bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from WEE1 Inhibition Alleviates Resistance to Immune Attack of Tumor Cells Undergoing Epithelial–Mesenchymal Transition
Authors: Hamilton Duane H. , Huang Bruce , Fernando Romaine I. , Tsang Kwong-Yok , Palena Claudia
Published: March 2024

Abstract
Aberrant expression of the T-box transcription factor brachyury in human carcinomas drives the phenomenon of epithelial–mesenchymal transition (EMT), a phenotypic modulation that facilitates tumor dissemination and resistance to conventional therapies, including chemotherapy and radiotherapy. By generating isogenic cancer cell lines with various levels of brachyury expression, we demonstrate that high levels of brachyury also significantly reduce the susceptibility of cancer cells to lysis by both antigen-specific T cells and natural killer cells. Our results indicated that resistance of brachyury-high tumor cells to immune-mediated attack was due to inefficient caspase-dependent apoptosis, manifested as inefficient nuclear lamin degradation in the presence of activated effector caspases. We correlated this phenomenon with loss of cell-cycle–dependent kinase 1 (CDK1), which mediates lamin phosphorylation. In support of a causal connection, pretreatment of tumor cells with a specific inhibitor of WEE1, a negative regulator kinase of CDK1, could counter the defective apoptosis of tumor cells expressing high levels of brachyury. Thus, our findings suggested that reconstituting CDK1 activity to threshold levels may be sufficient to restore immunosurveillance of mesenchymal-like cancer cells that have escaped previous immune detection or eradication. Cancer Res; 74(9); 2510–9. ©2014 AACR.
Data from Mechanistic Elucidation of the Antitumor Properties of Withaferin A in Breast Cancer
Authors: Nagalingam Arumugam , Kuppusamy Panjamurthy , Singh Shivendra V. , Sharma Dipali , Saxena Neeraj K.
Published: March 2024

Abstract
Withaferin A (WFA) is a steroidal lactone with antitumor effects manifested at multiple levels that are mechanistically obscure. Using a phospho-kinase screening array, we discovered that WFA activated phosphorylation of the S6 kinase RSK (ribosomal S6 kinase) in breast cancer cells. Pursuing this observation, we defined activation of extracellular signal–regulated kinase (ERK)–RSK and ETS-like transcription factor 1 (Elk1)–CHOP (C-EBP homologous protein) kinase pathways in upregulating transcription of the death receptor 5 (DR5). Through this route, WFA acted as an effective DR5 activator capable of potentiating the biologic effects of celecoxib, etoposide, and TRAIL. Accordingly, WFA treatment inhibited breast tumor formation in xenograft and mouse mammary tumor virus (MMTV)-neu mouse models in a manner associated with activation of the ERK/RSK axis, DR5 upregulation, and elevated nuclear accumulation of Elk1 and CHOP. Together, our results offer mechanistic insight into how WFA inhibits breast tumor growth. Cancer Res; 74(9); 2617–29. ©2014 AACR.
Data from Role of Chitinase 3–like-1 and Semaphorin 7a in Pulmonary Melanoma Metastasis
Authors: Ma Bing , Herzog Erica L. , Lee Chun Geun , Peng Xueyan , Lee Chang-Min , Chen Xiaosong , Rockwell Sara , Koo Ja Seok , Kluger Harriet , Herbst Roy S. , Sznol Mario , Elias Jack A.
Published: March 2024

Abstract
The prototypic chitinase-like protein Chi3l1 is induced in cancers and portends a poor prognosis, but whether it contributes to cancer progression is unknown. To address this gap in knowledge, we investigated the production of Chi3l1 in melanoma lung metastases. We found that Chi3l1 was induced during pulmonary melanoma metastasis and that this induction was regulated by the semaphorin Sema7a, interacting in stimulatory or inhibitory ways with its β1 integrin or Plexin C1 receptors, respectively. In mouse strains with genetic deletions of Chi3l1 or Sema7a, there was a significant reduction in pulmonary metastasis. Notably, antiserum raised against Chi3l1 or Sema7a phenocopied the reduction produced by genetic deletions. Melanoma lung metastasis was also decreased in the absence of IL13Rα2, a recently identified receptor for Chi3l1, consistent with a key role for Chi3l1 in melanoma spread. We confirmed roles for Sema7a and Chi3l1 in pulmonary metastasis of EMT6 breast cancer cells. Taken together, our studies establish a novel pathway through which Sem7a and its receptors regulate Chi3l1, revealing a host axis involving IL13Rα2 that plays a critical role in generating a pulmonary microenvironment that is critical to license metastasis. Cancer Res; 75(3); 487–96. ©2014 AACR.
Data from VEGF-Mediated Angiogenesis Links EMT-Induced Cancer Stemness to Tumor Initiation
Authors: Fantozzi Anna , Gruber Dorothea C. , Pisarsky Laura , Heck Chantal , Kunita Akiko , Yilmaz Mahmut , Meyer-Schaller Nathalie , Cornille Karen , Hopfer Ulrike , Bentires-Alj Mohamed , Christofori Gerhard
Published: March 2024

Abstract
An epithelial–mesenchymal transition (EMT) underlies malignant tumor progression and metastatic spread by enabling cancer cells to depart from the primary tumor, invade surrounding tissue, and disseminate to distant organs. EMT also enriches for cancer stem cells (CSC) and increases the capacity of cancer cells to initiate and propagate tumors upon transplantation into immune-deficient mice, a major hallmark of CSCs. However, the molecular mechanisms promoting the tumorigenicity of cancer cells undergoing an EMT and of CSCs have remained widely elusive. We here report that EMT confers efficient tumorigenicity to murine breast cancer cells by the upregulated expression of the proangiogenic factor VEGF-A and by increased tumor angiogenesis. On the basis of these data, we propose a novel interpretation of the features of CSCs with EMT-induced, VEGF-A–mediated angiogenesis as the connecting mechanism between cancer cell stemness and tumor initiation. Cancer Res; 74(5); 1566–75. ©2014 AACR.
Data from VEGF Regulates Region-Specific Localization of Perivascular Bone Marrow–Derived Cells in Glioblastoma
Authors: Burrell Kelly , Singh Sanjay , Jalali Shahrzad , Hill Richard P. , Zadeh Gelareh
Published: March 2024

