bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2876: Farnesyltransferase inhibitor induces p53-dependent cellular senescence in colon cancer cells
Authors: Zhu Limin , Noh Jung Hyun , Geryk-Hall Mandy , Hughes Dennis P.
Published: August 2012

Farnesyltransferase inhibitors (FTIs), small molecules that inhibit prenylation of multiple proteins such as Ras and Rho, have clinical promise for cancer. FTIs block malignant growth and promote cell death in a wide variety of tumor types in vitro and in vivo, but the mechanisms underlying the antitumor activity of FTIs are not completely understood. We have observed that colon cancer cell line OS187 responds to the FTI tipifarnib with a senescent phenotype, demonstrated as altered cell morphology, elevated senescence-associated β-galactosidase activity, upregulated protein and mRNA levels of senescence markers, and long-term cell cycle arrest. Our previous results also indicated that cells retaining intact p53 are more sensitive to tipifarnib than are cells with absent/inactive p53, suggesting a possible role for p53 in the senescent response to tipifarnib. To test this hypothesis, we used shRNA to reduce p53 expression in OS187 cells. Loss of p53 reversed the swollen and enlarged senescent-like cellular phenotype induced by tipifarnib. In addition, p53 knockdown cells showed less senescence-associated β-galactosidase activity than did scrambled control cells after 7-day treatment with tipifarnib. Cell viability assay suggested that p53 knockdown partially rescued the cytotoxic effects of tipifarnib at low concentrations. Western blot also demonstrated that cell cycle regulatory proteins such as p27, p21, and p15 were significantly upregulated in scrambled control cells while only a slight increase of these proteins were observed in p53 knockdown cells after tipifarnib treatment. Flow cytometry analysis showed that cells with decreased p53 level had a reduced percentage of cells in sub G1 phase when treated with a high concentration of tipifarnib, and fewer cells arresting in G2/M phase when treated with low concentration of tipifarnib compared to the scrambled control cells. This observation suggested that p53 may involve in different pathways depending on the strength of the stress signal imposed by different concentration of tipifarnib. In conclusion, these results confirm that p53 is important for inducing senescence when farnesylation is inhibited in OS187 cells. However, since p53 knockdown has very limited impact on the anti-proliferation effects of tipifarnib at high concentrations, other pathways are expected to play a role in mediating cell susceptibility to tipifarnib. Future experiments will focus on investigating the p53-independent pathways that are responsible for tipifarnib-induced senescence and apoptosis.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2876. doi:1538-7445.AM2012-2876
Abstract 3020: The role of NF-κB (p65) and Stat3 interplay in the regulation of radiation-induced, ICAM-1-mediated migration and invasion in glioma
Authors: Kesanakurti Divya , Maddirela Dilip , Chetty Chandramu , Rao Jasti S.
Published: August 2012

Aberrant activation of both NF-κB and Stat3 is associated with enhanced tumor metastasis in various cancers. In human glioma xenograft cells 4910 and 5310, irradiation (IR; 8 Gy) elevated Stat3 and p65 phosphorylation and persistent nuclear translocation in correlation with massive increases in cellular and soluble ICAM-1 levels. Co-immunoprecipitation experiments confirmed enhanced nuclear phospho-p65/phospho-Stat3 and acetyl-p65/phospho-Stat3 interactions along with enhanced invasiveness and migration in IR-treated cells. Promoter analysis indicated presence of adjacent NF-κB- and Stat3-consensus binding elements in the proximal promoter and the proximal intron-1 of the ICAM-1 promoter, whereas overlapping NF-κB/Stat3 binding motifs were identified in the distal intron-1 region. ChIP binding and promoter/luciferase reporter assays showed that IR induced prominent binding of both Stat3 and NF-κB to the proximal intron-1 region followed by the distal intron-1 and the proximal promoter regions on the ICAM-1 gene. Individual treatments with Stat3.si or JSH-23 blocked the occupancy of both Stat3 and p65 onto the ICAM-1 promoter and prevented subsequent ICAM-1-mediated migration and invasion. In addition, constitutive activation of Stat3 and NF-κB by Stat3C or TNF-α significantly elevated IR-induced migration and invasion. In vivo orthotropic experiments in nude mice showed that the Stat3.si treatment reduced tumor growth and IR-induced phopsho-Stat3/phospho-p65 and ICAM-1 expression when compared to pSV-treated controls. These results suggest that both p65 and Stat3 bind to the intronic region of ICAM-1 promoter to induce its optimal transcription in IR-treated cells and abrogation of IR-induced p65/Stat3 binding inhibits ICAM-1-mediated enhancement of malignancy in post-irradiated tumor cells. As such, the results of this study clearly have potential future therapeutic significance in glioma treatment.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3020. doi:1538-7445.AM2012-3020
Abstract 3026: Increase in proliferation and migration of breast cancer cells via PKC activation-induced down-regulation of BKca channel
Authors: Park Eun Sil , Mun Yun-Ja , Kang Kee Ryeon , Han Jaehee , Kang Dawon
Published: August 2012

