A step toward liver gene therapy: efficient correction of the genetic defect of hepatocytes isolated from a patient with Crigler-Najjar syndrome type 1 with lentiviral vectors.
BACKGROUND
Ex vivo liver gene therapy may be a future alternative to orthotopic liver transplantation for the treatment of some liver diseases. We previously described the transduction in suspension with lentiviral vectors and immediate hepatocyte transplantation (SLIT) protocol and its high transduction rate with normal human hepatocytes. We also reported SLIT efficiency in the animal model of Crigler-Najjar type 1 syndrome (CN-1), the Gunn rat. Here, we evaluated SLIT efficiency with diseased human hepatocytes.


METHODS
Hepatocytes of the liver from a 4-year-old patient presenting CN-1 were isolated. They were transduced with liver-specific lentiviral vectors expressing uridine-diphosphate-glucuronosyltransferase (hUGT1A1) or green fluorescent protein, and then analyzed in vitro for transduction efficiency and hUGT1A1 expression, or transplanted in nonobese diabetic/severe combined immunodeficiency (SCID) mice to evaluate long-term survival of transplanted cells.


RESULTS
More than 90% of CN-1 hepatocytes were transduced. Hepatocytes produced hUGT1A1 protein after lentiviral transduction. After having been subjected to the SLIT, lentivirally transduced CN-1 hepatocytes engrafted long term (up to 26 weeks posttransplantation) in recipient livers and expressed green fluorescent protein or hUGT1A1 vector.


CONCLUSIONS
The SLIT protocol allowed for a high transduction of CN-1 hepatocytes and restoration of the expression of the deficient protein. Furthermore, long-term survival of lentivirally transduced CN-1 hepatocytes in the liver of immunodeficient mice was demonstrated. This study is therefore an important step toward human application of lentiviral gene therapy.
Birraux
Jacques
J
University of Geneva Medical Centre (CMU), Research Laboratory of Pediatric Surgery, Geneva, Switzerland.
Menzel
Olivier
O
Wildhaber
Barbara
B
Jond
Caty
C
Nguyen
Tuan Huy
TH
Chardot
Christophe
C
D000818:Animals; D002675:Child, Preschool; D003414:Crigler-Najjar Syndrome / Q000235:genetics*; D015925:Cryopreservation / Q000379:methods; D005260:Female; D015316:Genetic Therapy; D005822:Genetic Vectors; D014453:Glucuronosyltransferase / Q000235:genetics*; D006498:Hepatectomy; D022781:Hepatocytes / Q000166:cytology / Q000502:physiology* / Q000637:transplantation*; D006801:Humans; D016086:Lentivirus / Q000235:genetics*; D051379:Mice; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D000099148:UGT1A1 Enzyme
D014453:Glucuronosyltransferase; D000099148:UGT1A1 Enzyme