bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract IA6: Genetic strategies for next generation of breast cancer therapies
Authors: Silva Jose M.
Published: August 2014

One of the major challenges for modern cancer treatments is the ability to eliminate tumor cells without affecting normal cell homeostasis. We propose to identify the “Achilles' heel” of cancer cells using the novel concept of the “noncancer addition.” Additionally to alterations in bona-fide cancer genes, in a tumor cell, multiple normal regulatory networks have been rearranged in order to adapt to the tumorigenic state. As consequence of this divergence, survival of cancer cells also depends on noncancer genes that are essential to maintain the tumor homeostasis. Therefore, we postulate that interfering with these noncancer dependencies will result in system failure, that is, the cessation of the tumorigenic state. We are combining the power of interactome-prediction models with the state-of-the-art RNAi-genetic approaches to block the function of every gene in the entire genome to find genetic synthetic lethal interactions between noncancer genes and bona-fide cancer genes.

Recently, we have applied this strategy searching for novel therapeutic alternatives for ErbB2 positive breast cancer patients. ErbB2 is a receptor tyrosine kinase found overexpressed in 20-40% of breast tumors (ErbB2+ tumors) and correlates with poor prognosis. Different targeted therapies have been developed to specifically inhibit its activity. Unfortunately, the majority of these patient tumors eventually progress acquiring resistance. Thus, there is the need to find alternative tumor targets to develop specific, more efficient, treatments for these patients.

By combining genome-wide RNAi loss-of-function screens with system biology interactome models we have recently found that the JAK/STAT3 pathway is activated in ErbB2 overexpressing breast cancers through an autocrine loop mediated by the secretion of Interleukin-6 (IL-6). Importantly, blockage of this pathway strongly reduced the viability of ErbB2+ cancer cells. The identification of Stat3 as a tumor target in ErbB2+ tumors opens new avenues for patients which tumors have become refractory to standard protocols.

Citation Format: Jose M. Silva. Genetic strategies for next generation of breast cancer therapies [abstract]. In: Proceedings of the AACR Special Conference on Chemical Systems Biology: Assembling and Interrogating Computational Models of the Cancer Cell by Chemical Perturbations; 2012 Jun 27-30; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2012;72(13 Suppl):Abstract nr IA6.
Abstract PR3: Considering alternate signaling mechanisms models in extrinsic apoptosis using programmatic modeling approaches
Authors: Lopez Carlos F. , Muhlich Jeremy L. , Bachman John J. , Sorger Peter K.
Published: August 2014

Experiments often result in observations that suggest conflicting biochemical mechanisms in signaling networks. Mathematical modeling of biological systems is a method used to probe such experimental observations and provide a consensus among seemingly discordant observations. However, probing multiple mechanistic hypotheses in biological modeling often involves the instantiation of complex systems of equations, which despite their usefulness, makes model revision, extension, and sharing extremely challenging. To address these modeling barriers, we have developed a modeling framework that brings a process-based approach to biological modeling. In our approach, biological models are written as programs that encode biological functions as needed. Our modeling framework, developed in the Python language, offers access to a large set of existing numerical and programming methods to biological systems modeling. Given our choice of programming language, the resulting models are easier to share, distribute, extend, and revise, adding transparency to the model-creation and execution process. We demonstrate our approach by exploring three competing mechanisms that describe the interactions among the Bcl-2 family of proteins and their contributions to mitochondrial outer membrane permeabilization in extrinsic apoptosis. We used our modeling framework to systematically explore these three proposed mechanisms resulting in the instantiation of sixteen biochemical model topologies for numerical exploration. Our preliminary results, based on simulations calibrated to experimental data, suggest that the so-called indirect mechanism does not accurately reproduce experimental observations. Results from simulations suggest that the so-called embedded model more seems to better align with existing experimental data. Despite these observations, further topology- and numerical-based characterization of the model topology and parameter-space is being carried out to more accurately understand the role of these Bcl-2 proteins in apoptosis. We aim to formulate hypotheses, testable through experiments, about the interaction among these proteins and their role in Bcl-2 cancer biology.

This proffered talk is also presented as Poster A9.

Citation Format: Carlos F. Lopez, Jeremy L. Muhlich, John J. Bachman, Peter K. Sorger. Considering alternate signaling mechanisms models in extrinsic apoptosis using programmatic modeling approaches [abstract]. In: Proceedings of the AACR Special Conference on Chemical Systems Biology: Assembling and Interrogating Computational Models of the Cancer Cell by Chemical Perturbations; 2012 Jun 27-30; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2012;72(13 Suppl):Abstract nr PR3.
Abstract PR9: Investigating combinatorial ligand addiction provides insights into rational drug combinations in cancer therapy
Authors: Pace Emily A. , Nielsen Ulrik B. , Schoeberl Birgit , Chai Diana H. , Parikh Anand , Kalra Ashish , Oyama Shinji , Johnson Bryan , Tan Gege , Fulgham Aaron , Bukhalid Raghida
Published: August 2014

