INTRACORONARY GENE-TRANSFER OF IMMUNOSUPPRESSIVE CYTOKINES TO CARDIACALLOGRAFTS - METHOD AND EFFICACY OF ADENOVIRUS-MEDIATED TRANSDUCTION

Objective: Allograft-targeted immunosuppressive gene therapy may inhib it recipient immune activation and provide an alternative to systemic immunosuppression. We studied the optimal technique and efficacy of in tracoronary gene transfer of viral interleukin-10 and human transformi ng growth factor-beta(1) in a rabbit model of heterotopic heart transp lantation. Methods: Replication-defective adenoviral vectors were cons tructed, expressing viral interleukin-10 (AdSvIL10) or transforming gr owth factor-beta(1) (AdCMVTGF-beta(1)). Intracoronary delivery of vect ors was accomplished ex vivo by either bolus injection or slow infusio n, The allografts were implanted heterotopically in recipient rabbits and collected 4 days after the operation. Vector dose was 4 x 10(9) to 6 x 10(10) pfu/gm of donor heart. Transfer was confirmed by DNA ampli fication for both genes. Gene product expression in tissue was quantif ied by immunoassay and visualized by immunohistochemical staining. Res ults: Allograft viral uptake was only 9.9% +/- 2.4% with bolus injecti on, but increased to 80.5% +/- 6.8% at 1 ml/min infusion rate (p = 5 x 10(-14)), Uptake ratio was not affected by vector quantity or slower infusion rates. Transforming growth factor-beta(1) was consistently de tected in allografts infected with AdCMVTGF-beta(1), but not with cont rol adenovirus or AdSvIL10. Expression was proportional to infused vec tor quantity and reached 10 ng/gm of allograft at infused 10(10) pfu/g m. Transforming growth factor-beta(1) was also detected in recipient's serum at less than 1 ng/ml. Viral interleukin-10 was detected in mino r amounts only (<1 ng/gm) in allografts infected with AdvIL10 up to 5 x 10(10) pfu/gm. Nevertheless, it was detected in recipient serum at c oncentrations up to 0.4 ng/ml. Conclusions: Intracoronary gene transfe r of immunosuppressive cytokines to cardiac allografts during cold pre servation is feasible. Slow infusion is superior to bolus injection. I n vivo effects on allograft rejection remain to be determined.