Regulating Tumor Cell Proliferation Research Articles

Ganglioside GM3 promotes HGF-stimulated motility of murine hepatoma cell through enhanced phosphorylation of cMet at specific tyrosine sites and PI3K/Akt-mediated migration signaling

Ganglioside GM3 plays a well-documented and important role in the regulation of tumor cell proliferation, invasion, and metastasis by modulating tyrosine kinase growth factor receptors. However, the effect of GM3 on the hepatocyte growth factor receptor (HGFR, cMet) has not been fully delineated. In the current study, we investigated how GM3 affects cMet signaling and HGF-stimulated cell motility and migration using three hepatic cancer cell lines of mouse (Hca/A2, Hca/16A3, and Hepa1-6). Decreasing GM3 expression with the use of P4, a specific inhibitor for ganglioside synthesis inhibited the HGF-stimulated phosphorylation of cMet and activity of PI3K/Akt signaling pathway. In contrast, the increased expression of GM3 as a result of adding exogenous GM3 enhanced the HGF-stimulated phosphorylation of cMet and activity of PI3K/Akt signaling pathway. Furthermore, HGF-stimulated cell motility and migration in vitro were inhibited by reduced expression of GM3 and enhanced by increased expression of GM3. All the observations indicate that ganglioside GM3 promotes HGF-stimulated motility of murine hepatoma cell through enhanced phosphorylation of cMet at specific tyrosine sites and PI3K/Akt-mediated migration signaling.

Annexin A1: A new biomarker for predicting nasopharyngeal carcinoma response to radiotherapy

Radiotherapy is the primary treatment for nasopharyngeal carcinoma (NPC), but radioresistance remains a serious obstacle to successful treatment in many cases. Therefore, the biomarkers for predicting NPC response to radiotherapy are very important for targeted therapy and individualized radiotherapy of NPC. Accumulating evidences have shown that Annexin A1 was correlated with NPC radioresistance. First, Annexin A1 is a potential tumor suppressor gene, and can regulate tumor cell proliferation and apoptosis, thus abnormal expression of Annexin A1 in NPC affects apoptosis of tumor cells induced by ionizing radiation and radiotherapeutic efficacy. Second, Annexin A1 is one of the proteins that are involved in p53-mediated radioresponse in NPC, and it might be related to NPC radioresistance. Third, the expression level of Annexin A1 is down-regulated in NPC, and is correlated with metastasis, recurrence and poor prognosis of NPC, thus Annexin A1 downregulation may increase NPC radioresistance, leading to poor prognosis. Last but not the least, Annexin A1 is closely related with tumor chemoresistance, whereas radioresistance is similar to chemoresistance in many aspects, thus Annexin A1 may also be involved in NPC radioresistance. Based on the above mentions, we hypothesize that Annexin A1 is closely correlated with NPC radioresistance and is an important new biomarker for predicting NPC response to radiotherapy.

Protein kinase Cα suppresses Kras-mediated lung tumor formation through activation of a p38 MAPK-TGFβ signaling axis

Protein kinase C alpha (PKCα) can activate both pro- and anti-tumorigenic signaling depending upon cellular context. Here, we investigated the role of PKCα in lung tumorigenesis in vivo. Gene expression data sets revealed that primary human non-small lung cancers (NSCLC) express significantly decreased PKCα levels, indicating that loss of PKCα expression is a recurrent event in NSCLC. We evaluated the functional relevance of PKCα loss during lung tumorigenesis in three murine lung adenocarcinoma models (LSL-Kras, LA2-Kras and urethane exposure). Genetic deletion of PKCα resulted in a significant increase in lung tumor number, size, burden and grade, bypass of oncogene-induced senescence, progression from adenoma to carcinoma and a significant decrease in survival in vivo. The tumor promoting effect of PKCα loss was reflected in enhanced Kras-mediated expansion of bronchio-alveolar stem cells (BASCs), putative tumor-initiating cells, both in vitro and in vivo. LSL-Kras/Prkca(-/-) mice exhibited a decrease in phospho-p38 MAPK in BASCs in vitro and in tumors in vivo, and treatment of LSL-Kras BASCs with a p38 inhibitor resulted in increased colony size indistinguishable from that observed in LSL-Kras/Prkca(-/-) BASCs. In addition, LSL-Kras/Prkca(-/-) BASCs exhibited a modest but reproducible increase in TGFβ1 mRNA, and addition of exogenous TGFβ1 to LSL-Kras BASCs results in enhanced growth similar to untreated BASCs from LSL-Kras/Prkca(-/-) mice. Conversely, a TGFβR1 inhibitor reversed the effects of PKCα loss in LSL-Kras/Prkca(-/-) BASCs. Finally, we identified the inhibitors of DNA binding (Id) Id1-3 and the Wilm's Tumor 1 as potential downstream targets of PKCα-dependent tumor suppressor activity in vitro and in vivo. We conclude that PKCα suppresses tumor initiation and progression, at least in part, through a PKCα-p38MAPK-TGFβ signaling axis that regulates tumor cell proliferation and Kras-induced senescence. Our results provide the first direct evidence that PKCα exhibits tumor suppressor activity in the lung in vivo.

