bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from Immunologic Consequences of Signal Transducers and Activators of Transcription 3 Activation in Human Squamous Cell Carcinoma
Authors: Albesiano Emilia , Davis Meghan , See Alfred P. , Han James E. , Lim Michael , Pardoll Drew M. , Kim Young
Published: March 2024

Abstract
Paracrine cross-talk between tumor cells and immune cells within the tumor microenvironment underlies local mechanisms of immune evasion. Signal transducer and activator of transcription 3 (STAT3), which is constitutively activated in diverse cancer types, is a key regulator of cytokine and chemokine expression in murine tumors, resulting in suppression of both innate and adaptive antitumor immunity. However, the immunologic effects of STAT3 activation in human cancers have not been studied in detail. To investigate how STAT3 activity in human head and neck squamous cell carcinoma (HNSCC) might alter the tumor microenvironment to enable immune escape, we used small interfering RNA and small-molecule inhibitors to suppress STAT3 activity. STAT3 inhibition in multiple primary and established human squamous carcinoma lines resulted in enhanced expression and secretion of both proinflammatory cytokines and chemokines. Although conditioned medium containing supernatants from human HNSCC inhibited lipopolysaccharide-induced dendritic cell activation in vitro, supernatants from STAT3-silenced tumor cells reversed this immune evasion mechanism. Moreover, supernatants from STAT3-silenced tumor cells were able to stimulate the migratory behavior of lymphocytes from human peripheral blood in vitro. These results show the importance of STAT3 activation in regulating the immunomodulatory mediators by human tumors and further validate STAT3 as a promising target for therapeutic intervention. Cancer Res; 70(16); 6467–76. ©2010 AACR.
Data from Vorinostat and Sorafenib Increase CD95 Activation in Gastrointestinal Tumor Cells through a Ca2+-De novo Ceramide-PP2A-Reactive Oxygen Species–Dependent Signaling Pathway
Authors: Park Margaret A. , Mitchell Clint , Zhang Guo , Yacoub Adly , Allegood Jeremy , Häussinger Dieter , Reinehr Roland , Larner Andrew , Spiegel Sarah , Fisher Paul B. , Voelkel-Johnson Christina , Ogretmen Besim , Grant Steven , Dent Paul
Published: March 2024

Abstract
The targeted therapeutics sorafenib and vorinostat interact in a synergistic fashion to kill carcinoma cells by activating CD95, and this drug combination is entering phase I evaluation. In this study, we determined how CD95 is activated by treatment with this drug combination. Low doses of sorafenib and vorinostat, but not the individual drugs, rapidly increased reactive oxygen species (ROS), Ca2+, and ceramide levels in gastrointestinal tumor cells. The production of ROS was reduced in Rho zero cells. Quenching ROS blocked drug-induced CD95 surface localization and apoptosis. ROS generation, CD95 activation, and cell killing was also blocked by quenching of induced Ca2+ levels or by inhibition of PP2A. Inhibition of acidic sphingomyelinase or de novo ceramide generation blocked the induction of ROS; however, combined inhibition of both acidic sphingomyelinase and de novo ceramide generation was required to block the induction of Ca2+. Quenching of ROS did not affect drug-induced ceramide/dihydro-ceramide levels, whereas quenching of Ca2+ reduced the ceramide increase. Sorafenib and vorinostat treatment radiosensitized liver and pancreatic cancer cells, an effect that was suppressed by quenching ROS or knockdown of LASS6. Further, sorafenib and vorinostat treatment suppressed the growth of pancreatic tumors in vivo. Our findings show that induction of cytosolic Ca2+ by sorafenib and vorinostat is a primary event that elevates dihydroceramide levels, each essential steps in ROS generation that promotes CD95 activation. Cancer Res; 70(15); 6313–24. ©2010 AACR.
Data from Targeting the Fanconi Anemia/BRCA Pathway Circumvents Drug Resistance in Multiple Myeloma
Authors: Yarde Danielle N. , Oliveira Vasco , Mathews Linda , Wang Xingyu , Villagra Alejandro , Boulware David , Shain Kenneth H. , Hazlehurst Lori A. , Alsina Melissa , Chen Dung-Tsa , Beg Amer A. , Dalton William S.
Published: March 2024

Abstract
The Fanconi anemia/BRCA (FA/BRCA) DNA damage repair pathway plays a pivotal role in the cellular response to replicative stress induced by DNA alkylating agents and greatly influences drug response in cancer treatment. We recently reported that FA/BRCA genes are overexpressed and causative for drug resistance in human melphalan-resistant multiple myeloma cell lines. However, the transcriptional regulation of the FA/BRCA pathway is not understood. In this report, we describe for the first time a novel function of the NF-κB subunits, RelB/p50, as transcriptional activators of the FA/BRCA pathway. Specifically, our findings point to constitutive phosphorylation of IκB kinase α and subsequent alterations in FANCD2 expression and function as underlying events leading to melphalan resistance in repeatedly exposed multiple myeloma cells. Inhibiting NF-κB by small interfering RNA, blocking the IκB kinase complex with BMS-345541, or using the proteasome inhibitor bortezomib drastically reduced FA/BRCA gene expression and FANCD2 protein expression in myeloma cells, resulting in diminished DNA damage repair and enhanced melphalan sensitivity. Importantly, we also found that bortezomib decreases FA/BRCA gene expression in multiple myeloma patients. These results show for the first time that NF-κB transcriptionally regulates the FA/BRCA pathway and provide evidence for targeting Fanconi anemia–mediated DNA repair to enhance chemotherapeutic response and circumvent drug resistance in myeloma patients. [Cancer Res 2009;69(24):9367–75]
Data from ARF Suppresses Tumor Angiogenesis through Translational Control of VEGFA mRNA
Authors: Kawagishi Hiroyuki , Nakamura Hideaki , Maruyama Mitsuo , Mizutani Shuki , Sugimoto Kazushi , Takagi Masatoshi , Sugimoto Masataka
Published: March 2024

