bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract B70: Conexic: A Bayesian framework to detect drivers and their function uncovers an endosomal signature in melanoma
Authors: Akavia Uri-David , Litvin Oren , Kim Jessica , Mozes Eyal , Kotliar Dylan , Tzur Yossi , Garraway Levi , Pe'er Dana
Published: May 2012

Genomics is revolutionizing our understanding of cancer biology. Tumor samples assayed for comprehensive chromosomal and gene expression data are accumulating at a staggering rate. A major challenge involves the development of analysis methods to uncover biological insights from these data, including the identification of the key mutations that drive cancer and how these events alter cellular regulation.

We have developed Conexic, a novel computational framework to integrate chromosomal copy number and gene expression data to detect genetic alterations in tumors that drive proliferation, and to model how these alterations perturb normal cell growth/survival. The underlying assumption to our approach is that significantly recurring copy number change, coinciding with its ability to predict the expression patterns varying across tumors, strengthens the evidence of a gene's causative role in cancer. This method not only pinpoints specific regulators within an a large region of copy number variation, but can shed light on the way in which gene regulation is altered

We applyed our Conexic framework to a melanoma dataset (Lin et al, Cancer Research, 2007) comprising 65 paired measurements of gene expression and copy number, with interesting results. Our analysis correctly identified many known ‘driver’ events, while also connecting these to many of their known targets (e.g. MITF). Our global integrative analysis reveals insight into how the drivers alter transcriptional programs. An interesting recurring characteristic is that there are a number of different ways by which drivers can be altered; e.g., the expression of a driver may be altered through copy number variation or other mechanisms, but its influence downstream remains the same.

In addition to confirming the role of known drivers in melanoma, our analysis suggests a number of novel drivers. Most strikingly, these point to a major role in regulation of protein trafficking and endosome biology in this malignancy. These results have linked endosomal processing and sorting to adhesion and survival. Preliminary experimental validation supports three novel drivers including TBC1D16, RAB7A and RAB27A. Together, these results affirm the potential of Conexic to elaborate novel driver modules with biological and possibly therapeutic importance in melanoma and other cancers.

Citation Information: Cancer Res 2009;69(23 Suppl):B70.
Abstract B71: A systems biology approach to study Ewing's sarcoma
Authors: Surdez Didier , Stoll Gautier , Tirode Franck , Laud Karine , Zinovyev Andrei , Barillot Emmanuel , Delattre Olivier
Published: May 2012

Ewing's sarcoma is the second most frequent pediatric bone tumor. It is characterized in 85% of cases by a chromosomal translocation which leads to the expression of a chimeric transcription factor: EWS-FLI1. This oncogene is transforming in various cell and animal models. Although important EWS-FLI1 transcription targets and altered pathways have been described, a comprehensive understanding of the disease remains to be achieved. For this purpose, a systems biology approach was used. In this project, gene profiling of (1) primary tumors and (2) of EWS-FLI1-inducible cell models were performed. Based on these experimental data and on literature knowledge, we constructed an annotated influence network of EWS-FLI1 effects that was validated using small scale silencing/qPCR experiments and home-made software. In a second step, in order to better understand the global impact of EWS-FLI1 in Ewing sarcoma context, EWS-FLI1 ChIP-seq experiments were performed. These results were combined to the gene expression profile data using rational approaches and/or PCA based analyses. This resulted in a genome wide analysis in which the genes were ranked upon their implication/significance in Ewing sarcoma. In the future, we intend to integrate these results to an extended version of our annotated influence network and confront high-throughput experimental results (siRNA screen) with predictive computational models of this network. This should allow identifying novel crucial genes/pathways in Ewing's sarcoma.

Citation Information: Cancer Res 2009;69(23 Suppl):B71.
Abstract B75: TWISTing Stemness, Inflammation, and Proliferation of Epithelial Ovarian Cancer Cells through MIR199A2/214
Authors: Yin Gang , Chen Rui , Fu Han-Hsuan , Alvero Ayesha , Glackin Carlotta , Mor Gil
Published: May 2012

Introduction: Twist1, a basic helix-loop-helix (bHLH) transcription factor, has an important role in tumor biology by regulating epithelial-mesenchymal transition. It also functions as a transcription repressor in NFκB-dependent cytokine expression. Recently, we reported the identification and characterization of epithelial ovarian cancer (EOC) cells with stem-like properties (Type I EOC cells). At the microRNA level, Type I EOC cells are characterized by low levels of hsa-miR-199a2 and hsa-miR-214. We also demonstrated the capacity of Type I EOC cells to differentiate into mature ovarian cancer cells (Type II EOC cells), which has lost stemness potential and express high levels of both hsa-miR-199a2 and hsa-miR-214. We also showed previously that hsa-miR-199a2 is able to regulate the levels of IKKβ and therefore have a direct effect on the NFκB pathway. Still, another group reported the regulatory control of hsa-miR-214 on the Akt pathway. In this study, we show that Twist1 is able to regulate the miR-199a2/214 cluster in EOC cells and can therefore control both NFκB and Akt pathways.

Methods: 5′-RACE and 3′-RACE was used to clone the full length pre-miR-199a gene. Real-time PCR was used to determine the levels of Twist1 and miR-199a2/214. Twist1 was knocked-down in Type II EOC cells using siRNA. The effect of Twist1 knockdown on levels of IKKβ was determined by Western blot analysis. Cytokines were quantified using Luminex technology.

