bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from Targeting Cdc37 Inhibits Multiple Signaling Pathways and Induces Growth Arrest in Prostate Cancer Cells
Authors: Gray Phillip J. , Stevenson Mary Ann , Calderwood Stuart K.
Published: March 2023

Abstract
Members of the 90-kDa heat shock protein (HSP90) family are known to bind and stabilize intermediates in a wide variety of cell signaling pathways and contribute to their dysregulation in cancer. An important intracellular cofactor for HSP90 is Cdc37, a protein with a broad role in fostering the activities of protein kinases. By targeting Cdc37 using RNA interference, we have shown that the loss of Cdc37 function induces irreversible growth arrest in androgen receptor-positive and -negative prostate carcinoma cells. In contrast to HSP90-directed agents, Cdc37 targeting seems to affect cancer cells through a distinct mechanism and does not significantly deplete the intracellular levels of most known HSP90 client proteins. Instead, Cdc37 depletion inhibits cellular kinase activity and flux through growth-promoting signal transduction cascades. We show that the loss of Cdc37 leads to reduced activity of the Erk, Akt, mTOR, and androgen-induced pathways. We have also discovered synergistic interactions between Cdc37 inactivation and the HSP90-inhibitory anticancer drug 17-(allylamino)-17-demethoxygeldanamycin (17AAG). These interactions involve enhanced degradation of proteins essential for growth and inhibition of 17AAG-induced expression of the antiapoptotic HSP70. Thus, Cdc37 is essential for maintaining prostate tumor cell growth and may represent a novel target in the search for multitargeted therapies based on the HSP90 chaperone system. [Cancer Res 2007;67(24):11942–50]
Data from RIN1 Is a Breast Tumor Suppressor Gene
Authors: Milstein Marc , Mooser Chelsea K. , Hu Hailiang , Fejzo Marlena , Slamon Dennis , Goodglick Lee , Dry Sarah , Colicelli John
Published: March 2023

Abstract
Breast cancer progression is driven by altered gene expression. We show that the RIN1 gene, which encodes a RAS effector regulating epithelial cell properties, is silenced in breast tumor cell lines compared with cultured human mammary epithelial cells. We also report that RIN1 is often reduced in human breast tumor cells compared with morphologically normal breast glandular cells. At least two silencing mechanisms seem to be involved. Overexpression of the transcription repressor SNAI1 (Snail) was observed in ZR75-1 cells, and SNAI1 knockdown restored RIN1 expression. In addition, DNA methylation within the RIN1 promoter and the first exon in KPL-1 cells suggested that epigenetic modifications may contribute to silencing, and demethylation was shown to restore RIN1 expression. Reexpression of RIN1 was shown to inhibit anchorage-independent growth in soft agar. In addition, RIN1 expression inhibited both the initiation and progression of tumorigenesis for two breast tumor cell lines in a mouse model, consistent with a tumor suppressor function. We also show that RIN1 acts as a negative regulator of tumor cell invasive growth and that this requires the ABL kinase–signaling function of RIN1, suggesting a mechanism through which RIN1 silencing may contribute to breast cancer progression. [Cancer Res 2007;67(24):11510–6]
Data from Deregulated Activity of Akt in Epithelial Basal Cells Induces Spontaneous Tumors and Heightened Sensitivity to Skin Carcinogenesis
Authors: Segrelles Carmen , Lu Jerry , Hammann Brian , Santos Mirentxu , Moral Marta , Cascallana José Luis , Lara M. Fernanda , Rho Okkyung , Carbajal Steve , Traag Jeanine , Beltrán Linda , Martínez-Cruz Ana Belén , García-Escudero Ramón , Lorz Corina , Ruiz Sergio , Bravo Ana , Paramio Jesús M. , DiGiovanni John
Published: March 2023

Abstract
Aberrant activation of the phosphoinositide-3-kinase (PI3K)/PTEN/Akt pathway, leading to increased proliferation and decreased apoptosis, has been implicated in several human pathologies including cancer. Our previous data have shown that Akt-mediated signaling is an essential mediator in the mouse skin carcinogenesis system during both the tumor promotion and progression stages. In addition, overexpression of Akt is also able to transform keratinocytes through transcriptional and posttranscriptional processes. Here, we report the consequences of the increased expression of Akt1 (wtAkt) or constitutively active Akt1 (myrAkt) in the basal layer of stratified epithelia using the bovine keratin K5 promoter. These mice display alterations in epidermal proliferation and differentiation. In addition, transgenic mice with the highest levels of Akt expression developed spontaneous epithelial tumors in multiple organs with age. Furthermore, both wtAkt and myrAkt transgenic lines displayed heightened sensitivity to the epidermal proliferative effects of the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) and heightened sensitivity to two-stage skin carcinogenesis. Finally, enhanced susceptibility to two-stage carcinogenesis correlated with a more sustained proliferative response following treatment with TPA as well as sustained alterations in Akt downstream signaling pathways and elevations in cell cycle regulatory proteins. Collectively, the data provide direct support for an important role for Akt signaling in epithelial carcinogenesis in vivo, especially during the tumor promotion stage. [Cancer Res 2007;67(22):10879–88]
Data from Origins and Prevalence of the American Founder Mutation of MSH2
Authors: Clendenning Mark , Baze Mark E. , Sun Shuying , Walsh Kyle , Liyanarachchi Sandya , Fix Dan , Schunemann Victoria , Comeras Ilene , Deacon Molly , Lynch Jane F. , Gong Gordon , Thomas Brittany C. , Thibodeau Stephen N. , Lynch Henry T. , Hampel Heather , de la Chapelle Albert
Published: March 2023