Abstract
Vascular endothelial growth factor A (VEGFA) is a specific mitogen for vascular endothelial cells that plays a critical role in cancer neoangiogenesis. Here, we report that the nucleolar tumor suppressor p19ARF suppresses VEGFA expression, acting at the level of mRNA translation without affecting the transcription of the VEGFA gene. Translational repression of VEGFA mRNA by p19ARF does not require p53, a major target of the ARF tumor suppressor pathway, but instead correlates with binding to nucleophosmin/B23. Maintaining VEGFA expression relies on nucleophosmin/B23, and downregulating this protein by RNAi or p19ARF leads to translational repression of VEGFA. p19ARF inhibits VEGFA-dependent tumor angiogenesis in nude mice. Additionally, p14ARF expression and microvessel density are inversely correlated in human colon carcinomas. Taken together, our results define a mechanism by which the ARF tumor suppressor targets the translational repression of specific oncogenes during neoplastic transformation. Cancer Res; 70(11); 4749–58. ©2010 AACR.
Data from CD117 and Stro-1 Identify Osteosarcoma Tumor-Initiating Cells Associated with Metastasis and Drug Resistance
Authors: Adhikari Amit S. , Agarwal Neeraj , Wood Byron M. , Porretta Constance , Ruiz Bernardo , Pochampally Radhika R. , Iwakuma Tomoo
Published: March 2024

Abstract
Emerging evidence indicates the presence of tumor-initiating cells (TIC) or cancer stem cells in osteosarcoma. However, no study has shown specific markers to identify osteosarcoma TICs with in vivo tumor formation ability. Additionally, there has been a lack of investigations gauging the contribution of osteosarcoma TICs to metastatic and drug-resistant properties. In this study, we have identified mouse and human osteosarcoma TICs using mesenchymal stem cell markers CD117 and Stro-1. These markers were preferentially expressed in spheres and doxorubicin-resistant cells. Both mouse and human cells expressing these markers were sorted and analyzed for their abilities of tumor formation with as few as 200 cells, self-renewability, multipotency, drug resistance, metastatic potential, and enrichment of a metastasis-associated marker (CXCR4) and a drug resistance marker (ABCG2). CD117+Stro-1+ cells efficiently formed serially transplantable tumors, whereas CD117−Stro-1− cells rarely initiated tumors. On orthotopic injections, CD117+Stro-1+ cell-derived tumors metastasized at a high frequency. Further, CD117+Stro-1+ cells showed high invasive and drug-resistant properties and were efficiently enriched for CXCR4 (20–90%) and ABCG2 (60–90%). These results suggest possible mechanisms for the high metastatic and drug-resistant properties of osteosarcoma TICs. In summary, CD117 and Stro-1 identify osteosarcoma TICs associated with the most lethal characteristics of the disease—metastasis and drug resistance—and these markers offer candidates for TIC-targeted drug delivery aimed at eradicating osteosarcoma. Cancer Res; 70(11); 4602–12. ©2010 AACR.
Data from Mitochondrial Chaperone Trap1 and the Calcium Binding Protein Sorcin Interact and Protect Cells against Apoptosis Induced by Antiblastic Agents
Authors: Landriscina Matteo , Laudiero Gabriella , Maddalena Francesca , Amoroso Maria Rosaria , Piscazzi Annamaria , Cozzolino Flora , Monti Maria , Garbi Corrado , Fersini Alberto , Pucci Piero , Esposito Franca
Published: March 2024

Abstract
TRAP1, a mitochondrial chaperone (Hsp75) with antioxidant and antiapoptotic functions, is involved in multidrug resistance in human colorectal carcinoma cells. Through a proteomic analysis of TRAP1 coimmunoprecipitation complexes, the Ca2+-binding protein Sorcin was identified as a new TRAP1 interactor. This result prompted us to investigate the presence and role of Sorcin in mitochondria from human colon carcinoma cells. Using fluorescence microscopy and Western blot analysis of purified mitochondria and submitochondrial fractions, we showed the mitochondrial localization of an isoform of Sorcin with an electrophoretic motility lower than 20 kDa that specifically interacts with TRAP1. Furthermore, the effects of overexpressing or downregulating Sorcin and/or TRAP1 allowed us to demonstrate a reciprocal regulation between these two proteins and to show that their interaction is required for Sorcin mitochondrial localization and TRAP1 stability. Indeed, the depletion of TRAP1 by short hairpin RNA in colorectal carcinoma cells lowered Sorcin levels in mitochondria, whereas the depletion of Sorcin by small interfering RNA increased TRAP1 degradation. We also report several lines of evidence suggesting that intramitochondrial Sorcin plays a role in TRAP1 cytoprotection. Finally, preliminary evidence that TRAP1 and Sorcin are both implicated in multidrug resistance and are coupregulated in human colorectal carcinomas is provided. These novel findings highlight a new role for Sorcin, suggesting that some of its previously reported cytoprotective functions may be explained by involvement in mitochondrial metabolism through the TRAP1 pathway. Cancer Res; 70(16); 6577–86. ©2010 AACR.
Data from Targeted Activation of RNA Helicase Retinoic Acid–Inducible Gene-I Induces Proimmunogenic Apoptosis of Human Ovarian Cancer Cells
Authors: Kübler Kirsten , Gehrke Nadine , Riemann Soheila , Böhnert Volker , Zillinger Thomas , Hartmann Evelyn , Pölcher Martin , Rudlowski Christian , Kuhn Walther , Hartmann Gunther , Barchet Winfried
Published: March 2024