Results: Characterization of full length of pre-miR-199a2 transcript reveals that the MIR199A2 gene contains a human microRNA cluster, miR-199a2/214, and pre-miR-199a2 within the human Dnm3os gene (NCBI GeneID 474332). PCR analysis showed low levels of Twist1 and miR-199a2/214 in Type I EOC cells but high levels of expression in Type II EOC cells. Knockdown of Twist1 in Type II cells induce a significant decrease in the levels both hsa-miR-199a2 and hsa-miR-214, and significant increase in IKKβ expression. Ectopic expression of hsa-miR-199a2 partially reverse the effect of Twist-1 knockdown on the levels of IKKβ. The combination of knockdown Twist1 and TNFα stimulation in Type II EOC cells significantly increase Rantes, an NFκB-dependent cytokine.

Conclusion: We demonstrate for the first time that Twist1 plays a feedback role for the NFkB pathway by repressing IKKβ expression through hsa-miR-199a2. Twist1 inhibition of IKKβ expression through regulating hsa-miR-199a2, represents a novel negative feedback loop for the NFκB pathway. The demonstration that Twist1 can regulate the miR-199a2/214 cluster and therefore both the IKKβ and Akt survival pathways, suggests the potential role of Twist1 in EOC differentiation. Furthermore it opens the opportunity to develop new approaches for targeting the ovarian cancer stem cells.

Citation Information: Cancer Res 2009;69(23 Suppl):B75.
Abstract B9: Arm1: A glutamate methyltransferase capable of methyl esterifying PCNA
Authors: Hoelz Derek J. , Albert Alexandra , Dobrolecki Lacey , Arnold Randy , Novotny Milos , Schnaper Lauren , Hickey Robert , Malkas Linda
Published: May 2012

Recently we reported on a novel post-translational modification (PTM) present on proliferating cell nuclear antigen (PCNA). This PTM (methyl esterification) was found acidic residues, primarily glutamic acids. Interestingly, methyl esterification of glutamic acids in proteins has only been observed in bacteria where a specialized glutamate carboxyl O-methyltransferase, CheR, has been shown to methyl esterify specific glutamic acid residues of receptors during chemotaxis signaling. In eukaryotic cells, however, only three carboxyl O-methyltransferases are known to exist and none of them have specificity for glutamic acid residues. Here we describe the isolation and identification of a protein encoded by an unknown open reading frame and expressed in human cells that may represent the first eukaryotic glutamate carboxyl O-methyltransferase, which can modify PCNA. Sequence alignment of this protein with other protein carboxyl O-methyltransferases has demonstrated the existence of multiple consensus and conserved regions, and secondary structural predictions confirm a potential to form a SAM-dependent methyltransferase fold in its C-terminus. We therefore named this protein acidic residue methyltransferase 1 (Arm1). The biological significance of this discovery and its potential role(s) in DNA replication and repair will be discussed.

Citation Information: Cancer Res 2009;69(23 Suppl):B9.
Abstract C29: Periostin, a regulator of adhesion, is a novel effector of tumor invasion
Authors: Wong Gabrielle , Michaylira Carmen Z. , Miller Charles G. , Gutierrez Christie M. , Stairs Douglas B. , Kalabis Jiri , Nakagawa Hiroshi , Diehl Alan J. , Herlyn Meenhard , Rustgi Anil K.
Published: May 2012

Squamous cell cancers in aggregate comprise the most common forms of epithelial cancers, arising from the skin, head/neck, oral cavity, esophagus, lung and anogenital tract, and they share a number of environmental exposures and genomic alterations. A prototype of this cancer is esophageal squamous cell carcinoma (ESCC), which is associated with a high mortality rate worldwide. Local invasion and metastasis are important factors which influence patient survival. Amongst the frequent oncogene alterations are EGFR and cyclin D1 overexpression, and that in tumor suppressor genes are p53 mutation and p120-catenin loss or mislocalization. However, the functional interplay between these genetic alterations contributing to cancer progression remains unclear. Our previous studies have addressed this by presenting a tumor microenvironment model for tumor cell invasion using a 3D organotypic culture system and transformed human esophageal cells overexpressing EGFR and mutant p53 (Genes and Development 2007). From this, we have now utilized laser capture microdissection (LCM) and microarray analysis to identify novel targets of EGFR overexpression and mutant p53 by comparing gene expression of invading versus non-invading transformed cells. Results from these studies reveal a marked upregulation of periostin, a gene involved in cell adhesion, whose expression is restricted to invading tumor cells, as validated by qPCR, immunohistochemistry and Western blot. Furthermore, its expression was found to be upregulated in human esophageal cancer compared to normal esophagus. Using genetic approaches, gain-of-function of periostin fosters tumor cell invasion, whereas silencing of periostin markedly attenuates tumor cell invasion. Bioinformatic analyses reveal that periostin is upregulated in other human squamous cell cancers as well. Promoter studies assessing the transcriptional regulation of periostin upon EGFR overexpression and p53 mutation have been done. Our novel findings indicate periostin induction upon convergence of EGFR signaling and p53 mutation provides a cue for tumor cell invasion in the tumor microenvironment, and that it may be part of tumor invasion signature in squamous cell cancers in general.

This work was supported by P01-CA098101, NIH/NRSA F32-DK075230 and T32 CA115299.

