bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract C04: HGFL promotes the development and progression mammary tumors and represents an attractive therapeutic target
Authors: Benight Nancy M. , Wagh Purnima , Waltz Susan E.
Published: August 2014

A standard therapy for breast cancer is the use of receptor tyrosine kinase (RTK) inhibitors. RTKs efficiently target multiple receptors, including the Ron transmembrane receptor, whose activation leads to increased survival, migration, and angiogenesis through signaling pathways such as PI3K/Akt and NF-κB. Ron overexpression is correlated with increased metastasis and poor outcomes in patients. The only ligand for Ron is hepatocyte growth factor-like protein (HGFL), which is an endocrine factor secreted into the circulation from the liver. Published reports demonstrated that ectopic overexpression of HGFL in cancer cells leads to increased metastasis. However, no studies have examined the role of endogenous HGFL in Ron activation during tumor development and metastasis. We sought to test the hypothesis that HGFL is required for Ron receptor activation and mammary tumorigenesis. We utilized mice with mammary-specific Ron overexpression (MMTV-Ron), which develop mammary tumors with 100% incidence. MMTV-Ron mice were crossed to HGFL deficient (HGFL-/-) mice to generate MMTV-Ron X HGFL-/- mice. We compared mammary hyperplasia and tumor latency in MMTV-Ron mice with and without HGFL serially at 2.5, 4, 6, 8 and 10 months. Additional mice were followed to established tumor kinetics and growth and to examine metastatic dissemination. MMTV-Ron X HGFL-/- mice displayed decreased mammary hyperplasia and a significant delay in the time to tumor formation compared to control mice. Kinase assays show that Ron activation is reduced in mice with HGFL loss. Tumor cell proliferation was different between the groups while markers of apoptosis (TUNEL IHC and cleaved caspase 3 expression) were significantly increased in MMTV-Ron X HGFL-/- mice compared to controls. Western blot analysis of tumor lysates showed diminished activation of NF-κB signaling associated with HGFL loss. In conclusion, our data show that HGFL promotes the development of mammary hyperplasia and mammary tumor formation by enhancing tumor cell survival, Ron activation and NF-κB signaling. These data suggest that HGFL represents an excellent target for inhibiting Ron mediated tumor growth in human disease.

Citation Format: Nancy M. Benight, Purnima Wagh, Susan E. Waltz. HGFL promotes the development and progression mammary tumors and represents an attractive therapeutic target. [abstract]. In: Proceedings of the Third AACR International Conference on Frontiers in Basic Cancer Research; Sep 18-22, 2013; National Harbor, MD. Philadelphia (PA): AACR; Cancer Res 2013;73(19 Suppl):Abstract nr C04.
Abstract C13: Novel molecular targets for breast cancer stem cells involved in HER-PI3K signaling
Authors: Gotoh Noriko
Published: August 2014

Breast cancer stem cells (BCSCs) are responsible for breast tumorigenesis. To develop more effective cancer therapies, it would be reasonable to target molecules that have a critical role in the maintenance of BCSCs. However, our understanding of the cellular and molecular mechanisms underlying BCSC properties is limited. We have previously shown that heregulin (HRG), a ligand for HER3, induced tumorsphere formation of breast cancer cells through a phosphatidyl inositol 3 (PI3) kinase pathway (Proc. Natl. Acad. Sci., USA, 109, p6584, 2012). To examine dynamic changes in gene expression patterns by stimulation of HER/PI3 kinase signaling, we performed time series microarray analyses under condition of HRG treatment and/or PI3 kinase inhibition by using a specific inhibitor, and obtained a HER/PI3 kinase gene signature that includes 635 genes. We found that the HER/PI3 kinase gene signature is more enriched in high-grade breast tumors than in low-grade tumors and is associated with high-risk patients for relapse. We found that several growth factors, cytokines and cytoplasmic molecules among the molecules in the HER/PI3 kinase gene signature, play important roles for tumorsphere formation derived from the clinical samples form the breast cancer patients. These findings suggest that novel molecular targets for BCSCs can be found in the HER/PI3 kinase signature.

Citation Format: Noriko Gotoh. Novel molecular targets for breast cancer stem cells involved in HER-PI3K signaling. [abstract]. In: Proceedings of the Third AACR International Conference on Frontiers in Basic Cancer Research; Sep 18-22, 2013; National Harbor, MD. Philadelphia (PA): AACR; Cancer Res 2013;73(19 Suppl):Abstract nr C13.
Abstract C17: Revealing targetable cancer cell vulnerabilities by in silico pathway profiling
Authors: Joshi Rashika , Komurov Kakajan
Published: August 2014

Successful targeted therapy of cancers relies on comprehensive understanding of vulnerabilities of the tumorigenic platform. Recent cancer genomics efforts have provided with an unprecedented opportunity to gain an unbiased insight into the functional make-up of human cancers. We have devised a computational strategy to leverage cancer genomic datasets to identify candidate pathway vulnerabilities in cancers. The core of our approach is in the delineation of molecular pathways that are activated in tumor cells as a result of secondary selection events during tumor evolution. These secondary pathways are likely to be required for tumorigenesis in the given tumor context. We applied this strategy on HER2+ breast cancers using breast cancer genomic data in the TCGA, the recently published METABRIC and the Cancer Cell Line Encyclopedia (CCLE) datasets. Although HER2+ breast cancers are treated with HER2-targeting agents in the clinic, the response is usually short-lived due to de novo and acquired resistance. We have found that HER2+ breast cancers are characterized by extensive and robust secondary upregulation of the endoplasmic reticulum (ER) quality control (ERQC) pathways, including the ER-associated degradation (ERAD) and PERK/eIF2 pathways. Our follow-up combined in silico and in vitro analyses on a panel of HER2+ and HER2- breast cancer cell lines revealed a model of HER2-driven tumorigenesis where yin-yang interplay between hyperactive HER2/mTOR signaling and the ERQC machinery regulate the ER protein homeostasis to control survival of HER2+ cells. Furthermore, we report that this delicate regulatory architecture can be exploited for therapeutic purposes, such that, combinatorial targeting of ERQC pathways to disturb the proteostatic balance synergized in selectively inducing proteotoxic cell death in therapy-refractory HER2+ cells. This study presents a computational method for identification of cancer pathway vulnerabilities, and suggests that exploiting proteotoxic ER stress may be an effective therapeutic strategy in therapy-refractory HER2+ breast cancers.

