Introduction. The use of thrombopoietin receptor agonists, especially eltrombopag, in the treatment of aplastic anemia (AA) patients who did not respond to the previous immunosuppressive therapy (IST), is accompanied by the development of a hematological response in 40–60 % of patients.Aim — to study the effi cacy of using eltrombopag in treatment programs for AA patients refractory to previous IST.Methods. The study included 20 AA patients who were treated at the National Research Center for Hematology from 2015 to 2020. These patients did not respond to the conducted IST (ATG + CsA). Eltrombopag was administered at a dose of 150 mg/day. The results of treatment were assessed at 3 and 6 months: the achievement of hematological improvement, partial and complete remission, as well as the identifi cation of possible clonal evolution were determined.Results. Eleven out of 20 (55 %) patients responded to treatment: 2 patients developed hematological improvement, 6 patients — partial remission, 3 patients — complete remission. All 11 patients responded to treatment within 12 months from the start of eltrombopag, but further positive dynamics of hematological parameters are possible. The median duration of treatment with eltrombopag was 11 (1–48) months. Most of the patients were treated with eltrombopag in combination with CsA. The duration of the course of treatment with eltrombopag depended on the response received (stable hematological improvement, remission, as well as the detection of clonal evolution) or its absence and the need for ATG or BMT. An aberrant karyotype was found in 2 AA patients who received eltrombopag: in one patient monosomy of chromosome 7 was detected 1 month after the start of treatment, in another patient, 37 months later, a clone with a derivative of chromosome 16 from t(1;16) and subclone with complex disorders of the karyotype without signs of myelodysplasia in the bone marrow.Conclusion. The inclusion of the TPO receptor agonist eltrombopag in the treatment program for AA patients allows for a stable hematological response and remission of AA for patients who have not responded to IST. The effectiveness of eltrombopag is determined by adherence to the treatment algorithm, the optimal duration of the course, and the dose of the drug used. There is still a need for long-term observation of the patient and control morphological and cytogenetic studies.

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.

The additional molecular and chromosomal abnormalities (ACA) in Phositive cells usually considered as a genetic marker of chronic myeloid leukemia (CML) progression. 457 patients in different CML phases received tyrosine kinase inhibitors (1st and 2nd generation) were studied. During therapy 50 cases with additional chromosomal abnormalities in Ph+ clone (22 of them in chronic CML phase) were revealed (median follow-up from CML diagnosis – 117 months, median imatinib therapy – 62 months). 86 % of patients in chronic phase with Ph+- cell abnormalities were cytogenetic resistance, and their 5-years overall survival was 80 % which was significantly lower than in patients without ACA (p < 0.005). The treatment results depend on chromosomal abnormalities detected. In patients with additional chromosome 8 imatinib therapy is effective, although complete cytogenetic response (CCR) is achieved only in the later therapy stages. In patients with additional translocations CCR also achieved with imatinib or 2nd generation TKI. Only a third of patients with additional Ph-chromosome or BCR/ABL amplification achieved complete suppression of Ph+ clone using 2nd generation TKI. The presence of additional chromosome 7 abnormalities and complex karyotype disorders involving isochromosome i(17)(q10) are poor prognostic factors of TKI treatment failures.

The types and incidence of chromosome aberrations in patients with multiple myeloma (MM) and their correlations with its clinical course and prognosis were studied. Standard cytogenetic studies of bone marrow cells and fluorescent in situ hybridization (FISH) with delivery of DNA probes to locuses 13q14, 14q32, 11q13, and 4p16 were carried out in 100 patients with MM before cytostatic therapy, The incidence of immunoglobulin heavy chain locus (14q32) restructuring was 50%, of t(11;14)(q13;q32) translocation 30%, of t(4;14)(p16;q32) translocation 15%, and of 13q14 deletion (del) 44%. None of the studied cytogenetic aberrations were detected in 30% cases. A significant relationship between disease prognosis and cytogenetic aberrations was revealed. Three groups of prognosis in myeloma have been distinguished: t(11; 14)(q13;q32) determine a favorable prognosis with overall survival (OS) 80% (p <= 0.001); 13q14 del is associated with development of resistance to standard therapy for MM in 18 of 25 patients; restructuring determines an intermediate prognosis in MM with OS 60% (p <= 0.05); and translocation is the most unfavorable aberration in MM (OS = 0%, median 8.5 months; p <= 0.001). Relationship between cytogenetic aberrations and clinical course and overall survival of patients is proven.

