bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 5044: Dual GSK-3β/CDK inhibitors induce multiple myeloma cell death and eliminate putative cancer stem cell populations
Authors: Dolloff Nathan G. , Hart Lori S. , Mayes Patrick A. , El-Deiry Wafik S.
Published: February 2011

Multiple myeloma (MM) is a plasma cell malignancy that recurs in 100% of patients despite high treatment response rates. The incurable nature of the disease and the low proliferative capacity of terminally differentiated MM plasma cells supports the existence of a cancer stem cell (CSC) component - a cell population with self-renewal and drug resistance properties capable of repopulating the disease after seemingly effective treatment and periods of undetectable disease. It is imperative to develop new therapies that target both terminally differentiated MM plasma cells and MM CSCs to extend disease-free survival and control the disease. We tested the ability of c-Myc-inducing compounds, such as GSK-3β and dual GSK-3β/CDK inhibitors, to enhance the activity of bortezomib against advanced MM cell lines (RPM-I8226 and NCI-H929). We have shown that this class of inhibitors has anti-tumor effects and sensitizes numerous solid malignancies to apoptosis-inducing agents (Mayes P.A., Dolloff N.G. et al., in revision). Moreover, c-Myc expression increases protein translation, and in doing so, enhances the cytotoxicity of proteasome inhibitors such as bortezomib (Nawrocki et al., 2008). We found that GSK-3β inhibitors (lithium choloride and sc-24020) failed to induce apoptosis or reduce viability of MM cell lines or to enhance the activity of bortezomib. By contrast, the dual GSK-3β/CDK inhibitors, alsterpaullone and the nucleoside analogue SLM3 (Mayes P.A., Dolloff, N.G. et al., in revision), were highly toxic to MM cells both alone and in combination with bortezomib. In addition to inducing massive apoptosis and reducing MM cell viability, alsterpaullone and SLM3 eliminated the presence of dye-effluxing side-populations, that have been shown to exhibit CSC behavior in vitro and in animal models of MM. By comparison, bortezomib, adriamycin and melphalan increased the percentage of MM side-population cells, suggesting that MM CSC cells are more resistant to these agents compared to their non-CSC counterparts. Interestingly, alsterpaullone and SLM3 eliminate a distinctly large sub-population of MM cells that are enriched in side-population cells, providing a further characterization of the MM CSC phenotype. We propose that using GSK-3β/CDK inhibitors may be effective therapies, as they target tumorigenic CSC populations, which are responsible for disease relapse. We are currently testing this hypothesis in mouse models of MM.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5044.
Abstract 5048: TAp73a enhances the cellular chemosensitivity to cisplatin in ovarian cancer cells
Authors: Zhang Pingde , Liu Stephanie S. , Ngan Hextan YS
Published: February 2011

Ovarian cancer is the most lethal gynecological malignancy. Most ovarian cancer patients relapse and subsequently die due to the development of resistance to chemotherapy. Strategies for overcoming resistance to chemotherapy and improving prognosis of ovarian cancer patients are very critical. P73, one member of the tumor suppressor p53 family, shares highly structural and functional similarity to p53. Established evidence showed that p73 mediated cellular sensitivity to chemotherapeutic agents in some human tumor cells. However, the role of p73 in the regulation of cellular response to DNA damaging agents in ovarian cancer is still poorly understood, and the function of the transcriptionally active isoform TAp73 as a tumor suppressor is still controversial. The aim of the present study was to examine the effect of TAp73a on cellular response to commonly used chemotherapeutic agent cisplatin in ovarian cancer cells. We established TAp73a overexpressed stable clones in two ovarian cancer cell lines: OVCA433 (wild-type p53) and SKOV3 (null p53). We found that TAp73a enhanced chemosensitivity of ovarian cancer cells to cisplatin by inhibiting cell growth and inducing cell cycle arrest and apoptosis in wild-type p53 OVCA433 cells. Additionally, TAp73a up-regulated the p53 expression and its downstream genes: PUMA, NOXA, p53AIP1, and down-regulated the phosphorylation of Akt as assessed by western blotting. The accumulation of p53 protein induced by TAp73a was found at the post-transcriptional level. Co-immunoprecipitation assay demonstrated that TAp73a directly bound to p53 protein, suggesting that such interaction might block the degradation of p53 protein in OVCA433 cells. In p53-null SKOV3 cells, the increase of chemosensitivity induced by TAp73a was also observed, indicating that TAp73a also functioned independent of p53 by up-regulating the cell cycle inhibitor p21 expression. In conclusion, these results suggested that TAp73a might play an important role in the enhancement of cellular sensitivity to chemotherapeutic drug cisplatin in ovarian cancer cells through p53 dependent and independent pathways.

Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5048.
Abstract 5051: Overexpression of talin 1 (TLN1) in invasive oral squamous cell carcinoma (OSCC)
Authors: Sheu Jim J. , Lai Ming-Tsung , Tseng Hsien-Chang , Lin Ying-Ju , Wan Lei , Hua Chun-Hung , Tsai Ming-Hsui , Chiu I-Wen , Chang Nai-Wen , Lin Chin-Fen , Chen Chih-Mei , Lee Min-Min , Tsai Fuu-Jen
Published: February 2011

Oral squamous cell carcinoma (OSCC) is a highly invasive disease displaying frequent tumor recurrence and lymph node metastasis. Genes involved in cytoskeleton remodeling, cell attachment, and cell mobility are suspicious candidates that promote invasiveness. A genomewide study based on Sty1 250K SNP array platform revealed the involvement of TLN1 gene in the 9p13.3 amplicon. Although large scale screening on 123 tumor tissues by dual color-fluorescence in situ hybridization (FISH) indicated low frequency (around 7.3%) of TLN1 amplification in OSCC, the overall gene expression levels were significantly higher in tumor tissues as compared to the adjacent normal oral epithelium (P < 0.01). As compared to other cytoskeleton crossing linkers that can trigger integrin activation, TLN1 stood out to be the most highly expressed one in OSCC. A mouse study revealed that TLN1 overexpression could be detected in invasive tumors but not in the lesions at early stages. Immunohistochemistry (IHC) data revealed stronger TLN1 immunereactivity in OSCC than in normal tissues. More intense staining pattern was detected at the leading edge of cancer cells under going angiogenesis or lymphogenesis. Patients with TLN1 overexpression had a significantly shorter overall survival and higher recurrence rate than those without. Collectively, our study suggested an important role of TLN1 in oral cancer progression.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5051.
Abstract 5054: The interplay between Disabled-2 and protein kinase C epsilon in the regulation of prostate cancer cell survival
Authors: Tseng Wei-Lien , Chen Yan-Yu , Cheng Ju-Chien , Tseng Ching-Ping
Published: February 2011

Prostate cancer is the leading cause of cancer-related death in men in the world. Previous studies have demonstrated that Disabled-2 (DAB2) is a tumor suppressor and down-regulation of DAB2 expression is accompanied with prostate cancer progression. On the other hands, protein kinase C epsilon (PKCε) functions as an oncogene and promotes human prostate cancer cell survival upon treatment of methyseleninic acid (MSA), an anti-cancer agent that induces prostate cancer cell apoptosis. However, little is known whether there is a crosstalk between DAB2 and PKCε. In this study, we first analyzed DAB2 and PKCε protein expression in three prostate cancer cell lines DU145, PC3 and C4-2. Western blot analysis revealed that DAB2 was relatively abundant in PC3 when compared with the DU145 and C4-2 cells. In contrast, the expression of PKCε was higher in DU145 and C4-2 cells when compared with the PC3 cells. Consistent with these findings, ectopic expression of DAB2 resulted in down-regulation of PKCε. These observations implicate a reverse association of DAB2 and PKCε and suggest that DAB2 is a regulator of PKCε expression. To further explore the interplay between DAB2 and PKCε, we determined the effect of DAB2 overexpression on DU145 cell susceptibility to MSA-induced cell death. Consistent with a previous report, overexpression of PKCε increased DU145 cell survival and knockdown of PKCε by small interfering RNA augmented cell death upon treatment of MSA but not Ibupofen. Accordingly, expression of DAB2 down-regulated PKCε and increased in cell susceptibility to MSA that resulted in a 40% decrease of cell survival. These findings suggest that DAB2 modulates PKCε expression and affects prostate cancer cell susceptibility to selective anti-cancer agents. This study represents the first report to unveil the interplay between DAB2, PKCε and prostate cancer cell survival.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5054.
Abstract 5055: TOPK phosphorylation of Prx1 at Ser32 prevents UVB-induced apoptosis in RPMI7951 melanoma cells through regulation of its peroxidase activity and blockade of intracellular accumulation of H2O2
Authors: Zykova Tatyana , Zhu Feng , Vakorina Tatyana I. , Zhang Jishuai , Higgins LeeAnn , Urusova Dasha V. , Bode Ann M. , Dong Zigang
Published: February 2011

A novel serine/threonine kinase, T-LAK cell-originated protein kinase (TOPK), is only expressed in a wide range of cancer cells. We previously showed that TOPK increases colorectal cancer growth in vivo and ex vivo and that melanoma cell lines with a high abundance of TOPK protein exhibit a marked resistance to As3+-induced apoptosis. Here we continue our investigation of the functional details of TOPK. LC/MS/MS data showed that TOPK can bind with peroxiredoxin 1 (Prx1), which regulates hydrogen peroxide-mediated signal transduction. The role of Prx1 phosphorylation induced by UV in melanoma is not clear. Here, we provide evidence that TOPK phosphorylates Prx1 at Ser32 in vitro. Analysis of the CD spectra of Prx1 and a mutant Prx1 (S32A) protein shows that the secondary structure of Prx1 was significantly altered by its phosphorylation at Ser32. UVB radiation induced phosphorylation of TOPK in a time-dependent manner in RPMI7951 human melanoma cells and phosphorylated TOPK co-localized with Prx1 in the nucleus. UVB treatment increased peroxidase activity of Prx1 in vitro and ex vivo. The level of H2O2 was increased in TOPK siRNA cells and in RPMI7951 cells stably overexpressing mutant Prx1 (S32A). In addition, apoptosis was dramatically increased in TOPK siRNA cells and decreased in WT-Prx1 stable cells after UVB treatment. Our results suggested that TOPK phosphorylation of Prx1 at Ser 32 prevents UVB-induced apoptosis in RPMI7951 melanoma cells through the regulation of Prx1 peroxidase activity by blocking intracellular accumulation of H2O2.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5055.
Abstract 5056: Signaling pathways critical for insulin receptor substrate (IRS)-mediated disruption of MCF-10A acini formation
Authors: Chan Bonita Tak-Yee , Kuiatse Isere , Lee Adrian V.
Published: February 2011

