title: Efficiency of onco-retroviral and lentiviral gene transfer into primary mouse and human B-lymphocytes is pseudotype dependent.
abstract: B lymphocytes are attractive targets for gene therapy of genetic diseases associated with B-cell dysfunction and for immunotherapy. Transduction of B lymphocytes was evaluated using green fluorescent protein (GFP)-encoding onco-retroviral and HIV-derived lentiviral vectors which were pseudotyped with ecotropic, amphotropic or vesicular stomatitis virus (VSV-G) envelopes. Transduction of mouse B lymphocytes activated with lipopolysaccharides (LPS) or by cross-linking CD40 in conjunction with interleukin-4 (IL-4) was significantly more efficient (p < 0.003) with ecotropic (11%) than with VSV-G pseudotyped onco-retroviral vectors (1%). Using high-titer cell-free ecotropic viral supernatant or by coculture with ecotropic onco-retroviral vector-producing cells, transduction efficiency increased significantly (p < 0.001) to approximately 50%, whereas transduction efficiency by coculture with VSV-G pseudotyped vector-producing cells remained low (< 2%). Similarly, transduction of mouse B lymphocytes was significantly more efficient (twofold, p < 0.01) with the ecotropic (7%) than with the VSV-G pseudotyped lentiviral vectors although gene transfer efficiency remained low because of dose-limiting toxicity of the concentrated vector preparations on the LPS-activated murine B cells. Consistent with murine B-cell transduction, human B cells activated with CD40L and IL-4 were also found to be relatively refractory to VSV-G pseudotyped onco-retroviral vectors (< 1%). However, higher transduction efficiencies could be achieved in activated primary human B lymphocytes using VSV-G pseudotyped lentiviral vectors instead (5%-6%). Contrary to the significant increase in mouse B-cell transduction efficiency with ecotropic vectors, the use of amphotropic onco-retroviral or lentiviral vectors did not increase transduction efficiency in primary human B cells. The present study shows that the transduction efficiency of onco-retroviral and lentiviral vectors in human and mouse B lymphocytes is pseudotype-dependent and challenges the widely held assumption that VSV-G pseudotyping facilitates gene transfer into all cell types.
authors.affiliation: Center for Molecular & Vascular Biology, University of Leuven, Leuven, B-3000, Belgium.
authors.forename: Wim
authors.initials: W
authors.lastname: Janssens
authors.forename: Marinee K L
authors.initials: MK
authors.lastname: Chuah
authors.forename: Luigi
authors.initials: L
authors.lastname: Naldini
authors.forename: Antonia
authors.initials: A
authors.lastname: Follenzi
authors.forename: Désiré
authors.initials: D
authors.lastname: Collen
authors.forename: Jean-Marie
authors.initials: JM
authors.lastname: Saint-Remy
authors.forename: Thierry
authors.initials: T
authors.lastname: VandenDriessche
mesh_terms: D000818:Animals; D001402:B-Lymphocytes / Q000378:metabolism*; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000633:toxicity; D006801:Humans; D016086:Lentivirus; D051379:Mice; D012190:Retroviridae; D014161:Transduction, Genetic