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IL13R\u03b11, the other binding partner of IL13, is ubiquitously expressed in healthy tissue, raising concerns about the therapeutic window of systemic administration. IL13 mutants with diminished binding affinity to IL13R\u03b11 were previously generated by structure-guided protein engineering. In this study, two such variants, termed C4 and D7, are characterized for their ability to mediate IL13R\u03b12-specific response as binding domains for CAR T cells. Despite IL13R\u03b11 and IL13R\u03b12 sharing similar binding interfaces on IL13, mutations to IL13 that decrease binding affinity for IL13R\u03b11 did not drastically change binding affinity for IL13R\u03b12. Micromolar affinity to IL13R\u03b11 was sufficient to pacify IL13-mutein CAR T cells in the presence of IL13R\u03b11-overexpressing cells in\u00a0vitro. Interestingly, effector activity of D7 CAR T cells, but not C4 CAR T cells, was demonstrated when cocultured with IL13R\u03b11\/IL4R\u03b1-coexpressing cancer cells. While low-affinity interactions with IL13R\u03b11 did not result in observable toxicities in mice, in\u00a0vivo biodistribution studies demonstrated that C4 and D7 CAR T cells were better able to traffic away from IL13R\u03b11+ lung tissue than were wild-type (WT) CAR T cells. These results demonstrate the utility of structure-guided engineering of ligand-based binding domains with appropriate selectivity while validating IL13-mutein CARs with improved selectivity for application to systemic IL13R\u03b12-expressing malignancies.<\/jats:p>","DOI":"10.1073\/pnas.2112006119","type":"journal-article","created":{"date-parts":[[2022,8,8]],"date-time":"2022-08-08T15:16:07Z","timestamp":1659971767000},"update-policy":"https:\/\/doi.org\/10.1073\/pnas.cm10313","source":"Crossref","is-referenced-by-count":30,"title":["Engineered IL13 variants direct specificity of IL13R\u03b12-targeted CAR T cell therapy"],"prefix":"10.1073","volume":"119","author":[{"given":"Lawrence A.","family":"Stern","sequence":"first","affiliation":[{"name":"Department of Hematology &amp; Hematopoietic Cell Transplantation, Beckman Research Institute, City of Hope National Medical Center, Duarte, CA 91010"},{"name":"Department of Immuno-Oncology, Beckman Research Institute, City of Hope National Medical Center, Duarte, CA 91010"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7425-6074","authenticated-orcid":false,"given":"Sharareh","family":"Gholamin","sequence":"additional","affiliation":[{"name":"Department of Hematology &amp; 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