Decreased circulating myeloid-derived suppressor cell count at the engraftment is one of the risk factors for multiple myeloma relapse after autologous hematopoietic stem cell transplantation.

Recent studies demonstrated that myeloid-derived suppressor cells (MDSCs) are involved in the pathogenesis and progression of multiple myeloma (MM). Nevertheless, data on the quantitative and functional changes in MDSCs during standard MM treatment remain poorly understood. Here, we determined that monocytic MDSCs (M-MDSC; CD14+HLA-DRlow/-) and granulocytic MDSCs (PMN-MDSC; Lin-HLA-DR-CD33+CD66b+) in MM patients in remission following induction therapy (IT) were significantly increased, while early MDSCs (E-MDSCs; Lin-HLA-DR-CD33+CD66b-) were decreased compared to the donor group. In progression, MM patients had the most pronounced decrease in E-MDSCs and enhanced levels of PMN-MDSCs. IT was accompanied with a decrease in the expression of arginase-1 (Arg-1). In MM patients with relapse or resistance to IT, Arg-1+ cell frequency in M-MDSCs and E-MDSCs, as well as PD-L1+ M-MDSCs, was increased, which may facilitate tumor immunosuppression. G-CSF administration led to a significant increment in the MDSC subsets. At the engraftment, circulating M-MDSC and PMN-MDSCs were temporarily increased, with a gradual decline to the pre-transplant levels in 12 months. The percentage of E-MDSCs was decreased at the leukocyte recovery. Patients with a higher (>Me) M-MDSC count at the engraftment had a shorter post-transplant leukopenia duration (Me 11 vs. 13 days; pU = 0.0086). The advanced MM stage, depth of response, and lower relative count of circulating E-MDSCs at the engraftment were independent risk factors associated with a lower progression-free survival. The obtained data allow us to hypothesize that MDSCs may play a positive role at the stage of leukocyte recovery by ameliorating the long-term anti-tumor response in MM.

Highly proliferative and functional PD-1+ and TIM-3+ T cells are transiently increased in multiple myeloma following autologous hematopoietic stem cell transplantation.

The aim of our prospective study was to assess recovery dynamics and functional characteristics of PD-1+ and TIM-3+ T cells in multiple myeloma (MM) patients following high-dose chemotherapy (HDCT) with autologous hematopoietic stem cell transplantation (AHSCT). Peripheral blood, autograft and bone marrow samples were obtained from 46 MM patients before conditioning, at the engraftment, following six and 12 months post-transplant. Frequencies of CD4+ and CD8+ T cells expressing PD-1 and TIM-3 and intracellular expression of Ki-67 and Granzyme B were evaluated. Counts of PD-1+ and TIM-3+ T cells at the engraftment were significantly higher comparing with the levels before HDCT and 6-12 months following AHSCT. The post-transplant increase in the studied subsets was due to a temporary enhancement in proliferation activity. The cytotoxic potential of PD-1- and TIM-3-expressing CD8+ T cells was higher at the engraftment comparing with the pre-transplant and remained at the same level for at least 12 months. The increase in CD4+PD-1+ and CD8+TIM-3+ T cells at the engraftment was associated with higher absolute counts of their reinfused counterparts. Circulating PD-1+ CD8+ and TIM-3+ CD4+ T cells were increased in patients after post-transplant relapse comparing with the ones in remission. Homeostatic proliferation plays a key role in the upregulation of inhibitory checkpoint receptors on functional T cells under lymphopenic conditions. In this regard, it is difficult to predict both the efficacy and adverse reactions of therapy with checkpoint inhibitors on the course of MM after HDCT with AHSCT. Précis. Homeostatic proliferation plays apparently a key role in the upregulation of PD-1 and TIM-3 on functional T cells after AHSCT and appears to be a normal physiological process, contrary to relapse-associated increase in PD-1+ and TIM-3+ T cells.

Quantitative and functional characteristics of circulating and bone marrow PD-1- and TIM-3-positive T cells in treated multiple myeloma patients.

