Successful allogeneic bone marrow retransplantation with the same donor after graft rejection: Application of a modified conditioning regimen

Summary A 26-year-old man with severe aplastic anemia was treated with high-dose Cyclophosphamide followed by the infusion of bone marrow cells from his HLA-identical sister. After initial take of the graft, rejection ensued by day 46 which was followed by a permanent complete aplasia. After 4 months, bone marrow retransplantation with the same donor was attempted after a more intensive conditioning regimen. This led to permanent engraftment with rapid normalization of the blood counts lasting now for over 12 months. The patient has since remained in excellent clinical condition without signs of graft-versus-host disease.     

HHV-6-related secondary graft failure following allogeneic bone marrow transplantation

We report the case of an 11-year-old boy who underwent allogeneic bone marrow transplantation (BMT) for relapsed acute lymphoblastic leukaemia. Despite adequate engraftment, on day 45 he developed marrow aplasia with haemophagocytosis. HHV-6 was detected in blood and bone marrow by nested PCR. Retrospective testing showed that viraemia had started on day 24. Following therapy with foscarnet and ganciclovir, viral load declined to undetectable levels and his donor marrow recovered contemporaneously. This case suggests that HHV-6 may be a treatable cause of graft failure following BMT and provides clinical and virological evidence for the anti-HHV-6 activity of ganciclovir and foscarnet.     

Recombinant human G-CSF-mobilized peripheral blood stem cells for second allogeneic transplant after bone marrow graft rejection in children

Two children affected by severe aplastic anaemia and sickle cell anaemia rejected the allogeneic bone marrow transplantation from an HLA-matched unrelated volunteer and an HLA-identical sibling, respectively. In both cases a second transplant using granulocyte-colony stimulating factor (G-CSF) mobilized peripheral blood stem cells (PBSC) was performed. Donors were the HLA-haploidentical mother and the same HLA-identical sibling who was employed for the first marrow allograft, respectively. Treatment with G-CSF and PBSC collection were well tolerated. Both patients had engraftment of donor haemopoiesis and did not experience severe graft-versus-host disease. These cases confirm that PBSC transplant should be considered as a feasible treatment to reverse graft failure in paediatric patients.     

Late graft rejection and second infusion of bone marrow in children with aplastic anaemia

Late graft rejection following allogeneic bone marrow transplantation (BMT) for aplastic anaemia is a significant clinical problem and is associated with a high risk of mortality. We report two children with severe aplastic anaemia (SAA) who developed very late graft rejection 2 years and 4 months and 10 years respectively after allogeneic BMT from HLA-identical siblings. Following a second BMT from their initial donors, engraftment has been sustained in both cases. The patients are alive with full donor chimaerism, 18 and 19 years from initial transplant. These cases illustrate that graft failure can be an extremely late event after allogeneic BMT for SAA, and that long-term sustained engraftment can be achieved in these patients with second BMT from the original donors.     

Predominant or complete recipient T-cell chimerism following alemtuzumab-based allogeneic transplantation is reversed by donor lymphocytes and not associated with graft failure

The clinical significance of mixed chimerism following allogeneic haematopoietic stem cell transplantation (HSCT) remains controversial. Its relevance and incidence are probably influenced by the conditioning regimen and incorporation of T‐cell depletion. The presence of recipient chimerism levels >40–50% following T‐cell replete reduced intensity transplantation correlates with a high risk of graft rejection, regardless of donor‐lymphocyte infusions, but it is unclear whether this finding translates to T‐cell depleted transplants. We conducted a retrospective single‐institution analysis of patients receiving alemtuzumab‐based HSCT. 27/152 (18%) evaluable cases had predominantly recipient T‐cell chimerism at 3 months or beyond. By contrast, coincident chimerism in the granulocyte lineage was predominantly of donor origin (median 100%) in all but one patient. Donor lymphocyte infusion effectively converted predominantly recipient T‐cell chimerism to ful donor chimerism in all evaluable cases including three cases with no detectable donor T cells. The only graft failure occurred in the patient with predominantly recipient myeloid chimerism in whom rejection occurred rapidly before donor lymphocytes could be administered. We conclude that predominant or complete recipient T‐cell chimerism following alemtuzumab‐based regimens does not have the same clinical implications as that following T‐cell replete transplants and can be effectively converted with donor lymphocytes without the need for lympho‐depleting agents or re‐conditioning.     

