Fig. 8: MAFF enhances tumor metastasis through IL11/STAT3 pathways in vivo and their combined expression predicts patient survival.

Spontaneous lung metastasis model of MDA-MB-231 was studied by injecting 2 × 10⁶ cells into the mammary fat pad of NSG mice (2 × 10⁶ cells/75 μl/mouse). a In MAFF knockdown group, spontaneous lung metastasis was significantly decreased while IL11 overexpression rescued the reduction. Lung weight was measured right after sacrificing mice. Also, lung tumor area was determined using ImageJ after H&E staining (n = 10 for shSCR and shMAFF, n = 9 for shMAFF+IL11). One-Way ANOVA with multiple comparisons. b IL11 and IL6 levels in mouse serum were determined using ELISA. While IL11 was decreased with MAFF knockdown, elevated IL11 levels were confirmed in shMAFF with IL11 overexpression group. However, IL6 levels remained unchanged in every group (n = 10 for shSCR, shMAFF, n = 7 or 6 for shMAFF+IL11). Unpaired t-test. c, d Expression of phospho-STAT3 and MECA32 were evaluated from primary tumor tissues and we observed consistent changes with in vitro studies suggesting that MAFF and IL11 pathways regulated phospho-STAT3 and angiogenesis (n = 10 for shSCR, shMAFF, n = 9 for shMAFF+IL11). Arrows indicate positive MECA-32 staining for vessels. One-Way ANOVA with multiple comparisons. e Distant metastasis-free survival of breast cancer patients was evaluated using KM plotter⁶³. Combined expression of MAFF and IL11, MAFF, and BACH1, or MAFF, IL11, and BACH1 showed the correlation with patient survival, indicating prognostic significance of these factors. Log-rank. Graphs represent the mean per group and error bars represent the SEM. ns: not significant.