bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3034: Determining the profiles and parameters for gene amplification testing of growth factor receptors in lung cancer.
Authors: Pros Eva , Lantuejoul Sylvie , Sanchez-Verde Lydia , Castillo Sandra D. , Suarez-Gauthier Ana , Conde Esther , Cigudosa Juan C. , Lopez-Rios Fernando , Torres-Lanzas Juan , Castellví Josep , Ramon y Cajal Santiago , Brambilla Elisabeth , Sanchez-Cespedes Montse
Published: August 2014

Growth factor receptors (GFRs) are amenable to therapeutic intervention in cancer and it is important to select patients appropriately.

One of the mechanisms for activation of GFRs is gene amplification (GA) but discrepancies arising from the difficulties associated with data interpretation and the lack of agreed parameters confound the comparison of results from different laboratories. Here, we attempt to establish appropriate conditions for standardization of the determination of GA in a panel of GFRs. A NSCLC tissue microarray panel containing 302 samples was screened for alterations at ALK,

FGFR1, FGFR2, FGFR3, ERBB2, IGF1R, KIT, MET and PDGFRA by FISH, immunostaining and/or real-time quantitative RT-PCR. Strong amplification was found for FGFR1, ERBB2, KIT/PDFGRA and MET, with frequencies ranging from 1 to

6%. Thresholds for overexpression and GA were established. Strong immunostaining was found in most tumors with ERBB2, MET and KIT amplification, although some tumors underwent strong immunostaining in the absence of GA. KIT and PDFGRA were always co-amplified, but only one tumor showed PDGFRA overexpression, indicating that KIT is the main target.

Amplification of FGFR1 predominated in squamous cell carcinomas, although the association with overexpression was inconclusive. Interestingly, alterations at ALK,

MET, EGFR, ERBB2 and KRAS correlated with augmented levels of phospho-S6 protein, suggesting activation of the mTOR pathway, which may prove useful to pre-select tumors for testing.

Overall, here, we provide with parameters for the determination of GA at ERBB2, MET, KIT, and PDGFRA which could be implemented in the clinic to stratify lung cancer patients for specific treatments.

Citation Format: Eva Pros, Sylvie Lantuejoul, Lydia Sanchez-Verde, Sandra D. Castillo, Ana Suarez-Gauthier, Esther Conde, Juan C. Cigudosa, Fernando Lopez-Rios, Juan Torres-Lanzas, Josep Castellví, Santiago Ramon y Cajal, Elisabeth Brambilla, Montse Sanchez-Cespedes. Determining the profiles and parameters for gene amplification testing of growth factor receptors in lung cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3034. doi:10.1158/1538-7445.AM2013-3034
Abstract 3040: Cavin-1 alters oncogenic effects of caveolin-1 microdomains in prostate cancer.
Authors: Moon Hyeongsun , Lee C Soon , Sharma Sowmya , Inder Kerry , Black Debra , Lê Cao Kim-Anh , Winterford Clay , Ling Patrick ,   the Australian Prostate Cancer BioResource, Craik David , Parton Robert , Russell Pamela , Hill Michelle
Published: August 2014

Caveolin-1 is associated with prostate cancer progression and has been suggested to be a biomarker and therapeutic target. Mature caveolin-1 resides in lipid raft domains at the plasma membrane, where it forms caveolae upon co-expression of cavin-1 (also known as PTRF; polymerase I and transcript release factor). In the absence of cavin-1, caveolin-1 does not form caveolae but are found on flat membrane. To determine if oncogenic caveolin-1 in prostate cancer is present in caveolae, we examined the relative expression of caveolin-1 and cavin-1 in normal, non-malignant and malignant prostate tissues. We found that caveolin-1 is induced in prostate cancer without cavin-1, an expression pattern mirror in the PC3 cell line. Previously we showed that expression of cavin-1 in PC3 cells recruits flat membrane caveolin-1 to caveolae and reduced transmigration. Here we report that cavin-1 expression reduces tumour size and metastasis of PC3 cells in vivo, using an orthotopic prostate cancer xenograft mouse model. To determine if cavin-1 acts by neutralizing oncogenic caveolin-1, we expressed cavin-1 in caveolin-1 negative LNCaP and 22Rv1 cells. While caveolin-1 over-expression increased anchorage-independent growth of LNCaP and 22Rv1 cells, cavin-1 over-expression had no effect. Furthermore, co-expression of cavin-1 in LNCaP+caveolin-1 cells reversed caveolin-1 effect. Taken together, these results suggest that caveolin-1 in prostate cancer is present outside of caveolae, and caveola formation by cavin-1 co-expression alters the oncogenic effect of non-caveolar caveolin-1 microdomains.

