Single center evaluation of TCRαβ+ T cell depletion using CliniMACS® plus platform: Optimizing cell selection for hematopoietic stem cell transplantation

Abstract Introduction TCRαβ+ T cell-depletion is a promising ex vivo graft engineering technique that effectively reduces the risk of graft-versus-host disease (GVHD) in haplo-hematopoietic stem cell transplantation. However, the magnetic sorting process typically results in a 30%-50% loss of CD34+ cells, posing challenges in balancing adequate CD34+ cell yields with thorough depletion of TCRαβ+ T cell. This retrospective study analyzed 398 TCRαβ+ T cell-depleted procedures performed at our center with the CliniMACS® Plus, aiming to advance clinical adoption of TCRαβ-depleted transplantation—particularly in higher-weight patients—through real-world evidence. Methods We retrospectively analyzed 398 αβ+ T cell-depleted procedures at our center between 2019-2024. Per clinical requirements for cell dose, we collected large-volume peripheral blood stem cell harvests containing substantial TCRαβ⁺ T-cell loads (up to 3.6×1010 cells), representing approximately 1.5 times the standard processing capacity. Cell separations were performed using CliniMACS® Plus (Miltenyi Biotec) for target cell depletion. Subset counts (CD34+, TCRαβ+ T, TCRγδ+ T, NK cells) were monitored via flow cytometry throughout the magnetic separation process. Results The characteristics of donors and patients were summarized in Table 1. Donor median age was 33 years (5–52); median weight was at 58.9 kg (15.2–106.0). Recipient median age was 6 years (0.5–58); median weight was at 20 kg (5.3–87.0), including 14 patients over 50 kg . Median yields were 85% for CD34+, 78% for TCRγδ+ T cells, and 87% for NK cells. TCRαβ+ T cell depletion efficiency log was 4.65 (3.71-5.81). The median compositions of graft were 25.76×10⁶/kg CD34+ cells; 19×103/kg TCRαβ+ T cells, 33.27×10⁶/kg TCRγδ+ T cells and 100.68×10⁶/kg NK cells. For patients over 50 kg, CD34+ cells were 18.5×10⁶/kg; TCRαβ+ T cells were 27.5×103/kg; TCRγδ+ T cells were 20.93×10⁶/kg; NK cells were 66.69×10⁶/kg (Table 2). All patients received TCRαβ⁺ T-cell infusions meeting our clinical per-kilogram body weight threshold. Primary engraftment rates were achieved in 90.45% (360/398) of all patients and 100% (14/14) of higher-weight patients (Table 2). Conclusion The CliniMACS Plus and matched reagents reliably support TCRαβ+ T cell depletion even at 150% standard processing volumes while maintaining high depletion efficiency (>3.7 log). This enables mega-dose CD34+ cell infusion (up to 10×10⁶/kg), even in high body weight patients. Key words: haplo-hematopoietic stem cell transplantation, TCRαβ+ T cell-depletion, TCD, CliniMACS.