bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from c-Myc and eIF4F Are Components of a Feedforward Loop that Links Transcription and Translation
Authors: Lin Chen-Ju , Cencic Regina , Mills John R. , Robert Francis , Pelletier Jerry
Published: March 2024

Abstract
The Myc/Max/Mad family of transcription factors and the eukaryotic initiation factor 4F (eIF4F) complex play fundamental roles in regulating cell growth, proliferation, differentiation, and oncogenic transformation. eIF4F is involved in the recruitment of ribosomes to mRNAs and is thought to generally be the rate-limiting phase of translation. Here, we show that c-Myc directly activates transcription of the three subunits of eIF4F (eIF4E, eIF4AI, and eIF4GI). These transcriptional effects are mediated through canonical E-boxes (5′CACGTG3′) present in the promoters of these genes. In addition, the c-Myc antagonist Mad1 down-regulates the expression of eIF4F subunits. We also show that MycER activation stimulates protein synthesis at the level of translation initiation. Increased eIF4F levels result in stimulation of c-Myc mRNA translation specifically, as assessed by quantitative reverse transcription–PCR. We use a murine model of lymphomagenesis to show the expression of eIF4F subunits is also up-regulated by c-Myc in vivo. Our results suggest the presence of a feedforward loop involving c-Myc and eIF4F that serves to link transcription and translation and that could contribute to the effects of c-Myc on cell proliferation and neoplastic growth. [Cancer Res 2008;68(13):5326–34]
Data from Tamoxifen Resistance in Breast Tumors Is Driven by Growth Factor Receptor Signaling with Repression of Classic Estrogen Receptor Genomic Function
Authors: Massarweh Suleiman , Osborne C. Kent , Creighton Chad J. , Qin Lanfang , Tsimelzon Anna , Huang Shixia , Weiss Heidi , Rimawi Mothaffar , Schiff Rachel
Published: March 2024

Abstract
Not all breast cancers respond to tamoxifen, and many develop resistance despite initial benefit. We used an in vivo model of estrogen receptor (ER)–positive breast cancer (MCF-7 xenografts) to investigate mechanisms of this resistance and develop strategies to circumvent it. Epidermal growth factor receptor (EGFR) and HER2, which were barely detected in control estrogen-treated tumors, increased slightly with tamoxifen and were markedly increased when tumors became resistant. Gefitinib, which inhibits EGFR/HER2, improved the antitumor effect of tamoxifen and delayed acquired resistance, but had no effect on estrogen-stimulated growth. Phosphorylated levels of p42/44 and p38 mitogen-activated protein kinases (both downstream of EGFR/HER2) were increased in the tamoxifen-resistant tumors and were suppressed by gefitinib. There was no apparent increase in phosphorylated AKT (also downstream of EGFR/HER2) in resistant tumors, but it was nonetheless suppressed by gefitinib. Phosphorylated insulin-like growth factor-IR (IGF-IR), which can interact with both EGFR and membrane ER, was elevated in the tamoxifen-resistant tumors compared with the sensitive group. However, ER-regulated gene products, including total IGF-IR itself and progesterone receptor, remained suppressed even at the time of acquired resistance. Tamoxifen's antagonism of classic ER genomic function was retained in these resistant tumors and even in tumors that overexpress HER2 (MCF-7 HER2/18) and are de novo tamoxifen-resistant. In conclusion, EGFR/HER2 may mediate tamoxifen resistance in ER-positive breast cancer despite continued suppression of ER genomic function by tamoxifen. IGF-IR expression remains dependent on ER but is activated in the tamoxifen-resistant tumors. This study provides a rationale to combine HER inhibitors with tamoxifen in clinical studies, even in tumors that do not initially overexpress EGFR/HER2. [Cancer Res 2008;68(3):826–33]
Data from Overexpression of Vascular Endothelial Growth Factor and the Development of Post-Transplantation Cancer
Authors: Basu Aninda , Contreras Alan G. , Datta Dipak , Flynn Evelyn , Zeng Liling , Cohen Herbert T. , Briscoe David M. , Pal Soumitro
Published: March 2024

Abstract
Cancer is an increasing and major problem after solid organ transplantation. In part, the increased cancer risk is associated with the use of immunosuppressive agents, especially calcineurin inhibitors. We propose that the effect of calcineurin inhibitors on the expression of vascular endothelial growth factor (VEGF) leads to an angiogenic milieu that favors tumor growth. Here, we used 786-0 human renal cancer cells to investigate the effect of cyclosporine (CsA) on VEGF expression. Using a full-length VEGF promoter-luciferase construct, we found that CsA markedly induced VEGF transcriptional activation through the protein kinase C (PKC) signaling pathway, specifically involving PKCζ and PKCδ isoforms. Moreover, CsA promoted the association of PKCζ and PKCδ with the transcription factor Sp1 as observed by immunoprecipitation assays. Using promoter deletion constructs, we found that CsA-mediated VEGF transcription was primarily Sp1 dependent. Furthermore, CsA-induced and PKC-Sp1–mediated VEGF transcriptional activation was partially inhibited by von Hippel-Lindau protein. CsA also promoted the progression of human renal tumors in vivo, wherein VEGF is overexpressed. Finally, to evaluate the in vivo significance of CsA-induced VEGF overexpression in terms of post-transplantation tumor development, we injected CT26 murine carcinoma cells (known to form angiogenic tumors) into mice with fully MHC mismatched cardiac transplants. We observed that therapeutic doses of CsA increased tumor size and VEGF mRNA expression and also enhanced tumor angiogenesis. However, coadministration of a blocking anti-VEGF antibody inhibited this CsA-mediated tumor growth. Collectively, these findings define PKC-mediated VEGF transcriptional activation as a key component in the progression of CsA-induced post-transplantation cancer. [Cancer Res 2008;68(14):5689–98]
Data from The Pervasive Presence of Fluctuating Oxygenation in Tumors
Authors: Cárdenas-Navia Laura I. , Mace Daniel , Richardson Rachel A. , Wilson David F. , Shan Siqing , Dewhirst Mark W.
Published: March 2024

