Diluting high yield hematopoietic stem cell apheresis for autologous transplantation results in high viability of fresh grafts beyond 72h

Abstract Background: Autologous Hematopoietic Cell Transplantation (HCT) remains a highly efficient therapy primarily for patients with Multiple Myeloma, Lymphomas, Solid Tumors and several Autoimmune Diseases. While fresh grafts for Multiple Myeloma stored for up to 48 hours has been robustly described for many years now, the storage of Hematopoietic Stem Cells (HPSC) beyond 72 hours has been questioned which serves as a barrier to access for starting programs specially for Lymphoma patients in which the most common conditioning protocol (BEAM) requires several days of administration. This situation is aggravated in Low-Middle-Income Countries (LMIC) where access to cryopreservation facilities is not always available. At the Instituto de Prevision Social all lymphoma patients are transplanted with fresh grafts using a shortened BEAM protocol with time to collect to infusion of 96 hours (BCNU d-4; Ara-C d-4, d-3, d-2; Etoposide d-4, d-3, d-2; Melphalan d-2) and minimum CD34+ cells of 2,5 x 10*6. According to the 2024 EBMT Handbook in case of storage of HPSC beyond 24 hours before cryopreservation the maximum total nucleated cells (TNC) should not exceed 2 x 10*8/ml (200,000/uL). Methods: This is a retrospective one center data collected over the course of 4 years. Patients transplanted with Lymphoma from July 2022 to October 2024 with BEAM protocol received unmanipulated fresh grafts while patients transplanted after November 2024 had their products diluted with patient plasma if TNC exceeded 2 x 10*8/ml. Viability checks were performed routinely by flow cytometry on collection day and on infusion day (+4). All patients were mobilized with GCSF +/- Plerixafor. Results: 33 patients diagnosed with Lymphoma were transplanted during the 2022-2025 period. Viability of all products on collection day was 95% or above. 29 HPSC were infused for 19 patients before the dilution protocol became standard with a median viability on infusion day (+4) of 80% (range 26%-99%). 15 HPSC were infused for 14 patients after the dilution protocol became standard with a median viability on infusion day (+4) of 94% (range 70%-99%). No graft failures were observed. Conclusion: While this is a very limited study due to number of participants, its retrospective nature and single center experience the data suggests diluting high yield apheresis products result in high viability of HPSC beyond 72 hours. Any HCT Program should aim to have cryopreservation facilities available as there are some patients that cannot be transplanted safely without said capacities. However, this may not be the case for Lymphoma patients. Diluting high TNC products results in more stable and predictable viabilities on infusion day. More studies are required to confirm these results.