Cw. Liao et al., A TARGET-SPECIFIC CHIMERIC TOXIN COMPOSED OF EPIDERMAL GROWTH-FACTOR AND PSEUDOMONAS EXOTOXIN A WITH A DELETION IN ITS TOXIN-BINDING DOMAIN, Applied microbiology and biotechnology, 43(3), 1995, pp. 498-507
We have fused the epidermal growth factor (EGF) to the amino terminus
of Pseudomonas exotoxin A (PE) to create a cytotoxic agent, designated
EGF-PE, which preferentially kills EGF-receptor-bearing cells. In thi
s study, we analyzed the effect of the Ia domain, the binding domain,
of PE on the cytotoxicity of ECF-PE towards EGF-receptor-bearing cells
and tried to develop a more potent EGF-receptor-targeting toxin. EGF-
PE molecules with sequential deletions at the amino terminus of PE wer
e constructed and expressed in E. coli strain BL21(DE3). The cytotoxic
ity of these chimeric toxins was then examined. Our results show that
the amino-terminal and carboxy-terminal regions of the Ia domain of PE
are important for the cytotoxicity of a PE-based targeting toxin. To
design a more potent PE-based EGF-receptor-targeting toxin, a chimeric
toxin, named EGF-PE(Delta 34-220), which had most of the Ia domain de
leted but retained amino acid residues 1-33 and 221-252 of this domain
, was constructed. EGF-PE(Delta 34-220) has EGF-receptor-binding activ
ity but does not show PE-receptor-binding activity and is mildly cytot
oxic to EGF-receptor-deficient NR6 cells. As expected, EGF-PE(Delta 34
-220) is a more potent cytotoxic agent towards EGF-receptor-bearing ce
lls than EGF-PE(Delta 1-252), where the entire Ia domain of PE was del
eted. In addition, EGF-PE(Delta 34-220) was shown to be extremely cyto
toxic to EGF-receptor-bearing cancer cells, such as A431, CE81T/VGH, a
nd KB-3-1 cells. We also found that EGF-PE(Delta 34-220) was highly ex
pressed in BL21(DE3) and could be easily purified by urea extraction.
Thus, EGF-PE(Delta 34-220) can be a useful cytotoxic agent towards EGF
-receptor-bearing cells.