Identification of Somatically Acquired Gene Mutations In the Kineome and Exome In CLL

Abstract 2410

The pathogenesis of CLL remains incompletely understood. While acquired chromosomal aberrations have been demonstrated to influence CLL biology and clinical behavior, it remains unclear what single gene defects other than p53 or ATM mutations contribute to or cause the CLL phenotype. In particular, recurrent gene mutations that are increasingly found in other hematological malignancies have not yet been identified in CLL. Further, the discovery of stereotypical immunoglobulin receptor genes in CLL has been advanced in support of the involvement of autoantigenes in the pathogenesis of CLL and thus a pro-proliferative or anti-apoptotic gene mutation may possibly not exist in CLL.

Given that kinase genes are often the targets for recurrent gene mutations and given the prospect for identification of drugable targets, we have analyzed 515 kinase genes in a discovery panel of 23 well characterized CLL samples (in DNA from CD19+ sorted cells) and tested their somatic nature in CD3+ cell-derived DNA from the same patients. A total of 8308 different exon sequences were analyzed. We identified six individual kinase gene mutations (WEE1, NEK1, BRAF, KDR, MAP4K3 and TRPM6), each occurring once in a separate CLL case as well as >1450 non-synonymous polymorphisms but no recurrent kinase gene mutations were identified. Interestingly, mutations in all these genes but NEK1 had previously been identified in cancer genome projects of cancers of the breast, colorectum, pancreas and glioblastome multiforme and WEE1, BRAF and KDR have established roles in cancer biology.

Next we used solution exon capture of genomic DNA followed by Solexa-based next gen exome sequencing on 8 paired CLL derived DNA samples (FACS-sorted CD19+ versus CD3+ paired samples), including 2 samples with known p53 mutations. Using stringent criteria for mutation calling, we identified 69 mutated genes including the 2 cases of p53 mutations. None of the mutated genes were recurrent. Work is in progress to estimate the frequency of mutations in these genes in a wider panel of CLL cases.

In summary, our data provide practical information about the mutations state of the CLL kineome and exome with implications for CLL biology/pathogenesis. This data should motivate additional exome sequence analyses of more CLL samples to fully capture and describe the mutation load in CLL.