National Cancer Institute

National Cancer Institute

NCI Second Symposium on

Second Symposium on Translational Genomics March 15–16, 2012 Natcher Auditorium National Institutes of Health, Bethesda, MD

Sponsor: Center for Cancer Research The Center of Excellence in Integrative Cancer Biology and Genomics Organizing Committee: Laufey Amundadottir, Curt Harris, Tim Harris, Javed Khan, Glenn Merlino, Thomas Ried, and Snorri Thorgeirsson

U.S. DEPARTMENT OF HEALTH AND HUMAN SERVICES • National Institutes of Health

Dear Colleagues, On behalf of the National Cancer Institute, the Center for Cancer Research, and the Center of Excellence in Integrative Cancer Biology and Genomics, it is our great pleasure to welcome you to this symposium. The mission of the Center of Excellence in Integrative Cancer Biology and Genomics (CEICBG) is to promote the innovative use of genetic and genomic approaches and technologies for basic science discoveries and clinical research applications for the prevention, diagnosis, and treatment of cancer. Towards this end, this symposium brings together internationally renowned experts in the areas of Noncoding RNAs, Next Generation Sequencing and Epigenomics and Genetic Variation. We thank you for your participation in this event. We hope this symposium offers you an opportunity to learn more about the current status of translational genomics, share your own research and discuss the use and implications of these advances for clinical applications. Sincerely, The Center of Excellence in Integrative Cancer Biology and Genomics symposium organizing committee: Laufey Amundadottir, Ph.D. Curt Harris, M.D. Tim Harris, Ph.D., D.Sc. Javed Khan, M.D. Glenn Merlino, Ph.D. Thomas Ried, M.D. Snorri Thorgeirsson, M.D., Ph.D.

Second Symposium on Translational Genomics Natcher Auditorium, National Institutes of Health, Bethesda, MD

AGENDA

Thursday, March 15, 2012 Welcome and Introductions 12:20 p.m.

Snorri Thorgeirsson, Head, Center of Excellence in Integrative Cancer Biology and Genomics

Opening Address 12:30 p.m.

Eric Green, National Human Genome Research Institute “Charting a Course for Genomic Medicine”

Keynote Address 1:00 p.m.

David Goldstein, Duke University "Complete sequencing and human disease: A genetic approach to genomics"

Session I: Noncoding RNAs: The Dark Matter of the Genome (Chairs: Laufey Amundadottir and Curt Harris) 1:35 p.m.

Joshua Mendell, University of Texas Southwestern Medical Center "MicroRNA reprogramming in cancer: mechanisms and consequences"

2:10 p.m.

Joanne Weidhaas, Yale University "The role of germ-line microRNA variants in cancer risk, biology and outcome"

2:45 p.m.

BREAK

3:05 p.m.

Curt Harris, National Cancer Institute “Interweaving Inflammation, MicroRNA and Senescence Networks in Human Cancer”

3:35 p.m.

Natasha Caplen, National Cancer Institute “Non-coding RNAs of the 8q24 locus”

4:05 p.m.

Carlo Croce, Ohio State University "Causes and Consequences of microRNA Dysregulation in Cancer"

4:35 p.m.

Poster Session

6:00 p.m.

Adjourn

1 Translational Genomics

March 15-16, 2012

Friday, March 16, 2012 Keynote Address 8:30 a.m.

Barbara Wold, California Institute of Technology "Translational Genomics: From 3-D Long-Distance Interactions to the Passenger/Driver Gene Repertoire in Cancer"

Session II: Next Generation Sequencing for Next Generation Medicine (Chairs: Tim Harris and David Goldstein) 9:05 a.m.

Tim Harris, Biogen Idec "The relevance of patient stratification in oncology for patients with multiple sclerosis and other neurological disorders"

9:40 a.m.

David Ting, Massachusetts General Hospital “Aberrant Expression of Satellite Repeats in Pancreatic Cancer”

10:15 a.m.

BREAK

10:35 a.m.

Elaine Mardis, Washington University “Integrated genomic data informs cancer treatment”

11:05 a.m.

Steve Lincoln, Complete Genomics “Comprehensive Detection of Variation in Thousands of Germ-Line and Tumor Genomes”

11:35 a.m.

Javed Khan, National Cancer Institute “Comprehensive Analysis of the Cancer Genome: Towards Precision Therapy in Cancer”

12:05 p.m.

LUNCH

Session III: Epigenomics and genetic variation (Chairs: Snorri Thorgeirsson and Thomas Ried) 1:05 p.m.

Manel Esteller, Bellvitge Biomedical Research Institute (IDIBELL), Barcelona “Human Cancer Epigenetics and Beyond”

1:40 p.m.

Peter Jones, University of Southern California “The Cancer Epigenome”

2:15 p.m.

Jean-Pierre Issa, Temple University School of Medicine “Integrated DNA methylation and chromatin maps to identify epigenetically silenced cancer drivers”

2:50 p.m.

BREAK

3:10 p.m.

Paul Meltzer, National Cancer Institute "Bringing the New Sequencing Technologies to the Oncology Clinic: Opportunities and Obstacles"

3:40 p.m.

Olufunmilayo Olopade, University Of Chicago “Deciphering Early Onset Breast Cancer in Women of African Ancestry via Integrated"-omics”

4:10 p.m.

Laufey Amundadottir, National Cancer Institute "The Pancreatic Cancer Transcriptome and Epigenome"

Closing Remarks 4:40 p.m.