Note: This abstract was not presented at the conference.

Citation Format: Rashika Joshi, Kakajan Komurov. Revealing targetable cancer cell vulnerabilities by in silico pathway profiling. [abstract]. In: Proceedings of the Third AACR International Conference on Frontiers in Basic Cancer Research; Sep 18-22, 2013; National Harbor, MD. Philadelphia (PA): AACR; Cancer Res 2013;73(19 Suppl):Abstract nr C17.
Abstract C24: High systemic cholesterol increases blood vessel endothelial permeability and favors metastasis: Are these events related?
Authors: Magalhaes Ana , Santos Catarina R. , Domingues Germana , Carvalho Tãnia , Dias Sergio
Published: August 2014

Here we hypothesize that systemic cholesterol favors metastasis by increasing blood vessel endothelial permeability. The majority of metastatic dissemination occurs through the hematogenous route and factors that alter blood vessel properties can affect the entry of cancer cells in circulation. It is generally accepted that high levels of systemic cholesterol, in particular low density lipoprotein (LDL), have a negative impact in endothelial function. This has been demonstrated mainly in large arteries but less is known about how LDL affects the microvasculature of different organs. High cholesterol has also been associated with poor prognosis in several cancers. In our lab, using orthotopic mouse models of breast cancer, we were able to show that mice in which increased LDL in circulation is induced by a high cholesterol diet, have more metastasis. Here we also show in an “in vitro” system, that LDL increases endothelial permeability and that this can be reverted by blocking the interaction of LDL with the LDL-receptor. Additionally we observe that this higher endothelial permeability is due to cholesterol enrichment of endothelial cells, as nystatin, a cholesterol sequestering agent is also able to revert the phenotype. Back to “in vivo”, using the same orthotopic breast cancer model, we are able to show that high systemic cholesterol increases the number of circulating cancer cells in the blood. We are now testing the hypothesis of whether this is due to increased permeability of tumor blood vessels or of the vessels surrounding the tumor.

Citation Format: Ana Magalhaes, Catarina R. Santos, Germana Domingues, Tãnia Carvalho, Sergio Dias. High systemic cholesterol increases blood vessel endothelial permeability and favors metastasis: Are these events related? [abstract]. In: Proceedings of the Third AACR International Conference on Frontiers in Basic Cancer Research; Sep 18-22, 2013; National Harbor, MD. Philadelphia (PA): AACR; Cancer Res 2013;73(19 Suppl):Abstract nr C24.
Abstract C35: Neural stem cells transduced with adenovirus Interferon-beta prolonged lives of MDA 231 lung tumor bearing SCID mice
Authors: Rachakatla Raja S. , Reuter Ashley , Hassan Sherif S.
Published: August 2014

Conventional cancer therapies are not selective and leave patients of metastatic breast cancer with poor prognosis and serious adverse effects. It has been shown that neural stem cells (NSCs) migrate specifically to cancer affected tissues and can be engineered to carry anti-cancer drugs; Therefore, NSCs may be used as drug delivery vehicles to specifically target cancer cells. Our goal was to determine if NSCs transduced with adenovirus expressing IFN-beta (NSC-IFN-beta) would target and kill MDA 231 human breast cancer in vitro and in vivo. In vitro: transwells were used to determine if NSCs and NSC-IFN-beta migrate towards and kill MDA 231 cells in vitro. NSCs and NSC-IFN-beta migrate towards MDA 231 cells (P < 0.05) and conditioned MDA 231 media (P < 0.05). However, only NSC-IFN-beta kill MDA 231 cells in vitro (P < 0.05). In vivo: A survival study was performed to test the efficacy of NSC-IFN-beta on MDA 231 lung tumor bearing SCID mice. Tumor bearing mice had greater lung weights (P < 0.05). Tumor bearing mice treated with NSC-IFN-beta lived longer than untreated mice (PBS) (median survival: 45 days versus 37 days, difference = 8 days, P = <0.05 and tumor bearing mice treated with NSCs (median survival: 45 days versus 31 days, difference = 14 days, P < 0.05). Conclusion: These results suggest that NSCs with IFN-beta may be able to target and reduce human breast cancer in a metastatic SCID mice tumor model.