Cancer, the second most common cause of death in the United States, is a collection of diseases caused by uncontrolled cell growth and metastasis. The main treatment for cancer is chemotherapy, which generally kills fast growing cells nonspecifically and has many side effects. A different type of cancer treatment, called targeted therapy, aims to avoid general toxicity by using drugs that block the activity of specific gene products, usually encoded by oncogenes, which have been shown to drive tumor growth. To date, targeted therapies, alone or in combination with chemotherapies, have mainly been successful in rare subsets of patients with tumors addicted to single oncogenes. This has created a rationale to mainly treat patients with an oncogene-addiction (such as those carrying mutated or overexpressed kinases) with targeted therapies like erlotinib and trastuzumab, which inhibit human epidermal growth factor receptor (EGFR) and human epidermal growth factor receptor 2 (HER2/ErbB2), respectively. Here, evidence is provided that targeted therapies are also effective in tumors that are dependent on multiple growth factors – a phenomenon that is called combinatorial ligand addiction. Specifically, it is shown that ligands that bind the EGFR family and the hepatocyte growth factor receptor (HGFR/MET) can activate protein kinase B (PKB/AKT) across a broad set of cancer cell lines, suggesting that ligand signaling is redundant and widespread. It is also shown that ErbB ligands have distinct signaling dynamics and strengths, which provides a rationale for investigating each component of the ErbB signaling network. Using a systematic approach, we found that ErbB3 is an important therapeutic target even though it is not overexpressed and lacks kinase activity. Furthermore, it is shown that cell lines with and without known oncogene-addiction express autocrine ligands and have improved growth inhibition with drug combinations that include autocrine ligand-blocking antibodies. This research demonstrates that combinatorial ligand addiction creates a new rationale for therapeutic combinations to improve efficacy and prevent resistance in cancer cells that are treated with current targeted drugs.

This proffered talk is also presented as Poster A22.

Citation Format: Emily A. Pace, Ulrik B. Nielsen, Birgit Schoeberl, Diana H. Chai, Anand Parikh, Ashish Kalra, Shinji Oyama, Bryan Johnson, Gege Tan, Aaron Fulgham, Raghida Bukhalid. Investigating combinatorial ligand addiction provides insights into rational drug combinations in cancer therapy [abstract]. In: Proceedings of the AACR Special Conference on Chemical Systems Biology: Assembling and Interrogating Computational Models of the Cancer Cell by Chemical Perturbations; 2012 Jun 27-30; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2012;72(13 Suppl):Abstract nr PR9.
Abstract A05: Phosphoinositide-3-kinase pathway promotes colon cancer stem cell proliferation
Authors: Chen Sugong , Fisher Robert C. , Shenoy Anitha K. , Lopez Maria Cecilia , Moldawer Lyle L. , Baker Henry V. , Huang Emina H.
Published: August 2014

Introduction. The cancer stem cell (CSC) model posits that a select cell-subpopulation exists in solid tumors that sustain tumorigenesis. Previously, we have shown that aldehyde-dehydrogenase (ALDH) enriches colon CSCs (CCSC) from sporadic colon cancer. In this study, Affymetrix microarray was used to screen for differences between ALDHhi and ALDHlo cells isolated from sporadic human colon adenocarcinoma. As a result, the phosphoinositide-3-kinase (PI3K) pathway was identified as a potential regulator of CCSC growth and differentiation. Functional assays demonstrate that inhibition of PI3K results in significant reduction in proliferation and colony formation of by CCSCs.

Experimental Procedures. Unbiased Expression-Profile Analysis: We implanted human CCSCs into flanks of NOD-SCID mice. Flow assisted cell sorting discriminated resulting tumor epithelia into ALDHhi and ALDHlo groups from 6 individual sporadic human adenocarcinomas, which were analyzed using Affymetrix GeneChip Human Genome U133 Plus 2.0 Array. Proliferation Assay: Bromodeoxyuridine (BrdU) incorporation measured the effect of LY294002 (Calbiochem) (1-100 μM) on proliferation. Colony Assay: Methylcellulose colony assays were used to assess the effect of LY294002 on colony formation.

Summary of Data. Expression Profile Analysis revealed 136 genes that were significantly up or down-regulated in the ALDHhi cell groups (>3-fold; p<0.001). The beta regulatory subunit of phosphoinositide-3 kinase demonstrated a 5.2 fold up-regulation in the ALDHhi vs. ALDHlo CCSCs. Cell Proliferation Assay: LY294002 significantly inhibits CCSC proliferation with IC50 ~ 50 μM and near complete inhibition at the dose of 100 μM (p<0.001). Colony Assay: LY294002 significantly reduces total colony area formed by CCSCs, with a similar trend as shown in BrdU proliferation assays (p<0.001).

Conclusions. We have previously shown that ALDH enriches colon cancer epithelia. Here we show that the PI3K signaling pathway is differentially expressed between ALDHhi and ALDHlo CCSCs derived from primary tumor xenografts. We demonstrate in vitro functional differences in CCSCs with drug-inhibition assays.

