Our long-term objective is to develop safe and efficient methods for liver-directed gene therapy for genetic disorders of hepatic metabolism, using inherited jaundice due to bilirubin-uridinediphospho-glucuronate glucuronosyltransferase (bilirubin-UGT) deficiency as a model target. Absence of hepatic bilirubin-UGT activity leads to accumulation of the bilirubin in plasma and consequent brain damage, resulting in the potentially lethal Crigler-Najjar syndrome type I (CN-I). Currently, liver transplantation is the only definitive therapy. Gunn rats, the animal model for CN-I, will be used for these studies. During the last four years, both viral and non-viral vehicles for delivery of normal human bilirubin-UGT genes to Gunn rat liver were devised and tested. In this continuation application, two strategies for correcting hepatic bilirubin-UGT deficiency in vivo will be pursued. In the first approach, recombinant viral vectors will be used to substitute hepatic bilirubin-UGT by delivering a normal gene. Recombinant adenoviruses are very efficient in transferring genes into the liver in vivo, but the transgene expression is transient because the virus is episomal and the host immune response precludes its repeated administration. Three methods for specific tolerization of the host to adenoviral antigens, developed in our laboratory, show promise for long-term adenovirus- mediated gene therapy without immunosuppression. To refine these methods for future clinical application, the mechanisms by which they induce specific tolerance will be elucidated. In addition, a much less immunogenic vector, based on the SV40 virus, will be developed and tested. The second approach will utilize RNA/DNA chimeric molecules to repair the genetic lesion in Gunn rats, using our highly efficient vectors for liver-specific nucleic acid delivery by receptor-mediated endocytosis. The gene transfer efficiency will be assessed by molecular, enzymatic and metabolic studies. Successful completion of these studies will provide a basis for gene therapies for CN-I and other inherited metabolic disorders, such as deficiency of alpha1-antitrypsin and urea cycle enzymes.
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