bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from Vascular Adhesion Protein-1 Enhances Tumor Growth by Supporting Recruitment of Gr-1+CD11b+ Myeloid Cells into Tumors
Authors: Marttila-Ichihara Fumiko , Auvinen Kaisa , Elima Kati , Jalkanen Sirpa , Salmi Marko
Published: March 2024

Abstract
Cancer growth is regulated by several nonmalignant cell types, such as leukocytes and endothelial cells, which reside in the stroma of the tumor. Vascular adhesion protein-1 (VAP-1) is an amine oxidase enzyme that is expressed on the surface of endothelial cells. It supports leukocyte traffic into inflamed tissues, but nothing is known about its possible role in cancer biology in vivo. Here, we report that B16 melanoma and EL-4 lymphoma remain smaller in VAP-1–deficient mice than in wild-type controls. We found an unexpected defect in tumor angiogenesis in the absence of VAP-1. VAP-1 also selectively enhanced the recruitment of Gr-1+CD11b+ myeloid cells into the tumors. Generation of mice expressing enzymatically inactive VAP-1 showed that the oxidase activity of VAP-1 was necessary to support neoangiogenesis, myeloid cell recruitment, and tumor growth in vivo. These data describe VAP-1 as the first adhesion molecule known to be involved in the recruitment of Gr-1+CD11b+ myeloid cells into tumors. They also suggest that VAP-1 is a potential new tool for immunotherapy of tumors that could be exploited to reduce tumor burden by controlling the traffic of Gr-1+CD11b+ myeloid cells. [Cancer Res 2009;69(19):7875–83]
Data from Intravesical Immunotherapy of Superficial Bladder Cancer with Chitosan/Interleukin-12
Authors: Zaharoff David A. , Hoffman Benjamin S. , Hooper H. Brooks , Benjamin Compton J. , Khurana Kiranpreet K. , Hance Kenneth W. , Rogers Connie J. , Pinto Peter A. , Schlom Jeffrey , Greiner John W.
Published: March 2024

Abstract
Intravesical BCG has been used successfully to treat superficial bladder cancer for three decades. However, 20% to 30% of patients will fail initial BCG therapy and 30% to 50% of patients will develop recurrent tumors within 5 years. Alternative or complementary strategies for the management of superficial bladder cancer are needed. Interleukin-12 (IL-12) is a potent TH1 cytokine with robust antitumor activity and the ability to potentiate immunologic memory. Unfortunately, intravesical IL-12 did not show antitumor efficacy in a recent clinical study of patients with recurrent superficial bladder cancer. We hypothesized that coformulation of IL-12 with chitosan, a biocompatible, mucoadhesive polysaccharide, could improve intravesical IL-12 delivery and provide an effective and durable alternative for the treatment of superficial bladder cancer. In antitumor studies, 88% to 100% of mice bearing orthotopic bladder tumors were cured after four intravesical treatments with chitosan/IL-12. In contrast, only 38% to 60% of mice treated with IL-12 alone and 0% treated with BCG were cured. Antitumor responses following chitosan/IL-12 treatments were durable and provided complete protection from intravesical tumor rechallenge. Urinary cytokine analysis showed that chitosan/IL-12 induced multiple TH1 cytokines at levels significantly higher than either IL-12 alone or BCG. Immunohistochemistry revealed moderate to intense tumor infiltration by T cells and macrophages following chitosan/IL-12 treatments. Bladder submucosa from cured mice contained residual populations of immune cells that returned to baseline levels after several months. Intravesical chitosan/IL-12 is a well-tolerated, effective immunotherapy that deserves further consideration for testing in humans for the management of superficial bladder cancer. [Cancer Res 2009;69(15):6192–9]
Data from Efficacy of Levo-1-Methyl Tryptophan and Dextro-1-Methyl Tryptophan in Reversing Indoleamine-2,3-Dioxygenase–Mediated Arrest of T-Cell Proliferation in Human Epithelial Ovarian Cancer
Authors: Qian Feng , Villella Jeannine , Wallace Paul K. , Mhawech-Fauceglia Paulette , Tario Joseph D. , Andrews Christopher , Matsuzaki Junko , Valmori Danila , Ayyoub Maha , Frederick Peter J. , Beck Amy , Liao Jianqun , Cheney Richard , Moysich Kirsten , Lele Shashikant , Shrikant Protul , Old Lloyd J. , Odunsi Kunle
Published: March 2024

Abstract
It has been reported that levo-1-methyl tryptophan (L-1MT) can block indoleamine-2,3-dioxygenase (IDO) expressed by human dendritic cells (DC), whereas dextro-1-methyl tryptophan (D-1MT) is inefficient. However, whether L-1MT or D-1MT can efficiently reverse IDO-induced arrest of human T-cell proliferation has not been clarified. Here, we show a marked immunosuppressive effect of IDO derived from INDO-transfected 293 cell, IDO+ ovarian cancer cells, and monocyte-derived DCs on CD4+ Th1 cells, CD8+ T cells, and natural killer cells derived from peripheral blood, ascites, and tumors of ovarian cancer patients. We found that, whereas L-1MT and D/L-1MT can restore proliferation of tumor-derived and peripheral blood T-cell subsets, D-1MT does not effectively restore IDO-induced arrest of T-cell proliferation. Although D-1MT inhibited kynurenine production at high concentrations, L-1MT was more effective in abrogating kynurenine generation and tryptophan depletion, whereas tryptophan was completely depleted by IDO even in the presence of high amounts of D-1MT. Together, the results indicate that, whereas the generation of tryptophan metabolites (kynurenines) by IDO is important in mediating suppression of T-cell proliferation, the degree to which tryptophan depletion is restored by 1MT is also critical in overcoming IDO-induced arrest of T-cell proliferation. [Cancer Res 2009;69(13):5498–504]
Data from Epigallocatechin-3-Gallate, a Histone Acetyltransferase Inhibitor, Inhibits EBV-Induced B Lymphocyte Transformation via Suppression of RelA Acetylation
Authors: Choi Kyung-Chul , Jung Myung Gu , Lee Yoo-Hyun , Yoon Joo Chun , Kwon Seung Hyun , Kang Hee-Bum , Kim Mi-Jeong , Cha Jeong-Heon , Kim Young Jun , Jun Woo Jin , Lee Jae Myun , Yoon Ho-Geun
Published: March 2024