The aim of the present work was to evaluate counts and functional properties of PD-1+ and TIM-3+ T cells in peripheral blood (PB) and bone marrow (BM) of multiple myeloma (MM) patients following the induction therapy. Sixty patients were enrolled in the study, CD4+ and CD8+ T cells expressing PD-1 and TIM-3, intracellular production of IFNγ and intracellular expression of Granzyme B were assessed. Relative counts of the majority of circulating PD-1+, TIM-3+ and PD-1+TIM-3+ T cells were higher in MM patients with disease progression compared with individuals in remission. Frequencies of almost all evaluated PD-1+ and TIM-3+ T cell subsets were higher in BM samples compared with PB; circulating CD4+PD-1+, CD8+PD-1+, CD8+TIM-3+, CD8+PD-1+TIM-3+ T cells positively correlated with the same BM subsets. Circulating CD4+ T cells, expressing PD-1 and TIM-3 (including co-expressing subset), as well as CD8+PD-1+TIM-3+ T cells, and BM CD8+PD-1+ T cells correlated with serum B2-M levels. Sufficient frequencies of GrB+ and IFNγ+ subsets in PD-1-expressing T cells indicated their retained functional properties. TIM-3-expressing T cells and double positive PD-1+TIM-3+ populations showed diminished cytotoxic and cytokine-producing ability and therefore might be attributed to the exhausted compartment. To identify T cell exhaustion, it is necessary to evaluate T cells co-expressing PD-1, TIM-3 and other inhibitory signal molecules and to study their functional properties. Sustained functionality of PD-1-positive T cells may explain low efficacy and frequent immune-mediated adverse events during anti-PD-1 therapy in MM.

Binding of the placental growth factor to VEGF receptor type 1 modulates human T cell functions.

The immunosuppressive properties of vascular endothelial growth factors (VEGFs) suggest a new role of angiogenic factors in T cell modulation in cancer and pregnancy. Most of VEGF effects on T cells are mediated through the VEGF receptor type 2 (VEGFR-2). This study aims to investigate the role of placental growth factor (PlGF) as a selective VEGFR-1 ligand in the modulation of human T cells functions. For this, PBMCs from healthy donors were stimulated with anti-CD3 mAbs (a-CD3) or Concanavalin A (ConA) in the absence or presence of PlGF and assessed for T cell proliferation, IL-10 production, programmed cell death, and the expression of inhibitory receptors (PD-1, CTLA-4, TIM-3) using radiometric (3 H-thymidine incorporation) and FACS analysis. We showed that most T cells in freshly isolated PBMCs lacked VEGFR-1. However, activation with a-CD3 or ConA strongly increased the percentages of VEGFR-1 expressing CD4+ and CD8+ T cells. PlGF in a wide dose range suppressed PBMC cell proliferation, inhibiting both CD4+ and CD8+ T cells. Blockade of VEGFR-1, but not VEGFR-2 with neutralizing Abs completely abolished the suppressive effect of PlGF. Furthermore, we found that treatment with PlGF up-regulated IL-10 production in CD4+ and CD8+ T cells, promoted CD8+ T cells apoptosis and enhanced the expression of inhibitory receptors (PD-1 and TIM-3) on activated T cells. Our in vitro findings suggest the involvement of PlGF/VEGFR-1 signaling in the modulation of T cell responses in a-CD3-stimulated PBMCs.

Increased circulating CD3+ T cells are associated with early relapse following autologous hematopoietic stem cell transplantation in patients with classical Hodgkin lymphoma.

Non-malignant host immune cells are the main substrate in classical Hodgkin lymphoma (HL) microenvironment. Reconstitution of lymphocyte populations following the high-dose chemotherapy (HDC) with autologous hematopoietic stem cell transplantation (auto-HSCT) can support tumor growth in HL patients. We investigated recovery dynamics of circulating CD3+, CD4+, CD8+, CD16+/CD56+, CD19+, CD4+FOXP3+ lymphocytes following auto-HSCT in 79 HL patients and assessed relationship between these populations and the development of early relapse. Studied populations were not statistically significant between patients with high or standard/intermediate risk of relapse. CD3+ T cells at the time of engraftment were increased in patients with the early relapse of HL compared to non-relapsed patients (PU = 0.0028). Area under the curve was 0.76 (р = .0037). In logistic regression models, CD3+ T cell count was associated with early relapse/progression as a trend. These findings elucidate several interactions between early systemic T cell recovery and tumor progression following HDC with auto-HSCT.