Insulin receptor substrate (IRS)-1 and IRS-2 are important adaptor proteins in insulin-like growth factor-I receptor (IGF-IR) signaling. IRS-1 and IRS-2 are expressed during mammary gland development and also in human breast tumors. In order to study the role of IRS-1 and IRS-2 in breast tumorigenesis, we overexpressed both IRS proteins in immortalized, non-transformed human mammary epithelial MCF-10A cells. It has been shown that MCF-10A cells form uniform, empty-lumen acini in three dimensional (3D) culture. We previously showed that overexpression of either IRS-1 or IRS-2 in MCF-10A cells disrupted the acini in 3D culture and resulted in extensive proliferation and disrupted apical-basal polarity. In both IRS-1 and IRS-2 overexpressing MCF-10A cells, an increase in tyrosine phosphorylation and association with p85 of both IRS proteins indicated that IRS-mediated signaling was activated. siRNA knockdown of the overexpressed IRS proteins reversed the disruption of MCF-10A acini, confirming that this is indeed an IRS-related event. In order to identify the critical effectors in IRS-mediated disruption of MCF-10A polarity and growth, we used small molecule inhibitors to screen multiple kinase signaling pathways upstream and downstream of the IRS proteins. Inhibition of Rac or atypical PKC (aPKC) didn't affect MCF-10A cell acini formation, however it abolished the IRS-mediated disruption of MCF-10A acini formation. Consistent with this, knockdown of Rac and aPKC by siRNA suppressed IRS-mediated disruption in acini formation. Rac belongs to the family of GTPase that regulates cell motility. aPKC is a key component of the PAR polarity complex, and has previously been shown to be involved in IRS-signaling in adipocytes. While the signaling mechanism of Rac and aPKC in the IRSs-induced disruption of acini is still unknown, we found that another component in the PAR polarity complex Par3 may associate with IRS proteins and leads to dissociation of the polarity complex. Taken together, overexpression of IRS-1 and IRS-2 disrupts the MCF-10A acini in 3D culture in part through Rac and aPKC pathways.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5056.
Abstract 5059: Detection of K-RAS mutation in clinical samples by one-step PNA-mediated real-time PCR clamping techniques
Authors: Cho Minhye , Choi Jae-Jin , Park Heekyung
Published: February 2011

Mutations in the K-RAS oncogene are frequently found in human cancers. K-RAS mutations may indicate prognosis and drug response, and many new cancer therapies are being targeted to the K-RAS pathway. Many researchers suggest that in colon cancer patients the K-RAS mutation status is strong predictor of resistance to therapy with tyrosine kinase inhibitors such as Erbitux® (cetuximab/ Imclone system Inc) or Vectibix® (panitumumab/ Amgen Inc). Therefore, it is important that detection of K-RAS mutations is fast and simple with high accuracy and high sensitivity for predict drug response and prognosis.

To detect a minority of mutant K-RAS among abundant wild-type alleles, we developed a high sensitivity and simple method for detection of K-RAS mutations using PNA-mediated real-time PCR clamping. PNA mediated real-time PCR clamping relies on the following two unique properties of PNA probes. 1) PNA-DNA duplexes generally have greater thermal stability than the corresponding DNA-DNA duplex and 2) PNA oligomers are not recognized by DNA polymerases and consequently can serve as sequence selective clamp during PCR amplification.

One-step PNA-mediated real-time PCR clamping method was optimized for detection of K-RAS mutations in exon 12 and 13 with high sensitivity. This method was simple and correct with detection limit approximately 0.1% mutant alleles using 10ng to 50ng of normal DNA as the template. The turnaround time for performing this assay was only 2.5hrs.

K-RAS mutations were detected in 24.4% (22/90) of clinical samples, of which 18 had mutations in codon 12 and 4 had these in codon 13. To confirm these results, same clinical samples were analyzed with sequencing assay. The concordant results were obtained from 88 (98%) of the 90 clinical samples by the two assays. The PNA-mediated real-time PCR clamping is a rapid and reliable tool for K-RAS mutation detection and allowed a minority of mutant in clinical field.

