In situ analysis of the variable heavy chain gene of an IgM/IgG-expressing follicular lymphoma: evidence for interfollicular trafficking of tumor cells.

It is generally assumed that follicular lymphomas (FL) not only morphologically resemble normal germinal centers but have retained some functional characteristics of their non-neoplastic counterparts as well. Recent IgV gene analyses on a panel of FLs however, strongly suggested that FLs do not retain the capacity of somatic hypermutation and are not being selected on basis of the quality of their mIgV regions. To extend these findings, we investigated the follicular organization and class switching in a FL that consisted of both IgM- and IgG-expressing tumor cells with a high somatic mutation load and significant intraclonal V(H) gene diversity. V(H)-C(mu) and V(H)-Cgamma gene transcripts were amplified and sequenced from samples of approximately 50 tumor cells, isolated from frozen tissue sections by laser microdissection. We identified many different subclones and obtained limited evidence of subclone dominance in individual follicles. Remarkably, several subclones were found scattered over different follicles. All samples contained IgM- and IgG-expressing tumor cells with, in general, non-identical mutation patterns, which is not in support of ongoing class switching. Accordingly, no switch circle recombination products were found. The findings indicate that the neoplastic follicles lack the organization and functions typical of reactive germinal centers.

Follicular lymphoma with a novel t(14;18) breakpoint involving the immunoglobulin heavy chain switch mu region indicates an origin from germinal center B cells.

With the use of DNA-fiber fluorescent in situ hybridization, a BCL2 protein positive follicular lymphoma with a novel BCL2 breakpoint involving the immunoglobulin heavy chain (IGH) switch mu (S(mu)) region instead of the J(H) or D(H) gene segments was identified. Sequence analysis showed that the genomic breakpoint is localized between the S(mu) region of the IGH complex and the first intron of BCL2. Reverse-transcriptase polymerase chain reaction showed expression of a unique hybrid IGH-BCL2 transcript involving the transcription initiation site I(mu). Sequence analysis of the V(H) region of the functional nontranslocated IGH allele showed multiple shared somatic mutations but also a high intraclonal variation (53 differences in 15 clones), compatible with the lymphoma cells staying in or re-entering the germinal center. This is the first example of a t(14;18) translocation that results from an illegitimate IGH class-switch recombination during the germinal center B-cell stage.