bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3339: Muscarinic acetylcholine receptor subtype 3 regulates gastric stem cell expansion and gastric cancer progression by controlling YAP activation
Authors: Hayakawa Yoku , Konishi Mitsuru , Sakitani Kosuke , Koike Kazuhiko , Wang Timothy
Published: July 2017

Within the gastrointestine, nerves help to regulate both normal and neoplastic stem cell dynamics. Several previous studies suggested that cholinergic nerve signaling plays an important role in gastrointestinal cancer development, but the exact underlying mechanism has not been clarified. In this study, we examined the role of muscarinic acetylcholine receptor subtype 3 (M3R) in gastric homeostasis and cancer development by using mouse models and human cancer cell lines. In situ hybridization revealed M3R expression in gastric stem cell zone, and its expression was markedly upregulated in gastric cancer cells. We knocked out M3R in Lgr5+ gastric stem cells in Lgr5-CreERT; M3Rflox/flox mice, and found that deletion of M3R inhibited clonal expansion of Lgr5+ cells in regenerative states. In a gastric tumor model of Mist1-CreERT; Apcflox/flox mice, knockout of M3R dramatically suppressed tumor growth. RNA sequencing analysis of these tumors revealed that several important pathways were significantly inhibited in M3R knockout samples, including YAP/TAZ pathway. We established M3R-expressing gastric cancer cell lines, and western blotting, luciferase assay, and RT-PCR analysis confirmed that acetylcholine (ACh) agonist activates YAP pathway through M3R. YAP is upregulated in approximately half of gastric cancer patients, and its expression is significantly associated with disease stage and histological form. This M3R-YAP axis activates the gastric stem cell niche and offers a compelling target for tumor treatment and prevention.

Note: This abstract was not presented at the meeting.

Citation Format: Yoku Hayakawa, Mitsuru Konishi, Kosuke Sakitani, Kazuhiko Koike, Timothy Wang. Muscarinic acetylcholine receptor subtype 3 regulates gastric stem cell expansion and gastric cancer progression by controlling YAP activation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3339. doi:10.1158/1538-7445.AM2017-3339
Abstract 3350: Cell-free DNA fragmentation patterns analyzed in over 15000 cancer patients reveal changes associated with tumor somatic mutations and result in improved sensitivity and specificity of somatic variant detection
Authors: Abdueva Diana , Eltoukhy Helmy , Chudova Darya , Talasaz AmirAli
Published: July 2017

Background: Cell-free DNA (cfDNA) isolated from plasma consists of DNA fragments surviving clearance of dying cells and bloodstream trafficking. In cancer, these fragments carry the footprint of tumor somatic variation as well as its microenvironment. Since genomic distribution of cell free DNA fragments was shown to reflect nucleosomal occupancy in hematopoietic cells, we hypothesized that (a) heterogeneous patterns of cfDNA positioning would be associated with distinct mutations in patient tumors and (b) integration of fragmentation patterns into analysis would allow increased sensitivity and specificity of somatic mutation detection.

Methods: cfDNA fragment length and position distributions as well as associated somatic genomic profiles of over 15,000 patients with advanced-stage clinical cancer were determined by a highly accurate, deep-coverage (15,000x) ctDNA NGS test targeting 70 genes (Guardant360). An integrative data analysis of variant-free fragmentomics domain across different driver mutations was performed to identify patterns associated with detected somatic alterations.

Results: We discovered distinct classes of fragmentomics subtypes significantly enriched in samples with different genomic subtypes. An independent cohort of samples with known HER2 immunohistochemistry status was interrogated to confirm discovered association between fragmentation patterns and HER2 status. Integrating fragmentomics amplification signature with ERBB2 copy number analysis has resulted in 42% increase in the sensitivity and 7% increase in specificity of detection. Observed lung adenocarcinoma fragmentomics subtypes co-occured with mutually exclusive genomic alterations and previously described intrinsic molecular subtypes of lung cancer.

Conclusions: Fragmentomics classification of cancer cfDNA provides independent evidence for observed somatic variation and underlying tumor microenviroment, leading to higher sensitivity and accuracy of variant detection.

Citation Format: Diana Abdueva, Helmy Eltoukhy, Darya Chudova, AmirAli Talasaz. Cell-free DNA fragmentation patterns analyzed in over 15000 cancer patients reveal changes associated with tumor somatic mutations and result in improved sensitivity and specificity of somatic variant detection [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3350. doi:10.1158/1538-7445.AM2017-3350
Abstract 3386: Brazilian melanoma genome project: mutational landscape based on whole-genome sequencing
Authors: Vazquez Vinicius L. , Evangelista Adriane , Silveira Henrique C. , Crovador Camila , Carvalho Andre L. , Scapulatempo-Neto Cristovam , Lee Jin , Landman Gilles , Kitagima Joao P. , Reis Rui M.
Published: July 2017

