title: Massive ex vivo expansion of human natural regulatory T cells (T(regs)) with minimal loss of in vivo functional activity.
abstract: Graft-versus-host disease (GVHD) is a frequent and severe complication after hematopoietic cell transplantation. Natural CD4(+)CD25(+) regulatory T cells (nT(regs)) have proven highly effective in preventing GVHD and autoimmunity in murine models. Yet, clinical application of nT(regs) has been severely hampered by their low frequency and unfavorable ex vivo expansion properties. Previously, we demonstrated that umbilical cord blood (UCB) nT(regs) could be purified and expanded in vitro using good manufacturing practice (GMP) reagents; however, the initial number of nT(regs) in UCB units is limited, and average yield after expansion was only 1 × 10(9) nT(regs). Therefore, we asked whether yield could be increased by using peripheral blood (PB), which contains far larger quantities of nT(regs). PB nT(regs) were purified under GMP conditions and expanded 80-fold to yield 19 × 10(9) cells using anti-CD3 antibody-loaded, cell-based artificial antigen-presenting cells (aAPCs) that expressed the high-affinity Fc receptor and CD86. A single restimulation increased expansion to ~3000-fold and yield to >600 × 10(9) cells while maintaining Foxp3 expression and suppressor function. nT(reg) expansion was ~50 million-fold when flow sort-purified nT(regs) were restimulated four times with aAPCs. Indeed, cryopreserved donor nT(regs) restimulated four times significantly reduced GVHD lethality induced by the infusion of human T cells into immune-deficient mice. The capability to efficiently produce donor cell banks of functional nT(regs) could transform the treatment of GVHD and autoimmunity by providing an off-the-shelf, cost-effective, and proven cellular therapy.
authors: Division of Bone Marrow Transplantation, Department of Pediatrics, University of Minnesota Cancer Center, Minneapolis, MN 55455, USA. hippe002@umn.edu
authors: Keli L
authors: KL
authors: Hippen
authors: Sarah C
authors: SC
authors: Merkel
authors: Dawn K
authors: DK
authors: Schirm
authors: Christine M
authors: CM
authors: Sieben
authors: Darin
authors: D
authors: Sumstad
authors: Diane M
authors: DM
authors: Kadidlo
authors: David H
authors: DH
authors: McKenna
authors: Jonathan S
authors: JS
authors: Bromberg
authors: Bruce L
authors: BL
authors: Levine
authors: James L
authors: JL
authors: Riley
authors: Carl H
authors: CH
authors: June
authors: Phillip
authors: P
authors: Scheinberg
authors: Daniel C
authors: DC
authors: Douek
authors: Jeffrey S
authors: JS
authors: Miller
authors: John E
authors: JE
authors: Wagner
authors: Bruce R
authors: BR
authors: Blazar
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