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L. Paschold et al.
from Moraxella species bind to the lymphoma BCR in a substantial fraction of IgD-positive NLPHL patients.9 This is striking since it suggests that an infectious trigger may drive lymphomagenesis (and potentially also lymphoma evolution) in a subset of patients with NLPHL. The Moraxella-associated cases from this study included only IgD-positive cases with BCR that share common features, such as long CDR3 sequences and specific IGH rearrange- ments.
In our study, we confirmed the unique IGHV/D/J rearrangement with a characteristically “longCDR3” con- figuration in approximately one-third of NLPHL patients, although acknowledging potential selection biases of our cohorts, such as patients with a history of relapse or transformation. The major determinant for antigen recog- nition is the CDR3 portion of the variable region of the BCR. The very unique characteristics of this region, which are shared between many NLPHL patients, togeth- er with the finding of ongoing mutational events in the majority of the studied patients, in our view clearly sug- gests that antigenic BCR interactions of the LP-cell clone must be pathophysiologically relevant, as reported for other lymphomas.42-46 Our present data do, however, clearly show that almost one-third of IgD-negative NLPHL also share the same characteristic IGHV/D/J rearrangement and might therefore have a similar patho- genesis. Future treatment strategies in early disease stages of cases with this BCR configuration might therefore con- sist in primary antimicrobial therapy to induce remission (as, for example, in Helicobacter-associated lymphoma) followed by radiation or chemotherapy only if remissions cannot be achieved. This may be especially feasible in NLPHL, since the disease has a rather indolent clinical course providing a “window of opportunity” for strate- gies to be attempted without compromising the curative potential of more aggressive treatment options.47
We noted a higher occurrence of atypical variant pat- terns in cases with relapse and transformation, which is in line with previous studies.3,10 However, given the long time until relapse/transformation in a substantial fraction of cases, we cannot exclude the possibility of future recurrences in cohort 1, despite the mean follow-up of 16 years.
From a diagnostic perspective, it was interesting for us to see that the B lineage patterns of NLPHL that can be deduced by IGH NGS can help to discriminate NLPHL cases from other related disorders that – by morphologi- cal evaluation – may occasionally pose challenges in dif- ferential diagnosis. In our analysis, we were able to dis- tinguish clonal T-cell/histiocyte-rich large B-cell lym- phoma repertoires, which resemble those of DLBCL, from the more diverse NLPHL repertoires. Furthermore, DLBCL cases that transformed from NLPHL showed more diverse B-cell repertoires than conventional DLBCL, which might argue that this entity should be kept apart from the general category of DLCBL and would be in line with the frequently observed favorable outcome of these cases after salvage therapy.14,48 As NGS becomes increas- ingly available, also for routine clinical applications, this technique could be used to support accurate discrimina- tion between lymphoma subgroups in cases that are mor- phologically challenging. This is especially relevant since these lymphomas often require different treatment modalities.
In addition, we identified a B lineage pattern that was
associated with subsequent NLPHL transformation. This pattern was essentially characterized by higher B lineage clonality and lower diversity already at the pre-transfor- mation NLPHL stage of the disease as well as absence of the paradigmatic “longCDR3” IGH rearrangement. When studying the LP-cell clone and its intraclonal variants sep- arately from the bulk B-cell repertoire (which also includes non-malignant bystander cells), we found the product of some sort of intraclonal diversification in the majority of all studied NLPHL cases both at initial diagno- sis and at relapse/transformation. However, transforming cases showed a greater magnitude of intraclonal diversifi- cation within the LP cell clone as another hallmark of this risk group. Collectively, these data indicate that higher age, IgD-negativity, high B-cell clonality with increased intraclonal diversification and the absence of a “longCDR3” LP rearrangement characterize NLPHL cases with a high risk of transformation to NHL. Clinically, this information could trigger more intense follow-up of patients at increased risk.
Beyond the diagnostic perspective, evolutionary pat- tern analysis suggested that, compared to transformed cases, relapsing cases more likely originated from a fully identical cell of origin. In transforming cases, intraclonal diversification was much more complex with the founder clones of initial diagnosis and transformation more likely being unrelated or separated by a sequence of mutational events. This, in turn, could be interpreted as higher anti- genic selection pressure (potentially via non-Moraxella antigens) acting on the LP-cell clone in the cases that ulti- mately transformed to NHL.
Of note, our strategy to derive the malignant clone from bulk sequencing of paired samples has the limitation that, unlike microdissection, it is an indirect method that is open to the possibility of misinterpretation. While we can confidently derive the malignant rearrangement from overlapping identical clones with high frequencies, some cases are less absolute. In particular, conclusions on cases NLPHL34 and NLPHL41, for which we did not identify the LP-cell clone but suggested that the transformation arose from a different cell of origin, must be treated with care.
Taken together, our data are strongly indicative of a pathophysiological role of antigens in driving lymphoma- genesis and transformation in NLPHL. They provide bio- logical insight into the cell of origin underlying relapse and transformation. Moreover, our data support the diag- nostic value of NGS in this type of lymphoma to substan- tiate the diagnosis in unclear cases and to predict the risk of transformation.
Disclosures
No conflicts of interest to disclose.
Contributions
SH and MB conceived and designed the research project; SH, MV, DE, AR, ACF, CW, WK, GO, and FF supplied critical materials; MB, EW and BT established the methods; JB, LP and DS performed experimental work; MB, LP and SH analyzed and interpreted the primary data; MB and LP drafted the man- uscript.
Acknowledgments
The authors thank Elena Hartung, Smaro Soworka and Marta Siedlecki for excellent technical assistance.
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