bluuebunny/crossref_metadata_2025 huggingface dataset:
Small-molecule inhibitor of USP7/HAUSP ubiquitin protease stabilizes and activates p53 in cells
Authors: Colland Frédéric , Formstecher Etienne , Jacq Xavier , Reverdy Céline , Planquette Cécile , Conrath Susan , Trouplin Virginie , Bianchi Julie , Aushev Vasily N. , Camonis Jacques , Calabrese Alessandra , Borg-Capra Catherine , Sippl Wolfgang , Collura Vincent , Boissy Guillaume , Rain Jean-Christophe , Guedat Philippe , Delansorne Rémi , Daviet Laurent
Published: August 2009

Deregulation of the ubiquitin/proteasome system has been implicated in the pathogenesis of many human diseases, including cancer. Ubiquitin-specific proteases (USP) are cysteine proteases involved in the deubiquitination of protein substrates. Functional connections between USP7 and essential viral proteins and oncogenic pathways, such as the p53/Mdm2 and phosphatidylinositol 3-kinase/protein kinase B networks, strongly suggest that the targeting of USP7 with small-molecule inhibitors may be useful for the treatment of cancers and viral diseases. Using high-throughput screening, we have discovered HBX 41,108, a small-molecule compound that inhibits USP7 deubiquitinating activity with an IC50 in the submicromolar range. Kinetics data indicate an uncompetitive reversible inhibition mechanism. HBX 41,108 was shown to affect USP7-mediated p53 deubiquitination in vitro and in cells. As RNA interference-mediated USP7 silencing in cancer cells, HBX 41,108 treatment stabilized p53, activated the transcription of a p53 target gene without inducing genotoxic stress, and inhibited cancer cell growth. Finally, HBX 41,108 induced p53-dependent apoptosis as shown in p53 wild-type and null isogenic cancer cell lines. We thus report the identification of the first lead-like inhibitor against USP7, providing a structural basis for the development of new anticancer drugs.[Mol Cancer Ther 2009;8(8):2286–95]
Xenogeneic immunization with human tyrosine hydroxylase DNA vaccines suppresses growth of established neuroblastoma
Authors: Huebener Nicole , Fest Stefan , Hilt Kerstin , Schramm Alexander , Eggert Angelika , Durmus Tahir , Woehler Anja , Stermann Alexander , Bleeke Matthias , Baykan Bianca , Weixler Silke , Gaedicke Gerhard , Lode Holger N.
Published: August 2009

Neuroblastoma (NB) is a challenging malignancy of the sympathetic nervous tissue characterized by a very poor prognosis. One important marker for NB is the expression of tyrosine hydroxylase (TH), the first-step enzyme of catecholamine biosynthesis. We could show stable and high TH gene expression in 67 NB samples independent of the clinical stage. Based on this observation, we addressed the question of whether xenogeneic TH DNA vaccination is effective in inducing an anti-NB immune response. For this purpose, we generated three DNA vaccines based on pCMV-F3Ub and pBUD-CE4.1 plasmids encoding for human (h)THcDNA (A), hTH minigene (B), and hTHcDNA in combination with the proinflammatory cytokine interleukin 12 (C), and tested prophylactic and therapeutic efficacy to suppress primary tumor growth and spontaneous metastasis. Here we report that xenogeneic TH DNA vaccination was effective in eradicating established primary tumors and inhibiting metastasis. Interestingly, this effect could not be enhanced by adding the Th1 cytokine interleukin 12. However, increased IFN-γ production and NB cytotoxicity of effector cells harvested from vaccinated mice suggested the participation of tumor-specific CTLs in the immune response. The depletion of CD8+T cells completely abrogated the hTH vaccine–mediated anti-NB immune response. Furthermore, rechallenging of surviving mice resulted in reduced primary tumor growth, indicating the induction of a memory immune response. In conclusion, xenogeneic immunization with TH-derived DNA vaccines is effective against NB, and may open a new venue for a novel and effective immunotherapeutic strategy against this challenging childhood tumor. [Mol Cancer Ther 2009;8(8):2392–401]
HYD1-induced increase in reactive oxygen species leads to autophagy and necrotic cell death in multiple myeloma cells
Authors: Nair Rajesh R. , Emmons Michael F. , Cress Anne E. , Argilagos Raul F. , Lam Kit , Kerr William T. , Wang Hong-Gong , Dalton William S. , Hazlehurst Lori A.
Published: August 2009

HYD1 is a D–amino acid peptide that was previously shown to inhibit adhesion of prostate cancer cells to the extracellular matrix. In this study, we show that in addition to inhibiting adhesion of multiple myeloma (MM) cells to fibronectin, HYD1 induces cell death in MM cells as a single agent. HYD1-induced cell death was necrotic in nature as shown by: (a) decrease in mitochondrial membrane potential (Δψm), (b) loss of total cellular ATP, and (c) increase in reactive oxygen species (ROS) production. Moreover, HYD1 treatment does not result in apoptotic cell death because it did not trigger the activation of caspases or the release of apoptosis-inducing factor and endonuclease G from the mitochondria, nor did it induce double-stranded DNA breaks. HYD1 did initiate autophagy in cells; however, autophagy was found to be an adaptive response contributing to cell survival rather than the cause of cell death. We were further able to show that N-acetyl-L-cysteine, a thiol-containing free radical scavenger, partially protects MM cells from HYD1-induced death. Additionally, N-acetyl-L-cysteine blocked HYD1-induced as well as basal levels of autophagy, suggesting that ROS can potentially trigger both cell death and cell survival pathways. Taken together, our data describe an important role of ROS in HYD1-induced necrotic cell death in MM cells. [Mol Cancer Ther 2009;8(8):2441–51]
Pharmacologic inhibitors of IκB kinase suppress growth and migration of mammary carcinosarcoma cellsin vitroand prevent osteolytic bone metastasisin vivo
Authors: Idris Aymen I. , Libouban Hélène , Nyangoga Hervé , Landao-Bassonga Euphemie , Chappard Daniel , Ralston Stuart H.
Published: August 2009