Abstract
Glioblastoma multiforme (GBM) is characterized by a pathogenic vasculature that drives aggressive local invasion. Recent work suggests that GBM cells recruit bone marrow–derived progenitor cells (BMDC) to facilitate recurrence after radiotherapy, but how this may be achieved is unclear. In this study, we established the spatiotemporal and regional contributions of perivascular BMDCs (pBMDC) to GBM development. We found an increased recruitment of BMDCs to GBM in response to tumor growth and following radiotherapy. However, in this study, BMDCs did not differentiate into endothelial cells directly but rather provided a perivascular support role. The pBMDCs were shown to associate with tumor vasculature in a highly region-dependent manner, with central vasculature requiring minimal pBMDC support. Region-dependent association of pBMDC was regulated by VEGF. In the absence of VEGF, following radiotherapy or antiangiogenic therapy, we documented an increase in Ang2 that regulated recruitment of pBMDCs to maintain the vulnerable central vasculature. Together, our results strongly suggested that targeting pBMDC influx along with radiation or antiangiogenic therapy would be critical to prevent vascular recurrence of GBM. Cancer Res; 74(14); 3727–39. ©2014 AACR.
Data from MET Signaling in Colon Cancer Stem-like Cells Blunts the Therapeutic Response to EGFR Inhibitors
Authors: Luraghi Paolo , Reato Gigliola , Cipriano Elia , Sassi Francesco , Orzan Francesca , Bigatto Viola , De Bacco Francesca , Menietti Elena , Han May , Rideout William M. , Perera Timothy , Bertotti Andrea , Trusolino Livio , Comoglio Paolo M. , Boccaccio Carla
Published: March 2024

Abstract
Metastatic colorectal cancer remains largely incurable, although in a subset of patients, survival is prolonged by new targeting agents such as anti-EGF receptor (anti-EGFR) antibodies. This disease is believed to be supported by a subpopulation of stem-like cells termed colon cancer–initiating cell (CCIC), which may also confer therapeutic resistance. However, how CCICs respond to EGFR inhibition has not been fully characterized. To explore this question, we systematically generated CCICs through spheroid cultures of patient-derived xenografts of metastatic colorectal cancer. These cultures, termed “xenospheres,” were capable of long-term self-propagation in vitro and phenocopied the original patient tumors in vivo, thus operationally defining CCICs. Xenosphere CCICs retained the genetic determinants for EGFR therapeutic response in vitro and in xenografts; like the original tumors, xenospheres harboring a mutated KRAS gene were resistant to EGFR therapy, whereas those harboring wild-type RAS pathway genes (RASwt) were sensitive. Notably, the effects of EGFR inhibition in sensitive CCICs could be counteracted by cytokines secreted by cancer-associated fibroblasts. In particular, we found that the MET receptor ligand hepatocyte growth factor (HGF) was especially active in supporting in vitro CCIC proliferation and resistance to EGFR inhibition. Ectopic production of human HGF in CCIC xenografts rendered the xenografts susceptible to MET inhibition, which sensitized the response to EGFR therapy. By showing that RASwt CCICs rely on both EGFR and MET signaling, our results offer a strong preclinical proof-of-concept for concurrent targeting of these two pathways in the clinical setting. Cancer Res; 74(6); 1857–69. ©2014 AACR.
Data from Context-Selective Death of Acute Myeloid Leukemia Cells Triggered by the Novel Hybrid Retinoid-HDAC Inhibitor MC2392
Authors: De Bellis Floriana , Carafa Vincenzo , Conte Mariarosaria , Rotili Dante , Petraglia Francesca , Matarese Filomena , Françoijs Kees-Jan , Ablain Julien , Valente Sergio , Castellano Rèmy , Goubard Armelle , Collette Yves , Mandoli Amit , Martens Joost H.A. , de Thé Hugues , Nebbioso Angela , Mai Antonello , Stunnenberg Hendrik G. , Altucci Lucia
Published: March 2024

Abstract
HDAC inhibitors (HDACi) are widely used in the clinic to sensitize tumorigenic cells for treatment with other anticancer compounds. The major drawback of HDACi is the broad inhibition of the plethora of HDAC-containing complexes. In acute promyelocytic leukemia (APL), repression by the PML-RARα oncofusion protein is mediated by an HDAC-containing complex that can be dissociated by pharmacologic doses of all trans retinoic acid (ATRA) inducing differentiation and cell death at the expense of side effects and recurrence. We hypothesized that the context-specific close physical proximity of a retinoid and HDACi-binding protein in the repressive PML-RARα-HDAC complex may permit selective targeting by a hybrid molecule of ATRA with a 2-aminoanilide tail of the HDAC inhibitor MS-275, yielding MC2392. We show that MC2392 elicits weak ATRA and essentially no HDACi activity in vitro or in vivo. Genome-wide epigenetic analyses revealed that in NB4 cells expressing PML-RARα, MC2392 induces changes in H3 acetylation at a small subset of PML-RARα–binding sites. RNA-seq reveals that MC2392 alters expression of a number of stress-responsive and apoptotic genes. Concordantly, MC2392 induced rapid and massive, caspase-8–dependent cell death accompanied by RIP1 induction and ROS production. Solid and leukemic tumors are not affected by MC2392, but expression of PML-RARα conveys efficient MC2392-induced cell death. Our data suggest a model in which MC2392 binds to the RARα moiety and selectively inhibits the HDACs resident in the repressive complex responsible for the transcriptional impairment in APLs. Our findings provide proof-of-principle of the concept of a context-dependent targeted therapy. Cancer Res; 74(8); 2328–39. ©2014 AACR.
Data from Axon Guidance Factor SLIT2 Inhibits Neural Invasion and Metastasis in Pancreatic Cancer
Authors: Göhrig Andreas , Detjen Katharina M. , Hilfenhaus Georg , Körner Jan L. , Welzel Martina , Arsenic Ruza , Schmuck Rosa , Bahra Marcus , Wu Jane Y. , Wiedenmann Bertram , Fischer Christian
Published: March 2024

