bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3395: Role of the activating protein 2 transcription factor in regulating cell invasion and migration in malignant glioma
Authors: Jain Saket , Poon Ho Yin , Godbout Roseline
Published: May 2015

Background: Malignant gliomas (MG) or high-grade astrocytomas are highly infiltrative brain tumors. Our previous work has shown that brain fatty acid binding protein (B-FABP) regulates the migratory properties of MG cells. B-FABP expression can be controlled by transcription factors such as NF1 and AP2. AP2 is a family of transcription factors involved in the regulation of genes responsible for cellular growth and differentiation during early development. Five AP2 genes have been identified in mice and humans: AP2α, AP2β, AP2γ, AP2δ and AP2ε. AP2 transcription factors are predominantly localized in the nucleus where they bind as homodimers or heterodimers to the consensus sequence GCCNNNGGC. Recent studies have shown that AP2α expression is inversely correlated with astrocytoma grade and that there are higher levels of AP2α in the cytoplasm of high-grade astrocytomas compared to low-grade astrocytomas.

Objective: To study the regulation of B-FABP expression in response to AP2 in MG.

Methodology: A panel of 10 MG cell lines was used to examine AP2α protein levels. Five of these cell lines were B-FABP +ve and five were B-FABP-ve. The effect of AP2 expression on endogenous B-FABP was examined by transfecting AP2 expression constructs into the B-FABP +ve U251 MG cell line. B-FABP levels were examined by western blot analysis and the subcellular localization of AP2 was studied by immunofluorescence analysis.

Results: Western blot analysis indicates that AP2α is expressed in four out of five B-FABP-ve MG cell lines but in only one of the five B-FABP +ve MG cell lines tested. AP2 overexpression studies indicate that B-FABP levels are significantly reduced upon overexpression of AP2α, AP2β or AP2ε. Immunofluorescence data suggest preferential localization of AP2β in the cytoplasm of B-FABP +ve U251 MG cells. Our preliminary result shows difference in affinity towards heterodimerization between AP2 family members.

Conclusion and future directions: AP2 is a negative regulator of B-FABP expression. Exclusion of AP2 from the nucleus allows transcription of B-FABP in MG cells, which in turn leads to increased cell migration. Experiments will be carried out to see the effect of AP2 heterodimerization on regulation of B-FABP transcription. Phosphorylation studies will be carried out to see whether AP2 phosphorylation affects its subcellular localization. This study may lead to new approaches to reduce B-FABP expression in MG, thereby reducing or preventing cell migration and tumor infiltration.

Citation Format: Saket Jain, Ho Yin Poon, Roseline Godbout. Role of the activating protein 2 transcription factor in regulating cell invasion and migration in malignant glioma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3395. doi:10.1158/1538-7445.AM2014-3395
Abstract 3397: Rational targeting of protein translation for cancer treatments
Authors: Chang Elizabeth , Nguyen Duc , Yang Qingyuan , Carrier France
Published: May 2015

Deregulation of protein translation is associated with a growing number of human diseases including tumorigenesis. New anti cancer therapies targeting the protein translation regulator mammalian Target Of Rapamycin (mTOR) highlight the importance of this signaling pathway. However, feedback mechanisms that can compensate for this general protein translation pathway limit the clinical efficiency of this approach. New drugs that could rationally target protein translation in cancer cells are therefore needed. We have identified the heterogenous ribonucleoprotein A18 (hnRNP A18) as a new regulator of protein translation in cancer cells. hnRNP A18 is up regulated in several human tumors including prostate, breast, colon and as shown here in melanoma tumors. hnRNP A18 regulates protein translation of its targeted transcripts by binding to a specific RNA signature motif in their 3’UTRs. Our data indicate that the hypoxia mimetic agent CoCl2 increases hnRNP A18 protein levels in melanoma cells but not in normal melanocytes. In addition, down regulation of hnRNP A18 sensitizes melanoma cells to CoCl2 and Temozolimide. The functional significance of this effect was evaluated in a mouse xenograft model where hnRNP A18 down regulation reduced melanoma tumor growth by more than 70%. RNA immunoprecipitation assay indicate that hnRNP A18 binds to a number of transcripts associated with tumor growth including the Hypoxia Inducible Factor 1α (HIF-1α). Levels of hnRNP A18 also directly affect HIF-1α protein levels in response to CoCl2. Taken together these data indicate that hnRNP A18 is a new regulator of protein translation that could be targeted to develop new mechanism based therapy for the rational inhibition of protein translation in cancer cells.

Citation Format: Elizabeth Chang, Duc Nguyen, Qingyuan Yang, France Carrier. Rational targeting of protein translation for cancer treatments. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3397. doi:10.1158/1538-7445.AM2014-3397
Abstract 3398: Reciprocal regulation of MYB and CXCR4 in pancreatic cancer cells: Potential pathobiological implications
Authors: Srivastava Sanjeev K. , Bhardwaj Arun , Arora Sumit , Singh Seema , Grizzle William E. , Singh Ajay P.
Published: May 2015