Purpose: K+ channels are involved in tumorigenesis through the regulation of cell proliferation, apoptosis, migration, and invasion. However, the involvement of the large conductance Ca2+-activated K+ (BKCa) channels in the cancerous phenomenon is controversial. This study was performed to investigate whether and how BKCa channels are related to the cellular processes for tumorigenesis. Methods: Cell proliferation and migration assays were performed in non-invasive (MCF-7) and invasive (MDA-MB-231) breast cancer cells in the absence or presence of BKCa channel modulators or PKC modulators. Patch-clamp technique and reverse-transcriptase (RT)-polymerase chain reaction (PCR) were performed to study the expression of the BKCa channel in breast cancer cells. The relationship between BKCa channels and cell proliferation/migration was analyzed using gene overexpression and knock-down techniques. Results: Patch-clamp technique and RT-PCR data showed that BKCa channels were expressed in both MCF-7 and MDA-MB-231 cells. Inhibitors of BKCa channels increased cell proliferation and migration, whereas BKCa channel activators suppressed them. Overexpression of BKCa in breast cancer cells reduced proliferation and migration, whereas silencing of BKCa enhanced the proliferation and migration. BKCa inhibitors (paxilline, ChTx, and IbTx) induced PKC activation, and the PKC activation down-regulated the BKCa channel expression and activity. In addition, the treatment of PMA (a PKC activator) decreased BKCa channel expression, whereas combination of PKC inhibitors maintained the BKCa channel expression levels without reduction. Conclusion: These results show that a decrease in BKCa expression levels, which could be modulated by PKC activation, increases cell proliferation and migration in breast cancer cells and suggest that modulation of BKCa expression may regulate tumorigenesis of breast cancer cells. Key words: Breast cancer; Cell migration; K+ channels; Proliferation

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3026. doi:1538-7445.AM2012-3026
Abstract 3030: Hypoxia induces BACH1 expression regulating colon cancer metastasis
Authors: Lee Jin Young , Jo Yeong Nang , Kim Jin-Ah , Lee Chang Woo , Lee Ok-Jun , Han Sang-Bae , Park Song-Kyu , Kang Jong Soon , Yun Jieun
Published: August 2012

BACH1 (BTB and CNC homology 1) is a basic region-leucine zipper (bZip) transcription factor that binds to MAREs (Maf recognition elements) and acts as a repressor of genes that regulate the oxidative stress response. It has been reported that BACH1 is a novel let-7 target and regulates breast cancer metastasis. Here we show that hypoxia induces BACH1 expression by regulating BACH1 promoter activity. In a mouse xenograft model, BACH1 depletion reduces the primary tumor growth of HCT-116 colorectal cancer cells. In addition, knockdown of BACH1 significantly decreases the invasive efficiency of colorectal cancer cells. Finally, we confirm that BACH1 expression is high in the patient samples of colorectal cancer and metastasis, unlike normal colon tissues. These results suggest BACH1 as a novel target for cancer therapeutic strategy.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3030. doi:1538-7445.AM2012-3030
Abstract 3033: Identification of the MUC16/CA125 interaction network: Results of a high-throughput yeast two-hybrid screening
Authors: Polterauer Stephan , Thapi Dharmarao , Schultz Nikolaus , Ouerfelli Ouathek , Chen Nancy , Rosales Nestor , Yan Xiu , Spriggs David R.
Published: August 2012

Background: MUC16 is a well-known serum marker for the diagnosis of ovarian cancer. MUC16 is involved in oncogenic transformation and is associated with migration, invasion, and tumor growth. Due to the large size of this mucin, the interacting partners involved in signaling and transformation have not been investigated. Materials and Methods: We performed a high-throughput yeast two-hybrid screening to investigate putative protein interactions with MUC16. A construct of the MUC16 carboxyterminus, lacking the transmembrane domain (≥mC89), was cloned into the activation domain of the yeast vector pGBKT7. Three different ovarian cancer cDNA prey libraries were tested against the bait μmC89. Of all blue yeast colonies cDNA prey-inserts were analysed with PCR and plasmids were recovered from yeast cultures. Basic Local Alignment Search Tool was used to identify corresponding genes in the GenBank Database. Gene expression- and correlation analyses of human ovarian cancer data of The Cancer Genome Atlas (TCGA) were performed. Immunoprecipitation analyses and magnetic immonoassays were performed to confirm potential interaction partners in human ovarian cancer cell lines. Results: Screening of approximately 30,000,000 colony-forming units identified 238 yeast colonies that maintained promoter activation when cultivated in selective media. PCR and sequencing of cDNA inserts of cultivated yeast colonies nominated 84 candidates as putative binding partners for MUC16. Analysis of TCGA data did not identify direct linkage between mRNA expression of MUC16 and the nominated partners in human ovarian cancers. Based on this analysis and on their biological function, six partners were chosen for further confirmatory analyses: AMHR2, eEF1α, EGF, GAL1, PPP2CB, and SEPT2. Results of the confirmation analyses are pending. Our findings suggest a role of these proteins in regulating MUC16 signaling and transformation mechanisms. Conclusions: We nominate six novel potential partners for interaction with the MUC16 carboxyterminus (≥mC89). These proteins might be involved in ovarian cancer signaling and transformation and might be useful novel targets for vaccines, antibodies and drug inhibitors for the treatment of ovarian cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3033. doi:1538-7445.AM2012-3033
Abstract 3040: Terbinafine inhibits oral squamous cell carcinoma growth through anti-cancer cell proliferation and anti-angiogenesis
Authors: Chien Ming-Hsien , Yang Shun Fa , Lee Wen Sen
Published: August 2012

