bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2114: Docosahexaenoic acid induces degradation of human papillomavirus E6 and E7 oncoproteins through activation of the ROS-mediated ubiquitin-proteasome system in cervical cancer cells.
Authors: Jing Kaipeng , Shin Soyeon , Jeong Soyeon , Park Ji-Hoon , Seo Kang-Sik , Heo Jun-Young , Han Jeongsu , Park Jong-Il , Kweon Gi-Ryang , Park Seung-Kiel , Yoon Wan-Hee , Hwang Byung-Doo , Lim Kyu
Published: August 2014

Sustained expression of high risk human papillomavirus (HR-HPV) E6/E7 oncogenes is essential for the maintenance of cervical cancer. We have previously reported that docosahexaenoic acid (DHA), an omega-3 polyunsaturated fatty acid, triggers apoptosis in HR-HPV positive cervical cancer cells. The aim of present study was to investigate whether and how the DHA-induced apoptosis is associated with modulation of E6/E7 viral oncoproteins in HR-HPV positive HeLa and SiHa cells. We found that DHA diminished E6/E7 protein levels in a time-dependent manner and restored the function of known E6/E7 binding targets, p53 and retinoblastoma (Rb) tumor suppressors. DHA increased the ubiquitination and proteasomal degradation of E6/E7 oncoproteins as well as three different ubiquitin-proteasome system (UPS) reporter substrates (GFP-CL1, GFP-ODC and UbG76V-GFP), supporting that DHA degrades E6/E7 viral proteins via the UPS. Using HeLa cells constitutively expressing UbG76V-GFP reporter, we confirmed these effects of DHA and further observed that similar to DHA, hydrogen peroxide (H2O2) was also able to accelerate E6/E7 degradation through promoting the UPS activity. In addition, reactive oxygen species (ROS) scavengers significantly prevented the UPS-mediated reduction in E6/E7 viral proteins and apoptosis caused by DHA and H2O2, suggesting that the augmented UPS activity and E6/E7 degradation caused by DHA depend on its effect on ROS generation. Together, our results show that DHA induces degradation of HPV E6/E7 oncoproteins through activation of the ROS-mediated UPS, and provide novel insights into the mechanism by which DHA exerts its inhibitory effect on virus-associated cancer. [This work was supported by the National Research Foundation of Korea (NRF) grant funded by Korea government (2012-0005767, 2012-0005456 and 2011-0003060)].

Citation Format: Kaipeng Jing, Soyeon Shin, Soyeon Jeong, Ji-Hoon Park, Kang-Sik Seo, Jun-Young Heo, Jeongsu Han, Jong-Il Park, Gi-Ryang Kweon, Seung-Kiel Park, Wan-Hee Yoon, Byung-Doo Hwang, Kyu Lim. Docosahexaenoic acid induces degradation of human papillomavirus E6 and E7 oncoproteins through activation of the ROS-mediated ubiquitin-proteasome system in cervical cancer cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2114. doi:10.1158/1538-7445.AM2013-2114
Abstract 2119: Silencing of KCNJ2, a potassium influx channel, increases cisplatin-induced cell death in oral cancer.
Authors: Li Yen Liang
Published: August 2014

Oral cancer is the sixth leading cancer in the world. Cisplatin is the first-line of chemotherapeutic drug for the treatment of oral cancer, however, cancer with cisplatin resistance is a significant cause of treatment failure. To investigate the mechanism of cisplatin resistance, we established cisplatin-resistant (CR) subline cells from SAS, CGHNC8, and CGHNC9 oral cancer cell lines. The transcriptomic profiling analysis was performed between parental and CR subline cells to global survey and identify cisplatin-resistant genes. Total of 345 genes were differentially expressed in both SAS and CGHNC8 cells, in which 16 genes were confirmed. KCNJ2 was shown highest over-expression in both CR subline cells, and was selected for further investigation. KCNJ2 is a molecular potassium influx channel, which allows potassium flowing into cells to maintain ion homeostasis. To determine potential effect of KCNJ2, we have established KCNJ2 stable knocked down cells. Results showed that KCNJ2 knockdown cells were more sensitive to cisplatin treatment, as demonstrated by significantly reduced potassium influx, accelerated apoptosis by increasing caspase activity, and led to reduced cellular clonogenic survival. Taking together, we have profiled transcriptomes and identified at least 16 genes which may associate with cisplatin resistance in oral cancer cells. KCNJ2 silencing sensitizes cisplatin-resistant cells, through suppression of potassium influx. This study serves as knowledge foundation for clinical application in the prediction or prognosis of cisplatin treatment for oral cancer.