Pancreatic cancer remains one of the most lethal malignancies with a median post-diagnosis survival of 3-8 months, and a 5 year survival of ∼6%. The poor outcome from pancreatic cancer is largely due to the presence of locally advanced or metastatic disease at the time of diagnosis. Therefore, it is imperative to identify novel molecular targets driving aggressive and metastatic progression of pancreatic cancer, and build up a better understanding of the underlying molecular mechanisms. In this regard, we have identified that MYB, encoding for an oncogenic transcription factor (a cellular progenitor of v-Myb oncogenes), is overexpressed in majority of pancreatic cancer cases and cell lines. Furthermore, our data from gain and loss of function studies (in BxPC3 and MiaPaCa cells, respectively) show that MYB overexpression is associated with pancreatic tumor growth and metastasis. From the mechanistic standpoint, we observe that MYB regulates the expression of CXCR4 through direct binding to its promoter. MYB-overexpressing BxPC3 cells exhibit significantly enhanced migration and invasion against a CXCL12 (sole ligand for CXCR4) gradient, whereas it is suppressed in MYB-silenced MiaPaCa cells as compared to their respective control cells. Furthermore, MYB-overexpressing cells exhibit greater resistance to serum-deprivation in CXCL12-supplemeted media, an effect that is abrogated upon CXCR4 silencing. Interestingly, in parallel studies, we have observed that treatment with CXCL12 also leads to MYB upregulation in pancreatic cancer cells in a CXCR4 dependent manner. Together, these data are suggestive of a novel positive feedback regulatory loop between CXCR4 and MYB and hold important significance for pancreatic cancer pathobiology.

Citation Format: Sanjeev K. Srivastava, Arun Bhardwaj, Sumit Arora, Seema Singh, William E. Grizzle, Ajay P. Singh. Reciprocal regulation of MYB and CXCR4 in pancreatic cancer cells: Potential pathobiological implications. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3398. doi:10.1158/1538-7445.AM2014-3398
Abstract 3399: Detection and validation of novel RNA cancer biomarkers by single molecule RNA fluorescence in situ hybridization (smRNA FISH)
Authors: Johansson Hans E. , Orjalo Arturo V. , Coassin Sally R. , Yin Raymund
Published: May 2015

RNA biomarkers are discovered and validated through advanced transcriptome analysis at a fast pace. These markers that allow the interrogation of the active genes hold promise for more precise staging, prognosis, and treatment of cancer. Long RNA markers comprise pre-mRNAs found in the cell nucleus, long non-coding RNAs (lncRNAs) that may be found in both the nucleus and the cytoplasm, as well as mature and mostly cytoplasmic mRNAs. Importantly, comparative studies in mouse have demonstrated that transcript levels are more strongly correlated with clinical traits than the corresponding protein levels. Because lncRNA and mRNA biomarkers have the potential to contribute to the development of targeted cancer therapies, there is an obvious need for a robust and dependable methodology to reliably detect and validate these RNA targets. RNA fluorescence in situ hybridization (RNA FISH) provides a powerful means to detect specific RNAs in single cells, while still maintaining tissue morphology. Significant advances in RNA FISH technology, such as the hybridization of multiple, fluorescently labeled 20-mer oligonucleotides to the RNA target, allow for the detection of single RNAs and yields information on the RNA's distinct spatial distribution within tissues and even within cells. In this study, we investigate emerging RNA biomarkers in prostate cancer cell lines and tissue utilizing RNA FISH. We also examine established breast cancer biomarkers such as human epidermal growth factor receptor 2 (HER2 / ERBB2), estrogen receptor α (ER α / ESR1), and progesterone receptor (PR / PGR) in breast cancer tumors and cell lines. The results and tools presented here will contribute to advancing the current capabilities of the detection and treatment of specific cancers, as well as in the continued discovery and development of cancer drug candidates.

Citation Format: Hans E. Johansson, Arturo V. Orjalo, Sally R. Coassin, Raymund Yin. Detection and validation of novel RNA cancer biomarkers by single molecule RNA fluorescence in situ hybridization (smRNA FISH). [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3399. doi:10.1158/1538-7445.AM2014-3399
Abstract 3404: Identifying genes involved in retinoic-acid-mediated breast tumor progression by total-genome-knockdown screen
Authors: Coyle Krysta M. , Dean Cheryl , Pan Lu-Zhe , Ahn Dae-Gyun , Sultan Mohammad , Salsman Jayme , Dellaire Graham , Giacomantonio Carman , Lee Patrick W. , Marcato Paola
Published: May 2015

Despite numerous advances in cancer care, breast cancer remains one of the leading causes of cancer-related death among Canadian women. Previous work by our lab has identified aldehyde dehydrogenase 1A3 (ALDH1A3) as a breast cancer stem cell marker, and patients with high levels of ALDH1A3 in their tumors have worse outcomes. Furthermore, we found that via its retinoic acid (RA) production function, ALDH1A3 also contributes to breast tumor growth and metastasis in a murine xenograft model. RA induces transcription of many genes (directly and indirectly), initiates cell signaling and regulates many cellular processes involved in cell growth and differentiation. To identify the key RA-inducible genes which mediate breast cancer progression we performed a genome-wide lentiviral-based shRNA screen using MDA-MB-231 breast cancer cells xenotransplanted into NOD/SCID mice with or without RA treatment. From the in vivo screen, 50 genes were identified as being putatively involved in RA-mediated breast tumor growth. 14 genes have been confirmed in vitro as differentially expressed in the presence of RA. We have generated individual shRNA knockdown clones of the genes and are validating their involvement in RA-mediated breast cancer progression in vivo. Analysis of the genes identified by this screen revealed potential interactions between RA and Wnt signaling, implicating extensive cross-talk between these two pathways. Completion of this study will define the mechanism of RA-mediated breast cancer progression and potentially identify new therapeutic targets for breast cancer treatment.

