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Chronic graft-v-host disease (GVHD) is the most frequent late complication of allogeneic bone marrow (BM) transplant. To test different treatments, we used a rat model of chronic GVHD which clinically, histologically, and immunologically resembles the disease occurring after human marrow transplant. The following treatments were administered: azathioprine 50 mg/kg/day plus prednisone 1 mg/kg/every other day; Cyclosporine (CsA) 30 mg/kg/day; CsA 30 mg/kg/day plus prednisone 1 mg/kg/every other day; thalidomide 50 mg/kg/day; thalidomide 10 mg/kg/day; CsA 10 mg/kg/day; and thalidomide 10 mg/kg/day plus CsA 10 mg/kg/day. All drugs were administered by gavage. Half of the animals (six of 12) treated with azathioprine plus prednisone every other day responded to treatment. The majority of animals (seven of 12) treated with CsA died early. The addition of prednisone every other day did not improve survival. The surviving animals treated with CsA (with or without prednisone) responded to treatment. Half of the animals treated with each of the above regimens had recurrent or progressive disease after therapy was discontinued. Thalidomide treatment was successful in the majority of animals (16 of 18). The addition of low-dose CsA to low-dose thalidomide resulted in a faster rate of response (12.1 +/- 1.8 v 28.2 +/- 1.6 days). We conclude that both thalidomide and thalidomide plus CsA appear promising in this model as treatment for chronic GVHD.

In an animal bone marrow transplant (BMT) model that mimics the human clinical condition, we evaluated the effectiveness of monoclonal antibody (MoAb) therapy in eliminating minimal residual disease (MRD) in a leukemic host. Leukemic rats were prepared with marrow ablative but noncurative doses of busulfan (BU) and cyclophosphamide (CY). Two days after syngeneic BMT, rats were treated with MoAb. Although all control rats died of leukemia relapse, 58% of those treated with MoAb were cured without any demonstrable effect on the rate of peripheral blood leukocyte recovery. Furthermore, the level of complement, an important effector in suppressing leukemia proliferation in the normal rat, was not adversely affected by BU/CY, BMT and MoAb. Thus, we demonstrated in an animal model that MoAb therapy may be a useful, nontoxic adjunct to high-dose chemotherapy and BMT in eliminating MRD.

The records of 549 bone marrow transplant (BMT) patients at The Johns Hopkins Oncology Center during a 9-year period were reviewed to determine the incidence of bronchiolitis obliterans (BrOb). Seven patients had BrOb. All seven died, and BrOb was a contributing cause of death in six patients. Only recipients of allogeneic BMT were at risk for developing BrOb (2% incidence). Three cases were incidentally discovered at autopsy in patients who died less than 120 days after BMT from ventilatory failure owing to interstitial pneumonitis. Four cases were patients who died greater than 120 days after BMT. Of this latter group, all had overt chronic graft-v-host disease (CGVHD). Among 120 day survivors of allogeneic BMT, 6% of those with CGVHD developed BrOb as compared with none of those without CGVHD (P = .008). Five percent of patients with reduced IgG levels at day 120 developed BrOb as compared with none of those with normal IgG (P = .04). The incidence of BrOb in 120-day survivors was 14% (4 of 29) in patients with both CGVHD and decreased serum IgG, whereas patients with CGVHD only (0 of 25), those with decreased IgG levels only (0 of 53), and those with no CGVHD and normal IgG levels (0 of 70) did not develop BrOb.