Citation Format: Yen Liang Li. Silencing of KCNJ2, a potassium influx channel, increases cisplatin-induced cell death in oral cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2119. doi:10.1158/1538-7445.AM2013-2119
Abstract 2134: Bone-targeted MMP inhibitors for the treatment of multiple myeloma.
Authors: Shay Gemma , Tauro Marilena , Laghezza Antonio , Tortorella Paolo , Hazlehurst Lori A. , Lynch Conor C.
Published: August 2014

Multiple Myeloma (MM) is a plasma cell malignancy that is hallmarked by areas of extensive bone destruction. Our group, and others have recently shown that individual host or tumor derived MMP-2 and MMP-9 play an important role in the progression of many cancers in the bone microenvironment, including MM. These data support the design of selective inhibitors of individual MMPs for the treatment of multiple myeloma. Previous clinical trials with broad-spectrum matrix metalloproteinase (MMP) inhibitors failed, arguably because the precise roles of individual MMPs had not been fully appreciated. Therefore, in the current study, we have developed novel bone seeking MMP inhibitors (BMMPIs) that are highly selective to MMP-2 and structurally are based on the bone targeting bisphosphonate, Tiludronate. In vitro, BMMPIs showed greater enzyme inhibition, and specificity for MMP-2 than previously synthesized bisphosphonates (e.g. Zoledronate, Etidronate and Tiludronate) with Ki values in the nanomolar range. Further treatment of the 5TGM1 mouse MM cell line with low micromolar concentrations of the BMMPI inhibitors significantly reduced cell viability through inhibition of cell growth (p<0.05). In vivo, 5TGMI tumor-bearing mice receiving BMMP inhibitors three times a week, showed significant increase in overall survival compared to vehicle and tiludronate treated mice over 80 days (p<0.05). This was associated with a reduction in tumor burden and protection from tumor-associated bone loss compared to vehicle treated mice as measured by x-ray and histomorphometry. These data suggest that given the roles for MMPs in tumor progression in bone, that our novel BMMPIs may be effective in the treatment of MM. We predict that MMP-2 specific BMMPIs may be more effective than current clinically used bisphosphonates for the treatment of MM, and may eliminate undesirable side effects of broad-spectrum MMP inhibitors due to their high specificity and bone seeking nature. We are currently continuing to characterize these BMMPI both as single agents, and in combination with existing MM treatments (eg. Bortezomib) for the treatment and eradication of MM.

Citation Format: Gemma Shay, Marilena Tauro, Antonio Laghezza, Paolo Tortorella, Lori A. Hazlehurst, Conor C. Lynch. Bone-targeted MMP inhibitors for the treatment of multiple myeloma. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2134. doi:10.1158/1538-7445.AM2013-2134
Abstract 2135: TGFβRI inhibition results in reduced collagen expression in pancreatic ductal adenocarcinoma.
Authors: Whatcott Clifford J. , Dumas Sabrina N. , Watanabe Aprill , LoBello Janine , Von Hoff Daniel D. , Han Haiyong
Published: August 2014

Pancreatic ductal adenocarcinoma (PDAC) is characterized by a robust stromal component termed the desmoplastic reaction. Transforming growth factor β (TGFβ) has been implicated in the activation of the cancer-associated fibroblast (CAF) that leads to pancreatic desmoplasia and the increased deposition of extracellular matrix proteins. Desmoplasia and its effects are thought to contribute to poor drug efficacy in PDAC treatment. We therefore hypothesized that TGFβ receptor (TGFβR) inhibition may serve to improve the activity of chemotherapeutics in PDAC by reducing the effects of desmoplasia. Using the TGFβRI inhibitors, LY2157299 and SB431542, we assessed the effects of TGFβR inhibition on cell proliferation and collagen production in both mono- and co-cultured pancreatic tumor cells and CAFs. We found that although the TGFβRI inhibitors had moderate effect on the proliferation of PDAC cells and co-cultured CAF cells, they significantly inhibited collagen 1 expression in the co-culture of MIA PaCa-2 and SU86.86 cells with CAFs (2.3 and 6.0-fold at the mRNA level, respectively). Similar qualitative reductions were also observed in immunofluorescence microscopy and immunoblots probing for collagen 1 protein levels. We also sought to determine if TGFβR inhibition would result in a reduction in collagen expression in the more biologically relevant in vivo model for PDAC, the LSL-KrasG12D/+;LSL-Trp53R172H/+;Pdx-1-Cre (KPC) mouse. KPC mice were treated orally, b.i.d., with 75mg/kg LY2157299 and by i.p., with gemcitabine (q3dx4) at 80mg/kg. Tumors in KPC mice treated with the combination of LY2157299 and gemcitabine showed a marked reduction in overall stromal content when compared to vehicle-treated mice. Our results suggest that TGFβR inhibition may be an effective means of ameliorating the effects of desmoplasia in PDAC, which may in turn improve the efficacy of current PDAC treatment regimens.

Citation Format: Clifford J. Whatcott, Sabrina N. Dumas, Aprill Watanabe, Janine LoBello, Daniel D. Von Hoff, Haiyong Han. TGFβRI inhibition results in reduced collagen expression in pancreatic ductal adenocarcinoma. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2135. doi:10.1158/1538-7445.AM2013-2135
Abstract 2138: Targeting cancer-associated fibroblasts using antibodies recognizing fibroblast activation protein-alpha.
Authors: Van Deventer James A. , Wittrup Karl D.
Published: August 2014

