Gene therapy targeting the blood-brain barrier improves neurological symptoms in a model of genetic MCT8 deficiency

The solute carrier monocarboxylate transporter 8 (MCT8) transports the thyroid hormones thyroxine and tri-iodothyronine (T3) across cell membranes. MCT8 gene deficiency, termed Allan-Herndon-Dudley syndrome, is an important cause of X-linked intellectual and motor disability. As no treatment of the neurological symptoms is available yet, we tested a gene replacement therapy in Mct8- and Oatp1c1-deficient mice as a well-established model of the disease. Here, we report that targeting brain endothelial cells for Mct8 expression by intravenously injecting the vector AAV-BR1-Mct8 increased T3 levels in the brain and ameliorated morphological and functional parameters associated with the disease. Importantly, the therapy resulted in a long-lasting improvement in motor coordination. Thus, the data support the concept that MCT8 mediates the transport of thyroid hormones into the brain and indicate that a readily accessible vascular target can help overcome the consequences of the severe disability associated with MCT8 deficiency.

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Insights Into the Mechanism of MCT8 Oligomerization

Journal of the Endocrine Society ◽

10.1210/jendso/bvaa080 ◽

2020 ◽

Vol 4 (8) ◽

Cited By ~ 1

Author(s):

Stefan Groeneweg ◽

Amanda van den Berge ◽

Elaine C Lima de Souza ◽

Marcel E Meima ◽

Robin P Peeters ◽

...

Keyword(s):

Protein Complexes ◽

Noncovalent Interactions ◽

Monocarboxylate Transporter ◽

Substantial Part ◽

Extracellular Loop ◽

Motor Disability ◽

Lithium Dodecyl Sulfate ◽

Non Covalent Interactions ◽

Mct8 Deficiency ◽

Covalent Interactions

Abstract Mutations in the thyroid hormone transporter monocarboxylate transporter 8 (MCT8) result in MCT8 deficiency, characterized by severe intellectual and motor disability. The MCT8 protein is predicted to have 12 transmembrane domains (TMDs) and is expressed as monomers, homodimers, and homo-oligomers. This study aimed to delineate the mechanism of MCT8 oligomerization. Coimmunoprecipitation studies demonstrated that lithium dodecyl sulfate effectively disrupts MCT8 protein complexes, indicating the involvement of non-covalent interactions. Successive C-terminal truncations of the MCT8 protein altered the oligomerization pattern only if introduced in the N-terminal half of the protein (TMD1-6). The truncation at extracellular loop 1 (E206X) still allowed homodimerization, but completely abrogated homo-oligomerization, whereas both were preserved by the C231X mutant (at TMD2), suggesting that the minimally required oligomerization sites are located proximal of Cys231. However, mutant constructs lacking the intracellular N-terminus or TMD1 and 2 were still capable to form homo-oligomers. Therefore, other domains distal of Cys231 are also likely to be involved in the formation of extensive multidomain interactions. This hypothesis was supported by structural modeling. Despite multiple approaches, MCT8 oligomerization could not be fully abrogated unless a substantial part of the protein was removed, precluding detailed studies into its functional role. Together, our findings suggest that MCT8 oligomerization involves extensive noncovalent interactions between the N-terminal halves of MCT8 proteins. Most mutations identified in patients with MCT8 deficiency have only minor effects on MCT8 oligomerization and, thus, impaired oligomerization does not appear to be an important pathogenic mechanism.

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Consequences of Monocarboxylate Transporter 8 Deficiency for Renal Transport and Metabolism of Thyroid Hormones in Mice

Endocrinology ◽

10.1210/en.2009-1053 ◽

2010 ◽

Vol 151 (2) ◽

pp. 802-809 ◽

Cited By ~ 41

Author(s):

Marija Trajkovic-Arsic ◽

Theo J. Visser ◽

Veerle M. Darras ◽

Edith C. H. Friesema ◽

Bernhard Schlott ◽

...