Abstract
Most malignant cells are poorly immunogenic and fail to elicit an effective antitumor immune response. In contrast, viral infections of cells are promptly detected and eliminated by the immune system. Viral recognition critically hinges on cytosolic nucleic acid receptors that include the proinflammatory RNA helicase retinoic acid–inducible gene-I (RIG-I). Here, we show that targeted delivery of RIG-I agonists induced ovarian cancer cells to upregulate HLA class I and to secrete the proinflammatory cytokines CXCL10, CCL5, interleukin-6, tumor necrosis factor-α, and IFN-β. Ovarian cancer cells stimulated via RIG-I became apoptotic and were readily phagocytosed by monocytes and monocyte-derived dendritic cells, which in turn upregulated HLA class I/II and costimulatory molecules and released CXCL10 and IFN-α. Our findings offer proof of principle that mimicking viral infection in ovarian cancer cells triggers an immunogenic form of tumor cell apoptosis that may enhance immunotherapy of ovarian cancer. Cancer Res; 70(13); 5293–304. ©2010 AACR.
Data from Selective Depletion of CD4+CD25+Foxp3+ Regulatory T Cells by Low-Dose Cyclophosphamide Is Explained by Reduced Intracellular ATP Levels
Authors: Zhao Jie , Cao Yuchun , Lei Zhang , Yang Zhuoshun , Zhang Biao , Huang Bo
Published: March 2024

Abstract
CD4+CD25+Foxp3+ regulatory T (Treg) cells have been shown to play important roles in mediating cancer development. Although cyclophosphamide (CY) has shown promise as a drug to selectively target Treg cells with low-dose in vivo, the underlying molecular mechanism remains unclear. In this report, we provide evidence that ATP, the energy molecule and signal element, accounts for the selective depletion of Treg cells by low-dose CY. Relative to conventional T cells or other cell types, ATP levels were much lower in Treg cells. This was due to Treg cells that downregulate one microRNA, miR-142-3p, and upregulate ecto-nucleoside triphosphate diphosphohydrolase CD39. The transfection of miR-142-3p or the blockade of CD39 could increase intracellular ATP levels of Treg cells, consequently decreasing the sensitivity of Treg cells to low-dose CY. On the other hand, the transfection of miR-142-3p inhibitor or the addition of soluble CD39 to the cultured CD4+CD25− T cells resulted in the decrease of intracellular ATP levels and increase of sensitivity of conventional T cells to low-dose CY. Furthermore, we found that the low levels of ATP attenuated the synthesis of glutathione, leading to the decrease of CY detoxification, thus increasing the sensitivity of Treg cells to low-dose CY. Therefore, we here identify a molecular pathway through which low-dose CY selectively ablates Treg cells. Our findings also imply that low levels of ATP are probably related to Treg cell function. Cancer Res; 70(12); 4850–8. ©2010 AACR.
Data from The Cytoskeleton Protein Filamin-A Is Required for an Efficient Recombinational DNA Double Strand Break Repair
Authors: Yue Jingyin , Wang Qin , Lu Huimei , Brenneman Mark , Fan Feiyue , Shen Zhiyuan
Published: March 2024

Abstract
The human actin-binding protein filamin-A (also known as ABP-280) cross-links actin into a dynamic three-dimensional structure. It interacts with >45 proteins of diverse functions, serving as the scaffold in various signaling networks. BRCA2 is a protein that regulates RAD51-dependent recombinational repair of DNA double strand breaks (DSB). Proximate to the COOH terminus of the BRCA2 protein, a conserved and DNA binding domain (BRCA2-DBD) interacts with filamin-A and BCCIP. In this study, we sought to test the hypothesis that filamin-A influences homologous recombinational repair of DSB and the maintenance of genomic stability. We used three pairs of cell lines with normal and reduced filamin-A expression, including breast cancer and melanoma cells. We found that lack or reduction of filamin-A sensitizes cells to ionizing radiation, slows the removal of DNA damage–induced γH2AX nuclear foci, reduces RAD51 nuclear focus formation and recruitment to chromatin in response to irradiation, and results in a 2-fold reduction of homologous recombinational repair of DSB. Furthermore, filamin-A–deficient cells have increased frequencies of micronucleus formation after irradiation. Our data illustrate the importance of the cytoskeleton structure in supporting the homologous recombinational DNA repair machinery and genome integrity, and further implicate a potential of filamin-A as a marker for prognosis in DNA damage–based cancer therapy. [Cancer Res 2009;69(20):7978–85]
Data from EWS/FLI and Its Downstream Target NR0B1 Interact Directly to Modulate Transcription and Oncogenesis in Ewing's Sarcoma
Authors: Kinsey Michelle , Smith Richard , Iyer Anita K. , McCabe Edward R.B. , Lessnick Stephen L.
Published: March 2024

Abstract
Most Ewing's sarcomas harbor chromosomal translocations that encode fusions between EWS and ETS family members. The most common fusion, EWS/FLI, consists of an EWSR1-derived strong transcriptional activation domain fused, in-frame, to the DNA-binding domain–containing portion of FLI1. EWS/FLI functions as an aberrant transcription factor to regulate genes that mediate the oncogenic phenotype of Ewing's sarcoma. One of these regulated genes, NR0B1, encodes a corepressor protein, and likely plays a transcriptional role in tumorigenesis. However, the genes that NR0B1 regulates and the transcription factors it interacts with in Ewing's sarcoma are largely unknown. We used transcriptional profiling and chromatin immunoprecipitation to identify genes that are regulated by NR0B1, and compared these data to similar data for EWS/FLI. Although the transcriptional profile overlapped as expected, we also found that the genome-wide localization of NR0B1 and EWS/FLI overlapped as well, suggesting that they regulate some genes coordinately. Further analysis revealed that NR0B1 and EWS/FLI physically interact. This protein-protein interaction is likely to be relevant for the development of Ewing's sarcoma because mutations in NR0B1 that disrupt the interaction have transcriptional consequences and also abrogate oncogenic transformation. Taken together, these data suggest that EWS/FLI and NR0B1 physically interact, coordinately modulate gene expression, and mediate the transformed phenotype of Ewing's sarcoma. [Cancer Res 2009;69(23):9047–55]
Data from Cancer Stem Cells and Aneuploid Populations within Developing Tumors Are the Major Determinants of Tumor Dormancy
Authors: Kusumbe Anjali P. , Bapat Sharmila A.
Published: March 2024

