Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Engineered IL13 variants direct specificity of IL13Rα2-targeted CAR T cell therapy

doi: 10.1073/pnas.2112006119

Figure Lengend Snippet: C4 and D7 IL13-mutein CAR T cells showed reduced recognition of IL13Rα1. IL13Rα1-depedendent degranulation ( A ) and IFN-γ production ( B ) for IL13 WT, E12Y, C4, and D7 CAR T cells cocultured with IL13Rα1+ A549, HT1080 parental, or HT1080 engineered to express either IL13Rα1 or both IL13Rα1 and IL4Rα as described in . ( C ) Secreted IFN-γ production by indicated T cell lines in response to increasing concentrations of immobilized rhIL13Rα1-Fc (mean ± SD; n = 3 wells). ( D ) IL13Rα1-dependent cell killing of the indicated CAR and target lines at a 1:10 E:T ratio for 2 days. A – D are representative of at least three independent experiments. ( E ) Winn assay evaluating A549 (0.1 × 10 6 ) tumor engraftment after coculture with CAR and mock T cell lines (E:T ratio of 10:1; 2 h preincubation and engraftment on day 0; n = 4). ( F ) NSG mice xenotransplanted with 0.5 × 10 6 HT1080 IL13Rα1/IL4Rα in a 1:1 ratio of media to Matrigel, followed by treatment with no T cells, 5 × 10 6 mock T cells, or the various mutein CAR T cells on day 4. All data shown on A – F are means ± SEM (**** P < 0.0001, *** P < 0.005, ** P < 0.01, * P < 0.05).

Article Snippet: For enzyme-linked immunosorbent assays (ELISAs), T cells were cultured overnight at 5 × 10 3 effectors per well in flat-bottom, 96-well plates (catalog 3361, Corning) that had been coated with 500, 250, 125, 62.5, or 31.25 ng/well rhIL13Rα1-Fc chimera (catalog 146-IR-100, R&D Systems) or IL13Rα2-Fc chimera (catalog 7147-IR-100, R&D Systems).

Techniques: