bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3273: rkmh: A MinHash toolbox for analyzing HPV coinfections
Authors: Dawson Eric T. , Wagner Sarah , Roberson David , Yeager Meredith , Boland Joseph , Garrison Erik , Schiffman Mark , Raine-Bennet Tina , Lorey Thomas , Castle Phillip , Chanock Stephen , Mirabello Lisa , Durbin Richard
Published: August 2018

Human papillomaviruses are common infections transmitted by direct physical contact. Two sexually transmitted types, HPV16 and HPV18, are responsible for almost 75% of cervical cancer cases worldwide; another 10 types of >200 known types are considered carcinogenic. Among the four HPV16 lineages (A, B, C, and D), 100-fold differences in odds ratios for adenocarcinoma are observed. It is therefore informative to distinguish between infection with HPV16 lineages, even though they differ by as little as 2% of their genome. We present a computational toolkit, rkmh, for characterizing viral coinfections. rkmh uses kmer matching strategies to rapidly determine the most similar type or lineage reference genome for a given read. The proportion of reads matching each reference genome can then be calculated and the ratios of the infecting viruses can be estimated. To assess the performance of rkmh we first simulated 100bp paired end Illumina read sets from the PAVE database of HPV reference genomes, then also evaluated performance on a real HPV16 sample sequenced on the Ion Torrent Proton platform (typical read length 250bp), and a set of 3,660 Oxford Nanopore minION reads generated from two HPV16 reference strains (typical read length over 6500bp). We demonstrate that rkmh can adequately classify HPV infections at the type and lineage level. We discuss further applications of the tool in metagenomics. rkmh is freely available at https://github.com/edawson/rkmh.

Citation Format: Eric T. Dawson, Sarah Wagner, David Roberson, Meredith Yeager, Joseph Boland, Erik Garrison, Mark Schiffman, Tina Raine-Bennet, Thomas Lorey, Phillip Castle, Stephen Chanock, Lisa Mirabello, Richard Durbin. rkmh: A MinHash toolbox for analyzing HPV coinfections [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3273.
Abstract 329: Highly potent and selective ATM kinase inhibitor M3541: A clinical candidate drug with strong antitumor activity in combination with radiotherapy
Authors: Fuchss Thomas , Mederski Werner W. , Zenke Frank T. , Dahmen Heike , Zimmermann Astrid , Blaukat Andree
Published: August 2018

M3541 is an ATP-competitive inhibitor of the Ataxia telangiectasia mutated (ATM) kinase (IC50 < 1 nM), which targets tumor cell survival and growth by inhibiting double-strand break (DSB) repair as well as checkpoint control. DSB repair is crucial for survival of malignant tumor cells, especially under treatment with DNA damaging chemo- and radiotherapy. As such, the rationale of pharmacological inhibition of ATM is to increase and maintain the extent of unrepaired DNA damage generated by radio-, chemotherapy, and targeted therapies to drive tumor cells into cell death. We have developed an orally administered, sub-nanomolar potent & selective kinase inhibitor of ATM, M3541, for cancer therapy in combination with DNA damaging modalities. Here, we present the optimization of initial hit matter & its structure-activity relationships leading to our clinical candidate M3541, its chemical structure (first-time disclosure) & preclinical characterization using biochemical, cellular & human tumor xenograft models. M3541 sensitizes tumor cell lines to radiation therapy in vitro and strongly enhances the anti-tumor activity of ionizing radiation in vivo. These effects are due to the inhibition of ATM kinase activity as demonstrated by the levels of the phosphorylation of its primary downstream target CHK2 in human tumor cell lines. M3541 is currently being investigated in a multicenter Ph I clinical trial.

Citation Format: Thomas Fuchss, Werner W. Mederski, Frank T. Zenke, Heike Dahmen, Astrid Zimmermann, Andree Blaukat. Highly potent and selective ATM kinase inhibitor M3541: A clinical candidate drug with strong antitumor activity in combination with radiotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 329.
Abstract 3304: The mutation landscape of cancers serves as a record of early malignant transformation
Authors: Polak Paz , Karlic Rosa , Kubler Kirsten , Foulkes William D. , Getz Gad
Published: August 2018

How the cell lineage influences a tissue's susceptibility to malignant transformation is a fundamental question in cancer biology, which has been barely addressed in cancer genomics thus far. Cell properties are encoded in the cell type-specific chromatin structure and we previously demonstrated that the cell-of-origin (COO) chromatin organization is a key determinant of the landscape of somatic mutations, which accumulated over lifetime serving as a memory of the historical cell lineage (Polak et al, Nature , 2015). We now show that this principle is generalizable to common tumor types and offers insights into the molecular events of cancer initiation. We extended our work to 2,641 genomes from 30 cancer types and epigenetic modifications from 98 normal tissues. In 25 cancer types, the tumor originated from a cell type that was its direct cellular counterpart or a close proxy; in only two, there was no match or a close proxy; and in the remaining three (esophageal, pancreatic ductal and biliary adenocarcinoma) the best matched cell type suggested metaplasia to stomack mucosa like tissue.The cellular context of breast tumor formation was investigated in more detail, showing that the COO, and not the gene inactivation event, determines the subtype. Basal-like tumors appeared to arise from luminal progenitor cells, while all other subtypes arose from mature luminal cells. Furthermore, irrespective of the inactivation mechanism (pathogenic germline, somatic truncating or epigenetic silencing event), all BRCA1/2- and RAD51C-altered basal-like tumors best matched to luminal progenitors while BRCA1/2- and CHEK2 -mutated luminal A/B subtypes best matched mature luminal cells. Finally, we observed that tumor type-specific driver genes reside in genomic regions that are defined by a highly active chromatin environment in their COOs. This highlights their essential role in cell type differentiation and implies the acquisition of somatic mutations early, when the chromatin architecture still reflected the COO. Taken together, our findings shed light on the crucial role of the COO in shaping the mutational landscape and tumor evolution.

