bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 842: The NALP3 inflammasome and IL-1â signaling link asbestos-induced inflammation with the development of malignant mesothelioma
Authors: Kadariya Yuwaraj , Menges Craig W. , Talarchek Jacqueline , Cai Qi , Klein-Szanto Andres J. , Pietrofesa Ralph A. , Christofidou-Solomidou Melpo , Mossman Brooke T. , Shukla Arti , Testa Joseph R.
Published: July 2016

Exposure to asbestos is causally associated with the development of malignant mesothelioma (MM), a cancer of cells lining the internal body cavity. MM is an aggressive cancer that is resistant to all current therapies. Once inhaled or ingested, asbestos causes inflammation in and around tissues that come in contact with these carcinogenic fibers. Recent studies suggest that inflammation is a major contributing factor in the development of many types of cancer, including MM. The NALP3 inflammasome, including the component ASC, is an important mediator of inflammation that senses extracellular insults, such as infection or asbestos, and activates a signaling cascade resulting in release of mature IL-1â and recruitment of inflammatory cells. To determine if inflammasome-mediated inflammation contributes to asbestos-induced MM, we chronically exposed Asc-deficient mice to asbestos and evaluated tumor incidence, latency and overall survival. Asc-deficient mice showed a significant delay in tumor onset compared to wild-type animals, suggesting that the NALP3 inflammasome plays a role in MM carcinogenesis. We also tested whether inflammation-related release of IL-1â promotes tumor development in an accelerated mouse model of asbestos-induced MM. Nf2+/-;Cdkn2a+/- mice exposed to asbestos in the presence of the IL-1â antagonist, anakinra, showed a 50% longer disease-free survival than in similarly exposed mice given vehicle control, consistent with IL-1â signaling contributing significantly to MM development. Collectively, these studies provide evidence for a link between asbestos-associated inflammation/IL-1â signaling and the development of MM; furthermore, these findings provide rationale for chemoprevention strategies targeting inflammation-related IL-1â signaling in high risk, asbestos-exposed populations.

Citation Format: Yuwaraj Kadariya, Craig W. Menges, Jacqueline Talarchek, Qi Cai, Andres J. Klein-Szanto, Ralph A. Pietrofesa, Melpo Christofidou-Solomidou, Brooke T. Mossman, Arti Shukla, Joseph R. Testa. The NALP3 inflammasome and IL-1â signaling link asbestos-induced inflammation with the development of malignant mesothelioma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 842.
Abstract 843: Spontaneous immortalization of human mammary epithelial cells from a woman with a germline STK11 mutation
Authors: Euhus David , Bu Dawei , Considine Michael , Cope Leslie
Published: July 2016

Spontaneous immortalization of benign mammary epithelial cells (MEC) in culture is very rare. Models of MEC immortalization suggest that cells escape M0 growth arrest (selection) by silencing p16 expression after

which they encounter a second growth arrest, M1 (agonescence), characterized by telomere shortening and profound genomic instability. Pre-selection, post-selection, and spontaneously immortalized MEC from a woman with a germline STK11 mutation were subjected to a comprehensive “OMICS” evaluation including miRNA, transcriptome, methylome, exome, and comparative genomic hybridization (CGH) in order to understand the molecular events associated with MEC immortalization. Principle component analysis of gene expression identified 4 significant components, the first of which showed the greatest separation between the different time points. The most notable expression changes were upregulation of FOXQ1, IL1-related genes, Ras-related genes, and fibronectin and downregulation of COL1, COL6, CK19, and CDH1. MEC gradually acquired global hypomethylation of non-CpG DNA and regional hypermethylation of CpG islands (including p16). A classifier based on methylation pattern showed that immortalized cells moved away from identity with The Cancer Genome Atlas (TCGA) benign samples towards TCGA breast cancer samples. Immortalization was associated with a modest accumulation of mutations and a modestly greater number of DNA copy number alterations than was observed in earlier passage cells but no evidence of general genomic instability. Immortal cells showed a large deletion at 10q13-15 and deletions at 10p11-15 and 17p11-13 that were associated with reduced expression of genes in those regions. Post-selection bulk cell populations are quite distinct from early passage cells, but are not direct precursors of the immortal cells as some DNA copy number alterations acquired by post-selection cells were not observed in the immortalized cells. Rather, in this instance, it is likely that post-selection cells established conditions conducive to the immortalization of a rare cell in the mix. This has implications for the development of tissue-based approaches for breast cancer risk stratification.

Citation Format: David Euhus, Dawei Bu, Michael Considine, Leslie Cope. Spontaneous immortalization of human mammary epithelial cells from a woman with a germline STK11 mutation. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 843.
Abstract 845: Diet stamps on bugs: early life dietary energy intake impacts gut microbiota
Authors: Xu Jinyu , Galley Jeffrey , Bailey Michael , Thomas-Ahner Jennifer , Clinton Steven , Olivo-Marston Susan
Published: July 2016

The complex and dynamic interactions between diet, gut microbiota (GM) structure and function, and colon carcinogenesis are only beginning to be elucidated. We examined the colonic microbiota and aberrant crypt foci (ACF) in C57BL/6N female mice fed various dietary interventions (control, energy restricted and high-fat) provided during two phases (initiation and progression) of azoxymethane (AOM)-induced early colon carcinogenesis. During progression (wks. 22- 60), a high-fat diet enhanced ACF formation compared to a control or energy restricted diet. In contrast, energy restriction during initiation phase (wks. 3-21) enhanced ACF burden at 60 weeks, regardless of the diet in progression phase. Alterations in GM structure during the initiation phase diet were partially maintained after changing diets during the progression phase. However, diet during the progression phase had major effects on the mucosal GM that energy restriction in progression phase increased Firmicutes and reduced Bacteroidetes compared to a high-fat diet, regardless of initiation phase diet, suggesting that diet may have both transient effects as well as a lasting impact on GM composition. Integration of early life and adult dietary impacts on the colonic microbial structure and function with host molecular processes involved in colon carcinogenesis will be key to defining preventive strategies.

