535. Long-Term Correction of Hyperbilirubinemia in a Rat Model of Crigler-Najjar Syndrome Type 1

Abstract

Top of pageAbstract Crigler-Najjar syndrome Type 1 is a recessively inherited disorder caused by a deficiency of the hepatic enzyme bilirubin uridine diphosphate-glucuronosyltransferase (UGT1A1), and is manifested as severe unconjugated hyperbilirubinemia. Unconjugated, water-insoluble bilirubin is not excreted and therefore accumulates in blood and in the central nervous system. This accumulation is fatal to a newborn if untreated. Phototherapy is currently utilized, owing to the ability of blue light to break bilirubin down into water-soluble fragments. However, its efficacy is reduced upon thickening of the skin and further reduced as a patient's surfavolume ratio changes around puberty. The alternative, liver transplantation, is neither easily affordable nor widely accessible. We have evaluated an adenoviral gene therapy modality for restoration of UGT1A1 function, over the long term, to the Gunn rat model of Crigler-Najjar Syndrome. We employed a helper-dependent adenoviral vector, whose lack of viral coding sequences allows greater persistence of transgene expression coupled to reduced chronic toxicity, as compared to early-generation adenoviral vectors. The vector contains the human UGT1A1 cDNA driven by the liver-restricted rat PEPCK promoter and a Woodchuck Posttranscriptional Regulatory Element (WPRE). Adenoviral vector was administered to Gunn rats via the tail vein at the following doses: 6, 30 and 100 (|[times]|1011 viral particles/kilogram). Treatment did not cause significant elevation of serum transaminases which are indicative of hepatocyte injury. Furthermore, moderate thrombocytopenia developed after vector administration but had resolved within a few days of injection. A clinically significant reduction in bilirubin levels was observed using the lowest adenoviral dose; this correction was maintained past 67 weeks. In rats receiving intermediate or high doses, complete correction of hyperbilirubinemia was sustained for over 2 years. HPLC analysis of serum processed from terminal bleeds demonstrated a dose-dependent reduction of unconjugated bilirubin, while bile HPLC analysis showed an increase in conjugated (glucuronidated) bilirubin. For an additional in vivo assay of UGT1A1 activity, HPLC analysis was carried out on bile samples taken before and after tail vein injection of bilirubin isomers BR-III and BR-XIII. Conjugation of these isomers requires UGT1A1 activity, and HPLC analysis showed an excess of bilirubin conjugating capacity in rats receiving intermediate or high doses. Taken together, these results support the efficacy of a single systemic administration of a helper-dependent adenoviral vector towards restoring UGT1A1 enzyme function over the long term. Acute toxicities occurred but were quickly resolved. The relatively favorable dose response makes this condition a candidate for considering clinical trials in the future.