Fig. 4: The SLIT2-ROBO1 axis facilitates metastasis in the liver.

a–c Representative CT combined with 3D organ reconstruction bioluminescence imaging to assess liver metastasis modelled by Kpc1199^Robo1-FL (b) or Panc02^Robo1-FL (c) cells in Slit2 ^fl/fl or Slit2 ^fl/fl/Alb-Cre mice with or without Lenti-loxp-Slit2 injection: Premetastatic niche (PMN) (upper) and macrometastatic niche (MMN) (lower) (n = 5 mice per group, mean ± SEM.; two-tailed unpaired Student’s t test). Scale colour bars: 2.00 × 10⁵−2.00 × 10⁷. d, e Representative CT combined with 3D organ reconstruction bioluminescence imaging displaying Kpc1199^Robo1-FL (upper) and Panc02^Robo1-FL (lower) cell-injected liver metastasis mouse models administered IgG or ROBO1 neutralizing antibody (n = 5 mice per group, mean ± SEM.; two-tailed unpaired t test). Scale colour bar: 2.00 × 10⁵−2.00 × 10⁷. Red, reconstructed liver; orange, reconstructed spleen; yellow, signals of tumour niches. (f–h) The administration of IgG or ROBO1 neutralizing antibody to the Kras^G12D/+/Trp53^R172H/+/Pdx1-Cre (KPC) mouse model from week 12 is shown. The number of metastatic niches in the liver with diameters over or under 500 μm in each group was determined (g), and the percentage of ROBO1⁺ metastatic niches in each group was assessed (h) (n = 5 mice per group, mean ± SEM.; two-tailed unpaired t test). (i) IHC-P staining of SLIT2 and ROBO1 in serial sections of livers with metastasis from KPC mice with or without ROBO1 neutralizing antibody treatment (n = 5 mice per group, 3 fields assessed per sample). Scale bar, 200 μm. (j–l) IHC-P staining of SLIT2 and ROBO1 in serial sections of livers with metastasis from KPC mice that underwent ROBO1 neutralizing antibody treatment starting from PMN (week 12-) or MMN (week 16-) occurrence. The number of metastatic niches in the liver with diameters over or under 500 μm in each group was determined (k), and the percentage of ROBO1⁺ metastatic niches in each group was assessed (l) (n = 5 mice per group, mean ± SEM.; two-tailed unpaired t test). (m) IHC-P staining of SLIT2 in serial sections of KPC liver metastases that experienced ROBO1 neutralizing antibody treatment starting from PMN (week 12) or MMN (week 16) occurrence (n = 5 mice per group, 3 fields assessed per sample). Scale bar, 200 μm. Source data are provided in the Source Data file.