Mutations in TDP-43 cause amyotrophic lateral sclerosis (ALS), a fatal paralytic disease characterized by degeneration and premature death of motor neurons. The contribution of mutant TDP-43-mediated ...damage within motor neurons was evaluated using mice expressing a conditional allele of an ALS-causing TDP-43 mutant (Q331K) whose broad expression throughout the central nervous system mimics endogenous TDP-43. TDP-43
Q331K
mice develop age- and mutant-dependent motor deficits from degeneration and death of motor neurons. Cre-recombinase-mediated excision of the TDP-43
Q331K
gene from motor neurons is shown to delay onset of motor symptoms and appearance of TDP-43-mediated aberrant nuclear morphology, and abrogate subsequent death of motor neurons. However, reduction of mutant TDP-43 selectively in motor neurons did not prevent age-dependent degeneration of axons and neuromuscular junction loss, nor did it attenuate astrogliosis or microgliosis. Thus, disease mechanism is non-cell autonomous with mutant TDP-43 expressed in motor neurons determining disease onset but progression defined by mutant acting within other cell types.
The mRNA transcript of the human STMN2 gene, encoding for stathmin-2 protein (also called SCG10), is profoundly impacted by TAR DNA-binding protein 43 (TDP-43) loss of function. The latter is a ...hallmark of several neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS). Using a combination of approaches, including transient antisense oligonucleotide-mediated suppression, sustained shRNA-induced depletion in aging mice, and germline deletion, we show that stathmin-2 has an important role in the establishment and maintenance of neurofilament-dependent axoplasmic organization that is critical for preserving the caliber and conduction velocity of myelinated large-diameter axons. Persistent stathmin-2 loss in adult mice results in pathologies found in ALS, including reduced interneurofilament spacing, axonal caliber collapse that drives tearing within outer myelin layers, diminished conduction velocity, progressive motor and sensory deficits, and muscle denervation. These findings reinforce restoration of stathmin-2 as an attractive therapeutic approach for ALS and other TDP-43-dependent neurodegenerative diseases.
The transcriptional coactivator PGC-1α induces multiple effects on muscle, including increased mitochondrial mass and activity. Amyotrophic lateral sclerosis (ALS) is a progressive, fatal, ...adult-onset neurodegenerative disorder characterized by selective loss of motor neurons and skeletal muscle degeneration. An early event is thought to be denervation-induced muscle atrophy accompanied by alterations in mitochondrial activity and morphology within muscle. We now report that elevation of PGC-1α levels in muscles of mice that develop fatal paralysis from an ALS-causing SOD1 mutant elevates PGC-1α-dependent pathways throughout disease course. Mitochondrial biogenesis and activity are maintained through end-stage disease, accompanied by retention of muscle function, delayed muscle atrophy, and significantly improved muscle endurance even at late disease stages. However, survival was not extended. Therefore, muscle is not a primary target of mutant SOD1-mediated toxicity, but drugs increasing PGC-1α activity in muscle represent an attractive therapy for maintaining muscle function during progression of ALS.
► Muscle is not a primary target for mutant SOD1-mediated toxicity ► Enhanced PGC-1α activity sustains muscle function throughout ALS-like disease ► Improved muscle endurance does not prolong survival in a model of inherited ALS ► Increased PGC-1α activity in muscle is an attractive palliative therapy for ALS
Neurotoxicity from accumulation of misfolded/mutant proteins is thought to drive pathogenesis in neurodegenerative diseases. Since decreasing levels of proteins responsible for such accumulations is ...likely to ameliorate disease, a therapeutic strategy has been developed to downregulate almost any gene in the CNS. Modified antisense oligonucleotides, continuously infused intraventricularly, have been demonstrated to distribute widely throughout the CNS of rodents and primates, including the regions affected in the major neurodegenerative diseases. Using this route of administration, we found that antisense oligonucleotides to superoxide dismutase 1 (SOD1), one of the most abundant brain proteins, reduced both SOD1 protein and mRNA levels throughout the brain and spinal cord. Treatment initiated near onset significantly slowed disease progression in a model of amyotrophic lateral sclerosis (ALS) caused by a mutation in SOD1. This suggests that direct delivery of antisense oligonucleotides could be an effective, dosage-regulatable means of treating neurodegenerative diseases, including ALS, where appropriate target proteins are known.
