bluuebunny/crossref_metadata_2025 huggingface dataset:
Retinoic Acid Induces Neuroblastoma Cell Death by Inhibiting Proteasomal Degradation of Retinoic Acid Receptor α
Authors: Nagai Jun-ichi , Yazawa Takuya , Okudela Koji , Kigasawa Hisato , Kitamura Hitoshi , Osaka Hitoshi
Published: September 2005

To seek a novel therapeutic approach to neuroblastoma (NBL), we used three NBL cell lines (SK-N-DZ, NH12, and SK-N-SH) to examine the underlining molecular mechanisms of cellular reactions and sensitivity to all-trans-retinoic acid (ATRA). SK-N-DZ cells expressed relatively high levels of retinoic acid receptor α (RAR-α) and underwent ATRA-induced cell death that was blocked by an RAR-α antagonist. By contrast, RAR-α expression gradually decreased in NH12 and SK-N-SH cells, which did not experience increased cell death in response to ATRA. We report here the ubiquitin-dependent down-regulation of RAR-α expression during ATRA treatment. Our data suggest that SK-N-DZ cells have a defect in RAR-α down-regulation, resulting in sustained high expression of RAR-α that confers high sensitivity to ATRA. Accordingly, treatment with a proteasome inhibitor dramatically increased ATRA-induced cell death in NH12 and SK-N-SH cell lines. Our results reveal the crucial involvement of the RAR-α signaling pathway in NBL cell death and show that three NBL cell lines are differentially sensitive to ATRA. These data suggest a potential novel therapy for NBL involving retinoic acid treatment combined with the inhibition of RAR-α degradation.
Frequent Nonrandom Activation of Germ-Line Genes in Human Cancer
Authors: Koslowski Michael , Bell Carolin , Seitz Gerhard , Lehr Hans-Anton , Roemer Klaus , Müntefering Horst , Huber Christoph , Sahin Ugur , Türeci Özlem
Published: September 2005

The growing class of cancer/germ-line genes is characterized by a unique expression pattern with transcription restricted to germ cells and cancer cells. It is not known which fraction of germ-line genes is ectopically activated in tumor cells and whether this fraction displays common features as compared with strictly germ-line genes remaining silent in cancer. Using an unbiased genome-wide scanning approach, representative samples of both cancer/germ-line genes as well as strictly germ-line-specific genes were determined. Comparative analysis disclosed highly significant diametric characteristics for these two categories of genes with regard to sex specificity, developmental stage of physiological expression during gametogenesis, chromosomal localization, and epigenetic regulation of expression. Our findings provide class predictors for germ cell-specific gene activation in cancer. The identification of highly congruent expression patterns in cancer and in DNA methyltransferase-deficient cells suggests an underlying common epigenetic mechanism for activation of germ-line genes in cancer.
p53 Upregulates Death Receptor 4 Expression through an Intronic p53 Binding Site
Authors: Liu Xiangguo , Yue Ping , Khuri Fadlo R. , Sun Shi-Yong
Published: September 2005

Death receptor 4 (DR4) is one of the tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) receptors and triggers apoptosis on ligation with TRAIL or overexpression. Our previous study demonstrated that DR4 expression could be regulated in a p53-dependent fashion. In the present study, we have demonstrated that DR4 is a p53 target gene and is regulated by p53 through a functional intronic p53 binding site (p53BS) based on the following lines of evidence: (a) the p53BS in the DR4 gene is almost identical to the one found in the first intron of the DR5 gene in terms of their locations and sequences; (b) DR4 p53BS bound to p53 protein in intact cells upon p53 activation as demonstrated by a chromatin immunoprecipitation assay; (c) a luciferase reporter vector carrying the DR4 p53BS upstream of an SV40 promoter exhibited enhanced luciferase activity when transiently cotransfected with a wild-type p53 expression vector in p53-null cell lines or stimulated with DNA-damaging agents in a cell line having wild-type p53; and (d) when the DR4 p53BS, together with its own corresponding promoter region in the same orientation as it sits in its natural genomic locus, was cloned into a basic luciferase vector without a promoter element, its transcriptional activity was strikingly increased by cotransfection of a wild-type p53 expression vector or treatment with DNA-damaging agents. However, wild-type p53 or DNA-damaging agents completely lost their activity to increase transcriptional activity of a reporter construct with deleted DR4 p53BS. Thus, we conclude that p53 directly regulates the expression of the DR4 gene via the novel intronic p53BS.
Combination Therapy with Conditionally Replicating Adenovirus and Replication Defective Adenovirus
Authors: Lee Choon-Taek , Park Kyung-Ho , Yanagisawa Kiyoshi , Adachi Yasushi , Ohm Joyce E. , Nadaf Sorena , Dikov Mikhail M. , Curiel David T. , Carbone David P.
Published: September 2005

