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PDF - 1109K, Blood and lymphatic vessel density in experimental BL2B95 tumors treated with IB in the CAM model.

Burkitt lymphoma is a rare malignancy arising from B cells. Current chemotherapeutic regimens achieve excellent overall survival rates in children, but less impressive rates in adults. There are cases with poor outcome caused by toxic effects of the therapy, tumor lysis syndrome, or metastatic spread of lymphomas to the central nervous system. Modulators of reactive oxygen species are currently discussed as potential drugs for the treatment of cancer. The NADPH oxidase 4 inhibitor imipramine-blue might satisfy the aforementioned requirements, and was studied here. We used MTT assay, crystal violet assay, and thymidine 3H-incorporation assay to analyze the effects of imipramine-blue on Burkitt lymphoma (BL2, BL2B95, BL30B95, BL41B95), neuroblastoma (KELLY, SH-SY5Y, SMS-KAN), cervix carcinoma (HeLa), breast cancer (MDA-MB231), angiosarcoma (AS-M), human embryonic kidney (HEK293WT), and nonmalignant (FLP1) cell lines. The effects of imipramine-blue on BL2B95 cells in vivo were investigated in xenografts on the chick chorioallantoic membrane (CAM). We report that imipramine-blue is a potent growth inhibitor for several cancer cell lines in vitro with IC50 values comparable to those of doxorubicin (0.16–7.7 μmol/L). Tumor size of BL2B95 cells inoculated in the CAM was reduced significantly (P < 0.05) after treatment with 10 μmol/L imipramine-blue. Lymphogenic dissemination of BL2B95 and the formation of blood and lymphatic vessels in experimental tumors were not affected. We show that imipramine-blue can be used to decrease the viability of cancer cell lines in vitro and in vivo. Imipramine-blue reduces the size of experimental Burkitt lymphoma significantly but does not affect the dissemination of BL2B95 cells, angiogenesis, and lymphangiogenesis. Mol Cancer Ther; 13(4); 833–41. ©2014 AACR.

PDF file, 109KB, S1: Genotype and allele frequency of the analyzed IL-10, IL-4R, TNFRI and II gene variations in patients with T-NHL (n = 117); S2: 2. 3-year survival rates for OS and EFS of patients suffering from T-NHL in relation to analyzed gene variation; S3: Real time PCR Taqman� assays used to analyze IL-10, IL-4R, TNFRI and TNFRII gene variations; S4: Primer for SNaPshot Assay used to analyze IL-10 gene variations.

Purpose: Peripheral T-cell non–Hodgkin lymphomas (T-NHL) represent a small but heterogeneous and clinically aggressive subset of NHLs with a poor outcome. Cytokines or their receptors might be associated with the clinical outcome of these lymphomas. Therefore, we tested whether gene variations and serum levels of soluble TNF receptor (TNFR)I (sTNFRI), sTNFRII, interleukin (IL)-10, or sIL-4R are predictive for treatment response in T-NHLs.Experimental Design: Peripheral blood DNA from 117 patients with T-NHL treated in prospective clinical trials was subjected to genotyping analysis. Whenever possible, pretreatment sera were obtained, and circulating levels of sTNFRI, sTNFRII, IL-10, and sIL-4R were determined with a specific capture enzyme-linked immunoassay.Results: Patients characterized by TNFRI-609GG (rs4149570) showed a trend toward better event free survival [EFS; univariate: P = 0.041; multivariate: HR, 1.76; confidence interval (CI), 0.99–3.14 with P = 0.056]. A protective role of IL-10–1087A, −824T, and −597A reported in another study was not confirmed in our cohort. Patients with circulating levels of soluble TNFRII ≥2.16 ng/mL had a 2.07-fold increased relative risk for shorter overall survival (OS; univariate: P = 0.0034; multivariate: HR, 2.07; CI, 0.92–4.70 with P = 0.081) and a 2.49-fold higher risk for shorter EFS (univariate: P = 0.00068; multivariate: HR, 2.49; CI, 1.22–5.08 with P = 0.012). Elevations of circulating levels of sTNFRI, IL-10, and sIL-4R are frequent, but the clinical response in these patients is not significantly different.Conclusions: Our findings suggest a critical role for TNF-TNFR signaling for the clinical outcome of patients with peripheral T-NHLs. Clin Cancer Res; 18(13); 3637–47. ©2012 AACR.

PDF file, 139KB, Overall and Event free survival of patients suffering from T-cell Non-Hodgkin lymphoma in relation to circulating serum levels of sTNFRI, IL-10 and sIL-4R. The survival curves (Kaplan-Meier plots) show a comparison of the OS (A,C,E) and EFS (B,D,F) of T-NHL patients with circulating serum levels of sTNFRI (A,B), IL-10 (C,D) and sIL-4R (E,F). The respective threshold used to define high and low producers is indicated in the figure legend (lower left). For high producers of sTNFRI (B) and sIL-4R (F) an insignificant trend for shorter EFS was observed (p=0.0886 and p=0.333). The other survival curves show no trends for associations of serum levels and survival periods. P refers to the log-rank test. Patients at risk represents the number of patients which still can develop an event (OS or EFS) at defined time points.

Supplementary Methods and Materials, Tables 1-3 from A Functional Polymorphism in the NAD(P)H Oxidase Subunit CYBA Is Related to Gene Expression, Enzyme Activity, and Outcome in Non–Hodgkin Lymphoma

NAD(P)H oxidase is a major endogenous source of reactive oxygen species (ROS). ROS may not only be involved in carcinogenesis but also in efficacy of chemotherapeutic agents like doxorubicin. By a comprehensive genotyping approach covering 48 genetic polymorphisms (single-nucleotide polymorphisms) in five subunits of phagocytic NAD(P)H oxidase, we asked whether they affect gene expression, enzymatic activity, and outcome of CHO(E)P chemotherapy. A highly consistent effect was observed for the CYBA 640A>G variant. In peripheral blood granulocytes of 125 healthy volunteers, the G allele of 640A>G was associated with lower NAD(P)H oxidase activity (P = 0.006). Moreover, the G allele was associated with lower mRNA and protein expression (both P = 0.02). Of clinical importance, the outcome of patients suffering from non-Hodgkin lymphoma and treated with CHO(E)P regimen was dependent on the CYBA 640A>G polymorphism. In an exploratory study (n = 401), carriers of 640GG had an event-free survival (EFS) risk ratio of 1.95 [95% confidence interval (95% CI), 1.31–2.90; P = 0.001] compared with 640AA. In a confirmatory set (n = 477), the risk ratios were 1.53 (1.04–2.25, P = 0.03). The complete set of 878 patients showed a relative risk of 1.72 (1.30–2.26) and 1.59 (1.14–2.21) for EFS and overall survival, respectively. Further molecular-biological experiments showed lower expression and reduced stability of transcripts with the G allele in lymphoblastoid cell lines. Transfection of allele-specific plasmids into HEK293 cells elicited lower activity for the G allele in a luciferase reporter gene construct. Thus, CYBA 640A>G was shown to be a functional polymorphism with possible consequences for patients receiving CHO(E)P chemotherapy and might have further implications for other ROS-mediated modalities. Cancer Res; 70(6); 2328–38

B cells have the unique property to somatically alter their immunoglobulin (IG) genes by V(D)J recombination, somatic hypermutation (SHM) and class-switch recombination (CSR). Aberrant targeting of these mechanisms is implicated in lymphomagenesis, but the mutational processes are poorly understood. By performing whole genome and transcriptome sequencing of 181 germinal center derived B-cell lymphomas (gcBCL) we identified distinct mutational signatures linked to SHM and CSR. We show that not only SHM, but presumably also CSR causes off-target mutations in non-IG genes. Kataegis clusters with high mutational density mainly affected early replicating regions and were enriched for SHM- and CSR-mediated off-target mutations. Moreover, they often co-occurred in loci physically interacting in the nucleus, suggesting that mutation hotspots promote increased mutation targeting of spatially co-localized loci (termed hypermutation by proxy). Only around 1% of somatic small variants were in protein coding sequences, but in about half of the driver genes, a contribution of B-cell specific mutational processes to their mutations was found. The B-cell-specific mutational processes contribute to both lymphoma initiation and intratumoral heterogeneity. Overall, we demonstrate that mutational processes involved in the development of gcBCL are more complex than previously appreciated, and that B cell-specific mutational processes contribute via diverse mechanisms to lymphomagenesis. This study has been supported by the German Ministry of Science and Education (BMBF) in the framework of the ICGC MMML-Seq project (01KU1002A-J) the MMML-MYC-SYS project (036166B) and the project ICGC DE-MINING (01KU1505E), the European Union in the framework of the BLUEPRINT Project (HEALTH-F5-2011-282510) and the KinderKrebsInitiative Buchholz/Holm-Seppensen. This work was supported by the BMBF-funded Heidelberg Center for Human Bioinformatics (HD-HuB) within the German Network for Bioinformatics Infrastructure (de.NBI) (#031A537A, #031A537C). Former grant support of MMML by the Deutsche Krebshilfe (2003–2011) is gratefully acknowledged. We acknowledge COSMIC and use of Cancer Gene Census. Part of the work was performed in association with SFB1074 (particularly subproject B1) funded by DFG. We wish to thank Barbara Hutter, Ivo Buchhalter, Zuguang Gu, and Natalie Jäger for skillful technical assistance. We thank the High-Throughput Sequencing Unit of the Genome and Proteome Core Facility and the Omics IT and Data Management Core Facility of the German Cancer Research Center (DKFZ, Heidelberg) as well as the Institute of Clinical Molecular Biology (IKMB, Christian-Albrechts-University Kiel) for excellent technical support and expertise. DH is a member of the Hartmut Hoffmann-Berling International Graduate School of Molecular and Cellular Biology (HBIGS) and of the MD/PhD-program of the University of Heidelberg. KK and UHT are funded by the Helmholtz International Graduate School for Cancer Research at the German Cancer Research Center. SHB, HK, and SH acknowledge support by LIFE (Leipzig Research Center for Civilization Diseases), Leipzig University. LIFE is funded by the European Union, the European Regional Development Fund (ERDF), the European Social Fund (ESF), and the Free State of Saxony. This work has been carried out with the help of the Interdisciplinary Bank of Biomaterials and Data of the University Hospital of Würzburg and the Julius Maximilian University of Würzburg (idbw). Peer Reviewed "Article signat per 70 autors/es:Daniel Hübschmann, Kortine Kleinheinz, Rabea Wagener, Stephan H. Bernhart, Cristina López, Umut H. Toprak, Stephanie Sungalee, Naveed Ishaque, Helene Kretzmer, Markus Kreuz, Sebastian M. Waszak, Nagarajan Paramasivam, Ole Ammerpohl, Sietse M. Aukema, Renée Beekman, Anke K. Bergmann, Matthias Bieg, Hans Binder, Arndt Borkhardt, Christoph Borst, Benedikt Brors, Philipp Bruns, Enrique Carrillo de Santa Pau, Alexander Claviez, Gero Doose, Andrea Haake, Dennis Karsch, Siegfried Haas, Martin-Leo Hansmann, Jessica I. Hoell, Volker Hovestadt, Bingding Huang, Michael Hummel, Christina Jäger-Schmidt, Jules N. A. Kerssemakers, Jan O. Korbel, Dieter Kube, Chris Lawerenz, Dido Lenze, Joost H. A. Martens, German Ott, Bernhard Radlwimmer, Eva Reisinger, Julia Richter, Daniel Rico, Philip Rosenstiel, Andreas Rosenwald, Markus Schillhabel, Stephan Stilgenbauer, Peter F. Stadler, José I. Martín-Subero, Monika Szczepanowski, Gregor Warsow, Marc A. Weniger, Marc Zapatka, Alfonso Valencia, Hendrik G. Stunnenberg, Peter Lichter, Peter Möller, Markus Loeffler, Roland Eils, Wolfram Klapper, Steve Hoffmann, Lorenz Trümper, ICGC MMML-Seq consortium, ICGC DE-Mining consortium, BLUEPRINT consortium, Ralf Küppers, Matthias Schlesner & Reiner Siebert"

Genome-wide association studies have led in the past to the discovery of susceptibility genes for many diseases including cancer and inflammatory conditions. However, a number of these studies did not realise their full potential. A first critical step in developing such large-scale studies is the choice of genotyping array with respect to the study goal. Coverage is the central criterion for array evaluation. We distinguish between estimates of global coverage across the genome, coverage for each chromosome, coverage for selected pathways and the coverage for genes of interest. Here, we focus on inflammatory and immunological pathways and genes relevant for haematopoietic stem cell transplantation. We compared three arrays: the Infinium Global Screening Array-24 v1.0, the Infinium OncoArray-500 K BeadChip and the Infinium PsychArray-24 v1.2 BeadChip. We employed the European population from the 1000 Genomes Project as reference genome. Global coverage was found to range between 12.2 and 14.2% whereas coverage for a selected pathway ranged from 6.2 to 13.2% and gene coverage ranged from 0 to 54.1%. The Global Screening Array outperformed both other arrays in terms of global coverage, for most chromosomes, most considered pathways and most genes. When selecting suitable arrays for a new study, the coverage of pathways or genes of interest should be considered in addition to global coverage. Local coverage should be regarded when discussing association findings inconsistent across studies and can be useful in data analysis and decision making for additional genotyping.

