MYC Gene FISH Testing in Aggressive B-Cell Lymphomas: Atypical Rearrangements May Result in Underreporting of Positive Cases

Multifunctional MYC oncogene overexpression resulting from genomic rearrangement plays a critical role in lymphomagenesis and lymphoma manifestation, particularly in aggressive B-cell lymphomas such as Burkitt lymphoma (BL), diffuse large B-cell lymphoma (DLBCL) and B-cell lymphoma unclassifiable with features intermediate between DLBCL and BL including double hit or triple hit lymphomas (DHL/THL). Accurate detection of MYC gene rearrangement, its presence or absence, has become increasingly important with its diagnostic and consequent therapeutic implications. In practice, FISH is the best test for MYC alterations. However, this is technically challenging, requiring knowledgeable and skillful cytogeneticists to design probe panels, correctly analyze and interpret atypical as well as typical signal patterns. For instance, up to 10% of patients with otherwise typical BL can be MYC rearrangement negative by FISH and, gene amplification and miRNA deregulation have been reported as possible reasons in occasional cases, although some of the FISH negative patients could be detected by better application of probes and interpretation of FISH findings. Here we retrospectively analyze FISH results from 879 consecutive MYC positive cases and share our experience with other FISH labs and physicians who are involved in diagnosis, differential diagnosis and treatment of aggressive lymphomas.

A MYC/IGH dual fusion translocation probe and a MYC break apart (ba) probe (Vysis) were applied to paraffin embedded tissue, lymph node biopsy, bone marrow and peripheral blood specimens. Of the 879 cases, MYC ba was positive in 258, MYC ba and MYC-IGH both positive in 331, MYC ba positive/MYC-IGH negative in 276, and MYC-IGH positive/MYC ba negative in 14 cases. Two subsets of cases with significant atypical signal patterns were observed. Firstly, in 14 cases with both probes tested, MYC ba was negative but MYC/IGH positive, including 6 BLs, 3 DLBCLs, 1 MCL, 1 Burkitt leukemia transformed from BL, 1 subtype unknown lymphoma and 2 DHLs with history of Hodgkin lymphoma and Burkitt-like lymphoma respectively and with concomitant BCL6 gene rearrangement in both. Eleven of these cases showed a single fusion signal pattern suggesting an insertion of IGH sequences into MYC or vise versa. This single insertion fusion pattern can also be resulted from complex chromosome changes as seen cytogenetically in 2 of the 11 cases. Secondly, 26 cases with various diagnoses including 4 DHLs and 2 THLs showed MYC rearrangement with a concomitant partial deletion of the MYC ba probe with 17 cases for 3' and 9 for 5' deletion. Three of these cases were also positive for concomitant probe deletion and amplification, two with 3' deletion and 5' amplification and one with 5' deletion and 3' amplification.

MYC gene rearrangement detection by FISH in aggressive B-cell lymphomas is widely used. However, FISH labs should realize the technical limitations of each probe/panel based on underlying mechanisms involving MYC rearrangement and minimize false negative results. From our experience and previous studies by other groups, it is recommended that MYC ba probe should not be tested alone but together with a MYC/IGH t(8;14) translocation probe to detect cryptic insertions and variant translocations which often present as MYC ba negative yet MYC/IGH positive with a single fusion signal pattern inserting IGH promoter/enhancer elements into MYC or part or all of the MYC probe into IGH locus. Another observation that can also be misinterpreted is that the MYC ba probe often shows a deletion of either 5' or 3' flanking sequence. This is not because of tissue sectioning but indicative of MYC rearrangement with a concomitant loss of the DNA sequences adjacent to the breakpoints. A flanking sequence deletion revealed at translocation breakpoints using a FISH ba probe is a common finding. As well documented, BCR/ABL translocation results in an ASS deletion upstream of ABL in ∼15% of CMLs, and a 5' probe deletion in ∼30% ALK rearrangement positive NSCLC cases. Three of 26 such cases in our study also showed simultaneous retained MYC probe amplification making FISH interpretation even more difficult. As a diagnostic, prognostic and predictive biomarker, MYC gene plays a pivotal role in aggressive B-cell lymphomas, and its accurate detection will help improve disease risk stratification and therapy selection.

No relevant conflicts of interest to declare.