bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from Intakes of Red Meat, Processed Meat, and Meat Mutagens Increase Lung Cancer Risk
Authors: Lam Tram Kim , Cross Amanda J. , Consonni Dario , Randi Giorgia , Bagnardi Vincenzo , Bertazzi Pier Alberto , Caporaso Neil E. , Sinha Rashmi , Subar Amy F. , Landi Maria Teresa
Published: March 2023

Abstract
Red and processed meat intake may increase lung cancer risk. However, the epidemiologic evidence is inconsistent and few studies have evaluated the role of meat mutagens formed during high cooking temperatures. We investigated the association of red meat, processed meat, and meat mutagen intake with lung cancer risk in Environment And Genetics in Lung cancer Etiology, a population-based case-control study. Primary lung cancer cases (n = 2,101) were recruited from 13 hospitals within the Lombardy region of Italy examining ∼80% of the cases from the area. Noncancer population controls (n = 2,120), matched to cases on gender, residence, and age, were randomly selected from the same catchment area. Diet was assessed in 1,903 cases and 2,073 controls and used in conjunction with a meat mutagen database to estimate intake of heterocyclic amines (HCA) and benzo(a)pyrene (BaP). Multivariable odds ratios (OR) and 95% confidence intervals (95% CI) for sex-specific tertiles of intake were calculated using unconditional logistic regression. Red and processed meat were positively associated with lung cancer risk (highest-versus-lowest tertile: OR, 1.8; 95% CI, 1.5–2.2; P trend < 0.001 and OR, 1.7; 95% CI, 1.4–2.1; P trend < 0.001, respectively); the risks were strongest among never smokers (OR, 2.4; 95% CI, 1.4–4.0; P trend = 0.001 and OR, 2.5; 95% CI, 1.5–4.2; P trend = 0.001, respectively). HCAs and BaP were significantly associated with increased risk of lung cancer. When separated by histology, significant positive associations for both meat groups were restricted to adenocarcinoma and squamous cell carcinoma but not small cell carcinoma of the lung. In summary, red meat, processed meat, and meat mutagens were independently associated with increased risk of lung cancer. [Cancer Res 2009;69(3):932–9]
Data from Quantitative Phosphoproteomics Reveals a Cluster of Tyrosine Kinases That Mediates Src Invasive Activity in Advanced Colon Carcinoma Cells
Authors: Leroy Cédric , Fialin Camille , Sirvent Audrey , Simon Valérie , Urbach Serge , Poncet Joël , Robert Bruno , Jouin Patrick , Roche Serge
Published: March 2023

Abstract
The nonreceptor tyrosine kinase Src is frequently overexpressed and/or activated in human colorectal carcinoma (CRC), and its increased activity has been associated with a poor clinical outcome. Src has been implicated in growth and invasion of these cancer cells by still not well-known mechanisms. Here, we addressed Src oncogenic signaling using quantitative phosphoproteomics. Src overexpression increased growth and invasiveness of metastatic SW620 CRC cells. Stable isotope labeling with amino acids in cell culture in combination with liquid chromatography tandem mass spectrometry allowed the identification of 136 proteins which exhibited a significant increase in and/or association with tyrosine phosphorylation upon Src expression. These mainly include signaling, cytoskeleton, and vesicular-associated proteins. Interestingly, Src also phosphorylated a cluster of tyrosine kinases, i.e., the receptors Met and EphA2, the cytoplasmic tyrosine kinase Fak, and pseudo-tyrosine kinase SgK223, which were required for its invasive activity. Similar results were obtained with metastatic Colo205 CRC cells that exhibit high endogenous Src activity. We concluded that Src uses a tyrosine kinases network to promote its invasive activity in CRC and this implicates a reverse signaling via tyrosine kinase receptors. Targeting these tyrosine kinases may be of significant therapeutic value in this cancer. [Cancer Res 2009;69(6):2279–86]
Data from Hypoxia-Inducible Factor-1α Suppresses Squamous Carcinogenic Progression and Epithelial-Mesenchymal Transition
Authors: Scortegagna Marzia , Martin Rebecca J. , Kladney Raleigh D. , Neumann Robert G. , Arbeit Jeffrey M.
Published: March 2023

Abstract
Hypoxia-inducible factor-1 (HIF-1) is a known cancer progression factor, promoting growth, spread, and metastasis. However, in selected contexts, HIF-1 is a tumor suppressor coordinating hypoxic cell cycle suppression and apoptosis. Prior studies focused on HIF-1 function in established malignancy; however, little is known about its role during the entire process of carcinogenesis from neoplasia induction to malignancy. Here, we tested HIF-1 gain of function during multistage murine skin chemical carcinogenesis in K14-HIF-1αPro402A564G (K14-HIF-1αDPM) transgenic mice. Transgenic papillomas appeared earlier and were more numerous (6 ± 3 transgenic versus 2 ± 1.5 nontransgenic papillomas per mouse), yet they were more differentiated, their proliferation was lower, and their malignant conversion was profoundly inhibited (7% in transgenic versus 40% in nontransgenic mice). Moreover, transgenic cancers maintained squamous differentiation whereas epithelial-mesenchymal transformation was frequent in nontransgenic malignancies. Transgenic basal keratinocytes up-regulated the HIF-1 target N-myc downstream regulated gene-1, a known tumor suppressor gene in human malignancy, and its expression was maintained in transgenic papillomas and cancer. We also discovered a novel HIF-1 target gene, selenium binding protein-1 (Selenbp1), a gene of unknown function whose expression is lost in human cancer. Thus, HIF-1 can function as a tumor suppressor through transactivation of genes that are themselves targets for negative selection in human cancers. [Cancer Res 2009;69(6):2638–46]
Data from Identification of Smyd4 as a Potential Tumor Suppressor Gene Involved in Breast Cancer Development
Authors: Hu Liping , Zhu Yiwei Tony , Qi Chao , Zhu Yi-Jun
Published: March 2023