Thomas Ried, Chair, Symposium Planning Committee


March 15-16, 2012

Speaker Abstracts


March 15-16, 2012

Charting a Course for Genomic Medicine. Eric D. Green. National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland, USA. The Human Genome Project’s generation of a reference human genome sequence was a landmark scientific achievement of historic significance. It also signified a critical transition for the field of genomics, as the new foundation of genomic knowledge started to be used in powerful ways by researchers and clinicians to tackle increasingly complex problems in biomedicine. To exploit the opportunities provided by the human genome sequence and to ensure the productive growth of genomics as one of the most vital biomedical disciplines of the 21st century, the National Human Genome Research Institute (NHGRI) is pursuing a broad vision for genomics research beyond the Human Genome Project. This vision includes using genomic data, technologies, and insights to acquire a deeper understanding of biology and to uncover the genetic basis of human disease. Some of the most profound advances are being catalyzed by revolutionary new DNA sequencing technologies; these methods are producing prodigious amounts of DNA sequence data, including data from large numbers of individual patients. Such capabilities are accelerating our understanding of the genetic basis for rare and common diseases, for cancer, and for drug response. Together, these developments are ushering in the era of genomic medicine.


March 15-16, 2012

Complete Sequencing and Human Disease: A Genetic Approach to Genomics. David Goldstein. Duke University Medical Center, Durham, North Carolina, USA. Genome wide association studies have proven successful in identifying regions of the genome that contain gene variants that influence both common diseases and drug responses. In most instances however it has not been possible to track these associations down to the causal variants that are responsible, and this greatly reduces the importance of the findings in terms of understanding disease biology, providing pointers to new treatments, and in prognosis. Sequencing based strategies on the other hand offer the promise of identifying the precise mutations and the genes they influence that are responsible for predisposition to common disease, congenital abnormalities, and drug responses. As sequencing approaches rapidly replace GWAS in the study of human disease it will become increasingly important to unite genomics research with laboratory based research and clinical research. Here I outline some of the near term opportunities and challenges in sequencing based discovery genetics.


March 15-16, 2012

MicroRNA Reprogramming in Cancer: Mechanisms and Consequences. Joshua T. Mendell. Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, Texas, USA. miRNA gain- and loss-of-function can potently influence cellular behavior in normal physiologic states and in diseases such as cancer. The regulation of miRNA expression and activity by cellular signaling cascades can therefore result in dramatic phenotypic outputs. We previously demonstrated extensive control of miRNA expression by wellcharacterized oncogenic and tumor suppressor networks including the Myc, Kras, and p53 pathways. We are now employing novel mouse models with gain and loss of miRNA function to investigate the roles of the miRNAs embedded within these signaling pathways and the pathologic consequences when their functions are disrupted. Insights gained from these functional studies have led to the development of novel therapeutic strategies for cancer and other pathologic states based on miRNA delivery. I will present our latest findings related to miRNA regulation in cancer and exploitation of these findings for the development of novel therapeutic approaches.


March 15-16, 2012

The Role of Germ-line MircoRNA Variants in Cancer Risk, Biology and Outcome. Joanne Weidhaas. Yale University, New Haven, Connecticut, USA. Our group has identified a novel class of cancer marker, microRNA (miRNA)-binding site variants. We have found these germ-line mutations in the 3’ untranslated region (UTR) of important oncogenes, where they disrupt miRNA-mediated gene regulation. We have shown that the first discovered variant in this class, the KRAS-variant (in a let-7 miRNA complementary site in the KRAS 3’UTR), predicts an increased risk of several types of cancer, including non-small cell lung cancer (NSCLC) 1, ovarian cancer 2 and triple negative breast cancer (TNBC) 3. In addition, we have found that the KRAS-variant predicts for multiple cancers in the same individual. Since miRNA expression is tissue dependent, we hypothesize that some of the organ specificity is determined by baseline miRNA regulation in the target tissues, as these variants are dynamically regulated by miRNAs. In addition, likely because miRNAs are fundamental in oncogenesis and critically involved in the response to cytotoxic cancer therapy 4, we and others have also found that tumors in patients with these miRNA-binding site variants are inherently biologically differently, and respond differently to cancer therapies 5. 1. Chin L, Ratner E, Leng S, Zhai R, Nullur S, Babar I, et al. A SNP in a let-7 microRNA complementary site in the KRAS 3' untranslated region increases non-small cell lung cancer risk. Cancer Res. 2008; 68: 8535-40. 2. Ratner E, Lu L, Boeke M, Barnett R, Nallur S, Chin L, et al. A KRAS-variant in Ovarian Cancer Acts as a Genetic Marker of Cancer Risk. Cancer Research. 2010; 15: 6509-15. 3. Paranjape T, Heneghan H, Lindner R, Keane F, Hoffman A, Hollestelle A, et al. A 3'-untranslated region KRAS variant and triple-negative breast cancer: a case-control and genetic analysis. Lancet Oncology. 2011. 4. Weidhaas J, Babar I, Nallue S, Trang P, Roush S, Boehm M, et al. MicroRNAs as Potential Agents to Alter Resistance to Cytotoxic Anticancer Therapy. Cancer Research. 2007; 67. 5. Graziano F, Canestrare E, Loupakis F, Ruzzo A, Galluccio N, Santini D, et al. Genetic modulation of the let-7 microRNA binding to KRAS 3'-untranslated region and survival of metastatic colorectal cancer patients treated with salvage cetuximab-irinotecan. The Pharmacogenomics Journal. 2010; 10(5): 458-64.


