IntroductionA viable option for high-risk, acute leukemia patients without matched donors is hematopoietic stem cell transplantation (HSCT) from human leukocyte antigen (HLA)-haploidentical 3-locimismatched family members who were readily available for almost all patients. 1,2 Until the 1990s, full-haplotype-mismatched, T cell-replete transplantations were unsuccessful because donoralloreactive T cells triggered a high incidence of severe graft-versushost disease (GVHD) despite posttransplantation immune suppression. 3,4 The breakthrough came with the use of a megadoses of extensively ex vivo T cell-depleted peripheral blood hematopoietic progenitor cells and a highly myeloablative conditioning regimen containing antithymocyte globulin (ATG), which exerts additional T-cell depletion in vivo. This approach ensures a high rate of primary engraftment in the absence of GVHD, 5 with more than 40% long-term event-free survival and excellent quality of life. 1,2 However, extensive ex vivo and in vivo T-cell depletion delays the recovery of immune responses against pathogens, leading to a high incidence of life-threatening infections. 1,2 Strategies to hasten posttransplantation immune reconstitution without triggering GVHD have included infusion of donor T cells after engineering with a suicide gene, 6 photodynamic purging, 7 and the use of an anti-CD25 monoclonal antibody (mAb) 8 to remove alloreactive cells. An alternative strategy might be based on donor CD4 ϩ CD25 ϩ T-regulatory cells (Tregs). In murine models of HSCT across major histocompatibility complex barriers, CD4 ϩ CD25 ϩ Tregs suppressed lethal GVHD 9 and favored posttransplantation immune reconstitution when coinfused with conventional T cells (Tcons). 10 The main obstacle to clinical application of human Tregs is their paucity in the peripheral blood. Although ex vivo-expanded polyclonal 11 or recipient-specific Tregs 12 were proposed to circumvent this potential barrier, we opted for closed, automated immunoselection 13 of naturally occurring Tregs. In the present study, for the first time in humans, we show that the early infusion of freshly isolated donor Tregs followed by Tcons at the time of full-haplotypemismatched HSCT prevented GVHD while favoring Tconmediated posttransplantation immune reconstitution. MethodsStudy design, conditioning regimen, stem cell mobilization, and supportive careIn 2008, the Umbria Regional Hospital Ethics Committee (CEAS Umbria) approved the protocol entitled "Adoptive Immunotherapy with Natural Regulatory T cells (Treg) and Effector T Cells in Allogeneic Hematopoietic Stem Cell Transplantation from 2-3 Loci Mismatched HLA-Haploidentical Family Donors for Patients with High Risk Haematologic Malignancies" (Protocol No 01/08). Written informed consent was obtained for all patients and donors in accordance with the Declaration of Helsinki. Inclusion criteria were: acute myeloid leukemia (AML) or acute lymphoid leukemia (ALL) in remission at high risk of relapse; acute leukemia with primary induction failure, in chemoresist...

Posttransplant relapse is still the major cause of treatment failure in high-risk acute leukemia. Attempts to manipulate alloreactive T cells to spare normal cells while killing leukemic cells have been unsuccessful. In HLA-haploidentical transplantation, we reported that donor-derived T regulatory cells (Tregs), coinfused with conventional T cells (Tcons), protected recipients against graft-versus-host disease (GVHD). The present phase 2 study investigated whether Treg-Tcon adoptive immunotherapy prevents posttransplant leukemia relapse. Forty-three adults with high-risk acute leukemia (acute myeloid leukemia 33; acute lymphoblastic leukemia 10) were conditioned with a total body irradiation-based regimen. Grafts included CD34(+) cells (mean 9.7 × 10(6)/kg), Tregs (mean 2.5 × 10(6)/kg), and Tcons (mean 1.1 × 10(6)/kg). No posttransplant immunosuppression was given. Ninety-five percent of patients achieved full-donor type engraftment and 15% developed ≥grade 2 acute GVHD. The probability of disease-free survival was 0.56 at a median follow-up of 46 months. The very low cumulative incidence of relapse (0.05) was significantly better than in historical controls. These results demonstrate the immunosuppressive potential of Tregs can be used to suppress GVHD without loss of the benefits of graft-versus-leukemia (GVL) activity. Humanized murine models provided insights into the mechanisms underlying separation of GVL from GVHD, suggesting the GVL effect is due to largely unopposed Tcon alloantigen recognition in bone marrow.

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Chronic lymphocytic leukemia (CLL) is an incurable B-cell neoplasm characterized by highly variable clinical outcomes. In recent years, genomic and molecular studies revealed a remarkable heterogeneity in CLL, which mirrored the clinical diversity of this disease. These studies profoundly enhanced our understanding of leukemia cell biology and led to the identification of new biomarkers with potential prognostic and therapeutic significance. Accumulating evidence indicates a key role of deregulated NOTCH1 signaling and NOTCH1 mutations in CLL. This review highlights recent discoveries that improve our understanding of the pathophysiological NOTCH1 signaling in CLL and the clinical impact of NOTCH1 mutations in retrospective and prospective trials. In addition, we discuss the rationale for a therapeutic strategy aiming at inhibiting NOTCH1 signaling in CLL, along with an overview on the currently available NOTCH1-directed approaches.