Article

Rhesus Macaque Natural CD4 Regulatory T Cells Exhibit Decreased Proliferation But Enhanced Suppression After Pulsing with Sirolimus

The Emory Transplant Center, Department of Surgery, Emory University School of Medicine, Atlanta, GA, USA.
American Journal of Transplantation (Impact Factor: 5.68). 02/2012; 12(6):1441-57. DOI: 10.1111/j.1600-6143.2011.03963.x
Source: PubMed

ABSTRACT

Although regulatory T cells (Tregs) suppress allo-immunity, difficulties in their large-scale production and in maintaining their suppressive function after expansion have thus far limited their clinical applicability. Here we have used our nonhuman primate model to demonstrate that significant ex vivo Treg expansion with potent suppressive capacity can be achieved and that Treg suppressive capacity can be further enhanced by their exposure to a short pulse of sirolimus. Both unpulsed and sirolimus-pulsed Tregs (SPTs) are capable of inhibiting proliferation of multiple T cell subpopulations, including CD4(+) and CD8(+) T cells, as well as antigen-experienced CD28(+) CD95(+) memory and CD28(-) CD95(+) effector subpopulations. We further show that Tregs can be combined in vitro with CTLA4-Ig (belatacept) to lead to enhanced inhibition of allo-proliferation. SPTs undergo less proliferation in a mixed lymphocyte reaction (MLR) when compared with unpulsed Tregs, suggesting that Treg-mediated suppression may be inversely related to their proliferative capacity. SPTs also display increased expression of CD25 and CTLA4, implicating signaling through these molecules in their enhanced function. Our results suggest that the creation of SPTs may provide a novel avenue to enhance Treg-based suppression of allo-immunity, in a manner amenable to large-scale ex vivo expansion and combinatorial therapy with novel, costimulation blockade-based immunosuppression strategies.

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    • "There are precedents for the loading of cells with hydrophobic drugs, with Pessina et al. [35] reporting that sufficient amounts of the anti-cancer drug paclitaxel were adsorbed by MSCs in a 24-h incubation to reduce tumor growth in vivo and Orange et al. [28] finding that FK-506eloaded dendritic cells could inhibit autoimmune arthritis. A rapid partition of ISDs into cells would appear to explain our observations of increased suppression by brief pretreatments of MSCs, fibroblasts, APCs and HUVECs and may explain at least part of the increased suppression reported for rapamycintreated endothelia and Tregs [30] [36]. It is well documented that rapamycin partitions into blood cells [25], and measurement of rapamycin in our treated MSCs showed sufficient levels of drug even after extensive washing to inhibit T-cell proliferation in co-culture. "
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    • "It has been reported that combination of high dose IL-2, rapamycin, anti-CD3 and (or) anti-CD28 beads or monoclonal antibodies and artificial APCs (aAPC) can expand Treg massively in mice and humans [10] [11] [12]. In contrast to these studies in mice and humans, few reports have addressed expansion of Treg from non-human primates (NHP), -important pre-clinical models in organ transplantation [13] [14] [15] [16] [17] [18]. Given the challenge of manufacturing Treg on a large scale and the potential advantage of banking these cells, cryopreservation may facilitate their clinical application [19]. "
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    • "Many of the studies investigating the role of mTOR in T regs have relied on the use of rapamycin (also known as sirolimus), which selectively inhibits mTORC1 at low doses but can also inhibit mTORC2 at higher doses (Delgoffe et al., 2011). Unlike conventional T cells, T regs are resistant to rapamycin-induced apoptosis (Strauss et al., 2009) and hence this drug can selectively block pro-inflammatory T cells while preserving T regs (Battaglia et al., 2006; Qu et al., 2007; Lu et al., 2010; Zuber et al., 2011) and their suppressive function (Singh et al., 2012). These data support the conclusion that activation of T regs does not require strong activity of the PI3K pathway. "
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