Crigler-Najjar syndrome is a recessively inherited disorder characterized by severe unconjugated hyperbilirubinemia caused by a deficiency of uridine diphospho-glucuronosyl transferase 1A1. Current therapy relies on phototherapy to prevent kernicterus, but liver transplantation presently is the only permanent cure. Gene therapy is a potential alternative, and recent work has shown that helper-dependent adenoviral (HD-Ad) vectors, devoid of all viral coding sequences, induce prolonged transgene expression and exhibit significantly less chronic toxicity than early-generation Ad vectors. We used a HD-Ad vector to achieve liver-restricted expression of human uridine diphospho-glucuronosyl transferase 1A1 in the Gunn rat, a model of the human disorder. Total plasma bilirubin levels were reduced from >5.0 mg͞dl to Ͻ Ͻ1.4 mg͞dl for >2 yr after a single i.v. administration of vector expressing the therapeutic transgene at a dose of 3 ؋ 10 12 viral particles per kg. HPLC analysis of bile from treated rats showed the presence of bilirubin glucuronides at normal WT levels >2 yr after one injection of vector, and i.v. injection of bilirubins III␣ and XIII␣ in the same animals revealed excess bilirubin-conjugating capacity. There was no significant elevation of liver enzymes (alanine aminotransferase) and only transient, moderate thrombocytopenia after injection of the vector. A clinically significant reduction in serum bilirubin was observed with a dose as low as 6 ؋ 10 11 viral particles per kg. We conclude that complete, long-term correction of hyperbilirubinemia in the Gunn rat model of Crigler-Najjar syndrome can be achieved with one injection of HD-Ad vector and negligible chronic toxicity.adenovirus ͉ bilirubin ͉ gene therapy
Bilirubin, the yellow pigment of jaundice, is a linear tetrapyrrole with a methylene group at its
center, C(10), a position of crucial importance to its conformation and metabolism. The presence of the central
methylene group allows the bilirubin to fold into an intramolecularly hydrogen-bonded conformation. This
paper describes the first synthesis of a bilirubin analogue with an oxo group at C(10). The change from CH2
to CO, from sp3 to sp2, is designed to stress the molecule at its hinge and relax its conformation. Such
compounds have been suggested as potential oxidative metabolites of bilirubin in vivo. 10-Oxo-mesobilirubin-XIIIα (1) is a red crystalline solid, unlike its parent, mesobilirubin-XIIIα, which is a bright yellow solid. It is
surprisingly polar, relative to the parent, yet it does not exhibit a significantly larger bicarbonate/chloroform
partition coefficient. Like the parent, 1 appears to adopt an intramolecularly hydrogen-bonded ridge-tile-like
conformation. In normal rats, 1 is metabolized to acylglucuronides, which are secreted into bile, but a portion
of the administered dose is secreted into bile intact. In mutant rats (Gunn rats) lacking bilirubin glucuronyl
transferase, 1 was excreted efficiently in bile in unchanged form, unlike the parent with a methylene group at
C(10). Thus, introduction of the oxygen function at C(10) has little effect on hepatic uptake but a dramatic
effect on canalicular secretion into bile.
A new class of highly fluorescent (phi(F) 0.3-0.8) low molecular weight water-soluble cholephilic compounds has been synthesized in two steps from dipyrrinones. The dipyrrinone nitrogens are first bridged by reaction with 1,1'-carbonyldiimidazole to form an N,N'-carbonyldipyrrinone (3H,5H-dipyrrolo[1,2-c:2',1'-f]pyrimidine-3,5-dione) nucleus, and a sulfonic acid group is then introduced at C(8) by reaction with concd H(2)SO(4). The resulting sulfonated N,N'-carbonyl-bridged dipyrrinones ("sulfoglows") are isolated as their sodium salts. When the alkyl substituents of the lactam ring are lengthened from ethyl to decyl, sulfoglows become increasingly lipophilic while maintaining water solubility. Low molecular weight sulfoglows were rapidly excreted intact in both bile and urine after intravenous infusion into rats, but higher molecular weight sulfoglows were excreted more selectively in bile. Hepatobiliary excretion of sulfoglows was partially, but not completely, blocked in mutant rats deficient in the multidrug-resistance associated transport protein Mrp2 (ABCC2). These observations point to the feasibility of developing simple sulfoglows with clinical diagnostic potential that are normally excreted in bile but appear in urine when hepatic elimination is impaired by cholestatic liver disease.
The metabolism and biliary excretion of a stretched bilirubin analog with a p-xylyl group replacing the central CH 2 hinge were investigated in normal rats, Gunn rats deficient in bilirubin conjugation, and TR 3 rats deficient in bilirubin glucuronide hepatobiliary transport. Unlike bilirubin, the analog was excreted rapidly in bile unchanged in all three rat strains after intravenous administration. In TR 3 rats biliary excretion of the analog was diminished, but still substantial, demonstrating that the ATP-binding cassette transporter Mrp2 is not required for its hepatic efflux. These effects are attributable to differences in the preferred conformations of bilirubin and the analog. ß 2001 Published by Elsevier Science B.V. on behalf of the Federation of European Biochemical Societies.
Fused pyrimidine derivatives R 0515 Synthesis and Hepatic Transport of Strongly Fluorescent Cholephilic Dipyrrinones. -(WOYDZIAK, Z. R.; BOIADJIEV, S. E.; NORONA, W. S.; MCDONAGH, A. F.; LIGHTNER*, D. A.; J. Org. Chem. 70 (2005) 21, 8417-8423; Dep. Chem., Univ. Nev., Reno, NV 89557, USA; Eng.) -Lindner 03-162
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