ISSN 1866-8836
Клеточная терапия и трансплантация

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Volume 5, Number 3
09/30/2016 12:09:00 pm
Volume 5, Number 3
Editor-in-Chief
Afanasyev B. V. (St. Petersburg, Russia)
Co-Editors-in-Chief
Wagemaker G. (Rotterdam, Netherlands)
Zander A.R. (Hamburg, Germany)
Deputy Editor
Fehse B. (Hamburg, Germany)
Managing Editor
Chukhlovin A. B. (St. Petersburg, Russia)
Editorial Board
Aleynikova O. V. (Minsk, Belarus)
Borset M. (Trondheim, Norway)
Büchner Th. (Münster, Germany)
Chechetkin A. V. (St. Petersburg, Russia)
Fibbe W. (Leiden, Netherlands)
Galibin O. V. (St. Petersburg, Russia)
Hölzer D. (Frankfurt a.M., Germany)
Klimko N. N. (St. Petersburg, Russia)
Kolb H.-J. (München, Germany)
Kröger N. (Hamburg, Germany)
Kulagin A. D. (St. Petersburg, Russia)
Lange C. (Hamburg, Germany)
Mamaev N. N. (St. Petersburg, Russia)
Mikhailova N. B. (St. Petersburg, Russia)
Moiseev I. S. (St. Petersburg, Russia)
Nagler A. (Tel-Aviv, Israel)
Nemkov A. S. (St. Petersburg, Russia)
Paramonov I. V. (Kirov, Russia)
Roumiantsev A. G. (Moscow, Russia)
Savchenko V. G. (Moscow, Russia)
Smirnov A. V. (St. Petersburg, Russia)
Uss A. L. (Minsk, Belarus)
Zubarovskaya L. S. (St.Petersburg, Russia)
In this Issue

Editorial

Short reports

Comparative analysis of different CVC dressing types convenience and economic effectiveness in allogeneic hemopoietic stem cell transplant recipients
Olga Ivanova, Julia Simakina, Marina Ermolova, Julia Oparina, Ekaterina Goncharova, Ludmila Zubarovskaya, Boris Afanasyev
Treatment of refractory intestinal acute GvHD using multipotent mesenchymal stromal cells (MMSC)
Darya S. Dubnyak, Larisa A. Kuzmina, Mikhail Y. Drokov, Natalia A. Petinati, Nina J. Drize, Vera V. Vasilyeva, Olga M. Koroleva, Ekaterina D. Mikhaltsova, Natalia N. Popova, Elena N.Parovichnikova, Valerii G. Savchenko
Alterations in multipotent mesenchymal stromal cells properties: in vitro model of their interactions with allogeneic lymphocytes
Nikolay M. Kapranov, Julia O. Davydova, Nataliya A. Petinati, Maria V. Bakshinskayte*, Irina V. Galtseva, Nina I. Drize, Larisa A. Kuzmina, Elena N. Parovichnikova, Valeriy G. Savchenko
Nursing care for adolescents with graft versus host disease after allogeneic hematopoietic stem cell transplantation
Anastasia A. Kraeva, Irina A. Badanova, Lubov A. Shepeleva, Maria U. Averyanova, Svetlana I. Gortinskaya-Oleschko, Ludmila S. Zubarovskaya, Boris V. Afanasyev
Relapse-free survival in multiple myeloma patients after autologous transplantation of peripheral blood stem cells depending of the content of CD45+CD19+ B cells in the apheresis product
Irina V. Kruchkova, Svetlana A. Sizikova, Galina U. Ushakova, Vera V. Sergeevicheva, Darya S. Batorova, Egor V. Batorov, Natalya V. Pronkina, Andrey V. Gilevich, Elena R. Chernykh
Usage of online platforms for remote evaluation of the quality of life in patients following allogeneic transplantation of hematopoietic stem cells on the territory of Russian Federation and ex-USSR
Darya S. Dubnyak, Mikhail Y. Drokov, Larisa A. Kuzmina, Vera V. Vasilyeva, Olga M. Koroleva, Ekaterina D. Mikhalcova, Natalia N. Popova, Dmitriy E. Vibornih, Sergey O. Khruschev, Elena N.Parovichnikova
Granzyme B expression in T-regulatory cells is a strong predictor of acute graftversus-host disease after day +30 in patients with classic immunosuppression after allo-HSCT
Mikhail Y. Drokov, Elena N. Parovichnikova, Julia O. Davydova, Larisa A. Kuzmina, Irina V. Galtseva, Nikolay M. Kapranov, Vera A. Vasilyeva, Darya S. Dubnyak, Olga M. Koroleva, Ekaterina D. Mikhalcova, Natalia N. Popova, Valery G. Savchenko
Posttransplant ruxolitinib combined with cyclophosphamide for graft versus host disease prophylaxis and relapse prevention in patients with myelofibrosis
Maria V. Barabanshikova, Ivan S. Moiseev, Elena V. Morozova, Julia J. Vlasova, Vadim V. Baykov, Ildar M. Barkhatov, Boris V. Afanasyev
Regulatory T cells affect early relapse following high-dose chemotherapy with autologous hematopoietic stem cell transplantation in multiple myeloma patients
Egor V. Batorov, Marina A. Tikhonova, Irina V. Kryuchkova, Vera V. Sergeevicheva, Svetlana A. Sizikova, Galina Ju. Ushakova, Dariya S. Batorova, Andrey V. Gilevich, Elena R. Chernykh
Successful usage of fludarabine for treatment of partial immunologic hematopoietic graft rejection after allogeneic hematopoietic stem cell transplantation in pediatric patients
Alexandra E. Burya, Kirill I. Kirgizov, Veronica V. Konstantinova, Ekaterina A. Prinstanskova, Natalia V. Sidorova, Bazarma B. Purbueva, Olesya S. Fink, Andrey A. Bologov, Elena V. Skorobatova
Novel HLA-alleles identified in populations of the Northern Caucasus
Maria A. Loginova1,2, Igor V. Paramonov2, Andrey V. Rylov2

1 The Rosplasma Center of Medical Research & Industry, Kirov, Russia
2 Kirov Research Institute of Hematology and Blood Transfusion, Киров, Россия
Functional disparity of graft-derived T lymphocytes: experimental data
Mikhail Y. Drokov, Elena N. Parovichnikova, Darya S. Dubnyak, Larisa A. Kuzmina, Irina V. Galtseva, Julia O. Davydova, Nikolay M. Kapranov, Vera A. Vasilyeva, , Olga M. Koroleva, Ekaterina D. Mikhalcova, Natalia N. Popova, Tatyana V. Gaponova, Valentina N. Dvirnyk, Olga N. Baiteriyakova, Grigory A. Efimov, Alexander S. Vdovin, Valery G. Savchenko and CIC 930
Update on the randomized trial of post-transplanation cyclophosphamide and rabbit ATG for graft-versus-host disease prophylaxis in chronic myeloproliferative neoplasms and myelodysplastic syndrome
Ivan S.Moiseev, Elena V.Morozova, Yulia V.Rudnizkaya, Yu.U Vlasova, Elena I.Darskaya, Olga A.Slesarchuk, Sergey N.Bondarenko, Boris V.Afanasyev 
Prognostic potential of positron emission tomography (PET) prior and after autologous stem cell transplant (ASCT) for chemoresistant Hodgkin lymphoma
Vsevolod G. Potapenko1,2, Natalia B. Mikhaylova1, Irina A. Skorokhod2, Daria A. Chaginskaya2, Victoria V. Ryabchikova2, Eleonora I. Podoltseva2, Victor V. Ipatov3, Igor V. Boykov3, Vyacheslav N. Semelev3, Dmitriy A. Gornostaev3, Nadezhda V. Medvedeva2, Boris V. Afanasyev1
Invasive fungal diseases in adolescents and young adults after allogeneic hematopoietic stem cell transplantation
Oksana Ayzsilnieks1, Marina Popova1, Alisa Volkova1, Olga Pinegina1, Svetlana Ignatyeva2, Kirill Ekushev1, Olga Slesarchuk1, Maria Vladovskaya1, Ludmila Zubarovskaya1, Nikolay Klimko1,2, Boris Afanasyev1
Characteristics of acute leukemia in Chui region of the Kyrgyz Republic
Abdukhalim R. Raimzhanov1, Shakhnoza A. Murzamatova1, Irina A. Tsopova2
Role of polyomavirus in emerging secondary hypofunction of marrow graft following allogeneic bone marrow transplantation in adults
Tatiana A. Rudakova, Yuriy A. Eismont, Ivan S. Moiseev, Ludmila S. Zubarovskaya, Alexander D. Kulagin, Boris V. Afanasyev
Role of allo-HSCT in the treatment of patients with T315I mutation in the TKI era
Yulia Yu Vlasova, Elena V. Morozova, Ivan S. Moiseev, Alexander L. Alyansky A, Tatiana L. Gindina, Boris V. Afanasyev
Influence of mixed chimerism upon outcomes of allogeneic stem cell transplantation (allo-SCT) in patients with non-malignant diseases
Larisa Vakhonina1,2, Igor Vyatkin1,2, Natalya Maysheva1,2, Grigory Tsaur1,2, Tatyana Riger1,2, Anna Demina1,2, Leonid Saveliev1,2,3, Oleg Arakaev1,2, Larisa Fechina1,2
Epidemiology and outcome of lymphomas in HIV-infected patients: a multicenter retrospective study
Marina Popova1, Tatyana Shneyder2, Igor Karaygin2, Olga Ponomarenko2, Ilya Zuyzgin2,11, Olga Ryabykina2, Olga Ruzhinskaya2, Olga Uspenskaya2, Nadezhda Medvedeva3, Anna Klimovich3, Vsevolod Potapenko3, Natalya Kotova3,11, Elena Zinina4, Natalya Popova4, Ylia Zhurba4, Alexander Myasnikov5, Svetlana Moshnina5, Alexey Evseev5, Tatyana Pospelova6, Kamil Kaplanov7, Tatyana Ksenzova8, Elena Karyagina9, Zhanna Stolypina9, Svetlana Dzola10, Alexander Levanov10, Evgenya Borzenkova1, Anastasya Nekrasova1, Natalya Mikhaylova1, Ludmila Zubarovskaya1, Boris Afanasyev1
Predictive value of cytokine levels for acute graft-versus-host disease in hematopoietic stem cell transplantation with post-transplant cyclophosphamide
Olga V. Pirogova1, Ivan S. Moiseev1, V.V. Beklenischev1, Elena A. Surkova2, Sergey V. Lapin2, Elena V. Babenko1, Alexander, L. Alyanskiy1, Boris V. Afanasyev1
Risk-adapted graft-versus-host disease prophylaxis with post-transplantation cyclophosphamide in related, unrelated and haploidentical stem cell transplantations
Olga V. Pirogova, Ivan S. Moiseev, Alexander L. Alyanski, Elena V. Babenko, Elena I. Darskaya, Olga A. Slesarchuk, Sergey N. Bondarenko, Boris V. Afanasyev
Haploidentical Hematopoietic Stem Cell Transplantation (Haplo-SCT) with Post-Transplant Cyclophosphamide for haematological malignancies: a single centre experience
Dmitry V.Motorin, Renat S.Badaev, Diana V.Babenetskaya, Nataliya A.Ilyina, Tatyana O.Silina, Georgiy G.Baratashvili, Vladimir V.Ivanov, Yuliya A.Alexeeva, Andrey Yu. Zaritskey
Efficiency of 5-azacytidine administration before allogeneic hematopoietic stem cell transplantation in acute myelobastic leukemia, myelodysplastic syndrome, and juvenile myelomonocytic leukemia
Varvara N.Ovechkina, Sergej N.Bondarenko, Elena V.Morozova, Olga Slesarchuk, Kirill A.Ekushev, Ludmila S. Zubarovskaya, Boris V.Afanasyev
Obituary. In memory of Professor Thomas Büchner
Professor Sergey F. Bagnenko, Full Member, Russian Academy of Sciences,
Rector, First State Pavlov Medical University of St. Petersburg
Professor Boris V. Afanasyev,
Director, Raisa Gorbacheva Memorial Research Institute for Pediatric Oncology, Hematology and Transplantation, First State Pavlov Medical University of St. Petersburg, Russia
Prof. Dr. Dr. h.c. Rüdiger Hehlmann,
Med. Fakultät Mannheim, Universität Heidelberg, Deutschland

Editorial

10th R.Gorbacheva Memorial Symposium “Hematopoietic Stem Cell Transplantation. Gene and Cellular Therapy”: an overview

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Boris V.Afanasyev

Professor Boris V. Afanasyev, R.Gorbacheva Memorial Research Institute of Children Oncology, Hematology and Transplantation, First State I. Pavlov Medical University, L.Tolstoy St 6-8, 197022, St.Petersburg, Russia. 

Phone: +7(812) 338 6265
E-mail: bvafan@gmail.com

Short reports

Comparative analysis of different CVC dressing types convenience and economic effectiveness in allogeneic hemopoietic stem cell transplant recipients

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Olga Ivanova, Julia Simakina, Marina Ermolova, Julia Oparina, Ekaterina Goncharova, Ludmila Zubarovskaya, Boris Afanasyev

First Pavlov State Medical University of St. Petersburg, St.Petersburg, Russian Federation

Contact: Olga Ivanova
E-mail: ivolga36936@yandex.ru

Introduction

The care for critically ill patients in our clinic requires in most cases the central venous access for therapeutic interventions and diagnostics. The use of central venous catheters (CVCs) has its drawbacks, one of which is development of catheter-associated infections (CAI) found in 20- 55% of patients with sepsis. CAI development leads to significant raise of treatment costs. The prevalence of CAI depends on intraluminal and exit site infections prophylaxis by using appropriate CVC maintenance equipment and techniques.

Materials and methods

A total of 112 of allogeneic hemopoietic stem cell transplantation recipients with AML, ALL, CML and AA were divided into 3 clinical groups based on dressing type used. All patients had CV accesss via v. jugularis interna dextra. In the 1st group (n=38) sterile self-adhesive non-woven pads (Cosmopor) were used. In the 2nd group (n=37) transparent Tegaderm Film dressings were used. In the 3rd group (n=37) transparent gel pads Tegaderm CHG were used. Nurses from CVC care team evaluated patients’ complaints and performed catheter site monitoring for possible presence of local inflammatory changes, allergic reactions and CVC displacement. In addition, dressings changing convenience, catheter site cultures results and economic aspects were analyzed.

