bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3425: MiR-215-5p is a tumor suppressor in colorectal cancer targeting EGFR ligand epiregulin and its transcriptional inducer HOXB9
Authors: Faltejskova-Vychytilova Petra , Merhautova Jana , Conesa-Zamora Pablo , Slaba Katerina , Machackova Tana , Svoboda Marek , Vecera Marek , Mlcochova Jitka , Juracek Jaroslav , Sana Jiri , Ahmad Parwez , Gablo Natalia , Slaby Ondrej
Published: July 2017

Growing evidence suggests that microRNAs (miRNAs) are involved in the development and progression of colorectal cancer (CRC). In the present study, deregulation and functioning of tumor suppressive miR-215-5p was evaluated in CRC. In total, 448 tumor tissues and 325 paired adjacent healthy tissues have been used for miRNA expression analyses. We proved that miR-215-5p is significantly down-regulated in tumor tissues compared to non-tumor adjacent tissues and its decreased levels correlate with the presence of lymph node metastases, tumor stage, and shorter overall survival in CRC patients. To identify specific cellular processes affected by ectopic expression of miR-215-5p, a series of in vitro experiments have been performed using transient transfection of miR-215-5p mimics into four CRC cell lines. Increased levels of miR-215-5p significantly reduced proliferation, clonogenicity, and migration of CRC cells, lead to cell cycle arrest in G2/M phase and p53-dependent induction of apoptosis. The ability of miR-215-5p to inhibit tumor growth was confirmed in vivo by use of NSG mice model and stable cell line over-expressing miR-215-5p. Finally, we proved epiregulin and HOXB9 to be the direct targets of miR-215-5p by luciferase assay and western blot analyses. Since epiregulin is EGFR ligand and HOXB9, is its transcriptional inducer, we suggest, that the main molecular link between miR-215-5p and CRC cells phenotypes presents the EGFR signaling pathway, which is one of the canonical pathogenic pathways in CRC. This work was supported by Ministry of Health of the Czech Republic, grant nr. 15-33158A, 15-34553A, 15-31627A, 15-34678A, 16-31314A, 16-31765A and by grant of Czech Grant Agency nr. 16-18257S.

Citation Format: Petra Faltejskova-Vychytilova, Jana Merhautova, Pablo Conesa-Zamora, Katerina Slaba, Tana Machackova, Marek Svoboda, Marek Vecera, Jitka Mlcochova, Jaroslav Juracek, Jiri Sana, Parwez Ahmad, Natalia Gablo, Ondrej Slaby. MiR-215-5p is a tumor suppressor in colorectal cancer targeting EGFR ligand epiregulin and its transcriptional inducer HOXB9 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3425. doi:10.1158/1538-7445.AM2017-3425
Abstract 344: Chondroitin sulfatases regulate Wnt signaling through effects on Shp2, phospho-Erk1,2, c-Myc, and histone methylation of DKK3 in prostate cancer and prostate stem cells
Authors: Tobacman Joanne Kramer , Bhattacharyya Sumit , Feferman Leo
Published: July 2017

The chondroitin sulfatases N-acetylgalactosamine 4-sulfatase (arylsulfatase B; ARSB) and N-acetylgalactosamine 6-sulfatase (galactose 6-sulfatase, GALNS) remove the 4-sulfate and 6-sulfate groups of chondroitin sulfates. ARSB acts to remove 4-sulfate groups of chondroitin 4-sulfate (C4S) or dermatan sulfate, and GALNS acts to remove 6-sulfate groups of chondroitin 6-sulfate (C6S) or chondroitin 4,6-disulfate, as well as keratan sulfate. In human prostate cancer tissues, ARSB activity was markedly reduced and GALNS activity increased. By immunohistochemistry of malignant prostate tissue captured by laser microdissection and by assay of human prostate stromal and epithelial cell lines, ARSB was predominant in stromal cells of the extracellular matrix, whereas GALNS predominated in prostate epithelial cells. When ARSB was reduced by silencing in human prostate stem cells or GALNS was increased by overexpression in the stem cells, SHP2 binding to chondroitin 4-sulfate increased, leading to inhibition of phosphatase action and sustained phosphorylation of Erk1,2. Erk activation enabled increase in c-Myc binding to nuclear DNA and increased expression of DNMT 1a, 3a, and 3b. DNMT activation increased following ARSB silencing or GALNS overexpression, and was blocked by ERK inhibition. Increased DNMT activity led to the increased promoter methylation of DKK3, and the subsequent inhibition of nuclear β-catenin nuclear translocation and Wnt signaling. Decline in Wnt signaling was manifested by reduced TCF-LEF nuclear binding, and decline in the mRNA expression of c-Myc and GATA. Since c-Myc activation can lead to increased expression of cell cycle regulators, the changes in chondroitin sulfation mediated by sulfatases can lead to profound effects in cell proliferation. Hence, the modification of sulfatase activity, manifested as increased chondroitin 4-sulfate, provides the platform for an extensive extracellular-intracellular signaling network that can regulate phosphorylation, DNA promoter methylation, and proliferation, and affect development and malignant transformation.

