الفهرس 
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Abstract
Multiple myeloma is a heterogeneous clonal B-cell neoplasm that affects terminally differentiated B cells [i.e. plasma cells (PC)] with the accumulation in bone marrow of malignant PC producing a monoclonal immunoglobulin known as a monoclonal component. Plasma cell myeloma is characterized, in addition to the serum monoclonal protein, by skeletal destruction with osteolytic lesions, pathological fractures, bone pain, renal impairments, hypercalcaemia, and anaemia. The genetic pathogenesis of the multiple myeloma and its clinical variants is still poorly understood. Specific non-random chromosomal aberrations have been implicated in the oncogenesis of this heterogeneous group of diseases, in the form of numerical and/or specific chromosomal translocations, which usually dysregulate specific oncogenes. However, cytogenetic analyses are hampered by the low proliferation rate of PC. Karyotypic abnormalities are detected at a frequency of 30% to 50% of myeloma tumours. Based on interphase FISH, several studies have shown that virtually all MM patients have chromosomal abnormalities in their plasma cells.
It is therefore the general aim of this study to further our knowledge about these genetic changes that characterise the pathogenesis of multiple myeloma and its clinical variants in more details. In order to accomplish this goal, we tried to characterise the various acquired chromosomal aberrations by interphase FISH, M-FISH and conventional cytogenetic techniques, assess possible correlations between these particular cytogenetics/molecular cytogenetics abnormalities with the type of the disease represented by the clinical/laboratory findings and finally evaluate the impacts of these chromosomal abnormalities on patients’ survival and outcome.
This study comprised 258 patients with different clinical variants of multiple myeloma, 83% MM, 11% SMM, and 6% MGUS. The median age was 67 years (range 27–93 years). The male: female ratio was 1.3:1 where 58% were male. The median bone marrow PC infiltrations in MM, SMM and MGUS were [25%, 18% and 4%] respectively. IgG represented 59% of all cases whereas IgA represented 27% and Light chain only 12%.
Interphase FISH studies on purified plasma cells from 249 patients were carried out using standard protocols of different probes preparations for detection of both numerical and structural aberrations. There was special emphasis on chromosomes 3, 6, 7, 9, 11 and 17 centromeres as examples of the most common numerical aberrations, and chromosome 13 deletion, rearrangements involving the immunoglobulin heavy chain at 14q32 region using certain probes (IgH dual break apart, 4p16, 6p21, 11q13, 16q23) and p53 gene loss on chromosome 17p13 as examples of structural aberrations.
Multicolour fluorescence in situ hybridization was performed using the commercial SpectraVysionTM Assay on metaphase spread from 25 patients with MM showing complex karyotypes by G-banding technique.
In this series, the abnormality rate using nuc FISH was 99% where only 3/249 cases failed to show abnormalities with any of the probes used. Among the numerical chromosome abnormalities identified in this study, trisomy of chromosome 9 was the highest (60%) followed by trisomies of chromosomes 3, 11, and 7 (48%, 46%, and 41%) respectively and hyperdiploidy was detected in 145 cases (58%). Regarding structural abnormalities, the overall frequency of any IgH rearrangement was 45%, chromosome 13 deletion 41% and p53 gene deletion 6%. The frequency of the specific IgH rearrangement was as follow; 16% t(11;14), 9% t(4;14) 3% t(14;16).
In the current study the numerical chromosome abnormalities were found in MGUS cases (68%), but at lower frequencies than those observed in MM (77%).We have also shown that, there were no significant specific pattern of distribution of individual trisomy among MM, SMM, or MGUS. Furthermore, structural abnormalities were also detected in all categories of the disease but at different frequencies. The abnormality rate in each group was as follow (MM 75%, SMM 59% and MGUS 50%). The frequency of del 13 was much higher in MM group compared to SMM or MGUS (44% VS 21% or 13% respectively) whereas all cases with deleted p53 gene on chromosome 17p13 were MM. On the other hand, the frequency of any IgH split was much lower in MGUS group (25%) compared to the MM and SMM groups (46% and 51%) respectively. Nevertheless, we found a strong inverse relationship between frequency of IgH rearrangement and age where the frequency in ≥70 years group was 38% compared to 62% for <70 years.
This study confirmed the strong association between t(4;14) or t(14;16) and del 13 where 83% of t(4;14) cases and 89% of t(14;16) cases had del 13. We also have confirmed the association of these changes [t(4;14), t(14;16), and del 13] and hypodiploidy, wherein (74%) of t(4;14) cases, (78%) of t(14;16) cases and (51%) of del 13 cases were associated with hypodiploidy. Furthermore, the frequency of these abnormalities in hypodiploidy in relation to hyperdiploidy was strongly significant confirming these associations. We also confirmed the infrequency of del 13 with t(11;14) wherein only 20% of t(11;14) cases had del 13 abnormality.
A striking confirming finding of our study was the strong association between t(11;14) and near diploid and the absence of any other abnormalities with these cases (more than 74% of cases), a pattern not seen for any other IgH rearrangement.
Concerning the clinical/laboratory implications of chromosomal aberrations, the frequency of elevated urea and hypoalbuminaemia in cases with del 13, any IgH rearrangement, t(4;14), and hypodiploidy were higher than with other abnormalities (~52-73%) suggesting the poorer prognosis of these abnormalities but this was only statistically significant for t(4;14).
In this series, increased level of β2 microglobulin was the most obvious parameter significantly associated with both numerical and structural chromosomal abnormalities. Furthermore, we have found that del 13, any IgH rearrangement, t(4;14), t(14;16) and all numerical abnormalities except chromosome 17 trisomy had a strong association with the presence of Bence-Jones protein in urine.
Regarding the influence of chromosomal abnormalities on the overall survival, chromosome 13 deletion, t(4;14), t(14;16), and p53 deletion were clearly associated with a poor survival in our series. Additionally, hypodiploidy showed a trend towards poorer survival but this was not statistically significant, confirming its lesser effect than del 13. Furthermore, none of the individual trisomies in addition to, t(11;14) had any effect on survival.
The value and strength of M-FISH in defining translocations and marker chromosomes have been confirmed in this study and we showed the ability of this technique in improving the characterization of complex or ill-defined karyotypes and verification of the commonly reported abnormalities. In this approach, we were able to elucidate the origin of 23 marker chromosomes in 11/25 cases with complex karyotype and revise the description of more than 100 unbalanced chromosomal changes. These imbalances involved and were widely distributed throughout all chromosomes.
Additionally, in two cases with positive IgH rearrangement involving 14q32 by nuc FISH, other chromosome 14 partners were also identified. In another two cases with no IgH rearrangements by nuc FISH we recognised rearrangements involving chromosome 14 by using M-FISH technique.