Markedly elevated serum IgE levels have been noted following allogeneic bone marrow transplantation (BMT) and have been correlated with graft-v- host disease (GVHD) in several studies. To investigate this phenomenon, we measured serum IgE levels in 387 allogeneic, 143 autologous, and 21 syngeneic BMT recipients before and at intervals after BMT. As a population, allogeneic BMT recipients displayed a biphasic elevation in IgE levels, with peak levels occurring either early (days 15 to 19) or late (days 80 to 89) posttransplant. Only in individuals in whom peak levels occurred early did IgE level correlate with liver disease, histological changes, and overall clinical stage of GVHD. The association of IgE elevation and GVHD does not appear to be direct since recipients of syngeneic (monozygotic twin) grafts had the highest incidence of IgE hyperresponsiveness as well as the highest absolute IgE levels. Similarly, 22 recipients of autologous marrow not treated with 4-hydroperoxycyclophosphamide had elevated IgE levels comparable to those seen in allogeneic graft recipients. We hypothesize that augmented IgE synthesis and its subsequent resolution is the natural consequence of immune reconstitution in the presence of potentially reaginic agents such as antibiotics and infectious agents. As such, IgE hyperresponsiveness in syngeneic graft recipients may reflect the maturational sequence of IgE regulatory elements in the absence of interference by GVHD, GVHD therapy, or minor histocompatibility disparities. The cell populations required for IgE response (T cells, B cells, and antigen-presenting cells) may be reconstituted in advance of the regulatory elements that limit IgE production in healthy subjects. Although this temporal relationship does not appear to hold in allogeneic BMT, the balance between positive and negative factors, which determines the rates of IgE synthesis and catabolism, may be altered by GVHD, infection, and liver dysfunction acting alone or in combination.

We report here the results of a phase I clinical trial using counterflow centrifugal elutriation (CCE) for the removal of donor T lymphocytes before allogeneic bone marrow transplantation (BMT). Thirty- eight patients received lymphocyte-depleted allografts from HLA- identical, MLR-nonreactive sibling donors. The patients entered onto the study were either at high risk on the basis of age (median, 39 years) or disease status (acute leukemia in early relapse [ER], chronic myelogenous leukemia [CML] in accelerated phase [AP], or therapy resistant [RES] lymphoma). All patients received a standard lymphocyte dose of 1 x 10(6) morphologic lymphocytes per kilogram ideal body weight (BW) and were maintained on cyclosporine A (CsA) for 170 days after BMT. Prompt engraftment occurred in 37 of 38 patients with a median time to absolute neutrophil count (ANC) greater than 500/microL of 18 days. Although acute graft-v-host disease (GVHD; clinical stage I or greater) was observed in 45%, it was limited to the skin in all but five patients. Survival was related to disease status at the time of BMT. Among patients with acute leukemia in first or second remission, CML in chronic phase (CP) or lymphoma in partial remission (PR), 64% are currently alive, in contrast to 31% of patients with acute leukemia in third remission or early relapse, CML in second CP or AP, or RES lymphoma. Median follow-up for all patients was 351 days (range, 105 to 711 days). We conclude that this procedure is safe and warrants further evaluation in a randomized efficacy trial.

To identify predictive parameters for incidence and severity of acute graft-versus-host disease (GVHD), 136 patients, transplanted with histocompatible marrow as therapy for aplastic anemia and hematologic malignancies, were examined using univariate and multivariate analyses. The risk of GVHD increased in patients with acute lymphocytic leukemia (p less than 0.05), in sex-mismatched donor-recipient pairs (p less than 0.01), and in patients older than 23.7 yr (p less than 0.05). No other commonly observed factors appeared to have any relationship to GVHD except the presence of certain alleles. The presence of a Cw4 allele or of the Bw21 specificities B49 and B50 were associated with significantly increased risks of GVHD (p less than 0.05), whereas the presence of Aw19 (or the related specificities A29, Aw30, Aw31 , Aw32, Aw33 ) was associated with a significantly decreased risk (p less than 0.01). Using these factors, a regression equation can be constructed that estimates the risk of a given patient to develop clinically significant acute GVHD.

Twenty-five patients with major ABO blood group incompatibility between donor and recipient underwent allogeneic bone marrow transplantation using erythrocyte depletion of the bone marrow infusate prior to administration. Over 95% of the original erythrocyte content of the marrows was removed, while retaining 75% of the mononuclear cell content and 57% of the granulocyte-monocyte colony-forming units. Recipients, well hydrated and premedicated with corticosteroids, diphenhydramine, and mannitol, tolerated infusions well. The frequency of engraftment, rate of recovery of peripheral blood leukocytes, granulocytes, and platelets, and the incidence of graft-versus-host disease was similar to that observed following ABO blood group compatible bone marrow transplantation. Erythroid development following ABO blood group incompatible transplantation was significantly impaired until hemagglutinins fell to 1:4 or lower, at which time recovery of erythrocytes was detected in the peripheral blood. The erythrocyte hypoplasia associated with incompatible hemagglutinins was temporary. Erythrocyte purging is a safe and effective technique to perform bone marrow transplantation across major ABO blood group incompatibilities.