Citation Format: Joanne Kramer Tobacman, Sumit Bhattacharyya, Leo Feferman. Chondroitin sulfatases regulate Wnt signaling through effects on Shp2, phospho-Erk1,2, c-Myc, and histone methylation of DKK3 in prostate cancer and prostate stem cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 344. doi:10.1158/1538-7445.AM2017-344
Abstract 3443: Functional and mechanistic characterization of oncogenic long intervening non-coding RNA Linc-CHD1L-1 in hepatocellular carcinoma
Authors: Tsang Felice H. , Ho Daniel W. , Ng Irene O. , Wong Jack C.
Published: July 2017

Hepatocellular carcinoma (HCC) is one of the most common and lethal cancers worldwide. Due to the asymptomatic nature of HCC, many patients were diagnosed at advance stages and available therapeutic options are very limited. Therefore, a better understanding of the molecular mechanism underlying hepatocarcinogenesis is urgently needed and is crucial for the development of effective treatment. Long intervening non-coding RNAs (lincRNAs) are emerging class of regulatory RNA involved in diverse molecular processes. In this study, we aimed to interrogate lincRNAs expression in HCC and identify novel lincRNAs that contribute to hepatocarcinogenesis. To investigate the global expression profile of lincRNAs in human HCCs, next-generation transcriptome sequencing (RNA-Seq) was performed in 16 pairs of HBV-associated primary HCCs and their corresponding non-tumorous liver tissues. Interestingly, the lincRNA expressions in human HCCs can be unambiguously segregated from their non-tumorous counterparts by unsupervised clustering analysis, suggesting that global lincRNA expression profiles between tumors and non-tumors were remarkably different. From this analysis, we identified linc-CHD1L-1 as the most significant up-regulated lincRNA in HCCs. Clinically, linc-CHD1L-1 was overexpressed in over 80% of HCC patients. Linc-CHD1L-1 expression level in tumors was also shown to be a promising marker for HCC diagnosis. Functionally, knockdown of linc-CHD1L-1 by shRNAs had significantly reduced HCC cell proliferation and cell migration in vitro. Besides, knockdown of linc-CHD1L-1 in HCC cells also substantially abrogated tumorigenicity in subcutaneous tumor model. Mechanistically, the expression of linc-CHD1L-1 was significantly and positively correlated to the expression of its neighboring genes located on the chromosome 1q. Knockout of linc-CHD1L-1 by CRISPR/Cas9 also significantly repressed its neighboring genes expression, suggesting that linc-CHD1L-1 may positively regulate gene expression in a cis-manner. Finally, DAVID pathway analysis performed on linc-CHD1L co-expressed protein coding genes further suggested that linc-CHD1L-1 was significantly co-expressed with genes that are involved in transcription and chromatin modifications. In conclusion, overexpression of linc-CHD1L-1 contributes to hepatocarcinogenesis by promoting HCC cell proliferation and migration through transcriptional and epigenetic mechanisms.

Citation Format: Felice H. Tsang, Daniel W. Ho, Irene O. Ng, Jack C. Wong. Functional and mechanistic characterization of oncogenic long intervening non-coding RNA Linc-CHD1L-1 in hepatocellular carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3443. doi:10.1158/1538-7445.AM2017-3443
Abstract 3444: LINC00152 regulates cell proliferation via p38 signaling and overexpression predicts poor patient survival in lung cancer
Authors: Feng Shumei , Zhang Jie , Su Wenmei , Bai Shengbin , Xiao Lei , Wang Zhuwen , Lin Jules , Reddy Rishindra , Chang Andrew , Beer David , Chen Guoan
Published: July 2017

Lung cancer is a molecularly-heterogenous disease and the leading cause of cancer mortality. The molecular basis for this heterogeneity remains incompletely understood. In the past few years, long non-coding RNAs (lncRNAs) have emerged as novel mechanisms in mediating cancer biology, although most lncRNAs are still undiscovered. LINC00152 has been identified as highly associated with the tumorigenesis and development in gastric cancer, colon cancer and hepatocellular carcinoma, however, the expression level and its oncogenic roles in lung cancer remains unknown. In the present study, we employed next generation RNA sequencing analysis to reveal dysregulated lncRNAs in lung cancer utilizing datasets of 461 lung adenocarcinomas (LUAD) and 156 normal lung tissues from 3 separate institutions. We found that LINC00152 was 4-fold (p <0.001) overexpressed in lung tumors as compared to their adjacent normal tissues with AUC > 0.8 in these 3 data sets. Patients with high LINC00152 expression have significantly poorer survival than those with low expression (log-rank test, p = 0.003). We verified this diagnostic/prognostic potential in an independent cohort of lung tumor tissues by quantitative RT-PCR. Cell proliferation and colony formation ability were decreased after knockdown of LINC00152 using siRNAs in lung cancer cell lines. The expression of LINC00152 was found primarily in the cytoplasm by qRT-PCR analysis. Trichostatin A treatment indicated that histone acetylation could be one of the mechanisms underlying LINC00152 overexpression in NSCLC and cell-based analyses showed p38 signaling was mainly affected by LINC00152 in vitro. Taken together, our study suggests that LINC00152 is involved in lung tumor growth, may have potential as diagnostic/prognostic marker and that further characterization of this lncRNA as a novel therapeutic target for lung cancer is warranted.

Note: This abstract was not presented at the meeting.