Low gene transfer rate is the most substantial hurdle in the practical application of gene therapy. One strategy to improve transfer efficiency is the use of a conditionally replicating adenovirus (CRAD) that can selectively replicate in tumor cells. We hypothesized that conventional E1-deleted adenoviruses (ad) can become replication-competent when cotransduced with a CRAD to selectively supply E1 in trans in tumors. The resulting selective production of large numbers of the E1-deleted ad within the tumor mass will increase the transduction efficiency. We used a CRAD (Δ24RGD) that produces a mutant E1 without the ability to bind retinoblastoma but retaining viral replication competence in cancer cells with a defective pRb/p16. Ad-lacZ, adenovirus-luciferase (ad-luc), and adenovirus insulin-like growth factor-1R/dominant-negative (ad-IGF-1R/dn; 482, 950) are E1-deleted replication-defective adenoviruses. The combination of CRAD and ad-lacZ increased the transduction efficiency of lacZ to 100% from 15% observed with ad-lacZ alone. Transfer of media of CRAD and ad-lacZ cotransduced cells induced the transfer of lacZ (media transferable bystander effect). Combination of CRAD and ad-IGF-1R/dn increased the production of truncated IGF-1R or soluble IGF-1R > 10 times compared with transduction with ad-IGF-1R/dn alone. Combined intratumoral injection of CRAD and ad-luc increased the luciferase expression about 70 times compared with ad-luc alone without substantial systemic spread. Combined intratumoral injection of CRAD and ad-IGF-1R/482 induced stronger growth suppression of established lung cancer xenografts than single injections. The combination of CRAD and E1-deleted ad induced tumor-specific replication of CRAD and E1-deleted ad and increased the transduction rate and therapeutic efficacy of these viruses in model tumors.
c-Jun NH2-Terminal Kinase-Mediated Up-regulation of Death Receptor 5 Contributes to Induction of Apoptosis by the Novel Synthetic Triterpenoid Methyl-2-Cyano-3,12-Dioxooleana-1, 9-Dien-28-Oate in Human Lung Cancer Cells
Authors: Zou Wei , Liu Xiangguo , Yue Ping , Zhou Zhongmei , Sporn Michael B. , Lotan Reuben , Khuri Fadlo R. , Sun Shi-Yong
Published: September 2005

Death receptor (DR) 4 or 5, on binding to its ligand, tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), triggers apoptosis via activating the caspase-8–mediated caspase cascade. Certain anticancer drugs up-regulate the expression of these receptors and thereby induce apoptosis or enhance TRAIL-induced apoptosis. In this study, we explored the ability of methyl-2-cyano-3,12-dioxooleana-1,9-dien-28-oate (CDDO-Me) to activate the extrinsic DR-mediated apoptotic pathway in human lung cancer cells. We found that CDDO-Me not only activated caspase-8 but also induced expression of DRs, particularly DR5, in a p53-independent mechanism. Correspondingly, CDDO-Me augmented TRAIL-induced apoptosis in these cells regardless of p53 status as evidenced by enhanced DNA fragmentation and activation of caspase cascades, suggesting that CDDO-Me–induced DRs are functionally active. Moreover, silencing of DR5 expression using small interfering RNA suppressed apoptosis induced by CDDO-Me alone or by combination of CDDO-Me and TRAIL, indicating that DR5 up-regulation is required for induction of apoptosis by CDDO-Me and for enhancement of TRAIL-induced apoptosis by CDDO-Me. CDDO-Me rapidly activated c-Jun NH2-terminal kinase (JNK) before DR up-regulation and caspase-8 activation. Moreover, application of the JNK-specific inhibitor SP600125 blocked CDDO-Me–induced increases in JNK activation, DR up-regulation, caspase-8 activation, and DNA fragmentation. These results show that activation of JNK pathway results in CDDO-Me–induced DR up-regulation, caspase-8 activation, and apoptosis. Collectively, we conclude that CDDO-Me induces apoptosis via the JNK-mediated DR up-regulation in human lung cancer cells.
Aberrant Expression of the Tyrosine Kinase Receptor EphA4 and the Transcription Factor Twist in Sézary Syndrome Identified by Gene Expression Analysis
Authors: van Doorn Remco , Dijkman Remco , Vermeer Maarten H. , Out-Luiting Jacoba J. , van der Raaij-Helmer Elisabeth M. H. , Willemze Rein , Tensen Cornelis P.
Published: September 2005

Sézary syndrome (Sz) is a malignancy of CD4+ memory skin-homing T cells and presents with erythroderma, lymphadenopathy, and peripheral blood involvement. To gain more insight into the molecular features of Sz, oligonucleotide array analysis was performed comparing gene expression patterns of CD4+ T cells from peripheral blood of patients with Sz with those of patients with erythroderma secondary to dermatitis and healthy controls. Using unsupervised hierarchical clustering gene, expression patterns of T cells from patients with Sz were classified separately from those of benign T cells. One hundred twenty-three genes were identified as significantly differentially expressed and had an average fold change exceeding 2. T cells from patients with Sz demonstrated decreased expression of the following hematopoietic malignancy-linked tumor suppressor genes: TGF-β receptor II, Mxi1, Riz1, CREB-binding protein, BCL11a, STAT4, and Forkhead Box O1A. Moreover, the tyrosine kinase receptor EphA4 and the potentially oncogenic transcription factor Twist were highly and selectively expressed in T cells of patients with Sz. High expression of EphA4 and Twist was also observed in lesional skin biopsy specimens of a subset of patients with cutaneous T cell lymphomas related to Sz, whereas their expression was nearly undetectable in benign T cells or in skin lesions of patients with inflammatory dermatoses. Detection of EphA4 and Twist may be used in the molecular diagnosis of Sz and related cutaneous T-cell lymphomas. Furthermore, the membrane-bound EphA4 receptor may serve as a target for directed therapeutic intervention.
Dendritic Cell Subsets Differentially Regulate Angiogenesis in Human Ovarian Cancer
Authors: Curiel Tyler J. , Cheng Pui , Mottram Peter , Alvarez Xavier , Moons Lieve , Evdemon-Hogan Melina , Wei Shuang , Zou Linhua , Kryczek Ilona , Hoyle Gary , Lackner Andrew , Carmeliet Peter , Zou Weiping
Published: September 2005

Angiogenesis is essential for both primary and metastatic tumor growth. Tumor blood vessel formation is complex and regulated by many factors. Ovarian carcinomas have a poor prognosis, often associated with multifocal intraperitoneal dissemination accompanied by intense neovascularization. To examine tumor angiogenesis in the tumor microenvironment, we studied malignant ascites of patients with untreated ovarian carcinoma. We observed high numbers of plasmacytoid dendritic cells (PDCs) and significant stromal-derived factor (CXCL-12/SDF)-1 in their malignant ascites, attracting PDCs into the tumor environment. We now show that tumor-associated PDCs induced angiogenesis in vivo through production of tumor necrosis factor α and interleukin 8. By contrast, myeloid dendritic cells (MDCs) were absent from malignant ascites. MDCs derived in vitro suppressed angiogenesis in vivo through production of interleukin 12. Thus, the tumor may attract PDCs to augment angiogenesis while excluding MDCs to prevent angiogenesis inhibition, demonstrating a novel mechanism for modulating tumor neovascularization. Because dendritic cells (DCs) have long been known to affect tumor immunity, our data also implicate DCs in regulation of tumor neoangiogenesis, suggesting a novel role of DCs in tumor pathology.
The Role of DNA Polymerase η in Translesion Synthesis Past Platinum–DNA Adducts in Human Fibroblasts
Authors: Bassett Ekaterina , King Nicole M. , Bryant Miriam F. , Hector Suzanne , Pendyala Lakshmi , Chaney Stephen G. , Cordeiro-Stone Marila
Published: September 2005

Cisplatin, a widely used chemotherapeutic agent, has been implicated in the induction of secondary tumors in cancer patients. This drug is presumed to be mutagenic because of error-prone translesion synthesis of cisplatin adducts in DNA. Oxaliplatin is effective in cisplatin-resistant tumors, but its mutagenicity in humans has not been reported. The polymerases involved in bypass of cisplatin and oxaliplatin adducts in vivo are not known. DNA polymerase η is the most efficient polymerase for bypassing platinum adducts in vitro. We evaluated the role of polymerase η in translesion synthesis past platinum adducts by determining cytotoxicity and induced mutation frequencies at the hypoxanthine guanine phosphoribosyltransferase (HPRT) locus in diploid human fibroblasts. Normal human fibroblasts (NHF1) were compared with xeroderma pigmentosum variant (XPV) cells (polymerase η-null) after treatment with cisplatin. In addition, XPV cells complemented for polymerase η expression were compared with the isogenic cells carrying the empty expression vector. Cytotoxicity and induced mutagenicity experiments were measured in parallel in UVC-irradiated fibroblasts. We found that equitoxic doses of cisplatin induced mutations in fibroblasts lacking polymerase η at frequencies 2- to 2.5-fold higher than in fibroblasts with either normal or high levels of polymerase η. These results indicate that polymerase η is involved in error-free translesion synthesis past some cisplatin adducts. We also found that per lethal event, cisplatin was less mutagenic than UVC. Treatment with a wide range of cytotoxic doses of oxaliplatin did not induce mutations above background levels in cells either expressing or lacking polymerase η, suggesting that oxaliplatin is nonmutagenic in human fibroblasts.
HOXB13 Induces Growth Suppression of Prostate Cancer Cells as a Repressor of Hormone-Activated Androgen Receptor Signaling
Authors: Jung Chaeyong , Kim Ran-Sook , Zhang Hong-Ji , Lee Sang-Jin , Jeng Meei-Huey
Published: September 2005

Androgen receptor (AR) signals play a decisive role in regulating the growth and differentiation of both normal and cancerous prostate cells by triggering the regulation of target genes, in a process in which AR cofactors have critical functions. Because of the highly prostate-specific expression pattern of HOXB13, we studied the role of this homeodomain protein in prostate cells. Expression of HOXB13 was limited to AR-expressing prostate cells. Reporter transcription assay demonstrated that HOXB13 significantly suppressed hormone-mediated AR activity in a dose-responsive manner, and suppression was specific to AR with which HOXB13 physically interacts. Overexpression of HOXB13 further down-regulated the androgen-stimulated expression of prostate-specific antigen, and suppression of endogenous HOXB13 stimulated transactivation of AR. Functionally, HOXB13 suppressed growth of LNCaP prostate cancer cells, which could be counteracted by additional hormone-activated AR. On the other hand, the growth-suppressive function of HOXB13 in AR-negative CV-1 cells was not affected by AR. These results suggest that HOXB13 functions as an AR repressor to modulate the complex AR signaling and subsequent growth regulation of prostate cancer cells. In addition to the loss of HOXB13 expression, maintaining AR may be an important step for prostate cancer cells to tolerate the suppressor function of HOXB13. Altogether, our data present a novel mechanism for the HOXB13-mediated repression of AR signaling, which can be interpreted to a growth-suppressive event.
Visualization of Intrathoracically Disseminated Solid Tumors in Mice with Optical Imaging by Telomerase-Specific Amplification of a Transferred Green Fluorescent Protein Gene
Authors: Umeoka Tatsuo , Kawashima Takeshi , Kagawa Shunsuke , Teraishi Fuminori , Taki Masaki , Nishizaki Masahiko , Kyo Satoru , Nagai Katsuyuki , Urata Yasuo , Tanaka Noriaki , Fujiwara Toshiyoshi
Published: September 2005

