Autophagy: a novel pathway for inhibitor of apoptosis protein-like protein-2 promoting breast cancer cell proliferation

Abstract Inhibitors of apoptosis protein-like protein-2 (ILP-2), which is a high expression in breast cancer, can cause apoptosis deficiency to promote proliferation on breast cancer cells. It is unclear that whether ILP-2 can promote breast cancer cell growth by regulating cellular autophagy. This study was designed to check the change of autophagy level in breast cancer cells (MCF-7 and MX-1) by MDC (monodansyl- cadaverin) staining through RNA interfering ILP-2 expression after addition of rapamycin (Rapa) treatment, and the results showed that the number of autophagic vesicles decreased significantly after the interference with ILP-2. Western blot analysis showed that expression of ILP-2, ATG14 and Beclin1 was downregulated and the ILC3-II/LC3-I ratio decreased in MCF-7 and MX-1 cells, while expression of P62 thereafter increased knockdown of ILP-2. A series of experiments such as CCK-8(Cell Counting Kit-8)assays, scratch assays, AO-EB༈Acridine Orange(AO)/EB Double Stain Kit ༉double staining analysis and western blot showed that knockdown of ILP-2 resulted in down-regulation of autophagy, significant increase in caspase-9, 3 expression and downregulation of MMP-9 (matrix metalloproteinase − 9) expression, increased apoptosis rate and inhibition of cell migration ability. Co-immunoprecipitation verified the existence of an interaction between ILP-2 and PI3K. Further Western blot results showed that after interference with ILP-2 expression, cellular autophagy levels and PI3K and Akt protein, LC3-II/LC3-I and Bcl-2 and Beclin1 protein expressions were significantly decreased. However, P62 protein expression was significantly increased. The results suggest that the possible pathways of ILP-2 are based on PI3K/Akt, Bcl-2/Bax, caspase-mediated signaling pathway to activate autophagy to inhibit apoptosis and promote tumor cell growth.