Construction and Expression of Novel Immunotoxin cpIL-4(13D)-PE38KDEL with Increased Activity

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J.-X. Cui^1,2, J.-F. Ji³, A.-G. Lv^1,2, and W.-F. Wu^1,2*

¹Institute of Applied Ecology, Chinese Academy of Sciences, No. 72, Wenhua Road, Shenyang City, Liaoning Province, 110016, P. R. China; fax: +86-24-23840383; E-mail: cjxlib@hotmail.com; wwf222@hotmail.com

²Graduate School of the Chinese Academy of Sciences, Beijing 100039, P. R. China; E-mail: lvag66@hotmail.com

³Beckman Research Institute, City of Hope National Medical Center, 1500 Duarte Rd. Duarte, CA 91010, USA; E-mail: jijfei@hotmail.com

^* To whom correspondence should be addressed.

Received March 31, 2004; Revision received July 2, 2004

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Interleukin-4 receptors (IL-4Rs) are expressed on a wide variety of human cancer cells, and therefore it may be a good option to treat IL-4R-bearing tumors with IL-4-fusing immunotoxins. In this study, the gene encoding human interleukin-4 mutein cpIL-4(13D) was obtained through overlapping polymerase chain reaction. A chimeric immunotoxin was constructed by genetically fusing the mutein cpIL-4(13D) to a modified version of Pseudomonas exotoxin A (PE38KDEL) and was expressed in Escherichia coli AD494 (DE3). The expression level of the fusion protein was about 30% of the total bacterial protein assessed by SDS-PAGE analysis. After purification by affinity chromatography and anion exchange chromatography, the chimeric protein was tested for its cytotoxicity. Our data show that cpIL-4(13D)-PE38KDEL has improved cytotoxicity on IL-4R-bearing tumor cells in comparison with other IL-4-fusing immunotoxins and might be useful in treating tumors with a large number of IL-4Rs.

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KEY WORDS: immunotoxin, interleukin-4, Pseudomonas exotoxin A

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