bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from Nitric Oxide Inhibits the Proliferation and Invasion of Pancreatic Cancer Cells through Degradation of Insulin Receptor Substrate-1 Protein
Authors: Sugita Hiroki , Kaneki Masao , Furuhashi Satoshi , Hirota Masahiko , Takamori Hiroshi , Baba Hideo
Published: April 2023

Abstract
Nitric oxide (NO), which plays a role in the posttranslational modification of proteins, exhibits tumoricidal activity. However, the mechanism remains largely unclear. We investigated whether the regulation of insulin receptor substrate (IRS)-1 protein expression and insulin/insulin-like growth factor (IGF) signaling by NO is involved in the proliferation and invasion of pancreatic cancer cells. NO donor inhibited insulin/IGF-I–stimulated phosphorylation of insulin receptor/IGF-I receptor, IRS-1, Akt/PKB, and glycogen synthase kinase-3β along with decreased expression of IRS-1 protein in MIAPaCa-2 cells, whereas NO donor enhanced the phosphorylation of extracellular signal-regulated kinase-1/2. In contrast, a selective inducible nitric oxide synthase inhibitor, 1400W, upregulated the expression of IRS-1 protein and the phosphorylation of IRS-1, Akt/PKB, and glycogen synthase kinase-3β, along with enhanced proliferation and invasion of Panc-1 cells expressing inducible nitric oxide synthase protein. NO donor induced IRS-1 protein reduction through increased ubiquitination and degradation. For the detection of the site responsible for NO-induced ubiquitination, IRS-1 deletion mutant genes were transfected and overexpressed in MIAPaCa-2 cells. The results indicate that the COOH terminus of the IRS-1 protein is required for NO donor–induced ubiquitination and protein degradation. Cells stably transfected with COOH-terminal deletion mutants of IRS-1 exhibited reduced IGF signaling and cell proliferation compared with vector alone–transfected cells, with no influence of NO on IGF signaling and invasion, although stable transfectants with full-length IRS-1 protein exhibited remarkable NO-induced reduction in IGF signaling, cell proliferation, and invasion. These findings indicate that NO inhibits the proliferation and invasion of pancreatic cancer cells, at least in part, through upregulation of IRS-1 protein degradation and resultant downregulation of the insulin/IGF-I-Akt pathway. Mol Cancer Res; 8(8); 1152–63. ©2010 AACR.
Data from Mutations in the Circadian Gene CLOCK in Colorectal Cancer
Authors: Alhopuro Pia , Björklund Mikael , Sammalkorpi Heli , Turunen Mikko , Tuupanen Sari , Biström Mia , Niittymäki Iina , Lehtonen Heli J. , Kivioja Teemu , Launonen Virpi , Saharinen Juha , Nousiainen Kari , Hautaniemi Sampsa , Nuorva Kyösti , Mecklin Jukka-Pekka , Järvinen Heikki , Orntoft Torben , Arango Diego , Lehtonen Rainer , Karhu Auli , Taipale Jussi , Aaltonen Lauri A.
Published: April 2023

Abstract
The circadian clock regulates daily variations in physiologic processes. CLOCK acts as a regulator in the circadian apparatus controlling the expression of other clock genes, including PER1. Clock genes have been implicated in cancer-related functions; in this work, we investigated CLOCK as a possible target of somatic mutations in microsatellite unstable colorectal cancers. Combining microarray gene expression data and public gene sequence information, we identified CLOCK as 1 of 790 putative novel microsatellite instability (MSI) target genes. A total of 101 MSI colorectal carcinomas (CRC) were sequenced for a coding microsatellite in CLOCK. The effect of restoring CLOCK expression was studied in LS180 cells lacking wild-type CLOCK by stably expressing GST-CLOCK or glutathione S-transferase empty vector and testing the effects of UV-induced apoptosis and radiation by DNA content analysis using flow cytometry. Putative novel CLOCK target genes were searched by using ChIP-seq. CLOCK mutations occurred in 53% of MSI CRCs. Restoring CLOCK expression in cells with biallelic CLOCK inactivation resulted in protection against UV-induced apoptosis and decreased G2-M arrest in response to ionizing radiation. Using ChIP-Seq, novel CLOCK-binding elements were identified near DNA damage genes p21, NBR1, BRCA1, and RAD50. CLOCK is shown to be mutated in cancer, and altered response to DNA damage provides one plausible mechanism of tumorigenesis. Mol Cancer Res; 8(7); 952–60. ©2010 AACR.
Data from Bortezomib-Resistant Nuclear Factor-κB Activity in Multiple Myeloma Cells
Authors: Markovina Stephanie , Callander Natalie S. , O'Connor Shelby L. , Kim Jihoon , Werndli Jae E. , Raschko Martha , Leith Catherine P. , Kahl Brad S. , Kim KyungMann , Miyamoto Shigeki
Published: April 2023

Abstract
Bortezomib (Velcade/PS341), a proteasome inhibitor used in the treatment of multiple myeloma (MM), can inhibit activation of nuclear factor-κB (NF-κB), a family of transcription factors often deregulated and constitutively activated in primary MM cells. NF-κB can be activated via several distinct mechanisms, including the proteasome inhibitor–resistant (PIR) pathway. It remains unknown what fraction of primary MM cells harbor constitutive NF-κB activity maintained by proteasome-dependent mechanisms. Here, we report an unexpected finding that constitutive NF-κB activity in 10 of 14 primary MM samples analyzed is refractory to inhibition by bortezomib. Moreover, when MM cells were cocultured with MM patient-derived bone marrow stromal cells (BMSC), microenvironment components critical for MM growth and survival, further increases in NF-κB activity were observed that were also refractory to bortezomib. Similarly, MM-BMSCs caused PIR NF-κB activation in the RPMI8226 MM cell line, leading to increased NF-κB–dependent transcription and resistance to bortezomib-induced apoptosis. Our findings show that primary MM cells frequently harbor PIR NF-κB activity that is further enhanced by the presence of patient-derived BMSCs. They also suggest that this activity is likely relevant to the drug resistance development in some patients. Further elucidation of the mechanism of PIR NF-κB regulation could lead to the identification of novel diagnostic biomarkers and/or therapeutic targets for MM treatment. (Mol Cancer Res 2008;6(8):1356–64)
Data from Suppression of IRF4 by IRF1, 3, and 7 in Noxa Expression Is a Necessary Event for IFN-γ–Mediated Tumor Elimination
Authors: Piya Sujan , Moon Ae Ran , Song Peter I. , Hiscott John , Lin Rongtuan , Seol Dai-Wu , Kim Tae-Hyoung
Published: April 2023