Citation Format: Raja S. Rachakatla, Ashley Reuter, Sherif S. Hassan. Neural stem cells transduced with adenovirus Interferon-beta prolonged lives of MDA 231 lung tumor bearing SCID mice. [abstract]. In: Proceedings of the Third AACR International Conference on Frontiers in Basic Cancer Research; Sep 18-22, 2013; National Harbor, MD. Philadelphia (PA): AACR; Cancer Res 2013;73(19 Suppl):Abstract nr C35.
Abstract C37: Reduced annexin A6 expression enhances EGFR degradation and sensitizes invasive breast cancer cells to EGFR-targeted tyrosine kinase inhibitors
Authors: Koumangoye Rainelli B. , Ochieng Josiah , Sakwe Amos M.
Published: August 2014

The role of annexin A6 (AnxA6), a calcium-dependent phospholipid binding protein, in breast carcinogenesis remains poorly understood. Some reports suggest that AnxA6 inhibits the activation and down-stream signaling of epidermal growth factor receptor (EGFR). On the contrary, other studies have shown that AnxA6 not only stabilizes activated receptors on the cell surface but also promotes the expression of differentiation markers in chondrocytes. Together with our recent study showing that AnxA6 expression is reduced in breast cancer, we investigated whether AnxA6 expression status influences breast cancer outcome in patients and whether this may be attributed to its modulation of EGFR expression. We show that low AnxA6 expression correlates with reduced distant metastasis-free and overall survival, but is associated with a higher probability for recurrence-free survivalof patients with basal-like breast cancer. To determine whether these effects are mediated via EGFR, a predictive marker of basal-like breast cancer, we examined the effects of altered expression of AnxA6 in AnxA6-high invasive triple negative breast cancer cells. Interestingly, depletion of AnxA6 in invasive BT-549 cells was accompanied by decreased expression of EGFR protein and mRNA. We also show that in invasive breast cancer cells, AnxA6 expression is required for sustained membrane localization of activated EGFR and persistent downstream signaling. Consequently, AnxA6-depleted cells exhibited decreased cell motility, and increased sensitivity to the tyrosine kinase inhibitors (TKIs), lapatinib and PD153035. On the contrary, over-expression of AnxA6 in AnxA6-low HCC1806 cells did not affect their motility but as expected, inhibited their growth in 3D matrigel cultures. These data suggest that AnxA6-modulation of EGFR expression and activity underlies at least in part, the changes in proliferation, metastatic potentials and sensitivity to EGFR-targeted TKIs of invasive breast cancer cells. Our data also suggest that AnxA6 expression status may predict the survival and likelihood to respond to EGFR-targeted therapies of patients with basal-like breast cancer.

Citation Format: Rainelli B. Koumangoye, Josiah Ochieng, Amos M. Sakwe. Reduced annexin A6 expression enhances EGFR degradation and sensitizes invasive breast cancer cells to EGFR-targeted tyrosine kinase inhibitors. [abstract]. In: Proceedings of the Third AACR International Conference on Frontiers in Basic Cancer Research; Sep 18-22, 2013; National Harbor, MD. Philadelphia (PA): AACR; Cancer Res 2013;73(19 Suppl):Abstract nr C37.
Abstract C38: Iroquois homeobox gene 3 is temporally regulated in response to vascular endothelial growth factor and modulates endothelial cell migration and tubulogenesis
Authors: Scarlett Kisha A. , Pattabiraman Vaishnavi , Anderson Leonard M.
Published: August 2014

Introduction: Differences in tumor-induced angiogenesis contributes to cancer-related health disparities. Understanding the transcriptional regulation of angiogenesis within endothelial cells (ECs) is essential to the development of anti-angiogenic therapies. Here we report that the homeobox gene, Irx3 is expressed in human microvascular endothelial cells (HMECs) and modulates their migration and ability to form networks during angiogenesis.

Methods: To test the effects of vascular endothelial growth factor (VEGF) on Irx3 expression, HMECs were treated with VEGF for 0 to 48 hours followed by extraction of total RNA and protein. Irx3 mRNA and protein expression were determined using qRT-PCR and western immunoblot respectively. Wound-healing and transwell migration assays were performed to investigate the role of Irx3 on EC migration in response to VEGF. For migration experiments, HMECs were transduced with adenoviral vectors resulting in the loss-of (Ad.mirIrx3-GFP) or gain-of (Ad.Irx3-V5) Irx3 function. We conducted a 672 transcription factor (TF) siRNA screen to identify transcriptional regulators of Irx3 expression in response to VEGF. HMECs were reverse-transfected with siRNAs and treated with VEGF for 12 hours. Total RNA was extracted for RT-PCR to determine Irx3 expression. Finally, we performed in vitro endothelial cell tubulogenesis assays using HMECs transduced with Ad.Irx3-V5, Ad.mirIrx3-GFP and their respective controls to study changes in endothelial tube formation as a result of differential Irx3 expression.

Results: VEGF treatment increases Irx3 expression at the mRNA and protein levels, with peak expression at 12 hours. HMECs transduced with Ad.mirIrx3-GFP exhibited decreased migration at 12 hours compared to HMECs transduced with Ad.Irx3-V5. Furthermore, when HMECs are transduced with Ad.Irx3-V5, they exhibit increased migration and invasiveness in transwell migration assays and form more complex networks during tube formation. The siRNA screen suggests that Irx3 is regulated by HEY1, a known transcriptional regulator of angiogenesis.

Conclusion: Our results provide compelling evidence that the homeobox gene Irx3 regulates EC migration, and is directly regulated by TFs known to govern transcriptional signaling during angiogenesis.

Grant Support: This work was supported in part by the following grants; the CVRI Center of Excellence Grant, NIH/NCRR/RCMI G12-RR03034, and the MBRS/RISE Program 2R25GM058268.