March 15-16, 2012

Interweaving Inflammation, MicroRNA and Senescence Networks in Human Cancer. Curtis C. Harris and Aaron J. Schetter. Laboratory of Human Carcinogenesis, Center for Cancer Research, National Cancer Institute, National Institute of Health, Bethesda, Maryland, USA. Chronic inflammation and infection are major causes of cancer. There are continued improvements to our understanding of the molecular connections between inflammation and cancer. Key mediators of inflammation-induced cancer include NFκB, reactive oxygen and nitrogen species, inflammatory cytokines, prostaglandins and specific microRNAs. The collective activity of these mediators is largely responsible for either a pro-tumorigenic or anti-tumorigenic inflammatory response through changes in cell proliferation, cell death, cellular senescence, DNA mutation rates, DNA methylation, and angiogenesis. As our understanding grows, inflammatory mediators will provide opportunities to develop novel diagnostic and therapeutic strategies. In this presentation, we provide a general overview of the connection between inflammation, microRNAs, senescence, and cancer and highlight how our improved understanding of these connections may provide novel preventive, diagnostic and therapeutic strategies to reduce the health burden of cancer.


March 15-16, 2012

Non-coding RNAs of the 8q24 Locus.

Konrad Huppi1, Oliver L. Ou1, Anthony Barsotti2, Jason J. Pitt1, Natalia Volfovsky3, Robert Stephens3, Siegfried Janz4, Vishala Neppalli4, Brady Wahlberg1, Tamara L. Jones1, Jaime Rodriguez-Canales5, Heidi Erickson6, J. Frederic Mushinski7, Carol Prives2, Michael Emmert-Buck5 and Natasha J. Caplen1. 1Genetics Branch, CCR, NCI, NIH, Bethesda, MD; 2Department of Biological Sciences, Columbia University, New York, NY; 3ABCC, NCI-Frederick, NIH, Fredrick, MD; 4Department of Pathology, University of Iowa, IA; 5 Laboratory of Pathology, CCR, NCI, NIH, Bethesda, MD; 6Department of Thoracic/Head & Neck Medical Oncology, UT MD Anderson Cancer Center, Houston, TX; 7Laboratory of Cancer Genetics and Biology, CCR, NCI, NIH Bethesda, Maryland, USA. The 8q24 locus has been found to be involved in many types of cancers as a consequence of somatic changes associated with chromosome instability including amplification, translocation or deletion or frequent viral integration (HPV). A number of SNPs in Genome Wide Association (GWA) studies have also implicated the 8q24 locus as a region of susceptibility for many types of cancer. The most likely 8q24 candidate target may be the MYC proto-oncogene that is a well-characterized transcription factor. However, the assumed correlation between MYC expression and disease is lacking suggesting a connection between 8q24 involvement and disease is much more complicated than simply targeting MYC. While other transcription units also reside within the 8q24 locus (PRNCR1, POU5F1P1, PVT1 and the miRNA cluster miR-1204~1208), they are remarkable in that no coding potential has been readily associated with any of these genes. Thus, the region has been referred to as the ―8q24 Gene Desert‖. With the renewed realization that many noncoding RNAs do have a functional role, the location of the miR-1204~1208 cluster of miRNAs within the PVT1 lincRNA transcriptional unit actually suggests an ―oasis of transcription‖ that could be the additional or alternative target to MYC. In this presentation we will discuss studies focused on examining the regulation of expression of long and small non-coding RNAs from the PVT1 locus and the potential functional consequence of the deregulated expression of these non-coding RNAs in cancer.


March 15-16, 2012

Causes and Consequesces of MicroRNA Dysregulation in Cancer. Carlo M. Croce. Department of Molecular Virology, Immunology and Medical Genetics, The Ohio State University Medical Center, Columbus, Ohio, USA. Since the discovery of miR-15a and miR-16-1 deletions in CLL15, many laboratories around the world have shown miRNA dysregulation in all tumours studied, including the most common, such as lung, breast, prostate and gastrointestinal cancers. Such dysregulation, like the dysregulation of oncogenes and tumour suppressor genes, can be caused by multiple mechanisms, such as deletion, amplification, mutation, transcriptional dysregulation and epigenetic changes. As miRNAs have multiple targets, their function in tumorigenesis could be due to their regulation of a few specific targets, possibly even one, or many targets. A future challenge will be to identify all of the targets of the miRNAs involved in cancer and establish their contribution to malignant transformation. An additional challenge will be the identification of all of the miRNAs that are dysregulated by pathways that are consistently dysregulated in various types of human cancers. This point is of particular importance, as instead of focusing on specific alterations in protein-coding oncogenes or tumour suppressor genes — which may be difficult to treat — we could focus on their downstream miRNA targets. If these miRNA targets are crucial for the expression of the malignant phenotype and the cancer cells depend on their dysregulation for proliferation and survival, we can expect that the use of miRNAs or anti-miRNAs will result in tumour regression. Genomic analyses for alteration in miRNA genes or for copy number alterations in various human tumours by deep sequencing is in progress but has not been completed. These studies could provide additional information concerning the involvements of miRNAs in cancer and in many other diseases. Over the past few years, we have observed a shift from conventional chemotherapy to targeted therapies, and miRNAs and anti-miRNAs will contribute extensively to the latter.


March 15-16, 2012

Translational Genomics: From 3-D Long-Distance Interactions to the Passenger/Driver Gene Repertoire in Cancer. Barbara Wold. California Institute of Technology, Pasadena, California, USA.


March 15-16, 2012

The Relevance of Patient Stratification in Oncology for Patients with Multiple Sclerosis and Other Neurological Disorders. Tim Harris. Biogen Idec, Cambridge, Massachusetts, USA. The identification of mutations in tumours by DNA sequencing and other methods is now common place in oncology clinical practice for helping to determine the best drug for the patient. For example, mutations in certain kinases can suggest particular inhibitors that the patient can be prescribed or the patient can be directed to a relevant clinical trial where inhibitors for that kinase are being tested. In diseases like multiple sclerosis and other auto-immune diseases there are no tumours to look at in this way. However, gene expression profiling and GWAS are beginning to shed more light on the biology of these autoimmune diseases. It is conceivable that it will be possible to determine subtypes of disease based on these methods so that patients in the future can be put on medicines that are more suitable for their particular disease subtype. In this presentation the challenges for patient stratification using genetics and genomics in autoimmune diseases and cancer will be compared and contrasted.


