Comparative Genomic Hybridization on Immunophenotypically Pure, Flow Sorted Plasmocyte Populations Shows an Improved Detection of Chromosomal Abnormalities in Monoclonal Gammopathy of Undetermined Significance and Multiple Myeloma.
Authors: Maes Brigitte , Franke Sabine , Voets Greet , Hensen Karen , Jongen Hanne , Peeters Veerle , Cartuyvels Reinoud , Bries Greet , Verhoef Gregor , Rummens Jean-Luc 
Published: October 2019

Abstract
Multiple myeloma (MM) is a clonal plasmocyte disorder often preceded by a premalignant condition, monoclonal gammopathy of undetermined significance (MGUS). Chromosomal abnormalities in MM are complex, of which some are at present considered the most important prognostic indicators in MM. Their role in the pathogenesis of MM and in the transformation from MGUS towards MM is however not fully understood, at least partly due to the low-level bone marrow infiltration hampering cytogenetic analysis. In this study, a unique combination of techniques was used to allow whole genome screening for numerical chromosomal abnormalities of immunophenotypically pure, aberrant plasmocyte populations selected from the bone marrow by flow cytometry. Fourteen bone marrow samples from patients with either MM (n= 11) (first diagnosis or relapsing) or MGUS (n=3), with plasmocyte percentages as low as 1 % (range 1 – 57 %), were analyzed. Aberrant/monoclonal plasmocyte populations were identified based on the expression of CD56 and/or light chain restriction within a CD138+/CD38++ gate and sorted using a FACSAria® (BD, US). Whole genomic DNA was extracted from the CD56+or−/κ or λ+ plasmocyte populations, amplified with degenerate oligonucleotide primer-PCR and used for comparative genomic hybridization (CGH). CGH results were confirmed by interphase fluorescent in situ hybridization. All cases (MM and MGUS) showed the presence of multiple chromosomal abnormalities with a minimum of 5 each. All chromosomes were at least one time involved. Relapsing MM showed a mean number of chromosomal changes of 10 compared to 7 and 6 in newly diagnosed MM and MGUS cases respectively, suggesting karyotypic instability during the course of the disease. The presence of chromosomal aberrations known to be frequently occurring in MM, was confirmed such as gains of 1q (4/11, 36%), 7q (5/11, 45%) and 5q11-q32 (7/11, 64%) and losses of 13q (6/11, 55 %) and 16q (4/11, 36%). This study also showed a high incidence of chromosomal abnormalities not previously or only rarely described in MM, such as gains of 4q11-q22 (9/11, 82 %) and 8q21-q23 (5/11, 45%) and loss of 20q (5/11, 45 %). Some abnormalities were detected in both MM and MGUS, such as gain of 5q and loss of 16q, and may be considered very early, primary aberrations. Gain of 4q11-q22 was detected in almost all MM cases but in none of the MGUS cases, suggesting this abnormality might be involved in disease progression. These results indicate a higher complexity and diversity of chromosomal abnormalities in both MM and MGUS, than has already been described. CGH on flow sorted, immunophenotypically pure, aberrant plasmocytes allows an adequate genetic analysis of MGUS, which should ultimately result in the identification of genetic changes involved in the transformation of MGUS towards MM.