Background: Acute graft versus host disease (GVHD) remains a major cause of morbidity and mortality after allogeneic hematopoietic cell transplantation. Preclinical studies have suggested that a T-cell subset with a CD4-/CD8- double-negative (DN) T-cell phenotype is capable of suppressing GVHD. Double-negative T cells can be mobilized into the peripheral blood with granulocyte colony-stimulating factor (G-CSF) and enriched by density centrifugation. The current study was performed to study the feasibility and safety of applying a density gradient separation technique for enrichment of CD34+ and DN T cells, while depleting CD4+ and CD8+ single-positive (SP) T cells from peripheral blood progenitor cells (PBPCs) for the purpose of allogeneic transplantation.
Methods: Twenty-five patients with advanced hematologic malignancies were treated with a myeloablative preparative regimen consisting of fractionated total body irradiation, etoposide, and cyclophosphamide. Human leukocyte antigen identical donors were mobilized with G-CSF PBPC collected by apheresis. The apheresis product was applied to a single-step density gradient, and the low-density cell population was collected. The low-density cell population was infused as the sole source of allogeneic cells after myeloablative therapy. Graft versus host disease prophylaxis consisted of cyclosporine with or without prednisone.
Results: CD34 cell recovery was efficient with a median 72% yield, providing for a median CD34+ cell dose of 6.5 x 10(6)/kg (range,1.0- 13.9 x 10(6)/kg). CD3+CD4+ or CD3+CD8+ SP T cells were depleted by a median of 94.4% (range, 58.8- 99.2%), and the ratio of CD34+:SP T cells increased 10-fold. Double-negative T cells were depleted by 92% (range, 18.8- 99.4%), thus the ratio of DN:SP T cells increased less than 2-fold in 71% of apheresis samples tested. Hematopoietic engraftment was rapid, and there was no occurrence of graft failure in examinable patients. Median time to absolute neutrophil count greater than 0.5 x 10(9)/L and platelet count greater than 20 x 10(9)/L was 10.5 and 12 days, respectively. The incidence of Grade 2-4 acute GVHD was 26% (95% confidence interval [CI], 6-45%), although not all patients were examinable due to an unexpectedly high nonrecurrence mortality that at Day 180 was 62% (95% CI, 40-83%).
Conclusions: These data suggest that T-cell subset manipulation via density gradient separation is a safe procedure and allowed rapid hematopoietic recovery. Selective enrichment of a donor DN T-cell subset was observed in only a few and was not associated with a reduced incidence of GVHD. However, the low-density selected cells still resulted in GVHD, and there was a high treatment-related mortality.
Copyright 2001 American Cancer Society.