Citation Format: Kisha A. Scarlett, Vaishnavi Pattabiraman, Leonard M. Anderson. Iroquois homeobox gene 3 is temporally regulated in response to vascular endothelial growth factor and modulates endothelial cell migration and tubulogenesis. [abstract]. In: Proceedings of the Third AACR International Conference on Frontiers in Basic Cancer Research; Sep 18-22, 2013; National Harbor, MD. Philadelphia (PA): AACR; Cancer Res 2013;73(19 Suppl):Abstract nr C38.
Abstract C41: Chc1L is a tumor suppressor involved in development of histiocyte-rich neoplasms
Authors: Spillane David , Wang Ding Yan , Shi Changxin , Wang Youdong , Gramolini Anthony , Stewart Keith , Newbigging Susan , Liu Mingyao , Wen Xiao-Yan
Published: August 2014

Human chromosome 13q14 is one of the hotspots of chromosomal deletion in prostate cancer, multiple myeloma, and chronic lymphocytic leukemia. Chromosome Condensation 1-like (CHC1L) is an uncharacterized gene in this region. CHC1L is found within the smallest common region of loss of heterozygosity in prostate cancer, and its decreased expression is linked to pathogenesis and progression of both prostate cancer and multiple myeloma. In the present study, we describe the generation and characterization of Chc1L gene knockout mice. Knockout mice develop normally and display an exaggerated proliferative response to LPS stimulation of cultured bone marrow and spleen cells at ten weeks of age. At approximately two years of age, both homozygous and heterozygous mice have markedly increased incidence of tumorigenesis, 80% and 56% respectively, compared to wild-type mice, 26%. The tumors are distributed most commonly to the spleen, mesenteric lymph nodes, liver and small intestine. The morphologic and immunophenotypic features of the tumor cells are consistent with those of tissue histiocytes, and most Chc1L knockout and heterozygote mice succumb to either Histiocytic Sarcoma or Histiocyte-Associated Lymphoma. These data provide the first direct evidence that Chc1L is a tumor suppressor gene involved in suppression of histiocyte-rich neoplasms.

Citation Format: David Spillane, Ding Yan Wang, Changxin Shi, Youdong Wang, Anthony Gramolini, Keith Stewart, Susan Newbigging, Mingyao Liu, Xiao-Yan Wen. Chc1L is a tumor suppressor involved in development of histiocyte-rich neoplasms. [abstract]. In: Proceedings of the Third AACR International Conference on Frontiers in Basic Cancer Research; Sep 18-22, 2013; National Harbor, MD. Philadelphia (PA): AACR; Cancer Res 2013;73(19 Suppl):Abstract nr C41.
Abstract C57: Observation of experimental and clinic curative effect on Z-C medicine treating malignancy
Authors: Xu Ze , Xu Jie , Zhu Shiping , Qio Shaoming , Wu Bin
Published: August 2014

• Objectives: To find traditional Chinese medicines of treating carcinoma which are good for curative effect and have no side effect.

• Methods We use liver cancer H2,2 to inoculate 260 Kun Ming little white mice which are divided into group Z-C1 and group Z-C4, group of chemical treatment (CTX) and group of contrapose.

• Results: We observe that the repression rate of mice H22 in liver cancer to of group Z-C1 for 2, 4, 6 weeks is 40%, 45% and 58%, and repression rate of mice H22 in liver cancer to of group Z-C4 for 2, 4, 6 weeks is 55%,68% and 70%, while repression rate of group CTX for 2, 4 and 6 weeks is 45%, 45% and 49%. For ten years' clinic abservation of 4277 middle and later period cancer patients, we find that Z-C medicine has good effect to prolong survival and has no side response for long time taking patients.

• Conclusions: Through experimental research and clinic validation, it is proved that Z-C1, Z-C4 medicine can improve the survival quality and immunity of middle and later period patients, and control the increase of carcinoma cells to get a further curative effect

Citation Format: Ze Xu, Jie Xu, Shiping Zhu, Shaoming Qio, Bin Wu. Observation of experimental and clinic curative effect on Z-C medicine treating malignancy. [abstract]. In: Proceedings of the Third AACR International Conference on Frontiers in Basic Cancer Research; Sep 18-22, 2013; National Harbor, MD. Philadelphia (PA): AACR; Cancer Res 2013;73(19 Suppl):Abstract nr C57.
Abstract IA1: Lgr5 stem cells in self-renewal and cancer
Authors: Clevers Hans
Published: August 2014

The intestinal epithelium is the most rapidly self-renewing tissue in adult mammals. We originally defined Lgr5 as a Wnt target gene, transcribed in colon cancer cells. Two knock-in alleles revealed exclusive expression of Lgr5 in cycling, columnar cells at the crypt base. Using an inducible Cre knock-in allele and the Rosa26-LacZ reporter strain, lineage tracing experiments were performed in adult mice. The Lgr5+ve crypt base columnar cells (CBC) generated all epithelial lineages throughout life, implying that it represents the stem cell of the small intestine and colon. Similar obserations were made in hair follicles and stomach epithelium.

Single sorted Lgr5+ve stem cells can initiate ever-expanding crypt-villus organoids in 3D culture. Tracing experiments indicate that the Lgr5+ve stem cell hierarchy is maintained in these organoids. We conclude that intestinal crypt-villus units are self-organizing structures, which can be built from a single stem cell in the absence of a non-epithelial cellular niche. The same technology has now been developed for the Lgr5+ve stomach stem cells.

Intestinal cancer is initiated by Wnt pathway-activating mutations in genes such as APC. As in most cancers, the cell of origin has remained elusive. Deletion of APC in stem cells, but not in other crypt cells results in progressively growing neoplasia, identifying the stem cell as the cell-of-origin of adenomas. Moreover, a stem cell/progenitor cell hierarchy is maintained in early stem cell-derived adenomas, lending support to the “cancer stem cell”-concept.

