Epigenetic Silencing of THY1 Tracks the Acquisition of the Notch1-EGFR Signaling in a Xenograft Model of CD44+/CD24low/CD90+ Myoepithelial Cells.

The surface glycoprotein THY is a marker of myoepithelial precursor cells, which are basal cells with epithelial-mesenchymal intermediate phenotype originating from the ectoderm. Myoepithelial precursor cells are lost during progression from in situ to invasive carcinoma. To define the functional role of Thy1-positive cells within the myoepithelial population, we tracked Thy1 expression in human breast cancer samples, isolated THY1-positive myoepithelial progenitor cells (CD44+/CD24low/CD90+), and established long-term cultures (parental cells). Parental cells were used to generate a xenograft model to examine Thy1 expression during tumor formation. Post-transplantation cell cultures lost THY1 expression through methylation at the THY1 locus and this is associated with an increase in EGFR and NOTCH1 transcript levels. Thy1-low cells are sensitive to the EGFR/HER2 dual inhibitor lapatinib. High THY1 expression is associated with poorer relapse-free survival in patients with breast cancer. THY1 methylation may track the shift of bipotent progenitors into differentiated cells. Thy1 is a good candidate biomarker in basal-like breast cancer. IMPLICATIONS: Our findings provide evidence that THY1 expression is lost in xenografts due to promoter methylation. Thy1-low cells with increased EGFR and Notch1 expression are responsive to target therapy. Because DNA methylation is often altered in early cancer development, candidate methylation markers may be exploited as biomarkers for basal-like breast cancer.

Exploring the molecular aspects associated with testicular germ cell tumors: a review.

Testicular germ cell tumors (TGCTs) represent the most common solid tumors affecting young men. They constitute a distinct entity because of their embryonic origin and their unique biological behavior. Recent preclinical data regarding biological signaling machinery as well as genetic and epigenetic mechanisms associated with molecular patterns of tumors have contribute to explain the pathogenesis and the differentiation of TGCTs and to understand the mechanisms responsible for the development of resistance to treatment. In this review, we discuss the main genetic and epigenetic events associated with TGCTs development in order to better define their role in the pathogenesis of these tumors and in cisplatin-acquired resistance.

Correction: Metabolic syndrome, endocrine disruptors and prostate cancer associations: biochemical and pathophysiological evidences.

[This corrects the article DOI: 10.18632/oncotarget.16725.].

ProEx C as Diagnostic Marker for Detection of Urothelial Carcinoma in Urinary Samples: A Review.

The gold standard for the detection of urothelial carcinoma is represented by urethro-cystoscopy and biopsy. Both procedures are invasive and expensive and therefore cytology is often used as first approach to investigate on a possible neoplasia, being a safe and cost-effective diagnostic modality of evaluation. Because cytology alone is not highly sensitive for detection of low grade urothelial carcinoma and recurrence of the disease, several adjunct markers and urine based tests for urothelial carcinoma have been developed, which can help in the final diagnosis. In particular, ProEx C is an immunohistochemical cocktail containing antibodies direct against topoisomerase IIα (TOP2A) and minichromosome maintenance 2 (MCM2) proteins. It proved to be a valid biomarker especially in detecting squamous intraepithelial lesions in cervical liquid-based samples and in discerning these lesions from their mimickers, as well as in ovarian, endometrial, vulvar, primary and metastatic melanomas, breast, pancreatic and renal cell carcinomas. This brief review covers the effective utility of ProEx C as adjunct tool in assessing the urothelial lesions in urine cytology, also providing prognostic and therapeutic information to help in clinical decisions.

Micrornas in prostate cancer: an overview.

Prostate cancer is the second highest cause of cancer mortality after lung tumours. In USA it affects about 2.8 million men and the incidence increases with age in many countries. Therefore, early diagnosis is a very important step for patient clinical evaluation and for a selective and efficient therapy. The study of miRNAs' functions and molecular mechanisms has brought new knowledge in biological processes of cancer. In prostate cancer there is a deregulation of several miRNAs that may function as tumour suppressors or oncogenes. The aim of this review is to analyze the progress made to our understanding of the role of miRNA dysregulation in prostate cancer tumourigenesis.

Epithelial-mesenchymal transition in prostate cancer: an overview.

