HLA-matched Bone Marrow Transplantation Research Articles

Treatment of Hemophagocytic Lymphohistiocytosis With Chemotherapy and Bone Marrow Transplantation: A Single-Center Study of 22 Cases

Abstract Twenty-two children with hemophagocytic lymphohistiocytosis were treated with a chemotherapy regimen consisting of VP16–213, corticosteroids, and intrathecal methotrexate. A sustained clinical and biologic complete remission was obtained in 15 children and a partial remission in one child; six children died early of opportunistic infection (n = 4) or of disease progression (n = 2). Of the 16 children who were placed in first remission, 10 received maintenance chemotherapy alone, while six underwent bone marrow transplantation (HLA matched in five, HLA mismatched in one). Of the children who received chemotherapy alone, only two are in long-term remission after cessation of treatment. The remaining eight patients relapsed after a mean period of 5.4 months (range 2 to 8 months). Further treatment using the same regimen induced second remissions of short duration; death occurred after a median period of 2.3 months (range 0.5 to 6 months). A total of nine patients received allogeneic bone marrow transplantation (BMT). Among the six children transplanted in remission, four are in long-term unmaintained remission, 1 to 6 years after HLA-matched BMT. However, the relapse that occurred in one patient 1 year post BMT is difficult to interpret because the donor, the patient's 5-year-old sister, also developed the disease 1 year later. An HLA-nonidentical BMT resulted in unmaintained remission for 1 year, with autologous hematologic reconstitution followed by disease relapse. HLA-nonidentical BMT failed in three other patients with active disease at time of transplant. The poor long-term results of chemotherapy alone justify the use of related HLA-matched BMT in complete remission.

Graft-Versus-Host Disease in Experimental Allogeneic Bone Marrow Transplantation

Considerable progress has been made in defining the relative contributions of CD4+ and CD8+ cells to GVHD. Studies in mice have shown that, in isolation, each T cell subset is able to induce lethal GVHD in irradiated hosts. For hosts differing at the MHC (H-2), CD4+ cells cause GVHD directed to H-2 class II antigens whereas CD8+ cells produce GVHD to H-2 class I antigens. With H-2-matched hosts expressing multiple minor H antigens, induction of lethal GVHD is largely under the control of CD8+ cells. Which particular minor H antigens provide the targets for GVHD in mice is still unclear. Clarifying this question is complicated by the finding that the target antigens for GVHD do not necessarily correlate with the targets for cytotoxicity measured in vitro; moreover, immunodominance occurs when T cells are exposed to multiple minor H antigens in vivo. In terms of clinical application, there is a need to devise animal models for improving the success of HLA-matched bone marrow transplantation. Selectively depleting the marrow inoculum of CD8+ cells and preimmunizing the donor against viral pathogens are two procedures which are currently under study.

"Minisatellite" DNA probes detect engraftment and/or chimerism in recipients of HLA-matched bone marrow transplants.

A study was carried out to determine whether the minisatellite DNA probes described by Jeffreys and coworkers could be used routinely to analyze engraftment, hematopoietic chimerism, and relapse in recipients of bone marrow transplants. The probes were informative for all of the recipient/donor pairs analyzed. Their limit of sensitivity was determined in reconstruction experiments and was found to vary from 2%, in the best cases, to 10%. We were able to confirm that engraftment and hematopoietic chimerism can, indeed, be sought routinely using this simple molecular approach. Only one set of probes is required for all patients and, unlike cytogenetic analysis, this analysis can be used whether or not blood or marrow cells are dividing.

Immunosuppressive Therapy Versus Bone Marrow Transplantation for Children With Aplastic Anemia

A total of 15 patients 1 to 16 years of age were treated for aplastic anemia (13 of a severe degree) and followed-up for a mean of 24 months (range 2 to 64 months). Six patients had an HLA-matched sibling and underwent allogeneic bone marrow transplantation. Nine patients who lacked a suitable donor were given immunosuppressive therapy. Antithymocyte globulin was the initial treatment for eight of these nine patients. Two patients who failed to respond to antithymocyte globulin were then treated with cyclosporine A. Pretreatment age, hematologic measurements, duration of follow-up, and interval prior to therapy were similar between the two groups. All of the patients receiving bone marrow transplants had a complete response and now have normal blood cell counts. Six of nine patients (67%) responded to antithymocyte globulin and are now transfusion free, although three have mild thrombocytopenia. Both patients given cyclosporine A had a good response and are also transfusion free. Patients who underwent marrow transplantation had a significantly shorter period of transfusion dependence for RBCs (9 v 4 weeks, P less than .005) and platelets (5 v 21 weeks, P less than .05). The patients given immunosuppressive therapy have significantly lesser platelet counts in follow-up but have similar values for both hemoglobin and absolute granulocyte counts. Although HLA-matched bone marrow transplantation leads to a faster and more complete recovery for children with aplastic anemia, immunosuppressive therapy can provide a good outcome for children with this disorder.

