Hematopoietic Stem Cell Transplantation Protocols Research Articles

Characterization and treatment of chronic active Epstein-Barr virus disease: a 28-year experience in the United States

AbstractChronic active EBV disease (CAEBV) is a lymphoproliferative disorder characterized by markedly elevated levels of antibody to EBV or EBV DNA in the blood and EBV RNA or protein in lymphocytes in tissues. We present our experience with CAEBV during the last 28 years, including the first 8 cases treated with hematopoietic stem cell transplantation in the United States. Most cases of CAEBV have been reported from Japan. Unlike CAEBV in Japan, where EBV is nearly always found in T or natural killer (NK) cells in tissues, EBV was usually detected in B cells in tissues from our patients. Most patients presented with lymphadenopathy and splenomegaly; fever, hepatitis, and pancytopenia were common. Most patients died of infection or progressive lymphoproliferation. Unlike cases reported from Japan, our patients often showed a progressive loss of B cells and hypogammaglobulinemia. Although patients with CAEBV from Japan have normal or increased numbers of NK cells, many of our patients had reduced NK-cell numbers. Although immunosuppressive agents, rituximab, autologous cytotoxic T cells, or cytotoxic chemotherapy often resulted in short-term remissions, they were not curative. Hematopoietic stem cell transplantation was often curative for CAEBV, even in patients with active lymphoproliferative disease that was unresponsive to chemotherapy. These studies are registered at http://www.clinicaltrials.gov as NCT00032513 for CAEBV, NCT00062868 and NCT00058812 for EBV-specific T-cell studies, and NCT00578539 for the hematopoietic stem cell transplantation protocol.

Primary ovarian dysfunction after hematopoietic stem cell transplantation during childhood: busulfan-based conditioning is a major concern

We evaluated the incidence of patient/treatment factors associated with primary ovarian failure (POF) after hematopoietic stem cell transplantation (HSCT) during childhood. Fifty girls over 12 years of age (15.0 +/- 2.2) who were referred to the pediatric endocrinology clinic between March 2002 and March 2010 after HSCT at the Catholic HSCT center were enrolled in the study. In total, 36 (72%) out of 50 patients developed POF. Twenty-three patients were diagnosed and treated as chronic graft-versus-host disease. As preparative regimens for HSCT, 23 patients received total body irradiation (TBI)-based regimen, 19 received busulfan (BU)-based regimen, 4 received both BU- and TBI-based, and 4 received reduced intensity conditioning regimen. In a univariate logistic regression analysis, the BU-based regimen (p = 0.028) showed a strong relationship with POF. The incidence of POF according to the route of BU administration, between orally and intravenously, were not different (p = 0.435). These results emphasize the importance of monitoring these patients at regular intervals and the need to develop complementary HSCT protocols for preventing POF in children.

Acquisition of G 0 state by CD34-positive cord blood cells after bone marrow transplantation

Hematopoietic stem cells are kept in a quiescent state in the hypoxic area of the bone marrow, which is essential for hematopoietic stem cell maintenance. However, when and how hematopoietic stem cells acquire their hypoxic state and maintain quiescence has not been fully elucidated. The aim of this study was to understand this process in human hematopoietic stem cells after bone marrow transplantation. Human CD34-positive cord blood cells were transplanted into nonobese diabetic/severe combined immunodeficient interleukin-2 receptor γ chain knockout mice. Cell cycle and hypoxia assay analyses were performed, to identify changes in the characteristics of human hematopoietic stem cells following transplantation. Quantitative real-time reverse transcription polymerase chain reaction analysis was used to analyze the transcriptional changes accompanying this transition. Engrafted primitive lineage-negative CD34-positive CD38-negative cells acquired hypoxic state and quiescence in the recipient bone marrow between 4 and 8 weeks, and between 8 and 12 weeks after transplantation, respectively. During 4 and 8 weeks after transplantation, changes in the transcription levels of G₀ regulatory factors, such as CCNC and RBL1, and stem cell regulators, such as Flt3, were also seen, which may be related to the characteristic changes in the cell cycle or oxygenation state. Behavioral changes of hematopoietic stem cells in their cell cycle and oxygenation state during and after bone marrow engraftment may provide insights into hematopoietic stem cell regulation, mediating the improvement of clinical hematopoietic stem cell transplantation protocols and the eradication of leukemia stem cells.

