A non-immunogenic adenoviral vector, coexpressing CTLA4lg and bilirubin-uridine-diphosphoglucuronateglucuronosyltransferase permits long-term, repeatable transgene expression in the Gunn rat model of Crigler-Najjar syndrome

N. R. Thummala, S. S. Ghosh, S. W. Lee, B. Reddy, A. Davidson, M. S. Horwitz, Jayanta Roy-Chowdhury, N. Roy Chowdury

Research output: Contribution to journalArticle

48 Citations (Scopus)

Abstract

Host immune responses limit the duration of expression of transgenes introduced by recombinant adenoviruses, preclude gene transfer upon vector readministration and cause liver injury. CTLA4lg inhibits immune response by blocking the co-stimulatory interaction between CD28 on T cells and B7 on antigen-presenting cells. We have constructed a recombinant adenovirus, Ad-hUGT1A1-CTLA4lg that co-expresses human bilirubin-uridinediphosphoglucuronate glucuronosyltransferase (hUGT1A1) and soluble murine CTLA4lg, both driven by CMV immediate-early promoters. After intravenous injection of this vector (6 × 1011 p.f.u.) into UGT1A1-deficient jaundiced Gunn rats, serum CTLA4lg levels peaked at 1.8-2.0 mg/ml on day 7 and declined thereafter to 0.2 mg/ml by day 180. Serum bilirubin declined from mean preinjection levels of 8.0 mg/dl to 0.48-0.6 mg/dl in 3 days, remained normal for 28 weeks, and then gradually increased to 8 mg/dl by day 350. A second injection of Ad-hUGT1A1-CTLA4lg normalized serum bilirubin. In two rats in this group that were followed longer, serum bilirubin increased to 3.1 and 3.5 mg/dl in 40 weeks, but was normalized again after a third injection. The antibody and cytotoxic lymphocyte (CTL) responses were negligible, and liver biopsy showed no inflammatory cell infiltration. Rats receiving a tertiary challenge with Ad-LacZ (expressing E. coli β-galactosidase) (5 × 1011 p.f.u.), 2 months after the second dose of Ad-hUGT1A1-CTLA4lg, showed β-galactosidase expression in over 80% of hepatocytes. In contrast, after Ad-hUGT1A1 (which expresses UGT1A1 alone) injection, serum bilirubin remained normal for only 4 weeks, and returned to preinjection levels by day 120. Bilirubin levels did not decline upon reinjection, and β-galactosidase was not expressed after Ad-LacZ. High levels of adenovirus-specific antibodies and CTL, and hepatic inflammation were found. This is the first demonstration that coexpression of CTLA4lg permits prolonged expression and repeatable gene transfer by an adenoviral vector.

Original languageEnglish (US)
Pages (from-to)981-990
Number of pages10
JournalGene Therapy
Volume9
Issue number15
DOIs
StatePublished - 2002

Fingerprint

Crigler-Najjar Syndrome
Gunn Rats
Uridine
Transgenes
Bilirubin
Galactosidases
Adenoviridae
Serum
Injections
Liver
B7 Antigens
Lymphocytes
Glucuronosyltransferase
Antibodies
Antigen-Presenting Cells
Licensure
Jaundice
Intravenous Injections
Hepatocytes
Escherichia coli

Keywords

  • CTLA4-lg
  • Gene therapy
  • Gunn rat
  • Recombinant adenovirus
  • UPD-glucuronysltransferase

ASJC Scopus subject areas

  • Genetics

Cite this

A non-immunogenic adenoviral vector, coexpressing CTLA4lg and bilirubin-uridine-diphosphoglucuronateglucuronosyltransferase permits long-term, repeatable transgene expression in the Gunn rat model of Crigler-Najjar syndrome. / Thummala, N. R.; Ghosh, S. S.; Lee, S. W.; Reddy, B.; Davidson, A.; Horwitz, M. S.; Roy-Chowdhury, Jayanta; Chowdury, N. Roy.

In: Gene Therapy, Vol. 9, No. 15, 2002, p. 981-990.

Research output: Contribution to journalArticle

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abstract = "Host immune responses limit the duration of expression of transgenes introduced by recombinant adenoviruses, preclude gene transfer upon vector readministration and cause liver injury. CTLA4lg inhibits immune response by blocking the co-stimulatory interaction between CD28 on T cells and B7 on antigen-presenting cells. We have constructed a recombinant adenovirus, Ad-hUGT1A1-CTLA4lg that co-expresses human bilirubin-uridinediphosphoglucuronate glucuronosyltransferase (hUGT1A1) and soluble murine CTLA4lg, both driven by CMV immediate-early promoters. After intravenous injection of this vector (6 × 1011 p.f.u.) into UGT1A1-deficient jaundiced Gunn rats, serum CTLA4lg levels peaked at 1.8-2.0 mg/ml on day 7 and declined thereafter to 0.2 mg/ml by day 180. Serum bilirubin declined from mean preinjection levels of 8.0 mg/dl to 0.48-0.6 mg/dl in 3 days, remained normal for 28 weeks, and then gradually increased to 8 mg/dl by day 350. A second injection of Ad-hUGT1A1-CTLA4lg normalized serum bilirubin. In two rats in this group that were followed longer, serum bilirubin increased to 3.1 and 3.5 mg/dl in 40 weeks, but was normalized again after a third injection. The antibody and cytotoxic lymphocyte (CTL) responses were negligible, and liver biopsy showed no inflammatory cell infiltration. Rats receiving a tertiary challenge with Ad-LacZ (expressing E. coli β-galactosidase) (5 × 1011 p.f.u.), 2 months after the second dose of Ad-hUGT1A1-CTLA4lg, showed β-galactosidase expression in over 80{\%} of hepatocytes. In contrast, after Ad-hUGT1A1 (which expresses UGT1A1 alone) injection, serum bilirubin remained normal for only 4 weeks, and returned to preinjection levels by day 120. Bilirubin levels did not decline upon reinjection, and β-galactosidase was not expressed after Ad-LacZ. High levels of adenovirus-specific antibodies and CTL, and hepatic inflammation were found. This is the first demonstration that coexpression of CTLA4lg permits prolonged expression and repeatable gene transfer by an adenoviral vector.",
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AU - Roy-Chowdhury, Jayanta

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