bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 830: Pin1 is a novel target of withaferin A, a mammary cancer chemopreventative steroidal lactone from Withania somnifera
Authors: Samanta Suman K. , Singh Shivendra V.
Published: July 2016

We have shown previously that naturally occurring steroidal lactone withaferin A (WA) inhibits breast cancer development in a transgenic mouse model. Mammary cancer chemoprevention by WA in vivo was associated with suppression of peptidyl-prolyl isomerase Pin1 protein. Pin1 is known to regulate diverse cellular processes including cell proliferation, cell cycle progression, apoptosis, and migration. Moreover, Pin1 is overexpressed in breast tumor and its expression is positively correlated with tumor grade. Thus, we studied functional significance of Pin1 downregulation by WA. Consistent with in vivo observations, Pin1 protein and mRNA expression was decreased upon WA treatment in human breast cancer cells irrespective of ER, HER-2 or p53 status. Overexpression of Pin1 partly reversed the growth inhibitory effect of WA in both MCF-7 (wild-type p53) and SUM159 (mutant p53) cells. However, WA-induced mitotic arrest and apoptosis were partly abrogated only in MCF-7 cells upon ectopic expression of Pin1 perhaps due to p53 stabilization. Molecular docking suggested covalent binding of WA with Cys113 of Pin1, which was confirmed by mass spectrometry. Overexpression of mutant (C113A) Pin1 augmented WA-mediated inhibition of cell survival as well as mitotic arrest and apoptosis in MCF-7 cells indicating that decreased catalytic activity of Pin1 is essential for growth inhibition by WA in breast cancer cells. Taken together, the present study suggests a critical role for Pin1 in WA-mediated growth inhibition of breast cancer. This study was supported by the grant CA142604 awarded by the National Cancer Institute.

Citation Format: Suman K. Samanta, Shivendra V. Singh. Pin1 is a novel target of withaferin A, a mammary cancer chemopreventative steroidal lactone from Withania somnifera. [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 830.
Abstract 837: Reproducibility of efficacy results for chemopreventive agents in the methylnitrosourea-induced rat mammary cancer model
Authors: Steele Vernon E. , Lubet Ronald A. , Grubbs Clinton J.
Published: July 2016

Perhaps the greatest question in biological science is: Can results be reproduced? This problem is of particular concern for a group whose primary mission is to screen for potentially effective agents and rank results by activity. The reproducibility of testing specific agents for chemopreventive activity, testing the same agent in various protocols, and testing multiple agents of a given mechanistic class in the MNU-induced model of ER+ breast cancer in Sprague-Dawley rats is presented. The model employs female virgin Sprague-Dawley rats on a standard 4% Teklad diet which develop ER+ mammary cancers (adenocarcinomas) following MNU injection at 50 days of age (DOA). Preventive agents were given by gavage or diet starting at 55 DOA, and animals monitored for tumor development until approximately 170 DOA. Seven agents which we had previously studied and which were strongly positive [tamoxifen, vorozole (an aromatase inhibitor), Targretin, gefitinib] or strongly negative (naproxen, Lipitor, metformin) were examined in the assays presented. These study results confirmed our previous data. We next tested these same agents in animals given a Western diet (high fat, low calcium) and again obtained the same results. We had thought that the altered physiology and potentially altered pharmacokinetics of rats on a Western diet might alter the observed chemopreventive efficacy. These studies show that the direct repetition of clearly positive and negative agents can be achieved and, in fact, repeated with a significant variation in diet. Importantly, in terms of reproducibility, we find that multiple agents of the same mechanistic class give similar results. Thus, various SERMs (tamoxifen, toremifene, Arzoxifene), RXR agonists (targretin, UAB-30, 4Me-UAB-30), EGFR inhibitors (gefitinib, Erlotinib, lapatinib) are all highly positive (70-95% reduction of cancer multiplicity). In contrast, the negative classes COX inhibitors (naproxen, sulindac, aspirin) or statins (atorvastatin or simvastatin) have been consistently negative (<20% reduction). Therefore, reproducibility of efficacy results appears to be quite solid in our chemoprevention agent development program for rat mammary cancer testing, and we can, with significant certainty, trust our results.

Citation Format: Vernon E. Steele, Ronald A. Lubet, Clinton J. Grubbs. Reproducibility of efficacy results for chemopreventive agents in the methylnitrosourea-induced rat mammary cancer model. [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 837.
Abstract 844: High-resolution microbiome profiling and meta-analysis yields insight into microbial consortia associated with colorectal cancer
Authors: White James R. , Drewes Julia , Sears Cynthia L.
Published: July 2016

Colorectal cancer (CRC) continues to be the second leading cause of cancer-related deaths globally. While multiple environmental factors are associated with incidence of CRC, multiple studies have reported that CRC is frequently linked with an altered colonic microbiota both in terms of taxonomic composition as well as structural organization. However, differences in study design and analysis methodologies among CRC microbiome profiling studies have generated inconsistent findings. To identify bacterial taxa consistently associated with CRC, we applied a uniform microbiome analysis methodology to sequence datasets followed by meta-analysis.

Raw 16S rRNA amplicon sequence datasets from 12 studies including two new Malaysian cohorts were collected and analyzed using the Resphera Insight high-resolution taxonomic assignment protocol. Taxonomic abundances within each dataset were then assessed for significant enrichment or depletion in colorectal carcinoma samples versus controls. Alpha diversity and differences in predicted functional capacity were assessed utilizing QIIME and PICRUSt, respectively.

