Abstract
Background:
Graft-versus-host disease (GVHD) is a severe complication of allogeneic stem-cell transplantation (allo-HSCT). Previously, we and others showed that activation of type I interferon (IFN-I) inducing pathways, such as cGAS/STING, can promote intestinal barrier integrity and limit GVHD. In contrast, there is limited knowledge regarding whether inhibiting the STING pathway can help prevent GVHD and about the optimal timing for such inhibition.
Methods:
The covalent STING inhibitor H-151 was evaluated at various time points in preclinical GvHD and GvL mouse models. For mechanistic studies, murine intestinal damage models were used and outcomes assessed using an organoid recovery assay in addition to established read-outs (histology, immune infiltrate). In vitro organoid assays were used to assess the impact of STING inhibition on epithelial regeneration after genotoxic stress from irradiation or chemotherapy. These assays were expanded to include co-cultures with allogeneic T cells to assess immune-modulatory effects. Human immune cells were treated with H151 in vitro and subsequently analyzed using flow cytometry and single-cell RNA sequencing (scRNA-seq). Ongoing scRNA-seq of epithelial and immune cells after transplantation in mice seeks to further unravel the underlying mechanisms and will be shared.
Results:
In preclinical models of GvHD, administering H151 before irradiation, but not at later time points, significantly decreased GvHD severity and enhanced survival. Recipients treated with H151 prior to conditioning showed a decreased loss of intestinal stem cells and reduced epithelial apoptosis after conditioning. Consequently, we observed enhanced intestinal regeneration, as demonstrated by improved organoid formation ex vivo. In vitro, H-151 treatment improved the resistance of organoids to genotoxic stress induced by irradiation or chemotherapy. Co-culture assays indicated that transient STING inhibition did not affect resistance to T cell-mediated damage, though prolonged inhibition impaired organoid growth, highlighting STING's role in intestinal homeostasis and regeneration.
In immune cell assays, H151 minimally affected T cell activation but reduced APC activation, suggesting that STING inhibition modulates inflammatory signals without compromising T cell function. Data from GvL models showed that transient STING inhibition did not impair the GvL effect. Ongoing scRNA-seq experiments aim to clarify the cellular mechanisms driving these outcomes, focusing on epithelial cells, T cells, and APCs following allo-BMT.
Conclusions:
Our findings show that transient modulation of the STING pathway before conditioning protects the intestinal epithelium and reduces GvHD without affecting the GvL effect. While STING activity is harmful during genotoxic damage, it is crucial for maintaining intestinal homeostasis and regeneration later. This study underscores the importance of timing in STING modulation and its potential as a therapeutic strategy to improve allo-HSCT outcomes.