Melanoma is the most aggressive form of skin cancer with increasing prevalence in Brazil. The comprehensive molecular profiling of these tumors and the recent genomic classification has improved the knowledge of melanoma biology and foster the identification of potential clinical biomarkers. The aim of this project, as part of the International Cancer Genome Consortium (ICGC) is to perform a genomic profile by whole genome sequencing (WGS) of Brazilian melanomas patients. We reported the WGS using Illumina paired-end sequencing strategy (>30 X-fold coverage) of 66 cases (45% primary and 55% metastatic) and respective matched normal. Overall we observed an average of 46K mutations/genome. The most frequent type of substitutions identified were: C>T (22.5%), T>C (6.4%), T>G (4.4%), T>A (2.9%), C>A (2.8%) and C>G (1.7%). A significative difference between frequency of C>T substitutions was observed among histological subtypes, i.e., acral lentiginous (11.3%), nodular (31.2%) and superficial spreading (25.9%). Among the classic genes involved in melanoma biology, we found that 31.8% of patients had BRAF mutations, being the V600E the most frequent (82.6%). A total of 6.1% of patients had NRAS, 3.0% showed KRAS, one case (1.5%) showed HRAS, and 12.1% showed NF1 mutations. TERT promoter mutation (C250T) was found in 28.8% of patients. TP53 mutations found 4.5% and RB1 in 1.5% of cases. Oncogenic KIT and PDGFRA mutations were observed in 6.1%and 1.5%, respectively. BRAF, NRAS, HRAS, KRAS and NF1 mutation were mutually exclusive. BRAF and TERT mutations were significantly more frequent in superficial than acral lentiginous melanomas. We showed that Brazilian melanoma patients exhibit a high number of single mutations and a similar mutation profile to the one reported in other populations. Acral/lentiginous melanoma subtype, which is a common subtype in our setting, showed a distinct mutation frequency. Further analysis will extend to other relevant genes and will also include the study of additional genomes to reach the goal of genomic landscape of 100 Brazilian melanomas cases.

Citation Format: Vinicius L. Vazquez, Adriane Evangelista, Henrique C. Silveira, Camila Crovador, Andre L. Carvalho, Cristovam Scapulatempo-Neto, Jin Lee, Gilles Landman, Joao P. Kitagima, Rui M. Reis. Brazilian melanoma genome project: mutational landscape based on whole-genome sequencing [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3386. doi:10.1158/1538-7445.AM2017-3386
Abstract 3397: Post-mortem examination of an aggressive case of medullary thyroid cancer characterised by catastrophic genomic abnormalities
Authors: O'Connor Darran P. , Das Sudipto , Kelly Deirdre , Moran Bruce , Han Kathleen , Mulligan Niall , Barrett Ciara , Buckley Patrick , McMahon Peter , McCaffrey J , Essen Henrik van , Connor Kate , Ylstra Bauke , Lambrechts Diether , Gallagher William M. , Kelly Catherine M.
Published: July 2017

Catastrophic genomic alterations may drive aggressive cancer phenotypes. We describe a diagnostically challenging and rapidly fatal case of medullary thyroid carcinoma (MTC) occurring in a young, morbidly obese man presenting with diffuse bone marrow involvement and disseminated intravascular coagulation. Whole-exome (WES) and shallow whole-genome sequencing (sWGS) were carried out for the primary tumour, adjacent normal, bone marrow tissues, multiple metastases and blood samples derived from the patient. We identified three germline single nucleotide polymorphisms (SNPs) within the RET proto-oncogene that remained undetected using routine hospital genetic testing procedures. One variant (L769L) has been previously reported to be associated with aggressive MTC presentation yet remains untested for in the routine diagnosis of MTC. Supported by findings from both WES and sWGS, we report for the first time in thyroid cancer on the occurrence of a “chromothripsis-like pattern”, which involved shattering of chromosome 4 leading to complete abrogation of normal chromosomal function, along with dramatic widespread copy number aberrations across both primary tumour and bone marrow samples. Based on the somatic variants identified, we describe the evolutionary pathway of the case, showing that bone marrow metastasis occurred separately to other metastatic sites. The presence of disease-associated SNPs within the RET proto-oncogene supports their inclusion as part of routine genetic testing for MTC cases. The copy number aberrations and chromothripsis-like pattern affected a much broader range of genes than single mutations and may have led to the widespread chromosomal instability evident, possibly contributing to the rapid fatal course of the case. These results provide a rationale for the application of comprehensive genomic analysis of cancers presenting with unusual and aggressive phenotypes to facilitate more appropriate therapeutic options and diagnoses.

Citation Format: Darran P. O'Connor, Sudipto Das, Deirdre Kelly, Bruce Moran, Kathleen Han, Niall Mulligan, Ciara Barrett, Patrick Buckley, Peter McMahon, J McCaffrey, Henrik van Essen, Kate Connor, Bauke Ylstra, Diether Lambrechts, William M. Gallagher, Catherine M. Kelly. Post-mortem examination of an aggressive case of medullary thyroid cancer characterised by catastrophic genomic abnormalities [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3397. doi:10.1158/1538-7445.AM2017-3397
Abstract 3434: Loss of regulation of β-Catenin expression by miR-214 and oncogenic role of miR-214 in metastatic melanoma cells
Authors: Prabhakar Kirthana , Jayanthy Ashika , Setaluri Vijay
Published: July 2017

