Arthur Edwards Broad Summer Research Program in Genomics Cancer Program 08/06/07 Genome-wide miRNA Expression Analysis in Lymphoma miRNAs Lymphoma

Why Lymphoma? Source: American Cancer Society, Men 38,670 Women 32,610 Deaths Cases Men 10,370 Women 9,360 Lymphoma Hodgkin’s lymphoma Hodgkin’s lymphoma Non-Hodgkin’s lymphoma Non-Hodgkin’s lymphoma 7,350 patients (est. 2007) 56,390 patients (est. 2007) Adapted from J.L Peck, SI /01/06 – Stays at MGH 07/23/07 – Plays at Fenway About 50% not so happy ending

Naive B-cells Antigen Memory B-cells Plasma cells Centroblasts Centrocytes Diffuse Large B-Cell Lymphoma (DLBCL) DLBCL is a result of abnormal B-cell development. Mediastinal Large B-Cell Lymphoma (MLBCL) – Subtype of DLBCL with worse prognosis than DLBCL. Significant clinical and genetic heterogeneity.

Challenges to the Lymphoma Field 1.Lack of understanding of its molecular mechanisms 2.Lack of diagnostic / prognostic tools 3.Lack of effective therapeutic drugs

miRNAs The start of something new: lin-4 and let-7 found in C.Elegans Cell Death Cell proliferation Differentiation Physiological Responses Expression Profiles Contain Diagnostic Information miRNAs as regulators of gene expression and biology Some miRNAs play functional roles in tumorgenesis particularly in lymphoma “miR-17 cluster cooperates with c-myc in lymphomagenesis” Lu et. al. (2005) – NatureHe et. al. (2005) – NatureLee et. al. (1993) – CellReinhart et. al. (2000) - Nature

Controlsn = Cell Line (HeLa) 9 Cell Line (MCF-7)9 Cell Line (MOLT-4)6 Blank Controls22 Lymphoma Samples Approach: genome-wide miRNA expression analysis Diagnosisn = Lymphoma Cell Line34 Paired DLBCL18 Primary MLBCL39 Primary DLBCL139 Normal B-Cell12 242

Overview of miRNA Profiling

Fig 1a. Plot of cell line controls and their relative expression values. Quality Control of Technical Procedure Fig 1b. Cell line controls cluster. Molt4 MCF-7 HeLa Plate 1 Plate 2 Plate 3 Code Category

Fig 2. Clustering of Normal B-Cell Samples miRNA Expression Profiles Differentiate B-Cell Development Naïve Centroblast Centrocyte Memory Code Category 121

miRNA Expression Differentiates Primary Lymphoma Samples Primary DLBCL Primary MLBCL Normal B-Cell Naïve Centroblast Memory Centrocyte miR-17 cluster Fig 3. Primary Lymphoma Samples Clustered Code Category miR-150

Non-Random Association of Patient Outcome Fig 4a. DLBCL vs. MLBCL MLBCL DLBCL Failure Cure No Data Code Category 12 Cluster 1 Cluster 2 Failure20 40 Cure p=0.018 Fisher’s Exact Test Fig 4b. Correlation between Cure and Failure

Fig 5. Lymphoma Cell Lines Clustered Lymphoma Cell Line Comparison Hodgkin’s Lymphoma DLBCL Mantle Cell Lymphoma MLBCL miR-155

Conclusions Classifying Primary Lymphoma Samples. CB / CC – like lymphoma. Naïve / memory - like lymphoma Another class miRNAs and prognosis. miRNAs may give clues on molecular signatures in prognosis of lymphoma patients. Lymphoma Cell Lines Hodgkin’s Lymphoma have a different miRNA expression pattern B-Cell Development Two distinct patterns of miRNA expression. CB / CC vs. Naïve / Memory

Unraveling the functional aspect of miRNAs in B-Cell development and in lymphomagenesis. More comprehensive data analysis with additional clinical information. Integrative data analysis (Affy data, miRNA data, SNP data and sequence data) Future Studies

Acknowledgements Broad Members –Jun Lu –Hao Zhang –Stefano Monti –Alper Uzun –Yuliya Kodysh –Arthur Liberzon –Xiaohui Xie –Jill Mesirov –Todd Golub Broad Summer Research Program in Genomics –Shawna Young –Bruce Birren –Maura Silverstein –Scott Breiding DFCI / Harvard Medical –Kunihiko Takeyama –Margaret Shipp MIR WAY