cells. Overexpression of TGF-β1 in tumor cells also provides additional oncogenic activities by circumventing the host immune surveillance. Therefore, this study ultimately aimed to test the hypothesis that suppression of TGF-β1 in tumor cells by RNA interference can have antitumorigenic effects. However, we demonstrated here that the interrelation between TGF-β isotypes should be carefully considered for the antitumor effect in addition to the selection of target sequences with highest efficacy. The target sequences were proven to be highly specific and effective for suppressing both TGF-β1 mRNA and protein expression in cells after infection with an adenovirus expressing TGF-β1 short hairpin RNA (shRNA). A single base pair change in the shRNA sequence completely abrogated the suppressive effect on TGF-β1. Surprisingly, the suppression of TGF-β1 induced TGF-β3 upregulation, and the suppression of TGF-β2 induced another unexpected downregulation of both TGF-β1 and TGF-β3. Taken together, this information may prove useful when considering the design for a novel cancer immunogene therapy.
Chen CL, Mahalingam D, Osmulski P, et al.Single-cell analysis of circulating tumor cells identifies cumulative expression patterns of EMT-related genes in metastatic prostate cancer.
Prostate. 2013; 73(8):813-26 [PubMed
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BACKGROUND: Prostate tumors shed circulating tumor cells (CTCs) into the blood stream. Increased evidence shows that CTCs are often present in metastatic prostate cancer and can be alternative sources for disease profiling and prognostication. Here we postulate that CTCs expressing genes related to epithelial-mesenchymal transition (EMT) are strong predictors of metastatic prostate cancer.
METHODS: A microfiltration system was used to trap CTCs from peripheral blood based on size selection of large epithelial-like cells without CD45 leukocyte marker. These cells individually retrieved with a micromanipulator device were assessed for cell membrane physical properties using atomic force microscopy. Additionally, 38 CTCs from eight prostate cancer patients were used to determine expression profiles of 84 EMT-related and reference genes using a microfluidics-based PCR system.
RESULTS: Increased cell elasticity and membrane smoothness were found in CTCs compared to noncancerous cells, highlighting their potential invasiveness and mobility in the peripheral circulation. Despite heterogeneous expression patterns of individual CTCs, genes that promote mesenchymal transitioning into a more malignant state, including IGF1, IGF2, EGFR, FOXP3, and TGFB3, were commonly observed in these cells. An additional subset of EMT-related genes (e.g., PTPRN2, ALDH1, ESR2, and WNT5A) were expressed in CTCs of castration-resistant cancer, but less frequently in castration-sensitive cancer.
CONCLUSIONS: The study suggests that an incremental expression of EMT-related genes in CTCs is associated with metastatic castration-resistant cancer. Although CTCs represent a group of highly heterogeneous cells, their unique EMT-related gene signatures provide a new opportunity for personalized treatments with targeted inhibitors in advanced prostate cancer patients.
BACKGROUND: As major regulators of normal chondrogenesis, the bone morphogenic protein (BMP) and transforming growth factor β (TGFB) signaling pathways may be involved in the development and progression of central chondrosarcoma. In order to uncover their possible implication, the aim of this study was to perform a systematic quantitative study of the expression of BMPs, TGFBs and their receptors and to assess activity of the corresponding pathways in central chondrosarcoma.
METHODS: Gene expression analysis was performed by quantitative RT-PCR in 26 central chondrosarcoma and 6 healthy articular cartilage samples. Expression of endoglin and nuclear localization of phosphorylated Smad1/5/8 and Smad2 was assessed by immunohistochemical analysis.
RESULTS: The expression of TGFB3 and of the activin receptor-like kinase ALK2 was found to be significantly higher in grade III compared to grade I chondrosarcoma. Nuclear phosphorylated Smad1/5/8 and Smad2 were found in all tumors analyzed and the activity of both signaling pathways was confirmed by functional reporter assays in 2 chondrosarcoma cell lines. Immunohistochemical analysis furthermore revealed that phosphorylated Smad1/5/8 and endoglin expression were significantly higher in high-grade compared to low-grade chondrosarcoma and correlated to each other.
