For many decades the main myeloablative conditioning regimen prior to allogeneic hematopoietic stem cell transplant (allo-HCT) consists of busulfan (16 mg/kg) and cyclophosphamide (120 mg/kg). The rate of transplanted-related mortality as a result of toxicity is still high. However alternative regimens are now available. These regimens allow to reduce toxicity and to get favorable outcomes. Aware of the serious toxic complications when using high doses of busulfan initiated the search for alternative programs. We studied conditioning regimen of the therapy with treosulfan 36 g/m²+ cyclophosphamide 120 mg/m² as myeloablative, but low toxic for allo-HCT in 18 patients with hematological malignancies from the high-risk group (allo-HCT has been performed not in first remission, patients with unfavorable cytogenetic abnormalities). The median of the age was 27 years (from 19 to 40 years). During the early posttransplant period no severe toxic complications were registered in the study group. Not bad results were got during 2 years after allo-HCT. The overall survival rate in the group of patients with acute myeloid leukemia accounted for 77.8% and in the group of patients with acute lymphoblastic leukemia – 66.7%. On the day +30 full donor chimerism was detected only in 12 patients (66%). However on day +90 in 8 cases (44.4%) the graft rejection was recorded.

To evaluate late nephrotoxicity of modified program NHL-BFM-90 (mNHL-BFM-90) in adult patients with poor-prognosis of diffuse large B-cell lymphoma (DLBCL). The data of laboratory tests of blood and urine of 40 patients (20 men and 20 women) with DLBCL treated during 2002--2009 years with modified NHL-BFM-90 protocol were analyzed. Comparison group included 19 patients (8 men and 11 women) treated with CHOP or R-CHOP. The median observation period after chemotherapy was 6 years. Laboratory tests of blood and urine were performed before chemotherapy and in 5 years after treatment. Results. Chronic kidney diseases were found in 32 (80%) patients treated with mNHL-BFM-90 vs in 12 (63%) treated with CHOP/R-CHOP. Only 2 (5%) patients had decreased creatinine clearance. The frequency of nephrotoxic complications was significantly higher in patients treated with mNHL-BFM-90 than with CHOP/R-CHOP. Treatment with high-dose chemotherapy was complicated by acute renal failure in 10 (25%) patients, 2 (20%) patients required hemodialysis in comparison with patients treated with standard chemotherapy (p= 0.01). However, in the long term observations there were no significant differences in the nephrotoxic complications between two groups.

The complex interaction between environmental factors and human genes makes a significant contribution to the development of chronic lymphoproliferative diseases (CLPD) in individuals with mutations in various genes, including the innate immune response genes. We examined 51 patients with chronic lymphocytic leukemia (CLL) and 70 multiple myeloma (MM) patients. The control group consisted of 47 healthy persons without hematological malignancies. The patients from control group were matched for gender and age characteristics of CLPD patients. In observed persons there was determined the prevalence of genetic polymorphisms (single-nucleotide polymorphism –SNP) in the innate immune response genes including 20 genetic polymorphisms (single-nucleotide polymorphism –SNP) in 14 genes of the innate immune response. In patients with CLL haplotype AA for the TLR3 gene in -421 position was revealed to occur significantly more often than in the control group (OR: 18.56; p= 0.005). In MM patients there was noted the relation between the risk of the development of the disease and gene polymorphism for IL-10-1082, TLR2-753 and TLR3-421. There were found genetic markers for rapidly progressing forms of CLL and MM (CG + GG haplotype of the gene IL-6 and haplotypes GG + GA gene IL-17A, respectively). It is possible to assume a probable link of gene polymorphisms for IL-6, IL-10, IL-17A, TLR2 and TLR3 with the development of the CLPD and recommend these markers as early additional diagnostic and prognostic criteria.

In the majority of patients with chronic lymphocytic leukemia (CLL) the complete remission (CR) and prolonged progression-free survival (PFS) can be achieved already after the first line chemoimmunotherapy. Nevertheless, a number of tumor cells (more than one per normal 10.000 cells) which determine the essence of minimal residual disease (MRD) is revealed by immunological and genetic methods in the bone marrow of significant number of patients in CR of the disease. MRD-negative status of a patient is associated with prolonged PFS and overall survival (OS) and being the only favorable prognostic factor determined after therapy. The delivery of targeted drugs for the therapy of in CLL (Ibrutinib, Idelalisib, Venclexta) possessing of high efficiency and low toxicity, give the hope to increase the number of MRD-negative remissions, and the change in treatment strategy for the solution of problems of control of residual tumor clone.