Burkitt lymphoma is a mature aggressive B-cell lymphoma derived from germinal center B cells(1). Its cytogenetic hallmark is the Burkitt translocation t(8;14)(q24;q32) and its variants, which juxtapose the MYC oncogene with one of the three immunoglobulin loci(2). Consequently, MYC is deregulated, resulting in massive perturbation of gene expression(3). Nevertheless, MYC deregulation alone seems not to be sufficient to drive Burkitt lymphomagenesis. By whole-genome, whole-exome and transcriptome sequencing of four prototypical Burkitt lymphomas with immunoglobulin gene (IG)-MYC translocation, we identified seven recurrently mutated genes. One of these genes, ID3, mapped to a region of focal homozygous loss in Burkitt lymphoma(4). In an extended cohort, 36 of 53 molecularly defined Burkitt lymphomas (68%) carried potentially damaging mutations of ID3. These were strongly enriched at somatic hypermutation motifs. Only 6 of 47 other B-cell lymphomas with the IG-MYC translocation (13%) carried ID3 mutations. These findings suggest that cooperation between ID3 inactivation and IG-MYC translocation is a hallmark of Burkitt lymphomagenesis.

MC3 Working Group: Rehan Akbani21, Pavana Anur22, Matthew H. Bailey1,2,3, Alex Buchanan9, Kami Chiotti9, Kyle Covington12,23, Allison Creason9, Li Ding1,2,3,20, Kyle Ellrott9, Yu Fan21, Steven Foltz1,2, Gad Getz8,14,15,16, Walker Hale12, David Haussler24,25, Julian M. Hess8,26, Carolyn M. Hutter27, Cyriac Kandoth28, Katayoon Kasaian29,30, Melpomeni Kasapi27, Dave Larson1 , Ignaty Leshchiner8, John Letaw31, Singer Ma32, Michael D. McLellan1,3,20, Yifei Men32, Gordon B. Mills33,34, Beifang Niu35, Myron Peto22, Amie Radenbaugh24, Sheila M. Reynolds36, Gordon Saksena8, Heidi Sofia27, Chip Stewart8, Adam J. Struck31, Joshua M. Stuart24,37, Wenyi Wang21, John N. Weinstein38, David A. Wheeler12,13, Christopher K. Wong24,39, Liu Xi12 & Kai Ye40,41 21Department of Bioinformatics and Computational Biology, The University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA. 22Molecular and Medical Genetics, OHSU Knight Cancer Institute, Oregon Health and Science University, Portland, OR 97239, USA. 23Castle Biosciences Inc, Friendswood, TX 77546, USA. 24UC Santa Cruz Genomics Institute, University of California Santa Cruz, Santa Cruz, CA 95064, USA. 25Howard Hughes Medical Institute, University of California Santa Cruz, Santa Cruz, CA 95064, USA. 26Massachusetts General Hospital Center for Cancer Research, Charlestown, MA 02114, USA. 27National Human Genome Research Institute, National Institutes of Health, Bethesda, MD 20894, USA. 28Marie-Josée and Henry R. Kravis Center for Molecular Oncology, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. 29Ontario Institute for Cancer Research, Toronto, ON M5G 0A3, Canada. 30Canada’s Michael Smith Genome Sciences Centre, BC Cancer, Vancouver, BC V5Z 4S6, Canada. 31Computational Biology Program, School of Medicine, Oregon Health and Science University, Portland, OR 97239, USA. 32DNAnexus Inc, Mountain View, CA 94040, USA. 33Department of Systems Biology, UT MD Anderson Cancer Center, Houston, TX 77030, USA. 34Precision Oncology, OHSU Knight Cancer Institute, Oregon Health and Science University, Portland, OR 97239, USA. 35Computer Network Information Center, Chinese Academy of Sciences, Beijing, China. 36Institute for Systems Biology, Seattle, WA 98109, USA. 37Department of Biomolecular Engineering, University of California Santa Cruz, Santa Cruz, CA 95064, USA. 38Department of Bioinformatics and Computational Biology and Department of Systems Biology, The University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA. 39Biomolecular Engineering Department, University of California Santa Cruz, Santa Cruz, CA 95064, USA. 40School of Elect, PCAWG novel somatic mutation calling methods working group: Matthew H. Bailey1,2,3, Beifang Niu35, Matthias Bieg42,43, Paul C. Boutros6,44,45,46, Ivo Buchhalter43,47,48, Adam P. Butler49, Ken Chen50, Zechen Chong51, Li Ding1,2,3,20, Oliver Drechsel52,53, Lewis Jonathan Dursi6,7, Roland Eils47,48,54,55, Kyle Ellrott9, Shadrielle M. G. Espiritu6, Yu Fan21, Robert S. Fulton1,3,20, Shengjie Gao56, Josep L. l. Gelpi57,58, Mark B. Gerstein5,18,19, Gad Getz8,14,15,16, Santiago Gonzalez59,60, Ivo G. Gut52,61, Faraz Hach62,63, Michael C. Heinold47,48, Julian M. Hess8,26, Jonathan Hinton49, Taobo Hu64, Vincent Huang6, Yi Huang65,66, Barbara Hutter43,67,68, David R. Jones49, Jongsun Jung69, Natalie Jäger47, Hyung-Lae Kim70, Kortine Kleinheinz47,48, Sushant Kumar5,19, Yogesh Kumar64, Christopher M. Lalansingh6, Ignaty Leshchiner8, Ivica Letunic71, Dimitri Livitz8, Eric Z. Ma64, Yosef E. Maruvka8,26,72, R. Jay Mashl1,2, Michael D. McLellan1,3,20, Andrew Menzies49, Ana Milovanovic57, Morten Muhlig Nielsen73, Stephan Ossowski52,53,74, Nagarajan Paramasivam43,47, Jakob Skou Pedersen73,75, Marc D. Perry76,77, Montserrat Puiggròs57, Keiran M. Raine49, Esther Rheinbay8,14,72, Romina Royo57, S. Cenk Sahinalp62,78,79, Gordon Saksena8, Iman Sarrafi62,78, Matthias Schlesner47,80, Jared T. Simpson6,17, Lucy Stebbings49, Chip Stewart8, Miranda D. Stobbe52,61, Jon W. Teague49, Grace Tiao8, David Torrents57,81, Jeremiah A. Wala8,14,82, Jiayin Wang1,40,66, Wenyi Wang21, Sebastian M. Waszak60, Joachim Weischenfeldt60,83,84, Michael C. Wendl1,10,11, Johannes Werner47,85, Zhenggang Wu64, Hong Xue64, Sergei Yakneen60, Takafumi N. Yamaguchi6, Kai Ye40,41, Venkata D. Yellapantula20,86, Christina K. Yung76 & Junjun Zhang76, PCAWG Consortium: Lauri A. Aaltonen87, Federico Abascal49, Adam Abeshouse88, Hiroyuki Aburatani89, David J. Adams49, Nishant Agrawal90, Keun Soo Ahn91, Sung-Min Ahn92, Hiroshi Aikata93, Rehan Akbani21, Kadir C. Akdemir50, Hikmat Al-Ahmadie88, Sultan T. Al-Sedairy94, Fatima Al-Shahrour95, Malik Alawi96,97, Monique Albert98, Kenneth Aldape99,100, Ludmil B. Alexandrov49,101,102, Adrian Ally30, Kathryn Alsop103, Eva G. Alvarez104,105,106, Fernanda Amary107, Samirkumar B. Amin108,109,110, Brice Aminou76, Ole Ammerpohl111,112, Matthew J. Anderson113, Yeng Ang114, Davide Antonello115, Pavana Anur22, Samuel Aparicio116, Elizabeth L. Appelbaum1,117, Yasuhito Arai118, Axel Aretz119, Koji Arihiro93, Shun-ichi Ariizumi120, Joshua Armenia121, Laurent Arnould122, Sylvia Asa123,124, Yassen Assenov125, Gurnit Atwal6,126,127, Sietse Aukema112,128, J. Todd Auman129, Miriam R. Aure130, Philip Awadalla6,126, Marta Aymerich131, Gary D. Bader126, Adrian Baez-Ortega132, Matthew H. Bailey1,2,3, Peter J. Bailey133, Miruna Balasundaram30, Saianand Balu134, Pratiti Bandopadhayay8,135,136, Rosamonde E. Banks137, Stefano Barbi138, Andrew P. Barbour139,140, Jonathan Barenboim6, Jill Barnholtz-Sloan141,142, Hugh Barr143, Elisabet Barrera59, John Bartlett98,144, Javier Bartolome57, Claudio Bassi115, Oliver F. Bathe145,146, Daniel Baumhoer147, Prashant Bavi148, Stephen B. Baylin149,150, Wojciech Bazant59, Duncan Beardsmore151, Timothy A. Beck152,153, Sam Behjati49, Andreas Behren154, Beifang Niu35, Cindy Bell155, Sergi Beltran52,61, Christopher Benz156, Andrew Berchuck157, Anke K. Bergmann158, Erik N. Bergstrom101,102, Benjamin P. Berman159,160,161, Daniel M. Berney162, Stephan H. Bernhart163,164,165, Rameen Beroukhim8,14,82, Mario Berrios166, Samantha Bersani167, Johanna Bertl73,168, Miguel Betancourt169, Vinayak Bhandari6,44, Shriram G. Bhosle49, Andrew V. Biankin133,170,171,172, Matthias Bieg42,43, Darell Bigner173, Hans Binder163,164, Ewan Birney59, Michael Birrer72, Nidhan K. Biswas174, Bodil Bjerkehagen147,175, Tom Bodenheimer134, Lori Boice176, Giada Bonizzato177, Johann S. De Bono178, Arnoud Boot179,180, Moiz S. Bootwalla166, Ake Borg181, Arndt Borkhardt182, Keith A. Boroevich183,184, Ivan Borozan6, Christoph Borst185, Marcus Bosenberg186, Mattia Bosio52,53,57, Jacqueline Boultwood187, Guillaume Bourque188,189, Paul C. Boutros6,44,45,46, G. Steven Bova190, David T. Bowen49,191, Reanne Bowlby30, David D. L. Bowtell103, Sandrine Boyault192, Rich Boyce59, Jeffrey Boyd193, Alvis Brazma59, Paul Brennan194, Daniel S. Brewer195,196, Arie B. Brinkman197, Robert G. Bristow44,198,199,200,201, Russell R. Broaddus99, Jane E. Brock202, Malcolm Brock203, Annegien Broeks204, Angela N. Brooks8,24,37,82, Denise Brooks30, Benedikt Brors67,205,206, Søren Brunak207,208, Timothy J. C. Bruxner113,209, Alicia L. Bruzos104,105,106, Alex Buchanan9, Ivo Buchhalter43,47,48, Christiane Buchholz210, Susan Bullman8,82, Hazel Burke211, Birgit Burkhardt212, Kathleen H. Burns213,214, John Busanovich8,215, Carlos D. Bustamante216,217, Adam P. Butler49, Atul J. Butte218, Niall J. Byrne76, Anne-Lise Børresen-Dale130,219, Samantha J. Caesar-Johnson220, Andy Cafferkey59, Declan Cahill221, Claudia Calabrese59,60, Carlos Caldas222,223, Fabien Calvo224, Niedzica Camacho178, Peter J. Campbell49,225, Elias Campo226,227, Cinzia Cantù177, Shaolong Cao21, Thomas E. Carey228, Joana Carlevaro-Fita229,230,231, Rebecca Carlsen30, Ivana Cataldo167,177, Mario Cazzola232, Jonathan Cebon154, Robert Cerfolio233, Dianne E. Chadwick234, Dimple Chakravarty235, Don Chalmers236, Calvin Wing Yiu Chan47,237, Kin Chan238, Michelle Chan-Seng-Yue148, Vishal S. Chandan239, David K. Chang133,170, Stephen J. Chanock240, Lorraine A. Chantrill170,241, Aurélien Chateigner76,242, Nilanjan Chatterjee149,243, Kazuaki Chayama93, Hsiao-Wei Chen114,121, Jieming Chen218, Ken Chen50, Yiwen Chen21, Zhaohong Chen244, Andrew D. Cherniack8,82, Jeremy Chien245, Yoke-Eng Chiew246,247, Suet-Feung Chin222,223, Juok Cho8, Sunghoon Cho248, Jung Kyoon Choi249, Wan Choi250, Christine Chomienne251, Zechen Chong51, Su Pin Choo252, Angela Chou170,246, Angelika N. Christ113, Elizabeth L. Christie103, Eric Chuah30, Carrie Cibulskis8, Kristian Cibulskis8, Sara Cingarlini253, Peter Clapham49, Alexander Claviez254, Sean Cleary148,255, Nicole Cloonan256, Marek Cmero257,258,259, Colin C. Collins62, Ashton A. Connor255,260, Susanna L. Cooke133, Colin S. Cooper178,196,261, Leslie Cope149, Vincenzo Corbo138,177, Matthew G. Cordes1,262, Stephen M. Cordner263, Isidro Cortés-Ciriano264,265,266, Kyle Covington12,23, Prue A. Cowin267, Brian Craft24, David Craft8,268, Chad J. Creighton269, Yupeng Cun270, Erin Curley271, Ioana Cutcutache179,180, Karolina Czajka272, Bogdan Czerniak99,273, Rebecca A. Dagg274, Ludmila Danilova149, Maria Vittoria Davi275, Natalie R. Davidson276,277,278,279,280, Helen Davies49,281,282, Ian J. Davis283, Brandi N. Davis-Dusenbery284, Kevin J. Dawson49, Francisco M. De La Vega216,217,285, Ricardo De Paoli-Iseppi211, Timothy Defreitas8, Angelo P. Dei Tos286, Olivier Delaneau287,288,289, John A. Demchok220, Jonas Demeulemeester290,291, German M. Demidov52,53,74, Deniz Demircioğlu292,293, Nening M. Dennis221, Robert E. Denroche148, Stefan C. Dentro49,290,294, Nikita Desai76, Vikram Deshpande72, Amit G. Deshwar295, Christine Desmedt296,297, Jordi Deu-Pons298,299, Noreen Dhalla30, Neesha C. Dhani300, Priyanka Dhingra301,302, Rajiv Dhir303, Anthony DiBiase304, Klev Diamanti305, Li Ding1,2,3,20, Shuai Ding306, Huy Q. Dinh159, Luc Dirix307, HarshaVardhan Doddapaneni12, Nilgun Donmez62,78, Michelle T. Dow244, Ronny Drapkin308, Oliver Drechsel52,53, Ruben M. Drews223, Serge Serge49, Tim Dudderidge150,221, Ana Dueso-Barroso57, Andrew J. Dunford8, Michael Dunn309, Lewis Jonathan Dursi6,7, Fraser R. Duthie133,310, Ken