An expanding body of work suggests that the tumor stroma plays an integral role in the initiation and development of epithelial cancers. Within the stroma, cancer-associated fibroblasts (CAFs) are involved in several functions that form integral aspects of tumor progression, including immune suppression, metabolism, and invasion and metastasis. Despite the demonstrated importance of CAFs within the tumor microenvironment, targeting CAFs in a therapeutic context remains a challenge. The work presented here details the isolation and characterization of antibody fragments targeting multiple epitopes of fibroblast activation protein-alpha (FAP), a membrane-bound protease selectively overexpressed on CAFs in numerous epithelial cancers. The combination of a minimalist antibody fragment library and yeast surface display allowed for the isolation of more than 50 unique antibodies recognizing the human or murine forms of FAP, with close to 30 exhibiting cross-reactivity. To narrow the pool of cross-reactive antibodies to a biologically relevant subset, the fragments were converted to an antibody-like format that could be secreted from yeast. Without purification, the soluble proteins were first used to investigate the abilities of individual fragments to recognize FAP transiently expressed on the surface of HEK293 cells. Proteins for which binding to FAP could be confirmed were then used in combination with antibody fragments displayed on the surface of yeast in order to determine which sets of fragments bind distinct epitopes of FAP. Flow cytometry-based assays revealed that the isolated fragments recognize two or more distinct epitopes of the antigen. A handful of fragments is currently being subjected to affinity maturation and conversion to IgGs, and work to further investigate the in vitro and in vivo characteristics of the proteins is ongoing. These proteins may allow for the ablation of cancer-associated fibroblasts or interference with the proteolytic activity of FAP in the tumor microenvironment, providing biological insights into the roles of CAFs and therapeutic approaches to cancer treatment. Furthermore, the general antibody isolation and characterization approaches used in this work are accessible to many single-investigator laboratories and should be applicable to the targeting of numerous cancer antigens.

Citation Format: James A. Van Deventer, Karl D. Wittrup. Targeting cancer-associated fibroblasts using antibodies recognizing fibroblast activation protein-alpha. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2138. doi:10.1158/1538-7445.AM2013-2138
Abstract 215: Neural precursor cells induce cell death of high-grade astrocytomas through stimulation of TRPV1.
Authors: Synowitz Michael , Stock Kristin , Kumar Jitender , Petrosino Stefania , Imperatore Roberta , St J Smith Ewan , Wend Peter , Purfürst Bettina , Nuber Ulrike A , Gurok Ulf , Matyash Vitali , Wälzlein Joo-Hee , Chirasani Sridhar R , Dittmar Gunnar , Cravatt Benjamin F , Momma Stefan , Lewin Gary R , Ligresti Alessia , De Petrocellis Luciano , Cristino Luigia , Tonn Joerg Christian , Di Marzo Vincenzo , Kettenmann Helmut , Glass Rainer
Published: August 2014

Primary astrocytomas of grade 3 or 4 according to the classification system of the World Health Organization (high-grade astrocytomas or HGAs) are preponderant among adults and are almost invariably fatal despite the use of multimodal therapy. Here we show that the juvenile brain has an endogenous defense mechanism against HGAs. Neural precursor cells (NPCs) migrate to HGAs, reduce glioma expansion and prolong survival time by releasing endovanilloids that activate the vanilloid receptor (transient receptor potential vanilloid subfamily member-1 or TRPV1) on HGA cells. TRPV1 is highly expressed in tumor and weakly expressed in tumor-free brain. TRPV1 stimulation triggers tumor cell death through the branch of the endoplasmic reticulum stress pathway that is controlled by activating transcription factor-3 (ATF3). The antitumorigenic response of NPCs is lost with aging. NPC-mediated tumor suppression can be mimicked in the adult brain by systemic administration of the synthetic vanilloid arvanil, suggesting that TRPV1 agonists have potential as new HGA therapeutics.

Citation Format: Michael Synowitz, Kristin Stock, Jitender Kumar, Stefania Petrosino, Roberta Imperatore, Ewan St J Smith, Peter Wend, Bettina Purfürst, Ulrike A Nuber, Ulf Gurok, Vitali Matyash, Joo-Hee Wälzlein, Sridhar R Chirasani, Gunnar Dittmar, Benjamin F Cravatt, Stefan Momma, Gary R Lewin, Alessia Ligresti, Luciano De Petrocellis, Luigia Cristino, Joerg Christian Tonn, Vincenzo Di Marzo, Helmut Kettenmann, Rainer Glass. Neural precursor cells induce cell death of high-grade astrocytomas through stimulation of TRPV1. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 215. doi:10.1158/1538-7445.AM2013-215
Abstract 2210: Integration of preclinical and clinical studies to identify pharmacogenetic markers associated with paclitaxel-induced neuropathy.
Authors: Wheeler Heather E. , Rodriguez-Antona Cristina , Wing Claudia , Komatsu Masaaki , Dolan M. Eileen
Published: August 2014

