bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from A Recurrent Chromosome Breakpoint in Breast Cancer at the NRG1/Neuregulin 1/Heregulin Gene
Authors: Huang Huai-En , Chin Suet-Feung , Ginestier Christophe , Bardou Valérie-Jeanne , Adélaïde José , Iyer N. Gopalakrishna , Garcia Maria J. , Pole Jessica C. , Callagy Grace M. , Hewitt Stephen M. , Gullick William J. , Jacquemier Jocelyne , Caldas Carlos , Chaffanet Max , Birnbaum Daniel , Edwards Paul A. W.
Published: March 2023

Abstract
Most studies of genomic rearrangements in common cancers have focused on regional gains and losses, but some rearrangements may break within specific genes. We previously reported that five breast cancer cell lines have chromosome translocations that break in the NRG1 gene and that could cause abnormal NRG1 expression. NRG1 encodes the Neuregulins 1 (formerly the Heregulins), ligands for members of the ErbB/epidermal growth factor-receptor family, which includes ErbB2/HER2. We have now screened for breaks at NRG1 in paraffin sections of breast tumors. Tissue microarrays were screened by fluorescence in situ hybridization, with hybridization probes proximal and distal to the expected breakpoints. This screen detects breaks but does not distinguish between translocation or deletion breakpoints. The screen was validated with array-comparative genomic hybridization on a custom 8p12 high-density genomic array to detect a lower copy number of the sequences that were lost distal to the breaks. We also precisely mapped the breaks in five tumors with different hybridization probes. Breaks in NRG1 were detected in 6% (19 of 323) of breast cancers and in some lung and ovarian cancers. In an unselected series of 213 cases with follow-up, breast cancers where the break was detected tended to be high-grade (65% grade III compared with 28% of negative cases). They were, like breast tumors in general, mainly ErbB2 low (11 of 13 were low) and estrogen receptor positive (11 of 13 positive).
Data from The Farnesoid X Receptor Is Expressed in Breast Cancer and Regulates Apoptosis and Aromatase Expression
Authors: Swales Karen E. , Korbonits Márta , Carpenter Robert , Walsh Desmond T. , Warner Timothy D. , Bishop-Bailey David
Published: March 2023

Abstract
Bile acids are present at high concentrations in breast cysts and in the plasma of postmenopausal women with breast cancer. The farnesoid X receptor (FXR) is a member of the nuclear receptor superfamily that regulates bile acid homeostasis. FXR was detected in normal and tumor breast tissue, with a high level of expression in ductal epithelial cells of normal breast and infiltrating ductal carcinoma cells. FXR was also present in the human breast carcinoma cells, MCF-7 and MDA-MB-468. Activation of FXR by high concentrations of ligands induced MCF-7 and MDA-MB-468 apoptosis. At lower concentrations that had no direct effect on viability, the FXR agonist GW4064 induced expression of mRNA for the FXR target genes, small heterodimer partner (SHP), intestinal bile acid binding protein, and multidrug resistance–associated protein 2 (MRP-2), and repressed the expression of the SHP target gene aromatase. In contrast to MRP-2, mRNA for the breast cancer target genes MDR-3, MRP-1, and solute carrier transporter 7A5 were decreased. Although multidrug resistance transporters were regulated and are known FXR target genes, GW4064 had no effect on the cell death induced by the anticancer drug paclitaxel. Our findings show for the first time that FXR is expressed in breast cancer tissue and has multiple properties that could be used for the treatment of breast cancer. (Cancer Res 2006; 66(20): 10120-6)
Data from Atm Heterozygosity Cooperates with Loss of Brca1 to Increase the Severity of Mammary Gland Cancer and Reduce Ductal Branching
Authors: Bowen T.J. , Yakushiji Hiroyuki , Montagna Cristina , Jain Sonia , Ried Thomas , Wynshaw-Boris Anthony
Published: March 2023

Abstract
The role of homozygous ataxia telangiectasia mutated (ATM) mutations in familial and sporadic forms of cancer is well established, but the contribution of ATM heterozygosity to mammary gland and other cancers has been controversial. To test the effect of Atm heterozygosity on mammary gland cancer, mice with complete loss of exon 11 of Brca1 specifically in mammary epithelium (Brca1-MG-Δex11) were studied in either Atm heterozygous or Atm wild-type backgrounds. Targeted deletion of Brca1 in mammary epithelium resulted in carcinomas and adenocarcinomas of varying histology with long (>9 months) latency. Latency to tumorigenesis was found to be unchanged in the Brca1-MG-Δex11;Atm heterozygous mice compared with Brca1-MG-Δex11;Atm wild-type mice. However, the mice displayed variable tumor severity and differences in mammary tissue development. Mammary tumors from Brca1-MG-Δex11;Atm heterozygous mice were anaplastic and undifferentiated in all 20 tumors tested, whereas tumors from mice that were Brca1-MG-Δex11 but wild-type for Atm displayed variable histologic profiles, with some anaplastic tumors and other differentiated and less invasive tumor types. Previously reported developmental defects for Brca1-deficient mice were also observed in our model with and without Atm heterozygosity, but Brca1-MG-Δex11;Atm heterozygous mice displayed decreased ductal branching during puberty, a phenotype that was not observed in Brca1-MG-Δex11;Atm wild-type mice. Our results provide evidence that Atm heterozygosity influences severity of mammary gland tumors in the Brca1-MG-Δex11 tumor-prone mouse and suggest that this mutation leads to a newly characterized developmental defect during glandular maturation.
Data from Malignant Transformation of Immortalized HaCaT Keratinocytes through Deregulated Nuclear Factor κB Signaling
Authors: Ren Qing , Kari Csaba , Quadros Marlene R.D. , Burd Randy , McCue Peter , Dicker Adam P. , Rodeck Ulrich
Published: March 2023

