Extended Data Fig. 1: Acquired radioresistance in experimental brain metastasis.

a, Schema of the experimental design. BLI: bioluminescence imaging. b, Kaplan–Meier curves showing survival in nonirradiated and irradiated experimental groups described in Fig. 1b (n = 10, mice nonirradiated; n = 9, mice irradiated with 10 x 3 Gy; n = 9, mice irradiated with 3 × 5.5 Gy; n = 10, mice irradiated with 3 x 3 Gy). P value was calculated using log-rank (Mantel–Cox) test between nonirradiated and irradiated experimental groups (nonirradiated versus 10 x 3 Gy, P = 0.2019; nonirradiated versus 3 × 5.5 Gy, P = 0.1649; nonirradiated versus 3 x 3 Gy, P = 0.3057). c, Schema of experimental design. d, Representative images of brain organotypic cultures with metastatic cells 72 hours after irradiation at 10 Gy. e, Quantification of photon flux values from metastatic cells growing in organotypic brain cultures after irradiation normalized to preirradiated BLI values. Values are shown in box-and-whisker plots where every dot represents an independent culture and the line in the box corresponds to the median. The boxes go from the upper to the lower quartiles and the whiskers go from the minimum to the maximum value (n = 9, nonirradiated brain slices with E0771-BrM; n = 9, irradiated brain slices with E0771-BrM). P value is calculated using two-tailed t test. f, Representative graph of positively enriched gene sets depicting induction of a stem cell signature among a preranked list of differentially expressed genes from resistant culture preparations ex vivo (Supplementary Table 1). The green curve corresponds to the Enrichment Score, while the normalized enrichment score (NES) and the FDR are shown below the graph. g, Representative images of oncospheres 72 hours after irradiation at 10 Gy. Scale bar, 250 µm. h, Quantification of oncosphere area. Values are shown in box-and-whisker plots where every dot represents a different well from which the mean area of all oncospheres were quantified, from an independent culture and the line in the box corresponds to the median. The boxes go from the upper to the lower quartiles and the whiskers go from the minimum to the maximum value (n = 4, nonirradiated wells with E0771-BrM oncospheres; n = 4, irradiated wells with E0771-BrM oncospheres). P value is calculated using two-tailed t test. i, Representative images of co-cultures between E0771-BrM and astrocytes (GFAP + ) 72 h after irradiation (10 Gy). Scale bar, 100 μm j, Quantification of GFAP- BrM cells after irradiation normalized to their respective nonirradiated controls from experiment in i. Values are shown in box-and-whisker plots where every dot represents an independent culture and the line in the box corresponds to the median. The boxes go from the upper to the lower quartiles and the whiskers go from the minimum to the maximum value (n = 4, in each respective experimental condition). P value is calculated using two-tailed t test.