Results

The catheter exit site bacterial colonization was lower in patients with Tegaderm CHG dressings (3.4%), than in patients with non-woven pads (5.7%) or Tegaderm Film (5.3%) dressings. In most cases the exit site inflammation was caused by Staph.Аureus, Staph. Epidermidis, Kl.Pneumoniae or Enterococcus spp. All 1st group (non-woven pads) patients reported frequent dressing deformation requiring its replacement and skin discomfort due to residual adhesive material. Patients from the 2nd (film pads) group complained of skin contraction in movement (23%) dressing deformation (22%), itching (18%), while 37% of patients had no complaints. In the 3rd group (transparent gel pads) 92% of patients were satisfied with dressing performance, while 2% reported itching and 6% movement restriction. From nurses’ points of view non-woven pads were inconvenient due to the difficulties of exit site monitoring, need of frequent changing of soiled, wet or deformed dressings. Transparent films were more convenient due to better visual control and less dense change schedule (once 2-3 days). Gel pads were most convenient as they required changing only one in 5-7 days and provided good catheter fixation. The daily cost was greater for daily changed non-woven pads (4.35 euro), than for film pads (2.67 euro) or gel pads (1.72 euro).

Conclusions

Transparent gel pads (Tegaderm CHG) is optimal in post-transplant patients due to less frequent change, which leads to less frequent skin traumas and exit sites infections, is more convenient for patients and nurses. In addition, although gel pads are relatively more expensive, their use provide economic advantages to lower daily cost (1.5-2.5 less than other dressing types) and less frequent CAIs.

Short reports

Treatment of refractory intestinal acute GvHD using multipotent mesenchymal stromal cells (MMSC)

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Darya S. Dubnyak, Larisa A. Kuzmina, Mikhail Y. Drokov, Natalia A. Petinati, Nina J. Drize, Vera V. Vasilyeva, Olga M. Koroleva, Ekaterina D. Mikhaltsova, Natalia N. Popova, Elena N.Parovichnikova, Valerii G. Savchenko

National Research Center For Hematology, Moscow, Russian Federation

Contact: Dr. Darya Dubnyak
E-mail: darya-dubnyak@yandex.ru

Introduction

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) in patients with hematological malignancies is frequently accompanied by severe complications. E.g., acute graft versus host disease (aGvHD) is common in recipients of peripheral stem cells or bone marrow, thus being among the main reasons of increased mortality or suboptimal life quality during post-transplant period. The aim of this study was to assess efficacy of multipotent mesenchymal stromal cells (MMSCs) in patients with refractory clinical forms of aGvHD with affection of gastrointestinal tract (GIT).

Patients and methods

One-hundred-eighty allo-HSCT have been performed at the National Research Center of Hematology (Moscow) between November 2012 and June 2016. Fifteen cases of refractory aGvHD with GIT involvement have been diagnosed. We studied efficacy of MMSC application as a second-line therapy for such GvHD cases: fifteen patients with refractory aGvHD and GIT involvement participated in the study, including eight males and 7 females, at a median age of 40 (21 to 57 years old). Eight patients were admitted with acute myeloid leukemia (AML), and five patients were treated for acute lymphoid leukemia (ALL). Patients with myelodysplastic syndrome (n=1), and chronic myeloid leukemia (n=1) were also included. Two patients underwent myeloablative conditioning regimen versus 13 cases with reduced intensity conditioning regimen. In 10 cases, peripheral blood stem cells have been used. Twelve patients were subjected to unrelated allo-HSCT (matched, 8, mismatched, 4). Three patients underwent allo-HSCT from related donors (matched, 2; mismatched, 1). We used different immunossupressive therapy as a preventive treatment of GvHD: MMF+CSA+MTX (n=9), CSA+MMF (n=3), MMF+post-translant cyclophosphamide on +3,4 day (n=1), post-translant cyclophosphamide on +3,+4 (n=1). In case of unrelated donor, ATG was added to the therapy. GvHD was established on an average of day +54 after HSCT. Six patients were diagnosed with aGVHD of gastrointestinal tract, 3 patients, with aGVHD affecting GIT and skin; two patients, with aGVHD of GIT and liver; four patients had aGVHD with GIT, liver and skin involvement. The most patients suffered from severe form of GvHD (grade 2 was confirmed in 1 case, grade 3, in 8 cases, grade 4, in 6patients).

Results

We administered prednisolone at the dose of 2mg/kg, as upfront therapy,. In 2 cases only partial response was achieved. We did not achieve any response to prednisolone in 12 patients. One patient with refractory form GvHD died due to malignancy progression. Fourteen patients who failed to respond for the first-line therapy were subjected to the second-line treatment. The therapy was started at an average of 14 days (8 to 58) since the diagnosis of aGvHD. In thirteen patients, MMSC were applied at 1…3 doses (1 dose =1x106/ kg MSCs) weekly. In one patient, ATG was administrated as a second-line therapy, however, without any response. In 2 of 13 patients, a “full” response to MMSC therapy was achieved. Moreover, we have got a partial response in 5 patients and did not observe any response in 6 patients. Four patients died from further GvHD progression accompanied by infectious complications. One patient further developed a chronic form of GvHD. In 7 cases, we tried a third-line therapy of aGvHD, with average onset time of 29 (17 to 97 days) since the aGvHD diagnosis. Different therapies were use in these cases. E.g., MMSC were applied in one patient with a partial response to therapy; ATG was injected in 2 patients without clinical effect. Usage of methotrexate in three patients brought a partial effect only in one case case and was entirely ineffective in two cases. Infliximab injection in 1 patient was also without effect. Three patients from the “third-line” group further died from progressing GvHD. A next-line therapy was tried in four patients who failed to respond to previous treatment. Tacrolimus was administered in 1 case, ATG, in 1 patient, rituximab, in 2 cases. These patients died from further GvHD progression and concomitant infectious complications. The three-year overall survival was 17.7%. Conclusions: At present, there is no precise concept for treatment of refractory aGvHD. Our study has shown that the second-line therapy with MMSC may be effective in 57% of the patients with severe aGvHD complicated by GIT affection . Moreover a “full” response was achieved in 14% of the cases. We didn’t observe any differences of efficacy in terms of MMSC source, i.e., if the same or another donor should be recruited. Third-line and subsequent therapies were proven to be entirely ineffective.

Short reports

Alterations in multipotent mesenchymal stromal cells properties: in vitro model of their interactions with allogeneic lymphocytes

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Nikolay M. Kapranov, Julia O. Davydova, Nataliya A. Petinati, Maria V. Bakshinskayte*, Irina V. Galtseva, Nina I. Drize, Larisa A. Kuzmina, Elena N. Parovichnikova, Valeriy G. Savchenko

National Research Center for Hematology, Russian Ministry of Health, *Biological Faculty, Department of Immunology, M.V. Lomonosov Moscow State University, Moscow, Russian Federation

Contact: Dr.Nikolay M. Kapranov
E-mail: kapranov.n@blood.ru

Background

Multipotent mesenchymal stromal cells (MSCs) possess immunomodulatory properties and were successfully used for treatment of autoimmune diseases and acute or chronic graft-versus-host disease (GVHD) after allogeneic hematopoietic cells transplantation. Therapy with MSCs is not always effective. It has been shown that immunomodulatory properties of the MSCs may be improved by means of various agents, such as IFN-γ, TNF-α, IL-17. Characteristics of MSCs after injection into the patients differ from those which have been studied in culture, due to their interactions with other cells in bloodstream and tissues. In this study, an attempt was made to develop a model of IFN-γ-treated MSCs (IFN-γ-MSC) and their in vitro interactions with allogeneic lymphocytes. The aim of the study was to identify the changes of MSCs and IFN-γ-MSC characteristics after co-cultivation with lymphocytes in vitro and to determine the level of expression of CD90 (Thy-1) – MSCs marker characterizing the degree of differentiation, in dynamics.

Materials and methods

MSCs were isolated from donor bone marrow (n=13) harvested at the Department of Bone Marrow Transplantation (NRCH) from healthy donors under their informed consent, and cultured by a standard method in aMEM with 10% fetal bovine serum (FBS). After 2-3 passages, the MSCs were seeded at 105 cells per flask with 25 cm2 bottom area, and 500 units/mL of IFN-γ were added a day later for 4 hours to half of the cultures. Then the media were changed either without lymphocytes or with 106 allogeneic lymphocytes in RPMI-1640 medium with 10% FBS. 5 mg/ml phytohemagglutinin (PHA) was added to the half of flasks with lymphocytes to activate them. All the flasks were cultured for up to 4 days at 37° C and 5% CO2 . After 1, 2, 3 and 4 days in vitro, the lymphocytes were washed from MSCs. MSCs were detached from the flasks with trypsin, and cell viability was determined by Trypan blue dye exclusion test. For each MSCs sample, the mean intensity of CD90-specific fluorescent signal (MFI) was determined by direct immunofluorescent staining with anti-CD90 PE antibody (BD Pharmingen) as with a BD FACSCanto II flow cytometer (BD Biosciences, USA). The data obtained are presented as a mean value ± standard error. Statistical analysis was performed using the Mann-Whitney test for distributions other than the normal and Student’s t-test (considered reliable at p < 0.05).

Results

The number of cells in cultures of MSCs and IFN-γ-MSCs did not differ significantly over 4 days of observation. The presence of non-activated lymphocytes had no effect on the parameters of growth and viability of MSCs. Co-cultivation of activated lymphocytes with MSCs resulted in reduction of viable MSCs to 51.6±5.6% as compared with IFN-γ-MSCs (68.1±6.2%, p=0.02). Viability of cultured MSCs without lymphocytes was 74.5±4.8% by day 4 from the beginning of experiment. CD90 expression on MSCs and IFN-γ-MSCs did not change significantly during all 4 days of the experiment (see Figure). When MSCs and IFN-γ-MSCs were co-cultured with non-activated lymphocytes, the CD90-related MFI was gradually reduced within 4 days when compared to control cultures (1.1; 1.3 1.6, and 1.6-fold, respectively, p=0.03 for control MSCs by day 4), and 0.98; 1.1; 1.5; 1.7-fold (p=0.0001 for IFN-γ-MSCs). When MSCs were co-cultured with activated lymphocytes, the CD90-specific MFI were decreased faster (1.3; 1.2; 1.5 and 2.3-fold, p=0.00001 for MSCs), and 1.1; 1.1; 1.6, and 2.1-fold, p=0.002 for IFN-γ-MSCs), thus suggesting possible induction of MSCs differentiation after contact with lymphocytes. 

Time-dependent alterations in MFI of CD90 on MSC in different culture conditions in dynamics

Conclusion 

The data obtained indicate that MSCs treatment with IFN-γ did not lead to a change in their growth characteristics. However, IFN-γ-MSCs have a greater resistance to activated lymphocytes, which makes them more effective for cell therapy

Short reports

Nursing care for adolescents with graft versus host disease after allogeneic hematopoietic stem cell transplantation

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Anastasia A. Kraeva, Irina A. Badanova, Lubov A. Shepeleva, Maria U. Averyanova, Svetlana I. Gortinskaya-Oleschko, Ludmila S. Zubarovskaya, Boris V. Afanasyev

Raisa Gorbacheva Memorial Research Institute for Children Oncology, Hematology and Transplantology, The First St. Petersburg I.Pavlov State Medical University, Russia

Contact: Anastasia A.Kraeva
E-mail: irulkina_pochta@mail.ru

Background

Graft versus host disease (GVHD) is a common severe complication after allogeneic hematopoietic stem cell transplantation (allo-SCT). Management of GVHD is based on multidisciplinary approach. In particular, nursing care and psychological support play an important role, especially, in children and adolescent patients. The aim of our study was to define incidence of acute and chronic GVHD in teenagers undergoing allo-SCT and to assess a role of nursing for the treatment process, patient care and psychological support. Patients and Methods The data were collected from 87 recipients of allo-SCT with acute and chronic GVHD. All the patients suffered from oncohematological disorders. Age of patients varied from 14 to 20 years.

Results

Frequency of acute GVHD was 59% (n=51), and incidence of chronic GVHD was 14% (n=12). Both types of GVHD, acute and chronic, were observed in 27% (n=24). Involvement of skin and mucous membranes was observed in 49% (n=43), affection of gastrointestinal tract was revealed in 31% (n=27), liver was involved in 19.5% (n=17). Incidence of GVHD after unrelated HLA-matched allo-SCT was 67% (n=58), after haploidentical allo-SCT, the incidence of GVHD was 18% (n=15), and only 15% (n=13) after related HLA-matched allo-SCT. All the patients also filled a questionnaire depicting quality of life (QoL). Up to 65% teenagers with GVHD had psychological problems (depression, misbehavior, abnormal communication with relatives and others). Most of these problems were directly induced by GVHD or by GVHD-associated limitations. QoL was also influenced by long-lasting steroid therapy, prolonged duration of hospitalization and problems in communication with counterparts. We developed recommendations for GVHD patients and their relatives on regime and care. Introduction of these recommendations improved management of patients with GVHD.

Conclusion

Clinical nurses should be aware of GVHD and associated problems in teenagers after allo-SCT. Patients with GVHD require high professional level of nursing (including psychological support for patients and relatives). Development of universal recommendations for patient care after allo-SCT will significantly improve the quality of treatment.

Short reports

Relapse-free survival in multiple myeloma patients after autologous transplantation of peripheral blood stem cells depending of the content of CD45+CD19+ B cells in the apheresis product

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Irina V. Kruchkova, Svetlana A. Sizikova, Galina U. Ushakova, Vera V. Sergeevicheva, Darya S. Batorova, Egor V. Batorov, Natalya V. Pronkina, Andrey V. Gilevich, Elena R. Chernykh

Research Institute of Fundamental and Clinical Immunology, Novosibirsk, Russia

Contact: Dr. Irina V. Kruchkova
E-mail: bmt-novosibirsk@mail.ru

Background

Currently 40% of autologous stem cell transplantation (auto-SCT) is performed for multiple myeloma (MM) patients, but so far there is no way the disease prognosis after auto-SCT having sufficiently high sensitivity and specificity. Objective To explore the relative content of CD45 + CD19 + B-lymphocytes in separated product from the MM patients and to investigate a possible relationship between the number of these cells and disease-free survival after autologous SCT.

Patients and Methods

The study included 17 patients with MM, who underwent high-dose chemotherapy with autologous SCT was performed over 2010-2012. The median age was 47 years (37-58 years). Mobilization of hematopoietic stem cells (HSC) was performed using high-dose cyclophosphamide (4 g/m2) followed by granulocyte colony-stimulating factor 5-10 mg/kg/day. Apheresis procedure was performed using a blood cell separator AS TEC 204 (Fresenius) and Spectra LRS 07 (COBE). At the time of auto-SCT, 6 patients were in complete remission, 9, in partial response and 2 had a resistant disease. Conditioning regimen included melphalan 140-200 mg/m2. 5.6±0.6×106 CD34+ HSC/kg (2.6-13.9×106 CCM/kg) were transplanted. CD45 + CD19 + B cell counts of in separation product were assessed by flow cytometry (BD FACSCalibur). The data are presented as Me (min-max) or M ± SE. Statistical analysis was performed using Statistica 6.0 (StatSoft). To assess significance of differences in continuous variables between independent groups of patients, we used a non-parametric U-Mann-Whitney (PU) test. To evaluate diagnostic values, we used ROC-analysis with calculation area under the curve (AUC). Survival analysis was carried out using the Kaplan-Meier method to estimate the reliability of log-ranktest.