Abstract
To identify genes involved in breast tumorigenesis, we applied the retroviral LoxP-Cre system to a nontumorigenic mouse mammary epithelial cell line NOG8 to create random chromosome deletion/translocation. We found that the disruption of one allele of Smyd4 (SET and MYND domain containing 4) gene through chromosome translocation led to tumorigenesis. The expression of Smyd4 was markedly decreased in tumor cells. Re-expression of Smyd4 resulted in growth suppression of tumor cells and inhibition of tumor formation in nude mice. Furthermore, the RNA interference–mediated suppression of Smyd4 expression in human MCF10A mammary epithelial cells caused their growth in soft agar. Microarray studies revealed that platelet-derived growth factor receptor α polypeptide (Pdgfr-α) was highly expressed in tumor cells compared with NOG8 cells. Re-expression of Smyd4 significantly reduced the expression of Pdgfr-α in tumor cells. In human breast cancers, reverse transcription-PCR results revealed that Smyd4 expression was totally silenced in 2 of 10 specimens. These findings indicate that Smyd4, as a potential tumor suppressor, plays a critical role in breast carcinogenesis at least partly through inhibiting the expression of Pdgfr-α, and could be a novel target for improving treatment of breast cancer. [Cancer Res 2009;69(9):4067–72]
Data from Inhibition of Constitutively Activated Nuclear Factor-κB Induces Reactive Oxygen Species- and Iron-Dependent Cell Death in Cutaneous T-Cell Lymphoma
Authors: Kiessling Michael K. , Klemke Claus D. , Kamiński Marcin M. , Galani Ioanna E. , Krammer Peter H. , Gülow Karsten
Published: March 2023

Abstract
Aberrant signaling of the nuclear facotr (NF-κB) pathway has been identified as a mediator of survival and apoptosis resistance in leukemias and lymphomas. Here, we report that cell death of cutaneous T-cell lymphoma cell lines induced by inhibition of the NF-κB pathway is independent of caspases or classic death receptors. We found that free intracellular iron and reactive oxygen species (ROS) are the main mediators of this cell death. Antioxidants such as N-Acetyl-l-cysteine and glutathione or the iron chelator desferrioxamine effectively block cell death in cutaneous T-cell lymphoma cell lines or primary T cells from Sézary patients. We show that inhibition of constitutively active NF-κB causes down-regulation of ferritin heavy chain (FHC) that leads to an increase of free intracellular iron, which, in turn, induces massive generation of ROS. Furthermore, direct down-regulation of FHC by siRNA caused a ROS-dependent cell death. Finally, high concentrations of ROS induce cell death of malignant T cells. In contrast, T cells isolated from healthy donors do not display down-regulation of FHC and, therefore, do not show an increase in iron and cell death upon NF-κB inhibition. In addition, in a murine T-cell lymphoma model, we show that inhibition of NF-κB and subsequent down-regulation of FHC significantly delays tumor growth in vivo. Thus, our results promote FHC as a potential target for effective therapy in lymphomas with aberrant NF-κB signaling. [Cancer Res 2009;69(6):2365–74]
Data from Adiponectin-Activated AMPK Stimulates Dephosphorylation of AKT through Protein Phosphatase 2A Activation
Authors: Kim Kun-yong , Baek Ahmi , Hwang Ji-Eun , Choi Yeon A. , Jeong Joon , Lee Myeong-Sok , Cho Dea Ho , Lim Jong-Seok , Kim Keun Il , Yang Young
Published: March 2023

Abstract
Low serum levels of adiponectin are a high risk factor for various types of cancer. Although adiponectin inhibits proliferation and metastasis of breast cancer cells, the underlying molecular mechanisms remain obscure. In this study, we show that adiponectin-activated AMPK reduces the invasiveness of MDA-MB-231 cells by stimulating dephosphorylation of AKT by increasing protein phosphatase 2A (PP2A) activity. Among the various regulatory B56 subunits, B56γ was directly phosphorylated by AMPK at Ser298 and Ser336, leading to an increase of PP2A activity through dephosphorylation of PP2Ac at Tyr307. We also show that both the blood levels of adiponectin and the tissue levels of PP2A activity were decreased in breast cancer patients and that the direct administration of adiponectin into tumor tissues stimulates PP2A activity. Taken together, these findings show that adiponectin, derived from adipocytes, negatively regulates the invasiveness of breast cancer cells by activating the tumor suppressor PP2A. [Cancer Res 2009;69(9):4018–26]
Data from Inhibition of Peroxisome Proliferator-Activated Receptor γ Increases Estrogen Receptor–Dependent Tumor Specification
Authors: Yin Yuzhi , Yuan Hongyan , Zeng Xiao , Kopelovich Levy , Glazer Robert I.
Published: March 2023

Abstract
Peroxisome proliferator-activated receptor γ (PPARγ) is a nuclear receptor that regulates gene transcription associated with intermediary metabolism, adipocyte differentiation, and tumor suppression and proliferation. To understand the role of PPARγ in tumorigenesis, transgenic mice were generated with mammary gland–directed expression of the dominant-negative transgene Pax8PPARγ. Transgenic mice were phenotypically indistinguishable from wild-type (WT) mice, but mammary epithelial cells expressed a greater percentage of CD29hi/CD24neg, CK5+, and double-positive CK14/CK18 cells. These changes correlated with reduced PTEN and increased Ras and extracellular signal-regulated kinase (ERK) and AKT activation. Although spontaneous tumorigenesis did not occur, transgenic animals were highly susceptible to progestin/7,12-dimethylbenz(a)anthracene–induced mammary carcinogenesis, which in contrast to WT mice resulted in a high tumor multiplicity and, most importantly, in the appearance of predominantly estrogen receptor α–positive (ER+) ductal adenocarcinomas. Tumors expressed a similar PTENlo/pERKhi/pAKThi phenotype as mammary epithelium and exhibited high activation of estrogen response element–dependent reporter gene activity. Tumorigenesis in MMTV-Pax8PPARγ mice was insensitive to the chemopreventive effect of a PPARγ agonist but was profoundly inhibited by the ER antagonist fulvestrant. These results reveal important new insights into the previously unrecognized role of PPARγ in the specification of mammary lineage and the development of ER+ tumors. [Cancer Res 2009;69(2):687–94]
Data from Thyroid Transcription Factor-1 Inhibits Transforming Growth Factor-β–Mediated Epithelial-to-Mesenchymal Transition in Lung Adenocarcinoma Cells
Authors: Saito Roy-Akira , Watabe Tetsuro , Horiguchi Kana , Kohyama Tadashi , Saitoh Masao , Nagase Takahide , Miyazono Kohei
Published: March 2023