Abstract
Pancreatic ductal adenocarcinoma (PDAC) metastasizes by neural, vascular, and local invasion routes, which limit patient survival. In nerves and vessels, SLIT2 and its ROBO receptors constitute repellent guidance cues that also direct epithelial branching. Thus, the SLIT2–ROBO system may represent a key pinch point to regulate PDAC spread. In this study, we examined the hypothesis that escaping from repellent SLIT2–ROBO signaling is essential to enable PDAC cells to appropriate their local stromal infrastructure for dissemination. Through immunohistochemical analysis, we detected SLIT2 receptors ROBO1 and ROBO4 on epithelia, nerves, and vessels in healthy pancreas and PDAC specimens, respectively. SLIT2 mRNA expression was reduced in PDAC compared with nontransformed pancreatic tissues and cell lines, suggesting a reduction in SLIT2–ROBO pathway activity in PDAC. In support of this interpretation, restoring the SLIT2 expression in SLIT2-deficient PDAC cells inhibited their bidirectional chemoattraction with neural cells, and more specifically, impaired unidirectional PDAC cell navigation along outgrowing neurites in models of neural invasion. Restoring autocrine/paracrine SLIT2 signaling was also sufficient to inhibit the directed motility of PDAC cells, but not their random movement. Conversely, RNA interference–mediated silencing of ROBO1 stimulated the motility of SLIT2-competent PDAC cells. Furthermore, culture supernatants from SLIT2-competent PDAC cells impaired migration of endothelial cells (human umbilical vein endothelial cells), whereas an N-terminal SLIT2 cleavage fragment stimulated such migration. In vivo investigations of pancreatic tumors with restored SLIT2 expression demonstrated reduced invasion, metastasis, and vascularization, with opposing effects produced by ROBO1 silencing in tumor cells or sequestration of endogenous SLIT2. Analysis of clinical specimens of PDAC showed that those with low SLIT2 mRNA expression exhibited a higher incidence and a higher fraction of tumor-infiltrated lymph nodes. Taken together, our findings argue that disrupting SLIT2–ROBO signaling in PDAC may enhance metastasis and predispose PDAC cells to neural invasion. Cancer Res; 74(5); 1529–40. ©2014 AACR.
Data from USP9X Downregulation Renders Breast Cancer Cells Resistant to Tamoxifen
Authors: Oosterkamp Hendrika M. , Hijmans E. Marielle , Brummelkamp Thijn R. , Canisius Sander , Wessels Lodewyk F.A. , Zwart Wilbert , Bernards René
Published: March 2024

Abstract
Tamoxifen is one of the most widely used endocrine agents for the treatment of estrogen receptor α (ERα)–positive breast cancer. Although effective in most patients, resistance to tamoxifen is a clinically significant problem and the mechanisms responsible remain elusive. To address this problem, we performed a large scale loss-of-function genetic screen in ZR-75-1 luminal breast cancer cells to identify candidate resistance genes. In this manner, we found that loss of function in the deubiquitinase USP9X prevented proliferation arrest by tamoxifen, but not by the ER downregulator fulvestrant. RNAi-mediated attenuation of USP9X was sufficient to stabilize ERα on chromatin in the presence of tamoxifen, causing a global tamoxifen-driven activation of ERα-responsive genes. Using a gene signature defined by their differential expression after USP9X attenuation in the presence of tamoxifen, we were able to define patients with ERα-positive breast cancer experiencing a poor outcome after adjuvant treatment with tamoxifen. The signature was specific in its lack of correlation with survival in patients with breast cancer who did not receive endocrine therapy. Overall, our findings identify a gene signature as a candidate biomarker of response to tamoxifen in breast cancer. Cancer Res; 74(14); 3810–20. ©2014 AACR.
Data from HDAC2 Provides a Critical Support to Malignant Progression of Hepatocellular Carcinoma through Feedback Control of mTORC1 and AKT
Authors: Noh Ji Heon , Bae Hyun Jin , Eun Jung Woo , Shen Qingyu , Park Se Jin , Kim Hyung Seok , Nam Boas , Shin Woo Chan , Lee Eun Kyung , Lee Kyungbun , Jang Ja-Jun , Park Won Sang , Lee Jung Young , Nam Suk Woo
Published: March 2024

Abstract
Aberrant regulation of histone deacetylase 2 (HDAC2) contributes to malignant progression in various cancers, but the underlying mechanism leading to the activation of oncogenic HDAC2 remains unknown. In this study, we show that HDAC2 expression is upregulated in a large cohort of patients with human hepatocellular carcinoma, and that high expression of HDAC2 was significantly associated with poor prognosis of patients with hepatocellular carcinoma. We found that mTORC1/NF-κBp50 signaling is necessary for the growth factor–induced HDAC2 and is sustained in hepatocellular carcinoma, but not in normal hepatic cells. Growth factor–induced mTORC1 activates the nuclear translocation of NF-κBp50, where it binds to the intragenic sequences of the HDAC2 gene and promotes its transcription. Hepatocellular carcinoma tissues derived from chemical-induced mouse and rat liver cancer models validated that mTORC1 activation and NF-κBp50 nuclear translocation are essential for the transcriptional activation of oncogenic HDAC2 in hepatocellular carcinoma. In addition, we demonstrate that HDAC2 is required to maintain mTORC1 activity by stabilizing the mTOR/RAPTOR complex. Elevated expression of HDAC2 triggers a positive feedback loop that activates AKT phosphorylation via the transcriptional modulation of phosphoinositide signaling molecules. Bioinformatics analysis of HDAC2 signature and immunoblot analysis of mesenchymal genes also evidenced that HDAC2 plays a role in the malignant behavior of tumor cells by Snail induction and simultaneously E-cadherin suppression in hepatocellular carcinoma cells. These findings establish a molecular mechanism responsible for the activation of oncogenic HDAC2, which explains how growth factor–induced HDAC2 maintains mitogenic signaling and function during hepatocellular malignant progression and provide a novel strategy for therapeutic intervention in liver cancer. Cancer Res; 74(6); 1728–38. ©2014 AACR.
Data from Src Kinase Is a Novel Therapeutic Target in Lymphangioleiomyomatosis
Authors: Tyryshkin Alexey , Bhattacharya Abhisek , Eissa N. Tony
Published: March 2024