Dutton-Regester311, Jenna Eagles272, Douglas F. Easton312,313, Stuart Edmonds314, Paul A. Edwards223,315, Sandra E. Edwards178, Rosalind A. Eeles178,221, Anna Ehinger316, Juergen Eils54,55, Roland Eils47,48,54,55, Adel El-Naggar99,273, Matthew Eldridge223, Kyle Ellrott9, Serap Erkek60, Georgia Escaramis53,317,318, Shadrielle M. G. Espiritu6, Xavier Estivill53,319, Dariush Etemadmoghadam103, Jorunn E. Eyfjord320, Bishoy M. Faltas280, Daiming Fan321, Yu Fan21, William C. Faquin72, Claudiu Farcas244, Matteo Fassan322, Aquila Fatima323, Francesco Favero324, Nodirjon Fayzullaev76, Ina Felau220, Sian Fereday103, Martin L. Ferguson325, Vincent Ferretti76,326, Lars Feuerbach205, Matthew A. Field327, J. Lynn Fink57,113, Gaetano Finocchiaro328, Cyril Fisher221, Matthew W. Fittall290, Anna Fitzgerald329, Rebecca C. Fitzgerald282, Adrienne M. Flanagan330, Neil E. Fleshner331, Paul Flicek59, John A. Foekens332, Kwun M. Fong333, Nuno A. Fonseca59,334, Christopher S. Foster335,336, Natalie S. Fox6, Michael Fraser6, Scott Frazer8, Milana Frenkel-Morgenstern337, William Friedman338, Joan Frigola298, Catrina C. Fronick1,262, Akihiro Fujimoto184, Masashi Fujita184, Masashi Fukayama339, Lucinda A. Fulton1 , Robert S. Fulton1,3,20, Mayuko Furuta184, P. Andrew Futreal340, Anja Füllgrabe59, Stacey B. Gabriel8, Steven Gallinger148,255,260, Carlo Gambacorti-Passerini341, Jianjiong Gao121, Shengjie Gao56, Levi Garraway82, Øystein Garred342, Erik Garrison49, Dale W. Garsed103, Nils Gehlenborg8,343, Josep L. l. Gelpi57,58, Joshy George110, Daniela S. Gerhard344, Clarissa Gerhauser345, Jeffrey E. Gershenwald346,347, Mark B. Gerstein5,18,19, Moritz Gerstung59,60, Gad Getz8,14,15,16, Mohammed Ghori49, Ronald Ghossein348, Nasra H. Giama349, Richard A. Gibbs12, Anthony J. Gill170,350, Pelvender Gill351, Dilip D. Giri348, Dominik Glodzik49, Vincent J. Gnanapragasam352,353, Maria Elisabeth Goebler354, Mary J. Goldman24, Carmen Gomez355, Santiago Gonzalez59,60, Abel Gonzalez-Perez298,299,356, Dmitry A. Gordenin357, James Gossage358, Kunihito Gotoh359, Ramaswamy Govindan3, Dorthe Grabau360, Janet S. Graham133,361, Robert C. Grant148,260, Anthony R. Green315, Eric Green27, Liliana Greger59, Nicola Grehan282, Sonia Grimaldi177, Sean M. Grimmond362, Robert L. Grossman363, Adam Grundhoff97,364, Gunes Gundem88, Qianyun Guo75, Manaswi Gupta8, Shailja Gupta365, Ivo G. Gut52,61, Marta Gut52,61, Jonathan Göke292,366, Gavin Ha8, Andrea Haake111, David Haan37, Siegfried Haas185, Kerstin Haase290, James E. Haber367, Nina Habermann60, Faraz Hach62,63, Syed Haider6, Natsuko Hama118, Freddie C. Hamdy351, Anne Hamilton267, Mark P. Hamilton368, Leng Han369, George B. Hanna370, Martin Hansmann371, Nicholas J. Haradhvala8,72, Olivier Harismendy102,372, Ivon Harliwong113, Arif O. Harmanci5,373, Eoghan Harrington374, Takanori Hasegawa375, David Haussler24,25, Steve Hawkins223, Shinya Hayami376, Shuto Hayashi375, D. Neil Hayes134,377,378, Stephen J. Hayes379,380, Nicholas K. Hayward211,311, Steven Hazell221, Yao He381, Allison P. Heath382, Simon C. Heath52,61, David Hedley300, Apurva M. Hegde38, David I. Heiman8, Michael C. Heinold47,48, Zachary Heins88, Lawrence E. Heisler152, Eva Hellstrom-Lindberg383, Mohamed Helmy384, Seong Gu Heo385, Austin J. Hepperla134, José María Heredia-Genestar386, Carl Herrmann47,48,387, Peter Hersey211, Julian M. Hess8,26, Holmfridur Hilmarsdottir320, Jonathan Hinton49, Satoshi Hirano388, Nobuyoshi Hiraoka389, Katherine A. Hoadley134,390, Asger Hobolth75,168, Ermin Hodzic78, Jessica I. Hoell182, Steve Hoffmann163,164,165,391, Oliver Hofmann392, Andrea Holbrook166, Aliaksei Z. Holik53, Michael A. Hollingsworth393, Oliver Holmes209,311, Robert A. Holt30, Chen Hong205,237, Eun Pyo Hong385, Jongwhi H. Hong394, Gerrit K. Hooijer395, Henrik Hornshøj73, Fumie Hosoda118, Yong Hou56,396, Volker Hovestadt397, William Howat352, Alan P. Hoyle134, Ralph H. Hruban149, Jianhong Hu12, Taobo Hu64, Xing Hua240, Kuan-lin Huang1,398, Mei Huang176, Mi Ni Huang179,180, Vincent Huang6, Yi Huang65,66, Wolfgang Huber60, Thomas J. Hudson272,399, Michael Hummel400, Jillian A. Hung246,247, David Huntsman401, Ted R. Hupp402, Jason Huse88, Matthew R. Huska403, Barbara Hutter43,67,68, Carolyn M. Hutter27, Daniel Hübschmann48,54,404,405,406, Christine A. Iacobuzio-Donahue348, Charles David Imbusch205, Marcin Imielinski407,408, Seiya Imoto375, William B. Isaacs409, Keren Isaev6,44, Shumpei Ishikawa410, Murat Iskar397, S. M. Ashiqul Islam244, Michael Ittmann411,412,413, Sinisa Ivkovic284, Jose M. G. Izarzugaza414, Jocelyne Jacquemier415, Valerie Jakrot211, Nigel B. Jamieson133,172,416, Gun Ho Jang148, Se Jin Jang417, Joy C. Jayaseelan12, Reyka Jayasinghe1 , Stuart R. Jefferys134, Karine Jegalian418, Jennifer L. Jennings419, Seung-Hyup Jeon250, Lara Jerman60,420, Yuan Ji421,422, Wei Jiao6, Peter A. Johansson311, Amber L. Johns170, Jeremy Johns272, Rory Johnson230,423, Todd A. Johnson183, Clemency Jolly290, Yann Joly424, Jon G. Jonasson320, Corbin D. Jones425, David R. Jones49, David T. W. Jones426,427, Nic Jones428, Steven J. M. Jones30, Jos Jonkers204, Young Seok Ju49,249, Hartmut Juhl429, Jongsun Jung69, Malene Juul73, Randi Istrup Juul73, Sissel Juul374, Natalie Jäger47, Rolf Kabbe47, Andre Kahles276,277,278,279,430, Abdullah Kahraman431,432,433, Vera B. Kaiser434, Hojabr Kakavand211, Sangeetha Kalimuthu148, Christof von Kalle405, Koo Jeong Kang91, Katalin Karaszi351, Beth Karlan435, Rosa Karlić436, Dennis Karsch437, Katayoon Kasaian29,30, Karin S. Kassahn113,438, Hitoshi Katai439, Mamoru Kato440, Hiroto Katoh410, Yoshiiku Kawakami93, Jonathan D. Kay117, Stephen H. Kazakoff209,311, Marat D. Kazanov441,442,443, Maria Keays59, Electron Kebebew444,445, Richard F. Kefford446, Manolis Kellis8,447, James G. Kench170,350,448, Catherine J. Kennedy246,247, Jules N. A. Kerssemakers47, David Khoo273, Vincent Khoo221, Narong Khuntikeo115,449, Ekta Khurana301,302,450,451, Helena Kilpinen117, Hark Kyun Kim452, Hyung-Lae Kim70, Hyung-Yong Kim415, Hyunghwan Kim250, Jaegil Kim8, Jihoon Kim453, Jong K. Kim454, Youngwook Kim455,456, Tari A. King457,458,459, Wolfram Klapper128, Kortine Kleinheinz47,48, Leszek J. Klimczak460, Stian Knappskog49,461, Michael Kneba437, Bartha M. Knoppers424, Youngil Koh462,463, Jan Komorowski305,464, Daisuke Komura410, Mitsuhiro Komura375, Gu Kong415, Marcel Kool426,465, Jan O. Korbel59,60, Viktoriya Korchina12, Andrey Korshunov465, Michael Koscher465, Roelof Koster466, Zsofia Kote-Jarai178, Antonios Koures244, Milena Kovacevic284, Barbara Kremeyer49, Helene Kretzmer164,165, Markus Kreuz467, Savitri Krishnamurthy99,468, Dieter Kube469, Kiran Kumar8, Pardeep Kumar221, Sushant Kumar5,19, Yogesh Kumar64, Ritika Kundra114,121, Kirsten Kübler8,14,72, Ralf Küppers470, Jesper Lagergren383,471, Phillip H. Lai166, Peter W. Laird472, Sunil R. Lakhani473, Christopher M. Lalansingh6, Emilie Lalonde6, Fabien C. Lamaze6, Adam Lambert351, Eric Lander8, Pablo Landgraf474,475, Luca Landoni115, Anita Langerød130, Andrés Lanzós230,231,423, Denis Larsimont476, Erik Larsson477, Mark Lathrop189, Loretta M. S. Lau478, Chris Lawerenz55, Rita T. Lawlor177, Michael S. Lawrence8,72,183, Alexander J. Lazar99,108, Xuan Le479, Darlene Lee30, Donghoon Lee5, Eunjung Alice Lee480, Hee Jin Lee417, Jake June-Koo Lee264,266, Jeong-Yeon Lee481, Juhee Lee482, Ming Ta Michael Lee340, Henry Lee-Six49, Kjong-Van Lehmann276,277,278,279,430, Hans Lehrach483, Dido Lenze400, Conrad R. Leonard209,311, Daniel A. Leongamornlert49,178, Ignaty Leshchiner8, Louis Letourneau484, Ivica Letunic71, Douglas A. Levine88,485, Lora Lewis12, Tim Ley486, Chang Li56,396, Constance H. Li6,44, Haiyan Irene Li30, Jun Li21, Lin Li56, Shantao Li5, Siliang Li56,396, Xiaobo Li56,396, Xiaotong Li5, Xinyue Li56, Yilong Li49, Han Liang21, Sheng-Ben Liang234, Peter Lichter68,397, Pei Lin8, Ziao Lin8,487, W. M. Linehan488, Ole Christian Lingjærde489, Dongbing Liu56,396, Eric Minwei Liu88,301,302, Fei-Fei Liu201,490, Fenglin Liu381,491, Jia Liu492, Xingmin Liu56,396, Julie Livingstone6, Dimitri Livitz8, Naomi Livni221, Lucas Lochovsky5,19,110, Markus Loeffler467, Georgina V. Long211, Armando Lopez-Guillermo493, Shaoke Lou5,19, David N. Louis72, Laurence B. Lovat117, Yiling Lu38, Yong-Jie Lu162,494, Youyong Lu495,496,497, Claudio Luchini167, Ilinca Lungu144,148, Xuemei Luo152, Hayley J. Luxton117, Andy G. Lynch223,315,498, Lisa Lype36, Cristina López111,112, Carlos López-Otín499, Eric Z. Ma64, Yussanne Ma30, Gaetan MacGrogan500, Shona MacRae501, Geoff Macintyre223, Tobias Madsen73, Kazuhiro Maejima184, Andrea Mafficini177, Dennis T. Maglinte166,502, Arindam Maitra174, Partha P. Majumder174, Luca Malcovati232, Salem Malikic62,78, Giuseppe Malleo115, Graham J. Mann211,246,503, Luisa Mantovani-Löffler504, Kathleen Marchal505,506, Giovanni Marchegiani115, Elaine R. Mardis1,193,507, Adam A. Margolin31, Maximillian G. Marin37, Florian Markowetz223,315, Julia Markowski403, Jeffrey Marks508, Tomas Marques-Bonet61,81,386,509, Marco A. Marra30, Luke Marsden351, John W. M. Martens332, Sancha Martin49,510, Jose I. Martin-Subero81,511, Iñigo Martincorena49, Alexander Martinez-Fundichely301,302,451 Yosef E. Maruvka8,26,72, R. Jay Mashl1,2, Charlie E. Massie223, Thomas J. Matthew37, Lucy Matthews178, Erik Mayer221,512, Simon Mayes513, Michael Mayo30, Faridah Mbabaali272, Karen McCune514, Ultan McDermott49, Patrick D. McGillivray19, Michael D. McLellan1,3,20, John D. McPherson148,272,515, John R. McPherson179,180, Treasa A. McPherson260, Samuel R. Meier8, Alice Meng516, Shaowu Meng134, Andrew Menzies49, Neil D. Merrett115,517, Sue Merson178, Matthew Meyerson8,14,82, William U. Meyerson4,5, Piotr A. Mieczkowski518, George L. Mihaiescu76, Sanja Mijalkovic284, Ana Mijalkovic Mijalkovic-Lazic284, Tom Mikkelsen519, Michele Milella253, Linda Mileshkin103, Christopher A. Miller1 , David K. Miller113,170, Jessica K. Miller272, Gordon B. Mills33,34, Ana Milovanovic57, Sarah Minner520, Marco Miotto115, Gisela Mir Arnau267, Lisa Mirabello240, Chris Mitchell103, Thomas J. Mitchell49,315,352, Satoru Miyano375, Naoki Miyoshi375, Shinichi Mizuno521, Fruzsina Molnár-Gábor522, Malcolm J. Moore300, Richard A. Moore30, Sandro Morganella49, Quaid D. Morris127,490, Carl Morrison523,524, Lisle E. Mose134, Catherine D. Moser349, Ferran Muiños298,299, Loris Mularoni298,299, Andrew J. Mungall30, Karen Mungall30, Elizabeth A. Musgrove133, Ville Mustonen525,526,527, David Mutch528, Francesc Muyas52,53,74, Donna M. Muzny12, Alfonso Muñoz59, Jerome Myers529, Ola Myklebost461, Peter Möller530, Genta Nagae89, Adnan M. Nagrial170, Hardeep K. Nahal-Bose76, Hitoshi Nakagama531, Hidewaki Nakagawa184, Hiromi Nakamura118, Toru Nakamura388, Kaoru Nakano184, Tannistha Nandi532, Jyoti Nangalia49, Mia Nastic284, Arcadi Navarro61,81,386, Fabio C. P. Navarro19, David E. Neal223,352, Gerd Nettekoven533, Felicity Newell209,311, Steven J. Newhouse59, Yulia Newton37, Alvin Wei Tian Ng534, Anthony Ng535, Jonathan Nicholson49, David Nicol221, Yongzhan Nie321,536, G. Petur Nielsen72, Morten Muhlig Nielsen73, Serena Nik-Zainal49,281,282,537, Michael S. Noble8, Katia Nones209,311, Paul A. Northcott538, Faiyaz Notta148,539, Brian D. O’Connor76,540, Peter O’Donnell541, Maria O’Donovan282, Sarah O’Meara49, Brian Patrick O’Neill542, J. Robert O’Neill543, David Ocana59, Angelica Ochoa88, Layla