Breast cancer is the most widely diagnosed cancer and the leading cause of cancer-related death in female. Triple-negative breast cancer (TNBC), defined by the lack of hormone receptors (ER and PR) and HER2, is the most aggressive and highly lethal subtype. The typical hormone- and HER2-targeted therapies used for most other breast cancer patients have limited effectiveness in TNBC patients due to the lack of hormone receptors and HER2 amplification. Currently, chemotherapy is the primary treatment option, but it often yields unsatisfactory results. Another major challenge for TNBC is the frequent occurrence of brain metastasis (BM), which considerably reduces the overall survival rate of patients. The treatment for BM is even more difficult because of the existence of the blood-brain barrier. Therefore, there is an unmet need for the development of novel targeted therapies for treating both TNBC and its brain metastases to improve patient outcomes.

Microtubule proteins, including α- and β-tubulins, are often recognized as the target when designing novel tubulin-binding chemotherapeutics. Alterations in the expression of different β-tubulin isoforms have been reported for different cancers, including breast cancer, and can impact tumor development, progression, as well as response to chemotherapy. However, the detailed function and mechanism of β-tubulin isoforms in breast cancer, particularly in TNBC, and brain metastasis is unclear. In this study, we found that TUBB2B, an isotype of β-tubulins, is significantly associated with TNBC as well as TNBC-BM. Its high expression correlates with poor overall survival in TNBC patients. Depletion of TUBB2B inhibits the growth of TNBC spheroid and promotes apoptosis in vitro. In vivo, TUBB2B silencing decreases the growth of TNBC primary tumor and its colonization in the brain. Mechanistically, we elucidate a new role for TUBB2B in promoting protein synthesis by interacting with and stabilizing eukaryotic translation elongation factor 1A1 (eEF1A1). Moreover, in the brain tumor microenvironment, TUBB2B activates astrocytes, which in turn increases the expression of TUBB2B in TNBC cells. This positive feedback loop between TUBB2B-enriched TNBC cells and astrocytes further promotes TNBC cells to proliferate in the brain. We also developed TUBB2B siRNA-gold nanoparticles and demonstrated suppression of TNBC growth in xenograft models. Thus, we demonstrated TUBB2B as an oncogenic gene promoting TNBC primary tumor growth and brain metastasis colonization. Our findings identify TUBB2B as a novel molecular target for TNBC treatment.

Super-enhancers (SEs) are defined by large clusters of adjacent active enhancers, abundant with transcription factors (TFs), coactivators, and mediators, that promote gene expression. Besides regulating crucial genes related to cell identity, SEs have been demonstrated to drive oncogene expression in various cancer types. In this study, we investigated TFs that act on the TNBC-specific SE, thus modulating TNBC cell growth and stemness. The AP1 transcription activator component, FOSL1, was identified as a major TF that binds to the SE region of TCOF1 gene, a novel oncogene previously identified by our lab. Depletion of FOSL1 in TNBC cells results in a significant reduction of TCOF1 mRNA and protein levels. Furthermore, we demonstrated that overexpression of FOSL1 activates the TCOF1 SE, thereby increasing TCOF1 expression. Lastly, TNBC spheroid growth and stemness are inhibited upon FOSL1 depletion. Taken together, this work identifies TCOF1 as a direct transcriptional target of FOSL1 and highlights the potential of targeting FOSL1 for TNBC treatment.