Paclitaxel is a tubulin-targeting agent used in the treatment of many cancers, including breast, lung, head and neck, and ovarian. Peripheral neuropathy is a common side effect of many chemotherapeutic agents, including paclitaxel, and limits their efficacy in patients. The overall goal of our work is to identify patients at risk for peripheral neuropathy using whole genome preclinical and clinical studies. Epstein-Barr virus transformed lymphoblastoid cell lines (LCLs) have emerged as a promising preclinical model system in the study of the genetics of drug toxicity because the cells provide a cost-effective testing system in which environmental factors such as concomitant medications can be controlled. Most importantly, publicly available genotype and sequencing data is available, to allow for genome-wide association studies between HapMap/1000 Genomes variants and pharmacologic phenotypes measured in the LCLs after drug treatment such as paclitaxel induced growth inhibition and apoptosis. The overall genetic architecture of paclitaxel-induced neuropathy is well captured by the cell-based models as evidenced by significant enrichment of paclitaxel-induced toxicity SNPs from the preclinical model in clinical genome-wide association studies of peripheral neuropathy. We utilized iCell Neurons, which are commercially available cells derived from induced pluripotent stem cells, to study the effects of gene modulation on paclitaxel-induced cell growth inhibition using Cell Titer Glo and on neurite parameters of outgrowth, processes and branching using the ImageXpress Micro system. Through transient knockdown in iCell Neurons of RFX2, a gene identified through clinical and preclinical studies to be associated with paclitaxel neuropathy, sensitivity to paclitaxel increased as measured by a decrease of neurite outgrowth, processes and branching. The ability to identify cancer patients at risk of paclitaxel-induced peripheral neuropathy could influence treatment choice, outcome and quality of life for breast, lung, head and neck, and ovarian cancer patients. This work was supported in part by NIH/NIGMS UO1GM61393.

Citation Format: Heather E. Wheeler, Cristina Rodriguez-Antona, Claudia Wing, Masaaki Komatsu, M. Eileen Dolan. Integration of preclinical and clinical studies to identify pharmacogenetic markers associated with paclitaxel-induced neuropathy. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2210. doi:10.1158/1538-7445.AM2013-2210
Abstract 2213: Influence of genetic polymorphism in UGT1A1, UGT1A7 and UGT1A9 on irinotecan, SN-38 and SN-38G pharmacokinetics.
Authors: Valenzuela Belen , González-Sales Mario , Escudero Vanessa , Navarro Elena , Perez-Ruixo Carlos , Rebollo Joseba , González-Manzano Ramon , Pérez-Ruixo Juan José
Published: August 2014

Background and objective: SN-38 is the active metabolite of irinotecan (CPT-11) and mainly responsible of hematological and intestinal toxicity after CPT-11 treatments. SN-38 is inactivated into SN-38G by uridine diphosphate glucuronosyltransferase (UGT) enzymes. Some polymorphism of these enzymes leads lower clearance (CL) of SN-38 and therefore bigger exposition and bigger risk of hematological and/or intestinal toxicity (1). Therefore, the goal of this study has been to evaluate the influence of genetic polymorphism in UGT1A1, UGT1A7 and UGT1A9 on the population pharmacokinetics of CPT-11 and its metabolites, SN-38 and SN-38G on Caucassian cancer population.

Methods: Plasma concentrations of CPT-11, SN-38 and SN-38G from 72 patients were pooled to develop a joint population pharmacokinetic model using NONMEM 7. The effect of age, sex, body surface area, total bilirubin, co-medication, tumor type, and UGT1A1, UGT1A7 and UGT1A9 genotypes on the model parameters was evaluated.

Results: The typical values (between-subject variability; %) of the irinotecan, SN-38 and SN-38G clearances were 42,9 L/h (56,4%), 1340 L/h (76,8%) and 188 L/h (70,1%), respectively. Clearance in patients with some allele (heterozygous or homozygous) associated with low enzymatic activity of UGT1A1*28 was a 35,7% lower (CI 95%: 6,7 - 64,7; p=0,003) than in wild-type patients. Additionally, linkage dissequilibrium indicates a strong interaction between functional UGT1A1, UGT1A7 and UGT1A9 polymorphisms related to the alteration of the UGT enzyme activity.

Conclusion: The impact of UGT1A1*28 genotype on the systemic exposure of SN-38 justifies its routine determination in patients receiving CPT-11 and subsequent dose adjustment.

Citation Format: Belen Valenzuela, Mario González-Sales, Vanessa Escudero, Elena Navarro, Carlos Perez-Ruixo, Joseba Rebollo, Ramon González-Manzano, Juan José Pérez-Ruixo. Influence of genetic polymorphism in UGT1A1, UGT1A7 and UGT1A9 on irinotecan, SN-38 and SN-38G pharmacokinetics. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2213. doi:10.1158/1538-7445.AM2013-2213
Abstract 2216: The role of cytosine modifications in cellular sensitivity to cytarabine.
Authors: Moen Erika L. , Mu Wenbo , Godley Lucy A. , Dolan M. Eileen , Zhang Wei
Published: August 2014