Citation Format: Hyeongsun Moon, C Soon Lee, Sowmya Sharma, Kerry Inder, Debra Black, Kim-Anh Lê Cao, Clay Winterford, Patrick Ling, the Australian Prostate Cancer BioResource, David Craik, Robert Parton, Pamela Russell, Michelle Hill. Cavin-1 alters oncogenic effects of caveolin-1 microdomains in prostate cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3040. doi:10.1158/1538-7445.AM2013-3040
Abstract 3047A: Differential subcellular localization and trafficking of RET isoforms.
Authors: Crupi Mathieu J. F. , Richardson Douglas S. , Mulligan Lois M.
Published: August 2014

The RET proto-oncogene encodes a receptor tyrosine kinase (RTK) necessary for development of the kidneys and enteric nervous system. Two distinct isoforms of the growth factor receptor arise through alternative splicing of the 3’ exons, and are termed RET9 and RET51 to denote the number of amino acids in each C-terminal tail. These isoforms induce unique transphosphorylation patterns on intracellular tyrosine residues, differentially bind downstream signalling proteins, and possess different intrinsic abilities to cause cellular transformation. Subcellular localization and trafficking of RTKs have increasingly been recognized as important post-receptor activation regulatory mechanisms, involving protein-protein interactions. Here, we explore the differential subcellular localization and trafficking of RET isoforms in human cell line models. We have observed that RET51 is efficiently matured and localized to the cell membrane, while immature RET9 accumulates in a perinuclear region and matures more slowly, leading to lower relative expression on the plasma membrane. Subsequent to RET activation, both isoforms at the cell surface are internalized through the clathrin coated-pit pathway into endosomes, where signaling persists. Although RET51 is internalized more rapidly than RET9, a portion of RET51 molecules recycles back to the plasma membrane, maintaining high expression levels at the cell surface. Recycling prevents efficient degradation of RET51, allowing for prolonged downstream signaling relative to RET9. We predict that progress through these trafficking steps is modulated by specific protein interactions between RET isoforms and sorting proteins, and these interactions are also determined at later stages by modifications on RET itself, most notably ubiquitination. Future studies could allow us to identify these key proteins/binding sites which may differ between RET9 and RET51. Since RET51 has a longer residency time in an endosomal compartment, where sustained Erk1/2 signaling promotes mitogenesis, this may in part explain the greater transforming potential of RET51. Together, our data suggest that differences in RET isoform subcellular localization and trafficking would lead to differences in signal transduction and may in part explain the functional differences observed between RET isoforms.

Citation Format: Mathieu J. F. Crupi, Douglas S. Richardson, Lois M. Mulligan. Differential subcellular localization and trafficking of RET isoforms. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3047A. doi:10.1158/1538-7445.AM2013-3047A
Abstract 3050: Transcriptome sequencing identifies oncogenic microRNA signatures related to NF-kB, TP53 and Notch family members in head and neck squamous cell carcinoma.
Authors: Saleh Anthony D. , Si Han , Yang Xingping , Coupar Jamie , Chen Zhong , Van Waes Carter
Published: August 2014

MicroRNA dysregulation has been shown to be nearly universal in cancer biology, driving oncogenic signaling networks. However, microRNA mediated global regulatory mechanisms underlying malignant phenotypes of head and neck squamous cell carcinoma (HNSCC) remain unclear. In this study, we have performed genome-wide small RNA sequencing on a panel of nine HNSCC cell lines and three normal primary oral keratinocyte cell lines, as well as matched primary tumor and normal tissues using the SOLiD parallel sequencing platform. A panel of microRNAs with altered expression was identified, including several novel oncogenic microRNAs which are upregulated in parallel with altered activation of NF-KB c-REL, TP53 family members ΔNp63α and TAp73, and potentially involved in the down-modulation of p53 and Notch1, two genes most often mutated or inactivated in HNSCC. To examine the hypothesis that these microRNAs could potentially be regulated by NF-KB and p53 pathways, NF-KB inducer TNF-α was used to treat an HNSCC cell line with high expression of mtTP53, ΔNp63α and TAp73. Genome-wide small RNA sequencing revealed significant changes in microRNA expression modulated by TNF-α that overlaps with the panel of constitutively dysregulated microRNAs in HNSCC. Furthermore, genome-wide ChIP-sequencing revealed that TNF-α modulated DNA binding activities of c-REL, ΔNp63α and TAp73 aligned within the regulatory regions of several differentially expressed microRNAs. Modulation of these microRNAs with anti-miR repression altered cell cycle and reduced proliferation in HNSCC cell lines, which confirmed the oncogenic activity of the microRNAs and their potential for therapeutic targeting.

Citation Format: Anthony D. Saleh, Han Si, Xingping Yang, Jamie Coupar, Zhong Chen, Carter Van Waes. Transcriptome sequencing identifies oncogenic microRNA signatures related to NF-kB, TP53 and Notch family members in head and neck squamous cell carcinoma. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3050. doi:10.1158/1538-7445.AM2013-3050

Note: This abstract was not presented at the AACR Annual Meeting 2013 because the presenter was unable to attend.
Abstract 3056: miR-125b suppresses p14ARF and modulates p53-dependent and p53-independent apoptosis in prostate cancer cells.
Authors: Shi Xu-Bao , Amir Sumaira , Ma Ai-Hong , Xue Lingru , deVere White Ralph W.
Published: August 2014

