bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 4847: CD95/Fas protects cancer cells from cell death
Authors: Hadji Abbas , Hau Annika , Peter Marcus E.
Published: August 2012

CD95 (also called Fas and APO-1) is a prototypical death receptor that regulates tissue homeostasis mainly in the immune system through the induction of apoptosis. During cancer progression CD95 is frequently downregulated or cells are rendered apoptosis resistant, raising the possibility that loss of CD95 is part of a mechanism for tumor evasion. However, complete loss of CD95 is rarely seen in human cancer and many cancer cells express large quantities of CD95 and are highly sensitive to CD95-mediated apoptosis in vitro. Furthermore, cancer patients frequently have raised levels of the physiological ligand for CD95, CD95L. These data raise the possibility that CD95 could actually promote the growth of tumors through its non-apoptotic activities. We have recently shown that cancer cells in general, regardless of their CD95 apoptosis sensitivity, depend on constitutive activity of CD95, stimulated by cancer-produced CD95L, for optimal growth (1). Consistently, loss of CD95 in mouse models of ovarian cancer and liver cancer reduces cancer incidence as well as the size of the tumors. We have now generated inducible CD95 and CD95L knock down cell lines to characterize the molecular mechanisms of the dependency of cancer cells on CD95. In all cancer cells knocking down CD95 or CD95L slows down growth and in many cells causes cell cycle arrest and even cell death. In CT26L cells knock down of CD95L causes all cells to die within 2 days. This form of cell death cannot be inhibited by inhibitors of apoptosis or necroptosis and has features of mitotic catastrophe. The data suggest that CD95 maintains viability of cancer cells by preventing DNA damage. 1. Chen et al (2010) CD95 promotes tumor growth. Nature, 465, 492-496.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4847. doi:1538-7445.AM2012-4847
Abstract 4848: A synthetic-lethal screen identifies novel Mcl1 regulators in neuroblastoma
Authors: Laetsch Theodore , Liu Xueyuan , Sliozberg Michael , Vido Michael , Vu Annette , Goldsmith Kelly , Hogarty Michael D.
Published: August 2012

Neuroblastoma (NB) is a lethal pediatric tumor. Despite multimodal therapy survival remains poor due to emergent chemoresistance governed by Bcl2-homology (BH) proteins. Agents that antagonize BH proteins, such as ABT-737, are in clinical development. ABT-737 does not antagonize Mcl1, a Bcl2-homologue correlated with ABT-737 resistance in many tumors. We defined the survival bias present in NBs using functional mitochondrial profiling in response to diverse BH3 death-domain peptides. Those with a functional Bcl2-dependence were exquisitely sensitive to ABT-737 in vitro and in vivo. Those with an Mcl1-dependence were relatively ABT-737 resistant (3-5 uM IC50s), yet Mcl1 knock-down restored ABT-737 activity in this subset (nM IC50s). We therefore undertook a synthetic-lethal screen for Mcl1 regulators using siRNA against deubiquitinating enzymes (DUBs), as Mcl1 is highly regulated with a short half-life due to ubiquitin-mediated degradation and this genomic space has many plausible druggable targets. We treated Mcl1-dependent NB cell lines (IMR5 and NLF) with siRNA targeting DUBs and ABT-737 at a sublethal concentration (200 nM). This screen identified 5 targets (of 98) demonstrating synthetic lethality (Z<-1.5) that was selective for Mcl1-dependent NBs. We validated these by measuring ABT-737 IC50s in vitro with and without siRNA knockdown and confirmed a >10-fold decrease in IC50 for 4 of 5 targets. Three further demonstrated a >10-fold decrease in ABT-737 IC50 with multiple independent siRNA supporting these as on-target effects (4/4 for PSMD14, 3/4 for PRPF8, and 2/4 for UBL5). We show that we can rescue the PSMD14 and UBL5 knock-down phenotypes by expression of an siRNA-resistant expression plasmid. While knock-down of all three targets decreases Mcl1 function, only knock-down of UBL5 (downregulates) and PSMD14 (upregulates) alter Mcl1 protein expression. We hypothesize that knock-down of PSMD14 and PRPF8 decrease Mcl1 activity through inhibitory post-translational modifications despite paradoxical or absent effects on Mcl1 protein expression, while UBL5 stabilizes Mcl1 as a ubiquitin-like PTM. Thus, we have demonstrated the robustness of a novel synthetic lethal screen to identify modulators of Mcl1 activity in vitro and identified three novel therapeutic targets regulating Mcl1 activity in NB.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4848. doi:1538-7445.AM2012-4848
Abstract 4909: Calcitriol and arsenic trioxide synergistically inhibited proliferation of HL-60 cell line mainly through apoptosis pathway
Authors: Bae Ji-Yeon , Kim Ji-Won , Kim Ji Yeon , Lim Yoojoo , Lee June Koo , Lee Eunyoung , Yoon Sung-Soo , Park Seonyang , Kim Byoung Kook , Kim Inho
Published: August 2012