Abstract
Tumor hypoxia is a persistent obstacle for traditional therapies in solid tumors. Strategies for mitigating the effects of hypoxic tumor cells have been developed under the assumption that chronically hypoxic tumor cells were the central cause of treatment resistance. In this study, we show that instabilities in tumor oxygenation are a prevalent characteristic of three tumor lines and previous characterization of tumor hypoxia as being primarily diffusion-limited does not accurately portray the tumor microenvironment. Phosphorescence lifetime imaging was used to measure fluctuations in vascular pO2 in rat fibrosarcomas, 9L gliomas, and R3230 mammary adenocarcinomas grown in dorsal skin-fold window chambers (n = 6 for each tumor type) and imaged every 2.5 minutes for a duration of 60 to 90 minutes. O2 delivery to tumors is constantly changing in all tumors, resulting in continuous reoxygenation events throughout the tumor. Vascular pO2 maps show significant spatial heterogeneity at each time point, as well as between time points. The fluctuations in oxygenation occur with a common periodicity within and between tumors, suggesting a common mechanism, but have tumor type–dependent spatial patterns. The widespread presence of fluctuations in tumor oxygenation has broad ranging implications for tumor progression, stress response, and signal transduction, which are altered by oxygenation/reoxygenation events. [Cancer Res 2008;68(14):5812–9]
Data from Diphenyl Difluoroketone: A Curcumin Derivative with Potent In vivo Anticancer Activity
Authors: Subramaniam Dharmalingam , May Randal , Sureban Sripathi M. , Lee Katherine B. , George Robert , Kuppusamy Periannan , Ramanujam Rama P. , Hideg Kalman , Dieckgraefe Brian K. , Houchen Courtney W. , Anant Shrikant
Published: March 2024

Abstract
Diphenyl difluoroketone (EF24), a molecule having structural similarity to curcumin, was reported to inhibit proliferation of a variety of cancer cells in vitro. However, the efficacy and in vivo mechanism of action of EF24 in gastrointestinal cancer cells have not been investigated. Here, we assessed the in vivo therapeutic effects of EF24 on colon cancer cells. Using hexosaminidase assay, we determined that EF24 inhibits proliferation of HCT-116 and HT-29 colon and AGS gastric adenocarcinoma cells but not of mouse embryo fibroblasts. Furthermore, the cancer cells showed increased levels of activated caspase-3 and increased Bax to Bcl-2 and Bax to Bcl-xL ratios, suggesting that the cells were undergoing apoptosis. At the same time, cell cycle analysis showed that there was an increased number of cells in the G2-M phase. To determine the effects of EF24 in vivo, HCT-116 colon cancer xenografts were established in nude mice and EF24 was given i.p. EF24 significantly suppressed the growth of colon cancer tumor xenografts. Immunostaining for CD31 showed that there was a lower number of microvessels in the EF24-treated animals coupled with decreased cyclooxygenase-2, interleukin-8, and vascular endothelial growth factor mRNA and protein expression. Western blot analyses also showed decreased AKT and extracellular signal-regulated kinase activation in the tumors. Taken together, these data suggest that the novel curcumin-related compound EF24 is a potent antitumor agent that induces caspase-mediated apoptosis during mitosis and has significant therapeutic potential for gastrointestinal cancers. [Cancer Res 2008;68(6):1962–9]
Data from Maintenance of Intratumoral Androgens in Metastatic Prostate Cancer: A Mechanism for Castration-Resistant Tumor Growth
Authors: Montgomery R. Bruce , Mostaghel Elahe A. , Vessella Robert , Hess David L. , Kalhorn Thomas F. , Higano Celestia S. , True Lawrence D. , Nelson Peter S.
Published: March 2024

Abstract
Therapy for advanced prostate cancer centers on suppressing systemic androgens and blocking activation of the androgen receptor (AR). Despite anorchid serum androgen levels, nearly all patients develop castration-resistant disease. We hypothesized that ongoing steroidogenesis within prostate tumors and the maintenance of intratumoral androgens may contribute to castration-resistant growth. Using mass spectrometry and quantitative reverse transcription–PCR, we evaluated androgen levels and transcripts encoding steroidogenic enzymes in benign prostate tissue, untreated primary prostate cancer, metastases from patients with castration-resistant prostate cancer, and xenografts derived from castration-resistant metastases. Testosterone levels within metastases from anorchid men [0.74 ng/g; 95% confidence interval (95% CI), 0.59–0.89] were significantly higher than levels within primary prostate cancers from untreated eugonadal men (0.23 ng/g; 95% CI, 0.03–0.44; P < 0.0001). Compared with primary prostate tumors, castration-resistant metastases displayed alterations in genes encoding steroidogenic enzymes, including up-regulated expression of FASN, CYP17A1, HSD3B1, HSD17B3, CYP19A1, and UGT2B17 and down-regulated expression of SRD5A2 (P < 0.001 for all). Prostate cancer xenografts derived from castration-resistant tumors maintained similar intratumoral androgen levels when passaged in castrate compared with eugonadal animals. Metastatic prostate cancers from anorchid men express transcripts encoding androgen-synthesizing enzymes and maintain intratumoral androgens at concentrations capable of activating AR target genes and maintaining tumor cell survival. We conclude that intracrine steroidogenesis may permit tumors to circumvent low levels of circulating androgens. Maximal therapeutic efficacy in the treatment of castration-resistant prostate cancer will require novel agents capable of inhibiting intracrine steroidogenic pathways within the prostate tumor microenvironment. [Cancer Res 2008;68(11):4447–54]
Data from Epigenetic Silencing of MicroRNA-34b/c and B-Cell Translocation Gene 4 Is Associated with CpG Island Methylation in Colorectal Cancer
Authors: Toyota Minoru , Suzuki Hiromu , Sasaki Yasushi , Maruyama Reo , Imai Kohzoh , Shinomura Yasuhisa , Tokino Takashi
Published: March 2024

