Tu185 - Combining Human CD4+ and CD8+ Treg to Inhibit Experimental Acute GVHD

Sonia Salle – Nantes Université, INSERM, Center for Research in Transplantation and Translational Immunology, UMR 1064, F-44000, Nantes, France; Céline Sérazin – Nantes Université, INSERM, Center for Research in Transplantation and Translational Immunology, UMR 1064, F-44000, Nantes, France; Juliette Lasselin – Nantes Université, INSERM, Center for Research in Transplantation and Translational Immunology, UMR 1064, F-44000, Nantes, France; Ignacio Anegon – Nantes Université, INSERM, Center for Research in Transplantation and Translational Immunology, UMR 1064, F-44000, Nantes, France; Carole Guillonneau – Nantes Université, INSERM, Center for Research in Transplantation and Translational Immunology, UMR 1064, F-44000, Nantes, France

Abstract Text: Combining CD4+Tregs and CD8+Tregs for cell therapy would increase the number of starting Tregs, reducing the volume of blood drawn from patients; would simplify cell isolation, saving time and costing less; and could be more efficient by recognizing different antigens. Here, we explored the differences and benefits of combining them.
Blood CD4+Tregs compared to CD8+Tregs displayed significantly lower methylation in the FOXP3-TSDR, 253 genes DE, and higher efficiency in suppressing CD4+ Tconv responses in vitro. In contrast, after overnight polyclonal activation, transcriptomic comparison showed no DE gene between subsets, similar efficacy in vitro, and their combination resulted in more efficient suppression than each individually. After separate expansion under pro-regulatory conditions, the combination of a half dose of CD4+Tregs and CD8+ Tregs in a model of humanized acute GVHD in NSG mice resulted in therapeutic effect similar to that of a full dose of CD4+ or CD8+ Tregs, and their addition improved long term mouse survival to 66%. To enrich for both FOXP3+ CD4+ and CD8+ Treg, we isolated T cells based on the CD45RClow/- expression marker and expanded them under pro-regulatory conditions. CD3+CD45RClow/- cells displayed similar suppressor efficacy to isolated CD4+ or CD8+Tregs both in vitro and in the GVHD NSG mouse model, without inducing any toxicity, when using a 1.5x doses, which should be largely offset by the production yield.
CD4+ and CD8+Tregs show similarities and their complementarity for Tconv control and it is possible to isolate them from blood using only the CD45RClow/neg marker and appropriate culture conditions.