March 15-16, 2012

Aberrant Expression of Satellite Repeats in Pancreatic Cancer. David T. Ting. Massachusetts General Hospital Cancer Center and Harvard Medical School, Boston Massachusetts, USA. Pancreatic cancer remains one of the most lethal cancers where the vast majority of patients are diagnosed too late and conventional therapies have largely been ineffective, making early detection modalities and novel avenues for drug development greatly needed. RNA sequencing has recently revealed a new side of pancreatic cancer with the discovery of aberrant expression of non-coding RNAs (ncRNAs), which offers new insight into pancreatic cancer biology for biomarker development and identification of unique therapeutic targets. A significant portion of these cancer associated ncRNAs emanate from pericentromeric satellite repeats, areas previously thought to be heavily silenced. Satellite RNAs represent a novel cancer biomarker as well as a previously unappreciated feature of cancer biology that may be involved in the metastatic spread of cancer cells. Advances in microfluidic technologies have provided methods to capture intact circulating tumor cells (CTCs) in the peripheral blood of cancer patients offering a modality to study the impact of satellite ncRNAs on the metastatic process as well as evaluate its utility as a non-invasive biomarker. The combination of a blood based tumor cell assay with a comprehensive characterization of the prevalence and impact of ncRNAs in pancreatic cancer development provides a discovery platform to formulate novel early detection biomarkers and therapeutic strategies against this deadly disease.


March 15-16, 2012

Integrated Genomic Data Informs Cancer Treatment Elaine R. Mardis. Washington University School of Medicine, St. Louis, Missouri, USA. Next-generation sequencing instruments and advanced bioinformatic data analysis approaches have enabled unprecedented exploration of the human genome, especially in the comparison of tumor to normal genomes from the same patient. While there has been an enormous amount of discovery work that has characterized the genomic landscape of both solid and liquid cancers, the translation of these approaches to clinical practice has been minimal. My talk will explore our efforts to pursue the use of cancer genomics on an individual patient basis, to provide therapeutic options for consideration by the patient and the treating oncologist. I will provide several vignettes from our work to-date, and make an argument for the broader application of this approach toward clinical utility.


March 15-16, 2012

Comprehensive Detection of Variation in Thousands of Germ-Line and Tumor Genomes. Stephen E. Lincoln. Complete Genomics, Inc., Mountain View, California, USA. Accurate, complete, genome sequences in significant numbers can provide a powerful resource for identifying variants, both somatic and germline, involved in cancer and many other human diseases. We have developed a technology platform, production laboratory and bioinformatics pipeline tailored to the specific problem of high-depth and high-volume whole human genome re-sequencing. These data have been used in studies of Mendelian diseases, somatic mutations in cancer, population genetics and basic genome biology. Recent algorithmic improvements, in combination with high depth sequencing (often over 100x intumor samples) improve sensitivity for minor alleles in heterogeneous samples, while at the same time reducing false positives. Of course, the availability of such technology leads one to consider the interpretation challenges presented by such data sets. Newer variant calling algorithms correctly present a complicated view of human genomes, including thousands of both small and large complex variants, and this fact can challenge current annotation methods and variantfiltering workflows which to date have mostly focused on simple SNPs. Nevertheless newer approaches to genome comparison and variant annotation allow these data sets to be effectively exploited and mined.


March 15-16, 2012

Comprehensive Analysis of the Cancer Genome: Towards Precision Therapy in Cancer. Javed Khan. Pediatric Oncology Branch, Center for Cancer Research, National Cancer Institute, Bethesda, Maryland, USA. Cancer is a disease of the disordered genome and genomic approaches continue to provide powerful tools to characterize many of the mechanisms involved in the oncogenic process, as well as identify biomarkers and targets for therapy. Previous microarray based studies have identified diagnostic biomarkers in pediatric cancers. I will begin by describing a study in which the discovery of a diagnostic biomarker has led to the identification of a therapeutic target that is being developed into a clinical trial. Rhabdomyosarcoma (RMS) is a childhood cancer originating from skeletal muscle, and patient survival is poor in the presence of metastatic disease. Few determinants that regulate metastasis development have been identified. The high expression of the diagnostic biomarker FGFR4, a receptor tyrosine kinase, in RMS suggested a role in tumorigenesis. Higher FGFR4 expression in RMS tumors was associated with advanced stage cancer and poor survival, while FGFR4 knockdown reduced tumor growth and lung metastases in mice. We identified 6 FGFR4 tyrosine kinase domain mutations which were found among 7 of 94 (7.5%) primary human RMS tumors. The mutants K535 and E550 increased autophosphorylation, STAT3 signaling, tumor proliferation, and metastatic potential. Cell lines expressing the K535 and E550 FGFR4 mutants were substantially more susceptible to apoptosis in the presence of a pharmacologic FGFR inhibitor than the cell control cell lines expressing the empty vector or wild type FGFR4. Together, these results demonstrate that mutationally activated FGFR4 acts as an oncogene and is the first known mutations in a receptor tyrosine in RMS. These findings support the potential therapeutic targeting of FGFR4 in RMS. I will next discuss how we are using a combination of whole genome, exome and transcriptome sequencing to identify other driver genes involved in metastasis and cell growth; which may represent potential targets of therapy. Previous studies have reported that metastatic tumor cells acquire additional mutations to those present in the primary due to an unstable cancer genome. However, it is not clear if all malignancies follow a similar pattern. To identify potential driver mutations and investigate genomic changes that occur during tumor progression, we performed comprehensive genomic analyses including whole genome, exome and transcriptome sequencing of a primary and two metastatic tumors from a patient with neuroblastoma taken at different time points during therapy. We found that despite 3.5 years of multimodal cytotoxic therapy the genomes of all three cancers remained relatively stable using SNP genotyping and array comparative genomic hybridization. Remarkably, there were only 5 expressed somatic non-synonymous mutations and all of them were present in the three tumors with no additional proteindisrupting mutation arising in the metastatic samples. The chromosomal structural stability together with the lack of evidence for progressive accumulation of expressed mutations indicates that this cancer arose from a single catastrophic chromosomal event combined with a small number of somatic mutations. These next generation sequencing techniques are currently being developed for precision therapy and I finally will discuss our current and future plans for leveraging these combinatorial approaches in clinical trials for children with high-risk, metastatic, refractory or recurrent cancers. 16 Translational Genomics