Abstract
Altered expression of microRNA (miRNA) is strongly implicated in cancer, and recent studies have shown that, in cancer, expression of some miRNAs cells is silenced in association with CpG island hypermethylation. To identify epigenetically silenced miRNAs in colorectal cancer (CRC), we screened for miRNAs induced in CRC cells by 5-aza-2′-deoxycytidine (DAC) treatment or DNA methyltransferase knockout. We found that miRNA-34b (miR-34b) and miR-34c, two components of the p53 network, are epigenetically silenced in CRC; that this down-regulation of miR-34b/c is associated with hypermethylation of the neighboring CpG island; and that DAC treatment rapidly restores miR-34b/c expression. Methylation of the miR-34b/c CpG island was frequently observed in CRC cell lines (nine of nine, 100%) and in primary CRC tumors (101 of 111, 90%), but not in normal colonic mucosa. Transfection of precursor miR-34b or miR-34c into CRC cells induced dramatic changes in the gene expression profile, and there was significant overlap between the genes down-regulated by miR-34b/c and those down-regulated by DAC. We also found that the miR-34b/c CpG island is a bidirectional promoter which drives expression of both miR-34b/c and B-cell translocation gene 4 (BTG4); that methylation of the CpG island is also associated with transcriptional silencing of BTG4; and that ectopic expression of BTG4 suppresses colony formation by CRC cells. Our results suggest that miR-34b/c and BTG4 are novel tumor suppressors in CRC and that the miR-34b/c CpG island, which bidirectionally regulates miR-34b/c and BTG4, is a frequent target of epigenetic silencing in CRC. [Cancer Res 2008;68(11):4123–32]
Data from The Wilms' Tumor Antigen Is a Novel Target for Human CD4+ Regulatory T Cells: Implications for Immunotherapy
Authors: Lehe Cynthia , Ghebeh Hazem , Al-Sulaiman Abdullah , Al Qudaihi Ghofran , Al-Hussein Khaled , Almohareb Fahad , Chaudhri Naeem , Alsharif Fahad , Al-Zahrani Hazza , Tbakhi Abdelghani , Aljurf Mahmoud , Dermime Said
Published: March 2024

Abstract
Compelling evidences indicate a key role for regulatory T cells (Treg) on the host response to cancer. The Wilms' tumor antigen (WT1) is overexpressed in several human leukemias and thus considered as promising target for development of leukemia vaccine. However, recent studies indicated that the generation of effective WT1-specific cytotoxic T cells can be largely affected by the presence of Tregs. We have generated T-cell lines and clones that specifically recognized a WT1-84 (RYFKLSHLQMHSRKH) peptide in an HLA-DRB1*0402–restricted manner. Importantly, they recognized HLA-DRB1*04–matched fresh leukemic cells expressing the WT1 antigen. These clones exerted a T helper 2 cytokine profile, had a CD4+CD25+Foxp3+GITR+CD127− Treg phenotype, and significantly inhibited the proliferative activity of allogeneic T cells independently of cell contact. Priming of alloreactive T cells in the presence of Tregs strongly inhibited the expansion of natural killer (NK), NK T, and CD8+ T cells and had an inhibitory effect on NK/NK T cytotoxic activity but not on CD8+ T cells. Furthermore, priming of T cells with the WT1-126 HLA-A0201–restricted peptide in the presence of Tregs strongly inhibited the induction of anti–WT1-126 CD8+ CTL responses as evidenced by both very low cytotoxic activity and IFN-γ production. Moreover, these Treg clones specifically produced granzyme B and selectively induced apoptosis in WT1-84–pulsed autologous antigen-presenting cells but not in apoptotic-resistant DR4-matched leukemic cells. Importantly, we have also detected anti–WT1-84 interleukin-5+/granzyme B+/Foxp3+ CD4+ Tregs in five of eight HLA-DR4+ acute myeloid leukemia patients. Collectively, our in vitro and in vivo findings strongly suggest important implications for the clinical manipulation of Tregs in cancer patients. [Cancer Res 2008;68(15):6350–9]
Data from Fibronectin Expression Modulates Mammary Epithelial Cell Proliferation during Acinar Differentiation
Authors: Williams Courtney M. , Engler Adam J. , Slone R. Daniel , Galante Leontine L. , Schwarzbauer Jean E.
Published: March 2024

Abstract
The mammary gland consists of a polarized epithelium surrounded by a basement membrane matrix that forms a series of branching ducts ending in hollow, sphere-like acini. Essential roles for the epithelial basement membrane during acinar differentiation, in particular laminin and its integrin receptors, have been identified using mammary epithelial cells cultured on a reconstituted basement membrane. Contributions from fibronectin, which is abundant in the mammary gland during development and tumorigenesis, have not been fully examined. Here, we show that fibronectin expression by mammary epithelial cells is dynamically regulated during the morphogenic process. Experiments with synthetic polyacrylamide gel substrates implicate both specific extracellular matrix components, including fibronectin itself, and matrix rigidity in this regulation. Alterations in fibronectin levels perturbed acinar organization. During acinar development, increased fibronectin levels resulted in overproliferation of mammary epithelial cells and increased acinar size. Addition of fibronectin to differentiated acini stimulated proliferation and reversed growth arrest of mammary epithelial cells negatively affecting maintenance of proper acinar morphology. These results show that expression of fibronectin creates a permissive environment for cell growth that antagonizes the differentiation signals from the basement membrane. These effects suggest a link between fibronectin expression and epithelial cell growth during development and oncogenesis in the mammary gland. [Cancer Res 2008;68(9):3185–92]
Data from Podoplanin Is a Novel Fos Target Gene in Skin Carcinogenesis
Authors: Durchdewald Moritz , Guinea-Viniegra Juan , Haag Daniel , Riehl Astrid , Lichter Peter , Hahn Meinhard , Wagner Erwin F. , Angel Peter , Hess Jochen
Published: March 2024