Hematopoietic stem cell transplantation in multiple sclerosis: a review of the clinical experience and a report of an international meeting

Hematopoietic stem cell transplantation (HSCT), both allogeneic and autologous, has become one of the hottest topics in clinical immunology. One of the main autoimmune diseases in which HSCT has been extensively tried during the last decade is multiple sclerosis (MS). A few original papers and many anecdotal reports have indicated a beneficial effect of this treatment in MS, leading to stabilization or improvement in a large proportion of the treated patients. However, although hundreds of MS patients have been treated with HSCT, different conditioning and treatment protocols have been used in each center, making it difficult to organize and summarize the results from all of these small studies. Moreover, there is currently no completed controlled study with HSCT in MS. In this review, the cumulative experiences from several centers and countries in the world are summarized, based on the data presented at a recent international meeting in Moscow, Russia, entitled ‘Stem Cell Transplantation in Multiple Sclerosis: Sharing the Experience’.

Strong Association between Respiratory Viral Infection Early after Hematopoietic Stem Cell Transplantation and the Development of Life-Threatening Acute and Chronic Alloimmune Lung Syndromes

Alloimmune lung syndromes (allo-LS), including idiopathic pneumonia syndrome, bronchiolitis obliterans syndrome, and bronchiolitis obliterans organizing pneumonia, are severe complications after hematopoietic stem cell transplantation (HSCT). In our cohort of 110 pediatric patients, 30 had allo-LS (27.3%), 18 with ideopathic pneumonia syndrome and 12 with bronchiolitis obliterans syndrome. Multivariate analysis showed that respiratory viral infection early after HSCT is an important predictor for the development of allo-LS (P <.0001). This was true for all viruses tested. In multivariate analysis, allo-LS was the only predictor for higher mortality (P = .04). Paradoxically, prolonged administration of immunosuppressive agents because of acute graft-versus-host disease had a protective effect on the development of allo-LS (P = .004). We hypothesize that early infection of the respiratory tract with a common cold virus makes the lungs a target for alloimmunity.

Post-Transplant Cyclophosphamide and Sirolimus Are Synergistic in Preventing Rejection and Inducing Stable Mixed Chimerism Independently of Regulatory T Cells.

Abstract 3540Poster Board III-477A strategy that can induce stable mixed chimerism across human leukocyte antigen (HLA) barriers would be beneficial in extending the application of hematopoietic stem cell transplantation (HSCT) to patients with severe sickle cell disease (SCD) who are in need of this potentially curative procedure. Indeed, we have recently demonstrated the feasibility of an HLA-matched sibling protocol employing low dose total body irradiation (TBI, 300cGy), the lymphocyte depleting agent alemtuzumab, and sirolimus to reverse the phenotype with minimal side effects. Due to the lack of HLA-matched siblings in the majority of patients, our goal is to develop a safe haploidentical regimen. In this work, we focused on determining optimal postgrafting immunosuppression and examined sirolimus and post-transplant cyclophosphamide (PT-cy), agents known to induce transplantation tolerance. To determine the optimal sequence for combining these drugs and whether this combination is synergistic in promoting stable donor chimerism despite the antiproliferative effects of sirolimus, we used a mismatched murine model with BalbC donors and C57Bl6 recipients. Twenty-five to 40 recipient mice received 200cGy TBI and PT-cy (200mg/kg intraperitoneally (IP) 2 days post transplant) with or without sirolimus (3mg/kg IP) for 14 to 30 days starting 1 day before or 4, 6, or 10 days post transplant. We found that in contrast to sirolimus or PT-cy alone, the combination of PT-cy and a limited course of sirolimus resulted in stable mixed chimerism: all mice that received PT-cy and sirolimus starting between 1 day before and 6 days after transplant attained donor chimerism levels ranging from 15-35%. Further, a 14 day course of sirolimus was sufficient to maintain stable mixed chimerism in our model (See Figures 1 and 2). To examine whether this synergistic effect is mediated by regulatory T cells, we administered anti-CD25 monoclonal antibody (CD25 mAb), an agent known to transiently deplete these cells in vivo. Fifteen mice received 200cGy TBI, sirolimus, PT-Cy, and either no CD25 mab, CD25 mab (1mg IP) on 7 and 3 days before and 1 day after transplant, or CD25 mab starting 14 days after transplant. CD25 mab was given biweekly for 5 weeks to mice in both groups. Donor engraftment levels did not differ in the three groups, with donor chimerism levels ranging from 30-40%. Our data show that the anti-proliferative effects of sirolimus do not inhibit the efficacy of the cytotoxic agent cyclophosphamide. Rather, our data demonstrate that the combination of PT-cy and a limited course of sirolimus synergistically promote mixed bone marrow chimerism in a complete mismatched setting. Further, the synergistic effect of this drug combination appears to be mediated independently from CD25+ regulatory T cell expression. In light of our previous success using sirolimus in an HLA-matched HSCT protocol, these findings lay the groundwork for developing PT-cy and sirolimus as a novel, safe, and effective means of promoting stable mixed chimerism in the haploidentical setting and thus greatly enhancing our ability to successfully apply this approach to patients with severe SCD. [Display omitted] [Display omitted] Disclosures:No relevant conflicts of interest to declare.