Late infections after allogeneic bone marrow transplantations: comparison of incidence in related and unrelated donor transplant recipients

Infectious complications are a major cause of morbidity and mortality after allogeneic bone marrow transplantation (BMT). We have evaluated the incidence of late infections (beyond day +50) in recipients of related (RD) and unrelated donor (URD) allogeneic BMT, factors associated with increased risks of infection, and the impact of the late infections on survival. Between 1989 and 1991, 249 patients received an RD (n = 151) or URD (n = 98) allogeneic BMT at the University of Minnesota and all late infections were investigated. Three hundred sixty-seven late infectious events developed in 162 patients between 50 days and 2 years after BMT. The incidence of any late infection was greater in URD versus RD recipients (84.7% v 68.2%, respectively; P = .009). In multivariate analysis, advanced graft- versus-host disease (GVHD) was significantly associated with late infections. The effect of GVHD was apparent only in RD recipients (relative risk [RR], 2.29; P = .003), whereas URD recipients, with or without GVHD, had more late infections compared with RD recipients without GVHD. Multivariate analysis showed that late posttransplantation infections were the dominant independent factor associated with increased nonrelapse mortality (RR, 5.5; P = .0001), resulting in improved 3-year survival for RD versus URD recipients (49.9% +/- 8% v 34.4% +/- 10%; P = .004). In this study, we observed that late infections are more frequent in URD recipients, resulting in substantially higher nonrelapse mortality. This prolonged period of increased infectious risk in URD recipients suggests the need for aggressive surveillance and therapy of late infections and perhaps prolonged antibiotic prophylaxis for all URD BMT recipients.     

Analysis of T-cell repertoire and mixed chimaerism in a patient with aplastic anaemia after allogeneic bone marrow transplantation

We analysed 26 T-cell receptor (TCR) beta chain subfamilies (VB) of a patient with aplastic anaemia (AA) who underwent allogeneic bone marrow transplantation (allo-BMT). The patient developed pancytopenia at d 80. The patient's T cells were skewed in 10 of 26 TCR-VB on d 83. These TCR-VB, especially VB15, which were almost entirely CD8-positive cells, were skewed throughout her clinical course. Chimaerism analysis of the CD8-positive cells indicated that they were of recipient origin. Therefore, some immune responses induced by the recipient CD8-positive T cells had an important role in pancytopenia in AA patients after allo-BMT.     

Comparative analysis of clinical outcomes after allogeneic bone marrow transplantation versus peripheral blood stem cell transplantation from a related donor in Japanese patients

A reduced incidence of graft versus host disease (GvHD) has been documented among Japanese allogeneic bone marrow transplantation (BMT) patients, as the Japanese are genetically more homogeneous than western populations. To clarify whether this ethnic difference affects the results of allogeneic peripheral blood stem cell transplantation (PBSCT), we conducted a nationwide survey to compare clinical outcomes of allogeneic PBSCT (n = 214) and BMT (n = 295) from a human leucocyte antigen-identical-related donor in Japanese patients. The cumulative incidence of grades II-IV acute GvHD was 37.4% for PBSCT and 32.0% for BMT. The cumulative incidence of extensive chronic GvHD at 1 year was significantly higher after PBSCT than BMT (42% vs. 27%; P < 0.01). The organ involvement patterns of GvHD were different between the two groups. By multivariate analyses, the incidence of chronic GvHD was significantly increased in PBSCT, whereas the stem cell source did not affect the incidence of acute GvHD, transplant-related mortality, relapse or survival. We concluded that Japanese PBSCT patients have an increased risk of chronic GvHD compared with BMT patients, but the incidence of acute GvHD was still lower than in western populations. Thus, the choice of haematopoietic stem cell source should be considered based on data for individual ethnic populations.     