Abstract
Expression and function of the oncogenic transcription factor activator protein (AP-1; mainly composed of Jun and Fos proteins) is required for neoplastic transformation of keratinocytes in vitro and tumor promotion as well as malignant progression in vivo. Here, we describe the identification of 372 differentially expressed genes comparing skin tumor samples of K5-SOS-F transgenic mice (Fosf/f SOS+) with samples derived from animals with a specific deletion of c-Fos in keratinocytes (FosΔep SOS+). Fos-dependent transcription of selected genes was confirmed by quantitative real-time PCR analysis using tumor samples and mouse back skin treated with the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA). One of the most differentially expressed genes encodes the small mucin–like glycoprotein Podoplanin (Pdpn), whose expression correlates with malignant progression in mouse tumor model systems and human cancer. We found Pdpn and Fos expression in chemically induced mouse skin tumors, and detailed analysis of the Pdpn gene promoter revealed impaired activity in Fos-deficient mouse embryonic fibroblasts, which could be restored by ectopic Fos expression. Direct Fos protein binding to the Pdpn promoter was shown by chromatin immunoprecipitation and a TPA-induced complex at a TPA-responsive element–like motif in the proximal promoter was identified by electrophoretic mobility shift assays. In summary, we could define a Fos-dependent genetic program in a well-established model of skin tumors. Systematic analysis of these novel target genes will guide us in elucidating the molecular mechanisms of AP-1–regulated pathways that are critically implicated in neoplastic transformation and/or malignant progression. [Cancer Res 2008;68(17):6877–83]
Data from Rb Depletion Results in Deregulation of E-Cadherin and Induction of Cellular Phenotypic Changes that Are Characteristic of the Epithelial-to-Mesenchymal Transition
Authors: Arima Yoshimi , Inoue Yasumichi , Shibata Tatsuhiro , Hayashi Hidemi , Nagano Osamu , Saya Hideyuki , Taya Yoichi
Published: March 2024

Abstract
The retinoblastoma tumor suppressor protein (Rb) is mutated or expressed at very low levels in several tumor types, including retinoblastoma and osteosarcoma, as well as small cell lung, colon, prostate, bladder, and breast carcinomas. Loss or reduction of Rb expression is seen most commonly in high-grade breast adenocarcinomas, suggesting that a relationship may exist between loss of Rb function and a less-differentiated state, increased proliferation, and high metastatic potential. In this study, we found that knockdown of Rb by small interfering RNA in MCF7 breast cancer cells disrupts cell-cell adhesion and induces a mesenchymal-like phenotype. The epithelial-to-mesenchymal transition (EMT), a key event in embryonic morphogenesis, is implicated in the metastasis of primary tumors. Additionally, Rb is decreased during growth factor– and cytokine-induced EMT and overexpression of Rb inhibits the EMT in MCF10A human mammary epithelial cells. Ectopic expression and knockdown of Rb resulted in increased or reduced expression of E-cadherin, which is specifically involved in epithelial cell-cell adhesion. Other EMT-related transcriptional factors, including Slug and Zeb-1, are also induced by Rb depletion. Furthermore, we confirmed that Rb binds to an E-cadherin promoter sequence in association with the transcription factor activator protein-2α. Finally, in breast cancer specimens, we observed a concurrent down-regulation of Rb and E-cadherin expression in mesenchymal-like invasive cancers. These findings suggest that Rb inactivation contributes to tumor progression due to not only loss of cell proliferation control but also conversion to an invasive phenotype and that the inhibition of EMT is a novel tumor suppressor function of Rb. [Cancer Res 2008;68(13):5104–12]
Data from Mitochondrial Cholesterol Contributes to Chemotherapy Resistance in Hepatocellular Carcinoma
Authors: Montero Joan , Morales Albert , Llacuna Laura , Lluis Josep M. , Terrones Oihana , Basañez Gorka , Antonsson Bruno , Prieto Jesús , García-Ruiz Carmen , Colell Anna , Fernández-Checa José C.
Published: March 2024

Abstract
Cholesterol metabolism is deregulated in carcinogenesis, and cancer cells exhibit enhanced mitochondrial cholesterol content whose role in cell death susceptibility and cancer therapy has not been investigated. Here, we describe that mitochondria from rat or human hepatocellular carcinoma (HC) cells (HCC) or primary tumors from patients with HC exhibit increased mitochondrial cholesterol levels. HCC sensitivity to chemotherapy acting via mitochondria is enhanced upon cholesterol depletion by inhibition of hydroxymethylglutaryl-CoA reductase or squalene synthase (SS), which catalyzes the first committed step in cholesterol biosynthesis. HCC transfection with siRNA targeting the steroidogenic acute regulatory protein StAR, a mitochondrial cholesterol–transporting polypeptide which is overexpressed in HCC compared with rat and human liver, sensitized HCC to chemotherapy. Isolated mitochondria from HCC with increased cholesterol levels were resistant to mitochondrial membrane permeabilization and release of cytochrome c or Smac/DIABLO in response to various stimuli including active Bax. Similar behavior was observed in cholesterol-enriched mitochondria or liposomes and reversed by restoring mitochondrial membrane order or cholesterol extraction. Moreover, atorvastatin or the SS inhibitor YM-53601 potentiated doxorubicin-mediated HCC growth arrest and cell death in vivo. Thus, mitochondrial cholesterol contributes to chemotherapy resistance by increasing membrane order, emerging as a novel therapeutic niche in cancer therapy. [Cancer Res 2008;68(13):5246–56]
Data from Genomic and Proteomic Analysis Reveals a Threshold Level of MYC Required for Tumor Maintenance
Authors: Shachaf Catherine M. , Gentles Andrew J. , Elchuri Sailaja , Sahoo Debashis , Soen Yoav , Sharpe Orr , Perez Omar D. , Chang Maria , Mitchel Dennis , Robinson William H. , Dill David , Nolan Garry P. , Plevritis Sylvia K. , Felsher Dean W.
Published: March 2024

