DETECTION OF B-CELL LYMPHOMAS BY MOLECULAR ANALYSIS OF IMMUNOGLOBULIN HEAVY CHAIN VARIABLE REGION GENES
LIU SM1, TSAI KJ2, CHEN TC3, KUO TT3 AND CHIU DTY1.
1School of Medical Techn., Chang Gung Univ., Kwei-san, Tao-yuan, 2Dept. of Medical Techn., Chung-San Medical College, Taichung, 3Dept. of Pathology, School of Medicine, Chang Gung Univ., Kwei-san, Tao-yuan, Taiwan
The detection of rearranged VDJ gene coded for immunoglobulin heavy chain by polymerase chain reaction (PCR) has recently been proposed as a rapid approach to assess B cell clonality in lymphoproliferative disorders. However, the 15-35% incidence of false negative results with this approach has been a constant and unresolved problem. The lowest detection rates were observed in follicular lymphoma and MALT-lymphoma, implicating a possible correlation between somatic mutation and PCR false-negativity. In the first part of this study, we evaluated the accuracy of PCR methods in detecting monoclonality in a well characterized panel of frozen and paraffin-embedded B-cell lymphoid proliferative tissues. Using the FR3-FR4 protocol, we evaluated 38 cases of different lymphoma and the over all detection rate was 65%. The lowest detection rate was in MALT-lymphoma. In the second part of this study, we investigated somatic hypermutation in immunoglobulin heavy chain variable region in follicular lymphoma and MALT-lymphoma. Toward this goal, we cloned and sequenced the immunoglobulin heavy chain variable region gene from two patients with follicular lymphomas and one patient with MALT-lymphoma. The obtained gene sequences were compared with the published germline sequences. The heavy chain variable (VH) gene in those three cases of lymphomas shows a much higher ratio of replacement to silent mutations (R/S) in the complementarity determing region (CDRs) (case 1=2.3, case 2= case 3=3) than the framework regions (FRs) (case 1=1.4, case 2=1.9, case 3=0.63), strongly suggesting the occurrence of frequent somatic mutations in the CDRs. Such mutation distribution patterns also suggest that the majority of tumor cells had been positively selected through their antigen receptor.
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