Abstract
Lymphangioleiomyomatosis (LAM) is a progressive cystic lung disease affecting some women with tuberous sclerosis complex (TSC). Sporadic LAM can develop in women without TSC, owing to somatic mutations in the TSC2 gene. Accumulating evidence supports the view of LAM as a low-grade, destructive, metastasizing neoplasm. The mechanisms underlying the metastatic capability of LAM cells remain poorly understood. The observed behavior of LAM cells with respect to their infiltrative growth pattern, metastatic potential, and altered cell differentiation bears similarity to cells undergoing epithelial–mesenchymal transition. Here, we report increased levels of active Src kinase in LAM lungs and in TSC2−/− cells, caused by a reduction of autophagy. Furthermore, increased Src kinase activation promoted migration, invasion, and inhibition of E-cadherin expression in TSC2−/− cells by upregulating the transcription factor Snail. Notably, Src kinase inhibitors reduced migration and invasion properties of TSC2−/− cells and attenuated lung colonization of intravenously injected TSC2−/− cells in vivo to a greater extent than control TSC2+/+ cells. Our results reveal mechanistic basis for the pathogenicity of LAM cells and they rationalize Src kinase as a novel therapeutic target for treatment of LAM and TSC. Cancer Res; 74(7); 1996–2005. ©2014 AACR.
Data from BRCA2 Phosphorylated by PLK1 Moves to the Midbody to Regulate Cytokinesis Mediated by Nonmuscle Myosin IIC
Authors: Takaoka Miho , Saito Hiroko , Takenaka Katsuya , Miki Yoshio , Nakanishi Akira
Published: March 2024

Abstract
Cytokinesis is the critical final step in cell division. BRCA2 disruption during cytokinesis is associated with chromosome instability, but mechanistic information is lacking that could be used to prevent cancer cell division. In this study, we report that BRCA2 phosphorylation by the mitotic polo-like kinase (PLK1) governs the localization of BRCA2 to the Flemming body at the central midbody, permitting an interaction with nonmuscle myosin IIC (NM-IIC). Formation of an NM-IIC ring-like structure at the Flemming body shows that the IIC-ring relies on its ATPase activity stimulated by interaction with BRCA2 and associated proteins. Notably, inhibiting this binding inactivated the ATPase activity, causing disassembly of the IIC-ring, defective formation of the midbody, and interruption of cytokinesis. An analysis of cancer-associated mutations in BRCA2 at the PLK1-binding site suggests that they may contribute to cytokinetic defects by altering BRCA2 localization. Our findings suggest that BRCA2-dependent IIC-ring formation is a critical step in proper formation of the midbody, offering an explanation for how chromosome instability may arise in breast cancer. Cancer Res; 74(5); 1518–28. ©2014 AACR.
Data from YAP-Induced Resistance of Cancer Cells to Antitubulin Drugs Is Modulated by a Hippo-Independent Pathway
Authors: Zhao Yulei , Khanal Prem , Savage Paul , She Yi-Min , Cyr Terry D. , Yang Xiaolong
Published: March 2024

Abstract
Although antitubulin drugs are used widely to treat human cancer, many patients display intrinsic or acquired drug resistance that imposes major obstacles to successful therapy. Mounting evidence argues that cancer cell apoptosis triggered by antitubulin drugs relies upon activation of the cell-cycle kinase Cdk1; however, mechanistic connections of this event to apoptosis remain obscure. In this study, we identified the antiapoptotic protein YAP, a core component of the Hippo signaling pathway implicated in tumorigenesis, as a critical linker coupling Cdk1 activation to apoptosis in the antitubulin drug response. Antitubulin drugs activated Cdk1, which directly phosphorylated YAP on five sites independent of the Hippo pathway. Mutations in these phosphorylation sites on YAP relieved its ability to block antitubulin drug-induced apoptosis, further suggesting that YAP was inactivated by Cdk1 phosphorylation. Notably, we found that YAP was not phosphorylated and inactivated after antitubulin drug treatment in taxol-resistant cancer cells. Our findings suggest YAP and its phosphorylation status as candidate prognostic markers in predicting antitubulin drug response in patients. Cancer Res; 74(16); 4493–503. ©2014 AACR.
Data from NUP98 Fusion Oncoproteins Promote Aneuploidy by Attenuating the Mitotic Spindle Checkpoint
Authors: Salsi Valentina , Ferrari Silvia , Gorello Paolo , Fantini Sebastian , Chiavolelli Francesca , Mecucci Cristina , Zappavigna Vincenzo
Published: March 2024

Abstract
NUP98 is a recurrent fusion partner in chromosome translocations that cause acute myelogenous leukemia. NUP98, a nucleoporin, and its interaction partner Rae1, have been implicated in the control of chromosome segregation, but their mechanistic contributions to tumorigenesis have been unclear. Here, we show that expression of NUP98 fusion oncoproteins causes mitotic spindle defects and chromosome missegregation, correlating with the capability of NUP98 fusions to cause premature securin degradation and slippage from an unsatisfied spindle assembly checkpoint (SAC). NUP98 fusions, unlike wild-type NUP98, were found to physically interact with the anaphase promoting complex/cyclosome (APC/C)Cdc20 and to displace the BubR1 SAC component, suggesting a possible mechanistic basis for their interference with SAC function. In addition, NUP98 oncoproteins displayed a prolonged half-life in cells. We found that NUP98 stability is controlled by a PEST sequence, absent in NUP98 oncoproteins, whose deletion reproduced the aberrant SAC-interfering activity of NUP98 oncoproteins. Together, our findings suggest that NUP98 oncoproteins predispose myeloid cells to oncogenic transformation or malignant progression by promoting whole chromosome instability. Cancer Res; 74(4); 1079–90. ©2013 AACR.
Data from A Mechanism of Hypoxia-Mediated Escape from Adaptive Immunity in Cancer Cells
Authors: Barsoum Ivraym B. , Smallwood Chelsea A. , Siemens D. Robert , Graham Charles H.
Published: March 2024

