Pulmonary bronchoalveolar cell and protein kinetics in dogs given total-body irradiation, autologous marrow grafts, and methotrexate

Patients receiving allogeneic marrow transplantation for hematologic malignancies commonly are conditioned with total body irradiation (TBI) and given methotrexate (MTX) in an attempt to prevent graft-versus-host disease. To study the effects of TBI with or without MTX on bronchoalveolar cells and proteins, we performed sequential bronchoalveolar lavages in dogs before and after irradiation. Ten dogs received 9 Gy TBI followed by autologous marrow grafts. Six dogs were given no additional treatment and four also received MTX at 0.4 mg/kg on days 1, 3, 6, and 11- and then weekly until day 100. TBI alone resulted in a significant decrease in alveolar macrophages and lymphocytes with recovery after day 30. The addition of MTX resulted in a more profound and prolonged decrease in alveolar macrophages and lymphocytes. The addition of MTX was also associated with a significant increase in alveolar granulocytes with a concomitant rise in lavage protein content in one animal. Lavage fluid IgA levels remained constant. We conclude that the irradiation and chemotherapy used in marrow transplantation has significant pulmonary effects and may contribute to the pulmonary complications following marrow transplantation.     

Effects of treating marrow with a CD3-specific immunotoxin for prevention of acute graft-versus-host disease

Data from human clinical trials and animal experiments have suggested that T lymphocytes in donor marrow help to facilitate engraftment after allogeneic bone marrow transplantation, possibly through a suppressive effect on the immunity of the recipient. In previous studies marrows from HLA-identical donors were treated ex vivo with a mixture of eight monoclonal antibodies together with rabbit complement to achieve a 3-log depletion of T cells and CD3-negative lymphoid cells. Transplantation of this marrow was associated with a 27% actuarial risk of graft failure in leukemic recipients conditioned with cyclophosphamide (120 mg/kg) and 15.75 Gy fractionated total body irradiation. In the present study, we employed an anti-CD3 ricin A-chain-containing immunotoxin (64.1-A) together with 20 mM NH4Cl to achieve a selective 3-log depletion of CD3-positive cells. The patient entry criteria and pretransplant conditioning regimen were identical to those used in previous studies. Despite the differences in marrow treatment, the clinical outcome of the present study was similar to that obtained previously. Graft-versus-host disease (GVHD) was largely prevented without the need for post-transplant immunosuppression, but two of the eight patients developed graft failure. These results indicate that CD3-negative cells have little or no ability to initiate GVHD. To the extent that graft failure in this study was not caused by stem cell damage or loss of CD3-negative cells during ex vivo processing of the marrow, it appears that the lymphoid cells required for facilitating allogeneic engraftment under these conditions are CD3-positive.     

Allogeneic marrow transplantation in the treatment of MOPP-resistant Hodgkin's disease

Eight patients with disseminated Hodgkin's disease resistant to MOPP (mechlorethamine, vincristine, procarbazine, and prednisone) chemotherapy were treated with high-dose chemoradiotherapy and marrow transplantation from an HLA-identical sibling. Two patients remain alive in unmaintained complete remission (CR) at 38 and 39 months after transplant. In the other six patients, reasons for failure included relapse of lymphoma (two patients), or death due to complications of the transplant procedure, including Legionnaire's disease, disseminated zoster, graft-v-host disease, and aspiration pneumonia secondary to severe mucositis. These results demonstrate that some patients with MOPP-resistant Hodgkin's disease can obtain prolonged CR following intensive chemoradiotherapy and allogeneic marrow transplantation.     

Pharmacokinetics of intravenous cyclosporine in bone marrow transplant patients. Comparison of two assay methods

The influence of assay method on steady-state cyclosporine (CsA) pharmacokinetics was studied in 18 patients with leukemia or aplastic anemia undergoing allogeneic marrow transplantation who received i.v. CsA for prophylaxis or treatment of graft-versus-host disease. Since CsA is extensively metabolized and subject to biliary elimination, CsA courses were divided according to the presence (serum bilirubin greater than 2.0 mg/dl) or absence (serum bilirubin less than or equal to 1.2 mg/dl) of hepatic dysfunction. All patients had normal renal function. Serum CsA concentrations were measured concurrently by radioimmunoassay (RIA) and high-performance liquid chromatography (HPLC). Serum concentration-time data were analyzed by standard nonlinear regression methods. Systemic clearance (Cls) calculated from HPLC results was higher than that derived from RIA results, regardless of hepatic function (P less than 0.05). These data indicate that results obtained by RIA overestimate CSP concentrations, presumably because of the presence of crossreactive CSP metabolites. These differences can significantly affect derived pharmacokinetic parameters. Therefore, clinicians who make dosage recommendations based on pharmacokinetic parameters should consider the effect of assay method.     

Recognition of target cell determinants associated with DLA-D-locus-encoded antigens by canine cytotoxic lymphocytes

Cytotoxic lymphocytes (CTL) were generated in bulk mixed leukocyte culture (MLC) using peripheral blood mononuclear cells from homozygous dogs identical to each other for DLA-A and -B but different for DLA-D loci. These CTL were tested against 51Cr-labeled targets (Con-A-stimulated PBMC) derived from the MLC sensitizing dogs and from other dogs homozygous for various DLA-D locus alleles. Indirect cell-mediated lympholysis (CML) assays, with and without cold target blocking and pretreatment of 51Cr-labeled targets with DLA alloantisera were used to study the target cell determinants recognized by these CTL. The resultant target cell lysis and cold target blocking of specific target cell lysis indicated target determinants associated with DLA-D-locus-encoded specificities, and different from those encoded by DLA-A and -B loci. The pattern of shared target determinants among different DLA-D HTC may be explained by: (A) two separate previously unrecognized MHC associated loci, each with 2 alleles defined so far; or (B) epitopes shared among certain DLA-D alleles but not present in others. This study demonstrates that DLA-D associated determinants different from previously described DLA-D specificities may serve as CML target cell determinants when there are no apparent DLA-A and -B loci encoded antigen differences between responder and stimulator cells used to generated MLC-derived CTL.