Abstract
IFN-γ plays a critical role in tumor immunosurveillance by affecting either immune cells or tumor cells; however, IFN-mediated effects on tumor elimination are largely unknown. In this study, we showed that IFN regulatory factors (IRF) modulated by IFNs up- and downregulated Noxa expression, a prodeath BH3 protein, in various cancer cells. Inhibition of Noxa expression using short hairpin RNA in tumor cells leads to resistance against lipopolysaccharide (LPS)-induced tumor elimination, in which IFN-γ is known as a critical effecter in mice. Chromatin immunoprecipitation analysis in both CT26 cells and SP2/0 cells, sensitive and resistant to LPS-induced tumor elimination, respectively, revealed that the responsiveness of IRF1, 3, 4, and 7 in the Noxa promoter region in response to IFN-γ might be crucial in LPS-induced tumor elimination. IRF1, 3, and 7 were upregulated by IFN-γ and activated Noxa expression, leading to the death of Noxa wild-type baby mouse kidney (BMK) cells but not of Noxa-deficient BMK cells. In contrast, IRF4 acts as a repressor for Noxa expression and inhibits cell death induced by IRF1, 3, or 7. Therefore, although IFN-γ alone are not able to induce cell death in tumor cells in vitro, Noxa induction by IFN-γ, which is regulated by the balance between its activators (IRF1, 3, and 7) and its repressor (IRF4), is crucial to increasing the susceptibility of tumor cells to immune cell-mediated cytotoxicity. Mol Cancer Res; 9(10); 1356–65. ©2011 AACR.
Data from Mutual Regulation of Hypoxia-Inducible Factor and Mammalian Target of Rapamycin as a Function of Oxygen Availability
Authors: Knaup Karl X. , Jozefowski Katrin , Schmidt Regina , Bernhardt Wanja M. , Weidemann Alexander , Juergensen Jan Steffen , Warnecke Christina , Eckardt Kai-Uwe , Wiesener Michael S.
Published: April 2023

Abstract
The mammalian target of rapamycin (mTOR) regulates cellular growth and proliferation, mainly by controlling cellular translation. Most tumors show constitutive activation of the mTOR pathway. In hypoxia, mTOR is inactivated, which is believed to be part of the program of the cell to maintain energy homeostasis. However, certain proteins are believed to be preferentially translated during hypoxia via 5′ terminal oligopyrimidine tract mechanisms with controversial discussion about the involvement of the mTOR-dependent ribosomal protein S6 (rpS6). The hypoxia-inducible transcription factor (HIF) is the master regulator of hypoxic adaptation and itself strongly implicated in tumor growth. HIF is translationally regulated by mTOR. The regulatory features and the involvement of molecular oxygen itself in this regulation of HIF by mTOR are poorly understood. mTOR inhibition leads to profound attenuation of HIFα protein in the majority of primary and cancer cells studied. Under severe hypoxia, no influence of mTOR inhibitors was observed; thus, stimulation of HIFα by mTOR may only be relevant under mild hypoxia or even normoxia. HIF expression and phosphorylated rpS6 negatively correlate in experimental tumors. In cell culture, prolonged hypoxia abolishes rpS6 phosphorylation, which seems to be partly independent of the upstream p70S6 kinase. We show that hypoxic repression of rpS6 is largely dependent on HIF, implicating a negative feedback loop, which may influence cellular translational rates and metabolic homeostasis. These data implicate that the hypoxic microenvironment renders tumor cells resistant to mTOR inhibition, at least concerning hypoxic gene activation, which would add to the difficulties of other established therapeutic strategies in hypoxic cancer tissues. (Mol Cancer Res 2009;7(1):88–98)
Data from Boswellic Acid Blocks Signal Transducers and Activators of Transcription 3 Signaling, Proliferation, and Survival of Multiple Myeloma via the Protein Tyrosine Phosphatase SHP-1
Authors: Kunnumakkara Ajaikumar B. , Nair Asha S. , Sung Bokyung , Pandey Manoj K. , Aggarwal Bharat B.
Published: April 2023

Abstract
Activation of signal transducers and activators of transcription-3 (STAT-3) has been linked with survival, proliferation, chemoresistance, and angiogenesis of tumor cells, including human multiple myeloma (MM). Thus, agents that can suppress STAT3 activation have potential as cancer therapeutics. In our search for such agents, we identified acetyl-11-keto-β-boswellic acid (AKBA), originally isolated from Boswellia serrata. Our results show that AKBA inhibited constitutive STAT3 activation in human MM cells. AKBA suppressed IL-6–induced STAT3 activation, and the inhibition was reversible. The phosphorylation of both Jak 2 and Src, constituents of the STAT3 pathway, was inhibited by AKBA. Interestingly, treatment of cells with pervanadate suppressed the effect of AKBA to inhibit the phosphorylation of STAT3, thus suggesting the involvement of a protein tyrosine phosphatase. We found that AKBA induced Src homology region 2 domain–containing phosphatase 1 (SHP-1), which may account for its role in dephosphorylation of STAT3. Moreover, deletion of the SHP-1 gene by small interfering RNA abolished the ability of AKBA to inhibit STAT3 activation. The inhibition of STAT3 activation by AKBA led to the suppression of gene products involved in proliferation (cyclin D1), survival (Bcl-2, Bcl-xL, and Mcl-1), and angiogenesis (VEGF). This effect correlated with the inhibition of proliferation and apoptosis in MM cells. Consistent with these results, overexpression of constitutive active STAT3 significantly reduced the AKBA-induced apoptosis. Overall, our results suggest that AKBA is a novel inhibitor of STAT3 activation and has potential in the treatment of cancer. (Mol Cancer Res 2009;7(1):118–28)
Data from Regulation of Expression of Stromal-Derived Factor-1 Receptors: CXCR4 and CXCR7 in Human Rhabdomyosarcomas
Authors: Tarnowski Maciej , Grymula Katarzyna , Reca Ryan , Jankowski Kacper , Maksym Radoslaw , Tarnowska Joanna , Przybylski Grzegorz , Barr Frederic G. , Kucia Magdalena , Ratajczak Mariusz Z.
Published: April 2023