Introduction: Inter-individual differences in cytosine modifications are a result of genetic and environmental influences. 5-methylcytosine and 5-hydroxymethylcytosine are covalent cytosine modifications that occur in CpG dinucleotides throughout the human genome. Objective: To characterize inter-individual variation in cytosine modifications in HapMap samples and investigate the contribution this may have on downstream phenotypes, such as cellular sensitivity to chemotherapeutics, such as cytarabine. Model: Cytosine modification levels at >485,000 CpGs were measured in the International HapMap lymphoblastoid cell lines (LCLs). We utilized the Illumina Infinium 450K HumanMethylation Beadchip array to measure cytosine modification levels in 73 unrelated African (YRI - Yoruba people from Ibadan, Nigeria) and 60 European ancestry (CEU - Caucasians from Utah, US) LCLs. In addition, we evaluated LCL sensitivity to cytarabine in these HapMap cell lines. Results: We performed a linear regression model in order to identify novel associations between cytosine modifications and cytarabine-induced cytotoxicity. From these analyses, we found that promoter methylation of CDC6 and CCDC37 was associated with cytarabine IC50 (p= 3.75 x 10−6 and p= 5.39x 10−6, respectively). Furthermore, it has been shown previously that SNPs associated with pharmacological traits in LCLs are enriched in expression quantitative trait loci (eQTLs). We interrogated SNPs associated with cytosine modification (mSNPs) to help us understand the role of mSNPs in pharmacologic trait-associated genetic variants. We found that SNPs associated with cytarabine are enriched in cis-acting mSNPs in LCLs derived from individuals of European descent. Next, we integrated genome-wide cytosine modification data with genotype and gene expression data available for HapMap LCLs to allow for a more comprehensive understanding of inter-individual genetic variation. We identified a SNP in SLFN5 that was significantly associated with cytarabine sensitivity that is a cis-acting eQTL and mSNP for SLFN5 gene expression and cytosine modification. Conclusions: Analyzing cytosine modification-drug phenotype data may identify novel candidate genes associated with sensitivity to cytarabine. In addition, integrating genotype and gene expression data allow for a better understanding of the biological function underlying pharmacologic trait-associated SNPs.

Citation Format: Erika L. Moen, Wenbo Mu, Lucy A. Godley, M. Eileen Dolan, Wei Zhang. The role of cytosine modifications in cellular sensitivity to cytarabine. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2216. doi:10.1158/1538-7445.AM2013-2216
Abstract 2219: The European Paediatric Oncology off-patent medicines Consortium (EPOC) pharmacokinetic/pharmacodynamic study of doxorubicin.
Authors: Boddy Alan V. , Steel Alison J. , Boos Joachim , Krischke Miriam , Andre Nicolas , Bisogno Gianni , Hempel Georg , Bode Gerlind , D'Incalci Maurizio , Kontny Nina
Published: August 2014

Background: Doxorubicin is a key component of a number of treatment regimens used in paediatric oncology despite the very limited pharmacology data on which current dosing regimens are based. In recognition of this, doxorubicin was included on the EMA priority list for paediatric off-patent medicines for which more information was required, particularly on pharmacokinetics.

Methods: The EPOC consortium, with collaborators in the UK, France, Germany and Italy, performs paediatric oncology pharmacology clinical trials. This study is a multicentre, multinational phase II pharmacokinetic study investigating age-dependency in the clearance of doxorubicin in children with solid tumours and leukaemia. All patients are being treated according to a tumour-specific national or European therapeutic trial. The aim was to recruit 100 patients, with a particular focus on children less than 3 years. The plasma levels of markers such as troponin T, are being measured to evaluate their use as clinical markers for cardiotoxicity. Data is being analysed using a population pharmacokinetic approach and the impact of pharmacogenetics is being investigated.

Results: The study has been open to recruitment in 20 clinical centres across the four participating countries. The target 100 patients have been recruited from all the contributing study protocols, with 26 patients less than 3 years. Of these 26 patients, 5 patients are less than 1 year. An interim pharmacokinetic analysis has been performed on the data from the first 31 patients. The results of the analysis are consistent with the primary aims of the overall study.

Conclusion: The data generated during this study should provide comparative data on the age-dependent pharmacokinetics and pharmacogenetics of doxorubicin. The results will contribute to optimising the safe use of doxorubicin in children with cancer.

Acknowledgements: Study funded under the seventh framework programme of the European Commission (FP7)

Citation Format: Alan V. Boddy, Alison J. Steel, Joachim Boos, Miriam Krischke, Nicolas Andre, Gianni Bisogno, Georg Hempel, Gerlind Bode, Maurizio D'Incalci, Nina Kontny. The European Paediatric Oncology off-patent medicines Consortium (EPOC) pharmacokinetic/pharmacodynamic study of doxorubicin. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2219. doi:10.1158/1538-7445.AM2013-2219
Abstract 2223: Identification of novel protein modulators of p53 family function using photochemical crosslinking of stapled peptides.
Authors: Felsenstein Kenneth M. , Headley Kathryn M. , Murray Kevin A. , Russell Elisabeth A. , Bernal Federico
Published: August 2014

Considered to be the “guardian of the genome,” the tumor suppressor p53 works to counteract genetic mutation through cell cycle arrest, DNA repair or apoptosis to prevent proliferation of damaged cells. Dysfunction of the p53 pathway is a large driver of tumorigenesis, with an estimated more than 50% of cancers bearing some p53 abnormality; hence, the normalization of p53 function is considered an important clinical goal for cancer therapy. Many of the interactions involving p53 have been extensively studied; however, absent structural information, the identification of the precise nature of the binding interactions proves challenging, more so if they are transient. Using the hydrocarbon stapled peptide methodology previously used by our group to study interactions mediated by alpha-helices, we have begun identifying novel binding targets of the p53 transactivation domain (TAD) as well as its related proteins p63 and p73 through biochemically specific, covalent photochemical crosslinking reactions. Methodologically, we have developed the use of photoreactive stapled alpha helices (pSAHs) representing a unique way to use highly selective, biochemically specific cross-linking reagents. These can be used more broadly to study protein-protein binding interactions in cell-free and cell based assays. The technique has allowed us to learn more about the biologically active state of proteins that interact with the p53-family member TADs as well as the identification of unexplored binding partners and their mechanism of action in disease.