The NF-κB signaling pathway is known to play an important role in the regulation of osteoclastic bone resorption and cancer cell growth. Previous studies have shown that genetic inactivation of IκB kinase (IKK), a key component of NF-κB signaling, inhibits osteoclastogenesis, but the effects of pharmacologic IKK inhibitors on osteolytic bone metastasis are unknown. Here, we studied the effects of the IKK inhibitors celastrol, BMS-345541, parthenolide, and wedelolactone on the proliferation and migration of W256 cells in vitro and osteolytic bone destruction in vivo. All compounds tested inhibited the growth and induced apoptosis of W256 cells as evidenced by caspase-3 activation and nuclear morphology. Celastrol, BMS-345541, and parthenolide abolished IL1β and tumor necrosis factor α–induced IκB phosphorylation and prevented nuclear translocation of NF-κB and DNA binding. Celastrol and parthenolide but not BMS-345541 prevented the activation of both IKKα and IKKβ, and celastrol inhibited IKKα/β activation by preventing the phosphorylation of TAK1, a key receptor–associated factor upstream of IKK. Celastrol and parthenolide markedly reduced the mRNA expression of matrix metalloproteinase 9 and urinary plasminogen activator, and inhibited W256 migration. Administration of celastrol or parthenolide at a dose of 1 mg/kg/day suppressed trabecular bone loss and reduced the number and size of osteolytic bone lesions following W256 injection in rats. Histomorphometric analysis showed that both compounds decreased osteoclast number and inhibited bone resorption. In conclusion, pharmacologic inhibitors of IKK are effective in preventing osteolytic bone metastasis in this model and might represent a promising class of agents to the prevention and treatment of metastatic bone disease associated with breast cancer. [Mol Cancer Ther 2009;8(8):2339–47]
Inhibition of hypoxia-inducible factor-1 function enhances the sensitivity of multiple myeloma cells to melphalan
Authors: Hu Yongzhen , Kirito Keita , Yoshida Kozue , Mitsumori Toru , Nakajima Kei , Nozaki Yumi , Hamanaka Satoshi , Nagashima Takahiro , Kunitama Masae , Sakoe Kumi , Komatsu Norio
Published: August 2009

Abnormal activation of hypoxia-inducible factor-1 (HIF-1), one of the most important transcription factors for the adaptation of cells to hypoxia, is frequently observed in numerous types of solid tumors. Dysregulation of HIF-1 induces tumor angiogenesis and enhances the expression of anti-apoptotic proteins and glycolysis-associated enzymes in cancer cells, which in turn leads to the promotion of tumor growth. In the present study, we examined the pathophysiologic role of HIF-1 in multiple myeloma. Furthermore, we explored the possibility that HIF-1 may be a molecular target for myeloma therapy. We identified constitutive expression of the hypoxia-inducible factor-1 α (HIF-1α)-subunit in established myeloma cell lines and in primary myeloma cells. Treatment with insulin-like growth factor-1 (IGF-1) significantly increased HIF-1α expression through activation of the AKT and mitogen-activated protein kinase signaling pathways. Inhibition of HIF-1 function either by echinomycin, a specific HIF-1 inhibitor, or a siRNA against HIF-1α resulted in enhanced sensitivity to melphalan in myeloma cells. This inhibition of HIF-1 also reversed the protective effect of IGF-1 on melphalan-induced apoptosis. Inhibition of HIF-1 drastically reduced both basal and IGF-1–induced expression of survivin, one of the most important anti-apoptotic proteins in myeloma cells. We conclude that HIF-1 inhibition may be an attractive therapeutic strategy for multiple myeloma. [Mol Cancer Ther 2009;8(8):2329–38]
Targeting Migration inducting gene-7 inhibits carcinoma cell invasion, early primary tumor growth, and stimulates monocyte oncolytic activity
Authors: Petty Aaron P. , Wright Stephen E. , Rewers-Felkins Kathleen A. , Yenderrozos Michelle A. , Vorderstrasse Beth A. , Lindsey J. Suzanne
Published: August 2009

Expression of Migration inducting gene-7 (Mig-7) is limited to tumor cells and to date not found in normal tissues. Multiple tumor microenvironment factors, such as epidermal and hepatocyte growth factors, in concert with αvβ5 integrin ligation, induce Mig-7 mRNA expression. Gain or loss of Mig-7 protein studies shows that Mig-7 promotes invasion of colon and endometrial carcinoma cells. These data led us to hypothesize that targeting Mig-7 through various methods could decrease invasion, enhance monocyte cell killing of tumor cells, and inhibit disease progression. To begin testing this hypothesis, an in vitro chemoinvasion assay of endometrial carcinoma cells treated with Mig-7–specific or control antibodies was used. Mig-7 antibody significantly reduced invasion by >60% compared with controls. In another approach to test this hypothesis, an in vitro analysis of peptide-stimulated human peripheral blood monocyte cells and their killing of MCF-7 breast carcinoma cells was used. Mig-7 peptide treatment increased monocyte cell tumor necrosis factor expression and killing of MCF-7 cells 30-fold over no peptide stimulation and 3-fold over MUC-1 or control peptide treatments. Furthermore, stably expressing Mig-7–specific short hairpin RNA resulted in significantly reduced Mig-7 protein levels and early primary tumor growth in a xenograft nude mouse model. Reduced phosphorylation of ERK1/2, Akt, and S6 kinase as well as decreased membrane-type 1 matrix metalloproteinase activity were mechanisms through which Mig-7 protein caused these effects. Based on these collective data, Mig-7 expression could be a potential candidate for future targeted cancer therapies. [Mol Cancer Ther 2009;8(8):2412–23]
Gene expression signatures and response to imatinib mesylate in gastrointestinal stromal tumor
Authors: Rink Lori , Skorobogatko Yuliya , Kossenkov Andrew V. , Belinsky Martin G. , Pajak Thomas , Heinrich Michael C. , Blanke Charles D. , von Mehren Margaret , Ochs Michael F. , Eisenberg Burton , Godwin Andrew K.
Published: August 2009