Currently available methods for detection of tumors in vivo such as X-ray, computed tomography, and ultrasonography are noninvasive and have been well studied; the images, however, are not specific for tumors. Direct optical imaging of tumor cells in vivo that can clearly distinguish them from surrounding normal tissues may be clinically useful. Here, we describe a new approach to visualizing tumors whose fluorescence can be detected using tumor-specific replication-competent adenovirus (OBP-301, Telomelysin) in combination with Ad-GFP, a replication-deficient adenovirus expressing green fluorescent protein (GFP). Human telomerase reverse transcriptase is the catalytic subunit of telomerase, which is highly active in cancer cells but quiescent in most normal somatic cells. We constructed an adenovirus 5 vector in which the human telomerase reverse transcriptase promoter element drives expression of E1A and E1B genes linked with an internal ribosome entry site and showed that OBP-301 replicated efficiently in human cancer cells, but not in normal cells such as human fibroblasts. When the human lung and colon cancer cell lines were infected with Ad-GFP at a low multiplicity of infection, GFP expression could not be detected under a fluorescence microscope; in the presence of OBP-301, however, Ad-GFP replicated in these tumor cells and showed strong green signals. In contrast, coinfection with OBP-301 and Ad-GFP did not show any signals in normal cells such as fibroblasts and vascular endothelial cells. We also found that established subcutaneous tumors could be visualized after intratumoral injection of OBP-301 and Ad-GFP. A549 human lung tumors and SW620 human colon tumors transplanted into BALB/c nu/nu mice were intratumorally injected with 8 × 105 plaque-forming units of Ad-GFP in combination with 8 × 106 plaque-forming units of OBP-301. Within 3 days of treatment, the fluorescence of the expressed GFP became visible by a three-chip color cooled charged-coupled device camera in these tumors, whereas intratumoral injection of Ad-GFP alone could not induce GFP fluorescence. Moreover, intrathoracic administration of Ad-GFP and OBP-301 could visualize disseminated A549 tumor nodules in mice after intrathoracic implantation. Our results indicate that intratumoral or intrathoracic injection of Ad-GFP in combination with OBP-301 might be a useful diagnostic method that provides a foundation for future clinical application.
Genetic Pathways to Glioblastoma
Authors: Ohgaki Hiroko , Dessen Pierre , Jourde Benjamin , Horstmann Sonja , Nishikawa Tomofumi , Di Patre Pier-Luigi , Burkhard Christoph , Schüler Danielle , Probst-Hensch Nicole M. , Maiorka Paulo César , Baeza Nathalie , Pisani Paola , Yonekawa Yasuhiro , Yasargil M. Gazi , Lütolf Urs M. , Kleihues Paul
Published: September 2005

We conducted a population-based study on glioblastomas in the Canton of Zurich, Switzerland (population, 1.16 million) to determine the frequency of major genetic alterations and their effect on patient survival. Between 1980 and 1994, 715 glioblastomas were diagnosed. The incidence rate per 100,000 population/year, adjusted to the World Standard Population, was 3.32 in males and 2.24 in females. Observed survival rates were 42.4% at 6 months, 17.7% at 1 year, and 3.3% at 2 years. For all of the age groups, younger patients survived significantly longer, ranging from a median of 8.8 months (<50 years) to 1.6 months (>80 years). Loss of heterozygosity (LOH) 10q was the most frequent genetic alteration (69%), followed by EGFR amplification (34%), TP53 mutations (31%), p16INK4a deletion (31%), and PTEN mutations (24%). LOH 10q occurred in association with any of the other genetic alterations and was predictive of shorter survival. Primary (de novo) glioblastomas prevailed (95%), whereas secondary glioblastomas that progressed from low-grade or anaplastic gliomas were rare (5%). Secondary glioblastomas were characterized by frequent LOH 10q (63%) and TP53 mutations (65%). Of the TP53 mutations in secondary glioblastomas, 57% were in hotspot codons 248 and 273, whereas in primary glioblastomas, mutations were more equally distributed. G:C→A:T mutations at CpG sites were more frequent in secondary than primary glioblastomas (56% versus 30%; P = 0.0208). This suggests that the acquisition of TP53 mutations in these glioblastoma subtypes occurs through different mechanisms.
Inhibition ofSmoothenedSignaling Prevents Ultraviolet B-Induced Basal Cell Carcinomas through Regulation of Fas Expression and Apoptosis
Authors: Athar Mohammad , Li Chengxin , Tang Xiuwei , Chi Sumin , Zhang Xiaoli , Kim Arianna L. , Tyring Stephen K. , Kopelovich Levy , Hebert Jennifer , Epstein Ervin H. , Bickers David R. , Xie Jingwu
Published: September 2005

Abnormal activation of the hedgehog-signaling pathway is the pivotal abnormality driving the growth of basal cell carcinomas (BCCs), the most common type of human cancer. Antagonists of this pathway such as cyclopamine may therefore be useful for treatment of basal cell carcinomas and other hedgehog-driven tumors. We report here that chronic oral administration of cyclopamine dramatically reduces (∼66%) UVBinduced basal cell carcinoma formation in Ptch1+/− mice. Fas expression is low in human and murine basal cell carcinomas but is up-regulated in the presence of the smoothened (SMO) antagonist, cyclopamine, both in vitro in the mouse basal cell carcinoma cell line ASZ001 and in vivo after acute treatment of mice with basal cell carcinomas. This parallels an elevated rate of apoptosis. Conversely, expression of activated SMO in C3H10T1/2 cells inhibits Fas expression. Fas/Fas ligand interactions are necessary for cyclopamine-mediated apoptosis in these cells, a process involving caspase-8 activation. Our data provide strong evidence that cyclopamine and perhaps other SMO antagonists are potent in vivo inhibitors of UVB-induced basal cell carcinomas in Ptch1+/− mice and likely in humans because the majority of human basal cell carcinomas manifest mutations in PTCH1 and that a major mechanism of their inhibitory effect is through up-regulation of Fas, which augments apoptosis.
Deregulated Akt3 Activity Promotes Development of Malignant Melanoma
Authors: Stahl Jill M. , Sharma Arati , Cheung Mitchell , Zimmerman Melissa , Cheng Jin Q. , Bosenberg Marcus W. , Kester Mark , Sandirasegarane Lakshman , Robertson Gavin P.
Published: September 2005

