*Purpose: We have reported that CXCR5⁺CD8⁺T_Ab-supp cells mediate suppression of alloantibody (alloAb) production and prolong graft survival in both hepatocellular and kidney transplant (KTx) mouse models. Understanding the mechanisms by which CD8⁺ T_Ab-supp cells and conventional pharmacotherapy interact is critical for determining the efficacy of this targeted immunotherapy against alloantibody (alloAb) production and AMR following transplant.

*Methods: We investigated the efficacy of GFP⁺CXCR5⁺CD8⁺T cell adoptive cell transfer (ACT) in modulating alloAb production after allogeneic KTx (A/J, H-2^a) into CCR5 KO (H-2^b) recipients under cover of conventional immunosuppressants (mTORi or calcineurin inhibitors, CNi) for 14 days following KTx. AlloAb titer was measured and immune cell subsets were quantified and phenotyped by flow cytometry on day 14 posttransplant.

*Results: ACT of CXCR5⁺CD8⁺T cells to CCR5 KO recipients on day 1 after KTx (D1) significantly suppresses in vivo alloAb titer (1258.7±113.8) compared to untreated recipients (5951.1±629.8; p=0.0001), while pre-KTx ACT (D-1) does not (4848.2±947.3, alloAb titer comparable to untreated mice; p=ns). Administration of mTORi significantly inhibits day 14 alloAb titer (495±76.9; p<0.0001), while CNi has no impact on alloAb production (6473±434.3; p=ns). ACT combined with mTORi significantly reduces day 14 post-KTx alloAb titer (387±62.8 vs 1259±114; p=0.0007 for ACT alone). Conversely, CNi treatment partially negates the beneficial effects of ACT on post-KTx alloAb titer (2385±113.8, p=0.03 vs ACT alone). Reduction in post-KTx alloAb by ACT was accompanied by reduced quantity of splenic germinal center (GC; GL-7⁺Fas⁺B220⁺) B cells compared to untreated CCR5 KO recipients (4037±515 vs 5815±655 cells/mm³; p=0.03). Addition of mTORi to ACT preserves ACT-mediated suppression of GC B cells (4498±940 cells/mm³), while CNi negates ACT-associated suppression of GC B cells (6130±834 cells/mm³; p=0.04). Analysis of persisting GFP⁺CXCR5⁺CD8⁺ T cells 14 days after ACT shows that concurrent treatment with mTORi is associated with comparable quantity (1671±139 cells/mm³; p=ns), while CNi is associated with reduced quantity (900±83 cells/mm³; p=.01) of transferred GFP⁺CXCR5⁺CD8⁺ T cells detected in the spleen compared to ACT alone (1601±116 cells/mm³).

*Conclusions: ACT-mediated suppression of alloAb titer after KTx in mice is effective only when administered posttransplant and is associated with a reduction in the quantity of splenic GC B cells. mTORi but not CNi optimizes the efficacy of ACT.

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