Abstract
Tumor formation involves substantial cell division and genetic instability, but the relationship between quiescent cancer stem cells (CSC) and dividing progenitors in these events is poorly understood. Likewise, the implication of aneuploid cells in solid tumors is uncertain. CSCs are postulated to contribute to tumor dormancy and present a formidable obstacle in limiting treatment outcomes for a majority of cancers, whereas the genetic heterogeneity conjured by aneuploid cells may influence tumor drug resistance. However, direct confirmation of these events remains forthcoming. In the present study, we addressed the identification of tumor dormancy in terms of isolation of therapy-refractory residual tumor cells from tumors that persist in a state of quiescence as label-retaining cells. The choices of label were PKH67/PKH26 dyes that irreversibly bind to the lipid bilayer on cell membranes and get equally partitioned among daughter cells subsequent to each cell division. Consequent characterization revealed that label-retaining cells encompass two different populations capable of remaining in a state of quiescence, i.e., stem-like cells and aneuploid cells. The former express a reversibility of quiescence through retention of functionality and also exhibit therapeutic refractoriness; the latter seem to be either quiescent or proliferation-arrested at steady-state. Subsequent to exposure to selective pressure of chemotherapy, a fraction of these cells may acquire the potential to proliferate in a drug-refractory manner and acquire stem-like characteristics. Collectively, the findings of the present study reveal that tumor-derived CSCs and aneuploid populations contribute to drug resistance and tumor dormancy in cancer progression. [Cancer Res 2009;69(24):9245–53]
Data from OTX2 Is Critical for the Maintenance and Progression of Shh-Independent Medulloblastomas
Authors: Adamson David C. , Shi Qun , Wortham Matthew , Northcott Paul A. , Di Chunhui , Duncan Christopher G. , Li Jianjun , McLendon Roger E. , Bigner Darell D. , Taylor Michael D. , Yan Hai
Published: March 2024

Abstract
OTX2 is a developmentally regulated transcription factor involved in early morphogenesis of the central nervous system. This gene is amplified and overexpressed in medulloblastoma cell lines, but the nature and extent of its genetic alterations in primary tumors have not been evaluated. Analysis of a large cohort of primary medulloblastomas revealed frequent focal copy number gain of a region minimally containing OTX2 as a single gene. OTX2 copy number gain was restricted to tumor subtypes that did not express a molecular signature of Wnt or Shh pathway activation. FISH analysis revealed copy number gain in a subset of cells within medulloblastoma samples, suggesting a late event in tumor progression. Gain of OTX2 copy number was associated with the presence of anaplastic histologic features and shorter survival in medulloblastoma patients. In support of a functional role, ectopic OTX2 expression enhanced proliferation and tumorigenicity of immortalized primary cells, whereas OTX2 knockdown in medulloblastoma cells prolonged the survival of animals bearing xenograft tumors. Mechanistic investigations revealed upregulation of MYC as a potential mechanism whereby OTX2 promotes tumor progression. Our findings define OTX2 as an important oncogenic driver in medulloblastoma. Cancer Res; 70(1); 181–91
Data from Extracellular Signal–Regulated Kinase Signaling Pathway Regulates Breast Cancer Cell Migration by Maintaining slug Expression
Authors: Chen Haoming , Zhu Genfeng , Li Yong , Padia Ravi N. , Dong Zheng , Pan Zhixing K. , Liu Kebin , Huang Shuang
Published: March 2024

Abstract
Cell migration is a critical step in cancer cell invasion. Recent studies have implicated the importance of the extracellular signal–regulated kinase (ERK) signaling pathway in cancer cell migration. However, the mechanism associated with ERK-regulated cell migration is poorly understood. Using a panel of breast cancer cell lines, we detected an excellent correlation between ERK activity and cell migration. Interestingly, we noticed that a 48-hour treatment with U0126 [specific mitogen-activated protein/ERK kinase (MEK)-1/2 inhibitor] was needed to significantly inhibit breast cancer cell migration, whereas this inhibitor blocked ERK activity within 1 hour. This observation suggests that ERK-dependent gene expression, rather than direct ERK signaling, is essential for cell migration. With further study, we found that ERK activity promoted the expression of the activator protein-1 (AP1) components Fra-1 and c-Jun, both of which were necessary for cell migration. Combination of U0126 treatment and Fra-1/c-Jun knockdown did not yield further reduction in cell migration than either alone, indicating that ERKs and Fra-1/c-Jun act by the same mechanism to facilitate cell migration. In an attempt to investigate the role of Fra-1/c-Jun in cell migration, we found that the ERK-Fra-1/c-Jun axis regulated slug expression in an AP1-dependent manner. Moreover, the occurrence of U0126-induced migratory inhibition coincided with slug reduction, and silencing slug expression abrogated breast cancer cell migration. These results suggest an association between ERK-regulated cell migration and slug expression. Indeed, cell migration was not significantly inhibited by U0126 treatment or Fra-1/c-Jun silencing in cells expressing slug transgene. Our study suggests that the ERK pathway regulates breast cancer cell migration by maintaining slug expression. [Cancer Res 2009;69(24):9228–35]
Data from PTPL1/PTPN13 Regulates Breast Cancer Cell Aggressiveness through Direct Inactivation of Src Kinase
Authors: Glondu-Lassis Murielle , Dromard Mathilde , Lacroix-Triki Magali , Nirdé Philippe , Puech Carole , Knani Dora , Chalbos Dany , Freiss Gilles
Published: March 2024

