According to the 5th edition of the World Health Organization (WHO) Classification of Tumours of Haematopoietic and Lymphoid Tissues[2], marginal zone lymphomas (MZLs) are a heterogeneous group of low-grade B-cell lymphomas with overlapping morphology and immunophenotypes. MZLs are divided into four main entities, including extranodal marginal zone lymphoma of mucosa-associated lymphoid tissue (EMZL), nodal marginal zone lymphoma (NMZL), paediatric nodal marginal zone lymphoma (pNMZL) and primary cutaneous marginal zone lymphoma (PCMZL). EMZL is the most common subtype, accounting for nearly 70% of all MZLs. However, despite MZLs sharing some common features, there are significant differences in cytogenetic and mutational profiles among them arising in different anatomic sites[2].
The BCL-6 gene, which is located at chromosome band 3q27 encoding a zinc-finger transcription repressor, functions as a nuclear transcriptional repressor required for germinal center (GC) development and T-helper 2-mediated antigen responses. The bcl6 protein is strongly expressed in B-cells and T-cells within the GC, and its down-regulation is a crucial step for the differentiation of GC lymphocytes into memory B cells or plasma cells, or for the induction of selective apoptosis following antigen stimulation[3]. EMZL originates from post-GC marginal zone B cells and mostly lacks BCL6 rearrangement, and typically does not express bcl-6. Relapsed EMZL cases can very rarely exhibit aberrant BCL6 expression following treatment, posing a significant diagnostic challenge for pathologists. To date, only three articles have been reported in PubMed[4–6]. Of note, there is significant heterogeneity among the samples reported in these three articles. The first article included composite lymphomas (EMZL with concomitant large cell components) and large cell variants of MZL. In that study, the protein expression of bcl6 and rearrangement of the BCL6 locus were exclusively observed in the large cell morphology of EMZL, rather than in the low-grade small cell components. Furthermore, the authors suggested that bcl6 could serve as an immunohistological marker for the transformation of MZBL, and that BCL6 rearrangement might indicate an association with lymphoma transformation or progression[4]. The second article studied 392 cases of EMZL, among which seven cases (1.8%, 7/392) carried a BCL6-involved chromosome translocation as determined by FISH analysis. The authors concluded that BCL6 translocation could contribute to lymphoma development[5]. Moreover, these seven cases showed the typical histologic and immunophenotypic characteristics of MALT lymphoma, and none displayed evidence of transformation[5]. Interestingly, only two of the seven cases expressed bcl6. The third paper described six cases of initially diagnosed stage I EMZL cases with positive bcl6 staining, all of which posed challenges in differentiation from Follicular lymphoma (FL). Dual bcl6/bcl2 immunostaining was performed to confirm the concurrent coexpression of bcl6 and bcl2 proteins in individual neoplastic cells within the follicular foci, and FISH analysis was used to confirm the absence of BCL2 rearrangement. Based on the overall features, these cases were ultimately diagnosed as EMZL[6]. That study also suggested that extranodal MZL cells exhibit plasticity in bcl6 protein expression patterns, which is influenced by the GC microenvironment, while BCL6 translocation status has not been determined[6].
Herein, both cases were primarily diagnosed as typical EMZLs without bcl6 expression. However, aberrant bcl6 protein expression within neoplastic cells appeared at the time of tumor recurrence or progression after treatment. One case (case 1) developed lymph nodal involvement eight years after standardized chemotherapy and was diagnosed with EMZL with partial high-grade transformation. This could be referred to as an asynchronous composite lymphoma, characterized by the occurrence of transformed aggressive B-cell lymphoma in extra-primary sites following a confirmed diagnosis of EMZL[7]. Notably, the specimens initially exhibited numerous intranuclear Dutcher bodies. However, the recurrent specimens unexpectedly exhibited strong bcl6 positivity and concurrent loss of bcl2 expression. Monoclonal plasmacytic differentiation appears to be common, often accompanied by high-grade transformation, particularly in relapse cases, which may indicate a poor prognosis or result from chemotherapy[8]. The other case (case 2) present with bone marrow involvement (stage Ⅳ) 2 years after confirmed EMZL following biopsy. The neoplastic cells showed weak positivity for bcl6, with no evidence of high-grade transformation. It is, however, unknown whether altered expression of the bcl6 protein is an early event in high-grade transformation, a consequence of chemotherapy, or others. Studies have previously reported differential upregulation or downregulation of CD10, BCL6, and MUM1 expression in cases of transformed DLBCL. Approximately 9–16% of DLBCL transformed from FL are classified as non-GCB type, whereas about 37% of DLBCL transformed from EMZL may exhibit GCB-type characteristics (CD10+ and/or bcl6+)[7]. However, the clinical significance of this rare phenomenon warrants further substantiation in a larger case series with long-term follow-up before concluding that abnormal bcl6 expression contributes to the high-grade transformation of EMZL.
In summary, what makes our cases special is that aberrant bcl6 expression were observed in recurrent or progressing EMZL specimens at new anatomic sites, whereas the primary specimens were negative for the bcl6 marker. Additionally, FISH analysis for BCL6 gene rearrangement was performed on initial and recurrent specimens in two cases, with the results being negative (though the decalcified bone marrow specimen from case 2 yielded no analyzable results). In this study, the abnormal expression of bcl6 protein in relapsed EMZL cases has been confirmed to be independent of BCL6 gene rearrangement. This finding suggests potential involvement of other mechanisms, including gene amplification, somatic mutations in promoter regions, epigenetic regulation, as well as transcriptional, post-transcriptional, and translational modifications[3, 9, 10].
The differential diagnosis of this case was performed as follows. (1) Follicular lymphoma (FL): Some cases of FL can exhibit marginal zone differentiation with loss of GC markers expression, resembling that of MZL, particularly in extranodal sites. Conversely, certain MZLs may present with a vaguely follicular nodular distribution with extensive GC colonization, mimicking the growth pattern of FL. Although typically negative, a very small subset of EMZL can express bcl6 or other GC markers, particularly in gastric EMZL. Consequently, distinguishing between FL and MZL is highly challenging for pathologists. In the present study, both primary tumors arose from extranodal sites and exhibited typical morphological and immunohistochemical features of EMZL, making the diagnosis definitive. Meanwhile, other low-grade B-cell lymphomas, such as FL, mantle cell lymphoma, and small lymphocytic lymphoma, were ruled out. The aberrant expression of bcl6 protein within neoplastic cells was observed in recurrent or progressing specimens, which raises the diagnostic pitfall of FL. Firstly, the progression of EMZL into a high-grade FL seems less likely unless this was a second malignant neoplasm occurrence. Secondly, the negative finding for BCL2 gene rearrangement helps to exclude FL in this case. Moreover, Akiko et al. also reminded us that large transformed lymphoma cells with a bcl6 + and/or CD10 + immunophenotype should not be diagnosed as FL[7]. (2) Diffuse large B cell lymphoma (DLBCL): DLBCL represent an aggressive and heterogeneous group of mature B-cell lymphomas that may arise de novo or transform from pre-existing low-grade B-cell lymphomas, for instance, MZLs or FLs. The histological features observed, including typical monocytoid B-cells with colonizing follicles, accompanied by regional diffuse proliferation of large blast-like lymphoid cells, frequent mitotic figures, and a high Ki-67 proliferation index, support the diagnosis of EMZL, partially transformed into a large B-cell lymphoma.