Keywords: K-RAS, Mutation, PNA, cancer, PCR clamping

This work was supported be the Small and Medium Business Administration funded by the Korean Government. (S1060400)

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5059.
Abstract 506: A time-dependent genome-wide gene interaction signature in U251 human glioma cell line exposed to ionizing radiation
Authors: Zach Leor , Hakim Ofir , Sung Myong Hee , Hager Gordon , Kevin Camphausen
Published: February 2011

Background: The effects of ionizing radiation on DNA integrity and functionality are well studied and reported. However, little is known about how irradiation affects large-scale nuclear architecture. We sought to investigate the latter using Chromosome Conformation Capture on Chip (4C) which assesses the spatial genomic organization around a locus of interest. Methods: U251 glioma cells were grown in standard media to 70% confluence, irradiated (6Gy) and harvested 6 and 24 hours after irradiation. We obtained all genomic loci in close physical proximity to an irradiation induced gene Alf1 on chromosome 6 as bait. 4C was performed in duplicate samples on control or irradiated cells, using custom NimbleGen microarrays that contained probes for all potential partner sites in the genome. Results: Alf1 locus was found to be engaged in multiple intra-chromosomal interactions with other regions in chromosome 6, and inter-chromosomal interactions with multiple areas in chromosomes 1,2,5,7,10 15 and 17 prior to irradiation. Irradiation induced changes in the Alf1 spatial environment as many of the contacts were lost, while additional interactions were induced. A partial recovery to the baseline nuclear architecture was evident 24 hrs after irradiation. Conclusions: Our preliminary results are intriguing and might suggest a new tool to investigate the effects of ionizing irradiation on nuclear high-order structure. A specific, time dependent post irradiation DNA architecture changes in tumor cells may correlate to the (and reproducible) levels of sensitivity to ionizing irradiation of different areas of the DNA in a specific tumor cell population.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 506.
Abstract 5063: Signatures of Ral GTPase status define key clinicopathologic characteristics and outcomes in cancer
Authors: Smith Steven C. , Baras Alexander S. , Wang Hong , Owens Charles R. , Theodorescu Dan
Published: February 2011

Ral GTPases, which include RalA and RalB, mediate essential phenotypes in model cancer systems; however, demonstration of the clinical relevance of these molecules has been lacking. To address this concern, we develop two types of gene expression signatures as surrogates for Ral GTPase status in tumors and characterize their nosologic and prognostic performance across several epithelial malignancies. Given Ral's well characterized role in regulating a variety of transcriptional pathways including AP1, NFkB, STAT3B, forkhead family members, and RREB1, the first type of signature is related to transcription downstream of RalA and RalB and was derived empirically from expression profiling of cells depleted of these molecules. This signature was then applied to human tumor gene expression data by using a nearest neighbor algorithm for classification. Consistent with prior findings of a role for these molecules in model surrogates of cancer progression(1,2), presence of the Ral transcriptional signature characterized the muscle invasive status of human bladder cancer (P<0.001), including prediction of subsequent development of invasion in non-invasive tumors (P<0.01). In prostate cancer, the signature characterized androgen independent status, and estrogen receptor status in human breast cancer, both datasets P<0.05. Supportive of a recent report of tumor suppressor function of Ral in a model of squamous cell cancer (3), the signature was inversely associated with stage in this tumor type.

Secondly, we have assayed constitutively-active RalA and RalB mutants, finding that either induces accelerated experimental metastasis of UMUC3 bladder cancer cells in mice (logrank P=0.01), similar to results for other tumor types (4). This made us question whether signatures of metastasis from tumor profiling could be associated with differential Ral activation. We assayed the activation of RalA and RalB across 40 cell lines, then mapped individual tumor expression of metastatic genes to these cell lines in silico. Using this methodology to compare tumors to cell lines of known Ral activation, we found association of higher Ral activation with nodal metastasis in bladder cancer and with visceral metastasis in breast cancer, both P<0.05. Taken together, these findings begin to make the as yet missing clinical case for the Ral pathway in human tumors and call for aggressive investigation of therapies to interrupt this pathway.

(1) Oxford, Cancer Res, 2005

(2) Lim, Curr Biol, 2006

(3) Sowalsky, Oncogene, 2009

(4) Yin, Mol Cell Biol, 2007

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5063.
Abstract 5068: Identification of RASSF1A as a novel RhoA antagonist: direct interaction with and Smurf1-mediated ubiquitination of RhoA
Authors: Ryu Byung-Kyu , Lee Min-Goo , Kang Min-Ju , Ha Tae-Kyu , Han Jikhyon , Her Nam-Gu , Chung Seong-In , Chi Sung-Gil
Published: February 2011