Keyword(s):

Thyroid Hormone ◽

Thyroid Hormones ◽

Psychomotor Retardation ◽

High Serum ◽

Monocarboxylate Transporter ◽

Iodothyronine Deiodinase ◽

Serum T4 ◽

Mct8 Deficiency ◽

Low Serum

Patients carrying inactivating mutations in the gene encoding the thyroid hormone transporting monocarboxylate transporter (MCT)-8 suffer from a severe form of psychomotor retardation and exhibit abnormal serum thyroid hormone levels. The thyroidal phenotype characterized by high-serum T3 and low-serum T4 levels is also found in mice mutants deficient in MCT8 although the cause of these abnormalities is still unknown. Here we describe the consequences of MCT8 deficiency for renal thyroid hormone transport, metabolism, and function by studying MCT8 null mice and wild-type littermates. Whereas serum and urinary parameters do not indicate a strongly altered renal function, a pronounced induction of iodothyronine deiodinase type 1 expression together with increased renal T3 and T4 content point to a general hyperthyroid state of the kidneys in the absence of MCT8. Surprisingly, accumulation of peripherally injected T4 and T3 into the kidneys was found to be enhanced in the absence of MCT8, indicating that MCT8 deficiency either directly interferes with the renal efflux of thyroid hormones or activates indirectly other renal thyroid hormone transporters that preferentially mediate the renal uptake of thyroid hormones. Our findings indicate that the enhanced uptake and accumulation of T4 in the kidneys of MCT8 null mice together with the increased renal conversion of T4 into T3 by increased renal deiodinase type 1 activities contributes to the generation of the low-serum T4 and the increase in circulating T3 levels, a hallmark of MCT8 deficiency.

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Monocarboxylate Transporter 8 Deficiency: From Pathophysiological Understanding to Therapy Development

Frontiers in Endocrinology ◽

10.3389/fendo.2021.723750 ◽

2021 ◽

Vol 12 ◽

Author(s):

Ferdy S. van Geest ◽

Nilhan Gunhanlar ◽

Stefan Groeneweg ◽

W. Edward Visser

Keyword(s):

Thyroid Hormone ◽

Clinical Stage ◽

Treatment Options ◽

Elevated Serum ◽

Physiological Role ◽

Monocarboxylate Transporter ◽

Peripheral Tissues ◽

Motor Disability ◽

Serum T3 ◽

Mct8 Deficiency

Genetic defects in the thyroid hormone transporter monocarboxylate transporter 8 (MCT8) result in MCT8 deficiency. This disorder is characterized by a combination of severe intellectual and motor disability, caused by decreased cerebral thyroid hormone signalling, and a chronic thyrotoxic state in peripheral tissues, caused by exposure to elevated serum T3 concentrations. In particular, MCT8 plays a crucial role in the transport of thyroid hormone across the blood-brain-barrier. The life expectancy of patients with MCT8 deficiency is strongly impaired. Absence of head control and being underweight at a young age, which are considered proxies of the severity of the neurocognitive and peripheral phenotype, respectively, are associated with higher mortality rate. The thyroid hormone analogue triiodothyroacetic acid is able to effectively and safely ameliorate the peripheral thyrotoxicosis; its effect on the neurocognitive phenotype is currently under investigation. Other possible therapies are at a pre-clinical stage. This review provides an overview of the current understanding of the physiological role of MCT8 and the pathophysiology, key clinical characteristics and developing treatment options for MCT8 deficiency.

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The Thyroid Hormone Analog DITPA Ameliorates Metabolic Parameters of Male Mice With Mct8 Deficiency

Endocrinology ◽

10.1210/en.2015-1234 ◽

2015 ◽

Vol 156 (11) ◽

pp. 3889-3894 ◽

Cited By ~ 17

Author(s):

Alfonso Massimiliano Ferrara ◽

Xiao-Hui Liao ◽

Honggang Ye ◽

Roy E. Weiss ◽

Alexandra M. Dumitrescu ◽

...

Keyword(s):