Abstract
Large germline deletions within the mismatch repair gene MSH2 account for a significant proportion (up to 20%) of all deleterious mutations of this gene which are associated with Lynch syndrome. An exons 1 to 6 deletion of MSH2, originally reported in nine families, has been associated with a founding event within the United States, which genealogic studies had previously dated to 1727, and the number of present day carriers was estimated to be 18,981. Here, we report the development of a robust multiplex PCR which has assisted in the detection of 32 new families who carry the MSH2 American Founder Mutation (AFM). By offering testing to family members, 126 carriers of the AFM have been identified. Extensive genealogic studies have connected 27 of the 41 AFM families into seven extended pedigrees. These extended families have been traced back to around the 18th century without any evidence of further convergence between them. Characterization of the genomic sequence flanking the deletion and the identification of a common disease haplotype of between 0.6 and 2.3 Mb in all probands provides evidence for a common ancestor between these extended families. The DMLE+2.2 software predicts an age of ∼500 years (95% confidence interval, 425–625) for this mutation. Taken together, these data are suggestive of an earlier founding event than was first thought, which likely occurred in a European or a Native American population. The consequences of this finding would be that the AFM is significantly more frequent in the United States than was previously predicted. [Cancer Res 2008;68(7):2145–53]
Data from Functional Diversity of Human Protection of Telomeres 1 Isoforms in Telomere Protection and Cellular Senescence
Authors: Yang Qin , Zhang Ran , Horikawa Izumi , Fujita Kaori , Afshar Yalda , Kokko Antti , Laiho Päivi , Aaltonen Lauri A. , Harris Curtis C.
Published: March 2023

Abstract
Protection of telomeres 1 (POT1) proteins in various organisms bind telomeres and regulate their structure and function. In contrast to mice carrying two distinct POT1 genes encoding two POT1 proteins (POT1a and POT1b), humans have the single POT1 gene. In addition to full-length POT1 protein (variant v1), the human POT1 gene encodes four other variants due to alternative RNA splicing (variants v2, v3, v4, and v5), whose functions are poorly understood. The functional analyses of the NH2-terminally and COOH-terminally truncated POT1 variants in this study showed that neither the single-stranded telomere-binding ability of the NH2-terminal oligonucleotide-binding (OB) folds nor the telomerase-dependent telomere elongation activity mediated by the COOH-terminal TPP1-interacting domain was telomere protective by itself. Importantly, a COOH-terminally truncated variant (v5), which consists of the NH2-terminal OB folds and the central region of unknown function, was found to protect telomeres and prevent cellular senescence as efficiently as v1. Our data revealed mechanistic and functional differences between v1 and v5: (a) v1, but not v5, functions through the maintenance of telomeric 3′ overhangs; (b) p53 is indispensable to v5 knockdown-induced senescence; and (c) v5 functions at only a fraction of telomeres to prevent DNA damage signaling. Furthermore, v5 was preferentially expressed in mismatch repair (MMR)-deficient cells and tumor tissues, suggesting its role in chromosome stability associated with MMR deficiency. This study highlights a human-specific complexity in telomere protection and damage signaling conferred by functionally distinct isoforms from the single POT1 gene. [Cancer Res 2007;67(24):11677–86]
Data from Novel Somatic and Germline Mutations in Cancer Candidate Genes in Glioblastoma, Melanoma, and Pancreatic Carcinoma
Authors: Balakrishnan Asha , Bleeker Fonnet E. , Lamba Simona , Rodolfo Monica , Daniotti Maria , Scarpa Aldo , van Tilborg Angela A. , Leenstra Sieger , Zanon Carlo , Bardelli Alberto
Published: March 2023

Abstract
A recent systematic sequence analysis of well-annotated human protein coding genes or consensus coding sequences led to the identification of 189 genes displaying somatic mutations in breast and colorectal cancers. Based on their mutation prevalence, a subset of these genes was identified as cancer candidate (CAN) genes as they could be potentially involved in cancer. We evaluated the mutational profiles of 19 CAN genes in the highly aggressive tumors: glioblastoma, melanoma, and pancreatic carcinoma. Among other changes, we found novel somatic mutations in EPHA3, MLL3, TECTA, FBXW7, and OBSCN, affecting amino acids not previously found to be mutated in human cancers. Interestingly, we also found a germline nucleotide variant of OBSCN that was previously reported as a somatic mutation. Our results identify specific genetic lesions in glioblastoma, melanoma, and pancreatic cancers and indicate that CAN genes and their mutational profiles are tumor specific. Some of the mutated genes, such as the tyrosine kinase EPHA3, are clearly amenable to pharmacologic intervention and could represent novel therapeutic targets for these incurable cancers. We also speculate that similar to other oncogenes and tumor suppressor genes, mutations affecting OBSCN could be involved in cancer predisposition. [Cancer Res 2007;67(8):3545–50]
Data from Altered MicroRNA Expression Confined to Specific Epithelial Cell Subpopulations in Breast Cancer
Authors: Sempere Lorenzo F. , Christensen Mette , Silahtaroglu Asli , Bak Mads , Heath Catherine V. , Schwartz Gary , Wells Wendy , Kauppinen Sakari , Cole Charles N.
Published: March 2023