Citation Information: Cancer Res 2009;69(23 Suppl):C29.
Abstract C32: Proteome analysis of pancreatic cancer tissue for identification of novel biomarkers
Authors: Ganepola Ganepola , Suman Paritosh , Gelber Cohava
Published: May 2012

Pancreatic cancer is the 4th largest cancer in US with mortality rate almost equal to incidence. The tumor usually presents at a late stage ruling out any curative surgery. Focus of pancreatic cancer research currently is mainly on early diagnosis and development of novel cancer therapeutics. Identifying sensitive, specific and reliable surrogate markers by genomic and proteomic methods1 in cancer tissue, blood or other bodily secretions can prove promising to reach this goal.

We are applying standard proteomic techniques like gel electrophoresis and MS (mass spectrometry) with a goal to identify potential biomarkers and new drug targets for the design and development of effective therapeutics in serum and pancreatic tissue samples.

Pancreatic cytosolic and membrane proteins were extracted from the 4 pairs of cancerous and adjacent normal tissue. Cytosolic protein fractions were run on PF-2D, traditional 1D and 2D SDS-PAGE. LC/MS/MS was used to identify differently expressed banding patterns. Multiple proteins were found to be either up (24) or downregulated (11). Compared to cytosolic proteins membrane proteins are priced for its accessibility on tumor tissue making these ideal for diagnosis and therapeutics. Isolating only plasma membrane proteins is very challenging due to low extraction efficiency, extremely low abundancy and poor solubility. Total cellular membrane protein was extracted and analyzed by SDS-PAGE and LC/MS/MS, and significant banding pattern differences were observed with 35 upregulated and 14 downregulated proteins with annexin 1, annexin A2 and annexin 5, tumor rejection antigen 1, prolyl 4-hydroxylase, calreticulin precursor, heat shock 70kDa protein 5, chaperonin and alpha 2 actin being the most significantly overexpressed. Findings were confirmed by probing western blots with anti-annexin 1 and 2 antibodies. Our findings suggest these differently expressed proteins can be explored as potential biomarkers after further validation. In contrast, analysis of serum samples by same techniques was futile due to exceedingly low abundance of protein and peptide fraction containing potential biomarkers due to masking by albumin and globulins as well as enzymatic degradation.

At present we are continuing the validation of our preliminary findings on larger sample size by using differential protein profiling technologies, immunohistochemistry and quantitative proteomics. Also we are planning to use affinity bait molecule with nanoparticles2 which can help overcome the limitations faced with serum samples in identification of de novo biomarkers.

Citation Information: Cancer Res 2009;69(23 Suppl):C32.
Abstract C71: Inhibition of polyisoprenylated methylated protein methylesterase by anticancer drugs
Authors: Schodlbauer Suzanne , Duverna Randolph , Lamango Nazarius S.
Published: May 2012

Polyisoprenylated methylated protein methylesterase (PMPMEase) is a little explored enzyme of the polyisoprenylation (isoprenylation) pathway (PP). Polyisoprenylation is a post-translational modification that is important for the function of such proteins as the heterotrimeric G proteins, Ras and Rab families of proteins. PMPMEase hydrolyzes a methyl ester formed by the S-adenosyl-L-methionine (SAM)-dependent enzyme, polyisoprenylated protein methyltransferase (PPMTase). PMPMEase's regulatory role is suggested by the fact it hydrolyzes the ester products of PPMTase. PMPMEase is a serine hydrolase, a class of hydrolytic enzymes that attack electron-deficient carbons of amide and ester bonds with the alkoxide ion of their catalytic serines. Because alkylating agents function through an aziridinium ion with an electron-deficient carbon atom, it is hypothesized that they may inhibit PMPMEase through active site adduct formation. Alkylating agents have been suspected of having other mechanisms of action against cells besides impeding replication via the cross-linking of DNA strands. The crystal structure of the human form of PMPMEase, human carboxylesterase 1, with tamoxifen bound to its active site has been solved, also suggesting its possible inhibition. Polyisoprenylated proteins are involved in processes that regulate cell viability. Mutations and overexpression of such proteins are implicated in about 30% of cancers. This study considers the possibility that the agents also reduce malignant cell growth by disrupting growth signaling pathways. Cyclophosphamide is an organophosphorous alkylating agent. Organophosphorous compounds irreversibly inhibit PMPMEase and other serine esterases. Busulfan, cyclophosphamide, mechlorethamine and tamoxifen were tested for porcine liver PMPMEase inhibition. At 1 mM concentrations, cyclophosphamide, mechlorethamine and tamoxifen inhibited 62, 35 and 55% of the PMPMEase activity. The results show that the anticancer drugs may owe part of their pharmacological activity to their modulation of oncogenic polyisoprenylated protein function through their inhibition of PMPMEase.

Citation Information: Cancer Res 2009;69(23 Suppl):C71.
Abstract C75: Granulin-expressing bone marrow cells promote the outgrowth of indolent tumors
Authors: McAllister Sandra S. , Scheel Christina , Nilsson Björn , Gifford Ann M. , Reinhardt Ferenc , Bray Mark , Carpenter Ann E. , Ebert Benjamin L. , Weinberg Robert A.
Published: May 2012