Transduction of donor hematopoietic stem-progenitor cells with Fas ligand enhanced short-term engraftment in a murine model of allogeneic bone marrow transplantation

Fas-mediated apoptosis is a major physiologic mechanism by which activated T cells are eliminated after antigen-stimulated clonal expansion generates a specific cellular immune response. Because activated T cells are the major effectors of allograft rejection, we hypothesized that genetically modifying allogeneic bone marrow (BM) cells prior to transplantation could provide some protection from host T-cell attack, thus enhancing donor cell engraftment in bone marrow transplantation (BMT). We undertook studies to determine the outcome of lentiviral vector-mediated transduction of Fas ligand (FasL) into lineage antigen-negative (lin(-)) mouse BM cells (lin(-) BMs), in an allogeneic BMT model. FasL-modified lin(-) BMs killed Fas-expressing T cells in vitro. Mice that received transplants of allogeneic FasL(+) lin(-) BMs had enhanced short-term engraftment, after nonmyeloablative conditioning, as compared to controls. We observed no major hepatic toxicity or hematopoietic or immune impairment in recipient mice at these time points. These results suggest potential therapeutic approaches by manipulating lymphohematopoietic stem-progenitor cells to express FasL or other immune-modulating genes in the context of BMT.     

Thymus-independent expansion of T lymphocytes in children after allogeneic bone marrow transplantation

The contribution of the thymus-dependent pathway and thymus-independent pathways for T cell regeneration after BMT in children is still unclear. We analyzed the kinetics of T cell regenerative pathways after allogeneic BMT. The number of CD4+CD45RA+ T cells, a thymus-dependent population, was very low until 3 months after BMT. The numbers of CD28- T cells and CD8+ T cells expressing CD8alpha/alpha homodimer (CD8alpha/alpha+ T cells), a thymus-independent population, increased shortly after BMT, beyond the levels of healthy children in some patients. The numbers of Vgamma9+Vdelta2+ and Valpha24+ T cells, which represent populations of extrathymic development, were less than 200/microl during the 6 months after BMT. There was a significant inverse correlation between the percentages of CD4+CD45RA+ and CD28-T cells at 1 month, and a positive correlation between the percentages of CD28- and CD8alpha/alpha+ T cells at 2 and 3 months after BMT. The mean age at BMT was higher in patients with a high level of CD8alpha/alpha+ T cells than in those without an increase in these cells, suggesting the influence of thymic function on the regenerative pathways. These results suggest that the thymus-independent pathway is the dominant source of T cells even in children shortly after allogeneic BMT.     

Role of biliary complications in chronic graft rejection after living donor liver transplantation

Postoperative biliary complications remain a substantial challenge after living donor liver transplantation, especially due to its heterogeneous clinical presentation.     

Influence of human leucocyte antigen disparity and graft lymphocytes on allogeneic engraftment and survival after umbilical cord blood transplant in adults

The dose of graft-nucleated cells and CD34(+) haematopoietic progenitor cells are predictors of allogeneic engraftment and survival in umbilical cord blood (UCB) recipients. In this single institution prospective phase II trial, flow cytometric analyses of CD34(+) progenitor and lymphocyte populations in unmodified single unit human leucocyte antigen (HLA)-disparate UCB grafts infused into 31 consecutive adults (median age 41 years, range 20-64) receiving myeloablative conditioning were compared with clinical outcomes. Median infused UCB graft-nucleated cells and CD34(+) dose was 2.2 x 10(7)/kg and 1.2 x 10(5)/kg respectively. Day to absolute neutrophil count >/=0.5 x 10(9)/l with full donor chimerism averaged 27 d (range 12-41). Univariate analyses demonstrated that UCB graft-infused cell doses of CD34(+) (P = 0.015), CD3(+) (P = 0.024) and CD34(+)HLADR(+)CD38(+) progenitors (P = 0.043) correlated with neutrophil engraftment. This same analysis did not demonstrate a correlation between CD34(+) (P = 0.11), CD3(+) (P = 0.28) or CD34(+)HLADR(+)CD38(+) (P = 0.108) cell dose and event-free survival (EFS). High-resolution matching for HLA-class II (DRB1) resulted in improved EFS (P = 0.02) and decreased risk for acute graft-versus-host disease (GVHD) (P = 0.004). Early mortality (prior to post-transplant day +28) occurred in three patients, while 26 patients achieved myeloid engraftment. These results suggest that UCB graft matching at DRB1 is an important risk factor for acute GVHD and survival, while higher UCB graft cell doses of CD34(+), committed CD34(+) progenitors and CD3(+) T cells favourably influence UCB allogeneic engraftment.     