Arsenic trioxide (As2O3) has shown substantial efficacy in the treatment of patients with acute promyelocytic leukemia (APL). However, since not all patients could achieve complete remission after As2O3 treatment, it is necessary to investigate a novel treatment method overcoming the resistance of current treatment. However, the therapeutic role of calcitriol against APL has not yet been fully elucidated. In our study, we investigated the anti-leukemic effect of calcitriol in combination with As2O3 on the human APL cell line HL-60. Inhibition of proliferation rate of HL-60 cells was measured by the trypan blue exclusion test was resulted in dose, time- dependent expressions. To evaluate the effects of combination treatment of As2O3 and calcitriol on the cell proliferation, we analyzed the combined effects of As2O3 and calcitriol using CalcuSyn software (BioSoft, Ferguson, MO, USA), which is based on the multiple drug-effect equation of Chou and Talalay (Chou TC, et al. Adv Enzyme Regul 1984): a combination index (CI) of <1, 1, or >1 is defined as synergistic, additive, or antagonistic, respectively. In all combinations, the CI values calculated were less than 1. To investigate the effects of these treatments on the apoptosis pathway, we analyzed the expression of Bax, Caspase-3, Bcl-2 with the Western blot and the RT-PCR. In the Western blot analysis, lower Bcl-2 and higher Bax and Caspase-3 expressions were observed in the As2O3 or calcitriol-treated groups compared to the control group. The combination treatment decreased Bcl-2 expression and increased Bax and Caspase-3 expressions more prominently than the single treatment using either As2O3 or calcitriol. In the RT-PCR analysis, these trends were validated again with the same results. These cells were then labeled with fluorescein-conjugated Annexin V-FITC and propidium iodide (PI), and analyzed with flow cytometry to differentiate whether the main cause of cell death was apoptosis or necrosis. To resultsze, calcitriol in combination with As2O3 synergistically inhibited proliferation of the human APL cell line HL-60. In addition, calcitriol combined with As2O3 more prominently activated the apoptosis pathway than single treatment using either calcitriol or As2O3. The main cause of cell death was also apoptosis. Our results suggest that these novel findings could be applied to improve the therapeutic outcomes of patients with APL.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4909. doi:1538-7445.AM2012-4909
Abstract 4912: Systematic, comparative network analysis on non-small cell lung cancer
Authors: Wong Serene , Kotlyar Max , Strumpf Dan , Cercone Nick , Shepherd Frances A. , Tsao Ming-Sound , Jurisica Igor
Published: August 2012

BACKGROUND: Most cancers lack any effective early disease markers, prognostic and predictive signatures. We fail treating cancer due to multiple ways cancer initiates and develops treatment resistance. While drug modes of action are complex and poorly understood, using Comparative Toxicogenomics Database is an effective way to identify drug combinations. Integrating signatures with deregulated network information may lead to identifying novel treatment option for individual patients. APPROACH: A systematic graph analysis was used to extract network structure differences between normal and tumor patient samples in non-small cell lung cancer. Three gene expression datasets with 27 squamous cell carcinoma, 129 adenocarcinoma, 20 large cell carcinoma and 141 normal samples, and 18 prognostic non-small cell lung cancer gene signatures were used to construct normal and tumor co-expression graphs. RESULTS: We enumerated all 5-node graphlets in normal and tumor graphs, and separated them into 3 categories: unique for normal graph, unique for tumor graph, or present in both. We further focused on subgraphs with the same membership across all 3 datasets and unique to tumor graph. Using gene enrichment analysis with a hypergeometric test we identified 9 subgraphs significantly enriched in the term “regulation of lymphocyte activation” (p<0.05), and genes related to chemokine receptors (CCR2, CCR7), interleukin (IL16), interleukin receptor (IL7R), interferon regulatory factor (IRF4), and T cells or B cells (PTPRCAP, SH2D1A, LCK, BTK, MS4A1). Importantly, this analysis identified protein interaction deregulated in tumors. Using the Comparative Toxicogenomics Database, we identified putative compounds that may “repair” the wiring of these subnetworks in tumor samples. 7 out of 7 identified compounds for the significantly up-regulated genes (p<0.05), and 20 out of 25 significantly down-regulated genes (p<0.05) are associated with non-small cell lung cancer. The other 5 identified compounds are known for other types of cancer or neoplasms, including lung neoplasms. Importantly, 13/38 edges have known/predicted protein interaction evidence, with a high prediction scores >0.9 (11 interactions) and 0.8 (2 interactions). CONCLUSIONS: Systematic integration and network analysis of non-small cell lung signatures identifies potential treatment options and insights to the difference in the underlying wiring related to immune system, an emerging hallmark of cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4912. doi:1538-7445.AM2012-4912
Abstract 4914: Integrated molecular profiles of primary breast tumors reveal differential interleukin signaling and improved prognostic power
Authors: Kristensen Vessela N.
Published: August 2012