Citation Format: Sugong Chen, Robert C. Fisher, Anitha K. Shenoy, Maria Cecilia Lopez, Lyle L. Moldawer, Henry V. Baker, Emina H. Huang. Phosphoinositide-3-kinase pathway promotes colon cancer stem cell proliferation. [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 A05.
Abstract A14: Transforming growth factor beta signaling regulates the fate of intestinal stem cells
Authors: Fischer Jared M. , Miller Ashleigh , Calabrese Peter P. , Shibata Darryl , Liskay R Michael
Published: August 2014

Transforming growth factor beta signaling has important roles in stem cell behavior, differentiation, cell motility, cell cycle and apoptosis. Here, we have utilized a novel mouse model that facilitates the sporadic inactivation of TgfβR2 via frameshift reversion of Cre in single, isolated cells and concomitant lineage tracing with a Cre-reporter. Short-term lineage tracing of TgfβR2-deficient crypts revealed reduced expansion suggesting a growth disadvantage among crypts. In contrast, long-term lineage tracing of TgfβR2-deficient crypts revealed an increased number suggesting a growth advantage within the crypt. To reconcile these apparently contradictory findings, we studied the dynamics of proliferative, Lgr5+ stem cells following TgfβR2 loss. We found that TgfβR2-deficient stem cell lineages required more time to reach monoclonality within the crypt. Using mathematical modeling of stem cell neutral drift, we found that increased “stemness” or a greater probability of a self-renewing asymmetrical outcome for TgfβR2-deficient stem cells was more consistent with our experimental results. Further analysis of TgfβR2-deficient stem cells in vivo revealed a decreased production of Paneth cells, a finding we corroborated in cultured intestinal organoids by treating with either a TgfβR1/2 inhibitor or low levels of the Tgfβ1 ligand. Recent studies by others suggested that quiescent stem cells are intermediaries between proliferative Lgr5+ stem cells and Paneth cells and that Tgfβ signaling is important in the formation of quiescent stem cells in other tissues. Therefore, we are currently asking whether Tgfβ signaling is important in the formation of quiescent stem cells in the intestine.

Citation Format: Jared M. Fischer, Ashleigh Miller, Peter P. Calabrese, Darryl Shibata, R Michael Liskay. Transforming growth factor beta signaling regulates the fate of intestinal stem cells. [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 A14.
Abstract A15: A reciprocal feedback loop between grainyhead-like-2 and ZEB1 controls EMT and tumor suppression
Authors: Cieply Benjamin , Ford Heide L. , Farris Joshua , Frisch Steven
Published: August 2014

The oncogenic Epithelial-Mesenchymal Transition (EMT) enhances malignant progression by promoting invasion and survival of tumor cells. EMT is induced by microenvironmental factors including TGF-β and Wnt agonists, and transcription factors including the E-box binding factors Twist, Snail and ZEB. Previously, we reported that a member of the mammalian Grainyhead family of wound healing-regulatory transcription factors, Grainyhead-like-2 (GRHL2), suppresses EMT and restores sensitivity to anoikis by repressing ZEB1 expression and inhibiting TGF-β signaling. Here, we elucidate the functional relationship between GRHL2 and ZEB1 in EMT/MET and tumor biology. At least three homeodomain proteins, Six1, LBX1, and HoxA5, transactivated the ZEB1 promoter; for Six1, this occurred through direct protein-promoter interaction. GRHL2 altered the Six1-DNA complex, inhibiting this transactivation. Correspondingly, GRHL2 expression prevented tumor initiation in xenograft assays, sensitized breast cancer cells to paclitaxel, and suppressed the emergence of CD44highCD24low cells in a cell line where this marker set correlates with cancer stem cell phenotype. GRHL2 was down-regulated in recurrent mouse tumors that had evolved to an oncogene-independent, EMT-like state, supporting a role for GRHL2 down-regulation in this tumor phenotype transition modeling disease recurrence. The combination of TGF-β and Wnt activation repressed GRHL2 expression through direct interaction of ZEB1 with the GRHL2 promoter, inducing EMT. These observations indicate that a reciprocal feedback loop between GRHL2 and ZEB1 controls epithelial vs. mesenchymal phenotypes and EMT-driven tumor progression.

Citation Format: Benjamin Cieply, Heide L. Ford, Joshua Farris, Steven Frisch. A reciprocal feedback loop between grainyhead-like-2 and ZEB1 controls EMT and tumor suppression. [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 A15.
Abstract A33: Aneuploidy due to inhibition of Aurora B induces p21 through activation of p38 by proteotoxic stress and ROS
Authors: Kumari Geeta , Ulrich Tanja , Gaubatz Stefan
Published: August 2014