Abstract
The protein tyrosine phosphatase PTPL1/PTPN13, the activity of which is decreased through allelic loss, promoter methylation, or somatic mutations in some tumors, has been proposed as a tumor suppressor gene. Moreover, our recent clinical study identified PTPL1 expression level as an independent prognostic indicator of a favorable outcome for patients with breast cancer. However, how PTPL1 can affect tumor aggressiveness has not been characterized. Here, we first show that PTPL1 expression, assessed by immunohistochemistry, is decreased in breast cancer and metastasis specimens compared with nonmalignant tissues. Second, to evaluate whether PTPL1 plays a critical role in breast cancer progression, RNA interference experiments were performed in poorly tumorigenic MCF-7 breast cancer cells. PTPL1 inhibition drastically increased tumor growth in athymic mice and also enhanced several parameters associated with tumor progression, including cell proliferation on extracellular matrix components and cell invasion. Furthermore, the inhibition of Src kinase expression drastically blocked the effects of PTPL1 silencing on cell growth. In PTPL1 knockdown cells, the phosphorylation of Src on tyrosine 419 is increased, leading to the activation of its downstream substrates Fak and p130cas. Finally, substrate-trapping experiments revealed that Src tyrosine 419 is a direct target of the phosphatase. Thus, by identification of PTPL1 as the first phosphatase able to inhibit Src through direct dephosphorylation in intact cells, we presently describe a new mechanism by which PTPL1 inhibits breast tumor aggressiveness. Cancer Res; 70(12); 5116–26. ©2010 AACR.
Data from Three-Dimensional Imaging and Quantification of Both Solitary Cells and Metastases in Whole Mouse Liver by Magnetic Resonance Imaging
Authors: Townson Jason L. , Ramadan Soha S. , Simedrea Carmen , Rutt Brian K. , MacDonald Ian C. , Foster Paula J. , Chambers Ann F.
Published: March 2024

Abstract
The metastatic cell population, ranging from solitary cells to actively growing metastases, is heterogeneous and unlikely to respond uniformly to treatment. However, quantification of the entire experimental metastatic cell population in whole organs is complicated by requirements of an imaging modality with the large field of view and high spatial resolution necessary to detect both single cells and metastases in the same organ. Thus, it is difficult to assess differential responses of these distinct metastatic populations to therapy. Here, we develop a magnetic resonance imaging (MRI) technique capable of quantifying the full population of metastatic cells in a secondary organ. B16F1 mouse melanoma cells were labeled with micron-sized iron oxide particles (MPIO) and injected into mouse liver via the mesenteric vein. Livers were removed immediately or at day 9 or 11, following doxorubicin or vehicle control treatment, and imaged using a 3T clinical magnetic resonance scanner and custom-built gradient coil. Both metastases (>200 μm) and MPIO-labeled single cells were detected and quantified from MR images as areas of hyperintensity or hypointensity (signal voids), respectively. We found that 1mg/kg doxorubicin treatment inhibited metastasis growth (n = 11 per group; P = 0.02, t test) but did not decrease the solitary metastatic cell population in the same livers (P > 0.05). Thus, the technique presented here is capable of quickly quantifying the majority of the metastatic cell population, including both growing metastases and solitary cells, in whole liver by MRI and can identify differential responses of growing metastases and solitary cells to therapy. [Cancer Res 2009;69(21):8326–31]
Data from p53 Pre- and Post-Binding Event Theories Revisited: Stresses Reveal Specific and Dynamic p53-Binding Patterns on the p21 Gene Promoter
Authors: Millau Jean-François , Bastien Nathalie , Bouchard Éric F. , Drouin Régen
Published: March 2024

Abstract
p53 is a master transcription factor that prevents neoplasia and genomic instability. It is an important target for anticancer drug design. Understanding the molecular mechanisms behind its transcriptional activities in normal cells is a prerequisite to further understand the deregulation effected by mutant p53 in cancerous cells. Currently, how p53 coordinates transcription programs in response to stress remains unclear. One theory proposes that stresses induce pre-binding events that direct p53 to bind to specific response elements, whereas a second posits that, in response to stress, p53 binds most response elements and post-binding events then regulate transcription initiation. It is critical to establish the relevance of both theories and investigate whether stresses induce specific p53-binding patterns correlated with effector gene induction. Using unique in cellulo genomic footprinting experiments, we studied p53 binding to the five response elements of p21 in response to stresses and monitored p21 mRNA variant transcription. We show clear footprints of p53 bound to response elements in living cells and reveal that the binding of p53 to response elements is transient, subject to dynamic changes during stress responses, and influenced by response element pentamer orientations. We show further that stresses lead to specific p53-binding patterns correlated with particular p21 mRNA variant transcription profiles and that p53 binding is necessary but not sufficient to induce p21 transcription. Our results indicate that pre- and post-binding events act together to regulate adapted stress responses; this paves the way to the unification of pre- and post-binding event theories. [Cancer Res 2009;69(21):8463–71]
Data from Epigenetic Silencing of miR-137 Is an Early Event in Colorectal Carcinogenesis
Authors: Balaguer Francesc , Link Alexander , Lozano Juan Jose , Cuatrecasas Miriam , Nagasaka Takeshi , Boland C. Richard , Goel Ajay
Published: March 2024