Dysregulation of β-catenin facilitates melanoma metastasis by activating the transcription of oncogenes like Myc, CyclinD1 and TCF-1. It is important to understand the regulation of β-catenin to develop strategies to target Wnt signaling for melanoma treatment. MicroRNAs are small non-coding RNAs that are primarily involved in post-transcriptional regulation of target genes. Hence, miRNAs can be used to manipulate protein levels by causing instability of the transcript or by inhibiting translation. MiR-214 has been documented as regulator of β-catenin in cancer cells other than melanoma cells. The role of miR-214 in regulation of β-catenin has not been investigated. In this study, we investigated regulation of β-catenin by miR-214 in melanoma cell lines and studied the effect of overexpression miR-214 on the malignant phenotype of metastatic melanoma cells and their sensitivity to MAP kinase inhibitors. Our data show that β-catenin is upregulated in a panel of metastatic melanoma cells and the expression levels of β-catenin is negatively correlated with endogenous expression of miR-214. However, overexpression of miR-214 by transfection with a miRNA mimic did not show significant decrease in β-catenin mRNA or protein expression. RNA-Seq showed that multiple isoforms of β-catenin transcript are present in melanoma where the binding site is absent or mutated. The alternatively spliced transcript with no binding site for miR-214 is seen to be more abundant than the other isoforms. Paradoxically, overexpression of miR-214 increased survival, proliferation and migration of metastatic melanoma. In addition, miR-214 overexpression decreased their sensitivity to vemurafenib and selumetinib. In melanoma cells with acquired resistance (in vitro), miR-214 overexpression further enhanced their resistance to MEK inhibitor, but not vemurafenib. In light of the published studies that proposed that miR-214 acts as tumor suppressor by negatively regulating β-catenin, our study is interesting since we find that in metastatic melanoma miR-214 has oncogenic role. Ongoing studies are aimed at understanding the mechanisms by which miR-214 promotes malignant behavior of melanoma cells.

Citation Format: Kirthana Prabhakar, Ashika Jayanthy, Vijay Setaluri. Loss of regulation of β-Catenin expression by miR-214 and oncogenic role of miR-214 in metastatic melanoma cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3434. doi:10.1158/1538-7445.AM2017-3434
Abstract 3437: Targeting microRNA-143 in glioblastoma in vivo increases tumor growth
Authors: Lozada-Delgado Eunice , Valiyeva Fatma , Marcos Maria , Vivas Pablo
Published: July 2017

The purpose of this study is to assess the biological role of the microRNA-143 (miR-143) in Glioblastoma multiforme (GBM). GBM is the most common and lethal of all brain tumors. In the United States, the incidence of GBM is about 17% of all primary brain tumors and about 60-75% of all Astrocytomas (American Brain Tumor Association, 2014). The standard therapy is surgical tumor removal followed by chemotherapy and radiotherapy. However, many patients recur after treatment and the median survival rate for GBM has remained 15 months for the past 20 years. Thus, novel therapies for GBM treatment are urgently necessary. MicroRNAs (miRNAs) are a class of small non-coding RNAs (18-22 nucleotides in length) that regulate gene expression at the post-transcriptional level. MiRNAs bind to the 3’-untranslate region (UTR) of messenger RNAs (mRNAs) and regulate protein synthesis. Several deregulated miRNAs have been identified in all cancer types including GBM. In this study we aim to thoroughly uncover the role of miR-143 by using GBM cell lines, mouse models and patient samples. Total RNA was isolated from FFPE samples from brain tumor patients. TaqMan-based Real-time PCR showed that the relative expression of miR-143 was higher in GBM patients compared to control individuals, and with paired surrounding non-cancerous tissue. Furthermore, GBM cells transiently transfected with a miR-143 oligonucleotide inhibitor showed reduced cell proliferation (68.5%) (clonogenicity assay), increased apoptosis and cell cycle arrest of GBM cells in the S phase (Flow cytometry and Western blots). In vivo studies using primary GBM cells injected in the flank of nude mice showed that repeated doses of miR-143-inhibitor liposomal formulation increased the tumor size compared with control mice. These contradictory results could be due to effects of the microenvironment where the tumor is growing. Further studies will be made using intracranial injections in an orthotopic xenograft mouse model to confirm this hypothesis. Western blot analysis and luciferase reporter assays are also underway to identify novel miR-143 target genes in GBM cells. This research project is being supported by: PRCTRC: NCRR (U54 RR 026139-01A1), NIMHD (8U54 MD 007587-03), and RCMI: MBRS-RISE, NCRR (2G12-RR003051) and NIMHD (8G12-MD007600) from the NIH.

Citation Format: Eunice Lozada-Delgado, Fatma Valiyeva, Maria Marcos, Pablo Vivas. Targeting microRNA-143 in glioblastoma in vivo increases tumor growth [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3437. doi:10.1158/1538-7445.AM2017-3437
Abstract 3439: MicroRNA-34a epigenetically silences epithelial-mesenchymal transitions (EMT)-TFs in metastatic breast cancer cells
Authors: Imani Saber , Wei Chunli , Cheng Jingliang , Fu Junjiang
Published: July 2017