Malignant melanoma is the skin cancer with the most significant impact on man, carrying the highest risk of death from metastasis. Both incidence and mortality rates continue to rise each year, with no effective long-term treatment on the horizon. In part, this reflects lack of identification of critical genes involved and specific therapies targeted to correct these defects. We report that selective activation of the Akt3 protein promotes cell survival and tumor development in 43 to 60% of nonfamilial melanomas. The predominant Akt isoform active in melanomas was identified by showing that small interfering RNA (siRNA) against only Akt3, and not Akt1 or Akt2, lowered the amount of phosphorylated (active) Akt in melanoma cells. The amount of active Akt3 increased progressively during melanoma tumor progression with highest levels present in advanced-stage metastatic melanomas. Mechanisms of Akt3 deregulation occurred through a combination of overexpression of Akt3 accompanying copy number increases of the gene and decreased PTEN protein function occurring through loss or haploinsufficiency of the PTEN gene. Targeted reduction of Akt3 activity with siRNA or by expressing active PTEN protein stimulated apoptotic signaling, which reduced cell survival by increasing apoptosis rates thereby inhibiting melanoma tumor development. Identifying Akt3 as a selective target in melanoma cells provides new therapeutic opportunities for patients in the advanced stages of this disease.
Functional Integrity of Nuclear Factor κB, Phosphatidylinositol 3′-Kinase, and Mitogen-Activated Protein Kinase Signaling Allows Tumor Necrosis Factor α-Evoked Bcl-2 Expression to Provoke Internal Ribosome Entry Site-Dependent Translation of Hypoxia-Inducible Factor 1α
Authors: Zhou Jie , Callapina Melvin , Goodall Gregory J. , Brüne Bernhard
Published: September 2005

Hypoxia-inducible factor (HIF)-1, a heterodimeric transcription factor composed of HIF-1α and HIF-1β subunits coordinates pathophysiologic responses toward decreased oxygen availability. It is now appreciated that enhanced protein translation of HIF-1α under normoxia accounts for an alternative regulatory circuit to activate HIF-1 by hormones, growth factors, or cytokines such as tumor necrosis factor α (TNF-α). Here, we aimed at understanding molecular details of HIF-1α translation in response to TNF-α. In tubular LLC-PK1 cells, activation of nuclear factor κB (NFκB) by TNF-α resulted in HIF-1α protein synthesis as determined by [35S]methionine pulse experiments. Protein synthesis was attenuated by blocking NFκB, phosphatidylinositol 3′-kinase (PI3k), and mitogen-activated protein kinase (MAPK). Use of a dicistronic reporter with the HIF-1α 5′-untranslated region (5′UTR) between two coding regions indicated that TNF-α promoted an internal ribosome entry site (IRES) rather than a cap-dependent translation. IRES-mediated translation required the functional integrity of the NFκB, PI3k, and MAPK signaling pathways. Although no signal cross-talk was noticed between NFκB, PI3k, and MAPK signaling, these pathways are needed to up-regulate the anti-apoptotic target protein Bcl-2 by TNF-α. Expression of Bcl-2 provoked not only IRES-dependent translation but also HIF-1α protein synthesis. We conclude that Bcl-2 functions as an important determinant in facilitating HIF-1α protein expression by TNF-α via an IRES-dependent translational mechanism. These observations suggest a link between Bcl-2 and HIF-1α expression, a situation with potential relevance to cancer biology.
Crucial Role for Interferon γ in the Synergism between Tumor Vasculature-Targeted Tumor Necrosis Factor α (NGR-TNF) and Doxorubicin
Authors: Sacchi Angelina , Gasparri Anna , Curnis Flavio , Bellone Matteo , Corti Angelo
Published: September 2005

NGR-TNF is a derivative of TNF-α, consisting of TNF fused to CNGRCG, a tumor vasculature-targeting peptide. Previous studies showed that NGR-TNF can exert synergistic antitumor effects with doxorubicin and with other chemotherapeutic drugs in murine models. In this study, we have investigated the role of endogenous IFN-γ on the antitumor activity of NGR-TNF in combination with doxorubicin. The study was carried out using murine B16F1 melanoma and TS/A mammary adenocarcinoma implanted subcutaneously in (a) immunocompetent mice, (b) athymic nude mice, and (c) IFN-γ–knockout mice. Synergism between NGR-TNF and doxorubicin was observed in immunocompetent mice but not in nude or IFN-γ–knockout mice. Preadministration of a neutralizing anti-IFN-γ antibody to immunocompetent mice inhibited the NGR-TNF/doxorubicin synergism, whereas administration of IFN-γ to nude and to IFN-γ–knockout mice restored the synergistic activity. The synergism in nude mice was restored also by transfecting tumor cells with the IFN-γ cDNA. Administration of NGR-TNF in combination with IFN-γ to nude mice, but not of NGR-TNF alone, doubled the penetration of doxorubicin in TS/A tumors. These findings point to a crucial role for locally produced IFN-γ in tumor vascular targeting with NGR-TNF and doxorubicin. Finally, addition of IFN-γ to the treatment of immunocompetent mice with NGR-TNF/doxorubicin induced only modest improvement in response, suggesting that exogenous IFN-γ can improve the therapeutic activity of these drugs only in case of suboptimal production of endogenous IFN-γ.
Notch1
and
Notch2
Have Opposite Effects on Embryonal Brain Tumor Growth
Authors: Fan Xing , Mikolaenko Irina , Elhassan Ihab , Ni XingZhi , Wang Yunyue , Ball Douglas , Brat Daniel J. , Perry Arie , Eberhart Charles G.
Published: September 2005

The role of Notch signaling in tumorigenesis can vary; Notch1 acts as an oncogene in some neoplasms, and a tumor suppressor in others. Here, we show that different Notch receptors can have opposite effects in a single tumor type. Expression of truncated, constitutively active Notch1 or Notch2 in embryonal brain tumor cell lines caused antagonistic effects on tumor growth. Cell proliferation, soft agar colony formation, and xenograft growth were all promoted by Notch2 and inhibited by Notch1. We also found that Notch2 receptor transcripts are highly expressed in progenitor cell-derived brain tumors such as medulloblastomas, whereas Notch1 is scarce or undetectable. This parallels normal cerebellar development, during which Notch2 is predominantly expressed in proliferating progenitors and Notch1 in postmitotic differentiating cells. Given the oncogenic effects of Notch2, we analyzed its gene dosage in 40 embryonal brain tumors, detecting an increased copy number in 15% of cases. Notch2 gene amplification was confirmed by fluorescence in situ hybridization in one case with extremely high Notch2 mRNA levels. In addition, expression of the Notch pathway target gene Hes1 in medulloblastomas was associated with significantly shorter patient survival (P = 0.01). Finally, pharmacological inhibition of Notch signaling suppresses growth of medulloblastoma cells. Our data indicate that Notch1 and Notch2 can have opposite effects on the growth of a single tumor type, and show that Notch2 can be overexpressed after gene amplification in human tumors.
Spontaneous Mutations in Digestive Tract of Old Mice Show Tissue-Specific Patterns of Genomic Instability
Authors: Ono Tetsuya , Ikehata Hironobu , Pithani Vishnu Priya , Uehara Yoshihiko , Chen Yali , Kinouchi Yoshitaka , Shimosegawa Toru , Hosoi Yoshio
Published: September 2005

