mTOR inhibitors [mTORi; Rapamycin, (Rapa)] and calcineurin inhibitors [CNi; Tacrolimus (Tac)] are immunosuppressives commonly used to suppress rejection of transplants. Despite widespread use following transplantation (Tx), effects of these therapies on alloantibody (alloAb) production postTx is unknown. To determine effects of mTORi and CNi on alloAb development, wild-type (C57BL/6, WT) mice received hepatocyte Tx (FVB/N donor). Select recipients were CD8-depleted (day -2, -1) and/or treated with Rapa or Tac (1-5 mg/kg/d, days 1-14). Graft survival was assessed and alloAb analyzed postTx by percent allotarget binding. In our model, WT recipients reject by CD8⁺CTL and alloAb-mediated mechanisms. CD8-depleted recipients are high alloAb producers and rejection is alloAb-dependent. We found that mTORi (6.3 ±1.5%) resulted in greater inhibition of alloAb development in vivo than CNi (20.3 ±2.5%) in low alloAb producing recipients. mTORi significantly suppressed alloAb production in CD8-depleted recipients (from 60.8% to <3.5% for all doses). In contrast, while CNi decreased alloAb production in a dose dependent manner, no dose inhibited alloAb to the degree as mTORi. mTORi prolonged graft survival in both WT (low alloAb producers, MST=d21) and CD8-depleted (high alloAb producers, MST=d27) recipients compared to controls (MST=d14). CNi prolonged graft survival only in WT recipients (MST=d27). Given the significant effect of mTORi on alloAb development, we were interested in the effects of mTORi on primed B cells in vivo. Alloprimed IgG1⁺ B cells were isolated from Tx recipients on day 5 and adoptively transferred (AT) into Rag1 KO mice. Rag1 KO mice AT with primed B cells were given exogenous “help” (αCD40 mAb, IL-4) and alloantigen (FVB/N splenocytes), followed by treatment with Rapa or Tac (1 mg/kg/d, days 1-7). Rag1 KO mice AT with naive B cells and similarly treated were controls. AT of alloprimed IgG1⁺ B cells (but not naïve B cells) into Rag1 KO mice resulted in high levels of alloAb on day 7 post-AT (73.09 ±7.3%). Treatment with mTORi resulted in dramatic suppression of alloAb from primed B cells (11.39 ±5.43%). These results demonstrate that mTORi is more effective than CNi for suppression of postTx alloAb production and primed B cell function in vivo. Furthermore, mTORi suppression of alloAb production in both low and high-alloAb producers is accompanied by significantly prolonged allograft survival.

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