Newly Designed Alginate-Based Microcapsules (AgMc) for the Molecular Therapy of Type 1 Diabetes

Abstract

Glucocorticoid-induced tumor necrosis factor receptor-related protein (GITR) plays a crucial role in either thymic t-Treg differentiation and expansion, or peripheral p-Treg expansion, thus representing a potential tool to contrast the autoimmune process. AgMc are usually intended to constitute an immunobarrier to prevent rejection of the embodied islet/insulin producing cell grafts in T1D. We aimed to reverse this principle, and engineered the AgMc membranes, in order to make them permeable to outflowing rat anti-mouse GITR IgM MAb, derived from microencapsulated rat G3C hybridoma cells. To this end, we changed our AgMc original formulation to obtain thicker and looser membranes. We observed intra-AgMc IgM retention and their progressive release into the culture medium in vitro. Prediabetic NOD mice were divided into two groups: (1) n=20, received graft (TX) of G3C containing microcapsules; (2) n=10 received TX of empty microcapsules. Both (1) and (2) were monitored for the onset of DM. BG in basal and after IPGTT, morphological, immuno-molecular, and cell immuno-phenotyping testing, as well as G3C plasma levels assay in (1) were performed. In (2): 5/10 animals developed DM within 70d; in (1) no animals developed DM at 180 of TX. In the spleen of G3C-AgMc-treated mice either CD4+FoxP3+GITRbright, or CD4+CD25-GITR(sp), or CD8+CD25+GITRlowFoxP3+, or CD8+CD25-GITRintFoxP3+ Tregs significantly increased from 4 week of TX as compared to controls. Pancreatic histology at 26 week of TX was associated with either completely destroyed islets (1) or intact islets with minor insulitis (2). Significant diabetes-free time of (1) vs. (2) may relate to increase of islet B-cell specific Tregs, expanded by G3C-released MAb, and antigenic stimulation, at early stage of the autoimmune disease process. In conclusion, the newly engineered AgMc, by permitting GITR triggering by anti-GITR IgM, may represent a new approach to expand Tregs and inhibit Teff, and to avert the onset of autoimmune DM in NOD mice. Disclosure R. Calafiore: None. P. Montanucci: None. G.P. Basta: None. G. Nocentini: None. T. Pescara: None. C. Riccardi: None.