Alpha-CaMKII Plays a Critical Role in Determining the Aggressive Behavior of Human Osteosarcoma

Osteosarcoma is among the most frequently occurring primary bone tumors, primarily affecting adolescents and young adults. Despite improvements in osteosarcoma treatment, more specific molecular targets are needed as potential therapeutic options. One target of interest is α-Ca(2+)/calmodulin-dependent protein kinase II (α-CaMKII), a ubiquitous mediator of Ca(2+)-linked signaling, which has been shown to regulate tumor cell proliferation and differentiation. Here, we investigate the role of α-CaMKII in the growth and tumorigenicity of human osteosarcoma. We show that α-CaMKII is highly expressed in primary osteosarcoma tissue derived from 114 patients, and is expressed in varying levels in different human osteosarcoma (OS) cell lines [MG-63, N-methyl-N'-nitro-N-nitrosoguanidine (MNNG)/HOS, and 143B). To examine whether α-CaMKII regulates osteosarcoma tumorigenic properties, we genetically inhibited α-CaMKII in two osteosarcoma cell lines using two different α-CaMKII shRNAs delivered by lentiviral vectors and overexpressed α-CaMKII by retrovirus. The genetic deletion of α-CaMKII by short hairpin RNA (shRNA) in MG-63 and 143B cells resulted in decreased proliferation (50% and 41%), migration (22% and 25%), and invasion (95% and 90%), respectively. The overexpression of α-CaMKII in HOS cells resulted in increased proliferation (240%), migration (640%), and invasion (10,000%). Furthermore, α-CaMKII deletion in MG-63 cells significantly reduced tumor burden in vivo (65%), whereas α-CaMKII overexpression resulted in tumor formation in a previously nontumor forming osteosarcoma cell line (HOS). Our results suggest that α-CaMKII plays a critical role in determining the aggressive phenotype of osteosarcoma, and its inhibition could be an attractive therapeutic target to combat this devastating adolescent disease.

Abstract 3107: Runt-related transcription factor 1 (RUNX1) is involved in transcriptional repression by estrogen receptor and breast cancer cell proliferation.

Abstract In the majority of breast cancers, estrogen and its cellular receptor, estrogen receptor (ER), are involved in regulating tumor cell proliferation. ER is both a prognostic marker and a therapeutic target in the management of hormone-dependent tumors. Current research efforts and the paradigm of ER transcriptional regulatory functions in breast cancer have focused on the role of ER in activating target gene transcription. Microarray studies and whole-genome ER binding site mapping experiments indicate, however, that ER can alternatively directly repress the expression of a significant subset of target genes. We posit that transcriptional repression by ER likely involves interactions with other co-regulatory transcription factors found in the proximity of ER binding sites in the cis-regulatory regions of repressed target genes and have identified the runt-related transcription factor 1 (RUNX1), a key regulator of hematopoiesis and leukemogenesis, as a candidate co-regulatory factor involved in transcriptional repression by ER. Here, we demonstrate that RUNX1 interacts with ER, specifically the ligand-binding domain, and is involved in the repression of ER target genes and estrogen-dependent and -independent proliferation of breast cancer cells. We also show that putative ER-RUNX1 target genes are associated with disease outcome in breast cancer patients. These findings not only provide the first evidence for the involvement of RUNX1 in hormonal carcinogenesis and additional insights into mechanisms of transcriptional regulation by ER but may potentially lead to more effective strategies for targeting ER and estrogen signaling in the prevention and treatment of breast cancer. Citation Format: Ka Liu, Richard L. Smith, Trang Nguyen-Vu, Nicholes R. Candelaria, Colin M. Rogerson, W. Evan Johnson, Edwin Cheung, Boris R. Jankovic, Wail Ba alawi, Jinsong Zhang, Vladimir B. Bajic, Chin-Yo Lin. Runt-related transcription factor 1 (RUNX1) is involved in transcriptional repression by estrogen receptor and breast cancer cell proliferation. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3107. doi:10.1158/1538-7445.AM2013-3107