"Minisatellite" DNA probes detect engraftment and/or chimerism in recipients of HLA-matched bone marrow transplants

"Minisatellite" DNA probes detect engraftment and/or chimerism in recipients of HLA-matched bone marrow transplants

Partial Chimerism After T-Cell-Depleted Allogeneic Bone Marrow Transplantation in Leukemic HLA-Matched Patients: A Cytogenetic Documentation

We evaluated serially by cytogenetics the blood and marrow chimerism of 38 leukemic recipients of HLA-matched bone marrow transplants (BMT) who were prepared by high doses of alkylating agents and fractionated total-body irradiation (2.2 Gy X 5). Donor or host mitoses were identified by examination of sex chromosomes in 32 patients or by evaluation of the polymorphism of other chromosomes after specific banding in six patients. Twenty-four patients were recipients of untreated BMT, and 14 were recipients of T-cell-depleted BMT. In the 24 patients who received untreated BMT, all showed successful engraftment, and only three had a transient mixed chimera. In the 14 recipients of T-cell-depleted BMTs, four rejected their grafts, and seven had mixed chimeras; these mixed chimeras were more frequent in blood lymphocytes than in marrow cells and could be detected up to 26 months after BMT. This high frequency of partial chimerism after T-cell-depleted BMT by comparison with a control group suggests that the donor's T cells play an important role in the eradication of host residual hematopoiesis after allogeneic BMT.

Partial chimerism after T-cell-depleted allogeneic bone marrow transplantation in leukemic HLA-matched patients: a cytogenetic documentation

We evaluated serially by cytogenetics the blood and marrow chimerism of 38 leukemic recipients of HLA-matched bone marrow transplants (BMT) who were prepared by high doses of alkylating agents and fractionated total- body irradiation (2.2 Gy X 5). Donor or host mitoses were identified by examination of sex chromosomes in 32 patients or by evaluation of the polymorphism of other chromosomes after specific banding in six patients. Twenty-four patients were recipients of untreated BMT, and 14 were recipients of T-cell-depleted BMT. In the 24 patients who received untreated BMT, all showed successful engraftment, and only three had a transient mixed chimera. In the 14 recipients of T-cell-depleted BMTs, four rejected their grafts, and seven had mixed chimeras; these mixed chimeras were more frequent in blood lymphocytes than in marrow cells and could be detected up to 26 months after BMT. This high frequency of partial chimerism after T-cell-depleted BMT by comparison with a control group suggests that the donor's T cells play an important role in the eradication of host residual hematopoiesis after allogeneic BMT.

IMPACT OF T-CELL DEPLETION ON OUTCOME OF ALLOGENEIC BONE-MARROW TRANSPLANTATION FOR STANDARD-RISK LEUKAEMIAS

71 leukaemic patients having HLA-matched bone-marrow transplants (BMT) were randomised to receive whole marrow (group A) or marrow depleted of T cells by treatment with monoclonal antibodies (anti CD4-CD5-CD8, group B; anti CD2-CD5-CD7, group C) plus complement. All patients received cyclophosphamide and total body irradiation before transplantation and cyclosporin after BMT. Marrow treatment removed 97% of T cells (median) in group B and 99% in group C. Although both serious and mild graft-versus-host disease (GVHD) were reduced in T-cell depleted patients, graft failure and relapse were increased. Graft failure was caused by GVHD and transplant complications in the controls and by rejection and relapse in the T-cell depleted groups; relapse-free survival did not differ between the groups. Without better control of host immunity and of the residual leukaemia T-cell depletion of the marrow, BMT should not be pursued in standard-risk patients.