Increasing evidence has revealed that miR-125b functions as a potential oncogene in different types of human cancer. Our previous studies demonstrated that miR-125b directly targets p53, Puma and Bak1 in human prostate cancer (CaP). To further elucidate the role of miR-125b in modulating the p53 network, we utilized a bioinformatics approach to identify miR-125b-regulated, CaP-relevant tumor suppressor gene, and identified p14ARF gene as a miR-125b target. We first evaluated the effect of miR-125b on the regulation of p14ARF expression, and found that miR-125b treatment induced reduction of p14ARF by 80% in LNCaP CaP cells and 60% in 22Rv1 CaP cells. Conversely, anti-miR-125b increased the p14ARF level by 40% in LNCaP and 30% in 22Rv1. We then performed luciferase assay to validate that the miR-125b binding site in the 3’UTR of p14ARF mRNA is responsible for the regulation of p14ARF by miR-125b. Since p14ARF accelerates Mdm2 degradation, resulting in p53 up-regulation, we thus assessed the effect of miR-125b on the expression of Mdm2 and p53. Compared with miRNA control, miR-125b treatment induced a dramatic increase in Mdm2 and a significant reduction of p53 in CaP cell lines, as well as in miR-125b-overexpressed xenograft tumor. Additionally, miR-125b treatment resulted in a reduction of interaction between p14ARF and Mdm2. Next, we examined the effect of miR-125b on CaP cell proliferation. Both LNCaP and 22Rv1 cells were transfected with miR-125b and cell proliferation was determined by clone formation assay. When compared with the control, miR-125b treatment increased clonogenic survival of CaP cells, while anti-miR-125b caused a dramatic reduction in the number of colonies. We evaluated the effect of downregulation of miR-125b on apoptosis in p53-positive 22Rv1 CaP cells. Data from TUNEL assay demonstrated that anti-miR-125 caused 63% of cells to undergo apoptosis. Similarly, in p53-negative PC3 CaP cells, miR-125b treatment induced a 2-fold increase in survival, while anti-miR-125b caused 50% of cells to undergo apoptosis. In summary, we observed that overexpression of miR-125b negatively regulates the expression of p14ARF, resulting in promotion of cell proliferation and inhibition of apoptosis. Inactivation of miR-125b using anti-miR-125b affects apoptosis involving both p53-dependent and p53-independent pathways. Therefore, our data suggest that miR-125b has a great potential to be used as a therapeutic target for CaP treatment.

Citation Format: Xu-Bao Shi, Sumaira Amir, Ai-Hong Ma, Lingru Xue, Ralph W. deVere White. miR-125b suppresses p14ARF and modulates p53-dependent and p53-independent apoptosis in prostate cancer cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3056. doi:10.1158/1538-7445.AM2013-3056
Abstract 3077: TMPRSS2-ERG regulation of miRNA in prostate cancer.
Authors: Kim Jung , Wu Longtao , Jin Hong-Jian , Zhao Jonathan , Yu Jindan
Published: August 2014

Prostate cancer is the cause of the second highest number of cancer related deaths in men in the United States, with the highest number of new estimated cases every year. In addition, the survival rate in localized and regional prostate cancer is 100%, while metastatic prostate cancer is about 29%, with a wide spectrum of biological deregulations. MicroRNA (miRNA) is a small non-coding RNA, about 22 nucleotides long, that mostly targets the 3’UTR of a gene for translational repression or mRNA degradation. More recently, miRNAs have been shown to play an important role in tumorigenesis, cancer progression, and metastasis. MiR-200c is one of the miRNA widely studied in cancer, as it has a role in regulating epithelial-mesenchymal transition (EMT). One of the ways in which miR-200c inhibits EMT is through its ability to downregulate ZEB1, an EMT activator that primarily functions by repressing E-cadherin. ERG is a transcription factor that is often fused with TMPRSS2 in prostate cancer and promotes cancer progression. In our study, we investigated the interaction of ERG and miR-200c, two factors that play important role in cancer progression, in prostate cancer.

First, we identified ERG regulated miRNAs through utilizing miRNA profiling in LNCaP cells that were overexpressing ERG and in VCaP cells that have ERG knocked down. We selected miRNAs that are repressed upon ERG overexpression and derepressed upon ERG knockdown and also overlapped the results with ERG ChIP-seq data to identify ERG regulated miRNAs. Through this process we discovered that miR-200c is a good ERG-targeted miRNA. To validate the bioinformatics results, we used ERG overexpression in LNCaP and ERG knockdown in VCaP to measure mature miR-200c levels by miRNA assay. In addition, ERG ChIP-qPCR and miR-200c promoter luciferase assays showed that ERG binds at the miR-200c promoter. Next, to identify functional roles of miR-200c in ERG overexpressed cells, we performed invasion and migration assays to show that miR-200c plays an important role in the inhibition of ERG-mediated cancer progression.

This study identifies an important factor, miR-200c, whose loss mediates the progression of TMPRSS2:ERG fusion-positive prostate cancer. According to our results, it might be possible to use miR-200c as a future therapeutic for ERG positive patients. This work was supported in part by the Research Scholar Award RSG-12-085-01 (to J.Y.) from the American Cancer Society, Department of Defense PC080665 (to J.Y.) and NRSA pre-doctoral fellowship T32 CA080621 (to J.K.).

