Combined oxidative phosphorylation deficiency Cell Models for Research

Combined oxidative phosphorylation deficiency (COXPD) is a group of rare mitochondrial disorders characterized by defects in multiple respiratory chain complexes. The exact prevalence is unknown, but mitochondrial diseases collectively affect approximately 1 in 5,000 individuals. COXPD presents with severe early-onset symptoms, including encephalopathy, cardiomyopathy, hypotonia, and lactic acidosis. Prognosis is poor, with many patients succumbing in infancy or early childhood. The clinical heterogeneity and lack of effective therapies underscore the urgent need for research models.

COXPD is ideal for studying mitochondrial function, nuclear-mitochondrial communication, and bioenergetics. The disease involves mutations in nuclear genes encoding subunits or assembly factors of respiratory chain complexes, offering a clear genotype-phenotype correlation. Public datasets, such as those from the Mitochondrial Disease Sequence Data Resource (MSeqDR) and ClinVar, provide genetic variants. Open questions include the molecular mechanisms of tissue-specific manifestations and potential therapeutic targets. Gene-edited cell models enable precise dissection of these pathways.

• • Although COXPD is not a cancer, the underlying pathways are relevant to mitochondrial dysfunction in cancer. Key pathways include:
• • Oxidative phosphorylation (OXPHOS): Defects in complex I, II, III, IV, or V lead to reduced ATP production and increased reactive oxygen species (ROS).
• • Mitochondrial dynamics: Imbalance in fusion/fission affects mitochondrial morphology and function.
• • Apoptosis: Mitochondrial dysfunction can trigger intrinsic apoptosis via cytochrome c release.
• • Metabolic reprogramming: Cells may shift to glycolysis (Warburg effect) to compensate for OXPHOS deficiency.

Data from ClinVar and MSeqDR.

• • Mitochondrial dysfunction in COXPD affects several signaling networks:
• • AMPK pathway: Activated by increased AMP/ATP ratio, promoting catabolism.
• • mTORC1 signaling: Suppressed due to energy stress, affecting cell growth.
• • Integrated stress response (ISR): Activation of eIF2α kinases (e.g., GCN2, PERK) in response to mitochondrial stress.
• • ROS-mediated signaling: Increased ROS can activate NF-κB and HIF-1α, influencing inflammation and adaptation.

Organoids, such as cerebral organoids, can recapitulate tissue-specific features and are valuable for studying neuronal involvement.

• • PDX models: Not common for COXPD, but patient-derived xenografts can be used for tumor models with mitochondrial defects.
• • GEMMs: Knockout mice for genes like Surf1, Bcs1l, and Ndufs4 have been generated, recapitulating disease phenotypes.
• • Induced models: Chemical inhibitors (e.g., rotenone for complex I) can induce mitochondrial dysfunction in vivo.

• • CRISPR-Cas9 gene editing enables the creation of isogenic cell lines with precise mutations in COXPD-associated genes. For example:
• • SURF1 knockout cell lines: Generated by introducing frameshift mutations, leading to loss of complex IV assembly.
• • MT-ATP6 knock-in cell lines: Introducing pathogenic point mutations (e.g., m.8993T>G) to model NARP/MILS.
• • BCS1L knockout lines: To study complex III deficiency.

These models are commercially available and sequence-verified, providing researchers with reliable tools for drug screening and functional studies.

Knockout and knock-in lines are used to validate the function of genes implicated in COXPD. For example, SURF1 knockout cells show reduced complex IV activity and increased ROS, confirming its role. Similarly, MT-ATP6 knock-in cells exhibit impaired ATP synthesis, validating the pathogenic variant.

Isogenic pairs (wild-type vs. mutant) are ideal for high-throughput screening of compounds that rescue mitochondrial function. For instance, screening for molecules that increase ATP production in MT-ATP6 mutant cells can identify potential therapeutics. Resistance mechanisms can be studied by exposing cells to mitochondrial toxins and selecting for resistant clones.

CRISPR synthetic lethality screens can identify genes that, when knocked out, are lethal only in COXPD mutant cells. This approach can reveal novel therapeutic targets and biomarkers. For example, targeting genes involved in glycolysis may be synthetically lethal in cells with OXPHOS defects.