The accumulation of high throughput molecular profiles of tumors at various levels has been a long and costly process worldwide. Combined analysis of gene regulation at various levels may point to specific biological functions and molecular pathways that are deregulated in multiple epithelial cancers and reveal novel subgroups of patients for tailored therapy and monitoring. We have collected high throughput data at several molecular levels derived from fresh frozen samples from primary tumors and matched blood from approximately 110 breast cancer patients (the MicMa dataset) with information about presence of disseminated tumor cells (DTC) and long term follow-up. The MicMa set has been used in parallel pilot studies of whole-genome mRNA expression (1), arrayCGH (2), DNA-methylation (3), whole genome SNP-CGH array (4), whole-genome miRNA expression analyses (5), TP53 mutation status dependent pathways (6) and high throughput paired-end sequencing (7). We have explored the extent to which these new profiles, acquired by molecular analyses at various levels recapitulate the initially discovered expression subclasses and their prognostic potential. Further, we attempted to integrate these by modeling mRNA, CNAs, miRNAs, and methylation in a pathway context using Paradigm (Pathway Recognition Algorithm using Data integration on Genomic Models) (8). We were able to show that combining mRNA expression and DNA copy-number leads to better discrimination of patients with respect to prognosis than any of the molecular levels studied separately. The pathways whose perturbations most strongly contributed to this classification were ERBB4, also previously found deregulated in breast and ovarian cancers, FoxM1 transcription, deregulated also in the ovarian and glioblastoma TCGA datasets, Angiopoientin receptor Tie2-mediated signaling and most notably the immune response (TCR) and interleukin signaling, IL4, IL6, IL12 and IL23 signaling. Massive interleukin signaling and similar sub-classification was observed also in DCIS (ductal carcinoma in situ) together with extracellular matrix and cell-cell adhesion regulating pathways. 1. Naume B et al. (2007) Mol. Oncol. 1, 160-171. 2. Russnes HG and Vollan HK et al. (2010) Sci. Transl. Med. 2, 38ra47. 3. Ronneberg J A et al. (2011) Mol. Oncol. 5, 61-76. 4. Van LP et al. (2010) Proc. Natl. Acad. Sci. U. S. A. 107, 16910-16915. 5. Enerly E and Steinfeld I et al. (2011) PLoS. One. 6, e16915. 6. Joshi H et al. submitted (2011). 7. Stephens P et al. (2009) Nature. 462, 1005-1010. 8. Kristensen V et al (2011)PNAS, 2001, Epub ahead

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4914. doi:1538-7445.AM2012-4914
Abstract 4918: Modeling polymersome delivery of anticancer therapeutics
Authors: Sorrell Ian , Webb Steven D. , Shipley Rebecca J. , Hearnden Vanessa L. , Colley Helen E. , Murdoch Craig
Published: August 2012

Polymersomes are synthetic nano-spheres that have the potential to specifically deliver anticancer therapeutics to tumor cells. They are an ideal delivery vector because of their nanometer size, structural architecture, stealth characteristics and the ability to attach tumor-specific ligands to their outer surface. The possible benefits of targeted polymersome drug delivery include reduced off-target toxic effects in healthy tissue and increased drug uptake by diseased tissue. In this study, in vitro experimentation is used to validate a mathematical description of polymersome-mediated therapy. Mathematical predictions are given for how the properties of polymersomes (size, membrane properties) and drug load affects rates of polymersome internalisation by tumor cells and therefore treatment efficacy. This allows for an optimisation of treatment design prior to clinical development. A key focus is polymersome-drug delivery via the clathrin-mediated endocytotic pathway. We include in our theoretical description specific details of this process (including specific binding ligands, receptor clustering, ligand tether length) and show how the characteristics of receptor targeting, number of ligands per polymersome, and polymersome concentration alters its uptake by tumor cells. The theoretical model is validated against in vitro studies using polymersome drug delivery to cell monolayers derived from different tumors. Next we describe how to scale-up the model to a tissue representation of the tumor mass. In this description the concentration of solutes (including oxygen and polymersomes) are tracked throughout the tumor using computational models based on realistic tumor vascular network structures with associated blood flow that have been constructed from confocal microscopic observations of a xenograft sub-cutaneously implanted tumor. Our results quantify the dependence of polymersome uptake rate and tumor distribution on polymersome properties and dosage. This multiscale approach identifies the key polymersome properties that influence treatment outcome and enables the treatment regime to be designed specifically in response to these features.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4918. doi:1538-7445.AM2012-4918
Abstract 4920: Gene network models identify targets for mammary tumor normalization
Authors: Brock Amy , Li Hu , Krause Silva , Kowalski Marek , Collins James J. , Ingber Donald E.
Published: August 2012