Citation Format: Shumei Feng, Jie Zhang, Wenmei Su, Shengbin Bai, Lei Xiao, Zhuwen Wang, Jules Lin, Rishindra Reddy, Andrew Chang, David Beer, Guoan Chen. LINC00152 regulates cell proliferation via p38 signaling and overexpression predicts poor patient survival in lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3444. doi:10.1158/1538-7445.AM2017-3444
Abstract 3447: Down-regulation of long non-coding RNA MALL-2 is associated with metastasis and poor survival in lung adenocarcinoma
Authors: Dang Ha X. , White Nicole M. , Rozycki Emily B. , Maher Christopher A.
Published: July 2017

Lung adenocarcinoma accounts for ~40% of lung cancers and is among the deadliest cancers worldwide. Patients with metastatic lung adenocarcinoma exhibit poor outcome with the average 5-year survival of ~1%. Long non-coding RNAs (lncRNAs) have recently emerged as having critical roles in tumorigenesis. While a subset of lncRNAs have been shown to promote tumor invasion and metastasis, little is known about the role of lncRNAs in metastatic lung adenocarcinoma. Therefore, we analyzed publicly available microarray and RNA-Seq gene expression datasets that include primary and metastatic tissues. Through this meta-analysis we discovered 98 metastasis associated lncRNAs that were dysregulated in metastatic tumors compared to non-metastatic tumors (MALLs). Notably, only two lncRNAs (MALL-2 and MALL-36) were found to be down-regulated in primary tumors, relative to normal tissue, and even further down-regulated in metastatic tumors relative to primary tumors. Gene set enrichment analysis revealed that MALL-2, the most down-regulated lncRNA in metastasis, was coexpressed with protein-coding genes enriched with biological concepts associated with repressing tumor growth and metastasis. Furthermore, we found that FOXA2, a metastasis repressor gene, was highly coexpressed with MALL-2 in lung cancer. A pan-cancer analysis using ~7,000 TCGA RNA-Seq tumor samples across 20 cancer types revealed that MALL-2 was also down-regulated and coexpressed with FOXA2 in multiple cancers suggesting the conserved regulatory relationship between the two genes. We confirmed that FOXA2 expression was decreased upon knock-down of MALL-2 and silencing MALL-2 resulted in a significant increase in cellular migration in lung adenocarcinoma cell lines. Last, we found that low expression of MALL-2 was consistently associated with poor overall survival across multiple independent lung adenocarcinoma cohorts. Taken together, our integrative analysis has revealed MALL-2 that acts as a repressor of lung adenocarcinoma metastasis by regulating FOXA2 and could serve as a biomarker of lung adenocarcinoma patient outcome.

Citation Format: Ha X. Dang, Nicole M. White, Emily B. Rozycki, Christopher A. Maher. Down-regulation of long non-coding RNA MALL-2 is associated with metastasis and poor survival in lung adenocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3447. doi:10.1158/1538-7445.AM2017-3447
Abstract 3449: Genistein inhibits renal cancer progression through long non-coding RNA HOTAIR suppression
Authors: Imai-Sumida Mitsuho , Majid Shahana , Dasgupta Pritha , Kulkarni Priyanka , Saini Sharanjot , Bhagirath Divya , Kato Taku , Maekawa Shigekatsu , Hashimoto Yutaka , Shiina Marisa , Deng Guoren , Shahryari Varahram , Tanaka Yuichiro , Dahiya Rajvir , Yamamura Soichiro
Published: July 2017

Genistein, a soy isoflavone, has been shown to have anticancer effects on various cancers in vitro and in vivo including renal cancer. Long non-coding RNAs (lncRNAs) are differentially expressed in various tissues and have important functions in cellular processes such as cell proliferation, motility and apoptosis in various malignancies. HOX transcript antisense RNA (HOTAIR) is a lncRNA localized in the Homeobox C gene cluster on chromosome 12. HOTAIR interacts with the polycomb repressive complex 2 (PRC2), which enhances H3K27 trimethylation and represses the expression of tumor suppressors. In various cancers, HOTAIR is highly expressed and involved in their progression and metastasis. In this study, we investigated the molecular mechanisms of genistein action through a novel pathway that represses HOTAIR. We found that HOTAIR expression is higher in renal cancer cell lines compared to normal controls. Genistein treatment was found to significantly decrease HOTAIR expression in renal cancer cells (786-O and ACHN cells). Genistein treatment also reduced expression of epithelial-to-mesenchyme transition (EMT)-related proteins (ZEB1, Vimentin and Snail), causing reduced cell migration, invasion, and increased apoptosis. We performed RNA immunoprecipitation assays, and found that genistein inhibits HOTAIR binding to PRC2. One of the other EMT markers, a tight junction protein ZO-1, is upregulated by genistein. We are currently investigating if genistein represses PRC2 recruitment to the ZO-1 promoter by inhibiting binding of HOTAIR to PRC2. Our results indicate that genistein is a potent therapeutic agent for renal cancer.

Citation Format: Mitsuho Imai-Sumida, Shahana Majid, Pritha Dasgupta, Priyanka Kulkarni, Sharanjot Saini, Divya Bhagirath, Taku Kato, Shigekatsu Maekawa, Yutaka Hashimoto, Marisa Shiina, Guoren Deng, Varahram Shahryari, Yuichiro Tanaka, Rajvir Dahiya, Soichiro Yamamura. Genistein inhibits renal cancer progression through long non-coding RNA HOTAIR suppression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3449. doi:10.1158/1538-7445.AM2017-3449
Abstract 345: PTEN tumor suppressive effects in glioblastoma are partly mediated by MET downregulation via a p53-miR-34a axis
Authors: Guessous Fadilla , Cruickshanks Nichola A. , Zhang Ying , Schiff David , Sarkaria Jann , Abounader Roger
Published: July 2017