Abstract
Thyroid transcription factor-1 (TTF-1) is expressed in lung cancer, but its functional roles remain unexplored. TTF-1 gene amplification has been discovered in a part of lung adenocarcinomas, and its action as a lineage-specific oncogene is highlighted. Epithelial-to-mesenchymal transition (EMT) is a crucial event for cancer cells to acquire invasive and metastatic phenotypes and can be elicited by transforming growth factor-β (TGF-β). Mesenchymal-to-epithelial transition (MET) is the inverse process of EMT; however, signals that induce MET are largely unknown. Here, we report a novel functional aspect of TTF-1 that inhibits TGF-β–mediated EMT and restores epithelial phenotype in lung adenocarcinoma cells. This effect was accompanied by down-regulation of TGF-β target genes, including presumed regulators of EMT, such as Snail and Slug. Moreover, silencing of TTF-1 enhanced TGF-β–mediated EMT. Thus, TTF-1 can exert a tumor-suppressive effect with abrogation of cellular response to TGF-β and attenuated invasive capacity. We further revealed that TTF-1 down-regulates TGF-β2 production in A549 cells and that TGF-β conversely decreases endogenous TTF-1 expression, suggesting that enhancement of autocrine TGF-β signaling accelerates the decrease of TTF-1 expression and vice versa. These findings delineate potential links between TTF-1 and TGF-β signaling in lung cancer progression through regulation of EMT and MET and suggest that modulation of TTF-1 expression can be a novel therapeutic strategy for treatment of lung adenocarcinoma. [Cancer Res 2009;69(7):2783–91]
Data from αvβ3/αvβ5 Integrins-FAK-RhoB: A Novel Pathway for Hypoxia Regulation in Glioblastoma
Authors: Skuli Nicolas , Monferran Sylvie , Delmas Caroline , Favre Gilles , Bonnet Jacques , Toulas Christine , Cohen-Jonathan Moyal Elizabeth
Published: March 2023

Abstract
The presence of hypoxic areas in glioblastoma is an important determinant in tumor response to therapy and, in particular, to radiotherapy. Here we have explored the involvement of integrins, up to now known as regulators of angiogenesis and invasion, in the regulation of tumor hypoxia driven from the tumor cell. We first show that hypoxia induces the recruitment of αvβ3 and αvβ5 integrins to the cellular membrane of U87 and SF763 glioblastoma cells, thereby activating the focal adhesion kinase (FAK). We then show that inhibiting αvβ3 or αvβ5 integrins in hypoxic cells with a specific inhibitor or with siRNA decreases the hypoxia-inducible factor 1α (HIF-1α) intracellular level. This integrin-dependent regulation of HIF-1α is mediated through the regulation of FAK, which in turn activates the small GTPase RhoB, leading to the inhibition of GSK3-β. Furthermore, silencing this pathway in glioma cells of established xenografts dramatically reduces glioma hypoxia, associated with a significant decrease in vessel density. Our present results unravel a new mechanism of hypoxia regulation by establishing the existence of an αvβ3/αvβ5 integrin–dependent loop of hypoxia autoregulation in glioma. Targeting this hypoxia loop may be crucial to optimizing radiotherapy efficiency. [Cancer Res 2009;69(8):3308–16]
Data from Dual Inhibition of Class IA Phosphatidylinositol 3-Kinase and Mammalian Target of Rapamycin as a New Therapeutic Option for T-Cell Acute Lymphoblastic Leukemia
Authors: Chiarini Francesca , Falà Federica , Tazzari Pier Luigi , Ricci Francesca , Astolfi Annalisa , Pession Andrea , Pagliaro Pasqualepaolo , McCubrey James A. , Martelli Alberto M.
Published: March 2023

Abstract
Recent investigations have documented that constitutively activated phosphatidylinositol 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) signaling is a common feature of T-cell acute lymphoblastic leukemia (T-ALL), where it strongly influences growth and survival. These findings lend compelling weight for the application of PI3K/Akt/mTOR inhibitors in T-ALL. However, our knowledge of PI3K/Akt/mTOR signaling in T-ALL is limited and it is not clear whether it could be an effective target for innovative therapeutic strategies. Here, we have analyzed the therapeutic potential of the dual PI3K/mTOR inhibitor PI-103, a small synthetic molecule of the pyridofuropyrimidine class, on both T-ALL cell lines and patient samples, which displayed constitutive activation of PI3K/Akt/mTOR signaling. PI-103 inhibited the growth of T-ALL cells, including 170-kDa P-glycoprotein overexpressing cells. PI-103 cytotoxicity was independent of p53 gene status. PI-103 was more potent than inhibitors that are selective only for PI3K (Wortmannin, LY294002) or for mTOR (rapamycin). PI-103 induced G0-G1 phase cell cycle arrest and apoptosis, which was characterized by activation of caspase-3 and caspase-9. PI-103 caused Akt dephosphorylation, accompanied by dephosphorylation of the Akt downstream target, glycogen synthase kinase-3β. Also, mTOR downstream targets were dephosphorylated in response to PI-103, including p70S6 kinase, ribosomal S6 protein, and 4E-BP1. PI-103 strongly synergized with vincristine. These findings indicate that multitargeted therapy toward PI3K and mTOR alone or with existing drugs may serve as an efficient treatment toward T-ALL cells, which require up-regulation of PI3K/Akt/mTOR signaling for their survival and growth. [Cancer Res 2009;69(8):3520–28]
Data from Genome-Wide Impact of Androgen Receptor Trapped clone-27 Loss on Androgen-Regulated Transcription in Prostate Cancer Cells
Authors: Nwachukwu Jerome C. , Mita Paolo , Ruoff Rachel , Ha Susan , Wang Qianben , Huang S. Joseph , Taneja Samir S. , Brown Myles , Gerald William L. , Garabedian Michael J. , Logan Susan K.
Published: March 2023