Fate mapping of individual crypt stem cells using a multicolor Cre-reporter revealed that, as a population, Lgr5 stem cells persist life-long, yet crypts drift toward clonality within a period of 1-6 months. Lgr5 cell divisions occur symmetrically. The cellular dynamics are consistent with a model in which the resident stem cells double their numbers each day and stochastically adopt stem or TA fates after cell division. Lgr5 stem cells are interspersed between terminally differentiated Paneth cells that are known to produce bactericidal products. We find that Paneth cells are CD24+ and express EGF, TGF-a, Wnt3 and the Notch ligand Dll4, all essential signals for stem-cell maintenance in culture. Co-culturing of sorted stem cells with Paneth cells dramatically improves organoid formation. This Paneth cell requirement can be substituted by a pulse of exogenous Wnt. Genetic removal of Paneth cells in vivo results in the concomitantloss of Lgr5 stem cells. In colon crypts, CD24+ cells residing between Lgr5 stem cells may represent the Paneth cell equivalents. We conclude that Lgr5 stem cells compete for essential niche signals provided by a specialized daughter cell, the Paneth cell.

Citation Format: Hans Clevers. Lgr5 stem cells in self-renewal and cancer. [abstract]. In: Proceedings of the Third AACR International Conference on Frontiers in Basic Cancer Research; Sep 18-22, 2013; National Harbor, MD. Philadelphia (PA): AACR; Cancer Res 2013;73(19 Suppl):Abstract nr IA1.
Abstract IA13: Glioma stem cells and cancer
Authors: Parada Luis F.
Published: August 2014

Malignant GBMs are incurable tumors that progress within a year's time. Genome wide studies have pinpointed prevalent cancer associated mutions in GBM and among these, the tumor suppressors p53, Pten and NF1 are among the most frequently found. We have developed mouse models of GBM via mutation of p53, Pten and NF1 in the brain. Among the many key insights gleaned from studies with our mouse models have been: 1) The first demonstration that stem cells are the major source of these tumors. 2) The demonstration that all tumor cells are not equivalent and that the Cancer Stem Cells, a subpopulation of cells, are the root of tumor progression. 3) The discovery of a distinct form of GBM that arises in a different cell population and that has unique properties. This novel form of GBM has until now been grouped together with the predominant form of GBM. Our work will lead to a separation of prognosis and treatment for the different forms. In addition, using high throughput screens, we have small compounds in development that specifically target GBM cells but not normally dividing cells. I will discuss these advances and the implications for glioma, solid tumors and cancer in general.

Citation Format: Luis F. Parada. Glioma stem cells and cancer. [abstract]. In: Proceedings of the Third AACR International Conference on Frontiers in Basic Cancer Research; Sep 18-22, 2013; National Harbor, MD. Philadelphia (PA): AACR; Cancer Res 2013;73(19 Suppl):Abstract nr IA13.
Abstract IA15: Genetic and functional tumor heterogeneity
Authors: Swanton Charles
Published: August 2014

Despite advances in genomic technologies, most advanced solid tumors remain incurable and drug resistance is almost inevitable with limited biomarkers available to personalize therapy. Two important lessons have emerged from the comprehensive genomic analyses of cancers, which may provide an explanation for difficulties that have been encountered in biomarker development. First, each tumor contains an individual assortment of multiple genomic aberrations, few of which are shared between patients with the same histopathological tumor subtype. Second, emerging evidence suggests that these anomalies appear to vary within the tumor, indicating substantial intratumor heterogeneity. Increasingly, molecular evidence suggests that intratumor heterogeneity may contribute to tumor growth through a branched (polytypic) rather than a linear pattern of tumor evolution. Branched evolutionary growth and intratumor heterogeneity results in coexisting cancer cell subclones with variegated genotypes and associated phenotypes that may be regionally separated within the same tumor and alter in dominance over time. Variegated phenotypes, resulting from intratumoral genetic heterogeneity and the emergence of new subclones at relapse, are likely to have important implications for developing novel targeted therapies and for preventing the emergence of drug resistance. Intratumor heterogeneity and sampling bias may also contribute to the recently reported failures in implementation of robust biomarkers in the clinical setting. In this talk, evidence for genetic, epigenetic and transcriptomic intratumour heterogeneity, impacting upon signal transduction pathway phenotypic heterogeneity, will be reviewed. Darwinian models for biomarker discovery will be discussed based on trunk and branched tumor growth models that may improve drug discovery and clinical outcome prediction.

Citation Format: Charles Swanton. Genetic and functional tumor heterogeneity. [abstract]. In: Proceedings of the Third AACR International Conference on Frontiers in Basic Cancer Research; Sep 18-22, 2013; National Harbor, MD. Philadelphia (PA): AACR; Cancer Res 2013;73(19 Suppl):Abstract nr IA15.
Abstract IA16: Dysregulated mTORC1 renders cells critically dependent on desaturated lipids for survival under tumor-like stress
Authors: Young Regina , Ackerman Daniel , Quinn Zachary , Mancuso Anthony , Gruber Michaela , Liu Liping , Giannoukos Dionysios , Bobovnikova-Marjon Ekaterina , Diehl J. Alan , Keith Brian , Simon M. Celeste
Published: August 2014

Solid tumors exhibit heterogeneous microenvironments, often characterized by limiting concentrations of oxygen (O2), glucose, and other nutrients. How oncogenic mutations alter stress response pathways, metabolism, and cell survival in the face of these challenges is incompletely understood. Here we report that constitutive mTORC1 activity renders hypoxic cells dependent on exogenous desaturated lipids, as levels of de novo synthesized unsaturated fatty acids are reduced under low O2. Specifically, we demonstrate that hypoxic Tsc2-/- cells deprived of serum lipids exhibit a magnified UPR response, but fail to appropriately expand their ER, leading to IRE1-dependent cell death that can be reversed by the addition of unsaturated lipids. UPR activation and apoptosis were also detected in Tsc2-deficient kidney tumors. Importantly, we observe this phenotype in multiple human cancer lines and suggest that cells committed to unregulated growth within ischemic tumor microenvironments are unable to balance lipid and protein synthesis due to a critical limitation in desaturated lipids.