Glioblastoma (GBM) is the most common and most lethal primary malignant brain tumor. Two common alterations in GBM are loss of the tumor suppressor PTEN and upregulation of the oncogenic receptor tyrosine kinase MET. We uncovered a new connection between PTEN and MET expressions and functions in GBM. We found that PTEN expression in GBM cell lines (U87, U373, A172, LNZ-308) and primary cells (GBM8) inhibits MET expression. We uncovered the molecular basis and functional significance of this connection. Based on our previous work and published literature, PTEN stabilizes p53, p53 induces miR-34a expression, and miR-34a inhibits MET expression. We therefore hypothesized that PTEN inhibits MET by regulating p53 and miR-34a and that the PTEN tumor suppressive effects are partly mediated by MET downregulation. To verify the hypothesis, we first showed that PTEN restoration leads to miR-34a upregulation in GBM cells. We then showed that this latter effect is p53-dependent as p53 knockdown with a specific shRNA partly reversed PTEN induction of miR-34a. We also showed that PTEN-induced downregulation of MET expression can be rescued by inhibition of miR-34a with an anti-miR3a. We therefore established a new molecular cascade that links PTEN, p53, miR-34a and MET in GBM. To determine the functional relevance of this cascade, we assessed the effects of PTEN on GBM cell death and invasion also in the settings of shRNA-mediated knockdown of p53 and anti-miR-mediated inhibition of miR-34a. PTEN strongly induced GBM cell death (measured by Annexin V/7AAD flow cytometry) and inhibited transwell invasion though a collagen IV matrix. The PTEN effects on cell death and invasion were partly reduced by p53 knock-down and miR-34a inhibition. This demonstrates that the tumor suppressive effects of PTEN in GBM are partly mediated by the PTEN/p53/miR-34a/MET cascade. In ongoing experiments, we are investigating the expressions of PTEN, MET, p53 and miR-34a in human GBM tissues to determine if there are potential correlations that are consistent with our above findings. Altogether, this study establishes a new molecular and functional link between PTEN and MET that involves p53 and miR-34a in GBM.

Citation Format: Fadilla Guessous, Nichola A. Cruickshanks, Ying Zhang, David Schiff, Jann Sarkaria, Roger Abounader. PTEN tumor suppressive effects in glioblastoma are partly mediated by MET downregulation via a p53-miR-34a axis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 345. doi:10.1158/1538-7445.AM2017-345
Abstract 3450: Identification of a novel lncRNA involved in Bcr-Abl mediated cellular transformation
Authors: Chen Jilong , Wang Xuefei
Published: July 2017

Aberrant expression of long noncoding RNAs (lncRNAs) is associated with various human cancers. However, little information is available about functional involvement of lncRNAs in tumorigenesis of chronic myeloid leukemia (CML) induced by Bcr-Abl oncogene. In this study, we performed a comprehensive analysis of lncRNAs in human CML cells using an lncRNA cDNA microarray combined with drug treatment. We identified an lncRNA termed lncRNA-up38 that was critically involved in Bcr-Abl-mediated cellular transformation. We found that expression of lncRNA-up38 was greatly upregulated by inhibiting Bcr-Abl kinase activity in K562 cells and leukemic cells derived from CML patients. Furthermore, our experiments demonstrated that lncRNA-up38 acted as a key regulator of 4EBP1 functioning during Bcr-Abl-mediated cellular transformation. Moreover, silencing lncRNA-up38 promoted leukemic cells survival and enhanced Bcr-Abl-mediated tumorigenesis, whereas forced expression of lncRNA-up38 sensitized leukemic cells to undergo apoptosis and inhibited Bcr-Abl-induced tumorigenesis. Additionally, our experiments have begun to address the mechanism of how lncRNA-up38 is regulated in the leukemic cells and showed that Bcr-Abl repressed lncRNA-up38 expression through c-Myc-dependent signaling. Taken together, these results reveal that Bcr-Abl-mediated cellular transformation is associated with downregulation of tumor-suppressor lncRNA-up38 and suggest a potential strategy for the treatment of Bcr-Abl-positive leukemia.

Note: This abstract was not presented at the meeting.

Citation Format: Jilong Chen, Xuefei Wang. Identification of a novel lncRNA involved in Bcr-Abl mediated cellular transformation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3450. doi:10.1158/1538-7445.AM2017-3450
Abstract 3458: Role of zyxin in mitosis and colon cancer
Authors: ZHOU JIULI , Dong Jixin , CHEN YUANHONG , CHEN XINGCHENG , Stauffer Seth , Zeng Yongji
Published: July 2017

Zyxin is a LIM domain protein that is concentrated at sites of cell-cell adhesion, where it is proposed to dock proteins involved in cytoskeletal dynamics and signaling. Recently, Zyxin has been shown to promote LATS degradation and YAP activation in Hippo Pathway. Here, We have identified a novel mechanism for phospho-regulation of Zyxin in mitosis and its biological significance in colon cancer. We found that the mitotic kinase cyclin-dependent kinase 1 (CDK1) phosphorylates Zyxin in vitro and in vivo at serine 281, serine 308 and serine 344 during antimitotic drug-induced G2/M phase arrest and unperturbed mitosis. Moreover, Zyxin mitotic phosphorylation-deficient mutant (Zyxin3A, harboring S281A/S308A/S344A) was sufficient to suppress colon cancer cell proliferation and migration. We further demonstrated that depletion of Zyxin resulted in impaired colon cancer tumorigenesis in vitro and in vivo through inhibiting F-actin cytoskeleton and YAP activity. Importantly, these observations could be fully rescued by re-expression of wild type Zyxin, but not Zyxin-3A mutant. Expression profile analysis revealed CDK8 (cyclin-dependent kinase 8) as a mediator downstream Zyxin/YAP. Inhibition of CDK8 phenocopied knockdown of Zyxin in colon cancer cells. Ectopic expression of CDK8 substantially restored the tumorigenic ability of Zyxin depletion cells in vitro and in vivo. Expression levels of Zyxin and CDK8 are both elevated in colon tumors and are negatively correlated with survival. Together, our findings reveal a novel layer of regulation for Zyxin in mitosis and define the Zyxin-YAP-CDK8 axis as a novel oncogenic regulator in colon cancer.