Key factors in experimental mouse hematopoietic stem cell transplantation

The first mouse model of hematopoietic stem cell transplantation (HSCT) was developed more than 50 years ago. HSCT is currently being widely used in a broad range of research areas, which include studies of the engraftment process, the pathogenesis of graft-versus-host disease and possible ways of its treatment and prophylaxis, attempts to use the graft-versus-leukemia/tumor effect in treating hematological and oncological malignancies, cancer vaccine development, induction of transplanted organ tolerance, and gene therapy. However, although this model is widely distributed, many laboratories use different protocols for the procedure. There are a number of papers discussing different HSCT protocols in clinical work, but no articles summarizing mouse laboratory models are available. This review attempts to bring together different details about HSCT in the mouse model, such as the types of transplantation, possible pretreatment regimens and their combinations, methods and sources of graft harvesting and preparation for the transplantation procedure, the influence of graft cell dose and content on the engraftment process, the transplantation method itself, possible complications, symptoms and techniques of their prophylaxis or treatment, as well as follow-up and engraftment assessment. We have also tried to reflect current knowledge of the biology of the engraftment.

Characterization of a lineage-negative stem-progenitor cell population optimized for ex vivo expansion and enriched for LTC-IC.

Current hematopoietic stem cell transplantation protocols rely heavily upon CD34+ cells to estimate hematopoietic stem and progenitor cell (HSPC) yield. We and others previously reported CD133+ cells to represent a more primitive cell population than their CD34+ counterparts. However, both CD34+ and CD133+ cells still encompass cells at various stages of maturation, possibly impairing long-term marrow engraftment. Recent studies demonstrated that cells lacking CD34 and hematopoietic lineage markers have the potential of reconstituting long-term in vivo hematopoiesis. We report here an optimized, rapid negative-isolation method that depletes umbilical cord blood (UCB) mononucleated cells (MNC) from cells expressing hematopoietic markers (CD45, glycophorin-A, CD38, CD7, CD33, CD56, CD16, CD3, and CD2) and isolates a discrete lineage-negative (Lin-) cell population (0.10% +/- 0.02% MNC, n=12). This primitive Lin- cell population encompassed CD34+/- and CD133+/- HSPC and was also enriched for surface markers involved in HSPC migration, adhesion, and homing to the bone marrow (CD164, CD162, and CXCR4). Moreover, our depletion method resulted in Lin- cells being highly enriched for long-term culture-initiating cells when compared with both CD133+ cells and MNC. Furthermore, over 8 weeks in liquid culture stimulated by a cytokine cocktail optimized for HSPC expansion, TPOFLK (thrombopoietin 10 ng/ml, Flt3 ligand 50 ng/ml, c-Kit ligand 20 ng/ml) Lin- cells underwent slow proliferation but maintained/expanded more primitive HSPC than CD133+ cells. Therefore, our Lin- stem cell offers a promising alternative to current HSPC selection methods. Additionally, this work provides an optimized and well-characterized cell population for expansion of UCB for a wider therapeutic potential, including adult stem cell transplantation.