Results

Median observation period after auto-SCT was 44.1 months (4-67.2 months). Median survival was 38.3 months. There were no significant differences between the patients in remission (n=8) and relapse (n=9), in age (48 vs. 46.5 years; Pu = 0.74), re-infused HSC number (4.8± 0.5 versus 6.5 ± 1.1×106/kg P=0.23), and disease status at the time of auto-SCT. Higher levels of CD45+CD19+ B cells in apheresis product were founded in MM patients with relapse (5.7 ± 0.9 vs 2.9 ± 0.9% in remission patients; P=0.046). By means of ROC analysis we have found AUC= 0.80 (95% CI: 0.588- 1.012; p = 0.040). The method showed 100% sensitivity and 60% specificity, with a likelihood ratio of 2.5 at the cut-off point of 2.5% for CD45 + CD19 + B cells. We have revealed significant differences (P log-rank=0.028) when analysing disease-free survival of MM patients after autologous SCT by Kaplan-Meier method, depending on the relative content of CD45 + CD19 + B cells in apheresis product.

Conclusion

In patients with MM, relative content of >2.5% CD45 + CD19 + B-cells in product apheresis may be considered an adverse prognostic factor indicating shorter period of disease-free survival (-16.8 months). Relative content of < 2.5% CD45+ CD19 + B-cells in apheresis product may be associated with prolonged disease-free survival period.

Short reports

Usage of online platforms for remote evaluation of the quality of life in patients following allogeneic transplantation of hematopoietic stem cells on the territory of Russian Federation and ex-USSR

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Darya S. Dubnyak, Mikhail Y. Drokov, Larisa A. Kuzmina, Vera V. Vasilyeva, Olga M. Koroleva, Ekaterina D. Mikhalcova, Natalia N. Popova, Dmitriy E. Vibornih, Sergey O. Khruschev, Elena N.Parovichnikova

National Research Center for Hematology, Moscow, Russian Federation

Сontact: Dr.Darya Dubnyak
E-mail: darya-dubnyak@yandex.ru 

Introduction

Transplantation of allogeneic hematopoietic stem cells (allo-HSCT) is now actively developing in Russia, despite a lot of information about clinical outcomes of allo-HSCT. There is little data about quality of patients’ life (QoL) at different stages, especially at late terms (over 5 years) after allo-HSCT. One of the main limiting factors for this research is the unwillingness and inability of most patients to visit the transplant center due to subjective «complete wellbeing» at later terms after allo-HSCT (for many reasons, including location on large distance). The aim of our study was to conduct a remote survey/interview and to identify the main factors influencing QoL in the patients’ after alloHSCT using an electronic version of the WHOQOL-BREF questionnaire.

Patients and methods

The research was performed from January to May 2016 in Bone Marrow Transplant Department at the National Research Center for Hematology. To conduct this study, an electronic interactive version of Russian WHOQOL-BREF questionnaire was developed. This survey was approved by WHO. It includes 26 questions classified in four areas («domains») of human life, i.e., physical health (domain 1), psychological health (domain 2), social relationships (domain 3) and environment (domain 4). Every answer was measured by means of a 1-to-5 point scale. The points were summarized and recalculation was made according to the algorithms approved by WHO. Finally, all the data collected were presented as separate statistics for each domain at a scale of 1 to 100 points. According to literature data, the average value at each scale is 75 ± 2.5 for Western population (Cummins, 2003). The survey covered 63 patients. They underwent allo-HSCT within a period of 2006 to 2016 including cases of AA (4); LPD (2); MDS (4); MM(1); MPD (2); ALL (8); AML (38); CML (4). The group consisted of 37 women (59%) and 26 men (41%), with a median age of 37 years (21 to 63 years old). Allogeneic bone marrow transplantation from related donors was performed in 47.6% of patients (n=30), from matched unrelated donor, in 41% (n = 26), and from unrelated partially-matched donors, in 11% (n=7). Acute GVHD (stage II-IV) was diagnosed in 30% of cases (n=19). Chronic GVHD (cGVHD) was detected in 41% (n = 26).

Results

The survey involved recipients of blood stem cells and bone marrow from 22 cities, 16 regions of Russia, 3 countries of the former USSR. A Cronbach’s α value was 0.894 of the value reliability (internal consistency) of the questionnaire, thus indicating to sufficient reliability (internal consistency) of the test for this group of patients. Primary data analysis involved information over the four evaluation domains for all the patients at different stages after allo-HSCT. Terms posttransplant proved to be the major factor influencing QoL. Design of a multiple regression model has shown the main factors affecting physical sphere in patients after allo-HSCT: age at the time of survey, presence of cGVHD, sex of patients. The determining quotient for these factors was 0.427 (e.c., assessment of physical sphere for the patients after allo-HSCT can be explained by these 3 factors in 42.7%). We determined standardized coefficients (beta) – factors which were used to compare the results of the effect of different independent factors (gender, age at the time of a survey, the time passed after alloHSCT, presence of acute and chronic GVHD in past period) upon dependent variables (the result of a survey by each of the «domains»). While assessing psychological sphere, we got an appropriate coefficient of determination which did not allow us to talk about significant effect of these factors (including aGVHD) on psychological sphere. According to our data, age at the time of the survey is a leading factor for the social sphere. It seems rational since significance of values in social sphere is reducing with age. When assessing environmental factors, we have shown that the time after allo-HSCT is a main factor (R2 = 0.193) which correlates with patient’s gradual adaptation for environment after long illness and reduced frequency/ duration of hospital staying.

Conclusion

According to our research, the time passed after allo-HSCT is one of the main factors affecting quality of life of patients after transplantation of allo-HSCT on the territory of the Russian Federation and former USSR. The results of an online QoL survey among patients after allo-HSCT 5 years after transplantation are comparable with average values in the population.

Short reports

Granzyme B expression in T-regulatory cells is a strong predictor of acute graftversus-host disease after day +30 in patients with classic immunosuppression after allo-HSCT

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Mikhail Y. Drokov, Elena N. Parovichnikova, Julia O. Davydova, Larisa A. Kuzmina, Irina V. Galtseva, Nikolay M. Kapranov, Vera A. Vasilyeva, Darya S. Dubnyak, Olga M. Koroleva, Ekaterina D. Mikhalcova, Natalia N. Popova, Valery G. Savchenko

National Research Center for Hematology, Ministry of Health, Moscow, Russian Federation

Сontact: Dr. Mikhail Drokov: Moscow
E-mail: mdrokov@gmail.com 

Introduction

Granzyme B is a serine protease commonly found in cytoplasmic granules of cytotoxic lymphocytes and natural killer cells. It is secreted with the pore-forming protein perforin and mediates apoptosis in target cells. Granzyme B mediated cytolysis is one of the regulatory mechanisms, together with IL-2 receptors (CD25) by which T-regulatory cells (T-regs) influence T-effectors. Despite the well-known fact that T-regs participate in pathogenesis of aGVHD, and their levels after allo-HSCT inversely correlate with a probability of aGVHD, the data about functional status of T-regs and aGVHD is still limited.

Patients and methods

Peripheral blood samples were collected in EDTA-tubes at day +30 after allo-HSCT. We use PBMC from 29 patients with hematological malignancies obtained by density gradient media. This method was used due to lymphopenia in this group of patients. Group with no aGVHD consisted of 22 patients (AML=12, ALL n=5, LPD=1, CML=2, AA=1, MDS=1) after allo-HSCT (17, MUD; 5, mismatched unrelated donor). MAC conditioning was used in 17 cases, whereas 5 patients received RIC. The group of aGVHD after day +30 included 7 patients. (AML=4, ALL n=3) after allo-HSCT from MUD (n=5), mismatch unrelated donor (n=1) and 1 recipient of HLA-identical sibling donor. Two patients received a myeloablative conditioning (MAC), and 5 patients received reduced-intensity conditioning regimens (RIC). All the patients in both groups received standard immunosuppression (MMF+CSA+ATG). All the patients developed II-IV grade aGVHD (grade II, 1; grade III, 4; grade IV, 2 patients), with a median onset on day +50 (34-150). Anti-CD4-APC-Cy7, anti-CD25-APC, anti-CD127-FITC and anti-Granzyme B-PE (Becton Dickinson, USA) antibodies were used to determine T-regulatory cells population. A Bland–Altman plot (difference plot) was used to test the agreement between the two different T-reg assays (CD4+CD25high, CD4+CD25highCD127low), and no significant differences were found (p=0.942). Due to this fact, CD4+CD25high cells were identified in our experiments as T-reg cells (Gregg et al., 2005). 30,000 CD4+ cells were analyzed on a BD FACSCanto II to achieve sufficient statistical power (Becton Dickinson, USA).

Granzyme B expression

Results

As seen from Fig. 1, the Granzyme B levels were higher in patients who never developed aGVHD. Fig.2 shows that a mean percentage of Granzyme B positive T-regs in GVHD-free patients was 7.26±1.89%, thus being significantly higher than in group who developed aGVHD after day +30 (2.04±0.93%; p=0.02). Noteworthy, according to our previous data, bacterial or viral infections (e.g CMV) and HLA-mismatches does not affect the Granzyme B expression levels in T-regs. Using ROC curve analysis (Fig.3), we obtained the area under curve (AUC) value of 0.74.


Short reports

Posttransplant ruxolitinib combined with cyclophosphamide for graft versus host disease prophylaxis and relapse prevention in patients with myelofibrosis

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Maria V. Barabanshikova, Ivan S. Moiseev, Elena V. Morozova, Julia J. Vlasova, Vadim V. Baykov, Ildar M. Barkhatov, Boris V. Afanasyev

Raisa Gorbacheva Memorial Institute of Children’s Oncology, Hematology and Transplantation First I. Pavlov State Medical University of St. Petersburg

Contact: Dr. Maria V. Barabanshikova
E-mail: mashaprian@mail.ru

Introduction

Myelofibrosis (MF) is a BCR-ABL–negative myeloproliferative disorder with progressive clinical course and usually poor prognosis. Allogeneic stem cell transplantation (alloHSCT) is currently the only treatment modality with curative potential in patients with MF. JAK1/JAK2 inhibitor ruxolitinib is effective in decreasing symptomatic splenomegaly and myelofibrosis-related symptoms. At the same time, it shows a significant immunomodulatory effect and is widely used for the treatment of acute and chronic GVHD. In our study, we administered ruxolitinib with posttransplant cyclophosphamide for GVHD prophylaxis and relapse prevention.

Patients and methods

We analyzed the results of alloHSCT in 4 patients aged 36-52 (median 41) years. One patient was diagnosed with post-essential thrombocythemia myelofibrosis, and three cases of primary myelofibrosis. By DIPSSplus, 2 patients had intermediate-2 and 2 patients had a high-risk disease. 3 patients were JAK2V617F positive, 1 patient was MPL-positive before alloHSCT. All the patients were treated with JAK1/2 inhibitors before alloHSCT, with median treatment duration of 6 months (3 to 18). Disease stabilization occurred in two cases, and two other patients achieved clinical improvement. In one patient, splenectomy was performed, due to poor spleen response. A reduced-intensity conditioning (Fludarabine 180 mg/m2 plus Busulfan 8-10 mg/kg) followed by alloHSCT from full-matched (3) and mismatched (HLA 9/10) (1) unrelated donor was performed. Graft-versus-host disease prophylaxis consisted of posttransplant cyclophosphamide 100 mg/kg at day +3, +4 and ruxolitinib (5 to 7,5 mg bid from day+5 until day +50 (2), and day +100 (2). G-CSF mobilized peripheral blood progenitor cells were used as a stem cell source. Median number of CD34+cells/kg was 6.7×106 (1.4 to 7.3). The trial is registered on clinicaltrials. gov, NCT02806375.

Results

Primary engraftment was documented in 4 patients. Median time to the leukocyte engraftment was 34 (19-79) days, to platelets’ engraftment, 57 days (20-112). 2 patients developed cytomegalovirus reactivation and were successfully treated with Gancyclovir. 2 patients experienced polyomavirus infection. We did not document any severe episodes of toxicity during ruxolitinib therapy in early posttransplant period. Four patients achieved hematological remission (splenomegaly reduction and constitutional symptoms resolution), molecular remission and full donor chimerism. a near-complete resolution of bone marrow fibrosis was observed in two patients (from grade 3-2 down to grade 1-0) accompanied by molecular remission and full donor chimerism. Two patients developed acute GVHD grade II and moderate overlap GVHD with skin and liver involvement after ruxolitinib discontinuation. GVHD resolved completely after cyclosporine A administration. None of the patients required steroid therapy. All patients are alive without any signs of disease progression with median follow-up of 5 months.

Conclusion

AlloHSCT is an effective treatment modality for myelofibrosis. Post-transplant JAK1/2 inhibition in combination with post-transplant Cyclophosphamide seems a promising therapeutic option to prevent GVHD and relapse. The trial will continue to recruit patients.

Short reports

Сritical conditions in complicated patients with oncohematological diseases

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Alfia F. Baibulatova, Bulat A. Bakirov

Bashkirian State Medical University, Ufa, Russian Federation

Contact: Dr. Alfia F. Baibulatova
E-mail: baf87@inbox.ru

Introduction

The aim of this study was to assess a direct connection between the development of multiple organ failure syndrome with duration of treatment and outcomes of cytostatic chemotherapy.

Patients and Methods

We performed a retrospective analysis of 138 adult patients with oncohematological disorders treated with different chemotherapy regimens, followed by admittance to intensive care unit (ICU). The data analysis was performed by assessing functional state of patient’s organs and systems using a SOFA scale for evaluation of critical states.

Results

Significant coagulopathies were diagnosed in more than 120 patients (86.9% of total group); acute respiratory failure, in 43 cases (31.2%); impaired consciousness, 9 (6.5%); acute renal failure, 25 (18,1% ); cardiovascular disorders, 58 (42.0%); acute liver failure, 34 (24.6%). It is also noteworthy that the multiple organ dysfunction occurred in 68 (49.3%) patients, and sepsis, in 24 patients (17.4%). In 78 patients (56.6%), a multiple organ failure (MOF) has been developed after chemotherapy. When summarizing clinical outcomes in the ICU-treated critical care patients, 91 subjects (65.9%) were returned to a somatic department in satisfactory state. Meanwhile, 47 patients were deceased (34.1%). Assessment of organ failure in MOF patients and their clinical course was quantified by means of SOFA scale. Among 47 deceased patients (34.1%) who died with complications refractory to critical care therapy, the SOFA values were estimated as 19.5 ± 0.5 points. In 91 patients, the degree of organ failure was 3 to 5 points, and 20 points, for 47 patients. Disorders of cardiovascular system and blood clotting dominated among those cases.

Conclusion

The data obtained can be used for prediction of lethal outcomes in oncohematological patients brought to the ICU due to evolving multiple organ failure.