Citation Format: Krysta M. Coyle, Cheryl Dean, Lu-Zhe Pan, Dae-Gyun Ahn, Mohammad Sultan, Jayme Salsman, Graham Dellaire, Carman Giacomantonio, Patrick W. Lee, Paola Marcato. Identifying genes involved in retinoic-acid-mediated breast tumor progression by total-genome-knockdown screen. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3404. doi:10.1158/1538-7445.AM2014-3404
Abstract 3407: Regulation of c-Myc and p53 by splicing factor SRp20
Authors: He Xiaolong , Yang Jilai , Wang Shijie
Published: May 2015

SRp20 is a member of the highly conserved SR protein family of splicing regulators. Our previous study found that SRp20 was overexpressed in ovarian and breast tumors compared to corresponding normal tissues and knockdown of SRp20 expression inhibited tumor cell growth. Suppression of SRp20 by 90% triggered apoptosis of tumor cells through intrinsic pathway, which was accompanied by a downregulation of anti-apoptotic protein, Bcl-2. However, it is unclear what mechanism mediates the induction of apoptosis and downregulation of Bcl-2 by SRp20 knockdown. To address this question, we conducted microarray analysis to identify the genes whose expression or splicing are altered in SRp20-knockdown A2780 cells. Among hundreds of identified genes, the expression of c-Myc and p53 was downregulated. The downregulation was further validated by quantitative PCR. Because of the fundamental importance of p53 and c-Myc in cancer biology and apoptosis, we further examined the expression of these two genes at protein levels. Western blotting showed that p53 is upregulated at protein level in SRp20 knockdown cells, contradictory to its expression at the mRNA level, while the expression of c-Myc is consistent at the protein level and at the mRNA level. These results suggest that SRp20 knockdown-induced apoptosis could be mediated by upregulated p53 protein or reduced c-Myc or both, given already established connection between these two proteins and apoptosis. How SRp20 is involved in the regulation of p53 and c-Myc remains to be determined and is currently under investigation.

Citation Format: Xiaolong He, Jilai Yang, Shijie Wang. Regulation of c-Myc and p53 by splicing factor SRp20. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3407. doi:10.1158/1538-7445.AM2014-3407
Abstract 341: Imatinib induces expression of Bim and apoptosis in chronic myelogenous leukemia cells via p38/H2AX pathway
Authors: Xiong Min , Niu Tianhui , Dong Yaqiong , Lu Chengrong
Published: May 2015

H2AX is a novel human tumor suppressor protein and plays an important role in apoptosis of cancer cells. Increasing published data indicate that H2AX phosphorylation (Ser139) contributes to its regulation of cancer cell apoptosis. Thus, charactering the mechanisms and signaling pathways involved in H2AX phosphorylation (Ser139) will certainly provide better understanding on H2AX function in cancer cells. Our previous report showed that caspase-3/Mst1 pathway regulates phosphorylation of H2AX at Ser139 in chronic myelogenous leukemia (CML) cells. Recently we found another pathway which regulates H2AX phosphorylation (Ser139) in CML cells. When the CML cells were subjected to imatinib treatment, MAPK family member ERK1/2 activity was inhibited, but p38 activity was increased, another member JNK1/2 had no response. Then we used p38 inhibitor SB202190 and siRNA against p38 to block p38 activation and observed that H2AX phosphorylation and apoptosis of CML cells was concurrently inhibited. Our results also showed that inhibition of MSK1/MSK2 did not affect H2AX phosphorylation (Ser139), although the p38 and ERK1/2 downstream MSK1/MSK2 could be activated by imatinib in CML cells. Overall, these data confirmed that p38 signaling pathway is also required for H2AX phosphorylation (Ser139) in CML cells. Our further evidences indicated that inhibition of H2AX phosphorylation (Ser139) regulated by p38 during apoptotic induction with imatinib suppressed the expression of apoptosis-related gene Bim. Taken together, these data demonstrated that p38/H2AX pathway regulates Bim expression and subsequent apoptosis in CML cells.

Citation Format: Min Xiong, Tianhui Niu, Yaqiong Dong, Chengrong Lu. Imatinib induces expression of Bim and apoptosis in chronic myelogenous leukemia cells via p38/H2AX pathway. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 341. doi:10.1158/1538-7445.AM2014-341
Abstract 3410: A novel regulatory region of the MYC oncogene decreases Myc transcriptional activity
Authors: Kalkat Manpreet , Wasylishen Amanda , Chan Pak-Kei Michael , Pandyra Aleksandra , Kim Sam Sulgi , Bros Christina , Raught Brian , Penn Linda Z.
Published: May 2015