Oesper544, Christopher Ogden221, Hideki Ohdan93, Kazuhiro Ohi375, Lucila Ohno-Machado244, Karin A. Oien523,545, Akinyemi I. Ojesina546,547,548, Hidenori Ojima549, Takuji Okusaka550, Larsson Omberg551, Choon Kiat Ong552, Stephan Ossowski52,53,74, German Ott553, B. F. Francis Ouellette76,554, Christine P’ng6, Marta Paczkowska6, Salvatore Paiella115, Chawalit Pairojkul523, Marina Pajic170, Qiang Pan-Hammarström56,555, Elli Papaemmanuil49, Irene Papatheodorou59, Nagarajan Paramasivam43,47, Ji Wan Park385, Joong-Won Park556, Keunchil Park557,558, Kiejung Park559, Peter J. Park264,266, Joel S. Parker518, Simon L. Parsons124, Harvey Pass560, Danielle Pasternack272, Alessandro Pastore276, Ann-Marie Patch209,311, Iris Pauporté251, Antonio Pea115, John V. Pearson209,311, Chandra Sekhar Pedamallu8,14,82, Jakob Skou Pedersen73,75, Paolo Pederzoli115, Martin Peifer270, Nathan A. Pennell561, Charles M. Perou129,518, Marc D. Perry76,77, Gloria M. Petersen562, Myron Peto22, Nicholas Petrelli563, Robert Petryszak59, Stefan M. Pfister426,465,564, Mark Phillips424, Oriol Pich298,299, Hilda A. Pickett478, Todd D. Pihl565, Nischalan Pillay566, Sarah Pinder567, Mark Pinese170, Andreia V. Pinho568, Esa Pitkänen60, Xavier Pivot569, Elena Piñeiro-Yáñez95, Laura Planko533, Christoph Plass345, Paz Polak8,14,15, Tirso Pons570, Irinel Popescu571, Olga Potapova572, Aparna Prasad52, Shaun R. Preston573, Manuel Prinz47, Antonia L. Pritchard311, Stephenie D. Prokopec6, Elena Provenzano574, Xose S. Puente499, Sonia Puig176, Montserrat Puiggròs57, Sergio Pulido-Tamayo505,506, Gulietta M. Pupo246, Colin A. Purdie575, Michael C. Quinn209,311, Raquel Rabionet52,53,576, Janet S. Rader577, Bernhard Radlwimmer397, Petar Radovic284, Benjamin Raeder60, Keiran M. Raine49, Manasa Ramakrishna49, Kamna Ramakrishnan49, Suresh Ramalingam578, Benjamin J. Raphael579, W. Kimryn Rathmell580, Tobias Rausch60, Guido Reifenberger475, Jüri Reimand6,44, Jorge Reis-Filho348, Victor Reuter348, Iker Reyes-Salazar298, Matthew A. Reyna579, Sheila M. Reynolds36, Esther Rheinbay8,14,72, Yasser Riazalhosseini189, Andrea L. Richardson323, Julia Richter111,128, Matthew Ringel581, Markus Ringnér181, Yasushi Rino582, Karsten Rippe405, Jeffrey Roach583, Lewis R. Roberts349, Nicola D. Roberts49, Steven A. Roberts584, A. Gordon Robertson30, Alan J. Robertson113, Javier Bartolomé Rodriguez57, Bernardo Rodriguez-Martin104,105,106, F. Germán Rodríguez-González83,332, Michael H. A. Roehrl44,123,148,234,585,586, Marius Rohde587, Hirofumi Rokutan440, Gilles Romieu588, Ilse Rooman170, Tom Roques262, Daniel Rosebrock8, Mara Rosenberg8,72, Philip C. Rosenstiel589, Andreas Rosenwald590, Edward W. Rowe221,591, Romina Royo57, Steven G. Rozen179,180,592, Yulia Rubanova17,127, Mark A. Rubin423,593,594,595,596, Carlota Rubio-Perez298,299,597, Vasilisa A. Rudneva60, Borislav C. Rusev177, Andrea Ruzzenente598, Gunnar Rätsch276,277,278,279,280,430, Radhakrishnan Sabarinathan298,299,599, Veronica Y. Sabelnykova6, Sara Sadeghi30, S. Cenk Sahinalp62,78,79, Natalie Saini357, Mihoko Saito-Adachi440, Gordon Saksena8, Adriana Salcedo6, Roberto Salgado600, Leonidas Salichos5,19, Richard Sallari8, Charles Saller601, Roberto Salvia115, Michelle Sam272, Jaswinder S. Samra115,602, Francisco Sanchez-Vega114,121, Chris Sander276,603,604, Grant Sanders134, Rajiv Sarin605, Iman Sarrafi62,78, Aya Sasaki-Oku184, Torill Sauer489, Guido Sauter520, Robyn P. M. Saw211, Maria Scardoni167, Christopher J. Scarlett170,606, Aldo Scarpa177, Ghislaine Scelo194, Dirk Schadendorf68,607, Jacqueline E. Schein30, Markus B. Schilhabel589, Matthias Schlesner47,80, Thorsten Schlomm84,608, Heather K. Schmidt1 , Sarah-Jane Schramm246, Stefan Schreiber609, Nikolaus Schultz121, Steven E. Schumacher8,323, Roland F. Schwarz59,403,405,610, Richard A. Scolyer211,448,602, David Scott428, Ralph Scully611, Raja Seethala612, Ayellet V. Segre8,613, Iris Selander260, Colin A. Semple434, Yasin Senbabaoglu276, Subhajit Sengupta614, Elisabetta Sereni115, Stefano Serra585, Dennis C. Sgroi72, Mark Shackleton103, Nimish C. Shah352, Sagedeh Shahabi234, Catherine A. Shang329, Ping Shang211, Ofer Shapira8,323, Troy Shelton271, Ciyue Shen603,604, Hui Shen615, Rebecca Shepherd49, Ruian Shi490, Yan Shi134, Yu-Jia Shiah6, Tatsuhiro Shibata118,616, Juliann Shih8,82, Eigo Shimizu375, Kiyo Shimizu617, Seung Jun Shin618, Yuichi Shiraishi375, Tal Shmaya285, Ilya Shmulevich36, Solomon I. Shorser6, Charles Short59, Raunak Shrestha62, Suyash S. Shringarpure217, Craig Shriver619, Shimin Shuai6,126, Nikos Sidiropoulos83, Reiner Siebert112,620, Anieta M. Sieuwerts332, Lina Sieverling205,237, Sabina Signoretti202,621, Katarzyna O. Sikora177, Michele Simbolo138, Ronald Simon520, Janae V. Simons134, Jared T. Simpson6,17, Peter T. Simpson473, Samuel Singer115,458, Nasa Sinnott-Armstrong8,217, Payal Sipahimalani30, Tara J. Skelly390, Marcel Smid332, Jaclyn Smith622, Karen Smith-McCune514, Nicholas D. Socci276, Heidi J. Sofia27, Matthew G. Soloway134, Lei Song240, Anil K. Sood623,624,625, Sharmila Sothi626, Christos Sotiriou244, Cameron M. Soulette37, Paul N. Span627, Paul T. Spellman22, Nicola Sperandio177, Andrew J. Spillane211, Oliver Spiro8, Jonathan Spring628, Johan Staaf181, Peter F. Stadler163,164,165, Peter Staib629, Stefan G. Stark277,279,618,630, Lucy Stebbings49, Ólafur Andri Stefánsson631, Oliver Stegle59,60,632, Lincoln D. Stein6,126, Alasdair Stenhouse633, Chip Stewart8, Stephan Stilgenbauer634, Miranda D. Stobbe52,61, Michael R. Stratton49, Jonathan R. Stretch211, Adam J. Struck31, Joshua M. Stuart24,37, Henk G. Stunnenberg396,635, Hong Su56,396, Xiaoping Su99, Ren X. Sun6, Stephanie Sungalee60, Hana Susak52,53, Akihiro Suzuki89,636, Fred Sweep637, Monika Szczepanowski128, Holger Sültmann67,638, Takashi Yugawa617, Angela Tam30, David Tamborero298,299, Benita Kiat Tee Tan639, Donghui Tan518, Patrick Tan180,532,592,640, Hiroko Tanaka375, Hirokazu Taniguchi616, Tomas J. Tanskanen641, Maxime Tarabichi49,290, Roy Tarnuzzer220, Patrick Tarpey642, Morgan L. Taschuk152, Kenji Tatsuno89, Simon Tavaré223,643, Darrin F. Taylor113, Amaro Taylor-Weiner8, Jon W. Teague49, Bin Tean Teh180,592,640,644,645, Varsha Tembe246, Javier Temes104,105, Kevin Thai76, Sarah P. Thayer393, Nina Thiessen30, Gilles Thomas646, Sarah Thomas221, Alan Thompson221, Alastair M. Thompson633, John F. Thompson211, R. Houston Thompson647, Heather Thorne103, Leigh B. Thorne176, Adrian Thorogood424, Grace Tiao8, Nebojsa Tijanic284, Lee E. Timms272, Roberto Tirabosco648, Marta Tojo106, Stefania Tommasi649, Christopher W. Toon170, Umut H. Toprak48,650, David Torrents57,81, Giampaolo Tortora651,652, Jörg Tost653, Yasushi Totoki118, David Townend654, Nadia Traficante103, Isabelle Treilleux655,656, Jean-Rémi Trotta61, Lorenz H. P. Trümper469, Ming Tsao124,539, Tatsuhiko Tsunoda183,657,658,659, Jose M. C. Tubio104,105,106, Olga Tucker660, Richard Turkington661, Daniel J. Turner513, Andrew Tutt323, Masaki Ueno376, Naoto T. Ueno662, Christopher Umbricht151,213,663, Husen M. Umer305,664, Timothy J. Underwood665, Lara Urban59,60, Tomoko Urushidate616, Tetsuo Ushiku339, Liis Uusküla-Reimand666,667, Alfonso Valencia57,81, David J. Van Den Berg166, Steven Van Laere307, Peter Van Loo290,291, Erwin G. Van Meir668, Gert G. Van den Eynden307, Theodorus Van der Kwast123, Naveen Vasudev137, Miguel Vazquez57,669, Ravikiran Vedururu267, Umadevi Veluvolu518, Shankar Vembu490,670, Lieven P. C. Verbeke506,671, Peter Vermeulen307, Clare Verrill351,672, Alain Viari177, David Vicente57, Caterina Vicentini177, K. Vijay Raghavan365, Juris Viksna673, Ricardo E. Vilain674, Izar Villasante57, Anne Vincent-Salomon635, Tapio Visakorpi190, Douglas Voet8, Paresh Vyas311,351, Ignacio Vázquez-García49,86,675,676, Nick M. Waddell209, Nicola Waddell209,311, Claes Wadelius677, Lina Wadi6, Rabea Wagener111,112, Jeremiah A. Wala8,14,82, Jian Wang56, Jiayin Wang1,40,66, Linghua Wang12, Qi Wang465, Wenyi Wang21, Yumeng Wang21, Zhining Wang220, Paul M. Waring523, Hans-Jörg Warnatz483, Jonathan Warrell5,19, Anne Y. Warren352,678, Sebastian M. Waszak60, David C. Wedge49,294,679, Dieter Weichenhan345, Paul Weinberger680, John N. Weinstein38, Joachim Weischenfeldt60,83,84, Daniel J. Weisenberger166, Ian Welch681, Michael C. Wendl1,10,11, Johannes Werner47,85, Justin P. Whalley61,682, David A. Wheeler12,13, Hayley C. Whitaker117, Dennis Wigle683, Matthew D. Wilkerson518, Ashley Williams244, James S. Wilmott211, Gavin W. Wilson6,148, Julie M. Wilson148, Richard K. Wilson1,684, Boris Winterhoff685, Jeffrey A. Wintersinger17,127,384, Maciej Wiznerowicz686,687, Stephan Wolf688, Bernice H. Wong689, Tina Wong1,30, Winghing Wong690, Youngchoon Woo250, Scott Wood209,311, Bradly G. Wouters44, Adam J. Wright6, Derek W. Wright133,691, Mark H. Wright217, Chin-Lee Wu72, Dai-Ying Wu285, Guanming Wu692, Jianmin Wu170, Kui Wu56,396, Yang Wu179,180, Zhenggang Wu64, Liu Xi12, Tian Xia693, Qian Xiang76, Xiao Xiao66, Rui Xing497, Heng Xiong56,396, Qinying Xu209,311, Yanxun Xu694, Hong Xue64, Shinichi Yachida118,695, Sergei Yakneen60, Rui Yamaguchi375, Takafumi N. Yamaguchi6, Masakazu Yamamoto120, Shogo Yamamoto89, Hiroki Yamaue376, Fan Yang490, Huanming Yang56, Jean Y. Yang696, Liming Yang220, Lixing Yang697, Shanlin Yang306, Tsun-Po Yang270, Yang Yang369, Xiaotong Yao408,698, Marie-Laure Yaspo483, Lucy Yates49, Christina Yau156, Chen Ye56,396, Kai Ye40,41, Venkata D. Yellapantula20,86, Christopher J. Yoon249, Sung-Soo Yoon463, Fouad Yousif6, Jun Yu699, Kaixian Yu700, Willie Yu701, Yingyan Yu702, Ke Yuan223,510,703, Yuan Yuan21, Denis Yuen6, Takashi Yugawa617, Christina K. Yung76, Olga Zaikova704, Jorge Zamora49,104,105,106, Marc Zapatka397, Jean C. Zenklusen220, Thorsten Zenz67, Nikolajs Zeps705,706, Cheng-Zhong Zhang8,707, Fan Zhang381, Hailei Zhang8, Hongwei Zhang494, Hongxin Zhang121, Jiashan Zhang220, Jing Zhang5, Junjun Zhang76, Xiuqing Zhang56, Xuanping Zhang66,369, Yan Zhang5,708,709, Zemin Zhang381,710, Zhongming Zhao711, Liangtao Zheng381, Xiuqing Zheng381, Wanding Zhou615, Yong Zhou56, Bin Zhu240, Hongtu Zhu700,712, Jingchun Zhu24, Shida Zhu56,396, Lihua Zou713, Xueqing Zou49, Anna deFazio246,247,714, Nicholas van As221, Carolien H. M. van Deurzen715, Marc J. van de Vijver523, L. van’t Veer716 & Christian von Mering433,717, The Cancer Genome Atlas (TCGA) and International Cancer Genome Consortium (ICGC) curated consensus somatic mutation calls using whole exome sequencing (WES) and whole genome sequencing (WGS), respectively. Here, as part of the ICGC/TCGA Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium, which aggregated whole genome sequencing data from 2,658 cancers across 38 tumour types, we compare WES and WGS side-by-side from 746 TCGA samples, finding that ~80% of mutations overlap in covered exonic regions. We estimate that low variant allele fraction (VAF < 15%) and clonal heterogeneity contribute up to 68% of private WGS mutations and 71% of private WES mutations. We observe that ~30% of private WGS mutations trace to mutations identified by a single variant caller in WES consensus efforts. WGS captures both ~50% more variation in exonic regions and un-observed mutations in loci with variable GC-content. Together, our analysis highlights technological divergences between two reproducible somatic variant detection efforts.