Thyroid Hormone ◽

Energy Expenditure ◽

Target Genes ◽

High Serum ◽

Metabolic Parameters ◽

Monocarboxylate Transporter ◽

Hormone Analog ◽

Serum T3 ◽

Mct8 Deficiency ◽

The Brain

Mutations in the gene encoding the thyroid hormone (TH) transporter, monocarboxylate transporter 8 (MCT8), cause mental retardation in humans associated with a specific thyroid hormone phenotype manifesting high serum T3 and low T4 and rT3 levels. Moreover, these patients have failure to thrive, and physiological changes compatible with thyrotoxicosis. Recent studies in Mct8-deficient (Mct8KO) mice revealed that the high serum T3 causes increased energy expenditure. The TH analog, diiodothyropropionic acid (DITPA), enters cells independently of Mct8 transport and shows thyromimetic action but with a lower metabolic activity than TH. In this study DITPA was given daily ip to adult Mct8KO mice to determine its effect on thyroid tests in serum and metabolism (total energy expenditure, respiratory exchange rate, and food and water intake). In addition, we measured the expression of TH-responsive genes in the brain, liver, and muscles to assess the thyromimetic effects of DITPA. Administration of 0.3 mg DITPA per 100 g body weight to Mct8KO mice brought serum T3 levels and the metabolic parameters studied to levels observed in untreated Wt animals. Analysis of TH target genes revealed amelioration of the thyrotoxic state in liver, somewhat in the soleus, but there was no amelioration of the brain hypothyroidism. In conclusion, at the dose used, DITPA mainly ameliorated the hypermetabolism of Mct8KO mice. This thyroid hormone analog is suitable for the treatment of the hypermetabolism in patients with MCT8 deficiency, as suggested in limited preliminary human trials.

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Rhes Is Involved in Striatal Function

Molecular and Cellular Biology ◽

10.1128/mcb.24.13.5788-5796.2004 ◽

2004 ◽

Vol 24 (13) ◽

pp. 5788-5796 ◽

Cited By ~ 47

Author(s):

Daniela Spano ◽

Igor Branchi ◽

Annamaria Rosica ◽

Maria Teresa Pirro ◽

Antonio Riccio ◽

...

Keyword(s):

Thyroid Hormones ◽

Motor Coordination ◽

Embryonic Stem ◽

Mental Deficiency ◽

Gene Encoding ◽

Behavioral Abnormalities ◽

And Function ◽

Hormone Control ◽

The Brain

ABSTRACT The development and the function of central nervous system depend on thyroid hormones. In humans, the lack of thyroid hormones causes cretinism, a syndrome of severe mental deficiency. It is assumed that thyroid hormones affect the normal development and function of the brain by activating or suppressing target gene expression because several genes expressed in the brain have been shown to be under thyroid hormone control. Among these, the Rhes gene, encoding a small GTP-binding protein, is predominantly expressed in the striatal region of the brain. To clarify the role of Rhes in vivo, we disrupted the Rhes gene by homologous recombination in embryonic stem cells and generated mice homozygous for the Rhes null mutation (Rhes−/−). Rhes −/− mice were viable but weighed less than wild-type mice. Furthermore, they showed behavioral abnormalities, displaying a gender-dependent increase in anxiety levels and a clear motor coordination deficit but no learning or memory impairment. These results suggest that Rhes disruption affects selected behavioral competencies.

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POLR3-Related Leukodystrophy: Exploring Potential Therapeutic Approaches

Frontiers in Cellular Neuroscience ◽

10.3389/fncel.2020.631802 ◽

2021 ◽

Vol 14 ◽

Author(s):

Stefanie Perrier ◽

Mackenzie A. Michell-Robinson ◽

Geneviève Bernard

Keyword(s):

Hypogonadotropic Hypogonadism ◽

Treatment Options ◽

Genetic Diseases ◽

Early Death ◽

Gene Replacement ◽

Therapeutic Approaches ◽

Disease Modifying Therapy ◽

Gene Replacement Therapy ◽

Progressive Neurodegeneration ◽

The Brain

Leukodystrophies are a class of rare inherited central nervous system (CNS) disorders that affect the white matter of the brain, typically leading to progressive neurodegeneration and early death. Hypomyelinating leukodystrophies are characterized by the abnormal formation of the myelin sheath during development. POLR3-related or 4H (hypomyelination, hypodontia, and hypogonadotropic hypogonadism) leukodystrophy is one of the most common types of hypomyelinating leukodystrophy for which no curative treatment or disease-modifying therapy is available. This review aims to describe potential therapies that could be further studied for effectiveness in pre-clinical studies, for an eventual translation to the clinic to treat the neurological manifestations associated with POLR3-related leukodystrophy. Here, we discuss the therapeutic approaches that have shown promise in other leukodystrophies, as well as other genetic diseases, and consider their use in treating POLR3-related leukodystrophy. More specifically, we explore the approaches of using stem cell transplantation, gene replacement therapy, and gene editing as potential treatment options, and discuss their possible benefits and limitations as future therapeutic directions.

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A Proton-Coupled Transport System for β-Hydroxy-β-Methylbutyrate (HMB) in Blood–Brain Barrier Endothelial Cell Line hCMEC/D3

Nutrients ◽

10.3390/nu13093220 ◽

2021 ◽

Vol 13 (9) ◽

pp. 3220

Author(s):

Kei Higuchi ◽

Sathish Sivaprakasam ◽

Souad R. Sennoune ◽

Jiro Ogura ◽

Yangzom D. Bhutia ◽

...