In an attempt to evaluate the possible role of mutations in the age-dependent increase of tumor incidence, we studied the mutational burden that accumulates in the aging process in different parts of the digestive tract in mice. The mutations were monitored in lacZ genes integrated in the mouse genome. The digestive tract was divided into the esophagus, stomach, proximal, medial, and distal part of the small intestine, and the colon. Epithelial tissues were separated from these tissues with the exception of the esophagus, in which case the whole tissue was examined. At a young age, the mutant frequencies as well as the molecular nature of the mutations were similar among the tissues examined. In old age, on the other hand, mutant frequencies were elevated to different degrees among the tissues; they were high in the small intestine and colon, intermediate in the stomach, and low in the esophagus. The molecular characteristics of the mutations also revealed distinct tissue-specificity; there were elevated rates of a small deletion mutation in the esophagus, G:C to T:A transversion in the proximal small intestine, and multiple mutations in the distal small intestine and colon. The results indicate that different parts of the digestive tract suffer from different kinds of mutational stress in the aging process. The nature of the multiple mutations suggests the presence of a mutator phenotype based on an imbalance in deoxyribonucleotide pools.
A Novel Role for the Coxsackie Adenovirus Receptor in Mediating Tumor Formation by Lung Cancer Cells
Authors: Qin Min , Escuadro Brian , Dohadwala Mariam , Sharma Sherven , Batra Raj K.
Published: September 2005

The Coxsackie Adenovirus Receptor (CAR) has primarily been studied in its role as the initial cell surface attachment receptor for Coxsackie and group C adenoviruses. Recent reports suggest that CAR mediates homotypic intercellular adhesion as part of the tight and/or adherens junction. Thus, CAR is well positioned to participate in intercellular interactions and signaling. Using an antisense (AS)-CAR plasmid vector, we silenced surface CAR expression in lung cancer cells that possessed a high basal expression of this molecule and monitored the resultant tumorigenesis. AS-CAR transfectants exhibit a profound loss in the ability to generate xenografts in scid/scid mice. The emergence of delayed-onset tumors in animals that received injection with AS-CAR transfectants correlates with the resurfacing of CAR expression, suggesting that such expression and tumor emergence are temporally related. To study the mechanism underlying the differences in tumorigenicity, control and AS-CAR cells were compared in terms of their in vitro growth potential. Whereas only subtle differences in the proliferative capacity of the two populations were evident when assayed with growth on plastic, significant differences became apparent when one compared the relative ability of these populations to form colonies in soft agar. These data indicate that silencing surface CAR expression abrogates xenograft tumorigenesis in vivo and colony formation in vitro and invoke the novel possibility that CAR expression is needed for the efficient formation of tumors by a subset of lung cancer cells.
Tumor Cell-Specific
BRCA1
and
RASSF1A
Hypermethylation in Serum, Plasma, and Peritoneal Fluid from Ovarian Cancer Patients
Authors: de Caceres Inmaculada Ibanez , Battagli Cristina , Esteller Manel , Herman James G. , Dulaimi Essel , Edelson Mitchell I. , Bergman Cynthia , Ehya Hormoz , Eisenberg Burton L. , Cairns Paul
Published: September 2005

Because existing surgical and management methods can consistently cure only early-stage ovarian cancer, novel strategies for early detection are required. Silencing of tumor suppressor genes such as p16INK4a, VHL, and hMLH1 have established promoter hypermethylation as a common mechanism for tumor suppressor inactivation in human cancer and as a promising target for molecular detection in bodily fluids. Using sensitive methylation-specific PCR, we screened matched tumor, preoperative serum or plasma, and peritoneal fluid (washes or ascites) DNA obtained from 50 patients with ovarian or primary peritoneal tumors for hypermethylation status of the normally unmethylated BRCA1 and RAS association domain family protein 1A tumor suppressor genes. Hypermethylation of one or both genes was found in 34 tumor DNA (68%). Additional examination of one or more of the adenomatous polyposis coli, p14ARF, p16INK4a, or death associated protein-kinase tumor suppressor genes revealed hypermethylation in each of the remaining 16 tumor DNA, which extended diagnostic coverage to 100%. Hypermethylation was observed in all histologic cell types, grades, and stages of ovarian tumor examined. An identical pattern of gene hypermethylation was found in the matched serum DNA from 41 of 50 patients (82% sensitivity), including 13 of 17 cases of stage I disease. Hypermethylation was detected in 28 of 30 peritoneal fluid DNA from stage IC-IV patients, including 3 cases with negative or atypical cytology. In contrast, no hypermethylation was observed in nonneoplastic tissue, peritoneal fluid, or serum from 40 control women (100% specificity). We conclude that promoter hypermethylation is a common and relatively early event in ovarian tumorigenesis that can be detected in the serum DNA from patients with ovary-confined (stage IA or B) tumors and in cytologically negative peritoneal fluid. Analysis of tumor-specific hypermethylation in serum DNA may enhance early detection of ovarian cancer.
Lysyl Oxidase
Is a Tumor Suppressor Gene Inactivated by Methylation and Loss of Heterozygosity in Human Gastric Cancers
Authors: Kaneda Atsushi , Wakazono Kuniko , Tsukamoto Tetsuya , Watanabe Naoko , Yagi Yukiko , Tatematsu Masae , Kaminishi Michio , Sugimura Takashi , Ushijima Toshikazu
Published: September 2005