The majority of breast cancer therapeutics aim at slowing tumor expansion through selective killing of tumor cells. Given the enormous plasticity of cells (highlighted by recent advances in the iPS field), we instead sought to induce a cell state transition from a mammary tumor cell phenotype to a normal mammary epithelial cell state. We have tackled the critical problem of identifying target genes for the normalization or differentiation of mammary tumor cells by employing a systems biology approach that integrates computational modeling with in vivo and in vitro studies. In vivo gene expression data of mammary tumor progression in the FVB C3(1) SV40Tag transgenic mouse model were collected at multiple time points as the disease progressed from normal gland to DCIS-like lesions to invasive carcinoma. Gene regulatory network models predicted novel transcription factor targets that appeared critical to the rise of the tumor phenotype. These putative gene targets were then assessed in a 3D spheroid assay. RNAi inhibition of two computationally predicted transcription factors reverted mammary tumor cells to a normalized mammary epithelial cell phenotype, with appropriate apical/basal polarization, lumen formation and reduced proliferative index. This integrated strategy has pinpointed transcription factors for RNAi therapeutic targeting in vivo but importantly, the method is not limited to breast cancer and may be useful for many different tumor types. In summary, our major advance has been the demonstration that a gene regulatory network reverse engineering approach can identify genes that are causatively involved in cancer progression and that represent viable therapeutic differentiation targets.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4920. doi:1538-7445.AM2012-4920
Abstract 4944: Crosstalk between NFκB and STAT3 in human multiple myeloma cells (U266)
Authors: Chudasama Vaishali L. , Mager Donald E.
Published: August 2012

Purpose: To evaluate a potential interaction between NFκB and STAT3 pathways and its consequences on treatment outcome (cell survival) in a human myeloma cell line (U266). Methods: A computational network model of signal transduction in U266 cells was used to simulate the interplay between NFκB and STAT3, suggesting that inhibition of just one of the pathways does not result in inhibition of NFκB and downstream processes, as NFκB expression is partially regulated by STAT3. To test this hypothesis, U266 cells were exposed continuously to either an IKK inhibitor (IKKi; 10 μM), a JAK inhibitor (JAKi; 10 μM), or bortezomib (Bort) (a proteasome inhibitor; 20 nM). The time-course of intracellular proteins were measured over 48 h (immunobloting), including: phospho-p65 (p-p65), p-stat3, p-IκBα, and BclXL. Cell proliferation was measured using WST-1 reagent assay kit at 72 h. PARP cleavage was measured as an indicator of apoptosis over a 24 h period. U266 cells were also exposed transiently (1 and 2 h) to Bort treatment, and the time-course of p-p65, BclXL, and PARP was measured. Results: Expression of p-p65 was unchanged following continuous exposure to IKKi and JAKi. However, p-p65 expression initially increased (1.7-fold) followed by a decrease below baseline following Bort exposure. BclXL expression remained unchanged after IKKi exposure, and it decreased (80 %) either rapidly or gradually after JAKi or Bort exposure. p-stat3 expression rapidly decreased after Bort (80 %) and was transiently increased after IKKi treatment (2 fold). For short-term Bort exposure, p-p65 expression transiently increased (2.5 fold) and then returned to baseline. BclXL expression decreased (50 %) and achieved a new steady-state baseline. Bort exposure for 2 h resulted in significant PARP cleavage (10 fold increase) in agreement with model simulations. Conclusions: Both NFκB and STAT3 pathways need to be inhibited to fully inhibit NFκB activity. Similarly, when NFκB and STAT3 are constitutively active, both transcription factors need to be inhibited to inhibit cell proliferation. Further research is needed to better understand mechanisms of p-stat3 inhibition and p-p65 transient stimulation and long-term inhibition following Bort exposure to U266 myeloma cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4944. doi:1538-7445.AM2012-4944
Abstract 4962: TLE1 is an anoikis regulator and is downregulated by Bit1 in breast cancer cells
Authors: Biliran Hector R. , Jennings Scott , Ireland Shubha Skale , Brunquell Chris , Chen Renwei , Ruoslahti Erkki
Published: August 2012

TLE1 is a Groucho related transcriptional repressor protein that exerts survival and anti-apoptotic function in several cellular systems and has been implicated in the pathogenesis of cancer. In the present study, we found that TLE1 is a critical regulator of anoikis in normal mammary epithelial and breast carcinoma cells. The induction of apoptosis following loss of cell attachment to the extracellular matrix (ECM) (anoikis) in untransformed mammary epithelial MCF10A cells was associated significant downregulation of TLE1 expression while forced expression of exogenous TLE1 in these cells promoted resistance to anoikis. In stark contrast, TLE1 expression was significantly upregulated following detachment from the ECM in malignant and anoikis resistant breast cancer cell lines, MDA-MB-231 and MCF7. Importantly, enforced suppression of TLE1 in these cells resulted in attenuation of anoikis resistance and decreased anchorage-independent growth with no impact on cell viability in adherent conditions. Consistently, enforced ectopic TLE1 expression further enhanced the anoikis resistance and anchorage-independent growth of MDA-MB-231 cells. The pro-survival and anti-apoptotic role of TLE1 during loss of cell attachment is in part attributed to its ability to inhibit the caspase-independent anoikis function of Bit1. Interestingly, the TLE1 protein is channelled to proteosomal degradation by Bit1 during anoikis. Taken together, these findings indicate a novel role for TLE1 in the maintenance of anoikis resistance in breast cancer cells. This conclusion is supported by an immunohistochemical analysis of a breast cancer tissue array illustrating that TLE1 is selectively upregulated in invasive breast tumors relative to noninvasive ductal carcinoma in situ (DCIS) and normal mammary epithelial tissues.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4962. doi:1538-7445.AM2012-4962
Abstract 4963: Tanshinone IIA induces JNK-dependent intrinsic apoptosis via suppressing astrocyte-elevated gene 1 expression in chronic myelogenous leukemia KBM-5 cells
Authors: Kim Ji-Hyun , Jeong Soo-Jin , Yun Sun-Mi , Jung Ji Hoon , Lee Hyo-Jung , Lee Hyo-Jeong , Lee Eun-Ok , Lee Min-Ho , Kim Sung-Hoon
Published: August 2012