Faithful transmission of genomic information to the two daughter cells requires highly coordinated cell cycle regulation. Aurora kinases have been implicated in mediating critical events during mitosis and cytokinesis. Aurora B is the enzymatic component of the chromosome passenger complex (CPC) which shows a characteristic localization during various phases of mitosis. Both the expression level and kinase activity of Aurora B is upregulated in several primary tumors including breast cancer and colon cancer and is associated with poor prognosis. Therefore, Aurora B is an attractive anti-cancer target and several chemical inhibitors of Aurora B are already in clinical trials. Inhibition of Aurora B results in cell cycle arrest. However, the signaling pathways responsible for the phenotypes observed after Aurora B inhibition have not yet been fully explored. We have recently shown that the p53 target gene p21 (CDKN1A) is activated both at protein and mRNA levels after pharmacological inhibition of Aurora B and that p21-activation is dependent on p53 and on the p38 MAP kinase. While the stabilization of p53 and its recruitment to the p21 promoter occur independently of p38, transcriptional elongation of p21 depends on active p38 after Aurora B is inhibited. We further show that partial inhibition of Aurora B is sufficient to induce p21. FISH (Fluorescence in-situ hybridization) assays indicate increased aneuploidy without inhibition of cytokinesis when Aurora B is partially inhibited. Increased aneuploidy is correlated with generation of reactive oxygen species (ROS) and proteotoxic stress, but not with DNA damage. In summary, our data suggest that aneuploidy due to partial inhibition of Aurora B results in proteotoxic stress and in the production of ROS, which contributes to the induction of p21 through activation of p38 MAPK. Significantly, Aurora B inhibitors and drugs that act on aneuploid cells (AICAR and 17AAG) synergize in the inhibition of cell proliferation and in inhibition of transformation. Thus, combining Aurora B inhibitors with drugs that act on aneuploid cells may result in a more effective cancer therapy than Aurora B inhibitors alone.

Note: This abstract was not presented at the conference.

Citation Format: Geeta Kumari, Tanja Ulrich, Stefan Gaubatz. Aneuploidy due to inhibition of Aurora B induces p21 through activation of p38 by proteotoxic stress and ROS. [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 A33.
Abstract A45: Investigating the role of ASCL1 in regulating differentiation of gliobastoma precursor cells
Authors: Park Nicole I. , Dirks Peter B.
Published: August 2014

Glioblastoma (GBM), the most common and lethal adult primary brain tumour, exhibits cellular heterogeneity and a subpopulation of tumour cells exhibits the neural precursor phenotype and drives tumourigenesis. These findings suggest that tumours may represent aberrant organogenesis with growth caused by unlimited proliferation and failure of differentiation of malignant precursor cells. It has yet to be determined the relative importance of aberrant self-renewal versus blocked differentiation in the brain tumour phenotype. As GBM-initiating cells have precursor cell properties, promotion of differentiation represents a potential strategy for treatment. Forcing glioblastoma precursor cells (GPCs) to differentiate terminally would limit clonal expansion and attenuate tumour growth. Data generated in our lab suggests that differentiation potential varies between different patient-derived GPC cultures. This study aims to determine what defines a blocked differentiation phenotype in human glioblastoma and to determine whether differentiation therapy is a feasible strategy for glioblastoma treatment.

A key regulator of neuronal differentiation is ASCL1 during normal development and adult neurogenesis. Microarray analysis of ASCL1 expression in GPCs (n = 33) revealed two subgroups of GPCs as having either high or low expression. Preliminary evidence suggests that GPCs with high expression of ASCL1 exhibit neuronal lineage commitment whereas GPCs with low expression of ASCL1 do not undergo lineage commitment. This was confirmed using immunocytochemistry, quantitative real-time PCR and Western blot analysis. Furthermore, an inverse correlation was observed between transcript levels of ASCL1 and DKK1, an antagonist of the Wnt signalling pathway, while a direct correlation was observed between ASCL1 and activated β-catenin (ABC). Treatment of GPCs with a GSK3β inhibitor resulted in the activation of Wnt signalling and concomitant increase in transcript levels of ASCL1 and neuronal lineage markers (i.e. TUJ1, MAP2).

This study aims to further investigate the role of ASCL1 in glioblastoma and to determine whether neuronal lineage commitment in the context of ASCL1 activity is mediated in a Wnt-dependent manner. Gain- and loss-of-function studies are in progress to determine whether ASCL1 is sufficient and/or necessary for neuronal lineage commitment and consequent effects on tumour-initiating properties (i.e. proliferation and self-renewal) will be examined in vitro and in vivo.

Citation Format: Nicole I. Park, Peter B. Dirks. Investigating the role of ASCL1 in regulating differentiation of gliobastoma precursor cells. [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 A45.
Abstract A49: Repression of tumor survival pathways by novel, selective inhibitors of MNK1 and MNK2 kinases
Authors: Rzymski Tomasz , Dreas Agnieszka , Milik Mariusz , Kucwaj Katarzyna , Zarebski Adrian , Skuta Malgorzata Szajewska , Cierpich Anna , Fabritius Charles , Brzozka Krzysztof
Published: August 2014

MNK1 and MNK2 are MAP kinase-interacting kinases involved in regulation of translation. Both kinases phosphorylate translation initiation factor eIF4E on a conserved serine 209. eIF4E can contribute to the oncogenic transformation both in vitro and in vivo and is highly expressed in diverse types of cancer. Interestingly, mice that lack both MNK1 and MNK2 do not have any apparent phenotype and therefore represent interesting possibility to develop targeted and safe anticancer therapies.