Abstract
Because the p300/CBP-mediated hyperacetylation of RelA (p65) is critical for nuclear factor-κB (NF-κB) activation, the attenuation of p65 acetylation is a potential molecular target for the prevention of chronic inflammation. During our ongoing screening study to identify natural compounds with histone acetyltransferase inhibitor (HATi) activity, we identified epigallocatechin-3-gallate (EGCG) as a novel HATi with global specificity for the majority of HAT enzymes but with no activity toward epigenetic enzymes including HDAC, SIRT1, and HMTase. At a dose of 100 μmol/L, EGCG abrogates p300-induced p65 acetylation in vitro and in vivo, increases the level of cytosolic IκBα, and suppresses tumor necrosis factor α (TNFα)–induced NF-κB activation. We also showed that EGCG prevents TNFα-induced p65 translocation to the nucleus, confirming that hyperacetylation is critical for NF-κB translocation as well as activity. Furthermore, EGCG treatment inhibited the acetylation of p65 and the expression of NF-κB target genes in response to diverse stimuli. Finally, EGCG reduced the binding of p300 to the promoter region of interleukin-6 gene with an increased recruitment of HDAC3, which highlights the importance of the balance between HATs and histone deacetylases in the NF-κB–mediated inflammatory signaling pathway. Importantly, EGCG at 50 μmol/L dose completely blocks EBV infection-induced cytokine expression and subsequently the EBV-induced B lymphocyte transformation. These results show the crucial role of acetylation in the development of inflammatory-related diseases. [Cancer Res 2009;69(2):583–92]
Data from c-Jun NH2-Terminal Kinase Activation Is Essential for DRAM-Dependent Induction of Autophagy and Apoptosis in 2-Methoxyestradiol–Treated Ewing Sarcoma Cells
Authors: Lorin Séverine , Borges Amélie , Ribeiro Dos Santos Lisandra , Souquère Sylvie , Pierron Gérard , Ryan Kevin M. , Codogno Patrice , Djavaheri-Mergny Mojgan
Published: March 2024

Abstract
Ewing sarcoma and osteosarcoma are two aggressive cancers that affect bones and soft tissues in children and adolescents. Despite multimodal therapy, patients with metastatic sarcoma have a poor prognosis, emphasizing a need for more effective treatment. We have shown previously that 2-methoxyestradiol (2-ME), an antitumoral compound, induces apoptosis in Ewing sarcoma cells through c-Jun NH2-terminal kinase (JNK) activation. In the present study, we provide evidence that 2-ME elicits macroautophagy, a process that participates in apoptotic responses, in a JNK-dependent manner, in Ewing sarcoma and osteosarcoma cells. We also found that the enhanced activation of JNK by 2-ME is partially regulated by p53, highlighting the relationship of JNK and autophagy to p53 signaling pathway. Furthermore, we showed that 2-ME up-regulates damage-regulated autophagy modulator (DRAM), a p53 target gene, in Ewing sarcoma cells through a mechanism that involves JNK activation. The silencing of DRAM expression reduced both apoptosis and autophagy triggered by 2-ME in Ewing sarcoma and osteosarcoma cells. Our results therefore identify JNK as a novel mediator of DRAM regulation. These findings suggest that 2-ME or other anticancer therapies that increase DRAM expression or function could be used to effectively treat sarcoma patients. [Cancer Res 2009;69(17):6924–31]
Data from [6]-Gingerol Suppresses Colon Cancer Growth by Targeting Leukotriene A4 Hydrolase
Authors: Jeong Chul-Ho , Bode Ann M. , Pugliese Angelo , Cho Yong-Yeon , Kim Hong-Gyum , Shim Jung-Hyun , Jeon Young-Jin , Li Honglin , Jiang Hualiang , Dong Zigang
Published: March 2024

Abstract
[6]-Gingerol, a natural component of ginger, exhibits anti-inflammatory and antitumorigenic activities. Despite its potential efficacy in cancer, the mechanism by which [6]-gingerol exerts its chemopreventive effects remains elusive. The leukotriene A4 hydrolase (LTA4H) protein is regarded as a relevant target for cancer therapy. Our in silico prediction using a reverse-docking approach revealed that LTA4H might be a potential target of [6]-gingerol. We supported our prediction by showing that [6]-gingerol suppresses anchorage-independent cancer cell growth by inhibiting LTA4H activity in HCT116 colorectal cancer cells. We showed that [6]-gingerol effectively suppressed tumor growth in vivo in nude mice, an effect that was mediated by inhibition of LTA4H activity. Collectively, these findings indicate a crucial role of LTA4H in cancer and also support the anticancer efficacy of [6]-gingerol targeting of LTA4H for the prevention of colorectal cancer. [Cancer Res 2009;69(13):5584–91]
Data from Interaction of E-cadherin and PTEN Regulates Morphogenesis and Growth Arrest in Human Mammary Epithelial Cells
Authors: Fournier Marcia V. , Fata Jimmie E. , Martin Katherine J. , Yaswen Paul , Bissell Mina J.
Published: March 2024

