bluuebunny/crossref_metadata_2025_split huggingface dataset:
Abstract B20: Mouse models of Ewing's sarcoma and fusion-related sarcoma: Correct phenotypes and cell-of-origin
Authors: Tanaka Miwa , Nakamura Takuro 
Published: May 2015

Abstract
Tumor-specific fusion genes associated with chromosomal translocations play an important role in development of bone and soft tissue sarcoma. Clarification of tumorigenic processes by fusion genes is valuable to understand disease mechanisms and to develop molecular targeted therapies. We have recently established ex vivo mouse models for Ewing's sarcoma (ES) by introducing EWS-FLI1 into mouse embryonic cells. The ES model enabled us to analyze tumorigenic processes from the initial step to the highly malignant stage, and to select important genetic pathways in the processes. The knockdown of the EWS-FLI1 target genes and members of the important signaling pathways significantly suppressed cell proliferation of ES in vitro. Importance of such pathways were further confirmed by using specific inhibitors for PARP, EZH2 or Wnt/β-catenin pathway both in vitro and in vivo. Moreover, Cre-loxp-mediated knockout of EWS-FLI1 showed that mouse ES growth is fusion gene-dependent. In addition, we have succeeded to generate synovial sarcoma, alveolar soft part sarcoma and Ewing-like sarcoma by introducing SYT-SSX1, ASPL-TFE3 or CIC-DUX4, respectively. These results indicate essential roles of fusion oncogenes in sarcomagenesis and tumor maintenance, and the epigenetic status of cell-of-origin. Our platform will allow us to explore and evaluate novel targeted therapies.
Citation Format: Miwa Tanaka, Takuro Nakamura. Mouse models of Ewing's sarcoma and fusion-related sarcoma: Correct phenotypes and cell-of-origin. [abstract]. In: Proceedings of the AACR Special Conference: The Translational Impact of Model Organisms in Cancer; Nov 5-8, 2013; San Diego, CA. Philadelphia (PA): AACR; Mol Cancer Res 2014;12(11 Suppl):Abstract nr B20.
Abstract IA21: Tumor heterogeneity and cell-of-origin of mouse small cell and non-small cell lung cancer
Authors: Berns Anton 
Published: May 2015

Abstract
Small cell lung cancer (SCLC) is one of the most lethal human malignancies, due to its high metastatic potential and chemo-resistance upon relapse. Using the Rbf/f;p53f/f mouse model for SCLC, we found that the tumors are often composed of phenotypically different cells, characterized by mesenchymal and neuroendocrine markers. These cells often share a common origin. Crosstalk between these cells can endow the neuroendocrine component with metastatic capacity, illustrating the potential relevance of tumor cell heterogeneity in dictating functional tumor properties. Also specific genetic lesions appear to be associated with metastatic potential.
Interestingly, the two cell types can also interconvert raising the question of their cell-of-origin. To investigate this in more detail, we inactivated Trp53 and Rb1 in distinct cell types in the adult lung by targeting Cre-recombinase expression to Clara, neuroendocrine, and alveolar type II cells using adenoviral vectors. We could show that neuroendocrine cells serve as the predominant although not the exclusive cell of origin of SCLC.
In contrast, mutant Kras-driven adenocarcinomas (one of the NSCLC subtypes) originates primarily from alveolar type II cells. However, in LSL-mutant-Kras;p53flox/flox mice also other cell types gave effectively rise to adenomas and adenocarcinomas. Our data indicate that both cell type specific features and the nature of the oncogenic lesion(s) are critical factors in determining the tumor initiating capacity of lung (progenitor) cells. Furthermore, the cell-of-origin appears to influence the malignant properties of the resulting tumors.
Citation Format: Anton Berns. Tumor heterogeneity and cell-of-origin of mouse small cell and non-small cell lung cancer. [abstract]. In: Proceedings of the AACR Special Conference: The Translational Impact of Model Organisms in Cancer; Nov 5-8, 2013; San Diego, CA. Philadelphia (PA): AACR; Mol Cancer Res 2014;12(11 Suppl):Abstract nr IA21.
