Human interleukin-18 (hIL-18), initially cloned as an IFN-gamma-inducing factor, has a key role in many inflaininatory diseases. We have previously developed a high production system for correctly folded active hIL-18 protein, leading to the revelation of the 3D-structure and the receptor binding mode. These findings can strongly indicate the experimental and medical applications of IL-18; however, the recombinant protein is prone to be inactivated forming multimers. Recently, therapeutic approaches using recombinant IL-18 have shown the effectiveness for treatment of cancer: indicating the necessity of a more stable protein for therapy with intertrial reliability. Here we have generated a highly stable hIL-18 with replacement of cysteine by serine based on the tertiary structure and the binding mechanism, retaining the biological activity. Similar rational designs can be applied to develop new therapeutic molecules of other cytokines. (C) 2004 Elsevier Inc. All rights reserved.