Abstract
Phosphatase and tensin homologue deleted on chromosome 10 (PTEN) is a dual-function phosphatase with tumor suppressor function compromised in a wide spectrum of cancers. Because tissue polarity and architecture are crucial modulators of normal and malignant behavior, we postulated that PTEN may play a role in maintenance of tissue integrity. We used two nonmalignant human mammary epithelial cell lines that form polarized, growth-arrested structures (acini) when cultured in three-dimensional laminin-rich extracellular matrix gels (lrECM). As acini begin to form, PTEN accumulates both in the cytoplasm and at cell-cell contacts where it colocalizes with the E-cadherin/β-catenin complex. Reduction of PTEN levels by shRNA in lrECM prevents formation of organized breast acini and disrupts growth arrest. Importantly, disruption of acinar polarity and cell-cell contact by E-cadherin function–blocking antibodies reduces endogenous PTEN protein levels and inhibits its accumulation at cell-cell contacts. Conversely, in Skbr-3 breast cancer cells lacking endogenous E-cadherin expression, exogenous introduction of E-cadherin gene causes induction of PTEN expression and its accumulation at sites of cell interactions. These studies provide evidence that E-cadherin regulates both the PTEN protein levels and its recruitment to cell-cell junctions in three-dimensional lrECM, indicating a dynamic reciprocity between architectural integrity and the levels and localization of PTEN. This interaction thus seems to be a critical integrator of proliferative and morphogenetic signaling in breast epithelial cells. [Cancer Res 2009;69(10):4545–52]
Data from Differentiation of Lung Adenocarcinoma, Pleural Mesothelioma, and Nonmalignant Pulmonary Tissues Using DNA Methylation Profiles
Authors: Christensen Brock C. , Marsit Carmen J. , Houseman E. Andres , Godleski John J. , Longacker Jennifer L. , Zheng Shichun , Yeh Ru-Fang , Wrensch Margaret R. , Wiemels Joseph L. , Karagas Margaret R. , Bueno Raphael , Sugarbaker David J. , Nelson Heather H. , Wiencke John K. , Kelsey Karl T.
Published: March 2024

Abstract
Pathologic differentiation of tissue of origin in tumors found in the lung can be challenging, with differentiation of mesothelioma and lung adenocarcinoma emblematic of this problem. Indeed, proper classification is essential for determination of treatment regimen for these diseases, making accurate and early diagnosis critical. Here, we investigate the potential of epigenetic profiles of lung adenocarcinoma, mesothelioma, and nonmalignant pulmonary tissues (n = 285) as differentiation markers in an analysis of DNA methylation at 1413 autosomal CpG loci associated with 773 cancer-related genes. Using an unsupervised recursively partitioned mixture modeling technique for all samples, the derived methylation profile classes were significantly associated with sample type (P < 0.0001). In a similar analysis restricted to tumors, methylation profile classes significantly predicted tumor type (P < 0.0001). Random forests classification of CpG methylation of tumors—which splits the data into training and test sets—accurately differentiated mesothelioma from lung adenocarcinoma over 99% of the time (P < 0.0001). In a locus-by-locus comparison of CpG methylation between tumor types, 1266 CpG loci had significantly different methylation between tumors following correction for multiple comparisons (Q < 0.05); 61% had higher methylation in adenocarcinoma. Using the CpG loci with significant differential methylation in a pathway analysis revealed significant enrichment of methylated gene-loci in Cell Cycle Regulation, DNA Damage Response, PTEN Signaling, and Apoptosis Signaling pathways in lung adenocarcinoma when compared with mesothelioma. Methylation profile–based differentiation of lung adenocarcinoma and mesothelioma is highly accurate, informs on the distinct etiologies of these diseases, and holds promise for clinical application. [Cancer Res 2009;69(15):6315–21]
Data from γ-Secretase Inhibitors Abrogate Oxaliplatin-Induced Activation of the Notch-1 Signaling Pathway in Colon Cancer Cells Resulting in Enhanced Chemosensitivity
Authors: Meng Raymond D. , Shelton Christopher C. , Li Yue-Ming , Qin Li-Xuan , Notterman Daniel , Paty Philip B. , Schwartz Gary K.
Published: March 2024

Abstract
Because Notch signaling is implicated in colon cancer tumorigenesis and protects cells from apoptosis by inducing prosurvival targets, it was hypothesized that inhibition of Notch signaling with γ-secretase inhibitors (GSI) may enhance the chemosensitivity of colon cancer cells. We first show that the Notch-1 receptor, as well as its downstream target Hes-1, is up-regulated with colon cancer progression, similar to other genes involved in chemoresistance. We then report that chemotherapy induces Notch-1, as oxaliplatin, 5-fluorouracil (5-FU), or SN-38 (the active metabolite of irinotecan) induced Notch-1 intracellular domain (NICD) protein and activated Hes-1. Induction of NICD by oxaliplatin was caused by an increase in the activity and expression of γ-secretase complex, as suppression of the protein subunit nicastrin with small interfering RNA (siRNA) prevented NICD induction after oxaliplatin. Subsequent inhibition of Notch-1 signaling with a sulfonamide GSI (GSI34) prevented the induction of NICD by chemotherapy and blunted Hes-1 activation. Blocking the activation of Notch signaling with GSI34 sensitized cells to chemotherapy and was synergistic with oxaliplatin, 5-FU, and SN-38. This chemosensitization was mediated by Notch-1, as inhibition of Notch-1 with siRNA enhanced chemosensitivity whereas overexpression of NICD increased chemoresistance. Down-regulation of Notch signaling also prevented the induction of prosurvival pathways, most notably phosphoinositide kinase-3/Akt, after oxaliplatin. In summary, colon cancer cells may up-regulate Notch-1 as a protective mechanism in response to chemotherapy. Therefore, combining GSIs with chemotherapy may represent a novel approach for treating metastatic colon cancers by mitigating the development of chemoresistance. [Cancer Res 2009;69(2):573–82]
Data from mTOR Signal and Hypoxia-Inducible Factor-1α Regulate CD133 Expression in Cancer Cells
Authors: Matsumoto Kazuko , Arao Tokuzo , Tanaka Kaoru , Kaneda Hiroyasu , Kudo Kanae , Fujita Yoshihiko , Tamura Daisuke , Aomatsu Keiichi , Tamura Tomohide , Yamada Yasuhide , Saijo Nagahiro , Nishio Kazuto
Published: March 2024

