Volume 270,
Number 40,
Issue of October 06, pp. 23345-23351, 1995
©1995 by The American Society for Biochemistry and Molecular Biology, Inc.
Self-potentiation
of Ligand-Toxin Conjugates Containing Ricin A Chain Fused with Viral
Structures
(Received for publication, January 20, 1995; and in revised form, July
5, 1995)
Roberto
Chignola ,
Cristina
Anselmi,
Mauro
Dalla
Serra
,
Antonia
Franceschi ,
Giulio
Fracasso,
Marcella
Pasti ,
Elena
Chiesa,
J. Michael
Lord
,
Giuseppe
Tridente ,
Marco
Colombatti
A chimeric protein was obtained by fusing together the ricin
toxin A chain (RTA) gene and a DNA fragment encoding the N terminus of
protein G of the vesicular stomatitis virus. Chimeric RTA (cRTA)
retained full enzymic activity in a cell-free assay, but was 10-fold
less toxic against human leukemic cells than either native RTA (nRTA)
or unmodified recombinant RTA (rRTA). However, conjugates made with
cRTA and human transferrin (Tfn) showed 10-20-fold greater cell
killing efficacy than Tfn-nRTA or Tfn-rRTA conjugates despite
equivalent binding of the three conjugates to target tumor cells. As a
consequence, by fusion of the KFT25 peptide to the RTA sequence, the
specificity factor (i.e. the ratio between nonspecific and
specific cytotoxicity) of Tfn-cRTA was increased 90-240 times
with respect to those of Tfn-nRTA and Tfn-rRTA. cRTA interacted with
phospholipid vesicles with 15-fold faster kinetics than nRTA at acidic
pH. Taken together, our results suggest that the ability of vesicular
stomatitis virus protein G to interact with cell membranes can be
transferred to RTA to facilitate its translocation to the cell cytosol.
Our strategy may serve as a general approach for potentiating the
cytotoxic efficacy of antitumor immunotoxins.
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