Various Gvhd Prophylaxis Regimens Influence Differently T-Memory Cell Subsets Recovery after Allogeneic Stem Cell Transplantation in Leukemia Patients
Authors: Drokov Mikhail , Popova Natalia , Parovichnikova Elena , Kuzmina Larisa , Davydova Julia , Kapranov Nikolay , Mikhaltsova Ekaterina , Koroleva Olga , Vasilyeva Vera , Dubnyak Daria , Konova Zoya , Sidorova Anna , Usikova Ekterina , Dmitrova Anna , Galtseva Irina V. , Maschan Mikhail , Savchenko Valery G. 
Published: October 2019

Abstract
Introduction. Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is a curative approach for most leukemia patients. It seems that the bone marrow (BM) resident T-memory cells are the main pool for reconstitution of adaptive immunity, providing graft-versus-leukemia effect after allo-HSCT. However it has been shown previously that T-naïve cell subsets are responsible for acute graft versus host disease (aGVHD) onset. Here we report an impact of different GVHD prophylaxis on the BM resident memory T-cells recovery after allo-HSCT.
Patients and methods. Our study comprised 39 leukemia patients (AML-26, ALL - 13) with a median age of 36 years (17 - 60 years old) who underwent allo-HSCT. Classical immunosuppressive regimen with CsA+MMF+MTX was applied in case of matched related (n=9) and matched unrelated (n=13) donors. Post-transplant high dose Cyclophosphamide (PT-Cy) on day +3, +4 as GVHD prophylaxis was used if allo-HSCT was performed from unrelated mismatched donors (n=8) or haplo-donors (n=4). Alfa/beta T-cells depletion was used as GVHD prophylaxis in young patients with a median age of 19 years (17-57 y.o.) who underwent haplo-HSCT (n=5). Acute GVHD grade II-III was diagnosed in 6 patients (27%) from the group with classical immunosuppressive regimen, in 2 patients (16,6%) from PT-Cy group. Acute skin GVHD grade II was diagnosed in 1 patient (20%) after T-cells depletion. Leukemia relapse was diagnosed after the day +100 in 4 patients (18%) after classic immunosuppressive regimen and 1 patient (8,3%) after PT-Cy. All 5 patients after T-cells depletion stayed in complete remission during follow-up period in 34 months after allo-HSCT. Bone marrow (BM) samples were collected on day +30, +60 and +90 after allo-HSCT in EDTA-tubes. Flow cytometry analysis was performed on BD FACS Canto II (Becton Dickinson, USA) to define T-memory subsets: T-naive and T-stem cell memory (Tnv+Tscm) - CD8+/CD4+ CD45R0-CCR7+CD28+; T-central memory (Tcm) - CD8+/CD4+ CD45R0+CCR7+CD28+; T-transitional memory (Ttm) - CD8+/CD4+ CD45R0+CCR7-CD28+; T-effector memory (Tem) - CD8+/CD4+ CD45R0+CCR7-CD28-; T-terminal effector (Tte) - CD8+/CD4+ CD45R0-CCR7-CD28-. Mann-Whitney U test was used for nonparametric data analysis. A p-value less than 0,05 was considered as significant. All data analysis was conducted utilizing SPSS ver. 23. (IBM, Chicago, Ill., USA).
Results. The data are presented in the picture 1. According to our data the number of BM resident Tnv+Tscm CD8+ or CD4+ cells are much lower after alternative regimes of GVHD prophylaxis (PT-Cy or T cells depletion) on day +30 after allo-HSCT. Moreover on day +60 Tnv+Tscm CD4+ T-cells are still extremely low after T-cells depletion comparing to PT-Cy (p=0,03) and classical regimen (p=0,034). We didn't found any correlation between the number of Tnv+Tscm, Tcm, Ttm, Tem, Tte and the development of leukemia relapse or acute GVHD.
Conclusion. New immunosuppressive approaches are now available for GVHD prophylaxis for patients who undergoes allo-HSCT from mismatched or haplo- donors that provides comparable results with classical immunosuppression in case of matched allo-HSCT. However Tnv+Tscm is the progenitor subset for all memory T-cells and its prompt recovery after allo-HSCT seems to be crucial for graft-versus-leukemia effect. Here we show the lowest number of Tnv+Tscm after alternative GVHD prophylaxis regimens that can be explained by more sever immunoablation. Through this it may lead to delayed reconstitution of adaptive immunity after alternative immunosuppression in patients after allo-HSCT.

Disclosures
No relevant conflicts of interest to declare.