bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 4227: The ribonucleotide reductase inhibitor Didox reverses tamoxifen resistance in breast cancer cells
Authors: Shah Khyati N. , Elford Howard L. , Faridi Jesika S.
Published: May 2015

Tamoxifen is the most prescribed endocrine therapy for estrogen receptor positive breast cancer patients. The emergence of endocrine resistance is a major hurdle in achieving a successful clinical outcome. Many advances have been made over the past few years to understand the biology of tamoxifen resistance. Activation of the AKT kinase is one of the primary mechanisms of intrinsic tamoxifen resistance. Previously, we reported that the Ribonucleotide Reductase M2 (RRM2) subunit is upregulated in AKT induced tamoxifen resistance and that inhibition of RRM2 by siRNA mediated interference reduces tamoxifen induced cell proliferation. In this study, we report that the inhibition of RRM2 by the small molecule inhibitor of ribonucleotide reductase activity Didox (3, 4-dihydroxybenzohydroxamic acid), significantly reduced tamoxifen induced cell proliferation in AKT overexpressing cells and tamoxifen resistant tumors generated by these cells. As well, Didox in combination with tamoxifen also inhibited cell proliferation in acquired tamoxifen resistant breast cancer cell lines. Employing comprehensive cell culture and in vivo models, we demonstrate that combining tamoxifen with Didox may reverse tamoxifen resistance in AKT expressing breast cancer cells.

Citation Format: Khyati N. Shah, Howard L. Elford, Jesika S. Faridi. The ribonucleotide reductase inhibitor Didox reverses tamoxifen resistance in breast cancer cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4227. doi:10.1158/1538-7445.AM2014-4227
Abstract 4236: Antitumor activity of a novel (-)-antofine analog in human lung cancer cells and paclitaxel-resistant lung cancer cells
Authors: Song Jayoung , Bae Song Yi , Shin Yoonho , Kim Won Kyung , Oh Jedo , Choi Tae Joon , Jeong Eun Ju , Park So Hyun , Jang Eun Jeong , Kang Ji In , Park Hyen Joo , Hong Ji-Young , Chung Hwa-Jin , Kwon Yongseok , Kim Sanghee , Lee Sang Kook
Published: May 2015

Our previous study revealed that (-)-antofine, a phenanthroindolizidine alkaloid from Cynanchum paniculatum Kitagawa (Asclepiadaceae), exhibited a potent growth inhibition activity against A549 human lung cancer cells. Based on the information several analogs were designed and synthesized to procure new entities. A novel (-)-antofine C-13a analog ((R)-4b) was found to be a potential growth inhibition and antitumor activities in A549 cells. Treatment with (R)-4b for 24 hours did not result in the induction of apoptotic cell death but moderately induced cell cycle arrest in the G0/G1 phase and inhibited the expression of cyclin D1, cyclin E, CDK4 and pRb. In addition, (R)-4b suppressed the phosphorylation of mTOR and its downstream effectors such as 4E-BP-1 and eIF4E. The paclitaxel-resistance was also overcome in human lung cancer cells (A549-Pa) via the suppression of p-glycoprotein expression. Furthermore, (R)-4b significantly inhibited the tumor growth of A549 and A549-Pa xenografts in a nude mouse model. Taken together, these findings suggest that (R)-4b might be a potential candidate for the development of cancer chemotherapeutic agents derived from natural products.

Citation Format: Jayoung Song, Song Yi Bae, Yoonho Shin, Won Kyung Kim, Jedo Oh, Tae Joon Choi, Eun Ju Jeong, So Hyun Park, Eun Jeong Jang, Ji In Kang, Hyen Joo Park, Ji-Young Hong, Hwa-Jin Chung, Yongseok Kwon, Sanghee Kim, Sang Kook Lee. Antitumor activity of a novel (-)-antofine analog in human lung cancer cells and paclitaxel-resistant lung cancer cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4236. doi:10.1158/1538-7445.AM2014-4236
Abstract 4244: Inhibition of HER2 and IGF2 triggers cell death in trastuzumab-resistant HER2 positive JIMT1 cells
Authors: Helu Xousaen M. , Leon Daisy De
Published: May 2015

Breast cancer is the second leading cause of death in women. Overexpression of Human Epidermal Growth Factor Receptor 2 (HER2) is detected in 20-30% of breast cancers. Increased levels of HER2 are linked to poor prognosis, more aggressive phenotype, and resistance to chemotherapy and hormonal treatment. HER2 is part of a four member family of cell surface receptors that are implicated in transmission of signals controlling cell growth and differentiation. HER2 has an intracellular domain with tyrosine kinase catalytic activity and dimerization of HER2 activates the MAP kinase and AKT pathways. HER2 receptor also heterodimerizes with the IGF1 receptor(IGF1R) in trastuzumab-resistant cell lines. The insulin-like growth factor 2 (IGF2) has an important role in fetal and cancer development signaling through the IGF1R, insulin receptor (InsR), and cross-talk with the estrogen receptor alpha and beta (ERα, ERß). Previous findings in our lab demonstrate an increased expression of IGF2 in trastuzumab-resistance cell line JIMT1 when compared to trastuzumab-sensitive cell lines BT474, SKBR3, and ZR75-30. We hypothesis that to overcome trastuzumab-resistance in JIMT1 cell lines, dual inhibition of HER2 and IGF2 is necessary to induce cell death. When JIMT1 cells were treated with resveratrol, both HER2 and IGF2 mRNA and protein levels were reduced and a decrease in cell viability was observed. We also analyzed the phosphorylation levels of HER2 and IGF1R. Our studies suggest that blocking both HER2 and IGF2 is necessary to decrease cell viability in trastuzumab-resistant JIMT1 cells.

