Person:

Faustman, Denise

Background: No targeted immunotherapies reverse type 1 diabetes in humans. However, in a rodent model of type 1 diabetes, Bacillus Calmette-Guerin (BCG) reverses disease by restoring insulin secretion. Specifically, it stimulates innate immunity by inducing the host to produce tumor necrosis factor (TNF), which, in turn, kills disease-causing autoimmune cells and restores pancreatic beta-cell function through regeneration. Methodology/Principal Findings Translating these findings to humans, we administered BCG, a generic vaccine, in a proof-of-principle, double-blind, placebo-controlled trial of adults with long-term type 1 diabetes (mean: 15.3 years) at one clinical center in North America. Six subjects were randomly assigned to BCG or placebo and compared to self, healthy paired controls (n = 6) or reference subjects with (n = 57) or without (n = 16) type 1 diabetes, depending upon the outcome measure. We monitored weekly blood samples for 20 weeks for insulin-autoreactive T cells, regulatory T cells (Tregs), glutamic acid decarboxylase (GAD) and other autoantibodies, and C-peptide, a marker of insulin secretion. BCG-treated patients and one placebo-treated patient who, after enrollment, unexpectedly developed acute Epstein-Barr virus infection, a known TNF inducer, exclusively showed increases in dead insulin-autoreactive T cells and induction of Tregs. C-peptide levels (pmol/L) significantly rose transiently in two BCG-treated subjects (means: 3.49 pmol/L [95% CI 2.95–3.8], 2.57 [95% CI 1.65–3.49]) and the EBV-infected subject (3.16 [95% CI 2.54–3.69]) vs.1.65 [95% CI 1.55–3.2] in reference diabetic subjects. BCG-treated subjects each had more than 50% of their C-peptide values above the 95th percentile of the reference subjects. The EBV-infected subject had 18% of C-peptide values above this level. Conclusions/Significance: We conclude that BCG treatment or EBV infection transiently modified the autoimmunity that underlies type 1 diabetes by stimulating the host innate immune response. This suggests that BCG or other stimulators of host innate immunity may have value in the treatment of long-term diabetes. Trial Registration ClinicalTrials.gov NCT00607230

Background: Soluble TNF superfamily (TNFSF) ligands are less stable and less active than their transmembrane (tm) analogues. This is a problem for the therapeutic use of recombinant TNFSF ligands in diverse diseases including cancer and autoimmunity. Creating TNFSF ligand analogues with improved targeting of their respective receptors is important for research and therapeutic purposes. Findings: Covalent internal cross-linking of TNF monomers by double mutations, S95C/G148C, results in stable trimers with improved TNFR2 function. The resulting mutein induced the selective death of autoreactive CD8 T cells in type-1 diabetic patients and demonstrates targeted proliferation and expansion of human CD4 Tregs. Conclusions: Stable TNF trimers, created by internal covalent cross-linking, show improved signaling. The high structural homology within the TNF superfamily provides an opportunity to extend internal cross-linking to other TNF superfamily proteins to produce active trimers with improved stability and receptor signaling, and with potential applications for cancer, autoimmunity, infections, and transplantation. Electronic supplementary material The online version of this article (doi:10.1186/s40591-015-0044-4) contains supplementary material, which is available to authorized users.

T-regulatory cells (Tregs) are a rare lymphocyte subtype that shows promise for treating infectious disease, allergy, graft-versus-host disease, autoimmunity, and asthma. Clinical applications of Tregs have not been fully realized because standard methods of expansion ex vivo produce heterogeneous progeny consisting of mixed populations of CD4 + T cells. Heterogeneous progeny are risky for human clinical trials and face significant regulatory hurdles. With the goal of producing homogeneous Tregs, we developed a novel expansion protocol targeting tumor necrosis factor receptors (TNFR) on Tregs. In in vitro studies, a TNFR2 agonist was found superior to standard methods in proliferating human Tregs into a phenotypically homogeneous population consisting of 14 cell surface markers. The TNFR2 agonist-expanded Tregs also were functionally superior in suppressing a key Treg target cell, cytotoxic T-lymphocytes. Targeting the TNFR2 receptor during ex vivo expansion is a new means for producing homogeneous and potent human Tregs for clinical opportunities.

Activated T-regulatory cells (aTregs) prevent or halt various forms of autoimmunity. We show that type 1 diabetics (T1D) have a Treg activation defect through an increase in resting Tregs (rTregs, CD4+CD25+Foxp3+CD45RA) and decrease in aTregs (CD4+CD25+Foxp3+CD45RO) (n= 55 T1D, n=45 controls, P=0.01). The activation defect persists life long in T1D subjects (T1D=45, controls=45, P=0.01, P=0.04). Lower numbers of aTregs had clinical significance because they were associated with a trend for less residual C-peptide secretion from the pancreas (P=0.08), and poorer HbA1C control (P=0.03). In humans, the tumor necrosis factor receptor 2 (TNFR2) is obligatory for Treg induction, maintenance and expansion of aTregs. TNFR2 agonism is a method for stimulating Treg conversion from resting to activated. Using two separate in vitro expansion protocols, TNFR2 agonism corrected the T1D activation defect by triggering conversion of rTregs into aTregs (n=54 T1D, P<0.001). TNFR2 agonism was superior to standard protocols and TNF in proliferating Tregs. In T1D, TNFR2 agonist-expanded Tregs were homogeneous and functionally potent by virtue of suppressing autologous cytotoxic T cells in a dose-dependent manner comparable to controls. Targeting the TNFR2 receptor for Treg expansion in vitro demonstrates a means to correct the activation defect in T1D.