bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract C47: 21-OH 6,19-epoxyprogesterone: A new dissociated glucocorticoid with anti-inflammatory properties but without antiapoptotic effects on chemotherapeutically treated mouse mammary tumor cells
Authors: Orqueda Andres J. , Veleiro Adriana , Joffé Elisa Bal de Kier , Kordon Edith , Burton Gerardo , Pecci Adali
Published: May 2012

Glucocorticoids (GCs) are widely used as coadyuvants in the treatment of solid tumors with chemotherapics such as paclitaxel (PXL) and doxorubicin (DOXO). However, clinical evidence supports the idea that GCs may induce resistance to these antitumor therapies. To improve cancer treatment quality, intense efforts are being made to design new dissociated steroids (steroid receptor glucocorticoid modulators (SRGMs). These compounds should mimic only the beneficial aspects of glucocorticoid therapy without inducing the undesired side effects in cancer patients. We have previously shown that dexamethasone (Dex), a well-known synthetic GC, inhibits PXL or DOXO induced apoptosis in breast tumor cells through a mechanism involving Bcl-XL and Bcl-2 induction. Here, our goal was to compare Dex effects with those triggered by 21-OH-6,19-epoxyprogesterone (21-OH-6,19-OP), which is a rigid steroid considered a passive GC antagonist. Molecular dynamic simulation studies predicted that 21-OH-6,19OP is unable to induce GR/21-OH-6,19-OP complex homodimerization and/or binding to coactivators, making these complex incapable to directly activate gene transcription (Alvarez et al J Med Chem. 2008. 13:1352). The effects of 21-OH-6,19-OP on apoptosis induced by DOXO or PXL in the epithelial mouse mammary tumor cell line LM3 was determined by analyzing Caspase-3 activity and gene and protein expression of antiapoptotic members of the Bcl family, i.e. Bcl-2 and Bcl-XL. The results show that, contrary to Dex, 21-OH-6,19-OP cannot reverse apoptosis induced by DOXO or PXL in LM3 cells. In fact, 21-OH-6,19-OP and Dex differentially regulate Bcl-2 and Bcl-XL expression. Dex's reversion on chemotherapic induced apoptosis (Dex+DOXO o Dex+PXL) correlates with an increase in Bcl-XL levels, while 21-O-H-6, 19-OP does not affect neither cell death nor Bcl-XL levels. The relevance of Bcl-XL involvement in DOXO resistance of cells treated with Dex was confirmed by knocking-down Bcl-XL with specific siRNA. Silencing Bcl-XL expression directly implicated loss of antiapoptotic GC effects. On the other hand, 21-OH-6,19-OP antiinflammatory action was also evaluated on the lung epithelial tumor cell line A549 by determining Interleukin-8 (IL-8) and Cyclooxygenase-2 (COX-2) expression in the presence of the Tumor Necrosis Factor-alpha (TNF-α). Expression levels of IL-8 and COX-2, measured by qRT-PCR, indicate that 21-OH-6,19-OP may have antiinflammatory effects as it reverses these inflammation markers expression induced by TNF-α in lung epithelial cells. Together, these findings suggest that 21-OH-6,19-OP is a SRGM that keeps glucocorticoid antiinflammatory properties without antiapoptotic effects, thus becoming a novel candidate for treatment of solid tumors as breast cancer.

Citation Information: Cancer Res 2009;69(23 Suppl):C47.
Abstract C48: Antiangiogenic properties of rare earth nanoparticles: Implications in ovarian cancer
Authors: Giri Shailendra , Rattan Ramandeep , Karakoti Ajay , Seal Sudeepta , Shridhar Vijayalakshmi
Published: May 2012

A substantial body of evidence suggests an association between vascular endothelial growth factor (VEGF) and the neovascularization that occurs with ovarian carcinogenesis. VEGF expression has been directly related to ascites accumulation. Phase II clinical trials in ovarian cancer using single-agent bevacizumab (a monoclonal antibody directed against VEGF) showed promising results, clearly highlighting the importance of anti-angiogenesis based-targeted therapies in ovarian cancer. One of the ways that VEGF mediates angiogenesis is by activating NADPH oxidase-ROS pathway. We have developed and engineered nanoparticles (NP) of cerium oxide (Nanoceria, NCe), a rare earth oxides, which have been tested for their ability to serve as free-radical scavengers to provide protection against chemical, biological, and radiological insults that promote the production of free radicals. Here we show for the first time that treatment of human umbilical vascular endothelial cells (HUVECs) with NCe resulted in inhibition of proliferation, capillary tube formation and VEGF165 induced activation of VEGFR2 and MMP2. Additionally, NCe treatment inhibited growth factor induced migration and invasion of ovarian cancer cells. Collectively, these results indicate that cerium oxide based NP could potentially be used as an antiangiogenic therapeutic agent in ovarian cancer.