Short reports

Regulatory T cells affect early relapse following high-dose chemotherapy with autologous hematopoietic stem cell transplantation in multiple myeloma patients

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Egor V. Batorov, Marina A. Tikhonova, Irina V. Kryuchkova, Vera V. Sergeevicheva, Svetlana A. Sizikova, Galina Ju. Ushakova, Dariya S. Batorova, Andrey V. Gilevich, Elena R. Chernykh

Research Institute of Fundamental and Clinical Immunology, Novosibirsk, Russia

Contact: Dr.Egor V. Batorov, Research Institute of Fundamental and Clinical Immunology, Novosibirsk, Russia
E-mail: Ebatorov@gmail.com

Introduction

Regulatory Т cells (Тregs) play an important role in pathogenesis of multiple myeloma (MM), providing tumor cell avoidance of immune surveillance. The latter, in turn, are able to induce the Treg expansion. However the data on their contents at different stages of disease and their relations to efficiency of the therapy performed are incomplete and controversial. The aim of this study was to investigate dynamics of CD4+FOXP3+ regulatory T cells (Treg) recovery following high-dose chemotherapy (HDCT) with autologous hematopoietic stem cell transplantation (AHSCT), and possible relationship between Tregs and clinical outcomes in multiple myeloma (MM) patients.

Patients and methods

Fifty-nine patients with MM (median age 49 years, range 32-64) underwent AHSCT in 2009-2015. For stem cell mobilization, patients were administered high-dose cyclophosphamide (4 g/m2) or another standard chemotherapy course + G-CSF, melfalan 140-200 mg/m was used as conditioning regimen. The average CD34+ hematopoietic stem cell dose was 5.9 ± 2.7 × 106 /kg (here and below data as М ± SD). The median observation time was 21 month. The counts of circulating CD4+FOXP3+ Tregs have been evaluated using flow cytometry (BD FACSCalibur, CellQuest Software) before HDCT with AHSCT at the day of engraftment, and following 6 and 12 months. The Mann-Whitney U test was used to calculate differences between groups of patients. ROC-analysis with area under the curve calculation was used to evaluate the predictive value. P < 0.05 was considered statistically significant confidence level.

Results

Pre-transplant count of CD4+FOXP3+ Tregs was 4.0±3.0 %. Percentage of Tregs restored rapidly, became higher than initial pre-transplant level (5.5 ± 3.3 %; pU=0.028) at the time of engraftment (~the 14th day following AHSCT) and then subsequently decreased for a year until it lowered to 2.3 ± 1.3 %. CD4+FOXP3+ Tregs at the time of engraftment were increased in patients with relapse of disease during 12 months following AHSCT (n=7) compared to non-relapsed patients (n=29): 7.9 ± 2.6 vs 4.6 ± 2.5 %; pU=0.0066. The prognostic value of CD4+FOXP3+ Treg relative counts at the day of engraftment was assessed for relapse rates at the 1st year post-transplant. Area under the curve (AUC) was 0.835 (95 % CI: 0.678-0.992; р=0.0066). CD4+FOXP3+ Treg relative count > 5.77 % allowed to predict early relapse (85.7 % sensitivity, 75.7 % specificity, with 3.55 likelihood ratio).

Conclusions

Relative counts of Tregs recovered rapidly following HDCT with AHSCT (at the day of engraftment), became higher than pre-transplant level and then subsequently decreased within a year to healthy donor values. The excess of Tregs at the time of engraftment is associated with early relapse.

Short reports

Successful usage of fludarabine for treatment of partial immunologic hematopoietic graft rejection after allogeneic hematopoietic stem cell transplantation in pediatric patients

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Alexandra E. Burya, Kirill I. Kirgizov, Veronica V. Konstantinova, Ekaterina A. Prinstanskova, Natalia V. Sidorova, Bazarma B. Purbueva, Olesya S. Fink, Andrey A. Bologov, Elena V. Skorobatova

BMT Department, Russian Children’s Research Hospital, Moscow, Russia

Contact: Dr.Alexandra E. Burya
E-mail: burya.a.e@gmail.com

Introduction

There are no any widely accepted recommendations for management of immunologic graft rejection after allogeneic hematopoietic stem cell transplantation (allo-HSCT). A number of possible approaches increase therapeutic risk due to high toxicity and severe side effects. Our aim was to evaluate the Fludarabine-based immunodepletion therapy followed by donor lymphocytes infusion (DLI) for treatment of early immunological hematopoietic graft rejection after allo-HSCT.

Patients and methods

Efficacy of Fludarabine therapy has been tested in BMT Department of the Russian Children Research Hospital. Seven patients at early terms after allo-HSCT were selected (6 patients with non-depleted graft, and one case of haploidentical HSCT with TcR α/β/CD19-depleted graft), treated from 2008 to 2016. Diagnoses: 2 patients with acute myeloid leukemia; 2, with aplastic anemia; 1, with MPS type I (Hurler syndrome); 1, with neuromyelitis optica (Devic’s disease); 1, with osteopetrosis. Treatment indications were as follows: decreased donor chimerism (common or/and restricted by CD34+ and CD3+) less than 95% at 1…6 months after HSCT. All the patients were transfusion-dependent at the moment of therapy (four patients were transfusion-dependent permanently after HSCT, and three required transfusions after the chimerism shift). The regimen used consisted of lymphocyte depletion therapy with fludarabine (150 mg/m2 for 5 days in one patient, and 90 mg/m2 for 3 days in other patients) followed by DLI on day 3 after the fludarabine treatment. Average dose of donor lymphocytes was 7.25х105 cells/kg, number of DLIs was 1 to 6 (depending on individual patient’s characteristics). Chimerism control was performed on day +30 after DLI. The average observation period was 3.5 years (10 months to 8 years).

Results

Five patients developed complete donor chimerism 1 month after DLI, and became transfusion independent. There were no any serious complications or side effects of the therapy. All patients who responded to the treatment have maintained complete donor cell engraftment during entire observation period. Two patients rejected their grafts despite treatment and will be transplanted again.

Conclusion

Our study shows that post-HSCT monotherapy with fludarabine followed by DLI is an effective and safe approach to therapy of partial immunological graft rejection, and may resolve a transfusion dependency.

Short reports

Novel HLA-alleles identified in populations of the Northern Caucasus

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Maria A. Loginova1,2, Igor V. Paramonov2, Andrey V. Rylov2

1 The Rosplasma Center of Medical Research & Industry, Kirov, Russia
2 Kirov Research Institute of Hematology and Blood Transfusion, Киров, Россия

Contact: Maria A.Loginova, Head, Stem Cell Donorship Department, Rosplasma, Kirov, Russia

E-mail: loginova-ma@rosplasma.ru

Objective

A study in genetic diversity of HLA alleles in populations of the Northern Caucasus.

Materials and Methods

High-resolution HLA typing of A, B, C, DRB1 and DQB1 loci was carried out by means of Sanger sequencing technique for 2,543 DNA samples from potential donors of hematopoietic stem cells, recruited in the Republics of Dagestan and Chechnya. To confirm and describe the new alleles, we used new pyrosequencing technologies (Roshe), next-generation sequencing (NGS, GenDx), and monoallelic sequencing (PROTRANS).

Results and Discussion

In the course of this study, we identified 15 new HLA alleles. Frequency of the new alleles in population is approximately 1: 170. The North Caucasian Republics are characterized by maximal frequency of new alleles among all the studied Russian populations, since these ethnic communities are underrepresented in bone marrow donor registries. This is related to the genetic profile of these populations. It should be noted that the 12 newly identified alleles are characterized by nucleotide substitutions which result into amino acid substitutions in domains constituting the peptide binding groove of HLA molecules, thus being involved into the “friend-or-foe” recognition. According to recent data, as by July 31, 2016, the nine novel alleles were registered by the WHO Nomenclature Committee for Factors of the HLA System: 3 HLA-loci A, 2 HLA-loci B, 1 HLA-loci C, 2 HLA-loci DRB1, 1 HLA loci DQB1. Interestingly, some of these alleles were identified in several non-related persons, i.e., A*26:106, B*57:78, A*24:314 were identified in two, five and seventeen cases correspondingly. This result suggests a turnover of these alleles in Russian population rather, than random genetic mutations. At present time, our laboratory is unable to identify specific allele differences for other 6 alleles as compared with known alleles.

Conclusion

The studies performed have shown relevance of further research in HLA-allele diversity among different Russian populations. A regional prevalence of HLA traits among potential HSC donors would increase efficiency of their matching to the patients at Russian clinics performing allogeneic stem cell transplantation.

Short reports

Functional disparity of graft-derived T lymphocytes: experimental data

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Mikhail Y. Drokov, Elena N. Parovichnikova, Darya S. Dubnyak, Larisa A. Kuzmina, Irina V. Galtseva, Julia O. Davydova, Nikolay M. Kapranov, Vera A. Vasilyeva, , Olga M. Koroleva, Ekaterina D. Mikhalcova, Natalia N. Popova, Tatyana V. Gaponova, Valentina N. Dvirnyk, Olga N. Baiteriyakova, Grigory A. Efimov, Alexander S. Vdovin, Valery G. Savchenko and CIC 930

National Research Center for Hematology, Ministry of Health, Moscow, Russian Federation 

Сontact: Dr.Mikhail Drokov
E-mail: mdrokov@gmail.com

Introduction

It is well known that bone marrow (BM) and peripheral blood stem cells (PBSC) have their advantages and disadvantages as the stem cell source. Median time of neutrophil recovery and hospitalization is shorter for PBSC than for BM. On the other hand, rates of acute graft-versus-host disease (aGVHD) are lower in case of using BM as a stem cell source. Almost all authors explain it by different amount of T cells (CD3+ cells) in grafts, but less data is available about functional ability of these T cells. It may be assessed by the amounts of secreted cytokines. Superantigens (SAgs) cause a nonspecific activation of T cells by binding the T cell receptor (TCR), independently of certain Vβ domains of TCR and crosslinking it to major histocompatibility complex (MHC) molecules of antigen presenting cell (APC). Due to this stimulation, CD4+ and CD8+ cells start to secrete effector cytokines and induce activation markers on their cell surface within few hours. Here we present preliminary data on functional assay (response to SAgs stimulation) of graft T cells obtained from healthy donors.

Patients and methods

Graft samples were collected from nine healthy donors (3 bone marrow samples and 6 PBSC specimens obtained after donor G-CSF stimulation (10μg/kg for 5 days). Aliquotes of 106 cells in RPMI-1640 supplied with 10% of autologous serum were incubated with SAgs at a recommended concentration. For T lymphocyte stimulation, we used Cytostim reagent (Miltenyi Biotec, Germany). Brefeldin A was added at specified amounts 2 hours after stimulation, in order to block anterograde transport from endoplasmic reticulum to Golgi structures. The cells were then incubated for 4 hours at 37ºC, 5% CO2. Anti-CD45-APC (BD, USA), anti-CD4-APC-Cy7 (BD, USA), anti-FoxP3-PerCP-Cy5.5 (BD, USA), anti-IL- 17A-PE (BD, USA), anti-CD294-FITC (Biolegend, USA), anti-INF-γ-PE-Cy7 and intracellular staining kit (Cytofix/Cytoperm, BD, USA) were used to determine T helper (Th) subpopulations (Fig. 1) (Th1:CD45+CD4+INF-γ+; Th2:CD45+CD4+CD294+; Treg:CD45+CD4+FoxP3+; Th17:CD45+CD4+IL-17A+). 30,000 of CD4+ cells were analyzed on a BD FACSCanto II (Becton Dickinson, USA) to achieve sufficient statistical power.

Results

As shown in Fig. 2 (left panel), all the bone marrow samples showed similar Th1:Th2:Treg:Th17 ratios, and did not functionally differ from normal peripheral blood (PB) samples. Almost all lymphocytes from bone marrow are presumed to originate from peripheral blood. Meanwhile, all the PBSC grafts had different Th1:Th2:Treg:Th17 ratios (Fig. 2, right panel). Due to wide use of CD34+ cell counts as a “universal index”, and variability in total nucleated cells contents, we attempted to calculate the number of Th1, Th2, Treg, Th17 cells per 100 of CD34+ cells of grafts received by the patients. As shown in Fig.3 (right), appropriate Th1-, Th2-, Treg-, Th17- to-CD34-ratios in PBSC proved to be quite variable. E.g., the relative numbers of Th cell subpopulation may vary up to 40 times from graft to graft. 

Gating strategy for Th subpopulations in PBSC

T helper subpopulation profile in bone marrow versus PBSC

 T helper subpopulation profile in bone marrow versus PBSC (per 100 CD34+ cells)

Conclusion

In vitro stimulation of graft T lymphocytes with SAgs can reflect some events which are similar to those in vivo observed in graft T cells after infusion. They may be used as a model to study the effects of graft composition upon GVHD development, graft failure and other processes in which the graft T cell may participate. Our preliminary data demonstrate that functional profile of the graft-derived T lymphocytes shows an extreme intersample variability and, probably, may affect different processes occurring in PBSC recipients, including clinical outcomes of allo-HSCT. Disclosures: No relevant conflicts of interest to declare.


Short reports

Update on the randomized trial of post-transplanation cyclophosphamide and rabbit ATG for graft-versus-host disease prophylaxis in chronic myeloproliferative neoplasms and myelodysplastic syndrome

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Ivan S.Moiseev, Elena V.Morozova, Yulia V.Rudnizkaya, Yu.U Vlasova, Elena I.Darskaya, Olga A.Slesarchuk, Sergey N.Bondarenko, Boris V.Afanasyev 

R.M. Gorbacheva Memorial Institute of Children Oncology, Hematology and Transplantation, First State I.Pavlov Medical University, St.Petersburg, Russia

Contact: Dr.Ivan S.Moiseev
E-mail: moisiv@mail.ru

Introduction

Our group have previously demonstrated superiority of graftversus-host disease prophylaxis (GVHD) with post-transplanation cyclophosphamide (PTCy) over horse anti-thymocyte globulin (ATG) in acute leukemia patients undergoing unrelated hematopoietic stem cell transplantation (HSCT)1. Nevertheless, in chronic myeloproliferative neoplasms (CMN) and myelodysplastic syndrome (MDS) there is a concern of increased graft failure after PTCy. Therefore, we have initiated the single center prospective randomized trial comparing PTCy and rabbit ATG in this group of patients.

Patients and methods

The inclusion criteria were as follows: patients with CMN and MDS, unrelated HSCT, only 10/10-HLA matched patients, peripheral blood stem cells as graft source, no severe organ dysfunction. All patients received conditioning with oral busulfan 10 mg/kg and fludarabine 180 mg/m2, and GVHD prophylaxis with tacrolimus adjusted for 5-15 ng/ml concentration and 30 days of 30 mg/kg mycophenolate mofetil. The third GVHD prophylaxis agent was either PTCy 50 mg/kg day+3, +4, or rabbit ATG (Thymoglobulin) 5 mg/kg. The strata for randomization was Seattle pre-transplant assessment of mortality (PAM) index2. The primary endpoint was incidence of graft failure and secondary endpoints were GVHD and survival. The trial was registered on clinicaltrials.gov, NCT02627573. So far, 17 patients were enrolled, 9 with MDS, and 8 with MPN.

