Detection of translocations affecting the BCL6 locus in B cell non-Hodgkin's lymphoma by interphase fluorescence in situ hybridization

Structural alterations in 3q27 affecting the BCL6 locus are among the most frequent changes in B-NHL. The aim of the present study was to establish an interphase-FISH assay for the detection of all diverse BCL6 translocations in B-NHL. Two different approaches were tested, one using a PAC-clone span ning the major breakpoint region (MBR) of BCL6 (span-assay), and another us ing two BAC clones flanking the MBR (flank-assay). Interphase FISH with the span-assay detected the various BCL6 translocations in seven B-NHL cell li nes. The dual-color flank-assay was evaluated in two laboratories independe ntly: in normal controls, the cutoff level for false-positive signals was 2 .6%, whereas the cutoff level for false-negatives in the seven cell lines w as 7.5%. To test the feasibility of the FISH strategies, 30 samples from pa tients with B-NHL with cytogenetic abnormalities of 3q27 were evaluated wit h both assays. In 21 cases, the span-assay indicated a BCL6 rearrangement. In 18 of the 21 cases, the dual-color flank-assay confirmed the translocati on including 12 different partner chromosomal loci. The three false-positiv e cases detected with the span-assay showed trisomy of chromosome 3 by cyto genetic analyses, and they were correctly classified as non-rearranged with the flank-assay. In summary, our FISH strategy using two differently label ed flanking BCL6 BAC probes provides a robust, sensitive, and reproducible method for the detection of common and uncommon abnormalities of BCL6 gene in interphase nuclei. The routine application of this assay to patients wit h B-NHL will allow the assessment of the diagnostic and prognostic signific ance of BCL6 rearrangements.