Citation Format: Xousaen M. Helu, Daisy De Leon. Inhibition of HER2 and IGF2 triggers cell death in trastuzumab-resistant HER2 positive JIMT1 cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4244. doi:10.1158/1538-7445.AM2014-4244
Abstract 4272: Applying TCGA data for breast cancer diagnostics and pathway analysis
Authors: Andersen Jannik , Shah Parantu , Korski Konstanty , Ibbs Matthew , Filas Violetta , Kosiedowski Michal , Pukacki Juliusz , Mazurek Cezary , Wu Yuanqing , Chang Edward , Toniatti Carlo , Draetta Giulio , Wiznerowicz Maciej
Published: May 2015

The main objective of our project is to obtain a comprehensive insight into oncogenic signaling in order to develop novel diagnostic tools for molecular subtypes of breast cancer (BRCA). We have provided about 100 BRCA samples along with full pathology and clinical annotations to the TCGA program. These cancer samples have been extensively characterized using all available genomics platforms and the obtained molecular data contributed to the molecular characterization of BRCA. This integrated effort identified four major molecular subtypes of breast cancer that can be identified across five profiling platform including genomics, transcriptomics and proteomics (Nature, 490 (7418):61-70). Building upon these datasets we aim to translate these genomics profiles into diagnostic tools that can be used in every day medical practice. In the first step, we have generated tissue microarrays (TMAs) from FFPE BRCA samples obtained from the patients enrolled into the TCGA project. Next, the TMA slides were used for immunohistochemistry (IHC) using about 100 antibodies specific for major cancer markers, including oncogenic kinases frequently activated or overexpressed in BRCA. The obtained IHC results have been scored and verified independently for each marker by two board-certified pathologists. To identify markers that correlate with or are specific to the molecular subtypes of BRCA, the TMA/IHC read-outs from each tumor sample were correlated with all TCGA genomic data. In addition, we integrate the pathology and genomics results with clinical data obtained from the TCGA-enrolled patients, including up to 4 years follow-up. Finally, we apply system biology tools to integrate the genomic data from TCGA with the proteomic analysis to understand causalities between changes in DNA, transcriptome and signal transduction pathways. Our long-term goal is to identify novel diagnostic biomarkers that will precisely identify each molecular subtype of BRCA and which may be predictive of patient response to therapy thus paving the way for novel personalized therapies for cancer.

Citation Format: Jannik Andersen, Parantu Shah, Konstanty Korski, Matthew Ibbs, Violetta Filas, Michal Kosiedowski, Juliusz Pukacki, Cezary Mazurek, Yuanqing Wu, Edward Chang, Carlo Toniatti, Giulio Draetta, Maciej Wiznerowicz. Applying TCGA data for breast cancer diagnostics and pathway analysis. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4272. doi:10.1158/1538-7445.AM2014-4272
Abstract 4279: Rapid biomarker discovery using large-scale, patient-derived cancer genomic cohorts
Authors: Sanga Sandeep , Nair Praveen , Mirsaidi Cyrus , Broudy Thomas
Published: May 2015

Target discovery and validation in oncology has largely relied on molecular and functional studies performed in cell lines. Recent advances in genomics have now created large databases based on well-characterized tumor tissue, which has enabled direct investigation of patient tumors for novel targets and predictive markers. Following these discoveries, it is routine to perform functional studies in cell line-based systems; however, it is often challenging to find a relevant cell line model and if found, there are often numerous factors which confound biology when using historical cell lines for functional studies. The result can be a process, which takes considerable time and does not readily translate to clinical relevance.

We report here our efforts to identify novel targets and markers through data mining large cohorts of cancer genomics data consisting of samples deriving from both drug-naïve and -treated patient-derived cancer samples. To serve as the drug-naïve reference, we look to the thousands of patient-derived tumor specimens, covering 30+ tissue types, genomically characterized by The Cancer Genome Atlas. For both drug-naïve and drug-treated primary data, we leverage Molecular Response's proprietary bank of viable cryopreserved tumor cells. The bank contains more than 144,000 tumor specimens, covering 25 tissue types and 76 clinical diagnoses. Each of the specimens has been profiled against a panel of drugs ex vivo, and levels of resistance recorded. Nearly 400 tumor specimens from Molecular Response's proprietary bank have been genomically characterized.

Station X has designed and developed a software environment called GenePool for the management, analysis and communication of genomic information for cohort-scale biomarker studies. The genomics data along with sample-associated clinical metadata deriving from Molecular Response's bank and The Cancer Genome Atlas were imported into GenePool. We evaluated consistency between drug-naïve samples deriving from the Molecular Response bank and TCGA, and subsequently mined for expression-based markers differentiating varying grades of drug sensitivities from drug-naïve specimens. We are currently evaluating these markers for further functional study in patient-derived models and plan to report to these findings.

