Abnormalities by Multicolor Flow Cytometry for Detection of Minimal Residual Disease in Recipients of Allo-HSCT Originating from Donors: A Cohort Study

Objective: In minimal residual disease (MRD) analysis after allogeneic hematopoietic stem cell transplantation (allo-HSCT), abnormal immunophenotyping is commonly considered as evidence of a secondary recurrence or complications, leading to overtreatment. We aimed to confirm whether such phenotypic abnormality might originate from donors using multicolor flow cytometry (MFC).Materials and Methods: The MRD of bone marrow specimens of 3395 patients who had received allo-HSCT were analyzed using the conventional two-tube, eight-color MFC panel. The frequencies of abnormal immunophenotypes were also evaluated in three groups of patients without malignancies.Results: The frequency of new abnormal polymorphisms was 0.088% (3/3395) among patients who received allo-HSCT. The abnormal cells seen in three patients in complete remission were Fc gamma receptor IIIB (Fc gamma RIIIB) gene deletion (CD16- neutrophils), CD2-CD159a-CD159c+ natural killer (NK) cells, and monoclonal B lymphocytosis (MBL), respectively. In addition, abnormal T-cells (CD4+CD8+) were detected in one donor before allo-HSCT. Identical abnormalities were found in the peripheral blood of the corresponding donors of the three patients via MFC. Among the individuals without malignancies, the incidence of Fc gamma RIIIB deletion was 0.2% (11/5256), that of NK cells with the absence of CD2 and single-positive CD159c was 0.05% (1/2000), that of monoclonal CD4/CD8 double-positive T-cells was 0.05% (1/2000), and that of MBL was 1.3% (14/1100). The frequency of NK cells with the absence of CD2 was 1.3% (1/79) and with CD8dim was 14% (11/79) in NK cell lymphoma. The following abnormalities could be identified by the two-tube, eight-color MFC panel: c Kappa/c lambda/CD19/CD5/CD20/ CD38/CD45/CD56 (adding CD10 and CD34 as the ninth and tenth colors) and CD16+CD56/CD5/CD3/CD7/CD4/CD8/CD2/CD45 (adding CD117 as the ninth color).Conclusion: Abnormalities in recipients of allo-HSCT detected by MRD analysis may originate from their donors. Screening of donor specimens with a suitable two-tube, eight-to ten-color MFC panel may be a promising method for minimizing misdiagnoses.