594 SPECIFIC INHIBITON OF NOTCH-2 AS A NOVEL THERAPY FOR INVASIVE BLADDER CANCER

You have accessJournal of UrologyBladder Cancer: Basic Research (I)1 Apr 2013594 SPECIFIC INHIBITON OF NOTCH-2 AS A NOVEL THERAPY FOR INVASIVE BLADDER CANCER Tetsutaro Hayashi, Kilian M. Gust, Wolfgang Jaeger, Shannon Awrey, Na Li, Manuel Altamirano-Dimas, Ralph Buttyan, Estelle Li, Ladan Fazli, and Peter C. Black Tetsutaro HayashiTetsutaro Hayashi Vancouver, Canada More articles by this author , Kilian M. GustKilian M. Gust Vancouver, Canada More articles by this author , Wolfgang JaegerWolfgang Jaeger Vancouver, Canada More articles by this author , Shannon AwreyShannon Awrey Vancouver, Canada More articles by this author , Na LiNa Li Vancouver, Canada More articles by this author , Manuel Altamirano-DimasManuel Altamirano-Dimas Vancouver, Canada More articles by this author , Ralph ButtyanRalph Buttyan Vancouver, Canada More articles by this author , Estelle LiEstelle Li Vancouver, Canada More articles by this author , Ladan FazliLadan Fazli Vancouver, Canada More articles by this author , and Peter C. BlackPeter C. Black Vancouver, Canada More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2013.02.1990AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Notch signaling guides decisions regarding cell fate, establishment of cell lineages, stem cell maintenance and differentiation during early development. Notch is active in many adult malignancies where it has been shown to regulate tumor cell proliferation and progression to aggressive disease. Previously, we showed that Notch is active in certain bladder cancer cells and that active Notch2 signaling mediates cell migratory and invasive behaviors. Here we report outcomes of Notch2 knockdown or treatment with a unique Notch2 inhibitor, NRR2Mab, on these and other aggressive behaviors in our bladder cancer cell models. METHODS UMUC3, UMUC13 and UMUC16 cells, representing invasive bladder cancer cells with high Notch2 expression, were transduced with Notch2 or non-targeting shRNA in a lentiviral vector and were stably propogated. A Notch2-inactivating monoclonal antibody (Mab), NRR2Mab, and an isotype-matched, non-targeting Mab were kindly provided by Genentech, Inc. (San Francisco, CA). Cells were grown under adherent (AH) or anchorage-independent conditions (AI) and the effects of Notch2 silencing or Mab inactivation were quantitatively assessed for changes in proliferation, migration and invasive activities using in vitro assays and, for expression of Notch2 or other common stem cell-related genes using quantitative RT-PCR, Western blotting and immunohistochemistry. In vivo effects were evaluated in an orthotopic bladder cancer xenograft model with bioluminescence imaging. RESULTS Cells transduced with the Notch2 shRNA or treated with NRR2Mab demonstrated markedly decreased Notch2 expression. Stable silencing of Notch2 or treatment with NRR2Mab in UMUC3 and 13 diminished cell proliferation under AI but not AH conditions. Likewise, when grown in AI conditions, UMUC3 and 13 were found to be enriched for expression of stem cell genes but Notch2 silencing or NRR2Mab diminished the expression of these genes. Coincidentally, Notch silencing and NRR2Mab treatment significantly inhibited cell migration and invasion of these cells. In vivo, stable Notch2 knockdown cells significantly inhibited xenograft growth for all 3 cell lines. NRR2Mab treatment of host mice also inhibited UMUC13 xenograft growth. CONCLUSIONS Our results provide preclinical evidence that Notch2 may be a useful target to block aggressive behaviors and to prevent growth and/or progression of bladder cancer. Notch2 inhibition with a specific inhibitory monoclonal antibody is practical and warrants further investigation as a novel treatment strategy for bladder cancer. © 2013 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 189Issue 4SApril 2013Page: e243 Advertisement Copyright & Permissions© 2013 by American Urological Association Education and Research, Inc.MetricsAuthor Information Tetsutaro Hayashi Vancouver, Canada More articles by this author Kilian M. Gust Vancouver, Canada More articles by this author Wolfgang Jaeger Vancouver, Canada More articles by this author Shannon Awrey Vancouver, Canada More articles by this author Na Li Vancouver, Canada More articles by this author Manuel Altamirano-Dimas Vancouver, Canada More articles by this author Ralph Buttyan Vancouver, Canada More articles by this author Estelle Li Vancouver, Canada More articles by this author Ladan Fazli Vancouver, Canada More articles by this author Peter C. Black Vancouver, Canada More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Understanding the functions of tumor stroma in resistance to ionizing radiation: Emerging targets for pharmacological modulation