Citation Format: Paz Polak, Rosa Karlic, Kirsten Kubler, William D. Foulkes, Gad Getz. The mutation landscape of cancers serves as a record of early malignant transformation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3304.
Abstract 3308: Urine epigenetic biomarkers for NSCLC diagnosis
Authors: Lerner Lane , Zheng Lily , Kottorou Anastasia , Chen Chen , Ito Tomoaki , Rodgers Kristen , Lee Beverly , Winn Robert , Benedetti Enrico , Wang Tza- Huei , Brock Malcolm V. , Herman James G. , Hulbert Alicia
Published: August 2018

Purpose:

The National Lung Cancer Screening Trial showed mortality reduction on patients with NSCLC by the use of Low-dose CT screening. The study exhibits an unreasonably high false positive rate of 96.6%, which can to lead to significant morbidity and mortality from unnecessary tests. This study aim to determine if methylated promoter regions of a panel of genes that are correlated with Non-Small Cell Lung Cancer can be detected in urine from patients with lung cancer versus those without.

Materials and Methods:

We conducted a prospective case-control study recruiting subjects from the Lung Cancer Spore Trial. Urine and pre-operative CT scans were obtained from all patients. We processed the urine using the Methylation on Beads assay to isolate and bisulfite treat circulating DNA and then employed Quantitative Methylation Specific Real-Time PCR to detect promoter methylation status of the genes: CDO1, TAC1, HOXA7, HOXA9, SOX17 and ZFP42. Sensitivity, specificity, PPV and NPV values were calculated for each gene methylation status.

Results:

34 patients were studied, including 23 patients with NSCLC and 11 patients with benign non-cancerous lesions. The sensitivity, specificity, PPV and NPV values for lung cancer detection in urine are shown in Table 1. Sensitivities ranged 43-96%, Specificities 64-91%, PPV 71-100% and NPV 40-67%. Promoter methylation of the gene panel CDO1, TAC1, HOXA7, HOXA9, SOX17 and ZPF42 has a 78% sensitivity, 91% specificity, 95% PPV and 67% NPV for lung cancer detection.

Conclusion:

Our study suggests that urine can provide a highly sensitive and specific non-invasive route for lung cancer detection. These results are very promising and could potentially reduce unnecessary morbidity and mortality in people undergoing screening for NSCLC by use of a non-invasive method that could be more accessible on primary care centers. However, further studies and validation with larger sample sizes and different populations are necessary before its application to clinical practice.

Patients with positive gene methylation from urine samples and its accuracy for NSCLC detection.Cancer (n=23)Cancer (n=23)Control (n=11)Control (n=11)nSensitivitynSpecificityPPVNPVCDO114611919353TAC11565010010058SOX1716704648050HOXA711483737940HOXA910431919143ZPF4222969187167Panel of genes18781919567

Citation Format: Lane Lerner, Lily Zheng, Anastasia Kottorou, Chen Chen, Tomoaki Ito, Kristen Rodgers, Beverly Lee, Robert Winn, Enrico Benedetti, Tza- Huei Wang, Malcolm V. Brock, James G. Herman, Alicia Hulbert. Urine epigenetic biomarkers for NSCLC diagnosis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3308.
Abstract 3313: Epigenetic biomarkers of prognosis in stage IIA colon cancer
Authors: Pagotto Sara , Porcellini Elisa , Bacalini Maria G. , Sarti Diletta , Garajova Ingrid , Broseghini Elisabetta , Laprovitera Noemi , Vasuri Francesco , Garagnani Paolo , Mariani-Costantini Renato , Basso Michele , Visone Rosa , Larocca Luigi M. , Ferracin Manuela , Veronese Angelo
Published: August 2018

Adjuvant therapy is a systemic treatment administered after primary tumor resection to reduce the risk of relapse and death in colorectal cancer (CRC) patients. Generally, adjuvant treatment is recommended for stage III and ‘high-risk' stage II CRC (NCCN guidelines). Nonetheless, adjuvant treatment is still debated for stage IIA (T3N0) patients, whose estimated recurrence rate is 15-20% in the absence of any further therapy after resection of the primary tumor. With the aim to find additional prognostic biomarkers that could improve current patient selection, we analyzed the global methylation profile of a discovery group of 10 relapsing/non-relapsing stage IIA colon cancer samples, without any other known risk factor (low-risk), by Illumina Epic 850K array. We identified a panel of 25 Illumina probes that were able to predict recurrence in chemotherapy-naïve patients. Then, we assessed the methylation of the most predictive Cytosine-Guanine (CpG) dinucleotides in 107 stage IIA colon cancer patients from two different cohorts, by EpiTYPER technology. We validated several CpGs as differentially methylated in relapsing vs. non-relapsing CRCs (p<0.05). For these CpG sites, we analyzed the methylation level also in the matching normal mucosa surrounding the tumor, to identify potential premalignant epigenetic lesions. In addition, we examined the association between the methylation level at specific CpG sites and prognostic clinical characteristics, overall survival and time-to-relapse. We concluded that the analysis of epigenetic alterations in archival colon cancer tissues could increase the panel of risk factors predictive for recurrence, and thus improve the selection of patients who could benefit from adjuvant therapy.