Abstract
Previous studies addressing functional aspects of nuclear factor κB (NF-κB) activation in normal and transformed keratinocytes revealed complex and seemingly contradictory roles of this transcription factor in this cell type. In normal skin, NF-κB signaling seems to inhibit squamous cell carcinoma development whereas, in squamous cell carcinoma themselves, deregulated NF-κB expression and/or signaling is frequently observed. To further investigate this paradox, we focused on NF-κB activation as it relates to the transformed phenotype of immortalized but nontumorigenic human keratinocytes (HaCaT cells). We observed that NF-κB activity contributed to survival and growth of cultured HaCaT keratinocytes as shown by use of pharmacologic NF-κB inhibitors, RNA interference, and inducible overexpression of a dominant interfering IκB construct. NF-κB activation was largely provided through interaction with extracellular matrix components because preventing cell attachment by forced suspension culture markedly reduced NFκB signaling associated with cell death (anoikis); conversely, anoikis was partially reversed by NF-κB activation induced either by tumor necrosis factor-α treatment or by overexpressing the NF-κB p65 subunit in HaCaT cells. Furthermore, overexpression of NF-κBp65 in HaCaT cells induced colony formation in soft agar and tumorigenicity in nude mice. In summary, as opposed to normal keratinocytes, immortalized HaCaT keratinocytes provide a cellular context in which deregulated NF-κB signaling supports multiple malignant traits in vitro and in vivo. (Cancer Res 2006; 66(10): 5209-15)
Data from Genome-Wide Expression Profiling Reveals EBV-Associated Inhibition of MHC Class I Expression in Nasopharyngeal Carcinoma
Authors: Sengupta Srikumar , den Boon Johan A. , Chen I-How , Newton Michael A. , Dahl David B. , Chen Meng , Cheng Yu-Juen , Westra William H. , Chen Chien-Jen , Hildesheim Allan , Sugden Bill , Ahlquist Paul
Published: March 2023

Abstract
To identify the molecular mechanisms by which EBV-associated epithelial cancers are maintained, we measured the expression of essentially all human genes and all latent EBV genes in a collection of 31 laser-captured, microdissected nasopharyngeal carcinoma (NPC) tissue samples and 10 normal nasopharyngeal tissues. Global gene expression profiles clearly distinguished tumors from normal healthy epithelium. Expression levels of six viral genes (EBNA1, EBNA2, EBNA3A, EBNA3B, LMP1, and LMP2A) were correlated among themselves and strongly inversely correlated with the expression of a large subset of host genes. Among the human genes whose inhibition was most strongly correlated with increased EBV gene expression were multiple MHC class I HLA genes involved in regulating immune response via antigen presentation. The association between EBV gene expression and inhibition of MHC class I HLA expression implies that antigen display is either directly inhibited by EBV, facilitating immune evasion by tumor cells, and/or that tumor cells with inhibited presentation are selected for their ability to sustain higher levels of EBV to take maximum advantage of EBV oncogene-mediated tumor-promoting actions. Our data clearly reflect such tumor promotion, showing that deregulation of key proteins involved in apoptosis (BCL2-related protein A1 and Fas apoptotic inhibitory molecule), cell cycle checkpoints (AKIP, SCYL1, and NIN), and metastasis (matrix metalloproteinase 1) is closely correlated with the levels of EBV gene expression in NPC. (Cancer Res 2006; 66(16): 7999-8006)
Data from Antiangiogenic Antithrombin Induces Global Changes in the Gene Expression Profile of Endothelial Cells
Authors: Zhang Weiqing , Chuang Yung-Jen , Jin Tianquan , Swanson Richard , Xiong Yan , Leung Lawrence , Olson Steven T.
Published: March 2023

Abstract
Antithrombin, a serpin family protease inhibitor crucial to hemostasis, acquires antiangiogenic properties on undergoing conformational alterations induced by limited proteolysis or elevated temperature. To better understand the biochemical mechanisms underlying antithrombin antiangiogenic activity, we did genome-wide expression profiling, coupled with quantitative reverse transcription-PCR, Northern blot, and Western blot analyses, to characterize the gene expression patterns that are induced by antiangiogenic antithrombin in cultured primary human umbilical vein endothelial cells. Overall, 35 genes with significantly increased expression and 93 genes with significantly reduced expression (≥2-fold changes) due to antiangiogenic antithrombin treatment were identified. More than half of the down-regulated genes have well-established proangiogenic functions in endothelial cells, including cell-surface and matrix proteoglycans (e.g., perlecan, biglycan, and syndecans 1 and 3) and mitogenesis-related signaling proteins (e.g., mitogen-activated protein kinase 3, signal transducers and activators of transcription 2, 3, and 6, and early growth response factor 1). In contrast, most up-regulated genes (e.g., caspase-3, p21, tissue inhibitor of metalloproteinases 1, 2, and 3, and adenomatosis polyposis coli) are known for their antiangiogenic functions which include the promotion of cell apoptosis and cell cycle arrest and the inhibition of tumor growth and metastasis. These results show that the antiangiogenic activity of antithrombin is mediated at least in part by a global genetic reprogramming of endothelial cells and strongly implicate an endothelial cell ligand-receptor signaling mechanism in this reprogramming. (Cancer Res 2006; 66(10): 5047-55)
Data from Conditional Expression of the CTCF-Paralogous Transcriptional Factor BORIS in Normal Cells Results in Demethylation and Derepression of MAGE-A1 and Reactivation of Other Cancer-Testis Genes
Authors: Vatolin Sergei , Abdullaev Ziedulla , Pack Svetlana D. , Flanagan Patrick T. , Custer Mary , Loukinov Dmitri I. , Pugacheva Elena , Hong Julie A. , Morse Herbert , Schrump David S. , Risinger John I. , Barrett J. Carl , Lobanenkov Victor V.
Published: March 2023