Despite initial efficacy of imatinib mesylate in most gastrointestinal stromal tumor (GIST) patients, many experience primary/secondary drug resistance. Therefore, clinical management of GIST may benefit from further molecular characterization of tumors before and after imatinib mesylate treatment. As part of a recent phase II trial of neoadjuvant/adjuvant imatinib mesylate treatment for advanced primary and recurrent operable GISTs (Radiation Therapy Oncology Group S0132), gene expression profiling using oligonucleotide microarrays was done on tumor samples obtained before and after imatinib mesylate therapy. Patients were classified according to changes in tumor size after treatment based on computed tomography scan measurements. Gene profiling data were evaluated with Statistical Analysis of Microarrays to identify differentially expressed genes (in pretreatment GIST samples). Based on Statistical Analysis of Microarrays [False Discovery Rate (FDR), 10%], 38 genes were expressed at significantly lower levels in the pretreatment biopsy samples from tumors that significantly responded to 8 to 12 weeks of imatinib mesylate, that is, >25% tumor reduction. Eighteen of these genes encoded Krüppel-associated box (KRAB) domain containing zinc finger (ZNF) transcriptional repressors. Importantly, 10 KRAB-ZNF genes mapped to a single locus on chromosome 19p, and a subset predicted likely response to imatinib mesylate–based therapy in a naïve panel of GIST. Furthermore, we found that modifying expression of genes within this predictive signature can enhance the sensitivity of GIST cells to imatinib mesylate. Using clinical pretreatment biopsy samples from a prospective neoadjuvant phase II trial, we have identified a gene signature that includes KRAB-ZNF 91 subfamily members that may be both predictive of and functionally associated with likely response to short-term imatinib mesylate treatment. [Mol Cancer Ther 2009;8(8):2172–82]
Wnt/Planar cell polarity signaling: A new paradigm for cancer therapy
Authors: Wang Yingqun
Published: August 2009

The evolutionarily conserved and developmentally important Wnt signaling pathway has traditionally been regarded as a critical player in tumorigenesis through the canonical Wnt/β-catenin cascade. Nevertheless, accumulating evidence based on recent research has revealed the previously unacknowledged role of noncanonical Wnt/planar cell polarity (PCP) signaling in cancer progression, invasion and metastasis, and angiogenesis. This review describes the PCP signaling pathway and its ever-expanding components and modulators, highlights the most recent studies that provide insight into the link between PCP signaling and cancer, and, finally, proposes a model by which PCP signaling may promote cancer development. This review underscores the emerging theme that deregulated PCP signaling contributes to tumorigenesis, providing new potential targets for cancer therapy. [Mol Cancer Ther 2009;8(8):2103–9]
Induction of endoplasmic reticulum stress response by TZD18, a novel dual ligand for peroxisome proliferator-activated receptor α/γ, in human breast cancer cells
Authors: Zang Chuanbing , Liu Hongyu , Bertz Janina , Possinger Kurt , Koeffler H. Phillip , Elstner Elena , Eucker Jan
Published: August 2009

Previously we reported that the peroxisome proliferator-activated receptor α/γ dual ligand TZD18 inhibited growth and induced apoptosis of leukemia and glioblastoma cells. Now we show that TZD18 also has the same effects against six human breast cancer cell lines. To obtain insights into the mechanism involved in TZD18-induced growth inhibition and apoptosis in breast cancer, the gene expression profiles of TZD18-treated and untreated MCF-7 and MDA-MB-231 cells were compared by microarray analysis. Results reveal that many genes implicated in endoplasmic reticulum stress signaling, such as CHOP (also known as DDIT3 or GADD153), GRP78 (HSPA5), and ATF4, are highly up-regulated, suggesting endoplasmic reticulum stress is induced. This is supported by our data that treatment of MCF-7 and MDA-MB-231 cells with TZD18 induces phosphorylation of PERK and the α subunit of eukaryotic initiation factor 2 (eIF2α), as well as an up-regulation of GRP78 and an activation of ATF6, all of which are specific markers for endoplasmic reticulum stress. Furthermore, this ligand increases the endoplasmic reticulum stress–related cell death–regulators such as CHOP, DR5, GADD34, Bax, and Bak in these cells. Importantly, knockdown of CHOP by small interference RNA antagonizes the TZD18-induced apoptosis, indicating a crucial role of CHOP in the apoptotic process triggered by TZD18. In addition, TZD18 also activates stress-sensitive mitogen-activated protein kinase (MAPK) pathways including p38, ERK, and JNK. The specific inhibitors of these MAPKs attenuated the TZD18-induced growth inhibition in these cells. These results clearly show that activation of these MAPKs is important for TZD18-induced growth inhibition. In summary, TZD18-treatment leads to the activation of endoplasmic reticulum stress response and, subsequently, growth arrest and apoptosis in breast cancer cells. [Mol Cancer Ther 2009;8(8):2296–307]
Oncolytic adenoviruses targeted to cancer stem cells
Authors: Short Joshua J. , Curiel David T.
Published: August 2009