Keyword(s):

Endothelial Cell ◽

Amino Acid ◽

Gene Family ◽

Cell Line ◽

Blood Brain Barrier ◽

Brain Barrier ◽

Monocarboxylate Transporter ◽

Endothelial Cell Line ◽

Solute Carrier ◽

The Brain

β-Hydroxy-β-methylbutyrate (HMB), a leucine metabolite, is used as a nutritional ingredient to improve skeletal muscle health. Preclinical studies indicate that this supplement also elicits significant benefits in the brain; it promotes neurite outgrowth and prevents age-related reductions in neuronal dendrites and cognitive performance. As orally administered HMB elicits these effects in the brain, we infer that HMB crosses the blood–brain barrier (BBB). However, there have been no reports detailing the transport mechanism for HMB in BBB. Here we show that HMB is taken up in the human BBB endothelial cell line hCMEC/D3 via H+-coupled monocarboxylate transporters that also transport lactate and β-hydroxybutyrate. MCT1 (monocarboxylate transporter 1) and MCT4 (monocarboxylate transporter 4) belonging to the solute carrier gene family SLC16 (solute carrier, gene family 16) are involved, but additional transporters also contribute to the process. HMB uptake in BBB endothelial cells results in intracellular acidification, demonstrating cotransport with H+. Since HMB is known to activate mTOR with potential to elicit transcriptomic changes, we examined the influence of HMB on the expression of selective transporters. We found no change in MCT1 and MCT4 expression. Interestingly, the expression of LAT1 (system L amino acid transporter 1), a high-affinity transporter for branched-chain amino acids relevant to neurological disorders such as autism, is induced. This effect is dependent on mTOR (mechanistic target of rapamycine) activation by HMB with no involvement of histone deacetylases. These studies show that HMB in systemic circulation can cross the BBB via carrier-mediated processes, and that it also has a positive influence on the expression of LAT1, an important amino acid transporter in the BBB.

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Gene replacement therapy in the central nervous system: Viral vector-mediated therapy of global neurodegenerative disease

Behavioral and Brain Sciences ◽

10.1017/s0140525x00037237 ◽

1995 ◽

Vol 18 (1) ◽

pp. 1-9 ◽

Cited By ~ 13

Author(s):

Edward A. Neuwelt ◽

Michael A. Pagel ◽

Alfred Geller ◽

Leslie L. Muldoon

Keyword(s):

Gene Therapy ◽

Central Nervous System ◽

Viral Vector ◽

Genetic Material ◽

Gene Replacement ◽

Vector System ◽

Gene Replacement Therapy ◽

The Central Nervous System ◽

The Brain ◽

Hsv 1

AbstractThis target article describes the current state of global gene replacement in the brain using viral vectors and assesses possible solutions to some of the many problems inherent in gene therapy of the central nervous system (CNS). Gene replacement therapy in the CNS is a potential means of producing a stable expression of normal human proteins in deficient cells and thus curing certain genetically inherited enzyme deficiencies and metabolic diseases as well as cancers. The two major issues to be addressed in CNS gene replacement are the delivery of genetic material to the brain and the expression of recombinant genetic material in target cells within the CNS. Focal inoculation of recombinant virions or other genetic vectors has limitations in global CNS disease. A new approach is the blood-brain barrier (BBB) disruption technique developed in this laboratory, in which hypertonic mannitol transiently shrinks the BBB endothelium, allowing passage of high molecular weight compounds and even viruses. Gene therapy of the CNS will require a viral vector system that allows long-term, nontoxic gene expression in neurons or glial cells. Retroviral vectors have limitations in CNS gene replacement, although they are suitable for expressing recombinant genes in intracerebral grafts, or toxic genes in brain tumors. Using mutant neurotropic viruses with reduced neurotoxicity (such as defective herpes simplex virus type I [HSV-1], the HSV-1 amplicon vector system we have developed, or adenovirus mutants) has potential for direct treatment of neurons. Injecting these vectors into rodent brains can lead to stable expression of foreign genetic material in postmitotic neuronal cells. We discuss our BBB disruption delivery technique, our defective HSV-1 amplicon vector system, and our feline model for the neuronal lysosomal storage disorder Gm2-gangliosidosis (Sandhoff disease), which may prove to be a useful model system for CNS gene therapy.

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