Citation Information: Cancer Res 2009;69(23 Suppl):C48.
Abstract C49: AKT inhibitor has potent antitumor activity in human lung cancer xenograft models
Authors: Banu Naheed , Hiran Tej , Fadela K. Chafai , Menon Krishna
Published: May 2012

Lung cancer accounts for more than 215,000 new cases annually in the United States, and is the single largest cause of cancer death accounting for more than 130,000 deaths each year. Non-small cell lung cancer (NSCLC) accounts for 80% of all bronchogenic neoplasms with 90% of diagnosed patients dying within five years; therefore, new therapies are strongly desired. Effective therapies will come from a more detailed understanding of the molecular mechanisms underlying both the disease and the clinical response. We are developing a novel molecule, termed Kevetrin™, which was effective in reducing tumor growth in human lung cancer xenograft models while being well-tolerated. Kevetrin appeared to exert its anti-tumor activity by lowering levels of activated AKT also known as protein kinase B, of the AKT signal transduction pathway resulting in reduced tumor cell survival. Activated AKT was measured by a sandwich ELISA specific for phosphorylated AKT protein. Kevetrin was also evaluated for anti-tumor activity in vivo in the NCI-H1975 and A549 lung xenograft tumor models. Tumor-bearing nude mice were treated intravenously (IV) with either 200 mg/kg Kevetrin three times every other day or 22 mg/kg paclitaxel four times every other day. After Kevetrin was incubated with cells in vitro, phosphorylated AKT levels were reduced leading us to test for anti-tumor activity in vivo. Results with NCI-H1975 tumors showed that Kevetrin significantly delayed median tumor growth by 34 days (142%) in the first experiment and 28 days (156%) in a repeat experiment compared to controls, whereas the tumor growth delay (TGD) of paclitaxel was just 4 days (17%) and 14 days (78%), respectively. No weight loss occurred during treatment with Kevetrin. Results with A549 tumors showed that Kevetrin significantly delayed median tumor growth by 11 days (33%) in the first experiment and 30 days (111%) in a repeat experiment compared to controls, whereas the TGD of paclitaxel was just 0 days (0%) and 3 days (11%), respectively. In this study, Kevetrin resulted in <5% weight loss during treatment. These results showed that Kevetrin exerted potent anti-tumor activity against two human lung xenograft tumor models, NCI-H1975 and A549, at a dose and schedule that was well-tolerated as indicated by a small transient weight loss during treatment. NCI-H1975 have the T790M and L858R mutations in epidermal growth factor receptor (EGFR) and A549 have the K-ras mutation and overexpress STAT3 and Nrf2; these mutations are associated with resistance to standard chemotherapy. The reduction of activated AKT may be a mechanism by which Kevetrin overcame resistance and showed efficacy in these tumor models. These studies demonstrate that Kevetrin has promising potential for the treatment of lung carcinoma, particularly in cases where tumors have become resistant to standard chemotherapy.

Citation Information: Cancer Res 2009;69(23 Suppl):C49.
Abstract C5: Epigenetic regulation of cancer stem cell marker CD133 by transforming growth factor-β
Authors: You Hanning , Rountree C. Bart
Published: May 2012

Background: Hepatocellular carcinoma (HCC) is the third leading cause of cancer mortality worldwide. CD133, a trans-membrane glycoprotein, is an important cell surface marker for both stem cells and cancer stem cells in various tissues including liver. CD133 expression was recently linked to poor prognosis in HCC patients. CD133+ liver cancer cells are characterized by resistance to chemotherapy, self-renewal, multi-lineage potential, increased colony formation, and in-vivo cancer initiation at limited dilution. Recent studies demonstrate that CD133 expression is regulated by DNA methylation. In this paper, we explored the role of Transforming Growth Factor β (TGFβ), a multi-functional cytokine that plays a critical role in chronic liver injury, in the regulation of CD133 expression.

Materials and Methods: CD133- Huh7 cells were sorted from bulk cultured cells, CD133 expression was analyzed using flow cytometry, immuno-blots and real-time RT-PCR after the cells were treated with TGFβ1, DNA methyltransferase inhibitors and inhibitory Smads. Pyrosequencing was introduced to evaluate CD133 promoter-1 methylation status.

Results: TGFβ1 is capable of up-regulating CD133 expression specifically within the Huh7 HCC cell line in a time and dose-dependent manner. Furthermore, forced expression of inhibitory Smads, including Smad6 and Smad7, attenuated TGFβ1-induced CD133 expression. Within CD133-Huh7 cells, TGFβ1 stimulation inhibited the expression of DNA methyltransferases DNMT1 and DNMT3β, which are critical in the maintenance of regional DNA methylation; and global DNMT activity in CD133-Huh7 cells was inhibited by TGFβ1. DNMT3 inhibition by TGFβ1 was partially rescued with overexpression of inhibitory Smads. Lastly, TGFβ1 treatment led to significant demethylation in CD133 promoter-1 in CD133-Huh7 cells.

Conclusion: TGFβ1 is able to regulate CD133 expression through inhibition of DNMT1 and DNMT3 expression, and subsequent demethylation of promoter-1. The mechanism by which TGFβ induces CD133 expression is partially dependent on the Smads pathway.

Citation Information: Cancer Res 2009;69(23 Suppl):C5.
Abstract C57: The role of stromal cells in tumor metastases
Authors: Duyverman Annique , Duda Dan G. , Steller Ernst J.A. , Fukumura Dai , Jain Rakesh K.
Published: May 2012

Background: In 1889, Stephen Paget hypothesized that metastatic cells - the “seeds” - only grow in the secondary sites - the soil” - with a permissive microenvironment. Here, we show that the “seeds” can carry their own “soil” - activated fibroblasts from primary site of metastatic tumors. Moreover, we report that carryover of these non-malignant host-derived cells during metastatic cell colonization in the lungs can increase cancer's metastatic efficiency.

Materials and Methods: We use multiple animal models and two lung tumor cell lines (LLC1 and LA-P0297) to study the composition, origin and role of fragments shed by tumors in the metastatic process.