Cancer stem cells (CSC) represent a distinct subpopulation of cancer cells of integral importance. CSCs embody the refractory nature observed among many cancers: very competent initial tumor establishment and extremely aggressive metastatic nature. Recent discoveries indicate that CSCs embody chemo- and radioresistance and have been correlated with advanced disease and resistance to current therapies, and thus help explain the treatment resistance of many cancers. As CSCs are critical for tumor initiation, progression, persistence, and the development of metastasis, the success or failure of treatment approaches may be influenced greatly by the presence and treatment sensitivity of these cells. There also seems to be a direct link between epithelial-mesenchymal transition phenomena and CSCs. Cancer cure is predicated upon effectively targeting and eradicating the CSC population. Oncolytic viruses have undergone many developments and through multiple generations offer an effective way to specifically target and eradicate CSCs, while still maintaining the ability to affect the general tumor cell population. Conditionally replicative adenoviruses (CRAd) are one virotherapy that is especially promising. Multiple advanced targeting and infectivity enhancement schemes have been developed to allow the necessary specificity and transduction efficiency required for an effective therapy. Furthermore, these advanced generation CRAds can be armed with therapeutic transgenes to generate greater antitumor effects. Although ultimately, the rewards of targeting and eradicating CSCs will be evaluated in clinical trials, there are numerous methods for isolating primary CSCs based on surface marker expression and multiple established cell lines representative of CSCs for preliminary evaluation.[Mol Cancer Ther 2009;8(8):2096–102]
Molecular predictors of response to a humanized anti–insulin-like growth factor-I receptor monoclonal antibody in breast and colorectal cancer
Authors: Zha Jiping , O'Brien Carol , Savage Heidi , Huw Ling-Yuh , Zhong Fiona , Berry Leanne , Lewis Phillips Gail D. , Luis Elizabeth , Cavet Guy , Hu Xiaolan , Amler Lukas C. , Lackner Mark R.
Published: August 2009

The insulin-like growth factor-I receptor (IGF-IR) pathway is required for the maintenance of the transformed phenotype in neoplastic cells and hence has been the subject of intensive drug discovery efforts. A key aspect of successful clinical development of targeted therapies directed against IGF-IR will be identification of responsive patient populations. Toward that end, we have endeavored to identify predictive biomarkers of response to an anti-IGF-IR-targeting monoclonal antibody in preclinical models of breast and colorectal cancer. We find that levels of the IGF-IR itself may have predictive value in these tumor types and identify other gene expression predictors of in vitro response. Studies in breast cancer models suggest that IGF-IR expression is both correlated and functionally linked with estrogen receptor signaling and provide a basis for both patient stratification and rational combination therapy with antiestrogen-targeting agents. In addition, we find that levels of other components of the signaling pathway such as the adaptor proteins IRS1 and IRS2, as well as the ligand IGF-II, have predictive value and report on the development of a pathway-focused panel of diagnostic biomarkers that could be used to test these hypotheses during clinical development of IGF-IR-targeting therapies. [Mol Cancer Ther 2009;8(8):2110–21]
Activity of dasatinib againstL576P KITmutant melanoma: Molecular, cellular, and clinical correlates
Authors: Woodman Scott E. , Trent Jonathan C. , Stemke-Hale Katherine , Lazar Alexander J. , Pricl Sabrina , Pavan Giovanni M. , Fermeglia Maurizio , Gopal Y.N. Vashisht , Yang Dan , Podoloff Donald A. , Ivan Doina , Kim Kevin B. , Papadopoulos Nicholas , Hwu Patrick , Mills Gordon B. , Davies Michael A.
Published: August 2009

Point mutations in the KIT receptor tyrosine kinase gene have recently been identified in mucosal, acral lentiginous, and chronically sun-damaged melanomas. We have identified the first human melanoma cell line with an endogenous L576P mutation, the most common KIT mutation in melanoma (∼30-40%). In vitro testing showed that the cell viability of the L576P mutant cell line was not reduced by imatinib, nilotinib, or sorafenib small molecule KIT inhibitors effective in nonmelanoma cells with other KIT mutations. However, the viability of the mutant cells was reduced by dasatinib at concentrations as low as 10 nM (P = 0.004). Molecular modeling studies found that the L576P mutation induces structural changes in KIT that reduce the affinity for imatinib (ΔΔGbind = −2.52 kcal/mol) but not for dasatinib (ΔΔGbind = +0.32 kcal/mol). Two metastatic melanoma patients with the L576P KIT mutation were treated with dasatinib, including one patient previously treated with imatinib. Both patients had marked reduction (>50%) and elimination of tumor F18-fluorodeoxyglucose (FDG)-avidity by positron emission tomography (PET) imaging after dasatinib treatment. These data support the selective inhibitory effect of dasatinib against cells harboring the most common KIT mutation in melanoma, and thus has therapeutic implications for acrallentiginous, chronic sun-damaged, and mucosal melanomas. [Mol Cancer Ther 2009;8(8):2079–85]
Regulation of Bcl-2 Expression by HuR in HL60 Leukemia Cells and A431 Carcinoma Cells
Authors: Ishimaru Daniella , Ramalingam Sivakumar , Sengupta Tapas K. , Bandyopadhyay Sumita , Dellis Stephanie , Tholanikunnel Baby G. , Fernandes Daniel J. , Spicer Eleanor K.
Published: August 2009