MicroRNA-34a (miR-34a) as key regulator of tumor suppression gene potentially plays a profound role in the tumorigenesis and progression of several cancers including breast cancer. In the present study, we invstigated the expression, targets and functional effects of miR-34a and its correlation with tumour characteristics and clinical outcome in the epithelial to mesenchymal transitions (EMT) stage of metastatic breast cancer (MBC). We focused to analyze EMT-inducing transcription factors (EMT-TFs)/miR-34a feedback loop, like TWIST1, SLUG, ZEB1/2, NOTCH1, on promotes MBC migration, invasion and metastasis. We found that miR-34a potently inhibits NOTCH1, ZEB1 protein expression activity in breast cancer cells. Importantly, the miR-34a do not have any inhibitory effects upon the SLUG and ZEB2. Using quantitative reverse transcription-PCR and western blot analysis, we showed that average pre-miR-34a expression is down regulated in human metastatic breast tissues as compared to normal human breast and negatively correlates with clinicopathological features of MBC patients. Ectopic expression of miR-34a in metastatic breast cancer cell-line BT-549 significantly inhibited cell proliferation, migrations, survival and cell invasion, but no clear effect on cancer cell growth. We provide indication that these regulatory pathways may stabilize epithelial and mesenchymal states, individually. We have consequently revealed that miR-34a is downregulated in MBC tissues and that it suppresses tumor growth by affecting several malignancy endpoints via downregulation of multiple EMT-TFs. Mutational or epigenetic inactivation of the EMT-TFs/miR-34a pathway probably alterations the equilibrium of these regulations toward EMT and the mesenchymal state and thereby contributes to metastasis. The results also suggest that miR-34a could serve as a potential therapeutic agent for MBC. Funding support: This work was supported by the NSFC (81172049, 81672887), Corresponding author email:fujunjiang@hotmail.com;fujunjiang@swmu.edu.cn

Citation Format: Saber Imani, Chunli Wei, Jingliang Cheng, Junjiang Fu. MicroRNA-34a epigenetically silences epithelial-mesenchymal transitions (EMT)-TFs in metastatic breast cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3439. doi:10.1158/1538-7445.AM2017-3439
Abstract 3441: A novel lncRNA-mediated YAP1 signaling pathway activation in epithelial ovarian cancer
Authors: Lin Xianzhi , Corona Rosario I. , Gayther Simon A. , Hazelett Dennis L. , Lawrenson Kate
Published: July 2017

Background: The long noncoding RNA (lncRNA) urothelial cancer associated 1 (UCA1) is upregulated in many cancer types, including ovarian cancer. In high grade serous ovarian cancers (HGSOCs), we had previously found that UCA1 expression negatively correlates with patients’ total survival and disease-free survival and that UCA1 knockout significantly impairs tumor growth in vivo. In addition, UCA1 promotes ovarian cancer cell growth via activating the YAP1 signaling pathway.

Purpose: Our present study aims to dissect the molecular mechanisms underlying YAP1 signaling activation by UCA1.

Methods: To characterize the UCA1 interactome, we utilized a modified in vivo RNA antisense purification method (UCA1-iRAP). This UCA1-iRAP method uses streptavidin-coupled magnetic beads to pull down biotin-labeled antisense probes which are complementary to the target RNA. In such a way, UCA1 RNA will be specifically enriched together with its in vivo-associated proteins, RNAs and/or genomic DNA, which can be profiled using mass spectrometry (MS) and next generation sequencing (NGS), respectively.

Results: Using UCA1-iRAP RNA-Seq and UCA1-iRAP protein MS, we comprehensively annotated the UCA1-interacting RNAs as well as UCA1-associated proteins. Interestingly many of the proteins identified were not known to have RNA-binding potential. Integration of NGS and MS data with multilayered functional and molecular characterization of UCA1 knockout, knockdown and overexpression models identified a YAP1 modulator involved in UCA1-mediated regulation of YAP1 signaling. Through repressing this YAP1 modulator via direct RNA-protein interaction, UCA1 promotes nuclear translocation of YAP1, resulting in activation of YAP1 target genes expression.

Conclusions: Our analyses indicate, for the first time, that UCA1 promotes ovarian cancer cell growth by activating YAP1 signaling via direct interaction with a YAP1 modulator. The interaction between UCA1 and this YAP1 modulator leads to activation of YAP1 signaling by nuclear translocation of YAP1. In summary, we characterize a novel UCA1-YAP1 signaling axis active in HGSOCs and also reveal that oncogenic lncRNA’s binding potential of cytoplasmic signaling proteins could be a common mechanism of post-translational control of signaling pathways deregulated in cancer.

Note: This abstract was not presented at the meeting.

Citation Format: Xianzhi Lin, Rosario I. Corona, Simon A. Gayther, Dennis L. Hazelett, Kate Lawrenson. A novel lncRNA-mediated YAP1 signaling pathway activation in epithelial ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3441. doi:10.1158/1538-7445.AM2017-3441
Abstract 3451: Depletion of Mastl kinase inhibits cancer growth by induction of mitotic catastrophe in breast cancer
Authors: Yina Yoon , Kim Jae Sung
Published: July 2017

Targeting mitotic kinase is an emerging anticancer strategy with promising pre-clinical results. Mastl is an essential mitotic kinase regulating mitotic progression by inactivation of a tumor suppressor protein phosphatase 2A (PP2A). However, it is unclear whether inhibition of Mastl kinase can kills tumor cells. Here, we show that Mastl inhibition selectively kills cancer cells by induction of mitotic catastrophe in breast cancer cells. We found that Mastl overexpression was tightly associated with tumor progression and poor prognosis in breast cancer. Mastl depletion with specific siRNAs induced cell death in Mastl high breast cancer cells, but not in Mastl low breast cancer cells. Interestingly, Mastl depletion did not affect the viability of normal cells. In addition, Mastl depletion reduced the oncogenic properties of breast cancer cells. Furthermore, we found that Mastl depletion caused mitotic catastrophe. Therefore, our data indicate that inhibition of Mastl can selectively kill breast cancer cells by induction of mitotic catastrophe, providing that targeting Mastl kinase is a very useful approach for breast cancer treatment.