ABSTRACT Terbinafine (TB), an oral antifungal agent used in the treatment of superficial mycosis, has been reported to exert an anti-tumor effect in various cancer cells. However, the effect of TB on oral cancer has not been evaluated. Herein we demonstrated that TB (0-60 μM) concentration-dependently decreased cell number in cultured human oral squamous cell carcinoma (OSCC), KB cells. The anti-proliferation effect of TB was also observed in two other OSCC cell lines, SAS and SCC 15. TB (60 μM) was not cytotoxic and its inhibition on KB cell growth was reversible. [3H]thymidine incorporation and flow cytometric analyses revealed that TB inhibited DNA synthesis and induced the G0/G1 cell cycle arrest. The TB-induced cell cycle arrest occurred when the cyclin-dependent kinase 2 (CDK2) activity was inhibited just as the protein levels of p21cip1 and p27kip1 were increased. The TB-induced G0/G1 cell cycle arrest was completely blocked when the expressions of p21cip1 and p27kip1 were knocked-down together. Taken together, these results suggest that the p21cip1- and p27kip1-associated signaling pathways might be involved in the TB-induced anti-proliferation in KB cells. In vivo, TB (50 mg/kg, i.p.) significantly inhibited the KB tumor size. In these TB-treated tumors, increases in the levels of p21cip1 and p27kip1 protein and decreases in the number of proliferating cell nuclear antigen (PCNA)-positive cells and the microvessel density (MVD) were observed. These findings demonstrate for the first time that TB might have potential to serve as a therapeutic tool in the treatment of oral cancer. 1

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3040. doi:1538-7445.AM2012-3040
Abstract 3047: MK1775, a selective Wee1 inhibitor shows antitumor activity against sarcoma cells
Authors: Kreahling Jenny M. , Foroutan Parastou , Gemmer Jennifer Y. , Martinez Gary , Reed Damon , Letson Douglas , Bui Marilyn , Gillies Rober J. , Altiok Soner
Published: August 2012

Sarcomas are rare and heterogeneous mesenchymal tumors affecting both pediatric and adult populations with more than 70 recognized histologies. Doxorubicin and ifosfamide have been the main course of therapy for treatment of sarcomas; however, the response rate to these therapies is about 10-20% in metastatic setting. Toxicity with the drug combination is high, response rates remain low, and improvement in overall survival, especially in the metastatic disease, remains negligible. Accordingly, new agents are needed for the treatment of this heterogeneous group of diseases. Wee1 is a critical component of the G2/M cell cycle checkpoint control and mediates cell cycle arrest by regulating the phosphorylation of CDC2. Inhibition of Wee1 by a selective small molecule inhibitor MK1775 has been reported to to enhance the cytotoxic effect of DNA damaging agents. However, its role in the treatment of mesenchymal tumors has not been explored. In this study we investigated the therapeutic efficacy of MK1775 in various sarcoma cell lines and patient-derived tumor explants ex vivo both alone and in combination with other chemotherapeutic drugs frequently used in the treatment of sarcomas. Our data demonstrate that MK1775 treatment as a single agent at clinically relevant concentrations leads to unscheduled entry into mitosis and initiation of apoptotic cell death in all sarcomas tested. Additionally, MK1775 significantly enhances the cytotoxic effect of the DNA damaging agents, gemcitabine and doxorubicin. In MK1775 treated cells CDC2 activity was enhanced and G2/M ceheckpoint was impaired as assessed by increased expression of phosphorylated histone H3, a marker of mitotic entry. The cytotoxic effect of Wee1 inhibition on sarcoma cells appears to be independent of p53 status as all sarcoma cell lines with different p53 mutation were highly sensitive to treatment. Additionally, in patient-derived bone and soft tissue sarcoma samples we showed that MK1775 in combination with gemcitabine causes significant apoptotic cell death suggesting that this treatment may represent a novel approach in the treatment of sarcomas. In patient-derived xenograft mouse models of sarcoma we are currently analyzing the therapeutic efficacy of MK1775 in vivo by utilizing magnetic resonance imaging (MRI) and diffusion MRI methods. Our results together with the high safety profile of MK1775 strongly suggest that this drug can be used as a potential therapeutic agent in the treatment of both adult as well as pediatric sarcoma patients.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3047. doi:1538-7445.AM2012-3047
Abstract 3052: Role of p53 in cdk inhibitor VMY-1-103-induced apoptosis in prostate cancer
Authors: Ringer Lymor , Sirajuddin Paul , Yenugonda Venkata M. , Brown Milton , Rodriguez Olga , Albanese Chris
Published: August 2012