Overexpression of the proto-oncogene bcl-2 promotes abnormal cell survival by inhibiting apoptosis. Expression of bcl-2 is determined, in part, by regulatory mechanisms that control the stability of bcl-2 mRNA. Elements in the 3′-untranslated region of bcl-2 mRNA have been shown to play a role in regulating the stability of the message. Previously, it was found that the RNA binding proteins nucleolin and Ebp1 have a role in stabilizing bcl-2 mRNA in HL60 cells. Here, we have identified HuR as a component of bcl-2 messenger ribonucleoprotein (mRNP) complexes. RNA coimmunoprecipitation assays showed that HuR binds to bcl-2 mRNA in vivo. We also observed an RNA-dependent coprecipitation of HuR and nucleolin, suggesting that the two proteins are present in common mRNP complexes. Moreover, nucleolin and HuR bind concurrently to bcl-2 AU-rich element (ARE) RNA in vitro, suggesting separate binding sites for these proteins on bcl-2 mRNA. Knockdown of HuR in A431 cells leads to down-regulation of bcl-2 mRNA and protein levels. Observation of a decreased ratio of bcl-2 mRNA to heterogeneous nuclear RNA in HuR knockdown cells confirmed a positive role for HuR in regulating bcl-2 stability. Recombinant HuR retards exosome-mediated decay of bcl-2 ARE RNA in extracts of HL60 cells. This supports a role for HuR in the regulation of bcl-2 mRNA stability in HL60 cells, as well as in A431 cells. Addition of nucleolin and HuR to HL60 cell extracts produced a synergistic protective effect on decay of bcl-2 ARE RNA. HuR knockdown also leads to redistribution of bcl-2 mRNA from polysomes to monosomes. Thus, HuR seems to play a positive role in both regulation of bcl-2 mRNA translation and mRNA stability. (Mol Cancer Res 2009;7(8):1354–66)
miR-93, miR-98, and miR-197 Regulate Expression of Tumor Suppressor Gene FUS1
Authors: Du Liqin , Schageman Jeoffrey J. , Subauste Maria C. , Saber Barbara , Hammond Scott M. , Prudkin Ludmila , Wistuba Ignacio I. , Ji Lin , Roth Jack A. , Minna John D. , Pertsemlidis Alexander
Published: August 2009

FUS1 is a tumor suppressor gene located on human chromosome 3p21, and expression of Fus1 protein is highly regulated at various levels, leading to lost or greatly diminished tumor suppressor function in many lung cancers. Here we show that selected microRNAs (miRNA) interact with the 3′-untranslated region (3′UTR) of FUS1, leading to down-regulation of protein expression. Using computational methods, we first predicted that FUS1 is a target of three miRNAs, miR-93, miR-98, and miR-197, and then showed that exogenous overexpression of these miRNAs inhibited Fus1 protein expression. We then confirmed that the three miRNAs target the 3′UTR region of the FUS1 transcript and that individual deletion of the three miRNA target sites in the FUS1 3′UTR restores the expression level of Fus1 protein. We further found that miR-93 and miR-98 are expressed at higher levels in small-cell lung cancer cell lines (SCLC) than in non–small-cell lung cancer cell lines (NSCLC) and immortalized human bronchial epithelial cells (HBEC), and that miR-197 is expressed at higher levels in both SCLCs and NSCLCs than in HBECs. Finally, we found that elevated miR-93 and miR-197 expression is correlated with reduced Fus1 expression in NSCLC tumor specimens. These results suggest that the three miRNAs are negative regulators of Fus1 expression in lung cancers. (Mol Cancer Res 2009;7(8):1234–43)
Up-Regulation of Krüppel-Like Factor 5 in Pancreatic Cancer Is Promoted by Interleukin-1β Signaling and Hypoxia-Inducible Factor-1α
Authors: Mori Akira , Moser Christian , Lang Sven A. , Hackl Christina , Gottfried Eva , Kreutz Marina , Schlitt Hans J. , Geissler Edward K. , Stoeltzing Oliver
Published: August 2009

Krüppel-like factor 5 (KLF5) is a transcription factor involved in cell transformation, proliferation, and carcinogenesis that can be up-regulated by RAS mutations. However, controversy persists as to whether it functions as a tumor suppressor or as an oncogene. Because KRAS is frequently mutated in pancreatic cancer, we investigated the regulation of KLF5 in this cancer entity. Our results show that KLF5 is overexpressed in pancreatic cancer cells and exceeds KLF5 expression of KRAS-mutated colon cancer cells. Surprisingly, inhibition of B-Raf/C-Raf or MAPK/Erk did not reduce KLF5 levels, suggesting that KLF5 expression is not promoted by KRAS-Raf-MEK-Erk signaling in pancreatic cancer. This finding is in striking contrast to reports on MEK-Erk–mediated KLF5 induction in colon cancer cells. Moreover, KLF5 expression levels neither correlated with the mutational status of KRAS nor with MEK phosphorylation in pancreatic cancer cells. Importantly, KLF5 was significantly up-regulated by interleukin (IL)-1β or hypoxia. The IL-1 β–mediated induction of KLF5 was diminished by blocking the p38 pathway. In addition, blocking IL-1R reduced the constitutive KLF5 expression, suggesting an autocrine activation loop. Moreover, KLF5 coimmunoprecipitated with hypoxia-inducible factor-1α (HIF-1α) and HIF-1αsiRNA reduced constitutive KLF5. Similarly, KLF5siRNA reduced the expression of the HIF-1α target gene GLUT-1. Furthermore, KLF5 expression was significantly elevated by high cell density, by anchorage-independent cell growth, and in tumor spheroids. Down-regulation of KLF5 by RNAi reduced the expression of the target genes, survivin, and platelet-derived growth factor-A. In conclusion, overexpression of KLF5 in human pancreatic cancer cells is not mediated by KRAS/Raf/MAPK/Erk signaling, but involves the IL-1β/IL-1R system, p38, and the transcription factor HIF-1α. (Mol Cancer Res 2009;7(8):1390–8)
Oncogenic Activating Mutations Are Associated with Local Copy Gain
Authors: Modrek Barmak , Ge Lin , Pandita Ajay , Lin Eva , Mohan Sankar , Yue Peng , Guerrero Steve , Lin William M. , Pham Thinh , Modrusan Zora , Seshagiri Somasekar , Stern Howard M. , Waring Paul , Garraway Levi A. , Chant John , Stokoe David , Cavet Guy
Published: August 2009

