Evidence for nonspecific adsorption of targeted retrovirus vector particles to cells.
The ability to specifically target a cell-type is important for the development of vectors for in vivo gene therapy. In order to produce retrovirus vectors targeting ovarian cancer cells, which specifically overexpress alpha folate receptor (alphaFR), a single chain antibody was fused as an N-terminal extension of the ecotropic and amphotropic murine leukemia virus (MLV) envelope glycoproteins. Vector particles bearing the modified glycoproteins were produced and analysed. Although conventional FACS studies indicated that viral particles bearing the modified Env could bind to ovarian cancer cells, targeted infection was not achieved. The initial step of virus-cell interaction was further studied using an immunofluorescence technique, which allows visualisation of single retrovirus particles. Vectors bearing chimeric or wild-type glycoproteins bound equally well to cells with or without the targeted receptor, although soluble chimeric glycoproteins bound specifically to FBP. Our results indicate that the incorporation of specific ligands to the virus envelope does not necessarily result in significant enhancement of vector particle binding. A similar interaction was also observed using Env-defective virus particles, suggesting that cellular factors incorporated into the lipid envelope play a dominant role in promoting initial adsorption of virus particles to cells. Significant implications arise from these observations on the interpretation of previous reports on 'targeted' vectors, and for the development of vectors for in vivo gene therapy protocols.
Pizzato
M
M
Wohl Virion Centre, Windeyer Institute of Medical Sciences, University College London, UK.
Blair
E D
ED
Fling
M
M
Kopf
J
J
Tomassetti
A
A
Weiss
R A
RA
Takeuchi
Y
Y
D016475:3T3 Cells; D000327:Adsorption; D000818:Animals; D000911:Antibodies, Monoclonal; D015153:Blotting, Western; D002352:Carrier Proteins / Q000235:genetics* / Q000276:immunology; D005260:Female; D005434:Flow Cytometry; D058973:Folate Receptors, GPI-Anchored; D018390:Gene Targeting; D005803:Genes, Immunoglobulin; D005818:Genetic Engineering; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000378:metabolism*; D007128:Immunoglobulin Fragments; D007135:Immunoglobulin Variable Region; D009052:Leukemia Virus, Murine / Q000235:genetics*; D051379:Mice; D010051:Ovarian Neoplasms / Q000628:therapy*; D011956:Receptors, Cell Surface
D000911:Antibodies, Monoclonal; D002352:Carrier Proteins; D058973:Folate Receptors, GPI-Anchored; D007128:Immunoglobulin Fragments; D007135:Immunoglobulin Variable Region; D011956:Receptors, Cell Surface