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L. Wang et al.
MYC can partner with immunoglobulin (Ig) and non-Ig genes in multiple types of B-cell lymphoma including dif- fuse large B-cell lymphoma (DLBCL), high grade B-cell lymphoma (previously known as B-cell lymphoma, unclassifiable, with features intermediate between DLBCL and BL)10-12 and rarely other small B-cell lym- phomas, such as follicular lymphoma, chronic lymphocyt- ic leukemia/small lymphocytic lymphoma and MCL. MYC alterations are often associated with an aggressive clinical course.13-19
Double-hit lymphoma (DHL) was defined broadly by Aukema et al.20 as a mature B-cell lymphoma with a chro- mosomal breakpoint affecting the MYC locus combined with additional translocations involving other genes, such as BCL2, BCL3, BCL6, or CCND1. The most common genetic combination in DHL is MYC/8q24 rearrangement and t(14;18)(q32;q21)/IGH-BCL2, which represents about 65% of cases.20-22 Significant advances in the understanding of DHL have been made in recent years, and large B-cell lymphomas with MYC and BCL2 and/or BCL6 rearrange- ments were included in the category of high-grade B-cell lymphoma with MYC and BCL2 and/or BCL6 rearrange- ments in the 2017 World Health Organization (WHO) classification, except for cases that fulfill criteria for a fol- licular lymphoma, MCL, or lymphoblastic lymphoma.23 As a result, MCL with MYC rearrangement (MYC-R), although fulfilling the earlier concept of DHL suggested by Aukema and colleagues, remains in the category of MCL.
MCL associated with MYC-R is rare and only case reports and small case series have been reported previous- ly.6,24-29 No study has explored the prognostic significance of MYC-R in MCL patients by comparing the survival of MCL patients with or without MYC-R. In addition, as we have studied cases of MCL by fluorescence in situ hybridization (FISH) to assess MYC we have come across cases of MCL with extra copies of MYC (MYC-EC), but without MYC-R. The prognostic impact of MYC-EC has not been well characterized previously.
In this study we had two aims. First, we addressed the prognostic impact of MYC-R in MCL patients, and in par- ticular, is the prognosis more akin to that of patients with DHL with MYC and BCL2 rearrangements (MYC/BCL2 DHL). Secondly, we addressed the question of the poten- tial prognostic impact of MYC-EC in the absence of MYC rearrangement in MCL patients.
Methods
Case selection
We searched the cytogenetic/FISH testing database of the Department of Hematopathology at The MD Anderson Cancer Center from January 1, 2004 to December 31, 2018 and identified 88 cases of MCL with 11q13/CCND1 and 8q24/MYC tested by FISH and/or conventional karyotyping. Only three cases were before 2010 and most cases were diagnosed in recent years. MYC-R in MCL is rare and there are no standard rules or criteria for which MCL should be tested for MYC FISH, so the choice of MYC FISH testing for MCL was solely at the discretion of the treating oncologist and diagnosing hematopathologist. However, in general it was performed on a small subset of blastoid/pleomor- phic MCL and occasional classic MCL cases. Ninety-five cases of high-grade B-cell lymphoma with concurrent MYC and BCL2 rearrangements confirmed by FISH (MYC/BCL2 DHL) from the same time period were used as a comparison group. Clinical infor-
mation was obtained by review of corresponding medical records, including lymphoma history, sites of involvement, stage, treat- ment and overall survival (OS). Morphologic, immunophenotypic and cytogenetic data were also reviewed. The diagnosis of all cases was made according to the criteria of the current WHO clas- sification.10,11 The study was approved by the Institutional Review Board.
Immunophenotypic methods
Immunohistochemical stains were performed using formalin- fixed, paraffin-embedded (FFPE) tissue sections, either at the time of diagnosis or retrospectively for the purpose of this study. The monoclonal antibodies used were specific for: CD3, CD5, CD10, CD20, BCL-2, BCL-6, IRF4/MUM-1, MYC, P53, Ki67, cyclin D1, and SOX-11. The positive cutoff was ≥30% for CD10, MUM-1, and BCL630; ≥20% for P5331; ≥40% for MYC16; ≥50% for BCL232 and >10% for SOX1133.
Flow cytometry immunophenotypic analysis was performed using either a FACScanto II or FACSCalibur cytometer (Becton- Dickinson Biosciences, San Jose, CA, USA) as described previous- ly.34,35 Lymphocytes were gated for analysis using side scatter versus forward scatter, and CD45 versus side scatter. The panel of antibodies employed included CD3, CD5, CD10, CD11c, CD19, CD20, CD22, CD23, CD30, CD38, CD43, CD45, CD79b, CD200, FMC-7, and surface Ig κ and l light chains. All antibodies were obtained from Becton-Dickinson Biosciences.
Conventional cytogenetics and fluorescence in situ hybridization
Conventional chromosomal analysis was performed on G- banded metaphase cells prepared from cell suspensions from tis- sue biopsy specimens or bone marrow aspirates using standard techniques. The karyotype was reported according to the International System for Human Cytogenetic Nomenclature (2016).36 FISH was performed on bone marrow smears or 4-μm- thick FFPE tissue sections according to the manufacturer’s instruc- tions. A total of 200 interphase nuclei for each probe were ana- lyzed. FISH probes used in this study included the following: locus specific identifier (LSI) IGH/CCND1 dual-color, dual fusion translocation probe; LSI MYC as well as BCL6 dual-color, break- apart probe; LSI IGH/BCL2 dual-color, dual-fusion translocation probe (Vysis/Abbott Laboratories, Des Plaines, IL, USA).
Statistical analysis
Overall survival (OS) was calculated from the date of initial diagnosis (for de novo cases) or the date that a MYC aberration was detected (for patients with MYC aberration detected at disease transformation or progression) to the date of death or last follow- up. Survival was analyzed using the Kaplan-Meier method and was compared by log-rank test (GraphPad Prism version 7 soft- ware). Fisher’s exact test was utilized to compare the difference between groups. Multivariate Cox proportional hazard analysis was performed using SPSS 24.0 software. Differences between groups were considered statistically significant when the P-value is less than 0.05.
Results
Mantle cell lymphoma patients with MYC rearrangement
Clinical characteristics
Twenty-seven MCL patients had MYC-R, including 20 men and 7 women, with a median age of 63 years (range, 47 to 85). Fourteen (52%) patients with MYC-R presented
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haematologica | 2021; 106(5)