Tightly regulated activity of the transcription factor MYC is essential for orderly cell proliferation. Upon deregulation, MYC elicits and promotes cancer progression. Proteasomal degradation is an essential element of MYC regulation, initiated by phosphorylation at Serine62 (Ser62) of the MB1 region. Here, we found that Ser62 phosphorylation peaks in mitosis, but that a fraction of nonphosphorylated MYC binds to the microtubules of the mitotic spindle. Consequently, the microtubule-destabilizing drug vincristine decreases wild-type MYC stability, whereas phosphorylation-deficient MYC is more stable, contributing to vincristine resistance and induction of polyploidy. PI3K inhibition attenuates postmitotic MYC formation and augments the cytotoxic effect of vincristine. Implications: The spindle's function as a docking site for MYC during mitosis may constitute a window of specific vulnerability to be exploited for cancer treatment.

Macrophages (Mφ) are abundantly present in the tumor microenvironment and may predict outcome in solid tumors and defined lymphoma subtypes. Mφ heterogeneity, the mechanisms of their recruitment, and their differentiation into lymphoma‐promoting, alternatively activated M2‐like phenotypes are still not fully understood. Therefore, further functional studies are required to understand biological mechanisms associated with human tumor‐associated Mφ (TAM). Here, we show that the global mRNA expression and protein abundance of human Mφ differentiated in Hodgkin lymphoma (HL)‐conditioned medium (CM) differ from those of Mφ educated by conditioned media from diffuse large B‐cell lymphoma (DLBCL) cells or, classically, by macrophage colony‐stimulating factor (M‐CSF). Conditioned media from HL cells support TAM differentiation through upregulation of surface antigens such as CD40, CD163, CD206, and PD‐L1. In particular, RNA and cell surface protein expression of mannose receptor 1 (MRC1)/CD206 significantly exceed the levels induced by classical M‐CSF stimulation in M2‐like Mφ; this is regulated by interleukin 13 to a large extent. Functionally, high CD206 enhances mannose‐dependent endocytosis and uptake of type I collagen. Together with high matrix metalloprotease9 secretion, HL‐TAMs appear to be active modulators of the tumor matrix. Preclinical in ovo models show that co‐cultures of HL cells with monocytes or Mφ support dissemination of lymphoma cells via lymphatic vessels, while tumor size and vessel destruction are decreased in comparison with lymphoma‐only tumors. Immunohistology of human HL tissues reveals a fraction of cases feature large numbers of CD206‐positive cells, with high MRC1 expression being characteristic of HL‐stage IV. In summary, the lymphoma‐TAM interaction contributes to matrix‐remodeling and lymphoma cell dissemination., The study highlights the ability of Hodgkin lymphoma cells to attract and to differentiate monocytes into M2‐like Mφ and the role of IL13 in regulating CD206 but also that CD206 contributes to the remodeling of the tumor microenvironment via take up glycoconjugates as well as type‐I collagen and immunosuppression by PD‐L1 upregulation.

