Combined patterns of IGHV repertoire and cytogenetic/molecular alterations in monoclonal B lymphocytosis versus chronic lymphocytic leukemia

Abstract

Background: Chronic lymphocytic leukemia (CLL)-like monoclonal B lymphocytosis (MBL) with (MBL(hi)) or without (MBL(lo)) absolute B-lymphocytosis precedes most CLL cases,the specific determinants for malignant progression remaining unknown.

Methodology/principal findings: For this purpose, simultaneous iFISH and molecular analysis of well-established cytogenetic alterations of chromosomes 11, 12, 13, 14 and 17 together with the pattern of rearrangement of the IGHV genes were performed in CLL-like cells from MBL and CLL cases. Our results based on 78 CLL-like MBL and 117 CLL clones from 166 subjects living in the same geographical area, show the existence of three major groups of clones with distinct but partially overlapping patterns of IGHV gene usage, IGHV mutational status and cytogenetic alterations. These included a group enriched in MBL(lo) clones expressing specific IGHV subgroups (e.g. VH3-23) with no or isolated good-prognosis cytogenetic alterations, a second group which mainly consisted of clinical MBL(hi) and advanced stage CLL with a skewed but different CLL-associated IGHV gene repertoire (e.g. VH1-69), frequently associated with complex karyotypes and poor-prognosis cytogenetic alterations, and a third group of clones with intermediate features, with prevalence of mutated IGHV genes, and higher numbers of del(13q)(+) clonal B-cells.

Conclusions/significance: These findings suggest that the specific IGHV repertoire and IGHV mutational status of CLL-like B-cell clones may modulate the type of cytogenetic alterations acquired, their rate of acquisition and/or potentially also their clinical consequences. Further long-term follow-up studies investigating the IGHV gene repertoire of MBL(lo) clones in distinct geographic areas and microenvironments are required to confirm our findings and shed light on the potential role of some antigen-binding BCR specificities contributing to clonal evolution.

Figure 1. Frequency of CLL-associated cytogenetic alterations (A) and the cytogenetic profile (B) for those IGHV genes most commonly detected in “low-count MBL”(MBL^lo), “high-count MBL” (MBL^hi) and CLL B-cell clones, as assessed by interphase fluorescence in situ hybridization (iFISH).

The three diagnostic categories studied are depicted by different colors (green, MBL^lo; red, MBL^hi; blue, CLL B-cell clones) and the absence vs presence of one vs≥2 chromosomal alterationsperclone, is indicated by empty circles, light colored and dark colored circles, respectively. For each IGHV subgroup, the clones are represented in the Y-axis according to the absolute number of clonal B cells per µL of PB (A) and the percentage of cells genetically altered, by iFISH (B). Different FISH patterns are defined by the following symbols in panel B:, del(13q14.3);, biallelic del(13q14.3);, del(13q14);, trisomy 12; Δ, del(11q);▿,del(17p) and; □, t(14q32); dotted contour lines in panel A highlight those clones phenotypically classified as SLL(small lymphocytic lymphoma); dotted blue lines in panel B indicate cells from the same B-cell clone showing different cytogenetic abnormalities; U = unmutated clones; a = clones with NOTCH1 mutation.

Figure 2. Principal component analysis (3-dimensionalX-Y-Z axis view of PC1 vs PC2 vs PC3, respectively) for comparison of “low-count MBL” (MBL^lo), “high-count MBL” (MBL^hi) and CLL B-cell clones according to the absolute number of clonal B cells/µL and the pattern of cytogenetic alterations (including the percentage of altered cells), using the Infinicyt^TMsoftware.

Overall, MBL^lo, MBL^hi and CLL cases are clustered into groups distinguished by different colors in A: magenta, gray, and black circles (A). The distribution of MBL^lo, MBL^hi, CLL-stage A and CLL-stage B/C clones are coloured differently in B: MBL^lo, green; MBL^hi, red, CLL stage A and B/C light blue and dark blue, respectively (B). The most informative parameters contributing to the best discrimination between 1×1 comparisons of the three groups are displayed in a decreasing order of percentage contribution to each of the principal component (C); Distribution of MBL^lo, MBL^hi and CLL clones among the three major groups defined in panel A by principal component analysis (D); CLL, chronic lymphocytic leukemia; MBL, monoclonal B lymphocytosis; PC: principal component.