Gene transfer and expression of a non-viral polycation-based vector in CD4+ cells.
CD4-selective targeting of an antibody-polycation-DNA complex was investigated. The complex was synthesized with the anti-CD4 monoclonal antibody B-F5, polylysine268 (pLL) and either the pGL3 control vector containing the luciferase reporter gene or the pGeneGrip vector containing the green fluorescent protein (GFP) gene. B-F5-pLL-DNA complexes inhibited the binding of 125I-B-F5 to CD4+Jurkat cells, while complexes synthesised either without B-F5 or using a non-specific mouse IgG1 antibody had little or no effect. Expression of the luciferase reporter gene was achieved in Jurkat cells using the B-F5-pLL-pGL3 complex and was enhanced in the presence of PMA. Negligible luciferase activity was detected with the non-specific antibody complex in Jurkat cells or with the B-F5-pLL-pGL3 complex in the CD4- K-562 cells. Using complexes synthesised with the pGeneGrip vector, the transfection efficiency in Jurkat and K-562 cells was examined using confocal microscopy. More than 95% of Jurkat cells expressed GFP and the level of this expression was markedly enhanced by PMA. Negligible GFP expression was seen in K-562 cells or when B-F5 was replaced by a non-specific antibody. Using flow cytometry, fluorescein-labelled complex showed specific targeting to CD4+ cells in a mixed cell population from human peripheral blood. These studies demonstrate the selective transfection of CD4+ T-lymphoid cells using a polycation-based gene delivery system. The complex may provide a means of delivering anti-HIV gene therapies to CD4+ cells in vivo.
Puls
R
R
Department of Pharmacology, University of Western Australia, Western Australia, Australia.
Minchin
R
R
D000818:Animals; D000911:Antibodies, Monoclonal / Q000008:administration & dosage*; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology*; D015870:Gene Expression; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000008:administration & dosage* / Q000378:metabolism; D049452:Green Fluorescent Proteins; D006801:Humans; D019169:Jurkat Cells; D008156:Luciferases / Q000235:genetics; D008164:Luminescent Proteins / Q000235:genetics; D051379:Mice; D011107:Polylysine; D011485:Protein Binding
D000911:Antibodies, Monoclonal; D008164:Luminescent Proteins; D049452:Green Fluorescent Proteins; D011107:Polylysine; D008156:Luciferases