Citation Format: Kenneth M. Felsenstein, Kathryn M. Headley, Kevin A. Murray, Elisabeth A. Russell, Federico Bernal. Identification of novel protein modulators of p53 family function using photochemical crosslinking of stapled peptides. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2223. doi:10.1158/1538-7445.AM2013-2223
Abstract 2226: Structure/function characterization of SBi4225, a novel inhibitor of the calcium-binding protein S100B.
Authors: Cavalier Michael C. , McKnight Laura , Weber David , Wilder Paul
Published: August 2014

The S100 family of proteins displays no enzymatic activity but has been shown to play roles in an array of biological functions such as calcium homeostasis, cellular growth, inflammation, and metabolism. The influence of these homodimeric proteins is mediated through protein-protein interactions in a typically calcium dependent manner and aberrant expression of S100 proteins has been linked to a variety of cancers, cardiomyopathies, and neurological disorders. Of particular interest, S100B is highly over-expressed in many cancers, including malignant melanoma (MM). In MM, S100B is used as prognostic indicator in evaluating treatment success and in predicting relapse. Studies now show that S100B is more than a clinical marker, and that it binds wild-type p53 in a calcium-dependent manner, sequestering it, and promoting its degradation. This results in the loss of p53-dependent tumor suppression activities. As inhibition of S100B expression by RNA interference restores both wild-type p53 protein levels and normal transcriptional activation/apoptosis activities, we have made it our long-term goal is to specifically inhibit the S100B-p53 interaction in order to restore the p53 tumor suppression function as a potential therapy for malignant melanoma. Although we have succeeded in identifying many S100B inhibitors (also known as SBiXs), optimization studies are a necessary part of the development of drugs with useful therapeutic activities. Included amongst these optimization studies are characterizations of structure-activity relationships (SAR). Our initial SAR studies have resulted in the development of a second series of SBiXs. Amongst this series is the novel S100B inhibitor, SBi4225. As part of the cyclical process of SAR studies, the binding of SBi4225 to S100B has been investigated using X-ray crystallography. As described within, SBi4225 binds to S100B in a position and orientation analogous to its predecessors, SBi4211 and SBi1. NMR data including chemical shift perturbations of 15N isotopically labeled protein backbone were consistent with the X-ray crystal structures and demonstrated the relevance of the SBi4225-S100B complex in solution. This new understanding of the SBi4225 structure/function relationship will undoubtedly impact the development and optimization of future SBiXs.

Citation Format: Michael C. Cavalier, Laura McKnight, David Weber, Paul Wilder. Structure/function characterization of SBi4225, a novel inhibitor of the calcium-binding protein S100B. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2226. doi:10.1158/1538-7445.AM2013-2226
Abstract 2235: Epigallocatechin-gallate suppresses tumorigenesis by directly targeting Pin1.
Authors: Urusova Darya V. , Shim Jung-Hyun , Kim Dong Joon , Jung Sung Keun , Zykova Tatyana , Carper Andria , Bode Ann M. , Dong Zigang
Published: August 2014

The most active anticancer component in green tea is epigallocatechin-3-gallate (EGCG). The human peptidyl prolyl cis/trans isomerase (Pin1) plays a critical role in oncogenic signaling. Herein, we report the X-ray crystal structure of the Pin1/EGCG complex resolved at 1.9 Å resolution. Notably, the structure revealed the presence of EGCG in both the WW and PP1ase domains of Pin1. The direct binding of EGCG with Pin1 was confirmed and the interaction inhibited Pin1 PP1ase activity. In addition, proliferation of cells expressing Pin1 was inhibited and tumor growth in a xenograft mouse model was suppressed. The binding of EGCG with Arg17 in the WW domain prevented the binding of c-Jun, a well-known Pin1 substrate. EGCG treatment corresponded with a decreased abundance of cyclin D1 and diminution of TPA-induced AP-1 or NFκB promoter activity in cells expressing Pin1. Overall, these results showed that EGCG directly suppresses the tumor promoting effect of Pin1.

Citation Format: Darya V. Urusova, Jung-Hyun Shim, Dong Joon Kim, Sung Keun Jung, Tatyana Zykova, Andria Carper, Ann M. Bode, Zigang Dong. Epigallocatechin-gallate suppresses tumorigenesis by directly targeting Pin1. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2235. doi:10.1158/1538-7445.AM2013-2235
Abstract 2272: Comparison of metformin, pheformin and buformin in mammary carcinogenesis in non-obese, non-diabetic rats.
Authors: Zhu Zongjian , Jiang Weiqin , McGinley John N. , Thompson Henry J.
Published: August 2014