The discovery of novel biomarkers has emerged as a critical need for therapeutic development in amyotrophic lateral sclerosis (ALS). For some subsets of ALS, such as the genetic superoxide dismutase ...1 (SOD1) form, exciting new treatment strategies, such as antisense oligonucleotide-mediated (ASO-mediated) SOD1 silencing, are being tested in clinical trials, so the identification of pharmacodynamic biomarkers for therapeutic monitoring is essential. We identify increased levels of a 7-amino acid endogenous peptide of SOD1 in cerebrospinal fluid (CSF) of human SOD1 mutation carriers but not in other neurological cases or nondiseased controls. Levels of peptide elevation vary based on the specific SOD1 mutation (ranging from 1.1-fold greater than control in D90A to nearly 30-fold greater in V148G) and correlate with previously published measurements of SOD1 stability. Using a mass spectrometry-based method (liquid chromatography-mass spectrometry), we quantified peptides in both extracellular samples (CSF) and intracellular samples (spinal cord from rat) to demonstrate that the peptide distinguishes mutation-specific differences in intracellular SOD1 degradation. Furthermore, 80% and 63% reductions of the peptide were measured in SOD1G93A and SOD1H46R rat CSF samples, respectively, following treatment with ASO, with an improved correlation to mRNA levels in spinal cords compared with the ELISA measuring intact SOD1 protein. These data demonstrate the potential of this peptide as a pharmacodynamic biomarker.
Abstract
To date, all approved chemotherapeutic agents which target the mitotic cell division interfere with spindle microtubule dynamics, leading to mitotic arrest and apoptosis. While effective, ...these drugs are subject to resistance mechanisms and they are also associated with a variety of side effects, including neurotoxicity. Their use in treating nervous system tumors is therefore not warranted.
One strategy to target mitosis, without damaging microtubules in non-dividing neurons, would be to inhibit key mitotic components, such as the mitotic kinesin Eg5, which is required for establishing a normal bipolar mitotic spindle. We have shown that glioblastoma cells depleted for Eg5 arrest in the next mitosis. After a prolonged arrest, they may slip out and become multinucleated, which will likely prevent further successful divisions or they may go into apoptosis. Further, mitotic arrest and induction of apoptosis in Eg5 depleted glioblastoma cells occur independent of p53, Rb-signalling and the PI3K-pathway suggesting that Eg5 is a potential therapeutic target for glioblastoma patients with different underlying genetic abnormalities
We have also tested the clinical feasibility of using a cell cycle targeting antisense oligonucleotide based therapy delivered directly to the central nervous system (CNS) as a novel treatment for glioblastoma tumors.
This work has demonstrated that intraventricular administration of ASOs can efficiently target cells in the CNS and be delivered to glioma-initiating neural stem cells transplanted into the cortex of naïve mice as well as to glioblastoma tumors in a genetically predisposed mouse model. This strategy is therefore a potential route of administration for treating glioblastoma tumors which originate in the CNS. Direct targeting of mitotic components in the brain should have a limited toxicity to non-cycling neurons and as a benefit, as long as the blood-brain barrier is intact direct CNS delivery should have minimal dose-limiting toxicity outside of the CNS. Ongoing studies will determine the effect of Eg5 inhibition on glioblastoma growth in vivo.
Citation Format: Cecilia C. Krona, Jihane Boubaker, Dinorah Friedmann-Morvinski, Alex Wong, Melissa McAlonis-Downes, Erich Koller, Aneeza S. Kim, Gene Hung, Frank Rigo, Seung Chun, Benjamin Vitre, Frank Bennett, Inder Verma, Don W. Cleveland. Antisense oligonucleotide depletion of the mitotic kinesin Eg5 by direct delivery to the brain could be a useful strategy for treating glioma tumors. abstract. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3302. doi:10.1158/1538-7445.AM2013-3302
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