Efficient transduction of CD34+CD38- cells with lentiviral vectors

G. M. Crooks, S. S. Case, M. A. Price, L. Naldini, D. B. Kohn

Research output: Contribution to journalArticle

Abstract

Current Moloney Murine Leukemia Virus (MLV) based vector strategies have failed to produce efficient transduction of human stem cells, a population which is quiescent and expresses low levels of virus receptor. A novel vector system in which a lentiviral vector (lenti) is pseudotyped with the vesicular stomatitis virus envelope G protein (VSV) has been developed in order to overcome the innate obstacles of stem cell transduction. We compared the efficiency of transducing CD34+ subpopulations using "lenti/VSV" with MLV pseudotyped with either VSV (MLV/VSV) or with GALV (MLV/GALV) under a variety of conditions and at increasing Multiplicities of Infection (MOI). Each construct contained the enhanced Green Fluorescent Protein (eGFP) as a reporter gene (using the CMV promoter in lenti/VSV and MLV/VSV and the LTR promoter in MLV/GALV). No toxicity was noted with MOI up to 3,000. Transduction of CD34+CD38+cells over three days in IL-3. IL-6 and SF (36S) resulted in high efficiency transduction of short term cultures with all three vectors (eGFP expression 14-58% with lenti/VSV (MOI 300-3,000), 12-39% with MLV/VSV (MOI 300-3,000) and 6-36% with MLV/GALV (MOI 300) (n=4). However, eGFP was only detectable in long term culture of CD34+CD38+ cells transduced with lenti/VSV (12-22% eGFP at 5 weeks). Three day transduction of more primitive progenitors, CD34+CD38- cells, in 36S with lenti/VSV resulted in 15-39% eGFP expression persisting for at least 5 weeks of culture (n=3) In contrast, MLV/VSV and MLV/GALV produced only 0-2% eGFP expression in short or long term cultures of CD34+CD38- cells. We next studied whether progenitors could be transduced by lenti/VSV during brief exposure to virus without growth factor stimulation. Exposure to lenti/VSV for 12 hours in the absence of growth factors resulted in eGFP expression in 7-12% of CD34+CD38+ cells (n=2) and 52% of CD34+CD38- cells (n=l) with stable expression in long term culture No eGFP expression was detectable in short or long term cultures after transduction with MLV/VSV or MLV/GALV in the absence of growth factors. The lenti/VSV system thus significantly improves the efficiency of gene transfer into a quiescent population of primitive progenitors and may provide a method for efficient hematopoietic stem cell transduction.

Original languageEnglish
Pages (from-to)814
Number of pages1
JournalExperimental Hematology
Volume26
Issue number8
Publication statusPublished - 1998

Fingerprint

Murine Leukemia Viruses
Gibbon ape leukemia virus
Infection
Intercellular Signaling Peptides and Proteins
Stem Cells
Viral Envelope Proteins
Moloney murine leukemia virus
Virus Receptors
enhanced green fluorescent protein
Interleukin-3
Hematopoietic Stem Cells
Reporter Genes
Population
Interleukin-6
Viruses

ASJC Scopus subject areas

  • Cancer Research
  • Cell Biology
  • Genetics
  • Hematology
  • Oncology
  • Transplantation

Cite this

Crooks, G. M., Case, S. S., Price, M. A., Naldini, L., & Kohn, D. B. (1998). Efficient transduction of CD34+CD38- cells with lentiviral vectors. Experimental Hematology, 26(8), 814.

Efficient transduction of CD34+CD38- cells with lentiviral vectors. / Crooks, G. M.; Case, S. S.; Price, M. A.; Naldini, L.; Kohn, D. B.

In: Experimental Hematology, Vol. 26, No. 8, 1998, p. 814.

Research output: Contribution to journalArticle

Crooks, GM, Case, SS, Price, MA, Naldini, L & Kohn, DB 1998, 'Efficient transduction of CD34+CD38- cells with lentiviral vectors', Experimental Hematology, vol. 26, no. 8, pp. 814.
Crooks, G. M. ; Case, S. S. ; Price, M. A. ; Naldini, L. ; Kohn, D. B. / Efficient transduction of CD34+CD38- cells with lentiviral vectors. In: Experimental Hematology. 1998 ; Vol. 26, No. 8. pp. 814.
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