Several bacterial species were consistently enriched in colorectal carcinoma samples relative to healthy controls including Fusobacterium nucleatum, Fusobacterium necrophorum, Leptotrichia trevisanii, Bacteroides fragilis, Parvimonas micra, Peptostreptococcus stomatis, and Gemella morbillorum. In contrast to other studies that have generally described Fusobacteria as enriched in CRC relative to controls, we identified multiple fusobacterial members that were often not strongly linked with CRC including Fusobacterium varium and Cetobacterium somerae. Evaluating functional predictions inferred from taxonomic composition, we observed consistent enrichment of specific KEGG categories in CRC including glycosyltransferases and carbohydrate digestion and absorption.

Applying a uniform high-resolution analysis protocol for microbiome profiling enabled detection of specific species that were strongly associated with CRC across multiple studies. These findings demonstrate that higher level analyses (e.g. those at the genus or family level) may be insufficient for detection of indicator taxa in some populations.

Citation Format: James R. White, Julia Drewes, Cynthia L. Sears. High-resolution microbiome profiling and meta-analysis yields insight into microbial consortia associated with colorectal 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 844.
Abstract 85: Macrophage inflammatory protein-1β and interleukin 15 modulate gene expression patterns associated with prostate cancer progression
Authors: Rohena-Rivera Krizia , Aponte-Colón Diana A. , Sánchez-Vázquez María M. , Cadilla Carmen L. , Martínez-Ferrer Magaly
Published: July 2016

Prostate Cancer (PCa) is the second-leading cause of cancer-related deaths in the United States. Inflammation is associated with PCa development and progression. Cytokines such as macrophage inflammatory protein-1β (MIP1β or CCL4) and Interleukin 15 (IL-15) are differentially expressed in prostate cancer patients with recurrent disease (MIP1β) or recurrence-free survival (IL-15). However, the specific pathway by which these cytokines affect PCa progression is unknown. We evaluated the changes in gene expression patterns using in vivo models of prostate cancer and microarray analysis. To generate prostate tumors in vivo we used a mouse orthotopic xenograft model. Androgen-receptor-positive (AR+) cells, 22RV1, were orthotopically injected in the anterior prostate lobes. MIP1β and IL-15 were administered bi-weekly with intraperitoneal injections during 4 weeks. Tumor tissue was collected and snap frozen for RNA extraction, and microarray analysis with the Affymetrix Human Gene 2.0 Gene Array. Differences in gene expression were analyzed with Ingenuity Pathway Analysis (IPA) software and confirmation was done via real-time PCR. Microarray analysis showed 179 genes differentially expressed between MIP1β and control. In addition, 952 genes were differentially expressed between IL-15 and control. Ingenuity Pathway Analysis revealed that MIP1β affected networks associated with cell development, proliferation and movement while IL-15 affected networks involved in lymphocyte development and movement, cell death, and the inhibition of cancer cell invasion. Knowledge of the role of MIP1β and IL-15 in the alteration of gene expression patterns is pertinent to elucidate the significance of these cytokines in prostate cancer progression.

Citation Format: Krizia Rohena-Rivera, Diana A. Aponte-Colón, María M. Sánchez-Vázquez, Carmen L. Cadilla, Magaly Martínez-Ferrer. Macrophage inflammatory protein-1β and interleukin 15 modulate gene expression patterns associated with prostate cancer progression. [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 85.
Abstract 857: Diagnostic test system for sensitive, specific and reproducible detection of EML4-ALK RNA fusion transcripts in the blood of patients with NSCLC
Authors: Mellert Hestia , Jackson Leisa , Maar Dianna , Shelton Dawne , Pestano Gary
Published: July 2016

Clinical testing for the detection of RNA fusions in tissue currently include FISH, IHC, PCR and NGS. However, approximately 30% of patients with advanced non-small cell lung cancer (NSCLC) are not candidates for tissue biopsies and in some cases where tissue is obtained, it is not always of sufficient quantity for molecular testing. For these reasons, the detection of nucleic acids in circulation has become increasingly relevant to clinical testing. In this study, we have focused on the development of an EML4-ALK diagnostic test system that includes the prospective collection of whole blood and the reproducible detection of mRNA fusion transcripts by a PCR-based technology. The focus was on the detection of EML4-ALK transcripts from donors with and without previously diagnosed NSCLC. Pre-analytic complexity was reduced by restricting the handling of samples to 72 hours from time of sample receipt to test result. Recovery methods for RNA extraction from donor plasma were then optimized to enrich for RNA recovered from both circulating-free, and RNA within blood vesicles, including platelets and exosomes. A two-step reverse transcription and Droplet Digital™ PCR (RT-ddPCR) method was evaluated in detection testing using a multiplexed EML4-ALK assay that includes variants 1, 2 and 3. Analytic assay specificity and sensitivity was examined for RT-ddPCR efficiency using a cell-line positive for EML4-ALK variant 1 and in vitro RNA designed to mimic variants 1, 2, and 3. Analytic sensitivity was determined to be 0.2% of fusion RNA spiked into a background of normal plasma. Precision studies were conducted with two different amounts of input RNA (high and low), over three consecutive days, three runs in one day and with two operators. The SD of detected fusion transcripts in this study did not exceed 25%. Finally, normal (n = 10) and FISH positive NSCLC donor (n = 9) plasma samples were processed for the recovery of RNA and tested for EML4-ALK fusions. All FISH positive cases were accurately detected by the RT-ddPCR test with EML4-ALK variant copies ranging from 13 to 150 copies. EML4-ALK fusion transcripts were not identified in normal donor plasma. We conclude that the developed test system is highly suited for a 72 hour test to result turnaround, reproducible and sensitive detection of diagnostic fusion RNA variants, including EML4-ALK in blood in the clinical laboratory.