Abstract
MicroRNAs (miRNAs) are a new class of short noncoding regulatory RNAs (18–25 nucleotides) that are involved in diverse developmental and pathologic processes. Altered miRNA expression has been associated with several types of human cancer. However, most studies did not establish whether miRNA expression changes occurred within cells undergoing malignant transformation. To obtain insight into miRNA deregulation in breast cancer, we implemented an in situ hybridization (ISH) method to reveal the spatial distribution of miRNA expression in archived formalin-fixed, paraffin-embedded specimens representing normal and tumor tissue from >100 patient cases. Here, we report that expression of miR-145 and miR-205 was restricted to the myoepithelial/basal cell compartment of normal mammary ducts and lobules, whereas their accumulation was reduced or completely eliminated in matching tumor specimens. Conversely, expression of other miRNAs was detected at varying levels predominantly within luminal epithelial cells in normal tissue; expression of miR-21 was frequently increased, whereas that of let-7a was decreased in malignant cells. We also analyzed the association of miRNA expression with that of epithelial markers; prognostic indicators such as estrogen receptor, progesterone receptor, and HER2; as well as clinical outcome data. This ISH approach provides a more direct and informative assessment of how altered miRNA expression contributes to breast carcinogenesis compared with miRNA expression profiling in gross tissue biopsies. Most significantly, early manifestation of altered miR-145 expression in atypical hyperplasia and carcinoma in situ lesions suggests that this miRNA may have a potential clinical application as a novel biomarker for early detection. [Cancer Res 2007;67(24):11612–20]
Data from Detection of DNA Copy Number Changes and Oncogenic Signaling Abnormalities from Gene Expression Data Reveals MYC Activation in High-Grade Papillary Renal Cell Carcinoma
Authors: Furge Kyle A. , Chen Jindong , Koeman Julie , Swiatek Pamela , Dykema Karl , Lucin Kseniji , Kahnoski Richard , Yang Ximing J. , Teh Bin Tean
Published: March 2023

Abstract
Papillary renal cell carcinoma (RCC) represents 10% to 15% of adult renal neoplasms; however, the molecular genetic events that are associated with the development and progression of sporadic papillary RCC remain largely unclear. Papillary RCCs can be divided into two subtypes based on histologic, cytogenetic, and gene expression differences. Type 1 tumors (∼60–70%) are generally low grade with favorable outcome, whereas type 2 tumors (∼30–40%) are associated with increased cytogenetic complexity, high tumor grade, and poor prognosis. In this study, computational analysis of gene expression data derived from papillary RCC revealed that a transcriptional signature indicative of MYC pathway activation is present in high-grade type 2 papillary RCC. The MYC signature is associated with amplification of chromosome 8q and overexpression of MYC that maps to chromosome 8q24. The importance of MYC activation was confirmed by both pharmacologic and short interfering RNA–mediated inhibition of active Myc signaling in a cell line model of type 2 papillary RCC. These results provide both computational and genetic evidence that activation of Myc is associated with the aggressiveness of papillary type 2 RCC. Therefore, it will be useful to consider inhibition of components of the MYC signaling pathway as avenues for therapeutic intervention in high-grade papillary RCC. [Cancer Res 2007;67(7):3171–6]
Data from Re-Expression of Transcription Factor ATF5 in Hepatocellular Carcinoma Induces G2-M Arrest
Authors: Gho Jennifer W-M. , Ip Wai-Ki , Chan Kathy Y-Y. , Law Priscilla T-Y. , Lai Paul B-S. , Wong Nathalie
Published: March 2023

Abstract
Transcription factors represent an important class of genes that play key roles in controlling cellular proliferation, cell cycle modulation, and attractive targets for cancer therapy. Here, we report on the novel finding of common ATF5 down-regulations in hepatocellular carcinoma (HCC), a highly malignant tumor with a dismal clinical course. Array-based mapping in HCC highlighted a high and consistent incidence of transcription factor ATF5 repressions on regional chr.19q13. By quantitative reverse transcription-PCR, profound down-regulations of ATF5 were further suggested in 78% of HCC tumors (60 of 77 cases) compared to their adjacent nontumoral liver (P = 0.0004). Restoration of ATF5 expression in 3 nonexpressing HCC cell lines demonstrated a consistent growth inhibitory effect (P < 0.029) but minimal induction on cellular apoptosis. Subsequent flow cytometric investigations revealed a G2-M cell cycle arrest in HCC cells that were ectopically transfected with ATF5 (P < 0.002). The differential expressed genes from the functional effects of ATF5 were examined by array profiling. Over a hundred genes were identified, among which ID1 contains the ATF/CREB target binding sequences within its promoter region. An inverse relationship between ATF5 expressions with ID1 transcriptions was verified in HCC (P = 0.019), and a direct interaction of ATF5 on the promoter of ID1 was further demonstrated from electromobility shift assay. Examination of causal events underlying the silencing of ATF5 in HCC suggested copy number losses, promoter hypermethylation, histone deacetylation, and DNA mutations to be the likely inactivating mechanisms. In conclusion, our finding supports a tumor suppressive role for ATF5 in HCC, and highlighted ID1 as a potential downstream target. [Cancer Res 2008;68(16):6743–51]
Data from Myc-Mediated Transcriptional Repression by Recruitment of Histone Deacetylase
Authors: Kurland John F. , Tansey William P.
Published: March 2023

Abstract
Myc is a transcription factor that features prominently in cancer. The oncogenicity of Myc stems from its ability to regulate expression of genes required for cell growth and proliferation. Although the mechanisms through which Myc activates transcription have been extensively studied, less is known about how Myc represses transcription. Recently, we reported that a conserved element within Myc–MbIII– is important for transcriptional repression. Here, we investigate the mechanism through which MbIII contributes to repression. We show that Myc represses transcription of target genes Id2 and Gadd153 by a process that involves histone deacetylation. We show that MbIII is important for repression of these genes and present evidence that this element contributes to repression by recruiting the histone deacetylase HDAC3 to the Id2 and Gadd153 promoters. These results describe a mechanistic role for MbIII in transcription, and reveal that recruitment of HDAC3 is a process by which Myc represses gene activity. [Cancer Res 2008;68(10):3624–9]
Data from Divergent Mechanisms of Glucocorticoid Resistance in Experimental Models of Pediatric Acute Lymphoblastic Leukemia
Authors: Bachmann Petra S. , Gorman Rosemary , Papa Rachael A. , Bardell Jane E. , Ford Jette , Kees Ursula R. , Marshall Glenn M. , Lock Richard B.
Published: March 2023

