Komlodi 2023 EUROMIT2023 Bologna

Komlódi T, Silva FSG, Duarte AI, Mena D, Garcia-Souza LF, Makrecka-Kuka M, Bento G, Grilo LF, Oliveira PJ, Gnaiger E (2023) Off-target effects of etomoxir: inhibition of mitochondrial Complex I and fatty acid oxidation. EUROMIT2023 International Conference on Mitochondrial Pathology.

Link: EUROMIT2023 International Conference on Mitochondrial Pathology

Komlodi Timea, Silva Filomena SG, Duarte Ana I, Mena Debora, Garcia-Souza Luiz F, Makrecka-Kuka Marina, Bento Guida, Grilo Luis F, Oliveira Paulo J, Gnaiger Erich (2023)

Event: EUROMIT2023 Bologna IT

Etomoxir is sold as an inhibitor of carnitine palmitoyltransferase-I (CPT-I) to block the transport of long-chain fatty acids into mitochondria. Etomoxir is used to inhibit fatty acid oxidation (FAO) in metabolic studies and drug development with tumor and non-tumor cells and in clinical trials attempting to switch energy metabolism from FAO to glucose oxidation. However, several studies suggest etomoxir may exert off-target effects, although the mechanism is not sufficiently known.

We determined the effects of etomoxir on mitochondrial electron transfer using high-resolution respirometry in living and permeabilized Huh7 human hepatocellular carcinoma cells and mitochondria isolated from mouse liver and brain. Palmitoylcarnitine (substrate of carnitine palmitoyl transferase-II CPT-II) or palmityl-CoA plus carnitine (CPT-I substrates) were used to test the effects of etomoxir on FAO in terms of oxidative phosphorylation and electron transfer capacity (F-pathway). NADH-linked substrates (N-pathway) and succinate (S-pathway) were applied as controls, and activities of respiratory Complexes CI, CII and CIV were measured to evaluate off-target effects of etomoxir.

Etomoxir (2.5 – 40 μM) did not selectively inhibit FAO with palmitoylcarnitine nor palmitoyl-CoA. In the presence of palmitoylcarnitine, 40 μM etomoxir showed a small inhibition of F- and N-pathways. This inhibition was pronounced at 200 μM etomoxir, when NS- and FNS-pathway capacities were inhibited. 40 μM and 200 μM etomoxir inhibited Complex I activity slightly and strongly, respectively.

Our results suggest that acute etomoxir treatment inhibits respiration through core pathways of mitochondrial energy metabolism particularly at high concentration. Both etomoxir and rotenone inhibited FAO by blocking Complex I. In conclusion, the off-target inhibition of mitochondrial respiration must be considered when using etomoxir in in-vitro studies and clinical trials.

• Keywords: Complex I, Etomoxir, High-resolution respirometry • Bioblast editor: Plangger M • O2k-Network Lab: HU Budapest Tretter L, AT Innsbruck Oroboros, AT Innsbruck Gnaiger E, AT Innsbruck MitoFit

Affiliations

Timea Komlódi^1,2, Filomena SG Silva³, Ana I Duarte^3,4,5, Débora Mena^3,5,6, Luiz F Garcia-Souza¹, Marina Makrecka-Kuka⁷, Guida Bento³, Luís F Grilo^3,5,6, Paulo J Oliveira³, Erich Gnaiger¹

Oroboros Instruments, Innsbruck, Austria;
Dept Biochem, Semmelweis Univ, Budapest, Hungary;
CNC-Center Neurosci and Cell Biol,
IIUC-Inst Interdisciplinary Research,
CIBB-Center for Innovative Biomed Biotechnol,
PDBEB-PhD Programme in Exp Biol Biomed, IIUC, Univ Coimbra, Portugal;
Lab Pharmaceut Pharmacol, Latvian Inst Organic Synthesis, Riga, Latvia

List of abbreviations, terms and definitions - MitoPedia

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Labels: MiParea: Respiration, Pharmacology;toxicology

Organism: Human, Mouse Tissue;cell: Nervous system, Liver Preparation: Permeabilized cells, Isolated mitochondria, Intact cells

Regulation: Fatty acid

Pathway: F, N, S, CIV HRR: Oxygraph-2k