March 15-16, 2012

Human Cancer Epigenetics and Beyond. Manel Esteller. Bellvitge Biomedical Research Institute (IDIBELL), Barcelona, Spain. For the last twenty-five years an increasing amount of evidence has shown the relevance of epigenetics in cell biology and tissue physiology, being DNA methylation aberrations in cancer the flag-ship for the recognition of its disturbance in human diseases. From the candidate gene approaches, new powerful technologies such as comprehensive DNA methylation microarrays and whole genome bisulfite sequencing has recently emerged that have reinforced the notion of epigenetic disruption in the crossroad of many sickness. From the poster-boy cases of MGMT and GSTP1 hypermethylation in the prediction of alkylating drug response and prostate cancer detection, respectively, to the personalized treatment of leukemia with small molecules targeted to fusion proteins involving histone modifiers such as DOT1L and MLL, the field has walked a long path. The current talk will focus in the epigenetic profiling, basically at the level of DNA methylation and histone modifications that is starting to provide clinical value in the diagnosis, prognosis and prediction of response to drug therapies, with an emphasis in neoplasia, but without forgetting the novel advances in other human disorders. For cancer, we have already a wide view of the undergoing DNA methylation events that expand beyond classical promoter CpG islands of tumor suppressor genes and we have a growing list of mutated chromatin remodeler genes that contributes to the tumorigenesis process. It is time to apply this knowledge in practical clinical situations like the diagnosis of cancers of unknown primary, the screening of malignancies in high-risk populations or a biomarker selection of the patients that should receive treatment with epigenetic drugs.


March 15-16, 2012

The Cancer Epigenome. Peter Jones. University of Southern California, Los Angeles, California, USA.


March 15-16, 2012

Integrated DNA Methylation and Chromatin Maps to Identify Epigenetically Silenced Cancer Drivers. Jean-Pierre Issa. Temple University School of Medicine, Philadelphia, Pennsylvania, USA.


March 15-16, 2012

Bringing the New Sequencing Technologies to the Oncology Clinic: Opportunities and Obstacles. Paul Meltzer, J. Keith Killian, Christopher Lau, Ogan Abaan, Marbin Pineda, Robert L. Walker, Jack Zhu, Sean Davis, and Sven Bilke. Genetics Branch, Center for Cancer Research, National Cancer Institute, Bethesda, Maryland, USA. Because the intrinsic biology of each patient’s tumor plays a major role in governing the response to therapy, biomarkers that reveal key aspects of that biology have the potential to guide clinical decision making. The tumor genome is unique among all the possible sources of biomarkers. DNA structural and sequence aberrations provide a genomic fingerprint unique to each tumor recording the deviation of the tumor genome from its normal counterpart. These aberrations can reveal the key genes and pathways driving tumor growth. Additionally, the stability of DNA means that it can be obtained from virtually any clinical sample making it an ideal analyte in the clinical laboratory. However, there are significant difficulties in bringing these concepts to clinical reality. First and foremost, cancer genomics is still in a discovery phase, and for many tumors the mutational spectrum is still unknown. Next, the available data suggests that in many common cancers, the pattern of mutations is exceedingly complex with few highly frequent driver mutations. There are also large numbers of passenger mutations in chaotic tumor genomes that have little clinical significance. These issues add substantially to the complexity of predicting the clinical biology of an individual tumor. Counterbalancing these challenges, dramatic technological advances in the characterization of tumor genomes are bringing large-scale sequencing closer to clinical reality. The development of large cancer genomics databases is beginning to provide the necessary backdrop for the ongoing and future clinical studies needed to bring the promise genomics to fruition. Pathways for bringing cancer genome analysis into the clinic will be discussed.


March 15-16, 2012

Deciphering Early Onset Breast Cancer in Women Of African Ancestry via Integrated "-omics." Olufunmilayo I. Olopade. Center for Clinical Cancer Genetics and Global Health, Department of Medicine, The University of Chicago, Chicago, Illinois, USA, African populations show significantly higher levels of genetic diversity than other populations as reflected in over 2,000 distinct ethnic groups and languages in Africa. Indeed, there is greater genetic diversity within and between different African populations than between African and Eurasian populations. During the Atlantic slave trade from the 15th through the 19th centuries, over 45 ethnic groups were brought to the Americas from African regions ranging from Senegambia to west Central Africa, and even some groups from south-east Africa. Additionally, modern descendants from these groups carry genetic admixtures of roughly 20% non-African origin. Genome-wide association studies (GWAS) have revealed several genetic loci that confer risk of breast cancer but none of the loci could be replicated in women of African ancestry. We are conducting a genome-wide association study (GWAS) of breast cancer in 1,715 cases and 2,162 controls of African ancestry from Nigeria, Barbados and the United States, using Illumina Hum-Omni2.5M platform. Subset analysis to classify genetic variants for Triple Negative Breast Cancer (TNBC) will be conducted. In addition, whole genome sequencing study will examine the rare genetic variants that confer excess risk of TNBC in women of African ancestry. Our goal is to answer two overarching, related research questions: 1) why are women of West African ancestry more likely to develop TNBC? 2) What are the complex roles of genetic and non-genetic factors in breast cancer etiology and progression? We propose a comprehensive approach that will define the genes and major pathways dysregulated in TNBC that disproportionately affects young women and women of African ancestry. Using high throughput sequencing, and computational biology coupled with rigorous population research methodology, we believe Integrated "-omics" will allow us to achieve major advances in the understanding of the mechanisms of TNBC, the most aggressive and most lethal of all breast cancers! In the long-term, we hope to formulate strategies to eradicate TNBC as a cause of death for high-risk women all over the world.


