title: CpG-depleted plasmid DNA vectors with enhanced safety and long-term gene expression in vivo.
abstract: Systemic delivery of cationic lipid-plasmid DNA (pDNA) complexes induces an acute inflammatory response with adverse hematologic changes and liver damage. Immunostimulatory CpG motifs in the pDNA are known to contribute substantially to this response. Here we constructed a pDNA vector (pGZB) that has been depleted of 80% of the CpG motifs present in the original vector. Compared with the unmodified vector, systemic administration of pGZB induced considerably fewer changes in blood parameters, reduced levels of inflammatory cytokines, and decreased liver damage. Despite the extensive sequence modifications within pGZB, there were still robust levels of transgene expression. Furthermore, in contrast to the transient expression observed from the unmodified vector, we observed sustained or increasing expression for up to 49 days from pGZB in the lung and liver of immunocompetent BALB/c mice. Studies adding CpG motifs in trans or in cis indicate that the reduced CpG content of pGZB was the major determinant for its persistent expression. This combination of decreased toxicity and sustained expression suggests that CpG-depleted pDNA vectors can greatly improve the safety and efficacy of synthetic gene delivery systems.
authors.affiliation: Genzyme Corporation, 31 New York Avenue, Framingham, Massachusetts 01701-9322, USA. nelson.yew@genzyme.com
authors.forename: Nelson S
authors.initials: NS
authors.lastname: Yew
authors.forename: Hongmei
authors.initials: H
authors.lastname: Zhao
authors.forename: Malgorzata
authors.initials: M
authors.lastname: Przybylska
authors.forename: I-Huan
authors.initials: IH
authors.lastname: Wu
authors.forename: Jennifer D
authors.initials: JD
authors.lastname: Tousignant
authors.forename: Ronald K
authors.initials: RK
authors.lastname: Scheule
authors.forename: Seng H
authors.initials: SH
authors.lastname: Cheng
mesh_terms: D000818:Animals; D015500:Chloramphenicol O-Acetyltransferase / Q000378:metabolism; D018899:CpG Islands; D005164:Factor IX / Q000378:metabolism; D015870:Gene Expression; D005822:Genetic Vectors / Q000235:genetics / Q000633:toxicity; D008099:Liver / Q000378:metabolism; D008168:Lung / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C
chemical_list: D005164:Factor IX; D015500:Chloramphenicol O-Acetyltransferase