Abstract
Global downregulation of microRNAs (miRNA) is a common feature in colorectal cancer (CRC). Whereas CpG island hypermethylation constitutes a mechanism for miRNA silencing, this field largely remains unexplored. Herein, we describe the epigenetic regulation of miR-137 and its contribution to colorectal carcinogenesis. We determined the methylation status of miR-137 CpG island in a panel of six CRC cell lines and 409 colorectal tissues [21 normal colonic mucosa from healthy individuals (N-N), 160 primary CRC tissues and their corresponding normal mucosa (N-C), and 68 adenomas]. TaqMan reverse transcription-PCR and in situ hybridization were used to analyze miR-137 expression. In vitro functional analysis of miR-137 was performed. Gene targets of miR-137 were identified using a combination of bioinformatic and transcriptomic approaches. We experimentally validated the miRNA:mRNA interactions. Methylation of the miR-137 CpG island was a cancer-specific event and was frequently observed in CRC cell lines (100%), adenomas (82.3%), and CRC (81.4%), but not in N-C (14.4%; P < 0.0001 for CRC) and N-N (4.7%; P < 0.0001 for CRC). Expression of miR-137 was restricted to the colonocytes in normal mucosa and inversely correlated with the level of methylation. Transfection of miR-137 precursor in CRC cells significantly inhibited cell proliferation. Gene expression profiling after miR-137 transfection discovered novel potential mRNA targets. We validated the interaction between miR-137 and LSD-1. Our data indicate that miR-137 acts as a tumor suppressor in the colon and is frequently silenced by promoter hypermethylation. Methylation silencing of miR-137 in colorectal adenomas suggests it to be an early event, which has prognostic and therapeutic implications. Cancer Res; 70(16); 6609–18. ©2010 AACR.
Data from microRNA-21 Negatively Regulates Cdc25A and Cell Cycle Progression in Colon Cancer Cells
Authors: Wang Peng , Zou Fangdong , Zhang Xiaodong , Li Hua , Dulak Austin , Tomko Robert J. , Lazo John S. , Wang Zhenghe , Zhang Lin , Yu Jian
Published: March 2024

Abstract
microRNAs (miRNA) are small noncoding RNAs that participate in diverse biological processes by suppressing target gene expression. Altered expression of miR-21 has been reported in cancer. To gain insights into its potential role in tumorigenesis, we generated miR-21 knockout colon cancer cells through gene targeting. Unbiased microarray analysis combined with bioinformatics identified cell cycle regulator Cdc25A as a miR-21 target. miR-21 suppressed Cdc25A expression through a defined sequence in its 3′-untranslated region. We found that miR-21 is induced by serum starvation and DNA damage, negatively regulates G1-S transition, and participates in DNA damage–induced G2-M checkpoint through down-regulation of Cdc25A. In contrast, miR-21 deficiency did not affect apoptosis induced by a variety of commonly used anticancer agents or cell proliferation under normal cell culture conditions. Furthermore, miR-21 was found to be underexpressed in a subset of Cdc25A-overexpressing colon cancers. Our data show a role of miR-21 in modulating cell cycle progression following stress, providing a novel mechanism of Cdc25A regulation and a potential explanation of miR-21 in tumorigenesis. [Cancer Res 2009;69(20):8157–65]
Data from Role of Peroxisome Proliferator-Activated Receptor-γ and Its Coactivator DRIP205 in Cellular Responses to CDDO (RTA-401) in Acute Myelogenous Leukemia
Authors: Tsao Twee , Kornblau Steven , Safe Stephen , Watt Julie C. , Ruvolo Vivian , Chen Wenjing , Qiu Yihua , Coombes Kevin R. , Ju Zhenlin , Abdelrahim Maen , Schober Wendy , Ling Xiaoyang , Kardassis Dimitris , Meyer Colin , Schimmer Aaron , Kantarjian Hagop , Andreeff Michael , Konopleva Marina
Published: March 2024

Abstract
Peroxisome proliferator-activated receptor-γ (PPARγ) is a member of the nuclear receptor (NR) family of transcription factors with important regulatory roles in cellular growth, differentiation, and apoptosis. Using proteomic analysis, we showed expression of PPARγ protein in a series of 260 newly diagnosed primary acute myelogenous leukemia (AML) samples. Forced expression of PPARγ enhanced the sensitivity of myeloid leukemic cells to apoptosis induced by PPARγ agonists 2-cyano-3,12-dioxooleana-1,9-dien-28-oic acid (CDDO) and 15-deoxy-12,14-15DPGJ2, through preferential cleavage of caspase-8. No effects on cell cycle distribution or differentiation were noted, despite prominent induction of p21 in PPARγ-transfected cells. In turn, antagonizing PPARγ function by small interfering RNA or pharmacologic PPARγ inhibitor significantly diminished apoptosis induction by CDDO. Overexpression of coactivator protein DRIP205 resulted in enhanced differentiation induction by CDDO in AML cells through PPARγ activation. Studies with DRIP205 deletion constructs showed that the NR boxes of DRIP205 are not required for this coactivation. In a phase I clinical trial of CDDO (RTA-401) in leukemia, CDDO induced an increase in PPARγ mRNA expression in six of nine patient samples; of those, induction of differentiation was documented in four patients and that of p21 in three patients, all expressing DRIP205 protein. In summary, these findings suggest that cellular levels of PPARγ regulate induction of apoptosis via caspase-8 activation, whereas the coactivator DRIP205 is a determinant of induction of differentiation, in response to PPARγ agonists in leukemic cells. Cancer Res; 70(12); 4949–60. ©2010 AACR.
Data from Tangeretin Sensitizes Cisplatin-Resistant Human Ovarian Cancer Cells through Downregulation of Phosphoinositide 3-Kinase/Akt Signaling Pathway
Authors: Arafa El-Shaimaa A. , Zhu Qianzheng , Barakat Bassant M. , Wani Gulzar , Zhao Qun , El-Mahdy Mohamed A. , Wani Altaf A.
Published: March 2024