MYC is a transcription factor that contributes to over 50% of cancers. While deregulated protein expression of MYC is an important factor driving tumorigenesis, remarkably little is known about the post-translational modifications (PTMs) that are necessary for MYC to function as an active oncogene. Given the important role of PTMs in the regulation of protein activity and stability, our overall objective is to further characterize novel PTMs of MYC, and the functional contribution of these modifications to the ability of the MYC to transform cells. Using a structure-function approach, we have identified six C-terminal lysines that can modulate the ability of MYC to transform cells. By utilizing lysine to arginine mutations (6KR) to abrogate signaling through these residues, we show that MYC 6KR increases anchorage-independent colony growth compared with MYC-WT. Moreover, this mutant was also more potent than WT in promoting xenograft tumour growth. Through target gene expression analysis and luciferase reporter assays, we show that 6KR has enhanced transcriptional activity compared with MYC-WT. To characterize the modifications that can occur at these lysine residues, we have performed mass spectrometry analysis and have identified a number of PTMs that can occur in this region. Furthermore, we have conducted BioID to identify MYC interactors that change in association with 6KR MYC compared to MYC-WT. We theorize that identifying the signaling pathways leading to post-translational modification of this region will be an important first step in the development of inhibitors targeting MYC induced transformation.

Citation Format: Manpreet Kalkat, Amanda Wasylishen, Pak-Kei Michael Chan, Aleksandra Pandyra, Sam Sulgi Kim, Christina Bros, Brian Raught, Linda Z. Penn. A novel regulatory region of the MYC oncogene decreases Myc transcriptional activity. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3410. doi:10.1158/1538-7445.AM2014-3410
Abstract 3413: A genetic analysis of splicing in neuroblastoma identifies opposing functions for FUBP1 splice variants in MYC regulation
Authors: Chen Justin , Hackett Christopher S. , Zhang Shile , Song Young K. , Molinaro Annette , Quigley David A. , Balmain Allan , Gustafson William C. , Dyke Terry A. Van , Kwok Pui-Yan , Khan Javed , Weiss William A.
Published: May 2015

Neuroblastoma, a tumor of the peripheral neural crest, is the most common cancer of infancy and the most common extracranial solid tumor of childhood. Large scale sequencing efforts have identified recurrent mutations in only a handful of genes in a minority of patients, but these efforts have so far focused on the exome, limiting identification of mutations leading to changes in alternative splicing.

Regulation of alternative splicing is a critical and tightly regulated cellular function involving a complex interplay between cis-acting sequences on the pre-mRNA transcript and accessory trans-acting splicing factors. To elucidate genetic control of splicing, we used a splicing quantitative trait loci (sQTL) analysis in a backcrossed mouse model of neuroblastoma. We utilized Affymetrix Exon Arrays to profile mRNA expression at the exon level for superior cervical ganglia and cerebella (n=102) which we coupled with genotypes at 350 SNP and microsattlelite locations. With whole-genome sequencing data available for the inbred parental strains, our approach provided us with localized sequences for each individual mouse.

In addition to identifying novel candidate trans-acting splicing factors, these sQTL identify novel candidate intronic regulatory sequences critical for splicing and reveal strain-specific splicing events leading to functional consequences in neuroblastoma. Among these, we show that a subtle splicing event within FUBP1 modulates levels of the MYC oncoprotein in human neuroblastoma-derived cell lines and correlates with outcome in children with neuroblastoma.

Citation Format: Justin Chen, Christopher S. Hackett, Shile Zhang, Young K. Song, Annette Molinaro, David A. Quigley, Allan Balmain, William C. Gustafson, Terry A. Van Dyke, Pui-Yan Kwok, Javed Khan, William A. Weiss. A genetic analysis of splicing in neuroblastoma identifies opposing functions for FUBP1 splice variants in MYC regulation. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3413. doi:10.1158/1538-7445.AM2014-3413
Abstract 3414: Genomic signatures of melanoma maintenance
Authors: Sherbaf Faranak Ghazi , Inaki Koichiro , Bertrand Denis , Woo Xing Y. , Hoon Dave , Hillmer Axel , Liu Edison
Published: May 2015

Melanoma is the malignant tumor of melanocytes and considered as one of the most aggressive human cancers. While the worldwide incidence rate of melanoma has increased during the last decade, current therapies for melanoma do not provide long-term effect. The highly heterogeneous expression signature of melanoma underlies the inefficient therapy and accentuates the inevitability of profound molecular understanding of melanoma, its maintenance and progression. In this study, we aimed to reveal the complex patterns of distinct molecular mechanisms underlying the progression and maintenance of melanoma using high-throughput sequencing technologies. Series of melanoma tumor tissues representing discrete stages of the malignant progression were deeply sequenced. Integrative analysis revealed hard-wired genomic aberrations and associated transcriptomic consequences. These genomic aberrations included clusters of oncogenes brought together by structural variations, which cooperated in their amplifications and subsequent up-regulated expressions. We believe that these hard-wired genetic changes are biologically significant; they sustain the survival of the tumor and offer the advantage of the distant metastasis. We reconstituted the architecture of melanoma progression and maintenance and we identified gene cassettes with a putative role in sustaining tumor growth. The correlation of these gene cassettes with the cancer phenotype and the feasibility of these putative oncogenes as candidates for targeted therapy were addressed using cell-based assays on a high content screening platform. Analysis of the matched cell lines derived from the same patient offered a valuable resource for in vitro manipulation, testing of drugs and functional validation. A detailed molecular characterization of the functionally validated genes and their contribution in the cancer signaling pathways will provide a better understanding of the molecular basis of tumor evolution.