AIM To analyse incidence rate of chromosomal aberrations in myelodysplastic syndromes (MDS), specification of clinicomorphological features of some cytogenetic variants. MATERIAL AND METHODS Chromosomal analysis by the method of G-differential staining of chromosomes was made in 209 patients with different variants of MDS. RESULTS; Clonal chromosomal aberrations occured in 60.8%. The following aberrations were found most frequently: deletion of the long arm of the chromosome 5 (del(5q)) - 34.6%, trisomy of chromosome 8 (14.1%), monosomy of chromosome 7 (13.4%), aberrations 3q21q26 (12.6%), aberrations of a long arm of X-chromosome (4.7%), the absence of Y-chromosome (3.1%). Complex aberrations of karyotype were found in 13.5% cases. Chromosomal aberrations determined not only clinical and morphological features but also the prognosis of the disease. CONCLUSION Cytogenetic examination is an essential component of MDS patients examination. It allows more precise classification of MDS variant and prognostification of the disease course.

AIM:To determine the type and rate of secondary chromosomal aberrations and role in pathogenesis of mantle cell lymphoma (MCL).MATERIAL AND METHODS:Standard cytogenetic examination (SCE) and fluorescent in situ hybridization (FISH) with tests for 11q22/ATM, 13q14, 17p13/p53, 9p21/p16 and chromosome 12 centromere were made in 28 patients.RESULTS:Secondary chromosomal aberrations were detected in 22 patients. Chromosomal abnormalities occurring in more than 2 cases include deletions 6q15-q23 (53% cases); 11q22/ATM (50%); 9p21 (36%, in half the cases deletion 9p21 was biallele); 13q14 (32%); trisomy 3/3q, monosomy 20 and deletions/translocations 1q and 1p (27% and 20%) and deletion 17p13/p53 (18%). Trisomies 12/12q, 6 and 18/18q, monosomies 2 and 16, deletions/translocations 2p and 16p were found in 2 cases each. The FISH technique identified 9 chromosomal anomalies missed at SCE. Deletions 6q15-q23 were seen in patients with privalent lesions of lymph nodes. Deletions 11q, 13q14, 9p21 and 17p13 occur more frequently in transformation and the blastoid variant. Monosomy 20 was found only in patients with large cell transformations. This was the only cytogenetic defect characteristic for transformed cases, in contrast to denovoblastoid ones. Thus, deletions 6q, 11q23, 13q14 and 9p21 are typical for MCL. Deletions 9p21 and 17p13 are more characteristic for large cell variants of the tumor. Deletion 17p13 occurs at the terminal stage of the disease in rapidly growing tumor mass and resistance to chemotherapy. FISH technique is effective in detection of submicroscopic rearrangements especially small deletions. No significant differences were found between transformed and de novo blastoid cases. This shows that the blastoid variant is not a specific biological form, it is rather a less typical manifestation of the disease due to some preclinical cytogenetic disorders.CONCLUSION:Progression and transformation of MCL are related to mutation of proapoptotic genes and genes proteins of which are inhibitors of active complexes of D1 cycline. Specification of these mechanisms requires further investigations in patients with different clinicomorphological forms of the disease at various stages of the disease.

AIM To genotype tumor cells in the recurrence of leukemia after allogenic transplantation of bone marrow (TBM). MATERIAL AND METHODS Standard cytogenetics and fluorescent hybridization in situ (FISH) with a probe to the centrometic sites of X/Y chromosomes were used in examination of 2 patients with acute promyelocytic and acute non-differentiated leukemia after allogenic TBM from donors of the opposite gender. Bone marrow was studied 1, 2, 3, 6, 9, 12, 15, 17, 18 months after the transplantation. RESULTS One of the patient in leukemia recurrence there were 72% cells with one X chromosome with unknown origin. 28% donor cells were with genotype XX. The primary archival cytological sample of the recipient's bone marrow 68% cells did not contain Y chromosome. Thus, the clone with Y loss is the recipient's clone and leukemia after transplantation developed from the recipient's cells. The other patient had only 8% dividing cells with her karyotype XX with translocation t(10;11) while 92% metaphases were donor's ones; the interphase cells ratio was 75% of host cells and 25% donor cells. This confirms leukemia origin from the recipient's cells. CONCLUSION High sensitive quantitative method FISH indicates a true correlation between the host and donor cells and is a method of choice for genotyping leukemic cells in recurrence after transplantation of bone marrow. While standard caryotyping depends on mytotic activity of donor and host cell populations, use of only one cytogenetic test for determination of leukemia origin after TBM may provoke diagnostic errors.