Incidence of cytogenetic aberrations in two B lineage subpopulations in multiple myeloma patients analyzed by combination of whole-genome profiling and FISH.

Multiple myeloma (MM) is an incurable malignant disease of the terminal developmental stage of B-lymphocytes. While genetic heterogeneity of MM is widely described, little is known about its genetic basis as well as primary damage during plasma cells (PC) development. In this study, we focused on genome-wide screening of DNA copy number changes using oligonucleotide-based array-CGH together with I-FISH of the IgH locus rearrangements in pair samples of bone marrow B-cells (CD19+) and CD138+ PC from newly diagnosed MM patients. The IgH disruption was found in 8.9% (4/45) of CD19+ samples and in 57.8% (26/45) of CD138+ samples. The genomic profiling using array-CGH identified copy number alterations (CNAs) in 10% (2/20) of CD19+ samples in regions known to be important for MM pathogenesis. In contrast, we found CNAs in 100% (16/16) of CD138+ samples. Most common chromosomal abnormalities were trisomies of odd-numbered chromosomes (3, 5, 7, 9, 11, 15, 19 and 21), gain 1q, gain Xq and monosomy of chromosome 13. We did not find any correlation between incidence of CNAs in CD19+ and CD138+ cells. In conclusion, effective utilization of FISH and array-CGH can identify genetic lesions in premalignant stages leading to better understanding and characterization of MM.

Prognostic significance of morphological assessment of plasma cells in multiple myeloma.

Multiple myeloma (MM) is a hematological malignancy caused by clonal proliferation of malignant plasma cells (PC). The aim of the work is to determine prognostic significance of morphological subtypes of PC in relation to overall treatment response, long-term survival and other conventional prognostic parameters. One hundred and thirty-nine newly diagnosed MM patients who underwent autologous transplantation in clinical trials conducted in one center were included. Percentual representation of subtypes of plasma cells in bone marrow was measured based on progressive analysis of nucleolus, nuclear chromatin and ratio of nuclei to the volume of cytoplasm (N/C ratio) creating 8 subtypes P000-P111 and four subclassifications of cells. Mature plasma cells (P000, P001) were found in 42.4% of patients; proplasmocytes I (P010, P011, P100) in 38.1% of patients, and proplasmocytes II (P101, P110) in 19.4% of patients. Patients who reached treatment response after autologous transplantation had statistically significant lower frequency of mature plasma cells than patients with no treatment response (median 24.0% vs. 36.0 %; p=0.032). Patients with mature plasma cells of subtype P000 an patients with value P000 ≥ 37% (median 46.8 months vs. 77.8 months; p = 0.020). Patients with proplasmocytes II subtype P110 rings valuable prognostic information and correlation with other prognostic factors as well as total treatment response and survival in MM patients who underwent autologous transplantation.

Centrosome amplification as a possible marker of mitotic disruptions and cellular carcinogenesis in multiple myeloma.

Centrosome amplification (CA) as a potential marker of mitotic disruptions in multiple myeloma (MM) was investigated in two populations of B-cell lineage: B-cells and plasma cells (PCs). Using immunofluorescent staining, it was shown that CA in B-cells is present in 3.2+/-2.5% in healthy donors versus 9.9+/-7.9% in MM patients (p<0.0001). Based on the calculated threshold value of CA in B-cells, 37% (14/38) of MM patients were positive. There was no significant correlation between CA-positive MM cases (based on PC samples evaluation) and the occurrence of cytogenetic abnormalities in PCs, including del(13)(q14), del(17)(p13), gain(1)(q21) and hyperdiploidy.

Negative prognostic significance of two or more cytogenetic abnormalities in multiple myeloma patients treated with autologous stem cell transplantation.

Malignant plasma cells in multiple myeloma (MM) are frequently characterized by complex karyotypes and chromosome instability. These cytogenetic changes are considered important prognostic indicators in MM patients. We have studied samples from 68 patients with newly diagnosed MM who were treated with high-dose chemotherapy and autologous stem cell transplantation. G-banding revealed abnormal karyotypes in 14 of 55 patients (25%) who had informative conventional cytogenetics. The combination of cytoplasmic immunoglobulin light chain labeling and interphase fluorescent in situ hybridization (cIg-FISH) revealed the presence of genetic aberrations in 53 of 68 patients (78%). Chromosome 13 abnormalities were found in 33 patients (50%) and IgH rearrangements in 36 patients (56.25%). In IgH positive patients we performed subsequent examinations of IgH affecting translocations t(4;14) and t(11;14) and we found translocation t(11;14) in 8 patients (12.5%) and t(4;14) in 10 patients (15.5%). The occurrences of others chromosomal abnormalities with known prognostic impact in MM were as follows: del(17)(p13) was present in 5 patients (9.8%) and gain 1q21 in 14 patients (36%). Analysis of survival of patients with different cytogenetic abnormalities revealed shorter overall survival (OS) in patients with IgH rearrangements (p=0.020) and trend to shorter OS in patients with gain 1q21 (p=0.064), respectively. Remarkably, patients with two or more aberrations had significantly shorter overall survival (p=0.001), time to progression (p=0.036) and progression free survival (p=0.008). Our results show a high incidence of chromosomal abnormalities in MM patients and confirm the prognostic impact of selected chromosomal aberrations as well as cumulative effect of multiple cytogenetic changes occurring simultaneously.