Abstract
Immune escape is a fundamental trait of cancer in which mechanistic knowledge is incomplete. Here, we describe a novel mechanism by which hypoxia contributes to tumoral immune escape from cytotoxic T lymphocytes (CTL). Exposure of human or murine cancer cells to hypoxia for 24 hours led to upregulation of the immune inhibitory molecule programmed cell death ligand-1 (PD-L1; also known as B7-H1), in a manner dependent on the transcription factor hypoxia-inducible factor-1α (HIF-1α). In vivo studies also demonstrated cellular colocalization of HIF-1α and PD-L1 in tumors. Hypoxia-induced expression of PD-L1 in cancer cells increased their resistance to CTL-mediated lysis. Using glyceryl trinitrate (GTN), an agonist of nitric oxide (NO) signaling known to block HIF-1α accumulation in hypoxic cells, we prevented hypoxia-induced PD-L1 expression and diminished resistance to CTL-mediated lysis. Moreover, transdermal administration of GTN attenuated tumor growth in mice. We found that higher expression of PD-L1 induced in tumor cells by exposure to hypoxia led to increased apoptosis of cocultured CTLs and Jurkat leukemia T cells. This increase in apoptosis was prevented by blocking the interaction of PD-L1 with PD-1, the PD-L1 receptor on T cells, or by addition of GTN. Our findings point to a role for hypoxia/HIF-1 in driving immune escape from CTL, and they suggest a novel cancer immunotherapy to block PD-L1 expression in hypoxic-tumor cells by administering NO mimetics. Cancer Res; 74(3); 665–74. ©2013 AACR.
Data from β-Catenin Inhibitor ICAT Modulates the Invasive Motility of Melanoma Cells
Authors: Domingues Mélanie J. , Rambow Florian , Job Bastien , Papon Laura , Liu Wanguo , Larue Lionel , Bonaventure Jacky
Published: March 2024

Abstract
Inhibitor of β-catenin and TCF (ICAT) inhibits β-catenin transcriptional activity by competing with T-cell factor/lymphoid enhancer factor. We documented high ICAT levels in human melanoma cells, in which β-catenin signaling is frequently deregulated, finding a correlation with the capacity to form metastases in nude mice. Ectopic expression of ICAT in melanoma cells did not affect their proliferation but increased cell motility and Matrigel invasion of metastatic cells in a manner relying upon stable ICAT–β-catenin interaction. This effect was associated with conversion of an elongated/mesenchymal phenotype to a round/amoeboid phenotype in the absence of similar effects on elongated morphology of nonmetastatic melanoma cells. Transition from mesenchymal to amoeboid movement was associated with decreased levels of NEDD9 and activated Rac1, a positive regulator of mesenchymal movement. Ectopic ICAT promoted colonization of melanoma cells in the lungs of nude mice, suggesting an increase in metastatic potential. Together, our results showed that by downregulating Rac signaling in metastatic melanoma cells, ICAT increased their invasive motility by promoting a morphologic variation that facilitates a favorable adaptation to their microenvironment. Cancer Res; 74(7); 1983–95. ©2014 AACR.
Data from Activation of the Glutamate Receptor GRM1 Enhances Angiogenic Signaling to Drive Melanoma Progression
Authors: Wen Yu , Li Jiadong , Koo Jasmine , Shin Seung-Shick , Lin Yong , Jeong Byeong-Seon , Mehnert Janice M. , Chen Suzie , Cohen-Sola Karine A. , Goydos James S.
Published: March 2024

Abstract
Glutamate-triggered signal transduction is thought to contribute widely to cancer pathogenesis. In melanoma, overexpression of the metabotropic glutamate receptor (GRM)-1 occurs frequently and its ectopic expression in melanocytes is sufficient for neoplastic transformation. Clinical evaluation of the GRM1 signaling inhibitor riluzole in patients with advanced melanoma has demonstrated tumor regressions that are associated with a suppression of the mitogen-activated protein kinase (MAPK) and phosphoinositide 3-kinase/protein kinase B (PI3K/AKT) pathways. Together, these results prompted us to investigate the downstream consequences of GRM1 signaling and its disruption in more detail. We found that melanoma cells with enhanced GRM1 expression generated larger tumors in vivo marked by more abundant blood vessels. Media conditioned by these cells in vitro contained relatively higher concentrations of interleukin-8 and VEGF due to GRM1-mediated activation of the AKT–mTOR–HIF1 pathway. In clinical specimens from patients receiving riluzole, we confirmed an inhibition of MAPK and PI3K/AKT activation in posttreatment as compared with pretreatment tumor specimens, which exhibited a decreased density of blood vessels. Together, our results demonstrate that GRM1 activation triggers proangiogenic signaling in melanoma, offering a mechanistic rationale to design treatment strategies for the most suitable combinatorial use of GRM1 inhibitors in patients. Cancer Res; 74(9); 2499–509. ©2014 AACR.
Data from Efficacy of CAR T-cell Therapy in Large Tumors Relies upon Stromal Targeting by IFNγ
Authors: Textor Ana , Listopad Joanna J. , Wührmann Lara Le , Perez Cynthia , Kruschinski Anna , Chmielewski Markus , Abken Hinrich , Blankenstein Thomas , Charo Jehad
Published: March 2024