Although activating mutations and gains in copy number are key mechanisms for oncogene activation, the relationship between the two is not well understood. In this study, we focused on KRAS copy gains and mutations in non–small cell lung cancer. We found that KRAS copy gains occur more frequently in tumors with KRAS activating mutations and are associated with large increases in KRAS expression. These copy gains tend to be more focal in tumors with activating mutations than in those with wild-type KRAS. Fluorescence in situ hybridization analysis revealed that some tumors have homogeneous low-level gains of the KRAS locus, whereas others have high-level amplification of KRAS, often in only a fraction of tumor cells. Associations between activating mutation and copy gains were also observed for other oncogenes (EGFR in non–small cell lung cancer, BRAF and NRAS in melanoma). Activating mutations were associated with copy gains only at the mutated oncogene locus but not other oncogene loci. However, KRAS activating mutations in colorectal cancer were not associated with copy gains. Future work is warranted to clarify the relationship among the different mechanisms of oncogene activation. (Mol Cancer Res 2009;7(8):1244–52)
NF-κB Activation, Dependent on Acetylation/Deacetylation, Contributes to HIF-1 Activity and Migration of Bone Metastatic Breast Carcinoma Cells
Authors: Bendinelli Paola , Matteucci Emanuela , Maroni Paola , Desiderio Maria Alfonsina
Published: August 2009

Here, we show that NF-κB-HIF-1 interaction contributed to breast cancer metastatic capacity by means of an incomplete epithelial/mesenchymal transition and influencing migration, as shown in 1833 (human) and 4T1 (mouse) metastatic cells after different stimuli. The 1833 and the transforming growth factor-β1–exposed 4T1 cells showed both epithelial (E-cadherins) and mesenchymal (N-cadherins and vimentin) markers, and common mechanisms contributed to the retention of certain epithelial characteristics and the control of migration. The complex NF-κB-HIF-1 reciprocal regulation and the enhanced c-Jun expression played a functional role in exacerbating the invasiveness of 1833 cells after p50/p65 transfection and of 4T1 cells exposed to transforming growth factor-β1. Twist expression seemed to exert a permissive role also regulating epithelial/mesenchymal transition markers. After c-Src wild-type (Srcwt) transfection, c-Src-signal transducer overexpression in 1833 cells increased HIF-1 transactivating activity and invasiveness, and changed E-cadherin/N-cadherin ratio versus mesenchymal phenotype. The transcription factor pattern and the motile phenotype of metastatic 1833 cells were influenced by p65-lysine acetylation and HDAC-dependent epigenetic mechanisms, which positively regulated basal NF-κB and HIF-1 activities. However, HDAC3 acted as a corepressor of NF-κB activity in parental MDA-MB231 cells, thus explaining many differences from the derived 1833 clone, including reduced HIF-1α and c-Jun expression. Invasiveness was differently affected by HDAC knockdown in 1833 and MDA-MB231 cells. We suggest that acetylation/deacetylation are critical in establishing the bone-metastatic gene signature of 1833 cells by regulating the activity of NF-κB and HIF-1, and further clarify the epigenetic control of transcription factor network in the motile phenotype of 1833 cells. (Mol Cancer Res 2009;7(8):1328–41)
Smad3 Is a Key Nonredundant Mediator of Transforming Growth Factor β Signaling in Nme Mouse Mammary Epithelial Cells
Authors: Dzwonek Joanna , Preobrazhenska Olena , Cazzola Silvia , Conidi Andrea , Schellens Ann , van Dinther Maarten , Stubbs Andrew , Klippel Anke , Huylebroeck Danny , ten Dijke Peter , Verschueren Kristin
Published: August 2009

Smad2 and Smad3 are intracellular mediators of transforming growth factor β (TGFβ) signaling that share various biochemical properties, but data emerging from functional analyses in several cell types indicate that these two Smad proteins may convey distinct cellular responses. Therefore, we have investigated the individual roles of Smad2 and Smad3 in mediating the cytostatic and proapoptotic effects of TGFβ as well as their function in epithelial-to-mesenchymal transition. For this purpose, we transiently depleted mouse mammary epithelial cells (Nme) of Smad2 and/or Smad3 mainly by a strategy relying on RNaseH-induced degradation of mRNA. The effect of such depletion on hallmark events of TGFβ-driven epithelial-to-mesenchymal transition was analyzed, including dissolution of epithelial junctions, formation of stress fibers and focal adhesions, activation of metalloproteinases, and transcriptional regulation of acknowledged target genes. Furthermore, we investigated the effect of Smad2 and Smad3 knockdown on the TGFβ-regulated transcriptome by microarray analysis. Our results identify Smad3 as a key factor to trigger TGFβ-regulated events and ascribe tumor suppressor as well as oncogenic activities to this protein. (Mol Cancer Res 2009;7(8):1342–53)
The Links between Transcription, β-catenin/JNK Signaling, and Carcinogenesis
Authors: Saadeddin Anas , Babaei-Jadidi Roya , Spencer-Dene Bradley , Nateri Abdolrahman Shams
Published: August 2009

Interactions between transcription and signaling are fundamentally important for understanding both the structure and function of genetic pathways and their role in diseases such as cancer. The finding that β-catenin/TCF4 and JNK/c-Jun cooperate has important implications in carcinogenesis. Previously, we found that binding of c-Jun and β-catenin/TCF4 to the c-jun promoter is dependent upon JNK activity, thus one role for this complex is to contribute to the repression and/or activation of genes that may mediate cell maintenance, proliferation, differentiation, and death, whereas deregulation of these signals may contribute to carcinogenesis. Here we address the functional links reported between activated β-catenin/JNK signaling pathways, their component genes, and their common targets, and discuss how alterations in the properties of these genes lead to the development of cancer. (Mol Cancer Res 2009;7(8):1189–96)
Roles of MKK1/2-ERK1/2 and Phosphoinositide 3-Kinase–AKT Signaling Pathways in Erlotinib-Induced Rad51 Suppression and Cytotoxicity in Human Non–Small Cell Lung Cancer Cells
Authors: Ko Jen-Chung , Ciou Shih-Ci , Jhan Jhih-Yuan , Cheng Chao-Min , Su Ying-Jhen , Chuang Show-Mei , Lin Szu-Ting , Chang Chia-Che , Lin Yun-Wei
Published: August 2009