Abstract
The underlying mechanism regulating the expression of the cancer stem cell/tumor-initiating cell marker CD133/prominin-1 in cancer cells remains largely unclear, although knowledge of this mechanism would likely provide important biological information regarding cancer stem cells. Here, we found that the inhibition of mTOR signaling up-regulated CD133 expression at both the mRNA and protein levels in a CD133-overexpressing cancer cell line. This effect was canceled by a rapamycin-competitor, tacrolimus, and was not modified by conventional cytotoxic drugs. We hypothesized that hypoxia-inducible factor-1α (HIF-1α), a downstream molecule in the mTOR signaling pathway, might regulate CD133 expression; we therefore investigated the relation between CD133 and HIF-1α. Hypoxic conditions up-regulated HIF-1α expression and inversely down-regulated CD133 expression at both the mRNA and protein levels. Similarly, the HIF-1α activator deferoxamine mesylate dose-dependently down-regulated CD133 expression, consistent with the effects of hypoxic conditions. Finally, the correlations between CD133 and the expressions of HIF-1α and HIF-1β were examined using clinical gastric cancer samples. A strong inverse correlation (r = −0.68) was observed between CD133 and HIF-1α, but not between CD133 and HIF-1β. In conclusion, these results indicate that HIF-1α down-regulates CD133 expression and suggest that mTOR signaling is involved in the expression of CD133 in cancer cells. Our findings provide a novel insight into the regulatory mechanisms of CD133 expression via mTOR signaling and HIF-1α in cancer cells and might lead to insights into the involvement of the mTOR signal and oxygen-sensitive intracellular pathways in the maintenance of stemness in cancer stem cells. [Cancer Res 2009;69(18):7160–4]
Data from Reactivation of Androgen Receptor–Regulated TMPRSS2:ERG Gene Expression in Castration-Resistant Prostate Cancer
Authors: Cai Changmeng , Wang Hongyun , Xu Youyuan , Chen Shaoyong , Balk Steven P.
Published: March 2024

Abstract
It seems clear that androgen receptor (AR)–regulated expression of the TMPRSS2:ERG fusion gene plays an early role in prostate cancer (PC) development or progression, but the extent to which TMPRSS2:ERG is down-regulated in response to androgen deprivation therapy (ADT) and whether AR reactivates TMPRSS2:ERG expression in castration-resistant PC (CRPC) have not been determined. We show that ERG message levels in TMPRSS2:ERG fusion-positive CRPC are comparable with the levels in fusion gene–positive primary PC, consistent with the conclusion that the TMPRSS2:ERG expression is reactivated by AR in CRPC. To further assess whether TMPRSS2:ERG expression is initially down-regulated in response to ADT, we examined VCaP cells, which express the TMPRSS2:ERG fusion gene, and xenografts. ERG message and protein rapidly declined in response to removal of androgen in vitro and castration in vivo. Moreover, as observed in the clinical samples, ERG expression was fully restored in the VCaP xenografts that relapsed after castration, coincident with AR reactivation. AR reactivation in the relapsed xenografts was also associated with marked increases in mRNA encoding AR and androgen synthetic enzymes. These results show that expression of TMPRSS2:ERG, similarly to other AR-regulated genes, is restored in CRPC and may contribute to tumor progression. [Cancer Res 2009;69(15):6027–32]
Data from Genome-Wide Promoter Analysis of the SOX4 Transcriptional Network in Prostate Cancer Cells
Authors: Scharer Christopher D. , McCabe Colleen D. , Ali-Seyed Mohamed , Berger Michael F. , Bulyk Martha L. , Moreno Carlos S.
Published: March 2024

Abstract
SOX4 is a critical developmental transcription factor in vertebrates and is required for precise differentiation and proliferation in multiple tissues. In addition, SOX4 is overexpressed in many human malignancies, but the exact role of SOX4 in cancer progression is not well understood. Here, we have identified the direct transcriptional targets of SOX4 using a combination of genome-wide localization chromatin immunoprecipitation–chip analysis and transient overexpression followed by expression profiling in a prostate cancer model cell line. We have also used protein-binding microarrays to derive a novel SOX4-specific position-weight matrix and determined that SOX4 binding sites are enriched in SOX4-bound promoter regions. Direct transcriptional targets of SOX4 include several key cellular regulators, such as EGFR, HSP70, Tenascin C, Frizzled-5, Patched-1, and Delta-like 1. We also show that SOX4 targets 23 transcription factors, such as MLL, FOXA1, ZNF281, and NKX3-1. In addition, SOX4 directly regulates expression of three components of the RNA-induced silencing complex, namely Dicer, Argonaute 1, and RNA Helicase A. These data provide new insights into how SOX4 affects developmental signaling pathways and how these changes may influence cancer progression via regulation of gene networks involved in microRNA processing, transcriptional regulation, the TGFβ, Wnt, Hedgehog, and Notch pathways, growth factor signaling, and tumor metastasis. [Cancer Res 2009;69(2):709–17]
Data from Epithelial to Mesenchymal Transition Contributes to Drug Resistance in Pancreatic Cancer
Authors: Arumugam Thiruvengadam , Ramachandran Vijaya , Fournier Keith F. , Wang Huamin , Marquis Lauren , Abbruzzese James L. , Gallick Gary E. , Logsdon Craig D. , McConkey David J. , Choi Woonyoung
Published: March 2024