Citation Format: Sandeep Sanga, Praveen Nair, Cyrus Mirsaidi, Thomas Broudy. Rapid biomarker discovery using large-scale, patient-derived cancer genomic cohorts. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4279. doi:10.1158/1538-7445.AM2014-4279
Abstract 4286: Improved FFPE DNA extraction for next generation sequencing using adaptive focused acoustics technology
Authors: Lin Ling , Vega Mikenah , Jones Robert T. , Ziaugra Liuda , Dolcen Deniz N. , Sunkavalli Ashwini , Schubert Laura , Lepine Jack R. , Thorner Aaron R. , Ducar Matthew D. , Hahn William C. , Meyerson Matthew L. , MacConaill Laura E. , Hummelen Paul Van
Published: May 2015

Formalin fixation followed by paraffin embedding (FFPE) is the most widely used method of tissue preservation and often used in the clinical setting for cancer specimens. However, this process of fixation has negative consequences on the quality of extracted DNA, and inevitably adversely affects downstream analyses such as next generation sequencing (NGS). DNA extraction from FFPE samples requires paraffin disassociation, tissue rehydration and de-crosslinking. Low efficiency in any step will yield low quantities of nucleic acids with incomplete reversal of formaldehyde crosslinks, and mixtures of single and double stranded DNA. In addition, low quality nucleic acids will often result in poor sequencing quality due to inefficient library preparation, lower number of reads with high quality, and variable recovery of sequence reads with high GC-content.

Here we describe the use of Adaptive Focused Acoustics technology (Covaris Inc. 2013) to remove paraffin and break down tissue, and we compared this method to traditional xylene-based paraffin removal. We found that Covaris had superior performance overall: (1) DNA yields were 6 times higher, and less specimens failed extraction: this is critically important when dealing with small tumor biopsies. (2) DNA was more readily available for PCR amplification. (3) Mean coverage of DNA fragments with high GC content was higher and more reproducible. (4) Library complexity was higher. In addition, the Covaris protocol required less hands-on time in the lab, and eliminated the need for hazardous organic solvents for paraffin removal.

In conclusion, the Covaris FFPE DNA extraction method is very efficient, enhances DNA availability, improves reproducibility and provides higher DNA quality for NGS.

Citation Format: Ling Lin, Mikenah Vega, Robert T. Jones, Liuda Ziaugra, Deniz N. Dolcen, Ashwini Sunkavalli, Laura Schubert, Jack R. Lepine, Aaron R. Thorner, Matthew D. Ducar, William C. Hahn, Matthew L. Meyerson, Laura E. MacConaill, Paul Van Hummelen. Improved FFPE DNA extraction for next generation sequencing using adaptive focused acoustics technology. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4286. doi:10.1158/1538-7445.AM2014-4286
Abstract 43: Podoplanin in head and neck squamous cell carcinoma: Prognostic value and clinicopathological implications
Authors: Koo Bon Seok , Lim Kyu
Published: May 2015

Background: Podoplanin is a mucin-type transmembrane glycoprotein that is important in lymphangiogenesis. We investigated the associations of podoplanin expression status with clinicopathological variables and the prognostic influence of podoplanin expression in head and neck squamous cell carcinoma (HNSCC), to determine its effectiveness as a marker of high-risk HNSCC subjects.

Patients and Methods: We analyzed by immunohistochemistry podoplanin expression and its associations with the clinicopathological characteristics of 119 surgically treated HNSCC cases. Additionally, we investigated the effects of podoplanin expression on overall survival and disease-specific survival in univariate and multivariate analyses.

Results: Of 119 HNSCC patients, high podoplanin expression was found in 61 cases (51.3%) and low expression in 58 (48.7%). High podoplanin expression was associated with lymph node metastasis, advanced AJCC stage, and poor histological grade (p < 0.05). In multivariate analysis, high podoplanin expression was significantly associated with circa three- and fivefold increases in the presence of positive lymph node metastasis and poor histological grade, respectively (p < 0.05). High levels of podoplanin expression were significantly associated with decreased overall survival and disease-specific survival (p < 0.05).

Conclusions: Our findings suggest that high podoplanin expression is associated with aggressive tumor behavior and poor prognosis in HNSCC.

Note: This abstract was not presented at the meeting.

Citation Format: Bon Seok Koo, Kyu Lim. Podoplanin in head and neck squamous cell carcinoma: Prognostic value and clinicopathological implications. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 43. doi:10.1158/1538-7445.AM2014-43
Abstract 430: Identification of novel methylation-silenced genes in oral squamous cell carcinomas
Authors: Abe Masanobu , Yamashita Satoshi , Mori Yoshiyuki , Abe Takahiro , Saijo Hideto , Ushijima Toshikazu , Takato Tsuyoshi
Published: May 2015

To identify novel methylation-silenced genes in oral squamous cell carcinomas (OSCCs), we took a combined approach of methylated DNA immunoprecipitation (MeDIP)-CpG island (CGI) microarray analysis and expression microarray analysis after treatment with a demethylating agent. MeDIP-CGI microarray revealed that promoter CGIs of 324 genes were methylated in two OSCC cell lines (Ca9-22, HSC-2). By treatment of the cell lines with 5-aza-2′-deoxycytidine, 125 of the 324 genes were up-regulated in at least one of the two cell lines. For 23 of the 125 genes that have not been focused on in the field of oral malignancy, we analyzed their promoter methylation statuses in 16 normal oral tissues, five OSCC cell lines, 26 OSCC tissues and 24 oral leukoplakia tissues. Ten promoter CGIs were methylated not only in OSCC cell lines but also OSCC tissues. The methylation profile indicated the presence of the CpG island methylator phenotype (CIMP) in OSCC tissues. Furthermore, the CIMP was observed in three of the 13 oral dysplastic leukoplakia tissues. The CIMP was considered as one of early molecular events in malignant transformation from oral premalignant lesions to OSCCs.