Prostate cancer is a main urological disease associated with significant morbidity and mortality. Radical prostatectomy and radiotherapy are potentially curative for localized prostate cancer, while androgen deprivation therapy is the initial systemic therapy for metastatic prostate disease. However, despite temporary response, most patients relapse and evolve into castration resistant cancer.Epithelial-mesenchymal transition (EMT) is a complex gradual process that occurs during embryonic development and/or tumor progression. During this process, cells lose their epithelial characteristics and acquire mesenchymal features. Increasing evidences indicate that EMT promotes prostate cancer metastatic progression and it is closely correlated with increased stemness and drug resistance.In this review, we discuss the main molecular events that directly or indirectly govern the EMT program in prostate cancer, in order to better define the role and the mechanisms underlying this process in prostate cancer progression and therapeutic resistance.

Metabolic syndrome, endocrine disruptors and prostate cancer associations: biochemical and pathophysiological evidences.

This review summarizes the main pathophysiological basis of the relationship between metabolic syndrome, endocrine disruptor exposure and prostate cancer that is the most common cancer among men in industrialized countries. Metabolic syndrome is a cluster of metabolic and hormonal factors having a central role in the initiation and recurrence of many western chronic diseases including hormonal-related cancers and it is considered as the world's leading health problem in the coming years. Many biological factors correlate metabolic syndrome to prostate cancer and this review is aimed to focus, principally, on growth factors, cytokines, adipokines, central obesity, endocrine abnormalities and exposure to specific endocrine disruptors, a cluster of chemicals, to which we are daily exposed, with a hormone-like structure influencing oncogenes, tumor suppressors and proteins with a key role in metabolism, cell survival and chemo-resistance of prostate cancer cells. Finally, this review will analyze, from a molecular point of view, how specific foods could reduce the relative risk of incidence and recurrence of prostate cancer or inhibit the biological effects of endocrine disruptors on prostate cancer cells. On the basis of these considerations, prostate cancer remains a great health problem in terms of incidence and prevalence and interventional studies based on the treatment of metabolic syndrome in cancer patients, minimizing exposure to endocrine disruptors, could be a key point in the overall management of this disease.

Role of DNA repair machinery and p53 in the testicular germ cell cancer: a review.

Notwithstanding the peculiar sensitivity to cisplatin-based treatment, resulting in a very high percentage of cures even in advanced stages of the disease, still we do not know the biological mechanisms that make Testicular Germ Cell Tumor (TGCT) "unique" in the oncology scene. p53 and MDM2 seem to play a pivotal role, according to several in vitro observations, but no correlation has been found between their mutational or expression status in tissue samples and patients clinical outcome. Furthermore, other players seem to be on stage: DNA Damage Repair Machinery (DDR) , especially Homologous Recombination (HR) proteins, above all Ataxia Telangiectasia Mutated (ATM), cooperates with p53 in response to DNA damage, activating apoptotic cascade and contributing to cell "fate". Homologous Recombination deficiency has been assumed to be a Germ Cell Tumor characteristic underlying platinum-sensitivity, whereby Poly(ADP-ribose) polymerase (PARP), an enzyme involved in HR DNA repair, is an intriguing target: PARP inhibitors have already entered in clinical practice of other malignancies and trials are recruiting TGCT patients in order to validate their role in this disease. This paper aims to summarize evidence, trying to outline an overview of DDR implications not only in TGCT curability, but also in resistance to chemotherapy.

Integrating role of T antigen, Rb2/p130, CTCF and BORIS in mediating non-canonical endoplasmic reticulum-dependent death pathways triggered by chronic ER stress in mouse medulloblastoma.

Distinct molecular pathways could be constitutively active in mouse T-Antigen positive and T-Antigen negative medulloblastoma cell lines, contributing to their phenotypic differences as well as to cellular responses, cell cycle progression, cell death and survival. The diversity of these responses may be due, at least in part, to distinct activities of Rb2/p130, CTCF and BORIS proteins in response to an altered network of signaling evoked by the T-Ag presence. Here, we provided evidence supporting a role for the T-Antigen in causing chronic endoplasmic reticulum (ER) stress and aberrant Caspase-12 expression and activation, subsequently driving to both massive cell death, and perhaps selection of cells with a higher malignant phenotype. Furthermore, we observed that the endoplasmic stress, either chronically caused by T-Ag or transiently induced by glucose deprivation, is accompanied by the formation of complexes between the retinoblastoma related protein Rb2/p130 and the chromatin insulator CCCTC-binding factor CTCF, or the CTCF-paralogue BORIS. Our study represents the first evidence supporting a role of the T-Antigen in inducing/maintaining chronic ER-stress, as well as, indicating a role of Rb2/p130, CTCF and BORIS as potential mediators of non-canonical ER-dependent death pathway in mouse medulloblastoma.