Abstract
A better understanding of drug resistance mechanisms is required to improve outcomes in patients with pancreatic cancer. Here, we characterized patterns of sensitivity and resistance to three conventional chemotherapeutic agents with divergent mechanisms of action [gemcitabine, 5-fluorouracil (5-FU), and cisplatin] in pancreatic cancer cells. Four (L3.6pl, BxPC-3, CFPAC-1, and SU86.86) were sensitive and five (PANC-1, Hs766T, AsPC-1, MIAPaCa-2, and MPanc96) were resistant to all three agents based on GI50 (50% growth inhibition). Gene expression profiling and unsupervised hierarchical clustering revealed that the sensitive and resistant cells formed two distinct groups and differed in expression of specific genes, including several features of “epithelial to mesenchymal transition” (EMT). Interestingly, an inverse correlation between E-cadherin and its transcriptional suppressor, Zeb-1, was observed in the gene expression data and was confirmed by real-time PCR. Independent validation experiment using five new pancreatic cancer cell lines confirmed that an inverse correlation between E-cadherin and Zeb-1 correlated closely with resistance to gemcitabine, 5-FU, and cisplatin. Silencing Zeb-1 in the mesenchymal lines not only increased the expression of E-cadherin but also other epithelial markers, such as EVA1 and MAL2, and restored drug sensitivity. Importantly, immunohistochemical analysis of E-cadherin and Zeb-1 in primary tumors confirmed that expression of the two proteins was mutually exclusive (P = 0.012). Therefore, our results suggest that Zeb-1 and other regulators of EMT may maintain drug resistance in human pancreatic cancer cells, and therapeutic strategies to inhibit Zeb-1 and reverse EMT should be evaluated. [Cancer Res 2009;69(14):5820–8]
Data from Identification of Inhibitors of ABCG2 by a Bioluminescence Imaging–Based High-Throughput Assay
Authors: Zhang Yimao , Byun Youngjoo , Ren Yunzhao R. , Liu Jun O. , Laterra John , Pomper Martin G.
Published: March 2024

Abstract
ABCG2 is a member of the ATP-binding cassette (ABC) family of transporters, the overexpression of which is associated with tumor resistance to a variety of chemotherapeutic agents. Accordingly, combining ABCG2 inhibitor(s) with chemotherapy has the potential to improve treatment outcome. To search for clinically useful ABCG2 inhibitors, a bioluminescence imaging (BLI)–based assay was developed to allow high-throughput compound screening. This assay exploits our finding that d-luciferin, the substrate of firefly luciferase (fLuc), is a specific substrate of ABCG2, and ABCG2 inhibitors block the export of d-luciferin and enhance bioluminescence signal by increasing intracellular d-luciferin concentrations. HEK293 cells, engineered to express ABCG2 and fLuc, were used to screen the Hopkins Drug Library that includes drugs approved by the Food and Drug Administration (FDA) as well as drug candidates that have entered phase II clinical trials. Forty-seven compounds showed BLI enhancement, a measure of anti-ABCG2 activity, of ≥5-fold, the majority of which were not previously known as ABCG2 inhibitors. The assay was validated by its identification of known ABCG2 inhibitors and by confirming previously unknown ABCG2 inhibitors using established in vitro assays (e.g., mitoxantrone resensitization and BODIPY-prazosin assays). Glafenine, a potent new inhibitor, also inhibited ABCG2 activity in vivo. The BLI-based assay is an efficient method to identify new inhibitors of ABCG2. As they were derived from a FDA-approved compound library, many of the inhibitors uncovered in this study are ready for clinical testing. [Cancer Res 2009;69(14):5867–75]
Data from Harnessing Competing Endocytic Pathways for Overcoming the Tumor-Blood Barrier: Magnetic Resonance Imaging and Near-Infrared Imaging of Bifunctional Contrast Media
Authors: Migalovich Helena Sheikhet , Kalchenko Vyacheslav , Nevo Nava , Meir Gila , Kohen Fortune , Neeman Michal
Published: March 2024

Abstract
Ovarian cancer is the most lethal gynecologic malignancy, often diagnosed at advanced stage leading to poor prognosis. In the study reported here, magnetic resonance imaging and near-infrared reflectance imaging were applied for in vivo analysis of two competing endocytic pathways affecting retention of bifunctional daidzein-bovine serum albumin (BSA)–based contrast media by human epithelial ovarian carcinoma cells. Suppression of caveolae-mediated uptake using nystatin or by BSA competition significantly enhanced daidzein-BSA-GdDTPA/CyTE777 uptake by tumor cells in vitro. In vivo, perivascular myofibroblasts generated an effective perivascular barrier excluding delivery of BSA-GdDTPA/CyTE777 to tumor cells. The ability to manipulate caveolae-mediated sequestration of albumin by perivascular tumor myofibroblasts allowed us to effectively overcome this tumor-stroma barrier, increasing delivery of daidzein-BSA-GdDTPA/CyTE777 to the tumor cells in tumor xenografts. Thus, both in vitro and in vivo, endocytosis of daidzein-BSA-GdDTPA/CyTE777 by ovarian carcinoma cells was augmented by albumin or by nystatin. In view of the cardinal role of albumin in affecting the availability and pharmacokinetics of drugs, this approach could potentially also facilitate the delivery of therapeutics and contrast media to tumor cells. [Cancer Res 2009;69(13):5610–7]
Data from Host-Derived Angiopoietin-2 Affects Early Stages of Tumor Development and Vessel Maturation but Is Dispensable for Later Stages of Tumor Growth
Authors: Nasarre Patrick , Thomas Markus , Kruse Karoline , Helfrich Iris , Wolter Vivien , Deppermann Carleen , Schadendorf Dirk , Thurston Gavin , Fiedler Ulrike , Augustin Hellmut G.
Published: March 2024

