Diabetes and other autoimmune endocrine diseases
CD8+ effector T cells largely contribute in the destruction of human pancreatic beta cells suggesting that MHC class I is abundant in the inflamed islets and can be employed to redirect regulatory T cells (Tregs) specificities to islets.
We developed novel methods allowing: (1) efficient Tregs expansion in which the endogenous TCR is removed using a non-viral CRISPR-based approach combined with lentiviral transduction for the insertion of an engineered TCR, (2) non-viral substitution of the CD4 co-receptor with CD8α chain, (3) human islets transplantation into the spleen of NSG mice to increase the interaction with transferred human T cells.
Nine day after electroporation, mean efficiency of TCR knockout (KO) in Tregs was 74% for TRAC (n=3) and 79% for TRBC (n=5), which significantly impaired TCR-stimulated suppression of Tregs in vitro. An HLA-A2 restricted TCR specific for preproinsulin (PPI15-24) was cloned into lentivirus and transduced in TCRKO T cells restoring up to 100% TCR expression. Importantly PPI15-24 tetramer staining was detected only on CD8 T cells demonstrating that antigen engagement by this TCR is CD8 co-receptor dependent. Using a non-viral approach, we inserted the CD8α chain into the CD4 locus with up to 25% efficiency. Finally, we transplanted 4000 human islets equivalent into spleens of NSG mice which reverted streptozotocin-induced diabetes in 6/9 mice up to 100 days.
Altogether, we generated the tools to engineer Tregs with HLA-class I restricted specificity. Preclinical testing of Tregs suppression in humanized mouse model for type 1 diabetes is currently under investigation