Performance of MYC, BCL2, and BCL6 break-apart FISH in small biopsies with large B-cell lymphoma: a retrospective Cytopathology Hematopathology Interinstitutional Consortium study

Abstract

Introduction: Fluorescence in situ hybridization (FISH) is an essential ancillary study used to identify clinically aggressive subsets of large B-cell lymphomas that have MYC, BCL2, or BCL6 rearrangements. Small-volume biopsies such as fine needle aspiration biopsy (FNAB) and core needle biopsy (CNB) are increasingly used to diagnose lymphoma and obtain material for ancillary studies such as FISH. However, the performance of FISH in small biopsies has not been thoroughly evaluated or compared to surgical biopsies.

Methods: We describe the results of MYC, BCL2, and BCL6 FISH in a series of 222 biopsy specimens, including FNAB with cell blocks, CNBs, and surgical excisional or incisional biopsies from 208 unique patients aggregated from 6 academic medical centers. A subset of patients had FNAB followed by a surgical biopsy (either CNB or excisional biopsy) obtained from the same or contiguous anatomic site as part of the same clinical workup; FISH results were compared for these paired specimens.

Results: FISH had a low hybridization failure rate of around 1% across all specimen types. FISH identified concurrent MYC and BCL2 rearrangements in 20 of 197 (10%) specimens and concurrent MYC and BCL6 rearrangements in 3 of 182 (1.6%) specimens. The paired FNAB and surgical biopsy specimens did not show any discrepancies for MYC or BCL2 FISH; of the 17 patients with 34 paired cytology and surgical specimens, only 2 of the 49 FISH probes compared (4% of all comparisons) showed any discrepancy and both were at the BCL6 locus. One discrepancy was due to necrosis of the CNB specimen causing a false negative BCL6 FISH result when compared to the FNAB cell block that demonstrated a BCL6 rearrangement.

Discussion: FISH showed a similar hybridization failure rate in all biopsy types. Ultimately, MYC, BCL2, or BCL6 FISH showed 96% concordance when compared across paired cytology and surgical specimens, suggesting FNAB with cell block is equivalent to other biopsy alternatives for evaluation of DLBCL or HGBCL FISH testing.

Keywords: BCL2 rearrangement; FISH; MYC rearrangement; diffuse large B-cell lymphoma; double-hit lymphoma; high-grade B-cell lymphoma.

Figure 1

(A) H&E-stained section of lymph node cell block demonstrating effacement by high-grade B-cell lymphoma with MYC and BCL2 rearrangements. (B) MYC interphase FISH showing separation of orange and green signals indicating a MYC rearrangement (see arrows). (C) Pap-stained smear slide showing aggregates of large atypical lymphoid cells. (D) May-Grünwald Giemsa (MGG)-stained smear slide of cytology smear slide similarly showing large cells. In general, Pap stains and MGG stains are complementary and helpful to obtain in all lymph node FNAB: Pap-stained slides demonstrate greater nuclear detail but make cells appear smaller and have poor cytoplasmic detail while May-Grünwald Giemsa stains show greater cytoplasmic detail and larger cell size but poor nuclear detail.

Figure 2

Diagnosis of all large B-cell lymphoma specimens with MYC, BCL2, and/or BCL6 FISH performed. The diagnosis was retrieved retrospectively with FISH results already incorporated. Most cases were diffuse large B-cell lymphoma (DLBCL) (n=173). High-grade B-cell lymphoma with MYC and BCL2 rearrangements (HGBCL-MYC/BCL2) (n=19) and high-grade B-cell lymphoma, NOS (HGBCL) (n=9) were smaller subsets. Burkitt lymphoma (n=5), primary mediastinal large B-cell lymphoma (PMLBCL; n=3) and T-cell, histiocyte-rich large B-cell lymphoma (TCHRLBCL; n=2) were rare in this cohort.

Figure 3

Sankey diagram of specimen type collected and specimen type undergoing FISH testing by institution (data access group). Of all the 222 specimens that underwent FISH testing in this cohort (right column), the majority are core biopsy, followed by excisional biopsy, and cell block. The core biopsy that have FISH performed include, of course, core biopsy only specimens but also FNAB with core biopsy and all FNA with cell block and core biopsy. One FNA with cell block and core biopsy had FISH performed on the cell block due to decalcification of the core biopsy (bone specimen). Abbreviations used: FNA (fine needle aspiration), CB (cell block from FNA), core biopsy (core needle biopsy), EB (excisional biopsy), MGH (Massachusetts General Hospital), MSKCC (Memorial Sloan Kettering Cancer Center), SFVAHCS (San Francisco Veterans Administration Health Care System), UCSF (University of California San Francisco), UVA (University of Virginia).

Figure 4

MYC FISH results across specimen types. MYC FISH results from all 222 specimens are arranged per specimen type that had FISH performed. MYC was rearranged in 18% of specimens and showed 4 “other” variant MYC FISH results, which included 3 specimens with 3’ signal loss and 1 with extra 5’ signaling. The unsuccessful or hybridization failure rate was overall low at 1.8% and similar across specimen types.

Figure 5

BCL2 FISH results across specimen types. BCL2 FISH results from all 222 specimens are arranged per specimen type that had FISH performed. BCL2 was rearranged in 34% of cases by FISH and showed 1 “other” variant result with 1 extra signal BCL2 FISH result on an add(3). The unsuccessful or hybridization failure rate was low at 1.0% and similar across specimen types.