Sara Trabanelli

Successful transfer of alloreactive haploidentical KIR ligand-mismatched natural killer cells after infusion in elderly high risk acute myeloid leukemia patients

Thirteen patients with acute myeloid leukemia, 5 with active disease, 2 in molecular relapse, and 6 in morphologic complete remission (CR; median age, 62 years; range, 53-73 years) received highly purified CD56(+)CD3(-) natural killer (NK) cells from haploidentical killer immunoglobulin-like receptor-ligand mismatched donors after fludarabine/cyclophosphamide immunosuppressive chemotherapy, followed by IL-2. The median number of infused NK cells was 2.74 × 10(6)/Kg. T cells were < 10(5)/Kg. No NK cell-related toxicity, including GVHD, was observed. One of the 5 patients with active disease achieved transient CR, whereas 4 of 5 patients had no clinical benefit. Both patients in molecular relapse achieved CR that lasted for 9 and 4 months, respectively. Three of 6 patients in CR are disease free after 34, 32, and 18 months. After infusion, donor NK cells were found in the peripheral blood of all evaluable patients (peak value on day 10). They were also detected in BM in some cases. Donor-versus-recipient alloreactive NK cells were shown in vivo by the detection of donor-derived NK clones that killed recipients targets. Adoptively transferred NK cells were alloreactive against recipients cells, including leukemia. In conclusion, infusion of purified NK cells is feasible in elderly patients with high-risk acute myeloid leukemia. This trial was registered at www.clinicaltrial.gov as NCT00799799.

The role of indoleamine 2,3-dioxygenase in the induction of immune tolerance: focus on hematology

The regulation of the interaction between the immune system and antigens, which may lead to the induction of immune tolerance, is critical both under physiologic conditions and in different pathological settings. In the past few years, major strides have been made in our understanding of the molecular and cellular bases of this process. Novel pathways have been identified and several novel therapeutic agents are currently under clinical investigation for those diseases in which the normal balance between activation and suppression of the immune response is altered. The tryptophan catabolic enzyme, indoleamine 2,3-dioxygenase (IDO), is one of the key players involved in the inhibition of cell proliferation, including that of activated T cells. Recent works have demonstrated a crucial role for IDO in the induction of immune tolerance during infection, pregnancy, transplantation, autoimmunity, and neoplasias, including hematologic malignancies. In this review, the role of IDO in the induction of immunologic tolerance is addressed with a specific focus on its recently discovered effect on hematologic malignancies.

Extracellular ATP Exerts Opposite Effects on Activated and Regulatory CD4+ T Cells via Purinergic P2 Receptor Activation

It has been reported that ATP inhibits or stimulates lymphoid cell proliferation depending on the cellular subset analyzed. In this study, we show that ATP exerts strikingly opposite effects on anti-CD3/CD28–activated and regulatory CD4+ T cells (Tregs), based on nucleotide concentration. We demonstrate that physiological concentrations of extracellular ATP (1–50 nM) do not affect activated CD4+ T cells and Tregs. Conversely, higher ATP concentrations have a bimodal effect on activated CD4+ T cells. Whereas 250 nM ATP stimulates proliferation, cytokine release, expression of adhesion molecules, and adhesion, 1 mM ATP induces apoptosis and inhibits activated CD4+ T cell functions. The expression analysis and pharmacological profile of purinergic P2 receptors for extracellular nucleotides suggest that activated CD4+ T cells are induced to apoptosis via the upregulation and engagement of P2X7R and P2X4R. On the contrary, 1 mM ATP enhances proliferation, adhesion, migration, via P2Y2R activation, and immunosuppressive ability of Tregs. Similar results were obtained when activated CD4+ T cells and Tregs were exposed to ATP released by necrotized leukemic cells. Taken together, our results show that different concentrations of extracellular ATP modulate CD4+ T cells according to their activated/regulatory status. Because extracellular ATP concentration highly increases in fast-growing tumors or hyperinflamed tissues, the manipulation of purinergic signaling might represent a new therapeutic target to shift the balance between activated CD4+ T cells and Tregs.