One hundred fifty-three patients who underwent autologous bone marrow transplant (ABMT) were studied retrospectively to determine the frequency, outcome, and risk-factors associated with varicella-zoster infections (VZV). Forty-three patients (28%) developed VZV infection after transplant. The median onset of infection was the fifth month, with 91% of cases occurring within the first year. Thirty-three patients (77%) had localized herpes zoster, and ten patients (23%) had varicella. Cutaneous dissemination developed in 15% of patients and probable visceral dissemination developed in 5%. Overall morbidity was 25% and included scarring, alopecia, postherpetic neuralgia, and neurologic dysfunction. There were no deaths from VZV infection. The majority of patients (79%) were treated with intravenous (IV) acyclovir. The only significant risk factor associated with VZV infection was the underlying disease. VZV infection occurred most frequently in patients with Hodgkin's and non-Hodgkin's lymphoma (46%) as compared with patients with leukemia (23%) or solid tumors (9%) (P less than .002). The frequency of VZV infection in ABMT patients appears to be comparable to that reported for allogeneic BMT patients and other immunocompromised patients.

Lethally irradiated LouM rats reconstituted with syngeneic bone marrow and then treated with cyclosporine (CsA) for 40 consecutive days following transplant developed a graft-v-host disease (GVHD)-like syndrome after CsA cessation. This model of GVHD was used to define and characterize a graft-v-tumor (GVT) effect against a syngeneic plasmacytoma CRL1662 cell line which expresses class II major histocompatibility (MHC) antigen (Ia). Nylon wool-nonadherent spleen cells from animals who developed syngeneic GVHD were capable of significant lysis against chromium-labeled tumor target cells in a four- hour chromium released cell mediated lympholysis assay; maximum lysis occurred five days following cessation of CsA when clinical signs first appeared. Cytolytic activity declined to baseline as GVHD symptoms resolved. Fractionation of splenocytes into lymphocyte subsets demonstrated that cytolytic lymphocytes (CTLs) of the OX8 phenotype (non-helper T) were capable of significant lysis against tumor target cells. Lysis of tumor cells was blocked by preincubation with monoclonal antibodies (MoAb) specific for the rat anti-class II MHC antigen but not with MoAb against class I. Incubation of tumor cells with gamma-interferon increased expression of tumor class II MHC antigens and significantly increased their susceptibility to lysis by nylon wool-nonadherent splenocytes from animals with syngeneic GVHD. These studies have demonstrated an in vitro GVT of syngeneic GVHD against an Ia-bearing tumor; the effector cell is a CTL of the OX8 phenotype specific for the class II MHC antigen.

We examined the effects of varying incubation conditions on the in vitro activity of 4-hydroperoxycyclophosphamide (4HC). 4HC activity against CFU-GM and against the K562 tumor cell line decreased with increasing the RBC concentration of the incubation mixture. Increasing the concentration of nucleated bone marrow cells in the incubation mixture also decreased the 4HC activity. Evaluation of 53 consecutive patients undergoing autologous bone marrow transplantation (BMT) revealed that the incubation RBC concentration during clinical purging showed a similar effect on CFU-GM recovery. Aldehyde dehydrogenase content of RBCs and nucleated marrow cells appears to be the cause of the inhibition of 4HC activity. Although there was no difference in individual CFU-GM sensitivity to 4HC among normals, previously treated patients undergoing autologous BMT showed significant variability in CFU-GM sensitivity to 4HC. The combined effects of incubation RBC concentration and individual patient 4HC sensitivity appear to account for most of the variability in CFU-GM recovery and speed of hematologic recovery after clinical purging with 4HC.