Tanshinone IIA (Tan IIA) is a phytochemical isolated from the root of Salviae miltiorrhizae (Danshen) and has been reported to have various biological activities such as anti-inflammation, anti-oxidation and anti-cancer. However, the underlying molecular mechanisms to control anti-cancer effect of Tan IIA in chronic myelogenous leukemia (CML) have not been fully understood yet. In the present study, we investigated whether Tan IIA has the anti-cancer potential by inducing apoptosis in CML cells. Tan IIA significantly decreased the viability of human CML KBM-5 cells, but not normal peripheral blood lymphocytes. Tan IIA induced apoptotic cell death by increasing the sub-G1 cell populations and stimulating atypical shrunken of nucleus and DNA fragmentation. Also, Tan IIA significantly reduced the mitochondrial membrane potential, stimulated the release of cytochrome C from mitochondria into the cytosol and activated intrinsic caspase cascade. Furthermore, Tan IIA enhanced phosphorylation of JNK and suppressed the expression of astrocyte-elevated gene-1 (AEG-1). In contrast, blocking JNK activity using JNK inhibitor SP600125 reversed Tan IIA-induced apoptosis and AEG-1 expression. Taken together, our findings suggest that Tan IIA induces JNK-dependent intrinsic apoptosis via suppressing AEG-1 expression in KBM-5 cells as a valuable candidate agent for CML therapy.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4963. doi:1538-7445.AM2012-4963
Abstract 4968: Phosphorylation of caspase-8 (Thr263) by ribosomal S6 kinase 2 (RSK2) mediates caspase-8 ubiquitination and stability
Authors: Peng Cong , Cho Yong-Yeon , Zhu Feng , Li Haitao , Li Xiang , Xie Hua , Bode Ann M. , Dong Zigang
Published: August 2012

The ribosomal S6 kinase 2 (RSK2) is a member of the p90 ribosomal S6 kinase (p90RSK) family of proteins and plays a critical role in proliferation, cell cycle and transformation. Here, we report that RSK2 phosphorylates caspase-8 and Thr263 was identified as a novel caspase-8 phosphorylation site. Phosphorylation of Thr263 is associated with the ubiquitination and degradation of caspase through the proteasome pathway. Results suggested that RSK2 regulates caspase-8 stability through its phosphorylation of this caspase. These data provide a direct link between RSK2 and caspase-8 and identify a novel molecular mechanism for caspase-8 modulation by RSK2.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4968. doi:1538-7445.AM2012-4968
Abstract 497: Arousal of the stroma: Fibroblasts lead pancreatic cancer cell invasion
Authors: Brentnall Teresa A. , Lai Lisa , Bronner Mary , Chen Ru
Published: August 2012

Background: Cancer-associated fibroblasts, comprised of activated fibroblasts or myofibroblasts, are found in the stroma surrounding solid tumors; these myofibroblasts are thought to promote invasion and metastasis of cancer cells. Mechanisms regulating the activation of the fibroblasts and the initiation of the invasive tumorigenesis are of great interest. Upregulation of the cytoskeletal protein, palladin, has been detected in the stromal myofibroblasts surrounding many solid cancers and in expression screens for genes involved in invasion. Using a pancreatic cancer model, we investigated the functional consequence of overexpression of exogenous palladin in normal fibroblasts in vitro and the affect it could have on the early stages of tumor invasion. Findings: Palladin expression in cancer-associated fibroblasts occurs very early in tumorigenesis. In vivo, concordant expression of palladin and the myofibroblast marker, alpha smooth muscle actin (α-SMA), occurs early at the dysplastic stages in peri-tumoral stroma and progressively increases in pancreatic tumorigenesis. In vitro introduction of exogenous 90kD palladin into normal human dermal fibroblasts (HDFs) induces activation of stromal fibroblasts into myofibroblasts as marked by induction of α-SMA and vimentin, and through the physical change of cell morphology. Moreover, palladin expression in the fibroblasts enhances cellular migration, invasion through the extracellular matrix, and creation of tunnels through which cancer cells can follow. The fibroblast-led invasion and creation of tunnels results from the development of invadopodia-like cellular protrusions (“feet”) which express invadopodia proteins and proteolytic enzymes. Palladin expression in fibroblasts is triggered by the co-culture of normal fibroblasts with k-ras-expressing epithelial cells. Abrogation of palladin with shRNA reverses the ability of cancer cells to invade by blocking the transition of fibroblasts into myofibroblasts and dampening of the myofibroblast-cancer cell partnership. Conclusions: Fibroblasts are active partners in pancreatic cancer cell invasion. Palladin expression can impart myofibroblast properties, in turn promoting the invasive potential of these peri-tumoral cells with invadopodia-driven degradation of extracellular matrix. Palladin expression in fibroblasts can be triggered by k-ras expression in adjacent epithelial cells. This data supports a model whereby palladin-activated fibroblasts facilitate stromal-dependent metastasis and outgrowth of tumorigenic epithelium.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 497. doi:1538-7445.AM2012-497
Abstract 5041: Characterization of the miR-125b1 promoter in breast cancer cell lines
Authors: Soberanis Fernanda Cisneros , Reyes Ernesto Soto , Castro Clementina , Targa Felix Recillas , Herrera Luis A.
Published: August 2012