Citation Format: Regina Young, Daniel Ackerman, Zachary Quinn, Anthony Mancuso, Michaela Gruber, Liping Liu, Dionysios Giannoukos, Ekaterina Bobovnikova-Marjon, J. Alan Diehl, Brian Keith, M. Celeste Simon. Dysregulated mTORC1 renders cells critically dependent on desaturated lipids for survival under tumor-like stress. [abstract]. In: Proceedings of the Third AACR International Conference on Frontiers in Basic Cancer Research; Sep 18-22, 2013; National Harbor, MD. Philadelphia (PA): AACR; Cancer Res 2013;73(19 Suppl):Abstract nr IA16.
Abstract PR02: “Guardians of repeats”: Novel and significant relationship of p53 and interferon
Authors: Leonova Katerina , Gudkov Andrei
Published: August 2014

Large parts of mammalian genomes are transcriptionally inactive and enriched with various classes of interspersed and tandem repeats. Here we show that the tumor suppressor protein p53, along with DNA methylation, plays an essential role in maintaining silencing of a large portion of the mouse genome. In the absence of functional p53, hypomethylation of DNA results in massive transcription of major classes of short interspersed repeats (SINEs B1 and B2), both strands of near-centromeric satellite DNAs consisting of tandem repeats, and multiple species of non-coding RNAs. In p53-deficient mouse fibroblasts treated with the DNA demethylating agent 5-aza-2'-deoxycytidine, the abundance of these new transcripts exceeds the level of β-actin mRNA by more than 150-fold. Accumulation of these largely double-stranded transcripts, in the absence of p53, is accompanied with a strong endogenous type I interferon response that induces apoptotic cell death. Transcriptional activation of repeats and non-coding RNAs is observed in a significant proportion of spontaneous mouse tumors, presumably reflecting naturally occurring DNA hypomethylation and p53 inactivation. These observations suggest that p53 and IFN cooperate to prevent accumulation of cells with activated repeats. This model, which we have named TRAIN (Transcription of Repeats Activates INterferon), assigns a novel role to p53 as a guardian of repeats. TRAIN model provides plausible explanation for frequent deregulation of interferon pathway observed in a variety of tumors. Overall, this work highlights the cooperation between p53 and interferon as key players needed to maintain genetic stability, and even further outlines their significance in tumor progression and the evolution of the species.

Citation Format: Katerina Leonova, Andrei Gudkov. “Guardians of repeats”: Novel and significant relationship of p53 and interferon. [abstract]. In: Proceedings of the Third AACR International Conference on Frontiers in Basic Cancer Research; Sep 18-22, 2013; National Harbor, MD. Philadelphia (PA): AACR; Cancer Res 2013;73(19 Suppl):Abstract nr PR02.
Abstract A18: The role of the microenvironment protein cathelicidin LL-37 in pancreatic ductal adenocarcinoma
Authors: Sainz Bruno , Frias Javier , Cioffi Michele , Alcala Sonia , Heeschen Christopher
Published: August 2014

This abstract is being presented as a short talk in the scientific program. A full abstract is printed in the Proffered Abstracts section (PR1) of the Conference Proceedings.

Citation Format: Bruno Sainz, Jr., Javier Frias, Michele Cioffi, Sonia Alcala, Christopher Heeschen. The role of the microenvironment protein cathelicidin LL-37 in pancreatic ductal adenocarcinoma. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Invasion and Metastasis; Jan 20-23, 2013; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2013;73(3 Suppl):Abstract nr A18.
Abstract A2: The importance of metabolic heterogeneity and its consequences on tumor invasion, metastatic growth, and treatment
Authors: Robertson-Tessi Mark , Gillies Robert J. , Gatenby Robert A. , Anderson Alexander R.A.
Published: August 2014

Heterogeneity of both the tumor microenvironment and cellular phenotype plays a key role in guiding tumor behavior. Metabolism acts as a central integrator to mediate cellular response to the changing environment, linking the effects of hypoxia, acidosis, proliferation and necrosis. Using a hybrid multi-scale mathematical model of tumor growth in a vascularized tissue, we investigate the selection pressures exerted by the tumor microenvironment as the cancer progresses. A key focus is the metabolism of both normal and tumor cells, and how small changes in phenotype can lead to the development of more metabolically aggressive groups of cells. Development of a glycolytic phenotype due to microenvironmental selection pressures leads to acid-mediated invasion. Results from the model suggest that these aggressive cells have a dramatic advantage as a metastatic seed, compared to less aggressive seeds from the same primary. In addition, there are significant differences in metastatic growth rate depending on the soil, suggesting that prevention or treatment of metastases in different organs may require different strategies. Application of pH buffering therapy has been shown in mouse models to slow down or prevent both primary tumor and metastatic growth. This treatment has been simulated in the model, and the results show that the effect of the treatment is highly sensitive to the timing of application. Additional results suggest that diffusible cytotoxic treatments such as chemotherapy may increase the metabolic aggressiveness of a tumor post-treatment due to the altered selection pressure caused by the drug. Chemotherapy removes the metabolic stratification of the tumor and allows more aggressive cells to grow towards blood vessels and normal tissue. A third treatment we consider is the use of anti-angiogenic therapy, which foments the development of aggressive phenotypes due to degradation of the tumor microenvironment. The model highlights the role of metabolic heterogeneity and microenvironmental selection in driving tumor invasion and metastatic development.