Abstract PR04: Linking development and cancer modeling in zebrafish identifies putative therapeutic targets in medullary thyroid cancer
Authors: Villefranc Jacques A. , Alazar Aida , Houvras Yariv 
Published: May 2015

Abstract
Medullary thyroid cancer (MTC) is a neuroendocrine cancer that arises from malignant transformation of calcitonin-producing C-cells. A majority of MTC is associated with somatically acquired mutations in the proto-oncogene Rearranged during transfection (RET), resulting in constitutive activation of downstream effectors of the ERK and AKT signaling pathways. While surgical treatment may be curative in patients with localized MTC, a significant fraction of patients harbor residual disease and ultimately relapse. Much of the research on MTC has converged on inhibition of RET, however, studies suggest that other signaling pathways may contribute to disease. As a result, there is a growing need to identify novel targets for MTC.
The zebrafish is well regarded as a model to study of development due its genetic tractability, conservation of pathways and rapid organ development; characteristics that also make it a fitting model to study human cancers. In this study we take advantage of these traits to 1) perform an in vivo chemical screen to identify modulators of C-cell development in zebrafish and 2) develop an in vivo oncogenic RET model in zebrafish to test the putative ‘hits’ from our screen. We hypothesize that the nexus of development and cancer modeling in the zebrafish will lead to therapies of potential use in treating MTC. We screened compounds modulating pathways involved in development, kinase signaling and chromatin modification for their ability to influence C-cell development and found that separate inhibition of the FGF and GSKβ; signaling pathways results in marked down-regulation of markers associated with C-cell differentiation. In parallel, we have introduced human oncogenic RET into an analogous cell type of neural crest origin in zebrafish which results in their early transformation, and are testing the ability of the identified ‘hits’ to suppress transformation of these cells. Our results implicate the FGF and GSKβ; pathways in C-cell development and encourage further investigation of their potential as therapeutic targets for MTC. Moreover, our studies highlight the use of zebrafish as model to identify novel targeted therapies in cancer.
This abstract is also presented as Poster B44.
Citation Format: Jacques A. Villefranc, Aida Alazar, Yariv Houvras. Linking development and cancer modeling in zebrafish identifies putative therapeutic targets in medullary thyroid cancer. [abstract]. In: Proceedings of the AACR Special Conference: The Translational Impact of Model Organisms in Cancer; Nov 5-8, 2013; San Diego, CA. Philadelphia (PA): AACR; Mol Cancer Res 2014;12(11 Suppl):Abstract nr PR04.
Abstract PR07: RNAi mouse models: Revolutionizing drug discovery in vivo
Authors: Premsrirut Prem , Dow Lukas , Hannon Gregory , Zuber Johannes , Lowe Scott , Zender Lars , Fellmann Christof 
Published: May 2015

Abstract
RNA interference is a powerful tool for studying gene function, however, the reproducible generation of RNAi tools including RNAi transgenic mice remains a significant limitation. One main hurdle is the identification of potent RNAi triggers, or short hairpin RNAs (shRNAs), that will induce stable and regulated gene silencing. Due to the lack of understanding of the requirements for shRNA biogenesis and target suppression, many predicted shRNAs fail to efficiently induce gene suppression. We have developed a “Sensor assay” that enables the biological identification of effective shRNAs at large scale and show that our assay reliably identifies potent shRNAs that are surprisingly rare and predominantly missed by existing algorithms. In addition, we have engineered a new miRNA scaffold, miR-E, that is more efficiently processed and thus produces more potent knockdown of target genes than our previous miR30 system. By combining our sensored miR-E based shRNAs with high efficiency ES cell targeting, we have developed a fast, scalable pipeline for the production of shRNA transgenic mice with reversible gene silencing. We show that RNAi can cause sufficient knockdown to recapitulate the phenotypes of knockout mice, particularly in cancer models. More importantly, unlike traditional knockout models, RNAi has the powerful advantage of reversibility, since the endogenous gene remains intact. Using this system, we generated a number of inducible RNAi transgenic lines and demonstrate how this approach can identify predicted phenotypes and also unknown functions for well-studied genes. In addition, through regulated gene silencing we are able to mimic drug therapy in mice without the actual drug molecule, allowing us to determine the therapeutic value and/or toxic effects associated with systemic gene suppression. Using this approach, we have been able to pinpoint potential toxicities associated with gene inhibition, results that will guide drug development to avoid target failures which will likely cause harmful and intolerable effects in patients. In a model of hepatocellular carcinoma, we demonstrate that short-term inhibition of a ribosomal protein is sufficient to induce stable cell cycle arrest of liver tumor cells. This system provides a cost-effective and scalable platform for the production of RNAi transgenic mice targeting any mammalian gene – mice with enormous predictive power that will shape our development of better tolerated therapies.
