Ατελείς αναδιατάξεις IGHD-IGHJ στη χρόνια λεμφοκυτταρική λευχαιμία

Abstract

CLL is characterized by the existence of subsets of cases with very similar if not identical (stereotyped) VH CDR3 sequences. Stereotyped VH CDR3 are often defined by the selective association of certain IGHD genes in specific reading frames (RF) with certain IGHJ genes. Although stereotypy strongly indicates a role for selection by structurally related epitopes, the possibility that CLL progenitors might exhibit inherently biased IGHD-IGHJ rearrangements, thus eventually accounting for restricted VH CDR3 motifs, cannot be excluded. To gain insight into the mechanisms shaping the IG repertoire and also determine the developmental stage when restrictions in VH CDR3 are imposed, we investigated the molecular features of partial IGHD-IGHJ rearrangements (PR) in a series of 829 patients with CLL. PR were detected in 272/829 cases (32.8%). 198 PR were potentially functional (PF) since they did not carry a stop codon at the IGHD-IGHJ junction. Four PF PR carried IGHD-IGHD gene fusions. Sequence analysis of the PF PR revealed: (i) increased frequency of IGHD2 subgroup genes (94/202 cases, 46.5%), especially IGHD2-2; (ii) equal distribution of the three RFs of the IGHD genes; (iii) increased recombination frequency between genes of the 5´ and 3´ end of the IGHD and IGHJ cluster genes respectively. No associations were identified between the occurrence of PF PR and IGHV gene usage or mutational status of the complete IGHV-IGHD-IGHJ rearrangement (CR) from the coding allele. Comparison of PR vs CR in CLL revealed: (i) selection of the IGHD3-3, IGHD3-10 and IGHD6-19 genes in specific RF in CR, p<0.001 (ii) preferential usage of RF encoding for hydrophilic peptides among CR. Further comparison of PR with 134 PF PRs from B ALL cases revealed that, the overall configuration of PR did not differ significantly in CLL vs. B ALL. Additionally, the comparison of IGHD-IGHJ junctions using the most frequent IGHD2 and IGHD3 subgroup genes in PF PR from CLL vs 82 PF PR from normal B cells (NBC) did not revealed any important differences. Finally, amplification of IGHD-IGHJ-IGHM transcripts in 2/24 B ALL cases carrying exclusively PR revealed the transcription of PRs in at least a proportion of cases. In conclusion, these results document that the early stages of IG gene rearrangements in CLL, B ALL and NBC do not show intrinsic, disease-specific differences. The detailed molecular characterization of the IGHD-IGHJ junctions in PR vs. CR in CLL provides further support that CLL development is not stochastic but directed by selection operating at the IG protein level.