Citation Format: Faranak Ghazi Sherbaf, Koichiro Inaki, Denis Bertrand, Xing Yi Woo, Dave Hoon, Axel Hillmer, Edison Liu. Genomic signatures of melanoma maintenance. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3414. doi:10.1158/1538-7445.AM2014-3414
Abstract 3416: Transcriptome analysis of papillary thyroid cancer using next-generation sequencing technologies in Korean patients
Authors: Lee Seungbok , Jee Hyeon-Gun , Lee Kyu Eun , Seo Jeong-Sun
Published: May 2015

Thyroid cancer is one of the most prevalent cancers worldwide, and papillary thyroid cancer (PTC) accounts for >80% of all thyroid cancers. Here we conducted RNA sequencing for 70 PTC and 42 paired-normal tissues in Korean patients, and their transcriptomes were analyzed in various aspects. First, somatic mutations were screened transcriptome-wide throughout samples; BRAF mutation (V600E), which is the most well-established mutation in thyroid cancer, was detected in 47 samples out of 70 (67.14%). HRAS (Q61R, Q61K) and NRAS (Q61R) mutations were shown in two samples, respectively. In gene fusion analysis, we discovered four fusion mutations using our stringent criteria and these include CCDC6-RET and PAX8-PPARG fusions, which have been previously reported for thyroid cancer. Other EZR-ERBB4 and PAX8-GLIS3 fusions are novel fusion mutations found in PTC, and the sample having EZR-ERBB4 mutation was diagnosed as the follicular variant subtype. This is an in-frame mutation and previous reports have shown that ERBB4 is a driver gene for carcinogenesis in multiple types of cancers. In addition, we also found ETV6-NTRK3 fusion in less conservative criteria, which was a well-known fusion in other cancer types. Each fusion mutation above was shown in only one sample, respectively. Through gene expression analysis, we clustered PTC cancer samples into four groups; three of them are shown to be enriched with BRAF V600E mutation, and the other includes samples with NRAS, HRAS mutation. In this study, we thoroughly explored the transcriptome of PTC samples using the next-generation sequencing method and proved well-correlation between mutation and expression profiles. These results not only replicated several findings of previous studies and suggested some novel genes or pathways involved in the PTC development in Koreans.

Citation Format: Seungbok Lee, Hyeon-Gun Jee, Kyu Eun Lee, Jeong-Sun Seo. Transcriptome analysis of papillary thyroid cancer using next-generation sequencing technologies in Korean patients. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3416. doi:10.1158/1538-7445.AM2014-3416
Abstract 3421: Serial whole exome sequencing showed the genetic aggravation of refractory osteosarcoma
Authors: Heo Su Jin , Kim Ji Woong , Kwon Woo Sun , Kim Hyo K. , Hong Min Hee , Yang Woo Ick , Kim Se-Kyu , Kim Hyo Song , Chung Hyun Cheol , Hwang Tae Hyun , Rha Sun Young
Published: May 2015

Osteosarcoma is the most common malignant bone tumor in children and adolescents with frequent lung metastasis. As the survival has not been improved with chemotherapy for the last two decades, novel therapeutic approaches are required to efficiently treat osteosarcoma. Genomic analyses have revealed biologically useful information and genetically altered therapeutic targets. The subject of this study was a 25 year old patient with conventional high-grade osteosarcoma with multiple metastasis of lung, lymph nodes and chest wall, who had failed from several palliative systemic chemotherapies and radiotherapy. We performed whole exome sequencing of the gDNA from peripheral blood mononucleated cells (PBMC) and formalin fixed paraffin embedded (FFPE) tissues of primary tumor, secondary (recurrence 1) and tertiary (recurrence 2) metastatic tumors. Samples were sequenced using one lane of paired-end, 100 bp reads on Illumina Hiseq for each sample. Here, we present an analysis result using VarScan and custom-made programs for the detection of somatic mutations and loss of heterozygosity (LOHs) in exome data from changes of genetic variations throughout the disease progression compared to blood germ-line sequencing data. As a result, we observed the 36 common somatic mutated genes, including p73 and TSHR, in three tumor types. And we indentified the somatic mutations affecting the functions of known cancer genes, RBM15, SYNE1, GNAQ and XPA, which were only present in the secondary and/or tertiary metastatic tumors but not in the primary tumor. Especially, GNAQ mutations which is found in the metastatic tumors, is known to be mutated in 50% of melanoma and driven constitutive activity of the MAPK pathway. Therefore, GNAQ mutation in recurrent osteosarcoma patient might be a potential marker for new therapeutic strategies of inhibiting MAPK pathway. In conclusion, the ability to track clonal evolution in cancers may provide new strategies and opportunities for drug development, especially in refractory and rare diseases.

