Pangenomic Study In Patients with Persistent Polyclonal Binucleated B-Cell Lymphocytosis (PPBL)

PPBL is a rare disease characterized by a chronic, stable, persistent and polyclonal B-cell lymphocytosis, the presence of binucleated lymphocytes in the peripheral blood and a polyclonal increase in serum immunoglobulin-M. We showed that PPBL was associated with recurrent chromosomal abnormalities and a typical cytogenetic profile including additional extra isochromosome +i (3)(q10) and premature chromosome condensation (PCC). The etiology of PPBL remains unclear. The probable role of tobacco has to be elucidated. Environment factors and persistent viral infection, particularly Epstein-Barr Virus infection, could be involved. Moreover, the possibility of PPBL to evolve toward a clonal proliferation, malignant lymphoma or secondary solid cancer led us to consider PPBL as a premalignant state. Isochromosome i (3)(q10) is routinely detected by conventional cytogenetic analysis (CCA) associated with fluorescent in situ hybridization (FISH) in 71% of the cases. In the peripheral blood, the polyclonal expansion of B-cells is variable, usually low with a mean of 49% of the lymphocytes (7–83%).

In order to study more extensively cytogenetic abnormalities, we used Affymetrix™ Cytogenetics Whole-Genome 2.7M Arrays® in 7 typical PPBL patients. The DNA was extracted from peripheral CD19⁺ B-cells and CD19⁻ cells purified using Miltenyi™ technology (AutoMACS Pro Separator®). The patients had a median age at 49 years (40–59), with a sex ratio at 2:7, smoker in 7/7. Using CCA/FISH, we detected +i (3)(q10) and PCC in 5/7 and 2/7 patients respectively. CD19⁺ B-cells were separated by immunomagnetic cell sorting with a purity of 95% (88% to 98%). We performed DNA arrays on CD19⁺ and CD19⁻ cells in 4 patients, CD19⁺ cells in 2 patients and CD19⁻ cells in 1 patient.

(1) Recurrent gene copy number (GCN) gains were detected on the long arm of chromosome 3 (3q) and identified only in CD19⁺ cells. Recurrent GCN gains were observed in 552/595 (92.8%) genes of 3q. GCN gains concerned either the totality of 3q or only telomeric part of 3q (3q13.3-3qter) or non consecutive genes on 3q. Moreover, we observed that GCN gains did not involve all the CD19⁺ B-cells (mosaicism phenomenon). (2) We observed in CD19⁺ B-cells other recurrent GCN aberrations, including 5 gains on Xp (3 patients) and 16 losses on 6q (2 patients). Non recurrent GCN aberrations involving the whole genome, except chromosomes 5 and 22, confirmed the presence of genetic instability in all patients. (3) Out of the genes amplified on 3q, a few genes have been reported to be implicated in hematological diseases (MLF1/LPP/ATR/BCL6). Interestingly, we identified with a high frequency partial or complete amplification of MDS1 and EVI1 complex locus (MECOM)(3q26.2).

(1) +i (3)(q10) identified by CCA is not a true complete duplication of 3q and can be associated with other recurrent GCN aberrations on chromosomes 6q and Xp. (2) Whereas MDS1 rearrangement (inversion or translocation) has been reported in cases of malignant myeloid disease (MDS, AML) as a poor prognosis factor, the role of MDS1 in lymphoid malignancies has to be elucidated. We are getting MDS1 quantitative PCR and direct sequencing. (3) We confirmed genetic instability in PPBL motivating a long term follow-up of PPBL patients.

No relevant conflicts of interest to declare.