Characterization of recombinant class I MHC alloantigen action upon the survival of disparate heart allografts

Previous studies have led to the development of allochimeric class I MHC proteins as agents that effectively induce donor-specific transplantation tolerance in a rat system with or without additional immunosuppression. Within the α1-helical region of RT1.Au, an epitope that conferred immunologic tolerance was discovered. Studies presented herein were designed to test our central hypothesis that allochimeric proteins onfer tolerance in a mouse model. To test this hypothesis, portal vein (PV) injection of wild-type H2Kd and H2Dd proteins were produced in a bacterial expression system and found to specifically prolong the survival of BALB/c (H2d) heart allografts in C57BL/10 (H2b) recipients. Although a single PV injection of 50 μg α1–α 3 H2Kd alone was ineffective, 50 μg α1 –α3 alone slightly prolonged BALB/c heart allograft survivals. In contrast, the combination of 25 μg α1–α 3 H2Kd and 25 μg α1–α 3 H2Dd proteins prolonged BALB/c graft survivals to 20.2 ± 6.4 days (p < 0.004). The effect was donor-specific, since a combination of 25 μg α1–α3 H2Kd and 25 μg α1–α3 H2Dd proteins failed to affect survivals of third-party C3H (H2k k) heart allografts, namely 9.0 ± 0.0 days in treated versus 7.8 ± 0.5 days in untreated hosts. Thus, the combination of two H2K d and H2Dd proteins is more effective in prolonging allograft survival than a single protein produced in a bacterial expression system. A single PV injection (day 0) of 25 μg α1–α 2 H2Kd and 25 μg α1–α 2 H2Dd proteins to C57BL/10 mice prolonged the survival of BALB/c heart allografts to 22.4 ± 4.5 days. Within a WF to ACI rat heart allograft system, a single PV injection of 20 μg 70–77 u-RT1.Aa induced specific tolerance of allografts. This therapy could be combined with CsA to induce transplantation tolerance. However, combination of 70–77u-RT1.Aa with CTLA4Ig, rapamycin, or AG-490 effectively blocked the induction of transplantation tolerance. Tolerance generated by allochimeric protein could be adoptively transferred to naive recipients. Intragraft cytokine mRNA levels showed a bias towards a Th2-type phenotype. Additionally, studies of cytokine signaling and activation of transcription factors revealed a requirement that these pathways remain available for signaling in order for transplantation tolerance to occur. These studies suggest that the generation of regulatory cells are required for the induction of transplantation tolerance through the use of allochimeric proteins. ^

Treatment of solid organ transplant recipients with autologous Epstein Barr virus-specific cytotoxic T lymphocytes (CTLs).

We have investigated the in vivo safety, efficacy, and persistence of autologous Epstein Barr virus (EBV)-specific cytotoxic T lymphocytes (CTLs) for the treatment of solid organ transplant (SOT) recipients at high risk for EBV-associated posttransplantation lymphoproliferative disease (PTLD). EBV-CTLs generated from 35 patients expanded with normal kinetics contained both CD8 and CD4 lymphocytes and produced significant specific killing of autologous EBV-transformed B lymphoblastoid cell lines (LCLs). Twelve SOT recipients at high risk for PTLD, or with active disease, received autologous CTL infusions without toxicity. Real-time polymerase chain reaction (PCR) monitoring of EBV-DNA showed a transient increase in plasma EBV-DNA suggestive of lysis of EBV-infected cells, although there was no consistent decrease in virus load in peripheral-blood mononuclear cells. Interferon-gamma enzyme-linked immunospot (ELISPOT) assay and tetramer analysis showed an increase in the frequency of EBV-responsive T cells, which returned to preinfusion levels after 2 to 6 months. None of the treated patients developed PTLD. One patient with liver PTLD showed a complete response, and one with ocular disease has had a partial response stable for over one year. These data are consistent with an expansion and persistence of adoptively transferred EBV-CTLs that is limited in the presence of continued immunosuppression but that nonetheless produces clinically useful antiviral activity.