Results

Nine patients were randomized in PTCy group and 8 in the ATG group. Median follow-up was 10 months (range 2-19). The incidence of primary and secondary graft failure was 0% vs 42%(3) in the PTCy and ATG groups (p=0.023), but the engraftment was slower in the PTCy group (20 days vs 15 days, p=0.009). Acute GVHD grade II developed in one PTCy patient and none in the ATG group, acute GVHD grade III-IV was not observed. None of patients developed moderate and severe chronic GVHD. Only 1 patient in PTCy group developed mild chronic GVHD (p=0.44). One patient in each of the groups relapsed (p=0.79). Overall survival was 100% vs 38% (p=0.002) in PTCy and ATG groups, respectively.

Conclusion

Despite significantly higher incidence of graft failure and lower survival in ATG group, it might be accidental, given the small group size, extremely high graft failure and lower survival than PAM-predicted in the ATG groups. The continuation of the trial is warranted. Unfortunately, the recruitment is very slow, so it may not be possible to complete the trial in the single center setting. The team is currently seeking for collaborators.

References

1. Moiseev IS, Pirogova OV, Alyanski AL et al. Biol Blood Marrow Transplant. 2016 Jun;22(6):1037-42.

1. Au BK, Gooley TA, Armand P et al. Biol Blood Marrow Transplant. 2015 May;21(5):848-54.

Short reports

Prognostic potential of positron emission tomography (PET) prior and after autologous stem cell transplant (ASCT) for chemoresistant Hodgkin lymphoma

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Vsevolod G. Potapenko1,2, Natalia B. Mikhaylova1, Irina A. Skorokhod2, Daria A. Chaginskaya2, Victoria V. Ryabchikova2, Eleonora I. Podoltseva2, Victor V. Ipatov3, Igor V. Boykov3, Vyacheslav N. Semelev3, Dmitriy A. Gornostaev3, Nadezhda V. Medvedeva2, Boris V. Afanasyev1

1First St.Petersburg I.Pavlov State Medical University, St.Petersburg, Russia
2Haematology Department, City Clinical Hospital №31, St.Petersburg, Russia
3S.M.Kirov Military Medical Academy, St.Petersburg, Russia

Contact: Dr. Vsevolod G.Potapenko, City Clinical Hospital №31, St.Petersburg, Russia
E-mail: potapenko.vsevolod@mail.ru

Aim of the study

To evaluate prognostic significance of pre- and post-transplant PET-scans in patients with relapsed or refractory Hodgkin disease.

Patients and methods

The retrospective data on 84 consecutive patients receiving high dose chemotherapy with АSCT were analyzed. Their median age was 26.6 (10-62) years. Median follow-up was 25 (1-85) months. Conditioning regimens were as follows: ВЕАМ (n=48), BEAM with bendamustine (n=32), CBV (n=4). Pre-transplant PET scanning (PET1) was performed in 83 pts. In 57 cases, repeated PET scans have been carried out (PET2). Survival was estimated using Kaplan–Meier method.

Results

Two-year overall and event-free survival rates were 70.6% and 58.7% respectively. Patients with CT-confirmed progression prior to ASCT had a worse prognosis. Predictive value of PET-status was shown in chemosensitive patients (partial/ complete response by CT-scan). The overall and event-free survival rates in PET1-negative and PET1-positive patients were 82% vs 62% (р=0.056) and 74% vs 44% (р=0.003), respectively. In PET2-negative and PET2-positive patients, overall and event-free survival were 90% vs 65% (р=0.013), and 72% vs 52% (р=0.014), respectively. Event-free and overall survival for PET1+PET2- group didn’t differ significantly from appropriate level in PET1- patients. Prognostic significance of PET2 was superior to predictive value of PET1 results.

Conclusion

CT-chemosensitivity exceeds PET in the prognostic significance. Patients with CT-proven response may benefit from PET scanning. While overall prognosis of PET-positive patients is worse, an additional post-transplant PET may be performed for further risk stratification. The PET1+PET2+ group is at higher risk, with regard of probable relapse.

Short reports

Invasive fungal diseases in adolescents and young adults after allogeneic hematopoietic stem cell transplantation

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Oksana Ayzsilnieks1, Marina Popova1, Alisa Volkova1, Olga Pinegina1, Svetlana Ignatyeva2, Kirill Ekushev1, Olga Slesarchuk1, Maria Vladovskaya1, Ludmila Zubarovskaya1, Nikolay Klimko1,2, Boris Afanasyev1

1R.Gorbacheva Memorial Research Institute of Children Oncology, Hematology and Transplantation, First St.Petersburg State I.Pavlov Medical University, St.Petersburg, Russia;
2Dept. of Clinical Mycology, I. Mechnikov North-Western State Medical University, St.Petersburg, Russia

Contact: Dr. Marina Popova
E-mail: marina.popova.spb@gmail.com

Background

Invasive fungal diseases (IFD) after allogeneic hematopoietic stem cell transplantation (allo-HSCT) are still actual problem despite improving prognosis due to recent introduction of novel antifungals and diagnostic procedures. There exist only limited data on IFD in adolescents and young adults (15 to 25 y.o.) after allo-HSCT. The aim of this study focuses on clinical characteristics, risk factors and outcomes of IFD in adolescents and young adults after allo-HSCT.

Patients and Methods

121 patients after first allo-HSCT were included into a retrospective single-center study from Jan 2013 to Nov 2015 (Table 1). Their age was from 15 to 25 years old, a median of 20 years. Underlying diseases were as follows: high risk acute leukemia, 74.4%; lymphoma, 11.6%; nonmalignant diseases, 7.4%, others, 6.6%. Allo-HSCT from matched unrelated donors was performed in 62.8%, matched related donors, 24.8%, and haploidentical grafting, 12.4%. Reduced-intensity conditioning was used in 75.2% of the cases. EORTC/MSG 2008 criteria for IFD diagnosis were used, only probable and proven IFD were taken into analysis. Active diagnostic strategy, including bronchoscopy, was used before allo-HSCT, with bronchoalveolar lavage sampling in patients with CT scanning of the lung lesions. “Active IFD” was defined as IFD diagnosed just before allo-HSCT. IFD-events means a newly arising IFD, and relapse of IFD in those patients who had it before allo-HSCT.

Results

IFD was diagnosed before allo-HSCT in 19.8% of total case number (24/121): invasive aspergillosis (IA), in 15 patients, hepatosplenic candidiasis (HSC), in 5 cases, and one patient had four IFD (IA+HSC, IA + Mucormycosis, IA + PCP). Complete response to antifungal therapy was in 9 patients (37.5%), partial response, in 6 subjects (25%), and “active IFD”, in 9 patients (37.5%). For a half of cases with active IFD (41.7%), a secondary antifungal prophylaxis was performed with Voriconazole. Patients free of IFD before allo-HSCT (n=85) received primary prophylaxis with Fluconazole (70.2%) or with Voriconazole/Posaconazole (5.8%). Cumulative incidence of IFD events by two years after allo-HSCT was 15.7% (19/121), including 18 cases of new IFD, and one IA relapse. Median time to the onset of IFD event was D+43 (14 to 577). Incidence of IA was 8.4% (10/121), with median time to onset D+27.5 (14 to 577). Invasive candidiasis (IC) was documented for 1.1% of total group, with median time to onset D+226.5 (54 to 399); Pneumocystis pneumonia (PCP), 0.5% (since day+43). The main sites of infection were lungs (86.7%). Risk factors for IFD development after allo-HSCT in the patients without pre-transplant IFD were: age < 18 y.o. (p=0.03); non-malignant underlying disease (p=0.016), and acute GvHD (p=0.036). Presence of IFD before allo-HSCT (p=0.67), grade 4 neutropenia for more than 20 days (p=0.08), active underlying disease at the time of allo-HSCT (p=0.38), reactivation of CMV infection after allo-HSCT (p=0.058), relapse of underlying disease within 100 days after allo-HSCT (p=0.65), and presence of chronic GvHD (p=0.19) were not significantly associated with IFD development after allo-HSCT. Ten patients with IA (41.7%) were treated with Voriconazole as a first line therapy, all the subjects with IC, with echinocandins, and a patient with PCP was treated with Trimethoprim/Sulfamethoxazole. A 12-week overall survival (OS) from the day of IFD diagnosis post-HSCT was 68.4% (Fig.1). The 12-week OS in cases of IA was 62.5%, and the patients with IC and PCP were alive at 12 weeks after IFD diagnosis. Two-year OS rates after allo-HSCT was 70.2%. There was no statistical difference in the two-year OS for the cases with (66.7%) or without IFD (71.1%) detected before allo-HSCT (p=0.91).

Conclusions

Incidence of IFD in adolescents and young adults before allo-HSCT was 19.8 %. Cumulative incidence of IFD at 2 years after allo-HSCT was 15.7%. The main type of IFD was invasive aspergillosis (62.5%). Only one patient with IA before allo-HSCT had a relapse of mycosis (5.3%). Risk factors for IFD after allo-HSCT (p < 0.05) were: age < 18 y.o., non-malignant underlying disease, and acute GvHD. 12-week OS value after the post-transplant IFD diagnosis was 68.4%. Due to effective diagnosis, timely treatment and secondary prophylaxis, the invasive fungal diseases, either before or after allo-HSCT, did not impair the outcome of HSCT in adolescents and young adults.

Short reports

Characteristics of acute leukemia in Chui region of the Kyrgyz Republic

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Abdukhalim R. Raimzhanov1, Shakhnoza A. Murzamatova1, Irina A. Tsopova2

1Kyrgyz Research Center of Hematology;
2Kyrgyz-Russian Slavonic University, Bishkek, Republic of Kyrgyzstan

Contact: Dr. Irina A.Tsopova
E-mail: tsopovaira@yandex.ru

Introduction

It is known about 35 new cases of acute leukemia (AL) being registered per 1 Mio population each year. Incidence of distinct AL types much depends on age and differs for various parts of the world. Purpose of the work was to study demographic parameters, structure and risk factors for AL based on the data of clinical/diagnostic Department of the Kyrgyz Research Center of Hematology (KRCH) considering the population inhabiting Chui Region of the Kyrgyz Republic.

Materials and methods

Complete cross-sectional analysis of data from medical cards of AL patients at KRCH was performed over four years.

Results and discussions

Period of examination: January 2013 – January 2016, the number of people registered with diagnosis of AL was 225 persons. Average age of the patients was 27±9,5 years. Of 225 cases, 136 patients (61%) were 17 to 40 years old (group 1); 62 patients (27%) were 41 to 60 years old (group 2), and 27 patients (12%) (group 3) were older than 60 years. Most of AL patients were male (73%), who significantly prevailed over female patients (р < 0.001), at a gender index of 1.1:1. Average age of male and female patients was, respectively, 33.7±11.1 and 38.2±12.3 years (р < 0,001). The ratio of acute myeloid leukemia (AML) to acute lymphoid leukemia (ALL) was 8:1 in group 3 of patients. Appropriate ratios in groups 1 and 2 were 5:4 and 5:2. As based on the card catalogue, twelve patients refused to receive medical treatment. 92% of the patients received one chemotherapy course, where 8% of patients died during or just after the therapy performed. 88% of these patients received second and third polychemotherapy courses, whereas 11 patients should receive therapy, but refused further treatment. Three patients are out of our access now. When analyzing distribution of the disease onset by months, we revealed increased incidence of AL from November till May, when the highest number of patients was noted (40% of cases being registered from December to February, with 20% in November). Such seasonal distribution showed that the number of patients substantially increased over winter months (р < 0.01). Some factors may represent significant risks for AL (р < 0.01), e.g., agricultural labor with potential exposure to UV irradiation, pesticides, some common factors (car emissions, food additives), smoking (60%), harboring the HTLV viruses (16%), hereditary predisposal for cancer (40%), and 4% could be attributed to stress factors.

Conclusion

Data analysis from medical cards of KRCH registry has shown that, among AL patients, who permanently inhabited Chui Region, 56% of patients were at active working age, and 27% of patients were in their advanced age. The majority of patients (р < 0.01) were males and were elder persons (р < 0,001). More common admission of the patients over the autumn-to-spring time correlates with the main (mostly, environmental) risk factors.

Short reports

Role of polyomavirus in emerging secondary hypofunction of marrow graft following allogeneic bone marrow transplantation in adults

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Tatiana A. Rudakova, Yuriy A. Eismont, Ivan S. Moiseev, Ludmila S. Zubarovskaya, Alexander D. Kulagin, Boris V. Afanasyev

R.M. Gorbacheva Memorial Research Institute of Children Oncology, Hematology and Transplantation, The First St. Petersburg State I. Pavlov Medical University, St. Petersburg, Russian Federation

Contact: Dr. Tatiana A. Rudakova
E-mail: t_a_rudakova@mail.ru

Background

Poor function of haematopoietic transplant is considered an important complication following allogeneic bone marrow transplantation (allo-HSCT). According to different reports, incidence of BK polyomavirus (BKV) reactivation following allo-HSCT can be as high as 33-54% (1). Relation between BK viruria (or viremia) and the risk of hemorrhagic cystitis is well studied in HSCT patients (1-3). Cases of BKV-associated encephalitis and pneumonitis have been also reported (2). Some studies suggest correlation between BK virus reactivation and cytopenia in both renal (1) and bone marrow transplant recipients (2-4). Taking into account susceptibility of different cell types to BKV infection, including peripheral leukocytes (4), one may suggest a clinical form of BKV disease manifesting as bone marrow transplant insufficiency.

Objective

The aim of our study was to assess a role of BKV infection in the development of secondary graft hypofunction following allogeneic HSCT.

Materials and methods

We carried out a retrospective analysis of 328 patients with different oncohematological disorders (18 to 77 y.o., a median of 31 years), who underwent allogeneic HSCT (matched related donors, 89; matched unrelated donors, 211; haploidentical donors, 28) during the period from 2014 to 2016. Myeloablative conditioning regimen was applied in 75 patients (23% of total), reduced-intensity regimens were used in 253 cases (77% of total). Characteristics of the group are shown in Table 1.

 Clinical characteristics of the group under study (n=328)

Fig. 1. Distribution of clinical samples tested for polyomavirusFig. 2. Prevalence of BK virus in primary graft failure(PGF) and secondary graft hypofunction (SGH).

