Jul 10, 1996

Expression of the human multidrug resistance and glucocerebrosidase cDNAs from adeno-associated vectors: efficient promoter activity of AAV sequences and in vivo delivery via liposomes

Human Gene Therapy
M BaudardM M Gottesman

Abstract

Recombinant adeno-associated viruses (rAAV) are attractive tools for gene therapy. We designed plasmids in which the human multidrug resistance gene (hMDR1) cDNA was placed downstream from portions of the 5' end of AAV including either a 234-bp cassette or the entire AAV p5 promoter. The drug-resistant phenotype conferred by the P-glycoprotein (Pgp) efflux pump encoded by the hMDR1 cDNA was used to select NIH-3T3 cells transfected with these plasmids. The 234-bp region alone showed promoter activity similar in strength to that of the entire p5 promoter or the retroviral Harvey murine sarcoma virus long terminal repeat (LTR); this result demonstrates that the 234-bp cassette might be used as a small and efficient promoter in rAAV designed to express large genes approaching the packaging limit of AAV particles. After transfection of AAV-MDR1 vectors, the integration of MDR1 sequences into the host cell genome was demonstrated by fluorescent in situ hybridization (FISH). In addition, Southern analysis of low-molecular-weight DNA extracted from drug-resistant cells grown under continuous selection pressure indicated the persistence of nonintegrated AAV-MDR1 plasmids. Coordinate expression of Pgp and human glucocerebrosidase (hGC) w...Continue Reading

  • References35
  • Citations19

References

  • References35
  • Citations19

Citations

Mentioned in this Paper

Abcb1b
Fluorescent in Situ Hybridization
Sarcoma
Shuttle Vectors
Transfection
Latent Virus Infection Process
Harvey murine sarcoma virus
Long Terminal Repeat
Intravenous Injections
Adeno-Associated Virus

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