Abstract
The androgen receptor (AR) directs diverse biological processes through interaction with coregulators such as AR trapped clone-27 (ART-27). Our results show that ART-27 is recruited to AR-binding sites by chromatin immunoprecipitation analysis. In addition, the effect of ART-27 on genome-wide transcription was examined. The studies indicate that loss of ART-27 enhances expression of many androgen-regulated genes, suggesting that ART-27 inhibits gene expression. Surprisingly, classes of genes that are up-regulated upon ART-27 depletion include regulators of DNA damage checkpoint and cell cycle progression, suggesting that ART-27 functions to keep expression levels of these genes low. Consistent with this idea, stable reduction of ART-27 by short-hairpin RNA enhances LNCaP cell proliferation compared with control cells. The effect of ART-27 loss was also examined in response to the antiandrogen bicalutamide. Unexpectedly, cells treated with ART-27 siRNA no longer exhibited gene repression in response to bicalutamide. To examine ART-27 loss in prostate cancer progression, immunohistochemistry was conducted on a tissue array containing samples from primary tumors of individuals who were clinically followed and later shown to have either recurrent or nonrecurrent disease. Comparison of ART-27 and AR staining indicated that nuclear ART-27 expression was lost in the majority of AR-positive recurrent prostate cancers. Our studies show that reduction of ART-27 protein levels in prostate cancer may facilitate antiandrogen-resistant disease. [Cancer Res 2009;69(7):3140–7]
Data from Integral Role of Transcription Factor 8 in the Negative Regulation of Tumor Angiogenesis
Authors: Inuzuka Takayuki , Tsuda Masumi , Tanaka Shinya , Kawaguchi Hideaki , Higashi Yujiro , Ohba Yusuke
Published: March 2023

Abstract
Angiogenesis is involved in various physiologic and pathological conditions, including tumor growth, and is tightly regulated by the orchestration of proangiogenic and antiangiogenic factors. Inhibition of vascular endothelial growth factor (VEGF), the best-established antiangiogenic treatment in cancer, has shown some effectiveness; however, the identification of novel regulators, whose function is independent of VEGF, is required to achieve better outcomes. Here, we show that transcription factor 8 (TCF8) is up-regulated in endothelial cells during angiogenesis, acting as a negative regulator. Furthermore, TCF8 is specifically expressed in the endothelium of tumor vessels. Tcf8-heterozygous knockout mice are more permissive than wild-type mice to the formation of tumor blood vessels in s.c. implanted melanoma, which seems to contribute to the more aggressive growth and the lung metastases of the tumor in mutant mice. Suppression of TCF8 facilitates angiogenesis in both in vitro and ex vivo models, and displays comprehensive cellular phenotypes, including enhanced cell invasion, impaired cell adhesion, and increased cell monolayer permeability due to, at least partly, MMP1 overexpression, attenuation of focal adhesion formation, and insufficient VE-cadherin recruitment, respectively. Taken together, our findings define a novel, integral role for TCF8 in the regulation of pathologic angiogenesis, and propose TCF8 as a target for therapeutic intervention in cancer. [Cancer Res 2009;69(4):1678–84]
Data from Activation of Meiosis-Specific Genes Is Associated with Depolyploidization of Human Tumor Cells following Radiation-Induced Mitotic Catastrophe
Authors: Ianzini Fiorenza , Kosmacek Elizabeth A. , Nelson Elke S. , Napoli Eleonora , Erenpreisa Jekaterina , Kalejs Martins , Mackey Michael A.
Published: March 2023

Abstract
Cancer is frequently characterized histologically by the appearance of large cells that are either aneuploid or polyploid. Aneuploidy and polyploidy are hallmarks of radiation-induced mitotic catastrophe (MC), a common phenomenon occurring in tumor cells with impaired p53 function following exposure to various cytotoxic and genotoxic agents. MC is characterized by altered expression of mitotic regulators, untimely and abnormal cell division, delayed DNA damage, and changes in morphology. We report here that cells undergoing radiation-induced MC are more plastic with regards to ploidy and that this plasticity allows them to reorganize their genetic material through reduction division to produce smaller cells which are morphologically indistinguishable from control cells. Experiments conducted with the large-scale digital cell analysis system are discussed and show that a small fraction of polyploid cancer cells formed via radiation-induced MC can survive and start a process of depolyploidization that yields various outcomes. Although most multipolar divisions failed and cell fusion occurred, some of these divisions were successful and originated a variety of cell progeny characterized by different ploidy. Among these ploidy phenotypes, a progeny of small mononucleated cells, indistinguishable from the untreated control cells, is often seen. We report here evidence that meiosis-specific genes are expressed in the polyploid cells during depolyploidization. Tumor cells might take advantage of the temporary change from a promitotic to a promeiotic division regimen to facilitate depolyploidization and restore the proliferative state of the tumor cell population. These events might be mechanisms by which tumor progression and resistance to treatment occur in vivo. [Cancer Res 2009;69(6):2296–304]
Data from Heterozygosity for Hypoxia Inducible Factor 1α Decreases the Incidence of Thymic Lymphomas in a p53 Mutant Mouse Model
Authors: Bertout Jessica A. , Patel Shetal A. , Fryer Benjamin H. , Durham Amy C. , Covello Kelly L. , Olive Kenneth P. , Goldschmidt Michael H. , Simon M. Celeste
Published: March 2023

