Project Abstract Non-Hodgkin lymphomas (NHLs) of B-cell type, a heterogenous group of lymphoid malignancies, are among the most common cancers worldwide, accounting for about 4% of all cancers. A dramatic rise in incidence of NHLs worldwide during the past decades has sparked intense research efforts to understand their pathogenesis. Genomics studies have uncovered many novel genomic alterations in NHLs, but these remain to be functionally validated and characterized. Among the most common of the genomic alterations is a missense mutation that results in leucine-to-proline substitution at position 265 in MYD88 (MYD88L265P), an adaptor protein that activates oncogenic NF-κB signaling. The MYD88L265P mutation is exceptionally frequent in lymphoplasmacytic lymphoma (LPL) and activated B-cell type of diffuse large B-cell lymphoma (ABC-DLBCL). While inhibition of MYD88L265P adversely impacts the survival of LPL and ABC-DLBCL cells, its role in lymphoma initiation remains to be clarified. Therefore, to elucidate the lymphomagenic potential of MYD88L265P we generated conditional transgenic mice overexpressing human wild-type (hMYD88WT) or mutant (hMYD88L265P) proteins in activated B-cells. Although abundance of both proteins and p65 NF-κB nuclear translocation was increased in transgenic GC B- cells, we observed that: (i) the MYD88L265P protein differed from the MYD88WT in its stability, ease of aggregation, and downstream activity; (ii) hMYD88WT did not produce detectable phenotypic alterations, but hMYD88L265P promoted with high frequency and long latency, a non-clonal, low-grade B-cell lymphoproliferative disorder resembling human LPL, which occasionally underwent transformation to ABC-DLBCL, suggesting that MYD88L265P is insufficient by itself to drive neoplastic transformation of mature B-cells, and that secondary cooperating genetic alterations are needed. In line with our findings, introduction of MYD88L265P into primary B- cells was recently shown to induce negative feedback mechanisms mediated by TNFAIP3, a negative regulator of NF-κB pathway residing on Chr6q, along with other important tumor suppressors. Notably, Chr6q deletions are observed in almost half of LPL cases with small somatic deletions present in up to 80% of patients with MYD88L265Pmutation and in ABC-DLBCL. Importantly, Chr6q losses are not detected in human MYD88WT LPL patients, indicating that repression of 6q-related signaling is a critical pathogenetic step specifically in MYD88L265P-induced LPL. These results indicate that MYD88L265P possesses unique biochemical and functional properties, and suggest that the hMYD88WT and hMYD88L265P transgenic mice constitute an ideal model system in which to investigate these properties, as well as the secondary cooperating genetic alterations that are necessary to fully develop a clonal LPL phenotype and its eventual progression to ABC-DLBCL. Here we propose to investigate the role of the MYD88L265P mutation, Chr6q deletion (Chr10q in mice) as well as other LPL- associated loss-of-function gene mutations in B-cell development and function as well as the pathogenesis of LPL and ABC-DLBCL, and to develop a preclinical mouse models of LPL and ABC-DLBCL for testing therapies.

Project Narrative Recent genomic studies have uncovered many novel putative oncogenes and tumor suppressors of B-cell lymphomas, but these remain to be functionally validated and characterized. Among the most common and co- occurring genomic alterations is a mutation in the MYD88 gene (MYD88L265P), and a large deletion of chromosome 6 (Chr6q deletion), which are exceptionally frequent among two important subtypes of B-cell lymphomas, lymphoplasmacytic lymphoma (LPL) and diffuse large B-cell lymphoma of activated B-cell type (ABC-DLBCL). Therefore, the studies proposed here to study the molecular mechanisms by which MYD88L265P mutation, Chr6q deletion and other LPL/ABC-DLBCL-associated genomic alterations induce lymphomagenesis will significantly move the field forward to provide discoveries which improve diagnosis, evaluate prognosis and lead to the development of novel and more effective therapies as well as cellular and organismal models to treat these and other types of B-cell lymphomas.