Citation Format: Masanobu Abe, Satoshi Yamashita, Yoshiyuki Mori, Takahiro Abe, Hideto Saijo, Toshikazu Ushijima, Tsuyoshi Takato. Identification of novel methylation-silenced genes in oral squamous cell carcinomas. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 430. doi:10.1158/1538-7445.AM2014-430
Abstract 4305: Biodistribution and clearance of non-targeted DyLight dyes in tumor-free nude mice for in vivo imaging
Authors: NLEND Marie C. , Desai Surbhi , Hong Suk J. , Anderson Mary Beth , Los Georgyi V. , Hermanson Greg T. , Diener Justin M. , Leevy Warren M. , Bell Peter A.
Published: May 2015

Far red to near infra red (NIR) fluorophores are commonly used in cell-based assays and for deep tissue in vivo imaging. The light emitting and physico-chemical characteristics of these fluorophores, e.g. excitation and emission spectra, relative hydrophilicity/hydrophobicity and molecule size, can significantly influence their performance in these applications. Currently there are no guidelines for selection of fluorophores optimal for targeted in vivo imaging. To address this issue we have designed a panel of DyLight dyes with maximum excitation wavelength in the range of 650 -785 nm, maximum emission wavelength in the range of 670 - 800 nm and different levels of sulfonation and/or PEGylation. We studied the kinetic of tissue distribution and clearance of these fluorophores as well as their effect on morphology of different tissues in mice after intravenous injection. Briefly, nude mice were anesthetized, injected with a dye (100 µL of dye at 0.5mg/ml in PBS) into retro-orbital plexus, and imaged using Carestream MSFX or Carestream XTREME Imager. Images were generated before dye injection, immediately after injection (0 hours) and 3, 6, 12 and 24 hours post injection. At the end of the experiment, animals were sacrificed; internal organs were collected and used for ex vivo analysis. The results of in vivo imaging experiments demonstrated that: hydrophobic fluorophores have a relatively slow kinetic of distribution and clearance, and often cleared via a hepatic pathway. Introduction of the negatively charged groups and or PEG chains increased dye solubility affected the kinetics of both fluorophore distribution and its clearance. In addition, negatively charged fluorophores had a tendency to be cleared predominantly via renal pathway. A histological analysis of the internal organs showed no detectable tissue damage. These results provide guidelines for the selection of optimal NIR dyes for in vivo imaging.

Citation Format: Marie C. NLEND, Surbhi Desai, Suk J. Hong, Mary Beth Anderson, Georgyi V. Los, Greg T. Hermanson, Justin M. Diener, Warren M. Leevy, Peter A. Bell. Biodistribution and clearance of non-targeted DyLight dyes in tumor-free nude mice for in vivo imaging. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4305. doi:10.1158/1538-7445.AM2014-4305
Abstract 4306: Bioconjugation of near infrared dyes by PEGylation improves pharmacokinetics and allows for better labeling and imaging of human gastrointestinal cancers
Authors: Maawy Ali A. , Hiroshima Yukihiko , Luiken George A. , Zhang Yong , Hoffman Robert M. , Bouvet Michael
Published: May 2015

NIR dyes are becoming increasingly popular due to their good tissue penetration, low tissue autofluorescence and resistance to hemoglobin quenching. However, they are still limited by lower hydrophilicity and poor chemical and photo-stability in-vivo. We aimed to try to improve these properties by covalent modification of these dyes by addition of polyethylene glycol (PEG) chains. We studied 2 commercially-available NIR dyes, DyLight 650 and 750, to evaluate how PEGylation affects the pharmacokinetics, biodistribution and imaging. PEGylated and non-PEGylated DyLight 650 and 750 dyes were conjugated to a chimeric anti-CEA antibody and were injected intravenously into nude mice. Serum samples were collected at various time points to determine serum concentrations and clearance kinetics. Biodistribution was determined by tissue sonication and analyzing tissue concentration profiles over time. PEGylated dyes had significantly higher serum concentrations than non-PEGylated dyes with p=0.005 for the 650 dyes and p<0.001 for the 750 dyes. PEGylated dyes had significantly lower liver concentrations (p=0.03 for the 650 dyes; p=0.002 for the 750 dyes) and higher renal concentrations (p=0.003 for the 650 dyes and p<0.001 for the 750 dyes) compared to non-PEGylated dyes. Human pancreatic tumors subcutaneously implanted into nude mice were labeled with antibody-dye conjugates and serially imaged. Labeling with PEGylated dyes resulted in significantly brighter tumors compared to the non-PEGylated dyes (p<0.001 for the 650 dyes; p=0.01 for 750 dyes). In a liver metastasis model of colon cancer, PEGylated dyes showed bright labeling of metastatic tissue with good contrast between normal and malignant tissue. PEGylation of the NIR dyes DyLight 650 and 750 significantly changes their pharmacokinetics and biodistribution, allowing higher serum concentrations, longer half-life, brighter tissue labeling and decreased accumulation in lymphoid organs. This allows for high fidelity imaging of metastatic disease to any lymphoid or hemoglobin dense organ such as the liver.