Abstract
Hypoxia inducible factors (HIF) are critical mediators of the cellular response to decreased oxygen tension and are overexpressed in a number of tumors. Although HIF1α and HIF2α share a high degree of sequence homology, recent work has shown that the two α subunits can have contrasting and tissue-specific effects on tumor growth. To directly compare the role of each HIFα subunit in spontaneous tumorigenesis, we bred a mouse model of expanded HIF2α expression and Hif1α+/− mice to homozygotes for the R270H mutation in p53. Here, we report that p53R270H/R270H mice, which have not been previously described, develop a unique tumor spectrum relative to p53R270H/− mice, including a high incidence of thymic lymphomas. Heterozygosity for Hif1α significantly reduced the incidence of thymic lymphomas observed in this model. Moreover, reduced Hif1α levels correlated with decreased stabilization of activated Notch1 and expression of the Notch target genes, Dtx1 and Nrarp. These observations uncover a novel role for HIF1α in Notch pathway activation during T-cell lymphomagenesis. [Cancer Res 2009;69(7):3213–20]
Data from Endothelin-1 Stimulates Lymphatic Endothelial Cells and Lymphatic Vessels to Grow and Invade
Authors: Spinella Francesca , Garrafa Emirena , Di Castro Valeriana , Rosanò Laura , Nicotra Maria Rita , Caruso Arnaldo , Natali Pier Giorgio , Bagnato Anna
Published: March 2023

Abstract
The lymphatic vasculature is essential for tissue fluid homeostasis and cancer metastasis, although the molecular mechanisms involved remain poorly characterized. Endothelin-1 (ET-1) axis plays a crucial role in angiogenesis and tumorigenesis. Here, we first report that ET-1 acts as a lymphangiogenic mediator. We performed in vitro and in vivo studies and show that lymphatic endothelial cells produce ET-1, ET-3, and express the endothelin B receptor (ETBR). In these cells, ET-1 promotes proliferation, invasiveness, vascular-like structures formation, and phosphorylation of AKT and p42/44 mitogen-activated protein kinase through ETBR. In normoxic conditions, ET-1 is also able to up-regulate the expression of vascular endothelial growth factor (VEGF)-C, VEGF receptor-3, and VEGF-A, and to stimulate hypoxia-inducible factor (HIF)-1α expression similarly to hypoxia. Moreover, HIF-1α silencing by siRNA desensitizes VEGF-C and VEGF-A production in response to ET-1 or hypoxia, implicating HIF-1α/VEGF as downstream signaling molecules of ET-1 axis. Double immunofluorescence analysis of human lymph nodes reveals that lymphatic vessels express ETBR together with the lymphatic marker podoplanin. Furthermore, a Matrigel plug assay shows that ET-1 promotes the outgrowth of lymphatic vessels in vivo. ETBR blockade with the specific antagonist, BQ788, inhibits in vitro and in vivo ET-1–induced effects, demonstrating that ET-1 through ETBR directly regulates lymphatic vessel formation and by interacting with the HIF-1α–dependent machinery can amplify the VEGF-mediated lymphatic vascularization. Our results suggest that ET-1 axis is indeed a new player in lymphangiogenesis and that targeting pharmacologically ETBR and related signaling cascade may be therapeutically exploited in a variety of diseases including cancer. [Cancer Res 2009;69(6):2669–76]
Data from Protein Kinase Cβ Is an Effective Target for Chemoprevention of Colon Cancer
Authors: Fields Alan P. , Calcagno Shelly R. , Krishna Murli , Rak Sofija , Leitges Michael , Murray Nicole R.
Published: March 2023

Abstract
Colon cancer develops over a period of 10 to 15 years, providing a window of opportunity for chemoprevention and early intervention. However, few molecular targets for effective colon cancer chemoprevention have been characterized and validated. Protein kinase CβII (PKCβII) plays a requisite role in the initiation of colon carcinogenesis in a preclinical mouse model by promoting proliferation and increased β-catenin accumulation. In this study, we test the hypothesis that PKCβII is an effective target for colon cancer chemoprevention using enzastaurin (LY317615), a PKCβ-selective inhibitor, in a mouse model of colon carcinogenesis. We find that enzastaurin potently reduces azoxymethane-induced colon tumor initiation and progression by inhibiting PKCβII-mediated tumor cell proliferation and β-catenin accumulation. Biochemically, enzastaurin reduces expression of the PKCβII- and β-catenin/T-cell factor–regulated genes PKCβII, cyclooxygenase II, and vascular endothelial growth factor, three genes implicated in colon carcinogenesis. Our results show that enzastaurin is an effective chemopreventive agent in a mouse model of sporadic colon cancer that significantly reduces both tumor initiation and progression by inhibiting expression of proproliferative genes. Thus, PKCβII is an important target for colon cancer chemoprevention and the PKCβ-selective inhibitor enzastaurin may represent an effective chemopreventive agent in patients at high risk for colon cancer. [Cancer Res 2009;69(4):1643–50]
Data from Central Role of c-Myc during Malignant Conversion in Human Hepatocarcinogenesis
Authors: Kaposi-Novak Pal , Libbrecht Louis , Woo Hyun Goo , Lee Yun-Han , Sears Nathaniel C. , Conner Elizabeth A. , Factor Valentina M. , Roskams Tania , Thorgeirsson Snorri S.
Published: March 2023