Abstract
Rhabdomyosarcomas (RMS) express CXCR4 and CXCR7 receptors that bind prometastatic α-chemokine stromal-derived factor-1 (SDF-1). In this report, we analyzed the activity of both promoters in a model of less metastatic human embryonal-RMS cell line (RD) and more metastatic alveolar-like RMS (RD cells transduced with paired box gene 3/forkhead homologue; PAX3-FKHR fusion gene). First, CXCR4 is barely detectable in RD and becomes upregulated in RD/PAX3-FKHR cells. In contrast, CXCR7 highly expressed in RD becomes downregulated in RD/PAX3-FKHR cells. Next, promoter deletion and mutation studies revealed that whereas (a) expression of CXCR4 in RD and RD/PAX3-FKHR cells required nuclear respiratory factor-1 (NRF-1) binding site and (b) was additionally upregulated by direct interaction of NRF-1 with PAX3-FKHR, CXCR7 promoter activity required a proximal nuclear factor-κB–binding motif. The requirement of these factors for CXCR4 and CXCR7 promoter activities was additionally supported after blocking NRF-1 and nuclear factor-κB. Furthermore, CXCR4 expression in PAX3-FKHR+ RMS cells seems to be enhanced because of the interaction of PAX3-FKHR and NRF-1 proteins in the proximal part of the promoter that prevents access of the negative regulator of transcription YY1 to its binding site. Finally, although hypoxia enhances CXCR4 and CXCR7 promoter activity and receptor expression in RD cells, it inhibits CXCR7 expression in RD/PAX3-FKHR cells. In conclusion, SDF-1 binding receptors CXCR4 and CXCR7 are differently regulated in RMS cells. The upregulation of CXCR4 and downregulation of CXCR7 expression by PAX3-FKHR or hypoxia may give SDF-1 an advantage to better engage the CXCR4 receptor, thus increasing RMS motility. Mol Cancer Res; 8(1); 1–14
Data from Identification of CD44 as a Surface Biomarker for Drug Resistance by Surface Proteome Signature Technology
Authors: Cain Jason W. , Hauptschein Robert S. , Stewart Jean K. , Bagci Tugba , Sahagian Gary G. , Jay Daniel G.
Published: April 2023

Abstract
We developed surface proteome signatures (SPS) for identification of new biomarkers playing a role in cancer drug resistance. SPS compares surface antigen expression of different cell lines by immunocytochemistry of a phage display antibody library directed to surface antigens of HT1080 fibrosarcoma cells. We applied SPS to compare the surface proteomes of two epithelial derived cancer cell lines, MCF7 and NCI/ADR-RES, which is drug resistant because of overexpression of the P-glycoprotein (P-gp) drug efflux pump. Surface proteomic profiling identified CD44 as an additional biomarker that distinguishes between these two cell lines. CD44 immunohistochemistry can distinguish between tumors derived from these lines and predict tumor response to doxorubicin in vivo. We further show that CD44 plays a role in drug resistance, independently of P-gp, in NCI/ADR-RES cells and increases expression of the antiapoptotic protein Bcl-xL. Our findings illustrate the utility of SPS to distinguish between cancer cell lines and their derived tumors and identify novel biomarkers involved in drug resistance. Mol Cancer Res; 9(5); 637–47. ©2011 AACR.
Data from Differential Hypoxic Regulation of Hypoxia-Inducible Factors 1α and 2α
Authors: Lin Qun , Cong Xiangyu , Yun Zhong
Published: April 2023

Abstract
The hypoxia-inducible transcription factors (HIF) 1α and HIF-2α play a critical role in cellular response to hypoxia. Elevated HIF-α expression correlates with poor patient survival in a large number of cancers. Recent evidence suggests that HIF-2α appears to be preferentially expressed in neuronal tumor cells that exhibit cancer stem cell characteristics. These observations suggest that expression of HIF-1α and HIF-2α is differentially regulated in the hypoxic tumor microenvironment. However, the underlying mechanisms remain to be fully investigated. In this study, we investigated the transcriptional regulation of HIF-1α and HIF-2α under different physiologically relevant hypoxic conditions. We found that transcription of HIF-2α was consistently increased by hypoxia, whereas transcription of HIF-1α showed variable levels of repression. Mechanistically, differential regulation of HIF-α transcription involved hypoxia-induced changes in acetylation of core histones H3 and H4 associated with the proximal promoters of the HIF-1α or HIF-2α gene. We also found that, although highly stable under acute hypoxia, HIF-1α and HIF-2α proteins become destabilized under chronic hypoxia. Our results have thus provided new mechanistic insights into the differential regulation of HIF-1α and HIF-2α by the hypoxic tumor microenvironment. These findings also suggest an important role of HIF-2α in the regulation of tumor progression under chronic hypoxia. Mol Cancer Res; 9(6); 757–65. ©2011 AACR.
Data from Inactivation of the MAL Gene in Breast Cancer Is a Common Event That Predicts Benefit from Adjuvant Chemotherapy
Authors: Horne Hisani N. , Lee Paula S. , Murphy Susan K. , Alonso Miguel A. , Olson John A. , Marks Jeffrey R.
Published: April 2023