Citation Format: Ali A. Maawy, Yukihiko Hiroshima, George A. Luiken, Yong Zhang, Robert M. Hoffman, Michael Bouvet. Bioconjugation of near infrared dyes by PEGylation improves pharmacokinetics and allows for better labeling and imaging of human gastrointestinal cancers. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4306. doi:10.1158/1538-7445.AM2014-4306
Abstract 4314: Metabolic manipulation of hypoxia and radiotherapy response by electron transport inhibitors
Authors: Gandolfo Maria Jose , Kyle Alastair H. , Minchinton Andrew I.
Published: May 2015

The presence of hypoxia in solid tumors is correlated with poor treatment outcome. Even small population of hypoxic cells within tumors can significantly decrease the efficacy of radiation therapy. One approach to treating hypoxia in tumours is to reduce overall tumor cell oxygen consumption by modulating mitochondrial respiration. This could reoxygenate hypoxic cells within the tumor and restore radiation sensitivity. To guide in vivo application we tested the effects of three respiratory chain inhibitors; the biguanides metformin and phenformin and rotenone on oxygen consumption in 3D tissue engineered discs.

Experiments were carried out to evaluate the effect of respiratory inhibition on the hypoxic fraction of HCT116 and HT29 tissue discs using immunostaining with the hypoxic cell marker pimonidazole. An oxygen sensing phosphorescence lifetime probe assessed pO2 on the basal side of the disc as a function of time after inhibitors were exposed to the apical surface. Clonogenic assays were then performed to confirm the radiosensitization of hypoxic cells within the tissue discs.

Pimonidazole staining and direct pO2 sensing confirmed that all three electron transfer inhibitors decrease the hypoxic fraction by reducing cellular oxygen consumption. Rotenone was found to completely eliminate the hypoxic fraction at nanomolar concentrations compared to metformin and phenformin which require milli to micromolar concentrations respectively. Furthermore, rotenone was found to have an effect on oxygen utilization within minutes of drug exposure compared to both metformin and phenformin, which took hours to be effective. In addition, clonogenics assays determined that all three electron transfer inhibitors radiosensitize tumor cells with rotenone being the most efficient compound on a molar basis.

The mitochondrial inhibitors of complex I metformin, phenformin and rotenone decrease oxygen utilization by tumor cells in a concentration dependent manner resulting in reduction of the hypoxic fraction and increased radiosensitivity.

Citation Format: Maria Jose Gandolfo, Alastair H. Kyle, Andrew I. Minchinton. Metabolic manipulation of hypoxia and radiotherapy response by electron transport inhibitors. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4314. doi:10.1158/1538-7445.AM2014-4314
Abstract 4316: BNIP3 suppresses mammary tumorigenesis and metastasis through negative regulation of Warburg effect and HIF-1α
Authors: Chourasia Aparajita Hoskote , Tracy Kristin , Boland Michelle , Sharifi Marina , Macleod Kay F.
Published: May 2015

BNIP3 is a hypoxia-inducible protein involved in mitochondrial dynamics and mitophagy that is up-regulated at pre-malignant stages of various human cancers, including breast cancer but down-regulated at later stages of tumorigenesis. To examine the role of BNip3 in tumor progression, we crossed BNip3 null mice to the MMTV-PyMT mouse model of breast cancer. Loss of BNip3 promoted primary tumor growth in vivo and tumor cell proliferation in vitro and was associated with increased aerobic glycolysis and reduced mitochondrial respiration. BNip3 null tumor cells also showed increased invasive properties that were cell intrinsic and transplantable to wild-type host mice. Consistently, there was reduced latency to lung metastasis in the MMTV-PyMT;BNip3-/- mice and increased numbers of metastases compared to control mice. BNip3 null tumors exhibited increased mitochondrial mass, but a greater fraction of mitochondria were dysfunctional with reduced membrane potential and decreased metabolite uptake compared to BNip3 wild-type tumor cells. Consistently, increased proliferation of BNip3 null tumor cells was associated with increased carbon flux from glucose to lipid and nucleic acids. Interestingly, although mitochondria were less efficient at uptake of pyruvate and glutamine, the rate of production of Krebs cycle intermediates was increased. This indicates that defective mitochondria due to inefficient mitophagy can induce the Warburg effect, as originally proposed by Warburg himself. Loss of BNip3 also resulted in more hypoxic tumors that expressed increased levels of Hif-1α and its target genes and showed increased angiogenesis, although blood vessels were abnormal possibly explaining increased tumor hypoxia, thus suggesting a negative feedback loop between Hif-1α and its target gene, BNip3. Inhibition of glycolysis or inhibition of HIF activity reduced proliferation in BNip3 null tumor cells to the levels observed in control cells. Significantly, BNip3 null tumor cells underwent autophagic flux as efficiently as wild-type cells demonstrating for the first time that inhibition of mitophagy has a distinct tumorigenic consequence to general inactivation of autophagy, as reported elsewhere. These results demonstrate that BNip3 has tumor and metastasis suppressor properties required to maintain mitochondrial integrity, promote oxidative metabolism and mitigate against the metastasis promoting activities of hypoxia and glycolytic metabolism.

