{"database":"biostudies-literature","file_versions":[],"scores":null,"additional":{"omics_type":["Unknown"],"volume":["100(3)"],"submitter":["Leong SR"],"pubmed_abstract":["DNA delivery of IL-12 has shown promise in reducing the toxic side effects associated with administration of recombinant human (h)IL-12 protein while maintaining the ability to inhibit tumor growth and abolish tumor metastases in animal models. We have developed a more potent version of IL-12 by using DNA shuffling and screening to improve its expression in human cells and specific activity on human T cells. The most improved evolved IL-12 (EvIL-12) derived from seven mammalian genes encoding both the p35 and p40 subunits of IL-12 showed a 128-fold improvement in human T cell proliferation compared with native hIL-12 during the initial screening of supernatants from transected cells. When purified hIL-12 and EvIL-12 proteins were compared in vitro in human T cell proliferation and Th1 differentiation assays, it was demonstrated that EvIL-12 exhibited a concomitant 10-fold increase in the specific activity of the protein compared with hIL-12. Furthermore, DNA shuffling improved the level of expression and homogeneity of the heterodimer synthesized by 293 human embryonic kidney cells transfected with EvIL-12 by at least 10-fold. Molecular analysis of the variant revealed strategic placement of amino acid substitutions that potentially may facilitate heterodimer formation and product expression. The enhanced expression and biological activity of EvIL-12 may improve the effectiveness of IL-12 gene-based vaccines and therapeutics without the toxic side effects sometimes associated with hIL-12 protein administration."],"journal":["Proceedings of the National Academy of Sciences of the United States of America"],"pagination":["1163-8"],"full_dataset_link":["https://www.ebi.ac.uk/biostudies/studies/S-EPMC298744"],"repository":["biostudies-literature"],"pubmed_title":["Optimized expression and specific activity of IL-12 by directed molecular evolution."],"pmcid":["PMC298744"],"pubmed_authors":["Dawes G","Leong SR","Ong R","Punnonen J","Stemmer WP","Chang JC"],"additional_accession":[]},"is_claimable":false,"name":"Optimized expression and specific activity of IL-12 by directed molecular evolution.","description":"DNA delivery of IL-12 has shown promise in reducing the toxic side effects associated with administration of recombinant human (h)IL-12 protein while maintaining the ability to inhibit tumor growth and abolish tumor metastases in animal models. We have developed a more potent version of IL-12 by using DNA shuffling and screening to improve its expression in human cells and specific activity on human T cells. The most improved evolved IL-12 (EvIL-12) derived from seven mammalian genes encoding both the p35 and p40 subunits of IL-12 showed a 128-fold improvement in human T cell proliferation compared with native hIL-12 during the initial screening of supernatants from transected cells. When purified hIL-12 and EvIL-12 proteins were compared in vitro in human T cell proliferation and Th1 differentiation assays, it was demonstrated that EvIL-12 exhibited a concomitant 10-fold increase in the specific activity of the protein compared with hIL-12. Furthermore, DNA shuffling improved the level of expression and homogeneity of the heterodimer synthesized by 293 human embryonic kidney cells transfected with EvIL-12 by at least 10-fold. Molecular analysis of the variant revealed strategic placement of amino acid substitutions that potentially may facilitate heterodimer formation and product expression. The enhanced expression and biological activity of EvIL-12 may improve the effectiveness of IL-12 gene-based vaccines and therapeutics without the toxic side effects sometimes associated with hIL-12 protein administration.","dates":{"release":"2003-01-01T00:00:00Z","publication":"2003 Feb","modification":"2021-02-20T06:56:40Z","creation":"2019-03-27T00:36:05Z"},"accession":"S-EPMC298744","cross_references":{"pubmed":["12529500"],"doi":["10.1073/pnas.0237327100"]}}