Dichloroacetate improves mitochondrial function, physiology, and morphology in FBXL4 disease models
Manuela Lavorato, … , Christoph Seiler, Marni J. Falk
Manuela Lavorato, … , Christoph Seiler, Marni J. Falk
Published July 26, 2022
Citation Information: JCI Insight. 2022;7(16):e156346. https://doi.org/10.1172/jci.insight.156346.
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Research Article Genetics Metabolism

Dichloroacetate improves mitochondrial function, physiology, and morphology in FBXL4 disease models

Abstract

Pathogenic variants in the human F-box and leucine-rich repeat protein 4 (FBXL4) gene result in an autosomal recessive, multisystemic, mitochondrial disorder involving variable mitochondrial depletion and respiratory chain complex deficiencies with lactic acidemia. As no FDA-approved effective therapies for this disease exist, we sought to characterize translational C. elegans and zebrafish animal models, as well as human fibroblasts, to study FBXL4–/– disease mechanisms and identify preclinical therapeutic leads. Developmental delay, impaired fecundity and neurologic and/or muscular activity, mitochondrial dysfunction, and altered lactate metabolism were identified in fbxl-1(ok3741) C. elegans. Detailed studies of a PDHc activator, dichloroacetate (DCA), in fbxl-1(ok3741) C. elegans demonstrated its beneficial effects on fecundity, neuromotor activity, and mitochondrial function. Validation studies were performed in fbxl4sa12470 zebrafish larvae and in FBXL4–/– human fibroblasts; they showed DCA efficacy in preventing brain death, impairment of neurologic and/or muscular function, mitochondrial biochemical dysfunction, and stress-induced morphologic and ultrastructural mitochondrial defects. These data demonstrate that fbxl-1(ok3741) C. elegans and fbxl4sa12470 zebrafish provide robust translational models to study mechanisms and identify preclinical therapeutic candidates for FBXL4–/– disease. Furthermore, DCA is a lead therapeutic candidate with therapeutic benefit on diverse aspects of survival, neurologic and/or muscular function, and mitochondrial physiology that warrants rigorous clinical trial study in humans with FBXL4–/– disease.

Authors

Manuela Lavorato, Eiko Nakamaru-Ogiso, Neal D. Mathew, Elizabeth Herman, Nina Shah, Suraiya Haroon, Rui Xiao, Christoph Seiler, Marni J. Falk

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Figure 7

FBXL4–/– human fibroblasts had reduced CS activity and abnormal mitochondrial structure that was improved with DCA treatment.

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FBXL4–/– human fibroblasts had reduced CS activity and abnormal mitocho...
(A) CS activity decreased in patient fibroblasts from participants 1 (n = 3) and 2 (n = 5) as compared with control fibroblasts (n = 3). (B–D) RC enzyme activity was not significantly changed in patient fibroblasts compared with control fibroblasts. (E) Extracellular lactate levels were significantly decreased in patient fibroblasts compared with control cells in control media. DCA treatment did not affect extracellular lactate levels in patient cells. n = 12 each condition. (F) Intracellular lactate level evaluation in untreated and DCA-treated (20 mM) fibroblasts from participant 1 and controls. n = 12 each condition. (G) CS activity after treatment of fibroblasts with 20 mM DCA (participants 1 and 2). CS activity level in treated fibroblasts was normalized for percentage in control media. Data are shown as the mean ± SEM. Significance was determined using unpaired Student’s t test. *P < 0.05, ***P < 0.001. Bonferroni’s correction method was applied for A and E–G to account for multiple comparisons, and significant findings still held. (H and I) Low-magnification images (original magnification, ×20) of FBXL4–/– fibroblasts from participant 1 after exposure to metabolic stressor and cotreatment with 20 mM DCA for 48 hours. Mitochondria were stained with MitoTracker green (Thermofisher Scientific). Scale bar: 200 μm. (J and K) High-magnification images of mitochondrial morphology in FBXL4–/– fibroblasts from participant 1 after exposure to metabolic stressor and cotreatment with 20 mM DCA for 48 hours (see representative fibroblast image from participant 2 in Supplemental Figure 5). Mitochondria (in green) were disarranged and fragmented when incubated with stressor. More elongated mitochondria were visible after coincubation with 20 mM DCA. Scale bar: 25 μm. (L and M) EM images of mitochondria in FBXL4–/– fibroblasts from participant 1, showing loss of matrix electron density and loss of mitochondrial cristae after incubation in metabolic stressor for 48 hours and rescue of mitochondrial ultrastructure, with defined cristae and normal matrix electron density after cotreatment with 20 mM DCA. Scale bar: 500 nm.