Genetic engineering of an immunotoxin to eliminate pulmonary vascular leak in mice


Vascular leak syndrome is a major and often dose-limiting side effect of immunotoxins and cytokines. We postulated that this syndrome is initiated by damage to vascular endothelial cells. Our earlier studies identified a three–amino acid motif that is shared by toxins, ribosome-inactivating proteins, and interleukin-2, all of which cause this problem. We have now generated a panel of recombinant ricin A chains with mutations in this sequence or in amino acids flanking it in the three-dimensional structure. These have been evaluated alone and as immunotoxins for activity, ability to induce pulmonary vascular leak in mice, pharmacokinetics, and activity in tumor-xenografted mice. One mutant was comparable to the ricin A chain used before in all respects except that it did not cause vascular leak at the same dose and, when used as an immunotoxin, was more effective in xenografted SCID mice.

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Figure 1: Ribbon diagram of RTA.
Figure 2: In vivo PVL of RFB4-RTA immunotoxins.
Figure 3: Effect of equitoxic doses of immunotoxins prepared with N97A vs. dgRTA on the survival of SCID-Daudi mice.

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We thank Lien Le, Ming-Mei Liu, Yuen Chinn, Ana Firan, Steve Ruback, and Stephanie Tuggle for exceptional technical assistance. We thank Shannon Flowers and Linda Owens for secretarial assistance and M. Lord for providing the rRTA clone. We are indebted to Jonathan Uhr and John Schindler for helpful comments concerning the manuscript. This work was supported by US National Institutes of Health grant CA-77701 and a grant from the Higher Education Coordinating Board of the state of Texas.

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Correspondence to Ellen S. Vitetta.

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Smallshaw, J., Ghetie, V., Rizo, J. et al. Genetic engineering of an immunotoxin to eliminate pulmonary vascular leak in mice. Nat Biotechnol 21, 387–391 (2003).

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