The double-hit signature identifies double-hit diffuse large B-cell lymphoma with genetic events cryptic to FISH

Abstract

High-grade B-cell lymphomas with MYC and BCL2 and/or BCL6 rearrangements (HGBL-DH/THs) include a group of diffuse large B-cell lymphomas (DLBCLs) with inferior outcomes after standard chemoimmunotherapy. We recently described a gene expression signature that identifies 27% of germinal center B-cell DLBCLs (GCB-DLBCLs) as having a double-hit-like expression pattern (DHITsig) and inferior outcomes; however, only half of these cases have both MYC and BCL2 translocations identifiable using standard breakapart fluorescence in situ hybridization (FISH). Here, 20 DHITsig+ GCB-DLBCLs apparently lacking MYC and/or BCL2 rearrangements underwent whole-genome sequencing. This revealed 6 tumors with MYC or BCL2 rearrangements that were cryptic to breakapart FISH. Copy-number analysis identified 3 tumors with MYC and 6 tumors with MIR17HG gains or amplifications, both of which may contribute to dysregulation of MYC and its downstream pathways. Focal deletions of the PVT1 promoter were observed exclusively among DHITsig+ tumors lacking MYC translocations; this may also contribute to MYC overexpression. These results highlight that FISH fails to identify all HGBL-DH/THs, while revealing a range of other genetic mechanisms potentially underlying MYC dysregulation in DHITsig+ DLBCL, suggesting that gene expression profiling is more sensitive for identifying the biology underlying poor outcomes in GCB-DLBCL.

Graphical abstract

Figure 1.

FISH-cryptic MYC and BCL2 rearrangements identified by WGS in DHITsig⁺ DLBCL tumors. (A) Diagram summarizing the occurrence of each genetic event identified in the 20 non–HGBL-DH/TH-BCL2 DHITsig⁺ genomes. The thresholds for MYC and BCL2 immunohistochemistry (IHC) positivity were 40% and 50%, respectively. (B-C) Cryptic BCL2 translocations identified in tumors that were negative for BCL2 rearrangement by breakapart FISH. (D) A cryptic MYC rearrangement identified in a tumor that was negative for MYC rearrangement by breakapart FISH.

Figure 2.

Somatic CNVs affecting MYC, MIR17HG, and PVT1 identified by WGS among DHITsig⁺ DLBCL tumors. Focal copy-number gains of MYC (A) or MIR17HG (B) identified among DHITsig⁺ DLBCL tumors. Red bars indicate regions where Control-FREEC identified a significant increase in copy number (P < .05). Compared with 162 GCB tumors without a MIR17HG amplification as determined by single-nucleotide polymorphism (SNP) arrays, expression of MIR17HG was significantly elevated among these 6 cases with MIR17HG copy gains (log2 fold change, 0.85; Wilcoxon P = .032). (C) The boundaries of focal PVT1 transcription start site (TSS) deletions identified in DHITsig⁺ tumors. *Indicates the PVT1 deletion identified in the double-minute chromosome.