Identification, selection, and expansion of non-gene modified alloantigen-reactive Tregs for clinical therapeutic use

Abstract

Transplantation is limited by the need for life-long pharmacological immunosuppression, which carries significant morbidity and mortality. Regulatory T cell (Treg) therapy holds significant promise as a strategy to facilitate immunosuppression minimization. Polyclonal Treg therapy has been assessed in a number of Phase I/II clinical trials in both solid organ and hematopoietic transplantation. Attention is now shifting towards the production of alloantigen-reactive Tregs (arTregs) through co-culture with donor antigen. These allospecific cells harbour potent suppressive function and yet their specificity implies a theoretical reduction in off-target effects. This review will cover the progress in the development of arTregs including their potential application for clinical use in transplantation, the knowledge gained so far from clinical trials of Tregs in transplant patients, and future directions for Treg therapy.

Keywords: Alloantigen-reactive Tregs; Cellular therapy; Clinical trials; Regulatory T cells; Transplantation; polyTregs.

Fig. 1

Original dataset. CD137⁺ alloantigen-expanded Tregs (green) are more suppressive than polyclonal Tregs (red), non-enriched Tregs (blue) and CD137^neg alloantigen-expanded Tregs (purple). (A) CD137 expression upon alloantigen stimulation of flow-sorted CD4⁺CD25⁺CD127^lo human Tregs. Tregs were stimulated with immature dendritic cells (iDCs) at a cell-to-iDC ratio of 4:1. Cells were assessed for CD137 expression by flow cytometry. Data from 9 healthy human donors are shown. (B) Experimental schematic for in vitro expansion of polyclonal and alloantigen expanded Tregs created with BioRender.com (C) Suppression assays were performed using 3H-thymidine incorporation; responder cells were stimulated with allogeneic iDCs. Polyclonally expanded, alloantigen-expanded non-CD137 enriched, alloantigen-expanded enriched CD137⁺ and CD137^neg Tregs were titrated into the culture. Responders alone were used as a negative control. Responders with alloantigen were used as a positive control. Six days later, thymidine was added to the culture and after 16 h of incubation cells were harvested. Data are represented as mean +/-SD, statistical analysis was performed using unpaired t-tests (*p < 0.05, **p < 0.01, ***p < 0.001). Representative data from one donor out of five is shown.