Genomic alterations and RNA/protein expression profiling

This workpackage includes different groups with complementary areas of expertise and activities covering all aspects of the project. In previous join studies these groups have analysed the primary genetic alterations, cell cycle deregulation mechanisms, and secondary genetic alterations in MCL. These investigations have identified several genetic and molecular targets that play a crucial role in the pathogenesis of MCL and may be used as prognostic factors. In the current project the efforts will be devoted to translate the information generated by global genomic and proteomic studies into the clinical practice by investigating clinical samples of study patients of the European MCL Network. The groups will work in an integrated network sharing samples and results that will allow building and testing different predictive models based on genomic alterations and expression profile signatures. Thus, The Nijmegen group will analyze the genomic alterations of MCL using matrix CGH. These results will be compared with microarrays expression data generated in Barcelona and Würzburg whereas the Ulm group will address the analysis and validation of potential target genes by real time quantitative PCR. The Barcelona, Würzburg, and Ulm groups will design predictive models based on small group of genes to be used by RQ-PCR and immunohistochemistry in clinical samples with limited amount of material. This small panel of genes will be selected from the results of the microarrays studies. The Hannover and Munich groups will develop different proteomics strategies to identify potential proteins of interest, both in tumour and blood samples of the patients that may be of help as new targets for monitorizing the progression of the disease and development of therapeutic strategies.

The different partners and activities of this workpackage are:

Ludwig Maximilian University, Munich, Germany

Activity: Proteomics, two-D Gels
M. Dreyling is assistant professor at the Dept. of Medicine III/university hospital Grosshadern/LMU. Besides his clinical responsibilities (assistant coordinator of the German Low Grade Lymphoma Study Group, project coordinator of the national competence network „malignant lymphomas“, co-coordinator of the clinical European MCL Network), his special research interests are the roles of cell cycle dysregulation and secondary molecular alterations as biological prognostic factors in mantle cell lymphoma.

Pathologie des Cellules Lymphoïdes, Université Claude Bernard Lyon-1, Lyon

Activities: comparative microarray analysis
(see WP III/1)

Hospital Clinic, University of Barcelona, Barcelona, Spain

Activities: Microarray Analysis, RQ-PCR
The Hematopathology Unit of the Hospital Clinic, University of Barcelona has a long experience in all aspects of diagnostic haematopathology including morphology, immunophenotyping, cytogenetics, and molecular biology. The research activity of the group during the last 10 years has concentrated on the genetic and molecular mechanisms involved in the pathogenesis of malignant lymphomas. In these fields, our group pioneered the recognition of Cyclin D1 as a specific alteration of MCL as well as the involvement of genetic alterations in different cell cycle regulators and DNA-damage response genes in the progression of this disease. The group participates actively in different international research networks including the European MCL Research Network since 1997 and the Leukemia/Lymphoma Molecular Profiling Project.

Institute of Pathology, University of Würzburg, Würzburg, Germany

Activities: Microarray analysis, tissue microarrays, immunohistochemistry
German Ott was trained as a haematopathologist at the Institute of Pathology, University of Würzburg/Germany (Prof. Dr. H.K. Müller-Hermelink). Over the past 10 years, his he has made important scientific contributions in the biological and genetic characterization of MALT-type lymphomas, follicular lymphomas, diffuse large B-cell lymphomas and mantle cell lymphomas. He currently is one of the reference pathologists in the European MCL Research Network.
Andreas Rosenwald is a resident in haematopathology at the Institute of Pathology, University of Würzburg/Germany. From 1999-2003, he worked as a research fellow in the laboratory of Dr. Louis Staudt at the National Cancer Institute, Bethesda, USA, where he used gene expression profiling (Lymphochip microarray technology) to work towards a molecular classification of B-cell Non-Hodgkin lymphomas and to predict their clinical outcome. Recently, Dr. Rosenwald has been appointed principal investigator of a research group at the University of Würzburg.
Both group leaders are active participants in the international Leukaemia/Lymphoma Molecular Profiling Project (LLMPP).

MHH, Hannover, Germany

Activity: Proteomics, SELDI
B Schlegelberger, M.D. is head of the Institute of Cell and Molecular Pathology, Hanover Medical School (MHH), Hannover, Germany. During the last 15 years, she has focussed on the cytogenetic characterization of malignant lymphomas, on cloning of genes involved in lymphoma-genesis, on the development of FISH assays, on the combination of immunophenotyping and FISH (FICTION) and on the delineation of deleted regions, suspected to contain tumour suppressor genes.

Department of Internal medicine III, University of Ulm, Germany

Activity: Genetic alterations and Expression Profile: RQ-PCR
S. Stilgenbauer has a longstanding experience is in the field of biological and clinical studies of haematopoietic tumours and is coordinator of the central reference laboratory of the German CLL Study Group (GCLLSG) and of several multicentre treatment trials on haematopoietic tumours.

Important achievements of the group within the last few years have been:

University Medical Center, Nijmegen, The Netherlands

Activity: Matrix CGH
J van Krieken is professor of tumour pathology at the department of Pathology, UMC Nijmegen the Netherlands. He has been trained as a pathologist with subspecialisation into hematopathology in Kiel/Germany (Prof Lennert) and Bethesda/USA (E. Jaffe). During the last 10 years, he has focussed on the genetic and immunological characterization of malignant lymphomas, on clinicopathologic studies, and on the development of new diagnostic tools, including molecular diagnostics.

Objectives:
Based on the recent development of innovative molecular techniques (matrix CGH, RNA array chips, RQ-PCR, proteomics), we propose a multimodal approach to evaluate the predictive value of large scale analysis of genomic alterations and gene expression profiles at mRNA and protein levels in the context of controlled clinical trials of the European MCL Network:
RNA array/RQ-PCR analysis


Proteomics
Secondary molecular alterations are known to be responsible for the progression and clinical course of malignant lymphomas. However, besides the well established role of p53 and p16 alterations in the secondary transformation to high malignant lymphoma, only little is known about such secondary alterations in MCL. Thus, aim of this project is the characterization of secondary prognostic markers based on 2 dimensional protein electrophoreses and subsequent identification of differentially expressed proteins (MALDI-TOF). At the same time, the innovative approaches of matrix CGH and proteome analysis (MALDI-TOF, SELDI) will be adapted to the analysis of the clinical patient samples. Results will be compared with the simultaneously performed RNA array analysis.