Lysyl oxidase (LOX) and HRAS-like suppressor (HRASLS) are silenced in human gastric cancers and are reported to have growth-suppressive activities in ras-transformed mouse/rat fibroblasts. Here, we analyzed whether or not LOX and HRASLS are tumor suppressor genes in human gastric cancers. Loss of heterozygosity and promoter methylation of LOX were detected in 33% (9 of 27) and 27% (26 of 96) of gastric cancers, respectively. Biallelic methylation and loss of heterozygosity with promoter methylation were also demonstrated in gastric cancers. Silencing of LOX was also observed in colon, lung, and ovarian cancer cell lines. As for mutations, only one possible somatic mutation was found by analysis of 96 gastric cancer samples and 58 gastric and other cancer cell lines. When LOX was introduced into a gastric cancer cell line, MKN28, in which LOX and HRASLS were silenced, it reduced the number of anchorage-dependent colonies to 57 to 61%, and the number of anchorage-independent colonies to 11 to 23%. Sizes of tumors formed in nude mice were reduced to 19 to 26%. Growth suppression in soft agar assay was also observed in another gastric cancer cell line, KATOIII. On the other hand, neither loss of heterozygosity nor a somatic mutation was detected in HRASLS, and its introduction into MKN28 did not suppress the growth in vitro or in vivo. These data showed that LOX is a tumor suppressor gene inactivated by methylation and loss of heterozygosity in gastric cancers, and possibly also in other cancers.
Antitumor Vaccination in Patients with Head and Neck Squamous Cell Carcinomas with Autologous Virus-Modified Tumor Cells
Authors: Karcher Jochen , Dyckhoff Gerhard , Beckhove Philipp , Reisser Christoph , Brysch Michael , Ziouta Yvonne , Helmke Burkhard H. , Weidauer Hagen , Schirrmacher Volker , Herold-Mende Christel
Published: September 2005

Prognosis of patients with advanced head and neck squamous cell carcinomas (HNSCC) is still poor. Therefore, we analyzed whether antitumor vaccination with a virus-modified autologous tumor cell vaccine is feasible and safe in HNSCC patients. Furthermore, we determined the influence on disease-free survival and overall survival and the vaccination-induced antitumor reactivity. In a nonrandomized pilot study, 20 patients were vaccinated postoperatively. Vaccine was prepared from the tumor cell cultures of patients by infection of the cells with Newcastle Disease Virus, followed by γ-irradiation, and vaccine was applied up to five times. Antitumor immune reactivity was determined in the skin by delayed type hypersensitivity skin reaction and in the blood by enzyme-linked immunospot assay. Establishment of tumor cell cultures was successful in about 80% of the cases. After vaccination, we observed no severe side effects. Percentages of survival of vaccinated patients with stage III and stage IV tumors (n = 18) were 61% at 5 years. Immune monitoring revealed significant increases of antitumor delayed type hypersensitivity reactivity especially in disease-free patients, and in a significant proportion of vaccinated patients the presence of tumor-reactive T-cells in the peripheral blood even 5 to 7 years after the last vaccination. Postoperative vaccination with virus-modified autologous tumor cells seems to be feasible and safe and may improve the prognosis of HNSCC patients with advanced tumors. This could be supported by antitumor immune responses that we observed especially in long-term surviving patients.
Modulation of Gene Expression by Tumor-Derived p53 Mutants
Authors: Scian Mariano J. , Stagliano Katherine E. R. , Ellis Michelle A. , Hassan Sajida , Bowman Melissa , Miles Michael F. , Deb Swati Palit , Deb Sumitra
Published: September 2005

p53 mutants with a single amino acid substitution are overexpressed in a majority of human cancers containing a p53 mutation. Overexpression of the mutant protein suggests that there is a selection pressure on the cell indicative of an active functional role for mutant p53. Indeed, H1299 cells expressing mutant p53-R175H, p53-R273H or p53-D281G grow at a faster rate compared with a control cell line. Using p53-specific small interfering RNA, we show that the growth rate of mutant p53-expressing cells decreases as mutant p53 level decreases, demonstrating that the increased cellular growth is dependent on p53 expression. Increased growth rate is not observed for H1299 cell clones expressing mutant p53-D281G (L22Q/W23S), which has been shown to be defective in transactivation in transient transcriptional assays. This shows that the increased growth rate imparted by mutant p53 in H1299 cells requires the transactivation function of mutant p53. By performing microarray hybridization analyses, we show that constitutive expression of three common p53 mutants (p53-R175H, p53-R273H, and p53-D281G) in H1299 human lung carcinoma cells evokes regulation of a common set of genes, a significant number of which are involved in cell growth regulation. Predictably, H1299 cells expressing p53-D281G (L22Q/W23S) are defective in up-regulating a number of these genes. The differences in expression profiles induced by individual p53 mutants in the cells may be representative of the p53 mutants and how they can affect gene expression resulting in the observed “gain of function” phenotypes (i.e., increased growth rate, decreased sensitivity to chemotherapeutic agents, and so forth).
The Palmitoylation of Metastasis Suppressor KAI1/CD82 Is Important for Its Motility- and Invasiveness-Inhibitory Activity
Authors: Zhou Bin , Liu Li , Reddivari Muralidhar , Zhang Xin A.
Published: September 2005