The Epstein-Barr virus (EBV) is found almost exclusively in the activated B-cell (ABC) subtype of diffuse large B-cell lymphoma (DLBCL), yet its contribution to this tumour remains poorly understood. We have focused on the EBV-encoded latent membrane protein-1 (LMP1), a constitutively activated CD40 homologue expressed in almost all EBV-positive DLBCLs and which can disrupt germinal centre (GC) formation and drive lymphomagenesis in mice. Comparison of the transcriptional changes that follow LMP1 expression with those that follow transient CD40 signalling in human GC B cells enabled us to define pathogenic targets of LMP1 aberrantly expressed in ABC-DLBCL. These included the down-regulation of S1PR2, a sphingosine-1-phosphate (S1P) receptor that is transcriptionally down-regulated in ABC-DLBCL, and when genetically ablated leads to DLBCL in mice. Consistent with this, we found that LMP1-expressing primary ABC-DLBCLs were significantly more likely to lack S1PR2 expression than were LMP1-negative tumours. Furthermore, we showed that the down-regulation of S1PR2 by LMP1 drives a signalling loop leading to constitutive activation of the phosphatidylinositol-3-kinase (PI3-K) pathway. Finally, core LMP1-PI3-K targets were enriched for lymphoma-related transcription factors and genes associated with shorter overall survival in patients with ABC-DLBCL. Our data identify a novel function for LMP1 in aggressive DLBCL. Copyright (c) 2019 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

More than half of all brain metastases show infiltrating rather than displacing growth at the macro-metastasis/brain parenchyma interface (MMPI), a finding associated with shorter survival. The Lymphoid Enhancer-binding Factor-1 (LEF1) is an epithelial-mesenchymal transition (EMT) transcription factor that is commonly overexpressed in brain-colonizing cancer cells. Here, we overexpressed LEF1 in an in vivo breast cancer brain colonization model. It shortened survival, albeit without engaging EMT at the MMPI. By differential proteome analysis, we identified a novel function of LEF1 as a regulator of the glutathione (GSH) system, the principal cellular redox buffer. LEF1 overexpression also conferred resistance against therapeutic GSH depletion during brain colonization and improved management of intracellular ROS. We conclude that besides EMT, LEF1 facilitates metastasis by improving the anti-oxidative capacity of epithelial breast cancer cells, in particular during colonization of the brain parenchyma. This article is protected by copyright. All rights reserved.

A network of autocrine and paracrine signals defines B cell homeostasis and is thought to be involved in transformation processes. Investigating interactions of these microenvironmental factors and their relation to proto-oncogenes as c-Myc (MYC) is fundamental to understand the biology of B cell lymphoma. Therefore, B cells with conditional MYC expression were stimulated with CD40L, insulin-like growth factor 1, α-IgM, Interleukin-10 (IL10) and CpG alone or in combination. The impact of forty different interventions on cell proliferation was investigated in MYC deprived cells and calculated by linear regression. Combination of CpG and IL10 led to a strong synergistic activation of cell proliferation (S-phase/doubling of total cell number) comparable to cells with high MYC expression. A synergistic up-regulation of CDK4, CDK6 and CCND3 expression by IL10 and CpG treatment was causal for this proliferative effect as shown by qRT-PCR analysis and inhibition of the CDK4/6 complex by PD0332991. Furthermore, treatment of stimulated MYC deprived cells with MLN120b, ACHP, Pyridone 6 or Ruxolitinib showed that IL10/CpG induced proliferation and CDK4 expression were JAK/STAT3 and IKK/NF-κB dependent. This was further supported by STAT3 and p65/RELA knockdown experiments, showing strongest effects on cell proliferation and CDK4 expression after double knockdown. Additionally, chromatin immunoprecipitation revealed a dual binding of STAT3 and p65 to the proximal promotor of CDK4 after IL10/CpG treatment. Therefore, the observed synergism of IL10R and TLR9 signalling was able to induce proliferation in a comparable way as aberrant MYC and might play a role in B cell homeostasis or transformation.

The interaction between vascular endothelial cells (ECs) and cancer cells is of vital importance to understand tumor dissemination. A paradigmatic cancer to study cell-cell interactions is classical Hodgkin Lymphoma (cHL) owing to its complex microenvironment. The role of the interplay between cHL and ECs remains poorly understood. Here we identify canonical WNT pathway activity as important for the mutual interactions between cHL cells and ECs. We demonstrate that local canonical WNT signaling activates cHL cell chemotaxis toward ECs, adhesion to EC layers and cell invasion using not only the Wnt-inhibitor Dickkopf, tankyrases and casein kinase 1 inhibitors but also knockdown of the lymphocyte enhancer binding-factor 1 (LEF-1) and β-catenin in cHL cells. Furthermore, LEF-1- and β-catenin-regulated cHL secretome promoted EC migration, sprouting and vascular tube formation involving vascular endothelial growth factor A (VEGF-A). Importantly, high VEGFA expression is associated with a worse overall survival of cHL patients. These findings strongly support the concept that WNTs might function as a regulator of lymphoma dissemination by affecting cHL cell chemotaxis and promoting EC behavior and thus angiogenesis through paracrine interactions.

// Alexandra Schrader 1, 2, 10, 11 , Katharina Meyer 3, 6 , Neele Walther 1 , Ailine Stolz 4 , Maren Feist 1, 7 , Elisabeth Hand 1, 6 , Frederike von Bonin 1 , Maurits Evers 3, 6, 13 , Christian Kohler 3, 6 , Katayoon Shirneshan 1 , Martina Vockerodt 5, 8, 10, 12 , Wolfram Klapper 5, 6, 7, 9 , Monika Szczepanowski 5, 6, 7, 9 , Paul G. Murray 8 , Holger Bastians 4 , Lorenz Trumper 1, 2, 5, 7 , Rainer Spang 3, 5, 6, 7 , Dieter Kube 1, 2, 5, 6, 7 1 Department of Haematology and Medical Oncology, University Medical Centre of the Georg-August University Gottingen, Gottingen, Germany 2 GRK1034 of the Deutsche Forschungsgemeinschaft, Georg-August University Gottingen, Gottingen, Germany 3 Department of Statistical Bioinformatics, Institute for Functional Genomics, University of Regensburg, Regensburg, Germany 4 Goettingen Center for Molecular Biosciences (GZMB) and University Medical Center, Institute of Molecular Oncology, Section for Cellular Oncology, Gottingen, Germany 5 Network Molecular Mechanism of Malignant Lymphoma (MMML) of the Deutsche Krebshilfe, Germany 6 BMBF-Network HamatoSys, Germany 7 BMBF-Network Myc-Sys, Germany 8 School of Cancer Sciences, University of Birmingham, Birmingham, UK 9 University-Hospital Schleswig-Holstein, Hematopathology Section and Lymph Node Registry Kiel, Kiel, Germany 10 Department of Anatomy, University Medical Centre of the Georg-August University Gottingen, Gottingen, Germany 11 Present address: Laboratory of Lymphocyte Signaling and Oncoproteome, Department I of Internal Medicine, University Hospital Cologne, Center for Integrated Oncology (CIO) Koln-Bonn, Cologne, Germany 12 Present address: Department of Anatomy, University Medical Centre of the Georg-August University Gottingen, Gottingen, Germany 13 Current address: The John Curtin School of Medical Research the Australian National University Canberra, Australia Correspondence to: Dieter Kube, e-mail: dieter.kube@med.uni-goettingen.de Keywords: lymphoma, B cell receptor signaling, guided clustering, cell cycle delay, chromosomal aberrations Received: September 23, 2015 Accepted: March 31, 2016 Published: May 7, 2016 ABSTRACT To discover new regulatory pathways in B lymphoma cells, we performed a combined analysis of experimental, clinical and global gene expression data. We identified a specific cluster of genes that was coherently expressed in primary lymphoma samples and suppressed by activation of the B cell receptor (BCR) through αIgM treatment of lymphoma cells in vitro . This gene cluster, which we called BCR.1, includes numerous cell cycle regulators. A reduced expression of BCR.1 genes after BCR activation was observed in different cell lines and also in CD10 + germinal center B cells. We found that BCR activation led to a delayed entry to and progression of mitosis and defects in metaphase. Cytogenetic changes were detected upon long-term αIgM treatment. Furthermore, an inverse correlation of BCR.1 genes with c-Myc co-regulated genes in distinct groups of lymphoma patients was observed. Finally, we showed that the BCR.1 index discriminates activated B cell-like and germinal centre B cell-like diffuse large B cell lymphoma supporting the functional relevance of this new regulatory circuit and the power of guided clustering for biomarker discovery.

Burkitt’s lymphoma (BL) and diffuse large B-cell lymphoma (DLBCL) are pathologically and clinically distinct subtypes of aggressive non-Hodgkin B-cell lymphoma. To learn more about their biology, we employed metabolomic and proteomic methods to study both established cell lines as well as cryopreserved and formalin-fixed paraffin-embedded (FFPE) tissue sections of BL and DLBCL. Strikingly, NMR analyses revealed DLBCL cell lines to produce and secrete significantly (padj = 1.72 × 10–22) more pyruvic acid than BL cell lines. This finding could be reproduced by targeted GC/MS analyses of cryopreserved tissue sections of BL and DLBCL cases. Enrichment analysis of an overlapping set of N = 2315 proteins, that had been quantified by nanoLC–SWATH-MS in BL and DLBCL cultured cells and cryosections, supported the observed difference in pyruvic acid content, as glycolysis and pyruvate metabolism were downregulated, while one-carbon metabolism was upregulated in BL compared to DLBCL. Furthermore, 92.1% of the overla...