Citation Format: Hestia Mellert, Leisa Jackson, Dianna Maar, Dawne Shelton, Gary Pestano. Diagnostic test system for sensitive, specific and reproducible detection of EML4-ALK RNA fusion transcripts in the blood of patients with NSCLC. [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 857.
Abstract 881: Dissecting chromatin dynamics in malignant progression
Authors: Yang Hanseul , Schramek Daniel , Adam Rene , Levorse John , Keyes Brice , Wang Ping , Zheng Deyou , Fuchs Elaine
Published: July 2016

Tumor-initiating cells within a cancer exhibit distinct patterns of transcription factors and gene expression compared to stem cells within their healthy tissue. Little is known about the key transcription factors that dictate chromatin remodeling and the accompanying transcriptional changes that ultimately hardwire the malignant behavior of tumor-initiating stem cells. Here, by in vivo chromatin and transcription profiling, we show that dramatic shifts in large open-chromatin (super-enhancer) landscapes underlie these differences. Focusing on one of the most common and life-threatening cancers world-wide, squamous cell carcinoma (SCC), we show that super-enhancers of SCC-stem cells contain the binding sites for a distinct set of putative master transcription factors. Many of their genes, including Ets2 and Elk3, are themselves regulated by SCC super-enhancers, suggesting a cooperative feed-forward loop. Malignant progression requires these genes, whose knockdown severely impairs tumor growth and prohibits progression from benign papillomas to SCCs. Interestingly, ETS2 is known to be phosphorylated and activated by RAS/MAPK signaling, and knockdown of ETS2 results in loss of expression of SCC super-enhancer-associated genes. Conversely, in vivo forced expression of an active ETS2 version harboring a phosphomimetic substitution at the MAPK-activation residue is sufficient to trigger cellular transformation without oncogenic RAS. Moreover, ETS2-overactivation in epidermal progenitors rewires the super-enhancer landscape and induces SCC super-enhancer associated genes including Fos, Junb, Klf5 and Elk3. Finally, we identify dramatic remodeling of the Cxcl1/2 locus from an H3K27me3-repressed to an ETS-regulated, super-enhancer-activated state, and provide evidence for their autocrine oncogenic role in SCCs through a cognate receptor Cxcr2. Together, our findings unearth an essential regulatory network required for the SCC chromatin landscape and unveil its importance in malignant progression.

This work was funded by grants from the National Institutes of Health (R01-AR31737) and NYSTEM #CO29559.

Citation Format: Hanseul Yang, Daniel Schramek, Rene Adam, John Levorse, Brice Keyes, Ping Wang, Deyou Zheng, Elaine Fuchs. Dissecting chromatin dynamics in malignant progression. [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 881.
Abstract 901: Transgenerational inheritance of increased mammary cancer risk in the offspring of high fat diet fed dams: Changes in oxidative stress pathways
Authors: Nguyen Nguyen M. , Andrade Fabia O. , DeAssis Sonia , Cruz Idalia , Benitez Carlos , Godschalk Roger , Hilakivi-Clarke Leena
Published: July 2016

Maternal high fat diet has been shown to increase mammary cancer risk among female offspring. We investigated here if the increase is seen across multiple generations. For that purpose, pregnant C57BL/6NTac dams (F0) were fed either a control (CON, 6% corn oil) or an isocaloric high n-6 polyunsaturated fat (HF) diet (18% corn oil) between gestation days 8 and 21. Offspring in subsequent generations (F1-F3) were not exposed to any further dietary modifications, and kept on CON diet. Mammary tumors were initiated by priming the mammary glands with 15 mg medroxyprogesterone (MPA) at week 6, followed by a 1 mg dose of 7,12-dimethylbetz(a)anthracene (DMBA) given on weeks 7-9 for a total of 3 doses. Animals were monitored for 20 weeks after the final dose of DMBA for tumors. Both F1 and F3 generation female HF offspring exhibited a significantly higher mammary tumor incidence and tumor burden than the control offspring. In F3 generation, the increase was seen from week 12 of tumor monitoring until the end of the study. The increase in risk was preceded by a higher number of terminal end buds in the F1 and F3 generation mammary glands. Malondialdehyde (MDA) adduct levels were found to be significantly elevated in the HF group of F3 offspring, suggesting that the increased mammary cancer risk in this generation was attributed to oxidative stress. Western blot analysis of mammary glands identified significantly elevated Keap1 (a repressor of the antioxidant response) and PERK levels in HF F3 offspring; and elevated levels of Beclin-1 and p62 in both F1 and F3 generations compared with CON offspring, corroborating with the idea that oxidative stress was the culprit. miRNA analysis of F3 mammary glands showed decreased expression of mir-324-5p, mir-136 and mir-378 that target Keap1 and p62, potentially explaining the increase in Keap1 and p62 levels. Our results suggest that oxidative stress and antioxidant response plays a role in explaining transgenerational increase in mammary cancer risk among offspring exposed to high fat diet through F0 dams.

