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KMID : 0350519930460020533
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Journal of Catholic Medical College
1993 Volume.46 No. 2 p.533 ~ p.545
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Identification of Clonality and Discrimination of B-and T-Lineage in Lymphoproliferative Diseases by Amplification of Rearranged Antigen Receptor Genes
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Abstract
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The phenomena of successive and orderly rearrangements of antigen receptor genes were recently found during the differentiation of lymphoid cells. Such gene rearrangements can be widely applied to clinics. By amplifying the junctional site of
rearrangements, minimal residual disease as well as the clonality can be detected more easily in lymphoid neoplasms.
We investigated the clonality and the pattern of gene rearrangements in lymphoproliferative diseases, and evaluate whether gene rearrangements could serve as a clonal marker in B- and T- cell neoplasms.
Each immunophenotype of non-Hodgkin´s lymphoma (NHL) and leukemia was determined by immunohistochemical staining and fluorescence-activated cell sorting scan analysis using cell-specific monoclonal antibodies. We amplified both the
complementarity-determining region 3 (CDR3) of rearranged immunoglobulin heavy chain (IgH) gene and the junctional site of rearranged T cell receptorγ(TCRγ) gene by polymerase chain reaction (PCR). Paraffin embedded tissues or cryopreserved
bone
marrow from 35 patients with lymphoproliferative diseases including immunophenotypically assigned 20 NHL (10 B-NHL, 10 T-NHL) and 6 leukemia (3 B-lineage, 3 T-lineage) and other lymphoid diseases (1 Hodgkin´s disease, 8 benign conditions) were
tested.
@ES The results were as follows :
@EN 1. PCR with primers corresponding to the consensus sequences of variable and joining segments flanking the CDR 3 of IgH resulted in a discrete band of amplified product, 100 to 120 base pairs (bp) in size, in all of B-cell (KM 3, NALM 6,
NAMALVA),
but failed in T-cell line (Molt 3). Conversely, PCR with consensus primers of TCRγ gene showed a discrete band of amplified product, 460 to 500 bp in size, in Molt 3 cell line, but no amplified material in B-cell lines.
2. Among 13 B-cell lymphoid neoplasms (10 NHL and 3 leukemia), 12 (92%) showed IgH generearrangements and 2 (15%)showed TCRγgene rearrangements.
3. Of 13 T-cell lymphoid neoplasms (10 NHL and 3 leukemia), 7 (54%) had TCRγ gene rearrangements, whereas 8 (62%) had IgH gene rearrangements. Twenty-six of 27 lymphoid neolasms demonstrated IgH or TCRγ gene rearrangements, or both.
4. Of 8 benign lymphoproliferative diseases, only one of 2 Castleman´s diseases and 2 of 5 reactive hyperplasia showed IgH and TCRγgene rearrangements, respectively.
5. As 10E3~10E4 fold dilution of the positive DNA could be amplified, the sensitivity of PCR technique was 0.1%~0.01%.
From these results, we suggest that both gene rearrangements and clonality can be detected by PCR in the great majority of patients with lymphoid neplasms. PCR amplification of CDR3 of IgH gene and TCRγ gene was a gensitive and accurate method
for
identifying the clonality and discriminating between B- and T-lineage lymphoid malignancies.
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