Lipid modulation and Oxphos uncoupling: a new therapeutic mode of action of anti-cancer agent 2-Hydroxyoleic acid (2OHOA) revealed in isolated mitochondria of U118MG glioblastoma cells
2Metabolic Disease Unit Hadassah Medical Center, Ein Kerem, The Hebrew University Jerusalem, Israel
3Mass Spectrometry Unit Institute for Drug Research, School of Pharmacy, The Hebrew University Jerusalem Israel, Israel
4Dorottya Utca, Lipodom Ltd, Hungary
5Department of Pharmacy, University of Salerno, Via Giovanni Paolo II, Italy
6Bionam Center for Biomaterials, University of Salerno, Via Giovanni Paolo II, Italy
7Department of Genetics, Hadassah Medical Center, Ein Kerem, The Hebrew University Jerusalem, Israel
8Laminar Pharmaceuticals, Laminar Pharmaceuticals,, Spain
9Molecular and Experimental Surgery, Clinic for General, Visceral, Vascular, and Transplant Surgery,, Medical Faculty, University Hospital Magdeburg,, Germany
10Department of Biology, University of the Balearic Islands, Spain
11Biological Research Centre, Institute of Biochemistry, Hungary
2-Hydroxyoleic acid (2OHOA), a synthetic fatty acid used as an anti-cancer agent based on its capacity to regulate membrane lipid composition and structure. 2OHOA incoporated into pre-existing lipids, generating hydroxylated lipids can activated sphingomyelin synthase 1 and elevate sphingomyelin levels. 2OHOA has also ability to depolarize the inner mitochondrial membrane. Previous study, we investigated a new anti-cancer modality of 2OHOA as an OxPhos uncoupler. It demonstrated beyond acting as a cancer cell selective OxPhos uncoupler, 2OHOA can also activated glycolysis to compensate the OxPhos deficiency of cancer cells. In this study, we mechanistically explored how 2OHOA can compromise mitochondrial function. Shotgun lipidomics, mass spectrometry, FACS and bioenergetic profiling were used to analyze mitochondria isolated from 2OHOA-treated glioblastoma cells. Lipidomics showed, 2OHOA treatment led to mitochondrial lipid hydroxylation. In molecular dynamics simulation, hydroxylation decreased CoQ diffusion in the liquid disordered membrane domain, which embeds respiratory complexes. Reduced CoQ diffusion is consistent with the inhibition of complex I–III activities when measured separately and not as part of the entire ETC. However, the stalled CoQ membrane diffusion cannot explain 2OHOA-mediated facilitation of state 3 respiration and complex IV electron transfer as measured within the ETC in intact mitochondria. 2OHOA inhibited β oxidation and glycolysis, suggesting that the enhanced/compensatory complex IV activity and state 3 respiration can sustain cancer cell viability in 2OHOA-treated cells and might, therefore, be a relevant therapeutic target for boosting 2OHOA cancer cell toxicity. This is the first mechanistic demonstration of modulation of an isolated organelle by membrane lipid therapy.