Abstract
Cell line models of glucocorticoid resistance in childhood acute lymphoblastic leukemia (ALL) almost invariably exhibit altered glucocorticoid receptor (GR) function. However, these findings are incongruous with those using specimens derived directly from leukemia patients, in which GR alterations are rarely found. Consequently, mechanisms of glucocorticoid resistance in the clinical setting remain largely unresolved. We present a novel paradigm of glucocorticoid resistance in childhood ALL, in which patient biopsies have been directly established as continuous xenografts in immune-deficient mice, without prior in vitro culture. We show that the GRs from six highly dexamethasone-resistant xenografts (in vitro IC50 >10 μmol/L) exhibit no defects in ligand-induced nuclear translocation and binding to a consensus glucocorticoid response element (GRE). This finding contrasts with five commonly used leukemia cell lines, all of which exhibited defective GRE binding. Moreover, whereas the GRs of dexamethasone-resistant xenografts were transcriptionally active, as assessed by the ability to induce the glucocorticoid-induced leucine zipper (GILZ) gene, resistance was associated with failure to induce the bim gene, which encodes a proapoptotic BH3-only protein. Furthermore, the receptor tyrosine kinase inhibitor, SU11657, completely reversed dexamethasone resistance in a xenograft expressing functional GR, indicating that pharmacologic reversal of glucocorticoid resistance in childhood ALL is achievable. [Cancer Res 2007;67(9):4482–90]
Data from SNAI1 Is Required for Tumor Growth and Lymph Node Metastasis of Human Breast Carcinoma MDA-MB-231 Cells
Authors: Olmeda David , Moreno-Bueno Gema , Flores Juana M. , Fabra Angels , Portillo Francisco , Cano Amparo
Published: March 2023

Abstract
The transcription factor, SNAI1 (Snail), has recently been proposed as an important mediator of tumor invasion because of its role in E-cadherin down-regulation and induction of epithelial-mesenchymal transition. In human breast cancer, the expression of SNAI1 and/or the homologous SNAI2 (Slug) has been associated with E-cadherin repression, local or distant metastasis, tumor recurrence, or poor prognosis in different tumor series. However, the specific contribution of either factor to breast tumor progression is still unclear. We have analyzed the role of SNAI1 in human breast cancer by loss of function studies and provide evidence of a major role for SNAI1 in both primary tumor growth and metastasis of human breast carcinoma MDA-MB-231 cells. Specific silencing of SNAI1 by short hairpin RNA induces a decrease in mesenchymal and proinvasive markers (MMP9, ID1, SPARC) in MDA-MB-231 cells, concomitant with reduced in vitro invasive behavior. More importantly, stable SNAI1 silencing in MDA-MB-231 cells leads to a dramatic reduction of in vivo tumor incidence and growth rate. Tumors induced by MDA-MB-231-SNAI1–silenced cells show extensive necrotic regions and a significant decrease in invasive and angiogenic markers. Moreover, SNAI1 silencing increases the sensitivity of MDA-MB-231 cells to chemotherapeutics relevant in breast cancer treatments, gemcitabine and docetaxel. Remarkably, analysis of cell lines derived from lymph node metastasis indicates that SNAI1 expression is required for metastatic dissemination. [Cancer Res 2007;67(24):11721–31]
Data from Identification of Cancer Stem Cell–Like Side Population Cells in Human Nasopharyngeal Carcinoma Cell Line
Authors: Wang Jing , Guo Li-Ping , Chen Li-Zhen , Zeng Yi-Xin , Lu Shih Hsin
Published: March 2023

Abstract
Side population (SP) cells have been isolated from several solid tumors. They lack distinct molecular markers for cancer stem cells (CSC) and increasing evidence suggests that they may play an important role in tumorigenesis and cancer therapy. However, there are no reports about the existence and function of SP cells in nasopharyngeal carcinoma (NPC) cells thus far. In this study, we scanned SP cells from five NPC cell lines and investigated stem cell characteristics, such as proliferation, self-renewal, and differentiation, using SP cells from the widely-used CNE-2 NPC cell line. We observed a strong tumorigenesis ability of SP cells following in vivo transplantation into nonobese diabetic/severe combined immunodeficient mice. Immunofluorescence revealed that cytokine 19 was highly expressed on SP cells. SP cells were found to be more resistant to chemotherapy and radiotherapy and this was related to the ATP-binding cassette half transporter member 2 of G family protein and Smoothened protein expression, respectively. Our results not only showed that SP cells in human NPC cell line CNE-2 had stem cell characteristics in vitro but also showed that they had a strong ability to form tumors in vivo. Importantly, we found the cell marker, cytokine 19, may serve as a potential molecular marker for further characterization of CSC. Taken together, our data shed light on tumorigenesis and therapeutic-resistant mechanisms, which are helpful for developing novel targets for effective clinical treatment of NPC. [Cancer Res 2007;67(8):3716–24]
Data from CHR-2797: An Antiproliferative Aminopeptidase Inhibitor that Leads to Amino Acid Deprivation in Human Leukemic Cells
Authors: Krige David , Needham Lindsey A. , Bawden Lindsay J. , Flores Nicolas , Farmer Hannah , Miles Lauren E.C. , Stone Erica , Callaghan Juliana , Chandler Stephen , Clark Vanessa L. , Kirwin-Jones Patricia , Legris Valérie , Owen Jo , Patel Thakor , Wood Steve , Box Gary , Laber David , Odedra Rajesh , Wright Annette , Wood L. Michael , Eccles Suzanne A. , Bone Elisabeth A. , Ayscough Andrew , Drummond Alan H.
Published: March 2023