Citation Format: Nguyen M. Nguyen, Fabia O. Andrade, Sonia DeAssis, Idalia Cruz, Carlos Benitez, Roger Godschalk, Leena Hilakivi-Clarke. Transgenerational inheritance of increased mammary cancer risk in the offspring of high fat diet fed dams: Changes in oxidative stress pathways. [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 901.
Abstract 905: Pterostilbene (PTER) suppresses breast cancer brain metastasis by targeting a c-Met mediated inflammation network
Authors: Xing Fei , Liu Yin , Sharma Sambad , Wu Kerui , Watabe Kounosuke
Published: July 2016

At the late stage of breast cancer, most patients develop metastatic lesions and the brain is one of the major metastatic sites. Brain metastasis profoundly affects the cognitive and sensory function as well as morbidity of patients, and the one year survival rate among these patients remains less than 20%. The result of our newly developed gene set enrichment analysis (GSEA)-based pathway screening indicates that c-Met pathway is highly activated among the patients who developed early brain metastasis. To identify a novel target of c-Met pathway in brain metastasis, we performed the microarray analysis in two brain metastatic cell lines with the expression of doxycycline-inducible sh-cMet. Our results indicate that a group of inflammatory cytokines including IL1b, IL8 and CXCL1 were significantly induced by c-Met activation. Furthermore, we found that IL1b was able to induce the secretion of HGF (hepatocyte growth factor) from tumor associated astrocytes (TAA) which in turn activates c-Met pathway in cancer cells. Our results of endothelial cell tube formation assays also strongly suggest that cMet-induced IL8 and the activation of its receptor, CXCL1, promote tumor angiogenesis which is essential for the metastatic growth of cancer cells. Natural compounds (NC) have been extensively studied for their anti-tumor effects. However, much less studies have been done on NC for the treatment/prevention of brain tumors mainly due to the obstacle of Blood-brain-barrier (BBB) permeability. To identify the NCs that can be used for treating brain metastasis, we performed pathway analysis which only focused on BBB-permeable NC. The results of GSEA indicate that Resveratrol-targeting genes were significantly enriched among those patients who developed brain metastasis compared to metastasis-free patients. Furthermore, we found that the one of derivatives of Resveratrol, Pterostilbene (PTER), showed significantly more potent activity than Resveratrol and suppressed brain metastasis in vitro and in vivo by targeting the c-Met oncogene. These findings suggest that TAA-mediated c-Met activation plays a key role in brain metastasis and that PTER is a potential therapeutic agent to treat brain metastasis by suppressing the c-Met expression.

Citation Format: Fei Xing, Yin Liu, Sambad Sharma, Kerui Wu, Kounosuke Watabe. Pterostilbene (PTER) suppresses breast cancer brain metastasis by targeting a c-Met mediated inflammation network. [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 905.
Abstract 906: Exosomes from mutant TP53 cancer cells polarize tumor associated macrophages
Authors: Cooks Tomer , Pateras Ioannis S. , Patel Keval M. , Forshew Tim , Gorgoulis Vassilis G. , Harris Curtis C.
Published: July 2016

Introduction: TP53 mutants are involved in the pathogenesis of most solid tumors and are known to gain oncogenic functions distinct from their original wild-type form. The existence of such gain-of-function (GOF) activities is supported by ample evidence, however only in a cell-autonomous fashion. Since tumor-associated-macrophages (TAM) are also a hallmark of solid tumors typically correlated with poor prognosis, we investigated the link between mutations in the TP53 gene (mutp53) occurring in epithelial tumor cells and the formation of a surrounding TAM population in-situ.

Methods: By designing a co-culture system we incubated human primary monocytes together with colorectal cancer (CRC) cells differing in their p53 status. Relevant macrophages markers were evaluated on RNA level and protein level. In addition, co-cultured macrophages were subjected to various functional assays (phagocytosis, migration, and invasion). In attempt to confirm clinical relevance, samples from a cohort of human CRC patients were analyzed using genomic and immunohistochemical methods. To identify the interaction between the tumor cells and the macrophages, we isolated exosomes from the CRC cells and subjected them to a Nanostring analysis to learn about their microRNAs composition.

Results: In this study, we discovered that mutp53 exerts a non-cell-autonomous effect over neighboring macrophages by using specific microRNAs (miRs) which are shuttled through an exosomal transfer resulting with a phenotype change of the affected macrophages. Mutp53-specific exosomes containing cargoes such as miR-1246 were shown to be used by macrophages at the receiving end, thus promoting the formation of TAM subset also observed in surgical specimens resected from cancer patients.

Conclusions: Altogether, these findings are consistent with a microenvironmental role for specific “hot-spot” p53 mutants tightening the interaction between the tumor cell and the immune compartment. Additionally, this study is the first to show a non-cell-autonomous role played by mutant p53 - the most common form of mutation found in human cancers. Deciphering the intricate regulation shared by the tumor cell and its surrounding macrophages may give rise to novel prognostic and diagnostic tools as well as to therapeutic approaches targeting TAM, specific tumor-promoting miRs and mutp53-specific subsets of exosomes.