Citation Format: Jinyu Xu, Jeffrey Galley, Michael Bailey, Jennifer Thomas-Ahner, Steven Clinton, Susan Olivo-Marston. Diet stamps on bugs: early life dietary energy intake impacts gut microbiota. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 845.
Abstract 849: Improved geometric deconvolution of bulk tumor genomic data
Authors: Roman Theodore , Xie Lu , Schwartz Russell
Published: July 2016

We develop new methods for improved computational deconvolution of clonal populations from heterogeneous bulk genomic data. Deconvolution methods have become an important tool in resolving intratumor heterogeneity from bulk tumor genomic data, but prevailing methods are still able only to resolve only coarse-grained approximations of true clonal substructure. We seek to improve a genomic deconvolution strategy called “simplicial complex inference” that is designed to take advantage of similarity between the clonal subpopulations of single tumors or tumor regions to better infer portraits of those clonal subpopulations and their distribution across tumor samples.

We evaluate the method through application to DNA copy-number (CN) data from a 472-tumor ovarian cancer (OV) panel and a 408-tumor lung squamous small cell carcinoma (LUSC) panel from The Cancer Genome Atlas (TCGA). We deconvolved the OV data into a mixture of six clonal models, two per data sample, each interpreted as a consensus copy number profile of one recurring progression stage found in a subset of OV samples. We analyzed the resulting models for enrichment of labels derived from RNASeq-based subtyping (Verhaak et al., 2013), revealing significant correspondence by chi-squared test between tumor subclusters identified by the deconvolution and OV subtypes (p = 0.0168). We further analyzed the clonal models to identify regions of significant amplification or loss, which we tested for enrichment of gene ontology (GO) terms and pathway terms through the DAVID toolkit. Ontological analysis showed significant enrichment for GO terms connected to genetic regulation and proliferation including “DNA binding,” “DNA packing,” and “protein-DNA complex” (p < 0.001). The LUSC data was partitioned into nine clonal models, three per sample. A log-rank test showed significant correspondence between mixture subclustering and survival for LUSC patients (p < 0.001). Regions of significant CN gain or loss identified in the clonal models were found by DAVID to be significantly enriched for pathway terms including “Cytokine network” (p = 0.0025) and “Cytokines and Inflammatory Response” (p = 0.0043). These findings suggest that more sophisticated models of clonal population structures can enhance our ability to deconvolve heterogeneous tumor genomic data and lead to better inference of clonal substructure.

Citation Format: Theodore Roman, Lu Xie, Russell Schwartz. Improved geometric deconvolution of bulk tumor genomic data. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 849.
Abstract 87: Clustered somatic mutations are frequent in transcription factor binding motifs in melanoma and other cutaneous malignancies
Authors: Colebatch Andrew J. , di Stefano Leon , Wong Stephen Q. , Papenfuss Anthony T. , McArthur Grant A.
Published: July 2016

We sought to explore the etiology, frequency and clinicopathologic associations of clustered noncoding hotspot mutations in cutaneous malignancies (melanoma, basal cell carcinoma (BCC) and Merkel cell carcinoma (MCC)). We did so by applying a sliding window approach which identified clusters in melanoma whole exome and genome sequencing data. These regions were then evaluated for recurrence in other skin malignancies by using whole exome data from BCC and MCC samples; motifs were extracted using a maximum expectation algorithm. We then sequenced 170 annotated melanoma clinical samples using a targeted amplicon panel which permitted analysis of clinicopathological correlations. Numerous recurrent clustered hotspot mutations were identified in melanoma whole exome and genome data sets in proximal promoters, with some gene-associated clusters present in up to 25% of cases. Mutations were clustered around canonical ETS and SP1 transcription factor binding sites and had a clear UV signature. These promoter mutations correlated with overall mutation load and NF1 mutation status, and showed significant enrichment in specific melanoma subtypes (desmoplastic and lentigo maligna melanoma), sun exposed anatomical locations, male gender, ulceration and increasing age. Identical clusters were identified in BCC and MCC samples, including several previously annotated in melanoma. In conclusion, we identify a novel class of mutation in cutaneous malignancies which rather than being defined by a particular genomic position are related to a position in a transcription factor binding motif. These mutations are also independent of cell of origin. Clinicopathologic correlation and mutation analysis support an etiologic role for chronic UV irradiation.

