Fig. 5

Neo-2/15 enhances the survival and persistence of CAR-NK cells in TME. a Quantification of CAR-NK cells in tumor tissues of orthotopic mice model. Data are presented as mean ± SD (***P < 0.001, n = 3). b Proportion of CAR-NK cells in the peripheral blood of mice in (a) was analyzed by flow cytometry. Data are presented as mean ± SD (*P < 0.05, ***P < 0.001, n = 6). c The number of CAR-NK cells in ascites was analyzed by flow cytometry. Data are presented as mean ± SD (*P < 0.05, n = 3). d Schematic of the intratumoral injection of CAR-NK cells treated with IL-2 or Neo-2/15 (left), and quantification of intratumoral BBζ at specified times post-administration (right). Data are presented as mean ± SD (*P < 0.05, **P < 0.01, n = 3). BBζ, stimulated with IL-2 or Neo-2/15 and pre-treated without (e) or with (f) 10058-F4, were co-cultured with tumor cells over specified durations. Western blot wase used to analyze the expression of Bax, Bak, Bcl-2, and Bcl-xl in BBζ. g Representation IF images for detecting the nuclear translocation of XBP1s in IL-2 or Neo-2/15 stimulated BBζ co-cultured with AsPC-1 cells for 8 hours at the E:T ratio of 1:1. XBP1s is marked in red, CD56 in green, and nuclei are stained with DAPI (blue). h IL-2 or Neo-2/15 expanded BBζ were pretreated with or without 10058-F4, following co-cultured with tumor cells for the 12 hours. Western blot analysis was performed to measure the expression of CHOP, caspase 3/12, total and phosphorylated IRE1α (left). XBP1s were analyzed in separated cytoplasmic and nuclear fractions, with GAPDH used as the cytoplasmic loading control and Lamin B used as the nuclear loading control (right)