Citation Format: Tomer Cooks, Ioannis S. Pateras, Keval M. Patel, Tim Forshew, Vassilis G. Gorgoulis, Curtis C. Harris. Exosomes from mutant TP53 cancer cells polarize tumor associated macrophages. [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 906.
Abstract 907: Brain microenvironment induced PTEN loss by microRNAs promotes brain metastasis
Authors: Zhang Lin , Zhang Siyaun , Yao Jun , Lowery Frank J. Lowery , Zhang Qingling , Huang Wen-Chien , Li Ping , Li Min , Wang Xiao , Zhang Chenyu , Wang Hai , Ellis Kenneth , Cheerathodi Mujeeburahiman , McCarty Joseph , Palmieri Diane , Steeg Patricia , Saunus Jodi S , Lakhani Sunil , Huang Suyun , Sahin Aysegul , Aldape Kenneth , Yu Dihua
Published: July 2016

Metastasis is the number one cause of cancer-related mortality. Major neoplastic diseases such as melanoma, lung, breast, and colon cancers have high incidences of brain metastases. One-year survival after diagnosis of brain metastasis is less than 20%. Cancer cells dynamically interacts with specific organ microenvironments to establish metastasis as depicted by the “seed and soil” hypothesis. Yet it is unclear when and how disseminated tumor cells acquire the essential traits from the brain microenvironment that primes their subsequent metastatic outgrowth.

Here we found that primary tumor cells with normal PTEN expression lose PTEN after dissemination to the brain, but not to other organs. Metastatic brain tumor cells that have experienced PTEN loss have PTEN levels restored once they leave the brain. This brain microenvironment-dependent, reversible PTEN mRNA and protein down-regulation is epigenetically regulated by microRNAs (miRNAs) from astrocyte-derived exosomes. Furthermore, this adaptive PTEN loss in brain metastatic tumor cells leads to an increased secretion of cytokine chemokine (C-C motif) ligand 2 (CCL2), which recruits Iba1+ myeloid cells that reciprocally enhance outgrowth of brain metastatic tumor cells via enhanced proliferation and reduced apoptosis.

Our findings signify the dynamic and reciprocal cross-talk between tumor cells and other brain stromal cells. Disseminated tumor cells acquire the essential traits from the microenvironment of brain that prime their outgrowth. Importantly, our finding provides new opportunities for effective anti-metastasis therapies: inhibiting CCL2 might be an effective therapeutic intervention of life-threatening brain metastases.

Citation Format: Lin Zhang, Siyaun Zhang, Jun Yao, Frank J. Lowery Lowery, Qingling Zhang, Wen-Chien Huang, Ping Li, Min Li, Xiao Wang, Chenyu Zhang, Hai Wang, Kenneth Ellis, Mujeeburahiman Cheerathodi, Joseph McCarty, Diane Palmieri, Patricia Steeg, Jodi S Saunus, Sunil Lakhani, Suyun Huang, Aysegul Sahin, Kenneth Aldape, Dihua Yu. Brain microenvironment induced PTEN loss by microRNAs promotes brain metastasis. [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 907.
Abstract 912: Mist1+ secretory progenitor cells can give rise to cancer in the intestine and colon
Authors: Hayakawa Yoku , Sakitani Kosuke , Kim Woosook , Tailor Yagnesh , Nagar Karan , Koike Kazuhiko , Asfaha Samuel , Wang Timothy C.
Published: July 2016

Intestinal crypts are maintained by long-lived intestinal stem cells (ISCs) which reside near the base of glands. Above the ISC pool, there are short-lived progenitors that can supply lineage-specific differentiated cell types into the villus. Notch and Wnt pathways play a key role for determining the stem/progenitor cell function and their cell fate, and regulating cancer development in the gastrointestine. Although it has been well established that ISCs are a major origin of intestinal cancer, it remains unknown whether intestinal progenitor population can give rise to cancer. We use Mist1-CreERT mice and induce Notch and Wnt activation by mating with LSL-Notch1-IC mice and/or Apc flox mice. We found that a bHLH transcriptional factor Mist1 is expressed in Paneth cells (Lysozyme+CD24high) as well as short-lived secretory progenitors (Lysozyme-CD24low) in the intestine. Mist1+ secretory progenitors are radio- and chemo-resistant, but do not show stem cell interconversion even after intestinal injury or Lgr5 ablation. However, aberrant Notch activation changes Mist1+ cells to Lgr5+ long-lived enterocyte progenitors, with loss of secretory lineage differentiation. Mist1+CD24low progenitor population can form intestinal organoids with Notch activation in vitro. Mist1+ secretory progenitors can give rise to intestinal cancer by simultaneous Notch and Wnt activation by loss of Apc gene, while loss of Apc in Mist1 lineage alone does not develop cancer. In the colon, Mist1 marks colonic secterory progenitors that become to lineage trace and can be a cancer-initiating cell after Notch activation or DSS-induced colonic injury. These results provide the clear evidence of cellular plasticity dependent on Notch signaling in the gut, and suggest short-lived progenitors as another cellular origin of cancer besides ISC pool.

