Adenovirus-mediated gene therapy to liver grafts: Successful gene transferby donor pretreatment

Background. We have previously shown excellent adenoviral (Ad) gene transfe ction to transplanted liver grafts with the clamp technique (CT) where vira l vector was delivered ex vivo and trapped in cold preserved liver grafts. In this study, we adopted a new gene therapy approach to achieve early tran sgene expression by donor pretreatment with viral vector and compared the e fficacy of these two methods by using Ad vector encoding enhanced green flu orescent protein (AdEGFP) marker gene. Methods. AdEGFP (1 x 10(9) plaque forming units) was delivered to the liver grafts by: (1) single intravenous injection to donor Lewis rats 48 hours b efore harvesting (2) ex vivo cold infusion into the harvested liver with CT or (3) a combination of both methods. Liver grafts were stored in Universi ty of Wisconsin solution at 4 degrees C for 18 hours and then orthotopicall y transplanted into syngeneic recipients, and the expression of EGFP was st udied. Results. With intravenous pretreatment of donor liver grafts, EGFP-expressi ng cells were detected as early as 3 hours after transplant, and moderate e xpression was seen by 12 hours. In contrast, EGFP was not detected until 12 to 24 hours after transplant with CT High levels of EGFP-producing cells w ere seen with each technique at 7 days (similar to 30% transfection efficie ncy). A combination of both methods did not enhance infectivity. Liver pres ervation injury was comparable between groups. Conclusions. Gene transfer by donor pretreatment with AdEGFP induces early and efficient gene transduction to liver grafts compared with back-table de livery with CT This method is simple and provides early transgene expressio n in liver grafts that potentially could be used to deliver genes to decrea se preservation injury or rejection.