Over the posttransplant period 219 patients were tested for BKV in different body fluids including blood plasma,urine, bone marrow aspirates, CSF etc. (Fig.1), using qualitative gene-specific PCR. BKV testing was carried out both in cases of local clinical symptoms (cystitis, encephalitis, graft failure etc.), and as a routine screening. Tests were positive in 166 (78%) cases and negative in 45 patients (2%). Altered graft functioning was evaluated as follows: primary graft failure (PGF) was identified in absence of donor hematopoiesis without clinical relapse; secondary graft hypofunction (SGH) was determined as post-engraftment cytopenia with sustained donor chimerism, or as a persisting dependence on transfusions and CSF injections with full donor chimerism and no signs of tumor relapse.

Results

Poor graft function was observed in 194 patients (59% of total); primary graft failure (PGF) was documented in 21 cases (6.4%); secondary graft hypofunction (SGH) was revealed in 172 patients (52.4%). BKV reactivation was found in 163 patients, including 42 cases with normally functioning graft, primary graft failure was diagnosed in 10 cases, whereas 111 patients exhibited SGH. BKV detection in bone marrow was a risk factor for SGH, as shown by a single-factor analysis. In the patients with BKV reactivation (n=163), PGF and SGH frequency was respectively 6.8% (n=11) and 68% (n=112). Among patients without BKV reactivation (n=56), PGF was reported in 1.7% (n=1), and SGH, in 46.4% (n=26, p < 0.002), as shown in Fig.2. In SGH cases, the median time for BKV detection was 46 days, thus preceding the onset of graft hypofunction (a median of 52 days). BKV reactivation episodes were observed within 14 days before the onset, or during graft hy ofunction in 74 patients (43% of the SGH group, or 22.5% of the total HSCT cohort). Meanwhile, bone marrow samples were positive for BKV in 46 patients, i.e., 27% of all SGH cases, or 14% of total allo-HSCT cohort. However, BKV reactivation (including bone marrow affection) did not correlate with total survival after HSCT.

Conclusion

Post-transplant BKV infection may be a predisposing or triggering factor in the development of hematopoietic graft hypofunction. Further studies are required in order to obtain more detailed information about BKV replication in bone marrow and its potential effects upon post-HSCT survival.

References

1. Erard V, Storer B, Corey L, Nollkamper J, Huang M-L, Limaye A, et al. BK virus infection in hematopoietic stem cell transplant recipients: frequency, risk factors, and association with postengraftment hemorrhagic cystitis. Clin Infect Dis 2004; 39: 186

2. Reploeg MD, Storch GA, Clifford DB. BK virus: a clinical review. Clin Infect Dis 2001; 33:191-202.

3. Pambrun E, Mengelle C, Fillola G, Laharrague P, Esposito L, Cardeau-Desangles I, Del Bello A, Izopet J, Rostaing L, Kamar N. An association between BK virus replication in bone marrow and cytopenia in kidney-transplant recipients. J Transplant 2014; 2014:252914. doi: 10.1155/2014/252914.

4. Dropulic LK, Jones RJ. Polyomavirus BK infection in blood and marrow transplant recipients. Bone Marrow Transplantation 2008; 41:11–18

Short reports

Role of allo-HSCT in the treatment of patients with T315I mutation in the TKI era

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Yulia Yu Vlasova, Elena V. Morozova, Ivan S. Moiseev, Alexander L. Alyansky A, Tatiana L. Gindina, Boris V. Afanasyev

R.M. Gorbacheva Institute of Pediatric Oncology Hematology and Transplantation St. Petersburg State Medical I. Pavlov University.

Contact: Dr.Yulia Yu. Vlasova
E-Mail: ij_vlasova@mail.ru

Introduction

Treatment of CML is based on the use of tyrosine kinase inhibitors (TKIs). Despite high effectiveness of the TKI, some patients in CP and significantly more patients in the AP and BC are resistant to it. Distinct point mutations at the kinase domain of ABL-tyrosine kinase are the most important mechanisms of TKI resistance. At present, T315I mutation is known to cause leukemia cells resistance for all known I and II generation TKIs.

Materials and Methods

We present cumulated results of 18 allogeneic bone marrow transplantations (allo-HSCT), performed in 16 patients with CML harboring T315I mutation, as well as data on pharmacological treatment of 37 patients that have been provided by hematological centers of the Russian Federation.

Results

Four patients were in chronic phase 1 (CP1); 7 other cases, with CP≥2; five patients, in acceleration phase (AP); two patients were in blast crisis (BC) at the time of allo-HSCT. Fourteen patients were male, and two females. Their median age was 34 years (17-55 years old). HSCT was performed from HLA-identical siblings (n=7), or unrelated donors (n=11). Therapy before allo-HSCT consisted of the first-line treatment (n=2), ≥ 2 lines of therapy (n=3), ≥ 3 treatment lines (n=11). Median time from diagnosis to HSCT was 39 months (14 to 139 mo). Median time from detection of the mutation to allo-HSCT was 10 months (2 to 38 mo). EBMT scores were as follows: 3-4 points (n=12), 5-7 points (n= 4). Conditioning regimens included myeloablative therapy (MAC) for 5 cases, and reduced-intensity conditioning (RIC) in 13 patients. Seven patients out of 16 survived during the observation terms. At the time of HSCT, two patients were in CP1; 4, in CP2; one patient was in AP. At present time, all the patients are in complete molecular remission. Three patients in the 1st remission, and four, in remission achieved after prophylactic administration of TKI. So far, median observation time for living patients is 48 months (8-79 mo). Overall survival of the patients with allo-HSCT was 42.9%, with median follow-up of 18 months. 18 patients of 37 survived after receiving pharmacological therapy (male, 25, and female, 12). Their age was 17 to 77 years, with a median of 49 years. At the time of diagnosis, 29 patients were in CP, 8, in AP. When the T315I mutation was detected, 23 patients were in CP; AP was registered in 11 cases, and 3 patients were in BC. Prior to detection of the mutation, 5 patients received 1st line TKI therapy, 22, 2nd line TKIs, and 10 were treated with 3rd line TKI. Appropriate therapy was as follows: ponatinib (n=3), I-II generation TKI (n=8), Chemotherapy+TKI\ hydroxyurea+IFN (n=17). At present time, six patients are in CP≥1, nine patients have developed CHR, ССR, or partial cytogenetic response. Current observation terms in living patients are 53-250 months, a median of 81 months. Overall survival among patients receiving pharmacological therapy is 48.4%, with a median follow-up of 81 months.

Conclusion

Interpretation of the newly found T315I mutation is important in view of clinical phase assessed at the time of this molecular diagnosis. The patients in CP1 may continue therapy with TKI. Allo-HSCT is a potential therapeutic option for CML patients in the AP and BC carrying T315I mutation, especially for patients in CP ≥2.

Short reports

Influence of mixed chimerism upon outcomes of allogeneic stem cell transplantation (allo-SCT) in patients with non-malignant diseases

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Larisa Vakhonina1,2, Igor Vyatkin1,2, Natalya Maysheva1,2, Grigory Tsaur1,2, Tatyana Riger1,2, Anna Demina1,2, Leonid Saveliev1,2,3, Oleg Arakaev1,2, Larisa Fechina1,2

1Regional Children’s Hospital No.1, Ekaterinburg, Russia
2Research Institute of Medical Cell Technologies, Ekaterinburg, Russia
3Ural State Medical University, Ekaterinburg, Russia

Contact: Dr. Larisa Vakhonina
E-mail: vakhonina_larisa@mail.ru

Introduction

AlloSCT is the only curative option for treatment of hematological disorders with suppressed hematopoiesis and primary immune deficiencies. Nonmyeloablative conditioning (MAC) regimens lead to long persistence of mixed chimerism (MC) in majority of the patients. Purpose of our study was to estimate relationships between the type of hematopoietic chimerism and development of GVHD following alloSCT being performed in patients with non-malignant diseases.

Patients and methods

Eleven patients (8 boys and 3 girls) at a median age of 9 years (range 3 to 17) were included in this study, including seven 7 patients with severe aplastic anemia (SAA); two, with Fancony anemia (FA); one case of thalassemia, one with Nijmegen syndrome. The patirnts were treated at the Pediatric Oncology and Hematology Center (Regional Children Hospital No.1) from 2008 to 2016. Stem cell donors were as follows: siblings (7 cases), nonrelated HLA-matched donors (n=4). Bone marrow aspirates and mobilized peripheral hematopoietic stem cells were used in 5 and 6 cases, respectively. Conditioning regimens included fludarabine, cyclophosphamide and horse ATG for SAA patients. For FA and Nijmegen syndrome patients, this schedule was augmented by low-dose busulfan treatment. In thalassemic patient, we used MAC with busulfan, fludarabin and horse ATG. In majority of cases, GVHD prophylaxis consisted of combined tacrolimus and methotrexate. In cases of unrelated alloSCT, the patients were additionally treated with mycophenolate mofetil. A median follow-up period was 30 mo (range 4-90). Quantitative evaluation of donor chimerism was done by multiplex amplification of STR loci with subsequent fragment analysis using «COrDIS Plus» kit («Gordiz», Russia). We studied chimerism for the both marrow and peripheral blood cells, as well as for CD3+ and CD19+ lymphocyte subpopulations. Donor genotype of ≥99% hematopoietic cells was considered a complete donor chimerism (CDC), whereas levels of < 99% were regarded as mixed chimeric state.

Results

All the patients did successfully engraft, and all these patients were alive at the present time. There were no severe life-threatening complications, infections, or graft rejection observed. Only two patients achieved CDC by the day +28, and only these patients retained complete chimerism at the day +100. At this time point, a mixed chimerism was most common in CD3+ lymphocytes. One year after SCT, a proportion of CDC patients increased up to 82% (2 patients are at earlier post-transplant terms). There were no correlations between the engraftment terms and chimerism values at the day +28, like as between the CD34+ cell dose transplanted, and chimerism levels (p>0.05 in both cases). Interestingly, severe GVHD was noted in only two female patients with CDC at day +28. The first case exhibited grade III aGVHD following HSCT from a non-related donor; the second case showed extensive form chronic GVVHD since 1 year after HSCT from a sibling. There were no other cases of severe (grade III-IV) aGVHD in the patients observed. Localized form of chronic GVHD was revealed in 4 cases (36%). In other patients, no signs of chronic GVHD were seen.

Conclusions

Despite limited number of observations, we assumed that fast achievement of CDC may be associated with severe GVHD. And, vice versa, long persistence of mixed chimerism seems to prevent the GVHD emergence. However, our findings should be confirmed in larger group of patients and, preferably, in multicenter settings.

Short reports

Epidemiology and outcome of lymphomas in HIV-infected patients: a multicenter retrospective study

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Marina Popova1, Tatyana Shneyder2, Igor Karaygin2, Olga Ponomarenko2, Ilya Zuyzgin2,11, Olga Ryabykina2, Olga Ruzhinskaya2, Olga Uspenskaya2, Nadezhda Medvedeva3, Anna Klimovich3, Vsevolod Potapenko3, Natalya Kotova3,11, Elena Zinina4, Natalya Popova4, Ylia Zhurba4, Alexander Myasnikov5, Svetlana Moshnina5, Alexey Evseev5, Tatyana Pospelova6, Kamil Kaplanov7, Tatyana Ksenzova8, Elena Karyagina9, Zhanna Stolypina9, Svetlana Dzola10, Alexander Levanov10, Evgenya Borzenkova1, Anastasya Nekrasova1, Natalya Mikhaylova1, Ludmila Zubarovskaya1, Boris Afanasyev1

1Raisa Gorbacheva Institute of Children Oncology, Hematology and Transplantation, First Pavlov State Medical University of St.Petersburg;
2Leningrad Regional Clinical Hospital, St.Petersburg;
3City Clinical Hospital №31, St.Petersburg;
4Surgut District Clinical Hospital, Surgut;
5Republican Clinical Hospital, Petrozavodsk;
6Center of Hematology, Novosibirsk;
7Volgograd Regional Clinical Oncologycal Hospital, Volgograd;
8Regional Clinical Hospital, Tumen’;
9City Clinical Hospital №15, St.Petersburg;
10Clinic of Profpathology and Hematology, Saratov State Medical University, Saratov;
11Petrov Research Institute of Oncology, St.Petersburg


Contact: Dr. Marina O.Popova
E-mail: marina.popova.spb@gmail.com

Background

Since the beginning of global epidemic, almost 70 million of people have been infected with the HIV virus, and about 35 million people have died of AIDS. HIV infected patients are at risk for cancer including lymphomas, despite the widespread accessibility of highly active antiretroviral therapy (HAART). In parallel with increasing number of patients living with HIV, the number of patients suffering from HIV-associated malignancies of hematopoietic and lymphoid tissues has increased as well. The aim of our study was to determine epidemiological characteristics and the outcome of lymphomas in HIV-infected patients in Russian Federation.

Materials and methods

We performed a retrospective multicenter study. The inclusion criterion was a diagnosis of lymphoma in HIV-infected patients. Seventy-three patients were enrolled over a period from May 2006 to Dec 2015. We analyzed data of medical histories, laboratory tests, treatment in hematological hospitals and AIDS Centers, as based on established local guidelines. The median follow-up of the patients was 24 (2-110) months.

Results

Epidemiological and clinical characteristics (diagnosis) of the patients’ are presented in Figure 1. Table 1 presents clinical features of the HIV patients with lymphoproliferative disorders, characteristics of lymphoma status, and treatment data. Overall survival at 2 years in HIV-infected patients with lymphomas was 64% (HL, 80%; NHL, 60%; MM, 100%), time to progression (TTP) was 17% (HL, 20%; NHL, 16.7%; MM, 0%). Favorable prognostic factors for OS were: chemotherapy (ChT) in combination with HAART, adequacy of ChT for the type and stage of lymphoma. Favorable prognostic factors for TTP were: adequate ChT for the type and stage of disorder. Adverse prognostic factors for OS and TTP were: LDH level >500 U/L; B symptoms. Usage of ChT+Rituximab improved overall survival (67.6% vs 42%, p=0.07) and reduces the probability of progression of CD20+ B-cell lymphoma (8% vs 37%, p=0.01). Patient’s age, ECOG status, disease stage, presence of B symptoms and International Prognostic Index (IPI) had no impact on the outcome, which indicates to presence of other factors, which could include HIV Status (CD4+ cell levels, and viral load).

Histological types of lymphoproliferative diseases in HIV-infected patients

 Clinical characteristics of the patients under study

Conclusions

Aggressive DLBCL in HIV-positive patients was diagnosed more often than other lymphoma types. The 2-year OS in patients with HIV and lymphomas was 64%. Adequate ChT, with regard to type and stage of lymphoma in combination with HAART improved overall survival. LDH levels higher than 500 U/L and B-symptoms were adverse prognostic factors. Usage of Rituximab for CD20 B-cell lymphomas reduced the probability of progression. HIV status seems to play an important prognostic role for the further outcomes. Continuous prospective studies are required in the field.

