Abstract
BackgroundAspartate β-hydroxylase (ASPH) is silent in normal adult tissues only to re-emerge during oncogenesis where its function is required for generation and maintenance of malignant phenotypes. Exosomes enable prooncogenic secretome delivering and trafficking for long-distance cell-to-cell communication. This study aims to explore molecular mechanisms underlying how ASPH network regulates designated exosomes to program development and progression of breast cancer.MethodsStable cell lines overexpressing or knocking-out of ASPH were established using lentivirus transfection or CRISPR-CAS9 systems. Western blot, MTT, immunofluorescence, luciferase reporter, co-immunoprecipitation, 2D/3-D invasion, tube formation, mammosphere formation, immunohistochemistry and newly developed in vitro metastasis were applied.ResultsThrough physical interactions with Notch receptors, ligands (JAGs) and regulators (ADAM10/17), ASPH activates Notch cascade to provide raw materials (especially MMPs/ADAMs) for synthesis/release of pro-metastatic exosomes. Exosomes orchestrate EMT, 2-D/3-D invasion, stemness, angiogenesis, and premetastatic niche formation. Small molecule inhibitors (SMIs) of ASPH’s β-hydroxylase specifically/efficiently abrogated in vitro metastasis, which mimics basement membrane invasion at primary site, intravasation/extravasation (transendothelial migration), and colonization/outgrowth at distant sites. Multiple organ-metastases in orthotopic and tail vein injection murine models were substantially blocked by a specific SMI. ASPH is silenced in normal adult breast, upregulated from in situ malignancies to highly expressed in invasive/advanced ductal carcinoma. Moderate-high expression of ASPH confers more aggressive molecular subtypes (TNBC or Her2 amplified), early recurrence/progression and devastating outcome (reduced overall/disease-free survival) of breast cancer. Expression profiling of Notch signaling components positively correlates with ASPH expression in breast cancer patients, confirming that ASPH-Notch axis acts functionally in breast tumorigenesis.ConclusionsASPH-Notch axis guides particularly selective exosomes to potentiate multifaceted metastasis. ASPH’s pro-oncogenic/pro-metastatic properties are essential for breast cancer development/progression, revealing a potential target for therapy.
Highlights
Aspartate β-hydroxylase (ASPH) is silent in normal adult tissues only to re-emerge during oncogenesis where its function is required for generation and maintenance of malignant phenotypes
ASPH’s prooncogenic/pro-metastatic properties are essential for breast cancer development/progression, revealing a potential target for therapy
ASPH physically interacts with notch receptors, ligands, and regulators to activate notch cascade in breast cancer Expression profile of ASPH-Notch axis was demonstrated in a panel of human breast cancer cell lines (Additional file 1: Figure S1A-B)
Summary
Aspartate β-hydroxylase (ASPH) is silent in normal adult tissues only to re-emerge during oncogenesis where its function is required for generation and maintenance of malignant phenotypes. This study aims to explore molecular mechanisms underlying how ASPH network regulates designated exosomes to program development and progression of breast cancer. ASPH is silent in normal adult tissues, during tumorigenesis, it is re-emerged and markedly upregulated by growth factor signaling pathways IN/IGF1/IRS1/RAS/ RAF/MAPK/ERK, IN/IGF1/IRS1/PI3K/AKT, and WNT/βcatenin [8, 10,11,12,13] attributed to development and progression of malignancies [14,15,16]. It is yet to be illustrated whether Notch cascade activated by ASPH is a major molecular mechanism to generate and maintain malignant cellular behaviors in breast cancer [8]
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