Citation Format: Mark Robertson-Tessi, Robert J. Gillies, Robert A. Gatenby, Alexander R.A. Anderson. The importance of metabolic heterogeneity and its consequences on tumor invasion, metastatic growth, and treatment. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Invasion and Metastasis; Jan 20-23, 2013; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2013;73(3 Suppl):Abstract nr A2.
Abstract A35: Aligned collagen is a prognostic signature for survival in human breast carcinoma
Authors: Conklin Matthew W. , Eickhoff Jens , Riching Kristin , Pehlke Carolyn , Eliceiri Kevin , Provenzano Paolo , Friedl Andreas , Keely Patricia J.
Published: August 2014

Evidence for the potent influence of stromal organization and function on invasion and metastasis of breast tumors is ever growing. Here we have performed a rigorous examination of the relationship of a tumor-associated collagen signature (TACS-3), to the long term survival rate of human patients diagnosed with invasive breast cancer. TACS-3 is characterized by bundles of straightened and aligned collagen fibers that are oriented perpendicular to the tumor boundary. An evaluation of TACS-3 was performed in biopsied tissue sections from 196 patients by second harmonic generation (SHG) imaging of the backscattered signal generated by collagen. Univariate analysis of a Cox proportional hazard model demonstrated that the presence of TACS-3 was associated with poor disease-specific and disease free survival, resulting in hazard ratios between 3.0-3.9. Furthermore, TACS-3 was confirmed to be an independent prognostic indicator regardless of tumor grade and size, ER or PR status, HER-2 status, node status, and tumor subtype. Interestingly, TACS-3 was positively correlated to expression of stromal syndecan-1, a receptor for several extracellular matrix proteins including collagens. Ongoing research is currently investigating both the biochemical signaling pathways that give rise to the TACS-3 phenotype as well as the extent of the influence the mechanical properties of an aligned matrix have on invasion. Because of the strong statistical evidence for poor survival in patients with TACS, and since the assessment can be performed in routine histopathologic samples imaged via SHG or using picrosirius, we propose that quantifying collagen alignment is a viable, novel paradigm for the prediction of human breast cancer survival.

Citation Format: Matthew W. Conklin, Jens Eickhoff, Kristin Riching, Carolyn Pehlke, Kevin Eliceiri, Paolo Provenzano, Andreas Friedl, Patricia J. Keely. Aligned collagen is a prognostic signature for survival in human breast carcinoma. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Invasion and Metastasis; Jan 20-23, 2013; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2013;73(3 Suppl):Abstract nr A35.
Abstract A45: An in vivo examination of the effects of ARID3B on ovarian cancer metastasis in nude mice
Authors: Roy Lynn , Samyesudhas Serene , Carrasco Martin , Dahl Karen Cowden
Published: August 2014

In 2011, 21,880 women were diagnosed with ovarian cancer and more than 13,850 died from the disease. Unfortunately, the prognosis for most women diagnosed with ovarian cancer is poor due to most patients being diagnosed with local and distant metastases. ARID3B, a DNA binding protein from the AT-rich interactive domain (ARID) family, is overexpressed in late stage neuroblastoma and ovarian cancer. In early development, ARID3B is essential as evidenced by the deaths of Arid3B null embryos by midgestation. The importance of ARID3B in early development and its overexpression in cancer suggest a potential role in cancer progression and metastasis. In addition, our laboratory identified two isoforms of ARID3B a full length (FL) and a short form (SH) that is missing the c-terminal exons 5-9. To determine the effects of the two ARID3B isoforms on metastasis in vivo, human ovarian cancer SKOV3IP cells (selected for their ability to metastasize) overexpressing red fluorescent protein (RFP), ARID3B-FL, or ARID3B-Sh were injected intraperitoneally into nude mice. The mice underwent live optical fluorescence imaging using the Multispectral FX (Kodak) imaging system at frequent intervals to assess tumor burden. The mice were sacrificed when they began to show signs of significant illness. Live imaging of the mice demonstrated that ARID3B-FL expressing SKOV3IP cells formed more and larger tumors more quickly than either SKOV3IP-RFP cells or SKOV3-ARID3B-Sh cells. After sacrificing the mice, tumors and organs were recovered from the abdomen and were imaged ex vivo for tumor burden. Tumors were separated from the organs, weighed, and imaged. The average weights of the tumors were 1.91g for SKOV3IP-RFP derived tumors, 3.22g for SKOV3IP ARID3B-Sh tumors, and 5.46g for SKOV3IP-FL derived tumors. The average tumor burden was greatest for the mice bearing SKOV3IP ARID3B-FL tumors. Gross examination of the tumors post-mortem and post fixation suggested that the tumors from the RFP and ARID3B-Sh expressing cell lines were less vascular or had less vascular leakage than those from the ARID3B-FL mice. Survival analysis showed that the median survival for SKOV3IP ARID3B-FL tumor bearing mice was significantly lower than mice bearing RFP or the ARID3B-Sh expressing tumors (RFP=51 days, ARID3B-Sh=52 days, ARID3B-FL=36 days; P=0.0074). Currently, our lab is addressing potential mechanisms for how ARID3B functions in metastasis. The data presented here suggest that ARID3B plays a significant role in peritoneal metastasis in ovarian cancer.