Abstract
Dysregulation of MAL (myelin and lymphocyte protein) has been implicated in several malignancies including esophageal, ovarian, and cervical cancers. The MAL protein functions in apical transport in polarized epithelial cells; therefore, its disruption may lead to loss of organized polarity characteristic of most solid malignancies. Bisulfite sequencing of the MAL promoter CpG island revealed hypermethylation in breast cancer cell lines and 69% of primary tumors analyzed compared with normal breast epithelial cells. Differential methylation between normal and cancer DNA was confined to the proximal promoter region. In a subset of breast cancer cell lines including T47D and MCF7 cells, promoter methylation correlated with transcriptional silencing that was reversible with the methylation inhibitor 5-aza-2′-deoxycytidine. In addition, expression of MAL reduced motility and resulted in a redistribution of lipid raft components in MCF10A cells. MAL protein expression measured by immunohistochemistry revealed no significant correlation with clinicopathologic features. However, in patients who did not receive adjuvant chemotherapy, reduced MAL expression was a significant predictive factor for disease-free survival. These data implicate MAL as a commonly altered gene in breast cancer with implications for response to chemotherapy. (Mol Cancer Res 2009;7(2):199–209)
Data from Blockade of Fatty Acid Synthase Induces Ubiquitination and Degradation of Phosphoinositide-3-Kinase Signaling Proteins in Ovarian Cancer
Authors: Tomek Katharina , Wagner Renate , Varga Franz , Singer Christian F. , Karlic Heidrun , Grunt Thomas W.
Published: April 2023

Abstract
Aberrations within the phosphoinositide-3-kinase (PI3K) pathway occur in greater than 45% of ovarian carcinomas. The PI3K cascade transmits signals from ErbB receptors downstream to S6 and 4EBP1, which are involved in protein biosynthesis. Many ovarian carcinomas reveal hyperactivation of ErbB1 (epidermal growth factor receptor) or ErbB2 (HER2/neu). Unfortunately, the benefit of anti-ErbB drugs is yet rather limited in ovarian carcinomas. Thus, novel targeting strategies are needed for ovarian carcinomas. The lipogenic enzyme fatty acid synthase (FASN) is overexpressed in approximately 80% of ovarian carcinomas. It stimulates cell growth and signifies poor prognosis. FASN inhibition impedes (ErbB) membrane receptor signaling and sensitizes cells against anti-ErbB drugs. Here, we show that the FASN inhibitor C75 and FASN-targeting siRNAs abrogate growth, induce apoptosis, and downregulate phosphorylation/expression of the PI3K effectors AKT, mTOR, p70S6K, S6, and 4EBP1. In contrast, FASN inhibition impairs expression but only weakly affects phosphorylation of ERK1/2 mitogen-activated protein kinases in ovarian carcinoma cells. Cycloheximide-mediated blockade of protein translation reveals that C75- or FASN siRNA–induced shutdown of FASN accelerates decomposition of signaling proteins. This effect is caused by C75- or FASN siRNA–dependent stimulation of ubiquitination followed by lysosomal-autophagosomal proteolysis. In contrast, PI3K inhibitor LY294002 blocks phosphorylation but does not reduce expression/stability of PI3K effectors. Forced expression of hyperactive (HA) AKT1, unlike HA-MEK1, impairs the growth-inhibitory action of C75. We provide first evidence that the anticancer action of FASN inhibitors is at least partially mediated by drug-dependent proteolysis of PI3K effectors. FASN is a promising cancer target, whose inhibition not only abrogates lipogenesis, which is indispensable for cancer growth, but also downregulates oncogenic PI3K signaling. Mol Cancer Res; 9(12); 1767–79. ©2011 AACR.
Data from Oncogenic Ras and Transforming Growth Factor-β Synergistically Regulate AU-Rich Element–Containing mRNAs during Epithelial to Mesenchymal Transition
Authors: Kanies Cindy L. , Smith J. Joshua , Kis Christian , Schmidt Carl , Levy Shawn , Khabar Khalid S.A. , Morrow Jason , Deane Natasha , Dixon Dan A. , Beauchamp R. Daniel
Published: April 2023

Abstract
Colon cancer progression is characterized by activating mutations in Ras and by the emergence of the tumor-promoting effects of transforming growth factor-β (TGF-β) signaling. Ras-inducible rat intestinal epithelial cells (RIE:iRas) undergo a well-described epithelial to mesenchymal transition and invasive phenotype in response to H-RasV12 expression and TGF-β treatment, modeling tumor progression. We characterized global gene expression profiles accompanying Ras-induced and TGF-β–induced epithelial to mesenchymal transition in RIE:iRas cells by microarray analysis and found that the regulation of gene expression by the combined activation of Ras and TGF-β signaling was associated with enrichment of a class of mRNAs containing 3′ AU-rich element (ARE) motifs known to regulate mRNA stability. Regulation of ARE-containing mRNA transcripts was validated at the mRNA level, including genes important for tumor progression. Ras and TGF-β synergistically increased the expression and mRNA stability of vascular endothelial growth factor (VEGF), a key regulator of tumor angiogenesis, in both RIE:iRas cells and an independent cell culture model (young adult mouse colonocyte). Expression profiling of human colorectal cancers (CRC) further revealed that many of these genes, including VEGF and PAI-1, were differentially expressed in stage IV human colon adenocarcinomas compared with adenomas. Furthermore, genes differentially expressed in CRC are also significantly enriched with ARE-containing transcripts. These studies show that oncogenic Ras and TGF-β synergistically regulate genes containing AREs in cultured rodent intestinal epithelial cells and suggest that posttranscriptional regulation of gene expression is an important mechanism involved in cellular transformation and CRC tumor progression. (Mol Cancer Res 2008;6(7):1124–36)
Data from Zebrafish Hagoromo Mutants Up-Regulate fgf8 Postembryonically and Develop Neuroblastoma
Authors: Amsterdam Adam , Lai Kevin , Komisarczuk Anna Z. , Becker Thomas S. , Bronson Roderick T. , Hopkins Nancy , Lees Jacqueline A.
Published: April 2023