This abstract is also presented as Poster B30.
Citation Format: Prem Premsrirut, Lukas Dow, Gregory Hannon, Johannes Zuber, Scott Lowe, Lars Zender, Christof Fellmann. RNAi mouse models: Revolutionizing drug discovery in vivo. [abstract]. In: Proceedings of the AACR Special Conference: The Translational Impact of Model Organisms in Cancer; Nov 5-8, 2013; San Diego, CA. Philadelphia (PA): AACR; Mol Cancer Res 2014;12(11 Suppl):Abstract nr PR07.
Abstract A06: NORE1A is a double-barreled Ras senescence effector linking Ras to p53 and Rb
Authors: Donninger Howard , Calvisi Diego , Barnoud Thibaut , Schmidt M. Lee , Clark Geoffrey J. 
Published: May 2015

Abstract
In addition to driving growth and transformation, activated forms of Ras are potent inducers of oncogene induced senescence (OIS). OIS appears to be a major barrier that must be overcome to permit Ras driven tumorigenesis. The signaling pathways utilized by Ras to induce senescence, and how they are subverted during tumor development remain poorly characterized.
NORE1A is a member of the RASSF family of tumor suppressors. It binds directly to activated Ras via the Ras effector domain and acts as a Ras death effector. Frequent loss of NORE1A expression is observed in many tumor types and hereditary genetic defects in NORE1A predispose carriers to cancer. We have found that NORE1A can connect Ras to the induction of p21CIP1 and cell cycle arrest. Therefore, we sought to determine if NORE1A might play a role in Ras induced senescence.
We now show that NORE1A is a potent mediator of Ras induced senescence. Knockdown of NORE1A suppresses the ability of Ras to induce senescence in multiple cell systems and enhances Ras driven transformation. We have identified two novel signaling pathways activated by Ras/NORE1A. First, NORE1A forms a Ras regulated, endogenous complex with the kinase HIPK2. HIPK2 can phosphorylate p53 to induce apoptosis or recruit acetyltransferases to acetylate p53 to induce senescence signaling. NORE1A suppresses HIPK2 apoptotic post-translational modifications of p53 but activates pro-senescence post-translational modifications. NORE1A also binds and destabilizes mdm2 to enhance the stability of nuclear p53. Primary human tumors show a close correlation between the expression levels of NORE1A and acetylated p53.
In addition to p53, we show that NORE1A also links Ras to the regulation of Retinoblastoma (Rb) protein. NORE1A promotes a potent, Ras dependent stabilization of the Rb protein. It also promotes dephosphorylation of RB, an activating event.
Thus, NORE1A is a double-barreled Ras senescence effector which connects Ras to two of the most important senescence regulating tumor suppressors in the cell.
Loss of NORE1A activity in tumors is usually due to epigenetic inactivation or aberrant protein degradation by calpains. Both of these mechanisms, in principal, may be subject to clinical intervention to restore NORE1A function. This may provide a novel approach to antagonizing Ras driven tumors.
Citation Format: Howard Donninger, Diego Calvisi, Thibaut Barnoud, M. Lee Schmidt, Geoffrey J. Clark. NORE1A is a double-barreled Ras senescence effector linking Ras to p53 and Rb. [abstract]. In: Proceedings of the AACR Special Conference on RAS Oncogenes: From Biology to Therapy; Feb 24-27, 2014; Lake Buena Vista, FL. Philadelphia (PA): AACR; Mol Cancer Res 2014;12(12 Suppl):Abstract nr A06. doi: 10.1158/1557-3125.RASONC14-A06
Abstract A08: Mechanistic dissection of Ral GTPase signaling in driving KRAS-dependent pancreatic cancer growth
Authors: Gentry Leanna R. , Martin Timothy D. , Reiner David J. , Der Channing J. 