Citation Format: Jung Kim, Longtao Wu, Hong-Jian Jin, Jonathan Zhao, Jindan Yu. TMPRSS2-ERG regulation of miRNA in prostate cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3077. doi:10.1158/1538-7445.AM2013-3077
Abstract 3078: Mifepristone suppresses triple-negative breast cancer cell growth via miR-153 mediated down-regulation of KLF5 expression.
Authors: Liu Rong , Chen Ceshi
Published: August 2014

Triple-negative breast cancer (TNBC) is one of the most malignant breast cancers without effective therapeutic approaches currently. It is reported that KLF5 is over-expressed in TNBC; and KLF5 depletion suppresses TNBC cell proliferation, cell survival and tumor growth in vivo significantly, which implies that KLF5 may be a therapeutic target for TNBC treatment. As a progesterone receptor antagonist, mifepristone (MIF) has been found to have anti-tumor activity in multiple hormone-dependent cancers, including luminal type breast cancers. Interestingly, we found that MIF is also able to suppress KLF5 protein expression in TNBC cell and inhibit TNBC cell growth in vitro and in vivo. We further demonstrated that MIF does not suppress KLF5 mRNA expression at all, but down-regulates KLF5 protein level via up-regulating miR-153 expression in TNBC cells. We found miR-153 mimics can down-regulates KLF5 protein level in TNBC cells similar to MIF does; and consistently, miR-153 inhibitor rescues MIF-induced down-regulation of KLF5 protein expression. Taken together, our findings suggest that MIF is able to inhibit TNBC growth possibly via suppressing KLF5 expression; and our research may provide a new possible chemotherapeutic for the treatment of TNBC.

Citation Format: Rong Liu, Ceshi Chen. Mifepristone suppresses triple-negative breast cancer cell growth via miR-153 mediated down-regulation of KLF5 expression. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3078. doi:10.1158/1538-7445.AM2013-3078
Abstract 3084: miR509 suppresses brain metastasis of breast cancer cells by modulating RhoC and TNF alpha.
Authors: Xing Fei , Okuda Hiroshi , Kobayashi Aya , Pandey Puspa R. , Sharma Sambad , Mo Yin-Yuan , Liu Yin , Wu Kerui , Watabe Kounosuke
Published: August 2014

Brain metastasis of breast cancer profoundly affects the cognitive and sensory functions as well as morbidity of patients, and the rate of one year survival among these patients remains less than 20%. Because of the location of metastatic lesions, a surgical approach is limited and most chemotherapeutic drugs are ineffective due to the blood brain barrier (BBB). Despite this clinical importance, the molecular basis of breast tumor metastasis to the brain is poorly understood. Recent active research in microRNA identified a series of this type of molecules that are involved in tumor progression in various tumors as oncogenes and tumor suppressors. In this study, we have isolated RNA from Formalin-Fixed Paraffin-Embedded (FFPE) samples obtained from primary breast tumors and also from brain metastatic lesions followed by microRNA profiling. The results of our profile analysis for these samples revealed that primary tumor samples expressed significantly high level of miR509 compared to brain metastatic lesions. We also found that this microRNA was capable of modulating two essential genes for invasion, RhoC and TNF alpha. Interestingly, high expression of TNF alpha was significantly and inversely correlated to brain metastasis-free survival of breast cancer patients. Importantly, we found that the expression of miR509 significantly suppressed the ability of cancer cells to metastasize to the brain in our animal model. We also examined the expression of TNF alpha in brain-metastatic lesions and found that it was indeed significantly up-regulated. Furthermore, the results of our in vitro experiments indicate that miR509 attenuates the abilities of invasion of breast cancer cells by modulating the RhoC expression. These results suggest that miR509 may serve as a biomarker or therapeutic targets for brain metastasis of breast cancer.

Citation Format: Fei Xing, Hiroshi Okuda, Aya Kobayashi, Puspa R. Pandey, Sambad Sharma, Yin-Yuan Mo, Yin Liu, Kerui Wu, Kounosuke Watabe. miR509 suppresses brain metastasis of breast cancer cells by modulating RhoC and TNF alpha. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3084. doi:10.1158/1538-7445.AM2013-3084

Note: This abstract was not presented at the AACR Annual Meeting 2013 because the presenter was unable to attend.
Abstract 3092: Deregulation of miR-10b in osteosarcoma.
Authors: Chen K. Amy , Sand Jordan P. , Janeway Katherine A. , Orkin Stuart H.
Published: August 2014

Osteosarcoma (OS) is the most common primary bone malignancy found in children and adolescents, but the underlying molecular pathogenesis is not yet well understood. Utilizing a genetically engineered mouse model of OS that faithfully recapitulates salient features of the human disease, we are able to examine potential molecular contributions, such as that of a class of small, regulatory RNAs called microRNAs (miRNAs), in the processes of tumor proliferation and tumor progression and/or metastasis.

MiRNA expression profile of tumors isolated from primary and secondary (metastatic) tumor sites of the mouse model revealed significant differential expression of miRNAs between the two groups. MiR-10b was significantly down-regulated in metastatic tumors in comparison with their primary counterparts, and thus was hypothesized to play important roles in the process of metastatic progression and/or tumorigenesis. To determine its impact on invasiveness and/or proliferation of the tumor cells, miR-10b was re-expressed in metastatic cell lines and introduced through tail-vein or intramuscular injections into scid/nude mice. Quantitative analysis using a bioluminescence imaging system showed delayed metastatic progression, lessened pulmonary tumor burden, as well as decreased primary tumor size in animals injected with tumor cells over-expressing miR-10b as compared to the controls. These data suggest that miR-10b down-regulation contributes to the processes of metastasis and tumorigenesis in OS through altered interaction with its target genes. Currently, putative miR-10b gene targets generated by microarray and RNA-seq analysis are being validated to provide a better understanding of the molecular mechanisms underlying miR-10b regulation in OS, and to facilitate the discovery of pathways amenable to manipulation with novel therapeutics.