In cancer cells, transcriptional gene silencing has been associated with genetic and epigenetic defects. The disruption of DNA methylation patterns and covalent histone marks has been associated with cancer development. Until recently, microRNA (miRNA) gene silencing was not well understood. In particular, miR-125b1 has been suggested to be a miRNA with tumor suppressor activity, and it has been shown to be deregulated in various human cancers. In this study, we characterized the promoter of the miR-125b1 and the modifications associated with gene silencing. We studied in silico the miR-125b1 locus to delimit the promoter region and then, we characterized the promoter activity by the luciferase assay, cloning a fragment in the 5′ extreme close to the transcriptional start site of the miR-125b1 gene. We found that this sequence has promoter activity and it is unidirectional. Subsequently, we analyzed the DNA methylation status in the CpG island promoter and found that it was methylated in breast cancer cell lines compared with a non-transformed breast cell line. To determine the effect of DNA methylation in the CpG island of miR-125b1 on the expression of this gene, we performed a qRT-PCR assay. We observed a significant reduction on the expression of miR-125b1 in cancer cells lines in comparison with a non-transformed cell line, suggesting that DNA methylation at the CpG island might reduce the expression of miR-125b1. Our data suggest that the fragment in the 5′ extreme close to the transcriptional start site of the miR-125b1 gene is a functional and unidirectional promoter. Also, the CpG island in this region is methylated in breast cancer cell lines and this methylation is associated with the silencing of the miR-125b1 gene. This work was supported by the Consejo Nacional de Ciencia y Tecnología (CONACyT: 83959) and the Programa de Apoyo a Proyectos de Investigación e Innovación Tecnológica of the Universidad Nacional Autónoma de México (PAPIIT, IN213311).

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5041. doi:1538-7445.AM2012-5041
Abstract 5049: Serum miRNAs as an early marker for breast cancer
Authors: Godfrey Ashley C. , Xu Zongli , Weinberg Clarice R. , Wade Paul A. , Deroo Lisa A. , Sandler Dale P. , Taylor Jack A.
Published: August 2012

MicroRNAs (miRNAs) are small, non-coding, single-stranded RNAs between 18-22 nucleotides that regulate gene expression by affecting mRNA translation and degradation. It has been proposed that miRNAs could be master regulators of gene expression by regulating entire gene networks. Expression of miRNAs is altered in tumor compared to normal tissue and there is growing evidence that miRNA profiles in serum may differ in cancer cases as compared to healthy controls. However, these studies use samples collected after diagnosis so treatment and/or disease effects cannot be excluded. We are using serum samples from the Sister Study, a prospective cohort study of 50,000 women who had never had breast cancer at the time of enrollment, but were at increased risk because they had a sister previously diagnosed with breast cancer. We are exploring whether serum miRNA profiles can be used as prediagnostic markers of breast cancer. In an initial pilot analysis comparing 5 women diagnosed with breast cancer 1-2 months after their blood draw to 5 controls, there were 25 miRNAs significantly different at the 0.05 level (paired T test). Of those 25 miRNAs, 7 have been previously associated with breast tumor tissue in published studies showing differential expression in tumor tissue or in breast cancer cell lines. We have now completed a larger follow up study examining 205 women diagnosed with breast cancer within 18 months of their blood draw compared to 205 controls frequency matched to cases by age, 2 month interval of blood draw and race. We are using Affymetrix miRNA microarrays to profile miRNA expression in serum samples, and have optimized this platform for use with less than .5mLs of serum. After normalization and background correction, we found 439 miRNAs (of 1105 human miRNAs on the chip) expressed above background in at least 50 samples. When the average expression of controls was compared to cases using logistic regression, we found 16 miRNAs significantly different at the .05 level. For cases, the mean time between blood draw and cancer diagnosis was 10 months. We hypothesize that, if miRNA is a prediagnostic marker, cases with a longer time between blood draw and diagnosis would have a more control-like miRNA profile. To examine this, we are conducting a more detailed analysis among cases with a short time between blood draw and diagnosis. The next steps of our analysis will be to examine how far in advance of cancer diagnosis can we detect case-control differences in miRNA expression profiles.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5049. doi:1538-7445.AM2012-5049
Abstract 505: Jagged 2 knockdown inhibits invasion in colorectal cancer cell lines
Authors: He Wan , Wong SC Cesar , Chan Charles ML , Ho WS , Au Thomas CC , Lam Money YY , Chan Anthony TC
Published: August 2012