Abstract
Combination of innocuous dietary components with anticancer drugs is an emerging new strategy for cancer chemotherapy to increase antitumor responses. Tangeretin is a citrus flavonoid known to inhibit cancer cell proliferation. Here, we show an enhanced response of A2780/CP70 and 2008/C13 cisplatin-resistant human ovarian cancer cells to various combination treatments of cisplatin and tangeretin. Pretreatment of cells with tangeretin before cisplatin treatment synergistically inhibited cancer cell proliferation. This combination was effective in activating apoptosis via caspase cascade as well as arresting cell cycle at G2-M phase. Moreover, phospho-Akt and its downstream substrates, e.g., NF-κB, phospho-GSK-3β, and phospho-BAD, were downregulated upon tangeretin-cisplatin treatment. The tangeretin-cisplatin–induced apoptosis in A2780/CP70 cells was increased by phosphoinositide-3 kinase (PI3K) inhibition and siRNA-mediated Akt silencing, but reduced by overexpression of constitutively activated Akt and GSK-3β inhibition. The overall results indicated that tangeretin exposure preconditions cisplatin-resistant human ovarian cancer cells for a conventional response to low-dose cisplatin-induced cell death occurring through downregulation of PI3K/Akt signaling pathway. Thus, effectiveness of tangeretin combinations, as a promising modality in the treatment of resistant cancers, warrants systematic clinical studies. [Cancer Res 2009;69(23):8910–7]
Data from Characterization of a Novel Mechanism of Genomic Instability Involving the SEI1/SET/NM23H1 Pathway in Esophageal Cancers
Authors: Li Yan , Nie Chang-Jun , Hu Liang , Qin Yanru , Liu Hai-bo , Zeng Ting-Ting , Chen Leilei , Fu Li , Deng Wen , Chen Shu-Peng , Jia Wei-Hua , Zhang Chunyu , Xie Dan , Guan Xin-Yuan
Published: March 2024

Abstract
Amplification of 19q is a frequent genetic alteration in many solid tumors, and SEI1 is a candidate oncogene within the amplified region. Our previous study found that the oncogenic function of SEI1 was associated with chromosome instability. In this study, we report a novel mechanism of genomic instability involving the SEI1-SET-NM23H1 pathway. Overexpression of SEI1 was observed in 57 of 100 of esophageal squamous cell carcinoma cases. Functional study showed that SEI1 had strong tumorigenic ability, and overexpression of SEI1 could induce the genomic instability by increasing micronuclei formation and reducing the number of chromosomes. Further study found that SEI1 was able to upregulate SET expression and subsequently promote the translocation of a small amount of NM23H1 from the cytoplasm to the nucleus. Nuclear NM23H1 can induce DNA damage through its DNA nick activity. Unlike CTL attack, only a small amount of NM23H1 translocated into the nucleus (<10%) induced by the overexpression of SEI1. Further study found that the small amount of NM23H1 only induced minor DNA damage and subsequently increased genomic instability, rather than inducing irreparable DNA damage and initiating apoptosis by CTL attack. Sister chromatid exchange experiment found that the translocation of small amount of NM23H1 into the nucleus induced by the overexpressions of SEI1/SET could increase the frequency of sister chromatid exchange. In addition, overexpression of SEI1 was associated with poor prognosis of esophageal squamous cell carcinoma. Taken together, these findings define a novel mechanism of genomic instability and malignant progression in esophageal cancers, a deadly disease of increasing incidence in developed countries. Cancer Res; 70(14); 5695–705. ©2010 AACR.
Data from A Gold(III) Porphyrin Complex with Antitumor Properties Targets the Wnt/β-catenin Pathway
Authors: Chow Kim Hei-Man , Sun Raymond Wai-Yin , Lam Janice B.B. , Li Carrie Ka-Lei , Xu Aimin , Ma Dik-Lung , Abagyan Ruben , Wang Yu , Che Chi-Ming
Published: March 2024

Abstract
Gold(III) complexes have shown promise as antitumor agents, but their clinical usefulness has been limited by their poor stability under physiological conditions. A novel gold(III) porphyrin complex [5-hydroxyphenyl-10,15,20-triphenylporphyrinato gold(III) chloride (gold-2a)] with improved aqueous stability showed 100-fold to 3,000-fold higher cytotoxicity than platinum-based cisplatin and IC50 values in the nanomolar range in a panel of human breast cancer cell lines. Intraductal injections of gold-2a significantly suppressed mammary tumor growth in nude mice. These effects are attributed, in part, to attenuation of Wnt/β-catenin signaling through inhibition of class I histone deacetylase (HDAC) activity. These data, in combination with computer modeling, suggest that gold-2a may represent a promising class of anticancer HDAC inhibitor preferentially targeting tumor cells with aberrant Wnt/β-catenin signaling. Cancer Res; 70(1); 329–37
Data from Wip1 Directly Dephosphorylates γ-H2AX and Attenuates the DNA Damage Response
Authors: Cha Hyukjin , Lowe Julie M. , Li Henghong , Lee Ji-Seon , Belova Galina I. , Bulavin Dmitry V. , Fornace Albert J.
Published: March 2024

Abstract
The integrity of DNA is constantly challenged throughout the life of a cell by both endogenous and exogenous stresses. A well-organized rapid damage response and proficient DNA repair, therefore, become critically important for maintaining genomic stability and cell survival. When DNA is damaged, the DNA damage response (DDR) can be initiated by alterations in chromosomal structure and histone modifications, such as the phosphorylation of the histone H2AX (the phosphorylated form is referred to as γ-H2AX). γ-H2AX plays a crucial role in recruiting DDR factors to damage sites for accurate DNA repair. On repair completion, γ-H2AX must then be reverted to H2AX by dephosphorylation for attenuation of the DDR. Here, we report that the wild-type p53–induced phosphatase 1 (Wip1) phosphatase, which is often overexpressed in a variety of tumors, effectively dephosphorylates γ-H2AX in vitro and in vivo. Ectopic expression of Wip1 significantly reduces the level of γ-H2AX after ionizing as well as UV radiation. Forced premature dephosphorylation of γ-H2AX by Wip1 disrupts recruitment of important DNA repair factors to damaged sites and delays DNA damage repair. Additionally, deletion of Wip1 enhances γ-H2AX levels in cells undergoing constitutive oncogenic stress. Taken together, our studies show that Wip1 is an important mammalian phosphatase for γ-H2AX and shows an additional mechanism for Wip1 in the tumor surveillance network. Cancer Res; 70(10); 4112–22. ©2010 AACR.
Data from Impaired Skin and Mammary Gland Development and Increased γ-Irradiation–Induced Tumorigenesis in Mice Carrying a Mutation of S1152-ATM Phosphorylation Site in Brca1
Authors: Kim Sang Soo , Cao Liu , Baek Hye Jung , Lim Sung-Chul , Li Cuiling , Wang Rui-Hong , Xu Xiaoling , Cho Kwan Ho , Deng Chu-Xia
Published: March 2024