We previously demonstrated that certain vigorously growing xenografted human carcinomas (“instigators”) stimulate the growth of otherwise-indolent carcinoma cells and metastases (“responders”) implanted at distant anatomical sites — a process we termed “systemic instigation” (S. McAllister, et al.; Cell, 2008). We showed that systemic instigation is largely the result of activation of bone marrow cells (BMCs) that subsequently contribute to the stroma of the responding tumors; however, the identity of the activated BMCs and their contribution to outgrowth of the once-indolent cells was unknown. Here, we demonstrate that Sca1+/cKit− BMCs of hosts bearing instigating tumors are activated prior to their mobilization into the circulation and are very potent in their ability to promote outgrowth of the responding tumors. Strikingly, responding tumors, which result from either the presence of a contralateral instigating tumor or the admixture of Sca1+/cKit− BMCs harvested from instigator-bearing mice, form with a desmoplastic stroma. Stromal desmoplasia, the accumulation of a myofibroblast-rich reactive connective tissue, is almost always observed in malignant human adenocarcinomas. The Sca1+/cKit− BMCs do not directly give rise to stromal myofibroblasts; instead, we found that factors secreted by these BMCs induce fibroblasts to express alpha-smooth muscle actin (ACTA2), a myofibroblast marker. Gene expression profiling of the Sca1+/cKit− BMCs from instigator- and non-instigator-bearing mice identified granulin, a pluripotent secreted growth factor, as the most highly upregulated gene in the activated BMCs. We found: i) that granulin is indeed expressed by bone marrow-derived cells that are recruited into the responding tumor stroma; ii) that treatment of human mammary fibroblasts with granulin induces ACTA2 expression; iii) that granulin treatment is sufficient to enhance responding tumor growth in vivo. Mining of existing microarray datasets revealed that high granulin expression is correlated with the expression of ACTA2 in human breast cancers and is associated with shorter disease-free survival of breast cancer patients. Our results indicate that the formation of desmoplastic stroma, a condition that is associated with invasive carcinomas, can be instigated systemically by certain tumors via activation of granulin-expressing Sca1+/cKit− in the bone marrow.

Citation Information: Cancer Res 2009;69(23 Suppl):C75.
Abstract IA-14: Molecular alterations that predict premalignancy in breast cancer
Authors: Tlsty Thea D.
Published: May 2012

Approximately 15–30% of women diagnosed with ductal carcinoma in situ (DCIS) develop a subsequent tumor event within 10 years after surgical lumpectomy. To date, little is known about the molecular pathways that confer this differential risk for developing subsequent disease. Since DCIS is non-obligate precursor of invasive carcinoma, the majority of DCIS lesions are not associated with subsequent invasive tumors. Likely, many women diagnosed with DCIS who opt for surgery, with or without adjuvant treatment, are being overtreated. Identification of characteristics of DCIS that predict subsequent tumor development would advance our understanding of breast cancer and allow us to stratify a woman's individual risk for subsequent invasive tumors and avoid over- and undertreatment of women with DCIS.

To address this clinical problem, we have recently identified biomarkers in DCIS biopsies that accurately predict future formation of tumors, years before it actually occurs. The overexpression of biological markers of stress activation, p16 and COX-2, reflect two biologically different processes that predict women with a better or worse prognosis. In the absence of proliferation, expression of these markers indicates activation of a senescence response and a low probability of tumor formation. In the presence of proliferation, these markers indicate abrogation of the pRb tumor suppressor pathway and are associated with a high likelihood of future tumor formation. We initially tested these markers in a pilot set of DCIS samples (32 cases and 38 controls) obtained from an extensively annotated population-based cohort of women diagnosed with DCIS, treated by lumpectomy alone, and followed for twenty years. These markers identified a highly aggressive subtype of DCIS that shares characteristics with basal-like invasive tumors. We have extended these studies to a larger retrospective cohort, 162 samples from the DCIS cohort (75 cases and 87 controls) and find that the prediction markers faithfully identify women at high and low risk for future tumor formation. Independent validation in a large retrospective cohort is ongoing. The finding that DCIS lesions with basal-like molecular characteristics have an increased risk of developing subsequent cancers suggest this phenotype can be assayed far in advance of the development of invasive disease and present clinical opportunities.

Citation Information: Cancer Res 2009;69(23 Suppl):IA-14.
Abstract IA-18: Mouse embryonic stem cells exist in a unique metabolic state
Authors: McKnight Steven L.
Published: May 2012

The cell division cycle of mouse embryonic stem (ES) cells is considerably more rapid than that of even the most aggressively growing cancer cells. In order to investigate whether mouse ES cells might exist in a unique metabolic state supportive of rapid growth, organic extracts were prepared and analyzed by liquid chromatography/mass spectrometry. Upon comparison of mouse ES cells with differentiating embryoid bodies, such assays revealed profound differences in the abundance of metabolites associated with one-carbon metabolism and purine biosynthesis. It was also noted that the abundance of both threonine and acetyl-CoA changed markedly as ES cells were cued to differentiate into embryoid bodies. These differences in metabolite abundance were traced to the copious, ES cell specific expression of the gene encoding threonine dehydrogenase (TDH). TDH gene expression was 1,000-fold higher in ES cells than any of 7 tissues assayed from adult mice. The TDH enzyme catalyzes the mitochondrial breakdown of threonine into glycine and acetyl-CoA. Glycine is further catabolized by mitochondrial enzymes associated with the glycine cleavage system to foster the charging (methylation) of tetrahydrofolate, a key intermediate in purine synthesis. The acetyl-CoA produced from TDH-mediated breakdown of threonine is fed into the TCA cycle. This TDH-dependent metabolic pathway is used in rapidly growing microbial organisms, including bacteria and yeast — and is a component of what has been termed the high flux backbone of microbial metabolism. “Drop-out” culture media were prepared individually missing each of the 20 amino acids. It was observed that mouse ES cells grow fine in all drop-out media except that lacking threonine. Cell division and DNA synthesis of mouse ES cells were abruptly impeded in threonine-deficient culture medium. Threonine dependence was not observed for any of a number of other cell lines that do not express the TDH gene, including HeLa cells, mouse embryonic fibroblast (MEF) cells, or NIH 3T3 cells. These studies provide evidence that mouse ES cells are unique in using threonine as a metabolic fuel.

