Cloning and cytotoxicity of fusion proteins of EGF and angiogenin.
Targeted toxins represent a new approach to specific cytocidal therapy. Immunotoxins based on plant and microbial toxins are very immunogenic. To develop a targeted therapy that is less immunogenic and easily invades target tissues, four fusion proteins containing human angiogenin targeted by human EGF have been constructed. EGF is a single chain polypeptide, which binds to epidermal growth factor receptor (EGFR) and is known to be internalized by endocytosis. Angiogenin has been separately fused either at the amino terminus or the carboxyl terminus of EGF via linkers, giving rise to angiogenin-gly-EGF, angiogenin-(gly)4ser-EGF and EGF-angiogenin, EGF-gly-angiogenin, respectively. The fusion proteins were over-expressed in Escherichia coli and purified from periplasmic eluents by affinity chromatography. EGF-angiogenin and EGF-gly-angiogenin maintained receptor-binding activity of EGF and RNase activity of angiogenin in a single peptide and actively inhibited growth of human EGFR-positive target cells in culture. They are expected to have a very low immunogenic potential in humans because of their endogenous origin and also to have another potential therapeutic advantage because these fusion proteins may have overcome conventional immunotoxin and possess increased ability to penetrate because of their small size.
D043925:Angiogenesis Inducing Agents; D000818:Animals; D016466:CHO Cells; D016923:Cell Death / Q000187:drug effects*; D002455:Cell Division / Q000187:drug effects; D003001:Cloning, Molecular; D006224:Cricetinae; D004815:Epidermal Growth Factor / Q000235:genetics* / Q000494:pharmacology; D066246:ErbB Receptors / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D006801:Humans; D000922:Immunotoxins; D011506:Proteins / Q000235:genetics* / Q000494:pharmacology; D011993:Recombinant Fusion Proteins / Q000276:immunology / Q000378:metabolism / Q000494:pharmacology*; D012259:Ribonuclease, Pancreatic; D012260:Ribonucleases / Q000378:metabolism; D014407:Tumor Cells, Cultured