Scholars@Duke publication: Dendritic cell (DC) reconstitution after nonmyeloablative allogeneic stem cell transplants

Dendritic cell (DC) reconstitution after nonmyeloablative allogeneic stem cell transplants

Publication , Conference

Morse, MA; Rizzieri, D; Hobeika, AC; Chao, N; Kirn Lyerly, H

Published in: Blood

December 1, 2000

Non-myeloablative allogeneic peripheral blood stem cell transplants {mini-allo PBSCT) are being evaluated as a lower toxicity alternative to conventional allogeneic transplantation for harnessing graft versus tumor effects. Induction of anti-tumor immunity post-transplant requires donor derived DC be available to process and present antigen to the donor lymphocytes. Because the kinetics of DC reconstitution after mini-allo PBSCT are unknown, we have initiated a study to evaluate DC number and function in 8 donor/recipient pairs prior to the preparative regimen and at various times after PBSC reinfusion. Peripheral blood mononuclear cells (PBMC) were separated over Ficoll and stained for FACS with antibodies to CD lie, HLA DR, and CD 14 to detect myeloid DC1, or lineage markers, CD123, HLA-DR, and CD1 le to detect lymphoid DC2. To determine the number of DC that can be generated from PBMC, we cultured the plastic adherent PBMC in serum free medium containing GM-CSF (800U/ml) and IL-4 (500U/ml) for 7 days and determined the percentage of large HLA DR+ cells by FACS. Diagnoses were AML 1, NHL 4, MDS 1, ET 1, and thalassemia 1. The preparative regimen was cyclophosphamide, fludarabine, and CAMPATH-1. Donor-derived G-CSF mobilized leukaphereses were depleted of T cells with CAMPATH-1 and reinfused. Five patients have reached day 14 following reinfusion. One failed to engraft, and the remainder achieved 43-99% donor chimerism in the bone marrow by RFLP analysis. Two patients died of disease progression. The median number of CD1 lc+CD14-HLA-DR+DC was 2.3% of the recipient PBMC prior to the preparative regimen, 1.5% of the donor leukapheresis PBMC, and 0.9% of the recipient peripheral blood at day 14 (P=NS). In two recipients who had CD1 lc-CD123bright DC2 measured after transplant, they were 0% and 0.7% of the PBMC compared to 0.13% and 0.26% in the donor leukapheresis. The median yield of DC generated from PBMC was 14% in the donor leukaphereses and 17% in the recipient peripheral blood at day 14 (P=NS). Three recipients had DC yields of < 1 % prior to the preparative regimen. This preliminary data suggests that myeloid DC 1 and DC precursors are present in the peripheral blood at the time of myeloid re-engraftment two weeks after mini-allo PBSCT. We are currently evaluating a) whether these DC are of donor or recipient origin, b) whether DC numbers stabilize at subsequent time points, c) whether DC function following transplant is diminished from pre-transplant levels. These results will guide studies of anti-tumor immunization following transplantation.