Reversing TCMR through inhibition of innate immune allorecognition

Introduction: A significant fraction of acute T cell-mediated rejection (TCMR) episodes are inadequately controlled by standard treatments, which markedly impede long-term graft survival. Given that TCMR is driven by innate allorecognition memory (Allo-IMem), we investigated whether specifically inhibiting Allo-IMem using targeted nanobiologics reverses established TCMR.
Methods: Wild-type C57BL/6 recipients of Balb/c islet allografts were treated at the peak of TCMR (days 7–12 post-transplant) with innate immune cell-specific high-density lipoprotein nanobiologics. These nanobiologics were either unloaded (control; U-NB) or loaded with the mTOR inhibitor rapamycin (mTORi-NB; 5 mg/kg). Outcomes were compared against treatment with free rapamycin (Rapa; 5 mg/kg).
Results: We found that Allo-IMem signals through the mTOR pathway in innate immune cells in vivo and that these memory responses are effectively inhibited by mTORi-NB, whereas free Rapa treatment is ineffective. By experimentally segregating Allo-IMem and skin allograft TCMR temporally, we revealed that: 1) established Allo-IMem significantly enhances subsequent acute TCMR; and 2) specific inhibition of Allo-IMem with mTORi-NB, in the complete absence of adaptive immune cells, drastically reduces subsequent TCMR and significantly enhances allograft survival compared to U-NB or Rapa. Remarkably, a short course of mTORi-NB initiated during acute TCMR (days 7–12) reversed or significantly delayed islet allograft rejection in 90% of recipients. Furthermore, mTORi-NB treatment during ongoing TCMR was therapeutically superior to Rapa in promoting the survival of heart allografts, and of islet allografts in a stringent model where recipients were reconstituted with donor-specific memory T cells prior to transplantation (Fig. 1). Single-cell RNA sequencing (scRNA-seq) of graft-infiltrating innate cells (day 10) demonstrated that mTORi-NB inhibited the cell cycle, glycolysis, and type I/II interferon signaling, while promoting TGF-β signaling pathways. Multiphoton intravital microscopy of islet allografts on day 10 revealed that mTORi-NB significantly reduced both the dynamics (speed and displacement) and intracellular Ca²⁺ signaling in effector T cells. Mechanistically, mTORi-NB drastically reduced the antigen-presentation capacity of graft-infiltrating innate cells, leading to a significant reduction in the proliferation and numbers of graft-infiltrating effector T cells. 

Conclusions: Our data demonstrate that Allo-IMem responses signal through mTOR in innate immune cells and potently drive TCMR. Consequently, inhibiting Allo-IMem using innate immune cell-specific nanobiologics effectively overturns established adaptive immune responses and reverses allograft rejection. This provides a novel, innate immune-targeted therapeutic approach for the treatment of acute TCMR in transplant recipients.