Abstract
CHR-2797 is a novel metalloenzyme inhibitor that is converted into a pharmacologically active acid product (CHR-79888) inside cells. CHR-79888 is a potent inhibitor of a number of intracellular aminopeptidases, including leucine aminopeptidase. CHR-2797 exerts antiproliferative effects against a range of tumor cell lines in vitro and in vivo and shows selectivity for transformed over nontransformed cells. Its antiproliferative effects are at least 300 times more potent than the prototypical aminopeptidase inhibitor, bestatin. However, the mechanism by which inhibition of these enzymes leads to proliferative changes is not understood. Gene expression microarrays were used to profile changes in mRNA expression levels in the human promyelocytic leukemia cell line HL-60 treated with CHR-2797. This analysis showed that CHR-2797 treatment induced a transcriptional response indicative of amino acid depletion, the amino acid deprivation response, which involves up-regulation of amino acid synthetic genes, transporters, and tRNA synthetases. These changes were confirmed in other leukemic cell lines sensitive to the antiproliferative effects of CHR-2797. Furthermore, CHR-2797 treatment inhibited phosphorylation of mTOR substrates and reduced protein synthesis in HL-60 cells, both also indicative of amino acid depletion. Treatment with CHR-2797 led to an increase in the concentration of intracellular small peptides, the substrates of aminopeptidases. It is suggested that aminopeptidase inhibitors, such as CHR-2797 and bestatin, deplete sensitive tumor cells of amino acids by blocking protein recycling, and this generates an antiproliferative effect. CHR-2797 is orally bioavailable and currently undergoing phase II clinical investigation in the treatment of myeloid leukemia. [Cancer Res 2008;68(16):6669–79]
Data from Induction of EBV–Latent Membrane Protein 1–Specific MHC Class II–Restricted T-Cell Responses against Natural Killer Lymphoma Cells
Authors: Kobayashi Hiroya , Nagato Toshihiro , Takahara Miki , Sato Keisuke , Kimura Shoji , Aoki Naoko , Azumi Makoto , Tateno Masatoshi , Harabuchi Yasuaki , Celis Esteban
Published: March 2023

Abstract
EBV-encoded latent membrane protein 1 (LMP1) has oncogenic potential and is expressed in many EBV-associated malignancies. Although LMP1 is regarded as a potential tumor-associated antigen for immunotherapy and several LMP1-specific MHC class I–restricted CTL epitopes have been reported, little is known regarding MHC class II–restricted CD4 helper T-lymphocyte (HTL) epitopes for LMP1. The goal of the present studies was to determine whether MHC class II–restricted CD4 T-cell responses could be induced against the LMP1 antigen and to evaluate the antitumor effect of these responses. We have combined the use of a predictive MHC class II binding peptide algorithm with in vitro vaccination of CD4 T cells using candidate peptides to identify naturally processed epitopes derived from LMP1 that elicit immune responses against EBV-expressing tumor cells. Peptide LMP1159-175 was effective in inducing HTL responses that were restricted by HLA-DR9, HLA-DR53, or HLA-DR15, indicating that this peptide behaves as a promiscuous T-cell epitope. Moreover, LMP1159-175–reactive HTL clones directly recognized EBV lymphoblastoid B cells, EBV-infected natural killer (NK)/T-lymphoma cells and naturally processed antigen in the form of LMP1+ tumor cell lysates presented by autologous dendritic cells. Because the newly identified epitope LMP1159-175 overlaps with an HLA-A2–restricted CTL epitope (LMP1159-167), this peptide might have the ability to induce simultaneous CTL and HTL responses against LMP1. Overall, our data should be relevant for the design and optimization of T-cell epitope–based immunotherapy against various EBV-associated malignancies, including NK/T cell lymphomas. [Cancer Res 2008;68(3):901–8]
Data from ABCG2 Expression and Side Population Abundance Regulated by a Transforming Growth Factor β–Directed Epithelial-Mesenchymal Transition
Authors: Yin Liqun , Castagnino Paola , Assoian Richard K.
Published: March 2023

Abstract
We describe here the regulation of ABCG2 expression and side population (SP) abundance in MCF7 human breast cancer cells. The level of ABCG2 mRNA and protein were increased in purified MCF7 SP relative to non-SP cells, and incubation with an ABCG2-specific inhibitor or ABCG2 short interfering RNA eliminated the MCF7 SP. The purified MCF7 SP could generate a heterogeneous population containing both SP and non-SP cells in culture. In vivo tumorigenicity experiments showed that the purified MCF7 SP has an increased ability to colonize the mouse mammary gland. Importantly, the MCF7 SP was depleted by a transforming growth factor-β (TGFβ)–directed epithelial-mesenchymal transition (EMT), and this effect was associated with a strong down-regulation of ABCG2 gene expression, and an increased sensitivity to mitoxantrone. ABCG2 expression and SP abundance were restored upon the removal of transforming growth factor-β and reversion of the cells to an epithelial phenotype. Knock-down of E-cadherin also reduced SP abundance, but this effect was not accompanied by the loss of ABCG2 mRNA or protein. We conclude that ABCG2 expression in MCF7 cells is regulated during an EMT, and that the EMT effect reflects posttranslational regulation of ABCG2 function by E-cadherin as well as transcriptional repression of the ABCG2 gene. [Cancer Res 2008;68(3):800–7]
Data from The Humoral Immune System Has a Key Prognostic Impact in Node-Negative Breast Cancer
Authors: Schmidt Marcus , Böhm Daniel , von Törne Christian , Steiner Eric , Puhl Alexander , Pilch Henryk , Lehr Hans-Anton , Hengstler Jan G. , Kölbl Heinz , Gehrmann Mathias
Published: March 2023

