Exome sequencing identifies recurrent BCOR alterations and the absence of KLF2, TNFAIP3 and MYD88 mutations in splenic diffuse red pulp small B-cell lymphoma

Abstract

Splenic diffuse red pulp lymphoma is an indolent small B-cell lymphoma recognized as a provisional entity in the World Health Organization 2008 classification. Its precise relationship to other related splenic B-cell lymphomas with frequent leukemic involvement or other lymphoproliferative disorders remains undetermined. We performed whole-exome sequencing to explore the genetic landscape of ten cases of splenic diffuse red pulp lymphoma using paired tumor and normal samples. A selection of 109 somatic mutations was then evaluated in a cohort including 42 samples of splenic diffuse red pulp lymphoma and compared to those identified in 46 samples of splenic marginal zone lymphoma and eight samples of hairy-cell leukemia. Recurrent mutations or losses in BCOR (the gene encoding the BCL6 corepressor) - frameshift (n=3), nonsense (n=2), splicing site (n=1), and copy number loss (n=4) - were identified in 10/42 samples of splenic diffuse red pulp lymphoma (24%), whereas only one frameshift mutation was identified in 46 cases of splenic marginal zone lymphoma (2%). Inversely, KLF2, TNFAIP3 and MYD88, common mutations in splenic marginal zone lymphoma, were rare (one KLF2 mutant in 42 samples; 2%) or absent (TNFAIP3 and MYD88) in splenic diffuse red pulp lymphoma. These findings define an original genetic profile of splenic diffuse red pulp lymphoma and suggest that the mechanisms of pathogenesis of this lymphoma are distinct from those of splenic marginal zone lymphoma and hairy-cell leukemia.

Figure 1.

Distribution of mutations in hairy-cell leukemia, splenic diffuse red pulp lymphoma and splenic marginal zone lymphoma. Each column represents a type of lymphoma and corresponding tissue (HCL: hairy cell lymphoma; SDRPL: splenic diffuse red pulp lymphoma; SMZL: splenic marginal zone lymphoma; PB: peripheral blood; SP: spleen; B+: CD19⁺ immunoselected sample). The orange colored boxes indicate the ten cases investigated by whole-exome sequencing. Six of them were also reanalyzed by targeted sequencing as internal controls, whereas the four remaining were not (NR: not resequenced). Each row (top) represents a gene linked to the corresponding RAS/MAPK, NF-κB or NOTCH signaling pathway. Three types of mutations (missense, nonsense, frameshift indel) are highlighted in different colors. Details of the mutations are available in Online Supplementary Table S3. Gene copy number (CN) was only reported as a gain (↑) or loss (↓) for relevant genes (❋), namely BCOR and TNFAIP3. The last row (bottom) represents the IGHV gene status, homology with germline sequence percentage, and cytogenetic findings (complex karyotype, trisomy 3, 12, 18 and deletion 7q) for each case when available.

Figure 2

BCOR mutations in splenic diffuse red pulp lymphoma, lymphoid and hematologic malignancies. The distribution of mutations along the BCOR sequence was drawn using cBioPortal (http://www.cbioportal.org). Mutation diagram circles are colored with respect to the corresponding mutation types. Six distinct mutations were detected in our SDRPL series (top), compared to mutations reported in lymphoid (middle) or hematologic (bottom) malignancies.^,44 BCOR: BCL-6 corepressor, non-ankyrin-repeat region (1202 - 1414); Ank_2: Ankyrin repeats (3 copies) (1467 - 1560); PUFD: BCORL-PCGF1-binding domain (1634 - 1747).

Figure 3

Copy number evaluation at the BCOR Xp11.4 locus in hairy-cell leukemia, splenic diffuse red pulp lymphoma and splenic marginal zone lymphoma. (A) Copy number (CN) variation was detected from exon sequencing of the target genes after normalization using DeCovA.^, Each row represents a lymphoma case (HCL: hairy-cell lymphoma; SDRPL: splenic diffuse red pulp lymphoma; SMZL: splenic marginal zone lymphoma). Each column represents an exon of two different genes (BCOR and KDM6A) located at locus Xp11. A progressive gradation of CN variation distinguishes loss (red) and gain (green). Right: mean copy number ratio of the BCOR locus, with some references in brackets corresponding to the next inserts (B–G). (B) Microarray-based comparative genomic hybridization confirmed a monoallelic microdeletion of approximately 669 kb (arr[GRCh37] Xp11.4(39576746_40245183)×1), encompassing the BCOR locus (SDRPL female patient VL_218), whereas the KDM6A gene was unaffected. (C–D) Copy number variation of chromosome X detected by whole-exome sequencing using a circular binary segmentation algorithm and compared to a paired reference genome (log₂ ratio). These two female patients with SDRPL acquired complete monosomy X. (E–G) Detailed karyotype results confirmed monosomy X (E), tetrasomy X (F) or trisomy X (G).