Breast cancer of the luminal subtype remains a major cause of morbidity and mortality among women worldwide, largely due to the development of resistance to endocrine therapies such as tamoxifen. One of the transcription factors involved in the proliferation of breast cancer cells is E2F2, whose activity is regulated by the CDK4/6–Rb complex. This study aimed to evaluate the potential of active compounds in the 75% ethanol extract of Annona muricata (soursop) leaves as CDK6 inhibitors and suppressors of E2F2 expression in MCF-7 breast cancer cells. LC–MS/MS analysis identified 152 unique compounds, most of which belonged to the acetogenin class. Activity prediction using SkelSpheres descriptors based on support vector regression (SVR) yielded an R2-testing value of 0.8925, indicating high predictive validity for CDK6 inhibitory activity. Several compounds, including annonacin, muricin D, and longifolicin, exhibited predicted IC50 values below 1 µM. Molecular docking results showed strong affinity between acetogenins and CDK6, with rerank scores comparable to the reference ligand ribociclib. In vitro cytotoxicity testing demonstrated that the extract had an IC50 value of 9.7 ppm against MCF-7 cells. Gene expression analysis using RT-qPCR revealed a significant reduction in E2F2 expression following extract treatment (0.5175 ± 0.1732) compared with the control group (1.0087 ± 0.1517; p = 0.000), comparable to the effect of tamoxifen. These findings suggest that acetogenins present in the 75% ethanol extract of A. muricata leaves possess potential as anticancer candidates by suppressing E2F2 expression, presumably through the inhibition of CDK6. © 2026 Oriental Scientific Publishing Company. All rights reserved.