Maintenance of both normal epithelial tissues and their malignant counterparts is supported by the host tissue stroma. The tumor stroma mainly consists of the basement membrane, fibroblasts, extracellular matrix, immune cells, and vasculature. Although most host cells in the stroma possess certain tumor-suppressing abilities, the stroma will change during malignancy and eventually promote growth, invasion, and metastasis. There is growing evidence that the stroma influences importantly the response to radiation therapy (RT). On the one hand, irradiation releases numerous inflammatory cytokines within the extracellular matrix and activates tumor specific antigens presentation, triggering an immune reaction that contributes to the antitumor effect seen after RT. On the other hand, the stroma significantly contributes to radioresistance but also increases the metastatic risk. Indeed, fibroblasts, which are major actors of the impact of stroma on tumor response, are involved in activation of autocrine and paracrine molecular signaling pathways regulating tumor cell proliferation, cell death, response to hypoxia, DNA repair systems and mesenchymal-epithelial transition. They are also actors of the peritumoral desmoplastic reaction, which decreases tumor radiosensitivity. The irradiated stroma can also contribute to tumor relapse after RT through recruitment of bone marrow-derived progenitors that contribute to local tumor relapse through neovascularization. It is therefore time to question preclinical models that would not take into account this impact of stroma. The increasing knowledge of the relationship between stroma and response to IR could help developing innovative strategies for potentially improve antitumor effect of RT.

PIKfyve Sensitivity of hERG Channels

Background/Aims: Human ether-a-go-go (hERG) channels contribute to cardiac repolarization and participate in the regulation of tumor cell proliferation. Mutations in hERG channels may cause long QT syndrome and sudden cardiac death due to ventricular arrhythmias. HERG channel activity is up-regulated by the serum- and glucocorticoid-inducible kinase isoforms SGK1 and SGK3. Related kinases are protein kinase B (PKB/Akt) isoforms. SGK´s and PKB/Akt´s activate phosphatidylinositol-3-phosphate-5-kinase PIKfyve, which in turn up-regulates several carriers and channels. An effect of PIKfyve on hERG channels, has, however, never been shown. The present study thus explored the putative influence of PIKfyve on hERG channel expression and activity. Methods: hERG channels were expressed in Xenopus oocytes with or without PIKfyve and/or PKB, expression of endogenous and injected hERG quantified by RT-PCR, and hERG channel activity determined utilizing dual electrode voltage clamp. Moreover, hERG protein abundance in the cell membrane was visualized utilizing specific antibody binding and subsequent confocal microscopy and quantified by chemiluminescence. Results: Coexpression of wild type PIKfyve increased hERG channel activity in hERG-expressing Xenopus oocytes. hERG channel activity was further increased by coexpression of PKB, an effect augmented by additional coexpression of PIKfyve, but not by additional coexpression of PKB/Akt-resistant PIKfyve mutant PIKfyve^S318A. Coexpression of PIKfyve increased hERG channel protein abundance in the cell membrane. Inhibition of hERG channel insertion into the cell membrane by Brefeldin A (5 µM) resulted in a decline of current, which was similar in Xenopus oocytes expressing hERG together with PIKfyve and in Xenopus oocytes expressing hERG alone. Conclusion: hERG is up-regulated by PIKfyve, which is in turn activated by PKB/Akt.

Association between CASP-8 gene polymorphisms and cancer risk in some Asian population based on a HuGE review and meta-analysis

Genetic variation in the CASP-8 gene reportedly can increase cancer susceptibility by regulating tumor cell proliferation and apoptosis. Several studies have investigated this possibility; however, the conclusions remain controversial. We made a Human Genome Epidemiology (HuGE) review and did a meta-analysis to explore the association between CASP-8 gene polymorphisms and cancer risk in Asian populations. Based on the inclusion criteria, 12 case-control studies comprising 7720 cancer cases and 9404 healthy controls were retrieved. Meta-analysis results showed that the rs3834129*del allele/carrier were associated with decreased risk of cancer in Asian populations [del allele: odd ratio (OR) = 0.79, 95% confidence interval (95%CI) = 0.75-0.83, P < 0.001; del carrier: OR = 0.77, 95%CI = 0.72-0.82, P < 0.001]. Subgroup analysis showed that the rs3834129*del allele/carrier are protective factors for cancer risk in Chinese populations (del allele: OR = 0.77, 95%CI = 0.73-0.81, P < 0.001; del carrier: OR = 0.75, 95%CI = 0.70-0.80, P < 0.001), but not in Indian and Korean populations. Furthermore, the rs6704688*T allele/carrier, rs3769827*C allele/carrier, rs3769825*C allele/carrier were associated with decreased risk of cancer in Asian populations (all P < 0.05). While the rs7608692*A allele was associated with increased risk of cancer risk in Asian populations (OR = 1.35, 95%CI = 1.02-1.78, P = 0.03). There was also no significant association between rs3769818, rs13030042, rs13030042, rs1045494, rs1045494, rs2823, or rs113686495, and cancer risk in Asian populations (all P > 0.05). This meta-analysis suggests that the rs3834129*del allele/ carrier, rs6704688*T allele/carrier, rs3769827*C allele/carrier, and rs3769825*C allele/carrier might be protective factors for cancer risk in Asian populations, while the rs7608692*A allele might be a risk factor for cancer risk in Asian populations.