CTCF and BORIS regulate Rb2/p130 gene transcription: a novel mechanism and a new paradigm for understanding the biology of lung cancer.

Although innumerable investigations regarding the biology of lung cancer have been carried out, many aspects thereof remain to be addressed, including the role played by the retinoblastoma-related protein Rb2/p130 during the evolution of this disease. Here we report novel findings on the mechanisms that control Rb2/p130 gene expression in lung fibroblasts and characterize the effects of Rb2/p130 deregulation on the proliferative features of lung cancer cells. We revealed for the first time that in lung fibroblasts the expression of Rb2/p130 gene is directly controlled by the chromatin insulator CCCTC-binding factor, CTCF, which by binding to the Rb2/p130 gene promoter induces, and/or maintains, a specific local chromatin organization that in turn governs the transcriptional activity of Rb2/p130 gene. However, in lung cancer cells the activity of CTCF in controlling Rb2/p130 gene expression is impaired by BORIS, a CTCF-paralogue, which by binding to the Rb2/p130 gene could trigger changes in the chromatin asset established by CTCF, thereby affecting CTCF regulatory activity on Rb2/p130 transcription. These studies not only provide essential basic insights into the molecular mechanisms that control Rb2/p130 gene expression in lung cancer, but also offer a potential paradigm for the actions of other activators and/or corepressors, such as CTCF and BORIS, that could be crucial in explaining how alterations in the mechanism regulating Rb2/p130 gene expression may accelerate the progression of lung tumors, or favor the onset of recurrence after cancer treatment.

Cross-talk between T-Ag presence and pRb family and p53/p73 signaling in mouse and human medulloblastoma.

The formation and progression of mudulloblastoma (MB) is poorly understood. However, somatic inactivation of pRb/p105, in combination with a somatic or a germ-line TP53 inactivation, leads to MB in a mouse model. Presently, there is no specific evidence of pathway/s alterations for the other two members of the retinoblastoma family, pRb2/p130 and/or p107 in MB. JC virus (JCV) is a human polyomavirus. Although there is no firm evidence that this virus plays a causal role in human neoplasia, it has been clearly proven that JCV is highly oncogenic when injected into the brain of experimental animals. The mechanism of JCV-induced tumorigenesis is not entirely clear. However, several studies relate the oncogenic properties of JCV mainly to its early protein large T-antigen (T-Ag), which is able to bind and inactivate both TP53 and Rb family proteins. Here, we compared the protein expression profiles of p53, p73, pRb family proteins, and PCNA, as main regulators of cell proliferation and death, in different cell lines of mouse primitive neuroectodermal tumors (PNET), either T-Ag-positive or -negative, and in human MB cell lines. Our goal was to determine if changes in the relative expression of these regulators could trigger molecular perturbations underlying MB pathogenesis in mouse and human cells. Our results support that the presence of JCV T-Ag may interfere with the expression of pRb family proteins, specific p73 isoforms, and p53. In turn, this "perturbation" may trigger a network of signals strictly connected with survival and apoptosis.

Epigenetic modulation of estrogen receptor-alpha by pRb family proteins: a novel mechanism in breast cancer.

Estrogen receptor-alpha (ER-alpha) plays a crucial role in normal breast development and has also been linked to mammary carcinogenesis and clinical outcome in breast cancer patients. However, ER-alpha gene expression can change during the course of disease and, consequently, therapy resistance can occur. The molecular mechanism governing ER-alpha transcriptional activity and/or silencing is still unclear. Here, we showed that the presence of a specific pRb2/p130 multimolecular complex on the ER-alpha promoter strongly correlates with the methylation status of this gene. Furthermore, we suggested that pRb2/p130 could cooperate with ICBP90 (inverted CCAAT box binding protein of 90 kDa) and DNA methyltransferases in maintaining a specific methylation pattern of ER-alpha gene. The sequence of epigenetic events for establishing and maintaining the silenced state of ER-alpha gene can be locus- or pathway- specific, and the local remodeling of ER-alpha chromatin structure by pRb2/p130 multimolecular complexes may influence its susceptibility to specific DNA methylation. Our novel hypothesis could provide a basis for understanding how the complex pattern of ER-alpha methylation and transcriptional silencing is generated and for understanding the relationship between this pattern and its function during the neoplastic process.