FAVORABLE PROGNOSIS FOR CHILDREN WITH APLASTIC ANEMIA TREATED WITH IMMUNOSUPPRESSIVE THERAPY

Survival rates for children with aplastic anemia (AA) who do not receive HLA-matched bone marrow transplantation (BMT) are less than 50%. In the past 5 years we have treated 9 patients (pts) with AA with immunosuppressive therapy (ImmRx); 8 pts are alive and do not require transfusion (Tx) with a median follow-up of 14.5 months (range 5-62). Seven pts had severe AA and 2 had moderate AA. AA was associated with hepatitis in 2 pts, benzene in 1 pt, arthritis in 1 pt and was idiopathic in 5 pts. All pts received anti-thymocyte globulin (ATG) 15 mg/kg/day for 14 consecutive days followed by 7 additional doses over the next 14 days. All pts received methylprednisolone or its equivalent at 1-2 mg/kg/day during the ATG to combat serum sickness. Two pts also received 1 haploidentical bone marrow infusion and oral androgens, 1 additional pt received oral androgens. Six pts (5 severe) had a good response to ATG and are Tx free. In one pt, persistent thrombocytopenia responded to cimetidine therapy. Of the 3 pts who failed ATG, 2 are Tx free after treatment with cyclosportne A. During the same time, 6 pts with severe AA had BMT with a complete response. The response to BMT was significantly faster than to ATG. Red cell Tx was required for 34 days post BMT vs. 66 days post ATG (p<0.05). In conclusion, although BMT is better than ImmRx in children with AA, when a suitable donor is unavailable, ImmRx may result in a favorable outcome.

Prevention of graft-versus-host disease in HLA-matched bone marrow transplantation for malignant diseases: a multicentric study of 62 patients using 3-pan-T monoclonal antibodies and rabbit complement.

In order to evaluate the effectiveness and reproducibility of T cell depletion in human leukocyte antigen (HLA)-matched bone marrow graft to prevent graft-v-host disease (GVHD), our multicentric study (nine different centers) investigated 62 consecutive patients with poor prognosis leukemia or hematosarcoma from June 1984 to November 1985. The data were updated October 1, 1986, and the mean follow-up was 18 +/- 4.3 months. T cells were depleted with a combination of 3-pan-T cell monoclonal antibodies (CD2 "D66"; CD5 "A50"; CD7 "I21") with a single incubation of rabbit complement (C'). The average number of T cells infused was 0.66 X 10(6) +/- 0.56/kg body weight. Twenty-six patients received chemoprophylaxis for GVHD, 16 received methotrexate, and ten received cyclosporin A. Only a single case of severe (greater than grade II) GVHD was observed, yet the incidence of graft failure was 19%. Factors that might have influenced the occurrence of graft failure appear to be the lack of radiotherapy in the conditioning regimen; the conditioning regimen itself (fractionated total body irradiation [TBI], 12 Gy, v single dose is better than TBI, 10 Gy, but still not statistically significant); and the age of the patients (high-risk after 30 years of age). In contrast, neither the number of nucleated cells reinfused nor the level of T cell depletion (provided the T cells were below critical numbers) seemed to have an influence, nor did chemoprophylaxis for GVHD or splenectomy in chronic granulocytic leukemia (CGL) patients. The survival of graft failure patients was very poor (one of 11; survival at 15 months of the initial graft). Thus, our study demonstrates the reproducibility and high effectiveness in preventing GVHD by immunodepletion of T cells in a large-scale multicentric assay, in which compliance with the protocol of immunodepletion was reasonably good. This study thus provides interesting clues to overcoming graft rejection.

Standardization of T-cell depletion in HLA matched bone marrow transplantation.

The IBM 2991 Blood Cell Processor has been used to isolate a mononuclear cell (MNC) fraction from the marrow of 31 allogeneic donors. The MNC fraction was then incubated with a combination of two murine monoclonal antibodies MBG6 (CD6) and RFT8 (CD8) followed by two rounds of treatment with rabbit complement resulting in a marrow inoculum significantly reduced in the number of T-lymphocytes. We report here new specifications for the use of Ficoll-Metrizoate, the method used to calculate T-lymphocyte depletion and the details of our attempts to improve T-depletion. Following marrow transplantation with this T-depleted fraction, 29 patients are evaluable for engraftment, one patient failed to engraft and one died too early for evaluation. Twenty-two had no acute graft-versus-host disease (aGvHD), at a minimum of 60 d, six had grade I acute GvHD and one grade III. No correlation was found between the absolute number of MNC infused and time to engraftment, nor any relationship between the number of residual viable T-lymphocytes in the infused marrow and the incidence of GvHD, but the patient with the most severe aGvHD also had the highest number of T-lymphocytes infused.