Citation Format: K. Amy Chen, Jordan P. Sand, Katherine A. Janeway, Stuart H. Orkin. Deregulation of miR-10b in osteosarcoma. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3092. doi:10.1158/1538-7445.AM2013-3092
Abstract 3112: The homeoprotein DLX4 induces topoisomerase IIα expression but reduces sensitivity of tumor cells to topoisomerase II poisons.
Authors: Trinh Bon Q. , Ko Song Yi , Barengo Nicolas , Lin Shiaw-Yih , Naora Honami
Published: August 2014

Chemotherapeutic drugs such as anthracyclines and etoposide that target topoisomerase II (TOP2) induce tumor cell death by causing accumulation of DNA double-strand breaks (DSBs). Experimental studies have shown that overexpression of topoisomerase IIα (TOP2A) increases sensitivity to TOP2 poisons. However, the clinical value of TOP2A in predicting responsiveness to TOP2 poisons has been controversial and it is not known why some TOP2A-overexpressing tumors respond poorly to these drugs. We identified that expression of TOP2A, the gene encoding TOP2, is induced by DLX4, a homeoprotein that is overexpressed in breast and ovarian cancers. Analysis of breast cancer datasets revealed that TOP2A-High cases that expressed DLX4 at low levels responded more favorably to anthracycline-based chemotherapy than TOP2A-High cases that also highly expressed DLX4. Overexpression of TOP2A alone in tumor cells increased sensitivity to TOP2 poisons. However, DLX4 decreased sensitivity to these drugs irrespective of its induction of TOP2A. We identified that DLX4 stimulates repair of DSBs and interacts with Ku proteins that are components of the non-homologous end-joining machinery. DLX4 did not reduce levels of TOP2 poison-induced DSBs in Ku-deficient cells, but reduced the level of DSBs when Ku was reconstituted in these cells. Taken together, our findings indicate that DLX4 induces TOP2A expression, but reduces the sensitivity of tumor cells to TOP2 poisons by stimulating Ku-dependent repair of DSBs. These dual and opposing effects of DLX4 could explain why some tumors with elevated TOP2A levels are not highly sensitive to TOP2 poisons.

Citation Format: Bon Q. Trinh, Song Yi Ko, Nicolas Barengo, Shiaw-Yih Lin, Honami Naora. The homeoprotein DLX4 induces topoisomerase IIα expression but reduces sensitivity of tumor cells to topoisomerase II poisons. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3112. doi:10.1158/1538-7445.AM2013-3112
Abstract 3132: Beta-catenin regulates c-Myc and CDKN1A expression in breast cancer cells.
Authors: Xu Jinhua , Chen Yinghua , Khramtsov Andrey , Huo Dezheng , Goss Kathleen , Olopade Olufunmilayo I.
Published: August 2014

Basal-like breast cancer (BLBC) is associated with a poor prognosis and lacks therapeutic targets. The molecular pathways leading to the development of basal-like tumors are not well understood. We reported recently that nuclear and cytosolic accumulation of beta-catenin was enriched in BLBC, suggesting Wnt pathway activation in this specific subtype. c-Myc has been placed as a downstream effector of Wnt/beta-catenin pathway in colorectal cancer. However, exactly how the Wnt pathway regulates c-Myc in breast cancer and the biological significance of this regulation are poorly understood.

In this study, we found that c-Myc is highly expressed in the basal-like subtype in comparison to the other four breast cancer subtypes by IHC staining. In addition, we confirmed the concordant expression of beta-catenin and c-Myc in basal-like breast tumors. Both c-Myc and beta-catenin proteins are highly expressed in the basal-like cell lines in comparison with non-basal-like cell lines. After silencing beta-catenin using siRNA, c-Myc expression was decreased in none-BLBC cells. In contrast, c-Myc mRNA and protein expression was up-regulated in the BLBC cell lines, suggesting that beta-catenin regulates c-Myc expression in a cell type dependent manner in breast cancer. Although we did not detect any TCF promoter activity in all breast cancer cell lines, decreased c-Myc promoter activity was observed after inhibiting beta-catenin by siRNA in non-BLBC cells; however, inhibition of beta-catenin or over-expression of dominant-negative LEF had no effect on c-Myc promoter activity in BLBC cell lines. In contrast, CDKN1A mRNA and p21 protein expression were significantly increased in all breast cancer cell lines upon beta-catenin silencing. Interestingly, inhibiting beta-catenin expression alone does not induce apoptosis in breast cancer cell lines despite c-Myc regulation,, but we observed a modest increase of cells in G1 and decrease of cells in S phase in the cell lines with beta-catenin silencing. Collectively, these findings suggest that the regulation of c-Myc in breast cancer cells is dependent on the molecular subtype, and that beta-catenin-mediated control of c-Myc and p21 may control the balance of cell death and proliferation in breast cancer by both TCF-dependent and TCF-independent mechanisms.