Abstract
Adoptive T-cell therapy using chimeric antigen receptor–modified T cells (CAR-T therapy) has shown dramatic efficacy in patients with circulating lymphoma. However, eradication of solid tumors with CAR-T therapy has not been reported yet to be efficacious. In solid tumors, stroma destruction, due to MHC-restricted cross-presentation of tumor antigens to T cells, may be essential. However, CAR-Ts recognize antigens in an MHC-independent manner on cancer cells but not stroma cells. In this report, we show how CAR-Ts can be engineered to eradicate large established tumors with provision of a suitable CD28 costimulatory signal. In an HER2–dependent tumor model, tumor rejection by HER2–specific CAR-Ts was associated with sustained influx and proliferation of the adoptively transferred T cells. Interestingly, tumor rejection did not involve natural killer cells but was associated instead with a marked increase in the level of M1 macrophages and a requirement for IFNγ receptor expression on tumor stroma cells. Our results argue that CAR-T therapy is capable of eradicating solid tumors through a combination of antigen-independent stroma destruction and antigen-specific tumor cell targeting. Cancer Res; 74(23); 6796–805. ©2014 AACR.
Data from Hypoxia Promotes Nuclear Translocation and Transcriptional Function in the Oncogenic Tyrosine Kinase RON
Authors: Chang Hong-Yi , Liu Hsiao-Sheng , Lai Ming-Derg , Tsai Yuh-Shyan , Tzai Tzong-Shin , Cheng Hong-Ling , Chow Nan-Haw
Published: March 2024

Abstract
Tumor hypoxia drives metastatic progression, drug resistance, and posttreatment relapses, but how cancer cells adapt and evolve in response to hypoxic stress is not well understood. In this study, we address this question with the discovery that the receptor tyrosine kinase RON translocates into the nucleus of hypoxic cancer cells. In response to hypoxia, nuclear RON interacts with the hypoxia-inducible factor HIF-1α in a manner that relies on RON tyrosine kinase activity, binding to the c-JUN promoter and activating it. Mechanistic investigations revealed unexpectedly that nuclear RON played a more important role in activation of the c-JUN promoter than HIF-1α, leading to increased cell proliferation, survival adaptation, in vitro migration, and tumorigenicity under hypoxic conditions. Taken together, our results pointed to a novel function for RON as a transcriptional regulator that promotes the survival of cancer cells subjected to hypoxia. These results suggest novel implications for the use of small-molecule inhibitors or monoclonal antibodies targeting the RON kinase in the prevention or treatment of advanced cancer. Cancer Res; 74(16); 4549–62. ©2014 AACR.
Data from Natural Killer Cells Are Essential for the Ability of BRAF Inhibitors to Control BRAFV600E-Mutant Metastatic Melanoma
Authors: Ferrari de Andrade Lucas , Ngiow Shin F. , Stannard Kimberley , Rusakiewicz Sylvie , Kalimutho Murugan , Khanna Kum Kum , Tey Siok-Keen , Takeda Kazuyoshi , Zitvogel Laurence , Martinet Ludovic , Smyth Mark J.
Published: March 2024

Abstract
BRAFV600E is a major oncogenic mutation found in approximately 50% of human melanoma that confers constitutive activation of the MAPK pathway and increased melanoma growth. Inhibition of BRAFV600E by oncogene targeting therapy increases overall survival of patients with melanoma, but is unable to produce many durable responses. Adaptive drug resistance remains the main limitation to BRAFV600E inhibitor clinical efficacy and immune-based strategies could be useful to overcome disease relapse. Tumor microenvironment greatly differs between visceral metastasis and primary cutaneous melanoma, and the mechanisms involved in the antimetastatic efficacy of BRAFV600E inhibitors remain to be determined. To address this question, we developed a metastatic BRAFV600E-mutant melanoma cell line and demonstrated that the antimetastatic properties of BRAF inhibitor PLX4720 (a research analogue of vemurafenib) require host natural killer (NK) cells and perforin. Indeed, PLX4720 not only directly limited BRAFV600E-induced tumor cell proliferation, but also affected NK cell functions. We showed that PLX4720 increases the phosphorylation of ERK1/2, CD69 expression, and proliferation of mouse NK cells in vitro. NK cell frequencies were significantly enhanced by PLX4720 specifically in the lungs of mice with BRAFV600E lung metastases. Furthermore, PLX4720 also increased human NK cell pERK1/2, CD69 expression, and IFNγ release in the context of anti-NKp30 and IL2 stimulation. Overall, this study supports the idea that additional NK cell-based immunotherapy (by checkpoint blockade or agonists or cytokines) may combine well with BRAFV600E inhibitor therapy to promote more durable responses in melanoma. Cancer Res; 74(24); 7298–308. ©2014 AACR.
Data from Targeting Cancer Stem–like Cells as an Approach to Defeating Cellular Heterogeneity in Ewing Sarcoma
Authors: Cornaz-Buros Sandrine , Riggi Nicolo , DeVito Claudio , Sarre Alexandre , Letovanec Igor , Provero Paolo , Stamenkovic Ivan
Published: March 2024

Abstract
Plasticity in cancer stem–like cells (CSC) may provide a key basis for cancer heterogeneity and therapeutic response. In this study, we assessed the effect of combining a drug that abrogates CSC properties with standard-of-care therapy in a Ewing sarcoma family tumor (ESFT). Emergence of CSC in this setting has been shown to arise from a defect in TARBP2-dependent microRNA maturation, which can be corrected by exposure to the fluoroquinolone enoxacin. In the present work, primary ESFT from four patients containing CD133+ CSC subpopulations ranging from 3% to 17% of total tumor cells were subjected to treatment with enoxacin, doxorubicin, or both drugs. Primary ESFT CSC and bulk tumor cells displayed divergent responses to standard-of-care chemotherapy and enoxacin. Doxorubicin, which targets the tumor bulk, displayed toxicity toward primary adherent ESFT cells in culture but not to CSC-enriched ESFT spheres. Conversely, enoxacin, which enhances miRNA maturation by stimulating TARBP2 function, induced apoptosis but only in ESFT spheres. In combination, the two drugs markedly depleted CSCs and strongly reduced primary ESFTs in xenograft assays. Our results identify a potentially attractive therapeutic strategy for ESFT that combines mechanism-based targeting of CSC using a low-toxicity antibiotic with a standard-of-care cytotoxic drug, offering immediate applications for clinical evaluation. Cancer Res; 74(22); 6610–22. ©2014 AACR.
Data from Changes in Pyruvate Metabolism Detected by Magnetic Resonance Imaging Are Linked to DNA Damage and Serve as a Sensor of Temozolomide Response in Glioblastoma Cells
Authors: Park Ilwoo , Mukherjee Joydeep , Ito Motokazu , Chaumeil Myriam M. , Jalbert Llewellyn E. , Gaensler Karin , Ronen Sabrina M. , Nelson Sarah J. , Pieper Russell O.
Published: March 2024