Hodgkin's lymphoma is unusual among B cell lymphomas, in so far as the malignant Hodgkin/Reed-Sternberg (HRS) cells lack a functional B cell receptor (BCR), as well as many of the required downstream signalling components. In Epstein-Barr virus (EBV)-positive cases of Hodgkin's lymphoma, HRS cells express the viral latent membrane proteins (LMP)-1 and -2A. LMP2A is thought to contribute to the pathogenesis of Hodgkin's lymphoma by providing a surrogate BCR-like survival signal. However, LMP2A has also been shown to induce the virus-replicative cycle in B cells, an event presumably incompatible with lymphomagenesis. In an attempt to resolve this apparent paradox, we compared the transcriptional changes observed in primary HRS cells with those induced by LMP2A and by BCR activation in primary human germinal centre (GC) B cells, the presumed progenitors of HRS cells. We found a subset of genes that were up-regulated by both LMP2A expression and BCR activation but which were down-regulated in primary HRS cells. These genes included EGR1, an immediate-early gene that is required for BCR-induced entry to the virus-replicative cycle. We present data supporting a model for the pathogenesis of EBV-positive Hodgkin's lymphoma in which LMP2A-expressing HRS cells lacking BCR signalling functions cannot induce EGR1 and are consequently protected from entry to the virus lytic cycle. The primary microarray data are available from GEO (http://www.ncbi.nlm.nih.gov/geo/) under series Accession No 46143.

Burkitt's lymphoma (BL) is a highly malignant, aggressive non-Hodgkin's lymphoma derived from germinal center B cells. Recently, global gene expression profiling of patient samples led to a molecular definition of BL with lymphocyte enhancer-binding factor 1 (LEF1) as a signature gene. Herein, we report the expression of nucleic LEF1 in 15 of 18 patients with BL and the identification of LEF1 target genes. Germinal center B cells were devoid of detectable nuclear LEF1 expression, as were mantle cell lymphoma (0 of 5), marginal zone lymphoma (0 of 6), follicular lymphoma (0 of 12), and diffuse large B-cell lymphoma (1 of 31). Whole-genome gene expression profiling after transient knockdown of LEF1 in BL cell lines identified new LEF1 target genes; these LEF1 targets are enriched with genes associated with cancers. The expression of LEF1 and LEF1-regulated genes in primary BL suggests that LEF1 is not only aberrantly expressed but also transcriptionally active. This study supports a functionally important role for LEF1 and its target genes in BLs.

CME Learning Objectives Discuss whether and how genetic variations influence clinical outcomes of Hodgkin lymphoma patients. Evaluate evidence that proximal IL10 promoter gene variations are associated with clinical courses of Hodgkin lymphoma patients. Compare host genetic variations from different cytokine/cytokine receptor gene variations. Background. Hodgkin lymphoma (HL) is a lymphoid malignancy characterized by the production of various cytokines possibly involved in immune deregulation. Interleukin-10 (IL-10) serum levels have been associated with clinical outcome in patients with HL. Because host genetic variations are known to alter the expression and function of cytokines and their receptors, we investigated whether genetic variations influence clinical outcome of patients with HL. Methods. A total of 301 patients with HL who were treated within randomized trials by the German Hodgkin Study Group were included in this exploratory retrospective study. Gene variations of IL-10 (IL-10-597AC, rs1800872; IL-10-824CT, rs1800871; IL-10-1087AG, rs1800896; IL-10-3538AT, rs1800890; IL-10-6208CG, rs10494879; IL-10-6752AT, rs6676671; IL-10-7400InDel), IL-13 (IL-13-1069CT, rs1800925; IL-13Q144R, rs20541), and IL-4R (IL-4RI75V, rs1805010; IL-4RQ576R, rs1801275) were genotyped. Results. Inferior freedom from treatment failure (FFTF) was found in patients harboring the IL-10-597AA, IL-10-824TT, or the IL-10-1087AA genotype. In contrast, the IL-10-1087G-824C-597C haplotype present in about 48% of analyzed HL patients is nominally significant for a better FFTF in a Cox-Regression model accounting for stage and treatment. No associations were observed between the other IL-10 gene variations, IL-13-1069CT, IL-13Q144R, IL-4RI75V, IL-4RQ576R and the clinical outcome of patients with HL. Conclusions. Our study provides further evidence that proximal IL-10 promoter gene variations are associated with clinical course of patients with HL. However, treatment success and survival rates are already at a very high rate, supporting the need to design studies focusing on identification of predictors to reduce the side effects of therapy.

Gene expression measurements are typically performed on a fixed-weight aliquot of RNA, which assumes that the total number of transcripts per cell stays nearly constant across all conditions. In cases where this assumption does not hold (e.g., when comparing cell types with different cell sizes) the expression data provide a distorted view of cellular events. Assuming constant numbers of total transcripts, increases in expression of some RNAs must be compensated for by decreases in expression of others. Therefore, we propose calibrating gene expression data to an external reference point, the number of cells in the sample, using whole-cell spike-ins. In a systematic dilution experiment, we mixed varying numbers of human cells with fixed numbers of Drosophila melanogaster cells and scaled the expression levels of the human genes relative to those of the Drosophila genes. This approach restored the original gene expression ratios generated by the dilutions. We then used Drosophila whole-cell spike-ins to uncover non-symmetric gene expression changes, in this case much larger numbers of induced than repressed genes, under perturbations of the human cell line P493–6. Drosophila whole-cell spike-ins are an experimentally and computationally easy and low-priced method to derive mRNA fold changes of absolute abundances from RNA sequencing (RNA-Seq) and quantitative real-time PCR (qPCR) data.

A network of autocrine and paracrine signals defines B cell homeostasis and is thought to be involved in transformation processes. Investigating interactions of these microenvironmental factors and their relation to proto-oncogenes as c-Myc (MYC) is fundamental to understand the biology of B cell lymphoma. Therefore, B cells with conditional MYC expression were stimulated with CD40L, insulin-like growth factor 1, α-IgM, Interleukin-10 (IL10) and CpG alone or in combination. The impact of forty different interventions on cell proliferation was investigated in MYC deprived cells and calculated by linear regression. Combination of CpG and IL10 led to a strong synergistic activation of cell proliferation (S-phase/doubling of total cell number) comparable to cells with high MYC expression. A synergistic up-regulation of CDK4, CDK6 and CCND3 expression by IL10 and CpG treatment was causal for this proliferative effect as shown by qRT-PCR analysis and inhibition of the CDK4/6 complex by PD0332991. Furthermore, treatment of stimulated MYC deprived cells with MLN120b, ACHP, Pyridone 6 or Ruxolitinib showed that IL10/CpG induced proliferation and CDK4 expression were JAK/STAT3 and IKK/NF-κB dependent. This was further supported by STAT3 and p65/RELA knockdown experiments, showing strongest effects on cell proliferation and CDK4 expression after double knockdown. Additionally, chromatin immunoprecipitation revealed a dual binding of STAT3 and p65 to the proximal promotor of CDK4 after IL10/CpG treatment. Therefore, the observed synergism of IL10R and TLR9 signalling was able to induce proliferation in a comparable way as aberrant MYC and might play a role in B cell homeostasis or transformation.

Motivation: Understanding the structure and interplay of cellular signalling pathways is one of the great challenges in molecular biology. Boolean Networks can infer signalling networks from observations of protein activation. In situations where it is difficult to assess protein activation directly, Nested Effect Models are an alternative. They derive the network structure indirectly from downstream effects of pathway perturbations. To date, Nested Effect Models cannot resolve signalling details like the formation of signalling complexes or the activation of proteins by multiple alternative input signals. Here we introduce Boolean Nested Effect Models (B-NEM). B-NEMs combine the use of downstream effects with the higher resolution of signalling pathway structures in Boolean Networks. Results: We show that B-NEMs accurately reconstruct signal flows in simulated data. Using B-NEM we then resolve BCR signalling via PI3K and TAK1 kinases in BL2 lymphoma cell lines. Availability and implementation: R code is available at https://github.com/MartinFXP/B-NEM (github). The BCR signalling dataset is available at the GEO database (http://www.ncbi.nlm.nih.gov/geo/) through accession number GSE68761. Contact: martin-franz-xaver.pirkl@ukr.de, Rainer.Spang@ukr.de Supplementary information: Supplementary data are available at Bioinformatics online.

Gene expression profiling has recently enabled the reclassification of aggressive non-Hodgkin lymphomas (aNHL) into distinct subgroups. In Burkitt lymphoma (BL) aberrant c-Myc activity results from IG-MYC translocations. However, MYC aberrations are not limited to BLs and then have a negative prognostic impact. In this study, we investigated to which extent aberrant c-Myc activity plays a functional role in other aNHL and whether it is independent from MYC translocations. Based on a combined microarray analysis of human germinal center (GC) B cells transfected with c-Myc and 220 aNHLs cases, we developed a "c-Myc index." This index measures the extent to which lymphomas express c-Myc responsive genes. It comprises genes that are affected in a variety of tumors compared to normal tissue. This supports the view that aberrant c-Myc expression in GC B cells triggers a tumor-like expression pattern. As expected, the "c-Myc index" is very high in molecular Burkitt lymphoma (mBL), but more importantly also high within other aNHL. It constitutes a negative prognostic marker independent of established risk factors and of the presence of a MYC translocation. Our data provide new insights into the role of c-Myc activity in different lymphomas and raises the question of treatment changes for those patients under risk.

An important pathogenic event in Epstein-Barr virus (EBV)-associated lymphomas is the suppression of virus replication, which would otherwise lead to cell death. Because virus replication in B cells is intimately linked to their differentiation toward plasma cells, we asked whether the physiologic signals that drive normal B-cell differentiation are absent in EBV-transformed cells. We focused on BLIMP1α, a transcription factor that is required for plasma cell differentiation and that is inactivated in diffuse large B-cell lymphomas. We show that BLIMP1α expression is down-regulated after EBV infection of primary germinal center B cells and that the EBV oncogene, latent membrane protein-1 (LMP-1), is alone capable of inducing this down-regulation in these cells. Furthermore, the down-regulation of BLIMP1α by LMP-1 was accompanied by a partial disruption of the BLIMP1α transcriptional program, including the aberrant induction of MYC, the repression of which is required for terminal differentiation. Finally, we show that the ectopic expression of BLIMP1α in EBV-transformed cells can induce the viral lytic cycle. Our results suggest that LMP-1 expression in progenitor germinal center B cells could contribute to the pathogenesis of EBV-associated lymphomas by down-regulating BLIMP1α, in turn preventing plasma cell differentiation and induction of the viral lytic cycle.

Background Recently published data indicate that host germline variations in immune genes can influence the outcome of lymphoma patients. Interleukin (IL)-4 and IL13 are crucial immune factors and may influence the course of the disease. Both cytokines signal through the interleukin-4 receptor (IL4R). Therefore, we investigated whether polymorphisms of IL4, IL13 and IL4R genes could predict the outcome of diffuse large B-cell lymphoma (DLBCL) patients. Methods In 228 DLBCL samples of the German High-Grade Non-Hodgkin's Lymphoma Study Group, the polymorphisms of IL4 (-524CT, rs2243250), IL13 (-1069CT, rs1800925) and IL4R (I75V, rs1805010; S503P, rs1805015; Q576R, rs1801275) were analyzed and the soluble interleukin-4 receptor (sIL4R) serum level was measured before the start of chemotherapy. Results Patients harboring IL4R V75 (IL4RI75V-AG and IL4RI75V-GG) had shorter overall survival (OS) (P = 0.044) and event-free survival (EFS) (P = 0.056) periods compared with I75 carriers (IL4RI75V-AA). Multivariate analysis adjusted to the International Prognostic Index revealed a relative risk of 1.9 for carriers of the IL4R V75 (P = 0.011) in relation to OS. DLBCL patients homozygous for the IL4R I75 and low sIL4R serum levels have the most favorable OS and EFS. Conclusions These data support the role for host germline gene variations of immunologically important factors like the IL4R I75V gene variation to predict the survival in DLBCL patients.

Although the latent membrane protein-1 (LMP1) of the Epstein-Barr virus (EBV) is believed to be important for the transformation of germinal centre (GC) B cells, the precise contribution of this viral oncogene to lymphoma development is poorly understood. In this study, we used a non-viral vector-based method to express LMP1 in primary human GC B cells. Gene expression profiling revealed that LMP1 induced in GC B cells transcriptional changes characteristic of Hodgkin's lymphoma cell lines. Strikingly, LMP1 down-regulated the expression of B-cell-specific genes including B-cell receptor components such as CD79A, CD79B, CD19, CD20, CD22, and BLNK. LMP1 also induced the expression of ID2, a negative regulator of B-cell differentiation. Our data suggest that in EBV-positive cases, LMP1 is likely to be a major contributor to the altered transcriptional pattern characteristic of Hodgkin/Reed-Sternberg cells, including the loss of B-cell identity.