Citation Format: Sara Pagotto, Elisa Porcellini, Maria G. Bacalini, Diletta Sarti, Ingrid Garajova, Elisabetta Broseghini, Noemi Laprovitera, Francesco Vasuri, Paolo Garagnani, Renato Mariani-Costantini, Michele Basso, Rosa Visone, Luigi M. Larocca, Manuela Ferracin, Angelo Veronese. Epigenetic biomarkers of prognosis in stage IIA colon cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3313.
Abstract 3316: Hypermethylation at the SMPD3 promoter is frequently detected in hepatocellular carcinoma tumor tissue and corresponding plasma circulating DNA, and potentially differs by tumor subtype
Authors: Kwee Sandi Alexander , Peplowska Karolina , Tiirikainen Maarit , Song Min-Ae , Wong Linda L.
Published: August 2018

Background: Circulating plasma DNAs may exhibit altered methylation reflective of aberrant epigenetics in tumor tissues, which may present a clinical opportunity for non-invasive assessment of tumor molecular phenotype. Hypermethylation at the promoter region of SMPD3 (Sphingomyelin phosphodiesterase 3), a tumor suppressor gene, has been identified in several genome-wide DNA methylation studies involving tissue specimens of hepatocellular carcinoma (HCC) (Shen et al., 2012; Song et al., 2013). The purpose of this study was to measure and compare SMPD3 promoter hypermethylation (specifically at cg10426893) in tumors, adjacent liver tissue, and circulating cell-free DNA (cfDNA) to examine its potential as a biomarker of HCC corresponding with tumor molecular subtype.

Methods: Blood and liver tissue specimens were collected following written informed consent from 54 patients undergoing partial liver resection for BCLC stage A HCC. DNA methylation was quantified through pyrosequencing analysis of bisulfite-treated DNAs from corresponding tumor tissue, adjacent liver tissue, plasma, and buffy-coat samples. Results were compared within patients using paired t-test. Transcriptomic molecular sub-classification based on the Hoshida classification system (2009) was performed on a subset of 39 tumors to explore potential associations between SMPD3 promoter hypermethylation and HCC molecular subtype.

Results: Levels of SMPD3 promoter methylation were significantly higher in tumor tissue compared to adjacent liver tissue (37.0% vs. 8.8%, p < 0.0001) as well as when compared to cfDNA samples (32.9% vs. 14.3%, p < 0.0001). SMPD3 promoter methylation levels in cfDNA and tumor tissues were also significantly higher than in corresponding buffy-coat samples (p = 0.0146 and p < 0.0001, respectively). Tumors belonging to the S3 molecular sub-class (n = 22) exhibited significantly higher SMPD3 promoter methylation levels than non-S3 tumors (n = 17) (44.1 % vs. 24.1%, p = 0.0166).

Conclusions: Significant increases in DNA methylation involving the promoter region of SMPD3 in HCC tumor tissues are reflected in cfDNA, supporting potential clinical liquid biopsy applications. Preliminarily, this epigenetic alteration appears to be associated with the S3 (differentiation) molecular subtype of HCC.

Citation Format: Sandi Alexander Kwee, Karolina Peplowska, Maarit Tiirikainen, Min-Ae Song, Linda L. Wong. Hypermethylation at the SMPD3 promoter is frequently detected in hepatocellular carcinoma tumor tissue and corresponding plasma circulating DNA, and potentially differs by tumor subtype [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3316.
Abstract 3320: Discovery and development of DNA methylation biomarkers in human papillomavirus related oropharyngeal squamous cell carcinoma
Authors: Ren Shuling , Gaykalova Daria A. , Guo Theresa W. , Favorov Alexander V. , Fertig Elana J. , Khan Zubair , DeJong Peter M. , Ando Mizuo , Liu Chao , Sakai Akihiro , Fukusumi Takahito , Haft Sunny , Sadat Sayed , Gutkind J Silvio , Koch Wayne M. , Califano Joseph A.
Published: August 2018

Purpose: Human papillomavirus (HPV) related oropharyngeal squamous cell carcinoma (OPSCC) has an improved prognosis compared to cancers associated with tobacco and alcohol exposure, and also exhibits a different composition of epigenetic alterations. In this methylome-wide association study, we identified DNA methylation-based biomarkers with potential utility in screening for HPV positive OPSCC. Experimental Design: In the discovery stage, genome wide DNA methylation was measured using methyl-CpG binding domain protein-enriched genome sequencing (MBD-seq) in 50 HPV positive OPSCC tissues and 25 normal tissues. We defined 51 differentially methylated regions (DMRs) with maximal methylation specificity to cancer samples. We used TCGA methylation array data to evaluate the performance of the proposed candidates. Top 20 candidates were validated using quantitative methylation specific PCR (QMSP) in independent 24 HPV related OPSCCs and 22 controls. Results: Supervised hierarchical clustering of 51 DMRs based on TCGA methylation data found that HPV positive OPSCC had significantly higher DNA methylation levels compared to normal samples and head and neck squamous cell carcinoma (HNSCC) without HPV or with different tumor sites. The methylation levels of all top 20 candidates in HPV positive OPSCC were significantly higher than those in normal samples in TCGA data. 16 of these 20 candidates were found by QMSP to have significant higher methylation levels compared with controls. 2 candidates (CCDC181 and OR6S1) had a sensitivity of 100%, while 10 candidates (KCNA3, ITGA4, BEND4, ELMO1, SFMBT2, HOXB4, ZNF439, ZNF93, VSTM2B and ZNF773) had a specificity of 100%. Our 20 candidates were found by the receiver operating characteristic analysis to have a prediction accuracy ranging from 56.3% to 99.8%. Conclusions: This study discovered 20 epigenetic markers for detection of HPV related OPSCC, which can potentially be applied to develop a population based screening test and improve disease management.

Citation Format: Shuling Ren, Daria A. Gaykalova, Theresa W. Guo, Alexander V. Favorov, Elana J. Fertig, Zubair Khan, Peter M. DeJong, Mizuo Ando, Chao Liu, Akihiro Sakai, Takahito Fukusumi, Sunny Haft, Sayed Sadat, J Silvio Gutkind, Wayne M. Koch, Joseph A. Califano. Discovery and development of DNA methylation biomarkers in human papillomavirus related oropharyngeal squamous cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3320.
Abstract 3323: Role of RNA methyltransferase METTL3 in melanoma
Authors: Dahal Ujwal , Le Kang , Gupta Mamta
Published: August 2018

Melanoma is the sixth most common cancer in the USA. Introduction of BRAF inhibitors and advances in cancer immunotherapy has greatly improved treatment of melanoma, however, the problem of tumour relapse and therapy resistance persists. Better understanding of the mechanism of development of the diseases is urgently needed to improve therapeutic intervention. Role of DNA and histone modifications have been well established in several diseases, however, RNA modifications, specifically, N6 methyl adenosine (m6A), has only recently begun to be shown to play critical role in cancer. m6A plays crucial roles in mRNA stability, splicing, export, translation and decay and is regulated by: methyltransferase complex proteins METTL3, METTL14, and WTAP; the demethylases FTO and ALKBH5, and the m6A reader proteins. In the present study, we uncover the role of RNA methyltransferase METTL3 in human melanoma cells. METTL3 expression was assessed in 8 melanoma cell lines (A375, A375MA2, A2058, WM164, 451Lu, WM793, SKMEL2, WM3918). Western blotting data revealed METTL3 to be upregulated in seven melanoma cell lines (except A2058) relative to normal melanocytes. To uncover the role of METTL3 in melanoma, we knockdown METTL3 with lentiviral shRNA and studied the effect on melanoma cells tumorigenesis. The colony formation assay revealed reduced colony number in the METTL3 knockdown A375, A375MA2 and WM793 cells relative to the scrambled control cells. The main cause of melanoma associated fatality is its metastatic dissemination we therefore, next sought to investigate whether METTL3 could affect melanoma cells migration and invasiveness. We performed migration and invasion assays to detect the migratory potential of control or METTL3 knockdown A375, A375MA2 and WM793 cells. Interestingly, both migration and invasion was found to be reduced in METTL3 knockdown cells in all the three cell lines examined. RNA dot blot assay demonstrated a decreased m6A level in total RNA of the METTL3 knockdown cells relative to control cells. p16 and p53 mRNA were found to be enriched in m6A immunoprecipitated samples as compared to IgG control, pointing towards a potential role of m6A in regulating expression of melanoma associated genes. Ongoing effort is aimed at identifying what known or novel regulators of the migratory/invasive cascade is regulated by METTL3 and whether it is through m6A mediated or independent mechanisms. In summary, we have shown that METTL3 is overexpressed in human melanoma and play an important role in metastasis.

Citation Format: Ujwal Dahal, Kang Le, Mamta Gupta. Role of RNA methyltransferase METTL3 in melanoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3323.
Abstract 3346: PRMT5-mediated methylation of YBX1 regulates NF-kB activity in colorectal cancer
Authors: Hartley Antja-Voy , Wang Benlian , Miyagi Masaru , Mundade Rasika , Hamilton James , Lu Tao
Published: August 2018

Colorectal cancer (CRC) is the second leading cause of cancer-related deaths in the US with a staggeringly less than 14% survival rate for patients with metastatic disease. This poor clinical outcome for CRC patients is largely attributed to the lack of understanding of the factors that drive CRC progression, leading to a dearth of available treatment strategies for this disease. Today, treatment modalities and clinical management of colorectal cancer (CRC) are fundamentally based on the success of controlling specific tumorigenic pathways. Interestingly, aberrant activation of the transcription factor nuclear factor ĸB (NF-ĸB) is increasingly recognized as a crucial player in CRC progression. Therefore, understanding the mechanisms underlying regulation of NF-ĸB holds great promise for devising new therapeutic strategies for CRC. Recently, we identified the oncogenic protein, Y-box binding protein 1 (YBX1), as a novel activator of NF-κB. Since YBX1 overexpression has also been linked to poor patient outcome in CRC, we sought to further understand whether this YBX1/NF-ĸB axis could be a potential target for CRC treatment. In this study, we discovered that YBX1 is methylated on arginine 205 (YBX1-R205me2), an event that is critical for YBX1-mediated NF-ĸB activation and target gene expression. Additionally, co-immunoprecipitation studies revealed that the R205 to alanine (A) mutant (YBX1-R205A) also significantly diminished the interaction between YBX1 and the p65 subunit of the transcriptionally active NF-ĸB, demonstrating a novel mechanism by which methylation of YBX1 mediates protein-protein interactions. Overexpression of YBX1-R205A significantly attenuated the migration, proliferation, and anchorage-independent growth of a panel of CRC cells, suggesting that YBX1-R205me2 is essential to the oncogenic functions exerted by the YBX1/NF-ĸB axis in CRC. Furthermore, we revealed that protein arginine methyltransferase 5 (PRMT5) is responsible for the methylation of YBX1-R205. Collectively, our novel findings present a complex picture of the sophisticated regulation of NF-ĸB through PRMT5-mediated YBX1-R205 methylation and suggest that pharmacological disruption of the YBX1/NF-κB axis using PRMT5 inhibitors could serve as the basis for new therapeutics that impede YBX1/NF-κB-driven CRC progression.