Abstract
Estrogen receptor (ER) expression and proliferative activity are established prognostic factors in breast cancer. In a search for additional prognostic motifs, we analyzed the gene expression patterns of 200 tumors of patients who were not treated by systemic therapy after surgery using a discovery approach. After performing hierarchical cluster analysis, we identified coregulated genes related to the biological process of proliferation, steroid hormone receptor expression, as well as B-cell and T-cell infiltration. We calculated metagenes as a surrogate for all genes contained within a particular cluster and visualized the relative expression in relation to time to metastasis with principal component analysis. Distinct patterns led to the hypothesis of a prognostic role of the immune system in tumors with high expression of proliferation-associated genes. In multivariate Cox regression analysis, the proliferation metagene showed a significant association with metastasis-free survival of the whole discovery cohort [hazard ratio (HR), 2.20; 95% confidence interval (95% CI), 1.40–3.46]. The B-cell metagene showed additional independent prognostic information in carcinomas with high proliferative activity (HR, 0.66; 95% CI, 0.46–0.97). A prognostic influence of the B-cell metagene was independently confirmed by multivariate analysis in a first validation cohort enriched for high-grade tumors (n = 286; HR, 0.78; 95% CI, 0.62–0.98) and a second validation cohort enriched for younger patients (n = 302; HR, 0.83; 95% CI, 0.7–0.97). Thus, we could show in three cohorts of untreated, node-negative breast cancer patients that the humoral immune system plays a pivotal role in metastasis-free survival of carcinomas of the breast. [Cancer Res 2008;68(13):5405–13]
Data from MetMAb, the One-Armed 5D5 Anti-c-Met Antibody, Inhibits Orthotopic Pancreatic Tumor Growth and Improves Survival
Authors: Jin Hongkui , Yang Renhui , Zheng Zhong , Romero Mally , Ross Jed , Bou-Reslan Hani , Carano Richard A.D. , Kasman Ian , Mai Elaine , Young Judy , Zha Jiping , Zhang Zemin , Ross Sarajane , Schwall Ralph , Colbern Gail , Merchant Mark
Published: March 2023

Abstract
The hepatocyte growth factor (HGF) and its receptor, c-Met, have been implicated in driving proliferation, invasion, and poor prognosis in pancreatic cancer. Here, we investigated the expression of HGF and c-Met in primary pancreatic cancers and described in vitro and in vivo models in which MetMAb, a monovalent antibody against c-Met, was evaluated. First, expression of HGF and MET mRNA was analyzed in 59 primary pancreatic cancers and 51 normal samples, showing that both factors are highly expressed in pancreatic cancer. We next examined HGF responsiveness in pancreatic cancer lines to select lines that proliferate in response to HGF. Based on these studies, two lines were selected for further in vivo model development: BxPC-3 (c-Met+, HGF−) and KP4 (c-Met+, HGF+) cells. As BxPC-3 cells are responsive to exogenous HGF, s.c. tumor xenografts were grown in a paracrine manner with purified human HGF provided by osmotic pumps, wherein MetMAb treatment significantly inhibited tumor growth. KP4 cells are autocrine for HGF and c-Met, and MetMAb strongly inhibited s.c. tumor growth. To better model pancreatic cancer and to enable long-term survival studies, an orthotopic model of KP4 was established. MetMAb significantly inhibited orthotopic KP4 tumor growth in 4-week studies monitored by ultrasound and also improved survival in 90-day studies. MetMAb significantly reduced c-Met phosphorylation in orthotopic KP4 tumors with a concomitant decrease in Ki-67 staining. These data suggest that the HGF/c-Met axis plays an important role in the progression of pancreatic cancer and that targeting c-Met therein may have therapeutic value. [Cancer Res 2008;68(11):4360–8]
Data from Distinct RAD51 Associations with RAD52 and BCCIP in Response to DNA Damage and Replication Stress
Authors: Wray Justin , Liu Jingmei , Nickoloff Jac A. , Shen Zhiyuan
Published: March 2023

Abstract
RAD51 has critical roles in homologous recombination (HR) repair of DNA double-strand breaks (DSB) and restarting stalled or collapsed replication forks. In yeast, Rad51 function is facilitated by Rad52 and other “mediators.” Mammalian cells express RAD52, but BRCA2 may have supplanted RAD52 in mediating RAD51 loading onto ssDNA. BCCIP interacts with BRCA2, and both proteins are important for RAD51 focus formation after ionizing radiation and HR repair of DSBs. Nonetheless, mammalian RAD52 shares biochemical activities with yeast Rad52, including RAD51 binding and single-strand annealing, suggesting a conserved role in HR. Because RAD52 and RAD51 associate, and RAD51 and BCCIP associate, we investigated the colocalization of RAD51 with BCCIP and RAD52 in human cells. We found that RAD51 colocalizes with BCCIP early after ionizing radiation, with RAD52 later, and there was little colocalization of BCCIP and RAD52. RAD52 foci are induced to a greater extent by hydroxyurea, which stalls replication forks, than by ionizing radiation. Using fluorescence recovery after photo bleaching, we show that RAD52 mobility is reduced to a greater extent by hydroxyurea than ionizing radiation. However, BCCIP showed no changes in mobility after hydroxyurea or ionizing radiation. We propose that BCCIP-dependent repair of DSBs by HR is an early RAD51 response to ionizing radiation–induced DNA damage, and that RAD52-dependent HR occurs later to restart a subset of blocked or collapsed replication forks. RAD52 and BRCA2 seem to act in parallel pathways, suggesting that targeting RAD52 in BRCA2-deficient tumors may be effective in treating these tumors. [Cancer Res 2008;68(8):2699–706]
Data from Nutlin-3a Activates p53 to Both Down-regulate Inhibitor of Growth 2 and Up-regulate mir-34a, mir-34b, and mir-34c Expression, and Induce Senescence
Authors: Kumamoto Kensuke , Spillare Elisa A. , Fujita Kaori , Horikawa Izumi , Yamashita Taro , Appella Ettore , Nagashima Makoto , Takenoshita Seiichi , Yokota Jun , Harris Curtis C.
Published: March 2023