Sphingosine kinase 1 promotes tumor progression and confers malignancy phenotypes of colon cancer by regulating the focal adhesion kinase pathway and adhesion molecules

Studies suggest a tumor-promoting function of sphingosine kinase 1 (SphK1) in some types of human tumors, however, its effect on colon cancer is still unclear. The aims of this study were to investigate the roles of SphK1 in the progression and tumor cell phenotypic changes in colon cancer. Moreover, the focal adhesion kinase (FAK) pathway and the expression of intercellular adhesion molecule‑1 (ICAM‑1) and vascular cell adhesion molecule‑1 (VCAM‑1) were detected to explore the mechanisms of SphK1 action. In this study, the expression of SphK1, FAK and phospho-FAK (p-FAK) was analyzed in 66 surgical specimens of primary colon cancer and matched adjacent normal tissues by immunohistochemistry and western blotting. In addition, N,N-dimethylsphingosine (DMS), SphK1 DNA and shRNA transfection were used to regulate the expression and activity of SphK1 in the LOVO colon cancer cell line. Tumor cell phenotypic changes were analyzed by cell viability, invasion and apoptosis assays. Results showed that the expression of SphK1, FAK and p-FAK in colon cancer tissues were significantly stronger compared to those in matched normal tissues. There was a close correlation between the expression of SphK1 and FAK or p-FAK and the co-expression of SphK1, FAK and p-FAK significantly associated with histological grade, Dukes' stage, lymph node metastasis and distant metastasis. Overexpression of SphK1 after DNA transfection enhanced tumor cell viability and invasiveness, but suppressed cell apoptosis. In contrast, suppression of SphK1 by DMS and shRNA reduced tumor cell viability and invasiveness, but promoted cell apoptosis. The expression of FAK, p-FAK, ICAM-1 and VCAM-1 in LOVO cells were increased with the overexpression of SphK1 but decreased with the suppression of SphK1. These findings indicate that SphK1 regulates tumor cell proliferation, apoptosis and invasion, which ultimately contributes to tumor progression and malignancy phenotype in colon cancer. FAK pathway, ICAM-1 and VCAM-1 may play critical roles in this SphK1‑mediated effect.

MTOR signalling pathway and gastric carcinoma

Aberrations of the mammalian target of rapamycin (mTOR) signaling are frequently observed in many types of cancer.Activation of mTOR regulates tumor cell proliferation,survival and metastasis,and then results tumorigenesis and tumor developmet.Blocking the mTOR signaling pathway by rapamycin and its derivatives can inhibit gastric cancer cell growth and promote tumor necrosis and such effects can be synergistically improved by combined use of other chemotherapeutic agents.Thus,rapamyein and its derivatives may be effective in the prevention and treatment of gastric cancer. Key words: Stomach neoplasms; Sirolimus; Mammalian target of rapamycin

Decreased melanoma proliferation and cell survival: turn down your SOX10

Decreased melanoma proliferation and cell survival: turn down your <scp>SOX</scp>10

Carbon Monoxide Mediates the Anti-apoptotic Effects of Heme Oxygenase-1 in Medulloblastoma DAOY Cells via K+ Channel Inhibition

Tumor cell survival and proliferation is attributable in part to suppression of apoptotic pathways, yet the mechanisms by which cancer cells resist apoptosis are not fully understood. Many cancer cells constitutively express heme oxygenase-1 (HO-1), which catabolizes heme to generate biliverdin, Fe(2+), and carbon monoxide (CO). These breakdown products may play a role in the ability of cancer cells to suppress apoptotic signals. K(+) channels also play a crucial role in apoptosis, permitting K(+) efflux which is required to initiate caspase activation. Here, we demonstrate that HO-1 is constitutively expressed in human medulloblastoma tissue, and can be induced in the medulloblastoma cell line DAOY either chemically or by hypoxia. Induction of HO-1 markedly increases the resistance of DAOY cells to oxidant-induced apoptosis. This effect was mimicked by exogenous application of the heme degradation product CO. Furthermore we demonstrate the presence of the pro-apoptotic K(+) channel, Kv2.1, in both human medulloblastoma tissue and DAOY cells. CO inhibited the voltage-gated K(+) currents in DAOY cells, and largely reversed the oxidant-induced increase in K(+) channel activity. p38 MAPK inhibition prevented the oxidant-induced increase of K(+) channel activity in DAOY cells, and enhanced their resistance to apoptosis. Our findings suggest that CO-mediated inhibition of K(+) channels represents an important mechanism by which HO-1 can increase the resistance to apoptosis of medulloblastoma cells, and support the idea that HO-1 inhibition may enhance the effectiveness of current chemo- and radiotherapies.