CONCLUSIONS: The BMP and TGFβ signaling pathways were found to be active in central chondrosarcoma cells. The correlation of Smad1/5/8 activity to endoglin expression suggests that, as described in other cell types, endoglin could enhance Smad1/5/8 signaling in high-grade chondrosarcoma cells. Endoglin expression coupled to Smad1/5/8 activation could thus represent a functionally important signaling axis for the progression of chondrosarcoma and a regulator of the undifferentiated phenotype of high-grade tumor cells.
An N, Rausch-fan X, Wieland M, et al.Initial attachment, subsequent cell proliferation/viability and gene expression of epithelial cells related to attachment and wound healing in response to different titanium surfaces.
Dent Mater. 2012; 28(12):1207-14 [PubMed
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OBJECTIVES: A tight seal between the epithelium and the dental implant surface is required to prevent bacterial inflammation and soft tissue recession and therefore to demonstrate a long-term success. Surface hydrophilicity was recently shown to promote osseointegration. The aim of this study was to investigate the influence of surface hydrophilicity in combination with surface topography of Ti implant surfaces on the behavior and activation/differentiation of epithelial cells using a set of in vitro experiments mimicking the implant-soft tissue contact.
METHODS: Hydrophobic acid-etched (A) and coarse-grit-blasted, acid-etched (SLA) surfaces and hydrophilic acid-etched (modA) and modSLA surfaces were produced. The behavior of an oral squamous cell carcinoma cell line (HSC-2) grown on all surfaces was compared through determination of cell attachment and proliferation/viability (CCK-8 and MTT assay), time-lapse microscopy of fluorescence labeled cells and determination of gene expression by real time polymerase chain reaction.
RESULTS: Within the surfaces with similar wettability cell spreading and cell movements observed by time-lapse microscopy after one day of incubation were most pronounced on smoother (A and modA) surfaces compared to rougher (SLA and modSLA) surfaces. Within the surfaces with similar roughness the hydrophilic surfaces (modA and modSLA) showed more cell spreading and cell activity compared to the hydrophobic surfaces (A and SLA). The relative gene expressions of cytokeratin14, integrin α6, integrin β4, vinculin, transforming growth factor (TGF)-β, TGF-β1, and TGF-β3 were decreased in HSC-2 on all four types of Ti surfaces compared to control surfaces (tissue culture polystyrene; p<0.01) and there was no significant difference of gene expression on the four different implant-surfaces.
SIGNIFICANCE: We have demonstrated that for proliferation and spreading of HSC-2 cells the smoother and hydrophilic surface is optimal (modA). These results suggest that surface hydrophilicity might positively influence the epithelial seal around dental implants. All tested titanium surfaces downregulate cell attachment, cell proliferation, expression of adhesion promoters, and cytokines involved in wound healing in HSC-2 cells compared to control surfaces.
Petrella BL, Armstrong DA, Vincenti MPInterleukin-1 beta and transforming growth factor-beta 3 cooperate to activate matrix metalloproteinase expression and invasiveness in A549 lung adenocarcinoma cells.
Cancer Lett. 2012; 325(2):220-6 [PubMed
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Cytokines present in the tumor microenvironment can promote the invasiveness and metastatic potential of cancer cells. We therefore investigated the effects of interleukin-1 beta (IL-1B) and transforming growth factor beta-3 (TGFB3) on the non-small cell lung carcinoma (NSCLC) cell line A549. We found that these cytokines synergistically activated matrix metalloproteinase (MMP)-1, MMP-3, and MMP-10 gene expression in these cells through mitogen-activated protein kinase (MAPK)-dependent pathways. Consistent with this, both cytokines stimulated epithelial to mesenchymal transition and MAPK-dependent invasion through Matrigel™. These studies identify IL-1B and TGFB3 as pro-invasive factors in NSCLC and potential therapeutic targets for tumor progression.
Reinertsen T, Halgunset J, Viset T, et al.Gene expressional changes in prostate fibroblasts from cancerous tissue.