Citation Format: JIULI ZHOU, Jixin Dong, YUANHONG CHEN, XINGCHENG CHEN, Seth Stauffer, Yongji Zeng. Role of zyxin in mitosis and colon cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3458. doi:10.1158/1538-7445.AM2017-3458
Abstract 3459: BRCA1 controls the cell division axis and governs ploidy and phenotype in human mammary cells
Authors: He Zhengcheng , Nemirovsky Oksana , Kannan Nagarajan , Eaves Connie , Maxwell Christopher A.
Published: July 2017

BRCA1 deficiency alters the relative proportions of progenitor cells in preneoplastic mammary tissue, and typically associates with breast cancers characterized by genomic instability and a basal-like cell phenotype. Oriented division of progenitor cells is one mechanism these cells use to maintain tissue homeostasis, and to suppress tumor formation. We now show that shRNA-mediated reduction of BRCA1 levels in non-tumorigenic and immortalized or freshly isolated, normal human mammary cells alters their plane of division with graded consequences that include the induction of aneuploidy in progeny cells, perturbation of polarity in spheroid cultures, and inhibition of clonal growth with favored expression of basal features. We also demonstrate a requirement for BRCA1 in establishing cortical asymmetry of NUMA-dynein complexes. Mutation of a single BRCA1 allele (BRCA1 185delAG/+) altered the division axis of isolated cells but their deficient spindle positioning was supervised by CDH1-positive adherens, which sustained oriented divisions and produced colonies with luminal features. These findings reveal a previously unrecognized consequence of mutant BRCA1 on the cell division axis, post-mitotic integrity and phenotype control in normal human mammary epithelial cells.

Note: This abstract was not presented at the meeting.

Citation Format: Zhengcheng He, Oksana Nemirovsky, Nagarajan Kannan, Connie Eaves, Christopher A. Maxwell. BRCA1 controls the cell division axis and governs ploidy and phenotype in human mammary cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3459. doi:10.1158/1538-7445.AM2017-3459
Abstract 346: Fatty acid synthase is a risk factor for bleomycin-induced lung fibrosis in mice
Authors: Jung Mi-Yeon
Published: July 2017

Pulmonary fibrosis is a significant complication in cancer patients treated with radiation or various chemotherapeutic agents. One such reagent, bleomycin, is used to treat a variety of cancers including, but not limited to, lymphoma, ovarian cancer, and melanoma. Its efficacy, however, is impacted by the incidence of significant pulmonary fibrosis, which often becomes dose limiting. In studies investigating the role(s) of metabolism in the profibrotic actions of transforming growth factor beta, we previously determined that increased fatty acid synthase (FASN) expression was observed in the fibrotic areas of lungs from idiopathic pulmonary fibrosis (IPF) patients compared to healthy controls. In that FASN levels are significantly increased in numerous human tumors including breast, prostate, colorectal, and ovarian, we wished to integrate these findings by examining whether inhibiting FASN activity could reduce pulmonary fibrosis. To that end, the cerulenin-derived FASN inhibitor C75 was tested in a murine treatment model of bleomycin-induced lung fibrosis. Following addition of C75, there was significant diminution in a number of profibrotic targets including collagen 1α1, fibronectin, connective tissue growth factor, and total lung collagen as assessed by hydroxyproline content. Not only was there a C75 dose-dependent decrease in the aforementioned targets, but lung function as assessed by peripheral blood oxygenation (on room air) was also stabilized/improved. Thus, FASN inhibition might reflect a possible target to maintain normal lung physiology in the context of cancer chemotherapy.

This work was supported by Public Health Service grants GM-055816 and GM-054200 from the National Institutes of General Medical Sciences.

Citation Format: Mi-Yeon Jung. Fatty acid synthase is a risk factor for bleomycin-induced lung fibrosis in mice [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 346. doi:10.1158/1538-7445.AM2017-346
Abstract 3463: Mitotic regulation of the tumor suppressor phosphatase PHLPP1
Authors: Grzechnik Agnieszka T. , Newton Alexandra C.
Published: July 2017

The PH domain Leucine-rich repeat Protein Phosphatases (PHLPP) are tumor suppressors originally discovered for their ability to directly dephosphorylate and inactivate the pro-survival kinase Akt, a key transducer of growth factor signaling. In the decade since this discovery, a number of other PHLPP targets have been elucidated, but still little is known about the molecular mechanisms governing the function and regulation of PHLPP itself. Here we report that PHLPP1 is hyperphosphorylated during mitosis and that, additionally, PHLPP1 suppresses the phosphorylation of the mitotic kinase CDK1. Specifically, we show that PHLPP1 undergoes an electrophoretic mobility shift in mitotic cells that is lost when lysates are treated in vitro with lambda phosphatase, suggesting that PHLPP1 is phosphorylated during mitosis. In addition to a cell cycle-dependent change in its post-translational modifications, PHLPP1 suppresses the phosphorylation of the main mitotic cyclin dependent kinase, CDK1, at Tyr15, an inhibitory phosphorylation that must be removed in order for cells to progress through mitosis. First, Tyr15 phosphorylation is increased in phlpp1-/- mouse embryonic fibroblasts (MEFs) compared to wild-type MEFs. Second, pharmacological inhibition of PHLPP1 in wild-type MEFs results in an increase in Tyr15 phosphorylation. Activation of key mitotic kinases, such as CDK1, and inactivation of phosphatases is required for the large increase in protein phosphorylation that occurs during mitosis. As such, reversible protein phosphorylation plays a role in regulating proper progression through mitosis, which is essential as errors in mitosis can result in aneuploidy, a hallmark of cancer. Our data support a previously undescribed role for PHLPP1 in regulating mitotic progression, potentially providing novel insight into how PHLPP suppresses tumorigenesis.