Modulation of cell cycle components by epigenetic and genetic events.

Cell cycle progression is monitored by surveillance mechanisms, or cell cycle checkpoints, that ensure that initiation of a later event is coupled with the completion of an early cell cycle event. Deregulated proliferation is a characteristic feature of tumor cells. Moreover, defects in many of the molecules that regulate the cell cycle have been implicated in cancer formation and progression. Key among these are p53, the retinoblastoma protein (pRb) and its related proteins, p107 and pRb2/p130, and cdk inhibitors (p15, p16, p18, p19, p21, p27), all of which act to keep the cell cycle from progressing until all repairs to damaged DNA have been completed. The pRb (pRb/p16(INK4a)/cyclin D1) and p53 (p14(ARF)/mdm2/p53) pathways are the two main cell-cycle control pathways frequently targeted in tumorigenesis, and the alterations occurring in each pathway depend on the tumor type. Virtually all human tumors deregulate either the pRb or p53 pathway, and oftentimes both pathways simultaneously. This review focuses on the genetic and epigenetic alterations affecting the components of mechanisms regulating the progression of the cell cycle and leading to cancer formation and progression.

Increased expression of geminin stimulates the growth of mammary epithelial cells and is a frequent event in human tumors.

Geminin is a potent inhibitor of origin assembly and re-replication in multicellular eukaryotes and is a negative regulator of DNA replication during the cell cycle. Thus, it was proposed as an inhibitor of cell proliferation and as a potential tumor suppressor gene. However, the protein was found specifically expressed in proliferating lymphocytes and epithelial cells and up-regulated in several malignancies. Therefore, geminin is now regarded as an oncogene but its role in tumor development remains unknown. In this study, we evaluated by Western blot analysis the expression of geminin in a series of human cancer cell lines of various histogenetic origin and in a series of human primary colon, rectal, and breast cancers. Expression of geminin was variable in different cell lines and not related to the expression level of the corresponding mRNA. Moreover, geminin was expressed at higher level in 56% and 58% of colon and rectal cancers, respectively, compared with the corresponding adjacent normal mucosa. A high expression of geminin was also detected by immunohistochemistry in 60% of human primary breast cancers. We also transfected a full-length geminin cDNA in a human non-tumorigenic and a cancer breast cell lines and obtained derivatives expressing high levels of the protein. Geminin overexpression stimulated cell cycle progression and proliferation in both normal and cancer cells and increased the anchorage--independent growth of breast cancer cells. These results demonstrate that expression of geminin is frequently deregulated in tumor cells and might play an important role in the regulation of cell growth in both normal and malignant cells.

Regulatory role of BMP2 and BMP7 in spermatogonia and Sertoli cell proliferation in the immature mouse.

AIM: The aim of this study was to determine the action of bone morphogenetic proteins (BMPs) on testicular cell proliferation during early postnatal life, a definite developmental time at which crucial changes in germ cell and Sertoli cell maturation occur. METHODS: We investigated the effect of BMP2 and BMP7, two factors which belong to the relatively distant decapentaplegic (DPP) and 60 A classes of the large BMP family, upon spermatogonial and Sertoli cell proliferation, and we examined the expression of activin/BMP type II and type I receptors. We used in vitro cultured testis fragments from 7-day-old mice, highly purified populations of somatic and germ cells and total testes from mice of different ages. Cell proliferation was assessed by BrdU labelling and [3H]-thymidine incorporation. Ribonuclease protection assays and Northern blotting were performed to analyse receptor expression. RESULTS AND CONCLUSIONS: We have demonstrated a stimulatory action of BMP2 and BMP7 in spermatogonia and Sertoli cell proliferation respectively. ActRIIB is the type II receptor expressed most in spermatogonia, whereas Sertoli cells specifically expressed BMPRIIB, in addition to ActRIIB. By contrast, the presence of ActRIIA was undetectable in either germ or somatic cells. The type I receptors ActRIA, ActRIB and BMPRIA were all found in both cell types, indicating that the observed effect of BMP2 and BMP7 on testicular cell proliferation may be mediated by a number of combinatorial interactions in the receptor complexes. These findings suggest that BMPs are involved in physiological paracrine signalling during the first wave of spermatogenesis.

