MAML1 drives Notch and Hedgehog oncogenic pathways by inhibiting Itch activity in triple-negative breast cancer

Triple-negative breast cancer (TNBC) is an aggressive and heterogeneous breast cancer subtype with poor patient outcomes. TNBC heterogeneity arises from multiple dysregulated pathways, including Notch and Hedgehog, which contribute to tumor initiation, progression, and drug resistance. Identifying common molecular regulators of TNBC aggressiveness is crucial for developing effective therapeutic strategies. Here, we demonstrate that the transcriptional coactivator MAML1 drives TNBC aggressiveness by regulating Notch1 and Gli1 stability through the E3 ubiquitin ligase Itch, functioning as an Itch-negative regulator. Mechanistically, MAML1 interacts with Itch via its PPQY motif and promotes K63-linked self-ubiquitylation of Itch, deregulating its expression/activity. Using a Maml1-deficient mouse model, we reveal an inverse correlation between MAML1 and Itch levels, where the loss of MAML1 stabilizes Itch and suppresses Notch1 and Gli1 activity. Conversely, MAML1 upregulation enhances Notch1 and Gli1 expression, driving accelerated TNBC tumor growth and faster multiorgan metastasis in vivo. Accordingly, we show that MAML1 is overexpressed in a cohort of TNBC patients, and the combined overexpression of MAML1/Notch1 and MAML1/Gli1 correlates with poor clinical outcomes by in silico analysis. Our findings establish a dual role for MAML1 as a transcriptional coactivator and a post-translational regulator of Itch, thereby amplifying Notch and Hedgehog oncogenic signaling. This study uncovers MAML1 as a key driver of TNBC progression and a potential therapeutic target for fighting TNBC aggressiveness and heterogeneity.