Patients with double/triple copy number gains on C-MYC, BCL2, and/or BCL6 treated with standard chemotherapy have a similarly poor prognosis than those with high-grade B cell lymphoma with C-MYC and BCL2 and/or BCL6 rearrangements: a single-center experience on a consecutive cohort of large B cell lymphomas

Large B cell lymphomas (LBCLs) comprise a heterogeneous group of lymphomas. While some of them, such as Burkitt lymphoma, have a well-defined morphology and molecular pathology, others are less well defined and disease outcome is highly variable. Especially, the group of diffuse large B cell lymphomas, not otherwise specified (DLBCL, NOS) consists of different morphological, immunohistochemical, and molecular subgroups and various subtypes significantly differing in clinical behavior and treatment outcome [1]. The WHO classification of hematopoietic tumors from 2008 introduced a provisional entity called large cell B cell lymphoma with features between a DLBCL, NOS and Burkitt lymphoma (BCLU) including large B cell lymphomas with Burkitt-like morphology, but distinct immunohistological features and the frequent presence of C-MYC rearrangements often together with a BCL2 and/or BCL6 rearrangement, so-called double/triple hit (DH) [2, 3]. These cases have been widely discussed in the literature and have been associated with a poor prognosis [4‐7]. However, rearrangements of C-MYC together with BCL2 and/or BCL6 have also been reported in otherwise classical DLBCL, NOS [8, 9]. In addition, ill-defined criteria for diagnosing BCLU have prevented a more consistent and comprehensible usage of the diagnosis [10, 11]. Therefore, the current WHO classification includes all B cell lymphomas with rearrangements of C-MYC and BCL2 and/or BLC6 into the category of high-grade B cell lymphomas with rearrangements of C-MYC and BCL2 and/or BLC6 (HGBL with MYC and BCL2 and/or BCL6 rearrangement) irrespective of morphology, while lymphomas without rearrangement, which would otherwise fit into the former category of BCLU, are now summarized as HGBL, NOS [12].

Beside C-MYC rearrangements, expression of C-MYC protein > 40% together with BCL2 expression in > 50% of lymphoma cells has been reported to be associated with a poor outcome which is however better than in HGBL, C-MYC. Therefore, overexpression is only seen as a prognostic factor [9, 12]. In addition, several studies point towards a role of copy number gains (CNG) on C-MYC and/or BCL2 and/or BCL6 [13‐15]. In fact, patients with CNG are reported to have a similarly poor prognosis than those with HGBL with MYC and BCL2 and/or BCL6. [13‐15].

There is currently no agreement if an analysis for a C-MYC, BLC2, and BCL6 rearrangement should be done in every DLBCL, NOS upon diagnosis or should be restricted to cases with GCB phenotype, a high-grade morphology, or C-MYC positivity [12].

From 2012 to 2016, we analyzed a consecutive cohort of DLBCL, NOS for the presence of C-MYC, BCL2, and BCL6 rearrangements and correlated our results with clinical-pathological parameters.

In our consecutive cohort of DLBCL, NOS, we identified 10 (12%) HGL-DH and three (4%) DLBCL, NOS with C-MYC rearrangement alone. Our data are in line with previous reports on the frequency of this type of lymphomas in a consecutive cohort, which range from 3 to 21% depending upon the composition of patient population [5]. Double/triple CNGs were found in 10% of patients, where recent reports suggest a frequency of 3.8 to 6% of DLBCL, NOS [13, 15]. The slightly higher number in our group may be due to the analysis of C-MYC, BCL2, as well as BCL6 in virtually all cases, which is seldom done in routine diagnostics.

Our data on HGL-DH once more confirm the need for a different therapeutic approach in patients with LBCL harboring a C-MYC and BCL2 and/or BCL6 rearrangement, though none of the suggested escalated regimens, such as DA-EPOCH-R (dose-adjusted etoposide, prednisolone, vincristine, cyclophosphamide, doxorubicin, and rituximab) or R-CHOP-ibrutinib, proved to be superior in a randomized phase III setting so far [6, 18‐23]. As previously reported by other authors, in our study, cases with double/triple CNG had a similarly poor prognosis than HGL-DH [13‐15]. Therefore, although the group of HGL-DH includes lymphomas with a defined aberration, this group does not suit all those LBCL with a poor prognosis. Thus, the LBCLs with MYC alterations are not a homogeneous group. In fact, while HGL-DH cases usually show overexpression of C-MYC and GCB subtypes, cases with double/triple CNG lack C-MYC expression > 40% and more often belong to the ABC-group, implicating that the mechanisms by which C-MYC and BCL2 and/or BCL6 alterations finally influence prognosis may be different ones [13, 14]. This is also supported by recent genetic data on DLBCL, NOS which show, depending on the method of analysis, up to five genetic subtypes of DLBCL, NOS, of which C-MYC rearranged or double/triple hit cases were usually part of but not exclusively formed the group of molecular high-grade lymphomas [24‐27]. Therefore, further investigations are needed to fully decrypt the molecular basis for a more aggressive behavior in order to delimit this group of patients from all others.

Our study has some limitations. First of all, it is based upon a rather small and heterogeneous group of patients including also cases of known secondary DLBCL, NOS, though the majority of these clinically usual more aggressive lymphomas belonged to the group of DLBCL without CNG. Second, it represents the experience of only a single center; thus, to further support our results, a larger prospective and multicenter trial would be necessary. Third, due to the small numbers of DH-CNG cases, a further subdivision (i.e., low versus high amplification; amplification with/or without additional rearrangement on either C-MYC or BLC2 or BCL6) could not be performed. Also, since the data were collected retrospectively, information upon polyploidy was not available for routine FISH and does not include centromere probes, but break apart probes for each specific locus.

According to the actual WHO, reporting double/triple CNG is not a mandatory component for the diagnosis of a DLBCL, NOS, but given the recent reports on clinical outcome, we suggest that double/triple CNG should be reported similar to overexpression of C-MYC and BCL2 [13‐15]. This would require FISH at least for C-MYC rearrangement in any newly diagnosed DLBCL, NOS.

To summarize, though one cannot draw any definite conclusions from our data by now, we believe our results warrant further investigations in larger sample sets and preferably a prospective fashion to fully elucidate the role of CNGs as well as the underlying molecular mechanisms resulting in aggressive behavior in LBCL.