March 15-16, 2012

The Pancreatic Cancer Transcriptome and Epigenome. Laufey Amundadottir. Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, National Cancer Institute, Bethesda, Maryland, USA. Pancreatic cancer is the tenth most commonly diagnosed cancer in both men and women in the U.S., but the fourth leading cause of cancer mortality. It is among the deadliest of all cancers with a 5 year overall survival of less than 5% and a median survival of 3-6 months. Conventional chemotherapy and radiation therapy have limited effects on overall survival. To enhance our understanding of the cellular pathways that are dysfunctional in pancreatic cancer, we have catalogued and compared transcribed sequences and DNA methylation patterns in tissues and cell lines derived from normal pancreatic tissues and pancreatic adenocarcinoma by next generation RNA-sequencing and methylated DNA immunoprecipitation (MeDIP) chip analysis. In this presentation I will detail our approach and results that highlighted genes, pathways and regulatory sub-networks implicated in cell proliferation, stress response, pancreatitis and pancreatic development as putatively important for the biology of pancreatic cancer.


March 15-16, 2012

Poster Abstracts


March 15-16, 2012

Poster List Speaker Number 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15. 16. 17. 18. 19. 20. 21. 22. 23. 24. 25. 26. 27. 28. 29. 30. 31. 32. 33. 34. 35. 36. 37.

Page Number

Abaan, Ogan………………………………………………… Abdelmonsif, Doaa………………………………………….. Andersen, Jesper…………………………………………… Bassett, Douglas……………………………………………. Bradley, Amanda …………………………………………… Camps, Jordi………………………………………………… Choudhury, Dipa Roy………………………………………. Duncan, Beverly…………………………………………….. Geiss, Gary………………………………………………….. Gao, Guimin…………………………………………………. Harris, Lyndsay……………………………………………… Hostetter, Galen…………………………………………….. Jeong, Kyeong Soo………………………………………… Killian, Keith……………………………………………….... Knudson, C. Michael……………………………………...... Kofman, Alexander…………………………………………. Lin, Eric………………………………………………………. Mittereder, Lara……………………………………………… O’Neill, Ray………………………………………………….. Padia, Janak………………………………………………… Reilly, Karlyne……………………………………………….. Reinhold, William……………………………………………. Riz, Irene…………………………………………………….. Robles, Ana…………………………………………………. Ruan, Jianhua………………………………………………. Ryan, Brid…………………………………………………… Sacchi, Nicoletta……………………………………………. Shiao, Yih-Horng…………………………………………… Sikdar, Nilabja………………………………………………. Simmons, John……………………………………………… Soldin, Offie………………………………………………….. Subramanian, Subbaya…………………………………….. Sun, Lei………………………………………………………. Thomson, Naomi……………………………………………. Tran, Bao…………………………………………………….. Varma, Sudhir……………………………………………….. Wakefield, Lalage……………………………………………

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March 15-16, 2012

38. 39. 40. 41. 42. 43.

Wang, Josh…………………………………………………... Xi, Sichuan…………………………………………………… Ye, Jian………………………………………………………. Yu, Guoqin…………………………………………………… Zeeberg, Barry………………………………………………. Zhang, Shuling……………………………………………….

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March 15-16, 2012

Poster 1 Pharmacogenomic Integration of Exome Sequence with Drug Sensitivity in the NCI60 Cell Lines. Ogan D. Abaan1, Sean R. Davis1, Sven Bilke1, Eric C. Polley3, Robert L. Walker1, Marbin Pineda1, Yuelin J. Zhu1, William Reinhold2, Susan L. Holbeck3, Richard M. Simon3, James H. Doroshow3, Yves Pommier2 and Paul S. Meltzer1. 1Genetics Branch, CCR, NCI; 2 Laboratory of Molecular Pharmacology, CCR, NCI; 3Division of Cancer Treatment and Diagnosis, NCI.

Acquired and constitutional genetic variations in cancers have a substantial influence on response to therapy. The NCI60 panel of cell lines provides a unique opportunity to investigate this problem 2006. These 60 human tumor cell lines represent 9 cancer types, and are richly characterized as the result of many previous studies, which have collected various biochemical and genomics data. In addition, there exists a comprehensive dataset of sensitivity and resistance profiles of over 100,000 compounds with potential antineoplastic activity. Here we sequenced the whole exomes of the NCI60 cell lines and identified single nucleotide variants and short insertions/deletions. Variants found by sequencing were in excellent agreement with variants previously identified by SNP analysis or targeted capillary sequencing. Observation from ti/tv ratio and nature of base changes revealed preservation of possible carcinogen specific DNA mutation signatures, especially for melanoma and lung cancer cell lines. Deleterious variants were identified at higher frequency in known cancer genes while TP53 mutations where the most frequent event among the NCI60. Finally, notable instances of correlation between gene/pathway mutations with sensitivity to certain drug/compounds were identified. For example, sensitivity to vemurafenib and selumetinib was highly correlated with BRAF mutations, and nutlin-3 to lack of TP53 mutations. Our analysis identifies a large number of genomic variants correlated with drug responsiveness, including a number of genes known to be targets of specific agents as well as numerous candidates for future investigation.