Citation Format: Su Jin Heo, Ji Woong Kim, Woo Sun Kwon, Hyo Ki Kim, Min Hee Hong, Woo Ick Yang, Se-Kyu Kim, Hyo Song Kim, Hyun Cheol Chung, Tae Hyun Hwang, Sun Young Rha. Serial whole exome sequencing showed the genetic aggravation of refractory osteosarcoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3421. doi:10.1158/1538-7445.AM2014-3421
Abstract 3422: RNAi screening identifies FGFR4 as a modulator of growth and survival in Ewing sarcoma
Authors: Azorsa David O. , Gonzales Irma M. , Arora Shilpi , Hagelstrom R. Tanner , Little Tanya H. , Arceci Robert J. , Mousses Spyro
Published: May 2015

Ewing sarcoma is the second most common cancer of bone and soft tissue arising in children and young adults. Standard treatment of Ewing sarcoma includes surgery, radiation, and chemotherapy largely consisting of combinations of non-targeted cytotoxic agents. Although the survival rate has improved for patients treated for localized disease, the survival rate of patients with metastatic tumor remains lower than 30%. In order to improve therapeutic options for Ewing Sarcoma, we employed a functional genomics approach based on RNA interference (RNAi) screening to identify genes whose silencing affected the proliferation and survival of Ewing sarcoma cells. Four Ewing sarcoma cell lines, TC-32, TC-71, SK-ES-1 and RD-ES, were transfected with a library of siRNA targeting 287 cancer-associated genes. The siRNA screening data were normalized and statistical cut-offs were determined for each cell line. From the list of siRNA ‘hits,’ we identified siRNAs targeting Fibroblast growth factor 4, FGFR4, to be among the most effective in inducing cytotoxicity in Ewing sarcoma cells. Further validation was conducted showing that siRNAs to FGFR4, but not FGFR1, FGFR2, or FGFR3, were able to significantly reduce Ewing Sarcoma cell viability. Furthermore, siRNA targeting FGFR4 were able to induce Caspase 3 activity. FGFR4 protein was shown to be expressed in Ewing sarcoma by western blot analysis, with expression levels lower than FGFR4 expression on rhabdomyosarcoma cells. Targeting FGFR4 activity in Ewing sarcoma cells using a pan-FGFR inhibitor, PD-173074, demonstrated that Ewing sarcoma cells were sensitive to FGFR inhibition. Since FGFR4 in a potential therapeutic target in rhadomyosarcoma, we compared the response of the Ewing sarcoma cells to PD-173074 to that of the rhabdomyosarcoma cell lines, RH-30 and RD. The IC50s of the four Ewing sarcoma cells ranged from 3.8-4.7 µM and were slightly lower than the IC50 for the rhabdomyosarcoma cell lines RH-30 (5.6 µM) and RD (7.9 µM). These results indicate that FGFR4 is a potential therapeutic target in Ewing sarcoma and that FGFR4 targeted therapy should be tested in preclinical xenograft models of Ewing sarcoma.

Citation Format: David O. Azorsa, Irma M. Gonzales, Shilpi Arora, R. Tanner Hagelstrom, Tanya H. Little, Robert J. Arceci, Spyro Mousses. RNAi screening identifies FGFR4 as a modulator of growth and survival in Ewing sarcoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3422. doi:10.1158/1538-7445.AM2014-3422
Abstract 3429: Synthetic lethal RNAi screens to nominate potential therapeutic combinations for uveal melanoma
Authors: Place Chelsea S. , Cowley Glenn S. , Root David E. , Garraway Levi A.
Published: May 2015

Uveal melanoma (UM) is the most common form of ocular cancer in adults and metastatic disease currently has no effective therapy. Over 80% of UM tumors harbor activating mutations in the heterotrimeric G protein α subunits GNAQ or GNA11. These mutations induce protein kinase C (PKC) and the MAP kinase pathway, which signals through RAF, MEK, and ERK. Clinical trials of MEK and PKC inhibitors are underway to treat UM, however, novel therapeutic strategies may be required for effective treatment. To identify rational combination therapies in the context of MEK or PKC inhibition, we performed a pooled, genome-wide synthetic lethal RNA interference screen. Candidate genes whose silencing sensitizes GNAQ/11-mutant UM cells to MEK or PKC inhibition were nominated and further validated across a panel of cell lines using a custom pool of shRNAs. Several genes scored well across all cell lines. The top MEK inhibitor sensitizer was BRAF, thereby highlighting the importance of MAPK signaling in UM. Several non-MAPK pathway genes were also nominated and may represent novel pathways for the development of combination therapies.

Citation Format: Chelsea S. Place, Glenn S. Cowley, David E. Root, Levi A. Garraway. Synthetic lethal RNAi screens to nominate potential therapeutic combinations for uveal melanoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3429. doi:10.1158/1538-7445.AM2014-3429
Abstract 3433: ARHGAP30 and DEDD as potential genomic drivers of invasive urothelial carcinomas
Authors: Hong Andrew L. , Riester Markus , Schinzel Anna C. , Schlauch Amy , Adam Rosalyn M. , Kwiatkowski David J. , Rosenberg Jonathan E. , Hahn William C.
Published: May 2015

Background: Urothelial carcinoma (UC) is diagnosed in 72,570 patients annually. 20-30% of these patients have high grade advanced disease. These patients have had historically poor prognosis with 5-year survival rates of 33% and 6% in patients with regional and distant staged disease, respectively. Recent genomic analysis suggests that amplification of 1q23.3 may be associated with poor survival in patients with advanced UC. Using functional genomic screens, we aim to identify the genomic driver(s) of this association.

Methods: First, we created an arrayed shRNA screen with 79 shRNAs targeting 15 genes within this amplification. We studied these effects across 4 and 6 bladder cell lines with and without amplification of 1q23.3, respectively. We then performed secondary screens through the overexpression of these genes. This was performed in 2 UC cells lines without amplification of 1q23.3, RT-4 (grade 1 UC) and RT-112 (grade 2 UC) along with an immortalized bladder cell line, TRT-HU1. These secondary screens include assessment of morphology, proliferation, clonogenicity, anchorage independence by soft agar colony formation, migration and invasion by Boyden chambers, and in-vivo tumor growth.

