SYNERGISTIC EFFECTS OF CYTOKINE GENE TRANSFER IN HIGH RESPONDER RAT RENAL ALLOGRAFT RECIPIENTS

Abstract

112 Gene therapy approach has the potential to introduce immunosuppressive molecules directly into the allograft (Tx), which should limit systemic side effects of conventional therapy. Although there is evidence that transfer of Th2-type cytokines may inhibit rejection in some rodent Tx models, numerous counter examples also exist. Recently, we have shown that ca. 60% of LEW rat recipients of LBNF1 kidney Tx survived >40 days following selective adenoviral (Ad)-mediated IL-4 gene transfer. This study analyzes the efficacy of cytokine gene transfer in high responder (WF to LEW) rat renal Tx model. Allogeneic renal Tx were performed between WF donors and LEW recipients. Donor kidneys were perfused in-situ with Ad-IL-4 [2.5×109 pfu] alone or with Ad-IL-10/Ad-IL-12(p40) [2×1010 pfu], stored at 4C for 1 1/2 h, and transplanted. Some recipients of kidney Tx transfected with Ad-IL-4 were treated concomitantly with low dose of CsA (1.5mg/kg/day ×10 days s.c.). All animals underwent contralateral native nephrectomy at day 5. Serum creatinine levels were measured, and Tx rejection was determined by recipient death. Intra-Tx cytokine gene expression was screened by competitive template RT-PCR. (Table)Table: depicts LEW rat survival after cytokine gene transfer into WF renal Tx. Serum creatinine levels (mg/dl) at day 10 were at average of 6.4, 4.4 and 7.1 in Ad-IL-4, Ad-IL-4/IL-10 and Ad-IL-4/IL-12(p40) groups, respectively, vs. 6.4 in untreated controls. By day 30, creatinine levels decreased to 2.5 mg/dl, and 2.7 mg/dl in Ad-IL-4/IL-10 and Ad-IL-4/IL-12 (p40) groups, respectively. In contrast, creatinine levels remained diminished throughout in Ad-IL-4/CsA group (ca. 1.9 mg/dl). Compared to Ad-IL-4-treated animals, elevated levels of IL-4 mRNA could be detected by RT-PCR in Ad-IL-4/IL-10 but not in Ad-IL-4/IL-12(p40) group. Intra-Tx IL-10 mRNA was increased in Ad-IL-4/IL-10 and Ad-IL-4/IL-12(p40) treated rats but not in those transfected with Ad-IL-4 alone. Significant CD3 T cell infiltration and IFN-γ expression were readily detectable despite long-term renal Tx acceptance in most of Ad-transfected recipients. This is the first report that documents synergistic effects of Th2-type/Th1-inhibitor cytokine gene transfer in renal Tx recipients. Unlike in low-responder model, co-application of Ad-IL4+Ad-IL-10/IL-12(p40) or Ad-IL-4+CsA were required to achieve long-term therapeutic effects in high-responder rat combination. Moreover, only adjunctive Ad-IL-4 + low dose CsA regimen afforded considerable protection from post-Tx uremia and early rejection episodes.