Abstract 517: 2OHOA-mediated furin inhibition: Disrupting Notch via processing, transcription, and ADAM downregulation

Abstract

Abstract Purpose Glioblastomas (GBMs) are highly treatment-resistant and aggressive brain tumors. Herewe assessed the potential of melitherapy to treat GBM, a strategy based on regulating themembrane’s structure and organization to modify aspects of intracellular signaling. In thiscontext, we have evaluated the effects of 2-hydroxyoleic acid (2OHOA) on GBM cells, acompound currently under study in a phase IIB/III clinical trial for newly diagnosed GBMpatients. The current study aims to determine whether 2OHOA modulates the Notch pathway aspart of its antitumoral mechanism, as seen its relevance driving the pathogenesis of GBM. Methods The effect of 2OHOA was evaluated on different components of the pathway usingseveral methods: W-blot, Q-PCR, and confocal microscopy. The relevance of HES1 in itsmechanism of action was evaluated by its overexpression and pathway activation assays byJagged1. To analyze Notch receptor processing, we performed subcellular fractionation andcolocalization studies. Furin activity was assessed by fluorescence assays measuring its substratecleavage and its binding affinity to 2OHOA was determined by surface plasmon resonance. Results 2OHOA suppresses both the Notch3 and Notch2 signaling pathways in GBM cell linesusing a two-fold mechanism. Notch3 signaling is abolished by repressing its transcription.Instead, Notch2 pathway is hindered by blocking its first cleavage through the inactivation offurin activity by physical association. Consequently, the Notch2 receptor is retained in the Golgicomplex, preventing its trafficking to the plasma membrane to initiate the signaling cascade.Moreover, overexpression of its target gene, HES1, partially dampened the drug's antiproliferative effect showing the relevance of this pathway in the 2OHOA pharmacologicalefficacy. In addition, considering the levels of Notch intracellular domain (NICD) obtained inJagged1 assays in conjunction with 2OHOA, we evaluated the second cleavage of the Notchprocessing, being also downregulated, as ADAM10 constitutes a furin substrate. Conclusions These findings elucidate that the antitumoral activity of 2OHOA involves theinhibition of Notch signaling, potentially driven by the direct inhibition of furin. This reveals anovel molecular target for this bioactive lipid in the therapeutic approach to treating GBM. Citation Format: Raquel Rodríguez-Lorca, Roberto Beteta-Göbel, Ramón Roman, Victoria Lladó, Pablo V. Escribá, Paula Fernández-García. 2OHOA-mediated furin inhibition: Disrupting Notch via processing, transcription, and ADAM downregulation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 517.