Molecular Cytogenetic Studies of Complex Karyotypes in Myelodysplastic Syndromes (MDS): Conventional Cytogenetics, FISH and Multiplex FISH (mFISH/mBAND)

Karyotype represents one of the most valuable prognostic markers for MDS patients with profound impact on differential diagnosis and therapeutic decisions despite of the fact that it is a heterogenous patient population with diverse pathogenesis and clinical course. In approximately 10–20% of MDS patients complex chromosomal aberrations (CCA; three or more numerical/structural cytogenetic aberrations) are present at diagnosis and this finding is associated with the treatment resistance and poor outcome. Precise identifications of chromosomal regions involved in CCA could help in detection of cryptic, recurrent, prognostically relevant aberrations and in identification of candidate genes involved in leukemogenesis and progression of the disease. In this study we used various modifications of molecular cytogenetic techniques to specify CCA more precisely and to determine exact localization and frequency of chromosomal breakpoints. During the last seven years we examined bone marrow samples of 630 adults with MDS and in 81 of them (12,9%) complex karyotypes were found (in 73 patients at diagnosis and in 8 cases during the disease progression). Out of 73 patients with CCA at diagnosis were 43 males and 30 females, age range 22 to 82 years (median 65,5 years). Primary MDS was diagnosed in 63 (86.3%) of them, in 10 cases therapy related MDS was ascertained. Diagnoses according to the WHO classifications: MDS NS 3x, RA 2x, RA/RCMD 6x, RCMD 5x, RAEB I 20x, RAEB II 20x, MDS-AML 16x and RARS 1x. 63 patients died, 10 patients are still alive (four of them after stem cell transplatation). Chromosomal aberrations found by G-banding were verified by FISH with appropriate locus specific probes (Abbott-Vysis, Des Plaines, Illinois, USA), and by multiplex FISH (mFISH/mBAND) with the “XCyte” probe kits (MetaSystems, Altlussheim, Germany).

Deletion of 5q31 region was proved in 63 patients (86.3%). In 29 cases mFISH analyses showed that parts of deleted No. 5 were translocated into other chromosomes. The most recurrent chromosomal partners of No. 5 in unbalanced translocations were Nos. 17 (9x), 12 (5x), 7 (5x), 2 (4x) and 3 (4x). Monosomy of No. 5 was confirmed in one case only, thus it was proved that monosomy 5 is not separated diagnostic unit in CCA. Except No. 5 the most frequently involved in CCA were chromosomes 7 (36x), 3 (33x), 12 (31x), 17 (31x) and 11 (23x). Presence of CCA at diagnosis was connected with poor response to the therapy and with short overall survival. For survival analysis patients were classified into three groups according to the molecular cytogenetic findings:

1. del(5)(q31) and additional CCA – 33 cases,

2. deleted chromosome 5 involved in CCA – 29 cases,

3. CCA without deletion of 5q – 10 cases.

The worst overall survival was found in group 2 (median 3 months), followed by group 1 (median 5 months) and 3 (median 6 months).

Patients with MDS and CCA should be considered as a unique entity with extremely poor prognosis. Exact molecular cytogenetic analysis by multiplex FISH of all chromosomal aberrations which are involved in CCA will lead to selection of the patients into new prognostic groups. Exploring the underlying mechanism of leukemogenesis could improve the therapeutic outcome for these patients and could contribute to the development of a new targeted treatment strategy.