Emerging evidence from studies of human pharmacoepidemiology indicate that metformin, an antidiabetic drug, is associated with reduced breast cancer risk in diabetic individuals. Data from preclinical mammary carcinogenesis studies in the rat indicate that metformin also has protective activity against chemically-induced mammary cancer in non diabetic rats, but the dose required for inhibition is above that used clinically for the treatment of diabetes. Human breast cancer cell growth also is inhibited by metformin, but at concentrations far in excess of anti-diabetic plasma concentrations of the drug. It is likely that pharmacokinetic/pharmacodyanmic factors contribute to this blunted sensitivity, possibly due to carcinogenesis-associated down regulation of organic cationic transporters (OCT) required for cellular uptake of metformin. Since some biguanides such as phenformin and buformin are not dependent on OCT for cellular accumulation, their effects against mammary carcinogenesis were evaluated in comparison to metformin as a proof-in-concept study that could set the stage for development of second generation biguanides. The hypothesis evaluated was that inhibition of mammary carcinogenesis by phenformin and buformin would be superior to metformin. Female Sprague Dawley rats (n=30/group) were injected with 1-methyl-1-nitrosourea (50 mg/kg) at 21 days of age and 7 days thereafter randomized to metformin, phenformin or buformin dietary groups and fed respective biguanides (10 mmol/kg) diets for 6 weeks. Both phenformin and buformin significantly reduced the number of mammary adenocarcinoma per rat (3.6, 1.6, and 0.7 cancers/rat for metformin, phenformin and buformin respectively) and tumor burden (1.61, 0.63, and 0.12 g/rat for metformin, phenformin and buformin respectively) and elongated cancer latency compared to metformin (40, 45, and 48 days post carcinogen for metformin, phenformin and buformin respectively) (p<0.05). Mammary cancer incidence was also lower in animals fed phenformin (77%) or buformin (43%) diet vs metformin (93%), but the difference was only significant between metformin and buformin (p<0.001). Cell culture experiments confirmed the carcinogenesis data. The comparative effects of these biguanides on the LKB1/AMPK axis will be reported. The associated mechanisms of cell growth inhibiton are under investigation. Supported by PHS grant CA52626 from the National Cancer Institute.

Citation Format: Zongjian Zhu, Weiqin Jiang, John N. McGinley, Henry J. Thompson. Comparison of metformin, pheformin and buformin in mammary carcinogenesis in non-obese, non-diabetic rats. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2272. doi:10.1158/1538-7445.AM2013-2272
Abstract 2274: Metformin use and risk of lung cancer in patients with diabetes.
Authors: Sakoda Lori C. , Achacoso Ninah S. , Quesenberry Charles P. , Peng Tiffany , Ehrlich Samantha F. , Ferrara Assiamira , Habel Laurel A.
Published: August 2014

Observational studies suggest that cancer incidence is lower in patients with type 2 diabetes treated with metformin, relative to patients treated with other diabetes medications. The few studies to examine metformin use in relation to lung cancer risk, however, have produced inconsistent results. To clarify this relationship, we conducted a retrospective cohort study of 50,722 adults aged 40 years or older included in the Kaiser Permanente Northern California Diabetes Registry, who completed a mailed survey administered in 1994 to 1996 on health-related traits and behaviors. All prescribed diabetes medications were identified for each patient from pharmacy records. Patients were followed for lung cancer from 1997 to 2009, with a median follow-up of 8.1 years. Using Cox regression, with age as the time scale, rate ratios (RR) and 95% confidence intervals (CI) were calculated to estimate risk of lung cancer associated with new use of metformin, adjusted for sex, race/ethnicity, smoking status, income, education level, diabetes duration, baseline measures of hemoglobin A1c and creatinine, and ever use of other types of diabetes medications. Measures of metformin use, along with ever use for each type of diabetes medication, were modeled as time-dependent variables. During 389,887 person-years of follow-up, 685 patients were diagnosed with lung cancer. The adjusted RR for ever use of metformin was 0.95 (95% CI, 0.79-1.14). For duration of metformin use, the adjusted RRs for <2.0, 2.0-4.9, and 5.0+ years were 1.01 (95% CI, 0.80-1.28), 0.94 (95% CI, 0.73-1.22), and 0.86 (95% CI, 0.63-1.18), respectively. In analyses stratified by smoking status, the adjusted RRs for ever use of metformin were 0.98 (95% CI, 0.79-1.21) in ever smokers and 0.59 (95% CI, 0.34-1.01) in never smokers. However, in never smokers, the number of users diagnosed with lung cancer was relatively small, and no dose-response decrease in risk with increasing duration of metformin use was found; the corresponding adjusted RRs for <2.0, 2.0-4.9, and 5.0+ years of metformin use were 0.52 (95% CI, 0.23-1.16), 0.58 (95% CI, 0.26-1.26), and 0.70 (95% CI, 0.31-1.58), respectively. Our results suggest that metformin use is not associated with lung cancer risk in former or current smokers with diabetes. Additional research is needed to determine whether metformin use might alter lung cancer risk in never smokers with diabetes.

Citation Format: Lori C. Sakoda, Ninah S. Achacoso, Charles P. Quesenberry, Tiffany Peng, Samantha F. Ehrlich, Assiamira Ferrara, Laurel A. Habel. Metformin use and risk of lung cancer in patients with diabetes. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2274. doi:10.1158/1538-7445.AM2013-2274
Abstract 2294: ROCK1 inhibition promotes the self-renewal of a novel mouse mammary cancer stem cell.
Authors: Maurer Jochen , Castro David J. , Oshima Robert G.
Published: August 2014

The differentiation of stem-like tumor cells may contribute to the cellular heterogeneity of breast cancers. We report the propagation of highly enriched mouse mammary cancer stem cells that retain the potential to differentiate both in vivo and in culture and their use to identify chemical compounds that influence both self-renewal and differentiation. We identify epithelial tumor initiating cells (ETICs) that express lineage markers of both basal and luminal mammary cell lineages and retain the potential, from even single cells, to generate heterogeneous tumors similar to the tumor of origin. ETICs can progress through a Rho associated coiled-coil containing protein kinase 1 (ROCK1) dependent, epithelial to mesenchymal transition to generate mesenchymal tumor initiating cells (MTICs) capable of initiating tumors of limited heterogeneity. The propagation of ETICs may allow for the identification of new therapeutic compounds that may inhibit or prevent progression of some types of breast cancer.

