PLA2G6-associated neurodegeneration (PLAN): Characterization of patients and drug screening

Neurodegeneration with brain iron accumulation (NBIA) envolve a group of rare neurodegenerative disorders characterized by brain iron accumulation, progressive extrapyramidal dysfunction (dystonia, stiffness, choreoathetosis), and presence of axonal spheroids, usually limited by the central nervous system.   Within the differents subtypes of NBIA, in this study we focussed in PLA2G6-associated neurodegeneration (PLAN), diseases caused by a mutation in the phospholipase A2 group VI (PLA2G6). PLA2G6 encodes the enzyme iPLA2b, a calcium-independent phospholipase A2 which is involved in lipid metabolism. The loss of iPLA2b’s function result in mitochondrial abnormalities and synaptic transmission impairment in neurons among other alterations.   In the current work we studied the pathophysiology of three confirmed cases of PLAN using fibroblasts derived from the patients: PLAN 10 (heterozygous mutation), PLAN 11(heterozygous mutation) and PLAN 8 (homozygous mutation). The aim of this study is to characterize, in patient-derived fibroblasts, the pathological alterations produced by PLA2G6 mutations. Main methods used were Western blot and Prussian Blue Staining. Our results confirm iron accumulation in patients-derived fibroblasts, impaired autophagy and ferritinophagy, especially in the patient that suffers the homozygous mutation (Plan 8). The purpose is to carry out a drug screening able to reverse the pathophysiology observed

Therapeutic approach with commercial supplements for pantothenate kinase-associated neurodegeneration with residual PANK2 expression levels

[Background]: Neurodegeneration with brain iron accumulation (NBIA) is a group of rare neurogenetic disorders frequently associated with iron accumulation in the basal nuclei of the brain characterized by progressive spasticity, dystonia, muscle rigidity, neuropsychiatric symptoms, and retinal degeneration or optic nerve atrophy. Pantothenate kinase-associated neurodegeneration (PKAN) is one of the most widespread NBIA subtypes. It is caused by mutations in the gene of pantothenate kinase 2 (PANK2) that result in dysfunction in PANK2 enzyme activity, with consequent deficiency of coenzyme A (CoA) biosynthesis, as well as low levels of essential metabolic intermediates such as 4′-phosphopantetheine, a necessary cofactor for essential cytosolic and mitochondrial proteins. [Methods]: In this manuscript, we examined the therapeutic effectiveness of pantothenate, panthetine, antioxidants (vitamin E and omega 3) and mitochondrial function boosting supplements (L-carnitine and thiamine) in mutant PANK2 cells with residual expression levels. [Results]: Commercial supplements, pantothenate, pantethine, vitamin E, omega 3, carnitine and thiamine were able to eliminate iron accumulation, increase PANK2, mtACP, and NFS1 expression levels and improve pathological alterations in mutant cells with residual PANK2 expression levels. [Conclusion]: Our results suggest that several commercial compounds are indeed able to significantly correct the mutant phenotype in cellular models of PKAN. These compounds alone or in combinations are of common use in clinical practice and may be useful for the treatment of PKAN patients with residual enzyme expression levels.This work was supported by FIS PI16/00786 and PI19/00377 grants, Instituto de Salud Carlos III, Spain and Fondo Europeo de Desarrollo Regional (FEDER-Unión Europea), Proyectos de Investigación de Excelencia de la Junta de Andalucía CTS-5725 and PY18-850

Pantothenate and L-Carnitine supplementation improves pathological alterations in cellular models of KAT6A syndrome

Mutations in several genes involved in the epigenetic regulation of gene expression have been considered risk alterations to different intellectual disability (ID) syndromes associated with features of autism spectrum disorder (ASD). Among them are the pathogenic variants of the lysine-acetyltransferase 6A (KAT6A) gene, which causes KAT6A syndrome. The KAT6A enzyme participates in a wide range of critical cellular functions, such as chromatin remodeling, gene expression, protein synthesis, cell metabolism, and replication. In this manuscript, we examined the pathophysiological alterations in fibroblasts derived from three patients harboring KAT6A mutations. We addressed survival in a stress medium, histone acetylation, protein expression patterns, and transcriptome analysis, as well as cell bioenergetics. In addition, we evaluated the therapeutic effectiveness of epigenetic modulators and mitochondrial boosting agents, such as pantothenate and L-carnitine, in correcting the mutant phenotype. Pantothenate and L-carnitine treatment increased histone acetylation and partially corrected protein and transcriptomic expression patterns in mutant KAT6A cells. Furthermore, the cell bioenergetics of mutant cells was significantly improved. Our results suggest that pantothenate and L-carnitine can significantly improve the mutant phenotype in cellular models of KAT6A syndrome.This research was funded by FIS PI16/00786 (2016) and FIS PI19/00377 (2019) grants, the Ministerio de Sanidad, Spain, and the Fondo Europeo de Desarrollo Regional (FEDER Unión Europea), Spanish Ministry of Education, Culture and Sport. This activity has been co-financed by the European Regional Development Fund (ERDF) and by the Regional Ministry of Economic Transformation, Industry, Knowledge and Universities of the Junta de Andalucía, within the framework of the ERDF Andalusia operational program 2014–2020 Thematic objective “01-Reinforcement of research, technological development and innovation” through the reference research project CTS-5725 and PY18-850.Peer reviewe