Abstract
The angiopoietin/Tie2 system has been identified as the second vascular-specific receptor tyrosine kinase system controlling vessel assembly, maturation, and quiescence. Angiopoietin-2 (Ang-2) is prominently up-regulated in the host-derived vasculature of most tumors, making it an attractive candidate for antiangiogenic intervention. Yet, the net outcome of Ang-2 functions on tumor angiogenesis is believed to be contextual depending on the local cytokine milieu. Correspondingly, Ang-2 manipulatory therapies have been shown to exert protumorigenic as well as antitumorigenic effects. To clarify the role of Ang-2 for angiogenesis and tumor growth in a definite genetic experimental setting, the present study was aimed at comparatively studying the growth of different tumors in wild-type and Ang-2–deficient mice. Lewis lung carcinomas, MT-ret melanomas, and B16F10 melanomas all grew slower in Ang-2–deficient mice. Yet, tumor growth in wild-type and Ang-2–deficient mice dissociated during early stages of tumor development, whereas tumor growth rates during later stages of primary tumor progression were similar. Analysis of the intratumoral vascular architecture revealed no major differences in microvessel density and perfusion characteristics. However, diameters of intratumoral microvessels were smaller in tumors grown in Ang-2–deficient mice, and the vasculature had an altered pattern of pericyte recruitment and maturation. Ang-2–deficient tumor vessels had higher pericyte coverage indices. Recruited pericytes were desmin and NG2 positive and predominately α-smooth muscle actin negative, indicative of a more mature pericyte phenotype. Collectively, the experiments define the role of Ang-2 during tumor angiogenesis and establish a better rationale for combination therapies involving Ang-2 manipulatory therapies. [Cancer Res 2009;69(4):1324–33]
Data from Comparative Analyses of Chromosome Alterations in Soft-Tissue Metastases within and across Patients with Castration-Resistant Prostate Cancer
Authors: Holcomb Ilona N. , Young Janet M. , Coleman Ilsa M. , Salari Keyan , Grove Douglas I. , Hsu Li , True Lawrence D. , Roudier Martine P. , Morrissey Colm M. , Higano Celestia S. , Nelson Peter S. , Vessella Robert L. , Trask Barbara J.
Published: March 2024

Abstract
Androgen deprivation is the mainstay of therapy for progressive prostate cancer. Despite initial and dramatic tumor inhibition, most men eventually fail therapy and die of metastatic castration-resistant (CR) disease. Here, we characterize the profound degree of genomic alteration found in CR tumors using array comparative genomic hybridization (array CGH), gene expression arrays, and fluorescence in situ hybridization (FISH). Bycluster analysis, we show that the similarity of the genomic profiles from primary and metastatic tumors is driven by the patient. Using data adjusted for this similarity, we identify numerous high-frequency alterations in the CR tumors, such as 8p loss and chromosome 7 and 8q gain. By integrating array CGH and expression array data, we reveal genes whose correlated values suggest they are relevant to prostate cancer biology. We find alterations that are significantly associated with the metastases of specific organ sites, and others with CR tumors versus the tumors of patients with localized prostate cancer not treated with androgen deprivation. Within the high-frequency sites of loss in CR metastases, we find an overrepresentation of genes involved in cellular lipid metabolism, including PTEN. Finally, using FISH, we verify the presence of a gene fusion between TMPRSS2 and ERG suggested by chromosome 21 deletions detected by array CGH. We find the fusion in 54% of our CR tumors, and 81% of the fusion-positive tumors contain cells with multiple copies of the fusion. Our investigation lays the foundation for a better understanding of and possible therapeutic targets for CR disease, the poorly responsive and final stage of prostate cancer. [Cancer Res 2009;69(19):7793–802]
Data from Aurora A Regulates Prometaphase Progression by Inhibiting the Ability of RASSF1A to Suppress APC-Cdc20 Activity
Authors: Song Su Jung , Song Min Sup , Kim Soon Jung , Kim So Yeon , Kwon Seung Hae , Kim Jhin Gook , Calvisi Diego F. , Kang Dongmin , Lim Dae-Sik
Published: March 2024

Abstract
The Aurora (Ipl) kinase family plays important roles in the regulation of mitosis and tumorigenesis. The tumor suppressor RASSF1A controls mitotic progression by regulating anaphase-promoting complex (APC)-Cdc20 activity and microtubule stability, but the mechanism by which this action is regulated has not been previously established. Here, we show that Aurora A and B associate with and phosphorylate RASSF1A on serine 203 in vivo at different times and in different subcellular compartments during mitosis. Notably, both depletion of Aurora A by RNA interference and expression of a nonphosphorylatable RASSF1A (S203A) mutant gene led to a marked delay in prometaphase progression. This is likely because of the failure of RASSF1A to dissociate from Cdc20, constitutive inhibition of APC-Cdc20, and accumulation of mitotic cyclins. In contrast, the delay in prometaphase progression caused by Aurora A depletion was largely normalized by phosphomimetic RASSF1A (S203D). Finally, RASSF1A phosphorylation on serine 203 was up-regulated in Aurora A–overexpressing human tumors. These findings indicate that Aurora A plays a critical role in RASSF1A-APC-Cdc20 regulatory mechanisms that control normal prometaphase progression and that are involved in tumorigenesis. [Cancer Res 2009;69(6):2314–23
Data from Caveolin-1 Regulates the Antagonistic Pleiotropic Properties of Cellular Senescence through a Novel Mdm2/p53-Mediated Pathway
Authors: Bartholomew Janine N. , Volonte Daniela , Galbiati Ferruccio
Published: March 2024

Abstract
We show that caveolin-1 is a novel binding protein for Mdm2. After oxidative stress, caveolin-1 sequesters Mdm2 away from p53, leading to stabilization of p53 and up-regulation of p21Waf1/Cip1 in human fibroblasts. Expression of a peptide corresponding to the Mdm2 binding domain of caveolin-1 is sufficient to up-regulate p53 and p21Waf1/Cip1 protein expression and induce premature senescence. Oxidative stress–induced activation of the p53/p21Waf1/Cip1 pathway and induction of premature senescence are compromised in caveolin-1 null mouse embryonic fibroblasts (MEF). We also show that reintroduction of caveolin-1 in oncogenic Ras (RasG12V)–transformed fibroblasts, which express residual levels of caveolin-1, is sufficient to promote cellular senescence. Moreover, caveolin-1 expression in MEFs is required for senescent fibroblast-induced stimulation of cell growth and tumorigenesis of both RasG12V-transformed fibroblasts and MDA-MB-231 breast cancer epithelial cells both in vitro and in vivo. Thus, our results propose caveolin-1 as a key mediator of the antagonistic pleiotropic properties of cellular senescence. [Cancer Res 2009;69(7):2878–86]
Data from Prediction of Clinical Outcome in Multiple Lung Cancer Cohorts by Integrative Genomics: Implications for Chemotherapy Selection
Authors: Broët Philippe , Camilleri-Broët Sophie , Zhang Shenli , Alifano Marco , Bangarusamy Dhinoth , Battistella Maxime , Wu Yonghui , Tuefferd Marianne , Régnard Jean-François , Lim Elaine , Tan Patrick , Miller Lance D.
Published: March 2024