Short reports

Predictive value of cytokine levels for acute graft-versus-host disease in hematopoietic stem cell transplantation with post-transplant cyclophosphamide

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Olga V. Pirogova1, Ivan S. Moiseev1, V.V. Beklenischev1, Elena A. Surkova2, Sergey V. Lapin2, Elena V. Babenko1, Alexander, L. Alyanskiy1, Boris V. Afanasyev1

1R.M. Gorbacheva Memorial Institute of Children Hematology, Oncology and Transplantation;
2Laboratory of Autoimmune Diagnostics, First St.Petersburg State I.P. Pavlov Medical University, St. Petersburg, Russian Federation

Background

Currently, acute graft-versus-host-disease (GVHD) remains one of the most serious complications of allogeneic hematopoietic stem cell transplantation (НSCT). The release of large amounts of cytokines causes activation of alloreactive T-cells in the early period after transplantation is an important step in pathogenesis of acute GVHD. The basic immunological events leading to GVHD occur during several weeks after hematopoietic graft transfusion. By this reason, cytokine profiling could be used to predict development of GVHD. The majority of studies on cytokines in allogeneic HSCT were performed with classical GVHD prophylaxis, consisting of non-specific immunosuppressive agents like calcineurin inhibitors, methotrexate, mycophenolate mofetil, anti-thymocyte globulin, etc. When using this type of prophylaxis, most studies agree in that higher levels of pro-inflammatory cytokines are associated with development of acute GVHD, while lower levels indicate a successful immunosuppression in abrogation of alloreactive response. Recently, many transplant centers are interested in a novel approach for the GVHD prevention based on Cyclophosphamide post-HSCT (PTCy). Currently, there is no clarity, whether cytokine changes after PTCy are similar to the situation with classical GVHD prophylaxis. It is also unclear if well-known predictive biomarkers remain valid after PTCy-based prophylaxis. Trying to answer these questions, we have conducted a pilot study to determine the dynamics of plasma cytokines in patients undergoing HSCT with PTCy for GVHD prophylaxis. For this pilot study, we have selected five cytokines, IL-17, IL-6, IL-8, IFN-γ, and TNF-α, that were shown to have predictive value for acute GVHD. We, therefore, compared their levels in patients who developed acute GVHD versus GVHD-free cases.

Patients and Methods

Among 192 patients transplanted with PTCy prophylaxis at The First State I.P. Pavlov Medical University between 2014 and 2015, we have identified 20 cases with acute GVHD and plasma samples available. These patients were matched to patients who did not develop acute GVHD at a 1:2 ratio. By decreasing priority, the matching criteria were as follows: donor type, graft source, intensity of the conditioning regimen, primary diagnosis, and disease status. Thus, the study group consisted of sixty adult patients with hematological malignancies who underwent HSCT. All patients have signed informed consent for the collection of blood samples and use of their personal data for research purposes. 28% were grafted from related donor, 72% – from unrelated. The patients received either myeloablative conditioning (MAC) with oral Busulfan (16 mg/kg) and Cyclophosphamide (100 mg/kg), or reduced-intensity conditioning (RIC) with oral Busulfan 8 mg/kg and Fludarabine 180 mg/m2 . All the patients received PTCy- based GVHD prophylaxis. For recipients transplanted from 10/10-HLA matched unrelated donors (MUD), GVHD prophylaxis included combination of Cyclophosphamide (50mg/kg) on day +3,+4, Tacrolimus with target concentration of 5-15 ng/ ml from day+5 through day+120, and Mycophenolate Mofetil (MMF) at 30 mg/kg, or 45 mg/kg for 8-9/10 mismatched unrelated donors (MMUD) from day+5 through day+35. For matched related donors (MRD) with bone marrow as graft source, GVHD prophylaxis was a single-agent Cyclophosphamide (50mg/kg) on day +3, +4. For blood testing, 10 mL of EDTA-preserved venous blood were obtained from the patients on days -7, 0, +7, +21 and +28. Plasma samples were obtained by centrifugation at 1000 g, and then stored in aliquotes at -80°C until the day of assays. We studied 5 plasma biomarkers, i.e., IL-17A, IL-6, IL-8, TNF-α and IFN-γ. Appropriate plasma levels were measured with ELISA technique using commercially available kits (Сytokine, St.Petersburg, Russia). All the assays were performed in compliance with protocols provided by the kit manufacturer. The cytokine concentrations were measured without knowledge of individual clinical data.

Results

Of sixty patients included into the study, twenty recipients developed aGVHD grade I-IV, whereas forty patients were GVHD-free. Eight of these 20 patients (40%) had grade I; 7 (35%) grade II; 5 (25%) grade III aGVHD. 6 patients (30%) developed a systemic aGVHD. There was no difference in cytokine concentrations between GVHD+ and GVHDgroups. The levels of all the cytokines under study did not show any correlations with a risk for aGVHD grades I-IV. We have not received significant differences between the concentrations of cytokines in the study groups at any time point (p>0.05). Noteworthy, additional comparisons between the groups with systemic GVHD and without GVHD have shown lower levels of several plasma cytokines in patients who did develop GVHD. A reduced mean level of IL-8 (98.74±39.47 vs 276.09±284.78 pg/ml, p=0.008 for GVHD+ and GVHD-, respectively), and IFN-γ (22.69±16.35 vs 60.07±42.00 pg/ml, p=0.005 for GVHD+ and GVHD-, respectively) at day +28 were significant risk factors for aGVHD occurrence. By ROC analysis, cutoff values were determined for IL-8, as (147.09 pg/ml; AUC= 0.833), and for IFN-γ (35.94 pg/ml; AUC= 0.848). Incidence of systemic aGVHD was significantly higher in patients with cytokine levels under the cutoff values (0% vs 35.3%, p=0.002, and 3.8% vs 38.5%, p=0,003, for IL-8 and IFN-γ, respectively).

Conclusions

In summary, our pilot trial has shown that time dynamics of cytokine levels after GVHD prophylaxis with PTCy might be different from those under conventional immunosuppressive treatment. Likewise, the well known predictive biomarkers might not work after PTCy. Further large prospective trials could elucidate the differences and determine reliable predictors for acute GVHD. Moreover, it also may be important to introduce additional time points for blood sampling before PTCy administration.

Short reports

Risk-adapted graft-versus-host disease prophylaxis with post-transplantation cyclophosphamide in related, unrelated and haploidentical stem cell transplantations

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Olga V. Pirogova, Ivan S. Moiseev, Alexander L. Alyanski, Elena V. Babenko, Elena I. Darskaya, Olga A. Slesarchuk, Sergey N. Bondarenko, Boris V. Afanasyev

R.M. Gorbacheva Memorial Institute of Children Hematology, Oncology and Transplantation, First St.Petersburg I.P. Pavlov State Medical University, St.Petersburg, Russian Federation Contact: Dr. Olga V.Pirogova E-mail: Dr.Pirogova@gmail.com

Introduction

There is a growing evidence of safety and efficacy of post-transplantation cyclophosphamide (PTCy) in related and haploidentical bone marrow (BM) transplantations, but the data regarding unrelated and peripheral blood stem cell (PBSC) transplants is limited. Hence, we conducted a prospective trial of risk-adapted graft-versus-host disease (GVHD) prophylaxis with PTCy that included different types of donors and graft sources.

Patients and methods

200 adult patients (median age 32 y.o., range 18-62) with hematologic malignancies, including AML (47.5%), ALL (26.5%), CML (12%), MDS (4%), and lymphomas (10%), were enrolled into NCT02294552 trial. 23% of patients were classified as salvage. 26% received the graft from matched related (MRD), 65% from matched/mismatched unrelated (MUD/MMUD), and 9% from haploidentical (haplo) donor. 43% received BM graft and 57%- PBSC graft. 18.5% had myeloablative conditioning and 81.5% - reduced-intensity conditioning. GVHD prophylaxis for matched BM graft consisted of single-agent PTCy 50 mg/kg days+3,+4; for matched PBSC graft we used PTCy+ tacrolimus+ mycophenolate mofetil (MMF) 30 mg/kg days 5-35, and for any mismatched graft, PTCy+ tacrolimus+ MMF 45 mg/kg at days 5-35. Median follow-up was 17 months (range 3-38).

Results

Grade II-IV, III-IV acute GVHD, and moderate-severe chronic GVHD in MRD, MUD/MMUD and haplo groups were respectively 8% (95% CI 3-21%) vs 18% (95% CI 13-26%) vs 26% (95% CI 11-61%), p=0.16; 4% (95% CI 1-16%) vs 6% (95% CI 3-12%) vs 0%, p=0.57; 15% (95% CI 7-29%) vs 10% (95% CI 5-17%) vs 16% (95% CI 5-59%), p=0.35. Non-relapse mortality (NRM) and relapse incidence were 8% (95% CI 3-22%) vs 14% (95% CI 9-21%) vs 16% (95% CI 6-45%), p=0.50, and 38% (95% CI 3066%) vs 23% (95% CI 16-32%) vs 45% (95% CI 27-76%), p=0.02 for MRD, MUD/MMUD and haplo group, respectively. 2-year overall survival (OS), event-free-survival (EFS), and GVHD-relapse free survival (GFRS) were 70% (95% CI 53-82%) vs 67% (95% CI 57-75%) vs 44% (95% CI 21-65%), p=0.007; 53% (95% CI 38-67%) vs 64% (95% CI 54-72%) vs 36% (95% CI 15-57%), p=0.009; 37% (95% CI 23-51%) vs 59% (95% CI 50-67%) vs 36% (95% CI 15-57%), p=0.035 for MRD, MUD/MMUD and haplo groups, respectively. There were no statistical differences in OS, EFS and GFRS between MUD and MMUD (p>0.05). In a multivariate analysis, only salvage status (HR 3.0 95%CI 1.94.7, p < 0.0001) and occurrence of sepsis (HR 1.7 95%CI 1.0-2.7, p=0.04) were predictive for EFS, while type of donor was not a significant factor (HR 1.1 95%CI 0.7-1.9, p=0.60) (Fig. 1). The incidences of complications were: hemorrhagic cystitis, 23%; sepsis, 24%; severe sepsis, 8%; invasive mycosis, 8%; CMV reactivation 45%; veno-occlusive disease, 2.5%; transplant-associated microangiopathy, 3.5%; grade 3/4 liver toxicity, 14%; grade 3/4 kidney toxicity, 1%. Conclusion With relatively short follow-up, we have demonstrated that the risk-adapted PTCy GVHD prophylaxis is safe and results in very low incidences of NRM, acute and chronic GVHD. Therefore, it could be used with any kind of graft source. It also alleviates the GVHD risk in MMUD and haplo donor settings. Relapse of underlying malignancy still significantly influences the outcome with this mode of prophylaxis.

Short reports

Haploidentical Hematopoietic Stem Cell Transplantation (Haplo-SCT) with Post-Transplant Cyclophosphamide for haematological malignancies: a single centre experience

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Dmitry V.Motorin, Renat S.Badaev, Diana V.Babenetskaya, Nataliya A.Ilyina, Tatyana O.Silina, Georgiy G.Baratashvili, Vladimir V.Ivanov, Yuliya A.Alexeeva, Andrey Yu. Zaritskey

Federal Almazov North-West Medical Research Centre, Saint Petersburg, Russia.

Contact: Dr.Dmitry V.Motorin
E-mail: almazov-bmt@mail.ru

Introduction

For many patients with hematological malignancies, hematopoietic stem cells transplantation is the only potentially curative treatment option, but only 30% of patients have fully HLA-matched donor. In case of urgent need to transplant, use of alternative haploidentical donor can be the only chance for cure. For a long time, usage of haploidentical SCT was limited due to high rate of severe GVHD and graft failure. Introduction of GVHD prophylaxis with рost-transplant Cyclophosphamide (PT-Cy) allowed overcome this barrier [1, 2]. Unfortunately many patients relapse after transplant. In this case, different treatment strategies can be used for induction of remission [4].

Materials and methods

Since 2014, 38 patients underwent 44 Haplo-SCTs at our centre. Six patients received second haplo-SCT because of graft failure or relapse. Five patients were transplanted from the same donor and one patient from another donor. Mean age was 38 years (21-61). Twenty two patients were males, and 16 females. The most common diagnosis was acute leukemia. Patients had AML (N=15), CML (N=7), HL (N=6), ALL (N=2), Burkitt lymphoma (N=2), T-lymphoblastic lymphoma (N=2), AL with mixed phenotype (N=1), NKleukemia (N=1), PMBCL (N=1) and primary myelofibrosis (N=1). 25 patients had relapsed or refractory disease. 15 patients proceeded to transplant without remission. Conditioning regimens were: Flu-Cy-Mel (N=22), Bu-FluCy (N=10), Flu-Cy-TBI (N= 3), Flamsa-RIC (N=3), other regimens (N=6). PT-Cy, MMF and CsA were used for GVHD prophylaxis. Stem cell sources were either peripheral blood (N=37) or bone marrow (N=7). Mean transplant CD34+cell dose was 4.09 Mln/kg (0.96 – 12.78).

Results

Mean time to engraftment was 27 days for platelets and 17 for neutrophils. 3 patients (7.8%) had primary transplant failure. Two of them had second transplantation from the same donor and one eventually engrafted. 8 patients proceeded to relapse (21%). Among them, 6 had acute leukemia (two had molecular relapse, and four, bone marrow relapse). Six patients were treated with DLI, azacitidine +DLI (in case of acute leukemia), 4 patients had second transplantation. Different chemotherapy regimens were used in relapse. Eventually, 6 patients achieved remission. 23 patients (61%) developed acute GVHD. 14 (37%) patients had Grade III-IV GVHD, all of them exhibited skin form, 7 (18%) patients had gastrointestinal form and 4 (11%) had hepatic form. Mean term of GVHD onset was 38 (13-130) days. 5 patients (13.2%) developed chronic GVHD (3 had skin damage, 1 had liver damage and 1 had muscle-skeletal form). Only 4 patients needed second-line GVHD treatment. Rate of hemorrhagic cystitis, a common complication of cyclophosphamide, was 8.9% (N=3). All cystitis episodes resolved on massive infusion therapy. By May 2016 OS of all patients was 55% (N=21), DFS was 47% (N=18). TRM was 16% (N=6). Rate of transplantrelated mortality (at < 30 days after transplantation) was 13% (N=5). Causes of death at late post-transplant period were as follows: infections (N=9), progression (N=3), GVHD (N=3), graft rejection (N=2).

Conclusion

Haplo-SCT is a possible alternative in patients without HLAcompatible donor. Conditioning regimen with Melphalan can be used instead of TBI-based regimens. GvHD prophylaxis with PT-Cy is feasible and effective. Different post-transplant strategies can be used for re-induction of remission in case of relapse. Disease status before transplantation has great influence on OS and PFS.

References

1. Kanakry CG, de Lima MJ, Luznik L. Alternative donor allogeneic hematopoietic cell transplantation for acute myeloid leukemia. SeminHematol. 2015;52(3):232-242.