Abstract
Brother of the Regulator of Imprinted Sites (BORIS) is a mammalian CTCF paralog with the same central 11Zn fingers (11ZF) that mediate specific interactions with varying ∼50-bp target sites. Regulated in vivo occupancy of such sites may yield structurally and functionally distinct CTCF/DNA complexes involved in various aspects of gene regulation, including epigenetic control of gene imprinting and X chromosome inactivation. The latter functions are mediated by meCpG-sensitive 11ZF binding. Because CTCF is normally present in all somatic cells, whereas BORIS is active only in CTCF- and 5-methylcytosine–deficient adult male germ cells, switching DNA occupancy from CTCF to BORIS was suggested to regulate site specificity and timing of epigenetic reprogramming. In addition to 11ZF-binding paternal imprinting control regions, cancer-testis gene promoters also undergo remethylation during CTCF/BORIS switching in germ cells. Only promoters of cancer testis genes are normally silenced in all somatic cells but activated during spermatogenesis when demethylated in BORIS-positive germ cells and are found aberrantly derepressed in various tumors. We show here that BORIS is also expressed in multiple cancers and is thus itself a cancer-testis gene and that conditional expression of BORIS in normal fibroblasts activates cancer-testis genes selectively. We tested if replacement of CTCF by BORIS on regulatory DNA occurs in vivo on activation of a prototype cancer-testis gene, MAGE-A1. Transition from a hypermethylated/silenced to a hypomethylated/activated status induced in normal cells by 5-aza-2′-deoxycytidine (5-azadC) was mimicked by conditional input of BORIS and is associated with complete switching from CTCF to BORIS occupancy at a single 11ZF target. This site manifested a novel type of CTCF/BORIS 11ZF binding insensitive to CpG methylation. Whereas 5-azadC induction of BORIS takes only few hours, derepression of MAGE-A1 occurred 1 to 2 days later, suggesting that BORIS mediates cancer-testis gene activation by 5-azadC. Indeed, infection of normal fibroblasts with anti-BORIS short hairpin RNA retroviruses before treatment with 5-azadC blocked reactivation of MAGE-A1. We suggest that BORIS is likely tethering epigenetic machinery to a novel class of CTCF/BORIS 11ZF target sequences that mediate induction of cancer-testis genes.
Data from Identification of SFRP1 as a Candidate Mediator of Stromal-to-Epithelial Signaling in Prostate Cancer
Authors: Joesting Margaret S. , Perrin Steve , Elenbaas Brian , Fawell Stephen E. , Rubin Jeffrey S. , Franco Omar E. , Hayward Simon W. , Cunha Gerald R. , Marker Paul C.
Published: March 2023

Abstract
Genetic changes in epithelial cells initiate the development of prostatic adenocarcinomas. As nascent tumors grow and undergo progression, epithelial tumor cells are intimately associated with stromal cells. Stromal cells within the tumor microenvironment acquire new properties, including the capacity to promote phenotypic and genetic progression in adjacent epithelial cells. Affymetrix microarrays were used to identify 119 genes differentially expressed between normal-derived and carcinoma-derived prostatic stromal cells. These included 31 genes encoding extracellular proteins that may act as stromal-to-epithelial paracrine signals. Further investigation of one of these genes, secreted frizzled related protein 1 (SFRP1), revealed that its expression parallels prostatic growth with high expression during prostatic development, low expression in the adult prostate, and elevated expression in prostatic tumor stroma. In addition, as prostatic epithelial cells progressed to a tumorigenic state under the influence of tumor stroma, SFRP1 became overexpressed in the progressed epithelial cells. To further understand the roles of SFRP1 in the prostate, we tested the affects of increased SFRP1 levels on prostatic tissues and cells. Treatment of developing prostates with SFRP1 in culture led to increased organ growth. Treatment of a human prostatic epithelial cell line with SFRP1 led to increased proliferation, decreased apoptosis, and decreased signaling through the Wnt/β-catenin pathway in vitro and increased proliferation in vivo. These data suggest that overexpression of SFRP1 by prostatic tumor stroma may account for the previously reported capacity of prostatic tumor stroma to provide a pro-proliferative paracrine signal to adjacent epithelial cells.
Data from The Neuromedin U-Growth Hormone Secretagogue Receptor 1b/Neurotensin Receptor 1 Oncogenic Signaling Pathway as a Therapeutic Target for Lung Cancer
Authors: Takahashi Koji , Furukawa Chiyuki , Takano Atsushi , Ishikawa Nobuhisa , Kato Tatsuya , Hayama Satoshi , Suzuki Chie , Yasui Wataru , Inai Kouki , Sone Saburo , Ito Tomoo , Nishimura Hitoshi , Tsuchiya Eiju , Nakamura Yusuke , Daigo Yataro
Published: March 2023

Abstract
Using a genome-wide cDNA microarray to search for genes that were specifically up-regulated in non–small cell lung cancers (NSCLC), we identified an abundant expression of neuromedin U (NMU) in the great majority of lung cancers. Immunohistochemical analysis showed a significant association of NMU expression with poorer prognosis of patients with NSCLC. Treatment of NSCLC cells with short interfering RNA against NMU suppressed its expression and inhibited the growth of the cells; on the other hand, the induction of exogenous expression of NMU conferred growth-promoting activity and enhanced cell mobility in vitro. We found that two G protein–coupled receptors, growth hormone secretagogue receptor 1b and neurotensin receptor 1, were also overexpressed in NSCLC cells, and that a heterodimer complex of these receptors functioned as an NMU receptor. The NMU-receptor interaction subsequently induced the generation of a second messenger, cyclic AMP, to activate its downstream genes including transcription factors and cell cycle regulators. Treatment of NSCLC cells with short interfering RNAs for growth hormone secretagogue receptor or neurotensin receptor 1 suppressed the expression of those genes and the growth of NSCLC cells. These data strongly implied that targeting the NMU signaling pathway would be a promising therapeutic strategy for the treatment of lung cancers. (Cancer Res 2006; 66(19): 9408-19)
Data from Distinct Transcription Profiles of Primary and Secondary Glioblastoma Subgroups
Authors: Tso Cho-Lea , Freije William A. , Day Allen , Chen Zugen , Merriman Barry , Perlina Ally , Lee Yohan , Dia Ederlyn Q. , Yoshimoto Koji , Mischel Paul S. , Liau Linda M. , Cloughesy Timothy F. , Nelson Stanley F.
Published: March 2023

