Mitochondrial diseases, characterized by dysfunction in the cellular powerhouse, the mitochondria, present a complex and heterogeneous group of disorders. As research in mitochondrial medicine advances, the need for effective therapies becomes increasingly apparent. Collaborative efforts among researchers, clinicians, regulatory bodies, patient advocacy groups and other stakeholders are crucial to overcome the challenges linked to the complexity of mitochondrial medicine, and to ensure the successful implementation of clinical trials in this field. This lecture explores the key aspects of trial readiness in the context of mitochondrial medicine, emphasizing the challenges and opportunities in designing and executing successful clinical trials. An overview of the ongoing clinical trials will be also provided.

The impairment of mitochondrial respiration, observed in neurodegenerative and cardiovascular disease, diabetes, cancer, and migraine headaches, has emerged as a biomarker of mitochondrial dysfunctions. Chronic fatigue, depression, and other behavior/mood disorders are also associated with mitochondrial malfunctioning, but so is our lifestyle! Our lab offers tests for insight into mitochondrial fitness, linking not only diseases but also behaviors and modern lifestyles that lead to health damage. Firstly, we focused on 88 (relatively) healthy volunteers, of which 32% were taking some medication (such as for high blood pressure or mood disorders), however, they considered themselves fit and healthy.  The blood was drawn 3h before PBMC (peripheral blood mononuclear cells) isolation, followed by an immediate Seahorse XF Cell Mito Stress Test (Agilent) on the SeahorseXF96e instrument (Agilent). Parameters of mitochondrial respiration were carefully examined. There was a significant difference between BHI (bioenergetic health index), reserve capacity, coupling efficiency, and proton leak, between people who took medication for chronic but manageable comorbidities and completely healthy individuals. Later, in another group we examined the alterations in NAD+ levels (by Q-NADMED Blood NAD+ assay kit, NADMED) and mitochondrial respiration parameters in a binge-drinking session (consuming 10 or more units of alcohol in less than three days). The decrease in NAD+ levels was positively correlated with the amount of alcohol consumed. Additionally, total NAD+ levels positively correlated with the BHI.  In another experiment, supplementation with niacin for 20 days, did not increase NAD+ levels in (relatively) healthy individuals. Apart from mitochondrial respiration and NAD+ levels, we focus on optimizing tests for mtDNA count and mitochondrial potential. All of these tests not only explore disease but also serve to monitor behaviors that lead to health damage or improvements.

Succinate dehydrogenase (SDH) connects the tricarboxylic acid (TCA) cycle and the respiratory chain. Mutations in SDH subunits have been associated with tumorigenesis and mitochondrial disease. In this project, we focused on subunit A of SDH (SDHA), primarily associated with inherited mitochondrial disease, and investigated the consequences of its loss or re-expression of mutant variants in HEK cells (SDHA KO). Lack of SDHA led to a downregulation of all SDH subunits and a secondary downregulation of the majority of mitochondrial complex I and IV subunits. Cellular respiratory capacity was severely decreased in the model, SDH-dependent respiration completely abolished and complex I-dependent respiration attenuated, reflecting the downregulation of respiratory chain complexes in general. Finally, the NAD⁺/NADH ratio was increased in SDHA KO, indicating complex rearrangement of the TCA. It resulted in higher glycolytic activity and lipid accumulation.

Supported by Czech Science Foundation (21-18993S), Grant Agency of Charles University (283423) and Czech Health Research Council (NU22-01-00499).