March 15-16, 2012

Poster 2 Molecular Predictors of the Outcome for Anthracycline - Based Adjuvant Chemotherapy in Egyptian High Risk Female Breast Cancer Patients. Ehsan Mohamad Hassan Abd Al-Rahman1, Eman Mohamed Soliman Kamha1, Nashaat Saad Lotfy2 and Doaa Ali Abd Al-Monsef1. 1Medical Biochemistry Department, Clinical Oncology and 2Nuclear Medicine, Faculty of Medicine, University of Alexandria, Egypt.

Anthracyclines represent one of the most important chemotherapeutics in breast cancer. However, they cause cardiac damage. Besides, some tumors might be anthracyclineresistant. The aim of the present work was to study the predictive value of estrogen receptors (ER) and progesterone receptors (PR) proteins. Furthermore, the predictive value of topoisomerase IIa (TOPOIIa) gene aberrations (amplification or deletion) and breast cancer 1, early onset (BRCA1) gene methylation for the outcome of 5-fluorouracil / Adriamycin / cyclophosphamide (FAC) adjuvant chemotherapy in Egyptian high risk female breast cancer patients. The present retrospective cohort study was conducted in Alexandria Main University Hospital, Egypt. It included fifty high risk female breast cancer patients (according to St Gallen guidelines 2007) with operable breast cancer. All of them have received FAC adjuvant chemotherapy between January 2007 and December 2007 and were followed for 2 years. Pretreatment breast tumor samples were obtained from formalin fixed/paraffin-embedded tissue blocks. Log rank survival analysis showed that TOPOIIa gene aberrations, methylated BRCA1 gene, negative ER protein and negative ER/PR protein states were associated with significantly superior disease free survival (DFS) rates after FAC therapy. Cox regression analysis showed that ER protein and BRCA1 gene methylation states might be independent predictors for the outcome of FAC adjuvant chemotherapy while TOPOIIa gene state mightn't. However, if ER protein and BRCA1 gene methylation states can be used in tailoring chemotherapy or not, further studies have to be done on a bigger number of cases with longer follow-up period. Additionally, large-scale prospective studies will be needed to clearly define TOPOIIa gene and PR protein predictive values. Patients having BRCA1 gene methylation might be at risk of having distant metastasis. So, if proved by large-scale studies, such patients could be recommended for intensified follow-up and treatment. We thank our clinical and laboratory colleagues who have contributed to the current research. Special thanks to Prof. Dr. Osama Essa, Professor of Community Medicine, Faculty of Medicine, University of Alexandria, Egypt, for helpful discussions and statistical analysis. We wish to thank Dr. Nadia Abaas Lecturer of Pathology, Faculty of Medicine, University of Alexandria, for her assistance in IHC work.


March 15-16, 2012

Poster 3 Translational Genomics Analyses of Cholangiocarcinoma Identify Patients Who May Respond to Tyrosine Kinase Inhibitors. Jesper B Andersen, Matthew Gillen, Elizabeth A Conner, Valentina M Factor and Snorri S Thorgeirsson. Laboratory of Experimental Carcinogenesis, CCR, NCI. Background & Aims: We recently developed a characterization strategy using 104 resected cholangiocarcinomas (CCAs). Patients were classified into distinct subclasses, based on survival, recurrence, and KRAS mutations. Patient group III with the worst prognosis was characterized by genes associated with proteasomal activity and inflammation. Microdissection of tumor compartments and immunostaining showed deregulation of HER2, EGFR and MET suggesting improved therapeutic response to TKIs in combination with anti-inflammatory drugs. Methods: We integrative data from our patient cohort and 7 human CCA cell lines to investigate optimal treatment options for CCA. CCA lines were exposed to trastuzumab, lapatinib, and celecoxib. Targets were validated by transcriptomics, Western and immunostaining. The efficacy of co-administering celecoxib and trastuzumab was examined in vitro and in TKI-resistant xenografts (30mg/kg celecoxib gavage bid, 0.45mg/dose trastuzumab i.p. twice weekly). Results: To assess receptor tyrosine kinases as potential drug-targets, we first integrated the CCA lines with the cohort to establish concordance between cell lines and tumor subclasses, then exposed them to trastuzumab and lapatinib for 7 days. Lapatinib was more effective showing 50-80% growth inhibition compared to 15-20% inhibition achieved by trastuzumab. Classification was confirmed by transcriptomics separating the lines based on drug-response. TKI-resistance was observed in 3/7 lines. A combination of celecoxib and trastuzumab was effective in inhibiting growth of TKI-resistant lines in vitro (P < 0.0001). In the xenograft model celecoxib and trastuzumab significantly inhibited tumor growth (P < 0.004) compared to control and mono-therapies. Conclusion: Our study has provided new insights into both pathogenesis and optimal treatment options for CCA. We identified a subgroup of patients with poor outcome who may benefit from TKIs. TKI-resistance was effectively treated both in vitro and in vivo by co-administration with celecoxib. Our data demonstrates that targeting AKT downstream of KRAS sensitizes TKI-resistant CCA, and provides a new treatment option.


March 15-16, 2012

Poster 4 Biological Interpretation of NGS Re-sequencing Studies Leveraging Context-Rich Biomedical Content.