Abstract
Glioblastomas are invasive and aggressive tumors of the brain, generally considered to arise from glial cells. A subset of these cancers develops from lower-grade gliomas and can thus be clinically classified as “secondary,” whereas some glioblastomas occur with no prior evidence of a lower-grade tumor and can be clinically classified as “primary.” Substantial genetic differences between these groups of glioblastomas have been identified previously. We used large-scale expression analyses to identify glioblastoma-associated genes (GAG) that are associated with a more malignant phenotype via comparison with lower-grade astrocytomas. We have further defined gene expression differences that distinguish primary and secondary glioblastomas. GAGs distinct to primary or secondary tumors provided information on the heterogeneous properties and apparently distinct oncogenic mechanisms of these tumors. Secondary GAGs primarily include mitotic cell cycle components, suggesting the loss of function in prominent cell cycle regulators, whereas primary GAGs highlight genes typical of a stromal response, suggesting the importance of extracellular signaling. Immunohistochemical staining of glioblastoma tissue arrays confirmed expression differences. These data highlight that the development of gene pathway-targeted therapies may need to be specifically tailored to each subtype of glioblastoma. (Cancer Res 2006; 66(1): 159-67)
Data from Genetic Reclassification of Histologic Grade Delineates New Clinical Subtypes of Breast Cancer
Authors: Ivshina Anna V. , George Joshy , Senko Oleg , Mow Benjamin , Putti Thomas C. , Smeds Johanna , Lindahl Thomas , Pawitan Yudi , Hall Per , Nordgren Hans , Wong John E.L. , Liu Edison T. , Bergh Jonas , Kuznetsov Vladimir A. , Miller Lance D.
Published: March 2023

Abstract
Histologic grading of breast cancer defines morphologic subtypes informative of metastatic potential, although not without considerable interobserver disagreement and clinical heterogeneity particularly among the moderately differentiated grade 2 (G2) tumors. We posited that a gene expression signature capable of discerning tumors of grade 1 (G1) and grade 3 (G3) histology might provide a more objective measure of grade with prognostic benefit for patients with G2 disease. To this end, we studied the expression profiles of 347 primary invasive breast tumors analyzed on Affymetrix microarrays. Using class prediction algorithms, we identified 264 robust grade-associated markers, six of which could accurately classify G1 and G3 tumors, and separate G2 tumors into two highly discriminant classes (termed G2a and G2b genetic grades) with patient survival outcomes highly similar to those with G1 and G3 histology, respectively. Statistical analysis of conventional clinical variables further distinguished G2a and G2b subtypes from each other, but also from histologic G1 and G3 tumors. In multivariate analyses, genetic grade was consistently found to be an independent prognostic indicator of disease recurrence comparable with that of lymph node status and tumor size. When incorporated into the Nottingham prognostic index, genetic grade enhanced detection of patients with less harmful tumors, likely to benefit little from adjuvant therapy. Our findings show that a genetic grade signature can improve prognosis and therapeutic planning for breast cancer patients, and support the view that low- and high-grade disease, as defined genetically, reflect independent pathobiological entities rather than a continuum of cancer progression. (Cancer Res 2006; 66(21): 10292-301)
Data from CXC Chemokine Receptor-4 Antagonist Blocks Both Growth of Primary Tumor and Metastasis of Head and Neck Cancer in Xenograft Mouse Models
Authors: Yoon Younghyoun , Liang Zhongxing , Zhang Xin , Choe Mison , Zhu Aizhi , Cho Heidi T. , Shin Dong M. , Goodman Mark M. , Chen Zhuo (Georgia) , Shim Hyunsuk
Published: March 2023

Abstract
Squamous cell carcinoma of the head and neck (SCCHN) metastasizes to the lymph nodes and lungs. We have generated previously an orthotopic mouse model for head and neck metastasis and did in vivo selection of SCCHN cells through four rounds of serial metastases. A subpopulation of 686LN cells with high metastatic potential (686LN-Ms) was isolated. When the highly metastatic cells were compared with their low metastatic parental cells (686LN-Ps), we found that CXC chemokine receptor-4 (CXCR4) mRNA levels were significantly higher in the 686LN-Ms cells than the 686LN-Ps cells. Interestingly, the metastatic subclones had lost epithelial morphology and acquired mesenchymal features, which were maintained during cell expansion in vitro. This was featured by decreased E-cadherin and involucrin and increased vimentin and integrin β1. These results imply that CXCR4 and epithelial-mesenchymal transition markers can be potential biomarkers to identify the subpopulation of cells with high metastatic potential. Using the orthotopic SCCHN animal model, we showed that anti-CXCR4 treatment suppressed primary tumor growth by inhibiting tumor angiogenesis and prevented lung metastasis. Because the reduction of metastasis seen in the treated group could have resulted from 2-fold reduction in primary tumor size compared with that in the control group, we examined the effects of the CXCR4 antagonist in an experimental metastatic animal model in which 686LN-Ms cells were i.v. injected. 686LN-Ms cells failed to metastasize in the CXCR4 antagonist-treated group, whereas they metastasized to the lungs in the control group. Our data indicate that CXCR4 is an important target to inhibit tumor progression in SCCHN. [Cancer Res 2007;67(15):7518–23]
Data from SLC5A8 Triggers Tumor Cell Apoptosis through Pyruvate-Dependent Inhibition of Histone Deacetylases
Authors: Thangaraju Muthusamy , Gopal Elangovan , Martin Pamela M. , Ananth Sudha , Smith Sylvia B. , Prasad Puttur D. , Sterneck Esta , Ganapathy Vadivel
Published: March 2023

