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The use of fluorescent in situ hybridization for detection of the t(2;5)(p23;q35) translocation in anaplastic large-cell lymphoma.

BACKGROUND
Anaplastic large-cell lymphoma (ALCL) is a recently recognized, distinctive type of non-Hodgkin's lymphoma characterized by anaplastic large-cell cytology and expression of a member of the TNF-receptor family CD30. A characteristic chromosomal translocation has been identified in ALCL of T-or null-cell lineage which juxtaposes a novel tyrosine kinase (anaplastic lymphoma kinase, ALK) located at 2p23 with the nucleophosmin gene (NPM) at 5q35. A chimeric mRNA transcript is produced, and the translocation results in constitutive expression of a truncated form of the ALK protein, p80. There is controversy concerning whether or not the translocation occurs in Hodgkin's disease. The aim of this study was to develop a methodology for fluorescent in situ hybridization (FISH) to detect the t(2;5)(p23;q35), and to compare the results with conventional cytogenetics, reverse-transcriptase PCR and immunostaining for the p80 protein.


PATIENTS AND METHODS
Twenty-five cases of malignant lymphoma (11 ALCL and 14 HD) were studied. Immunohistochemistry was performed to confirm the diagnosis and for analysis of p80 expression. Conventional cytogenetics were analyzed on G-banded metaphase spreads. FISH was performed using whole chromosome paints for chromosomes 2 and 5 on metaphase spreads and YAC probes for interphase nuclei. Reverse-transcriptase PCR using primers for ALK and NPM was used to amplify the translocation breakpoint in extracted mRNA.


RESULTS
Among 11 cases of ALCL examined by FISH, the translocation was detected in 4. Two of these cases also had RT-PCR and p80 staining performed, with positive results. Among 7 cases where the t(2;5) was not detected by FISH, 3 cases were examined by RT-PCR with negative results and 4 cases by p80 staining, also negative. The RT-PCR was negative in all 14 cases of Hodgkin's disease, 4 of which were also examined by FISH and found to be negative.


CONCLUSION
Fluorescent in situ hybridization is a useful method for detection of the t(2;5)(p23;q35) in anaplastic large-cell lymphoma. The results concur with those of RT-PCR for the chimeric transcript and immunostaining for the p80 protein. The frequency with which the translocation was found was 36% in this small series, and no evidence of the translocation was found in cases of Hodgkin's disease.