Background: The notch pathway is involved in control of cell differentiation in various tissues including pancreas, muscle and bone marrow. Jagged 2 (JAG2) is a ligand for the notch pathway and was found to be overexpressed in malignant plasma cells from patients with multiple myeloma, causing the secretion and release of IL-6, VEGF, and IGF-1 in the microenvironment supporting myeloma cell growth. In breast cancers, it was found to be overexpressed at the hypoxic invasive front and was significantly correlated with metastasis free survival. Furthermore, up-regulation of JAG2 expression in mouse lung adenocarcinoma cells promoted its metastasis in mice. However, the expression pattern and function of JAG2 in colorectal cancer (CRC) is not clear. Therefore in this study, we examined the expression of JAG2 in CRC cell lines and determined the effect(s) of JAG2 knockdown by RNA interference in CRC cell lines. Materials and Methods: The expression of JAG2 protein was examined in four CRC cell lines (SW480, SW620, HCT116, DLD-1) and also in 1 normal colon epithelial cell line (CCD18co) using the western blot. To investigate the effects JAG2 knockdown in CRC cell lines, a pool of 4 siRNAs against JAG2 was transfected into the 4 CRC cell lines listed above. The effects on cell growth was then determined by the CellTiter 96® AQueous One Solution Cell Proliferation Assay (the MTS assay) and the Matrigel invasion assay was performed to assess its effects on invasive capability. Results: Western blot results showed that JAG2 protein was expressed in all 4 colorectal cancer cell lines including SW480, SW620, HCT116 and DLD-1 but not in the CCD18co normal colon epithelial cell line. The MTS assay showed that cell growth was not affected by JAG2 knockdown in the CRC cell lines. Interestingly, the invasive capability of HCT116 and DLD-1cells was reduced after JAG2 knockdown. Conclusions: Results from this study suggest that JAG2 may play a role in the invasiveness of CRC cell lines but not in cell proliferation. JAG2 may be a promising therapeutic target for inhibition of metastasis in colorectal cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 505. doi:1538-7445.AM2012-505
Abstract 5059: Exome and targeted sequencing to discover and validate candidate genes in advanced and lethal prostate cancer
Authors: Kumar Akash , MacKenzie Alexandra P. , O'Roak Brian J. , Coleman Ilsa , Morrissey Colm , Lee Choli , White Thomas A. , Dumpit Ruth F. , True Lawrence D. , Corey Eva , Vessella Robert L. , Nelson Peter S. , Shendure Jay
Published: August 2012

Prostate carcinoma is the most common malignancy and the second leading cause of cancer mortality in men. In order to understand the genetic composition of lethal and advanced prostate cancer, we used exome sequencing to identify protein-altering mutations within a panel of 23 prostate cancer xenografts that represent a wide spectrum of advanced disease phenotypes. Although corresponding normal tissue was not available for most tumors, we were able to take advantage of increasingly deep catalogs of human genetic variation to remove most germline variants. On average, each tumor genome contains ∼200 novel nonsynonymous variants, of which the vast majority was specific to individual carcinomas. Based on the prevalence of rare nonsynonymous variants, we identified a subset of 30 genes including TP53, DLK2, GPC6 and SDF4 as putative candidates involved in tumor progression. To ascertain the frequency of mutations in these candidate genes across a large panel of tumors, we developed a molecular inversion probe (MIP) based protocol that enables efficient targeting and sequencing of tens of genes across hundreds to thousands of samples. The method requires as little as 50 nanograms of DNA as input, and is compatible with DNA isolated from both frozen and formalin formalin-fixed paraffin embedded (FFPE) tissues. We have applied this method to capture and deeply sequence the nearly ∼1,000 exons of the 30 candidate genes across 150 prostate cancer metastases on a single lane of an Illumina HiSeq2000 platform. Unexpectedly, our exome survey identified three tumors with substantially higher mutation frequencies, with 2,000-4,000 novel coding variants per exome. We used whole genome sequencing to characterize the global patterns of mutation in one of these tumors, and found a high prevalence of transition mutations within CpG islands. Experiments to determine whether hypermutation is the result of loss of DNA repair pathway function are ongoing. Collectively, our results indicate that exome sequencing combined with efficient screening of candidate genes is a powerful method to understand the genetics of advanced prostate cancer. We also report a previously undescribed subtype of prostate cancers exhibiting “hypermutated” genomes, with potential implications for resistance to cancer therapeutics.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5059. doi:1538-7445.AM2012-5059
Abstract 5062: Integrated analysis of the whole exome sequencing (WES) identifies novel mutations and suggests patient selection hypothesis
Authors: Lezhnin Sergey
Published: August 2012