Citation Format: Lynn Roy, Serene Samyesudhas, Martin Carrasco III, Karen Cowden Dahl. An in vivo examination of the effects of ARID3B on ovarian cancer metastasis in nude mice. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Invasion and Metastasis; Jan 20-23, 2013; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2013;73(3 Suppl):Abstract nr A45.
Abstract A56: Bone morphogenetic protein antagonist DMH1 inhibits metastasis in a mouse model of breast cancer
Authors: Owens Philip , Pickup Michael W. , Gorska Agnieszka E. , Novitskiy Sergey V. , Hong Charles C. , Moses Harold L.
Published: August 2014

Bone Morphogenetic Proteins (BMPs) are secreted cytokines/growth factors belonging to the Transforming Growth Factor β (TGFβ) superfamily. BMPs have recently been shown to be overexpressed in human breast cancers; however, loss of BMP signaling in mammary carcinomas has also been shown to accelerate metastases. Human breast cancers display active BMP signaling by phosphorylated Smads 1, 5 and 8 in both the tumor and the stromal microenvironment. We hypothesized that inhibiting BMP signaling in both the tumor epithelium as well as the surrounding microenvironment would prevent tumor progression. To test this hypothesis we used DMH1, which is a selective and specific antagonist of the type I BMP receptor. Mice that express the MMTV.PyVmT oncogene were either treated with DMSO vehicle or DMH1 for six weeks following initial mammary tumor palpation. Following six weeks, animals were euthanatized and evaluated for lung metastases. Whole mount staining of lungs revealed a statically significant reduction in lung metastasis in DMH1 treated animals compared to vehicle treated (p-value = 0.03). Primary tumors demonstrated reduced proliferation (BrdU incorporation) and increased apoptosis (cleaved-caspase-3) when treated with DMH1. RNA isolated from peripheral blood at the time of sacrifice indicated decreased circulating tumor cells measured by real-time PCR. Overall, the BMP pathway represents an understudied pathway that may be suitable for targeted therapy in breast cancer.

Citation Format: Philip Owens, Michael W. Pickup, Agnieszka E. Gorska, Sergey V. Novitskiy, Charles C. Hong, Harold L. Moses. Bone morphogenetic protein antagonist DMH1 inhibits metastasis in a mouse model of breast cancer. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Invasion and Metastasis; Jan 20-23, 2013; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2013;73(3 Suppl):Abstract nr A56.
Abstract A71: CD146 is a novel target of CD44-signaling mediating breast tumor invasion
Authors: Ouhtit Allal , Abdraboh Mohamed , Hollenbach Andrew D. , Gupta Ishita , Shanmuganathan Somya , Riyami Hamad Al , Errami Youssef , Farsi Yahya Al , Raj Madhwa HG
Published: August 2014

Cancer cell metastasis is one of the most critical steps in tumor development and is responsible for more than 80% of cancer related deaths. The cell adhesion molecule CD44 promotes cancer cell motility and metastasis. Despite this knowledge, the molecular mechanism through which CD44 promotes tumor development and cell metastasis is still unclear. The CD146 (MUC 18), another member of the cell adhesion molecules family, first identified in highly metastatic melanomas. The absence of CD146 in normal melanocytes and its high expression in melanomas suggests its tumor promoting actions. However, CD146 appears to suppress breast tumor progression. Furthermore, despite the association between CD146 expression and development of melanoma, the expression patterns and the role of CD146 in normal and metastatic breast tissues are still controversial. In a pilot study, we have identified CD146 as a potential downstream target of CD44-signaling in the MDA-435 breast cancer cell line, using subtractive hybridization and northern blot analyses. In this paper we provide evidence that CD146 is a downstream target for CD44, in a way that CD146 expression is regulated according to tumor microenvironment. Finally, this study demonstrates a new role for CD44 in regulating neovascularization and promoting cancer cell transmigration of blood vessels via CD146.

Citation Format: Allal Ouhtit, Mohamed Abdraboh, Andrew D. Hollenbach, Ishita Gupta, Somya Shanmuganathan, Hamad Al Riyami, Youssef Errami, Yahya Al Farsi, Madhwa HG Raj. CD146 is a novel target of CD44-signaling mediating breast tumor invasion. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Invasion and Metastasis; Jan 20-23, 2013; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2013;73(3 Suppl):Abstract nr A71.
Abstract A72: TGF-β2 underpins CD44-promoted breast tumor cell invasion
Authors: Ouhtit Allal , Madani Samineh , Gupta Ishita , Shanmuganathan Somya , Al-Riyami Hamad , Al-Farsi Yahya
Published: August 2014

Hyaluronan (HA) mediates communication between cancer cells and the environment via interactions with the cell surface receptor CD44. The long-term objective of this study is to increase our understanding of the mechanisms by which CD44-HA interaction promotes BC metastasis, and further identify and validate CD44-downstream transcriptional targets for anti-metastatic therapy. Here, we have developed a tetracycline (tet)-regulated expression of CD44 gene in the BC cell line MCF-7 (B5 clone) and identified TGF-beta2 (Transforming Growth Factor beta-2; 3 fold induction) as a potential CD44s-downstream transcriptional target by microarray analysis. The same RNA samples, used for microarray analysis and their corresponding protein lysates, were examined for CD44 expression by RT-PCR and western-Blot methods, respectively. TGF-beta2 mRNA levels were significantly elevated following the removal of tet at 18, 24, and 48 h post-HA stimulation compared to controls. Furthermore, the TGF-beta2 precursor protein increased in a time-dependent pattern upon HA-stimulation and in the absence of tet. Inhibition of CD44 gene by RNAi method decreased TGF-beta2 expression upon HA-stimulation and in the absence of tet. Our data put together strongly support the hypothesis that TGF-beta2 is a potential target of HA/CD44-downstream-signaling mediating BC cell invasion. Finally, CREB might be the transcription factor linking CD44 to TGF-beta2 transcription.

Citation Format: Allal Ouhtit, Samineh Madani, Ishita Gupta, Somya Shanmuganathan, Hamad Al-Riyami, Yahya Al-Farsi. TGF-β2 underpins CD44-promoted breast tumor cell invasion. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Invasion and Metastasis; Jan 20-23, 2013; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2013;73(3 Suppl):Abstract nr A72.