Evolution of a cytokine using DNA family shuffling

DNA shuffling of a family of over 20 human interferon-alpha (Hu-IFN-alpha) genes was used to derive variants with increased antiviral and antiprolifer ation activities in murine cells, A clone with 135,000-fold improved specif ic activity over Hu-IFN-alpha 2a was obtained in the first cycle of shuffli ng, After a second cycle of selective shuffling, the most active clone was improved 285,000-fold relative to Hu-IFN-alpha 2a and 185-fold relative to Hu-IFN-alpha 1, Remarkably, the three most active clones were more active t han the native murine IFN-alpha s. These chimeras are derived from up to fi ve parental genes but contained no random point mutations. These results de monstrate that diverse cytokine gene families can be used as starting mater ial to rapidly evolve cytokines that are more active, or have superior sele ctivity profiles, than native cytokine genes.