Investigation of CD27-CD70 co-stimulation in human regulatory T cells for enhanced cellular therapy in transplantation

CD4+FOXP3+ regulatory T cells (Tregs) are essential for maintaining immune homeostasis and are a focus of investigation in the treatment of transplant rejection and autoimmune diseases. Current clinical protocols for Treg cellular therapy production are heavily reliant on in vitro expansion techniques. However, post-expansion Tregs demonstrate heterogeneity with some subpopulations displaying unstable regulatory phenotype and function. The aim of this study was to investigate this phenomenon in depth, in order to improve the efficacy and safety of Treg cellular therapy. Cell surface CD27 is a useful marker for the identification of Tregs with potent suppressive capacity after expansion. Signalling via the CD27-CD70 co-stimulatory receptor-ligand interaction is important for T cell function but its role in Tregs is not known. In this study, the CD27+ Treg population is characterised in depth. The differential expression of CD27 and CD70 is shown to identify distinct human Treg populations. Stable and potent suppressive function after in vitro expansion is confined principally to cells with a CD27+CD70- phenotype. However, this suppressive function is not reliant on CD27 expression. Prolonged in vitro expansion of Tregs results in stable CD70 expression with concomitant CD27 downregulation in a subset of Tregs, and these CD27-CD70+ Tregs provide pro-inflammatory co-stimulation to conventional T cells via CD70. Genetic deletion (using CRISPR/Cas9) or monoclonal antibody-based blockade of CD70 rescues Tregs from hypofunctionality, highlighting a potential therapeutic intervention. Taken together, our data identify CD70 as a molecular target via which in vitro-expanded human Tregs exert detrimental pro-inflammatory effects. For clinical application, the availability and ongoing development of antibodies that target the CD27/CD70 pathway provides an opportunity for the therapeutic manipulation of Tregs. Furthermore, relatively straightforward genetic engineering techniques could be exploited to disrupt CD70 expression and improve the efficacy of Treg cellular therapy.