Abstract P22: Progestin (R5020) Induced Cell Apoptosis Via Mitochondria-Mediated Cell Death Pathway In MCF-7 Cell With PRB Overexpression

Progestin has been widely reported to exert growth stimulatory or inhibitory effects on breast cancer cells depending on the cellular context. Our previous research showed that Promegestone (R5020) treatment induced apoptosis in MCF-7 cells with overexpression of PRB (MCF-7PRB), which paradoxically associated with massive downregulation of the known pro-apoptotic protein such as PARP, BID, Caspase 7 and 8. The objective of this study was to find out the mechanism of R5020-induced apoptosis in MCF-7PRB cells. Proteomic profiling of total cell lysates of control and R5020 treated MCF-7PRB cells were conducted using Tandem Mass Tag (TMT). HYPOXIA was found to be the topmost enriched Hallmark in R5020 treated cells. Western blotting analysis confirmed that HIF1A, BNIP3, and BNIP3L/NIX were massively upregulated in R5020 treated cells. BNIP3 and BNIP3L/NIX, the downstream apoptosis mediator of HIF1A can cause mitochondria dysfunction by forming pores on the mitochondria outer membrane or interacting with BCL-2 or BCL-xL. Indeed, mitotracker staining showed fragmented mitochondria, and this is associated with increased cytochrome C leakage into the cytoplasm and increased level of pro- and cleaved caspase 9 in R5020 treated cells. Furthermore, R5020 induced marked increases in many mitochondrial proteins including Apoptosis-Inducing Factor (AIF/AIFM1), AIFM2, Endonuclease G (EndoG), High Temperature Requirement A2 (HTRA2/ OMI), Second Mitochondria-Derived Activator of Caspases (Smac/DIABLO) and PARL (Presenilin-associated rhomboid-like), endonuclease G and Htra2/Omi, all of which are proapoptotic. For example, AIF/AIFM1and AIFM2 have been shown to induce caspase independent apoptosis by recruiting DNA nuclease such as ENGOG to the nuclease and causing chromatin condensation and DNA fragmentation. In conclusion, progestin induces mitochondria mediated apoptosis in breast cancer cells by activation of hypoxia pathway, upregulation of many proapoptotic mitochondrial proteins and downregulation of anti-apoptotic proteins BCL-2 and BCL-XL. Citation Format: Qian Yee Woo, Pheck Khee Lau, Meng Wei, Shi Hao Lee, Kye Siong Leong, Natasa Bajalovic, Valerie C.L Lin. Progestin (R5020) Induced Cell Apoptosis Via Mitochondria-Mediated Cell Death Pathway In MCF-7 Cell With PRB Overexpression [abstract]. In: Proceedings of Frontiers in Cancer Science 2024; 2024 Nov 13-15; Singapore. Philadelphia (PA): AACR; Cancer Res 2025;85(15_Suppl):Abstract nr P22.