Abstract 1052: Synergy between inhibition of novel histone demethylase (LSD2) and DNA methyltransferase (DNMT) and histone deacetylase (HDAC) in modulating gene expression and inhibiting growth in human breast cancer cells

Abstract Epigenetic gene silencing plays a critical role in breast tumor aggression. Our previous studies have shown that abnormal activities of histone lysine demethylases (KDMs) are associated with aberrant gene expression in breast cancer development. However, the precise molecular mechanisms underlying the activity of KDMs in chromatin remodeling and regulation of gene transcription in breast cancer are still elusive. Recently, a novel mammalian histone demethylase LSD2 (also known as KDM1B or AOF1) was identified. We have shown that LSD2 possesses the activity to demethylate H3K4 in breast cancer cells. This clearly suggests the existence of a more sophisticated FAD-dependent histone demethylase family whose members play a role in chromatin remodeling and transcription regulation in breast cancer. Here we demonstrated that inhibition of LSD2 mRNA expression by shRNA suppressed migratory behavior in human breast cancer MDA-MB-231 cells. RNAi -mediated LSD2 deficiency led to a significant re-expression of ERα mRNA, in ER negative breast cancer cells and upregulated ER-mediated transcription activity as evidenced by ERE luciferase assay. Microarray studies indicated a unique subset of genes whose expression was significantly changed by LSD2 KD in MDA-MB-231 cells. The genes identified are extensively involved in regulation of tumor cell proliferation, metastasis, cell signaling, transcription regulation, and chromatin remodeling. For example, a panel of the upregulated genes potentially plays a role in cell adhesion and cell death such as CLDN1 &amp; 11, CDH11, CASP5 &amp; 10, PDCD4, and NCR3. In addition, expression of several tumor suppressor genes such as ERBB2IP, MTSS1, and S100A2 were up-regulated by LSD2 KD in MDA-MB-231 cells. Moreover, treating LSD2 KD cells with the low doses of DNMT inhibitor, 5-aza-2-deoxycitidine (DAC) led to a robust reexpression of ERα and CASP5 mRNA in MDA-MB-231 cells. Treatment with the HDAC inhibitor vorinostat in LSD2 KD MDA-MB-231 cells also lead to the reexpression of ERα as well as other genes of interest such as CASP5, ESRRG, and MTSS1. Further simultaneous treatment of LSD2 KD cells with DAC and vorinostat produced a more robust transcriptional re-activation of ERα, CASP5, and MTSS1 mRNA expression. Finally, LSD2 KD MDA-MB-231 cells exhibited increased sensitivity to growth inhibition induced by combination therapy of vorinostat and DAC. Taken together, these data implicate an important role for LSD2 in mediating expression of epigenetically silenced genes in breast cancer cells and suggest that combined inhibition of LSD2 with other epigenetic targets could have the potential to improve the treatment of breast cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1052. doi:1538-7445.AM2012-1052

Abstract 4613: A role of integrin-associated CD151 in human ovarian cancer

Abstract Despite being implicated in multiple types of human epithelia-origin cancer, laminin-binding (LB) integrins (e.g., α3α1 and α6β4) have rarely been investigated for their roles in human ovarian tumors. More recently, we and others have shown that CD151, a member of the tetraspanin family, not only forms tight surface complexes with these adhesion receptors, but also mediates the malignancy of human carcinomas largely in a LB integrin-dependent manner. Here we report our studies on the role of CD151 in human ovarian cancer. Initially, we stained human ovarian tumor tissue microarrays with CD151-specific antibody. Our data showed that the majority of ovarian tumors exhibited a reduced expression of CD151 protein, compared to the fallopian tube, implying a putative suppressing role of this tetraspanin in ovarian cancer. With this hint we next evaluated the impact of CD151 ablation on the behaviors and proliferation of cultured human ovarian cancer cells. While CD151 removal had a minimal impact on cell motility or invasion, it markedly enhanced cell proliferation. Also, CD151-deficeint cells appeared to have reduced cell-cell contacts, suggestive of epithelial-mesenchymal transition (EMT)-like changes. However, there was no dramatic difference in the surface expression of E-cadherin protein, according to our FACS analyses. To substantiate these observations, we also injected ovarian cancer cells with or without CD151 knockdown into immuno-deficient nude mice subcutaneously or orthotopically (intra-peritoneal). Results from these ex vivo analyses showed that disruption of CD151 accelerated ovarian tumor growth and production of ascites fluid in mice, further supporting a role of CD151 in ovarian tumor progression. While tumors in the control group had homogenous epithelial-like appearance, those derived from CD151-ablated cells exhibited mucin-related vacuoles and diverse morphologies. Our subsequent DNA array and protein analyses indicated that there was indeed a marked increase in the expression of a mucin family member in CD151-deficient tumor cells. Collectively, these results provide strong evidence that CD151 suppresses human ovarian cancer progression by regulating tumor cell proliferation and extracellular microenvironment. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4613. doi:1538-7445.AM2012-4613