Citation Information: Cancer Res 2009;69(23 Suppl):IA-18.
Abstract IA-21: Malignant progression and the stem-cell state
Authors: Weinberg Robert A.
Published: May 2012

The process of invasion and metastasis represents a complex, multi-step succession of events that, when completed, enables a cancer cell to spawn a macroscopic, clinically detectable tumor. Given the complexity of this process, much effort has been expended in attempting to elucidate how cancer cells acquire the entire spectrum of cell-biological phenotypes that in aggregate are needed to negotiate the steps of the invasion-metastasis cascade, as it is often called. The discovery of the epithelial-mesenchymal transition (EMT) program and its importance in conferring multiple malignancy-associated traits provides one solution to this puzzle, as the expression of one or more EMT-inducing transcription factors can serve to pleiotropically induce many of these cellular phenotypes. Indeed, the expression of these transcription factors may occur in primary tumors through the actions of paracrine signals that are received by carcinoma cells from the nearby tumor-associated stroma.

Several years ago, we and others discovered that the product of the EMT program is not, as we had anticipated, a mesenchymal cell, but instead is a cell exhibiting many stem-cell properties and concomitantly expressing a variety of mesenchymal markers. Indeed, extension of this work now suggests that similar dynamics operate in both immortalized mammary epithelial cells and their transformed derivatives. This indicates that the EMT program not only enables cancer cells to physically disseminate, but in addition, because of the self-renewal potential associated with the stem-cell state, confers on these disseminated cells an ability to spawn the large number of cells forming a metastasis.

Recent research has revealed that stem-like cells can also be found in cultures of mammary epithelial cells that have undergone various degrees of transformation. Of interest is the fact that these cells retain stem-cell-like properties for extended periods of time in culture. Moreover, non-stem-like cells in culture appear to be able to spontaneously convert into stem-cell-like derivatives, a process that appears to be accelerated by cell transformation. This raises the possibility that the typically depicted hierarchy of stem cells generating non-stem cells may need to be revised to accommodate the reversal of this process, that is a bidirectional trafficking between cells in the stem-cell and non-stem-cell compartments of a normal or neoplastic tissue.

Citation Information: Cancer Res 2009;69(23 Suppl):IA-21.
Abstract IA-23: TERT, senescence, and transformation
Authors: Maida Yoshiko , Hayashizaki Yoshihide , Masutomi Kenkichi , Hahn William C.
Published: May 2012

Constitutive expression of telomerase in human cells prevents the onset of senescence and crisis by maintaining telomere homeostasis. However, accumulating evidence suggests that the human telomerase catalytic subunit (hTERT) contributes to cell physiology independent of its ability to elongate telomeres. We have recently found that hTERT interacts with the RNA component of mitochondrial RNA processing endoribonuclease (RMRP), a gene that is mutated in the inherited pleiotropic syndrome Cartilage-Hair Hypoplasia (CHH). hTERT and RMRP form a distinct ribonucleoprotein complex that exhibits RNA dependent RNA polymerase (RdRP) activity and produces double-stranded RNAs that can be processed into small interfering RNA in a Dicer-dependent manner. These observations identify a mammalian RdRP composed of hTERT in complex with RMRP. Since mutations in RMRP are found in CHH, these findings suggest that perturbation of the hTERT-RMRP complex is involved in the pathogenesis of this disorder. The involvement of hTERT in two syndromes characterized by stem cell failure (CHH and dyskeratosis congenita) suggests that ribonucleoprotein complexes containing hTERT play a critical role in stem cell biology.

Citation Information: Cancer Res 2009;69(23 Suppl):IA-23.
Abstract IA-8: p53: From structure to drug discovery
Authors: Fersht Alan R.
Published: May 2012

The ability to produce quantities of stable p53 and the use of new biophysical methods are giving novel insights into the oligomerization of p53 and how this may be important in its regulation. We have discovered that the association and dissociation of p53 is extremely slow and can take more time than the half-life of p53 in the cell. The rate of association equilibria can be catalyzed by the binding to DNA and proteins and also be regulated by the binding to proteins, such as members of the S100 family, for example. I will discuss these phenomena and the latest developments in rescuing unstable oncogenic mutants of p53 by lead compounds for novel anti-cancer drugs.

Citation Information: Cancer Res 2009;69(23 Suppl):IA-8.
Abstract IA-9: Using switchable mouse genetic models to validate therapeutic cancer targets
Authors: Evan Gerard I.
Published: May 2012

Cancers are lethal diseases in desperate need of new therapeutic approaches: despite dramatic advances in the use of conventional chemo and radiotherapy and the growth of more “targeted” drugs, many cancers are still incurable. The principal problem is that we have no systematic or informed way of predicting which, out of the legion of aberrant processes in pancreatic cancer cells, is the best to target with drugs or where, within that process, the most effective molecular targets lie. To address this problem, we employ a novel class of genetically engineered mouse (GEM) in which individual oncogenes and/or tumor suppressor genes may be systemically toggled off and on, reversibly and at will, in vivo. In this way we can identify the most effective therapeutic targets irrespective of contemporary (and ephemeral) prejudices as to their “druggability.” This approach will be illustrated by two such GEMs: one mouse that models pharmacological inhibition of Myc, a core component of the replicative machinery of all tumor and normal cells and a downstream conduit for many (perhaps all) oncogenic growth signals, and a second mouse that models pharmacological restoration of the p53 tumor suppressor that is functionally inactivated in most human cancers. Since it is the endogenous oncogene or tumor suppressor that is toggled in such GEMs, they can be easily applied to any existing or new preclinical mouse cancer model. Using these two GEMs, we have directly ascertained the therapeutic impact, efficacy, and side effects of Myc inhibition and p53 restoration — establishing both mechanism of action and therapeutic index.

