miR-26a chimera inhibits mouse breast cancer growth and protects from chemotherapy-induced myelosuppression. (A) The binding of c-Kit aptamer to TUBO cells. (B) miR-26a chimera suppressed the growth and induced apoptosis of TUBO cells. Additional 5-FU treatment (1 μg/ml) enhanced these effects of miR-26a chimera. Cell count (left) and percentage of annexin V⁺ cells (%) (right) of the cells cultured with miR-26a chimera or the control (ctrl) were determined by flow cytometry. Asterisks denote the significant difference between ctrl chimera vs miR-26a chimera and between ctrl chimera plus 5-FU vs miR-26a chimera plus 5FU. Data (mean ± SD) were pooled from 2 experiments. (C-E) miR-26a, Ezh2, and Bak1 expressions in c-Kit⁺ or c-Kit⁻ cells harvested from TUBO-derived tumors in BALB/c mice treated with miR-26a chimera (670 pmol per 20 g) for 3 days. Data (mean ± SD) were pooled from 2 experiments involving a total of 6 mice per group. (F,G) miR-26a and Bak1 expressions in BM detected by quantitative polymerase chain reaction at 3 days after IV injection with the miR-26a chimera (670 pmol per 20 g). Data (mean ± SD) were pooled from 2 experiments involving a total of 6 mice per group. (H) Bak1 expression in c-Kit⁺ BM cells harvested from tumor (TUBO)-bearing BALB/c mice treated with miR-26a chimera (670 pmol per 20 g; 3 times) and 5-FU 50 mg/kg. Representative plots of Bak1 levels in cKit⁺ BM cells at day 3 after 5-FU treatment (left). Mean fluorescence intensity (MFI) of Bak1 staining in cKit⁺ BM cells (right). Data (mean ± SD) were pooled from 2 experiments involving a total of 3 mice per group. (I) MFI of Bak1 staining in cKit⁺ tumor cells harvested from the tumor (TUBO)-bearing BALB/c mice treated with miR-26a chimera (670 pmol per 20 g; 3 times) and 5-FU 50 mg/kg. Data (mean ± SD) were pooled from 2 experiments involving a total of 3 mice per group. (J) Tumor volume over time. BALB/c mice bearing TUBO cells were treated with miR-26a chimera (670 pmol per 20 g; 5 times, gray arrows) and 5-FU 50 mg/kg (3 times, blue arrows). Data (mean ± SD) were pooled from 2 experiments involving a total of 6 mice per group. There was significant difference between the 5-FU–only group vs miR-26a chimera–only group (2-way repeated-measures analysis of variance [ANOVA] followed by Bonferroni post-test for day 0 to day 18; interaction P < .001). (K) Tumor volume over time after combinational treatment with 5-FU and chimeras. Data (mean ± SD) were pooled from 2 experiments involving a total of 6 mice per group. There was a significant difference between 5-FU plus ctrl chimera group vs 5-FU plus miR-26a chimera group (2-way repeated-measures ANOVA followed by Bonferroni post-test for day 0 to day 21; interaction P < .0001). (L) The numbers of WBCs and platelets (PLTs) in the tumor bearing mice 5 days after 5-FU treatment. Data (mean ± s.d.) pooled from 2 experiments involving a total of 6 mice per group. *P < .05, **P < .01. Error bars indicate SD.