Herewith, we report the development of first selective small molecule inhibitors of MNK1 and MNK2 kinases and their in vitro cellular activity. Selvita developed a series of small molecule type I, ATP-competitive inhibitors targeting both MNK1 and MNK2 with mid nM activity range. Selected compounds were tested on a panel of 456 kinases and showed very high selectivity. Additionally, in cellular models such as serum stimulated SW480 cells, synthesized MNK1/2 inhibitors caused dose dependent inhibition of phosphorylation of eIF4e at Ser209 in line with the kinase activity profile. The observed cellular activity on biomarker inhibition was more potent than observed for reported in the literature MNK1/2 inhibitors such as cercosporamide and CGP57380. High activity of MNK1/2 inhibitors on in vitro biomarkers correlated with efficacy on cancer cells challenged with various stress conditions, typical for tumor microenvironment. These initial findings confirm that selective inhibition of MNK1/2 may repress major tumor survival pathways induced under stress and support further development of this class of compounds as a novel anticancer therapy with a promising therapeutic window.

Citation Format: Tomasz Rzymski, Agnieszka Dreas, Mariusz Milik, Katarzyna Kucwaj, Adrian Zarebski, Malgorzata Szajewska Skuta, Anna Cierpich, Charles Fabritius, Krzysztof Brzozka. Repression of tumor survival pathways by novel, selective inhibitors of MNK1 and MNK2 kinases. [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 A49.
Abstract B09: Liver-specific knockout of GRP94 in mice disrupts cell adhesion, activates liver progenitor cells, and accelerates liver tumorigenesis
Authors: Chen Wan-Ting , Tseng Chun-Chih , Pfaffenbach Kyle , Kanel Gary , Luo Biquan , Stiles Bangyan L. , Lee Amy S.
Published: August 2014

Liver cancer is one of the most common solid tumors with poor prognosis and high mortality. Mutation or deletion of the tumor suppressor PTEN is strongly correlated with human liver cancer. Glucose-regulated protein 94 (GRP94) is a major endoplasmic reticulum (ER) chaperone protein, but its in vivo function is still emerging. To study the role of GRP94 in maintaining liver homeostasis and tumor development, we created two liver-specific knockout mouse models with the deletion of Grp94 alone, or in combination with Pten, using the albumin-cre system. We demonstrated that while deletion of GRP94 in the liver led to hyperproliferation of liver progenitor cells, deletion of both GRP94 and PTEN accelerated development of liver tumors, including both hepatocellular carcinoma (HCC) and cholangiocarcinoma (CC) suggestive of progenitor cell origin. Furthermore, at the premalignant stage, we observed disturbance of cell adhesion proteins but no detectable overt liver injury. We further discovered that when GRP94 was deleted in PTEN-null livers, ERK was selectively activated, suggesting that this could be a potential molecular mechanism for enhanced liver tumorigenesis in this mouse model. Conclusion: We identify GRP94 as a novel regulator of cell adhesion, liver homeostasis, and tumorigenesis.

Citation Format: Wan-Ting Chen, Chun-Chih Tseng, Kyle Pfaffenbach, Gary Kanel, Biquan Luo, Bangyan L. Stiles, Amy S. Lee. Liver-specific knockout of GRP94 in mice disrupts cell adhesion, activates liver progenitor cells, and accelerates liver tumorigenesis. [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 B09.
Abstract B48: Breast tumor cell TACE-shed MCSF promotes pro-angiogenic macrophages through NF-κB signaling
Authors: Rego Stephen L. , Helms Rachel , Dreau Didier
Published: August 2014

Mortalities associated with breast cancer, the most common malignancy in women, are mainly due to metastases. Tumor associated macrophages (TAMs) significantly contribute to the development of metastasis through promotion of the rate limiting step of angiogenesis through secretion of vascular endothelial growth factor (VEGF). Macrophage differentiation and migration are activated by macrophage colony stimulating factor (MCSF) and chemokine (C-C motif) ligand 2 (CCL2), respectively. Whereas tumor cells secrete CCL2, MCSF release is mediated by ectodomain shedding through tumor necrosis factor alpha converting enzyme activation (TACE). How tumor cells support macrophage promotion of angiogenesis remains unclear. In the present study, we determined whether MCSF shed by TACE from tumor cells promotes angiogenesis through the activation of NF-κB signaling in macrophages and subsequent VEGF release. These studies were modeled in vitro using a panel of murine mammary cells mimicking breast cancer progression from normal murine mammary gland (NMuMG) cells to metastatic 4T1 cells along with J774 macrophages. TACE and MCSF expressions were higher in metastatic cells compared to epithelial cells (p<0.05). After incubation with 4T1 tumor conditioned media (CM), macrophages secreted significantly more VEGF and expressed of the activated NF-κB, and the resulting macrophage secretions promoted significantly high levels of endothelial cell tubes (p<0.05). Furthermore, a similar activation of macrophages was generated by the combination of CCL2, also highly expressed by tumor cells, and MCSF. These results highlight the key role of tumor cell TACE-shed MCSF and secreted CCL2 in synergistically stimulating pro-angiogenic macrophages.

Citation Format: Stephen L. Rego, Rachel Helms, Didier Dreau. Breast tumor cell TACE-shed MCSF promotes pro-angiogenic macrophages through NF-κB 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 B48.
Abstract C03: A comprehensive study of genomic alterations reveals deregulation of the cell cycle in most esophageal squamous cell carcinomas
Authors: Bai Jian , Wang Qiyan , Gong Kenan , Cai Hong , Ke Yang , Zeng Changqing
Published: August 2014

Objective: Esophageal squamous cell carcinoma (ESCC) is a major cause of cancer related mortality. To determine the molecular basis of ESCC development, this investigation sought to comprehensively identify the characteristic genome-wide changes in ESCC, including exonic mutations and structural alterations. The clinical implications of these genetic changes are also analyzed.