Abstract
We screened an existing collection of zebrafish insertional mutants for cancer susceptibility by histologic examination of heterozygotes at 2 years of age. As most mutants had no altered cancer predisposition, this provided the first comprehensive description of spontaneous tumor spectrum and frequency in adult zebrafish. Moreover, the screen identified four lines, each carrying a different dominant mutant allele of Hagoromo previously linked to adult pigmentation defects, which develop tumors with high penetrance and that histologically resemble neuroblastoma. These tumors are clearly neural in origin, although they do not express catecholaminergic neuronal markers characteristic of human neuroblastoma. The zebrafish tumors result from inappropriate maintenance of a cell population within the cranial ganglia that are likely neural precursors. These neoplasias typically remain small but they can become highly aggressive, initially traveling along cranial nerves, and ultimately filling the head. The developmental origin of these tumors is highly reminiscent of human neuroblastoma. The four mutant Hagoromo alleles all contain viral insertions in the fbxw4 gene, which encodes an F-box WD40 domain–containing protein. However, although one allele clearly reduced the levels of fbxw4 mRNA, the other three insertions had no detectable effect on fbw4 expression. Instead, we showed that all four mutations result in the postembryonic up-regulation of the neighboring gene, fibroblast growth factor 8 (fgf8). Moreover, fgf8 is highly expressed in the tumorigenic lesions. Although fgf8 overexpression is known to be associated with breast and prostate cancer in mammals, this study provides the first evidence that fgf8 misregulation can lead to neural tumors. (Mol Cancer Res 2009;7(6):841–50)
Data from Epigenetic Upregulation of Urokinase Plasminogen Activator Promotes the Tropism of Mesenchymal Stem Cells for Tumor Cells
Authors: Pulukuri Sai Murali Krishna , Gorantla Bharathi , Dasari Venkata Ramesh , Gondi Christopher S. , Rao Jasti S.
Published: April 2023

Abstract
A major obstacle for the effective treatment of cancer is the invasive capacity of the tumor cells. Previous studies have shown the capability of mesenchymal stem cells (MSC) to target these disseminated tumor cells and to serve as therapeutic delivery vehicles. However, the molecular mechanisms that would enhance the migration of MSCs toward tumor areas are not well understood. In particular, very little is known about the role that epigenetic mechanisms play in cell migration and tropism of MSCs. In this study, we investigated whether histone deacetylation was involved in the repression of urokinase plasminogen activator (uPA) expression in MSCs derived from umbilical cord blood (CB) and bone marrow (BM). Induction of uPA expression by histone deacetylase inhibitors trichostatin A and sodium butyrate was observed in CB- and BM-derived MSCs examined. In vitro migration assays showed that induction of uPA expression by histone deacetylase inhibitors in CB- and BM-derived MSCs significantly enhanced tumor tropism of these cells. Furthermore, overexpression of uPA in CB-MSCs induced migration capacity toward human cancer cells in vitro. In addition, our results showed that uPA-uPAR knockdown in PC3 prostate cancer cells significantly inhibited tumor-specific migration of uPA-overexpressing MSCs. These results have significant implications for the development of MSC-mediated, tumor-selective gene therapies. Mol Cancer Res; 8(8); 1074–83. ©2010 AACR.
Data from Temporal and Spatial Cooperation of Snail1 and Twist1 during Epithelial–Mesenchymal Transition Predicts for Human Breast Cancer Recurrence
Authors: Tran David D. , Corsa Callie Ann S. , Biswas Hirak , Aft Rebecca L. , Longmore Gregory D.
Published: April 2023

Abstract
Epithelial–mesenchymal transition (EMT) is a normal developmental program that is considered to also play an important role in cancer metastasis. Ultimate inducers of EMT are transcriptional repressors that individually can induce experimental EMT, yet in many cells, particularly cancer cells, multiple inducers are expressed simultaneously. Why, and if, and how they interact to regulate EMT is unanswered. Using RNA interference technology to affect protein knockdown and avoid potential overexpression artifact coupled with transient TGFβ treatment to better mimic in vivo conditions we show, in both nontumorigenic and tumorigenic epithelial cancer cells, that Snail1 is uniquely required for EMT initiation, whereas Twist1 is required to maintain late EMT. Twist1, present in resting epithelial cells, is dispensable for EMT initiation. Mechanistically, in response to transient TGFβ treatment, transient Snail1 expression represses Twist1 transcription directly, which is subsequently upregulated, as Snail1 levels decrease, to sustain E-cadherin downregulation and growth arrest of EMT. Persistent Twist1 expression is associated with a p38 and extracellular signal–regulated kinase signal feedback loop that sustains growth-inhibitory signals characteristic of quiescent micrometastatic tumors. This Snail1–Twist1 temporal and spatial cooperation was also observed in vivo during human breast cancer progression to metastasis. Twist1 level, but not Snail1 level, and Twist1:Snail1 ratio in disseminated micrometastatic bone marrow tumor cells was found to correlate with survival and treatment resistance and is highly predictive of metastatic or recurrent disease. Mol Cancer Res; 9(12); 1644–57. ©2011 AACR.
Data from Extracellular Matrix-Induced Gene Expression in Human Breast Cancer Cells
Authors: Garamszegi Nandor , Garamszegi Susanna P. , Shehadeh Lina A. , Scully Sean P.
Published: April 2023

