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.

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.

Nonadherent Spheres With Multiple Myeloma Surface Markers Contain Cells that Contribute to Sphere Formation and Are Capable of Internalizing Extracellular Double-Stranded DNA.

BACKGROUND: The most prominent features of cancer stem cells are asymmetric cell division, tumorigenicity, and clonogenicity. Recently one more feature of poorly differentiated cell types of various origin, including cancer stem cells, has been described. Namely, these cells can internalize extracellular DNA natively, without additional transfection procedures. PATIENTS AND METHODS: Using our approach to trace internalization of a TAMRA (carboxy tetramethyl-rhodamine [fluorescent dye])-DNA labeled probe by poorly differentiated cell types, we isolated and characterized the cells from free-floating spheres derived from the bone marrow clonogenic aspirate of a multiple myeloma patient. RESULTS: Nonadherent spheres display a B-cell phenotype (CD73/CD20+/CD45+/CD19dim). Further, free-floating spheres contain 1% to 3% cells with a clonogenic potential, and these cells display a marker of poorly differentiated cell types (TAMRA+). Upon association with a group of ∼ 10 free-floating TAMRA- cells, this peculiar cell type forms a sphere-forming cluster that initiates secondary aggregation of cells into a spheric structure. TAMRA+ and TAMRA- cells secrete distinct sets of cytokines indicative of the paracrine regulation. Grafting experiments of intact whole spheres versus cell suspensions prepared from dispersed spheres indicate that successful engraftment only occurs in the former case. CONCLUSION: Nonadherent 3-D cell colonies (spheres) encompass B cells with CD73/CD20+/CD45+/CD19dim phenotype, as well as double-stranded DNA-internalizing cells. The latter cell type appears to function as a sphere-forming center. Different cells in the spheres communicate with each other by secreting specific sets of cytokines. For successful engraftment and tumor growth in mice, intact spheres containing ∼ 106 cells must be used.