Citation Format: Yoon Yina, Jae Sung Kim. Depletion of Mastl kinase inhibits cancer growth by induction of mitotic catastrophe in breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3451. doi:10.1158/1538-7445.AM2017-3451
Abstract 3470: The promyelocytic leukemia (PML) protein modulates rate and localization of homologous recombination in human cell lines with the alternative lengthening of telomeres (ALT) pathway
Authors: Barry Walter , Heaphy Christopher , Meeker Alan
Published: July 2017

The promyelocytic leukemia protein (PML) is tightly associated with a defining feature of the cancer specific, telomerase-independent telomere-maintenance pathway known as alternative lengthening of telomeres (ALT); therefore, PML is an important potential target for chemotherapeutics. Cells running ALT contain large accumulations of telomeric DNA and RNA in the nucleoplasm that are frequently coated in a polymer shell of PML protein. These structures are known as ALT-associated PML bodies (APBs). APBs have been postulated to be major sites of the telomeric sequence homologous recombination believed to be the basis of telomere maintenance in ALT. A CRISPR based strategy was used to knock out the PML gene in two widely used ALT positive osteosarcoma-derived human cell lines; U2OS, and SAOS2. Knockout clones from both cell lines continued to proliferate past passage 90 without crisis, and knockouts were validated at the genetic and protein levels by Sanger sequencing and western blotting, respectively. Chromosome orientation fluorescent in situ hybridization (COFISH) was carried out to determine the number of telomere sequence exchanges occurring in a single cell cycle in these knockouts. A twofold increase in telomere-specific exchanges, presumably telomere sister chromatid exchanges (T-SCE), over the parental line was detected. Intrachromosomal exchanges not specific to telomeres were also analyzed by the harlequin chromosome staining method. A significant increase in these types of exchanges was also observed. In conclusion, the presence of PML is not required for ALT to continue to function, and may be acting to modulate homologous recombination frequency and location in ALT positive cells.

Citation Format: Walter Barry, Christopher Heaphy, Alan Meeker. The promyelocytic leukemia (PML) protein modulates rate and localization of homologous recombination in human cell lines with the alternative lengthening of telomeres (ALT) pathway [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3470. doi:10.1158/1538-7445.AM2017-3470
Abstract 3471: Replication of megabase-size alpha-satellite DNA arrays in human centromere
Authors: Erliandri Indri
Published: July 2017

Alphoid centromere array is composed by 171 bp AT rich alphoid monomer that tandemly arranged into high order repeat unit up to 4 MB array. Hallmark of centromeric DNA is CENPB box, a 17 bp motif that is a binding site of CENPB. As the chromosomal site that regulates the precise and accurate chromosome segregation, understanding DNA replication in centromere will reveal insight into faithful chromosome segregation that potentially influence cellular behavior in normal and cancer cell. The present of active replicons within alphoid centromere array are detected by enrichment of nsDNA abundance. This finding is supported by evident of Orc2, Cdt1 and Treslin accumulation on the alphoid centromere array. Chromatin fiber analysis revealed that inter origin distance within alphoid centromere array is irregular. CENPB depletion that is affected centromeric chromatin structure enhances DNA replication activity within alphoid centromeric array, indicates centromeric chromatin landscape alteration affected centromeric DNA replication dynamic.

Citation Format: Indri Erliandri. Replication of megabase-size alpha-satellite DNA arrays in human centromere [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3471. doi:10.1158/1538-7445.AM2017-3471
Abstract 3473: Functional analysis of the KIF22 in human prostate cancer cells
Authors: Sakamaki Rina , Saito Shungo , Kitano Tatsuya , Nittami Tadashi , Seo Jieun , Takagi Akimitsu , Ishiguro Hitoshi , Uemura Hiroji , Watanabe Masatoshi
Published: July 2017

In the previous study, we identified genes that were differentially expressed between monolayers and spheroids in prostate cancer cell lines. The KIF22 gene was extracted as one of candidate genes related with spheroid formation and multicellular resistance. KIF22 is one of the kinesin superfamily proteins that are microtubule-dependent molecular motor proteins with DNA-binding capacity. KIF22 plays an important role in eukaryotic cell mitosis and macromolecule transportation. Alteration of KIF22 expression and function may lead to cancer development and progression. In this study, we explored the function of the KIF22 in prostate cancer cell lines, DU145 and LNCaP using the KIF22 siRNA. The KIF22 mRNA levels in LNCaP and DU-145 monolayers were detected, and time-dependently increased in both spheroids. The inhibition of KIF22 mRNA using siRNA affected cell proliferation in DU145 and LNCaP spheroids as well as monolayers. In addition, this suppression caused G2/M arrest and apoptosis in both cell lines. In clinical samples, its mRNA expression was significantly higher in tumor portions than in non-cancerous portions. However, the inhibition of KIF22 mRNA did not affect cell migration and invasion in DU145 and LNCaP cells. These findings suggest that KIF22 may play an important role in prostate cancer proliferation, especially spheroid formation, and have the potential to be targeted for prostate cancer treatments such as combination therapy with docetaxel.