Abstract
Tumor cells up-regulate glycolysis but convert pyruvate into lactate instead of oxidizing it. Here, we show that pyruvate, but not lactate, is an inhibitor of histone deacetylases (HDAC) and an inducer of apoptosis in tumor cells and that SLC5A8, a Na+/monocarboxylate cotransporter, is obligatory for this process. We found that SLC5A8 is expressed in nontransformed breast epithelial cell lines but silenced by DNA methylation in tumor cell lines. The down-regulation of the gene is also evident in primary breast tumors. When MCF7 breast tumor cells are transfected with SLC5A8 cDNA, the cells undergo pyruvate-dependent apoptosis. Butyrate and propionate also induce apoptosis in SLC5A8-expressing cells, whereas lactate does not. The differential ability of these monocarboxylates to cause apoptosis in SLC5A8-expressing MCF7 cells correlates with their ability to inhibit HDACs. Apoptosis induced by SLC5A8/pyruvate in MCF7 cells is associated with up-regulation of p53, Bax, tumor necrosis factor–related apoptosis-inducing ligand (TRAIL), TRAIL receptor (TRAILR) 1, and TRAILR2 and down-regulation of Bcl2 and survivin. Lactate dehydrogenase isozymes are differentially expressed in nontransformed cells and tumor cells such that the latter convert pyruvate into lactate. Silencing of SLC5A8 coupled with conversion of pyruvate into lactate in tumor cells correlates with increased HDAC activity in these cells compared with nontransformed cells. Our studies thus identify pyruvate as a HDAC inhibitor and indicate that the Na+-coupled pyruvate transport underlies the tumor-suppressive role of SLC5A8. We propose that tumor cells silence SLC5A8 and convert pyruvate into lactate as complementary mechanisms to avoid pyruvate-induced cell death. (Cancer Res 2006; 66(24): 11560-4)
Data from Distinct Roles for IFN Regulatory Factor (IRF)-3 and IRF-7 in the Activation of Antitumor Properties of Human Macrophages
Authors: Romieu-Mourez Raphaëlle , Solis Mayra , Nardin Alessandra , Goubau Delphine , Baron-Bodo Véronique , Lin Rongtuan , Massie Bernard , Salcedo Margarita , Hiscott John
Published: March 2023

Abstract
When properly activated, macrophages can be tumoricidal, thus making them attractive additions to standard cancer therapies. To this end, tolerance and activity of human autologous IFN-γ-activated macrophages, produced in large scale for clinical use (MAK cells), have been assessed in pilot trials in cancer patients. In the present study, we tested the hypothesis that activation of IFN regulatory factor (IRF)-3 and IRF-7, with subsequent type I IFN production, may be involved in the acquisition of new antitumor functions by macrophages. Adenoviral vectors were generated for the delivery of constitutively active forms of IRF-3 (Ad-IRF-3) or IRF-7 (Ad-IRF-7) into primary human macrophages. Cell death was observed in Ad-IRF-3-transduced macrophages, whereas Ad-IRF-7-transduced macrophages produced type I IFNs and displayed increased expression of genes encoding tumor necrosis factor (TNF)–related apoptosis-inducing ligand, interleukin (IL)-12, IL-15, and CD80, persisting for at least 96 hours. Expression of iNOS, TNF-α, FasL, IL-1, and IL-6 genes was unaltered by Ad-IRF-7 transduction. Interestingly, Ad-IRF-3 or Ad-IRF-7 transduction negatively regulated the transcription of protumorigenic genes encoding vascular endothelial growth factor and matrix metalloproteinase-2. Furthermore, Ad-IRF-7-transduced macrophages exerted a cytostatic activity on different cancer cell lines, including SK-BR-3, MCF-7, and COLO-205; the latter cells were shown previously to be insensitive to MAK cells. In conclusion, transduction of active forms of IRF-3 or IRF-7 differentially modulate the apoptotic and antitumor properties of primary macrophages, with active IRF-7 leading to the acquisition of novel antitumor effector functions. (Cancer Res 2006; 66(21): 10576-85)
Data from Colon Cancer Cell–Derived Tumor Necrosis Factor-α Mediates the Tumor Growth–Promoting Response in Macrophages by Up-regulating the Colony-Stimulating Factor-1 Pathway
Authors: Zins Karin , Abraham Dietmar , Sioud Mouldy , Aharinejad Seyedhossein
Published: March 2023

Abstract
The interplay between malignant and stromal cells is essential in tumorigenesis. We have previously shown that colony-stimulating factor (CSF)-1, matrix metalloprotease (MMP)-2, and vascular endothelial growth factor (VEGF)-A production by stromal cells is enhanced by CSF-1–negative SW620 colon cancer cells. In the present study, the mechanisms by which colon cancer cells up-regulate host factors to promote tumorigenesis were investigated. Profiling of tumor cell cytokine expression in SW620 tumor xenografts in nude mice showed increased human tumor necrosis factor (TNF)-α mRNA expression with tumor growth. Incubation of macrophages with small interfering (si) RNAs directed against TNF-α or TNF-α–depleted SW620 cell conditioned medium versus SW620 cell conditioned medium failed to support mouse macrophage proliferation, migration, and expression of CSF-1, VEGF-A, and MMP-2 mRNAs. Consistent with these results, human TNF-α gene silencing decreased mouse macrophage TNF-α, CSF-1, MMP-2, and VEGF-A mRNA expression in macrophages cocultured with human cancer cells. In addition, inhibition of human TNF-α or mouse CSF-1 expression by siRNA reduced tumor growth in SW620 tumor xenografts in mice. These results suggest that colon cancer cell–derived TNF-α stimulates TNF-α and CSF-1 production by macrophages, and that CSF-1, in turn, induces macrophage VEGF-A and MMP-2 in an autocrine manner. Thus, interrupting tumor cell–macrophage communication by targeting TNF-α may provide an alternative therapeutic approach for the treatment of colon cancer. [Cancer Res 2007;67(3):1038–45]
Data from In vivo Real-time Tracking of Single Quantum Dots Conjugated with Monoclonal Anti-HER2 Antibody in Tumors of Mice
Authors: Tada Hiroshi , Higuchi Hideo , Wanatabe Tomonobu M. , Ohuchi Noriaki
Published: March 2023

