Genome Wide SNP Analysis Reveals Frequent Cryptic Clonal Chromosomal Aberrations Including Uniparental Disomy (UPD) in Waldenstrom's Macroglobulinemia.
Authors: Poulain Stephanie , Roumier Christophe , Cheok Meyling , Zoutina Sylvie , Daudignon Agnes , Nibourel Olivier , Soenen Valérie , Lai Jean Luc , Fernandez Jose , Preudhomme Claude , Duthilleul Patrick , Morel Pierre , Leleu Xavier 
Published: October 2019

Abstract 3932

Poster Board III-868

Background

Waldenstrom's macroglobulinemia (WM) is a rare lymphoproliferative disorder characterized by bone marrow (BM) infiltration of lymphoplasmacytic cells that secrete monoclonal IgM antibody. Approximately 50% of patients (pts) with WM exhibit a normal karyotype using either conventional chromosome banding analysis (CBA) or FISH approach. However, CBA is a low resolution method and FISH only target previously described abnormalities. Comparative genomic hybridization (CGH) array delineated the minimal deleted region on 6q deletion, the most frequent aberration in WM, and pointed out the role of NFKB pathway key regulators genes. However, tumour suppressor gene or oncogene involved in WM physiopathology is not known, to date. Partial uniparental disomy (UPD) induced by copy neutral loss of heterozygosity (LOH) are important mechanisms for tumour suppressor gene inactivation or oncogene activation in cancer. Recently, Single Nucleotide Polymorphism (SNP) based arrays had been described as a powerful high resolution method allowing both the detection of LOH and copy number alteration (CNA) analysis in the same experiment, a major advantage over CGH array. Our aim was to identify new CNA and LOH involved in WM pathogenesis using SNP arrays analysis. Material and Methods. BM samples of 12 pts with WM (7 males, mean age: 67 years, 7 symptomatic pts) were analysed. DNA was extracted following CD19 B cells selection. Genome-Wide Human SNP Array 6.0 (Affymetrix chips) was used to detect both LOH and CNA. In 6 pts, paired samples (tumor / normal T lymphocytes) were used as an intra-individual reference to identify germline polymorphisms. In the 6 left pts, CN polymorphisms were excluded using copy-number variants database (http://projects.tcag.ca/variation) and the reference genotyping data from the HapMap project. Size, position and location of genes were identified with UCSC Genome Browser HG18 assembly, LOH and CNA using genotyping console 3.02 software (Affymetrix) and Partek genomic suite. FISH analysis was performed to detect deletion 6q; 13q14, 11q22, TP53, trisomy 4 and 12 chromosomal aberrations using Vysis probes.

Results

SNP array detected 35 CNA (23 gains, 12 losses) in 9/12 pts, with a mean of 2.9 abnormalities per patient (range 0 to 10). 91% of CNA were observed in symptomatic pts. 50% of CNA were < 5 mb in size, the lower limit of detection by CBA. 15 cryptic aberrations were identified among the somatic CNA spread over 7 pts. These 11 gains and 4 losses are identified by *. Deletions were observed on chromosomal segments 1p36*, 3p21.*, 6q16-q27 (2pts), 8p21*, 8p23-q24, 9p21*, 11q22 (2pts), 16p13*, 19p13*. One cryptic homozygous deletion in 13q14 was identified. Gains were observed on chromosomal segments 3p26, 4p16-q35 (3pts), 4q24*, 5p15 (2pts)*, 5p14, 7p22, 12p13-q24, 16p13*, 17q11*, 18p11-q23, 19p13-q13, Xq21* (5pts), Xq22*, Xq25*, Xp22-q28, Xq27-q28*(2pts).

We also identified 42 LOH in 9/12 pts (mean of 3.5 per genome, range 0 to 7) dispersed on 17/23 chromosomes. The LOH observed in the absence of CNA loss are consistent with UPD in 35 cases (83%). 59% of LOH were observed in symptomatic pts. The interstitial or telomeric UPD regions varied in size, from 0.4 to 154mb. Two recurrent regions were identified on chromosome 4 (1.2mb involving genes DCLK2 and LRBA) and 13 (0.8mb involving genes KPNA3, ARL11 and SETDB2), probably subsequent to mitotic recombination. No UPD was observed on chromosome 6q.

SNP arrays detected all FISH and CBA findings, except in one pt with subclonal deletion of TP53. Of the 5 pts with normal CBA or FISH, SNP array detected a CNA and 4 UPD in one patient and 7 UPD in an other patient. The remaining 3 cases (25%) had no detected CNA or LOH.

Overall, SNP array detected a total of 77 genetic aberrations (CNA + LOH) (6.4 per genome, range 0 -16) in this cohort allowing the selection of 976 relevant genes (159 implicated in cellular growth and proliferation, 165 in cancer, 69 in cell cycle).

Conclusion

New cryptic clonal chromosomal lesions were detected using high resolution SNP array in this study. Several abnormalities were recurrent. We described a high frequency of UPD in WM, that might contribute to the inactivation of tumour suppressor genes by mutations or epigenetic alterations and subsequently to the regulation of tumor progression in WM. Further confirmation of the role of these candidate genes is underway.

Disclosures:

Leleu: Celgene: Research Funding; Janssen Cilag: Research Funding; Chugai: Research Funding.