Results: We have shown that, in addition to single cells (81%), tumors shed fragments consisting of both tumor and host cells. We show that these fragments are more viable in the circulation compared to tumor cells alone (p<0.05).When we selectively deplete the host cells in the tumor microenvironment the spontaneous formation of metastases is significantly reduced three weeks after primary tumor resection compared to control mice (p<0.05). Next, survival was significantly increased when we deplete host-cells in mice with metastases after co-implantation of CAFs with LLC1, but had no effect in mice implanted with cancer cells alone (p<0.01) Lastly, we have assessed the nature of these host stromal cells using immunohistochemistry and RT-PCR and identified them being of fibroblast origin.

Discussion: We found that tumor-associated stromal cells can escape primary tumors along with cancer cells, survive in blood circulation and in metastatic nodules. Pre-existence of a tissue-like structure in the form of heterotypic cell fragments may increase the viability of cancer cells in blood circulation and modulate metastasic efficiency. How this mechanism contributes to the organo-tropism of certain metastatic tumors is not known, but demonstration of the involvement of tumor “soil” in both the primary tumor as well as the metastatic site, has important conceptual and clinical implications for colonization during metastasis progression.

Citation Information: Cancer Res 2009;69(23 Suppl):C57.
Abstract C6: Barrett's esophagus and esophageal adenocarcinoma: A paradigm for the cancer stem cell model?
Authors: Grotenhuis Brechtje A. , Dinjens Winand N.M. , Sonneveld Petra , Sacchetti Andrea , Van Dekken Herman , Wijnhoven Bas P.L. , Van Lanschot Jan J.B. , Fodde Riccardo
Published: May 2012

Introduction: In recent years, accumulating experimental evidence has suggested that tumors have a hierarchical organisation where only a subset of parenchymal cells, the cancer stem cells (CSCs), has tumor-initiating properties. To date, several antibodies directed against surface antigens have been employed to prospectively isolate CSCs from a broad spectrum of malignancies, though not from esophageal adenocarcinoma. Here, we conducted a series of experiments to test whether Barrett's esophagus and esophageal adenocarcinoma may serve as a disease model for the cancer stem cell concept.

Methods: We employed a panel of previously established CSC markers (CD24, CD29, CD44, CD133, CD166, EpCAM) for immunohistochemistry (IHC) analysis and in vivo transplantation assays. Transplantation of serially diluted bulk cancer cells of human tumors into NOD-SCID mice was performed to investigate the presence of CSCs in esophageal adenocarcinoma. Analogous assays were carried out by injecting esophageal cancer cells sorted by FACS using the same panel of CSC markers as employed for IHC analysis.

Results: IHC revealed that the intensity of CD24 and CD29 staining tended to increase along the metaplasia — dysplasia — carcinoma sequence. Cells positive for CD24 or CD29 were mainly located in the basal compartment of the metaplastic and dysplastic epithelium. CD44 showed membranous staining mainly in adenocarcinomas, whereas CD166 revealed pronounced cytoplasmic staining in all stages.

Serial transplantation experiments with bulk tumor cells indicated that the frequency of tumor-initiating cells in the Lin- population of esophageal adenocarcinoma is approximately 1:50,000 cells. However, to date no tumor growth was observed when tumor cells of subpopulations expressing the above-described CSC markers were transplanted at lower multiplicities.

Conclusions: Tumor-initiating cells in esophageal adenocarcinoma are rare, reflecting a hierarchal organisation. However, surface-antigens that have been established as CSC markers in other malignancies do not seem to enrich for tumor initiating cells in esophageal adenocarcinoma.

Citation Information: Cancer Res 2009;69(23 Suppl):C6.
Abstract C60: The integrative genomics viewer
Authors: Robinson James , Thorvaldsdottir Helga , Nazaire Marc-Danie , Winckler Wendy , Reich Michael , Getz Gad , Mesirov Jill
Published: May 2012

Rapid improvements in next generation sequencing and array-based profiling methods have created a paradigm shift in how scientists approach the study of the cancer genome. Projects such as The Cancer Genome Atlas (TCGA) are harnessing these advances to probe the cancer genome at unprecedented resolution, generating comprehensively catalogs of the genetic alterations of hundreds of tumors.

Visualization plays an important role in assimilating the large, muti-modal datasets produced by TCGA and similar projects. To be maximally effective tools must flexibly integrate genomic, functional, and phenotypic data from a variety of platforms, and efficiently handle large datasets. To address these critical challenges we developed the Integrative Genomics Viewer (IGV), a flexible, scalable, high-performance tool for the concurrent visualization of multiple large scale datasets. The IGV supports integrated views of any data type that can be mapped to genomic coordinates, including, but not limited to, next generation sequence data, copy number, gene expression, RNAi, ChIP-seq, RNA-seq, and genome annotations. Data can be annotated, filtered, grouped, and sorted according to clinical and phenotypic attributes.

A particular strength of IGV is its ability to handle very large datasets, for example deep coverage sequence data from whole genome scans, or highresolution copy number data from thousands of samples, on standard desktop computers. IGV permits the user to move seamlessly across these datasets at any level of detail, and quickly zoom out to whole genome or drill down to base pair resolution. The ability to dynamically and flexibly integrate multiple data modalities and view them at any scale allows the user to elucidate complex biological relationships that would not otherwise be apparent.

The IGV is freely available without restriction. It is currently under active development and use in The Cancer Genome Atlas, 1000 Genomes Project, and other large genomic projects.