Citation Information: Cancer Res 2009;69(23 Suppl):IA-9.
Identification of Unique MEK-Dependent Genes in GNAQ Mutant Uveal Melanoma Involved in Cell Growth, Tumor Cell Invasion, and MEK Resistance
Authors: Ambrosini Grazia , Pratilas Christine A. , Qin Li-Xuan , Tadi Madhavi , Surriga Oliver , Carvajal Richard D. , Schwartz Gary K.
Published: May 2012

Purpose: Metastatic uveal melanoma represents the most common intraocular malignancy with very poor prognosis and no effective treatments. Oncogenic mutations in the G-protein α-subunit q and 11 have been described in about 85% of uveal melanomas and confer constitutive activation. Multiple signaling pathways are induced as a consequence of GNAQ/11 activation, which include the MEK/ERK kinase cascade. We analyzed the transcriptional profile of cell lines treated with a mitogen-activated protein (MAP)/extracellular signal–regulated (ERK) kinase (MEK) inhibitor to identify gene targets of activated GNAQ and to evaluate the biologic importance of these genes in uveal melanoma.

Experimental Design: We conducted microarray analysis of uveal melanoma cell lines with GNAQ mutations treated with the MEK inhibitor selumetinib. For comparison, we used cells carrying BRAFV600E and cells without either mutation. Changes in the expression of selected genes were then confirmed by quantitative real-time PCR and immunoblotting.

Results: We found that GNAQ mutant cells have a MEK-dependent transcriptional output and identified a unique set of genes that are downregulated by MEK inhibition, including the RNA helicase DDX21 and the cyclin-dependent kinase regulator CDK5R1 whereas Jun was induced. We provide evidence that these genes are involved in cell proliferation, tumor cell invasion, and drug resistance, respectively. Furthermore, we show that selumetinib treatment regulates the expression of these genes in tumor tissues of patients with metastatic GNAQ/11 mutant uveal melanoma.

Conclusions: Our findings define a subset of transcriptionally regulated genes by selumetinib in GNAQ mutant cells and provide new insights into understanding the biologic effect of MEK inhibition in this disease. Clin Cancer Res; 18(13); 3552–61. ©2012 AACR.
Abstract A112: Molecular variants of E7/ HPV58 in women with normal citology from the Colombian cohort. Preliminary results
Authors: Buitrago Oscar , Martin Carolina , Huertas Antonio , Moreno Pablo , Hernandez Gustavo , Martinez Teresa , Molano Monica
Published: May 2012

A112

Introduction

Human papillomavirus type 58 (HPV 58) is the second viral type more prevalent in women with normal cytology from the Colombian population. Also this type is highly prevalent in women with high grade squamous intraepithelial cervical lesions (HGSIL) and in women with invasive cervical cancer worldwide. There are few studies of E7/HPV 58 variants, however some of them have shown an association of variants presence with a higher risk of HGSIL and cervical cancer in Asian populations. In Colombia there are not studies of presence and persistence of E7/HPV58 variants.

Objective

To identify molecular variants of E7/HPV58 in cervical scrapes of women with normal cytology that belongs to the base line of the Bogotá, Colombia Cohort.

Materials and Methods

1845 cervical scrapes of women with normal cytology that belong to the base line of the Bogotá, Colombia cohort were analysed. HPV detection was done using the GP5+/GP6+ primers and 37 different HPV types were typed using a PCR-RLB assay. The E7 region of samples HPV 58 positives was amplified using the E7P1- E7P2 primers that amplify a fragment of 316 bp (561 to 877 nucleotides). E7/HPV 58 variants were detected using automated direct sequencing. The reference sequence of HPV 58 was used to compare sequences.

Results

34 samples were positive for HPV 58 (1.84%). Until this moment 13 samples have been analysed for E7/HPV 58 variants. 12 of these samples have the A694/G744/A761 variant (94.7%), one sample have the A599/A694/G744/A761 variant (5,5%), and none of the samples had the reference sequence of HPV58.

Conclusions

Two variants have been identified in the samples analysed: 1 previously identified in Asian population and 1 new variant (A599/A694/G744/A761). There are few diversity of E7/ HPV58 variants in the analysed samples. The cohort analysis will give important information about the role of these variants in the persistence of HPV 58 infection and in the development of cervical intraepithelial lesions.

Citation Information: Cancer Prev Res 2008;1(7 Suppl):A112.
Abstract A113: Association between NSAIDs and PSA, prostate volume, and prostate cancer
Authors: Fowke Jay , Motley Saundra , Byerly Susan , Cookson Michael , Chang Sam , Smith Joseph
Published: May 2012

A113

Purpose

NSAIDs such as aspirin are commonly used to prevent cardiovascular disease, and several prior studies suggest NSAIDs may also reduce prostate inflammation and prostate cancer risk. We investigated the association between NSAID use, PSA, and prostate volume, hypothesizing lower PSA and volume levels with NSAID use.