Leukaemic relapse of donor origin after allogeneic bone marrow transplantation from a donor who later developed bronchogenic carcinoma

Donor-derived leukaemia is exceptional after allogeneic bone marrow transplantation (BMT). A woman with chronic myeloid leukaemia received an allogeneic BMT from a human leucocyte antigen-identical brother. The donor, a 50-year-old non-smoker, died of squamous cell bronchogenic carcinoma 1 year later. At 4 years post BMT, the patient became BCR/ABL positive and relapsed with acute myeloid leukaemia, which was shown to be donor-derived cytogenetically and molecularly. Retrospective analysis showed that the donor-leukaemic clone had started to evolve as early as 6 months post BMT. Sequencing of p53 ruled out Li-Fraumeni syndrome. Predisposition to malignancy might be an underlying mechanism of donor-cell leukaemia.     

An unbalanced monocyte macrophage polarization in the bone marrow microenvironment of patients with poor graft function after allogeneic haematopoietic stem cell transplantation

Poor graft function (PGF) is a severe complication of allogeneic haematopoietic stem cell transplantation (allo‐HSCT). Murine studies have demonstrated that effective haematopoiesis depends on the specific bone marrow (BM) microenvironment. Increasing evidence shows that BM macrophages (MФs), which constitute an important component of BM immune microenvironment, are indispensable for the regulation of haematopoietic stem cells (HSCs) in the BM. However, little is known about the number and function of BM MФs or whether they directly interact with HSCs in PGF patients. In the current prospective case‐control study, PGF patients showed a significant increase in classically activated inflammatory MФs (M1; 2·18 ± 0·11% vs. 0·82 ± 0·06%, P < 0·0001), a striking reduction in alternatively activated anti‐inflammatory MФs (M2; 3·02 ± 0·31% vs. 21·89 ± 0·90%, P < 0·0001), resulting in a markedly increased M1/M2 ratio (0·82 ± 0·06 vs. 0·06 ± 0·002; P < 0·0001) in the BM compared with good graft function patients. Meanwhile, standard monocyte subsets were altered in PGF patients. Dysfunctional BM MФs, which were characterized by reduced proliferation, migration and phagocytosis, were evident in PGF patients. Furthermore, BM MФs from PGF patients with high tumour necrosis factor‐α and interleukin 12 levels and low transforming growth factor‐β levels, led to impaired BM CD34⁺ cell function. In summary, our data indicate that an unbalanced BM M1/M2 ratio and dysfunctional MФs may contribute to the occurrence of PGF following allo‐HSCT.     

Eosinophilic folliculitis in a patient after allogeneic bone marrow transplantation: case report and review of the literature

We describe a patient with eosinophilic folliculitis (EF) that developed 3 months after allogeneic bone marrow transplantation (BMT) for chronic myelogenous leukemia. Skin biopsy specimen revealed numerous eosinophil infiltrations from the hair follicles to the sebaceous glands. No sign of skin graft-versus-host disease (GVHD) was found. The skin lesions and peripheral eosinophilia subsided within 5 weeks. In the following 8 months, the rash did not recur and GVHD has not developed. Taken together with previous reports, the present case demonstrates that it is important to recognize this distinct skin condition to avoid an inference that it may manifest itself as one of the signs of skin GVHD.     