Citation Format: Agnieszka T. Grzechnik, Alexandra C. Newton. Mitotic regulation of the tumor suppressor phosphatase PHLPP1 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3463. doi:10.1158/1538-7445.AM2017-3463
Abstract 3469: Identification of novel human telomerase associated protein by mass spectrometry
Authors: Wang Jing , Bi Xuezhi , Li Shang
Published: July 2017

Telomeres are nucleoprotein structures at the ends of eukaryotic chromosomes that prevent them from degradation, recombination, and end-to-end fusion. Telomere consists of DNA repeats, which in human is of the sequence TTAGGG, and it is bound by a group of proteins called shelterin complex. Telomere is synthesized by telomerase. Telomerase is absent in most human somatic cells while it is up-regulated in 90% of cancer cells, in which telomerase plays an essential role in cancer cells’ replicative immortality. However, telomerase is only present at approximately 250 molecules per cancer cell. To exert its function of elongating telomere, telomerase has to find the end of telomeres which needed to be elongated. However, how telomerase is precisely recruited to telomere ends is not clearly understood. To better understand the recruitment process of telomerase, we initially screened and subsequently identified novel binding proteins of hTERT, the catalytic subunit of telomerase, through a pull-down assay using cancer cell lines with endogenously Flag-tagged hTERT. Using mass spectrometry (MS), we were able to identify a promising interesting protein. Further, direct interaction between TERT and this protein was validated. Then, we found that this protein can also interact with the shelterin component, TRF1 (Telomeric repeat-binding factor 1), which binds to double stranded telomeric DNA. Furthermore, from previous published reports, this protein was shown to interact with single-stranded telomeric DNA binding protein POT1 (Protection of Telomeres 1) in the shelterin complex. Functionally, overexpression of this protein in cancer cell lines can result in the elongation of telomeres, while knock-down or knock-out of the protein results in the shortening of telomere. In conclusion, we identified an hTERT-binding protein that affects telomere maintenance which may be involved in telomerase recruitment process. In the long run, it could serve as a potential therapeutic target for cancers.

Citation Format: Jing Wang, Xuezhi Bi, Shang Li. Identification of novel human telomerase associated protein by mass spectrometry [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3469. doi:10.1158/1538-7445.AM2017-3469
Abstract 3482: SNORD-X in glioblastoma: regulation and functional analysis
Authors: Gupta Shikha , Jha Prerana , Sarkar Chitra , Kulshreshtha Ritu
Published: July 2017

Glioblastoma Multiforme (GBM) is a kind of brain tumor which arises from astrocytes present in the brain. It represents approximately 15% of all primary brain tumor and has been considered as one of the deadliest type of cancer with very poor prognosis. Thus, there is an urgent need to identify novel targets for GBM therapy. Recently, non-coding RNAs (ncRNAs) such as miRNAs and lncRNAs have emerged as one of the promising tools as diagnostic or prognostic biomarkers and as novel targets for therapy. There are recent evidences that small nucleolar RNAs (snoRNAs), a subtype of ncRNAs might also play a key role in various cancers. In this context, we aimed at studying the role of snoRNAs in GBM. For this, we did snoRNA profiling in GBM patients and found that a particular cluster of snoRNA called SNORD-X was highly downregulated in adult as well as pediatric GBM patients. We also found SNORD-X promoter to be methylated and shows maternal imprinting. We next did functional analyses of SNORD-X by generation of an overexpression construct. Our preliminary data suggests that overexpression of SNORD-X in GBM cell lines (U87MG and A172) increases cell proliferation in vitro. It also enhances the colony forming capability of GBM cells. Further, its overexpression causes decrease in apoptosis in a caspase-dependent manner. A target analysis of SNORD-X was done using RNAhybrid software. We found EGR1, DCUN1D3 and BRCA1 to be the direct targets of SNORD-X using a combination of qRT-PCR and wild-type/mutated 3’UTR luciferase analyses. Overall, our work shows for the first time correlation and functional analysis of SNORD-X cluster in GBM.

Citation Format: Shikha Gupta, Prerana Jha, Chitra Sarkar, Ritu Kulshreshtha. SNORD-X in glioblastoma: regulation and functional analysis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3482. doi:10.1158/1538-7445.AM2017-3482
Abstract 3485: Identification and characterization of circular RNAs in cancer
Authors: Thrue Charlotte A. , Petri Andreas , Løvendorf Marianne B. , Dybkær Karen , Papaioannou Dimitrios , Garzon Ramiro , Kauppinen Sakari
Published: July 2017

Circular RNAs (circRNAs) are an abundant class of non-coding RNAs that are formed by a backsplice event resulting in formation of covalently closed circular RNA molecules. The efficiency of backsplicing is inferior to linear splicing, however, due to their long half-lives circRNAs can accumulate to high levels in cells. Functional characterization of a few circRNAs has shown that they can act as endogenous microRNA sponges and transcriptional regulators; one example is CDR1as/ciRS-7, which has been shown to act as a sponge for miR-7. However, the functions of most circRNAs are still unknown.