Citation Format: Jinhua Xu, Yinghua Chen, Andrey Khramtsov, Dezheng Huo, Kathleen Goss, Olufunmilayo I. Olopade. Beta-catenin regulates c-Myc and CDKN1A expression in breast cancer cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3132. doi:10.1158/1538-7445.AM2013-3132
Abstract 3134: Exomic landscape of somatic variation in uterine leiomyomas.
Authors: Mäkinen Netta , Vahteristo Pia , Aaltonen Lauri A.
Published: August 2014

Uterine leiomyomas, or fibroids, are benign tumors for which the lifetime risk in women by the age of 50 has been estimated to be nearly 70%. Fibroids arise from smooth muscle cells of the myometrium. Regardless of their benign nature, leiomyomas can cause difficult symptoms, such as excessive bleeding, abdominal pain, pregnancy complications, and infertility. They are also the most common cause for hysterectomy. Considering the prevalence of these lesions, relatively little is known about the mechanisms of their growth and tumorigenesis. We have previously showed by exome sequencing that mediator complex subunit 12 (MED12) exon 2 is mutated in approximately 70% of sporadic uterine leiomyomas. The aim of this study is to examine molecular genetic background of MED12 mutation positive lesions, as well as to identify additional recurrent somatic variants from those fibroids that do not harbor MED12 mutations and cannot be explained for example by mutations in fumarase hydratase (FH) which has already previously been associated with the predisposition to uterine leiomyomas. We performed whole exome sequencing of 28 uterine leiomyomas and their corresponding myometrium pairs. Exome sequencing data of the uterine leiomyomas was filtered against data from the respective normal tissue, the 1000Genomes Project (93 FIN samples, Phase 1 release v2), rs-coded SNPs taken from the Database of Single Nucleotide Polymorphisms (dbSNP, Build 132), and 70 in-house control exomes to exclude germ line variation from the analysis. Both exomic regions and sequences within two base pairs of the exon-intron boundaries were analyzed. Synonymous coding and intronic variants were not included to the study. All potentially significant mutations and their somatic status were validated by Sanger sequencing. Based on our results, uterine leiomyomas harbor less mutations than malignant tumors. We have not identified mutations in genes, such as TP53 or PTEN, which are associated with tumor malignancy. In addition to MED12, there are no other recurrent point mutations or small insertion-deletion type of mutations in sporadic uterine leiomyomas. Our results emphasize the major role of MED12 in tumorigenesis of these lesions.

Citation Format: Netta Mäkinen, Pia Vahteristo, Lauri A. Aaltonen. Exomic landscape of somatic variation in uterine leiomyomas. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3134. doi:10.1158/1538-7445.AM2013-3134
Abstract 3148: Mapping mobile DNAs in human cancer cell lines.
Authors: Zampella John G. , Yang Wan Rho , Rodric Nemanja , Huang Cheng Ran Lisa , Welch Jane , Gnanakkan Veena P. , Cornish Toby C. , Boeke Jef D. , Burns Kathleen H.
Published: August 2014

Genomic instability represented by genetic and epigenetic changes is a hallmark of oncogenesis and tumor progression. The study of genomic instability using conventional methods has necessarily been restricted to regions of unique DNA and excludes repetitive DNA sequences. Repeat elements derived from mobile DNAs comprise a large portion of the human genome and are known sources of genetic variation in humans. Active transposable elements (TE) have recently been shown to cause genetic heterogeneity in tumors through somatic insertions. The functional impact of inherited and somatically acquired TE in cancer development and progression remains unproven. To begin to understand the roles of these mobile DNA elements in transformed cells, we used a novel genomic method to map TEs in a panel of well-characterized human cancer cell lines. Using transposon insertion profiling (TIP)-chip we discover transposition competent families of L1 and Alu TEs. This technique recovered many known reference and polymorphic insertions, novel polymorphisms shared by unrelated tumor cell lines, as well as unique, potentially tumor-specific insertions. Insertions were discovered in several important oncogenes such as KRAS, ALK and BRAF. Most insertions discovered were intronic or intergenic, however 16 TEs were putative exonic. Analysis of the novel and polymorphic insertions revealed a relative enrichment of L1 and Alu insertions in cancer gene loci that are mutated in human cancers or implicated in tumor development in forward genetic screens (p=7.74e-10). We also report associations of individual insertion sites with cellular phenotypes, including DNA methylation status, RNA and protein expression, and drug sensitivities. Epigenetic, gene expression and gene pathway associations were evaluated to delineate local (cis) from pathway (trans) effects. We identified several examples of cis effects of TEs on gene structure as well as nearly 250 curated proliferation pathways affected by a TE insertion. These data suggest that TE insertions may function locally to alter genomic structure or act more globally in concert with other genetic mechanisms to promote the development and progression of several common tumor types.