The mammalian cell cycle is a tightly regulated process that controls cell division and replication. The tumor suppressor p53 plays a crucial role in both the G1/S and mitotic checkpoints, as well as in apoptosis in response to DNA damage. P53 activates p21, which is an inhibitor of cdk/cyclin E complexes, to cause cell cycle arrest. P53 can also function as a mediator of apoptosis by activating either the instrinsic or extrinsic pathway of apoptosis by activating transcriptional targets such as Bax, Bid, and TRAIL R4/5. In prostate cancer (PCa), p53 mutations are frequently associated with advanced staged disease. We recently have developed a novel analog of the CDK inhibitor purvalanol B, termed VMY-1-103 (VMY) which functions as a potent inhibitor of proliferation. To assess the efficacy of this compound in vitro, we tested the ability of VMY to cause cell cycle arrest and apoptosis in LNCaP, DU145, and PC3 prostate cancer cell lines. VMY caused an increase in cells in G2/M in all cell lines tested. Additionally, VMY treatment resulted in a large amount of apoptosis in LNCaP cells, which are p53 wild-type. In contrast, DU145 cells, which are p53 mutant, and PC3 cells, which are p53 null, were significantly less susceptible to VMY-mediated apoptosis, strongly suggesting that functional p53 was required for VMY to induce apoptosis in PCa cells. In order to test this hypothesis, a series of experiments were carried out in these PCa cell lines. Knockdown of p53 by siRNA in LNCaP cells resulted in a decreased sensitivity to VMY, as these cells no longer underwent cell cycle arrest or apoptosis. Further modulation of p53 was performed by overexpressing a wild-type construct in PC3 cells, which greatly sensitized the cells to cell death following VMY treatment. Conversely, introduction of several mutant p53 constructs into LNCaP cells did not decrease sensitivity to the drug whatsoever. Importantly, in DU145 cells, which have two mutations of p53 on opposite alleles, apoptosis was significantly increased when cells were pre-treated with PRIMA-1 to restore WT p53 function. Therefore, our data establishes p53 plays a crucial regulator of sensitivity to VMY and its ability to induce apoptosis in prostate cancer cells. In addition we show that modification of mutant p53 by PRIMA in a p53 deficient cell lines, and the ensuing restoration of drug sensitivity, enhances the potential of VMY as a novel therapeutic for prostate cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3052. doi:1538-7445.AM2012-3052
Abstract 3062: Ada3, a component of STAGA HAT complex, controls cell cycle by regulating c-myc
Authors: Mohibi Shakur , Gurumurthy Channabasavaiah , Mirza Sameer , Nag Alo , Katafiasz Bryan , Bele Aditya , Naramura Mayumi , Band Hamid , Band Vimla
Published: August 2012

The human homologue of the yeast Ada3 (alteration/deficiency in activation) protein is an essential component of various Histone acetyl transferase complexes (HATs). To gain insight into the physiological importance of Ada3, we established a conditional knockout mouse model and showed that loss of Ada3 led to embryonic lethality at an early stage of development. Using the derived Ada3lox/lox mouse embryonic fibroblast (MEF) cell lines we show that Ada3 deletion causes defects in histone acetylation. Also we show that Ada3 enhances catalytic activity of p300 HAT. We further showed that Ada3 plays an important role in cell proliferation; depletion of Ada3 from cells caused cell lethality. This proliferation defect was reversed by introduction of human Ada3 in Ada3 null MEFs. Ada3 null MEFs also showed delay in transition from G0-G1 phase of the cell cycle to the S phase. These cells also manifested decreased hyperphosphorylation of Retinoblastoma protein (Rb); an increased protein half life of CDK inhibitor p27Kip1 and decrease in Cdk2 kinase activity. The increase in p27 protein was a result of decreased Skp2 protein and mRNA levels which is a direct target of c-myc oncogene. We further show that mRNA levels of c-myc, a direct transcriptional target of Ada3 containing STAGA HAT complex, are reduced on deletion of Ada3. These data suggest an Ada3-myc-skp2-p27 pathway that controls mammalian cell cycle. Taken together, our data suggests an essential role of Ada3 in embryonic development and cell cycle progression.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3062. doi:1538-7445.AM2012-3062
Abstract 3119: A synthetic lethal screen identifies genetic determinants for gemcitabine sensitivity in pancreatic cancer
Authors: Landry Jerome C. , Pantazides Brooke G. , Park Wungki , Shelton Joseph W. , Maithel Shishir K. , El-Rayes Bassel , Yu David S.
Published: August 2012

