Fig. 5.

Tbx3 is necessary and sufficient for breast CSC expansion. (A) (Left) Average percentage of CD44⁺/CD24⁻/ESA⁺ cells in MCF7 cultures transduced with lentiviruses encoding short hairpins targeting a scrambled sequence (Cntrl), GFP, or Tbx3 and treated with 1 nM 17-β-estradiol (E2) or vehicle (EtOH). *P < 0.0015. (Right) Average percentage of CD44⁺/CD24⁻/ESA⁺ cells in SUM149 and SUM159 cultures transduced with lentiviruses encoding short hairpins targeting Tbx3 and treated with recombinant FGF9. Data are mean ± SEM; n = 4 biological replicates. *P < 0.003; **P < 0.007. (B) Normalized tumorsphere-forming potential of SUM159, SUM149, or MCF7 cultures transduced with hairpins targeting a scramble sequence (Cntrl) or Tbx3. Data as mean ± SEM; n = 4 experiments. *P < 0.001; **P < 0.005; ***P < 0.02. (C) Normalized tumorsphere formation of breast cancer cells isolated from a primary human breast cancer (TUM177) transduced with lentiviruses containing two different short hairpin sequences targeting Tbx3. *P < 0.002; **P < 0.0008. (D) Normalized sphere-forming ability of immortalized human mammary epithelia cells (HMEC) or MCF7 cells ectopically overexpressing human Tbx3; n = 4 experiments, 2 biological replicates. *P = 0.002; **P = 0.003. (E) Tumor formation of MCF7 cells overexpressing Tbx3 or empty vector (EV) injected in limiting dilution into NOD/SCID mice. *Nonparametric χ² statistic was used as described in Fig. 1.