Citation Format: John G. Zampella, Wan Rho Yang, Nemanja Rodric, Cheng Ran Lisa Huang, Jane Welch, Veena P. Gnanakkan, Toby C. Cornish, Jef D. Boeke, Kathleen H. Burns. Mapping mobile DNAs in human cancer cell lines. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3148. doi:10.1158/1538-7445.AM2013-3148
Abstract 3149: Genetic alteration of lung adenocarcinomas with micropapillary component.
Authors: Furukawa Masashi , Toyooka Shinichi , Yamamoto Hiromasa , Tada Ryuhei , Hashida Shinsuke , Shien Kazuhiko , Soh Junichi , Asano Hiroaki , Tsukuda Kazunori , Ouchida Mamoru , Ichimura Kouichi , Miyoshi Shinichiro
Published: August 2014

Purpose: Adenocarcinoma with micropapillary component (AMPC) is known as one of aggressive subtypes among lung adenocarcinoma. The molecular profiles of AMPC have not been well characterized from the view point of difference from other adenocarcinomas. In this study, we analyzed molecular alterations of AMPC to investigate its molecular pathogenesis. Methods: We reviewed pathological reports of patients who underwent surgical resection for lung cancers between April 2004 and May 2012. Twenty-eight patients were recorded to possess micropapillary component among 674 patients diagnosed as lung adenocarcinoma. A total of 52 resected lung adenocarcinomas without micropapillary component were selected in the same period so as to adjust age, sex, and smoking status to those of the patients with micropapillary component. Mutational status was determined by the following two methods: SNaPshot assay based on multiplex PCR, primer extension, and capillary electrophoresis that was designed to assess for 38 somatic mutations in eight genes (AKT1, BRAF, EGFR, KRAS, MEK1, NRAS, PIK3CA, and PTEN), and a PCR-based sizing assay that assesses for EGFR exon 19 deletions, EGFR exon 20 insertions, and HER2 exon 20 insertions. EML4-ALK fusion was screened by ALK immunohistochemistry and confirmed using reverse transcriptional PCR assay and break-apart FISH assay.Results: As for genetic alterations, 13 (46.4%) of 28 lung adenocarcinomas with micropapillary component harbored mutually exclusive mutations: 9 (32.1%) EGFR mutation, 1 (3.3%) KRAS mutation and 3 (9.9%) EML4-ALK fusion. Adenocarcinomas without micropapillary component harbored 11 (21.2%) EGFR mutations and 8 (15.3%) KRAS mutations in a mutually exclusive manner, but had no EML4-ALK fusion genes. There were no mutations in AKT1, BRAF, EGFR, KRAS, MEK1, NRAS, PIK3CA, PTEN and HER2 genes in the both groups. EML4-ALK fusions appeared to occur significantly more frequent in AMPC than in control cases (P = 0.039).Conclusion: Although the size of our study was small, our study suggests the molecular pathogenesis of AMPC may be different from other adenocarcinoma, associated with its unique character among lung adenocarcinomas.

Citation Format: Masashi Furukawa, Shinichi Toyooka, Hiromasa Yamamoto, Ryuhei Tada, Shinsuke Hashida, Kazuhiko Shien, Junichi Soh, Hiroaki Asano, Kazunori Tsukuda, Mamoru Ouchida, Kouichi Ichimura, Shinichiro Miyoshi. Genetic alteration of lung adenocarcinomas with micropapillary component. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3149. doi:10.1158/1538-7445.AM2013-3149
Abstract 3167: Deciphering the genomic heterogeneity in malignant melanoma by genomic profiling of clonal tumor populations.
Authors: Lorber Thomas , Dietsche Tanja , Gsponer Joël , Rufle Alexander , Barret Michael , Sommer Lukas , Glatz Katharina , Ruiz Christian , Bubendorf Lukas
Published: August 2014

1) Background

Melanoma is a heterogeneous disease and even though consistent genetic patterns among different subtypes have been revealed, clinical trials have failed to significantly improve survival. However, most of the genomic studies do not take into consideration the underlying genomic heterogeneity. In this study, we used genomic profiling of sorted clonal tumor populations in order to investigate the genomic heterogeneity and the clonal evolution in malignant melanoma.

2) Methods

Multiple biopsies from melanoma patients were subjected to flow-sorting based on DNA content. The tumor specific marker SOX10 was used as an additional parameter to exclude contribution of normal tissue. Resulting clonal tumor populations were genomically characterized by usage of high resolution aCGH. A TMA was constructed for validation of the findings.

3) Results

DNA content based flow sorting of small human melanoma biopsies was successfully implemented and revealed the co-existence of distinct clonal populations (diploid as well as aneuploid) within these samples. The use of the additional marker SOX10 allowed us to uncover and sort pure diploid tumor populations from diploid fractions admixed with normal cells. These findings were confirmed by aCGH.

4) Conclusions

Human malignant melanoma is composed of different clonal populations with population-specific genomic aberrations. The use of SOX10 allowed unraveling of pure tumor populations within the diploid peaks, which would have been obscured by the use of DNA content alone. Further genomic analyses of these sorted clonal tumor populations are fundamental for the understanding of the genomic heterogeneity and its potential impact on metastasis and therapy response in malignant melanoma.