Abstract
MYC overexpression has been implicated in the pathogenesis of most types of human cancers. MYC is likely to contribute to tumorigenesis by its effects on global gene expression. Previously, we have shown that the loss of MYC overexpression is sufficient to reverse tumorigenesis. Here, we show that there is a precise threshold level of MYC expression required for maintaining the tumor phenotype, whereupon there is a switch from a gene expression program of proliferation to a state of proliferative arrest and apoptosis. Oligonucleotide microarray analysis and quantitative PCR were used to identify changes in expression in 3,921 genes, of which 2,348 were down-regulated and 1,573 were up-regulated. Critical changes in gene expression occurred at or near the MYC threshold, including genes implicated in the regulation of the G1-S and G2-M cell cycle checkpoints and death receptor/apoptosis signaling. Using two-dimensional protein analysis followed by mass spectrometry, phospho-flow fluorescence-activated cell sorting, and antibody arrays, we also identified changes at the protein level that contributed to MYC-dependent tumor regression. Proteins involved in mRNA translation decreased below threshold levels of MYC. Thus, at the MYC threshold, there is a loss of its ability to maintain tumorigenesis, with associated shifts in gene and protein expression that reestablish cell cycle checkpoints, halt protein translation, and promote apoptosis. [Cancer Res 2008;68(13):5132–42]
Data from OX40 Agonist Therapy Enhances CD8 Infiltration and Decreases Immune Suppression in the Tumor
Authors: Gough Michael J. , Ruby Carl E. , Redmond William L. , Dhungel Birat , Brown Alexis , Weinberg Andrew D.
Published: March 2024

Abstract
Acquisition of full T-cell effector function and memory differentiation requires appropriate costimulatory signals, including ligation of the costimulatory molecule OX40 (TNFRSF4, CD134). Tumors often grow despite the presence of tumor-specific T cells and establish an environment with weak costimulation and immune suppression. Administration of OX40 agonists has been shown to significantly increase the survival of tumor-bearing mice and was dependent on the presence of both CD4 and CD8 T cells during tumor-specific priming. To understand how OX40 agonists work in mice with established tumors, we developed a model to study changes in immune cell populations within the tumor environment. We show here that systemic administration of OX40 agonist antibodies increased the proportion of CD8 T cells at the tumor site in three different tumor models. The function of the CD8 T cells at the tumor site was also increased by administration of OX40 agonist antibody, and we observed an increase in the proportion of antigen-specific CD8 T cells within the tumor. Despite decreases in the proportion of T regulatory cells at the tumor site, T regulatory cell function in the spleen was unaffected by OX40 agonist antibody therapy. Interestingly, administration of OX40 agonist antibody caused significant changes in the tumor stroma, including decreased macrophages, myeloid-derived suppressor cells, and decreased expression of transforming growth factor-β. Thus, therapies targeting OX40 dramatically changed the tumor environment by enhancing the infiltration and function of CD8 T cells combined with diminished suppressive influences within the tumor. [Cancer Res 2008;68(13):5206–15]
Data from Truncated ETV1, Fused to Novel Tissue-Specific Genes, and Full-Length ETV1 in Prostate Cancer
Authors: Hermans Karin G. , van der Korput Hetty A. , van Marion Ronald , van de Wijngaart Dennis J. , Ziel-van der Made Angelique , Dits Natasja F. , Boormans Joost L. , van der Kwast Theo H. , van Dekken Herman , Bangma Chris H. , Korsten Hanneke , Kraaij Robert , Jenster Guido , Trapman Jan
Published: March 2024

Abstract
In this study, we describe the properties of novel ETV1 fusion genes, encoding N-truncated ETV1 (dETV1), and of full-length ETV1, overexpressed in clinical prostate cancer. We detected overexpression of novel ETV1 fusion genes or of full-length ETV1 in 10% of prostate cancers. Novel ETV1 fusion partners included FOXP1, an EST (EST14), and an endogenous retroviral repeat sequence (HERVK17). Like TMPRSS2, EST14 and HERVK17 were prostate-specific and androgen-regulated expressed. This unique expression pattern of most ETV1 fusion partners seems an important determinant in prostate cancer development. In transient reporter assays, full-length ETV1 was a strong transactivator, whereas dETV1 was not. However, several of the biological properties of dETV1 and full-length ETV1 were identical. On stable overexpression, both induced migration and invasion of immortalized nontumorigenic PNT2C2 prostate epithelial cells. In contrast to dETV1, full-length ETV1 also induced anchorage-independent growth of these cells. PNT2C2 cells stably transfected with dETV1 or full-length ETV1 expression constructs showed small differences in induced expression of target genes. Many genes involved in tumor invasion/metastasis, including uPA/uPAR and MMPs, were up-regulated in both cell types. Integrin β3 (ITGB3) was clearly up-regulated by full-length ETV1 but much less by dETV1. Based on the present data and on previous findings, a novel concept of the role of dETV1 and of full-length ETV1 overexpression in prostate cancer is proposed. [Cancer Res 2008;68(18):7541–9]
Data from Silencing of HSP90 Cochaperone AHA1 Expression Decreases Client Protein Activation and Increases Cellular Sensitivity to the HSP90 Inhibitor 17-Allylamino-17-Demethoxygeldanamycin
Authors: Holmes Joanna L. , Sharp Swee Y. , Hobbs Steve , Workman Paul
Published: March 2024

