Post Transplantation Bilirubin Nanoparticles Ameliorate Murine Graft Versus Host Disease via a Reduction of Systemic and Local Inflammation

Abstract

Allogeneic stem cell transplantation is a curative immunotherapy where patients receive myeloablative chemotherapy and/or radiotherapy, followed by donor stem cell transplantation. Graft versus host disease (GVHD) is a major complication caused by dysregulated donor immune system, thus a novel strategy to modulate donor immunity is needed to mitigate GVHD. Tissue damage by conditioning regimen is thought to initiate the inflammatory milieu that recruits various donor immune cells for cross-priming of donor T cells against alloantigen and eventually promote strong Th1 cytokine storm escalating further tissue damage. Bilirubin nanoparticles (BRNP) are water-soluble conjugated of bilirubin and polyethylene glycol (PEG) with potent anti-inflammatory properties through its ability to scavenge reactive oxygen species generated at the site of inflammation. Here, we evaluated whether BRNP treatment post-transplantation can reduce initial inflammation and subsequently prevent GVHD in a major histocompatibility (MHC) mismatched murine GVHD model. After myeloablative irradiation, BALB/c mice received bone marrow and splenocytes isolated from C57BL/6 mice, with or without BRNP (10 mg/kg) daily on days 0 through 4 post-transplantation, and clinical GVHD and survival was monitored for 90 days. First, BRNP treatment significantly improved clinical GVHD score compared to untreated mice (3.4 vs 0.3, p=0.0003), and this translated into better overall survival (HR 0.0638, p=0.0003). Further, BRNPs showed a preferential accumulation in GVHD target organs leading to a reduced systemic and local inflammation evidenced by lower pathologic GVHD severity as well as circulating inflammatory cytokines such as IFN-gamma. Lastly, BRNP treatment post-transplantation facilitated the reconstitution of CD4(+) iNK T cells and reduced expansion of proinflammatory CD8 alpha(+) iNK T cells and neutrophils especially in GVHD organs. Lastly, BRNP treatment decreased ICOS+ or CTLA-4(+) T cells but not PD-1(+) T cells suggesting a decreased level of T cell activation but maintaining T cell tolerance. In conclusion, we demonstrated that BRNP treatment post-transplantation ameliorates murine GVHD via diminishing the initial tissue damage and subsequent inflammatory responses from immune subsets.