Abstract
Extracellular matrix (ECM) molecules modify gene expression through attachment-dependent (focal adhesion-related) integrin receptor signaling. It was previously unknown whether the same molecules acting as soluble peptides could generate signal cascades without the associated mechanical anchoring, a condition that may be encountered during matrix remodeling and degradation and relevant to invasion and metastatic processes. In the current study, the role of ECM ligand-regulated gene expression through this attachment-independent process was examined. It was observed that fibronectin, laminin, and collagen type I and II induce Smad2 activation in MCF-10A and MCF-7 cells. This activation is not caused by transforming growth factor (TGF)-β ligand contamination or autocrine TGF involvement and is 3- to 5-fold less robust than the TGF-β1 ligand. The resulting nuclear translocation of Smad4 in response to ECM ligand indicates downstream transcriptional responses occurring. Coimmunoprecipitation experiments determined that collagen type II and laminin act through interaction with integrin α2β1 receptor complex. The ECM ligand-induced Smad activation (termed signaling crosstalk) resulted in cell type and ligand-specific transcriptional changes, which are distinct from the TGF-β ligand-induced responses. These findings show that cell-matrix communication is more complex than previously thought. Soluble ECM peptides drive transcriptional regulation through corresponding adhesion and non-attachment-related processes. The resultant gene expressional patterns correlate with pathway activity and not by the extent of Smad activation. These results extend the complexity and the existing paradigms of ECM-cell communication to ECM ligand regulation without the necessity of mechanical coupling. (Mol Cancer Res 2009;7(3):319–29)
Data from The MicroRNA Profile of Prostate Carcinoma Obtained by Deep Sequencing
Authors: Szczyrba Jaroslaw , Löprich Elke , Wach Sven , Jung Volker , Unteregger Gerhard , Barth Stephanie , Grobholz Rainer , Wieland Wolf , Stöhr Robert , Hartmann Arndt , Wullich Bernd , Grässer Friedrich
Published: April 2023

Abstract
Prostate cancer is a leading cause of tumor mortality. To characterize the underlying molecular mechanisms, we have compared the microRNA (miRNA) profile of primary prostate cancers and noncancer prostate tissues using deep sequencing. MiRNAs are small noncoding RNAs of 21 to 25 nucleotides that regulate gene expression through the inhibition of protein synthesis. We find that 33 miRNAs were upregulated or downregulated >1.5-fold. The deregulation of selected miRNAs was confirmed by both Northern blotting and quantitative reverse transcription-PCR in established prostate cancer cell lines and clinical tissue samples. A computational search indicated the 3′-untranslated region (UTR) of the mRNA for myosin VI (MYO6) as a potential target for both miR-143 and miR-145, the expression of which was reduced in the tumor tissues. Upregulation of myosin VI in prostate cancer was previously shown by immunohistochemistry. The level of MYO6 mRNA was significantly induced in all primary tumor tissues compared with the nontumor tissue from the same patient. This finding was matched to the upregulation of myosin VI in established prostate cancer cell lines. In luciferase reporter analysis, we find a significant negative regulatory effect on the MYO6 3′UTR by both miR-143 and miR-145. Mutation of the potential binding sites for miR-143 and miR-145 in the MYO6 3′UTR resulted in a loss of responsiveness to the corresponding miRNA. Our data indicate that miR-143 and miR-145 are involved in the regulation of MYO6 expression and possibly in the development of prostate cancer. Mol Cancer Res; 8(4); 529–38. ©2010 AACR.
Data from PRMT5, a Novel TRAIL Receptor-Binding Protein, Inhibits TRAIL-Induced Apoptosis via Nuclear Factor-κB Activation
Authors: Tanaka Hiroshi , Hoshikawa Yutaka , Oh-hara Tomoko , Koike Sumie , Naito Mikihiko , Noda Tetsuo , Arai Hiroyuki , Tsuruo Takashi , Fujita Naoya
Published: April 2023

Abstract
Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is a member of the TNF superfamily and has selective antitumor activity. Although TNF-α-induced intracellular signaling pathways have been well studied, TRAIL signaling is not fully understood. Here, we identified a novel TRAIL receptor-binding protein, protein arginine methyltransferase 5 (PRMT5), as a result of proteomic screening. PRMT5 selectively interacted with death receptor 4 and death receptor 5 but not with TNF receptor 1 or Fas. PRMT5 gene silencing sensitized various cancer cells to TRAIL without affecting TRAIL resistance in nontransformed cells. PRMT5 contributed to TRAIL-induced activation of inhibitor of κB kinase (IKK) and nuclear factor-κB (NF-κB), leading to induction of several NF-κB target genes. Although IKK inhibition increased sensitivity to both TRAIL and TNF-α, PRMT5 knockdown potentiated TRAIL-mediated cytotoxicity alone. PRMT5 had no effect on TNF-α-mediated NF-κB signaling. These results show the selectivity of PRMT5 for TRAIL signaling. The PRMT5 small interfering RNA-mediated susceptibility to TRAIL was rescued by ectopic expression of active IKKβ, confirming the involvement of PRMT5 in TRAIL resistance by activating the NF-κB pathway. Collectively, our findings suggest the therapeutic potential of PRMT5 in TRAIL-based cancer treatments.(Mol Cancer Res 2009;7(4):557–69)
Data from Circulating Tumor Cells from Patients with Advanced Prostate and Breast Cancer Display Both Epithelial and Mesenchymal Markers
Authors: Armstrong Andrew J. , Marengo Matthew S. , Oltean Sebastian , Kemeny Gabor , Bitting Rhonda L. , Turnbull James D. , Herold Christina I. , Marcom Paul K. , George Daniel J. , Garcia-Blanco Mariano A.
Published: April 2023

