To characterise the genetics of splenic marginal zone lymphoma (SMZL), we performed whole exome sequencing of 16 cases and identified novel recurrent inactivating mutations in Kruppel-like factor 2 (KLF2), a gene whose deficiency was previously shown to cause splenic marginal zone hyperplasia in mice. KLF2 mutation was found in 40 (42%) of 96 SMZLs, but rarely in other B-cell lymphomas. The majority of KLF2 mutations were frameshift indels or nonsense changes, with missense mutations clustered in the C-terminal zinc finger domains. Functional assays showed that these mutations inactivated the ability of KLF2 to suppress NF-κB activation by TLR, BCR, BAFFR and TNFR signalling. Further extensive investigations revealed common and distinct genetic changes between SMZL with and without KLF2 mutation. IGHV1-2 rearrangement and 7q deletion were primarily seen in SMZL with KLF2 mutation, while MYD88 and TP53 mutations were nearly exclusively found in those without KLF2 mutation. NOTCH2, TRAF3, TNFAIP3 and CARD11 mutations were observed in SMZL both with and without KLF2 mutation. Taken together, KLF2 mutation is the most common genetic change in SMZL and identifies a subset with a distinct genotype characterised by multi-genetic changes. These different genetic changes may deregulate various signalling pathways and generate cooperative oncogenic properties, thereby contributing to lymphomagenesis.

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Purpose: Diffuse large B-cell lymphoma (DLBCL) is a heterogeneous group of aggressive lymphomas with the activated B-cell-like subtype characterized by constitutive NF-kB activation. Activating mutations of CARD11 and inactivating mutations of A20 are frequent events in DLBCL. However, the full extent of genetic alterations in the NF-kB pathway regulators and their potential prognostic value in DLBCL remain to be investigated. We investigated the genetic abnormalities of CARD11, A20, and ABIN-1/2/3 (the A20 binding inhibitor of NF-kB) and their clinicopathologic correlation in gastrointestinal DLBCL.Experimental Design: The somatic mutation and copy number changes of CARD11, A20, and ABIN-1/ 2/3 were investigated in 71 gastrointestinal DLBCLs by PCR/sequencing, and interphase FISH/array comparative genomic hybridization, respectively. The mutations identified were functionally characterized by NF-kB reporter assays and immunoprecipitation experiments.Results: Recurrent somatic mutations were found in CARD11 (10%), A20 (17%), ABIN-1 (4%), and ABIN-2 (3%), but not in ABIN-3. In comparison with the wild-type, all CARD11 mutants were potent NF-kB activators in vitro. On the basis of the destructive nature of the observed mutations, and the findings by reporter assays and immunoprecipitation studies, most if not all of the somatic mutations that were seen in A20, ABIN-1, and ABIN-2 could impair their normal functions. Among these genetic abnormalities, A20 somatic mutation was significantly associated with both poor overall survival and event-free survival.Conclusions: We show further evidence of NF-kB pathway genetic abnormalities in DLBCL, which are potentially valuable in the prognosis and design of future therapeutic strategies.