Citation Format: Andrew J. Colebatch, Leon di Stefano, Stephen Q. Wong, Anthony T. Papenfuss, Grant A. McArthur. Clustered somatic mutations are frequent in transcription factor binding motifs in melanoma and other cutaneous malignancies. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 87.
Abstract 870: Derivation and characterization of antibodies from immune checkpoint blockade treated cancer patients
Authors: Branum Mark , Smith Laura , Brett Heather , Algate Paul , Greenfield Brad , Brown William , Swiderek Kristine M.
Published: July 2016

The IgG+ memory B cell repertoire of cancer patients demonstrating durable responses to immune checkpoint blockade therapy has been interrogated to identify tumor specific antibodies. Matched serum and PBMC samples have been collected from 10 melanoma patients with durable responses to treatment with ipilimumab. A panel of 7 well characterized melanoma cell lines with a diverse set of oncogenic driver mutations has been employed to assess durable responder patient serum reactivity to cell surface determinants in a live-cell flow cytometry assay. IgG+ memory B cells obtained from several prioritized durable responder melanoma patients have been plated out on 384 well plates at monoclonal density. The memory B cells have been activated in a short term culture system optimized to expand the clonal B cells and induce secretion of IgG. Antibodies secreted from clonally expanded B cells have been screened using live cell high throughput multiplex flow cytometry assays against a panel of melanoma cell lines. A prioritized subset of antibodies, selected on the basis of tumor cell line binding profiles and antibody v-region sequences, has been cloned and expressed as recombinant IgG1. A cell surface target for a prioritized antibody has been identified and the in vitro analysis and characterization of the antibody including functional activity and immunohistochemistry staining on tumor and normal tissues has been carried out in preparation for in vivo testing of efficacy.

Citation Format: Mark Branum, Laura Smith, Heather Brett, Paul Algate, Brad Greenfield, William Brown, Kristine M. Swiderek. Derivation and characterization of antibodies from immune checkpoint blockade treated cancer patients. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 870.
Abstract 875: Acquisition of chemoresistance in tumor cells requires crosstalk between dying and remnant live tumor cells via HMGB1
Authors: Zhou Junmin , Chen Xianghong , Gilvary Danielle L. , Tejera Melba Marie , Eksioglu Erika A. , Trinh Thu Le , Wei Sheng , Djeu Julie
Published: July 2016

Chemoresistance often develops during drug treatment in cancer patients but its mechanism remains unclear. To understand acquisition of drug resistance, we investigated whether tumor cells dying during drug treatment might signal to neighboring live cells to program themselves for survival in the drug-containing environment. The experimental design was to identify a factor in the supernatants of docetaxel (DTX)-treated human DU145 and PC3 prostate tumor cells that could cause live tumor cells to resist apoptosis and cell death under docetaxel. We discovered that dying cells release HMGB1 which can bind TLR4 or RAGE on live tumor cells causing the induction of cytoplasmic Clusterin (CLU). We demonstrated that CLU is a potent prosurvival protein that can trap Bax from translocating to the mitochondria to cause caspade 3 activation and subsequent apoptosis. Proof of this pathway came from the ability of anti-HMGB1to prevent supernatants from dying cells to induce CLU in newly-plated tumor cells. The receptors for HMGB1 were identified by use of anti-TLR4 and anti-RAGE, which prevented recombinant HMGB1 or dying cell supernatants from inducing CLU in newly-plated tumor cells, causing them to subsequently succumb to DTX toxicity. Moreover, overexpression of anti-sense CLU in tumor cells abrogated their ability to respond to recombinant HMGB1 or dying cell supernatants to induce drug resistance. Most importantly, standard chemotherapeutic agents including gemcitabine, taxol, Ara-C, doxorubicin, cisplatin, etoposide and carboplatin, can cause the release of HMGB1 from dying tumor cells, implicating this crosstalk between dying and live tumor cells as a common pathway towards multi-drug resistance.

Citation Format: Junmin Zhou, Xianghong Chen, Danielle L. Gilvary, Melba Marie Tejera, Erika A. Eksioglu, Thu Le Trinh, Sheng Wei, Julie Djeu. Acquisition of chemoresistance in tumor cells requires crosstalk between dying and remnant live tumor cells via HMGB1. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 875.
Abstract 893: Diffuse gastric adenocarcinoma often harbors KMT2C mutations resulting in malignant phenotypes and worse overall survival
Authors: Cho Soo-Jeong , Yoon Changhwan , Lee Jun Ho , Chang Kevin K. , Aksoy Bulent A. , Park Do Joong , Yoon Sam S.
Published: July 2016

Purpose: The Lauren diffuse type of gastric adenocarcinoma (DGA) is genomically stable compared to the intestinal type, but the primary driver mutations in DGA are still being defined. Lysine (K) -specific methyltransferase 2C (KMT2C), also known as mixed lineage leukemia 3 (MLL3), is a histone methyltransferase involved in transcriptional coactivation and can be mutated in gastric cancer. The purpose of this study was to determine the frequency of KMT2C mutation in DGA and to elucidate the role of KMT2C in DGA tumorigenesis and progression.

Methods: Whole exome sequencing was performed on matched tumor samples of 27 patients with DGA who underwent potentially curative surgical resection. Immunohistochemistry (IHC) was performed for KMT2C protein expression on these same tumors and on an additional 289 tumors from a separate cohort of patients with DGA. KMT2C overexpression in DGA cell lines (MKN-45 and SNU-668) using lentiviral transduction and KMT2C shRNA knockdown in normal gastric epithelial cells were examined in several in vitro assays.