RASSF1A is a tumor suppressor which plays a critical role in the regulation of various aspects of cellular functions, including cell proliferation, apoptosis, and motility. RASSF1A is frequently inactivated in various types of human cancers by transcriptional silencing of the gene due to aberrant promoter hypermethylation. However, molecular mechanisms underlying its tumor suppressive functions were largely undefined. We characterized RASSF1A regulation of a small GTPase RhoA and its implication in tumor cell migration and invasion. Effect of RASSF1A expression on RhoA activity was examined by immunoblot, immunopricipitation, and ubiquitinylation assays, and RASSF1A modulation of RhoA-mediated cell migration and invasion was determined using wound healing and in vitro invasion assay. We identified that RASSF1A inhibits serum- or LPA-induced RhoA activation in both cancer cells and noncancerous cells. SiRNA-mediated knockdown of RASSF1A increases RhoA activity and RhoA-mediated tumor cell migration. RASSF1A promotes the protesomal degradation of GTP-bound RhoA via Smurf1-mediated ubiquitinylation, but does not affect the activity and stability of Rac1 and CDC42. Immunofluoroscence and immunopricipitation assays revealed that RASSF1A colocalizes and interacts with RhoA and Smurf1. The N-terminal region of RASSF1A interacts with Smurf1, and its C-terminal residues (amino acids 266-272) are critical for the interaction with RhoA. The mutant RASSF1A (delNC-RASSF1A) having a sequence alteration in the Smurf1 and RhoA binding region fails to inhibit the RhoA-mediated tumor cell migration and invasion. Consistent with theses results, loss or abnormal reduction of RASSF1A expression exhibits a tight correlation with RhoA activation in both cancer cell lines and primary tumors of various tissues origins. Our study demonstrates first that RASSF1A regulates cell motility through the proteosomal degradation of activated RhoA through Smurf1-mediated ubiquitinylation, indicating that genetic and epigenetic inactivation of RASSF1A during tumorigenic process increases the invasive and metastatic potential of tumor cells by deregulation of RhoA.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5068.
Abstract 5069: c-Abl contributes to Mdm2-Mdmx complex formation protecting p53 levels
Authors: Waning David L. , Mayo Lindsey D.
Published: February 2011

This study was designed to better understand the signaling events through c-Abl kinase that impact p53 activity. The p53 tumor suppressor protein is a key protein in maintaining the integrity of the genome by inducing either cell cycle arrest or apoptosis following cellular stress signals. Two human family members, Mdm2 and Mdmx, are primarily responsible for inactivating transcription initiation of p53 or destabilizing p53 through ubiquitination. In response to genotoxic stress, post-translational modifications to Mdm2 and Mdmx relieve inhibition on p53, which is in part mediated through the c-Abl kinase. c-Abl is necessary for maintaining p53 expression levels and for maximal accumulation of p53 following DNA damage. While p53 is not a direct substrate of c-Abl there are c-Abl phosphorylation sites present in Mdm2 and Mdmx. Considering that c-Abl regulates both Mdm2 and Mdmx, we designed this study to understand the role of c-Abl phosphorylation of Mdm2 in protecting p53 activity. We hypothesized that protection of p53 would be linked to Mdm2-Mdmx complex formation and modulated by c-Abl phosphorylation. Here we present a mechanism of p53 stabilization whereby c-Abl promotes Mdm2-Mdmx heterodimer complex formation. For this study we used multiple complementary approaches including: recombinant in vitro protein assays, therapeutic drug treatments of cells and targeted mutagenesis of Mdm2 phosphorylation sites to show the role of c-Abl in protecting p53. Additionally, blocking c-Abl phosphorylation has therapeutic implications in treatment of disease through modulation of p53 activity. There are several small molecule inhibitors that affect c-Abl activity, such as imatinib, and a better understanding of the mechanistic role of c-Abl in the stoichiometric balance between Mdm2 and Mdmx may lead to better approaches for engaging p53 activity. This could lead to improvements in current disease treatment and the development of novel therapeutics for cancers harboring wild-type p53.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5069.
Abstract 507: Elevated manganese superoxide dismutase (SOD2) activity as a mechanism for the low dose dadiation- and thiol-induced adaptive responses in human RKO36 colon carcinoma cells and BFS2C mouse fibrosarcoma cells
Authors: Murley Jeffrey S. , Baker Kenneth L. , Kataoka Yasushi , Weichselbaum Ralph R. , Grdina David J.
Published: February 2011

Exposure of cells to low non-lethal doses of ionizing radiation (≤ 10 cGy) or WR1065, the active free thiol form of amifostine, can induce pro-survival pathways that result in protection against the damaging effects of a 2 Gy dose of ionizing radiation. One such signaling pathway involves the elevation of active manganese superoxide dismutase (SOD2). SOD2 is a mitochondrial matrix protein that serves as the primary mitochondrial defense against superoxide formation. Its primary function is to facilitate the dismutation of two molecules of superoxide anion (O2−) produced by normal respiratory processes or following exposure to ionizing radiation into water and hydrogen peroxide. To characterize the role of SOD2 in the radiation- and thiol-induced adaptive responses, RKO36 human colon carcinoma cells and BFS2C mouse fibrosarcoma cells were exposed to 10 cGy x-rays and 40 μM or 4 mM WR1065 and SOD2 activity measured 24 h later. Significant increases in SOD2 activity in RKO36 cells (3.7-fold, P < 0.001) and BFS2C cells (3.0-fold, P = 0.005) were observed 24 h after exposure to 10 cGy. SOD2 activity was also observed to be significantly elevated in RKO36 cells (3.5-fold, P = 0.014 and 3.4-fold, P = 0.017) and BFS2C cells (3.7-fold, P = 0.007 and 2.5 fold, P = 0.021) 24 h after treatment with 40 μM or 4 mM WR1065, respectively. The protective effect of the low dose radiation- and thiol-induced elevation in active SOD2 was examined using the endpoint of micronuclei formation as a measure of chromosomal damage. Exposure of RKO36 and BFS2C cells to 2 Gy resulted in a significant increase in the mean number of micronuclei observed relative to unirradiated control cells (8.8 versus 2.2, P < 0.001; 5.4 versus 2.1, P < 0.001, respectively). The frequency of micronuclei formation was significantly reduced in both RKO36 cells (P = 0.003) and BFS2C cells (P = 0.011) exposed to 10 cGy 24 h prior to the 2 Gy dose. A significant reduction in micronuclei formation was also observed in RKO36 and BFS2C cells treated with 40 μM or 4 mM WR1065 30 min or 24 h prior to irradiation with 2 Gy. Treatment of both cell lines with SOD2 siRNA reduced SOD2 activity relative to mock-transfected control cells which resulted in the abrogation of both the radiation- and thiol-induced protective effect. This work was supported by NIH/NCI grant R01 CA132998 and DOE grant DE- PS02-08ER08-21 (D.J.G.).

