Targeting the Transcription Factor Id2 Reveals Novel Regulatory Mechanisms in Chronic Graft-versus-Host Disease 2257336

Abstract Introduction Chronic graft-versus-host disease (cGVHD) is a severe complication of hematopoietic stem cell transplantation (HSCT) complication driven by immune dysregulation. A key feature is aberrant T follicular helper (Tfh) and germinal center B (GCB) cells, which promote pro-inflammatory cytokine release, pathogenic antibody production, and tissue fibrosis. Leveraging cGVHD patient T cell single-cell RNA sequencing data from the Kean Lab, we selected Id2, a transcription factor that inhibits Tfh differentiation while supporting antibody production, as a target for the treatment of cGvHD. Methods We used CRISPR/Cas9 to generate Id2-knockout (KO) naïve murine T cells and adoptively transferred into pre-conditioned B10.BR recipients of C57BL/6 bone marrow (BM) ± T cells to generate murine cGVHD model with bronchiolitis obliterans (BO) lung disease. Results Id2-KO T cells maintained ≥90% KO efficiency until study end (d49). Mice receiving Id2-KO vs control T cells had significantly improved pulmonary function (p < 0.0001), normalizing resistance, compliance, and elastance to BM-only (no disease) levels. Consistent with cGVHD amelioration, a 2-fold reduction in lung collagen deposition (p < 0.0001) and 4-fold decrease in serum allo-reactive antibody (p = 0.0286) were noted. On D49, despite similar GCB and Tfh ratios to control, Id2-KO T cells had enhanced Tfh differentiation: Bcl6+T cells increased from 4.9% to 10.8% (p = 0.0038) with 1.8-fold higher Bcl6 MFI(p = 0.0005). Id2-KO Tfh cells were functionally impaired with a 50% reduction in IL-4 production (p = 0.0078) and a 1.9-fold decrease in SLAMF1+ Tfh cells (from 63.3% to 33%, p < 0.0001), indicating disrupted Tfh-GC B cell interactions. Conclusion Id2 restrains Tfh differentiation while supporting their functional capacity to help GCB cells produce pathogenic antibody. Genetic Id2 loss uncouples Tfh differentiation from effector activity, positioning Id2 inhibition as a novel therapeutic strategy to selectively disrupt pathogenic Tfh-GCB crosstalk in cGVHD Funding Source NIH Topic Categories Transplantation Immunology (TRAN)