Abstract
The role of adjuvant chemotherapy in patients with stage IB non–small-cell lung cancer (NSCLC) is controversial. Identifying patient subgroups with the greatest risk of relapse and, consequently, most likely to benefit from adjuvant treatment thus remains an important clinical challenge. Here, we hypothesized that recurrent patterns of genomic amplifications and deletions in lung tumors could be integrated with gene expression information to establish a robust predictor of clinical outcome in stage IB NSCLC. Using high-resolution microarrays, we generated tandem DNA copy number and gene expression profiles for 85 stage IB lung adenocarcinomas/large cell carcinomas. We identified specific copy number alterations linked to relapse-free survival and selected genes within these regions exhibiting copy number–driven expression to construct a novel integrated signature (IS) capable of predicting clinical outcome in this series (P = 0.02). Importantly, the IS also significantly predicted clinical outcome in two other independent stage I NSCLC cohorts (P = 0.003 and P = 0.025), showing its robustness. In contrast, a more conventional molecular predictor based solely on gene expression, while capable of predicting outcome in the initial series, failed to significantly predict outcome in the two independent data sets. Our results suggest that recurrent copy number alterations, when combined with gene expression information, can be successfully used to create robust predictors of clinical outcome in early-stage NSCLC. The utility of the IS in identifying early-stage NSCLC patients as candidates for adjuvant treatment should be further evaluated in a clinical trial. [Cancer Res 2009;69(3):1055–62]
Data from Constitutive Activation of the DNA Damage Signaling Pathway in Acute Myeloid Leukemia with Complex Karyotype: Potential Importance for Checkpoint Targeting Therapy
Authors: Cavelier Cindy , Didier Christine , Prade Naïs , Mansat-De Mas Véronique , Manenti Stéphane , Recher Christian , Demur Cécile , Ducommun Bernard
Published: March 2024

Abstract
Genomic instability in solid tumors participates in the oncogenetic process and is associated with the activation of the DNA damage response pathway. Here, we report the activation of the constitutive DNA damage and checkpoint pathway associated with complex karyotypes in samples from patients with acute myeloid leukemia (AML). We show that antagonizing CHK1 kinase with a small inhibitory compound or by RNA interference strongly reduces the clonogenic properties of high–DNA damage level AML samples, particularly those with complex karyotypes. Moreover, we observe a beneficial effect of CHK1 inhibition in high–DNA damage level AML samples treated with 1-β-d-arabinofuranosylcytosine. In contrast, CHK1 inhibition has no effect on the clonogenic properties of normal hematopoietic progenitors. All together, our results indicate that CHK1 inhibition may represent an attractive therapeutic opportunity in AML with complex karyotype. [Cancer Res 2009;69(22):8652–61]
Data from Vascular Cell Adhesion Molecule-1 Is a Regulator of Ovarian Cancer Peritoneal Metastasis
Authors: Slack-Davis Jill K. , Atkins Kristen A. , Harrer Christine , Hershey E. Daniel , Conaway Mark
Published: March 2024

Abstract
Ovarian cancers metastasize by attaching to and invading through the mesothelium, a single layer of mesothelial cells lining the peritoneal cavity. The presence of invasive peritoneal metastases is associated with a poor prognosis for ovarian cancer (5-year survival <25%). Vascular cell adhesion molecule-1 (VCAM-1) is a cell surface receptor that mediates leukocyte attachment and extravasation across endothelial and mesothelial monolayers at sites of inflammation. Membranous VCAM-1 expression was observed on the mesothelium of 13 of 14 women with ovarian cancer compared with 6 of 15 who were cancer-free. Using a cell culture model system of mesothelial invasion, highly tumorigenic SKOV-3 and ES-2 cells were 2.5 to 3 times more efficient in transmigration through the mesothelial monolayer compared with poorly tumorigenic OVCAR-3 cells. Blocking antibodies to, or small interfering RNA knockdown of, VCAM-1 or its ligand α4β1 integrin significantly decreased, but did not completely inhibit, transmigration of SKOV-3 cells through mesothelial monolayers. Furthermore, using a mouse model of ovarian cancer metastasis, treatment with VCAM-1 function-blocking antibodies decreased tumor burden and increased survival. Together, these observations implicate VCAM-1-α4β1 integrin interactions in the regulation of ovarian cancer cell mesothelial invasion and metastatic progression and offer the possibility of novel therapeutic targets. [Cancer Res 2009;69(4):1469–76]
Data from A Key Regulatory Role of the Transcription Factor NFATc2 in Bronchial Adenocarcinoma via CD8+ T Lymphocytes
Authors: Maxeiner Joachim H. , Karwot Roman , Sauer Kerstin , Scholtes Petra , Boross Ildiko , Koslowski Michael , Türeci Özlem , Wiewrodt Rainer , Neurath Markus F. , Lehr Hans A. , Finotto Susetta
Published: March 2024

Abstract
The Ca2+-regulated calcineurin/nuclear factor of activated T cells (NFAT) cascade controls alternative pathways of T-cell activation and peripheral tolerance. Here, we describe reduction of NFATc2 mRNA expression in the lungs of patients with bronchial adenocarcinoma. In a murine model of bronchoalveolar adenocarcinoma, mice lacking NFATc2 developed more and larger solid tumors than wild-type littermates. The extent of central tumor necrosis was decreased in the tumors in NFATc2(−/−) mice, and this finding was associated with reduced tumor necrosis factor-α and interleukin-2 (IL-2) production by CD8+ T cells. Adoptive transfer of CD8+ T cells of NFATc2(−/−) mice induced transforming growth factor-β1 in the airways of recipient mice, thus supporting CD4+CD25+Foxp-3+glucocorticoid-induced tumor necrosis factor receptor (GITR)+ regulatory T (Treg) cell survival. Finally, engagement of GITR in NFATc2(−/−) mice induced IFN-γ levels in the airways, reversed the suppression by Treg cells, and costimulated effector CD4+CD25+ (IL-2Rα) and memory CD4+CD127+ (IL-7Rα) T cells, resulting in abrogation of carcinoma progression. Agonistic signaling through GITR, in the absence of NFATc2, thus emerges as a novel possible strategy for the treatment of human bronchial adenocarcinoma in the absence of NFATc2 by enhancing IL-2Rα+ effector and IL-7Rα+ memory-expressing T cells. [Cancer Res 2009;69(7):3069–76]
Data from Equally Potent Inhibition of c-Src and Abl by Compounds that Recognize Inactive Kinase Conformations
Authors: Seeliger Markus A. , Ranjitkar Pratistha , Kasap Corynn , Shan Yibing , Shaw David E. , Shah Neil P. , Kuriyan John , Maly Dustin J.
Published: March 2024

