Mode of action of the mitochondrial Ca2+ uptake enhancer efsevin for treatment of cardiac arrhythmia

Wilting, Fabiola (2023): Mode of action of the mitochondrial Ca2+ uptake enhancer efsevin for treatment of cardiac arrhythmia. Dissertation, LMU München: Medizinische Fakultät

Abstract

Cardiovascular diseases (CVDs) like arrhythmia cause immense health and economic burdens in Europe and worldwide and are the number one cause of death worldwide. Mitochondria have been shown to act as regulators of cardiac rhythm, but their potential as therapeutic targets in the treatment of CVDs has not been further investigated. Recently, the synthetic compound efsevin was identified as a potent candidate drug for the treatment of cardiac arrhythmia due to its suppressive effect on arrhythmia in a zebrafish model. Its antiarrhythmic activity was linked to an interaction with the voltage-dependent anion channel 2 (VDAC2), a pore-forming channel in the outer mitochondrial membrane (OMM). VDAC2 facilitates the transport of a variety of metabolites and ions, including Ca2+, through the OMM. However, its potential to directly modulate calcium transport into the mitochondria was questionable due to its large pore size. In this work, the mode-of-action of efsevin was characterized. I could proof in a cardiomyocyte cell line, that efsevin leads to increased fast Ca2+ uptake into the mitochondria. This increased Ca2+ calcium uptake was linked to an anti-arrhythmic effect and confirmed in freshly isolated RyR2R4496C/WT cardiomyocytes. In addition, the efsevin-VDAC2 interaction was biophysically described. It was shown that efsevin modulates the open probability of VDAC2 and stabilizes the channel in its cation-selective closed state. Molecular ligand-protein docking was used to identify the binding site of efsevin on the inner wall of the channel, adjacent to the N-terminal α-helix. The binding site is formed by amino acids N207, K237, and N238. Replacement of these three amino acids with alanines abolished the efsevin sensitivity of the channel. This biophysical effect was also demonstrated in cultured murine cardiomyocytes in which either wild-type zVDAC2 or the efsevin-insensitive zVDAC2AAA mutant were overexpressed. Both channels, wild-type zVDAC2 and the zVDAC2AAA mutant, restored mitochondrial Ca2+ uptake. However, only wild-type zVDAC2 was sensitive to efsevin leading to an increased mitochondrial Ca2+ uptake. In conclusion, in this work, I established pharmacological modulation of fast mitochondrial Ca2+ uptake as a possible pharmacological target and was able to show the capability of zVDAC2 to modulate Ca2+ levels. Deciphering the direct interaction of efsevin and zVDAC2 reveals new insights into cardiomyocyte functioning and provides the basis for the computer-aided development of optimized molecules that could function as tools in research and ultimately as therapeutics for CVDs.

• Dokumententyp: Dissertationen (Dissertation, LMU München)
• Keywords: VDAC, VDAC2, arrhythmia, efsevin, CVD, calcium
• Themengebiete: 600 Technik, Medizin, angewandte Wissenschaften ; 600 Technik, Medizin, angewandte Wissenschaften > 610 Medizin und Gesundheit
• Fakultäten: Medizinische Fakultät
• Sprache der Hochschulschrift: Englisch
• Datum der mündlichen Prüfung: 3. Februar 2023
• 1. Bericht­erstatter:in: Nicke, Annette