Abstract
Studies with tracking of single nanoparticles are providing new insights into the interactions and processes involved in the transport of drug carriers in living mice. Here, we report the tracking of a single particle quantum dot (Qdot) conjugated with tumor-targeting antibody in tumors of living mice using a dorsal skinfold chamber and a high-speed confocal microscope with a high-sensitivity camera. Qdot labeled with the monoclonal anti-HER2 antibody was injected into mice with HER2-overexpressing breast cancer to analyze the molecular processes of its mechanistic delivery to the tumor. Movement of single complexes of the Qdot-antibody could be clearly observed at 30 frames/s inside the tumor through a dorsal skinfold chamber. We successfully identified six processes of delivery: initially in the circulation within a blood vessel, during extravasation, in the extracelullar region, binding to HER2 on the cell membrane, moving from the cell membrane to the perinuclear region, and in the perinuclear region. The six processes were quantitatively analyzed to understand the rate-limiting constraints on Qdot-antibody delivery. The movement of the complexes at each stage was “stop-and-go.” The image analysis of the delivery processes of single particles in vivo provides valuable information on antibody-conjugated therapeutic nanoparticles, which will be useful in increasing therapeutic efficacy. [Cancer Res 2007;67(3):1138–44]
Data from 2-Methoxyestradiol Inhibits the Anaphase-Promoting Complex and Protein Translation in Human Breast Cancer Cells
Authors: Bhati Rajendra , Gökmen-Polar Yesim , Sledge George W. , Fan Cheng , Nakshatri Harikrishna , Ketelsen David , Borchers Christoph H. , Dial Michael J. , Patterson Cam , Klauber-DeMore Nancy
Published: March 2023

Abstract
2-Methoxyestradiol (2ME2), an estradiol metabolite with antiproliferative and antiangiogenic activities, is in phase I/II clinical trials for breast cancer. 2ME2 inhibits microtubule polymerization and causes cells to arrest in G2-M. The purpose of this study was to further elucidate the molecular mechanism of 2ME2. MDA-MB-435 breast cancer cells were treated with 2ME2 (2 μmol/L) or vehicle alone. RNA was extracted and genomic profiling was done using 22k Agilent microarrays. Expression Analysis Systematic Explorer was used to determine enrichment of Gene Ontology categories. Protein isolates were subjected to Western blot analysis. Protein synthesis was measured with a [35S]methionine pulse assay. An MDA-MB-435 cell line with two β-tubulin mutations (2ME2R) was used to determine whether novel mechanisms were tubulin-dependent. Gene Ontology categories enriched include genes that regulate the mitotic spindle assembly checkpoint, apoptosis, and the cytosolic ribosome. The target of the mitotic spindle assembly checkpoint is the anaphase-promoting complex (APC). APC inhibition was confirmed by measuring protein levels of its targets securin and cyclin B1, which were increased in 2ME2-treated cells. Because gene expression in the cytosolic ribosome category was decreased, we evaluated whether 2ME2 decreases protein translation. This was confirmed with a pulse assay, which showed decreased isotope incorporation in 2ME2-treated cells, which was maintained in the tubulin-resistant 2ME2R cells. APC inhibition was not maintained in 2ME2R cells. 2ME2 induces tubulin-dependent cell cycle arrest through regulation of genes involved in the mitotic spindle assembly checkpoint, which results in inhibition of the APC and tubulin-independent inhibition of protein translation. [Cancer Res 2007;67(2):702–8]
Data from A Hypermutation Phenotype and Somatic MSH6 Mutations in Recurrent Human Malignant Gliomas after Alkylator Chemotherapy
Authors: Hunter Chris , Smith Raffaella , Cahill Daniel P. , Stephens Philip , Stevens Claire , Teague Jon , Greenman Chris , Edkins Sarah , Bignell Graham , Davies Helen , O'Meara Sarah , Parker Adrian , Avis Tim , Barthorpe Syd , Brackenbury Lisa , Buck Gemma , Butler Adam , Clements Jody , Cole Jennifer , Dicks Ed , Forbes Simon , Gorton Matthew , Gray Kristian , Halliday Kelly , Harrison Rachel , Hills Katy , Hinton Jonathon , Jenkinson Andy , Jones David , Kosmidou Vivienne , Laman Ross , Lugg Richard , Menzies Andrew , Perry Janet , Petty Robert , Raine Keiran , Richardson David , Shepherd Rebecca , Small Alexandra , Solomon Helen , Tofts Calli , Varian Jennifer , West Sofie , Widaa Sara , Yates Andy , Easton Douglas F. , Riggins Gregory , Roy Jennifer E. , Levine Kymberly K. , Mueller Wolf , Batchelor Tracy T. , Louis David N. , Stratton Michael R. , Futreal P. Andrew , Wooster Richard
Published: March 2023

