[Repeated haploidentical allogeneic hematopoietic stem cell transplantation with TCR αß/CD19 depletion in patient with primary myelofibrosis. Case report].

Indications of allogeneic hematopoietic stem cell transplantation (allo-HSCT) in patients with primary myelofibrosis are intermediate-2 and high-risk group of DIPSS (Dynamic International Prognostic Scoring System), beginning of the disease in childhood. The other adverse factors affect engraftment and survival after allo-HSCT, example partialy matched donor. But the result of allo-HSCT from matched related donors and result of allo-HSCT from haploidentical donors are comparable. The method for haploidentical hematopoietic stem cell transplantation is T-cell-depletion. This is clinical case of T-cell-depleted haploidentical hematopoietic stem cell transplantation in patient with primary myelofibrosis, the diagnosis was established in childhood.

[Development of program therapy for patients with acute myeloid leukemia under the age of 60 years, based on the principles of differentiated effects].

AIM: To analyze the results of treatment in patients with acute myeloid leukemia (AML) within protocols AML-17 and modified AML-17 (mOML-17) as part of two consecutive pilot studies in order to develop the best treatment strategy for AML patients aged below 60 years. MATERIALS AND METHODS: The study included 89 AML patients who were aged below 60 years and received treatment within the AML-17 and mOML-17 protocols. Cytogenetic and molecular genetic studies were performed in all patients. The presence of mutations in the FLT3, NPM1, CEBPa genes was assessed by fragment analysis. 35 patients underwent a study for mutTP53, mutRUNX1 using next generation sequencing (NGS). The minimum residual population of tumor cells was evaluated by multicolor flow cytometry. Statistical analysis was performed using the procedures of the SAS 9.3 package. RESULTS: Complete remission (CR) was achieved in 89.7% of patients treated with intensive chemotherapy (CT) courses and in 52.4% of patients treated with low-dose CT courses. 8.8% of intensively treated patients were refractory to therapy, and 38% did not respond to low-dose exposure. The early mortality rate was 3%. The overall survival and disease-free 3-year survival for patients included in 2 consecutive studies was were 60% and 67%, respectively. The level of minimal residual disease (MRD) after the first course of induction CT was an important prognostic indicator. The three-year relapse-free survival for patients in whom CR was achieved after the first course of induction CT and in whom MRD was not detected (MRD-negative status was obtained) was 90% compared to 43% for patients who were MRD positive after the first course of induction CT (p=0.00001). CONCLUSION: The key factor that significantly affects the long-term results of therapy is the rate of MRD after the first course of induction CT.

[Multipotent mesenchymal stromal cells application for acute graft versus host disease treatment].

AIM: Analysis of the effectiveness of the MSCs aministration as the second- or third-line therapy of acute GVHD (aGVHD) resistant to glucocorticosteroid treatment. MATERIALS AND METHODS: The study included 35 patients who received MSCs obtained from the bone marrow of healthy donors as a treatment of steroid-resistant aGVHD. The clinical parameters of patients, MSCs cultural characteristics, the MSC expression profile for various genes including those involved in immunomodulation, expression of cells surface markers, the source of MSCs, as well as the frequency and number of MSC administrations were analyzed. RESULTS: Response to therapy was achieved in 74% of cases, a complete response was reached in 13 (37%) patients, partial response/clinical improvement was demonstrated in 13 (37%). This treatment was ineffective in 9 patients. The prediction of a group of patients with good response to MSC therapy turned to be impossible. The differences between the effective and ineffective for the GVHD treatment MSCs samples were found. The effective ones were characterized with a decreased total MSCs production and an increase in the main histocompatibility complex and PDL-1 antigens expression. CONCLUSION: These data allow to select optimal samples for aGVHD treatment that can improve clinical results. aGVHD treatment with MSCs has shown efficacy comparable to other treatment approaches. Given the low percentage of complications and the absence of significant adverse effects, MSC therapy seems to be one of the optimal approaches to the treatment of resistant forms of GVHD.

Results of program acute myeloid leukemia therapy use in National Medical Research Center for Hematology of the Ministry of Health of Russian Federation.