Methods: Exome sequencing was applied to detect somatic mutations in nine pairs of ESCC and matched blood samples, followed by validation with MassARRAY genotyping and Sanger sequencing. Whole-genome SNP arrays were employed to study copy number changes and the loss of heterozygosity in 55 ESCCs.

Results: 108 non-synonymous mutations in 102 genes were identified and validated in nine patients, suggesting a low somatic mutation rate (average 12 per tumor) in our ESCC cases. The chromatin modification process was found to be enriched in gene ontology analysis. Mutated TP53 was seen in five of the nine tumors (55.6%) undergone exome sequencing and in 23/46 tumors in the validation set. Integrating with the SNP array data, we found that tumor genomes with TP53 mutations were much more unstable than those without these mutations. In the landscape of genomic alterations, deletion of 9p21.3 covering CDKN2A (30.9%), amplification of 11q13.3 covering CCND1 (30.9%), and point mutation of TP53 (50.9%) occurred in two-thirds of ESCCs, suggesting deregulation of G1 in cell cycle is the key event in ESCC. Furthermore, six minimal common regions were found to be significantly altered in ESCC and three (9p21.3, 7p11.2, 3p12.1) were associated with lymph node metastasis.

Conclusions: Frequent TP53 mutation and genomic instability are highly correlated in ESCC. The amplification of CCND1, deletion of CDKN2A/B, and somatic mutation of TP53 play pivotal roles via G1 deregulation and therefore classify this cancer into major groups. These findings provide clinical significance for future molecular diagnosis and therapeutic targeting.

Citation Format: Jian Bai, Qiyan Wang, Kenan Gong, Hong Cai, Yang Ke, Changqing Zeng. A comprehensive study of genomic alterations reveals deregulation of the cell cycle in most esophageal squamous cell carcinomas. [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 C03.
Abstract C22: Differential Tks5 isoform expression contributes to metastatic invasion of lung adenocarcinoma
Authors: Li Carman Man-Chung , Chen Guoan , Dayton Talya , Kim-Kiselak Caroline , Beer David , Winslow Monte , Jacks Tyler
Published: August 2014

Metastasis accounts for the vast majority of cancer related deaths, yet the molecular mechanisms that drive metastatic spread remain poorly understood. Here we report that Tks5, which has been linked to formation of proteolytic cellular protrusions known as invadopodia, undergoes an isoform switch during metastatic progression in a genetically-engineered mouse model of lung adenocarcinoma. Non-metastatic primary tumor-derived cells predominantly expressed a short isoform Tks5short, while metastatic primary tumor- and metastasis-derived cells acquired increased expression of the full-length isoform Tks5long. This elevation of Tks5long-to-Tks5short ratio correlated with a commensurate increase in invadopodia activity in metastatic cells compared to non-metastatic cells. Further characterization of these isoforms by knockdown and over-expression experiments demonstrated that Tks5long promoted invadopodia in vitro and increased metastasis in transplant models and an autochthonous model of lung adenocarcinoma. Conversely, Tks5short decreased invadopodia stability and proteolysis, acting as a natural dominant-negative inhibitor to Tks5long. Importantly, high Tks5long and low Tks5short expressions in human lung adenocarcinomas correlated with metastatic disease and predicted worse survival of early-stage patients. These data indicate that tipping the Tks5 isoform balance to a high Tks5long-to-Tks5short ratio promotes invadopodia-mediated invasion and metastasis.

Citation Format: Carman Man-Chung Li, Guoan Chen, Talya Dayton, Caroline Kim-Kiselak, David Beer, Monte Winslow, Tyler Jacks. Differential Tks5 isoform expression contributes to metastatic invasion of lung adenocarcinoma. [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 C22.
Abstract IA11: MicroRNAs in oncogenic and tumor suppressor signaling pathways
Authors: Mendell Joshua T.
Published: August 2014

MicroRNA (miRNA) gain- and loss-of-function can potently influence cellular behavior in normal physiologic states and in diseases such as cancer. The regulation of miRNA expression and activity by cellular signaling cascades can therefore result in dramatic phenotypic outputs. We previously demonstrated extensive control of miRNA expression by well-characterized oncogenic and tumor suppressor networks including the Myc, Kras, and p53 pathways. We are now employing novel mouse models with gain and loss of miRNA function to investigate the physiologic functions of the miRNAs embedded within these signaling pathways and the pathologic consequences when their functions are disrupted. Insights gained from these studies are revealing unanticipated roles for miRNAs in vivo and are informing our understanding of how dysregulated miRNA activity contributes to tumorigenesis. I will present our latest findings related to the regulation and function of mammalian miRNAs in normal physiology and in cancer and how these findings may be exploited for the development of novel therapeutic approaches.