To study the potential role of circRNAs in cancer, we identified exonic circRNA candidates by analysing ENCODE RNAseq data from a panel of cancer cell lines. We used chimeric alignment detection implemented in the STAR aligner and subsequent filtering of output files to identify chimeric alignments consistent with circRNA backsplice sites. The circular nature of the identified putative circRNAs was validated by testing their resistance to RNase R digestion, and sequences surrounding the backsplice sites were used as target recognition sequences for designing LNA-modified gapmer antisense oligonucleotides (ASOs) to specifically knock down the circular isoforms of the RNA transcripts.

Data from ongoing studies on knockdown of several abundant circRNAs in cancer cell lines, assessment of the specificity of backsplice-targeting ASOs relative to the linear mRNA counterparts, and analyses to identify biological effects of modulating circRNA levels in cultured cells will be presented.

Citation Format: Charlotte A. Thrue, Andreas Petri, Marianne B. Løvendorf, Karen Dybkær, Dimitrios Papaioannou, Ramiro Garzon, Sakari Kauppinen. Identification and characterization of circular RNAs in cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3485. doi:10.1158/1538-7445.AM2017-3485
Abstract 3493: LncRNA as potential target in drug-resistant melanomas
Authors: Esteve-Puig Rosaura , Sanlorenzo Martina , Vujic Igor , Lai Kevin , Vujic Marin , Mould Dallas , Lin Kevin , Oses-Prieto Juan , Chand Shreya , Posch Christian , Burlingame Alma , Ortiz-Urda Susana
Published: July 2017

Melanoma is the most common lethal form of skin cancer. Activating mutations in BRAF and RAS oncogenes account for over 80% of all melanomas and hyper-activate the MAPK cascade. Despite major advances in targeted melanoma therapies over the past decade, drug resistance limits their efficacy.

Long noncoding RNAs (lncRNAs) are a new class of genes that do not encode proteins, which have been characterized as epigenetic regulatory molecules. LncRNAs have emerged as important players in cancer, potential predictors of therapeutic responses and targets for new treatments; however, their functions and precise molecular mechanisms remains largely unexplored.

Our study aims to find novel potential targets involved in the acquisition of resistance to MAPK inhibitors.

We identified an intergenic lncRNA (lincRNA) highly differentially expressed in drug-resistant melanoma samples, called MAPK Inhibitor Resistance Associated Transcript (MIRAT). We determined that MIRAT is localized mainly in the cytoplasm and plays a role in modulating the signal transduction pathways without regulating the genomic neighborhood. We discovered that MIRAT is upregulated in early drug tolerance to MAPK inhibition. Mechanistically, the gene overexpression and silencing of MIRAT, as well as preliminary RNA-protein interaction results, showed that it modulates the MAPK signal transduction pathway by binding to a downstream target of MEK.

Collectively, these results highlight the relevance of cytoplasmic lincRNA’s and their role in oncogenic signaling pathways, and further suggest that lncRNAs and their protein-binding partners may serve as novel potential targets for overcoming drug resistance in melanoma.

Note: This abstract was not presented at the meeting.

Citation Format: Rosaura Esteve-Puig, Martina Sanlorenzo, Igor Vujic, Kevin Lai, Marin Vujic, Dallas Mould, Kevin Lin, Juan Oses-Prieto, Shreya Chand, Christian Posch, Alma Burlingame, Susana Ortiz-Urda. LncRNA as potential target in drug-resistant melanomas [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3493. doi:10.1158/1538-7445.AM2017-3493
Abstract 35: The dual inhibition of Met and EGFR by ME22S, a novel Met/EGFR bispecific monoclonal antibody, suppresses the proliferation and invasion of laryngeal cancer
Authors: Lee Bok-Soon , Kim Dae Ho , Kim Yeon Soo , Chang Jae Won , Koo Bon Seok , Won Ho Ryun , Kim Haeng-Jun , Cha Hyun-Young , Lee Ji Min , Kim Kyung-Ah , Kim Chul-Ho
Published: July 2017

Background and purpose: It has been reported that the abnormal activation of receptor tyrosine kinases is associated with the development of many human carcinomas and the high activation of EGFR and Met mediates the tumorigenicity of laryngeal carcinoma. In this study, we have done the therapeutic efficacy of ME22S, a novel EGFR/Met bispecific antibody, in laryngeal carcinoma in vitro and in vivo was thoroughly evaluated.

Methods: The effects of ME22S on cell viability was assessed through MTT assays, then Western blotting and immunocytochemistry were used to determine the expression of EGFR and Met. Also, wound healing and invasion assays were performed to observe the inhibitory effects of ME22S.

Results: We found the ability of ME22S reducing the expression of both EGFR and Met and significantly inhibiting the cell migration, invasion, and proliferation of SNU899 and HN3 in vitro. Also the notably reduced levels of p-Met, p-ERK, and p-AKT were found when the cells were treated with only ME22S alone or with HGF together. Meanwhile, ME22S, interestingly enough, caused caspase-3-dependent apoptotic cell death when HN3 cells were treated with ME22S for 72h, decreased the HGF-induced Slug expression, and also inhibited the tumor growth of HN3 cells in a xenograft model in vivo.

Conclusion: Taken together, our findings suggest that the dual inhibition of EGFR and Met through ME22S largely suppresses the invasion and growth of laryngeal carcinoma both in vitro and in vivo, hence, can be a practical approach as a novel therapeutic strategy for the treatment of laryngeal carcinoma.