Citation Format: Yoku Hayakawa, Kosuke Sakitani, Woosook Kim, Yagnesh Tailor, Karan Nagar, Kazuhiko Koike, Samuel Asfaha, Timothy C. Wang. Mist1+ secretory progenitor cells can give rise to cancer in the intestine and colon. [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 912.
Abstract 916: Slit2 inhibits breast cancer growth and metastasis by modulating tumor microenvironment
Authors: Ahirwar Dinesh K. , Nasser Mohd W. , Elbaz Mohamad , Shilo Kontestine , Ganju Ramesh
Published: July 2016

Breast cancer (BC) continues to be a major health issue particularly in developed countries. The Slit2 gene is hyper-methylated in BC and acts through Roundabout Homolog1 (Robo1) receptor. Slit2/Robo1 signaling has been shown to inhibit the migration of a variety of cells including BC cells. Recently, it has also been reported to improve chemotherapy outcome in intestinal cancer. However, Slit2/Robo1-mediated mechanisms that regulate tumor growth and metastasis are not well known. Using syngeneic mouse model, we report that the over-expression of Slit2 using Adeno-Slit2 transduction successfully restrains tumor growth and also restrict the metastasis to lung, compared to Adeno-null control. To explore the Slit2 mediated cellular events, we analyzed the stromal cells recruitment to the tumor and observed reduced number of tumor associated macrophages (TAMs) in the Adeno-Slit2 group compared to Adeno-null group. We further analyzed the underlying molecular mechanisms and found that the soluble Slit2 inhibits TGF-β1 mediated alternative activation of macrophages to TAMs. Treatment with TGF-β1 significantly enhances the markers of TAMs by upregulating β-catenin levels. Interestingly, we observed that Soluble Slit2 inhibits TGF-β1 mediated activation of TAMs by inhibiting downstream signaling molecules Smad2 and β-catenin. We also analyzed Slit2 expression in human breast cancer TMA and an inverse correlation of Slit2 expression was observed with the incidence of breast cancer, metastasis and macrophage recruitment. This study highlights the ability of Slit2 to prevent macrophage alternative activation to TAMs, thereby restrict tumor growth and lung metastasis. Anti-cancer activity of Adeno-Slit2 suggests therapeutic potential of Slit2 to treat advanced breast cancer.

Citation Format: Dinesh K. Ahirwar, Mohd W. Nasser, Mohamad Elbaz, Kontestine Shilo, Ramesh Ganju. Slit2 inhibits breast cancer growth and metastasis by modulating tumor microenvironment. [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 916.
Abstract 921: Pericellular-interactions promote malignant progression of glioblastoma via novel NOTCH-1/HIF-1α pathway
Authors: Kim Eunsoo , Lee Jungwhoi , Ryu Seung-Wook , Choi Chulhee , Choi Kyungsun
Published: July 2016

Malignant tumors generally develop hypoxic condition in the core region of tumor mass, which induces stabilization of hypoxia inducible factor-1 alpha (HIF-1α) by preventing proteasome-dependent degradation. HIF-1α has been shown to regulate various oncogenes for malignant progression of many types of tumors including glioblastoma multiforme (GBM). In the present study, we showed that cell-to-cell interactions at high density can induce malignant transformation of tumor cells by activation of HIF-1α even under normoxic condition. To delineate the biological effect of cell culture density, we maintained cancer cells in different density. Four types of GBM cell lines (U251-MG, LN215-MG, U373-MG, and CRT-MG) were used to examine the oncogenic phenotypes of cancer cells cultured in different densities. Cells cultured in a high-density condition showed higher rate of proliferation and migration activity and higher resistance to temozolomide, an anti-cancer drug commonly used for GBM treatment. We found that the pericellular interactions stimulate the cleavage of notch-1 intracellular domain (NICD), and the cleaved NICD protects the HIF-1α from hydroxylation and subsequent proteasomal degradation, resulting in nuclear translocation of HIF-1α. Collectively, we suggest that the cell-cell interactions in high-density culture conditions induce NICD-mediated HIF-1α activation, which leads to malignant phenotypic changes. Therefore, treatment of gamma-secretase inhibitor (GSI), which can block NOTCH-1 signaling pathway, in combination with TMZ could be a promising therapeutic strategy in TMZ-resistant tumor cells.

This research was supported by a grant of the Korea Health Technology R&D Project through the Korea Health Industry Development Institute (KHIDI), funded by the Ministry of Health & Welfare, Republic of Korea (grant number : HI14C0042)

Citation Format: Eunsoo Kim, Jungwhoi Lee, Seung-Wook Ryu, Chulhee Choi, Kyungsun Choi. Pericellular-interactions promote malignant progression of glioblastoma via novel NOTCH-1/HIF-1α 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 921.
Abstract 922: PGE2/HuR/CEBPD axis in macrophages contributes to immunosuppression and inhibits phagocytosisof nasopharyngeal carcinoma
Authors: Hsiao Yu-Wei , Wang Ju-Ming
Published: July 2016