Indoleamine 2,3-dioxygenase-expressing leukemic dendritic cells impair a leukemia-specific immune response by inducing potent T regulatory cells

Background The immunoregulatory enzyme indoleamine 2,3-dioxygenase, which catalyzes the conversion of tryptophan into kynurenine, is expressed in a significant subset of patients with acute myeloid leukemia, resulting in the inhibition of T-cell proliferation and the induction of regulatory T cells. Acute myeloid leukemia cells can be differentiated into dendritic cells, which have increased immunogenicity and have been proposed as vaccines against leukemia. Design and Methods Leukemic dendritic cells were generated from acute myeloid leukemia cells and used as stimulators in functional assays, including the induction of regulatory T cells. Indoleamine 2,3-dioxygenase expression in leukemic dendritic cells was evaluated at molecular, protein and enzymatic levels. Results We demonstrate that, after differentiation into dendritic cells, both indoleamine 2,3-dioxygenase-negative and indoleamine 2,3-dioxygenase-positive acute myeloid leukemia samples show induction and up-regulation of indoleamine 2,3-dioxygenase gene and protein, respectively. Indoleamine 2,3-dioxygenase-positive acute myeloid leukemia dendritic cells catabolize tryptophan into kynurenine metabolite and inhibit T-cell proliferation through an indoleamine 2,3-dioxygenase-dependent mechanism. Moreover, indoleamine 2,3-dioxygenase-positive leukemic dendritic cells increase the number of allogeneic and autologous CD4+CD25+ Foxp3+ T cells and this effect is completely abrogated by the indoleamine 2,3-dioxygenase-inhibitor, 1-methyl tryptophan. Purified CD4+CD25+ T cells obtained from co-culture with indoleamine 2,3-dioxygenase-positive leukemic dendritic cells act as regulatory T cells as they inhibit naive T-cell proliferation and impair the complete maturation of normal dendritic cells. Importantly, leukemic dendritic cell-induced regulatory T cells are capable of in vitro suppression of a leukemia-specific T cell-mediated immune response, directed against the leukemia-associated antigen, Wilms’ tumor protein. Conclusions These data identify indoleamine 2,3-dioxygenase-mediated catabolism as a tolerogenic mechanism exerted by leukemic dendritic cells and have clinical implications for the use of these cells for active immunotherapy of leukemia.

Larger Size of Donor Alloreactive NK Cell Repertoire Correlates with Better Response to NK Cell Immunotherapy in Elderly Acute Myeloid Leukemia Patients

Purpose: In acute myeloid leukemia (AML), alloreactive natural killer (NK) cells are crucial mediators of immune responses after haploidentical stem cell transplantation. Allogeneic NK cell infusions have been adoptively transferred with promising clinical results. We aimed at determining whether the composition of NK graft in terms of frequency of alloreactive NK cells influence the clinical response in a group of elderly AML patients undergoing NK immunotherapy. Experimental Design: Seventeen AML patients, in first complete remission (CR; median age 64 years, range 53–73) received NK cells from haploidentical KIR-ligand–mismatched donors after fludarabine/cyclophosphamide chemotherapy, followed by IL2. To correlate donor NK cell activity with clinical response, donor NK cells were assessed before and after infusion. Results: Toxicity was moderate, although 1 patient died due to bacterial pneumonia and was censored for clinical follow-up. With a median follow-up of 22.5 months (range, 6–68 months), 9 of 16 evaluable patients (0.56) are alive disease-free, whereas 7 of 16 (0.44) relapsed with a median time to relapse of 9 months (range, 3–51 months). All patients treated with molecular disease achieved molecular CR. A significantly higher number of donor alloreactive NK cell clones was observed in responders over nonresponders. The infusion of higher number of alloreactive NK cells was associated with prolonged disease-free survival (0.81 vs. 0.14, respectively; P = 0.03). Conclusions: Infusion of purified NK cells is feasible in elderly AML patients as post-CR consolidation strategy. The clinical efficacy of adoptively transferred haploidentical NK cells may be improved by infusing high numbers of alloreactive NK cells. Clin Cancer Res; 22(8); 1914–21. ©2016 AACR. See related commentary by Muntasell and López-Botet, p. 1831