Whole exome sequencing was done on 257 breast and 102 ovarian tumors using next generation sequencing. Seven gigabases of sequencing data were generated for each sample or 100x coverage of the whole exome, which enable to determine sequence variations with >99% confidence. The results of the mutation calls were compared with recently published TCGA breast and ovarian datasets. Genes with large number of passenger mutations were called out and frequency of passenger mutations per sample was linked to MMR signature. In ovarian tumors, a high prevalence of p53 and BRCA1 mutation was confirmed in serous tumors. A signature of BRCA1/2 deficient tumors was developed and cross-validated between the datasets. In no-serous tumors, a high prevalence of mutations MAPK pathway and beta-catenin pathway were found. FOXL2 mutation was identified in 100% of granulose ovarian tumors and linked to high levels of EMT signature. In addition we showed a significant overlap between genes found to be mutated in TCGA datasets and in our dataset, suggesting a larger number of potentially significantly mutated genes than previously reported. In breast tumors, known high frequency of mutations was confirmed in PIK3CA and TP53 genes and validated with respective pathway specific gene expression signatures. In addition, prevalent mutations were identified in MAPK and PI3K pathways and linked with respective pathway signatures. These findings strengthen and enrich our understanding of molecular subtypes of ovarian and breast tumors and can be used for identification of novel targets and for development of responder hypothesis.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5062. doi:1538-7445.AM2012-5062
Abstract 5075: Genome-wide analysis of DNA copy number variations in osteosarcoma
Authors: Gokgoz Nalan , Wunder Jay S. , Andrulis Irene L.
Published: August 2012

Objectives: Copy number variations (CNVs) such as deletions, amplifications and duplications across the whole genome may contribute to Osteosarcoma (OS) tumorigenesis. Detection of genomic regions containing these changes is extremely important for both the basic understanding of OS and its diagnosis. In this study we performed Whole Genome Genotyping (WGG) using high-density SNP arrays to detect CNVs in OS tumors and blood. A major advantage of this technology is the simultaneous detection of both allelic frequencies and CNVs. Methods: WGG analysis was performed using 35 high-grade intramedullary OS and 8 parosteal sarcoma tumor DNAs. 10 matching germline DNAs from blood were also included into the analysis. 250 nanograms of DNA were hybridized onto the Illumina 610 Quad BeadChips using the Infinium II WGG assay. The WGG data was analysed using Bead Studio and Partek software. Results: SNP array based Whole Genome Genotyping revealed a large number of chromosomal aberrations in OS. Preliminary analysis of 35 OS tumor DNAs showed high levels of recurrent amplification of 1p36, 3p12.1, 6q14, 12q13-15, 17p11.2, 20p12; and deletion of 3q26.31, 6p21, 6q16.3, 7q21, 7q35, 8q24.13, 9p21 and 17p13 regions. Parosteal tumor DNAs displayed a high level of amplification of the 12q13-15 amplicon. Amplification status of genes from 17p11.2 (PMP22, NCOR1, COPS3 and TOM1L2) and 12q14 (CDK4 and MDM2) was validated by Real-Time PCR. Some tumors exhibited whole chromosome arm deletions and/or duplications. Conclusions: OS tumors exhibit chromosomal alterations that can be detected using WGG. Comparison of normal and tumour DNAs from the same subjects can be used to identify CNV associated with OS development and progression. Current analyses include comparisons of CNV data and gene expression profiles of the same OS tumours to discover genes that may be biologically relevant targets in OS.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5075. doi:1538-7445.AM2012-5075
Abstract 5078: Identification of novel gene fusions by RNA-seq in gastric cancer
Authors: Cho Geun-A , Kim Hwang-Phill , Shin Jong-Yeon , Lee Won-Cheol , Yoon Young-Kwang , Han Sae-Won , Kim Jong-Il , Seo Jeong-Sun , Kim Tae-You
Published: August 2012

The recent discoveries of gene fusions in prostate cancer, lung cancer and gastric cancer suggested that gene fusion could be recurrent and tumorigenic also in carcinoma not only in haematological malignancies. Here, we have found 19 novel fusion genes and fusion transcripts in 12 gastric cancer cell lines. Using the paired-end RNA-seq approach, we obtained a list of 282 fusion genes and fusion transcripts candidates in 12 gastric cancer cell lines. Among these 282 candidates, we selected 25 candidates to examine: 22 candidates whose number of junction read was greater than 10 and 3 additional candidates whose loci are closely related to 22 previously selected candidates. We validated the candidates via Sanger sequencing. As a result, 19 out of 25 candidates were validated: 7 inter-chromosomal fusions and 12 intra-chromosomal fusions including 6 read-through events. We also performed long-range PCR to confirm genomic rearrangements, through which six candidates did not show any evidence of DNA rearrangements. These six candidates were assumed to be fusion transcripts without DNA-level rearrangements while the others are fusion genes that result from chromosomal rearrangements. Furthermore, we investigated the frequencies of all the 19 validated candidates in normal and tumour tissue pairs of 17 gastric cancer patients to determine whether these fusions are recurrent in other tumours and exist in actual cancer patients. Two inter-chromosomal fusion genes, CCDC67-VAV2 and PVT1-ATE1 were detected only in tumour tissues but in low frequencies: 2/17(11.8%) and 1/17(5.9%) respectively. BCAP29-DUS4L, a read-through event, was detected in 12 tumour tissues, 10 of which were only observed in tumour tissues not in the matched normal pairs. Taken together, we have found 13 fusion genes and 6 fusion transcripts in 12 gastric cancer cell lines, some of which are tumour-specific or highly recurrent in tumour tissues. We believe this study might contribute to find oncogenic fusion genes which can be potential biomarkers or therapeutic targets.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5078. doi:1538-7445.AM2012-5078