High-level gene transfer to cord blood progenitors using gibbon ape leukemia virus pseudotype retroviral vectors and an improved clinically applicable protocol.

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  • Additional Information
    • Source:
      Publisher: M.A. Liebert Country of Publication: United States NLM ID: 9008950 Publication Model: Print Cited Medium: Print ISSN: 1043-0342 (Print) Linking ISSN: 10430342 NLM ISO Abbreviation: Hum Gene Ther Subsets: MEDLINE
    • Publication Information:
      Original Publication: New York : M.A. Liebert, c1990-
    • Subject Terms:
      Gene Transfer Techniques*; Fetal Blood/*virology ; Genetic Vectors/*therapeutic use ; Leukemia Virus, Gibbon Ape/*genetics ; Stem Cells/*virology; Antigens, CD34/analysis ; Cell Line ; Centrifugation ; Coculture Techniques ; Culture Media ; Fetal Blood/cytology ; Fetal Blood/drug effects ; Fibronectins/pharmacology ; Hexadimethrine Bromide/pharmacology ; Humans ; Leukemia Virus, Gibbon Ape/growth & development ; Stem Cells/drug effects ; Stem Cells/metabolism
    • Abstract:
      The best methods for transducing hematopoietic progenitor cells usually involve either direct co-cultivation with virus-producing cells or human stromal supportive cells. However, these methods cannot be safely or easily applied to clinical use. Therefore, we aimed at improving retrovirus-mediated gene transfer into hematopoietic progenitors derived from cord blood CD34+ cells using viral supernatant to levels achieved at least with direct co-cultivation and under conditions that are suitable for clinical applications. In a first set of experiments, CD34+ cells were infected with supernatant containing amphotropic retroviral particles carrying the nls-lacZ reporter gene and the effects of centrifugation, cell adhesion to fibronectin, and Polybrene on the transduction of both clonogenic progenitors (CFC) and long-term culture initiating cells (LTC-IC) were studied. Transduction efficiency was evaluated on the percentage and total number of progenitors expressing the beta-galactosidase activity. Results show that a 48-hr infection of CD34+ cells with viral supernatant combining centrifugation at 1000 x g for 3 hr followed by adhesion to fibronectin allows transduction levels for both CFC and LTC-IC to be reached that are as good as using direct co-cultivation. In a second set of experiments, CD34+ cells were infected using this optimized protocol with pseudotyped retroviral particles carrying the gibbon ape leukemia virus (GALV) envelope protein. Under these conditions, between 50 and 100% of CFC and LTC-IC were transduced. Thus, we have developed a protocol capable of highly transducing cord blood progenitors under conditions suitable for a therapeutical use.
    • Accession Number:
      0 (Antigens, CD34)
      0 (Culture Media)
      0 (Fibronectins)
      4C905MSK4W (Hexadimethrine Bromide)
    • Publication Date:
      Date Created: 19980224 Date Completed: 19980325 Latest Revision: 20131121
    • Publication Date:
      20240829
    • Accession Number:
      10.1089/hum.1998.9.2-225
    • Accession Number:
      9472782