Published: May 2015

Abstract
The high frequency (95%) of KRAS mutations in pancreatic ductal adenocarcinomas (PDAC), together with strong evidence for the KRAS dependency of PDAC growth, makes the K-Ras oncoprotein an attractive therapeutic target. However, to date the development of direct inhibitors of K-Ras has not been successful, with most efforts focused on targeting components of K-Ras downstream effector signaling. While the pharmaceutical industry has centered on inhibitors of the Raf or PI3K effector signaling pathways, there is substantial evidence that K-Ras-dependent cancer growth must rely on the activities of other effector pathways. Of those pathways, the Ral guanine nucleotide exchange factor RalGEF) effector pathway, leading to activation of the Ras-like (Ral) small GTPases is well-validated to drive PDAC growth. Interestingly, while the two Ral isoforms share significant sequence identity (82%) as well as the same set of effectors, our studies found that they have strikingly different roles in PDAC growth. While RalA but not RalB is necessary for PDAC anchorage-independent growth in vitro and tumorigenic growth in vivo, RalB is the critical driver of PDAC invasion in vitro and metastasis in vivo. The mechanistic basis for their different biological roles in cancer remains to be elucidated. We hypothesize that their divergent C-terminal sequences and the posttranslational modifications that they signal for result in distinct subcellular localizations that then contribute to Ral isoform-selective effector interactions that promote distinct biological consequences in PDAC. To evaluate our hypothesis, we initiated studies to determine the effector(s) that mediates RalA-dependent anchorage-independent growth and tumorigenesis in vivo, the effector(s) that mediates RalB-dependent invasion and metastasis in vivo; and the role of the C-terminal hypervariable domain in causing distinct RalA- and RalB-dependent phenotypes and effector utilization. We found that both RalA and RalB subcellular localization is dependent on the C-terminal CAAX tetrapeptide signaled modifications by Rce1 and ICMT. We also determined that RalB preferentially activated mTORC1 to regulate pancreatic tumor cell Matrigel migration, through the Sec5 effector and regulation of exocyst function.
Citation Format: Leanna R. Gentry, Timothy D. Martin, David J. Reiner, Channing J. Der. Mechanistic dissection of Ral GTPase signaling in driving KRAS-dependent pancreatic cancer growth. [abstract]. In: Proceedings of the AACR Special Conference on RAS Oncogenes: From Biology to Therapy; Feb 24-27, 2014; Lake Buena Vista, FL. Philadelphia (PA): AACR; Mol Cancer Res 2014;12(12 Suppl):Abstract nr A08. doi: 10.1158/1557-3125.RASONC14-A08
Abstract A11: Oncogenic Ras signaling involves sustained perinuclear localization of the effector kinases p-ERK1/2 and CK2 via KSR-1 and endosomal trafficking
Authors: Lee Sook , Basu Sandip K. , Walia Vijay , Morrison Deborah K. , Johnson Peter F. 
Published: May 2015

Abstract
In primary cells oncogenic Ras can induce cellular senescence, an important tumor suppression mechanism that must be bypassed in tumor cells. Oncogene-induced senescence is regulated in part by the transcription factor C/EBPβ, which is post-translationally activated by Ras signaling. We previously reported that C/EBPβ activation by H-RasV12 is suppressed in immortalized and transformed cells by a novel 3'UTR-dependent mechanism termed 3'UTR regulation of protein activity, or UPA. UPA acts by restricting the Cebpb mRNA to a peripheral cytoplasmic region, which is spatially distinct from a perinuclear compartment containing the C/EBPβ-activating kinase, p-ERK1/2. UPA can thereby block C/EBPβ phosphorylation/activation and suppress its pro-senescence functions in tumor cells. Here we identify CK2 as another C/EBPβ kinase that undergoes perinuclear relocalization in RasV12-expressing cells. Accordingly, CK2-mediated phosphorylation of C/EBPβ was inhibited by the Cebpb 3'UTR in tumor cells but not primary cells. The MAPK signaling scaffold KSR-1 and endocytic trafficking were both required for RasV12-induced activation of C/EBPβ and perinuclear targeting of p-ERK and CK2. CK2 co-localized and interacted with KSR-1 and Rab11 (a marker of recycling endosomes), while p-ERK appeared to be associated with a different endosomal population. RasV12 expression led to sustained perinuclear localization of p-ERK and CK2 in both transformed and senescent cells. In contrast, growth factors (GFs) only transiently induced this response, which occurred with delayed kinetics (4-6 hr) coinciding with transitory activation of C/EBPβ. Our results indicate that oncogenic Ras signaling involves constitutive activation of the delayed GF signaling phase, characterized by restriction of the effector kinases p-ERK and CK2 to perinuclear endosomes. In this compartment they can access critical targets such as C/EBPβ in a UPA-regulated manner to promote either transformation or senescence.