Abstract 2143: Smg-GDS regulates cellular proliferation in squamous cell carcinomas of the head and neck

Abstract Squamous cell carcinoma of the head and neck (SCCHN) is a major worldwide source of morbidity and mortality, with over 26,000 estimated new cases and 6,000 deaths from pharyngeal and laryngeal cancer in 2011 in the the United States alone. SCCHN is an aggressive disease which is characterized by local invasion and lymphatic and distant metastasis. Ras and Rho family GTPases are known to regulate tumor cell proliferation in numerous human cancers including SCCHN. SmgGDS, an unusual guanine nucleotide exchange factor, has previously been shown in other systems to activate these small GTPases and directly regulate cellular proliferation through control of small GTPase prenylation and cellular localization. Little is known about the role of SmgGDS in the progression of SCCHN. These studies were begun to characterize the expression of SmgGDS in SCCHN and assess the role that SmgGDS plays in SCCHN proliferation. SCCHN tumor tissue microarray analysis demonstrated increased SmgGDS expression in tumor tissue from various head and neck sites compared to normal tissues, with evidence for higher SmgGDS expression at the pushing tumor boarders in invasive cancers. Significantly increased amounts of SmgGDS protein and RNA expression were also noted in head and neck tumor cell lines compared to cultured normal head and neck tissue cell lines. To assess the function of Smg-GDS expression in SCCHN, UM-SCC-1 cells were transfected with siRNA directed toward the 60.7 and 55.8 kD SmgGDS isoforms, with significant decreases in S-phase distribution for all treatments. Blocking both 60.7 and 55.8 kD SmgGDS isoforms with siRNA induced an 89% decrease in S-phase distribution as determined by Click-iT Edu cell cycle proliferation assay. These studies confirm an active role for SmgGDS protein function in the promotion of cancer cell proliferation in SCCHN, and provide the basis for further investigations into the role of small GTPases in head and neck neoplasia. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2143. doi:1538-7445.AM2012-2143

Abstract 1300: Far upstream element binding proteins (FBP)-1 and FBP-2 in non-small cell lung cancer: Key factors regulating tumor cell proliferation, migration, and invasion

Abstract The far upstream element (FUSE)-binding proteins (FBP)- 1, FBP-2, and FBP-3 represent a family of single-strand nucleic acid binding factors, regulating transcriptional as well as post-transcriptional processes. Elevated expression and pro-tumorigenic functions of all FBPs have been described for human liver cancer [1, 2]. First data indicated that FBP-1 affects microtubule dynamics through regulating MT-destabilizing factors in non-small cell lung cancer (NSCLC) [3], however, comprehensive studies analyzing the expression and functional relevance of all FBPs in NSCLCs are missing so far. In order to define the expression of FBPs in lung cancer, FBP-transcript and protein levels were analysed in primary human NSCLC tissue samples. Semiquantitative real-time PCR as well as Tissue-Micro-Array (TMA) analyses revealed an elevated expression of FBP-1 and FBP-2 in most NSCLC tissue samples (&amp;gt;60%) in comparison to non-tumorous specimens. Interestingly, nuclear accumulation of FBP-1 significantly correlated with FBP-2 expression (r=0.33; p&amp;lt;0.01), suggesting common regulatory mechanisms. In vitro, transient inhibition of FBP-1 by gene-specific siRNAs in NSCLC cell lines (Calu-6) was associated with decreased tumor cell viability (-76%; MTT assay), proliferation (-83%; BrdU assay), and increased apoptosis (2.8-fold; Nicoletti FACS assay). In contrast, inhibition of FBP-2 reduced cell viability of Calu-1 cells (-32%; MTT assay) but predominantly reduced tumor cell migration (-62%; two-dimensional scratch assay) as well as tumor cell invasion (-81%; sprouting assay) in all analyzed NSCLC cell lines (Calu-1, Calu-6, and A549), suggesting that both FBP isoforms facilitate partly distinct tumor-supporting effects. Surprisingly, efficient single-gene inhibition of FBP-1 or FBP-2 in A549 cells did not affect tumor cell viability. In contrast, the concomitant knock down of both FBP isoforms resulted in significantly decreased cell viability (-69%), suggesting that some FBP family members may compensate the loss of other members. Actually, FBP-2 negatively regulates FBP-1 expression in A549 cells, resulting in increased FBP-1 transcript and protein levels after FBP-2 inhibition. Therefore, functional compensation prevents A549 cells from anti-tumorigenic effects after FBP-2 knock down. In summary, this study provides evidence that coordinated overexpression of FBP-1 and FBP-2 is a frequent event in NSCLCs and that both factors are support tumor growth and NSCLC cell dissemination. Interestingly, partial functional redundance and mutual negative regulation of FBPs indicate that these factors may fine-tune the oncogenic behavior of NSCLC cells.