2. Dietrich S, Finel H, Martinez C. Post-transplant cyclophosphamide-based haplo-identical transplantation as alternative to matched sibling or unrelated donor transplantation for non-Hodgkin lymphoma: a registry study by the European society for blood and marrow transplantation. Leukemia. 2016; 30(10):2086-2089.

3. Leo Luznik, Paul V. O’Donnell, J. Fuchs Ephraim. Post-transplantation cyclophosphamide for tolerance induction in HLA-haploidentical BMT. Semin Oncol .2012; 39(6):683-693

4. Amer M Zeidan, Patrick M Forde, Heather Symons. HLA-haploidentical donor lymphocyte infusions for patients with relapsed hematologic malignancies after related HLA-haploidentical bone marrow transplantation. Biol Blood Marrow Transplant. 2014; 20(3): 314–318.

Short reports

Efficiency of 5-azacytidine administration before allogeneic hematopoietic stem cell transplantation in acute myelobastic leukemia, myelodysplastic syndrome, and juvenile myelomonocytic leukemia

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Varvara N.Ovechkina, Sergej N.Bondarenko, Elena V.Morozova, Olga Slesarchuk, Kirill A.Ekushev, Ludmila S. Zubarovskaya, Boris V.Afanasyev

R.Gorbacheva Memorial Research Institute of Children Oncology, Hematology, and Transplantation, First St.Petersburg I.Pavlov State Medical University, St.Petersburg, Russian Federation

Contact: Dr. Varvara N.Ovechkina,
E-mail: ovetchkina@gmail.com

Introduction

The aim of our study was to assess overall survival (OS), time to progression and event-free survival (EFS) in patients with acute myelobastic leukemia (AML), myelodysplastic syndrome (MDS), juvenile myelomonocytic leukemia (JMML) who underwent therapy with hypomethylating agents (HMA) before before allogeneic hematopoietic stem cell transplantation (HSCT).

Patients and Methods

We have performed analysis of 54 cases including 15 AML (28%), 33 MDS ( 61%), 4 JMML (7%), 2 CMML(4%), at a mean age of 33 years old (3 mo to 61 years old). Male-to-female ratio was 54:46. Cytogenetic risk group at diagnosis was estimated as favorable in 22 cases (41%); intermediate, in 13 patients (24%), and unfavorable, in 18 cases (35%). In course of preparation for HSCT, 60% of the patients were treated with 5-azacytidine (5-Aza), 35% were administered Decytabine (Dec), and 5% of the patients were treated with the both drugs. The mean number of treatment rounds was (1 to 12). All the patients then underwent allo-HSCT including 11 transplants (20% of total) from a related HLA compatible donor; 30 patients (55%) were transplanted from matched unrelated donors. In 5 patients (10%), hematopoietic grafts were taken from unrelated HLA-compatible donors, and 8 transplants (15%) were performed from haploidentical donors.

Results

Eight patients exhibited progression despite the therapy performed; in 30 cases (56%), we have observed stabilization (S) of the disease, fourteen patients (26%) achieved partial remission (PR), and complete remission (CR) was observed in two cases (4%). CR or PR were registered after an average of 3 rounds of the therapy (1 to 5 courses). Progression of disease by the time of HSCT was documented in 10 patients (18%). The median OS value in the group was 778 days (95%CI, 346-1025 days). The OS terms differed significantly between the patients with AML (382 days; 95% CI, 134-1029) and MDS (1036 days; 95% CI, 619-1452, р=0.05). The median of time-to-progression comprised 593 days (95% CI, 169-1016). Posttransplant relapse developed in 19 cases (35% of total group). At the present time, 23 patients (43% of total) are alive. The main causes of death were as follows: relapse/progression of primary disease (13.4%), infection (10.3%), hemorrhagic complications (4.1%), graft-versushost disease (3.1%), and second malignancy (1.3%). The EFS median was 661 day (95% CI, 1-1346). Median EFS values were significantly higher for the patients who achieved stabilization or remission state (956 days; 95% CI, 689-1224) and for those who retained this state by the time of allo-HSCT (1062 days; 95% CI, 790-1333) as compared with patients who did not respond to hypomethylating therapy (300 days; 95% CI, 108-491) and had developed progression at the time of allo-HSCT (306 days; 95% CI, 171-442, p=0.02).

Conclusion

Hypomethylating agents may be drugs of choice for the patients with AML, MDS, CMML, and JMML during preparation for allo-HSCT. Taking into account the time-to-progression values in this cohort of patients, the start of HMA therapy should coincide with decision on unrelated donor search. Post-transplant EFS results proved to be better in HMA responders. To obtain more reliable results, further studies are required with larger numbers of patients

Short reports

High-dose cyclophosphamide after allogeneic stem cell transplantation as a graft-versus-host-disease prophylactic agent in adult acute leukemia: Experience of two Mexican hospitals

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Roberto Ovilla-Martínez1 , Uendy Pérez-Lozano2 , Diego Cruz-Contreras

1Hematology Department “Hospital Angeles Lomas”, Mexico City, México.
2Hematology Department of High Speciality Medical Unit “Manuel Ávila Camacho”, Mexican Institute of Social Insurance,
Puebla, México

Contact: Dr. Diego Cruz-Contreras
E-mail: dr_diego_cruz@hotmail.com

Introduction

Usage of high-dose Cyclophosphamide (HDCY) after allogeneic stem cell transplantation (alloSCT) as a prophylactic agent against acute graft-versus-host-disease (aGVHD) has been widespread worldwide with good results. Our aim was to assess incidence of aGVHD when applying HDCY as a prophylaxis agent in our patients.

Materials and methods.

Forty-eight patients were allo-transplanted between January 2012 and January 2016 including 22 patients (46%) from fully HLA-matched related donors, and 26 (54%), from haploidentical donors. Their median age was 34 (2-65) years old. Acute lymphoblastic leukemia was the most common diagnosis (79% of patients). Twenty-nine (60%) patients received myeloablative conditioning (MAC), and 19 (40%) were administered a reduced intensity conditioning (RIC). In alloSCT from fully HLA-matched related donors, we used peripheral blood stem cells in 12 (54%) patiens, and bone marrow in 10 (46%) patients; in all haploSCT we used bone marrow as a source of stem cells. All the patients received aGVHD prophylaxis with HDCY (50 mg/kg) day+3 and +4, haploSCT patients received also Tacrolimus and Mycophenolate Mofetil.

Results

Median neutrophilic engraftment term was on the +15 (MAC) and +17 (RIC). Graft failure was observed in 6 patients (MAC) and 3 patients (RIC). Incidence of aGVHD in total group was 41.6% (n=20), including grade I (5 patients); grade II (8 cases); grade III (3 patients), and grade IV in 4 cases. Skin was the most affected organ (n=11). Four patients relapsed, with median time of relapse at 266 (108-756) days post-transplant. Conditioning regimen was not a risk factor for aGVHD (p=0.349). A special risk factor for aGVHD was a transplant from female donor to male recipient (p=0.019). The presence of aGVHD was not predictive for death (RR 2.110, 95% IC, 0.645-6.902, p= 0.214). Median follow-up terms were 108 (6-999) days for MAC regimen, and 240 (27-939) days for RIC patients. Median OS was 15 months for haplo-SCT and 18 months for alloSCT from related donors. Median disease free survival (DFS) for haploSCT and alloSCT from related donor was, respectively, 28 months and 27 months. OS value was 39% for haploSCT and 56% for alloSCT from related donor.

Conclusions

HDCY application for aGVHD prophylaxis is a feasible scheme to use in Mexico being cheaper tolerable and effective. In our study, no patient died for aGVHD. Among total group, only 14% (48) had grade III/IV aGVHD.

References

1. Kanakry CG, et al. Multi-institutional study of post-transplantation cyclophosphamide as single- agent graft-versushost disease prophylaxis after allogeneic bone marrow transplantation using myeloablative busulfan and fludarabine conditioning. J Clin Oncol. 2014; 32(31):3497–3505.

1. Luznik L, et al. High-dose cyclophosphamide as single-agent, short-course prophylaxis of graft- versus-host disease. Blood. 2010; 115(16):3224–3230.

Short reports

Haploidentical bone marrow stem cell transplantation for hematological diseases: experience of single public Mexican hospital

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Uendy Pérez-Lozano, José Ruiz-Ovalle, Diego Cruz-Contreras

Hematology Department of High Speciality Medical Unit “Manuel Ávila Camacho”, Mexican Institute of Social Security, Puebla, Mexico

Contact: Dr. Diego Cruz
E-mail: dr_diego_cruz@hotmail.com

Introduction

Unavailability of complete HLA-matched related donors and lack of a bone marrow Registry in Mexico is one of our main problems when performing allogeneic stem cell transplantations (alloSCT) in the patients who require hematopoietic grafting. The aim of our work was to study haploidentical stem cell transplantation (haploSCT) as an alternative way for the patients lacking available stem cell donors.

Materials and methods

The study group included 20 patients (16 ALL, 1 SMD, 1 AML, 1 CML). Their median age was 30 (3-58) years old. Nine patients (45%) received reduced-intensity conditioning (RIC) regimens, and 11 (55%) received myeloablative conditioning (MAC) regimen. Bone marrow was used as a stem cell source. All the patients received high-dose cyclophosphamide (HDCY) on day +3 and +4, Tacrolimus and MMF as acute graft-versus-host-disease (aGVHD) prophylaxis.

Results

Overall survival (OS) in both cohorts was 58% (OS rate was 60% for MAC cohort, with a median follow-up of 23.8 months, and 57% for RIC group, follow-up of 14 months). Two patients (18%) relapsed in each cohort. Two patients died from infectious complications in MAC cohort, whereas none of the patients was lost for these reasons in RIC cohort. Two (18%) patients from each group had grade IV aGVHD, but neither patient died from aGVHD. Median values for neutrophilic and platelet engraftment in RIC cohort was 15.3/27.8 days, respectively. Apрropriate values for MAC cohort were 19.4/30.7 days. All the patients from RIC cohort reached 100% chimerism as compared with 55% in MAC group.

Conclusions

HaploSCT is a feasible transplant mode which may be performed in Mexico for all patients with agressive malignant hematological diseases who have no an available donor.

References

1. Kanakry CG, Fuchs EJ, Luznik L. Modern approaches to HLA-haploidentical blood or marrow transplantation. Nat Rev Clin Oncol. 2016; 13(1):10–24

2. Luznik L, et al. HLA-haploidentical bone marrow transplantation for hematologic malignancies using nonmyeloablative conditioning and high-dose, posttransplantation cyclophosphamide. Biol Blood Marrow Transplant. 2008; 14(6):641–650.

Short reports

Obituary. In memory of Professor Thomas Büchner

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Professor Sergey F. Bagnenko, Full Member, Russian Academy of Sciences,
Rector, First State Pavlov Medical University of St. Petersburg
Professor Boris V. Afanasyev,
Director, Raisa Gorbacheva Memorial Research Institute for Pediatric Oncology, Hematology and Transplantation, First State Pavlov Medical University of St. Petersburg, Russia
Prof. Dr. Dr. h.c. Rüdiger Hehlmann,
Med. Fakultät Mannheim, Universität Heidelberg, Deutschland

Professor Thomas Büchner

Professor Thomas Büchner, Professor Emeritus at the University of Münster, passed away on Friday, August 5, 2016. For the last decades, he was considered a renowned clinician, worldwide expert in leukemia, and a real symbol of German academic elite.

Thomas Büchner was born in Berlin on September 22, 1934. He graduated from the Albert-Ludwig University in Freiburg i.B. (1955). Following residences in Vienna, Innsbruck and Munich, he has took a Doctor’s degree in Freiburg (1961) with a Thesis: ‘Autoradiographic Studies of Cell Kinetics in Mice’. Then, over the next decades, his academic and clinical career was closely connected with Münster University Clinic. As an assistant, he worked with DNA synthesis and electron microscopy of human chromosomes, and flow cytometric techniques. In 1971, his publication “Inflammatory cells in Blood and Tissues” was awarded with the Theodor-Frerich Price from The German Society for Internal Medicine, followed by a professorship in Internal Medicine and Haematology at the Wilhelms-University in Münster. Since 1976, he managed a Laboratory of Special Haematology at the Medical Clinic of Münster University.

Clinical studies by Professor Büchner focused on acute myeloid leukemia (AML). In 1978, he became Chairman of the AML Cooperative Group. The well-known TAD protocol for leukemia treatment was established on the basis of these cooperative studies. In extension of his leukemia program, Prof. Büchner, together with his Münster colleague Günther Schellong (1926-2015), started the Acute Leukemias Symposium which is an important scientific and educational event since 1986. Within a program of leukemia studies, a Bone Marrow Transplantation Center was arranged at Münster University in the 90’s, along with the establishment of a Leukemia Studies Department. Professor Büchner held his academic positions until 1999 when was elected Emeritus Professor at Münster University. In 1999 Thomas became cofounder of the German Competence Network on Acute and Chronic Leukemias and started the German AML-Intergroup study that compared 5 randomized German AML studies by up-front randomization into a common standard arm. At the EU level, Prof. Büchner played a decisive role in establishing the European LeukemiaNet, a Europe-wide network promoting cure of leukemia by cooperative research.

During the last 15 years, Prof. Büchner was involved in research and treatment of acute myeloid leukemia in older patients. Several programs in clinical research were performed since 2005, including Biology and Treatment Strategy of AML in Its Subgroups: Multicenter Randomized Trial by the German Acute Myeloid Leukemia Cooperative Group (AMLCG).

Professor Büchner was a member of leading hematology and oncology associations in the field, including the American Society of Hematology, the American Association for Cancer Research and the German Society for Haematology and Medical Oncology (DGHO, Honorary Member since 2008).

Along with his professional merits, Prof. Büchner was well known for treating Raisa Maximovna Gorbachova (1932– 1999). In 2007 in memoriam of his wife Michail Gorbachov in cooperation with Pavlov University funded the construction of the Raisa Gorbacheva memorial Institute for Hematology and Transplantation, which started to work with support of leading German haematologists, including Prof. Büchner.

Prof. Büchner supported cooperation with Russian clinicians and scientists in the field of leukemia treatment from the very beginning of these contacts in the late 80’s. These long-lasting professional and friendly relationships have given the opportunity to know Thomas Büchner as the great person, tutor, physician and scientist he was. Not only his professional excellence inspired admiration, but also his sense of tact and dignity coupled with a dry humor formed the impression of an extraordinary person.

Prof. Büchner almost annually held special lectures at the Raisa Gorbacheva Memorial Symposium on Hematopoietic Stem cell Transplantation in St. Petersburg and other places. This year he was also expected at the 10th R. Gorbacheva Symposium in 2016. We shall always remember his life, clinical studies and good deeds for the patients. The fundamental academic school in Hematology and Oncology founded by Professor Büchner, will serve as heritage of his life. Apart from his scientific merits Thomas was a good hearted, sincere, always reliable and helpful colleague.

Thomas Büchner will always remain in our hearts.