Background: Survivin, an inhibitor of apoptosis proteins (IAPs), is more strongly expressed in triple negative-breast cancer (TNBC) than other subtypes of breast cancer and closely associated with aggressiveness characterized by rapid progression and poor prognosis. The main function of survivin is to regulate cell division and prevent apoptosis. However, other survivin mechanisms are complex and not fully understood. The aim of this study was to evaluate the effects of survivin knockout on TNBC progressiveness relating to proliferation, apoptosis, stemness, cellular stress response, and metastasis mechanisms. Methods and results: The CRISPR/Cas9 (clustered regularly interspaced short palindrom repeat-associated Cas9) system was utilized to establish survivin knockout in the BT549 TNBC cell line. Both knockout and wild-type cells were used to study the role of survivin in various biological mechanisms. Apoptosis-, pluripotency-, and cellular stress-related proteins were examined via proteomic arrays. The cell cycle, apoptosis, and expression of breast cancer stem cell markers were analyzed via flow cytometry. Metastasis-related markers were evaluated via qRT‒PCR. Here, we report that survivin knockout inhibits proliferation and induces apoptosis in TNBC cells. Moreover, survivin knockout suppressed the expression of pluripotent markers and promoted a shift toward activation response to cellular stress, such as genotoxic, hypoxic, and oxidative stress. Additionally, survivin knockout suppressed metastasis. Conclusion: The loss of survivin leads to attenuation of TNBC progression by altering various mechanisms, particularly by suppressing stemness and altering the cellular stress response. We propose that knocking out survivin could be a potential strategy for breast cancer therapy, especially TNBC. © The Author(s), under exclusive licence to Springer Nature B.V. 2025.