Citation Format: Antja-Voy Hartley, Benlian Wang, Masaru Miyagi, Rasika Mundade, James Hamilton, Tao Lu. PRMT5-mediated methylation of YBX1 regulates NF-kB activity in colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3346.
Abstract 3353: ZNF414 as a functionally relevant transcription factor in pancreatic and breast cancer cells
Authors: Martínez Alejandra Rodríguez , Vuorinen Elisa M. , Shcherban Anastasia , Rajala Nina K. , Nykter Matti , Kallioniemi Anne
Published: August 2018

Background: The Zinc Finger Protein 414 (ZNF414) is a member of the krüppel C2H2-type zinc-finger protein family. ZNF414 is a cargo protein for Karyopherin α7 (KPNA7), a nuclear importer expressed during embryogenesis, absent in most adult tissues but re-expressed in cancer cells. KPNA7 is involved in promoting proliferation and maintaining nuclear morphology in several breast and pancreatic cancer cell lines. Similar effects on cell growth have been evidenced for ZNF414 using in vitro knock-down experiments. Other than this, the function of ZNF414 remains uncharacterized but, as a zinc finger protein with nuclear localization, it is likely to act as a transcription factor. This study aimed at identifying target genes and DNA binding motifs of ZNF414 in pancreatic and breast cancer cells. Methods: We used next generation sequencing methods on Hs700T and MCF-7 cell lines. RNA-seq was used to identify genes regulated by ZNF414 and ChIP-exo and ATAC-seq analyses to uncover its genomic binding regions. Samples for RNA-seq were collected 12h and 24h after transfection with siRNAs targeting ZNF414. For ChIP-exo and ATAC-seq experiments, cells were transfected to transiently overexpress V5-tagged ZNF414. Results: RNA-seq data analyses revealed 33 and 296 differentially expressed genes (DEGs) in Hs700T cells at 12h and 24h time points, respectively. The corresponding amounts of DEGs in MCF-7 cell line were 177 and 556. There were 23 and 108 DEGs in common to both cell lines at 12h and 24h, respectively. Interestingly, gene ontology analyses revealed enrichment of many functional categories related to cellular proliferation in both cell lines, providing a molecular explanation for the observed ZNF414-elicited phenotype. ChIP-exo data was processed using MACE tool and then combined with the ATAC-seq open chromatin signal to identify the most reliable peaks. We looked at the genomic location of these peaks and observed clear enrichment in the promoter and 5'UTR regions, indicating that ZNF414 is a classical transcription factor. Using MEME for de novo motif discovery analyses, we identified several putative binding motifs common to both cell lines, some of which were also found in promoters of DEGs. Conclusion: This study uncovered the transcriptional regulation that underlies the role of ZNF414 as inducer of cellular proliferation in pancreatic and breast cancer cells.

Citation Format: Alejandra Rodríguez Martínez, Elisa M. Vuorinen, Anastasia Shcherban, Nina K. Rajala, Matti Nykter, Anne Kallioniemi. ZNF414 as a functionally relevant transcription factor in pancreatic and breast cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3353.
Abstract 3354: Chemically induced degradation of the transcription factor BCL6
Authors: Kerres Nina , Steurer Steffen , Schlager Stefanie , Bader Gerd , Caligiuri Maureen , Dank Christian , Engen John R. , Ettmayer Peter , Gerlach Daniel , Gerstberger Thomas , Han Bingsong , Iacob Roxana E. , Kessler Dirk , Lancia David R. , Moriz Mayer , Mischerikow Nikolai , Rumpel Klaus , Schnitzer Renate , Voss Tilman , Zheng Xiaozhang , Zoephel Andreas , Kraut Norbert , McConnell Darryl , Pearson Mark , Koegl Manfred
Published: August 2018

The transcription factor BCL6 is a known driver of oncogenesis in lymphoid malignancies, including diffuse large B-cell lymphoma (DLBCL). Disruption of its interaction with transcriptional repressors interferes with the oncogenic effects of BCL6. We have used structure-based drug design to develop highly potent compounds that block this interaction. A subset of these inhibitors also cause rapid ubiquitylation and degradation of BCL6 in cells. These compounds display significantly stronger induction of expression of BCL6-repressed genes and antiproliferative effects than compounds that merely inhibited co-repressor interaction. The fact that the magnitude of effects elicited by this class of BCL6 degrading compounds greatly exceeds that of our equipotent non-degrading inhibitors offers exciting opportunities for the development of BCL6-based lymphoma therapeutics. To support further research, the most potent BCL6 degrading inhibitor is made freely available to the research community as an in vitro tool compound. Please see http://www.opnMe.com for further infos.