The association of inflammation and cancer progression via modulation of phagocytosis and suppression of adaptive immunity by tumor-associated macrophages (TAM) to promote tumorigenesis has been implicated in a number of recent studies. To date, the underlying mechanisms involving the interactions of these two cellular processes remain elusive, particularly in the involvement of posttranscriptional regulation. In this study, we demonstrate that prostaglandin E2 (PGE2) activates transcription factor CCAAT/enhancer binding protein delta (CEBPD) in macrophages by inducing nucleocytoplasmic shuttling of RNA binding protein Hu antigen R (HuR), which plays a direct role in stabilizing CEBPD mRNA in macrophages. Increase of CEBPD resulted in elevated expression of IL-10, a well-studied immunosuppressor, and PTX3, which contribute to suppression of phagocytosis of cancer cells by macrophages. In addition, the conditioned medium of CEBPD-transfected macrophages showed an immunosuppressive effect to attenuate phagocytosis of cancer cells by macrophages, suggesting an autocrine mode of regulation. Immunohistochemistry studies demonstrated that the cytosolic level of HuR protein correlates with increased CEBPD in TAM and malignant nasopharyngeal carcinoma (NPC). Collectively, these results provide new evidences that the inflammatory PGE2/HuR/CEBPD axis in macrophages plays protumor roles in controlling immunosuppression and attenuating phagocytosis of cancer cells by macrophages.

Citation Format: Yu-Wei Hsiao, Ju-Ming Wang. PGE2/HuR/CEBPD axis in macrophages contributes to immunosuppression and inhibits phagocytosisof nasopharyngeal 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 922.
Abstract 933: Protein kinase a regulation of malignant brain tumor migration
Authors: Ausland Jennifer N. , Newman Robert H. , Martin Patrick M.
Published: July 2016

Glioblastoma multiforme (GBM) is the most common and deadly type of primary brain tumor, accounting for over 13,000 new cases in the U.S. annually and a two-year survival rate of < 5%. GBMs are highly invasive and classified by the World Health Organization as a grade IV astrocytoma. Recent studies have shown that protein kinase A (PKA), a key regulator of signal transduction and various biological functions, is essential for maintaining polarity during cell migration and that localized activation of PKA at the leading edge is an early event in directional cell migration of other cell types. Though PKA has previously been shown to promote proliferation and migration in lung, colorectal, adrenocortical, and prostate cancer cell lines, the molecular mechanisms that link PKA activity with GBM physiology remain unclear. Wound healing assays in human GBM cell lines U-251 MG, U-87 MG and A172, demonstrated that GBM cell migration increased following the addition of 20 μM epidermal growth factor (EGF). Similar results were found with 5 μM treatment of the adenylate cyclase activator, forskolin, and inhibited by pre-treatment with 10 μM of the PKA inhibitor, H89. In order to assay the amount of PKA activity, we measured the phosphorylation of cAMP response element binding protein (CREB) in these cell lines. Western blot studies revealed an increase in CREB phosphorylation in cells treated with EGF and forskolin. Phosphorylation of CREB was decreased with the addition of H89 treatment. Our western blot analysis supports our wound healing data demonstrating a correlation between increased PKA-mediated CREB phosphorylation and migration in the absence of serum. Taken together, these data suggest that PKA activation by EGF is involved in GBM cell migration. Moreover, our findings describe a mechanism by which PKA regulates the migration of human brain tumors.

Citation Format: Jennifer N. Ausland, Robert H. Newman, Patrick M. Martin. Protein kinase a regulation of malignant brain tumor migration. [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 933.
Abstract 939: Cell-free DNA associated with extracellular vesicle: Biomarker or bioactivity
Authors: Shelke Ganesh V. , Jang Su Chul , Yin Yanan , Lässer Cecilia , Lötvall Jan
Published: July 2016

Extracellular vesicles (EVs) including exosomes transfer bioactive cargo, including proteins and nucleic acids. Recently, EVs associated DNA were shown to be surrogate biomarker present in body fluids harboring various oncogenic mutations. Cell-free, extracellular DNA is reported in cell cultures and many body fluids, but its relative location in relation to EVs and function is not known. Our study demonstrates that DNase sensitive nucleic acids were present on the surface of EV isolates from mast cell lines. Association of EV-DNA was revealed by increase of zeta potential and particle number upon DNase treatment. Additionally, cytoplasmic DNA traces were found in recipient mesenchymal stem cell after EVs treatment. In conclusion, we define that surface association of DNA with EVs that could regulate recipient cellular function.

Citation Format: Ganesh V. Shelke, Su Chul Jang, Yanan Yin, Cecilia Lässer, Jan Lötvall. Cell-free DNA associated with extracellular vesicle: Biomarker or bioactivity. [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 939.
Abstract 945: Tissue tranglutaminase-2 promotes gastric cancer progression via ERK1/2 pathway
Authors: Su Liping , Wang Xiaofeng , Zhou Quan , Wu Xiongyan , Liu Bingya
Published: July 2016

Gastric cancer is one of the most common tumors worldwide with extensive local tumor invasion, metastasis and poor prognosis. Thus, understanding the mechanisms regulating progression of gastric cancer is the key for developing effective therapeutic strategies. Tissue tranglutaminase-2 (TG2), a multifunctional member of the transglutaminase (TGase) family, has already been proved to play an important role in tumor initiation and progression. However, the specific role and underlying molecular mechanisms of TG2 in the progression of gastric cancer (GC) remain unknown. Here we showed that TG2 was highly expressed in GC tissues and positively associated with depth of tumor invasion and late TNM stage. With gain and lose-of function approaches, we found that TG2 promotes GC cell proliferation, migration and invasion, as well as tumorigenesis and peritoneal metastasis in vivo. These events were associated with activating of ERK1/2 pathway, and ERK1/2 inhibitor U0126 inhibited cell proliferation, migration and invasion induced by overexpression of TG2. Taken together, these findings suggest that TG2 contributes to tumorigenesis and progression of GC by activating the ERK1/2 signaling pathway and is a potential therapeutic target of metastatic gastric cancer.