Patient-Derived Cellular Models for Polytarget Precision Medicine in Pantothenate Kinase-Associated Neurodegeneration

The term neurodegeneration with brain iron accumulation (NBIA) brings together a broad set of progressive and disabling neurological genetic disorders in which iron is deposited preferentially in certain areas of the brain. Among NBIA disorders, the most frequent subtype is pantothenate kinase-associated neurodegeneration (PKAN) caused by pathologic variants in the PANK2 gene codifying the enzyme pantothenate kinase 2 (PANK2). To date, there are no effective treatments to stop the progression of these diseases. This review discusses the utility of patient-derived cell models as a valuable tool for the identification of pharmacological or natural compounds for implementing polytarget precision medicine in PKAN. Recently, several studies have described that PKAN patient-derived fibroblasts present the main pathological features associated with the disease including intracellular iron overload. Interestingly, treatment of mutant cell cultures with various supplements such as pantothenate, pantethine, vitamin E, omega 3, α-lipoic acid L-carnitine or thiamine, improved all pathophysiological alterations in PKAN fibroblasts with residual expression of the PANK2 enzyme. The information provided by pharmacological screenings in patient-derived cellular models can help optimize therapeutic strategies in individual PKAN patients.This work was supported by PI19/00377 and PI22/00142 grants, Instituto de Salud Carlos III, Spain and Fondo Europeo de Desarrollo Regional (FEDER-Unión Europea), Proyectos de Investigación de Excelencia de la Junta de Andalucía PY18-850 and UPO-FEDER 2018 (UPO-1380614).Peer reviewe

UPRmt activation improves pathological alterations in cellular models of mitochondrial diseases

Background: Mitochondrial diseases represent one of the most common groups of genetic diseases. With a prevalence greater than 1 in 5000 adults, such diseases still lack effective treatment. Current therapies are purely palliative and, in most cases, insufficient. Novel approaches to compensate and, if possible, revert mitochondrial dysfunction must be developed. Results: In this study, we tackled the issue using as a model fibroblasts from a patient bearing a mutation in the GFM1 gene, which is involved in mitochondrial protein synthesis. Mutant GFM1 fibroblasts could not survive in galactose restrictive medium for more than 3 days, making them the perfect screening platform to test several compounds. Tetracycline enabled mutant GFM1 fibroblasts survival under nutritional stress. Here we demonstrate that tetracycline upregulates the mitochondrial Unfolded Protein Response (UPR), a compensatory pathway regulating mitochondrial proteostasis. We additionally report that activation of UPR improves mutant GFM1 cellular bioenergetics and partially restores mitochondrial protein expression. Conclusions: Overall, we provide compelling evidence to propose the activation of intrinsic cellular compensatory mechanisms as promising therapeutic strategy for mitochondrial diseases.This work was supported by FIS PI16/00786 (2016) and FIS PI19/00377 (2019) grants, the Ministerio de Sanidad, Spain and the Fondo Europeo de Desarrollo Regional (FEDER Unión Europea), Spanish Ministry of Education, Culture and Sport. This activity has been co-financed by the European Regional Development Fund (ERDF) and by the Regional Ministry of Economic Transformation, Industry, Knowledge and Universities of the Junta de Andalucía, within the framework of the ERDF Andalusia operational program 2014–2020 Thematic objective "01—Reinforcement of research, technological development and innovation" through the reference research project CTS-5725 and PY18-850

Neurodegeneration, Mitochondria, and Antibiotics

Neurodegenerative diseases are characterized by the progressive loss of neurons, synapses, dendrites, and myelin in the central and/or peripheral nervous system. Actual therapeutic options for patients are scarce and merely palliative. Although they affect millions of patients worldwide, the molecular mechanisms underlying these conditions remain unclear. Mitochondrial dysfunction is generally found in neurodegenerative diseases and is believed to be involved in the pathomechanisms of these disorders. Therefore, therapies aiming to improve mitochondrial function are promising approaches for neurodegeneration. Although mitochondrial-targeted treatments are limited, new research findings have unraveled the therapeutic potential of several groups of antibiotics. These drugs possess pleiotropic effects beyond their anti-microbial activity, such as anti-inflammatory or mitochondrial enhancer function. In this review, we will discuss the controversial use of antibiotics as potential therapies in neurodegenerative diseases.This project was supported by FIS PI19/00377 (2019) and FIS PI22/00142 (2022) grants, Instituto de Salud Carlos III, Spain; and the Fondo Europeo de Desarrollo Regional (FEDER Unión Europea), Spanish Ministry of Education, Culture, and Sport. This activity was co-financed by the European Regional Development Fund (ERDF) and by the Regional Ministry of Economic Transformation, Industry, Knowledge, and Universities of the Junta de Andalucía, within the framework of the ERDF Andalusia operational program 2014–2020 Thematic objective “01—Reinforcement of research, technological development and innovation” through the reference research project CTS-5725 and PY18-850.Peer reviewe