Abstract
Hepatocarcinogenesis is a multistage process in which precursor lesions progress into early hepatocellular carcinomas (eHCC) by sequential accumulation of multiple genetic and epigenetic alterations. To decode the molecular events during early stages of liver carcinogenesis, we performed gene expression profiling on cirrhotic (regenerative) and dysplastic nodules (DN), as well as eHCC. Although considerable heterogeneity was observed at the regenerative and dysplastic stages, overall, 460 differentially expressed genes were detected between DN and eHCC. Functional analysis of the significant gene set identified the MYC oncogene as a plausible driver gene for malignant conversion of the DNs. In addition, gene set enrichment analysis revealed global activation of the MYC up-regulated gene set in eHCC versus dysplasia. Presence of the MYC signature significantly correlated with increased expression of CSN5, as well as with higher overall transcription rate of genes located in the 8q chromosome region. Furthermore, a classifier constructed from MYC target genes could robustly discriminate eHCC from high-grade and low-grade DNs. In conclusion, our study identified unique expression patterns associated with the transition of high-grade DNs into eHCC and showed that activation of the MYC transcription signature is strongly associated with the malignant conversion of preneoplastic liver lesions. [Cancer Res 2009;69(7):2775–82]
Data from microRNA-205 Regulates HER3 in Human Breast Cancer
Authors: Iorio Marilena V. , Casalini Patrizia , Piovan Claudia , Di Leva Gianpiero , Merlo Andrea , Triulzi Tiziana , Ménard Sylvie , Croce Carlo M. , Tagliabue Elda
Published: March 2023

Abstract
An increasing amount of experimental evidence shows that microRNAs can have a causal role in breast cancer tumorigenesis as a novel class of oncogenes or tumor suppressor genes, depending on the targets they regulate. HER2 overexpression is a hallmark of a particularly aggressive subset of breast tumors, and its activation is strictly dependent on the trans-interaction with other members of HER family; in particular, the activation of the PI3K/Akt survival pathway, so critically important in tumorigenesis, is predominantly driven through phosphorylation of the kinase-inactive member HER3. Here, we show that miR-205, down-modulated in breast tumors compared with normal breast tissue, directly targets HER3 receptor, and inhibits the activation of the downstream mediator Akt. The reintroduction of miR-205 in SKBr3 cells inhibits their clonogenic potential and increases the responsiveness to tyrosine-kinase inhibitors Gefitinib and Lapatinib, abrogating the HER3-mediated resistance and restoring a potent proapoptotic activity. Our data describe miR-205 as a new oncosuppressor gene in breast cancer, able to interfere with the proliferative pathway mediated by HER receptor family. Our study also provides experimental evidence suggesting that miR-205 can improve the responsiveness to specific anticancer therapies. [Cancer Res 2009;69(6):2195–200]
Data from The EGF/CSF-1 Paracrine Invasion Loop Can Be Triggered by Heregulin β1 and CXCL12
Authors: Hernandez Lorena , Smirnova Tatiana , Kedrin Dmitriy , Wyckoff Jeffrey , Zhu Liyin , Stanley E. Richard , Cox Dianne , Muller William J. , Pollard Jeffrey W. , Van Rooijen Nico , Segall Jeffrey E.
Published: March 2023

Abstract
An important step in the process of metastasis from the primary tumor is invasive spread into the surrounding stroma. Using an in vivo invasion assay, we have previously shown that imposed gradients of epidermal growth factor (EGF) or colony-stimulating factor-1 (CSF-1) can induce invasion through an EGF/CSF-1 paracrine loop between cancer cells and macrophages. We now report that invasion induced by other ligands also relies on this EGF/CSF-1 paracrine invasive loop. Using an in vivo invasion assay, we show that MTLn3 breast cancer cells overexpressing ErbB3 exhibit enhanced invasion compared with control MTLn3 cells in response to the ErbB3 ligand HRG-β1. The invasive response of both MTLn3-ErbB3 and transgenic MMTV-Neu tumors to HRG-β1 is inhibited by blocking EGF receptor, CSF-1 receptor, or macrophage function, indicating that invasiveness to HRG-β1 is dependent on the EGF/CSF-1 paracrine loop. Furthermore, we show that CXCL12 also triggers in vivo invasion of transgenic MMTV-PyMT tumors in an EGF/CSF-1–dependent manner. Although the invasion induced by HRG-β1 or CXCL12 is dependent on the EGF/CSF-1 paracrine loop, invasion induced by EGF is not dependent on HRG-β1 or CXCL12 signaling, showing an asymmetrical relationship between different ligand/receptor systems in driving invasion. Our results identify a stromal/tumor interaction that acts as an engine underlying invasion induced by multiple ligands. [Cancer Res 2009;69(7):3221–7]
Data from Deletion of PTEN Promotes Tumorigenic Signaling, Resistance to Anoikis, and Altered Response to Chemotherapeutic Agents in Human Mammary Epithelial Cells
Authors: Vitolo Michele I. , Weiss Michele B. , Szmacinski Marta , Tahir Khola , Waldman Todd , Park Ben Ho , Martin Stuart S. , Weber David J. , Bachman Kurtis E.
Published: March 2023

Abstract
Many cancers, including breast cancer, harbor loss-of-function mutations in the catalytic domain of phosphatase and tensin homologue deleted on chromosome 10 (PTEN) or have reduced PTEN expression through loss of heterozygosity and/or epigenetic silencing mechanisms. However, specific phenotypic effects of PTEN inactivation in human cancer cells remain poorly defined without a direct causal connection between the loss of PTEN function and the development or progression of cancer. To evaluate the biological and clinical relevance of reduced or deleted PTEN expression, a novel in vitro model system was generated using human somatic cell knockout technologies. Targeted homologous recombination allowed for a single and double allelic deletion, which resulted in reduced and deleted PTEN expression, respectively. We determined that heterozygous loss of PTEN in the nontumorigenic human mammary epithelial cell line MCF-10A was sufficient for activation of the phosphoinositide 3-kinase/AKT and mitogen-activated protein kinase pathways, whereas the homozygous absence of PTEN expression led to a further increased activation of both pathways. The deletion of PTEN was able to confer growth factor–independent proliferation, which was confirmed by the resistance of the PTEN−/− MCF-10A cells to small-molecule inhibitors of the epidermal growth factor receptor. However, neither heterozygous nor homozygous loss of PTEN expression was sufficient to promote anchorage-independent growth, but the loss of PTEN did confer apoptotic resistance to cell rounding and matrix detachment. Finally, MCF-10A cells with the reduction or loss of PTEN showed increased susceptibility to the chemotherapeutic drug doxorubicin but not paclitaxel. [Cancer Res 2009;69(21):8275–83]
Data from Endothelial Nitric Oxide Synthase Mediates Lymphangiogenesis and Lymphatic Metastasis
Authors: Lahdenranta Johanna , Hagendoorn Jeroen , Padera Timothy P. , Hoshida Tohru , Nelson Gregory , Kashiwagi Satoshi , Jain Rakesh K. , Fukumura Dai
Published: March 2023