Pancreatic cancer is notorious for its devastating disease course and prognosis. Gemcitabine is a widely used regimen for pancreatic cancer treatment but lack of response and resistance often limits its use. To identify the genes and mechanisms involved in determining gemcitabine sensitivity in pancreatic cancer, we completed a loss of function genetic screen using RNA interference to identify genes which when silenced are synthetically lethal with a low dose of gemcitabine in human pancreatic cancer cells. We optimized an assay using ATR or CHK1 siRNA as positive controls and ATM or non-targeting (NT) siRNA as negative controls. Our siRNA library included 4,474 siRNAs corresponding to 1,006 unique human genes arrayed in a one-gene:one-well format in 96-well plates. Genes chosen for our library consisted predominantly of nuclear enzymes, including but not limited to kinases, phosphatases, ubiquitinylation proteins, sumoylation proteins, acetylases, deactylases, nucleases, helicases, and chromatin modifiers, which we reasoned were more likely to function directly in DNA repair processes and be targetable. MIA PaCa-2 cells were transfected with 25 nM siRNA and treated 48 hours later with or without 13 nM gemcitabine for 72 hours prior to the cell proliferation assay using WST-1 reagent. Using this assay, we identified 87 genes which when silenced caused gemcitabine sensitization and 42 genes which when silenced caused gemcitabine resistance. Positive hits included 16 known genome maintenance genes including well-characterized ATR signaling pathway genes - CHK1, RAD9, HUS1, and CDC25A, 5 putative ATM/ATR substrates, and 20 genes identified in previously published DNA damage sensitivity screens. 21 of the genes are somatically mutated or differentially expressed in pancreatic cancer, suggesting that our gemcitabine sensitivity genes may function as useful predictive biomarkers or novel targets for the treatment of pancreatic cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3119. doi:1538-7445.AM2012-3119
Abstract 3121: Systematic analysis of the DNA synthesis fidelity of somatic mutations of polymerase beta found in prostate cancer
Authors: Chen Desheng , LeCompte Kimberly , Pursell Zachary , Makridakis Nick
Published: August 2012

Human DNA polymerase beta (polβ) is a monomeric protein of 335 amino acids and functions in base excision repair and meiotic recombination. Human polα is over-expressed in many cancer cells like ovarian, prostate, breast, colon and chronic myelogenous leukemia. To obtain clues of possible functions in vivo, we have determined the fidelity of DNA synthesis of polα with forward mutation assay which scored the sequence errors caused in the lacZ-α gene in M13mp2 during DNA synthesis to fill a 407-nucleotide gap. Three of seven of the somatic polα mutants, p.E232K, p.E123K and pE216K, which we identified previously in prostate tumors, were tested. The results indicated that the somatic mutants of human DNA polymerase beta display altered fidelity. The fidelity for large deletions and complex changes of the pE232K and pE123K muatants are three and 31 times lower respectively, compared to wild type polα. The T>G transversion at position 103, which is the most common feature of polB base substitution error spectrum, is observed six times and two times lower for the pE123K and p.E216K mutants than wild polymerase beta. Our data show that somatic mutations in prostate cancer change both the synthesis fidelity and/or mutation type compared with wild type polymerase beta.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3121. doi:1538-7445.AM2012-3121
Abstract 3132: Dynamic changes of epigenetic abnormalities during initiation and progression of adult T-cell leukemia/lymphoma (ATLL)
Authors: Oka Takashi , Al-Kader Lamia Abd , Sato Hiaki , Washio Kana , Shinnou Yoko , Takata Katsuyoshi , Murakami Ichiro , Utsunomiya Atae , Yoshino Tadashi
Published: August 2012

HTLV-1 causes ATLL in 3-5% of infected individuals after a long latent period of 40 to 60 years. ATLL is divided into four stages: namely, smoldering, chronic, lymphoma and acute types. The smoldering and chronic types are indolent, but the acute and lymphoma types are aggressive ATLL characterized by resistance to chemotherapy and a poor prognosis. Such a long latent period suggests that a multi-step leukemogenic/lymphomagenic mechanism is involved in the development of ATLL, although the critical events in the progression have not been characterized. To determine whether epigenetic abnormalities are playing the important role in the progression of ATLL, we analyzed the methylation profiles, showing that number of CpG island methylated genes increased with disease progression and aberrant hyper-methylation in specific genes was detected even in HTLV-1 carriers and correlated with progression to ATLL. The CpG island methylator phenotype (CIMP) was observed most frequently in lymphoma type ATLL and was also closely associated with the progression and crisis of ATLL. The high number of methylated genes and increase of CIMP incidence were shown to be unfavorable prognostic factors and correlated with a shorter overall survival with the Kaplan-Meyer analysis. Dynamic changes of aberrant DNA methylation status in the specific genes were observed during the progression of an ATLL patient. The present findings strongly suggest that the multi-step accumulation of aberrant CpG methylation in specific target genes and the presence of CIMP are deeply involved in the initiation and progression of ATLL not only epidemiologically but also in the clinical course of a specific ATLL patient.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3132. doi:1538-7445.AM2012-3132
Abstract 3134: Integrin α6β4 upregulates EGFR ligands in pancreatic cancer cells
Authors: Carpenter Brittany L. , Chen Min , O'Connor Kathleen L.
Published: August 2012