Results: Thirty-eight percent of the 27 sequenced DGA specimens harbored somatic mutations in KMT2C. The next most frequent mutation, CDH1, which encodes E-cadherin, was found in 28% of specimens. Patients with KMT2C mutation tended to have worse overall survival (OS) compared to patients without the mutation (p = 0.194). Additional tumor tissue for IHC analysis was only available from 11 of 27 patients. Among this group, patients with tumors expressing KMT2C protein had better OS compared to patients whose tumors did not express KMT2C (p = 0.034). In an external validation set of 289 DGA samples, KMT2C expression again correlated with better OS (p = 0.009). In the DGA cell lines, KMT2C overexpression reduced proliferation by 32-39%, soft agar colony formation by 75-77%, spheroid cell formation by 77-78%, cell migration by 52-60%, and cell invasion by 50-74%. KMT2C overexpression also increased resistance to 5-FU and cisplatin chemotherapy. KMT2C shRNA knockdown in gastric epithelial cells increased proliferation, soft agar colony formation, migration, and invasion by 1.7- to 25-fold.

Conclusions: KMT2C is frequently mutated in DGA, and loss of KMT2C protein expression correlates with worse OS. Restoration of KMT2C expression in DGA cell lines reduces cancer stem-like cell and malignant phenotypes. Thus, KMT2C may be a novel therapeutic target for patients with DGA.

Citation Format: Soo-Jeong Cho, Changhwan Yoon, Jun Ho Lee, Kevin K. Chang, Bulent A. Aksoy, Do Joong Park, Sam S. Yoon. Diffuse gastric adenocarcinoma often harbors KMT2C mutations resulting in malignant phenotypes and worse overall survival. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 893.
Abstract 900: In vivo modeling of NRAS-mutant melanoma reveals differential preventative efficacy amongst SPF30 sunscreens
Authors: Holderbaum Andrea M. , Hennessey Rebecca C. , Gillahan James E. , Bonilla Anamaria , Delaney Conor , Kladney Raleigh D. , Tober Kathleen L. , Oberyszyn Tatiana M. , Burd Christin E.
Published: July 2016

Cutaneous melanoma claims the life of one American every hour, and while the etiology of this tumor type is not entirely understood, exposure to ultraviolet (UV) radiation (280-340nm) is a major risk factor. For this reason, the use of UV-blocking sunscreens is strongly advocated; however, few studies have tested the relative efficacy of these agents in preventing melanoma formation in vivo. Here, we employed a new genetically engineered mouse model (TpN61R) to examine the ability of 6 chemically distinct SPF30 sunscreens to prevent melanoma. In this TpN61R model, topical 4-hydroxytamoxifen (4OHT) treatment induces the melanocyte-specific expression of oncogenic NRas as well as inactivation of the p16INK4a tumor suppressor. These genetic lesions co-occur in ∼24% of all human melanomas, making the TpN61R model biologically relevant. Since NRAS mutations are an early and UV-independent event in human melanoma, TpN61R mice were first painted with 4OHT and then exposed to a single dose of 4.5 kJ/m2 UVB light one day later. Exposed skin from TpN61R mice showed transient cyclobutane pyrimidine dimer (CPD) formation, however no evidence of edema or inflammation was observed. Despite these mild effects, UVB exposure reduced the melanoma-free survival of TpN61R mice by 80% and increased tumor incidence rate from 1.2 to 3.4 tumors/mouse. Further experiments using a variety of UVB doses (0.25, 1.0, 2.3 and 9.0 kJ/m2) in the TpN61R model revealed a dose-dependent increase in early melanoma incidence. Together, these data establish the exquisite cooperation of UV light and oncogenic NRas mutations in driving melanoma. Taking advantage of this unique model, we tested the preventative efficacy of SPF30 sunscreens with differing chemical composition. Sunscreens components included UVA (avobenzene), UVB (homosalate, octisalate) and broad spectrum (oxybenzone, octocrylene, zinc oxide) blocking agents. Application of sunscreen prior to UVB exposure decreased DNA damage, delayed melanoma onset and reduced tumor incidence in a sunscreen-dependent manner. Thus, SPF30 sunscreens do not equally prevent UVB-induced, NRAS mutant melanoma. This work establishes the first in vivo system to test sunscreen efficacy in NRAS-driven melanoma and will help direct the development of improved melanoma preventatives.

Citation Format: Andrea M. Holderbaum, Rebecca C. Hennessey, James E. Gillahan, Anamaria Bonilla, Conor Delaney, Raleigh D. Kladney, Kathleen L. Tober, Tatiana M. Oberyszyn, Christin E. Burd. In vivo modeling of NRAS-mutant melanoma reveals differential preventative efficacy amongst SPF30 sunscreens. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 900.
Abstract 934: Tributyltin-induced dysregulation of inflammatory cytokine levels in human and mouse immune cells
Authors: Lawrence Shanieek , Pellom Samuel T. , Shanker Anil , Whalen Margaret
Published: July 2016

