CD4-TARGETED THERAPY ABROGATES ACCELERATED REJECTION OF LIVER ALLOGRAFTS IN PRESENSITIZED RAT RECIPIENTS

Abstract

127 Allograft (Tx) rejection in sensitized patients represents one of the most challenging problems in clinical transplantation. We have developed a model of accelerated rejection in which LEW rats presensitized with BN skin Tx (day-7) reject LBNF1 cardiac Tx (transplanted at day 0) in <36 h. We have also shown that treatment with RIB-5/2, a nondepleting CD4 mAb, may induce tolerance in this sensitized Tx model. However, to achieve tolerogenic effect, animals require intermittent infusion of CD4 mAb in the "sensitization" phase, i.e. starting at the time of skin Tx; CD4-targeted therapy after cardiac engraftment is ineffective. This study was designed to establish a model of accelerated rejection of orthotopic liver Tx (OLT) and then to test the effects of RIB-5/2 mAb treatment in sensitized rat recipients. First, we have determined that normal LEW rats survived >100 days following transplantation of LBNF1 livers (Gr. I; n=6), similarly to isograft controls (n=3). Sensitization with BN skin Tx at day-7 prompted accelerated rejection of liver Tx, and resulted in animal death within 8.9±2.2 days (Gr.II; n=11). Then, in order to mimic a clinical scenario, we asked whether CD4-targeted therapy introduced in presensitized hosts at the time of liver engraftment may affect the accelerated rejection cascade. Accordingly, LEW rats bearing BN skin Tx for a week were treated by RIB-5/2 mAb immediately after OLT (7× 5mg/rat i.v. day 0 to 21). Indeed, all animals in this treatment group are currently surviving at >50 days post-Tx and maintain good liver function (Gr. III; n=6). Hence, by day 4, serum GOT levels (IU/L) in Gr. III recipients were 140±51 vs. 348±53 and 840±51 in Gr. I and Gr. II rats, resp. (p<0.01; n=3/group). By day 50, sGOT levels in the treatment group were 233±92 IU/L (n=3); no circulating RIB-5/2 mAb were detectable at 3 weeks after cessation of RIB-5/2 mAb therapy. Liver samples were collected from recipient groups at day 8 post-Tx for histological evaluation, and the degree of rejection was scored according to the Banff criteria by blinded pathologist. Unlike in isografts, which remained essentially normal, severe lobular disarray with diffuse necrosis and fibrin deposition in hepatic sinusoids, was noted in Gr. II sensitized rats, consistent with ferocious accelerated OLT rejection. In marked contrast, Gr. III liver Tx harvested from RIB-5/2 mAb treated presensitized hosts, showed only mild bile duct obstruction and proliferation, consistent with "mild" acute OLT rejection. Liver Tx harvested at day 8 from Gr. I unsensitized hosts showed signs of "moderate-severe" acute rejection. Finally, we analyzed host systemic IgM and IgG allo-Ab responses in sera samples by flow cytometry (n=3/group/time-point). No differences in IgM and IgG allo-Ab levels could be detected at day 7 post-Tx between animal groups. Interestingly, however, by day 28, RIB-5/2 mAb treatment depressed IgM (mean channel fluorescence±SD = 28±30 vs. 376±108 in Gr. I; p<0.01), and prevented the switch from IgM to IgG (0 vs. 435±27 in Gr. I; p<0.01). In conclusion, this study documents the efficacy of a clinically relevant nondepleting CD4 mAb regimen to abrogate accelerated rejection and induce long-term acceptance of liver Tx in sensitized rat recipients. Mab-facilitated depression of host cellular and humoral immunity may contribute to OLT survival in this stringent transplantation model.