The SOCS3-Independent Expression of IDO2 Supports the Homeostatic Generation of T Regulatory Cells by Human Dendritic Cells

Dendritic cells (DCs) are professional APCs that have a role in the initiation of adaptive immune responses and tolerance. Among the tolerogenic mechanisms, the expression of the enzyme IDO1 represents an effective tool to generate T regulatory cells. In humans, different DC subsets express IDO1, but less is known about the IDO1-related enzyme IDO2. In this study, we found a different pattern of expression and regulation between IDO1 and IDO2 in human circulating DCs. At the protein level, IDO1 is expressed only in circulating myeloid DCs (mDCs) and is modulated by PGE2, whereas IDO2 is expressed in both mDCs and plasmacytoid DCs and is not modulated by PGE2. In healthy subjects, IDO1 expression requires the presence of PGE2 and needs continuous transcription and translation, whereas IDO2 expression is constitutive, independent from suppressor of cytokine signaling 3 activity. Conversely, in patients suffering from inflammatory arthritis, circulating DCs express both IDO1 and IDO2. At the functional level, both mDCs and plasmacytoid DCs generate T regulatory cells through an IDO1/IDO2-dependent mechanism. We conclude that, in humans, whereas IDO1 provides an additional mechanism of tolerance induced by proinflammatory mediators, IDO2 is stably expressed in steady-state conditions and may contribute to the homeostatic tolerogenic capacity of DCs.

Decreased expression of indoleamine 2,3-dioxygenase 1 in dendritic cells contributes to impaired regulatory T cell development in immune thrombocytopenia

Along with their immunogenic role, dendritic cells (DCs) are also critical in maintaining tolerance to self-antigens by inducing regulatory T cells (Tregs) via the expression of the immunomodulatory enzyme indoleamine 2,3-dioxygenase 1 (IDO1). In turn, Tregs modulate the maturation and/or function of DCs. In immune thrombocytopenia (ITP), the interaction between DCs and Tregs has never been investigated although decreased number/function of Tregs as well as altered DCs have been described. Here, we ask whether, in ITP: (1) IDO1 expression/activity is decreased in mature DCs; (2) IDO1-mediated Treg generation is impaired; and (3) DC maturation is abnormally modulated by Tregs. We found that in ITP, DCs show reduced capability of upregulating the expression/activity of IDO1. This finding results in the reduced ability of mature DCs of converting T cells into Tregs. In turn, Tregs are characterized by decreased interleukin-10 production and show lower ability of inhibiting DC maturation. In conclusion, these data point out the role of IDO1 in the impaired regulatory T cell development of ITP patients and suggest that the cross-talk between Tregs and DCs is hampered and plays a pathogenetic role.

CD127+ innate lymphoid cells are dysregulated in treatment naïve acute myeloid leukemia patients at diagnosis

Innate lymphoid cells (ILCs), defined as lineage negative (Lin−) CD127+ cells, have emerged as novel important immune effector cells, playing a critical role also in tumor immunosurveillance.[1][1]–[5][2] Recently, it was shown that the number of ILCs in acute myeloid leukemia (AML) patients is

Circulating CD4+CD161+CD196+ Th17 cells are not increased in immune thrombocytopenia

Immune thrombocytopenia (ITP) is an autoimmune disorder in which, for still unknown reasons, platelet surface proteins become antigenic and stimulate the immune system to produce autoantibodies and self-reactive cytotoxic T lymphocytes. These findings result in immune-induced platelet destruction

Induction of Regulatory T Cells by Dendritic Cells through Indoleamine 2,3- dioxygenase: A Potent Mechanism of Acquired Peripheral Tolerance

Indoleamine 2,3-dioxygenase (IDO) is an intracellular heme-containing enzyme that catalyzes the initial rate-limiting step in tryptophan degradation along the kynurenine pathway. Recent works have demonstrated a crucial role for IDO in the induction of immune tolerance during infections, pregnancy, transplantation, autoimmunity, and neoplasias. IDO is widely expressed in human tissues and cell subsets, including dendritic cells, where it modulates their function by increasing tolerogenic capacities. The aim of the present paper is to highlight the most recent data about IDO expression in dendritic cells and its role as a potent inducer of T regulatory cells.