Targeted inhibition of tumor proliferation, survival, and metastasis by pentacyclic triterpenoids: Potential role in prevention and therapy of cancer

Over the last two decades, extensive research on plant-based medicinal compounds has revealed exciting and important pharmacological properties and activities of triterpenoids. Fruits, vegetables, cereals, pulses, herbs and medicinal plants are all considered to be biological sources of these triterpenoids, which have attracted great attention especially for their potent anti-inflammatory and anti-cancer activities. Published reports in the past have described the molecular mechanism(s) underlying the various biological activities of triterpenoids which range from inhibition of acute and chronic inflammation, inhibition of tumor cell proliferation, induction of apoptosis, suppression of angiogenesis and metastasis. However systematic analysis of various pharmacological properties of these important classes of compounds has not been done. In this review, we describe in detail the pre-clinical chemopreventive and therapeutic properties of selected triterpenoids that inhibit multiple intracellular signaling molecules and transcription factors involved in the initiation, progression and promotion of various cancers. Molecular targets modulated by these triterpenoids comprise, cytokines, chemokines, reactive oxygen intermediates, oncogenes, inflammatory enzymes such as COX-2, 5-LOX and MMPs, anti-apoptotic proteins, transcription factors such as NF-κB, STAT3, AP-1, CREB, and Nrf2 (nuclear factor erythroid 2-related factor) that regulate tumor cell proliferation, transformation, survival, invasion, angiogenesis, metastasis, chemoresistance and radioresistance. Finally, this review also analyzes the potential role of novel synthetic triterpenoids identified recently which mimic natural triterpenoids in physical and chemical properties and are moving rapidly from bench to bedside research.

Opposing function of MYBBP1A in proliferation and migration of head and neck squamous cell carcinoma cells

BackgroundHead and neck squamous cell carcinoma (HNSCC) is one of the most prevalent and lethal cancers worldwide and mortality mostly results from loco-regional recurrence and metastasis. Despite its significance, our knowledge on molecular, cellular and environmental mechanisms that drive disease pathogenesis remains largely elusive, and there are limited therapeutic options, with only negligible clinical benefit.MethodsWe applied global gene expression profiling with samples derived from a recently established mouse model for oral cancer recurrence and identified a list of genes with differential expression between primary and recurrent tumors.ResultsOne differentially expressed gene codes for Myb-binding protein 1a (MYBBP1A), which is known as a transcriptional co-regulator that physically interacts with nuclear transcription factors, such as NFκB and p53. We confirmed significantly reduced MYBBP1A protein levels on tissue sections of recurrent mouse tumors compared to primary tumors by immunohistochemistry, and found aberrant MYBBP1A protein levels also in tumor samples of HNSCC patients. Interestingly, silencing of MYBBP1A expression in murine SCC7 and in human HNSCC cell lines elicited increased migration but decreased cell growth.ConclusionWe provide experimental evidence that MYBBP1A is an important molecular switch in the regulation of tumor cell proliferation versus migration in HNSCC and it will be a major challenge for the future to proof the concept whether regulation MYBBP1A expression and/or function could serve as a novel option for anti-cancer therapy.

Retinoblastoma tumor-suppressor protein phosphorylation and inactivation depend on direct interaction with Pin1

Inactivation of the retinoblastoma protein (pRb) by phosphorylation triggers uncontrolled cell proliferation. Accordingly, activation of cyclin-dependent kinase (CDK)/cyclin complexes or downregulation of CDK inhibitors appears as a common event in human cancer. Here we show that Pin1 (protein interacting with NIMA (never in mitosis A)-1), a peptidylprolyl isomerase involved in the control of protein phosphorylation, is an essential mediator for inactivation of the pRb. Our results indicate that Pin1 controls cell proliferation by altering pRb phosphorylation without affecting CDK and protein phosphatase 1 and 2 activity. We demonstrated that Pin1 regulates tumor cell proliferation through direct interaction with the spacer domain of the pRb protein, and allows the interaction between CDK/cyclin complexes and pRb in mid/late G1. Phosphorylation of pRb Ser 608/612 is the crucial motif for Pin1 binding. We propose that Pin1 selectively boosts the switch from hypo- to hyper-phosphorylation of pRb in tumor cells. In addition, we demonstrate that the CDK pathway is responsible for the interaction of Pin1 and pRb. Prospectively, our findings therefore suggest that the synergism among CDK and Pin1 inhibitors holds great promise for targeted pharmacological treatment of cancer patients, with the possibility of reaching high effectiveness at tolerated doses.