Citation Format: Nina Kerres, Steffen Steurer, Stefanie Schlager, Gerd Bader, Maureen Caligiuri, Christian Dank, John R. Engen, Peter Ettmayer, Daniel Gerlach, Thomas Gerstberger, Bingsong Han, Roxana E. Iacob, Dirk Kessler, David R. Lancia, Mayer Moriz, Nikolai Mischerikow, Klaus Rumpel, Renate Schnitzer, Tilman Voss, Xiaozhang Zheng, Andreas Zoephel, Norbert Kraut, Darryl McConnell, Mark Pearson, Manfred Koegl. Chemically induced degradation of the transcription factor BCL6 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3354.
Abstract 3356: The EWS/FLI1 transcriptome is characterized by marked heterogeneity across Ewing sarcoma cell lines
Authors: Kitchen-Goosen Susan M. , Bowman Megan J. , Adams Marie , Berger Penny , Grohar Patrick J.
Published: August 2018

BACKGROUND: Ewing sarcoma (ES) is the second most common pediatric bone cancer. ES cells are dependent on the activity of the EWS/FLI1 transcription factor for cell survival. EWS/FLI1 drives proliferation, dysregulates the cell cycle and establishes a block in differentiation. Despite the known dependence of Ewing sarcoma on EWS/FLI1, the exact identity of all the downstream targets of EWS/FLI1 is not known. While gene signatures have been reported for EWS/FLI1, these fail to capture the marked heterogeneity in the EWS/FLI1 transcriptome or correlate these expression changes with cellular phenotype. METHODS: Next generation RNA sequencing with deep coverage was used to generate a comprehensive analysis of the EWS/FLI1 transcriptome by silencing EWS/FLI1 in 5 different ES cell lines at two different time points. These results were organized into gene expression networks and signaling nodes were identified by weighted correlation network analysis (WGCNA). The networks were validated by a 500-gene signature of EWS/FLI1 targets using a next generation sequencing capture assay and the Illumina platform. The cellular endpoint of EWS/FLI1 silencing was evaluated by caspase 3,7 cleavage to measure apoptosis, BGAL staining to measure senescence and scratch assays to capture migration. RESULTS: Next generation RNA sequencing with siRNA silencing of EWS/FLI1 revealed marked heterogeneity in the EWS/FLI1 transcriptome among cell lines. Targets of EWS/FLI1 that were common across cell lines exhibited small magnitude gene expression changes (log fold <2), while large magnitude gene expression changes were unique to individual cell lines (log fold>2). When the log fold change in expression was restricted to greater than 2, only 13 common repressed targets and 4 common induced targets were identified in all 5 cell lines. In addition, silencing of EWS/FLI1 induced dramatically different phenotypes across cell lines, where p53 wild-type cell lines underwent senescence and the p53 mutant cell lines did not. Most cell lines did not continue to proliferate with EWS/FLI1 silencing with the exception being the widely used A673 cell line. Cellular migration did not change with EWS/FLI1 silencing in many of the cell lines in contrast to A673 cells. CONCLUSIONS: EWS/FLI1 drives different gene expression signatures and different phenotypes in unique cell lines. This diversity needs to be considered as EWS/FLI1 directed therapies are developed.

Citation Format: Susan M. Kitchen-Goosen, Megan J. Bowman, Marie Adams, Penny Berger, Patrick J. Grohar. The EWS/FLI1 transcriptome is characterized by marked heterogeneity across Ewing sarcoma cell lines [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3356.
Abstract 3365: MAP3K10 mutation as a biomarker for predicting response to chemotherapy in gastric cancer patients
Authors: Kwon Chae Hwa , Lee Sun Jin , Choi Yuri , Park Do Youn
Published: August 2018

Gastric cancer is one of most common malignant tumors. Although chemotherapy can improve the survival of gastric cancer, the benefit is limited because most cancer cells eventually become irresponsive to chemotherapeutic drugs. To investigate mechanism of resistance to chemotherapy, we performed whole-exome sequencing (WES) on resistant tumor and paired normal samples from 14 patients received adjuvant chemotherapy. We found that 3 patients had novel somatic mutations in mitogen-activated protein kinase kinase kinase 10 (MAP3K10), an upstream kinase of mitogen activated protein kinase (MAPK). MAP3K10R304C, MAP3K10H211R, or MAP3K10A30E were detected each patient, respectively. The survival rate was higher and apoptosis rate was smaller in the cells expressing each MAP3K10 mutations compared with parental controls and cells expressing wild-type MAP3K10, when treated with cisplatin or 5-FU. Moreover, cells expressing each MAP3K10 mutations exhibited higher levels of phosphorylated MEK1/2 and p38MAPK at baseline and when treated with cisplatin or 5-FU than wild-type AGS or MKN45 cells or those expressing wild-type MAP3K10. In summary, MAP3K10R304C, MAP3K10H211R, and MAP3K10A30E MAP3K10 confer resistance to cisplatin or 5-FU via activation of MAPK pathway in gastric cancer cells. Thus, somatic MAP3K10 mutations could be a potential biomarker for predicting resistance to chemotherapy for gastric cancer patients