Purpose: Current chemotherapy can achieve high response rates in aggressive non-Hodgkin's lymphoma (NHL), but the factors that influence regression and survival remain unknown. The present exploratory study tested the hypothesis whether interleukin-10 (IL-10) polymorphisms predict clinical outcome, leukocytopenia, or infectivity during therapy. IL-10 was chosen because immune alterations are a major risk factor for NHL, and IL-10 is a cytokine involved in inflammatory processes associated with clinical outcome. Experimental Design: Five hundred patients with aggressive NHL treated with CHOP/CHOEP were analyzed for IL-10 gene polymorphisms, including distal loci -7400InDel, -6752AT (rs6676671), and -6208CG (rs10494879) in comparison with proximal loci -3538AT (rs1800890), -1087AG (rs1800896), and -597AC (rs1800872) according to the incidence and outcome of the lymphoma. Results: No differences in allele frequencies or haplotypes were found comparing a cohort of patients with aggressive NHL/diffuse large B-cell lymphoma with a healthy control group. Patients with aggressive NHL characterized by IL-10-7400DelDel had shorter overall survival periods compared with the other genotypes (P = 0.004). The 3-year rate is 43.4% for IL-10-7400DelDel and 73.4% for IL-10-7400InIn and IL-10-7400InDel together. A significant increased risk for event-free survival is found for carriers of the genotype IL-10-6752TT-6208CC-3538AA (P = 0.047). Multivariate analysis of IL-10-7400 gene variation in relation to overall survival adjusted to international prognostic index revealed a relative risk of 1.9 for carriers of IL-10-7400DelDel (P = 0.037). No associations were found analyzing diffuse large B-cell lymphoma patients separately. Conclusion: Our results indicate that IL-10 gene variations could be associated to the clinical course of aggressive NHL, which points out the importance of host factors and respective genetic elements for treatment response.

Background— Extracellular matrix proteins, such as laminins, and endothelial cells are known to influence cardiomyocyte performance; however, the underlying molecular mechanisms remain poorly understood. Methods and Results— We used a forward genetic screen in zebrafish to identify novel genes required for myocardial function and were able to identify the lost-contact ( loc ) mutant, which encodes a nonsense mutation in the integrin-linked kinase ( ilk ) gene. This loc/ilk mutant is associated with a severe defect in cardiomyocytes and endothelial cells that leads to severe myocardial dysfunction. Additional experiments revealed the epistatic regulation between laminin-α4 (Lama4), integrin, and Ilk, which led us to screen for mutations in the human ILK and LAMA4 genes in patients with severe dilated cardiomyopathy. We identified 2 novel amino acid residue–altering mutations (2828C>T [Pro943Leu] and 3217C>T [Arg1073X]) in the integrin-interacting domain of the LAMA4 gene and 1 mutation (785C>T [Ala262Val]) in the ILK gene. Biacore quantitative protein/protein interaction data, which have been used to determine the equilibrium dissociation constants, point to the loss of integrin-binding capacity in case of the Pro943Leu ( K d =5±3 μmol/L) and Arg1073X LAMA4 ( K d =1±0.2 μmol/L) mutants compared with the wild-type LAMA4 protein ( K d =440±20 nmol/L). Additional functional data point to the loss of endothelial cells in affected patients as a direct consequence of the mutant genes, which ultimately leads to heart failure. Conclusions— This is the first report on mutations in the laminin, integrin, and ILK system in human cardiomyopathy, which has consequences for endothelial cells as well as for cardiomyocytes, thus providing a new genetic basis for dilated cardiomyopathy in humans.

The Interleukin 10 (IL-10) gene is highly polymorphic, and the IL-10(-1087AG) (rs1800896) gene variation is the only so far studied intensively in association with certain diseases. Conflicting data have been published about an association of IL-10(-1087AG) gene variation with lower rates of complete remission and lower overall survival (OS) in patients with diffuse large B-cell lymphoma. To further investigate this in malignant lymphoma, we established the IL-10 genotypes in patients from the NHL-B1/ B2 studies from the German High-Grade Non-Hodgkin's Lymphoma Study Group. In our study, allele frequencies of lymphoma patients are comparable as in healthy controls. No increase of IL-10(-1087G) alleles was found. In addition we did not find any difference in OS or event-free survival between patients with IL-10(-1087AA) and the other genotypes. Comparable results were obtained for the IL-10 loci at -3538 (A/T), -1354 (A/G), -824 (C/T) and -597 (A/C) (rs1800890, rs1800893, rs1800871 and rs1800872).

Classical Hodgkin lymphoma (cHL) has a typical clinical manifestation, with dissemination involving functionally neighboring lymph nodes. The factors involved in the spread of lymphoma cells are poorly understood. Here we show that cHL cell lines migrate with higher rates compared with non-Hodgkin lymphoma cell lines. cHL cell migration, invasion and adhesion depend on autocrine WNT signaling as revealed by the inhibition of WNT secretion with the porcupine inhibitors Wnt-C59/IWP-2, but did not affect cell proliferation. While application of recombinant WNT5A or WNT5A overexpression stimulates HL cell migration, neither WNT10A, WNT10B nor WNT16 did so. Time-lapse studies revealed an amoeboid type of cell migration modulated by WNT5A. Reduced migration distances and velocity of cHL cells, as well as altered movement patterns, were observed using porcupine inhibitor or WNT5A antagonist. Knockdown of Frizzled5 and Dishevelled3 disrupted the WNT5A-mediated RHOA activation and cell migration. Overexpression of DVL3-K435M or inhibition of ROCK (Rho-associated protein kinase) by Y-27632/H1152P disrupted cHL cell migration. In addition to these mechanistic insights into the role of WNT5A in vitro, global gene expression data revealed an increased WNT5A expression in primary HL cells in comparison with normal B-cell subsets and other lymphomas. Furthermore, the activity of both porcupine and WNT5A in cHL cells had an impact on lymphoma development in the chick chorionallantoic membrane assay. Massive bleeding of these lymphomas was significantly reduced after inhibition of WNT secretion by Wnt-C59. Therefore, a model is proposed where WNT signaling has an important role in regulating tumor-promoting processes.

Item does not contain fulltext Epstein-Barr virus (EBV) and Plasmodium falciparum have overlapping distributions and are thought to have causal interactions, particularly with regard to the aetiology of endemic Burkitt's lymphoma. Using real-time PCR, we quantified and compared EBV DNA levels in the blood before and after antimalarial treatment of age- and gender-matched groups of Gabonese children who presented with either mild or severe P. falciparum malaria. Following treatment, the prevalence of EBV DNA declined in the mild malaria group but increased in the severe malaria group, and a significantly higher proportion of the latter had EBV DNA detectable in their blood when they were healthy and parasite free (67% vs. 39%; P=0.013). High EBV DNA loads were associated with more malaria attacks and with elevated plasma concentrations of both TNF-alpha and IL-12p40. Significantly more under 5 year olds had EBV DNA, highlighting the strong age dependence of the interaction between the two pathogens. These findings confirm that EBV is reactivated during acute P. falciparum malaria but, importantly, also reveal that: (i) EBV activity persists at a higher frequency in children with a history of severe malaria; and (ii) higher peripheral blood EBV DNA loads are associated with susceptibility to more frequent P. falciparum episodes and with altered cytokine activity.

CD4 + T cells classically recognize antigens that are endocytosed and processed in lysosomes for presentation on major histocompatibility complex (MHC) class II molecules. Here, endogenous Epstein-Barr virus nuclear antigen 1 (EBNA1) was found to gain access to this pathway by autophagy. On inhibition of lysosomal acidification, EBNA1, the dominant CD4 + T cell antigen of latent Epstein-Barr virus infection, slowly accumulated in cytosolic autophagosomes. In addition, inhibition of autophagy decreased recognition by EBNA1-specific CD4 + T cell clones. Thus, lysosomal processing after autophagy may contribute to MHC class II–restricted surveillance of long-lived endogenous antigens including nuclear proteins relevant to disease.

Item does not contain fulltext Interleukin-10 (IL-10), a cytokine involved in many aspects of the immune response shows interindividual variations in their expression. However, genetic variations of the 5'-flanking region of the IL-10 gene (PIL-10) are poorly characterised with respect to different stimuli. New extended haplo- and genotypes are identified present at differing frequencies in three geographically separated populations. Their influence on IL-10 expression have been assessed in vitro after stimulation of leukocytes with lipopolysaccharide (LPS), dibutyryl-cAMP or following immortalisation with Epstein-Barr virus (lymphoblastoid cell line (LCL)). Interindividual differences of IL-10 production were found to be related to single-nucleotide polymorphisms (SNP) haplotype -6752/-6208 in LCLs (P

Burkitt lymphoma is a rare malignancy arising from B cells. Current chemotherapeutic regimens achieve excellent overall survival rates in children, but less impressive rates in adults. There are cases with poor outcome caused by toxic effects of the therapy, tumor lysis syndrome, or metastatic spread of lymphomas to the central nervous system. Modulators of reactive oxygen species are currently discussed as potential drugs for the treatment of cancer. The NADPH oxidase 4 inhibitor imipramine-blue might satisfy the aforementioned requirements, and was studied here. We used MTT assay, crystal violet assay, and thymidine 3H-incorporation assay to analyze the effects of imipramine-blue on Burkitt lymphoma (BL2, BL2B95, BL30B95, BL41B95), neuroblastoma (KELLY, SH-SY5Y, SMS-KAN), cervix carcinoma (HeLa), breast cancer (MDA-MB231), angiosarcoma (AS-M), human embryonic kidney (HEK293WT), and nonmalignant (FLP1) cell lines. The effects of imipramine-blue on BL2B95 cells in vivo were investigated in xenografts on the chick chorioallantoic membrane (CAM). We report that imipramine-blue is a potent growth inhibitor for several cancer cell lines in vitro with IC50 values comparable to those of doxorubicin (0.16–7.7 μmol/L). Tumor size of BL2B95 cells inoculated in the CAM was reduced significantly (P < 0.05) after treatment with 10 μmol/L imipramine-blue. Lymphogenic dissemination of BL2B95 and the formation of blood and lymphatic vessels in experimental tumors were not affected. We show that imipramine-blue can be used to decrease the viability of cancer cell lines in vitro and in vivo. Imipramine-blue reduces the size of experimental Burkitt lymphoma significantly but does not affect the dissemination of BL2B95 cells, angiogenesis, and lymphangiogenesis. Mol Cancer Ther; 13(4); 833–41. ©2014 AACR.

Epstein-Barr virus (EBV) is associated with several human malignancies including Burkitt's lymphoma (BL), Hodgkin's disease (HD) and nasopharyngeal carcinoma. A variety of cytokines and receptors have been described to be activated by EBV. Here we show that the IL-2 receptor (IL-2R) alpha-chain, which is weakly expressed on normal resting lymphoid cells, is activated by EBV. Comparison of EBV-negative BL cell lines and their EBV convertants showed an enhanced CD25 expression in EBV-positive BL cells. Transient expression of the oncogenic virus protein latent membrane protein-1 (LMP1) in L428 Hodgkin's lymphoma cells and in Burkitt's lymphoma cells (BL2, BL41, BL30) cells leads to enhanced CD25 expression. Both C-terminal activating regions (CTARs) of LMP1 are involved in CD25 activation. Inhibition of LMP1-mediated NFkappaB enhancement by a constitutive repressive form of IkappaB-alpha resulted in decreased CD25 surface expression, indicating that NFkappaB is involved in CD25 gene regulation. Furthermore, LMP1-mediated CD25 activation was associated with enhanced levels of the soluble form of CD25 (sCD25) in L428 Hodgkin's lymphoma cells but not in BL cells. LMP1 associated enhanced expression of membrane CD25 and soluble CD25 may have immunomodulatory functions and could be involved in biology of EBV-associated diseases.