Results: RIGER analysis of the arrayed shRNA screen identifies DEDD, F11R, ARHGAP30 and PVRL4 knockdown as inducing decreased proliferation (FDR <0.25) when comparing 1q23.3 amplified to non-amplified lines. Overexpression of those genes preliminarily shows: 1) ARHGAP30 overexpression changes the morphology from an epithelial to an amoeboid state in a subpopulation of both cancer cell lines. 2) DEDD overexpression confers a modest proliferative advantage in both UC cell lines. 3) No increased clonogenic potential or invasion/migration is observed when these genes are overexpressed. 4) DEDD and ARHGAP30 confer anchorage independence in RT-112 although this is not seen in RT-4 or TRT-HU1.

Conclusions: ARHGAP30 and DEDD are potential drivers within the amplification of 1q23.3 and possibly mediators of advanced urothelial carcinoma. Detailed biological mechanisms and cooperativity of these genes requires further investigation.

Citation Format: Andrew L. Hong, Markus Riester, Anna C. Schinzel, Amy Schlauch, Rosalyn M. Adam, David J. Kwiatkowski, Jonathan E. Rosenberg, William C. Hahn. ARHGAP30 and DEDD as potential genomic drivers of invasive urothelial carcinomas. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3433. doi:10.1158/1538-7445.AM2014-3433
Abstract 3437: Differential transcript expression in nasopharyngeal carcinoma by cDNA microarray analysis
Authors: Yip Timothy T.C. , Kwong Dora L.W. , Ngan Roger K.C. , Lim Cadmon K.P. , Cheng Wai Wai , Ma Victor W.S. , Law Stephen C.K. , Tse Loretta
Published: May 2015

By cDNA microarray, expression profiling was carried out in 360 paired tumor & normal biopsies from 60 nasopharyngeal cancer (NPC) patients at 3 time points, namely, at diagnosis of NPC, on treatment with an anti-inflammatory drug, Celecoxib (Celec) and on radiotherapy (RT). We found 1 cluster of 82 transcripts with statistically significant difference in expression between tumor & normal tissues at diagnosis, a second cluster of 214 tumor versus normal transcripts on Celec treatment and finally a third cluster of 202 tumor versus normal transcripts on RT. Differential expression of selected tumor associated transcripts was confirmed by RT-PCR. These include 8 tumor up-regulated transcripts with 3 encoding for immunological/inflammatory molecules, IL8, CXCL9 & CXCL10, 1 matrix metallopeptidase, MMP12, which break down extracellular matrix and thus is associated with cancer metastasis, 1 osteonectin protein which is involved in cell shape changes and thus related to cancer invasion, SPARC, 1 bone morphogenic protein with a role in tissue differentiation, GREM1, 1 GTPase-activating protein for phorbol ester receptor, CHN1 and a sorting nexin family member with a role in intracellular trafficking, SNX10. Genes which were prominently down-regulated in tumor were a variety of polymorphic xenobiotic detoxification enzyme genes or genes with antioxidant function (e.g. GSTA1, GSTA2, GSTA3, CYP4B1, ALDH1A1, ADHFE1, MAOB, NQO1 & OAT) which can detoxify carcinogens or other harmful chemicals or have anti-oxidant effects. Many genes were also down-regulated by RT in tumor tissues and one notable example was TOP2A which encodes topoisomerase 2A involving in chromosome condensation & chromatid separation during DNA replication, CLDN1 which has tight junction adhesion function, ANLN which is required for cytokinesis, and STMN1 which can regulate microtubule filament. These genes had good potential in applying for clinical applications in diagnosis or treatment of NPC in the future. Further exploration of the clinical applications of these genes is underway.

Citation Format: Timothy T.C. Yip, Dora L.W. Kwong, Roger K.C. Ngan, Cadmon K.P. Lim, Wai Wai Cheng, Victor W.S. Ma, Stephen C.K. Law, Loretta Tse. Differential transcript expression in nasopharyngeal carcinoma by cDNA microarray analysis. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3437. doi:10.1158/1538-7445.AM2014-3437
Abstract 3438: Integrated analysis of germline and somatic variants in renal clear cell carcinoma
Authors: Wang Jiayin , Lu Charles , Xie Mingchao , Tripathi Piyush , McLellan Michael , Chen Feng , Johnson Kimberly J. , Ding Li
Published: May 2015