APMIS. 2012; 120(7):558-71 [PubMed
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Prostate cancer is the most common type of cancer in men. It is assumed that the tumor microenvironment of the prostate contributes to invasion and metastasis. Stroma-epithelial crosstalk has shown to change with progression of prostate cancer, and thereby the stromal compartment might be an attractive target in diagnostic and therapeutic approaches to prostate cancer. The purpose of this project was to study the reciprocal influence between fibroblasts and cancer cells in prostate cancer. Prostate fibroblast primary cultures from areas with cancer and hyperplasia were cocultivated with cells of the PC-3 lineage. Gene expression profiles of both cell types were studied to reveal possible associations to cancer invasion and metastasis. There were 383 differentially expressed genes between fibroblasts from cancerous areas and fibroblasts from areas with hyperplasia before cocultivation with PC-3 cells. Several of the differentially expressed gene classes are associated with cancer development and metastasis. After cocultivation, there were 26 differentially expressed genes between cancerous and hyperplastic fibroblasts. There were only three differentially expressed genes between PC-3 cells that had been cocultivated with cancerous fibroblasts and PC-3 cells that had been cocultivated with hyperplastic fibroblasts. The fibroblasts from cancer areas showed a different expression pattern from the characteristics reported as reactive stroma in previous studies. We found tenascin C to be downregulated, which is contrary to previous findings. TGF-β3 and TGF-βR3 were also downregulated, which has been associated with disturbance of TGF-β signaling during prostate cancer progression. Cocultivation with PC-3 cells seems to make the cancerous and hyperplastic fibroblasts more alike each other, as the number of differentially expressed genes decreases. It is desirable to find out if the reduction in differential gene expression is attributable to that hyperplastic fibroblasts become more alike the cancerous fibroblasts or vice versa. Also, we think that the lower expression levels of c-Jun and c-Fos in cancerous fibroblasts without coculture may cause loss of normal fibroblast differentiation, proliferation and inflammatory response, and hence, favor the proliferation and invasion of cancer cells.
BACKGROUND: The extracellular matrix (ECM) has a key role in facilitating the progression of ovarian cancer and we have shown recently that the secreted ECM protein TGFBI modulates the response of ovarian cancer to paclitaxel-induced cell death.
RESULTS: We have determined TGFBI signaling from the extracellular environment is preferential for the cell surface αvß3 integrin heterodimer, in contrast to periostin, a TGFBI paralogue, which signals primarily via a ß1 integrin-mediated pathway. We demonstrate that suppression of ß1 integrin expression, in ß3 integrin-expressing ovarian cancer cells, increases adhesion to rTGFBI. In addition, Syndecan-1 and -4 expression is dispensable for adhesion to rTGFBI and loss of Syndecan-1 cooperates with the loss of ß1 integrin to further enhance adhesion to rTGFBI. The RGD motif present in the carboxy-terminus of TGFBI is necessary, but not sufficient, for SKOV3 cell adhesion and is dispensable for adhesion of ovarian cancer cells lacking ß3 integrin expression. In contrast to TGFBI, the carboxy-terminus of periostin, lacking a RGD motif, is unable to support adhesion of ovarian cancer cells. Suppression of ß3 integrin in SKOV3 cells increases resistance to paclitaxel-induced cell death while suppression of ß1 integrin has no effect. Furthermore, suppression of TGFBI expression stimulates a paclitaxel resistant phenotype while suppression of fibronectin expression, which primarily signals through a ß1 integrin-mediated pathway, increases paclitaxel sensitivity.
CONCLUSIONS: Therefore, different ECM components use distinct signaling mechanisms in ovarian cancer cells and in particular, TGFBI preferentially interacts through a ß3 integrin receptor mediated mechanism to regulate the response of cells to paclitaxel-induced cell death.
Malik M, Catherino WHDevelopment and validation of a three-dimensional in vitro model for uterine leiomyoma and patient-matched myometrium.
Fertil Steril. 2012; 97(6):1287-93 [PubMed
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OBJECTIVE: To develop and validate a three-dimensional (3D) culture system of leiomyoma and myometrial cells.
DESIGN: In vitro study of immortalized cultures of patient-matched leiomyoma and myometrium.
SETTING: University hospital.
PATIENT(S): Women undergoing hysterectomy for symptomatic leiomyomas.
INTERVENTION(S): Immortalized cell cultures, quantitative reverse-transcriptase polymerase chain reaction (qRT-PCR), cytoimmunofluorescence, and Western blot analysis.
MAIN OUTCOME MEASURE(S): Morphologic features; expression of COL1A1, versican, fibronectin, dermatopontin, and transforming growth factor β3 (TGF-β3); and integrin-mediated 3D structural formation.