Methods

The Nashville Men’s Health Study utilizes a multi-centered, rapid-recruitment protocol to collect clinical, biological, behavioral, and body measurement data from 1,277 men over age 40 years and scheduled for diagnostic prostate biopsy. Approximately 95% of eligible men approached for recruitment agree to participate. NSAID use was ascertained by survey and clinical interview, and medical charts were reviewed to ascertain current PSA levels, prostate volume, and clinical diagnoses following biopsy. The distributions of PSA level and prostate volume data were natural log transformed prior to analysis. In a linear model, mean PSA and volume scores across NSAID categories were adjusted for age; race (black/white); family history (yes/no); number of prior PSA tests (1,2, 3 or more); BMI; WHR; height; or treatment for BPH (steroid reductase inhibitors, other), diabetes (Yes/No), CVD (Yes/No), hyperlipidemia (Yes/No); and the diagnosis of PIN, atypical findings suspicious for cancer, low-grade cancer, or high-grade cancer following biopsy. PSA and prostate volume scores were back-transformed and geometric mean values are reported.

Results

Approximately 46% of subjects reported taking an NSAID, primarily aspirin (37% of subjects). After adjusting for age, race, family prostate cancer history, obesity, and treatment for BPH, CVD, hyperlipidemia, and diabetes; aspirin was significantly associated with lower PSA levels (7.3 vs. 8.0 ng/ml, p=0.01). This effect of aspirin on PSA was greatest among men with a prostate volume of 60 mls or more (8.6 vs. 9.7 ng/ml, p=0.06), men diagnosed with prostate cancer (6.1 vs. 7.3 ng/ml, p<0.01), or men with both cancer and prostate enlargement (7.3 vs. 12.7 ng/ml, p<0.01). NSAID use was not significantly associated with prostate volume (47.6 vs. 46.0 mls, p=0.16).

Conclusions

Aspirin use was significantly associated with lower PSA levels. Controlling for BMI, WHR, and the use of medications to treat CVD, hyperlipidemia, BPH, and diabetes had little effect on the association between aspirin use and PSA levels, suggesting aspirin was not simply a proxy for these comorbid conditions or their associated treatments. These results may suggest that aspirin use decreases the ability to detect prostate cancer and may contribute to prior investigations reporting a protective association between NSAID use and prostate cancer risk. Implications for prostate cancer screening recommendations and for clinical decision-making will require further investigation.

Citation Information: Cancer Prev Res 2008;1(7 Suppl):A113.
Abstract A117: A meta-analysis of the incidence of non-AIDS cancers in HIV-infected individuals
Authors: Shiels Meredith , Cole Stephen , Poole Charles
Published: May 2012

A117

Objective

To compare the incidence rate of all non-AIDS cancers combined among HIV-infected individuals with the general population rate and to estimate modification of the association by gender and AIDS diagnosis.

Design

A meta-analysis was carried out using data from 11 studies of non-AIDS cancer incidence in HIV-infected individuals.

Methods

Standardized incidence rate ratios (SIR) for non-AIDS cancers in HIV-infected individuals compared to the general population and 95% confidence limits (CL) were abstracted from each study. The results were assessed for between-study heterogeneity and funnel plot asymmetry. Meta-regression was used to estimate modification of the general-population SIR by gender and AIDS.

Results

There was pronounced heterogeneity between studies, thus, an overall pooled estimate was not calculated. The SIR of non-AIDS cancers in HIV-infected individuals, adjusted for AIDS and study design was greater among men (2.3; 95% CL: 1.7, 3.1) than women (1.5; 95% CL: 0.9; 2.3; ratio of SIRs = 1.6; 95% CL: 0.7, 3.2). The SIR also appeared slightly greater among those with an AIDS diagnosis; however, when the analysis was adjusted for gender and study design, the SIRs for those with (1.88; 95% CL: 1.34, 2.66) and without AIDS (1.98; 95% CL: 1.35, 2.88) were similar (ratio of SIRs: 0.95; 95% CL: 0.46, 1.96).

Conclusions

The incidence rate of non-AIDS cancers was elevated among HIV-infected individuals. Additionally, HIV-infected men appeared to have a greater SIR for non-AIDS cancers than HIV-infected women. However, no difference was seen between the SIRs for non-AIDS cancers among those with and without AIDS.

Citation Information: Cancer Prev Res 2008;1(7 Suppl):A117.
Abstract A14: The role of Livin in oncogenesis reveals a novel mode of gene regulation
Authors: Itay Lazar , Perlman Riki , Kadouri Luna , Ben-Yehuda Dina
Published: May 2012

A14

Acquired resistance to apoptosis is a hallmark of all types of cancer. The Inhibitor of Apoptosis Protein (IAP) family members is able to inhibit apoptosis induced by a variety of stimuli mainly by binding to and inhibiting caspases.

We and others previously identified the IAP Livin. We further showed that Livin functions as a regulator of apoptosis that can undergo cleavage by effector caspases to produce a truncated form with paradoxical pro-apoptotic activity.

In this study we describe the role of Single Nucleotide Polymorphism (SNP) in the regulation of Livin in normal and malignant cells and demonstrate how the 528 (C/T) SNP regulates Livin protein expression. We show that Livin is expressed in a non-random monoallelic manner as only the 528T allele is expressed in normal blood and in early-stage melanoma. In advanced melanoma, by mechanism of DNA overrepresentation, both 528T and 528C mRNA alleles of Livin are expressed, resulting in expression of the Livin protein.

The variable expression of the Livin protein plays a major role in the survival of melanoma patients: We show that high Livin expression is an independent adverse prognostic marker. Importantly, we show that low-intermediate expression of Livin is associated with favorable prognosis in comparison to no Livin expression.