Abstract
The tumor suppressor BRCA1 interacts with many proteins and undergoes multiple modifications on DNA damage. ATM, a key molecule of the DNA damage response, phosphorylates S1189 of BRCA1 after γ-irradiation. S1189 of BRCA1 is known as a unique ATM phosphorylation site in BRCA1 exon 11. To study the functions of ATM-dependent phosphorylation of BRCA1-S1189, we generated a mouse model carrying a mutation of S1152A (S1152 in mouse Brca1 corresponds to S1189 in human BRCA1) by gene targeting. Brca1S1152A/S1152A mice were born at the expected ratio, unlike that seen in previous studies of Brca1-null mice. However, 36% of Brca1S1152A/S1152A mice exhibited aging-like phenotypes including growth retardation, skin abnormalities, and delay of the mammary gland morphogenesis, with an increase in apoptosis. Mutant mice were hypersensitive to high doses of γ-irradiation, displaying shortened life span and reduction in intestinal villus size, associated with increased apoptosis. Aging-unaffected 18-month-old Brca1S1152A/S1152A female mice also showed mammary gland abnormalities with increased levels of cyclin D1 and phospho-ER-α, such as Brca1-Δ11 mutation. On low-dose γ-irradiation, they suffered a marked increase in tumor formation with an abnormal coat pattern. Furthermore, Brca1S1152A/S1152A embryonic fibroblasts failed to accumulate p53 on γ-irradiation with delayed phosphorylation of p53-S23. These observations indicate that ATM-mediated phosphorylation of S1189 is required for BRCA1 functions in the modulation of DNA damage response and in the suppression of tumor formation by regulating p53 and apoptosis. [Cancer Res 2009;69(24):9291–300]
Data from Identification of ASF/SF2 as a Critical, Allele-Specific Effector of the Cyclin D1b Oncogene
Authors: Olshavsky Nicholas A. , Comstock Clay E.S. , Schiewer Matthew J. , Augello Michael A. , Hyslop Terry , Sette Claudio , Zhang Jinsong , Parysek Linda M. , Knudsen Karen E.
Published: March 2024

Abstract
The cyclin D1b oncogene arises from alternative splicing of the CCND1 transcript, and harbors markedly enhanced oncogenic functions not shared by full-length cyclin D1 (cyclin D1a). Recent studies showed that cyclin D1b is selectively induced in a subset of tissues as a function of tumorigenesis; however, the underlying mechanism(s) that control tumor-specific cyclin D1b induction remain unsolved. Here, we identify the RNA-binding protein ASF/SF2 as a critical, allele-specific, disease-relevant effector of cyclin D1b production. Initially, it was observed that SF2 associates with cyclin D1b mRNA (transcript-b) in minigene analyses and with endogenous transcript in prostate cancer (PCa) cells. SF2 association was altered by the CCND1 G/A870 polymorphism, which resides in the splice donor site controlling transcript-b production. This finding was significant, as the A870 allele promotes cyclin D1b in benign prostate tissue, but in primary PCa, cyclin D1b production is independent of A870 status. Data herein provide a basis for this disparity, as tumor-associated induction of SF2 predominantly results in binding to and accumulation of G870-derived transcript-b. Finally, the relevance of SF2 function was established, as SF2 strongly correlated with cyclin D1b (but not cyclin D1a) in human PCa. Together, these studies identify a novel mechanism by which cyclin D1b is induced in cancer, and reveal significant evidence of a factor that cooperates with a risk-associated polymorphism to alter cyclin D1 isoform production. Identification of SF2 as a disease-relevant effector of cyclin D1b provides a basis for future studies designed to suppress the oncogenic alternative splicing event. Cancer Res; 70(10); 3975–84. ©2010 AACR.
Data from Myocardin Functions as an Effective Inducer of Growth Arrest and Differentiation in Human Uterine Leiomyosarcoma Cells
Authors: Kimura Yasunori , Morita Tsuyoshi , Hayashi Ken'ichiro , Miki Tsuneharu , Sobue Kenji
Published: March 2024

Abstract
Myocardin is an important transcriptional regulator in smooth and cardiac muscle development. We noticed that the expression of myocardin was markedly downregulated in human uterine leiomyosarcoma cells. Restoration of myocardin expression induced the reexpression of smooth muscle marker proteins and the formation of well-developed actin fibers. A concomitant increase in the expression of a cyclin-dependent kinase inhibitor, p21, led to significantly reduced cell proliferation, via p21's inhibition of the G1-S transition. A p21 promoter-reporter assay showed that myocardin markedly increased p21's promoter activity. Furthermore, a serum response factor (SRF)–binding cis-element CArG box in the p21 promoter region was required for this myocardin effect. Chromatin immunoprecipitation and DNA-protein binding assays showed that myocardin indirectly bound to the CArG box in the p21 promoter through the interaction with SRF. Furthermore, immunohistochemistry revealed that the levels of myocardin and p21 were both lower in leiomyosarcoma samples than in normal smooth muscle tissue. Taken together, our results indicate that the downregulation of myocardin expression facilitates cell cycle progression via the reduction of p21 expression in human leimyosarcomas and suggest that myocardin could be a useful therapeutic target for this disease. Cancer Res; 70(2); 501–11