Because of late diagnosis and lack of effective treatments, pancreatic cancer is the 4th leading cause of cancer death in the US, with a death to incidence ratio that is nearly equal. In an effort to improve patient survival, there needs to be a focus on molecular mechanisms that confer a metastatic phenotype. The integrin α6β4 is overexpressed in pancreatic carcinoma and plays a role in cancer progression by promoting tumor invasion and metastasis as well as cell survival. We find that integrin α6β4 overexpression leads to upregulation of cancer promoting genes such as amphiregulin and epiregulin, ligands of the epidermal growth factor receptor (EGFR). The purpose of this study is to address how integrin α6β4 upregulates these ligands. We have identified large CpG islands within the amphiregulin and epiregulin promoters (-1500 to +500 bp from TSS) using CpG plot and CpG Island searcher. This provides a potential site for epigenetic modifications, which are essentially reversible, and therefore an ideal target for treatment. To test whether integrin α6β4 contributes to this regulation through DNA methylation pancreatic cancer cells expressing low levels of α6β4 were treated with DNA methyltransferase inhibitor 5-aza-2′deoxycitidine. Using Q-PCR, our results show that inhibition of DNA methyltransferase increases amphiregulin and epiregulin expression at the mRNA level. In contrast, when cells with high levels of integrin α6β4 were treated with the hypermethylating agent, S-adenosylmethionine, it leads to a decrease in amphiregulin and epiregulin expression. This implicates α6β4 as a regulator of this process allowing us insight into how this pathway might be regulated. Using ELISA, we also provide evidence that integrin α6β4 promotes increased autocrine secretion of amphiregulin after promoter demethylation. This is of particular interest to our lab because autocrine secretion of amphiregulin and epiregulin make it possible for cancer cells to both produce and respond to their own growth factors, promoting cellular invasion and metastasis. This study indicates that overexpression of integrin α6β4 in pancreatic cancer is associated with the hypomethylation of amphiregulin and epiregulin promoters, which may play an important role in the development of a malignant phenotype.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3134. doi:1538-7445.AM2012-3134
Abstract 3145: Smyd3 catalyzes a novel methylation mark and regulates cancer cell proliferation
Authors: VanAller Glenn S. , Reynoird Nicolas , Barbash Olena , Huddleston Michael J. , Le BaoChau , Annan Roland S. , Garcia Ben A. , Tummino Peter J. , Gozani Or , Kruger Ryan G.
Published: August 2012

SET and MYND domain-containing protein 3 (Smyd3) is a lysine methyltransferase over-expressed in liver, breast and rectal carcinomas. We have shown that knockdown of Smyd3 has an anti-proliferative effect in cancer cell lines and that this effect can be rescued with wild type but not catalytically inactive Smyd3, directly linking Smyd3 catalytic activity to proliferation. Published reports indicate Smyd3 methylates lysine 4 of histone H3, lysine 20 of histone H4 and VEGFR1. However, activity testing of a peptide library and intact recombinant nucleosomes indicate that a unique methylation mark is catalyzed by Smyd3. LC-MS/MS analysis of in vitro reaction products have identified the location of this novel site of methylation. The existence of this novel mark and its dependence on Smyd3 activity was confirmed in cancer cells and mouse embryonic fibroblasts. We speculate that overexpression of Smyd3 causes aberrant methylation and plays an important role in the deregulation of gene expression leading to uncontrolled cellular proliferation in cancer making Smyd3 a promising target for anti-cancer therapy.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3145. doi:1538-7445.AM2012-3145
Abstract 315: Immune cells promote hepatocellular carcinoma stemness phenotype
Authors: Wan Shanshan , Vatan Linda , Simeone Diane M. , Kryczek Ilona E. , Zou Weiping , Welling Theodore H.
Published: August 2012

Hepatocellular carcinoma (HCC) is a common cause of cancer-related deaths and recurrence rates can be as high as 50% following resection. Evidence in several tumor types suggests that cancer stem cells (CSCs) contribute to tumorigenicity, therapeutic resistance and recurrence even in the presence of anti-tumor immunity. Furthermore, CSC phenotype may be influenced by tumor microenvironment. Therefore, we aimed to determine whether HCC stemness phenotype may be influenced by immune cell subsets. We have previously identified CD44 as a HCC-CSC marker as evidenced by enhanced tumorigenicity and sphere forming capacity in CD44+ HCC cells. To determine the effects of immune cells on HCC stemness, transwell co-culture experiments were performed using GFP-labeled HepG2 and T cells or monocytes/macrophages. The percentage of CD44+ HCC cells were increased during co-culture with anti-CD3/anti-CD28 activated T cells from donor PBMCs (n=9), suggesting that immune effector cells could enrich CD44+ HepG2 cells. In addition, CD44+ HCC cells were enriched during co-culture with HCC-associated macrophages (TAM) (n=3 HCC patients) or monocytes from healthy donor PBMCs (n=11). CD44+ HCC cells were positively correlated with TAM in primary HCC patients (R=0.69, n=12, p<0.05), suggesting a potential role of TAM in promoting HCC-CSCs. Concordantly, sphere formation capacity was increased in HepG2 cells following co-culture with monocytes. Furthermore, co-culture with HepG2 elevated IL-6 and IL-8 produced by monocytes. IL-6 and IL-8, when added exogenously to HepG2 cells, synergistically expanded CD44+ HCC cells and promoted sphere formation capacity. In conclusion, HCC stemness phenotype is expanded by T cells, HCC associated macrophage and monocytes, as well as cytokines, IL-6 and IL-8. These findings suggest that immune cell derived factors in the tumor microenvironment may contribute to HCC recurrence or progression by promoting a stemness phenotype.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 315. doi:1538-7445.AM2012-315
Abstract 3156: MicroRNA profiling for the detection of melanoma
Authors: Qu Kevin Z. , Liu Yan , Zhang Ke , Owen Renius X. , Sferruzza Anthony D.
Published: August 2012

