Expression of chimeric envelope proteins in helper cell lines and integration into Moloney murine leukemia virus particles.
New retroviral constructs with a grafted specificity of infection could become useful gene delivery vehicles with applications in systemic gene therapy. We have constructed retroviral vectors to target gene transfer to human tumor cells. Chimeric envelope proteins have been expressed to obtain viral particles with a defined specificity of infection. Two tumor cell-specific recognition domains were cloned and fused with the viral envelope gene. A recognition domain specific for ErbB-2 expressing tumor cells was derived from a monoclonal antibody directed against the ErbB-2 receptor in the form of a single chain antibody domain (scFv-erbB-2). The receptor binding domain was derived from the heregulin gene (HRG70). This domain provides recognition specificity for ErbB-3 and ErbB-4 receptor expressing tumor cells. The recognition domains were inserted at the amino terminal end into the MoMLV envelope gene. Helper cell lines were established which express the recombinant envelope protein genes, the gag and pol genes and packageable retroviral RNA. The analysis of the helper cell line revealed that the recombinant ErbB-2 scFv-envelope protein was expressed, but not incorporated into viral particles. The scFv-erbB-2 envelope protein was not inserted into the cell membrane and the assembly of retroviral particles was not completed. In contrast, the HRG70-envelope protein was expressed on the surface of the helper cells and incorporated into retroviral particles. The HRG70-envelope protein, however, did not alter the host range of infection. Only cells expressing the ecotropic viral receptor could be infected.
Schnierle
B S
BS
Institute for Experimental Cancer Research, Tumor Biology Center, Freiburg, Germany.
Moritz
D
D
Jeschke
M
M
Groner
B
B
D016475:3T3 Cells; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D003001:Cloning, Molecular; D017931:DNA Primers / Q000235:genetics; D015870:Gene Expression; D015686:Gene Products, env / Q000235:genetics*; D015316:Genetic Therapy; D005822:Genetic Vectors; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D008979:Moloney murine leukemia virus / Q000235:genetics*; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000235:genetics; D011991:Receptors, Virus / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics; D006377:T-Lymphocytes, Helper-Inducer / Q000378:metabolism* / Q000821:virology; D014162:Transfection; D014407:Tumor Cells, Cultured
D017931:DNA Primers; D015686:Gene Products, env; D018719:Erb-b2 Receptor Tyrosine Kinases; D011991:Receptors, Virus; D011993:Recombinant Fusion Proteins