Abstract
Lymphatic metastasis is a critical determinant of cancer prognosis. Recently, several lymphangiogenic molecules such as vascular endothelial growth factor (VEGF)-C and VEGF-D were identified. However, the mechanistic understanding of lymphatic metastasis is still in infancy. Nitric oxide (NO) plays a crucial role in regulating blood vessel growth and function as well as lymphatic vessel function. NO synthase (NOS) expression correlates with lymphatic metastasis. However, causal relationship between NOS and lymphatic metastasis has not been documented. To this end, we first show that both VEGF receptor-2 and VEGF receptor-3 stimulation activate eNOS in lymphatic endothelial cells and that NO donors induce proliferation and/or survival of cultured lymphatic endothelial cells in a dose-dependent manner. We find that an NOS inhibitor, L-NMMA, blocked regeneration of lymphatic vessels. Using intravital microscopy that allows us to visualize the steps of lymphatic metastasis, we show that genetic deletion of eNOS as well as NOS blockade attenuates peritumor lymphatic hyperplasia of VEGF-C–overexpressing T241 fibrosarcomas and decreases the delivery of metastatic tumor cells to the draining lymph nodes. Genetic deletion of eNOS in the host also leads to a decrease in T241 tumor cell dissemination to the lymph nodes and macroscopic lymph node metastasis of B16F10 melanoma. These findings indicate that eNOS mediates VEGF-C–induced lymphangiogenesis and, consequently, plays a critical role in lymphatic metastasis. Our findings explain the correlation between NOS and lymphatic metastasis seen in a number of human tumors and open the door for potential therapies exploiting NO signaling to treat diseases of the lymphatic system. [Cancer Res 2009;69(7):2801–8]
Data from Smad Ubiquitination Regulatory Factor 2 Promotes Metastasis of Breast Cancer Cells by Enhancing Migration and Invasiveness
Authors: Jin Chaoyang , Yang Yu-an , Anver Miriam R. , Morris Nicole , Wang Xiangchun , Zhang Ying E.
Published: March 2023

Abstract
Controlled protein degradation mediated by ubiquitin/proteasome system (UPS) plays a crucial role in modulating a broad range of cellular responses. Dysregulation of the UPS often accompanies tumorigenesis and progression. Here, we report that Smad ubiquitination regulatory factor 2 (Smurf2), a HECT-domain containing E3 ubiquitin ligase, is up-regulated in certain breast cancer tissues and cells. We show that reduction of Smurf2 expression with specific short interfering RNA in metastatic breast cancer cells induces cell rounding and reorganization of the actin cytoskeleton, which are associated with a less motile and invasive phenotype. Overexpression of Smurf2 promotes metastasis in a nude mouse model and increases migration and invasion of breast cancer cells. Moreover, expression of Smurf2CG, an E3 ligase–defective mutant of Smurf2, suppresses the above metastatic behaviors. These results establish an important role for Smurf2 in breast cancer progression and indicate that Smurf2 is a novel regulator of breast cancer cell migration and invasion. [Cancer Res 2009;69(3):735–40]
Data from miR-205 Exerts Tumor-Suppressive Functions in Human Prostate through Down-regulation of Protein Kinase Cε
Authors: Gandellini Paolo , Folini Marco , Longoni Nicole , Pennati Marzia , Binda Mara , Colecchia Maurizio , Salvioni Roberto , Supino Rosanna , Moretti Roberta , Limonta Patrizia , Valdagni Riccardo , Daidone Maria Grazia , Zaffaroni Nadia
Published: March 2023

Abstract
Limited information is available concerning the expression and role of microRNAs in prostate cancer. In this study, we investigated the involvement of miR-205 in prostate carcinogenesis. Significantly lower miR-205 expression levels were found in cancer than in normal prostate cell lines as well as in tumor compared with matched normal prostate tissues, with a particularly pronounced reduction in carcinomas from patients with local-regionally disseminated disease. Restoring the expression of miR-205 in prostate cancer cells resulted in cell rearrangements consistent with a mesenchymal-to-epithelial transition, such as up-regulation of E-cadherin and reduction of cell locomotion and invasion, and in the down-regulation of several oncogenes known to be involved in disease progression (i.e., interleukin 6, caveolin-1, EZH2). Our evidence suggests that these events are driven by the concurrent repression of specific predicted miR-205 targets, namely N-chimaerin, ErbB3, E2F1, E2F5, ZEB2, and protein kinase Cε. Strikingly, the latter seemed to play a direct role in regulating epithelial-to-mesenchymal transition. In fact, its down-regulation led to a cell phenotype largely reminiscent of that of cells ectopically expressing miR-205. Overall, we showed for the first time that miR-205 exerts a tumor-suppressive effect in human prostate by counteracting epithelial-to-mesenchymal transition and reducing cell migration/invasion, at least in part through the down-regulation of protein kinase Cε. [Cancer Res 2009;69(6):2287–95]
Data from Anticancer Chemotherapy Inhibits MHC Class I–Related Chain A Ectodomain Shedding by Downregulating ADAM10 Expression in Hepatocellular Carcinoma
Authors: Kohga Keisuke , Takehara Tetsuo , Tatsumi Tomohide , Miyagi Takuya , Ishida Hisashi , Ohkawa Kazuyoshi , Kanto Tatsuya , Hiramatsu Naoki , Hayashi Norio
Published: March 2023