Citation Information: Cancer Prev Res 2008;1(7 Suppl):A14.
Abstract A19: Identification of epigenetic hits can predict breast cancer risk
Authors: Fischer John , Rossetti Stefano , Cheney Richard , Sacchi Nicoletta
Published: May 2012

A19

RARB2 is a master tumor suppressor that mediates the growth-inhibitory action of retinoic acid (RA). Homozygosis for epigenetically silent RARB2 alleles, which results into loss of RARB2 tumor suppressor activity, leads to RA-resistance, and apparently precedes the acquisition of morphological transformation of breast epithelial cells (Bistulfi et al., Cancer Research, 2006). RARB2 epigenetic silencing is marked by chromatin repressive changes, including DNA methylation of the RARB2 CpG island (Sirchia et al., Oncogene, 2000). In a mechanistic study, we demonstrated that methylation arises at a specific epicenter in the RARB2 CpG island, which we call the RARB2 methylation epicenter (RME) (Ren et al., MCB, 2005). Here we show that RME methylation is detectable in both benign and ductal carcinoma in situ within the same patient tissue sample. This finding implies sequential epigenetic silencing of RARB2 alleles. Timely identification of the first epigenetic hit in breast tissue of women at risk of breast cancer could prevent progression to RME methylation homozygosity, the consequent loss of RARB2-mediated tumor suppressor function, RA-resistance, and morphological epithelial cell transformation.

This work was supported by the Roswell Park Alliance Foundation Award (NS), the Breast Cancer Coalition of Rochester (NS), the Susan Komen Foundation (SR).

Citation Information: Cancer Prev Res 2008;1(7 Suppl):A19.
Abstract A22: HIF-1 mediates up-regulation of urokinase-type activator receptor (uPAR) in cervical cancer cells
Authors: Nishi Hirotaka , Isaka Keiichi
Published: May 2012

A22

Hypoxia occurs during development of cervical cancer and is considered to correlate with its invasion. Hypoxia induces cancer cells to have more invasive property through the urokinase plasminogen activator receptor (uPAR) expression. We sought to determine the regulator(s) of uPAR expression during hypoxia. According to the invasion assays, cervical cancer cell lines, CaSki and CA, under hypoxic condotion (1% O2) showed more invasive property than those under normoxia. By western blot analysis, hypoxia enhanced the endogenous hypoxia inducible factor (HIF)-1alpha and uPAR protein expression. uPAR mRNA level was also up-regulated by hypoxia using realtime RT-PCR. Overexpression of HIF-1alpha which is induced by hypoxia activated the transcriptional activity of the uPAR promoter by luciferase assays. HIF-1 protein bound the putative HIF-1 response element on the uPAR promoter using the NoShift Transcription Factor Assay kit, and additional luciferase assays show that this is essential for uPAR transactivation by HIF-1. HIF-1 overexpression enhanced the endogenous uPAR expression and introduction of siRNA for HIF-1alpha diminishes uPAR expression during hypoxia. These results indicate the up-regulation of uPAR by hypoxia in cervical cancer cells is mediated through HIF-1. Using cervical cancer cell tissues, we also demonstrated that the level of uPAR mRNA in normal cervix is significantly lower than cervical cancer. Our results provide evidence that regulation of uPAR expression by HIF-1 represents a mechanism for cervical cancer invasion during hypoxia.

Citation Information: Cancer Prev Res 2008;1(7 Suppl):A22.
Abstract A5: Inhibition of tumor cell migration and angiogenesis by sulphated polysaccharides from Sargassum wightii (Greville)
Authors: Delma Caroline , Ramalingam Karthikeyan , Pandian Vijayabaskar , Baskar Albert , Savarimuthu Ignacimuthu , Thangavelu Balasubramanian , Somasundaram Somasundaram
Published: May 2012

A5

Seaweeds are potential renewable resources of the marine environment. They are low in fats but contain vitamins and bioactive compounds like terpenoids and sulphated polysaccharides. Sulphated polysaccharides from brown algae are found to exhibit diverse biological activities like anticoagulant, antihyperlipidemic, antiviral, antioxidant, antitumor, antimetastatic and antiangiogenic activities. In this study, the antiproliferative, antiinvasive and antiangiogenic properties of sulphated polysaccharides from brown seaweed, Sargassum wightii (Greville) against colon carcinoma cell line COLO 320DM were investigated. Crude sulphated polysaccharide was extracted from S. wightii collected from Mandapam coast, South east coast of India. The antiproliferative activity of the seaweed polysaccahrides on COLO 320DM was evaluated by MTT assay and DNA fragmentation studies. The inhibitory potential on the migration of this invasive cell line was assessed by wound healing assay. The effect of sulphated polysaccharides against angiogenesis was investigated by chorio allantoic membrane (CAM) assay in chick embryo. Sulphated polysaccharides from S. wightii exhibited moderate growth inhibitory activity against COLO 320DM at a concentration of 1mg/ml at 24 hours. DNA fragmentation studies revealed characteristic ladder formation which indicates that seaweed polysaccharides have effected the induction of apoptosis. The seaweed extract was found to inhibit the migration of the colon carcinoma cell line. The antiinvasive property increased with increasing concentrations. However, the activity was more or less similar at 24 and 48 hours. The CAM assay showed that sulphated polysaccharides from the brown seaweed exhibited promising activities against neoangiogenesis. These results clearly indicate that sulphated polysaccharides from brown algae are potential molecular leads for antimetastatic drug development.

Citation Information: Cancer Prev Res 2008;1(7 Suppl):A5.