D Richards1, R Flannery1, A Krämer1, A Kutchma1, J Lerman1, J Leschly1, S Majumdar1, N Marshall1, M Molloy1, A Muthiah1, A Ning1, R O'Connor1, K Patel1, V Rajaraman1, R Rebres1, S Sanga1, A Sarver1, H Su1, W Zhou1, X Zhu1 and D Bassett1. 1Ingenuity Systems, Redwood City, California. Biological interpretation of thousands of potentially deleterious variants is a bottleneck in extracting valuable causal insights from DNA re-sequencing studies, often requiring months of effort after completion of the reference genome alignment and variant calling steps. We have developed a fast, easy-to-use application, Ingenuity® Variant Analysis (www.ingenuity.com), that leverages an extensive knowledge base of millions of expertcurated mutation and biomedical findings from the literature to empower real-time interactive filtering and rapid prioritization of variants. It enables clinical researchers to quickly zero in on the few variants that are most compelling for follow-up. Using a combination of causal analytics, genetic analysis at the variant, gene, and pathway levels, and the ability to visualize how variants impact disease progression, we will demonstrate the application of a context-rich knowledge base to discover cancer driver variants and novel causal variants for human genetic disease.


March 15-16, 2012

Poster 5 Clonal Patterns of Chromosomal Aberrations in Cervical Cancers and Precursor lesions.

Amanda Bradley1, Kerstin Heselmeyer-Haddad1, Timo Gaiser2, Woei-Jyh Lee3, Alejandro Schaffer3, Sonia Andersson4 and Thomas Ried1. 1Genetics Branch, CCR, NCI, NIH, Bethesda, Maryland; 2University Hospital Mannheim, Germany; 3NCBI, NIH, Bethesda, Maryland; 4Karolinska Institute, Karolinska University Hospital-Solna, Stockholm, Sweden. We developed a nine-probe FISH assay to investigate the dynamics of clonal evolution of cervical precancerous and cancerous lesions. Five oncogenes, COX2 on 1q, TERC on 3q, TERT on 5p, MYC on 8q, and ZNF217 on 20q, and three tumor suppressor genes, ING5 on 2p, FHIT on 3p and CHEK1 on 11q, and a centromere 7 probe as a ploidy control were hybridized subsequently in three-probe panels on cytospins produced from histologically defined areas microdissected from 50 µm sections. Archival specimens were obtained from the Karolinska University Hospital, Sweden, and the University Hospital Mannheim, Germany. Objectives: • Determine gain and loss patterns of the targeted genes within cervical intraepithelial neoplasia (CIN) and invasive cervical carcinomas • Characterize clonal patterns with the goal to understand clonal evolution during cervical carcinogenesis We plan to analyze 20 cancers, 20 CIN3 lesions and 20 lower-grade CIN lesions. We have completed the analysis of two cancer specimens and three severely dysplastic lesions (CIN3). All five lesions showed distinct clonal cell populations characterized by specific signal patterns with a TERC gain (3q) being the only aberration present in all of the clones. Both cancers and one CIN3 lesion consisted mainly of diploid cell populations in which the TERC gain was accompanied by a loss of FHIT (3p) suggestive of an isochromosome 3q as the major chromosomal aberration event. The other two CIN3 lesions consisted mainly of tetraploid cell populations with a TERC gain but no concomitant FHIT loss. In three of the lesions only two or three changes were observed: 1) FHIT loss and TERC gain, 2) FHIT loss, TERC gain and CHEK1 loss, and 3) TERC gain and COX2 gain. The other two lesions showed clonal changes involving five to seven markers. Our goal is to improve our understanding of genome dynamics during cervical carcinogenesis.


March 15-16, 2012

Poster 6 Genome-wide Molecular and Functional Analysis Identified LNX2 as a Novel Gene Involved in Colorectal Carcinogenesis.

Jordi Camps1, Jason J. Pitt2, Georg Emons1,3, Amanda B. Hummon1, Chanelle M. Case1, Marian Grade1,3, Tamara L. Jones2, Quang T. Nguyen1, B. Michael Ghadimi3, Tim Beissbarth3, Michael J. Difilippantonio1, Natasha J. Caplen2 and Thomas Ried1. 1Cancer Genomics Section; 2Gene Silencing Section, Genetics Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD, USA; 3 Department of General and Visceral Surgery, University Medicine Göttingen, Göttingen, Germany. Colorectal cancer (CRC) is one of the most frequent malignancies in many parts of the world and a leading cause of cancer deaths in both men and women. The identification of rationale therapeutic targets is one possibility to provide personalized medicine to cancer patients. Our approach consisted of identifying overexpressed genes located at sites of recurrent chromosomal amplifications, as these regions are likely to harbor genes required for cancer cell survival. Thirty-one colon cancers, 25 rectal cancers, and 15 CRC cell lines were analyzed by high-resolution array CGH and microarray gene expression profiling. RNA interference (RNAi)-based analysis identified a subset of genes whose loss-offunction (LOF) reduced the cellular viability of CRC cell lines. Consistent with previous reports, the vast majority of CRC assayed exhibited amplification of the chromosome band 13q12.13-q12.3. Among the genes residing within the 13q12.13-q12.3 amplified region, we focused on those showing an overexpression level of at least two-fold higher in the tumor compared to normal mucosa. Of this subset, we identified NUPL1, LNX2, POLR1D, CDX2, POMP, and SLC7A1, for which cell survival was impaired after gene silencing. As little is known about the function of these proteins, we decided to use an unbiased systems biology approach to identify genes, pathways and networks altered following RNAimediated LOF of each of these candidate genes. To do this, we perturbed the expression of each candidate gene through application of two or more siRNAs corresponding to each gene, followed by whole genome expression profiling to monitor cellular transcriptional responses for each gene specific LOF. One candidate, LNX2, ligand of numb-protein X 2, encodes an intracellular scaffolding protein involved in regulating notch signaling; silencing LNX2 resulted in reduced NOTCH1 levels and NOTCH signaling activity, in downregulation of the transcription factor TCF7L2, and in markedly reduced WNTsignaling. Further analysis of the whole genome deregulated effectors showed a statistical significant enrichment of TCF7L2 targets (P