Citation Format: Joshua T. Mendell. MicroRNAs in oncogenic and tumor suppressor signaling pathways. [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 IA11.
Abstract IA2: Somatic alterations in human cancer genomes
Authors: Meyerson Matthew
Published: August 2014

The advent of high-throughput genome sequence analysis has enabled systematic discovery of somatic genome alterations through genome, exome, and transcriptome sequencing of paired human cancer and germline nucleic acids. These discoveries have led to the identification of somatic genome alterations in new pathways including epigenetic regulators, RNA splicing factors, and immune response regulators, in addition to previously known signal transduction, cell cycle, and transcriptional pathways. I will discuss alterations in these pathways by copy number disruption, mutation, and re-arrangement, especially as seen through the lens of recent and ongoing large-scale studies of lung adenocarcinoma and squamous cell lung carcinoma genomes, from The Cancer Genome Atlas and beyond'and will discuss the implications for the understanding of cancer pathogenesis and for the development of targeted and immunomodulatory therapies.

Citation Format: Matthew Meyerson. Somatic alterations in human cancer genomes. [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 IA2.
Abstract IA27: Cell fate determinants as regulators of cancer metastasis
Authors: Kang Yibin
Published: August 2014

An emerging class of genes dictating malignant behavior is cell fate regulators. In the orderly development of a given tissue, the balance between cellular differentiation and division/migration is precisely controlled by a number of lineage specific transcription factors to maintain normal tissue architecture. Deregulation of such cell fate determinants have been increasingly linked to cancer metastasis, and have been shown to play critical roles in regulating cancer stem cell activity, epithelial-mesenchymal transition (EMT), tumor invasion and metastatic colonization.

In our study of the mammary gland development, we found that the transcription factor Elf5, a key regulator of alveologenesis in the mammary gland, also plays a critical role in regulating the stem/progenitor function of the mammary epithelium. Targeted deletion of Elf5 in the mammary glands leads to accumulation of cell types with dual luminal/basal properties such as coexpression of K8 and K14 and an increase in CD61(+) luminal progenitor population during pregnancy. Further interrogation suggests that the abnormal increase in K14(+) K8(+) cells may represent the CD61(+) luminal progenitors blocked in differentiation. Remarkably, Elf5 deficiency in mammary epithelium also triggers an increase of adult mammary stem activity as evidenced by the accumulation of mammary stem cell (MaSC)-enriched cell population in both pregnant and virgin mice and further confirmed by mammosphere and transplantation assays.

In addition to controlling cell fate determination, Elf5 also inhibits EMT in both mammary gland development and metastasis. This specific role for Elf5 in suppressing EMT and metastasis was uncovered via analyses of Elf5 conditional knockout animals, various in vitro and in vivo models of EMT and metastasis, an MMTV-neu transgenic model of mammary gland tumor progression, and clinical breast cancer samples. Furthermore, we demonstrate that this role of Elf5 is mediated by the direct transcriptional repression of Snail2/Slug, a master regulator of mammary stem cells and a known inducer of EMT. These findings establish a broad molecular function of Elf5 that extends from being a key cell lineage regulator during normal mammary gland development to serving as a suppressor of EMT and metastasis in breast cancer.

Citation Format: Yibin Kang. Cell fate determinants as regulators of cancer metastasis. [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 IA27.
Abstract A10: Stromal CAIX expression in oral squamous cell carcinoma and corresponding lymph node metastases
Authors: Chandarana Shamir , Bose Pinaki , Klimowicz Alexander C. , Petrillo Stephanie K. , Konno Mie , Rudmik Luke , Nakoneshny Steven C. , Matthews Wayne T. , Magliocco Anthony M. , Dort Joseph C. , Brockton Nigel T.
Published: August 2014

Background: Hypoxia is an almost ubiquitous feature of human solid tumours. However, the direct measurement of oxygen concentration in head and neck squamous cell carcinoma (HNSCC) has been conducted almost exclusively in the metastatic lymph nodes, not the primary tumor. Hypoxia is associated with resistance to both radiotherapy (RT) and chemotherapy in squamous cell cancers (SCCs). However, endogenous markers of hypoxia (EMH) have not reproducibly predicted clinical outcome. However, historically, the method used to assign hypoxic status according to directly measured oxygen concentration (polarographic electrode) and EMH (typically immunohistochemistry) have fundamentally differed.

Methods: Demographic data and clinical outcomes of 61 patients with oral squamous cell carcinoma (OSCC) treated with primary surgery +/- adjuvant radiation were collected. Tissue microarray's with representative samples of patients' tumors were constructed from paraffin-embedded blocks. Automated quantitative immunohistochemistry (AQUA) was performed to measure expression and distribution of carbonic anhydrase IX (CAIX). Kaplan-Meier survival analysis and the log-rank test was used to assess the impact of vimentin-defined stromal CAIX (in both primary tumor and nodal metastases) on survival.

Results: High stromal CAIX expression in the primary tumor was associated with significantly worse 5-year disease-specific survival. There was a statistically significant but relatively weak correlation between stromal CAIX expression in the primary tumor and the corresponding lymph nodes (r2:0.359; p=0.013). Stromal CAIX expression in the lymph node metastases was not significantly associated with 5-year disease-specific survival.

