title: Hepatic lentiviral gene transfer is associated with clonal selection, but not with tumor formation in serially transplanted rodents.
abstract: Lentiviral (LV) vectors are promising tools for long-term genetic correction of hereditary diseases. In hematopoietic stem cell gene therapies adverse events in patients due to vector integration-associated genotoxicity have been observed. Only a few studies have explored the potential risks of LV gene therapy targeting the liver. To analyze hepatic genotoxicity in vivo, we transferred the fumarylacetoacetate hydrolase (FAH) gene by LV vectors into FAH((-/-)) mice (n = 97) and performed serial hepatocyte transplantations (four generations). The integration profile (4,349 mapped insertions) of the LV vectors was assessed by ligation-mediated polymerase chain reaction and deep sequencing. We tested whether the polyclonality of vector insertions was maintained in serially transplanted mice, linked the integration sites to global hepatocyte gene expression, and investigated the effects of LV liver gene therapy on the survival of the animals. The lifespan of in vivo gene-corrected mice was increased compared to 2-(2-nitro-4-trifluoromethylbenzoyl)-1,3-cyclohexanedione (NTBC) control animals and unchanged in serially transplanted animals. The integration profile (4,349 mapped insertions) remained polyclonal through all mouse generations with only mild clonal expansion. Genes close to the integration sites of expanding clones may be associated with enhanced hepatocyte proliferation capacity.
  
  
  CONCLUSIONS
  We did not find evidence for vector-induced tumors. LV hepatic gene therapy showed a favorable risk profile for stable and long-term therapeutic gene expression. Polyclonality of hepatocyte regeneration was maintained even in an environment of enforced proliferation.
authors.affiliation: Gastroenterology, Hepatology and Endocrinology, Hannover Medical School, Germany.
authors.forename: Ina
authors.initials: I
authors.lastname: Rittelmeyer
authors.forename: Michael
authors.initials: M
authors.lastname: Rothe
authors.forename: Martijn H
authors.initials: MH
authors.lastname: Brugman
authors.forename: Marcus
authors.initials: M
authors.lastname: Iken
authors.forename: Axel
authors.initials: A
authors.lastname: Schambach
authors.forename: Michael P
authors.initials: MP
authors.lastname: Manns
authors.forename: Christopher
authors.initials: C
authors.lastname: Baum
authors.forename: Ute
authors.initials: U
authors.lastname: Modlich
authors.forename: Michael
authors.initials: M
authors.lastname: Ott
mesh_terms: D000818:Animals; D002999:Clone Cells; D018628:Gene Dosage; D015316:Genetic Therapy / Q000009:adverse effects*; D005822:Genetic Vectors; D022781:Hepatocytes / Q000637:transplantation*; D006867:Hydrolases / Q000235:genetics*; D016086:Lentivirus / Q000235:genetics*; D008113:Liver Neoplasms / Q000235:genetics*; D051379:Mice; D008810:Mice, Inbred C57BL; D016133:Polymerase Chain Reaction / Q000379:methods
chemical_list: D006867:Hydrolases; C020156:fumarylacetoacetase