Alpha-lipoic acid supplementation corrects pathological alterations in cellular models of pantothenate kinase-associated neurodegeneration with residual PANK2 expression levels

[Background]: Neurodegeneration with brain iron accumulation (NBIA) disorders are a group of neurodegenerative diseases that have in common the accumulation of iron in the basal nuclei of the brain which are essential components of the extrapyramidal system. Frequent symptoms are progressive spasticity, dystonia, muscle rigidity, neuropsychiatric symptoms, and retinal degeneration or optic nerve atrophy. One of the most prevalent subtypes of NBIA is Pantothenate kinase-associated neurodegeneration (PKAN). It is caused by pathogenic variants in the gene of pantothenate kinase 2 (PANK2) which encodes the enzyme responsible for the first reaction on the coenzyme A (CoA) biosynthesis pathway. Thus, deficient PANK2 activity induces CoA deficiency as well as low expression levels of 4′-phosphopantetheinyl proteins which are essential for mitochondrial metabolism.[Methods]: This study is aimed at evaluating the role of alpha-lipoic acid (α-LA) in reversing the pathological alterations in fibroblasts and induced neurons derived from PKAN patients. Iron accumulation, lipid peroxidation, transcript and protein expression levels of PANK2, mitochondrial ACP (mtACP), 4′′-phosphopantetheinyl and lipoylated proteins, as well as pyruvate dehydrogenase (PDH) and Complex I activity were examined.[Results]: Treatment with α-LA was able to correct all pathological alterations in responsive mutant fibroblasts with residual PANK2 enzyme expression. However, α-LA had no effect on mutant fibroblasts with truncated/incomplete protein expression. The positive effect of α-LA in particular pathogenic variants was also confirmed in induced neurons derived from mutant fibroblasts.[Conclusions]: Our results suggest that α-LA treatment can increase the expression levels of PANK2 and reverse the mutant phenotype in PANK2 responsive pathogenic variants. The existence of residual enzyme expression in some affected individuals raises the possibility of treatment using high dose of α-LA.This work was supported by FIS PI16/00786 and PI19/00377 grants, Instituto de Salud Carlos III, Spain and Fondo Europeo de Desarrollo Regional (FEDER-Unión Europea), Proyectos de Investigación de Excelencia de la Junta de Andalucía CTS-5725 and PY18-850.Peer reviewe

Activation of the Mitochondrial Unfolded Protein Response: A New Therapeutic Target?

Mitochondrial dysfunction is a key hub that is common to many diseases. Mitochondria’s role in energy production, calcium homeostasis, and ROS balance makes them essential for cell survival and fitness. However, there are no effective treatments for most mitochondrial and related diseases to this day. Therefore, new therapeutic approaches, such as activation of the mitochondrial unfolded protein response (UPRmt), are being examined. UPRmt englobes several compensation processes related to proteostasis and antioxidant mechanisms. UPRmt activation, through an hormetic response, promotes cell homeostasis and improves lifespan and disease conditions in biological models of neurodegenerative diseases, cardiopathies, and mitochondrial diseases. Although UPRmt activation is a promising therapeutic option for many conditions, its overactivation could lead to non-desired side effects, such as increased heteroplasmy of mitochondrial DNA mutations or cancer progression in oncologic patients. In this review, we present the most recent UPRmt activation therapeutic strategies, UPRmt’s role in diseases, and its possible negative consequences in particular pathological conditions

mtUPR Modulation as a Therapeutic Target for Primary and Secondary Mitochondrial Diseases

Mitochondrial dysfunction is a key pathological event in many diseases. Its role in energy production, calcium homeostasis, apoptosis regulation, and reactive oxygen species (ROS) balance render mitochondria essential for cell survival and fitness. However, there are no effective treatments for most primary and secondary mitochondrial diseases to this day. Therefore, new therapeutic approaches, such as the modulation of the mitochondrial unfolded protein response (mtUPR), are being explored. mtUPRs englobe several compensatory processes related to proteostasis and antioxidant system mechanisms. mtUPR activation, through an overcompensation for mild intracellular stress, promotes cell homeostasis and improves lifespan and disease alterations in biological models of mitochondrial dysfunction in age-related diseases, cardiopathies, metabolic disorders, and primary mitochondrial diseases. Although mtUPR activation is a promising therapeutic option for many pathological conditions, its activation could promote tumor progression in cancer patients, and its overactivation could lead to non-desired side effects, such as the increased heteroplasmy of mitochondrial DNA mutations. In this review, we present the most recent data about mtUPR modulation as a therapeutic approach, its role in diseases, and its potential negative consequences in specific pathological situations

Additional file 1 of Alpha-lipoic acid supplementation corrects pathological alterations in cellular models of pantothenate kinase-associated neurodegeneration with residual PANK2 expression levels

Additional file 1. Supplementary figures