The effects of Tributyltin (TBT), a widespread environmental contaminant, on the induction of in vivo and ex vivo cytokine production and secretion were investigated. Human monocyte-depleted peripheral blood mononuclear cells and different groups of Balb/C mice were stimulated with desired concentrations of TBT (200-2.5 nM for human studies and 200-25 nM for mouse studies). The quantitative determination of IFNγ, TNFα and IL-1β was performed in mouse sera by MagPix analysis and western blot. The levels of these cytokines were measured at different time points: 6, 12, 24 and 48 h after injection. Analysis of IFNγ and TNFα in human cells were conducted by ELISA and western blot. MAPK and NF-κB pathways were inhibited to determine TBT-induced dysregulation of IFNγ and TNFα. Inhibition of the p38 and p44/42 pathways blocked the TBT-induced elevation of IFNγ, while the inhibition of the p38 pathway blocked the TBT-induced elevation of TNFα. Results showed that TBT increased the levels of IFNγ, TNFα and IL-1β in the sera of mice. IFNγ and TNFα secretion from human monocyte-depleted peripheral blood mononuclear cells also increased, showing striking agreement in the response to TBT between the mouse and human systems. These data suggest that TBT exposure can result in an inflammatory environment, which could predispose the host to various immunopathologies.

Grant U54CA163066 from the National Institute of Health

Citation Format: Shanieek Lawrence, Samuel T. Pellom, Anil Shanker, Margaret Whalen. Tributyltin-induced dysregulation of inflammatory cytokine levels in human and mouse immune cells. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 934.
Abstract 935: Oral cancer cells may hijack stem cell altruism to survive during extreme hypoxia, and exposure to chemotherapeutic drugs
Authors: Bhuyan Rashmi , Li Hong , Gayan Sukanya , Pal Bidisha , Bayat-Mokhtari Reza , Phukan Jyotirmoy , Baishya Debabrata , Sarma Anupam , Talukdar Joyeeta , Alkurdi Manaf Muhammad , Tasabehji Wael , Bhuyan Seema , Gogoi Gayatri , Pulu Ista , Yeger Herman , Das Bikul
Published: July 2016

The mechanism of tumor hypoxia induced reprogramming of cancer cells are not understood well. Hypoxia might activate evolutionary preserved cellular defense mechanism that could contribute to tumor progression and metastasis. We recently described that during hypoxia/oxidative stress, human embryonic stem cells (hES) exhibit an altruistic defense mechanism, where a few cells reprogram to a highly undifferentiated state, and secrete glutathione to protect rest of the community of cells from oxidative stress induced DNA damage (Das B et al, Stem Cells, 2012). The altruistic stem cells phenotype exhibited high HIF-2alpha and low p53 activity. After a few weeks, the reprogrammed cells, although highly fit to survives, underwent spontaneous apoptosis/differentiation by re-activating the p53/MDM2 oscillation. Thus, the reprogrammed cells sacrificed its own fitness to enhance the fitness of the rest of the community, an altruistic behavior (Das B et al. Stem Cells, 2012). Here, we investigated the potential hijacking of the altruistic defense mechanism by oral cancer cells during exposure to hypoxia. We exposed the four oral cancer cell lines SCC-25, SCC-15, SCC-9 and SCC-5 to extreme hypoxia followed by re-oxygenation for 72 hours. We found that while majority of cancer cells underwent apoptosis, a few cancer cell lines survived, exhibited side-population (SP) phenotype, high level of HIF-2alpha, Nanog, Sox-2 and MYC. The SP cells exhibited migratory activity, as well as high tumorigenic and metastatic activity in NOD/SCID mice. ChIP assay indicated that HIF-2alhpa interact with MYC and NOTCH1. The conditioned media of SP cells exhibited high level of glutathione, and ability to protect non-SP cells from cisplatin-induced toxicity. These results indicate that HIF-2alpha and MYC may cooperate to reprogram a few oral cancer cells to altruistic stemness phenotype. The altruistic phenotype that exhibited cytoprotective activity against cisplatin mediated toxicity. Thus, similar to bacteria, where altruistic biofilm exhibit novel drug resistance mechanism, stem cell altruism may serve as a novel drug resistance mechanism in oral cancer.

Citation Format: Rashmi Bhuyan, Hong Li, Sukanya Gayan, Bidisha Pal, Reza Bayat-Mokhtari, Jyotirmoy Phukan, Debabrata Baishya, Anupam Sarma, Joyeeta Talukdar, Manaf Muhammad Alkurdi, Wael Tasabehji, Seema Bhuyan, Gayatri Gogoi, Ista Pulu, Herman Yeger, Bikul Das. Oral cancer cells may hijack stem cell altruism to survive during extreme hypoxia, and exposure to chemotherapeutic drugs. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 935.
Abstract 949: Epigenetic silencing of microRNA-34a promotes cholangiocarcinoma growth by regulating Notch pathway
Authors: Kwon Hyunjoo , Song Kyoungsub , Han Chang , Zhang Jinqing , Ungerleider Nathan , Yao Lu , Wu Tong
Published: July 2016