Abstract
AHA1 (activator of HSP90 ATPase) is a cochaperone of the ATP-dependent molecular chaperone, HSP90, which is involved in the maturation, stabilization/degradation, and function of oncogenic proteins. HSP90 operates in a multimeric complex driven by the binding and hydrolysis of ATP. Treatment of cells with the HSP90 inhibitor 17-allylamino-17-demethoxygeldanamycin (17-AAG) results in the degradation of client proteins via the ubiquitin-proteasome pathway. As AHA1 increases the ATPase activity of HSP90, we hypothesized that modulation of AHA1 expression could influence the activity of client proteins and/or the cellular response to 17-AAG. We show that the basal expression of AHA1 is different across a panel of human cancer cell lines, and that treatment with 17-AAG resulted in sustained AHA1 up-regulation. Increasing the expression of AHA1 did not affect the sensitivity to 17-AAG, but did increase C-RAF activity and the levels of phosphorylated MEK1/2 and ERK1/2 without affecting total levels of these proteins or of client proteins C-RAF, ERBB2, or CDK4. Conversely, small interfering RNA–selective knockdown of >80% of AHA1 expression decreased C-RAF activity and reduced the levels of MEK1/2 and ERK1/2 phosphorylation. Moreover, the AHA1 knockdown resulted in a significant (P < 0.05) increase in sensitivity to 17-AAG, due in part to a 2- to 3-fold increase in apoptosis. These results show that the reduction of AHA1 levels could decrease the phosphorylation of key signal transduction proteins, and for the first time, separate the activation and stabilization functions of HSP90. Furthermore, AHA1 knockdown could sensitize cancer cells to 17-AAG. We conclude that modulation of AHA1 might be a potential therapeutic strategy to increase sensitivity to HSP90 inhibitors. [Cancer Res 2008;68(4):1187–97]
Data from A Role for TAZ in Migration, Invasion, and Tumorigenesis of Breast Cancer Cells
Authors: Chan Siew Wee , Lim Chun Jye , Guo Ke , Ng Chee Peng , Lee Ian , Hunziker Walter , Zeng Qi , Hong Wanjin
Published: March 2024

Abstract
TAZ (WWTR1), identified as a 14-3-3 binding protein with a PDZ binding motif, modulates mesenchymal stem cell differentiation. We now show that TAZ plays a critical role in the migration, invasion, and tumorigenesis of breast cancer cells. TAZ is conspicuously expressed in human breast cancer cell lines in which its expression levels generally correlate with the invasiveness of cancer cells. Overexpression of TAZ in low-expressing MCF10A cells causes morphologic changes characteristic of cell transformation and promotes cell migration and invasion. Conversely, RNA interference–mediated knockdown of TAZ expression in MCF7 and Hs578T cells reduces cell migration and invasion. TAZ knockdown in MCF7 cells also retards anchorage-independent growth in soft agar and tumorigenesis in nude mice. Significantly, TAZ is overexpressed in ∼20% of breast cancer samples. These results indicate that TAZ plays a role in the migration, invasion, and tumorigenesis of breast cancer cells and thus presents a novel target for the detection and treatment of breast cancer. [Cancer Res 2008;68(8):2592–8]
Data from Hypermethylator Phenotype in Sporadic Colon Cancer: Study on a Population-Based Series of 582 Cases
Authors: Barault Ludovic , Charon-Barra Céline , Jooste Valérie , de la Vega Mathilde Funes , Martin Laurent , Roignot Patrick , Rat Patrick , Bouvier Anne-Marie , Laurent-Puig Pierre , Faivre Jean , Chapusot Caroline , Piard Francoise
Published: March 2024

Abstract
The CpG island methylator phenotype (CIMP) is a distinct phenotype in colorectal cancer, associated with specific clinical, pathologic, and molecular features. However, most of the studies stratified methylation according to two subgroups (CIMP-High versus No-CIMP/CIMP-Low). In our study, we defined three different subgroups of methylation (No-CIMP, CIMP-Low, and CIMP-High) and evaluated the prognostic significance of methylation status on a population-based series of sporadic colon cancers. A total of 582 colon adenocarcinomas were evaluated using methylation-specific PCR for 5 markers (hMLH1, P16, MINT1, MINT2, and MINT31). No-CIMP status was defined as no methylated locus, CIMP-Low status as one to three methylated loci, and CIMP-High status as four or five methylated loci. Clinicopathologic and molecular characteristics were correlated to the methylation status. Crude and relative survival was compared according to methylation status. In the microsatellite-stable (MSS) group, CIMP-High was significantly associated with proximal location (P = 0.011) and BRAF mutation (P < 0.001). KRAS mutations were more associated with CIMP-High and CIMP-Low status (P = 0.008). A shorter 5-year survival was observed in MSS cancer patients with CIMP-Low or CIMP-High status. These results remained significant in multivariate analysis adjusted for age, stage, and BRAF and KRAS mutational status [CIMP-Low: hazard ratio (HR), 1.85; 95% confidence interval (95% CI), 1.37–2.51; CIMP-High, HR, 2.90; 95% CI, 1.53–5.49 compared with No-CIMP]. Within the high-level microsatellite instability group, no difference in survival was observed between the different CIMP groups. Our results show the interest of defining three subgroups of patients according to their methylation status (No-CIMP/CIMP-Low/CIMP-High). Methylation is an independent prognostic factor in MSS colon cancer. [Cancer Res 2008;68(20):8541–6]
Data from EBV Is Necessary for Proliferation of Dually Infected Primary Effusion Lymphoma Cells
Authors: Mack Amanda A. , Sugden Bill
Published: March 2024