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 507.
Abstract 5070: Identification of DKK2 as a candidate gene that facilitates tumor reoccurrence upon MYC inactivation
Authors: Adam Stacey J. , Xu Liwen , Karlsson Asa , Felsher Dean W.
Published: February 2011

The MYC oncogene is prevalently activated in human hematopoietic cancers, especially T-cell acute lymphoblastic leukemia (T-ALL) and Burkitt's lymphoma. Utilizing the Tet-system to generate a conditional model of MYC-induced T-ALL, our laboratory has demonstrated that MYC inactivation induces sustained tumor regression. However, after prolonged MYC inactivation some tumors eventually escape MYC dependence and reoccur. Our results support the hypothesis that tumors escape complete dependence on MYC by acquiring additional genetic or epigenetic changes, leading to altered gene function that may substitute totally or partially for MYC. Specifically, using Spectral Karyotypic Analysis (SKY) and array Comparative Genomic Hybridization (aCGH) we have obtained evidence that the Wnt/ β-catenin related gene, DKK2, may play a role in the mechanism by which tumors escape dependence upon MYC. We demonstrate DKK2 facilitates the ability of MYC-driven lymphomas to reoccur even in the presence of sustained oncogene inactivation. We have evidence that cellular context may play a pivotal role in DKK2's ability to aid tumor reoccurrence. Our data also suggest a mechanism by which DKK2 functions, alone or in conjunction with other Wnt/β-catenin pathway members, to maintain tumorigenesis or facilitate tumor reoccurrence in murine and human MYC-driven tumors in the presence of sustained MYC inactivation. Discovering that DKK2 overexpression may aid in lymphoma reoccurrence is a highly novel finding as DKK2 has never been implicated as an oncogene in lymphoma. Results presented here provide critical new insights into the mechanism by which MYC-driven cancers may obtain chemotherapeutic resistance and reoccur even in the presence of sustained drug therapy.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5070.
Abstract 5071: Defining the role of canonical MAPK signaling in glioma initiation and maintenance
Authors: Lastwika Kristin J. , Robinson James P. , VanBrocklin Mathew W. , Brandner Sebastian , Homen Sheri L.
Published: February 2011

Mutations in receptor tyrosine kinase (RTK) growth factor receptors e.g., EGFR, PDGFR, MET and ERBB2 which result in constitutive downstream signaling of the canonical mitogen activated protein kinase (MAPK) (e.g., RAS/RAF/MEK/ERK) and phosphatidylinositol 3-kinase (PI3K)/AKT pathways, are found in almost all World Health Organization (WHO) grade II, III, and IV astrocytomas. KRAS, BRAF and CRAF mutations occur in approximately 6%, 72% and 2% of low grade gliomas (WHO Grade I), respectively. A recent study also found a 5-fold increase in RAF serine/threonine kinase activity in a significant series of grade III and IV gliomas, coupled with a 12-fold increase in BRAF and a 2-fold increase in CRAF protein levels. By using an established glioma mouse model, it has been demonstrated that KRas, CRAF or BRAF expression in nestin expressing neural progenitor cells, combined with either Akt activation or Ink4a/Arf loss, leads to the development of high grade gliomas. Together, these data clearly demonstrate the importance of MAPK signaling in glioma development. However, KRAS, BRAF and CRAF are all differently regulated. To further validate a role for canonical MAPK signaling in glioma formation, we have evaluated the effect of constitutive MEK1 activity in glioma formation in vitro and in vivo. While expression of activated MEK1 alone is not sufficient for tumorigenesis, the combination of activated MEK1 and Akt, or MEK1 with Ink4a/Arf loss leads to glioma formation. To assess the efficacy of targeting MAPK signaling in gliomas, we treated a large panel human GBM cell lines with the MEK inhibitor PD0325901 alone and in combination with the PI3K/mTOR inhibitor BEZ235. Cell lines with the highest levels of ERK activation were the most sensitive to MEK inhibition while those with the highest levels of AKT activation were protected from apoptosis following MAPK inhibition. Treatment with BEZ235, in combination with PD0325901, caused significantly higher levels of apoptosis in the glioma cell lines. Importantly, neither treatment induced apoptosis in normal human astrocytes. Our studies demonstrate that activation and signaling through the MAPK pathway following Ink4a/Arf loss or Akt activation can give rise to high grade gliomas; moreover MAPK inhibition, when combined with PI3K/mTOR inhibition may prove to be an effective treatment for gliomas.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5071.
Abstract 5073: SIAH2 antagonizes the growth-inhibitory activity of the TGFβ signaling pathway and enhances the growth-promoting activity of the RAS signaling pathway during the genesis and progression of human cancer
Authors: Liao Yang , Edens Maryanne , Leof Edward B. , Tang Amy H.
Published: February 2011