AIM: To analyze treatment results of 172 patients with acute myeloid leukemia (AML) aged 18-60 years in National Medical Research Center for Hematology of MHRF. MATERIALS AND METHODS: Inductive and consolidation program for 139 (80%) patients was based on a standardized protocol: 4 courses "7+3" with different anthracycline use (2 courses of daunorubicin, idarubicin, mitoxantrone) and continuous use of cytarabine on the second inductive course. In 20% of patients cytarabine courses at the dose of 1 g/m2 2 times a day for 1-3 days combined with idarubicin and mitoxantrone were used as two consolidation courses. Allogenic bone marrow transplantation was performed in the first complete remission (CR) period in 40% of patients. RESULTS: The frequency of CR achievement in all patients was 78.6%, refractory forms were observed in 13.9% of patients, early mortality - in 7.5% of patients. Seven-year overall survival (OS) rate was 40.7%, relapse free survival (RFS) - 43.2%. When estimating effectiveness depending on cytogenetic risk group it was demonstrated that 5-year OS and RFS in patients with translocation (8; 21) cannot be considered as satisfying, it accounted for 50 and 34%, respectively. At the same time in patients with 16th chromosome inversion (inv16) these characteristics accounted for 68.6 and 63.5%. Acquired results forced reconsidering of the consolidation program in AML patients of this subgroup. The median time to allogenic blood stem cells transplantation (allo-BSCT) in patients with first CR was 6.5 months that was taken as a reference point in landmark analysis of patients in whom allo-BSCT was not performed. Landmark analysis showed that in AML patients of favorable prognosis group allo-BSCT does not significantly reduce the probability of relapse (0 and 36%) and does not influence RFS (33 and 64%). In patients of border-line and poor prognosis allo-BSCT significantly reduces relapse probability (26 and 66%; 20 and 100%) and significantly increases a 7-year RFS (68.7 and 30%; 45.6 and 0%). Allo-BSCT also results in significant RFS increase and reduces the probability of relapse (25 и 78%) in patients in whom CR was achieved only after the second induction course. At the same time allo-BSCT does not influence patients who achieved CR after the first treatment course: 55 and 50%. CONCLUSION: Multivariate analysis showed that cytogenetic risk group (HR=2.3), time of CR achievement (HR=2.9), and allo-BSCT transplantation (HR=0.16) are independent factors for disease relapse prognosis after achieving CR.

Recombinant MHC Tetramers for Isolation of Virus-Specific CD8+ Cells from Healthy Donors: Potential Approach for Cell Therapy of Posttransplant Cytomegalovirus Infection.

Patients undergoing allogeneic hematopoietic stem cell transplantation have a high risk of cytomegalovirus reactivation, which in the absence of T-cell immunity can result in the development of an acute inflammatory reaction and damage of internal organs. Transfusion of the virus-specific donor T-lymphocytes represents an alternative to a highly toxic and often ineffective antiviral therapy. Potentially promising cell therapy approach comprises transfusion of cytotoxic T-lymphocytes, specific to the viral antigens, immediately after their isolation from the donor's blood circulation without any in vitro expansion. Specific T-cells could be separated from potentially alloreactive lymphocytes using recombinant major histocompatibility complex (MHC) multimers, carrying synthetic viral peptides. Rapid transfusion of virus-specific T-cells to patients has several crucial advantages in comparison with methods based on the in vitro expansion of the cells. About 30% of hematopoietic stem cell donors and 46% of transplant recipients at the National Research Center for Hematology were carriers of the HLA-A*02 allele. Moreover, 94% of Russian donors have an immune response against the cytomegalovirus (CMV). Using recombinant HLA-A*02 multimers carrying an immunodominant cytomegalovirus peptide (NLV), we have shown that the majority of healthy donors have pronounced T-cell immunity against this antigen, whereas shortly after the transplantation the patients do not have specific T-lymphocytes. The donor cells have the immune phenotype of memory cells and can be activated and proliferate after stimulation with the specific antigen. Donor lymphocytes can be substantially enriched to significant purity by magnetic separation with recombinant MHC multimers and are not activated upon cocultivation with the antigen-presenting cells from HLA-incompatible donors without addition of the specific antigen. This study demonstrated that strong immune response to CMV of healthy donors and prevalence of HLA-A*02 allele in the Russian population make it possible to isolate a significant number of virus-specific cells using HLA-A*02-NLV multimers. After the transfusion, these cells should protect patients from CMV without development of allogeneic immune response.

Long-term survival of donor bone marrow multipotent mesenchymal stromal cells implanted into the periosteum of patients with allogeneic graft failure.

The present study involved three patients with graft failure following allogeneic hematopoietic stem cell transplantation (allo-HSCT). We obtained multipotent mesenchymal stromal cells (MSCs) from the original hematopoietic cell donors and implanted these cells in the periosteum to treat long-term bone marrow aplasia. The results showed that in all patients endogenous blood formation was recovered 2 weeks after MSC administration. Donor MSCs were found in recipient bone marrow three and 5 months following MSC implantation. Thus, our findings indicate that functional donor MSCs can persist in patient bone marrow.