Monitoring Of CD3～+T Cell Chimerism After Allogeneic Hematopoietic Stem Cell Transplantation

【Objective】:Applying quantitative methods to analyse CD3+T cell donor chimerism(DC) and its kinetics after allogeneic hematopoietic stem cell transplantation(allo-HSCT), to study the relationship between CD3+T cell DC and the outcome of disease and to compare DC of T cell fraction with the chimeric status of peripheral blood(PB)or bone marrow(BM).①The engraftment of donor T cell after allo-HSCT conditioned by modified Bu/CY regiment.②The relationship between T-DC and acute graft versus host disease (aGVHD).③The connection among the kinetics of T-DC and disease relapse or graft rejection(GR).【Methods】:①PB or BM was obtained at different time points of 19 patients with hematologic malignancies after allo-HSCT conditioned by modified Bu/CY regiment. High purity CD3+T cells were gained using magnetic cell separation technique(MACS). The genomic DNA was extracted from PB/BM and T cells respectively. The technique of multiplex amplification of short tandem repeats by fluorescence labeling polymerase chain reaction(STR-PCR) combined with automated capillary electrophoresis were used to analyse donor chimerism. At the same time, the chimeric status of donor T cells were evaluated in the study of engraftment kinetics and prediction of aGVHD.②PB and BM was collected from patients who had relapsed (3 patients) or GR (2 patients) after allo-HSCT. The chimeric status of donor T cells, whole blood and BM were analysed using the same methods as above in order to investigate the relationship between the donor T cell chimerism and disease relapse or graft rejection.【Results】:①All 19 patients achieved successful engraftment. Donor chimerism of whole blood or BM showed full donor chimerism(FDC) on day14 to day21 post-transplant. But the engraftment of donor T cells were delayed, the median time of achieving donor T cell FDC was +60 (+14~+180) days. At the early period of transplantation (+14d), the T cell engfted faster in patients undergoing unrelated donor transplantation than sibling donor transplantation. Among 12 patients whose T cell achieved FDC or high donor percentage(≥90%) in 28 days after allo-HSCT ,8 patients developed different degree of aGVHD, while in 7 patients whose donor T cell chimerism was below 90% in +28d , only 1 patients presented aGVHD(P<0.01).②Two of the 3 relapsed patients turned from FDC to mixed chimerism(MC) in whole blood and BM at the time of relapse, as well as T cell fraction. The DC of other one was MC in BM, while still FDC in whole blood and T cell fraction. The DC of PB or BM in 2 patients who had GR turned from FDC to MC, and T-DC of both were all below 30% .【Conclusion】:①Donor T cell engraftment was slower after allo-HSCT conditioned by modified Bu/CY regiment, which can maintain MC corresponding long period after transplantation.②The engraftment of donor T cell was influenced by regiment of GVHD prophylaxis . T cell engfted faster in patients with unrelated donors than others with sibling donors because of more intensive GVHD prophylactic regiment.③The patients whose T cell achieved FDC or high donor percentage(≥90%) in +28 days after allo-HSCT had high risk of developing aGVHD.④The value of T-DC in predicting disease relapse, especially in patients who relapsed later post-transplant still need further research.⑤Low percentage of T-DC maybe a sensitive parameter to predict GR, but also still need further investigation.⑥The monitoring of donor T cell chimerism which can be addition of whole blood or BM DC after allo-HSCT , but can not instead of it.⑦Individual adoptive immunotherapy should be implemented according to the results of lineage-specific chimerism, especially the chimeric status of donor T cell fraction.