Citation Format: Aparajita Hoskote Chourasia, Kristin Tracy, Michelle Boland, Marina Sharifi, Kay F. Macleod. BNIP3 suppresses mammary tumorigenesis and metastasis through negative regulation of Warburg effect and HIF-1α. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4316. doi:10.1158/1538-7445.AM2014-4316
Abstract 4322: The landscape of mitochondrial DNA mutations in human cancer
Authors: Ju Young Seok , Alexandrov Ludmil , Gerstung Moritz , Martincorena Inigo ,   ICGC Breast Cancer Group; ICGC Chronic Myeloid Disorders Group; ICGC Prostate Cancer Group, Stratton Mike , Campbell Peter J.
Published: May 2015

Recent sequencing studies have extensively explored the somatic alterations present in the nuclear genomes of human cancers. Although mitochondria in cancer cells have important roles in energy metabolism and survival, the origins and impact of somatic mutations in mitochondrial DNA (mtDNA) are unclear. Here, we provide a comprehensive analysis of somatic alterations in cancer mitochondria from 1,675 genome sequences across 31 tumor types. We identified 1,907 somatic substitutions which were mainly C:G>T:A and T:A>C:G. These mutations exhibited dramatic replicative strand bias, being almost all C>T and A>G on the mitochondrial heavy strand. This strand-asymmetric mitochondrial mutational signature is different from those found in nuclear cancer genomes but is similar to processes shaping human mtDNA sequence over evolutionary time. The number of mtDNA mutations showed considerable heterogeneity across tumor types, being significantly higher in tumors from gastrointestinal and hepatobiliary systems. With respect to selection, observed missense mutations were nearly neutral and often gradually drifted towards homoplasmy over time. In contrast, mutations resulting in protein truncation showed evidence of negative selection and were more frequently heteroplasmic. Our findings indicate that the endogenous mutational mechanism has far greater impact than any other external mutagens in mitochondria, and is fundamentally linked to mtDNA replication.

Citation Format: Young Seok Ju, Ludmil Alexandrov, Moritz Gerstung, Inigo Martincorena, ICGC Breast Cancer Group; ICGC Chronic Myeloid Disorders Group; ICGC Prostate Cancer Group, Mike Stratton, Peter J. Campbell. The landscape of mitochondrial DNA mutations in human cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4322. doi:10.1158/1538-7445.AM2014-4322
Abstract 4334: Greater resistance of mutant p53 tumor cells to hypoxia or chloroquine with glucose or pyruvate supplementation is diminished under glucose starvation
Authors: Rieber Manuel
Published: May 2015

BACKGROUND. Poorly vascularized solid tumors are exposed to regions of low oxygen and glucose deprivation. These restrictions may promote either defensive autophagy or cell death through excessive reactive oxygen species (ROS), mediated by activated NADPH oxidases (NOX) which transport electrons from NADPH on the cytoplasmic side of plasma membranes to reduce molecular oxygen at the extracellular space, increasing superoxide anion radical (O2•-). .OBJECTIVES. To investigate in genetically-matched human melanoma cells with wt or mutant p53, responses to: a) autophagy inhibitor chloroquine and to glucose-lowering metformin or to hypoxia in glucose-supplemented medium; b) pyruvate in glucose-free medium under normoxia or hypoxia; c) sub-physiological glucose. EXPERIMENTAL. Responses to normoxia or hypoxia were assesed by live-dead fluorometric assays, or by crystal violet staining of surviving cells. Mechanisms influencing response to glucose limitation were studied by specific immune blotting. RESULTS. Lower susceptibility to chloroquine ± metformin involving greater basal AMPK activation was seen in physiological 5 mM glucose in mutant p53 cells. These cells were also preferentially protected from prolonged hypoxia by glucose supplementation or by pyruvate supplementation of glucose-free medium under normoxia, but not under hypoxia. Sub-physiological glucose without pyruvate induced apoptosis-associated PARP cleavage irrespective of p53 status. However, treatment with impermeable exogenous superoxide dismutase (SOD) to remove extracellular superoxide signalling across cell membranes, preferentially counteracted PARP cleavage only in wt p53 cells with lower LDHA expression after 48 hr in 2.5 mM glucose. CONCLUSIONS. Glucose availability increases resistance to hypoxia or to chloroquine ± metformin in mutant p53 cells. However, glucose starvation promotes toxicity linked to extracellular (O2•-), preferentially in mutant p53 cells.