Background: Numerous reports have suggested a link between aberrant expression of microRNAs (miRNAs) and various forms of cancer, including melanoma. We examined miRNA expression patterns in melanoma and nevi to identify candidate biomarkers for differentiating between these melanocytic lesions. Method: The study used formalin-fixed paraffin-embedded (FFPE) tissues from 78 patients with melanoma and 98 with nevi (18 compound nevi, 20 intradermal nevi, 20 junctional nevi, 20 blue nevi, and 20 spitz nevi). All lesions were confirmed by at least 2 pathologists. Total RNA was extracted from the FFPE tissue and hybridized onto the Affymetrix GeneChip miRNA array (Affymetrix, Santa Clara, CA), which measure 850 human microRNAs. After determining an optimal set of miRNAs for differentiation using random forest algorithm, we retested specimens for this set using quantitative real-time PCR (qPCR) on a TaqMan miRNA system (Applied Biosystems, Foster City, CA). Results: Expression levels of 257 miRNAs differed significantly between melanomas and nevi (p values < 0.001); 129 miRNAs were downregulated and 128 were upregulated in melanomas relative to nevi. 80 of these miRNAs exhibited area under the ROC curves (AUROCs) of >0.75 for distinguishing melanomas from nevi, and 74 exhibited more than a two-fold difference. An optimized set of 7 miRNAs provided good differentiation of melanomas from nevi, with an AUORC of 0.94 and 87.7% concordance with pathology-determined tissue type. miRNA array data were further validated by qPCR: for 5 of the 7 miRNAs identified above, qPCR results showed significant correlation with the miRNA array results (average Pearson correlation coefficient = 0.95). Conclusions: These data suggest that the miRNA expression pattern from FFPE tissue holds promise for distinguishing melanomas from nevi. Use of this miRNA set as a diagnostic marker requires further validation and exploration in a large-scale study.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3156. doi:1538-7445.AM2012-3156
Abstract 3165: microRNA-148a: A novel oncogenic microRNA in glioblastoma
Authors: Kim Jungeun , Zhang Ying , Guessous Fadila , Abounader Roger
Published: August 2012

Glioblastoma is the most common and deadly malignant brain tumor. One main reason of resistance of glioblastoma to therapy is the ability of the tumor cells to migrate and invade the surrounding brain, rendering total surgical removal of the tumor practically impossible. MicroRNAs (miRNA) are small noncoding regulatory RNA molecules with emerging critical roles in cancer. MicroRNAs modulate protein expression by binding to the 3-untranslated region of mRNA and promoting RNA degradation or inhibiting transcription. In this study, we analyzed data from The Cancer Genome Atlas (TCGA) and found that microRNA-148a (miR-148a) was one of few miRNAs that inversely correlated with GBM patient survival. However, nothing is known about the role of this miRNA in cancer and GBM. We therefore started a systematic study of miR-148a in GBM. We first measured miR-148a levels in GBM and found that they were > 3-fold higher in GBM stem cells than in normal astrocytes and ∼15 times higher in tumor specimens than in and normal brain. We then investigated the functional effects of miR-148a on GBM malignancy. Inhibition of miR-148a in GBM stem with anti-miR-148a strongly induced apoptosis. Overexpression of miR-148a in GBM stem cells strongly promoted their self-renewal ability. Transfection of pre-miR-148a into GBM cells strongly promoted cell proliferation, migration and invasion though a membrane coated with collagen IV, a commonly expressed matrix protein in GBM. Therefore, miR-148a acts as an oncogene in GBM cells by promoting cell proliferation, migration and invasion and inhibiting stem cell differentiation. To elucidate the mode of action and targets of miR-148a in GBM, we used a combined approach of miRNA target prediction analysis and immunoblotting to identify oncogenes that are targeted and inhibited by miR-148a. We selected proteins that are known to play roles in GBM and cancer, and were able to confirm inhibition by miR-148a of MIG6 and BIM. MIG6 is a negative regulator of EGFR (an important oncogene in GBM), and BIM is a central pro-apoptotic molecule. Therefore, we predicted that miR-148a might act as an oncogenic microRNA by inducing EGFR expression and activation and inhibiting apoptosis. Using immunoblotting, we demonstrated that miR-148a does in fact induce EGFR expression and activation in GBM cells. Ongoing and future studies will assess the in vivo effects of miR-148a and evaluate its usefulness as a new target for GBM therapy. Altogether, we have uncovered miR-148a a new oncogenic microRNA in GBM that acts via regulation of EGFR and BIM.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3165. doi:1538-7445.AM2012-3165
Abstract 3173: Identification of miR-4423 as a primate-specific microRNA highly expressed in airway epithelium and associated with lung cancer
Authors: Perdomo Catalina , Campbell Joshua , Gerrein Joseph , Tellez Carmen , Gower Adam , Vick Jessica , Garrison Carly , Anderlind Christina , Jackson George R. , Schembri Frank , Gomperts Brigitte N. , Hayden Patrick , Belinsky Steven A. , Lenburg Marc E. , Lenburg Marc E. , Spira Avrum
Published: August 2012