Abstract
During cancer progression, malignant cells undergo epithelial-mesenchymal transitions (EMT) and mesenchymal-epithelial transitions (MET) as part of a broad invasion and metastasis program. We previously observed MET events among lung metastases in a preclinical model of prostate adenocarcinoma that suggested a relationship between epithelial plasticity and metastatic spread. We thus sought to translate these findings into clinical evidence by examining the existence of EMT in circulating tumor cells (CTC) from patients with progressive metastatic solid tumors, with a focus on men with castration-resistant prostate cancer (CRPC) and women with metastatic breast cancer. We showed that the majority (>80%) of these CTCs in patients with metastatic CRPC coexpress epithelial proteins such as epithelial cell adhesion molecule (EpCAM), cytokeratins (CK), and E-cadherin, with mesenchymal proteins including vimentin, N-cadherin and O-cadherin, and the stem cell marker CD133. Equally, we found that more than 75% of CTCs from women with metastatic breast cancer coexpress CK, vimentin, and N-cadherin. The existence and high frequency of these CTCs coexpressing epithelial, mesenchymal, and stem cell markers in patients with progressive metastases has important implications for the application and interpretation of approved methods to detect CTCs. Mol Cancer Res; 9(8); 997–1007. ©2011 AACR.
Data from β-Arrestin/Ral Signaling Regulates Lysophosphatidic Acid–Mediated Migration and Invasion of Human Breast Tumor Cells
Authors: Li Timothy T. , Alemayehu Mistre , Aziziyeh Adel I. , Pape Cynthia , Pampillo Macarena , Postovit Lynne-Marie , Mills Gordon B. , Babwah Andy V. , Bhattacharya Moshmi
Published: April 2023

Abstract
The lipid mediator lysophosphatidic acid (LPA) plays a role in cancer progression and signals via specific G protein–coupled receptors, LPA1-3. LPA has been shown to enhance the metastasis of breast carcinoma cells to bone. However, the mechanisms by which LPA receptors regulate breast cancer cell migration and invasion remain unclear. Breast cancer cell proliferation has been shown to be stimulated by Ral GTPases, a member of the Ras superfamily. Ral activity can be regulated by the multifunctional protein β-arrestin. We now show that HS578T and MDA-MB-231 breast cancer cells and MDA-MB-435 melanoma cells have higher expression of β-arrestin 1 mRNA compared with the nontumorigenic mammary MCF-10A cells. Moreover, we found that the mRNA levels of LPA1, LPA2, β-arrestin 2, and Ral GTPases are elevated in the advanced stages of breast cancer. LPA stimulates the migration and invasion of MDA-MB-231 cells, but not of MCF-10A cells, and this is mediated by pertussis toxin–sensitive G proteins and LPA1. However, ectopic expression of LPA1 in MCF-10A cells caused these cells to acquire an invasive phenotype. Gene knockdown of either β-arrestin or Ral proteins significantly impaired LPA-stimulated migration and invasion. Thus, our data show a novel role for β-arrestin/Ral signaling in mediating LPA-induced breast cancer cell migration and invasion, two important processes in metastasis. (Mol Cancer Res 2009;7(7):1064–77)
Data from Up-Regulation of Connective Tissue Growth Factor in Endothelial Cells by the Microtubule-Destabilizing Agent Combretastatin A-4
Authors: Samarin Jana , Rehm Margot , Krueger Bettina , Waschke Jens , Goppelt-Struebe Margarete
Published: April 2023

Abstract
Incubation of microvascular endothelial cells with combretastatin A-4 phosphate (CA-4P), a microtubule-destabilizing compound that preferentially targets tumor vessels, altered cell morphology and induced scattering of Golgi stacks. Concomitantly, CA-4P up-regulated connective tissue growth factor (CTGF/CCN2), a pleiotropic factor with antiangiogenic properties. In contrast to the effects of other microtubule-targeting agents such as colchicine or nocodazole, up-regulation of CTGF was only detectable in sparse cells, which were not embedded in a cell monolayer. Furthermore, CA-4P induced CTGF expression in endothelial cells, forming tube-like structures on basement membrane gels. Up-regulation of CTGF by CA-4P was dependent on Rho kinase signaling and was increased when p42/44 mitogen-activated protein kinase was inhibited. Additionally, FoxO transcription factors were identified as potent regulators of CTGF expression in endothelial cells. Activation of FoxO transcription factors by inhibition of phosphatidylinositol 3-kinase/AKT signaling resulted in a synergistic increase in CA-4P-mediated CTGF induction. CA-4P-mediated expression of CTGF was thus potentiated by the inhibition of kinase pathways, which are targets of novel antineoplastic drugs. Up-regulation of CTGF by low concentrations of CA-4P may thus occur in newly formed tumor vessels and contribute to the microvessel destabilization and antiangiogenic effects of CA-4P observed in vivo. (Mol Cancer Res 2009;7(2):180–8)
Data from Downregulation of Sec23A Protein by miRNA-375 in Prostate Carcinoma
Authors: Szczyrba Jaroslaw , Nolte Elke , Wach Sven , Kremmer Elisabeth , Stöhr Robert , Hartmann Arndt , Wieland Wolf , Wullich Bernd , Grässer Friedrich A.
Published: April 2023

Abstract
Prostate carcinoma (CaP) is a leading cause of cancer-related death in men. We have previously determined the microRNA (miRNA) profile of primary CaP in comparison with nontumor prostate tissue. miRNAs are small, noncoding RNAs that inhibit protein synthesis on a posttranscriptional level by binding to the 3′-untranslated region (3′-UTR) of their target genes. In primary CaP tissue, we have previously found by miRNA sequencing that miR-375 and miR-200c were upregulated 9.1- and 4.5-fold, respectively. A computational analysis predicted the 3′-UTR of the SEC23A gene as a potential target for both miR-375 and miR-200c. Here, we show that the 3′-UTR of SEC23A mRNA is indeed a target for miR-375 and miR-200c and that both miRNAs downregulate Sec23A protein expression when ectopically expressed in human 293T cells. In primary samples of CaP, we found a direct correlation between reduction of SEC23A mRNA and overexpression of miR-375 but not of miR-200c. The reduced levels of Sec23A protein were inversely correlated to the increased amount of miR-375 in the LNCaP and DU145 CaP cell lines when compared with normal prostate fibroblasts. In primary CaP, we also detected decreased amounts of Sec23A protein when compared with corresponding normal prostate tissue. Ectopically overexpressed Sec23A in LNCaP and DU145 CaP cells significantly reduced the growth properties, indicating that Sec23A might play a role in the induction or growth of prostate carcinoma. Sec23A overexpression reduced cell growth but did not induce apoptosis, whereas inhibition of Sec23A stimulated cell proliferation. Mol Cancer Res; 9(6); 791–800. ©2011 AACR.
Data from Metformin Induces Both Caspase-Dependent and Poly(ADP-ribose) Polymerase-Dependent Cell Death in Breast Cancer Cells
Authors: Zhuang Yongxian , Miskimins W. Keith
Published: April 2023