Citation Format: Manuel Rieber. Greater resistance of mutant p53 tumor cells to hypoxia or chloroquine with glucose or pyruvate supplementation is diminished under glucose starvation. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4334. doi:10.1158/1538-7445.AM2014-4334
Abstract 4345: Dysregulated miR-205 signaling promotes symmetric division of self-renewing mammary stem cells and mammary tumorigenesis
Authors: Wu Meng-Ju , Chao Chi-Hong , Chang Chao-Ching , Chang Chun-Ju
Published: May 2015

Epigenetic control plays a key role in regulation of tumorigenesis in response to the microenvironmental stimuli, however, the regulatory mechanism involved in the process is largely unclear. We now report that a critical epigenetic regulator miR-205 is repressed by Jagged1, a ligand shown to be secreted from the tumor stroma, to promote the cancer stem cell phenotype. Loss of miR-205 in mammary epithelial cells leads to epithelial-mesenchymal transition, disrupted epithelial cell polarity, and enhanced symmetric division to expand the stem cell population. Furthermore, mice deficient of miR-205 spontaneously develop mammary lesions, and activation of miR-205 significantly diminishes the breast cancer stemness. These data provides the first evidence to link microenvironment and microRNA regulation to the disrupted epithelial polarity and aberrant stem cell division, leading to an expansion of the stem cell population and tumorigenesis. Together, this study elucidates a new role for miR-205 in regulation of stem cell fate, providing a promising therapeutic target to eradicate the genesis of breast cancer.

Citation Format: Meng-Ju Wu, Chi-Hong Chao, Chao-Ching Chang, Chun-Ju Chang. Dysregulated miR-205 signaling promotes symmetric division of self-renewing mammary stem cells and mammary tumorigenesis. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4345. doi:10.1158/1538-7445.AM2014-4345
Abstract 4351: Targeting miRNA-driven aggressive behavior of pancreatic cancer stem cells by a novel drug
Authors: Bao Evan , Azmi Asfar , Zhang Ellen , Ali Shadan , Ahmad Aamir , Zaiem Feras , Bao Bin , Sarkar Fazlul H.
Published: May 2015

Pancreatic cancer (PC) is a lethal malignant disease, which is in part due to lack of early detection biomarkers. Once diagnosed, the majority of PC patients are already in advanced stages. Currently there are no effective treatments in improving the overall survival. Therefore, the majority of PC patients die within 5-6 months due to therapeutic resistance and early metastatic disease. Recently, a sub-population of cancer stem cells (CSCs) (<0.1%) have been identified from many different tumors including PC, and these cells are believed to be responsible for treatment resistance, tumor recurrence and metastasis. Many studies have shown that the existence of CSCs has important clinical implications because CSCs are involved in cell migration/invasion, drug resistance, and metastasis, which leads to poor clinical outcome. Therefore, selective killing or complete elimination of CSCs by novel approaches would likely become newer treatment strategy for PC in the improvement of overall survival. However, the regulation of CSC characteristics during tumor development and progression has not been well understood. In this study, cell and molecular biology techniques were used to investigate the CSC characteristics and its regulation to find a new effective approach for the treatment of PC. We found that cancer stem-like cells (CSLCs) which is reminiscent of CSCs isolated from human PC cell line, MiaPaCa-2 cells as triple-marker positive cells (CD44+/CD133+/EpCAM+ cells) cells displayed aggressive phenotypes, such as increased clonogenicity, cell migration, and self-renewal capacity, along with over-expression of CSC marker microRNAs (miRNAs) such as miR-21 and miR-210. Under-expression of miR-21 and miR-210 by transfection of anti-miRNAs led to decreased clonogenicity, cell migration and self-renewal capacity, along with reduced expression of CSC-associated markers/mediators such as CD44, EpCAM, EZH2, Notch-1, and Snail in CSLCs. Similarly, under-expression of CSC-associated mediator Notch-1 led to decreased cell migration, self-renewal capacity, and expression of EpCAM and Snail in CSLCs. Finally, CDF, a novel synthetic small molecule showed reduced aggressive behavior of CSLCs such as cell survival/growth, migration, clonogenicity, self-renewal capacity, and decreased expression of miR-21 and miR-210. From these results, we conclude that CDF may serve as an effective anti-tumor agent via targeting miRNA-driven aggressive behavior of CSLCs.

Citation Format: Evan Bao, Asfar Azmi, Ellen Zhang, Shadan Ali, Aamir Ahmad, Feras Zaiem, Bin Bao, Fazlul H. Sarkar. Targeting miRNA-driven aggressive behavior of pancreatic cancer stem cells by a novel drug. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4351. doi:10.1158/1538-7445.AM2014-4351
Abstract 4360: Hypoxia-induced miR-372 targets p62 to affect the progression of oral carcinoma
Authors: Yeh Li-Yin , Lin Shu-Chun , Liu Chung-Ji , Wong Yong-Kie , Chang Kuo-Wei
Published: May 2015

Oral squamous cell carcinoma (OSCC) is one of the prevalent neoplasms worldwide. Recent studies have shown that miRNAs were involved in the pathogenesis of various human malignancies, which function by post-transcriptional down-regulation of target genes. Also, hypoxia is a feature of most tumors and plays an important role in regulating tumor progression and resistance to therapy. In our previous study, we found that miR-372 was highly expressed in OSCC tissues. Other studies have shown the oncogenic roles of miR-372 in regulating the cell cycle and apoptosis in tumor cells by targeting different targets. This study identified that miR-372 was up-regulated by hypoxia in OSCC cells. The plasma level of miR-372 in OSCC patients significantly increased with the advance of OSCC. p62 is a multidomain protein that interacts with key components to drive diverse impacts on signaling regulation including detoxification pathway. Using in silico prediction modules, qRT-PCR analysis, Western blot analysis, reporter assay and exogenous expression construct, we discovered that p62 was a novel target of miR-372. Ectopic miR-372 expression and knockdown of p62 both enhanced the migration of OSCC cells, but it did not affect the proliferation of OSCC cells. Furthermore, we demonstrated that p62 was able to induce the expression of phase II enzyme NQO1, which resulted in the attenuation of reactive oxygen species and migration in OSCC cells. Furthermore, correlation among the expression of miR-372, p62 and NQO1 was found in OSCC tissues. This study concludes that hypoxia-induced miR-372 expression modulates the progression of OSCC by targeting p62, which disrupts the ROS homeostasis in tumor cells.

