Leucine-rich repeat neuronal protein 1 as a novel biomarker of breast cancer stem cells and poor prognosis in patients with breast cancer

Cancer stem cells (CSCs) play critical roles in tumor progression, therapeutic resistance, and disease relapse in breast cancer. Leucine-rich repeat neuronal protein-1 (LRRN1), previously identified as a regulator of pluripotency in human embryonic stem cells, has been implicated in oncogenic processes across multiple solid tumors. However, the functional role of LRRN1 in breast cancer stem cells (BCSCs) and its clinical significance in breast cancer remain incompletely understood. LRRN1 expression was evaluated in breast cancer patient cohorts and patient-derived xenograft (PDX) models. BCSC-associated phenotypes were assessed by mammosphere formation, aldehyde dehydrogenase (ALDH) activity, migration assay, orthotopic xenograft tumor growth, and extreme limiting dilution analysis (ELDA) to determine tumor-initiating cell (TIC) frequency following genetic manipulation of LRRN1 expression. AKT isoform expression was analyzed by quantitative RT-PCR and Western blotting. Transcriptomic alterations associated with LRRN1 silencing were examined using RNA sequencing followed by pathway-level analyses. Circulating anti-LRRN1 autoantibody levels were measured using a Luminex-based immunoassay and correlated with clinical outcomes. LRRN1 expression was significantly enriched in BCSCs compared with non-BCSC populations in PDX tumors. Genetic silencing of LRRN1 attenuated mammosphere formation, ALDH activity, migratory capacity, in vivo tumor growth, and tumor-initiating cell (TIC) frequency, whereas LRRN1 overexpression enhanced BCSC-associated properties across distinct breast cancer subtypes. At the molecular level, LRRN1 expression was associated with altered AKT isoform expression-most prominently AKT3-without detectable changes in AKT phosphorylation. RNA-seq analysis further revealed coordinated alterations in PI3K-AKT-related and cytoskeleton-associated signaling pathways following LRRN1 depletion. Clinically, elevated tumor LRRN1 mRNA expression and increased circulating anti-LRRN1 autoantibody levels were associated with poorer recurrence-free and overall survival, particularly in early-stage luminal breast cancer. Circulating anti-LRRN1 autoantibody titers positively correlated with tumor LRRN1 expression. LRRN1 is enriched in breast cancer stem cell populations and functionally contributes to CSC-related phenotypes, tumor-initiating capacity, and tumor growth. Its expression is linked to isoform-specific regulation of AKT and transcriptomic changes in AKT-related signaling pathways. Both tumor LRRN1 expression and the corresponding circulating autoantibody response may serve as prognostic biomarkers, highlighting the translational potential of LRRN1 for breast cancer risk stratification and therapeutic development.