Abstract
Nutlin-3, an MDM2 inhibitor, activates p53, resulting in several types of cancer cells undergoing apoptosis. Although p53 is mutated or deleted in ∼50% of all cancers, p53 is still functionally active in the other 50%. Consequently, nutlin-3 and similar drugs could be candidates for neoadjuvant therapy in cancers with a functional p53. Cellular senescence is also a phenotype induced by p53 activation and plays a critical role in protecting against tumor development. In this report, we found that nutlin-3a can induce senescence in normal human fibroblasts. Nutlin-3a activated and repressed a large number of p53-dependent genes, including those encoding microRNAs. mir-34a, mir-34b, and mir-34c, which have recently been shown to be downstream effectors of p53-mediated senescence, were up-regulated, and inhibitor of growth 2 (ING2) expression was suppressed by nutlin-3a treatment. Two candidates for a p53-DNA binding consensus sequence were found in the ING2 promoter regulatory region; thus, we performed chromatin immunoprecipitation and electrophoretic mobility shift assays and confirmed p53 binding directly to those sites. In addition, the luciferase activity of a construct containing the ING2 regulatory region was repressed after p53 activation. Antisense knockdown of ING2 induces p53-independent senescence, whereas overexpression of ING2 induces p53-dependent senescence. Taken together, we conclude that nutlin-3a induces senescence through p53 activation in normal human fibroblasts, and p53-mediated mir34a, mir34b, and mir34c up-regulation and ING2 down-regulation may be involved in the senescence pathway. [Cancer Res 2008;68(9):3193–203]
Data from CCN3/Nephroblastoma Overexpressed Matricellular Protein Regulates Integrin Expression, Adhesion, and Dissemination in Melanoma
Authors: Vallacchi Viviana , Daniotti Maria , Ratti Francesca , Di Stasi Delia , Deho Paola , De Filippo Annamaria , Tragni Gabrina , Balsari Andrea , Carbone Antonino , Rivoltini Licia , Parmiani Giorgio , Lazar Noureddine , Perbal Bernard , Rodolfo Monica
Published: March 2023

Abstract
CCN3/nephroblastoma overexpressed belongs to the CCN family of genes that encode secreted proteins associated with the extracellular matrix (ECM) and exert regulatory effects at the cellular level. Overexpression of CCN3 was shown in metastatic melanoma cells compared with cells of the primary tumor from the same patient. Analysis of short-term cultures from 50 primary and metastatic melanomas revealed a heterogeneous expression pattern of both the 46-kDa full-length cytoplasmic/secreted protein and the 32-kDa nuclear-truncated form. The different protein expression patterns were not associated with gene alterations or polymorphisms. Like the metastatic cells expressing high levels of the 46-kDa CCN3, cells transfected to overexpress CCN3 showed increased adhesion to ECM proteins, whereas inhibition of CCN3 expression by small interfering RNA decreased adhesion to laminin and vitronectin. CCN3 overexpression induced increased expression of laminin and vitronectin integrin receptors α7β1 and αvβ5 by increasing their mRNA production. Moreover, CCN3 secreted by melanoma cells acted as an adhesion matrix protein for melanoma cells themselves. Analysis of CCN3 protein expression with respect to melanoma progression detected the protein in all visceral metastases tested and in most nodal metastases from relapsing patients but in only a few nodal metastases from nonrelapsing patients and cutaneous metastases. Consistently, xenotransplantation in immunodeficient mice showed a higher metastatic potential of melanoma cells overexpressing CCN3. Together, these data indicate a role for CCN3 in melanoma cell interaction with the ECM by regulating integrin expression, resulting in altered cell adhesion and leading melanoma progression to aggressive disease. [Cancer Res 2008;68(3):715–23]
Data from Revealing Targeted Therapy for Human Cancer by Gene Module Maps
Authors: Wong David J. , Nuyten Dimitry S.A. , Regev Aviv , Lin Meihong , Adler Adam S. , Segal Eran , van de Vijver Marc J. , Chang Howard Y.
Published: March 2023

Abstract
A major goal of cancer research is to match specific therapies to molecular targets in cancer. Genome-scale expression profiling has identified new subtypes of cancer based on consistent patterns of variation in gene expression, leading to improved prognostic predictions. However, how these new genetic subtypes of cancers should be treated is unknown. Here, we show that a gene module map can guide the prospective identification of targeted therapies for genetic subtypes of cancer. By visualizing genome-scale gene expression in cancer as combinations of activated and deactivated functional modules, gene module maps can reveal specific functional pathways associated with each subtype that might be susceptible to targeted therapies. We show that in human breast cancers, activation of a poor-prognosis “wound signature” is strongly associated with induction of both a mitochondria gene module and a proteasome gene module. We found that 3-bromopyruvic acid, which inhibits glycolysis, selectively killed breast cells expressing the mitochondria and wound signatures. In addition, inhibition of proteasome activity by bortezomib, a drug approved for human use in multiple myeloma, abrogated wound signature expression and selectively killed breast cells expressing the wound signature. Thus, gene module maps may enable rapid translation of complex genomic signatures in human disease to targeted therapeutic strategies. [Cancer Res 2008;68(2):369–78]
Data from RhoA-Dependent Regulation of Cell Migration by the Tumor Suppressor hSNF5/INI1
Authors: Caramel Julie , Quignon Frédérique , Delattre Olivier
Published: March 2023

