Frequency and structure of t(14;18) major breakpoint regions in non-Hodgkin's lymphomas typed according to the Kiel classification: analysis by direct DNA sequencing.
We have examined 165 unselected cases of non-Hodgkin's lymphomas for rearrangements involving the t(14;18) major breakpoint region using a polymerase chain reaction (PCR) and direct sequencing of amplified major breakpoint region bcl-2/JH junctional regions. The lymphomas, diagnosed according to the updated Kiel classification, consisted of 33 centroblastic-centrocytic, 37 centroblastic, 27 immunocytic, 10 immunoblastic, 10 centrocytic, 2 lymphoblastic, 2 Ki-1-positive anaplastic large cell, 14 peripheral T-cell, and 4 unclassified lymphomas. In addition 18 chronic lymphocytic leukemias, 2 hairy cell leukemias, and 6 plasmacytomas were studied. In 17 cases a bcl-2/JH gene fusion sequence was amplified by PCR. A bcl-2/JH gene fusion was detected only in three lymphoma subgroups: 13 of 33 centroblastic-centrocytic (39%), 2 of 37 centroblastic (6%), and 2 of 27 immunocytic (8%) were positive. In two cases, major breakpoint region bcl-2 rearrangements verified by genomic Southern analysis were not detected by PCR. Direct sequencing of all 17 PCR-amplified, previously uncharacterized t(14;18) junctional regions provided corroborating evidence for the specificity of the assay. The procedure gave sequencing results even from limited amounts of lymphoma cells as obtained by fine needle aspiration of lymph nodes or from clinically uninvolved sites. Clone-specific sequences were identified due to the involvement of different JH segments, the variations among the exact JH and bcl-2 breakpoint positions, and the extensive incorporation of junctional region (D-) N-nucleotides. These clone-specific sequences allow accurate identification of clinically occult lymphoma cells and reduce the threat of false positive results. The finding of exceptionally long intervening sequences in some of the junctions and the partial homology with published DH segments in three cases support the view that some of the putative N-regions harbor DH regions.
Kneba
M
M
Department of Hematology, University Clinics, Göttingen, Germany.
Eick
S
S
Herbst
H
H
Willigeroth
S
S
Pott
C
C
Bolz
I
I
Bergholz
M
M
Neumann
C
C
Stein
H
H
Krieger
G
G
D015703:Antigens, CD / Q000032:analysis; D001483:Base Sequence; D001853:Bone Marrow / Q000473:pathology; D002883:Chromosomes, Human, Pair 14; D002887:Chromosomes, Human, Pair 18; D003001:Cloning, Molecular; D004273:DNA, Neoplasm / Q000235:genetics*; D005787:Gene Frequency; D006801:Humans; D007438:Introns; D008198:Lymph Nodes / Q000473:pathology; D008228:Lymphoma, Non-Hodgkin / Q000145:classification / Q000235:genetics* / Q000276:immunology / Q000473:pathology; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes; D016133:Polymerase Chain Reaction; D014178:Translocation, Genetic
D015703:Antigens, CD; D004273:DNA, Neoplasm; D015345:Oligonucleotide Probes