Each year in the United States, there are approximately 54,000 new cases of kidney and upper urinary tract cancers and 13,000 deaths. Renal clear cell carcinoma (RCC) is the most common type of kidney cancer comprising more than 75% of cases. It is known that certain genetic syndromes strongly increase risk for renal cell carcinoma including Von Hippel Lindau disease (VHL) that is caused by germline mutations in the VHL tumor suppressor gene. However, there are a large number of cases with familial RCC that are not explained by germline VHL mutations. Publically available data generated through large scale genomic sequencing projects such as The Cancer Genome Atlas (TCGA) provides new opportunities for identifying both known and novel germline cancer susceptibility variants. Moreover, the availability of matched tumor and normal genomic sequencing data from cancer patients (including RCCC) through TCGA allows for integrated analyses of germline and somatic interactions. Toward this end, we analyzed the TCGA exome sequencing data from 499 RCC cases. We employed GATK, VarScan, and Pindel tools to identify germline single nucleotide variants and indels and identified 10,492 candidate germline rare truncation variants (X truncation variants were in cancer genes). All the truncation candidates in commonly mutated cancer driver genes were manually reviewed, resulting in a total of 119 high confidence germline truncation variants in cancer genes. To date, we have identified one nonsense mutation in the VHL gene. In addition, there was one nonsense mutation and a frame-shift deletion in WRN and a nonsense and a splice-site mutation in BAP1, a recently identified RCC predisposition gene. Moreover, we also identified 6,079 candidate missense variants in known cancer driver genes, among a total of 179,066 rare missense variants. Future directions for this project will be to use the burden analysis to determine genes with significant enrichment for deleterious variants. We will also examine loss of heterozygosity patterns to evaluate germline somatic interactions.

Citation Format: Jiayin Wang, Charles Lu, Mingchao Xie, Piyush Tripathi, Michael McLellan, Feng Chen, Kimberly J. Johnson, Li Ding. Integrated analysis of germline and somatic variants in renal clear cell carcinoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3438. doi:10.1158/1538-7445.AM2014-3438
Abstract 3440: Cell-based screening identify gene expression signature correlated with sensitivity to an MEK1/2 inhibitor
Authors: Zhang Jing
Published: May 2015

The mitogen-activated extracellular signal-regulated kinase (MEK) family is one of the key kinase regulators involved in tumor oncogenic growth and progression. Multiple MEK inhibitors are currently under clinical investigation for cancer treatment and more agents targeting MEK are under preclinical development. The mechanisms underlying resistance to MEK inhibitor are unclear. Here we examined the anti-proliferation activity of a new small molecule MEK inhibitor in a panel of cell lines harboring either ras or raf mutation. We found 23 lines that are sensitive to the inhibitor and 24 lines are resistant. We compared gene expression by Affymetrix U219 arrays and gene copy number by Affymetrix SNP6.0 arrays in these two cell line groups. With a p-value cutoff set at0.01, we identified 549 genes differentially expressed and 328 genes demonstrated different copy numbers between the two groups. 23 genes were both detected by gene expression profiling and gene copy number data. These genes include MEK family members MAP2K3 and MAP2K7, as well as other important molecule markers in cancer biology, including PIK3R2 and BRCA2. The results suggested a potential genomic signature that could be predictive of response to the MEK inhibitor in vitro. Further investigation and validation in patient-derived xenograft (PDX) models will undoubtfully provide valuable information for selecting patients that are likely to receive clinical benefit.

Citation Format: Jing Zhang. Cell-based screening identify gene expression signature correlated with sensitivity to an MEK1/2 inhibitor. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3440. doi:10.1158/1538-7445.AM2014-3440
Abstract 3454: Integrin-linked kinase and proximal signaling contributes to the cisplatin resistance of ovarian cancer cells
Authors: Soto Daniel , Valiyeva Fatima , Echevarria Ileabett , Reyes Jeyshka , Vivas-Mejia Pablo E.
Published: May 2015

Platinum-based chemotherapy has been used for more than four decades for ovarian cancer patients. Although initially most ovarian tumors respond well to cisplatin treatment, many of the tumors become resistant to chemotherapy. Several drug resistant-associated genes have been identified by gene expression arrays in drug sensitive and drug resistant cells. Recent evidence indicates that posttranscriptional and posttranslational regulation of gene expression governs the function of several proteins associated with cancer initiation, progression and drug resistance. To reveal additional proteins associated with cisplatin resistance, we performed a proteomic analysis in cisplatin sensitive (A2780) and cisplatin resistant (A2780CP20) ovarian cancer cells. Forty-seven, out of 148 identified proteins, were differentially abundant in A2780CP20 compared with A2780 cancer cells. Western blot analysis was used to confirm the proteomic results. Particularly, the Integrin-Linked Kinase-Associated Serine/Threonine Phosphatase 2C (ILKAP) was decreased in A2780CP20 compared with A2780 cells. The phosphorylation levels of the Integrated-Linked Kinase ILK (the major ILKAP effector), protein kinase B (AKT), and glycogen synthase kinase 3β (GSK3β) (two ILK downstream effectors) were higher in A2780CP20 compared with A2780 cells. Small-interference RNA (siRNA)-targeted ILK, reduced the phosphorylation levels of AKT and GSK3β; and reduced cell proliferation and invasion of the A2780CP20 cells. Most important, combination of inactive ILK-targeted siRNA doses plus inactive cisplatin doses reduced cell growth in a synergistic-like way. These results indicate that protein phosphorylation of the ILK/AKT/GSK3β cascade play a vital role in cisplatin resistance, and propose ILK as a therapeutic target for advanced and drug resistant ovarian cancer.

Citation Format: Daniel Soto, Fatima Valiyeva, Ileabett Echevarria, Jeyshka Reyes, Pablo E. Vivas-Mejia. Integrin-linked kinase and proximal signaling contributes to the cisplatin resistance of ovarian cancer cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3454. doi:10.1158/1538-7445.AM2014-3454