Abstract
Epstein Barr virus (EBV) and Kaposi's sarcoma–associated herpesvirus (KSHV) are found together in ∼80% of primary effusion lymphomas (PEL), but their contribution to these cancers is unclear. We found that dominant-negative derivatives of EBNA1 inhibited EBV-positive PEL cells from forming colonies. Those rare PEL cells that proliferated after expression of the dominant-negative derivatives usually expressed these derivatives at low or undetectable levels and continued to maintain their EBV genomes. Those proliferating cells expressing higher levels of the derivatives expressed mutant derivatives that could not bind DNA. These findings indicate that EBV is required to sustain proliferation, as measured by colony formation of dually infected PEL cells. The dominant-negative derivatives of EBNA1 had no effect on the colony-forming ability of five EBV-negative, KSHV-negative hematopoietic cell lines. Surprisingly, they did inhibit the colony-forming ability of EBV-negative, KSHV-positive PEL cells. The small fraction of cells that continued to proliferate expressed only mutants of the EBNA1 derivatives that could no longer bind DNA. These findings indicate that the site-specific DNA-binding activity of EBNA1 or its derivatives when expressed efficiently in EBV-negative, KSHV-positive PEL cells inhibits their colony formation possibly through their binding to the KSHV genome. [Cancer Res 2008;68(17):6963–8]
Data from SLITs Suppress Tumor Growth In vivo by Silencing Sdf1/Cxcr4 within Breast Epithelium
Authors: Marlow Rebecca , Strickland Phyllis , Lee Ji Shin , Wu Xinyan , PeBenito Milana , Binnewies Mikhail , Le Elizabeth K. , Moran Angel , Macias Hector , Cardiff Robert D. , Sukumar Saraswati , Hinck Lindsay
Published: March 2024

Abstract
The genes encoding Slits and their Robo receptors are silenced in many types of cancer, including breast, suggesting a role for this signaling pathway in suppressing tumorigenesis. The molecular mechanism underlying these tumor-suppressive effects has not been delineated. Here, we show that loss of Slits, or their Robo1 receptor, in murine mammary gland or human breast carcinoma cells results in coordinate up-regulation of the Sdf1 and Cxcr4 signaling axis, specifically within mammary epithelium. This is accompanied by hyperplastic changes in cells and desmoplastic alterations in the surrounding stroma. A similar inverse correlation between Slit and Cxcr4 expression is identified in human breast tumor tissues. Furthermore, we show in a xenograft model that Slit overexpression down-regulates CXCR4 and dominantly suppresses tumor growth. These studies classify Slits as negative regulators of Sdf1 and Cxcr4 and identify a molecular signature in hyperplastic breast lesions that signifies inappropriate up-regulation of key prometastatic genes. [Cancer Res 2008;68(19):7819–27]
Data from The ARF Tumor Suppressor Can Promote the Progression of Some Tumors
Authors: Humbey Olivier , Pimkina Julia , Zilfou Jack T. , Jarnik Michal , Dominguez-Brauer Carmen , Burgess Darren J. , Eischen Christine M. , Murphy Maureen E.
Published: March 2024

Abstract
p14/p19ARF (ARF) is a tumor suppressor gene that is frequently mutated in human cancer. ARF has multiple tumor suppressor functions, some of which are mediated by signaling to p53. Surprisingly, a significant fraction of human tumors retain persistently high levels of ARF, suggesting that ARF may possess a prosurvival function. We show that ARF protein is markedly up-regulated in cells exposed to nutrient starvation. Cells with silenced ARF show reduced autophagy and reduced viability when placed under conditions of starvation. We show for the first time that ARF silencing can limit the progression of some tumors, such as lymphoma, but not others, such as E1A/Ras-induced tumors. Specifically, myc-driven lymphomas with mutant p53 tend to overexpress ARF; we show that silencing ARF in these tumors greatly impedes their progression. These data are the first to show that ARF can act in a p53-independent manner to promote the progression of some tumors. [Cancer Res 2008;68(23):9608–13]
Data from Enterococcus faecalis Induces Aneuploidy and Tetraploidy in Colonic Epithelial Cells through a Bystander Effect
Authors: Wang Xingmin , Allen Toby D. , May Randal J. , Lightfoot Stanley , Houchen Courtney W. , Huycke Mark M.
Published: March 2024

Abstract
Intestinal commensals are potential important contributors to the etiology of sporadic colorectal cancer, but mechanisms by which bacteria can initiate tumors remain uncertain. Herein, we describe mechanisms that link Enterococcus faecalis, a bacterium known to produce extracellular superoxide, to the acute induction of chromosomal instability. Immortalized human and nontransformed murine colonic epithelial cells, along with a mouse colonic ligation model, were used to assess the effect of E. faecalis on genomic DNA stability and damage. We found that this human intestinal commensal generated aneuploidy, tetraploidy, and γH2AX foci in HCT116, RKO, and YAMC cells. In addition, direct exposure of E. faecalis to these cells induced a G2 cell cycle arrest. Similar observations were noted by exposing cells to E. faecalis–infected macrophages in a dual-chamber coculture system for detecting bystander effects. Manganese superoxide dismutase, catalase, and tocopherols attenuated, and caffeine and inhibitors of glutathione synthase exacerbated, the aneugenic effects and linked the redox-active phenotype of this intestinal commensal to potentially transforming events. These findings provide novel insights into mechanisms by which E. faecalis and intestinal commensals can contribute to cellular transformation and tumorigenesis. [Cancer Res 2008;68(23):9909–17]
Data from Systemic Targeting of CpG-ODN to the Tumor Microenvironment with Anti–neu-CpG Hybrid Molecule and T Regulatory Cell Depletion Induces Memory Responses in BALB-neuT Tolerant Mice
Authors: Sharma Sanjay , Dominguez Ana Lucia , Manrique Soraya Zorro , Cavallo Federica , Sakaguchi Shimon , Lustgarten Joseph
Published: March 2024

