Recombinant adenoviral delivery for in vivo expression of scFv antibody fusion proteins.
Antibodies and their recombinant fragments have enormous potential for therapy of malignant and other diseases, but there can be problems associated with their production and purification in the quantities required for therapeutic use. We investigated the use of gene therapy for the production of such recombinant antibody fragments in vivo. We generated two recombinant adenoviruses expressing the single chain Fvs (scFvs) fused to murine GM-CSF (mGM-CSF). The scFvs used are MFE-23 which binds to a human tumour-associated marker carcino-embryonic antigen (CEA) and B1.8 which binds the hapten 4-hydroxy-3-nitro-5-iodo-phenylacetyl (NIP). Using scFvs to target GM-CSF to tumour cells should reduce the systemic toxicity of GM-CSF but retain its ability as a cytokine to induce systemic immune responses to tumour targets. Cell lines infected with the recombinant adenoviruses in vitro express and secrete high levels of the scFv.mGM-CSF fusion proteins. The scFv retains specificity while the mGM-CSF portion is fully bioactive and there is no detectable degradation of the fusion product. We also demonstrated effective in vivo expression of the scFv.mGM-CSF proteins. C57BI/6 mice injected intravenously with the adenovirus encoding the MFE-23.mGM-CSF fusion produce high levels of the fusion protein by 2 days after infection. The scFv.mGM-CSF protein can be detected in the serum, at biologically active levels, for at least 20 days and the level of protein produced is related to the amount of adenovirus injected. This approach has the potential to streamline the testing of the many therapeutic strategies based on recombinant scFvs and we are currently testing these constructs in an animal model for antitumour activity.
Whittington
H A
HA
University Department of Oncology, Bristol Oncology Centre, UK.
Ashworth
L J
LJ
Hawkins
R E
RE
D000256:Adenoviridae; D000818:Animals; D019677:Artificial Gene Fusion; D002455:Cell Division; D015870:Gene Expression; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000235:genetics*; D006367:HeLa Cells; D006801:Humans; D007128:Immunoglobulin Fragments; D007167:Immunotherapy; D051379:Mice; D008810:Mice, Inbred C57BL
D007128:Immunoglobulin Fragments; D016178:Granulocyte-Macrophage Colony-Stimulating Factor