Figure 5

Inhibition of breast cancer cell proliferation, migration and invasion by a functional anti-ATP synthase α- subunit antibody. A, MDA-MB-231 and MCF-10F were plated at a density of 5,000 cells/well in serum-free media for overnight fasting. The α-subunit of ATP synthase was added at concentrations of 10 μg/ml (low dose), 50 μg/ml (medium dose) or 100 μg/ml (high dose). CCK-8 solution was added after 24 h, and the absorbance used to calculate percent proliferation was measured on a Thermo max plate reader at a wavelength of 490 nm. Growth of MDA-MB-231 cells treated with high dose MAb9E10 was significantly suppressed compared with IgG control treatment. MCF-10F cell proliferation was not inhibited by MAb9E10 treatment. B, Migration of MDA-MB-231 and MCF-10F cells were measured using wound-healing assays. MDA-MB-231 cells treated with MAb9E10 100 μg/ml showed lower migration ability comparing with mIgG control; ** P < 0.01. No significant inhibition of migration was observed when MCF-10F cells were treated with MAb9E10. C, Invasion of MDA-MB-231 cells was measured using transwell inserts coated with fibronectin (10 μg/ml). After treatment with anti-ATP synthase α-subunit antibody or control mIgG, cells that migrated through the filters onto the lower surface were fixed, stained and photographed (200×). In each individual experiment, the invaded cells were counted from at least three randomly selected fields. Results were averaged from at least three individual experiments. MDA-MB-231 cells treated with MAb9E10 100 μg/ml also showed lower invasion ability comparing with mIgG control (Figure 5C). Almost no inhibition of invasion when MCF-10F cells treated with MAb9E10. The bar graph displays means ± S.D; ** P < 0.01.