The cancer metastasis suppressor protein KAI1/CD82 is a member of the tetraspanin superfamily. Recent studies have demonstrated that tetraspanins are palmitoylated and that palmitoylation contributes to the organization of tetraspanin webs or tetraspanin-enriched microdomains. However, the effect of palmitoylation on tetraspanin-mediated cellular functions remains obscure. In this study, we found that tetraspanin KAI1/CD82 was palmitoylated when expressed in PC3 metastatic prostate cancer cells and that palmitoylation involved all of the cytoplasmic cysteine residues proximal to the plasma membrane. Notably, the palmitoylation-deficient KAI1/CD82 mutant largely reversed the wild-type KAI1/CD82’s inhibitory effects on migration and invasion of PC3 cells. Also, palmitoylation regulates the subcellular distribution of KAI1/CD82 and its association with other tetraspanins, suggesting that the localized interaction of KAI1/CD82 with tetraspanin webs or tetraspanin-enriched microdomains is important for KAI1/CD82’s motility-inhibitory activity. Moreover, we found that KAI1/CD82 palmitoylation affected motility-related subcellular events such as lamellipodia formation and actin cytoskeleton organization and that the alteration of these processes likely contributes to KAI1/CD82’s inhibition of motility. Finally, the reversal of cell motility seen in the palmitoylation-deficient KAI1/CD82 mutant correlates with regaining of p130CAS-CrkII coupling, a signaling step important for KAI1/CD82’s activity. Taken together, our results indicate that palmitoylation is crucial for the functional integrity of tetraspanin KAI1/CD82 during the suppression of cancer cell migration and invasion.
Caspase Inhibitors, but not c-Jun NH2-Terminal Kinase Inhibitor Treatment, Prevent Cisplatin-Induced Hearing Loss
Authors: Wang Jing , Ladrech Sabine , Pujol Remy , Brabet Philippe , Van De Water Thomas R. , Puel Jean-Luc
Published: September 2005

Cisplatin (CDDP) is a highly effective chemotherapeutic agent but with significant ototoxic side effects. Apoptosis is an important mechanism of cochlear hair cell loss following exposure to an ototoxic level of CDDP. This study examines intracellular pathways involved in hair cell death induced by CDDP exposure in vivo to develop effective therapeutic strategies to protect the auditory receptor from CDDP-initiated hearing loss. Guinea pigs were treated with systemic administration of CDDP. Cochlear hair cells from CDDP-treated animals exhibited classic apoptotic alterations in their morphology. Several important signaling events that regulate the death of CDDP-injured cochlear hair cells were identified. CDDP treatment induced the activation and redistribution of cytosolic Bax and the release of cytochrome c from injured mitochondria. Activation of caspase-9 and caspase-3, but not caspase-8, was detected after treatment with CDDP, and the cleavage of fodrin by activated caspase-3 was observed within damaged hair cells. Intracochlear perfusions with caspase-3 inhibitor (z-DEVD-fmk) and caspase-9 inhibitor (z-LEHD-fmk) prevent hearing loss and loss of sensory cells, but caspase-8 inhibitor (z-IETD-fmk) and cathepsin B inhibitor (z-FA-fmk) do not. Although the stress-activated protein kinase/c-Jun NH2-terminal kinase (JNK) signaling pathway is activated in response to CDDP toxicity, intracochlear perfusion of d-JNKI-1, a JNK inhibitor, did not protect against CDDP ototoxicity but instead potentiated the ototoxic effects of CDDP. The results of the present study show that blocking a critical step in apoptosis may be a useful strategy to prevent harmful side effects of CDDP ototoxicity in patients having to undergo chemotherapy.
Uric Acid Promotes Tumor Immune Rejection
Authors: Hu De-En , Moore Alistair M. , Thomsen Lindy L. , Brindle Kevin M.
Published: September 2005

Uric acid released from dying cells has been shown recently to act as a danger signal for the immune system, stimulating dendritic cell maturation and enhancing T-cell responses to foreign antigens. Stimulation of dendritic cell maturation by uric acid has been proposed as a mechanism by which the immune system could generate responses against tumors. We show here that uric acid levels are elevated in tumors undergoing immune rejection and that the inhibition of uric acid production, by systemic administration of allopurinol, or the removal of uric acid, by administration of uricase, delayed tumor immune rejection, whereas subcutaneous administration of crystalline uric acid enhanced the rejection process.
Ontogeny and Oncogenesis Balance the Transcriptional Profile of Renal Cell Cancer
Authors: Wang Ena , Lichtenfels Rudolf , Bükur Jurgen , Ngalame Yvonne , Panelli Monica C. , Seliger Barbara , Marincola Francesco M.
Published: September 2005

Global transcript analysis is increasingly used to describe cancer taxonomies beyond the microscopic reach of the eye. Diagnostic and prognostic portraits are formulated by ranking cancers according to transcriptional proximity. However, the role that distinct biological factors play in defining these portraits remains undefined. It is likely that the transcriptional repertoire of cancers depends, on one hand, on the anamnestic retention of their ontogenesis and, on the other, on the emergence of novel expression patterns related to oncogenesis. We compared the transcriptional profile of primary renal cell cancers (RCCs) with that of normal kidney tissue and several epithelial cancers of nonrenal origin to weigh the contribution that ontogeny and oncogenesis make in molding their genetic profile. Unsupervised global transcript analysis demonstrated that RCCs retain transcriptional signatures related to their ontogeny and cluster close to normal renal epithelium. When renal lineage-associated genes are removed from the analysis and cancer-specific genes are analyzed, RCCs segregate with other cancers with limited lineage specificity underlying a predominance of the oncogenic process over lineage specificity. However, a RCC-specific set of oncogenesis-related genes was identified and surprisingly shared by sarcomas. In summary, the transcriptional portrait of primary RCCs is largely dominated by ontogeny. Genes responsible for lineage specificity may represent poor molecular targets for immune or drug therapy. Most genes associated with oncogenesis are shared with other cancers and may represent better therapeutic targets. Finally, a small subset of genes is associated with lineage-specific oncogenesis, and these may provide information regarding the biological behavior of RCCs and facilitate diagnostic classification of RCCs.