Cytotoxic effects of vascular smooth muscle cells of the chimeric toxin, heparin binding TGF alpha-Pseudomonas exotoxin.
OBJECTIVE
Smooth muscle cell proliferation appears to be very important in restenosis after angioplasty. A chimeric toxin created by genetically fusing the gene encoding TGF alpha (targets the EGF receptor) to the gene encoding Pseudomonas exotoxin (PE) preferentially kills rapidly proliferating smooth muscle cells. Recently, a heparin binding EGF-like growth factor (HB-EGF) has been identified. The HB domain enhances the mitogenic activity for smooth muscle cells. The purpose of this study was to design a new chimeric toxin, having both heparin binding and EGF receptor binding function, and to determine whether it is more cytotoxic to smooth muscle cells.


METHODS
By recombinant DNA techniques, a new chimeric toxin, HB-TGF alpha-PE4EKDEL, was synthesised. Cytotoxic assays were performed by assessing the capacity to inhibit protein synthesis of rat vascular smooth muscle cells.


RESULTS
The toxin preferentially killed rapidly proliferating smooth muscle cells (p < 0.025). The HB domain increased the cytotoxicity of the molecule when compared to the other chimeric toxins tested against smooth muscle cells. The cytotoxic effect of the new molecule was significantly decreased by exogenously added heparin (p < 0.05).


CONCLUSIONS
The presence of a heparin binding domain increases the smooth muscle cell cytotoxicity of the TGF alpha fusion toxin, perhaps because HB-TGF alpha-PE4EKDEL functions as a molecule with two ligands. It will be important to determine whether the greater smooth muscle cell cytotoxicity that exists in vitro will facilitate the specific targeting and killing of rapidly proliferating cells in vivo.
Fu
Y M
YM
National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892.
Mesri
E A
EA
Yu
Z X
ZX
Kreitman
R J
RJ
Pastan
I
I
Epstein
S E
SE
D000818:Animals; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D066246:ErbB Receptors / Q000378:metabolism; D005098:Exotoxins / Q000494:pharmacology*; D006493:Heparin / Q000378:metabolism; D000922:Immunotoxins / Q000494:pharmacology*; D007930:Leucine / Q000378:metabolism; D009132:Muscles / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D011485:Protein Binding; D051381:Rats; D017207:Rats, Sprague-Dawley; D011993:Recombinant Fusion Proteins / Q000494:pharmacology*; D016211:Transforming Growth Factor alpha / Q000494:pharmacology*; D000097668:Pseudomonas aeruginosa Exotoxin A
D066246:ErbB Receptors; D005098:Exotoxins; D006493:Heparin; D000922:Immunotoxins; D007930:Leucine; D011993:Recombinant Fusion Proteins; D016211:Transforming Growth Factor alpha; C081440:transforming growth factor(alpha)-Pseudomonas exotoxin A (35); D000097668:Pseudomonas aeruginosa Exotoxin A