Citation Format: Thomas Lorber, Tanja Dietsche, Joël Gsponer, Alexander Rufle, Michael Barret, Lukas Sommer, Katharina Glatz, Christian Ruiz, Lukas Bubendorf. Deciphering the genomic heterogeneity in malignant melanoma by genomic profiling of clonal tumor populations. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3167. doi:10.1158/1538-7445.AM2013-3167
Abstract 3180: In vitro tool for discovering oncogenes and tumor suppressor genes in a system's manner.
Authors: Saginc Gaye , Ukil Leena , Woo Xingyi , Menghi Francesca , Bertrand Denis , Lee Charlie Wah Heng , Inaki Koichiro , Liu Edison Tak Bun
Published: August 2014

The molecular complexity of cancer still remains as the limiting factor for developing efficient therapies with minimal side effects. However, scientists succeeded in determining common alterations in most human cancer types. Each of these alterations results from disruption of an anti-cancer regulation mechanism and/or activation of cancer-promoting networks by genetic and epigenetic changes. As a consequence, cancer cells become capable of mimicking stem cells’ ability to proliferate indefinitely while maintaining their cellular identity, or to self-renew, which is crucial for any type of stem cell. Indeed, some of the networks which are deregulated in cancer normally regulate self-renewal in stem cells. However, self-renewal is tightly regulated in stem cells, while cancer cells manage to escape the internal and external regulation mechanisms. Hence, cancer cells differ from stem cells by displaying disruptions in feedback mechanism controlling the rate of cell division, aberrant differentiation programs and error-prone replication, so they become invasive and even metastatic. We hypothesize that the stemness regulatory “cassette” that serves limitless replicative potential and undifferentiated state is a measure of cancer virulence and progression. We believe that this gene cassette is established through different mechanisms in each cell type, so it can be separated from invasion and instability in a precise and defined manner. To test our hypotheses, we set up a system reconstruction model consisting of induced pluripotent cells and transformed cells as stem and cancer cell models respectively, and their source of primary mouse embryonic fibroblast cells. From this model system, we created a system's map for transformation using differential expression patterns from whole genome expression arrays. We discovered gene networks that are common between stem and cancer cells, but different from their primary cells of origin. We also found deregulated/activated cassettes which are unique for cancer cells and which are hardwired by genomic rearrangements (copy number and structural variations) identified by DNA paired end tag (DNA-PET) sequencing and single nucleotide variations using RNA sequencing. We are now investigating the role of candidate genes selected from this analysis in cancer and stem cell maintenance through a high content siRNA screening.

Citation Format: Gaye Saginc, Leena Ukil, Xingyi Woo, Francesca Menghi, Denis Bertrand, Charlie Wah Heng Lee, Koichiro Inaki, Edison Tak Bun Liu. In vitro tool for discovering oncogenes and tumor suppressor genes in a system's manner. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3180. doi:10.1158/1538-7445.AM2013-3180
Abstract 3193: NPV-LDE-225 (Erismodegib) inhibits human prostate cancer stem cell growth in NOD/SCID IL2Rγ null mice by regulating Bmi-1 and microRNA-128.
Authors: Nanta Rajesh , Shankar Sharmila , Srivastava Rakesh K.
Published: August 2014

Prostate cancer stem cells (CSCs) are defined by their extensive self-renewal, differentiation, and tumor initiation properties. It is now clear that CSCs are involved in tumor growth and recurrence, and resistance to conventional treatments. Thus, the strategy that suppresses stemness and consequently tumorigenic potential of CSCs could be considered for the management of prostate cancer. The objectives of this study were to examine the molecular mechanisms by which NPV-LDE-225 / Erismodegib (smoothened inhibitor) regulates stem cell characteristics in prostate cancer. Effects of NPV-LDE-225 on CSC's viability, sphere formation, apoptosis, transcriptional activity, and epithelial-mesenchymal transition (EMT) were measured. NPV-LDE-225 inhibited cell viability and spheroid formation, and induced apoptosis by activation of caspase-3 and cleavage of PARP. NPV-LDE-225 induced expression of Bax and Bak, and inhibited the expression of Bcl-2, Bcl-XL, XIAP, cIAP1, cIAP2 and survivin. NPV-LDE-225 inhibited Gli transcriptional activity, Gli-DNA interaction, and the expression of Gli1, Gli2, Patched1 and Patched 2 in prostate CSCs. Interestingly, NPV-LDE-225 induced PDCD4 and apoptosis and inhibited cell viability by suppressing miR-21. Furthermore, NPV-LDE-225 inhibited pluripotency maintaining factors Nanog, Oct4, cMyc and Sox-2. The inhibition of Bmi-1 by NPV-LDE-225 was regulated by up-regulation of miR-128. NPV-LDE-225 suppressed EMT by up-regulating E-cadherin and inhibiting N-cadherin, Snail, Slug and Zeb1 through regulating miR-200 family. Finally, NPV-LDE-225 inhibited CSC tumor growth which was associated with the suppression of Gli1, Gli2, Patched-1, Patched-2, Cyclin D1 and PCNA, and cleaved caspase-3 and PARP in tumor tissues derived from NOD/SCID IL2Rγ null mice. Overall, our findings suggest that inhibition of the Shh signaling pathway in CSCs is a potential therapeutic strategy for prostate cancer.

Citation Format: Rajesh Nanta, Sharmila Shankar, Rakesh K. Srivastava. NPV-LDE-225 (Erismodegib) inhibits human prostate cancer stem cell growth in NOD/SCID IL2Rγ null mice by regulating Bmi-1 and microRNA-128. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3193. doi:10.1158/1538-7445.AM2013-3193