Citation Information: Cancer Res 2009;69(23 Suppl):C60.
Abstract C61: Reducing cancer incidence and mortality in African American, Arab American, and Chaldean communities in the Detroit metropolitan area utilizing a community-based participatory process
Authors: Dykes Vicky A. , Akhtar Adil , Berry Stanley , Jaiyesimi Ishmael , Douglas-Nikitin Vonda , Shetty Sugandh , Vicini Frank A. , Fakhouri Monty
Published: May 2012

Purpose: We reviewed the impact of a community based participatory process (CBPP) addressing cancer education, prevention, and screening in two ethnic/minority populations to determine their impact on personal health related decisions.

Materials and Methods: From 2005 – 2008, 2,281 community members participated in CBPP outreach efforts (cancer education, prevention and screening forums) conducted by the Beaumont Cancer Institute in cooperation with the Arab American and Chaldean Council, the National Cancer Institute and the American Cancer Society. The study population consisted of 1,067 (46.7%) individuals completing a post-cancer forum survey: 642 (60.2%) from the African American (AA) and 425 (39.8%) from the Arab American & Chaldean (AAC) communities. Data were collected on behavioral intent and knowledge including: (1) forum content, (2) participants' cancer screening history, (3) symposium logistics (4) educational materials and (5) participation in subsequent cancer screening tests.

Results: After attending at least one cancer education forum, 329 (30.8%) of the 1,067 participant respondents (14.4% of all attendees) underwent some type of cancer screening: 108 women (10.1%) underwent a screening mammogram, and 94 (8.8%) a pap smear. Thirty-eight men underwent (3.6%) a screening PSA, and 89 (8.3%) had other forms of cancer screenings (e.g. colonoscopy). Overall, 60.2% of AA versus 35% of ACC attendees completed a post-survey and 19.6% of AA versus 10% of AAC attendees underwent some type of cancer screening.

Conclusions: The results of this study suggest that CBPP's based around perceived and/or serious diseases negatively affecting a target population can positively impact their personal health related decisions.

Citation Information: Cancer Res 2009;69(23 Suppl):C61.
Abstract C63: Structure of the Skp1-Fbx4 complex reveals a new class of the SCF ubiquitin ligase
Authors: Li Yunfeng , Hao Bing
Published: May 2012

Ubiquitin-mediated proteolysis governs the levels of many cellular proteins via a triple-enzyme cascade, involving ubiquitin-activating (E1), conjugating (E2) and ligating (E3) activities. The E3 ligases recruit both E2 and specific substrates and catalyze the final step in ubiquitination reaction. SCF complexes are the largest family of E3 ubiquitin-protein ligases that consist of Cul1, Rbx1, Skp1 and a member of the F-box protein family. F-box proteins are the substrate-recognition components of SCF; they interact with Skp1 through the N-terminal ∼40-residue F box motif, to recruit substrates through their variable C-terminal protein-protein interaction domains, including WD40 repeats (Fbxw subfamily), leucine-rich repeats (LRRs; Fbxl subfamily) and other different or unknown domains (Fbxo subfamily).

Fbx4, which is the F-box protein responsible for the recognition of cell-cycle regulator cyclin D1 and telomeric DNA-binding protein Pin2, belongs to the Fbxo family that lack recognizable protein-protein interaction motif in their C-terminus. Reduced Fbx4 expression appears to be responsible, at least in part, for overexpression of cyclin D1 in human cancers. On the other hand, overexpression of Fbx4 in human cells promotes degradation of endogenous Pin2 and results in progressive telomere elongation. In order to define the fundamental structural properties of Fbx4, we have determined the crystal structure of the human Fbx4 protein in complex with Skp1 by single-wavelength anomalous dispersion method. The structure of the binary complex reveals that Skp1 and the F-box domain of Fbx4 retain the same overall fold as in other Fbxl and Fbxw structures determined, and that binding of the F box to Skp1 is very similar to that in other Skp1-F box complexes. Interestingly, the C-terminal domain of Fbx4 is projected away from Skp1 and F box, to adopt an α/β fold with a seven-stranded β sheet surrounded by seven α helices, which is distinct from LRR (in Fbxl) and WD40 (in Fbxw), reinforcing the notion that Fbx4 represents a new family of substrate-recognition subunits in SCF E3s. Moreover, mutagenesis experiments have revealed a potential substrate-binding site in Fbx4 C-terminal domain. Taken together, our structural and biochemical data provide a framework for understanding the molecular mechanism of substrate recognition by the SCFFbx4 ubiquitin ligase.

Citation Information: Cancer Res 2009;69(23 Suppl):C63.
Abstract C65: Mechanisms to maintain extra centrosomes in cancer cells
Authors: Godinho Susana A. , Kwon Mijung , Ganem Neil J. , Pellman David
Published: May 2012

