Comparison of Various Envelope Proteins for Their Ability to Pseudotype Lentiviral Vectors and Transduce Primitive Hematopoietic Cells from Human Blood

Abstract

Substantial effort has been invested in developing methodologies for efficient gene transfer into human, repopulating, hematopoietic stem cells. Oncoretroviral vectors are limited by the lack of nuclear mitosis in quiescent stem cells during ex vivo transduction, whereas the preintegration complex of lentiviral vectors contains nuclear-localizing signals that permit genome integration without mitosis. We have developed a flexible and versatile system for generating lentiviral vector particles and have pseudotyped such particles with amphotropic, ecotropic, feline endogenous virus (RD114) or vesicular stomatitis virus (VSV-G) envelope proteins. Particles of all four types could be concentrated approximately 100-fold by ultracentrifugation or ultrafiltration. RD114 or amphotropic particles were more efficient than VSV-G-pseudotyped particles at transducing human cord blood CD34(+) cells and clonogenic progenitors within that population. Amphotropic particles transduced cytokine-mobilized, human peripheral blood CD34(+) cells capable of establishing hematopoiesis in immunodeficient mice more efficiently than the other two types of particles. We conclude that the use of amphotropic pseudotyped lentiviral vector particles rather than the commonly used VSV-G-pseudotyped particles should be considered in potential applications of lentiviral vectors for gene transfer into this therapeutically relevant target cell population.

A new xenograft model of myeloma bone disease demonstrating the efficacy of human mesenchymal stem cells expressing osteoprotegerin by lentiviral gene transfer

“…Third generation self-inactivating lentivirus were produced as described previously. 23 Briefly, 293 T cells were transfected with four plasmids containing (1) the lentiviral gag-pol genes, (2) the vesicular stomatitis virus glycoprotein envelope gene, (3) the reverse transcriptase, and (4) a bicistronic vector (pCL10.1) as described. Biological titres of the supernatant were determined by limiting dilution on HeLa cells.…”

Gene therapy to inhibit xenoantibody production using lentiviral vectors in non-human primates

“…[33][34][35][36][37] We elected to use SIVderived lentiviral vectors for this study based on reports that they are more efficient for transduction of simian stem cells. 19,20,[38][39][40] Our results using SIV-based aGT constructs are consistent with those findings.…”

Surface-engineering of lentiviral vectors

“…Lentiviral vectors pseudotyped with the glycoproteins from Ross River virus show superiority over vectors pseudotyped with VSV-G for in vivo gene delivery to the liver of mice [41]. Likewise, screening of a large panel of pseudotyped vectors established the superiority of the gibbon ape leukemia virus (GALV) and the cat endogenous retroviral glycoproteins (RD114) for transduction of progenitor and differentiated hematopoietic cells [22,34,[42][43][44][45]. Finally, owing to the capacity of the influenza virus, Sindis virus, MLV and GALV glycoproteins, among others, to pseudotype MLV and lentiviral vectors [22,25,46], several Engineering Lentiviral Vectors S85 approaches can be undertaken to modify their tropism in order to modify the host range of vectors (see below).…”