Abstract
MHC class I–related chain A (MICA) is a ligand for the NKG2D-activating immunoreceptor that mediates activation of natural killer (NK) cells. The ectodomain of MICA is shed from tumor cells, which may be an important means of evading antitumor immunity. We previously reported that patients with hepatocellular carcinoma (HCC) display high levels of soluble MICA in circulation, which could be downregulated by chemotherapy. The present study shows that anti-HCC drugs suppress MICA ectodomain shedding by inhibiting expression of a disintegrin and metalloproteinase 10 (ADAM10). Both ADAM10 and CD44, a typical substrate of the ADAM10 protease, were expressed in human HCC tissues and HCC cells but not in normal liver tissues or cultured hepatocytes. Small interfering RNA–mediated knockdown experiments revealed that ADAM10 is a critical sheddase for both MICA and CD44 in HCC cells. Of interest is the finding that epirubicin clearly downregulated ADAM10 expression and MICA shedding in HCC cells; its suppressive effect on MICA shedding was abolished in ADAM10-depleted cells. Epirubicin treatment also enhanced the NKG2D-mediated NK sensitivity of HCC cells. Patients with HCC had significantly higher levels of serum-soluble CD44, which correlated well with serum-soluble MICA levels, thus suggesting a close link between ADAM10 activity and MICA shedding in these patients. Soluble MICA and CD44 levels were downregulated with a significant correlation in patients treated by transarterial chemoembolization using epirubicin. In conclusion, anticancer drugs can modulate expression of ADAM10, which is critically involved in MICA ectodomain shedding. Epirubicin therapy may have a previously unrecognized effect on antitumor immunity in HCC patients. [Cancer Res 2009;69(20):8050–7]
Data from Effective Immunotherapy against Murine Gliomas Using Type 1 Polarizing Dendritic Cells—Significant Roles of CXCL10
Authors: Fujita Mitsugu , Zhu Xinmei , Ueda Ryo , Sasaki Kotaro , Kohanbash Gary , Kastenhuber Edward R. , McDonald Heather A. , Gibson Gregory A. , Watkins Simon C. , Muthuswamy Ravikumar , Kalinski Pawel , Okada Hideho
Published: March 2023

Abstract
In an attempt to develop effective vaccines against central nervous system (CNS) tumors, we evaluated the ability of vaccines with standard dendritic cells (DC) versus type 1 polarizing DCs (DC1) to induce glioma-specific type 1 CTLs with CNS tumor-relevant homing properties and the mechanism of their action. C57BL/6 mouse–derived bone marrow cells were cultured with mouse granulocyte/macrophage colony-stimulating factor (GM-CSF) for 6 days, and CD11c+ cells were subsequently cultured with GM-CSF, rmIFN-γ, rmIFN-α, rmIL-4, and polyinosinic-polycytidylic acid stabilized by lysine and carboxymethylcellulose for 24 hours to generate DC1s. In analogy to their human counterparts, mouse DC1s exhibited surface marker profiles of mature DCs and produced high levels of IL-12 and CXCL10. Importantly for their application as cancer vaccines, such DC1s stably retained their type 1 phenotype even when exposed to type 2–promoting or regulatory T cell (Treg)–promoting environments. Consistently, mouse DC1s induced antigen-specific type 1 CTLs more efficiently than nonpolarized DCs in vitro. DC1s given s.c. migrated into draining lymph nodes, induced antigen-specific CTLs, and suppressed Treg accumulation. In addition, s.c. immunization with DC1s loaded with glioma-associated antigen (GAA)–derived CTL epitope peptides prolonged the survival of CNS GL261 glioma-bearing mice, which was associated with efficient CNS glioma homing of antigen-specific CTLs. Intratumoral injections of GAA peptide-loaded DC1s further enhanced the anti-CNS glioma effects of DC1-based s.c. immunization. Interestingly, the antitumor functions were abrogated with CXCL10−/− mouse–derived DC1s. Collectively, these findings show the anti-CNS glioma effects of DC1-based therapy and a novel role of CXCL10 in the immunologic and therapeutic activity of DC-based cancer vaccines. [Cancer Res 2009;69(4):1587–95]
Data from Targeting Fibroblast Growth Factor Receptors Blocks PI3K/AKT Signaling, Induces Apoptosis, and Impairs Mammary Tumor Outgrowth and Metastasis
Authors: Dey Julien H. , Bianchi Fabrizio , Voshol Johannes , Bonenfant Debora , Oakeley Edward J. , Hynes Nancy E.
Published: March 2023

Abstract
Members of the fibroblast growth factor receptor (FGFR) family have essential roles in normal physiology and in cancer where they control diverse processes. FGFRs have been associated with breast cancer development. Thus, models to study the role of FGFR in breast cancer and their targeting potential are important. We present an in vitro and in vivo analysis of FGFRs in the breast cancer model cell lines 67NR and 4T1. We show that both tumor cell lines coexpress FGFRs and ligands and display autocrine FGFR signaling activity. Fibroblast growth factor receptor substrate 2 (FRS2), a downstream mediator of FGFR, is constitutively tyrosine phosphorylated and multiple signaling pathways are active. Treatment of 67NR and 4T1 cultures with TKI258, an FGFR tyrosine kinase inhibitor (TKI), caused a rapid decrease in FRS2 phosphorylation; decreased the activity of extracellular signal–regulated kinase 1/2 (ERK1/2), AKT, and phospholipase Cγ; and blocked proliferation of both tumor lines. Furthermore, TKI258 induced 4T1 apoptotic cell death via blockade of the phosphoinositide 3-kinase/AKT pathway. In vivo, one dose of TKI258 rapidly lowered FRS2 phosphorylation and ERK1/2 and AKT activity in mammary tumors. Long-term TKI258 treatment of 4T1 tumor– and 67NR tumor–bearing mice had a significant effect on primary tumor outgrowth and 4T1 tumor–induced lung metastases. A microarray analysis was carried out to identify targets with roles in TKI258 antitumor activity and potential prognostic markers in human breast tumors. Of interest are the downregulated matrix metalloproteases (MMP), in particular MMP9, which is essential for metastatic spread of 4T1 tumors. Cancer Res; 70(10); 4151–62. ©2010 AACR.