Citation Format: Sook Lee, Sandip K. Basu, Vijay Walia, Deborah K. Morrison, Peter F. Johnson. Oncogenic Ras signaling involves sustained perinuclear localization of the effector kinases p-ERK1/2 and CK2 via KSR-1 and endosomal trafficking. [abstract]. In: Proceedings of the AACR Special Conference on RAS Oncogenes: From Biology to Therapy; Feb 24-27, 2014; Lake Buena Vista, FL. Philadelphia (PA): AACR; Mol Cancer Res 2014;12(12 Suppl):Abstract nr A11. doi: 10.1158/1557-3125.RASONC14-A11
Abstract A20: ARHGEF2 inhibition impairs cell growth and viability in KRAS-dependent cancer cells
Authors: Vargas Maria Jose Sandi , Kent Oliver A. , Normand Josee , Rottapel Robert 
Published: May 2015

Abstract
Sporadic tumours carrying activating mutations of the RAS family of GTPases are characterized by a progressive and multiple step tumorigenesis. Among the three isoforms, K-RAS is the most commonly mutated (Prior et al., 2012). ARHGEF2 is a microtubule-associated guanine nucleotide exchange factor (GEF) for the Rho family of small GTPases whose expression is up-regulated (more than 30 times) upon Ras neoplastic transformation in Rat fibroblasts (Zuber et al., 2000). A truncated version of the protein can transform NIH 3T3 cells (Whitehead et al., 1995) and induce tumour formation in nude mice (Brecht et al., 2005). We recently identified ARHGEF2 as a transcriptional target of the RAS/MAPK pathway and its expression contributes to cell survival and growth in RAS-transformed cells (Cullis et al., in revision). We aimed to determine if ARHGEF2 is required for tumour cell viability in different KRAS mutant cancer cell lines. We used an RNAi approach using two different ARHGEF2 and KRAS shRNAs to monitor cell growth and viability in six different cancer cell lines harbouring a KRAS mutation: four pancreatic adenocarcinomas (PL45, Panc 02.03, Panc 04.03 and KP4), one endometrial adenocarcinoma (HEC-1-A) and one colorectal adenocarcinoma (LS 174T). We found that ARHGEF2 is required for cell growth and survival in half of the cell lines analyzed. We established an ARHGEF2 and KRAS dependency index for those cell lines and further analysis allowed us to determine a positive correlation (correlation coefficient of 0.64, p value &lt;0.03) between ARHGEF2 and KRAS dependency. In conclusion, our findings suggest that: i/ ARHGEF2 plays a role in the tumorigenesis and survival of KRAS dependent cell lines, but further in vitro and in vivo studies are required; ii/ ARHGEF2 inhibition could be a potential therapeutic target in tumours harbouring KRAS mutations.
References:
Prior, IA et al. A comprehensive survey of Ras mutations in cancer. Cancer Res. 2012.
Zuber, J. et al. A genome-wide survey of RAS transformation targets. Nat Gen. 2000.
Whitehead, I et al. Expression cloning of lfc, a novel oncogene with structural similarities to guanine nucleotide exchange factors and to the regulatory region of protein kinase C. J Biol Chem. 1995.
Brecht, M. et al. Activation of gef-h1, a guanine nucleotide exchange factor for RhoA, by DNA transfection. Int J Cancer. 2005.
Cullis, J. et al. The Rho GEF-H1 is required for RAS oncogene-diven pancreatic cancer. In revision.
Citation Format: Maria Jose Sandi Vargas, Oliver A. Kent, Josee Normand, Robert Rottapel. ARHGEF2 inhibition impairs cell growth and viability in KRAS-dependent cancer cells. [abstract]. In: Proceedings of the AACR Special Conference on RAS Oncogenes: From Biology to Therapy; Feb 24-27, 2014; Lake Buena Vista, FL. Philadelphia (PA): AACR; Mol Cancer Res 2014;12(12 Suppl):Abstract nr A20. doi: 10.1158/1557-3125.RASONC14-A20