Aim.To evaluate the efficacy of DSP30 in combination with IL2 in cultivating blood cells/bone marrow/lymph nodes in chronic lymphocytic leukemia (CLL) patients to detect clonal abnormalities.Materials and methods.The study included 50 patients with CLL, all of whom underwent both chromosome banding analysis (CBA) (46 patients with DSP30+IL2 and LPS+TPA; 4 patients with only DSP30+IL2) and FISH with DNA probes to detect trisomy 12 and deletions of 13q14, 11q22 and 17p13.Results.Under cell cultivation with DSP30+IL2 and LPS+TPA, CBA was successfully performed in 41 (82 %) and 38 (83 %) patients. Chromosome aberrations were observed in 36 (72 %) and 15 (33%) cases, while a complex karyotype was detected in 13 (26%) and 5 (11%) cases, respectively. A significant difference was found between the number of metaphases with chromosomal abnormalities obtained by cultivation with DSP30+IL2 and LPS+TPA (V = 490.5, p < 0.05). CBA revealed balanced translocations in 6 patients, with the involvement of the IgH/14q324 locus being confirmed in 4 cases. Unbalanced translocations and various combinations of translocations were detected in 11 and 6 patients, respectively. In 5 cases, according to CBA, the results of 13q14, 11q22, 17p13 deletions identified by FISH were accompanied by balanced or unbalanced translocations in these loci. Unbalanced t(12;16)(q14;q23) — a case of partial trisomy — was detected only by CBA with DSP30+IL2.Conclusions.An abnormal karyotype was detected in CLL patients twice as more frequently under cultivation with DSP30+IL2 compared to LPS+TPA. CBA is an important method allowing the structure of chromosomal abnormalities to be specified and translocations to be identified. As a result, patients running the highest risk of CLL — those with a complex karyotype — can be singled out for selecting an optimal strategy of their management.

Aim. To evaluate the efficacy of DSP30 in combination with IL2 in cultivating blood cells/bone marrow/lymph nodes in chronic lymphocytic leukemia (CLL) patients to detect clonal abnormalities. Materials and methods. The study included 50 patients with CLL, all of whom underwent both chromosome banding analysis (CBA) (46 patients with DSP30+IL2 and LPS+TPA; 4 patients with only DSP30+IL2) and FISH with DNA probes to detect trisomy 12 and deletions of 13q14, 11q22 and 17p13. Results. Under cell cultivation with DSP30+IL2 and LPS+TPA, CBA was successfully performed in 41 (82 %) and 38 (83 %) patients. Chromosome aberrations were observed in 36 (72 %) and 15 (33%) cases, while a complex karyotype was detected in 13 (26%) and 5 (11%) cases, respectively. A significant difference was found between the number of metaphases with chromosomal abnormalities obtained by cultivation with DSP30+IL2 and LPS+TPA (V = 490.5, p < 0.05). CBA revealed balanced translocations in 6 patients, with the involvement of the IgH/14q324 locus being confirmed in 4 cases. Unbalanced translocations and various combinations of translocations were detected in 11 and 6 patients, respectively. In 5 cases, according to CBA, the results of 13q14, 11q22, 17p13 deletions identified by FISH were accompanied by balanced or unbalanced translocations in these loci. Unbalanced t(12;16)(q14;q23) — a case of partial trisomy — was detected only by CBA with DSP30+IL2. Conclusions. An abnormal karyotype was detected in CLL patients twice as more frequently under cultivation with DSP30+IL2 compared to LPS+TPA. CBA is an important method allowing the structure of chromosomal abnormalities to be specified and translocations to be identified. As a result, patients running the highest risk of CLL — those with a complex karyotype — can be singled out for selecting an optimal strategy of their management.