Citation Format: Rina Sakamaki, Shungo Saito, Tatsuya Kitano, Tadashi Nittami, Jieun Seo, Akimitsu Takagi, Hitoshi Ishiguro, Hiroji Uemura, Masatoshi Watanabe. Functional analysis of the KIF22 in human prostate cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3473. doi:10.1158/1538-7445.AM2017-3473
Abstract 349: Eribulin induces cortical localization of E-cadherin, p120-catenin and beta-catenin in breast cancer cells
Authors: Clark Alison D. , Dybdal-Hargreaves Nicholas F. , Mooberry Susan L.
Published: July 2017

Eribulin provides a survival advantage for breast cancer patients, particularly those with triple-negative and HER2-negative subtypes.1,2 Mechanistically, eribulin was shown to reverse epithelial-to-mesenchymal transition (EMT) in breast cancer models.3 E-cadherin is a central regulator responsible for maintenance of the epithelial phenotype and loss of cortical E-cadherin and its downstream signaling is an early and significant event in EMT. Short term treatment of HCC1937 breast cancer cells with eribulin initiated cortical E-cadherin localization in manner different from other microtubule targeting agents. Cortical E-cadherin plays essential roles in cell adhesion and in regulating the Wnt pathway, whose aberrant activity is implicated in EMT. At the adherens junctions p120-catenin and β-catenin bind to E-cadherin, an interaction facilitated by microtubules. Cortical sequestration of p120- and β-catenin at the adherens junctions stabilizes these complexes and inhibits their ability to stimulate signaling pathways that contribute to EMT. We tested whether eribulin, which binds to the plus ends of microtubules, promotes EMT reversal by influencing p120- and β-catenin localization and activity. Our results show that eribulin induces cortical localization of p120-catenin and β-catenin with E-cadherin in HCC1937 cells, consistent with the formation of normal adherens junctions. Eribulin also stimulated the phosphorylation of Ser33/37 and Thr41 on β-catenin which promotes proteasomal degradation, which would be expected to further inhibit oncogenic signaling. Collectively, our results suggest that eribulin contributes to reversal of EMT through modulation of downstream signaling initiated by cortical localization of p120- and β-catenin at the adherens junctions. Funding for this work was provided by Eisai Inc.

1. Cortes J., et al., Lancet 377:914-923, 2011

2. Twelves, C., et al., Breast Cancer Res Treat 148:553-561, 2014

3. Yoshida, T., et al., Br J of Cancer 110: 1497-1505, 2014

Citation Format: Alison D. Clark, Nicholas F. Dybdal-Hargreaves, Susan L. Mooberry. Eribulin induces cortical localization of E-cadherin, p120-catenin and beta-catenin in breast cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 349. doi:10.1158/1538-7445.AM2017-349
Abstract 3497: Network-based systematic inference of lncRNA activity in breast cancer
Authors: Ung Matthew , Mattox Daniel , Wang George , Cheng Chao
Published: July 2017

The production of lncRNA is widespread across the human genome, yet the functions of lncRNAs and the phenotypes resulting from their activation have remained largely unstudied. In cancer, several lncRNAs have been found to associate with disease development and poor patient prognosis including HOTAIR and MALAT1. Recent studies have been performed using lncRNA microarrays or re-annotated probes from coding gene microarrays to systematically investigate their roles in cancer. However, datasets from lncRNA microarrays are typically not accompanied by comprehensive clinical information that can be used to draw associations between lncRNA expression and clinical phenotypes. Furthermore, lncRNA stability and cellular localization is highly variable suggesting that expression-based inference of lncRNA activity may not always be an accurate measure of its activity. To address this issue, we introduce an activity-based approach whereby we analyze the expression of lncRNA target genes to infer lncRNA activity. Specifically, we construct a breast cancer specific lncRNA-target gene network from RNA-seq data using a network reconstruction algorithm. We then apply this network to several breast cancer microarray datasets to systematically investigate lncRNA activity and their association with clinical phenotypes. Notably, we find that certain lncRNAs are active in specific molecular subtypes of breast cancer even though their expression levels do not reveal significant differences. Lastly, we performed survival analysis to systematically screen for novel lncRNAs whose activity associate with patient prognosis. Our results suggest that activity-based inference of lncRNAs can identify novel lncRNAs that may serve as therapeutic targets or biomarkers in breast cancer.

Citation Format: Matthew Ung, Daniel Mattox, George Wang, Chao Cheng. Network-based systematic inference of lncRNA activity in breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3497. doi:10.1158/1538-7445.AM2017-3497
Abstract 350: Pregnancy at early age is associated with a reduction of progesterone-responsive cells and epithelial Wnt signaling in human breast tissue
Authors: Mechera Robert , Münst Simone , Däster Silvio , Piscuoglio Salvatore , Ng Charlotte , Meier-Abt Fabienne , Weber Walter P. , Soysal Savas D.
Published: July 2017

Background: Pregnancy at early age is the most significant modifiable factor which consistently decreases lifetime breast cancer risk. However, the underlying mechanisms haven’t been conclusively identified. Studies in mice suggest a reduction in progesterone-receptor (PR) sensitive epithelial cells as well as a downregulation of the Wnt signaling pathway as being one of the main mechanisms for the protective effect of early pregnancy. The aim of our study was to validate these findings in humans.