Aberrant expression and regulation of microRNAs (miRNAs) have been found in multiple cancers and their dysregulation importantly contribute to the tumorigenic processes. Among these miRNAs, miRNA-34a (miR-34a) functions as a tumor suppressor and its dysregulation has been reported in several cancers. In this study, we investigated epigenetic regulation and biological function of miR-34a in human cholangiocarcinoma cells. Our data indicate that miR-34a expression is epigenetically silenced in human cholangiocarcinoma cells (CCLP1, SG231, Hucct1 and TFK1) compared to the nonmalignant biliary epithelial cells (H69). Treatment of human cholangiocarcinoma cells with the DNA methylation inhibitor, 5’aza-2’deoxycytidine (5-Aza-CdR), or the EZH2 (enhancer of zeste 2 polycomb repressive complex 2) inhibitor, GSK126, enhanced the expression of miR-34a. In parallel, knockdown of EZH2 by shRNA also significantly decreased miR-34a expression in human cholangiocarcinoma cells. We further showed that DNA methylation and EZH2-mediated histone H3 lysine 27 trimethylation (H3K27Me3) repressed miR-34a gene expression, as determined by methylation-specific PCR and chromatin immunoprecipitation. The DNA methylation and EZH2-mediated H3K27Me3 independently repress miR-34a expression in cholangiocarcinoma cells since treatment of 5-Aza-CdR and GSK126 had no effect on H3K27me3 and DNMT1 levels, respectively. Functional analyses revealed that miR-34a mimic decreased cholangiocarcinoma cell proliferation, colony formation and migration, in vitro. Moreover, we identified Notch1, Notch2 and Jagged 1, which are the major receptors and ligands of the Notch pathway, as miR-34a target genes in cholangiocarcinoma cells. Accordingly, forced overexpression of miR-34a significantly decreased the expression of Notch1, Notch2 and Jagged1, as determined by Western blotting and qRT-PCR analyses. Our results provide novel evidence that miR-34a expression is epigenetically silenced by DNA methylation and EZH2 in human cholangiocarcinoma cells. Thus, epigenetic induction of miR-34a or miR-34a replacement therapy may represent a novel therapeutic approach for the treatment of cholangiocarcinoma.

Citation Format: Hyunjoo Kwon, Kyoungsub Song, Chang Han, Jinqing Zhang, Nathan Ungerleider, Lu Yao, Tong Wu. Epigenetic silencing of microRNA-34a promotes cholangiocarcinoma growth by regulating Notch pathway. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 949.
Abstract 953: miR-31 targets ARID1A to enhance the progression of oral carcinoma
Authors: Lu Wen-Cheng , Lin Shu-Chun , Chang Kuo-Wei
Published: July 2016

miR-31 is an oncogenic microRNA in several malignancies including oral squamous cell carcinoma (OSCC). However, the function of miR-31 in modulating the stem cell properties in OSCC has not yet been explored. AT-rich interacting domain (ARID) containing protein members play pluripotent roles in modulating chromatin accessibility of the transcription machineries for gene expression. ARID1A has been shown suppressive to several malignancies and the driver mutations in ARID1A gene are noted in tumors. In this study, we identified that miR-31 targets ARID1A in OSCC, and a reverse expression between miR-31 and ARID1A was noted in OSCC tumors. Furthermore, the miR-31 induced migration, invasion, CSC cell population and stemness gene expression were rescued by ARID1A expression in OSCC cells. ARID1A suppressed the transcription activity of a panel of stemness markers through its affinity to A-T rich region in the promoters. A reverse correlation between ARID1A and these stem cell markers was also seen in OSCC tumors. miR-31+/ARID1A− OSCC cell subclone exhibited higher potency in sphere formation, as well as tumorigenesis and neck metastasis in nude mice relative to other cell subclones. In addition, tumors carrying high miR-31 expression and low ARID1A expression had the worst survival. In summary, this study provides novel mechanistic clues demonstrating that miR-31 targets ARID1A to enrich the stemness properties for OSCC progression.

Citation Format: Wen-Cheng Lu, Shu-Chun Lin, Kuo-Wei Chang. miR-31 targets ARID1A to enhance the progression of oral carcinoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 953.
Abstract 957: Regulation of PCGEM1 in castration resistant prostate cancer
Authors: Ho Tsui-Ting , Mo Yin-Yuan
Published: July 2016

PCGEM1 has been shown to be prostate cancer specific long non-coding RNA (lncRNA). Our previous study indicates that PCGEM1 plays a role in castration resistance through regulation of androgen receptor (AR) splice variants. Although it is known that PCGEM1 is often upregulated in prostate cancer, little is known how PCGEM1 is regulated. In the present study, we show that p54/nrb is a positive regulator for PCGEM1. PCGEM1 is higher in castration resistant CWR22Rv1 than in androgen sensitive LNCaP cells. Moreover, androgen deprivation induces PCGEM1 in LNCaP cells. Histone ChIP assays suggest that the active binding site is within 500 bp upstream of putative transcriptional start site. By using biotin-labelled PCR product and pulldown assays combined with mass spectrometry analysis, we show that p54/nrb interacts with the promoter of PCGEM1. While ectopic expression increases, suppression of p54/nrb by RNAi or knockout (KO) suppresses PCGEM1, supporting a role for p54/nrb in PCGEM1 regulation. Moreover, rescue experiments indicate that re-expression of p54/nrb in KO cells restores the ability to induce PCGEM1. Finally, 3, 3’-Diindolylmethane (DIM), a known chemoprevention agent derived from plants such as broccoli, is capable of suppressing PCGEM1 expression by preventing the interaction of p54/nrb with the PCGEM1 promoter. In particular, DIM reduces tumor growth by suppression of PCGEM1 and promoting apoptosis in the castrated xenograft mouse model. Together, these results demonstrate a novel mechanism of how PCGEM1 and AR splicing are regulated. As a result, PCGEM1 may ultimately contribute to castration resistance.