Citation Format: Chae Hwa Kwon, Sun Jin Lee, Yuri Choi, Do Youn Park. MAP3K10 mutation as a biomarker for predicting response to chemotherapy in gastric cancer patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3365.
Abstract 3366: APOBEC3 family of cytidine deaminases in sensitizing triple-negative breast cancer cells to cisplatin and carboplatin
Authors: Conner Kayla L. , Shaik Asra N. , White Jordan , Lei Wen , Cote Michele L. , Patrick Steve M.
Published: August 2018

Cisplatin is a DNA damaging chemotherapeutic which forms interstrand crosslinks (ICL) and intrastrand adducts that induce replication fork collapse and lead to apoptosis. These adduct sites can be repaired through several repair pathways, the most important being nucleotide excision repair (NER). Cisplatin and carboplatin ICLs form a unique structure that forces cytosines flanking the adduct site to become extrahelical. Our lab has previously shown that base excision repair (BER) and mismatch repair (MMR) mediate sensitivity to cisplatin and carboplatin as a consequence of nonproductive processing of DNA flanking the ICL structure. The APOBEC3 (A3) protein family of cytidine deaminases have recently been implicated in cancer development and have potential to deaminate the extrahelical cytosines formed by cisplatin ICLs. Polβ knockdown MDA-MB-231 cells are resistant to cisplatin in vivo compared to wild-type cells (p<0.001). Knockdown of APOBEC3 family members in MDA-MB-231 breast cancer cells results in a resistant phenotype to cisplatin and carboplatin treatment compared to the control. Utilizing CRISPR-Cas9 genome editing, we generated knockouts of A3 members. A3D knockdown in the APOBEC3C (A3C) knockout results in resistance to cisplatin compared to the control, suggesting that multiple A3 enzymes can deaminate the ICL induced extrahelical cytosine. APOBEC3A (A3A) knockdown in a cell line with high expression of A3A (SK-BR-3) shows no difference in cisplatin and carboplatin treatment, suggesting that A3A does not deaminate extrahelical cytosines. Overexpression of A3C or A3D in cell lines with low expression of A3 proteins (SK-BR-3) results in sensitivity to cisplatin. These results together suggest that A3C, A3D, and A3G can deaminate the ICL extrahelical cytosines, resulting in the activation of BER. The subsequent nucleotide misincorporation by Polβ followed by MMR protein binding would physically block NER proteins from repairing the ICL, therefore conferring sensitivity to cisplatin and carboplatin.

Citation Format: Kayla L. Conner, Asra N. Shaik, Jordan White, Wen Lei, Michele L. Cote, Steve M. Patrick. APOBEC3 family of cytidine deaminases in sensitizing triple-negative breast cancer cells to cisplatin and carboplatin [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3366.
Abstract 3367: USP7 regulates CSB protein recovery after UV-induced DNA damage
Authors: Wei Shengcai
Published: August 2018

Cockayne syndrome group B (CSB) protein functions in transcription-coupled repair (TCR), a sub-pathway of nucleotide excision repair (NER). We previously demonstrated that ultraviolet radiation (UVR) induces a valosin-containing protein (VCP)/p97-dependent ubiquitin-mediated CSB degradation. In this study, we further revealed that CSB protein is ubiquitinated immediately after ultraviolet-light irradiation and undergoes proteasomal degradation. The recovery of CSB protein level is dependent on USP7. Disruption of USP7 function sensitizes CSB degradation to lower UVR doses. We further demonstrated USP7's interaction with CSB in vivo by immunoprecipitation and in vitro by GST pulldown assays. Interestingly, in vitro CSB strongly binds to TRAF domain, which also interacts with UV stimulated scaffold protein A (UVSSA). Additionally, USP7 also interacts with VCP/p97 using its TRAF domain and C-terminal in in vitro. Furthermore, VCP/p97 was demonstrated to associates with USP7 in vivo. These results suggest that USP7 rescues CSB from ubiquitin-mediated proteolysis after (VCP)/p97-mediated CSB extraction from chromatin, preventing the CSB-ubiquitin conjugates being presented to proteasome for degradation.

Citation Format: Shengcai Wei. USP7 regulates CSB protein recovery after UV-induced DNA damage [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3367.
Abstract 3376: Early onset of chromosomal instability in breast preneoplasia detected by single-cell genomics
Authors: Fagan-Solis Katerina , Simpson Dennis A. , Martelotto Luciano , Reis-Filho Jorge S. , Petrini John H. , Gupta Gaorav P.
Published: August 2018

Chromosomal rearrangements and copy number aberrations are a major mechanism for driver gene alterations in breast cancer. However, the origins of chromosomal instability during breast tumorigenesis are poorly understood. We have recently shown that oncogene expression in normal mammary epithelial cells induces replication stress and a DNA damage response (DDR). The double-strand break sensor Mre11 is required for the oncogene-induced DDR, and suppression of Mre11 is sufficient to promote the development of Her2/Neu-driven breast cancer in a transgenic mouse model. Here we have applied single-cell genomics to primary mammary epithelial cells from this mouse model and demonstrate a profound effect of oncogene expression on chromosomal instability in preneoplastic cells, which is further augmented in Mre11-mutant cells. Her2/Neu-induced chromosomal breakpoints are highly enriched in the vicinity of large genes, suggesting a possible role for gene transcription and/or R-loops in the origin of oncogene-induced genomic fragility. Mre11 hypomorphism does not affect the number of chromosomal breakpoints but alters the genomic aberration signature to one that is over-represented by larger-size deletions. These findings indicate that chromosomal instability is an early event during Her2/Neu-initiated breast neoplasia. Furthermore, DDR perturbation—in the form of Mre11 hypomorphism—is not required for oncogene-induced genomic instability but rather alters the pattern of observed copy number aberrations and promotes the proliferative expansion of genomically unstable clones. Thus, single-cell genomics applied to murine models of breast preneoplasia reveals insights into the origins of chromosomal instability in breast cancer that may inform novel strategies to prognosticate or prevent the progression of high-risk premalignant breast lesions.