Citation Format: Bok-Soon Lee, Dae Ho Kim, Yeon Soo Kim, Jae Won Chang, Bon Seok Koo, Ho Ryun Won, Haeng-Jun Kim, Hyun-Young Cha, Ji Min Lee, Kyung-Ah Kim, Chul-Ho Kim. The dual inhibition of Met and EGFR by ME22S, a novel Met/EGFR bispecific monoclonal antibody, suppresses the proliferation and invasion of laryngeal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 35. doi:10.1158/1538-7445.AM2017-35
Abstract 3502: RNAi screen identified novel long non-coding RNAs involved in cell growth control, migration, and invasion
Authors: Kuo Tse-Chun , Hsu Sheng-Chieh , Chou Hui-Ju , Li Pei-Shan , Wu Szu-Huei , Yu Yen-Ling , Song Jen-Shin , Kung Hsing-Jien
Published: July 2017

Triple-negative breast cancer (TNBC) is a highly aggressive form of breast cancer often associated with poor prognosis. Owing to the lack of therapeutic targets, patients with TNBC tend to rely on standard chemotherapy; however, chemo-resistance often develops in TNBC as a result of deregulated cell death control like in many other tumors. Long non-coding RNAs (lncRNAs) are transcripts longer than 200bp and have little or no protein-coding capacity. Although once considered as transcriptional noise, significant numbers of lncRNAs are now known as modulators of protein expression and functions and are implicated in a range of developmental processes and diseases. As many of them influence oncogenes and tumor suppressors in cancer, they represent excellent biomarkers as well as potential targets for therapy. In order to gain more insight on cell death regulation and to seek potential targets for TNBC, we employed an RNAi screen targeting 2231 lncRNAs in MDA-MB-231 TNBC cells. By using changes in cell metabolic activity (MTT assay) as readouts for primary screen and changes in cell number as confirmation, our unbiased screen identified two novel lncRNAs that drastically reduced cell proliferation when depleted. In addition to cell growth, the two lncRNAs also contribute to cancer cell migration and invasion. Interestingly, our gene profiling analyses and RNA immunoprecipitation studies show the two lncRNAs are involved in signaling cascades known to promote mammary tumorigenesis including EGF and β-catenin. We are now trying to delineate the detailed molecular mechanism(s) by which the two lncRNAs contribute to TNBC tumorigenesis. Together, our current results suggest a possibility for the two novel lncRNAs as drug targets in triple-negative breast cancer.

Citation Format: Tse-Chun Kuo, Sheng-Chieh Hsu, Hui-Ju Chou, Pei-Shan Li, Szu-Huei Wu, Yen-Ling Yu, Jen-Shin Song, Hsing-Jien Kung. RNAi screen identified novel long non-coding RNAs involved in cell growth control, migration, and invasion [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3502. doi:10.1158/1538-7445.AM2017-3502
Abstract 351: mTOR-ERK co-targeting strategies for head and neck cancer therapy
Authors: Wang Zhiyong , Delgado Esteban , Yamaguchi Kosuke , Iglesias-Bartolomé Ramiro , Amornphimoltham Panomwat , Molinolo Alfredo , Chen Qianming , Gutkind J.Silvio
Published: July 2017

Despite therapeutic advances in the past decade, head and neck squamous cell carcinoma (HNSCC) continues to be a major public health concern. Although recent studies of human cancer genomes identified remarkable complexity of genomic alterations in HNSCC, surprisingly, most aberrations have fallen within key driver signaling pathways. Among them, PI3K/AKT/mTOR pathway is most frequently activated in over 80% of the HNSCC patients. We showed that the majority of HNSCC lesions exhibit high levels of phosphorylated ribosomal S6 protein (pS6), key downstream target of the AKT-mTOR pathway, and that inhibition of mTOR by the use of rapamycin causes a rapid decrease in the level of pS6 and the apoptotic death of HNSCC tumor xenografts, thereby causing tumor regression. Our initial studies identified the AKT-mTOR pathway as a potential therapeutic target for HNSCC and provided strong rational to initiate multiple clinical trials. Indeed, inhibitors of PI3K/AKT/mTOR block mTOR activity and exert a beneficial response in HNSCC patients. However, therapeutic resistance was evidenced in several trials, which might be due to the activation of adaptive survival signaling in tumor cells. To explore the therapeutic option that can overcome the resistance, we performed a synthetic lethality screen in HNSCC cell lines using shRNA libraries. We found that multiple molecules involved in ERK pathway were highly represented. Furthermore, treating HNSCC cell lines harboring activated mTOR with small molecule inhibitors that target PI3K/AKT/mTOR pathway can sequentially induce ERK activation, which is consistent with the findings observed in numerous clinical trials. We demonstrated that co-targeting mTOR and ERK using trametinib, a MEK1/2 inhibitor or cetuximab, an EGFR monoclonal antibody, exhibited the synergistic effect by sensitizing HNSCC to priming with the mTOR inhibitor. Specifically, mTOR-ERK co-targeting prevented the growth of HNSCC tumor xenografts by decreasing cell proliferation, reducing lymphangiogenesis and activating apoptosis. In addition, we have recently used a phosphoproteomics approach to identify the molecular mechanisms of ERK feedback activation caused by PI3K/AKT/mTOR inhibition. Overall, our promising findings from these preclinical studies suggest that the use of mTOR-ERK co-targeting strategies may provide the novel alternative approach to achieve durable responses (cure) in HNSCC cancer patients.

Citation Format: Zhiyong Wang, Esteban Delgado, Kosuke Yamaguchi, Ramiro Iglesias-Bartolomé, Panomwat Amornphimoltham, Alfredo Molinolo, Qianming Chen, J.Silvio Gutkind. mTOR-ERK co-targeting strategies for head and neck cancer therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 351. doi:10.1158/1538-7445.AM2017-351