Erlotinib (Tarceva) is a selective epidermal growth factor receptor tyrosine kinase inhibitor in the treatment of human non–small cell lung cancer (NSCLC). In this study, we investigated the roles of ERK1/2 and AKT signaling pathways in regulating Rad51 expression and cytotoxic effects in different NSCLC cell lines treated with erlotinib. Erlotinib decreased cellular levels of phosphorylated ERK1/2, phosphorylated AKT, Rad51 protein, and mRNA in erlotinib-sensitive H1650, A549, and H1869 cells, leading to cell death via apoptosis, but these results were not seen in erlotinib-resistant H520 and H1703 cells. Erlotinib decreased Rad51 protein levels by enhancing Rad51 mRNA and protein instability. Enforced expression of constitutively active MKK1 or AKT vectors could restore Rad51 protein levels, which were inhibited by erlotinib, and decrease erlotinib-induced cytotoxicity. Knocking down endogenous Rad51 expression by si-Rad51 RNA transfection significantly enhanced erlotinib-induced cytotoxicity. In contrast, overexpression of Rad51 by transfection with Rad51 vector could protect the cells from cytotoxic effects induced by erlotinib. Blocking the activations of ERK1/2 and AKT by MKK1/2 inhibitor (U0126) and phosphoinositide 3-kinase inhibitor (wortmannin) suppressed the expression of Rad51 and enhanced the erlotinib-induced cell death in erlotinib-resistant cells. In conclusion, suppression of Rad51 may be a novel therapeutic modality in overcoming drug resistance of erlotinib in NSCLC. (Mol Cancer Res 2009;7(8):1378–89)
Differential Influence of Normal and Cancer-Associated Fibroblasts on the Growth of Human Epithelial Cells in an In vitro Cocultivation Model of Prostate Cancer
Authors: Paland Nicole , Kamer Iris , Kogan-Sakin Ira , Madar Shalom , Goldfinger Naomi , Rotter Varda
Published: August 2009

The prostate is composed of a number of different cell populations. The interaction between them is crucial for the development and proper function of the prostate. However, the effect of the molecular cross talk between these cells in the course of carcinogenesis is still unclear. Employing an approach wherein immortalized epithelial cells and immortalized human fibroblasts were cocultured, we show that normal associated fibroblasts (NAF) and cancer-associated fibroblasts (CAF) differentially influenced the growth and proliferation of immortalized human prostate epithelial cells. Whereas NAFs inhibited the growth of immortalized epithelial cells but promoted the growth of metastatic PC-3 cells, CAFs promoted the growth of immortalized epithelial cells but not of PC-3. Cytokine arrays revealed that NAFs secreted higher levels of tumor necrosis factor-α compared with CAFs whereas CAFs secreted higher levels of interleukin-6 (IL-6) compared with NAFs. The growth-inhibiting effects of NAFs were counteracted by the addition of IL-6, and the growth-promoting effects exerted by the CAFs were counteracted by tumor necrosis factor-α. Furthermore, CAFs induced the migration of endothelial cells in an IL-6–dependent manner. Here, we show that normal fibroblast cells have a protective function at very early stages of carcinogenesis by preventing immortalized epithelial cells from proliferating and forming new blood vessels whereas CAFs aid immortalized epithelial cells to further develop. (Mol Cancer Res 2009;7(8):1212–23)
Functional Role of Cell Surface CUB Domain-Containing Protein 1 in Tumor Cell Dissemination
Authors: Deryugina Elena I. , Conn Erin M. , Wortmann Andreas , Partridge Juneth J. , Kupriyanova Tatyana A. , Ardi Veronica C. , Hooper John D. , Quigley James P.
Published: August 2009

The function of CUB domain-containing protein 1 (CDCP1), a recently described transmembrane protein expressed on the surface of hematopoietic stem cells and normal and malignant cells of different tissue origin, is not well defined. The contribution of CDCP1 to tumor metastasis was analyzed by using HeLa carcinoma cells overexpressing CDCP1 (HeLa-CDCP1) and a high-disseminating variant of prostate carcinoma PC-3 naturally expressing high levels of CDCP1 (PC3-hi/diss). CDCP1 expression rendered HeLa cells more aggressive in experimental metastasis in immunodeficient mice. Metastatic colonization by HeLa-CDCP1 was effectively inhibited with subtractive immunization-generated, CDCP1-specific monoclonal antibody (mAb) 41-2, suggesting that CDCP1 facilitates relatively late stages of the metastatic cascade. In the chick embryo model, time- and dose-dependent inhibition of HeLa-CDCP1 colonization by mAb 41-2 was analyzed quantitatively to determine when and where CDCP1 functions during metastasis. Quantitative PCR and immunohistochemical analyses indicated that CDCP1 facilitated tumor cell survival soon after vascular arrest. Live cell imaging showed that the function-blocking mechanism of mAb 41-2 involved enhancement of tumor cell apoptosis, confirmed by attenuation of mAb 41-2–mediated effects with the caspase inhibitor z-VAD-fmk. Under proapoptotic conditions in vitro, CDCP1 expression conferred HeLa-CDCP1 cells with resistance to doxorubicin-induced apoptosis, whereas ligation of CDCP1 with mAb 41-2 caused additional enhancement of the apoptotic response. The functional role of naturally expressed CDCP1 was shown by mAb 41-2–mediated inhibition of both experimental and spontaneous metastasis of PC3-hi/diss. These findings confirm that CDCP1 functions as an antiapoptotic molecule and indicate that during metastasis CDCP1 facilitates tumor cell survival likely during or soon after extravasation. (Mol Cancer Res 2009;7(8):1197–211)
Molecular Mechanisms of Human Papillomavirus-Induced Carcinogenesis
Authors: Lehoux Michaël , D’Abramo Claudia M. , Archambault Jacques
Published: August 2009