Somatic mutations in codons 532--547 of JAK2 exon 12 are highly specific to confirm the diagnosis of polycythemia vera (PV). The aim of this study was to develop the pyrosequencing method for the detection and quantification of JAK2 exon 12 allele burden. The nucleotide sequencing of the JAK2 exon 12 fragment was carried out with the use of “PyroMark Q24”. To verify the presence of mutations, the DNA sequences extracted from the clinical samples were cloned and obtained clones were sequenced with the use of reagents and equipment of the “Applied Biosystems” (USA). A sample containing the mutation was analyzed again after confirming the presence of mutations in clones by Sanger sequencing with “PyroMark Q24” for the quantification of the JAK2 exon 12 allele burden. Among 48 of JAK2 V617F-negative PV patients, who had a high clinical and hematological probability of PV diagnosis, a N542-E543 del mutation of JAK2 in exon 12 has been detected in the 62-years old male patient. The detection and quantification of the JAK2 exon 12 allele burden by virtue of using a pyrosequencing method for this patient was carried out three times from August 2013 to June 2015. Our results show the JAK2 N542-E543 del allele burden to increase more than twice during 23 months of following up of the patient. These data suggest about increase of the number of the transformed clonal cells. The proposed method allows detecting and carrying out quantitative determination of the mutant JAK2 exon 12 allele burden for monitoring the efficacy of the therapy.

Monitoring of minimal residual disease (MRD) proved to be a valuable tool for predicting relapse in patients with acute lymphoblastic leukemia (ALL). However, the universal use of MRD monitoring in routine clinical practice is limited because current MRD assays have some methodological difficulties, high price and ambiguity of assay results interpretation. Here we describe next generation sequencing based system for the detection of clonal rearrangements in immunoglobulin genes loci. We performed testing of the system on 17 initial bone marrow samples from B-ALL patients. We revealed 1 to 6 characteristic immunoglobulin genes rearrangements in each of 16 samples. These results are in a good accordance with the results obtained by traditional BIOMED-2 assay. Further improvement of the reported system will provide highly reliable and sensitive technique for MRD monitoring.

The use of a freeze-dried plasma has a number of advantages over fresh frozen plasma (FFP) as for the provision of transfusion care in the field conditions, as well for the carrying out diagnostic studies of the hemostatic system. The execution of diagnostic tests requires for the availability of Biological Standards, plasma calibrator and control material with high stability of target indices in the storage of lyophilized reagents. Plasma lyophilization process causes significant changes in coagulation indices and is accompanied by an increase in plasma pH. The aim of the study was to obtain a freeze-dried human plasma without the pronounced changes in the activity of clotting factors under lyophilization and storage. The acidification of pooled FFP from at least 20 donors before lyophilization to pH values of 7.0–7.1 by HEPES buffer solution at a final concentration of 50 mM was shown to increase the stability of the hemostatic system factors and retain physiological plasma pH after lyophilisation. In conditions of an accelerated storage just so lyophilized plasmas are characterized by the greater stability of all determined coagulation indices. Thus, the proposed approach makes it possible to obtain storage-stable, lyophilized forms of diagnostic reagents and control materials.

The basic method of the treatment of haemophilia A and B is the replacement therapy by preparations obtained from plasma of donors, or by recombinant DNA technology. Among the major shortcomings of unmodified drugs VIII and IX factors should be allocated between the low time circulation of the drug in the body of the patient and the delivery of antibodies (inhibitors) to a protein preparation that significantly reduces their effectiveness. The improvement of the efficiency and safety of recombinant preparations is achieved by removing B-domain of factor VIII molecule, pegylation or development of preparations of fusion proteins (fusion molecules of the preparation to the albumin or Fc-fragment of IgG). These modifications, especially, enhance the stability of the molecule of the active substance to increase half-life and reduce immunogenicity of recombinant products. Replacement therapy with traditional preparations in hemophilia patients delivering antibodies (inhibitors) to the preparation is ineffective. Taking this into account, in recent years, new preparations have been developed for alternative approaches treatment of hemophilia. There were developed the preparation based on bispecific monoclonal antibody mimicking the function of factor VIII; monoclonal antibodies and aptamer blocking the activity of tissue factor pathway inhibitor, and the preparation based on antisense oligonucleotide blocking mRNA responsible for the synthesis of antithrombin III. The main advantage of these new formulations is that they do not cause the production of antibodies to coagulation factors and so can be used for the treatment of patients with inhibitors in blood.

Even in the tyrosine kinase inhibitors era, the prognosis of patients with chronic myeloid leukemia in myeloid blast crisis remains to be unfavorable. Myeloid blast crisis is characterized to be refractory to cytotoxic agents and high risk of recurrence. We described the patient with chronic myeloid leukemia in myeloid blast crisis who was successfully treated by the combination of the second generation tyrosine kinase inhibitor and 5-azacytidine.

Bernard–Soulier syndrome (BSS) is the rare platelet disorder. It is caused by the deficiency or defect of the glycoprotein (GP) Ib-IX-V complex on the surface of platelets – the main receptor for von Willebrand factor (vWF). Binding of vWF with GP Ib-IX-V complex initiates primary hemostasis and provides platelets adhesion at the site of the injured blood vessel. Dissociation of plasma, vascular and thrombocytic hemostasis gives rise to the development of the microcirculatory or mixed hemorrhagic syndrome. The typical features of such types are considered in the clinical case of the BSS female patient. Diagnostic algorithm for BSS includes the evaluation of functional properties, morphology and biochemical characteristics of platelets with the compulsory use of flow cytometry, allowing to reveal the deficiency of GP Ib-IX-V complex on the surface of platelets.