Secondary eosinophilia in a patient with hypereosinophilic syndrome after allogeneic bone marrow transplantation from a sibling donor

A 23-year-old man first visited a local hospital in 1998 because of exertional dyspnea. Peripheral blood examination revealed mild leukocytosis with 82% eosinophils, and he was treated with prednisolone. As the eosinophilia did not improve, he was referred to Tokai University Hospital in March 1999 for further diagnosis and treatment. The patient was diagnosed as having hypereosinophilic syndrome (HES) because of unexplained hypereosinophilia persisting for more than 6 months, resulting in cardiac dysfunction. His disease was progressive in spite of immunosuppressive therapy, interferon-alpha and cytotoxic chemotherapy. Since he had an HLA-identical brother, allogeneic bone marrow transplantation (BMT) was performed in October 1999. After completion of the immunosuppressive therapy on day 79 after BMT, the number of eosinophils gradually increased again. Although we suspected recurrence of the disease, DNA fingerprinting revealed that the peripheral granulocytes were 100% donor type. An increase of interleukin-5 (IL-5) produced by peripheral lymphocytes and a decrease of the Th1/2 ratio suggested that the eosinophilia was related to GVHD. The eosinophilia was eventually controlled by cyclosporin. We conclude that DNA fingerprinting and examination of the IL-5 level and Th1/2 ratio are useful for differentiating between relapse and GVHD in cases of eosinophilia occurring after BMT for HES.     

Studies of allogeneic bone marrow and spleen cell transplantation in a murine model using ultraviolet-B light

Ultraviolet irradiation inhibits alloreactive and mitogen-induced responses and might reduce both graft-versus-host and host-versus-graft reactions after bone marrow transplantation (BMT). We have studied proliferative responses to mitogens and reactivity in mixed lymphocyte culture after irradiation with ultraviolet (UV)-B light using splenocytes from Balb/c (H-2d) and CBA (H-2k) mice. Response to mitogens and in MLC was strongly inhibited by 20 J/m2 and abolished at 50 J/m2. Clonogenic cell recovery (CFU-GM; CFU-S) after UV-B irradiation was also reduced. When bone marrow and spleen cells were transplanted from parent (Balb/c) animals into F1 hybrid (Balb/c X CBA) recipients, all animals died with features indicative of graft-versus- host disease (GVHD) in 34 days. If the grafts were first irradiated with 100 J/m2 of UV-B at a mean wavelength of 310 nm, then 76% survived to day 80 when they were killed and shown to have normal marrow cellularity. The remainder died in marrow aplasia or of GVHD. H-2 typing in a group of surviving recipients showed either donor hematopoiesis only (8 of 15), mixed allogeneic chimerism (5 of 15), or recipient type hematopoiesis (2 of 15). Higher doses (200 to 300 J/m2) were detrimental to survival with 88% of recipients dying in marrow aplasia. Syngeneic BMT in Balb/c mice showed slower hematopoietic reconstitution when the grafts were first irradiated with 100 J/m2. After BMT from Balb/c to CBA mice all recipients of unirradiated grafts died within 54 days. By contrast, after graft irradiation with 100 J/m2 survival of recipient animals to day 80 was 59%. If these grafts were treated with 50 J/m2 survival was only 26% with an increase in deaths due to GVHD. Hematopoiesis at day 80 in a group of survivors studied by Ig heavy chain allotyping indicated donor type hematopoiesis in 6 of 10 (50 J/m2) and 2 of 9 (100 J/m2). These data indicate that UV-B irradiation inhibits lymphocyte reactivity and can prevent GVHD. However, there is clear in vitro and in vivo evidence of stem cell damage, such that autologous marrow recovery was demonstrated in a proportion of recipients. In parent----F1 UV-irradiated transplants, sustained hematopoietic recovery was effected in the majority by donor stem cells.