EPSTEIN-BARR VIRUS INFECTION AND IMMUNITY IN BONE MARROW TRANSPLANT RECIPIENTS

Studies on patients for up to one year following allogeneic, HLA-matched bone marrow transplants have shown no increased incidence of salivary Epstein-Barr (EB) virus secretion and no significant rise in EB-virus-specific antibody titers. EB-virus-specific cytotoxic T cells could be detected in the peripheral blood of all patients by six months posttransplant. For up to one year posttransplantation in vitro EB virus infection of peripheral blood B lymphocytes from the majority of patients leads to an abortive infection followed by cell death, and without the establishment of continuously growing cell lines. This abnormality appeared to be due to patients' monocytes, which formed a defective feeder cell layer in culture, and it could be circumvented by culturing the EB-virus-infected B cells from patients on a feeder layer of x-irradiated adherent cells from normal peripheral blood. These findings may explain the relative lack of EB-virus-associated lymphoma seen in bone marrow transplant recipients when compared with other groups of transplant patients.

Graft rejection following HLA matched T-lymphocyte depleted bone marrow transplantation.

Bone marrow graft rejection following HLA-matched bone marrow transplantation (BMT) for leukaemia has been a rare problem. However, with the introduction of T-lymphocyte depleted BMT, graft rejection is recognized as a new complication. At the Royal Free Hospital (RFH) in London T-depletion is achieved using two monoclonal antibodies with complement mediated lysis. The methodology was extended to other centres and in total 56 patients have received T-depleted, HLA matched BMT. Twelve of 56 patients have had graft rejection. At the RFH three of 41 (7%) patients have had rejection whereas at collaborating centres nine of 15 (60%) patients have had rejection. We have investigated these rejections in order to identify factor(s) responsible. Rejection was not restricted by patient or donor characteristics, nor disease status. Patient management, chemotherapy conditioning, efficiency of T-depletion, graft versus host disease (GvHD), and infection post BMT, were not consistently implicated. The major difference between the RFH and all other centres was in the radiotherapy (RT) conditioning: The RFH prescribed a single fraction of 7.5 Gy total body irradiation (TBI) whilst collaborating centres gave 10 or 12 Gy fractionated TBI. We conclude that the different incidence of rejection (7% v. 60%) relates primarily to the RT conditioning although the mechanisms(s) of rejection remain unknown. We conclude that where T-depleted BMT is used, compensation by more intensive RT conditioning is required in order to avert graft rejection.

BONE-MARROW TRANSPLANTATION FOR INBORN ERROR OF PHAGOCYTIC CELLS ASSOCIATED WITH DEFECTIVE ADHERENCE, CHEMOTAXIS, AND OXIDATIVE RESPONSE DURING OPSONISED PARTICLE PHAGOCYTOSIS

Two girls had delayed umbilical cord detachment, recurrent bacterial infection, inability to form pus, and marked leucocytosis. Their phagocytes were defective in tests of adherence, random migration, chemotaxis, and oxidative burst. NK activity was virtually absent. This rare disorder, due to an inherited absence of a 180 kilodalton membrane glycoprotein on polymorphonuclear cells, is usually lethal within 2 years. Allogeneic HLA-matched bone-marrow transplantation done at ages 4 months in one patient and 2 years in the other after intensive conditioning was successful and resulted in nearly complete correction of phagocytic cell function and NK activity within two months. One patient died 9 months after transplantation from severe chronic graft-versus-host disease with obstructive bronchopneumopathy. The other is doing well 1 year after transplantation and showing stable chimerism and normal phagocytic function.

H-2-restriction of T cells mediating lethal graft-versus-host-disease to minor histocompatibility determinants.

Although the etiology and pathogenesis of graft-versus-host disease (GVHD) continues to excite controversy and speculation, it is now generally accepted that the disease reflects alloaggres-sion mediated by mature post-thymic T cells (1–3). The pioneering studies of Siraonsen (1) established that the most florid forms of GVHD occur when the donor and host differ at the major histocompatibility complex (MHC). In recent years, however, it has become clear that severe and often fatal forms of GVHD occur in MHC-com-patible combinations, e.g. following HLA-matched bone marrow transplantation in man (3). Although the etiology of this form of GVHD is still debated, our own studies in mice suggest that responses to minor histocompatibility antigens (minor HA)4 are the principal cause of the disease, at least in this species (4).

Autologous Hematologic Recovery from Aplastic Anemia following High Dose Cyclophosphamide and HLA-Matched Allogeneic Bone Marrow Transplantation

A 17-year-old woman with severe aplastic anemia was treated with high-dose cyclophosphamide followed by infusion of bone marrow cells from her HLA-identical, ABO-incompatible brother. The marrow graft failed to take. Subsequently, the patient revealed an autologous marrow reconstitution leading to a near-complete hematologic recovery which is now persisting for over 20 months.