Multiple centrosomes in tumor cells create the potential for multipolar divisions that can lead to aneuploidy and cell death. Nevertheless, many cancer cells successfully divide because of mechanisms that suppress multipolar mitoses. Using a genome-wide RNAi screen in Drosophila S2 cells, we defined several mechanisms that suppress multipolar mitoses. We also found that HSET, a normally non-essential kinesin motor was essential for the viability of cancer cells containing extra centrosomes. Interestingly, using fibronectin micropatterns, we found that interphase cell shape and adhesion pattern can determine the success of the subsequent mitosis in cells with extra centrosomes. Thus, cell adhesion is an important morphological feature that contributes to centrosome clustering. Importantly, during tumor progression, changes in cell architecture and adhesion patterns, such as loss of E-cadherin and acquisition of an elongated cell shape, are often observed. This is referred to as the epithelial-to-mesenchymal transition (EMT). The presence of extra centrosomes is often correlated with more malignant tumors, which lost their epithelial phenotype most likely through EMT. Thus, we hypothesized that the ability of cells to cluster extra centrosomes varies between epithelial cells and non-epithelial cells. To test this idea we used a panel of non-transformed mammalian epithelial and non-epithelial cell lines and quantified their ability to cluster extra centrosomes. We treated cells with DCB to generate tetraploid cells containing extra centrosomes and followed them by live-cell imaging. We found that non-epithelial cells cluster their extra centrosomes much more efficiently than epithelial cells. This finding suggests that loss of cell-cell adhesion might facilitate centrosome clustering. Consistent with this idea, our preliminary data indicate that induction of EMT in the epithelial cell lines MCF10A and MDCK increases the number of tetraploid cells that undergo a bipolar mitosis. We propose that changes that take place during tumor progression, such as loss of cell-cell adhesion, might facilitate centrosome clustering and therefore increase the ability of cancer cells to maintain extra centrosomes. We are currently investigating the nature of the changes that occur during EMT that facilitate centrosome clustering.

Citation Information: Cancer Res 2009;69(23 Suppl):C65.
Abstract C67: Hypoxia-inducible factor-2α regulates macrophage functions in both acute and tumor inflammation
Authors: Imtiyaz Hongxia Zhang , Williams Emily P. , Hickey Michele M. , Patel Shetal A. , Durham Amy C. , Yuan Li-Jun , Keith Brian , Simon M. Celeste
Published: May 2012

Hypoxia and inflammation in the tumor microenvironment profoundly influence carcinogenesis and tumor progression. In order to connect hypoxic signaling to inflammatory responses and their roles in oncogenesis, we investigated how one of the master regulators of hypoxic hypoxia-inducible factor (HIF)-2α, adaptations, regulates macrophage functions in both acute inflammatory diseases and cancer (a disease involving various inflammatory components). Myeloid-specific deletion of HIF-2α results in defective expression of a panel of proinflammatory cytokines in macrophages responding to innate stimuli. Also, mice lacking myeloid HIF-2α are resistant to lipopolysaccharide-induced endotoxemia and display a marked inability to mount inflammation in response to several other acute irritants, revealing that this transcription factor is essential for macrophage inflammatory responses. Tumor-associated macrophages (TAMs) represent the major inflammatory cell type within tumors and HIF-2α is robustly expressed in these cells. Clinically, increased TAM HIF-2α level correlates with high tumor grade and poor prognosis in breast and uterine cervical cancer, respectively. To characterize roles of HIF-2α in TAMs, we induced hepatocellular carcinoma (HCC) and colitis-associated cancer (CAC), two established inflammation-associated tumor models, in mice lacking myeloid HIF-2α expression. Interestingly, we found that HIF-2α -deficient TAMs migrate less efficiently into tumors, leading to reduced tumor cell proliferation and decreased malignancy in mutant mice. Further studies revealed that the deficient macrophages were attributable to compromised migration capacity of HIF-2α to insufficient expression of M-CSF receptor, CXCR4 and extracellular fibronectin-1. Therefore, our study demonstrated HIF-2α as an important regulator in both acute inflammation and cancer, suggesting that the myeloid HIF-2α pathway could be exploited as therapeutic targets for treating these diseases.

Citation Information: Cancer Res 2009;69(23 Suppl):C67.
Abstract C68: Using a chemical genetic approach to define the role of CDK2 in normal and transformed cell lines
Authors: Huskey Noelle E. , Horiuchi Dai , Kusdra Leonard , Zhang Chao , Shokat Kevan , Goga Andrei
Published: May 2012

Cyclin dependent kinases (CDKs) drive progression through the eukaryotic cell cycle, making them an attractive target for cancer therapeutics. In particular, CDK2 is thought to regulate mammalian S-phase entry and progression, and frequently shows increased activity in a variety of tumors. Surprisingly, recent studies using conventional genetic approaches suggest that CDK2 is dispensable for cellular proliferation, both during normal development and in malignant cell lines. Here we use an alternative chemical genetic approach to determine whether acute ablation of CDK2 activity has the same affect as genetic deletion. We have replaced the endogenous CDK2 allele in mouse embryonic fibroblasts (MEFs) with a mutant analogue sensitive CDK2 allele, allowing us to selectively inactivate CDK2 using bulky ATP analogs. In contrast to genetically deleting CDK2, the acute inhibition of CDK2 resulted in severe defects in proliferation of non-transformed MEFs, as well as MEFs that were transformed using various oncogenes. These findings reveal a requirement for CDK2 in mammalian cell proliferation and suggest its potential as a therapeutic target in tumor cells.