Citation Format: Li-Yin Yeh, Shu-Chun Lin, Chung-Ji Liu, Yong-Kie Wong, Kuo-Wei Chang. Hypoxia-induced miR-372 targets p62 to affect the progression of oral carcinoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4360. doi:10.1158/1538-7445.AM2014-4360
Abstract 4364: Posttranscriptional regulation of tetraspanins CD151 & CD9 in breast & prostate cancers
Authors: Bond Danielle R. , Passfield Crystal , Cairns Murray , Ashman Leonie K. , Weidenhofer Judith
Published: May 2015

Rationale

The tetraspanins CD151 & CD9 are known to play a prominent role in cancer progression by modulating cell adhesion and migration/invasion. CD151 is commonly overexpressed (metastasis enhancer) and tends to predict poor prognosis in many solid malignancies, whereas the metastasis suppressor CD9 is typically down-regulated which correlates with poor patient outcomes. However, progress in developing CD151 and CD9 as prognostic biomarkers & therapeutic targets has been hampered by a lack of understanding of how they are regulated in normal cells and how they are deregulated in cancers.

Objective

Many micro-RNAs are deregulated in prostate and breast cancers. Bio-informatic analysis has identified 250 miRNAs that potentially regulate CD151 or CD9. Therefore, we investigated whether miRNAs regulate CD151 and CD9 in a range of normal and cancerous breast and prostate cell lines in vitro.

Methods & Results

A dual luciferase reporter assay using the 3′UTR of CD151 and CD9 provided evidence that breast & prostate cancer cell lines endogenously express miRNA that regulate tetraspanins and that the extent of regulation varies across the cell lines. Analysis of miRNA expression in normal and cancer cell lines using miRNA arrays and qPCR has confirmed that some miRNAs which are predicted to target CD9, are up-regulated in many prostate and/or breast cancer cell lines compared to non-tumourigenic cells. Furthermore, miRNAs were identified with altered expression that were predicted to target CD151. These miRNA have also been shown to directly regulate CD151 or CD9 using the dual luciferase 3′UTR reporter assay with miRNA mimics.

Conclusion

Altered miRNA expression may contribute to prostate and breast cancers by manipulating tetraspanins to allow cancer progression and metastasis. Therefore, miRNA may be effective prognostic and/or therapeutic targets for prostate and breast cancers in the future.

Citation Format: Danielle R. Bond, Crystal Passfield, Murray Cairns, Leonie K. Ashman, Judith Weidenhofer. Posttranscriptional regulation of tetraspanins CD151 & CD9 in breast & prostate cancers. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4364. doi:10.1158/1538-7445.AM2014-4364
Abstract 4388: Targeted VHL activation by RNAa inhibits renal cell carcinoma
Authors: Kang Jong Soon , Kang Moo Rim , Yun Jieun , Oh Soo Jin , Park Ki Hwan , Lee Chang Woo
Published: May 2015

Von Hippel Lindau (VHL) is a tumor suppressor protein classically known to target the α subunit of hypoxia inducible factor (HIF) for proteasomal degradation. Emerging evidence has underscored a novel role for VHL in both cell cycle regulation and extracelluar matrix assembly. Recent studies have reported that chemically synthesized small duplex RNAs complementary to target promoter sequence, also known as small activating RNA (saRNA), can specifically induce gene expression in cancer cells, which might be associated with epigenetic changes and exhibited long-lasting gene activation for nearly 2 weeks. In the present study, we designed 5 candidate dsRNAs (dsVHL-58, dsVHL-93, dsVHL-395, dsVHL-515 and dsVHL-675) targeted for the VHL promoter at sites ranging from -675 to -58 relative to the transcription start site. Compared with controls, dsVHL-675 induced VHL expression by approximately 10-fold in renal cell carcinoma 769-P (VHL methylated) cells. Transfection of a dsVHL-675 into 769-P cells significantly inhibited cell proliferation in dose dependent manner. In addition, single transfection of dsVHL-675 induced VHL gene expression for 14 days. Functional analysis of dsVHL-675 transfected cells revealed that dsVHL-675 induced apoptosis by activating caspase 3 in dose dependent manner. Knockdown experiments confirmed that induction of apoptosis by dsVHL-675 required Ago2 and TNRC6A. This study suggests that targeted activation of VHL by dsVHL-675 may be explored as a novel therapy for the treatment of renal cell carcinoma.

Citation Format: Jong Soon Kang, Moo Rim Kang, Jieun Yun, Soo Jin Oh, Ki Hwan Park, Chang Woo Lee. Targeted VHL activation by RNAa inhibits renal cell carcinoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4388. doi:10.1158/1538-7445.AM2014-4388