Abstract

Hyperactivation of the PI3K/AKT pathway is a hallmark of metastatic triple-negative breast cancer (TNBC), but its drivers in TNBC retaining wild-type PTEN are poorly understood. Here, we identify Testis-Specific Y-Like Protein 5 (TSPYL5) -the top metastasis-associated gene from an unbiased bioinformatics screen- as a master regulator that resolves this paradox. Clinically, TSPYL5 amplification and overexpression are robust predictors of metastatic progression and poor prognosis. High-resolution single-cell and spatial analyses reveal that TSPYL5 defines a malignant subpopulation with stem-like, genomically unstable, and pro-metastatic properties. Functionally, TSPYL5 is sufficient to drive spontaneous polymetastasis from orthotopic tumors and is indispensable for post-intravasation colonization, culminating in overt liver metastases in 60% of animals-a phenotype absent in controls. Mechanistically, TSPYL5 sequesters the deubiquitinase USP10, thereby preventing it from stabilizing the tumor suppressor PTEN. This TSPYL5-USP10 interaction triggers the proteasomal degradation of PTEN, circumventing its wild-type status to hyperactivate PI3K/AKT signaling and unleash a ZEB1-driven metastatic program. This study delineates a complete TSPYL5-USP10-PTEN axis, providing a new paradigm for the post-translational tumor suppressor inactivation in TNBC. Our work validates TSPYL5 as a biomarker for PI3K pathway dependency and establishes the TSPYL5-USP10 interface as a tractable therapeutic target to restore PTEN function and combat metastatic TNBC.