Citation Format: Tsui-Ting Ho, Yin-Yuan Mo. Regulation of PCGEM1 in castration resistant prostate cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 957.
Abstract 966: Circulating exosomal miRNAs as potential biomarker in liposarcoma
Authors: Casadei Lucia , Creighton Chad , Batte Kara , Chelouche-Lev Dina , Croce Carlo M. , Pollock Raphael E.
Published: July 2016

Liposarcoma (LPS) is the most common soft tissue sarcoma histological subtype. Despite the development of combined modality treatments in recent years, a significant proportion of patients respond poorly to chemotherapy, leading to local recurrence or distant metastasis. Thus, early detection of recurrent or metastatic disease or early decision making could improve patient prognosis. However, there are no useful biomarkers for these purposes. Thus, the discovery of novel biomarkers to detect tumors, predict their drug sensitivity, and monitor their progression is one of the most important challenges that must be overcome.

miRNAs are short (approximately 22 nucleotides in length) non-coding RNAs (ncRNAs) that regulate gene expression by binding to specific mRNA targets and promoting their degradation and/or translational inhibition; since their discovery in body fluids (secreted in cell-borne membrane vesicles or associated with macromolecular complexes), circulating blood-borne microRNAs are being investigated as potent minimally invasive biomarkers of several diseases including tumors.

In this study, our goal is to identify potential biomarkers in the peripheral blood of LPS patients that could be useful for LPS diagnostic, prognostic and therapeutic purposes.

To determine the miRNA expression profiles of LPS in peripheral blood (PB) and in peripheral blood exosomes (PBX), a series of 16 human LPS patient specimens and 8 healthy controls was analyzed on a miRNA microarray platform capable of detecting 800 human miRNAs (nCounter Human microRNA expression Assay Nanostring v3). Validation of the array was performed by qRT-PCR on a new set of samples (10 human LPS and 9 healthy controls).

To identify differentially expressed miRNAs, we compared miRNAs expression profiles from PBX of 15 patients to 8 healthy samples. A total of 26 miRNAs were significantly upregulated and 3 miRNAs were significantly down regulated (p<0.004). When we compared miRNA expression profiles from PB to healthy controls, fewer miRNAs were consistently deregulated (18 miRNAs were upregulated and 3 miRNAs were downregulated, p<0.02). Moreover the miRNAs found to be consistently deregulated in PB were all also significantly deregulated in the PBX. Validation of the miRNA panel in an independent cohort of LPS patients confirmed this signature.

Our study has identified a specific miRNA signature in circulating exosomes that may have a novel role in the diagnosis, prognosis and or treatment of LPS patients in a clinical setting.

Citation Format: Lucia Casadei, Chad Creighton, Kara Batte, Dina Chelouche-Lev, Carlo M. Croce, Raphael E. Pollock. Circulating exosomal miRNAs as potential biomarker in liposarcoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 966.
Abstract 967: Vemurafenib regulates miRNA-211 expression in melanoma - effects on extracellular vesicle RNA cargo and function
Authors: Lunavat Taral R. , Cheng Lesley , Einarsdottir Berglind , Lässer Cecilia , Nilsson Jonas A. , Hill Andrew F. , Lötvall Jan
Published: July 2016

In melanoma, more than 50% of the patients harbor BRAF somatic missense mutations, most affecting the V600 region. Vemurafenib, a potent BRAF inhibitor used for the treatment of late stage melanoma, has shown promising results by causing programmed cell death in melanoma cells. Recently, extracellular vesicles including apoptotic bodies, microvesicles and exosomes has been shown to contain genetic information especially RNA with distinct repertoire of RNA, which could be transferred from one cell to another. Here we investigate the effects of Vemurafenib on melanoma cells and the RNA content in their subsets of extracellular vesicles. Upon vemurafenib treatment, we found that the melanoma cells harboring the BRAF mutation significantly increases the RNA and protein cargo in the different subsets of extracellular vesicles. The RNA profile also changes substantially in apoptotic bodies and microvesicles with significant changes in the ribosomal RNA ratio. Further, using small RNA sequencing, we found that extracellular vesicles from treated cells harbor unique miRNAs. The significantly different miRNA detected with sequencing analysis was validated in vitro with PCR and confirmed that miR-211 is upregulated in cells as well as subsets of extracellular vesicles after treatment. Furthermore, miR-211 was validated to be significantly upregulated in tumors tissues harvested from patient derived xenograft (PDX) as well as cell line derived xenograft and its vesicular subsets. This shows that vemurafenib induces miR-211 upregulation in BRAF mutant cells and PDX especially in the subsets of extracellular vesicles. miR-211 induction upon oncogene treatment could be potentially used as a biomarker in melanoma patients with BRAF mutation.