Approximately 20% of all cancers are associated with infectious agents. Among them, human papillomaviruses (HPVs) are very common and are now recognized as the etiological agent of cervical cancer, the second most common cancer in women worldwide, and they are increasingly linked with other forms of dysplasia. Carcinogenesis is a complex and multistep process requiring the acquisition of several genetic and/or epigenetic alterations. HPV-induced neoplasia, however, is in part mediated by the intrinsic functions of the viral proteins. In order to replicate its genome, HPV modulates the cell cycle, while deploying mechanisms to escape the host immune response, cellular senescence and apoptosis. As such, HPV infection leads directly and indirectly to genomic instability, further favouring transforming genetic events and progression to malignancy. This review aims to summarize our current understanding of the molecular mechanisms exploited by HPV to induce neoplasia, with an emphasis on the role of the 2 viral oncoproteins E6 and E7. Greater understanding of the role of HPV proteins in these processes will ultimately aid in the development of antiviral therapies, as well as unravel general mechanisms of oncogenesis.
Human Papillomaviruses: Genetic Basis of Carcinogenicity
Authors: Burk Robert D. , Chen Zigui , Van Doorslaer Koenraad
Published: August 2009

Persistent infection by specific oncogenic human papillomaviruses (HPVs) is established as the necessary cause of cervix cancer. DNA sequence differences between HPV genomes determine whether an HPV has the potential to cause cancer. Of the more than 100 HPV genotypes characterized at the genetic level, at least 15 are associated, to varying degrees, with cervical cancer. Classification based on nucleotide similarity places nearly all HPVs that infect the cervicovaginal area within the α-PV genus. Within this genus, phylogenetic trees inferred from the entire viral genome cluster all cancer-causing types together, suggesting the existence of a common ancestor for the oncogenic HPVs. However, in separate trees built from the early open reading frames (ORFs; i.e. E1, E2, E6, E7) or the late ORFs (i.e. L1, L2), the carcinogenic potential sorts with the early region of the genome, but not the late region. Thus, genetic differences within the early region specify the pathogenic potential of α-HPV infections. Since the HPV genomes are monophyletic and sites are highly correlated across the genome, diagnosis of oncogenic types and non-oncogenic types can be accomplished using any region across the genome. Here we review our current understanding of the evolutionary history of the oncogenic HPVs, in particular, we focus on the importance of viral genome heterogeneity and discuss the genetic basis for the oncogenic phenotype in some but not all α-PVs.
Epidemiology of Mucosal Human Papillomavirus Infection and Associated Diseases
Authors: Trottier Helen , Burchell Ann N.
Published: August 2009

This article describes the epidemiology of mucosal human papillomavirs (HPV) in adults and children, its mode of transmission and its associated diseases. Over 40 genotypes of HPV infect the epithelial lining of the anogenital tract and other mucosal areas of the body. HPV is the most common sexually transmitted infection globally, with high prevalences found in both females and males. The predominant route of transmission is via sexual contact, although mother-to-child transmission is also possible. HPV infection may exist asymptomatically or may induce the formation of benign or malignant tumours in the genital, oral or conjunctival mucosa. Although most infections clear spontaneously, those that persist result in substantial morbidity and invoke high costs associated with the treatment of clinically relevant lesions. Some 13–18 mucosal HPV types are considered to have high oncogenic potential. HPV is recognized unequivocally as the main causal factor for cervical cancer, and is further responsible for a substantial proportion of many other anogenital neoplasms and head and neck cancers. Infections with HPV types that have low oncogenic risk, such as HPV-6 and 11, are associated with benign lesions of the anogenital areas known as condylomata acuminata (genital warts), oral papillomas, conjunctival papillomas, as well as low-grade squamous intra-epithelial lesions of the cervix. Perinatally acquired HPV can also cause recurrent respiratory papillomatosis in infants and young children. The implementation of HPV vaccination therefore has the potential to prevent a substantial proportion of HPV-related disease in the future.
Prevention of Human Papillomavirus Infections and Associated Diseases by Vaccination: A New Hope for Global Public Health
Authors: Harper Diane M.
Published: August 2009

Cervarix® and Gardasil®, 2 human papillomavirus (HPV) vaccines, have been approved and implemented globally in young adolescent women with the hope of reducing the incidence of cervical cancer several decades hence. This program is dependent on the concept of ‘immunobridging’: antibody titers generated in young adolescents that are the same or higher than generated in HPV-naive 16- to 26-year-old women, the population in which efficacy is proven. Likewise, realizing a decline in cervical cancer from young adolescent female vaccination depends on the duration of vaccine efficacy, and the population coverage reached. While we patiently wait for results from our young adolescent vaccination programs, newly released data indicates that the immunogenicity and efficacy of the vaccines for young adult women with prior HPV exposure is equal or superior to that seen for young adolescents. This same concept of immunobridging supported by limited efficacy data offers the potential to reduce cervical cancer precursors within just a few years in our young sexually active adult women, a population secondary to our young adolescents. The HPV vaccines are not therapeutic. Neither vaccine will inhibit an already HPV-infected basal epithelial cell which continues to transform differentiated epithelial layers into cervical dysplasias. There is a clinical hope, though, already supported by early data, that the vaccines are capable of neutralizing HPV virions in host tissues from both auto-inoculated infections and infections in other organs than the cervix, thereby making it possible for these vaccines to prevent less common HPV-associated cancers of the penis, vagina, vulva, anus, oral cavity and oro-pharynx. Both vaccines have been shown to be generally safe in the phase II and phase III randomized controlled trials over 3–6.4 years. Post-marketing surveillance of Cervarix and Gardasil continues to show that they are safe for most women despite rarely occurring serious events.