Abstract
Malignant gliomas have a very poor prognosis. The current standard of care for these cancers consists of extended adjuvant treatment with the alkylating agent temozolomide after surgical resection and radiotherapy. Although a statistically significant increase in survival has been reported with this regimen, nearly all gliomas recur and become insensitive to further treatment with this class of agents. We sequenced 500 kb of genomic DNA corresponding to the kinase domains of 518 protein kinases in each of nine gliomas. Large numbers of somatic mutations were observed in two gliomas recurrent after alkylating agent treatment. The pattern of mutations in these cases showed strong similarity to that induced by alkylating agents in experimental systems. Further investigation revealed inactivating somatic mutations of the mismatch repair gene MSH6 in each case. We propose that inactivating somatic mutations of MSH6 confer resistance to alkylating agents in gliomas in vivo and concurrently unleash accelerated mutagenesis in resistant clones as a consequence of continued exposure to alkylating agents in the presence of defective mismatch repair. The evidence therefore suggests that when MSH6 is inactivated in gliomas, alkylating agents convert from induction of tumor cell death to promotion of neoplastic progression. These observations highlight the potential of large scale sequencing for revealing and elucidating mutagenic processes operative in individual human cancers. (Cancer Res 2006; 66(8): 3987-91)
Data from DNA Methylation Pathway Alterations in an Autochthonous Murine Model of Prostate Cancer
Authors: Morey Shannon R. , Smiraglia Dominic J. , James Smitha R. , Yu Jihnhee , Moser Michael T. , Foster Barbara A. , Karpf Adam R.
Published: March 2023

Abstract
We examined the DNA methylation pathway in an autochthonous murine prostate cancer model, transgenic adenocarcinoma of mouse prostate (TRAMP). We observed that, compared with strain-matched normal prostates, primary and metastatic TRAMP tumors display increased cytosine DNA methyltransferase (Dnmt) activity, Dnmt1 and Dnmt3b protein expression, and Dnmt1, Dnmt3a, and Dnmt3b mRNA expression. Increased expression of Dnmt genes correlates with increased expression of cyclin A and E2F target genes, implicating increased cell proliferation and Rb inactivation in Dnmt overexpression. We analyzed DNA methylation in TRAMP and found that global levels of 5-methyl-2′-deoxycytidine are unaltered, whereas specific tumors display centromeric repeat hypomethylation. To interrogate locus-specific methylation, we did restriction landmark genomic scanning (RLGS) on normal prostates and primary tumors. In primary tumors, 2.3% of ∼1,200 analyzed loci display aberrant DNA hypermethylation, whereas a considerably smaller number of events show hypomethylation. The pattern of RLGS changes was nonrandom, indicating a coordinated methylation defect. Two specific genes identified by RLGS were studied in detail. Surprisingly, methylation of a downstream exon of p16(INK4a) (p16) was the highest frequency hypermethylation event identified in TRAMP, where it is associated with increased p16 mRNA and protein expression. In contrast, hypermethylation of the 5′ CpG island region of the homeobox gene Irx3 in TRAMP is associated with reduced gene expression. In summary, our data reveal a systemic DNA methylation pathway defect in TRAMP reminiscent of human prostate cancer, supporting the use of this model to investigate the functional role of DNA methylation pathway alterations in prostate cancer development. (Cancer Res 2006; 66(24): 11659-67)
Data from Mechanisms of Mitotic Cell Death Induced by Chemotherapy-Mediated G2 Checkpoint Abrogation
Authors: Vogel Celia , Hager Christian , Bastians Holger
Published: March 2023

Abstract
The novel concept of anticancer treatment termed “G2 checkpoint abrogation” aims to target p53-deficient tumor cells and is currently explored in clinical trials. The anticancer drug UCN-01 is used to abrogate a DNA damage–induced G2 cell cycle arrest leading to mitotic entry and subsequent cell death, which is poorly defined as “mitotic cell death” or “mitotic catastrophe.” We show here that UCN-01 treatment results in a mitotic arrest that requires an active mitotic spindle checkpoint, involving the function of Mad2, Bub1, BubR1, Mps1, Aurora B, and survivin. During the mitotic arrest, hallmark parameters of the mitochondria-associated apoptosis pathway become activated. Interestingly, this apoptotic response requires the spindle checkpoint protein Mad2, suggesting a proapoptotic function for Mad2. However, although survivin and Aurora B are also required for the mitotic arrest, both proteins are part of an antiapoptotic pathway that restrains the UCN-01–induced apoptosis by promoting hyperphosphorylation of Bcl-2 and by inhibiting the activation of Bax. Consequently, inhibition of the antiapoptotic pathway by genetic ablation of survivin or by pharmacologic inhibitors of Aurora B or cyclin-dependent kinase 1 lead to a significant enhancement of apoptosis and therefore act synergistically with UCN-01. Thus, by defining the mechanism of cell death on G2 checkpoint abrogation we show a highly improved strategy for an anticancer treatment by the combined use of UCN-01 with abrogators of the survivin/Aurora B–dependent antiapoptotic pathway that retains the selectivity for p53-defective cancer cells. [Cancer Res 2007;67(1):339–45]
Data from Integrin α4β1 Promotes Monocyte Trafficking and Angiogenesis in Tumors
Authors: Jin Hui , Su Jingmei , Garmy-Susini Barbara , Kleeman Jeanine , Varner Judy
Published: March 2023

Abstract
Monocytes and macrophages extensively colonize solid tumors, where they are thought to promote tumor angiogenesis. Here, we show that integrin α4β1 (VLA4) promotes the invasion of tumors by myeloid cells and subsequent neovascularization. Antagonists of integrin α4β1, but not of other integrins, blocked the adhesion of monocytes to endothelium in vitro and in vivo as well as their extravasation into tumor tissue. These antagonists prevented monocyte stimulation of angiogenesis in vivo, macrophage colonization of tumors, and tumor angiogenesis. These studies indicate the usefulness of antagonists of integrin α4β1 in suppressing macrophage colonization of tumors and subsequent tumor angiogenesis. These studies further indicate that suppression of myeloid cell homing to tumors could be a useful supplementary approach to suppress tumor angiogenesis and growth. (Cancer Res 2006; 66(4):2146-52)
Data from High-throughput Loss-of-Heterozygosity Study of Chromosome 3p in Lung Cancer Using Single-Nucleotide Polymorphism Markers
Authors: Tai Amy L.S. , Mak William , Ng Phoebe K.M. , Chua Daniel T.T. , Ng Mandy Y.M. , Fu Li , Chu Kevin K.W. , Fang Yan , Qiang Song You , Chen Muhan , Zhang Minyue , Sham Pak C. , Guan Xin-Yuan
Published: March 2023