Citation Information: Cancer Res 2009;69(23 Suppl):C68.
Abstract C69: Elastic properties of highly metastatic cells using nano-capillary wrinkling
Authors: Hurth Cedric , Pratap Divya , Yang Jianing , Zenhausern Frederic
Published: May 2012

We have adapted an existing emerging method to characterize elastic properties of thin (30–150 nm) polystyrene (PS) films based on the observation of the wrinkling patterns of the film (J. Huang et al, Science 2007, 317, 650–653). Our goal is to evaluate the stiffness of cell layers cultured on top of such polystyrene films when they enter metastasis as compared to healthy epithelial cells. Recent reports using atomic force microscopy (AFM) suggest differences in cell stiffness at the local single cell level (J.K. Gimzewski et al, Nature Nanotechnology 2007, 2, 780–783). We plan to measure similar properties when cells are arranged in a sheet and compare them to AFM measurements. A relevant system is the melanoma cell lines such as A 375 and SK-Mel28, for instance. We characterized the wrinkling patterns obtained using a stereomicroscope to provide a first straightforward assessment of the elastic properties by counting the number of wrinkles and measuring their average lengths. By comparing the wrinkling pattern when either healthy or melanoma cells are grown on top of the thin PS films to that obtained when bare thin PS film floats on water and developing a theoretical model based on 2 films of different but close Young moduli associated in parallel, one can extract quantities of interest to characterize the elastic properties of normal vs. cancer cells. In addition, we propose to use digital holographic microscopy (DHM) to improve the observation of the wrinkling patterns and allow access to other relevant viscoelastic information such as relaxation of the films. Because we propose an innovative technique coupled with an emerging form of microscopy yet unexplored for cancer research, i.e. DHM, to study the mechanical elastic forces within an interconnected assembly of metastatic cells with the goal of complementing current methods of clinical diagnostic, we hope to answer the current need for new ways of studying the mechanisms of cancer.

Citation Information: Cancer Res 2009;69(23 Suppl):C69.
Abstract C77: A mesenchymal-like subpopulation within squamous cell carcinomas with chemotherapy resistance and phenotypic plasticity
Authors: Basu Devraj , Nguyen Thierry-Thien , Montone Kathleen , Zhang Gao , Wang Li Ping , Rustgi Anil , Lee John , Weinstein Gregory , Herlyn Meenhard
Published: May 2012

A major barrier to curative therapy for most malignancies is intrinsic drug resistance within discrete tumor cell subpopulations, which are presently ill-defined. Previous studies show that carcinoma cell lines abundantly expressing mesenchymal markers have increased resistance to both conventional chemotherapy and epidermal growth factor receptor (EGFR)-targeted agents. However it is unclear whether small mesenchymal-like subpopulations contribute to therapy resistance in individual carcinomas with predominantly epithelial features. Here we identified a mesenchymallike subset expressing low E-cadherin (Ecad lo) and high vimentin (Vim hi) within human squamous cell carcinomas of the esophagus (ESCC) and head and neck (HNSCC). This minority subset was readily isolated from two HNSCC lines (SCC9 and OCTT2) and was identifiable in vivo, not only in mouse xenografts of these lines but also in the clinical specimen from which a HNSCC cell line (OCTT2) was derived. The minority Ecad lo/Vim hi subset contained more low-turnover cells and was less proliferative overall, correlating with increased resistance to the conventional chemotherapeutic agent paclitaxel. The Ecad lo/Vim hi subpopulation also demonstrated diminished activity in the MAP kinase/PI3-kinase pathways, likely arising from decreased EGFR expression and correlating with increased in vitro resistance to the EGFR-targeted agent, cetuximab. This mesenchymal-like subpopulation displayed comparable drug resistancein vivo, becoming enriched by cetuximab treatment in a mouse xenograft derived directly from a clinical HNSCC specimen. Furthermore, the Ecad lo/Vim hi and Ecad hi/Vim lo subsets were dynamic and reversible in their phenotypes, showing the capacity to repopulate each other from single cell clones. Taken together, these results show that a low-turnover, mesenchymal-like subpopulation in squamous cell carcinomas has diminished EGFR pathway function and elevated intrinsic resistance to both conventional and targeted chemotherapies. They further suggest that this mesenchymal-like subset retains the plasticity to repopulate other malignant subpopulations post therapy.

Citation Information: Cancer Res 2009;69(23 Suppl):C77.
Abstract C9: An in vitro model of osteoblastic bone metastasis in prostate cancer
Authors: Lescarbeau Reynald M. , Kaplan David L.
Published: May 2012

Prostate cancer progression is characterized by a striking affinity to spread to bone, where it forms metastases predominately of an osteoblastic phenotype, resulting in a significant source of morbidity. Currently, animal models for studying the development of these osteoblastic metastases are limited due to a lack of spontaneous osteoblastic metastasis development. In order to recapitulate osteoblastic metastasis development, an in vitro, three dimensional culture system was created utilizing silk fibroin scaffolds. LNCaP cells were seeded to silk scaffolds with human bone marrow derived mesenchymal stem cells (hMSCs) and cultured for 6 weeks. Examination after co-culture revealed through histological analysis that hMSCs underwent osteogenic differentiation in response to LNCaP paracrine signals. Paracrine interactions between hMSCs and LNCaPs were studied and IL-6, known to promote androgen independent growth, was found to be transiently expressed by hMSCs over the first two weeks of co-culture. In addition to secreted soluble signaling factors, the effect of extracellular matrix signals on the LNCaP cells was examined. LNCaP cells grown in a decellularized matrix, derived from hMSCs having undergone osteogenic differentiation, showed a 12 fold survival advantage in androgen depleted conditions over controls. This data indicates the ability of the co-culture system to stimulate differentiation of hMSCs towards an osteogenic linage and release soluble and extracellular matrix cues which stimulate androgen independent disease progression. The in vitro co-culture system presents a scalable and high throughput method for studying the molecular mechanisms underlying the osteoblastic component of prostate cancer bone metastasis.