Abstract
There is substantial evidence that metformin, a drug used to treat type 2 diabetics, is potentially useful as a therapeutic agent for cancer. However, a better understanding of the molecular mechanisms through which metformin promotes cell-cycle arrest and cell death of cancer cells is necessary. It will also be important to understand how the response of tumor cells differs from normal cells and why some tumor cells are resistant to the effects of metformin. We have found that exposure to metformin induces cell death in all but one line, MDA-MB-231, in a panel of breast cancer cell lines. MCF10A nontransformed breast epithelial cells were resistant to the cytotoxic effects of metformin, even after extended exposure to the drug. In sensitive lines, cell death was mediated by both apoptosis and a caspase-independent mechanism. The caspase-independent pathway involves activation of poly(ADP-ribose) polymerase (PARP) and correlates with enhanced synthesis of PARP and nuclear translocation of apoptosis-inducing factor (AIF), which plays an important role in mediating cell death. Metformin-induced, PARP-dependent cell death is associated with a striking enlargement of mitochondria. Mitochondrial enlargement was observed in all sensitive breast cancer cell lines but not in nontransformed cells or resistant MDA-MB-231. Mitochondrial enlargement was prevented by inhibiting PARP activity or expression. A caspase inhibitor blocked metformin-induced apoptosis but did not affect PARP-dependent cell death or mitochondrial enlargement. Thus, metformin has cytotoxic effects on breast cancer cells through 2 independent pathways. These findings will be pertinent to efforts directed at using metformin or related compounds for cancer therapy. Mol Cancer Res; 9(5); 603–15. ©2011 AACR.
Data from Interplay Between pVHL and mTORC1 Pathways in Clear-Cell Renal Cell Carcinoma
Authors: Kucejova Blanka , Peña-Llopis Samuel , Yamasaki Toshinari , Sivanand Sharanya , Tran Tram Anh T. , Alexander Shane , Wolff Nicholas C. , Lotan Yair , Xie Xian-Jin , Kabbani Wareef , Kapur Payal , Brugarolas James
Published: April 2023

Abstract
mTOR complex 1 (mTORC1) is implicated in cell growth control and is extensively regulated. We previously reported that in response to hypoxia, mTORC1 is inhibited by the protein regulated in development and DNA damage response 1 (REDD1). REDD1 is upregulated by hypoxia-inducible factor (HIF)-1, and forced REDD1 expression is sufficient to inhibit mTORC1. REDD1-induced mTORC1 inhibition is dependent on a protein complex formed by the tuberous sclerosis complex (TSC)1 and 2 (TSC2) proteins. In clear-cell renal cell carcinoma (ccRCC), the von Hippel-Lindau (VHL) gene is frequently inactivated leading to constitutive activation of HIF-2 and/or HIF-1, which may be expected to upregulate REDD1 and inhibit mTORC1. However, mTORC1 is frequently activated in ccRCC, and mTORC1 inhibitors are effective against this tumor type; a paradox herein examined. REDD1 was upregulated in VHL-deficient ccRCC by in silico microarray analyses, as well as by quantitative real-time PCR, Western blot, and immunohistochemistry. Vhl disruption in a mouse model was sufficient to induce Redd1. Using ccRCC-derived cell lines, we show that REDD1 upregulation in tumors is VHL dependent and that both HIF-1 and HIF-2 are, in a cell-type-dependent manner, recruited to, and essential for, REDD1 induction. Interestingly, whereas mTORC1 is responsive to REDD1 in some tumors, strategies have evolved in others, such as mutations disrupting TSC1, to subvert mTORC1 inhibition by REDD1. Sequencing analyses of 77 ccRCCs for mutations in TSC1, TSC2, and REDD1, using PTEN as a reference, implicate the TSC1 gene, and possibly REDD1, as tumor suppressors in sporadic ccRCC. Understanding how ccRCCs become refractory to REDD1-induced mTORC1 inhibition should shed light into the development of ccRCC and may aid in patient selection for molecular-targeted therapies. Mol Cancer Res; 9(9); 1255–65. ©2011 AACR.
Data from Inhibition of G1 to S Phase Progression by a Novel Zinc Finger Protein P58TFL at P-bodies
Authors: Minagawa Kentaro , Katayama Yoshio , Nishikawa Shinichiro , Yamamoto Katsuya , Sada Akiko , Okamura Atsuo , Shimoyama Manabu , Matsui Toshimitsu
Published: April 2023

Abstract
We recently reported the translocation of the immunoglobulin (Ig) light chain κ locus gene with a possible tumor suppressor gene, TFL, in transformed follicular lymphoma. However, the functional significance in cell transformation remains to be elucidated. Here, we first identified two gene products, P58TFL and P36TFL, derived by alternative splicing. The expression was prominent in normal human lymphocytes but defective in some leukemia/lymphoma cell lines. Overexpression of either protein in a mouse pro-B cell line, Ba/F3, and a human leukemia cell line, Jurkat, inhibited G1 to S phase progression through suppression of retinoblastoma protein (Rb) phosphorylation. The dominant gene product, P58TFL, colocalized with mRNA-processing body markers, eukaryotic translation initiation factor 2C and DCP1 decapping-enzyme homolog A, but not with a stress granule maker, T-cell intracellular antigen 1, in the cytoplasm. Taken together with the unique CCCH-type zinc finger motif, the present study suggests that P58TFL could play an important role in the regulation of cell growth through posttranscriptional modification of cell cycle regulators, at least partially, upstream of Rb. (Mol Cancer Res 2009;7(6):880–9)