Detection of Chromosome 11 q 13 Breakpoints by Interphase Fluorescence In Situ Hybridization

We assessed cytologic specimens from 11 mantle cell lymphomas (MCLs) and 32 other B-cell nonHodgkin lymphomas (NHLs) for 11q13 breakpoints using a 2-color fluorescence in situ hybridization (FISH) assay that uses an 11q13 probe centered on the CCND1 gene and a centromeric chromosome 11 probe (CEP11). The number of nuclei in 200 cells were counted, and results were expressed as an 11q13/ CEP11 ratio. All MCLs showed a high percentage of interphase nuclei with 3 or more 11q13 signals (mean, 74.8%; range 57%-90%). In contrast, in other B-cell NHLs the mean percentage of cells with 3 or more 11q13 signals was 9.2%. All MCLs had an elevated 11q13/CEP11 ratio (mean, 1.38). The mean ratio for other B-cell NHLs was 0.99. Two non-MCL cases, 1 large B-cell and 1 B-cell unclassified NHL, had high 11q13/CEP11 ratios of 1.15 and 1.30, respectively. Conventional cytogenetic analysis performed on the former case revealed a t(5;11)(q31;q13). Interphase FISH analysis using 11q13 and CEP11 probes is a convenient ancillary method for assisting in the diagnosis of MCL. This commercially available assay is simple to use on cytology or imprint specimens, and results can be obtained within 24 hours. Mantle cell lymphoma (MCL) is a clinically aggressive, generally incurable type of non-Hodgkin lymphoma (NHL) representing up to 6% of all NHLs in the United States.1,2 MCL can be clinically and histologically difficult to distinguish from other types of small B-cell NHL and leukemia, such as follicular small cleaved cell or small lymphocytic lymphoma/chronic lymphocytic leukemia (SLL/CLL), which have very different prognoses and treatment.2 One way to support the diagnosis of MCL is to identify the t(11;14)(q13;q32), now widely accepted as the hallmark of MCL, in which the CCND1 (PRAD-1, bcl-1) gene at 11q13 is juxtaposed with the joining region of the immunoglobulin (Ig) heavy chain gene at 14q32.2,3 The CCND1 gene is up-regulated, resulting in overexpression of cyclin D1, which acts at the G1 to S phase checkpoint of the cell cycle to drive cell proliferation.4 Unfortunately, currently available methods to demonstrate the t(11;14) have substantial limitations. Conventional cytogenetics can detect the t(11;14) in up to 75% of MCLs, but karyotyping is a cumbersome procedure that requires fresh tissue, meticulous sample preparation, and adequate metaphases.5 Southern blot analysis can detect bcl-1 locus rearrangements in up to 70% of cases, but this approach requires multiple probes and is laborious and time consuming.6,7 Polymerase chain reaction (PCR) methods are available to detect the t(11;14) involving the major translocation cluster (MTC) region of bcl-1, but only 30% to 40% of MCLs have chromosome 11 breakpoints that occur in the MTC region.8 Other chromosome 11 breakpoints cannot be assessed routinely by PCR methods at this time. D ow naded rom http/academ ic.p.com /ajcp/article-t/114/2/248/1757910 by gest on 16 Feruary 2019 Hematopathology / ORIGINAL ARTICLE Am J Clin Pathol 2000;114:248-257 249 © American Society of Clinical Pathologists Alternatively, others have assessed cyclin D1 expression in MCL, because the t(11;14) results in cyclin D1 overexpression regardless of breakpoint location.2 However, methods to assess cyclin D1 expression also have limitations. Northern blot methods are time consuming and laborious and require relatively large samples of fresh or frozen tissue from which high quality RNA can be extracted. Reverse transcriptase–PCR methods require much less RNA but are not well suited for quantitative methods.9 Immunohistochemical assays for cyclin D1 overexpression are valuable in the assessment of tissue sections.10 However, these methods are not optimized for assessing cytologic smears or cytocentrifuged slides, and, in our experience, the results using cytologic preparations are less reliable than those obtained using tissue sections. For these reasons, we became interested in fluorescence in situ hybridization (FISH) methods and, in particular, a recently developed, commercially available FISH assay that simultaneously assesses 11q13 and centromeric chromosome 11 loci in interphase nuclei. Although this assay assesses only chromosome 11, the ease and convenience of this approach is attractive. In a cohort of 11 MCL cases and 32 non-MCL B-cell NHLs, all 11 MCLs had 3 or more 11q13 signals consistent with 11q13 breakpoints. In contrast, other types of B-cell NHL rarely had a substantial number of cells with more than 2 11q13 signals. Materials and Methods