Citation Format: Liping Su, Xiaofeng Wang, Quan Zhou, Xiongyan Wu, Bingya Liu. Tissue tranglutaminase-2 promotes gastric cancer progression via ERK1/2 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 945.
Abstract 954: Analysis of noncoding RNA expression in a mouse model of PTEN-deficient prostate cancer
Authors: De Velasco Marco A. , Kura Yurie , Sakai Kazuko , Hatanaka Yuji , Fujita Yoshihiko , Togashi Yosuke , Terashima Masato , Yoshikawa Kazuhiro , Nishio Kazuto , Uemura Hirotsugu
Published: July 2016

Prostate cancer is a heterogeneous disease that is driven by combined genetic and epigenetic alterations. A significant portion of the mammalian genome consists of non-coding regions of RNA. Increasing evidence has shown that these noncoding RNAs (ncRNAs) have significant roles in the epigenetic regulation of several cellular processes, and their dysregulation can contribute to tumorigenesis and promote disease progression in many cancer types. In order to gain better insights into the potential roles of ncRNAs in prostate cancer, we used a genetically engineered mouse model of prostate cancer to perform a comparative analysis of the cancer transcriptome and the landscape of ncRNAs in castration-naïve prostate tumors and the progression to castration-resistant disease. Whole transcription arrays were used to explore both messenger (mRNA) and long intergenic non-coding RNA (LincRNA) transcript expression in normal prostate tissue from wildtype mice and prostate tumors from conditional PTEN-knockout mice. Comparative analyses were performed between prostate tumor samples from castration-naïve mice and, at 4 weeks and 10 weeks post-surgical castration. Altered expression of ncRNAs was most commonly observed and accounted for 56.2% (2370/4216), 47.6% (4460/9366) and 41.57% (1545/3717) of the total genes differentially expressed between normal mouse prostate and prostate tumors from castration-naïve, 4 weeks post-castration and 10 weeks post-castration, respectively. We also profile the expression of relevant cancer-related lincRNAs present in these models. Overall, our analyses provide data to support a role of LincRNA dysregulation in the pathogenesis of PTEN-deficient prostate cancer, and suggest that these mouse models might provide a potential preclinical tool to test candidate prospective therapies targeting ncRNAs.

Citation Format: Marco A. De Velasco, Yurie Kura, Kazuko Sakai, Yuji Hatanaka, Yoshihiko Fujita, Yosuke Togashi, Masato Terashima, Kazuhiro Yoshikawa, Kazuto Nishio, Hirotsugu Uemura. Analysis of noncoding RNA expression in a mouse model of PTEN-deficient 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 954.
Abstract 975: Hypoxic long noncoding RNA NDRG1-OT1 inhibit NDRG1 in MCF-7 breast cancer cell line
Authors: Lin Hsin-Chen , Lai Liang-Chuan , Chen Hsing-Guang , Chen Hung-Hsin , Chang Mei-Hung , Tsai Mong-Hsun , Chung Eric Y.
Published: July 2016

Hypoxia is a crucial factor that associated with solid tumor progression by driving multiple signaling pathway. Our lab previously reported that NDRG1 was oxygen-responsive genes and could affect cellular function in a breast cancer cell line MCF-7. Long-non coding RNAs (lncRNAs) have recently emerged as important regulators of tumor progression by participating in several process of gene expression. However, it is unclear whether lncRNAs are responsed to oxygen concentrations in breast cancer. Therefore, the aim of this study is to identify lncRNAs which are responsive upon changes in oxygen concentrations, and its regulatory mechanism in the breast cancer MCF-7 cells. We investigated the differentially expressed lncRNAs in different oxygen concentrations using next-generation sequencing (NGS) and validated these results through quantitative reverse transcription polymerase chain reaction (qRT-PCR). We found an lncRNA, NDRG1-OT1, that was up-regulated during hypoxia and down-regulated during reoxygenation. Next, we identified the differentially expressed genes regulated by NDRG1-OT1 using microarrays and confirmed these results through qRT-PCR and Western blotting. We found an oxygen responsive gene, NDRG1, was inhibited by NDRG1-OT1. Furthermore, Ingenuity Pathway Analysis(IPA) indicated that NDRG1-OT1 could involve in the Ubiquitin-proteasome pathway. We considered NDRG1-OT1 might regulate NDRG1 stability and ubiquitination, and we are proving these possibility by immunoblotting and immunoprecipitation. These findings may provide new insights into epigenetic regulation of breast cancer during hypoxia.

Citation Format: Hsin-Chen Lin, Liang-Chuan Lai, Hsing-Guang Chen, Hung-Hsin Chen, Mei-Hung Chang, Mong-Hsun Tsai, Eric Y. Chung. Hypoxic long noncoding RNA NDRG1-OT1 inhibit NDRG1 in MCF-7 breast cancer cell line. [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 975.