The family is a highly conserved gene group that regulates cell-cell interaction, embryogenesis, and tissue commitment. This review summarizes the existing data on the third subtype of the family, Notch3. The role of Notch3 in different types of cancers is discussed, as well as implications of its modification and new strategies to affect Notch3 signaling activity.

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Purpose Notch1, a trans-membrane receptor, has been recently shown to aid in the determination of thyroid cell fate associated with tumorigenesis. The present study aimed to investigate the clinical relevance of Notch1 and its role in the regulation of differentiated thyroid cancer (DTC) behavior. Experimental design We examined Notch1 expression level and its relationship with clinicopathologic features and outcomes of DTC. Notch1 intracellular domain (NICD) was further characterized both in vitro and in vivo by gain-of-function assays using an inducible system. Results Notch1 expression levels were down-regulated in primary DTC tissue samples compared with contralateral non-tumor and benign thyroid tissues. Decreased Notch1 expression in DTC was associated with advanced patient age (p=0.032) and the presence of extrathyroidal invasion (p=0.005). Patients with lower Notch1 expression had a significantly higher recurrence rate (p=0.038). Restoration of NICD in a stably doxycycline-inducible metastatic DTC cell line reduced cell growth and migration profoundly. Using an orthotopic thyroid cancer model, NICD induction significantly reduced the growth of the primary thyroid tumor and inhibited the development of lung metastasis. SERPINE1 was discovered by microarray as the most significant gene down-regulated by NICD. Further validation showed that induction of NICD reduced SERPINE1 expression in a dose-dependent manner while restoration of a relative higher level of SERPINE1was observed with NICD back to minimal level. Additionally, SERPINE1 knock-down inhibited DTC cell migration. Conclusions Notch1 regulates the aggressive phenotypes of DTC, which could be mediated by SERPINE1 inhibition. Notch1/SERPINE1 axis warrants further investigation as a novel therapeutic target for advanced DTC.

BACKGROUND Thyroid tumorigenesis is characterized by a progressive loss of differentiation exhibited by a range of disease variants. The Notch(1-4) receptor family regulates developmental progression in both normal and cancerous tissues. We sought to characterize the third Notch isoform (Notch3) across the various differentiated states of thyroid cancer and determine its clinical impact. METHODS Notch3 expression was analyzed in a tissue microarray of normal and pathologic thyroid biopsies from 155 patients. The functional role of Notch3 was then investigated by upregulating its expression in a follicular thyroid cancer (FTC) cell line. RESULTS Notch3 expression regressed across decreasingly differentiated, increasingly malignant thyroid specimens, correlated with clinicopathological attributes reflecting poor prognosis, and independently predicted survival following univariate and multivariate analysis. Overexpression of the active Notch3 intracellular domain (NICD3) in an FTC, gain-of-function line led to functional CBF1-binding and increased thyroid-specific gene transcription. NICD3 induction also reduced tumor burden in vivo, and initiated the intrinsic apoptotic cascade, alongside suppressing cyclin and Bcl-2 family expression. CONCLUSIONS Loss of Notch3 expression may be fundamental to the process of dedifferentiation that accompanies thyroid oncogenesis. Conversely, activation of Notch3 in thyroid cancer exerts an antiproliferative effect and restores elements of a differentiated phenotype. These findings provide preclinical rationale for evaluating Notch3 as a disease prognosticator and therapeutic target in advanced thyroid cancer.

Novel therapies for neuroendocrine (NE) cancers are desperately needed as they frequently present as metastatic disease and cause debilitating symptoms by secreting excessive hormones. Induction of Notch isoforms has a tumor suppressive effect in NE cancer cell lines, and we have observed that histone deacetylase inhibitors (HDACi) potently activate Notch. In this study, we describe the potential for Burkholderia thailandensis-derived class I HDACi thailandepsin A (TDP-A) as a Notch activator and therapeutic agent against NE cancer. IC50 for TDP-A was determined to be 4-6 nM in NE cancer cell lines (BON, MZ-CRC-1, and TT) without cytotoxicity to lung fibroblasts. The binding characteristics of TDP-A to its target HDAC1 was examined using bioluminescence resonance energy transfer (BRET). Western blot and flow cytometry analysis showed that TDP-A induces cell cycle arrest and apoptosis in a dose-dependent manner. TDP-A dose-dependently activated the Notch pathway as measured by increasing functional CBF1-luciferase reporter signal and mRNA and protein expressions of Notch isoforms, which were attenuated by pretreatment with γ-secretase inhibitor DAPT. Furthermore, TDP-A lead to changes in expression level of downstream targets of Notch pathway and reduced expression of NE cancer markers. An in vivo study demonstrated that TDP-A suppressed NE cancer progression. These results show that TDP-A, as a Notch activator, is a promising agent against NE cancers.