Citation Information: Cancer Res 2009;69(23 Suppl):C9.
Abstract IA-1: From chromosome engineering to chromatin remodeler: CHD5 is a tumor suppressor mapping to human 1p36
Authors: Mills Alea A.
Published: May 2012

Deletions encompassing the 1p36 region of human chromosome 1 have been associated with a spectrum of malignancies, however the tumor suppressor had not been identified. In an effort to identify tumor suppressors within this interval, we used chromosome engineering to generate mouse models with loss- and gain of regions of the genome corresponding to human 1p36. This strategy allowed us to identify a 4.3 megabase interval with potent tumor suppressive activity. Whereas loss of this region caused a marked increase in cellular proliferation that culminated in cancer in vivo, gain of the same interval triggered excessive tumor suppression as observed by augmented cellular senescence in cultured cells and apoptosis in vivo. Using genetic and molecular analyses, we identified the chromatin remodeling protein Chromodomain, helicase, DNA-binding domain protein 5 (Chd5) as the tumor suppressor within the interval, and revealed that Chd5 functions as a master regulator that facilitates p16/Rb- and p19/p53-modulated pathways. Analyses of human gliomas revealed that CHD5 is frequently deleted, suggesting that CHD5 loss contributes to malignancy. Here I will present our current efforts aimed at elucidating the epigenetic role of CHD5 in tumor suppression.

Citation Information: Cancer Res 2009;69(23 Suppl):IA-1.
Abstract IA-12: Oncology drug discovery and development in the 21st century
Authors: Hait William N.
Published: May 2012

Oncology drug discovery and development is poised to enter an era of unparalleled growth. Based on an increasingly detailed understanding of human genetics and cancer biology, new targets have emerged that go beyond nucleic acids and microtubules to members of signal transduction pathways that are upstream of these fundamental cellular structures. In addition, an appreciation of the effects of the tumor microenvironment on cancer cell pathophysiology and metabolism has focused increasing attention on targets in these areas. With more than 200 compounds in the Pharmaceutical Industry's pipeline, substantial improvements in the lives of cancer patients are anticipated.

Yet major hurdles exist. Despite the increased knowledge and investment in drug development, the entry of new drugs into the clinic has not kept pace. Key barriers include but are not limited to: 1. Understanding the role of a drug target in the context of an individual's malignancy; 2. Predicting characteristics of a patient and his or her cancer that is most likely to benefit from a specific therapy; 3. Predicting the toxicity of new agents; 4. Outmoded clinical trial designs; 5. Developing rational drug combinations; 6. Accrual to clinical trials; 7. Appropriate resourcing of regulatory agencies and; 8. the price of developing cancer drugs.

This presentation will address each of these challenges and suggest ways in which progress can be made.

Citation Information: Cancer Res 2009;69(23 Suppl):IA-12.
Abstract IA-13: Mechanisms regulating anchorage independent survival
Authors: Schafer Zachary , Grassian Alex , Song Loling , Mailleux Arnaud , Schmelzle Tobias , Irie Yoko , Gao Grace , Overholtzer Michael , Jiang Zhenyang , Gerhart-Hines Zachary , Puigsever Pere , Brugge Joan S.
Published: May 2012

One of the earliest recognized features of tumor cells in culture is their ability to proliferate and survive without attachment to extracellular matrix. This ‘anchorage independence’ of tumor cells distinguishes their behavior from normal cells and is believed to be important to allow tumor cells to proliferate outside of their natural niches during early stages of tumor formation as well as during invasion and metastasis. My presentation will focus on findings from our laboratory on the mechanisms responsible for death of cells deprived of matrix and how oncogenes and other factors promote anchorage independence.

We have found that three distinct death processes — apoptosis, metabolic impairment, and a novel non-apoptotic mechanism, referred to as entosis, involving invasion of one cell within another and lysosomal degradation — contribute to death of cells deprived of their appropriate matrix. The major topic of my presentation will be new unpublished findings on metabolic pathways that are regulated by matrix attachment. We have found that detachment of mammary epithelial cells from ECM causes an ATP deficiency due to loss of glucose transport. Overexpression of ErbB2 rescues the ATP deficiency by restoring glucose uptake through PI(3)K activation. Interestingly, we found that the ATP deficiency could also be rescued by antioxidant treatment without rescue of glucose uptake. This restoration was found to be dependent on stimulation of fatty acid oxidation (FAO), which is inhibited by detachment-induced reactive oxygen species (ROS). The significance of these findings was supported by studies in 3D acinar cultures and anchorage-independent colony formation. These results reveal both the importance of matrix attachment in regulating metabolic activity and an unanticipated mechanism for cell survival in altered matrix environments through antioxidant restoration of ATP generation.

Citation Information: Cancer Res 2009;69(23 Suppl):IA-13.
Abstract IA-22: Cellular responses to telomerase perturbations
Authors: Li S. , Matsuguchi T. , Stohr B.A. , Xu L. , Perry K. , Ren B. , Blackburn Elizabeth H.
Published: May 2012

Telomerase is a ribonucleoprotein that replenishes the DNA at telomeres, thereby counteracting telomere shortening. Telomerase is highly active in many human malignancies. Telomerase functions in cancer maintenance include replenishing telomeric DNA and maintaining cell immortality. The telomerase RNA component provides a short template that is copied to make telomeric DNA. We found that mutating the template to cause various aberrant telomeric sequences to be added in human cancer cells rapidly caused massive ATM-dependent and ATM-independent chromosome fusions and cell death.

