MOLECULAR ANALYSIS OF REARRANGED VH GENES DURING B-CELL CHRONIC LYMPHOCYTIC-LEUKEMIA - INTRACLONAL STABILITY IS FREQUENT BUT NOT CONSTANT

Several genetic mechanisms have been shown to diversify the expressed antibody repertoire of commited B lymphocytes. These include V gene re placement, ongoing gene rearrangement and somatic hypermutation. These mechanisms may be operational at discrete points in the B cell differ entiation pathway and generate idiotype diversity in various malignant B cell tumors. In particular, V region mutations have been establishe d as a major mechanism of tumor escape from anti-idiotype immunotherap y in some lymphoma. On the other hand, previous studies on a few selec ted cases have shown that this mutation process does not affect the B cell clone during chronic lymphocytic leukemia. However, to what exten t this intraclonal stability is a general phenomenon during B cell CLL is not clear. Therefore, we randomly selected 6 patients suffering fr om classical B cell CLL (sIgM (+), CD5 (+), CD19 (+)) at different sta ges of the disease and analysed the intraclonal variability of the exp ressed variable region of the heavy chain (VH). After PCR amplificatio n of the cDNA corresponding to the rearranged VDJ regions, the product s were cloned and sequenced. In five cases, multiple clone analysis di d not show any intraclonal variability whatever the stage of the disea se. Furthermore, in a single case, this intraclonal stability was conf irmed during a three year period of time when the disease progressed. The sixth case behaved differently since we found multiple nucleotide substitutions, apparently accumulating as the malignant clone expanded . Besides the theoretical difficulties that these changes can induce d uring immunotherapy, two findings merit further discussion: 1) the dis tribution of the ongoing mutations affecting the VH region was not sug gestive of an antigen driven selection, 2) this intraclonal variabilit y was specific for the VH region, since the VL region showed no intrac lonal variation.