Study of the immune reconstitution in patients receiving a reduced intensity stem cell transplantation for Aplastic Anemia

Aplastic anemia is a rare hematological disease, prototype of bone marrow failure syndromes, characterized by a progressive reduction of bone marrow stem cells that leads to classic histological finding of “fatty” or “empty” marrow. Clinical picture is dominated by bone marrow failure symptoms, such as fatigue, petechiae and/or fever that can rapidly deteriorate to potentially fatal bleeding, or life-threatening blood sepsis. Aplastic Anemia commonly affects 1 to 2/106 individuals per year, with an estimated incidence about 2-3 times higher in Eastern countries. Although the etiology remains unknown in most cases, approximately one fifth of patients recognize a triggering event like: exposure to toxins (eg. Benzene) and / or drugs (eg. NSAIDs, gold salts, etc .) and / or viral agents (eg. Parvovirus B19, EBV, etc.). Rarely the disease follows an episode of self-limiting, sero-negative hepatitis. Regardless of this, pathogenesis of Aplastic Anemia is immune mediated, and is determined by cytotoxic action of T-lymphocytes against the hematopoietic stem cells, which eventually dieby apoptosis. Due to this auto-immune mechanism, a quantitative and qualitative defect of stem cells is typically detectable. However, very recently, findings that stem cells of Aplastic Anemia patients may contain short telomeres, telomerase-genes complex mutations, or mutations in specific key genes of DNA epigenetic regulation system, have suggested that the functional damage of bone marrow does not derive only from an extrinsic immune-mediated mechanisms, but also form an intrinsic one, arising from a primitive genetic deficiency of stem cell compartment. The pathogenesis of immune-mediated Aplastic Anemia realizes high response rate that patients show after immunosuppressive treatment. Over the years several treatment regimens have been experimented, but the highest rate of responses is inevitably associated with combinative use of anti-lymphocyte serum (preferentially horse ATG) and cyclosporine. Alternative regimens, including Cyclophosphamide or Alemtuzumab remain limited to clinical trials. However, the majority of patients cannot be defined completely cured with immunosuppression, for consistent rate of incomplete responses or relapses, and for the risk of clonal progression, present even in responding patients. For these reasons, probably related to the primitive intrinsic stem cell defect aforementioned, hematopoietic stem cell transplantation is currently the only chance of long-term cure in these patients. In this study we investigated the clinical efficacy and immunological reconstitution dynamics of a reduced intensity conditioning regimen including Fludarabine, Cyclophosphamide, and Alemtuzumab (namely, FCC conditioning regimen) in the treatment of Aplastic Anemia patients. The results of our study clearly confirm excellent clinical results of the FCC-conditioning, for the reduced incidence of graft rejection and GVHD, and prolonged overall and event-free survival. Moreover, our result show clearly a peculiar immune reconstitution, where global post-transplant lymphocytopenia is associated with 1) central re-education of CD4+ T-lymphocytes, 2) virus-mediated expansion of CD8+ T-lymphocytes, 3) presence of cells with a regulatory phenotype (T-regs and B-regs), and 4) establishment of a stable mixed chimerismstatus, that all together institute post-transplant tolerance, refractory in triggering potentially harmful immunological phenomena like GvHD