RESULT(S): Cells in 3D culture maintained spindle morphology. There was elevated expression of collagen 1A1 (6.66 ± 1.5), total versican (4.78 ± 0.5), fibronectin (3.94 ± 0.3), and TGF-β3 (2.21 ± 0.1) as was seen in progenitor tissue. Dermatopontin gene was down-regulated (0.29 ± 0.1), also similar to values in the surgical tissue sample. Myometrial cells in 3D culture responded to TGF-β3 with increased gene expression of collagen 1A1, fibronectin, and versican, and decreased expression of dermatopontin gene recapitulating the leiomyoma phenotype. Integrin-β1-inhibiting antibody disrupted the cell-extracellular matrix (ECM) communication and induced apoptosis.
CONCLUSION(S): Three-dimensional 3D leiomyoma cell cultures maintain the molecular phenotype of progenitor tissue, produce ECM, and interact with the ECM directly. This model system allows for assessment of the mechanism of aberrant ECM formation as well as the effectiveness of various potential therapies.
Kolacinska A, Fendler W, Szemraj J, et al.Gene expression and pathologic response to neoadjuvant chemotherapy in breast cancer.
Mol Biol Rep. 2012; 39(7):7435-41 [PubMed
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Pathologic complete response after neoadjuvant systemic treatment appears to be a valid surrogate for better overall survival in breast cancer patients. Currently, together with standard clinicopathologic assessment, novel molecular biomarkers are being exhaustively tested in order to look into the heterogeneity of breast cancer. The aim of our study was to examine an association between 23-gene real-time-PCR expression assay including ABCB1, ABCC1, BAX, BBC3, BCL2, CASP3, CYP2D6, ERCC1, FOXC1, GAPDH, IGF1R, IRF1, MAP2, MAPK 8, MAPK9, MKI67, MMP9, NCOA3, PARP1, PIK3CA, TGFB3, TOP2A, and YWHAZ receptor status of breast cancer core biopsies sampled before neoadjuvant chemotherapy (anthracycline and taxanes) and pathologic response. Core-needle biopsies were collected from 42 female patients with inoperable locally advanced breast cancer or resectable tumors suitable for downstaging, before any treatment. Expressions of 23 genes were determined by means of TagMan low density arrays. Analysis of variance was used to select genes with discriminatory potential between receptor subtypes. We introduced a correction for false discovery rates (presented as q values) due to multiple hypothesis testing. Statistical analysis showed that seven genes out of a 23-gene real-time-PCR expression assay differed significantly in relation to pathologic response regardless of breast cancer subtypes. Among these genes, we identified: BAX (p = 0.0146), CYP2D6 (p = 0.0063), ERCC1 (p = 0.0231), FOXC1 (p = 0.0048), IRF1 (p = 0.0022), MAP2 (p = 0.0011), and MKI67 (p = 0.0332). The assessment of core biopsy gene profiles and receptor-based subtypes, before neoadjuvant therapy seems to predict response or resistance and to define new signaling pathways to provide more powerful classifiers in breast cancer, hence the need for further research.
Uehara E, Shiiba M, Shinozuka K, et al.Upregulated expression of ADAM12 is associated with progression of oral squamous cell carcinoma.
Int J Oncol. 2012; 40(5):1414-22 [PubMed
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ADAMs are a disintegrin and metalloproteinase family of membrane-associated metalloproteinases characterized by their multidomain structure, and have been reported to be associated with various malignant tumors. The aim of this study was to identify crucial members of the ADAM family in oral squamous cell carcinoma (OSCC), and to reveal their biological function and clinical significance. To clarify whether ADAM family genes are involved in OSCC, changes in the expression profile were investigated by real-time quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR) analysis and immunohistochemical analysis. Functional analysis was performed by comparing cellular proliferation of siADAM-transfected cell lines and parental cell lines. Real-time qRT-PCR analysis identified significantly upregulated expression of ADAM12 in OSCC-derived cell lines. This was validated in OSCC samples using real-time qRT-PCR and immuno-histochemical staining. ADAM12 expression was correlated with TNM classification; significantly greater expression of ADAM12 was observed in tumors with higher T classification and more advanced stages. Moreover, siADAM12-transfected cells showed both a suppressed proliferation rate and increased transforming growth factor (TGF)-β3 expression. Our data indicate that ADAM12 is overexpressed in OSCC and might accelerate cellular proliferation. Its function may be associated with TGF-β signaling. This study suggests that controlling the expression or activity of ADAM12 could be a useful strategy in the development of an effective cure for OSCC.