A better understanding of the antagonistic interaction and cross-talk between the pivotal oncogenic and tumor suppressor signaling pathways, and the concerted cancer signaling networks that control tumorigenesis and metastasis should give rise to novel therapeutic interventions. Hyperactivated ERBB/RAS is known to drive neoplastic transformation, tumorigenesis and metastasis in a large percentage of human cancers, and thus finding novel measures to countervail activated EGFR/HER2/RAS signals to reverse tumorigenesis and metastasis remains an important goal in cancer biology. Instead of targeting an upstream signaling component, we targeted the most downstream signaling module in the ERBB/RAS pathway, the SIAH-dependent proteolytic machinery. SIAHs are evolutionarily conserved RING E3 ligases that are the critical downstream “gatekeeper” signaling modules in the RAS signal transduction pathway. We show that human SIAH E3 ligase is upregulated in proliferating tumor cells in human tumor specimens, and that proper SIAH function is required for RAS signal transduction. By inhibiting SIAH2 E3 ligase, we have blocked both tumorigenesis and metastasis in eleven of the most aggressive human cancer cell lines known in athymic nude mice. Our observations highlight the important roles that SIAH2 plays in RAS-dependent tumorigenesis and metastasis in human cancer. In this study, we aim to identify the molecular mechanism underlying SIAH2 deficiency-mediated growth inhibition in human cancer cells. We have found that SMAD2/3/4 - the essential downstream mediators of TGFβ signaling, are novel SIAH2 substrates in that SIAH2 physically interacts with, ubiquitinates and degrades SMAD2/3/4 (not SMAD1). The identification of SIAH2-mediated SMAD2/3/4 modulation may provide an additional molecular mechanism to explain the well documented, but less understood RAS-TGFβ antagonism described in normal development, tumorigenesis and metastasis. We propose that the tumor promoting activity of SIAH2 may be mediated through the degradation of SMAD2/3/4 proteins which enhancing the growth-promoting activity of the RAS pathway while suppressing the growth inhibitory action of the TGFβ pathway. Since the substrate specificity of ubiquitin-mediated proteolysis is primarily determined by the E3 ligase, our results provide new insights into the role of altered proteolysis in mediating the RAS-TGFβ antagonism and cross-talk observed in the genesis and progression of human cancers.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5073.
Abstract 5075: PI3K activation restores KrasG12D oncogenic ability in a mouse model of follicular thyroid cancer by directly opposing multiple Kras-initiated negative feedback signals
Authors: Dima Mariavittoria , Baker Kristen , Cristofano Antonio Di
Published: February 2011

Activating mutations of Ras family members and aberrant activation of the PI3K/AKT pathway are quite common in thyroid cancer originating from the follicular epithelium.

We have recently shown in a relevant in vivo model that, contrary to many other cell types, Kras activation is unable to induce oncogenic transformation of the thyroid follicular cells. However, PI3K activation is able to potently cooperate with Kras oncogenic mutation, thus inducing aggressive and metastatic thyroid follicular carcinomas. We now show that one key event mediated by PI3K in this cross-talk is the inhibition of multiple Kras-initiated negative feedback signals shutting down MAPK signaling and the induction of proliferation. Through these mechanisms, PI3K signaling restores Kras-initiated MAPK activation, induction of proliferation, and oncogenic transformation.

In particular, our data reveal that activation of p38α MAPK, as well as up-regulation of MKP2 and MKP3 phosphatases, play a pivotal role in mediating the growth-suppressive negative feedback initiated by Kras oncogenic activation.

Activation of p38α MAPK acts by inducing cell cycle arrest and the establishment of cellular senescence, while MKP2 and MKP3 stunt ERK1/2 activation through direct dephosphorylation of these kinases.

Strikingly, we have found that PI3K acts through Akt1 (but not Akt2) to potently suppress p38 phosphorylation and activation. Furthermore, PI3K activation results in the transcriptional repression of MKP2 and MKP3 promoters. Through these two molecular mechanisms, PI3K activation contributes to unleashing to its full extent Kras oncogenic ability.