Increased expression of dystroglycan inhibits the growth and tumorigenicity of human mammary epithelial cells.

Dystroglycan (DG) is an adhesion molecule formed by two subunits, alpha (extracellular) and beta (transmembrane) DG, which are codified by a single gene and form a continuous link from the extracellular matrix to the intracellular cytoskeleton. Reduction or loss of expression of DG has been observed in human cancer cell lines and primary tumors and has been suggested to promote tumor development and invasiveness. In this study, the human breast epithelial non-tumorigenic MCF10F and the breast cancer MCF7 cell lines were engineered to stably express an exogenous DG cDNA and the effects on the phenotype of both cell lines were evaluated. The MCF10F transfected cells displayed an increased expression of both DG subunits which was associated with inhibition of the anchorage-dependent growth, accumulation of cells in the G0/G1 phase of the cell cycle and increased adhesion to a substratum. The MCF7 transfected cells were unable to restore alpha-DG despite an increased expression of the beta-DG subunit. Anchorage-dependent and independent growth and the in vivo tumorigenicity were reduced in these derivatives that also displayed a reduced adhesion to a substratum and were shown to release alpha-DG in the culture medium. These findings confirm and extend previous evidence that transformation of mammary epithelial cells is associated with loss of their ability to retain alpha-DG on the cell membrane. Moreover, they indicate that DG is involved in cell functions other than cell adhesion to the extracellular matrix, and that its loss of function might predispose to tumor progression by compromising regulatory controls over cell growth and proliferation.

Dystroglycan expression is frequently reduced in human breast and colon cancers and is associated with tumor progression.

Dystroglycan (DG) is an adhesion molecule responsible for crucial interactions between extracellular matrix and cytoplasmic compartment. It is formed by two subunits, alpha-DG (extracellular) and beta-DG (transmembrane), that bind to laminin in the matrix and dystrophin in the cytoskeleton, respectively. In this study we evaluated by Western blot analysis the expression of DG in a series of human cancer cell lines of various histogenetic origin and in a series of human primary colon and breast cancers. Decreased expression of DG was observed in most of the cell lines and in both types of tumors and correlated with higher tumor grade and stage. Analysis of the mRNA levels suggested that expression of DG protein is likely regulated at a posttranscriptional level. Evaluation of alpha-DG expression by immunostaining in a series of archival cases of primary breast carcinomas confirmed that alpha-DG expression is lost in a significant fraction of tumors (66%). Loss of DG staining correlated with higher tumor stage (P = 0.022), positivity for p53 (P = 0.033), and high proliferation index (P = 0.045). A significant correlation was also observed between loss of alpha-DG and overall survival (P = 0.013 by log-rank test) in an univariate analysis. These data indicate that DG expression is frequently lost in human malignancies and suggest that this glycoprotein might play an important role in human tumor development and progression.

Oxidative DNA damage as a marker of aging in WI-38 human fibroblasts.

Cause-effect relationships between oxidative stress, DNA damage and aging were investigated in WI-38 human diploid fibroblasts at 21, 41 or 58 population doublings (PDs), corresponding to young, middle age or old fibroblasts, respectively. Oxidative DNA damage was evaluated by immunohistochemical detection of 8-hydroxy-2'deoxyguanosine (8-OHdG) adducts or by single cell microgel electrophoresis (COMET assay). Aging was evaluated by growth rate, senescence-associated-beta-galactosidase (SA-beta galactosidase) activity, cell cycle distribution, and expression of p21. Our results demonstrate that (i) oxidative DNA damage is proportional to the age of cells (ii) DNA damage in old/58 PDs cells reflects both an increased susceptibility to oxidative stress, induced by acute exposure to sub-lethal concentrations of hydrogen peroxide (H(2)O(2)), and a reduced efficiency of repair mechanisms. We also show that mild chronic oxidative stress, induced by prolonged exposure to 5 microM H(2)O(2), accelerates aging in fibroblasts. In fact, this treatment increased 8-OHdG levels, SA-beta-galactosidase activity, and G0/G1 cell cycle arrest in middle age/41 PDs, making them similar to H(2)O(2)-untreated old/58 PDs cells. Although other mechanisms may concur in mediating the effects of H(2)O(2), these results lend support to the concept that oxidative stress may be a key determinant of aging. Measurements of oxidative DNA damage might therefore be exploited as reliable marker of cellular aging.