Abstract
Recent findings show that exposure to temozolomide (TMZ), a DNA-damaging drug used to treat glioblastoma (GBM), can suppress the conversion of pyruvate to lactate. To understand the mechanistic basis for this effect and its potential utility as a TMZ response biomarker, we compared the response of isogenic GBM cell populations differing only in expression of the DNA repair protein methyltransferase (MGMT), a TMZ-sensitivity determinant, after exposure to TMZ in vitro and in vivo. Hyperpolarized [1-(13)C]-pyruvate–based MRI was used to monitor temporal effects on pyruvate metabolism in parallel with DNA-damage responses and tumor cell growth. TMZ exposure decreased conversion of pyruvate to lactate only in MGMT-deficient cells. This effect coincided temporally with TMZ-induced increases in levels of the DNA-damage response protein pChk1. Changes in pyruvate to lactate conversion triggered by TMZ preceded tumor growth suppression and were not associated with changes in levels of NADH or lactate dehydrogenase activity in tumors. Instead, they were associated with a TMZ-induced decrease in the expression and activity of pyruvate kinase PKM2, a glycolytic enzyme that indirectly controls pyruvate metabolism. PKM2 silencing decreased PK activity, intracellular lactate levels, and conversion of pyruvate to lactate in the same manner as TMZ, and Chk1 silencing blocked the TMZ-induced decrease in PKM2 expression. Overall, our findings showed how TMZ-induced DNA damage is linked through PKM2 to changes in pyruvate metabolism, and how these changes can be exploited by MRI methods as an early sensor of TMZ therapeutic response. Cancer Res; 74(23); 7115–24. ©2014 AACR.
Data from RB Family Tumor Suppressor Activity May Not Relate to Active Silencing of E2F Target Genes
Authors: Vormer Tinke L. , Wojciechowicz Kamila , Dekker Marleen , de Vries Sandra , van der Wal Anja , Delzenne-Goette Elly , Naik Sjalin H. , Song Ji-Ying , Dannenberg Jan-Hermen , Hansen Jacob B. , te Riele Hein
Published: March 2024

Abstract
The retinoblastoma protein pRB and its two homologs p130 and p107 form the family of pocket proteins and play a major role in cell-cycle regulation and suppression of human and mouse tumorigenesis. Pocket proteins regulate the activity of E2F transcription factors during G1–S transition. Two mechanisms have been described: (i) pocket protein binding blocks the transactivation domain of activator E2Fs, inhibiting E2F-dependent transcription and (ii) E2F-bound pocket proteins can recruit chromatin remodeling proteins containing an LxCxE motif (x encoding any amino acid), resulting in active repression of E2F target genes. To investigate the importance of pRB's LxCxE-interacting motif in cell-cycle control and tumor suppression, we generated mouse embryonic fibroblasts and mice expressing a mutant pRB protein carrying an asparagine for phenylalanine substitution at position 750, abrogating LxCxE binding. Because p130 may compensate for loss of pRB, we studied pRBN750F activity in the presence and absence of p130. The pRB–LxCxE interaction was not required for cell-cycle arrest upon mitogen deprivation and cell-cell contact, but did contribute to RASV12- and radiation-induced cell-cycle arrest. Remarkably, the pRB–LxCxE interaction was not required for suppression of in vitro and in vivo transformation, even in the absence of p130. These results indicate that pRB's tumor suppressor activity is not effectuated by active silencing of E2F target genes, but rather by regulation of activator E2Fs or another unidentified mechanism. Furthermore, the in vitro response of pocket protein–perturbed cells to mitogen deprivation and cell–cell contact seems a better predictor of tumor development than the response to ectopic RASV12 expression. Cancer Res; 74(18); 5266–76. ©2014 AACR.
Data from BRCA1 Suppresses Epithelial-to-Mesenchymal Transition and Stem Cell Dedifferentiation during Mammary and Tumor Development
Authors: Bai Feng , Chan Ho Lam , Scott Alexandria , Smith Matthew D. , Fan Cheng , Herschkowitz Jason I. , Perou Charles M. , Livingstone Alan S. , Robbins David J. , Capobianco Anthony J. , Pei Xin-Hai
Published: March 2024

Abstract
BRCA1 mutation carriers are predisposed to developing basal-like breast cancers with high metastasis and poor prognosis. Yet, how BRCA1 suppresses formation of basal-like breast cancers is still obscure. Deletion of p18Ink4c (p18), an inhibitor of CDK4 and CDK6, functionally inactivates the RB pathway, stimulates mammary luminal stem cell (LSC) proliferation, and leads to spontaneous luminal tumor development. Alternately, germline mutation of Brca1 shifts the fate of luminal cells to cause luminal-to-basal mammary tumor transformation. Here, we report that disrupting Brca1 by either germline or epithelium-specific mutation in p18-deficient mice activates epithelial-to-mesenchymal transition (EMT) and induces dedifferentiation of LSCs, which associate closely with expansion of basal and cancer stem cells and formation of basal-like tumors. Mechanistically, BRCA1 bound to the TWIST promoter, suppressing its activity and inhibiting EMT in mammary tumor cells. In human luminal cancer cells, BRCA1 silencing was sufficient to activate TWIST and EMT and increase tumor formation. In parallel, TWIST expression and EMT features correlated inversely with BRCA1 expression in human breast cancers. Together, our findings showed that BRCA1 suppressed TWIST and EMT, inhibited LSC dedifferentiation, and repressed expansion of basal stem cells and basal-like tumors. Thus, our work offers the first genetic evidence that Brca1 directly suppresses EMT and LSC dedifferentiation during breast tumorigenesis. Cancer Res; 74(21); 6161–72. ©2014 AACR.
Data from Favorable Prognostic Impact in Loss of TP53 and PIK3CA Mutations after Neoadjuvant Chemotherapy in Breast Cancer
Authors: Jiang Yi-Zhou , Yu Ke-Da , Bao Jing , Peng Wen-Ting , Shao Zhi-Ming
Published: March 2024