Abstract
Imatinib is an inhibitor of the Abl tyrosine kinase domain that is effective in the treatment of chronic myelogenic leukemia. Although imatinib binds tightly to the Abl kinase domain, its affinity for the closely related kinase domain of c-Src is at least 2,000-fold lower. Imatinib recognition requires a specific inactive conformation of the kinase domain, in which a conserved Asp-Phe-Gly (DFG) motif is flipped with respect to the active conformation. The inability of c-Src to readily adopt this flipped DFG conformation was thought to underlie the selectivity of imatinib for Abl over c-Src. Here, we present a series of inhibitors (DSA compounds) that are based on the core scaffold of imatinib but which bind with equally high potency to c-Src and Abl. The DSA compounds bind to c-Src in the DFG-flipped conformation, as confirmed by crystal structures and kinetic analysis. The origin of the high affinity of these compounds for c-Src is suggested by the fact that they also inhibit clinically relevant Abl variants bearing mutations in a structural element, the P-loop, that normally interacts with the phosphate groups of ATP but is folded over a substructure of imatinib in Abl. Importantly, several of the DSA compounds block the growth of Ba/F3 cells harboring imatinib-resistant BCR-ABL mutants, including the Thr315Ile “gatekeeper” mutation, but do not suppress the growth of parental Ba/F3 cells. [Cancer Res 2009;69(6):2384–92
Data from Senescent Stromal-Derived Osteopontin Promotes Preneoplastic Cell Growth
Authors: Pazolli Ermira , Luo Xianmin , Brehm Sarah , Carbery Kelly , Chung Jun-Jae , Prior Julie L. , Doherty Jason , Demehri Shadmehr , Salavaggione Lorena , Piwnica-Worms David , Stewart Sheila A.
Published: March 2024

Abstract
Alterations in the tissue microenvironment collaborate with cell autonomous genetic changes to contribute to neoplastic progression. The importance of the microenvironment in neoplastic progression is underscored by studies showing that fibroblasts isolated from a tumor stimulate the growth of preneoplastic and neoplastic cells in xenograft models. Similarly, senescent fibroblasts promote preneoplastic cell growth in vitro and in vivo. Because senescent cells accumulate with age, their presence is hypothesized to facilitate preneoplastic cell growth and tumor formation in older individuals. To identify senescent stromal factors directly responsible for stimulating preneoplastic cell growth, we carried out whole-genome transcriptional profiling and compared senescent fibroblasts with their younger counterparts. We identified osteopontin (OPN) as one of the most highly elevated transcripts in senescent fibroblasts. Importantly, reduction of OPN protein levels by RNA interference did not affect senescence induction in fibroblasts; however, it dramatically reduced the growth-promoting activities of senescent fibroblasts in vitro and in vivo, showing that OPN is necessary for paracrine stimulation of preneoplastic cell growth. In addition, we found that recombinant OPN was sufficient to stimulate preneoplastic cell growth. Finally, we show that OPN is expressed in senescent stroma within preneoplastic lesions that arise following 7,12-dimethylbenz(a)anthracene/12-O-tetradecanoylphorbol-13-acetate treatment of mice, suggesting that stromal-derived OPN-mediated signaling events affect neoplastic progression. [Cancer Res 2009;69(3):1230–9]
Data from NKX3.1 Activates Expression of Insulin-like Growth Factor Binding Protein-3 to Mediate Insulin-like Growth Factor-I Signaling and Cell Proliferation
Authors: Muhlbradt Erin , Asatiani Ekaterina , Ortner Elizabeth , Wang Antai , Gelmann Edward P.
Published: March 2024

Abstract
NKX3.1 is a homeobox gene that codes for a haploinsufficient prostate cancer tumor suppressor. NKX3.1 protein levels are down-regulated in the majority of primary prostate cancer tissues. NKX3.1 expression in PC-3 cells increased insulin-like growth factor binding protein-3 (IGFBP-3) mRNA expression 10-fold as determined by expression microarray analysis. In both stably and transiently transfected PC-3 cells and in LNCaP cells, NKX3.1 expression increased IGFBP-3 mRNA and protein expression. In prostates of Nkx3.1 gene-targeted mice Igfbp-3 mRNA levels correlated with Nkx3.1 copy number. NKX3.1 expression in PC-3 cells attenuated the ability of insulin-like growth factor-I (IGF-I) to induce phosphorylation of type I IGF receptor (IGF-IR), insulin receptor substrate 1, phosphatidylinositol 3-kinase, and AKT. The effect of NKX3.1 on IGF-I signaling was not seen when cells were exposed to long-R3-IGF-I, an IGF-I variant peptide that does not bind to IGFBP-3. Additionally, small interfering RNA–induced knockdown of IGFBP-3 expression partially reversed the attenuation of IGF-IR signaling by NKX3.1 and abrogated NKX3.1 suppression of PC-3 cell proliferation. Thus, there is a close relationship in vitro and in vivo between NKX3.1 and IGFBP-3. The growth-suppressive effects of NKX3.1 in prostate cells are mediated, in part, by activation of IGFBP-3 expression. [Cancer Res 2009;69(6):2615–22]