Methods: We collected benign breast tissue of 123 women who had been stratified according to age at first pregnancy and the occurrence of subsequent breast cancer, and performed immunohistochemistry for PR, Wnt4 and the Wnt-target Versican.

Results: The number of PR positive epithelial cells was significantly lower in the group of women with early pregnancy and no subsequent breast cancer compared to the group of nulliparous women with subsequent invasive breast cancer (p=0.017). In women with early pregnancy, expression of Versican and Wnt4 was significantly lower compared to nulliparous women (p=0.0064 and p=0.0156 respectively), and Versican expression was also significant lower compared to women with late pregnancy (p<0.0001).

Discussion: Our results confirm prior observations in mice and suggest a role of downregulation of epithelial Wnt signaling in the protective effect of early pregnancy in humans. This results in a decreased proliferation of stem/progenitor cells; therefore, the Wnt signaling pathway may represent a potential target for breast cancer prevention in humans.

Citation Format: Robert Mechera, Simone Münst, Silvio Däster, Salvatore Piscuoglio, Charlotte Ng, Fabienne Meier-Abt, Walter P. Weber, Savas D. Soysal. Pregnancy at early age is associated with a reduction of progesterone-responsive cells and epithelial Wnt signaling in human breast tissue [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 350. doi:10.1158/1538-7445.AM2017-350
Abstract 3501: Prognostic value of a 4 lncRNA signature in patients with non-small cell lung cancer: a lncRNA expression analysis
Authors: Long XiaoRan , Wang HuiYun , Ge ChunLei , Song Xin , Zhang MeiYin , Zhen XiaoFeng
Published: July 2017

Recently, increasing evidence indicates that lncRNA plays an important role in the cancer development and progression. In this study, we are aimed to identify lncRNA signature for predicting survival of non-small cell lung cancer (NSCLC) patients. A total of 366 NSCLC cases, which were divided into a training and validation set, were detected by lncRNA microarray (containing 2412 lncRNA probes), and another 73 cases as an independent set were analyzed by qRT-PCR. LncRNA microarray analysis identified a total of 305 differentially expressed lncRNAs (138 up-regulated and 167 down-regulated) between NSCLC and normal tissues in the training set. We then validated the microarray data by qRT-PCR analysis of 5 lncRNAs (2 up-regulated and 3 down-regulated) in 30 paired NSCLC and adjacent lung tissues randomly selected from the training set. We then used a stepwise forward process to select the best combination of lncRNAs with the most significance in prediction of NSCLC prognosis and a 4-lncRNA prognostic signature was established, which was significantly associated with overall survival (hazard ratio [HR]: 3.563, 95% confidence interval [CI]: 2.118-5.995, P <0.001) and disease-free survival (HR: 3.236, 95% CI: 2.116-4.940, P <0.001) in the training set. This prognostic signature was validated in the validation (OS, HR: 3.186, 95% CI: 1.849-5.488, P <0.001; DFS, HR: 2.967, 95% CI: 1.949-4.517, P <0.001) and independent sets (OS, HR: 4.703, 95% CI: 1.923-11.50, P <0.001; DFS, HR: 4.549, 95% CI: 2.139-9.674, P <0.001). This 4-lncRNA signature has been demonstrated as an independent prognostic factor, which significantly improves the prognostic accuracy of TNM staging system in a combined model as assessed by ROC curve analyses in the three sets, respectively. Therefore, the 4-lncRNA signature has potential prognostic role in NSCLC patients.

Citation Format: XiaoRan Long, HuiYun Wang, ChunLei Ge, Xin Song, MeiYin Zhang, XiaoFeng Zhen. Prognostic value of a 4 lncRNA signature in patients with non-small cell lung cancer: a lncRNA expression analysis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3501. doi:10.1158/1538-7445.AM2017-3501
Abstract 3514: Assessing the role of CREB in fibrolamellar hepatocellular carcinoma with PRKACA-DNAJB1 fusion protein
Authors: Karki Anju , LaQuaglia Michael J. , Dios Amanda M. , Sadri-Vakili Ghazaleh , Vakili Khashayar
Published: July 2017

Introduction: Fibrolamellar Hepatocellular Carcinoma (FLHCC) is a rare form of liver cancer that occurs predominantly in adolescents or young adults. Curing or prolonging the lives of patients with FLHCC is extremely challenging. Despite aggressive surgical resections of primary tumors as well as metastatic lesions, outcomes tend to be poor due to lack of effective chemotherapy. Previous studies have demonstrated the presence of PRKACA-DNAJB1 fusion protein in the majority of FLHCC patients. DNAJB1 is a heat shock protein and PKA is a kinase that phosphorylates numerous substrates including cAMP-regulatory element-binding protein (CREB), a proto-oncogenic transcription factor. Therefore, we sought to examine whether the presence of the fusion protein in FLHCC is associated with increased CREB phosphorylation.

Methods: We analyzed tumor and matched non-neoplastic liver tissue from 7 pediatric patients with a pathologic diagnosis of FLHCC. We assessed the presence of the PRKACA-DNAJB1 fusion protein using PKA antibody, as well as total CREB and phosphorylated (p-CREB) using immunoblot analyses with specific antibodies.