Complex karyotype (CK) in patients with myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML) is a factor of unfavorable prognosis, associated with high frequency of refractory forms and relapses. Precise identification of chromosome abnormalities in CK by conventional cytogenetic analysis (CCA) is limited due to low resolution of this method. Molecular cytogenetic techniques (FISH, mFISH, mBAND) have significantly higher sensitivity and allow to identify complex chromosome abnormalities, marker chromosomes, submicroscopic deletions and specify chromosomes breakpoints. This study included 15 MDS and 11 AML patients whose complex karyotypes were characterized by these methods. mFISH and iFISH are performed in all 26 patients, mBAND is performed in two cases. CCA revealed an average of 7 karyotype abnormalities (from 3 to 21). Structural rearrangements were found in all cases (n = 26): reciprocal translocations (38.4%), deletions (65.3%), additional chromosomal material of unknown origin (69.2%), marker chromosomes (53.8%). Numerical anomalies typical for MDS and AML (n = 26) were found: trisomy 8 (15.3%), monosomy 5 (42.3%), 7 (34.6%) and 17 (11.5%). Molecular cytogenetic analysis revealed additional chromosomal abnormalities and / or additional chromosome breakpoints in 19 cases. Translocations were found: reciprocal in 22 cases, complicated in 13 cases. In karyotype of 23 patients we found chromosome anomalies 5 (76.9%), 7 (57.6%), 11 (34.6%) and 17 (46.1%). The deletion 5q was confirmed in five cases, the deletion of 7q in one case. True monosomy 7 is confirmed in one case. On the results of molecular cytogenetic techniques in all other cases with monosomy 5, 7, and 17 revealed fragments of these chromosomes involved in translocations were combined deletions of loci 5q31, 7q31 and 17p13. All marker chromosomes and chromosomes with additional material of unknown origin were recognized as complex translocations or derivative chromosomes with breakpoints in both arms. Combination of CCA and molecular cytogenetic techniques is necessary for the complete characterization of complex karyotypes in MDS and AML and for precise for characteristic of CK in MDS and AML and determination of exact breakpoints loci of potential oncogenes and tumor suppressor genes.

Aim. To study and compare cytogenetic abnormalities in the bone marrow (BM) and peripheral blood (PB) CD34+ hematopoietic progenitor cells and in the BM mesenchymal stromal cells (MSCs) in patients with myelodysplastic syndrome (MDS).Subjects and methods. The results of a cytogenetic analysis of the total population of BM cells (BMC), CD34+ hematopoietic progenitor cells from BM and PB, and BM MSCs were analyzed in 35 patients (29 patients with MDS and 6 with MDS transformed into acute myeloid leukemia (AML)) and 7 healthy BM donors. Cytogenetic examinations were performed by G-banding of chromosomes and fluorescence in situ hybridization (FISH).Results. The BMC karyotype was abnormal in 17 (49%) of the 35 patients (13 with MDS and 4 with AML); the others were found to have a normal BM karyotype. The FISH analysis confirmed the same cytogenetic abnormalities in the BM and PB CD34+ hematopoietic progenitor cells in all the examinees with an abnormal karyotype. The mean abnormal clone sizes in the total population of BMCs and BM and PB CD34+ progenitor cells did not differ and constituted 65.8, 73.1, and 74.8%, respectively. The patients with a normal BM karyotype had no chromosome abnormalities in the CD34+ cells either. The karyotype of MSCs was analyzed in 23 (19 with MDS and 4 with AML) of the 35 patients. No karyotype abnormalities were revealed in the patients with MDS transformed into AML. There were structural chromosome aberrations in 2 (11%) of the 19 patients with MDS (one with constitutional inv(9)(p13q21) was found to have non-clonal translocation t(2;22)(p10;q11) and the other had a clone with an additional segment of the long arm of chromosome 2 in 35% of the cells. No numerical MSC karyotype abnormalities were detected. A normal MSC karyotype was defined in 7 healthy BM donors.Conclusion. The cytogenetic analysis of hematopoietic and mesenchymal progenitor cells showed that the chromosome abnormalities revealed in these cell populations were different in the patients with MDS. The isolated CD34+ cells displayed the same cytogenetic abnormalities as in a total population of BMC. Examination of the latter could reveal no cytogenetic abnormalities in the majority of the patients. A normal BMC karyotype was detectable in the patients with AML. Two (11%) patients with MDS were found to have structural chromosome abnormalities that differed from those detected in the total population of BMC and in isolated CD34+ cells. The differences of chromosome abnormalities in the hematopoietic and mesenchymal progenitor cells point to the fact that the stromal microenvironment is not part of the abnormal clone in MDS, however, it may be of great importance in the pathogenesis of the disease.