Discovery of recurrent t(6;7)(p25.3;q32.3) translocations in ALK-negative anaplastic large cell lymphomas by massively parallel genomic sequencing

Abstract

The genetics of peripheral T-cell lymphomas are poorly understood. The most well-characterized abnormalities are translocations involving ALK, occurring in approximately half of anaplastic large cell lymphomas (ALCLs). To gain insight into the genetics of ALCLs lacking ALK translocations, we combined mate-pair DNA library construction, massively parallel ("Next Generation") sequencing, and a novel bioinformatic algorithm. We identified a balanced translocation disrupting the DUSP22 phosphatase gene on 6p25.3 and adjoining the FRA7H fragile site on 7q32.3 in a systemic ALK-negative ALCL. Using fluorescence in situ hybridization, we demonstrated that the t(6;7)(p25.3;q32.3) was recurrent in ALKnegative ALCLs. Furthermore, t(6;7)(p25.3; q32.3) was associated with down-regulation of DUSP22 and up-regulation of MIR29 microRNAs on 7q32.3. These findings represent the first recurrent translocation reported in ALK-negative ALCL and highlight the utility of massively parallel genomic sequencing to discover novel translocations in lymphoma and other cancers. IntroductionRecurrent chromosomal translocations are common pathogenetic events in hematologic malignancies. 1 Among peripheral (postthymic) T-cell lymphomas, however, the only well-characterized translocations are those involving the anaplastic lymphoma kinase gene ALK. 2 ALK is an important prognostic marker and therapeutic target in T-cell anaplastic large cell lymphomas (ALCLs) 3,4 ; however, approximately half of ALCLs lack ALK expression, despite nearly identical morphology and phenotype. 5 ALKnegative ALCLs can occur either cutaneously or systemically. We previously identified recurrent IRF4 translocations in cutaneous ALCLs, 6 but recurrent translocations in the more lethal, systemic form of ALK-negative ALCL have not been reported.Massively parallel ("Next Generation") DNA sequencing technology represents a quantum advance in the ability to understand cancer genomes. To identify translocations in ALK-negative ALCL, we performed massively parallel sequencing of a mate-pair DNA library constructed from a systemic ALK-negative ALCL. Using a unique bioinformatic algorithm for translocation discovery, we identified a translocation, t(6;7)(p25.3;q32.3), and demonstrated this translocation in additional ALK-negative ALCLs. This represents the first recurrent translocation reported in systemic ALKnegative ALCL, and demonstrates the utility of mate-pair library sequencing as a tool for translocation discovery. MethodsBriefly, mate-pair library construction followed the manufacturer's protocol (Illumina) using approximately 5-kb genomic DNA fragments. Sequencing was performed on an Illumina GAIIx, and results were mapped to the genome using a binary indexing algorithm. 7 Candidate translocations were validated by polymerase chain reaction (PCR), Sanger sequencing, and fluorescence in situ hybridization (FISH). Gene and microRNA expression levels were assessed using quantitative real-time PCR. The study was approved by the Mayo Clinic Institutional Review Board. Detail...

Mucosal CD30-positive T-cell lymphoproliferations of the head and neck show a clinicopathologic spectrum similar to cutaneous CD30-positive T-cell lymphoproliferative disorders

“…Rearrangements of 6p25.3 are recurrent in ALK-negative anaplastic large cell lymphomas and are most common in primary cutaneous anaplastic large cell lymphoma, where they are seen in about 28% of cases. 8,[23][24][25] This finding is an additional feature shared between patients with primary mucosal CD30-positive T-cell lymphoproliferations and primary cutaneous cases. A 6p25.3 rearrangement also was seen in a patient with orbital disease (case 15) but unfortunately staging data were not available.…”

Targeted next generation sequencing of clinically significant gene mutations and translocations in leukemia

“…35 Therefore, even with sufficient coverage this method may not be capable of identifying all translocation events, and additional methodologies such as the inclusion of mate pair libraries may be required. 36 Unlike current methods employed in the clinical molecular oncology laboratory, target-capture-based next generation sequencing offers greatly increased scalability, requires less technician labor, and is becoming less expensive. For example, mutations in many genes recently implicated in leukemia prognosis, such as CEBPA and DMNT3A occur throughout the coding region, and require sequencing of multiple PCR products and/or exons for full evaluation.…”

Aspects moléculaires des lymphomes T périphériques (1)

“…Le facteur de transcription IRF4/MUM1 est connu pour être un important régulateur de la différencia-tion lymphoïde normale, fortement exprimé dans les plasmocytes et les lymphocytes T activés [72]. Dans la majorité des translocations impliquant IRF4, le partenaire n'est pas identifié et l'expression d'IRF4 ne semble pas dérégulée [73]. En revanche, quand le gène affecté en 6p25 est DUSP22, son partenaire se situe dans plus de la moitié des cas en 7q32.3, à proximité du site D'autres partenaires d'ALK ont par la suite été rapportés [15,17,55] (Tableau III).…”