Bispecific CD8 Treg modulators regulate a novel regulatory CD8 T cell network and eliminate pathogenic CD4 T cells in live cell co-culture system

Abstract

Abstract INTRODUCTION Others have described a subset of CD8 T cells (CD8 Treg) with immunosuppressive characteristics in inflammatory disease settings. CD8 Treg activation through canonical T cell receptors results in their oligoclonal expansion and perforin dependent elimination of pathogenic CD4 T cells. We have described the CD8 Treg network in Celiac patients and its potential to eliminate pathogenic CD4 T cells. Here we describe bispecific CD8 Treg modulators that activate CD8 Tregs, resulting in pathogenic CD4 T cell death. METHODS Novel bispecific CD8 Treg modulators were tested for target specificity by Octet and cell binding. The functional impact of CD8 Treg modulators was evaluated using flow and Luminex assays, and in a live cell co-culture system using Celiac patient derived CD8 Tregs and gliadin activated CD4 T cell targets. CD8 Treg modulators with specific molecular formats were then ranked for functional efficacy. RESULTS We tested a panel of bispecific CD8 Treg modulators with monovalent binding affinities in the low nanomolar range that selectively bind the CD8 Treg subset. Within 48 hours of CD8 Treg modulator addition to T cell co-cultures, CD8 Tregs had a rapid increase of cytolytic markers and degranulation in response to CD4 T cell targets. We observed a concomitant increase of activated CD4 T cell death and decrease of pro-inflammatory cytokines. CONCLUSIONS Our results indicate that a targeted approach to engage and activate CD8 Treg reduces pathogenic CD4 T cells. As this newly described CD8 Treg cell network is observed in many autoimmune diseases, we postulate that bispecific CD8 T cell modulators may represent a novel, selective and broadly applicable therapeutic approach for the treatment of human autoimmune diseases.