Inhibition of ATG5-mediated autophagy maintains PMAIP1 stability to promote cell apoptosis and suppress triple-negative breast cancer progression.

OBJECTIVE: Triple-negative breast cancer (TNBC) is a highly aggressive subtype of breast cancer characterized by a high recurrence rate and a lack of effective targeted therapies. The purpose of this study was to investigate the interaction between the pro-apoptotic factor phorbol-12-myristate-13-acetate-induced protein 1 (PMAIP1) and autophagy-related protein 5 (ATG5), as well as their regulatory mechanisms in TNBC cell apoptosis and autophagy, to identify potential therapeutic targets for TNBC. METHODS: TNBC-related datasets were retrieved from The Cancer Genome Atlas and selected by Prediction Analysis of Microarray 50 analysis to assess the expression of PMAIP1 in the samples. Additionally, the expression of PMAIP1 in the TNBC cell lines (MDA-MB-231) was detected using quantitative real-time polymerase chain reaction. In MDA-MB-231 cells, the expression of PMAIP1 and ATG5 was overexpressed or knocked down, and autophagy was inhibited using chloroquine (20 μM). Gene and protein expression levels were evaluated using quantitative real-time polymerase chain reaction and Western blot, respectively. Immunofluorescence was used to observe microtubule-associated protein 1 light chain 3 puncta formation to assess autophagy levels. Furthermore, cell apoptosis, proliferation, migration, and invasion were analyzed using terminal deoxynucleotidyl transferase dUTP nick-end labeling assay, colony formation assay, and Transwell assay. RESULTS: Compared to the control group, the expression of PMAIP1 was significantly elevated in TNBC tissues and MDA-MB-231 cells. Furthermore, overexpression of PMAIP1 led to a marked increase in apoptosis levels and a remarkable reduction in autophagy levels in MDA-MB-231 cells, while knockdown of PMAIP1 showed the opposite effects. Additionally, knockdown of ATG5 expression or treatment with chloroquine not only resulted in an increase in PMAIP1 expression in a time-dependent manner, but also reduced autophagy levels and enhanced apoptosis levels of cells. Furthermore, simultaneous knockdown of PMAIP1 and ATG5 considerably up-regulated apoptosis levels while down-regulating autophagy levels. Moreover, knockdown of PMAIP1 alone promoted the viability, invasion, and migration abilities of TNBC cells, while dual knockdown reversed these effects. CONCLUSION: Inhibition of ATG5-mediated autophagy maintains PMAIP1 stability, thereby promoting cell apoptosis and suppressing TNBC progression.