Lo PK, Kanojia D, Liu X, et al.CD49f and CD61 identify Her2/neu-induced mammary tumor-initiating cells that are potentially derived from luminal progenitors and maintained by the integrin-TGFβ signaling.
Oncogene. 2012; 31(21):2614-26 [PubMed
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Human epidermal growth factor receptor 2 (HER2)/Neu is overexpressed in 20-30% of breast cancers and associated with aggressive phenotypes and poor prognosis. For deciphering the role of HER2/Neu in breast cancer, mouse mammary tumor virus (MMTV)-Her2/neu transgenic mice that develop mammary tumors resembling human HER2-subtype breast cancer have been established. Several recent studies have revealed that HER2/Neu is overexpressed in and regulates self renewal of breast tumor-initiating cells (TICs). However, in the MMTV-Her2/neu transgenic mouse model, the identity of TICs remains elusive, despite previous studies showing supportive evidence for existence of TICs in Her2/neu-induced mammary tumors. Through systematic screening and characterization, we identified that surface markers CD49f, CD61 and ESA were aberrantly overexpressed in Her2-overexpressing mammary tumor cells. Analysis of these markers and CD24 detected anomalous expansion of the luminal progenitor population in preneoplastic mammary glands of Her2/neu transgenic mice, indicating that aberrant luminal progenitors originated in Her2-induced mammary tumors. The combined markers, CD49f and CD61, further delineated the CD49f(high)CD61(high)-sorted fraction as a TIC-enriched population, which displayed increased tumorsphere formation ability, enhanced tumorigenicity both in vitro and in vivo and drug resistance to pacitaxel and doxorubicin. Moreover, the TIC-enriched population manifested increased transforming growth factor-β (TGFβ) signaling and exhibited gene expression signatures of stemness, TGFβ signaling and epithelial-to-mesenchymal transition. Our findings that self-renewal and clonogenicity of TICs were suppressed by pharmacologically inhibiting the TGFβ signaling further indicate that the TGFβ pathway is vital for maintenance of the TIC population. Finally, we showed that the integrin-β3 (CD61) signaling pathway was required for sustaining active TGFβ signaling and self-renewal of TICs. We for the first time developed a technique to highly enrich TICs from mammary tumors of Her2/neu transgenic mice, unraveled their properties and identified the cooperative integrin-β3-TGFβ signaling axis as a potential therapeutic target for HER2-induced TICs.
BACKGROUND: The transforming growth factor (TGF)-β superfamily comprises cytokines such as TGF-β and Bone Morphogenetic Proteins (BMPs), which have a critical role in a multitude of biological processes. In breast cancer, high levels of TGF-β are associated with poor outcome, whereas inhibition of TGF-β-signaling reduces metastasis. In contrast, BMP-7 inhibits bone metastasis of breast cancer cells.
METHODS: In this study, we investigated the effect of BMP-7 on TGF-β-induced invasion in a 3 dimensional invasion assay.
RESULTS: BMP-7 inhibited TGF-β-induced invasion of the metastatic breast cancer cell line MCF10CA1a, but not of its premalignant precursor MCF10AT in a spheroid invasion model. The inhibitory effect appears to be specific for BMP-7, as its closest homolog, BMP-6, did not alter the invasion of MCF10CA1a spheroids. To elucidate the mechanism by which BMP-7 inhibits TGF-β-induced invasion, we analyzed invasion-related genes. BMP-7 inhibited TGF-β-induced expression of integrin α(v)β(3) in the spheroids. Moreover, targeting of integrins by a chemical inhibitor or knockdown of integrin β(3) negatively affected TGF-β-induced invasion. On the other hand, overexpression of integrin β(3) counteracted the inhibitory effect of BMP7 on TGF-β-induced invasion.
CONCLUSION: Thus, BMP-7 may exert anti-invasive actions by inhibiting TGF-β-induced expression of integrin β(3).