INDUCTION OF DONOR SPECIFIC ACQUIRED THYMIC TOLERANCE BY INTRATHYMIC INOCULATION OF A SINGLE SYNTHETIC ALLO-MHC CLASS I PEPTIDE [RT1.Au

Abstract

499 This study extends the finding that intrathymic (IT) injection of closely related WAG (RT1u) class I peptides induces transplant tolerance to WF(RT1u) to the use of well defined, strain specific WF MHC class I peptides [RT1.Au]. It is reasoned that the effectiveness of TCR-MHC / peptide interaction which controls T cell ontogeny may be influenced by the presence of allopeptides in the thymus. Seven peptides were synthesized from RT1.Au sequences as published by Walter et al (1995) and Joly et al(1995), respectively. Following in-vivo immunization, we determined that while two peptides [U-5 & U-7] were immunogenic, 5 others [U-1, U-2, U-3, U-4,& U-6] were not immunogenic in ACI recipients. We then examined the effects of IT injection of a combination or single doses of the immunogenic RT1.Au peptides on cardiac allograft survival in the WF-to-ACI rat combination. The results showed that a combination of equal amounts [150 μg or 300 μg each] of the two peptides combined with 0.5 ml ALS on day -7 relative to cardiac transplantation led to 60 & 100% permanent WF graft survival (>100 days) respectively in ACI recipients. We then studied the effects of the individual peptides on graft survival. While 150 μg U-5 injected IT on day -7 did not prolong graft survival, 300 μg U-5 significantly prolonged graft survival to MST of 17.3 ± 3.6 days from 9.8 days in naive recipients. In contrast, IT injection of 150, 300, or 600μg U-7 with or without ALS on day -7 did not affect graft rejection. On the other hand, IT injection of 150, 300, and 600 μg U-5 combined with 0.5 ml ALS on day -7 led to permanent graft survival (>100 days) in 4/6, 6/6, and 4/4 ACI recipients, respectively, compared to an MST of 15.4 days in ALS alone treated controls; thus demonstrating that peptide U-5 alone mediates the observed effects on graft prolongation. Similar treatment with 300 μg U-5 combined with ALS led to acute rejection of third party (Lewis) grafts. Intravenous injection of 300 μg U-5 with ALS also did not prolong WF graft survival in ACI recipients. The long-term unresponsive ACI recipients challenged with 2nd-set allografts accepted permanently donor-type (WF) but not 3rd party (Lewis) cardiac allografts. In conclusion, we have demonstrated that a short segment of RT1.Au represents the tolerogenic epitope that induces acquired systemic tolerance. These studies have relevant clinical implications since only a single peptide can be potentially used in future large animal and human studies.