Cytomegalovirus (CMV) infection was detected in 65 of 143 (45%) autologous bone marrow transplant (BMT) patients. CMV pneumonitis occurred in only 2% of the patients and CMV retinitis occurred in none. Infection occurred in half of the 40 initially seronegative patients and 47% of the 94 initially seropositive patients. Among initially seropositive patients, platelet recovery was slower in infected patients than in those not infected (97 v 35 days median, P = .003), and neutrophil recovery was slightly delayed in infected patients (31 days v 24 days, P = .02). Although the incidence of CMV infection was comparable in autologous and allogeneic BMT patients, CMV pneumonitis was less frequent in autologous BMT patients (2% v 12%, P less than .001). The risk for CMV pneumonitis in autologous BMT patients was comparable with that in allogeneic BMT patients without graft-v-host disease (GVHD) (2% v 6%), but significantly lower than the risk in allogeneic BMT patients with GVHD (2% v 23%, P less than .001).

Autologous bone marrow transplants (BMTs) can repopulate the hematologic system of patients treated with marrow-ablative chemotherapy and/or radiotherapy. However, treatment of the bone marrow graft to eliminate residual tumor cells prior to reinfusion can delay the return of peripheral blood elements, presumably from damage to or loss of hematopoietic stem cells responsible for hematologic recovery. To develop a model predictive of hematologic recovery, we studied the progenitor cell contents of 4-hydroperoxycyclophosphamide (100 micrograms/mL)-purged bone marrow grafts of 40 consecutive patients undergoing autologous BMT at this center. Granulocyte-macrophage colonies (CFU-GM) were grown from all grafts after treatment with this chemotherapeutic agent, but erythroid (BFU-E) and mixed (CFU-GEMM) colonies were grown from only 44% and 33% of the grafts respectively. The recovery of CFU-GM after purging ranged from 0.07% to 23%. The logarithm of CFU-GM content of the treated grafts was linearly correlated with the time to recovery of peripheral blood leukocytes (r = -0.80), neutrophils (r = -0.79), reticulocytes (r = -0.60), and platelets (r = -0.66). The CFU-GM content of purged autologous bone marrow grafts may reflect the hematopoietic stem cell content of the grafts and thus predict the rate of hematologic recovery in patients undergoing autologous BMT.

Cyclosporin-A (CsA) was given to 22 patients who received allogeneic bone marrow transplants as therapy for aplastic anemia and hematologic malignancies. The drug was given daily for 180 days starting with the day of marrow infusion. Engraftment was not impaired and myelotoxicity was not observed. Cutaneous graft-versus-host disease (GVHD) developed in five patients and all either spontaneously resolved or promptly responded to therapy with steroids. Five patients developed systemic GVHD and all responded to therapy with steroids, but only two survived. Interstitial pneumonia was seen in six patients and was fatal in all of them. Liver function abnormalities were seen in 14 patients but could not positively be correlated with CsA administration. Renal function abnormalities were seen in 17 patients. Amphotericin-B therapy contributed significantly to the renal failure. Serum levels of CsA, measured by radioimmunoassay, could not be correlated with the presence of liver or renal function abnormalities. Overall survival so far has been 50.0%. Second malignancies were not observed, but one patient relapsed with leukemia at 343 days.

A 22-yr-old man with acute myelocytic leukemia received a bone marrow transplant from a genotypically HLA-identical female sibling after cyclophosphamide preparation. He remained in complete remission for 18 mo, when he developed a chloroma in the perineum. The chloroma was treated with local radiotherapy. The chloroma recurred 8 mo later and was treated with radiotherapy followed by combination chemotherapy. At 34 mo after transplant, marrow relapse and chloroma were documented. The first chloroma contained host cells by fluorescent Y-chromatin body analyses of interphase nuclei. All metaphase cells and karyotypes from peripheral blood and marrow samples showed no evidence of host cells from 3 wk after transplant through the time of marrow relapse. Data from autosomal and sex chromosome studies indicate that the marrow relapse occurred in cells of donor origin. A new consistent chromosome abnormality [45, X, -X, t(8;21) (q22; q22)] was observed in a majority of donor cells. The patient received a second bone marrow transplant from the same donor after preparation with busulfan and cyclophosphamide and attained a complete remission with full hematologic engraftment.