PROLONGATION OF HEART XENOGRAFT SURVIVAL IN A CONCORDANT HAMSTER TO RAT MODEL FOLLOWING THERAPY WITH A RATIONALLY DESIGNED IMMUNOSUPPRESSIVE PEPTIDE

Abstract

83 Peptide derived from the HLA class I heavy chain were shown to modulate immune response in vitro and in vivo in a non allele restricted fashion. The immunomodulatory effect of these peptides correlated with the peptide mediated inhibition of heme oxygenase 1 (HO-1), also called HSP32. By computer based rational design, we developed novel peptides with enhanced immunosuppressive activity, more potent inhibitors of HO-1 in vitro. One peptides, RDP1258, was assessed for its ability to prolong heart graft survival in the concordant Hamster to LEW.1A rat model. RDP1258 was given IP on day -7 and -1 before transplantation (at 1 mg/kg) alone or with Cobra Venom Factor (CVF) given IM from day-1 for ten days (at 0.125mg/kg).Methods: Xenograft survival was studied using a Kaplan Meyer analysis. Grafts from recipients treated with CVF or with CVF/RDP1258 were studied for leukocyte infiltrate (Point counting technique), HO-1 staining by immunohistochemistry, and for TNFα and iNOS mRNA by a competitive RT-PCR method. Results: Peptide or CVF monotherapy had no effect on graft survival. However, combination of CVF/RDP1258 resulted in a significant prolongation of graft survival (7.9±1 vs 4.5±0 days, p<0.001). Immunohistochemical analysis of graft infiltrating leukocytes on day 3 after transplantation showed a trend for a decrease in the number of cells in CVF/RDP1258 treated recipients compared to control animals treated with CVF monotherapy (1.7 vs 3.3%, p<0.07). In addition, RDP1258 administration resulted in enhanced expression of HO-1 in spleen compared to controls. This is consistent with observations made with other inhibitors of HO-1, inducing an up-regulation of HO-1 expression in vivo. In other combination (ACI rats), measurement of activity in spleen cell extracts, after RDP1258 treatment, confirmed an up-regulation of HO-1 production associated with an increased heme oxygenase enzyme activity. Interestingly, xenografts of RDP1258 treated animals displayed a two time lower number of TNFα mRNA (p<0.07) compared to controls treated with CVF monotherapy (1.73±0.31 vs 3.50±0.61 copies/HPRT). The iNOS mRNA level was not changed (0.32±0.08 vs 0.32±0.1 copies/HPRT).Conclusions: Our results demonstrated prolongation of xenograft survival following therapy with immunosuppressive peptide, associated with elevated expression of HO-1 and decreased production of TNFα mRNA. These observations are consistent with recent observation that up-regulation of HO-1 resulted in the inhibition of several immune effector functions.