Citation Format: Katerina Fagan-Solis, Dennis A. Simpson, Luciano Martelotto, Jorge S. Reis-Filho, John H. Petrini, Gaorav P. Gupta. Early onset of chromosomal instability in breast preneoplasia detected by single-cell genomics [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3376.
Abstract 338: A new investigational ATM Inhibitor, M3541, synergistically potentiates fractionated radiotherapy and chemotherapy in cancer cells and animal models
Authors: Zimmermann Astrid , Zenke Frank , Dahmen Heike , Sirrenberg Christian , Grombacher Thomas , Vassilev Lyubomir T. , Fuchss Thomas , Blaukat Andree
Published: August 2018

Physical or chemical DNA damaging agents are among the most widely used classes of cancer therapeutics today. Radiotherapy and topoisomerase inhibitors induce double strand breaks (DSB) considered most dangerous if left unrepaired because they can induce growth arrest, chromosomal abnormality and/or cell death. ATM is an essential kinase which orchestrates one of the two major pathways for repair of DSBs via the high fidelity homologous recombination (HR) mechanism. In addition, ATM acts as an upstream signaling kinase, regulating cell cycle checkpoint control in the response to DSB damage. Inhibitors of ATM kinase are expected to suppress DSB DNA repair, block checkpoint controls and enhance the therapeutic effect of radiation therapy and other DSB-inducing treatment modalities. We have developed a novel, highly potent and selective inhibitor of ATM, M3541, which modifies cellular response to DSB damage leading to potentiation of radiation therapy and DSB-inducing chemotherapy. Here, we present data from the preclinical evaluation of M3541 therapeutic potential in cellular and xenograft models of cancer. ATM inhibitor sensitized multiple tumor cell lines to ionizing radiation and a topoisomerase inhibitor in vitro. Oral administration of M3541 to nude mice bearing human tumor xenografts in a clinically relevant radiation regimen strongly enhanced the antitumor activity, leading to complete tumor regression. These activities resulted from inhibition of ATM kinase activity and modulation of its downstream effects in the xenograft tissues. M3541 is currently under clinical investigation in a Phase I trial.

Citation Format: Astrid Zimmermann, Frank Zenke, Heike Dahmen, Christian Sirrenberg, Thomas Grombacher, Lyubomir T. Vassilev, Thomas Fuchss, Andree Blaukat. A new investigational ATM Inhibitor, M3541, synergistically potentiates fractionated radiotherapy and chemotherapy in cancer cells and animal models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 338.
Abstract 3380: BRIP1/FANCJ is vital for efficient response to and repair of replication-associated DNA damage
Authors: Clark David W. , Mani Chinnadurai , Palle Komariah
Published: August 2018

BRCA1-Interacting Protein 1 (BRIP1), a DNA helicase and tumor suppressor also known as FANCJ, is implicated in breast and ovarian cancers as well as the genetic instability syndrome Fanconi anemia (FA) which predisposes patients to multiple neoplasms. This study investigates the role of FANCJ in repair of replication-associated DNA damage which can lead to DNA strand breaks, chromosomal instability, and tumorigenesis. FANCJ has long been known to function in repair of DNA crosslinks; however, its role in protecting cells from replication-associated DNA damage is less well known. We have shown that loss of FANCJ sensitizes cells to damage caused by compounds that block DNA replication, such as hydroxyurea (HU) and the topoisomerase I poison camptothecin (CPT). Loss of FANCJ disrupts the cell's response to replication-associated DNA damage by impairing Chk1 and Chk2 phosphorylation and stability and activation of multiple DNA repair proteins. FANCJ-deficient cells exposed to low levels of CPT failed to progress through S-phase indicating that FANCJ is important for intra-S phase DNA damage checkpoint and recovery from replication fork blocks. FANCJ depletion leads to a 90% decrease in homologous recombination repair in assays designed to measure repair of DNA double strand breaks as well as a loss of expression of repair-associated proteins and DNA repair foci in response to damage from HU and CPT. Loss of FANCJ impairs DNA replication and leads to accumulation of gross chromosomal aberrations. Importantly, some of the DNA damage response functions of FANCJ are independent of its helicase activity, as a helicase-deficient mutant is able to compensate for loss of FANCJ in activation and stability of FANCD2 and other HR proteins. Collectively our findings indicate a novel role for FANCJ as a key regulator of the DNA damage response to errors in DNA replication potentially leading to mutagenesis and tumorigenesis.

Citation Format: David W. Clark, Chinnadurai Mani, Komariah Palle. BRIP1/FANCJ is vital for efficient response to and repair of replication-associated DNA damage [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3380.