Citation Format: Jochen Maurer, David J. Castro, Robert G. Oshima. ROCK1 inhibition promotes the self-renewal of a novel mouse mammary cancer stem cell. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2294. doi:10.1158/1538-7445.AM2013-2294
Abstract 2316: Mutations in the p53 gene family reveal conservation of structure-function in the L1 and L3 loops and a response element code for transcriptional specificity.
Authors: Ciribilli Yari , Monti Paola , Bisio Alessandra , Nguyen H. T. , Ethayathulla A S. , Resnick Micheal A. , Viadiu Hector , Fronza Gilberto , Inga Alberto
Published: August 2014

The p53, p63 and p73 family of sequence-specific transcription factors share similar biochemical properties and structural features resulting in the recognition of DNA with identical amino acids and similar binding affinity. However, loss of individual members results in very different phenotypes in mice as well as humans. We have addressed functional relationships between p53 family proteins using a combination of mutants, target response elements (REs) and sequence-specific transactivation assays based in yeast and human cells. p53 alleles exhibiting enhanced transcriptional activation and altered promoter selectivity were previously identified in the L1 and L3 loop of the DNA binding domain of the protein. Included was S121F that results in defective p21 induction and confers a strong apoptotic phenotype in mammalian cells. Guided by result with p53-S121F, -T123A and -S240N mutants, the corresponding p63 and p73 alleles were investigated for ability to drive transactivation at over fifty p53 REs in isogenic yeast reporter strains. The phenotypes observed with p53 alleles were reproduced with the p63- and p73- alleles. Focusing on p73-S139F (corresponding to p53-S121F) transactivation at many RE sequences, we identify and experimentally confirm an RE target sequence code that predicts enhanced or reduced transactivation potentials. The correlation is particularly strong for induction at p53 targets within promoters of genes associated with apoptosis. Furthermore, p73 S139F showed an enhanced DNA-binding cooperativity in vitro, and loop L1 shows conformational changes in the crystal structure of the protein-DNA complex. The altered function phenotype was confirmed not only in gene reporter assays but also for endogenous gene expression in the p53 null HCT116 human cell line, suggesting that target recognition can predict relative changes in transcription rates at different loci. Based on the correlation between RE code and associated gene functions, we propose the selection during evolution of an RE sequence code within target promoters that can affect intrinsic conformational flexibility / DNA binding affinity of p53 proteins thereby modulating in vivo selectivity, specifically favoring the activation of apoptotic target genes

Citation Format: Yari Ciribilli, Paola Monti, Alessandra Bisio, H. T. Nguyen, A S. Ethayathulla, Micheal A. Resnick, Hector Viadiu, Gilberto Fronza, Alberto Inga. Mutations in the p53 gene family reveal conservation of structure-function in the L1 and L3 loops and a response element code for transcriptional specificity. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2316. doi:10.1158/1538-7445.AM2013-2316
Abstract 2319: Histone demethylase JMJD2C is a coactivator for hypoxia-inducible factor 1 that is required for breast cancer progression.
Authors: Luo Weibo , Chang Ryan , Zhong Jun , Pandey Akhilesh , Semenza Gregg L.
Published: August 2014

Hypoxia-inducible factor 1 (HIF-1) activates transcription of genes encoding proteins that play key roles in breast cancer biology. We hypothesized that interaction of HIF-1 with epigenetic regulators may increase HIF-1 transcriptional activity and thereby promote breast cancer progression. We report that the histone demethylase jumonji domain containing protein 2C (JMJD2C) selectively interacts with HIF-1α, but not HIF-2α, and that HIF-1α mediates recruitment of JMJD2C to the hypoxia response elements (HREs) of HIF-1 target genes. JMJD2C decreases trimethylation of histone H3 at lysine 9, and enhances HIF-1 binding to HREs, thereby activating transcription of BNIP3, LDHA,PDK1, and SLC2A1, which encode proteins that are required for metabolic reprogramming, as well as LOXL2 and L1CAM, which encode proteins that are required for lung metastasis. JMJD2C expression is significantly associated with expression of GLUT1, LDHA, PDK1, LOX, LOXL2, and L1CAM mRNA in human breast cancer biopsies. JMJD2C knockdown inhibits breast tumor growth and spontaneous metastasis to the lungs of mice following mammary fat pad injection. Taken together, these findings establish a novel epigenetic mechanism that stimulates HIF-1-mediated transactivation of genes encoding proteins involved in metabolic reprogramming and lung metastasis in breast cancer.

Citation Format: Weibo Luo, Ryan Chang, Jun Zhong, Akhilesh Pandey, Gregg L. Semenza. Histone demethylase JMJD2C is a coactivator for hypoxia-inducible factor 1 that is required for breast cancer progression. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2319. doi:10.1158/1538-7445.AM2013-2319