Abstract
Loss of DNA copy number at the short arm of chromosome 3 is one of the most common genetic changes in human lung cancer, suggesting the existence of one or more tumor suppressor genes (TSG) at 3p. To identify most frequently deleted regions and candidate TSGs within these regions, a recently developed single-nucleotide polymorphism (SNP)-mass spectrometry-genotyping (SMSG) technology was applied to investigate the loss of heterozygosity (LOH) in 30 primary non–small-cell lung cancers. A total of 386 SNP markers that spanned a region of 70 Mb at 3p, from 3pter to 3p14.1, were selected for LOH analysis. The average intermarker distance in the present study is ∼180 kb. Several frequently deleted regions, including 3p26.3, 3p25.3, 3p24.1, 3p23, and 3p21.1, were found. Several candidate TSGs within these frequently detected LOH regions have been found, including APG7L at 3p25.3, CLASP2 at 3p23, and CACNA2D3 at 3p21.1. This study also showed that SMSG technology is a very useful approach to rapidly define the minimal deleted region and to identify target TSGs in a given cancer. (Cancer Res 2006; 66(8): 4133-8)
Data from Nonclassic Functions of Human Topoisomerase I: Genome-Wide and Pharmacologic Analyses
Authors: Miao Ze-Hong , Player Audrey , Shankavaram Uma , Wang Yong-Hong , Zimonjic Drazen B. , Lorenzi Philip L. , Liao Zhi-Yong , Liu Hong , Shimura Tsutomu , Zhang Hong-Liang , Meng Ling-Hua , Zhang Yong-Wei , Kawasaki Ernest S. , Popescu Nicholas C. , Aladjem Mirit I. , Goldstein David J. , Weinstein John N. , Pommier Yves
Published: March 2023

Abstract
The biological functions of nuclear topoisomerase I (Top1) have been difficult to study because knocking out TOP1 is lethal in metazoans. To reveal the functions of human Top1, we have generated stable Top1 small interfering RNA (siRNA) cell lines from colon and breast carcinomas (HCT116-siTop1 and MCF-7-siTop1, respectively). In those clones, Top1 is reduced ∼5-fold and Top2α compensates for Top1 deficiency. A prominent feature of the siTop1 cells is genomic instability, with chromosomal aberrations and histone γ-H2AX foci associated with replication defects. siTop1 cells also show rDNA and nucleolar alterations and increased nuclear volume. Genome-wide transcription profiling revealed 55 genes with consistent changes in siTop1 cells. Among them, asparagine synthetase (ASNS) expression was reduced in siTop1 cells and in cells with transient Top1 down-regulation. Conversely, Top1 complementation increased ASNS, indicating a causal link between Top1 and ASNS expression. Correspondingly, pharmacologic profiling showed l-asparaginase hypersensitivity in the siTop1 cells. Resistance to camptothecin, indenoisoquinoline, aphidicolin, hydroxyurea, and staurosporine and hypersensitivity to etoposide and actinomycin D show that Top1, in addition to being the target of camptothecins, also regulates DNA replication, rDNA stability, and apoptosis. Overall, our studies show the pleiotropic nature of human Top1 activities. In addition to its classic DNA nicking-closing functions, Top1 plays critical nonclassic roles in genomic stability, gene-specific transcription, and response to various anticancer agents. The reported cell lines and approaches described in this article provide new tools to perform detailed functional analyses related to Top1 function. [Cancer Res 2007;67(18):8752–61]
Data from Phosphoprotein Pathway Mapping: Akt/Mammalian Target of Rapamycin Activation Is Negatively Associated with Childhood Rhabdomyosarcoma Survival
Authors: Petricoin Emanuel F. , Espina Virginia , Araujo Robyn P. , Midura Brieanne , Yeung Choh , Wan Xiaolin , Eichler Gabriel S. , Johann Donald J. , Qualman Stephen , Tsokos Maria , Krishnan Kartik , Helman Lee J. , Liotta Lance A.
Published: March 2023

Abstract
Mapping of protein signaling networks within tumors can identify new targets for therapy and provide a means to stratify patients for individualized therapy. Despite advances in combination chemotherapy, the overall survival for childhood rhabdomyosarcoma remains ∼60%. A critical goal is to identify functionally important protein signaling defects associated with treatment failure for the 40% nonresponder cohort. Here, we show, by phosphoproteomic network analysis of microdissected tumor cells, that interlinked components of the Akt/mammalian target of rapamycin (mTOR) pathway exhibited increased levels of phosphorylation for tumors of patients with short-term survival. Specimens (n = 59) were obtained from the Children's Oncology Group Intergroup Rhabdomyosarcoma Study (IRS) IV, D9502 and D9803, with 12-year follow-up. High phosphorylation levels were associated with poor overall and poor disease-free survival: Akt Ser473 (overall survival P < 0.001, recurrence-free survival P < 0.0009), 4EBP1 Thr37/46 (overall survival P < 0.0110, recurrence-free survival P < 0.0106), eIF4G Ser1108 (overall survival P < 0.0017, recurrence-free survival P < 0.0072), and p70S6 Thr389 (overall survival P < 0.0085, recurrence-free survival P < 0.0296). Moreover, the findings support an altered interrelationship between the insulin receptor substrate (IRS-1) and Akt/mTOR pathway proteins (P < 0.0027) for tumors from patients with poor survival. The functional significance of this pathway was tested using CCI-779 in a mouse xenograft model. CCI-779 suppressed phosphorylation of mTOR downstream proteins and greatly reduced the growth of two different rhabdomyosarcoma (RD embryonal P = 0.00008; Rh30 alveolar P = 0.0002) cell lines compared with controls. These results suggest that phosphoprotein mapping of the Akt/mTOR pathway should be studied further as a means to select patients to receive mTOR/IRS pathway inhibitors before administration of chemotherapy. [Cancer Res 2007;67(7):3431–40]