Molecular analysis of the leukaemic B cell in adult and childhood acute lymphoblastic leukaemia

Abstract

Immunoglobulin heavy chain gene (IgH gene) rearrangements are found in the majority of cases of B-lineage acute lymphoblastic leukaemia (ALL). We have examined bone marrow samples taken at presentation or relapse from 109 patients (79 adults and 30 children) and have performed sequence analysis of the complementarity determining region 3 (CDR3) on 65 alleles from 54 patients. We aimed to define immunoglobulin heavy chain (IgH) variable segment family use and investigate biological and structural features of the B cell in adult and childhood ALL. Using the FR1 fingerprinting method, a rearranged band was identified in 70 (89%) of 79 adult ALL and in 29 (97%) of 30 childhood ALL. This study found no preferential use or selection of IgH VH genes and no statistically significant structural differences between normal and leukaemic B cells in either adult and childhood ALL. An equal proportion of amplifiable cases of adult and childhood ALL uses more than one VH family gene (24/70, 34%, and 8/29, 27.5%, respectively). There were no significant differences in the structure or size of the CDR3 region and the variable (V) or joining (J) segment use in ALL patients compared to normal B cells. We observed that the N2 region was shorter than N1 in children whereas the opposite was observed in adults (not statistically significant). The J4 segment was a more common rearrangement in children than in adults, and in both groups J4 was more frequently associated with multiple D segment VDJ rearrangements. An increase in VH6 use in leukaemic alleles compared to normal B lymphocytes (2%) was observed but it was not statistically significant in our group of patients. Amongst children and adults, in-frame CDR3 junctions occurred in 78% and 64% of rearranged alleles, respectively, compared to 75% of in-frame sequences reported by others to occur among normal B cells.

Mechanisms of relapse in acute leukaemia: involvement of p53 mutated subclones in disease progression in acute lymphoblastic leukaemia

“…Resistance to chemotherapy-induced apoptosis may confer a survival advantage to a p53-mutated subclone which by clonal expansion could mediate relapse. This may be of biological importance in ALL where previous studies have demonstrated oligoclonality in 25Ð30% of cases at diagnosis (Beishuizen et al, 1991;Coyle et al, 1996). We did not find a higher incidence of p53 mutations at relapse of AML.…”

Immunoglobulin heavy-chain gene rearrangement in adult acute lymphoblastic leukemia reveals preferential usage of JH-proximal variable gene segments

“…Information on 102 alleles (in adults) and 91 alleles in children had been previously reported in a preliminary study. 39 Except for V H 6, the profile appears to be similar to the germline V H (rearrangable) profile.…”

“…For comparison, previous data on V H family usage in childhood B-lineage ALL (91 clones), 39 B-cell lymphoma (36 clones), and CLL (56 clones) have been displayed in Table 4 and Figure 2C. The childhood B-lineage ALL profile appears to be most similar to the adult pre-B ALL profile, with the exception that V H 6, though overrepresented (10 of 91; 10%), is not as greatly elevated as in the adult ALL group.…”

“…39 In patients with less than 5 g DNA total (fewer than 30% of adult ALL patients) the IgH pattern was analyzed by radionucleotide-incorporated PCR. 41 In brief, 6 PCR reactions, spiked with the radionucleotide ␣ 32 P-deoxycytidine triphosphate, were set up per sample, by means of 1 each of 6 sense family-specific (V H 1 through V H 6) framework 1 primers in combination with an antisense J H primer, as previously described.…”

“…Here we expand our preliminary study 39 and report on the immunoglobulin IGH gene analysis carried out on 127 adult ALL patients (245 alleles) with the use of IGH fingerprinting to define the V H gene pattern of rearrangement. Our aim was to investigate the pattern of V H usage in ALL patients, free of antigen-driven selection pressure, and test the hypothesis that preimmune B cells (as in adult B-lineage ALL clones) undergo random, nonbiased, nonrestricted V H segment usage.…”