Smoking is a significant risk factor for lung cancer, the leading cause of cancer-related deaths worldwide. Our group has previously shown that epithelial gene expression is altered throughout the airway of smokers and that some of these changes are regulated by microRNAs. Moreover, we have previously identified gene expression differences in cytologically normal bronchial airway epithelial cells between smokers with and without lung cancer that can serve as an early diagnostic biomarker for lung cancer. Here, we use next-generation sequencing of small RNAs to identify novel microRNAs expressed in airway epithelium and associated with lung cancer. We identify miR-4423 as a primate-specific microRNA highly expressed in the airway epithelium. In vitro, the expression of miR-4423 increases as Normal Human Bronchial Epithelial cells are differentiated into mucociliary epithelium at an Air Liquid Interface, while its mRNA targets decrease in expression. Furthermore, the expression of miR-4423 is reduced in lung tumors and in the cytologically normal bronchial airway epithelium of smokers with lung cancer. In gain-of-function experiments, ectopic expression of miR-4423 in lung cancer cell lines resulted in reduced colony formation in soft agar. Taken together, these data support the power of next-generation sequencing in identifying novel cell type- specific transcripts and provides evidence that this newly characterized microRNA may play a role in promoting the differentiation and/or maintenance of airway epithelium, and can reduce anchorage-independent lung cancer cell growth.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3173. doi:1538-7445.AM2012-3173
Abstract 3210: Glutamine synthetase is a genetic determinant of cell-type specific glutamine independence in breast epithelia
Authors: Kung Hsiu-Ni , Marks Jeffrey R. , Chi Jen-tsan Ashley
Published: August 2012

Although significant variations in the metabolic profiles exist among different cells, little is understood in terms of genetic regulations of such cell-type specific metabolic phenotypes and nutrient requirement. While many cancer cells depend on exogenous glutamine for survival to justify the therapeutic targeting of glutamine metabolism, the mechanisms of glutamine dependence and likely response and resistance of such glutamine-targeting strategies among cancers are largely unknown. In this study, we have found a systematic variation in the glutamine dependence among breast tumor subtypes associated with mammary differentiation: basal but not luminal-type breast cells are more glutamine dependent and may be susceptible to glutamine-targeting therapeutics. Glutamine independence of luminal-type cells is associated mechanistically with lineage-specific expression of glutamine synthetase (GS). Luminal cells can also rescue basal cells in co-culture without glutamine, indicating a potential for glutamine symbiosis within breast ducts. The luminal-specific expression of GS is directly induced by GATA3 and represses glutaminase expression. Such distinct glutamine dependency and metabolic symbiosis is coupled with the acquisition of the GS and glutamine independence during the mammary differentiation program. Understanding the genetic circuitry governing distinct metabolic patterns is relevant to many symbiotic relationships among different cells and organisms. In addition, the ability of GS to predict patterns of glutamine metabolism and dependency among tumors is also crucial in the rational design and application of glutamine and other metabolic pathway targeted therapies.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3210. doi:1538-7445.AM2012-3210
Abstract 3212: N-acetylcysteine protects melanoma cells from glucose depletion through unequal modulation of SOD1 and Copper chaperone for SOD1 depending on p53 mutational status
Authors: Chavez-Perez Valery , Strasberg-Rieber Mary , Rieber Manuel
Published: August 2012

BACKGROUND.-Cancer cells from solid tumors undergo metabolic stress, when nutrients become limiting, within poorly vascularized regions. Severe deprivation of glucose and serum-derived growth factors can promote cell death by metabolic stress, partly through excessive reactive oxygen species (ROS) production.OBJECTIVES.- To investigate whether survival in response to metabolic stress is influenced by N-acetylcysteine (NAC), an anti-oxidant glutathione precursor, and whether it is linked to modulation of superoxide dismutases (SOD1 and SOD2) that catalyze intracellularly the conversion of O2− to H2O2. EXPERIMENTAL- matched human melanoma cells with unequal p53 mutational status were exposed to overnight depletion of glucose with either 0.5% or 10% serum supplementation. Differential cell survival was assesed by crystal violet staining. Mechanisms influencing response to glucose limitation were studied by specific immune blotting. RESULTS.- A 24 hour glucose depletion suppressed cell survival in 0.5% dialyzed serum but this was counteracted by 2 mM NAC irrespective of cellular p53 status. A shorter glucose depletion in 0.5% serum revealed normoxic induction of hypoxia-inducible factor (HIF1α) reciprocal with concomitant down-regulation of SOD1 together with the Copper chaperone for SOD1 (CCS) and intact PARP, with no comparable effect on mitochondrial SOD2. Under these restrictive conditions, NAC addition partly restored SOD1 and PARP but not CCS expression in mutant p53(R175H) cells. No comparable NAC restoration of these proteins was evidenced in matched wt p53 melanoma under glucose depletion in 0.5 % dialyzed serum. However, SOD1 but not CCS was highest following glucose deprivation with 10% dialyzed serum in wt p53 melanoma cells, compared to identically treated mutant p53 cells. NAC addition increased CCS irrespective of p53 status with glucose depletion in 10% serum, and preferentially induced SOD1 in mutant p53 cells under comparable conditions. CONCLUSIONS.-human melanoma cells are protected from transient glucose depletion by anti-oxidants like NAC through differential modulation of SOD1 and CCS depending on serum availability and p53 mutational status.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3212. doi:1538-7445.AM2012-3212