Increased circulating CD4+FOXP3+ T cells associate with early relapse following autologous hematopoietic stem cell transplantation in multiple myeloma patients.

We investigated dynamics of CD4+FOXP3+ T cell recovery following the high-dose chemotherapy (HDC) with autologous hematopoietic stem cell transplantation (auto-HSCT) in multiple myeloma (MM) patients. Circulating CD4+FOXP3+ T cells of 79 MM patients were evaluated using flow cytometry before HDC with auto-HSCT, at the day of engraftment, and following 6 and 12 months. Percentage of CD4+FOXP3+ T cells restored rapidly following auto-HSCT, became higher than pre-transplant level at the day of engraftment and then subsequently decreased for a year. CD4+FOXP3+ T cells at the time of engraftment were increased in patients with the relapse or progression of MM during 12 months following auto-HSCT (n=10) compared to non-relapsed patients (n=50): 6.7% (5.3-8.9%) vs 4.9% (2.8-6.6%); PU = 0.026. Area under the curve was 0.72 (95% CI: 0.570-0.878; р=0.026). Circulating CD4+FOXP3+ T cell count was not associated with the percentage of myeloma plasma cells in a bone marrow but depended on its amount in autografts. CONCLUSIONS: Relative count of CD4+FOXP3+ T cells restored rapidly following auto-HSCT (at the day of engraftment), became higher than pre-transplant level and then subsequently decreased for a year. Their excess at the time of engraftment is associated with early relapse.

CD4+ memory T cells retain surface expression of CD31 independently of thymic function in patients with lymphoproliferative disorders following autologous hematopoietic stem-cell transplantation.

High-dose chemotherapy with autologous hematopoietic stem-cell transplantation (AHSCT) causes severe and long-lasting immunodeficiency in patients with lymphoproliferative disorders. The thymus begins to restore the T-cell repertoire approximately from the sixth month post-transplant. We assessed the dynamics of post-transplant recovery of CD4+CD45RA+CD31+ T cells, "recent thymic emigrants" (RTEs), and a poorly described subtype of CD4+CD45RA-CD31+ T cells in 90 patients with lymphoproliferative disorders following high-dose chemotherapy with AHSCT. Relative and absolute counts of CD4+CD31+ naïve and memory T cells were evaluated before AHSCT, at the day of engraftment, and 6- and 12-month post-transplant. The pre-transplant count of CD4+CD45RA+CD31+ T cells was lower than in healthy controls, and did not reach donors' values during the 12-month period. The pre-transplant number of CD4+CD45RA-CD31+ T cells was higher than in healthy controls and was restored rapidly following AHSCT. Post-transplant mediastinal radiotherapy reduced counts of RTEs and elongated recovery period. Non-thymic tissue irradiation did not reduce this subset. The obtained data indicate that homeostatic proliferation may decrease the significance of CD31 expression on CD4+CD45RA+ T cells as a marker of RTEs, and suggest that evaluation of RTEs recovery by flow cytometry requires an accurate gating strategy to exclude CD31+ memory T cells.

Mesenchymal stromal cells improve early lymphocyte recovery and T cell reconstitution after autologous hematopoietic stem cell transplantation in patients with malignant lymphomas.

Mesenchymal stromal cells (MSCs) possess a multi-lineage potential and immunoregulatory activities and provide a great potential in cell-based technologies. However, MSC suppressive activity raises concerns regarding the possible adverse effect of MSCs on the immune recovery. The influence of autologous MSC co-transplantation on recovery of T cell subsets in patients receiving autologous hematopoietic stem cell transplantation (AHSCT) for malignant lymphomas and multiple myeloma were characterized. Co-transplantation of MSCs improved lymphocyte recovery most effectively in patients with low input of hematopoietic stem cells or low absolute lymphocyte count in apheresis product. MSC co-transplantation improved early recovery of both memory and naive T cells with more prominent effect on naive CD4(+) T cells. Patients with MSC co-transplantation showed more effective reconstitution of recent thymic emigrants. These data indicate the positive impact of MSCs on immune reconstitution and note MSC co-transplantation is feasible to optimize the outcomes of AHSCT in malignant lymphoma patients.