Abstract
Malignant rhabdoid tumors (MRT) are extremely aggressive pediatric tumors caused by the inactivation of the hSNF5/INI1 tumor suppressor gene, which encodes a core member of the SWI/SNF chromatin remodeling complex. Roles for hSNF5/INI1 in cell cycle and differentiation have been documented. Based on the observation that MRTs are highly invasive, we investigated a role for hSNF5/INI1 in cell migration. MRT cell lines exhibit high migration properties that are dramatically reduced upon hSNF5/INI1 expression. This effect is associated with the disorganization of the actin stress fiber network and is mediated by the inhibition of the activity of the small GTPase RhoA, through a nuclear, SWI/SNF-dependent transcriptional mechanism. We further show that the knockdown of hSNF5/INI1 in epithelial 293T or MCF7 cells results in increased cell size, loss of cell-cell adhesions, and enhanced migration, associated with an increased RhoA activity. Finally, we show that the SNF5 homology domain is required for hSNF5/INI1-mediated inhibition of migration, and that a missense mutation (S284L) associated with cancer is sufficient to impair hSNF5/INI1 function in migration. We conclude that the inhibition of migration is another crucial tumor suppressor function of hSNF5/INI1, in addition to its previously described functions in proliferation and differentiation, and that its loss-of-function in MRTs may account for the high invasiveness and metastatic potential of these tumors. [Cancer Res 2008;68(15):6154–61]
Data from Human Tumor Antigens Tn and Sialyl Tn Arise from Mutations in Cosmc
Authors: Ju Tongzhong , Lanneau Grainger S. , Gautam Tripti , Wang Yingchun , Xia Baoyun , Stowell Sean R. , Willard Margaret T. , Wang Wenyi , Xia Jonathan Y. , Zuna Rosemary E. , Laszik Zoltan , Benbrook Doris M. , Hanigan Marie H. , Cummings Richard D.
Published: March 2023

Abstract
Neoplastic lesions typically express specific carbohydrate antigens on glycolipids, mucins, and other glycoproteins. Such antigens are often under epigenetic control and are subject to reversion and loss upon therapeutic selective pressure. We report here that two of the most common tumor-associated carbohydrate antigens, Tn and sialyl Tn (STn), result from somatic mutations in the gene Cosmc that encodes a molecular chaperone required for formation of the active T-synthase. Diverse neoplastic lesions, including colon cancer and melanoma-derived cells lines, expressed both Tn and STn antigen due to loss-of-function mutations in Cosmc. In addition, two human cervical cancer specimens that showed expression of the Tn/STn antigens were also found to have mutations in Cosmc and loss of heterozygosity for the cross-linked Cosmc locus. This is the first example of somatic mutations in multiple types of cancers that cause global alterations in cell surface carbohydrate antigen expression. [Cancer Res 2008;68(6):1636–46]
Data from Inhibition of Glycogen Synthase Kinase 3β Attenuates Neurocognitive Dysfunction Resulting from Cranial Irradiation
Authors: Thotala Dinesh K. , Hallahan Dennis E. , Yazlovitskaya Eugenia M.
Published: March 2023

Abstract
There are now more than 10 million cancer survivors in the United States. With these numbers, chronic sequelae that result from cancer therapy have become a major health care problem. Although radiation therapy of the brain has improved cancer cure rates, learning disorders and memory deficits are a common consequence of this therapy. Here we show that glycogen synthase kinase 3β (GSK-3β) is required for radiation-induced hippocampal neuronal apoptosis and subsequent neurocognitive decline. Inhibition of GSK-3β either by small molecules (SB216763 or SB415286) or by ectopic expression of kinase-inactive GSK-3β before irradiation significantly attenuated radiation-induced apoptosis in hippocampal neurons. GSK-3β inhibition with SB216763 or SB415286 also decreased apoptosis in the subgranular zone of the hippocampus in irradiated mice, leading to improved cognitive function in irradiated animals. Studies of the molecular mechanisms of the cytoprotective effect showed that GSK-3β activity in hippocampal neurons was not significantly altered by radiation, pointing to the indirect involvement of this enzyme in radiation-induced apoptosis. At the same time, radiation led to increased accumulation of p53, whereas inhibition of the basal level of GSK-3β activity before radiation prevented p53 accumulation, suggesting a possible mechanism of cytoprotection by GSK-3β inhibitors. These findings identify GSK-3β signaling as a key regulator of radiation-induced damage in hippocampal neurons and suggest that GSK-3β inhibitors may have a therapeutic role in protecting both pediatric and adult cancer patients and may help to improve quality of life in cancer survivors. [Cancer Res 2008;68(14):5859–68]
Data from Exploration of Tumor-Suppressive MicroRNAs Silenced by DNA Hypermethylation in Oral Cancer
Authors: Kozaki Ken-ichi , Imoto Issei , Mogi Seiki , Omura Ken , Inazawa Johji
Published: March 2023

Abstract
In the last few years, microRNAs (miRNA) have started a revolution in molecular biology and emerged as key players in the carcinogenesis. They have been identified in various tumor types, showing that different sets of miRNAs are usually deregulated in different cancers. To identify the miRNA signature that was specific for oral squamous cell carcinoma (OSCC), we first examined expression profiles of 148 miRNAs in a panel of 18 OSCC cell lines and the immortalized oral keratinocyte line RT7 as a control. Compared with RT7, the expression of 54 miRNAs (36.5%) was frequently down-regulated in OSCC lines (<0.5-fold expression, ≥66.7% of 18 lines). Among these 54 miRNAs, we further analyzed four of these miRNAs (i.e., miR-34b, miR-137, miR-193a, and miR-203), located around CpG islands, to identify tumor-suppressive miRNAs silenced through aberrant DNA methylation. The expression of those four genes was restored by treatment with 5-aza-2′-deoxycytidine in OSCC cells lacking their expression. In addition, expression levels of the four miRNAs were inversely correlated with their DNA methylation status in the OSCC lines. In primary tumors of OSCC with paired normal oral mucosa, down-regulation of miRNA expression through tumor-specific hypermethylation was more frequently observed for miR-137 and miR-193a than for miR-34b and miR-203. Moreover, the ectopic transfection of miR-137 or miR-193a into OSCC lines lacking their expressions significantly reduced cell growth, with down-regulation of the translation of cyclin-dependent kinase 6 or E2F transcription factor 6, respectively. Taken together, our results clearly show that miR-137 and miR-193a are tumor suppressor miRNAs epigenetically silenced during oral carcinogenesis. [Cancer Res 2008;68(7):2094–105]