Abstract
We investigated the loss of somatic mutations in TP53 and PIK3CA in breast cancer tissue after neoadjuvant chemotherapy (NCT) and the clinical relevance of the observed mutation profiles. Samples were derived from three cohorts: Cohort 1 consisting of 206 patients undergoing NCT with matched pre- and postchemotherapy tumor tissues; Cohort 2 consisting of 158 additional patients undergoing NCT; and Cohort 3, consisting of 81 patients undergoing chemotherapy with prechemotherapy tumor tissues. In the first cohort, somatic mutations in TP53 or PIK3CA were identified in 24.8% of the pre-NCT tumor samples but in only 12.1% of the post-NCT tumor samples (P < 0.001). Patients with initial TP53 and PIK3CA mutations who became negative for the mutations after NCT had a higher Miller–Payne score (P = 0.008), improved disease-free survival, and improved overall survival than those with no change or the opposite change. The association of loss of mutations in TP53 and PIK3CA and improved survival was successfully validated in the second cohort. In addition, 28.4% of the tumors showed intratumoral heterogeneity of somatic mutations in TP53 or PIK3CA, whereas 71.6% were homogeneous, either with or without the mutations. Our data reveal the novel concept that chemotherapy may reduce mutation frequency in patients with breast cancer. Furthermore, the loss of somatic mutations in TP53 and PIK3CA may be translated to biomarkers for prognosis via further verification, which may optimize the choice of sequential therapy and improve patient survival. Cancer Res; 74(13); 3399–407. ©2014 AACR.
Data from In Vivo Disruption of an Rb–E2F–Ezh2 Signaling Loop Causes Bladder Cancer
Authors: Santos Mirentxu , Martínez-Fernández Mónica , Dueñas Marta , García-Escudero Ramón , Alfaya Begoña , Villacampa Felipe , Saiz-Ladera Cristina , Costa Clotilde , Oteo Marta , Duarte José , Martínez Victor , Gómez-Rodriguez Ma José , Martín Ma Luisa , Fernández Manoli , Viatour Patrick , Morcillo Miguel A. , Sage Julien , Castellano Daniel , Rodriguez-Peralto Jose L. , de la Rosa Federico , Paramio Jesús M
Published: March 2024

Abstract
Bladder cancer is a highly prevalent human disease in which retinoblastoma (Rb) pathway inactivation and epigenetic alterations are common events. However, the connection between these two processes is still poorly understood. Here, we show that the in vivo inactivation of all Rb family genes in the mouse urothelium is sufficient to initiate bladder cancer development. The characterization of the mouse tumors revealed multiple molecular features of human bladder cancer, including the activation of E2F transcription factor and subsequent Ezh2 expression and the activation of several signaling pathways previously identified as highly relevant in urothelial tumors. These mice represent a genetically defined model for human high-grade superficial bladder cancer. Whole transcriptional characterizations of mouse and human bladder tumors revealed a significant overlap and confirmed the predominant role for Ezh2 in the downregulation of gene expression programs. Importantly, the increased tumor recurrence and progression in human patients with superficial bladder cancer is associated with increased E2F and Ezh2 expression and Ezh2-mediated gene expression repression. Collectively, our studies provide a genetically defined model for human high-grade superficial bladder cancer and demonstrate the existence of an Rb–E2F–Ezh2 axis in bladder whose disruption can promote tumor development. Cancer Res; 74(22); 6565–77. ©2014 AACR.
Data from Myeloid IKKβ Promotes Antitumor Immunity by Modulating CCL11 and the Innate Immune Response
Authors: Yang Jinming , Hawkins Oriana E. , Barham Whitney , Gilchuk Pavlo , Boothby Mark , Ayers Gregory D. , Joyce Sebastian , Karin Michael , Yull Fiona E. , Richmond Ann
Published: March 2024

Abstract
Myeloid cells are capable of promoting or eradicating tumor cells and the nodal functions that contribute to their different roles are still obscure. Here, we show that mice with myeloid-specific genetic loss of the NF-κB pathway regulatory kinase IKKβ exhibit more rapid growth of cutaneous and lung melanoma tumors. In a BRAFV600E/PTEN−/− allograft model, IKKβ loss in macrophages reduced recruitment of myeloid cells into the tumor, lowered expression of MHC class II molecules, and enhanced production of the chemokine CCL11, thereby negatively regulating dendritic-cell maturation. Elevated serum and tissue levels of CCL11 mediated suppression of dendritic-cell differentiation/maturation within the tumor microenvironment, skewing it toward a Th2 immune response and impairing CD8+ T cell–mediated tumor cell lysis. Depleting macrophages or CD8+ T cells in mice with wild-type IKKβ myeloid cells enhanced tumor growth, where the myeloid cell response was used to mediate antitumor immunity against melanoma tumors (with less dependency on a CD8+ T-cell response). In contrast, myeloid cells deficient in IKKβ were compromised in tumor cell lysis, based on their reduced ability to phagocytize and digest tumor cells. Thus, mice with continuous IKKβ signaling in myeloid-lineage cells (IKKβCA) exhibited enhanced antitumor immunity and reduced melanoma outgrowth. Collectively, our results illuminate new mechanisms through which NF-κB signaling in myeloid cells promotes innate tumor surveillance. Cancer Res; 74(24); 7274–84. ©2014 AACR.