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Detection of the t(2;5)(p23;q35) and NPM-ALK fusion in non-Hodgkin's
lymphoma by two-color fluorescence in situ hybridization
P Mathew, WG Sanger, DD Weisenburger, M Valentine, V Valentine, D Pickering, C Higgins, M Hess, X Cui, DK Srivastava and SW Morris
Department of Pediatrics, Medical College of Ohio, Toledo, USA.
The non-Hodgkin's lymphoma (NHL) subset commonly referred to as large cell
lymphoma (LCL) has historically been characterized by it's marked
cytological, immunological, and clinical heterogeneity. One potential
defining feature of these lymphomas, the t(2;5)(p23;q35), occurs in 25% to
30% of anaplastic LCLs and is also found in cases with diffuse large cell
or immunoblastic morphology. We recently identified nucleophosmin (NPM) and
anaplastic lymphoma kinase (ALK) as the genes on chromosomes 5 and 2,
respectively, that are juxtaposed by this translocation. To provide a
complementary approach to the use of classical cytogenetics or polymerase
chain reaction-based methods for the detection of this abnormality, we have
developed a two-color fluorescent in situ hybridization (FISH) assay for
the t(2;5) that may be used for the analysis of both interphase nuclei and
metaphase chromosomes. Three overlapping chromosome 5 cosmid clones located
immediately centromeric to the NPM gene locus and an ALK P1 clone located
telomeric to the chromosome 2 breakpoint were labeled with digoxigenin or
biotin, respectively, and used to visualize the derivative chromosome 5
produced by the t(2;5), evident as juxtaposed or overlapping red and green
fluorescent signals. This NPM-ALK FISH assay was initially validated by
analysis of a series of cytogenetically characterized cell lines, with the
presence of the der(5) chromosome showed specifically only in those lines
known to contain the t(2;5). The assay was then applied in a blinded
fashion to a series of eight cytogenetically t(2;5)-positive clinical
specimens and seven known t(2;5)-negative cases, including three NHL and
four Hodgkin's disease biopsy samples. Whereas the t(2;5)-negative cases
were negative by FISH, all eight t(2;5)-positive cases were positive. One
additional case, initially thought to be positive for the translocation by
cytogenetics, was proven to not be a classic t(2;5) by interphase and
metaphase FISH. These data indicate that the FISH assay described is a
highly specific and rapid test that should prove to be a useful adjunct to
the currently available methods for detection of the t(2;5).
Volume 89,
Issue 5,
pp. 1678-1685,
03/01/1997
Copyright © 1997 by The American Society of Hematology
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