To the editor:
Follicular lymphoma in situ (FLIS) is characterized by the presence of a B-cell population with immunophenotypic and genotypic features of follicular lymphoma (FL) but exclusively localized to germinal centers (GCs) in morphologically reactive lymph nodes.1 FLIS lesions are monoclonal and carry the t(14;18)(q32;q21) translocation, which juxtaposes the BCL2 gene to the immunoglobulin heavy chain (IGH) locus, causing constitutive expression of the antiapoptotic protein BCL2. Most of the cases described showed no evidence of manifest FL (mFL) at diagnosis, however, some individuals had synchronous evidence of mFL at another site or developed mFL during follow-up.2,3 This latter finding suggests that FLIS might represent a very early stage in the development of mFL. However, the genetic events responsible for progression of FLIS to mFL are largely unknown. Here we report a case of FLIS with BCL2 expression synchronously presenting with a BCL2 negative mFL, both lesions clonally related and carrying the t(14;18)(q32;q21) translocation. The case occurred in a 78-year-old man with stage III disease and FLIPI intermediate risk. The diagnostic biopsy was composed of 2 lymph nodes. The larger one showed the typical morphology of FL grade 1/II with the characteristic phenotype (CD20+, CD10+, BCL6+), except for lack of BCL2 expression. The small lymph node showed normal architecture, but with strong BCL2 expression in some morphologically reactive GCs diagnostic of FLIS (Figure 1A-B). FISH analysis using a BCL2 break-apart probe (LSI BCL2 BAP, Vysis) confirmed a BCL2 breakpoint in both areas suggestive of the t(14;18)(q32;21) translocation (Figure 1B; insert). IGH PCR clonality analysis performed in microdissected tissue showed identical monoclonal peaks in both lesions (Figure 1C-D). Sequence analysis of the BCL2 breakpoint region confirmed the clonal identity of both lesions (Figure 1E). To explain the discordant BCL2 reactivity, exon 1 of the BCL2 gene, where the epitope of the BCL2 antibody resides, was amplified and sequenced. A point mutation resulting in amino acid substitution (c.144 C > G; p.I48M ATC–ATG) was found in the mFL, whereas the FLIS showed a wild type sequence (Figure 1F-G). Array CGH (244K platform, Agilent Technologies) analysis was performed on DNA from microdissected tissue of both FLIS and mFL.4 mFL showed gains in 6p22.2-p12.3 and losses in 6q14.1-qter and 8pter-p23.1. Subsequent FISH using probes for 6q21 and 6q27 confirmed 6q deletions in 80% of the mFL cells (Figure 1H-J). In contrast, the FLIS counterpart showed no pathogenetic alterations by array CGH. Nevertheless, 32 benign polymorphisms (CNPs) were observed, of which 16 (50%) were also detected in the mFL confirming the same patient origin.
This is the first description of a FLIS clonally related to the mFL where secondary genetic alterations are demonstrated, probably representing clonal evolution as sign of disease progression. Our findings provide evidence that FLIS is in fact a very early lesion carrying the t(14;18)(q32;q21) translocation without additional secondary genetic alterations.5,6 In this case, acquisition of numeric chromosomal aberrations and BCL2 mutation indicates that the synchronous FLIS represents cells of the founder clone rather than early colonization of reactive GCs by mFL.
Acknowledgments: The authors thank the technical staff of the laboratories in Tübingen and in Kiel for expert technical assistance. The work of R.S. on clonal evolution of t(14;18)-positive lymphomas is supported from Bundesministerium für Bildung und Forschung through the network “HämatoSys” The work of L.Q.-M., F.F. and I.B. is supported by the SFB 685.
Contribution: I.B., I.S. and A.H. performed research and analyzed data; G.G. contributed with vital patient information; P.A. analyzed data; and R.S., F.F. and L.Q.-M. designed the research, analyzed data and wrote the paper.
Conflict-of-interest disclosure: The authors declare no competing financial interests.
Correspondence: Leticia Quintanilla-Martinez, Institute of Pathology, University Hospital Tuebingen, Liebermeisterstrasse 8, 72076 Tuebingen, Germany; e-mail: email@example.com.
I.B. and I.S. contributed equally to this article.
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