P-BT-2 - Two Distinct CD45 Positive Stem Cell Populations Within a Single Stem Cell Product – a Clinical Practice Conundrum

Flow cytometric evaluation of donor Hematopoietic Progenitor Cells, Apheresis [HPC(A)] is a critical quality control measure for appropriate stem cell transplant dose and/or cryopreservation. Two major determinants of product manipulation are the CD34+ population - to ensure adequate dosage upon transplant; and the CD3+ population - to minimize the risk of severe graft versus host disease (GVHD). Here we describe a unique situation wherein flow cytometry of a single donor product revealed two distinct populations of CD45+/CD34+ cells, presenting a unique best practice conundrum.

Study

Design/Methods: Multi-channel flow cytometric evaluation on the donor HPC(A) was performed per the standard operating procedure (SOP). Additional flow cytometric characterization was performed within the Division of Hematopathology.

Results/Findings:

The donor product was received through the NMDP. Initial flow cytometric evaluation revealed two CD45+/CD34+ populations with one population demonstrating distinctly lower CD45 expression (hereafter referred to as CD45^dim/CD34+). Repeat analysis revealed similar results. Gating to include both populations resulted in a dose of 10.57 x 10⁶ CD34+ cells/kg. Exclusion of the CD45^dim/CD34+ population resulted in a dose of 7.33 x 10⁶ CD34+ cells/kg. The CD3+ concentration in the product was 2.45 x 10⁸ CD3+ cells/kg. The requested dose for transplant was 5 x 10⁶ CD34+ cells/kg. Cryopreservation based on the 7.33 x 10⁶ CD34+ cells/kg concentration presented the caveat of a higher CD3+ cell content, increasing the risk of GVHD. Cryopreservation based on the 10.57 x 10⁶ CD34+ cells/kg concentration risked underdosing and engraftment issues if the CD45^dim /CD34+ cells were later determined to be non-hematopoietic stem cells. Another significant concern was that this population was potentially an undiagnosed malignancy in the donor. The product was sent for additional flow cytometric immunophenotyping in the Division of Hematopathology. The CD45^dim/CD34+ population was determined to be myeloid blasts with no immunophenotypic abnormalities. Following confirmation of benignity, a collection dose of 10.57 x 10⁶ CD34+ cells/kg, 2.45 x 10⁸ CD3+ cells/kg and 0.36 x 10⁹ ALC/kg were reported for transplant.

Conclusions: Flow cytometric characterization of stem cell products is crucial to ensuring appropriate dosage and engraftment. Here we describe a rare situation in which a phenotypically distinct CD45^dim/CD34⁺ cell population was identified and underwent expert hematopathology evaluation with flow cytometric immunophenotypic analysis to exclude an undiagnosed hematological malignancy. Confirmation of benignity also allowed the total CD3+ dose to be adjusted, lowering the risk of severe GVHD.