Characterizing Translocations Involving the DUSP22-IRF4 Gene Region in ALK-Negative Anaplastic Large Cell Lymphomas Using Next Generation Sequencing.
Andrew L Feldman, Ahmet Dogan, David I Smith, Mark E Law, Stephen M Ansell, Sarah H Johnson, Julie C Porcher, Nazan Ozsan, Eric D Wieben, Bruce W Eckloff, George Vasmatzis. Mayo Clinic, Rochester; Ege University, Izmir, Turkey
Background: We recently described novel translocations involving the IRF4 region on 6p25.3 in ALK-negative ALCLs (cutaneous and occasionally systemic). To characterize the breakpoints and partner loci in these translocations, we utilized Next Generation sequencing of mate-pair libraries prepared from genomic DNA. Mate-pair library preparation juxtaposes DNA fragments originating ∼5000 bp apart; this allows coverage of the entire genome to identify translocations at a fraction of the cost of whole genome sequencing.
Design: Mate-pair libraries from 4 ALK-negative ALCLs (3 cutaneous, 1 systemic) each were applied to one lane of an Illumina flow cell and sequenced on an Illumina GAIIx. Sequence data were mapped to the genome using a binary indexing algorithm and analyzed for >2 non-identical mate-pairs in which one end mapped to 6p25.3 and the other mapped to a distant locus. PCR products using primers flanking the breakpoints were Sanger sequenced. Fluorescence in situ hybridization (FISH) was performed using home-brew probes. RNA was quantitated using real-time PCR.
Results: Two cases had t(6;7)(p25.3;q32.3) with breakpoints disrupting DUSP22 (telomeric to IRF4 on 6p25.3) and telomeric to microRNAs 29A/B on 7q32.3. One had der(6)t(6;9)(p25.3;p24.3) with breakpoints between DUSP22 and IRF4 (consistent with loss of the telomeric DUSP22 fragment) and telomeric to SMARCA2 on 9p24.3. One case had a complex t(Y;6)(q11.221;p25.3) with two 6p25.3 breakpoints flanking DUSP22 (consistent with DUSP22 deletion). t(6;7) and t(6;9) were confirmed by Sanger sequencing. FISH for t(6;7) was positive in 13/29 (6 cutaneous, 7 systemic) ALCLs with 6p25.3 translocations. 6p25.3 translocations were associated with 50-fold reduction in DUSP22 expression (p=0.002, t-test) but no change in IRF4 expression. Cases with 7q32.3 breakpoints showed 5-fold up-regulation of MIR29B (p=0.007).
Conclusions: Translocations involving 6p25.3 in ALK-negative ALCLs lead to disruption or deletion of DUSP22. Down-regulation of DUSP22 expression, rather than IRF4 dysregulation, appears to be the common feature of these tumors. DUSP22 encodes a dual-specificity phosphatase that inhibits T-cell signaling, and may represent a novel tumor suppressor gene. Other biologic effects may derive from the partner loci, of which 7q32.3 is most common. These include overexpression of MIR29B and perhaps dysregulation of SMARCA2, which is amplified in some B-cell lymphomas.
Wednesday, March 2, 2011 9:30 AM
Poster Session V # 150, Wednesday Morning