Improving immune reconstitution after cord blood transplantation using ex vivo expanded virus-specific T cells: a phase I clinical study

Abstract

Ischemic stroke (IS) is the leading cause of adult disability and IS patients have few options to augment the rehabilitation process. The use of autologous mesenchymal stromal cells (MSCs) as a treatment of stroke has shown promise. One significant limitation to the use of autologousMSCs for stroke is that cells cannnot be transfused until 5 weeks post-IS due to the time required for expansion. We hypothesize that treating patients with MSCs within 48 hours of a stroke may improve outcomes. We plan to expedite the time-to-infusion by using banked allogeneic-MSCs. Here we have compared the use of an automated cell culture device (the Quantum by Terumo BCT), which uses 2.1m of hollow fibers in a bioreactor (equivalent to w120 T-175cm flasks), to our current flask-based method. In flasks, human bone marrow (BM) mononuclear cells (BMMC) were seeded at 5 105 BMMC/cm in T-175cm flasks and split 1:4 when near confluence. In the Quantum, w25 mL of unprocessed BM was added to the bioreactor. After w14 days, cells were harvested and 2.0-3.5 107 cells were re-seeded in a new bioreactor for w6 days. After 3-4 passages in flasks, an average of 2.89 108 MSCs were expanded from fourBMdonors.MSCsharvestedafter2passages in theQuantumyieldedanaverage of 6.63 108 cells. MSCs expanded in both flasks and the Quantum were analyzed by flow cytometry and met the ISCT minimum criteria for MSC (CD73, CD105,CD90 expression). Expanded MSCs were sterile, were free from chromosomal abnormalities, and differentiated into adipocytes, chondrocytes, and osteoblasts in vitro.The production of 5 108MSCs inflasks required 101-hours of labor compared to 21-hours with the Quantum. We estimated that during the manufacture of MSCs in flasks, there were 600 open events compared to 9 in the Quantum. This project is supported by NHLBI-PACT, contract # HHSN268 201000007C.