Abstract
We have shown that neu transgenic mice are immunotolerant and that immunizations with dendritic cells (DC) pulsed with neu-derived antigens were not able to control tumor growth in these animals. We tested whether, by modulating the tumor microenvironment with Toll-like receptor ligands, it could be possible to induce the activation of antitumor responses in neu mice. Our results indicate that only intratumoral (i.t.) injections of CpG-ODN induce an antitumor response in neu mice. To target the CpG-ODN to the tumor site anywhere within the body, we chemically conjugated an anti–Her-2/neu monoclonal antibody (mAb) with CpG-ODN. The anti–neu-CpG hybrid molecule retained its ability to bind to Her-2/neu+ tumors, activate DCs, and induce antitumor responses. Our results indicated that injections of anti–neu-CpG induced the rejection of primary tumors in 100% of BALB/c mice and only in ∼30% of BALB-neuT mice. After challenging the BALB/c and BALB-neuT mice, we observed that BALB/c mice developed a protective memory response; in contrast, BALB-neuT mice succumbed to the challenge. After injections of anti–neu-CpG, T regulatory cells (T-reg) were drastically reduced at the tumor site, but a large number were still present in the lymphoid organs. When BALB-neuT mice were treated with anti–neu-CpG plus anti-GITR mAb, but not with anti-CD25 mAb, 100% of the BALB-neuT mice rejected the primary tumor and developed a protective memory response indicating the critical role of T-regs in regulating the repertoire against self antigens. Taken together, these results indicate that CpG-ODN–targeted therapy and depletion of T-regs optimally activate a primary response and generate a protective memory response against self-tumor antigens. [Cancer Res 2008;68(18):7530–40]
Data from Activation of Signal Transducer and Activator of Transcription 3 through a Phosphomimetic Serine 727 Promotes Prostate Tumorigenesis Independent of Tyrosine 705 Phosphorylation
Authors: Qin Haiyan R. , Kim Han-Jong , Kim Joon-Young , Hurt Elaine M. , Klarmann George J. , Kawasaki Brian T. , Duhagon Serrat Maria A. , Farrar William L.
Published: March 2024

Abstract
Aberrantly activated signal transducer and activator of transcription 3 (Stat3) is implicated in the development of various human cancers. Y705 phosphorylation is conventionally thought to be required for Stat3 signal-dependent activation and seems to play an essential role in some malignancies. Recently, it was shown that Stat3 is activated through novel and noncanonical mechanisms, including phosphorylation at S727. Here, we investigate S727 phosphorylation of Stat3 and its subsequent effects in prostate cancer development, independent of Y705 phosphorylation, using mutated Stat3 in the human prostate cancer cell line LNCaP. We show mutation of S727 to the phosphomimetic residue Glu, and inactivation of Y705 (Y705F/S727E) resulted in a remarkable growth advantage in low-serum, enhanced anchorage-independent growth in soft agar, and increased tumorigenicity in nonobese diabetic/severe combined immunodeficient (NOD/SCID) mice, possibly by direct activation of downstream proto-oncogenes c-myc, mcl-1, and survivin. Y705F/S727E mutant cells were more invasive than Y705F/S727A (inactivation of Y705 and S727) mutant cells, and more Y705F/S727E mutant Stat3 was localized in the nuclei relative to Y705F/S727A mutant Stat3 at the steady state. Furthermore, the Y705F/S727E but not the Y705F/S727A mutant induced anchorage-independent growth of noncancerous prostate epithelial cells (RWPE-1). We further show that Stat3 is phosphorylated at S727 in 65% of malignant prostate tissues (n = 20) relative to 25% of normal prostate tissues (n = 4). Moreover, there is a positive correlation between phosphoS727-Stat3 expression and Gleason score in these prostate cancer tissues (P = 0.05). Our data suggest for the first time that S727 phosphorylation is sufficient to activate Stat3, thereby driving prostate tumorigenesis independent of Y705 phosphorylation. [Cancer Res 2008;68(19):7736–41]
Data from AKT1 Inhibits Homologous Recombination by Inducing Cytoplasmic Retention of BRCA1 and RAD51
Authors: Plo Isabelle , Laulier Corentin , Gauthier Laurent , Lebrun Fabienne , Calvo Fabien , Lopez Bernard S.
Published: March 2024

Abstract
AKT1 is frequently up-regulated in sporadic breast cancer, whereas BRCA1 is frequently mutated in familial breast cancer. Because BRCA1 is involved in homologous recombination (HR), we addressed whether AKT1 also has an effect on this process. We showed that AKT1 repressed HR through cytoplasmic retention of BRCA1 and RAD51 proteins, resulting in a BRCA1-deficient–like phenotype. This process does not require direct BRCA1 phosphorylation by AKT1. The cytoplasmic retention of BRCA1 and RAD51 correlated with activated AKT1 in tumor cell lines and in biopsies from sporadic breast cancers. Under nonpathologic conditions, fibroblast growth factor, which activates AKT1 and stimulates proliferation in fibroblasts, impaired excessive HR without fully inhibiting it, promoting genome stability. Our study reveals that the regulation of BRCA1 and RAD51 is altered in a high frequency of sporadic breast cancers and highlights the role of extracellular AKT signaling-dependent regulation of HR and genome stability. [Cancer Res 2008;68(22):9404–12]