Citation Format: Taral R. Lunavat, Lesley Cheng, Berglind Einarsdottir, Cecilia Lässer, Jonas A. Nilsson, Andrew F. Hill, Jan Lötvall. Vemurafenib regulates miRNA-211 expression in melanoma - effects on extracellular vesicle RNA cargo and function. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 967.
Abstract 973: A long noncoding RNA-regulating enhancer links Ewing sarcoma susceptibility to stress response
Authors: Aryee Dave NT , Tomazou Eleni , Kauer Maximilian , Grünewald Thomas , Tirode Franck , Delattre Olivier , Kovar Heinrich
Published: July 2016

Ewing sarcoma is the second most frequent bone cancer in Caucasian children and young adults, but hardly occurs in Africans. It is characterized by a relatively quiet genome with only one highly recurrent chromosomal aberration, a EWS-ETS (predominantly EWS-FLI1) gene rearrangement. The resulting fusion gene encodes for an aberrant ETS transcription factor which drives sarcomagenesis. In a large GWAS study, three Ewing sarcoma susceptibility loci were discovered on chromosomes 1, 10 and 15, but the molecular mechanisms, how these loci affect tumorigenesis, remained largely unknown.

By epigenomic mapping, EWS-FLI1 ChIP-seq, and reporter gene assays we found one of the susceptibility loci to localize to an EWS-FLI1 bound enhancer in the intron of a long non-coding antisense RNA, which we termed lncESST1 (Ewing Sarcoma Susceptibility Transcript 1). Deletion of the ETS binding region in the enhancer by CRISPR/Cas9 mediated gene editing lead to a drastic decrease in lncESST1 expression. Modulation of lncESST expression by siRNA or enhancer deletion significantly reduced soft agar colony formation of Ewing sarcoma cells without affecting growth under adherent culture conditions. Detachment of Ewing sarcoma cells from the plastic surface and growth under spheroid culture conditions induced stress granule formation as an immediate response, and a steady increase in lncESST1 expression over a period of at least 72 hours. We report that lncESST1 binds to a stress granule component with which it shares the promoter, and that loss of lncESST1 expression due to enhancer deletion perturbs stress granule formation. Our study therefore unveils a novel mechanism of lncRNA mediated cancer susceptibility.

Supported by FWF grant I1225-B19

Citation Format: Dave NT Aryee, Eleni Tomazou, Maximilian Kauer, Thomas Grünewald, Franck Tirode, Olivier Delattre, Heinrich Kovar. A long noncoding RNA-regulating enhancer links Ewing sarcoma susceptibility to stress response. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 973.
Abstract 977: RNA-sequencing analysis implicates novel non-coding RNAs in human papillomavirus-associated head and neck squamous cell carcinoma
Authors: Zou Angela E. , Krishnan Aswini R. , Xuan Yinan , Saad Maarouf A. , Korrapati Avinaash , Advani Sunil J. , Wang-Rodriguez Jessica , Ongkeko Weg
Published: July 2016

Head and neck squamous cell carcinoma (HNSCC) is an aggressive disease marked by frequent recurrence and metastasis and stagnant survival rates. Given the recent emergence of human papillomavirus (HPV) as a major etiological factor in head and neck cancers, there exists a pressing need to better resolve the molecular heterogeneity of HNSCCs, define more precise diagnostic and prognostic tools, and develop a wider repertoire of targeted therapies. Non-coding RNAs (ncRNAs), including long non-coding RNAs (lncRNAs) and microRNAs (miRNAs), have been increasingly viewed as promising sources of insight into cancer onset and progression, with their diverse regulatory roles, tissue specificity, and aberrant expression in various human diseases now established by a multitude of studies. Using RNA-sequencing data from 466 HNSCC patients in The Cancer Genome Atlas (TCGA), we sought to investigate the transcriptome-wide dysregulation of both classes of ncRNAs in HPV-associated HNSCCs. Differential expression analyses controlled for patient smoking status revealed 5 lncRNAs and 9 miRNAs exhibiting significantly altered regulation between HPV-negative and HPV-positive HNSCC tumors (FDR < 1 × 10-10). Among these, 8 ncRNAs were additionally correlated with HNSCC tumor stage, invasion, and anatomic site, among other clinical characteristics (Kruskal-Wallis, p < 0.05). We subsequently validated the dysregulation of several candidate HPV-associated ncRNAs in vitro after ectopically expressing HPV16 E6/E7 in a panel of normal oral epithelial and HPV-negative HNSCC cell lines. To explore the functional consequences of our identified HPV-associated ncRNAs, we further investigated lnc-SP9-1:1, an lncRNA upregulated by 73.89 fold in HPV-positive compared to HPV-negative HNSCCs (FDR < 1 × 10-10). Cell proliferation, migration, and matrigel invasion assays were performed to evaluate the effects of siRNA-mediated lnc-SP9-1:1 knockdown in E6/E7 overexpressing cell lines. While the specific roles of the vast majority of non-coding transcripts remain unknown, their consistent dysregulation in HPV-positive HNSCCs, coupled with their ongoing functional characterization, promises to unravel new layers of understanding of the molecular underpinnings of HPV-associated HNSCCs and nominate new biomarkers and therapeutic candidates for this specific disease.

Citation Format: Angela E. Zou, Aswini R. Krishnan, Yinan Xuan, Maarouf A. Saad, Avinaash Korrapati, Sunil J. Advani, Jessica Wang-Rodriguez, Weg Ongkeko. RNA-sequencing analysis implicates novel non-coding RNAs in human papillomavirus-associated head and neck squamous cell carcinoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 977.