We also showed previously that human cancer cells responded rapidly to abrupt depletion of telomerase RNA by rapid and distinctive cellular/transcriptional responses, despite no obvious bulk telomere shortening or loss of telomere integrity. This and evidence from others indicates that, in addition to its telomere-elongating role, telomerase may play other roles in cells: cells lacking or overexpressing telomerase show a variety of responses even in the absence of detectable effects on telomere integrity or functionality. We are performing ChIP—SEQ experiments to identify genomic sites of telomerase interaction. New results of experiments exploring these roles of telomerase in cancer will be presented. Together, these and other findings indicate that telomerase, both via its roles in telomere maintenance and other roles, provides potential new inroads into cancer therapies.

Citation Information: Cancer Res 2009;69(23 Suppl):IA-22.
Abstract IA-3: The regulation of p53 tumor suppressing activities
Authors: Lozano Guillermina
Published: May 2012

The p53 tumor suppressor is often altered in tumors via single nucleotide changes that alter critical amino acids in the DNA binding domain. These p53 mutants lose DNA binding activity and some mutants exhibit a gain-of-function phenotype that is manifested as metastasis in vivo. We have discovered that p53 mutants are normally unstable similar to wild type p53, and become stable in some tumors leading to the increased metastasis. Mdm2, a negative regulator of wild type p53 also negatively regulates mutant p53. Decreased Mdm2 gene dosage delays a cancer phenotype in tumor prone mouse models. In humans, an Mdm2 single nucleotide polymorphism (SNP) increases the levels of MDM2 and correlates with an earlier age of tumor onset. We have generated mice with the human MDM2 SNP and show that the SNP accelerates a tumor phenotype in mice.

Citation Information: Cancer Res 2009;69(23 Suppl):IA-3.
Abstract IA-5: Dissecting tumor suppressor gene networks in vivo
Authors: Lowe Scott W.
Published: May 2012

Cancer arises through an evolutionary process whereby normal cells acquire mutations that erode growth controls, leading to the inappropriate expansion of aberrantly proliferating cells. Such mutations can involve activation of oncogenes or inactivation of tumor-suppressor genes, each contributing one or more new capabilities to the developing cancer cell. However, cancer is not an inevitable consequence of oncogenic mutations; instead, cells acquiring such mutations can be eliminated or kept in check by innate tumor-suppressor programs that can be activated in these damaged cells. Our laboratory studies tumor-suppressor networks controlling apoptosis and senescence and how their disruption influences malignant behavior. We previously showed that apoptosis and cellular senescence are potent barriers to oncogene-driven tumorigenesis and that each contributes to the antitumor action of many chemotherapeutic drugs. Thus, not only do mutations that disrupt apoptosis and senescence promote tumor progression but, depending on the particular lesion, they can also reduce the efficacy of cancer therapy. To facilitate our research, we have recently developed new mouse cancer models based on the genetic manipulation of stem and progenitor cells ex vivo followed by transplantation of the altered cells into the appropriate organ of syngeneic recipient mice. This approach allows us to rapidly study the impact of many genes and gene combinations on tumorigenesis in a “mosaic” setting where tumor-initiating cells are embedded in normal tissues. Furthermore, we have developed powerful methods for using RNAi to suppress gene function in vivo in either a stable and reversible manner. Current efforts in the laboratory strive to integrate mosaic mouse models, RNA interference, and cancer genomics to identify new components of tumor suppressor gene networks and characterize their impact on tumorigenesis and treatment response. In addition, we are developing new RNAi interference methods to explore the role of tumor suppressor genes in tumor maintenance, and the cell death mechanisms involved in tumor regression.

Citation Information: Cancer Res 2009;69(23 Suppl):IA-5.
Abstract IA-6: Single nucleotide polymorphisms in the p53 pathway
Authors: Levine Arnold J.
Published: May 2012

The p53 protein and the signal transduction pathway controlled by that protein responds to a wide variety of stress signals which can disrupt the fidelity of DNA replication and cell division. To prevent these errors or mutations the p53 pathway responds by initiating a process of cell cycle arrest, senescence or apoptosis which either permits repair of these errors or kills the clone of cells that contain these mutations. Central to this pathway are a series of proteins that respond to the stress signals and regulate the levels and activity of the p53 pathway. We have identified a number of single nucleotide polymorphisms (SNP) in those genes that regulate p53 activity and functions and these SNPs can play a role in the incidence of cancers in a population, the age of onset of cancers and the response to therapy. Some of the haplotypes containing these SNPs appear to be under positive evolutionary selection pressures in some human populations. The reason for this appears to be the role of p53 in the implantation of embryos into the uterus and the impact of some of these SNPs upon the fecundity of mice and humans. This process is mediated by the p53 regulated gene, Leukemia Inhibitory Factor or LIF, a cytokine that is essential for the implantation of embryos. SNPs in the p53 gene, the MDM-4 gene and the LIF gene regulate the efficiency of implantation of embryos in humans. It could well be that the p53 protein is also involved in the surveillance of developmental abnormalities.

Citation Information: Cancer Res 2009;69(23 Suppl):IA-6.
Abstract IA-7: Fragment-based cancer drug discovery
Authors: Fesik Stephen W.
Published: May 2012

Fragment-based approaches offer several advantages over traditional methods for discovering new drug molecules. The concepts of fragment-based methods for lead identification and optimization will be presented and applications towards drug discovery will be discussed.

Citation Information: Cancer Res 2009;69(23 Suppl):IA-7.