Conclusions: Previous literature presenting the prognostic impact of directly measured hypoxia, using a polarographic electrode, was based predominantly on measurements from lymph nodes, not the primary tumor. Furthermore, comparisons with EMH were based solely on percentage of tumor cells that were positively stained, not the intensity of staining. We present the results of a tumor comparment-specific, quantitative fluorescent IHC method and the survival associated with EMH in the tumor microenvironment in the primary tumor and lymph node metastases.

Citation Format: Shamir Chandarana, Pinaki Bose, Alexander C. Klimowicz, Stephanie K. Petrillo, Mie Konno, Luke Rudmik, Steven C. Nakoneshny, Wayne T. Matthews, Anthony M. Magliocco, Joseph C. Dort, Nigel T. Brockton. Stromal CAIX expression in oral squamous cell carcinoma and corresponding lymph node metastases. [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 A10.
Abstract A13: Interleukin-1β promotes brain metastasis of breast cancer by activating Notch signaling through reactive astrocytes
Authors: Xing Fei , Kobayashi Aya , Okuda Hiroshi , Watabe Misako , Pandey Puspa R. , Pai Sudha K. , Liu Yin , Wu Kerui , Watabe Kounosuke
Published: August 2014

Metastatic diseases are responsible for the majority of the deaths in breast cancer patients, and brain is one of the most common metastatic sites. The metastatic tumor in the brain profoundly affects the cognitive and sensory functions as well as morbidity of patients, and the one year survival rate among these patients remains less than 20%. However, little is known about the pathogenesis of brain metastasis, and therefore, it is of paramount importance to elucidate the molecular mechanism of metastatic process in order to define a specific therapeutic target. Our analysis of existing microarray database indicate that interleukin-1β (IL-1β) is highly expressed in MDA-MB231BrM2a (231BrM) cell line which is capable of preferentially metastasizing to the brain. IL-1β is known to have an ability of “activating” astrocytes that are often found in the metastatic niche, and therefore, it is speculated that the reactive astrocytes can promote growth and invasion of metastasized cells in the brain. To explore this possibility, we examined the effect of IL-1β and the conditioned medium of 231BrM cells on rat primary astrocytes in vitro. Our results of qRT-PCR and Western blot analysis indicate that IL-1β and the conditioned medium significantly up-regulated the Notch ligand, JAG1, in astrocytes, and that this up-regulation was abrogated by the NF-kB inhibitors, PDTC and RO 106-9920. We also found that the interaction between CSCs and astrocytes significantly enhances the self-renewal ability of CSCs. To further examine the interaction of astrocytes and tumor cells in the brain, we intracardially injected 231BrM cells in nude mice, and tumor growth was monitored by Xenogen bioimager. After 4 weeks, metastatic tumor growth was observed at multiple sites of the brain, and the results of immunohistochemical analysis indicated that astrocytes adjacent to the metastatic lesions strongly expressed JAG1 and the reactive astrocyte marker, GFAP (Glial fibrillary acidic protein). Finally, we found that a BBB-permeable Notch inhibitor was able to significantly suppress metastatic growth of breast cancer in the brain in our animal model. These results represent a novel paradigm for the understanding of how metastatic breast CSCs re-establish their niche for their self-renewal in a totally different microenvironment, which opens a new avenue to identify a novel and specific target for the brain metastatic disease.

Citation Format: Fei Xing, Aya Kobayashi, Hiroshi Okuda, Misako Watabe, Puspa R. Pandey, Sudha K. Pai, Yin Liu, Kerui Wu, Kounosuke Watabe. Interleukin-1β promotes brain metastasis of breast cancer by activating Notch signaling through reactive astrocytes. [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 A13.
Abstract A19: Ecrg4 downregulation in glioma enables transformed cells to escape immunosurveillance by tumor-associated macrophages/microglia
Authors: Lee Jisook , Dang Xitong , Borboa Alexandra , Coimbra Raul , Baird Andrew , Eliceiri Brian
Published: August 2014

Tumor-associated microglia generally promote tumor invasion, adopting an immunosuppressive phenotype that is distinct from their pro-inflammatory functions. Based on bioinformatic mining of novel sentinel genes encoding biologically active peptides, we have recently identified Ecrg4, as a novel secreted pro-inflammatory mediator of macrophage activation. While downregulation of Ecrg4 expression occurs in various tumor cells, in this study, we demonstrate that the biological activity of Ecrg4 in cancer functions through a paracrine mechanism, where tumor cell-exposed Ecrg4 acts upon tumor-associated microglia. In orthotopic glioma models, tumor cells expressing Ecrg4 induce proliferation and morphological changes in tumor-associated microglia. These Ecrg4-mediated effects on microglia correlate with a reduction in tumor burden and prolonged survival. Together, these results identify Ecrg4 as a novel mediator of glioma progression functioning through a paracrine mechanism that regulates the inflammatory state of tumor-associated microglia/macrophages.

Citation Format: Jisook Lee, Xitong Dang, Alexandra Borboa, Raul Coimbra, Andrew Baird, Brian Eliceiri. Ecrg4 downregulation in glioma enables transformed cells to escape immunosurveillance by tumor-associated macrophages/microglia. [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 A19.