These data not only uncover novel oncogenic pathways controlled by PI3K signaling, but also further strengthen the rationale for the dual targeting of PI3K and MAPK pathways in thyroid cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5075.
Abstract 5078: FGFR2 gene amplification status in gastric carcinoma and clinicopathologic significance
Authors: Jung Eun-Jung , Kim Woo Ho , Jung Eun-Ji , Kim Min A.
Published: February 2011

Background: Fibroblast growth factor receptor 2 (FRFG2) is a member of FGFR receptor tyrosine kinase family. FGFR2 gene amplification or missense mutation has been observed in various human cancers, including gastric carcinoma. Recent studies showed that, anti FGFR2 agents inhibit tumor progression in other tumors. Gastric carcinoma is one of the most frequent causes of cancer-related death worldwide. Evaluate the status of FGFR2 gene amplification in gastric carcinoma is beneficial in targeted cancer therapy.

Methods: We evaluated FGFR2 amplification status in the surgically resected 312 gastric carcinoma cases, using fluorescence in situ hybridization (FISH) and real-time quantitative-polymerase chain reaction (q-PCR), and compared the results. We compared clinicopathologic parameters with the presence of FGFR2 amplification, and performed survival analysis.

Results: Among 312cases, 14 cases (4.5%) showed FGFR2 amplification by FISH. FGFR2 amplification was associated to higher pT stage (p=0.015), pN stage (p=0.031) and distant metastasis (p=0.023). In survival analysis, FGFR2 amplification was significantly associated with lower cancer-specific survival in univariate analysis (p=0.012). The Real-time q-PCR results were concord with the FISH result.

Conclusions: Gastric carcinoma with FGFR2 amplification showed more advanced disease and poor survival. Evaluate the FGFR2 amplification status may identify a subset of cancers that are sensitive to targeted therapy using FGFR2 inhibitors.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5078.
Abstract 5079: Gadd45a differentially modulates breast tumorigenesis by promoting or suppressing tumor growth in response to distinct oncogenic stress
Authors: Tront Jennifer S. , Haung Yajue , Hoffman Barbara , Liebermann Dan
Published: February 2011

Gadd45a plays a pivotal role as a stress sensor that modulates cellular responses to various stress stimuli, including oncogenic stress. Generating novel MMTV-ras and MMTV-myc mouse models which are either wildtype or null for Gadd45a, we identified Gadd45a as a tumor suppressor in ras-mediated breast tumorigenesis. In sharp contrast, recent findings have shown that Gadd45a promotes myc-driven breast cancer. Furthermore, the function of Gadd45a as either a tumor promoter or suppressor was observed to be mediated through distinct, mutually exclusive, signaling pathways. Whereas Gadd45a tumor suppressor function is mediated via activation of JNK and p38 stress kinases promoting ras-induced apoptosis and senescence, Gadd45a function as a tumor promoter is exerted by negatively regulating MMP10 expression via the GSK3b/Beta-catenin signaling cascade, resulting in increased tumor vascularization. Analyzing specimens obtained from human breast cancer patients has shown relatively high levels of Gadd45a expression in low to moderate grade tumors compared to normal breast tissue, and no Gadd45a expression in high grade tumors. Correlating the status of Gadd45a expression in human breast cancer to activated oncogenes and Gadd45a signaling, as well as to patient survival following treatment is in progress. Collectively, these observations highlight the significance of the type of oncogenic alteration on how stress response genes function during initiation and progression of tumorigenesis.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5079.
Abstract 508: Synergistic effect of β-catenin knockdown and radiation combination toward AMC-HN9 head and neck cancer cells is associated with down-regulation of Ku expression.
Authors: Chang Hyo Won , Kim Hyo Jung , Moon So Young , Kim Mi Ra , Kim Seong Who , Kim Sang Yoon
Published: February 2011

Based on our previous report showing that up-regulation of Ku70/Ku80 in AMC-HN9 (HN9) cells is required for cell-cycle re-entry and cell proliferation following irradiation, further study to investigate the mechanism by which β-catenin knockdown sensitizes the cells to irradiation have been conducted. β-Catenin, as a multifunctional protein involved in cadherin-mediated cell-cell adhesion and Wnt signaling transduction, have been well known to regulates cell growth and survival following radiation in various types of cancer cells. However, the molecular mechanisms by which β-catenin affects radiation sensitivity are still not fully understood, which led us to explore, with regard to regulation of Ku70/Ku80 expression, the molecular mechanism of the synergistic effects of β-catenin silencing and radiation combination in radio-resistant head and neck cancer cells.

β-Catenin silencing using small interfering RNA(siRNA) down-regulated β-catenin expression up to 72 h both in membranous and nuclear fraction of HN9 cells. β-Catenin knockdown induced cell cycle arrest on G1 phase and thus decreased cell proliferation, but didn't cause any DNA damage response and cell death. Whereas expression of Ku70/Ku80 was up-regulated in HN9 cell following irradiation (4Gy), in the cells treated with combination of radiation and β-catenin siRNA, Ku70/Ku80 expression were dramatically decreased. In addition, when β-catenin silenced cells were exposed to single dose of radiation, irradiation-induced cell death was much more increased from 3.1% to 13% and accompanied with caspase-3 activation and PARP cleavage.

Taken together, these results suggest that inhibition of Ku expression through β-catenin silencing is associated with its radio-sentitizing effect in HN9 cells and further studies to clarify the pathway linking β-catenin signaling and regulation of Ku70/Ku80 expression are warranted.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 508.