Results: Immunoblot analysis using anti-PKA antibody confirmed the occurrence of PRKACA-DNAJB1 fusion transcripts in 6/7 (86%) FLHCC tumor samples. Immunoblot analyses of the 6 patients with the fusion protein revealed a trend towards a decrease in p-CREB expression in tumor samples compared to non-neoplastic tissue controls (p=0.15).

Conclusions: The pattern of p-CREB in FLHCC with PRKACA-DNAJB1 fusion protein suggests that the fusion protein does not exert its proto-oncogenic effects through the transcriptional activity of CREB.

Citation Format: Anju Karki, Michael J. LaQuaglia, Amanda M. Dios, Ghazaleh Sadri-Vakili, Khashayar Vakili. Assessing the role of CREB in fibrolamellar hepatocellular carcinoma with PRKACA-DNAJB1 fusion protein [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3514. doi:10.1158/1538-7445.AM2017-3514
Abstract 352: The role of p62 (SQTSM1) in the transforming growth factor β signaling pathway
Authors: Ng Evelyn , Gunaratne Adrian , Guglielmo John Di
Published: July 2017

Transforming growth factor beta (TGFβ) is a cytokine that regulates many cellular processes, including cellular adhesion, proliferation and apoptosis. Its canonical downstream effectors include Smad2/3 proteins, which are phosphorylated and then translocate to the nucleus to alter gene transcriptional programs and promote processes such as epithelial-to-mesenchymal transition (EMT). Previous studies in our lab have shown that atypical Protein Kinase C (aPKC) isoforms associate with TGFβ receptors and modulate receptor trafficking and signal transduction. An aPKC-associated protein, p62 (SQSTM1) has been implicated in TGFβ-dependent EMT, however the mechanisms remain unclear.Here, we investigate the localization of p62 and its potential roles in modulating TGFβ signaling via knockdown and overexpression studies. Using antibody feeding and immunofluorescence microscopy, we support previous findings showing that p62 localizes to late endosomes. In addition, using a co-immunoprecipitation approach, we observed that p62 may associate with TGFβ type II receptor. To explore the functional role of p62 in TGFβ signaling, we conducted protein silencing using siRNA. We observed TGFβ-independent decreases in E-cadherin expression. However these changes were independent of the phosphorylation status or nuclear translocation of Smad2.P62 also plays an important role in autophagy by targeting proteins for degradation. Recently, prolonged TGFβ stimulation has been shown to induce cellular autophagy. Therefore, it is in our interest to characterize the relationship between TGFβ-dependent EMT and autophagy, and to determine whether p62 has a regulatory role between these two processes.

Citation Format: Evelyn Ng, Adrian Gunaratne, John Di Guglielmo. The role of p62 (SQTSM1) in the transforming growth factor β signaling pathway [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 352. doi:10.1158/1538-7445.AM2017-352
Abstract 3536: The role of Nuclear factor I transcription factors in glioma
Authors: Chen Kok Siong , Richards Linda J. , Bunt Jens
Published: July 2017

Gliomas are the most common brain tumors in adults. Among the malignant gliomas, grade IV glioma, or glioblastoma (GBM), is the most aggressive form. Despite harsh treatments such as surgery, radiotherapy and chemotherapy, the median survival of GBM patients remains at just 12-15 months. GBM tumors could originate from cells of the glial lineage that have escaped the normal glial differentiation mechanisms. Therefore, by applying knowledge about normal glial development, we might understand how pathways that normally drive differentiation are affected in these tumors. If reactivating these pathways induces the differentiation of proliferative tumor cells, we could use this as a novel target for therapy.

Our research is focused on the Nuclear factor I (NFI) transcription factors, one of the key factors to induce glial differentiation during normal development and is implicated in glioma. In insertional mutagenesis glioma mouse models, Nfi genes are consistently disrupted, suggesting a role for NFI in glioma initiation. Furthermore, loss of NFIB is common in human astrocytoma, while loss of NFIA is associated with oligodendroglioma. To determine whether loss of Nfi alters glioma initiation, progression or tumor histology, we have crossed inducible glioma mice with conditional Nfi deletion mice to follow tumor initiation and progression using fluorescence imaging and immunohistochemistry.

Based on analyses of glioma mRNA expression data sets, NFIB expression correlates inversely with tumor grade and survival. Using immunofluorescence, we determined that both NFIA and NFIB are mainly expressed within the non-proliferative cells in GBM tissue, suggesting that NFI may play a direct role in tumor cell differentiation. To establish whether NFI expression can indeed inhibit tumor proliferation and induce differentiation, U251 GBM cells were transfected with NFI. While overexpression of NFIA and NFIB reduced cell proliferation, an NFI dominant-negative protein enhanced proliferation. We are currently validating these finding in vivo by electroporating NFI expression constructs into patient-derived GBM xenografts in mice. Our preliminary data show NFIB electroporated cells are indeed differentiated and non-proliferative.

Our data suggests that NFI-mediated glial differentiation can be induced in glioma to inhibit tumor growth and may prevent recurrence. Hence, activation of NFI could be a potential target for a glial differentiation-based approach to treat glioma.

Citation Format: Kok Siong Chen, Linda J. Richards, Jens Bunt. The role of Nuclear factor I transcription factors in glioma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3536. doi:10.1158/1538-7445.AM2017-3536